Eubacterium, Fusicatenibacter, Romboutsia, and Roseburia. Metabolomics analysis showed that JTP interfered with 13 biomarkers to play a therapeutic role in type 2 diabetes mellitus. They were L-Valine, Choline, L-Aspartic acid, Serotonin, L-Lysine, L-Histidine, 3-Hydroxybutyric acid, Pyruvic acid, N-Acetylornithine, Arachidonic acid, L-Tryptophan, L-Alanine, and L-Methionine. KEGG metabolic pathway analysis of the above differential metabolites and gut microbiota by using the MetaboAnalyst database and Picrust software. It was found that JTP treated type 2 diabetes mellitus by affecting metabolic pathways such as amino acid metabolism, carbohydrate metabolism, and lipid metabolism. Spearman correlation analysis revealed high correlations for 7 pharmacological indicators, 12 biomarkers, and 11 gut microbiota. In this study, the therapeutic effect and potential mechanism of JTP on type 2 diabetes mellitus were preliminarily demonstrated by gut microbiota and metabolomics, which could provide a theoretical basis for the treatment of T2DM with JTP.The occurrence and development of type 2 diabetes mellitus (T2DM) are closely related to gut microbiota. Jiaotai pill (JTP) is used to treat type 2 diabetes mellitus, with definite efficacy in clinical practice. However, it is not clear whether the therapeutic effect is produced by regulating the changes in gut microbiota and its metabolism. In this study, T2DM rat models were established by a high-fat diet and low-dose streptozotocin (STZ). Based on the pharmacodynamic evaluation, the mechanism of JTP in the treatment of type 2 diabetes mellitus was investigated by fecal metabolism and 16S rRNA gene sequencing. The results showed that JTP decreased blood glucose  and blood lipid  levels and alleviated insulin resistance  in T2DM rats. 16S rRNA gene sequencing results revealed that JTP increased microbiota diversity and reversed the disorder of gut microbiota in T2DM rats, and therefore achieved the therapeutic effect in T2DM. JTP regulated 13 differential flora, which were Actinobacteria, Bacteroidetes, Firmicutes, Proteobacteria, Eubacteriaceae, Prevotellaceae, Ruminococcaceae, Clostridium_IV, Clostridium_XlVa,  Type 2 diabetes mellitus is the most common metabolic disease . With thTraditional Chinese medicine (TCM) has accumulated a wealth of experience in the treatment of diabetes in thousands of years of clinical practice, and TCM can reduce blood glucose through multiple pathways and multiple targets, with obvious advantages in the treatment of diabetes and its complications. Increasing evidence suggests a close association between TCM and gut microbes, which interact with each other. TCM intervenes in intestinal microecology by regulating the number and proportion of gut microbiota, inflammatory factors, signaling pathways, genes, etc., which has the effects of regulating gut microbiota, protecting the intestinal mucosal barrier, restoring intestinal microbial diversity, and enhancing immune function. Intestinal microecology also affects the metabolism and absorption of TCM in the body, which can enhance or reduce the efficacy and change the toxicity of herbal medicines. Jiaotai Pill was first recorded in “Han’s Medical Circular,” and consists of Coptis chinensis, and Cinnamon. It is clinically effective in the treatment of type 2 diabetes mellitus. Modern research has found that Coptis chinensis significantly inhibits the growth of pathogenic bacteria and promotes the growth of beneficial bacteria , 7. MeanThe gut microbiota is both a player and a regulator of metabolic processes . The metIn this study, based on the pharmacodynamic evaluation of the efficacy of JTP in the treatment of type 2 diabetes mellitus, we performed untargeted metabolomics and 16S rRNA gene sequencing studies on stool samples to investigate the changes in endogenous metabolites and intestinal bacteria during the treatment of type 2 diabetes mellitus with JTP. Then, the relationship between host phenotype, intestinal microbiota, and metabolites was analyzed by calculating Spearman correlation coefficients. To investigate the mechanism of JTP for the treatment of type 2 diabetes mellitus from the perspective of multi-level integration of biomarkers and gut microbiota.Coptis chinensis Franch of Buttercup family and the dried bark of Cinnamomi cortex Presl of Camphoraceous family by the Teaching and Research Department of Chinese Medicine Identification of Heilongjiang University of Chinese Medicine, respectively.Coptis chinensis and Cinnamon were purchased from the Harbin Branch of Beijing Tong Ren Tang Pharmacy and identified as the dried rhizome of According to the ratio of Coptis chinensis: Cinnamon (10:1), weigh 500 g of Coptis chinensis and 50 g of Cinnamon, decoct and extract twice, add 10 times the amount of water each time, decoct for 40 min. The two decoctions were combined, concentrated and freeze-dried. A total of 88 g of lyophilized powder was obtained, the powder yield was 16%. Metformin hydrochloride was purchased from Shanghai Yuanye Biotechnology Co., Ltd. (batch number: 201812161).ad libitum. The level of fasting blood glucose (FBG) and body mass were measured. Rats with normal blood glucose values were screened and randomly divided into four groups, which were the control group (Con), model group (Mod), metformin group (MET), and Jiaotai Pill group (JTP). The model and treatment groups were fed a high-fat diet for 4 weeks with two consecutive injections of 25 mg/kg of STZ . The control group was fed a conventional maintenance diet for 4 weeks and injected with an equal volume of citrate buffer intraperitoneally. After 1 week, blood glucose was stabilized, Jiaotai Pill (6.8 g of raw drug/kg) was gavaged in the JTP group, metformin (0.2 g/kg) was gavaged once daily for 10 weeks in the metformin group, and an equal volume of distilled water was gavaged in the remaining groups. All experiments were performed following the Declaration of Helsinki and were approved by the Animal Health and Ethics Committee of Heilongjiang University of Chinese Medicine (2021012709).In this study, 32 men SD rats, weighing 150 ± 20 g, 7–8 weeks old, provided by Liaoning Changsheng Biotechnology Co., Ltd. were used, license number: SCXK (Liao) 2020–0001. The rats were housed in an SPF class animal room with a room temperature of 25 ± 1°C, relative humidity of 50 ± 5%, and a light cycle of 12 h light/night. Rats were acclimatized for a week with a standard rodent diet, and water was available ad libitum, blood was taken by the tail break method, and the level of fasting blood glucose (FBG) was measured.After the rats fasted for 12 h but water Blood was taken from the abdominal aorta of the rats and centrifuged at 3,000 rpm for 10 min at 4°C to collect serum. Total cholesterol (TC), Triglyceride (TG), and Low-density lipoprotein (LDL) in rat serum were measured by an automated biochemical instrument. The enzyme-linked immunoassay was used to detect glycosylated hemoglobin (HbA1c), fasting serum insulin level (FINS), and interleukin 10 (IL-10) in rats.After blood collection, the rat pancreas was carefully separated and the pancreatic tissues were immersed in 10% neutral-buffered formalin to prepare pathological sections.The feces of rats were collected by stimulated defecation and placed into sterilized 2 ml EP tubes. The collected feces samples were uniformly sent to Wuhan UW Medical Laboratory Co., Ltd. for testing, and the Illumina Miseq platform  was used for standard bioinformatic analysis of the bacterial 16S rRNA V3-V4 region. The main procedure is as follows: take 30 ng of qualified genomic DNA samples and corresponding fusion primers to configure the PCR reaction system, set the PCR reaction parameters for PCR amplification, use Agencourt AMPure XP magnetic beads to purify the PCR amplification products, dissolve them in Elution Buffer, label them, and complete the library construction. The libraries were tested for fragment range and concentration using an Agilent 2100 Bioanalyzer. The libraries that passed the assay were selected for sequencing on the HiSeq platform (Illumina) according to the insert size.The collected fecal samples were weighed about 100 mg, placed in 2 ml centrifuge tubes, and thawed on ice. Added 500 μL of ultrapure water, extracted by ultrasonication at 4°C for 5 min, vortex shaking for 30 s, centrifuged at 10,000 rpm for 15 min at 4°C. A total of 300 μL of supernatant was placed in 1.5 ml centrifuge tubes as the first step of extraction. Discard the remaining supernatant in the fecal sediment, add 500 μL of methanol, ultrasonic extract at 4°C for 5 min, vortex shake for 30 s, and centrifuge at 10,000 rpm for 15 min at 4°C. A total of 300 μL of supernatant was taken as the second extraction solution, and the two extracts were combined. After vortex shaking for 30 s and filtration using a 0.22 μm filter membrane, the samples were transferred to the injection vial and all samples were analyzed in positive and negative ion modes. A total of 10 mg of each sample to be tested was mixed and then processed as QC samples according to the pre-treatment method described above.Chromatographic separation was performed on a Thermo Scientific™ Ultra Performance Liquid Chromatograph , We used an Acquity HSS T3  at 40°C, The mobile phase was optimized, and 0.1% formic acid in water and 0.1% formic acid in acetonitrile was selected as mobile phases A and B, respectively. The injection volume was 1 μL in both positive and negative ion modes. The mobile phases were used at a flow rate of 0.4 ml/min with a gradient of 0–3 min at 1.0–10.0% B, 3–5 min at 10.0–20.0% B, 5–8.5 min at 20.0–40.0% B, 8.5–13.5 min at 40.0–99.0% B, 13.5–14 min at 99.0% B, 14–15 min at 99.0–1.0% B.1 and 17,500 FWHM on MS2, the AGC target was set to 1E6 on MS1 and 2E5 on MS2, maximum allowed ion injection time was set to 100 ms on MS1 and 50 ms on MS2, Top three ddms2 for identification, isolation window was set to 1.5 m/z, secondary mass spectrometry collision energy was set to 20, 40, and 60; vertex excitation was 4–8 s, dynamic exclusion was 8 s, acquisition data type was profile mode. The multivariate statistical analysis was used to qualify the potential biomarkers in both positive and negative ion modes.Mass spectrometry (MS) data were acquired by using Q-Exactive focus mass spectrometer  in both positive and negative ion modes. The optimized conditions were as follows: spray voltage was set to 3,500/3,200 V (±), capillary ion transport temperature was set to 350°C, sheath gas flow rate was 45 arb, the auxiliary gas flow rate was 15 arb, capillary (ion transport) temperature was set to 320°C, S-lens voltage was set to 75. The full scan range was set to 60–900 m/z, the resolution was set to 70,000 FWHM on MSThe spliced Tags were clustered into Operational taxonomic unit (OTUs) using the software USEARCH (v7.0.1090_i86linux32). Sequence Variants (ASVs), ASVs are sequences that are 100% similar. In turn, the feature table  is obtained. The RDP classifier Bayesian algorithm was used to analyze the OTU representative sequences taxonomically, and the cluster composition of each sample was counted. The function of the microbiota was predicted using Picrust software.1Raw data were analyzed using Compound Discoverer 3.2  for matching and identification of differential biomarkers. Between-group difference analysis was performed using SIMCA-P . Metabolic Pathway Analysis was performed by using MetaboAnalyst 5.0.Spearman correlation was used to analyze the association of differential biomarkers with differential gut microbiota.t-test or Mann-Whitney U-test was performed to test the differences between the two groups. Statistical analyses were performed using SPSS Statistics , and p < 0.05 were considered significant.Data in a normal distribution were expressed as (mean ± SD). Student’s p < 0.01), and the FBG was significantly higher than 11.1 mmol/L after the second injection of STZ. After 7 days, the FBG of T2DM rats tended to be stable and reached the evaluation criteria of the type 2 diabetes mellitus model  of rats showed an increasing trend in the high-fat diet stage. Compared with the control group, the FBG of the T2DM model group was significantly increased after the first injection of STZ (us model .p < 0.01). FBG in the metformin group (MET) and Jiaotai pill (JTP) groups was significantly lower than that in the model group (p < 0.01)  .p < 0.01), TC levels were significantly increased (p < 0.05), and IL-10 levels were highly significantly decreased (p < 0.01) in the T2DM model rats.Compared with the control group, the serum levels of HbA1c, FINS, TG, and LDL were highly significantly increased (p < 0.01), the level of TC had a decreasing trend, the level of HbA1c was significantly lower (p < 0.05), and the level of IL-10 was highly significant higher (p < 0.01), the levels of HbA1c, FINS, TC, TG, and LDL in the MET group were highly significantly lower (p < 0.01), and IL-10 levels were highly significantly increased (p < 0.01)  .In the control group, the islets had clear boundaries, regular morphology, neatly arranged cells, and uniform size, while in the type 2 diabetes mellitus (T2DM) model group, the islets had fewer cells, disorganized cell bodies, irregular morphology, most nuclei of unequal size, and vacuolar degeneration. Compared with the model group, the islets in metformin group (MET) and Jiaotai pill (JTP) groups were observed to have a regular shape, more β-cells, neat edges, and clear boundaries .In this experiment, based on the 97% similarity OTU obtained, the trends of the species accumulation curve  and specAt the phylum level , the relWe performed PICRUST analysis to predict the function of gut microbiota based on the KEGG database. The relative abundance of 30 pathways was predicted. The top ten pathways were Carbohydrate metabolism, Amino acid metabolism, Metabolism of cofactors and vitamins, Metabolism of terpenoids and polyketides, Replication and repair, Metabolism of other amino acids, Lipid metabolism, Energy metabolism, Cell motility, and Glycan biosynthesis and metabolism. And among them, the expression of Cell motility in the model group was lower than that of the control group. A callback trend was observed after the intervention with JTP. The expression of Carbohydrate metabolism, Amino acid metabolism, Metabolism of cofactors and vitamins, Metabolism of terpenoids and polyketides, Replication and repair, Metabolism of other amino acids, Lipid metabolism, Energy metabolism, Cell motility, and Glycan biosynthesis and metabolism were enriched in the model group. After JTP intervention reversed the abundance of Metabolism of cofactors and vitamins, Replication and repair, Metabolism of other amino acids, Energy metabolism, Glycan biosynthesis and metabolism .Total Ion Chromatography (TIC) of quality control (QC) samples were obtained in both positive and negative ion modes, consistently showing good peak shape and relatively uniform distribution, thus verifying that the UPLC-Q-Exactive focus was stable throughout the detection process .The data of the fecal metabolic profile of rats at key time points  in the T2DM model replication cycle were analyzed, and a certain grouping of model and control rats appeared from day 14 of modeling, but there was an intersection and a significant difference between day 35 of modeling and day 0. This indicates that endogenous substances were changed during the transformation of healthy rats into T2DM rats . Analyzi+: R2Y-0.881, Q2-0.779; ESI–: R2Y-1.000, Q2-0.981  analysis [ESIQ2-0.981 ] was useQ2-0.981 . The datQ2-0.981 , VariablQ2-0.981 , and volQ2-0.981 , and finentified , 2, 11 ientified , includi+: R2Y-0.995, Q2-0.973; ESI–: R2Y-0.952, Q2-0.900  analysis [ESIQ2-0.900 ] of the Q2-0.900 . The resQ2-0.900 . JTP sigQ2-0.900 .p < 0.05 and | r| > 0.5 were selected as correlating.Correlations were calculated using spearman for thirteen Jiaotai pill (JTP)-regulated potential biomarkers and the seven pharmacological indicators . Positiv3-Hydroxybutyric acid was positively correlated with FBG, HbA1c, FINS, TG, LDL and negatively correlated with IL-10. Arachidonic acid was positively correlated with IL-10 and negatively correlated with FBG, FINS, HbA1c, and TG. Choline was positively correlated with LDL, FBG, and FINS. L-Aspartic acid was positively correlated with LDL. L-Histidine was positively correlated with IL-10 and negatively correlated with FINS, FBG, and LDL. L-Methionine was positively correlated with LDL, FINS, FBG, HbA1c, and TG, and negatively correlated with IL-10. L-Lysine was negatively correlated with FBG, FINS, and HbA1c. L- Tryptophan was positively correlated with IL-10, and negatively correlated with FBG, HbA1c, FINS, LDL, and TG. L-Valine was positively correlated with FBG, FINS, HbA1c, and LDL, and negatively correlated with IL-10. N-Acetylornithine was positively correlated with FBG, FINS, HbA1c, LDL, TG, and TC, and negatively correlated with IL-10. Pyruvic acid was positively correlated with IL-10, and negatively correlated with FINS, FBG, HbA1c, and LDL. Serotonin was positively correlated with FBG, FINS, and TG, and negatively correlated with IL-10.p < 0.05 and | r| > 0.5 were selected as correlating.Thirteen JTP-regulated potential biomarkers were correlated with the 13 differential gut microbiota using spearman calculations . PositivAt the phylum level, the Actinobacteria were positively correlated with N-Acetylornithine, L-Methionine, L-Valine, 3-Hydroxybutyric acid, Choline, L-Aspartic acid, Serotonin, and negatively correlated with Arachidonic acid, L-Tryptophan, L-Lysine, Pyruvic acid, L-Histidine. Bacteroidetes were negatively correlated with L-Histidine, Pyruvic acid. Firmicutes were positively correlated with L-Histidine, Pyruvic acid, and Arachidonic acid and negatively correlated with L-Methionine. Proteobacteria were positively correlated with Serotonin, L-Valine, N-Acetylornithine, 3-Hydroxybutyric acid, L-Methionine, and Choline, and negatively correlated with L-Tryptophan, L-Histidine, Arachidonic acid, L-Lysine, Pyruvic acid was negatively correlated.At the genus level, the Clostridium_IV was positively correlated with Arachidonic acid, L-Tryptophan, Pyruvic acid, and L-Histidine, and negatively correlated with L-Valine, N-Acetylornithine, 3-Hydroxybutyric acid, Serotonin, and L-Methionine were negatively correlated. Clostridium_XlVa was positively correlated with Pyruvic acid, L-Histidine, Arachidonic acid, and L-Tryptophan and negatively correlated with 3-Hydroxybutyric acid. Eubacterium was positively correlated with L-Lysine. Fusicatenibacter was positively correlated with L-Methionine, Serotonin, 3-Hydroxybutyric acid, and N-Acetylornithine, and negatively correlated with L-Histidine, L-Lysine, Pyruvic acid, L-Alanine, L-Tryptophan, and Arachidonic acid. Romboutsia was positively correlated with L-Valine and N-Acetylornithine and negatively correlated with L-Tryptophan, Arachidonic acid, Pyruvic acid, and L-Lysine. Roseburia was positively correlated with L-Histidine, Arachidonic acid, Pyruvic acid, and L-Tryptophan, and negatively correlated with 3-Hydroxybutyric acid, Serotonin, L-Methionine, N-Acetylornithine, L-Valine.At the family level, the Eubacteriaceae were positively correlated with L-Lysine. Prevotellaceae was positively correlated with L-Methionine, and negatively correlated with L-Histidine, L-Lysine, Pyruvic acid, and Arachidonic acid. Ruminococcaceae was positively correlated with Arachidonic acid, L-Tryptophan, and negatively correlated with N-Acetylornithine, 3-Hydroxybutyric acid, Serotonin, L-Methionine, L-Valine.The biomarkers and gut microbiota with correlation were correlated with pharmacological indicators, and the Sankey diagram  shows thType 2 diabetes mellitus model rats established by high-fat diet and low dose STZ had dry, gray, and loose hair, depression, gradual loss of body shape, irritability, high urine output, and sour and pungent taste. The study showed that the rats fed with high-fat diets exhibited insulin resistance, increased pancreatic islet β cells, and impaired islet β-cell function . It was Based on 16S rRNA sequencing analysis, we observed a decrease in alpha diversity of gut microbiota in T2DM group rats through sobs, Chao, Shannon, and ace indexes. Microbial species diversity is the basis of intestinal nutrient absorption, metabolism, and immune barrier regulation , 18. In In our study, the dominant microorganisms in the rat gut mainly included Firmicutes, Bacteroidetes, Proteobacteria, and Actinobacteria. The stability of these flora plays an important role in immune regulation, energy metabolism, and substance metabolism, which are essential factors for the maintenance of human health and a mirror reflecting the internal environment of the organism . CompareThe study also confirmed that the abundance of Gram-negative pathogenic bacteria was increased when type 2 diabetes mellitus occurred. Gram-positive bacteria  decreased in abundance. The increase of Gram-negative bacteria will cause an increase in cytosolic toxicogenic LPS . In partPICRUST analysis revealed that gut microbiota is involved in metabolic pathways such as amino acid metabolism, carbohydrate metabolism, and lipid metabolism affecting type 2 diabetes mellitus. Metabolomics was used to further analyze the metabolism of rats. The JTP callback of metabolic pathways associated with type 2 diabetes mellitus is described below.in vitro and is an effective therapeutic supplement for T2DM , which affect insulin resistance and dyslipidemia, and play an important role in type 2 diabetes mellitus . L-Phenyfor T2DM . L-Histifor T2DM . In thisCarbohydrate metabolism was significantly altered in T2DM rats. Pyruvic acid is the end product of glycolysis and the starting point for gluconeogenesis, and can be generated by transamination and participate in energy metabolism to provide energy to living cells and organisms , 37. TheAt the same time, Pyruvic acid can be fermented into succinate, lactate, and acetyl-CoA by gut microbiota and further metabolized into SCFAs. This affects changes in levels of glycolysis/gluconeogenesis and pyruvate metabolism . In thisLipid metabolism is highly associated with the development of type 2 diabetes mellitus (T2DM). Arachidonic acid is a polyunsaturated fatty acid that inhibits lipogenesis and promotes lipolysis and playsThis study mainly focuses on the changes in gut microbiota and metabolite detection in JTP intervention in T2DM rats. The animal indicators are currently limited to the phenotypic part. In the future, experiments are needed to further improve the exploration of pathways and mechanisms.In summary, Jiaotai pill (JTP) decreased blood glucose and lipid levels and reduced insulin resistance in type 2 diabetes mellitus (T2DM) rats. Potential biomarkers and key functional bacteria were identified for JTP treatment in T2DM rats. These results suggest that JTP can increase microbiota diversity and restore gut microbiota balance. It improves metabolic pathways such as amino acid metabolism, carbohydrate metabolism, and lipid metabolism associated with type 2 diabetes mellitus, and plays a therapeutic role in T2DM rats.https://www.ncbi.nlm.nih.gov/bioproject/922696.The datasets presented in this study can be found in online repositories. The name of the repository and accession number can be found below: This animal study was reviewed and approved. All experiments were performed following the Declaration of Helsinki and were approved by the Animal Health and Ethics Committee of Heilongjiang University of Chinese Medicine (2021012709).XhW and JL designed the study. XW conducted the animal trial, sample collection, and physical analysis. QL and SY performed the bioinformatics analysis of 16S rRNA sequencing. JL and YZ performed the untargeted metabolomics data. JL and CP wrote the manuscript. XhW, JL, and ZF contributed to the discussion of the work and assisted in drafting the manuscript. All authors read and approved the final manuscript.


Evaluation of the Shear bond strength over zirconia and titanium alloy and degree of conversion of extraoral compared to intraoral self-adhesive resin cements.Nine bonding protocols were carried out on zirconia 4Y-TZP and titanium alloy (Ti-6Al-4V). Seven resin cement  were tested in the shear bond strength test and the degree of conversion measurements.The significantly highest value was obtained for Monobond Plus + Multilink Hybrid Abutment, the extraoral resin cement for both titanium alloy (35.1 MPa) and zirconia (32.9 MPa). For each resin, significantly higher DC values were obtained for the dual-cure mode compared with the self-cure mode. Regardless of the cure mode, Nexus Universal reached the highest DC (78.4%).In this study, the extraoral self-curing resin cement showed the higher bond strength values on zirconia and titanium alloy when associated with a universal primer. Some intraoral dual-cure resin cements showed closed performances when used with universal primers. There is no direct correlation between the degree of conversion of the resin cement and the shear bond strength obtained on the prosthetic materials tested. Zirconia and titanium alloys are widely used materials in prosthetic dentistry. In practice, they need to be carefully grit-blasted to achieve good micromechanical retention  . MoreoveZirconia can be bonded on dental tissues, such as cementing zirconia crowns on prepared teeth abutments  or on titanium base abutment (Ti-base) in prosthetic labs  . Some reDepending on their chemical composition, these resin cements can be self-cured (chemopolymerized), light-cured (photopolymerized) or dual-cured (association of chemopolymerization and photopolymerization) . There aThe aim of this study was first to evaluate the shear bond strength of one extraoral and various intraoral resin cements to zirconia and titanium alloy according to nine bonding protocols in self-curing mode (in order to assess their bonding strength in the least favorable polymerization mode) and then to measure their degree of conversion in self-curing mode or with additional light curing. The first null hypothesis was that there are no differences in shear bond strengths between the nine protocols tested resin for each substrate. The second null hypothesis was that there were no differences in the degree of conversion between the various resin cements in self-curing mode or with an additional light-cured mode.The materials used in this study are summarized in Table Nine bonding protocols Table  were car2O3 (50 µm) for 10 s at 2 bars, rinsed with water/air spray, then with a 99% ethanol solution and dried.Blocks of zirconia and titanium alloy were included in self-curing acrylic resin and then abraded with water-cooled sandpaper (800 grits) to expose a flat surface (>7 mm2). All these samples were grit-blasted with Aln = 180), a cylindrical Teflon mold was placed to build a 3 mm-high cylinder of resin cement with a flat base of 7 mm² (diameter = 3 mm). The tested bonding protocols were performed following the manufacturer’s instructions (which can lead to various protocols for a specific resin cement). The resin cement was set in self-curing mode (in the dark) under 50 g pressure for 60 min. Then, the mold was removed, and the excess, if present, was gently removed with a scalpel. All the samples were stored in demineralized water at 37 °C for 24 h. Detailed bonding protocols are summarized in Table On each sample (n = 10), SBS values were determined in a universal testing machine . The shear force was applied at the resin cement/prosthetic material interface, with a chisel-shaped blade parallel to the prosthetic material surface. A cross-head speed of 0.5 mm/min was chosen.For each group  at 30× magnification, and the failure modes were classified according to the following four types: cohesive failure within the prosthetic material, adhesive failure at the interface between the resin cement and the prosthetic material, mixed failure , cohesive failure within the resin cement.n = 7), 6 cylindrical specimens (diameter 6 mm x height 3 mm) were made with Teflon molds: 3 were performed in with light-activation for 60 s at 1200 mW per square cm  immediately after mixing, and 3 were performed only in self-curing mode (mixed in the dark). All samples were then stored in the dark in distilled water at 37 °C for one week before the degree of conversion (DC) measurement.For each resin cement tested  with a NicoletTM iS10 spectrometer  in attenuated total reflectance (ATR) mode following proposed protocol from previous publications . These s−1) and that of the aromatic C=C bond (1608 cm−1) were measured, first in the monomer state and then after the polymerization process. DC was determined by evaluating the change in the height ratio of the aliphatic C=C peak and the aromatic C=C peak during the polymerization process [For each spectrum, the height of the absorption band of the aliphatic C=C bond  were used to investigate the difference in SBS between the different groups. The failure mode was analyzed by Fisher’s exact test for single comparisons between groups and pairwise analysis.p < 0.05. Statistical calculations were performed using R software .For DC results, a 2-way ANOVA followed by Tukey’s post hoc test was performed for the factors “resin” and “mode of polymerization”. In all tests, the significance level was The mean SBS values and standard deviations are reported in Table The significantly highest SBS value was obtained for MP + MHA, the extraoral resin cement for both Ti-6Al-4V (35.1 MPa) and zirconia (32.9 MPa). In contrast, TTC had the lowest SBS values regardless of the prosthetic material (15.9 MPa for Ti-6Al-4V and 12.7 MPa for zirconia).vs. 29.4 MPa for GMP + GC-O; 24.7 MPa for PSA-U vs. 24.3 MPa for CUB + PSA-U), whereas it was significantly increased on titanium alloy for GC-O (25.9 MPa for GC-O vs. 32.0 MPa for GMP + GC).The use of universal primer or universal adhesive did not significantly improve the bond strength on zirconia , whereas TTC had the lowest DC (60.3%).It is difficult to establish an immediate in vitro threshold bond strength, despite that such a measurement could ensure long-term clinical adhesion. However, long-term clinical studies show good bonding performance on zirconia and Ti-6Al-4V when the standard protocols are followed correctly –26.In this study, macro-shear bond strength test was used to assess SBS values of the various adhesive protocols to Zirconia and Ti-6Al-4V. The macro-shear bond strength test is one of the two most frequently used tests, along with the micro-tensile bond strength test, to assess the strength of an adhesive procedure –29. SomeBonding strength values on Ti-6Al-4V and zirconia were measured concomitantly, due to the dual indication of these two materials in certain prosthetic indications such as bonding crowns on abutments in implantology. It is therefore interesting to know the performance of the same protocol on these two substrates. However, in mouth, the two materials have different clinical indications. Zirconia can be bonded in a large number of different fixed prosthetic indications , 31, whiThe extraoral resin cement bonding protocol MP + MHA was chosen for study here because of its widespread use in the literature with good clinical retention results –36. As aThe two highest values found for each prosthetic material were MHA and GC-O using their dedicated universal primers . There is a general trend of specific primers such as MP or GMP to be more effective than universal adhesives used as primers on titanium alloy and zirconia . MP is aHere, all dental universal adhesives tested as an alternative to universal primers also contained 10-MDP or glyceryl dimetacrylates as functional monomers but led to lower bond strength values than when a universal primer was used. All curing procedures were carried out in a dark room with a self-curing mode to avoid activating light-curing polymerization of resin cements. Under these conditions, universal adhesives used as nonspecific primers were not light-cured either. This could also contribute to the lower bond strength values, even though manufacturers report self-cure contact with their associated resin cements.Intraoral self-adhesive resin cements showed different behaviors when used directly on prosthetic materials without their specific primers or associated universal adhesives. GC-O had a completely different behavior between bonding on zirconia or titanium alloy. On zirconia, GC-O with or without its universal silane GMP exhibited almost the same SBS value, whereas significantly low values were observed on Ti6-Al-4V. This could be linked to the absence of MDTP in its composition, whereas this molecule is present on GMP.For the other self-adhesive resin cements tested without their associated universal primer or universal adhesive on zirconia, we note that there were a number of factors that may explain the lower performance compared to GC-O. Kerr, who produces the OBU and the N-U, is the only brand that uses GPDM instead of 10-MDP in self-adhesive resin cements or dental adhesives. Prior research has shown that 10-MDP has a higher affinity for zirconia than GPDM . MoreoveAll the material in dual cure mode yielded a higher DC compared to the self-cured mode. This outcome has been reported in many studies , 53. LigGPDM is a monomer found only in N-U, which has two methacrylate groups and one phosphate group. These two methacrylate groups can promote better cross-linking in polymerized networks despite other functional monomers, such as 10-MDP . This coDual curing is therefore concluded to optimize the degree of conversion in any clinical or laboratory situation, which can have a major impact on the long-term adhesion stability, particularly in terms of water absorption and resin solubility in the bonding interface , which cSurprisingly, MHA, the only extraoral and self-cured resin cement, had higher DC with light curing than self-cure mode. This resin cement is recommended for Ti-base abutment sealing in laboratory and no light-curing is advised. This recommendation is explained by the astonishing presence of camphorquinone, a light curing initiatior in its composition. Therefore, although not recommended by the manufacturer, additional light-curing of this material in the laboratory could result in beneficial properties, especially as the high temperatures caused by possible heating do not appear to alter its mechanical and adhesive properties. This procedure would also be a useful technique for improving its biological properties in the oral environment .It would be interesting to carry out adhesion tests with ageing, in particular to evaluate the evolution over time of the simplified intra-oral adhesives studied in this study, which contain a certain hydrophilicity due to their self-adhesive chemistry.Because of their in vitro nature, these results cannot be directly transposed to the clinic. Long-term studies are needed to draw clinical conclusions.This study being in vitro, it includes a certain number of limitations due to this fact, but also to its construction scheme:Finally, as a follow-up to this study, the behavior of the simplified intra-oral adhesives from this study on dental tissues will be investigated in addition to the current investigation on prosthetic tissues.In this study, the extraoral self-curing resin cement showed the higher bond strength values on zirconia and Ti-6Al-4V alloy when associated with a universal primer. Some intraoral dual-cure resin cements showed closed performances when used with universal primers.The extreme simplification of bonding procedures based on the use of self-adhesive resin cements containing functional monomers alone or monomers associated with a universal adhesive led to variable results depending on brands and formulations. These results may nevertheless be sufficient for these applications given that they yielded bond strength values close to those of conventional procedures.There is no direct correlation between the degree of conversion of the resin cement and the shear bond strength obtained on the prosthetic materials tested. Additional light curing is advised in every case of bonding to enhance the degree of conversion.However, further clinical studies validating these results are required before generalizing these conclusions.


Increasing evidence suggests that reducing pulse pressure amplification (PPA) plays an important role in pathogenesis and progression of cardiovascular disease. This is a cross-sectional, observational, and analytical study in which we evaluated the associated factors with a greater chance of reducing PPA in 136 healthy children and adolescents aged 8 to 19 years old stratified by gender and age group.Arterial stiffness and vascular and hemodynamic parameters were non-invasively measured using Mobil-O-Graph® , a cuff-based oscillometric device. PPA was expressed as the peripheral-to-central pulse pressure ratio (PPp / PPc). Participants with PPA < 1.49 were considered as part of the arterial stiffness group.In a univariate model, the increase in total vascular resistance, the reflection coefficient and the augmentation pressure were more likely to have arterial stiffness in all groups. The factors most likely to have arterial stiffness (as assessed by the reduction of the PPA) in the multivariate model were increasing age, the reflection coefficient and cardiac index in the total sample, male group and child and adolescent groups. In addition to age in the female group, cardiac output, stroke volume, and AIx@75 were the factors most likely to present arterial stiffness.The results show for the first time in children and adolescents that the factors most likely to reduce PPA are related to the reflection wave, which determines aortic pressures and, therefore, left ventricular afterload. Arterial stiffness is a natural aging process that can be accelerated by the cumulative effects of risk factors such as high blood pressure, hyperglycemia, adiposity, hypercholesterolemia , sedentaArterial stiffness can be assessed by invasive and non-invasive methods. However, there is a lack of validation studies in pediatrics comparing invasive measures such as cardiac catheterization and non-invasive measures due to the ethical concerns involved with invasive studies in children for research purposes only . The mosStudies in adults show that PPA has a significant inverse relationship between age and its reduction has a negative impact on mortality, cardiovascular events, and damage to noble organs such as the heart, kidney and brain , 10. TheAlthough the atherosclerotic process often starts early in childhood, the clinical manifestations of cardiovascular disease (CVD) appear in adulthood . Thus, tThis is a cross-sectional study to assess PPA in children from 8 years old and adolescents up to 19 years old, considered healthy, with normal body mass index (BMI). Children and adolescents with acute or chronic diseases were excluded from the study. A total of 177 students were recruited from public schools in 9 metropolitan regions of Belo Horizonte. However, 41 participants were excluded for the following reasons: 8 did not show up on the day scheduled for the exam, 9 did not sign the Informed Parental Consent Form or the Assent Form, 17 had a history of chronic respiratory disease (recurrent wheezing or diagnosis of asthma), 3 used medications for epilepsy, 2 were using Ritalin and 2 had a high BMI. Thus, the final sample consisted of 65 female and 71 male participants.The sample size was calculated according to Duarte et al. . The samAfter parents or guardians provided written informed consent, the participants answered the questionnaires to assess their socioeconomic level, quality of life and habitual physical activity level. The questionnaires were self-administered and when participants had difficulty understanding an item, they were helped by the trained researchers. Anthropometric data were subsequently collected, followed by cardiovascular measurements.2). Children and adolescents with percentiles between 5 and 85% were included in the study. Waist (WC) and hip (HC) circumferences were measured and the ratios between WC/Height and WC/HC were obtained. Cut-off points based on WHO guidelines (2007) were used to analyze weight, height, and BMI [BMI was obtained by the ratio between weight (kg) and height squared  was used to assess habitual physical activity level, which investigates the practice of physical activity in children and adolescents in the seven days prior to filling it out. It consists of nine questions related to physical activity performed at school and during leisure time during the week and on weekends. It also includes questions related to the time spent in front of the television and the computer. Each question has scores from 1 to 5. Those who spent 150 min performing moderate physical activities during the week were considered active .Health-related quality of life was evaluated using the Pediatric Quality of Life Inventory Version 4.0 (Peds-QL 4.0) . It is aThe assessment of respiratory health status was assessed using the International Study of Asthma and Allergies in Childhood (ISAAC) questionnaire, with the aim of excluding volunteers with a significant respiratory history or medical diagnosis of asthma, as this is a very prevalent disease in this age group . This quThe socioeconomic classification was performed as recommended by the Brazilian Association of Research Companies . The socCardiovascular parameters were performed according to previous study by our group . The MobThe cuffs were selected according to the circumference of the participant’s left arm, where the measurements were taken. The measurement was performed with the participant sitting, in a calm and quiet place, after a minimum rest of 5 min. The room temperature was maintained between 21 degrees according to the manufacturer's recommendation. The researchers responsible for data collection were trained to handle the Mobil-O-Graphy and apply the questionnaires.All participants were instructed to maintain a postprandial state of at least three hours and 24 h of abstaining from physical exercise, consumption of caffeine or other stimulant food before this stage.The use of Mobil-O-Graphy to assess cardiovascular parameters in children and adolescents has previously been used by our research group , 23, 24 The variable PPA was categorized into values less than and greater than or equal to the median (1.49). The group of children and adolescents with PPA < 1.49 was called the arterial stiffness group, and with PPA ≥ 1.49 as the control group.Categorical variables were presented as counts and percentages, and numerical variables as mean ± standard deviation (median). Numerical variables were submitted to the Shapiro–Wilk normality test.p < 0.20 in the univariate analysis were included in a saturated model, to which the final model was reached using the backward strategy, with variables with p < 0.05 and the variable age independent of significance to control effects. The results were presented as OR, 95% confidence interval of the OR and respective p-values. The quality of the fit was analyzed using the Hosmer–Lemeshow (HL) test. The analysis was developed in the free R version 4.0.2 program.Univariate logistic regression models were built to assess the association between each of the analyzed variables and arterial stiffness indices. The results were presented as odds ratio (OR) and the respective p-values for the whole sample, and stratified by gender and age group. Variables with The study involved 136 individuals, 81 (59.6%) adolescents and 71 (52.2%) males. Moreover, 62 participants (45.6%) were in social class A or B, and 74 (54.4%) in social class C. Regarding the habitual physical activity level, 95 participants (69.8%) were considered active and 41 participants (30.2%) were considered sedentary. Table The minimum and maximum PPA values were 0.95 and 2.05, respectively. The coefficient of variation was 12.45%. Anthropometric data, socioeconomic class, habitual physical activity level, and quality of life were not associated with a greater chance of having arterial stiffness, as assessed by PPA.Table p < 0.01), AIx@75 (p = 0.01), reflection coefficient (p < 0.01) and augmentation pressure in the univariate analysis for the entire sample represented a greater chance of having arterial stiffness (p < 0.01). On the other hand, an increase in cardiac output (p < 0.01) and cardiac index (p = 0.03) presented less chance of increasing arterial stiffness (Table p = 0.016) and cardiac index (p < 0.001) in the multivariate model were associated with a lower chance of having arterial stiffness, and an increase in reflection coefficient was associated with a higher chance of increasing PPA (p < 0.001) (Table An increase in TVR (x p = 0.0 presente1) Table .Table 3Up = 0.03), reflection coefficient (p < 0.01) and augmentation pressure (p = 0.01) in the univariate analysis of the group of male individuals represented a greater chance of having arterial stiffness (Table p = 0.026) and cardiac index (p < 0.001) in the multivariate model were associated with a lower chance of having arterial stiffness, and an increase in reflection coefficient was associated with a higher chance (p < 0.001)  Table .p < 0.01), and a higher chance of an increase in the TVR (p < 0.01), in the reflection coefficient (p < 0.01), augmentation pressure (p < 0.01) and AIx@75 (p < 0.01) in the univariate analysis (Table p = 0.003) in the multivariate model, while the increase in systolic volume and AIx@75 represented greater chances   Table .p < 0.01) and augmentation pressure (p < 0.01) in the group of children in the univariate analysis were associated with a greater chance of having arterial stiffness (Table p = 0.036) in the multivariate model, and an increase in cardiac output was associated with a lower chance (p < 0.001) (Table An increase in reflection coefficient (ss Table . An incr1) Table .p = 0.01) in the univariate analysis was less likely to present arterial stiffness, while the increase in TVR (p < 0.01), reflection coefficient (p < 0.01), AP (p < 0.01) and AIx@75 (p < 0.01) were more likely to have arterial stiffness (Table p = 0.003) in the age-controlled multivariate model, while the increase in reflection coefficient represented a greater chance (p < 0.001) (Table The increase in cardiac output (ss Table . In addi1) Table .In the present study, we evaluated the factors associated with reduced PPA in children and adolescents considered healthy, stratified by gender and age group. Participants with PPA < 1.49 were considered from the arterial stiffness group. An increase in the TVR, the reflection coefficient and the augmentation pressure in the univariate model were more likely to have arterial stiffness in all groups. The factors most likely to have arterial stiffness as assessed by the PPA in the multivariate model were the age increase and the reflection coefficient in the total sample, male group and in the child and adolescent groups. Moreover, age and AIx@75 were the factors most likely to present arterial stiffness in the female group. It is important to point out that the reflection coefficient measures the magnitude of the reflection wave, while the AIx@75 measures the percentage increase in the reflection wave in relation to the PPc.PPA can be calculated in several ways: 1- difference between PPp and PPc (PPA = PPp—PPc), 2- relationship between PPp and PPc (PPA = PPp / PPc), 3- relationship between (PPp—PPc) / PPc and 4- absolute difference between SBPp and SBPc . PPA in In comparing the coefficient of variation of the different PPA indices, we observed that the lowest coefficient of variation was the PPp / PPc ratio (12.54%). Furthermore, the PPp—PPc, (PPp—PPc) / PPc and SBPp – SBPc indexes were 38.87%, 36.89% and 46.67%, respectively.PPA has been extensively studied in adults and the results show that PPA has a significant inverse relationship between age, and its reduction has a negative impact on mortality, cardiovascular events and on the damage to noble organs such as the heart, kidney and brain . In contThe variable age was included in the multiple regression analysis model in the present study, regardless of significance to control for effects. Age is the main non-modifiable factor associated with reduced PPA because normal vascular aging is the main causal factor for arterial stiffness of the great arteries , 28.Increases in TVR, reflection coefficient, augmentation pressure and AIx@75 were more likely to have arterial stiffness in the univariate analysis of the entire studied sample. TVR is defined by the relationship between pressure and blood flow. The reflection coefficient is a measure of the magnitude of the reflection wave . This inIncreases in TVR, reflection coefficient and augmentation pressure represented a greater chance of having arterial stiffness in the male group in the univariate analysis. Similarly to the total sample, increasing age and cardiac index were associated with a lower chance of having arterial stiffness in the multivariate model, and an increase in the reflection coefficient was associated with a higher chance of having arterial stiffness.Univariate PPA analysis of the female and adolescent groups were similar. The increase in the cardiac output was associated with lower chance to have arterial stiffness, and increases in total vascular resistance (TVR), reflection coefficient and augmentation pressure, were associated with a greater chance of having arterial stiffness in adolescent group. In addition to these factors, the increase in AIx@75 in the female group also contributed to a greater chance of having arterial stiffness, that is, a reduction in PPA. In the multivariate model, an increase in cardiac output and age were associated with a lower chance of having arterial stiffness in the female group, while the increase in systolic volume and AIx@75 represented a greater chance of arterial stiffness. The AIx@75 is a measure, in percentage, of the increase in the augmentation pressure in relation to the PPc, which occurs due to the early return of the reflection wave . An incrIncreases in reflection coefficient and augmentation pressure in the group of children were associated with a greater chance of having arterial stiffness. However, an increase in the cardiac index was associated with a lower chance of having arterial stiffness in the age-controlled multivariate model, while the increase in the reflection coefficient represented a greater chance of arterial stiffness.The results of the present study show that the factors which present a higher risk of reduction in PPA in multivariate analysis logistic regression are related to the reflection wave. PWV, the gold standard for arterial stiffness, was not associated with a reduction in PPA. These results suggest that PPA is related to wave propagation velocity, reflected wave amplitude, reflection point, and the ventricular ejection duration and pattern. In contrast, aortic PWV represents intrinsic arterial stiffness. Furthermore, the influence of age is greater in AIx@75 than in aortic PWV before 50 years. Then, the influence of age after 50 years is greater on PWV . PP signStudies show, in adults, that APP has a significant inverse relationship between age and its reduction has a negative impact on mortality, cardiovascular events and damage to noble organs such as the heart, kidney and brain . Thus, lOur study has the following strengths. First; the sample was representative of the different regions of the Belo Horizonte city, gender, and age groups. The adjusted models considered the assumptions necessary to construct linear regression models  and presented no multicollinearity problem. Second, we excluded volunteers with a significant respiratory history or medical diagnosis of asthma using the International Study of Asthma and Allergies in Childhood (ISAAC) questionnaire. Asthma is a prevalent disease in this age group. We  and otheHowever, our findings present some limitations. First, due to its cross-sectional design, our study cannot be conclusive regarding the causal relationship between variables. Second, although the sample was adequate, it may have been too small for sub-group analysis. Third, we did not collect family history data for cardiovascular disease and dietary habits; and fourth, we did not exclude participants with a history of obstructive sleep-disorder breathing. We recently demonstrated that children with obstructive sleep-disorder breathing have increased arterial stiffness, evidenced by the increase in AIx@75 . Herein,The results of the present study show that the factors that present a higher risk of reduction in PPA in univariate logistic regression are related to the reflection wave, meaning the reflection coefficient, augmentation pressure and AIx@75. PWV the gold standard for arterial stiffness was not associated with a reduction in PPA. PPA is related to wave propagation velocity, reflected wave amplitude, reflection point, and the ventricular ejection duration and pattern, while aortic PWV represents intrinsic arterial stiffness. Taken together, these data suggest that the assessment of these arterial stiffness indices, PPA and PWV, are not interchangeable, and that PPA, like AIx@75, may be a more sensitive marker in young individuals.


When serum cortisol was included, heterogeneity stayed at 0%. A larger number of high-quality studies, especially those specific to serum cortisol, are recommended, to elucidate the relationship between OMT and cortisol. This research suggests OMT reduces cortisol more than sham treatment before versus after OMT treatment, and though the change is small when comparing after one treatment, it may have clinical usage if multiple OMT sessions are performed.The effectiveness of osteopathic manipulative treatment (OMT) in the modification of various hormones has been studied; however, there is still a need for quantitative measurements to determine how large of an influence exists. The goal of this meta-analysis is to investigate the implications OMT has on cortisol levels. A systematic literature search restricted to English was performed from October 2022 to November 2022 using Google Scholar, OSTMED.DR, and PubMed and included articles from 2000 onward. Articles were excluded if they did not include a measurement for the control group in their study. We identified 4120 studies for potential inclusion. Of these, a total of four studies met the inclusion criteria, with a total of 135 participants . Out of the 135 participants, 126 participants  made up the salivary cortisol studies, and the remaining nine participants made up the serum cortisol studies . The National Institutes of Health (NIH) bias assessment tool was utilized to measure the risk of bias. Standard mean differences were calculated for effect size. A mean difference in cortisol of 0.10μg/dl  was found when comparing all pre- versus post-cortisol levels with OMT versus sham control groups. OMT demonstrated a 0.10μg/dl larger decrease in cortisol than sham control treatments. The standard mean difference was found to be -0.46  making this finding a medium effect size without significance. Heterogeneity for the salivary analysis measured by I Osteopathic manipulative treatment (OMT) serves as a modality to potentially assist in the treatment of various illnesses. Through manipulation treatment, osteopathic physicians offer an additional method by which healing can be promoted in patients. While these treatments may have effects on various biomarkers, the exact changes are unknown. Bodily manipulation is well established in-vitro, with fibroblasts indicating changes in what is released, how cells change, and bodily interconnection -4. It foThe common problem with OMT research is that studies often have low power, low sample size, or questionable experimental design . To bettFor something like cortisol, which often is only in concentrations of micrograms per deciliter in the blood or salivary secretions, the minute differences are one such thing that may be overlooked . CortisoWith cortisol established as a necessary component of the stress response, its potential use in treatment modalities, and OMT possibly being able to affect cortisol concentration, an exploration of OMT and cortisol via meta-analysis is necessary. The objective of this study is to combine study results as a systematic review to find the overall change of cortisol in OMT versus sham treatments. Through this finding, the hope is to explore if there is a difference found in OMT that can have potential use in treatment.Materials and methodsStudies were included based on specific inclusion criteria. To be included, a study needed to have pre- and post-treatment data, a defined control group, standard deviation as well as effect size and mean , sample size for OMT and sham groups, cortisol , and a specified and recognized OMT treatment. Additionally, cortisol values were measured in μg/dL units in this nlysis, so all included studies needed data that was convertible to these units. Exclusion criteria were the following: studies that were not written in English, studies published prior to 2000, and studies that used alternative means to measure cortisol other than salivary or serum cortisol .As for database search, Google Scholar, Osteopathic Medical Digital Repository (OSTMED.DR®), and PubMed were used. PubMed and OSTMED.DR had limited search results, and all search results found on them were also found on Google Scholar. The search term used in the search was “osteopathic” AND “cortisol.” After the initial article results, articles were eliminated first based on the language. This was done by using each database's option to only find English-only articles. After this, articles were excluded based on the other inclusion and exclusion criteria.The search was divided among three researchers. Article search results were compiled together onto a shared document and each of the three researchers was given part of the search results to go through. Data gathered by the researchers were compiled into a shared document. If a researcher disputed an article for analysis, the article was given to the other two researchers doing the search to analyze. If the researchers still disputed whether the article should be included, the article was presented to the principal investigator of the project for a final say. Lastly, articles that did not include a control group measurement in their study were excluded from the final group. An example of this is Henderson et al., where placebo group changes in salivary cortisol were reported in terms of percent change . PercentData collected included control and OMT group sample size, standard deviation, mean difference (or pre- and post-mean) for control and OMT groups, and OMT technique comparison. If multiple time points were given, the time points measuring closest to before and after the treatment was used. Risk of bias was measured using the National Institutes of Health (NIH) bias assessment tool. For effect size measurement, the standard mean difference was calculated .2, in which a value above 50% was considered significant heterogeneity. Sensitivity testing was used to determine the highest contributors to heterogeneity for the results. Subgroup analysis was conducted for salivary cortisol as well as serum cortisol, with a total comparison conducted as well. This subgroup analysis helped in determining if an overall pattern was apparent in cortisol changes or if one measurement of cortisol was irrelevant to cortisol level change. As for bias assessment, studies were rated as good, fair, or poor. Analysis was conducted including and not including studies found to be poor in order to better see if there was an effect if they were excluded; 95% confidence intervals were generated for the mean difference and standard mean difference effect size measurement.Statistical analysis and sensitivity testing were performed using RevMan software . HeterogResultsA total of four studies met the inclusion criteria for this study with a total of 135 participants . Out of this number, 126 participants  made up the salivary cortisol studies, and 9 participants made up the serum cortisol studies  -23. ContBias assessment of included studies is summarized in Table Meta-analysis findings are summarized in Table Discussion2= 0%) compared to sham treatment. Analysis of salivary cortisol alone versus a combined analysis of serum cortisol with salivary cortisol led to the same findings. The one study found to be poor on bias assessment ultimately did not make a substantial difference to findings; removing it led to near identical findings [The meta-analysis of four studies performed here found that osteopathic manipulative treatment overall led to a 0.10 μg/dl decrease  in salivary cortisol compared to sham treatments. This change is a significant change with low heterogeneity (p= 0.003, Ifindings .Further analysis of the results, however, shows that the effect size, as calculated by standard mean difference, suggests the change in cortisol determined through statistical analysis had low power. Considering the change in cortisol after a single treatment is such a low concentration, this finding is unsurprising. Additional treatments of OMT over time may show greater changes; there is reason to suggest a potential multiplicative effect of OMT if used multiple times. Previous research on OMT has found both acute and chronic differences due to OMT treatment -27. A loWhen comparing OMT effects on cortisol with chiropractic effects on cortisol, a similar decrease in cortisol is seen, albeit inconclusive -30. AlthHigh cortisol also shows a correlation with cognitive impairment in dementia, cardiovascular risk, complications in the elderly, and problems with depression -34. In aIn the future, this meta-analysis can be used as a guideline to pursue other patterns between biomarker levels and OMT. A low number of studies concerning the effects of OMT on serum levels of biomarkers prevents significant statistical power from being achieved. With more studies and conclusive findings, changes in biomarker levels after OMT can be viewed more frequently as an additional parameter when assessing the effectiveness of treatment. While encouraging, the inconsistent use of both serum and salivary measures for cortisol as well as the low effect size, pose limitations on the results of this study. A larger number of studies with consistent measuring protocols and larger sample sizes would fortify any correlations between OMT and changes in biomarker levels in future research. Nevertheless, the difference of 0.10 μg/dl lower serum cortisol levels compared to sham treatments may be additive with subsequent OMT and has the potential to provide benefit to patient care.


Non-invasive, prompt, and proper detection tools for kidney graft injuries (KGIs) are awaited to ensure graft longevity. We screened diagnostic biomarkers for KGIs following kidney transplantation using extracellular vesicles  from the urine samples of patients.One hundred and twenty-seven kidney recipients at 11 Japanese institutions were enrolled in this study; urine samples were obtained prior to protocol/episode biopsies. EVs were isolated from urine samples, and EV RNA markers were assayed using quantitative reverse transcription polymerase chain reaction. Diagnostic performance of EV RNA markers and diagnostic formulas comprising these markers were evaluated by comparison with the corresponding pathological diagnoses.EV CXCL9, CXCL10, and UMOD were elevated in T-cell-mediated rejection samples compared with other KGI samples, while SPNS2 was elevated in chronic antibody-mediated rejection (cABMR) samples. A diagnostic formula developed through Sparse Logistic Regression analysis using EV RNA markers allowed us to accurately (with an area under the receiver operator characteristic curve [AUC] of 0.875) distinguish cABMR from other KGI samples. EV B4GALT1 and SPNS2 were also elevated in cABMR, and a diagnostic formula using these markers was able to distinguish between cABMR and chronic calcineurin toxicity accurately (AUC 0.886). In interstitial fibrosis and tubular atrophy (IFTA) urine samples and those with high Banff chronicity score sums (BChS), POTEM levels may reflect disease severity, and diagnostic formulas using POTEM detected IFTA (AUC 0.830) and high BChS (AUC 0.850).KGIs could be diagnosed with urinary EV mRNA analysis with relatively high accuracy. The kidney transplant is becoming increasingly successful worldwide and in Japan ; howeverKGIs are first suspected by clinical findings, urine or blood chemical analyses, and imaging results; the definitive diagnosis is made on histology. Kidney allograft biopsies can now be performed relatively safely following the development of improved technology; however, the procedure is still somewhat invasive and not consistently feasible. Thus, biomarkers that can distinguish between the different types of KGI with high accuracy and low invasiveness are yet to be discovered –8.Recently discovered extracellular vesicles  in urine are gaining attention as a source of biomarkers, and we have previously investigated a diagnostic system of KGI using quantitative mRNA analysis of urine EVs –13. We h, Hyogo Prefectural Nishinomiya Hospital, Graduate School of Medicine, Hokkaido University, Sapporo Hokuyu Hospital, Graduate School of Biochemical and Health Sciences, Hiroshima University, Tokai University School of Medicine, and Osaka City University Graduate School of Medicine) approved the study in accordance with the approval of the ethical committee of the principal institution, Sapporo City General Hospital .This study was performed according to the Helsinki Declaration and complied with both the Ethical Guidelines for Medical and Biological Research Involving Human Subjects and the Ethical Guidelines of Japanese Society for Clinical Renal Transplantation. The ethical committee of each participating facility  reviewed all preparations by the Banff criteria  accordinAll stored urine samples were thawed for 5 min (15 min in the case of 50 mL samples) in a warm (37° C) water bath, and supernatants were collected after 800xG centrifugation. Next, we performed EV recovery and extracted mRNA from 10 ml of supernatant using ExoComplete Kit  according to the instruction manual [p was < 0.05 were considered statistically significant using the Welch t-test. Sparse Logistic Regression analysis was used to calculate each diagnostic formula as previously reported [Thirty-nine genes were analyzed by quantitative RT-qPCR as previously described . These ireported . BrieflyBackground and demographic information for the 127 patients enrolled in this study are shown in Table The results of qPCR analyses are shown in Fig. In contrast, the detection of cABMR was more challenging. Elevation of SPNS2 and SPDEF expression was observed in cABMR, although this was also the case for other KGIs. Previously, we reported that ANXA1 increased in TCMR and cABMR ; howevercCNIT is not graft rejection; however, it is difficult to clinically distinguish it from cABMR. Thus, we investigated ways to distinguish between cABMR and cCNIT by EV RNA. B4GALT1 and SPNS2 gene expressions were enhanced in cABMR but suppressed in cCNIT  relies on clinical manifestations such as a decrease in urine volume and fever, urine and blood analyses, blood chemical analysis, and radiographic evaluation such as ultrasonography, computed tomography, or radioisotope imaging. However, the definitive diagnosis is still made by graft pathology , 4. A grIn this study, we evaluated KGI using the measurement of mRNA obtained from urine EVs after having previously introduced the usefulness of a single gene, ANXA1, in the detection of graft injury in a single center analysis of kidney injury model . SubsequHere, 39 candidate genes selected based on our preparation study were analyzed using qPCR from 127 patients. CXCL9, CXCL10, SPDEF, SPNS2, and UMOD showed statistical differences between some graft rejection types. Among these, CXCL9/CXCL10 and UMOD were shown to be significant biomarkers of TCMR, as their expression showed robust enhancements in samples from patients with TCMR; in contrast, there was no increase in the expression of these genes in samples from patients with antibody-mediated rejection. Previous literature has stated that the chemokines CXCL9 and CXCL10 are significant biomarkers for detecting allograft rejection in animal models and a clinical multiple-institute study. Our present study clearly supports these results , 15, 16.B4GALT1 expression was increased in cABMR but decreased in cCNIT. Both KGIs induce gradual arteriole stenosis and consecutive tissue injury as a result of chronic ischemic changes. B4GALT4 is a promising gene biomarker for distinguishing between these two events and has critical significance given the contrary management of immunosuppressant dosing for these conditions. B4GALT1 is a gene encoding glycosyltransferase and influences B cell activation  and has SLC12A1 was identified as a candidate marker for reflecting the severity of IFTA by qPCR analysis. NKCC exists on the cell surface and has two variants, NKCC1 and NKCC2; NKCC2 is expressed only in kidney tissue and encoded by SLC12A1 . HoweverFinally, POTEM and HAVCR1 were nominated as the candidate biomarkers for the detection of BChS, which is supposed to correlate with graft loss. HAVCR1, also called TIM-1, is a known biomarker of kidney injury and has been proven to be a candidate marker for chronic KGI. While POTEM was also identified as a candidate biomarker, further study is needed regarding its mechanism of involvement in the progression of chronic graft damage.The results of the present study were obtained from transplant institutions in Japan; however, the sample size was limited. In addition, the recent progress of immunosuppressants along with a thorough understanding of the mechanism of graft rejection and graft pathology, lessen the probability of acute cellular rejection, making the distribution of KGI unequable. Moreover, immunosuppression protocol and follow-up policy were carried out differently by individual facilities. Lastly, donor type was skewed to living donors because of the extreme shortage of deceased organ donors in our country.This study focused on urine EVs as a biomarker source and developed a diagnostic modality for graft injury after kidney transplantation using qPCR of mRNA obtained from EVs. This modality was able to incorporate further specific gene explorations beyond those presented here. Moreover, by calculating the diagnostic formula using multiple gene combinations, detected graft injury was detected more accurately. The present study was a nationwide, multicenter study; however, further studies with larger sample sizes are required to validate its results. This study presents the groundwork for the future development of solutions for tough KGI, cABMR, and graft fibrosis. To promote this development, an upcoming prospective long-term study is schemed to ensure graft longevity.


The all-inside anterior cruciate ligament reconstruction (ACLR) technique was developed to improve patient outcomes by reducing the procedure’s invasiveness, minimizing complications and pain, and enabling faster postoperative recovery. This study presents a detailed description of the all-inside ACLR technique, which involves the use of quadrupled semitendinosus (ST) graft and suspension devices at both tibial and femoral sites, as well as valuable tips for avoiding complications that may arise during the procedure. The surgical procedure employs retrograde drills to create bony sockets for graft passage, which are then fixed with suspension devices at both the tibial and femoral sites. This technique has no specific restrictions and may be applied to all patients with anterior cruciate ligament (ACL) injuries. The literature reports the advantages, good clinical outcomes, and medium- to long-term graft survival achieved with the all-inside ACLR technique. However, the complications and disadvantages associated with the technique must be considered. Anterior cruciate ligament reconstruction (ACLR) is one of the most common procedures performed in orthopedic surgery; it is estimated that in the United States alone, 400,000 ACLRs are performed annually . Over thThe all-inside ACLR technique, unlike other ACLR procedures, has no significant restriction and may be applied to all patients with an anterior cruciate ligament (ACL) rupture, including adolescents, due to growth cartilage preservation performing sockets entirely within the epiphysis; this approach reduces bone growth damage risk and postoperative pain ,9,10. ThTherefore, several previous studies have comprehensively evaluated the all-inside ACLR technique, analyzing the clinical outcomes, graft rupture rates, and rehabilitation protocols ,10,11,12The study aims to describe our all-inside ACLR technique, which uses a quadrupled ST graft fixed with both tibial and femoral suspension devices, to provide procedure standardization and share our tips and tricks to avoid the complications that may occur with this ACLR technique.A thorough clinical examination under anesthesia is performed before positioning the patient on the operating table to confirm anteroposterior (AP) knee instability using the Lachman and Anterior Drawer test. Rotational stability is also assessed by performing the pivot shift test and varus/valgus stress to rule out other ligamentous injuries . The patThe surgical procedure begins with graft harvesting and preparation on a dedicated workstation by an assistant. Several tendons can be harvested as grafts, including the ST, gracilis, quadriceps, peroneus longus, and allografts ,15,16. OThe surgical procedure begins with an incision positioned 2 cm medial to the anterior tibial tuberosity and 4 cm distal to the joint line. A planar dissection is performed and arrives at the sartorius fascia, which is incised to identify the hamstrings. With the aid of an open tendon stripper, only the ST is harvested. The graft is cleaned and bathed with saline solution supplemented with 1 g vancomycin powder and is then placed on a workstation for subsequent graft preparation. The gracilis is usually retained unless the size of the ST is inadequate or has been prematurely amputated inadvertently.® No. 2; Arthrex, Naples, FL, USA) while keeping the two suture tails free  A. Then, ils free C. FiberWed state D.® No. 0; Arthrex), which annularly fixes the graft using multiple loops and creates an annular tag at a predefined distance of 15 mm  or 20 mm  from both ends of the graft  is passed through the femoral buttonholes of the adjustable-loop suspension device , leaving the tails free at the level of the tibial fixation device  and anteromedial (AM) arthroscopic portals are used .Tips. Perform careful Hoffa’s fat pad debridement for adequate articular visualization.The AM portals should be used as viewing portals to visualize the ACL femoral footprint better. With the knee positioned in 90° of flexion and the scope in the AM portal, the clinician inserts the outside-in aimer arm through the AL portal and places the head of the guide over the center of the native ACL femoral footprint, which is identified by taking the lateral intercondylar ridge and the lateral bifurcate ridge as a reference point ,8. The cTips. In case the footprint is not optimally visualized, the presence of a ligamentous remnant or narrow intercondylar gap is strongly suggested to establish a third portal, the accessory anteromedial portal (AAM), which provides good arthroscopic visualization of the femoral footprint, to make an accurate femoral tunnel.Tips. The aiming arm on the femoral side should be set at about 80 degrees, with a variability of about 5 to 10 degrees, depending on the lateral femoral condyle’s anatomical conformation, the soft tissues’ thickness, and the knee’s intercondylar notch depth. This aimer arm positioning allows easy visualization of the correct button flipping, checked by direct arthroscopic viewing using the AL portal as the entry point, and positioning of the scope at the lateral subcutaneous layer. Attempt to respect the femoral entry point of the guidewire, located approximately 2.5 cm proximal to the lateral femoral epicondyle, being careful not to stand too far anterior so as not to compromise the trochlear cartilage.The surgeon drills the outer lateral femoral cortical with the guide wire through the bullet until it exits the center of the intra-articular guide B,C. DireTips. ®, Arthrex) should be used during the anterograde drill advancement   F,G.Tips. Always look at the femoral socket to check that it has been correctly created circumferentially and that there are no soft tissues/bone remnants/reamer debris locked inside that could interfere with the graft’s passage. During the drilling phase, it is recommended to position the suction shaver from the AM portal to aspirate the debris from drilling.For a comprehensive view and to achieve a proper tibial socket, it is recommended that the AL and AAM portals be used for arthroscopic viewing, using the AM portal as the working portal.Through the AM portal, the tip of the ACUFEX™ Director ACL Tip Aimer (Smith&Nephew) is placed at an angle of 55–60° at the level of the ACL tibial footprint, anterior to the posterior border of the anterior horn of the lateral meniscus A,B. The ®, Arthrex) is placed as protection, and the same retrograde drilling  procedure is performed at the femoral level   B,C.®, Arthrex) and avoid soft tissue interposition. Specifically, the proximal section of the jaws is used to slide through the tibial shuttle suture, and the distal serrated tip of the jaws is used as a femoral shuttle suture grasper , are suggested to avoid contact between the graft and the skin.Tips. The AM portal must be dilated to permit graft introduction.The femoral shuttle suture is used through the AM portal to assist the passage of the femoral button of the adjustable-loop suspension device  through the femoral socket A,B.A “flip-then-fill” technique is used  and the Once the femoral button has been flipped E, the grTips. It may be helpful to partially position the femoral side of the graft at the level of the femoral socket and then pass the graft to the tibial socket so that the graft depth in the femoral socket may be better accommodated during tensioning. Usually, during tensioning of the femoral graft side, only 1 to 1.5 cm of the graft is pulled into the femoral socket. The graft progression inside the femoral socket is confirmed by the position of the annular tag at the femoral graft edge A,B.Tips. Proper flipping of the femoral button may be checked by direct arthroscopic viewing, using the AL portal as the entry point and positioning the scope at the lateral subcutaneous layer.The tibial button of the adjustable-loop suspension device  is passed through the tibial socket by pulling the tibial shuttle sutures C,D.Tips. ® No. 0; Arthrex), placed 15–20 mm from the extremity of the graft and tagged in the middle of the graft with an annular mark, allows for better adjustment during the graft pull maneuver at both the femoral and tibial sockets under arthroscopic visualization.A multi-strand high-strength suture (FiberWireTips. ® (Arthrex), the free tails of the suture tape should be retrieved along with the sutures of the tibial adjustable-loop suspension device.If an internal brace is used, consisting of a suture tape such as FiberTapeAfter ensuring that the tibial graft side has been appropriately engaged in the tibial socket, the surgeon begins to pull the white sutures of the tibial adjustable-loop suspension device alternately, to bring the tibial button in contact with the tibial cortical E–G.The tibial button should be fixed while keeping the knee in extension/hyper-extension and neutral rotation. The interposition of soft tissue between the tibial cortical and the tibial button during the tensioning phase should be avoided A.Cycling of the knee in flexion–extension through its range of motion (ROM) is performed to help accommodate the graft. Further tension is applied at this point by manually pulling the white sutures of the femoral adjustable-loop suspension device. An arthroscopic examination of the successful tension and tightness of the graft positioned in its anatomical site is performed using a probe B,C.After graft tensioning, the knee should be checked to ensure that full ROM is available. The white sutures of the tibial button are then tied at the top of the button and the remaining tails are cut off. At the femoral level, the white sutures of the femoral button may be secured using a knot-pusher under arthroscopic subcutaneous viewing, entering from the AL portal and placing the scope at the lateral subcutaneous layer, and, finally, the remaining tails are cut off D.Finally, layer suturing is performed at the graft harvest site, using No. 0 Vicryl sutures, along with skin suturing of the arthroscopic portals and the graft harvest area with No. 2-0 Prolene sutures.Tips. An excessively lengthy graft will dimple on the femoral or tibial socket floor and is, therefore, unacceptable since it fails to maintain proper graft tension.Tips. To promote greater tibial button–tibial bone contact and in cases where there is the suspicion of having maintained a short bone bridge at the level of the tibial socket, it is advisable to use an XTENDOBUTTON Fixation Device (Smith&Nephew), which has been designed to provide an extended tibial surface area of contact.Tips. If an internal brace (suture tape) is used, it must be held under tension during the tibial button tensioning maneuver and must then be tied and stabilized at the level of the tibial button by passing the extremities of the suture tape tails through the holes freed by the removal of the blue carrier sutures, or they could be anchored at the tibial level using fully threaded knotless SwiveLock (Arthrex) anchors, which have been designed for use with suture tape. During the internal brace fixation process, it becomes imperative to meticulously administer an equivalent or slightly reduced tension compared to the graft. This precautionary measure is crucial to prevent stress shielding.Tips. In case the graft appears loose after performing proper tensioning in the tibial site, it is recommended to perform the following procedures in this order:(1)Check the amount of graft in the tibial socket according to the position of the annular tag at a predetermined distance from both ends of the graft and the annular tag in the middle of the graft.(2)Tighten the femoral side further by approximately 5 mm.(3)Perform additional cycling of the knee.(4)Remove the graft from the tibial socket and repeat the retrograde reaming by increasing the drilling by 5 mm, put the shuttle suture back in place, retrieve it without soft tissue bridge interposition, and proceed with graft passage and tensioning.This study’s main finding is that the all-inside ACLR technique represents a feasible procedure that may be performed in all patients with ACL injury, including adolescents with open physis, reporting clinical and functional outcomes that are in line with other ACLR techniques ,18.Encouraging results were reported in the literature regarding the PROMs of patients undergoing ACLR using the all-inside technique; in general, similar or superior functional outcomes and a reduction in postoperative pain were described, compared with other ACLR techniques ,18. In tSchurz et al.  reportedThe all-inside ACLR technique allows anatomical graft placement at the tibial and femoral sides, providing physiological benefits for graft revascularization and ligamentization . This teThe all-inside ACLR technique allows for various graft choices, such as the autologous hamstring, quadriceps tendon, peroneus longus, and allografts. With double cortical suspension fixation, the necessary graft length is only about 6–7 cm; therefore, for example, the required length of 26–28 cm for graft preparation may be achieved with only ST . Conte eUsing half instead of complete tunnels, which preserves bone stock, is another advantage of the all-inside ACLR technique, especially in ACLR revision or multiple ligament reconstructions. Moreover, postoperative pain and swelling may be reduced since this technique does not violate the extra-articular cortex and periosteum. Several authors ,25,26,27Lastly, in the all-inside ACLR technique, an independent femoral guide facilitates precise anatomical placement on the femoral ligament footprint, heightening the probability of reinstating the knee’s natural kinematics. Furthermore, it mitigates potential hazards, including forming an excessively short tunnel, posterior wall blowout, and encroachment upon the lateral wall of the intercondylar notch, which instead represents significant issues to be considered with other ACLR techniques .®, Arthrex) to reinforce ACLR. Subsequently, several authors [Internal brace application in ACLR surgery is a relatively recent concept. Mackay et al.  were the authors ,16,30 re authors . In a re authors  included authors .The all-inside ACLR technique is complex, and orthopedic surgeons must be aware of the associated pitfalls. This technique’s overall complication rate is 5.89% ,21; in dThe all-inside ACLR technique also has some drawbacks. The learning curve of some procedures, such as graft preparation, drilling, and socket creation, is more complex and challenging compared to other ACLR techniques ; for thiThe all-inside ACLR technique was developed to improve ACLR surgical outcomes. This technique offers advantages such as less invasive surgery, reduced bleeding, faster recovery, and better graft tension control. Half, instead of complete, tunnel creation implemented quadrupled ST or quadriceps tendon as the grafts of choice. The all-inside ACLR technique applies to all patients with an ACL injury, including adolescents, and provides good clinical outcomes postoperatively, with medium- to long-term graft survival. Technique standardization may help avoid the potential complications associated with this procedure.


Portal vein (PV) reconstruction is a key factor for successful living-donor liver transplantation (LDLT). Anatomical variations of right PV (RPV) are encountered among potential donors.To evaluate a single center experience of reconstruction techniques for the right hemi-liver grafts with PV variations during the period between May 2004 and 2022.A total of 915 recipients underwent LDLT, among them 52 (5.8%) had RPV anatomical variations. Type II PV was found in 7 cases (13.5%), which were reconstructed by direct venoplasty. Type III PV was found in 27 cases (51.9%). They were reconstructed by direct venoplasty in 2 cases (3.8%), Y graft interposition in 2 cases (3.8%), and in situ double PV anastomoses in 23 cases (44.2%). Type IV PV was found in 18 cases (34.6%) and was reconstructed by Y graft interposition in 9 cases (17.3%), and in situ double PV anastomoses in 9 cases (17.3%).Early right posterior PV stenosis occurred in 2 recipients (3.8%). Early PV thrombosis occurred in 3 recipients (5.8%). The median follow-up duration was 54.5 months (4 – 185). The 1-, 3-, and 5-years survival rates were 91.9%, 86%, and 81.2%, respectively. Late PV stenosis occurred in 2 recipients (3.8%) and was managed conservatively.Utilization of potential living donors with RPV anatomic variations may help to expand the donor pool. We found that direct venoplasty and in situ dual PV anastomoses techniques were safe, feasible, and associated with successful outcomes. Liver transplantation is widely accepted as the only effective treatment for patients with end stage liver disease and selected hepatic malignancies . In EgypThe first adult LDLT was performed in 1994 . On the Portal vein (PV) reconstruction is a key factor to achieve a successful LDLT surgery. Anatomical variations of the right PV (RPV) are commonly encountered among potential donors. The incidence of portal venous variations had been heterogenous between different series ranging between 10% and 35% , 9. DiffThe current study was conducted to estimate the incidence of RPV anatomical variations among liver donors and evaluate different reconstruction techniques utilized in portal flow reconstruction for the right hemi-liver graft with PV anatomical variations. To the best of our knowledge, this is the first study from Egyptian centers addressing the feasibility and the outcomes of adult LDLT utilizing right hemi-liver grafts with PV anatomical variations.We retrospectively reviewed the data of all recipients who underwent LDLT at Liver Transplantation Unit, Gastrointestinal Surgery Center, Mansoura University during the period between May 2004 and May 2022. Recipients who underwent LDLT utilizing right hemi-liver grafts from living donors with RPV anatomical variations were identified and included in the current study. Recipients’ data were retrieved from a prospectively maintained database for all recipients undergoing LDLT. An informed consent was obtained from each recipient and donor. The study was approved by the Institutional Review Board and Local Ethical Committee at the Faculty of Medicine, Mansoura University, Egypt with reference number (R.23.01.2003).Preoperative evaluation protocol had been described previously , 14. In For donors, evaluation of PV anatomical variation was dependent of computed tomographic portography. Classification of PV anatomical variations was based upon Cheng classification where: Type I, classical anatomical division of the main PV; Type II, trifurcation of the main PV; Type III, the right posterior PV arises as the first branch of the main PV; Type IV, the right anterior PV arises from the left PV .The operative technique had been described before , 14. A sFor liver grafts with RPV anatomical variations, different techniques were utilized for PV reconstruction. If the two RPV branches were close to each other, as in type II PV, a common orifice was created by direct venoplasty utilizing polypropylene 6/0 sutures at the back table to be anastomosed as a single orifice to recipient PV. In some donors, the RPV was divided at the level of the trifurcation leaving a small bridge between the two PV branches. The intervening bridge was widened by venoplasty utilizing polypropylene 6/0 sutures at the back table. Similarly, a single PV anastomosis is created with the recipient PV Figs. .Fig. 1DiIf the two RPV branches were far from each other, as in type III and IV PV, we utilized either autologous Y-graft interposition technique or in situ direct dual PV anastomoses technique. For autologous Y-graft interposition technique, the recipient’s PV bifurcation was excised with the proximal main PV trunk and used as a Y-graft. The recipient right and left portal branches were anastomosed to the two separate graft RPV branches utilizing polypropylene 6/0 sutures at the back table. The distal end of the autologous Y-graft was then anastomosed to the remnant recipient main PV as a single anastomosis in the recipient Fig. C.For in situ direct dual PV anastomoses technique, the two separate graft RPV branches were anastomosed each to the recipient right and left PV branches utilizing polypropylene 6/0 sutures. This required an acceptable angulation of the recipient main PV axis Figs. D and 3. Intraoperative Doppler ultrasound was used to confirm the patency of all vascular anastomoses. A portal vein flow around 100 – 300 ml/min/100 g liver graft was considered adequate. After ensuring adequate hemostasis and biliostasis, abdominal drains were placed around the graft and near the raw surface.After surgery, all cases were transferred to the intensive care unit (ICU) for monitoring. Patients underwent detailed laboratory evaluation twice daily during the ICU stay. Oral intake and ambulation were allowed once bowel sounds were restored. Patients were transferred to the ward on the fifth postoperative day, depending on clinical improvement. Doppler ultrasound (US) played the main role in the follow-up of the recipients. Our postoperative protocol included doppler US examinations once daily during the first week, day after other during the second and third weeks and before hospital discharge, and then once weekly during the following 2 months. Further radiological workup was done upon suspicion of any postoperative complications.After discharge, patients were followed up regularly on outpatient visits. Follow-up visit included detailed history taking, clinical examination, detailed laboratory evaluation including trough level of immunosuppression drugs, and Doppler US evaluation of hepatic vasculature.Postoperative morbidity is defined as any adverse event occurring during the first 90 postoperative days and is graded according to the Clavien-Dindo classification. Major complications are defined as class 3 or higher . Postopep value less than 0.05 was considered statistically significant.Shapiro–Wilk test was used to assess the normality of the data. Categorical variables were expressed as number and percentage, and continuous variables were expressed as median and range. Survival analysis was performed by Kaplan–Meier analysis. Statistical analysis was performed using the SPSS 22 software . A During the period between May 2004 and May 2022, 915 recipients underwent LDLT at Liver Transplantation Unit, Gastrointestinal Surgery Center, Mansoura University. Fifty-two recipients (5.8%) underwent LDLT utilizing right hemi-liver grafts from living donors with RPV anatomical variations and were included in the current study.The preoperative demographic data of the study cases were summarized in Table The operative data of the study cases were summarized in Table Type II PV was found in 7 cases (13.5%), which were reconstructed by direct venoplasty of the double PV stomas and single PV anastomosis in the recipient. Type III PV was found in 27 cases (51.9%). They were reconstructed by direct venoplasty of the double PV stomas and single PV anastomosis in the recipient in 2 cases (3.8%), Y graft interposition (from recipient PV) in the bench surgery and single PV anastomosis in the recipient in 2 cases (3.8%), and in situ double PV anastomoses in the recipient in 23 cases (44.2%). Type IV PV was found in 18 cases (34.6%) and was reconstructed by Y graft interposition (from recipient PV) in the bench surgery and single PV anastomosis in the recipient in 9 cases (17.3%), and in situ double PV anastomoses in the recipient in 9 cases (17.3%).The postoperative data of the study cases were summarized in Table The long-term follow-up data of the study cases were summarized in Table The global demands for liver transplantation have been progressively increasing worldwide in the last decades. On the other hand, there is a global shortage of organs that can meet the current demands resulting in a major organ shortage problem . The proThe availability of an appropriate living donor is the most vital step for LDLT. However, several factors can lead to rejection of the available living donor . TransplPortal venous reconstruction is a crucial and technically demanding step in liver transplantation . Recent In the past, the anatomical variations of the RPV were considered contraindications for living donation. However, the improvement of surgical techniques and familiarity of the surgical procedure allowed transplantation teams to accept living donors with RPV anatomical variations allowing further expansion of the potential donor pool. Different surgical techniques had been described to overcome the RPV anatomical variations like back wall venoplasty (unification venoplasty), autologous Y-graft interposition utilizing either the recipient’s own PV confluence or cryopreserved iliac veins, in situ direct dual PV anastomoses, and Malatya approach utilizing autologous saphenous vein graft –13. HoweIn the current study we evaluated our center experience of different reconstruction techniques utilized in portal flow reconstruction for the right hemi-liver graft with PV anatomical variations. In the beginning of our experience, we utilized autologous Y-graft interposition technique utilizing the recipient’s own PV confluence for reconstruction of widely separated two RPV stomas. Autologous Y-graft interposition technique was used in 2 cases 3.8%) of Type III PV and 9 cases (17.3%) of Type IV PV in our series. It should be noted that our group experienced several complications after utilizing autologous Y-graft interposition technique especially thrombosis of right posterior PV (3 cases—5.8%). This is commonly attributed to twisting or kinking of the long right posterior PV , 22, 23.% of TypeIn the current practice, our group adopted the use of in situ double PV anastomoses in the recipient instead of autologous Y graft interposition technique for cases with widely separated two RPV stomas. In situ double PV anastomoses technique was used in 23 cases (44.2%) of Type III PV and 9 cases (17.3%) of Type IV PV in our series with adequate intraoperative PV flow. It should be noted that we did not experience any PV complications after adoption of in situ double PV anastomoses technique by our group. Kuriyama et al. recently evaluated the feasibility and outcomes of direct dual PV anastomoses in LDLT using the right hemi-liver graft with anatomical PV variations. They utilized direct dual PV anastomoses technique in 9 adult recipients with Type III PV variation and achieved successful results of all recipients .In situ double PV anastomoses technique is simple and can be easily performed. Although it may be associated with some twist of the axis of main PV, however, it is not associated with significant PV flow changes. In few recipients with type III or IV PV, the two separate graft RPV stomas were anastomosed each to the corresponding recipient right anterior and posterior PV branches with closure of the recipient left PV stump to prevent over twisting of the axis of main PV. The main drawbacks of this technique are the prolonged warm ischemic time during performance of the dual PV anastomoses, and it could not be performed for recipients with higher PV thrombosis grades .An additional important issue among potential donors with RPV anatomical variations is the association with right hepatic duct variations , 27. SucThe current study had some limitations. Firstly, it is a single center retrospective study which is liable to selection bias. Secondly, the number of included cases is relatively small owing to the unique nature of such vascular variation; however, a future multi-center study may help to recruit more cases.In conclusion, anatomical variations of the RPV are commonly encountered among potential living donors. Transplant surgeons should be familiar with such anatomical variations and different techniques for their reconstruction. We found that direct venoplasty of the double PV stomas and in situ dual PV anastomoses techniques are safe, feasible, and associated with successful outcomes. Utilization of potential living donors with RPV anatomic variations may help to expand the donor pool for LDLT.