Sympathetic hyperactivity is one of the main mechanisms of secondary hypertension. Reducing renal sympathetic activity through surgery can effectively reduce blood pressure. Many cases have shown that renal denervation (RDN) can selectively block renal artery sympathetic nerve activity to control refractory hypertension. This surgery is a minimally invasive surgery, and the risk of surgery-related adverse events is significantly reduced compared with surgery. Therefore, the purpose of this study is to explore the efficacy of radiofrequency ablation of renal artery sympathetic nerve in the treatment of secondary hypertension. Eight patients with secondary hypertension diagnosed by the cardiovascular department of our hospital and treated with RDN were followed up for 3-18 months, of which 5 cases were followed up for more than 12 months and 8 cases were followed up for more than 3 months. Eight patients were treated with radiofrequency ablation of renal artery catheter. The parameters such as preoperative blood pressure, antihypertensive drugs, organ function, intraoperative ablation resistance, power, time, and temperature were determined. The related changes of blood pressure, antihypertensive drugs, and visceral function and the occurrence of side effects at 1 week and 1, 3, 6, and 12 months after operation were related to the operation. In conclusion, RDN has a significant clinical effect in the treatment of refractory hypertension, with stable postoperative blood pressure drop, reduced drug dosage, and less side effects. The hyperactivation of sympathetic nerve is one of the main mechanisms of secondary hypertension, and reducing renal sympathetic nerve activity by surgery can effectively reduce blood pressure –3. Many In this paper, we have thoroughly investigated the curative effect of renal artery sympathetic radiofrequency ablation in the treatment of secondary hypertension. For this purpose, a total of 8 patients with secondary hypertension confirmed by the Department of Cardiovascular Medicine of our hospital and receiving RDN were followed up for 3-18 months, among which 5 patients were followed up for >12 months and 8 patients were followed up for >3 months. Eight patients were treated by radiofrequency ablation of renal artery catheter line RDN, preoperative blood pressure, antihypertensive drugs, viscera function, intraoperative ablation resistance, parameters such as power, time, temperature, and postoperative 1-week and 1-, 3-, 6-, and 12-month blood pressure, antihypertensive drugs, relevant changes of the viscera function, and the occurrence of side effects related to the operation situation, and the clinical efficacy and safety of RDN in the treatment of secondary hypertension were evaluated.The remaining parts or sections of this paper are organized according to the following plan which is given below.In subsequent section, that is, Section-Eight patients were selected with secondary hypertension diagnosed in our hospital during 2020.06-2021.06. Inclusion criteria: patients who were diagnosed as secondary hypertension without prior formal treatment, renal and adrenal space occupation detected by CT, or related endocrine or immune system examination suggested secondary causes. The diagnosis was in accordance with the 1999 WHO/ISH cutoff criteria for hypertension, i.e., systolic blood pressure > 140 mmHg (1 mmHg = 0.133 kPa) and/or diastolic blood pressure > 90 mmHg. The statistical nodes of follow-up data were 1 week and 1, 3, 6, and 12 months after surgery. The data of this study were obtained from inpatient and outpatient departments.Thermo Cool Smart Touch is a multielectrode cavity catheter with an adjustable bend tip designed for electrophysiological mapping of the heart and radiofrequency current transfer to the catheter's tip electrodes for ablation. Pipe body size is 7.5 F (with 8 F ring electrode). For ablation, the catheter must be used in conjunction with a radiofrequency ablation instrument and a diffusion plate . The catheter has force sensing technology that provides a real-time measurement of the contact pressure between the catheter tip and the heart wall.ω. All 9 patients had pain of different degrees during operation, and all of them were treated with analgesic drugs. Postoperative local hematoma at puncture site occurred in 1 case, and the rest had no adverse reactions. The patients were followed up 1 week and 1, 3, 6, and 12 months after the operation, including family self-measured blood pressure, 24 h ambulatory blood pressure, renal function, N, NE, dopamine, and surgery-related side effects. Index decline criteria: defined as the reduction of systolic blood pressure in the postoperative office = 10 mmHg, and the antihypertensive drugs decreased compared with the preoperative level. Combined with the specific surgical plan to adjust the parameters, the traditional RDN operation was explored and improved.Improve the preoperative examination, eliminate the contraindications, and make a comprehensive preoperative discussion according to the condition of the operation plan. Femoral artery was punctured in all patients, RFA/8F arterial sheath was indwelled, and heparin 3000 U was injected into the sheath. A 6 F JR3.5 angiography catheter was used to send the tip of the catheter into the left and right renal artery openings under fluoroscopy for renal arteriography to understand the diameter, length, and presence of obvious stenosis or dissection of renal artery, and determine the surgical plan. By guiding sheath pipe thread into the renal artery bifurcation, connect the ablation catheter handle by renal artery radiofrequency ablation instrument; using X-ray imaging equipment along the thread to promote completely into the renal artery catheter until electrode, slowly push the handle on the electrode button electrode joint blood vessel walls, and start the renal artery radiofrequency ablation instrument to stick wall impedance detection. According to the parameters of renal artery radiofrequency ablation instrument, 360° ring radiofrequency ablation was performed. Generally, 6-12 points are ablated at 10-12 W at each point, the rf time is 30-40 s, the RF temperature fluctuates between 30 and 60°C, and the RF impedance is 80-180 X (mean) ± S (standard deviation), comparison between groups was compared by t-test, and P < 0.05 was considered statistically significant.SPSS 17.0 statistical software was used for analysis, measurement data were expressed as β-blockers, 40.0% ARB, and 100% CCB. 80.0% took diuretics and about 60.0% took direct vasodilators.In this study, a total of 5 patients were followed up for 12 months. The mean preoperative systolic blood pressure was 165.84 ± 7.21 mmHg, and the mean diastolic blood pressure was 97.88 ± 10.27 mmHg. They took 5.98 ± 0.87 kinds of antihypertensive drugs, including 100% P < 0.05). The mean diastolic blood pressure decreased at 1 week, 1 month, 3 months, 6 months, and 12 months after surgery, and there was no statistical significance except 1 week after surgery, but there was statistical significance at 1 month, 3 months, 6 months, and 12 months after surgery (P < 0.05). The differences of mean systolic blood pressure and mean diastolic blood pressure 1 week after surgery and 6 and 12 months after surgery were statistically significant (P < 0.05). The mean systolic blood pressure at 1 month after surgery was significantly lower than that at 12 months after surgery (P < 0.05).As shown in Among the 8 patients, 5 were male, 3 were female, 4 were smokers, 3 were diabetic, 4 were cardiac insufficiency, 4 were renal insufficiency, and 2 were Sjogren's syndrome. During the operation, all the 8 patients had different degrees of pain, which were cured by fentanyl or morphine. Local hematoma at the puncture site occurred in 1 case after surgery, which was considered to be related to inadequate compression at the femoral artery puncture site after surgery. The hematoma was gradually absorbed and dispersed after compression dressing. One case had left renal artery reverse dissection during surgery, and no abnormality was found after stent implantation. Postoperative pseudoaneurysm at puncture site occurred in 1 case, and hematoma was gradually absorbed and dissipated after local pressure dressing and fistula reanastomosis. No obvious hypotension, infection of puncture site, renal artery stenosis, puremia, thromboembolism, arteriovenous fistula requiring blood transfusion and/or surgical treatment, and other adverse events and side effects occurred during and after operation. Blood pressure decreased steadily after RDN treatment, and renal function was not significantly damaged, so RDN was relatively safe.The etiology of secondary hypertension is clear , but the hidden etiology needs efforts to find, and its onset is more dangerous and harmful –9. AccorStudies have shown that the sympathetic nervous system is a special regulatory system between the kidney and the brain and plays an important role in the blood pressure regulation of hypertensive patients. Local norepinephrine isotope dilution was used to measure renal norepinephrine outflow in patients with untreated hypertension –20. MoniAt the beginning of the last century, renal sympathetic nerve to nerve in the surgical treatment in hypertension , 31, advP < 0.05), indicating that RDN treatment for refractory hypertension has definite clinical efficacy. Moreover, with the extension of time, the antihypertensive effect is still meaningful, and the efficacy of RDN for refractory hypertension is not temporary or transient. The mean systolic blood pressure at 1 week and 1 month after surgery had no significant difference compared with that at 3 and 6 months after surgery, but the blood pressure decreased significantly compared with that at 12 months (P < 0.05), indicating that the blood pressure decreased most significantly at 1 month after surgery, and the blood pressure decreased steadily with the extension of time. Antihypertensive drugs decreased before and after operation, and the difference was statistically significant (P < 0.05). Follow-up data of eGFR, Scr, and BUN at 1 week, 3 months, and 12 months after operation showed no significant difference, indicating that RDN had no significant effect on renal function. There were statistically significant differences in the decrease of epinephrine and norepinephrine (P < 0.05) indicating that the renal sympathetic nerve activity was decreased compared with that before surgery, but there were no statistically significant differences among groups 1 week, 3 months, and 12 months after surgery. These results indicate that RDN can effectively reduce renal sympathetic nerve activity. Due to renal sympathetic nerve ablation, sympathetic nerve activity is weakened, RAAS system activity is weakened, blood pressure is decreased, and the corresponding antihypertensive drugs are also decreased, which can not only reduce drug-related side effects and reduce medical costs but also stabilize blood pressure, reduce blood pressure-related risks, and ensure the health of patients. At the same time, the preoperative and postoperative heart rate and liver function were followed up in this study, and the difference was not statistically significant. The results showed that RDN had no significant effect on liver function and heart rate.Results of this study showed that systolic blood pressure and diastolic blood pressure of 5 patients followed up for 1 year at 1 week, 1 month, 3 months, 6 months, and 12 months after surgery were significantly decreased compared with those before surgery, and the difference was statistically significant  identify indications: clear which part of the group for RDN is the primary problem, RDN can effectively reduce the renal sympathetic nerve activity, lower blood pressure, the concept can not only be used in the treatment of refractory hypertension, its indications should be renal sympathetic nerve activity hyperfunction caused by a series of disease. (2) How to find and evaluate indicators of sympathetic overactivation: although epinephrine, norepinephrine, dopamine, aldosterone, and other indicators can reflect renal sympathetic activity, reliable indicators with high specificity and sensitivity are still lacking at present, and more studies are still needed to further explore appropriate indicators for evaluation. (3) Further improvement of surgical methods: the structure and shape of renal arteries and sympathetic nerves are different in different individuals, so the surgical methods are also different. How to explore a better surgical method and block more sympathetic nerves as much as possible is worth further discussion and research.RDN not only has significant antihypertensive effect but also can rapidly and steadily relieve blood pressure after surgery. It is also safe and has few side effects. No serious complications occurred during the 12-month follow-up after surgery, which is suitable for clinical promotion.


Lachnospiraceae in control group, Enterobacteriaceae in NYHA class III, and Escherichia in NYHA class IV. The concentration of PAGln was significantly different between CHF and control groups and increased with the severity of heart failure. Finally, the correlation analysis represented that Parabacteroides and Bacteroides were negatively correlated to brain natriuretic peptide (BNP) and PAGln; Romboutsia and Blautia adversely associated with PAGln; Klebsiella was positively interrelated with BNP; Escherichia-Shigella was positively correlated with PAGln and BNP; Alistipes was contrasted with BNP; and Parabacteroides was negatively correlated with the left ventricular end-diastolic diameter (LVEDD). This study presented that the intestinal flora and its metabolite PAGln were altered with different grades of CHF and illustrated the effects of the gut flora and its metabolite on CHF.Chronic Heart Failure (CHF) is the end result of nearly all cardiovascular disease and is the leading cause of deaths worldwide. Studies have demonstrated that intestinal flora has a close relationship with the development of Cardiovascular Disease (CVD) and plays a vital role in the disease evolution process. Phenylacetylglutamine (PAGln) a metabolite of the intestinal flora, is one of the common chronic kidney disease toxins. Its concentrations in plasma were higher in patients with major adverse cardiovascular events (MACE) however, its variation in patients with various degrees of CHF has rarely been reported. Therefore, we collected stool and plasma samples from 22 healthy controls, 29 patients with NYHA Class III and 29 patients with NYHA Class IV CHF (NYHA stands for New York Heart Association) from the Department of Cardiology of Shanghai Fengxian District Central Hospital. Next, we analyzed these samples by performing bacterial 16S ribosomal RNA gene sequencing and liquid chromatography tandem mass spectrometry. The result shows: The Chao 1 index was significantly lower in both NYHA class III and NYHA class IV than it was in the control group. The beta diversity was substantially dissimilar across the three groups. The linear discriminant analysis effect size analysis (LEfSe) showed that the bacterial species with the largest differences were  Chronic Heart Failure (CHF) is a major health problem worldwide and is the end result of almost the entire forms of cardiovascular disease –3. The pWe selected 58 patients of CHF hospitalized in the department of cardiology of Shanghai Fengxian District Central Hospital from June 2021 to June 2022 for this study (CHF group). There were 26 male cases and 32 female cases, aged 52–89 years. In terms of comorbid diseases, there were 22 patients with coronary heart disease, 3 patients with hypertension, and 33 patients with coronary heart disease combined with hypertension. According to the New York Heart Association (NYHA) cardiac function classification, 29 patients were categorized as class III and 29 patients were categorized as class IV. Furthermore, 22 patients with the same CHF risk factors but without clinical manifestations and a previous medical history of CHF were chosen as the control group. There were 13 male patients and 9 female patients, aged 65–86 years. In terms of comorbid diseases, there were 9 patients of coronary heart disease and 13 patients of coronary heart disease combined with hypertension. The research was authorized by the Ethics Board of Shanghai Fengxian District Central Hospital, and all the patients (or their direct relatives) signed the informed consent form.The exclusion standards included (1) hypertrophic or restrictive cardiomyopathy, myocarditis, acute myocardial infarction, and congenital heart disease; (2) diabetes mellitus, diarrhea, Crohn’s disease, ulcerative colitis, infectious diseases, and malignancy; (3) severe hepatic and renal insufficiency; (4) having taken antibiotics, steroid hormones, herbal medicines , or probiotics (such as yogurt) in the 3–6 months before admission; and (5) a substantial change in diet in the week before admission.We collected 5 ml of venous blood within 24 h of admission from both groups, the samples were mixed in EDTA anticoagulation tubes, and 1 ml of plasma was separated and preserved at −80°C in an ultra-low temperature refrigerator. The plasma PAGln level was measured through liquid chromatography tandem mass spectrometry. The plasma samples (100 μL) were vortexed with methanol (400 μL), sonicated and centrifuged . The supernatant was subsequently removed and analyzed on the machine. Next, chromatographic separation was performed on a column Acquity UPLC HSS T3 1.8 μm  with the following conditions: flow rate, 0.30 ml/min; column temperature, 40°C; injection volume, 5 μl; mobile phase composition, A, water (0.1% formic acid) and B, methanol; and sample gradient elution program, 0–1.0 min 10% B. The PAGln was detected using an AB Sciex Triple Quad 5,500 mass spectrometer with the setting thermoelectric spray ion source mode. The data were collected using the multiple reaction monitoring mode with the parameters 263.2 Da for Q1 Mass and 145.2 Da for Q3 Mass. The declustering potential was −29 V, and the collision energy was −15 V. The MultiQuant software was used for the integration, and the PAGln content was calculated using the standard curve.We collected 5.0 g of fresh early morning stools from patients in the CHF and control groups within 24 h after admission in sterile tubes, and the sampling tubes were placed in self-sealing bags. The samples were delivered to the laboratory within 2 hours and stored in an ultra-low temperature refrigerator set at −80°C. The DNA extraction for the microbial community was performed according to the instructions of the DNA Extraction Kit (DP712), and the purity and concentration of the DNA were tested using a Thermo NanoDrop One after DNA extraction.The regions V3 to V4 of the bacterial 16S ribosomal RNA (rRNA) gene were amplified by the primers 338F (5′-ACTCCTACGGGAGGCAGCAG-3′) and 806R (5′-GGACTACHVGGGTWTCTAAT-3′). The polymerase chain reactions (PCRs) were performed in a volume of 50:25 μl of 2 × Premix Taq, 1 μL of the forward and reverse primers, and 50 ng of DNA. The PCR cycling was performed at a BioRad S1000 thermal cycler . The reaction conditions of the PCR system were 94°C pre-denaturation for 5 min; 30 cycles of denaturation at 94°C for 30 s, annealing at 52°C for 30 s, extension at 72°C for 30 s; and finally extension at 72°C for 10 min. The PCR products were purified by electrophoresis on 1% agarose gels. Product concentrations of PCR were compared using GeneTools analysis software . The volume needed for each sample was computed based on the equal mass principle and each PCR product was mixed. The PCR mix was recovered by using the EZNA Gel Extraction Kit , and the target DNA fragments were recovered with elution with TE buffer.The library were constructed following the standard procedure of the NEBNext Ultra II DNA Library Prep Kit for Illumina . The raw sequences were then sequenced using an Illumina Nova 6000 platform for PE250 sequencing, and the sequencing data were processed using fastp (0.14.1) software. The sequence information for the different treatment groups was clustered into Operational Taxonomic Units (OTUs) for species classification according to the UPARSE clustering method, and the similarity of the OTUs was set to 97%. The species annotation information was obtained by comparing them with the SILVA (16S) database.trialos.com (SL2021-KY-25-02).Informed and written agreement was acquired from all patients before the trial. The Ethics Committee of Shanghai Fengxian District Central Hospital approved the experimental protocol. The trial was registered at Chi-square test. For the normally distributed measures, the data from two groups were compared using the independent samples t-test; and the data from multiple groups were compared by the one-way ANOVA test. For the non-normally distributed measures, the data from two groups were compared by using a Mann-Whitney U test, and the data from multiple groups were compared by using the Kruskal-Wallis test. Spearman’s rank correlation was employed to analyze the correlation between variables. All the statistical analyses were performed on SPSS (version 26.0), and the significance was setting to p < 0.05. The alpha-diversity (Shannon and Chao1), non-metric multidimensional scaling, redundancy analysis/canonical correlation analysis (CCA), and correlation analysis were determined on the Magigene Cloud Platform1.Enumeration data were compared with p > 0.05). Heart rate, serum creatinine, blood urea nitrogen, and diuretics were significant differences across the three groups (p < 0.01).The general baseline characteristics of all participants are presented in p < 0.001). The difference in LVEF between the three groups was statistically significant (p < 0.01). Pairwise comparison showed significant difference. Additionally, the differences in LVESD and LVEDD between the control group and NYHA class IV were statistically significant (p < 0.01), LVESD and LVEDD between NYHA class III and NYHA class IV were statistically significant (p < 0.01), and the differences in LVESD and LVEDD between the control group and NYHA class III were not statistically significant (p > 0.05). Also, the results of the comparison between each paired group showed that the differences in the BNP and PAGln levels in the three groups were statistically significant (p < 0.01).The comparison of the echocardiographic indices, brain natriuretic peptide (BNP), and PAGln levels of all the participants is shown in r = 0.924, p < 0.001), LVESD , and LVEDD  were positively correlated with PAGln, and LVEF  was negatively correlated with PAGln.In The species composition and similarity of the intestinal flora in the CHF and control groups were compared using a Venn diagram. As p > 0.05). The Chao 1 index was lower in both NYHA class III and NYHA class IV than in the control group, and the difference was statistically significant(p < 0.05). The above results indicated that the more severe the degree of heart failure, the lower the number of flora species in heart failure patients. Furthermore, the Shannon index was not statistically significant (p > 0.05) among the control, NYHA class III, and NYHA class IV groups, indicating that the intestinal flora diversity was essentially the same in all three groups of patients. Non-metric multidimensional scaling is a method that is used to characterize the differences in microbial communities by reducing the dimensionality of the sample distance matrix, simplifying the data structure, and showing the distribution of the samples at specific distances. It can be used to analyze the dissimilarity in the microbial community composition among the three groups of samples. As shown in p < 0.05). Bacteroidetes was the second most represented strain in the three groups, with 32.8, 32.8, and 32.1% in the control, NYHA class III, and NYHA class IV respectively, with no statistically significant difference among the three groups (p > 0.05). The percentage of the remaining phyla, such as Proteobacteria and Actinobacteria, was relatively low. The abundance percentages of Escherichia-Shigella, Agathobacter, Megamonas, Bifidobacterium, Klebsiella, Streptococcus, and Lactobacillus in the three groups were significantly different at the genus level (p < 0.05). Linear discriminant analysis effect size (LEfSe) was used to conduct the colony difference analysis to find the biomarkers. The method integrates statistical analysis of the differences and the score of the influence of that difference on the grouping results while emphasizing statistical significance and biological relevance. The results are shown in Lachnospiraceae in control group, Enterobacteriaceae in NYHA class III, and Escherichia in NYHA class IV.Wilcox rank sum test was used to analyze the significance of the differences in the α-diversity index  of the three groups. As shown in Parabacteroides and Bacteroides were negatively correlated with BNP and PAGln, and Romboutsia and Blautia were negatively correlated with PAGln. Moreover, Klebsiella was positively correlated with BNP, and Escherichia-Shigella was positively correlated with BNP and PAGln. Alistipes was negatively correlated with BNP, and Parabacteroides was negatively correlated with LVEDD.A CCA analysis can explain the relationship between clinical phenotypic data (such as PAGln in this study) and the gut microbial community . The totp < 0.05). According to the above results, the more severe the level of heart failure, the lower the number of flora species in heart failure patients. The β-diversity analysis revealed significant separation between the three groups of sample points and clustering within a certain range, indicating differences in the gut flora composition between the three groups. This indicates that there is a connection in the severity of heart failure and the distribution and components of the gut flora. The research demonstrated that the intestinal perfusion of CHF patients was insufficient, and the intestinal structure and function were changed, causing a clear reduction in the diversity of the intestinal flora (p < 0.05), with a marked reduction in the Firmicutes abundance in the CHF group. It was also discovered that Bacteroidete’s abundance was raised and Firmicute’s abundance was reduced in the CHF patients, and the abundance of Firmicutes and intestinal microbial diversity were positively correlated with the intake of dietary fiber (Escherichia-Shigella (1.7:7.2:6.6), Bifidobacterium (0.8:4.2:1.6), Klebsiella (0.5:1.34:4.7), and Lactobacillus (0.6:1.17:4.28) had significantly different abundance shares at the genus level (p < 0.05). The LEfSe analysis demonstrated that at the genus level, Agathobacter was significantly higher in the control, and Escherichia and Bifidobacterium were significantly higher in NYHA class III. Then, Klebsiella and Lactobacillus were significantly higher in NYHA class IV. The genera that were statistically different in both test analyses were Bifidobacterium, Klebsiella, and Lactobacillus. Moreover, Bifidobacterium was higher in NYHA class III than that in NYHA class IV, and Klebsiella in NYHA class III was lower than that in NYHA class IV. This suggests that Bifidobacterium and Klebsiella may be the key organisms that change under different degrees of CHF. Studies have shown that Klebsiella is elevated in cardiovascular disease and that it regulates the trimethylamine synthesis pathway, which can be metabolized by intestinal microbes to produce trimethylamine N-oxide (TMAO). Additionally, researches have found that elevated plasma TMAO levels in CHF patients predict a higher risk of death , inhibiting the generation of pro-inflammatory cell factors, regulating host organism immunity, and enhancing the gut environment by depressing ammonia concentration in feces and intestinal pH. Therefore, the increased abundance of Bifidobacterium with heart failure could be the consequence of a feedback mechanism of the organism, acting against pathogenic bacteria and toxic mediators. For instance, some studies have shown an increased abundance of Bifidobacterium in heart failure patients , LVESD , LVEDD  were positively correlated with PAGln, and LVEF  was negatively correlated with PAGln. The CCA analysis showed that five significant clinical phenotypic data explained 72.7% of the species distribution. The arrows representing BNP and PAGln had the longest lengths, indicating that BNP and PAGln had the greatest effect on the change in the flora; the BNP and PAGln arrows had similar lengths, indicating that both have similar effect sizes on the flora of the CHF group. Hyperactivation of the sympathetic nervous system is a distinctive characteristic of CHF, and prolonged activation can lead to the worsening of CHF. Thus, PAGln can act on the β2-adrenergic receptors in the heart, resulting in hyperexcitation of the sympathetic nervous system, which can exacerbate heart failure , and the plasma PAGln concentrations were substantially higher in the patients in NYHA class IV than those in NYHA class III (p < 0.001), indicating that the more severe the degree of heart failure, the higher the PAGln concentration. The correlation analysis revealed that Parabacteroides and Bacteroides were negatively correlated with BNP and PAGln, and Romboutsia and Blautia were negatively correlated with PAGln. Klebsiella was positively correlated with BNP, and Escherichia-Shigella was positively correlated with BNP and PAGln. Furthermore, Alistipes was negatively correlated with BNP, and Parabacteroides was negatively correlated with LVEDD. Inflammation and myocardial fibrosis are the basic pathological mechanisms of heart failure. With heart failure, there is a lack of intestinal motility, impaired absorption in the mucosal villi, and damage to the mucosal barrier of the intestinal epithelium, which subsequently leads to increased intestinal permeability and failure to remove bacteria and food residues from the intestinal lumen promptly. This promotes the proliferation of Gram-negative bacteria such as Klebsiella, resulting in the easy entry of toxic mediators such as lipopolysaccharides (LPS) into the circulation, further stimulating the activation of the inflammation and immune response and exacerbating the progression of heart failure. Therefore, we suggest that Klebsiella exacerbates the progression of heart failure by releasing LPS. Also, we speculate that PAGln is a microbial metabolite that is present in Klebsiella, and the role and mechanism of Klebsiella in the development of heart failure should be investigated more thoroughly in the future. In addition, there were significant differences in the LVEF, LVESD, LVEDD, and BNP indices across the three groups. These indices are often involved in the diagnosis and differential diagnosis of heart failure, assessing the extent of the disease, and evaluating the prognosis, and are widely used in clinical practice. In mice given an ob/ob and high-fat diet, Wang et al.  can be found at: NCBI repository, accession number The studies involving human participants were reviewed and approved by Ethics Committee of Shanghai Fengxian District Central Hospital. The patients/participants provided their written informed consent to participate in this study.BC and ZZ conducted the investigation and wrote the original draft. BC, ZZ, YS, and HD reviewed and edited the manuscript. HW and ZQ provided the conceptualization and funding. ZQ was the supervisor and conducted project administration. All authors contributed to the article and approved the submitted version.


Patients with lupus nephritis (LN) are in urgent need for early diagnosis and therapeutic interventions targeting aberrant molecular pathways enriched in affected kidneys.We used mRNA-sequencing in effector (spleen) and target  tissues from lupus and control mice at sequential time points, and in the blood from 367 individuals (261 systemic lupus erythematosus (SLE) patients and 106 healthy individuals). Comparative cross-tissue and cross-species analyses were performed. The human dataset was split into training and validation sets and machine learning was applied to build LN predictive models.In murine SLE, we defined a kidney-specific molecular signature, as well as a molecular signature that underlies transition from preclinical to overt disease and encompasses pathways linked to metabolism, innate immune system and neutrophil degranulation. The murine kidney transcriptome partially mirrors the blood transcriptome of patients with LN with 11 key transcription factors regulating the cross-species active LN molecular signature. Integrated protein-to-protein interaction and drug prediction analyses identified the kinases TRRAP, AKT2, CDK16 and SCYL1 as putative targets of these factors and capable of reversing the LN signature. Using murine kidney-specific genes as disease predictors and machine-learning training of the human RNA-sequencing dataset, we developed and validated a peripheral blood-based algorithm that discriminates LN patients from normal individuals (based on 18 genes) and non-LN SLE patients (based on 20 genes) with excellent sensitivity and specificity (area under the curve range from 0.80 to 0.99).Machine-learning analysis of a large whole blood RNA-sequencing dataset of SLE patients using human orthologs of mouse kidney-specific genes can be used for early, non-invasive diagnosis and therapeutic targeting of LN. The kidney-specific gene predictors may facilitate prevention and early intervention trials. Prediction of patients with systemic lupus erythematosus (SLE) that will develop nephritis and early diagnosis represents an unmet need because of the limited value of known predictors and the invasiveness of kidney biopsy.Even with best treatment up to 40% of patients fail to reach a complete renal response suggesting that early diagnosis and prompt treatment including targeting of renal specific pathways is needed.Distinct, renal-specific molecular pathways are associated with the development of nephritis and its progression from subclinical to full blown disease in murine SLE.The mouse kidney transcriptome mirrors the human whole-blood transcriptome in lupus nephritis (LN).Upstream and downstream regulators of the cross-species (murine and human) kidney-specific gene signatures have been identified as putative targets in LN and novel cross-species drug signatures for kidney disease in lupus.Using the mouse kidney-specific transcriptome and through training by machine-learning techniques of a large whole-blood RNA-sequencing dataset of SLE patients, we developed and validated an algorithm that predicts patients that will develop LN based on a small number (no more than 20) of genes.Common cross-species (murine and human) genes could be prioritised as potential therapeutic targets for LN or tested as an alternative, non-invasive ‘liquid biopsy’ marker of kidney disease in patients with SLE.The mouse kidney-specific set of gene predictors may be used towards monitoring human kidney disease in SLE patients and enrolment in LN prevention and early treatment studies.In lupus nephritis (LN), current therapy fails to induce remission in more than 50% of patients. Even in cases with clinical remission, repeat kidney biopsies often exhibit residual inflammation and increased fibrosis, with 15%–20% of patients eventually developing end-stage kidney disease.Subclinical (silent) LN represents an early stage in the natural history of the diseaseIn this paper, we performed sequential mRNA-sequencing studies in effector (spleen) and target tissues  from lupus and healthy mice, as well as in the whole blood of patients with systemic lupus erythematosus (SLE) (including patients with active or responding LN or neuropsychiatric lupus) and healthy individuals. Comparative cross-tissue and cross-species analyses yielded common, cross-species, nephritis-specific genes that could be prioritised as potential therapeutic targets. Using machine-learning algorithms, we constructed a clinical-transcriptome predictive model that can be tested as a non-invasive ‘liquid biopsy’ marker of kidney disease in patients with SLE, to be used for monitoring of kidney disease in SLE, as well as enrollment in LN prevention and early treatment studies.Patients with SLE (n=261) who met the SLICC 2012 or EULAR/ACR 2019 classification criteria and age-matched and sex-matched healthy individuals (n=106) were recruited from the Departments of Rheumatology and Nephrology at the University Hospitals of Heraklio, ‘Attikon’ University Hospital and the respective Blood Transfusion Units. Active LN was defined by the presence of proteinuria more than 0.5 g/day and active urine sediment. A kidney biopsy was performed in all patients with evidence of active kidney disease. Patients either developed active LN de novo or had had a history of LN and were flaring at the time of sampling. Responding LN was defined by preservation or improvement of kidney function with reduction of proteinuria to less than 50% after 6 months of therapy or less than 0.5–0.7 g/day by 12 months.NZB/W-F1 mice were sacrificed at the prepuberty (1 month old), preautoimmunity (3 months old) and nephritic (6 months old with proteinuria more than 200 mg/dL for three consecutive days) stage of SLE. Age-matched C57BL/6 mice were used as controls. Spleen, kidneys and brain were removed for RNA extraction.2 search engine.RNA libraries were prepared using the Illumina Truseq kit. Paired-end 37 bp (for mouse) and 67 bp (for human) mRNA-sequencing was performed on the Illumina HiSeq2000 and HiSeq4000, respectively, at the University of Geneva Medical School.The human mRNA-sequencing dataset was randomly split into training (70%) and validation (30%) sets. Using the training set and feature selection algorithms, the smallest set of human orthologs that most accurately predicted the outcome of interest was selected. Using these orthologs as predictors, models were fit and compared for their ability to predict human disease. To improve performance, clinical predictors (not included in the definition of active or responding LN) were added to the final model. Accuracy, sensitivity, specificity and area under (AUC) the receiver operating curve (ROC) were determined in the validation set.https://1drv.ms/u/s!Au_gakpSntTbrGO3-3RQ39ByOId1?e=MLF007.Detailed information for all methods can be found in 10.1136/annrheumdis-2021-222069.supp1Supplementary dataFCGR2B, PTPRC, ITGAM, NCF1 and RASGRP1 , kidneys and brain (major end-organ tissues) from NZB/W-F1 lupus mice and age-matched C57BL/6 healthy counterparts. Tissues were collected at the clinical (nephritic) stage of the disease when nervous system involvement also occurs. Differentially expressed genes (DEGs) in lupus versus healthy mice tissues were analysed. Using genes differentially expressed within kidneys of the NZB/W-F1 lupus mice but not in other tissues studied, we defined a ‘kidney-specific signature’ comprising 726 DEGs  . Enriche RASGRP1 .PTPRC, NCF1 and ITGAM genes, as well as the IRF8,Next, we examined the molecular events underlying transition from the preclinical to clinical stage of lupus kidney disease by comparing DEGs between the tissues from lupus mice probed at the prepuberty versus the nephritic stage. Genes that were differentially expressed uniquely within kidneys of the NZB/W-FI lupus mice but not in other tissues studied defined the ‘kidney-specific LN-transition signature’ comprising 507 DEGs   that werTo analyse the sequential molecular events underlying the evolution towards LN, we identified DEGs in tissues from lupus vs healthy mice demonstrating a strain-specific effect in a time-series analysis. DEGs within kidneys demonstrating the lupus-specific pattern were combined with genes within kidneys that were differentially expressed across all stages of the disease. Combined signatures were compared across tissues and genes that were differentially expressed uniquely within kidneys—but not in other tissues—defined the ‘sequential kidney-specific signature’, composed of 1668 genes . FunctioKidney biopsy, an invasive procedure linked to increased risk for adverse events, is currently essential to confirm diagnosis and guide therapeutic decisions in LN; however, it is still an imperfect predictor of response to treatment. Previous studies have reported shared molecular signatures within LN kidneys of mice and humans,NCF2, ITGAM, NCF1, RASGRP1 and FCGR2ANext, we examined whether the human peripheral blood from patients with LN shares common gene expression aberrations with the mouse kidney-specific gene signatures. Using the human orthologous genes of the mouse genome, we examined if the mouse ‘kidney-specific signature’ is present in the blood of patients with active LN as compared with healthy individuals. A total 272 genes (193 upregulated and 79 downregulated) were common between the two datasets , referreCD53, ITGB2 and LAPTM5 were the highest-degree hub genes, underscoring their role in evolution of LN. The risk locus ITGAX was also identified, further supporting its pathogenic roleA similar cross-species analysis was performed to determine whether the mouse ‘kidney-specific LN-transition signature’ intersects with the human blood transcriptome of patients with active LN versus non-LN patients. Ninety-seven common genes (67 upregulated and 30 downregulated) were identified , comprisTo characterise the ‘sequential kidney-specific signature’ in the context of human LN, we compared the human orthologous genes of the mouse signature with the DEGs between active LN patients and healthy individuals and revealed 609 common genes that defined the ‘shared sequential kidney-specific signature’ . These gIn silico analysis of upstream regulators, downstream kinases and drug signatures for the identification of novel therapeutic targets in LN: Kinases TRRAP, AKT2, CDK16 and SCYL1 as putative targets for reversing the LN signatureGenetic association studies have identified TFs to play a major pathogenic role in SLE.‘shared active LN signature’ , we detected the top 50 drugs or small molecule compounds . To remove noise and keep the smallest set of human orthologs of the mouse genes which best predicts outcome, we performed feature selection using recursive feature elimination with a random forest (machine-learning) model under a 10-fold cross-validation. Based on model accuracy, a set of 50 human orthologs were selected. Next, prediction models were fit to identify which performs best with the selected genes. The glmnet model using 18 genes—including and POLE — and IL10RA gene . Male sex and younger age of SLE patients were associated with higher probability of active LN. In the validation dataset, the glm model displayed accuracy 81.7% (95% CI (0.70% to 0.90%)), sensitivity 63.2% and specificity 90.2% (AUC 0.80) in distinguishing patients with active LN from SLE patients without history of LN  in a non-invasive manner. We sought that the kidney-specific gene expression profile of lupus mice at the clinical (nephritic) versus the preclinical (prepuberty) stage of the disease  could reflect the whole-blood gene expression profile of SLE patients with active LN versus SLE patients without history of LN (non-LN patients). Thus, we used the human orthologs of the mouse ‘kidney-specific LN-transition signature’ as predictors, and applied feature selection under a 10-fold cross-validation. Based on accuracy, 20 genes best predicted the outcome. Models were fit to identify which performs best with the selected genes. Model performance was further improved by the addition of age, sex and presence of anti-dsDNA, as predictors of outcome. As expected, due to the higher likelihood of patients with proliferative LN to have anti-DNA antibodies, the presence of anti-dsDNA was the most important predictor of kidney disease, followed by the expression of ry of LN , demonstry of LN . TogethePatients with LN are in need for an early diagnosis and therapeutic targeting of aberrant molecular pathways enriched within the affected kidneys. Here, we performed sequential mRNA-sequencing in three tissues of lupus and healthy mice, and in the whole-blood of SLE and healthy individuals. Through cross-tissue analysis, we defined a murine kidney-specific molecular signature and a molecular signature that underlines progression from the predisease stage to overt clinical disease. We also demonstrated that the murine kidney transcriptome mirrors—in part—the human whole blood transcriptome of LN patients and found upstream and downstream transcriptional regulators that may be prioritised as potential therapeutic targets. Finally, we developed a blood gene-based predictive model for human LN that can be tested as an alternative, non-invasive ‘liquid biopsy’ marker of kidney disease in patients with SLE. Pending further confirmation, this marker could identify patients in need of monitoring for development of LN, as well as enrolment in LN prevention and early treatment studies.To improve therapeutic interventions and optimise the use of animal models, gene expression profiling across three samples and species is important in defining how mouse biology can be extrapolated to humans.PTPRC, ITGAM, NCF1 and IRF8 genes), reinforcing their pathogenic role in LN and the progression from preclinical to clinical kidney disease. Validating our results, the VEGF, TLR2 and SOCS3 genes were also differentially expressed in the kidneys from NZB/W-F1 mice 9 months old vs 6 months old as well as the kidneys from patients with LN.et al,ITGAM and FCGR2B were also differentially expressed in the ‘kidney-specific gene signature’. The FPR2, IL18R1, ITGAM and NCF4 genes were also differentially expressed in the myeloid lineage from paediatric patients with LN,MDP1, PTGR1 and MX2 were also differentially expressed within the kidneys from LN patients, as assessed by microarraysTMEM167A, TNFAIP8 and VCAM1 were also differentially expressed in kidney tubular cells from LN patients.In view of the heterogeneity of lupus, we used next-generation sequencing as an unbiased and not requiring a priori hypothesis approach to uncover novel molecular pathways implicated in major end-organ injury in SLE. Initially we performed mRNA-sequencing of a peripheral lymphoid organ  and two end-organ tissues (kidneys and brain) from the NZB/W-F1 lupus model at the prepuberty, preautoimmunity and nephritic stage of SLE and identified the molecular profile which is expressed uniquely within kidneys of this model—but not in other tissues studied—and the molecular profile that characterises unique-to-kidney molecular events underlying LN transition from the preclinical to clinical stage of kidney disease. In this process, we identified pathways enriched within each signature and found that hub genes correspond to lupus susceptibility risk loci (such as the CEACAM1, TYMP, NCOA7 and AIM2 were also differentially expressed in interferon stimulating genes identified through single-cell RNA-sequencing within the kidneys from LN patientsSERPINA1, IL1RN and ABCB1 genes were also differentially expressed in kidney tubular cells from LN patients.Blood transcriptome analysis identified similarities as well as differences from the molecular signatures detected within kidneys in patients with LN, underscoring that limitations exist in the use of blood for uncovering kidney disease processes.To decipher cross-species specific targets in LN, we used systems biology approaches and combined our experimental data with simulation-based analyses. We report upstream and downstream regulators of the cross-species kidney-specific gene signatures as specific targets in LN and describe novel cross-species drug signatures for kidney disease in lupus, suggesting non-immune-based approaches to be tested in LN therapeutics, as ‘add on’ therapy to conventional immune therapy. We must underscore that due to limitations in the analysis, identified TFs are not restricted to immune cells therefore therapies targeting them could have off-target effects with potential toxicity.Although current therapeutic decisions in LN are guided by its histological classification,In conclusion, common cross-species, nephritis-specific genes could be used as potential therapeutic targets for LN or tested as a surrogate, non-invasive ‘liquid biopsy’ marker of kidney disease in patients with SLE. These kidney-specific genes can be used to design prevention and early intervention trials, following their validation in longitudinal studies.


Lactobacillus reuteri CCFM1040 (CCFM1040) was found to be an inhibitor of the common pathogenesis of AR and asthma in our previous studies. This study presented a randomized, placebo-controlled trial to investigate the clinical effects of CCFM1040 on both diseases. The total symptom score (TSS), the quality of life (QoL), and the modulation in the gut microbiota of patients with AR, the Asthma Control and Test (ACT) of patients with asthma, and the safety of both AR and asthma were measured. In patients with AR, CCFM1040 numerically decreased TSS, Rhinoconjunctivitis Quality of Life Questionnaire (RQLQ), 3 nasal scores in TSS , and sleep and significantly improved (P = 0.014) non-nose/eye symptoms. The ACT score was numerically increased in patients with asthma . Significant microbial (from class level to genus level) and metabolic differences (P < 0.05) were found in patients with AR. No adverse reactions were observed. No effect on the blood and urine routine indexes. CCFM1040 has a potential benefit on both diseases. Further studies based on these findings will help to optimize the management of AR and asthma.The coexistence of allergic rhinitis (AR) and asthma reinforces the concept of “one airway, one disease,” which has prompted the exploration for a single intervention to treat both diseases.  Allergic rhinitis (AR) and asthma are the commonest chronic inflammatory disorders of the airways worldwide and can affect people of any age, impairing quality of life (QoL) , 2. Currvia aspiration  was found to be significantly reduced in the intestines of individuals with both AR and asthma  was isolated from agricultural unpasteurized milk. The effect of CCFM1040 on targets with type 2 immune inflammation and gut microbiota was identified in mice . Patients with a 1-month history of antibiotics use, pregnancy, and breastfeeding were excluded. The informed consent form approved by the Research Ethics Committee was signed by all the participants before starting any study procedure.The patients were randomized at a ratio of 1:1 to receive CCFM1040 or placebo, using computer-generated block randomization. The study flowchart is shown in The patients filled out self-report online questionnaires at baseline, the midpoint (week 4), and the end of the study. The total symptom score (TSS) form and the Rhinoconjunctivitis Quality of Life Questionnaire (RQLQ) were filled out by patients with AR and AR with asthma, and the Asthma Control and Test (ACT), by those with asthma and AR with asthma.The TSS form measures both nasal symptoms  and eye symptoms (itchy/red eyes and watery eyes), which scored 0 (no symptoms), 1 (mild), 2 (moderate), or 3 (severe). These scores are summed to produce the TSS (range: 0–18).The ACT measures the adequacy of asthma control and change in asthma control as reported by the patients: a score of 20–25 points denotes that asthma is well-controlled, 16–19 points, partially controlled asthma, and 5–15 points, uncontrolled asthma.The RQLQ is a valid scale that assesses the QoL according to how the symptoms affect the patients with AR. It consists of 28 questions covering seven domains . The average score for all items ranges from 0  to 6 (extremely).16S rRNA genes were amplified with specific primers (341F and 806R) for the V3-V4 region. The amplicon libraries were pooled and subjected to sequencing on an Illumina MiSeq High-Flux Sequencing Platform , which was described previously . The eviously , 23.U/Kruskal-Wallis (nonparametric) tests. The hierarchical structure of taxonomic classifications to the median abundance (quantitatively) and the nonparametric Wilcoxon Rank Sum test  was used to depict the taxonomic differences between microbial communities   classifiable read counts was generated after Illumina MiSeq sequencing of 90 samples. The number of counts in the samples ranged from 277 to 15,226. All data were subjected to total sum scaling normalization. The R phyloseq  and vegaBlood routine and urine routine were detected before and after treatment. Adverse events were recorded at each visit.t-test and the analysis of covariance (ANCOVA). Both 95% CI and P-values were provided for differences between the CCFM1040 group and the placebo group.The data on rhinitis symptoms, asthma symptoms, and QoL were analyzed using GraphPad Prism version 8  and SPSS version 22.0 . The data are expressed as mean ± SEM. The differences between the groups were compared using the A total of 99 patients with AR (77), AR with asthma (11), and asthma (11) were enrolled. All the patients were of Han nationality, had lived in their current residence for more than 1 year, and had a carbohydrate-based diet; 45 patients with AR completed the questionnaires and had their fecal samples analyzed, and eight patients with asthma completed the questionnaires  to  for nasal congestion,  to  for watery eyes, and  to  for rhinorrhea. A higher reduction in mean sneezing score was observed in the CCFM1040 group than in the placebo group after 4 weeks of intervention , while a recovery was observed until 8 weeks . For itchy/red eyes and itching, no differences relative to placebo were observed in the CCFM1040 group.After 4 weeks of the intervention, the TSS of the patients with AR was lower in the CCFM1040 group than in the placebo group , and this efficacy was magnified until the 8th week  . FurtherP = 0.105 vs. placebo)  than a decrease in the placebo group during 8 weeks of the intervention  .P = 0.506 vs. placebo) to   and  and were greater for sleep  and  across the 8-week intervention period  . Regardin period .16S rRNA gene sequencing was not significantly (P > 0.05) different between the groups . After an 8-week intervention, the influence of CCFM1040 in the gut microbiota was analyzed. The α-diversity indices were examined to analyze the microbial diversity (P > 0.05) in the CCFM1040 group compared with the placebo group. The hierarchical clustering of the heatmap revealed significant alterations (P < 0.05) from the class level to the feature level, which resulted in distinct clustering of microbes in the CCFM1040 group and the placebo group (P > 0.05) was found between the two groups before the intervention. After the intervention, CCFM1040 and placebo groups both altered some of the abundances of the microbes, but they were completely different. The gut bacteria from the phylum Proteobacteria to the genus Escherichia_Shigella and the order Intestinibacter were significantly decreased in the CCFM1040 group compared with the baseline, whereas the placebo markedly decreased the abundance of the gut microbes under the class Bacilli compared with both the baseline and the CCFM1040 group. These results indicated that CCFM1040 modulated the gut microbiota structure in a fundamentally different way compared with the placebo in patients with AR.The baseline gut microbiota profile as per iversity . The Obsbo group –4. It ise groups . After aP < 0.05), and the novobiocin biosynthesis was dramatically inhibited (P < 0.05) after the CCFM1040 intervention compared with the placebo. Meanwhile, compared with the baseline, CCFM1040 significantly altered (P < 0.05) 18 biological pathways, which were mainly associated with carbohydrate, energy and lipid metabolisms, xenobiotic biodegradation and metabolism, metabolism of cofactors and vitamins, and immune system. Placebo, however, altered only two metabolic pathways: apoptosis and aminobenzoate degradation as compared with before the intervention.To characterize the functional composition profiles after CCFM1040 intervention, we predicted the gut microbial functions. A total of 328 biological pathways were identified. As shown in CCFM1040 treatment was well-tolerated, and no side effects or adverse events were recorded. After the 8-week intervention, blood and urine specimens were provided by more than 80% of the patients. There was no significant impact of the intervention on the patients' blood and urine routine .Lactobacillus on both diseases in adult patients.Both AR and asthma have been emphasized the “one airway, one disease,” because they share similar type 2 inflammation. It is exciting but unknown to explore a single intervention to treat both diseases. Previously, we had conducted animal experiments on CCFM1040 and demonstrated its favorable effect on targets with type 2 inflammation by modulating gut microbiota and metabolism. In this study, we investigated the beneficial effect of CCFM1040 on patients with both AR and asthma. To the best of our knowledge, this is the first study that reported the simultaneous effect of Our data showed that CCFM1040 not only improves symptoms and the QoL in patients with AR but also strengthens the control in patients with asthma. This is an important finding because it supports the hypothesis that supplementation with one single intervention, such as the “missing” agricultural microbes, might be used to treat AR and asthma simultaneously.P < 0.05), which suggest that these indicators have adequate sensitivity in the microbiological intervention trial.Currently, there are no accepted criteria for determining the optimal parameters from regulatory authorities or academia for clinical microbiological intervention trials. In general, patient-rated TSS and RQLQ are preferred as a primary measure of efficacy in AR trials for drug development, but some researchers reported that RQLQ may insufficiently or inaccurately capture the efficacy of anti-AR drugs , 30. ForThe multiple causes of AR and asthma in patients complicate the management of these conditions . Based oP < 0.05) in patients with AR. This is consistent with the previous results: microorganisms can improve certain indicators of AR  and altered composition of the gut microbiota , sufficient patients with AR were recruited, while few with asthma and AR with asthma only were recruited. During the study, our patients were not comprehensive enough when filling out the online questionnaires and were reluctant to provide stool samples. This had a major impact on the comparison between CCFM1040 and placebo due to a drastic decrease (42%) in the number of samples we could analyze  and the Collaborative Innovation Center of Food Safety and Quality Control in Jiangsu Province, and the 111 Project (Grant No. BP0719028).The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.


Experiencing a greater number of stressful days is associated with a heightened risk of cardiovascular disease. This study examined whether positive affect reactivity, the trait-like change in positive affect in response to daily stressors, moderates the association between the number of stressful days and blood pressure. Participants were 664 adults from the National Study of Daily Experiences II (Midlife in the United States sub-study). Participants reported stressors and positive affect across 8 days and provided resting blood pressure measures during a separate clinic visit. A greater number of stressful days was associated with increased systolic blood pressure, but only among individuals with higher positive affect reactivity . Results suggests that individuals who maintain positive affect when experiencing stressors may have lower risk of heightened systolic blood pressure, contributing to the growing evidence that positive affective reactivity may be protective against daily stress.


The relationship between arteriolar hyalinosis and renal progression in immunoglobulin A nephropathy (IgAN) is not fully understood. We aimed to investigate the clinicopathological features and outcomes of IgAN with or without arteriolar hyalinosis.A total of 762 diagnosed with IgAN patients were retrospectively analyzed. We classified IgAN patients into two groups with or without arteriolar hyalinosis. Then, the clinicopathological characteristics of the two groups were compared. We used Kaplan–Meier survival analysis to compare the composite kidney outcome of the two groups and applied multivariate Cox regression analyses to test the association between arteriolar hyalinosis and composite kidney outcome.p = 0.002) is an independent risk factor for renal prognosis in IgAN patients.Overall, 412 (54.1%) patients had arteriolar hyalinosis, including 173 patients diagnosed with hypertension. IgAN patients with arteriolar hyalinosis were older and had higher proteinuria, urea, uric acid, and blood pressure, while lower eGFR than those without arteriolar hyalinosis. Subgroup analysis showed similar results in IgAN patients with hypertension. Kaplan–Meier survival analysis showed that IgAN patients with arteriolar hyalinosis had worse composite kidney outcome than those without arteriolar hyalinosis. In addition, subgroup analysis revealed that patients with hypertension have worse composite kidney outcome than those without hypertension. Multivariate Cox regression analyses confirm that arteriolar hyalinosis (HR 2.57; 95% CI 1.41–4.69; Our study demonstrated that arteriolar hyalinosis is a common vascular lesion in IgAN patients. Arteriolar hyalinosis connects closely with hypertension, and arteriolar hyalinosis is an independent risk factor for renal prognosis in patients with IgAN. Immunoglobulin A nephropathy (IgAN) is the most common primary glomerular nephropathy globally . A systeIt is well-known that the clinical features of IgAN mainly include proteinuria, microscopic hematuria, high blood pressure (one or more symptoms) , And theIn recent years, studies on the intrarenal vascular lesions of IgAN have attracted significant attention. For example, intrarenal vascular lesions, including arterial intimal thickening, arteriolar hyalinosis, and microangiopathy, are common in IgAN . A previA total of 762 patients with IgAN diagnosed by renal biopsy were admitted to the nephrology department, Xinqiao Hospital, Army Medical University between January 2012 and December 2018. These clinicopathological materials were retrospectively analyzed. This study was approved by the ethics committee of Xinqiao Hospital and adhered to the principles of the Declaration of Helsinki. Our study has obtained informed consent from all the patients.Patients with (a) diagnosed with IgAN by renal biopsy and (b) a regular medical history with follow-up data in Xinqiao Hospital (c) the number of glomeruli ≥ 8 were included. While patients with (a) follow-up less than 6 months, (b) no exact blood pressure value, (c) had already progressed to end-stage renal disease (ESRD), (d) diabetes or malignant tumors or other autoimmune disorders, (e) thrombotic microangiopathy in the renal biopsy were excluded. The flowchart is shown in We collected the clinical data of every patient at the time of renal biopsy. These data include sex, age, height, weight, blood pressure, and inflammation markers which include C-reactive protein (CRP) and tumor necrosis factor α (TNF-α), hemoglobin, platelet, serum complementary 3 (C3), serum complementary 4 (C4), serum albumin, urea, serum creatinine, serum urea acid, total cholesterol and total proteinuria of 24 h, and estimated glomerular filtration rate (eGFR) was calculated using the CKD epidemiology collaboration (CKD-EPI) equation . HyperteAll renal biopsy specimens were processed routinely for immunofluorescence microscopy and light microscopy, Sections were stained for direct immunofluorescence with fluorescein isothiocyanate–conjugated antibodies specific for human IgA, IgG, IgM, C3, C4, and C1q. Immunofluorescence results were graded on a scale of 0 to 3, and a score ≥1 was regarded as immunofluorescence positive in this study. Sections used for light microscopy were stained with hematoxylin and eosin, periodic acid–Schiff, and Masson’s trichrome stain. Sections for light microscopy were graded according to the Oxford MEST-C score of IgAN . Two patArteriolar hyalinosis consists of the deposition of hyaline material into the vascular wall coupled with matrix protein synthesis. Accumulation of a large amount of amorphous, eosinophilic, glassy material replaces the media and has resulted in thickening and rigidity of the wall and apparent attenuation of the lumen . And in The primary outcome of interest was a composite kidney outcome defined as more than a 30% reduction in eGFR , or ESRDt-test was applied for normally distributed data, and the Mann-Whitney U test was applied for non-normally distributed data. The chi-squared test was applied for enumeration data. And the Kaplan–Meier method works to estimate renal survival time. A log-rank test was used to compare differences between renal survival curves. Univariate and multivariate Cox regression analyses were performed to test the association between arteriolar hyalinosis and the primary event of interest and reported hazard ratio (HR) with 95% confidence interval (CI) and p values. Multivariate Cox models included univariate, Model 1, Model 2, and Model 3. Model 1 included clinical variables such as age, sex, BMI, TNF-α, mean arterial pressure (MAP), proteinuria of 24 h, total cholesterol, uric acid, eGFR, and Model 2 is model 1 plus pathological variable such as global glomerulosclerosis, mesangial hypercellularity (M), endocapillary proliferation (E), segmental glomerulosclerosis (S) and tubular atrophy/interstitial fibrosis (T) and crescents (C), deposition of C3, deposition of C4. Model 3 is model 2 plus Renin-angiotensin-aldosterone System (RAAS) blockade, other immunosuppressants, and glucocorticoid. All p values were two-tailed, and values < 0.05 were considered statistically significant. The statistical analysis was performed by the SPSS statistics  and GraphPad Prism .All continuous variables were expressed as the mean ± standard deviation  or median with interquartile range  for data distribution. Enumeration data were expressed in percentages. The 2 (IQR 60.97–110.93). In addition, over 50% of the patients were diagnosed with arteriolar hyalinosis under renal biopsy, and the representative images of arteriolar hyalinosis are shown in p < 0.001), had a higher proportion of hypertension , heavier proteinuria , higher serum uric acid , and higher total cholesterol , while a lower level of eGFR . Furthermore, renal biopsy findings showed a higher proportion of global glomerulosclerosis  and a higher deposition rate of C3  in IgAN with the arteriolar hyalinosis group. Besides that, there were also significant differences in mesangial hypercellularity , segmental glomerulosclerosis , and tubular atrophy/interstitial fibrosis  between the two groups in the Oxford classification. Interestingly, our work found that more patients in IgAN with arteriolar hyalinosis were treated with glucocorticoid  and immunosuppressants .A total of 762 patients with IgAN were included; the essential characteristics are shown in p < 0.001; p < 0.001; p < 0.001; p = 0.002). Furthermore, compared with patients without arteriolar hyalinosis, we found that patients with a higher ratio of arteriolar hyalinosis (26–50%) have a much higher risk of composite kidney outcome  patients in IgAN without arteriolar hyalinosis group and 57 (13.8%) patients in IgAN with arteriolar hyalinosis group reached the composite kidney outcome. Kaplan-Meier curve analysis showed survival rate in IgAN with arteriolar hyalinosis was significantly lower than that of patients without arteriolar hyalinosis , uric acid , serum creatinine , and cholesterol  than those without hypertension. In addition, in the pathological aspects, we found that the patients with hypertension had a higher percentage of T1/T2 , global glomerulosclerosis , and 26–50% of arteriolar hyalinosis  but a lower proportion of C1/C2  than patients without hypertension. Moreover, the Kaplan–Meier curve showed that the survival percentage of patients with hypertension was not significantly different from patients without hypertension in the no-arteriolar hyalinosis group  patients were diagnosed with hypertension. The clinicopathological features are listed in p = 0.027), but after adjusting for age, sex, BMI, TNF-α, proteinuria, total cholesterol, uric acid, eGFR, global glomerulosclerosis, MEST-C scores, mesangial deposition of C3, mesangial deposition of C4, RAAS blockade, other immunosuppressants and glucocorticoid (Model 3), the outcome suggests that the hypertension is not the independent risk factor for the progression of the composite kidney event in the cohort of arteriolar hyalinosis patients .As shown in In the current study, we found that more than half of patients with IgAN have arteriolar hyalinosis. Patients with arteriolar hyalinosis have poor renal outcomes than patients without arteriolar hyalinosis, supporting that arteriolar hyalinosis is an independent predictor of the prognosis of IgAN patients.The pathologic features of IgAN were complicated and extensive. Some just manifested like minimal change disease, some showed the focal segmental glomerular sclerosis, but some yet had mild mesangial proliferation and immunofluorescence indicated the deposition of IgA . At presMeanwhile, intrarenal vascular lesions were common in pathological reports of renal biopsy ,14. As wFurthermore, intrarenal vascular lesions remain to be shared in IgAN. Some studies have reported that the manifestations of vascular lesions are arterial wall fibrotic intimal thickening, arteriolar hyalinosis, arteriolar endotheliocyte swelling, arteriolar inflammatory cell infiltration, and arteriolar thrombosis in IgAN. The risk factors of these lesions are associations with hypertension, higher serum creatinine and uric acid, high urinary protein excretion, global glomerulosclerosis, tubular atrophy, and interstitial fibrosis . And in Our study confirmed significant differences in clinical and pathological features between the patients with arteriolar hyalinosis and without arteriolar hyalinosis in IgAN. Myllymaki et al.  analyzedAnd univariate analysis  showed tMeanwhile, there are striking differences between the prognosis of the two groups, which implies that arteriolar hyalinosis is a pathological feature of poor prognosis. Furthermore, as the degree of arteriolar hyalinosis increases, the prognosis worsens. In addition, we have observed the relationship between arteriolar hyalinosis and hypertension. Wang et al.  analyzedA few limitations are able to make bias between the fact and our study. The first is that this was a retrospective study from the single-center, with limited sample capacity and a not too long follow-up period. But at the same time, the end event of our research is a composite kidney event. Therefore, our outcome needs to be proved by more extensive samples in multicenter by longer follow-up time. Second, we have just classified arteriolar hyalinosis into three degrees according to the relationship between the extent of deposits and the vessel wall diameter, but we have not calculated the index of arteriolar hyalinosis and not collected the data of fundoscopy or echocardiography. Above all may lead to a little deviation between this study and those of the public. These points need to be further evaluated in subsequent studies.In conclusion, our results indicated that a large proportion of IgAN patients have arteriolar hyalinosis, which is an independent risk factor for renal progression. And the higher the degree of affected arterioles is, the greater the risk of worse renal outcomes will be. Examination of arteriolar hyalinosis for IgAN patients under renal biopsy is essential to renal pathologists and clinicians.


Laparoscopic surgery in Trendelenburg position may impede mechanical ventilation (MV) due to positioning and high intra-abdominal pressure. We sought to identify the positive end-expiratory pressure (PEEP) levels necessary to counteract atelectasis formation (“Open-Lung-PEEP”) and to provide an equal balance between overdistension and alveolar collapse (“Best-Compromise-PEEP”).2O in supine position without capnoperitoneum and in Trendelenburg position with capnoperitoneum.In 30 patients undergoing laparoscopic gynecological surgery, relative overdistension and alveolar collapse were assessed with electrical impedance tomography (EIT) during a decremental PEEP trial ranging from 20 to 4 cmH2O with 8.7 (4.7–15.5)% of overdistension and 1.7 (0.4–2.2)% of collapse. Best-Compromise-PEEP was 8 (6.5–10) cmH2O with 4.2 (2.4–7.2)% of overdistension and 5.1 (3.9–6.5)% of collapse. In Trendelenburg position with capnoperitoneum, Open-Lung-PEEP was 18 (18–20) cmH 2 O (p < 0.0001 vs supine position) with 1.8 (0.5–3.9)% of overdistension and 0 (0–1.2)% of collapse and Best-Compromise-PEEP was 18 (16–20) cmH2O (p < 0.0001 vs supine position) with 1.5 (0.7–3.0)% of overdistension and 0.2 (0–2.7)% of collapse. Open-Lung-PEEP and Best-Compromise-PEEP were positively correlated with body mass index during MV in supine position but not in Trendelenburg position.In supine position, the median Open-Lung-PEEP was 12 (8–14) cmHThe PEEP levels required for preventing alveolar collapse and for balancing collapse and overdistension in Trendelenburg position with capnoperitoneum were significantly higher than those required for achieving the same goals in supine position without capnoperitoneum. Even with high PEEP levels, alveolar overdistension was negligible during MV in Trendelenburg position with capnoperitoneum.This study was prospectively registered at German Clinical Trials registry (DRKS00016974). Postoperative pulmonary complications (PPC) are common after general anesthesia with mechanical ventilation and may lead to prolonged hospitalisation, increased mortality and morbidity . Increas2), are not suitable for identifying regional phenoma such as atelectasis formation and pulmonary overdistension during mechanical ventilation. Electrical impendence tomography (EIT) is a noninvasive and radiation-free imaging modality based on the measurement of electrical potentials at the chest wall surface that can be used for direct monitoring of regional ventilation [2O [Unfortunately, standard monitoring parameters used during mechanical ventilation, such as peripheral oxygen saturation  as well as the PEEP level associated with equal amounts of overdistension and alveolar collapse (“Best Compromise PEEP”) in Trendelenburg position with capnoperitoneum and compare these results with the PEEP values needed to reach the same conditions in supine position without capnoperitoneum. Additionally, we assessed the hemodynamic changes associated with the abovementioned PEEP levels in comparison to baseline taken after induction of general anesthesia.2, presence of metallic implants or open injuries in the thoracic area, suspected or confirmed pregnancy, chronic pulmonary diseases or functional capacity below four metabolic equivalents of activity were excluded.We conducted a prospective clinical study in the Department of Gynecology and Obstetrics at the University Medical Center Schleswig–Holstein, Campus Kiel, Germany. The study received approval from the Ethics Committee of the Medical Faculty of Christian-Albrechts-University Kiel, Germany (D426/19) and was registered at German Clinical Trials registry on 14/03/2019 before including the first patient (DRKS00016974). 30 patients were enrolled between March 2019 and January 2020. Written informed consent was obtained from all study participants. We included patients aged more than 18 years with scheduled laparoscopic surgery in Trendelenburg position under general anesthesia. Patients with body mass index (BMI) above 35 kg/m2O in supine position and 8 cmH2O in Trendelenburg position with capnoperitoneum.General anesthesia was induced with Sufentanil (0.2–0.4 µg/kg), Propofol (1.5–2 mg/kg) and Rocuronium (0.6 mg/kg) and maintained with Sufentanil and Sevoflurane according to routine clinical practice in our institution. Fluid replacement was conducted with balanced crystalloid solution according to the treating anesthesiologist’s discretion . Patients were ventilated in a pressure-controlled mode with inspiratory pressure adjusted to achieve a tidal volume of 6–8 ml per kg ideal body weight and respiratory rate of 12–15/min, applying a PEEP of 5 cmHth−6th intercostal space, measured at the parasternal line. EIT measurements were conducted at a scan rate of 50 images per second using the PulmoVista 500 EIT device . Patients were intubated with an endotracheal tube of 7.5 mm internal diameter. EIT data were recorded under two conditions: 1) supine position without capnoperitoneum after induction of general anesthesia; 2) 30° Trendelenburg position after establishment of capnoperitoneum with a positive intra-abdominal pressure of 10–12 mmHg. In each of these two conditions, a decremental PEEP trial was conducted measuring alveolar collapse and overdistension. For this purpose, the PEEP level was first increased to 20 cmH2O in two or three one-minute steps. Subsequently, the set PEEP was decreased in one-minute steps of 2 cmH2O from 20 cmH2O to 4 cmH2O. During the collection of EIT data, vital parameters such as heart rate, arterial pressure and SpO2 were registered at baseline  and during each step of the PEEP trial. Additionally, the use of vasoactive medications was noted. After the end of the PEEP trial, mechanical ventilation was continued with clinically selected PEEP levels .Before induction of general anesthesia, a 16-electrode belt was attached to the patient’s chest circumference at the level of the 4rs) was calculated by dividing tidal volume by the corresponding driving pressure. For calculation of pixel Crs, local tidal volume was set equal to local tidal impedance change and driving pressure was estimated in pressure-controlled mode by subtracting PEEP from the set inspiratory pressure.EIT data was analysed offline using the EIT Data Analysis Tool 6.1 and PulmoVista PC Software 1.2 , and Microsoft Excel 2010 . The percentages of alveolar collapse and overdistension for each PEEP level were calculated using the algorithm proposed by Costa and colleagues  as impleOpen Lung PEEP was defined as the lowest PEEP level with less than or equal to 3% of alveolar collapse. Best Compromise PEEP was defined as the PEEP level associated with equal amounts of alveolar overdistension and alveolar collapse.The primary objective of the study was to identify the lowest PEEP level at which no alveolar collapse could be detected with EIT (“Open Lung PEEP”) in a 30° Trendelenburg position with capnoperitoneum. Further objectives were to identify the PEEP level with equal amounts of overdistension and alveolar collapse (“Best compromise PEEP”) in the 30° Trendelenburg position with existing capnoperitoneum, to identify Open Lung PEEP and Best Compromise PEEP in the horizontal supine position and to describe the PEEP level with the highest global respiratory system compliance. Furthermore, we sought to assess blood pressure, heart rate and oxygen saturation at the abovementioned PEEP levels and to investigate the correlation between Open Lung PEEP, Best Compromise PEEP and BMI.To obtain solid estimates for Open Lung PEEP and Best Compromise PEEP, adequately reflecting the clinical spectrum of weight characteristics in the range below BMI 35, we estimated that a sample size of 30 patients would be suitable. Statistical analysis was performed using GraphPad Prism 8.0 . All data sets were tested for normal distribution using D’Agostino Pearson omnibus normality test. Normally distributed variables are reported as mean with standard deviation, and not normally distributed variables as median with 25-75th percentile. Comparisons were performed using the paired t-test for normally distributed data and Wilcoxon matched pairs test for not normally distributed data. For comparison of repeated measurements, Friedman’s test with Dunn’s post hoc analysis was used. For comparison of categorical variables, Fisher exact test was applied.Twenty-seven patients with completed sets of EIT measurements were included in the final analysis. Patient flow is presented in the CONSORT diagram Fig. . 3 of in2O. In Trendelenburg position with capnoperitoneum, the same criterion was fulfilled under ventilation with a PEEP of 18 (18–20) cmH2O (p < 0.0001 vs supine position). Best Compromise PEEP was also reached with significantly higher PEEP values of 18 (16–20) cmH2O in Trendelenburg position with capnoperitoneum in comparison to 8 (6.5–10) cmH2O in supine position without capnoperitoneum (p < 0.0001). Figure rs was significantly lower in Trendelenburg position with capnoperitoneum in comparison to supine position without capnoperitoneum (p < 0.0001) for both Open Lung and Best Compromise PEEP  cmHEEP Fig. , while tp < 0.0001), while there was no significant difference between Open Lung PEEP and Best Compromise PEEP in Trendelenburg position with capnoperitoneum (p = 0.09).In supine position without capnoperitoneum, Open Lung PEEP was significantly higher than Best Compromise PEEP  but not in Trendelenburg position with capnoperitoneum . Similar results were also observed while ventilating with Best Compromise PEEP with lower heart rate and higher mean arterial blood pressure in Trendelenburg position compared to baseline and supine position (p < 0.0001). There was no significant difference in SpO2 between different positions and baseline. During PEEP trial, 3 of 27 patients needed vasoactive drugs in Trendelenburg position with capnoperitoneum and 14 of 27 patients in supine position (p = 0.0001).In the Trendelenburg position with capnoperitoneum, a lower heart rate of 60 (54–65) beats/min and higher mean arterial blood pressure of 89.5 (83–98.3) mmHg were documented with Open Lung PEEP in comparison to baseline (heart rate 68 (62–76) beats/min, mean arterial blood pressure 71.5 (64.8–81.8 mmHg) and supine position (heart rate 66 (59–75) beats/min, mean arterial blood pressure 73.0 (69.3–81.3) mmHg)  as well as the PEEP level required for equally balancing overdistension and collapse (“Best Compromise PEEP”) was significantly higher in Trendelenburg position with capnoperitoneum than in supine position. Both Open Lung PEEP and Best Compromise PEEP were positively correlated with BMI in supine position but not in Trendelenburg position with capnoperitoneum.2O is being used for ventilation both in open abdominal and in laparoscopic surgery [2O can be considered in Trendelenburg position with capnoperitoneum, without additional risk for hemodynamic compromise or lung overdistension. BMI could be used as additional criterium for choosing individualised PEEP value in supine position without capnoperitoneum using the regression equations presented in Fig. Currently, there is no international consensus on best PEEP level for prevention of regional hypoventilation and PPC. Frequently, a conservative PEEP level of less than 5 cmH surgery . Our fin2O). Due to the nature of the EIT analysis performed for the present study, “Best Compromise PEEP” and “Open Lung PEEP” in these patients would still be identified within the range of 20 – 4 cmH2O, precluding any possible differentiation in the higher PEEP range.Our seemingly contradictory finding of a positive correlation between the required PEEP value and BMI in the supine position but not in the Trendelenburg position with capnoperitoneum requires cautious interpretation. The mechanism by which a higher BMI leads to higher PEEP requirements is most likely closely related to intra-abdominal pressure, which is positively correlated to BMI during general anesthesia . In our 2O for counteraction of atelectasis formation in Trendelenburg position, although the PEEP trial was conducted before establishment of capnoperitoneum. This may explain the higher PEEP values for prevention of alveolar collapse and equally balancing overdistension and collapse reported in our study. Nevertheless, Pereira and colleagues were able to demonstrate the positive effect of EIT guided ventilation using postoperative CT scans showing less atelectasis formation compared to a control group ventilated with a fixed PEEP of 4 cmH2O.Our suggestion of using higher PEEP values for MV in Trendelenburg position is in line with the findings of another study on EIT-guided PEEP ventilation conducted by Pereira and colleagues . They de2O had a positive effect on regional ventilation distribution as compared to ventilation with PEEP of 5 cmH2O [2O.Investigating patients undergoing robot-assisted laparoscopic prostatectomy with capnoperitoneum in Trendelenburg position, Shono and colleagues reported that a higher PEEP level of 15 cmH 5 cmH2O . However2O for improvement of pulmonary ventilation [2O was argued to be not effective to compensate the effects of capnoperitoneum and Trendelenburg position, while a higher PEEP of 12 cmH2O was associated with increased intrapulmonary shunt and hemodynamic depression. In contrast, we observed preserved oxygenation and hemodynamic stability with the application of EIT-guided PEEP in Trendelenburg position with capnoperitoneum. All in all, high PEEP values during ventilation with Trendelenburg position with capnoperitoneum were well tolerated in our study.Another study on optimal PEEP level in Trendelenburg position with capnoperitoneum during laparoscopic surgery used ventilation parameters such as dynamic compliance, dead space to tidal volume ratio and intrapulmonary shunt ratio for evaluation of lung-protective ventilation and suggested ventilation with a moderate PEEP of 8 cmHtilation . In this2O for prevention of regional hypoventilation in patients undergoing laparascopic surgery in supine position while monitoring regional ventilation with EIT [Karsten and colleagues suggested ventilation with PEEP of 10 cmHwith EIT . These r2O during laparoscopic surgery in supine position for morbidly obese patients with BMI 49 ± 8 kg/m2 according to monitoring of regional ventilation with EIT, which was performed before the beginning of surgery and induction of capnoperitoneum [Erlandsson and co-workers suggested ventilation with PEEP 15 ± 1 cmHritoneum . These vritoneum .Some intra-individual variation of Open Lung and Best Compromise PEEP was observed in supine as well as in Trendelenburg position. This suggests that intraoperative monitoring of regional ventilation with EIT on an individual basis could contribute to improvement of lung-protective ventilation strategies. This may not be cost-effective for patients with general low risk of PPC and operation in supine position, but might be of benefit in high risk patients and body positions with increased intra-abdominal and transpulmonary pressure.Our study has some strengths. To our knowledge, it is the first study that assessed Open Lung PEEP and Best Compromise PEEP with EIT both after induction of general anesthesia and in Trendelenburg position with capnoperitoneum. Previous studies performing PEEP titration with EIT did so after induction of general anesthesia but not after establishment of capnoperitoneum , 21, 22.2 were excluded from the present study, our results cannot be extrapolated to morbidly obese patients.Our study has several limitations. As this study was conducted in the department of gynecology and obstetrics, only younger female patients were included, limiting the generalizability of our findings. Our findings should be examined in a mixed gender cohort, to eliminate possibility of gender specific findings. Moreover, as patients with BMI > 35 kg/mIn general, the perioperative risk of pulmonary complications is low in a relatively young and healthy patient population as studied in our manuscript. Therefore, it is unclear whether individualized adjustment of PEEP will have any clinically relevant effects on patient-centered outcomes in this patient population.The effects of Trendelenburg position and capnoperitoneum were not investigated separately. The sample size was relatively small and was further reduced by a drop-out rate of 10%. However, given the relatively homogeneous patient population, we were still able to obtain robust values for median and interquartile ranges of Open Lung and Best Compromise PEEP. Post hoc power calculations using the actually achieved effect sizes revealed a statistical power of > 0.95 for detecting a difference in PEEP level between supine position and Trendelenburg position.Our study was neither designed nor powered to investigate the influence of EIT-guided PEEP postoperative pulmonary complications. Nevertheless, none of the patients investigated in this study had additional oxygen requirements or needed invasive or non-invasive mechanical ventilation after the end of surgery.A randomized study to assess the effect of EIT-guided PEEP settings during Trendelenburg position with capnoperitoneum on the occurrence of PPCs would be desirable to confirm our results and further specify the optimal PEEP level for prevention of PPCs after mechanical ventilation during general anesthesia.2O do not lead to relevant pulmonary overdistension and could be considered for preventing regional pulmonary hypoventilation and alveolar collapse. In supine position without capnoperitoneum, significantly lower PEEP levels that could be adjusted according to BMI may be suitable for achieving a balanced compromise between alveolar collapse and overdistension.During mechanical ventilation in Trendelenburg position with capnoperitoneum, higher PEEP levels of 16 to 20 cmH