The morphology of the contact surface between cast-in-place engineering structures and soil is generally random. Previous research focusing on the shear mechanical properties of soil–concrete interfaces has predominantly concentrated on the role of interface roughness by constructing regular concrete surface types, largely neglecting the potential impact of the roughness morphology . In this study, concrete blocks with the same interface roughness and different roughness morphologies were constructed based on the sand-cone method, including random rough surface, triangular groove surface, rectangular groove surface, trapezoid groove surface, and semicircular groove surface. A series of direct shear tests were conducted on the rough and smooth sand–concrete interfaces, as well as on natural sand. Through these tests, we examined the shear mechanical behavior and strength of the sand–concrete interfaces, and analyzed the underlying shear mechanisms. The results showed that: (i) the interface morphology had little effect on the variation in the shear stress–displacement curve of sand–concrete interfaces, and it had a significant influence on the shear strength of the interfaces; (ii) under the same normal stress, the shear strength of the sand–concrete interfaces with a random rough surface was the greatest, followed by the triangular groove surface, while the shear strength of the rectangular groove surface proved the lowest; (iii) the shear strength of the sand–concrete interfaces with the same roughness was affected by the size of the contact area between the concrete plane and the sand, that is, a larger contact area correlated with a decrease in shear strength. It can be concluded that the shear strength value of a sand–concrete surface with the triangular groove is the closest to the shear strength of a random rough interface. By gaining a deep understanding of the effects of different contact surface morphologies on shear strength and shear behavior, significant insights can be provided for optimizing engineering design and enhancing engineering performance. Soil–structure interaction features are among the keen research topics in geotechnical engineering, with significant achievements in investigating the mechanical features of soil–structure interfaces, such as studying the damage constitutive model of the soil–structure interface  and the A deeper understanding and analysis of the effects of different morphology characteristics on the shear mechanical properties of interfaces can help us gain better insights into the properties of real engineering interfaces and provide more rational guidance and optimization strategies for engineering design and construction. Therefore, investigating the impact of interface morphology characteristics on the shear mechanical properties of interfaces holds significant theoretical and practical significance. In the present work, we designed five distinct rough-morphology concrete surfaces using the sand-cone method, each with the same degree of roughness. The designs comprised random surfaces, rectangular groove surfaces, triangular groove surfaces, semicircular groove surfaces, and trapezoidal groove surfaces. A series of direct shear tests on sand–concrete interfaces at three relative densities were conducted. The shear stress displacements on contact surfaces with varying roughness morphologies and relative densities were analyzed. Additionally, the influences of different roughness morphologies on the shear strength of the soil–concrete interface under the three relative densities were investigated. The shear failure mechanism of different roughness morphology concrete surfaces was revealed.The ZJ strain controlled direct shear apparatus produced by Nanjing Soil Instrument Factory Co., Ltd. , was adopted for the direct shear tests . NaturalBased on the slurry intrusion test in sand, concrete blocks with random roughness surfaces were obtained . The rouDr = 30%, 50%, 70%, respectively) were designed for the direct shear test, and each relative density corresponded to 7 sets of shear tests, as shown in σ, applied in the shear test were 50 kPa, 100 kPa, 200 kPa, and 300 kPa. Thus, 28 sand samples needed to be made for each relative density. The main steps in the testing process were as follows:Three relative densities of sand samples ((i) A preliminary verification as to whether the direct shear instrument and data acquisition system could operate normally was carried out. (ii) The circular container of the lower box of the shear box was removed, and the prepared concrete block was placed into the lower box of the shear box, as shown in In the cases of random rough concrete surfaces, the variation law of shear stress–displacement curves was found not to have been affected by the relative density and normal stress values, as evidenced by the virtual absence of any softening trend, as shown in Previous studies had shown that while the shear strength of interfaces rose with the increase in interface roughness, this augmentation remained less than the internal shear strength of sand ,25, whilThe shear mechanical properties of the soil–structure contact surface are a keen research topic in geotechnical engineering, and the previous studies have mainly focused on the effect of concrete surface roughness on interface shear mechanical properties by constructing concrete blocks with regular rough surfaces. However, the morphology of the contact surface between the cast-in-place concrete structure and the surrounding soil in engineering is random, and the morphology of regular rough surfaces constructed in different studies also varies. Therefore, it was necessary to study the potential impact of concrete surfaces with various roughness morphologies on interface mechanical properties. In this study, based on the roughness value of a random concrete surface, four regular types of concrete specimen contact surfaces with the same roughness but different groove morphologies were produced, as well as a smooth concrete surface sample. A series of direct shear tests were performed. The relationship between shear stress and shear displacement during the shear process was analyzed. The effects of the interface morphology on the shear strength of the sand–concrete interfaces were investigated, and the main conclusions were as follows: (1) The rough morphology of concrete surface had little effect on the variations in the shear stress–displacement curves of the sand–concrete interfaces. (2) The shear strength of the sand–concrete interfaces varied with the differences in concrete surface morphology, and the shear strength of the concrete surface with random rough morphology was proved to be the greatest. (3) Among the concrete surfaces with regular rough morphologies, the shear strength of the triangular groove concrete surface was the highest, which was closest to the shear strength of the random rough concrete surface, while the shear strength of the rectangular groove concrete surface proved to be the lowest. (4) The shear strength of sand–concrete interfaces with the same roughness value was affected by the size of the contact area between the concrete plane and sand. The greater the contact area between the concrete plane and the sand, the lower the shear strength of the sand–concrete interface was found to be.There are currently numerous evaluation methods of roughness, among which the sand-cone evaluation method was selected for this study. The average depth of the random concrete surface was the same as that of the four regular concrete surfaces . The results in the present research showed that different contact surface types with the same roughness value yielded different shear strength values. It was thus inferred that solely evaluating concrete surface roughness on the basis of its average depth may not reflect all of the properties of the concrete surface in question. Accordingly, the present authors concluded that in the future, rough concrete surfaces should be constructed on the basis of other roughness evaluation methods to further investigate the properties of the sand–concrete interfaces. In addition, further research is also needed on roughness evaluation methods, so that the research results obtained can be better applied to engineering practice.


This easy-to-control variation yielded a gradually less electron-rich cavity in going from zero to three quinone units, as shown from the strength of host–guest interactions with silver ions. Such macrocycle-Ag2 complexes themselves were shown to display an unprecedented, sharp distinction between terminal alkynes, which strongly bound to such complexes, and internal alkynes, internal alkenes and terminal alkenes, which do hardly bind.We developed an improved, robust synthesis of a series of pillar[ This isn]arenes are macrocyclic hosts with a cylindrical, pillar-like rigid structure that have been investigated extensively with respect to their host–guest chemistry [n]arenes are composed of n hydroquinone monomers covalently connected with methylene bridges at the 2,5-para positions; at present, pillar[5]arenes (P5s) and pillar[6]arenes (P6s) have attracted most attention, due to their relative ease of formation [5]arenes display a 5-fold symmetry and a cavity diameter of about 5 Å, i.e., similar to that of α-cyclodextrin [6]arenes have a hexagon-like structure with a cavity size of, ca. 6.7 Å [n]arenes are electron-rich due to the electron-donating hydroxyl or alkoxy groups on both rims. This particular feature has driven most of the aforementioned rich host–guest chemistry, and is, for example, used in a wide variety of separation and extraction applications quinones (n = 3–7) arene (MeP5) was synthesized on a multigram scale via a literature method (see Supporting Information) [6]arene (EtP6), as needed for the subsequent synthesis and follow-up studies of pillar[6]arene[1]quinone, pillar[6]arene[2]quinone and pillar[6]arene[3]quinone . For EtP6, a variety of synthetic procedures have also been reported, e.g., using a catalyst template [EtP6 using [ChCl][FeCl3]2 as the catalyst template  in DCM/THF 1:1 solvent mixtures  after the addition of CF3COOAg .Next, we investigated the complexation of all six compounds (a)–(f) see  with CF3research , and basoacetate . In contCF3COOAg . Therefo1H NMR titration experiments of MeP5, EtP5, EtP6 and P6Q1 allowed us to calculate the binding strengths. Different protons show different chemical shifts, as the guest molecules in the cavity have different effects on them. Protons on the EtP6 benzene ring displayed the largest chemical shift (∆δ up to 0.005 ppm), while the largest chemical shift for P6Q1 was, ca. 0.002 ppm. We then used the observed ∆δ values of the peaks for the protons in the benzene ring to calculate the binding constants for EtP6 and P6Q1 , we calculated the binding constant by taking the combination of two CF3COOAg as one guest for the calculation, so the data were set into a 1:1 complex calculation system, in which the guest concentration taken into account in the binding constant calculations was half of that of the experimental CF3COOAg concentration + peak gives the same trend in binding strength as that of the NMR titration experiment, but due to the sensitivity of the ESI-MS experiment also, data for the P6Q2 and P6Q3 hosts could be obtained, EtP5 ≈ EtP6 > MeP5 > P6Q1 > P6Q2 > P6Q3, using NL values of 1.58, 1.55, 1.43, 1.05, 0.86 and 0.15 × 109, respectively  and compared the ratios of the NL values of different [host + CF3COOAg + Ag]+ peaks in ESI-MS experiments. The resulting ratios are shown in EtP5 ≈ EtP6 > MeP5 > P6Q1 > P6Q2 > P6Q3. The occurrence of this trend thus strongly supports the hypothesis that the binding strength toward the silver ions can be precisely tuned via the number of quinone moieties in the P6Qn host.Since both the NMR-based K values and the ESI-MS-based NL values indicate Ag3COOAg entities is strongly preferred over binding only one; while the binding of one CF3COOAg follows roughly the same trend over the various hosts as two CF3COOAg units, the values are much smaller: from +14 to +21 kcal/mol for one CF3COOAg, to −23 to −43 kcal/mol for two CF3COOAg. These data confirm the earlier observation that only complexes with two CF3COOAg units are observed, both for P5 by Huang et al.   in water was added. The resulting red-colored mixture was stirred at room temperature for 30 min to 3 h, washed with water and concentrated under reduced pressure. The crude product was purified via column chromatography to afford P6Q1 , P6Q2  or P6Q3  [n]arenes and CF3COOAg were elucidated using the well-established methodology of Job Plots. To ascertain these ratios, the combined concentration of the host and guest species ([host + guest]) was held constant at 2.0 mM. The host-to-guest ratio was then systematically varied to create a spectrum of ratios, ranging from 9:1 to 1:9. Importantly, the silver ion-pair was treated as a singular guest entity, effectively halving the [Ag+] concentration, as each ‘guest’ now comprised two Ag+ ions. This reevaluation allowed us to explore the range from 0.2 mM of the Ag22+ guest (resulting from 0.4 mM of Ag+ in a 1.8 mM pillararene solution) to 1.8 mM of the Ag22+ guest (achieved by introducing 3.6 mM of Ag+ into a 0.2 mM pillararene solution).The stoichiometric ratios underlying the complexation between pillar+ peaks. This analysis enabled the exploration of relative ratios between different peak configurations, thereby offering information regarding the relative binding strengths.In the competitive ESI-MS competition experiments, two distinct host species were combined with two equivalents of the guest, CF3COOAg with pillararenes is defined as3COOAg, Ep is the free energy of pillararenes and 3COOAg.All DFT calculations were carried out using the Gaussian 16 suite of computational programs ). The ge6]arene hosts and silver ions are strong, and can be tuned in detail via partial oxidation of the pillararene aromatic rings. Such host–guest, Ag+∙∙∙π complexes themselves give rise to the exclusive and unprecedented in-cavity complexation of terminal alkynes over internal alkynes, internal alkenes and terminal alkenes. This sharp distinction provides a novel basis for specific separations of terminal alkynes from other compounds and will be studied further in our laboratories.A series of NMR titration and ESI-MS experiments in combination with high-end DFT data show that the host–guest interactions between pillar[


Environmental tobacco smoke (ETS) is known to cause lung inflammatory and injurious responses. Smoke exposure is associated with the pathobiology related to lung fibrosis, whereas the mechanism by which ETS exposure augments lung fibrogenesis is unclear. We hypothesized that ETS exposure could exacerbate fibrotic responses via collagen dynamic dysregulation and complement activation. C57BL/6J and p16–3MR mice were exposed to ETS followed by bleomycin administration. ETS exposure exacerbated bleomycin-induced collagen and lysyl oxidase overexpression in the fibrotic lesion. ETS exposure also led to augmented bleomycin-induced upregulation of C3 and C3AR, which are pro-fibrotic markers. Moreover, overexpressed collagens and C3 levels were highly significant in males than females. The old mice (17 months old) were exposed to ETS and treated with bleomycin to induce fibrogenesis, since fibrogenesis is an aging-associated disease. Fewer gene and protein dysregulations trends were identified between ETS exposure with the bleomycin group and the bleomycin alone group in old mice. Based on our findings, we suggested that ETS exposure increases the risk of developing severe lung fibrotic responses via collagen overexpression and lysyl oxidase-mediated collagen stabilization in the fibrotic lesion. ETS exposure also potentially affected the complement system activation induced by bleomycin. Further, male mice were more susceptible than females during fibrogenesis exacerbation. Tobacco smoke is not only a global public health concern , but alsTobacco smoke exposure can be considered active smoking exposure or passive smoking exposure. ETS is a mixture of side-stream cigarette smoke (CS) and main-stream CS exhaled by active smokers. It is well known that both CS and ETS could induce a significant inflammatory response, abnormal respiratory syndromes, and CS exposure aggravates fibrotic progression –9. CS exECM is formed with extracellular proteins as basal material to support cells in the three-dimension microenvironment. The dynamic production and degradation of ECM are involved in multiple disease mechanisms and overexpression of ECM is commonly observed in fibrotic progression. The major ECM proteins are collagens, fibronectin, and elastin. Among collagens, types 1 and 3 are the major fibrous collagens to form ECM in alveoli and type 4 majorly exists in the basal membrane for supporting epithelium , 17. EnzHere, we hypothesized that sub-chronic exposure to ETS augments fibrogenesis induced by bleomycin (BLM) administration via collagen dynamic dysregulation. C57BL/6J and p16–3MR mice were exposed to ETS for 30 days and followed by bleomycin administration for 14 days. The duration of ETS exposure and post-bleomycin administration is based on previous publications to avoid emphysema development induced by CS exposure and recovery of scaring injury after 21 days post-bleomycin , 23–29. C57BL/6 and p16–3MR  mice were housed in a 12-hour-light/12-hour-dark cycle in the inhalation core facility at the University of Rochester. First, both young and old groups of each strain were exposed to ETS for a total of 30 days, 5 days/week, via the Teague machine with 100 total particular matter . Air-exposed control groups of mice were housed separately. After 6 weeks of exposure, bleomycin (BLM) (1.5U/kg) or PBS (50μl) were oropharyngeal administered. Mice were sacrificed after 14 days. During the sacrifice, pentobarbital  injection) was given, lung mechanics were measured during the sacrifice, and lung lobes were then snap-frozen for protein/RNA analysis. Lung lobes inflated with 1% low-melting agarose were used for histological analysis. Ganciclovir  was given to p16–3MR mice via intraperitoneal (i.p.) injection of every alternative day for a total of 8 times post-BLM.Young (4–5 months) and old (15–17 months) groups of 2O positive. The impedance of the equipment and tracheal tube was removed with appropriate calibration. These measurements were repeated 3 times for each animal  were determined with the FlexiVent device . Mice were first anesthetized with an intraperitoneal injection of sodium pentobarbital (100 mg/kg). After a tracheotomy, an 18-gauge cannula was introduced into the exposed trachea and attached to a computer-controlled rodent ventilator . Following the initial ventilation, compliance, resistance, and elastance were measured by a computer-generated program at 3cmHh animal .g for 15 min at 4°C, after that, the aqueous phase was moved to new RNase-free tubes and mixed with isopropanol for 2 hr at −20°C. The mixtures were spun down at 20,000g for 15 min at 4°C, and the supernatant was removed. RNA pellets were washed with 75% EtOH and then centrifuged at 20,000g for 15 min at 4°C. Discarded the EtOH and re-suspended RNA with RNase-free water. All of the RNA samples were cleaned up by RNeasy Plus Mini Kit following the manufacturer’s protocol. RNA samples were kept at − 80°C until analysis. RNA samples were quantified using a NanoDrop spectrophotometer . A total of 100ng of RNA samples were used for NanoString analysis. We have used the nCounter Fibrosis Panel  in this study. Briefly, the hybridization reaction mix was prepared by RNA samples, reporter codeset, and capture codeset. The mixtures were incubated at 65°C for 16 hrs and stored at 4°C until further profiling. NanoString running cartridge was used to load all the sample mixtures and performed gene expression profiling via nCounter SPRINT Profiler . After profiling, gene expression results were analyzed by nSolver 4.0 software, and reported as normalized count for data presentation and statistical analysis. Raw RCC files were uploaded to ROSALIND for advanced analysis and generated the signaling pathway scores as directed enrichment scores with at least 10% change, and p ≤ 0.05 were considered as significantly altered.Snap-frozen lungs were mechanically homogenized with QIAzol reagent, then each sample was mixed with chloroform and vortexed vigorously for 10 sec. The samples were centrifuged at 20,000Snap-frozen lungs were mechanically homogenized in RIPA buffer  with protease inhibitor . Protein concentrations were quantified by Pierce BCA Protein Assay . Total 30 μg of protein was prepared for each lane for electrophoresis via precast gels . After electrophoresis, proteins were transferred onto a nitrocellulose membrane  through a semi-dry system. The membranes were washed with tris-buffered saline containing 0.1% Tween 20 (TBS-T) for 15 min, then blocked with 5% bovine serum albumin (BSA) for 1 hr at room temperature (RT). Next, the membranes were probed with primary antibodies overnight at 4°C: anti-Collagen 1α1 ; anti-Collagen 4α1 ; anti-LOX ; anti-fibronectin ; anti-MMP2 ; anti-C3 , and the appropriate secondary antibody  for 1 hr and 30 min at RT. After each antibody incubation, the membrane was washed 3 times in TBS-T at RT, 15 min per wash. The luminescence signals were developed using a chemiluminescence substrate . The membranes exposure and band intensities were detected via the Bio-Rad ChemiDoc MP imaging system . Protein quantification was done by densitometry, and β-ACTIN  was used as housekeeping control. Densitometry analysis was performed by Image Lab software  and data was presented as fold change compared to the PBS group.The H&E staining has been performed as previously described . BrieflyGomori’s trichrome staining has been used to quantify collagen deposition in the lungs from different experimental groups. Lung sections (5 μm) were deparaffinized, rehydrated, and finally rinsed with tap water. Sections were then subjected to Gomori’s trichrome staining . In brief, sections were placed in Bouin’s solution at 56°C for 1h, and then washed with water until the yellow color is removed. Next, sections were immersed in Working Weigert’s Iron Hematoxylin Stain for 10 min and washed with water for 10 min. After, the sections were soaked in Trichrome Stain for 15 min and immediately after in acetic acid 1% for 1 minute. The sections were rinsed with water for 30 sec and dehydrated. Lastly, the slides were mounted by Permount  for microscopy observation. For each sample, 10 pictures (20x magnification) have been taken by Nikon microscopy (Eclipse Ni-U) and the amount of collagen has been quantified by Colour Deconvolution via FIJI ImageJ.The staining protocol has been performed as previously described . Lung seAll the data were analyzed through one-way ANOVA with Šidák correction using GraphPad Prism software . Results were presented as mean ± SEM, and p < 0.05 was considered significant.The animal experiments performed in this study was following the standards from the United States Animal Welfare Act, NIH, and protocol approved by the Animal Research Committee of the University of Rochester.Figure S1). There was no difference in BLM-induced lung injury between either young and old mice, or male or female mice . We also performed the Trichrome staining to determine dysregulated collagen deposition among different conditions. BLM (Air + BLM and ETS + BLM)-induced significant collagen deposition compared to Air + PBS or ETS + PBS groups, and ETS exposure did not affect the collagen deposition in the injured areas from either young or old mice .Mice were exposed to ETS for 30 days and then BLM was administered oropharyngeal to evaluate how ETS exposure contributes to BLM-induced lung injury . Signifiold mice . ETS expold mice . The altLung mechanics  were measured on day 14 post-BLM administration. There was no significant alteration in resistance, while significantly decreased elastance and increased compliance was found in ETS + PBS compared to the Air + PBS group . BLM Ai. SimilarCOL1A1, COL1A2, COL4A1, COL4A2, and COL5A3. Gene expression levels of COL4A1 and COL5A3 showed a significant increase in ETS + BLM compared to Air + BLM, while COL1A1, COL1A2, and COL4A2 showed non-significant increase trends . The MMPs inhibitor, TIMP1, showed increased gene expression levels in Air + BLM and ETS + BLM groups compared to either Air + PBS or ETS + PBS groups  or ETS (ETS + BLM) exposure compared to the respective PBS groups (Air + PBS and ETS + PBS) .To further determine the role of ETS exposure in BLM-induced lung injury and dysregulated repair in young mice, RNA was isolated from lung homogenates prepared from young mice, and fibrotic genes were measured via NanoString profiling . BLM adme trends . Other tM groups . A similS groups . One of Based on the dysregulated gene list, pathway analysis was performed to determine how ETS affects the lung injury induced by BLM . BLM admFigure S4). The significantly increased protein level of activated MMP2 was found in females after ETS exposure and BLM compared to Air + PBS, while no difference between Air + BLM and ETS + BLM groups was found in males . To determine the protein abundance in the lesion area, IHC staining was performed. The protein expressions of COL1A1, COL4A1, and LOX were increased after BLM administration following either air (Air + BLM) or ETS (ETS + BLM) exposure compared to Air + PBS or ETS + PBS respectively. The protein level of LOX in the lesion areas was significantly increased in the ETS + BLM group compared to Air + BLM, while COL4A1 and COL1A1 showed non-significant increased protein expression in the injury area of the ETS + BLM group compared to Air + BLM group  were upregulated after BLM administration either with air (Air + BLM) or ETS exposure (ETS + BLM)  were tested as well. Increased CDKN1A and CDKN2B gene expression levels were found in Air + BLM and ETS + BLM groups compared to Air + PBS group, and ETS exposure with BLM (ETS + BLM) showed further increased gene expression of CDKN1A when compared to Air + BLM group without significant difference  or ETS (ETS + BLM) exposure  was slightly activated after BLM administration (BLM + PBS), and ETS exposure followed by BLM (ETS + BLM) showed a further increase . More imS + BLM) , while ofference . Other Sexposure .C3AR1 between the ETS + BLM and Air + BLM groups. Then, we tested the protein abundance of C3 to confirm the activation of the complement system. A significant increase of C3 alpha chain (C3A) was seen after ETS exposure and BLM compared to Air + PBS or Air + BLM groups, and there is no change between Air + PBS and Air + BLM groups . A significantly increased protein level of C3A was identified in males, and no alternation of C3A protein abundance was identified in females . Increased protein abundances of C3 alpha chain fragment (C3B) and C3 alpha chain fragment 2 (C3C) were also significantly upregulated in Air + BLM and ETS + BLM groups compared to the Air + PBS group. ETS + BLM group showed further increased protein abundance of C3B and C3C compared to Air + BLM, but without significant difference . Interestingly, the gene expression of C3 showed a higher basal level in the old mice than in the young mice. However, a lower basal transcript level was detected in C3AR1 gene expression . Despite the different basal levels of gene abundance of C3 and C3AR1, the dysregulated trend under ETS exposure post-BLM showed similarly between young and old mice .We found a significant difference in the gene expression of fference , and simCOL1A1, COL1A2, COL3A1, COL4A1, COL4A2, COL5A1, and COL5A3) were observed after BLM administration, and ETS exposure did not affect the transcript levels  and MMP2 showed upregulation in both Air + BLM and ETS + BLM groups compared to Air + PBS, whereas no alternation between Air + BLM and ETS + BLM groups  were upregulated after BLM administration regardless followed by Air (Air + BLM) or ETS (ETS + BLM) exposure . In old mice, both TIMP1 and TIMP2 gene levels were upregulated after BLM administration, and there was no significant difference between Air + BLM and ETS + BLM groups . The gene levels of TGFβ and its receptors  showed upregulation in Air + BLM and ETS + BLM group compared to either Air + PBS or ETS + PBS groups, and there was no significant difference between Air + BLM and ETS + BLM groups .To determine the impact of ETS exposure on lung fibrotic progression after chronological aging, 18 months old mice were used in this study. Increased gene expression levels of collagens . Most of the collagens gene dysregulation trends are similar between young and old mice under different conditions, while transcript levels of COL3A1, COL5A1, and COL14A1, showed further upregulation post-BLM in old mice compared to young mice. However, the basal gene expression of COL4A1 showed lower in old mice despite the dysregulation trends are similar between young and old mice . Unlike the gene expression of collagens, the transcript levels of lysyl oxidases and proteases showed decreased basal levels in older mice, although the dysregulation trend is comparable between young and old mice .Besides the gene expression measurement, protein abundances were also tested from lung homogenates . The incSince the ETS exposure augmented premature senescence induced by BLM, we also utilized p16–3MR mice which can remove the p16 overexpressing senescent cells with the help of ganciclovir GCV). GCV was given to individual mice during BLM administration after ETS exposure on alternative days . After dV. GCV waExposure to ETS, a mixture of tobacco smoke from the side-stream of cigarettes and the exhaled smoke from active smokers, has shown correlations with asthma and COPD development. However, no clinical evidence showed how ETS exposure contributes to fibrogenesis development , 32. RecCOL4A1 and COL5A1 recorded significantly increased RNA expression levels in ETS+BLM compared to the Air+ETS group, and non-significant increased COL1A1, COL1A2, and COL4A2 were found as well in the ETS+BLM group compared to Air+BLM. Besides, our study also showed further increased protein levels of COL1A1 in males rather than females in the ETS+BLM group compared to the Air+BLM group, which partially agreed that males are more vulnerable during fibrogenesis , and inflammatory cell-flux infiltration . In thisogenesis . A recenogenesis . The samogenesis . Anotherogenesis . Our stuLOX, LOXL1, and LOXL2 showed no significant difference between the ETS+BLM and Air+BLM groups, a significantly increased protein abundance of LOX was found especially in the fibrotic lesion area in the ETS+BLM group compared to Air+BLM group. More importantly, ETS exposure impacts bleomycin-induced upregulated protein abundance of active LOX in males only, whereas no difference is identified in females. Our results of lysyl oxidase differentiated expression in different groups in a sex-dependent manner contribute to the statement that males showed worse lung injury during fibrotic progress than females . A non-significant further increased CDKN1A was found in ETS+BLM compared to the Air+BLM group. The protein encoded by CDKN1A is p21, which can cause cell cycle arrest by inhibiting CDK2 and CDK4 (CDKN1A in the alveolar epithelium (CDKN1A upregulation. Another SASP gene, SERPINE1, showed upregulation after bleomycin administration and a further upward trend was found in the ETS+BLM group compared to the Air+BLM group. It has shown that the upregulation of SERPINE1 induced the phosphorylation of p53 and increased the protein level of p21, which activated p53-p21-Rb-associated cell cycle repression (SERPINE1 by ETS could augment the senescence process induced by bleomycin.Our gene expression results and pathway analysis show further activation of ECM degradation, collagen biosynthesis and modification, and ECM synthesis. We, therefore, focused on MMPs and lysyl oxidases, which are the enzymes responsible for ECM remodeling. Increased protein abundance of MMP2 was found either in lung fibrosis animal models or fibrotic clinical samples . Our res females . Other t females , 42, we  females . After band CDK4 . Either ithelium . Our respression . Our resC3AR after ETS exposure. Longer exposure duration and higher doses of ETS should be considered to activate C3 and C3AR. Another study showed that an increased protein abundance of C3A was found after bleomycin treatment (Besides the CDKN family and SERPINE1, the transcriptomic panel also showed irregular activation of the complement system, which plays an important role during cellular senescence and aging . It has reatment . Increasreatment . In the reatment . Our resAs mentioned before, fibrogenesis is an aging disease usually occurring in elders. However, based on our normalized gene expressions of collagen, lysyl oxidase, and proteases, older mice did not show significant dysregulation levels than younger mice in most fibrotic markers. It has been reported that the bleomycin-induced fibrosis model serves better in the old mice, in which the chronological aging could delay fibroblast-dominated repair capacity, hence inducing persistent scarring and ECM deposition, whereas young mice showed recovery in lung scarring , 51. BleIn conclusion, we showed that ETS exposure exacerbated bleomycin-induced overexpression of collagen subtypes, lysyl oxidase, and C3A-receptor signaling. Removing p16 high-expressed cells did not help alleviate the lung injury induced by bleomycin. More importantly, our results showed that male mice were more susceptible than females during fibrogenesis exacerbation. Our study provided potential signaling that ETS exposure not only activated the TGFβ/SMAD2 pathway but also C3A-receptor signaling, which might be the potential reason for exacerbated abnormal collagens dynamics during fibrogenesis.


The developed method is the first MEKC for concurrent determination of EMP, LIN and MET with high separation efficiency, low solvent consumption and regard as an easy green and white analytical tool. Moreover, Greenness and whiteness assessment were done via the most widely used Analytical Eco-Scale, the innovative AGREE tool and the RGB 12 algorithm.The work introduces green and white sustainable micellar electrokinetic chromatography (MEKC) procedure that could analyze therapeutically related drugs, empagliflozin (EMP), linagliptin (LIN) and metformin (MET) which are antidiabetic drugs with different mechanism of action, in their different pharmaceutical combinations. The method not only comply with the green analytical concepts, but also it is in line with sustainable analytical concepts as it is economic by applying the same operating conditions to analyze different pharmaceuticals in quality control (QC) labs which is a crucial step in QC labs and research centers to save time, effort, and money. Moreover, the method functionality regarding its scope with its achieved levels of accuracy, precision, low detection, and quantitation limits is tested using white assessment tool and compared with reported methods. The proposed MEKC coupled with a diode array detector (DAD) has been developed and validated for micro estimation of EMP and LIN in their low critical concentrations with MET in a ratio of  and . Separation was achieved within 6 min using fused silica capillary (40 cm × 50 µm id) using 20 mM Tris buffer (pH 10) in presence of 50 mM sodium dodecyl sulphate and 10% v/v methanol. The concentration ranges of the studied anti-diabetic drugs were 10–500, 10–100 and 2.5–100 µg. mLThe online version contains supplementary material available at 10.1186/s13065-023-00997-0. The greenness and whiteness of analytical methodology is vital and significant with the aim of improving the health hazards and environmental effect. It is important not only to comply with the green analytical concepts, but also should be in line with sustainable analytical concepts .®) has been accepted officially by FDA (January 2020) which merged into one fixed dose tablet three classes of commonly used anti-diabetic medications working together to reduce blood glucose level in type 2 diabetes mellitus(T2DM). These drugs are characterized by diverse mechanism of action, with both short and long-term glycemic control, achieving low hypoglycemic risk in order to accomplish effectiveness and tolerability for different individuals [A novel triple combo pill (Trijardy XR January 020 whichMetformin (MET) Fig. Literature review showed that limited methods of analysis were established for simultaneous determination of the triple combination such as HPLC –8, HPTLCTo our knowledge, MEKC method has not been applied yet for the determination of the combo pill triple combination therapy and their related binary combinations of EMP/MET or LIN/EMP except for LIN/MET mixture, one CE method was found in literature .The current work represents green and white MEKC procedure for analyzing the three therapeutically related anti-diabetic drugs (EMP/LIN/MET) in their different binary and ternary pharmaceutical combinations. The method not only comply with the green analytical concepts, but also it is in line with sustainable analytical concepts as it is economic by applying one analytical technique with one operating condition which saves time, money and efforts in QC laboratories. Moreover, the two green chemistry metrics along with the white RGB 12 model revealed that our proposed MEKC is the most green and white compared to other published methods.Agilent CE instruments 7100 series from Agilent Technologies, Waldbronn, Germany, equipped with a DAD was used. It was also equipped with a data handling system comprising a computer loaded with software “Agilent Openlab CDS ChemStation”. The used capillary obtained from Agilent Technologies, Inc. It was a deactivated fused silica capillary that had the following dimensions: 48.5 cm total length, 40 cm effective length, and 50 µm id. For the purpose of accurate weighing of buffer and surfactant, a 3-digits Sartorius BL 310 balance was used, while the analytical balance used for accurate weighing of the drugs and the dosage forms a 4-digit Kern AEJ 220-4 M balance (Germany) was used. In addition, a Jenway 3305 pH Meter was used for pH measurements.Pharco Pharmaceuticals, Co.  kindly offered MET and LIN, while EMP was supplied by Pharaonia pharmaceuticals . HPLC grade methanol , Tris buffer Extra pure and sodium phosphate dibasic , sodium hydroxide, hydrochloric acid, sodium dodecyl sulphate  and deionized water were used.® and Cidophage® labelled to contain 5 mg, 25 mg and 1000 mg per tablet, respectively.Empagliform® tablets labeled to contain 12.5 mg EMP/ 500 mg MET, Gliptalina® tablets labeled to contain 2.5 mg LIN/ 500 mg MET and Glyxambi® tablets labeled to contain 10 mg EMP/ 5 mg LIN were purchased from the Egyptian market. Trijardy XR® tablets labeled to contain 25 mg EMP/ 5 mg LIN/ 1000 mg MET, due to the unavailability in the local market, laboratory-prepared tablets having the same drugs’ amount were prepared. This was done by using the individual dosage forms of each drug alone Trajenta®, EmpaglimaxTris buffer  was prepared by dissolving 0.24 g of Tris buffer extra pure in a 100-mL volumetric flask using deionized water, pH was adjusted to 10 with 0.5 M hydrochloric acid. To prepare 50 mM of sodium dodecyl sulphate (SDS) in Tris buffer, the following was done: a weight of 1.44 g SDS was added to the prepared buffer, followed by 15 min sonication till reach the complete dissolution. The finally used background electrolyte (BGE) is 20 mM Tris buffer adjusted to pH 10 with 0.5 M hydrochloric acid. This buffer contains 50 mM sodium dodecyl sulphate (SDS) and 10%v/v methanol.The fused silica capillary (40 cm × 50 µm id) was conditioned at the start of each working day first for 10 min using 0.5 M NaOH followed by another 10 min with deionized water, then for 5 min using NaOH (0.1 M), again deionized water was utilized for 5 min and the last 10 min of conditioning Tris buffer solution was used. For the in-between injections, two min flushing was done using the selected Tris buffer. The capillary was kept at 25 °C. The BGE used was a 20 mM Tris buffer (pH 10) contains 50 mM SDS and 10%v/v methanol, prepared as described under Sect. "−1 of MET and 1000 µg. mL−1 of both EMP and LIN were prepared in methanol. These stock solutions were stable for at least one month when kept refrigerated at 4 ºC. Suitable aliquots from EMP, LIN and MET stock solutions were transferred separately into a series of 10-mL volumetric flasks in order to prepare their working solutions. This was followed by completing the volume using deionized water to reach the concentration ranges 10–500, 10–100 and 2.5–100 µg. mL−1 for MET, EMP and LIN, respectively.Stock solutions 5000 µg. mL®, Mix III), or 5 mg LIN/ 10 mg EMP was extracted with 25 mL methanol in separate 25-mL volumetric flasks. Following sonication for 20 min and further filtration (Whatman No. 1 filter paper 110 mm diameter), volumes of 0.125 mL (Mix I), 0.25 mL (Mix II), 0.25 mL (Mix III) and 1.25 mL (Mix IV) were diluted with deionized water in separate 10-mL volumetric flasks to get working tablet solutions equivalent to 12.5 EMP/2.5 LIN/500 MET (Mix I), 12.5 EMP/500 MET (Mix II), 2.5 LIN/500 MET (Mix III) / 25 LIN and 50 EMP (Mix IV), respectively. Final concentrations of compounds in µg. mL−1.A number of ten tablets were weighed and finely powdered. The content of one powdered tablet equivalent to 25 mg EMP/5 mg LIN/1000 mg MET , 12.5 mg EMP/500 mg MET , 2.5 mg LIN/500 mg MET  technique at various pH range (3–10) using phosphate buffer was tried but it failed to analyze the triple combination due to pka value of EMP , it is considered as a weakly ionizable compound . It is nAs a result, MEKC was performed by adding a surfactant such as the anionic SDS exceeding its critical micelle concentration (CMC) to Tris buffer solution. In this mode, both neutral and charged molecules can be separated so it improved the peak shape and symmetry of EMP .The partConsequently, to solve this problem a modification was performed by the addition of organic modifier such as methanol to the BGE. Addition of methanol causes alterations in the viscosity and dielectric constant of the electrolyte, thus a significant reduction in the velocity of BGE and separation of the coeluted drugs EMP and LIN with reasonable resolution and migration time , 28.The following parameters were studied; buffer type and pH, buffer strength, SDS concentration, percentage of organic modifier, applied voltage, applied pressure, injection time, operating temperature, and detection wavelength.Two buffer systems (20 mM) were investigated namely, phosphate buffer and Tris buffer. In all the analytical conditions, phosphate buffer, on the contrary of tris buffer, was associated with high currents (40–65 µA), which lead to excessive heating of the capillary and their breakage , 29. As The role of the buffer pH is crucial relative to the degree of ionization of drugs. The pKa values are: 12.57, 12.4 and 8.6 for EMP, MET and LIN respectively. ConsequTris buffer solutions (pH 10.0) at different strength values  were tested for its effect on migration time and peak shape. Low buffer strength decreased the resolution between EMP and LIN with value 1.4 (< 2), while upon increasing the strength of Tris buffer, it insignificantly increased the migration time of the EMP, LIN and MET. A strength of 20 mM was optimum regarding the cited drugs peaks symmetry and reasonable migration times.The effect of SDS concentration on EMP, LIN and MET separation was studied by addition of 25, 50 and 100 mM SDS in presence of 10% v/v methanol in the BGE. As SDS concentration increased, the migration time slightly increased as well. Despite the short migration time upon using 25 mM SDS, bad resolution was observed between EMP and LIN peaks. Therefore, 50 mM of SDS was selected for separation of MET, EMP and LIN as it is the best compromise between peaks shape and analysis time taking into consideration that they are well separated at the studied SDS concentrations . Upon using 15% methanol, migration time of the compounds increased. The high content of methanol > 20% can hinder micelle formation . TherefoThe absorption spectra of EMP, LIN and MET, Additional file Instrumental parameters optimization including applied voltage, applied pressure, injection time and operating temperature are discussed in the supplementary material file.Final separation was achieved within 6 min using a BGE of Tris buffer  containing 50 mM of the anionic SDS and 10% v/v MeOH as an organic modifier at 25 ºC as an operating temperature, 30 kV as the applied voltage, 50 mbar and 15 s as pressure and injection time, respectively. The three anti-diabetic compounds  were detected at wavelength 230 nm. A typical electropherogram of MET, EMP, and LIN, analyzed using the proposed MEKC conditions was shown in Fig. The proposed MEKC technique was fully validated as per the ICH guidelines .−1. Regression and statistical parameters were calculated as shown in Table –8) [Linearity was assessed in the concentration ranges of 10–100 (EMP), 2.5–100 (LIN), and 10–500 (MET) µg. mLable –8) –37.Table−1) and LOQ  values for EMP, LIN and MET, respectively.LOD and LOQ of EMP, LIN and MET were determined based on S/N ratio of 3: 1 for LOD and 10: 1 for LOQ. The proposed MEKC method yielded low LOD  ranged from 0.39 to 1.66%, 0.25 to 1.84% and 0.47 to 1.98% for EMP, LIN and MET, respectively. Inter-day precision ranged from 0.42 to 0.66%, 0.91 to 1.78% and 0.57 to 1.96% for EMP, LIN and MET, respectively.Moreover, the proposed method could analyze accurately and precisely EMP and LIN in their low critical concentrations with MET  and .Various parameters in the proposed MEKC method were slightly changed such as: pH of buffer , buffer strength , SDS concentration , percent of organic modifier  in addition to detection wavelength  were varied.The impact of these minor changes on the migration time (tm) values and peaks area were considered by calculating the RSD% of the peak areas and SD of the migration times for each compound. The low values of RSD% < 1.62) along with almost unchanged migration times values obtained after inducing minor intentional changes in the methods parameters revealing high degree of method’s robustness. . Also, the MEKC method specificity was further confirmed by obtaining acceptable values for the system suitability parameters as EMP, LIN and MET peaks were well separated from each other and showed good resolution values (above 2.25). Moreover, the overlying of the spectra recorded at different wavelength intervals during the registration of each electrophoretic peak in the analysis of dosage forms obtained from the DAD indicates the purity of the peaks Fig. : Fig. S4Stock solutions and standard solutions of EMP, LIN and MET were found to be stable for one month at 4 ℃ and for 8 h at 25 ℃, respectively. No changes in peak areas and no extra peaks were found.The applicability of the suggested MEKC method was extended to the analysis of ternary pharmaceutical mixture of Mix I, and three binary mixtures Mix II, Mix III and Mix IV. These prepared tablet solutions "" sectionThe analytical methodology should strike a compromise between being environmentally friendly and functionality in terms of completing the scope of application with a high degree of accuracy and precision, saving time and money, and consuming less waste and harmful chemicals. Consequently, the greenness and whiteness of the current work were assessed and compared with reported HPLC , HPTLC , UPLC 1 and specUsing the semi quantitative analytical Eco-scale, which depends on subtracting the penalty points (PP) out of total score 100, our proposed MEKC method had a score equal to 86 more than 75, so it is considered an excellent green analysis  12 model . Up to 1To sum up, upon applying the three approaches we found that our proposed MEKC method is superior to reported chromatographic and spectrophotometric methods concerning high separation efficiency with short analysis time, small quantities of reagents and solvents with minimal negative impact on the environment and human health. Thus, it was valuable to develop a MEKC method for rapid, green, white, and effective determination of the three antidiabetic medications.A simple, rapid, inexpensive green and white MEKC method has been developed for the first time for the simultaneous determination of the combo pill triple therapy containing LIN and EMP in their lowest critical concentration ratios with MET. To our knowledge, this triple combination and EMP binary combination with MET or LIN were not previously determined using MEKC or even the conventional CE method. The MEKC method for the analysis of MET, EMP and LIN was totally optimized and validated as per the ICH guidelines. Moreover, greenness and whiteness appraisal for the suggested MEKC method were done using two different green metrics  and white model  ascertaining that our MEKC method was greener and whiter when compared to other published methods.The proposed MEKC method is characterized by high resolving power that allowed this mode to rapidly analyze in relatively short periods of time a large number of potential drugs with good resolution in the same run despite the large difference between the doses of LIN or EMP with MET. Moreover, minimal sample and aqueous buffer consumption adds the advantages of method’s greenness and economic. This promotes our proposed MEKC method to be used in quality control laboratories not only for its high-through-put but also because it is efficient, simple green and white analytical tool.Additional file 1: Instrumental parameters optimization. Figure S1. Effect of pH of 20 mM Tris buffer on the asymmetry factor of EMP (a), electropherogram showing EMP shouldered peak obtained upon trying BGE at pH 9 as representative example (b). Figure S2. Effect of (a) SDS concentration (b) percent of organic modifier (c) applied voltage on the migration time of MET, EMP, and LIN. Figure S3. The absorption spectra of (a) MET (b) EMP and (c) LIN of the standard mixture extracted from DAD. Figure S4. The peaks purity plot and profile from tablet extract of the three compounds MET, EMP and LIN respectively measured at 230 nm for the analysis of MET/LIN/EMP (Mix I) as a representative example. Table S1. System suitability parameters for the MEKC determination of MET, EMP and LIN ternary mixture. Table S2. Intra-day and Inter-day precision and accuracy for the determination of MET, EMP and LIN using the proposed MEKC method. Table S3. Evaluation of the robustness of the proposed MEKC method for the determination of MET, EMP and LIN ternary mixture. Table S4. Analytical eco-scale for assessment of greenness of the proposed MEKC method.


In order to efficiently investigate the effect of the mesoscale heterogeneity of a concrete core and the randomness of circular coarse aggregate distribution on the stress wave propagation procedure and the response of PZT sensors in traditional coupling mesoscale finite element models (CMFEMs), firstly, a mesoscale homogenization approach is introduced to establish coupling homogenization finite element models (CHFEMs) with circular coarse aggregates. CHFEMs of rectangular concrete-filled steel tube (RCFST) members include a surface-mounted piezoelectric lead zirconate titanate (PZT) actuator, PZT sensors at different measurement distances, a concrete core with mesoscale homogeneity. Secondly, the computation efficiency and accuracy of the proposed CHFEMs and the size effect of representative area elements (RAEs) on the stress wave field simulation results are investigated. The stress wave field simulation results indicate that the size of an RAE limitedly affects the stress wave fields. Thirdly, the responses of PZT sensors at different measurement distances of the CHFEMs under both sinusoidal and modulated signals are studied and compared with those of the corresponding CMFEMs. Finally, the effect of the mesoscale heterogeneity of a concrete core and the randomness of circular coarse aggregate distribution on the responses of PZT sensors in the time domain of the CHFEMs with and without debond defects is further investigated. The results show that the mesoscale heterogeneity of a concrete core and randomness of circular coarse aggregate distribution only have a certain influence on the response of PZT sensors that are close to the PZT actuator. Instead, the interface debond defects dominantly affect the response of each PZT sensor regardless of the measurement distance. This finding supports the feasibility of stress wave-based debond detection for RCFSTs where the concrete core is a heterogeneous material. As typical structural components carrying increasing vertical or axial load, large-scale concrete-filled steel tube (CFST) members have been extensively applied as columns in high-rise buildings or arches and piers in long-span bridges. The non-uniformly distributed temperature in the curing procedure of the concrete core of CFST members after pouring concrete and the unavoidable long-term shrinking and creeping of concrete core may cause interface debond defects between the concrete core and steel tube, weakening the desired confinement effect of the steel tube on the concrete core and negatively affecting the ductility and load-carrying capacity of the CFST members. Therefore, the development of reliable interface debond defect detection methodologies for the CFST members is of great emergency. Most of the traditional non-destructive testing (NDT) techniques, which have been successfully applied in defect detection for reinforced concrete (RC) or steel structures, including infrared thermal imaging ,2, electRecently, defect detection for concrete materials, composites, RC and steel structures based on stress wave measurement using piezoelectric lead zirconate titanate (PZT) actuators and sensors have attracted much attention ,11,12,13One of the most potential defects in CFST members is the interface debond defect between the concrete core and steel tube. In order to accurately detect interface debond defects in CFST members, Xu et al. ,22 develHowever, one of the main concerning shortages in the above-mentioned numerical analysis on stress wave travel in CFST members is that the concrete core was considered as a homogeneous material with uniform material parameters. Considering the fact that concrete is a typical heterogeneous material, composed of mortar, coarse and fine aggregates and an interfacial transition zone (ITZ) between aggregates and the mortar, etc., it has been a common concern if the mesoscale heterogeneity and randomness of aggregate distribution in the concrete core dominantly affect stress wave travel within CFSTs and PZT sensor responses at different measurement distances when compared with interface debond defects in CFSTs . With thThe mesoscale homogenization method is computationally efficient for the behavior simulation of mesoscale concrete materials and structures, where the mesoscale concrete is modeled with a number of representative area elements (RAEs) two-dimensionally (in 2D) or representative volume elements (RVEs) three-dimensionally (in 3D). The concrete material of each RVE or RAE is homogenized with uniform mechanical properties ,33,34. WIn the study by Wang et al. , the effIn this paper, multi-physics coupling mesoscale finite element models (CMFEMs) composed of a surface-mounted PZT actuator, a surface-mounted or embedded PZT sensor, and a 2D mesoscale RCFST specimen with randomly distributed circular aggregates are firstly established. After that, the corresponding coupling homogenization finite element models (CHFEMs) with different RAE dimensions are established to study the effect of RAE dimensions on both stress wave fields and the response of PZT sensors at different measurement distances. For simplicity, the stress wave fields in mesoscale finite element models (MFEMs) and the corresponding homogenization finite element models (HFEMs) with and without debond defects are determined considering different RAE dimensions. The effects of RAE dimensions on the simulation results of stress wave fields are analyzed comprehensively. Therefore, the PZT sensor responses with different measurement distances in CMFEMs and the corresponding CHFEMs with and without debond defects under various excitation signals are also determined considering different RAE dimensions. The results show that the mesoscale heterogeneity of a concrete core and the randomness of circular aggregate distribution only have a certain influence on the response of PZT sensors that are close to the surface-mounted PZT actuator. Compared with the mesoscale heterogeneity of a concrete core in the form of the random distribution of circular aggregates, the interface debond defect dominantly affects the stress wave fields and the response of PZT sensors at various measurement distances.In recent years, the effect of the morphology of coarse aggregates on the properties of concrete materials and structures has been an active topic . In thisThe piezoelectric equation for the employed PZT patches can be found in the previous studies ,37,38,39In addition, the equation of motion for the coupled RCFST–PZT model is described in the form of vectors and generalized matrices ,35:(2)M0In this study, the plane dimensions of the concrete core of the 2D RCFST member are 400 mm by 400 mm and the thickness of the concrete core is 10 mm. The concrete core is surrounded by a plane steel tube with a thickness of 5 mm. The employed PZT patches acting as either sensors or actuators are of a length of 10 mm and a thickness of 0.3 mm. The polarization direction of each PZT patch is its thickness direction. In order to investigate the influence of different measurement distances between the PZT actuator and the PZT sensors, a number of mesoscale RCFST–PZT coupling models with different PZT sensor locations are established with the RAM approach ,35.The size and quantity of circular aggregates in a specific RCFST member are specified according to an ideal gradient curve shown as follows .(3)P with the Walraven method.i-th and j-th circular aggregates and In a Cartesian coordinate system, the coordinates of the center of a circle are randomly generated within the RCFST member using the Monte Carlo method. In order to avoid circular aggregate overlap and contact, the distance between any two circular aggregates must be greater than a certain value. The overlap of two circular aggregates is judged based on the distance between their centers according to Equation (5). The aggregate with a large radius should be put in first because the effective delivery area after the successful delivery of an aggregate is gradually reduced.Here, three CMFEMs with different randomly distributed circular coarse aggregates mimicking the mesoscale heterogeneity and randomness of a concrete core, i.e., CMFEM 1, 2 and 3, are established as shown in As a further investigation of the previous study by Wang et al. , CHFEMs In order to study the computational accuracy of the CHFEMs, CHFEMs with different RAE dimensions are established for each CMFEM. The material properties including density, Poisson’s ratio and elastic modulus of each RAE with different dimensions can be determined according to the method proposed by Wang et al. , where eFor the debond defect detection of RCFST members using stress wave fields and PZT sensor response measurements, high-frequency signals are commonly used as inputs on a PZT actuator mounted on the surface of the steel tube of an RCFST. The high-frequency excitation signal leads to a short stress wave wavelength, where fine meshing and short integration time steps are required. Here, the typical 2D RAE dimensions of 20 mm by 20 mm, 25 mm by 25 mm and 40 mm by 40 mm for each CMFEM model are considered to distinguish the size effect of RAEs on the stress wave fields and PZT sensors measurement simulation results. Since the advantage of the computational efficiency of CHFEM has been demonstrated in the recent research by Wang et al. , a compaFor the mesoscale simulation of the stress wave field within different mesoscale RCFSTs, it is convenient to exclude the direct and inverse piezoelectric effects of PZT actuators and sensors and the coupling effect between them and the RCFST member. In this study, the stress wave propagation procedure within different mesoscale finite element models (MFEMs) of RCFSTs and the corresponding homogenization finite element models (HFEMs) employing different RAE sizes under the excitation of a pulse force are investigated for comparison.The applied pulse force signal for stress wave fields and modulated sinusoidal excitation signal for the PZT sensor measurement simulation is the same as that in . The modThe measurement results of different PZT sensors under both sinusoidal and modulated signals are simulated using both the CMFEMs and the corresponding CHFEMs, where the circular coarse aggregates are randomly distributed and the concrete core is heterogeneous at the mesoscale.In the study by Wang et al. , the infWithout a loss of generality, the stress wave fields of the healthy MFEM 1 without interface debond defects and the corresponding HFEMs with different RAE dimensions at different time instants are investigated and compared at first. The three side dimensions of 20 mm, 25 mm and 40 mm for square RAEs are considered.In In addition to a comparison of the element numbers, DOFs and the computation time for stress wave propagation simulation between the healthy MFEM 1 and its corresponding HFEMs with different RAE dimensions are shown in In order to further investigate the effect of different RAE dimensions on the stress wave field simulation results and to compare it with that of the interface debond defect, the stress wave fields of the MFEM 1 with a debond defect and the corresponding HFEMs with different RAE dimensions at different time instants are simulated. As shown in The stress wave fields of the MFEM 1 with an interface debond defect and its corresponding HFEMs with different RAE dimensions are shown and compared in When comparing the stress wave fields of the healthy RCFSTs at each identical time instant shown in In practice, for the debond detection of RCFST members, it is difficult to get the stress wave fields at different time instants. PZT sensors are usually used to measure the stress wave at certain locations and the measurements are used to detect the debond defect. Therefore, a further numerical study on the response of PZT sensors in CMFEMs and the CHFEMs at different measurement distances is carried out to distinguish the influences of the mesoscale heterogeneity of concrete on PZT sensor measurement and those of debond defects in the following chapter.The effects of both themesoscale heterogeneity of concrete cores in the form of randomly distributed circular coarse aggregates and debond defects on the output of PZT sensors at different measurement distances from the PZT actuator are investigated. Multi-physics field simulation on the measurement of PZT sensors of three CMFEMs and the corresponding CHFEMs with different RAE dimensions under both sinusoidal excitation and modulated excitation is performed. The frequency of the sinusoidal input signal is 20 kHz and the frequency of the modulated excitation signal described in Equation (3) is 20 kHz. Both input signals are of an amplitude of 10 V. The distances between the PZT sensor and actuator are assigned to be 80 mm, 160 mm, 240 mm, 320 mm and 410 mm. The distance of 410 mm means that the PZT sensor is installed on the opposite side surface of the RCFST, representing the surface-mounted PZT actuator and sensor installation strategy. Considering the similarity of the stress wave fields of the HFEMs with different RAE dimensions shown in In order to investigate the effect of interface debond defect on PZT sensor measurements, a numerical simulation on the PZT response of the three CMFEMs with identical debond defects, CMFEM 1D, CMFEM 2D and CMFEM 3D, is also carried out. Comparing the PZT sensor response at identical measurement distance of CMFEMs with and without a debond defect as shown in In this section, the time-domain responses of PZT sensors at different measurement distances from the PZT actuator of the three CMFEMs with and without an interface debond defect under the modulated sinusoidal excitation signal are investigated and the results are shown in The similarity of the PZT sensor measurements at an identical CMFEM measurement distance to different mesoscale structures can be found in In the previous study by the authors of , the infTo further compare the influence of the debond on the measurement of PZT sensors at different measurement distances, the responses of PZT sensors of the CHFEMs with an interface debond defect using different RAE dimensions are shown in A further investigation on the influence of RAE dimensions of CHFEMs on PZT sensor responses under the modulated sinusoidal excitation signal is performed. From all the above-mentioned simulation results of both CMFEMs and CHFEMs, it is concluded that debond defects are the dominant factor affecting the PZT sensor’s response at identical measurement distances. A close look at the influence of the heterogeneity of concrete cores and randomness of coarse aggregate distribution in RCFSTs on the responses of PZT sensors at different measurement distances is taken in a quantitative manner in the following chapter.Further analysis of the maximum amplitude of the time-domain response of the PZT sensors under a sinusoidal input signal and of the wavelet packet energy of the response of PZT sensors under a modulated input signal is carried out.As shown in Here, the effect of different measurement distances on the response of PZT sensors is further elucidated under sinusoidal signals. The average amplitude and variance in PZT sensors at identical measurement distances in the three CMFEMs and three CHFEMs shown in Here, the influence of the mesoscale hetereogeneity of a concrete core and the randomness of coarse aggregate distribution and of the debond defect on the responses of the PZT sensors of three CMFEMs and the three CHFEMs corresponding to CMFEM 1 under the modulated sinusoidal signal is further distinguished.A wavelet packet analysis of each sensor response is carried out to determine the wavelet packet energy of the output signals collected by each PZT sensor , and theAs shown in Multi-physics and multi-scale CMFEMs composed of a PZT sensor, a PZT actuator and a 2D RCFST cross-section are constructed, and the corresponding CHFEMs with different RAE dimensions are established. The feasibility of using CHFEMs for stress wave field and PZT sensor response simulation and their computational efficiency are illustrated. Then, the difference in the influences of the mesoscale heterogeneity of a concrete core in the form of the randomness of coarse aggregate distribution in a RCFST and of the debond defect on the responses of PZT sensors at different measurement distances are elucidated. Based on the analysis of the numerical simulation results, the following conclusions can be made.(1) The feasibility and the computational efficiency of the proposed mesoscale homogenization approach for RCFST stress wave fields and PZT sensor response simulation are numerically confirmed. The stress wave fields of the CMFEMs and their corresponding CHFEMs with different RAE dimensions are similar, and the influence of an interface debond defect on the stress wave field is dominant. The concrete core with mesoscale homogeneity in the form of random coarse aggregate distribution in RCFSTs has no obvious influence on the stress wave fields of RCFST specimens.(2) By comparing the PZT sensor measurement simulation results of healthy CMFEMs and those of the corresponding CHFEMs, the heterogeneity in the form of random concrete core distribution only affects the responses of PZT sensors close to the PZT actuator. The influence of mesoscale heterogeneity on PZT sensor measurement becomes weaker when the measurement distance increases. RAE dimensions have no obvious influence on the PZT responses of CHFEMs. This finding also works for CMFEMs and CHFEMs of RCFST members with interface debond defects.(3) From the simulation results of the CMFEMs and the CHFEMs, it is clear that the interface debond defects of RCFSTs have a more obvious effect on the response of each PZT sensor when compared with those of a heterogenous and random distribution a concrete core at the mesoscale in RCFSTs. (4) The findings from this study imply that the interface debond defect detection method using stress wave measurement from a PZT sensor is reasonable in practice even though the concrete core in RCFST specimens is a heterogenous and random material. The mesoscale heterogeneity and randomness of concrete cores affect PZT sensor responses locally, and the interface debond defect is the dominant factor affecting the PZT sensor measurement no matter what the measurement distance is.


We investigated sensitivity of the 2020 Revised Comprehensive Diagnostic Criteria (RCD) and the 2019 ACR/EULAR classification criteria across the four identified IgG4-related disease (IgG4-RD) phenotypes: “Pancreato-Hepato-Biliary”, “Retroperitoneum and Aorta”, “Head and Neck-limited” and “Mikulicz’ and Systemic” in a well-characterized patient cohort.We included adult patients diagnosed with IgG4-RD after comprehensive clinical assessment at Oslo University Hospital in Norway. We assigned patients to IgG4-RD phenotypes based on pattern of organ involvement and assessed fulfillment of RCD and 2019 ACR/EULAR classification criteria. Differences between phenotype groups were analyzed using one-way ANOVA for continuous variables, and contingency tables for categorical variables.p < 0.001). Among the 57 (72.2%) patients meeting the 2019 ACR/EULAR classification criteria, proportion was again lowest in the “Retroperitoneum and Aorta” group .The study cohort included 79 IgG4-RD patients assigned to the “Pancreato-Hepato-Biliary” (22.8%), Retroperitoneum and Aorta” (22.8%) “Head and Neck-limited” (29.1%), and “Mikulicz’ and Systemic” (25.3%) phenotype groups, respectively. While 72/79 (91.1%) patients in total fulfilled the RCD, proportion differed across phenotype groups and was lowest in the “Retroperitoneum and Aorta” group (66.7%, The results from this study indicate that IgG4-RD patients having the “Retroperitoneum and Aorta” phenotype less often fulfill diagnostic criteria and classification criteria than patients with other IgG4-RD phenotypes. Accordingly, this phenotype is at risk of being systematically selected against in observational studies and randomized clinical trials, with potential implications for patients, caregivers and future definitions of IgG4-RD. IgG4-related disease (IgG4-RD) is a fibroinflammatory systemic disease that can involve nearly any organ. Core features include tissue infiltration of IgG4-positive plasma cells causing tumefactive lesions and/or organomegaly, frequently accompanied by elevated serum IgG4 concentration .IgG4-RD is a diagnostic challenge, owing to its heterogeneous presentations and lack of pathognomonic features. Diagnosis requires correlation of clinical, serological, radiological and/or histopathological findings . ComprehThe unsettled status of diagnostic criteria for IgG4-RD is not unexpected. It reflects that development of accurate diagnostic criteria for complex diseases with overlap to mimicking conditions is inherently challenging, as evident from the near complete absence of diagnostic criteria in rheumatology . InsteadThe 2019 ACR/EULAR IgG4-RD classification criteria were developed by an international expert group. In the two separate validation cohorts, the reported sensitivities of the criteria were 85.5% and 82.0%, respectively, while specificities were 99.2% and 97.8% [Following publication of the 2019 ACR/EULAR classification criteria, Wallace et al. used data from the validation cohorts to identify four distinct clinical phenotypes of IgG4-RD with different patterns of organ involvement: (i) “Pancreato-Hepato-Biliary”; (ii) “Retroperitoneum and Aorta”; (iii) “Head and Neck-limited” and (iv) “Mikulicz’ and Systemic” . ImportaHere, we aimed to assess sensitivity of the RCD and the 2019 ACR/EULAR classification criteria across the four phenotypes. We included a well-characterized Norwegian cohort of patients with IgG4-RD diagnosed by expert clinical assessment, stratified by phenotype, and assessed criteria performance. As our study cohort did not include patients diagnosed with mimicking conditions, we were not able to assess the specificity of the criteria.At the Department of Rheumatology at Oslo University Hospital (OUH) we consecutively include consenting adult patients (≥ 18 years) diagnosed with IgG4-RD by expert clinical assessment in the Norwegian systemic connective tissue disease and vasculitis registry (NOSVAR) . For thiElevated serum IgG4 levels were defined as > 1.35 g/L for the CDC and RCD criteria , as per Organ involvement was determined by clinical, histopathological and/or radiological findings, where other causes were deemed unlikely. Multi-organ involvement was defined as ≥ 2 involved organs. Two rheumatologists  assessed fulfilment of the CDC, RCD and 2019 ACR/EULAR classification criteria, and assigned patients to one out of four phenotypes based on pattern of organ involvement .Written informed consent was given for the included IgG4-RD patients as requirement for inclusion in NOSVAR. The study was conducted in compliance with the Helsinki Declaration and approved by the regional ethics committee (REK #342136).Both the CDC and RCD include three variables: (i) clinical and radiological findings suggestive of IgG4-RD; (ii) elevated serum IgG4 level (defined as > 1.35 g/L); and (iii) compatible histopathological findings [The 2019 ACR/EULAR classification criteria employ a three-step approach, which includes (i) an obligatory entry criterion ; (ii) a set of exclusion criteria; and (iii) a list of classification items with weighted scores assigned to various clinical, serological, and histopathological features. Following exclusion of mimickers, we classified patients as IgG4-RD cases if they (i) met the entry criterion, (ii) had no exclusion criteria and (iii) scored ≥ 20 points by the defined classification items [Fulfilment of CDC, RCD and 2019 ACR/EULAR classification criteriaReasons for failure to fulfil the criteriaIn this cohort of well-characterized patients diagnosed with IgG4-RD based on expert clinical assessment, we aimed to describe, both on a group and phenotypic level:Descriptive statistics were applied, using IBM SPSS version 26 for Windows . Continuous variables are reported as means and standard deviations, and between-group differences analyzed using one-way ANOVA. Categorical variables are reported as absolute number and percentage, and between-group differences analyzed using contingency tables.p = 0.002), more often female (p = 0.024), and demonstrated a trend toward more non-white patients. The “Retroperitoneum and Aorta” group had the highest mean CRP (p < 0.001) and ESR (p = 0.001) and was characterized by the lowest mean serum IgG4 concentration, less frequent multi-organ disease (p = 0.03), and fewer biopsies (p < 0.001). The “Mikulicz’ and Systemic” group had the highest mean serum IgG4 concentration and mean number of involved organs (p < 0.001 for both).The IgG4 study cohort included 79 patients Table . In the p < 0.001).In total, 72 patients (91.1%) fulfilled the CDC and RCD. Discrepancy between CDC and RCD only occurred twice: two patients deemed “possible” IgG4-RD by CDC were considered “definite” by RCD. This discrepancy related to the histopathological domain of these criteria. Both patients had dense lymphoplasmacytic infiltrate with fibrosis, and > 10 IgG4 + plasma cells per hpf. The tissue IgG4 + /IgG + plasma cell ratio was < 0.40 , but there was evidence of storiform fibrosis and obliterative phlebitis . Given these minor differences, we decided to focus on RCD for all further analyses. Fulfilment of RCD was lower in the “Retroperitoneum and Aorta” group (66.7%) than in the remaining groups: 100% in “Pancreato-Hepato-Biliary”, 100% in “Head and Neck-limited” and 95.0% in “Mikulicz’ and Systemic” phenotype. The between-group difference was statistically significant (p < 0.001).Fifty-seven patients (72.2%) in the IgG4-RD cohort fulfilled the 2019 ACR/EULAR classification criteria, with 100% meeting the criteria in both the “Pancreato-Hepato-Biliary” and “Mikulicz’ and Systemic” groups. The percentage of patients fulfilling the classification criteria was lower in the “Retroperitoneum and Aorta” group (27.8%) and the “Head and Neck-limited” group (60.9%)  had elevated baseline serum IgG4 (> 2.01 g/L), and 11 (50.0%) had performed biopsy, excluding fine needle aspiration.n = 3), (ii) fulfilment of one or more exclusion criteria (n = 5) or (iii) failure to achieve the required 20 points (n = 14).The reasons why the 22 patients did not meet the 2019 ACR/EULAR classification criteria are summarized in Fig. Of the 13 patients in the “Retroperitoneum and Aorta” group who failed to fulfil the 2019 ACR/EULAR classification criteria (Table n = 1), positive anti-RNP (n = 1) or positive MPO-ANCA (n = 3). In the MPO-ANCA positive group, 2 patients were presumed to have coexisting IgG4-RD and microscopic polyangiitis. Of the 7 patients who failed to fulfil the entry criterion and/or fulfilled an exclusion criterion, 5 (71.4%) achieved the required 20 points in the subsequent domain of the classification criteria.Of the 9 patients in the “Head and Neck-limited” group who failed to fulfil the 2019 ACR/EULAR classification criteria Table , the cliAmong the 22 patients who did not fulfil the 2019 ACR/EULAR classification criteria, 16 (72.7%) fulfilled the RCD, with 5, 2 and 9 patients considered “definite”, “probable” and “possible” IgG4-RD, respectively , the net result may be lost opportunities for treatment of individual patients, and skewed apprehension of disease features.To our knowledge, our study is the first to describe fulfilment of RCD and 2019 ACR/EULAR classification criteria across the four phenotypes, highlighting potentially important differences across phenotypes. Fulfilment of classification criteria is usually a prerequisite for inclusion in studies in the field of rheumatology. Hence, the subset of patients fulfilling such criteria largely shape our understanding of a disease over time . ImportaOur cohort demonstrated similar disease characteristics and phenotypic distribution as the multinational phenotype derivation cohort , and mosConsidering the inherent ambiguity when diagnosing a complex and heterogenous disease, we chose to describe the patients not fulfilling the 2019 ACR/EULAR classification criteria, for transparency and to allow recalculation based on alternative interpretations by the readers.The most common reason for not fulfilling the 2019 ACR/EULAR classification criteria in our IgG4-RD cohort was inability to achieve the required 20 points in the final step of the criteria . This waImportantly, clinical experience indicates that the “Retroperitoneum and Aorta” and “Head and Neck-limited” phenotypes are more treatment refractory than the remaining groups . Taken tAs we did not have access to patients with mimicking conditions in this study, we were unable to calculate the specificity of any criteria. It is reasonable to assume that RCD has a low specificity for IgG4-RD, as it focuses on largely nonspecific features of the disease. This is particularly true for cases designated as “possible” IgG4-RD, which largely rests on elevated serum IgG4, a finding seen in many inflammatory conditions. Accordingly, we do not suggest the superiority of these criteria, nor do we support favoring their use to identify patients for clinical trials. Rather, the main finding in our study is the potential limitation of the 2019 ACR/EULAR classification criteria for certain phenotypes, which may have implications for future research. Whether increasing the weighed score assigned to “typical” retroperitoneal fibrosis and/or including orbital pseudotumor as a weighted manifestation alleviate this shortcoming without significantly sacrificing specificity is unclear but warrants further discussion. We encourage further research to evaluate the specificity of the criteria in large cohorts that include patients diagnosed with mimicking conditions.18FDG PET/CT in many patients. Hence, it seems unlikely that the failure to achieve the required 20 points reflects inability to capture additional, mild and/or asymptomatic disease manifestations.The strength of our study is a well-described cohort followed at a tertiary referral center with rheumatologists, pathologists, radiologists, and other specialists experienced in IgG4-RD. Furthermore, the work-up included advanced imaging, including The limitations of this study include its single center design with partly retrospectively collected data and predominantly White patients. Another limitation is the lack of baseline (pre-treatment) serum IgG4 in some patients, the fact that some patients did not have a biopsy performed, and the inherent diagnostic ambiguity in this field.Our study demonstrated that the 2019 ACR/EULAR classification criteria did not capture most patients with the “Retroperitoneum and Aorta” and “Head and Neck-limited” phenotypes of IgG4-RD. Hence, through a lower ability to capture these subgroups, results from studies based on these criteria, may not be representative for the whole disease population.