ObjectiveTo establish a high-performance liquid chromatography（HPLC） for the quantitative analysis of multiple components in Gegen Qinlian tablets， and to comprehensively evaluate the quality of samples from different manufacturers by integrating chemical pattern recognition and technique for order preference by similarity to ideal solution（TOPSIS）， in order to provide a reference basis for quality evaluation and control of Gegen Qinlian tablets. MethodsHPLC was employed to determine the contents of 10 components in 28 batches of Gegen Qinlian tablets collected from 6 manufacturers， and taking the detection results as variables， SIMCA 14.1 and SPSS 26.0 were employed for cluster analysis（CA）， principal component analysis（PCA）， and orthogonal partial least squares-discriminant analysis（OPLS-DA） to identify key components affecting the quality. Then， TOPSIS analysis was employed to rank the quality of Gegen Qinlian tablets from the 6 manufacturers and establish a comprehensive quality evaluation method. ResultsA quantitative method for Gegen Qinlian tablets was established. After methodological validation， the method was found to be stable and reliable， and could be used for the quantitative analysis of this preparation. The contents of 3′-hydroxy puerarin， puerarin， 3′-methoxy puerarin， daidzein， coptisine hydrochloride， epiberberine， jatrorrhizine hydrochloride， berberine hydrochloride， palmatine hydrochloride and baicalin in 28 batches of samples were 3.58-7.35， 24.88-42.32， 4.20-9.36， 4.33-7.60， 2.52-6.44， 0.93-4.10， 0.58-3.05， 10.68-22.92， 0.82-4.82， 11.73-60.16 mg·g^-1， respectively. Among them， puerarin， berberine hydrochloride and baicalin all met the limit requirements for this preparation specified in the 2025 edition of the Pharmacopoeia of the People's Republic of China. CA and PCA clustered the 28 batches of samples into 5 categories， PCA extracted 2 principal components with a cumulative variance contribution rate of 90.588%， and OPLS-DA screened out 4 differential markers with variable importance in the projection（VIP） values>1.0， namely baicalin， 3′-hydroxy puerarin， coptisine hydrochloride and palmatine hydrochloride， which might be the main components affecting the quality of Gegen Qinlian tablets. TOPSIS analysis showed that the comprehensive score of each evaluation index（C_i） values of different manufacturers were different. Among them， the C_i of manufacturer B was ranked higher， indicating potentially superior quality， while the C_i of manufacturer A was ranked lower， suggesting potentially inferior quality. ConclusionThis study establishes a quantitative method for Gegen Qinlian tablets， and the content uniformity of the same manufacturer is good， while there are differences in the contents of active components among different manufacturers. Through the chemical pattern recognition analysis， it is found that the content differences of Gegen Qinlian tablets may be related to baicalin， 3′-hydroxy puerarin， coptisine hydrochloride and palmatine hydrochloride.

OBJECTIVE To establish a high-performance liquid chromatography（HPLC） fingerprint and a multi-component content determination method for Ziziphus jujuba stir-fried with Coptis chinensis juice， a characteristic processed product of Longsha Medical School， and to provide a basis for the inheritance and clinical application of this distinctive variety. METHODS HPLC method was employed to establish the fingerprints of 12 batches of Z. jujuba stir-fried with C. chinensis juice. Similarity evaluation and common peak identification were subsequently carried out. Quantitative determination was performed on coclaurine， magnoflorine， coptisine hydrochloride， epiberberine， jatrorrhizine hydrochloride， spinosin， berberine hydrochloride， palmatine hydrochloride， 6"'-feruloylspinosin， and jujuboside A of samples. RESULTS The fingerprint similarities of all 12 batches of samples were higher than 0.960. A total of 17 common peaks were calibrated， among which 9 peaks were identified as coclaurine， magnoflorine， coptisine hydrochloride， epiberberine， jatrorrhizine hydrochloride， spinosin， berberine hydrochloride， palmatine hydrochloride， and 6"'-feruloylspinosin， respectively. The quantitative results showed that the mass fractions of coclaurine， magnoflorine， coptisine hydrochloride， epiberberine， jatrorrhizine hydrochloride， spinosin， berberine hydrochloride， palmatine hydrochloride， 6"'-feruloylspinosin， and jujuboside A in the 12 batches of samples were 0.044-0.091， 0.908-1.698， 0.325-0.549， 0.163-0.287， 0.103-0.172， 0.712-0.952， 1.185-1.843， 0.341-0.598， 0.228-0.405， and 0.050-0.091 mg/g， respectively. CONCLUSIONS The established HPLC fingerprint and multi-component determination method is stable and reliable， and can provide a basis for the quality standard formulation and clinical application of Z. jujuba stir-fried with C. chinensis juice.

Objective To establish primary and simian virus 40(SV40)T antigen transformed human vascular en-dothelial cell models,and provide available resources for endothelial research.Methods Human umbilical vein endothelial cells(HUVEC),human umbilical artery endothelial cells(HUAEC),great saphenous vein endothelial cells(GSVEC)and endothelial cells form endometrium and liver tissue were isolated and cultured respectively.Then,the primary endothelial cells were transformed by lentivirus containing SV40 big T and small T antigens,and continuously subcultured in vitro.The expression of CD31 was detected by flow cytometry,species identification-and mycoplasma detection by PCR,and cell identity was identified by STR detection.The transformed ECs were checked for HLA types.Some of them were tested for RNA expression profile and infected by Cas9 lentivirus to es-tablish stable clones.Results Totally 187 cell lines of transformed HUVEC,1 of transformed HUAEC,5 of trans-formed GSVEC,1 of transformed endothelial cells from endometrium and 1 of transformed endothelial cells from liv-er tissue,and 9 monoclonal HUVEC cell lines stably expressing Cas9 protein were established.All the transformed umbilical endothelial cells were CD31 positive ranging from 20%-90%for 20 cases,while for the rest 168 cases the positive rate was more than 90%.RNA expression revealed stable activation of cell proliferation(cell cycle and DNA synthesis).Their species were identified as human origin.The STR results were consistent with those of the primary culture and unique,and there was no mycoplasma contamination.All these cells could be obtained with the sharing services of National Science and Technology Infrastructure,the National Biomedical Cell-line Resource cen-ters(NSTI-BMCR).Conclusions A series of primary and SV40 T antigen transformed human vascular endothelial cell models have been established,which provide a tool for the study of cardiovascular diseases,inflammation,tumors and immune-related diseases.

ObjectiveBased on the AMP-activated protein kinase （AMPK）/mammalian target of rapamycin （mTOR） signaling pathway， this study aimed to observe the effect of the Huazhuo Jiedu prescription on renal fibrosis in 5/6 nephrectomy rats and explore its underlying mechanism. MethodsA total of 67 SPF-grade male SD rats were used， of which 11 were randomly selected as the normal group. A chronic renal failure （CRF） model was established using 5/6 nephrectomy. The successfully modeled rats were randomly assigned to the model group， losartan potassium group （4.5 mg·kg^-1）， and low- （1.175 g·kg^-1）， medium- （2.35 g·kg^-1） and high-dose （4.7 g·kg^-1） Huazhuo Jiedu prescription groups， with 9 rats per group. Each group received an equivalent volume of saline or the corresponding concentration of Huazhuo Jiedu prescription by gavage once daily for 8 weeks. Hematoxylin-eosin （HE） and Masson staining were used to observe renal tissue pathological changes. Transmission electron microscopy examined renal ultrastructure. Immunohistochemistry （IHC） detected expressions of α-smooth muscle actin （α-SMA） and transforming growth factor-β₁ （TGF-β₁）. Western blot analyzed expression levels of microtubule-associated protein Ⅰ light chain 3Ⅱ （LC3Ⅱ）， Beclin1， p62， AMPK， phosphorylated AMPK （p-AMPK）， mTOR， and phosphorylated mTOR （p-mTOR）. ResultsCompared with the normal group， the model group exhibited glomerular shrinkage， mesangial and interstitial thickening， and tubular vacuolar degeneration， with no evident autophagosomes or autophagolysosome structures. Expression levels of α-SMA and TGF-β₁ were significantly increased （P0.01）， while p-AMPK/AMPK， Beclin1， and LC3Ⅱ were significantly decreased （P0.01）， and p-mTOR/mTOR and p62 were significantly increased （P0.01）. Compared with the model group， the medium- and high-dose Huazhuo Jiedu prescription groups and the losartan potassium group showed varying degrees of pathological improvement. Autophagosomes with double- or multiple-layer membranes and autophagolysosomes with monolayer membranes containing undegraded organelles were observed. Renal α-SMA and TGF-β₁ protein expression levels were markedly reduced （P0.05， P0.01）， p-mTOR/mTOR and p62 were significantly decreased （P0.05， P0.01）， and p-AMPK/AMPK， Beclin1， and LC3Ⅱ expression levels were significantly increased （P0.05， P0.01）. ConclusionHuazhuo Jiedu prescription may improve renal fibrosis in 5/6 nephrectomy rats by regulating the AMPK/mTOR signaling pathway and enhancing autophagy.

Objective:To investigate the mechanism of inhibition for Janus tyrosine kinase,signal transducer and activator of transcription(JAK/STAT)on ferroptosis,myocardial remodeling,and JAK/STAT melatonin receptor 1-nuclear factor E2-related factor 2-glutathione peroxidase 4(MT1-Nrf2-GPX4)signaling axis in rats with heart failure(HF)under echocardiography.Methods:Forty SD rats were selected,and they were randomly divided into blank control group,model group,interleukin-6(IL-6)group and tyrosine phosphorylation inhibitor(AG490)group according to random number table,with 10 rats in each group.HF rat models were established in three groups except the blank control group.The expression levels of JAK/STAT mRNA in myocardial tissue among 4 groups were compared and analyzed.The small animal ultrasound imaging system was used to measure a series of cardiac function indexes included interventricular septum(IVS)thickness,left ventricular posterior wall(LVPWs)thickness,left ventricular posterior wall(LVPWd)thickness,left ventricular end-diastolic diameter(LVEDD),left ventricular end-systolic diameter(LVESD)and ejection fraction(EF).The Fe2+concentration and lesions of myocardial tissue,as well as the expressions of MT1,Nrf2 and GPX4 proteins,in the four groups were compared and analyzed.Results:The increases of JAK1 and STAT3 mRNA levels,and Fe2+concentration in the model group,IL-6 group and AG490 group were higher than those in the blank control group,and the protein expression levels of MT1,Nrf2 and GPX4 in them were lower than those in the blank control group,and the differences of the above indicators between the model group and the blank control group were statistically significant(t=11.810,16.250,17.150,10.460,9.272,11.180,P＜0.05),and the differences of the above indicators between the IL-6 group and the blank control group were statistically significant(t=9.834,19.030,16.320,14.450,13.250,14.070,P＜0.05),and the differences of them between the AG490 group and the blank control group were statistically significant(t=8.025,11.050,17.590,4.173,4.179,4.183,P＜0.05),respectively.The levels of STAT3 mRNA level and Fe2+concentration in IL-6 group were higher than those in the model group,and the expression levels of MT1,Nrf2 and GPX4 proteins in that were lower than those in the model group,and the differences were statistically significant(t=2.224,2.582,3.550,3.078,2.624,P＜0.05),respectively.The levels of JAK1,STAT3 mRNA and Fe2+concentration in the AG490 group were lower than those in the model group,and the levels of MT1,Nrf2 and GPX4 in that were higher than those in the model group,and the differences were statistically significant(t=4.052,6.930,8.598,6.247,5.055,6.799,P＜0.05),respectively.The IVS,LVPWs,LVPWd and EF of the model group,IL-6 group and AG490 group were lower than those of the blank control group,and the LVEDD and LVESD of them were higher than those of the blank control group,and the differences of these indicators between the model group and the blank control group were statistically significant(t=13.740,11.100,5.654,7.049,13.440,14.260,P＜0.05),and the differences of them between the IL-6 group and the blank control group were statistically significant(t=16.090,13.140,8.103,9.174,15.940,17.010,P＜0.05),and the differences of them between the AG490 group and the blank control group were statistically significant(t=5.474,4.947,2.682,4.071,8.608,13.300,P＜0.05),respectively.The LVPWd and EF in the IL-6 group were lower than those in the model group,and the LVEDD and LVESD in the IL-6 group were higher than those in the model group,and the differences of them were statistically significant(t=2.417,2.327,2.236,2.818,P＜0.05),respectively.The IVS,LVPWs,LVPWd and EF in the AG490 group were higher than those in the model group,and the LVEDD and LVESD in that were lower than those in the model group,and the differences were statistically significant(t=8.598,7.147,3.514,3.361,4.914,2.970,P＜0.05),respectively.Conclusion:Inhibition of JAK/STAT expression can improve the cardiac structure of HF rats,and relieve ventricular remodeling,and inhibit ferroptosis of cardiomyocyte,and promote the expression of MT1,Nrf2 and GPX4 proteins.

AIM To establish the preparation process and quality standard for Zhenggu Pills.METHODS With decoction time,decoction frequency and water addition as influencing factors,comprehensive score for extract yield and transfer rates of epicatechin and naringin as an evaluation index,the decoction process was optimized by orthogonal test.With sugarless paste relative density,medicinal powder fineness,sugarless paste-corn starch ratio,drying temperature and drying time as influencing factors,soft material traits,pill formability,moisture and disintegration time limit as evaluation indices,the formability process was optimized by single factor test.TLC was adopted in the qualitative identification of Dipsaci Radix,salt-processed Psoraleae Fructus,cooked Rhei Radix et Rhizoma and Notoginseng Radix et Rhizoma.HPLC was used for the content determination of paeoniflorin and naringin.RESULTS The optimal decoction process was determined to be 0.5 h for decoction time,two times for decoction frequency,and 10 times for water addition,the comprehensive score was 0.93.The optimal formability process was determined to be 1.21-1.22 for sugarless paste relative density,80 mesh for medicinal powder fineness,1∶0.17-1∶0.18 for sugarless paste-corn starch ratio,70 ℃ for drying temperature,and 24 h for drying time,good soft material traits and pill formability were observable,and moisture and disintegration time limit accored with 2020 edition of Chinese Pharmacopoeia requirements.The TLC spots were clear without negative interference.Two constituents showed good linear relationships within 61.30-490.41 μg/mL(r=0.999 8)and 3.27-26.18 μg/mL(r=0.999 8),whose average recoveries were 100.15％and 98.15％with the RSDs of 0.55％and 2.30％,respectively.CONCLUSION This stable,reliable and specific method can be used for the production and quality evaluation of Zhenggu Pills.

AIM To establish the quantitative models for gallic acid,mononucleoside,loganin,resveratrol,and rhein in Yishen Xiezhuo Mixture.METHODS HPLC was adopted in the content determination of various effective components,after which the near-infrared spectroscopy(NIRS)data were collected in 128 batches of samples and pretreatment was conducted,competitive adaptive reweighting sampling(CARS)algorithm was used for screening wavelength,partial least square method(PLS)regression analysis was performed.RESULTS There were no significant differences between the predicted values obtained by PLS models and measured values obtained by HPLC for various effective components(P＞0.05).CONCLUSION The quantitative models established by NIRS combined with chemometrics display good predictive performance,which can be used for the rapid determination of effective components in Yishen Xiezhuo Mixture,and provide a reference for the rapid monitoring of other traditional Chinese medicine preparations in production processes.

BACKGROUND: The clinical impact of post-percutaneous coronary intervention (PCI) quantitative flow ratio (QFR) in patients treated with PCI for chronic total occlusion (CTO) was still undetermined. METHODS: All CTO vessels treated with successful anatomical PCI in patients from PANDA III trial were retrospectively measured for post-PCI QFR. The primary outcome was 2-year vessel-oriented composite endpoints (VOCEs, composite of target vessel-related cardiac death, target vessel-related myocardial infarction, and ischemia-driven target vessel revascularization). Receiver operator characteristic curve analysis was conducted to identify optimal cutoff value of post-PCI QFR for predicting the 2-year VOCEs, and all vessels were stratified by this optimal cutoff value. Cox proportional hazards models were employed to calculate the hazard ratio (HR) with 95% CI. RESULTS: Among 428 CTO vessels treated with PCI, 353 vessels (82.5%) were analyzable for post-PCI QFR. 31 VOCEs (8.7%) occurred at 2 years. Mean value of post-PCI QFR was 0.92 ± 0.13. Receiver operator characteristic curve analysis shown the optimal cutoff value of post-PCI QFR for predicting 2-year VOCEs was 0.91. The incidence of 2-year VOCEs in the vessel with post-PCI QFR < 0.91 (n = 91) was significantly higher compared with the vessels with post-PCI QFR ≥ 0.91 (n = 262) (22.0% vs. 4.2%, HR = 4.98, 95% CI: 2.32-10.70). CONCLUSIONS Higher post-PCI QFR values were associated with improved prognosis in the PCI practice for coronary CTO. Achieving functionally optimal PCI results (post-PCI QFR value ≥ 0.91) tends to get better prognosis for patients with CTO lesions.

Objective To identify disulfidptosis-related gene(DRG)in acute myocardial infarction(AMI)by bioinformatics,analyze the molecular pattern of DRGs in AMI,and construct a DRGs-related prediction model.Methods AMI-related datasets were downloaded from the Gene Expression Omnibus database,and DRGs with differential expression were screened in AMI.CIBERSORT method was used to analyze the immune infiltration.Based on the differentially expressed DRGs,the AMI patients were classified into distinct subtypes via consensus clustering,followed by immune infiltration analysis,differential expression analysis,gene ontology and Kyoto encyclopedia of genes and genomes enrichment analysis,and gene set variation analysis.Weighted gene co-expression network analysis(WGCNA)was then performed to construct subtype-associated modules and identify hub genes.Finally,least absolute shrinkage and selection operator,random forest,and support vector machine-recursive feature elimination were used to screen feature genes to construct a DRGs-related prediction model.The model's diagnostic efficacy was evaluated by nomogram and receiver operating characteristic(ROC)curve analysis,followed by external validation.Results Nine differentially expressed DRGs were identified between AMI patients and controls.Based on the expression levels of these nine DRGs,AMI patients were divided into two DRGs subtypes,C1 and C2.Increased infiltration of monocytes,M0 macrophages,and neutrophils was observed in AMI patients and C1 subtype(all P<0.05),indicating a close correlation between DRGs and immune cells.There were 257 differentially expressed genes between the C1 and C2 subtypes,which were related to biological processes such as myeloid leukocyte activation and positive regulation of cytokines.Fcγ receptor-mediated phagocytosis and NOD-like receptor signaling pathway activity were enhanced in C1 subtype.WGCNA analysis suggested that the brown module exhibited the strongest correlation with DRG subtypes(r=0.67),from which 23 differentially expressed genes were identified.The feature genes screened by three machine learning methods were interpolated to obtain a DRGs-related prediction model consisting of three genes(AQP9,F5 and PYGL).Nomogram and ROC curves(AUC_train=0.891,AUC_test=0.840)showed good diagnostic efficacy.Conclusions DRGs were closely related to the occurrence and progression of AMI.The DRGs-related prediction model consisting of AQP9,F5 and PYGL may provide targets for the diagnosis and personalized treatment of AMI.
Humans ; Myocardial Infarction/diagnosis* ; Transcriptome ; Computational Biology ; Gene Expression Profiling ; ROC Curve ; Gene Regulatory Networks ; Nomograms ; Disulfidptosis

Objective To investigate the epidemiological characteristics of common respiratory viruses and atypical pathogens in late-term neonates during the pandemic period of Coronavirus Dis-ease 2019(COVID-19)and the impact of non-pharmaceutical interventions(NPIs)on neonatal re-spiratory pathogens.Methods A total of 2,452 late-term hospitalized neonates caused by respirato-ry infection symptoms at the Children's Hospital of Soochow University from January 2018 to February 2023 were included in the study.Data from January 2018 to December 2019 were used as pre-pan-demic control group,and data from January 2020 to December 2021 were used as post-pandemic con-trol group.The changes in nucleic acid test results for eight common pathogens in hospitalized chil-dren during the pandemic period of COVID-19 were retrospectively analyzed.Data from March 2020 to February 2023 were compared to summarize the epidemiological characteristics of neonatal respira-tory pathogens in different seasons after the normalization of the pandemic period of COVID-19.Results Among the 2,452 hospitalized neonates,364(14.85％)tested positive for pathogens,with respiratory syncytial virus(RSV)having the highest detection rate of 262 cases(10.69％),followed by parainfluenza viruses(PIVS)with 64 cases(2.61％).The total number of positive cases for the eight pathogens before and after the pandemic control measures were 229(17.1％)and 96(12.3％)respectively.The detection rate after the pandemic control measures was significantly lower than be-fore(P＜0.05).The detection rates of all eight pathogens decreased after the pandemic control measures,but only the difference in the RSV detection rate was statistically significant(P＜0.05).The peak month for RSV-positive cases was delayed by one month after the pandemic control measures compared to pandemic control before,and there were two peaks in RSV incidence in 2022.PIVS was more prevalent in spring,summer,and autumn before the pandemic control measures,but became more prevalent in winter afterward,with a higher peak in positive cases detected after the control measures than pandemic control before;the seasonality of PIVS infection changed.Conclusion After the implementation of pandemic control measures for COVID-19,the detection rate and epidemiology of common respiratory pathogens in late-term neonates with respiratory infections in Suzhou have changed.NPIs such as wearing masks,hand hygiene,maintaining social distance,and avoiding un-necessary outings have a certain preventive effect on the outbreak of respiratory pathogens.