OBJECTIVE To investigate the intervention effect and potential mechanism of breviscapine on hepatic fibrosis （HF） in rats based on the transforming growth factor-β（1 TGF-β1）/Smad2/extracellular signal-regulated protein kinase 1（ERK1） and Kelch-like epichlorohydrin-associated protein 1（Keap1）/nuclear factor-erythroid 2-related factor 2（Nrf2）/heme oxygenase-1（HO-1） pathways. METHODS Totally 60 rats were randomly divided into normal control group， model group， breviscapine low-dose， medium-dose and high-dose groups （5.4， 10.8， 21.6 mg/kg）， and colchicine group （positive control， 0.45 mg/kg）， with 10 rats in each group， half male and half female. Except for the normal control group， HF model of the other groups was induced by carbon tetrachloride. Subsequently， each drug group was given corresponding medicine by gavage once a day for 28 days. The liver appearance of rats in each group was observed and their liver coefficients were calculated. The levels of alanineaminotransferase （ALT） and aspartate aminotransferase （AST）in serum， those of ALT， AST， superoxide dismutase （SOD），malondialdehyde （MDA） and glutathione peroxidase （GSH- Px） in liver tissue were detected. The liver tissue inflammatory and fibrotic changes were observed. The protein and mRNA expressions of TGF-β1， Smad2， ERK1， Nrf2， Keap1 and HO-in liver tissue were detected. RESULTS Compared with the normal control group， the model group showed large areas of white nodular lesions in the liver， obvious inflammatory cell infiltration and collagen fiber deposition. The body weight， the levels of SOD and GSH-Px in liver tissue， the protein and mRNA expressions of Nrf2 and HO-1 were significantly lowered in the model group （P＜0.05）； the liver coefficient， the percentage of Masson staining positive area， ALT and AST levels of serum and liver tissue， MDA level of liver tissue， the protein and mRNA expressions of TGF-β1， Smad2， ERK1 and Keap1 were significantly increased （P＜0.05）. Compared with the model group， the liver lesions of rats in each drug group were improved， and the above quantitative indexes were generally reversed （P＜0.05）. CONCLUSIONS Breviscapine has a good intervention effect on HF rats， which may be related to inhibiting TGF-β1/Smad2/ERK1 pathway for anti-fibrosis and regulating Keap1/Nrf2/HO-1 pathway to inhibit oxidative stress.

Objective:To observe of the effects of micro-video breakthrough education and thematic workshops in the care of internal fistulae in maintenance haemodialysis patients, to provide a basis for improving prognosis of patients.Methods:This study was a randomized controlled trial. A total of 90 cases of maintenance haemodialysis patients with endocardial fistula were selected in Huzhou Hospital of Traditional Chinese Medicine Affiliated to Zhejiang University of Traditional Chinese Medicine from January to December in 2022 as the observation samples using convenience sampling method. They were divided into the experimental group and the control group with 45 cases in each group by the method of random number table. The control group used the conventional nursing care for maintenance haemodialysis endocardial fistula to administer nursing care, the experimental group introduced the micro-video breakthrough education and thematic workshop as a combined mechanism of intervention. The self-care behaviors with arteriovenous fistula and other observation indexes between the two groups were compared.Results:There were 23 males and 22 females in the control group, aged (58.07 ± 3.00) years old. There were 24 males and 21 females in the experimental group, aged (58.93 ± 4.07) years old. After care, the total internal fistula self-care behaviour score in the experimental group was (49.44 ± 1.84) points, and (38.93 ± 2.19) points in the control group, the difference between the two groups were statistical significant ( t = 24.65, P<0.05). For health management adherence, the score of fluid intake, dialysis regimen, medication and dietary dimension were (15.82 ± 1.37), (12.87 ± 1.39), (17.24 ± 1.33) and (21.60 ± 1.03) points in the experimental group, and (11.20 ± 0.92), (9.98 ± 1.14), (12.11 ± 1.01) and (17.40 ± 1.10) points in the control group, the difference between the two groups were statistical significant ( t values were 10.78 to 20.62, all P<0.05). For positive psychological capital, the scores of self-efficacy, resilience, hope and optimism were (38.69 ± 1.22), (39.27 ± 1.10), (33.29 ± 1.52) and (33.40 ± 1.39) points, and were (31.16 ± 1.26), (31.04 ± 1.15), ( 26.13 ± 1.52) and (27.09 ± 7.28) points in the control group, the difference between the two groups were statistical significant ( t values were 5.71 to 34.77, all P<0.05). The total complication rate of the experimental group was 8.89% (4/45), lower than 28.89% (13/45) in the control group, and the differences were all statistically significant ( χ2 = 5.87, P<0.05). Conclusions:The combined application of micro-video breakthrough education and special workshops in maintenance hemodialysis patients with internal fistula is beneficial to the improvement of their internal fistula self-care ability, health management compliance and positive psychological capital, and can reduce the probability of internal fistula complications.

Objective To investigate the reversal effect and mechanism of cinobufagin(CBG)on cisplatin resist-ance in human ovarian cancer cells.Methods A2780 cell line and its cisplatin-resistant cell line A2780/DDP are common ovarian cancer cells in clinic,so these two cell lines were selected as the research objects.The cell viabil-ity was detected by cell Counting Kit-8(CCK-8)assay,and the cell proliferation ability was detected by plate clo-ning and 5-ethynyl-2′-deoxyuridine(EdU)assay.Hoechst staining was used to observe cell apoptosis.Cell scratch test and Transwell test were used to evaluate cell migration and invasion ability.Western blot and quantitative reverse transcription PCR(RT-qPCR)were used to detect the protein and mRNA expressions of phosphatidylinosi-tol 3-kinase/protein kinase(PI3K/AKT)signaling pathway and epithelial-mesenchymal transition(EMT).Re-sults Compared with A2780 cells,the drug resistance indexes of A2780/DDP cells were 5.636,5.864,5.695,respectively.After treatment of A2780/DDP cells with CBG(2,4,6 mg/ml),the reversal resistance indexes were 1.617,2.570,3.461,respectively.CBG treatment significantly increased the level of apoptosis and inhibi-ted the proliferation,migration and invasion of the cells in a concentration-dependent manner(P<0.05).Western blot results showed that compared with A2780 cells,the relative ratio of P-PI3K/PI3K and P-AKT/AKT protein levels,as well as the protein expression of N-cadherin,Vimentin,and Snail were higher in the control group(A2780/DDP)cells,while the protein expression of E-cadherin was lower(tP-PI3K/PI3K=8.115,tP-AKT/AKT=17.62,tN-cadherin=6.126,tVimentin=4.001,tSnail=17.333,tE-cadherin=4.620,P<0.01);As the dose of CBG increased,the protein expression levels of P-PI3K,P-AKT,N-cadherin,Vimentin,and Snail in drug-resistant cells de-creased,while the protein expression level of E-cadherin increased(FP-PI3K=268.5,FP-AKT=190.5,FN-cadherin=24.02,FVimentin=57.65,FSnail=87.24,FE-cadherin=135.8,P<0.05).qRT-PCR results showed that with the in-crease of CBG concentration,the mRNA expression levels of PI3K,AKT,N-cadherin,Vimentin and Snail de-creased,while the mRNA expression level of E-cadherin gradually increased(FPI3K=101.1,FAKT=558.3,FN-cadherin=86.97,FVimentin=105.9,FSnail=85.71,FE-cadherin=80.96,P<0.01).Conclusion CBG can reverse cisplatin resistance of ovarian cancer A2780/DDP cell line,and its mechanism may be related to the regulation of PI3K/AKT signaling pathway and inhibition of EMT by CBG.

The efficient clinical treatment of oral squamous cell carcinoma(OSCC)is still a challenge that demands the development of effective new drugs.Phenformin has been shown to produce more potent anti-tumor activities than metformin on different tumors,however,not much is known about the influence of phenformin on OSCC cells.We found that phenformin suppresses OSCC cell proliferation,and promotes OSCC cell autophagy and apoptosis to significantly inhibit OSCC cell growth both in vivo and in vitro.RNA-seq analysis revealed that autophagy pathways were the main targets of phenformin and identified two new targets DDIT4(DNA damage inducible transcript 4)and NIBAN1(niban apoptosis regulator 1).We found that phenformin significantly induces the expression of both DDIT4 and NIBAN1 to promote OSCC autophagy.Further,the enhanced expression of DDIT4 and NIBAN1 elicited by phenformin was not blocked by the knockdown of AMPK but was suppressed by the knockdown of transcription factor ATF4(activation transcription factor 4),which was induced by phenformin treatment in OSCC cells.Mechanistically,these results revealed that phenformin triggers endoplasmic reticulum(ER)stress to activate PERK(protein kinase R-like ER kinase),which phosphorylates the transitional initial factor eIF2,and the increased phosphorylation of eIF2 leads to the increased translation of ATF4.In summary,we discovered that phenformin induces its new targets DDIT4 and especially NIBAN1 to promote autophagic and apoptotic cell death to suppress OSCC cell growth.Our study supports the potential clinical utility of phenformin for OSCC treatment in the future.

AIM:To investigate the effects of Shaoyao-Gancao decoction(SGD)on inflammation and mucosal barrier damage in rats with 2,4,6-trinitrobenzenesulfonic acid(TNBS)-induced inflammatory bowel disease(IBD).METHODS:Forty-eight male SD rats were randomly divided into normal group,model group,high-dose(500 mg/kg),medium-dose(250 mg/kg)and low-dose(125 mg/kg)SGD groups,and balsalazide sodium(1 g/kg)group.All rats were pre-administered for 3 d,and on the 4th day of the experiment,they were fasted for 24 h.Except for the normal group,the rats in the other 5 groups were given enema mixed with TNBS(100 mg/kg)and 50％ethanol,and continued to be adminis-tered for 5 d after modeling.After modeling,the disease activity index(DAI)was evaluated.After the experiment,the levels of nitric oxide(NO)and myeloperoxidase(MPO)in serum and colonic tissues of rats were determined.RT-qPCR and Western blot were used to determine tumor necrosis factor-α(TNF-α),cyclooxygenase-2(COX-2),inducible nitric oxide synthase(iNOS)and nuclear factor-κB(NF-κB)in the colon of rats.The expression of tight junction proteins zonu-la occludens-1(ZO-1)and claudin 2 in rat colon tissues was determined by immunofluorescence staining.RESULTS:Compared with normal group,the weight of rats in model group was decreased,the colon was shortened,DAI and colon tissue macroscopic scores were significantly increased(P<0.05),colon pathological injury was serious,and NO and MPO levels in serum and colon tissues of the rats in model group were significantly increased(P<0.05).The mRNA and pro-tein expression levels of TNF-α,COX-2,iNOS and NF-κB in colon tissues were significantly increased(P<0.01),while the expression levels of ZO-1 and claudin 2 were significantly decreased(P<0.01).Compared with model group,the body weight and colon shortening of rats in SGD groups were alleviated,DAI and macroscopic scores of colon tissues were significantly decreased(P<0.05),the pathological damage of colon was improved,and the levels of NO and MPO in se-rum and colon tissues of rats were significantly decreased(P<0.05).The mRNA and protein expression levels of TNF-α,COX-2,iNOS and NF-κB in colon tissues were significantly decreased(P<0.05),while the expression levels of ZO-1 and claudin 2 were significantly increased(P<0.05).CONCLUSION:Treatment with SGD effectively attenuates the inflam-matory response and intestinal mucosal barrier damage caused by TNBS-induced IBD in rats.

Objective To observe the impact of ultrasonic image quality on the consistency of artificial intelligence(Al)assisted diagnosis system and manual measurements of biological indicators of developmental dysplasia of hip(DDH).Methods Hip ultrasonic data of 75 DDH and 345 non-DDH children were retrospectively analyzed,and the quality of ultrasonic images were subjectively scored.An evaluation model of ultrasonic image quality was constructed based on 140 ultrasonic images acquired from 140 cases(group A,containing 25 DDH and 115 non-DDH)using entropy weighting method,the weight of anatomic structures and impact factors related to DDH were obtained.The comprehensive image quality scores of other ultrasonic images acquired from 280 cases(group B,including 50 DDH and 230 non-DDH)were calculated,and the images in group B were classified into grade A,B and C in descending order.The consistency of AI and manual measurements of DDH biological indicators in group B was assessed.Results The weight of each anatomic structure and impact factors of DDH obtained with the model were as follows:The lower edge of iliac branch＞ilium＞glenoid labrum＞bony margin＞femoral head＞motion artifacts.In group B,grade A was observed in 67(9 DDH and 58 non-DDH),grade B was found in 160(26 DDH and 134 non-DDH),while grade C was noticed in 53(15 DDH and 38 non DDH)images.Except for β,femoral head coverage(FHC)and femoral head length diameter,the consistencies between AI and manual measurements of other indicators of DDH were grade A＞B＞C.In group B,AI and manual measurements were more consistent in DDH than in non-DDH cases.Conclusion Ultrasonic image quality affected the consistency between AI and manual measurements of biological indicators of DDH.When image quality was not good enough,further attention should be paid to measurement of FHC and sizes of femoral head.

Objective Based on the limited time theory,taking tonsillectomy day operation as an example,the whole process nursing list of day operation was constructed,and its application effect was evaluated.Methods The nursing service list of tonsillectomy was constructed based on literature research,field observation and expert correspondence.A total of 197 patients who underwent tonsillectomy in the day ward of a tertiary A general hospital in Hangzhou,Zhejiang Province were selected.Among them,98 patients from April to June 2023 were taken as an experimental group and treated with the whole-process nursing list of daytime tonsillectomy constructed in this study,and 99 patients from January to March 2023 were taken as a control group and routine nursing was adopted.The anxiety degree,postoperative pain score,postoperative complication rate and patients'satisfaction with nursing service were compared between the 2 groups.Results Anxiety scores before operation and at discharge,pain scores at 6 h,12 h and 24 h after operation,and incidence of complications such as infection and gastrointestinal reaction within 14 days after operation in the experimental group were lower than those in the control group,with statistical significance(P＜0.05).The satisfaction of nursing service in the experimental group(99.40±0.69)score was significantly higher than that in the control group(90.36±1.04)score,and the difference was statistically significant(P＜0.001).Conclusion The use of tonsillectomy day whole-process nursing list can improve perioperative anxiety,relieve postoperative pain and reduce the incidence of postoperative complications,and improve patient satisfaction.

Objective:To establish a multiplex detection method for monitoring viruses post-kidney transplantation based on droplet digital PCR (ddPCR) technology, evaluate its detection performance, and discuss its potential clinical application.Methods:We developed a ddPCR assay for the simultaneous detection of polyomavirus type 1 virus (BKV), polyomavirus type 2 virus (JCV), and torque teno virus (TTV), assessing its conformity rate, repeatability, and detection limit. 69 clinical urine samples were collected at the Shanghai Jiao Tong University School of Medicine affiliated with Renji Hospital between June and September 2023. The qPCR and ddPCR methods were employed to analyze the results, respectively. Spearman correlation analysis, Bland-Altman analysis, and Wilcoxon paired rank sum test were applied to analyze the detection results of the two methodologies.Results:The constructed ddPCR method had a 7/7 concordance rate for the seven reference samples. The precision reference samples were tested ten times, respectively, and their coefficients of variation were less than 10%. The detection limit was 10 copies/μl. The concordance rates of ddPCR and qPCR for BKV and JCV detection were more than 95% (66/69, 68/69). The correlation coefficients (R) of BKV and JCV viral loads were 0.874 and 0.840, respectively. The Bland-Altman analysis showed that the mean differences in BKV and JCV viral load detection between the two methods were -0.34 and -0.187, respectively. The Wilcoxon paired rank sum test showed that there were no statistically significant differences in the detection results of polyomaviruses between the two methods ( P>0.05). Conclusion:A ddPCR-based multiplex detection method for viruses post-renal transplantation was successfully developed, confirming its superior attributes of high accuracy, high sensitivity, and multi-detection capability. This method could be used for the quantitative detection of BKV, JCV, and TTV in clinical urine samples.

Objective:To investigate the effects of sclerostin (SOST) and WNT/CTNNB1 signaling pathway on the cell cycle, migration and invasion of human uveal melanoma (UM) cells and its related mechanism.Methods:UM tissues from 20 cases of epithelioid UM and 16 cases of spindle cell type UM were collected.The contents of SOST, Wnt-1 and Catenin beta-1 proteins in the collected tissues were detected by immunohistochemical staining.Three human UM tissue derived cell lines OCM-1 (primary spindle cell type), Mum-2B (metastatic epithelioid) and Mum-2C (metastatic spindle cell type) were selected and divided into three groups, blank control group not transfected, empty vector group transfected with SOST negative control vector and SOST siRNA group transfected with SOST siRNA.After 24-hour transfection, the mRNA and protein expression levels of SOST, CTNNB1, WNT protein family 1 (WNT1), CCND1, matrix metalloproteinase (MMP)2 and MMP9 were detected by real-time fluorescence quantitative PCR and Western blot, respectively.The invasion and migration ability of the transfected cells were measured by transwell method, and the cell cycle distribution was detected by flow cytometry.Another 9 female BALB/c nude mice were selected and randomized into OCM-1 group, OCM-1 empty vector group and SOST shRNA group, inoculated with OCM-1 without lentivirus infection, OCM-1 with blank lentivirus infection and OCM-1 with SOST shRNA lentivirus infection, respectively.Six weeks after inoculation, the in situ formation of tumor was observed.The interaction between SOST and low density lipoprotein receptor related protein(LRP)-5/6 in OCM-1 cells was explored by co-immunoprecipitation assay.The study protocol was approved by the Ethics Committee of Tianjin Medical University Eye Hospital (2018KY[L]-20).Results:Immunohistochemical staining results showed that the SOST expression level was higher and the expression levels of Wnt-1 and Catenin beta-1 were lower in spindle cell type UM tissues than in epithelioid UM tissues, and the differences were all statistically significant (all at P<0.01). The real-time fluorescence quantitative PCR results showed that the relative expression of SOST mRNA was significantly lower and the relative expressions of CCND1, WNT1 and MMP9 mRNA were significantly higher in SOST siRNA groups than in corresponding empty vector groups in the three cell lines (all at P<0.05). In OCM-1 and Mum-2C cell lines, the relative expressions of CTNNB1 mRNA were significantly higher in SOST siRNA groups than in empty vector groups (all at P<0.01). Western blot results showed that the relative expression of SOST protein was significantly lower and the relative expressions of Wnt-1, Catenin beta-1, cyclin-D1, MMP2 and MMP9 proteins were significantly higher in SOST siRNA groups than in empty vector groups (all at P<0.01). Transwell assay showed that the cell invasion and migration ability of SOST siRNA group was significantly higher than that of blank control group and empty vector group in the three cell lines (all at P<0.01). Flow cytometry showed that the proportion of G1-phase cells and the G1/S-phase ratio were significantly lower in SOST siRNA group than in blank control groups and empty vector groups (all at P<0.01). The eyeball volume of OCM-1 group, OCM-1 empty vector group and SOST shRNA group was (42.7±4.6), (49.0±22.9) and (135.2±32.7)mm 3, respectively, showing a significant overall difference ( F=19.963, P<0.01). The eyeball volume of SOST shRNA group was larger than that of OCM-1 group and OCM-1 empty vector group, and the differences were statistically significant (both at P<0.05). Co-immunoprecipitation results showed that SOST could interact with LRP-5 and LRP-6 by binding to them, respectively. Conclusions:Silencing SOST can promote the invasion and migration of UM cells, and increase the proportion of UM cells in the division phase.Silencing SOST can promote tumor growth in eyes of nude mice.SOST may play this function by interacting with the membrane receptor LRP-5/LRP-6 and then regulating the WNT/CTNNB1 signal pathway.

Purpose To investigate the clinicopathological characteristics,diagnosis,and differential diagnosis of invasive mucinous adenocarcinoma(IMA)and mixed invasive mucinous and non-mucinous adenocarcinoma(mIMA).Methods The clinical data were collected in 36 patients with primary IMA and 17 patients with mIMA,and the expression of TTF-1,CK7,CK20,SATB2,CDX2,EGFR,HNF4a,etc.was detected by immunohistochemical EnVision two-step method.The Sanger se-quencing and the FISH were used for KRAS mutation and NRG1 gene rearrangement detection.The clinicopathological character-istics were analyzed with review of relevant literature.Results There were 9 cases(25.0％)and 3(8.3％)cases of papillary and micropapillary structures in IMA,while 13 cases(76.5％)(P＜0.001)and 9 cases(52.9％)(P=0.001)were present in mIMA.There were 5 cases(13.9％)of high nuclear grade of IMA and 10 cases(58.8％)of high nuclear grade of mIMA(P=0.002).TTF-1 had a positive rate of 37.5％in IMA,but 60.0％and 80.0％in the mucinous adenocarcinoma and non-mucinous adenocarcinoma components of mIMA(P=0.021),respectively.The positive rates of CK7,CK20,and CDX2 in IMA were 90.6％,21.9％,and 9.4％,and the positive rates in mucinous adenocarcinoma and non-mucinous adenocarcinoma components of mIMA were 100％,20％,20％and 100％,6.7％,6.7％,respectively and no SATB2 expression was found in all cases.There was no significant difference in the expres-sion of total EGFR and two EGFR mutation-specific antibodies(L858R,DEL19)between IMA and mIMA.There were 3 cases of mucinous adenocarcinoma with L858R positive in mIMA,and 2 of them were negative for non-mueinous adenocarcinoma.In another case,the non-mueinous adenocarcinoma component of mIMA expressed DEL19,but the mucinous adenocarcinoma component was not expressed.The positive rate of HNF4a in IMA was 72.0％(18/25),and those of HNF4a in mucinous adenocarcinoma and non-mucinous adenocarcinoma in mIMA were 41.7％(5/12)and 33.3％(4/12),respectively(P=0.048).KRAS gene sequencing was carried out in 19 cases of IMA,among which 9 cases(47.4％)had mutations,G12D and G12V were most commonly detected,and 4 cases of mIMA were sequenced,but none of them showed KRAS mutations.FISH detection showed that 2 cases(7.1％)IMAs had NRG1 translocation rearrangement.Conclusion Pulmonary mIMA is more aggressive than IMA.For example,mIMA has significantly more papillary structure,micropapillary structure,and high nu-clear grade cases than IMA.The differences in immunohisto-chemical expression and KRAS mutation between the two are sta-tistically significant.