Objective To investigate the in vitro inhibitory effects of regulatory T cells ( Treg ) from unpregnant mice and pregnancy-induced regulatory T cells ( piTreg) on the proliferation of na?ve T cells and their differences .Methods The numbers of piTreg cells from allogeneic pregnant mice ( C57/B6 fe-male×BALB/c male) on day 12.5 (E12.5d) of gestation and Treg cells from unpregnant C57/B6 mice were detected respectively by flow cytometry .The percentages of piTreg cells and Treg cells in CD 4+T cells of age-matched female mice and their intracellular expression of Foxp 3 were analyzed .The in vitro inhibitory effects of piTreg cells and Treg cells on the CFSE-labeled na?ve T cells ( effector cells ) were compared in a one-way mixed lymphocyte culture system using mitomycin C-inactivated CD4-T cells as stimulator cells . Results The level of piTreg cells in splenic mononuclear cells was significantly higher than that of Treg cells (P<0.001) from normal mice.Foxp3 was highly expressed in both piTreg cells and Treg cells , howev-er slightly increased in piTreg cells .Moreover , piTreg cells had a significant stronger in vitro inhibitory effect on na?ve T cells proliferation than that of Tregs cells (P<0.006), which was in a cell-dependent manner. Conclusion The present study suggests that the piTreg cells have a stronger inhibitory effect on na ?ve T cell proliferation as compared with Terg cells from unpregnant mice , The differential activity of CD 4+CD25+Treg might be mediated by the paternal antigens during pregnancy .

Objective To investigate the protection effects and possible mechanism of apoptotic effect of compound Xiancao granules on renal ischemia reperfusion injury ( IRI ) in rats. Methods Renal IRI rat model was established by clipping bilateral renal artery.The rats were divided into model control group (n=10), compound Xiancao granules group (n=10) and sham operation group (n=10).All Serum creatinine (Cr) and blood urea nitrogen (BUN) were determined to evaluate kidney function after IRI.Superoxide dismutase (SOD), glutathione peroxidase (GSH-PX) and malondialdehyde (MDA) activity in kidney were measured by colorimetric method. Expression of apoptotic regulatory genes Bcl-2 and Bax in renal tissue were detected by Western blotting.Renal tissue sections were stained by hematoxylin and eosin (HE) and examined under a light microscope. Results SOD and GSH-PX levels of the compound Xiancao granules group (278.1±16.2),(155.96±20.58) U?mg-1 were significantly higher than those of the model control group (196.3±12.1),(109.34±17.81) U?mg-1 (P<0.05). MDA, BUN and Cr (12.49±1.07) nmol?mL-1,(8.9±2.7) mmol?L-1,(149.7±8.5) μmol?L-1 were significantly reduced in the compound Xiancao granules group as compared with those of the model control group (17.32±1.26) nmol?mL-1,(14.6± 3.3) mmol?L-1,(206.1±11.2) μmol?L-1(P<0.05).Bax and Bcl-2 mRNA expression levels of sham operation group were significantly lower, and the Bax and Bcl-2 mRNA expression levels of model control group were significantly enhanced ( Bcl-2:P<0. 05, Bax: P<0. 01 ) . Bax mRNA expression level of compound Xiancao granules group was significantly decreased as compared with that of model control group ( P<0.05) , but Bcl-2 mRNA expression level of compound Xiancao granules group was significantly enhanced as compared with that of model control group (P<0.01). Conclusion These results suggested that compound Xiancao granules has protection effect on renal IRI in rats. The mechanisms may be related to its antioxidant activity and expression of the apoptotic genes such as Bcl-2 and Bax.

Objective:To investigate the effects of tamoxifen on high glucose-induced epithelial-to-mesenchymal transition of rat peritoneal mesothelial cells and the underlying mechanism.Methods:The peritoneal mesothelial cells of normal male SD rats were selected between January 2015 and June 2016 and then cultured and divided into blank control, high-glucose stimulation and drug intervention groups. High-glucose stimulation group: primary cultured rat peritoneal mesothelial cells (RPMCs) were treated with 60 mmol/L high-concentration glucose to induce epithelial-to-mesenchymal transition. Drug intervention group: (1) RPMCs were treated with 60 mmol/L high-concentration glucose and different concentrations (0.5 μmol/L, 2 μmol/L) of tamoxifen. After 72 hours of stimulation, protein was extracted. (2) RPMCs were treated with 60 mmol/L high-concentration glucose and 2 μmol/L tamoxifen with or without 2 μmol/L ER-α antagonist for 1 hour to extract protein and for 6 hours to extract RNA. (3) RPMCs were treated with high-concentration glucose and 2 μmol/L tamoxifen with or without 1 μmol/L 1 μM proteasome inhibitor for 1 hour to extract protein. Western blot analysis was performed to analyze change in E-cadherin, α-SMA, Smad2, p-Smad2, Smad3, p-Smad3 and Smad4 protein. Real-time fluorescence quantitative PCR was performed to detect the change in mRNA expression of Smad2, Smad3, connective tissue growth factor and plasminogen activator inhibitor 1.Results:Tamoxifen attenuated epithelial-to-mesenchymal transition on RPMCs induced by high-level glucose, showing increased expression of epithelial cell marker E-cadherin and decreased expression of α-SMA in a concentration-dependent manner ( tE-cadherin = 2.31, tα-SMA =-2.53, both P < 0.05).TGF-β1/R-Smad signal pathway was activated by high-concentration glucose. Phosphorylation of Smad2/3 and mRNA expression of CTGF and PAI-1 were increased. Tamoxifen remarkably reduced protein and mRNA level of above mentioned protein and related target genes ( tp-Smad2 = -3.38, tCTGF = -3.81, P < 0.05), which could be blocked by ER-α antagonist. Finally, proteasome inhibitor could weaken the inhibitory effects of tamoxifen on p-Smad2/3 and increase p-Smad2/3 protein level ( tp-Smad2 = 3.94, P < 0.05). Conclusion:Tamoxifen activates ER-α on RPMCs, weakens the activation of TGF-β1/R-Smad signal pathway through decreasing p-Smad2 protein level, and effectively inhibits the progression of high-concentration glucose-induced epithelial-to-mesenchymal transition possibly through degrading p-Smad2 protein through proteasome. The role of tamoxifen in epithelial-to-mesenchymal transition may provide a possible guide for research, prevention and treatment of peritoneal fibrosis.

Objective To detect the expression of death-associated protein kinase like 1(DAPL1)and mismatch repair gene(Mut L homologue 1,MLH1)methylation level in bronchoalveolar lavage fluid(BALF)of patients with lung cancer,and further investigate the diagnostic value of gene methylation in early lung cancer and its relationship with clinicopathological features.Methods A total of 142 patients with suspected early stage lung cancer admitted the Second People's Hospital of Mengcheng County from January 2022 to January 2024 were retrospectively selected and divided into lung cancer group(n=82)and lung benign lesion group(n=60)according to the final pathological results.Quantitative real time polymerase chain reaction(qRT-PCR)was used to detect the methylation levels of DAPL1 and MLH1 in BALF samples.The clinical diagnostic value of DAPL1 and MLH1 methylation in early stage lung cancer and its relationship with clinicopathological features of lung cancer patients were analyzed.Results The methylation levels of DAPL1 and MLH1 gene in BALF in lung cancer group were 53.66％(44/82)and 56.10％(46/82),respectively,which were significantly higher than those in benign disease group[11.67％(7/60)and 18.33％(11/60)],and the difference was statistically significant(x2=56.544,20.565,all P＜0.05).The sensitivity of DAPL1 andMLH1 gene methylation in the diagnosis of early lung cancer was 53.66％(44/82)and 56.10％(46/82),the specificity was 88.33％(53/60)and 81.67％(49/60),and the accuracy were 68.31％(97/142)and 66.90％(95/142),respectively.The sensitivity and accuracy of DAPL1 methylation combined with MLH1 methylation in the diagnosis of early lung cancer was 86.59％(71/82),and 85.92％(122/142),respectively,both of which were higher than that of a single index(Z=24.411,16.450,all P＜0.05).The methylation levels of DAPL1 and MLH1 genes in BALF of lung cancer patients were closely correlated with clinical stage,smoking history and lymph node metastasis(x2=5.493,13.083;8.167,6.946;9.303,4.523,all P＜0.05).Spearman correlation showed that DAPL1 and MLH1 gene methylation were positively correlated with clinical stage,smoking history and lymph node metastasis in patients with lung cancer(r=0.523,0.602;0.548,0.498;0.630,0.524,all P＜0.05).Conclusion The methylation of DAPL1 and MLH1 genes in BALF has high clinical diagnostic value for early lung cancer,and the methylation levels of both genes are related to the disease progression and smoking history of lung cancer patients.

Objective:To investigate the status of frailty and activities of daily living (ADL) in maintenance hemodialysis (MHD) patients, and to explore the effect of frailty phenotype on ADL.Methods:The patients who underwent MHD in Kidney Disease Center of the First Affiliated Hospital from March 2019 to March 2020 were enrolled in this study. The demographic and laboratorial data were collected by cross-sectional survey method. Fried frailty phenotype scale and ADL scale were used to evaluate the frailty and ADL, respectively. The differences of basic data and different frailty phenotypes between the normal function group and the function decline group were compared in terms of ADL, physical self-maintenance ability and instrumental ADL ability. Pearson correlation analysis was used to analyze the correlation between frailty and ADL, and binary logistic regression analysis was used to analyze the influencing factors of ADL.Results:A total of 676 MHD patients were included in this study, including 434 males (64.2%) and 242 females (35.8%). The age was (59.2±19.4) years old, and the median dialysis age was 59.0 (25.3, 110.0) months. There were 159 frailty patients (23.5%), 230 pre-frailty patients (34.0%), and 287 non-frailty patients (42.5%). The ADL was decreased in 163 patients (24.1%), including 131 patients (19.4%) with decreased physical self-maintenance ability and 161 patients (23.8%) with decreased instrumental ADL ability. Pearson correlation analysis showed that the frailty score was positively correlated with total ADL score ( r=0.728, P<0.001), physical self-maintenance ability score ( r=0.669, P<0.001) and instrumental ADL ability score ( r=0.729, P<0.001). Binary logistic regression analysis results showed that older age and lower physical activity, fatigue, slowed steps and lower grip strength in the frailty phenotypes were the independent influencing factors of ADL, physical self-maintenance ability and instrumental ADL ability (all P<0.05). Conclusions:The prevalence of frailty is 23.5% in MHD patients, and 24.1% of MHD patients have decreased ADL. Elder age and lower physical activity, fatigue, reduced step counts, and lower grip strength in frailty phenotypes are the independent influencing factors for poor ADL, poor physical self-maintenance ability and poor instrumental ADL ability.

Objective To investigate the effects of serum uric acid (SUA) on all?cause death and cardiovascular death in patients of maintaining peritoneal dialysis (PD). Methods One thousand and sixty?three PD patients in the First Affiliated Hospital of Zhejiang University Medical College were included. The SUA levels at 6 months after PD start were measured. Patients with SUA≥420 μmol/L were grouped in hyperuricemia group (492 cases) and patients with SUA<420 μmol/L were grouped in normal uric acid group (571 cases). The effects on all ? cause mortality and cardiovascular mortality were retrospectively analyzed. Results The median age of the patients was 51(41, 62) years; 557 cases were male (52.40％); the median follow?up time was 33(20, 54) months (6?96 months); 167 cases (15.71％) died during the follow?up period, including 64 cases (6.02％) withcardiovascular causes. The mortality in hyperuricemia group was 19.11％(94/492) and the cardiovascular mortality was 7.93％(39/492), both rates were higher than those in normal uric acid group, and the differences were statistically significant (P=0.005, P=0.015, respectively). Hyperuricemia (SUA≥420μmol/L) at 6 months after PD start (HR=1.572, 95％CI 1.155-2.141, P=0.004), high uric acid level (continuous variable) at 6 months after PD start (HR=1.002, 95％CI 1.001-1.004, P=0.008), and age≥65 years (HR=3.571, 95％CI 2.556-4.990, P<0.001), serum albumin≤30 g/L (HR=1.907, 95％CI 1.278-2.845, P=0.002), high Charlson comorbidity index (HR=1.209, 95％CI 1.032-1.417, P=0.019) at the beginning of PD start were independent risk factors for all ? causes death in PD patients. Hyperuricemia (SUA≥420 μmol/L) at 6 months after PD start (HR=1.734, 95％CI 1.033-2.912, P=0.037) and age≥65 years (HR=1.761, 95％CI 1.024-3.209, P=0.041), with diabetes (HR=2.775, 95％CI 1.358-5.671, P=0.005) at the beginning of PD start were independent risk factors for cardiovascular death in PD patients. Conclusions SUA at 6 months after PD is an independent risk factor for all?cause death and cardiovascular death in PD patients.

Objective:To observe the clinical characteristics and prognosis of patients with rapidly progressive glomerulonephritis (RPGN) caused by lupus nephritis, antineutrophil cytoplasmic antibodies (ANCA) - associated vasculitis, or primary glomerulonephritis who were treated with peritoneal dialysis (PD) and then withdrew PD because of renal recovery.Methods:Data of the above patients were retrospectively analyzed. The patients were diagnosed as RPGN and received PD therapy in Kidney Disease Center, the First Affiliated Hospital, College of Medicine, Zhejiang University from February 2009 to August 2018. The patients were divided into early withdrawal group (PD time≤183 days, n=24) and late withdrawal group (PD time>183 day, n=24). The differences of clinical characteristics between the two groups were compared. The cumulative incidence of adverse events in both groups was analyzed using Kaplan-Meier curves. Cox proportional hazards model was used to analyze the risk factors influencing the prognosis of patients. Results:Forty-eight RPGN patients were included. The median time of maintaining PD was 178(76, 378) days. Compared with the late withdrawal group, the patients in early withdrawal group had lower levels of urine volume, serum albumin and parathyroid hormone, and lower rates of gross hematuria and hypertension at the beginning of PD, and received higher rates of methylprednisolone impulse, combined immunosuppressive agents, and hemodialysis or continuous renal replacement therapy (all P<0.05). At the time of PD withdrawal, the levels of serum creatinine, serum calcium, serum albumin and parathyroid hormone in the early withdrawal group were significantly lower than those in the late withdrawal group (all P<0.05). The Kaplan-Meier curves showed that there was no significant difference in the cumulative survival of patients in both groups (log-rank test χ2=3.485, P=0.062). Cox regression analysis revealed serum creatinine≥209 μmol/L at the time of PD withdrawal was an independent risk factor for poor prognosis ( HR=5.253, 95% CI 1.757-15.702, P=0.003). Conclusions:PD can be used for RPGN patients caused by lupus nephritis, ANCA-associated vasculitis and primary nephritis. Serum creatinine≥209 μmol/L at the time of PD withdrawal is an independent risk factor for poor prognosis.

Objective To investigate the risk factors and prognosis of peritoneal dialysis (PD)related bacterial peritonitis.Methods The clinical data of patients with PD related bacterial peritonitis from January 2006 to September 2016 in our hospital were retrospectively analyzed and followed up until December 2016.Patients were divided into two groups according to the frequency of peritonitis,single episode group and multiple episodes group (no less than two episodes of peritonitis).According to efficacy of therapy,the episodes of peritonitis were divided into two groups,cured group (no relapse,recurrence or repeat episodes) and failure group (relapse,recurrent or repeat infection after the therapy of initial episode).Logistic regression and Cox regression were used to analyze the risk factors for outcomes.Results Five hundred and fifty-nine patients had PD related bacterial peritonitis,including 339 patients in the single episode group and 220 patients in the multiple episodes group.Logistic analysis showed low serum albumin level (OR=787,P ＜ 0.001) and malnutrition (OR=0.422,P ＜ 0.001) at baseline were independent risk factors for multiple episodes (P ＜ 0.001).The technical survival was better in the single episode group than that in the multiple episodes group (75.2％ vs 36.2％,P=0.001) while the difference of survival rate was not significant between the two groups (48.2％ vs 24.1％,P=0.592).Five hundred and thirteen episodes of peritonitis were analyzed,including 147 episodes in failure group (88 relapse episodes,16 recurrent episodes and 43 repeat episodes) and 366 episodes in cured group.There were 78 patients in failure group and 253 patients in cured group.Logistic analysis showed prolonged response time (OR=1.200,P ＜ 0.001),Gram-positive bacteria infection (OR=1.736,P=0.022),higher hs-CRP level (OR=1.004,P=0.013),lower serum albumin level (OR=0.936,P=0.008) were independent risk factors for failure of therapy.Multivariate Cox regression showed prolonged response time (HR=1.120,P=0.032),Gram-positive bacteria infection (HR=2.462,P=0.002),higher hs-CRP level (HR=1.007,P=0.009) were independent risk factors for failure of therapy and higher serum albumin level (HR=0.942,P=0.048) was an independent protection factor.Conclusions Low serum albumin level and malnutrition at baseline are independent risk factors for patients with multiple peritonitis episodes.Prolonged response time,Gram-positive bacteria infection,the high hs-CRP level are independent risk factors for relapse or recurrent or repeat episodes while high serum albumin level was an independent protection factor.