Objective To investigate the dynamic expressions of exchange protein directly activated by cyclic adenosine monophosphate (cAMP) (Epac) in rat model of hepatic fibrosis(HF).Methods Forty-two male SD rats were divided into control group (n = 6) and model group (n = 36)which was divided into six subgroups of day 4, week 1, week 2, week 4,week 6 and week 8 with six rats in each subgroup. The rat model of HF was established by intraperitoneal injection of dimethylnitrosamine (DMN). The pathological changes of liver were observed by Hematoxylin-Eosin and Masson staining. Reverse transcription-polymerase chain reaction (RT-PCR),immunohistochemistry and Western blot were employed to detect the mRNA and protein expressions of Epac1, Epac2 and transforming gronth factor (TGF)β1 during the process of modeling and localization in the liver. The statistical analysis was done using one-factor ANOVA, LSD-t test,Dunnett T3 test and Pearson linear correlation analysis. Results Rat model of liver fibrosis was established successfully. In control group, Epac1 (0. 031 28±0. 008 96) and Epac2 protein (0.034 43±0. 002 45) mainly expressed in the cytoplasm of hepatocytes. In model group, the level of Epac1 decreased at day 4 (0. 023 97±0. 003 81) and week 1 (0. 015 81±0. 002 48) ,then began to increase at week 2 of modeling and peaked at week 6 (0. 039 54±0. 001 43), which had statistical significance compared to the control group (t= 5.47,11.58 and - 6.18, respectively; all P＜0.05). Epac2 protein expression declined after modeling, reached the lowest level at week 4 (0. 011 21 ±0. 001 32), which had statistical significance compared to the control group (t= 24. 50, P＜0. 05). TGFβ1 protein expression increased after modeling and peaked at week 4 (0. 011 30±0.001 03) which had statistical significance (t= -23. 36, P＜0. 05) compared to the control group (0. 002 08 ±0. 000 18). The expressions of Epac1, Epac2 and TGFβ1 mRNA were consistent with the trend of protein levels.Correlation analysis showed that Epac1 protein was positively correlated with the course of HF (r =0. 703, P＜0.01 ), while Epac2 protein was negatively correlated (r = - 0. 409, P＜0.05). Conclusions During the progression of HF, Epac1 expression tends to decrease firstly and increase afterwards,while Epac2 expression declines continually. Epac may be involved in the pathogenesis of HF.

Objective To investigate the changes and clinical significance of the frequency of circulating CD4+ CD25+ regulatory T cells (Treg) in nonalcoholic fatty liver (NAFL) patients.Methods CD4+ CD25+ Treg in the peripheral blood from 50 NAFL patients and 50 healthy subjects were quantitatively analyzed using flow cytometry. Group t test or Mann-Whitney U test and Spearman's rank correlation test were used for statistical analysis.Results The proportion of circulating CD4+CD25+ Treg in NAFL patients was (5.39 ± 1.94)％,which was significantly higher than that in healthy controls [(4.21±1.52)％](t=3.385,P＜0.01).Further analysis revealed that the frequency of Treg was positively correlated with triglyceride (TG) level and body mass index (r=0.307 and 0.251,respectively; P=0.002 and 0.012,respectively),and negatively correlated with high density lipoproteincholesterol (HDL-C) (r=-0.306,P=0.002).Meanwhile,Treg in patients with high body mass index,high TG,low HDL-C,hypertension and metabolic syndrome (MS) were all higher than those in controls (t=2.294,2.533,3.154,2.010 and 4.454,respectively; all P＜0.05).But there was no significant difference between patients with high fasting blood glucose and controls (U=1143.500,P=0.471).Conclusion The increased frequency of peripheral Treg in NAFL patients may have some relations with the imbalance of proinflammation and anti-inflammation in NAFL patients who coexisting with MS.

Objective To explore the dynamic expressions and the significance of Notch/Jagged signal pathway in rat model of hepatic fibrosis. Methods A total of 42 healthy male SD rats were randomly divided into normal control group (n=6) and model group (n= 36). The model group was further divided into six subgroup according to different time points: subgroups of 4 days, 1, 2, 4, 6 and 8 weeks with six rats in each subgroup. The rat model of hepatic fibrosis was induced by dimethylnitrosamine (DMN). The serum levels of alanine aminotransferase (ALT), aspertate aminotransferase (AST), albumin (Alb) and hyaluronic acid (HA) were detected dynamically after 4 days, 1,2,4,6 and 8 weeks of injection. The liver tissues were observed under optical microscope after HE and Masson staining. Notch-1, Jagged-1 mRNA and protein in liver were detected by reverse transcriptase polymerase chain reaction (RT-PCR) and immunohistochemistry. The comparison of means among groups was done by univariate ANOVA. Results The hepatic fibrosis model was successfully induced by DMN injection and pseudolobules were found after 4 weeks of injection. The serum levels of ALT, AST, Alb and HA were all increased after 4 day of injection and peaked at week 4 which were all significantly higher than those in control group (F=83.10, 104.63, 54.24, 203.81,respectively; all P＜0.05). The expressions of Notch-1, Jagged-1 mRNA and protein in model group were all significantly increased than those in control group (F=282. 44, 369.14, 374.17, 256. 14,respectively;P＜0. 01). And the expressions of Notch-1, Jagged-1 were closely correlated with the hepatic fibrosis stages and transforming growth factor β1 (TGFβ1) expression (r=0. 821, 0. 917,0. 767,0. 844, respectively; P＜0. 01 ). Conclusions The Notch/Jagged pathway may participate in the development of hepatic fibrosis, which is closely correlated with the progression and severity of liver fibrosis.

ObjectiveTo investigate whether CD4+ CD25+ CD127dim/- regulatory T lymphocytes (Treg) can induce the proliferation of hepatic stellate cells (HSC) and expression of fibrosis-related factors on HSC in vitro and further to explore the mechanism of Treg inducing fibrogenesis. MethodsHSC LX-2 cells were subcultured.CD4+ CD25+ CD127dim/- cells were purified using magnetic cell separation. The HSC were co-cultured with Treg by direct contact or by Transwell system in vitro. The HSC cultured alone was used as control. Cell proliferation was measured by CCK-8 assay.The expression of transforming growth factor (TGF)-β1 was detected by enzyme-linked inmunosorbent assay (ELISA), and the expressions of hyaluronic acid (HA) and precollagen Ⅲ (PC Ⅲ ) were detected by radio immunoassay (RIA). The data were analyzed by LSD-t test. ResultsHSC proliferation was strongest when Treg∶ HSC= 1.5∶ 1. The absorbance in direct contact co-culture group and Transwell system co-culture group was (0. 713±0. 032) cpm and (0. 735±0. 028) cpm, respectively, both of which were higher than that in control group [(0. 677 ± 0. 029) cpm](t = 5. 4003 and 8. 7878,respectively; both P＜0. 01). The concentrations of TGF-β1 in the supernatant were (781. 59 ±76.45) pg/mL and (813. 53±60. 62) pg/mL, respectively in direct contact co-culture group and Transwell system co-culture group, which were significantly higher than that in control group [(722.51±59. 66) pg/mL](t = 4.0014 and 6. 1653, respectively; both P＜0.01).The concentrations of HA were (433. 575±27.90) ng/mL and (445.40±23.73) ng/mL, respectively in direct contact co-culture group and Transwell system co-culture group, which were higher compared to that in control group [-(415. 83±19.44) ng/mL](t =3. 3124 and 5. 5231, respectively; both P＜0.01). Likewise, the concentrations of PCⅢ were (21. 93± 1.71) and (23. 125± 1.87) ng/mL in direct contact group and Transwell group, respectively compared to (20. 10± 1.49) ng/mL in control group (t = 4. 8082 and 7. 9436, respectively; both P ＜ 0.01).Furthermore, the absorbance,concentrations of TGF-β1, HA and PC Ⅲ in Transwell co-culture group were all higher compared to direct contact group (t = 3. 3875, 2.1639, 2. 2107 and 3.1354, respectively; all P＜0. 05).ConclusionsThe cell proliferation and the expressions of fibrosis-related factors in HSC increase greatly after co-cultured with CD4+ CD25+ CD127dim/-Treg. Therefore, Treg may play an important role in inducing liver fibrogenesis.

Objective To investigate the roles of five scoring systems including model for endstage liver disease (MELD), Child-Turcotte-Pugh (CTP), Mayo, MESO and MELD-Na scoring systems, in predicting the prognosis of patients with chronic severe hepatitis. Methods The clinical data of 213 patients with chronic severe hepatitis were retrospectively studied. The five scoring systems were applied respectively to evaluate the scores in survival group and death group. The capability of these five scoring systems to predict the prognosis of severe hepatitis were compared by the receiver operating characteristic (ROC) curve, area under curve (AUC) and cut-off value.Measurement data were compared by group t test. The comparisons of AUC among scoring systems were done using MEDCLAC software. Results The scores of death group evaluated by MELD, CTP,Mayo, MESO or MELD-Na scoring systems (30.6 ± 9.5, 11.3 ± 1.5, 10.4 ± 1.3, 2.3 ± 0.8 and 39.0 ± 11.8, respectively) were consistently higher than those of survival group (21.1± 6.8, 10.6 ±1.6, 9.0±1.5, 1.6±0.5 and 22.6±8.2, respectively) (P＜0.01). The values of AUC of these five systems were 0.810, 0.623, 0.749, 0.829 and 0.885, respectively. The Youden's indexes of these five systems were 0.507, 0.175, 0.389, 0.528 and 0.650, respectively. Conclusions The CTP scoring systems can not predict the prognosis of chronic severe hepatitis very well. The Mayo scoring systems can partially predict the prognosis. On the contrary, MELD, MESO and MELD-Na systems can successfully predict the disease prognosis, and the score of MELD-Na system shows the best correlation with the prognosis.

Objective To investigate the practical efficacy and safety of ultrasound with microbubbles mediated rAAV2-EGFP to retina of rat after intravitreal and subretinal injection. Methods Gene transfer was examined by rAAV2-EGFP intravitreal and subretinal injection into the Wistar rats with or without microbubbles. The eyes were exposed to US (1 MHz,2 W/cm2, duration 5 minutes,duty cycle 50%,pulse recurrent frequency 100 Hz). The onset of EGFP gene expression, lightness of fluorescence, area of fluorescence and its distribution in the fundus in vivo via fluorescence stereosocope were investigated on the 4th,7th, 35th,49th and 120th day respectively. The value of gene transfer was quantified through the EGFP fluorescence quantitative methods by Axiovision 3. 1 software. HE staining was used to observe tissue damage. Results There was no fluorescence observed by fluorescence stereosocope after intravitreal injection after two-month study. After subretinal injection, ultrasound-targeted microbubbles destruction (UTMD) strongly increased gene transfer efficiency. UTMD used in the experiment did no harm to the rat retina structure. Conclusions UTMD could not enhance rAAV2-EGFP transfecion efficiency to rat retina after intravitreal injection but the transduction could be enhanced significantly after subretinal injection.

Objective:To establish a calculation model for the operational efficiency and resource allocation of clinical departments in hospitals, for references for hospitals to optimize resource allocation.Methods:The informations including hospitalization time, nursing grade, etc. of inpatients admitted by 32 clinical departments in a tertiary public hospital from January to December in 2021 were extracted. A data envelopment analysis method was conducted on the operation efficiency and input edundancy of the departments. The K-means algorithm was used to divide inpatients into 3 categories according to the level of medical workload. Taking the numbers of doctors, nurses and beds as the input indicators, and the numbers of patients in the 3 categories as the output indicators, a BCC model 1 was established to evaluate the efficiency of resources invested by clinical departments into professional human value. At the same time, a BCC model 2 was established with the total number of patients admitted and medical income as the output indicators to evaluate the efficiency of resources invested by clinical departments into economic benefits.Results:A total of 38 147 inpatients were enrolled. There were 14 departments with overall technical efficiency (OTE) =1.000 in the BCC model 1, 10 departments with OTE=1.000 in the BCC model 2, and 8 departments with OTE=1.000 in the 2 models. As for the input redundancy, 6 departments had high input redundancy in the BCC model 1, 11 departments had high input redundancy in the BCC model 2, and 4 departments had high input redundancy in both models.Conclusions:The model established by this study could effectively evaluate the operational efficiency and input redundancy of clinical departments, identify departments with high workload and low economic benefits, and provide reference for the rational allocation of medical resources in hospitals.