Objective To express and purify EGF-TCS fusion protein and observe the targeted and selective killing effect on cancer cells of the fusion protein.Methods The recombinant expression plasmid PQE30/EGF-TCS was transformed to E.coli.M15 and the fusion protein(EGF-TCS) was expressed.Ni-NTA Agrose affinity chromatography was used to purify the protein,flow cytometry to detect EGFR expression rate in cancer cells(BEL-7402,MCF-7,BGC-823) and normal liver LO2 cells,and the killing test to verify selective killing ability of EGF-TCS;The cell apoptosis detection by flow cytometry and microscopic observation were used to confirm the selective killing ability of EGF-TCS.Results Recombinant expression plasmid PQE30/EGF-TCS was expressed in E.coli.M15 stably and effectively.The purity of EGF-TCS was over 95% by chromatography.EGFR expression rate was highest in hepatoma cells BEL-7402(72.33%) and lowest in normal liver LO2 cells(5.51%).The killing ability of recombinant protein was more effective to cancer cells(IC50 of BEL-7402,MCF-7 and BGC-823 was 11.4,22.47 and 12.53 ?g/ml respectively) and was weak to normal cells(IC50 53.19 ?g/ml).Conclusion The recombinant protein EGF-TCS that induces apoptosis of cancer cells was successfully constructed by gene engineering technology.

Objective To observe the effects of Weipixiao on the ultrastructure of gastric mucosa capillaries in rats with precancerous lesions of gastric cancer ( PLGC). Methods The rats were randomized into six groups, including normal group, model group, Vatacoenayme Tablets group ( 0.2 g·kg-1·d-1) , and high-, middle-, and low- dose Weipixiao groups ( 15, 7.5, and 3.75 g·kg-1·d-1 respectively) . The rats received spontaneous intake of N-methyl-N’-nitro-nitrosoguanidine ( MNNG, 200 μg/mL) solution combined with irregular diet and intragastric administration of purgative herbs Xiao Chengqi Decoction ( 2 mL, containing 1 g/mL crude drug) for 18 weeks to induce spleen-deficiency PLGC. The pathological changes in gastric mucosa and the ultrastructure of gastric mucosa capillaries were observed under the transmission electron microscope. Results The model has been established successfully. Transmission electron microscopy results in the model group showed as severely swollen endothelial cells of gastric mucosa capillaries, severely-narrowing or even blocked vascular lumens, rough and discontinuous basement membrane, and swollen, degenerated or even absent pericytes. And the ultrastructure of gastric mucosa capillaries in high-, middle-, and low- dose Weipixiao groups were improved to some degrees, the effect of low-dose Weipixiao group being the best. Conclusion The improvement of the mucosal microcirculation of spleen-deficiency PLGC rats may be one of the pathohistological mechanisms of Weipixiao for spleen-deficiency PLGC.

AIM:To observe the effect of Wnt/β-catenin signaling pathway on diabetic ulcer.METHODS:Diabetic animal model was established in the female Wistar rats by intraperitoneal injection of low-dose streptozotocin fol-lowing high-fat diet feeding.A circular wound was made on the dorsum of the rats in both control group and diabetic group. The condition of wound healing was recorded and the structures of the wound tissues were observed by HE staining in the 2 groups at 3, 7 and 14 d after wounding.The expression ofβ-catenin, GSK-3βand Rspo-3 at mRNA and protein levels in the wound tissues was detected by RT-PCR and ELISA.RESULTS:In diabetic group, the wound healing rate was lower (P<0.05), and the inflammatory cells, fibroblast cells and new capillaries in the wound tissues were fewer than those in control group.The expression of β-catenin and Rspo-3 at mRNA and protein levels in the wound tissues in control group was significantly higher than those in diabetic group, and the expression of GSK-3βwas exactly the opposite (P<0.05). CONCLUSION:The down-regulation of Wnt/β-catenin probably resultes from the decreased level of Rspo-3, which may be one of the reasons for delaying the diabetic ulcer healing.

Objective To observe the effect of Weipixiao, a compound recipe which has the actions of strengthening spleen, resolving stasis and removing toxins, on the histopathological changes of gastric mucosal tissue in rats with gastric precancerous lesions ( GPL) . Methods SD rats were randomly divided into normal group, model group, Vitacoenzyme group (0.2 g·kg-1·d-1), and high-, middle-, and low-dose Weipixiao groups ( in the dose of 15, 7.5, 3.75 g·kg-1·d-1, respectively) . Except for the normal control group, the rats in other groups received spontaneous intake of N-methyl-N’-nitro-nitrosoguanidine ( MNNG) solution combined with irregular diet and oral use of purgative herbs for 18 weeks to induce GPL. From the 9th week, the mediation groups were simultaneously given corresponding medicine for 10 weeks. At the end of the experiment, the histopathological changes of gastric mucosal tissue in all groups were observed. Results Pathological scores of intestinal metaplasia and epithelial dysplasia in rat gastric mucosa of the model group were significantly increased ( P<0.01 compared with those of the normal group) , but were decreased in three Weipixiao groups to various degrees, particularly in low-dose Weipixiao group ( P<0.05 or P<0.01) . Conclusion Weipixiao can block and reverse gastric intestinal metaplasia and dysplasia in GPL rats to certain degrees, and low-dose Weipixiao may have better long-term effect for the prevention and treatment of GPL.

Objective:To investigate the effects of 3-methyladenine(3-MA)on extracellular matrix deposition in early diabetic nephropathy(DN)and its mechanism.Methods:A streptozotocin(STZ)-induced type 1 diabetes mouse model was used, and the mice were divided into vehicle control group, diabetes group(STZ group), 3-MA group, and chloroquine(CQ)group, 8 mice in each group. After 6 weeks of intervention, both kidneys were harvested, and the kidney-to-body weight ratio was recorded. Western blotting was performed to detect protein expressions of renal cortex fibronectin, α-smooth muscle actin(α-SMA), LC3, Beclin 1, p62, and vascular endothelial growth factor(VEGF). Immunohistochemistry was used to observe kidney fibronectin staining. Bioinformatics analysis was conducted on shared genes between diabetic nephropathy(DN)gene targets and 3-MA predicted gene targets.Results:Both 3-MA and CQ exhibited certain hypoglycemic effects in diabetic mice. Compared to the STZ group, the kidney-to-body weight ratio decreased in the 3-MA group( P<0.05). Western blotting showed that 3-MA reduced the expression of renal cortex matrix-related proteins fibronectin and α-SMA in diabetic mice( P<0.05 or P<0.01). Immunohistochemistry also revealed that 3-MA reduced fibronectin staining in the kidneys of diabetic mice. Both 3-MA and CQ inhibited the protein expression of renal cortex Beclin 1 in diabetic mice(both P<0.05), while 3-MA increased the expression of renal cortex p62( P<0.05). Bioinformatics analysis indicated a connection between shared genes of DN gene targets and 3-MA predicted gene targets with the VEGF signaling pathway. Western blotting results further showed that 3-MA reduced renal cortex VEGF expression in diabetic mice( P<0.01). Conclusion:3-MA can alleviate extracellular matrix deposition in the kidneys of early DN mice by inhibiting the VEGF signaling pathway.

Objective:To analyze the risk factors for bile leakage in patients undergoing cystic echinococcosis surgery to establish a nomogram model to predict the risk of bile leakage.Methods:The clinical data were retrospectively analyzed that 203 patients with cystic echinococcosis treated in Qinghai University Affiliated Hospital from January 2015 to October 2018. Logistic regression was used to screen out independent risk factors of biliary leakage in patients undergoing cystic hydatid surgery. Then the R software was used to establish a predictive nomogram model. Bootstrap method was used to validate the nomogram model and ROC curve was used to explore the predictive efficacy of the model in predicting the bile leakage.Results:There were 54 cases of biliary leakage in 203 patients with cystic echinococcosis, the incidence was 26.1%. ALP ( OR=11.193, 95% CI 5.066-24.731), GGT ( OR=2.728, 95% CI 1.246-5.975), cyst diameter ( OR=3.491, 95% CI 1.550-7.861), hilar cyst ( OR=2.503, 95% CI 1.176-5.329) were all independent risk factors for biliary leakage of cystic echinococcosis. The consistency index was 0.835 for predicting PPC risk, and the area was 0.823 (95% CI 0.754-0.892) under the curve for predicting the risk of cystic echinococcal bile leakage. Conclusions:ALP, GGT, cyst diameter and hepatic hilar cyst are independent risk factors for cystic echinococcal bile leakage. Nomogram model can evaluate the risk of cystic echinococcal bile leakage more intuitively, with potentially high clinical application value.

Objective To explore the mechanism of 3-methyladenine(3-MA)for alleviating early diabetic renal injury.Methods Mouse models of streptozotocin(STZ)-induced diabetes mellitus were randomized into model group and 3-MA treatment group for daily treatments with normal saline and 10 mg/kg 3-MA by gavage for 6 weeks,respectively.Body weight and fasting blood glucose of the mice were recorded every week.After the treatments,the kidneys of the mice were collected for measurement kidney/body weight ratio,examination of glomerular size with PAS staining,and detection of α-SMA and PCNA expressions using Western blotting and immunohistochemistry.SV40 MES 13 cells cultured in normal glucose(5.6 mmol/L)and high glucose(30 mmol/L)were treated with 24.4 mmol/L mannitol and 5 mmol/L 3-MA for 24 h,respectively,and the changes in cell viability and PCNA expression were examined using CCK8 assay and Western blotting.Bioinformatics analysis of the intersecting gene targets of diabetic kidney disease(DKD)and 3-MA was performed,and the results were verified by Western blotting both in vivo and in vitro.Results In the diabetic mice,treatment with 3-MA produced a short-term hypoglycemic effect,reduced the kidney/body weight ratio and glomerular hypertrophy,and decreased the expressions of α-SMA and PCNA in the renal cortex.In the in vitro study,3-MA significantly lowered the viability and reduced PCNA expression in SV40 MES 13 cells exposed to high glucose.The results of bioinformatic analysis identified AKT1 as the key gene in the therapeutic mechanism of 3-MA for DKD.Western blotting confirmed that 3-MA inhibited the phosphorylation of AKT and S6 in both the renal cortex of diabetic mice and high glucose-treated SV40 MES 13 cells.Conclusion 3-MA suppresses mesangial cell proliferation and alleviates early diabetic renal injury in mice possibly by inhibiting AKT signaling.

Objective To explore the mechanism of 3-methyladenine(3-MA)for alleviating early diabetic renal injury.Methods Mouse models of streptozotocin(STZ)-induced diabetes mellitus were randomized into model group and 3-MA treatment group for daily treatments with normal saline and 10 mg/kg 3-MA by gavage for 6 weeks,respectively.Body weight and fasting blood glucose of the mice were recorded every week.After the treatments,the kidneys of the mice were collected for measurement kidney/body weight ratio,examination of glomerular size with PAS staining,and detection of α-SMA and PCNA expressions using Western blotting and immunohistochemistry.SV40 MES 13 cells cultured in normal glucose(5.6 mmol/L)and high glucose(30 mmol/L)were treated with 24.4 mmol/L mannitol and 5 mmol/L 3-MA for 24 h,respectively,and the changes in cell viability and PCNA expression were examined using CCK8 assay and Western blotting.Bioinformatics analysis of the intersecting gene targets of diabetic kidney disease(DKD)and 3-MA was performed,and the results were verified by Western blotting both in vivo and in vitro.Results In the diabetic mice,treatment with 3-MA produced a short-term hypoglycemic effect,reduced the kidney/body weight ratio and glomerular hypertrophy,and decreased the expressions of α-SMA and PCNA in the renal cortex.In the in vitro study,3-MA significantly lowered the viability and reduced PCNA expression in SV40 MES 13 cells exposed to high glucose.The results of bioinformatic analysis identified AKT1 as the key gene in the therapeutic mechanism of 3-MA for DKD.Western blotting confirmed that 3-MA inhibited the phosphorylation of AKT and S6 in both the renal cortex of diabetic mice and high glucose-treated SV40 MES 13 cells.Conclusion 3-MA suppresses mesangial cell proliferation and alleviates early diabetic renal injury in mice possibly by inhibiting AKT signaling.

Objective:To investigate the effect of hepatitis B virus (HBV) infection in patients with hepatic alveolar echinococcoisis after surgical resection and determine the differences of liver function between patients with different HBV-DNA levels.Methods:Patients were selected from January 2014 to July 2018 in the Affiliated Hospital of Qinghai University. Twenty-eight patients with hepatitis B and hepatic alveolar echinococcoisis were included in the experimental group, and 20 patients with hepatic alveolar echinococcoisis but without hepatitis B virus were included in the control group. Based on HBV-DNA level, the experimental group was divided into low-level group (HBV-DNA level<200 IU/ml, n=6), intermediate-level group (HBV-DNA level 200-20 000 IU/ml, n=15) and high level group (HBV-DNA level>20 000 IU/ml, n=7). Comparison of complications and liver function after liver resection in two groups.Univariate and multivariate logistic regression were used to analyze the influential factors of postoperative complications in patients.Comparison of postoperative liver function indexes in patients with different HBV-DNA levels. Results:In the control group, postoperative total bilirubin 10.6(8.3, 16.9) μmol/L, direct bilirubin 5.3(3.4, 10.0) μmol/L, prothrombin time 13.6(13.0, 15.8)s, and the incidence of complications 25.0%(5/20), were better than the experimental group 12.6(8.4, 46.9) μmol/L, 6.7(3.1, 26.4) μmol/L, 15.4(13.5, 18.1)s, 78.6% (22/28), the differences were statistically significant significance (all P<0.05). Multivariate logistic analysis showed that patients with HBV infection ( OR=4.593, 95% CI: 1.128-18.708) and intraoperative blood loss ≥1 000 ml ( OR=2.200, 95% CI: 1.106-4.378) were the hepatic alveolar echinococcoisis independent risk factors for postoperative complications. There were no significant differences in total bilirubin and albumin between the three groups of patients with different HBV-DNA levels ( P>0.05). Conclusion:Patients with HBV and hepatic alveolar echinococcoisis have worse liver function and are more prone to complications after surgical resection, but there is no significant difference in liver function among patients with different HBV-DNA levels.

During coronavirus disease 2019 epidemic, the treatment of severe trauma has been impacted. The Consensus on emergency surgery and infection prevention and control for severe trauma patients with 2019 novel corona virus pneumonia was published online on February 12, 2020, providing a strong guidance for the emergency treatment of severe trauma and the self-protection of medical staffs in the early stage of the epidemic. With the Joint Prevention and Control Mechanism of the State Council renaming "novel coronavirus pneumonia" to "novel coronavirus infection" and the infection being managed with measures against class B infectious diseases since January 8, 2023, the consensus published in 2020 is no longer applicable to the emergency treatment of severe trauma in the new stage of epidemic prevention and control. In this context, led by the Chinese Traumatology Association, Chinese Trauma Surgeon Association, Trauma Medicine Branch of Chinese International Exchange and Promotive Association for Medical and Health Care, and Editorial Board of Chinese Journal of Traumatology, the Chinese expert consensus on emergency surgery for severe trauma and infection prevention during coronavirus disease 2019 epidemic ( version 2023) is formulated to ensure the effectiveness and safety in the treatment of severe trauma in the new stage. Based on the policy of the Joint Prevention and Control Mechanism of the State Council and by using evidence-based medical evidence as well as Delphi expert consultation and voting, 16 recommendations are put forward from the four aspects of the related definitions, infection prevention, preoperative assessment and preparation, emergency operation and postoperative management, hoping to provide a reference for severe trauma care in the new stage of the epidemic prevention and control.