BACKGROUND: More and more researches prove that cell apoptosis could be induced by glutamine, also there are more researches on studying the indirect and direct nervous-protective effects of insulin, but the nervous-protective effects of insulin on impairment induced by glutamine, as well as its mechanism still need further investigation.OBJECTIVE: To investigate the nervous-protective effects of insulin on impairment induced by glutamine in PC12 cells, and to explore its molecular mechanism.DESIGN: A prospective controlled study based on cells.SETTING: Department of Neurology, Zhejiang Hospital; Department of Neurology of Sun Yat-wen University Hospital.MATERIALS: The study was carried out at the Laboratory of the Third Affiliated Hospital and the Experimental Animal Center of Sun Yat-sen University from March 2002 to March 2003. PC12 cells were purchased from the same animal center.METHODS: Traumatic models were made in PC 12 cells by treated with 0.5 mmol/L glutamine for 20 minutes, and the insulin of different concentration were used for protection, after 24 hours, protective effects of insulin were assessed with MTT method, Hoechst33258 fluorescence staining, DNA agar gelatin electrophoresis, meanwhile the expression of PKB/Akt protein were also detected./Akt protein in experimental group.RESULTS: The A value of50 mU/L, 100 mU/L, 200 mU/L, 400 mU/L insulin groups were 0. 214 ±0. 062, 0. 234 ±0. 067, 0. 260 ±0. 076 and 0. 265 ± 0. 069, respectively, but the value of single glutamine group was 0. 201 ± 0. 079, statistical analysis indicated that compared with single glutamine group, there were no significant difference in 50 mU/L, 100 mU/L insulin groups( P > 0.05), but 200 mU/L, 400 mU/L insulin groups were found statistically different from single glutamine group(t=-2.398,-2. 716, P < 0.05); "DNA Ladder" could not be observed in 400 mU/L insulin group by electrophoresis;It was proved that Insulin could enhance the expression of PKB/Akt protein.CONCLUSION: Insulin has nervous-protective effects on impairment induced by glutamine in PC12 cells, furthermore it also has property of anti-apoptosis, and its protective mechanism might be associated with enhancement of the expression of PKB/Akt protein.

Objective Clinical effects of dot from body skin irradiated xenogeneic skin improved composite graft.Methods Selected 80 patients with severe burn patients randomized after admission on the basis of conventional treatment,3-5 days line Crust treatment group transplanted autologous point-like skin irradiation pigskin coverage.Results Treatment of patients in group one week dressing see irradiated pigskin Tiefu full two weeks the pigskin was dry scab-like.Wound healing treatment group (29 ± 5) days (P ＜ 0.01) was significantly shorter than the control group(39 ±4) days.Wound healing rate of treatment is significantly higher than the control group (P ＜0.01).Wound infection rate reduced greatly reduce labor intensity and dressing,and reduce pain.1 year after scar formation in patients with good flexibility and functionality.Conclusions Punctate since improved composite body skin irradiated xenogeneic skin transplantation can improve skin graft survival rate,promote wound healing,treatment of a large area of the burn wound repair is feasible and effective.

Objective To investigate the prevalence of pulmonary acariasis among the employees working on traditional Chinese medicinal materials and observe the effect of treatment. Methods History inquiry, detection of mites in sputum, blood examination for eosinophils and specific antibodies, x-ray chest film were carried out for 327 workers involving in traditional Chinese medicinal materials. Mites were found in sputum in 121 persons who were then treated with metronidazole, twice a day with a daily dosage of 0.8g for seven days as a course of treatment. Two courses were conducted with an interval of 7-10 day. Prevalence and morbidity in different groups of occupation, age, and sex were analyzed. Results The overall infection rate of mites in sputum was 37.0% (121/327) with an average morbidity of 12.5% (41/327). Among the four types of worker investigated, the highest infection rate (51.8%), and morbidity (18.6%) were in those working in transfer warehouse; the second highest infection rate (40.7%) and morbidity (15.7%) were in employees in factory of Chinese traditional medicine. Both groups showed a significant difference with others(?2inf=11.36,P0.05). After treatment with metronidazole, 88.4% showed negative in sputum examination for mites and the efficacy of the treatment for pulmonary acariasis was 92.3%. Conclusions Employees engaged in traditional Chinese medicinal materials are one of the groups at the highest risk of pulmonary acariasis. Metronidazole is effective in treating the infection.

Objective To assess the value of four different techniques of detecting the Mycobacterium tuberculosis (MTB) in bronchoalveolar lavage fluid (BALF) in the diagnosis of tracheobronchial tuberculosis. Methods A total of 98 patients diagnosed as tracheobronchial tuberculosis were selected from May 1,2013 to June 30,2016. The clinical data was analyzed retrospectively,and the positive rates of MTB of the 960 cultrue, the direct smears , the modified Ziehl?Neelsen stain method and Xpert MTB/RIF assay were compared. Results The positive rates of the 960 cultrue,the direct smears,the modified Ziehl?Neelsen stain method and Xpert MTB/RIF assay were 20.4%(20/98),15.3%(15/98),70.4%(69/98) and 74.5%(73/98),respectively. Among the four techniques ,the positive rates of the modified Ziehl?Neelsen stain method and Xpert MTB/RIF assay were significantly higher than those of the 960 cultrue and the direct smears(P<0.05,respectively). However,no significant difference was found between the modified Ziehl?Neelsen stain method and the Xpert MTB/RIF assay (P > 0.05). Conclusions The modified Ziehl?Neelsen stain method and Xpert MTB/RIF assay for detecting the MTB in BALF have high clinical value in the diagnosis of tracheobronchial tuberculosis.

Objective To investigate the efficacy and safety of compound lysostaphin disinfectant solution in the latter stage of the treatment of bum wounds infection,and to evaluate its clinical value.Methods The selected 60 cases of bum wound infection were grouped into treatment and control group at random.The treatment group recieved compound lysostaphin disinfectant solution(Shanghai Hi-tech Co.bio-engineering,Ltd.)with debridement and wet,once a day;the control group with normal saline+gentamicin wet gauze,once a day.After cleaning the wound two groups 40 cases of underwent regular skin debridement,20 cases in each group.The bacteria changes in the wound were detected,of bacteria removal the accumulated rate of wound healing time and healing rate were compared.Results 3,6,9 days after treatment the accumulated rate of bacteria removal was 45.8%,73.9%, 89.5%higherthan the control group(13.8%,27.6%,57.4%)respectively(P＜0.01).Wound healing time in the treatment group was(12±5)d,shorter than the control group(16±4)d Was significantly(P＜0.01). The rate of wound healing in the treatment group was significantly higher(P＜0.01).No adverse reactions were found.Conclusion The treatment application of compound iysostaphin disinfectant in the later stage of bum wound is effective in control of infection,increases the survival rate of skin and shortens wound healing time,so it is safe and effective.

Objective To assess the efficacy of pirfenidone combined with PD-L1 inhibitor for treatment of bladder cancer in a mouse model and its effect on tumor immune microenvironment modulation.Methods Forty C57BL/6 mouse models bearing ectopic human bladder cancer xenografts were randomized into control group,PD-L1 inhibitor group,pirfenidone group and combined treatment group(n=10).After successful modeling,PD-L1 inhibitor treatment was administered via intraperitoneal injection at 12.5 mg/kg every 3 days,and oral pirfenidone(500 mg/kg)was given on a daily basis.The survival rate of the mice and tumor growth rate were compared among the 4 groups.The expressions of CD3,CD8,CD45,E-cadherin and N-cadherin in the tumor tissues were detected with immunohistochemistry after the 21-day treatment,and bone marrow-derived suppressor cells(MDSCs)were observed with immunofluorescence staining;serum levels of alanine aminotransferase(ALT),aspartate aminotransferase(AST),urea nitrogen(BUN),creatinine(CRE)and lactate dehydrogenase(LDH-L)were analyzed using an automated biochemical analyzer.Results Treatment with PD-L1 inhibitor and pirfenidone alone both significantly decreased tumor growth rate and tumor volume at 21 days(P<0.05),but the combined treatment produced an obviously stronger inhibitory effect(P<0.05).PD-L1 inhibitor and pirfenidone alone significantly increased E-cadherin expression and decreased N-cadherin expression in the tumor tissue(P<0.05).The two treatments both significantly increased the percentage of CD3+,CD8 and CD45+ T cells and decreased the percentage of Ly-6G+CD11b+MDSCs in the tumor tissue,and these changes were more obvious in the combined treatment group(P<0.05).No significant differences were found in serum ALT,AST,BUN,CRE or LDH-L levels among the 4 groups(P>0.05).Conclusion Combined treatment with pirfenidone and PD-L1 inhibitor significantly inhibits the progression of bladder cancer in mice possibly by regulating tumor immune microenvironment and inhibiting epithelial-mesenchymal transition of the tumor cells.

Objective To assess the efficacy of pirfenidone combined with PD-L1 inhibitor for treatment of bladder cancer in a mouse model and its effect on tumor immune microenvironment modulation.Methods Forty C57BL/6 mouse models bearing ectopic human bladder cancer xenografts were randomized into control group,PD-L1 inhibitor group,pirfenidone group and combined treatment group(n=10).After successful modeling,PD-L1 inhibitor treatment was administered via intraperitoneal injection at 12.5 mg/kg every 3 days,and oral pirfenidone(500 mg/kg)was given on a daily basis.The survival rate of the mice and tumor growth rate were compared among the 4 groups.The expressions of CD3,CD8,CD45,E-cadherin and N-cadherin in the tumor tissues were detected with immunohistochemistry after the 21-day treatment,and bone marrow-derived suppressor cells(MDSCs)were observed with immunofluorescence staining;serum levels of alanine aminotransferase(ALT),aspartate aminotransferase(AST),urea nitrogen(BUN),creatinine(CRE)and lactate dehydrogenase(LDH-L)were analyzed using an automated biochemical analyzer.Results Treatment with PD-L1 inhibitor and pirfenidone alone both significantly decreased tumor growth rate and tumor volume at 21 days(P<0.05),but the combined treatment produced an obviously stronger inhibitory effect(P<0.05).PD-L1 inhibitor and pirfenidone alone significantly increased E-cadherin expression and decreased N-cadherin expression in the tumor tissue(P<0.05).The two treatments both significantly increased the percentage of CD3+,CD8 and CD45+ T cells and decreased the percentage of Ly-6G+CD11b+MDSCs in the tumor tissue,and these changes were more obvious in the combined treatment group(P<0.05).No significant differences were found in serum ALT,AST,BUN,CRE or LDH-L levels among the 4 groups(P>0.05).Conclusion Combined treatment with pirfenidone and PD-L1 inhibitor significantly inhibits the progression of bladder cancer in mice possibly by regulating tumor immune microenvironment and inhibiting epithelial-mesenchymal transition of the tumor cells.

African swine fever(ASF)is an acute and highly pathogenic hemorrhagic disease of pigs,causing huge economic losses to pig industry.In order to quantitatively detect clinical samples of ASF and inactivated ASFV antigens,the IgG of ASF positive serum was used as capture anti-body and the HRP-labeled p72 monoclonal antibody was used as detecting antibody.The standard curve was drawn with the cell-cultured ASFV,and a sandwich ELISA detection of antigen was es-tablished.The specificity,sensitivity and stability of the method were evaluated.The effects of dif-ferent inactivation methods and adjuvant addition on antigen detection were further evaluated.The results showed that the minimum detection limits of the recombinant protein and the ASFV were 0.1 mg/L and 103.7 TCID50/mL,respectively.There was no cross-reaction with five common porcine pathogenic viruses,and the coefficient variations between batches was less than 10％.The total co-incidence rate with real-time fluorescence quantitative PCR was 92％(23/25).The sensitivity of antigen detection was significantly reduced when antigen was treated by BEI inactivation,and the detection results were severely interfered by aluminum adjuvant and nano-adjuvant.In summary,the sandwich ELISA antigen detection method established is specific,sensitive,and repeatable,with a good consistency to the qPCR method,which provides an effective clinical diagnostic meth-od for ASFV antigen.

Objective:To observe any effect of an enriched environment (EET) on cognitive functioning and sonic hedgehog (SHH) signaling in rats modeling cerebral ischemia.Methods:Sixty adult male Sprague-Dawley rats were randomly divided into a blank control group, a model group and a training group. A model of cerebral ischemia was established in the model and training groups by thread thrombus. The training group was given an EET. After 7 and 14 days, the rats′ cognition was tested using the Morris water maze and the platform jumping test. Apoptosis of brain cells in the hippocampus was detected by using TUNEL staining, and the expression of SHH, Gli2 and PTCH1 proteins in the hippocampus were measured using qRT-PCRs, western blotting and immunohistochemistry.Results:After 14 days the average escape latency in the Morris water maze test had shortened more in the training group than in the model group, while the average swimming speed, the number of platform crossings and the time spent in the quadrant had increased significantly more. They also received fewer electric shocks and spent significantly less time on the platform in the platform jumping test on average. Apoptosis in the hippocampus after 14 days was significantly less in the training group with significantly greater expression of SHH and Gli2 protein and significantly less PTCH1 protein expression.Conclusion:An EET can significantly improve cognition after cerebral ischemia, at least in rats. Its mechanism may be related to enhanced activation of the SHH signaling pathway.