Differentiation of Rabbit Bone Marrow Mesenchymal Stem Cells to Myogenic Ceils and Expression of VEGF After Gene Transfection

Objectives To induce the differentiation of rabbit bone marrow mesenchymal stem cells （MSCs） to myogenic cells in vitro, and to investigate the expression of vascular endothelial growth factor （VEGF） gene in MSCs transfected by AdTrackCMV-VEGF165. Methods MSCs were isolated and purified from rabbit bone marrow by percoll （11）73 g/ml） and then cultured in low glucose DMEM with 10% FBS. AdTrackCMV-VEGF165 eukaryotic expression vector was constructed and transfected into the MSCs. After being incubated with 5-azacytidine （5- Aza）, the expression of troponin I in MSCs was assayed by immunohistochemistry and the expression of VEGF gene was identified by northern blot and western blot. The concentration of VEGF in the supernatant was measured by ELASA. Results MSCs were isolated and cultured successfully from rabbit bone marrow. The positive cTnI stain of some MSCs after the induction of 5-aza indicated that the cells were differentiated to myogenic cells. Northern blot and western blot showed that the expression of VEGF 165 mRNA was much higher in the hVEGF165 gene transfected cells than that of the control. The concentration of VEGF in the supernatant got to the peak 3-5 days after hVEGF165 gene transfection （1011- 1027 pg/ml） and decreased gradually thereafter, but still higher than that of control group or pAdTrackCMV group （349 pg/mLvs 116 pg/mL or 125pg/ml respectively, MSCs could be induced P〈 0.01）. Conclusions to differentiate to myogenic cells by 5-aza in vitro and could express VEGF by VEGF gene transfection.     

Apoptotic effect of Epigallocatechin-3-gallate on the human gastric cancer cell line MKN45 via activation of the mitochondrial pathway

To investigate whether Epigallocatechin-3-gallate （EGCG） can induce apoptosis of the gastric cancer cell line MKN45 and its apoptotic pathway.METHODS： To determine this, apoptotic rates of MKN45 cells after EGCG treatment with or without caspase-3 inhibitor were evaluated by Annexin V-FITC ＋ PI staining The influence of EGCG on the activity of caspase-3 in the MKN45 cells was determined by ELISA. By Rhodamine123 staining, the membrane potential change of the mitochondrion was also investigated, and mRNAs and protein expression of the bcl-2 family were analyzed by RT-PCR and Western blot.RESULTS： EGCG can induce apoptosis of MKN45 cells in time- and dose-dependent manner. Eight hours after EGCG treatment, the activity of caspase-3 in the MKN45 increased, especially 12 h after treatment. The mitochondrial membrane potential was significantly weakened 4 h after EGCG insult. The mRNA and protein expression levels of pro-apoptotic members, such as Bax, Bid and Bad, were upregulated gradually as treated time increased. Moreover, the mRNA and protein expression levels of anti-apoptotic members, such as Bcl-xL and Bcl-2, were inhibited. CONCLUSION： These data support that EGCG can induce apoptosis of the human gastric cancer cell line MKN45, and the effect is in a time- and dose-dependent manner. The apoptotic pathway triggered by EGCG in MKN45 is mitochondrial-dependent.     

Expression of bcl-2 protein in chronic hepatitis C： Effect of interferon alpha 2b with ribavirin therapy

AIM: Mechanisms responsible for persistence of HCV infection and liver damage in chronic hepatitis C are not clear. Apoptosis is an important form of host immune response against viral infections. Anti-apoptotic protein bcl-2 expression on liver tissue as well as the influence of interferon alpha 2b (IFNa2b) and ribavirin (RBV) were analyzed in patients with chronic hepatitis C. METHODS: In 30 patients with chronic hepatitis C (responders - R and non-responders - NR) treated with IFNα2b+RBV, protein bcl-2 was determined in hepatocytes and in liver associated lymphocytes before and after the treatment. RESULTS: The treatment diminished bcl-2 protein accumulation in liver cells in_patients with hepatitis C (P<0.05). Before and after the therapy, we detected bcl-2 protein in R in 87±15% and 83±20% of hepatocytes and in 28±18% and 26±10% of liver-associated lymphocytes, respectively. In NR, the values before treatment decreased from 94±32% to 88±21% of hepatocytes and 39±29% to 28±12% of lymphocytes with bcl-2 expression. There was no statistical correlation between bcl-2 expression on liver tissue with inflammatory activity, fibrosis and biochemical parameters before and after the treatment. CONCLUSION: IFNα2b+RBV treatment, by bcl-2 protein expression decrease, enables apoptosis of hepatocytes and associated liver lymphocytes, which in turn eliminate hepatitis C viruses.     

c-Jun N-terminal kinase-mediated Rubicon expression enhances hepatocyte lipoapoptosis and promotes hepatocyte ballooning

AIM: To clarify the relationship between autophagy and lipotoxicity-induced apoptosis, which is termed "lipoapoptosis," in non-alcoholic steatohepatitis. METHODS: Male C57BL/6J mice were fed a high-fat diet(HFD) for 12 wk, after which the liver histology and expression of proteins such as p62 or LC3 were evaluated. Alpha mouse liver 12(AML12) cells treated with palmitate(PA) were used as an in vitro model. RESULTS: LC3-Ⅱ, p62, and Run domain Beclin-1 interacting and cysteine-rich containing(Rubicon) proteins increased in both the HFD mice and in AML12 cells in response to PA treatment. Rubicon expression was decreased upon c-Jun N-terminal kinase(JNK) inhibition at both the m RNA and the protein level in AML12 cells. Rubicon knockdown in AML12 cells with PA decreased the protein levels of both LC3-Ⅱ and p62. Rubicon expression peaked at 4 h of PA treatment in AML12, and then decreased. Treatment with caspase-9 inhibitor ameliorated the decrease in Rubicon protein expression at 10 h of PA and resulted in enlarged AML12 cells under PA treatment. The enlargement of AML12 cells by PA with caspase-9 inhibition was canceled by Rubicon knockdown.CONCLUSION: The JNK-Rubicon axis enhanced lipoapoptosis, and caspase-9 inhibition and Rubicon had effects that were cytologically similar to hepatocyte ballooning. As ballooned hepatocytes secrete fibrogenic signals and thus might promote fibrosis in the liver, the inhibition of hepatocyte ballooning might provide antifibrosis in the NASH liver.     

乙型肝炎病毒感染肾小管上皮细胞Toll样受体4的表达及其作用

Objective To explore the expression and role of Toll receptor 4(TLR4)in human proximal tubular epithelial cell line HK-2,infected by HBV. Methods The serum of HBV DNA copies between 107-108/ml was collected. Before and after infected by HBV DNA positive serum. the HK-2 cells' morphology and the expression of α-smooth muscle actin(α-SMA)were observed by microscopy and immunofluorescence, and the effects of different concentrations of lipopolysaccharides(U)S.TLR4-stimulating factor)and CLI-095(TLR4 Inhibitor)on the proliferation rate of HK-2 cells were observed by MTT assays. After HBV serum and 10μg/ml LPS and 5μl/ml CLI-095 acted on HK-2 cells,TLR4 protein expression was measured by immunofluorescence and Western-blotting assay, and HBsAg and HBeAg in cell culture medium were detected by ELISA. and HBV DNA copies by fluorescence quantitative PCR. Results The longer HBV infected HK-2 cells, the more irregular of the cells' shape, the fewer number of the cells were left. But compared with HBV infected after 24 hours, α-SMA was more expressed after HBV infected 12 hours. After infected by HBV serum in 24 hours.HK-2 cells' proliferation rate was positively correlation in a dose range of LPS, but was negatively correlated with the CLI-095(P＜0.05＝.The levels of HBsAg and HBeAg in cell culture medium were largest when the LPS concentration was at 10μg/ml and CLI-095 at 5μg/ml.The expression of TLR4 significantly increased in HK-2 cells treated with LPS compared with those with CLI-095.but HBV DNA levels and HBsAg and HBeAg expression levels were lower. Conclusions HBV infection may promote cell transdifferentiation and cell injury. The stimulation of HK-2 infected with HBV by LPS may upregulate the expression of TLR4 and reduce the copies of HBV DNA.     

乙型肝炎病毒感染肾小管上皮细胞Toll样受体4的表达及其作用

Objective To explore the expression and role of Toll receptor 4(TLR4)in human proximal tubular epithelial cell line HK-2,infected by HBV. Methods The serum of HBV DNA copies between 107-108/ml was collected. Before and after infected by HBV DNA positive serum. the HK-2 cells' morphology and the expression of α-smooth muscle actin(α-SMA)were observed by microscopy and immunofluorescence, and the effects of different concentrations of lipopolysaccharides(U)S.TLR4-stimulating factor)and CLI-095(TLR4 Inhibitor)on the proliferation rate of HK-2 cells were observed by MTT assays. After HBV serum and 10μg/ml LPS and 5μl/ml CLI-095 acted on HK-2 cells,TLR4 protein expression was measured by immunofluorescence and Western-blotting assay, and HBsAg and HBeAg in cell culture medium were detected by ELISA. and HBV DNA copies by fluorescence quantitative PCR. Results The longer HBV infected HK-2 cells, the more irregular of the cells' shape, the fewer number of the cells were left. But compared with HBV infected after 24 hours, α-SMA was more expressed after HBV infected 12 hours. After infected by HBV serum in 24 hours.HK-2 cells' proliferation rate was positively correlation in a dose range of LPS, but was negatively correlated with the CLI-095(P＜0.05＝.The levels of HBsAg and HBeAg in cell culture medium were largest when the LPS concentration was at 10μg/ml and CLI-095 at 5μg/ml.The expression of TLR4 significantly increased in HK-2 cells treated with LPS compared with those with CLI-095.but HBV DNA levels and HBsAg and HBeAg expression levels were lower. Conclusions HBV infection may promote cell transdifferentiation and cell injury. The stimulation of HK-2 infected with HBV by LPS may upregulate the expression of TLR4 and reduce the copies of HBV DNA.     

乙型肝炎病毒感染肾小管上皮细胞Toll样受体4的表达及其作用

Objective To explore the expression and role of Toll receptor 4(TLR4)in human proximal tubular epithelial cell line HK-2,infected by HBV. Methods The serum of HBV DNA copies between 107-108/ml was collected. Before and after infected by HBV DNA positive serum. the HK-2 cells' morphology and the expression of α-smooth muscle actin(α-SMA)were observed by microscopy and immunofluorescence, and the effects of different concentrations of lipopolysaccharides(U)S.TLR4-stimulating factor)and CLI-095(TLR4 Inhibitor)on the proliferation rate of HK-2 cells were observed by MTT assays. After HBV serum and 10μg/ml LPS and 5μl/ml CLI-095 acted on HK-2 cells,TLR4 protein expression was measured by immunofluorescence and Western-blotting assay, and HBsAg and HBeAg in cell culture medium were detected by ELISA. and HBV DNA copies by fluorescence quantitative PCR. Results The longer HBV infected HK-2 cells, the more irregular of the cells' shape, the fewer number of the cells were left. But compared with HBV infected after 24 hours, α-SMA was more expressed after HBV infected 12 hours. After infected by HBV serum in 24 hours.HK-2 cells' proliferation rate was positively correlation in a dose range of LPS, but was negatively correlated with the CLI-095(P＜0.05＝.The levels of HBsAg and HBeAg in cell culture medium were largest when the LPS concentration was at 10μg/ml and CLI-095 at 5μg/ml.The expression of TLR4 significantly increased in HK-2 cells treated with LPS compared with those with CLI-095.but HBV DNA levels and HBsAg and HBeAg expression levels were lower. Conclusions HBV infection may promote cell transdifferentiation and cell injury. The stimulation of HK-2 infected with HBV by LPS may upregulate the expression of TLR4 and reduce the copies of HBV DNA.     

Induction of macrophage inflammatory cytokines by Ox-LDL is ABCA1 dependent

Objective The current study aimed to evaluate whether the induction of macrophage inflammatory cytokines by Ox-LDL is related to the expression of ABCA 1 pathway. Methods After THP 1/PMA macrophages were transfected with ABCA1 antisense oligonucleotides （100nmol/L） followed by treatment with Ox-LDL （30mg/L）, the expressions of ABCA1, ICAM-1 and MCP-1 mRNA and protein were determined by real-time fluorescent quantitative RT-PCR, Western blot or ELISA. Results Ox-LDL induced expressions of ABCA1, ICAM-1, and MCP-1 at both mRNA and protein levels from THPI/PMA macrophages. Transfection with ABCAI antisense oligonucleotides reduced ABCA1 mRNA levels after 3 and 6 hours and protein levels after 12 and 24 hours. The expression of ICAM-1 and MCP-1 induced by Ox-LDL was also decreased after inhibition of ABCA 1 protein expression by ABCA 1 antisense oligonucleotide decreased. Conclusion The induction of macrophage inflammatory cytokines by Ox-LDL is partially dependent on expression ofABCA1. Our studies disclose new functions of ABCA1 in macrophages Objective The current study aimed to evaluate whether the induction of macrophage inflammatory eytokines by Ox-LDL is related to the expression of ABCA 1 pathway. Methods After THP 1/PMA macrophages were transfected with ABCA1 antisense oligonucleotides （100nmol/L） followed by treatment with Ox-LDL （30mg/L）, the expressions of ABCA1, ICAM-1 and MCP-1 mRNA and protein were determined by real-time fluorescent quantitative RT-PCR, Western blot or ELISA. Results Ox-LDL induced expressions of ABCA1, ICAM-1, and MCP-1 at both mRNA and protein levels from THPI/PMA macrophages. Transfection with ABCAI antisense oligonucleotides reduced ABCA1 mRNA levels after 3 and 6 hours and protein levels after 12 and 24 hours. The expression of ICAM-1 and MCP-1 induced by Ox-LDL was also decreased after inhibition of ABCA 1 protein expression by ABCA 1 antisense oligonucleotide decreased. Conclusion The induction of macrophage inflammatory cytokines by Ox-LDL is partially dependent on expression ofABCA1. Our studies disclose new functions of ABCA1 in macrophages     

英夫利昔治疗瘘管型克罗恩病的研究进展

瘘管是克罗恩病（CD）常见的一种并发症,往往导致患者生活质量明显下降.许多临床试验证实英夫利昔对瘘管型CD安全有效.对传统治疗方案无效者,早期规律给予英夫利昔可很好地控制疾病进展和复发.对单药治疗效果不佳者,联合英夫利昔和内外科治疗可提高疗效.本文就近年英夫利昔治疗瘘管型CD的研究进展作一综述.     

大黄素诱导急性胰腺炎胰腺细胞凋亡机制的实验研究

目的 从凋亡信号传导的角度探讨中药大黄素治疗大鼠急性胰腺炎的分子生物学机制。方法 将44只雄性Wistar大鼠随机分为正常组，非治疗组，大黄素组，以腹腔注射雨蛙肽物方法诱导大鼠急性胰腺炎模型。并于大黄素治疗后6，24，48，72，96小时处死大鼠。应用HE染色比较胰腺组织病理学改变，应用Tunel法检测胰腺细胞凋亡指数，应用RT-PCR技术检测治疗前后凋亡调控基因Bak和BaxmRNA表达。结果 大黄素干预治疗急性胰腺炎后96小时淀粉酶值显低于未治疗组，胰腺细胞凋亡指数显高于未治疗组，凋亡调控基因BakmRNA的表达与未治疗组之间无显性差异。而BaxmRNA的表达显高于未治疗组。结论 大黄素治疗实验性急性胰腺炎的机制可能与干预凋亡调控基因有关。诱导凋亡调控基因Bax表达增强可能是干预凋亡信号传导的重要机制，而与Bak基因表达无关。     

苯并咪唑类药物治疗包虫病研究进展

包虫病主要是由细粒棘球绦虫和多房棘球绦虫的幼虫引起的严重的人兽共患寄生虫病。在我国,由于受手术复发、死亡等手术水平的限制,药物治疗仍然是目前最主要的治疗方法,每年药物治疗的数量远大于手术治疗数。药物在术前缩小病灶、减轻痛苦,延长患者寿命发挥着重要作用。本文综述苯并咪唑类药物及其新剂型的动物及临床治疗包虫病现况,为探索包虫病药物治疗研究的发展方向提供依据。     

Use of Gaming in Self-Management of Diabetes in Teens

With the growing prevalence of diabetes in teens and frequent concomitant problems with adherence, adolescents are a frequent target for diabetes self-management support and education. Due to widespread use of technology among teens in general, the use of serious games, games used for purposes beyond entertainment with the intention to educate and support health behavior for teens with diabetes self-management, is an emerging and promising practice. This report explores games intended for teens with diabetes, how the use of games may enhance clinical practice, and provides suggestions for future research and better utilization of these technologies. Current research on the use of gaming for promoting diabetes management in teens is fairly limited, with some initial support for improvements in both behavioral and clinical outcomes among teens. More research is clearly needed in order to further determine how gaming can best be utilized to impact health outcomes in these teens, as well as potential mechanisms of change.     

莫昔沙星的抗结核作用及其应用研究进展

全球结核病的疫情非常严重,尤其是耐多药结核病不断增加,WHO于20世纪90年代就一再呼吁予以重视。解决此世界性难题的首要措施就是积极落实现代结核病控制策略,亦即直接监视督导下短程化疗（DOTS）,在此基础上要研发新药,使耐多药结核病的治疗和缩短结核病疗程两个跨世纪性课题有所突破^[1]。     

2011年全国血吸虫病监测点疫情分析

目的了解2011年全国81个血吸虫病监测点疫情变化趋势。方法收集2011年全国81个血吸虫病监测点螺情、人畜病情和相关因素的调查数据,分析不同地区疫情特征及变化情况。结果2011年全国血吸虫病监测点血清学检测阳性率以及居民感染率分别为7.78%和0.54%,家畜感染率为0.97%,较2010年均有所下降。2011年81个血吸虫病监测点共查出感染性钉螺面积98.30 hm2,活螺平均密度为0.222 6只/0.1 m2,感染性钉螺平均密度为0.000 3只/0.1 m2,钉螺感染率为0.14%。有螺地带敞放家畜12 422头,比2010年上升46.99%。结论2011年全国血吸虫病监测点疫情较2010年下降不明显。仍需加大家畜传染源控制措施的实施力度。