Apoptosis and its modulation in hepatic ischemia and reperfusion injury

Hepatic ischemia/reperfusion injury (HIRI) exists in lots of process of clinical pathology and operation. Apoptosis is an active process controlled by some gene and factors such as Fas/FasL, caspases and Bcl-2 families. More and more studies suggest that HIRI is associated with apoptosis. This article summarized the mechanisms and gene modulation of apoptosis during HIRI and the significance of suppressing apoptosis in preventing HIRI.     

Expression of human inducible nitric oxide synthase in fibroblast cell line V79 and effects of H4Bon iNOS activity

AIM: To explore the feasibility of human iNOS transfected into V₇₉ cells by gene transfer and investigate the effects of H₄B on iNOS activity. METHODS: Human iNOS was transfected into V₇₉ cells with the karyocyte expressive vector. The cloned cells were selected by G₄₁₈. The expression of iNOS mRNA was quantified by RT-PCR and iNOS expression was observed by immunofluorescence. NO product in cells was determined by measuring nitrite (NO^-₂) release using the Griess reaction. RESULTS: V₇₉ cells infected human iNOS was proved to have iNOS mRNA at 462 bp by RT-PCR, and iNOS protein in the cytochylema by immunofluorescence. When the cells were incubated without H₄B, the content of NO in pcDNA₃ cells was minimal, with NO^-₂ production (82.32±13.08) just above the normal group (74 38±9 80, P>0.05, n=6) There was no significant difference between pcDNA₃ cells incubated with or without H₄B, (P>0.05, n=6) NO^-₂ production by pcDNA₃-iNOS cells without H₄B was higher (105 58±13 33) (n=6, P<0.01vs the normal cells or pcDNA cells). However, in pcDNA₃-iNOS cells incubated with H₄B, NO^-₂ production was much higher (236 57±3183) (n=6, P<0.01vs the all former groups). CONCLUSION: iNOS activity was increased by adding H₄B in pcDNA₃-iNOS cells, and the fibroblast can be a target cell of iNOS gene transfer.     

Relationship between thromboxane synthase gene and prostacyclin synthase gene expression and pulmonary hypertension induced by hypoxic hypercapnia

AIM: To study the effect of chronic hypoxic hypercapnia on gene expression of thromboxane synthase and prostacyclin synthase in pulmonary arterioles. METHODS: Sprague-Dawley rats were randomly divided into two groups: control group and hypoxic hypercapnic group. TXS mRNA and PGI₂-SmRNA were observed in pulmonary arterioles by in situ hybridization. RESULTS: mPAP, weight ratio of right ventricle (RV) to left ventricle plus septum(LV+S), contents of TXB₂ and 6-keto-PGF1_α in plasma and lung and TXS mRNAin pulmonary arterioles were much higher in rats of hypoxic hypercapnic group than those of control group. Differences of PGI₂-SmRNA in pulmonary arterioles were not significant in two groups. Light microscopy showed hypertrophy of vessel smooth muscle cells and vessel cavity straitness were found in hypoxic hypercapnic group. CONCLUSION: Changes of gene expressions of thromboxane synthase and prostacyclin synthase and imbalance of TXA₂/PGI₂ may play an important role in hypoxic hypercapnic pulmonary hypertension.     

Effect of CLCN2 antisense oligonucleotide on U251 cells injury by cisplatin

AIM: To investigate the effect of chloride channel CLCN2 antisense oligonucleotide on the cell injury of malignant U251 glioma cells induced by cisplatin (DDP). METHODS: The experiment was divided into 4 groups: control group (nonsense oligonucleotide), CLCN2 antisense oligonucleotide group, DDP group (DDP＋nonsense oligonucleotide), DDP＋CLCN2 antisense oligonucleotide group. The viability of U251 cells was measured by MTT assay, CLCN2 mRNA level was determined by RT-PCR, cell apoptosis was measured by TUNEL assay. RESULTS: Compared to the control group, the cell viability, CLCN2 and cyclinD1 mRNA decreased in CLCN2 antisense oligonucleotide group, DDP treated group and CLCN2 antisense oligonucleotide with DDP treated group, cells apoptosis increased. Compared to DDP group, the cell viability (P<0.05) and CLCN2 mRNA decreased in CLCN2 antisense oligonucleotide with DDP treated group, and cells apoptosis increased (P<0.01). Compared to CLCN2 antisense oligonucleotide group, CLCN2 mRNA significantly decreased (P<0.01) in CLCN2 antisense oligonucleotide with DDP treated group. CONCLUSION: CLCN2 antisense oligonucleotide inhibits the expression of CLCN2 mRNA in U251 cells. Inhibition of CLCN2 mRNA facilitates the cell injury of U251 cells induced by DDP. The decrease in CLCN2 mRNA is involved in the mechanism of cell injury by DDP.     

大黑鳃金龟消化与解毒相关基因的组织表达研究

[目的]明确大黑鳃金龟幼虫消化酶和解毒相关基因在不同组织中的表达情况,筛选中肠特异表达基因。[方法]利用RT-PCR方法对大黑鳃金龟幼虫中肠消化酶(胰蛋白酶、氨肽酶、羧肽酶)基因和解毒相关基因(细胞色素P450、谷胱甘肽S-转移酶、酯酶)在不同组织中的表达进行分析。[结果]所调查的大黑鳃金龟的6个谷胱甘肽S-转移酶在中肠中都有表达,有3个基因在中肠中特异性表达。细胞色素P450家族中CYP 6A20和CYP 4C1 2个基因在中肠中特异表达,3个氨肽酶基因(APN1、APN2和AP2)只在中肠中表达。[结论]大部分大黑鳃金龟的消化酶和解毒酶基因在中肠中表达量最高。     

外源基因对小单孢菌40027菌株产生抗生素的影响

研究了外源基因对小单孢菌40027菌株产生抗生素水平的影响。结果表明:整合了质粒pSET152和质粒pHZ1904的小单孢菌40027菌株比原始菌株产生福堤霉素A的产量低,在产素时间上缩短;整合了质粒pSET152和pHZ1904小单孢菌40027菌株之间在产素水平和产素时间上没有显著差异。表明整合了质粒pSET152和pHZ1904的小单孢菌40027菌株与原始菌株在产素水平和产素时间上的差异是由质粒pSET152造成的,dnd基因簇对小单孢菌40027菌株的产素水平和产素时间没有影响。     

植物叶片衰老研究进展

叶片衰老是植物叶片生长发育过程中衰变的重要信号,在此综述了植物叶片衰老的特性、机理、与叶片衰老相关的基因、影响叶片衰老的因子(即器官、植物激素、氧化胁迫等),同时又介绍了叶片衰老相关基因表达的调控,并对开展植物叶片衰老研究的综合发展趋势作了分析。     

大豆三系的选育及恢复基因的SSR初步定位研究

以栽培大豆ZD8319为母本,分别与SG01、JX03和PI004杂交,再用父本回交5次,选育出阜CMS1A、阜CMS2A和阜CMS3A三个高度不育的不育系。正反交结果表明,这3个不育系为质核互作不育系（CMS）,花粉为典败型不育,且不育性稳定。与3个不育系广泛测交,筛选与不育系配合力高、育性恢复力强的强优势组合,从中鉴定出恢复度高的恢复系5个（蒙-06,YC04,阜84-5-4,ZY9010,Z9001）,使栽培大豆质核互作不育系三系配套。通过测交发现,阜CMS1A、阜CMS2A比阜CMS3A育性易被恢复。用这2个不育系和5个恢复系的核基因组DNA为模板,进行简单重复序列（simple sequence repeat, SSR）分析,从60对核心引物中筛选出在2个不育系和5个恢复系间有特异带的SSR引物,分别为Satt143、Satt168、Satt441。表明这3个SSR标记是与恢复基因有关的3个等位位点。并把恢复基因初步定位于2个分离群体上的6个连锁群上。     

The construction of bicistronic eukaryotic vector carrying green fluorescent protein and hytk gene and its expression in bladder carcinoma

AIM: To facilitate the suicide gene delivery into neoplasm, a chimeric gene of HSV-tk and green fluorescent protein (gfp) was constructed. METHODS: Molecular cloning technique was used to construct this kind of eukaryotic vector. The internal ribosome entry site (IRES) of encephalomyocarditis virus (EMCV), which could coordinate expression of two genes in a single vector, was optioned. By using liposome-mediated transfection, eukaryotic expression vector tgCMV/hytk-IRES-gfp was transfected into human bladder carcinoma cells EJ. RESULTS: A bicistronic eukaryotic vector carrying gfp and hygromycin phosphotransferase-thymidine kinase fusion (hytk) gene was constructed. The results of PCR and microscopy detection show that the hytk-IRES-gfp gene was successfully transferred into EJ cells. There were no differences in the growth pattern or the morphology between EJ and EJ/hytk-GFP cells. In vitro experiments demonstrated dose- and time-dependent cell killing by transduction of the hytk-IRES-gfp gene followed by GCV treatment. The IC50 (the concentration required to elicit 50% growth inhibition) was 2.16 mg/L in treatment with GCV for 72 hours. CONCLUSION: These results suggest that this new kind of eukaryotic vector could serves as a new tool and method for neoplasm gene therapy.     

Effect of L-arginine on expression of bcl-2 and bax mRNA in pulmonary injury induced by ischemia-reperfusion in rabbits

AIM:To investigate the effect of L-arginine (L-Arg) on expression of bcl-2, bax mRNA during pulmonary ischemia and reperfusion injury (PIRI) in rabbits.METHODS:Single lung ischemia and reperfusion animal model was used in vivo. The rabbits were randomly divided into three groups: sham operated group (sham, n=12), ischemia-reperfusion group (I/R, n=12) and I/R+ L-arginine group (L-Arg, n=12). Changes of several parameters, which included apoptotic index (AI), wet to dry ratio of lung tissue weight (W/D) and index of quantitative assessment of histologic lung injury (IQA), were measured at 300 min after reperfusion in lung tissue. Meanwhile the location and expression of bcl-2, bax mRNA as well as the ratio of bcl-2 mRNA/bax mRNA were observed. The lung tissue was prepared for light microscopic and electron microscopic observation at 60, 180 and 300 min after reperfusion. RESULTS:As compared with I/R group, in intima and extima of small pulmonary artery, alveoli, and bronchiole epithelia, the expression of bcl-2 mRNA and the ratio of bcl-2 mRNA/bax mRNA were increased, and the expression of bax mRNA was decreased in L-Arg treatment group. The values of AI, W/D and IQA showed significantly lower than that in I/R group at 180 minutes after reperfusion in lung tissue (P<0.01 and P<0.05). Meanwhile, abnormal changes of the lung tissue in morphologically were markedly lessened in L-Arg treatment group.CONCLUSION:L-arginine produces a notable protective effect on PIRI in rabbits by up-regulating bcl-2 mRNA expression, down-regulating bax mRNA expression in lung tissue and regulating the balance of bcl-2 mRNA and bax mRNA to decrease apoptosis.