高效氯氰菊酯对小鼠卵巢生殖功能的影响及维生素E的干预作用

为探究高效氯氰菊酯 (beta-cypermethrin, β-CP) 对雌性小鼠卵巢生殖功能的影响及维生素E (vitamin E, VE) 的干预作用,将雌性昆白小鼠随机分为6 组:空白对照组 (花生油处理)、β-CP不同剂量 (10、20、40 mg/kg) 处理组、VE保护组 (20 mg/kg β-CP+20 mg/kg VE) 和VE组 (20 mg/kg VE),连续灌胃10 d。灌胃结束后取小鼠卵巢组织,观察组织结构的病理变化,采用免疫组化法、蛋白免疫印迹试验及RT-PCR方法检测卵巢中StAR蛋白含量及casp-3、casp-8、INHα和INHβB 基因mRNA表达的变化。结果显示:与对照相比,10、20和40 mg/kg的β-CP处理均使小鼠卵巢组织结构发生了损伤,使组织中StAR蛋白的浓度分别降低了18.8%、36.3%和40.3%,casp-3基因的表达分别升高了16.0%、26.7%和52.9%,INHα基因的表达分别升高了34.5%、83.6%和228.7%,INHβB基因的表达分别升高了7.5%、39.2%和52.7%;20和40 mg/kg的β-CP处理使得casp-8基因的表达分别升高了27.1%和36.7%。上述处理组与对照组的差异均达显著水平 (P＜0.05)。与 20 mg/kg β-CP处理组相比,VE 保护组的StAR蛋白含量也显著增多 (P＜0.05)。研究表明,β-CP对小鼠卵巢具有毒性作用,这与β-CP抑制StAR蛋白的合成,上调casp-3、casp-8、INHα及INHβB基因的表达有关;添加VE对小鼠卵巢有一定的保护作用,这与VE可减弱β-CP对StAR蛋白合成的抑制作用有关。     

矮凤梨短营养生长期突变材料‘14-1’的性状鉴定

菠萝栽培种(Ananas comosus var. comosus)的营养生长期较长,制约了其新品种选育效率。收集和筛选营养生长期较短的种质,可为短营养生长期菠萝新品种的选育创制中间材料。从境外收集了23份易成花的野生种质材料,自2008年起进行栽培、繁殖和性状鉴定。通过对营养生长期的比较和观察,从矮凤梨(A. comosus var. nanus)的体胚苗中发现了1份营养生长期显著缩短的突变材料‘14-1’,其营养生长期约6个月(原品种约12个月),1年开花2次,具有极易成花等特点。‘14-1’无性繁殖后代遗传性状稳定,可作为短营养生长期菠萝育种的亲本资源。     

采用胚挽救获得Ogura CMS甘蓝与甘蓝型油菜的种间杂种

为将甘蓝型油菜细胞质雄性不育系(Ogura CMS)的恢复基因转至甘蓝Ogura CMS材料上,以6份甘蓝Ogura CMS材料为母本,以含恢复基因的甘蓝型油菜为父本,人工杂交结合胚挽救培养,研究取材时期、培养基成分和杂交组合对胚珠成苗的影响,同时对胚挽救培养获得的植株是否为真实F1杂种进行鉴定。结果发现,胚珠培养以剥蕾授粉后16d取材时胚珠成苗数最多,成苗率为4.56%;培养基以MS+GA 0.1 mg·L^-1+NAA 0.1 mg·L^-1+0.5%水解酪蛋白(CH)+0.5%活性炭(AC)成苗效果最好,成苗率高达6.24%;M09CMS×RFO-46组合成苗数最多,成苗率为5.96%。67株胚挽救培养得到的植株经流式细胞仪、SSR分子标记和形态学鉴定,得出65株为真杂种,真杂种率高达97%。     

Effect of proline-spirooxindole on viability and apoptosis of non-small-cell lung cancer A549 cells

AIM:To investigate the effect of proline-spirooxindole on the viability and apoptosis of human non-small-cell lung cancer A549 cells. METHODS:The effect of proline-spirooxindole on the viability of A549 cells was determined by CCK-8 assay. The apoptosis was analyzed by flow cytometry. The effects of proline-spirooxindole on the expression of PARP and p53 and the phosphorylation of mTOR were determined by Western blot. RESULTS:After A549 cells were treated with proline-spirooxindole (25, 50 and 100 mg/L), the cell viability was decreased (P<0.01) compared with DMSO control group. The apoptotic rate was increased compared with DMSO control group (P<0.01). The protein expression of p53 was up-regulated, the increased apoptotic protein cleaved PARP was observed, and the phosphorylation of mTOR was inhibited (P<0.01). CONCLUSION:Proline-spirooxindole inhibits the viability of A549 cells and induces apoptosis, which may be related to the phosphorylation of mTOR.     

Effect of FGFR1 expression knock-down on biological characteristics of infantile hemangioma endothelial cells

AIM: To study the effect of fibroblast growth factor receptor 1 (FGFR1) expression knock-down on the viability, apoptosis, invasion and migration of infantile hemangioma endothelial cells (HemECs). METHODS: FGFR1 was down-regulated by FGFR1 small interfering RNA (si-FGFR1) transfection. The viability of the cells was measured by CCK-8 assay. The apoptotic rate was analyzed by flow cytometry and the invasion and migration abilities were determined by Transwell assay. The protein levels of phosphatidylinositol 3-kinase (PI3K), protein kinase B (AKT), and phosphorylated AKT (p-AKT) were examined by Western blot. RESULTS: Transfection of si-FGFR1 into HemECs had significant effects on inhibiting cell viability (P<0.05), promoting apoptosis (P<0.05), and decreasing cell invasion and migration abilities (P<0.05). The results of Western blot showed that knockdown of FGFR1 gene expression in the cells reduced the protein levels of PI3K and p-AKT (P<0.05), and had no significant effect on AKT protein level. CONCLUSION: Knock-down of FGFR1 expression changes the biological characteristics of endothelial cells in infantile hemangiomas by regulating PI3K/AKT signaling pathway.     

All-trans retinoic acid attenuates blood-brain barrier injury induced by cerebral ischemia-reperfusion in rats through JNK/P38 MAPK signaling pathway

AIM: To investigate the effect of all-trans retinoic acid (ATRA) on blood-brain barrier after cerebral ischemia-reperfusion (CIR) injury in rats and its possible role mechanism.METHODS: Male SD rats were randomly divided into sham group, model (CIR) group and CIR+ATRA (10, 30 and 90 mg/kg) groups. The rat model of CIR injury was established by MCAO thread occlusion method. After ischemia for 1.5 h and reperfusion for 24 h, the neurological functional behavioral score, cerebral infarction volume, brain water content and Evans blue content were determined. The activity of matrix metalloprotein-9 (MMP-9) was measured by gelatin zymography. The protein levels of claudin-5, occludin, ZO-1, JNK, p-JNK, P38, p-P38 and MMP-9 in the brain tissues were determined by Western blot.RESULTS: Compared with CIR model group, ATRA at 30 mg/kg significantly improved neurological function, and decreased cerebral infarction volume, brain water content, Evans blue content and the degradation of tight junction proteins in ischemic area (P<0.01). The activity and protein expression of MMP-9 in ischemic brain tissue were decreased (P<0.01). The phosphorylation of JNK and P38 was inhibited and the protein levels of p-JNK and p-P38 were decreased (P<0.01).CONCLUSION: ATRA reduces the damage of brain tissue and the destruction of blood-brain barrier induced by CIR in rats. The protective effect may be related to inhibiting the activation of JNK/P38 MAPK signaling pathway and MMP-9.     

Role of miR-219 and TGFBR2 in pathogenesis of renal fibrosis

AIM:To study the role of microRNA-219 (miR-219) in regulation of transforming growth factor-β receptor type 2 (TGFBR2) in renal fibrosis. METHODS:The renal fibrosis patients (n=70) were selected in this stu-dy, and 20 cases of healthy people were selected as control group. RT-qPCR was used to detect the expression of miR-219 in the serum of the patients with renal fibrosis and control group, and the expression of miR-219 in NRK49F cells after stimulation with angiotensin Ⅱ(AngⅡ) was detected. The protein expression of α-smooth muscle actin (α-SMA) in the NRK49F cells transfected with miR-219 mimics after stimulation with AngⅡ was determined by Western blot. The potential target gene TGFBR2 of miR-219 was screened and verified by the method of luciferase reporter gene. RT-qPCR and Western blot were used to detected the effect of miR-219 mimics on the expression of TGFBR2 at mRNA and protein levels, and the mRNA expression of α-SMA, connective tissue growth factor (CTGF), type I collagen α1 (COL1A1) and COL3A1 in the NRK49F cells was also detected, respectively. The unilateral ureteral occlusion (UUO) mouse model was established and the expression of miR-219 in the renal tissue was monitored. The morphological change of renal fibrosis was observed in the UUO mice after injection of miR-219, and the mRNA expression levels of COL1A1 and COL3A1 were detected. RESULTS:The expression level of miR-219 in the patients with renal fibrosis was significantly lower than that in control group, and the expression of miR-219 in the UUO mice was decreased significantly (P<0.01). The expression level of miR-219 was significantly decreased in the NRK49F cells after AngⅡ stimulation, and miR-219 mimics inhibited the protein expression of α-SMA(P<0.01). miR-219 mimics had a targeted regulatory effect on TGFBR2 gene, which inhibited the mRNA and protein expression of TGFBR2. miR-219 mimics inhibited the mRNA expression of α-SMA, CTGF, COL1A1 and COL3A1. miR-219 also down-regulated the mRNA expression of COL1A1 and COL3A1 in the UUO mice and inhibited the process of renal fibrosis. CONCLUSION:miR-219 inhibits the development of renal fibrosis by inhibiting the expression of TGFBR2, which may become a new target for the diagnosis and treatment of renal fibrosis.     

Calreticulin expression is necessary for protective immune effect of T-cell vaccine in EAE mice

AIM:To study the role of cell membrane ectopic calreticulin (CALR) expression on the protective immunie effect of T-cell vaccine (TCV) on experimental autoimmune encephalomyelitis (EAE). METHODS:EAE model was established by myelin oligodendrocyte glycoprotein 35-55 (MOG_35-55) immunization in C57BL/6 mice, and the mice were immunized with MOG_35-55-specific CALR⁺ and CALR^- T-lymphocytes. Symptomatic scores were compared at the maximum of the disease. On the 15th day after immunization, the proportion of CD4⁺ CD25⁺ Foxp3⁺ regulatory T cells (Treg) in the spleen, and the expression of interferon-γ (IFN-γ), interleukin-4 (IL-4), IL-10 and IL-17A in the serum were measured. RESULTS:Increased expression of CALR in activated T cells after γ-irradiation was observed. Blockade of CALR on the vaccinating T-cell surface reduced the protective effect of TCV. Furthermore, blockade of CALR reduced the number of Treg in the spleen and up-regulated pro-inflammatory cytokines. CONCLUSION:CALR expression in the T cells is necessary for the protective immunity induced by TCV in EAE mice.     

Metformin sensitizes human breast cancer MDA-MB-231 cells to tamo-xifen through down-regulation of c-Myc

AIM: To investigate whether metformin enhances the sensitivity of human breast cancer MDA-MB-231 cells to tamoxifen by down-regulating c-Myc. METHODS: The cell viability, colony formation, apoptosis, and migration and invasion abilities of MDA-MB-231 cells were detected by CCK-8 assay, colony formation experiment, flow cytometry and Transwell assay. The expression level of c-Myc was quantified by Western blot and immunohistochemical analysis. The antitumor effects of metformin and tamoxifen were investigated in vivo in a MDA-MB-231 triple-negative breast cancer xenograft model in the SCID mice. RESULTS: Metformin in combination with tamoxifen exerted synergistic effects on inhibition of the viability, colony formation, migration and invasion, and induced the apoptosis compared with the controls and either agent treatment alone in the MDA-MB-231 cells. The levels of c-Myc was down-regulated in vitro by treatment with metformin and/or tamoxifen (P<0.01). Moreover, metformin or in combination with tamoxifen also reduced the growth of MDA-MB-231 breast cancer tumors in the SCID mice by down-regulation of c-Myc in vivo. CONCLUSION: Metformin in combination with tamoxifen exerts synergistic effects on inhibition of the proliferation, migration, invasion and tumor growth of human triple-negative breast cancer MDA-MB-231 cells by down-regulating c-Myc expression, suggesting that metformin in combination with tamoxifen merits further evaluation as a target.     

Effect of Jieyuwan on depression-like behaviors and expression of BDNF in hippocampus and prefrontal cortex of WKY rats

AIM:To investigate the mechanism of depression and its development, and to study the expression of brain-derived neurotrophic factor (BDNF) in the hippocampus and prefrontal cortex of Wistar-Kyoto (WKY) rats treated with Jieyuwan. METHODS:Adult male WKY rats were used as an animal model of endogenous depression. Wistar rats of the same strain were selected as control group. WKY rats were randomly divided into model group, citalopram group and Jieyuwan group. After intragastric administration for 21 d, the changes of depression-like behaviors were observed by sucrose preference test and forced swimming test. Immunofluorescence and Western blot were used to detect the expression of BDNF in the hippocampus and prefrontal cortex. RESULTS:WKY rats showed significant depression-like behaviors, and the expression of BDNF was significantly decreased in the hippocampus and prefrontal cortex (P<0.01). The reduction of neuronal axons in hippocampus was also observed. After drug treatment, the depression-like behaviors of WKY rats were significantly attenuated, and the expression of BDNF and the number of axons were increased (P<0.01). CONCLUSION:Jieyuwan effectively attenuates the depression-like behaviors of WKY rats, and BDNF is a key factor in its antidepressant effect. Our findings further confirm the involvement of BDNF in the development of depression.