Cepharanthine induces autophagy via PI3K/AKT/mTOR signaling pathway in ovarian cancer SKOV3 cells

AIM: To investigate the autophagy of human ovarian cancer SKOV3 cells induced by cepharanthine and to explore its mechanism. METHODS: The effect of cepharanthine on the viability of ovarian cancer SKOV3 cells was measured by CCK-8 assay. The SKOV3 cells were treated with cepharanthine, and then the formation of autophagosome was observed with acridine orange staining under fluorescence microscope. The protein levels of LC3, AKT, p-AKT, mTOR, p-mTOR and GAPDH in the SKOV3 cells treated with cepharanthine were determined by Western blot.RESULTS: Cepharanthine significantly inhibited the viability of ovarian cancer SKOV3 cells in a dose-dependent manner (P<0.05). The number of the intracellular acidic autophagosomes with bright red fluorescence was significantly increased after cepharanthine treatment in the SKOV3 cells. The expression of LC3-Ⅱ in SKOV3 cells was significantly enhanced after cepharanthine treatment. Furthermore, treatment with cepharanthine in the SKOV3 cells also resulted in a significant down-regulation of phosphorylated form of AKT and mTOR (P<0.01), while the total protein level was not changed. Combination of cepharanthine and 3-methyladenine resulted in a substantial decrease in the cell viability compared with using cepharanthine alone.CONCLUSION: Cepharanthine significantly inhibits the growth of human ovarian cancer SKOV3 cells and induces the autophagy, which may be correlated with down-regulation of PI3K/AKT/mTOR signaling pathway.     

A pre-synchronization program at early postpartum might increase the chances of Bos indicus cows cycling prior to 50 days regardless of the length of calf separation

The aim of this study was to establish if pre-synchronization would enhance the number of animals cycling prior to conventional breeding at 45 days irrespective of the length of calf separation. Multiparous Bos indicus cows were allotted in four groups (n = 10). Control group (C) dams remained with their calves; groups G24, G48 and G72, which were partially weaned for 24, 48 and 72 h, respectively, were estrus synchronized using a controlled internal drug. These procedures were performed at 25 days and again at 45 days postpartum. The number of follicles, presence of a corpus luteum and back fat thickness (BFT) were determined by ultrasound. The proportion of cows with estrus and ovulation at day 25 postpartum was statistically different between the control and treated groups, with the values being 20, 60, 50 and 70 for the control, G24, G48 and G72 groups respectively (P < 0.05). At days 45 postpartum, the proportion of cows with estrus and ovulation was different in group G48 compared with the other groups (P <0.05). The average BFT and body condition score for the four experimental groups in the two periods were similar (P >0.05). Animals with a higher proportion of follicles from 17 to 21 mm, BFT values above 3.5 mm and a regular body condition were significantly different regardless of whether the dams remained with their calves or were separated, regardless of the length of this event. It can be concluded that (1) a pre-synchronization program at day 25 could trigger the onset of ovarian activity and facilitate a breeding program at day 50 and (2) temporary weaning enhances the effect of a pre-synchronization program.     

Effect of hPTTG1 siRNA on proliferation and apoptosis of ovarian cancer cells

AIM: To observe the proliferation and apoptosis of ovarian cancer cells by silencing the expression of human pituitary tumor-transforming gene 1 ( hPTTG1 ) using RNA interference technique.METHODS: The chemically synthesized siRNA targeting hPTTG1 was transfected into ovarian cancer cell line A2780 in vitro. The expression levels of hPTTG1 and c-myc were examined by RT-PCR and Western blotting. Cell proliferation was measured by MTT colorimetric assay and -TdR incorporation test. Cell apoptosis was detected by flow cytometry with annexin V/PI and TUNEL labeling.RESULTS: The expression of hPTTG1 at mRNA and protein levels was inhibited after transfection of hPTTG1 siRNA. The inhibitory efficiency was 70.5%±3.9% and 63.8%±4.5%, respectively. The absorbance began to decrease 24 h after transfection of hPTTG1 siRNA,and the highest inhibitory rate was 42.9%±5.2% at 48 h post-transfection. Radioactive incorporation of -TdR in hPTTG1 siRNA group was lower than that in normal and negative groups. The survival rate declined while the apoptotic rate and necrotic rate increased in hPTTG1 siRNA group. Apoptotic index in hPTTG1 siRNA group was higher than that in normal and negative groups. The expression of c-myc at mRNA and protein levels was down-regulated.CONCLUSION: Cell proliferation is inhibited and cell apoptosis is induced by hPTTG1 siRNA through down-regulating the expression of c-myc. hPTTG1 can be regarded as a candidate gene for ovarian cancer gene therapy.     

Effect of 3 isoforms of Ikaros on proliferation of human ovarian cancer SKOV3 cells

AIM: To investigate the effect of Ikaros isoforms on the proliferation of human ovarian cancer SKOV3 cells. METHODS: Three isoforms of Ikaros, IK1, IK2 and IK6, were transfected into ovarian cancer SKOV3 cells. CCK-8 assay and cell counting were used to detect the effects of Ikaros isoforms on the proliferation of SKOV3 cells. The cell cycle was analyzed by flow cytometry. The cell cycle-related proteins were detected by Western blot. RESULTS: IK1 and IK2 expression inhibited SKOV3 cells proliferation. Flow cytometry analysis indicated that IK1 and IK2 induced SKOV3 cell cycle arrest at the G₁ phase. IK6 isoform exerted no obvious effect on the proliferation or cell cycle of SKOV3 cells. Compared with control EV group, IK1 group and IK2 group showed a dramatic elevation in the expression of the cell cycle inhibitor p21, along with a substantial decrease in the expression of the cell cycle inducers cyclin D1 and cyclin D2, which did not change in IK6 group. CONCLUSION: IK1 and IK2 significantly inhibit the proliferation of ovarian cancer SKOV3 cells and induce cell cycle arrest at G₁ phase by regulation of cell cycle-related proteins cyclin D1, cyclin D2 and p21, while IK6 isoform exerts no obvious effect on the proliferation and cell cycle of SKOV3 cells.     

体外受精不同时间等因素对体外受精效果的影响

本试验对牛体外受精不同时间、不同的培养液成分和培养方法等对奶牛体外受精后的卵裂率、囊胚发育率的影响进行了研究。试验包括:(1)牛体外受精不同时间(8h、20h)对奶牛体外受精后的卵裂率、囊胚发育率的影响;(2)不同的培养液成分对奶牛体外受精早期胚胎发育的影响。研究结果表明:(1)牛体外受精时间20h对奶牛体外受精后的卵裂率(78%)好于体外受精8h组(76%),但囊胚发育率前者不如后者好(20.51%VS23.68%),两者间差异不显著(P0.05)。(2)作为早期胚胎的培养液IVD101、TCM199培养系的卵裂率分别为76%、74%,而囊胚率却分别为22.37%、22.97%,TCM199培养系好于IVD-101,但两者间差异不显著(P0.05)。     

Inducing synchronous ovarian maturation in the crayfish,Procambarus clarkii,via eyestalk interventional injection as compared with eyestalk ablation and combined injection of serotonin and domperidone

We examined the feasibility of inducing synchronous ovarian maturation in the crayfish, Procambarus clarkii, via eyestalk interventional injection (EI) using a Bletilla striata polysaccharide (BSP) gelatin containing tranexamic acid (TRA), domperidone (DOM) or serotonin (5‐HT). In total, 360 females were randomly assigned to seven experimental groups and a control group. The average survival rate (SR%) and average synchronous ovarian maturation rate (SMR%) in survivors in the EI groups, EI‐TRA, EI‐DOM, EI‐5HT were compared with bilateral eyestalk ablation (BEA), unilateral eyestalk ablation (UEA), abdominal injection (AI) of DOM (0.5 mg crayfish^?1) alone (AI‐DOM) or combined with 5‐HT (0.5 mg crayfish^?1) (AI‐DOM+5‐HT). The experiment covered a prolonged period of 32 days until two ovigerous females were observed in BEA. EI‐DOM achieved a SMR (66.67 ± 9.62%) higher than the control (22.05 ± 3.06%) (P < 0.05), but lower than BEA (88.89 ± 11.11%), UEA (63.33 ± 18.56) and AI‐DOM+5‐HT (59.26 ± 10.14%) (P > 0.05). EI‐DOM also achieved a higher SR (53.33 ± 3.85%) than BEA (15.56 ± 2.22), UEA (24.44 ± 5.88) (P < 0.05) and AI‐DOM+5HT (51.11 ± 4.44) (P > 0.05). However, EI‐TRA (SR = 28.89 ± 4.44, SMR = 37.78 ± 2.22) and EI‐5‐HT (SR = 15.56 ±4.44, SMR = 22.22 ± 11.11) failed to induce significantly higher SMR than the control. These findings suggest that EI methods, such as EI‐DOM, may have positive characteristics for the development of inexpensive and less labour‐intensive techniques for induction of ovarian maturation in decapods.     

西兰苔籽中异硫氰酸盐鉴定及诱导OVCAR-3细胞凋亡研究

[目的]研究西兰苔籽中异硫氰酸盐的抗肿瘤活性。[方法]利用GC—MS联甩仪对西兰苔籽中的异硫氰酸盐进行了分析和鉴定,并通过MTT试验和相差显微镜观察研究了其抗肿瘤活性。[结果]通过GC—MS分析从西兰苔籽提取物中成功鉴定出5种异硫氰酸盐,占挥发性物质的80％。体外细胞毒性试验表明,异硫氰酸盐能有效抑制卵巢癌细胞OVCAR-3的生长,且抑制率随药物浓度的增大而增大,呈明显的剂量关系。异硫氰酸盐浓度为78．3μg/ml时,对OVCAR-3细胞生长的抑制率为50％。对照组的OVCAR-3细胞生长良好,低浓度加药组（10、30、60μg/ml）OVCAR-3细胞的形态和数目发生了变化,圆形脱壁细胞逐渐增多。随着药物浓度的继续增加,OVCAR-3细胞的形态变得很不规则,凋亡小体出现。[结论]该试验为我国天然抗癌药物研究奠定了基础。     

大连地区方氏云鳚卵巢发育的组织学特征及其分期

本文根据1985～1987年在大连地区捕获的方氏云鳚为材料,对其卵母细胞发育各阶段的形态特征进行了描述和探讨。并对方氏云鳚产卵类型及卵子形成过程中生长环的形成等问题进行了阐述。     

Expression of miRNA-22 in ovarian cancer and effect of miRNA-22 over-expression on SKOV-3 ovarian cancer cell proliferation,migration and invasion

AIM: To examine the expression of miRNA-22 in the ovarian tissues and the effect of miRNA-22 over-expression on the proliferation, migration and invasion in SKOV-3 cells. METHODS: The expression levels of miRNA-22 in different ovarian tissues and SKOV-3 cells were determined by qPCR. miRNA-22 was over-expressed by transfection of miRNA-22 mimic. The cell viability was examined by CCK-8 assay. The cell migration was measured by wound healing test. The cell invasion was analyzed by Transwell assay. The protein expression levels of VEGF and P53 were determined by Western blot. RESULTS: Compared with the normal ovarian tissue, the expression level of miRNA-22 was remarkably decreased in the ovarian tumor tissues. After transfection with miRNA-22 mimic, the expression level of miRNA-22 in the SKOV-3 cells was significantly increased, while the cell viability, migration and invasion were obviously decreased. Moreover, the protein expression of VEGF and P53 was dramatically inhibited after over-expression of miRNA-22. CONCLUSION: The decreased miRNA-22 expression may be correlated with the development of ovarian can-cer. Over-expression of miRNA-22 decreases the cell viability, migration and invasion by reducing the protein expression of VEGF and P53.