It is well known that during mammalian ovarian follicular development, the majority of follicles undergo atresia at various stages of their development. However, the mechanisms controlling this selection process remai...It is well known that during mammalian ovarian follicular development, the majority of follicles undergo atresia at various stages of their development. However, the mechanisms controlling this selection process remain unknown. In this study, we investigated apoptosis in granulosa cells during goat follicular atresia by terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL). The changes in the levels of steroids, insulin-like growth factors (IGFs) and IGF receptors were studied by radioimmunoassay (RIA) and semi-quantitative reverse transcrip-tion-PCR. We found that the percentage of apoptotic granulosa cells in the atretic (A) follicles was significantly higher than that in the slightly atretic (SA) and healthy (H) follicles. The level of estradiol and the ratio of estradiol to progesterone in H follicles were significantly higher than those in A follicles. On the other hand, the level of progesterone was not significantly different among these follicle types. We also found that the level of IGF-Ⅰ in H follicles was higher than in SA and A follicles, whereas the amount of IGF-Ⅱ did not vary significantly. The expression of IGF receptor also decreased in A follicles as compared to that in H and SA follicles. These results suggested that estradiol and IGF-Ⅰ might be involved in controlling apoptosis in granulosa cells during follicular atresia.展开更多
To construct the recombinant expression functionally in Chinese hamster ovary cells in order vector for CD80-IgG fusion gene and to express it to be used as an effective method to eliminate the immune escape of leukem...To construct the recombinant expression functionally in Chinese hamster ovary cells in order vector for CD80-IgG fusion gene and to express it to be used as an effective method to eliminate the immune escape of leukemic cells, the cDNA encoding the signal and extracellular domains of murine CD80 was generated by PCR amplification from plasmid pcDNMB7 containing the full length cDNA of murine CD80 and those of murine IgG1, in which the Fc fragment was obtained through RT-PCR amplification from murine spleen cells. These two cDNAs were then cloned in tandem into eukaryotic expression vector pcDNA3.0 and the resultant recombinant plasmid pcDNA/CD80-IgG was then transfected to Chinese hamster ovary cells with liposome transfection reagent. The cell clones constitutively expressing CD80-IgG fusion protein were obtained by G418 screening. Western blotting and dot ELISA assay were used to detect the expression of the fusion protein in the supernatants of these cells. Meanwhile, the fusion protein expressed was then purified with affinity chromatography, and its biological activity was demonstrated by flow cytometry, MTr colorimetry and ELISA assay. The experimental resuits showed that these two inserts were successfully cloned into plasmid pcDNA3.0, and the highly purified fusion protein was obtained. This fusion protein was proved to be able to upregulate the density of CD80 on leukemic cells, deliberately promote the proliferative reactions of mouse allogenic lymphocytes and increase the killing activity against WEHI-3 cells from 49.7 % up to 84.6 %. In addition, this fusion protein could also enhance the IL-2 secretion from allogenic lymphocytes activated by tumorspecific antigens. It is concluded that the recombinant vector constructed can be functionally expressed in the mammalian cells, thus providing a solid foundation for the further investigation on the mechanism to eliminate the immune escape of leukemic cells in vivo.展开更多
t Objective： The aim of our study was to investigate the expression of guanine nucleotide exchange factor Dock180 in ovarian tumor, and its significance in the initiation and progression of ovarian cancer. Methods： ...t Objective： The aim of our study was to investigate the expression of guanine nucleotide exchange factor Dock180 in ovarian tumor, and its significance in the initiation and progression of ovarian cancer. Methods： Immunohistochemical staining with SP method was conducted to identify the expression of Dock180 protein in epithelial ovarian tumor in 68 cases. Results： Dock180 present with higher expression in ovarian cancer, as compared with than that in low malignant tumor and benign ovarian tumor （P 〈 0,01）, In ovarian cancer, Dock180 expression was increased with the increased FIGO stage and grade. Conclusion： Dock180 overexpression may play an important role in the development and progression of ovarian cancer and it could be used as a new measurement of malignant biological behavior of ovarian cancer.展开更多
OBJECTIVE Epidermal growth factor receptor (EGFR) isdysregulated in many human malignancies and is a potentialtarget for therapeutic intervention, but there is a majordisagreement among researchers about both the freq...OBJECTIVE Epidermal growth factor receptor (EGFR) isdysregulated in many human malignancies and is a potentialtarget for therapeutic intervention, but there is a majordisagreement among researchers about both the frequency andpossible clinical importance of EGFR overexpression in ovariancancer. We investigated the expression and significance of theEGFR mRNA and vascular endothelial growth factor (VEGF)mRNA in ovarian carcinoma.METHODS Reverse transcription polymerase chain reaction(RT-PCR) was employed to determine the expression of EGFRmRNA and VEGF mRNA in 79 ovarian specimen (including15 normal, 13 benign and 51 malignant, from 79 patients). Therelationship between EGFR and VEGF expression was analyzed.RESULTS The positive rates of the expression of EGFR mRNAand VEGF mRNA were significantly higher in the patientswith ovarian carcinoma than those in both the patients withbenign ovarian tumors and in the normal controls. There wascorrelation between EGFR mRNA expression and clinical stages.The positive rate of the expression of EGFR mRNA in Stage Ⅲ-Ⅳ was higher than that in Stage Ⅰ-Ⅱ of ovarian carcinoma (P <0.05). The expression of VEGF mRNA was correlated with theclinical stages and lymph node metastasis. The expression levelsof VEGF mRNA in Stage Ⅲ-Ⅳ and in the group with lymph nodemetastasis were significantly higher than those in Stage Ⅰ-Ⅱ and inthe group without lymph node metastasis, respectively (P < 0.05).The expression of EGFR mRNA was positively correlated with theexpression of VEGF mRNA (r = 0.438, P < 0.05).CONCLUSION The expressions of EGFR mRNA and VEGFmRNA are positively correlated to the occurrence of ovariancarcinoma and its metastasis. The detection of EGFR and VEGFmay be helpful for the targeted chemotherapy.展开更多
目的:观察妇科养荣胶囊(Fu Ke Yang Rong capsule,FKYRC)对卵巢损伤小鼠卵巢储备功能和生育力的保护作用。方法:通过成年雌性小鼠一次性腹腔注射120 mg/kg环磷酰胺和10 mg/kg白消安建立卵巢损伤小鼠模型,于造模前7 d至造模后60 d每日对...目的:观察妇科养荣胶囊(Fu Ke Yang Rong capsule,FKYRC)对卵巢损伤小鼠卵巢储备功能和生育力的保护作用。方法:通过成年雌性小鼠一次性腹腔注射120 mg/kg环磷酰胺和10 mg/kg白消安建立卵巢损伤小鼠模型,于造模前7 d至造模后60 d每日对卵巢损伤小鼠用高、中、低剂量(6 g/kg、4 g/kg和2 g/kg)FKYRC(H组、M组、L组)连续灌胃给药;同时,于造模前7 d一次性皮下注射给予Gn RHa(38 mg/kg)作为阳性对照组(Gn RHa组),用生理盐水(0.2 m L/d)代替FKYRC连续灌胃作为卵巢损伤对照组(模型组),另设正常对照组:非卵巢损伤正常小鼠腹腔注射给予等体积DMSO(NC组)。分别于造模后30 d和60 d取材,计数卵巢组织中各级卵泡数量,检测血清E2、FSH、抑制素B(INHB)和抗苗勒氏管激素(AMH)水平,检测卵巢组织中翼状螺旋/叉头转录因子2(FOXL2)和AMH蛋白的表达水平。并观察造模60 d后各组妊娠率和窝仔数。结果:各给药组和模型组小鼠血清E2低于NC组(P<0.05),但FSH水平组间无统计学差异(P>0.05)。造模各组卵泡数明显低于NC组(P<0.05),且H组和Gn RHa组造模后60 d时卵巢组织中窦前卵泡与窦卵泡数较高,但各干预组与模型组比较无统计学差异(P>0.05)。模型组小鼠卵巢组织中FOXL2和AMH蛋白表达水平显著低于各FKYRC干预组(P<0.05),H组的妊娠率(80.00%)显著高于模型组(36.36%)(P<0.05)。结论:连续FKYRC(6 g/kg)给药60 d后可明显改善烷化剂所致卵巢损伤小鼠的卵巢储备功能和生育能力,其机制可能与上调颗粒细胞中FOXL2和AMH的表达水平相关。展开更多
基金supported by the National Natural Science Foundation of China(No.30070556)the Major State Basic Research Development Program of China(No.G200016107).
文摘It is well known that during mammalian ovarian follicular development, the majority of follicles undergo atresia at various stages of their development. However, the mechanisms controlling this selection process remain unknown. In this study, we investigated apoptosis in granulosa cells during goat follicular atresia by terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL). The changes in the levels of steroids, insulin-like growth factors (IGFs) and IGF receptors were studied by radioimmunoassay (RIA) and semi-quantitative reverse transcrip-tion-PCR. We found that the percentage of apoptotic granulosa cells in the atretic (A) follicles was significantly higher than that in the slightly atretic (SA) and healthy (H) follicles. The level of estradiol and the ratio of estradiol to progesterone in H follicles were significantly higher than those in A follicles. On the other hand, the level of progesterone was not significantly different among these follicle types. We also found that the level of IGF-Ⅰ in H follicles was higher than in SA and A follicles, whereas the amount of IGF-Ⅱ did not vary significantly. The expression of IGF receptor also decreased in A follicles as compared to that in H and SA follicles. These results suggested that estradiol and IGF-Ⅰ might be involved in controlling apoptosis in granulosa cells during follicular atresia.
文摘To construct the recombinant expression functionally in Chinese hamster ovary cells in order vector for CD80-IgG fusion gene and to express it to be used as an effective method to eliminate the immune escape of leukemic cells, the cDNA encoding the signal and extracellular domains of murine CD80 was generated by PCR amplification from plasmid pcDNMB7 containing the full length cDNA of murine CD80 and those of murine IgG1, in which the Fc fragment was obtained through RT-PCR amplification from murine spleen cells. These two cDNAs were then cloned in tandem into eukaryotic expression vector pcDNA3.0 and the resultant recombinant plasmid pcDNA/CD80-IgG was then transfected to Chinese hamster ovary cells with liposome transfection reagent. The cell clones constitutively expressing CD80-IgG fusion protein were obtained by G418 screening. Western blotting and dot ELISA assay were used to detect the expression of the fusion protein in the supernatants of these cells. Meanwhile, the fusion protein expressed was then purified with affinity chromatography, and its biological activity was demonstrated by flow cytometry, MTr colorimetry and ELISA assay. The experimental resuits showed that these two inserts were successfully cloned into plasmid pcDNA3.0, and the highly purified fusion protein was obtained. This fusion protein was proved to be able to upregulate the density of CD80 on leukemic cells, deliberately promote the proliferative reactions of mouse allogenic lymphocytes and increase the killing activity against WEHI-3 cells from 49.7 % up to 84.6 %. In addition, this fusion protein could also enhance the IL-2 secretion from allogenic lymphocytes activated by tumorspecific antigens. It is concluded that the recombinant vector constructed can be functionally expressed in the mammalian cells, thus providing a solid foundation for the further investigation on the mechanism to eliminate the immune escape of leukemic cells in vivo.
基金Supported by grants from the National Natural Science Foundation of China (No. C30772330)the Natural Science Foundation Project of Chongqing (No. 2010BB5387)partly by the Chongqing Municipal Health Bureau Foundation Project (No. 2010-2-062)
文摘t Objective： The aim of our study was to investigate the expression of guanine nucleotide exchange factor Dock180 in ovarian tumor, and its significance in the initiation and progression of ovarian cancer. Methods： Immunohistochemical staining with SP method was conducted to identify the expression of Dock180 protein in epithelial ovarian tumor in 68 cases. Results： Dock180 present with higher expression in ovarian cancer, as compared with than that in low malignant tumor and benign ovarian tumor （P 〈 0,01）, In ovarian cancer, Dock180 expression was increased with the increased FIGO stage and grade. Conclusion： Dock180 overexpression may play an important role in the development and progression of ovarian cancer and it could be used as a new measurement of malignant biological behavior of ovarian cancer.
文摘OBJECTIVE Epidermal growth factor receptor (EGFR) isdysregulated in many human malignancies and is a potentialtarget for therapeutic intervention, but there is a majordisagreement among researchers about both the frequency andpossible clinical importance of EGFR overexpression in ovariancancer. We investigated the expression and significance of theEGFR mRNA and vascular endothelial growth factor (VEGF)mRNA in ovarian carcinoma.METHODS Reverse transcription polymerase chain reaction(RT-PCR) was employed to determine the expression of EGFRmRNA and VEGF mRNA in 79 ovarian specimen (including15 normal, 13 benign and 51 malignant, from 79 patients). Therelationship between EGFR and VEGF expression was analyzed.RESULTS The positive rates of the expression of EGFR mRNAand VEGF mRNA were significantly higher in the patientswith ovarian carcinoma than those in both the patients withbenign ovarian tumors and in the normal controls. There wascorrelation between EGFR mRNA expression and clinical stages.The positive rate of the expression of EGFR mRNA in Stage Ⅲ-Ⅳ was higher than that in Stage Ⅰ-Ⅱ of ovarian carcinoma (P <0.05). The expression of VEGF mRNA was correlated with theclinical stages and lymph node metastasis. The expression levelsof VEGF mRNA in Stage Ⅲ-Ⅳ and in the group with lymph nodemetastasis were significantly higher than those in Stage Ⅰ-Ⅱ and inthe group without lymph node metastasis, respectively (P < 0.05).The expression of EGFR mRNA was positively correlated with theexpression of VEGF mRNA (r = 0.438, P < 0.05).CONCLUSION The expressions of EGFR mRNA and VEGFmRNA are positively correlated to the occurrence of ovariancarcinoma and its metastasis. The detection of EGFR and VEGFmay be helpful for the targeted chemotherapy.
