RGMb蛋白是反义导向分子(repulsive guidance molecule,RGM)家族成员之一,可在细胞水平上介导骨形态发生蛋白(bone morphogenetic protein,BMP)的信号通路。大量研究报道,RGMb参与调控细胞的增殖、分化、凋亡以及细胞间的黏附能力。本...RGMb蛋白是反义导向分子(repulsive guidance molecule,RGM)家族成员之一,可在细胞水平上介导骨形态发生蛋白(bone morphogenetic protein,BMP)的信号通路。大量研究报道,RGMb参与调控细胞的增殖、分化、凋亡以及细胞间的黏附能力。本研究旨在探讨RGMb对子宫内膜腺细胞的作用及其分子机制。应用siRNA与过表达技术处理子宫内膜腺细胞系(Ishikawa),采用qPCR与Western印迹技术确定其转染效率,以及BMP相关信号通路(MAPK与Smad)成员的表达。结果显示:下调RGMb基因的表达显著降低了p-ERK1/2(1.861±0.1864 vs 0.885±0.0869,P=0.0090)与p-Smad1/5/8(1.624±0.1238 vs 1.093±0.0890,P=0.0253)的表达水平。过表达RGMb基因显著升高了p-ERK1/2(1.237±0.1114 vs 2.089±0.1658,P=0.0130)与p-Smad1/5/8(1.139±0.0562 vs 1.98±0.1449,P=0.0056)的表达水平。而RGMb基因下调和过表达对p-P38 MAPK蛋白表达水平均无显著影响(P>0.05)。采用CCK-8和qPCR技术检测RGMb对Ishikawa细胞增殖活力及增殖相关基因的影响。结果显示:转染80 nmol/L RGMb siRNA显著降低Ishikawa细胞的增殖活力(0.479±0.0271 vs 0.3487±0.0094,P=0.0104),同时降低增殖相关基因CCND1(1±0.0366 vs 0.6719±0.0236,P=0.0017)和CDK2(1±0.0370 vs 0.853±0.0135,P=0.0202)表达水平;转染1μg/mL RGMb基因过表达质粒显著提高Ishikawa细胞增殖活力(0.283±0.0030 vs 0.3714±0.0140,P=0.0001),同时增加CCND1(1±0.0178 vs 1.375±0.0356,P=0.0007)和CDK2(1±0.0188 vs 1.376±0.0513,P=0.0023)基因的表达水平。以上结果表明,RGMb可能通过p-ERK1/2与p-Smad1/5/8影响Ishikawa细胞增殖活力,为进一步研究RGMb调控子宫机能的分子机制提供科学依据。展开更多
[Objective] The experiment aimed to set up a method for isolating and culturing endometrial stromal cells (BESC) and endometrial glandular epithelial cells(BEGEC) of buffalo as well as laid foundation for studying bio...[Objective] The experiment aimed to set up a method for isolating and culturing endometrial stromal cells (BESC) and endometrial glandular epithelial cells(BEGEC) of buffalo as well as laid foundation for studying biological mechanism of embryo implantation and uterine diseases. [Method] The enzymatic digestion method, scraping method, serial filtration and differential velocity adherent technique were used to isolate BESC and BEGEC, then immunocytochemical method and TRYPAN-Blue assay were used to determine the purity and survival rate of isolated cells. [Result] The BESC and BEGEC were successfully isolated and cultured while immunocytochemical method and cell count method demonstrated that the purity was over 90%. The result of TRYPAN-Blue assay shown that survival rate of BESC and BEGEC was 91% and 78% respectively. [Conclusion] The enzymatic digestion method, scraping method, serial filtration and differential velocity adherent technique could isolate BESC and BEGEC with high purity.展开更多
Objective: To observe the effects of Bushenyiqihexue Formula (补肾益气和血方 Formula for Tonifying the Kidney, Replenishing qi and Harmonizing Blood, FTKRQHB) on the endometrial gland apoptosis in the mice with blasto...Objective: To observe the effects of Bushenyiqihexue Formula (补肾益气和血方 Formula for Tonifying the Kidney, Replenishing qi and Harmonizing Blood, FTKRQHB) on the endometrial gland apoptosis in the mice with blastocyst implantation dysfunction. Methods: The mice with the first-day pregnancy were divided into the control, model and treatment groups, with 30 in each group, and blastocyst implantation dysfunction was induced by subcutaneous injection of mifepristone in the mice of the model and treatment groups. The pregnancy rate and implantation number of blastocysts were measured and the expressions of proliferating cell nuclear antigen (PCNA), Bax, Bcl-2, and activated caspase-3 were detected in all the three groups. Results: The model group had significantly depressed pregnancy rate, implantation number of blastocysts and apoptosis index, and elevated proliferation index of endometrial gland as compared with the control group (P<0.05 or P<0.01). Administration of FTKRQHB (the treatment group) resulted in significant increases in pregnancy rate, implantation number of blastocysts and apoptosis index of the endometrial gland, and a significant decrease in the proliferation index of the endometrial gland as compared with the model group (P<0.05 or P<0.01). The differences in the four indexes between the treatment group and control group were not significant statistically. The Bax and activated caspase-3 expressions in endometrial gland in the model group became significantly lower than that of the control group (P<0.01), whereas those in the treatment group were significant higher than that of the model group (P<0.01). However, the Bax and activated caspase-3 expressions in endometrial gland were similar in both treatment and control groups. Conclusion: Promoting the increases in Bax and activated caspase-3 expressions in the endometrial gland and bringing into balance between apoptosis and proliferation of the glandular cells at the implantation window phase by FTKRQHB may contribute to the effects of promoting the establishment of endometrial receptivity and improving blastocyst implantation dysfunction.展开更多
文摘RGMb蛋白是反义导向分子(repulsive guidance molecule,RGM)家族成员之一,可在细胞水平上介导骨形态发生蛋白(bone morphogenetic protein,BMP)的信号通路。大量研究报道,RGMb参与调控细胞的增殖、分化、凋亡以及细胞间的黏附能力。本研究旨在探讨RGMb对子宫内膜腺细胞的作用及其分子机制。应用siRNA与过表达技术处理子宫内膜腺细胞系(Ishikawa),采用qPCR与Western印迹技术确定其转染效率,以及BMP相关信号通路(MAPK与Smad)成员的表达。结果显示:下调RGMb基因的表达显著降低了p-ERK1/2(1.861±0.1864 vs 0.885±0.0869,P=0.0090)与p-Smad1/5/8(1.624±0.1238 vs 1.093±0.0890,P=0.0253)的表达水平。过表达RGMb基因显著升高了p-ERK1/2(1.237±0.1114 vs 2.089±0.1658,P=0.0130)与p-Smad1/5/8(1.139±0.0562 vs 1.98±0.1449,P=0.0056)的表达水平。而RGMb基因下调和过表达对p-P38 MAPK蛋白表达水平均无显著影响(P>0.05)。采用CCK-8和qPCR技术检测RGMb对Ishikawa细胞增殖活力及增殖相关基因的影响。结果显示:转染80 nmol/L RGMb siRNA显著降低Ishikawa细胞的增殖活力(0.479±0.0271 vs 0.3487±0.0094,P=0.0104),同时降低增殖相关基因CCND1(1±0.0366 vs 0.6719±0.0236,P=0.0017)和CDK2(1±0.0370 vs 0.853±0.0135,P=0.0202)表达水平;转染1μg/mL RGMb基因过表达质粒显著提高Ishikawa细胞增殖活力(0.283±0.0030 vs 0.3714±0.0140,P=0.0001),同时增加CCND1(1±0.0178 vs 1.375±0.0356,P=0.0007)和CDK2(1±0.0188 vs 1.376±0.0513,P=0.0023)基因的表达水平。以上结果表明,RGMb可能通过p-ERK1/2与p-Smad1/5/8影响Ishikawa细胞增殖活力,为进一步研究RGMb调控子宫机能的分子机制提供科学依据。
基金Supported by the Innovation Foundation For Postgraduate of Guangxi University(2008105930905D001) the Tackle Key Program in Science and Technology of Science and Technology Bureau of Guangxi Province(0815008-2-4)~~
文摘[Objective] The experiment aimed to set up a method for isolating and culturing endometrial stromal cells (BESC) and endometrial glandular epithelial cells(BEGEC) of buffalo as well as laid foundation for studying biological mechanism of embryo implantation and uterine diseases. [Method] The enzymatic digestion method, scraping method, serial filtration and differential velocity adherent technique were used to isolate BESC and BEGEC, then immunocytochemical method and TRYPAN-Blue assay were used to determine the purity and survival rate of isolated cells. [Result] The BESC and BEGEC were successfully isolated and cultured while immunocytochemical method and cell count method demonstrated that the purity was over 90%. The result of TRYPAN-Blue assay shown that survival rate of BESC and BEGEC was 91% and 78% respectively. [Conclusion] The enzymatic digestion method, scraping method, serial filtration and differential velocity adherent technique could isolate BESC and BEGEC with high purity.
基金supported by the National Natural Science Foundation of China (No. 30171193)
文摘Objective: To observe the effects of Bushenyiqihexue Formula (补肾益气和血方 Formula for Tonifying the Kidney, Replenishing qi and Harmonizing Blood, FTKRQHB) on the endometrial gland apoptosis in the mice with blastocyst implantation dysfunction. Methods: The mice with the first-day pregnancy were divided into the control, model and treatment groups, with 30 in each group, and blastocyst implantation dysfunction was induced by subcutaneous injection of mifepristone in the mice of the model and treatment groups. The pregnancy rate and implantation number of blastocysts were measured and the expressions of proliferating cell nuclear antigen (PCNA), Bax, Bcl-2, and activated caspase-3 were detected in all the three groups. Results: The model group had significantly depressed pregnancy rate, implantation number of blastocysts and apoptosis index, and elevated proliferation index of endometrial gland as compared with the control group (P<0.05 or P<0.01). Administration of FTKRQHB (the treatment group) resulted in significant increases in pregnancy rate, implantation number of blastocysts and apoptosis index of the endometrial gland, and a significant decrease in the proliferation index of the endometrial gland as compared with the model group (P<0.05 or P<0.01). The differences in the four indexes between the treatment group and control group were not significant statistically. The Bax and activated caspase-3 expressions in endometrial gland in the model group became significantly lower than that of the control group (P<0.01), whereas those in the treatment group were significant higher than that of the model group (P<0.01). However, the Bax and activated caspase-3 expressions in endometrial gland were similar in both treatment and control groups. Conclusion: Promoting the increases in Bax and activated caspase-3 expressions in the endometrial gland and bringing into balance between apoptosis and proliferation of the glandular cells at the implantation window phase by FTKRQHB may contribute to the effects of promoting the establishment of endometrial receptivity and improving blastocyst implantation dysfunction.
