Background Within the follicular fluid,extracellular vesicles(EVs)guide oocyte growth through their cargo microRNAs(miRNAs).Here,we inv estigated the role of EVs and their cargo miRNAs by linking the miRNAs found in E...Background Within the follicular fluid,extracellular vesicles(EVs)guide oocyte growth through their cargo microRNAs(miRNAs).Here,we inv estigated the role of EVs and their cargo miRNAs by linking the miRNAs found in EVs,derived from the fluid of an individual follicle,to the ability of its oocyte to become a blastocyst(competent)or not(non-competent).Methods Bovine antral follicles were dissected,categorized as small(2-4 mm)or large(5-8 mm)and the corresponding oocytes were subjected to individual maturation,fertilization and embryo culture to the blastocyst stage.Follicular fluid was pooled in 4 groups(4 replicates)based on follicle size and competence of the corresponding oocyte to produce a blastocyst.Follicular fluid-derived EVs were isolated",characterized,and subjected to miRNAsequencing(Illumina Miseq)to assess differential expression(DE)in the 4 groups.Functional validation of the effect of miR-34c on embryo development was performed by supplementation of mimics and inhibitors during in vitro maturation(IVM).Results We identified 16 DE miRNAs linked to oocyte competence when follicular size was not considered.Within the large and small follicles,46 DE miRNAs were driving blastocyst formation in each group.Comparison of EVs from competent small and large follicles revealed 90 DE miRNAs.Cell regulation,cell differentiation,cell cycle,and metabolic process regulation were the most enriched pathways targeted by the DE miRNAs from competent oocytes.We identified bta-miR-34c as the most abundant in follicular fluid containing competent oocytes.Supplementation of miR-34c mimic and inhibitor during IVM did not affect embryo development.However,blastocyst quality,as evidenced by higher cell numbers,was significantly improved following oocyte IVM in the presence of miR-34c mimics,while miR-34c inhibitors resulted in the opposite effect.Conclusion This study demonstrates the regulatory effect of miRNAs from follicular fliived EVs on oocyte competence acquisitividing a further basis for understanding the significance of miRNAs in oocyte maturation and embryonic development Up-regulation of miR-34c in EVs from follicular fluid containing competent oocytes and the positive impact of miR-34c mimics added during IVM on the resulting blastocysts indicate its pivotal role in oocyte competence.展开更多
Granulosa cells（GCs） are somatic cells of ovary, the behaviors of GCs are important for ovarian function. MicroRNAs（miRNAs） are a class of endogenous 18–24 nucleotide（nt） non-coding RNAs, some of which have bee...Granulosa cells（GCs） are somatic cells of ovary, the behaviors of GCs are important for ovarian function. MicroRNAs（miRNAs） are a class of endogenous 18–24 nucleotide（nt） non-coding RNAs, some of which have been shown to be important regulators of GCs function. miR-34c involved in the regulation of various biological processes and was identified to be a pro-apoptotic and anti-proliferative factor in many cell types. However, the roles of miR-34c in GCs function remain unknown. In this study, we used Annexin V-FITC and Ed U assays to demonstrate that miR-34c exerted pro-apoptotic and anti-proliferative effects in porcine GCs. Dual-luciferase reporter assays, quantitative real-time PCR（q RT-PCR） and Western blotting identified Forkhead box O3a（Fox O3a） as a direct target gene of miR-34c. The overexpression of FoxO3a rescued the phenotypic change caused by miR-34c in porcine GCs. In conclusion, miR-34c regulate the function of porcine GCs by targeting FoxO3a.展开更多
目的：探讨miRNA-34c在人脑胶质瘤组织中的表达情况及临床意义。方法应用定量PCR方法对18例WHO分级Ⅱ～Ⅳ级的甲醛固定石蜡包埋的脑胶质瘤标本和外伤或脑出血患者内减压手术获得的5例脑组织标本分别进行miR-34c-3p和miR-34c-5p含量的检...目的：探讨miRNA-34c在人脑胶质瘤组织中的表达情况及临床意义。方法应用定量PCR方法对18例WHO分级Ⅱ～Ⅳ级的甲醛固定石蜡包埋的脑胶质瘤标本和外伤或脑出血患者内减压手术获得的5例脑组织标本分别进行miR-34c-3p和miR-34c-5p含量的检测，比较不同级别胶质瘤和正常脑组织中miR-34c-3p和miR-34c-5p的表达差异，分析miR-34c-3p和miR-34c-5p的表达和胶质瘤恶性程度的相关性。结果与正常脑组织相比，胶质瘤中miR-34c-3p和miR-34c-5p的表达皆显著减少（ P ＜0＊．05），而且miR-34c-3p和miR-34c-5p的表达随着胶质瘤级别或恶性程度的增加分别减少，呈显著的负相关（ miR-34c-3p的spearman相关系数＝－0．856， P ＜0．01；miR-34c-5p的spearman相关系数＝－0．767， P ＜0．01）。结论 MiR-34c在人脑胶质瘤细胞中的表达显著降低，而且表达随着肿瘤分级或恶性程度的增高而减少，与胶质瘤级别的升高呈显著的负相关。 MiR-34c在胶质瘤的进展中有重要的意义，可作为脑胶质瘤临床分级的重要指标。展开更多
Objective This study aimed to examine the role of long non-coding RNA PCED1B antisense RNA 1(PCED1B-AS1)in the development of hepatocellular carcinoma(HCC).Methods A total of 62 pairs of HCC tissues and adjacent non-t...Objective This study aimed to examine the role of long non-coding RNA PCED1B antisense RNA 1(PCED1B-AS1)in the development of hepatocellular carcinoma(HCC).Methods A total of 62 pairs of HCC tissues and adjacent non-tumor tissues were obtained from 62 HCC patients.The interactions of PCED1B-AS1 and microRNA-34a(miR-34a)were detected by dual luciferase activity assay and RNA pull-down assay.The RNA expression levels of PCED1B-AS1,miR-34a and CD44 were detected by RT-qPCR,and the protein expression level of CD44 was determined by Western blotting.The cell proliferation was detected by cell proliferation assay,and the cell invasion and migration by transwell invasion assay.The HCC tumor growth after PCED1B-AS1 was downregulated was determined by in vivo animal study.Results PCED1B-AS1 was highly expressed in HCC tissues,which was associated with poor survival of HCC patients.Furthermore,PCED1B-AS1 interacted with miR-34a in HCC cells,but they did not regulate the expression of each other.Additionally,PCED1B-AS1 increased the expression level of CD44,which was targeted by miR-34a.The cell proliferation and invasion assay revealed that miR-34a inhibited the proliferation and invasion of HCC in vitro,while CD44 exhibited the opposite effects.Furthermore,PCED1B-AS1 suppressed the role of miR-34a.Moreover,the knockdown of PCED1B-AS1 repressed the HCC tumor growth in nude mice in vivo.Conclusion PCED1B-AS1 may play an oncogenic role by regulating the miR-34a/CD44 axis in HCC.展开更多
基金supported by the European Union’s Horizon 2020 research and innovation program under the Marie Sklodowska-Curie grant agreement No 860960by Bijzonder Onderzoeksfonds GOA(Geconcerteerde onderzoeksacties)2018000504(GOA030-18 BOF)supported by the Research Foundation Flanders(FWO)(grant numbers:1228821N and 12A2H24N)。
文摘Background Within the follicular fluid,extracellular vesicles(EVs)guide oocyte growth through their cargo microRNAs(miRNAs).Here,we inv estigated the role of EVs and their cargo miRNAs by linking the miRNAs found in EVs,derived from the fluid of an individual follicle,to the ability of its oocyte to become a blastocyst(competent)or not(non-competent).Methods Bovine antral follicles were dissected,categorized as small(2-4 mm)or large(5-8 mm)and the corresponding oocytes were subjected to individual maturation,fertilization and embryo culture to the blastocyst stage.Follicular fluid was pooled in 4 groups(4 replicates)based on follicle size and competence of the corresponding oocyte to produce a blastocyst.Follicular fluid-derived EVs were isolated",characterized,and subjected to miRNAsequencing(Illumina Miseq)to assess differential expression(DE)in the 4 groups.Functional validation of the effect of miR-34c on embryo development was performed by supplementation of mimics and inhibitors during in vitro maturation(IVM).Results We identified 16 DE miRNAs linked to oocyte competence when follicular size was not considered.Within the large and small follicles,46 DE miRNAs were driving blastocyst formation in each group.Comparison of EVs from competent small and large follicles revealed 90 DE miRNAs.Cell regulation,cell differentiation,cell cycle,and metabolic process regulation were the most enriched pathways targeted by the DE miRNAs from competent oocytes.We identified bta-miR-34c as the most abundant in follicular fluid containing competent oocytes.Supplementation of miR-34c mimic and inhibitor during IVM did not affect embryo development.However,blastocyst quality,as evidenced by higher cell numbers,was significantly improved following oocyte IVM in the presence of miR-34c mimics,while miR-34c inhibitors resulted in the opposite effect.Conclusion This study demonstrates the regulatory effect of miRNAs from follicular fliived EVs on oocyte competence acquisitividing a further basis for understanding the significance of miRNAs in oocyte maturation and embryonic development Up-regulation of miR-34c in EVs from follicular fluid containing competent oocytes and the positive impact of miR-34c mimics added during IVM on the resulting blastocysts indicate its pivotal role in oocyte competence.
基金supported by the National Natural Science Foundation of China (31201771)the earmarked fund for the China Agriculture Research System (CARS-36)
文摘Granulosa cells（GCs） are somatic cells of ovary, the behaviors of GCs are important for ovarian function. MicroRNAs（miRNAs） are a class of endogenous 18–24 nucleotide（nt） non-coding RNAs, some of which have been shown to be important regulators of GCs function. miR-34c involved in the regulation of various biological processes and was identified to be a pro-apoptotic and anti-proliferative factor in many cell types. However, the roles of miR-34c in GCs function remain unknown. In this study, we used Annexin V-FITC and Ed U assays to demonstrate that miR-34c exerted pro-apoptotic and anti-proliferative effects in porcine GCs. Dual-luciferase reporter assays, quantitative real-time PCR（q RT-PCR） and Western blotting identified Forkhead box O3a（Fox O3a） as a direct target gene of miR-34c. The overexpression of FoxO3a rescued the phenotypic change caused by miR-34c in porcine GCs. In conclusion, miR-34c regulate the function of porcine GCs by targeting FoxO3a.
文摘目的：探讨miRNA-34c在人脑胶质瘤组织中的表达情况及临床意义。方法应用定量PCR方法对18例WHO分级Ⅱ～Ⅳ级的甲醛固定石蜡包埋的脑胶质瘤标本和外伤或脑出血患者内减压手术获得的5例脑组织标本分别进行miR-34c-3p和miR-34c-5p含量的检测，比较不同级别胶质瘤和正常脑组织中miR-34c-3p和miR-34c-5p的表达差异，分析miR-34c-3p和miR-34c-5p的表达和胶质瘤恶性程度的相关性。结果与正常脑组织相比，胶质瘤中miR-34c-3p和miR-34c-5p的表达皆显著减少（ P ＜0＊．05），而且miR-34c-3p和miR-34c-5p的表达随着胶质瘤级别或恶性程度的增加分别减少，呈显著的负相关（ miR-34c-3p的spearman相关系数＝－0．856， P ＜0．01；miR-34c-5p的spearman相关系数＝－0．767， P ＜0．01）。结论 MiR-34c在人脑胶质瘤细胞中的表达显著降低，而且表达随着肿瘤分级或恶性程度的增高而减少，与胶质瘤级别的升高呈显著的负相关。 MiR-34c在胶质瘤的进展中有重要的意义，可作为脑胶质瘤临床分级的重要指标。
基金supported by the Medical Science and Technology Research Foundation of Guangdong Province(No.A2020559).
文摘Objective This study aimed to examine the role of long non-coding RNA PCED1B antisense RNA 1(PCED1B-AS1)in the development of hepatocellular carcinoma(HCC).Methods A total of 62 pairs of HCC tissues and adjacent non-tumor tissues were obtained from 62 HCC patients.The interactions of PCED1B-AS1 and microRNA-34a(miR-34a)were detected by dual luciferase activity assay and RNA pull-down assay.The RNA expression levels of PCED1B-AS1,miR-34a and CD44 were detected by RT-qPCR,and the protein expression level of CD44 was determined by Western blotting.The cell proliferation was detected by cell proliferation assay,and the cell invasion and migration by transwell invasion assay.The HCC tumor growth after PCED1B-AS1 was downregulated was determined by in vivo animal study.Results PCED1B-AS1 was highly expressed in HCC tissues,which was associated with poor survival of HCC patients.Furthermore,PCED1B-AS1 interacted with miR-34a in HCC cells,but they did not regulate the expression of each other.Additionally,PCED1B-AS1 increased the expression level of CD44,which was targeted by miR-34a.The cell proliferation and invasion assay revealed that miR-34a inhibited the proliferation and invasion of HCC in vitro,while CD44 exhibited the opposite effects.Furthermore,PCED1B-AS1 suppressed the role of miR-34a.Moreover,the knockdown of PCED1B-AS1 repressed the HCC tumor growth in nude mice in vivo.Conclusion PCED1B-AS1 may play an oncogenic role by regulating the miR-34a/CD44 axis in HCC.
