Background:MiRNAs act as pivotal post-transcriptional gene mediators in the regulation of diverse biological processes,including proliferation,development and apoptosis.Our previous study has showed that miR-101-3p is...Background:MiRNAs act as pivotal post-transcriptional gene mediators in the regulation of diverse biological processes,including proliferation,development and apoptosis.Our previous study has showed that miR-101-3p is differentially expressed in dairy goat ovaries compared single with multiple litters.The objective of this research was to explore the potential function and molecular mechanism of miR-101-3p via its target STC1 in goat ovarian growth and development.Results:cDNA libraries were constructed using goat granulosa cells transfected with miR-101-3p mimics and negative control by RNA-sequencing.In total,142 differentially expressed unigenes(DEGs)were detected between two libraries,including 78 down-regulated and 64 up-regulated genes.GO and KEGG enrichment analysis showed the potential impacts of DEGs on ovarian development.STC1 was singled out from DEGs for further research owing to it regulates reproductive-related processes.In vitro,bioinformatics analysis and 3′-UTR assays confirmed that STC1 was a target of miR-101-3p.ELISA was performed to detect the estrogen(E2)and progesterone(P4)levels.CCK8,EdU and flow cytometry assays were performed to detect the proliferation and apoptosis of granulosa cells.Results showed that miR-101-3p regulated STAR,CYP19A1,CYP11A1 and 3β-HSD steroid hormone synthesis-associated genes by STC1 depletion,thus promoted E2 and P4 secretions.MiR-101-3p also affected the key protein PI3K,PTEN,AKT and mTOR in PI3K-AKT pathway by STC1,thereby suppressing proliferation and promoting apoptosis of granulosa cells.In vivo,the distribution and expression levels of miR-101-3p in mouse ovaries were determined through fluorescence in situ hybridisation(FISH).Immunohistochemistry results showed that STC1 expression was suppressed in mouse ovaries in miR-101-3p-agonist and siRNA-STC1 groups.Small and stunted ovarian fragments,decreased numbers of follicles at diverse stages were observed using Hematoxylin-eosin(HE)staining,thereby showing unusual ovarian development after miR-101-3p overexpression or STC1 depletion.Inhibition of miR-101-3p manifested opposite results.Conclusions:Taken together,our results demonstrated a regulatory mechanism of miR-101-3p via STC1 in goat granulosa cells,and offered the first in vivo example of miR-101-3p and STC1 functions required for ovarian development.展开更多
Objective: To investigate the effects of CAL-101, particularly when combined with bortezomib(BTZ) on mantle cell lymphoma(MCL) cells, and to explore its relative mechanisms.Methods: MTT assay was applied to detect the...Objective: To investigate the effects of CAL-101, particularly when combined with bortezomib(BTZ) on mantle cell lymphoma(MCL) cells, and to explore its relative mechanisms.Methods: MTT assay was applied to detect the inhibitory effects of different concentrations of CAL-101. MCL cells were divided into four groups: control group, CAL-101 group, BTZ group, and CAL-101/BTZ group. The expression of PI3K-p110σ, AKT, ERK, p-AKT and p-ERK were detected by Western blot. The apoptosis rates of CAL-101 group, BTZ group, and combination group were detected by flow cytometry. The location changes of nuclear factor kappa-B(NF-κB) of 4 groups was investigated by NF-κB Kit exploring. Western blot was applied to detect the levels of caspase-3 and the phosphorylation of AKT in different groups. Results: CAL-101 dose- and time-dependently induced reduction in MCL cell viability. CAL-101 combined with BTZ enhanced the reduction in cell viability and apoptosis. Western blot analysis showed that CAL-101 significantly blocked the PI3K/AKT and ERK signaling pathway in MCL cells. The combination therapy contributed to the inactivation of NF-κB and AKT in MCL cell lines. However, cleaved caspase-3 was up-regulated after combined treatment. Conclusion: Our study showed that PI3K/p110σ is a novel therapeutic target in MCL, and the underlying mechanism could be the blocking of the PI3K/AKT and ERK signaling pathways. These findings provided a basis for clinical evaluation of CAL-101 and a rationale for its application in combination therapy, particularly with BTZ.展开更多
目的探讨miR-101在非小细胞肺癌发生、发展中的作用以及新的分子调控机制。方法将miR-101转染到非小细胞肺癌A549细胞株,比较测量人类非小细胞肺癌组织及癌旁组织的miR-101表达水平。采用生物信息学技术预测miR-101在c-Fos的3′UTR潜在...目的探讨miR-101在非小细胞肺癌发生、发展中的作用以及新的分子调控机制。方法将miR-101转染到非小细胞肺癌A549细胞株,比较测量人类非小细胞肺癌组织及癌旁组织的miR-101表达水平。采用生物信息学技术预测miR-101在c-Fos的3′UTR潜在结合靶点,然后采用Western blot法和基因敲除方法检测c-Fos的表达和功能。结果分析miR-101在非小细胞肺癌组织及癌旁组织表达,miR-101在非小细胞肺癌组织中低表达(0.00010±0.00002 vs 0.00021±0.00003,P=0.036),过表达miR-101抑制非小细胞肺癌A549细胞增殖并诱导细胞凋亡。并且在非小细胞肺癌细胞中敲除c-Fos和过表达miR-101有相似的效果。结论 miR-101通过c-Fos靶点调节非小细胞肺癌细胞的生长,因此调控miR-101和c-Fos可以应用于非小细胞肺癌潜在的分子治疗。展开更多
基金supported by the National Natural Science Foundation of China(31601925)Shaanxi Science and Technology Innovation Project Plan(2020ZDLNY02–01,2020ZDLNY02–02,2018ZDCXL-NY-01-04,2018ZDCXL-NY-01-02 and 2017ZDXM-NY-081)Natural Science Foundation of Shaanxi Province(2020JQ-868)。
文摘Background:MiRNAs act as pivotal post-transcriptional gene mediators in the regulation of diverse biological processes,including proliferation,development and apoptosis.Our previous study has showed that miR-101-3p is differentially expressed in dairy goat ovaries compared single with multiple litters.The objective of this research was to explore the potential function and molecular mechanism of miR-101-3p via its target STC1 in goat ovarian growth and development.Results:cDNA libraries were constructed using goat granulosa cells transfected with miR-101-3p mimics and negative control by RNA-sequencing.In total,142 differentially expressed unigenes(DEGs)were detected between two libraries,including 78 down-regulated and 64 up-regulated genes.GO and KEGG enrichment analysis showed the potential impacts of DEGs on ovarian development.STC1 was singled out from DEGs for further research owing to it regulates reproductive-related processes.In vitro,bioinformatics analysis and 3′-UTR assays confirmed that STC1 was a target of miR-101-3p.ELISA was performed to detect the estrogen(E2)and progesterone(P4)levels.CCK8,EdU and flow cytometry assays were performed to detect the proliferation and apoptosis of granulosa cells.Results showed that miR-101-3p regulated STAR,CYP19A1,CYP11A1 and 3β-HSD steroid hormone synthesis-associated genes by STC1 depletion,thus promoted E2 and P4 secretions.MiR-101-3p also affected the key protein PI3K,PTEN,AKT and mTOR in PI3K-AKT pathway by STC1,thereby suppressing proliferation and promoting apoptosis of granulosa cells.In vivo,the distribution and expression levels of miR-101-3p in mouse ovaries were determined through fluorescence in situ hybridisation(FISH).Immunohistochemistry results showed that STC1 expression was suppressed in mouse ovaries in miR-101-3p-agonist and siRNA-STC1 groups.Small and stunted ovarian fragments,decreased numbers of follicles at diverse stages were observed using Hematoxylin-eosin(HE)staining,thereby showing unusual ovarian development after miR-101-3p overexpression or STC1 depletion.Inhibition of miR-101-3p manifested opposite results.Conclusions:Taken together,our results demonstrated a regulatory mechanism of miR-101-3p via STC1 in goat granulosa cells,and offered the first in vivo example of miR-101-3p and STC1 functions required for ovarian development.
基金supported by the National Natural Science Foundation of China (Grant No. 30672208, 81270603, and 31301161)Tianjin Natural Science Foundation of China (Grant No. 13JCYBJC22800)
文摘Objective: To investigate the effects of CAL-101, particularly when combined with bortezomib(BTZ) on mantle cell lymphoma(MCL) cells, and to explore its relative mechanisms.Methods: MTT assay was applied to detect the inhibitory effects of different concentrations of CAL-101. MCL cells were divided into four groups: control group, CAL-101 group, BTZ group, and CAL-101/BTZ group. The expression of PI3K-p110σ, AKT, ERK, p-AKT and p-ERK were detected by Western blot. The apoptosis rates of CAL-101 group, BTZ group, and combination group were detected by flow cytometry. The location changes of nuclear factor kappa-B(NF-κB) of 4 groups was investigated by NF-κB Kit exploring. Western blot was applied to detect the levels of caspase-3 and the phosphorylation of AKT in different groups. Results: CAL-101 dose- and time-dependently induced reduction in MCL cell viability. CAL-101 combined with BTZ enhanced the reduction in cell viability and apoptosis. Western blot analysis showed that CAL-101 significantly blocked the PI3K/AKT and ERK signaling pathway in MCL cells. The combination therapy contributed to the inactivation of NF-κB and AKT in MCL cell lines. However, cleaved caspase-3 was up-regulated after combined treatment. Conclusion: Our study showed that PI3K/p110σ is a novel therapeutic target in MCL, and the underlying mechanism could be the blocking of the PI3K/AKT and ERK signaling pathways. These findings provided a basis for clinical evaluation of CAL-101 and a rationale for its application in combination therapy, particularly with BTZ.
文摘目的探讨miR-101在非小细胞肺癌发生、发展中的作用以及新的分子调控机制。方法将miR-101转染到非小细胞肺癌A549细胞株,比较测量人类非小细胞肺癌组织及癌旁组织的miR-101表达水平。采用生物信息学技术预测miR-101在c-Fos的3′UTR潜在结合靶点,然后采用Western blot法和基因敲除方法检测c-Fos的表达和功能。结果分析miR-101在非小细胞肺癌组织及癌旁组织表达,miR-101在非小细胞肺癌组织中低表达(0.00010±0.00002 vs 0.00021±0.00003,P=0.036),过表达miR-101抑制非小细胞肺癌A549细胞增殖并诱导细胞凋亡。并且在非小细胞肺癌细胞中敲除c-Fos和过表达miR-101有相似的效果。结论 miR-101通过c-Fos靶点调节非小细胞肺癌细胞的生长,因此调控miR-101和c-Fos可以应用于非小细胞肺癌潜在的分子治疗。
