Ovarian follicle development is associated with the physiological functions of granulosa cells(GCs),including proliferation and apoptosis.The level of miR-24-3p in ovarian tissue of high-yielding Yorkshire×Landra...Ovarian follicle development is associated with the physiological functions of granulosa cells(GCs),including proliferation and apoptosis.The level of miR-24-3p in ovarian tissue of high-yielding Yorkshire×Landrace sows was significantly higher than that of low-yielding sows.However,the functions of miR-24-3p on GCs are unclear.In this study,using flow cytometry,5-ethynyl-2′-de-oxyuridine(EdU)staining,and cell count,we showed that miR-24-3p promoted the proliferation of GCs increasing the proportion of cells in the S phase and upregulating the expression of cell cycle genes,moreover,miR-24-3p inhibited GC apoptosis.Mechanistically,on-line prediction,bioinformatics analysis,a luciferase reporter assay,RT-qPCR,and Western blot results showed that the target gene of miR-24-3p in proliferation and apoptosis is cyclin-dependent kinase inhibitor 1B(P27/CDKN1B).Furthermore,the effect of miR-24-3p on GC proliferation and apoptosis was attenuated by P27 overexpression.These findings suggest that miR-24-3p regulates the physiological functions of GCs.展开更多
Background Atresia and degeneration,a follicular developmental fate that reduces female fertility and is triggered by granulosa cell(GC)apoptosis,have been induced by dozens of miRNAs.Here,we report a miRNA,miR-423,th...Background Atresia and degeneration,a follicular developmental fate that reduces female fertility and is triggered by granulosa cell(GC)apoptosis,have been induced by dozens of miRNAs.Here,we report a miRNA,miR-423,that inhibits the initiation of follicular atresia(FA),and early apoptosis of GCs.Results We showed that miR-423 was down-regulated during sow FA,and its levels in follicles were negatively correlated with the GC density and the P4/E2 ratio in the follicular fluid in vivo.The in vitro gain-of-function experiments revealed that miR-423 suppresses cell apoptosis,especially early apoptosis in GCs.Mechanically speaking,the miR-423 targets and interacts with the 3’-UTR of the porcine SMAD7 gene,which encodes an apoptosis-inducing factor in GCs,and represses its expression and pro-apoptotic function.Interestingly,FA and the GC apoptosis-related lncRNA NORHA was demonstrated as a ceRNA of miR-423.Additionally,we showed that a single base deletion/insertion in the miR-423 promoter is significantly associated with the number of stillbirths(NSB)trait of sows.Conclusion These results demonstrate that miR-423 is a small molecule for inhibiting FA initiation and GC early apoptosis,suggesting that treating with miR-423 may be a novel approach for inhibiting FA initiation and improving female fertility.展开更多
BACKGROUND Gastric carcinoma(GC)is the third most frequent cause of cancer-related death,highlighting the pressing need for novel clinical treatment options.In this regard,microRNAs(miRNAs)have emerged as a promising ...BACKGROUND Gastric carcinoma(GC)is the third most frequent cause of cancer-related death,highlighting the pressing need for novel clinical treatment options.In this regard,microRNAs(miRNAs)have emerged as a promising therapeutic strategy.Studies have shown that miRNAs can regulate related signaling pathways,acting as tumor suppressors or tumor promoters.AIM To explore the effect of miR-204-3p on GC cells.METHODS We measured the expression levels of miR-204-3p in GC cells using quantitative real-time polymerase chain reaction,followed by the delivery of miR-204-3p overexpression and miR-204-3p knockdown vectors into GC cells.CCK-8 was used to detect the effect of miR-204-3p on the proliferation of GC cells,and the colony formation ability of GC cells was detected by the clonal formation assay.The effects of miR-204-3p on GC cell cycle and apoptosis were detected by flow cytometry.The BABL/c nude mouse subcutaneous tumor model using MKN-45 cells was constructed to verify the effect of miR-204-3p on the tumorigenicity of GC cells.Furthermore,the study investigated the effects of miR-204-3p on various proteins related to the MAPK signaling pathway,necroptosis signaling pathway and apoptosis signaling pathway on GC cells using Western blot techniques.RESULTS Firstly,we found that the expression of miR-204-3p in GC was low.When treated with the lentivirus overexpression vector,miR-204-3p expression significantly increased,but the lentivirus knockout vector had no significant effect on miR-204-3p.In vitro experiments confirmed that miR-204-3p overexpression inhibited GC cell viability,promoted cell apoptosis,blocked the cell cycle,and inhibited colony formation ability.In vivo animal experiments confirmed that miR-204-3p overexpression inhibited subcutaneous tumorigenesis ability in BABL/c nude mice.Simultaneously,our results verified that miR-204-3p overexpression can inhibit GC cell proliferation by inhibiting protein expression levels of KRAS and p-ERK1/2 in the MAPK pathway,as well as inhibiting protein expression levels of p-RIP1 and p-MLK1 in the necroptosis pathway to promote the BCL-2/BAX/Caspase-3 apoptosis pathway.CONCLUSION MiR-204-3p overexpression inhibited GC cell proliferation by inhibiting the MAPK pathway and necroptosis pathway to promote apoptosis of GC cells.Thus,miR-204-3p may represent a new potential therapeutic target for GC.展开更多
BACKGROUND Diabetes is a chronic metabolic disease,and a variety of miRNA are involved in the occurrence and development of diabetes.In clinical studies,miR-124 is highly expressed in the serum of patients with diabet...BACKGROUND Diabetes is a chronic metabolic disease,and a variety of miRNA are involved in the occurrence and development of diabetes.In clinical studies,miR-124 is highly expressed in the serum of patients with diabetes and in pancreatic isletβ-cells.However,few reports exist concerning the role and mechanism of action of miR-124 in diabetes.AIM To investigate the expression of miR-124 in diabetic mice and the potential mechanism of action in isletβ-cells.METHODS The expression levels of miR-124 and enhancer of zeste homolog 2(EZH2)in pancreatic tissues of diabetic mice were detected.The targeted relationship between miR-124 and EZH2 was predicted by Targetscan software and verified by a double luciferase reporter assay.Mouse isletβ-cells Min6 were grown in a high glucose(HG)medium to mimic a diabetes model.The insulin secretion,proliferation,cell cycle and apoptosis of HG-induced Min6 cells were detected after interference of miR-124a and/or EZH2.RESULTS The expression of miR-124 was upregulated and EZH2 was downregulated in the pancreatic tissue of diabetic mice compared with control mice,and the expression of miR-124 was negatively correlated with that of EZH2.miR-124 was highly expressed in HG-induced Min6 cells.Inhibition of miR-124 promoted insulin secretion and cell proliferation,induced the transition from the G0/G1 phase to the S phase of the cell cycle,and inhibited cell apoptosis in HG-induced Min6 cells.EZH2 was one of the targets of miR-124.Co-transfection of miR-124 inhibitor and siRNA-EZH2 could reverse the effects of the miR-124 inhibitor in HG-induced Min6 cells.CONCLUSION miR-124 is highly expressed in diabetic mice and HG-induced Min6 cells and regulates insulin secretion,proliferation and apoptosis of isletβ-cells by targeting EZH2.展开更多
Background: To investigate the therapeutic activity of the miR-221/222 inhibitor against OSCC in vitro and in vivo. Materials and Methods: HSC3 and HSC6 were treated with miR-221/222 inhibitor and the empty vector res...Background: To investigate the therapeutic activity of the miR-221/222 inhibitor against OSCC in vitro and in vivo. Materials and Methods: HSC3 and HSC6 were treated with miR-221/222 inhibitor and the empty vector respectively. After the recombinants were transfected into HSC3 and HSC6 with Lipofectamine<sup>TM</sup> MAX, the expression of miR-221/222 and PUMA was analyzed by RT-PCR. The proliferation and migration of HSC3 and HSC6 were detected by CCK-8 assay and Wound-healing assay. Cell cycle and apoptosis were detected by flow cytometry. The effect of the miR-221/222 inhibitor was also assessed in OSCC xenografts in BALB/c-nu mice. Results: Transfection of the miR-221/222 inhibitor increased cell apoptosis and upregulated PUMA expression in OSCC cell lines HSC3 and HSC6 with the significantly reduced expression of miR-221 and miR-222. Furthermore, the miR-221/222 inhibitor suppressed cell growth and invasion and blocked the cell cycle at the G1 phase. Obvious anti-tumor activity was achieved in BALB/c-nu mice by treatment with the miR-221/222 inhibitor, together with the upregulation of PUMA protein in tumors retrieved from the mice. Conclusions: There was a significant inhibitory effect of the miR-221/222 inhibitor on the growth of OSCC both in vitro and in vivo, and there might be a regulatory loop between miR-221/222 and PUMA. These findings demonstrated that downregulation of miR-221/222 could induce cell apoptosis, and it might be considered as a candidate target for gene therapy of OSCC.展开更多
Objective:Investigating the effects of miR-873 on apoptosis and autophagy in bronchial epithelial cells,as well as its regulatory role on Beclin1.Methods:Following transfection of miR-873 mimic into 16HBE cells for 48...Objective:Investigating the effects of miR-873 on apoptosis and autophagy in bronchial epithelial cells,as well as its regulatory role on Beclin1.Methods:Following transfection of miR-873 mimic into 16HBE cells for 48 hours,the mRNA level of miR-873 was quantified by qRT-PCR,and cell viability was evaluated by CCK-8 assay.The levels of IL-2,IL-6,IL-10,and TNF-αin the cell supernatant were determined using ELISA assay,while cell apoptosis was detected by TUNEL staining.LC-3 protein expression was examined by immunofluorescence,and mRNA and protein expression levels of Beclin1 were analyzed by qRT-PCR and Western blot,respectively.Moreover,dual-luciferase reporter gene technology was employed to investigate the binding site between miR-873 and Beclin1.Results:Transfection of miR-873 mimic into 16HBE cells significantly upregulated the mRNA level of miR-873,which led to the inhibition of cell proliferation and the promotion of secretion of pro-inflammatory cytokines IL-2,IL-6,and TNF-α,while suppressing the secretion of anti-inflammatory cytokine IL-10.Moreover,miR-873 induced cell apoptosis and inhibited the expression of LC-3.Dual-luciferase reporter gene assay further confirmed the presence of binding sites between miR-873 and Beclin1 gene.Besides,miR-873 could target and suppress the mRNA and protein expression levels of Beclin1.Conclusion:miR-873 might modulate cell autophagy by targeting the Beclin1 gene,which can potentially promote inflammation and apoptosis in bronchial epithelial cells.展开更多
Objective:To investigate the role of miR-129-5p in regulating HMGB1 expression in pancreatic cancer cell apoptosis.Methods:The untreated pancreatic cancer SW1990 cells were used as the control group.Mimics-NC(empty ve...Objective:To investigate the role of miR-129-5p in regulating HMGB1 expression in pancreatic cancer cell apoptosis.Methods:The untreated pancreatic cancer SW1990 cells were used as the control group.Mimics-NC(empty vector),miR-129-5p mimics,inhibitor-NC(empty vector)and miR-129-5p inhibitor were transfected into SW1990 cells by liposome transfection method as the mimics-NC group,miR-129-5p overexpression group(miR-129-5p mimics group),inhibitor-NC group and miR-129-5p low expression group(miR-129-5p inhibitor group).The binding site of miR-129-5p and HMGB1 was predicted by online target gene prediction website Target genes,and the targeting relationship between miR-129-5p and HMGB1 was verified by dual luciferase gene report experiment.The expression of miR-129-5p in each group was detected by qRT-PCR,and the expression of HMGB1 protein and apoptosis-related proteins Caspase 3 and Bcl-2 by Western blot.Hoechst staining was used to observe the changes of apoptosis.Results:Compared with the mimics-NC group and control group,miR-129-5p mimics transfection significantly up-regulated miR-129-5p level(P<0.01),inhibited HMGB1(P<0.01)and Bcl-2(P<0.05)protein expression,pro-moted Caspase 3 protein expression(P<0.05),and promoted apoptosis;compared with the inhibitor-NC group and control group,miR-129-5p inhibitor transfection significantly down-regulated miR-129-5p level(P<0.05),promoted HMGB1 and Bcl-2 protein expression(all P<0.05),inhibited Caspase 3protein expression(P<0.01),and inhibited apoptosis.The results of dual luciferase reporter gene assay showed that miR-129-5p could inhibit the fluorescence activity of wildtype HMGB1 cells and target the expression of HMGB1.Conclusion:miR-129-5p promotes the apoptosis of pancreatic cancer SW1990 cells by targeting inhibition of HMGB1 expression.展开更多
Objective:To investigate the effects of co-transfection of miR-520c-3p and miR-132 on proliferation and apoptosis of hepatocellular carcinoma Huh7.Methods:Hepatocellular carcinoma Huh7 was cultured in vitro and lipido...Objective:To investigate the effects of co-transfection of miR-520c-3p and miR-132 on proliferation and apoptosis of hepatocellular carcinoma Huh7.Methods:Hepatocellular carcinoma Huh7 was cultured in vitro and lipidosome was used to transfect miR-520c-3p and miR-132 respectively or together.The effects of transfection of miR-520c-3p and miR-132 on proliferation and apoptosis of Huh7 were detected by CCK8 and Annexin V staining and flow cytometry,and the expression level of the targeted gene of over-expressed miR-520c-3p and miR-132 was determined by Western blot and realtime PCR.Results:Compared with the control group,the proliferation ability of Huh7 of the single transfected and co-transfected miR-520c-3p and miR-132 decreased significantly,and the apoptosis ratio increased distinctly(P<0.05).Besides,the affect of the co-transfection group was better than that of the single transfection group.The protein levels of GPC3(Glypican-3) and YAP(Yes-associated protein),the target genes transfected only by miR-520c-3p and miR-132,respectively,reduced obviously(P<0.05),which was similar with the co-infected cells,but cells transfected by miR-132 only showed a decrease of YAP.Conclusions:The co-transfection of miR-520c-3p and miR-132 can target-regulate the expression of GPC3 and YAP,enhance the exhibition effect on proliferation of hepatocellular carcinoma Huh7 and induce cell apoptosis synergistically.展开更多
Objective：Previous studies have identified that kazrin is a constituent of desmosome and influences intercellular adhesion,growing development and morphology.We previously cloned another new isoform,kazrin F and foun...Objective：Previous studies have identified that kazrin is a constituent of desmosome and influences intercellular adhesion,growing development and morphology.We previously cloned another new isoform,kazrin F and found that it has anti-apoptotic effects on human glioma cell line.To further explore whether kazrin F is involved in tumorigenesis,we investigated its expression and role in cervical cancer（CC） cells.Methods：The role of kazrin F and miR-186 in CC was determined by 3-（4,5-dimethylthiazol-2-yl）-2,5-diphenyltetrazolium bromide（MTT） assay,colony formation,transwell,and apoptosis assays.Using enhanced green fluorescent protein（EGFP） reporter assays,reverse transcription-quantitative polymerase chain reaction（RT-qPCR） and western blot analysis,we identified kazrin F post-transcriptional regulation by miR-186.Results：We demonstrate that kazrin F is highly expressed in CC tissues compared with the adjacent noncancerous tissues and promotes cell proliferation,colony formation,migration and invasion in HeLa and C33 A cells by suppressing apoptosis and facilitating epithelial-to-mesenchymal transition（EMT）.Furthermore,miR-186 was confirmed as a regulator of kazrin F dysregulation.An EGFP reporter assay proved that miR-186 directly targets the 3＇-untranslated region（3＇UTR） of kazrin F and downregulates its expression,and miR-186 expression showed an inverse correlation with kazrin F levels in CC tissues.In addition,overexpression of miR-186 suppressed the malignant behaviors of CC cells.The ectopic expression of kazrin F rescued the inhibitory effects of miR-186.Conclusions：Our findings indicate that the upregulation of kazrin F due to downregulated miR-186 levels contributes to malignancy,and highlight the significance of kazrin F in CC tumorigenesis.展开更多
基金supported by the National Natural Science Foundation of China(32272849)the National Key R&D Program of China(2021YFF1000602)the earmarked fund for CARS-35-PIG。
文摘Ovarian follicle development is associated with the physiological functions of granulosa cells(GCs),including proliferation and apoptosis.The level of miR-24-3p in ovarian tissue of high-yielding Yorkshire×Landrace sows was significantly higher than that of low-yielding sows.However,the functions of miR-24-3p on GCs are unclear.In this study,using flow cytometry,5-ethynyl-2′-de-oxyuridine(EdU)staining,and cell count,we showed that miR-24-3p promoted the proliferation of GCs increasing the proportion of cells in the S phase and upregulating the expression of cell cycle genes,moreover,miR-24-3p inhibited GC apoptosis.Mechanistically,on-line prediction,bioinformatics analysis,a luciferase reporter assay,RT-qPCR,and Western blot results showed that the target gene of miR-24-3p in proliferation and apoptosis is cyclin-dependent kinase inhibitor 1B(P27/CDKN1B).Furthermore,the effect of miR-24-3p on GC proliferation and apoptosis was attenuated by P27 overexpression.These findings suggest that miR-24-3p regulates the physiological functions of GCs.
基金supported by the National Key R&D Program of China(2022YFD1600903)the National Natural Science Foundation of China(32072693)the College Students’Innovative Entrepreneurial Training Plan Program(202110307028).
文摘Background Atresia and degeneration,a follicular developmental fate that reduces female fertility and is triggered by granulosa cell(GC)apoptosis,have been induced by dozens of miRNAs.Here,we report a miRNA,miR-423,that inhibits the initiation of follicular atresia(FA),and early apoptosis of GCs.Results We showed that miR-423 was down-regulated during sow FA,and its levels in follicles were negatively correlated with the GC density and the P4/E2 ratio in the follicular fluid in vivo.The in vitro gain-of-function experiments revealed that miR-423 suppresses cell apoptosis,especially early apoptosis in GCs.Mechanically speaking,the miR-423 targets and interacts with the 3’-UTR of the porcine SMAD7 gene,which encodes an apoptosis-inducing factor in GCs,and represses its expression and pro-apoptotic function.Interestingly,FA and the GC apoptosis-related lncRNA NORHA was demonstrated as a ceRNA of miR-423.Additionally,we showed that a single base deletion/insertion in the miR-423 promoter is significantly associated with the number of stillbirths(NSB)trait of sows.Conclusion These results demonstrate that miR-423 is a small molecule for inhibiting FA initiation and GC early apoptosis,suggesting that treating with miR-423 may be a novel approach for inhibiting FA initiation and improving female fertility.
文摘BACKGROUND Gastric carcinoma(GC)is the third most frequent cause of cancer-related death,highlighting the pressing need for novel clinical treatment options.In this regard,microRNAs(miRNAs)have emerged as a promising therapeutic strategy.Studies have shown that miRNAs can regulate related signaling pathways,acting as tumor suppressors or tumor promoters.AIM To explore the effect of miR-204-3p on GC cells.METHODS We measured the expression levels of miR-204-3p in GC cells using quantitative real-time polymerase chain reaction,followed by the delivery of miR-204-3p overexpression and miR-204-3p knockdown vectors into GC cells.CCK-8 was used to detect the effect of miR-204-3p on the proliferation of GC cells,and the colony formation ability of GC cells was detected by the clonal formation assay.The effects of miR-204-3p on GC cell cycle and apoptosis were detected by flow cytometry.The BABL/c nude mouse subcutaneous tumor model using MKN-45 cells was constructed to verify the effect of miR-204-3p on the tumorigenicity of GC cells.Furthermore,the study investigated the effects of miR-204-3p on various proteins related to the MAPK signaling pathway,necroptosis signaling pathway and apoptosis signaling pathway on GC cells using Western blot techniques.RESULTS Firstly,we found that the expression of miR-204-3p in GC was low.When treated with the lentivirus overexpression vector,miR-204-3p expression significantly increased,but the lentivirus knockout vector had no significant effect on miR-204-3p.In vitro experiments confirmed that miR-204-3p overexpression inhibited GC cell viability,promoted cell apoptosis,blocked the cell cycle,and inhibited colony formation ability.In vivo animal experiments confirmed that miR-204-3p overexpression inhibited subcutaneous tumorigenesis ability in BABL/c nude mice.Simultaneously,our results verified that miR-204-3p overexpression can inhibit GC cell proliferation by inhibiting protein expression levels of KRAS and p-ERK1/2 in the MAPK pathway,as well as inhibiting protein expression levels of p-RIP1 and p-MLK1 in the necroptosis pathway to promote the BCL-2/BAX/Caspase-3 apoptosis pathway.CONCLUSION MiR-204-3p overexpression inhibited GC cell proliferation by inhibiting the MAPK pathway and necroptosis pathway to promote apoptosis of GC cells.Thus,miR-204-3p may represent a new potential therapeutic target for GC.
基金Supported by The Medical Science and Technology Key Project of Henan Province,No.2018020733.
文摘BACKGROUND Diabetes is a chronic metabolic disease,and a variety of miRNA are involved in the occurrence and development of diabetes.In clinical studies,miR-124 is highly expressed in the serum of patients with diabetes and in pancreatic isletβ-cells.However,few reports exist concerning the role and mechanism of action of miR-124 in diabetes.AIM To investigate the expression of miR-124 in diabetic mice and the potential mechanism of action in isletβ-cells.METHODS The expression levels of miR-124 and enhancer of zeste homolog 2(EZH2)in pancreatic tissues of diabetic mice were detected.The targeted relationship between miR-124 and EZH2 was predicted by Targetscan software and verified by a double luciferase reporter assay.Mouse isletβ-cells Min6 were grown in a high glucose(HG)medium to mimic a diabetes model.The insulin secretion,proliferation,cell cycle and apoptosis of HG-induced Min6 cells were detected after interference of miR-124a and/or EZH2.RESULTS The expression of miR-124 was upregulated and EZH2 was downregulated in the pancreatic tissue of diabetic mice compared with control mice,and the expression of miR-124 was negatively correlated with that of EZH2.miR-124 was highly expressed in HG-induced Min6 cells.Inhibition of miR-124 promoted insulin secretion and cell proliferation,induced the transition from the G0/G1 phase to the S phase of the cell cycle,and inhibited cell apoptosis in HG-induced Min6 cells.EZH2 was one of the targets of miR-124.Co-transfection of miR-124 inhibitor and siRNA-EZH2 could reverse the effects of the miR-124 inhibitor in HG-induced Min6 cells.CONCLUSION miR-124 is highly expressed in diabetic mice and HG-induced Min6 cells and regulates insulin secretion,proliferation and apoptosis of isletβ-cells by targeting EZH2.
文摘Background: To investigate the therapeutic activity of the miR-221/222 inhibitor against OSCC in vitro and in vivo. Materials and Methods: HSC3 and HSC6 were treated with miR-221/222 inhibitor and the empty vector respectively. After the recombinants were transfected into HSC3 and HSC6 with Lipofectamine<sup>TM</sup> MAX, the expression of miR-221/222 and PUMA was analyzed by RT-PCR. The proliferation and migration of HSC3 and HSC6 were detected by CCK-8 assay and Wound-healing assay. Cell cycle and apoptosis were detected by flow cytometry. The effect of the miR-221/222 inhibitor was also assessed in OSCC xenografts in BALB/c-nu mice. Results: Transfection of the miR-221/222 inhibitor increased cell apoptosis and upregulated PUMA expression in OSCC cell lines HSC3 and HSC6 with the significantly reduced expression of miR-221 and miR-222. Furthermore, the miR-221/222 inhibitor suppressed cell growth and invasion and blocked the cell cycle at the G1 phase. Obvious anti-tumor activity was achieved in BALB/c-nu mice by treatment with the miR-221/222 inhibitor, together with the upregulation of PUMA protein in tumors retrieved from the mice. Conclusions: There was a significant inhibitory effect of the miR-221/222 inhibitor on the growth of OSCC both in vitro and in vivo, and there might be a regulatory loop between miR-221/222 and PUMA. These findings demonstrated that downregulation of miR-221/222 could induce cell apoptosis, and it might be considered as a candidate target for gene therapy of OSCC.
基金National Natural Science Foundation of China(No.82160011)。
文摘Objective:Investigating the effects of miR-873 on apoptosis and autophagy in bronchial epithelial cells,as well as its regulatory role on Beclin1.Methods:Following transfection of miR-873 mimic into 16HBE cells for 48 hours,the mRNA level of miR-873 was quantified by qRT-PCR,and cell viability was evaluated by CCK-8 assay.The levels of IL-2,IL-6,IL-10,and TNF-αin the cell supernatant were determined using ELISA assay,while cell apoptosis was detected by TUNEL staining.LC-3 protein expression was examined by immunofluorescence,and mRNA and protein expression levels of Beclin1 were analyzed by qRT-PCR and Western blot,respectively.Moreover,dual-luciferase reporter gene technology was employed to investigate the binding site between miR-873 and Beclin1.Results:Transfection of miR-873 mimic into 16HBE cells significantly upregulated the mRNA level of miR-873,which led to the inhibition of cell proliferation and the promotion of secretion of pro-inflammatory cytokines IL-2,IL-6,and TNF-α,while suppressing the secretion of anti-inflammatory cytokine IL-10.Moreover,miR-873 induced cell apoptosis and inhibited the expression of LC-3.Dual-luciferase reporter gene assay further confirmed the presence of binding sites between miR-873 and Beclin1 gene.Besides,miR-873 could target and suppress the mRNA and protein expression levels of Beclin1.Conclusion:miR-873 might modulate cell autophagy by targeting the Beclin1 gene,which can potentially promote inflammation and apoptosis in bronchial epithelial cells.
基金National Natural Science Foundation of China (No.82260134)Natural Science Foundation of Guangxi (No.2023GXNSFAA026118)+1 种基金High-level Talents Research Project of the Affiliated Hospital of Youjiang Medical College for Nationalities (No.R202011702)Guangxi Graduate Education Innovation Project (No.YCSW2023506)。
文摘Objective:To investigate the role of miR-129-5p in regulating HMGB1 expression in pancreatic cancer cell apoptosis.Methods:The untreated pancreatic cancer SW1990 cells were used as the control group.Mimics-NC(empty vector),miR-129-5p mimics,inhibitor-NC(empty vector)and miR-129-5p inhibitor were transfected into SW1990 cells by liposome transfection method as the mimics-NC group,miR-129-5p overexpression group(miR-129-5p mimics group),inhibitor-NC group and miR-129-5p low expression group(miR-129-5p inhibitor group).The binding site of miR-129-5p and HMGB1 was predicted by online target gene prediction website Target genes,and the targeting relationship between miR-129-5p and HMGB1 was verified by dual luciferase gene report experiment.The expression of miR-129-5p in each group was detected by qRT-PCR,and the expression of HMGB1 protein and apoptosis-related proteins Caspase 3 and Bcl-2 by Western blot.Hoechst staining was used to observe the changes of apoptosis.Results:Compared with the mimics-NC group and control group,miR-129-5p mimics transfection significantly up-regulated miR-129-5p level(P<0.01),inhibited HMGB1(P<0.01)and Bcl-2(P<0.05)protein expression,pro-moted Caspase 3 protein expression(P<0.05),and promoted apoptosis;compared with the inhibitor-NC group and control group,miR-129-5p inhibitor transfection significantly down-regulated miR-129-5p level(P<0.05),promoted HMGB1 and Bcl-2 protein expression(all P<0.05),inhibited Caspase 3protein expression(P<0.01),and inhibited apoptosis.The results of dual luciferase reporter gene assay showed that miR-129-5p could inhibit the fluorescence activity of wildtype HMGB1 cells and target the expression of HMGB1.Conclusion:miR-129-5p promotes the apoptosis of pancreatic cancer SW1990 cells by targeting inhibition of HMGB1 expression.
基金supported by Supported by Education Department of Hubei Province science and technology research project(NO.B2015230)Applied Fundamental Research Project of Wuhan Municipal Science and Technology Bureau(Grant No.2015061701011630)+1 种基金Medical Scientific Research Project of Health and Familly Planning Commission of Wuhan Municipality(Grant No.WX16E12)the fourth batch of "Hanyang Talent Associate Program"
文摘Objective:To investigate the effects of co-transfection of miR-520c-3p and miR-132 on proliferation and apoptosis of hepatocellular carcinoma Huh7.Methods:Hepatocellular carcinoma Huh7 was cultured in vitro and lipidosome was used to transfect miR-520c-3p and miR-132 respectively or together.The effects of transfection of miR-520c-3p and miR-132 on proliferation and apoptosis of Huh7 were detected by CCK8 and Annexin V staining and flow cytometry,and the expression level of the targeted gene of over-expressed miR-520c-3p and miR-132 was determined by Western blot and realtime PCR.Results:Compared with the control group,the proliferation ability of Huh7 of the single transfected and co-transfected miR-520c-3p and miR-132 decreased significantly,and the apoptosis ratio increased distinctly(P<0.05).Besides,the affect of the co-transfection group was better than that of the single transfection group.The protein levels of GPC3(Glypican-3) and YAP(Yes-associated protein),the target genes transfected only by miR-520c-3p and miR-132,respectively,reduced obviously(P<0.05),which was similar with the co-infected cells,but cells transfected by miR-132 only showed a decrease of YAP.Conclusions:The co-transfection of miR-520c-3p and miR-132 can target-regulate the expression of GPC3 and YAP,enhance the exhibition effect on proliferation of hepatocellular carcinoma Huh7 and induce cell apoptosis synergistically.
基金supported in part by the National Natural Science Foundation of China(No.8157279091629302+1 种基金 31270818)the Natural Science Foundation of Tianjin (No.12JCZDJC25100,14JCYBJC26400,16JCYBJC42400)
文摘Objective：Previous studies have identified that kazrin is a constituent of desmosome and influences intercellular adhesion,growing development and morphology.We previously cloned another new isoform,kazrin F and found that it has anti-apoptotic effects on human glioma cell line.To further explore whether kazrin F is involved in tumorigenesis,we investigated its expression and role in cervical cancer（CC） cells.Methods：The role of kazrin F and miR-186 in CC was determined by 3-（4,5-dimethylthiazol-2-yl）-2,5-diphenyltetrazolium bromide（MTT） assay,colony formation,transwell,and apoptosis assays.Using enhanced green fluorescent protein（EGFP） reporter assays,reverse transcription-quantitative polymerase chain reaction（RT-qPCR） and western blot analysis,we identified kazrin F post-transcriptional regulation by miR-186.Results：We demonstrate that kazrin F is highly expressed in CC tissues compared with the adjacent noncancerous tissues and promotes cell proliferation,colony formation,migration and invasion in HeLa and C33 A cells by suppressing apoptosis and facilitating epithelial-to-mesenchymal transition（EMT）.Furthermore,miR-186 was confirmed as a regulator of kazrin F dysregulation.An EGFP reporter assay proved that miR-186 directly targets the 3＇-untranslated region（3＇UTR） of kazrin F and downregulates its expression,and miR-186 expression showed an inverse correlation with kazrin F levels in CC tissues.In addition,overexpression of miR-186 suppressed the malignant behaviors of CC cells.The ectopic expression of kazrin F rescued the inhibitory effects of miR-186.Conclusions：Our findings indicate that the upregulation of kazrin F due to downregulated miR-186 levels contributes to malignancy,and highlight the significance of kazrin F in CC tumorigenesis.
