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BACKGROUND:Grape seed extract can inhibit chondrocyte apoptosis and aging,and improve osteoarthritis.However,the effects of grape seed extract on the apoptosis of chondrocytes in the growth plate and tibial growth are still unclear. OBJECTIVE:To investigate the effect of grape seed extract on interleukin-1β-induced apoptosis in rat growth plate cells and on tibial bone growth. METHODS:(1)Cell experiment:Growth plate chondrocytes from Sprague-Dawley rats were isolated,cultured,and identified.The cells were then randomly divided into control group,model group,grape seed extract group,miR-138-5p NC group and miR-138-5p inhibitor group.In the model group,20 ng/mL interleukin-1β was used to induce apoptosis in rat growth plate chondrocytes.In the grape seed extract group,20 ng/mL interleukin 1β was added along with 10 μmol/L grape seed extract solution for 48 hours.Cells in the miR-138-5p NC and inhibitor groups were transfected with 5 nmol/L miR-138-5p NC and 5 nmol/L miR-138-5p inhibitor for 12 hours,respectively,followed by addition of 20 ng/mL interleukin-1β.qRT-PCR was used to detect miR-138-5p and caspase-3 expression.Luciferase reporter assay was used to analyze the relationship between miR-138-5p and caspase-3 targeting.Cell counting kit-8 was used to detect cell proliferation activity.Flow cytometry was used to detect cell apoptosis.Caspase-3 and Bcl-2 protein expressions were detected by western blot.(2)Animal experiment:The animals were divided into normal control group,grape seed extract group and miR-138-5p inhibitor group.The effects of grape seed extract on epiphyseal closure and tibial growth of the tibial plateau in rats were observed. RESULTS AND CONCLUSION:miR-138-5p had a targeting relationship with caspase-3.Compared with the control group,cell proliferation was significantly reduced,apoptosis was significantly increased(P＜0.01),miR-138-5p,Bcl-2 expression was reduced(P＜0.01),and caspase-3 expression was increased(P＜0.01)in the model group.Compared with the mod group,the grape seed extract group showed a significant increase in cell proliferation,a significant decrease in apoptosis(P＜0.01),an increase in miR-138-5p and Bcl-2 expression(P＜0.01)and a decrease in caspase-3 expression(P＜0.01).Compared with the grape seed extract group,the miR-138-5p inhibitor group showed a significant decrease in cell proliferation,a significant increase in apoptosis(P＜0.01),a decrease in miR-138-5p and Bcl-2 expression(P＜0.05 or P＜0.01),and an increase in caspase-3 expression(P＜0.05).Intragastric administration of grape seed extract could delay epiphyseal closure of the tibial plateau and promote tibial bone growth in rats,whereas miR-138-5p inhibitor intervention inhibited the effect of grape seed extract on tibial bone growth in rats.To conclude,grape seed extract can inhibit apoptosis of rat growth plate chondrocytes through regulating the miR-138-5p/caspase-3 pathway,improve epiphyseal closure of rat tibial plateau and promote tibial bone growth.
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Objective:To investigate the effects of serum exosomes of patients with gastric cancer on cisplatin resistance, clonal formation, migration, invasion and apoptosis of the AGS gastric cancer cells, and the corresponding molecular mechanisms.Methods:The experimental study was conducted. The exosomes of patients with gastric cancer was separated from their serum, and the expression of lncRNA HOTTIP was analyzed using the quantitative real time polymerase chain reaction (qRT-PCR). Normal gastric epithelial cell line GES1, gastric cancer cell line AGS and human embryonic kidney cell 293T were cultured in vitro. AGS cells were incubated with exosomes (Exo),with phos-phate buffered saline (PBS) treatment as control, and transfected with si-NC or si-HOTTIP-3, named as Exo group, PBS group, si-NC+Exo group, and si-HOTTIP-3+Exo group. The AGS cells were trans-fected with si-NC, si-HOTTIP-1, si-HOTTIP-2, si-HOTTIP-3, oe-HOTTIP, vector, oe-HOTTIP+miR-138-5p mimic, oe-HOTTIP+mimic NC, miR-138-5p inhibitor, inhibitor NC, miR-138-5p inhibitor+si-TJP1 and miR-138-5p inhibitor+si-NC. They were recorded as si-NC group, si-HOTTIP-1 group, si-HOTTIP-2 group, si-HOTTIP-3 group, oe-HOTTIP group, vector group, oe-HOTTIP+miR-138-5p mimic group, oe-HOTTIP+mimic NC group, miR-138-5p inhibitor group, inhibitor NC group, miR-138-5p inhibitor+si-TJP1 group and miR-138-5p inhibitor+si-NC group. The 293T cells transfected with mimic NC+HOTTIP wt, miR-138-5p mimic+HOTTIP wt, mimic NC+HOTTIP mut, miR-138-5p mimic+HOTTIP mut, mimic NC+TJP1 3'UTR wt, miR-138-5p mimic+TJP1 3'UTR wt, mimic NC+TJP1 3'UTR mut, miR-138-5p mimic+TJP1 3'UTR mut were recorded as the mimic NC+HOTTIP wt group, miR-138-5p mimic+HOTTIP wt group, mimic NC+HOTTIP mut group, miR-138-5p mimic+HOTTIP mut group, mimic NC+TJP1 3'UTR wt group, miR-138-5p mimic+TJP1 3'UTR wt group, mimic NC+TJP1 3'UTR mut group, miR-138-5p mimic+TJP1 3'UTR mut group. The cell counting kit-8 (CCK8) was used to analyze the cisplatin sensitivity of gastric cancer cells. The colony formation experiment was used to analyze the colony formation of gastric cancer cells. The Transwell experiment was used to analyzed cell migration and invasion of gastric cancer cells. The flow cytometry experiment was used to analyze cell apoptosis of gastric cancer cells. The Western bolt assay was used to analyze the expression of exosome marker proteins, including the CD63 and CD81, and the protein of TJP1, the drug-resistance related proteins, including the P-gp and MCL-1. The dual-luciferase assay was used to analyze the targeted relationships among lncRNA HOTTIP, miR-138-5p and TJP1. Observation indicators: (1) expression of lncRNA HOTTIP; (2) resistance of gastric cancer cells to cisplatin regulated by exosome-mediated lncRNA HOTTIP; (3) regulation of cisplatin resistance in gastric cancer cells mediated by miR-138-5p through lncRNA HOTTIP overexpression; (4) targeting of TJP1 gene 3′-untranslated region (UTR) by miR-138-5p; (5) regulation of cisplatin resistance in gastric cancer cells by TJP1 through miR-138-5p inhibition. Measurement data with normal distribution were represented as Mean± SD, and comparison between groups was conducted using the t test. The one-way ANOVA was used for comparison for multiple groups and the Tukey′s test was used for further pairwise compari-son. Count data were described as absolute numbers, and the chi-square test was used for comparison. Correlation analysis was conducted using the Pearson′s test. Results:(1) Expression of lncRNA HOTTIP. The expression of lncRNA HOTTIP in the serum exosome of patients with gastric cancer was higher than that in healthy volunteers, showing a significant difference ( P<0.05). Results of transmi-ssion electron microscopy examination showed that the serum exosomes were circular or oval in shape. Results of Western bolt assay showed the expression of marker proteins of CD63 and CD81 in serum exosomes. (2) Resistance of gastric cancer cells to cisplatin regulated by exosome-mediated lncRNA HOTTIP. Compared with the PBS group, the biochemical half maximal inhibitory concentra-tion (IC50), the number of clone formation, the number of invasive cell, the number of migratory cell, expression of P-gp protein, expression of MCL-1 protein in the Exo group increased, while the cell apoptosis rate decreased, showing significant differences between them ( P<0.05). Compared with the si-NC+Exo group, the IC50, the number of clone formation, the number of invasive cell, the number of migratory cell, expression of P-gp protein, expression of MCL-1 protein in the si-HOTTIP-3+Exo group decreased, while the cell apoptosis rate increased, showing significant differences between them ( P<0.05). (3) Regulation of cisplatin resistance in gastric cancer cells mediated by miR-138-5p through lncRNA HOTTIP overexpression. Compared with the vector group, the IC50, the number of clone formation, the number of invasive cell, the number of migratory cell, expression of P-gp protein, expression of MCL-1 protein in the oe-HOTTIP group increased, while the cell apoptosis rate decreased, showing significant differences between them ( P<0.05). Compared with the oe-HOTTIP+mimic NC group, the IC50, the number of clone formation, the number of invasive cell, the number of migratory cell, expression of P-gp protein, expression of MCL-1 protein in the oe-HOTTIP+miR-138-5p mimic group increased, while the cell apoptosis rate decreased, showing significant differences between them ( P<0.05). (4) Targeting of TJP1 gene 3′-UTR by miR-138-5p. Results of dual-luciferase assay showed that the luciferase activity in 293T cells treatment with mimics of control+vectors of wild type of TJP1 gene 3′-UTR and 293T cells treatment with mimics of miR-138-5p+vectors of wild type of TJP1 gene 3′-UTR was 1.00±0.09 and 0.21±0.03, respectively, showing a significant difference between them ( t=15.02, P<0.05). (5) Regulation of cisplatin resistance in gastric cancer cells by TJP1 through miR-138-5p inhibition. Compared with the inhibitor group, the IC50, the number of clone formation, the number of invasive cell, the number of migratory cell, expression of P-gp protein, expression of MCL-1 protein in the miR-138-5p inhibitor group increased, while the cell apoptosis rate decreased, showing significant differences between them ( P<0.05). Compared with the miR-138-5p inhibitor+si-NC group, the IC50, the number of clone formation, the number of invasive cell, the number of migratory cell, expression of P-gp protein, expression of MCL-1 protein in the miR-138-5p inhibitor+si-TJP1 group decreased, while the cell apoptosis rate increased, showing significant differences between them ( P<0.05). Conclusion:Serum exosomes-mediated lncRNA HOTTIP can promote cisplatin resistance, clonal formation, migration, invasion and apoptosis of gastric cancer cells and inhibit cell apoptosis of gastric cancer cells through regulating the expression of miR-138-5p/TJP1.
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Objective:To study the effect of Long non-coding RNA(LncRNA)small nucleolar RNA host gene 12(SNHG12)regulating miR-138-5p/hypoxia inducible factor-1(HIF-1α)axis on improving the damage of hypoxia/reoxygenation(H/R)human vas-cular endothelial cells.Methods:Human umbilical vein endothelial cells(HUVECs)were cultured in vitro and randomly divided into control group,H/R model group,H/R+LncRNA SNHG12 overexpression group,H/R+miR-138-5p mimics group,H/R+co-transfec-tion group and H/R+co-transfection negative control group,each transfection group was transfected separately,and except for control group,the remaining groups were given hypoxia for 5 hours and then reoxygenated for 1 hour to induce the cell models,and then the cell viability of each group was detected by CCK-8 experiment;the cell apoptosis in each group was detected by flow cytometry experi-ment,and the apoptosis rate of each group was compared;the levels of reactive oxygen species(ROS),lactate dehydrogenase(LDH)and inflammatory factors IL-6,IL-17 and IL-18 in each group were measured by the kit;the expressions of miR-138-5p and HIF-1α mRNA in cells of each group were measured by real-time quantitative PCR(qRT-PCR)experiment;the expressions of apoptotic pro-teins caspase-9,Bcl-2-associated X protein(Bax)and HIF-1α in each group were evaluated by Western blot.Results:Compared with control group,the apoptosis rate,cellular ROS,LDH,IL-6,IL-17 and IL-18 levels,cellular HIF-1α mRNA and protein levels,cellular caspase-9,Bax and HIF-1α protein levels were increased in H/R model group(P<0.05),the cell viability and miR-138-5p level were decreased(P<0.05).Compared with H/R model group and H/R+co-transfection group,the cell viability,cell HIF-1αmRNA and protein levels were increased in H/R+LncRNA SNHG12 overexpression group(P<0.05),the apoptosis rate,cellular ROS,LDH,IL-6,IL-17 and IL-18 levels,cellular caspase-9 and Bax protein levels,and miR-138-5p level were decreased(P<0.05);the cell viability,cellular HIF-1α mRNA and protein levels were decreased in H/R+miR-138-5p mimics group(P<0.05),the apoptosis rate,cellular ROS,LDH,IL-6,IL-17 and IL-18 levels,cellular caspase-9 and Bax protein levels were increased(P<0.05).Com-pared with H/R model group,there was no significant difference in cell index levels between the H/R+co-transfection negative control group and the H/R+co-transfection group(P>0.05).Conclusion:LncRNA SNHG12 can upregulate HIF-1α expression by downregulat-ing miR-138-5p expression,inhibit H/R-induced inflammation and oxidative stress in HUVECs,and reduce cell damage and apoptosis.
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AIM: To study the effects and mechanism of down-regulating lncRNA LINC00176 on cisplatin resistance and autophagy in lung cancer A549/DDP cells. METHODS: The qRT-PCR method was used to determine the expression changes of LINC00176 in normal bronchial epithelial 16HBE cells and lung cancer A549, A549/DDP, NCI-H1299 and SK-MES-1 cells. A549/DDP cells were divided into Control group, si-NC group, and si-LINC00176 group, si-LINC00176+ Anti-miR-NC group, and si-LINC00176+ Anti-miR-138-5p group. MTT experiment detected the half inhibitory concentration (IC
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AIM: To study the effect of miR-138-5p on the function of β cell in gestational diabetes mellitus (GDM) and its related mechanism. METHODS: The expression of miR-138-5p in peripheral blood of 15 GDM pregnant women and 15 normal pregnant women were compared by RT-qPCR. miR-138-5p mimic and inhibitor were transfected into INS-1 cells, respectively, and their expression level was over expressed or inhibited. RT-qPCR was used to verify the transfection efficiency.MTT proliferation experiment, Annexin V-FITC apoptosis experiment and insulin release experiment were used to detect the effects of miR-138-5p on INS-1 cell proliferation, apoptosis and insulin release ability. The target gene of miR-138-5p was screened by TargetScan, a miRNA target gene prediction software. The functional rescue experiment confirmed whether miR-138-5p could exert its influence on INS-1 cell proliferation, apoptosis and insulin release ability by targeting its target gene. Western blot was used to detect the molecular signaling pathway of miR-138-5p in INS-1 cells.RESULTS: The expression of miR-138-5p in peripheral blood of GDM pregnant women was significantly lower than that of normal pregnant women. RT-qPCR showed that miR-138-5p mimic and inhibitor could significantly promote or inhibit the expression of miR-138-5p in INS-1 cells. The results of MTT proliferation experiment, Annexin V-FITC apoptosis experiment and insulin release experiment indicated that over expression of miR-138-5p could significantly promote the proliferation of INS-1 cells, inhibit the apoptosis of cells and promote the insulin release ability of cells. However, down-regulating the expression of miR-138-5p could significantly inhibit the proliferation of INS-1 cells, promote apoptosis and inhibit insulin release. HIF-1α was selected as the target gene of miR-138-5p by TargetScan. The double luciferase gene report and Western blot showed that miR-138-5p could inhibit the expression of HIF-1α in INS-1 cells. The functional rescue experiment confirmed that miR-138-5p could affect the proliferation, apoptosis and insulin release of INS-1 cells by regulating the expression of HIF-1α. Western blot showed that miR-138-5p may play a role in INS-1 cells by affecting PI3K, Akt and p-PI3K, p-Akt protein after phosphorylation in PI3K/Akt signaling pathway. CONCLUSION: miR-138-5p may reguLate HIF-1α expression in a targeted manner, thereby affecting the PI3K/AKT signaling pathway, promoting the proliferation and inhibition of the parent cells' apoptosis, and promoting their insulin-releasing ability to protect the function of β cell in GDM.
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BACKGROUND: Cisplatin resistance (DDP-resistance) remains one of the major causes of poor prognosis in females with ovarian cancer. Long non-coding RNAs (lncRNAs) have been shown to participate in the regulation of cellular processes, including chemoresistance. The aim of this study was to explore the role of HOX transcript antisense RNA (HOTAIR) in DDP-resistant ovarian cancer cells. METHODS: DDP-resistant ovarian cancer cell lines (SKOV3/DDP and A2780/DDP) were established. Real-time PCR, western blot, dual-luciferase reporter assay, and flow cytometry were then used to evaluate the effect of HOTAIR/miR-138-5p axis on chemoresistance of DDP-resistant ovarian cancer cells to DDP. RESULTS: We found that HOTAIR was upregulated in DDP-resistant cells, while miR-138-5p was downregulated. Knockdown of HOTAIR increased the expression of miR-138-5p in DDP-resistant cells and miR-138-5p is directly bound to HOTAIR. Upregulation of miR-138-5p induced by HOTAIR siRNA or by its mimics enhanced the chemosensitivity of DDP-resistant cells and decreased the expression of EZH2 (enhancer of zeste 2 polycomb repressive complex 2 subunit) and SIRT1 (sirtuin 1). Furthermore, the HOTAIR silencing-induced chemosensitivity of DDP-resistant cells was weakened by miR-138-5p inhibitor. CONCLUSIONS: These data demonstrate that HOTAIR acts as a sponge of miR-138-5p to prevent its binding to EZH2 and SIRT1, thereby promoting DDP-resistance of ovarian cancer cells. Our work will shed light on the development of therapeutic strategies for ovarian cancer treatment.
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Humains , Femelle , Tumeurs de l'ovaire/génétique , Cisplatine/pharmacologie , Résistance aux médicaments antinéoplasiques/génétique , ARN long non codant/génétique , Régulation de l'expression des gènes tumoraux/effets des médicaments et des substances chimiques , Régulation positive , Apoptose/effets des médicaments et des substances chimiques , microARN/antagonistes et inhibiteurs , Lignée cellulaire tumorale , Techniques de knock-out de gènes/méthodes , Sirtuine-1/antagonistes et inhibiteurs , Réaction de polymérisation en chaine en temps réel , Protéine-2 homologue de l'activateur de Zeste/antagonistes et inhibiteursRÉSUMÉ
Objective@#Abstract: Objective: To investigate the expression and significance of miR-138 and programmed cell death protein 1 (PD-1) in the patients with hepatitis B virus (HBV)-related hepatocellular carcinoma (HCC). @*Methods@#A total of 30 patients with HBV-related HCC, 20 with HBV-related cirrhosis (LC) and 30 with chronic hepatitis B (CHB) were recruited from Jiaozuo People′s Hospital. The blood samples from all patients and the peritoneal effusion samples from HCC and LC patients were collected. The levels of miR-138 and soluble PD-1 (sPD-1) in blood and peritoneal effusion samples were detected by real-time PCR and ELISA, respectively. The expressions of PD-1 in T lymphocytes were measured with flow cytometry and western blot. The targeting effect of miR-138 on the 3′-non-coding region (3′-UTR) of PD-1 gene was verified by the dual-luciferase reporter gene system. @*Results@#The relative expression levels of miR-138 in the peritoneal effusion and plasma of HBV-related HCC patients were significantly lower than those in LC and CHB patients (P＜0.05). The serum sPD-1 levels and the expression levels of PD-1 in CD3 + T lymphocytes of HBV-related HCC patients were significantly higher than those in LC and CHB patients (P＜0.05). The relative expression levels of miR-138 were negatively correlated with serum sPD-1 levels and the expression levels of PD-1 in CD3 + T lymphocytes (P＜0.05). The dual-luciferase reporter gene system and western blot results demonstrated that there was a targeting relationship between miR-138 and the 3′-UTR of PD-1 gene. After miR-138 was transfected, the expression level of PD-1 was significantly down-regulated. @*Conclusion@#miR-138 participates in the development and progression of HBV-related HCC probably by targeting PD-1.
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PURPOSE: Adipogenic differentiation of adipose tissue-derived mesenchymal stem cells (AMSCs) is critical to many disease-related disorders, such as obesity and diabetes. Studies have demonstrated that miRNA-138 (miR-138) is closely involved in adipogenesis. However, the mechanisms affected by miR-138 remain unclear. This work aimed to investigate interactions between miR-138 and lipoprotein lipase (LPL), a key lipogenic enzyme, in AMSCs. MATERIALS AND METHODS: Human AMSCs (hAMSCs) isolated from human abdomen tissue were subjected to adipogenic differentiation medium. Quantitative real-time polymerase chain reaction and Western blot assay were applied to measure the expressions of miR-138, LPL, and the two adipogenic transcription factors cytidine-cytidine-adenosine-adenosine-thymidine enhancer binding protein alpha (C/EBPα) and peroxisome proliferator-activated receptor gamma (PPARγ). The relationship between miR-138 and LPL was predicted utilizing the miRTarBase database and validated by dual luciferase reporter assay. RESULTS: Showing increases in C/EBPα and PPARγ expression levels, hAMSCs were induced into adipogenic differentiation. During adipogenesis of hAMSCs, miR-138 expression was significantly downregulated. Overexpression of miR-138 by transfection inhibited hAMSCs adipogenic differentiation in vitro. Mechanically, LPL was a target of miR-138. LPL expression was upregulated during adipogenesis of hAMSCs, and this upregulation was reversed by miR-138 overexpression. Functionally, silencing of LPL by transfection exerted similar inhibition of the expressions of C/EBPα and PPARγ. Meanwhile, LPL ectopic expression was able to partly abolish the suppressive effect of miR-138 overexpression on adipogenic differentiation of hAMSCs. CONCLUSION: Upregulation of miR-138 inhibits adipogenic differentiation of hAMSCs by directly downregulating LPL.
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Humains , Abdomen , Adipogenèse , Technique de Western , Protéines de transport , Expression génique ectopique , Techniques in vitro , Lipoprotein lipase , Lipoprotéines , Luciferases , Cellules souches mésenchymateuses , Obésité , Récepteur PPAR gamma , Réaction de polymérisation en chaine en temps réel , Facteurs de transcription , Transfection , Régulation positiveRÉSUMÉ
Myocardial infarction (MI) is a common presentation for ischemic heart disease, which is a leading cause of death. Emodin is a Chinese herbal anthraquinone used in several diseases. However, the effect of emodin in hypoxia-induced injury in cardiomyocytes has not been clearly elucidated. Our study aimed to clarify the functions of emodin in hypoxia-induced injury in rat cardiomyocytes H9c2 and explore the underlying mechanism. The effects of emodin on cell viability and apoptosis were analyzed by the Cell counting kit-8 assay and flow cytometry assay, respectively. The cell proliferation- and cell apoptosis-related proteins were detected by western blot. qRT-PCR was used to determine the relative expression of miR-138. Cell transfection was performed to alter miR-138 and MLK3 expression. miR-138 target was performed by dual luciferase activity assay. Sirt1/AKT and Wnt/β-catenin pathways-related factors phosphorylation were analyzed by western blot. Emodin inhibited hypoxia-induced injury in H9c2 cells by promoting cell viability and reducing cell apoptosis. miR-138 was down-regulated by hypoxia treatment but up-regulated by emodin. Up-regulation of miR-138 alleviated hypoxia-induced cell injury. Down-regulation of miR-138 attenuated the growth-promoting effect of emodin on hypoxia-induced injury, whereas up-regulation of miR-138 enhanced the growth-promoting effects of emodin. The underlying mechanism might be by inactivating Sirt1/AKT and Wnt/β-catenin pathways. MLK3 was negatively regulated by miR-138 expression and inactivated Sirt1/AKT and Wnt/β-catenin pathways. Emodin alleviated hypoxia-induced injury in H9c2 cells via up-regulation of miR-138 modulated by MLK3, as well as by activating Sirt1/AKT and Wnt/β-catenin pathways.
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Animaux , Rats , Hypoxie cellulaire/effets des médicaments et des substances chimiques , Survie cellulaire/effets des médicaments et des substances chimiques , Émodine/usage thérapeutique , Myocytes cardiaques/anatomopathologie , Prolifération cellulaire/effets des médicaments et des substances chimiques , Hypoxie/complications , Transduction du signal , Régulation positive , Lignée cellulaire , Myocytes cardiaques/effets des médicaments et des substances chimiques , microARNRÉSUMÉ
Objective To detect the expression of miR-138 in lens tissues of agerelated cataract and explore the effects of miR-138 on the proliferation and apoptosis of human lens epithelial cells and its possible target genes.Methods Real-time quantitative PCR (RT-qPCR) was applied for the detection of the expression of miR-138 and prediction of target gene sirtuin (silent information regulator 1) (SIRT1) in patients with age-related cataract (cataract group) and anterior lens capsules (normal control group).Then miR-138 mimics,mimic controls,miR-138 inhibitors and inhibitor controls were transfected into the human lens epithelial cell line (SRA01/04),and the expression of SIRT1 mRNA and protein was detected by RT-qPCR and Western blot,accordingly.At 72 hours after transfection,the cells were exposed to 200 μmol · L-1 H2O2 for 1 hour,followed by detection of the activity of Caspase-3 by the Caspase-3 activity assay kit,and identification of the targeted relationship between miR-138 and SIRT1 by dual luciferase reporter assays.Results Compared with the normal control group,the expression of miR-138(3.64 ±0.19) was significantly increased (P ＜0.001),but the expression of SIRT1 mRNA(0.32 ± 0.06) was significantly decreased (P ＜ 0.001) in the cataract group.Moreover,The expression levels of SIRT1 mRNA(0.42 ± 0.05) and protein(0.46 ± 0.05) in cells transfected with miR-138 mimics were significantly decreased,while the activity of Caspase-3 (3.24 ± 0.17) was significantly elevated when compared with cells transfected with minic controls (all P ＜ 0.05);Compared with cells transfected with inhibitor controls,the expressions of SIRT1 mRNA(2.95 ±0.13) and protein(1.98 ±0.12) were significantly upregulated,whereas Caspase-3 activity(0.42 ±0.05) was significantly decreased in cells transfected with miR-138 inhibitors (all P ＜0.05).And fmally,dual luciferase reporter assays showed the confirmation SIRT1 as a direct target of miR-138.Conclusion miR-138 is highly expressed in the lens capsule of age-related cataract patients,and it can promote the apoptosis of lens epithelial cells by negatively regulating the expression of SIRT1.
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microRNA (miRNA, miR) has attracted great interest in recent years. miR-138 is closely related to the occurrence, development and prognosis of different kinds of malignant tumors.Present studies have shown that miR-138 is frequently down-regulated in tumor cells and tissues. Down-regulation expression of miR-138 can promote the proliferation, invasion and metastasis in a variety of malignant tumor cells. Up-regulation expression of miR-138 may play a crucial role in tumor suppression, and enhance sensitivity of tumor cells to radiotherapy and chemotherapy. Therefore, miR-138 can be considered as a biomarker of tumor prognosis, and also one of the tumor therapeutic targets.
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Objective To determine the effect of miR-138 on SD rat renal ischemia-reperfusion injury its mechanism.Methods 60 adult SD rats were randomly divided into Sham group,renal ischemia-reperfusion (RIRI) group,RIRI + vector group,RIRI + miR-138 group,RIRI + miR-138 shRNA group,and then we established renal reperfusion injury model by occlusion bilateral renal pedicle.Creatinine,blood urea nitrogen in serum are tested to compare differences in renal function in each group,and TUNEL staining was taked to detect apoptosis,real time PCR were taked to test the exression of p53 and it's pathway.Results When compared with the sham group,expression of kidney function BUN/Crin serum,percentage of apoptoiscells of reperfusion injury group wrer all increased,and the differences were statistically significant (P ＜ 0.05).Upregulate miR-138 increased these indexes,and the difference was statistically significant (P ＜0.05),and down regulate miR-138 would protect the injure of renal ischemia-reperfusion.Further studies showed that miR-138 may play the role by inhibiting the p53 signaling pathway.Conclusion MiR-138 has a significant aggravating effect on renal ischemia-reperfusion injury,and the mechanism may be related to p53 signaling pathway.
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miRNA was 20～25 nt endogenous non-coding RNA. miRNAs are encoded small RNAs that hybridize with messenger RNAs, resulting in degradation or translational inhibition of targeted transcripts. In order to investigate the function of miR-138 on mouse mammary epithelial cells, technique for gene silencing-miRNA inhibitor (anti-miRNA) was applied to make miR-138 silence, qRT-PCR was showed valid for inhibitor miR-138. And Western blot, CASY(r)-technology was put in use to study some change of mouse mammary epithelial cells after miR138 inhibitor. It was shown that miR-138 suppresses the exepress of PRL-R(P