Résumé
Background: Expression of microRNA⁃320 (miR⁃320) is down regulated in acute pancreatitis, and the mechanism of its effect on acute pancreatitis is still unclear. Aims: To investigate the effect of miR⁃320 on intestinal injury in rats with acute pancreatitis and its mechanism. Methods: Rats were randomly divided into sham operation group, model group, miR⁃ 320 agonist group (agomir miR ⁃ 320 group), miR ⁃ 320 agonist control group (agomir NC group), JAK2 inhibitor group (AG490 group), and NF⁃κB pathway inhibitor group (PDTC group). The rat model of acute pancreatitis was established by retrograde injection of 5% sodium taurocholate to the bile duct. The automatic biochemical analyzer was used to detect serum levels of amylase and lipase; ELISA assay was used to detect serum levels of TNF⁃α and IL⁃1β; HE staining was used to observe the pathological changes of rat pancreas and ileum; TUNEL staining was used to observe cell apoptosis in rat ileum; real⁃time fluorescent quantitative PCR (RT⁃qPCR) was used to detect the expression of miR⁃320 in ileum tissue; Western blotting method was used to detect the expressions of JAK2/STAT3 and NF⁃κB signaling pathway related proteins in ileum. Results: Compared with sham operation group, the pancreas and ileum were severely injured in model group, and the pathological score and ileum cell apoptosis were significantly increased (P<0.05), serum levels of amylase, lipase, TNF⁃ α, and IL⁃1β were significantly increased (P<0.05), the expression of miR⁃320 in ileum tissue was significantly decreased (P<0.05), the ratios of p⁃JAK2/JAK2, p⁃STAT3/STAT3, p⁃p65/p65, and p⁃IκBα/IκBα in ileum tissue were significantly increased (P<0.05). Compared with model group, the pathological damages of pancreas and ileum in agomir miR ⁃ 320 group, AG490 group and PDTC group were reduced, and the pathological score and ileum cell apoptosis were significantly decreased (P<0.05), serum levels of amylase, lipase, TNF ⁃ α, and IL ⁃ 1β were significantly decreased (P<0.05), the expression of miR⁃320 in ileum tissue was significantly increased (P<0.05), the ratios of p⁃JAK2/JAK2, p⁃STAT3/STAT3, p⁃ p65/p65, and p⁃IκBα/IκBα in ileum tissue were significantly decreased (P<0.05). Conclusions: MiR⁃320 can improve the intestinal injury in rats with acute pancreatitis by inhibiting the activation of JAK2/STAT3 and NF⁃κB signaling pathways.
Résumé
Objective Long non-coding RNA(lncRNA) are aberrantly expressed in breast cancer(BC) and strongly associated with its survival prognosis. The aim of this study is to investigate the expression and effect of IncRNA SPATA31D5P on the invasion and migration capacity of breast cancer cells through adsorption of miR-320a. Methods Totally 30 cases of BC tissues and paraneoplastic tissues were collected, and the expression levels of SPATA31D5P in BC tissues and BC cell lines were detected by Real-time PCR. MDA-MB-231 cells were transfected with SPATA31D5P siRNA interference vector, and cell proliferation, invasion and migration capacity were determined using the cell counting kit-8 assay (CCK-8), 5-ethynyl-2'- deoxyuridine(EdU), Transwell and wound-healing assay respectively. And cell cycle and apoptosis were detected by flow cytometry. Bioinformatics approachs were used to screen for miRNAs that could bind complementarily to SPATA31D5P, and the regulatory effect of SPATA31D5P on miR-320a was detected by Real-time PCR and dual luciferase reporter assay. Results SPATA31D5P levels were significantly higher in BC tissues than in adjacent normal breast tissues, and SPATA31D5P expression was higher in each BC cell line than in normal breast epithelial cells MCF10 A. The level of SPATA31D5P in the interference group was 0. 288±0. 052, which was lower than that of the blank control group 1. 114±0. 096 and negative control (NC) group 1. 079±0. 128 (P< 0. 01). The proliferation activity of MDA- MB-231 cells in the interfered group was significantly reduced and apoptotic rate was obviously increased compared to the NC and control groups (P<0. 01) ;the Gj phase block was observed in the interfered group; the scratch healing rate and number of perforated cells in the interference group were (14. 36 ± 1. 75) % and (26±1.52), which were lower than (52. 25± 1.87)% and ( 67. 33 ± 2. 91 ) of the NC group (PcO.Ol). Dual luciferase experiments confirmed that SPATA31D5P could directly regulate miR-320a expression and luciferase activity. Conclusion SPATA31D5P is highly expressed in BC, interfering with SPATA31D5P expression effectively inhibits the proliferation, migration and invasion of MDA-MB-231 cells, and the mechanism may be related to the targeted regulation of miR-320a.
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Aim To investigate the effect of miR-320a up-regulation on the apoptosis and migration of Bel7402 cells induced by ribonucleic acid Ⅱ.Methods The different expression levels of miR-320a in normal liver cells and hepatocellular carcinoma (HCC) cells were detected by qRT-PCR.Bel-7402 cell was transfected with miR-320a mimic,and the miR-320a expression levels were measured by qRT-PCR.The effect of ribonucleic acid Ⅱ on proliferation of Bel-7402 and Bel-7402-miR-320a cells was measured by CCK-8 assay,and cell cycle and apoptosis were detected by flow cytometry.The migration and invasion ability of ribonucleic acid Ⅱ on Bel-7402 cells were tested by Transwell method.The expression of p53,Cyclin D1,Bax,Bcl-2,MMP-3 proteins were examined by Western blot.Results miR-320a expression levels in HCC cell line Bel-7402 were significantly lower than those in normal cell line HL-7702.Bel-7402 cells were successfully transfected with miR-320a mimic,named Bel-7402-miR-320a.CCK-8 showed that ribonucleic acid Ⅱ could effectively inhibit the proliferation of Bel7402 and Bel-7402-miR-320a cells in vitro in a dosedependent manner at the range of 100,200,300,400,500 mg · L-1.The IC50 of ribonucleic acid Ⅱexposure on Bel-7402 and Bel-7402-miR-320a cells for 12 h and 24 h was 250,200 mg · L-t and 150,120 mg · L-1,respectively;flow cytometric analysis indicated that over-expression of miR-320a could arrest Bel-7402 and Bel-7402-miR-320a cells induced by ribonucleic acid Ⅱ in G0/G1 phase,and promote the apoptosis of HCC cells.Transwell method showed that Bel-7402-miR-320a + Ribonucleic acid Ⅱ group could significantly inhibit the migration of HCC cells compared with control group and Bel-7402 + Ribonucleic acid Ⅱ group.Western blot results showed that the expression of p53,Bax proteins increased,while the Cyclin D1,Bcl-2,MMP-3 proteins were down-regulated in Bel-7402 and Bel-7402-miR-320a cells induced by ribonucleic acid Ⅱ.Conclusions The expression of miR-320a is lower in HCC cells than that in normal cell line.While ribonucleic acid Ⅱ could promote the apoptosis of liver cancer cells by arresting the cell cycle protein expression of Cyclin D1,activating p53 signaling pathway,down-regulating Bcl-2,up-regulating Bax and destroying Bcl-2/Bax proportions,and inhibiting the migration and invasion of HCC cells by downregulating MMP-3.Overexpression of miR-320a could increase the sensitivity and boost the pharmacological effects of ribonucleic acid Ⅱ on HCC cells.
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[Objective]To investigate the inhibitory effect and mechanism of the microRNA-320d(miR-320d)on epithelial mesenchymal transition in endometrial carcinoma JEC cells.[Methods]JEC endometrial carcinoma cell lines were transfected with miR-320d mimics or negative control mimic,respectively,as M320d or NCM group. Control group was established with untreated JEC endometrial carcinoma cells. miR-320d content in each group was detected by RT-PCR method. Transwell assay was used to detect the migration and invasion ability of the 3 groups. Western-blot assay was used to detect the expressions ofα-Catenin,E-cad-herin,Vimentin and PBX3 protein in 3 groups. Antagonistic effect of PBX3 overexpression on miR-320d inhibition of EMT was detect-ed by western blot assay. The relationship between miR-320d and PBX3 was detected by dual luciferase assay.[Results]The expres-sion level of miR-320d in M320d group was significantly up-regulated,and the expression level of miR-320d was 808.25 ± 15.58 times higher than that of control group(P<0.05). The number of migrating cells in M320d group was 29.56 ± 0.59,which was signif-icantly lower than that of control group at 94.48 ± 1.02(P < 0.05). The number of invasive cells in M320d group was 7.33 ± 0.84, which was significantly lower than that of group control 86.28 ± 3.51(P < 0.05). Compared with control group ,the expression of α-Catenin and E-cadherin protein was significantly increased ,the expression of Vimentin protein was significantly decreased ,and the expression of PBX3 protein was significantly decreased. After PBX3 overexpression,the expression ofα-Catenin and E-cadherin protein were significantly decreased,the expression of Vimentin protein were significantly increased. Dual luciferase assay showed that PBX3 is a downstream target gene of miR-320d(P<0.05).[Conclusion]miR-320d may inhibit the expression of EMT related protein through the downstream target gene PBX3 and inhibit the epithelial mesenchymal transition function of endometrial carcinoma JEC cells.