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OBJECTIVE@#To study the effect of silencing LncRNA SNHG7 on hypoxia/reoxygenation (H/R)-induced cardiomyocyte injury and its targeted regulation on miR-181b-5p.@*METHODS@#Rat cardiomyocytes H9c2 were cultured in vitro and randomly divided into control group, H/R group, H/R + si-NC group, H/R + si-SNHG7 group, H/R + si-SNHG7 + anti-miR-NC group and H/R + si-SNHG7 + anti-miR-181b-5p group. The content of lactate dehydrogenase (LDH), malondialedhyde (MDA) and the activity of superoxide dismutase (SOD) were detected. Flow cytometry was carried out to detect the rate of apoptosis. qRT-PCR was used to detect the expression of SNHG7 and miR-181b-5p. Dual luciferase report experiment was used to verify the targeting relationship between SNHG7 and miR-181b-5p. Western blotting was used to detect the expression of Bax and Bcl-2.@*RESULTS@#Compared with the control group, the H/R group showed significantly increased SNHG7 expression in cardiomyocytes, reduced miR-181b-5p expression, higher levels of LDH and MDA, reduced activity of SOD, increased cell apoptosis rate, higher level of Bax protein, and reduced level of Bcl-2 protein (all P< 0.05). Compared with the H/R and H/R + si-NC groups, the H/R + si-SNHG7 group had significantly reduced level of LDH and MDA, increased activity of SOD, reduced apoptosis rate, reduced level of Bax protein, increased level of Bcl-2 protein (all P< 0.05). The dual luciferase report experiment confirmed that SNHG7 could target miR-181b-5p. Interference with the expression of miR-181b-5p could reduce the effect of silencing SNHG7 on H/R-induced cardiomyocyte oxidative stress and apoptosis.@*CONCLUSION@#Silencing SNHG7 may inhibit H/R-induced cardiomyocyte oxidative stress and apoptosis by up-regulating the expression of miR-181b-5p, thereby exerting a protective effect on cardiomyocytes.
Subject(s)
Animals , Rats , Apoptosis , Hypoxia , MicroRNAs/genetics , Myocardial Reperfusion Injury , Myocytes, Cardiac , RNA, Long Noncoding/geneticsABSTRACT
Objective To establish an efficient method of genotyping for Leprdb/ +mouse offsprings by TaqMan probe quantitative fluorescence PCR. Methods Genome DNA was extracted from tails of 228 Leprdb/ +mouse offsprings. PCR primers and TaqMan probes were designed according to the mutation sites of Lepr gene(rs1801133). Real time PCR assay was applied and SNP loci were typed with SDS software. The genotyping of 2-month old Leprdb/dbmice was validated by the phenotype and Hardy-Weinberg equilibrium test was performed. Results 228 samples were detected by the established TaqMan fluorescence quantitative PCR assay. 64 mice were of GG genotype, with a genotype frequency of 0.1929. 123 mice were of GT genotype, with a genotype frequency of 0.5395. 41 mice were of TT genotype, with a genotype frequency of 0.2807. Compared with the phenotype typing,the sensitivity of the TaqMan fluorescence quantitative PCR was 97.56% and the specificity was 99.47%. Conclusions TaqMan probe quantitative fluorescence PCR assay is a simple and efficient method,and can be used to detect the genotype of Leprdb/ +mouse offsprings.
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Objective To study the expression of lipoic acid synthase(LIAS)in the liver and kidney of Leprdb/db mice with deficient leptin receptor. Methods Eight 10-week old male Leprdb/ +mice and Leprdb/dbmice were included in this study. The body weight of rats in the two groups was measured. Fasting blood glucose(FPG)was measured with blood glucose test strips for all mice after fasting for 8 hours. Blood samples were obtained from the abdominal aorta and the animals were sacrificed. The liver and kidney were weighed. The right lobe of liver and the left kidney samples were fixed in 4% paraformaldehyde for pathological examination. Serum samples were separated and the sereum contents of CHO, TG,HDL and LDL were detected. The mitochondria of liver and kidney tissues were extracted with a mitochondrial isolation kit, and the protein was extracted. The expression of LIAS protein was detected by western blot. Results Histopathological observation showed that the liver and kidney tissues of Leprdb/ +mice have intact and clear structure. But the liver tissue of Leprdb/dbmice showed fatty degeneration, the kidney tissue showed glomerular hypertrophy, basement membrane thickening, mesangial area widened, including mesangial cells and mesangial matrix increased. The GLU,CHO,TG,LDL and AST of Leprdb/dbmice were significantly increased compared with those of Leprdb/ +mice(P<0.05). Compared with Leprdb/ +mice,the LIAS protein expression was significantly increased in the liver and kidney mitochondria of Leprdb/dbmice(P<0.05). Conclusions There is impaired glucose and lipid metabolism in the Leprdb/dbmice which has defect leptin receptor,and the expression of LIAS protein in liver and kidney of the Leprdb/dbmice is higher than that of Leprdb/ +mice.
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Objective To study the adverse effect of omethoate on DNA in mouse sperm cells in vivo. Single cell gel electrophoresiss (SCGE) or comet assay was used to observe the damage of DNA. Methods 40 male kunming mice were randomly divided into 4 groups and were treated with omethoate by gavage at 0 mg/kg, 0.5 mg/kg, 1 mg/kg and 2 mg/kg respectively, once a day for 35 consecutive days. Results The movement and density of sperms decreased as the dose of omethoate increased, the length of comet tails in group of 1 mg/kg and group of 2 mg/kg was much longer than that in the control, the other indexes of comet assay also showed significant DNA damage. Conclusion Omethoate exposure can induce DNA damage in the sperm cells of male mice.
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Objective To study the effect of omethoate on the activity of characteristic enzymes in mouse testicles and the antagonism of teapolyphenol. Methods 80 Kunming male mice were divided into 8 groups, omethoate was given by gavage: the control group given 0.9% NaCl solution, a group given teapolyphenol of 180 mg/kg bw, three groups given omethoate of 1, 2 and 4 mg/kg bw respectively, three groups given teapolyphenol of 180 mg/kg bw and two hours later followed by omethoate of 1, 2 and 4 mg/kg bw respectively. The gavages were conducted for 6 consecutive days. The activities of the characteristic enzymes (AKP, ACP, LDH and LDH-x) in mouse testicles were measured by biochemistry method and variance analysis was done with the SPSS 11.0 software. Results As omethoate dose increased, the body weight and the testicle weight decreased obviously, the activities of AKP, ACP, LDH in mouse testicles significantly increased compared with the control. Given teapolyphenol 2 hours before given omethoate, the significant changes as above did not emerged. Conclusion Teapolyphenol has an antagonism against the adverse effects of omethoate on the activity of characteristic enzymes in mouse testicles.