RESUMO
Chronic hypoxia is a key pathological change in patients with cyanotic congenital heart defect (CCHD). It has been demonstrated that enhanced myocardial unfolded protein response (UPR) increases the capacity to buffer endoplasmic reticulum (ER) stress and to avoid subsequent apoptosis caused by the hypoxia that underlies CCHD. The present study was performed to determine the regulatory role of microRNAs (miRNAs) in this cytoprotective UPR process. The results revealed that miR199a5p was markedly downregulated in the cardiac tissue of patients with CCHD and in human myocardial cells cultured in hypoxic conditions. The two major UPR modulators, 78 kDa glucoseregulated protein (GRP78) and activating transcription factor 6 (ATF6), were potential target genes of miR199a5p in CCHD myocardial specimens. In addition, the miR199a5p mimic and inhibitor were evidently able to change GRP78 and ATF6 gene expression and ER stressassociated apoptosis in hypoxiatreated cardiomyocytes. The interaction between miR199a5p and the ATF6 and GRP78 3'UTR binding sites in myocardial cells was also confirmed by luciferase assay. Thus, it is concluded that myocardial downregulation of miR199a5p favors the UPR against hypoxiainduced ER stress in CCHD, which contributes to myocardial protection.
Assuntos
Cianose/congênito , Cianose/genética , Regulação para Baixo/genética , Estresse do Retículo Endoplasmático/genética , Cardiopatias/congênito , Cardiopatias/genética , MicroRNAs/genética , Miócitos Cardíacos/patologia , Fator 6 Ativador da Transcrição/genética , Fator 6 Ativador da Transcrição/metabolismo , Adolescente , Apoptose/genética , Hipóxia Celular/genética , Células Cultivadas , Criança , Pré-Escolar , Chaperona BiP do Retículo Endoplasmático , Proteínas de Choque Térmico/genética , Proteínas de Choque Térmico/metabolismo , Humanos , MicroRNAs/metabolismo , Miocárdio/metabolismo , Miocárdio/patologia , Miócitos Cardíacos/metabolismo , Resposta a Proteínas não Dobradas/genéticaRESUMO
MicroRNAs (miRs) regulate a number of physiological and pathological processes, including myocardial chronic hypoxia. Previous studies revealed that the expression of miR-146b is increased in vitro and in vivo following the induction of hypoxia. In the present study, the role of miR146b in hypoxic cardiomyocytes, and the mechanisms underlying its activity, were investigated. The expression of miR146b was measured in tissue samples from patients with congenital heart disease by reverse transcriptionquantitative polymerase chain reaction. The rat H9c2 cardiomyocyte cell line was transfected with an miR146b inhibitor or the experimental controls, and the cells were maintained under hypoxic conditions for 72 h. The expression of miR146b increased following the induction of hypoxia. Transfection with the miR146b inhibitor enhanced the release of lactate dehydrogenase and increased hypoxiainduced apoptosis, as determined by terminal deoxynucleotidyl transferase dUTP nickend labeling, Hoechst 33258 staining, JC1 assay (measuring mitochondrial membrane permeability) and annexin V/propidium iodide analysis. A decreased expression of Bcl2 was observed, whereas the expression levels of cleavedcaspase 3 and Bax were increased. Western blot analysis and a dual luciferase reporter assay confirmed that ribonuclease L is a direct target of miR146b. Furthermore, inhibition of miR-146b increased the activation of nuclear factor-κB and signal transducer and activator of transcription 3. In conclusion, the inhibition of miR146b may increase hypoxia-induced cardiomyocyte apoptosis.
Assuntos
Apoptose , Hipóxia/genética , MicroRNAs/genética , Miócitos Cardíacos/patologia , Animais , Hipóxia Celular , Linhagem Celular , Células Cultivadas , Cardiopatias Congênitas/complicações , Cardiopatias Congênitas/genética , Cardiopatias Congênitas/patologia , Humanos , Hipóxia/complicações , Hipóxia/patologia , Lactente , MicroRNAs/antagonistas & inibidores , Miócitos Cardíacos/citologia , Miócitos Cardíacos/metabolismo , Ratos , Transfecção , Regulação para CimaRESUMO
AIM: To probe into the clinical significance of cellular immune function in patients with viral myocarditis. METHODS: Choose our hospital in January 2008-December 2009 admitted during the 75 patients of viral myocarditis as the experimental group, 67 healthy patients served as control groups, respectively, and hospital admission within 24 h after the venous blood, before and after treatment T cell subsets, natural killer cells (NK) activity and tumor necrosis factor (TNF) were detected and analyzed. RESULTS: Compared with admission, 24 h after admission test in peripheral blood T lymphocyte subsets CD3, CD4, CD8 and CD4/CD8 were improved and CD3, CD4 and CD4/CD8 were statistically significant differences (P<0.05), but still slightly lower in the control group, while the difference was not statistically significant. At the same time, compared with admission, 24 h after admission of natural killer cells in the experimental group (NK) and tumor necrosis factor were improved, the differences were statistically significant (P<0.05), but still slightly lower than the control group, but the difference was not statistically significant. CONCLUSION: More in-depth study of viral myocarditis in patients with immune injury mechanism will help to better guide the treatment, and has important clinical significance.