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MiR-29b-3p aggravates cardiac hypoxia/reoxygenation injury via targeting PTX3.

ABSTRACT: Our current research aimed to decipher the role and underlying mechanism with regard to miR-29b-3p involving in myocardial ischemia/reperfusion (I/R) injury. In the present study, cardiomyocyte H9c2 cell was used, and hypoxia/reoxygenation (H/R) model was established to mimic the myocardial I/R injury. The expressions of miR-29b-3p and pentraxin 3 (PTX3) were quantified deploying qRT-PCR and Western blot, respectively. The levels of LDH, TNF-α, IL-1β and IL-6 were detected to evaluate cardiomyocyte apoptosis and inflammatory response. Cardiomyocyte viability and apoptosis were examined employing CCK-8 assay and flow cytometry, respectively. Verification of the targeting relationship between miR-29b-3p and PTX3 was conducted using a dual-luciferase reporter gene assay. It was found that miR-29b-3p expression in H9c2 cells was up-regulated by H/R, and a remarkable down-regulation of PTX3 expression was demonstrated. MiR-29b-3p significantly promoted of release of inflammatory cytokines of H9c2 cells, and it also constrained the proliferation and promoted the apoptosis of H9c2 cells. Additionally, PTX3 was inhibited by miR-29b-3p at both mRNA and protein levels, and it was identified as a direct target of miR-29b-3p. PTX3 overexpression could reduce the inflammatory response, increase the viability of H9c2 cells, and inhibit apoptosis. Additionally, PTX3 counteracted the function of miR-29b-3p during the injury of H9c2 cells induced by H/R. In summary, miR-29b-3p was capable of aggravating the H/R injury of H9c2 cells by repressing the expression of PTX3.

SUBMITTER: He D

PROVIDER: S-EPMC7817735 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:Aimsthe adult zebrafish heart regenerates spontaneously after injury and has been used to study the mechanisms of cardiac repair. However, no zebrafish model is available that mimics ischemic injury in mammalian heart. We developed and characterized zebrafish cardiac injury induced by hypoxia/reoxygenation (H/R) and the regeneration that followed it.Methods and resultsadult zebrafish were kept either in hypoxic (H) or normoxic control (C) water for 15 min; thereafter fishes were returned to C water. Within 2-6 hours (h) after reoxygenation there was evidence of cardiac oxidative stress by dihydroethidium fluorescence and protein nitrosylation, as well as of inflammation. We used Tg(cmlc2:nucDsRed) transgenic zebrafish to identify myocardial cell nuclei. Cardiomyocyte apoptosis and necrosis were evidenced by TUNEL and Acridine Orange (AO) staining, respectively; 18 h after H/R, 9.9±2.6% of myocardial cell nuclei were TUNEL(+) and 15.0±2.5% were AO(+). At the 30-day (d) time point myocardial cell death was back to baseline (n?=?3 at each time point). We evaluated cardiomyocyte proliferation by Phospho Histone H3 (pHH3) or Proliferating Cell Nuclear Antigen (PCNA) expression. Cardiomyocyte proliferation was apparent 18-24 h after H/R, it achieved its peak 3-7d later, and was back to baseline at 30d. 7d after H/R 17.4±2.3% of all cardiomyocytes were pHH3(+) and 7.4±0.6% were PCNA(+) (n?=?3 at each time point). Cardiac function was assessed by 2D-echocardiography and Ventricular Diastolic and Systolic Areas were used to compute Fractional Area Change (FAC). FAC decreased from 29.3±2.0% in normoxia to 16.4±1.8% at 18 h after H/R; one month later ventricular function was back to baseline (n?=?12 at each time point).Conclusionszebrafish exposed to H/R exhibit evidence of cardiac oxidative stress and inflammation, myocardial cell death and proliferation. The initial decrease in ventricular function is followed by full recovery. This model more closely mimics reperfusion injury in mammals than other cardiac injury models.

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MiR-29b-3p aggravates cardiac hypoxia/reoxygenation injury via targeting PTX3.

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