The effect of amniotic membrane extract (AME) against hypoxia induced cytopathy in H9c2 cell line

Abstract

In this study, we aimed to appraise the protective impact of Amnion membrane protein extracts (AMPs) on H9c2 cells under hypoxia condition. Material and Method: AMPs were obtained from cezarian section and isolated by mechanical methods. Then, pretreated H9c2 cells with AMPs subjected to 24h hypoxia to induced injury. Cell viability, cell toxicity and H9c2 proliferation were evaluated by MTT, LDH, and Ki67 protein expression, respectively. Furthermore gene and protein expression of of nuclear factor erythroid 2-related factor 2 (Nrf2) and Peroxisome proliferator-activated receptor gamma (PPARγ), High mobility group box-1 (HMGB1) and Myeloid differentiation primary response 88 (MyD88) were detected by qPCR and western-blotting. The mitochondrial membrane potential (ΔΨm) was estimated by JC-1 staining. Moreover, cell apoptosis, Reactive oxygen species (ROS) production, Ca2+ alteration and cell senescence were measured by relevant technique. Results: Pre-treatment of AMPs resulted in significant induction in cell viability and inhibition of LDH with also increased in the Ki67 expression under hypoxia condition in H9c2 cells. Accordingly, these protective effects of AMPs were associated with reduction in apoptosis, ROS levels, Ca2+, and cell senescence while ΔΨm was increased. Furthermore, AMPs treatment resulted in down-regulation of HMGB1 and MyD88 levels and up-regulation of Nrf2 and PPAR-γ in H9c2 cell exposed to the hypoxia.