Inhibition of Mitofusin-2 Promotes Cardiac Fibroblast Activation via the PERK/ATF4 Pathway and Reactive Oxygen Species

Abstract

Mitofusin-2 (Mfn2) is a key outer mitochondrial membrane protein, which maintains normal mitochondrial dynamics and function. However, its role in cardiac fibroblast activation remains poorly understood. In the present study, a rat model of transverse aortic constriction (TAC) was established to observe the cardiac fibroblast activation in vivo. TGF-β1 treatment for 24 hours was used to induce cardiac fibroblast activation in vitro. As a result, the expression of Mfn2 decreased in the hypertrophic heart tissues and cardiac fibroblasts treated with TGF-β1. siMfn2 and adenovirus were applied to mediate Mfn2 gene silencing and overexpression in cardiac fibroblasts to elucidate the relationship between Mfn2 and cardiac fibroblast activation, as well as the possible underlying mechanisms. Knockdown of Mfn2 further promoted TGF-β1-induced cardiac fibroblast activation, while forced expression of Mfn2 attenuated this pathological reaction. The PERK/ATF4 pathway, one of the branches of endoplasmic reticulum (ER) stress, was identified to be involved in this process. Knockdown and overexpression of Mfn2 lead to aggravation or alleviation of the PERK/ATF4 pathway. Blocking this pathway by silencing ATF4 with siATF4 attenuated the pathological process. During the activation of cardiac fibroblasts, knockdown of Mfn2 also increased the production of reactive oxygen species (ROS), while ROS scavenger N-acetyl-l-cysteine (NAC) could attenuate the effect caused by knockdown of Mfn2. Our data suggested that inhibition of Mfn2 could promote cardiac fibroblast activation by activating the PERK/ATF4 signaling pathway and increasing the generation of ROS.

Inhibition of P2X7 Purinergic Receptor Ameliorates Cardiac Fibrosis by Suppressing NLRP3/IL-1β Pathway

“…Treatments. Neonatal rat cardiac fibroblasts were isolated from the hearts of decapitated Sprague-Dawley rats (age, 0-3 days) according to the methods described previously [19]. The CFs were grown in a culture flask with DMEM mixed with 10% FBS, and 100 U/ml of both streptomycin and penicillin.…”

“…Immunofluorescence Methodology. Immunofluorescence staining on cells was performed as described previously [19]. The primary antibody for incubation was anti-α-SMA (Abcam, 1 : 200) or P2X7R (Alomone Labs, 1 : 200).…”

Photodynamic therapy regulates fate of cancer stem cells through reactive oxygen species

“…PERK is found to be localized to MAMs and promotes endoplasmic reticulum-mitochondria coupling[ 162 ]. Second, Mfn2, a kind of skeleton in MAMs, can regulate the endoplasmic reticulum associated autophagy and apoptosis by downregulating the activity of PERK[ 163 , 164 ]. PACS2, another important component of MAM, also participates in the autophagy process.…”

Effects of Oxidative Stress on Protein Translation: Implications for Cardiovascular Diseases

“…Although ATF4 is viewed as stress-defense facilitator [112], several studies have reported the contribution of ATF4 to cardiovascular pathologies during oxidative stress. As such, in the rat model of transverse aortic constriction, the pathological stimuli-induced overexpression of ATF4 and increased ROS levels were found to contribute to cardiac fibroblast activation, the cellular reprogramming process that causes cardiac hypertrophy [113]. To detect ROS, the authors used 2 ,7 -dichlorofluorescein diacetate (H2DCFDA), the fluorogenic dye that measures a wide range of ROS, including hydroxyl, peroxyl, and other species within the cell [114]; thus, the particular ROS specie(s) modulating ATF4 s activity is unknown.…”