SIRT3 sulfhydration using hydrogen sulfide inhibited angiotensin II-induced atrial fibrosis and vulnerability to atrial fibrillation via suppression of the TGF-β1/smad2/3 signalling pathway.

Atrial fibrosis is associated with the occurrence of atrial fibrillation (AF) and regulated by the transforming growth factor-β1 (TGF-β1)/Smad2/3 signalling pathway. Unfortunately, the mechanisms of regulation of TGF-β1/Smad2/3-induced atrial fibrosis and vulnerability to AF remain still unknown. Previous studies have shown that sirtuin3 (SIRT3) sulfhydration has strong anti-fibrotic effects. We hypothesised that SIRT3 sulfhydration inhibits angiotensin II (Ang-II)-induced atrial fibrosis via blocking the TGF-β1/Smad2/3 signalling pathway. In this study, we found that SIRT3 expression was decreased in the left atrium of patients with AF compared to that in those with sinus rhythm (SR). In vitro, SIRT3 knockdown by small interfering RNA significantly expanded Ang-II-induced atrial fibrosis and TGF-β1/Smad2/3 signalling pathway activation, whereas supplementation with Sodium Hydrosulfide (NaHS, exogenous hydrogen sulfide donor and sulfhydration agonist) and SIRT3 overexpression using adenovirus ameliorated Ang-II-induced atrial fibrosis. Moreover, we observed suppression of the TGF-β1/Smad2/3 pathway when Ang-II was combined with NaHS treatment, and the effect of this co-treatment was consistent with that of Ang-II combined with LY3200882 (Smad pathway inhibitor) on reducing atrial fibroblast proliferation and cell migration in vitro. Supplementation with dithiothreitol (DTT, a sulfhydration inhibitor) and adenovirus SIRT3 shRNA blocked the ameliorating effect of NaHS and AngII co-treatment on atrial fibrosis in vitro. Finally, continued treatment with NaHS in rats ameliorated atrial fibrosis and remodelling, and further improved AF vulnerability induced by Ang-II, which was reversed by DTT and adenovirus SIRT3 shRNA, suggesting that SIRT3 sulfhydration might be a potential therapeutic target in atrial fibrosis and AF.

Hu HJ ，Wang XH ，Zhang ZZ ，Ou Y ，Ning ZH ，Yang JY ，Huang H ，Tang HF ，Jiang ZS ... -  《-》

MFGE8 attenuates Ang-II-induced atrial fibrosis and vulnerability to atrial fibrillation through inhibition of TGF-β1/Smad2/3 pathway.

Atrial fibrillation (AF) is characterized by potentiated growth of atrial fibroblasts and excessive deposition of the extracellular matrix. Atrial fibrosis has emerged as a hallmark of atrial structural remodeling linked to AF. Nonetheless, the specific mechanism underlying the progression of atrial fibrosis to AF is still largely unknown. MFGE8 (milk fat globule-EGF factor 8) is a soluble glycoprotein associated with many human diseases. Recently, a number of studies revealed that MFGE8 plays a crucial role in heart disease. Yet, MFGE8 regulation and function in the process of atrial fibrosis and vulnerability to AF remain unexplored. In this study, we found that the expression of MFGE8 was downregulated in the atriums of patients with AF compared with individuals without AF. In addition, the expression of MFGE8 was lower in atriums of angiotensin II (Ang-II)-stimulated rats as compared with the sham group. In vitro, silencing of MFGE8 by small interfering RNA significantly increased Ang-II-induced atrial fibrosis, whereas administration of recombinant human MFGE8 (rhMFGE8) attenuated the atrial fibrosis. Moreover, we found that the activated TGF-β1/Smad2/3 pathway after Ang-II treatment was significantly potentiated by the MFGE8 knockdown but inhibited by rhMFGE8 in vitro. Inhibition of integrin β3 which is the receptor for MFGE8, suppressed the TGF-β1/Smad2/3 activating effects of the MFGE8 knockdown in Ang-II-treated rat atrial fibroblasts. Finally, we administered rhMFGE8 to rats; it attenuated atrial fibrosis and remodeling and further reduced AF vulnerability induced by Ang-II, indicating that MFGE8 might have the potential both as a novel biomarker and as a therapeutic target in atrial fibrosis and AF.

Ge Z ，Chen Y ，Wang B ，Zhang X ，Yan Y ，Zhou L ，Zhang Y ，Xie Y ... -  《-》

Upregulation of Transient Receptor Potential Canonical Type 3 Channel via AT1R/TGF-β1/Smad2/3 Induces Atrial Fibrosis in Aging and Spontaneously Hypertensive Rats.

Fibroblast proliferation and migration are central in atrial fibrillation (AF) promoting structure remodeling, which is strongly associated with aging and hypertension. Transient receptor potential canonical-3 channel (TRPC3) is a key mediator of cardiac fibrosis and the pathogenesis of AF. Here, we have observed the increased TRPC3 expression that induced atrial fibrosis which possibly is either mediated by the aging process or related to hypertensive progression. In this study, we measured the pathological structure remodeling by H&E staining, Masson staining, and transmission electron microscope (TEM). The protein expression levels of fibrotic biomarkers and TRPC3 were measured by Western blotting with atrial tissues from normotensive Wistar Kyoto rats (WKY 4m-o (4 months old)), old WKY (WKY 24m-o (24 months old)), spontaneously hypertensive rat (SHR 4m-o (4 months old)), and old SHR (SHR 24m-o (24 months old)). To illuminate the molecular mechanism of TRPC3 in atrial fibrosis of aging rats and SHR, we detected the inhibited role of TRPC3 selective blocker ethyl-1-(4-(2,3,3-trichloroacrylamide) phenyl)-5-(trifluoromethyl)-1H-pyrazole-4-carboxylate,pyrazole-3 (Pyr3) on angiotensin II (Ang II) induced fibrosis in neonatal rat atrial fibroblasts. The pathological examination showed that the extracellular matrix (ECM) and collagen fibrils were markedly increased in atrial tissues from aged and hypertensive rats. The protein expressions of fibrotic biomarkers (collagen I, collagen III, and transforming growth factor-β1 (TGF-β1)) were significantly upregulated in atrial tissues from the WKY 24m-o group, SHR 4m-o group, and SHR 24m-o group compared with the WKY 4m-o group. Meanwhile, the expression level of TRPC3 was significantly upregulated in WKY 24m-o and SHR 4m-o atrial tissues compared to WKY 4m-o rats. In isolated and cultured neonatal rat atrial fibroblasts, Ang II induced the atrial fibroblast migration and proliferation and upregulated the expression levels of TRPC3 and fibrotic biomarkers. TRPC3 selected blocker Pyr3 attenuated the migration and proliferation in neonatal rat atrial fibroblasts. Furthermore, Pyr3 significantly alleviated Ang II-induced upregulation of TRPC3, collagen I, collagen III, and TGF-β1 through the molecular mechanism of the TGF-β/Smad2/3 signaling pathway. Similarly, knocking down TRPC3 using short hairpin RNA (shTRPC3) also attenuated Ang II-induced upregulation of TGF-β1. Pyr3 preconditioning decreased Ang II-induced intracellular Ca2+ transient amplitude elevation. Furthermore, AT1 receptor was involved in Ang II-induced TRPC3 upregulation. Hence, upregulation of TRPC3 in aging and hypertension is involved in an atrial fibrosis process. Inhibition of TRPC3 contributes to reverse Ang II-induced fibrosis. TRPC3 may be a potential therapeutic target for preventing fibrosis in aging and hypertension.

He R ，Zhang J ，Luo D ，Yu Y ，Chen T ，Yang Y ，Yu F ，Li M ... -  《-》

Angiotensin II upregulates Kv1.5 expression through ROS-dependent transforming growth factor-beta1 and extracellular signal-regulated kinase 1/2 signalings in neonatal rat atrial myocytes.

Kv1.5 potassium channel represents a promising target for atrial fibrillation (AF) therapy. During AF, the renin-angiotensin system is markedly activated. Recent evidence indicates that angiotensin II (Ang II) can upregulate Kv1.5 channel, but the mechanism remains unknown. In this study, we report that Ang II-mediated transforming growth factor-beta1 (TGF-β1)/Smad2/3 and extracellular signal-regulated kinase (ERK) 1/2 signalings are involved in atrial Kv1.5 expression. In neonatal rat atrial myocytes, quantitative PCR and Western blotting revealed that Ang II upregulated TGF-β1, synapse-associated protein 97 (SAP97) and Kv1.5 expression in a time- and concentration-dependent manner. The Ang II-induced upregulation of Kv1.5, SAP97 and phosphorylated Smad2/3 (P-Smad2/3) were reversed by the Ang II type 1 (AT1) receptor antagonist losartan, an anti-TGF-β1 antibody and the ERK 1/2 inhibitor PD98059 but not by the AT2 receptor antagonist PD123319. mRNA knockdown of either Smad2 or Smad3 blocked Ang II-induced expression of Kv1.5 and SAP97. These data suggest that AT1 receptor/TGF-β1/P-Smad2/3 and ERK 1/2 signalings are involved in Ang II-induced Kv1.5 and SAP97 expression. Flow cytometry and Western blotting revealed that losartan and the anti-TGF-β1 antibody diminished Ang II-induced reactive oxygen species (ROS) generation and that the antioxidants diphenyleneiodonium and N-acetyl cysteine inhibited Ang II-induced expression of P-Smad2/3, phosphorylated ERK (P-ERK) 1/2, Kv1.5, SAP97, suggesting that ROS participate in Kv1.5 and SAP97 regulation by modulating Ang II-induced P-Smad2/3 and P-ERK 1/2 expression. In conclusion, we demonstrate that ROS-dependent Ang II/AT1 receptor/TGF-β1/P-Smad2/3 and Ang II/ERK 1/2 signalings are involved in atrial Kv1.5 and SAP97 expression. Antioxidants would be beneficial for AF treatment through inhibiting atrial Kv1.5 expression.

Lu G ，Xu S ，Peng L ，Huang Z ，Wang Y ，Gao X ... -  《-》

Neferine mitigates angiotensin II-induced atrial fibrillation and fibrosis via upregulation of Nrf2/HO-1 and inhibition of TGF-β/p-Smad2/3 pathways.

Jiang XX ，Zhang R ，Wang HS 《Aging-US》