Yong Xia

Corrigendum: Editorial: Epigenetic and transcriptional networks underlying ventricular and atrial arrhythmias

Background: Sterile inflammation contributes to the pathogenesis of cardiac dysfunction caused by various conditions including pressure overload in hypertension. Mitochondrial DNA (mtDNA) released from damaged mitochondria has been implicated in cardiac inflammation. However, the upstream mechanisms governing mtDNA release and how mtDNA activates sterile inflammation in pressure-overloaded hearts remain largely unknown. Here, we investigated the role of inducible NO synthase (iNOS) on pressure overload-induced cytosolic accumulation of mtDNA and whether mtDNA activated inflammation through the cyclic GMP-AMP synthase (cGAS)-stimulator of interferon genes (STING) pathway.

Endothelial dysfunction often accompanies sepsis. We aimed to explore the role of PCSK9 in septic endothelial dysfunction. Sepsis was induced by lipopolysaccharide (LPS) treatment of human umbilical vein endothelial cells (HUVECs) in vitro and cecal ligation and puncture (CLP) surgery in mice in vivo. Evolocumab (EVC) and Pep 2–8, PCSK9 inhibitors, were subsequently used to determine the role of PCSK9 in sepsis-induced endothelial dysfunction in vitro and in vivo, respectively. In addition, the TLR4 agonist, Kdo2-Lipid A ammonium (KLA), was used to determine the related mechanism. Protein expression of eNOS, VE-cadherin, PCSK9, TLR4, MyD88, p-p65, p65, NLRP3, ASC, and caspase-1 p20 in mice aortas and HUVECs was measured by western blotting, while mRNA expression of TNFα, IL-1β, and IL-18 was determined by qRT-PCR. The level of inflammatory cytokines of mouse aortas was visualized by immunohistochemistry. Vasodilation of the aorta was detected by vascular reactivity experiments. The 96-h survival rate after CLP was assessed. Our findings showed that the expression of eNOS and VE-cadherin decreased, and PCSK9 expression increased, in septic HUVECs or mice. Inhibition of PCSK9 increased eNOS and VE-cadherin expression. Activation of the TLR4/MyD88/NF-κB and NLRP3 pathways may be responsible for PCSK9-induced endothelial dysfunction in sepsis. Vascular reactivity tests and survival studies showed that inhibition of PCSK9 could prevent the decrease in endothelium-dependent vasodilation function and improve the survival rates of septic mice. In summary, our results suggested that increased PCSK9 expression during sepsis activated the TLR4/MyD88/NF-κB and NLRP3 pathways to induce inflammation, which resulted in vascular endothelial dysfunction and decreased survival rates. Thus, inhibition of PCSK9 may be a potential clinical therapeutic target to improve vascular endothelial function in sepsis.

Editorial: Epigenetic and transcriptional networks underlying ventricular and atrial arrhythmias

Endothelial nitric oxide synthase (eNOS)‐derived nitric oxide (NO) plays a crucial role in maintaining vascular homeostasis. As a hallmark of eNOS activation, phosphorylation of eNOS at Ser1177 induced by activated protein kinase B (PKB/Akt) is pivotal for NO production. The complete activation of Akt requires its phosphorylation of both Thr308 and Ser473. However, which site plays the main role in regulating phosphorylation of eNOS Ser1177 is still controversial. The purpose of the present study is to explore the specific regulatory mechanism of phosphorylated Akt in eNOS activation. Inhibition of Akt Thr308 phosphorylation by a specific inhibitor or by siRNA in vitro led to a decrease in eNOS phosphorylation at Ser1177 and to lower NO concentration in the cell culture medium of HUVECs. However, inhibiting p‐Akt Ser473 had no effect on eNOS phosphorylation at Ser1177. Next, we administered mice with inhibitors to downregulate p‐Akt Ser473 or Thr308 activity. Along with the inhibition of p‐Akt Thr308, vascular p‐eNOS Ser1177 protein was simultaneously downregulated in parallel with a decrease in plasma NO concentration. Additionally, we cultured HUVECs at various temperature conditions (37, 22, and 4 °C). The results showed that p‐Akt Ser473 was gradually decreased in line with the reduction in temperature, accompanied by increased levels of p‐Akt Thr308 and p‐eNOS Ser1177. Taken together, our study indicates that the phosphorylation of Akt at Thr308, but not at Ser473, plays a more significant role in regulating p‐eNOS Ser1177 levels under physiological conditions.

Inducible nitric oxide synthase (iNOS) plays critical roles in the inflammatory response and host defense. Previous research on iNOS regulation mainly focused on its gene expression level, and much less is known about the regulation of iNOS function by N-glycosylation. In this study, we report for the first time that iNOS is N-glycosylated in vitro and in vivo. Mass spectrometry studies identified Asn(695) as an N-glycosylation site of murine iNOS. Mutating Asn(695) to Gln(695) yields an iNOS that exhibits greater enzyme activity. The essence of nitric oxide synthase catalytic reaction is electron transfer process, which involves a series of conformational changes, and the linker between the flavin mononucleotide-binding domain and the flavin adenine dinucleotide-binding domain plays vital roles in the conformational changes. Asn(695) is part of the linker, so we speculated that attachment of N-glycan to the Asn(695) residue might inhibit activity by disturbing electron transfer. Indeed, our NADPH consumption results demonstrated that N-glycosylated iNOS consumes NADPH more slowly. Taken together, our results indicate that iNOS is N-glycosylated at its Asn(695) residue and N-glycosylation of Asn(695) might suppress iNOS activity by disturbing electron transfer.