内皮素-1 受体 B 型（ETBR）过表达与细胞间黏附分子-1（ICAM-1）下调相关，导致实验性自身免疫性心肌炎中的炎症减轻。

ET-1 receptor type B (ETBR) overexpression associated with ICAM-1 downregulation leads to inflammatory attenuation in experimental autoimmune myocarditis.

机构信息

Cardiology, The First Affiliated Hospital of Nanchang University, Nanchang, China.

Cardiology, Shenzhen Third People's Hospital, Shenzhen, Guangdong, China.

出版信息

PeerJ. 2023 Oct 24;11:e16320. doi: 10.7717/peerj.16320. eCollection 2023.

DOI:10.7717/peerj.16320

PMID:37901475

原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC10607261/

Abstract

BACKGROUND

An experimental autoimmune myocarditis rat model was established by subcutaneous injection of porcine myocardial myosin (PCM). The effect of ET-1 receptor type B (ETBR) overexpression on autoimmune myocarditis was observed tail vein injection of ETBR overexpression lentivirus in rats. We further investigated the mechanisms involved in the regulation of autoimmune myocarditis by ETBR overexpression.

METHODS

Six rats were randomly selected from 24 male Lewis rats as the NC group, and the remaining 18 rats were injected with PCM on Day 0 and Day 7, to establish the experimental autoimmune myocarditis (EAM) rat model. The 18 rats initially immunized were randomly divided into three groups: the EAM group, ETBR-oe group, and GFP group. On Day 21 after the initial immunization of rats, cardiac echocardiography and serum brain natriuretic peptide (BNP) analysis were performed to evaluate cardiac function, myocardial tissue HE staining was performed to assess myocardial tissue inflammatory infiltration and the myocarditis score, and mRNA expression of IFN-γ, IL-12, and IL-17 was detected by qRT-PCR. Subsequently, immunohistochemical analysis was performed to detect the localization and expression of the ETBR and ICAM-1 proteins, and the expression of ETBR and ICAM-1 was verified by qRT-PCR and western blotting methods.

RESULTS

On Day 21 after initial immunization, left ventricular end-diastolic diameter (LVEDd), left ventricular end-systolic diameter (LVEDs), and serum BNP concentrations increased in the hearts of rats in the EAM group compared with the NC group ( < 0.01), and ejection fraction (EF) and fractional shortening (FS) decreased compared with those of the normal control (NC) group ( < 0.01). LVEDd, LVEDs, and serum BNP concentrations decreased in the ETBR-oe group compared with the EAM group, while EF and FS increased significantly ( < 0.01). HE staining showed that a large number of inflammatory cell infiltrates, mainly lymphocytes, were observed in the EAM group, and the myocarditis score was significantly higher than that of the NC group ( < 0.01). Compared with that of the EAM group, myocardial tissue inflammatory cell infiltration was significantly reduced in the ETBR-oe group, and the myocarditis scores were significantly lower ( < 0.01). The mRNAs of the inflammatory factors IFN-γ, IL-12 and IL-17 in myocardial tissue of rats in the EAM group exhibited elevated levels compared with those of the NC group ( < 0.01) while the mRNAs of IFN-γ, IL-12 and IL-17 were significantly decreased in the ETBR-oe group compared with the EAM group ( < 0.01). Immunohistochemistry showed that the staining depth of ETBR protein in myocardial tissue was greater in the EAM group than in the NC group, and significantly greater in the ETBR-oe group than in the EAM group, while the staining depth of ICAM-1 was significantly greater in the EAM group than in the NC group, and significantly lower in the ETBR-oe group than in the EAM group. The ICAM-1 expression level was significantly higher in the EAM group than in the NC group ( < 0.01), and was significantly lower in the ETBR-oe groupthan in the EAM group ( < 0.01).

摘要

背景

通过皮下注射猪心肌肌球蛋白（PCM）建立实验性自身免疫性心肌炎大鼠模型。通过尾静脉注射 ETBR 过表达慢病毒观察 ETBR 过表达对自身免疫性心肌炎的影响。我们进一步研究了 ETBR 过表达调节自身免疫性心肌炎的机制。

方法

从 24 只雄性 Lewis 大鼠中随机选择 6 只作为 NC 组，其余 18 只大鼠于第 0 天和第 7 天注射 PCM，建立实验性自身免疫性心肌炎（EAM）大鼠模型。最初免疫的 18 只大鼠随机分为三组：EAM 组、ETBR-oe 组和 GFP 组。在大鼠初次免疫后第 21 天，进行心脏超声心动图和血清脑钠肽（BNP）分析，评估心功能；心肌组织 HE 染色评估心肌组织炎症浸润和心肌炎评分；qRT-PCR 检测 IFN-γ、IL-12 和 IL-17 的 mRNA 表达。随后，进行免疫组织化学分析，检测 ETBR 和 ICAM-1 蛋白的定位和表达，并通过 qRT-PCR 和 Western blot 方法验证 ETBR 和 ICAM-1 的表达。

结果

初次免疫后第 21 天，EAM 组大鼠左心室舒张末期直径（LVEDd）、左心室收缩末期直径（LVEDs）和血清 BNP 浓度均高于 NC 组（ < 0.01），射血分数（EF）和短轴缩短率（FS）均低于 NC 组（ < 0.01）。与 EAM 组相比，ETBR-oe 组 LVEDd、LVEDs 和血清 BNP 浓度降低，EF 和 FS 显著升高（ < 0.01）。HE 染色显示，EAM 组大量炎症细胞浸润，主要为淋巴细胞，心肌炎评分明显高于 NC 组（ < 0.01）。与 EAM 组相比，ETBR-oe 组心肌组织炎症细胞浸润明显减少，心肌炎评分明显降低（ < 0.01）。EAM 组大鼠心肌组织中炎症因子 IFN-γ、IL-12 和 IL-17 的 mRNA 水平均高于 NC 组（ < 0.01），而 ETBR-oe 组的 IFN-γ、IL-12 和 IL-17 的 mRNA 水平均低于 EAM 组（ < 0.01）。免疫组织化学显示，EAM 组心肌组织中 ETBR 蛋白的染色深度大于 NC 组，ETBR-oe 组的染色深度大于 EAM 组，而 ICAM-1 蛋白的染色深度在 EAM 组大于 NC 组，ETBR-oe 组的染色深度小于 EAM 组。EAM 组 ICAM-1 表达水平明显高于 NC 组（ < 0.01），ETBR-oe 组明显低于 EAM 组（ < 0.01）。