Rubicon 通过 LC3 相关的吞噬作用介导的清除作用促进库普弗细胞的 M2 极化，从而改善肝移植。

Rubicon promotes the M2 polarization of Kupffer cells via LC3-associated phagocytosis-mediated clearance to improve liver transplantation.

机构信息

Department of Hepatobiliary Surgery, The Second Affiliated Hospital of Chongqing Medical University, Chongqing, China.

Department of Hepatobiliary Surgery, The First Affiliated Hospital of Chongqing Medical University, Chongqing, China.

出版信息

Cell Immunol. 2022 Aug;378:104556. doi: 10.1016/j.cellimm.2022.104556. Epub 2022 May 30.

DOI:10.1016/j.cellimm.2022.104556

PMID:35700602

Abstract

BACKGROUND

Acute rejection (AR) after liver transplantation (LT) is closely related to the survival of patients after surgery. Enhancement of the ability of Kupffer cells (KCs) to eliminate apoptotic cells can effectively alleviate AR.

METHODS

Rubicon lentivirus (LV) and Rubicon small interfering RNA (siRNA) were transfected into KCs extracted from the liver tissue of mice. Primary KCs were extracted and cocultured with zymosan and apoptotic T lymphocytes. The levels of CD86, CD163, IL-10, TNF-α, TGF-β, JAK1, STAT6, AKT1, mTOR and peroxisome proliferator-activated receptor-γ (PPARγ) were assessed via Western blotting (WB) and q-PCR. The levels of CD86 and CD163 were assessed via flow cytometry. mCherry-GFP-LC3 adenovirus (AV) was transfected into KCs. The recruitment of LC3II and the fusion of phagosomes and lysosomes were detected using immunofluorescence. Rubicon adeno-associated virus (AAV) was transfected into the liver tissue of mice via the portal vein, and models of immune tolerance (IT) and AR following LT were established. Pathological changes in the liver tissue were detected using HE staining. Apoptotic cells were assessed via TUNEL staining. The polarization state of KCs was detected via immunohistochemical staining.

RESULTS

Rubicon-mediated LC3-associated phagocytosis (LAP) promotes the ability of KCs to degrade and clear apoptotic T lymphocytes. Polyunsaturated fatty acids (PUFAs), the product of apoptotic T lymphocyte degradation, activate PPARγ, which further promotes the M2 polarization of KCs. Enhanced degradation mediated by Rubicon contributes to promoting the M2 polarization of KCs and a microenvironment supportive of IT.

CONCLUSIONS

Rubicon-mediated LAP promotes the clearance capability and M2 polarization of KCs via PUFA-dependent PPARγ activation to improve LT.

摘要

背景

肝移植（LT）后急性排斥（AR）与患者术后生存密切相关。增强枯否细胞（KCs）清除凋亡细胞的能力可以有效缓解 AR。

方法

Rubicon 慢病毒（LV）和 Rubicon 小干扰 RNA（siRNA）转染入从小鼠肝组织中提取的 KCs。提取原代 KCs 与酵母聚糖和凋亡 T 淋巴细胞共培养。通过 Western blot（WB）和 q-PCR 评估 CD86、CD163、IL-10、TNF-α、TGF-β、JAK1、STAT6、AKT1、mTOR 和过氧化物酶体增殖物激活受体-γ（PPARγ）的水平。通过流式细胞术评估 CD86 和 CD163 的水平。将 mCherry-GFP-LC3 腺病毒（AV）转染入 KCs。用免疫荧光法检测 LC3II 的募集和吞噬体与溶酶体的融合。经门静脉转染 Rubicon 腺相关病毒（AAV）至小鼠肝组织，建立 LT 后免疫耐受（IT）和 AR 模型。用 HE 染色检测肝组织的病理变化。用 TUNEL 染色评估凋亡细胞。用免疫组化染色检测 KCs 的极化状态。

结果

Rubicon 介导的 LC3 相关吞噬作用（LAP）增强了 KCs 降解和清除凋亡 T 淋巴细胞的能力。凋亡 T 淋巴细胞降解产物多不饱和脂肪酸（PUFA）激活 PPARγ，进一步促进 KCs 的 M2 极化。Rubicon 介导的增强降解有助于促进 KCs 的 M2 极化和有利于 IT 的微环境。