Liao Ke, Niu Fang, Hu Guoku, Guo Ming-Lei, Sil Susmita, Buch Shilpa
Department of Pharmacology & Experimental Neuroscience, University of Nebraska Medical Center , Omaha, NE, USA.
Tissue Barriers. 2020 Apr 2;8(2):1748983. doi: 10.1080/21688370.2020.1748983. Epub 2020 Apr 16.
The blood-brain barrier (BBB) is a tight barrier that is critical for preventing the entry of pathogens and small molecules into the brain. HIV protein Tat (Tat) is known to disrupt the tight junctions of the BBB. Autophagy is an intracellular process that involves degradation and recycling of damaged organelles to the lysosome and has recently been implicated in the BBB disruption. The role of autophagy in Tat-mediated BBB disruption, however, remains elusive. Herein we hypothesized that Tat induces endothelial autophagy resulting in decreased expression of the tight junction protein ZO-1 leading to breach of the BBB. In this study, we demonstrated that exposure of human brain microvessel endothelial cells (HBMECs) to Tat resulted in induction of autophagy in a dose- and time-dependent manner, with upregulation of BECN1/Beclin 1, ATG5 and MAP1LC3B proteins and a concomitant downregulation of the tight junction protein ZO-1 ultimately leading to increased endothelial cell monolayer paracellular permeability in an BBB model. Pharmacological and genetic inhibition of autophagy resulted in the abrogation of Tat-mediated induction of MAP1LC3B with a concomitant restoration of tight junction proteins, thereby underscoring the role of autophagy in Tat-mediated breach of the BBB. Additionally, our data also demonstrated that Tat-mediated induction of autophagy involved PELI1/K63-linked ubiquitination of BECN1. Increased autophagy and decreased ZO-1 was further recapitulated in microvessels isolated from the brains of HIV Tg26 mice as well as the frontal cortex of HIV-infected autopsied brains. Overall, our findings identify autophagy as an important mechanism underlying Tat-mediated disruption of the BBB.
血脑屏障（BBB）是一种紧密屏障，对于防止病原体和小分子进入大脑至关重要。已知HIV蛋白Tat会破坏血脑屏障的紧密连接。自噬是一种细胞内过程，涉及将受损细胞器降解并循环至溶酶体，最近被认为与血脑屏障破坏有关。然而，自噬在Tat介导的血脑屏障破坏中的作用仍不清楚。在此，我们假设Tat诱导内皮细胞自噬，导致紧密连接蛋白ZO-1表达降低，从而导致血脑屏障破坏。在本研究中，我们证明，将人脑微血管内皮细胞（HBMECs）暴露于Tat会以剂量和时间依赖性方式诱导自噬，BECN1/Beclin 1、ATG5和MAP1LC3B蛋白上调，同时紧密连接蛋白ZO-1下调，最终导致血脑屏障模型中内皮细胞单层的细胞旁通透性增加。自噬的药理学和基因抑制导致Tat介导的MAP1LC3B诱导被废除，同时紧密连接蛋白得以恢复，从而强调了自噬在Tat介导的血脑屏障破坏中的作用。此外，我们的数据还表明，Tat介导的自噬诱导涉及PELI1/K63连接的BECN1泛素化。在从HIV Tg26小鼠大脑分离的微血管以及HIV感染尸检大脑的额叶皮质中，自噬增加和ZO-1减少的情况进一步得到证实。总体而言，我们的研究结果确定自噬是Tat介导的血脑屏障破坏的重要潜在机制。
