Alcohol and HIV-Derived Hepatocyte Apoptotic Bodies Induce Hepatic Stellate Cell Activation

Abstract

Recently, we found that both HIV and acetaldehyde, an alcohol metabolite, induce hepatocyte apoptosis, resulting in the release of large extracellular vesicles called apoptotic bodies (ABs). The engulfment of these hepatocyte ABs by hepatic stellate cells (HSC) leads to their profibrotic activation. This study aims to establish the mechanisms of HSC activation after engulfment of ABs from acetaldehyde and HIV-exposed hepatocytes (ABAGS+HIV). In vitro experiments were performed on Huh7.5-CYP (RLW) cells to generate hepatocyte ABs and LX2 cells were used as HSC. To generate ABs, RLW cells were pretreated for 24 h with acetaldehyde, then exposed overnight to HIV1ADA and to acetaldehyde for 96 h. Thereafter, ABs were isolated from cell suspension by a differential centrifugation method and incubated with LX2 cells (3:1 ratio) for profibrotic genes and protein analyses. We found that HSC internalized ABs via the tyrosine kinase receptor, Axl. While the HIV gag RNA/HIV proteins accumulated in ABs elicited no productive infection in LX2 and immune cells, they triggered ROS and IL6 generation, which, in turn, activated profibrotic genes via the JNK-ERK1/2 and JAK-STAT3 pathways. Similarly, ongoing profibrotic activation was observed in immunodeficient NSG mice fed ethanol and injected with HIV-derived RLW ABs. We conclude that HSC activation by hepatocyte ABAGS+HIV engulfment is mediated by ROS-dependent JNK-ERK1/2 and IL6 triggering of JAK-STAT3 pathways. This can partially explain the mechanisms of liver fibrosis development frequently observed among alcohol abusing PLWH.

Hepatocyte-Specific Triggering of Hepatic Stellate Cell Profibrotic Activation by Apoptotic Bodies: The Role of Hepatoma-Derived Growth Factor, HIV, and Ethanol

“…We previously observed robust hepatocyte (RLW) apoptosis from the combination of the acetaldehyde-generating system (AGS) and HIV [ 9 ], followed by the generation of HIV and MDA-expressing ABs (AB AGS+HIV(RLW) ) to induce profibrotic activation in HSC (LX2 cells) [ 11 ]. Here, LX2 cells were exposed to hepatic AB AGS+HIV(RLW).…”

“…We demonstrated the highest hepatocyte apoptosis induced by the combined exposure to acetaldehyde and HIV [ 9 , 10 ], which exceeds the effects of single exposures. The apoptotic bodies (ABs) derived from these cells express enormous HIV proteins, HIV RNA, and malondialdehyde (MDA), capable of inducing hepatic stellate cell (HSC) profibrotic activation after ABs internalization [ 11 ]. While HSC profibrotic activation and the consequent extracellular matrix (ECM) deposition are homeostatic responses to hepatocyte depletion [ 12 ], acetaldehyde- and HIV-induced massive hepatocyte depletion engenders excessive continuous ECM deposition, which is a pivotal event in the incidence of hepatic fibrosis [ 13 ].…”

“…A brief review of this crosstalk must be established as we commence hepatocyte apoptosis. Unlike other studies, which utilized the UV method to generate hepatocyte ABs [ 15 , 16 ], in the current study, we generated ABs from acetaldehyde and HIV-treated hepatocyte-like cells (RLW cells) [ 11 ]. While we did not seek to discredit the UV method of ABs generation, we preferred to mimic natural exposures for ABs generation to avoid the possibility that in HIV-infected hepatocytes, UV may affect ABs cargo [ 17 ].…”

“…As we demonstrated before, there is an internalization of the phosphatidylserine-expressing hepatocyte-derived ABs by AXL, a phosphatidylserine recognition receptor expressed on HSC leading to their profibrotic activation [ 11 ]. However, a profibrotic activation may not be unique to the liver and require the involvement of hepatic ABs.…”

“…In addition, internalization of ABs derived from immune cells has been reported [ 20 ]. As we established, in the liver, ABs from hepatocytes serve as a non-immune source for HSC profibrotic activation [ 9 , 11 ]. However, the liver is a graveyard for activated immune cells, which accumulate many HIV-infected circulating lymphocytes coming to the liver to die [ 21 , 22 ].…”