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Neuroinflammation is a major component in the transition to and perpetuation of neuropathic pain states. Spinal neuroinflammation involves activation of TLR4, localized to enlarged, cholesterol-enriched lipid rafts, designated here as inflammarafts. Conditional deletion of cholesterol transporters ABCA1 and ABCG1 in microglia, leading to inflammaraft formation, induced tactile allodynia in naive mice. The apoA-I binding protein (AIBP) facilitated cholesterol depletion from inflammarafts and reversed neuropathic pain in a model of chemotherapy-induced peripheral neuropathy (CIPN) in wild-type mice, but AIBP failed to reverse allodynia in mice with ABCA1/ABCG1–deficient microglia, suggesting a cholesterol-dependent mechanism. An AIBP mutant lacking the TLR4-binding domain did not bind microglia or reverse CIPN allodynia. The long-lasting therapeutic effect of a single AIBP dose in CIPN was associated with anti-inflammatory and cholesterol metabolism reprogramming and reduced accumulation of lipid droplets in microglia. These results suggest a cholesterol-driven mechanism of regulation of neuropathic pain by controlling the TLR4 inflammarafts and gene expression program in microglia and blocking the perpetuation of neuroinflammation.

Apolipoprotein A-I binding protein (AIBP) has been shown to augment cholesterol efflux from endothelial cells and macrophages. In zebrafish and mice, AIBP-mediated regulation of cholesterol levels in the plasma membrane of endothelial cells controls angiogenesis. The goal of this work was to evaluate metabolic changes and atherosclerosis in AIBP loss-of-function and gain-of-function animal studies. Here, we show that mice fed a high-cholesterol, high-fat diet had exacerbated weight gain, liver steatosis, glucose intolerance, hypercholesterolemia, hypertriglyceridemia, and larger atherosclerotic lesions compared with mice. Feeding mice a high-cholesterol, normal-fat diet did not result in significant differences in lipid levels or size of atherosclerotic lesions from mice. Conversely, adeno-associated virus-mediated overexpression of AIBP reduced hyperlipidemia and atherosclerosis in high-cholesterol, high-fat diet-fed mice. Injections of recombinant AIBP reduced aortic inflammation in mice fed a short high-cholesterol, high-fat diet. Conditional overexpression of AIBP in zebrafish also reduced diet-induced vascular lipid accumulation. In experiments with isolated macrophages, AIBP facilitated cholesterol efflux to HDL, reduced lipid rafts content, and inhibited inflammatory responses to lipopolysaccharide.jlr Our data demonstrate that AIBP confers protection against diet-induced metabolic abnormalities and atherosclerosis.

Transforming growth factor-␤ (TGF-␤) is implicated in the pathogenesis of liver disease. TGF-␤ is involved both in liver regeneration and in the fibrotic and cirrhotic transformation with hepatitis viral infection. Hepatitis C virus (HCV) infection often leads to cirrhosis and hepatocellular carcinoma. HCV nonstructural 5A (NS5A) protein is a multifunctional protein that modulates cytokine-mediated signal transduction pathways. To elucidate the molecular mechanism of HCV pathogenesis, we examined the effect of NS5A protein on TGF-␤-stimulated signaling cascades. We show that NS5A protein inhibited the TGF-␤-mediated signaling pathway in hepatoma cell lines as determined by reporter gene assay. To further investigate the role of NS5A, we examined the protein/protein interaction between NS5A and TGF-␤ signal transducers. Both in vitro and in vivo binding data showed that NS5A protein directly interacted with TGF-␤ receptor I (T␤R-I) in hepatoma cell lines. This interaction was mapped to amino acids 148 -238 of NS5A. We also found that NS5A protein co-localized with T␤R-I in the cytoplasm of Huh7 cells and inhibited TGF-␤-mediated nuclear translocation of Smad2. Furthermore, we demonstrate that NS5A protein abrogated the phosphorylation of Smad2 and the heterodimerization of Smad3 and Smad4. To further explore the relevance to viral infection, we examined the effect of the HCV subgenomic replicon on the TGF-␤ signaling pathway. We show that the HCV subgenomic replicon also inhibited TGF-␤-induced signaling cascades. These results indicate that HCV NS5A modulates TGF-␤ signaling through interaction with T␤R

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