Increased Lysyl oxidase-like 2 (LOXL2) gene expression accurately designates advanced liver fibrosis induced by the Hepatitis C Virus (HCV) regardless race, blood specimen or co-infection with the Human Immunodeficiency Virus (HIV)

Abstract

Background: The LOX gene encodes a member of the lysyl-oxidase (LOX) gene family, which is involved in biogenesis of connective tissue, and 4 LOX-like proteins (LOXL1 – 4) have been identified so far. This study was aimed to investigate gene expression of LOXL1 – 4 in association with fibrogenesis in Hepatitis C Virus (HCV) infected patients with and without co-infection with the Human Immundeficiency Virus (HIV) in peripheral blood mononuclear cells (PBMC), peripheral whole blood (PwB) and liver tissue (LT) samples. Materials and Methods: PwB samples from 34 therapy-naive patients (34/34 Caucasians, 18/34 males, 28/34 with HCV-genotype-1) with chronic HCV infection treated at the University Hospital Essen (Germany) and PBMC samples from 23 HIV/HCV co-infected subjects (11/23 African Americans and none Caucasian, 20/23 males, 23/23 with HCV-genotype-1) recruited at the NIAID/NIH (MD/U.S.A.) were enrolled in this study. Liver biopsies obtained at baseline were evaluated using the Batts-Ludwig staging (BL-S) system for HCV mono-infected patients and the Ishak score (IS) for HIV/HCV co-infected donors. 34 PwB and 23 PBMC samples (and 7 LT samples from HIV/HCV co-infected participants) were subjected to microarray analysis. For all statistical analyses (i.e. pair-wise group comparison) 1-way-ANOVA was performed. Overall, a P< 0.05 was considered significant. Results: PwB samples from 34 HCV mono-infected patients were grouped according to BL-S as early fibrosis (BL-S: 0 – 1, “EF”, N= 20,) and late fibrosis (BL-S: 2 – 4, “LF”, N= 14) and PBMC samples from 23 HIV/HCV co-infected patients were grouped based on IS as early fibrosis (IS: 0 – 2, “EF”, N= 20) and late fibrosis (IS: 3 – 6, “LF”, N= 14). We consistently detected significantly increased LOXL-2 gene expression levels in association with “LF” in HCV mono-infected patients (PwB, P< 0.041) and HIV/HCV co-infected subjects (PBMC, P< 0.038). In addition, linear regression analysis was performed between fibrosis grades (IS: 0 – 6) and LOXL1 – 4 gene expression levels measured in LT samples of 7 HIV/HCV co-infected subjects; here, a significant correlation was found only for LOXL1 (r= 0.8, P> 0.031). These results are currently being validated by quantitative real-time polymerase chain reaction (qRT-PCR). Conclusions: This study suggests significant involvement of LOX-like proteins, especially LOXL2, in the fibrogenic liver transformation process during chronic HCV infection. Previous research reported: 1. Association between increased mRNA levels of LOXL2 and type-III-procollagen in liver fibrosis in mice. 2. Abnormal deposition of collagen in hepatocytes obtained from humans with Wilson's disease. 3. LOXL2 involvement in fibroblast activation. These findings (1.-3.) along with the present data demonstrating consistent significance in two demographically very heterogeneous cohorts further highlight the pathogenic role suggested for LOXL in fibrogenesis and the importance of generating LOXL2-targeted therapies for the prevention/treatment of liver fibrosis.