The Effect of Carotenoids on Transforming Growth Factor β1‐induced Fibrogenesis in LX‐2 cells, a Human Hepatic Stellate Cell Line

Abstract

Non‐alcoholic fatty liver disease (NAFLD) is the most common chronic liver disease in the United States. In response to oxidative stress or inflammation, simple steatosis can progress to non‐alcoholic steatohepatitis (NASH), an advanced form of NAFLD. Fibrosis is one of key features of NASH with increased accumulation of extracellular matrix (ECM) in the liver. Hepatic stellate cells (HSCs) play important roles in the development of liver fibrosis as they are the primary source of ECM in the damaged liver. We previously demonstrated that astaxanthin (ASTX), a xanthophyll carotenoid, prevents the activation of HSCs and attenuated the expression of fibrogenic markers, such as α‐smooth muscle actin (α‐SMA) and procollagen type I α1 (COL1A1) in LX‐2 (a human HSC cell line) as well as primary mouse and human HSCs. The objective of this study was to compare the efficacy of other carotenoids, including lycopene, lutein, zeaxanthin and canthaxanthin, in inhibiting HSC activation with that of ASTX in LX‐2 cells. Lycopene is a carotene while the other carotenoids are xanthophylls with very similar structure. LX‐2 cells were treated with 0, 10, 25, 50 and 100 μM of each carotenoid for 24 h to measure cytotoxicity. While most cells were viable with all concentrations of ASTX and zeaxanthin, cells treated with 100 μM of lycopene, lutein or canthaxanthin were ~60–70% viable. To evaluate the effect of the carotenoids on the activation of LX‐2 cells by transforming growth factor β1 (TGFβ1), a potent fibrogenic cytokine, the cells were pretreated with 25 μM each carotenoid for 24 h, followed by the treatment with 4 ng/mL for 24 h. TGFβ1 treatment markedly increased mRNA of α‐SMA and COL1A1 in LX‐2 cells by ~4‐fold and 5‐fold, respectively. ASTX repressed TGFβ1‐induced expression of α‐SMA and COL1A1 by ~60% and ~30%, respectively. Lutein also decreased the expression of α‐SMA and COL1A1 by 85% and 70%, respectively. Basal and TGFβ1‐induced α‐SMA and COL1A1 protein levels were noticeably decreased by lutein and ASTX with lutein being more potent. TGFβ1‐induced COL1A1 protein expression was attenuated by lycopene, zeaxanthin and canthaxanthin, albeit less than lutein and ASTX. In conclusion, the results suggest that ASTX and lutein inhibit TGFβ1‐induced HSC activation and fibrogenic gene expression while zeaxanthin and canthaxanthin did not exert the same effect despite their similar structure to ASTX and lutein.Support or Funding InformationUnited States Department of Agriculture, AFRI (2012‐67018‐19290)