Amelioration of hepatic fibrosis via beta-glucosylceramide-mediated immune modulation is associated with altered CD8 and NKT lymphocyte distribution

Background: While CD8 lymphocytes possess pro-fibrogenic properties and NK (non-T) cells are antifibrogenic, the role of NKT lymphocytes in liver fibrosis is still unclear. beta-Glucosylceramide (GC), a naturally occurring glycolipid, exerts modulatory effects on these cells. Aim: To explore the role of NKT cells in hepatic fibrosis via GC. Methods: Hepatic fibrosis was induced by biweekly intraperitoneal (IP) carbon tetrachloride (CCl4) administrations for 7 weeks in 5 groups (A-E) of male C57Bl/6 mice. Mice were treated with daily IP GC injections in groups A and C, or daily oral doses in groups B and D. GC was administered either for the duration of the study period (in groups A and B), or for the last 3 weeks of CCl4 induction (groups C and D). GC-treated mice were compared with non-treated fibrotic controls (group E) and naive rodents (group F). Liver fibrosis, injury parameters and FACS analysis of lymphocytes were assessed. Results: Marked amelioration (P < 0.0001) of hepatic fibrosis observed in all GC-treated mice without altering reactive oxygen species production. As determined by Sirius red-stained liver tissue sections and measured by Bioquant (R) morphometry; all CCl4-administered groups significantly (P < 0.0001) increased the relative fibrosis area compared with naive animals. The increases were 14.4 +/- 1.03-fold in group A, 7.9 +/- 0.37-fold in group B, 5.2 +/- 0.2-fold in group C, 10.3 +/- 0.4-fold in group D and 23.8 +/- 1.9-fold in group E. Western blot analysis for alpha smooth muscle actin from liver extracts followed a similar pattern, increasing in groups A-E. A significant decrease in liver damage was observed in all GCtreated groups, as noted by a decrease in transaminase serum levels (P < 0.005). The beneficial effect of GC was associated with a significant decrease in the intra-hepatic NKT and CD8 lymphocytes as well as their attenuation of both T(h)1 and T(h)2 cytokines. Conclusions: Administration of GC had a significant anti-fibrotic effect following CCl4 administration. This effect was associated with an altered NKT and CD8 lymphocyte distribution and a cytokine shift. Immune modulation using GC may have a role in the treatment of fibrosis and other immune-mediated disorders.