Amelioration of hepatic fibrosis via Padma Hepaten is associated with altered natural killer T lymphocytes.
Hepatic fibrosis is the end-stage consequence of chronic liver disease, affecting many people worldwide. Unlike the anti-fibrotic effect of natural killer (NK) cells, CD8 and NK T subsets are considered as profibrogenic subsets. Padma Hepaten is a multi-compound herbal preparation derived from Tibetan medicine and has proven efficacy in some clinical trials and tests at the cellular level. In this study, we evaluate the immune efficacy of Padma Hepaten administered intraperitoneally (i.p.) and/or orally in a mice model of hepatic fibrosis. Hepatic fibrosis was induced by 6 weeks of biweekly i.p. carbon tetrachloride (CCl4) injections in male C57Bl6 mice. There were four groups, including naive mice, non-treated fibrotic mice and fibrotic mice treated by Padma Hepaten at weeks 5-6 of fibrosis induction either orally or by i.p. injections. Padma Hepaten was prepared at 10 mg/ml in saline and 250 microl (2.5 mg) were administered four times per week. After week 6, animals were killed. To isolate a Padma Hepaten-associated effect on lymphocytes, splenocytes were harvested from either naive or Padma Hepaten-treated non-fibrotic donors. Isolated splenocytes were therefore reconstituted into two groups of irradiated recipients. Recipients were then administered the same CCl4 regimen. Hepatic fibrosis was determined by sirius red staining of liver sections and by assessment of alpha smooth muscle actin expression compared with beta-actin (both by mRNA as well as the protein liver extract western blotting). Hepatic fibrosis and alanine aminotransferase serum levels were decreased significantly in both Padma Hepaten-treated groups compared with the non-treated fibrotic group. Padma Hepaten treatment was associated with attenuation of lymphocyte subsets in both treated groups. Using a chemiluminescence technique to assess total anti-oxidant capacities (TAC), it was found that both the plasmas and livers of mice treated by CCl4 had significantly higher TAC compared with controls. However, the levels of TAC in animals treated either by CCl4 alone or CCl4 with Padma Hepaten were similar. Adoptive transfer of Padma Hepaten-treated lymphocytes was associated with fibrosis amelioration compared with recipients with naive lymphocytes. CCl4 generates higher levels of anti-oxidant capacities, probably as a response to oxidative stress. Padma Hepaten administration attenuated hepatic fibrogenesis significantly, accompanied by attenuation of lymphocyte but not anti-oxidant capacities.
Ginsburg
I
I
Institute of Dental Research, Faculty of Dental Medicine, Hadassah-Hebrew University Medical Center, Jerusalem, Israel.
Koren
E
E
Horani
A
A
Mahamid
M
M
Doron
S
S
Muhanna
N
N
Amer
J
J
Safadi
R
R
D000199:Actins / Q000032:analysis / Q000235:genetics; D019264:Adoptive Transfer; D000818:Animals; D015415:Biomarkers / Q000032:analysis; D015153:Blotting, Western; D002251:Carbon Tetrachloride; D005434:Flow Cytometry / Q000379:methods; D008099:Liver / Q000737:chemistry / Q000276:immunology* / Q000473:pathology; D008103:Liver Cirrhosis / Q000276:immunology* / Q000473:pathology / Q000628:therapy*; D008214:Lymphocytes / Q000276:immunology; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D023421:Models, Animal; D055611:Natural Killer T-Cells / Q000276:immunology*; D008517:Phytotherapy / Q000379:methods*; D010936:Plant Extracts / Q000008:administration & dosage*; D014916:Whole-Body Irradiation
D000199:Actins; D015415:Biomarkers; C543472:Padma Hepaten; D010936:Plant Extracts; D002251:Carbon Tetrachloride