Influence of proinflamatory macrophages (M-1) on proliferative activity and PPAR-γ expression in neoplastic rat hepatocytes in vitro

Abstract

We sought to analyse the proliferative activity and PPARγ expression in neoplastic and non-neoplastic rat hepatocytes, exposed to immunologically trained macrophages M1 (Mf-M1). Ten-week-old female Wistar rats were divided into two groups: I - control (n=5) and II neoplstic (n=5). To induce HCC in the neoplastic group, genotoxic diethylnitrosamine (DEN) was administered after a partial hepatectomy (PH). Hepatocytes were isolated by liver perfusion method and the mononuclear blood cells were isolated using Lymphoprep density-gradient centrifugation. After differentiation, blood cells were treated with barley-derived β-glucan (BBG) (10 μg/ml). Adhered heaptocytes and Mf were cultured in 3D Quasi-Vivo System during 24 h. Then, hepatic proliferation and PPAR γ expression were analysed after 72 h and the 1st ,the 2nd and the 3rd week of incubation. The proliferation index of control hepatocytes ranged between 0.41±0.04 – 0.43±0.03 after 72 h and after the 3rd week of incubation respectively. Exposure of these cells to Mf resulted in marked increase of IP after the 1st, the 2nd and the 3rd week of incubation. When hepatocytes were influenced by proinflammatory Mf- M1, their proliferation was maintained at control stage. DEN-obtained hepatocytes, not influenced by macrophages, exhibit enhanced proliferation. When these cells where co-cultured with Mf-M1, marked (P≤0.05) inhibition of cell proliferation was observed. In such condition, a high negative relationship (r= -0.93) between proliferative activity of the hepatocytes and the PPARγ concentration was observed. We conclude that immunologically trained macrophages M1, are capable to activation of PPARγ in rat neoplastic hepatocytes derived from experimentally induced HCC. In turn intensified expression of PPARγ may inhibited proliferation of neoplastic cell in vitro.