P75NTR protein promotes L02 cells proliferation in vitro via TrkANGFR/P75NTR heterodimer signal pathway

Abstract

Objective To observe the expression of nerve growth factor(NGF)and its receptors (TrkANGFRand P75NTR) in hepatocytes and to explore the biological effects of exogenous P75NTR protein on hepatocytes.Methods L02 hepatocytes were cultured in vitro.The expression of NGF,TrkANNGFR and P75NTR in L02 cells were assessed by immunocytochemistry and fluorescent quantitation polymerase chain reaction (PCR). The effects on L02 cell proliferation by exogenous P75NTR,NGF,NGF+ P75NTR,anti-TrkANGFR and anti-P75NTR were detected by XTT assay.The effect of exogenous P75NTR on L02 cell apoptosis was measured by flow cytometry (Annexin V/PI) and the effect of exogenous P75NTR on L02 cell cycle was determined by flow cytometry (PI).Results L02 cell proliferation was promoted by P75NTR and in dose-dependent manner.The A value of NGF group and NGF+ P75NTR group was 0.4916±0.0565 and 0.5839 ± 0.0733,respectively,and there was statistical significance compared with control group (0.3601 ± 0.0310,P＜0.05).The A value ot anti-TrkANGFR group was 0.2689±0.0229,and there was statistical significance compared with control group (P=0.003).The A value of anti P75NTR was 0.3524 ± 0.0312,and there was no statistical significance compared with control group (P=1.000). Exogenous P75NTR had anti-apoptosis effect on L02 cells,the antiapoptosis effect was strongest when 100 ng/ml P75NTR was added and the expression quantity was 3.70 ±0.26.However there was no statistical significant compared with control group (4.10 ± 0.62,P=1.000).P75NTR affected the cell cycle S phase of L02 cells and in dose-dependent manner,which was inverted U shaped curve.The effect was strongest when the concentration was 100 ng/ml (25.60 ±0.40) and there was statistical significance compared with control group (20.10 ±1.00,P=0.000).Exogenous NGF,P75NTR and NGF + P75NTR up regulated the gene expression of NGFmRNA,TrkANGFRmRNA and P75NTR mRNA in L02 cells and there was statistical significance compared with control group (P＜0.05).There was no significant difference in the gene expression of NGFmRNA,TrkANGFFR mRNA and P75NTR mRNA between anti-TrkANGFR,anti-P75NTR groUp and control group (P＞0.05).Conclusion NGF and its receptors TrkANGFR and P75NTR were expressed in L02 cells.The appropriate dose of exogenous P75NTP protein promoted L02 cells proliferation via TrkANGFR/P75NTR heterodimer signal pathway. Key words: Nerve tissue proteins; Receptors,nerve growth factor; Liver; Cell division; Apoptosis; Cells,cultured; Gene expression