Objective: Our previous investigation demonstrated that angiotensin II could induce proliferation and differentiation of hepatic stellate cells, and also could up-regulate its extracellular matrix synthesis. The objective of this study was to determine the effects of 10(-5) mol/L angiotensin II on gene expression of hepatic stellate cells.
Methods: After incubation with 10(-5) mol/L angiotensin II for 48 hours, the cultured hepatic stellate cells were collected. The mRNA and total protein were obtained from cell lysate and then suppression subtractive hybridization (SSH), 2D-gel electrophoresis and MALDI-TOF mass spectrometry were used to identify these cDNAs and proteins.
Results: A total of 36 clones from the subtracted cDNA library were sequenced and compared to sequences in the GenBank using BLAST. Of the 36 differentially expressed gene fragments from the subtracted library, 13 differentially expressed gene fragments showed significant homology to other known proteins, such as ribosomal protein, beta-actin FE-3, leucyl-tRNA synthetase, CD147, pyruvate kinase, peroxiredoxin 1, and BAT3, while 2 other gene fragments encoding protein BC097361 and BC057380 and their functions were not disclosed. About 1110 and 1008 protein spots were detected by employing the ImageMaster 2D Platinum 4.9 proteome image analysis system in angiotensin-treated hepatic stellate cells and control cells separately. Among these spots 108 proteins were up-regulated while the other 153 proteins were down-regulated. Several up-regulated proteins were chosen to be excised and in-gel digest MALDI-TOF-MS and Database analysis showed that among the high expression proteins, there were prohibitin, RBL-NDP kinase 1.8x10(4) subunit, electron transfer flavoprotein alpha-subunit, guanine nucleotide binding protein, alpha 15, and heat shock 7.0x10(4) protein 5.
Conclusion: Our results suggest that the up-regulation of hepatic stellate cell mRNA influences proliferation, differentiation and apoptosis of those cells. The proteins of signal transduction, metabolizing regulation, apoptosis suppression, and fibrogenesis regulation of hepatic stellate cells were up-regulated.