Lipopolysaccharides indirectly stimulate apoptosis and global induction of apoptotic genes in fibroblasts

Following Gram-negative bacterial infection there is a reduction in matrix-producing cells. The goal of the present study was to examine the apoptotic effects of lipopolysaccharide (LPS) on fibroblastic cells and to investigate the role that the host response plays in this reaction. This was accomplished in vivo by subcutaneous inoculation of LPS in wild type and TNFR1(-/-)R2(-/-) mice. The direct effects of LPS on fibroblast apoptosis was studied in vitro with normal diploid human fibroblasts. The results indicate that LPS in vivo induces apoptosis of fibroblasts. By RNA profiling we demonstrated that LPS stimulates global expression of apoptotic genes and down-regulates anti-apoptotic genes. Fluorometric studies demonstrated that LPS in vivo significantly increased caspase-8 and caspase-3 activity and by use of specific inhibitors, the activation of caspase-3 was shown to be initiated by caspase-8 with no contribution from caspase-9. In vitro studies demonstrated that LPS did not induce apoptosis of fibroblasts, whereas tumor necrosis factor (TNF) did. In addition, the pattern of apoptotic gene expression induced by TNF in vitro was nearly identical to that induced by LPS in vivo, as measured by RNase protection assay. Moreover, pre-treatment of cells with TNF greatly enhanced apoptosis induced by a second stimulation with TNF 24 h later, suggesting that the global induction of proapoptotic genes was functionally significant. Thus, LPS acts to modulate the expression of a large number of genes that favor apoptosis of fibroblastic cells that is dependent upon activation of caspase-8 and is largely mediated by TNF.