RESUMO
BACKGROUND: In chronic renal disease, renal tubular epithelial cell (RTC) Fas expression is up-regulated, leading to apoptotic RTC deletion and tubular atrophy. In vitro, cytokine- or hypoxia-induced up-regulation of Fas expression is associated with RTC apoptosis. In contrast, constitutively expressed, low level RTC Fas does not mediate apoptosis, suggesting that Fas may be coupled to expression level-dependent RTC signaling pathways. Fas is known to signal through JNK in many systems, but the requirement of JNK activation for apoptosis remains controversial. METHODS: To determine if RTC Fas regulates JNK activity and apoptosis, human RTC were transfected with graded concentrations of a eukaryotic expression vector for murine Fas. Apoptosis was measured by annexin V, TUNEL and PARP cleavage assays. JNK activity was determined by immune complex kinase assay and/or immunoblots with phospho-specific JNK antibodies, in the presence or absence of co-expressed dominant negative JNK constructs. RESULTS: Fas antibody stimulation of RTC with high Fas expression levels (to model RTC phenotype in renal disease) caused a tenfold increase in apoptosis, while RTC with low level Fas expression (to model normal RTC phenotype) were apoptosis-resistant. Fas ligation activated JNK in RTC expressing low levels of Fas, but not in apoptosis-sensitive RTC with increased Fas expression. Dominant negative JNK co-expression failed to inhibit apoptosis in RTC expressing high levels of Fas, suggesting that JNK is neither necessary, nor sufficient, for Fas-dependent apoptosis. CONCLUSIONS: At high levels of expression, RTC Fas promotes apoptosis in a JNK-independent manner. At low basal expression, Fas induces JNK activation, but not apoptosis, consistent with novel roles for RTC Fas as a mediator of cell stress or chronic inflammation.