BACKGROUND: Cytokines stimulate the expression of the adhesion molecule VCAM-1 in renal tubular epithelial cells. We have recently shown that VCAM-1 can also be upregulated by low molecular weight breakdown products of the matrix constituent hyaluronan (HA) (J Immunol 1998; 161: 3431-3437). The mechanisms of VCAM-I expression in response to HA remain to be defined. METHODS: Using a defined mouse cortical tubular (MCT) cell line we investigated the effect of protein kinase C (PKC) and tyrosine kinase (TK) inhibition on the HA-stimulated VCAM-1 expression by cell ELISA and RT PCR or Northern blotting. Furthermore, we examined the effect of PKC and TK inhibition on NF-kappaB. RESULTS: We found that the PKC inhibitor GF109203X (acting on conventional, novel and atypical isoforms) inhibited the HA-stimulated VCAM-1 expression in MCT cells dose-dependently up to 90%, whereas chelerythrine (acting on conventional and novel isoforms) had no effect. Downregulation of PKC with PMA did not prevent the HA-stimulated VCAM-1 expression, suggesting that Ca2+- and diacylglycerol-independent (atypical) isoforms of PKC are involved. The TK inhibitor genistein also inhibited the HA-stimulated VCAM-1 expression at the mRNA and protein level up to 70%. Interestingly, the HA-stimulated nuclear translocation of NF-kappaB could not be prevented with GF109203X and genistein. CONCLUSION: These data demonstrate that the HA-stimulated VCAM-1 expression in MCT cells involves PKC and TK pathways. The absence of an effect of PKC and TK inhibitors on the nuclear translocation of NF-kappaB suggests that additional transcription factors are involved for VCAM-1 expression.
BACKGROUND: Cytokines stimulate the expression of the adhesion molecule VCAM-1 in renal tubular epithelial cells. We have recently shown that VCAM-1 can also be upregulated by low molecular weight breakdown products of the matrix constituent hyaluronan (HA) (J Immunol 1998; 161: 3431-3437). The mechanisms of VCAM-I expression in response to HA remain to be defined. METHODS: Using a defined mouse cortical tubular (MCT) cell line we investigated the effect of protein kinase C (PKC) and tyrosine kinase (TK) inhibition on the HA-stimulated VCAM-1 expression by cell ELISA and RT PCR or Northern blotting. Furthermore, we examined the effect of PKC and TK inhibition on NF-kappaB. RESULTS: We found that the PKC inhibitor GF109203X (acting on conventional, novel and atypical isoforms) inhibited the HA-stimulated VCAM-1 expression in MCT cells dose-dependently up to 90%, whereas chelerythrine (acting on conventional and novel isoforms) had no effect. Downregulation of PKC with PMA did not prevent the HA-stimulated VCAM-1 expression, suggesting that Ca2+- and diacylglycerol-independent (atypical) isoforms of PKC are involved. The TK inhibitor genistein also inhibited the HA-stimulated VCAM-1 expression at the mRNA and protein level up to 70%. Interestingly, the HA-stimulated nuclear translocation of NF-kappaB could not be prevented with GF109203X and genistein. CONCLUSION: These data demonstrate that the HA-stimulated VCAM-1 expression in MCT cells involves PKC and TK pathways. The absence of an effect of PKC and TK inhibitors on the nuclear translocation of NF-kappaB suggests that additional transcription factors are involved for VCAM-1 expression.
Authors: Judith C Kim; Diana Whitaker-Menezes; Masatoshi Deguchi; Brigette S Adair; Robert Korngold; George F Murphy Journal: Am J Pathol Date: 2002-09 Impact factor: 4.307