INTERLEUKIN-1-BETA REGULATES VERO CELL INTERLEUKIN-1 RECEPTOR-TYPE-I MESSENGER-RIBONUCLEIC-ACID EXPRESSION

Embryos cocultured with Vero cells display enhanced development In vit ro, This could be due to an interaction between the embryo and cellula r layer mediated by paracrine cytokines. The interleukin-1 (IL-1) syst em is composed of two IL-1 agonists, IL-1 receptor antagonist (IL-1ra) , and two major IL-1 receptors (IL-1R tI and tII). In this study, we m easured Vero cell expression of IL-1 system mRNAs with reverse transcr iption polymerase chain reaction (RT-PCR) and validated the results wi th an immunohistochemistry study. RT-PCR revealed beta-actin and IL-1R tl mRNA expression in Vero cell cocultures without detectable IL-1 be ta and IL-1ra mRNA expression. To determine the effect of IL-1 beta on IL-1R tI mRNA expression in Vero cells, quantitative competitive PCR methodology was developed. A competitive cDNA fragment was coamplified as an internal standard with the target cDNA sequence of IL-1R tI, sh owing a 50% decrease in Vero cell IL-1R tl cDNA cultured in the presen ce of IL-1 beta (100 IU/ml) compared to control Vero cell cultures (62 .5 fg vs. 125 fg). Down-regulation of IL-1R tl mRNA by IL-1 beta is do se-dependent, with increasing concentrations (0-1000 IU/ml) of IL-1 be ta producing progressive attenuation of IL-1R tl expression. Treatment with anti-IL-1 beta antibody ablated the inhibitory effect of IL-1 be ta (100 IU/ml) on IL-1R tl mRNA expression. Immunostaining confirmed t he presence of IL-1R tl protein in Vero cells. These results demonstra te the presence of IL-1R tl in Vero cell monolayers and regulation of IL-1R tl mRNA by IL-1 beta.