Human keratinocyte cell lines differ in the expression of the collagenolytic matrix metalloproteinases-1,-8, and-13 and of TIMP-1

We investigated cells and conditioned media of the three human keratinocyte cell lines HaCaT (non-tumorigenic), A5 (benign, tumorigenic) and II-4RT (m alignant, tumorigenic) with regard to production and secretion of the colla genases-1 to -3 (MMP-1, MMP-8 and MMP-13) and TIMP-1 using semi-nested RT-P CR, Western blots, ELISA, immunocytochemistry and casein zymography. Transcripts of MMP-1, -8, -13 and TIMP-1 were detected in all cell lines by RT-PCR and the corresponding proteins were found in the cytoplasm of all t hree cell lines by Western blot analysis and/or immunocytochemistry. The co nditioned media of the malignant II-4RT cells contain significantly more MM P-1 and MMP-8 than those of HaCaT or A5 as evidenced by immunoblotting and ELISA. In addition to the presence of latent MMP-1, zymography also detecte d the active form of this enzyme. TIMP-1 was found only in extracts of all three cell lines, predominantly in A5. This study clearly indicates that the epithelial tumor cells synthesize dif ferent collagenases and TIMP-1. The malignant clone secretes increased amou nts of distinct collagenases compared to the non-tumorigenic cell line, the reby verifying a correlation between biological behaviour and the amount of collagenases. In addition, we provide clear evidence that MMP-8 is not exc lusively found in polymorphonuclear granulocytes, but also in keratinocyte cell lines.