Expression of lysosome-associated membrane protein 1 (Lamp-1) and galectins in human keratinocytes is regulated by differentiation
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Lysosomes and their components are suspected to be involved in epidermal differentiation. In this study, lysosomal enzyme activities, expression of the lysosome-associated membrane protein 1 (Lamp-1) and expression of the epidermal galectins-1, -3 and -7 were investigated in human keratinocytes cultured at different cell densities (subconfluence, confluence and postconfluence) in order to induce differentiation. Detected by Western blot and immunofluorescence, Lamp-1 expression is transiently upregulated at culture confluence, but reduced at postconfluence. Northern blot analyses performed on subconfluent, confluent and post-confluent cultures of keratinocytes show that Lamp-1 mRNA expression is also upregulated at culture confluence, but downregulated at postconfluence. Measurements of lysosomal enzyme activities indicate a transient upregulation at culture confluence, whereas cathepsins B, C and L are particularly downregulated at postconfluence. Cell density and differentiation of epidermal cells also differentially regulates galectin expression in autocrine cultures. As the expression of galectin-1 mRNA is high in subconfluent cells, it is assumed to be associated with their proliferative state. On the other hand, as the mRNA levels for galectins-3 and -7 are notably upregulated at culture confluence (galectin-7) or at postconfluence (galectin-3), their expression is thought to be related to the differentiated state of keratinocytes. However, we collected evidence by confocal microscopy that galectin-3 and Lamp-1 do not colocalize in vitro in keratinocytes. Altogether, our results suggest that the upregulated Lamp-1 expression at confluence could be involved in keratinocyte differentiation, but apparently not through interaction with galectin-3.
KeywordsKeratinocytes Lysosomes Lamp-1 Galectins Culture confluence
The authors are grateful to Prof. M. Jadot for his helpful ideas and critical remarks. The technical assistance provided by R. Deom, F. Dubois and F. Herphelin is gratefully acknowledged. We thank Dr B. Bienfait (Namur-Bouge) for providing skin specimens. Special thanks are addressed to Drs D.R. Roop (Houston), R.L. Eckert (Cleveland) and F. Van Den Brûle (Liège) for their gift of keratins, involucrin and galectins cDNA probes, respectively.
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