Calcium regulates the expression of insulin-like growth factor binding protein-3 by the human keratinocyte cell line HaCaT

The insulin-like growth factor (IGF) system is essential for epidermal home ostasis, Insulin-like growth factor binding protein 3 (IGFBP-3), a modulato r of IGF action that also exhibits IGF-independent activity, is localized t o selected keratinocytes in the basal epidermal layer and may thus contribu te to keratinocyte differentiation. We have utilized the human keratinocyte cell line, HaCaT, to examine the effect of calcium on the regulation of co mponents of the IGF system. Western ligand and northern blot analyses revea led secreted IGFBP-3 and TGFBP-3 mRNA were reduced by an elevation in calci um levels in the culture medium. At 1.0 and 1.2 mM CaCl2 culture conditions IGFBP-3 abundance was reduced to 36% +/- 1.6% and 26% +/- 7.1.%, respectiv ely, of that from cells grown at 0.03 mM CaCl2, IGFBP-3 mRNA levels in 0.7 mM and 1.2 mM CaCl2 were reduced to 46% +/- 17.4% and 24% +/- 4.6%, respect ively, compared with IGFBP-3 mRNA levels at 0.03 mM CaCl2. The observed red uction of IGFBP-3 was not associated with IGFBP-3 proteolysis. In contrast IGF-I receptor protein and mRNA levels remained unchanged. The IGF-I stimul ated proliferative response of HaCaT keratinocytes showed that under low (0 .03 mM) and high (1.2 mM) CaCl2, conditions TGF-I at 100 and 1000 ng per mi similarly increased cell number 2.4- and 2.7-fold, respectively, with simi lar dose-response curves. HaCaT keratinocytes grown under medium (0.7 mM) a nd high (1.2 mM), but not low (0.03 mM), CaCl2 conditions for 21 d revealed an induction of profilaggrin mRNA, a marker of keratinocyte differentiatio n. These studies indicate that the exposure of HaCaT keratinocytes to eleva ted calcium levels is associated with a decline in IGFBP-3 but not IGF-I re ceptor levels. These findings suggest a potential mechanism for the distrib ution of IGFBP-3 in the epidermis, which may be involved in the process of keratinocyte differentiation.