Keratinocyte differentiation induced by calcium, phorbol ester or interferon-γ elicits distinct changes in the retinoid signalling pathways

Background: Retinoids influence keratinocyte proliferation and differentiation via binding to nuclear retinoic acid receptors (RARα, -γ) and retinoid X receptor α (RXRα). The effect of keratinocyte differentiation on expression of nuclear retinoid receptors and on the conversion of retinol into retinoic acid has not been examined earlier in depth. Objectives: Our aim was to examine the expression of retinoid receptors and a retinoid-regulated gene CRABPII, as well as the metabolism of exogenous [3H]retinol in cultured human keratinocytes induced to differentiate by exposure to either calcium, phorbol 12-myristate 13-acetate (PMA), or interferon-γ (IFNγ). Methods: Normal human keratinocytes were cultured and exposed to differentiation-inducing agents. The mRNA and protein expression of retinoid receptors were examined using real-time PCR and Western blot. [3H]Retinol uptake and metabolism was monitored by HPLC with on-line radioactivity detection. Results: In calcium-exposed cells, increased expression of RARγ and RXRα, enhanced metabolism of [3H]retinol to 3,4-didehydro-RA (ddRA), and an induction of CRABPII mRNA and protein was noted. In contrast, treatment with PMA and IFNγ reduced the RARγ and RXRα protein expression (preventable by the proteasome inhibitor MG132), increased the accumulation of [3H]RA and/or [3H]ddRA in the cells, and changed the CRABPII transcription. Conclusions: Retinoid signalling is profoundly altered upon differentiation of keratinocytes and the effects depend on how cellular differentiation is initiated.