Tang W, Ziboh V A
Department of Dermatology, School of Medicine, University of California, Davis 95616.
J Invest Dermatol. 1991 Jan;96(1):134-8. doi: 10.1111/1523-1747.ep12515934.
Extracellular calcium concentrations markedly affect the pattern of proliferation and differentiation in cultured keratinocytes. When medium contains 0.1 mM calcium or above, the cells lose their proliferative ability, rapidly stratify, and terminally differentiate. Because 1,25(OH)2D3 (a modulator of Ca++ homeostasis) enhances the differentiation of keratinocytes, we investigated whether a link exists between 1,25(OH)2D3-induced release of inositol-1,4,5-trisphosphate (Ins(1,4,5)P3) from PtdIns 4,5-P2 and intracellular calcium [Ca++]i release from keratinocytes. Specifically, primary culture of keratinocytes were loaded with fluorescence dye Fura-2AM (10 microM) and changes in fluorescence intensity were monitored at the excitation wavelengths of 340 and 380 nm and emission wavelength of 505 nm. Additions of two agonists, 1,25(OH)2D3 (1.2 x 10(-9) M) and 13-Cis retinoic acid (0.2 x 10(-9) M), to dye-loaded keratinocytes induced rapid release of [Ca++]i, respectively, followed by gradual return to the prestimulated state. Addition of Ins(1,4,5)P3 (10 microM) to saponin-treated (leaky) keratinocytes also resulted in a rapid release of [Ca++]i. In contrast, the addition of inositol-1,3,4,5-tetrakisphosphate Ins(1,3,4,5)P4 at similar concentrations exerted negligible effect. Taken together, these results support the view that 1,25(OH)2D3-induced [Ca++]i release in keratinocytes may be via the Ins(1,4,5)P3-induced early release of intracellular [Ca++]i. This may explain, at least in part, 1,25(OH)2D3-enhanced keratinocyte differentiation.
细胞外钙浓度显著影响培养的角质形成细胞的增殖和分化模式。当培养基中钙浓度达到或高于0.1 mM时，细胞失去增殖能力，迅速分层并终末分化。由于1,25(OH)2D3（一种钙稳态调节剂）可增强角质形成细胞的分化，我们研究了1,25(OH)2D3诱导磷脂酰肌醇4,5-二磷酸（PtdIns 4,5-P2）释放肌醇-1,4,5-三磷酸（Ins(1,4,5)P3）与角质形成细胞内钙（[Ca++]i）释放之间是否存在联系。具体而言，将角质形成细胞原代培养物用荧光染料Fura-2AM（10 microM）负载，并在激发波长340和380 nm以及发射波长505 nm处监测荧光强度变化。向负载染料的角质形成细胞中添加两种激动剂，即1,25(OH)2D3（1.2×10(-9) M）和13-顺式视黄酸（0.2×10(-9) M），分别诱导[Ca++]i迅速释放，随后逐渐恢复到刺激前状态。向皂素处理（通透）的角质形成细胞中添加Ins(1,4,5)P3（10 microM）也导致[Ca++]i迅速释放。相反，添加浓度相似的肌醇-1,3,4,5-四磷酸Ins(1,3,4,5)P4则产生可忽略不计的影响。综上所述，这些结果支持以下观点：1,25(OH)2D3诱导角质形成细胞中[Ca++]i释放可能是通过Ins(1,4,5)P3诱导细胞内[Ca++]i早期释放实现的。这可能至少部分解释了1,25(OH)2D3增强角质形成细胞分化的现象。
