Characterization of osteogenic differentiation and gene expression profile of multiple miliary osteoma cutis

Multiple miliary osteoma cutis (MMOC) is a rare skin disorder characterized by heterotopic ossification in dermis and subcutis. The etiology and disease mechanism is unknown, but earlier data indicates that it seems to be different from other diseases containing heterotopic intramembranous ossification. The purpose of this study was to further investigate the pathogenesis of MMOC by comparing patients’ osteoma affected skin area to their unaffected healthy skin area, as well as to skin from controls. This comparison was made by using osteogenic differentiation studies in cell cultures and by gene expression experiments. Both patient and control dermal fibroblast-like cells were able to differentiate into osteoblast-like matrix mineralizing cells. The differentiation seems to diminish with bone morphogenetic protein (BMP) 4 and 2/7 exposures and seems to correlate with proportion of stem cell associated cell surface marker CD105 positive cells. Microarray analysis revealed altered gene expression patterns between skin samples of patients and controls, as well as between samples taken from different skin areas. BMP receptor II and G-protein α-stimulatory subunit genes were among downregulated and β-catenin among upregulated genes in patient compared to control samples. Genes for homeobox proteins were among downregulated and gene for secreted frizzled related protein 2 among upregulated genes when compared patients’ osteoma area to their unaffected area. Differences in unaffected skin area in patients and controls suggest that the nature of MMOC may be more systemic than previously thought. Detected differences in separate skin areas may, in turn, provide new information for understanding the localized characteristics of MMOC. BMPs’ inhibitory effect to osteogenic differentiation of human dermal fibroblast-like cells may provide a new perspective to the clinical use of BMPs. Skin biopsies were taken from three patients and two controls for RNA extraction and hybridization on Affymetrix microarrays.

多发性粟粒性皮肤骨瘤（Multiple miliary osteoma cutis, MMOC）是一种罕见的皮肤疾病，以真皮及皮下组织异位骨化为特征。其病因与发病机制目前尚未明确，但既往研究数据显示，该病与其他伴膜内异位骨化的疾病存在差异。本研究旨在通过对比患者骨瘤受累皮肤区域、未受累健康皮肤区域与健康对照者的皮肤样本，进一步探究MMOC的发病机制。本次对比通过细胞培养成骨分化实验与基因表达实验完成。患者与健康对照者的皮肤成纤维样细胞均可分化为成骨细胞样基质矿化细胞。该分化过程可因骨形态发生蛋白（bone morphogenetic protein, BMP）4及BMP2/7的处理而减弱，且与干细胞相关细胞表面标志物CD105阳性细胞的占比相关。微阵列分析结果显示，患者与健康对照者的皮肤样本之间，以及不同皮肤区域的样本之间，基因表达模式均存在显著差异。相较于健康对照样本，患者样本中骨形态发生蛋白受体II（BMP receptor II）与G蛋白α刺激性亚基基因呈下调表达，而β-连环蛋白（β-catenin）则呈上调表达。将患者的骨瘤受累皮肤区域与未受累区域相比，同源框蛋白编码基因呈下调表达，而分泌型卷曲相关蛋白2（secreted frizzled related protein 2）编码基因则呈上调表达。患者未受累皮肤区域与健康对照者皮肤区域的差异提示，MMOC的本质可能比此前认为的更具全身性。而不同皮肤区域之间检出的差异，则可为阐明MMOC的局部特征提供新的研究线索。骨形态发生蛋白对人皮肤成纤维样细胞成骨分化的抑制作用，可为骨形态发生蛋白的临床应用提供新的研究视角。本研究从3例患者及2例健康对照者处采集皮肤活检样本，用于RNA提取及Affymetrix微阵列杂交实验。