Ratkaj Ivana, Bujak Maro, Jurišić Davor, Baus Lončar Mirela, Bendelja Krešo, Pavelić Krešimir, Kraljević Pavelić Sandra
University of Rijeka, Department of Biotechnology, Rijeka, Croatia.
Cell Physiol Biochem. 2012;30(4):927-42. doi: 10.1159/000341470. Epub 2012 Sep 12.
Dupuytren's disease (DD) is a nodular palmar fibromatosis that causes irreversible permanent contracture of fingers and results in the loss of hand function. Surgery still remains the only available solution for DD patients but cannot permanently cure the disease nor reduce high recurrence rates. With this rationale, we designed a study aimed at an improved understanding of the molecular mechanisms underlying DD. Our major focus was an analysis of the global gene expression profile and signalling pathways in DD cells with the aim of identifying novel biomarkers and/or therapeutic targets.
Primary cells were cultured from surgically removed diseased and healthy tissue. Microarray expression analysis (HG-U133A array, Affymetrix) and qPCR was performed with total RNA isolated from primary DD cells. Mechanistic studies involving inhibition of p38 phosphorylation were performed on normal human fibroblasts' and primary DD cells' in vitro models. Expression of stem cell markers in primary fibroblasts/myofibroblasts was assessed as well.
We identified 3 p38MAPK signalling pathway regulatory genes, THBS1, GADD45α and NUAK1, all involved in cellular proliferation and production of the extracellular matrix proteins. Inhibition of the p38MAPK signalling pathway induced down-regulation of myofibroblast markers, α-smooth muscle actin and palladin. A stem-cell like subpopulation positive for CD90 marker was identified among primary DD cells.
The study reveals involvement of the p38 MAPK pathway as a possible signalling cascade in the pathogenesis of Dupuytren's disease. Moreover, a particular stem cell-like CD90(+) subpopulation was identified that might contribute to DD development.
杜普伊特伦挛缩病(DD)是一种结节性掌部纤维瘤病,可导致手指不可逆的永久性挛缩并导致手部功能丧失。手术仍然是DD患者唯一可用的治疗方法,但无法永久治愈该疾病,也无法降低高复发率。基于这一原理,我们设计了一项研究,旨在更好地理解DD潜在的分子机制。我们的主要重点是分析DD细胞中的全局基因表达谱和信号通路,以识别新的生物标志物和/或治疗靶点。
从手术切除的患病组织和健康组织中培养原代细胞。使用从原代DD细胞中分离的总RNA进行微阵列表达分析(HG-U133A阵列,Affymetrix)和qPCR。在正常人成纤维细胞和原代DD细胞的体外模型上进行了涉及抑制p38磷酸化的机制研究。还评估了原代成纤维细胞/肌成纤维细胞中干细胞标志物的表达。
我们鉴定出3个p38丝裂原活化蛋白激酶(MAPK)信号通路调节基因,即血小板反应蛋白1(THBS1)、生长停滞和DNA损伤诱导蛋白45α(GADD45α)和NUAK1,它们均参与细胞增殖和细胞外基质蛋白的产生。抑制p38 MAPK信号通路可诱导肌成纤维细胞标志物α-平滑肌肌动蛋白和帕拉丁的下调。在原代DD细胞中鉴定出一个对CD90标志物呈阳性的干细胞样亚群。
该研究揭示了p38 MAPK通路可能作为杜普伊特伦挛缩病发病机制中的信号级联参与其中。此外,还鉴定出一个特定的干细胞样CD90(+)亚群,其可能对DD的发展有贡献。