Boban Ivana, Barisic-Dujmovic Tatjana, Clark Stephen H
Department of Genetics and Developmental Biology, University of Connecticut Health Center, Farmington, Connecticut 06030, USA.
J Cell Physiol. 2008 Jan;214(1):230-7. doi: 10.1002/jcp.21182.
To test the hypothesis of an extra-dermal origin of dermal fibroblasts, parabiosis, and transplantation models were developed utilizing a collagen promoter green fluorescent protein (GFP) reporter transgene expressed in dermal fibroblasts. Parabiotic pairs were treated with bleomycin to induce the skin fibrosis that was evaluated for a dense deposition of collagen and inflammatory cell infiltrates in the thickened dermis in comparison with parabiotic pairs treated with saline. Although, in all cases, repeated injection of bleomycin for 4 weeks induced skin fibrosis, only a few GFP positive cells were detected in skin samples from some of the treated non-transgenic mice. Unexpectedly, similar results were observed in saline treated controls. Furthermore, bone marrow chimeras were created in which non-transgenic recipient mice received injections of bone marrow cell preparations isolated from pOBCol3.6GFP transgenic mice. After bone marrow chimerism had been successfully established, fibrotic lesions in the skin were induced by local bleomycin injections. Donor GFP expressing cells were observed in the skin from all recipient mice. However, no difference in the presence of GFP expressing cells was observed between non-treated mice or mice treated with bleomycin or saline. A large number of GFP expressing cells were observed in the lung preparations from all chimeric mice. Mac-3 antibody immunostaining confirmed a macrophage phenotype for these GFP expressing cells suggesting the expression of the pOBCol3.6GFP transgene in a non-collagen producing cell. Based on these observations, we found no evidence of circulating dermal fibroblast progenitors that participate in the development of bleomycin-induced skin fibrosis.
为了验证真皮成纤维细胞的真皮外起源假说，利用在真皮成纤维细胞中表达的胶原蛋白启动子绿色荧光蛋白（GFP）报告转基因，建立了联体共生和移植模型。联体共生对用博来霉素处理以诱导皮肤纤维化，与用盐水处理的联体共生对相比，评估增厚真皮中胶原蛋白的密集沉积和炎性细胞浸润情况。尽管在所有情况下，连续4周注射博来霉素均诱导了皮肤纤维化，但在一些经处理的非转基因小鼠的皮肤样本中仅检测到少数GFP阳性细胞。出乎意料的是，在盐水处理的对照组中也观察到了类似结果。此外，构建了骨髓嵌合体，其中非转基因受体小鼠接受了从pOBCol3.6GFP转基因小鼠分离的骨髓细胞制剂注射。成功建立骨髓嵌合后，通过局部注射博来霉素诱导皮肤出现纤维化病变。在所有受体小鼠的皮肤中均观察到表达供体GFP的细胞。然而，在未处理的小鼠或用博来霉素或盐水处理的小鼠之间，在表达GFP的细胞存在情况上未观察到差异。在所有嵌合小鼠的肺制剂中均观察到大量表达GFP的细胞。Mac-3抗体免疫染色证实这些表达GFP的细胞具有巨噬细胞表型，这表明pOBCol3.6GFP转基因在非胶原蛋白产生细胞中表达。基于这些观察结果，我们没有发现参与博来霉素诱导的皮肤纤维化发展的循环真皮成纤维细胞祖细胞的证据。
