瘢痕疙瘩周围皮肤成纤维细胞周期分析及P53基因突变检测

目的：探讨瘢痕疙瘩周围皮肤中是否有生物学活性异常的成纤维细胞， 以期进一步了解瘢痕疙瘩的发展机制。方法：对所取新鲜组织标本进行细胞培养，通过碘化丙啶（PI）染色，激光流式细胞仪分析细胞周期，比较各组成纤维细胞处于增殖期细胞的百分比。采用PCR技术对P53基因外显子4、5、6进行扩增并测序。结果：瘢痕疙瘩周围皮肤增殖期成纤维细胞百分比与瘢痕疙瘩中央部相似（P＞0．05），介于瘢痕疙瘩边缘部与正常皮肤之间，与两者的差异均有显著性意义（P＜0．05）。所有体外培养瘢痕疙瘩周围皮肤成纤维细胞（6／6）均存在P53外显子4的点突变，3例（2／6）P53外显子5存在移码突变，均与同病人瘢痕疙瘩成纤维细胞基因突变一致，而两组正常皮肤成纤维细胞则未发现有任何形式的基因突变，结论：瘢痕疙瘩周围0.5cm皮肤内存在生物学活性异常成纤维细胞，这可能为瘢痕疙瘩浸润性生长的结果及其治疗后易复发的原因之一。

Cell cycle analysis and P53 gene mutation detection of fibroblasts derived from the surrounding skin of keloids

Objective To investigate whether there are abnormal fibroblasts in the surrounding skin of keloids for better understanding of the etiological feature of keloids. Methods Fresh samples were used for cell culture. Flow cytometry was used for analyzing cell cycles of fibroblasts derived from keloids and the surrounding skin. Proliferative cell proportions were compared between every two groups. P53 exon 4, 5 and 6 were amplified by PCR. DNA was sequenced to examine the structure of the destination gene. Results The proliferative cell proportion of the fibroblasts derived from the surrounding skin is not so high as that from the verge of keloids, but is higher than the normal skin fibroblasts derived from other part of the keloid patients or persons without keloids (P<0.05). Mutations (point and frameshift mutations) of P53 exon 4 (6/6) and exon 5(2/6) in fibroblasts derived from the surrounding skin of keloids were identified. Fibroblasts derived from keloids have the same mutations as its surrounding skin. Conclusion There are abnormal fibroblasts in the surrounding skin of keloids. It may be the consequence of keloid infiltrative growth and a reason of easy recurrence of keloid after therapy.;