Ladin D A, Hou Z, Patel D, McPhail M, Olson J C, Saed G M, Fivenson D P
Department of Surgery (Plastics), Henry Ford Hospital, Detroit, Mich, USA.
Wound Repair Regen. 1998 Jan-Feb;6(1):28-37. doi: 10.1046/j.1524-475x.1998.60106.x.
Keloids are the result of a dysregulated wound-healing process and are characterized by formation of excess scar tissue that proliferates beyond the boundaries of the inciting wound. In this study, we investigated the expression of key proteins involved in regulating apoptosis in keloids. Twenty archival paraffin-embedded keloid samples were randomly selected for an immunoperoxidase assay with antibodies against fas, p53, bcl-2, and bcl-x proteins using the target antigen-retrieval technique. Apoptosis was assessed in keloids and normal skin and in keloid and normal fibroblasts by the TdT-mediated dUTP nick-end labeling (tunel) assay on tissue sections, fibroblast cultures, and by flow cytometry for cell suspensions. We found that 18 of 20 keloids expressed p53 protein; bcl-2 was expressed by keloid fibroblasts in 19 of 20 keloids, and all specimens had prominent fas expression throughout the tissue. The distribution of these three antigens was regional within each lesion and followed a consistent pattern of p53 and bcl-2 expression colocalized to the hypercellular, peripheral areas of each keloid in a perinuclear pattern (p < .001). In contrast, an inverse distribution of fas expression was shown, with staining being more diffuse across the cell surfaces and limited to the central, more hypocellular regions in16 of 17 keloids (p < .001). There was no specific staining pattern in these keloids with antihuman bcl-x. In vitro studies on cultured keloid fibroblasts (derived from six patients) revealed maintenance of the p53+, bcl-2+ phenotype up to passage 10. Neither neonatal nor normal adult skin fibroblasts expressed either antigen but could be induced to express p53 by exposure to adriamycin. Keloid lesions and keloid fibroblasts were found to have lower rates of apoptosis than normal controls. Keloid fibroblasts displayed enhanced apoptosis rates in response to hydrocortisone, gamma interferon, and hypoxia treatment as compared with normal adult fibroblasts. Focal dysregulation of p53 combined with upregulation of bcl-2 may help produce a combination of increased cell proliferation and decreased cell death in the younger, hypercellular areas of the keloid. This phenotype is reversed in the older areas of the keloid and may prevent malignant degeneration, thus favoring normal apoptosis as evidenced by prominent fas expression.
瘢痕疙瘩是伤口愈合过程失调的结果，其特征是形成过多的瘢痕组织，该组织增生超出了引发伤口的边界。在本研究中，我们调查了瘢痕疙瘩中参与调节细胞凋亡的关键蛋白的表达情况。随机选择20个存档石蜡包埋的瘢痕疙瘩样本，采用靶抗原修复技术，用抗fas、p53、bcl-2和bcl-x蛋白的抗体进行免疫过氧化物酶检测。通过对组织切片、成纤维细胞培养物进行TdT介导的dUTP缺口末端标记（TUNEL）检测以及对细胞悬液进行流式细胞术，评估瘢痕疙瘩和正常皮肤以及瘢痕疙瘩和成纤维细胞中的细胞凋亡情况。我们发现，20个瘢痕疙瘩中有18个表达p53蛋白；20个瘢痕疙瘩中有19个的瘢痕疙瘩成纤维细胞表达bcl-2，并且所有标本在整个组织中均有明显的fas表达。这三种抗原在每个病变内呈区域分布，并且p53和bcl-2的表达以核周模式共定位于每个瘢痕疙瘩的高细胞外周区域，呈现一致的模式（p <.001）。相比之下，fas表达呈相反分布，在17个瘢痕疙瘩中的16个中，染色在细胞表面更弥散，且局限于中央、细胞较少的区域（p <.001）。这些瘢痕疙瘩用抗人bcl-x检测时没有特异性染色模式。对培养的瘢痕疙瘩成纤维细胞（来自6名患者）的体外研究表明，直至传代10代，p53+、bcl-2+表型得以维持。新生儿和成体正常皮肤成纤维细胞均不表达这两种抗原，但暴露于阿霉素后可诱导表达p53。发现瘢痕疙瘩病变和瘢痕疙瘩成纤维细胞的细胞凋亡率低于正常对照。与成体正常成纤维细胞相比，瘢痕疙瘩成纤维细胞在氢化可的松、γ干扰素和缺氧处理后显示出更高的细胞凋亡率。p53的局部失调与bcl-2的上调相结合，可能有助于在瘢痕疙瘩较年轻、细胞较多的区域产生细胞增殖增加和细胞死亡减少的组合。这种表型在瘢痕疙瘩较老的区域会逆转，并且可能防止恶性退变，因此有利于正常细胞凋亡，如明显的fas表达所示。
