Galkowska Hanna, Olszewsk Waldemar L, Wojewodzka Urszula, Mijal Joanna, Filipiuk Ewa
Department of Surgery and Transplantation and the Laboratory of Cell Ultrastructure, Medical Research Center, Polish Academy of Sciences and Central Clinical Hospital, Ministry of Internal Affaires, Warsaw, Poland.
Surgery. 2003 Aug;134(2):213-20. doi: 10.1067/msy.2003.223.
Epithelialization of cutaneous ulcers is a long-lasting process. To study the pathomechanism of impaired epithelialization, we evaluated the role of cell cycle- and apoptosis-related proteins in the regenerating epidermis. We characterized immunohistochemically the expression of cell cycle regulators p63, CD29, PCNA, p53, pro- and antiapoptotic proteins bcl2, bax, caspase 3 and DNA breaks, as well as keratin 10, 16 and 17.
Studies were carried out in 12 patients with diabetic foot, and 10 patients with varicose ulcers of the calf. Skin biopsy specimens were obtained from the border area of ulcers and the topographically corresponding sites of normal skin of patients undergoing orthopedic surgery. Biopsy specimens were stained by use of specific primary antibodies, a kit based on biotin-avidin-peroxidase complex technique, and DAB substrate. Results were expressed as a mean staining intensity.
At the edge of both types of ulcers, keratinocytes were p63+, CD29+, PCNA+ and p53-. The mean intensity of p63 and CD29 staining was slightly higher than in controls. The intensity of bcl2 staining was higher at the edge of diabetic ulcers compared with venous ulcers, whereas the intensity of bax staining was similar. The expression of caspase 3 was lower at the edge of venous ulcers and higher in diabetic ulcers and the intensity of TUNEL staining was lower at the edge of both types of ulcers compared with controls. Keratinocytes at the edge and distally to both types of ulcers expressed cytokeratin 16 and 17. There was no expression of cytokeratin 10 at the edge of ulcers. Together, there was a slight tendency for higher expression of cell cycle-related proteins in venous and of apoptosis-related proteins in diabetic ulcers epidermis; however, the differences were minor.
The impaired epithelialization of chronic leg ulcers is not caused by an inadequate epidermal stem cell proliferation, differentiation, or apoptosis. It may rather reflect the distorted organization of wound bed, caused by infection and impaired nutrition supply, altering keratinocyte migration. To accelerate healing of an ulcer, modeling of the granulation tissue by regulatory cytokines but not stimulation of keratinocyte growth seems to be indicated.
皮肤溃疡的上皮化是一个持久的过程。为了研究上皮化受损的发病机制，我们评估了细胞周期和凋亡相关蛋白在再生表皮中的作用。我们通过免疫组织化学方法对细胞周期调节因子p63、CD29、增殖细胞核抗原（PCNA）、p53、促凋亡和抗凋亡蛋白bcl2、bax、半胱天冬酶3以及DNA断裂，还有角蛋白10、16和17的表达进行了特征描述。
对12例糖尿病足患者和10例小腿静脉曲张溃疡患者进行了研究。从溃疡边缘区域以及接受骨科手术患者正常皮肤的地形对应部位获取皮肤活检标本。活检标本使用特异性一抗、基于生物素 - 抗生物素蛋白 - 过氧化物酶复合物技术的试剂盒以及DAB底物进行染色。结果以平均染色强度表示。
在两种类型溃疡的边缘，角质形成细胞p63阳性、CD29阳性、PCNA阳性且p53阴性。p63和CD29染色的平均强度略高于对照组。与静脉性溃疡相比，糖尿病性溃疡边缘bcl2染色强度更高，而bax染色强度相似。静脉性溃疡边缘半胱天冬酶3的表达较低，糖尿病性溃疡中较高，并且与对照组相比，两种类型溃疡边缘TUNEL染色强度均较低。两种类型溃疡边缘及远端的角质形成细胞表达细胞角蛋白16和17。溃疡边缘未检测到细胞角蛋白10的表达。总体而言，静脉性溃疡表皮中细胞周期相关蛋白有稍高表达的趋势，糖尿病性溃疡表皮中凋亡相关蛋白有稍高表达的趋势；然而，差异较小。
慢性腿部溃疡上皮化受损并非由表皮干细胞增殖、分化或凋亡不足引起。它可能更反映了由感染和营养供应受损导致的伤口床组织紊乱，从而改变了角质形成细胞的迁移。为了加速溃疡愈合，似乎需要用调节性细胞因子对肉芽组织进行塑形，而不是刺激角质形成细胞生长。
