Sun Li-xia, Wang Zheng, Yang Bin, Liu Juan, Qiu Ping, Chen Jia-qi
Zhongshan Ophthalmic Center, Sun Yat-sen University, Guangzhou 510060, China.
Zhonghua Yan Ke Za Zhi. 2005 Jun;41(6):492-7.
To study the proliferation activity of corneal epithelial cells and the apoptosis of keratocytes in the rabbit cornea after treatment with 20% ethanol. The results were compared to the cornea treated with mechanical scraping of the epithelial cells.
The experimental group consisted of 42 rabbits. One of the two corneas of each rabbit was incised by 8 mm epithelial trephine using in LASEK and was exposed to 20% ethanol (in distilled water) in the trephine for 40 seconds. In the other eye, the corneal epithelium in the central area was mechanically scraped. The rabbits in the experimental group were randomly divided into seven sub-groups. Each sub-group had six rabbits and the rabbits were killed at 0, 4 hours, 1, 3, 5, 8 and 30 days after the surgery. Three rabbits without any treatment were used as blank controls. Immunohistochemical staining (Ki-67 antigen) was performed to detect the proliferation of corneal epithelial cells. Apoptotic cells were detected by TUNEL assay. Number of keratocytes in the central anterior stroma of cornea was determined by counting the total number of cells under 400 x magnification field in hematoxylin-eosin-stained corneal sections.
In the ethanol-treated eyes, the number of Ki-67 positive cells peaked 5 days after the treatment in the central corneal epithelium and 1 day after the treatment in the peripheral corneal epithelium. TUNEL-positive cells were detected in the anterior central stromal keratocytes under the epithelial incisions and the number of TUNEL-positive cells reached a peak 4 hours after the treatment. There was no statistically significant difference in the number of central anterior stromal keratocytes between the ethanol-treated group and the controls (P = 0.68). In the mechanical scraping group, the number of Ki-67 positive cells in the peripheral corneal epithelium peaked 3 days after the treatment. The number of Ki-67 positive cells in the peripheral corneal epithelium of the scraping group was greater than that of the ethanol-treated group. TUNEL-positive cells were detected in the anterior central stromal keratocytes and reached a peak 4 hours after the scraping. The number of central anterior stromal keratocytes was decreased maximally 1 day after the scraping (P < 0.01).
The cornea injury caused by treatment with 20% ethanol for 40 seconds is milder than that caused by mechanical scraping. The wound-healing process in the ethanol-treated cornea is faster than that of the cornea treated with scraping. Corneal epithelium treated with 20% ethanol for 40 seconds can protect the stromal keratocytes.
研究20%乙醇处理后兔角膜上皮细胞的增殖活性及角膜基质细胞的凋亡情况，并与机械刮除上皮细胞处理的角膜进行结果对比。
实验组有42只兔。每只兔的两只眼中，一只眼采用准分子激光上皮下角膜磨镶术（LASEK）用8mm上皮环钻切开，在环钻内暴露于20%乙醇（溶于蒸馏水）中40秒。另一只眼，中央区域的角膜上皮进行机械刮除。实验组的兔被随机分为7个亚组。每个亚组有6只兔，分别在术后0、4小时、1、3、5、8和30天处死。3只未做任何处理的兔作为空白对照。采用免疫组织化学染色（Ki-67抗原）检测角膜上皮细胞的增殖情况。通过末端脱氧核苷酸转移酶介导的缺口末端标记法（TUNEL法）检测凋亡细胞。通过对苏木精-伊红染色的角膜切片在400倍放大视野下计数角膜中央前基质层的角膜基质细胞总数来确定其数量。
在乙醇处理的眼中，中央角膜上皮中Ki-67阳性细胞数量在处理后5天达到峰值，周边角膜上皮中在处理后1天达到峰值。在上皮层切口下方的中央前基质层角膜基质细胞中检测到TUNEL阳性细胞，其数量在处理后4小时达到峰值。乙醇处理组与对照组中央前基质层角膜基质细胞数量无统计学显著差异（P = 0.68）。在机械刮除组中，周边角膜上皮中Ki-67阳性细胞数量在处理后3天达到峰值。刮除组周边角膜上皮中Ki-67阳性细胞数量多于乙醇处理组。在中央前基质层角膜基质细胞中检测到TUNEL阳性细胞，在刮除后4小时达到峰值。刮除后1天中央前基质层角膜基质细胞数量减少最多（P < 0.01）。
20%乙醇处理40秒引起的角膜损伤比机械刮除引起的损伤轻。乙醇处理的角膜伤口愈合过程比刮除处理的角膜快。20%乙醇处理40秒可保护角膜基质细胞。
