目的：通过培养、分离、纯化、鉴定眼睑基底细胞癌相关成纤维细胞(CAFs)及正常眼睑皮肤成纤维细胞(NFs)，将两种成纤维细胞的生物学特性和成纤维活化蛋白(FAP)表达情况进行对比研究，以进一步探究FAP以及CAFs在眼睑基底细胞癌发病及其发展中的作用及意义。

方法：将眼睑基底细胞癌及眼睑正常皮肤组织块贴壁培养获得原代CAFs及NFs，于倒置相差显微镜下观察纯化的两种细胞第3代的细胞形态并用细胞免疫化学染色(SP法)鉴定两种细胞； MTT法检测细胞生长增殖活力； 通过RT-qPCR、Western Blot分别检测其FAP mRNA、蛋白在两种细胞中的表达差异。

结果：眼睑CAFs呈长梭形或纺锤形，胞质突明显减少，而胞浆内具有较多颗粒，细胞体积差异明显，排列紊乱且密集，丧失接触抑制，存在明显的重叠生长现象； NFs呈长梭形，但胞浆内颗粒少见，细胞体积较均一，放射状排列，存在接触抑制，未见重叠生长。眼睑CAFs的增殖速度明显快于NFs。RT-qPCR法检测眼睑CAFs中FAP mRNA相对于内参的表达量为3.672±0.221，明显高于眼睑NFs中的含量1.034±0.024(P<0.05)。Western Blot法检测两种细胞中FAP蛋白表达水平发现，FAP在眼睑CAFs中高表达，而眼睑NFs中几乎不表达(P<0.05)。

结论：眼睑CAFs与眼睑NFs在形态结构、生长增殖、生长因子表达等生物学特征方面发生了显著改变，提示肿瘤微环境随眼睑基底细胞癌的发生发展相应地发生了一些变化，进而诱导NFs生物学特性和功能发生改变，最终转变为CAFs。此外，眼睑CAFs较NFs高表达分泌FAP，表明肿瘤细胞微环境中FAP可能参与肿瘤的发生发展，与眼睑基底细胞癌浸润性生长有一定关系。

AIM: To investigate the role and significance of carcinoma-associated fibroblasts(CAFs)in the occurrence and development of eyelid basal cell carcinoma(BCC), and study on the biological characteristics and the expression of fibroblast activation protein(FAP)of two kinds of fibroblasts associated with eyelid BCC and normal eyelid skin fibroblasts(normal fibroblasts, NFs).

METHODS: CAFs and NFs were obtained by tissue primary cultured. The cell morphology of the 3^rd generation purified cells were observed under an inverted phase contrast microscope and identified them using their biomarker by immunohistochemistry for CK, VIM, α-SMA and FAP. Cell growth and proliferation were measured by MTT assay. The expression of FAP mRNA and protein in cells was detected by RT-qPCR and Western Blot.

RESULTS: The CAFs of the eyelid was long fusiform or spindle-shaped, with reduced cytoplasmic processes, more cytoplasmic granules, different cell sizes, disordered arrangement, overlapping growth, and loss of contact inhibition. NFs were in the form of extensive shuttle, radial arrangement, the cytoplasmic particles were rare, there was no overlapping growth phenomenon, and no contact inhibition. The proliferation rate of eyelid CAFs was faster than that of NFs. And the expression of FAP mRNA in CAFs was 3.672±0.221, which was significantly higher than that in NFs(1.034±0.024)(P<0.05). In addition, the expression level of FAP protein in CAFs was high, while NFs were not expressed(P< 0.05).

CONCLUSION: There were significant differences in the biological characteristics of CAFs and NFs, such as morphological structure, growth and the proliferation, growth factor expression and so on. It was therefore suggested that the tumor microenvironment of eyelid basal cell carcinoma had changed, and further induced the biological characteristics and function of NFs, and finally transformed into CAFs. In addition, eyelid CAFs expressed higher expression of FAP than NFs, indicating that FAP may be involved in the occurrence and development of tumor cells in tumor microenvironment, which is associated with the invasive growth of CAFs in eyelid.