关键词: 成纤维细胞, Graves眼病, 中药疗法, 基质金属蛋白酶类, CD40L

Abstract: Background　Regarding thyroid-associated ophthalmopathy（TAO），although modern medicine has made some progress in basic and clinical research，and accumulated a certain amount of materials and evidence on pathogenesis，there are still few studies on TAO treatment，and there are still a lack of effective therapeutic methods and drugs in clinical practice.Furthermore，for TAO，TCM-based syndrome differentiation and treatment has a certain therapeutic effect with few side effects and low recurrence rate，but the mechanism of action is not clear，and the target of drug action is unclear.Objective　To observe the effects of CD40L on the expression of MMP-1，MMP-2，TIMP-1，TIMP-2 on the surface of retroocular fibroblasts（RFs） and the intervention results of three TCM composite drugs.Methods　In this experiment，the serum contained in three TCM composite drugs（xiehuopingtusan，huoxuepingtusan and yangmupingtusan） was derived from 35 male SPF SD rats in September 2016，and the human retrobulbar connective tissue was derived from two patients with ocular trauma（from Department of Ophthalmology，the First Affiliated Hospital of Zhengzhou University in 2014）.Successfully cultured RFs were randomly divided into 7 groups：blank group（cultured in a normal medium），serum group（cultured in a medium containing 10% normal rat serum），CD40L group（cultured in a culture medium containing 100 ng/ml CD40L），CD40L+serum group（cultured in a medium containing 100 ng/ml CD40L and 10% normal rat serum），xiehuopingtusan group（cultured in a medium containing 100 ng/ml CD40L and 10% xiehuopingtusan-containing rat serum with），huoxuepingtusan group（cultured in a medium containing 100 ng/ml CD40L and 10% huoxuepingtusan-containing rat serum），and yangmupingtusan group（cultured in a medium containing 100 ng/ml CD40L and 10% yangmupingtusan-containing serum）.Immunocytochemical staining was used to detect the expression of MMP-1，MMP-2，TIMP-1 and TIMP-2 in each group.Results　The activities of MMP-1 and MMP-2 in CD40L group were lower than those of the blank group（P<0.05）.The activities of MMP-1 and MMP-2 in CD40L+serum group were lower than those of serum group，and of the three TCM composite drug groups（P<0.05）.The activities of MMP-1 and MMP-2 in yangmupingtusan group were higher than those in xiehuopingtusan group and huoxuepingtusan group（P<0.05）.The activities of TIMP-1 and TIMP-2 in CD40L group were higher than those in blank group（P<0.05）.The activities of TIMP-1 and TIMP-2 in CD40L+serum group were higher than those in serum group，and those in three TCM composite drug groups（P<0.05）.The activities of TIMP-1 and TIMP-2 in yangmupingtusan group were lower than those in xiehuopingtusan group and huoxuepingtusan group（P<0.05）.Mean ratios of MMP-1/TIMP-1 and MMP-2/TIMP-2 in the CD40L group were lower than those in the blank group（P<0.05）.Mean ratios of MMP-1/TIMP-1 and MMP-2/TIMP-2 in CD40L+serum group were lower than those in serum group（P<0.05）.Mean ratios of MMP-1/TIMP-1 and MMP-2/TIMP-2 in three TCM composite drug groups were higher than those in CD40L+serum group（P<0.05）.Mean ratios of MMP-1/TIMP-1 and MMP-2/TIMP-2 in yangmupingtusan group were higher than those in the xiehuopingtusan group and huoxuepingtusan group（P<0.05）.Conclusion　The treatment of TAO by TCM composite drugs may be achieved by blocking the CD40-CD40L co-stimulatory pathway，subsequently leading to the imbalance of MMP/TIMP.

Key words: Fibroblasts, Graves ophthalmopathy, Drug therapy（TCD）, Matrix metalloproteinases, CD40L