Authors: Ke Meiqing, Zhang Xingru, Zhou Guizhen, Ma Yanchun, Li Qingsong, Xiang Minhong
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Background
Conjunctivochalasis (CCh) is a common age-related ocular surface diseases.Researches showed that the inflammatory factors are upregulated in conjunctiva and tear of CCh patients, inferring inflammation participates in the pathogenesis of CCh, however, its mechanism is unclear now.Studies also showed that the expression of matrix metalloproteinases (MMPs) in CCh conjunctiva is elevated.However, the association between inflammatory factors and MMPs is unknown.
Objective
This research was to observe the effects of tumor necrosis factor-α (TNF-α) and interleukin-1β (IL-1β) on the expression of MMP-1, MMP-3, tumor necrosis factor-stimulated gene-6(TSG-6) and pentraxin-3 (PTX3) in cultured CCh fibroblasts.
Methods
Conjunctival fibroblasts were isolated and cultured from CCh-derived and cataract-derived human conjunctiva explants.The cells were treated using 20 ng/ml PBS, TNF-α or IL-1β for 4 hours, respectively.The expression levels of MMP-1, MMP-3, TSG-6 and PTX3 genes and proteins were detected by quantitative real time PCR and Western blot assay, respectively.
Results
The second-generation cells showed a long and fusiform shape with ovoid nucleus and radial agreement, and the cell processors connected each other.The differences of the relative expression levels of MMP-1, MMP-3, TSG-6 and PTX3 mRNA in the cells were significantly different between CCh group and cataract group after being treated by PBS, TNF-α and IL-1β (MMP-1 mRNA: Fgroup=2.611, P=0.116; Fintervene=161.564, P=0.000; MMP-3 mRNA: Fgroup =5.201, P=0.029; Fintervene =211.021, P=0.000; TSG-6 mRNA: Fgroup =47.209, P=0.000; Fintervene=119.340, P=0.000; PTX3 mRNA: Fgroup =40.512, P=0.000; Fintervene =93.935, P=0.000). Compared with the cataract group with PBS treatment, the expression levels of MMP-1, MMP-3, TSG-6 and PTX3 mRNA were significantly elevated in the CCh group with PBS treatment or the cataract group with TNF-α and IL-1β treatment (all at P<0.05). Compared with the CCh group with PBS treatment, the expression levels of MMP-1, MMP-3, TSG-6 and PTX3 mRNA were significantly elevated in the CCh group with TNF-α and IL-1β treatment (all at P<0.05). The differences of the relative expression levels of MMP-1, MMP-3, TSG-6 and PTX3 proteins in the cells were significantly different between CCh group and cataract group after being treated by PBS, TNF-α and IL-1β (MMP-1: Fgroup=84.702, P=0.000; Fintervene=48.900, P=0.000; MMP-3: Fgroup=112.818, P=0.000; Fintervene=194.980, P=0.000; TSG-6: Fgroup=56.867, P=0.000; Fintervene=70.356, P=0.000; PTX3: Fgroup=1.488, P=0.231; Fintervene=89.872, P=0.000), and the expression changes of MMP-1, MMP-3, TSG-6 and PTX3 proteins were coincident with the genes among the groups with various treatments.
Conclusions
The expressions of MMP-1, MMP-3, TSG-6 and PTX3 in conjunctival fibroblasts are upregulated in CCh eyes.The interaction of TNF-α and IL-1β with MMPs is probably involved with the pathogenesis and development MMPs probably.