Experimental Study On The Early Expression Of Surfactant Proteins A And D In Fusarium Solani Infected Cornea

Purpose:To investigate the early expression of surfactant proteins A and D in Fusarium Solani infected cornea and to explore the role of surfactant proteins A and D in innate immunity of fungal keratitis.Methods:Eighty Wistar rats were divided into four groups randomly. They are control group, simple injury group, fungal keratitis group 1 and fungal keratitis group 2. The right eye were taken as experimental eye. Control group has five rats with no treatment. Twenty five rats in Simple injury group were not inoculated with Fusarium solani. Twenty five rats in fungal keratitis group 1 and twenty five rats in fungal keratitis group 2 were taken as fusarium solani keratitis model. Five rats in simple injury group, fungal keratitis group 1 and fungal keratitis group 2 were killed at 6h,12h,24h,48h and 96h respectively after the experimental model being establised. We observed the eye changes of the rats under slit lamp microscope, the corneal tissue inflammation by histopathological slices (To take the right cornea in control group, simple injury group and fungal keratitis group 1. To take the left cornea in fungal keratitis group 2). We also detected the SP-A, SP-D mRNA expression of cornea tissue by reverse transcriptase polymerase chain reaction (RT-PCR) and SP-A, SP-D protein expression and co-expression intensity of cornea tissue by immunohistochemical method.Results:(1) Slit lamp examination:The result of 6h fungal infected corneal shew, palpebral conjunctival hiperemia(+) and no obvious edema on corneal. There was no significant differences compared with simple injury group. The result of 12h fungal infected corneal shew, palpebral conjunctival hiperemia(++) and edema appeared on corneal. The result of 24h fungal infected corneal shew, palpebral conjunctival hiperemia(+++), and ground glass-like edema appeared on corneal. The result of 48h and 96h fungal infected corneal shew, obvious edema on corneal, yellow-white ulcers in different sizes and immune ring formed. (2) Histopathological examination:The result of 6h fungal infected corneal shew that, central corneal epithelium was absent, collagen fibers were arranged regularly, no significant neutrophil infiltration. There was no signifiant difference compared with simple injury group. The result of 12h fungal infected corneal shew few neutrophil infiltration in shallow stromal layer. The result of 24h fungal infected corneal shew that, edema appeared on corneal, central epithelium of corneal and Bowman's layer disappeared, collagen fibers were irregularly arranged at stromal layer, and neutrophil infiltration increased in shallow stromal layer. The result of 48h and 96h fungal infected corneal shew obvious edema on corneal, corneal stroma were more disordered, massive neutrophil infiltration appeared in stronal layer. (3) Immunohistochemistry:There was low SP-A protein and SP-D protein expression in corneal epithelial cells of normal rats, simple injury group and fungal keratitis group 2. The expression of the three groups had no significant difference (p> 0.05). The SP-A protein and SP-D protein expression was not increased significantly in corneal epithelial cells of 6h fungal infected corneal. The expression has no significant difference compared with normal cornea, simple injury group and fungal keratitis group 2(p> 0.05). The expression of 12h fungal infected corneal was increased gradually. It was highest at 48h, and decreased at 96h. The expressions at different time were in significant difference(p <0.01), and the synchronous expression of control group, simple injury group and fungal keratitis group 2 were also in significant difference (p<0.01). (4) RT-PCR detection:The SP-A mRNA, SP-D mRNA expression were low in the corneal of normal rats, simple injury group and fungal keratitis group 2. There were no significant difference (p> 0.05) among the three groups. The SP-A mRNA, SP-D mRNA expression of fungal infected corneal increased gradually. It was highest at 24h. The expression at 48h and 96h were decreased. The expressions at different time were in significant difference(p<0.01), and the synchronous expression of control group, simple injury group and fungal keratitis group 2 were also in significant difference (p<0.01).Conclusion:The SP-A, SP-D expression in the corneal tissue is significantly increased at the early pioried of fusarium solani injection on the cornea. The expression is decreased along with the progress of the disease, but it is still higher than the expression of simple injury group. This result presented that SP-A, SP-D may be the pattern recognition receptor of Fusarium solani. Therefor, SP-A and SP-D may play a role in the early innate immunity response of the corneal resistance to Fusarium solani infection.