The Pretreatment With Recombinant Human Slit2 Protein Relieves Endotoxin-induced Uveitis In Rat

Objective:To investigate the effect of recombinant human Slit2 protein on endotoxin-induced uveitis in rat.Methods:1.Exploring rhSlit2 administration methods and concentrations.2.Rh Slit2(10 ng/eye,30 ng/eye,100 ng/eye)was injected into the vitreous cavity of rats,respectively.After 24 hours,2 g/L lipopolysaccharides was injected into the rat posterior foot pads(200 ug/rat),respectively.24 hours later,the clinical scores were determined according to the inflammation of the anterior chamber.3.The protein concentrations and the total number of cells in aqueous humor were detected.4.The ocular morphological changes were observed by HE staining after 24 hours of LPS injection.5.Real-time qPCR was used to detect the expression of inflammatory factors.The protein levels of inflammatory factors were measured by Western blot.6.The protein levels of Akt and P-Akt were measured by Western blot.Results:1.Among the methods of administration explored in this study,intravitreal injection of rh-Slit2 is the best method;LPS+rhSlit2(100ng/eye)group has a significant inhibitory effect on anterior chamber inflammation.2.The clinical scores of LPS+rh Slit2(100 ng/eye)group(1.40±0.84)was significantly lower than LPS+PBS group(5.60±0.97,P<0.01).3.In the LPS+PBS group,a large number of cellulose-like exudates were found in the anterior chamber.Compared with the LPS+PBS group,the cellulose-like exudation of LPS+rhSlit2(100 ng/eye)group in the anterior chamber decreased,and the number of inflammatory cells around the ciliary body and intravitreal cavity were decreased.4.The number of intraocular inflammatory cells of LPS+rh Slit2(100ng/eye)group[(65.21±18.38)×10~6/ml]was lower than LPS+PBS group[(319.60±28.88)×10~6/ml](P<0.01),and the protein concentrations of aqueous humor also got the same trend(P<0.01);.5.The mRNA levels of IL-6(19.69±5.28)and TNF-α(1.78±0.33)in LPS+rh Slit2(100 ng/eye)group were lower than LPS+PBS group[IL-6(77.09±12.61),TNF-α(4.06±0.58)](P<0.01).The protein levels of IL-6(1.92±0.60),TNF-α(1.33±0.11);The protein levels of IL-6,TNF-αalso got the same trend(P<0.01).6.The protein levels of P-Akt(0.18±0.37)in LPS+rhSlit2(100 ng/eye)group were lower than LPS+PBS group(P<0.01).Conclusion:The intravitreal injection of rhSlit2 can ameliorate the inflammation of endotoxin-induced uveitis in SD rats.RhSlit2 may inhibit LPS-induced inflammatory by inhibiting the activation of the PI3K/Akt signaling pathway.