Nerve Growth Factor Promotes The Repair Of Corneal Nerves In Diabetic Rats

Objective:To explore the effect of nerve growth factor(NGF)on corneal nerves of diabetic rats,and to explain the molecular mechanism.Methods:Type 1 diabetes model rats were induced by intraperitoneal injection of streptozotocin(STZ).Six weeks after the model were successfully established,corneal sensitivity were tested between the control and diabetic rats,then the left or the right eyes of control and diabetic rats were made as Ctrl + BSA group or Ctrl +NGF group and DM + BSA group or DM + NGF group.Based on the groups,the eyes of four groups were received topical application of BSA or NGF eye drops(15ug/mL)three times a day(9 am,13 pm and 17 pm)for two weeks,and then the corneal sensitivity and part of corneal nerves density were detected,in the next,the same area of epithelium of remaining corneas were scraped and continued eye drops for one week,during the week,the repaired area of corneal epithelium were measured through Sodium fluorescein staining in 0,24,36,48hours after scraped and the corneal sensitivity were measured in 2,4 and 7 days after scraped.At last,the cornea,corneal epithelium,corneal stroma and trigeminal ganglion were separated from rats,Immunofluorescence staining was used to evaluate the corneal nerves density and the co-expressing of TrkA and corneal nerves,ELISA was used to test the level of beta NGF in corneal epithelium,stroma of cornea and trigeminal ganglion,Western blot was used to test the TrkA,p75NTR,?-tubulin-III,Akt and p-Akt in corneal stroma or trigeminal ganglion.Results:The corneal sensitivity of diabetic rats was lower than that of Ctrl rats in the sixth,eighth and ninth week,and the repair rate of corneal sensitivity were also slower,however,topical application of NGF eye drops could promote the repair of corneal sensitivity in diabetic rats.The central corneal nerves density of DM + BSA group were lower than that of Ctrl + BSA group and the density of DM + NGF group was higher than that of DM + BSA group in the eighth and ninth week.The repair rate of corneal epithelium in diabetic rats was also slower than that of control rats,topical application of NGF eye drops could promote the repair of corneal epithelium in control and diabetic rats.The levels of NGF-? were not significantly different in the corneal epithelium,corneal stroma and trigeminal ganglion between DM + BSA group and Ctrl + BSA group,but the level in the groups treated with NGF eye drops were higher in corneal epithelium and corneal stroma than that treated with BSA eye drops.The levels of TrkA in the corneal stroma of diabetic rats were lower than that of control rats;what is more,the level in corneal stroma was much lower when treated with NGF,but the level in trigeminal ganglion was increased.Akt and p-Akt were down regulated in the corneal stroma of DM + BSA group,compared with them in Ctrl + BSA group,treated with NGF can promote them increased,and p75NTR,?-tubulin-III,Akt and p-Akt were no significant difference in trigeminal ganglion.Conclusion:Corneal nerve density,corneal sensitivity and corneal epithelial wound healing were decreased in diabetic rats,NGF can promote the repair of corneal nerve of diabetic rats,and then improved diabetic keratopathy,the mechanism of NGF repairing corneal nerves is associated with NGF-TrkA-Akt signaling pathway.