Protective Effect Of α-melanocyte Stimulating Hormone On Ocular Surface Of Scopolamine-induced Dry Eye Rat Model

Background Dry eye is a multifactorial disease of the tears and ocular surface that results in symptoms of discomfort,visual disturbance,and tear film instability with potential damage to the ocular surface.It is accompanied by increased osmolarity of the tear film and inflammation of the ocular surface.Previous studies have identified the inflammation plays a key role in the occurrence and development of dry eye.Thereby,the inhibition of inflammation is very important for dry eye treatment.a-MSH is a 13-aa-long neuropeptide.Both a-MSH and adrenocorticotropin(ACTH)are derived from the post-translational processing of the pro-opiomelanocortin(POMC)molecμLe.And a-MSH corresponds to the first 13 amino acids of the ACTH sequence,so that they are same in some effects.Clinical studies indicated that topical ACTH is an effective treatment option for patients suffering from keratoconjunctivitis sicca.It also has been demonstrated that a-MSH has a strong anti-inflammatory effect and plays a vital role in the corneal epithelium wound healing.Objective To investigate the therapeutic effects and possible mechanisms of Alpha-Melanocyte Stimulating Hormone(α-MSH)on the rat dry eye model induced by scopolamine.Methods The eye drop containing a-MSH was topically administered(0.2,2,and 20 ng per day)on a scopolamine-induced dry eye model.The following clinical indications of dry eye were evaluated on day 0,7,14,21,and 28:tear break-up time(BUT),corneal fluorescein staining,and tear volume.Global specimens were collected on day 28 and then the following examinations were performed:histologic investigation,and periodic acid-schiff(PAS)assay to detect goblet cells.The levels of IL-1β and TNF-a of the corneas were also measured by RT-PCR.Results1.Tear film break-up time Since the 7th day.variance analysis shows that the BUT among the five groups was statistically significant.seventh day.the BUT of the 10-4mg/ml α-MSH was significantly longer compared with 0.9%NaCl solution(P = 0.021).On the 14th day,in addition to no differences between 0.9%NaCl solution and 10-5mg/ml α-MSHtreatment group.any two groups of the rest were statistically different.On the 21st day.it is very different between the three different concentrations of α-MSH treatment group with the control group and 0.9%NaCl-treated group.On the 28st day,in addition to no differences between 0.9%NaCl solution and 10-5mg/ml α-MSH treatment group(P=0.239),10-4mg/ml α-MSH group and 10-3mg/ml α-MSH group(P=0.553).any two groups of the rest were statistically different.2.The tear volumeVariance analysis shows the amount of tear secretion of the five groups has no difference each time point.On the 14th day.the tear volume of the 10-4mg/mlα-MSH-treated group which is(4.88 ±0.74)mm,did not increase significantly compared with the control group(P=0.355);10-4mg/ml α-MSH group and 10-3mg/mlα-MSH group secreted more tear than the 0.9%NaCl-treated group(P=0.023,P=0.046).Only the normal group and the four treated group were different(both P=0.000).On the 28th day,except for the 10-5mg/ml α-MSH group and group,the rest were all different.3.Fluorescein StainingWe can see obvious corneal fluorescein sodium coloring in 0.9%NaCl-treated eyes.It performanced as densely punctate,even small pieces;while administration of α-MSH resulted in its decrease compared with 0.9%NaCl-treated group.And with the increase of the concentration of a-MSH,it is more apparent,almost close to the normal control group.4.Tear ferning testSince the 7th day,the tear ferns and its branch of group decreased.While at the same time point,ferns form α-MSH group is more and better.5.HE stainingHE staining revealed that α-MSH-treated corneas had smoother epithelium compared with the NS-treated group.Morphologically,there were noticeable epithelial hyperplasia,cellular level increased,and basal cell edema and disorganized in 0.9%NaCl solution-treated eyes that were not found in α-MSH treated eyes.6.Periodic acid-Schiff staining Goblet cell number was decreased in the condition of dry7 eye,while increased to a similar level to the naive ones when given a-MSH treatment in a dose-dependent manner.7.Real-time PCR showed α-MSH can inhibit the expression of inflammatory cytokines,such as TNF-a mRNA and IL-1 β mRNA in rat cornea.Conclusions Taken together,topical application of a-MSH presented clinical improvements on dry eye by promoting the secretion of tears,stabilizing the tear film,maintaining the integrity of epithelium,increasing the number of goblet cells,and alleviating the ocular surface inflammation.And the concentration of the 10-4 mg/ml a-MSH may be the best choice.The results indicate that scopolamine-induced dry eye model is valuable for dry eye study and a-MSH has potential as a therapeutic agent in clinical treatment of dry eye.