Neuroprotective Effect Of Methyl 3,4-dihydroxybenzoate On Retinal Ganglion Cells In Rats And Its Mechanisms

Background : Optical nerve injury is a kind of diseases that companied with the pathological features of retinal ganglion cells(RGCs)apoptosis and optic nerve atrophy.It can eventually lead to blind.At present,the most common drugs used in clinical such as glucocorticoid,ganglioside,vitamin E and so on.They have good effects on optic nerve injury,but also accompanied by different levels of toxic and side effects,and their curative effects have yet to be improved.It has recently been reported that certain traditional Chinese medicine or its extracts have the effect of improving eye blood flow,prolonging the neurons survival time and promoting neurite growth,which may be superior to traditional therapeutic drugs for optic nerve injury.In recent years it has been reported that some traditional Chinese medicine or natural extracts can improve eye blood flow and promote neurite growth.Methyl 3,4-dihydroxybenzoate(MDHB)is a monomer extracted from Kalimeris indica(Linn.)Sch.or Hedyotis diffusa Willd.The early experiment results showed that MDHB in vitro could protect cortical neurons from A? toxicity,but also protect RGC-5 cells from damages by antioxidant,anti-apoptosis and improving mitochondrial membrane potential.Objective: To study the protective effects of MDHB on N-methyl-D-aspartate(NMDA)induced damage of retinal ganglion cells in rat and its mechanisms.Material and Methods: MDHB was injected intraperitoneally into adult SD rats,detecting the concentrations of MDHB through the blood-ocular barrier into the retina at the end of the second minute and the thirtieth minute.The experimental rats were divided into 6 groups: blank control group,model group,positive drug group(memantine,10mg/kg),MDHB low dose(25mg/kg)group,MDHB middle dose(50mg/kg)group and MDHB high dose(100mg/kg)group.Except the rats of blank control group,the other rats were anaesthetized through chloral hydrate,intravitreally injected with NMDA to cause retinal ganglion damage.The rats in the model group were injected intraperitoneally with normal saline,the positive drug group were administered by memantine(10mg/kg),all the MDHB groups were administered with MDHB(25mg/kg,50mg/kg,100mg/kg,respectively),and every group was injected intraperitoneally for 5 consecutive days.The vision of the rats was testesd by black-white box,the changes of visual electrophysiology were recorded by electroretinogram(ERG),the hematein eosin(HE)staining and immunohistochemistry with the Brn-3a antibody were performed to the count of retinal ganglion cells,the Western blot analysis was used to detect Bax,Bcl-2,Cleaved caspase-3,Caspase-3,Akt,and BDNF.Results: 1.MDHB can rapidly penetrate the blood-ocular barrier and reach the retina,after about 2minutes it can be detected in the retina via HPLC.2.The result tested by black and white box showed that,the MDHB low dose(25mg/kg)group and middle dose(50mg/kg)group could improve the visual ability of the rats(P<0.01)compared with the model group.HE staining and immunohistochemistry showed that the MDHB middle dose(50mg/kg)group could keep RGCs alive(P<0.05).The results of ERG showed that MDHB could enhance the photoreaction of RGCs(P<0.05).3.Western blot analysis showed that the ratio of Bax/Bcl-2,Celaved caspase-3/Caspase-3 in MDHB middle dose(50mg/kg)and MDHB high dose(100mg/kg)group decreased(P<0.05),and the ratio of p-Akt/Akt increased(P<0.05).Conclusion: MDHB can alleviate NMDA-induced retinal ganglion cell damage and improve the visual behavior of experimental animals.The action mechanisms of MDHB may be related to its anti-apoptotic and neurotrophic effects.MDHB may become a new drug in optic nerve injury treatment.