Neuroprotective Effect On The Optic Nerve In Norwegian Brown Rat Model Of Optic Nerve Crush By Thermal Stimulius In Optic Nerve Head

【Objective】To explore the neuroprotective effect of subthreshold transpupillary thermotherapy(TTT) on optic nerve head of Norwegian Brown rat with partial optic nerve crush and the effect of Hsp60 expression on optic nerve head.This study provides methodological and theoretical basis for clinical application,and also explores new way to prevent and treat glaucoma and other neurodegenerative disease.【Methods】TTT was performed on the eyes of Norwegian Brown rats with an 810-nm diode laser(60 mW for 60 seconds with a 50-microm spot size),then optic nerve crush(ONC)model was made.Fluorogold was injected bilaterally into superior colliculi 5 days before retina flat-mounted(1W,2W,4W).The RGCs survival of control group,ONC group,sham operation group,TTT+ ONC group, TTT group was compared respectivly.As a result,the protective effect of subthreshold TTT on retinal ganglion cells(RGCs) in the rat with partial optic nerve crush can be verified.In order to test the effect of quercetin(Q,an inhibitor of Hsp erpression):In control groups, quercetin(4mg/kg) was injected intraperitoneally to inhibit the induction of heat shock proteins(Hsps) 6 hours before TTT in 7 consecutive days.The RGCs survival of TTT+ONC group,TTT+ONC+Q group,Q group, Saline group was compared respectively,to investigate inhabitation about neuroprotective of RGCs on ONC model.Meanwhile,immunohistochemical staining was performed to test strength and location of Hsp60 expression in optic nerve head.The injury caused by TTT irradiation by light microscope and transmission electron microscope was also observed.【Results】1 week after exposure with subthreshod TTT:The density of labeled RGCs was 2340.33±132.36 cells/mm~2 in control group,1336.17±110.30 cells/mm~2 in ONC group,2242.17±197.08 cells/mm~2 in sham operation group,1669±132.43 cells/mm~2 in TTT+ONC group,2373.33±174.43 cells/mm~2 in TTT group,1130.67±245.63 cells/mm~2 in TTT+ONC+Q group,2295±137.06 cells/mm~2 in Q group,2295±137.06 cells/mm~2 in Saline group,respectively;There was significant difference in density of labeled RGCs between TTT+ONC group and ONC group (t=-7.65,P=0.001).There was significant difference in density of labeled RGCs between TTT+ONC+Q group and TTT+ONC group(t=3.876,P=0.012).The density of labeled RGCs in TTT+ONC+Q group was lower than that of TTT+ONC group.There was no significant difference in density of labeled RGCs between TTT+ONC+Q group and ONC group(t=1.529,P=0.187).2 weeks after exposure with subthreshod TTT:The density of labeled RGCs was 2307.17±153.42 cells/mm~2 in control group,984.00±128.02 cells/mm~2 in ONC group,2246.67±182.71 cells/mm~2 in sham operation group,1299.50±237.63 cells/mm~2 in TTT+ONC group,2295±161.72 cells/mm~2 in TTT group,477.17±162.3 cells/mm~2 in TTT+ONC+Q group,2273.40±118.97 cells/mm~2 in Q group, 2307.80±103.74 cells/mm~2 in Saline group,respectively;There was significant difference in density of labeled RGCs between TTT+ONC group and ONC group(t=-2.910,P=0.044).The density of labeled RGCs in TTT+ONC group was higher than that of ONC group.There was significant difference in density of labeled RGCs between TTT+ONC+Q group and TTT+ONC group (t=6.126,P=0.002).The density of labeled RGCs in TTT+ONC+Q was lower than that of TTT+ONC group.There was significant difference in density of labeled RGCs between TTT+ONC+Q group and ONC group(t=8.553,P=0.000).The density of labeled RGCs in TTT+ONC+Q group was lower than that of ONC group.4 weeks after exposure with subthreshod TTT:The density of labeled RGCs was 2282.17±121.53 cells/mm~2 in control group,824.00±244.57 cells/mm~2 in ONC group,2238.5±119.10 cells/mm~2 in sham operation group,1091.67±115.03 cells/mm~2 in TTT+ONC group,2259.67±162.16 cells/mm~2 in TTT group,353.83±102.96 cells/mm~2 in TTT+ONC+Q group,2257.40±118.25 cells/mm~2 in Q group, 2306.80±122.89 cells/mm~2 in Saline group,respectively;There was significant difference in density of labeled RGCs between TTT+ONC group and ONC group(t=-2.889,P=0.045).The density of labeled RGCs in TTT+ONC group was higher than that of ONC group.There was significant difference in density of labeled RGCs between TTT+ONC+Q group and TTT+ONC group (t=20.624,P=0.000).The density of labeled RGCs in TTT+ONC+Q group was lower than that of TTT+ONC group.There was significant difference in density of labeled RGCs between TTT+ONC+Q group and ONC group(t=4.552,P=0.006).The density of labeled RGCs in TTT+ONC+Q group was lower than that of ONC group. 24h,72h,1W after TTT,immunohistochemical staining showed strongly positive stain in optic nerve head and the highest positive stain at 72h after TTT.Light microscope and transmission electron microscope showed that there was limited damage in periphery retina and the optic nerve head.【Conclusions】TTT has protective effect on retinal ganglion cells in the BN rat with partial optic nerve crush.TTT may initiate the endogenous protective mechanism by inducing the overexpression of Hsp60 in optic nerve head,to inhibit the secondary degeneration of retinal ganglion cells.