Axonal Protective Effect Of PEG On Crushed Optic Nerves In Adult Rats

Classically, axonal degeneration has been regarded as a passive process which is mainly resulting from the lack of nutrition from the soma and a nonreversible primary lesion process which is no way to rescue. But recently it has been recognized that with proper interference axons can survive. Polyethylene glycol, a hydrophilic polymer, has been shown to repair cell memo- brines and inhibit free radical production in vitro and in vivo. Immediate PEG administration can promote the functional recovery in various models of brain ischemia and trauma and this is believed to be due to its significant neuroprotective effect.In part one, we established a new rat optic nerve model suitable for investigating Wallerian degeneration in the central nervous system (CNS). A pair of forceps with a width of 1 mm and clipping pressure of 148 g was used to crush the exposed left optic nerves (ON) of 12 adult female SD rats for 10 s at 2 mm from the optic disc. The animals were randomly divided into 2 groups (n=6 for each group) after surgery. Six animals received administration of retrograde fluorescent dye, FluroGold (FG), onto the bilateral superior colliculi immediately after ON crush, and survived for 72 h. The retinas were removed for flat-mount and ONs were cut for cryostat serial sections after these animals were killed. The other 6 animals were perfused 72 h after ON crush and the ON were cryo-sectioned forβ-tubulin and NF-M immuno-staining and HE histological staining, respectively. Three days after the left ON crush, no FG- labeled retinal ganglion cells were detected in the left retina. A lesion zone of 1 mm width appeared within the crushed ON, and the continuity of the ON matrix was maintained. Most FG-labeled and NF-M immune positive axons were interrupted within the distal segment of the crushed ON where most ?-tubulin immune positive structures were destroyed. In the present results indicate that the crushed axons are totally separated after ON crush but with maintained continuity of the ON matrix, and classic Wallerian degeneration take place in the distal segment of crushed ON. Thus, we have established a simple and pragmatic rat model for the study of Wallerian degeneration in the CNS and its treatment.In part two, we studied axonal protective effect of PEG on crushed optic nerves in adult rat. The left ON in the subjected rat was crushed at 2 mm from the optic disc and PEG or PBS was administered to the lesion site immediately after the lesion. The RGCs were retrogradely labeled with a gold fluorescent dye(FG) by superior colliculi (SC) administration and the number of functional RGCs (which is still connected to SC) was calculated in the wholemounted retinas. The optic nerve is stained by NF 160 respectively and the number of stained fibers in the different distance to the lesion site was calculated and compared. There were no functional RGCs at 3 days after ON crush (ONC) in both the control and PEG animals. In control animals, the number of functional RGCs of the whole retina was 222+91.09/retina at 7 days and 523.6+51.92/Retina at 14 days after ON crush. Immediate PEG administration significantly preserved the functional RGCs 449.25+36.25/retina at 7 days and 896+16.67/Retina at 14 days. 3 days after ONC, at 500μm the proximal nerve and 500μm the distal nerve and 250μm the distal nerve part from the crushed site, PEG treated (18.00±2.17, 19.00±3.24 and 17.45±2.17, p>0.05) when compared with those in PBS treated (12.95±3.97, 17.26±1.97 and 13.28±6.94) control animals; at 250μm the proximal nerve part from the crushed site, PEG treated (12.85±4.09, p<0.01) when compared with those in PBS treated (6.38±3.92) control animals. 7 days after ONC, at 500μm the proximal nerve and 500μm the distal nerve and 250μm the distal nerve part from the crushed site, PEG treated (21.98±1.52, 18.77±2.58 and 18.90±3.72, p<0.01) when compared with those in PBS treated (13.79±5.09, 13.67±3.19 and 12.63±3.91) control animals; at 250μm the proximal nerve part from the crushed site, PEG treated (17.98±2.15, p<0.05) when compared with those in PBS treated (12.94±7.16) control animals. 14 days after ONC, at 500μm the proximal nerve and 250μm the proximal nerve part from the crushed site, PEG treated(11.50±1.73 and 14.33±4.05, p<0.01)when compared with those in PBS treated (6.94±1.05 and 6.17±2.13)control animals; at 500μm the distal nerve part from the crushed site, PEG treated(13.67±2.37, p<0.05)when compared with those in PBS treated (10.16±3.03)control animals; at 250μm the distal nerve part from the crushed site, PEG treated(11.53±4.26, p>0.05)when compared with those in PBS treated (9.72±3.14)control animals. The result of fibers at different time but at the same site as follows: at 500μm the proximal nerve part from the crushed site, 3 days after ONC, PEG treated (18.00±2.17, p>0.05) when compared with those in PBS treated (12.95±3.97) control animals; 7 days and 14 days PEG treated (21.98±1.52 and 11.50±1.73, p<0.01) when compared with those in PBS treated (13.79±5.09 and 6.94±1.05) control animals. at 250μm the proximal nerve part from the crushed site, 3 days and 14 days after ONC, PEG treated(12.85±4.09 and 14.33±4.05, p<0.01 )when compared with those in PBS treated (6.38±3.92 and 6.17±2.13) control animals; 7 days PEG treated(17.98±2.15, p<0.05)when compared with those in PBS treated (12.94±7.16) control animals. at 250μm the distal nerve part from the crushed site, 3 days and 14 days after ONC, PEG treated(17.45±2.17 and 11.53±4.26, p>0.05 )when compared with those in PBS treated (13.28±6.94 and 9.72±3.14) control animals; 7 days PEG treated(18.90±3.72, p<0.01)when compared with those in PBS treated (12.63±3.91) control animals. at 500μm the distal nerve part from the crushed site, 3 days after ONC, PEG treated (19.00±3.24, p>0.05) when compared with those in PBS treated (17.26±1.97) control animals; 7 days after ONC, PEG treated (18.77±2.58, p<0.01) when compared with those in PBS treated (13.67±3.19) control animals; 14 days after ONC, PEG treated (13.67±2.37, p<0.05) when compared with those in PBS treated (10.16±3.03) control animals. So, we think that PEG significant promotes the survival of the nerve fibers.Our study indicates that PEG has the neuroprotective effect on promoting the survival of the RGCs and axons.