Bikbova Guzel, Oshitari Toshiyuki and Yamamoto Shuichi
Department of Ophthalmology and Visual Science, Chiba University Graduate School of Medecine, 1-8-1 Inohana, Chuo-ku, Chiba 2608670, Japan
Purpose: To invent the combined therapeutic agents with neuroprotective abilities for diabetic retinopathy.
Methods: All of the procedures were performed in accordance with the ARVO Statement for the Use of Animals in Ophthalmic and Vision Research. Retinal explants of 7 adult SD rats were three-dimensionally cultured in collagen gel, and incubated in serum free media, AGEs, AGEs+100=µM citicoline, AGEs+10ng/ml NT-4, AGEs+100µM TUDCA, AGEs+100µM citicoline + TUDCA (doublet), AGEs+100µM citicoline+TUDCA+10ng/ml NT-4 (triplet). The number of regenerating neurites was counted under a phase-contrast microscope after 7 days of culture. After counting, retinal explants were fixed, cryosectioned, and stained by TUNEL and DAPI. The ratio of TUNEL-positive cells to the number of DAPI-staining nuclei in the ganglion cell layer was calculated. Statistical analyses were performed by one-way ANOVA.
Results: In retinas incubated with triplet, the number of neurites was significantly increased more than other groups (P < 0,0001) and the number of TUNEL-positive cells was fewer than other groups (P<0.0001). In retinas culture in doublet, the number of neurits was more than that of single treatment groups (P < 0,0001) and the number of TUNEL-positive cells were fewer than of singe treatment group (P < 0,0001)
Conclusions: Doublet (without NT-4) and triplet (with NT-4) significantrly increased the number of regenerated neurits and decreased the number of TUNEL positive cells, thas they may be considered as promising agents for neuroprotective and regenerative therapy for diabetic retinopathy.