| Background: Diabetic retinopathy (DR) becomes one of the leading causes for the visual loss in recent years. However the mechanism of pathogenesis of DR is still not well-elucidated. DR is usually considered to be a disease of the retinal blood vessels. But some investigations indicated that microvasculopathy may not be the sole cause for DR. Diabetic neuropathy may play an important role in the pathogenesis of DR. The abnormalities in electroretinogram (ERG) have been reported to precede retinal vasculopathy or accompany early vascular lesions. The results from our laboratory's research on the STZ induced diabetic rats also indicated the changes of ERG in the early stage of diabetes. The abnormalities in ERG appeared 3 days after the onset of diabetes and became most obvious 2 weeks later. Subsequent studies also showed that an obvious breakdown of blood retinal barrier (BRB) could been observed 2 weeks after the onset of diabetes. Retinal Mtlller cells (RMCs), as the principal glial cells of retina, are very important for the maintenance of retinal normal structure, metabolism, and function. They can also modulate the function of BRB with their processes wrapping the capillary vessels. The abnormalities of ERG and BRB are relevant to RMCs. This indicates that in the early stage of diabetes, hyperglycemia, it's relative products and other cytokines may affect RMCs which dysfunction may damage both retinal neurons and vascular cells directly or indirectly under the diabetic glycemia. RMCs might be involved in thepathogenesis of DR. This research, based on our laboratory's former study on STZ-diabetic rats, put emphasis on the relationship between RMCs and vascular endothelial cells in incipient DR so as to provide some experimental bases for :idating the pathogenesis of DR.Aims: (1) To observed the effects of high glucose and serum from ratwith incipient diabetes on the proliferation of cultured RMCs in vitro; (2) Toevaluate the effects of high-glucose-cultured RMCs conditioned media(RMCs-CM) and cocultured RMCs in high glucose on the viability of bovineaortal endothelial cells (BAECs ) .Methods: (1) The RMCs were cultured for 72 h in DMEM whichcontains different concentrations of glucose (5.5, 15, 30, and 60mM) with themannitol as a hypertonic control. A tetrazolium (MTT) reduction method wasused to evaluate their viability by measuring the absorbance (A value) ; (2) TheMTT method was used to evaluate the viability of the RMCs cultured in DMEMwith different concentrations (50, 100, and 200 mL. L-1) of sera from rats with 2weeks of STZ-induced diabetes; (3) The RMCs were cultured for 48 h indifferent concentrations of glucose (5.5, 15, 30, and 60mM) of DMEM, and thesupernatants were collected, filtered and stored as conditioned medium(RMCs-CM) . Then the RMCs-CM diluted with DMEM (low glucose) indifferent concentrations (250, 500, and 1000 mL.L-1) were used to cultureBAECs, and MTT assay was carried out after 48 h; (4) The BAECs wereseeded onto the 96-well plates where the MMC (2g/ml, 2h) treated RMCs hadbeen attached to the bottom, then different concentrations of glucose of DMEMwas added into the plates, and the MTT assay was carried out after 48 h.Results: (1) The A values evaluating the viability of the RMCs culturedin different concentrations (5.5, 15, 30, and 60mM) of glucose and the corresponding hypertonic mannitol controls were (0.264+0.006 vs 0.264+ 0.006) ), (0.293+0.007 vs 0.285+0.007) , (0.326+0.009 vs 0.318+ 0.009) ,and (0.280+0.007 vs 0.297+0.010) respectively; (2) The A values evaluating the viability of the RMCs cultured in different concentrations (50, 100, and 200 mL.L-1) of sera from diabetic rats and age-matched control rats were(0.208+0.020 vs 0.231+0.020), (0.262+0.013 vs 0.304+0.014), and (0.327+0.021 vs 0.380+0.024) , and their inhibition rates were 10.0%, 12.2%, and 13.9%; (3)...
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