维生素B1和B6对低血糖导致的视网膜神经元损伤的保护作用

Neuroprotective effect of vitamins B1 and B6 on injury of retinal neurons induced by low glucose in vitro

目的评价维生素B1和B6对低血糖导致的视网膜神经元损伤的保护作用。方法新生大鼠视网膜单细胞悬液接种到96孔细胞培养板中。培养24h后，低血糖处理6h，同时加入不同浓度的维生素B1和B6，并抑制神经胶质细胞生长。继续培养至第4d，用MTT微量比色法检测细胞活力，测定乳酸脱氢酶（LDH）释放情况，评价低血糖引起的细胞损伤。结果低血糖处理组与未处理组相比，视网膜神经元活力显著下降（P〈0．01）；维生素B1浓度为100～300μmol／L，维生素B6浓度为100μmol／L时，视网膜神经元活力明显高于未加药组（P〈0．05，P〈0．01）；维生素B1和B6在100μmol／L时显著抑制低血糖损伤引起的LDH释放。低血糖处理结束后开始应用维生素则无明显神经保护作用。结论维生素B1和B6在一定药物浓度时对低血糖引起的视网膜神经元损伤有显著保护作用，低血糖损伤后延迟给药则未发现神经保护作用。

Objective B vitamins are usually used to the treatment of retina and optic nerve diseases as the necessary coenzyme of cell metabolism and take part in many physiological and biochemical procedure of nerve system. Our study aimed to evaluate the neuroproteetivity of vitamins B1 and B6 on the retinal neurons injury induced by low concentration of glucose. Methods Neonatal Wistar rat retinal neuron suspensions （ 1.2 × 10^6 - 1.3 × 10^6/mL） were planted on 96-well cell culture plates. After 24 hours in vitro culture,the low concentration of glucose was given in culture medium for 6 hours. Vitamin B1 （30, 100,300 μmol/L）and B6（ 10,100,1 000 μmol/L）were added simultaneously at the beginning or just after the insult respectively in different groups. Continued culturing with vitamins B1 and B6 were taken until the 4th day, and vitamin did not added into the low glucose medium in positive control group, and normal concentration of glucose was used in the normal control group. MTT chromatometry was applied to assess the viability of retinal neurons. Releasing of lactate dehydrogenase （LDH） was measured to assess the injury of retinal neurons by Koh and Choi＇ s methods. Results Cell viability of retinal neurons were declined significantly after low concentration of glucose insult in comparison with non-treating cultures（ t = 5. 046,P = 0. 001 ）. Addition of vitamins B1 and B6 demonstrated different degree of neuroprotective effects on retinal neurons. The effective action concentrations of vitamin B1 were 100 μmol/L（P =0. 018） and 300 μmol/L（P =0. 001） ,and that of vitamin B6 was 100 μmol/L（P =0. 001） in comparison with positive control group. LDH releasing was significantly depressed at the concentration of 100 μmol/L in both vitamin B1 and vitamin B6. However, the vitamins exhibited weak efficiency of neuroprotection in low concentration of glucose group. Conclusion Vitamin B1 and B6 have the protection on retinal neurons cultrued with low concentration of glucose in certain concentration. However, delayed supplying of these vitamins do not effectively rescue the retinal neurons from low concentration of glucose injury.