吡喃葡萄糖甙对培养大鼠视网膜神经细胞的影响

Effect of 4 pyrone 3 β D glucopyraoside on rat retinal neurons in vitro

目的 观察灯盏细辛主要成份之一４吡喃酮３βＤ吡喃葡萄糖甙（４ｐｙｒｏｎｅ３βＤｇｌｕｃｏｐｙｒａｏｓｉｄｅ， ＰＧＰ）对培养大鼠视网膜神经细胞的影响，并筛选出其有效浓度。方法 ６ 只生后２～３ｄ ＳｐｒａｇｕｅＤａｗｌｅｙ 大鼠，视网膜采用胰酶消化法制成细胞悬液后接种于经鼠尾胶原处理的９６ 孔培养板（每孔１×１０５ 个细胞），同时加入１ｇ·Ｌ－ １ ～１０－ ５ ｇ·Ｌ－ １ ＰＧＰ及ＤＭＥＭ，分别培养１、３、５ｄ，用ＭＴＴ法测量ＯＤ值。对部分ｄ５ 的细胞行Ｎｉｓｓｅｌ体染色检查。结果 培养在鼠尾胶原上的视网膜神经细胞生长良好，部分细胞伸出突起，且少数突起相互连接。Ｎｉｓｓｅｌ体染色检查示培养５ｄ的存活细胞大部分为神经细胞。各种浓度的ＰＧＰ对培养细胞形态无影响，药物浓度≥１０－ ２ｇ·Ｌ－ １时，各培养时期细胞数目明显减少，ＯＤ值明显降低（Ｐ＜ ０．００１～０．０５），在该浓度以下各培养时期细胞数目未见明显减少，ＯＤ值仅有轻微下降（Ｐ＞ ０．０５）。结论 ＰＧＰ无直接促进培养大鼠视网膜神经细胞生长的作用，药物浓度≤１０－ ３ｇ·Ｌ－ １ 时，对培养细胞无毒副作用。

Objectives To observe the effect of 4 pyrone 3 β D glucopyraoside(PGP),one of the main integrants of Erigeron Breviscapus(Vant.) Hand Mazz, on rat retinal neurons in vitro and to find out its nontoxicity concentration for cultured retinal neurons.Methods The retinae of 6 postnatal 2～3 days Sprague Dawley rats were dissociated into cell suspension with trypsin digestion. The cell suspension was implanted in 96 well culture plates covered with murine tail collagen in each well(1×10 5 cells/well). The concentrations of PGP solution were 1g·L -1 ～10 -5 g·L -1 . The period of culture was 1,3 and 5 days respectively, Then, the OD of living cells in each well was tested by MTT colorimetric microassay. Some of the 5 day culture cells were identified by Nissel technique.Results The retinal neurons cultured in the well coated by murine tail collagen tissue grew very well. Some cells processed axons and some axons connected network form. Most of the living cells were retinal neurons by Nissel identification. The morphological contour of cultured retinal cells showed no significant changes in each concentrations of PGP. However, the numbers of cells and OD decreased significantly at the concentrations of above or equal to 10 -2 g·L -1 (P<0.01～0.05);The number of cells and OD decreased slightly under this concentration (P>0.05).Conclusion PGP has no direct effect on retinal neuron growth in vitro, and it has no toxicity on cells under the concentration of 10 -3 g·L -1 .