鸡胚前脑细胞在各种支持物特别是人羊膜基膜上生长的定量研究

QUANTITATIVE ANALYSIS OF FOREBRAIN CELLS OF CHICK EMBRYO GROWTH ON VARIOUS SUBSTRATA ESPECIALLY THE HUMAN AMNIOTIC BASEMENT MEMBRANE

作者采用Ⅰ型胶原、人羊膜基膜(HABM)基质面、Fibronectin(FN)、多聚鸟氨酸(PO)、PO+FN、PO+Laminin(LN)、鼠尾胶原和 HABM 上皮面作为8天鸡胚前脑细胞生长的支持物,观察这些支持物对前脑细胞生存生长的影响。前脑细胞以每个孔不同的细胞密度在 PO、LN 和HABM 上皮面培养。发现每个孔4×10~5细胞密度细胞生长最好。用 MTT 做为线粒体脱氢酶底物的自动比色法,对前脑细胞在各种支持物上的生长进行了定量测定。结果表明,所测光密度值(OD)与细胞生存、生长成正比,且 HABM 上皮面是前脑细胞生长最好的支持物。各种支持物促进前脑细胞生长的顺序浓次是:HABM 上皮面>鼠尾胶原>PO+LN>PO+FN>PO>FN>HABM 基质面>Ⅰ型胶原。HABM 上皮面比基质面促进前脑细胞生长作用明显强(P<0.01);鼠尾胶原比Ⅰ型胶原强(P<0.05);PO+LN 比 PO 作用强(P<0.05)。这提示 LN 很可能是 HABM、鼠尾胶原和 PO+LN 中促进神经生长的主要活性因子。

By using collagen type Ⅰ,stromal surface of human amniotic basement membrane (HABM),fibronectin(FN),polyornithine(PO),PO-precoated fibronectin,PO- precoated laminin(LN),collagen gel(from rat tail)and the epithelial surface of HABM as substrata to test their ability in promoting growth and neurite outgrowth of cells from the forebrain of E8-chick embryo.Forebrain cells of various cell densities per well were cultured on PO,LN and the epithelial surface of HABM.It was found that the neurite outgrowth was most extensive in cultures at cell density of 4×10~5 per well.An automated colorimetric assay,using MTT as substrate for mitochondrial dehydrogenases,was used to quantify the growth of forebrain cells on different substrata.It was found that the optical density(OD)is directly pro- portional to the number of viable cells per well seeded and/or the extent of cells growth.HABM was the best substratum for forebrain cells growth.The promoting ability of various substrata for forebrain cells in decreasing order was as following: the epithelial surface of HABM> collagen gel>PO-precoated LN>PO-precoated FN>PO>FN>the stromal surface of HABM>collagen type Ⅰ.The epithelial surface of HABM was significantly better in promoting forebrain cells growth than the stromal surface(P<0.01).Collagen gel was better than collagen type Ⅰ(P<0.05), while the PO-precoated LN was better than the only PO coated surface(P<0.05). The results indicated that LN was the common potent factor in HABM,collagen gel and the PO-precoated LN in promoting forebrain cells growth and neurite outgrowth.