Lucke S, Ziegler B, Komolov I, Hahn H J
Acta Histochem. 1986;79(1):33-41.
DNA synthesis and labelling index of isolated neonatal rat islets were investigated during a 4 to 5 culture period in TCM 199 (10 mmol/l glucose) under selected conditions, as change of serum concentration in the medium or addition of a proliferation stimulating fraction. Additionally it should be proved whether the DNA-synthesis of pancreatic islets, measured as [3H]-thymidine incorporation, is representative for the amount of cells during the S-phase (= "growth fraction"). There is a correlation between the incorporation of [3H]-thymidine into the TCA-precipitate and the labelling index of islets determined on HE-stained radioautographs. For this reason and because of being less time-consuming, the measurement of DNA synthesis is a suitable screening method for selection of factors increasing the growth fraction of pancreatic beta-cells. Only the addition of FGF-S3 (growth stimulating raw prepared isolated from bovine brain) during culture caused a significant increase of DNA synthesis and labelling index of the islets. The labelling index of radioautographs stained by immunofluorescence method for insulin did not differ significantly from that determined at HE-stained radioautographs. Insulin secretion and content of the islets were not influenced by the addition of FGF-S3.
在选定条件下,于含10 mmol/l葡萄糖的TCM 199培养基中,对分离的新生大鼠胰岛进行4至5天培养,研究其DNA合成及标记指数,条件包括改变培养基中血清浓度或添加增殖刺激因子。此外,还应证明以[3H] - 胸腺嘧啶核苷掺入量衡量的胰岛DNA合成是否代表S期(即“生长分数”)的细胞数量。[3H] - 胸腺嘧啶核苷掺入三氯乙酸沉淀中的量与在苏木精 - 伊红染色放射自显影片上测定的胰岛标记指数之间存在相关性。因此,且由于耗时较少,DNA合成的测量是一种筛选增加胰腺β细胞生长分数的因子的合适方法。仅在培养期间添加FGF - S3(从牛脑分离制备的生长刺激原料)会导致胰岛的DNA合成和标记指数显著增加。用胰岛素免疫荧光法染色的放射自显影片的标记指数与苏木精 - 伊红染色放射自显影片上测定的标记指数无显著差异。添加FGF - S3对胰岛的胰岛素分泌和含量无影响。