咖啡因对睾丸生殖细胞PCNA表达、DNA含量影响的体外研究

The effects of caffeine on the PCNA expression, DNA content of germ cells in testis of fetal and newborn rats

为了深入了解咖啡因对胎儿、新生儿生殖细胞的影响及其机制，本实验采用低（０３ｍＭ）、中（０６ｍＭ）、高（１２ｍＭ）浓度咖啡因体外培养ＳＤ孕１８天胎鼠（１８ｄ）、０天（０ｄ）及４天（４ｄ）乳鼠睾丸组织块，培养时间分别为１周、２周、３周，观察咖啡因对睾丸内生殖细胞数量的影响并用免疫组化、计算机图像分析等技术检测生殖细胞ＰＣＮＡ（增殖细胞核抗原）的表达、ＤＮＡ的含量等与ＤＮＡ合成有关的指标，以研究咖啡因作用的可能机制。结果如下：（１）１８天胎鼠睾丸培养组织生殖细胞受咖啡因影响最少，０天乳鼠次之，４天乳鼠受影响较大。（２）低浓度的咖啡因对生殖细胞影响较少；中等浓度的咖啡因在培养三周后，生殖细胞的数量、ＤＮＡ的含量才降低；而高浓度的咖啡因在培养二周后，以上指标已有下降，三周更明显。（３）生殖细胞数量的减少往往伴随ＤＮＡ含量的降低，也与ＰＣＮＡ表达程度的减少有一定的一致性。由此推测高浓度咖啡因长时间培养后使生殖细胞数量减少，可能与抑制ＰＣＮＡ表达的阳性程度，干扰ＤＮＡ复制有关。

Low (03mM)、medium(06mM) and high (12mM) doses of caffeine were used in tissue culture of testicular organ explant taken from 18 d fetal rats and 0 d and 4 days neonatal rats to investigate the effects of caffeine on the number, the PCNA expression and the DNA content of germ cells. Techenics of immunohistochemistry and computer image analyses were used. The culture time was 1 week, 2 weeks and 3 weeks, respectively. The results were as follows: (1) The effects of caffeine on the parameters of 18 days fetal testes explant were the least; those on the parameters of 0 d were moderate; while those on the parameters of 4 d testes were the most. (2) Low doses of caffeine exerted little influence on germ cells. Medium doses of caffeine decreased the number of germ cells and DNA content after 3 weeks of culture. High doses of caffeine, however, reduced these important parameters just after 2 weeks of culture. (3) The decrease of the number of germ cells were accompanied by the reduction of DNA content and accorded to the degree of PCNA expression in several group. It may be deduced that high doses of caffeine inhibit the proliferation of germ cells after culturing for a long time through reducing the degree of PCNA expression and intervening the synthesis of DNA.