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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（０３ｍＭ）、中（０６ｍＭ）、高（１２ｍＭ）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。
In order to further understand the effect of caffeine on fetal and neonatal germ cells and its mechanism, we used SD (18mM), low (0.3mM), medium (0.6mM) and high (1.2mM) Fetal rats (18d), 0d (0d) and 4d (4d) testes were cultured for 1 week, 2 weeks and 3 weeks respectively. The effects of caffeine on the number of testicular germ cells were observed. And computer image analysis to detect the expression of PCNA (proliferating cell nuclear antigen), DNA content and other indicators related to DNA synthesis in order to study the possible mechanism of caffeine. The results are as follows: (1) The embryonic germ cells cultured on the 18th day in fetal testes had the least effect on caffeine, followed by 0-day-old suckling rats and 4-day old suckling rats. (2) Low concentrations of caffeine had little effect on germ cells; the number of germ cells and the content of DNA in medium concentration of caffeine decreased only after three weeks of culture, while the high concentration of caffeine Indicators have declined, three weeks more obvious. (3) The decrease of germ cells often accompanied with the decrease of DNA content, but also with the decrease of PCNA expression. It is speculated that high concentrations of caffeine after a long time to reduce the number of germ cells, may inhibit the positive expression of PCNA, interfere with DNA replication.