Catalano A, Huang P H
Foundation 41, East Sydney, New South Wales, Australia.
Teratog Carcinog Mutagen. 1992;12(5):231-41. doi: 10.1002/tcm.1770120506.
Diethylnitrosamine (DEN) and methylazoxymethanol acetate (MAM) are not transplacental carcinogenic but embryotoxic to Wistar rats when administered by i.p. injection on day 12 of gestation. MAM, a weak teratogen to rats during this period, induced a dose dependent increase in the number of resorptions to 15% and 40% of the litters following doses of 15 and 25 mg/kg bw, respectively. Rats similarly treated with 70, 150, and 180 mg DEN/kg bw resulted in increases in total DNA mass of day 13 embryos by 31%, 45% and 52%, respectively, compared to the saline treated controls. Twenty percent reduction in total DNA amount was detected following 25 mg MAM/kg bw. Benzoylated DEAE-cellulose (BD-cellulose) chromatography fractionates DNA on the basis of secondary structure by stepwise elution of double-stranded DNA with 1.0M NaCl solution (SE-DNA) followed by elution of DNA containing single-stranded regions with caffeine solution (CE-DNA). Day 13 embryonic DNA was monitored by in vivo labelling with [methyl-3H]-thymidine (3H-TdR) on days 6 and 7 of gestation. Significant increases in percentages of caffeine-eluted DNA (%CE-DNA) compared to control values were detected 24 h after treatment of day 12 embryos with 70, 150, and 180 mg DEN/kg bw. Such increases were not observed after MAM. Incorporation of [methyl-14C]-thymidine (14C-TdR) into embryonic DNA demonstrated the effects of treatment with these compounds on DNA synthesis in vivo. When compared to saline controls, DEN induced significant increases in 14C-TdR incorporation into embryo DNA, 1 h prior to analysis, but the increases were not proportional to the doses administered. Similar analysis of MAM treated samples showed no significant changes to %CE-DNA values. The relative %CE-DNA is expressed as the ratio of the percentage of caffeine-eluted 14C-labelled DNA to %CE-DNA (i.e., %CE-14C-DNA:%CE-3H-DNA). In the majority of control embryos the 14C-specific activity of CE-DNA was higher than the 14C-specific activity of SE-DNA. No significant change to relative %CE-DNA values of embryos to those of the controls was observed 24 h after treatment of day 12 gestation rats with single doses of DEN and MAM. The results of this study support the hypothesis that initiation mechanisms of teratogenesis and transplacental carcinogenesis are different. The pertinence of %CE-DNA and relative %CE-DNA values to teratogenesis and transplacental carcinogenesis is also discussed.
二乙基亚硝胺（DEN）和乙酸甲基偶氮甲醇（MAM）并非经胎盘致癌物质，但在妊娠第12天经腹腔注射给予Wistar大鼠时，它们具有胚胎毒性。在此期间，MAM对大鼠是一种弱致畸剂，分别给予15和25mg/kg体重的剂量后，导致吸收胎数呈剂量依赖性增加，分别达到窝仔数的15%和40%。用70、150和180mg DEN/kg体重对大鼠进行类似处理，与生理盐水处理的对照组相比，第13天胚胎的总DNA质量分别增加了31%、45%和52%。给予25mg MAM/kg体重后，检测到总DNA量减少了20%。苯甲酰化二乙氨基乙基纤维素（BD-纤维素）色谱法根据二级结构对DNA进行分级分离，先用1.0M NaCl溶液逐步洗脱双链DNA（SE-DNA），然后用咖啡因溶液洗脱含有单链区域的DNA（CE-DNA）。在妊娠第6天和第7天用[甲基-3H]-胸腺嘧啶核苷（3H-TdR）进行体内标记来监测第13天胚胎的DNA。用70、150和180mg DEN/kg体重处理第12天胚胎24小时后，与对照值相比，咖啡因洗脱DNA（%CE-DNA）的百分比显著增加。用MAM处理后未观察到这种增加。将[甲基-14C]-胸腺嘧啶核苷（14C-TdR）掺入胚胎DNA证明了这些化合物处理对体内DNA合成的影响。与生理盐水对照组相比，在分析前1小时，DEN诱导14C-TdR掺入胚胎DNA显著增加，但增加幅度与给药剂量不成比例。对MAM处理样品的类似分析显示%CE-DNA值无显著变化。相对%CE-DNA表示为咖啡因洗脱的14C标记DNA百分比与%CE-DNA的比值（即%CE-14C-DNA:%CE-3H-DNA）。在大多数对照胚胎中，CE-DNA的14C比活性高于SE-DNA的14C比活性。用单剂量DEN和MAM处理妊娠第12天的大鼠24小时后，未观察到胚胎相对%CE-DNA值与对照组相比有显著变化。本研究结果支持致畸作用和经胎盘致癌作用的起始机制不同这一假说。还讨论了%CE-DNA和相对%CE-DNA值与致畸作用和经胎盘致癌作用的相关性。
