Teratogenic effects of bis-diamine on the developing myocardium.
BACKGROUND
Bis-diamine induces conotruncal anomalies and disproportional ventricular development in rat embryos when administered to the mother. To evaluate the mechanisms of disproportional ventricular development in the anomalous heart, we analyzed the morphology of the embryonic heart and investigated cardiomyocytic DNA synthesis and apoptosis.


METHODS
A single dose of 200 mg of bis-diamine was administered to pregnant rats Wistar on day 9.5 of pregnancy. The embryos were removed on each embryonic day from 10.5 to 18.5. Expression of cardiotrophin-1 and hepatocyte growth factor was investigated on the sections, and cardiotrophin-1, hepatocyte growth factor and myocyte enhancer factor 2 mRNA expression was examined by reverse transcriptase-polymerase chain reaction. Myocardial DNA synthesis was investigated using 5-bromo-2'-deoxyuridine and the labeling index was calculated for each heart. Apoptosis was also analyzed using TUNEL reaction and electrophoresis of DNA fragmentation.


RESULTS
The embryos treated with bis-diamine had conotruncal anomalies associated with thin left ventricular wall in the later stage. The labeling index on embryonic day 15.5 and 16.5 was significantly lower than those in the controls. Hepatocyte growth factor and cardiotrophin-1 mRNA expression was upregulated on embryonic day 12.5 and 15.5 in bis-diamine-treated hearts. Fewer apoptotic cells were detected in the hearts of bis-diamine-treated embryos than in control hearts from embryonic day 14.5 to 16.5.


CONCLUSIONS
The ventricular disproportion in the bis-diamine-treated heart may be caused by the early myocardial differentiation delay and poor proliferation and reduced apoptosis associated with anomalous circulatory condition in the later stage.
Okamoto
Nobuhiko
N
Department of Pediatrics, Shiga University of Medical Science, Seta, Otsu, Shiga 520-2192, Japan. cxq07667@nifty.com
Nakagawa
Masao
M
Fujino
Hidetoshi
H
Nishijima
Setsuko
S
Hanato
Takashi
T
Narita
Tsutomu
T
Takeuchi
Yoshihiro
Y
Imanaka-Yoshida
Kyoko
K
D000818:Animals; D017209:Apoptosis / Q000187:drug effects; D001483:Base Sequence; D001973:Bromodeoxyuridine; D002454:Cell Differentiation / Q000187:drug effects; D002455:Cell Division / Q000187:drug effects; D017931:DNA Primers; D003959:Diamines / Q000633:toxicity*; D006321:Heart / Q000187:drug effects* / Q000196:embryology; D007150:Immunohistochemistry; D020287:In Situ Nick-End Labeling; D051381:Rats; D017208:Rats, Wistar; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013723:Teratogens / Q000633:toxicity*
D017931:DNA Primers; D003959:Diamines; D013723:Teratogens; D001973:Bromodeoxyuridine