Serum proteins and disazo dye teratogenesis.

Serum proteins of pregnant rats were analyzed by paper electrophoresis following administration of the teratogenic disazo dyes trypan blue, Evans blue, Niagara blue 4B, Niagara sky blue 6B, Congo red, and Niagara blue 2B. Sera were obtained from blood taken by cardiac puncture just before dye injection, 48 hours after dye injection, and again at autopsy. All of the dyes tested caused a reduction in serum albumin and total protein concentration and a n elevation in the concentration of beta or alpha-2 globulins a t some time during pregnancy. The remaining serum fractions reacted variably to the dye treatment. In spite of the rather consistent results presented in this paper, and similar findings reported for other teratogens, there is no direct proof that altered serum proteins, per se, are causally related to the production of malformations. Clinical and experimental observations suggest that altered electrophoretic patterns of serum proteins may accompany pregnancies terminating in the birth of malformed offspring. Abnormal electrophoretic patterns were noted in the serum proteins of rats treated with various teratogens (see table 1 for references). Fairly consistent among these were decrease in serum albumin and increase in serum alpha globulin. Similar changes were found in sera from high-risk pregnant women (Langman et al., '59), in 78% of whom pregnancy ended in abortion, birth of premature infants with or without malformation, or abnormal full-term children. These observations have generated speculation about the possible relation of altered serum proteins and congenital malformations. The results reported below were obtained during an investigation of the teratogenic activities of several disazo dyes structurally related to trypan blue (Beaudoin, '68). They support previously observed effects of certain teratogens on serum proteins in rats and suggest a relation between teratogenic potency and altered protein metabolism. MATERIAL AND METHODS Virgin female Wistar Albino rats (Albino Farms, Red Bank, New Jersey) were used. They were maintained on Rockland Complete Rat Diet ad libitum, with supplemental feedings of lettuce. Day 0 of TERATOLOGY, 2: 85-90. pregnancy was considered to begin on the morning sperm were found in the vaginal smear. On day 8 pregnant rats received an intraperitoneal injection of a 2% aqueous solution of the dye to be tested. The dose was calculated on the basis of the maternal weight on day 8. The doses used were 14 mg/100 g for the more potent dyes, trypan blue, Evans blue, and Niagara blue 4B; and 20 mg/100 g for the less potent dyes, Niagara sky blue 6B, Congo red, and Niagara blue 2B (Beaudoin, '68). All dyes were purchased from Matheson, Coleman and Bell Co., Norwood, Ohio. Blood was withdrawn via cardiac puncture just before dye was injected, 48 hours after dye injection, and at the time of autopsy. Sera were analyzed by the paperelectrophoretic method as described previously (Beaudoin, '66). Total protein content of the serum was measured with a Bausch and Lomb Serum Protein Meter. Pregnancy was terminated on day 20 and the fetuses were recovered, weighed, and fixed in Bouin's fluid. The types and frequencies of the malformations produced are listed in table 2 and have been previously reported (Beaudoin, '68). . RESULTS Table 3 summarizes the results of this experiment. The dyes are listed in the order of decreasing teratogenicity. Blood ISupported by USPHS grant HD 00400 from the National Institute of Child Health and Human Development.