Effect Of Lactational Exposure To TCDD On The Development Of Offspring Mice And The Expression Of CYP1A1 In Lungs

ObjectiveThe contaminant 2, 3, 7, 8-tetrachlordibenzo-p-dioxin (TCDD) is one of the most toxical compounds and belongs to the category of high carcinogenicity. It is called"environmental hormone"for the ability of disordering the endocrine system, development and reproduction of animals. At present, the acute toxicity of TCDD and the effect of gestational exposure to TCDD on offspring have been well investigated in animals, but the effect of lactational exposure to TCDD on offspring has been little. In this study, the female mice got TCDD by intraperitoneal injection immediately after parturition and the offspring by breast feeding. The present study would delicate the toxic effect of TCDD lactational exposure on offspring mice by observing development of offspring mice, histological change and expression of cytochrome P4501A1 enzyme (CYP1A1) in lungs. The results would provide evidence for the study about the chronic effect of lactational exposure to TCDD on offspring.Methods1 Animals Adult Kunming mice with 24 female and 8 male were grouped by 3:1. After parturition, the female mice with 7~10 pups were retained. Then the female mice and its offspring, regarded as a unit, were divided into 5 groups randomly and each group had three units.2 ChemicalsTCDD was dissolved in methylbenzene with a concentration of 10μg/ml when purchased, and diluted with earthnut oil by 1:4. The control agent was the mixed liquor with earthnut oil and methylbenzene by 1:4.3 GroupsThe five groups were classified by a dose of 40μg TCDD/kg body weight, 20μg TCDD/kg body weight, 40μg control solvent/kg body weight, 20μg control solvent/kg body weight and 0μg control solvent/kg body weight, and noted as 40μg TCDD, 20μg TCDD, 40μg solvent control, 20μg solvent control and black control group respectively. After parturition, the female mice were weighted and intraperitoneally injected.4 The observed indexes of development for offspring mice4.1 The growth of offspring (the time of eyes-opening, superior-teeth eruption, vagina patefaction and testes descensus, the body weight change along with time)4.2 The development of nerve (turning-over test and distance between outer toes of hind feet)4.3 The effect on immune organs (the weight and quotient of thymus gland and spleen) 5 The structure change in lungs and expression of CYP1A1The structure change in lungs of offspring was observed by light microscope, and the expression of CYP1A1 in lungs was detected by immunohistochemistry (CYP1A1 took on brown particles).Results1 The effect of TCDD on the growth of offspring mice1.1 Body weightThere was no significant difference for body weight of offspring between five groups when the offspring were born (P>0.05). Along with the time of breast feeding, the body weight in 40μg and 20μg TCDD groups was light significantly compared with control groups (P<0.05). The result demonstrated that lactational exposure to TCDD had an adverse and long-term effect on body weight of offspring mice.1.2 The time of eyes-openingThe eyes-opening time in 40μg TCDD group was earlier significantly than in 40μg solvent control group and 20μg TCDD group (P<0.05), which indicated that lactational exposure to TCDD could make the eyes of offspring mice open in advance, and promote the growth and development of offspring mice in some degree.1.3 The eruption time of superior teethThe eruption time of superior teeth in 40μg and 20μg TCDD groups was earlier significantly than in 40μg and 20μg solvent control groups (P<0.05), which indicated that lactational exposure to TCDD could make the superior teeth of offspring mice erupt in advance, and the result was coincident with eyes-open time.1.4 The development of reproductive organsThe vagina-open time of female offspring in 40μg and 20μg TCDD groups was earlier significantly than in 40μg and 20μg solvent control groups (P<0.05), but the testes-descent time of male offspring in 40μg and 20μg TCDD groups was later significantly than in 40μg and 20μg solvent control groups (P<0.05), and the testes-descent time in 40μg TCDD group was late significantly compared with 20μg TCDD group. It demonstrated that lactational exposure to TCDD could promote the female offspring mice but suppress the male offspring mice in reproductive organs.2 The effect of TCDD on the development of nerve2.1 Turning-over testThere was significant difference in the correct rates of turning-over test between five groups at the 9th,11th day (P<0.05), and no difference at the 13th day(P>0.05). Compared between two groups optionally byχ2 partitions, the standard of test had to be corrected,α'=0.0045. At the 9th day, the correct rate in 40μg TCDD group was decreased significantly compared with 40μg solvent control group (P<0.0045). At the 11th day, the correct rate in 40μg TCDD group was decreased significantly compared with 20μg TCDD group and 40μg solvent control group (P<0.0045). It suggested that lactational exposure to TCDD could delay the nerval development of mice offspring in some degree.2.2 The distance between outer toes of hind feetThe distance between outer toes of offspring's hind feet in 40μg and 20μg TCDD groups was shorter significantly than in 40μg and 20μg solvent control groups at the 15th, 30th day (P<0.05), which suggested that lactational exposure to TCDD could decrease the ability of movement and coordination of offspring.3 The effect of TCDD on immune organs of offspring mice3.1 Female offspring miceThe weight of thymus and spleen in 40μg and 20μg TCDD groups was decreased and the quotient of thymus was increased compared with 40μg and 20μg solvent control groups, but there was no significant difference in them (P>0.05). There was no obvious regulation in quotient of spleen between five groups.3.2 Male offspring miceThe weight of thymus in 40μg and 20μg TCDD groups was increased compared with 40μg and 20μg solvent control groups, and the weight of spleen was decreased, but there was no significant difference (P>0.05). The quotient of thymus in 40μg and 20μg TCDD groups was increased significantly compared with 40μg and 20μg solvent control groups (P<0.05), and the quotient of spleen in 40μg TCDD group was significantly increased (P<0.05), too.It suggested that the effect of lactational exposure to TCDD on the immune organs of offspring mice maybe has sex difference: the quotients of immune organs in male offspring increased but there was no significant change in the female offspring.4 The histological change in lungs and expression of CYP1A14.1 The histological change in lungs of offspring miceSerious patho-structure of lungs showed up in 40μg and 20μg TCDD groups: Inflammatory cell infiltrated, alveolar wall thickened and dropsied, alveolar space enlarged and brown particles (CYP1A1) increased obviously. In addition, the pathological changes in lungs were more serious and the brown particles were more increased in 40μg TCDD group than in 20μg TCDD group. Nevertheless, in solvent control groups and black control group, the structure of lungs was normal, the pulmonary alveoli was regular and the brown articles had little expression.4.2 The expression of CYP1A1 in lungs of offspring miceCYP1A1 could be semi-quantitated by integral optical density. The integral optical density in 40μg and 20μg TCDD groups was increased significantly compared with 40μg and 20μg solvent control groups (P<0.05). And within the comparison of each group, the integral optical density in the female offspring mice of 40μg and 20μg TCDD groups was respectively increased significantly compared with the male offspring mice(P<0.05). It suggested that lactational exposure to TCDD could increase the expression of CYP1A1 in lungs of offspring mice and significant sex difference was certified: CYP1A1 in the female offspring mice was more than in the male offspring mice.Conclusion1 The effect of lactational exposure to TCDD on growth of offspring mice was different according to different indexes and sex: lactational exposure to TCDD slowed the body weight increasing of offspring mice; made the eyes open and superior teeth erupt in advance; promoted the development of reproductive organs of female offspring mice and suppressed the development of reproductive organs of male offspring mice2 Lactational exposure to TCDD delayed the nerve development, and decreased the ability of movement and coordination for offspring mice.3 Lactational exposure to TCDD significantly increased the quotients of thymus and spleen in the male offspring mice, but there was no significant change in the female offspring mice. It needs further research on whether the immunotoxicity of TCDD on offspring mice has sex difference or not4 Lactational exposure to TCDD made the structure in lungs of offspring mice disordered, the inflammatory cell infiltrated, alveolar wall thickened and dropsied, alveolar space enlarged. TCDD could increase the expression of CYP1A1 in lungs of offspring mice and and there was significant difference between sex: the expression of CYP1A1 in lungs of female offspring mice was more than in lungs of male offspring mice.