| Heavy metal pollution is rapidly widespread with fast urbanization and causes serious environmental pollution and health problems.Lead(Pb)is well-recognized for its great hazards to human and wildlife health.It has negative influences on multiple organs and systems of birds.However,the systematic research of its effects of lead on bird reproduction and liver glucolipid metabolism are scarcely studied.Japanese quails(Coturnix japonica),a domesticated bird worldwide,is an ideal experimental animal model in developmental biology and toxicology.In this study,Japanese quails were exposed to environment-related lead concentrations of 50,500,and 1000 ppm,respectively.We examined the effects of chronic lead exposure on growth and development,thyroid histology,thyroid hormone secretion,thyroid induced ovarian development,ovarian cell apoptosis,estradiol secretion,steroidogenesis,and the effects on liver development and glycolipid metabolism through PI3K/Akt signaling pathway.The main research results were as follows:1.High Pb exposure caused thyroid gland underdevelopment.Histopathological damages featured by deformed and atrophied follicles and disorganized epithelial cells in thyroid were induced by high lead exposure.Compared with that in control group,thyroid follicular diameter was significantly reduced in 500 ppm and 1000 ppm Pb groups.By the time of 36 days exposure,T3 and T4 concentrations of Pb exposure groups were significantly decreased in comparison with those of control.The mRNA expression of Dio1 and Dio2 in all three Pb groups were significantly lower than that in the control.Significant up-regulation in Dio3 mRNA levels was detected in 50 ppm Pb group.The TRa and TR? transcript levels showed significant upregulation in 50 ppm group.Moreover,Pb accumulation in the ovary had a positive correlation with the supplemental levels of lead acetate,and high dose Pb caused food and water intake decrease,body weight loss and growth inhibition.High dose Pb exposure also induced ovarian histopathological damages with the characters of granulosa cells disorganization,follicle atresia and interstitial cell degeneration.The number of primary and developing follicles decreased markedly in the ovary of 500 and 1000 ppm groups.The ovary follicle size in 50,500 and 1000 ppm Pb groups was significantly reduced compared to that in control.The steroidogenesis related genes(StAR,P450scc,Cyp17,3?-HSD,17?-HSD,P450arom)were significantly down-regulation in the ovary of 500 and 1000 ppm groups while the expressions of ERa and ER? in all Pb groups was significantly up-regulated in contrast with those in control group.Moreover,significant decrease of estradiol was also found in 500 and 1000 ppm groups.TUNEL assay showed that high dose Pb caused increasing cell apoptosis in ovary.The mRNA levels of apoptosis pathways related genes(Cyt-c,Casp-3 and Casp-9)were significantly up-regulated in all Pb groups.But,significant down-regulation in Bcl-2 mRNA levels was detected in 500 and 1000 ppm Pb groups.The study implied that Pb caused ovarian underdevelopment and dysfunction by inducing ovary and thyroid histopathological injury,thyroid hormone and estradiol inhibition and ovarian steroidogenesis disruption.2.High doses of lead(500 and 1000 ppm)increased mortality while feed conversion rate and liver weight index decreased significantly.The results indicated that the Pb level was increased significantly in liver of quail exposed to ascending concentrations of lead,especially in the high Pb dose groups.We observed that severe liver damage caused by high doses of lead accumulation,including blood sinusoids irregularity and dilation,hepatic lipid vacuolization,hepatocellular cytoplasm hyalinization and vacuolization,severe congestion of hepatic sinusoid.Pb led to marked reductions in the serum levels of TG.Moreover.the number of lipid droplets in hepatocytes of quails decreased significantly after lead exposure found by oil red O staining.Fewer and more sporadically distributed lipid droplets were observed in the 500 and 1000 ppm Pb groups.PAS stain of liver sections showed an obvious increase in liver glycogen in all Pb-treated quail samples.The percentage of glycogen area of 50,500 and 1000 ppm Pb groups were significantly higher than that of control group.The transcript level of PI3K/Akt signaling pathway related genes showed different patterns under lead exposure.The PI3K/Akt pathway regulates glucose metabolism through GSK-3 and FoxOl,and the expression levels of these genes were downregulated in 500 and 1000 ppm pb groups.As the rate control enzyme of liver gluconeogenesis,we observed significantly decreased G6Pase and PEPCK expression in high Pb groups.Expression of GS,regarded important genes for hepatic glycogen synthesis,was upregulated in all pb groups.The expression of three rate-limiting enzymes in the glycolytic pathway(HK,PFK1.and PK)were significantly down-regulation in 500 and 1000 ppm pb groups.The PI3K/Akt pathway regulates lipid metabolism through mTOR and SREBPlc.and the expression levels of these genes were significantly lower in 500 and 1000 ppm Pb groups.With regard to de novo lipogenesis genes,we observed significantly downregulated ACL,ACC,FAS and SCD-1 expression in 500 and 1000 ppm Pb groups than that in control group.We also found that the expression of triglyceride synthesis genes(GPAT and DGAT)was significantly downregulated in 500 and 1000 ppm Pb groups.The expression of CPT-1 and CPT-2,major enzymes for fatty acid ?-oxidation,was upregulated in all pb groups.Our results indicate that lead exposure leads to glycogen accumulation and reduced lipid accumulation and triglyceride amounts by inducing liver histopathological injury,the PI3K/Akt signaling pathway inhibition.which ultimately affects the normal liver function of birds and damages their health.The study would contribute to understand the mechanism of metal pollution on birds,provide new insight into the potential health risks of Pb to birds,and it also provides advices on poultry farming in China. |
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