Development of Hypothalamic-Pituitary-Adrenal Axis and Adrenal Requires Fetal Steroids Synthesis

During pregnancy, fetal corticosterone is derived both from that transferred from mother and that secreted from their own adrenal. The P450scc (Cyp11a1) enzyme, which catalyzes the first step of steroidogenesis, contributes to de novo synthesis of steroid hormones. Thus, the corticosterone in Cyp11a1 null embryos all comes from their mother through placenta, and this provides an animal model to study the disorder due to the lack of de novo synthesis of steroid hormones. The normal level of plasma corticosterone in the Cyp11a1 null mice indicates the transplacental corticosterone provides enough corticosterone level in fetal circulation. Even so, the developmental disorders were still detected. The null mice had enlarged and randomly distributed oil droplets in the adrenal at 16.5 dpc and the structure of adrenal was altered in parallel with random distribution of proliferating cells. The medulla migration was normal in the Cyp11a1 null mice, but this null adrenal could not synthesize epinephrine. In HPA axis of null mice, pituitary POMC had higher expression than the wildtype, which was consistent with the higher ACTH level in the plasma. Our results suggest that: (1) Inability of the fetal steroid synthesis during pregnancy affects adrenal organization. (2) The development of adrenal medulla required de novo corticosterone, which synthesize from vicinal cortex. (3) The de novo synthesis of steroid hormone is essential for normal development of the negative feedback function in HPA axis.Table of Content Abstract in Chinese 1 Abstract 2 Introduction I. Steroid hormones Classes & Functions 3 Steroidogenesis 3 II. Adrenal Gross Anatomy 4 Cortical Zonation 4 Zonal specific markers in cortex 5 Medulla 5 Development of the embryonic mouse adrenal 6 III. Corticosterone & Stress Stress response systems 6 Sympatho-adrenomedullary systems 6 HPA axis 7 Negative feedback in HPA axis 7 Non-glucocorticoids inhibitors involved in the negative feedback 8 IV. Corticosterone & Adrenal Function Adrenalmedullary function and corticosterone 8 Cortical steroidogenic ability and corticosterone 9 V. Maternal corticosterone & Fetal Development Transplacental corticosterone 9 Corticosterone and fetal development 10 VI. Models of Steroids Deficiency Congenital adrenal hyperplasia 11 Cyp21 deficient mouse 11 Models of lipoid CAH 11 Other steroids deficiency models 12 VII. Aims & Motivation 13 Materials and Methods The generation of Cyp11a1 KO mice 14 The genotyping of Cyp11a1 KO mice 14 Wax embedding 14 Hematoxylin and eosin staining 14 Proliferating cell nuclear antigen (PCNA) staining 15 Oil Red O staining 15 TUNEL assay 15 Immunohistochemistry 16 Hormone assays 16 Real-Time PCR 16 Epinephrine measurement 17 Chromaffin reaction 17 Results Lack of daily rhythm of corticosterone in Cyp11a1 null fetus 18 Activated HPA axis in Cyp11a1 null fetus 18 Lower HSD11b expression in Cyp11a1 null mice brain 19 Disorganized embryonic adrenal of Cyp11a1 null mice 19 Lipid accumulation in null adrenal 20 Increased apoptosis and impaired cell proliferation in Cyp11a1 null mice 20 Lack of adrenergic chromaffin cells in Cyp11a1 null mice 20 Less catecholamine synthesis in Cyp11a1 null medulla 21 Discussions Maternal and fetal corticosterone secretion 22 Lack of de novo synthesized corticosterone 22 Steroid secretion defect leads to disorganized adrenal 23 Medulla function and corticosterone from vicinal cortex 23 Neurosteroids and negative feedback 24 11beta-HSD-1 involved in glucocorticoid action 25 Negative feedback in Cyp11a1 null mice 26 Future Experiments 27 References 28 Tables 35 Figures 3