Abstract
Maternal obesity during pregnancy is associated with chronic low-grade inflammation and results in long-term health risks in the offspring. This thesis has investigated the hypothesis that elevated inflammatory responses in obese mothers would alter development of the fetal brain. In particular, non-neuronal cells of the mediobasal hypothalamus, including those that form the blood-brain barrier (BBB), were examined for maternal obesity-induced developmental alterations. The studies here also investigated the hypothesis that the placenta of an obese mother plays an active role in creating the inflammatory environment, in which the fetus is abnormally exposed to inflammatory responses in the placenta of an obese mother.
The first experimental chapter (Chapter 3) was focused on the placenta, and histomorphology and inflammatory responses in mice placenta were studied from normal-weight mothers (control) and a high-fat diet fed obese mothers (mHFD). We hypothesised that elevated inflammatory responses in obese mothers would elevate inflammatory responses in the placenta and disrupt placental structure.
mHFD placentas showed changes in the composition of the placental layers and cell proliferation compared to the controls. Placental inflammatory responses were elevated during late gestation (GD17.5), with increased macrophage activation and elevated cytokine gene expression. With increased placental inflammatory responses, an increase in cytokine protein levels (IL6, IFNy and IL17A) were observed in the fetal circulation of mHFD group compared to the controls at GD17.5.
This chapter provides an understanding of the changes that take place in the placenta when a mother is obese during pregnancy.
The second experimental chapter (Chapter 4) was to study the possible consequences of elevated fetal cytokines of obese mothers on fetal brain development. The fetal hypothalamus, a key brain region that regulates energy homeostasis, was the focus of this study. Glial cells (astrocytes and microglia) in the fetal hypothalamus were studied, as generation of glial cells is initiated during the fetal period and mediated by cytokines. We hypothesised that elevated fetal cytokines of obese mothers would disrupt the development of glial cells in the hypothalamus.
The supraoptic nucleus (SON) and arcuate nucleus (ARC) of the hypothalamus of GD17.5 mHFD fetuses showed significantly increased GFAP expression compared to the controls. By contrast, only a small number of microglia were present in the fetal hypothalamus at GD17.5, and neither mRNA, cell number or morphology were different between dietary groups.
The role of cytokine(s) in regulating GFAP expression or fetal astrocyte cell numbers was tested in vitro. To do this, astrocytes from the fetal SON and ARC were placed in culture with either IL6, IFNy and IL17A, the cytokines that were shown to be elevated in the fetuses of obese mothers (Chapter 3). IL6 treatment resulted in a significant increase in proliferation of both ARC and SON astrocytes, but neither IFNy or IL17A had any effect.
This chapter shows that maternal obesity elevates astrocyte expression in the fetal hypothalamus in vivo, and IL6 in vitro could stimulate proliferation of astrocytes, which could explain how astrocyte expression increases in the fetuses of obese mothers in vivo; elevated IL6 in the fetal circulation of obese mothers could increase proliferation of hypothalamic astrocytes.
The third experimental chapter (Chapter 5) was to study the BBB in the offspring of obese mothers, to test the hypothesis that perturbed development of the BBB in these offspring impairs communication between blood-borne molecules and the offspring's brain.
Tanycytes, a gate-keeper of blood-borne molecules into the ARC, were reduced in the offspring of obese mothers compared to the controls. Experiments were also conducted to study several features of capillary endothelial cells that are important for the formation of the BBB; expression of tight junctions, fenestrations and leptin transporters were perturbed in the ARC of the offspring of obese mothers compared to the controls. These experiments suggest that the ARC of the offspring of obese mothers could be abnormally exposed to blood-borne molecules.
Intraperitoneal injection of Evans Blue dye was used to test permeability of the BBB by quantifying its diffusion into the ARC. Consistent with the findings of changes in a number of markers that indicated disruption of the BBB in the offspring of obese mothers, Evans Blue diffusion into the ARC was significantly higher in the offspring of obese mothers compared to the controls.
This chapter shows that maternal obesity can compromise the formation of the BBB in the offspring, leading to an increase in exposure to blood-borne molecules into the ARC.
In conclusion, this thesis has shown: 1) maternal obesity elevates inflammatory responses in the placenta and fetuses, 2) elevated inflammatory responses in the fetuses of obese dams alter the development of astrocytes in the hypothalamus, and 3) maternal obesity during pregnancy disrupts the proper formation of the BBB in the offspring.