Multidrug resistance phosphoglycoprotein (P-gp) is expressed in the placenta and fetal blood-brain barrier (BBB) and plays a critical role in reducing fetal accumulation of xenobiotics. In other tissues, P-gp activity is inhibited by selective serotonin reuptake inhibitors (SSRIs) and by lethal doses of LPS modeling a bacterial infection. However, nothing is known with respect to the effects of SSRIs or nonlethal infection on P-gp activity in the placenta or fetal tissues. In the studies presented in this thesis, we hypothesized that (1) the SSRI sertraline and (2) a nonlethal maternal bacterial infection would decrease P-gp activity in the placenta and fetal BBB. The first study shows that sertraline affects P-gp activity at these barrier sites in a tissue-specific manner. The second study shows that nonlethal infection does not significantly affect P-gp activity at either site. However, nonlethal infection may still influence substrate biodistribution by altering hepatic elimination of these substrates.

Multidrug resistance phosphoglycoprotein (P-gp) is expressed in the placenta and fetal blood-brain barrier (BBB) and plays a critical role in reducing fetal accumulation of xenobiotics. In other tissues, P-gp activity is inhibited by selective serotonin reuptake inhibitors (SSRIs) and by lethal doses of LPS modeling a bacterial infection. However, nothing is known with respect to the effects of SSRIs or nonlethal infection on P-gp activity in the placenta or fetal tissues. In the studies presented in this thesis, we hypothesized that (1) the SSRI sertraline and (2) a nonlethal maternal bacterial infection would decrease P-gp activity in the placenta and fetal BBB. The first study shows that sertraline affects P-gp activity at these barrier sites in a tissue-specific manner. The second study shows that nonlethal infection does not significantly affect P-gp activity at either site. However, nonlethal infection may still influence substrate biodistribution by altering hepatic elimination of these substrates.

Glucocorticoid (GC) signalling via the GC receptor (GR) regulates many aspects of lung development. To determine the need for epithelial GC-GR signalling, triple transgenic (TT) mice with doxycycline (dox) – inducible suppression of GR exclusively in the distal lung epithelium (DLE) were created. Following exposure to dox, E18.5 TT fetuses showed a reduction in GR mRNA levels and elimination of GR protein expression exclusively in the DLE. Newborn TT pups had decreased viability and TT fetal lungs had increased tissue to airspace ratios, decreased levels of proximal epithelial protein CC10, of all surfactant proteins (ATII cell proteins), of ion conductance channels β and γENaC, of water channel AQP5, and of ATI cell protein T1α. Thus DLE GC-GR signalling is important for neonatal viability and increased mortality of TT pups could be due to impaired epithelial differentiation, leading to decreased surfactant protein expression, delayed fluid clearance and/or increased lung cellularity.

Glucocorticoid (GC) signalling via the GC receptor (GR) regulates many aspects of lung development. To determine the need for epithelial GC-GR signalling, triple transgenic (TT) mice with doxycycline (dox) – inducible suppression of GR exclusively in the distal lung epithelium (DLE) were created. Following exposure to dox, E18.5 TT fetuses showed a reduction in GR mRNA levels and elimination of GR protein expression exclusively in the DLE. Newborn TT pups had decreased viability and TT fetal lungs had increased tissue to airspace ratios, decreased levels of proximal epithelial protein CC10, of all surfactant proteins (ATII cell proteins), of ion conductance channels β and γENaC, of water channel AQP5, and of ATI cell protein T1α. Thus DLE GC-GR signalling is important for neonatal viability and increased mortality of TT pups could be due to impaired epithelial differentiation, leading to decreased surfactant protein expression, delayed fluid clearance and/or increased lung cellularity.

This study examined the DNA methylation profile of endothelial nitric oxide synthase (eNOS) in the placental villous trophoblast in first, second trimester and healthy term placentas. Syncytiotrophoblast DNA revealed a heterogeneous methylation pattern in the first trimester eNOS proximal promoter and transitioned to a densely methylated pattern at term. Healthy, term syncytiotrophoblast and cytotrophoblast obtained through cytotrophoblast isolation technique provided purified cell samples for RNA and DNA extraction. Real-time PCR (rt-PCR) verified the presence and quantity of eNOS mRNA. In summary, the main findings of this thesis are heterogeneous methylation in first trimester compared to hypermethylation at term; greater eNOS expression and variable methylation in term syncytiotrophoblast compared to cytotrophoblast. A heterogeneous methylation pattern in eNOS has been identified in this study and recent stem cell studies in our lab, and we propose it represents plasticity in gene expression, particularly in early development.

This study examined the DNA methylation profile of endothelial nitric oxide synthase (eNOS) in the placental villous trophoblast in first, second trimester and healthy term placentas. Syncytiotrophoblast DNA revealed a heterogeneous methylation pattern in the first trimester eNOS proximal promoter and transitioned to a densely methylated pattern at term. Healthy, term syncytiotrophoblast and cytotrophoblast obtained through cytotrophoblast isolation technique provided purified cell samples for RNA and DNA extraction. Real-time PCR (rt-PCR) verified the presence and quantity of eNOS mRNA. In summary, the main findings of this thesis are heterogeneous methylation in first trimester compared to hypermethylation at term; greater eNOS expression and variable methylation in term syncytiotrophoblast compared to cytotrophoblast. A heterogeneous methylation pattern in eNOS has been identified in this study and recent stem cell studies in our lab, and we propose it represents plasticity in gene expression, particularly in early development.

The strength of time-dependent correlations known as stride interval (SI) dynamics have been proposed as an indicator of neurologically healthy gait. Most recently, it has been hypothesized that these dynamics may be necessary for gait efficiency although the supporting evidence to date is limited. To gain a better understanding of this relationship, this study investigated stride interval dynamics, the energy cost of walking, and physical activity in a pediatric population. The findings indicate that differences in energy cost are not reflected in the stride interval dynamics of able-bodied children. Interestingly, increasing physical activity levels were associated with decreasing variance in stride interval dynamics between subjects, though this finding only approached significance (p=0.054). Lastly, this study found that stride interval dynamics in children as young as nine years were comparable to stride interval dynamics found in healthy young adults.

The Sonic hedgehog (Shh) signaling pathway plays a critical role in murine gastric development. When Shh is knocked out in the mouse embryonic stomach, glandular epithelial hyperplasia occurs. Furthermore, this phenotype was mimicked in Gli3−/−, but not Gli2−/− stomachs. I utilized three additional mouse models that modulate Gli3 activity to better understand the role of Gli3 in the developing stomach - the Gli3Δ699/Δ699 ,Gli3P1−4/P1−4, and Kif7−/− mice. The Gli3P1−4/P1−4 stomach displayed glandular epithelial overgrowth, as did the Kif7−/− stomach to a lesser extent; the Gli3Δ699/Δ699 stomach displayed glandular hypoplasia. Moreover, the Gli3P1−4/P1−4 and Kif7−/− stomachs have a thicker circular smooth muscle, and the Gli3Δ699/Δ699 had a thinner one relative to wild-type. It appears that altering the balance of Gli3 in favour of its activator results in gastric glandular epithelial and circular smooth muscle hyperplasia, and a balance favouring the Gli3 repressor results in hypoplasia.

The strength of time-dependent correlations known as stride interval (SI) dynamics have been proposed as an indicator of neurologically healthy gait. Most recently, it has been hypothesized that these dynamics may be necessary for gait efficiency although the supporting evidence to date is limited. To gain a better understanding of this relationship, this study investigated stride interval dynamics, the energy cost of walking, and physical activity in a pediatric population. The findings indicate that differences in energy cost are not reflected in the stride interval dynamics of able-bodied children. Interestingly, increasing physical activity levels were associated with decreasing variance in stride interval dynamics between subjects, though this finding only approached significance (p=0.054). Lastly, this study found that stride interval dynamics in children as young as nine years were comparable to stride interval dynamics found in healthy young adults.

The Sonic hedgehog (Shh) signaling pathway plays a critical role in murine gastric development. When Shh is knocked out in the mouse embryonic stomach, glandular epithelial hyperplasia occurs. Furthermore, this phenotype was mimicked in Gli3−/−, but not Gli2−/− stomachs. I utilized three additional mouse models that modulate Gli3 activity to better understand the role of Gli3 in the developing stomach - the Gli3Δ699/Δ699 ,Gli3P1−4/P1−4, and Kif7−/− mice. The Gli3P1−4/P1−4 stomach displayed glandular epithelial overgrowth, as did the Kif7−/− stomach to a lesser extent; the Gli3Δ699/Δ699 stomach displayed glandular hypoplasia. Moreover, the Gli3P1−4/P1−4 and Kif7−/− stomachs have a thicker circular smooth muscle, and the Gli3Δ699/Δ699 had a thinner one relative to wild-type. It appears that altering the balance of Gli3 in favour of its activator results in gastric glandular epithelial and circular smooth muscle hyperplasia, and a balance favouring the Gli3 repressor results in hypoplasia.