Gestational hypoxia and programming of disease

Abstract

Fetal hypoxia remains the most common complication throughout pregnancy and has a wide aetiology including, but not limited to, placental insufficiency, high altitude living and maternal factors such as smoking and pulmonary disease. Intrauterine growth restriction and subsequent low birth weight are common features of pregnancies complicated by reduced oxygenation. Suboptimal organ development may arise, thereby substantially increasing vulnerability to cardiovascular and renal diseases in adulthood. Recently it has been established that prenatally programmed vulnerability to disease may be exacerbated by a postnatal ‘second-hit’, such as a high salt diet or ageing. The primary aim of this thesis was to investigate the effects of maternal hypoxia on the programming of cardiovascular and renal disease in mice, and secondly whether excess dietary salt intake throughout life may exacerbate disease outcomes. Pregnant CD1 mice were housed in a hypoxia chamber set to 12% oxygen throughout the last five days of pregnancy, from embryonic day (E) 14.5 until birth (P0). Control animals remained in normoxic room conditions (21% oxygen) throughout pregnancy. Upon littering, animals were removed from the hypoxia chamber and raised in normoxic room conditions. A subset of mice was fed a high salt diet from 10 weeks of age until post-mortem. Mice were housed in metabolic cages for 24 hours to assess renal function under basal conditions, and in response to a 24 hour water deprivation challenge, at 4 and 12 months of age. Blood pressure and microvascular function and structure were assessed in offspring at 12 months of age. Kidneys, hearts and aortas were collected for stereological and histological analyses. Prenatal hypoxia reduced nephron number in the kidneys of male offspring, leading to glomerular hypertrophy, renal fibrosis and mild albuminuria by 12 months of age. However, female offspring exposed to the same hypoxic insult in utero presented with normal number of glomeruli and renal pathology equivalent to control animals by 12 months of age. Both male and female hypoxia-exposed offspring had elevated mean arterial pressure at 12 months of age. A high salt diet throughout adult life increased cardiac tissue fibrosis, particularly in male hypoxia-exposed offspring, and exacerbated renal pathology in male offspring only. Pressurized myography was performed at the time of post-mortem to assess functional, structural and mechanical properties of mesenteric resistance arteries at 12 months of age. Both male and female offspring exposed to maternal hypoxia had significantly impaired endothelial function, which was not exacerbated by the high salt diet. The combination of prenatal hypoxia and a postnatal high salt diet caused marked stiffening of the microvasculature as well as loss of elastin integrity and increased collagen content in the aorta, consistent with vascular stiffness. This suggests that kidneys from female offspring are somehow protected from the in utero insult; however, this protection does not extend to the cardiovascular system. We also investigated the impact of prenatal hypoxia on the renal tubule system and collecting ducts. First, we developed a methodology to estimate lengths of renal tubules and collecting ducts using a combination of immunohistochemistry and design-based stereology. We have demonstrated that the renal tubules and collecting ducts undergo significant elongation postnatally. Furthermore, salt-induced renal hypertrophy was associated with elongation of the thin descending limb of Henle and the proximal tubule. We applied this technique to the kidneys of male hypoxia-exposed offspring with a congenital nephron deficit, and observed significant elongation of the proximal and distal tubule compared to controls. Male hypoxia-exposed offspring had an altered response to a 24 h water deprivation challenge at 12 months of age, which was associated with altered cellular composition of the collecting duct and reduced Aquaporin 2 expression and distribution in the kidney. This highlights that renal tubule segments and collecting ducts are susceptible to prenatal insult and may, in combination with reduced nephron number, contribute to the predisposition to cardiovascular and renal disease. In conclusion, this thesis has demonstrated that prenatal hypoxia increases vulnerability of offspring, particularly males, to cardiovascular and renal disease in adulthood. Furthermore, a postnatal diet high in salt exacerbated microvascular stiffness, cardiac fibrosis and renal histopathology, suggesting that consumption of a healthier diet may attenuate predisposition to disease. This indicates that the postnatal environment is an important consideration in disease progression, particularly for those born of low birth weight.