The effect of maternal nutrition on the developmental origins of respiratory disease

Abstract

Environmental challenges during early life have been shown to result in greater risk of chronic diseases such as diabetes and coronary disease in later life. Factors such as unbalanced nutrition before birth result in metabolic and structural adaptations that lead to persistent modifications to offspring phenotype. There is evidence that respiratory disease is influenced by developmental environment. Reduced fetal growth is associated with impaired lung development, increasing risk of developing asthma and Chronic Obstructive Pulmonary Disease (COPD) in later life. To investigate the mechanisms underlying these effects, it is necessary to utilise animal models that emulate the phenotypes observed in human studies.
The aim of this thesis was to investigate whether exposure to a maternal low protein diet in utero affects offspring lung morphology, bronchial hyperresponsiveness (BHR) and global methylation and/or gene expression in Wistar rats. Pregnant Wistar rats were allocated to either control (C, 18% casein) or protein restricted (PR, 9% casein) diet. Lung tissue was harvested (225 days) from male offspring (F1, (28 days, C=15; PR=10, 120 days, C=11; PR=7, 225 days: C=6; PR=6) F2, (28 days, C=24; PR=17) and F3 (21 days, C=16; PR=5)). Lungs were removed and the left lung was sectioned and haematoxylin and eosin stained, imaged at 10x magnification and using stereological methods, point counted to give volume fractionation of selected areas. Other morphological measurements were made to estimate surface area (10x magnification), and alveolar wall thickness (63x magnification). Primary bronchi were dissected out, mounted in a myograph and bronchoconstriction in response to a range of bronchoconstrictors and bronchodilators was assessed. RNA and DNA were extracted using TRIzol ® and global methylation assessed using MethylampTM Global DNA Methylation Quantification Ultra Kit.
There was no significant difference in either lung volume or lung structure between the F1 or F2 offspring of control of PR exposed mothers except for a significant increase (p=0.046) in the amount of smooth muscle around blood vessels in the protein restricted group in the F1 225 day old group. However, in assessment of constrictor responses of isolated bronchi, a significant difference in response between groups was found with both carbachol, an acetylcholine mimetic, and U44619, A thromboxane mimetic, with the F1 offspring from protein restricted mothers exhibiting significantly increased BHR at 75 days of age (p=<0.001) compared to controls. No significant difference in global DNA methylation in lung tissue was found between F1 of F2 offspring of mothers exposed to the control of PR diet during pregnancy. Gene expression levels of selected candidate genes in lung tissue were then assessed using qPCR and again no differences were found between groups.
In this study there is no evidence to suggest that in utero exposure to a maternal low protein diet has affected offspring lung physiology, structure, methylation patterns or gene expression. However, there could be differences in the amount of smooth muscle found around the vessels in response to this nutritional challenge as protein restricted animals appear to have more smooth muscle compared to controls. BHR was observed in the 75 day old rats indicating a possible shift towards a priming of asthma phenotype which could be induced with post natal allergen challenges.

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Identifiers

ORCID for John W. Holloway: orcid.org/0000-0001-9998-0464

ORCID for Susan J. Wilson: orcid.org/0000-0003-1305-8271

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