Comparison of apoptosis related gene expression in p7-p14 mouse lenses

Lens development exhibits a seamless progression from embryogenesis through ageing. Lens maturation and growth proceed throughout life; lens fibre cells differentiate from lens epithelial cells at the equator of the lens, migrate inwards at the bow region and mature around pre-existing fibre cells at the centre of the lens. Lens fibre cells lose their nuclei via an apoptosis like mechanism as they mature. We therefore decided to study further the relationship of apoptosis related gene expression during vital stages of lens maturation in the mouse. This is an important question as defects in lens function and/or maturation result in clouding of the lens, or cataract, which can result in considerable visual impairment. Keywords: development/age comparison We used manufactured Sigma Panorama mouse apoptosis gene arrays (cat# G1039), in conjunction with RNA extracts from day 7 and day 14 postnatal mice. These stages were chosen because the eyes are closed at day 7, but open on day 14. The postnatal period before day 14 is a period of rapid lens growth, accompanied by lens fibre cell differentiation and organelle loss via apoptosis signalling pathways. Formation of the organelle free zone is complete at day 14 when the eyes open, allowing for clear vision. 4 biological repeats were carried out for day 7, and 3 for day 14 lenses. Each array carries duplicate spots, giving 8 and 6 repeats respectively. Analysis of all day 7 repeats versus all day 14 repeats was carried out by t-test, we also required a fold change of 2. Statistically significant genes were tested by PCR using 3 biological replicates for each stage. We also used the array data to estimate which genes were highly expressed at each stage. We asked that at least one of the spots representing each gene be above background + 2SD in all replicates for a given stage (day7 or day14).