Apolipoprotein B and triacylglyceride physiology in the shortfinned eel, Anguilla australis.

Abstract

Lipids play an essential role in reproduction, especially in oviparous species such as teleost fish, in which the developing embryo does not receive nutrients from the mother after fertilisation. To date, mammalian paradigms have been applied to understanding lipid physiology and transport in teleost fish; however, the fact that notable differences exist in fish lipid biology (i.e. high blood lipid levels, absence of albumin in the eel and accumulation of large amounts of lipids into the gamete) is likely to be reflected in transport mechanisms and their regulation. This thesis aimed to explore several of these paradigms in-depth and shed light on the similarities and differences between human and teleost fish lipid physiology. Firstly, a specific antiserum for apolipoprotein B (apoB) was developed (Chapter Two) and subsequent analysis suggested the presence of only one form of apoB in the shortfinned eel (Anguilla australis). This finding was reinforced by inferring the evolutionary relationships of apoB and the apoB mRNA editing enzyme (apoBEC1) in a number of vertebrates (Chapter Three). The presence of a single apoB was also linked to the evident hyperlipidemia seen in lower vertebrates. Secondly, the fasting-induced mobilisation of stored lipids and their subsequent re-packaging in the liver was examined in eels in different stages of early oogenesis (Chapter Four). The changes in relative transcript copy numbers of intestinal and hepatic triacylglyceride (TAG) packagers (namely apoB and microsomal triacylglyceride transport protein (MTTP)) as well as the TAG receivers and modifiers (apolipoprotein E (apoE), lipoprotein lipase (LPL), fatty acid transport protein one (FATp1), vitellogenin receptor (vtg-R) and low density lipoprotein receptor (LDL-R)) in the ovary were followed using quantitative polymerase chain reaction (qPCR). Relative transcript copy numbers of the intestinal packagers decreased around the same stage as the relative transcript copy numbers of the hepatic packagers and ovarian receivers increased. The importance of androgens during early reproduction in multiple vertebrate species prompted the effects of androgens on the above mentioned packaging and uptake to be evaluated (Chapter Five). Levels of 11-ketotestosterone and 17-methyltestosterone were artificially raised in two separate experiments and qPCR results showed that the majority of the changes observed mirrored those seen in the wild-caught eels, indicating their possible induction by androgen treatment. Lastly, the involvement in ovarian TAG uptake of two receptors (vtg-R and LDL-R) indicated to be important in TAG uptake was examined in vitro. Antisera were used to block these receptors and oil droplet accumulation was examined in the ooplasm of the ovarian explants after 21 days of incubation. Blocking the LDL-R significantly reduced the accumulation of oil droplets, whilst blocking the vtg-R appeared to have no effect. These results indicate that, as with other oviparous vertebrates (and in contrast to humans) receptor-mediated endocytosis is likely to be the main pathway of TAG uptake into the ovary. This thesis highlighted several important evolutionary and physiological differences between mammals and lower vertebrates and has considerably advanced the field as a whole.