Transgenic And Knockout Animals In Studies Of Food Intake Body Composition And Obesity

In addition to the studies of GH described earlier in this chapter, transgenic and targeted gene-disruption technologies are used to elucidate the mechanisms controlling food intake, body composition, and the pathogenesis of obesity. Within the past few years, the applications of these genetic techniques in whole animals have identified a number of new molecules and physiologic pathways involved in the regulation of body wt, and have led to important new insights into the pathophysiology of eating disorders and obesity. These include the hormone leptin, the short and long forms of the leptin receptor, uncoupling proteins, agouti and agouti-related proteins, melanocortin-recep-tor isoforms, melanin-concentrating hormone, orexin, mahogany, and the proteins responsible for tub and fat, two monogenic mouse models of obesity. These efforts, in addition to characterization of several spontaneous obese mutants, provided a number of novel genetic models in which a single gene or pathway has been experimentally modified to test specific hypotheses. Both the spontaneous and experimentally generated mutants, or their intercrosses, are remarkably useful models in elucidating the

Table 1

Transgenic Models with Alterations in GH Release or Action^

Transgenic animals

GH-transgenic mice with MT or PEPCK promoters (1-3,5,15)

GH-transgenic mice with adenoviral vectors (19,20)

GH-transgenic rats (17)

GHRH-transgenic mice (54)

GH-ablated mice (90)

Growth-retarded transgenic rats (93)

GH-antagonist transgenic mice (96)

Phenotypes

Expression of heterologous (human, bovine, ovine, rat) GH genes in the liver, kidney, and other organs leads to increases in plasma IGF-I, growth and adult body size, while release of endogenous GH is suppressed. Synthesis and release of GH does not depend on physiological control mechanisms (such as GHRH and SRIF) but can be modified by composition of the diet, according to the properties of the employed promoter.

This form of "gene therapy" was successful in achieving transient or prolonged expression of mouse or rat GH in genetically GH-deficient mice and in partially correcting their phenotypic defects.

Ectopic overexpression of human GH under control of mWAP promoter in transgenic rats produced phenotypic characteristics similar to those of MT-hGH and PEPCK-hGH transgenic mice.

Overexpression of GHRH produces stimulation and hyperplasia of somatotrophs, progressive enlargement of the anterior or pituitary, and increases in somatic growth and adult body size. This transgenic model allows study of the effects of excessive secretion of homologous (mouse) GH of eutopic (pituitary) origin.

Transgenic expression of DTA-chain structural gene under control of rat GH promoter led to selective destruction of somatotrophs, absence of GH in the circulation, and reduced growth.

Targeted expression of hGH in the hypothalamic GHRH neurons of transgenic rats reduced GHRH expression, presumably via negative GH feedback. The animals had fewer somatotrophs, reduced GH levels and dwarf phenotype.

Overexpression of antagonistic GH analogs under control of MT promoter in transgenic mice reduces growth and adult body size, apparently by interfering with the action of endogenous GH and producing a state of GH resistance.

Table 1 (continued)

Transgenic animals Phenotypes

GH-R/GHBP knockouts (102) Targeted disruption of the GH receptor/

GH-binding protein gene in mice produces GH resistance. Plasma GH levels are elevated, while the levels of IGF-I, postweaning growth, and adult body size are drastically reduced.

information on GH transgenics in species other than mice and rats (including domestic animals and fish) is outside the scope of this chapter.

physiology of energy metabolism, nutrient partition, the pathophysiology of obesity, and in the development of novel anti-obesity drugs.

Studies using these models lead to the following conclusions: (1) Genetic and hormonal control of food intake and body composition is extremely complex and involves interaction of multiple regulatory loops; (2) There is a great deal of redundancy in this controlling system and, similarly to many other complex biological systems, this redundant assemblage appears to provide a reliable physiological mechanism to meet an organism's constant energy needs for growth, reproduction, and maintenance; and (3) Obesity, an easily detected phenotypic indicator, is generally not a one-gene/one-disease event, but rather a result of diverse underlying metabolic and physiologic dysregulations involving a number of different molecules and independent pathways.

The intricate interactions between various hormones involved in the control of food intake and obesity are well illustrated by the relationships between GH and leptin, or between novel mahogany peptide, the agouti peptide, alpha-melanocyte-stimulating hormone (a-MSH), and melanocortin receptor 4 (MC4R). Through its potent lipolytic action, GH reduces adipose tissue mass, the source of leptin. Moreover, GH can reduce leptin expression (as measured by leptin mRNA levels) in pig adipocytes (114). Pathologic overproduction of GH in acromegalic patients is associated with reduced levels of leptin (115). In turn, leptin can affect GH release (116,117). This includes stimulation of both basal and GHRH-induced GH release (116), and blocking the inhibitory effect of fasting on the release of GH (118). Interestingly, both starvation and obesity are associated with the suppression of circulating GH levels (119,120). Chronic caloric restriction in rats prevents age-related decline in pulsatile GH release (121).

Similarly complex interactions of various molecules and receptors have been recently discovered in the hypothalamic melanocortin signaling system, which is also involved in the control of feeding. Melanocortin signaling transduced by MC4R tonically inhibits feeding (122). The agouti protein that is normally expressed in the hypothalamus (123) acts as an MC4R antagonist (124). Increased expression of the agouti protein results in increased appetite and obesity (125). MC4R-gene knockout produces a similar phenotypic change (126). However, for the agouti protein to antagonize the MC4R signaling, the mahogany (attractin) protein is required (127,128). It has been proposed that the role of the mahogany gene product in this pathway is to present agouti antagonist to MC4R, thereby reducing signaling. Thus, increased expression of mahogany would result in increased agouti binding and, consequently, increased appetite and body wt. Alterations in melanocortin signaling also affect the NPY-leptin pathway (129,130).

As shown in the following sections, the discoveries of these intricate interrelationships would not be possible without transgenic and gene-knockout animal models.

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