Effect of the genetic background and specific mutation on adenylate cyclase activity in obesity syndromes

Adenylate cyclase activity and its modulation by guanine nucleotides and isoproterenol were assessed in adipocyte membranes of mice with mutations causing different genetic obesity syndromes. The object was to determine whether the defect in inhibitory modulation observed in the obese (ob/ob) mouse was also present in the diabetes (db/db) mouse. The data show that adipocyte adenylate cyclase in both the ob/ob and the db/db mouse is resistant to activation by isoproterenol. The response to guanosine triphosphate (GTP) differed between the two mutants, such that an inhibitory phase was visible in the db/db but not in the ob/ob membranes. Moreover, pertussis toxin attenuated the inhibitory effect of GTP and significantly stimulated cyclase activity in the db/db but not in the ob/ob membranes. The data show that the two mutations affect the expression of adenylate cyclase activity via different mechanisms.     

Hypothalamo-pituitary regulation of the c-myc gene in rat liver

Expression of the c-myc gene was studied in the livers of male and female Wistar rats. Furthermore, the effects on hepatic c-myc expression of neonatal and adult castration, with or without testosterone supplementation, as well as of continuous administration of GH to intact males, were analysed. Expression of c-myc was low in 6-day-old animals of both sexes, reached a maximum at 35 days of age and declined to the level of adult animals at 70 days. In prepubertal animals, expression was higher in females, but was higher in males after the onset of puberty, the postpubertal female rat liver exhibiting 50-70% of the expression in males. Treatment of adult male rats with bovine GH in osmotic minipumps for 1 week reduced c-myc expression to the level of female rats. Castration, both neonatally and of adults, also feminized hepatic c-myc expression. Testosterone supplementation of the castrated animals increased the expression towards the level in sham-operated controls. These results indicate that the c-myc gene is regulated by the hypothalamo-pituitary-liver axis via the sex-differentiated pattern of GH secretion, in analogy with other sex-differentiated hepatic functions, such as metabolism of steroids and xenobiotics. Neuroendocrine regulation of a gene such as c-myc, which is involved in the control of cell proliferation and differentiation, represents another aspect of the complex influence of GH on various somatic functions.     

Tissue-specific expression and developmental regulation of the rat apolipoprotein B gene.

Expression of the apolipoprotein B (apoB) gene was examined in a variety of fetal, neonatal, and adult rat tissues by probing RNA blots with a cloned rat apoB cDNA. Among 10 adult male tissues surveyed, small intestine had the highest concentration of apoB mRNA. Its abundance in liver and adrenal gland was 40% and 0.5%, respectively, of that in small bowel, while none was detected in colon, kidney, testes, spleen, lung, heart, or brain. ApoB mRNA is as abundant in 18-day fetal liver as at any subsequent period of hepatic development. In contrast, the concentration of apoB mRNA remains low in fetal intestine until the last (21st) day of gestation, when it increases sharply to levels that are several-fold higher than in the liver. ApoB mRNA levels in fetal membranes harvested during this late gestational period were 10 times greater than in fetal liver. Since the major lipoprotein species in 19-day fetal plasma is low density lipoprotein, these observations suggest that fetal liver, and particularly its functional homologue, the yolk sac, are the principal sites of fetal lipoprotein synthesis at this stage of development. A 20-fold increase in placental apoB mRNA concentrations during the last 48 hr of pregnancy (to a level that is 50% of that encountered in fetal membrane RNA) suggests a specific role for this organ in maternal-fetal lipid transport immediately prior to parturition. Pulse-labeling experiments using 21-day fetal tissue slices showed that the liver synthesizes both apoB-100 (B-PI) and apoB-48 (B-PIII) albeit in somewhat different ratios than the adult organ. Fetal intestine produces almost exclusively the smaller apoB species, while fetal membranes and placenta synthesize only the larger peptide. The postnatal pattern of apoB mRNA accumulation is similar in liver and intestine. Profound decreases were observed during the late suckling and weaning periods, followed by an increase to adult levels. These final concentrations were similar to those encountered at birth. Analysis of these developmental changes offers an opportunity to generate testable hypotheses about the factors that modulate apoB synthesis.     

The effect of old age on apolipoprotein E and its receptors in rat liver

Apolipoprotein E (apoE) is associated with aging and some age-related diseases. The majority of apoE is produced by hepatocytes for the receptor-mediated uptake of lipoproteins. Here, the effects of age on the hepatic expression and distribution of apoE and its receptors were determined using immunofluorescence, Western blots, and quantitative PCR in rat liver tissue and isolated hepatocytes. The expression of apoE mRNA and protein was not influenced significantly by aging. Immunofluorescence studies in isolated hepatocytes showed that apoE was more likely to be co-localized with early endosomes, golgi, and microtubules in isolated old hepatocytes. The mRNA expression of the receptor involved in sequestration of apoE, heparan sulfate proteoglycan was reduced in old age, without any significant effect on the expression of either the low-density lipoprotein receptor or low density-lipoprotein receptor-related protein. Old age is associated with changes in hepatic apoE intracellular trafficking and heparan sulfate proteoglycan expression that might contribute to age-related disease.     

Abnormal regulation of the leptin gene in the pathogenesis of obesity

A subset of obese humans has relatively low plasma levels of leptin. This finding has suggested that in some cases abnormal regulation of the leptin gene in adipose tissue is etiologic in the pathogenesis of the obese state. The possibility that a relative decrease in leptin production can lead to obesity was tested by mating animals carrying a weakly expressed adipocyte specific aP2-human leptin transgene to C57BL/6J ob/ob mice (which do not express leptin). The transgene does not contain the regulatory elements of the leptin gene and is analogous to a circumstance in which the cis elements and/or trans factors regulating leptin RNA production are abnormal. The ob/ob mice carrying the transgene had a plasma leptin level of 1.78 ng/ml, which is ≈one-half that found in normal, nontransgenic mice (3.72 ng/ml, P < 0.01). The ob/ob animals expressing the leptin transgene were markedly obese though not as obese as ob/ob mice without the transgene. The infertility as well as several of the endocrine abnormalities generally evident in ob/ob mice were normalized in the ob/ob transgenic mice. However, the ob/ob transgenic mice had an abnormal response when placed at an ambient temperature of 4°C, suggesting that different thresholds exist for the different biologic effects of leptin. Leptin treatment of the ob/ob transgenic mice resulted in marked weight loss with efficacy similar to that seen after treatment of wild-type mice. In aggregate these data suggest that dysregulation of leptin gene can result in obesity with relatively normal levels of leptin and that this form of obesity is responsive to leptin treatment.     

Concentration-affinity equivalence in gene regulation: convergence of genetic and environmental effects.

It is proposed that equivalent phenotypic effects can be obtained by either structural changes in macromolecules involved in gene regulation or changes in activity of the structurally unaltered macromolecules. This equivalence between changes in activity (concentration) and changes in structure can come into play within physiologically plausible limits and seems to represent an important interface between environment and genome--namely, between environmentally determined and genetically determined gene expression. The equivalence principle helps explain the appearance of phenocopies. It also points to a general pathway favorable to the occurrence, during evolution, of frequent episodes corresponding to Waddington's genetic assimilation and is likely to represent one component of the system responsible for the high frequency of recurrence of parallel evolution.     

The influence of genetic background on the expression of the obese (ob) gene in the mouse

Summary A new congenic strain of obese mice, C57BK/KsJ-ob, has been developed for comparison with the C57BL/6J-ob congenic strain. While obese mice of both strains are characterized by obesity, hyperphagia, and hyperglycemia, the C57BL/Ks obese mice have severe diabetes, marked hyperglycemia, temporarily elevated plasma insulin concentrations, and typical degenerative changes in the islets of Langerhans. In contrast, the C57BL/6J obese mice have mild hyperglycemia and marked hyperinsulinemia coupled with hypertrophy and hyperplasia of the islets of Langerhans. The severe diabetic condition produced by obese (ob) on the C57BL/KsJ background is similar, if not identical, to that produced by the diabetes (db) gene on the same background. The metabolic disorder produced by these mutations is associated with the capacity of the islets to respond to an increased demand for insulin. The islet response, whether atrophy or hypertrophy, appears to be due to the interaction of the obese and diabetes genes with modifiers in the genetic background rather than the specific consequences of the particular gene. The markedly different diabetic syndromes that result when the obese mutation is on different genetic backgrounds emphasize the importance of strict genetic control in studies with obese-hyperglycemic mutants.Supported in part by NIH Research Grants AM 14461 from the National Institute of Arthritis, Metabolism, and Digestive Diseases; CA 05873 from the National Cancer Institute and allocations from the South Waite Foundation and the Virginia and D.K. Ludwig Foundation.The Jackson Laboratory is fully accredited by the American Association for Accreditation of Laboratory Animal Care.     

Assessment of genetic linkage and parent-of-origin effects on obesity

CONTEXT: Obesity is a growing health care problem worldwide and is a major underlying risk factor for common diseases such as diabetes. Parent-of-origin effect has been reported to be involved in the development of obesity. But the genes with imprinting effects related to obesity are largely unknown. OBJECTIVE: The objective of the study was to identify obesity-related genetic loci, both with and without imprinting effects. DESIGN AND SUBJECTS: We conducted genome-wide linkage analyses for obesity with and without consideration of imprinting effects in a large sample including more than 4000 individuals. In addition to body mass index (BMI), we also used a more stringent and accurate obesity definition, which simultaneously considers BMI and percentage of fat mass (PFM) in a gender-specific manner. Simulations were performed to identify the genome-wide significant and suggestive significant thresholds. RESULTS: In nonimprinted linkage analyses, we detected suggestive linkage at 2q31 (LOD = 2.23) and 16q22 (LOD = 1.87) for BMI and 2q37 (LOD = 2.23) for BMI and PFM. Interestingly, 2q37 also achieved a significant maternal linkage with BMI and PFM (LOD=3.34) in imprinted linkage analyses. Imprinted linkage analyses revealed suggestive linkage evidence for BMI at three additional genomic regions, including 3p14 (LOD = 2.20, paternal), 3q24 (LOD = 1.97, maternal), and 19q13 (LOD = 1.81, maternal). CONCLUSION: We reported linkage and imprinting effects for obesity on several chromosome regions and suggested the potential importance of parent-of-origin effects and phenotype definition of obesity in delineating the genetic basis of obesity.     

Impact of apolipoprotein E genetic polymorphisms on liver disease: An essential review

Cirrhosis is an advanced stage of liver disease, compromising liver function with systemic health implications and poor quality of life. Hepatitis C virus (HCV) infection and alcoholic liver disease are the main causes of this pathology. However, since genetic factors may play a large role in the progression and severity of liver disease, and as apolipoprotein E (apoE) has been recognised to be mainly synthesised in the liver, apoE polymorphism studies are important to better understand the causal mechanisms in liver diseases. In this review, we summarise up-to-date studies addressing how apoE polymorphisms influence liver cirrhosis and liver transplantation outcomes and potential protective mechanisms. Although more clinical studies are needed to support these findings, the apoE ɛ4 allele seems to be protective against the progression of liver cirrhosis in the majority of aetiologies and the postoperative serum apoE phenotype of the transplanted subject receptors was converted to that of the donor, indicating that >90% of apoE in plasma is synthesised in the hepatic system.     

The effects of fat feeding on apolipoprotein AI secretion from rat small intestinal epithelium

The small intestine is known to be an important synthetic site for certain apolipoproteins, which are subsequently secreted from the enterocyte into the mesenteric lymph. We have studied apolipoprotein AI and CIII content of the enterocyte during the course of fat feeding in order to determine their relative synthetic and secretory rates. Rat intestinal enterocytes were isolated from the entire jejunal villus after fat feeding in vivo. The apo AI content fell 50% as determined by RIA one and two hours after fat feeding. By four hours, the intracellular cellular levels had returned to prefeeding levels. These changes in apolipoprotein AI levels were not seen in the terminal ileum. Apolipoprotein CIII levels remained unchanged afer fat feeding. To determine the effect of free fatty acids on apolipoprotein AI secretion, organ culture explants were incubated for four hours in the presence and absence of 360 microM oleic acid bound to albumin. Apolipoprotein AI detected in the incubation media reflected release from the lamina propria (which was not colchicine sensitive), and secretion from the enterocyte (which was inhibited by colchicine). In the absence of oleic acid, enterocyte secretion of apolipoprotein AI accounted for about half of the apo AI recovered in the medium. In the presence of oleic acid, the total apolipoprotein AI content of the tissue increased by 50 percent. A similar increase in colchicine sensitive secretion was observed. The secretion of apolipoprotein AI from explants was more rapid in the presence of oleic acid and began without the half hour lag noted when oleic acid was absent. The mid intestine was most active in the secretion of apolipoprotein AI. These data are consistent with the hypothesis that in the first few hours after feeding the rate of secretion of apolipoprotein AI exceeds the synthetic capacity of the small intestinal epithelium.