Michael W. Bradbury

Uptake of Long Chain Free Fatty Acids Is Selectively Up-regulated in Adipocytes of Zucker Rats with Genetic Obesity and Non-insulin-dependent Diabetes Mellitus

To examine whether fatty acid transport is abnormal in obesity, the kinetics of [3H]oleate uptake by hepatocytes, cardiac myocytes, and adipocytes from adult male Wistar (+/+), Zucker lean (fa/+) and fatty (fa/fa), and Zucker diabetic fatty (ZDF) rats were studied. A tissue-specific increase in oleate uptake was found in fa/fa and ZDF adipocytes, in which the Vmax was increased 9-fold (p < 0.005) and 13-fold (p < 0.001), respectively. This increase greatly exceeded the 2-fold increase in the surface area of adipocytes from obese animals, and did not result from trans-stimulation secondary to increased lipolysis. Adipocyte tumor necrosis factor-α mRNA levels, assayed by Northern hybridization, increased in the order +/+ < fa/fa < ZDF. Oleate uptake was also studied in adipocytes from 20-24-day-old male +/+, fa/+, and fa/fa weanlings. These animals were not obese, and had equivalent plasma fatty acid and glucose levels. Tumor necrosis factor-α mRNA levels in +/+ and fa/fa cells also were similar. Nevertheless, Vmax was increased 2.9-fold (p < 0.005) in fa/fa compared +/+ cells. These studies indicate 1) that regulation of fatty acid uptake is tissue-specific and 2) that up-regulation of adipocyte fatty acid uptake is an early event in Zucker fa/fa rats. These findings are independent of the role of any particular fatty acid transporter. Adipocyte mRNA levels of three putative transporters, mitochondrial aspartate aminotransferase, fatty acid translocase, and fatty acid transporting protein (FATP) were also determined; mitochondrial aspartate aminotransferase and FATP mRNAs correlated strongly with fatty acid uptake.

Selective up-regulation of fatty acid uptake by adipocytes characterizes both genetic and diet-induced obesity in rodents.

Long chain fatty acid transport is selectively up-regulated in adipocytes of Zucker fatty rats, diverting fatty acids from sites of oxidation toward storage in adipose tissue. To determine whether this is a general feature of obesity, we studied [3H]oleate uptake by adipocytes and hepatocytes from 1) homozygous male obese (ob), diabetic (db), fat (fat), and tubby (tub) mice and from 2) male Harlan Sprague-Dawley rats fed for 7 weeks a diet containing 55% of calories from fat. V max andK m were compared with controls of the appropriate background strain (C57BL/6J or C57BLKS) or diet (13% of calories from fat). V max for adipocyte fatty acid uptake was increased 5–6-fold in ob, db, fat, and tub mice versus controls (p < 0.001), whereas no differences were seen in the corresponding hepatocytes. Similar changes occurred in fat-fed rats. Of three membrane fatty acid transporters expressed in adipocytes, plasma membrane fatty acid-binding protein mRNA was increased 9–11-fold in ob and db, which lack a competent leptin/leptin receptor system, but was not increased in fatand tub, i.e. in strains with normal leptin signaling capability; fatty acid translocase mRNA was increased 2.2–6.5-fold in tub, ob, and fatadipocytes, but not in db adipocytes; and only marginal changes in fatty acid transport protein 1 mRNA were found in any of the mutant strains. Adipocyte fatty acid uptake is generally increased in murine obesity models, but up-regulation of individual transporters depends on the specific pathophysiology. Leptin may normally down-regulate expression of plasma membrane fatty acid binding protein.

Cellular uptake of long chain free fatty acids: the structure and function of plasma membrane fatty acid binding protein

Publisher Summary Non-esterified long chain fatty acids (LCFA) are important energy substrates, building blocks for the complex lipid components of cellular membranes, and precursors of biological mediators, such as the prostaglandins and leukotrienes. The chapter discusses the mechanisms of cellular uptake of long chain fatty acids. The apparent saturability of LCFA uptake challenged the long-held view that LCFA uptake occurred solely by passive diffusion. Saturability is a necessary, but not sufficient, condition to establish the existence of facilitated transport. The significance of putative LCFA transport systems lies in the roles they play in human physiology, pathophysiology, and disease. The chapter summarizes adipocytes and HepG2 cells that suggests important roles for regulated plasma membrane expression of mAspAT in LCFA transport systems that contribute to the pathogenesis of obesity, type 2 diabetes, and EtOH-associated fatty liver, whereas identification of mAspAT, FAT, and an FATP in particular human placental plasma membranes suggests that they are important for transplacental LCFA transport. Facilitated transport mechanisms increasingly appear to be critical providers of energy substrates to skeletal and cardiac muscle. Finally, a selective LCFA uptake defect is reportedly the basis of a syndrome of recurrent bouts of nonketotic, hypoglycemic acidosis and liver failure requiring transplantation in children. Current work in the field of fatty acids transport provides convincing evidence that cellular LCFA is a regulatable, protein mediated process of considerable, potential clinical importance.

Genomic imprinting : a gene regulatory phenomenon with important implications for micromanipulation-assisted in vitro fertilization (IVF)

1 Molecular Biology, Mt. Sinai Medical Center, P.O. Box 1175, New York, New York 10029. 2 Ob/Gyn & Reproductive Science, Mt. Sinai Medical Center, P.O. Box 1175, New York, New York 10029. 3 Geriatrics and Adult Development, Mt. Sinai Medical Center, P.O. Box 1175, New York, New York 10029. sources to their effort to conceive. These circumstances define two important concerns with respect to management of infertility in this patient group. First, it is highly desirable to identify those individuals for whom even IVF is insufficient and who must consequently seek adoption. Second, it is of utmost importance to maximize the opportunity of achieving fertilization and implantation once an IVF cycle is undertaken. The following discussion focuses on the second of these two imperatives by describing a new potential approach to increasing the chance of conception after oocyte retrieval and insemination. The strategy involves micromanipulation of the fertilized egg and is based upon the fact that 5-10% of zygotes produced in IVF cycles are polyspermic. While the fate of polyspermic zygotes in vivo is not known with certainty, it must be assumed that the presence of two, or, rarely, more than two, male haploid genomes in the zygote is inconsistent with normal development. While a 5-10% rate of polyspermy superficially appears acceptable, it must be recognized that some IVF cycles will result in a single fertilization event, with that event characterized by polyspermy. In such an instance, the chance of conception is lost completely. A more common circumstance is that two or three oocytes will be fertilized, one of which will be polyspermic. In that case the number of embryos available for transfer will be reduced, with an attendant significant reduction in the likelihood of achieving a pregnancy. The micromanipulation procedure discussed as a potential treatment for polyspermy is pronucleus removal. In theory it should be possible to extract supernumerary male pronuclei from polyspermic zygotes, thus restoring the normal diploid number of chromosomes. However, recent experiments with animals indicate that if the single female pronucleus is removed, leaving behind two genomes derived from the male parent, the conceptus will not survive, and serious pathological consequences may ensue. These experiments have collectively identified a phenomenon known as genomic imprinting, a gene modification system which confers distinct regulatory characteristics upon a subset of

Rapid Sexing of Preimplantation Mouse Embryos by Blastomere Biopsy and Enzymatic Amplification

When infertile couples reach the point in their evaluation where in vitro fertilization (IVF) is considered as a remedy, they have usually exhausted all other modes of treatment In the case of the female, previous testing and therapy may include diagnostic and/or therapeutic laparoscopics, hysterosalpingography, and prolonged cycle monitoring with or without hormone treatment. In the case of the males, various hormone treatment regimens, varicocoele surgery and tests such as the zona-free hamster penetration assay are typical. Given these circumstances, two important features characterize the typical IVF couple. First, their age is relatively advanced, and the female is often approaching the end of her reproductive life. Second, these couples have arrived at a juncture in their treatment course in which the likelihood of pregnancy is relatively low, and consequendy, in which loss of a pregnancy, established after years of emotional stress, medical intervention and financial expense, is a disastrous event.