Obesogens, stem cells and the developmental programming of obesity.

Obesogens are chemicals that directly or indirectly lead to increased fat accumulation and obesity. Obesogens have the potential to disrupt multiple metabolic signalling pathways in the developing organism that can result in permanent changes in adult physiology. Prenatal or perinatal exposure to obesogenic endocrine disrupting chemicals has been shown to predispose an organism to store more fat from the beginning of its life. For example, excess oestrogen or cortisol exposure in the womb or during early life resulted in an increased susceptibility to obesity and metabolic syndrome later in life. This review focuses on the effects of environmental chemicals, such as the model obesogen, tributyltin (TBT), on the development of obesity. We discuss evidence linking the obesogenic effects of TBT with its ability to activate the peroxisome proliferator-activated receptor gamma and stimulate adipogenesis. We also discuss how TBT and other environmental obesogens may lead to epigenetic changes that predispose exposed individuals to subsequent weight gain and obesity. This suggests that humans, who have been exposed to obesogenic chemicals during sensitive windows of development, might be pre-programmed to store increased amounts of fat, resulting in a lifelong struggle to maintain a healthy weight and exacerbating the deleterious effects of poor diet and inadequate exercise.

Treatment of chronic hepadnavirus infection in a woodchuck animal model with an inhibitor of protein folding and trafficking.

A novel strategy for anti-viral intervention of hepatitis B virus (HBV) through the disruption of the proper folding and transport of the hepadnavirus glycoproteins is described. Laboratory reared woodchucks chronically infected with woodchuck hepatitis virus (WHV) were treated with N-nonyl-deoxynojirimycin (N-nonyl-DNJ), an inhibitor of the endoplasmic reticulum (ER) alpha-glucosidases. The woodchucks experienced significant dose dependent decreases in enveloped WHV, resulting in undetectable amounts in some cases. The reduction in viremia correlated with the levels of hyperglucosylated glycan in the serum of treated animals. This correlation supports the mechanism of action associated with the drug and highlights the extreme sensitivity of the virus to this type of glycan inhibitor. At N-nonyl-DNJ concentrations that prevented WHV secretion, the glycosylation of most serum glycoproteins appeared unaffected, suggesting great selectivity for this class of therapeutics. Indeed, this may account for the low toxicity of the compound over the treatment period. We provide the first evidence that glucosidase inhibitors can be used in vivo to alter specific steps in the N-linked glycosylation pathway and that this inhibition has anti-viral effects.

The nature of australia antigen and its relation to antigen-antibody complex formation.

There is considerable data to support the hypothesis that Australia antigen is an infectious agent that causes hepatitis in man. (a) Association with acute viral hepatitis. (b) Association with chronic hepatitis. (c) Virus-like appearance under the electron microscope (200-A particles). (d) Transmission of Au(1) from man to man. (e) Transmission and passage of partially purified Au(1) to an animal host (infant African green monkey). (f) Localization [with fluorescent anti-Au(1)] of Au(1) in the nuclei of liver cells of patients with hepatitis and/or Au(1) in their blood. (g) Distribution of Au(1) in institutions, disease groups, and populations is consistent with the distribution of an infectious agent. (h) RNA identified in Au(1) particles isolated from blood. (i) Apparent replication of Australia antigen in tissue cultures of human liver cells. There is also considerable evidence that Australia antigen has many of the characteristics of a serum protein polymorphism. Since neither of these hypotheses has been rejected they can be combined to make a third hypothesis, namely, that Australia antigen is an infectious agent which causes hepatitis in some people infected with it and that it has the characteristics of an (inhertied) serum protein polymorphism. We propose calling agents of this postulated class "Icrons."

Australia antigen (a hepatitis-associated antigen): purification and physical properties.

Australia antigen [Au(1)], a particle associated with viral hepatitis, was isolated from the plasma of a patient with chronic anicteric hepatitis and leukemia who had received radioactive phosphorus. We have found that the immunoreactivity and appearance of Au(1) in the electron microscope were not altered by treatment with enzymes including trypsin, pronase, lipase, phospholipase C, ribonuclease, deoxyribonuclease, amylase, and neuraminidase. In contrast, other serum constituents were degraded by these enzymes. Therefore, treatment of the patient's plasma with many enzymes was exploited as an initial step for the isolation of Au(1). Subsequently, Au(1) was purified from the enzyme-treated (32)P-labeled plasma by gel filtration through Sephadex G-200 and centrifugation through sucrose and in cesium chloride gradients. There were no detectable human serum components in the purest fractions, as tested by immunoelectrophoresis and immunodiffusion. The density of the purified Au(1) was 1.21 in CsCl. The particle measured about 200 A in diameter, was predominantly spherical in shape and appeared to be composed of subunits. Nucleic acids were not detected by spectrophotometric, radiochemical, and chemical analyses. Immunoreactivity of purified Au(1) was destroyed by heating for 1 hr at 85 degrees C but was stable at 56 degrees C. Treatment with Carnoy's solution (3 parts ethanol:1 part glacial acetic acid) followed by pronase disrupted the particles as seen with the electron microscope. These findings, combined with other published information on Australia antigen and viral hepatitis, suggest that the bulk of Australia antigen in the blood of this patient is an incomplete virus or virus capsid.

The localization of Australia antigen by immunofluorescence.

We have studied the localization of Australia antigen, a particulate substance associated with hepatitis, by means of the fluorescent antibody technique. Preparations were made from 61 liver biopsy specimens taken from patients with infectious hepatitis, serum hepatitis, and a variety of other diseases. When tested with fluorescein-conjugated rabbit anti-Au(1) antisera all 26 patients who had Au(1) in their serum had specific fluorescence in their liver cells. The fluorescence appeared in three forms: as discrete particles within the nucleus, diffuse fluorescence of the entire nucleus, and fluorescence of the nuclear rim. Occasionally there were also fluorescent particles in the cytoplasm. Other specimens were tested with the fluorescent antibody including a variety of human tissues, buffy coat smears, peripheral lymphocyte cultures, and cells obtained from bile and duodenal drainage. Among these specimens, fluorescence was found in the cytoplasm of a few cells in the bone marrow of two patients with hepatitis and Au(1) in their serum, and in the liver, spleen, mesentery, and testis of one patient with leukemia, chronic hepatitis, and Au(1) in his serum. We have shown that the presence of fluorescent particles in the liver cells is strongly associated with the presence of Au(1) in the serum and the diagnosis of viral hepatitis. We believe that this study adds support to the hypothesis that Australia antigen is an antigenic determinant of a virus capable of causing hepatitis.

Biosynthesis and supramolecular assembly of procollagen IV in neonatal lung.

The rate of biosynthesis of procollagen IV, the principal collagen of basement membranes, and the concentration of specific RNAs coding for procollagen IV were measured in neonatal rat lungs. Both decreased sharply at birth and then recovered again a few days later. The supramolecular assembly of procollagen IV was followed in neonatal rat, mouse, and chick lungs, which actively elaborate endothelial and alveolar basement membranes, and in chick embryo gizzard which is rich in smooth muscle. The tetramer of four procollagen IV molecules linked covalently through their amino ends was isolated as an assembly intermediate from all these tissues. While noncovalent association of the carboxyl ends of two procollagen IV molecules occurred readily, the subsequent establishment of covalent cross-links was substantially slower in the junctional complexes of the carboxyl ends than of the amino ends. Both disulfide bonds and other, unidentified covalent links formed. The six component carboxyl peptides of a junctional complex became progressively covalently linked into two kinds of carboxyl peptide pairs. We conclude that both amino-linked tetramers and carboxyl-linked dimers of procollagen IV molecules are intermediates in the biological assembly of the collagen networks of these basement membranes.

Albumin Naskapi: a new variant of serum albumin.

An apparently new Variant of human serum albumin, albumin Naskapi, has been found in high frequency in the Naskapi Indians of Quebec and, in lower frequency,in other North American Indians. The family and population data of the albumin are consistent with its inheritance as a simple autosomal trait Controlled by a gene designated Al Naskapi. This gene is allelic with the gene Al(A) which controls the common albumin.Both homozygotes and heterozygotes have been distinguished. This is the first report of a homozygote for an albumin Variant.

Autosomal linkage between the albumin and Gc loci in humans.

Naskapi and Montagnais families segregating for albumin Naskapi give evidence for close linkage of the Gc and albumin loci with a high probability. One possible case of crossover is included in the data.

Serum ferritin as a predictor of host response to hepatitis B virus infection.

With hemodialysis patients, a high serum ferritin before there was serological evidence of hepatitis B virus infection increased the likelihood that the infection would be persistent. This finding suggested that hepatitis B virus is likely to infect and actively replicate in liver cells with the propensity for increased ferritin synthesis. The virus itself could stimulate the synthesis of ferritin in a cyclic positive feedback mechanism that increases intracellular ferritin concentration and, eventually, intracellular iron. Transformed liver cells have low iron content, do not replicate hepatitis B virus, and require iron for growth. Infected, nonmalignant liver cells could supply iron to the transformed cells and nourish their expansion.

Organizer-specific homeobox genes in Xenopus laevis embryos.

The dorsal blastopore lip of the early Xenopus laevis gastrula can organize a complete secondary body axis when transplanted to another embryo. A search for potential gene regulatory components specifically expressed in the organizer was undertaken that resulted in the identification of four types of complementary DNAs from homeobox-containing genes that fulfill this criterion. The most abundant of these encodes a DNA-binding specificity similar to that of the Drosophila melanogaster anterior morphogen bicoid. The other three are also homologous to developmentally significant Drosophila genes. These four genes may participate in the regulation of the developmental potential of the organizer.

Australia antigen: detection and transmission in shellfish.

Australia antigen was found in clams contaminated by drainage of untreated sewage from a coastal hospital. In closed-system aquariums, the antigen was ingested by clams and transmitted to previously uninfected clams. In opensystem aquariums, the titer of Australia antigen decreased with time, suggesting viral concentration rather than replication.

Developmental effects: oestrogen-induced vaginal changes and organotin-induced adipogenesis.

The emerging paradigm, the foetal origin of adult disease, is a new framework for considering the effects of endocrine disrupters on human and animal health. Prenatal diethylstilbestrol (DES) exposure resulted in various reproductive tract abnormalities in women, which is called as DES syndrome. Similar abnormalities have been demonstrated in experimental animals exposed perinatally to oestrogens. Developmental oestrogen exposure induces persistent proliferation of vaginal epithelial cells in mice. The persistent changes in the vagina of mice neonatally exposed to oestrogens results from persistent phosphorylation of erbB2 and oestrogen receptor alpha, sustained expression of EGF-like growth factors and phosphorylation of JNK1, IGF-I receptor and Akt. The ubiquitous environmental contaminant, tributyltin chloride (TBT) is well known to induce the development of male sex characteristics (imposex) in gastropods. We recently found that TBT and its congeners induce the differentiation of adipocytes in vitro and increase adipose mass in vivo in vertebrates. TBT is a nanomolar affinity ligand for retinoid X receptor (RXR) in the rock shell and for both the RXRalpha and the peroxisome proliferator-activated receptor gamma (PPARgamma) in the amphibian (Xenopus laevis), mouse, and human. TBT promotes adipogenesis in the murine 3T3-L1 cell model and perturbs key regulators of adipogenesis and lipogenic pathways in vivo, primarily through activation of RXRalpha and PPARgamma. Moreover, in utero exposure to TBT leads to strikingly elevated lipid accumulation in adipose depots, liver, and testis of neonate mice and results in increased adipose mass in adults. In X. laevis, ectopic adipocytes form in and around gonadal tissues following organotin, RXRalpha or PPARgamma ligand exposure. TBT represents the first example of an environmental endocrine disrupter that promotes adverse effects from gastropods to mammals. Prenatal (TBT) and early postnatal exposures (oestrogens) stand as strong examples of endocrine disrupting compounds that permanently alter developmental programming.

DNA polymerase activity as an index of lymphocyte stimulation: studies in Down's syndrome.

The ability of peripheral blood lymphocytes to respond to phytohemagglutinin (PHA) in vitro was studied in patients with Down's syndrome. The response was measured by the increase in DNA polymerase activity and the rate of incorporation of tritiated thymidine by the cultured lymphocytes. These activities were significantly lower in PHA-stimulated lymphocytes from patients with Down's syndrome compared with age- and sex-matched, mentally retarded patients without Down's syndrome from the same institution and the normal healthy volunteers. The impairment in response to PHA does not seem to be related to the presence of Australia antigen in patients with Down's syndrome or to institutionalization itself. In contrast to DNA polymerase activity and thymidine-(3)H uptake, there was no significant difference in the percentage of blast transformation in the three groups studied. The poor response of the lymphocytes from patients with Down's syndrome to a mitogenic stimulus could reflect an impairment of cellular immune functions in these patients which may be one of the factors contributing to the vulnerability of these patients to repeated or persistent infections.

Up-regulation of the alligator CYP3A77 gene by toxaphene and dexamethasone and its short term effect on plasma testosterone concentrations.

In this study we describe an alligator hepatic CYP3A gene, CYP3A77, which is inducible by dexamethasone and toxaphene. CYP3A plays a broad role in biotransforming both exogenous compounds and endogenous hormones such as testosterone and estradiol. Alligators collected from sites in Florida that are contaminated with organochlorine compounds exhibit differences in sex steroid concentrations. Many organochlorine compounds induce CYP3A expression in other vertebrates; hence, CYP3A induction by organochlorine contaminants could increase biotransformation and clearance of sex steroids by CYP3A and provide a plausible mechanism for the lowering of endogenous sex steroid concentrations in alligator plasma. We used real time PCR to examine whether known and suspected CYP3A inducers (dexamethasone, metyrapone, rifampicin, and toxaphene) up-regulate steady state levels of hepatic CYP3A77 transcript to determine if induction patterns in female juvenile alligators are similar to those reported in other vertebrates and whether toxaphene, an organochlorine compound found in high concentrations in Lake Apopka alligators, induces this gene. Estrogen receptor alpha (ERalpha), estrogen receptor beta (ERbeta), androgen receptor (AR), glucocorticoid receptor (GR), progesterone receptor (PR), and steroid-xenobiotic receptor (SXR) transcripts were also measured to determine whether any of these nuclear receptors are also regulated by these compounds in alligators. Dexamethasone (4.2-fold) and toxaphene (3.5-fold) significantly induced CYP3A77 gene transcript, whereas rifampicin (2.8-fold) and metyrapone (2.1-fold) up-regulated ERbeta after 24h. None of the compounds significantly up-regulated AR, ERalpha, GR, PR, or SXR over this time period. Plasma testosterone (T) did not change significantly after 24h in alligators from any of the treatment groups. Dexamethasone treated animals exhibited a strong relationship between the 24h plasma T concentrations and CYP3A77 (R(2)=0.9, positive) and SXR (R(2)=0.77, negative) transcripts, which suggests that the expression of these genes is related to plasma T in alligators. In light of our findings, we hypothesized that higher steady state CYP3A77 (and possibly SXR) gene expression would be observed in alligators collected from Lake Apopka, a polluted lake containing organochlorine compounds known to induce CYP3A isoforms in other taxa. Therefore, we measured basal levels of CYP3A77 and SXR gene transcripts in wild juvenile alligators collected from Orange Lake (reference lake), Lake Woodruff (reference lake), and Lake Apopka (contaminated lake). We found that no differences existed in CYP3A77 or SXR gene expression among animals from the lakes sampled suggesting that exposure to organochlorine compounds at concentrations present in Lake Apopka does not lead to variation in the expression of these genes, although capture stress could be interfering with these results since the glucocorticoid dexamethasone induces CYP3A77 transcript in alligators.

Iron-binding proteins and risk of cancer in Taiwan.

The relationship of serum ferritin and transferrin levels to risk of cancer was examined in a population of 21,513 Chinese male government workers in Taiwan who have been followed prospectively since 1975. On the basis of a previous study in the Solomon Islands, increased ferritin and decreased transferrin levels were predicted for those men who developed cancer. The results were consistent with the prediction. The mean serum ferritin was higher at the start of the study in 192 men who had died of cancer or who had developed primary hepatocellular carcinoma (PHC) as of July 1983, as compared to their controls. The mean serum transferrin level was lower in men who had died of cancers other than PHC. The estimate of relative risk of cancer death for a man with 200 ng ferritin/ml and 200 mg transferrin/dl, as compared to a man with levels of 20 ng/ml and 400 mg/dl, respectively, is 2.9. These serum iron-binding protein levels are at the extremes of the "normal" range. Men who subsequently died of cancer had lower hemoglobin, lower hematocrit, lower albumin, and higher globulin levels at the start of the study than did the controls. These results are consistent with the hypothesis that increased iron stores increase the risk of cancer. However, direct assessment of iron stores prior to disease was not possible, and the same constellation of findings may be consistent with other explanations.

Forecasting the development of primary hepatocellular carcinoma by the use of risk factors: studies in West Africa.

An association between hepatitis B virus (HBV) and primary hepatocellular carcinoma (PHC) has been found in several studies in Africa, Asia, and elsewhere. In this paper we considered the interrelations between several events related to HBV infection, which include the presence of: 1) hepatitis B surface antigen (HBsAg), 2) antibody to hepatitis B core antigen (anti-HBc), 3) antibody to the surface antigen (anti-HBs), 4) chronic liver disease, 5) elevated alpha-fetoprotein, and 6) PHC. With the use of preliminary epidemiologic data, risk factors related to these events were calculated. We suggested that the interactions between these events and HBV infection in parents be used to estimate the risk of PHC for an individual in this environment.

Studies of parents of children with acute leukemia.

In a case-control study, 70 mothers and 24 fathers of children with acute leukemia (AL) were compared with 70 mothers and 24 fathers of normal children. Three significant differences (p smaller than 0.05) were found when 35 factors were compared among the mother pairs and one difference among the father pairs. Mothers of children with AL, though alike in most respects to their matched controls, had a significantly lower number of monocytes than their controls. This was a new observation. The mothers of the children with AL also had higher levels of gamma-globulin, IgA, and IgG (Philadelphia only), which confirmed previous observations. The fathers and mothers had higher levels of basophils. These findings direct attention to the immune systems, particularly the mononuclear cells, of the parents of children with AL, as a focus for further studies on the etiology and pathogenesis of childhood leukemia.

Iron nutrition and tumor growth: decreased tumor growth in iron-deficient mice.

Groups of 15 mice of three different laboratory strains (BALB/c, C3H/He, DBA/2) were fed on a low iron diet (5 mg iron/kg diet), and three similar groups of 15 mice were maintained on a normal iron diet (312 mg iron/kg diet). When the low iron diet group became iron deficient, tumor cells (5 x 10(5) cells/mouse) of CA07-A (colon adenocarcinoma), HE129 (hepatoma), and M119 (mammary adenocarcinoma) were inoculated s.c. in BALB/c, C3H/He, and DBA/2 mice, respectively. All mice developed tumors, tumors grew more slowly, and the mean tumor sizes were smaller in the low iron diet group at nearly all weekly observations in all three strains of mice. No apparent differences in the behavior, activity (e.g., movement, climbing, running, grooming, etc.), and appearance were observed between low iron diet and normal iron diet mice. The mean body weight of mice at transplantation was less in the low iron than in the normal iron groups for the BALB/c strain but higher in the low iron groups of C3H/He and DBA/2 mice, indicating that food intake of mice on a low iron diet was not impaired. These results suggest that iron nutrition of the host affects tumor growth; tumor cells grow better in an iron-rich environment. This knowledge should be considered when designing treatment for patients with cancer. Iron oversupply in cancer patients might enhance tumor growth and adversely affect cancer therapy.