Interest in the biology of white adipose tissue has increased dramatically since the discovery of leptin in 1994. The identification of the product of the gene obese (ob) threw light on the role of adipose tissue in the physiopathology of obesity-related diseases and spurred the identification of numerous other adipokines, many of a proinflammatory nature. It has become increasingly evident that white adipose tissue-derived cytokines mediate between obesity-related exogenous factors (nutrition and lifestyle) and the molecular events that lead to metabolic syndrome, inflammation, and cardiovascular diseases. Here we review recent adipokine research, with particular attention to the roles of adiponectin, leptin, resistin, visfatin, apelin, omentin, and chemerin in such conditions.

To investigate the effect of the ob gene in the heterozygous condition, plasma glucose and insulin responses of adult heterozygous lean (ob/+) mice were compared with mice of the homozygous lean (+/+) and homozygous obese (ob/ob) genotypes. The ob/+ mice consumed 24% more food than +/+ mice although body weights were similar. Plasma glucose and insulin concentrations were respectively 16% and 176% higher in ob/+ mice than +/+ mice in the freely fed state, and 44% and 88% higher during glucose tolerance tests. In 24 hour fasted ob/+ mice, plasma glucose concentrations were 23% higher than +/+ mice but plasma insulin concentrations were not significantly different. Arginine produced a greater insulin response (172%) and a greater fall in glycaemia (200%) in ob/+ mice. A significant difference in the hypoglycaemic effect of insulin in ob/+ and +/+ mice was not observed. These results demonstrate an effect of the ob gene on glucose homeostasis in heterozygous lean (ob/+) mice. The abnormalities were qualitatively similar but considerably less severe than those in ob/ob mice, suggesting that ob/+ mice might prove useful to study factors predisposing to inappropriate hyperglycaemia.

Macrophage products, such as cytokines, prostanoids, nitric oxide, and reactive oxygen intermediates, influence the function and viability of macrophages and neighboring cells. Given that the liver has one of the largest resident macrophage populations in the body, it is not surprising that hepatic macrophages [i.e., Kupffer cells (KC)] are involved in the pathogenesis of many kinds of liver disease. This review summarizes the abnormalities that have been demonstrated in bone marrow, peritoneal and hepatic macrophage of leptin-resistant (fa/fa) rats and leptin-deficient (ob/ob) mice, two animal models for nonalcoholic fatty liver disease (NAFLD). Evidence supports the concept that altered KC function influences the viability of other cells, such as lymphocytes and hepatocytes, in fatty livers, thereby contributing to the pathogenesis of NAFLD in animals with reduced leptin activity. Further work is needed to determine whether KC dysfunction is a component of more generalized mechanisms that lead to NAFLD.

BACKGROUND

The T-20 Versus Optimized Background Regimen Only (TORO) 1 and TORO 2 clinical trials are open-label, controlled, parallel-group, phase 3 studies comparing enfuvirtide plus an optimized background (OB) of antiretrovirals (n = 661) with OB alone (n = 334) in treatment-experienced HIV-1-infected patients.

METHODS

The primary objective at week 48 was to investigate durability of efficacy, as measured by the percentage of patients maintaining their week 24 response or improving. Efficacy analyses used the intent-to-treat population.

RESULTS

A total of 73.7% of patients randomized to the enfuvirtide group remained on treatment through week 48 versus 21.3% originally randomized to the control group. At week 48, a higher proportion of week 24 responders maintained their response or were new responders in the enfuvirtide group than in the control group in each responder category: HIV-1 RNA level>> or =1.0 log(10) change from baseline, <400 copies/mL and <50 copies/mL (37.4%, 30.4%, and 18.3% in the enfuvirtide group vs. 17.1%, 12.0%, and 7.8% in the control group, respectively; P < 0.0001 for all comparisons). CD4 cell count increases from baseline were twice as great in the enfuvirtide group as in the control group.

CONCLUSIONS

These data demonstrate durable efficacy of enfuvirtide plus OB over 48 weeks.

Neural stem cells depend on insulin-like growth factor I (IGF-I) for differentiation. We analysed how activation and inhibition of the PI 3-kinase-Akt signalling affects the number and differentiation of mouse olfactory bulb stem cells (OBSCs). Stimulation of the pathway with insulin and/or IGF-I, led to an increase in Akt phosphorylated on residues Ser473 and Thr308 (P-Akt(Ser473) and P-Akt(Thr308), respectively) in proliferating OBSCs, and in differentiating cells. Conversely, P-Akt(Ser473) levels decreased by 50% in the OB of embryonic day 16.5-18.5 IGF-I knockout mouse embryos. Overexpression of PTEN, a negative regulator of the PI 3-kinase pathway, caused a reduction in the basal levels of P-Akt(Ser473) and P-Akt(Thr308) and a minor reduction in IGF-I-stimulated P-Akt(Ser473). Although PTEN overexpression decreased the proportion of neurons and astrocytes in the absence of insulin/IGF-I, it did not alter the proliferation or survival of OBSCs. Accordingly, overexpression of a catalytically inactive PTEN mutant promoted OBSCs differentiation. Inhibition of PI 3-kinase by LY294002 produced strong and moderate reductions in IGF-I-stimulated P-Akt(Ser473) and P-Akt(Thr308), respectively. Consequently, LY294002 reduced the proliferation of OBSCs and the number of neurons and astrocytes, and also augmented cell death. These findings indicate that OBSC differentiation is more sensitive to lower basal levels of P-Akt than proliferation or death. By regulating P-Akt levels in opposite ways, IGF-I and PTEN contribute to the fine control of neurogenesis in the olfactory bulb.

Chlorella virus DNA ligase, the smallest eukaryotic ligase known, has pluripotent biological activity and an intrinsic nick-sensing function, despite having none of the accessory domains found in cellular ligases. A 2.3-A crystal structure of the Chlorella virus ligase-AMP intermediate bound to duplex DNA containing a 3'-OH-5'-PO4 nick reveals a new mode of DNA envelopment, in which a short surface loop emanating from the OB domain forms a beta-hairpin 'latch' that inserts into the DNA major groove flanking the nick. A network of interactions with the 3'-OH and 5'-PO4 termini in the active site illuminates the DNA adenylylation mechanism and the crucial roles of AMP in nick sensing and catalysis. Addition of a divalent cation triggered nick sealing in crystallo, establishing that the nick complex is a bona fide intermediate in the DNA repair pathway.

Mechanisms regulating circulating leptin are incompletely understood. We developed a radioimmunoassay for mouse leptin to examine the influence of age, dietary fat content, and fasting on plasma concentrations of leptin in the background strain for the ob/ob mouse, the C57BL/6J mouse. Plasma leptin increased with age [5.3 +/- 0.6 ng/ml at 2 mo (n = 23) vs. 14.2 +/- 1.6 ng/ml at 11 mo (n = 15), P < 0.001]. Across all age groups (2-11 mo, n = 160), log plasma leptin correlated with body weight (r = 0.68, P < 0.0001), plasma insulin (r = 0.38, P < 0.001), and amount of intra-abdominal fat (r = 0.90, P < 0.001), as revealed by magnetic resonance imaging. Plasma leptin was increased by a high-fat diet (58% fat for 10 mo) and reduced by fasting for 48 h. The reduction of plasma leptin was correlated with the reduction of plasma insulin (r = 0.43, P = 0.012) but not with the initial body weight or the change in body weight. Moreover, the reduction in plasma leptin by fasting was impaired by high-fat diet. Thus plasma leptin in C57BL/6J mice 1) increases with age or a high-fat diet; 2) correlates with body weight, fat content, and plasma insulin; and 3) is reduced during fasting by an action inhibited by high-fat diet and related to changes of plasma insulin.

Insulin resistance in type 2 diabetes is characterized by defects in muscle glucose uptake and hepatic overproduction of both glucose and lipids. These hepatic defects are perplexing because insulin normally suppresses glucose production and increases lipid synthesis in the liver. To understand the mechanisms for these seemingly paradoxical defects, we examined the activation of atypical protein kinase C (aPKC) and protein kinase B (PKB), two key signaling factors that operate downstream of phosphatidylinositol 3-kinase and regulate various insulin-sensitive metabolic processes. Livers and muscles of three insulin-resistant rodent models were studied. In livers of type 2 diabetic non-obese Goto-Kakazaki rats and ob/ob-diabetic mice, the activation of PKB was impaired, whereas activation of aPKC was surprisingly maintained. In livers of non-diabetic high fatfed mice, the activation of both aPKC and PKB was maintained. In contrast to the maintenance of aPKC activation in the liver, insulin activation of aPKC was impaired in muscles of Goto-Kakazaki-diabetic rats and ob/ob-diabetic and non-diabetic high fat-fed mice. These findings suggest that, at least in these rodent models, (a) defects in aPKC activation contribute importantly to skeletal muscle insulin resistance observed in both high fat feeding and type 2 diabetes; (b) insulin signaling defects in muscle are not necessarily accompanied by similar defects in liver; (c) defects in hepatic PKB activation occur in association with, and probably contribute importantly to, the development of overt diabetes; and (d) maintenance of hepatic aPKC activation may explain the continued effectiveness of insulin for stimulating certain metabolic actions in the liver.

Insulin resistance of diaphragms of ob/ob mice has been repeatedly demonstrated previously both in vitro and in vivo. In the present study, transport and metabolism of glucose with and without insulin stimulation were compared in a skeletal muscle more likely than diaphragm or heart to be representative of the overall striated muscle mass, i.e. isolated soleus muscle. Compared with soleus muscle from lean controls, unstimulated lactate release in the presence of exogenous glucose was depressed from 16.2 to 12.3 nmol/60 min per mg wet wt in soleus from ob/ob mutants; glycolysis was decreased from 6.6 to 3.7 and [14C]glucose oxidation to 14CO2 from 0.90 to 0.33 nmol glucose/60 min per mg wet wt. Uptake of 2-deoxyglucose (2-DOG), both with and without insulin, was very much less for soleus from ob/ob than from lean mice, at 2-DOG concentrations ranging from 0.1 to 10 mM, and in mice of 6-15 wk. When 2-DOG concentration was 1 mM, its basal uptake was 0.53 nmol/30 min per mg wet wt for soleus of ob/ob as against 0.96 for soleus of lean mice. The absolute increment due to 1 mU/ml insulin was 0.49 in muscle of ob/ob as against 1.21 in that of lean mice. When the resistance to insulin action was decreased by pretreatment in vivo by either streptozotocin injection or fasting, the decreased basal 2-DOG uptake of subsequently isolated soleus muscle was not improved. Inhibition of endogenous oxidation of fatty acids by 2-bromostearate, while greatly increasing 14CO2 production from [14C]glucose, did not affect basal [5-3H]glucose metabolism or 2-DOG uptake. It is suggested that transport and/or phosphorylation of glucose under basal, unstimulated conditions are depressed in soleus muscle of ob/ob mice, whether or not resistance to insulin and hyperinsulinemia are also present. Although the origin of the decreased basal glucose uptake remains unknown it might be related to a similar decrease in basal glucose uptake by ventromedial hypothalamic cells, an event presumably resulting in a tendency to hyperphagia. Decreased basal glucose uptake by soleus muscle of ob/ob mice might explain the hyperglycemia, and hence partly the hyperinsulinemia and excessive fat deposition of those animals.

OBJECTIVE

Whole body and myocardial insulin resistance are features of non-insulin-dependent diabetes mellitus (NIDDM) and left-ventricular dysfunction (LVD). We determined whether abnormalities of insulin receptor substrate-1 (IRS1), IRS1-associated PI3K (IRS1-PI3K), and glucose transporter 4 (GLUT4) contribute to tissue-specific insulin resistance.

RESULTS

We collected skeletal muscle (n = 27) and myocardial biopsies (n = 24) from control patients (n = 7), patients with NIDDM (n = 9) and patients with LVD (n = 8), who were characterized by euglycaemic-hyperinsulinaemic clamp and positron emission tomography. Comparative studies were carried out in three mouse models. We demonstrate an unrecognized reduction of IRS1 in skeletal muscle of LVD patients and an unexpected increase in cardiac IRS1-PI3K activity in NIDDM and LVD patients. In NIDDM, there was a concomitant reduction in sarcolemmal GLUT4, whereas in patients with LVD sarcolemmal GLUT4 was increased. We confirm activation of IRS1-PI3K and reduction in sarcolemmal GLUT4 in the insulin resistant ob/ob mouse heart where we also demonstrate perturbation of GLUT4 docking and fusion. A direct relationship between PI3K and GLUT4 was demonstrated in mice expressing activated PI3K in the heart and increased GLUT4 at the sarcolemma was confirmed in a mouse model of LVD.

CONCLUSIONS

Our data show that the mechanisms of myocardial insulin resistance are different between NIDDM and LVD.

BACKGROUND

Peptide YY (PYY) is a 36 amino-acid peptide secreted from ileal L cells following meals. The cleaved subpeptide PYY[3-36] is biologically active and may constitute the majority of circulating PYY-like immunoreactivity. The peptide family that includes PYY, pancreatic peptide and neuropeptide Y is noted for its orexigenic effect following intracerebroventricular administration.

OBJECTIVE

To investigate the effects of peripheral (intraperitoneal and chronic subcutaneous) infusions of PYY[3-36] on food intake, body weight and glycemic indices.

UNASSIGNED

Food intake was measured in normal mice and in several rodent models of obesity and type II diabetes. In marked contrast to the reported central orexigenic effects, in the present study, PYY[3-36] acutely inhibited food intake by up to 45%, with an ED(50) of 12.5 microg/kg in fasted female NIH/Swiss mice. A 4-week infusion reduced weight gain in female ob/ob mice, without affecting the cumulative food intake. In diet-induced obese male mice, PYY[3-36] infusion reduced cumulative food intake, weight gain and epididymal fat weight (as a fraction of carcass) with similar ED(50)'s (466, 297 and 201 microg/kg/day, respectively) and prevented a diet-induced increase in HbA1c. Infusion at 100 microg/kg/day for 8 weeks in male fa/fa rats reduced the weight gain (288+/-11 vs 326+/-12 g in saline-infused controls; P<0.05), similar to effects in a pair-fed group. In female ob/ob and db/db mice, there was no acute effect of PYY[3-36] on plasma glucose concentrations. In male diabetic fatty Zucker rats, PYY[3-36] infused for 4 weeks reduced HbA1c and fructosamine (ED(50)'s 30 and 44 microg/kg/day).

CONCLUSIONS

Peripheral PYY[3-36] administration reduced the food intake, body weight gain and glycemic indices in diverse rodent models of metabolic disease of both sexes. These findings justify further exploration of the potential physiologic and therapeutic roles of PYY[3-36].

Besides being mandatory in the metabolic system, adipokines like leptin directly affect immunity. Leptin was found to be necessary in T helper 1 (Th1)-dependent inflammatory processes, whereas effects on Th2 cells are rarely understood. Here, we focused on leptin in T-helper cell polarization and in Th2-mediated intestinal inflammation in vivo. The induction of cytokine-producing Th1 or Th2 cells from naive CD4(+) T cells under polarizing conditions in vitro was generally decreased in cells from leptin-deficient ob/ob mice compared with wild-type mice. To explore the in vivo relevance of leptin in Th2-mediated inflammation, the model of oxazolone-induced colitis was employed in wild-type, ob/ob, and leptin-reconstituted ob/ob mice. Ob/ob mice were protected, whereas wild-type and leptin-reconstituted ob/ob mice developed colitis. The disease severity went in parallel with local production of the Th2 cytokine IL-13. A possible explanation for the protection of ob/ob mice in Th1- as well as in Th2-dependent inflammation is provided by a decreased expression of the key transcription factors for Th1 and Th2 polarization, T-bet and GATA-3, in naive ob/ob T cells. In conclusion, these results support the regulatory function of the adipokine leptin within T-cell polarization and thus in the acquired immune system and support the concept that there is a close interaction with the endocrine system.

OBJECTIVE

The purpose of this study was to clarify the effects of maternal obesity on insulin sensitivity and secretion in offspring.

METHODS

Fifty-one offspring of both sexes of obese (Ob group) and 15 offspring of normal-weight (control group) mothers were studied. Plasma glucose, insulin, and C-peptide were measured during an oral glucose tolerance test (OGTT). Insulin sensitivity was calculated using the oral glucose insulin sensitivity index, and insulin secretion and beta-cell glucose sensitivity were computed by a mathematical model. Fasting leptin and adiponectin were also measured. Body composition was assessed by dual-X-ray absorptiometry.

RESULTS

No birth weight statistical difference was observed in the two groups. Of the Ob group, 69% were obese and 19% were overweight. The Ob group were more insulin resistant than the control group (398.58 +/- 79.32 vs. 513.81 +/- 70.70 ml(-1) x min(-1) x m(-2) in women, P < 0.0001; 416.42 +/- 76.17 vs. 484.242 +/- 45.76 ml(-1) x min(-1) x m(-2) in men, P < 0.05). Insulin secretion after OGTT was higher in Ob group than in control group men (63.94 +/- 21.20 vs. 35.71 +/- 10.02 nmol x m(-2), P < 0.01) but did not differ significantly in women. beta-Cell glucose sensitivity was not statistically different between groups. A multivariate analysis of variance showed that maternal obesity and offspring sex concurred together with BMI and beta-cell glucose sensitivity to determine the differences in insulin sensitivity and secretion observed in offspring.

CONCLUSIONS

Obese mothers can give birth to normal birth weight babies who later develop obesity and insulin resistance. The maternal genetic/epigenetic transmission shows a clear sexual dimorphism, with male offspring having a higher value of insulin sensitivity (although not statistically significant) associated with significantly higher insulin secretion than female offspring.

Maternal obesity is increasing at an alarming rate. We previously showed that maternal obesity induces an inflammatory response and enhances adipogenesis in fetal skeletal muscle at midgestation. The objective of this study was to evaluate effects of maternal obesity on adipogenesis, inflammatory signaling, and insulin pathways at late gestation when ovine fetal skeletal muscle matures. Nonpregnant ewes were assigned to a control diet (Con, fed 100% of National Research Council nutrient recommendations, n = 6) or obesogenic diet (OB, fed 150% of National Research Council recommendations, n = 6) from 60 d before to 135 d after conception (term 148 d) when the fetal semitendenosus skeletal muscle was sampled. Expression of the adipogenic marker, peroxisome proliferator-activated receptor-gamma, was increased in OB compared with Con fetal semitendenosus muscle, indicating up-regulation of adipogenesis. More intramuscular adipocytes were observed in OB muscle. Phosphorylation of inhibitor-kappaB kinase-alpha/beta and nuclear factor-kappaB RelA/p65 were both increased in OB fetal muscle, indicating activation of nuclear factor-kappaB pathway. Phosphorylation of c-Jun N-terminal kinase and c-Jun (at Ser 63 and Ser 73) was also elevated. Toll-like receptor 4 expression was higher in OB than Con fetal muscle. Moreover, despite higher insulin concentrations in OB vs. Con fetal plasma (2.89 +/- 0.53 vs. 1.06 +/- 0.52 ng/ml; P < 0.05), phosphorylation of protein kinase B at Ser 473 was reduced, indicating insulin resistance. In conclusion, our data show maternal obesity-induced inflammatory signaling in late gestation fetal muscle, which correlates with increased im adipogenesis and insulin resistance, which may predispose offspring to later-life obesity and diabetes.

BACKGROUND

Appetite, food intake and weight are frequently altered in psychiatric disorders such as major depression and schizophrenia. Leptin is an adipocyte hormone, as the product of the ob gene, regulating food intake and energy balance providing the hypothalamus with information on the amount of body fat. Leptin seems to be strongly associated with lipid metabolism. Moreover, leptin is involved in the control of other behaviors and in brain development. There are few studies about the amounts of plasma leptin in mood disorder and schizophrenia with inconsistent findings. The relationship between leptin and major depressive disorder is still unknown. We planned to investigate the relationship of the serum leptin concentration, cholesterol, and BMI between patients with major depressive disorder, schizophrenic patients and healthy control subjects.

METHODS

In the present study, the BMI, plasma cholesterol and leptin levels, BDI, and BPRS were compared in 69 patients with major depressive disorder, 78 schizophrenic patients, and 51 healthy controls.

RESULTS

The major findings of our study included (1) leptin and cholesterol levels were low in patients with major depressive disorder, but high in schizophrenic patients; (2) negative correlations between BDI scores and serum cholesterol or leptin levels in the patients with major depressive disorder; (3) an inconsistently positive correlation between mean leptin levels, cholesterol, and BMI among different groups; (4) positive correlations between serum cholesterol or leptin levels and the length of illness in the schizophrenic patients.

CONCLUSIONS

In this study, our results indicate that that leptin and cholesterol might play differently important pathophysiological roles in these psychiatric disorders.

The unique vulnerability of the olfactory system to Alzheimer's disease (AD) provides a quintessential translational tool for understanding mechanisms of synaptic dysfunction and pathological progression in the disease. Using the Tg2576 mouse model of β-amyloidosis, we show that aberrant, hyperactive olfactory network activity begins early in life, before detectable behavioral impairments or comparable hippocampal dysfunction and at a time when amyloid-β (Aβ) deposition is restricted to the olfactory bulb (OB). Hyperactive odor-evoked activity in the piriform cortex (PCX) and increased OB-PCX functional connectivity emerged at a time coinciding with olfactory behavior impairments. This hyperactive activity persisted until later in life when the network converted to a hyporesponsive state. This conversion was Aβ-dependent, because liver-X receptor agonist treatment to promote Aβ degradation rescued the hyporesponsive state and olfactory behavior. These data lend evidence to a novel working model of olfactory dysfunction in AD and, complimentary to other recent works, suggest that disease-relevant network dysfunction is highly dynamic and region specific, yet with lasting effects on cognition and behavior.

OBJECTIVE

To determine the capacity of human subchondral osteoarthritic osteoblasts (Ob) to produce interleukin (IL)-1beta, IL-6, transforming growth factor-beta (TGF-beta) and prostaglandin E(2) (PGE(2)), and determine if a relationship exists between IL-1beta, TGF-beta, PGE(2) and IL-6 production.

METHODS

We measured the abundance of IL-1beta, IL-6, TGF-beta and PGE(2) using very sensitive ELISA in conditioned-media of human primary subchondral Ob from normal individuals and osteoarthritic patients. Selective inhibition of IL-6 or IL-6 receptor signaling was performed to determine its effect on PGE(2) production whereas the inhibiton of PGE(2) production was performed to determine its effect on IL-6 production. The expression of bone cell markers and urokinase plasminogen activator (uPA) activity was also determined.

RESULTS

Osteoarthritic Ob produced all these factors with greater variability than normal cells. Interestingly, the production of IL-6 and PGE(2) by osteoarthritic Ob separated patients into two subgroups, those whose Ob produced levels comparable to normal (low producers) and those whose Ob produced higher levels (high producers). In those cells classified as high osteoarthritic Ob, PGE(2) and IL-6 levels were increased two- to three-fold and five- to six-fold, respectively, compared with normal. In contrast, while using their IL-6 and PGE(2) production to separate osteoarthritic Ob into low and high producers, we found that IL-1beta levels were similar in normal and all osteoarthritic Ob. Using the same criteria, TGF-beta levels were increased in all osteoarthritic Ob compared with normal. Reducing PGE(2) synthesis by Indomethacin [a cyclo-oxygenase (COX) -1 and -2 inhibitor] reduced IL-6 levels in all osteoarthritic Ob, whereas Naproxen (a more selective COX-2 inhbitor) reduced PGE(2) and IL-6 levels only in the high osteoarthritic group. Conversely, PGE(2) addition to osteoarthritic Ob enhanced IL-6 production in both groups. Moreover, the addition of parathyroid hormone also stimulated IL-6 production to similar normal levels in both osteoarthritic groups. In contrast, using an antibody against IL-6 or IL-6 receptors did not reduce PGE(2) levels in either group. The evaluation of alkaline phosphatase activity, osteocalcin release, collagen type I and uPA activity in osteoarthritic Ob failed to show any differences between these cells regardless to which subgroup they were assigned.

CONCLUSIONS

These results indicate that IL-6 and PGE(2) production by subchondral Ob can discriminate two subgroups of osteoarthritic patients that cannot otherwise be separated by their expression of cell markers, and that endogenous PGE(2) levels influence IL-6 synthesis in osteoarthritic Ob.

BACKGROUND

Multiple myeloma (MM) expands almost exclusively in the bone marrow and generates devastating bone lesions, in which bone formation is impaired and osteoclastic bone resorption is enhanced. TGF-beta, a potent inhibitor of terminal osteoblast (OB) differentiation, is abundantly deposited in the bone matrix, and released and activated by the enhanced bone resorption in MM. The present study was therefore undertaken to clarify the role of TGF-beta and its inhibition in bone formation and tumor growth in MM.

RESULTS

TGF-beta suppressed OB differentiation from bone marrow stromal cells and MC3T3-E1 preosteoblastic cells, and also inhibited adipogenesis from C3H10T1/2 immature mesenchymal cells, suggesting differentiation arrest by TGF-beta. Inhibitors for a TGF-beta type I receptor kinase, SB431542 and Ki26894, potently enhanced OB differentiation from bone marrow stromal cells as well as MC3T3-E1 cells. The TGF-beta inhibition was able to restore OB differentiation suppressed by MM cell conditioned medium as well as bone marrow plasma from MM patients. Interestingly, TGF-beta inhibition expedited OB differentiation in parallel with suppression of MM cell growth. The anti-MM activity was elaborated exclusively by terminally differentiated OBs, which potentiated the cytotoxic effects of melphalan and dexamethasone on MM cells. Furthermore, TGF-beta inhibition was able to suppress MM cell growth within the bone marrow while preventing bone destruction in MM-bearing animal models.

CONCLUSIONS

The present study demonstrates that TGF-beta inhibition releases stromal cells from their differentiation arrest by MM and facilitates the formation of terminally differentiated OBs, and that terminally differentiated OBs inhibit MM cell growth and survival and enhance the susceptibility of MM cells to anti-MM agents to overcome the drug resistance mediated by stromal cells. Therefore, TGF-beta appears to be an important therapeutic target in MM bone lesions.

The genome of Ob, a tobamovirus that overcomes the N gene-mediated hypersensitive response (HR), was cloned as a cDNA, and its nucleotide sequence was determined. The genomic organization of Ob is similar to that of other tobamoviruses, consisting of 6506 nucleotides and containing at least four open reading frames. These open reading frames encode a 126-kD polypeptide with a 183-kD readthrough product, a 30.6-kD movement protein, and an 18-kD coat protein. A bacteriophage T7 promoter sequence was fused to the full-length cDNA clone to obtain infectious RNA transcripts. These transcripts, when inoculated onto tobacco plants, induced disease symptoms indistinguishable from plants inoculated with Ob viral RNA. To determine which viral factor is responsible for the resistance-breaking character of Ob, a recombinant virus was constructed in which the movement protein gene of tobacco mosaic virus was replaced with that of Ob. Cultivar Xanthi NN tobacco plants infected with this virus responded with an HR, indicating that the Ob movement protein alone does not act to overcome the N gene-mediated response. Following mutagenesis of the infectious Ob cDNA clone with hydroxylamine, populations of transcripts from the mutagenized DNA were inoculated onto Xanthi NN tobacco, and a variant that induced the HR was identified. The mutant was analyzed and found to contain a single nucleotide change in the 126-kD gene. Recreating the mutation in the Ob cDNA clone by site-directed mutagenesis resulted in a virus that caused symptoms identical to the chemically induced mutant.

Inhibition of Wnt/beta-catenin/T-cell factor (TCF) signaling induces proliferation of mesenchymal stem cells and/or suppresses their differentiation into osteoblasts (OBs). Osteolysis in multiple myeloma (MM) is related to the suppression of canonical Wnt signaling caused by DKK1, a soluble inhibitor of this pathway secreted by MM cells. Bortezomib (Bzb) can induce OB differentiation in vitro and in vivo and its anti-MM efficacy linked to bone anabolic effects. However, the molecular basis of the action of Bzb on bone is not completely understood. In the present study, we show that Bzb promotes matrix mineralization and calcium deposition by osteoprogenitor cells and primary mesenchymal stem cells via Wnt-independent activation of beta-catenin/TCF signaling. Using affinity pull-down assays with immunoblotting and immunofluorescence, we found that Bzb induced stabilization of beta-catenin. Nuclear translocation of stabilized beta-catenin was associated with beta-catenin/TCF transcriptional activity that was independent of the effects of Wnt ligand-receptor-induced signaling or GSK3beta activation. Blocking the activation of beta-catenin/TCF signaling by dominant negative TCF attenuated Bzb-induced matrix mineralization. These results provide evidence that Bzb induces OB differentiation via Wnt-independent activation of beta-catenin/TCF pathway and suggest that proteasome inhibition therapy in MM may function in part by subverting tumor-induced suppression of canonical Wnt signaling in the bone microenvironment.

Several models have been proposed to suggest how the evolution of sex-determining mechanisms might contribute to speciation. Here, we describe the inheritance of sex in 19 fish species from the rapidly evolving flock of cichlids in Lake Malawi, Africa. We found that many of these species have a male heterogametic (XY) system on linkage group 7. Some species also segregate for a female heterogametic (ZW) system on linkage group 5 that is coincident with a dominant orange-blotch (OB) color pattern in females. The ZW system is epistatically dominant to the XY system when both are segregating within a family. Several lines of evidence suggest that additional sex-determining loci are segregating in some species. These results are consistent with the idea that genetic conflicts play an important role in the evolution of these species flocks and suggest that evolution of sex-determining mechanisms has contributed to the radiation of cichlid fish in East Africa.

Semaphorin 3A (Sema3A) is a membrane-associated secreted protein that has chemorepulsive properties for neuropilin-1 (npn-1)- expressing axons. Although mice lacking the Sema3A protein display skeletal abnormalities and heart defects, most axonal projections in the CNS develop normally. We show here that Sema3A is expressed in the lamina propria surrounding the olfactory epithelium (OE) and by ensheathing cells in the nerve layer of the ventral olfactory bulb (OB) throughout development. Subsets of sensory neurons expressing npn-1 are distributed throughout the OE and extend fibers to the developing OB. In wild-type mice, npn-1-positive (npn-1(+)) axons extend to lateral targets in the rostral OB and medial targets in the caudal OB, avoiding regions expressing Sema3A. In Sema3A homozygous mutant mice, many npn-1(+) axons are misrouted into and through the ventral nerve layer, beginning as early as embryonic day 13 and continuing at least until birth. At postnatal day 0, npn-1(+) glomeruli are atypically located in the ventral OB of Sema3A(-/-) mice, indicating that aberrant axon trajectories are not corrected during development and that connections are made in inappropriate target regions. In addition, subsets of OCAM(+) axons that normally project to the ventrolateral OB and some lactosamine-containing glycan(+) axons that normally target the ventral OB are also misrouted in Sema3A mutants. These observations indicate that Sema3A expression by ensheathing cells plays an important role in guiding olfactory axons into specific compartments of the OB.

The risk of obesity (OB) in adulthood is strongly influenced by maternal body composition. Here we examined the hypothesis that maternal OB influences white adipose tissue (WAT) transcriptome and increases propensity for adipogenesis in the offspring, prior to the development of OB, using an established model of long-term metabolic programming. Employing an overfeeding-based rat model, in which exposure to OB is limited to preconception and gestation alone, we conducted global transcriptomic profiling in WAT, and gene/protein expression analysis of lipogenic and adipogenic pathways and examined adipogenic differentiation of WAT stromal-vascular cells ex vivo. Using reduced representation bisulfite sequencing we also evaluated genome-scale changes in DNA methylation in offspring WAT. Maternal OB led to extensive changes in expression of genes (± 1.8-fold, P ≤ .05), revealing a distinct up-regulation of lipogenic pathways in WAT. mRNA expression of a battery of sterol regulatory element-binding protein-1-regulated genes was increased in OB-dam offspring, which were confirmed by immunoblotting. In conjunction with lipogenic gene expression, OB-dam offspring showed increased glucose transporter-4 mRNA/protein expression and greater AKT phosphorylation following acute insulin challenge, suggesting sensitization of insulin signaling in WAT. Offspring of OB dams also exhibited increased in vivo expression of adipogenic regulators (peroxisome proliferator-activated receptor-γ, CCAAT enhancer binding protein α [C/EBP-α] and C/EBP-β), associated with greater ex vivo differentiation of WAT stromal-vascular cells. These transcriptomic changes were associated with alterations in DNA methylation of CpG sites and CGI shores, proximal to developmentally important genes, including key pro-adipogenic factors (Zfp423 and C/EBP-β). Our findings strongly suggest that the maternal OB in utero alters adipocyte commitment and differentiation via epigenetic mechanisms.

OBJECTIVE

Leptin, a hormone mainly produced by fat cells, acts primarily on the hypothalamus regulating energy expenditure and food intake. Leptin receptors are expressed in several tissues and the possible physiological role of leptin is being extensively investigated, with the result that important peripheral actions of the hormone in the organism are being discovered. Recent studies have demonstrated leptin and leptin receptor expression in gastric epithelial cells. In the present study, we report the presence of the long leptin receptor isoform (OB-Rb) in human, rat, and mouse small intestine, supporting the hypothesis of leptin as a hormone involved in gastrointestinal function.

METHODS

The presence of the leptin receptor was determined by immunocytochemical methods using antibodies against the peptide corresponding to the carboxy terminus of the long isoform of the leptin receptor. Human duodenal biopsies from normal individuals undergoing gastrointestinal endoscopy, and intestinal fragments of Wistar rats and Swiss mice were processed for the study.

RESULTS

Immunoreactivity for the long leptin receptor isoform was observed in the three studied species. Staining was located throughout the cytoplasm of the enterocytes, of both villi and crypts, and in the basolateral plasma membrane. Immunolabelling for OB-Rb protein was also found in the brush border of human enterocytes of formol and paraformaldehyde fixed samples.

CONCLUSIONS

This report demonstrates the presence of the long leptin receptor isoform in the absorptive cells of rat, mouse, and human small intestine, suggesting that leptin could have a physiological role in the regulation of nutrient absorption.