IntroductionSubcutaneous adipose tissue is an interesting source of autologous stem cells with a fundamental role in the pathophysiology of obesity, metabolic syndromes and insulin resistance. We hypothesize that obesity could alter the stromal-vascular fraction (SVF) and adipose stem cell (ASCs) functions, which could compromise its regenerative behavior. Furthermore, we aimed to evaluate whether ASCs derived from post bariatric surgery ex-obese women maintain their functions in a similar fashion as do those from individuals who have never been obese.MethodsThe SVF of subcutaneous adipose tissue from control (n = 6, body mass index – BMI - 27.5 ± 0.5 kg/m2), obese (n = 12, BMI 46.2 ± 5.1 kg/m2) and post bariatric surgery ex-obese (n = 7, initial BMI 47.8 ± 1.3 kg/m2; final BMI 28.1 ± 1.1 kg/m2) women were isolated and evaluated by flow cytometry. ASCs were tested for lipid accumulation by perilipin, adipose differentiation-related protein (ADRP) and Oil Red O staining after adipogenic stimulus. The cytokines secreted by the ASCs and after lipid accumulation induction were also evaluated.ResultsThe subcutaneous adipose tissue of obese and post bariatric surgery ex-obese women was enriched in pericytes (p = 0.0345). The number of supra-adventitial cells was not altered in the obese patients, but it was highly enriched in the post bariatric surgery ex-obese women (p = 0.0099). The ASCs of the post bariatric surgery ex-obese patients secreted more MCP-1 (monocyte chemoattractant protein-1; p = 0.0078). After lipid accumulation induction, the ASCs of the patients in all groups secreted less IL-6 than the ASCs with no adipogenic stimulus (p < 0.0001). Obese ASCs with lipid accumulation secreted the highest amount of IL-6 (p < 0.001) whereas the ASCs from the controls secreted the highest amount of adiponectin (p < 0.0001). The ASCs from the post bariatric surgery ex-obese patients showed the highest levels of lipid accumulation whereas those from the obese women had the lowest levels (p < 0.0001).ConclusionsSVF content and ASC behavior are altered in the subcutaneous adipose tissue of morbid obese women; these changes are not completely restored after bariatric surgery-induced weight loss. The cellular alterations described in this study could affect the regenerative effects of adipose stem cells. Further investigations are required to avoid jeopardizing the development of autologous stem cell-based therapies.

In mammals, lipid droplets (LDs) are ubiquitous organelles that modulate immune and inflammatory responses through the production of lipid mediators. In insects, it is unknown whether LDs play any role during the development of immune responses. We show that Aedes aegypti Aag2 cells – an immune responsive cell lineage – accumulates LDs when challenged with Enterobacter cloacae, Sindbis, and Dengue viruses. Microarray analysis of Aag2 challenged with E.cloacae or infected with Dengue virus revealed high transcripts levels of genes associated with lipid storage and LDs biogenesis, correlating with the increased LDs numbers in those conditions. Similarly, in mosquitoes, LDs accumulate in midgut cells in response to Serratia marcescens and Sindbis virus or when the native microbiota proliferates, following a blood meal. Also, constitutive activation of Toll and IMD pathways by knocking-down their respective negative modulators (Cactus and Caspar) increases LDs numbers in the midgut. Our results show for the first time an infection-induced LDs accumulation in response to both bacterial and viral infections in Ae. Aegypti, and we propose a role for LDs in mosquito immunity. These findings open new venues for further studies in insect immune responses associated with lipid metabolism.

Background: Leptin is an adipokine with well-known effects on the central nervous system including the induction of energy expenditure and satiety. Leptin also has major relevance when activating immune cells and modulating inflammatory response. In obesity, increases in white adipose tissue accumulation and leptin levels are accompanied by hypothalamic resistance to leptin. Even though the adipose tissue is a leptin-rich environment, the local actions of leptin regarding adipogenesis were not thoroughly investigated until now. Here we evaluate the contributions of leptins direct signaling in preadipocytes and adipose tissue-derived stromal cells (ASCs) for adipogenesis.Methods: Adipocytes were differentiated from the murine lineage of preadipocytes 3T3-L1 or ASCs from subcutaneous and visceral (retroperitoneal) fat depots from C57Bl/6J mice. Differentiating cells were treated with leptin in addition to or in replacement of insulin. The advance of adipogenesis was assessed by the expression and secretion of adipogenesis-and lipogenesis-related proteins by Western blot and immunoenzimatic assays, and the accumulation of lipid droplets by fluorescence microscopy.Results: Leptin treatment in 3T3-L1 preadipocytes or ASCs increased the production of the adipogenesis-and lipogenesis-related proteins PLIN1, CAV-1, PPARγ, SREBP1C, and/or adiponectin at earlier stages of differentiation. In 3T3-L1 preadipocytes, we found that leptin induced lipid droplets' formation in an mTOR-dependent manner. Also, leptin induced a proinflammatory cytokine profile in 3T3-L1 and ASCs, modulating the production of TNF-α, IL-10, and IL-6. Since insulin is considered an essential factor for preadipocyte differentiation, we asked whether leptin would support adipogenesis in the absence of insulin. Importantly, leptin induced the formation of lipid droplets and the expression of adipogenesis-related proteins independently of insulin during the differentiation of 3T3-L1 cells and ASCs. Conclusions:Our results demonstrate that leptin induces intracellular signaling in preadipocytes and adipocytes promoting adipogenesis and modulating the secretion Palhinha et al.Leptin Induces Adipogenesis of inflammatory mediators. Also, leptin restores adipogenesis in the absence of insulin. These findings contribute to the understanding of the local signaling of leptin in precursor and mature adipose cells. The proadipogenic role of leptin unraveled here may be of especial relevance during obesity, when its central signaling is defective.

Leptin directly activates macrophages and lymphocytes, but the role of leptin in neutrophil activation and migration is still controversial. Here, we investigate the in vivo mechanisms of neutrophil migration induced by leptin. The intraperitoneal injection of leptin (1 mg/kg) induces a time- and concentration-dependent neutrophil influx. We did not observe the enhancement of lipid bodies/droplets in neutrophils, after leptin treatment, as we had observed previously in peritoneal macrophages. The participation of leukotriene B4 (LTB4) in neutrophil recruitment triggered by leptin was investigated using different strategies. Leptin-induced neutrophil recruitment occurs both in the absence of 5-lipoxygenase activity in 5-lipoxygenase (5-LO)−/− mice and after the administration of either 5-LO inhibitor (Zileuton) or the LTB4 receptor antagonist (U-75302). Moreover, no direct induction of LTB4 by leptin could be observed. Neutrophil influx could not be prevented by the mammalian target of rapamycin (mTOR) inhibitor, rapamycin, contrasting with the leptin-induced signaling for lipid body formation in macrophage that is mTOR-dependent. Leptin administration led to tumor necrosis factor-alpha (TNFα) production by the peritoneal cells both in vivo and in vitro. In addition, neutrophil recruitment was inhibited in tumor necrosis factor receptor 1 (TNFR1−/−) mice, indicating a role for TNF in leptin-induced neutrophil recruitment to the peritoneal cavity. Leptin-induced neutrophil influx was PI3Kγ-dependent, as it was absent in PI3Kγ−/− mice. Accordingly, leptin induced the peritoneal cells to produce CXCL1, both in vivo and in vitro, and the neutrophil influx was ablated after using an antibody against CXCL1. Our results establish TNFα/TNFR1- and CXCL1-dependent signaling as important pathways for leptin-induced neutrophil migration in vivo.

Objectives: To evaluate the prevalence of goiter and nodular disease in patients with class III obesity, and to correlate results with serum leptin levels and insulin resistance (IR) parameters. Subjects and methods: A cross-sectional study was performed to assess thyroid ultrasound (US) patterns, HOMA-IR, serum leptin, and TSH levels in obese patients and controls. Results: Thyroid volume was positively correlated with body mass index (BMI) (r = 0.240, p = 0.039) and with HOMA-IR (r = 0.329; p < 0.01). Thyroid US patterns were similar between groups. However, when data from the male group was considered, greater thyroid volume was detected in the obese group compared with controls (10.8 vs. 8.5 cm 3 ; p = 0.04). Also, nodules were more frequently detected (67% vs. 18%), as were nodules requiring FNAB (33.3% vs. 0%, p ≥ 0.05-0.09), in this group. Conclusion: Although IR did not correlate directly with the presence of nodules, the results support the hypothesis of a direct association between insulin resistance and thyroid volume. Arq Bras Endocrinol Metab. 2013;57(2):120-5 Keywords Obesity; insulin resistance; goiter; thyroid nodule RESUMO Objetivos: Avaliar a prevalência de bócio e doença nodular tireoidiana em pacientes com obesidade grau III e correlacionar os resultados com os níveis de leptina e parâmetros de resistência à ação da insulina (RI). Sujeitos e métodos: Estudo seccional foi desenvolvido realizando ultrassonografia (US) tireoidiana e níveis séricos de HOMA-IR e TSH nos pacien tes obesos e nos controles. Resultados: Volume tireoidiano foi positivamente correlacionado com índice de massa corporal (IMC) (r = 0,240, p = 0,039) e com HOMA (r = 0,329; p < 0,01). Volume tireoidiano e prevalência de doença nodular tireoidiana foram similares entre os grupos. Quando avaliado o subgrupo masculino, maiores volumes tireoidianos foram detectados no grupo dos obesos comparados aos controles (10,8 vs. 8,5 cm 3 ; p = 0,04), nódulos foram mais frequentes (67% vs. 18%), assim como nódulos com indicação de punção (33,3% vs. 0%, p ≥ 0,05-0,09). Conclusão: Embora RI não se correlacione diretamente com a presença de nódulos, os resultados suportam a hipótese da direta associação entre resistência à ação da insulina e volume tireoidiano. Arq Bras Endocrinol Metab.2013;57(2):120-5 Descritores Obesidade; resistência à insulina; bócio; nódulo da glândula tireoide

Background/ObjectivesThe pathological condition of obesity is accompanied by a dysfunctional adipose tissue. We postulate that subcutaneous, preperitoneal and visceral obese abdominal white adipose tissue depots could have stromal vascular fractions (SVF) with distinct composition and adipose stem cells (ASC) that would differentially account for the pathogenesis of obesity.MethodsIn order to evaluate the distribution of SVF subpopulations, samples of subcutaneous, preperitoneal and visceral adipose tissues from morbidly obese women (n = 12, BMI: 46.2±5.1 kg/m2) were collected during bariatric surgery, enzymatically digested and analyzed by flow cytometry (n = 12). ASC from all depots were evaluated for morphology, surface expression, ability to accumulate lipid after induction and cytokine secretion (n = 3).ResultsA high content of preadipocytes was found in the SVF of subcutaneous depot (p = 0.0178). ASC from the three depots had similar fibroblastoid morphology with a homogeneous expression of CD34, CD146, CD105, CD73 and CD90. ASC from the visceral depot secreted the highest levels of IL-6, MCP-1 and G-CSF (p = 0.0278). Interestingly, preperitoneal ASC under lipid accumulation stimulus showed the lowest levels of all the secreted cytokines, except for adiponectin that was enhanced (p = 0.0278).ConclusionsASC from preperitoneal adipose tissue revealed the less pro-inflammatory properties, although it is an internal adipose depot. Conversely, ASC from visceral adipose tissue are the most pro-inflammatory. Therefore, ASC from subcutaneous, visceral and preperitoneal adipose depots could differentially contribute to the chronic inflammatory scenario of obesity.

Dengue is characterized as one of the most important arthropod-borne human viral diseases, representing a public health problem. Increased activation of immune cells is involved in the progression of infection to severe forms. Recently, our group demonstrated the contribution of platelet-monocyte interaction to inflammatory responses in dengue, adding to evolving evidence that platelets have inflammatory functions and can regulate different aspects of innate immune responses. Furthermore, stimuli-specific-activated platelets can promote phenotypic changes and metabolic reprogramming in monocytes. Thus, this study aimed to evaluate the roles of dengue virus (DENV)-activated platelets on immunometabolic reprogramming of monocytes in vitro, focusing on lipid droplet (LD) biogenesis. We demonstrated that platelets exposed to DENV in vitro form aggregates with monocytes and signal to LD formation and CXCL8/IL-8, IL-10, CCL2, and PGE 2 secretion. Pharmacologic inhibition of LD biogenesis prevents PGE 2 secretion, but not CXCL8/IL-8 release, by platelet-monocyte complexes. In exploring the mechanisms involved, we demonstrated that LD formation in monocytes exposed to DENV-activated platelets is partially dependent on platelet-produced MIF. Additionally, LD formation is higher in monocytes, which have platelets adhered on their surface, suggesting that beyond paracrine signaling, platelet adhesion is an important event in platelet-mediated modulation of lipid metabolism in monocytes. Together, our results demonstrate that activated platelets aggregate with monocytes during DENV infection and signal to LD biogenesis and the secretion of inflammatory mediators, which may contribute to dengue immunopathogenesis.

Uncaria tomentosa (Willd.) DC. e U. guianensis (Aubl.) Gmel., conhecidas como unha-de-gato, são trepadeiras lenhosas nativas das florestas tropicais amazônica e central-americana. As espécies contêm, em diferentes proporções, alcalóides indólicos e oxindólicos, triterpenóides glicosilados, esteróides e proantocianidinas. U. tomentosa é quimicamente identificada pelo perfil e conteúdo de alcalóides oxindólicos, ao passo que U. guianensis não possui um marcador químico efetivo. Nesse trabalho descreve-se o isolamento de canferol-3,7-O-(α)-dirramnosídeo (canferitrina) pela primeira vez em espécies do gênero Uncaria. A triagem para essa substância em folhas, galhos ou cascas das duas espécies por CCD e CLAE-DAD-EM demonstrou a presença de canferitrina apenas nas folhas e galhos de U. guianensis, numa proporção cerca de trinta e seis vezes maior nas folhas do que nos galhos. Estes resultados revelaram a seletividade da U. guianensis em produzir o flavonóide glicosilado, sugerindo esta substância como um marcador químico potencial para a espécie.Uncaria tomentosa (Willd.) DC. and U. guianensis (Aubl.) Gmel., known as cat's claw, are large woody vines native to the Amazonian and Central American rainforests. The species contain, in different proportions, indole and oxindole alkaloids, triterpenoid glycosides, sterols and proanthocyanidins. U. tomentosa can be chemically identified by its oxindole alkaloid profile and content, whereas U. guianensis has no satisfactorily established chemical markers. This work describes, for the first time, the isolation of kaempferol-3,7-O-(α)-dirhamnoside (kaempferitrin) in Uncaria species. Screening for this compound in leaves, stems or bark of both species through TLC and HPLC-DAD-MS showed the presence of kaempferitrin only in the leaves and stems of U. guianensis, at a ratio almost thirty six times greater in the leaves than in the stems. These results reveal the selectivity of U. guianensis to produce this bioactive flavonoid glycoside, and suggest this compound as a potential chemical marker for the species.