Deficiency of the Cytoskeletal Protein SPECC1L Leads to Oblique Facial Clefting

Genetic mutations responsible for oblique facial clefts (ObFC), a unique class of facial malformations, are largely unknown. We show that loss-of-function mutations in SPECC1L. are pathogenic for this human developmental disorder and that SPECC1L is a critical organizer of vertebrate facial morphogenesis. During murine embryogenesis, Speed 1 1 is expressed in cell populations of the developing facial primordial, which proliferate and fuse to form the face. In zebrafish, knockdown of a SPECC1L homolog produces a faceless phenotype with loss of jaw and facial structures, and knockdown in Drosophila phenocopies mutants in the integrin signaling pathway that exhibit cell-migration and -adhesion defects. Furthermore, in mammalian cells, SPECC1L colocalizes with both tubulin and actin, and its deficiency results in defective actin-cytoskeleton reorganization, as well as abnormal cell adhesion and migration. Collectively, these data demonstrate that SPECC1L functions in actin-cytoskeleton reorganization and is required for proper facial morphogenesis.

Free Cyclitol, Soluble Carbohydrate and Protein Contents in Vigna unguiculata and Phaseolus vulgaris Bean Sprouts

Seeds sprouts have been used as a good source of basic nutrients and nutraceutical compounds. The high nutritional value of seeds derives from the deposition of compounds during development. However some of these molecules are used in metabolic processes like germination, which leads to a considerable variation in their concentrations once these events are completed. In this work, we investigate the levels of inositols (myo-inositol, D-pinitol and ononitol), soluble carbohydrates and proteins in cotyledons of Phaseolus vulgaris and Vigna unguiculata sprouts. Sprouting increased myo-inositol and glucose content and reduction of raffinose and ononitol was observed. The protein levels increased in P. vulgaris and decreased in V. unguiculata sprouting. The level of sucrose was maintained in both sprouts. D-Pinitol was detected only in quiescent seeds. Our results suggested that bean sprout is an important source of proteins, sucrose, glucose and myo-inositol. Additionally, bean sprouts have low levels of raffinose, an antinutritional compound.

Toll-Like Receptor 4 Signaling Leads to Severe Fungal Infection Associated with Enhanced Proinflammatory Immunity and Impaired Expansion of Regulatory T Cells

Toll-like receptors (TLRs) present in innate immune cells recognize pathogen molecular patterns and influence immunity to control the host-parasite interaction. The objective of this study was to characterize the involvement of TLR4 in the innate and adaptive immunity to Paracoccidioides brasiliensis, the most important primary fungal pathogen of Latin America. We compared the responses of C3H/HeJ mice, which are naturally defective in TLR4 signaling, with those of C3H/HePas mice, which express functional receptors, after in vitro and in vivo infection with P. brasiliensis. Unexpectedly, we verified that TLR4-defective macrophages infected in vitro with P. brasiliensis presented decreased fungal loads associated with impaired synthesis of nitric oxide, interleukin-12 (IL-12), and macrophage chemotactic protein 1 (MCP-1). After intratracheal infection with 1 million yeasts, TLR4-defective mice developed reduced fungal burdens and decreased levels of pulmonary nitric oxide, proinflammatory cytokines, and antibodies. TLR4-competent mice produced elevated levels of IL-12 and tumor necrosis factor alpha (TNF-alpha), besides cytokines of the Th17 pattern, indicating a proinflammatory role for TLR4 signaling. The more severe infection of TLR4-normal mice resulted in increased influx of activated macrophages and T cells to the lungs and progressive control of fungal burdens but impaired expansion of regulatory T cells (Treg cells). In contrast, TLR4-defective mice were not able to clear their diminished fungal burdens totally, a defect associated with deficient activation of T-cell immunity and enhanced development of Treg cells. These divergent patterns of immunity, however, resulted in equivalent mortality rates, indicating that control of elevated fungal growth mediated by vigorous inflammatory reactions is as deleterious to the hosts as low fungal loads inefficiently controlled by limited inflammatory reactions.

Gradual Decline in Malaria-Specific Memory T Cell Responses Leads to Failure to Maintain Long-Term Protective Immunity to Plasmodium chabaudi AS Despite Persistence of B Cell Memory and Circulating Antibody

The mechanisms responsible for the generation and maintenance of immunological memory to Plasmodium are poorly understood and the reasons why protective immunity in humans is so difficult to achieve and rapidly lost remain a matter for debate. A possible explanation for the difficulty in building up an efficient immune response against this parasite is the massive T cell apoptosis resulting from exposure to high-dose parasite Ag. To determine the immunological mechanisms required for long-term protection against P. chabaudi malaria and the consequences of high and low acute phase parasite loads for acquisition of protective immunity, we performed a detailed analysis of T and B cell compartments over a period of 200 days following untreated and drug-treated infections in female C57BL/6 mice. By comparing several immunological parameters with the capacity to control a secondary parasite challenge, we concluded that loss of full protective immunity is not determined by acute phase parasite load nor by serum levels of specific IgG2a and IgG1. Abs, but appears to be a consequence of the progressive decline in memory T cell response to parasites, which occurs similarly in untreated and drug-treated mice with time after infection. Furthermore, by analyzing adoptive transfer experiments, we confirmed the major role of CD4(+) T cells for guaranteeing long-term full protection against P. chabaudi malaria. The Journal of Immunology, 2008, 181: 8344-8355.

Inflammation of the Hypothalamus Leads to Defective Pancreatic Islet Function

Type 2 diabetes mellitus results from the complex association of insulin resistance and pancreatic beta-cell failure. Obesity is the main risk factor for type 2 diabetes mellitus, and recent studies have shown that, in diet-induced obesity, the hypothalamus becomes inflamed and dysfunctional, resulting in the loss of the perfect coupling between caloric intake and energy expenditure. Because pancreatic beta-cell function is, in part, under the control of the autonomic nervous system, we evaluated the role of hypothalamic inflammation in pancreatic islet function. In diet-induced obesity, the earliest markers of hypothalamic inflammation are present at 8 weeks after the beginning of the high fat diet; similarly, the loss of the first phase of insulin secretion is detected at the same time point and is restored following sympathectomy. Intracerebroventricular injection of a low dose of tumor necrosis factor a leads to a dysfunctional increase in insulin secretion and activates the expression of a number of markers of apoptosis in pancreatic islets. In addition, the injection of stearic acid intracerebroventricularly, which leads to hypothalamic inflammation through the activation of tau-like receptor-4 and endoplasmic reticulum stress, produces an impairment of insulin secretion, accompanied by increased expression of markers of apoptosis. The defective insulin secretion, in this case, is partially dependent on sympathetic signal-induced peroxisome proliferator receptor-gamma coactivator Delta a and uncoupling protein-2 expression and is restored after sympathectomy or following PGC1 alpha expression inhibition by an antisense oligonucleotide. Thus, the autonomic signals generated in concert with hypothalamic inflammation can impair pancreatic islet function, a phenomenon that may explain the early link between obesity and defective insulin secretion.

Gene optimization leads to robust expression of human respiratory syncytial virus nucleoprotein and phosphoprotein in human cells and induction of humoral immunity in mice

Human respiratory syncytial virus (HRSV) is the major pathogen leading to respiratory disease in infants and neonates worldwide. An effective vaccine has not yet been developed against this virus, despite considerable efforts in basic and clinical research. HRSV replication is independent of the nuclear RNA processing constraints, since the virus genes are adapted to the cytoplasmic transcription, a process performed by the viral RNA-dependent RNA polymerase. This study shows that meaningful nuclear RNA polymerase II dependent expression of the HRSV nucleoprotein (N) and phosphoprotein (F) proteins can only be achieved with the optimization of their genes, and that the intracellular localization of N and P proteins changes when they are expressed out of the virus replication context. Immunization tests performed in mice resulted in the induction of humoral immunity using the optimized genes. This result was not observed for the non-optimized genes. In conclusion, optimization is a valuable tool for improving expression of HRSV genes in DNA vaccines. (c) 2009 Elsevier B.V. All rights reserved.

Ab Initio Calculations of Structural Evolution and Conductance of Benzene-1,4-dithiol on Gold Leads

By performing at) initio density functional theory (DFT) calculations and electronic transport simulations based on the OFT nonequilibrium Green`s functions method we investigate how the conformational changes of a benzene-1,4-dithiol molecule bonded to gold affect the molecular transport as the electrodes are separated from each other. In particular we consider the full evolution of the stretching process until the Junction breaking point and compare results obtained with a standard semilocal exchange and correlation functional to those computed with a self-interaction corrected method. We conclude that the inclusion of self-interaction corrections is fundamental for describing both the molecule conductance and its stability against conformational fluctuations.

Long-term intermittent feeding, but not caloric restriction, leads to redox imbalance, insulin receptor nitration, and glucose intolerance

Calorie restriction is a dietary intervention known to improve redox state, glucose tolerance, and animal life span. Other interventions have been adopted as study models for caloric restriction, including nonsupplemented food restriction and intermittent, every-other-day feedings. We compared the short- and long-term effects of these interventions to ad libitum protocols and found that, although all restricted diets decrease body weight, intermittent feeding did not decrease intra-abdominal adiposity. Short-term calorie restriction and intermittent feeding presented similar results relative to glucose tolerance. Surprisingly, long-term intermittent feeding promoted glucose intolerance, without a loss in insulin receptor phosphorylation. Intermittent feeding substantially increased insulin receptor nitration in both intra-abdominal adipose tissue and muscle, a modification associated with receptor inactivation. All restricted diets enhanced nitric oxide synthase levels in the insulin-responsive adipose tissue and skeletal muscle. However, whereas calorie restriction improved tissue redox state, food restriction and intermittent feedings did not. In fact, long-term intermittent feeding resulted in largely enhanced tissue release of oxidants. Overall, our results show that restricted diets are significantly different in their effects on glucose tolerance and redox state when adopted long-term. Furthermore, we show that intermittent feeding can lead to oxidative insulin receptor inactivation and glucose intolerance. (C) 2011 Elsevier Inc. All rights reserved.