Peroxisome proliferator-activated receptor beta/delta exerts a strong protection from ischemic acute renal failure.

Ischemic acute renal failure is characterized by damages to the proximal straight tubule in the outer medulla. Lesions include loss of polarity, shedding into the tubule lumen, and eventually necrotic or apoptotic death of epithelial cells. It was recently shown that peroxisome proliferator-activated receptor beta/delta (PPARbeta/delta) increases keratinocyte survival after an inflammatory reaction. Therefore, whether PPARbeta/delta could contribute also to the control of tubular epithelium death after renal ischemia/reperfusion was tested. It was found that PPARbeta/delta+/- and PPARbeta/delta-/- mutant mice exhibited much greater kidney dysfunction and injury than wild-type counterparts after a 30-min renal ischemia followed by a 36-h reperfusion. Conversely, wild-type mice that were given the specific PPARbeta/delta ligand L-165041 before renal ischemia were completely protected against renal dysfunction, as indicated by the lack of rise in serum creatinine and fractional excretion of Na+. This protective effect was accompanied by a significant reduction in medullary necrosis, apoptosis, and inflammation. On the basis of in vitro studies, PPARbeta/delta ligands seem to exert their role by activating the antiapoptotic Akt signaling pathway and, unexpectedly, by increasing the spreading of tubular epithelial cells, thus limiting potentially their shedding and anoikis. These results point to PPARbeta/delta as a remarkable new target for preconditioning strategies.

Cross-presenting dendritic cells are required for control of Leishmania major infection.

Leishmania major infection induces self-healing cutaneous lesions in C57BL/6 mice. Both IL-12 and IFN-γ are essential for the control of infection. We infected Jun dimerization protein p21SNFT (Batf3(-/-) ) mice (C57BL/6 background) that lack the major IL-12 producing and cross-presenting CD8α(+) and CD103(+) DC subsets. Batf3(-/-) mice displayed enhanced susceptibility with larger lesions and higher parasite burden. Additionally, cells from draining lymph nodes of infected Batf3(-/-) mice secreted less IFN-γ, but more Th2- and Th17-type cytokines, mirrored by increased serum IgE and Leishmania-specific immunoglobulin 1 (Th2 indicating). Importantly, CD8α(+) DCs isolated from lymph nodes of L. major-infected mice induced significantly more IFN-γ secretion by L. major-stimulated immune T cells than CD103(+) DCs. We next developed CD11c-diptheria toxin receptor: Batf3(-/-) mixed bone marrow chimeras to determine when the DCs are important for the control of infection. Mice depleted of Batf-3-dependent DCs from day 17 or wild-type mice depleted of cross-presenting DCs from 17-19 days after infection maintained significantly larger lesions similar to mice whose Batf-3-dependent DCs were depleted from the onset of infection. Thus, we have identified a crucial role for Batf-3-dependent DCs in protection against L. major.

Estimation of Aedes aegypti (Diptera: Culicidae) population size and adult male survival in an urban area in Panama

Traditional mosquito control strategies rely heavily on the use of chemical insecticides. However, concerns about the efficiency of traditional control methods, environmental impact and emerging pesticide resistance have highlighted the necessity for developing innovative tools for mosquito control. Some novel strategies, including release of insects carrying a dominant lethal gene (RIDL®), rely on the sustained release of modified male mosquitoes and therefore benefit from a thorough understanding of the biology of the male of the species. In this report we present the results of a mark-release-recapture study aimed at: (i) establishing the survival in the field of laboratory-reared, wild-type male Aedes aegypti and (b) estimating the size of the local adult Ae. aegypti population. The study took place in Panama, a country where recent increases in the incidence and severity of dengue cases have prompted health authorities to evaluate alternative strategies for vector control. Results suggest a life expectancy of 2.3 days for released male mosquitoes (confidence interval: 1.78-2.86). Overall, the male mosquito population was estimated at 58 males/ha (range 12-81 males/ha), which can be extrapolated to an average of 0.64 pupae/person for the study area. The practical implications of these results are discussed.

Blockade of both alpha1A- and alpha1B-adrenergic receptor subtype signaling is required to inhibit neointimal formation in the mouse femoral artery.

Attenuation of early restenosis after percutaneous coronary intervention (PCI) is important for the successful treatment of coronary artery disease. Some clinical studies have shown that hypertension is a risk factor for early restenosis after PCI. These findings suggest that alpha(1)-adrenergic receptors (alpha(1)-ARs) may facilitate restenosis after PCI because of alpha(1)-AR's remarkable contribution to the onset of hypertension. In this study, we examined the neointimal formation after vascular injury in the femoral artery of alpha(1A)-knockout (alpha(1A)-KO), alpha(1B)-KO, alpha(1D)-KO, alpha(1A)-/alpha(1B)-AR double-KO (alpha(1AB)-KO), and wild-type mice to investigate the functional role of each alpha(1)-AR subtype in neointimal formation, which is known to promote restenosis. Neointimal formation 4 wk after wire injury was significantly (P < 0.05) smaller in alpha(1AB)-KO mice than in any other group of mice, while blood pressures were not altered in any of the groups of mice after wire injury compared with those before it. These results suggest that lack of both alpha(1A)- and alpha(1B)-ARs could be necessary to inhibit neointimal formation in the mouse femoral artery.

PARP inhibition attenuates histopathological lesion in ischemia/reperfusion renal mouse model after cold prolonged ischemia.

We test the hypothesis that PARP inhibition can decrease acute tubular necrosis (ATN) and other renal lesions related to prolonged cold ischemia/reperfusion (IR) in kidneys preserved at 4°C in University of Wisconsin (UW) solution. Material and Methods. We used 30 male Parp1(+/+) wild-type and 15 male Parp1(0/0) knockout C57BL/6 mice. Fifteen of these wild-type mice were pretreated with 3,4-dihydro-5-[4-(1-piperidinyl)butoxyl]-1(2H)-isoquinolinone (DPQ) at a concentration of 15 mg/kg body weight, used as PARP inhibitor. Subgroups of mice were established (A: IR 45 min/6 h; B: IR + 48 h in UW solution; and C: IR + 48 h in UW solution plus DPQ). We processed samples for morphological, immunohistochemical, ultrastructural, and western-blotting studies. Results. Prolonged cold ischemia time in UW solution increased PARP-1 expression and kidney injury. Preconditioning with PARP inhibitor DPQ plus DPQ supplementation in UW solution decreased PARP-1 nuclear expression in renal tubules and renal damage. Parp1(0/0) knockout mice were more resistant to IR-induced renal lesion. In conclusion, PARP inhibition attenuates ATN and other IR-related renal lesions in mouse kidneys under prolonged cold storage in UW solution. If confirmed, these data suggest that pharmacological manipulation of PARP activity may have salutary effects in cold-stored organs at transplantation.

PARP inhibition attenuates histopathological lesion in ischemia/reperfusion renal mouse model after cold prolonged ischemia.

We test the hypothesis that PARP inhibition can decrease acute tubular necrosis (ATN) and other renal lesions related to prolonged cold ischemia/reperfusion (IR) in kidneys preserved at 4°C in University of Wisconsin (UW) solution. Material and Methods. We used 30 male Parp1(+/+) wild-type and 15 male Parp1(0/0) knockout C57BL/6 mice. Fifteen of these wild-type mice were pretreated with 3,4-dihydro-5-[4-(1-piperidinyl)butoxyl]-1(2H)-isoquinolinone (DPQ) at a concentration of 15 mg/kg body weight, used as PARP inhibitor. Subgroups of mice were established (A: IR 45 min/6 h; B: IR + 48 h in UW solution; and C: IR + 48 h in UW solution plus DPQ). We processed samples for morphological, immunohistochemical, ultrastructural, and western-blotting studies. Results. Prolonged cold ischemia time in UW solution increased PARP-1 expression and kidney injury. Preconditioning with PARP inhibitor DPQ plus DPQ supplementation in UW solution decreased PARP-1 nuclear expression in renal tubules and renal damage. Parp1(0/0) knockout mice were more resistant to IR-induced renal lesion. In conclusion, PARP inhibition attenuates ATN and other IR-related renal lesions in mouse kidneys under prolonged cold storage in UW solution. If confirmed, these data suggest that pharmacological manipulation of PARP activity may have salutary effects in cold-stored organs at transplantation.

A novel ABCG-like transporter of Trypanosoma cruziis involved in natural resistance to benznidazole

Benznidazole (BZ) is one of the two drugs used for Chagas disease treatment. Nevertheless therapeutic failures of BZ have been reported, which were mostly attributed to variable drug susceptibility among Trypanosoma cruzistrains. ATP-binding cassette (ABC) transporters are involved in a variety of translocation processes and some members have been implicated in drug resistance. Here we report the characterisation of the first T. cruzi ABCG transporter gene, named TcABCG1, which is over-expressed in parasite strains naturally resistant to BZ. Comparison ofTcABCG1 gene sequence of two TcI BZ-resistant strains with CL Brener BZ-susceptible strain showed several single nucleotide polymorphisms, which determined 11 amino acid changes. CL Brener transfected with TcI transporter genes showed 40-47% increased resistance to BZ, whereas no statistical significant increment in drug resistance was observed when CL Brener was transfected with the homologous gene. Only in the parasites transfected with TcI genes there was 2-2.6-fold increased abundance of TcABCG1transporter protein. The analysis in wild type strains also suggests that the level of TcABCG1transporter is related to BZ natural resistance. The characteristics of untranslated regions of TcABCG1genes of BZ-susceptible and resistant strains were investigated by computational tools.

Functional and molecular evolutionary analysis of the "Brevix Radix" gene family in mono-and dicotyledonous plants

ABSTRACTIn contrast to animals, plants cannot move from their place of birth and, therefore, need to adapt to their particular habitat in order to survive. Thus, plant development is remarkably plastic, making plants an ideal system for the isolation of genes that account for intraspecific natural variation and possibly environmental adaptation. However, to date, this approach mostly identified null alleles and missed mutations with subtle effects. For instance, BREVIS RADIX (BRX) has been isolated as a key regulator of root growth through a naturally occurring loss-of-function allele in the Arabidopsis thaliana accession Uk-1 and is the founding member of a highly-conserved plant-specific gene family.In this work, we show that a strong selective pressure is acting on the BRX gene family and dates back before the monocot-dicot divergence. However, functional diversification is observed mainly in dicotyledon BRX family genes and is correlated with acceleration in the evolutionary rates in the N-terminal regions. Population genetic data revealed that BRX is highly conserved across Arabidopsis accessions and presents signatures of adaptation. Interestingly, a seven amino acid deletion polymorphism in BRX sequence was found in a few accessions, which seems to be responsible for their enhanced primary root growth. Nevertheless, BRX might not only be active in the root, as suggested by its expression in the shoot. Indeed, leaves and cotyledons of brx mutants are significantly smaller than wild- type. This phenotype is a direct consequence of the absence of BRX function in the shoot rather than an indirect effect of an altered root system growth. Interestingly, cotyledons of brx plants reflect the same physiological defects as the root. Moreover, phenotypes in BRX gain-of-function plants, such as epinastic leaves and increased epidermal cell size, could be associated with an increase in leaf brassinosteroid content.Collectively, these results indicate that BRX contributes to local adaptation by ubiquitously regulating plant growth, probably through the modulation of brassinosteroid biosynthesis.RÉSUMÉContrairement à la plupart des animaux, les plantes ne peuvent se mouvoir et doivent ainsi s'adapter à leur environnement pour survivre. Pour cette raison, elles représentent un système idéal pour l'identification de gènes contribuant à la variation naturelle intra- spécifique, ainsi qu'à l'adaptation. Cependant, cette approche a, jusqu'à présent, surtout permis d'isoler des allèles nuls et non des mutations conférant des effets plus subtiles. C'est le cas du gène Β REVIS RADIX (BRX), un régulateur clé de la croissance racinaire, qui a été identifié grâce à un allèle non-fonctionnel présent dans l'accession naturelle d'Arabidopsis thaliana Uk-1. BRX et ses homologues des plantes mono- et dicotylédones forment une famille très conservée et spécifique aux plantes.Dans ce travail, nous démontrons que la famille de gènes BRX est soumise à une forte pression de sélection qui remonte avant la divergence entre mono- et dicotylédones. Cependant, une diversification fonctionnelle a été observée chez les gènes des dicotylédones et corrèle avec une accélération de la vitesse d'évolution dans leur région N- terminale. Une analyse génétique de différentes accessions naturelles d'Arabidopsis a révélé que BRX est hautement conservé et présente des signatures d'adaptation. Remarquablement, un polymorphisme de délétion de sept acides aminés a été détecté dans quelques accessions et a pour conséquence une plus forte croissance de la racine primaire. Néanmoins, il semble que le rôle de BRX ne se limite pas qu'à la racine, comme indiqué par son expression dans les parties aériennes de la plante. En effet, les mutants brx présentent des cotylédons et des feuilles significativement plus petits que le type sauvage, une conséquence directe de l'absence d'activité de BRX dans ces organes. Nous avons aussi noté que les cotylédons des mutants brx, à l'instar des racines, ont une perception altérée de l'auxine et peuvent être complémentés par l'application exogène de brassinostéroïdes. De plus, dans des plantes présentant un gain de fonction BRX, les feuilles sont épinastiques et les cellules de leur épiderme plus grandes. Ces phénotypes sont accompagnés d'une augmentation de la concentration de brassinostéroïdes dans les feuilles. Conjointement, ces résultats démontrent que BRX contribue à une adaptation locale de la plante par la régulation générale de sa croissance, probablement en modulant la biosynthèse des brassinostéroïdes.

CD1d-restricted NK T cells are dispensable for specific antibody responses and protective immunity against liver stage malaria infection in mice.

Immunization with a single dose of irradiated sporozoites is sufficient to induce protection against malaria in wild-type mice. Although this protection is classically attributed to conventional CD4+ and CD8+ T cells, several recent reports have suggested an important role for CD1-restricted NK T cells in immunity to malaria. In this study, we directly compared the ability of C57BL/6 wild-type and CD1-deficient mice to mount a protective immune response against Plasmodium berghei sporozoites. Our data indicate that CD1-restricted NK T cells are not required for protection in this model system. Moreover, specific IgG antibody responses to the P. berghei circumsporozoite repeat sequence were also unaffected by CD1 deficiency. Collectively, our data demonstrate that CD1-restricted NK T cells are dispensable for protective immunity to liver stage P. berghei infection.

Inverse agonism and neutral antagonism at alpha(1a)- and alpha(1b)-adrenergic receptor subtypes.

We have characterized the pharmacological antagonism, i.e., neutral antagonism or inverse agonism, displayed by a number of alpha-blockers at two alpha1-adrenergic receptor (AR) subtypes, alpha(1a)- and alpha(1b)-AR. Constitutively activating mutations were introduced into the alpha(1a)-AR at the position homologous to A293 of the alpha(1b)-AR where activating mutations were previously described. Twenty-four alpha-blockers differing in their chemical structures were initially tested for their effect on the agonist-independent inositol phosphate response mediated by the constitutively active A271E and A293E mutants expressed in COS-7 cells. A selected number of drugs also were tested for their effect on the small, but measurable spontaneous activity of the wild-type alpha(1a)- and alpha(1b)-AR expressed in COS-7 cells. The results of our study demonstrate that a large number of structurally different alpha-blockers display profound negative efficacy at both the alpha(1a)- and alpha(1b)-AR subtypes. For other drugs, the negative efficacy varied at the different constitutively active mutants. The most striking difference concerns a group of N-arylpiperazines, including 8-[2-[4-(5-chloro-2-methoxyphenyl)-1-piperazinyl]ethyl]-8-azaspiro [4, 5] decane-7,9-dione (REC 15/3039), REC 15/2739, and REC 15/3011, which are inverse agonists with profound negative efficacy at the wild-type alpha(1b)-AR, but not at the alpha(1a)-AR.

Tau hyperphosphorylation and increased BACE1 and RAGE levels in the cortex of PPARβ/δ-null mice.

The role of peroxisome proliferator activator receptor (PPAR)β/δ in the pathogenesis of Alzheimer's disease has only recently been explored through the use of PPARβ/δ agonists. Here we evaluated the effects of PPARβ/δ deficiency on the amyloidogenic pathway and tau hyperphosphorylation. PPARβ/δ-null mice showed cognitive impairment in the object recognition task, accompanied by enhanced DNA-binding activity of NF-κB in the cortex and increased expression of IL-6. In addition, two NF-κB-target genes involved in β-amyloid (Aβ) synthesis and deposition, the β site APP cleaving enzyme 1 (Bace1) and the receptor for advanced glycation endproducts (Rage), respectively, increased in PPARβ/δ-null mice compared to wild type animals. The protein levels of glial fibrillary acidic protein (GFAP) increased in the cortex of PPARβ/δ-null mice, which would suggest the presence of astrogliosis. Finally, tau hyperphosphorylation at Ser199 and enhanced levels of PHF-tau were associated with increased levels of the tau kinases CDK5 and phospho-ERK1/2 in the cortex of PPARβ/δ(-/-) mice. Collectively, our findings indicate that PPARβ/δ deficiency results in cognitive impairment associated with enhanced inflammation, astrogliosis and tau hyperphosphorylation in the cortex.

A new factor stimulating peptidoglycan hydrolysis to separate daughter cells in Caulobacter crescentus.

Cell division in Gram-negative bacteria involves the co-ordinated invagination of the three cell envelope layers to form two new daughter cell poles. This complex process starts with the polymerization of the tubulin-like protein FtsZ into a Z-ring at mid-cell, which drives cytokinesis and recruits numerous other proteins to the division site. These proteins are involved in Z-ring constriction, inner- and outer-membrane invagination, peptidoglycan remodelling and daughter cell separation. Three papers in this issue of Molecular Microbiology, from the teams of Lucy Shapiro, Martin Thanbichler and Christine Jacobs-Wagner, describe a novel protein, called DipM for Division Involved Protein with LysM domains, that is required for cell division in Caulobacter crescentus. DipM localizes to the mid-cell during cell division, where it is necessary for the hydrolysis of the septal peptidoglycan to remodel the cell wall. Loss of DipM results in severe defects in cell envelope constriction, which is deleterious under fast-growth conditions. State-of-the-art microscopy experiments reveal that the peptidoglycan is thicker and that the cell wall is incorrectly organized in DipM-depleted cells compared with wild-type cells, demonstrating that DipM is essential for reorganizing the cell wall at the division site, for envelope invagination and cell separation in Caulobacter.

A molecular platform for the diagnosis of multidrug-resistant and pre-extensively drug-resistant tuberculosis based on single nucleotide polymorphism mutations present in Colombian isolates of Mycobacterium tuberculosis

Developing a fast, inexpensive, and specific test that reflects the mutations present in Mycobacterium tuberculosis isolates according to geographic region is the main challenge for drug-resistant tuberculosis (TB) control. The objective of this study was to develop a molecular platform to make a rapid diagnosis of multidrug-resistant (MDR) and extensively drug-resistant TB based on single nucleotide polymorphism (SNP) mutations present in therpoB, katG, inhA,ahpC, and gyrA genes from Colombian M. tuberculosis isolates. The amplification and sequencing of each target gene was performed. Capture oligonucleotides, which were tested before being used with isolates to assess the performance, were designed for wild type and mutated codons, and the platform was standardised based on the reverse hybridisation principle. This method was tested on DNA samples extracted from clinical isolates from 160 Colombian patients who were previously phenotypically and genotypically characterised as having susceptible or MDR M. tuberculosis. For our method, the kappa index of the sequencing results was 0,966, 0,825, 0,766, 0,740, and 0,625 forrpoB, katG, inhA,ahpC, and gyrA, respectively. Sensitivity and specificity were ranked between 90-100% compared with those of phenotypic drug susceptibility testing. Our assay helps to pave the way for implementation locally and for specifically adapted methods that can simultaneously detect drug resistance mutations to first and second-line drugs within a few hours.

Glucose-dependent insulinotropic polypeptide (GIP) and its receptor (GIPR): cellular localization, lesion-affected expression, and impaired regenerative axonal growth.

Glucose-dependent insulinotropic polypeptide (GIP) was initially described to be rapidly regulated by endocrine cells in response to nutrient ingestion, with stimulatory effects on insulin synthesis and release. Previously, we demonstrated a significant up-regulation of GIP mRNA in the rat subiculum after fornix injury. To gain more insight into the lesion-induced expression of GIP and its receptor (GIPR), expression profiles of the mRNAs were studied after rat sciatic nerve crush injury in 1) affected lumbar dorsal root ganglia (DRG), 2) spinal cord segments, and 3) proximal and distal nerve fragments by means of quantitative RT-PCR. Our results clearly identified lesion-induced as well as tissue type-specific mRNA regulation of GIP and its receptor. Furthermore, comprehensive immunohistochemical stainings not only confirmed and exceeded the previous observation of neuronal GIP expression but also revealed corresponding GIPR expression, implying putative modulatory functions of GIP/GIPR signaling in adult neurons. In complement, we also observed expression of GIP and its receptor in myelinating Schwann cells and oligodendrocytes. Polarized localization of GIPR in the abaxonal Schwann cell membranes, plasma membrane-associated GIPR expression of satellite cells, and ependymal GIPR expression strongly suggests complex cell type-specific functions of GIP and GIPR in the adult nervous system that are presumably mediated by autocrine and paracrine interactions, respectively. Notably, in vivo analyses with GIPR-deficient mice suggest a critical role of GIP/GIPR signal transduction in promoting spontaneous recovery after nerve crush, insofar as traumatic injury of GIPR-deficient mouse sciatic nerve revealed impaired axonal regeneration compared with wild-type mice.

Influence of the LILRA3 Deletion on Multiple Sclerosis Risk: Original Data and Meta-Analysis.

BACKGROUND Multiple sclerosis (MS) is a neurodegenerative, autoimmune disease of the central nervous system. Genome-wide association studies (GWAS) have identified over hundred polymorphisms with modest individual effects in MS susceptibility and they have confirmed the main individual effect of the Major Histocompatibility Complex. Additional risk loci with immunologically relevant genes were found significantly overrepresented. Nonetheless, it is accepted that most of the genetic architecture underlying susceptibility to the disease remains to be defined. Candidate association studies of the leukocyte immunoglobulin-like receptor LILRA3 gene in MS have been repeatedly reported with inconsistent results. OBJECTIVES In an attempt to shed some light on these controversial findings, a combined analysis was performed including the previously published datasets and three newly genotyped cohorts. Both wild-type and deleted LILRA3 alleles were discriminated in a single-tube PCR amplification and the resulting products were visualized by their different electrophoretic mobilities. RESULTS AND CONCLUSION Overall, this meta-analysis involved 3200 MS patients and 3069 matched healthy controls and it did not evidence significant association of the LILRA3 deletion [carriers of LILRA3 deletion: p = 0.25, OR (95% CI) = 1.07 (0.95-1.19)], even after stratification by gender and the HLA-DRB1*15:01 risk allele.