Earthworm leukocytes react with different mammalian antigen-specific monoclonal antibodies

We identified conserved molecules (enzymes, peptides, cytokines) that might play a role in invertebrate innate immunity. We found these molecules by immunoserological and immunohistochemical methods in association with coelomocytes, leukocytes located in the coelomic cavity of the earthworm Eisenia foetida. We detected the enzyme Cu-Zn-superoxide-dismutase (SOD), cytokines (tumor necrosis factor-alpha, TNFalpha; transforming growth factor-alpha, TGFalpha; and alpha peptide hormone, thyreotrope stimulating hormone, TSH) in earthworm coelomocytes with monoclonal antibodies developed originally against human and/or mouse antigens. Three coelomocyte subpopulations were identified according to their form, size and granularity by microscopic and flow cytometric analysis. These cell populations showed different reactivity with antibodies against mammalian cell surface (CD) markers and different intracellular antigens. Two coelomocyte types showed cell surface positivity with anti-Thy-1 (CD90), CD24 and TNF-alpha antibodies. Strong cytoplasmic reaction was shown with anti-TNF-alpha and anti-SOD mAbs and a weaker but unambiguous reaction with thyroid stimulating hormone (TSH) in two cell populations. The third population was negative for all of the monoclonal antibodies. Our flow cytometric results were confirmed by confocal microscopy both on the cell surfaces and intracellularly.     

Dynamic prestress in a globular protein

A protein at equilibrium is commonly thought of as a fully relaxed structure, with the intra-molecular interactions showing fluctuations around their energy minimum. In contrast, here we find direct evidence for a protein as a molecular tensegrity structure, comprising a balance of tensed and compressed interactions, a concept that has been put forward for macroscopic structures. We quantified the distribution of inter-residue prestress in ubiquitin and immunoglobulin from all-atom molecular dynamics simulations. The network of highly fluctuating yet significant inter-residue forces in proteins is a consequence of the intrinsic frustration of a protein when sampling its rugged energy landscape. In beta sheets, this balance of forces is found to compress the intra-strand hydrogen bonds. We estimate that the observed magnitude of this pre-compression is enough to induce significant changes in the hydrogen bond lifetimes; thus, prestress, which can be as high as a few 100 pN, can be considered a key factor in determining the unfolding kinetics and pathway of proteins under force. Strong pre-tension in certain salt bridges on the other hand is connected to the thermodynamic stability of ubiquitin. Effective force profiles between some side-chains reveal the signature of multiple, distinct conformational states, and such static disorder could be one factor explaining the growing body of experiments revealing non-exponential unfolding kinetics of proteins. The design of prestress distributions in engineering proteins promises to be a new tool for tailoring the mechanical properties of made-to-order nanomaterials.     

The genetic basis of pollinator adaptation in a sexually deceptive orchid

In plants, pollinator adaptation is considered to be a major driving force for floral diversification and speciation. However, the genetic basis of pollinator adaptation is poorly understood. The orchid genus Ophrys mimics its pollinators' mating signals and is pollinated by male insects during mating attempts. In many species of this genus, chemical mimicry of the pollinators' pheromones, especially of alkenes with different double-bond positions, plays a key role for specific pollinator attraction. Thus, different alkenes produced in different species are probably a consequence of pollinator adaptation. In this study, we identify genes that are likely involved in alkene biosynthesis, encoding stearoyl-acyl carrier protein (ACP) desaturases (SAD), in three closely related Ophrys species, O. garganica, O. sphegodes, and O. exaltata. Combining floral odor and gene expression analyses, two SAD homologs (SAD1/2) showed significant association with the production of (Z)-9- and (Z)-12-alkenes that were abundant in O. garganica and O. sphegodes, supporting previous biochemical data. In contrast, two other newly identified homologs (SAD5/6) were significantly associated with (Z)-7-alkenes that were highly abundant only in O. exaltata. Both molecular evolutionary analyses and pollinator preference tests suggest that the alkenes associated with SAD1/2 and SAD5/6 are under pollinator-mediated divergent selection among species. The expression patterns of these genes in F(1) hybrids indicate that species-specific expression differences in SAD1/2 are likely due to cis-regulation, while changes in SAD5/6 are likely due to trans-regulation. Taken together, we report a genetic mechanism for pollinator-mediated divergent selection that drives adaptive changes in floral alkene biosynthesis involved in reproductive isolation among Ophrys species.     

Bandoniozyma gen. nov., a Genus of Fermentative and Non-Fermentative Tremellaceous Yeast Species

Background

Independent surveys across the globe led to the proposal of a new basidiomycetous yeast genus within the Bulleromyces clade of the Tremellales, Bandoniozyma gen. nov., with seven new species.

Methodology/Principal Findings

The species were characterized by multiple methods, including the analysis of D1/D2 and ITS nucleotide sequences, and morphological and physiological/biochemical traits. Most species can ferment glucose, which is an unusual trait among basidiomycetous yeasts.

Conclusions/Significance

In this study we propose the new yeast genus Bandoniozyma, with seven species Bandoniozyma noutii sp. nov. (type species of genus; CBS 8364^T  =  DBVPG 4489^T), Bandoniozyma aquatica sp. nov. (UFMG-DH4.20^T  =  CBS 12527^T  =  ATCC MYA-4876^T), Bandoniozyma complexa sp. nov. (CBS 11570^T  =  ATCC MYA-4603^T  =  MA28a^T), Bandoniozyma fermentans sp. nov. (CBS 12399^T  =  NU7M71^T  =  BCRC 23267^T), Bandoniozyma glucofermentans sp. nov. (CBS 10381^T  =  NRRL Y-48076^T  =  ATCC MYA-4760^T  =  BG 02-7-15-015A-1-1^T), Bandoniozyma tunnelae sp. nov. (CBS 8024^T  =  DBVPG 7000^T), and Bandoniozyma visegradensis sp. nov. (CBS 12505^T  =  NRRL Y-48783^T  =  NCAIM Y.01952^T).     

Ghrelin Influences Novelty Seeking Behavior in Rodents and Men

Recent discoveries indicate an important role for ghrelin in drug and alcohol reward and an ability of ghrelin to regulate mesolimbic dopamine activity. The role of dopamine in novelty seeking, and the association between this trait and drug and alcohol abuse, led us to hypothesize that ghrelin may influence novelty seeking behavior. To test this possibility we applied several complementary rodent models of novelty seeking behavior, i.e. inescapable novelty-induced locomotor activity (NILA), novelty-induced place preference and novel object exploration, in rats subjected to acute ghrelin receptor (growth hormone secretagogue receptor; GHSR) stimulation or blockade. Furthermore we assessed the possible association between polymorphisms in the genes encoding ghrelin and GHSR and novelty seeking behavior in humans. The rodent studies indicate an important role for ghrelin in a wide range of novelty seeking behaviors. Ghrelin-injected rats exhibited a higher preference for a novel environment and increased novel object exploration. Conversely, those with GHSR blockade drastically reduced their preference for a novel environment and displayed decreased NILA. Importantly, the mesolimbic ventral tegmental area selective GHSR blockade was sufficient to reduce the NILA response indicating that the mesolimbic GHSRs might play an important role in the observed novelty responses. Moreover, in untreated animals, a striking positive correlation between NILA and sucrose reward behavior was detected. Two GHSR single nucleotide polymorphisms (SNPs), rs2948694 and rs495225, were significantly associated with the personality trait novelty seeking, as assessed using the Temperament and Character Inventory (TCI), in human subjects. This study provides the first evidence for a role of ghrelin in novelty seeking behavior in animals and humans, and also points to an association between food reward and novelty seeking in rodents.     

Identification of galectin-1 as a critical factor in function of mouse mesenchymal stromal cell-mediated tumor promotion

Bone marrow derived mesenchymal stromal cells (MSCs) have recently been implicated as one source of the tumor-associated stroma, which plays essential role in regulating tumor progression. In spite of the intensive research, the individual factors in MSCs controlling tumor progression have not been adequately defined. In the present study we have examined the role of galectin-1 (Gal-1), a protein highly expressed in tumors with poor prognosis, in MSCs in the course of tumor development. Co-transplantation of wild type MSCs with 4T1 mouse breast carcinoma cells enhances the incidence of palpable tumors, growth, vascularization and metastasis. It also reduces survival compared to animals treated with tumor cells alone or in combination with Gal-1 knockout MSCs. In vitro studies show that the absence of Gal-1 in MSCs does not affect the number of migrating MSCs toward the tumor cells, which is supported by the in vivo migration of intravenously injected MSCs into the tumor. Moreover, differentiation of endothelial cells into blood vessel-like structures strongly depends on the expression of Gal-1 in MSCs. Vital role of Gal-1 in MSCs has been further verified in Gal-1 knockout mice. By administering B16F10 melanoma cells into Gal-1 deficient animals, tumor growth is highly reduced compared to wild type animals. Nevertheless, co-injection of wild type but not Gal-1 deficient MSCs results in dramatic tumor growth and development.These results confirm that galectin-1 is one of the critical factors in MSCs regulating tumor progression.     

Effect of early detection and treatment on malaria related maternal mortality on the north-western border of Thailand 1986-2010

Introduction

Maternal mortality is high in developing countries, but there are few data in high-risk groups such as migrants and refugees in malaria-endemic areas. Trends in maternal mortality were followed over 25 years in antenatal clinics prospectively established in an area with low seasonal transmission on the north-western border of Thailand.

Methods and Findings

All medical records from women who attended the Shoklo Malaria Research Unit antenatal clinics from 12^th May 1986 to 31^st December 2010 were reviewed, and maternal death records were analyzed for causality. There were 71 pregnancy-related deaths recorded amongst 50,981 women who attended antenatal care at least once. Three were suicide and excluded from the analysis as incidental deaths. The estimated maternal mortality ratio (MMR) overall was 184 (95%CI 150–230) per 100,000 live births. In camps for displaced persons there has been a six-fold decline in the MMR from 499 (95%CI 200–780) in 1986–90 to 79 (40–170) in 2006–10, p<0.05. In migrants from adjacent Myanmar the decline in MMR was less significant: 588 (100–3260) to 252 (150–430) from 1996–2000 to 2006–2010. Mortality from P.falciparum malaria in pregnancy dropped sharply with the introduction of systematic screening and treatment and continued to decline with the reduction in the incidence of malaria in the communities. P.vivax was not a cause of maternal death in this population. Infection (non-puerperal sepsis and P.falciparum malaria) accounted for 39.7 (27/68) % of all deaths.

Conclusions

Frequent antenatal clinic screening allows early detection and treatment of falciparum malaria and substantially reduces maternal mortality from P.falciparum malaria. No significant decline has been observed in deaths from sepsis or other causes in refugee and migrant women on the Thai–Myanmar border.     

Diclofenac Prolongs Repolarization in Ventricular Muscle with Impaired Repolarization Reserve

Background

The aim of the present work was to characterize the electrophysiological effects of the non-steroidal anti-inflammatory drug diclofenac and to study the possible proarrhythmic potency of the drug in ventricular muscle.

Methods

Ion currents were recorded using voltage clamp technique in canine single ventricular cells and action potentials were obtained from canine ventricular preparations using microelectrodes. The proarrhythmic potency of the drug was investigated in an anaesthetized rabbit proarrhythmia model.

Results

Action potentials were slightly lengthened in ventricular muscle but were shortened in Purkinje fibers by diclofenac (20 µM). The maximum upstroke velocity was decreased in both preparations. Larger repolarization prolongation was observed when repolarization reserve was impaired by previous BaCl₂ application. Diclofenac (3 mg/kg) did not prolong while dofetilide (25 µg/kg) significantly lengthened the QT_c interval in anaesthetized rabbits. The addition of diclofenac following reduction of repolarization reserve by dofetilide further prolonged QT_c. Diclofenac alone did not induce Torsades de Pointes ventricular tachycardia (TdP) while TdP incidence following dofetilide was 20%. However, the combination of diclofenac and dofetilide significantly increased TdP incidence (62%). In single ventricular cells diclofenac (30 µM) decreased the amplitude of rapid (I_Kr) and slow (I_Ks) delayed rectifier currents thereby attenuating repolarization reserve. L-type calcium current (I_Ca) was slightly diminished, but the transient outward (I_to) and inward rectifier (I_K1) potassium currents were not influenced.

Conclusions

Diclofenac at therapeutic concentrations and even at high dose does not prolong repolarization markedly and does not increase the risk of arrhythmia in normal heart. However, high dose diclofenac treatment may lengthen repolarization and enhance proarrhythmic risk in hearts with reduced repolarization reserve.