Reduction in diabetes-induced craniofacial defects by maternal immune stimulation

BACKGROUND: Maternal diabetes can induce a number of developmental abnormalities in laboratory animals and humans, including facial deformities and defects in neural tube closure. The incidence of birth defects in newborns of diabetic women is approximately 3-5 times higher than among non-diabetics. In mice, non-specific activation of the maternal immune system can reduce fetal abnormalities caused by diverse etiologies, including diabetes induced neural tube defects. This study was conducted to determine whether non-specific maternal immune stimulation could reduce diabetes-induced craniofacial defects as well. METHODS: Maternal immune function was stimulated before streptozocin (STZ) treatment by maternal footpad injection with Freund's complete adjuvant (FCA), maternal intraperitoneal (i.p.) injection with granulocyte-macrophage colony-stimulating factor (GM-CSF), or maternal i.p. injection with interferon-gamma (IFNgamma). Streptozocin (200 mg/kg i.p.) was used to induce hyperglycemia (26-35 mmol blood glucose) in female ICR mice before breeding. Fetuses from 12-18 litters per treatment group, were collected at Day 17 of gestation. RESULTS: Craniofacial defects were observed in fetuses from all hyperglycemic groups. The incidence of defects was significantly decreased in fetuses from dams immune stimulated with IFNgamma or GM-CSF. The most common defects were reduced maxillary and mandibular lengths. Both were prevented by maternal stimulation with GM-CSF. CONCLUSION: Maternal immune stimulation reduced the incidence of diabetic craniofacial embryopathy. The mechanisms for these protective effects are unknown but may involve maternal or fetal production of cytokines or growth factors that protect the fetus from the dysregulatory effects of hyperglycemia.     

Daily nurse requirements planning based on simulation of patient flows

Nurses account for approximately 50 % of total hospital budgets and their allocation to medical units and shifts can significantly affect the quality of care provided to patients. The adoption of flexible shift schedules and the assessment of actual nursing time can enable sensible resource planning, balancing the quality of care with efficiency in resource use. Starting from the concept that nurse requirements are triggered by patient needs, which are stochastic in nature both for clinical activities and their duration, this paper proposes an innovative Nurse Requirement Planning model grounded on the concept of the clinical pathway (the “standard” sequence of diagnostic, therapeutic and care activities a patient with certain pathology should undertake over time) with its inner routing probability and patient dependence on nurses, which can be correlated to the time needed to perform nursing tasks. In merging and modelling these two aspects, the method summarizes the best features of acuity-quality and timed-task/activity techniques, well known although not usually applied for reasons of demands on clinicians’ time. Instead, in this paper, for each shift of the day, hospital management is enabled to choose the optimal number of nurses to meet actual requirements according to a desired service level and personnel saturation by means of a tool that simulates the patient flow in a medical unit based on automatic data retrieval from hospital databases. The validation and verification of the proposal were undertaken in a stroke unit.     

Diverse lineages of pathogenic Leptospira species are widespread in the environment in Puerto Rico,USA

BackgroundLeptospirosis, caused by Leptospira bacteria, is a common zoonosis worldwide, especially in the tropics. Reservoir species and risk factors have been identified but surveys for environmental sources are rare. Furthermore, understanding of environmental Leptospira containing virulence associated genes and possibly capable of causing disease is incomplete, which may convolute leptospirosis diagnosis, prevention, and epidemiology.Methodology/Principal findingsWe collected environmental samples from 22 sites in Puerto Rico during three sampling periods over 14-months (Dec 2018-Feb 2020); 10 water and 10 soil samples were collected at each site. Samples were screened for DNA from potentially pathogenic Leptospira using the lipL32 PCR assay and positive samples were sequenced to assess genetic diversity. One urban site in San Juan was sampled three times over 14 months to assess persistence in soil; live leptospires were obtained during the last sampling period. Isolates were whole genome sequenced and LipL32 expression was assessed in vitro.We detected pathogenic Leptospira DNA at 15/22 sites; both soil and water were positive at 5/15 sites. We recovered lipL32 sequences from 83/86 positive samples (15/15 positive sites) and secY sequences from 32/86 (10/15 sites); multiple genotypes were identified at 12 sites. These sequences revealed significant diversity across samples, including four novel lipL32 phylogenetic clades within the pathogenic P1 group. Most samples from the serially sampled site were lipL32 positive at each time point. We sequenced the genomes of six saprophytic and two pathogenic Leptospira isolates; the latter represent a novel pathogenic Leptospira species likely belonging to a new serogroup.Conclusions/SignificanceDiverse and novel pathogenic Leptospira are widespread in the environment in Puerto Rico. The disease potential of these lineages is unknown but several were consistently detected for >1 year in soil, which could contaminate water. This work increases understanding of environmental Leptospira diversity and should improve leptospirosis surveillance and diagnostics.     

Spontaneous allelic variant in deafness–blindness gene Ush1g resulting in an expanded phenotype

Relationships between novel phenotypic behaviors and specific genetic alterations are often discovered using target-specific, directed mutagenesis or phenotypic selection following chemical mutagenesis. An alternative approach is to exploit deficiencies in DNA repair pathways that maintain genetic integrity in response to spontaneously induced damage. Mice deficient in the DNA glycosylase NEIL1 show elevated spontaneous mutations, which arise from translesion DNA synthesis past oxidatively induced base damage. Several litters of Neil1 knockout mice included animals that were distinguished by their backwards-walking behavior in open-field environments, while maintaining frantic forward movements in their home cage environment. Other phenotypic manifestations included swim test failures, head tilting and circling. Mapping of the mutation that conferred these behaviors showed the introduction of a stop codon at amino acid 4 of the Ush1g gene. Ush1g^bw/bw null mice displayed auditory and vestibular defects that are commonly seen with mutations affecting inner-ear hair-cell function, including a complete lack of auditory brainstem responses and vestibular-evoked potentials. As in other Usher syndrome type I mutant mouse lines, hair cell phenotypes included disorganized and split hair bundles, as well as altered distribution of proteins for stereocilia that localize to the tips of row 1 or row 2. Disruption to the bundle and kinocilium displacement suggested that USH1G is essential for forming the hair cell's kinocilial links. Consistent with other Usher type 1 models, Ush1g^bw/bw mice had no substantial retinal degeneration compared with Ush1g^bw/+ controls. In contrast to previously described Ush1g alleles, this new allele provides the first knockout model for this gene.     

CD5-Dependent CK2 Activation Pathway Regulates Threshold for T Cell Anergy

CD5 activates casein kinase 2 (CK2), a serine/threonine kinase that constitutively associates with the CK2-binding domain at the end of its cytoplasmic tail. To determine the physiological significance of CD5-dependent CK2 activation in T cells, we generated a knock-in mouse that expresses a CD5 protein containing a microdeletion with selective inability to interact with CK2 (CD5ΔCK2BD). The levels of CD5 on developing and mature T cell populations from CD5ΔCK2BD mice and CD5 wild-type (WT) mice were similar. The thymus of CD5ΔCK2BD mice contained fewer double-positive thymocytes than did that of both CD5WT and CD5 knockout (KO) mice, although the numbers of all other immature and mature T cell populations were unaltered. CD5ΔCK2BD T cells hypoproliferated and exhibited enhanced activation-induced cell death when stimulated with anti-CD3 or cognate peptide in comparison with CD5WT T cells. We also found that functional CD5-dependent CK2 signaling was necessary for efficient differentiation of naive CD4(+) T cells into Th2 and Th17 cells, but not Th1 cells. We previously showed that experimental autoimmune encephalomyelitis (EAE) in CD5KO mice was less severe and delayed in onset than in CD5WT mice. Remarkably, CD5ΔCK2BD mice recapitulated both EAE severity and disease onset of CD5KO mice. Increasing the immunization dose of myelin oligodendrocyte glycoprotein 35-55 peptide, a model that mimics high-dose tolerance, led to decreased severity of EAE in CD5WT mice but not in CD5KO or CD5ΔCK2BD mice. This property was recapitulated in in vitro restimulation assays. These results demonstrate that CD5-CK2 signaling sets the threshold for T cell responsiveness and is necessary for efficient generation of Th2 and Th17 cells.     

Mass spectral analyses of hemoglobin adducts formed after in vitro exposure of erythrocytes to hydroxymethylvinyl ketone

Hydroxymethylvinyl ketone (HMVK) is a reactive oxidation product of 3-butene-1,2-diol, a metabolite of 1,3-butadiene. The potential for HMVK (0.1 and 1mM) to form hemoglobin (Hb) adducts in erythrocytes from Sprague-Dawley rats was investigated at physiological conditions (pH 7.4, 37 degrees C) using electrospray ionization mass spectrometry (ESI/MS). With the 0.1mM HMVK globin samples, the results indicate HMVK adduction on the alpha2, beta2 and beta3 chains. With the 1.0mM HMVK globin samples, adducts were detected on the beta2 and beta3 chains. However, no correlation was observed between incubation time and the extent of adduct formation, and additional adducts were detected when globin samples were fractionated by HPLC before the ESI/MS analyses. For specific localizations of adducts on the globin chains, trypsin digested peptides from the 1mM HMVK globin samples were subjected to liquid chromatography/mass spectrometry analyses. The results, which are consistent with formation of HMVK adducts on several specific peptides within the alpha- and beta-chains, suggest selectivity in the interaction of HMVK with the different cysteine residues in Hb. Because adducts were also detected in peptides containing no cysteine residues and multiple HMVK moieties were detected on some of the cysteine-containing peptides, the results suggest other amino acids may be also reactive with HMVK. Adduct profiles and their relative intensities were consistent between the 1 and 2h samples providing evidence for the HMVK reactions being fast and selective. The finding that fewer peptides were adducted in the 0.1mM HMVK globin samples provides further evidence for selectivity of the HMVK reaction. Collectively, the results show HMVK readily and selectively forms adducts on Hb. Characterization of these adducts will facilitate development of useful biomarkers of exposure to HMVK and its precursor 1,3-butadiene.     

Perception of ultraviolet light by crab spiders and its role in selection of hunting sites

The perception of ultraviolet (UV) light by spiders has so far been only demonstrated in salticids. Crab spiders (Thomisidae) hunt mostly on flowers and need to find appropriate hunting sites. Previous studies have shown that some crab spiders that reflect UV light use UV contrast to enhance prey capture. The high UV contrast can be obtained either by modulation of body colouration or active selection of appropriate backgrounds for foraging. We show that crab spiders (Thomisus sp.) hunting on Spathiphyllum plants use chromatic contrast, especially UV contrast, to make themselves attractive to hymenopteran prey. Apart from that, they are able to achieve high UV contrast by active selection of non-UV reflecting surfaces when given a choice of UV-reflecting and non-UV reflecting surfaces in the absence of odour cues. Honeybees (Apis cerana) approached Spathiphyllum plants bearing crab spiders on which the spiders were high UV-contrast targets with greater frequency than those plants on which the UV contrast of the spiders was low. Thus, crab spiders can perceive UV and may use it to choose appropriate backgrounds to enhance prey capture, by exploiting the attraction of prey such as honeybees to UV.     

Chronic Administration of Lamotrigine Downregulates COX-2 mRNA and Protein in Rat Frontal Cortex

Chronic administration to rats of mood-stabilizers that are effective against mania in bipolar disorder, is reported to downregulate markers of the brain arachidonic acid cascade. We hypothesized that chronic administration of lamotrigine, which is used to treat depression and rapid cycling in bipolar disorder, might do so as well. Male CDF rats were administered a therapeutically relevant dose of lamotrigine (10 mg/kg) or vehicle intragastrically once daily for 42 days. Protein levels of isoforms of phospholipase A₂ (PLA₂) and of cyclooxygenase (COX), and the mRNA level of COX-2, were quantified in the frontal cortex using immunoblotting and RT-PCR, respectively. Compared to vehicle-treated rats, chronic lamotrigine significantly decreased frontal cortex protein and mRNA levels of COX-2 without altering protein levels of the PLA₂ isoforms. Consistent with the hypothesis, lamotrigine and other mood-stabilizers have a common downregulatory action on COX-2 expression in rat brain, which may account in part for their efficacy in bipolar disorder.     

The Xenopus Cell Cycle: An Overview

Oocytes, eggs and embryos from the frog Xenopus laevis have been an important model system for studying cell-cycle regulation for several decades. First, progression through meiosis in the oocyte has been extensively investigated. Oocyte maturation has been shown to involve complex networks of signal transduction pathways, culminating in the cyclic activation and inactivation of Maturation Promoting Factor (MPF), composed of cyclin B and cdc2. After fertilisation, the early embryo undergoes rapid simplified cell cycles which have been recapitulated in cell-free extracts of Xenopus eggs. Experimental manipulation of these extracts has given a wealth of biochemical information about the cell cycle, particularly concerning DNA replication and mitosis. Finally, cells of older embryos adopt a more somatic-type cell cycle and have been used to study the balance between cell cycle and differentiation during development.