Potentiation of arginine-induced glucagon secretion by adenosine

Adenosine and the synthetic adenosine agonists 2-chloroadenosine and N⁶-(L-2-phenylisopropyl)-adenosine were tested for effects on hormone secretion from the rat isolated perfused pancreas. These nucleosides, at concentrations of 5 μM, markedly potentiated both phases of arginine-induced glucagon release; the two synthetic agonists were more effective than adenosine. In the absence of arginine, each of the nucleosides induced a transient burst of glucagon. In contrast, adenosine and both synthetic agonists inhibited arginine-induced insulin secretion to varying degrees and caused only negligible insulin release when perfused without arginine. The adenosine antagonist 8-($\text{p}$-sulfophenyl)-theophylline prevented the actions of adenosine on hormone release from the pancreas. Our data suggest that adenosine potentiation of arginine-induced glucagon release may be mediated via adenosine receptors on alpha cell membranes; such a mechanism could provide an important endogenous control over glucagon secretion.     

Tumor promoters and epidermal growth factor stimulate anchorage-independent growth of adenovirus-tranformed rat embryo cells.

Previous studies indicated that the potent tumor promoter 12--0--tetradecanoyl-phorbol-13-acetate (TPA) enhances transformation of rat embryo cells (2 degrees RE) by a mutant of human Ad5 (H5ts125). This study examines the effect of TPA, its structural analogs and epidermal growth factor (EGF) on anchorage-independent growth of a cloned population of H5ts125-transformed 2 degrees RE cells (clone E11). Both TPA and EGF (approximately 10(-8) M) induced a 3--5 fold increase in agar cloning efficiency of E11 cells. In addition, macroscopic colonies appeared earlier and were larger and more diffuse. The TPA analogs phorbol--12,13--didecanoate (PDD) and ingenol--3,20--dibenzoate also enhanced growth in agar of E11 cells, whereas phorbol, 4 alpha PDD and 4--0--meTPA, which are inactive as tumor promoters, failed to enhance agar growth. In contrast to the results obtained with E11 cells, TPA, PDD or ingenol--3,20--bidenzoate failed to induce growth in agar of normal 2 degrees RE cells. Dexamethasone (10(-5)--10(-6) M), trans retinoic acid (10(-5)--10(-6) M) and the protease inhibitors leupeptin, antipain and elastatinol did not inhibit the ability of TPA to enhance the growth of E11 cells in agar. The TPA-enhanced anchorage independence was a stable property, since subclones of E11 cells isolated from TPA-agar plates had a higher agar cloning efficiency than the parental E11 cells when retested in the absence of TPA. This effect of TPA does not appear to reflect simple selection of a subpopulation of cells. When the parental E11 cells were first cloned in monolayer culture in the absence of TPA, all ten randomly picked clones showed enhanced growth in agar in the presence of TPA. In addition, prior growth of E11 cells in monolayer culture in the presence of TPA did not enhance their subsequent growth in agar. This system therefore provides an example in which TPA appears to enhance the acquisition of a stable cell property, and thus may be a useful model for studying mechanisms of tumor promotion and progression.     

A scanning electron microscopy study of micro-arterial damage and repair.

The effects of microvascular clamps on the femoral vessels of rats were studied, using the SEM. The early changes observed were (1) local fusiform dilatation of the area secondary to necrosis of the muscular wall, (2) flattening of the longitudinal ridges in the endothelial (3) loss of laminar flow, (4) endothelial sloughing, (5) platelet aggregation, and (6) leukocyte adherence and diapedesis. The repair of the endothelium occurred by an early replication of the adjacent undamaged endothelial cells -- with their subsequent migration across a platelet bed. The coverage was complete in one week, although reorientation of the neo-endothelial cells took longer. On the basis of this study and our clinical experience, we think the ideal microvascular clamp would possess the following characteristics: small size, light weight, mechanical simplicity, flat jaws (one to two mm in diameter) coated with a non-slip surface, and calibrated to produce a pressure less than 30 gm per mm2. In addition the clamp should be unaffected by blood, autoclaving, or repeated use. No such clamp is commercially available now, but we hope that one will be available in the near future.     

Host diversity and behavior determine patterns of interspecies transmission and geographic diffusion of avian influenza A subtypes among North American wild reservoir species

Wild birds can carry avian influenza viruses (AIV), including those with pandemic or panzootic potential, long distances. Even though AIV has a broad host range, few studies account for host diversity when estimating AIV spread. We analyzed AIV genomic sequences from North American wild birds, including 303 newly sequenced isolates, to estimate interspecies and geographic viral transition patterns among multiple co-circulating subtypes. Our results show high transition rates within Anseriformes and Charadriiformes, but limited transitions between these orders. Patterns of transition between species were positively associated with breeding habitat range overlap, and negatively associated with host genetic distance. Distance between regions (negative correlation) and summer temperature at origin (positive correlation) were strong predictors of transition between locations. Taken together, this study demonstrates that host diversity and ecology can determine evolutionary processes that underlie AIV natural history and spread. Understanding these processes can provide important insights for effective control of AIV.     

The Giardia ventrolateral flange is a lamellar membrane protrusion that supports attachment

Attachment to the intestinal epithelium is critical to the lifestyle of the ubiquitous parasite Giardia lamblia. The ventrolateral flange is a sheet-like membrane protrusion at the interface between parasites and attached surfaces. This structure has been implicated in attachment, but its role has been poorly defined. Here, we identified a novel actin associated protein with putative WH2-like actin binding domains we named Flangin. Flangin complexes with Giardia actin (GlActin) and is enriched in the ventrolateral flange making it a valuable marker for studying the flanges’ role in Giardia biology. Live imaging revealed that the flange grows to around 1 μm in width after cytokinesis, then remains uniform in size during interphase, grows in mitosis, and is resorbed during cytokinesis. A flangin truncation mutant stabilizes the flange and blocks cytokinesis, indicating that flange disassembly is necessary for rapid myosin-independent cytokinesis in Giardia. Rho family GTPases are important regulators of membrane protrusions and GlRac, the sole Rho family GTPase in Giardia, was localized to the flange. Knockdown of Flangin, GlActin, and GlRac result in flange formation defects. This indicates a conserved role for GlRac and GlActin in forming membrane protrusions, despite the absence of canonical actin binding proteins that link Rho GTPase signaling to lamellipodia formation. Flangin-depleted parasites had reduced surface contact and when challenged with fluid shear force in flow chambers they had a reduced ability to remain attached, confirming a role for the flange in attachment. This secondary attachment mechanism complements the microtubule based adhesive ventral disc, a feature that may be particularly important during mitosis when the parental ventral disc disassembles in preparation for cytokinesis. This work supports the emerging view that Giardia’s unconventional actin cytoskeleton has an important role in supporting parasite attachment.     

Gene expression changes reflect clinical response in a placebo-controlled randomized trial of abatacept in patients with diffuse cutaneous systemic sclerosis

IntroductionSystemic sclerosis is an autoimmune disease characterized by inflammation and fibrosis of the skin and internal organs. We sought to assess the clinical and molecular effects associated with response to intravenous abatacept in patients with diffuse cutaneous systemic.MethodsAdult diffuse cutaneous systemic sclerosis patients were randomized in a 2:1 double-blinded fashion to receive abatacept or placebo over 24 weeks. Primary outcomes were safety and the change in modified Rodnan Skin Score (mRSS) at week 24 compared with baseline. Improvers were defined as patients with a decrease in mRSS of ≥30 % post-treatment compared to baseline. Skin biopsies were obtained for differential gene expression and pathway enrichment analyses and intrinsic gene expression subset assignment.ResultsTen subjects were randomized to abatacept (n = 7) or placebo (n = 3). Disease duration from first non-Raynaud’s symptom was significantly longer (8.8 ± 3.8 years vs. 2.4 ± 1.6 years, p = 0.004) and median mRSS was higher (30 vs. 22, p = 0.05) in the placebo compared to abatacept group. Adverse events were similar in the two groups. Five out of seven patients (71 %) randomized to abatacept and one out of three patients (33 %) randomized to placebo experienced ≥30 % improvement in skin score. Subjects receiving abatacept showed a trend toward improvement in mRSS at week 24 (−8.6 ± 7.5, p = 0.0625) while those in the placebo group did not (−2.3 ± 15, p = 0.75). After adjusting for disease duration, mRSS significantly improved in the abatacept compared with the placebo group (abatacept vs. placebo mRSS decrease estimate −9.8, 95 % confidence interval −16.7 to −3.0, p = 0.0114). In the abatacept group, the patients in the inflammatory intrinsic subset showed a trend toward greater improvement in skin score at 24 weeks compared with the patients in the normal-like intrinsic subset (−13.5 ± 3.1 vs. −4.5 ± 6.4, p = 0.067). Abatacept resulted in decreased CD28 co-stimulatory gene expression in improvers consistent with its mechanism of action. Improvers mapped to the inflammatory intrinsic subset and showed decreased gene expression in inflammatory pathways, while non-improver and placebos showed stable or reverse gene expression over 24 weeks.ConclusionsClinical improvement following abatacept therapy was associated with modulation of inflammatory pathways in skin.

Trial registration

ClinicalTrials.gov {"type":"clinical-trial","attrs":{"text":"NCT00442611","term_id":"NCT00442611"}}NCT00442611. Registered 1 March 2007.

Electronic supplementary material

The online version of this article (doi:10.1186/s13075-015-0669-3) contains supplementary material, which is available to authorized users.     

Inequality as a Powerful Predictor of Infant and Maternal Mortality around the World

Background

Maternal and infant mortality are highly devastating, yet, in many cases, preventable events for a community. The human development of a country is a strong predictor of maternal and infant mortality, reflecting the importance of socioeconomic factors in determinants of health. Previous research has shown that the Human Development Index (HDI) predicts infant mortality rate (IMR) and the maternal mortality ratio (MMR). Inequality has also been shown to be associated with worse health in certain populations. The main purpose of the present study was to determine the correlation and predictive power of the Inequality Adjusted Human Development Index (IHDI) as a measure of inequality with the Infant Mortality Rate (IMR), Maternal Mortality Rate (MMR), Early Neonatal Mortality Rate (ENMR), Late Neonatal Mortality Rate (LNMR), and the Post Neonatal Mortality Rate (PNMR).

Methods and Findings

Data for the present study were downloaded from two sources: infant and maternal mortality data were downloaded from the Global Burden of Disease 2013 Cause of Death Database and the Human Development Index (HDI) and Inequality-Adjusted Human Development Index (IHDI) data were downloaded from the United Nations Development Program (UNDP). Pearson correlation coefficients were estimated, following logarithmic transformations to the data, to examine the relationship between HDI and IHDI with MMR, IMR, ENMR, LNMR, and PNMR. Steiger’s Z test for the equality of two dependent correlations was utilized in order to determine whether the HDI or IHDI was more strongly associated with the outcome variables. Lastly, we constructed OLS regression models in order to determine the predictive power of the HDI and IHDI in terms of the MMR, IMR, ENMR, LNMR, and PNMR.Maternal and infant mortality were both strongly and negatively correlated with both HDI and IHDI; however, Steiger’s Z test for the equality of two dependent correlations revealed that IHDI was more strongly correlated than HDI with MMR (Z = 4.897, p < 0.001), IMR (Z = 2.524, p = 0.012), ENMR (Z = 2.936, p = 0.003), LNMR (Z = 2.272, p = 0.023), and PNMR (Z = 2.277, p = 0.023). Furthermore, side-by-side OLS regression models revealed that, when IHDI was used as the predictor variable instead of HDI, the R ² value was 0.053 higher for MMR, 0.025 higher for IMR, 0.038 higher for ENMR, 0.029 higher for LNMR, and 0.026 higher for PNMR.

Conclusions

Even when both the HDI and the IHDI correlate with the infant and maternal mortality rates, the IHDI is a better predictor for these two health indicators. Therefore, these results add more evidence that inequality is playing an important role in determining the health status of various populations in the world and more efforts should be put into programs to fight inequality.     

Characterization of modeled inhibitory binding sites on isoform one of the Na+/H+ exchanger

Mammalian Na⁺/H⁺ exchanger isoform one (NHE1) is a plasma membrane protein responsible for pH regulation in mammalian cells. Excess activity of the protein promotes heart disease and is a trigger of metastasis in cancer. Inhibitors of the protein exist but problems in specificity have delayed their clinical application. Here we examined amino acids involved in two modeled inhibitor binding sites (A, B) in human NHE1. Twelve mutations (Asp159, Phe348, Ser351, Tyr381, Phe413, Leu465, Gly466, Tyr467, Leu468, His473, Met476, Leu481) were made and characterized. Mutants S351A, F413A, Y467A, L468A, M476A and L481A had 40–70% of wild type expression levels, while G466A and H473A expressed 22% ~ 30% of the wild type levels. Most mutants, were targeted to the cell surface at levels similar to wild type NHE1, approximately 50–70%, except for F413A and G466A, which had very low surface targeting. Most of the mutants had measurable activity except for D159A, F413A and G466A. Resistance to inhibition by EMD87580 was elevated in mutants F438A, L465A and L468A and reduced in mutants S351A, Y381A, H473A, M476A and L481A. All mutants with large alterations in inhibitory properties showed reduced Na⁺ affinity. The greatest changes in activity and inhibitor sensitivity were in mutants present in binding site B which is more closely associated with TM4 and C terminal of extracellular loop 5, and is situated between the putative scaffolding domain and transport domain. The results help define the inhibitor binding domain of the NHE1 protein and identify new amino acids involved in inhibitor binding.