Influence of substrate supply on cardiac efficiency, as measured by pressure-volume analysis in ex vivo mouse hearts

In the present study, we tested the reliability of measurements of pressure-volume area (PVA) and oxygen consumption (MVo(2)) in ex vivo mouse hearts, combining the use of a miniaturized conductance catheter and a fiber-optic oxygen sensor. Second, we tested whether we could reproduce the influence of increased myocardial fatty acid (FA) metabolism on cardiac efficiency in the isolated working mouse heart model, which has already been documented in large animal models. The hearts were perfused with crystalloid buffer containing 11 mM glucose and two different concentrations of FA bound to 3% BSA. The initial concentration was 0.3 +/- 0.1 mM, which was subsequently raised to 0.9 +/- 0.1 mM. End-systolic and end-diastolic pressure-volume relationships were assessed by temporarily occluding the preload line. Different steady-state PVA-MVo(2) relationships were obtained by changing the loading conditions (pre- and afterload) of the heart. There were no apparent changes in baseline cardiac performance or contractile efficiency (slope of the PVA-MVo(2) regression line) in response to the elevation of the perfusate FA concentration. However, all hearts (n = 8) showed an increase in the y-intercept of the PVA-MVo(2) regression line after elevation of the palmitate concentration, indicating an FA-induced increase in the unloaded MVo(2). Therefore, in the present model, unloaded MVo(2) is not independent of metabolic substrate. This is, to our knowledge, the first report of a PVA-MVo(2) relationship in ex vivo perfused murine hearts, using a pressure-volume catheter. The methodology can be an important tool for phenotypic assessment of the relationship among metabolism, contractile performance, and cardiac efficiency in various mouse models.     

Essential amino acids of the hantaan virus N protein in its interaction with RNA

The nucleocapsid (N) protein of hantavirus encapsidates viral genomic and antigenomic RNAs. Previously, deletion mapping identified a central, conserved region (amino acids 175 to 217) within the Hantaan virus (HTNV) N protein that interacts with a high affinity with these viral RNAs (vRNAs). To further define the boundaries of the RNA binding domain (RBD), several peptides were synthesized and examined for the ability to bind full-length S-segment vRNA. Peptide 195-217 retained 94% of the vRNA bound by the HTNV N protein, while peptides 175-186 and 205-217 bound only 1% of the vRNA. To further explore which residues were essential for binding vRNA, we performed a comprehensive mutational analysis of the amino acids in the RBD. Single and double Ala substitutions were constructed for 18 amino acids from amino acids 175 to 217 in the full-length N protein. In addition, Ala substitutions were made for the three R residues in peptide 185-217. An analysis of protein-RNA interactions by electrophoretic mobility shift assays implicated E192, Y206, and S217 as important for binding. Chemical modification experiments showed that lysine residues, but not arginine or cysteine residues, contribute to RNA binding, which agreed with bioinformatic predictions. Overall, these data implicate lysine residues dispersed from amino acids 175 to 429 of the protein and three amino acids located in the RBD as essential for RNA binding.     

Quantitative genetics of vector competence for La Crosse virus and body size in Ochlerotatus hendersoni and Ochlerotatus triseriatus interspecific hybrids

La Crosse virus is a leading cause of pediatric encephalitis in the United States. The mosquito Ochlerotatus triseriatus is an efficient vector for La Crosse virus, whereas the closely related O. hendersoni transmits only at very low rates. Quantitative trait loci (QTL) affecting the ability to orally transmit this virus and adult body size were identified in 164 F(2) female individuals from interspecific crosses of O. hendersoni females and O. triseriatus males using a combination of composite interval mapping (CIM), interval mapping (IM) for binary traits, and single-marker mapping. For oral transmission (OT), no genome locations exceeded the 95% experimentwise threshold for declaring a QTL using IM, but single-marker analysis identified four independent regions significantly associated with OT that we considered as tentative QTL. With two QTL, an increase in OT was associated with alleles from the refractory vector, O. hendersoni, and likely reflect epistatic interactions between genes that were uncovered by our interspecific crosses. For body size, two QTL were identified using CIM and a third tentative QTL was identified using single-marker analysis. The genome regions associated with body size also contain three QTL controlling OT, suggesting that these regions contain either single genes with pleiotropic effects or multiple linked genes independently determining each trait.     

Cyclin D1 splice variant and risk for non-Hodgkin lymphoma

To investigate the role of cell cycle gene variations in lymphomagenesis, we evaluated associations (odds ratios [OR] and 95% confidence intervals [CI]) in polymorphisms from seven candidate genes in 1,172 non-Hodgkin lymphoma (NHL) cases and 982 population-based controls. The cyclin D1 (CCND1) splice variant G870A (rs603965) increased NHL risk (OR_AA = 1.4, 95% CI = 1.1–1.8, P-trend = 0.021), which was consistent for four B-cell subtypes. As CCND1 expression indicates poor NHL prognosis, our results, if true, would support its potentially dual importance in NHL etiology and survival.Electronic Supplementary Material Supplementary material is available to authorised users in the online version of this article at .     

The distribution and evolutionary history of the PRP8 intein

Background  

We recently described a mini-intein in the PRP8 gene of a strain of the basidiomycete Cryptococcus neoformans, an important fungal pathogen of humans. This was the second described intein in the nuclear genome of any eukaryote; the first nuclear encoded intein was found in the VMA gene of several saccharomycete yeasts. The evolution of eukaryote inteins is not well understood. In this report we describe additional PRP8 inteins (bringing the total of these to over 20). We compare and contrast the phylogenetic distribution and evolutionary history of the PRP8 intein and the saccharomycete VMA intein, in order to derive a broader understanding of eukaryote intein evolution. It has been suggested that eukaryote inteins undergo horizontal transfer and the present analysis explores this proposal.     

RNAi knock-downs support roles for the mucin-like (AeIMUC1) gene and short-chain dehydrogenase/reductase (SDR) gene in Aedes aegypti susceptibility to Plasmodium gallinaceum

The mosquito midgut represents the first barrier encountered by the Plasmodium parasite (Haemosporida: Plasmodiidae) when it is ingested in blood from an infected vertebrate. Previous studies identified the Aedes aegypti (L.) (Diptera: Culicidae) mucin-like (AeIMUC1) and short-chain dehydrogenase/reductase (SDR) genes as midgut-expressed candidate genes influencing susceptibility to infection by Plasmodium gallinaceum (Brumpt). We used RNA inference (RNAi) by double-stranded RNA (dsRNA) injections to examine ookinete survival to the oocyst stage following individual gene knock-downs. Double-stranded RNA gene knock-downs were performed 3 days prior to P. gallinaceum infection and oocyst development was evaluated at 7 days post-infection. Mean numbers of parasites developing to the oocyst stage were significantly reduced by 52.3% in dsAeIMUC1-injected females and by 36.5% in dsSDR-injected females compared with females injected with a dsβ-gal control. The prevalence of infection was significantly reduced in dsAeIMUC1- and dsSDR-injected females compared with females injected with dsβ-gal; these reductions resulted in a two- and three-fold increase in the number of uninfected individuals, respectively. Overall, these results suggest that both AeIMUC1 and SDR play a role in Ae. aegypti vector competence to P. gallinaceum.