High-throughput phenotypic assessment of cardiac physiology in four commonly used inbred mouse strains

Mice with genetic alterations are used in heart research as model systems of human diseases. In the last decade there was a marked increase in the recognition of genetic diversity within inbred mouse strains. Increasing numbers of inbred mouse strains and substrains and analytical variation of cardiac phenotyping methods require reproducible, high-throughput methods to standardize murine cardiovascular physiology. We describe methods for non-invasive, reliable, easy and fast to perform echocardiography and electrocardiography on awake mice. This method can be used for primary screening of the murine cardiovascular system in large-scale analysis. We provide insights into the physiological divergence of C57BL/6N, C57BL/6J, C3HeB/FeJ and 129P2/OlaHsd mouse hearts and define the expected normal values. Our report highlights that compared to the other three strains tested C57BL/6N hearts reveal features of heart failure such as hypertrophy and reduced contractile function. We found several features of the mouse ECG to be under genetic control and obtained several strain-specific differences in cardiac structure and function.     

A Thermostable Salmonella Phage Endolysin,Lys68, with Broad Bactericidal Properties against Gram-Negative Pathogens in Presence of Weak Acids

Resistance rates are increasing among several problematic Gram-negative pathogens, a fact that has encouraged the development of new antimicrobial agents. This paper characterizes a Salmonella phage endolysin (Lys68) and demonstrates its potential antimicrobial effectiveness when combined with organic acids towards Gram-negative pathogens. Biochemical characterization reveals that Lys68 is more active at pH 7.0, maintaining 76.7% of its activity when stored at 4°C for two months. Thermostability tests showed that Lys68 is only completely inactivated upon exposure to 100°C for 30 min, and circular dichroism analysis demonstrated the ability to refold into its original conformation upon thermal denaturation. It was shown that Lys68 is able to lyse a wide panel of Gram-negative bacteria (13 different species) in combination with the outer membrane permeabilizers EDTA, citric and malic acid. While the EDTA/Lys68 combination only inactivated Pseudomonas strains, the use of citric or malic acid broadened Lys68 antibacterial effect to other Gram-negative pathogens (lytic activity against 9 and 11 species, respectively). Particularly against Salmonella Typhimurium LT2, the combinatory effect of malic or citric acid with Lys68 led to approximately 3 to 5 log reductions in bacterial load/CFUs after 2 hours, respectively, and was also able to reduce stationary-phase cells and bacterial biofilms by approximately 1 log. The broad killing capacity of malic/citric acid-Lys68 is explained by the destabilization and major disruptions of the cell outer membrane integrity due to the acidity caused by the organic acids and a relatively high muralytic activity of Lys68 at low pH. Lys68 demonstrates good (thermo)stability properties that combined with different outer membrane permeabilizers, could become useful to combat Gram-negative pathogens in agricultural, food and medical industry.     

Examination of the Combined Effects of Chondroitinase ABC,Growth Factors and Locomotor Training following Compressive Spinal Cord Injury on Neuroanatomical Plasticity and Kinematics

While several cellular and pharmacological treatments have been evaluated following spinal cord injury (SCI) in animal models, it is increasingly recognized that approaches to address the glial scar, including the use of chondroitinase ABC (ChABC), can facilitate neuroanatomical plasticity. Moreover, increasing evidence suggests that combinatorial strategies are key to unlocking the plasticity that is enabled by ChABC. Given this, we evaluated the anatomical and functional consequences of ChABC in a combinatorial approach that also included growth factor (EGF, FGF2 and PDGF-AA) treatments and daily treadmill training on the recovery of hindlimb locomotion in rats with mid thoracic clip compression SCI. Using quantitative neuroanatomical and kinematic assessments, we demonstrate that the combined therapy significantly enhanced the neuroanatomical plasticity of major descending spinal tracts such as corticospinal and serotonergic-spinal pathways. Additionally, the pharmacological treatment attenuated chronic astrogliosis and inflammation at and adjacent to the lesion with the modest synergistic effects of treadmill training. We also observed a trend for earlier recovery of locomotion accompanied by an improvement of the overall angular excursions in rats treated with ChABC and growth factors in the first 4 weeks after SCI. At the end of the 7-week recovery period, rats from all groups exhibited an impressive spontaneous recovery of the kinematic parameters during locomotion on treadmill. However, although the combinatorial treatment led to clear chronic neuroanatomical plasticity, these structural changes did not translate to an additional long-term improvement of locomotor parameters studied including hindlimb-forelimb coupling. These findings demonstrate the beneficial effects of combined ChABC, growth factors and locomotor training on the plasticity of the injured spinal cord and the potential to induce earlier neurobehavioral recovery. However, additional approaches such as stem cell therapies or a more adapted treadmill training protocol may be required to optimize this repair strategy in order to induce sustained functional locomotor improvement.     

Effects of Exogenous Recombinant APC in Mouse Models of Ischemia Reperfusion Injury and of Atherosclerosis

Activated protein C (APC) is a serine protease that has both anticoagulant and cytoprotective properties. The cytoprotective effects are protease activated receptor 1 (PAR-1) and endothelial protein C receptor (EPCR) dependent and likely underlie protective effects of APC in animal models of sepsis, myocardial infarction and ischemic stroke. S360A-(A)PC, a variant (A)PC that has no catalytic activity, binds EPCR and shifts pro-inflammatory signaling of the thrombin-PAR-1 complex to anti-inflammatory signaling. In this study we investigated effects of human (h)wt-PC, hS360A-PC, hwt-APC and hS360A-APC in acute (mouse model of acute myocardial ischemia/reperfusion (I/R) injury) and chronic inflammation (apoE^−/− mouse model of atherosclerosis). All h(A)PC variants significantly reduced myocardial infarct area (p<0.05) following I/R injury. IL-6 levels in heart homogenates did not differ significantly between sham, placebo and treatment groups in I/R injury. None of the h(A)PC variants decreased number and size of atherosclerotic plaques in apoE^−/− mice. Only hS360A-APC slightly affected phenotype of plaques. IL-6 levels in plasma were significantly (p<0.001) decreased in hwt-APC and hS360A-PC treated mice. In the last group levels of monocyte chemotactic protein 1 (MCP-1) were significantly increased (p<0.05). In this study we show that both hwt and hS360A-(A)PC protect against acute myocardial I/R injury, which implies that protection from I/R injury is independent of the proteolytic activity of APC. However, in the chronic atherosclerosis model hwt and hS360-(A)PC had only minor effects. When the dose, species and mode of (A)PC administration will be adjusted, we believe that (A)PC will have potential to influence development of chronic inflammation as occurring during atherosclerosis as well.     

Fatty acid composition and biological activities of Isochrysis galbana T-ISO,Tetraselmis sp. and Scenedesmus sp.: possible application in the pharmaceutical and functional food industries

Organic and water extracts of Isochrysis galbana T-ISO (=Tisochrysis lutea), Tetraselmis sp. and Scenedesmus sp. were evaluated for their antioxidant activity, acetylcholinesterase (AChE) inhibition, cytotoxicity against tumour cell lines, and fatty acids and total phenolic content (TPC). I. galbana T-ISO had the highest TPC (3.18 mg GAE g^?1) and radical scavenging activity, with an IC₅₀ value of 1.9 mg mL^?1 on the acetone extract. The extracts exhibited a higher ability to chelate Fe²⁺ than Cu²⁺, and the maximum Fe²⁺ chelating capacity was observed in the hexane extract of Scenedesmus sp. (IC₅₀=0.73 mg mL^?1) and Scenedesmus sp. (IC₅₀?=?0.73 mg mL^?1). The highest ability to inhibit AChE was observed in the water and ether extracts of Scenedesmus sp., with IC₅₀ values of 0.11 and 0.15 mg mL^?1, respectively, and in the water extract of I. galbana (IC₅₀?=?0.16 mg mL^?1). The acetone extract of I. galbana T-ISO significantly reduced the viability of human hepatic carcinoma HepG2 cells (IC₅₀?=?81.3 μg mL^?1) as compared to the non-tumour murine stromal S17 cell line, and displayed a selectivity index of 3.1 at the highest concentration tested (125 μg mL^?1). All species presented a highly unsaturated fatty acids profile. Results suggest that these microalgae, particularly I. galbana T-ISO, could be a source of biomolecules for the pharmaceutical industry and the production of functional food ingredients and can be considered as an advantageous alternative to several currently produced microalgae.     

Differences in nutritional status between rural and urban Yucatec Maya children: The importance of early life conditions

Early-life conditions shape childhood growth and are affected by urbanization and the nutritional transition. To investigate how early-life conditions (across the “first” and “second” 1000 days) are associated with rural and urban children's nutritional status, we analyzed anthropometric data from Maya children in Yucatan, Mexico. We collected weight, height and triceps skinfold measures, then computed body mass and fat mass indices (BMI/FMI), in a cross-sectional sample of 6-year-olds (urban n = 72, rural n = 66). Demographic, socioeconomic and early-life variables (birthweight/mode, rural/urban residence, household crowding) were collected by maternal interview. We statistically analyzed rural-urban differences in demographic, socioeconomic, early-life, and anthropometric variables, then created linear mixed models to evaluate associations between early-life variables and child anthropometric outcomes. Two-way interactions were tested between early-life variables and child sex, and between early-life variables and rural-urban residence. Results showed that rural children were shorter-statured, with lower overweight/obesity and cesarean delivery rates, compared to urban children. Household crowding was a negative predictor of anthropometric outcomes; the strongest effect was in boys and in urban children. Birthweight positively predicted anthropometric outcomes, especially weight/BMI. Birth mode was positively (not statistically) associated with any anthropometric outcome. Cesarean delivery was more common in boys than in girls, and predicted increased height in urban boys. In conclusion, urbanization and household crowding were the most powerful predictors of Maya 6-year-old anthropometry. The negative effects of crowding may disproportionately affect Maya boys versus girls and urban versus rural children. Early-life conditions shape Maya children's nutritional status both in the “first” and “second” 1000 days.