A rapid response microviscosimeter

An in vitro instrument is described which is designed to measure effective viscosity of blood in arteriolar size tubes at physiologically nominal flow rates, mimicking flow in the microcirculation. The 41-micron microviscosimeter is accurate within 2% when tested against viscosity standards and is reproducible within 2% using blood samples. Because the full-scale instrument response time is 3 s, either fresh or anticoagulated blood samples may be used. Measured over the nominal range of blood flow rate (Q), effective blood viscosity was found to be an increasing, decreasing, or flat function of Q, depending upon the particular individual being tested. A reference group of 81 young, healthy subjects was used to define viscous resistance (VR), a new parameter that provides for quantitative viscosity comparisons between individuals or groups without hematocrit manipulation of blood samples. As examples of the microviscosimeter's use, a group of 118 subjects was used to test for VR variation between various group subsets. No difference in VR was found between men and women; exercisers had lower VR than nonexercisers; and overweight subjects had more viscous blood than non-overweight subjects. The instrument will be useful for in vitro investigations of effective viscosity and viscous resistance in the microcirculation.     

Efficiency and specificity of gene isolation by exon amplification

Exon amplification is an increasingly popular approach to the identification of transcribed sequences and will complement other strategies to isolate genes. We have used this system to amplify candidate exons from 32 cosmids, including 8 cosmids which span a well characterized 185-kb region of the human major histocompatibility class II region on Chromosome (Chr) 6. We have examined the efficiency, specificity, and reproducibility of the system in isolating exons from genes known to be present on particular cosmids and have determined the nature and frequency of artefact amplifications in routine cosmid screening. We were able to clone at least one exon from 88% (7/8) of all known genes tested (including exons which are differentially spliced) and obtained artefacts from 19% (6/32) of the cosmids tested. Such artefacts generally arise from the amplification of noncoding sequences flanked by regions with high homology to acceptor and donor splice junctions. We show that the exon amplification procedure can be used successfully with a wide variety of cosmids which have different numbers of genes and gene structures and describe several approaches to the characterization of novel exons cloned in this study.     

Effect of a thromboxane receptor antagonist on PGD2- and allergen-induced bronchoconstriction

In this study we investigated the effect of the selective and potent thromboxane A2 (TxA2) receptor antagonist GR32191 on smooth muscle contraction induced by the TxA2 analogue U46619, prostaglandin (PG) D2, PGF2 alpha, and methacholine (MCh) in guinea pig airways in vitro and the airways response provoked by inhaled PGD2 and MCh in asthmatic subjects in vivo. GR32191 antagonized competitively the contractile responses of all three prostanoids to a similar degree but had no effect on MCh-induced contractions. In asthmatic subjects GR32191, in a single oral dose of 80 mg, did not affect base-line airway caliber or MCh-induced broncho-constriction but caused significant inhibition of PGD2-induced bronchoconstriction, displacing the concentration-response curves to the right by greater than 10-fold. The effect of the same oral dose of GR32191 on allergen-induced immediate bronchoconstriction was subsequently investigated in allergic asthmatic subjects. In individual subjects, GR32191 inhibited to varying degrees the overall bronchoconstrictor response, with the maximum effect occurring between 10 and 30 min after allergen challenge. These studies suggest that prostanoids contribute to the immediate bronchoconstriction induced by inhaled allergen in allergic asthmatics, and that this effect is mediated by stimulation of a thromboxane receptor.     

Aldehydes, hydrogen peroxide, and organic radicals as mediators of ozone toxicity

It is generally agreed that unsaturated fatty acids (UFA) are an important class of target molecule for reaction with ozone when polluted air is inhaled. Most discussions have implicated the UFA in cell membranes, but lung lining fluids also contain fatty acids that are from 20 to 40% unsaturated. Since UFA in lung lining fluids exist in a highly aquated environment, ozonation would be expected to produce aldehydes and hydrogen peroxide, rather than the Criegee ozonide. In agreement with this expectation, we find that ozonations of emulsions of fatty acids containing from one to four double bonds give one mole of H2O2 for each mole of ozone reacted. Ozonation of oleic acid emulsions and dioleoyl phosphatidyl choline gives similar results. with two moles of aldehydes and one mole of H2O2 formed per mole of ozone reacted. The net reaction that occurs when ozone reacts with pulmonary lipids is suggested to be given by equation 1. [formula: see text]. From 5 to 10% yields of Criegee ozonides also appear to be formed. In addition, a direct reaction of unknown mechanism occurs between ozone and UFA in homogeneous organic solution, in homogeneous solutions in water, in aqueous emulsions, and in lipid bilayers to give organic radicals that can be spin trapped. These radicals are suggested to be responsible for initiating lipid peroxidation of polyunsaturated fatty acids. Thus, aldehydes, hydrogen peroxide, and directly produced organic radicals are suggested to be mediators of ozone-induced pathology.     

FTO demethylase activity is essential for normal bone growth and bone mineralization in mice

The Fto gene locus has been linked to increased body weight and obesity in human population studies, but the role of the actual FTO protein in adiposity has remained controversial. Complete loss of FTO protein in mouse and of FTO function in human patients has multiple and variable effects. To determine which effects are due to the ability of FTO to demethylate mRNA, we genetically engineered a mouse with a catalytically inactive form of FTO. Our results demonstrate that FTO catalytic activity is not required for normal body composition although it is required for normal body size and viability. Strikingly, it is also essential for normal bone growth and mineralization, a previously unreported FTO function.     

A Change in Orientation: Recognition of Rotated Patterns by Bumble Bees

In three experiments, bumble bees were trained to discriminate between a reinforcing pattern (S+) and a nonreinforcing one (S–) which differed only in the configuration of four artificial petals. They were subsequently tested for recognition of the S+ rotated by 90° (S + 90). Experiment 1 used petals of four colors, and the other experiments used four symbols. The symbols either remained unchanged when the whole pattern was rotated (e.g., + in Experiment 2) or changed appearance (e.g., < in Experiment 3). The bees failed to recognize the S + 90 in the first two experiments, but in Experiment 3, the choice proportion for S + 90 in the presence of a New pattern was significantly higher than chance. Bumble bees can recognize a rotated pattern, possibly by using mental rotation, provided that a cue as to the extent of the pattern transformation is given.     

Determinants of the Changes in Glycemic Control with Exercise Training in Type 2 Diabetes: A Randomized Trial

Aims

To assess the determinants of exercise training-induced improvements in glucose control (HbA_1C) including changes in serum total adiponectin and FFA concentrations, and skeletal muscle peroxisome proliferator-activated receptor-γ coactivator-1α (PGC-1α) protein content.

Methods

A sub-cohort (n = 35; 48% men; 74% Caucasian) from the HART-D study undertaking muscle biopsies before and after 9 months of aerobic (AT), resistance (RT), or combination training (ATRT).

Results

Changes in HbA_1C were associated with changes in adiponectin (r = −0.45, P = 0.007). Participants diagnosed with type 2 diabetes for a longer duration had the largest increase in PGC-1α (r = 0.44, P = 0.008). Statistical modeling examining changes in HbA_1C suggested that male sex (P = 0.05), non-Caucasian ethnicity (P = 0.02), duration of type 2 diabetes (r = 0.40; P<0.002) and changes in FFA (r = 0.36; P<0.004), adiponectin (r = −0.26; P<0.03), and PGC-1α (r = −0.28; P = 0.02) explain ∼65% of the variability in the changes in HbA_1C.

Conclusions

Decreases in HbA_1C after 9 months of exercise were associated with shorter duration of diabetes, lowering of serum FFA concentrations, increasing serum adiponectin concentrations and increasing skeletal muscle PGC-1α protein expression.

Trial Registration

ClinicalTrials.gov {"type":"clinical-trial","attrs":{"text":"NCT00458133","term_id":"NCT00458133"}}NCT00458133     

The evolution of large‐scale body size clines in Plethodon salamanders: evidence of heat‐balance or species‐specific artifact?

A major goal in macroecology is to determine how body size varies geographically, and explain why such patterns exist. Recently, a grid‐cell assemblage analysis found significant body size trends with latitude and temperature in Plethodon salamanders, and support for the heat‐balance hypothesis as a possible explanation for these trends. Here we demonstrate that the heat‐balance hypothesis is unlikely to have generated this pattern, and that there is no overall body size trend with temperature in Plethodon. Using data from 3155 local Plethodon assemblages, we find no support for body size clines with latitude, and no relationship between body size and temperature. We also found that body size did not covary with elevation, in contrast to what was predicted by heat‐balance. We then examined the various scenarios under which body size clines across grid‐cell assemblages could evolve via heat‐balance, and found that none were tenable in light of the existing data. Instead, a single, widely distributed species was responsible for the pattern across grid‐cell assemblages. Finally, we examined why phylogenetic eigenvector regression does not account for phylogenetic non‐independence among taxa, and should not be used to account for shared evolutionary history in assembly‐level analyses. Assemblage‐level patterns are a useful means of assessing biogeographic trends, and are an important complement to within‐species and cross‐species patterns. However, while the use of grid‐cell assemblage approaches from digital databases is expedient, their results must be examined critically, and whenever possible, compared with data obtained from local species assemblages (particularly for ecological mechanisms that operate at the level of individuals). Finally, our results emphasize the importance of using corroborative data to evaluate alternative hypotheses, so that potential mechanisms that explain bioegeographic patterns are properly assigned.     

Gastrointestinal Viral Load and Enteroendocrine Cell Number Are Associated with Altered Survival in HIV-1 Infected Individuals

Human immunodeficiency virus type 1 (HIV-1) infects and destroys cells of the immune system leading to an overt immune deficiency known as HIV acquired immunodeficiency syndrome (HIV/AIDS). The gut associated lymphoid tissue is one of the major lymphoid tissues targeted by HIV-1, and is considered a reservoir for HIV-1 replication and of major importance in CD4+ T-cell depletion. In addition to immunodeficiency, HIV-1 infection also directly causes gastrointestinal (GI) dysfunction, also known as HIV enteropathy. This enteropathy can manifest itself as many pathological changes in the GI tract. The objective of this study was to determine the association of gut HIV-1 infection markers with long-term survival in a cohort of men who have sex with men (MSM) enrolled pre-HAART (Highly Active Antiretroviral Therapy). We examined survival over 15-years in a cohort of 42 HIV-infected cases: In addition to CD4+ T cell counts and HIV-1 plasma viral load, multiple gut compartment (duodenum and colon) biopsies were taken by endoscopy every 6 months during the initial 3-year period. HIV-1 was cultured from tissues and phenotyped and viral loads in the gut tissues were determined. Moreover, the tissues were subjected to an extensive assessment of enteroendocrine cell distribution and pathology. The collected data was used for survival analyses, which showed that patients with higher gut tissue viral load levels had a significantly worse survival prognosis. Moreover, lower numbers of serotonin (duodenum) and somatostatin (duodenum and colon) immunoreactive cell counts in the gut tissues of patients was associated with significant lower survival prognosis. Our study, suggested that HIV-1 pathogenesis and survival prognosis is associated with altered enteroendocrine cell numbers, which could point to a potential role for enteroendocrine function in HIV infection and pathogenesis.