Burnt sugarcane harvesting: Particulate matter exposure and the effects on lung function, oxidative stress, and urinary 1-hydroxypyrene

Non-mechanised sugarcane harvesting preceded by burning exposes workers and the people of neighbouring towns to high concentrations of pollutants. This study was aimed to evaluate the respiratory symptoms, lung function and oxidative stress markers in sugarcane workers and the residents of Mendonca, an agricultural town in Brazil, during the non-harvesting and harvesting periods and to assess the population and individual exposures to fine particulate matter (PM2.5). Sugarcane workers and healthy volunteers were evaluated with two respiratory symptom questionnaires, spirometry, urinary 1-hydroxypyrene levels, and the measurement of antioxidant enzymes and plasma malonaldehyde during the non-harvesting and harvesting periods. The environmental assessment was determined from PM2.5 concentration. PM2.5 level increased from 8 mu g/m(3) during the non-harvesting period to 23.5 mu g/m(3) in the town and 61 mu g/m(3) on the plantations during the harvesting period. Wheezing, coughing, sneezing, and breathlessness increased significantly in both groups during the harvesting period, but more markedly in workers. A decrease in lung function and antioxidant enzyme activity was observed in both populations during harvesting; this decrease was greater among the sugarcane workers. The urinary 1-hydroxypyrene levels only increased in the sugarcane workers during the harvesting period. The malonaldehyde levels were elevated in both groups, with a higher increase observed in the workers. This research demonstrates the exposure of sugarcane workers and the inhabitants of a neighbouring town to high PM2.5 concentrations during the sugarcane harvest period. This exposure was higher among the sugarcane workers, as illustrated by both higher PM2.5 concentrations in the sugarcane fields and higher urinary 1-hydroxypyrene levels in the volunteers in this group. The higher incidence of respiratory symptoms, greater decrease in lung function and more marked elevation of oxidative stress markers among the sugarcane workers during the harvest confirms the greater effect magnitude in this population and a dose-dependent relationship between pollution and the observed effects. (C) 2012 Elsevier B.V. All rights reserved.

Renal function evaluation in patients with American Cutaneous Leishmaniasis after specific treatment with pentavalent antimonial

Background: Renal evaluation studies are rare in American Cutaneous Leishmaniasis (ACL). The aim of this study is to investigate whether specific treatment reverts ACL-associated renal dysfunction. Methods: A prospective study was conducted with 37 patients with ACL. Urinary concentrating and acidification ability was assessed before and after treatment with pentavalent antimonial. Results: The patients mean age was 35.6 +/- 12 years and 19 were male. Before treatment, urinary concentrating defect (U/P-osm < 2.8) was identified in 27 patients (77%) and urinary acidification defect in 17 patients (46%). No significant glomerular dysfunction was observed before and after specific ACL treatment. There was no reversion of urinary concentrating defects, being observed in 77% of the patients before and in 88% after treatment (p = 0.344). Urinary acidification defect was corrected in 9 patients after treatment, reducing its prevalence from 40% before to only 16% after treament, (p = 0.012). Microalbuminuria higher than 30 mg/g was found in 35% of patients before treatment and in only 8% after treatment. Regarding fractional excretion of sodium, potassium, calcium, phosphorus and magnesium, there was no significant difference between pre and post-treatment period. Conclusion: As previously described, urinary concentrating and acidification defects were found in an important number of patients with ACL. Present results demonstrate that only some patients recover urinary acidification capacity, while no one returned to normal urinary concentration capacity.

Chronic Stress Improves the Myocardial Function without Altering L-type Ca+2 Channel Activity in Rats

Background: Chronic stress is associated with cardiac remodeling; however the mechanisms have yet to be clarified. Objective: The purpose of this study was test the hypothesis that chronic stress promotes cardiac dysfunction associated to L-type calcium Ca2+ channel activity depression. Methods: Thirty-day-old male Wistar rats (70 - 100 g) were distributed into two groups: control (C) and chronic stress (St). The stress was consistently maintained at immobilization during 15 weeks, 5 times per week, 1h per day. The cardiac function was evaluated by left ventricular performance through echocardiography and by ventricular isolated papillary muscle. The myocardial papillary muscle activity was assessed at baseline conditions and with inotropic maneuvers such as: post-rest contraction and increases in extracellular Ca2+ concentration, in presence or absence of specific blockers L-type calcium channels. Results: The stress was characterized for adrenal glands hypertrophy, increase of systemic corticosterone level and arterial hypertension. The chronic stress provided left ventricular hypertrophy. The left ventricular and baseline myocardial function did not change with chronic stress. However, it improved the response of the papillary muscle in relation to positive inotropic stimulation. This function improvement was not associated with the L-type Ca2+ channel. Conclusion: Chronic stress produced cardiac hypertrophy; however, in the study of papillary muscle, the positive inotropic maneuvers potentiated cardiac function in stressed rats, without involvement of L-type Ca2+ channel. Thus, the responsible mechanisms remain unclear with respect to Ca2+ influx alterations. (Arq Bras Cardiol 2012;99(4):907-914)

Atorvastatin Prevents the Downregulation of Aquaporin-2 Receptor After Bilateral Ureteral Obstruction and Protects Renal Function in a Rat Model

OBJECTIVE To assess the effects of atorvastatin (ATORV) on renal function after bilateral ureteral obstruction (BUO), measuring inulin clearance and its effect on renal hemodynamic, filtration, and inflammatory response, as well as the expression of Aquaporin-2 (AQP2) in response to BUO and after the release of BUO. METHODS Adult Munich-Wistar male rats were subjected to BUO for 24 hours and monitored during the following 48 hours. Rats were divided into 5 groups: sham operated (n = 6); sham + ATORV (n = 6); BUO (n = 6); BUO + ATORV (10 mg/kg in drinking water started 2 days before BUO [n = 5]; and BUO + ATORV (10 mg/kg in drinking water started on the day of the release of BUO [n = 5]). We measured blood pressure (BP, mm Hg); inulin clearance (glomerular filtration rate [GFR]; mL/min/100 g); and renal blood flow (RBF, mL/min, by transient-time flowmeter). Inflammatory response was evaluated by histologic analysis of the interstitial area. AQP2 expression was evaluated by electrophoresis and immunoblotting. RESULTS Renal function was preserved by ATORV treatment, even if initiated on the day of obstruction release, as expressed by GFR, measured by inulin clearance. Relative interstitial area was decreased in both BUO + ATORV groups. Urine osmolality was improved in the ATORV-treated groups. AQP2 protein expression decreased in BUO animals and was reverted by ATORV treatment. CONCLUSION ATORV administration significantly prevented and restored impairment in GFR and renal vascular resistance. Furthermore, ATORV also improved urinary concentration by reversing the BUO-induced downregulation of AQP2. These findings have significant clinical implication in treating obstructive nephropathy. UROLOGY 80: 485.e15-485.e20, 2012. (c) 2012 Elsevier Inc.

Renal function evaluation in patients with American Cutaneous Leishmaniasis after specific treatment with pentavalent antimonial

Background Renal evaluation studies are rare in American Cutaneous Leishmaniasis (ACL). The aim of this study is to investigate whether specific treatment reverts ACL-associated renal dysfunction. Methods A prospective study was conducted with 37 patients with ACL. Urinary concentrating and acidification ability was assessed before and after treatment with pentavalent antimonial. Results The patients mean age was 35.6 ± 12 years and 19 were male. Before treatment, urinary concentrating defect (U/Posm <2.8) was identified in 27 patients (77%) and urinary acidification defect in 17 patients (46%). No significant glomerular dysfunction was observed before and after specific ACL treatment. There was no reversion of urinary concentrating defects, being observed in 77% of the patients before and in 88% after treatment (p = 0.344). Urinary acidification defect was corrected in 9 patients after treatment, reducing its prevalence from 40% before to only 16% after treament, (p = 0.012). Microalbuminuria higher than 30 mg/g was found in 35% of patients before treatment and in only 8% after treatment. Regarding fractional excretion of sodium, potassium, calcium, phosphorus and magnesium, there was no significant difference between pre and post-treatment period. Conclusion As previously described, urinary concentrating and acidification defects were found in an important number of patients with ACL. Present results demonstrate that only some patients recover urinary acidification capacity, while no one returned to normal urinary concentration capacity.

Regulation of corticosterone function during early weaning and effects on gastric cell proliferation

Objectives: The development of the gastrointestinal tract depends on many elements, including glucocorticoids. In the current study, we evaluated the effects of early weaning on corticosterone function and the growth of rat gastric mucosa. Methods: By using Wistar rats submitted to early weaning at 15 d, we analyzed plasma corticosterone, corticosteroid-binding globulin (CBG), and glucocorticoid receptor (GR) distribution in the gastric epithelium. Results: With the use of radioimmunoassay, we found that early weaning increased corticosterone concentration at day 16 and 17 in test subjects as compared with controls, whereas it was equivalent between groups at day 18. CBG binding capacity decreased during treatment, and it was significantly lower at day 18. At this age, GR levels and distribution in the gastric mucosa were also reduced as compared with suckling counterparts. To reduce corticosterone activity during early weaning and to explore cell proliferation responses, we administered RU486 to 15-d-old pups. We found that cytoplasmic GR reached a peak after 48 h, whereas nuclear levels remained constant, thereby confirming the inhibition of receptor function. Next, by checking gastric proliferative responses, we observed that RU486 induced higher DNA synthesis and mitotic indices in test subjects as compared with control groups. Conclusions: We demonstrated that early weaning changed corticosterone activity by increasing hormone levels, reducing CBG binding capacity, and decreasing GR distribution in the gastric epithelium. These modifications seem to be important to the reorganization of gastric growth after the abrupt interruption of suckling.

Altered phenotype and function of dendritic cells in individuals with chronic periodontitis

OBJECTIVE: To investigate the effects of periodontal bacterial lysates on maturation and function of mature monocyte-derived dendritic cells (m-MDDCs) derived from individuals with chronic periodontitis (CP) or healthy periodontal tissue (HP). DESIGN: m-MDDCs derived from peripheral blood monocytes, cultured for 7 days in presence of interleukin (IL)-4 and granulocyte-macrophage colony stimulating factor (GM-CSF), were stimulated with lysates of Streptococcus sanguinis, Prevotella intermedia, Porphyromonas gingivalis, or Treponema denticola on day 4, and were then phenotyped. IL-10, IL-12 and IFN-gamma concentration in the supernatant of cultures were measured. RESULTS: Expression of HLA-DR was lower in bacterial-unstimulated mature m-MDDC from CP compared to HP (p=0.04), while expression of CD1a and CD123 were higher in CP. The expression pattern of HLA-DR, CD11c, CD123, and CD1a did not change on bacterial stimulation, regardless of the bacteria. Stimulation with P. intermedia upregulated CD80 and CD86 in CP cells (p≤0.05). Production of IL-12p70 by bacterial-unstimulated m-MDDCs was 5.8-fold greater in CP compared to HP. Bacterial stimulation further increased IL-12p70 production while decreasing IL-10. Significantly more IFN-gamma was produced in co-cultures of CP m-MDDCs than with HP m-MDDCs when cells were stimulated with P. intermedia (p=0.009). CONCLUSIONS: Bacterial-unstimulated m-MDDC from CP exhibited a more immature phenotype but a cytokine profile biased towards proinflammatory response; this pattern was maintained/exacerbated after bacterial stimulation. P. intermedia upregulated co-stimulatory molecules and IFN-gamma expression in CP m-MDDC. These events might contribute to periodontitis pathogenesis

Computational analyses on the structure-function relation in ion channels

Ion channels are protein molecules, embedded in the lipid bilayer of the cell membranes. They act as powerful sensing elements switching chemicalphysical stimuli into ion-fluxes. At a glance, ion channels are water-filled pores, which can open and close in response to different stimuli (gating), and one once open select the permeating ion species (selectivity). They play a crucial role in several physiological functions, like nerve transmission, muscular contraction, and secretion. Besides, ion channels can be used in technological applications for different purpose (sensing of organic molecules, DNA sequencing). As a result, there is remarkable interest in understanding the molecular determinants of the channel functioning. Nowadays, both the functional and the structural characteristics of ion channels can be experimentally solved. The purpose of this thesis was to investigate the structure-function relation in ion channels, by computational techniques. Most of the analyses focused on the mechanisms of ion conduction, and the numerical methodologies to compute the channel conductance. The standard techniques for atomistic simulation of complex molecular systems (Molecular Dynamics) cannot be routinely used to calculate ion fluxes in membrane channels, because of the high computational resources needed. The main step forward of the PhD research activity was the development of a computational algorithm for the calculation of ion fluxes in protein channels. The algorithm - based on the electrodiffusion theory - is computational inexpensive, and was used for an extensive analysis on the molecular determinants of the channel conductance. The first record of ion-fluxes through a single protein channel dates back to 1976, and since then measuring the single channel conductance has become a standard experimental procedure. Chapter 1 introduces ion channels, and the experimental techniques used to measure the channel currents. The abundance of functional data (channel currents) does not match with an equal abundance of structural data. The bacterial potassium channel KcsA was the first selective ion channels to be experimentally solved (1998), and after KcsA the structures of four different potassium channels were revealed. These experimental data inspired a new era in ion channel modeling. Once the atomic structures of channels are known, it is possible to define mathematical models based on physical descriptions of the molecular systems. These physically based models can provide an atomic description of ion channel functioning, and predict the effect of structural changes. Chapter 2 introduces the computation methods used throughout the thesis to model ion channels functioning at the atomic level. In Chapter 3 and Chapter 4 the ion conduction through potassium channels is analyzed, by an approach based on the Poisson-Nernst-Planck electrodiffusion theory. In the electrodiffusion theory ion conduction is modeled by the drift-diffusion equations, thus describing the ion distributions by continuum functions. The numerical solver of the Poisson- Nernst-Planck equations was tested in the KcsA potassium channel (Chapter 3), and then used to analyze how the atomic structure of the intracellular vestibule of potassium channels affects the conductance (Chapter 4). As a major result, a correlation between the channel conductance and the potassium concentration in the intracellular vestibule emerged. The atomic structure of the channel modulates the potassium concentration in the vestibule, thus its conductance. This mechanism explains the phenotype of the BK potassium channels, a sub-family of potassium channels with high single channel conductance. The functional role of the intracellular vestibule is also the subject of Chapter 5, where the affinity of the potassium channels hEag1 (involved in tumour-cell proliferation) and hErg (important in the cardiac cycle) for several pharmaceutical drugs was compared. Both experimental measurements and molecular modeling were used in order to identify differences in the blocking mechanism of the two channels, which could be exploited in the synthesis of selective blockers. The experimental data pointed out the different role of residue mutations in the blockage of hEag1 and hErg, and the molecular modeling provided a possible explanation based on different binding sites in the intracellular vestibule. Modeling ion channels at the molecular levels relates the functioning of a channel to its atomic structure (Chapters 3-5), and can also be useful to predict the structure of ion channels (Chapter 6-7). In Chapter 6 the structure of the KcsA potassium channel depleted from potassium ions is analyzed by molecular dynamics simulations. Recently, a surprisingly high osmotic permeability of the KcsA channel was experimentally measured. All the available crystallographic structure of KcsA refers to a channel occupied by potassium ions. To conduct water molecules potassium ions must be expelled from KcsA. The structure of the potassium-depleted KcsA channel and the mechanism of water permeation are still unknown, and have been investigated by numerical simulations. Molecular dynamics of KcsA identified a possible atomic structure of the potassium-depleted KcsA channel, and a mechanism for water permeation. The depletion from potassium ions is an extreme situation for potassium channels, unlikely in physiological conditions. However, the simulation of such an extreme condition could help to identify the structural conformations, so the functional states, accessible to potassium ion channels. The last chapter of the thesis deals with the atomic structure of the !- Hemolysin channel. !-Hemolysin is the major determinant of the Staphylococcus Aureus toxicity, and is also the prototype channel for a possible usage in technological applications. The atomic structure of !- Hemolysin was revealed by X-Ray crystallography, but several experimental evidences suggest the presence of an alternative atomic structure. This alternative structure was predicted, combining experimental measurements of single channel currents and numerical simulations. This thesis is organized in two parts, in the first part an overview on ion channels and on the numerical methods adopted throughout the thesis is provided, while the second part describes the research projects tackled in the course of the PhD programme. The aim of the research activity was to relate the functional characteristics of ion channels to their atomic structure. In presenting the different research projects, the role of numerical simulations to analyze the structure-function relation in ion channels is highlighted.

Genomic and non genomic effects of elevated concentration of anabolic steroids in human neuronal cells

Nandrolone and other anabolic androgenic steroids (AAS) at elevated concentration can alter the expression and function of neurotransmitter systems and contribute to neuronal cell death. This effect can explain the behavioural changes, drug dependence and neuro degeneration observed in steroid abuser. Nandrolone treatment (10-8M–10-5M) caused a time- and concentration-dependent downregulation of mu opioid receptor (MOPr) transcripts in SH-SY5Y human neuroblastoma cells. This effect was prevented by the androgen receptor (AR) antagonist hydroxyflutamide. Receptor binding assays confirmed a decrease in MOPr of approximately 40% in nandrolonetreated cells. Treatment with actinomycin D (10-5M), a transcription inhibitor, revealed that nandrolone may regulate MOPr mRNA stability. In SH-SY5Y cells transfected with a human MOPr luciferase promoter/reporter construct, nandrolone did not alter the rate of gene transcription. These results suggest that nandrolone may regulate MOPr expression through post-transcriptional mechanisms requiring the AR. Cito-toxicity assays demonstrated a time- and concentration dependent decrease of cells viability in SH-SY5Y cells exposed to steroids (10-6M–10-4M). This toxic effects is independent of activation of AR and sigma-2 receptor. An increased of caspase-3 activity was observed in cells treated with Nandrolone 10-6M for 48h. Collectively, these data support the existence of two cellular mechanisms that might explain the neurological syndromes observed in steroids abuser.

Analysing the possible influence of transposon TCl4.7 insertion on the function of the genome of Cydia pomonella granulovirus

CpGV-MCp5 is a natural mutant of the Cydia pomonella Granulovirus (Mexican isolate) (CpGV-M) that harbors an insect host transposon termed TCl4.7 in its genome. TCl4.7 is located between the open reading frames Cp15 and Cp16 and separates two homologous regions hr3 and hr4, which have been recently shown to be origins of replication of CpGV-M. The MCp5 has a significant replication disadvantage in the presence of the wild-type CpGV-M. In this study, the possible effects of TCl4.7 transposon insertion on the genome function of its insertion site has been analysed. The role of Cp15 and Cp16 in the context of the virus infection cycle was examined by generating a CpGV-Bacmid (CpBAC) and Cp15 knock-out (CpBACCp15KO) and Cp16 knock-out (CpBACCp16KO) mutants. The mutant CpBACCp15KO was not able to replicate in CM larvae suggesting that Cp15 was essential for virus replication. In contrast, the mutant CpBACCp16KO infected CM larvae and produced viable occlusion bodies (OBs) demonstrating that Cp16 is a non-essential gene for virus in vivo infection of C. pomonella. The temporal transcription of Cp15 and Cp16, as well as of Cp31 (F protein) as a control, was analysed using RT-PCR and quantitative real-time PCR. It suggested a general delay or reduction of gene transcription of MCp5 compared to the parental CpGV-M. Western blot analyses using anti-Cp15 and anti-Cp16 polyclonal antibodies, however, did not show any immuno-reactive response. Thus, a direct influence of TCl4.7 on the expression of Cp15 and Cp16 could not be substantiated. To investigate whether the interruption of hr3 and hr4 palindromes affects the virus replication, two mutant bacmids with a deletion of hr3 and hr4 (CpBAChr3/hr4-KO) and another with an insertion of a Kanamycin resistance cassette between hr3 and hr4 (CpBAChr3-kan-hr4) were generated. Both mutant bacmids replicated and produced infectious virus OBs, which did not significantly differ in their median lethal concentration (LC50) and median survival time (ST50) compared to the parental CpBAC. Interestingly, the mutant CpBAChr3-kan-hr4 was very effectively out-competed by parental CpBAC, when CM larvae were co-infected with known ratios of OBs of CpBAC and the mutant CpBAChr3-kan-hr4. These observations suggested a functional co-operation between hr3 and hr4 which was interrupted by the KanR insertion in CpBAChr3-kan-hr4 and possibly by TCl4.7 transposon insertion in the mutant MCp5. This hypothesis may explain the observed replication disadvantage of the mutants MCp5 and CpBAChr3-kan-hr4 in the presence of the parental viruses CpGV-M and CpBAC, respectively.

Regulatory T cell-mediated suppression of Th9 cell development and effector function

T helper (Th) 9 cells are an important subpopulation of the CD4+ T helper cells. Due to their ability to secrete Interleukin-(IL-)9, Th9 cells essentially contribute to the expulsion of parasitic helminths from the intestinal tract but they play also an immunopathological role in the course of asthma. Recently, a beneficial function of Th9 cells in anti-tumor immune responses was published. In a murine melanoma tumor model Th9 cells were shown to enhance the anti-melanoma immune response via the recruitment of CD8+ T cells, dendritic cells and mast cells. In contrast to Th9 effector cells regulatory T cells (Tregs) are able to control an immune response with the aid of different suppressive mechanisms. Based on their ability to suppress an immune response Tregs are believed to be beneficial in asthma by diminishing excessive allergic reactions. However, concerning cancer they can have a detrimental function because Tregs inhibit an effective anti-tumor immune reaction. Thus, the analysis of Th9 suppression by Tregs is of central importance concerning the development of therapeutic strategies for the treatment of cancer and allergic diseases and was therefore the main objective of this PhD thesis.rnIn general it could be demonstrated that the development of Th9 cells can be inhibited by Tregs in vitro. The production of the lineage-specific cytokine IL-9 by developing Th9 cells was completely suppressed at a Treg/Th9 ratio of 1:1 on the transcriptional (qRT-PCR) as well as on the translational level (ELISA). In contrast, the expression of IRF4 that was found to strongly promote Th9 development was not reduced in the presence of Tregs, suggesting that IRF4 requires additional transcription factors to induce the differentiation of Th9 cells. In order to identify such factors, which regulate Th9 development and therefore represent potential targets for Treg-mediated suppressive mechanisms, a transcriptome analysis using “next-generation sequencing” was performed. The expression of some genes which were found to be up- or downregulated in Th9 cells in the presence of Tregs was validated with qRT-PCR. Time limitations prevented a detailed functional analysis of these candidate genes. Nevertheless, the analysis of the suppressive mechanisms revealed that Tregs probably suppress Th9 cells via the increase of the intracellular cAMP concentration. In contrast, IL-9 production by differentiated Th9 cells was only marginally affected by Tregs in vitro and in vivo analysis (asthma, melanoma model). Hence, Tregs represent very effective inhibitors of Th9 development whereas they have only a minimal suppressive influence on differentiated Th9 cells.rn

The relation between mass and concentration in x-ray galaxy clusters at high-redshift.

In this Thesis work we have studied the properties of high-redshift galaxy clusters through the X-ray emission from their intracluster gas. In particular, we have focused on the relation between concentration and mass that is related to the density of the universe at the formation time of the clusters and therefore, it is a powerful cosmological probe. Concentration is expected to be a decreasing function of mass but a complete characterization of this relation has not been reached yet. We have analysed 22 clusters observed withe the Chandra satellite at high redshift and we have investigated the concentration-mass relation.

Effect of Acute Psychological Stress on Prefrontal GABA Concentration Determined by Proton Magnetic Resonance Spectroscopy

Objective Impaired function of the central gamma-aminobutyric acid (GABA) system, which provides the brain’s major inhibitory pathways, is thought to play an important role in the pathophysiology of anxiety disorders. The effect of acute psychological stress on the human GABA-ergic system is still unknown, however. The purpose of this study was to determine the effect of acute stress on prefrontal GABA levels. Method A recently developed noninvasive magnetic resonance spectroscopy method was used to measure changes in the GABA concentration of the prefrontal cortex in 10 healthy human subjects during a threat-of-shock condition and during a safe condition (two sessions on different days). The main outcome measure was the mean GABA concentration within a 3×3×2-cm3 voxel selected from the medial prefrontal cortex. Results Prefrontal GABA decreased by approximately 18% in the threat-of-shock condition relative to the safe condition. This reduction was specific to GABA, since the concentrations of N-acetyl-aspartate, choline-containing compounds, and glutamate/glutamine levels obtained in the same spectra did not change significantly. Conclusions This result appeared compatible with evidence from preclinical studies in rodents, which showed rapid presynaptic down-regulation of GABA-ergic neurotransmission in response to acute psychological stress. The molecular mechanism and functional significance of this reduced inhibitory effect of acute psychological stress in relation to impaired GABA-ergic function in anxiety disorders merit further investigation.

Expression and function of 5-hydroxytryptamine 4 receptors in smooth muscle preparations from the duodenum, ileum, and pelvic flexure of horses without gastrointestinal tract disease

OBJECTIVE: To evaluate the expression of the 5-hydroxytryptamine 4 (5-HT4) receptor subtype and investigate the modulating function of those receptors on contractility in intestinal tissues obtained from horses without gastrointestinal tract disease. SAMPLE POPULATION: Smooth muscle preparations from the duodenum, ileum, and pelvic flexure collected immediately after slaughter of 24 horses with no history or signs of gastrointestinal tract disease. PROCEDURES: In isometric organ baths, the contractile activities of smooth muscle preparations in response to 5-hydroxytryptamine and electric field stimulation were assessed; the effect of tegaserod alone or in combination with 5-hydroxytryptamine on contractility of intestinal specimens was also investigated. Presence and distribution of 5-HT4 receptors in intestinal tissues and localization on interstitial cells of Cajal were examined by use of an immunofluorescence technique. RESULTS: Widespread 5-HT4 receptor immunoreactivity was observed in all intestinal smooth muscle layers; 5-HT4 receptors were absent from the myenteric plexus and interstitial cells of Cajal. In electrical field-stimulated tissue preparations of duodenum and pelvic flexure, tegaserod increased the amplitude of smooth muscle contractions in a concentration-dependent manner. Preincubation with tegaserod significantly decreased the basal tone of the 5-HT-evoked contractility in small intestine specimens, compared with the effect of 5-HT alone, thereby confirming that tegaserod was acting as a partial agonist. CONCLUSIONS AND CLINICAL RELEVANCE: In horses, 5-HT4 receptors on smooth muscle cells appear to be involved in the contractile response of the intestinal tract to 5-hydroxytryptamine. Results suggest that tegaserod may be useful for treatment of reduced gastrointestinal tract motility in horses.

Type 2 diabetes is associated with reduced ATP-binding cassette transporter A1 gene expression, protein and function

Objective Increasing plasma glucose levels are associated with increasing risk of vascular disease. We tested the hypothesis that there is a glycaemia-mediated impairment of reverse cholesterol transport (RCT). We studied the influence of plasma glucose on expression and function of a key mediator in RCT, the ATP binding cassette transporter-A1 (ABCA1) and expression of its regulators, liver X receptor-α (LXRα) and peroxisome proliferator-activated receptor–γ (PPARγ). Methods and Results Leukocyte ABCA1, LXRα and PPARγ expression was measured by polymerase chain reaction in 63 men with varying degrees of glucose homeostasis. ABCA1 protein concentrations were measured in leukocytes. In a sub-group of 25 men, ABCA1 function was quantified as apolipoprotein-A1-mediated cholesterol efflux from 2–3 week cultured skin fibroblasts. Leukocyte ABCA1 expression correlated negatively with circulating HbA1c and glucose (rho = −0.41, p<0.001; rho = −0.34, p = 0.006 respectively) and was reduced in Type 2 diabetes (T2DM) (p = 0.03). Leukocyte ABCA1 protein was lower in T2DM (p = 0.03) and positively associated with plasma HDL cholesterol (HDL-C) (rho = 0.34, p = 0.02). Apolipoprotein-A1-mediated cholesterol efflux correlated negatively with fasting glucose (rho = −0.50, p = 0.01) and positively with HDL-C (rho = 0.41, p = 0.02). It was reduced in T2DM compared with controls (p = 0.04). These relationships were independent of LXRα and PPARγ expression. Conclusions ABCA1 expression and protein concentrations in leukocytes, as well as function in cultured skin fibroblasts, are reduced in T2DM. ABCA1 protein concentration and function are associated with HDL-C levels. These findings indicate a glycaemia- related, persistent disruption of a key component of RCT.