Molecular cloning and characterization of the porcine prostaglandin transporter (SLCO2A1): evaluation of its role in F4 mediated neonatal diarrhoea

<p>Abstract</p> <p>Background</p> <p>Because prostaglandins are involved in many (patho)physiological processes, <it>SLCO2A1 </it>was already characterized in several species in an attempt to unravel specific processes/deficiencies. Here, we describe the molecular cloning and characterization of the porcine ortholog in order to evaluate its possible involvement in F4 enterotoxigenic <it>E. coli </it>mediated neonatal diarrhoea, based on a positional candidate gene approach study.</p> <p>Results</p> <p>Porcine <it>SLCO2A1 </it>is organized in 14 exons, containing an open reading frame of 1935 bp, encoding a 12-transmembrane organic anion cell surface transporter of 644 aa. The -388 to -5 upstream region comprises a (CpG)₄₈island containing a number of conserved promoter elements, including a TATA box. A potential alternative promoter region was found in the conserved -973 to -700 upstream region. No consensus polyadenylation signal was discovered in the 3' UTR. Repeat sequences were found in 15% of all the non coding sequences.</p> <p>As expected for a multifunctional protein, a wide tissue distribution was observed. mRNA expression was found in the adrenal gland, bladder, caecum, colon (centripetal coil/centrifugal coil), diaphragm, duodenum, gallbladder, heart, ileum, jejunum, kidney, liver, longissimus dorsi muscle, lung, lymph node, mesenterium, rectum, spleen, stomach, tongue and ureter, but not in the aorta, oesophagus and pancreas.</p> <p>The promoter region and the exons (including the splice sites) of <it>SLCO2A1 </it>were resequenced in 5 F4ab/ac receptor positive and 5 F4ab/ac receptor negative pigs. Two silent and 2 missense (both S → L at position 360 and 633) mutations were found, but none was associated with the F4ab/ac receptor phenotype. In addition, no phenotype associated differential mRNA expression or alternative/abberant splicing/polyadenylation was found in the jejunum.</p> <p>Conclusion</p> <p>The molecular cloning and characterization of porcine <it>SLCO2A1 </it>not only contributes to the already existing knowledge about the transporter in general, but enables studies on porcine prostaglandin related processes/deficiencies as patient and/or model. Here we examined its possible involvement as receptor in F4 enterotoxigenic <it>E. coli </it>mediated neonatal diarrhoea. Because no phenotype associated differences could be found in the gene sequence nor in its jejunal transcription profile of F4ab/ac receptor positive/negative pigs, SLCO2A1 can most likely be excluded as receptor for F4 bacteria.</p

Retinal microvascular parameters are not associated with reduced renal function in a study of individuals with type 2 diabetes

Abstract The eye provides an opportunistic “window” to view the microcirculation. There is published evidence of an association between retinal microvascular calibre and renal function measured by estimated glomerular filtration rate (eGFR) in individuals with diabetes mellitus. Beyond vascular calibre, few studies have considered other microvascular geometrical features. Here we report novel null findings for measures of vascular spread (vessel fractal dimension), tortuosity, and branching patterns and their relationship with renal function in type 2 diabetes over a mean of 3 years. We performed a nested case-control comparison of multiple retinal vascular parameters between individuals with type 2 diabetes and stable (non-progressors) versus declining (progressors) eGFR across two time points within a subset of 1072 participants from the GoDARTS study cohort. Retinal microvascular were measured using VAMPIRE 3.1 software. In unadjusted analyses and following adjustment for age, gender, systolic blood pressure, HbA1C, and diabetic retinopathy, no associations between baseline retinal vascular parameters and risk of eGFR progression were observed. Cross-sectional analysis of follow-up data showed a significant association between retinal arteriolar diameter and eGFR, but this was not maintained following adjustment. These findings are consistent with a lack of predictive capacity for progressive loss of renal function in type 2 diabetes

Modulation of endogenous antioxidant defense and the progression of kidney disease in multi-heritage groups of patients with type 2 diabetes: PRospective EValuation of Early Nephropathy and its Treatment (PREVENT).

BACKGROUND: Diabetes is the western world's leading cause of end-stage renal disease. Glucose-dependent, oxidative stress is linked to the development of renal inflammation and sclerosis, which, in animal models of diabetes, can be prevented by anti-oxidative treatment. Patients of non-Caucasian heritage have low activity of the selenoprotein, antioxidant enzyme, glutathione peroxidase (GPx) and its co-factor vitamin E, which may be linked to their increased propensity to developing end-stage renal disease. RESEARCH DESIGN AND METHODS: We have designed a double-blind, randomized, placebo controlled study with selenium and/or vitamin E versus placebo as the interventions for patients with type 2 diabetes and chronic kidney disease (CKD) stages 1-3. A 2 × 2 factorial design will allow a balanced representation of the heritage groups exposed to each intervention. The primary biochemical outcome is change in GPx activity, and clinical outcome measure is the actual, rate of-and/or percentage change in estimated glomerular filtration rate (eGFR) from baseline. Analysis will be with a marginal model for longitudinal data using Generalized Estimating Equations corrected for measures of baseline serum antioxidant enzyme activities (GPx, superoxide dismutase and catalase), micronutrient levels (vitamins E and C), measures of inflammation (interleukin 6, c-reactive protein and monocyte chemoattractant protein-1) and markers of oxidative damage (plasma 8-isoprostaglandin F2α and urinary 8-hydroxydeoxyguanosine). EXPECTED RESULTS: The study will assess the relationship between GPx activity, oxidative stress, inflammation and eGFR. It will test the null hypothesis that antioxidant therapy does not influence the activity of GPx or other antioxidant enzymes and/or alter the rate of change in eGFR in these patient groups. CONCLUSIONS: Outcome data on the effect of antioxidants in human diabetic renal disease is limited. Previous post hoc analyses have not shown a beneficial effect of vitamin E on renal function. A recent trial of a pharmaceutical antioxidant agent, improved eGFR, but in patients with advanced diabetes-related chronic kidney disease its use was associated with an increased incidence of cardiovascular events. We will explore whether the nutritional antioxidants, vitamin E and selenium alone, or in combination in patients at high risk of renal disease progression, forestalls a reduction in eGFR. The study will describe whether endogenous antioxidant enzyme defenses can be safely modified by this intervention and how this is associated with changes in markers of oxidative stress. Trial registration ISRCTN 97358113. Registered 21st September 2009

Overweight status is associated with extensive signs of microvascular dysfunction and cardiovascular risk

The aim of this present study was to investigate if overweight individuals exhibit signs of vascular dysfunction associated with a high risk for cardiovascular disease (CVD). One hundred lean and 100 overweight participants were recruited for the present study. Retinal microvascular function was assessed using the Dynamic Retinal Vessel Analyser (DVA), and systemic macrovascular function by means of flow-mediated dilation (FMD). Investigations also included body composition, carotid intimal-media thickness (c-IMT), ambulatory blood pressure monitoring (BP), fasting plasma glucose, triglycerides (TG), cholesterol levels (HDL-C and LDL-C), and plasma von Willebrand factor (vWF). Overweight individuals presented with higher right and left c-IMT (p = 0.005 and p = 0.002, respectively), average 24-h BP values (all p <0.001), plasma glucose (p = 0.008), TG (p = 0.003), TG: HDL-C ratio (p = 0.010), and vWF levels (p = 0.004). Moreover, overweight individuals showed lower retinal arterial microvascular dilation (p = 0.039) and baseline-corrected flicker (bFR) responses (p = 0.022), as well as, prolonged dilation reaction time (RT, p = 0.047). These observations emphasise the importance of vascular screening and consideration of preventive interventions to decrease vascular risk in all individuals with adiposity above normal range