Search

A mathematical model of ascending Henle limb (AHL) epithelium has been fashioned using kinetic representations of [Na.sup.+]-[K.sup.+]-2[Cl.sup.-] cotransporter (NKCC2), KCC4, and type 3 [Na.sup.+]/[H.sup.+] exchanger (NHE3), with transporter densities selected to yield the reabsorptive [Na.sup.+] flux expected for rat tubules in vivo. Of necessity, this model predicts fluxes that are higher than those measured in vitro. The kinetics of the NKCC and KCC are such that [Na.sup.+] reabsorption by the model tubule is responsive to variation in luminal NaCl concentration over the range of 30 to 130 mM, with only minor changes in cell volume. Peritubular KCC accounts for about half the reabsorptive [Cl.sup.-] flux, with the remainder via peritubular [Cl.sup.-] channels. Transcellular [Na.sup.+] flux is turned off by increasing peritubular KCl, which produces increased cytosolic Cl- and thus inhibits NKCC2 transport. In the presence of physiological concentrations of ammonia, there is a large acid challenge to the cell, due primarily to N[H.sup.+.sub.4] entry via NKCC2, with diffusive N[H.sub.3] exit to both lumen and peritubular solutions. When NHE3 density is adjusted to compensate this acid challenge, the model predicts luminal membrane proton secretion that is greater than the HC[O.sub.-.sub.3]-reabsorptive fluxes measured in vitro. The model also predicts luminal membrane ammonia cycling, with uptake via NKCC2 or [K.sup.+] channel, and secretion either as N[H.sup.+.sub.4] by NHE3 or as diffusive N[H.sub.3] flux in parallel with a secreted proton. If such luminal ammonia cycling occurs in vivo, it could act in concert with luminal [K.sup.+] cycling to facilitate AHL [Na.sup.+] reabsorption via NKCC2. With physiological ammonia, peritubular KCl also blunts NHE3 activity by inhibiting N[H.sup.+.sub.4] uptake on the Na-K-ATPase, and alkalinizing the cell. NKCC; KCC; NHE3; sodium transport; potassium transport; ammonia doi:10.1152/ajprenal.00231.2009

Adenosine mediates vascular responses through four receptor subtypes: [A.sub.1], [A.sub.2A], [A.sub.2B], and [A.sub.3]. The role of [A.sub.2A] receptors in aortic vascular tone was investigated using [A.sub.2A] adenosine receptor (AR) knockout ([A.sub.2A]KO) and corresponding wild-type ([A.sub.2A]WT) mice. Isolated aortic rings from [A.sub.2A]WT and [A.sub.2A]KO mice were precontracted with phenylephrine ([10.sup.-7] M), and concentration responses for adenosine analogs and selective agonists/ antagonists were obtained. Nonselective adenosine analog (NECA; [EC.sub.50] = 6.78 [micro]M) and CGS-21680 ([A.sub.2A]AR selective agonist; [EC.sub.50] = 0.013 [micro]M) produced concentration-dependent relaxation (maximum of 25% and 28% relaxation at [10.sup.-5] M NECA and CGS-21680, respectively) in [A.sub.2A] WT aorta. In [A.sub.2A]KO aorta, NECA ([EC.sub.50] = 0.075 [micro]M) induced concentration-dependent contraction (maximum contraction of 47% at [10.sup.-6] M; P < 0.05 compared with [A.sub.2A] WT), whereas CGS-21680 produced no response. SCH-58261 ([10.sup.-6] M; [A.sub.2A]AR selective antagonist) abolished both NECA- and CGS-21680-mediated vasorelaxation in [A.sub.2A] WT (P < 0.05), whereas no change was observed in [A.sub.2A]KO. When DPCPX ([10.sup.-5] M; [A.sub.1] selective antagonist) was used in NECA concentration response, greater vasorelaxation was observed in [A.sub.2A]WT (50% vs. 25% in controls at [10.sup.-5] M; P < 0.05), whereas lower contraction was seen in [A.sub.2A]KO tissues (5% vs. 47% in controls at [10.sup.-6] M; P < 0.05). Aortic endothelial function, determined by response to acetylcholine, was significantly higher in WT compared with KO (66% vs. 51%; P < 0.05). BAY 60-6583 ([A.sub.2B] selective agonist) produced similar relaxation in both KO and WT tissues. In conclusion, [A.sub.2A]AR KO mice had significantly lower aortic relaxation and endothelial function, suggesting that the [A.sub.2A] AR plays an important role in vasorelaxation, probably through an endothelium-dependent mechanism. endothelium; mouse aorta; vasorelaxation; adenosine agonists; antagonists doi: 10.1152/ajpheart.00192.2009.

Acid-base transport in the brush border of the proximal tubule in response to the presence of carbon dioxide and carbonic acid was investigated in a mathematical model that allows simulation of carbonic anhydrase (CA) activity and its inhibition. CA was shown to cause an imbalance in pH gradients and depressed sodium bicarbonate reabsorption through the brush border by acting as a barrier to the diffusion of bicarbonate ions.

Abstract Background Circulating tumour cells (CTCs) have shown prognostic relevance in metastatic breast, prostate, colon and pancreatic cancer. For further development of CTCs as a biomarker, we compared the performance of different protocols for CTC detection in murine breast cancer xenograft models (MDA-MB-231, MDA-MB-468 and KPL-4). Blood samples were taken from tumour bearing animals (20 to 200 mm2) and analysed for CTCs using 1. an epithelial marker based enrichment method (AdnaTest), 2. an antibody independent technique, targeting human gene transcripts (qualitative PCR), and 3. an antibody-independent approach, targeting human DNA-sequences (quantitative PCR). Further, gene expression changes associated with epithelial-to-mesenchymal transition (EMT) were determined with an EMT-specific PCR assay. Methods We used the commercially available Adna Test, RT-PCR on human housekeeping genes and a PCR on AluJ sequences to detect CTCs in xenografts models. Phenotypic changes in CTCs were tested with the commercially available “Human Epithelial to Mesenchymal Transition RT-Profiler PCR Array”. Results Although the AdnaTest detects as few as 1 tumour cell in 1 ml of mouse blood spiking experiments, no CTCs were detectable with this approach in vivo despite visible metastasis formation. The presence of CTCs could, however, be demonstrated by PCR targeting human transcripts or DNA-sequences - without epithelial pre-enrichment. The failure of CTC detection by the AdnaTest resulted from downregulation of EpCAM, whereas mesenchymal markers like Twist and EGFR were upregulated on CTCs. Such a change in the expression profile during metastatic spread of tumour cells has already been reported and was linked to a biological program termed epithelial-mesenchymal transition (EMT). Conclusions The use of EpCAM-based enrichment techniques leads to the failure to detect CTC populations that have undergone EMT. Our findings may explain clinical results where low CTC numbers have been reported even in patients with late metastatic cancers. These results are a starting point for the identification of new markers for detection or capture of CTCs, including the mesenchymal-like subpopulations.

Abstract Background Left ventricular diastolic dysfunction (LVDD) is considered a precursor of diabetic cardiomyopathy, while insulin resistance (IR) is a precursor of type 2 diabetes mellitus (T2DM) and independently predicts heart failure (HF). We assessed whether IR and abnormalities of the glucose metabolism are related to LVDD. Methods We included 208 patients with normal ejection fraction, 57 (27%) of whom had T2DM before inclusion. In subjects without T2DM, an oral glucose tolerance test (oGTT) was performed. IR was assessed using the Homeostasis Model Assessment of Insulin Resistance (HOMA-IR). The lower limit of the top quartile of the HOMA-IR distribution (3.217) was chosen as threshold for IR. LVDD was verified according to current guidelines. Results IR was diagnosed in 38 (18%) patients without a history of diabetes. The prevalence of LVDD was 92% in subjects with IR vs. 72% in patients without IR (n = 113), respectively (p = 0.013). In the IR group, the early diastolic mitral inflow velocity (E) in relation to the early diastolic tissue Doppler velocity (averaged from the septal and lateral mitral annulus, E'av) ratio (E/E'av) was significantly higher compared to those without IR (9.8 [8.3-11.5] vs. 8.1 [6.6-11.0], p = 0.011). This finding remains significant when patients with IR and concomitant T2DM based on oGTT results were excluded (E/E'av ratio 9.8 [8.2-11.1)] in IR vs. 7.9 [6.5-10.5] in those without both IR and T2DM, p = 0.014). There were significant differences among patients with and without LVDD regarding the HOMA-IR (1.71 [1.04-3.88] vs. 1.09 [0.43-2.2], p = 0.003). The HOMA-IR was independently associated with LVDD on multivariate logistic regression analysis, a 1-unit increase in HOMA-IR value was associated with an odds ratio for prevalent LVDD of 2.1 (95% CI 1.3-3.1, p = 0.001). Furthermore, the E/E'av ratio increases along the glucose metabolism status from normal glucose metabolism (7.6 [6.2-10.1]) to impaired glucose tolerance (8.8 [7.4-11.0]) and T2DM (10.5 [8.1-13.2]), respectively (p < 0.001). Conclusions Insulin resistance is independently associated with LVDD in subjects without overt T2DM. Patients with IR and glucose metabolism disorders might represent a target population to prevent the development of HF. Screening programs for glucose metabolism disturbances should address the assessment of diastolic function and probably IR.

Abstract Background Increased muscle mass index of the left ventricle (LVMi) is an independent predictor for the development of symptoms in patients with asymptomatic aortic stenosis (AS). While the onset of clinical symptoms and left ventricular systolic dysfunction determines a poor prognosis, the standard echocardiographic evaluation of LV dysfunction, only based on measurements of the LV ejection fraction (EF), may be insufficient for an early assessment of imminent heart failure. Contrary, 2-dimensional speckle tracking (2DS) seems to be superior in detecting subtle changes in myocardial function. The aim of the study was to assess these LV function deteriorations with global longitudinal strain (GLS) analysis and the relations to LVMi in patients with AS and normal EF. Methods 50 patients with moderate to severe AS and 31 controls were enrolled. All patients underwent echocardiography, including 2DS imaging. LVMi measures were performed with magnetic resonance imaging in 38 patients with AS and indexed for body surface area. Results The total group of patients with AST showed a GLS of -15,2 ± 3,6% while the control group reached -19,5 ± 2,7% (p < 0,001). By splitting the group with AS in normal, moderate and severe increased LVMi, the GLS was -17,0 ± 2,6%, -13,2 ± 3,8% and -12,4 ± 2,9%, respectively (p = 0,001), where LVMi and GLS showed a significant correlation (r = 0,6, p < 0,001). Conclusions In conclusion, increased LVMi is reflected in abnormalities of GLS and the proportion of GLS impairment depends on the extent of LV hypertrophy. Therefore, simultaneous measurement of LVMi and GLS might be useful to identify patients at high risk for transition into heart failure who would benefit from aortic valve replacement irrespectively of LV EF.

Abstract Background Diabetes mellitus (DM) has reached epidemic proportions and is an important risk factor for heart failure (HF). Left ventricular diastolic dysfunction (LVDD) is recognized as the earliest manifestation of DM-induced LV dysfunction, but its pathophysiology remains incompletely understood. We sought to evaluate the relationship between proinflammatory cytokine levels (TNF-alpha, IL-6) and tissue Doppler derived indices of LVDD in patients with stable coronary artery disease. Methods We enrolled 41 consecutive patients (mean age 65+/-10 years) submitted for coronary angiography. Echocardiographic assessment was performed in all patients. Pulsed tissue Doppler imaging was performed at the mitral annulus and was characterized by the diastolic early relaxation velocity Em. Conventional transmitral flow was measured with pw-doppler. Early (E) transmitral flow velocity was measured. LVDD was defined as E/Em ratio ≥ 15, E/Em 8-14 was classified as borderline. Plasma levels of TNF-alpha and IL-6 were determined in all patients. A standardized oral glucose tolerance test was performed in subjects without diabetes. Results Patients with E/Em ratio ≥ 15, classified as LVDD and those with E/Em ratio 8-14 (classified as borderline) had significantly higher IL-6 (P = 0,001), TNF-alpha (P < 0,001) and NT-pro- BNP (P = 0,001) plasma levels compared to those with normal diastolic function. TNF-alpha and IL-6 levels remains significantly elevated after adjustment for sex, age, left ventricular ejection function, body mass index, coronary heart disease, smoking, hypertension and diabetes mellitus with linear regression analysis. Furthermore, in subjects LVDD or borderline LV diastolic function, 75% had diabetes or IGT, respectively. When subjects without diabetes were excluded, both IL-6 (P = 0,006) and TNF-alpha (P = 0,002) remained significantly elevated in subjects with E/Em ratio ≥ 15. Conclusion This study reveals that increased plasma levels of IL-6 and TNF-alpha were associated with LVDD. These findings suggest a link between low-grade inflammation and the presence of LVDD. An active proinflammatory process may be of importance in the pathogenesis of diastolic dysfunction.