23Na Magnetic Resonance Imaging of the Lower Leg of Acute Heart Failure Patients during Diuretic Treatment

Objective

Na⁺ can be stored in muscle and skin without commensurate water accumulation. The aim of this study was to assess Na⁺ and H₂O in muscle and skin with MRI in acute heart failure patients before and after diuretic treatment and in a healthy cohort.

Methods

Nine patients (mean age 78 years; range 58–87) and nine age and gender-matched controls were studied. They underwent ²³Na/¹H-MRI at the calf with a custom-made knee coil. Patients were studied before and after diuretic therapy. ²³Na-MRI gray-scale measurements of Na⁺-phantoms served to quantify Na⁺-concentrations. A fat-suppressed inversion recovery sequence was used to quantify H₂O content.

Results

Plasma Na⁺-levels did not change during therapy. Mean Na⁺-concentrations in muscle and skin decreased after furosemide therapy (before therapy: 30.7±6.4 and 43.5±14.5 mmol/L; after therapy: 24.2±6.1 and 32.2±12.0 mmol/L; p˂0.05 and p˂0.01). Water content measurements did not differ significantly before and after furosemide therapy in muscle (p = 0.17) and only tended to be reduced in skin (p = 0.06). Na⁺-concentrations in calf muscle and skin of patients before and after diuretic therapy were significantly higher than in healthy subjects (18.3±2.5 and 21.1±2.3 mmol/L).

Conclusions

²³Na-MRI shows accumulation of Na⁺ in muscle and skin in patients with acute heart failure. Diuretic treatment can mobilize this Na⁺-deposition; however, contrary to expectations, water and Na⁺-mobilization are poorly correlated.     

High ABCC2 and Low ABCG2 Gene Expression Are Early Events in the Colorectal Adenoma-Carcinoma Sequence

Development of colorectal cancer (CRC) may result from a dysfunctional interplay between diet, gut microbes and the immune system. The ABC transport proteins ABCB1 (P-glycoprotein, Multidrug resistance protein 1, MDR1), ABCC2 (MRP2) and ABCG2 (BCRP) are involved in transport of various compounds across the epithelial barrier. Low mRNA level of ABCB1 has previously been identified as an early event in colorectal carcinogenesis (Andersen et al., PLoS One. 2013 Aug 19;8(8):e72119). ABCC2 and ABCG2 mRNA levels were assessed in intestinal tissue from 122 CRC cases, 106 adenoma cases (12 with severe dysplasia, 94 with mild-moderate dysplasia) and from 18 controls with normal endoscopy.We found significantly higher level of ABCC2 in adenomas with mild to moderate dysplasia and carcinoma tissue compared to the levels in unaffected tissue from the same individual (P = 0.037, P = 0.037, and P<0.0001) and in carcinoma and distant unaffected tissue from CRC cases compared to the level in the healthy individuals (P = 0.0046 and P = 0.036). Furthermore, ABCG2 mRNA levels were significantly lower in adenomas and carcinomas compared to the level in unaffected tissue from the same individuals and compared to tissue from healthy individuals (P<0.0001 for all). The level of ABCB2 in adjacent normal tissue was significantly higher than in tissue from healthy individuals (P = 0.011).In conclusion, this study found that ABCC2 and ABCG2 expression levels were altered already in mild/moderate dysplasia in carcinogenesis suggesting that these ABC transporters are involved in the early steps of carcinogenesis as previously reported for ABCB1. These results suggest that dysfunctional transport across the epithelial barrier may contribute to colorectal carcinogenesis.     

RAPID EVOLUTION OF REPRODUCTIVE ISOLATION BETWEEN INCIPIENT OUTCROSSING AND SELFING CLARKIA SPECIES

A major goal of speciation research is to understand the processes involved in the earliest stages of the evolution of reproductive isolation (RI). One important challenge has been to identify systems where lineages have very recently diverged and opportunities for hybridization are present. We conducted a comprehensive examination of the components of RI across the life cycle of two subspecies of Clarkia xantiana, which diverged recently (ca. 65,000 bp). One subspecies is primarily outcrossing, but self‐compatible, whereas the other is primarily selfing. The subspecies co‐occur in a zone of sympatry but hybrids are rarely observed. Premating barriers resulted in nearly complete isolation in both subspecies with flowering time and pollinator preference (for the outcrosser over the selfer) as the strongest barriers. We found that the outcrosser had consistently more competitive pollen, facilitating hybridization in one direction, but no evidence for pollen–pistil interactions as an isolating barrier. Surprisingly, postzygotic isolation was detected at the stage of hybrid seed development, but in no subsequent life stages. This crossing barrier was asymmetric with crosses from the selfer to outcrosser most frequently failing. Collectively, the results provide evidence for rapid evolution of multiple premating and postzygotic barriers despite a very recent divergence time.     

Effects of the new adiponectin paralogous protein CTRP-3 and of LPS on cytokine release from monocytes of patients with type 2 diabetes mellitus

Aims/Hypothesis: It was the aim to investigate the hypothesis that the new C1q/TNF-family member CTRP-3 (C1q/TNF-related protein-3) acts anti-inflammatory in human monocytes from healthy controls and patients with type 2 diabetes mellitus (T2D). Methods: Monocytes were isolated from 20 healthy controls and 30 patients with T2D. IL-6 and TNF concentrations were measured by ELISA. CTRP-3 was expressed in insect cells and used for stimulation experiments. Results: Basal IL-6 and TNF were not different in control and in T2D monocytes. LPS-stimulation (1 μg/ml) significantly (p < 0.001) increased IL-6 and TNF in the supernatants of control and in T2D monocytes to a similar extent. CTRP-3 (1 μg/ml) significantly (p = 0.03) inhibited LPS-induced IL-6 in control monocytes but not in T2D monocytes. TNF upon co-stimulation with LPS and CTRP-3 was significantly (p = 0.012) lower in control than in T2D monocytes. LPS-induced TNF concentration was significantly and positively correlated with serum total cholesterol and LDL cholesterol in T2D patients. Conclusions: CTRP-3 inhibits LPS-induced IL-6 and TNF release. This anti-inflammatory effect is lost in T2D. Serum cholesterol concentration affects the pro-inflammatory potential of LPS to induce TNF release from T2D monocytes in the presence or absence of CTRP-3. CTRP-3 might partly account for the pro-inflammatory state in T2D.     

Dual effect of acid pH on purinergic P2X3 receptors depends on the histidine 206 residue

Whole cell patch clamp investigations were carried out to clarify the pH sensitivity of native and recombinant P2X(3) receptors. In HEK293 cells permanently transfected with human (h) P2X(3) receptors (HEK293-hP2X(3) cells), an acidic pH shifted the concentration-response curve for alpha,beta-methylene ATP (alpha,beta-meATP) to the right and increased its maximum. An alkalic pH did not alter the effect of alpha,beta-meATP. Further, a low pH value increased the activation time constant (tau(on)) of the alpha,beta-meATP current; the fast and slow time constants of desensitization (tau(des1), tau(des2)) were at the same time also increased. Finally, acidification accelerated the recovery of P2X(3) receptors from the desensitized state. Replacement of histidine 206, but not histidine 45, by alanine abolished the pH-induced effects on hP2X(3) receptors transiently expressed in HEK293 cells. Changes in the intracellular pH had no effect on the amplitude or time course of the alpha,beta-meATP currents. The voltage sensitivity and reversal potential of the currents activated by alpha,beta-meATP were unaffected by extracellular acidification. Similar effects were observed in a subpopulation of rat dorsal root ganglion neurons expressing homomeric P2X(3) receptor channels. It is suggested that acidification may have a dual effect on P2X(3) channels, by decreasing the current amplitude at low agonist concentrations (because of a decrease in the rate of activation) and increasing it at high concentrations (because of a decrease in the rate of desensitization). Thereby, a differential regulation of pain sensation during e.g. inflammation may occur at the C fiber terminals of small DRG neurons in peripheral tissues.     

Sphingosine 1-phosphate phosphatase 2 is induced during inflammatory responses

Sphingosine 1-phosphate (S1P) levels in cells and, consequently, its bioactivity as a signalling molecule are controlled by the action of enzymes responsible for its synthesis and degradation. In the present report, we examined alterations in expression patterns of enzymes involved in S1P-metabolism (sphingosine kinases including their splice variants, sphingosine 1-phosphate phosphatases, and sphingosine 1-phosphate lyase) under certain inflammatory conditions. We found that sphingosine kinase type 1 (SPHK1) mRNA could be triggered in a cell type-specific manner; individual SPHK1 splice variants were induced with similar kinetics. Remarkably, expression and activity of S1P phosphatase 2 (SPP2) was found to be highly upregulated by inflammatory stimuli in a variety of cells (e.g., neutrophils, endothelial cells). Bandshift analysis using oligonucleotides spanning predicted NFkappaB sites within the SPP2 promoter and silencing of NFkappaB/RelA via RelA-directed siRNA demonstrated that SPP2 is an NFkappaB-dependent gene. Silencing of SPP2 expression in endothelial cells, in turn, led to a marked reduction of TNF-alpha-induced IL-1beta mRNA and protein and to a partial reduction of induced IL-8, suggesting a pro-inflammatory role of SPP2. Notably, up-regulation of SPP2 was detected in samples of lesional skin of patients with psoriasis, an inflammatory skin disease. This study provides detailed insights into the regulation of SPP2 gene expression and suggests that SPP2 might be a novel player in pro-inflammatory signalling.