The mechano-sensitivity of cardiac ATP-sensitive potassium channels is mediated by intrinsic MgATPase activity.

Cardiac ATP-sensitive K+ (KATP) channel activity plays an important cardio-protective role in regulating excitability in response to metabolic stress. Evidence suggests that these channels are also mechano-sensitive and therefore may couple KATP channel activity to increased cardiac workloads. However, the molecular mechanism that couples membrane stretch to channel activity is not currently known. We hypothesized that membrane stretch may alter the intrinsic MgATPase activity of the cardiac KATP channel resulting in increased channel activation. The inside-out patch-clamp technique was used to record single-channel and macroscopic recombinant KATP channel activity in response to membrane stretch elicited by negative pipette pressure. We found that stretch activation requires the presence of the SUR subunit and that inhibition of MgATPase activity with either the non-hydrolysable ATP analog AMP-PNP or the ATPase inhibitor BeFx significantly reduced the stimulatory effect of stretch. We employed a point mutagenic approach to determine that a single residue (K1337) in the hairpin loop proximal to the major MgATPase catalytic site in the SUR2A subunit is responsible for the difference in mechano-sensitivity between SUR2A and SUR1 containing KATP channels. Moreover, using a double cysteine mutant substitution in the hairpin loop region revealed the importance of a key residue-residue interaction in this region that transduces membrane mechanical forces into KATP channel stimulation via increases in channel MgATPase activity. With respect to KATP channel pharmacology, glibenclamide, but not glicalizide or repaglinide, was able to completely inhibit KATP channel mechano-sensitivity. In summary, our results provide a highly plausible molecular mechanism by which mechanical membrane forces are rapidly converted in changes in KATP channel activity that have implications for our understanding of cardiac KATP channels in physiological or pathophysiological settings that involve increased workload.

The Oral ß-Lactamase SYN-004 (Ribaxamase) Degrades Ceftriaxone Excreted into the Intestine in Phase 2a Clinical Studies.

SYN-004 (ribaxamase) is a ß-lactamase designed to be orally administered concurrently with intravenous ß-lactam antibiotics, including most penicillins and cephalosporins. Ribaxamase's anticipated mechanism of action is to degrade excess ß-lactam antibiotic that is excreted into the small intestine. This enzymatic inactivation of excreted antibiotic is expected to protect the gut microbiome from disruption and thus prevent undesirable side effects, including secondary infections such as Clostridium difficile infections, as well as other antibiotic-associated diarrheas. In phase 1 clinical studies, ribaxamase was well tolerated compared to a placebo group and displayed negligible systemic absorption. The two phase 2a clinical studies described here were performed to confirm the mechanism of action of ribaxamase, degradation of ß-lactam antibiotics in the human intestine, and were therefore conducted in subjects with functioning ileostomies to allow serial sampling of their intestinal chyme. Ribaxamase fully degraded ceftriaxone to below the level of quantitation in the intestines of all subjects in both studies. Coadministration of oral ribaxamase with intravenous ceftriaxone was also well tolerated, and the plasma pharmacokinetics of ceftriaxone were unchanged by ribaxamase administration. Since ribaxamase is formulated as a pH-dependent, delayed-release formulation, the activity of ribaxamase in the presence of the proton pump inhibitor esomeprazole was examined in the second study; coadministration of these drugs did not adversely affect ribaxamase's ability to degrade ceftriaxone excreted into the intestine. These studies have confirmed the in vivo mechanism of action of ribaxamase, degradation of ß-lactam antibiotics in the human intestine (registered at ClinicalTrials.gov under NCT02419001 and NCT02473640).

Late sodium current inhibition alone with ranolazine is sufficient to reduce ischemia- and cardiac glycoside-induced calcium overload and contractile dysfunction mediated by reverse-mode sodium/calcium exchange.

Excessive reverse-mode (RM) sodium/calcium exchanger 1.1 (NCX1.1) activity, resulting from intracellular sodium accumulation caused by reduced Na+/K+-ATPase activity, increased Na-H exchanger 1 activity. The induction of the voltage-gated sodium channel late current component (late INa), is a major pathway for intracellular calcium (Ca2+i) loading in cardiac ischemia-reperfusion (IR) injury and cardiac glycoside toxicity. Inhibition of late INa with the antianginal agent ranolazine is protective in models of IR injury and cardiac glycoside toxicity. However, whether inhibition of late INa alone is sufficient to provide maximal protection or additional inhibition of RM NCX1.1 provides further benefit remains to be determined conclusively. Therefore, the effects of ranolazine were compared with the INa inhibitor lidocaine in models of IR injury and ouabain toxicity, RM NCX1.1-mediated Ca2+ overload, and patch-clamp assays of RM NCX1.1 currents. Ranolazine and lidocaine (10 µM) similarly reduced Ca2+i overload and improved left ventricle work recovery in whole-heart models of IR injury or exposure to ouabain (80 µM). Ranolazine (10 µM), but not lidocaine (10 µM), reduced RM NCX1.1-mediated Ca2+i overload in ventricular myocytes. Furthermore, ranolazine inhibited RM NCX1.1 currents (IC50 1.7 µM), without affecting forward mode currents, revealing that ranolazine has novel RM NCX1.1 inhibitory actions. However, because lidocaine provides similar protection to ranolazine in whole-heart models but does not inhibit RM NCX1.1, we conclude that induction of late INa is upstream of RM NCX1.1 activity and selective inhibition of late INa alone is sufficient to reduce Ca2+i overload and contractile dysfunction in IR injury and cardiac glycoside toxicity.

Preventing progression to chronicity in first onset, subacute low back pain: an exploratory study.

OBJECTIVES: To evaluate the effects of a behavioral medicine intervention, relative to an attention control, in preventing chronic pain and disability in patients with first-onset, subacute low back pain (LBP) with limitations in work-role function. DESIGN: A 2-group, experimental design with randomization to behavioral medicine or attention control groups. SETTING: Orthopedic clinic at a Naval Medical Center. PARTICIPANTS: Sixty-seven participants with first-onset LBP of 6 to 10 weeks of duration and impairment in work function, of whom 50 completed all 4 therapy sessions and follow-up 6 months after pain onset. INTERVENTION: Four 1-hour individual treatment sessions of either behavioral medicine, focused on back function and pain education, self-management training, graded activity increases, fear reduction, and pain belief change; or attention control condition, focused on empathy, support, and reassurance. MAIN OUTCOME MEASURES: The primary outcome was proportion of participants classified as recovered, according to pre-established clinical cutoffs on standardized measures, signifying absence of chronic pain and disability at 6 months after pain onset. Secondary analyses were conducted on pain, disability, health status, and functional work category. Intervention credibility and pain belief manipulation checks were also evaluated. RESULTS: Chi square analyses comparing proportions recovered at 6 months after pain onset for behavioral medicine and attention control participants found relative rates of 52% versus 31% in the modified intent-to-treat sample (P=.09) and 54% versus 23% for those completing all 4 sessions and 6-month follow-up (P=.02). At 12 months, 79% of recovered and 68% of chronic pain participants still met criteria for their respective groups (P<.0001). Recovered participants also had higher rates of functional work status recovery at 12 months (recovered: 96% full duty and 4% light duty; chronic pain: 61% full duty, 18% light duty, and 21% medical discharge, respectively; P=.03). CONCLUSIONS: Early intervention using a behavioral medicine rehabilitation approach may enhance recovery and reduce chronic pain and disability in patients with first-onset, subacute LBP. Effects are stronger for participants attending all 4 sessions and the follow-up assessment.

2-Aminoadipic acid is a biomarker for diabetes risk.

Improvements in metabolite-profiling techniques are providing increased breadth of coverage of the human metabolome and may highlight biomarkers and pathways in common diseases such as diabetes. Using a metabolomics platform that analyzes intermediary organic acids, purines, pyrimidines, and other compounds, we performed a nested case-control study of 188 individuals who developed diabetes and 188 propensity-matched controls from 2,422 normoglycemic participants followed for 12 years in the Framingham Heart Study. The metabolite 2-aminoadipic acid (2-AAA) was most strongly associated with the risk of developing diabetes. Individuals with 2-AAA concentrations in the top quartile had greater than a 4-fold risk of developing diabetes. Levels of 2-AAA were not well correlated with other metabolite biomarkers of diabetes, such as branched chain amino acids and aromatic amino acids, suggesting they report on a distinct pathophysiological pathway. In experimental studies, administration of 2-AAA lowered fasting plasma glucose levels in mice fed both standard chow and high-fat diets. Further, 2-AAA treatment enhanced insulin secretion from a pancreatic ß cell line as well as murine and human islets. These data highlight a metabolite not previously associated with diabetes risk that is increased up to 12 years before the onset of overt disease. Our findings suggest that 2-AAA is a marker of diabetes risk and a potential modulator of glucose homeostasis in humans.

The ATP-sensitive K(+) channel ABCC8 S1369A type 2 diabetes risk variant increases MgATPase activity.

Pancreatic ß-cell ATP-sensitive K(+) (K(ATP)) channels are composed of Kir6.2 and SUR1 subunits encoded by the KCNJ11 and ABCC8 genes, respectively. Although rare monogenic activating mutations in these genes cause overt neonatal diabetes, the common variants E23K (KCNJ11) and S1369A (ABCC8) form a tightly heritable haplotype that is associated with an increased susceptibility to type 2 diabetes (T2D) risk. However, the molecular mechanism(s) underlying this risk remain to be elucidated. A homology model of the SUR1 nucleotide-binding domains (NBDs) indicates that residue 1369 is in close proximity to the major MgATPase site. Therefore, we investigated the intrinsic MgATPase activity of K(ATP) channels containing these variants. Electrophysiological and biochemical techniques were used to study the MgATPase activity of recombinant human K(ATP) channels or glutathione S-transferase and NBD2 fusion proteins containing the E23/S1369 (nonrisk) or K23/A1369 (risk) variant haplotypes. K(ATP) channels containing the K23/A1369 haplotype displayed a significantly increased stimulation by guanosine triphosphate compared with the E23/S1369 haplotype (3.2- vs. 1.8-fold). This effect was dependent on the presence of the A1369 variant and was lost in the absence of Mg(2+) ions or in the presence of the MgATPase inhibitor beryllium fluoride. Direct biochemical assays also confirmed an increase in MgATPase activity in NBD2 fusion proteins containing the A1369 variant. Our findings demonstrate that the A1369 variant increases K(ATP) channel MgATPase activity, providing a plausible molecular mechanism by which the K23/A1369 haplotype increases susceptibility to T2D in humans homozygous for these variants.

Fatigue varies by social class in African Americans but not Caucasian Americans.

Socioeconomic status explains many ethnic disparities in health; however, mechanisms are hard to identify. Fatigue-a frequent complaint in patients and normals-is associated with poorer quality of life. We wondered if ethnicity and social class interact to explain fatigue. A total of 40 African Americans (AAs) and 64 Caucasian Americans (CAs) completed short forms of the Profile of Mood States (POMS-SF) and Multidimensional Fatigue Symptom Inventory (MFSI-SF). Participants were divided into high-middle and low social class groups (as per Hollingshead, 1958a). After controlling for gender, body mass index, depressive symptoms, and response bias, ethnicity and social class interacted for POMS-SF fatigue. AAs in the high-middle classes reported more fatigue than AAs in the low classes and CAs in the high-middle classes. Fatigue did not differ by class for CAs nor by ethnicity in the lower classes. Similar findings emerged for MFSI-SF general fatigue. Social class is important for understanding fatigue in AAs but not CAs.