An unknown endogenous inhibitor of Na/Ca exchange can enhance the cardiac muscle contractility

The cardiac sarcolemma Na/Ca exchanger is a key system for controlling the intracellular calcium levels during the excitation-contraction coupling. Here, we test the hypothesis that the heart tissue contains a putative endogenous factor having a capacity to modulate the Na/Ca exchanger and muscle contractility. The concentrated cardiac extracts inhibit the Na(i)- or Ca(i)-dependent (45)Ca uptakes in isolated cardiac sarcolemma vesicles as well as the Na(o)-dependent Ca efflux, monitored by extravesicular Ca probe fluo-3. The inhibitory activity has been purified approximately 2000-fold by normal and reversed-phase HPLC procedures. The inhibitory activity is eluted from the Sephadex G-10 in the range of 350-550 Da, suggesting that the inhibitory factor is a low-molecular-weight substance. The mass spectra analysis shows a number of signals within m/z 380-560; however, it is not clear at this moment whether these recordings represent the mass of putative inhibitory factor or irrelevant impurities. The endogenous inhibitory factor of Na/Ca exchange does not resemble the properties (HPLC retention time, mass spectra, amino acid analysis, etc.) of autoinhibitory XIP peptide. The addition of inhibitory factor to muscle strip of guinea pig ventricles induces 2- to 5-fold enhancement of isometric contractions, thereby exhibiting a strong positive inotropic effect. This effect is a dose-dependent phenomenon, which can be reversed by washing the inhibitory factor from the organ bath. Assuming a molecular weight of 350-550 Da, the effective concentrations of putative inhibitor must be <10(-6) M. Therefore, the present findings demonstrate that the mammalian heart contains a low-molecular-weight factor that can inhibit Na/Ca exchange and enhance the cardiac contractility.     

Continuum rheology of muscle contraction and its application to cardiac contractility.

A set of constitutive equations is proposed to describe the mechanics of contraction of skeletal and heart muscle. Fiber tension is assumed to depend on the degree of chemical activation, the stretch ratio, and the rate of stretching of the fibers. The time rate of change of activation is governed by a differential equation. The proposed constitutive equations are used to model the time courses of isotonic and isometric twitches during contraction and relaxation phases of the muscle response to stimulation. Various contractility indices of the left ventricle are considered next by using the proposed constitutive equations. The present analysis introduces a new interpretation of the index of contractility (dP/dt)/P used in cardiac literature. It is shown that this index may not be related at all to the maximum speed of shortening and that it may be dependent on both preload and afterload. The development of pressure during isovolumetric contraction of the left ventricle is shown to be governed by a differential equation describing the time rate of change of tension during isometric contraction of myocardium fibers.     

L-selectin and SDF-1 enhance the migration of mouse and human cardiac mesoangioblasts

Efficient delivery of stem cells to heart regions is still a major problem for cell therapy. Here, we report experiments aimed to improve migration of mouse and human cardiac mesoangioblasts to the damaged heart. Cardiac mesoangioblasts were induced to transmigrate through the endothelium by factors released by cardiomyocytes or cytokines, among which stromal-derived factor 1 (SDF-1) was the most potent. Cardiac mesoangioblasts were also delivered into the left ventricular (LV) chamber of mice after coronary artery ligation (CAL), and their in vivo homing to the damaged heart was found to be quite modest. Pretreatment of cardiac mesoangioblasts with SDF-1 or transient expression of L-selectin induced a two- to three-fold increase in their transmigration and homing to the damaged heart. Therefore, combined pretreatment with SDF-1 and L-selectin generated modified cardiac mesoangioblasts, 50% of which, after injection into the LV chamber of mice early after CAL, home directly to the damaged free wall of the heart. Finally, modified mouse cardiac mesoangioblasts, injected into the LV chamber regenerate a larger surface of the ventricle in long-term experiments in comparison with their control counterparts. This study defines the requirements for efficient homing of cardiac mesoangioblasts to the damaged heart and offers a new potent tool to optimize efficiency of future cell therapy protocols for cardiovascular diseases.     

Extracellular ATP has a potent effect to enhance cytosolic calcium and contractility in single ventricular myocytes

The effect of extracellular ATP on the contraction of single rat cardiac myocytes was investigated, together with the effect on the transient change in cytosolic Ca2+ (Cai) elicited by excitation and on the relationship between these two parameters. In unstimulated single myocytes, ATP caused a small increase in Cai (measured as the ratio of fluorescence of Indo-1 at 410 to that at 490 nm. In myocytes bathed in a medium containing 1.0 mM [Ca2+] at 23 degrees C and stimulated at 1 Hz, ATP (1 microM) resulted in a two-threefold increase in amplitude of contraction, as measured by video cinemicrographic techniques. The duration of the Cai-transient was not altered but its amplitude was markedly enhanced, as was the amplitude of contraction. The relation between Cai and contraction-amplitude was not altered by ATP, when measured over a range of extracellular [Ca2+], suggesting that ATP does not affect the myofilament-Ca2+ interaction. The primary site of action of ATP in increasing Cai is at the sarcolemma since the addition to suspensions of myocytes of caffeine (10 mM), which depletes the sarcoplasmic reticulum Ca2+ load, does not prevent the subsequent increase of Cai due to ATP. Further, lowering of the extracellular [Ca2+] to less than 1 microM with EGTA abolishes the response of Cai to ATP, though not the response to caffeine. Thus in rat cardiac myocytes ATP stimulates trans-sarcolemmal influx of Ca2+: ADP, AMP and adenosine are ineffective. ATP markedly augments the amplitude of the Cai transient elicited by electrical stimulation thus rendering it a potent inotropic agent.     

Application of low-intensity ultrasound to enhance simultaneous nitrification/iron-based autotrophic denitrification

Biotechnology Letters - Intermittent ultrasound with an intensity of 0.2&nbsp;W/ml was applied during simultaneous nitrification/iron-based autotrophic denitrification to evaluate its impacts...     

Impaired cardiac contractility response to hemodynamic stress in S100A1-deficient mice

Ca(2+) signaling plays a central role in cardiac contractility and adaptation to increased hemodynamic demand. We have generated mice with a targeted deletion of the S100A1 gene coding for the major cardiac isoform of the large multigenic S100 family of EF hand Ca(2+)-binding proteins. S100A1(-/-) mice have normal cardiac function under baseline conditions but have significantly reduced contraction rate and relaxation rate responses to beta-adrenergic stimulation that are associated with a reduced Ca(2+) sensitivity. In S100A1(-/-) mice, basal left-ventricular contractility deteriorated following 3-week pressure overload by thoracic aorta constriction despite a normal adaptive hypertrophy. Surprisingly, heterozygotes also had an impaired response to acute beta-adrenergic stimulation but maintained normal contractility in response to chronic pressure overload that coincided with S100A1 upregulation to wild-type levels. In contrast to other genetic models with impaired cardiac contractility, loss of S100A1 did not lead to cardiac hypertrophy or dilation in aged mice. The data demonstrate that high S100A1 protein levels are essential for the cardiac reserve and adaptation to acute and chronic hemodynamic stress in vivo.     

Changes in cardiac contractility related to calcium-mediated changes in phosphorylation of myosin-binding protein C.

Ca ions can influence the contraction of cardiac muscle by activating kinases that specifically phosphorylate the myofibrillar proteins myosin-binding protein C (MyBP-C) and the regulatory light chain of myosin (RLC). To investigate the possible role of Ca-regulated phosphorylation of MyBP-C on contraction, isolated quiescent and rhythmically contracting cardiac trabeculae were exposed to different concentrations of extracellular Ca and then chemically skinned to clamp the contractile system. Maximum Ca-activated force (F(max)) was measured in quiescent cells soaking in 1) 2.5 mM Ca for 120 min, 2) 1.25 mM for 120 min, or 3) 1.25 mM for 120 min followed by 10 min in 7.5 mM, and 4) cells rhythmically contracting in 2.5 mM for 20 min. F(max) was, respectively, 21.5, 10.5, 24.7, and 32.6 mN/mm(2). Changes in F(max) were closely associated with changes in the degree of phosphorylation of MyBP-C and occurred at intracellular concentrations of Ca below levels associated with phosphorylation of RLC. Monophosphorylation of MyBP-C by a Ca-regulated kinase is necessary before beta-adrenergic stimulation can produce additional phosphorylation. These results suggest that Ca-dependent phosphorylation of MyBP-C modulates contractility by changing thick filament structure.     

The human anger face evolved to enhance cues of strength

Animals typically deploy their morphology during conflict to enhance competitors' assessments of their fighting ability (e.g. bared fangs, piloerection, dewlap inflation). Recent research has shown that humans assess others' fighting ability by monitoring cues of strength, and that the face itself contains such cues. We propose that the muscle movements that constitute the human facial expression of anger were selected because they increased others' assessments of the angry individual's strength, thereby increasing bargaining power. This runs contrary to the traditional theory that the anger face is an arbitrary set of features that evolved simply to signal aggressive intent. To test between these theories, the seven key muscle movements constituting the anger face were systematically manipulated one by one and in the absence of the others. Raters assessed faces containing any one of these muscle movements as physically stronger, supporting the hypothesis that the anger face evolved to enhance cues of strength.     

The contribution of Kv7 channels to pregnant mouse and human myometrial contractility

Premature birth accounts for approximately 75% of neonatal mortality and morbidity in the developed world. Despite this, methods for identifying and treating women at risk of preterm labour are limited and many women still present in preterm labour requiring tocolytic therapy to suppress uterine contractility. The aim of this study was to assess the utility of Kv7 channel activators as potential uterine smooth muscle (myometrium) relaxants in tissues from pregnant mice and women. Myometrium was obtained from early and late pregnant mice and from lipopolysaccharide (LPS)‐injected mice (day 15 of gestation; model of infection in pregnancy). Human myometrium was obtained at the time of Caesarean section from women at term (38–41 weeks). RT‐PCR/qRT‐PCR detected KCNQ and KCNE expression in mouse and human myometrium. In mice, there was a global suppression of all KCNQ isoforms, except KCNQ3, in early pregnancy (n= 6, P < 0.001 versus late pregnant); expression subsequently increased in late pregnancy (n= 6). KCNE isoforms were also gestationally regulated (P < 0.05). KCNQ and KCNE isoform expression was slightly down‐regulated in myometrium from LPS‐treated‐mice versus controls (P < 0.05, n= 3–4). XE991 (10 μM, Kv7 inhibitor) significantly increased spontaneous myometrial contractions in vitro in both human and mouse myometrial tissues (P < 0.05) and retigabine/flupirtine (20 μM, Kv7 channel activators) caused profound myometrial relaxation (P < 0.05). In summary, Kv7 activators suppressed myometrial contraction and KCNQ gene expression was sustained throughout gestation, particularly at term. Consequently, activation of the encoded channels represents a novel mechanism for treatment of preterm labour.     

Autoantibodies enhance agonist action and binding to cardiac muscarinic receptors in chronic Chagas' disease

Chronic Chagasic patient immunoglobulins (CChP-IgGs) recognize an acidic amino acid cluster at the second extracellular loop (el2) of cardiac M(2)-muscarinic acetylcholine receptors (M(2)AChRs). These residues correspond to a common binding site for various allosteric agents. We characterized the nature of the M(2)AChR/CChP-IgG interaction in functional and radioligand binding experiments applying the same mainstream strategies previously used for the characterization of other allosteric agents. Dose-response curves of acetylcholine effect on heart rate were constructed with data from isolated heart experiments in the presence of CChP or normal blood donor (NBD) sera. In these experiments, CChP sera but not NBD sera increased the efficacy of agonist action by augmenting the onset of bradyarrhythmias and inducing a Hill slope of 2.5. This effect was blocked by gallamine, an M(2)AChR allosteric antagonist. Correspondingly, CChP-IgGs increased acetylcholine affinity twofold and showed negative cooperativity for [(3)H]-N-methyl scopolamine ([(3)H]-NMS) in allosterism binding assays. A peptide corresponding to the M(2)AChR-el2 blocked this effect. Furthermore, dissociation assays showed that the effect of gallamine on the [(3)H]-NMS off-rate was reverted by CChP-IgGs. Finally, concentration-effect curves for the allosteric delay of W84 on [(3)H]-NMS dissociation right shifted from an IC(50) of 33 nmol/L to 78 nmol/L, 992 nmol/L, and 1670 nmol/L in the presence of 6.7 x 10(- 8), 1.33 x 10(- 7), and 2.0 x 10(- 7) mol/L of anti-el2 affinity-purified CChP-IgGs. Taken together, these findings confirmed a competitive interplay of these ligands at the common allosteric site and revealed the novel allosteric nature of the interaction of CChP-IgGs at the M(2)AChRs as a positive cooperativity effect on acetylcholine action.     

Corticosteroids enhance the binding of recombinant interferon-gamma to cultured human monocytes

Human peripheral blood monocytes possess receptors on their surface for human interferon-gamma (IFN-gamma). Inasmuch as the biologic effects of IFN-gamma are thought to be mediated through binding of the molecule to this receptor, modulation of receptor number may be a means for enhancing the effect of IFN-gamma. We examined this possibility by treating human monocytes in culture with dexamethasone. At a dexamethasone concentration of 200 nM, there was a twofold increase in the number of 125I-recombinant IFN-gamma molecules bound to the cell. An effect was noted at concentrations of dexamethasone greater than 50 nM, and was observed as early as after 18 hr of treatment. The observed effect was the result of both a 50% decrease in receptor number in untreated cells and an increase to pretreatment levels or greater in cells incubated with dexamethasone. These results suggest that dexamethasone may modulate the effects of IFN-gamma on monocytes by changes in receptor number.     

Specialized proresolving mediators enhance human B cell differentiation to antibody-secreting cells

The resolution of inflammation is an active and dynamic process critical in maintaining homeostasis. Newly identified lipid mediators have been recognized as key players during the resolution phase. These specialized proresolving mediators (SPM) constitute separate families that include lipoxins, resolvins, protectins, and maresins, each derived from essential polyunsaturated fatty acids. New results demonstrate that SPM regulate aspects of the immune response, including reduction of neutrophil infiltration, decreased T cell cytokine production, and stimulation of macrophage phagocytic activity. The actions of SPM on B lymphocytes remain unknown. Our study shows that the novel SPM 17-hydroxydosahexaenoic acid (17-HDHA), resolvin D1, and protectin D1 are present in the spleen. Interestingly, 17-HDHA and resolvin D1, but not protectin D1, strongly increase activated human B cell IgM and IgG production. Furthermore, increased Ab production by 17-HDHA is due to augmented B cell differentiation toward a CD27(+)CD38(+) Ab-secreting cell phenotype. The 17-HDHA did not affect proliferation and was nontoxic to cells. Increase of plasma cell differentiation and Ab production supports the involvement of SPM during the late stages of inflammation and pathogen clearance. The present study provides new evidence for SPM activity in the humoral response. These new findings highlight the potential applications of SPM as endogenous and nontoxic adjuvants, and as anti-inflammatory therapeutic molecules.     

Application of low intensity ultrasound to enhance the activity of anammox microbial consortium for nitrogen removal

In this study, effect of low intensity ultrasound on the activity of anammox microbial consortium for nitrogen removal was investigated through batch experiments at the same irradiation frequency of 25 kHz. Total nitrogen removal rate increased by about 25.5% when ultrasound intensity of 0.3 w cm⁻² was applied at an optimal irradiation time of 4 min, and further experiments demonstrated that this effect could last for about 6 days. Analysis of extracellular polymeric substances indicated that the maximum increase of carbohydrate, protein and total extracellular substances was obtained on the first day after ultrasound, which was 28.8%, 30.5% and 29.7%, respectively. As the time prolonged, the production rate of extracellular carbohydrate, protein decreased gradually. Transmission electron microscopy observation demonstrated that ultrasounded cell wall of anammox microbial consortium became thinner resulting in increased release of extracellular substances. The results suggested that application of low intensity ultrasound may enhance the activity of anammox microbial consortium and ultimately the potential for nitrogen removal.     

Exploiting zebrafish to help understand human cardiac arrhythmia

Commentary to:

Hassel D, Scholz EP, Trano N et al. Deficient zebrafish Ether-a-Go-Go-related gene channel gating causes short-QT syndrome in zebrafish reggae mutants. Circulation 2008; 117:866-75.     

Gamma interferon is able to enhance the oxidative metabolism of human neutrophils

The oxidative metabolism of human neutrophils has been studied after incubation of the cells with recombinant interferon-y. Neutrophils incubated for 2-4 hours with 2-50 U/ml recombinant interferon-y undergo a higher respiratory burst measured both as Oz consumption and Oz- production when stimulated with formyl-methionyl-leucyl-phenylalanine, Concanavalin A or phorbol myristate acetate. The potentiating effect of interferon-y requires more than one hour of incubation, is optimal at 20-50 U/ml and depends on the presence of serum in the incubation medium. The interferon effect depends on new protein synthesis. Cycloheximide at doses which do not alter the respiratory response of normal cells completely prevents the potentiating effect of interferon.     

Agonists trigger G protein-mediated activation of the CPI-17 inhibitor phosphoprotein of myosin light chain phosphatase to enhance vascular smooth muscle contractility

Myosin light chain phosphatase (MLCP) plays a pivotal role in smooth muscle contraction by regulating Ca(2+) sensitivity of myosin light chain phosphorylation. A smooth muscle phosphoprotein called CPI-17 specifically and potently inhibits MLCP in vitro and in situ and is activated when phosphorylated at Thr-38, which increases its inhibitory potency 1000-fold. We produced a phosphospecific antibody for this site in CPI-17 and used it to study in situ phosphorylation of endogenous CPI-17 in arterial smooth muscle in response to agonist stimulation. In the intact femoral artery, CPI-17 phosphorylation was negligible at the resting state and was not increased during contraction induced by K(+) depolarization. The Ca(2+)-sensitizing agonists histamine and phenylephrine induced nearly equivalent contractions, but histamine generated significantly higher levels of CPI-17 phosphorylation. In alpha-toxin-permeabilized strips at pCa 6.7, contractile force and CPI-17 phosphorylation were proportional in response to histamine, guanosine 5'-O-(gamma-thiotriphosphate), and histamine plus guanyl-5'-yl thiophosphate, implying that histamine increased CPI-17 phosphorylation through activation of G proteins. Inhibitors of Rho-kinase (Y27632) and protein kinase C (PKC; GF109203X) reduced contraction and CPI-17 phosphorylation in parallel, suggesting that CPI-17 functions downstream of Rho kinases and PKC. The results show that agonists such as histamine signal through phosphorylation of CPI-17 to produce Ca(2+) sensitization of smooth muscle contraction.     

Tumor necrosis factor-alpha increases airway smooth muscle oxidants production through a NADPH oxidase-like system to enhance myosin light chain phosphorylation and contractility

Tumor necrosis factor plays a critical role in airway smooth muscle hyperresponsiveness observed in asthma. However, the mechanisms underlying this phenomenon are poorly understood. We investigated if tumor necrosis factor-stimulated airway smooth muscle produced reactive oxygen species, leading to muscular hyperresponsiveness. Tumor necrosis factor increased intracellular and extracellular oxidants production in guinea pig airway smooth muscle cells and tissue homogenates. This production was abolished by inhibitors of NADPH oxidase (diphenylene iodinium or apocynin) and was enhanced by NADPH, whereas inhibitors of mitochondrial respiratory chain, nitric-oxide synthase, cyclooxygenase, and xanthine oxidase had no effect. NADPH oxidase subunits p22(phox) and p47(phox) were detected in smooth muscle cells and tissue homogenates by Western blot, immunohistochemistry, and spectral analysis. Furthermore, oxidants production was significantly reduced by transient transfection of smooth muscle cells with p22(phox) antisense oligonucleotides. Intracellular antioxidants and diphenylene iodinium abolished tumor necrosis factor-induced muscular hyperresponsiveness and increased in phosphorylation of the myosin light chain. Finally, NADPH oxidase subunits p22(phox) and p47(phox) were also detected in human airway smooth muscle. Collectively, these results demonstrate that tumor necrosis factor-stimulated airway smooth muscle produces oxidants through a NADPH oxidase-like system, which plays a pivotal role in muscle hyperresponsiveness and myosin light chain phosphorylation.     

Ability of human leukocytic pyrogen to enhance phytohemagglutinin induced murine thymocyte proliferation

Human leukocytic pyrogen, a monokine produced by stimulated human mononuclear phagocytes, will enhance the murine thymocyte proliferation response to phytohemagglutinin (lymphocyte activating factor (LAF) activity). During all steps of purification of human LP, pyrogenicity and LAF activity are coincidental suggesting a single identity for the two monokines. The LAF assay for human LP is highly sensitive and can detect human LP at a concentration of 10^?12M. Further experiments suggests that human LP and LAF activities could be destroyed by heating to 70 °C. Furthermore, while in vivo pyrogenicity of human LP can be blocked by ibuprofen, the in vitro LAF activity of the same molecule is unaffected by ibuprofen. Immune rabbit serum directed against human LP could also block in vitro LAF activity either by preincubation with LP or by blocking during culture.