Nora Eccles Harrison Cardiovascular Research and Training Institute

Salt Lake City, UT, United States

Nora Eccles Harrison Cardiovascular Research and Training Institute

Salt Lake City, UT, United States
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Cho S.,University of Utah | Cho S.,Nora Eccles Harrison Cardiovascular Research and Training Institute | Dong S.,University of Utah | Parent K.N.,Michigan State University | Chen M.,University of Utah
Journal of Drug Targeting | Year: 2016

Background: Cytotoxic T lymphocyte (CTL) vaccine carriers are known to enhance the efficacy of vaccines, but a search for more effective carriers is warranted. Elastin-like polypeptides (ELPs) have been examined for many medical applications but not as CTL vaccine carriers. Purpose: We aimed to create immune tolerant ELPs using a new polypeptide engineering practice and create CTL vaccine carriers using the ELPs. Results: Four sets of novel ELPs, termed immune-tolerant elastin-like polypeptide (iTEP) were generated according to the principles dictating humoral immunogenicity of polypeptides and phase transition property of ELPs. The iTEPs were non-immunogenic in mice. Their phase transition feature was confirmed through a turbidity assay. An iTEP nanoparticle (NP) was assembled from an amphiphilic iTEP copolymer plus a CTL peptide vaccine, SIINFEKL. The NP facilitated the presentation of the vaccine by dendritic cells (DCs) and enhanced vaccine-induced CTL responses. Discussion: A new ELP design and development practice was established. The non-canonical motif and the immune tolerant nature of the iTEPs broaden our insights about ELPs. ELPs, for the first time, were successfully used as carriers for CTL vaccines. Conclusion: It is feasible to concurrently engineer both immune-tolerant and functional peptide materials. ELPs are a promising type of CTL vaccine carriers. © 2015 Taylor & Francis.


PubMed | Oxford Genetics, University of Ulsan and Nora Eccles Harrison Cardiovascular Research and Training Institute
Type: Journal Article | Journal: The Journal of physiology | Year: 2014

Cellular processes are exquisitely sensitive to H+ and Ca2+ ions because of powerful ionic interactions with proteins. By regulating the spatial and temporal distribution of intracellular [Ca2+] and [H+], cells such as cardiac myocytes can exercise control over their biological function. A well-established paradigm in cellular physiology is that ion concentrations are regulated by specialized, membrane-embedded transporter proteins. Many of these couple the movement of two or more ionic species per transport cycle, thereby linking ion concentrations among neighbouring compartments. Here, we compare and contrast canonical membrane transport with a novel type of Ca(2+)-H+ coupling within cytoplasm, which produces uphill Ca2+ transport energized by spatial H+ ion gradients, and can result in the cytoplasmic compartmentalization of Ca2+ without requiring a partitioning membrane. The mechanism, demonstrated in mammalian myocytes, relies on diffusible cytoplasmic buffers, such as carnosine, homocarnosine and ATP, to which Ca2+ and H+ ions bind in an apparently competitive manner. These buffer molecules can actively recruit Ca2+ to acidic microdomains, in exchange for the movement of H+ ions. The resulting Ca2+ microdomains thus have the potential to regulate function locally. Spatial cytoplasmic Ca(2+)-H+ exchange (cCHX) acts like a pump without a membrane and may be operational in many cell types.


Swietach P.,Oxford Genetics | Leem C.-H.,University of Ulsan | Spitzer K.W.,Nora Eccles Harrison Cardiovascular Research and Training Institute | Vaughan-Jones R.D.,Oxford Genetics
Journal of Physiology | Year: 2014

Cellular processes are exquisitely sensitive to H+ and Ca2+ ions because of powerful ionic interactions with proteins. By regulating the spatial and temporal distribution of intracellular [Ca2+] and [H+], cells such as cardiac myocytes can exercise control over their biological function. A well-established paradigm in cellular physiology is that ion concentrations are regulated by specialized, membrane-embedded transporter proteins. Many of these couple the movement of two or more ionic species per transport cycle, thereby linking ion concentrations among neighbouring compartments. Here, we compare and contrast canonical membrane transport with a novel type of Ca2+-H+ coupling within cytoplasm, which produces uphill Ca2+ transport energized by spatial H+ ion gradients, and can result in the cytoplasmic compartmentalization of Ca2+ without requiring a partitioning membrane. The mechanism, demonstrated in mammalian myocytes, relies on diffusible cytoplasmic buffers, such as carnosine, homocarnosine and ATP, to which Ca2+ and H+ ions bind in an apparently competitive manner. These buffer molecules can actively recruit Ca2+ to acidic microdomains, in exchange for the movement of H+ ions. The resulting Ca2+ microdomains thus have the potential to regulate function locally. Spatial cytoplasmic Ca2+-H+ exchange (cCHX) acts like a 'pump' without a membrane and may be operational in many cell types. © 2014 The Physiological Society.


Wu W.,Nora Eccles Harrison Cardiovascular Research and Training Institute | Gardner A.,Nora Eccles Harrison Cardiovascular Research and Training Institute | Sanguinetti M.C.,University of Utah
Journal of Physiology | Year: 2014

At depolarized membrane potentials, the conductance of some voltage-gated K+ channels is reduced by C-type inactivation. This gating process is voltage independent in Kv1 and involves a conformational change in the selectivity filter that is mediated by cooperative subunit interactions. C-type inactivation in hERG1 K+ channels is voltage-dependent, much faster in onset and greatly attenuates currents at positive potentials. Here we investigate the potential role of subunit interactions in C-type inactivation of hERG1 channels. Point mutations in hERG1 known to eliminate (G628C/S631C), inhibit (S620T or S631A) or enhance (T618A or M645C) C-type inactivation were introduced into subunits that were combined with wild-type subunits to form concatenated tetrameric channels with defined subunit composition and stoichiometry. Channels were heterologously expressed in Xenopus oocytes and the two-microelectrode voltage clamp was used to measure the kinetics and steady-state properties of inactivation of whole cell currents. The effect of S631A or T618A mutations on inactivation was a graded function of the number of mutant subunits within a concatenated tetramer as predicted by a sequential model of cooperative subunit interactions, whereas M645C subunits increased the rate of inactivation of concatemers, as predicted for subunits that act independently of one another. For mutations located within the inactivation gate proper (S620T or G628C/S631C), the presence of a single subunit in a concatenated hERG1 tetramer disrupted gating to the same extent as that observed for mutant homotetramers. Together, our findings indicate that the final step of C-type inactivation of hERG1 channels involves a concerted, all-or-none cooperative interaction between all four subunits, and that probing the mechanisms of channel gating with concatenated heterotypic channels should be interpreted with care, as conclusions regarding the nature of subunit interactions may depend on the specific mutation used to probe the gating process. © 2014 The Physiological Society.


Swietach P.,Oxford Genetics | Spitzer K.W.,Nora Eccles Harrison Cardiovascular Research and Training Institute | Vaughan-Jones R.D.,Oxford Genetics
Cardiovascular Research | Year: 2015

Aims Contraction of the heart is regulated by electrically evoked Ca2+ transients (CaTs). H+ ions, the end products of metabolism, modulate CaTs through direct interactions with Ca2+-handling proteins and via Na+-mediated coupling between acid-extruding proteins (e.g. Na+/H+ exchange, NHE1) and Na+/Ca2+ exchange. Restricted H+ diffusivity in cytoplasm predisposes pH-sensitive Ca2+ signalling to becoming non-uniform, but the involvement of readily diffusible intracellular Na+ ions may provide a means for combatting this. Methods and results CaTs were imaged in fluo3-loaded rat ventricular myocytes paced at 2 Hz. Cytoplasmic [Na+] ([Na+]i) was imaged using SBFI. Intracellular acidification by acetate exposure raised diastolic and systolic [Ca2+] (also observed with acid-loading by ammonium prepulse or CO2 exposure). The systolic [Ca2+] response correlated with a rise in [Na+]i and sarcoplasmic reticulum Ca2+ load, and was blocked by the NHE1 inhibitor cariporide (CO2/HCO3 --free media). Exposure of one half of a myocyte to acetate using dual microperfusion (CO2/HCO3 --free media) raised diastolic [Ca2+] locally in the acidified region. Systolic [Ca2+] and CaT amplitude increased more uniformly along the length of the cell, but only when NHE1 was functional. Cytoplasmic Na+ diffusivity (DNa) was measured in quiescent cells, with strophanthidin present to inhibit the Na+/K+ pump. With regional acetate exposure to activate a local NHE-driven Na+-influx, DNa was found to be sufficiently fast (680 μm2/s) for transmitting the pH-systolic Ca2+ interaction over long distances. Conclusions Na+ ions are rapidly diffusible messengers that expand the spatial scale of cytoplasmic pH-CaT interactions, helping to co-ordinate global Ca2+ signalling during conditions of intracellular pH non-uniformity. © 2014 The Author.


Jou C.J.,Nora Eccles Harrison Cardiovascular Research and Training Institute | Barnett S.M.,Nora Eccles Harrison Cardiovascular Research and Training Institute | Bian J.-T.,Nora Eccles Harrison Cardiovascular Research and Training Institute | Weng H.C.,Nora Eccles Harrison Cardiovascular Research and Training Institute | And 2 more authors.
Circulation Research | Year: 2013

RATIONALE:: Genetic testing for Long QT Syndrome is now a standard and integral component of clinical cardiology. A major obstacle to the interpretation of genetic findings is the lack of robust functional assays to determine the pathogenicity of identified gene variants in a high-Throughput manner. OBJECTIVE:: The goal of this study was to design and test a high-Throughput in vivo cardiac assay to distinguish between disease-causing and benign KCNH2 (hERG1) variants, using the zebrafish as a model organism. METHODS AND RESULTS:: We tested the ability of previously characterized Long QT Syndrome hERG1 mutations and polymorphisms to restore normal repolarization in the kcnh2-knockdown embryonic zebrafish. The cardiac assay correctly identified a benign variant in 9 of 10 cases (negative predictive value 90%), whereas correctly identifying a disease-causing variant in 39/39 cases (positive predictive value 100%). CONCLUSIONS:: The in vivo zebrafish cardiac assay approaches the accuracy of the current benchmark in vitro assay for the detection of disease-causing mutations, and is far superior in terms of throughput rate. Together with emerging algorithms for interpreting a positive long QT syndrome genetic test, the zebrafish cardiac assay provides an additional tool for the final determination of pathogenicity of gene variants identified in long QT syndrome genetic screening. © 2013 American Heart Association, Inc.


Swenson D.J.,University of Utah | Swenson D.J.,Scientific Computing and Imaging Institute | Geneser S.E.,Scientific Computing and Imaging Institute | Stinstra J.G.,Scientific Computing and Imaging Institute | And 5 more authors.
Annals of Biomedical Engineering | Year: 2011

The electrocardiogram (ECG) is ubiquitously employed as a diagnostic and monitoring tool for patients experiencing cardiac distress and/or disease. It is widely known that changes in heart position resulting from, for example, posture of the patient (sitting, standing, lying) and respiration significantly affect the body-surface potentials; however, few studies have quantitatively and systematically evaluated the effects of heart displacement on the ECG. The goal of this study was to evaluate the impact of positional changes of the heart on the ECG in the specific clinical setting of myocardial ischemia. To carry out the necessary comprehensive sensitivity analysis, we applied a relatively novel and highly efficient statistical approach, the generalized polynomial chaos-stochastic collocation method, to a boundary element formulation of the electrocardiographic forward problem, and we drove these simulations with measured epicardial potentials from whole-heart experiments. Results of the analysis identified regions on the body-surface where the potentials were especially sensitive to realistic heart motion. The standard deviation (STD) of ST-segment voltage changes caused by the apex of a normal heart, swinging forward and backward or side-to-side was approximately 0.2 mV. Variations were even larger, 0.3 mV, for a heart exhibiting elevated ischemic potentials. These variations could be large enough to mask or to mimic signs of ischemia in the ECG. Our results suggest possible modifications to ECG protocols that could reduce the diagnostic error related to postural changes in patients possibly suffering from myocardial ischemia. © 2011 Biomedical Engineering Society.


PubMed | Oxford Genetics and Nora Eccles Harrison Cardiovascular Research and Training Institute
Type: Journal Article | Journal: Cardiovascular research | Year: 2015

Contraction of the heart is regulated by electrically evoked Ca(2+) transients (CaTs). H(+) ions, the end products of metabolism, modulate CaTs through direct interactions with Ca(2+)-handling proteins and via Na(+)-mediated coupling between acid-extruding proteins (e.g. Na(+)/H(+) exchange, NHE1) and Na(+)/Ca(2+) exchange. Restricted H(+) diffusivity in cytoplasm predisposes pH-sensitive Ca(2+) signalling to becoming non-uniform, but the involvement of readily diffusible intracellular Na(+) ions may provide a means for combatting this.CaTs were imaged in fluo3-loaded rat ventricular myocytes paced at 2 Hz. Cytoplasmic [Na(+)] ([Na(+)]i) was imaged using SBFI. Intracellular acidification by acetate exposure raised diastolic and systolic [Ca(2+)] (also observed with acid-loading by ammonium prepulse or CO exposure). The systolic [Ca(2+)] response correlated with a rise in [Na(+)]i and sarcoplasmic reticulum Ca(2+) load, and was blocked by the NHE1 inhibitor cariporide (CO/HCO(-)-free media). Exposure of one half of a myocyte to acetate using dual microperfusion (CO/HCO(-)-free media) raised diastolic [Ca(2+)] locally in the acidified region. Systolic [Ca(2+)] and CaT amplitude increased more uniformly along the length of the cell, but only when NHE1 was functional. Cytoplasmic Na(+) diffusivity (DNa) was measured in quiescent cells, with strophanthidin present to inhibit the Na(+)/K(+) pump. With regional acetate exposure to activate a local NHE-driven Na(+)-influx, DNa was found to be sufficiently fast (680 m(2)/s) for transmitting the pH-systolic Ca(2+) interaction over long distances.Na(+) ions are rapidly diffusible messengers that expand the spatial scale of cytoplasmic pH-CaT interactions, helping to co-ordinate global Ca(2+) signalling during conditions of intracellular pH non-uniformity.


PubMed | Nora Eccles Harrison Cardiovascular Research and Training Institute
Type: Journal Article | Journal: American journal of physiology. Heart and circulatory physiology | Year: 2011

Sodium-hydrogen exchanger (NHE), the principal sarcolemmal acid extruder in ventricular myocytes, is stimulated by a variety of autocrine/paracrine factors and contributes to myocardial injury and arrhythmias during ischemia-reperfusion. Platelet-activating factor (PAF; 1-o-alkyl-2-acetyl-sn-glycero-3-phosphocholine) is a potent proinflammatory phospholipid that is released in the heart in response to oxidative stress and promotes myocardial ischemia-reperfusion injury. PAF stimulates NHE in neutrophils and platelets, but its effect on cardiac NHE (NHE1) is unresolved. We utilized quiescent guinea pig ventricular myocytes bathed in bicarbonate-free solutions and epifluorescence to measure intracellular pH (pH(i)). Methylcarbamyl-PAF (C-PAF; 200 nM), a metabolically stable analog of PAF, significantly increased steady-state pH(i). The alkalosis was completely blocked by the NHE inhibitor, cariporide, and by sodium-free bathing solutions, indicating it was mediated by NHE activation. C-PAF also significantly increased the rate of acid extrusion induced by intracellular acidosis. The ability of C-PAF to increase steady-state pH(i) was completely blocked by the PAF receptor inhibitor WEB 2086 (10 M), indicating the PAF receptor is required. A MEK inhibitor (PD98059; 25 M) also completely blocked the rise in pH(i) induced by C-PAF, suggesting participation of the MAP kinase signaling cascade downstream of the PAF receptor. Inhibition of PKC with GF109203X (1 M) and chelerythrine (2 M) did not significantly affect the alkalosis induced by C-PAF. In summary, these results provide evidence that PAF stimulates cardiac NHE1, the effect occurs via the PAF receptor, and signal relay requires participation of the MAP kinase cascade.


PubMed | Nora Eccles Harrison Cardiovascular Research and Training Institute
Type: Journal Article | Journal: Pacing and clinical electrophysiology : PACE | Year: 2011

Ectopy-induced cardiomyopathy is an increasingly recognized cause of reversible left ventricular (LV) dysfunction. The underlying mechanisms remain unknown. Our goal was to create an animal model for ectopy-induced cardiomyopathy.Eleven mongrel dogs underwent the implantation of a dual-chamber pacemaker. Four dogs served as the control group and seven as the paced group. In the paced group, the pacemaker was connected to two endocardial right ventricular leads, one inserted into the atrial port and the other one into the ventricular port with an atrioventricular delay adjusted to ensure the presence of coupled pacing simulating ventricular bigeminy. Echocardiographic measurements of LV size (LV end-diastolic diameter [LV-EDD], LV end-systolic diameter [LV-ESD]), LV ejection fraction (LVEF), and mitral regurgitation (MR) were obtained at baseline and after 4 weeks of monitoring or pacing in all dogs except one who had lead dislodgement.In the control group (n = 4), no significant changes in LV dimensions or function were noted. In the paced group (n = 6), LV-EDD and LV-ESD increased from 3.58 0.65 cm and 2.47 0.55 cm to 4.15 0.59 cm and 3.21 0.47 cm, respectively (P < 0.01). In addition, LVEF decreased from 60 7% to 46 9% (P < 0.05). No changes in MR were noted.We have shown that coupled pacing simulating ventricular bigeminy was feasible and resulted in increased LV dimensions and decreased LV function. By controlling the percentage of pacing, the coupling interval and the location of the pacing lead, this new model will allow the assessment of the relative roles of these variables in the development of ectopy-induced cardiomyopathy.

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