Publications by authors named "Kenneth S Campbell"

79 Publications

Renal Angiotensinogen Is Predominantly Liver Derived in Nonhuman Primates.

Arterioscler Thromb Vasc Biol 2021 Sep 9:ATVBAHA121316590. Epub 2021 Sep 9.

Saha Cardiovascular Research Center, University of Kentucky, Lexington. (M.K., L.C., D.Y., H.S., M.K.F., P.I.H., A.D., R.E.T., H.S.L.).

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http://dx.doi.org/10.1161/ATVBAHA.121.316590DOI Listing
September 2021

Building biorepositories in the midst of a pandemic.

J Clin Transl Sci 2021 Feb 5;5(1):e92. Epub 2021 Feb 5.

Clinical & Translational Science Center, Caryl and Israel Englander Institute for Precision Medicine, Weill Cornell Medicine, New York, NY, USA.

Biospecimen repositories play a vital role in enabling investigation of biologic mechanisms, identification of disease-related biomarkers, advances in diagnostic assays, recognition of microbial evolution, and characterization of new therapeutic targets for intervention. They rely on the complex integration of scientific need, regulatory oversight, quality control in collection, processing and tracking, and linkage to robust phenotype information. The COVID-19 pandemic amplified many of these considerations and illuminated new challenges, all while academic health centers were trying to adapt to unprecedented clinical demands and heightened research constraints not witnessed in over 100 years. The outbreak demanded rapid understanding of SARS-CoV-2 to develop diagnostics and therapeutics, prompting the immediate need for access to high quality, well-characterized COVID-19-associated biospecimens. We surveyed 60 Clinical and Translational Science Award (CTSA) hubs to better understand the strategies and barriers encountered in biobanking before and in response to the COVID-19 pandemic. Feedback revealed a major shift in biorepository model, specimen-acquisition and consent process from a combination of investigator-initiated and institutional protocols to an enterprise-serving strategy. CTSA hubs were well equipped to leverage established capacities and expertise to quickly respond to the scientific needs of this crisis through support of institutional approaches in biorepository management.
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http://dx.doi.org/10.1017/cts.2021.6DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8134891PMC
February 2021

Mathematical modeling of myosin, muscle contraction, and movement.

Arch Biochem Biophys 2021 Jun 24:108979. Epub 2021 Jun 24.

Department of Physiology and Division of Cardiovascular Medicine, University of Kentucky, Lexington, KY, USA.

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http://dx.doi.org/10.1016/j.abb.2021.108979DOI Listing
June 2021

Hypertrophic cardiomyopathy β-cardiac myosin mutation (P710R) leads to hypercontractility by disrupting super relaxed state.

Proc Natl Acad Sci U S A 2021 Jun;118(24)

Department of Pediatrics (Cardiology), Stanford University School of Medicine, Palo Alto, CA 94304;

Hypertrophic cardiomyopathy (HCM) is the most common inherited form of heart disease, associated with over 1,000 mutations, many in β-cardiac myosin (MYH7). Molecular studies of myosin with different HCM mutations have revealed a diversity of effects on ATPase and load-sensitive rate of detachment from actin. It has been difficult to predict how such diverse molecular effects combine to influence forces at the cellular level and further influence cellular phenotypes. This study focused on the P710R mutation that dramatically decreased in vitro motility velocity and actin-activated ATPase, in contrast to other MYH7 mutations. Optical trap measurements of single myosin molecules revealed that this mutation reduced the step size of the myosin motor and the load sensitivity of the actin detachment rate. Conversely, this mutation destabilized the super relaxed state in longer, two-headed myosin constructs, freeing more heads to generate force. Micropatterned human induced pluripotent derived stem cell (hiPSC)-cardiomyocytes CRISPR-edited with the P710R mutation produced significantly increased force (measured by traction force microscopy) compared with isogenic control cells. The P710R mutation also caused cardiomyocyte hypertrophy and cytoskeletal remodeling as measured by immunostaining and electron microscopy. Cellular hypertrophy was prevented in the P710R cells by inhibition of ERK or Akt. Finally, we used a computational model that integrated the measured molecular changes to predict the measured traction forces. These results confirm a key role for regulation of the super relaxed state in driving hypercontractility in HCM with the P710R mutation and demonstrate the value of a multiscale approach in revealing key mechanisms of disease.
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http://dx.doi.org/10.1073/pnas.2025030118DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8214707PMC
June 2021

Impact of regulatory light chain mutation K104E on the ATPase and motor properties of cardiac myosin.

J Gen Physiol 2021 Jul;153(7)

Pennsylvania State University College of Medicine, Hershey, PA.

Mutations in the cardiac myosin regulatory light chain (RLC, MYL2 gene) are known to cause inherited cardiomyopathies with variable phenotypes. In this study, we investigated the impact of a mutation in the RLC (K104E) that is associated with hypertrophic cardiomyopathy (HCM). Previously in a mouse model of K104E, older animals were found to develop cardiac hypertrophy, fibrosis, and diastolic dysfunction, suggesting a slow development of HCM. However, variable penetrance of the mutation in human populations suggests that the impact of K104E may be subtle. Therefore, we generated human cardiac myosin subfragment-1 (M2β-S1) and exchanged on either the wild type (WT) or K104E human ventricular RLC in order to assess the impact of the mutation on the mechanochemical properties of cardiac myosin. The maximum actin-activated ATPase activity and actin sliding velocities in the in vitro motility assay were similar in M2β-S1 WT and K104E, as were the detachment kinetic parameters, including the rate of ATP-induced dissociation and the ADP release rate constant. We also examined the mechanical performance of α-cardiac myosin extracted from transgenic (Tg) mice expressing human wild type RLC (Tg WT) or mutant RLC (Tg K104E). We found that α-cardiac myosin from Tg K104E animals demonstrated enhanced actin sliding velocities in the motility assay compared with its Tg WT counterpart. Furthermore, the degree of incorporation of the mutant RLC into α-cardiac myosin in the transgenic animals was significantly reduced compared with wild type. Therefore, we conclude that the impact of the K104E mutation depends on either the length or the isoform of the myosin heavy chain backbone and that the mutation may disrupt RLC interactions with the myosin lever arm domain.
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http://dx.doi.org/10.1085/jgp.202012811DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8077168PMC
July 2021

Diverse and complex muscle spindle afferent firing properties emerge from multiscale muscle mechanics.

Elife 2020 12 28;9. Epub 2020 Dec 28.

Coulter Department of Biomedical Engineering, Emory University and Georgia Institute of Technology, Atlanta, United States.

Despite decades of research, we lack a mechanistic framework capable of predicting how movement-related signals are transformed into the diversity of muscle spindle afferent firing patterns observed experimentally, particularly in naturalistic behaviors. Here, a biophysical model demonstrates that well-known firing characteristics of mammalian muscle spindle Ia afferents - including movement history dependence, and nonlinear scaling with muscle stretch velocity - emerge from first principles of muscle contractile mechanics. Further, mechanical interactions of the muscle spindle with muscle-tendon dynamics reveal how motor commands to the muscle (alpha drive) versus muscle spindle (gamma drive) can cause highly variable and complex activity during active muscle contraction and muscle stretch that defy simple explanation. Depending on the neuromechanical conditions, the muscle spindle model output appears to 'encode' aspects of muscle force, yank, length, stiffness, velocity, and/or acceleration, providing an extendable, multiscale, biophysical framework for understanding and predicting proprioceptive sensory signals in health and disease.
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http://dx.doi.org/10.7554/eLife.55177DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7769569PMC
December 2020

Fast-relaxing cardiomyocytes exert a dominant role in the relaxation behavior of heterogeneous myocardium.

Arch Biochem Biophys 2021 01 30;697:108711. Epub 2020 Nov 30.

Department of Biomedical Engineering, Yale University, New Haven, CT, USA; Department of Cellular and Molecular Physiology, Yale School of Medicine, New Haven, CT, USA. Electronic address:

Substantial variation in relaxation rate exists among cardiomyocytes within small volumes of myocardium; however, it is unknown how this variability affects the overall relaxation mechanics of heart muscle. In this study, we sought to modulate levels of cellular heterogeneity in a computational model, then validate those predictions using an engineered heart tissue platform. We formulated an in silico tissue model composed of half-sarcomeres with varied relaxation rates, incorporating single-cell cardiomyocyte experimental data. These model tissues randomly sampled relaxation parameters from two offset distributions of fast- and slow-relaxing populations of half-sarcomeres. Isometric muscle twitch simulations predicted a complex relationship between relaxation time and the proportion of fast-versus slow-relaxing cells in heterogeneous tissues. Specifically, a 50/50 mixture of fast and slow cells did not lead to relaxation time that was the mean of the relaxation times associated with the two pure cases. Rather, the mean relaxation time was achieved at a ratio of 70:30 slow:fast relaxing cells, suggesting a disproportionate impact of fast-relaxing cells on overall tissue relaxation. To examine whether this behavior persists in vitro, we constructed engineered heart tissues from two lines of fast- and slow-relaxing human iPSC-derived cardiomyocytes. Cell tracking via fluorescent nanocrystals confirmed the presence of both cell populations in the 50/50 mixed tissues at the time of mechanical characterization. Isometric muscle twitch relaxation times of these mixed-population engineered heart tissues showed agreement with the predictions from the model, namely that the measured relaxation rate of 50/50 mixed tissues more closely resembled that of tissues made with 100% fast-relaxing cells. Our observations suggest that cardiomyocyte diversity can play an important role in determining tissue-level relaxation.
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http://dx.doi.org/10.1016/j.abb.2020.108711DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7785692PMC
January 2021

Titin-Truncating Mutations Associated With Dilated Cardiomyopathy Alter Length-Dependent Activation And Its Modulation Via Phosphorylation.

Cardiovasc Res 2020 Nov 2. Epub 2020 Nov 2.

National Heart and Lung Institute, Imperial College London, London, W12 0NN, UK.

Aims: Dilated cardiomyopathy (DCM) is associated with mutations in many genes encoding sarcomere proteins. Truncating mutations in the titin gene TTN are the most frequent. Proteomic and functional characterisations are required to elucidate the origin of the disease and the pathogenic mechanisms of TTN-truncating variants.

Methods And Results: We isolated myofibrils from DCM hearts carrying truncating TTN mutations and measured the Ca2+ sensitivity of force and its length dependence. Simultaneous measurement of force and adenosine triphosphate (ATP) consumption in skinned cardiomyocytes was also performed. Phosphorylation levels of troponin I (TnI) and myosin binding protein-C (MyBP-C) were manipulated using protein kinase A and λ phosphatase. mRNA sequencing was employed to overview gene expression profiles. We found that Ca2+ sensitivity of myofibrils carrying TTN mutations was significantly higher than in myofibrils from donor hearts. The length dependence of the Ca2+ sensitivity was absent in DCM myofibrils with TTN-truncating variants. No significant difference was found in the expression level of TTN mRNA between the DCM and donor groups. TTN exon usage and splicing were also similar. However, we identified downregulation of genes encoding Z-disk proteins, while the atrial-specific regulatory myosin light chain gene, MYL7, was upregulated in DCM patients with TTN-truncating variants.

Conclusion: Titin-truncating mutations lead to decreased length-dependent activation and increased elasticity of myofibrils. Phosphorylation levels of TnI and MyBP-C seen in the left ventricles are essential for the length-dependent changes in Ca2+ sensitivity in healthy donors, but they are reduced in DCM patients with TTN-truncating variants. A decrease in expression of Z-disk proteins may explain the observed decrease in myofibril passive stiffness and length-dependent activation.

Translational Perspective: Our findings may have implications in the development of new strategies for DCM treatment in patients with TTN-truncating variants as well as in the development of new drugs.
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http://dx.doi.org/10.1093/cvr/cvaa316DOI Listing
November 2020

Multiscale Modeling of Cardiovascular Function Predicts That the End-Systolic Pressure Volume Relationship Can Be Targeted via Multiple Therapeutic Strategies.

Front Physiol 2020 19;11:1043. Epub 2020 Aug 19.

Department of Biomedical Engineering, Yale University, New Haven, CT, United States.

Most patients who develop heart failure are unable to elevate their cardiac output on demand due to impaired contractility and/or reduced ventricular filling. Despite decades of research, few effective therapies for heart failure have been developed. In part, this may reflect the difficulty of predicting how perturbations to molecular-level mechanisms that are induced by drugs will scale up to modulate system-level properties such as blood pressure. Computer modeling might help with this process and thereby accelerate the development of better therapies for heart failure. This manuscript presents a new multiscale model that uses a single contractile element to drive an idealized ventricle that pumps blood around a closed circulation. The contractile element was formed by linking an existing model of dynamically coupled myofilaments with a well-established model of myocyte electrophysiology. The resulting framework spans from molecular-level events (including opening of ion channels and transitions between different myosin states) to properties such as ejection fraction that can be measured in patients. Initial calculations showed that the model reproduces many aspects of normal cardiovascular physiology including, for example, pressure-volume loops. Subsequent sensitivity tests then quantified how each model parameter influenced a range of system level properties. The first key finding was that the End Systolic Pressure Volume Relationship, a classic index of cardiac contractility, was ∼50% more sensitive to parameter changes than any other system-level property. The second important result was that parameters that primarily affect ventricular filling, such as passive stiffness and Ca reuptake via sarco/endoplasmic reticulum Ca-ATPase (SERCA), also have a major impact on systolic properties including stroke work, myosin ATPase, and maximum ventricular pressure. These results reinforce the impact of diastolic function on ventricular performance and identify the End Systolic Pressure Volume Relationship as a particularly sensitive system-level property that can be targeted using multiple therapeutic strategies.
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http://dx.doi.org/10.3389/fphys.2020.01043DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7466769PMC
August 2020

Effects of mavacamten on Ca sensitivity of contraction as sarcomere length varied in human myocardium.

Br J Pharmacol 2020 12 21;177(24):5609-5621. Epub 2020 Oct 21.

Department of Integrative Physiology and Neuroscience, Washington State University, Pullman, Washington, USA.

Background And Purpose: Heart failure can reflect impaired contractile function at the myofilament level. In healthy hearts, myofilaments become more sensitive to Ca as cells are stretched. This represents a fundamental property of the myocardium that contributes to the Frank-Starling response, although the molecular mechanisms underlying the effect remain unclear. Mavacamten, which binds to myosin, is under investigation as a potential therapy for heart disease. We investigated how mavacamten affects the sarcomere-length dependence of Ca -sensitive isometric contraction to determine how mavacamten might modulate the Frank-Starling mechanism.

Experimental Approach: Multicellular preparations from the left ventricular-free wall of hearts from organ donors were chemically permeabilized and Ca activated in the presence or absence of 0.5-μM mavacamten at 1.9 or 2.3-μm sarcomere length (37°C). Isometric force and frequency-dependent viscoelastic myocardial stiffness measurements were made.

Key Results: At both sarcomere lengths, mavacamten reduced maximal force and Ca sensitivity of contraction. In the presence and absence of mavacamten, Ca sensitivity of force increased as sarcomere length increased. This suggests that the length-dependent activation response was maintained in human myocardium, even though mavacamten reduced Ca sensitivity. There were subtle effects of mavacamten reducing force values under relaxed conditions (pCa 8.0), as well as slowing myosin cross-bridge recruitment and speeding cross-bridge detachment under maximally activated conditions (pCa 4.5).

Conclusion And Implications: Mavacamten did not eliminate sarcomere length-dependent increases in the Ca sensitivity of contraction in myocardial strips from organ donors at physiological temperature. Drugs that modulate myofilament function may be useful therapies for cardiomyopathies.
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http://dx.doi.org/10.1111/bph.15271DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7707091PMC
December 2020

Heart Failure in Humans Reduces Contractile Force in Myocardium From Both Ventricles.

JACC Basic Transl Sci 2020 Aug 22;5(8):786-798. Epub 2020 Jul 22.

Department of Physiology, University of Kentucky, Lexington, Kentucky.

This study measured how heart failure affects the contractile properties of the human myocardium from the left and right ventricles. The data showed that maximum force and maximum power were reduced by approximately 30% in multicellular preparations from both ventricles, possibly because of ventricular remodeling (e.g., cellular disarray and/or excess fibrosis). Heart failure increased the calcium (Ca) sensitivity of contraction in both ventricles, but the effect was bigger in right ventricular samples. The changes in Ca sensitivity were associated with ventricle-specific changes in the phosphorylation of troponin I, which indicated that adrenergic stimulation might induce different effects in the left and right ventricles.
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http://dx.doi.org/10.1016/j.jacbts.2020.05.014DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7452203PMC
August 2020

Regulation of Myofilament Contractile Function in Human Donor and Failing Hearts.

Front Physiol 2020 25;11:468. Epub 2020 May 25.

Division of Cardiovascular Medicine, University of Kentucky, Lexington, KY, United States.

Heart failure (HF) often includes changes in myocardial contractile function. This study addressed the myofibrillar basis for contractile dysfunction in failing human myocardium. Regulation of contractile properties was measured in cardiac myocyte preparations isolated from frozen, left ventricular mid-wall biopsies of donor ( = 7) and failing human hearts ( = 8). Permeabilized cardiac myocyte preparations were attached between a force transducer and a position motor, and both the Ca dependence and sarcomere length (SL) dependence of force, rate of force, loaded shortening, and power output were measured at 15 ± 1°C. The myocyte preparation size was similar between groups (donor: length 148 ± 10 μm, width 21 ± 2 μm, = 13; HF: length 131 ± 9 μm, width 23 ± 1 μm, = 16). The maximal Ca-activated isometric force was also similar between groups (donor: 47 ± 4 kN⋅m; HF: 44 ± 5 kN⋅m), which implicates that previously reported force declines in multi-cellular preparations reflect, at least in part, tissue remodeling. Maximal force development rates were also similar between groups (donor: = 0.60 ± 0.05 s; HF: k = 0.55 ± 0.04 s), and both groups exhibited similar Ca activation dependence of values. Human cardiac myocyte preparations exhibited a Ca activation dependence of loaded shortening and power output. The peak power output normalized to isometric force (PNPO) decreased by ∼12% from maximal Ca to half-maximal Ca activations in both groups. Interestingly, the SL dependence of PNPO was diminished in failing myocyte preparations. During sub-maximal Ca activation, a reduction in SL from ∼2.25 to ∼1.95 μm caused a ∼26% decline in PNPO in donor myocytes but only an ∼11% change in failing myocytes. These results suggest that altered length-dependent regulation of myofilament function impairs ventricular performance in failing human hearts.
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http://dx.doi.org/10.3389/fphys.2020.00468DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7261867PMC
May 2020

Force-dependent recruitment from myosin OFF-state increases end-systolic pressure-volume relationship in left ventricle.

Biomech Model Mechanobiol 2020 Dec 28;19(6):2683-2692. Epub 2020 Apr 28.

Department of Mechanical Engineering, University of Kentucky, 269 Ralph G. Anderson Building, Lexington, KY, 40506-0503, USA.

Finite element (FE) modeling is becoming increasingly prevalent in the world of cardiac mechanics; however, many existing FE models are phenomenological and thus do not capture cellular-level mechanics. This work implements a cellular-level contraction scheme into an existing nonlinear FE code to model ventricular contraction. Specifically, this contraction model incorporates three myosin states: OFF-, ON-, and an attached force-generating state. It has been speculated that force-dependent transitions from the OFF- to ON-state may contribute to length-dependent activation at the cellular level. The current work investigates the contribution of force-dependent recruitment out of the OFF-state to ventricular-level function, specifically the Frank-Starling relationship, as seen through the end-systolic pressure-volume relationship (ESPVR). Five FE models were constructed using geometries of rat left ventricles obtained via cardiac magnetic resonance imaging. FE simulations were conducted to optimize parameters for the cellular contraction model such that the differences between FE predicted ventricular pressures for the models and experimentally measured pressures were minimized. The models were further validated by comparing FE predicted end-systolic strain to experimentally measured strain. Simulations mimicking vena cava occlusion generated descending pressure volume loops from which ESPVRs were calculated. In simulations with the inclusion of the OFF-state, using a force-dependent transition to the ON-state, the ESPVR calculated was steeper than in simulations excluding the OFF-state. Furthermore, the ESPVR was also steeper when compared to models that included the OFF-state without a force-dependent transition. This suggests that the force-dependent recruitment of thick filament heads from the OFF-state at the cellular level contributes to the Frank-Starling relationship observed at the organ level.
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http://dx.doi.org/10.1007/s10237-020-01331-6DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7606253PMC
December 2020

Cardiac myosin regulatory light chain kinase modulates cardiac contractility by phosphorylating both myosin regulatory light chain and troponin I.

J Biol Chem 2020 04 21;295(14):4398-4410. Epub 2020 Feb 21.

Randall Centre for Cell and Molecular Biophysics and British Heart Foundation Centre of Research Excellence, King's College London, London SE1 1UL, United Kingdom

Heart muscle contractility and performance are controlled by posttranslational modifications of sarcomeric proteins. Although myosin regulatory light chain (RLC) phosphorylation has been studied extensively and , the precise role of cardiac myosin light chain kinase (cMLCK), the primary kinase acting upon RLC, in the regulation of cardiomyocyte contractility remains poorly understood. In this study, using recombinantly expressed and purified proteins, various analytical methods, and kinase assays, and mechanical measurements in isolated ventricular trabeculae, we demonstrate that human cMLCK is not a dedicated kinase for RLC but can phosphorylate other sarcomeric proteins with well-characterized regulatory functions. We show that cMLCK specifically monophosphorylates Ser of human cardiac troponin I (cTnI) in isolation and in the trimeric troponin complex and in the native environment of the muscle myofilament lattice. Moreover, we observed that human cMLCK phosphorylates rodent cTnI to a much smaller extent and , suggesting species-specific adaptation of cMLCK. Although cMLCK treatment of ventricular trabeculae exchanged with rat or human troponin increased their cross-bridge kinetics, the increase in sensitivity of myofilaments to calcium was significantly blunted by human TnI, suggesting that human cTnI phosphorylation by cMLCK modifies the functional consequences of RLC phosphorylation. We propose that cMLCK-mediated phosphorylation of TnI is functionally significant and represents a critical signaling pathway that coordinates the regulatory states of thick and thin filaments in both physiological and potentially pathophysiological conditions of the heart.
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http://dx.doi.org/10.1074/jbc.RA119.011945DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7135997PMC
April 2020

The Heart by Numbers.

Biophys J 2019 12 29;117(12):E1-E3. Epub 2019 Nov 29.

Department of Medicine and Division of Cardiology, University of California, Los Angeles, Los Angeles, California.

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http://dx.doi.org/10.1016/j.bpj.2019.11.010DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6990371PMC
December 2019

Diabetic microcirculatory disturbances and pathologic erythropoiesis are provoked by deposition of amyloid-forming amylin in red blood cells and capillaries.

Kidney Int 2020 01 5;97(1):143-155. Epub 2019 Sep 5.

Department of Pharmacology and Nutritional Sciences, University of Kentucky, Lexington, Kentucky, USA; Department of Neurology, University of Kentucky, Lexington, Kentucky, USA. Electronic address:

In the setting of type-2 diabetes, there are declines of structural stability and functionality of blood capillaries and red blood cells (RBCs), increasing the risk for microcirculatory disturbances. Correcting hyperglycemia is not entirely effective at reestablishing normal cellular metabolism and function. Therefore, identification of pathological changes occurring before the development of overt hyperglycemia may lead to novel therapeutic targets for reducing the risk of microvascular dysfunction. Here we determine whether RBC-capillary interactions are altered by prediabetic hypersecretion of amylin, an amyloid forming hormone co-synthesized with insulin, and is reversed by endothelial cell-secreted epoxyeicosatrienoic acids. In patients, we found amylin deposition in RBCs in association with type-2 diabetes, heart failure, cancer and stroke. Amylin-coated RBCs have altered shape and reduced functional (non-glycated) hemoglobin. Amylin-coated RBCs administered intravenously in control rats upregulated erythropoietin and renal arginase expression and activity. We also found that diabetic rats expressing amyloid-forming human amylin in the pancreas (the HIP rat model) have increased tissue levels of hypoxia-inducible transcription factors, compared to diabetic rats that express non-amyloid forming rat amylin (the UCD rat model). Upregulation of erythropoietin correlated with lower hematocrit in the HIP model indicating pathologic erythropoiesis. In the HIP model, pharmacological upregulation of endogenous epoxyeicosatrienoic acids protected the renal microvasculature against amylin deposition and also reduced renal accumulation of HIFs. Thus, prediabetes induces dysregulation of amylin homeostasis and promotes amylin deposition in RBCs and the microvasculature altering RBC-capillary interaction leading to activation of hypoxia signaling pathways and pathologic erythropoiesis. Hence, dysregulation of amylin homeostasis could be a therapeutic target for ameliorating diabetic vascular complications.
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http://dx.doi.org/10.1016/j.kint.2019.07.028DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6943180PMC
January 2020

Muscle thixotropy-where are we now?

J Appl Physiol (1985) 2019 06 9;126(6):1790-1799. Epub 2019 May 9.

Department of Physiology, College of Medicine, University of Kentucky , Lexington, Kentucky.

Relaxed skeletal muscle has an inbuilt resistance to movement. In particular, the resistance manifests itself as a substantial stiffness for small movements. The stiffness is impermanent, because it forms only when the muscle is stationary for some time and is reduced upon active or passive movement. Because the resistance to movement increases with time at rest and is reduced by movement, this behavior has become known as muscle thixotropy. In this short review, we describe the phenomenon of thixotropy and illustrate its significance in postural control with particular emphasis on human standing. We show how thixotropy came to be unambiguously associated with muscle mechanics and we review present knowledge of the molecular basis of thixotropic behavior. Specifically, we examine how recent knowledge about titin, and about the control of cross-bridge cycling, has impacted on the role of non-cross-bridge mechanisms and cross-bridge mechanisms in explaining thixotropy. We describe how thixotropic changes in muscle stiffness that occur during transitions from posture to movement can be tracked by analyzing physiological tremor. Finally, because skeletal muscle contains sensory receptors, and because some of these receptors are themselves thixotropic, we outline some of the consequences of muscle thixotropy for proprioception.
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http://dx.doi.org/10.1152/japplphysiol.00788.2018DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6734056PMC
June 2019

Closing the therapeutic loop.

Arch Biochem Biophys 2019 03 9;663:129-131. Epub 2019 Jan 9.

Department of Mechanical Engineering and Department of Surgery, University of Kentucky, United States.

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http://dx.doi.org/10.1016/j.abb.2019.01.006DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6377839PMC
March 2019

A short history of the development of mathematical models of cardiac mechanics.

J Mol Cell Cardiol 2019 02 29;127:11-19. Epub 2018 Nov 29.

Departments of Biomedical Engineering and Cellular and Molecular Physiology, Yale University, New Haven, USA.

Cardiac mechanics plays a crucial role in atrial and ventricular function, in the regulation of growth and remodelling, in the progression of disease, and the response to treatment. The spatial scale of the critical mechanisms ranges from nm (molecules) to cm (hearts) with the fastest events occurring in milliseconds (molecular events) and the slowest requiring months (growth and remodelling). Due to its complexity and importance, cardiac mechanics has been studied extensively both experimentally and through mathematical models and simulation. Models of cardiac mechanics evolved from seminal studies in skeletal muscle, and developed into cardiac specific, species specific, human specific and finally patient specific calculations. These models provide a formal framework to link multiple experimental assays recorded over nearly 100 years into a single unified representation of cardiac function. This review first provides a summary of the proteins, physiology and anatomy involved in the generation of cardiac pump function. We then describe the evolution of models of cardiac mechanics starting with the early theoretical frameworks describing the link between sarcomeres and muscle contraction, transitioning through myosin-level models to calcium-driven systems, and ending with whole heart patient-specific models.
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http://dx.doi.org/10.1016/j.yjmcc.2018.11.015DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6525149PMC
February 2019

Diabetes with heart failure increases methylglyoxal modifications in the sarcomere, which inhibit function.

JCI Insight 2018 10 18;3(20). Epub 2018 Oct 18.

Loyola University Chicago, Department of Cell and Molecular Physiology, Chicago, Illinois, USA.

Patients with diabetes are at significantly higher risk of developing heart failure. Increases in advanced glycation end products are a proposed pathophysiological link, but their impact and mechanism remain incompletely understood. Methylglyoxal (MG) is a glycolysis byproduct, elevated in diabetes, and modifies arginine and lysine residues. We show that left ventricular myofilament from patients with diabetes and heart failure (dbHF) exhibited increased MG modifications compared with nonfailing controls (NF) or heart failure patients without diabetes. In skinned NF human and mouse cardiomyocytes, acute MG treatment depressed both calcium sensitivity and maximal calcium-activated force in a dose-dependent manner. Importantly, dbHF myocytes were resistant to myofilament functional changes from MG treatment, indicating that myofilaments from dbHF patients already had depressed function arising from MG modifications. In human dbHF and MG-treated mice, mass spectrometry identified increased MG modifications on actin and myosin. Cosedimentation and in vitro motility assays indicate that MG modifications on actin and myosin independently depress calcium sensitivity, and mechanistically, the functional consequence requires actin/myosin interaction with thin-filament regulatory proteins. MG modification of the myofilament may represent a critical mechanism by which diabetes induces heart failure, as well as a therapeutic target to avoid the development of or ameliorate heart failure in these patients.
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http://dx.doi.org/10.1172/jci.insight.121264DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6237482PMC
October 2018

Author Correction: Abnormal contractility in human heart myofibrils from patients with dilated cardiomyopathy due to mutations in TTN and contractile protein genes.

Sci Rep 2018 Sep 24;8(1):14485. Epub 2018 Sep 24.

National Heart and Lung Institute, Imperial College London, London, W12 0NN, United Kingdom.

A correction to this article has been published and is linked from the HTML and PDF versions of this paper. The error has been fixed in the paper.
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http://dx.doi.org/10.1038/s41598-018-32408-zDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6155132PMC
September 2018

Differential Effects of Isoproterenol on Regional Myocardial Mechanics in Rat using 3D cine DENSE Cardiovascular Magnetic Resonance.

J Biomech Eng 2018 Aug 4. Epub 2018 Aug 4.

Department of Surgery, University of Kentucky, Lexington, KY, USA.

The present study assessed the acute effects of isoproterenol on left ventricular (LV) mechanics in healthy rats with the hypothesis that ß-adrenergic stimulation influences the mechanics of different myocardial regions of the LV wall in different ways. To accomplish this, magnetic resonance images were obtained in the LV of healthy rats with or without isoproterenol infusion. The LV contours were divided into basal, mid-ventricular, and apical regions. Additionally, the mid-ventricular myocardium was divided into three transmural layers with each layer partitioned into four segments (i.e., septal, inferior, lateral, and anterior). Peak systolic strains and torsion were quantified for each region. Isoproterenol significantly increased peak systolic radial strain and circumferential-longitudinal shear strain, as well as ventricular torsion, throughout the basal, mid-ventricle, and apical regions. In the mid-ventricle, isoproterenol significantly increased peak systolic radial strain, and induced significant increases in peak systolic circumferential strain and longitudinal strain in the septum. Isoproterenol consistently increased peak systolic circumferential-longitudinal shear strain in all mid-ventricular segments. Ventricular torsion was significantly increased in nearly all segments except the inferior sub-endocardium. The effects of isoproterenol on LV systolic mechanics (i.e., 3D strains and torsion) in healthy rats depend on the region. This region-dependency is also strain component-specific. These results provide insight into the regional response of LV mechanics to ß-adrenergic stimulation in rats, and could act as a baseline for future studies on subclinical abnormalities associated with the inotropic response in heart disease.
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http://dx.doi.org/10.1115/1.4041042DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7580659PMC
August 2018

Force-Dependent Recruitment from the Myosin Off State Contributes to Length-Dependent Activation.

Biophys J 2018 08 11;115(3):543-553. Epub 2018 Jul 11.

Department of Biomedical Engineering, Yale University, New Haven, Connecticut.

Cardiac muscle develops more force when it is activated at longer lengths. The concentration of Ca required to develop half-maximal force also decreases. These effects are known as length-dependent activation and are thought to play critical roles in the Frank-Starling relationship and cardiovascular homeostasis. The molecular mechanisms underpinning length-dependent activation remain unclear, but recent experiments suggest that they may include recruitment of myosin heads from the off (sometimes called super-relaxed) state. This manuscript presents a mathematical model of muscle contraction that was developed to investigate this hypothesis. Myosin heads in the model transitioned between an off state (that could not interact with actin), an on state (that could bind to actin), and a single attached state. Simulations were fitted to experimental data using multidimensional parameter optimization. Statistical analysis showed that a model in which the rate of the off-to-on transition increased linearly with force reproduced the length-dependent behavior of chemically permeabilized myocardium better than a model with a constant off-to-on transition rate (F-test, p < 0.001). This result suggests that the thick-filament transitions are modulated by force. Additional calculations showed that the model incorporating a mechanosensitive thick filament could also reproduce twitch responses measured in a trabecula stretched to different lengths. A final set of simulations was then used to test the model. These calculations predicted how reducing passive stiffness would impact the length dependence of the calcium sensitivity of contractile force. The prediction (a 60% reduction in ΔpCa) mimicked the 58% reduction in ΔpCa in myocardium from rats that expressed a giant isoform of titin and had low resting tension. Together, these computational results suggest that force-dependent recruitment of myosin heads from the thick-filament off state contributes to length-dependent activation and the Frank-Starling relationship.
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http://dx.doi.org/10.1016/j.bpj.2018.07.006DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6084639PMC
August 2018

Evaluation of a Novel Finite Element Model of Active Contraction in the Heart.

Front Physiol 2018 23;9:425. Epub 2018 Apr 23.

Department of Mechanical Engineering, University of Kentucky, Lexington, KY, United States.

Finite element (FE) modeling is becoming a widely used approach for the investigation of global heart function. In the present study, a novel model of cellular-level systolic contraction, which includes both length- and velocity-dependence, was implemented into a 3D non-linear FE code. To validate this new FE implementation, an optimization procedure was used to determine the contractile parameters, associated with sarcomeric function, by comparing FE-predicted pressure and strain to experimental measures collected with magnetic resonance imaging and catheterization in the ventricles of five healthy rats. The pressure-volume relationship generated by the FE models matched well with the experimental data. Additionally, the regional distribution of end-systolic strains and circumferential-longitudinal shear angle exhibited good agreement with experimental results overall, with the main deviation occurring in the septal region. Moreover, the FE model predicted a heterogeneous distribution of sarcomere re-lengthening after ventricular ejection, which is consistent with previous studies. In conclusion, the new FE active contraction model was able to predict the global performance and regional mechanical behaviors of the LV during the entire cardiac cycle. By including more accurate cellular-level mechanisms, this model could provide a better representation of the LV and enhance cardiac research related to both systolic and diastolic dysfunction.
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http://dx.doi.org/10.3389/fphys.2018.00425DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5924776PMC
April 2018

Abnormal contractility in human heart myofibrils from patients with dilated cardiomyopathy due to mutations in TTN and contractile protein genes.

Sci Rep 2017 11 1;7(1):14829. Epub 2017 Nov 1.

National Heart and Lung Institute, Imperial College London, London, W12 0NN, United Kingdom.

Dilated cardiomyopathy (DCM) is an important cause of heart failure. Single gene mutations in at least 50 genes have been proposed to account for 25-50% of DCM cases and up to 25% of inherited DCM has been attributed to truncating mutations in the sarcomeric structural protein titin (TTNtv). Whilst the primary molecular mechanism of some DCM-associated mutations in the contractile apparatus has been studied in vitro and in transgenic mice, the contractile defect in human heart muscle has not been studied. In this study we isolated cardiac myofibrils from 3 TTNtv mutants, and 3 with contractile protein mutations (TNNI3 K36Q, TNNC1 G159D and MYH7 E1426K) and measured their contractility and passive stiffness in comparison with donor heart muscle as a control. We found that the three contractile protein mutations but not the TTNtv mutations had faster relaxation kinetics. Passive stiffness was reduced about 38% in all the DCM mutant samples. However, there was no change in maximum force or the titin N2BA/N2B isoform ratio and there was no titin haploinsufficiency. The decrease in myofibril passive stiffness was a common feature in all hearts with DCM-associated mutations and may be causative of DCM.
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http://dx.doi.org/10.1038/s41598-017-13675-8DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5665940PMC
November 2017

No Difference in Myosin Kinetics and Spatial Distribution of the Lever Arm in the Left and Right Ventricles of Human Hearts.

Front Physiol 2017 13;8:732. Epub 2017 Oct 13.

Department of Cell Biology and Center for Commercialization of Fluorescence Technologies, University of North Texas, Health Science Center, Fort Worth, TX, United States.

The systemic circulation offers larger resistance to the blood flow than the pulmonary system. Consequently, the left ventricle (LV) must pump blood with more force than the right ventricle (RV). The question arises whether the stronger pumping action of the LV is due to a more efficient action of left ventricular myosin, or whether it is due to the morphological differences between ventricles. Such a question cannot be answered by studying the entire ventricles or myocytes because any observed differences would be wiped out by averaging the information obtained from trillions of myosin molecules present in a ventricle or myocyte. We therefore searched for the differences between single myosin molecules of the LV and RV of failing hearts . We show that the parameters that define the mechanical characteristics of working myosin (kinetic rates and the distribution of spatial orientation of myosin lever arm) were the same in both ventricles. These results suggest that there is no difference in the way myosin interacts with thin filaments in myocytes of failing hearts, and suggests that the difference in pumping efficiencies are caused by interactions between muscle proteins other than myosin or that they are purely morphological.
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http://dx.doi.org/10.3389/fphys.2017.00732DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5645524PMC
October 2017

MyoVision: software for automated high-content analysis of skeletal muscle immunohistochemistry.

J Appl Physiol (1985) 2018 01 5;124(1):40-51. Epub 2017 Oct 5.

Department of Physiology, College of Medicine, University of Kentucky , Lexington, Kentucky.

Analysis of skeletal muscle cross sections is an important experimental technique in muscle biology. Many aspects of immunohistochemistry and fluorescence microscopy can now be automated, but most image quantification techniques still require extensive human input, slowing progress and introducing the possibility of user bias. MyoVision is a new software package that was developed to overcome these limitations. The software improves upon previously reported automatic techniques and analyzes images without requiring significant human input and correction. When compared with data derived by manual quantification, MyoVision achieves an accuracy of ≥94% for basic measurements such as fiber number, fiber type distribution, fiber cross-sectional area, and myonuclear number. Scientists can download the software free from www.MyoVision.org and use it to automate the analysis of their own experimental data. This will improve the efficiency and consistency of the analysis of muscle cross sections and help to reduce the burden of routine image quantification in muscle biology. NEW & NOTEWORTHY Scientists currently analyze images of immunofluorescently labeled skeletal muscle using time-consuming techniques that require sustained human supervision. As well as being inefficient, these techniques can increase variability in studies that quantify morphological adaptations of skeletal muscle at the cellular level. MyoVision is new software that overcomes these limitations by performing high-content analysis of muscle cross sections with minimal manual input. It is open source and freely available.
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http://dx.doi.org/10.1152/japplphysiol.00762.2017DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6048460PMC
January 2018

The effects of pH and P on tension and Ca sensitivity of ventricular myofilaments from the anoxia-tolerant painted turtle.

J Exp Biol 2017 11 22;220(Pt 22):4234-4241. Epub 2017 Sep 22.

Department of Biology, Saint Louis University, St Louis, MO 63109, USA

We aimed to determine how increases in intracellular H and inorganic phosphate (P) to levels observed during anoxic submergence affect contractility in ventricular muscle of the anoxia-tolerant Western painted turtle, Skinned multicellular preparations were exposed to six treatments with physiologically relevant levels of pH (7.4, 7.0, 6.6) and P (3 and 8 mmol l). Each preparation was tested in a range of calcium concentrations (pCa 9.0-4.5) to determine the pCa-tension relationship for each treatment. Acidosis significantly decreased contractility by decreasing Ca sensitivity (pCa) and tension development (<0.001). Increasing [P] also decreased contractility by decreasing tension development at every pH level (<0.001) but, alone, did not affect Ca sensitivity (=0.689). Simultaneous increases in [H] and [P] interacted to attenuate the decreased tension development and Ca sensitivity (<0.001), possibly reflecting a decreased sensitivity to P when it is present as the dihydrogen phosphate form, which increases as pH decreases. Compared with that of mammals, the ventricle of turtles exhibits higher Ca sensitivity, which is consistent with previous studies of ectothermic vertebrates.
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http://dx.doi.org/10.1242/jeb.164137DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6514463PMC
November 2017

Regional quantification of myocardial mechanics in rat using 3D cine DENSE cardiovascular magnetic resonance.

NMR Biomed 2017 Aug 8;30(8). Epub 2017 May 8.

Department of Mechanical Engineering, University of Kentucky, Lexington, KY, USA.

Rat models have assumed an increasingly important role in cardiac research. However, a detailed profile of regional cardiac mechanics, such as strains and torsion, is lacking for rats. We hypothesized that healthy rat left ventricles (LVs) exhibit regional differences in cardiac mechanics, which are part of normal function. In this study, images of the LV were obtained with 3D cine displacement encoding with stimulated echoes (DENSE) cardiovascular magnetic resonance in 10 healthy rats. To evaluate regional cardiac mechanics, the LV was divided into basal, mid-ventricular, and apical regions. The myocardium at the mid-LV was further partitioned into four wall segments (i.e. septal, inferior, lateral, and anterior) and three transmural layers (i.e. sub-endocardium, mid-myocardium, and sub-epicardium). The six Lagrangian strain components (i.e. E , E , E , E , E , and E ) were computed from the 3D displacement field and averaged within each region of interest. Torsion was quantified using the circumferential-longitudinal shear angle. While peak systolic E differed between the mid-ventricle and apex, the other five components of peak systolic strain were similar across the base, mid-ventricle, and apex. In the mid-LV myocardium, E decreased gradually from the sub-endocardial to the sub-epicardial layer. E demonstrated significant differences between the four wall segments, with the largest magnitude in the inferior segment. E was uniform among the four wall segments. E varied along the transmural direction and among wall segments, whereas E differed only among the wall segments. E was not associated with significant variations. Torsion also varied along the transmural direction and among wall segments. These results provide fundamental insights into the regional contractile function of healthy rat hearts, and form the foundation for future studies on regional changes induced by disease or treatments.
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http://dx.doi.org/10.1002/nbm.3733DOI Listing
August 2017

Super-relaxation helps muscles work more efficiently.

J Physiol 2017 02;595(4):1007-1008

Department of Physiology and Division of Cardiovascular Medicine, University of Kentucky, MS508 Chandler Medical Center, 800 Rose Street, Lexington, KY, 40536-0298, USA.

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http://dx.doi.org/10.1113/JP273629DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5309356PMC
February 2017
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