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Single Cross-Bridge Kinetics in Transgenic Mouse Hearts Expressing FHC Mutations

表达 FHC 突变的转基因小鼠心脏中的单桥动力学

基本信息

批准号：
8249067
负责人：
JULIAN BOREJDO
金额：
$ 39.64万
依托单位：
UNIVERSITY OF NORTH TEXAS HLTH SCI CTR
依托单位国家：
美国
项目类别：
财政年份：
2009
资助国家：
美国
起止时间：
2009-04-15 至 2014-03-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8249067
关键词：
ATP Hydrolysis ATP phosphohydrolase Abbreviations Actins Address Age Animal Model Applications Grants Asses Binding Biological Calcium Calmodulin Cardiac Cardiac Muscle Contraction Cardiovascular Diseases Clinical Contractile Proteins Contracts Data Detection Development Disease Dissociation Dyspnea Electrocardiogram Environment Familial Hypertrophic Cardiomyopathy Fatigue Fiber Fluorescence Fluorescence Spectroscopy Gene Transfer Techniques Genes Goals Head Heart Heart Diseases Heart Hypertrophy Heart failure Human Hypertrophic Cardiomyopathy Hypertrophy Individual Isometric Contraction Kinetics Label Lead Left Light Link Malignant - descriptor Measurement Measures Mechanics Mediating Microscopic Mind Modality Molecular Molecular Biology Monitor Muscle Muscle Contraction Muscle Fibers Mutant Strains Mice Mutation Myocardium Myofibrils Myopathy Myosin ATPase Myosin Alkali Light Chains Myosin Light Chain Kinase Myosin Light Chains Myosin Regulatory Light Chains Nanotechnology Optics Organ Pathology Patients Performance Phenotype Physiological Physiology Point Mutation Preparation Principal Investigator Process Proteins Public Health Recombinants Research Research Personnel Resolution Role Rotation Sarcomeres Site Skin Solutions Solvents Spectrum Analysis Structure Techniques Technology Testing Thick Filament Thin Filament Time Transgenic Animals Transgenic Mice Transgenic Organisms Ventricular Ventricular Myosins Work aqueous base blood pump cost disease phenotype disease-causing mutation experience fluorescence microscope fluorophore innovation mortality multidisciplinary mutant nano nanomechanics papillary muscle premature public health relevance single molecule sudden cardiac death ventricular hypertrophy

项目摘要

Familial hypertrophic cardiomyopathy (FHC) is an autosomal dominant disease originating from mutations in genes that encode for the major contractile proteins of the heart, including the ventricular myosin regulatory (RLC) and essential (ELC) light chains. FHC results in ventricular and septal hypertrophy, myofibrillar disarray and is the leading cause of sudden cardiac death in young individuals. This research is aimed at elucidating the molecular mechanisms involved in triggering of FHC at the level of a single myosin cross-bridge. We propose to test the hypothesis that FHC is caused by inefficient utilization of ATP by cardiac muscle due to alteration of myosin cross-bridge kinetics in transgenic mouse hearts expressing disease-causing mutations in myosin RLC and ELC. We will examine this hypothesis at the single molecule level in papillary muscle fibers from transgenic mouse hearts which carry disease-causing mutations in the regulatory and/or essential light chains of myosin. We strongly believe that the unambiguous determination of myosin cross-bridge kinetics must be carried out at the level of a single cross-bridge and the results compared to cross-bridge mechanics derived from measurements on skinned and intact muscle fibers. The advantage of the single molecule approach is its ability to avoid averaging over ensembles of molecules with different kinetics such as a mixture of WT and FHC molecules, and the ability to unambiguously determine the kinetics of "healthy" and "diseased" muscle. Since human patients are heterozygous for FHC mutations and their thick filaments contain interspersed WT and HCM mutant heads it is extremely important to correlate the single molecule information with the phenotype of FHC assessed at the muscle fiber level. Specifically we ask whether the durations (Aim 1A) and lifetimes (Aim 1B) of detached and strongly-bound states are the same in a single cross-bridge from FHC hearts and in healthy transgenic controls. The information derived using this single molecule technology will be paralleled with functional studies of force development, ATPase on skinned papillary muscle fibers as well as force and calcium transients on intact muscle fibers from transgenic mice (Aim 2A). The ultimate objective is to link the single molecule derived data with the cellular findings to fully understand the mechanism of action of the individual RLC and ELC mutations causing FHC (Aim 2B). The fundamental question that is being addressed is why and how these individual mutations in RLC and/or ELC cause variable disease phenotypes in humans ranging from relatively mild to malignant clinical FHC phenotypes. We believe that integration of molecular biology approaches with high resolution optics and nano-fluorescence spectroscopy will enable us to successfully answer important questions regarding the molecular basis of FHC-mediated pathology in the heart and the role of RLC and ELC in cardiac muscle contraction.

家族性肥厚性心肌病是一种常染色体显性遗传病

项目成果

期刊论文数量（8）

专著数量（0）

科研奖励数量（0）

会议论文数量（0）

专利数量（0）

Elimination of autofluorescence background from fluorescence tissue images by use of time-gated detection and the AzaDiOxaTriAngulenium (ADOTA) fluorophore.

DOI：
10.1007/s00216-012-6623-1
发表时间：
2013-02
期刊：
ANALYTICAL AND BIOANALYTICAL CHEMISTRY
影响因子：
4.3
作者：
Rich, Ryan M.;Stankowska, Dorota L.;Maliwal, Badri P.;Sorensen, Thomas Just;Laursen, Bo W.;Krishnamoorthy, Raghu R.;Gryczynski, Zygmunt;Borejdo, Julian;Gryczynski, Ignacy;Fudala, Rafal
通讯作者：
Fudala, Rafal

Real-time imaging of exocytotic mucin release and swelling in Calu-3 cells using acridine orange.

使用吖啶橙对 Calu-3 细胞中的胞吐粘蛋白释放和肿胀进行实时成像。

DOI：
10.1016/j.ymeth.2013.09.004
发表时间：
2014
期刊：
Methods (San Diego, Calif.)
影响因子：
0
作者：
Shumilov,Dmytro;Popov,Alexander;Fudala,Rafal;Akopova,Irina;Gryczynski,Ignacy;Borejdo,Julian;Gryczynski,Zygmunt;Grygorczyk,Ryszard
通讯作者：
Grygorczyk,Ryszard

Elimination of autofluorescence in fluorescence correlation spectroscopy using the AzaDiOxaTriAngulenium (ADOTA) fluorophore in combination with time-correlated single-photon counting (TCSPC).

DOI：
10.1007/s00216-013-6879-0
发表时间：
2013-05
期刊：
ANALYTICAL AND BIOANALYTICAL CHEMISTRY
影响因子：
4.3
作者：
Rich, Ryan M.;Mummert, Mark;Gryczynski, Zygmunt;Borejdo, Julian;Sorensen, Thomas Just;Laursen, Bo W.;Foldes-Papp, Zeno;Gryczynski, Ignacy;Fudala, Rafal
通讯作者：
Fudala, Rafal

Detection of hyaluronidase activity using fluorescein labeled hyaluronic acid and Fluorescence Correlation Spectroscopy.

DOI：
10.1016/j.jphotobiol.2012.07.007
发表时间：
2012-11-05
期刊：
JOURNAL OF PHOTOCHEMISTRY AND PHOTOBIOLOGY B-BIOLOGY
影响因子：
5.4
作者：
Rich, Ryan M.;Mummert, Mark;Foldes-Papp, Zeno;Gryczynski, Zygmunt;Borejdo, Julian;Gryczynski, Ignacy;Fudala, Rafal
通讯作者：
Fudala, Rafal

Fluorescence detection of MMP-9. I. MMP-9 selectively cleaves Lys-Gly-Pro-Arg-Ser-Leu-Ser-Gly-Lys peptide.

DOI：
10.2174/138920111795470967
发表时间：
2011-05
期刊：
Current pharmaceutical biotechnology
影响因子：
2.8
作者：
Fudala R;Ranjan AP;Mukerjee A;Vishwanatha JK;Gryczynski Z;Borejdo J;Sarkar P;Gryczynski I
通讯作者：
Gryczynski I