Identification and Imaging of Skeletal Muscle Response to Graded Nerve Crush

骨骼肌对分级神经挤压反应的识别和成像

• 批准号: 10646172
• 负责人: David Micah Brogan
• 金额: $ 15.45万
• 依托单位: WASHINGTON UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-07-01 至 2026-06-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10646172
• 关键词: AKT Signaling Pathway; Affect; Animal Model; Assessment tool; Atrophic; Biological; Biological Assay; Calpain; Caring; Cells; Clinical; Crush Injury; Custom; Data Correlations; Denervation; Desmin; Early Intervention; Enzyme-Linked Immunosorbent Assay; Evaluation; Event; Excision; Experimental Designs; Feedback; Female; Fibrosis; Filament; Fluorescence; Fluorescent Probes; Functional Imaging; Functional disorder; Future; Genetic Transcription; Goals; Grant; Hamsters; Hand Strength; Health; Hindlimb; Hippocampus; Image; In Vitro; Injectable; Injections; Injury; Investigation; Journals; Lesion; Limb structure; Literature; Mediating; Mediator; Mentors; Methodology; Methods; Modeling; Molecular; Molecular Biology; Molecular Probes; Molecular Target; Mus; Muscle; Muscle denervation procedure; Muscular Atrophy; Musculoskeletal; Myofibrils; Myogenin; Nature; Near-infrared optical imaging; Nerve; Nerve Crush; Neuregulins; Operative Surgical Procedures; Optics; Orthopedics; Outcome; PI3K/AKT; Pathway interactions; Patients; Penetration; Peptide Hydrolases; Peripheral Nerves; Process; Prognosis; Proteins; Recovery; Recovery of Function; Reporting; Research; Research Methodology; Resected; Resources; Role; Scientist; Secondary to; Series; Severities; Signal Transduction; Skeletal Muscle; Surgeon; System; Techniques; Technology; Testing; Time; Training; Translations; Trauma; Traumatic injury; Up-Regulation; Visualization; Walking; Wallerian Degeneration; Work; Writing; axon injury; bone imaging; clinical translation; depolymerization; design; diagnostic tool; differential expression; disability; ex vivo imaging; experience; functional outcomes; iatrogenic injury; imaging probe; imaging system; improved; improved outcome; in vivo; in vivo imaging; insight; interest; male; molecular marker; mouse model; nerve injury; nerve repair; nerve transection; neuromuscular; novel; novel diagnostics; optical imaging; preclinical imaging; predictive marker; protein aminoacid sequence; repaired; response; restoration; sciatic nerve; skeletal muscle wasting; skills; stem; time interval; tool; transcription factor

Project Summary Atrophy and fibrosis of skeletal muscle after neuromuscular trauma is a significant impediment to the restoration of function after severe neuromuscular trauma. Despite this, dynamic assessment tools for muscle wasting and dysfunction are limited, leaving a critical gap in the orthopedic surgeon’s ability to assess the degree of neurogenic muscle injury and its ultimate prognosis. This gap stems in part from an incomplete understanding of the role of increased expression of transcriptional factors and proteases related to atrophy, and inability to dynamically assess them clinically. Calpain is one of these proteases central to the myofibril destruction of neurogenic atrophy, and therefore has potential to serve as a marker of muscle atrophy. However, translation of this relationship into a diagnostic tool is limited by a lack of techniques for real time assessment of calpain activity. The proposed work seeks to explore the potential for use of optical probes to identify muscle atrophy by examining the relationship between nerve injury and muscle contractility and calpain activity. Aim 1a will determine if calpain expression and activity will increase proportionally with nerve injury and muscle dysfunction. Equal numbers of male and female mice will be subjected to a varying degree of unilateral sciatic nerve crush injury. At a subsequent surgery, at staged intervals, functional recovery will be assessed with walking track analysis and grip strength testing. Hindlimb muscles will undergo ex-vivo contractility testing, as well as histomorphometric analysis and relevant transcriptional factors will be assayed. Calpain activity will be quantified with ELISA kits and with use of a pre-clinical imaging system to detect near-infrared fluorescence (NIR) within the hindlimb muscles after administration of an injectable calpain sensitive probe. In Aim 1b, a unilateral sciatic nerve transection and repair will be performed in the mice, and the same series of functional tests, transcriptional assays and NIR imaging with the optical probe will be undertaken. Similarly, Aim 1c will utilize the same methodologic assessments, at the same time intervals, after removal of a segment of sciatic nerve. The increasing degree of nerve injuries and proposed assessments will help to delineate the canonical pathways of muscle atrophy after nerve injury, and the proposed optical probe will provide a powerful new diagnostic tool. As an orthopedic surgeon with a practice devoted to the care of mangled limbs, I understand the clinical impact of such injuries, but need protected time and resources to develop the skills to study these at a molecular level. In addition to the investigations described above, I will participate in graduate coursework to improve my understanding of molecular biology, as well as optical and biological imaging. I will regularly participate in scholarly activities such as journal clubs and grant seminars through the Musculoskeletal Research Center to enhance my grant writing abilities and improve my understanding of experimental methodologies. In addition to my primary and secondary mentor, I have assembled a mentoring committee to give feedback on results and assist with experimental design. This constellation of planned activities, along with the proposed research methods above will provide me the requisite training and experience to develop as a clinician scientist with an interest in optical imaging of skeletal muscle atrophy.

项目总结