The overall purpose of this proposed study is the understanding of cellular mechanisms involved in the pathologic fatty degeneration of muscle. Fatty infiltration in skeletal muscle is observed following sports injuries such as muscle strain injuries and Achilles tendon rupture. It is also observed in the degenerative state after rotator cuff tears as well as in the aging process. While fatty degeneration of skeletal muscle occurs in many different conditions and is known to decrease muscle function, the cellular processes involved in fatty infiltration have not been investigated in human muscle.
Hypotheses:
1. There is a high amount of fibro-adipogenic progenitors (FAPs) with an adipogenic pattern in pathologic skeletal muscle following a muscle strain injury and a full Achilles tendon rupture. We hypothesize that there is an increased number of FAPs with an adipogenic signature already in the acute phase after a strain injury, but a significantly higher number in the chronic stage as well as in the muscle following an Achilles tendon rupture. 2. Mechanical cues are a major driver of the phenotypic drift of FAPs. The lack of mechanical stimuli initiates the adoption of an adipogenic pathway in naïve FAPs, whereas naïve FAPs exposed to mechanical stimuli will maintain their undifferentiated phenotype. 3. The adherence of FAPs to a soft material will activate the adoption of an adipogenic phenotype, whereas a stiff material will favor a more fibrotic phenotype in naïve FAP's isolated from healthy skeletal muscle.
18 years and older, male and female, with an acute muscle strain injury (<14 days post injury) in the calf or hamstring muscles (group 1). Pathological changes visible on an ultrasound scan as hypoechoic areas.
18 years and older, male and female, with a chronic muscle strain injury (> 6 months prior to inclusion) in the calf or hamstring muscles (group 2), sequela including recurrent strain injuries, pain and/ or decrease in muscle function. Pathological changes visible on an ultrasound scan as hypo-/ hyperechoic areas.
18 years and older, male and female, with a full Achilles tendon rupture (> 1 year prior to inclusion) with pathological changes: Fatty degeneration visible on an ultrasound scan (group 3).
Type I and II Diabetes, Connective tissue and/or rheumatic diseases, or any observed organ dysfunctions
Daily smoking
Persons with daily intake of non-steroidal anti-inflammatory drugs (NSAIDs) within three months prior to time of contact
Allergic reactions to local anesthesia
Healthy individuals with an acute or previous muscle strain injury or a previous Achilles tendon rupture
Cohort
Prospective
Biopsies
Procedure/SurgeryBiopsies from the pathological muscle and healthy, unaffected muscle
Presence and number of Fibro-adipogenic progenitors
Determination of the number and the phenotypic appearance of fibro-adipogenic progenitors in the injured compared to the healthy muscle
Cell cluster analysis
Analysis of cell clusters present in the injured compared to the healthy muscle by single nuclei RNA sequencing
Importance of mechanical stimuli for Fibro-adipogenic progenitors
Importance of mechanical stimuli on the phenotypic drift of fibro-adipogenic progenitors ("FAPS". Isolated FAP´s will be subjected to either mechanical stimulation (by using the FlexCell system) or unloading ("FAPs embedded in a free-floating matrix). FAPs will also be cultured on stiff versus soft surfaces to study thereafter the effect of the substrate properties on the phenotypic appearance of FAPs. The phenotypic appearance of FAPs will be tested by qRT-PCR, protein analysis and compared to the well-described effect on phenotypic apprearance of FAPs when subjected to pro-fibrotic or pro-adipogenic differentiation medium.
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