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P&F Grant Awards

Year 7


Grant # 15

Promoting rotator cuff muscle regeneration with paracrine adipose signaling.

PI: Gretchen Meyer, PhD

 

Specific Aims

Rotator cuff (RC) tears are one of the most common causes of musculoskeletal pain and dysfunction, estimated to affect more than 10 million people in the US alone (1). Widespread degeneration of cuff tissues, particularly RC muscles, is a hallmark of chronic tears and is associated with both a progressive decline in shoulder function and high re-tear rates following surgical repair (2-5). In the case of a chronic tear, up to 75% of the muscle volume may be replaced with fat, both internally (intramuscular) (6,7) and externally (epimuscular) (8), in a process termed fatty atrophy. Little is known about the influence of fat expansion on RC tissues. It is generally considered undesirable due to its correlation with poor surgical outcomes (4,9), however no studies have directly examined its influence on cuff degeneration, function, or recovery. Given that over 50% of surgical cases include substantial fatty expansion (6,8), addressing this significant gap in knowledge could significantly impact shoulder function and, ultimately, the quality of life for a large patient population.

Adipose depots influence neighboring tissue function through endocrine/paracrine signaling (10,11). The secreted factors, and pathways modulated, vary substantially between depots. White depots (e.g., visceral or subcutaneous fat), are associated with the release of inflammatory factors, many upstream of the muscle atrophy pathway (12). However, brown fat, a specialized thermogenic depot, secretes various growth factors known to positively influence muscle health (13-15). Recently a new adipose type, termed inducible-brown or beige fat, has been identified with a phenotype intermediate between classical white and brown depots (16,17). Beige fat has been touted for its therapeutic promise as it can be pushed along the white-brown spectrum, in a process termed “browning” (18-20).

Interestingly, our preliminary data demonstrates that epimuscular (EM) fat in the human RC is a unique beige depot whose transcriptional resemblance to brown fat is decreased in the torn vs. intact cuff, suggesting that its phenotype can be modulated. This result reveals a potential therapeutic strategy, as paracrine signaling from “browned” beige fat has been shown to regulate bone density via an insulin-like growth factor (IGF) pathway (21). Considering IGF signaling is also a major pathway for muscle hypertrophy (22), we believe the “brownness” of EM fat could act as a regulatory switch for muscle tone. However, a significant gap in understanding the paracrine regulation of muscle function by adipose tissues stands in the way of targeting EM fat therapeutically. To begin to address this critical knowledge gap, we will adapt mouse models of acute regeneration and chronic degeneration in the rotator cuff to test our central hypothesis that paracrine signaling by brown fat is anabolic for muscle and shifting adipose phenotype away from brown exacerbates muscle degenerative changes and blunts regenerative capacity. This hypothesis represents a paradigm shift in thinking about RC pathology as it suggests that the large stores of fat in the cuff, previously considered a negative feature of the disease, present a novel therapeutic opportunity to prevent muscle degeneration or enhance muscle regeneration. We will test our hypothesis via two Specific Aims:

Specific Aim #1. Evaluate paracrine regulation of muscle regeneration by phenotype-specific fat. Approach: RC muscle regeneration will be stimulated using an injection of toxin into the infraspinatus (IS) muscle. Adipose grafts from brown, beige or white depots of GFP mice will be transplanted adjacent to the injured IS to examine their effect on muscle regeneration. Regenerative efficiency will be assayed via ex-vivo force production and histological measures of hypertrophy, fibrosis, and angiogenesis. GFP expression will be examined histologically to identify direct contribution of adipose-derived cells to myogenesis. The gene expression signature of excised adipose grafts will be evaluated to identify putative signaling mediators. We hypothesize that muscle regeneration will be more robust in the presence of brown fat compared to beige or white grafts and will be correlated with expression of myogenic growth factors.

Specific Aim #2. Investigate the effects of adipose phenotype in a chronic model of rotator cuff degeneration. Approach: Chronic degeneration of the rotator cuff will be induced using our well-established model of dual tenotomy of the infraspinatus and supraspinatus tendons. Adipose tissue will be isolated and transplanted as in Aim 1 to evaluate its potential for mitigating cuff degeneration. Following 16 weeks of progressive degeneration, muscle quality will be assayed as in Aim 1. Adipose grafts will be excised and their expression signature compared with results from Aim 1 to identify phenotypic shifts as a result of exposure to the chronic degenerative environment. We hypothesize that beige grafts will shift toward a whiter profile in the chronic model and this shift will be associated with increased expression of inflammatory mediators and decreased muscle quality.

We expect that the completion of these Aims will: (1) develop an in vivo system to manipulate and evaluate muscle-fat cross-talk in a pathological environment, (2) define the influence of fat phenotype on skeletal muscle pathology and, and (3) point to novel signaling mechanisms for future exploration. We believe this data will form a springboard for the development of strategies exploiting the large stores of adipose tissue in the torn rotator cuff for therapeutic benefit, not only for muscle but for tendon and bone healing as well.