Spontaneous muscle regenerative potential is limited, as severe injuries incompletely recover and result in chronic inflammation. Current therapies are restricted to conservative management, not providing a complete restitutio ad integrum; therefore, alternative therapeutic strategies are welcome, such as cell-based therapies with stem cells or Peripheral Blood Mononuclear Cells (PBMCs). Here, we described two different in vitro myogenic models: a 2D perfused system and a 3D bioengineered scaffold within a perfusion bioreactor. Both models were assembled with human bone marrow-derived mesenchymal stem cells (hBM-MSCs) and human primary skeletal myoblasts (hSkMs) to study induction and maintenance of myogenic phenotype in presence of PBMCs. When hBM-MSCs were cultured with human primary skeletal myoblasts (hSkMs) in medium supplemented with 10 ng/mL of bFGF; cells showed increased expression of myogenic-related gene, such as Desmin and Myosin Heavy Chain II (MYH2) after 21 days, and a prevalent expression of anti-inflammatory cytokines (IL10, 15-fold). Next, PBMCs were added in an upper transwell chamber and hBM-MSCs significantly upregulated myogenic genes throughout the culture period, while pro-inflammatory cytokines (e.g., IL12A) were downregulated. In 3D, hBM-MSCs plus hSkMs embedded in fibrin-based scaffolds, cultured in dynamic conditions, showed that all myogenic-related genes tended to be upregulated in the presence of PBMCs, and Desmin and MYH2 were also detected at protein level, while pro-inflammatory cytokine genes were significantly downregulated in the presence of PBMCs. In conclusion, our works suggest that hBM-MSCs have a versatile myogenic potential, enhanced and modulated by PMBCs. Moreover, our 3D biomimetic approach seemed to better resemble the tissue architecture allowing an efficient in vitro cellular cross-talk

Summary Statement. This experimental study showed that platelet rich fibrin matrix can improve muscle regeneration and long-term vascularization without local adverse effects. Introduction. Even though muscle injuries are very common, few scientific data on their effective treatment exist. Growth Factors (GFs) may have a role in accelerating muscle repair processes and a currently available strategy for their delivery into the lesion site is the use of autologous platelet-rich plasma (PRP). The present study is focused on the use of Platelet Rich Fibrin Matrix (PRFM), as a source of GFs. Materials and Methods. Bilateral muscular lesions were created on the longissimus dorsi muscle of Wistar rats. One side of the lesion was filled with a PRFM while the contralateral was left untreated (controls). Animals were sacrificed at 5, 10, 40 and 60 days from surgery. Histological, immunohistochemical and histomorphometric analyses were performed to evaluate muscle regeneration, neovascularization, fibrosis and inflammation. The presence of metaplasic zones, calcifications and heterotopic ossification were also assessed. Results. PRFM treated muscles exhibited an improved muscular regeneration, an increase in neovascularization, and a slight reduction of fibrosis compared with controls. No differences were detected for inflammation. Metaplasia, ossification and heterotopic calcification were not detected. Conclusions. This preliminary morphological experimental study shows that PRFM use can improve muscle regeneration and long-term vascularization. Since autologous blood products are safe, PRFM may be a useful and handily product in clinical treatment of muscle injuries

Injury to muscles is very common. We have previously observed that basic fibroblast growth factor (b-FGF), insulin growth factor type 1 (IGF-1) and nerve growth factor (NGF) are potent stimulators of the proliferation and fusion of myoblasts in vitro. We therefore injected these growth factors into mice with lacerations of the gastrocnemius muscle. The muscle regeneration was evaluated at one week by histological staining and quantitative histology. Muscle healing was assessed histologically and the contractile properties were measured one month after injury. Our findings showed that b-FGF, IGF and to a less extent NGF enhanced muscle regeneration in vivo compared with control muscle. At one month, muscles treated with IGF-1 and b-FGF showed improved healing and significantly increased fast-twitch and tetanus strengths. Our results suggest that b-FGF and IGF-1 stimulated muscle healing and may have a considerable effect on the treatment of muscle injuries

Myostatin (GDF-8) is known to play an important role in muscle regeneration, and myostatin is also expressed during the early phases of fracture healing. In this study we used fluorescent immunohistochemistry to define the temporal and spatial localization of myostatin during muscle and bone repair following deep penetrant injury in a mouse model. We then used hydrogel delivery of exogenous myostatin in the same injury model to determine the effects of myostatin exposure on muscle and bone healing. Results show that while myostatin was constitutively expressed in the cytoplasm of intact skeletal muscle fibers, a pool of intense myostatin staining was observed amongst injured skeletal muscle fibers 12-24 hours post-surgery. Myostatin was also expressed in the soft callus chondrocytes 4 days following osteotomy. Hydrogel delivery of 10 or 100 ug/ml recombinant myostatin decreased fracture callus cartilage area relative to total callus area in a dose-dependent manner by 41% and 80% (p<0.05), respectively, compared to vehicle treatment. Myostatin treatment also dose-dependently decreased fracture callus total bone volume by 23% and 47% (p<0.05), with the higher dose of recombinant myostatin yielding the greatest decrease in callus bone volume. Finally, exogenous myostatin treatment caused a significant, dose-dependent increase in fibrous tissue formation in skeletal muscle. Together, these findings suggest that myostatin may inhibit bone repair after traumatic musculoskeletal injury through both autocrine (soft-callus chondrocytes) and paracrine (surrounding injured muscle fibers) mechanisms. Thus, early pharmacological inhibition of myostatin is likely to improve the regenerative potential of both muscle and bone following deep penetrant musculoskeletal injury

Objectives

Rotator cuff tears are among the most frequent upper extremity injuries. Current treatment strategies do not address the poor quality of the muscle and tendon following chronic rotator cuff tears. Hypoxia-inducible factor-1 alpha (HIF-1α) is a transcription factor that activates many genes that are important in skeletal muscle regeneration. HIF-1α is inhibited under normal physiological conditions by the HIF prolyl 4-hydroxylases (PHDs). In this study, we used a pharmacological PHD inhibitor, GSK1120360A, to enhance the activity of HIF-1α following the repair of a chronic cuff tear, and measured muscle fibre contractility, fibrosis, gene expression, and enthesis mechanics.

Objectives

Rotator cuff tears are among the most common and debilitating upper extremity injuries. Chronic cuff tears result in atrophy and an infiltration of fat into the muscle, a condition commonly referred to as ‘fatty degeneration’. While stem cell therapies hold promise for the treatment of cuff tears, a suitable immunodeficient animal model that could be used to study human or other xenograft-based therapies for the treatment of rotator cuff injuries had not previously been identified.

The nervous system is known to be involved in inflammation and repair. We aimed to determine the effect of physical activity on the healing of a muscle injury and to examine the pattern of innervation. Using a drop-ball technique, a contusion was produced in the gastrocnemius in 20 rats. In ten the limb was immobilised in a plaster cast and the remaining ten had mobilisation on a running wheel. The muscle and the corresponding dorsal-root ganglia were studied by histological and immunohistochemical methods.

In the mobilisation group, there was a significant reduction in lymphocytes (p = 0.016), macrophages (p = 0.008) and myotubules (p = 0.008) between three and 21 days. The formation of myotubules and the density of nerve fibres was significantly higher (both p = 0.016) compared with those in the immobilisation group at three days, while the density of CGRP-positive fibres was significantly lower (p = 0.016) after 21 days.

Mobilisation after contusional injury to the muscle resulted in early and increased formation of myotubules, early nerve regeneration and progressive reduction in inflammation, suggesting that it promoted a better healing response.