The omentum harbors unique conditions in the peritoneal cavity to promote healing and regeneration for diaphragm muscle repair in mdx mice.

Although the primary cause of Duchenne muscular dystrophy (DMD) is a genetic mutation, the inflammatory response contributes directly to severity and exacerbation of the diaphragm muscle pathology. The omentum is a lymphoid organ with unique structural and immune functions serving as a sanctuary of hematopoietic and mesenchymal progenitors that coordinate immune responses in the peritoneal cavity. Upon activation, these progenitors expand and the organ produces large amounts of growth factors orchestrating tissue regeneration. The omentum of mdx mouse, a DMD murine model, is rich in milky spots and produces growth factors that promote diaphragm muscle regeneration. The present review summarizes the current knowledge of the omentum as an important immunologic structure and highlights its contribution to resolution of dystrophic muscle injury by providing an adequate environment for muscle regeneration, thus being a potential site for therapeutic interventions in DMD.

Omentum acts as a regulatory organ controlling skeletal muscle repair of mdx mice diaphragm.

Duchenne muscular dystrophy is a lethal X-linked muscle wasting disease due to mutations of the dystrophin gene leading to distinct susceptibility to degeneration and fibrosis among skeletal muscles. This study aims at verifying whether intense mdx diaphragm remodeling could be attributed to influences from the omentum, a lymphohematopoietic tissue rich in progenitor cells and trophic factors. Mdx omentum produces growth factors HGF and FGF and increased amounts of VEGF with pleiotropic actions upon muscular progenitors and myoblast differentiation. Histology revealed that the absence of the omentum reduced inflammation and collagen deposition in the diaphragm. The diaphragm from omentectomized mdx mice presents impaired repair with a predominance of collagen type I deposition, decreased muscle regeneration and a reduction in collagen type IV and indication of altered basal lamina integrity in the diaphragm. Omentectomy further reduced inflammatory infiltration and NFκ-B activation but a change in the pattern of muscle inflammation with low numbers of the F4/80+CD206+ M-2 macrophage subset. Although omentectomized mice had high levels of Pax7, myogenin and TNF-α, the percentage of myofibers undergoing regeneration was low thus suggesting that a lack of the omentum halts the muscle differentiation program. Such results support that omentum exerts a regulatory function inducing an inflammatory process that favors regeneration and inhibits fibrosis selectively in the diaphragm muscle thus being a potential site for therapeutic interventions in DMD.

Neoantigen Expression in Steady-State Langerhans Cells Induces CTL Tolerance.

The skin hosts a variety of dendritic cells (DCs), which act as professional APC to control cutaneous immunity. Langerhans cells (LCs) are the only DC subset in the healthy epidermis. However, due to the complexity of the skin DC network, their relative contribution to either immune activation or immune tolerance is still not entirely understood. To specifically study the function of LCs in vivo, without altering the DC subset composition in the skin, we have generated transgenic mouse models for tamoxifen-inducible de novo expression of Ags in LCs but no other langerin+ DCs. Therefore, this system allows for LC-restricted Ag presentation to T cells. Presentation of nonsecreted OVA (GFPOVA) by steady-state LCs resulted in transient activation of endogenous CTL in transgenic mice. However, when these mice were challenged with OVA by gene gun immunization in the contraction phase of the primary CTL response they did not respond with a recall of CTL memory but, instead, with robust Ag-specific CTL tolerance. We found regulatory T cells (Tregs) enriched in the skin of tolerized mice, and depletion of Tregs or adoptive experiments revealed that Tregs were critically involved in CTL tolerance. By contrast, when OVA was presented by activated LCs, a recallable CTL memory response developed in transgenic mice. Thus, neoantigen presentation by epidermal LCs results in either robust CTL tolerance or CTL memory, and this decision-making depends on the activation state of the presenting LCs.

Persistent activation of omentum influences the pattern of muscular lesion in the mdx diaphragm.

The mdx (X chromosome-linked muscular dystrophy) mouse develops a multi-staged disorder characterized by muscle degeneration and reactive fibrosis. Skeletal muscles of mdx mice are not equally susceptible to degeneration. The aim of this study was to verify whether the intense remodeling of the mdx diaphragm could be attributed to influences from the peritoneal microenvironment and omentum, a lymphohematopoietic tissue rich in progenitor cells and trophic factors. At ages corresponding to increased muscular regeneration (12 weeks) and activation of fibrosis (24 weeks), the mdx omentum exhibited (1) morphological and functional characteristics of activation with enlarged milk-spots, an accumulation of CD4(+), CD8(+) and CD19(+)B220(+) B lymphocytes; (2) the formation of clusters positive for proliferating cell nuclear antigen, mainly in B220(+)-rich areas organized in a follicular structure with a germinative center without any challenge by external antigen inducers; (3) clusters with cells positive for fibroblast growth factor-2, numerous Sca-1(+)CD3(-)CD19(-)Mac-1(-) progenitor cells and increased CD4(+), CD8(+) and CD3(+)NK1.1(+) cells in the peritoneal cavity. Omentectomy reduced areas with F4/80(+) inflammatory infiltrate the activity of matrix metalloproteases 9 and 2, collagen deposition and areas with regenerating myofibers in the diaphragm. Thus, persistent activation of the omentum influences the pattern of inflammation and regeneration of the mdx diaphragm partly via the activation of progenitor cells and the production of growth factors that influence the physiopathology of the muscular tissue remodeling.

Nicotinic acetylcholine receptor activation reduces skeletal muscle inflammation of mdx mice.

Mdx mice develop an inflammatory myopathy characterized at different ages by myonecrosis with scattered inflammatory infiltrates followed by muscular regeneration and later persistent fibrosis. This work aimed to verify the putative anti-inflammatory role of nicotinic acetylcholine receptor (nAChR) in the mdx muscular lesion. Mitigation of myonecrosis and decreased TNFα production were accompanied by increased numbers of F4/80 macrophages expressing nAChRα7. In vivo treatment with nicotine attenuated muscular inflammation characterized by reduced metalloprotease MMP-9 activity, TNFα and NFkB content and increased muscular regeneration. Our data indicate that nAChR activation influences local inflammatory responses in the muscular lesion of mdx mice.

Pattern of metalloprotease activity and myofiber regeneration in skeletal muscles of mdx mice.

Matrix metalloproteases (MMPs) are key regulatory molecules in the formation, remodeling, and degradation of extracellular matrix components in both physiological and pathological processes. Skeletal muscles of mdx dystrophic mice show distinct patterns of inflammation and regeneration, suggesting that factors within the microenvironment influence the adaptive responses of muscles with predominantly slow-twitch or fast-twitch fibers. This study aimed to verify the pattern of MMP activity in gastrocnemius, soleus, and diaphragm muscles and correlate it with the regenerative capability at distinct stages of the mdx myopathy. Marked inflammation and myonecrosis was associated with increased MMP-9 activity and TNF-alpha (tumor necrosis factor-alpha) production, whereas muscle regeneration, evidenced by NCAM (neural cell adhesion molecule) expression and MMP-2 activity, varied at different stages of the disease. Soleus muscles showed a high percentage of NCAM-positive myofibers in the early stages (2 weeks) of the disease, but they appeared in the gastrocnemius muscles at 12 weeks and in the diaphragm at 24 weeks. Increased MMP-2 activity in the diaphragm throughout all stages of the disease suggests important tissue remodeling, which is probably associated with persistent inflammation. The results indicate that the microenvironment of distinct skeletal muscle may influence a particular kinetic pattern of MMP activity, which ultimately favors persistent inflammation and myofiber regeneration at different stages of the myopathy in mdx mice.