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ABSTRACT: OBJECTIVES:: Traumatized muscle is a complex healing environment containing cells with robust reparative and regenerative potential interacting in a cytokine milieu that influences the function and differentiation of these cells, leading to a spectrum of healing responses. In particular, BMP-4 is of interest as a potential modulator of healing, as its dysregulation has been associated with fibrosis and HO formation. We propose a descriptive study of altered BMP-4 expression traumatized muscle tissue and to evaluate its role in the fibroregulatory function of resident mesenchymal progenitor cells at the protein- and gene-expression level. METHODS:: Protein-level expression of BMP-4 from cells resident in traumatized muscle specimens was evaluated using ELISA and also using SDS-PAGE to compare BMP-4 in homogenized muscle tissue specimens. BMP-4, COMP, and osteocalcin expression localization was analyzed via immunohistochemistry. RT-PCR was performed on to evaluate fibroregulatory gene expression in MPCs after treatment with BMP-4. RESULTS:: BMP-4 was present in all traumatized muscle tissue specimens. Immunohistochemistry demonstrated that traumatized muscle fibers contained greater numbers of cells expressing BMP-4 in a more disorganized fashion compared to control samples. RT-PCR demonstrated that COMP, GDF10, and ITGB2 were up-regulated, whereas TNFA was significantly down-regulated. COMP expression was co-localized in the traumatized muscle tissue with osteocalcin. CONCLUSIONS:: BMP-4 has an effect on MPCs that appears to promote fibrotic tissue formation. These findings suggests that BMP-4, while promoting osteoinduction, may also act upon MPCs to promote formation of a fibrotic osteoinductive matrix. Thus, this signaling axis might be a potential target for HO prevention.
Journal of orthopaedic trauma 09/2012; · 1.78 Impact Factor
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ABSTRACT: The success of peripheral nerve regeneration is governed by the rate and quality of axon bridging and myelination that occurs across the damaged region. Neurite growth and the migration of Schwann cells is regulated by neurotrophic factors produced as the nerve regenerates, and these processes can be enhanced by mesenchymal stem cells (MSCs), which also produce neurotrophic factors and other factors that improve functional tissue regeneration. Our laboratory has recently identified a population of mesenchymal progenitor cells (MPCs) that can be harvested from traumatized muscle tissue debrided and collected during orthopaedic reconstructive surgery. The objective of this study was to determine whether the traumatized muscle-derived MPCs exhibit neurotrophic function equivalent to that of bone marrow-derived MSCs. Similar gene- and protein-level expression of specific neurotrophic factors was observed for both cell types, and we localized neurogenic intracellular cell markers (brain-derived neurotrophic factor and nestin) to a subpopulation of both MPCs and MSCs. Furthermore, we demonstrated that the MPC-secreted factors were sufficient to enhance in vitro axon growth and cell migration in a chick embryonic dorsal root ganglia (DRG) model. Finally, DRGs in co-culture with the MPCs appeared to increase their neurotrophic function via soluble factor communication. Our findings suggest that the neurotrophic function of traumatized muscle-derived MPCs is substantially equivalent to that of the well-characterized population of bone marrow-derived MPCs, and suggest that the MPCs may be further developed as a cellular therapy to promote peripheral nerve regeneration. Copyright © 2012 John Wiley & Sons, Ltd.
Journal of Tissue Engineering and Regenerative Medicine 05/2012; · 3.28 Impact Factor
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ABSTRACT: Peripheral nerve damage frequently accompanies musculoskeletal trauma and repair of these nerves could be enhanced by the targeted application of neurotrophic factors (NTFs), which are typically expressed by endogenous cells that support nerve regeneration. Injured muscle tissues express NTFs to promote reinnervation as the tissue regenerates, but the source of these factors from within the muscles is not fully understood. We have previously identified a population of mesenchymal progenitor cells (MPCs) in traumatized muscle tissue with properties that support tissue regeneration, and our hypothesis was that MPCs also secrete the NTFs that are associated with muscle tissue reinnervation. We determined that MPCs express genes associated with neurogenic function and measured the protein-level expression of specific NTFs with known functions to support nerve regeneration. We also demonstrated the effectiveness of a neurotrophic induction protocol to enhance the expression of the NTFs, which suggests that the expression of these factors may be modulated by the cellular environment. Finally, neurotrophic induction affected the expression of cell surface markers and proliferation rate of the MPCs. Our findings indicate that traumatized muscle-derived MPCs may be useful as a therapeutic cell type to enhance peripheral nerve regeneration following musculoskeletal injury.
Molecular Biotechnology 09/2011; 51(2):128-36. · 2.17 Impact Factor
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ABSTRACT: Heterotopic ossification (HO) occurs at a high frequency in severe orthopaedic extremity injuries; however, the etiology of traumatic HO is virtually unknown. Osteogenic progenitor cells have previously been identified within traumatized muscle. Although the signaling mechanisms that lead to this dysregulated differentiation pathway have not been identified, it is assumed that inflammation and fibrosis, which contribute to an osteoinductive environment, are necessary for the development of HO. The hypothesis of this study was that cytokines related to chronic inflammation, fibrogenesis, and osteogenesis become up-regulated following severe muscle trauma where HO forms. Classification of these cytokines by their differential expression relative to control muscle will provide guidance for further study of the mechanisms leading to HO. Real-time RT-PCR analysis revealed no significant up-regulation of cytokines typically associated with HO (e.g., BMP-4, as observed in the genetic form of HO, fibrodysplasia ossificans progressiva). Instead, the cytokine gene expression profile associated with the traumatized muscle included up-regulation of cytokines associated with osteogenesis and fibrosis (i.e., BMP-1 and TGF-β(1)). Using immunohistochemistry, these cytokines were localized to fibroproliferative lesions, which have previously been implicated in HO. This study identifies other cell and tissue-level interactions in traumatized muscle that should be investigated further to better define the etiology of HO.
Journal of Orthopaedic Research 03/2011; 29(10):1613-20. · 2.81 Impact Factor
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ABSTRACT: Heterotopic ossification (HO), characterized by the formation of mature bone in the soft tissues, is a complication that can accompany musculoskeletal injury, and it is a frequent occurrence within the military population that has experienced orthopaedic combat trauma. The etiology of this disease is largely unknown. Our laboratory has developed strategies to investigate the cellular and molecular events leading to HO using clinical specimens that were obtained during irrigation and debridement of musculoskeletal injuries. Our approach enables to study (1) the cell types that are responsible for pathological transformation and ossification, (2) the cell- and tissue-level signaling that induces the pathologic transformation, and (3) the effect of extracellular matrix topography and force transduction on HO progression. In this review, we will report on our findings in each of these aspects of HO etiology and describe our efforts to recapitulate our findings in an animal model for traumatic HO.
Advances in experimental medicine and biology 01/2011; 720:39-50. · 1.09 Impact Factor
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Youngmi Ji,
Nijaguna B Prasad,
Elizabeth A Novotny,
Sukhbir Kaur,
Abdel Elkahloun,
Yidong Chen,
Rui-Zhu Zhang,
Mon-Li Chu,
Sunita K Agarwal,
Stephen J Marx,
Francis S Collins,
Settara C Chandrasekharappa
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ABSTRACT: Multiple endocrine neoplasia type 1 (MEN1) is an autosomal dominant familial cancer syndrome characterized primarily by endocrine tumors of the parathyroids, anterior pituitary, and enteropancreatic endocrine tissues. Affected individuals carry a germ-line loss-of-function mutation of the MEN1 gene, and tumors arise after loss of the second allele. Homozygous loss of Men1 in the germ line of mice results in early embryonic lethality, with defective development of neural tube, heart, liver, and craniofacial structures. We generated immortalized wild-type (WT) and menin-null mouse embryo fibroblast (MEF) cell lines and evaluated their characteristics, including global expression patterns. The WT and menin-null cell lines were aneuploid, and the nulls did not display tumorigenic characteristics in soft agar assay. Expression arrays in menin-null MEFs revealed altered expression of several extracellular matrix proteins that are critical in organogenesis. Specifically, transcripts for fibulin 2 (Fbln2), periostin (Postn), and versican [chondroitin sulfate proteoglycan (Cspg2)], genes critical for the developing heart and known to be induced by transforming growth factor-beta (TGF-beta), were decreased in their expression in menin-null MEFs. Fbln2 expression was the most affected, and the reduction in menin-null MEFs for Fbln2, Postn, and Cspg2 was 16.18-, 5.37-, and 2.15-fold, respectively. Menin-null MEFs also showed poor response to TGF-beta-induced Smad3-mediated transcription in a reporter assay, supporting a role for menin in this pathway. Postn and Cspg2 expression in WT, unlike in null MEFs, increased on TGF-beta treatment. The expression changes associated with the loss of the tumor suppressor menin provide insights into the defective organogenesis observed during early embryonic development in Men1-null mouse embryos.
Molecular Cancer Research 11/2007; 5(10):1041-51. · 4.29 Impact Factor
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Sunita K Agarwal,
A Lee Burns,
Karen E Sukhodolets,
Patricia A Kennedy,
Victor H Obungu,
Alison B Hickman,
Michael E Mullendore,
Ira Whitten,
Monica C Skarulis,
William F Simonds, [......],
Steven K Libutti,
H Richard Alexander,
Aniello Cerrato,
Michael J Parisi,
Sonia Santa Anna-A,
Brian Oliver,
Settara C Chandrasekharappa,
Francis S Collins,
Allen M Spiegel,
Stephen J Marx
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ABSTRACT: Multiple endocrine neoplasia type 1 (MEN1), among all syndromes, causes tumors in the highest number of tissue types. Most of the tumors are hormone producing (e.g., parathyroid, enteropancreatic endocrine, anterior pituitary) but some are not (e.g., angiofibroma). MEN1 tumors are multiple for organ type, for regions of a discontinuous organ, and for subregions of a continuous organ. Cancer contributes to late mortality; there is no effective prevention or cure for MEN1 cancers. Morbidities are more frequent from benign than malignant tumor, and both are indicators for screening. Onset age is usually earlier in a tumor type of MEN1 than of nonhereditary cases. Broad trends contrast with those in nonneoplastic excess of hormones (e.g., persistent hyperinsulinemic hypoglycemia of infancy). Most germline or somatic mutations in the MEN1 gene predict truncation or absence of encoded menin. Similarly, 11q13 loss of heterozygosity in tumors predicts inactivation of the other MEN1 copy. MEN1 somatic mutation is prevalent in nonhereditary, MEN1-like tumor types. Compiled germline and somatic mutations show almost no genotype/phenotype relation. Normal menin is 67 kDa, widespread, and mainly nuclear. It may partner with junD, NF-kB, PEM, SMAD3, RPA2, FANCD2, NM23beta, nonmuscle myosin heavy chain II-A, GFAP, and/or vimentin. These partners have not clarified menin's pathways in normal or tumor tissues. Animal models have opened approaches to menin pathways. Local overexpression of menin in Drosophila reveals its interaction with the jun-kinase pathway. The Men1+/- mouse has robust MEN1; its most important difference from human MEN1 is marked hyperplasia of pancreatic islets, a tumor precursor stage.
Annals of the New York Academy of Sciences 05/2004; 1014:189-98. · 3.15 Impact Factor
