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ECM Degradation, Matricryptic Peptides, and Stem Cell Recruitment

Agrawal, Vineet (2011) ECM Degradation, Matricryptic Peptides, and Stem Cell Recruitment. Doctoral Dissertation, University of Pittsburgh.

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    Abstract

    Biologic scaffolds composed of extracellular matrix (ECM) have been used to promote site-specific, functional remodeling of tissue in both preclinical animal models and human clinical applications. Although the mechanisms of action of ECM scaffolds are not completely understood, proteolytic degradation of the ECM scaffold and subsequent progenitor cell recruitment are thought to be important mediators of the constructive remodeling process.Proteolytic degradation of the ECM scaffolds results in the generation and release of cryptic peptides with novel bioactive properties not associated with their parent molecules such as angiogenic, antimicrobial, mitogenic, and chemotactic properties. While previous studies have suggested that degradation products of ECM scaffolds are chemotactic for progenitor cells in vitro, the present thesis expands upon these findings in vivo.In a non-regenerating model of mid-second phalanx digit amputation, treatment with ECM degradation rpodcuts resulted in the accumulation of a heterogeneous population of cells with in vitro differentiation potential along osteogenic, adipogenic, and neuroectodermal lineages. Focusing specifically on the Sox2+ population of cells found at the site of injury, work in the present thesis showed that Sox2+ cells co-express bone marrow and periosteal stem cell markers CD90 and Sca1, but not dermal stem cell marker CD133 or circulating stem cell marker c-kit (CD117). Additionally, bone marrow chimeric studies utilizing wild type C57/BL6 and Sox2 eGFP/+ mice showed that the Sox2+ cells are not derived from the bone marrow, but more likely from a local tissue source such as the periosteum. Fractionation of the ECM degradation products resulted in the identification of a highly conserved cryptic peptide derived from the C-terminal telopeptide of the collagen type IIIα molecule with chemotactic activity for multiple progenitor cells in vitro, IAGVGGEKSGGF. Administration of the cryptic peptide in a model of digit amputation resulted in the accumulation of Sox2+, Sca1+, Lin- cells at the site of amputation. Peptide treatment also resulted in the formation of a bone nodule at the site that coincided with the spatial location of Sox2+ cells. In vitro, the peptide accelerated osteogenesis of mesenchymal stem cells and increased the expression of osteogenic and chondrogenic genes.The result of this body of work shows that degradation products of ECM scaffolds contain cryptic peptides with the ability to influence chemotaxis and differentiation of progenitor cells in vitro and in vivo. The ability to influence stem cell phenotype and fate may be useful in designing new therapies for regenerative medicine approaches to complex, composite tissue reconstruction. Additionally, the findings of the present thesis may serve as the basis for future studies investigating the importance of ECM degradation in the downstream constructive remodeling events at a site of ECM implantation in soft tissue models of injury.


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    Details

    Item Type: University of Pittsburgh ETD
    Creators/Authors:
    CreatorsEmailORCID
    Agrawal, Vineetvineet.agrawal85@gmail.com
    ETD Committee:
    ETD Committee TypeCommittee MemberEmailORCID
    Committee ChairWells, Alan
    Committee MemberAlmarza, Alejandro
    Committee MemberFox, Ira J
    Committee MemberMarra, Kacey G
    Committee MemberBadylak, Stephen F
    Committee MemberOury, Timothy D
    Title: ECM Degradation, Matricryptic Peptides, and Stem Cell Recruitment
    Status: Unpublished
    Abstract: Biologic scaffolds composed of extracellular matrix (ECM) have been used to promote site-specific, functional remodeling of tissue in both preclinical animal models and human clinical applications. Although the mechanisms of action of ECM scaffolds are not completely understood, proteolytic degradation of the ECM scaffold and subsequent progenitor cell recruitment are thought to be important mediators of the constructive remodeling process.Proteolytic degradation of the ECM scaffolds results in the generation and release of cryptic peptides with novel bioactive properties not associated with their parent molecules such as angiogenic, antimicrobial, mitogenic, and chemotactic properties. While previous studies have suggested that degradation products of ECM scaffolds are chemotactic for progenitor cells in vitro, the present thesis expands upon these findings in vivo.In a non-regenerating model of mid-second phalanx digit amputation, treatment with ECM degradation rpodcuts resulted in the accumulation of a heterogeneous population of cells with in vitro differentiation potential along osteogenic, adipogenic, and neuroectodermal lineages. Focusing specifically on the Sox2+ population of cells found at the site of injury, work in the present thesis showed that Sox2+ cells co-express bone marrow and periosteal stem cell markers CD90 and Sca1, but not dermal stem cell marker CD133 or circulating stem cell marker c-kit (CD117). Additionally, bone marrow chimeric studies utilizing wild type C57/BL6 and Sox2 eGFP/+ mice showed that the Sox2+ cells are not derived from the bone marrow, but more likely from a local tissue source such as the periosteum. Fractionation of the ECM degradation products resulted in the identification of a highly conserved cryptic peptide derived from the C-terminal telopeptide of the collagen type IIIα molecule with chemotactic activity for multiple progenitor cells in vitro, IAGVGGEKSGGF. Administration of the cryptic peptide in a model of digit amputation resulted in the accumulation of Sox2+, Sca1+, Lin- cells at the site of amputation. Peptide treatment also resulted in the formation of a bone nodule at the site that coincided with the spatial location of Sox2+ cells. In vitro, the peptide accelerated osteogenesis of mesenchymal stem cells and increased the expression of osteogenic and chondrogenic genes.The result of this body of work shows that degradation products of ECM scaffolds contain cryptic peptides with the ability to influence chemotaxis and differentiation of progenitor cells in vitro and in vivo. The ability to influence stem cell phenotype and fate may be useful in designing new therapies for regenerative medicine approaches to complex, composite tissue reconstruction. Additionally, the findings of the present thesis may serve as the basis for future studies investigating the importance of ECM degradation in the downstream constructive remodeling events at a site of ECM implantation in soft tissue models of injury.
    Date: 05 August 2011
    Date Type: Completion
    Defense Date: 27 July 2011
    Approval Date: 05 August 2011
    Submission Date: 29 July 2011
    Access Restriction: 5 year -- Restrict access to University of Pittsburgh for a period of 5 years.
    Patent pending: No
    Institution: University of Pittsburgh
    Thesis Type: Doctoral Dissertation
    Refereed: Yes
    Degree: PhD - Doctor of Philosophy
    URN: etd-07292011-121054
    Uncontrolled Keywords: amputation; chemotaxis; differentiation; digit; epimorphic regeneration; extracellular matrix; progenitor cell; stem cell
    Schools and Programs: School of Medicine > Cellular and Molecular Pathology
    Date Deposited: 10 Nov 2011 14:55
    Last Modified: 06 Jun 2012 10:43
    Other ID: http://etd.library.pitt.edu/ETD/available/etd-07292011-121054/, etd-07292011-121054

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