MECHANISMS OF EQUINE PERIOSTEAL OSTEOGENIC ACTIVATION

• Sponsoring Institution: National Institute of Food and Agriculture
• Project Status: COMPLETE
• Funding Source: ANIMAL HEALTH
• Reporting Frequency: Annual
• Accession No.: 1021783
• Project Start Date: Dec 1, 2019
• Project End Date: Sep 30, 2021
• Program Code: [(N/A)]- (N/A)

Recipient Organization
UNIVERSITY OF ILLINOIS
2001 S. Lincoln Ave.
URBANA,IL 61801

Performing Department
Veterinary Research & Extension

Non Technical Summary
Periosteum is a thin layer of specialized connective tissue that covers the outer surface of bone. The progenitor cells in the inner cellular layer of periosteum are responsible for appositional bone growth during skeletal development, increasing bone girth as bones lengthen. Of more importance to clinical issues of equine bone repair, periosteal osteoprogenitors are primarily responsible for bone repair by endochondral ossification; the mechanism of bone repair in the great majority of skeletal trauma cases. All but non-displaced fractures heal by endochondral ossification and, therefore, are dependent on periosteal osteoprogenitors for successful healing. Our preliminary experiments have demonstrated that periosteal-derived cells are far less capable of in vitro osteogenic differentiation than progenitors isolated from the endosteal (bone marrow) compartment. This deficit is linked to dramatic down-regulation of Osterix and BMP-2 expression; both integral to bone formation. The proposed experiments will determine whether collagen-based substrates can prevent the ex vivo loss of osteogenic capacity in periosteal cells (Aim 1) and whether inflammatory mediators, as are released at the time of skeletal trauma, can activate periosteal cell bone formation. The outcomes of these experiments will provide valuable information on the in vitro maintenance of periosteal osteogenic capacity for tissue engineering and regenerative applications, and on the cellular mechanisms that stimulate periosteal bone formation, both in vivo and in vitro.

Animal Health Component

10%

Research Effort Categories

Basic

80%

Applied

10%

Developmental

10%

Classification

Knowledge Area (KA)	Subject of Investigation (SOI)	Field of Science (FOS)	Percent
311	3810	1030	100%

Knowledge Area
311 - Animal Diseases;

Subject Of Investigation
3810 - Horses, ponies, and mules;

Field Of Science
1030 - Cellular biology;

Keywords

periosteum

periosteal osteogenic capacity

periosteal bone formation

equine orthopaedics

Goals / Objectives
Aim 1: To determine whether collagen type I substrates maintain and/or restore osteogenic capacity of periosteal cells.Aim 2. To determine whether exposure to inflammatory cytokines stimulates BMP expression and osteogenesis in periosteal cells.

Project Methods
Please note that tissue and cell collections will be carried out on horses admitted to the VTH for euthanasia, immediately after the animals are euthanized. As a result, work with live animals is not a part of this study.Periosteum will be collected from horses euthanized for reasons other than musculoskeletal pathology. Periostral explants and isolated cells will be maintained in culture to determine the effects of substrate (native periosteal tissue vs collagen scaffolds vs cell culture plastic) and exposure to inflammatory mediators on the activity of these cells. Cell morphology, quantitative PCR and enzymatic activities will be used to assess the effects of these factors on the osteogenic phenotype and synthetic activities of periosteal cells.

Progress 12/01/19 to 09/30/21

Outputs
Target Audience:The research findings will be of interest and relevance to orthopaedic and musculoskeletal researchers and clinicians; particularly, those with an interest in fracture repair and skeletal remodelling. Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided? Nothing Reported How have the results been disseminated to communities of interest? Nothing Reported What do you plan to do during the next reporting period to accomplish the goals?We are continuing our experiments on the role of inflammatroy cytokines, BMPs and TGF-bs on periosteal osteogenic activity. We are particularly interested in investigating the possibility of a positive feedback loop between cytokine effects and downstream induction of BMPs/TGF-bs.

Impacts
What was accomplished under these goals? The experiments in Aim 1 have demonstrated that mantaining periostal cells as tissue explants, as opposed to isolated cels, slows the loss of the differentiated phenotype moderately but does not prevent de-differentation. Culturing cells on collagen-based substrate improves cell adhesion to the surface but does not protect against phenotypic loss. Experiments in Aim 2 have demonstrated an up-regulation in ostegenic factors after periostal cell exposure to inflammatroy cytokines. In parallel, we have been assessing the effect of exogenous BMP-2 and other putative periosteal growth factors, such as PgE2, PDGF, and IGF-1, on periosteal ostegenesis. We have found tat TGF-b ligands (particularly TGF-b3) are strong indicers of osteogenic genes in periosteal cells that have otherwise lost their phenotypic profle in vitro.

Publications

• Type: Conference Papers and Presentations Status: Under Review Year Published: 2021 Citation: An abstract has been published in the Proceedings of the 2021 Veterinary Orthopedic Society meeting and a second abstract will be published in the Proceedings of the 2022 Annual Scientific Meeting of the Veterinary Orthopedic Society. A manuscript detailing the results of this study is currently being prepared.

Progress 12/01/19 to 09/30/20

Outputs
Target Audience:The research findings will be of interest and relevance to orthopaedic and musculoskeletal researchers and clinicians;particularlythose with an interest in fracture repair and skeletal remodelling. Changes/Problems:We have added a focus on the role of TGF-b signaling in the induction of periosteal osteogenesis, as a consequence of ourearly experimental results. What opportunities for training and professional development has the project provided? Nothing Reported How have the results been disseminated to communities of interest? Nothing Reported What do you plan to do during the next reporting period to accomplish the goals?We are continuing our experiments on the role of inflammatroy cytokines, BMPs and TGF-bs on periosteal osteogenic activity.We are particularly interested in investigating the possibility of a positive feedback loop between cytokine effects anddownstream induction of BMPs/TGF-bs.

Impacts
What was accomplished under these goals? The experiments in Aim 1 have demonstrated that mantaining periostal cells as tissue explants, as opposed to isolated cells,slows the loss of the differentiated phenotype moderately but does not prevent de-differentation. Culturing cells on collagen-basedsubstrate improves cell adhesion to the surface but does not protect against phenotypic loss. Experiments in Aim 2 have demonstrated an up-regulation in ostegenic factors after periostal cell exposure to inflammatorycytokines. In parallel, we have been assessing the effect of exogenous BMP-2 and other putative periosteal growth factors,such as PgE2, PDGF, and IGF-1, on periosteal ostegenesis. We have found that TGF-b ligands (particularly TGF-b3) arestrong indicers of osteogenic genes in periosteal cells that have otherwise lost their phenotypic profle in vitro.

Publications

• Type: Conference Papers and Presentations Status: Under Review Year Published: 2021 Citation: If accepted, the abstract will be published in the Proceedings of the 2021 Annual Scientific Meeting of the Veterinary Orthopedic Society and will also be published in a 2021 volume of Veterinary and Comparative Orthopedics and Traumatology.