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Localized Osteoarthritis Disease-Modifying Changes due to Intra-articular Injection of Micronized Dehydrated Human Amnion/Chorion Membrane

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Abstract

Osteoarthritis (OA) is the leading cause of joint disability, and there are no FDA-approved disease-modifying drugs. The intra-articular delivery of micronized dehydrated human amnion/chorion membrane (AmnioFix, MiMedx, GA) has been shown to have a chondro-protective effect on articular cartilage in the medial meniscus transection (MMT) pre-clinical model of OA and has entered human clinical trials. AmnioFix is a well-characterized extracellular matrix (ECM)-derived therapeutic that contains hundreds of bioactive molecules, but little is known about its therapeutic mechanism in OA. The objective of this study was to elucidate, via local gene expression analysis, the molecular mechanisms of action of AmnioFix during OA development and progression. Lewis rats underwent MMT surgery, and AmnioFix or saline was injected intra-articularly 24 h post-surgery. At 5, 7, and 21 days post-surgery, articular cartilage, synovial membrane, and osteophyte tissues from multiple regions were collected and analyzed by microarray RT-PCR. Results demonstrated regional variation in the effects of amnion treatment on gene expression. Although gene expression was unaltered in articular cartilage and osteophyte tissue, pro- and anti-inflammatory markers were more highly expressed in the medial synovial membrane for the AmnioFix treatment group compared to the saline and naïve control groups. These data suggest that the previously observed chondro-protective effects of AmnioFix treatment may be mediated via alteration of the synovial membrane microenvironment. This work is the first to provide insight into the mechanisms of action of AmnioFix within the articular joint space.

Lay Summary

AmnioFix (MiMedx, Marietta, GA), obtained from placenta, is a promising candidate for treatment of osteoarthritis (OA). AmnioFix has previously demonstrated protection of the cartilage on joint surfaces in an animal model of OA. However, this is the first study to elucidate the mechanisms of action of AmnioFix on multiple tissues within the joint space. Our results demonstrated that AmnioFix acts primarily through the synovial membrane, where it induces the expression of immunomodulatory markers. We suggest that the immunomodulatory properties of AmnioFix influence the microenvironment of the synovial membrane, thereby affecting joint homeostasis and inducing a chondro-protective effect on the articular cartilage. Future work should investigate the phenotype/s of cells recruited to the synovium following AmnioFix treatment, as well as the interaction of AmnioFix with synovial tissue-resident cells. Furthermore, the effects of AmnioFix on other joint disorders such as rheumatoid arthritis, a disease involving inflammation of the synovium, warrants further investigation.

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Acknowledgments

The authors would like to acknowledge MidMedx (Marietta, GA) for providing AmnioFix from five donors for this study, and Clinton Smith (from Project ENGAGES, Georgia Institute of Technology) for assistance with sectioning of histological samples. The authors would also like to acknowledge the Genomics core and the Histology core at the Georgia Institute of Technology, as well as the National Science Foundation Graduate Research Fellowship Program (# DGE-1148903).

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Authors and Affiliations

  1. Wallace H. Coulter Department of Biomedical Engineering, Georgia Institute of Technology, Atlanta, GA, USA

    Giuliana E. Salazar-Noratto & Catriana C. Nations

  2. Parker H. Petit Institute for Bioengineering and Biosciences, Georgia Institute of Technology, 315 Ferst Dr. NW, Atlanta, GA, 30332, USA

    Giuliana E. Salazar-Noratto & Robert E. Guldberg

  3. Woodruff School of Mechanical Engineering, Georgia Institute of Technology, Atlanta, GA, USA

    Hazel Y. Stevens & Robert E. Guldberg

Authors
  1. Giuliana E. Salazar-Noratto
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  2. Catriana C. Nations
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  3. Hazel Y. Stevens
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  4. Robert E. Guldberg
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Corresponding author

Correspondence to Robert E. Guldberg.

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Conflict of Interest

Authors Giuliana E. Salazar-Noratto, Catriana C. Nations, and Hazel Y. Stevens have no conflicts to declare. Author Robert E. Guldberg has consulted for MiMedx and owns stock options in the company.

Ethical Approval

All applicable international, national, and/or institutional guidelines for the care and use of animals were followed. All procedures performed in studies involving animals were in accordance with the ethical standards of the Georgia Institute of Technology Institutional Animal Care and Use Committee (IACUC protocol #A14023).

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Supplementary Figure 1

Normalized (to control) relative expression (ΔΔCt) of genes significantly different, in the medial articular cartilage, between saline and AmnioFix groups; graphed as means ± SEM. ** indicates significant difference between saline and AmnioFix group for the specific timepoint at p value < 0.01 and *** at p value < 0.001. ### indicates time differences with day 7 for the specified treatment group at p value < 0.001. (PNG 62 kb)

High Resolution Image (TIF 119 kb)

Supplementary Figure 2

Relative expression of genes (ΔCt) of the medial side of the articular cartilage which showed significant differences between all groups (control, saline and AmnioFix), graphed as means ± SEM. * indicates significant difference with the control group for the specified timepoint at p value < 0.05, ** at p value < 0.01, and *** at p value < 0.001. $ indicates significant timepoint difference within group at p value < 0.05, $$ at p value < 0.01, and $$$ at p value < 0.01. (PNG 130 kb)

High Resolution Image (TIF 161 kb)

Supplementary Figure 3

Normalized (to control) relative expression (ΔΔCt) of genes associated with (pro- and anti-)inflammation that have significantly different expression in the medial synovium upon treatment with AmnioFix, graphed as means ± SEM. * indicates significant difference between saline and AmnioFix group for the specific timepoint at p value < 0.05, ** at p value < 0.01, and *** at p value < 0.001. # indicates time differences with day 5 for the specified treatment group at p value < 0.05, ## at p value < 0.01, and ### at p value < 0.001. % indicated time differences with day 7 for the specified treatment group at p value < 0.05, %% at p value < 0.01, and %%% at p value < 0.001. (PNG 66 kb)

High Resolution Image (TIF 138 kb)

Supplementary Figure 4

Normalized (to control) relative expression (ΔΔCt) of (A) genes associated with ECM composition and (B) genes associated with cell death that have significantly different expression in the medial synovium upon treatment with AmnioFix, graphed as means ± SEM. * indicates significant difference between saline and AmnioFix group for the specific timepoint at p value < 0.05, ** at p value < 0.01, and *** at p value < 0.001. # indicates time differences with day 5 for the specified treatment group at p value < 0.05, ## at p value < 0.01, and ### at p value < 0.001. % indicated time differences with day 7 for the specified treatment group at p value < 0.05, %% at p value < 0.01, and %%% at p value < 0.001. (PNG 44 kb)

High Resolution Image (TIF 109 kb)

Supplementary Figure 5

Normalized (to control) relative expression (ΔΔCt) of genes associated with ECM remodeling that have significantly different expression in the medial synovium upon treatment with AmnioFix, graphed as means ± SEM. * indicates significant difference between saline and AmnioFix group for the specific timepoint at p value < 0.05, ** at p value < 0.01, and *** at p value < 0.001. # indicates time differences with day 5 for the specified treatment group at p value < 0.05, ## at p value < 0.01, and ### at p value < 0.001. % indicated time differences with day 7 for the specified treatment group at p value < 0.05, %% at p value < 0.01, and %%% at p value < 0.001. (PNG 49 kb)

High Resolution Image (TIF 132 kb)

Supplementary Figure 6

Relative expression (ΔCt) of genes of the medial side of the synovial membrane which showed significant differences between the control and the operated legs (MMT surgery), regardless of treatment. * indicates significant difference with the control group for the specified timepoint at p value < 0.05, ** at p value < 0.01, and *** at p value < 0.001. # indicates significant difference with day 5 within the group at p value < 0.05, ## at p value < 0.01, and ### at p value < 0.001. $ indicates significant difference with day 7 within the group at p value < 0.05, $$ at p value < 0.01, and $$$ at p value < 0.01. (PNG 72 kb)

High Resolution Image (TIFF 93 kb)

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Salazar-Noratto, G.E., Nations, C.C., Stevens, H.Y. et al. Localized Osteoarthritis Disease-Modifying Changes due to Intra-articular Injection of Micronized Dehydrated Human Amnion/Chorion Membrane. Regen. Eng. Transl. Med. 5, 210–219 (2019). https://doi.org/10.1007/s40883-018-0087-6

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