2017 ISAKOS Biennial Congress ePoster #305
Human Subacromial Bursa Cells Display Superior Engraftment Vs. Bone Marrow Stromal Cells in Murine Tendon Repair
Felix Dyrna, MD, Münster GERMANY
Leo Pauzenberger, MD, Vienna AUSTRIA
Philip Zakko, BS, Farmington, CT UNITED STATES
Mary Beth McCarthy, BA, Farmington, United States of Ame UNITED STATES
David W. Rowe, MD, Farmington, CT UNITED STATES
Augustus D. Mazzocca, MS, MD, Farmington, CT UNITED STATES
Nathaniel Dyment, PhD, Farmington, CT UNITED STATES
Uconn Health Center, Farmington, CT, UNITED STATES
FDA Status Cleared
The subacromial bursa, while often removed during rotator cuff repair, may harbor a more suitable cell source for tendon repair than BMSCs, as they display superior engraftment and survival in tendon tissue.
Today bone marrow aspirate (BMA) is a primary source for cell-based therapies. But also, cells within peritendinous structures, such as the paratenon and surrounding bursa, contribute to the tendon healing response and offer promising cell populations for cell-based repair strategies. However, studies have not compared the efficacy of this cell population to the gold standard bone marrow stromal cells (BMSCs) in improving tendon repair.
Subacromial bursa and BMA was harvested from the same donor (N=5) during primary rotator cuff repair. Cells were expanded to P1-P2 in a 5% O2 incubator and transfected with an ubiquitin-mcherry lentiviral vector. In Vivo Model. On day of surgery, cells were incorporated into fibrin gels and implanted into full-length, central-width patellar tendon defects of NSG mice (n=70). Control mice received a defect plus fibrin without cells in one limb and defect only in the other (n=40). Mice were harvested at 1, 2, 5, and 8 weeks for analyses. In Vitro Model. Fibrin cell gel constructs were made in the FlexCell Tissue Train system. Gels were cultured either in ambient or low (5%) O2 for 14 days prior to analysis. Immunohistochemistry. Limbs were fixed, decalcified, and frozen embedded. Serial sections (7-8µm) were made at 5 different. Slides were stained for 1) collagen I and collagen III, 2) tenascin C (TNC) and EdU, and 3) F4/80 and collagen IV. All samples were scanned on the Prairie Ultima IV multiphoton microscope prior to IHC.
In Vivo Model. Based on two photon 3D imaging of the entire defect space, the bursa cells showed greater engraftment into the host tissue. The engrafted cells also took on the 3D morphology of the surrounding tissue as cells that infiltrated the adjacent tendon struts arranged in linear arrays similar to native tendon fibroblasts. In addition, bursa cells displayed greater survivability with 82% of limbs still containing cells at 8 weeks vs. only 50% of BMSC limbs. Implantation of human cells also stimulated a greater response from the host cells as the healing bridge that forms over the anterior surface of the tendon was greater in the cell groups than no-cell controls. However, the number of EdU+ cells was not significantly different between the groups. Finally, the human cells expressed higher levels of collagen I, collagen III, and TNC within the fibrin implant compared to surrounding host tissue. In Vitro Model. The BMSC gels displayed greater compaction than the Bursa gels (89±4% vs 74±3%). The bursa cells assembled thicker collagen matrix that displayed higher coherency (i.e., greater isotropy) than the BMSC gels. Additionally, gels cultured at 5% O2 yielded thicker and more organized than ambient O2 gels.
We compared the ability of bursa-derived cells to produce organized matrix and engraft into host tissue compared with the gold standard cell source for tendon repair, BMSCs. It was critical for this study to isolate both cell populations from the same patient to minimize the effect of patient differences. The stable transfection of the Ub-cherry lentivirus in combination with the 3D two photon imaging of the entire defect volume allowed us to trace the fate of the donor cells reliably over several weeks. The bursa cells displayed superior engraftment and survival in the patellar tendon implants while also synthesizing greater amounts of organized collagen in the in vitro gels.