Abstract
BackgroundMicrofracture is one of the most widely used techniques for the repair of articular cartilage. However, microfracture often results in filling of the chondral defect with fibrocartilage, which exhibits poor durability and sub-optimal mechanical properties. Stromal cell-derived factor-1 (SDF-1) is a potent chemoattractant for mesenchymal stem cells (MSCs) and is expressed at high levels in bone marrow adjacent to developing cartilage during endochondral bone formation. Integrating SDF-1 into an implantable collagen scaffold may provide a chondro-conductive and chondro-inductive milieu via chemotaxis of MSCs and promotion of chondrogenic differentiation, facilitating more robust hyaline cartilage formation following microfracture.ObjectiveThis work aimed to confirm the chemoattractive properties of SDF-1 in vitro and develop a one-step method for incorporating SDF-1 in vivo to enhance cartilage repair using a rat osteochondral defect model.MethodsBone marrow-derived MSCs (BMSCs) were harvested from the femurs of Sprague–Dawley rats and cultured in low-glucose Dulbecco’s modified Eagle’s medium containing 10% fetal bovine serum, with the medium changed every 3 days. Passage 1 MSCs were analyzed by flow cytometry with an S3 Cell Sorter (Bio-Rad). In vitro cell migration assays were performed on MSCs by labeling cells with carboxyfluorescein diacetate, succinimidyl ester (CFDA-SE; Bio-Rad). For the microfracture model, a 1.6-mm-diameter osteochondral defect was created in the femoral trochleae of 20 Sprague–Dawley rats bilaterally until bone marrow spillage was seen under saline irrigation. One knee was chosen at random to receive implantation of the scaffold, and the contralateral knee was left unfilled as an empty control. Type I collagen scaffolds (Kensey Nash) were coated with either gelatin only or gelatin and SDF-1 using a dip coating process. The rats received implantation of either a gelatin-only scaffold (N = 10) or gelatin-and-SDF-1 scaffold (N = 10) at the site of the microfracture. Femurs were collected for histological analyses at 4- and 8-week time points post-operatively, and sections were stained with Safranin O/Fast Green. The samples were graded blindly by two observers using the Modified O’Driscoll score, a validated scoring system for chondral repair. A minimum of 10 separate grading scores were made per sample and averaged. Quantitative comparisons of cell migration in vitro were performed with one-way ANOVA. Cartilage repair in vivo was also compared among groups with one-way ANOVA, and the results were presented as mean ± standard deviation, with P-values < 0.05 considered as statistically significant.ResultsMSC migration showed a dose–response relationship with SDF-1, with an optimal dosage for chemotaxis between 10 and 100 ng/ml. After scaffold implantation, the SDF-1-treated group demonstrated complete filling of the cartilage defect with mature cartilage tissue, exhibiting strong proteoglycan content, smooth borders, and good incorporation into marginal cartilage. Modified O’Driscoll scores after 8 weeks showed a significant improvement of cartilage repair in the SDF-1 group relative to the empty control group (P < 0.01), with a trend toward improvement when compared with the gelatin-only-scaffold group (P < 0.1). No significant differences in scores were found between the empty defect group and gelatin-only group.ConclusionIn this study, we demonstrated a simple method for improving the quality of cartilage defect repair in a rat model of microfracture. We confirmed the chemotactic properties of SDF-1 on rat MSCs and found an optimized dosage range for chemotaxis between 10 and 100 ng/ml. Furthermore, we demonstrated a strategy to incorporate SDF-1 into gelatin–collagen I scaffolds in vivo at the site of an osteochondral defect. SDF-1-treated defects displayed robust hyaline cartilage resurfacing of the defect with minimal fibrous tissue, in contrast to the empty control group. The results of the in vitro and in vivo studies together suggest that SDF-1-mediated signaling may significantly improve the quality of cartilage regeneration in an osteochondral defect.
Highlights
While the etiology of joint degeneration and osteoarthritis occurs at the molecular, cellular, and tissue level, current treatments of isolated chondral defects are primarily surgical (Furman et al, 2006; Daher et al, 2009; Anderson et al, 2011)
Chemoattraction of bone marrow-derived MSCs (BMSCs) by Stromal cell-derived factor-1 (SDF-1) was tested with the Transwell culture system in vitro
To determine the optimal dosage of SDF-1 for chemoattraction of BMSCs, SDF-1 was added to the Transwell system at concentrations of 1, 10, and 100 ng/ml
Summary
While the etiology of joint degeneration and osteoarthritis occurs at the molecular, cellular, and tissue level, current treatments of isolated chondral defects are primarily surgical (Furman et al, 2006; Daher et al, 2009; Anderson et al, 2011). It is thought that microfracture alone does not recruit a sufficient amount of reparative cells and growth factors to promote adequate native tissue repair (Richter, 2009). One potential therapeutic strategy to augment the chondrogenic milieu following microfracture is by incorporating homing factors into cell-free scaffolds to promote the recruitment of MSCs to the site of injury (Mishima and Lotz, 2008; Chanda et al, 2010; Fong et al, 2011; Eseonu and De Bari, 2015; Truong et al, 2017; Lu et al, 2018). Integrating SDF-1 into an implantable collagen scaffold may provide a chondro-conductive and chondro-inductive milieu via chemotaxis of MSCs and promotion of chondrogenic differentiation, facilitating more robust hyaline cartilage formation following microfracture
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