Background:
Poor healing of the tendon-bone interface after rotator cuff repair is one of the main causes of surgical failure. Previous studies demonstrated that demineralized cortical bone (DCB) ...could improve healing of the enthesis.
Purpose:
To evaluate the outcomes of hierarchically demineralized cortical bone (hDCB) coated with stem cell–derived extracellular matrix (hDCB-ECM) in the repair of the rotator cuff in a rabbit model.
Study Design:
Controlled laboratory study.
Methods:
Tendon-derived stem cells (TDSCs) were isolated, cultured, and identified. Then, hDCB was prepared by the graded demineralization procedure. Finally, hDCB-ECM was fabricated via 2-week cell culture and decellularization, and the morphologic features and biochemical compositions of the hDCB-ECM were evaluated. A total of 24 rabbits (48 samples) were randomly divided into 4 groups: control, DCB, hDCB, and hDCB-ECM. All rabbits underwent bilateral detachment of the infraspinatus tendon, and the tendon-bone interface was repaired with or without scaffolds. After surgery, 8 rabbits were assessed by immunofluorescence staining at 2 weeks, and the others were assessed by micro–computed tomography (CT) examination, immunohistochemical staining, histological staining, and biomechanical testing at 12 weeks.
Results:
TDSCs were identified to have universal stem cell characteristics including cell markers, clonogenicity, and multilineage differentiation. The hDCB-ECM contained 3 components (bone, partial DCB, and DCB coated with ECM) with a gradient of calcium and phosphorus elements, and the ECM had stromal cell-derived factor 1, biglycan, and fibromodulin. Macroscopic observations demonstrated the absence of infection and rupture around the enthesis. The results of immunofluorescence staining showed that hDCB-ECM promoted stromal cell recruitment. Results of micro-CT analysis, immunohistochemical staining, and histological staining showed that hDCB-ECM enhanced bone and fibrocartilage formation at the tendon-bone interface. Biomechanical analysis showed that the hDCB-ECM group had higher ultimate tensile stress and Young modulus than the DCB group.
Conclusion:
The administration of hDCB-ECM promoted healing of the tendon-bone interface.
Clinical Relevance:
hDCB-ECM could provide useful information for the design of scaffolds to repair the tendon-bone interface, and further studies are needed to determine its effectiveness.
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Gels derived from decellularized small intestinal submucosa (SIS) have been used to repair ischemic myocardium and deliver protein drug. However, their material properties and effects ...on cell behavior are not well understood, in part because of the difficulty of gelling in vitro. In this study, soluble SIS matrix, which was easily handled and could effectively gel, was successfully prepared using a modified method. Fourier transform infrared spectroscopy confirmed that the SIS gel contained not only collagen but also sulfated glycosaminoglycans (sGAGs). Interestingly, the sustained release of vascular endothelial growth factor and basic fibroblast growth factor within the SIS gel was detected, and no initial burst release was observed. The SIS gel was more capable of evoking neovascularization than collagen type I gel, as determined by tube formation experiments in human umbilical vein endothelial cells, the mouse aortic ring assay, and animal experiments. The upregulated expression of kinase insert domain receptor (KDR), Notch1, and Ang2, the key genes in angiogenesis that were evaluated in HUVECs seeded on the SIS gel, confirmed that angiogenesis bioactive factors contained in the SIS gel are indeed active and effective. The SIS gel significantly promoted neovascularization compared to the collagen type I gel in vivo. Histology revealed adequate host tissue response in engraftment both types of gels. Together, these data demonstrate that the SIS gel is a promising and attractive candidate for tissue engineering, especially in promoting vessel formation.
The material properties of small intestinal submucosa (SIS) gel and the effect of these properties upon cell behavior are not well understood, in part due to the difficulty of gelling in vitro. In this study, soluble SIS matrix, which was easily handled and gelled was prepared using modified method. The material properties and biocompatibility of SIS gel were explored. The sustained release of growth factors from this gel was observed along with its degradation in vitro. The results demonstrate that the SIS gel promote angiogenesis in vitro and in vivo. The SIS gel biological properties suggest that the constituent ECM molecules released from the gel remain activity. These findings suggested that the SIS gel was a promising candidate for tissue engineering, especially in promoting vessel formation.
Abstract Cell-based tissue engineered tendons have potential to improve clinical outcomes following rotator cuff repair, especially in large or massive rotator cuff tears, which pose a great clinical ...challenge. The aim of this study was to develop a method of constructing a functional engineered tendon patch for rotator cuff repair with cyclic mechanical stimulation. Decellularized tendon slices (DTSs) were seeded with BMSCs and subjected to cyclic stretching for 1, 3, or 7 days. The mechanical properties, morphologic characteristics and tendon-related gene expression of the constructs were investigated. Viable BMSCs were observed on the DTS after 7 days. BMSCs penetrated into the DTSs and formed dense cell sheets after 7 days of mechanical stretching. Gene expression of type I collagen, decorin, and tenomodulin significantly increased in cyclically stretched BMSC-DTS constructs compared with the unstrained control group ( P < 0.05). The ultimate tensile strength and stiffness of the cyclically stretched tendon constructs were similar to the unstrained control group ( P > 0.05). In conclusion, mechanical stimulation of BMSC-DTS constructs upregulated expression of tendon-related proteins, promoted cell tenogenic differentiation, facilitated cell infiltration and formation of cell sheets, and retained mechanical properties. The patch could be used as a graft to enhance the surgical repair of rotator cuff tears.
Biglycan (BGN) has been identified as one of the critical components of the tendon‐derived stem cells (TDSCs) niche and may be related to tendon formation. However, so far, no study has demonstrated ...whether the soluble BGN could induce the tenogenic differentiation of TDSCs in vitro. The aim of this study was to investigate the effect of BGN on the tenogenic differentiation of TDSCs. The proliferation and tenogenic differentiation of TDSCs exposed to different concentrations of BGN (0, 50, 100, and 500 ng/ml) were determined by the live/dead cell staining assay, CCK‐8 assay, quantitative real‐time polymerase chain reaction (qRT‐PCR), and western blot analysis. The BGN signaling pathway of TDSCs (with and without 50 ng/ml of BGN) was determined by western blot analysis and qRT‐PCR analysis. At a concentration of 50 ng/ml, BGN increased the expression of the tenogenic markers THBS‐4 and TNMD at both the messenger RNA (mRNA) and protein levels. Meanwhile, 50 ng/ml of BGN inhibited the expression of the chondrogenic and osteogenic markers SOX9, ACN, and RUNX2 at both the mRNA and protein levels. Moreover, BGN (50 ng/ml) affected the expression of the components of the extracellular matrix of TDSCs. Additionally, BGN activated the Smad1/5/8 pathway as indicated by an increase in phosphorylation and demonstrated by inhibition experiments. Upregulation in the gene expression of BMP‐associated receptors (BMPRII, ActR‐IIa, and BMPR‐Ib) and Smad pathway components (Smad4 and 8) was observed. Taken together, BGN regulates tenogenic differentiation of TDSCs via BMP7/Smad1/5/8 pathway and this regulation may provide a basic insight into treating tendon injury.
Biglycan (BGN) regulates tenogenic differentiation of tendon‐derived stem cells (TDSCs) via BMP7/Smad1/5/8 pathway and this regulation may provide a basic insight into treating tendon injury.
Abstract Porcine small intestinal submucosa (SIS) has been widely used in repairing various tissues and organs. Despite this, some SIS products have the capacity to cause variable inflammatory ...responses after implantation resulting in severe adverse effects due to porcine cell existence. In this study, we described a multi-step method including mechanical disassociation, degrease, enzyme digestion, detergent treatment, freeze-drying and sterilization by irradiation for preparation of SIS. The efficacy of acellularization was evaluated by histological observation and the content of porcine immunoreceptor DAP12 gene. The change of growth factors contents within SIS accompanying with decellularization was quantitatively assessed by ELISA. Inflammatory reaction of SIS implanted subcutaneously in a rat was investigated. The histological examination revealed no remaining cells after enzyme digestion. Moreover, qPCR analysis demonstrated that the content of a porcine immunoreceptor gene DAP12 DNA in final SIS product (SISv) was only 1.05% of that in SIS samples (SISi) prepared by mechanical disassociation. Degrease with methanol/chloroform dramatically reduced the contents of VEGF, b-FGF, TGF-β, and TNF-α within SISii, but further treatment could not significantly reduced the contents of growth factors. SIS implanted into rats showed that inflammatory cells was more accumulated surrounded to SISi at 1, and 2 weeks, but reduced in SISv samples. The degree of inflammatory reaction for SISv was significantly less than that of SISi.
Background:
Achilles tendon (AT) defects frequently occur in trauma and chronic injuries. Currently, no method can satisfactorily reconstruct the AT with completely restored function.
Purpose:
To ...evaluate the postoperative outcomes of AT defect reconstruction with decellularized bovine tendon sheets (DBTSs) in a rabbit model.
Study Design:
Controlled laboratory study.
Methods:
DBTSs were prepared from bovine tendons after compression, decellularization, antigen extraction, freeze drying, and sterilization. Platelet-rich plasma (PRP) was obtained by differential centrifugation. Sixty-three rabbits were used in this study, and the AT defect model was created bilaterally. All rabbits were divided into 3 groups (n = 21). In the DBTS group and the DBTS + PRP group, 2-cm-long AT was excised and reconstructed by DBTSs or PRP-treated DBTSs. In the control group, the rabbits underwent AT transection, and stumps were sutured. After surgery, all rabbits were assessed by ultrasonography and magnetic resonance imaging and then sacrificed for histological examination and biomechanical testing at 4, 8, or 12 weeks.
Results:
Gross observations demonstrated the absence of immunologic incompatibility and rejection. Histological examination showed that DBTSs promoted host cell infiltration and new fibrous tissue integration as compared with the control group. In each group, there was an AT-like structure formation and aligned collagen fiber deposition at 12 weeks. Mechanical properties of the reconstructed AT were not significantly different among the 3 groups at 4, 8, and 12 weeks after surgery (P > .05). Ultrasonography and magnetic resonance imaging results illustrated that the reconstructed AT from each group maintained remodeling, and there was no significant difference in the echogenicity scoring (P > .05) and percentages of good and excellent (P > .05) among the 3 groups.
Conclusion:
DBTSs, which retain the native tendon structure and bioactive factors, had the ability to remodel and integrate into the rabbit AT and improve the healing process.
Clinical Relevance:
DBTSs could serve as an effective bioscaffold to reconstruct AT defects.
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