Direct Intra-articular Injection Research Articles

Enhanced intra-articular therapy for rheumatoid arthritis using click-crosslinked hyaluronic acid hydrogels loaded with toll-like receptor antagonizing peptides

Rheumatoid arthritis (RA) is a chronic inflammatory disorder that results in the deterioration of joint cartilage and bone. Toll-like receptor 4 (TLR4) has been pinpointed as a key factor in RA-related inflammation. While Toll-like receptor antagonizing peptide 2 (TAP2) holds potential as an anti-inflammatory agent, its in vivo degradation rate hinders its efficacy. We engineered depots of TAP2 encapsulated in click-crosslinked hyaluronic acid (TAP2+Cx-HA) for intra-articular administration, aiming to enhance the effectiveness of TAP2 as an anti-inflammatory agent within the joint cavity. Our data demonstrated that FI-TAP2+Cx-HA achieves a longer retention time in the joint cavity compared to FI-TAP2 alone. Mechanistically, we found that TAP2 interacts with TLR4 on the cell membranes of inflammatory cells, thereby inhibiting the nuclear translocation of NF-κB and maintaining it in an inactive cytoplasmic state. In a rat model of RA, the TAP2+Cx-HA formulation effectively downregulated the inflammatory cytokines TNF-α and IL-6, while upregulating the anti-inflammatory cytokine IL-10 and the therapeutic protein 14-3-3ζ. This led to a more rapid restoration of cartilage thickness, increased deposition of glycosaminoglycans, and new bone tissue formation in the regenerated cartilage, in comparison to a single TAP2 treatment after a six-week period. Our results suggest that TAP2+Cx-HA could serve as a potent intra-articular treatment for RA, offering both symptomatic relief and promoting cartilage regeneration. This innovative delivery system holds significant promise for improving the management of RA and other inflammatory joint conditions. Statement of significanceIn this study, we developed a therapy by creating toll-like receptor 4 (TLR4)-antagonizing peptide (TAP2)-loaded click-crosslinked hyaluronic acid (TAP2+Cx-HA) depots for direct intra-articular injection. Our study demonstrates that FI-TAP2+Cx-HA exhibits a more than threefold longer lifetime in the joint cavity compared to FI-TAP2 alone. Furthermore, we found that TAP2 binds to TLR4 and masks the nuclear localization signals of NF-κB, leading to its sequestration in an inactive state in the cytoplasm. In a rat model of RA, TAP2+Cx-HA effectively suppresses inflammatory molecules, specifically TNF–α and IL-6, while upregulating the anti-inflammatory cytokine IL-10 and the therapeutic protein 14-3-3ζ. This resulted in faster regeneration of cartilage thickness, increased glycosaminoglycan deposits in the regenerated cartilage, and a twofold increase in new bone tissue formation compared to a single TAP2 treatment.

USING MESENCHYMAL STROMAL CELLS AND THEIR SECRETOME TO IMPROVE OSTEOCHONDRAL REPAIR IN A MURINE MODEL

Osteochondral injuries are a recognised factor in the development of osteoarthritis (OA). Mesenchymal stromal cells (MSCs) represent a promising biological therapeutic option as an OA-modifying treatment, and they also secrete factors that may have an anti-catabolic effect and/or encourage endogenous repair. We aim to study the effects of (i) intra-articular injection of human bone-marrow-derived MSCs and (ii) their secretome on recovery in a murine knee osteochondral injury model.The MSC secretome was generated by stimulating human bone-marrow-derived MSCs with tumour necrosis factor alpha (TNFα). Mice (n=48) were injected with i) MSC secretome, ii) MSCs or iii) cell culture medium (control). Pain was assessed by activity monitoring, and cartilage repair, subchondral bone volume and synovial inflammation were evaluated using histology and microCT.Both MSC- and MSC-secretome-injected mice showed significant pain reduction at day 7 when compared to control mice, but only the MSC-injected mice maintained a significant improvement over the controls at day 28. Cartilage repair was significantly improved in MSC-injected mice. No significant effects were observed with regards to synovial inflammation or subchondral bone volume.The MSC secretome demonstrates regenerative effects but this does not appear to be as sustained as a MSC cell therapy. Further studies are required to investigate if this can be overcome using different dosing regiments for injection of the MSC secretome. As we further understand the regenerative properties of the MSC secretome, we may be able to enhance the clinical translatability of these therapies. Direct intra-articular injection of MSCs for the treatment of OA also appears promising as a potential future strategy for OA management.Acknowledgements: MS is supported by a grant from the Wellcome Trust (PhD Programme for Clinicians)

Certain aspects of mesenchymal stem cells in small ruminants: An overview

In recent years, there has been tremendous progress in the fields of tissue engineering and cellular therapy. Bone tissue engineering is an alternative for regeneration of large bone defects caused by trauma and other bone pathologies. Treatments employed for bone regeneration are based on the use of bioactive molecules, stem cells and scaffold among others. Among these treatments, mesenchymal stem cells (MSCs) have number of advantages: i) available in large quantities from patients (ii) capable of suppressing large immune response and (iii) migrate to sites of injury. A number of farm animal models have been used for tissue-engineered products. Within each animal model, studies were categorised further according to MSC delivery/culture techniques. These techniques included direct application, fibrin glue/gel/clot, intra-articular injection, scaffold, tissue-engineered construct, meniscus tissue, pellets/aggregates and hydrogel. Till date MSCs have been isolated from various sources in small ruminants like bone marrow, adipose tissues, amniotic fluids and characterised by flow cytometry, specific staining methods and immunocytochemistry methods. Moreover, consequent to their isolation and characterization they have differentiated into osteocytes, chondrocytes, adipocytes, neural-like cells, oocyte-like cells etc. This review focuses on the biology of MSCs, including isolation, characterization, purification, in vitro differentiation potential and goat and sheep models that can be used to test tissue-engineered constructs.

Hyaluronic Acid Supplement as a Chondrogenic Adjuvant in Promoting the Therapeutic Efficacy of Stem Cell Therapy in Cartilage Healing.

The main aim of this study is to investigate the therapeutic efficacy of direct intra-articular injection of bone-marrow-derived stem/stromal cells (BMSCs) and the adjuvant role of hyaluronic acid (HA) in facilitating rabbit articular cartilage repair. First, rabbit BMSCs were treated with a medium containing different concentrations of HA. Later, HA’s influence on BMSCs’ CD44 expression, cell viability, extracellular glycosaminoglycan (GAG) synthesis, and chondrogenic gene expression was evaluated during seven-day cultivation. For the in vivo experiment, 24 rabbits were used for animal experiments and 6 rabbits were randomly allocated to each group. Briefly, chondral defects were created at the medial femoral condyle; group 1 was left untreated, group 2 was injected with HA, group 3 was transplanted with 3 × 106 BMSCs, and group 4 was transplanted with 3 × 106 BMSCs suspended in HA. Twelve weeks post-treatment, the repair outcome in each group was assessed and compared both macroscopically and microscopically. Results showed that HA treatment can promote cellular CD44 expression. However, the proliferation rate of BMSCs was downregulated when treated with 1 mg/mL (3.26 ± 0.03, p = 0.0002) and 2 mg/mL (2.61 ± 0.04, p = 0.0001) of HA compared to the control group (3.49 ± 0.05). In contrast, 2 mg/mL (2.86 ± 0.3) of HA treatment successfully promoted normalized GAG expression compared to the control group (1.88 ± 0.06) (p = 0.0009). The type II collagen gene expression of cultured BMSCs was significantly higher in BMSCs treated with 2 mg/mL of HA (p = 0.0077). In the in vivo experiment, chondral defects treated with combined BMSC and HA injection demonstrated better healing outcomes than BMSC or HA treatment alone in terms of gross grading and histological scores. In conclusion, this study helps delineate the role of HA as a chondrogenic adjuvant in augmenting the effectiveness of stem-cell-based injection therapy for in vivo cartilage repair. From a translational perspective, the combination of HA and BMSCs is a convenient, ready-to-use, and effective formulation that can improve the therapeutic efficacy of stem-cell-based therapies.

Over-Production of Therapeutic Growth Factors for Articular Cartilage Regeneration by Protein Production Platforms and Protein Packaging Cell Lines.

Simple SummaryOsteoarthritis (OA) is the most common form of arthritis across the world. Most of the existing drugs for OA treat the symptoms of pain and inflammation. There are no drugs that can dure the disease. There are a number of new treatments for OA including cell therapy and gene therapy. This articles outlines the concept behind TissueGene-C, a new biological drug for OA. This new treatment includes cartilage cells mixed with a genetically modified cell line called GP2-293, which is effectively a “drug factory”, over-producing the growth factors that are important for cartilage regeneration and changing the environment inside joints. The mixture is injected into the affected knee joint. These cells are designed to be short-lived and cannot reproduce. Therefore, after they have done their job, they die and are cleared by immune cells. This is a new and modern approach to treating OA and TissueGene-C is the prototype cell therapy for OA. In the future, it is entirely possible to combine different clones of genetically engineered cells like GP2-293 that have been designed to over-produce a growth factor or biological drug with cells from the cartilage endplate of the intervertebral disc to treat degeneration in the spine.This review article focuses on the current state-of-the-art cellular and molecular biotechnology for the over-production of clinically relevant therapeutic and anabolic growth factors. We discuss how the currently available tools and emerging technologies can be used for the regenerative treatment of osteoarthritis (OA). Transfected protein packaging cell lines such as GP-293 cells may be used as “cellular factories” for large-scale production of therapeutic proteins and pro-anabolic growth factors, particularly in the context of cartilage regeneration. However, when irradiated with gamma or x-rays, these cells lose their capacity for replication, which makes them safe for use as a live cell component of intra-articular injections. This innovation is already here, in the form of TissueGene-C, a new biological drug that consists of normal allogeneic primary chondrocytes combined with transduced GP2-293 cells that overexpress the growth factor transforming growth factor β1 (TGF-β1). TissueGene-C has revolutionized the concept of cell therapy, allowing drug companies to develop live cells as biological drug delivery systems for direct intra-articular injection of growth factors whose half-lives are in the order of minutes. Therefore, in this paper, we discuss the potential for new innovations in regenerative medicine for degenerative diseases of synovial joints using mammalian protein production platforms, specifically protein packaging cell lines, for over-producing growth factors for cartilage tissue regeneration and give recent examples. Mammalian protein production platforms that incorporate protein packaging eukaryotic cell lines are superior to prokaryotic bacterial expression systems and are likely to have a significant impact on the development of new humanized biological growth factor therapies for treating focal cartilage defects and more generally for the treatment of degenerative joint diseases such as OA, especially when injected directly into the joint.

Cell-to-Cell Culture Inhibits Dedifferentiation of Chondrocytes and Induces Differentiation of Human Umbilical Cord-Derived Mesenchymal Stem Cells.

Background Human umbilical cord-derived mesenchymal stem cells (hUC-MSCs) possess great promise as a therapeutic to repair damaged cartilage. Direct intra-articular injection of mesenchymal stem cells has been shown to reduce cartilage damage and is advantageous as surgical implantation and associated side effects can be avoided using this approach. However, the efficacy of stem cell-based therapy for cartilage repair depends highly on the direct interactions of these stem cells with chondrocytes in the joint. In this study, we have carried out an in vitro cell-to-cell contact coculture study with human articular chondrocytes (hACs) and hUC-MSCs, with the goal of this study being to evaluate interactions between hACs and hUC-MSCs. Methods Low-density monolayer cultures of hUC-MSCs and hACs were mixed at a ratio of 1 : 1 in direct cell-to-cell contact groups. Results were analyzed using quantitative reverse transcription polymerase chain reaction (qRT-PCR), western blot, enzyme-linked immunosorbent assay (ELISA), and immunofluorescence. Results A mixed coculture of hUC-MSCs and hACs was found to exhibit synergistic interactions with enhanced differentiation of hUC-MSCs and reduced dedifferentiation of chondrocytes. Mixed cultures after 21 days were found to exhibit sufficient chondrogenic induction. Conclusions The results from this study suggest the presence of mutual effects between hUC-MSCs and hACs even culture at low density and provide further support for the use of intra-articular injection strategies for cartilage defect treatment.

Injectable Click-Crosslinked Hyaluronic Acid Depot To Prolong Therapeutic Activity in Articular Joints Affected by Rheumatoid Arthritis.

The aim of this study was to design a click-crosslinked hyaluronic acid (HA) (Cx-HA) depot via a click crosslinking reaction between tetrazine-modified HA and trans-cyclooctene-modified HA for direct intra-articular injection into joints affected by rheumatoid arthritis (RA). The Cx-HA depot had significantly more hydrogel-like features and a longer in vivo residence time than the HA depot. Methotrexate (MTX)-loaded Cx-HA (MTX-Cx-HA)-easily prepared as an injectable formulation-quickly formed an MTX-Cx-HA depot that persisted at the injection site for an extended period. In vivo MTX biodistribution in MTX-Cx-HA depots showed that a high concentration of MTX persisted at the intra-articular injection site for an extended period, with little distribution of MTX to normal tissues. In contrast, direct intra-articular injection of MTX alone or MTX-HA resulted in rapid clearance from the injection site. After intra-articular injection of MTX-Cx-HA into rats with RA, we noted the most significant RA reversal, measured by an articular index score, increased cartilage thickness, extensive generation of chondrocytes and glycosaminoglycan deposits, extensive new bone formation in the RA region, and suppression of tumor necrosis factor-α or interleukin-6 expression. Therefore, MTX-Cx-HA injected intra-articularly persists at the joint site in therapeutic MTX concentrations for an extended period, thus increasing the duration of RA treatment, resulting in an improved relief of RA.

Efficacy evaluation of one-stage revision combined with intra-articular injection of antifungal agents in the treatment of chronic periprosthetic fungal infection

Objective: To investigate the clinical effect of one-stage revision combined with intra-articular injection of antifungal agents in the treatment of chronic periprosthetic fungal infection. Methods: A retrospective analysis of 11 patients(4 hips, 7 knees) admitted with chronic periprosthetic fungal infection at Department of Orthopaedics, First Affiliated Hospital of Xinjiang Medical University from January 2004 to April 2016.There were males and females with an age of 67 years (range:47-77 years). Each patient underwent single-stage revision including aggressive soft-tissue debridement. Liquid samples and tissue samples were immediately sent to the microbiology laboratory for drug sensitivity testing and histological analysis. Removed the infected components and cement thoroughly, pouring powdered vancomycin into the medullary cavity and direct intra-articular injection of fungussensitive antibiotics. The patients with infected hips received an uncemented prosthesis and 0.5 g of gentamicin loaded commercial cement was received by the patients with infected knee.After that, a new prosthesis was implanted.Long-term combination therapy of antibacterial agents and antifungal agents were given after operation. Recurrence of infection and clinical outcomes were evaluated. The follow-up period was 5 years (range: 2-12 years). Results: One patient died of acute heart failure on the eighth postoperative day.Three infection cases were recurred.Eight cases had satisfactory outcomes and required no additional surgical or medical treatment for recurrence of infection. The Harris hip score assessed preoperatively and at latest follow-up was increased from 39.25±5.12 to 79.50±4.79, the difference was statistically significant (t=-11.356, P=0.001).The Hospital for Special Surgery knee score was improved from preoperative 46.25±5.61 to final follow-up 80.50±5.06, and the difference was statistically significant (t=-9.930, P=0.002). Conclusion: Treatment of chronic fungal periprosthetic joint infection with single-stage revision can be fairly effective for achieving acceptable functional outcomes.

The Role of Gene Therapy in Cartilage Repair.

The key principle of gene delivery to articulations by direct intra-articular injection is to release complementary DNA (cDNA)-encoding medical products that will lead to maintained, endogenous production of the gene products within the articulation. In fact, this has been accomplished for both in vivo and ex vivo gene delivery, using several vectors, genes, and cells in some animal models. Some clinical trials for rheumatoid arthritis and osteoarthritis (OA) using retrovirus vectors for ex vivo gene delivery and adeno-associated virus (AAV) for in vivo delivery have been reported. AAV is of special attention because, contrary to other viral vectors, it can enter deep within joint cartilage and transduce chondrocytes in situ. This quality is of special significance in OA, in which modifications in chondrocyte metabolism are believed to be crucial to the pathophysiology of the disease. The clinical effectiveness of TissueGene-C (TG-C), a cell and gene therapy for OA consisting of nontransformed and transduced chondrocytes (3:1) retrovirally transduced to overexpress TGF-β1 has been reported in patients with knee OA. The most common complications of TG-C were peripheral edema (9%), arthralgia (8%), articular swelling (6%), and injection site pain (5%). TG-C was associated with relevant ameliorations in function and pain. Gene therapy appears to be a viable method for the management of articular cartilage defects and OA.

Pericellular collagen I coating for enhanced homing and chondrogenic differentiation of mesenchymal stem cells in direct intra-articular injection

BackgroundDirect intra-articular injection (DIAI) of mesenchymal stem cells (MSCs) is a promising technique for cartilage repair. However, the repair process was hindered by the absence of scaffold and poor cell–matrix interactions.MethodsIn this study, we developed a pericellular collagen I coating (PCC) on MSCs. The overall performances of MSC-PCC homing, chondrogenic differentiation, and cartilage regeneration have been comprehensively evaluated in a New Zealand rabbit model. Firstly, we examined the morphology and physical characteristics of PCC. Secondly, MSC ex-vivo cartilage slice adhesion and in-vivo cartilage defect homing were observed using multiscale methods. Thirdly, the precartilage condensation of cell pellets formed by aggregation of MSCs was examined to evaluate the cartilage-inducing potential of PCC. Finally, the cartilage regeneration by DIAI of PCC-coated MSCs was observed and scored macroscopically and histologically.ResultsIn general, the cell adhesion and homing assay revealed that PCC facilitated MSC adhesion on cartilage slices, enhancing MSC homing and retention to cartilage defect. This increased homing ratio was accompanied by an increasing cell–cell contact. Compared with naked MSCs, the cell pellets formed by PCC-coated MSCs exhibited more evident appearance of condensation. In pellets, cell–cell interaction has been significantly stimulated, inducing the expression of condensation marker N-cadherin, and subsequent chondrogenic marker collagen II and aggrecan. By 12 weeks after DIAI, cartilage defects have been repaired by MSCs to varying degrees. Overall, PCC significantly enhances the quality of cartilage regeneration judging from macroscopic observation, ICRS score, histological examination, and collagen type I, II, and X immunohistochemical staining.ConclusionsThe capacity and viability of MSCs can be enhanced by collagen I coating, which provides cues for enhancing cell homing and differentiation. Our method provides a novel strategy for stem cell therapy.

Gene Delivery to Joints by Intra-Articular Injection.

Most forms of arthritis are incurable, difficult to treat, and a major cause of disability in Western countries. Better local treatment of arthritis is impaired by the pharmacokinetics of the joint that make it very difficult to deliver drugs to joints at sustained, therapeutic concentrations. This is especially true of biologic drugs, such as proteins and RNA, many of which show great promise in preclinical studies. Gene transfer provides a strategy for overcoming this limitation. The basic concept is to deliver cDNAs encoding therapeutic products by direct intra-articular injection, leading to sustained, endogenous synthesis of the gene products within the joint. Proof of concept has been achieved for both in vivo and ex vivo gene delivery using a variety of vectors, genes, and cells in several different animal models. There have been a small number of clinical trials for rheumatoid arthritis (RA) and osteoarthritis (OA) using retrovirus vectors for ex vivo gene delivery and adeno-associated virus (AAV) for in vivo delivery. AAV is of particular interest because, unlike other viral vectors, it is able to penetrate deep within articular cartilage and transduce chondrocytes in situ. This property is of particular importance in OA, where changes in chondrocyte metabolism are thought to be fundamental to the pathophysiology of the disease. Authorities in Korea have recently approved the world's first arthritis gene therapy. This targets OA by the injection of allogeneic chondrocytes that have been transduced with a retrovirus carrying transforming growth factor-β1 cDNA. Phase III studies are scheduled to start in the United States soon. Meanwhile, two additional Phase I trials are listed on Clinicaltrials.gov , both using AAV. One targets RA by transferring interferon-β, and the other targets OA by transferring interleukin-1 receptor antagonist. The field is thus gaining momentum and promises to improve the treatment of these common and debilitating diseases.

Single-Stage Revision for Chronic Fungal Periprosthetic Joint Infection: An Average of 5 Years of Follow-Up

BackgroundPeriprosthetic infections caused by fungal pathogens are a rare entity, and there exist no definite guidelines according to which these infections can be successfully managed. In these situations, we wondered whether patients could be treated successfully for their fungal infections with single-stage revision. MethodsA retrospective analysis between January 2004 and October 2014 included 11 patients (4 hips and 7 knees) with chronic fungal periprosthetic joint infection who underwent single-stage revision, including aggressive soft-tissue debridement, thorough removal of infected components and cement, pouring powdered vancomycin into the medullary cavity and direct intra-articular injection of fungus-sensitive antibiotics, and a reasonable combination of antifungal agents and antibacterial medications. Recurrence of infection and clinical outcomes were evaluated. The average follow-up was 5 years (range, 2-10 years). ResultsThere were 3 failures during the study period; 1 patient died during the perioperative period because of acute heart failure on the eighth postoperative day. Of the 11 patients, 7 patients had satisfactory outcomes and required no additional surgical or medical treatment for recurrence of infection. The mean postoperative Harris hip score and Hospital for Special Surgery knee score was 77 points (67-88 points; P < .05) and 78 points (73-84 points; P < .05), respectively, at the most recent assessment. ConclusionTreatment of chronic fungal periprosthetic joint infection with single-stage revision can be fairly effective for achieving acceptable functional outcomes, which indicated that this may be a feasible alternative strategy in selected patients.

Mesenchymal stem cell and osteoarthritis: a literature review

Article type Review article The most common disease in the aged population is osteoarthritis (OA) that is resulting in progressive dysfunction following isolated cartilage injuries, subchondral bone remodeling, tissue loss, marginal osteophytes, and loss of joint space. Mesenchymal stem cells (MSCs) are multipotent stem cells; they are able to produce many or all joint tissues. Bone marrow and adipose tissue are rich sources of mesenchymal cells that are useful for the reconstruction of injured tissues such as bone, cartilage, or cardiac muscle. Recently, some studies have been performed on the use of the direct intra-articular injection of mononuclear cells (MNCs) and MSCs as potential therapeutic targets in OA. In this review, the history of MSCs in the treatment of OA are explained. Injection of Bone Marrow Aspirates Concentrate (BMAC) has significantly improved both joint pain and function in radiologic findings; some studies suggested that the injection would be even more effective in early to moderate phases of OA. Injection of MSCs in combination with growth factors may be better solution for the treatment.

Mesenchymal Stem Cells for Treating Articular Cartilage Defects and Osteoarthritis.

Articular cartilage damage and osteoarthritis are the most common joint diseases. Joints are prone to damage caused by sports injuries or aging, and such damage regularly progresses to more serious joint disorders, including osteoarthritis, which is a degenerative disease characterized by the thinning and eventual wearing out of articular cartilage, ultimately leading to joint destruction. Osteoarthritis affects millions of people worldwide. Current approaches to repair of articular cartilage damage include mosaicplasty, microfracture, and injection of autologous chondrocytes. These treatments relieve pain and improve joint function, but the long-term results are unsatisfactory. The long-term success of cartilage repair depends on development of regenerative methodologies that restore articular cartilage to a near-native state. Two promising approaches are (i) implantation of engineered constructs of mesenchymal stem cell (MSC)-seeded scaffolds, and (ii) delivery of an appropriate population of MSCs by direct intra-articular injection. MSCs may be used as trophic producers of bioactive factors initiating regenerative activities in a defective joint. Current challenges in MSC therapy are the need to overcome current limitations in cartilage cell purity and to in vitro engineer tissue structures exhibiting the required biomechanical properties. This review outlines the current status of MSCs used in cartilage tissue engineering and in cell therapy seeking to repair articular cartilage defects and related problems. MSC-based technologies show promise when used to repair cartilage defects in joints.

Meniscus repair using mesenchymal stem cells - a comprehensive review.

The menisci are a pair of semilunar fibrocartilage structures that play an essential role in maintaining normal knee function. Injury to the menisci can disrupt joint stability and lead to debilitating results. Because natural meniscal healing is limited, an efficient method of repair is necessary. Tissue engineering (TE) combines the principles of life sciences and engineering to restore the unique architecture of the native meniscus. Mesenchymal stem cells (MSCs) have been investigated for their therapeutic potential both in vitro and in vivo. This comprehensive review examines the English literature identified through a database search using Medline, Embase, Engineering Village, and SPORTDiscus. The search results were classified based on MSC type, animal model, and method of MSC delivery/culture. A variety of MSC types, including bone marrow-derived, synovium-derived, adipose-derived, and meniscus-derived MSCs, has been examined. Research results were categorized into and discussed by the different animal models used; namely murine, leporine, porcine, caprine, bovine, ovine, canine, equine, and human models of meniscus defect/repair. Within each animal model, studies were categorized further according to MSC delivery/culture techniques. These techniques included direct application, fibrin glue/gel/clot, intra-articular injection, scaffold, tissue-engineered construct, meniscus tissue, pellets/aggregates, and hydrogel. The purpose of this review is to inform the reader about the current state and advances in meniscus TE using MSCs. Future directions of MSC-based meniscus TE are also suggested to help guide prospective research.Electronic supplementary materialThe online version of this article (doi:10.1186/s13287-015-0077-2) contains supplementary material, which is available to authorized users.

Non-traditional 3D cell culture scaffold in canine mesenchymal stem cells

Three dimensional cell culture using microcarrier beads is a proven platform for large scale cell culture. Introducing a novel, functionally preserved, and clinically relevant ECM carrier could provide additional benefits and therapeutic opportunities. Therefore, we sought to use 3D cell culture to produce a cellularized, biomaterial (scaffold) generated from culturing canine MSCs on 250-micron diameter ECM disks. The newly generated cellularized scaffold could be tailored for veterinary or human clinical use, either for direct intra-articular injection, topical therapies or building block for an engineered tissue. Depositing a cellularized matrix onto an ECM-based microcarrier creates a potentially unique platform with more potent and/or clinically relevant growth factor profile. The resulting scaffold has been characterized for structure, cell attachment and proliferation, and ability to preserve and protect cellular integrity during injection.

The population of human bone marrow mesenchymal stem cells supplemented with basic fibroblast growth factor for chondrogenic differentiation

s / Osteoarthritis and Cartilage 23 (2015) A82eA416 A382 as meniscal progenitor cells (MPC) derived from diseased human meniscus. Also osteoarthritic cartilage of DDR-1-deficient mice harbors a progenitor cell population. Conclusions: Taken together, we propose that specific progenitor cell populations found in situ are responsible for tissue regeneration in a wide variety of cartilage tissues in humans andwill be of importance for cell biological therapeutic interventions in osteoarthritis. 634 THE POPULATION OF HUMAN BONE MARROWMESENCHYMAL STEM CELLS SUPPLEMENTED WITH BASIC FIBROBLAST GROWTH FACTOR FOR CHONDROGENIC DIFFERENTIATION H. Mera yz, Y. Tamamura y, M. Itokazu yx, M. Asada y, S. Wakitani k. yMukogawa Women's Univ., Nishinomiya, Japan; z Intl. Med. Device Alliance, Fndn. for BioMed. Res. and Innovation, Kobe, Japan; xOsaka City Univ., Osaka, Japan; kDept. of Artificial Joint & Biomaterials, Hiroshima Univ. Graduate Sch. of BioMed. Sci., Hiroshima, Japan Purpose:We are studying the cell basedand pharmaceuticalapproaches for cartilage repair, combined with surgical bone marrow stimulation technique to supply the cells contributing to wound heal. Bone marrow tissue has heterogeneous cell-populations, including hematopoietic stem cells, and we prefer to enhance the cells with as better chondrogenic ability as possible in order to promote cartilage repair process. It has been known that direct intra-articular injection of basic fibroblast growth factor (bFGF) promotes articular cartilage repair of osteochondral defect in animal model, however bFGF also has an unfavorable action for clinical application, osteophyte formation. In vitro studies have shown that the supplementation of bFGF for human bone marrow derived mesenchymal stem cells (hBMSCs) during expansion phase maintains the chondrogenic potential as well as enhance their proliferation. However, the population of hBMSCs supplemented with bFGF has not been investigated enough yet. This study aimed to analyze some characters, size and MSC markers, of the population of hBMSCs with bFGF. Methods: Bone marrow aspirates were obtained from healthy donors by routine iliac crest aspiration with their agreement and seeded to culture flasks, mixed with 2-fold volume of growth medium containing 15% lot adjusted FBS, supplemented with or without 10ng/ml bFGF (PeproTech, NJ, USA ). Non-adherent cells were removed at 3days and culture medium was changed every 3 days. At 11 to 14 days, P0 end, hBMSCs were passed to P1 and cultured with 5% FBS containing growth medium, keeping the condition of bFGF. The P1 cells were prepared for immunocytochemistry and for the confirmation of the chondrogenic ability. The size of the cells was measured with the automated Coulter cell counter at the end of P0 and P1. For immunocytochemistry, the cells at day4 on P1 stage were fixed with 4% PFA, and then Endoglin on cell surface were stained. The cells harvested at P1 end were used to perform pellet culture for 21 days. The histology sections of the pellets were stained with Safranin-O/Fast green, and the glycosaminoglycan (GAG) contents were measured and normalized with DNA (GAG/DNA), to evaluate their chondrogenic ability. The data of two groups, with or without bFGF on P0 and P1, were compared using unpaired t-test. Statistical tests were considered significant at the level of P < 0.05. Results: The size of the bFGF supplemented-hBMSCs was significantly smaller than hBMSCs without bFGF at P0 and P1, both. Most of the cells regardless with bFGF supplementation were expressed Endoglin, however bFGF supplemented-hBMSCs seemed to express Endoglin more. The pellets from the cells expanded with bFGF were bigger and histology of them showed Safranin-O stained more and contained more GAG/DNA than another as previously reported. Conclusions: The population of hBMSCs supplemented with bFGF has some distinct characters, smaller size and probably more expression of Endoglin than hBMSCs without bFGF. Endoglin has been reported as an accessory receptor for transforming growth factor-b (TGF-b) and might modulate the downstream signal molecules, such as smad-1/5/8, of hBMSCs. Gene expression level of Endoglin needs to be measured, and other MSC markers as well are going to be investigated. The cells involving in the cartilage repair would be characterized more, and if we could target and enhance this kind of the cells with any other agent than bFGF during cartilage repair process, better cartilage repair might be acquired. Synovial Tissue Biology & Biochemistry 635 DIFFERENTIAL SYNOVIAL EXPRESSION PATTERNS BETWEEN PATIENTS WITH PROGRESSION OF CARTILAGE DAMAGE AND PROGRESSION OF OSTEOPHYTE FORMATION IN EARLY OSTEOARTHRITIS A.B. Blom y, M.H. van den Bosch y, H. Cats z, F.H. van den Hoogen yz, F.P. Lafeber x, W.B. van den Berg y, P.L. van Lent y, P.M. van der Kraan y. yRadboud Univ. Med. Ctr. Nijmegen, Nijmegen, Netherlands; z Sint Maartenskliniek, Nijmegen, Netherlands; xUniv. Med. Ctr., Utrecht,

Humoral and cell-mediated immune response, and growth factor synthesis after direct intraarticular injection of rAAV2-IGF-I and rAAV5-IGF-I in the equine middle carpal joint.

Intraarticular (IA) administration of viral vectors expressing a therapeutic transgene is an attractive treatment modality for osteoarthritis (OA) as the joint can be treated as a contained unit. Humoral and cell-mediated immune responses in vivo can limit vector effectiveness. Transduction of articular tissues has been investigated; however, the immune response to IA vectors remains largely unknown. We hypothesized that IA rAAV2 and rAAV5 overexpressing insulin-like growth factor-I (IGF-I) would result in long-term IGF-I formation but would also induce neutralizing antibodies (NAb) and anti-capsid effector T cells. Twelve healthy horses were assigned to treatment (rAAV2 or rAAV5) or control (saline) groups. Middle carpal joints were injected with 5×10(11) vector genomes/joint. Synovial fluid was analyzed for changes in composition, NAb titers, immunoglobulin isotypes, proinflammatory cytokines, and IGF-I. Serum was analyzed for antibody titers and cytokines. A T cell restimulation assay was used to assess T cell responses. Injection of rAAV2- or rAAV5-IGF-I did not induce greater inflammation compared with saline. Synovial fluid IGF-I was significantly increased in both rAAV2- and rAAV5-IGF-I joints by day 14 and remained elevated until day 56; however, rAAV5 achieved the highest concentrations. A capsid-specific T cell response was not noted although all virus-treated horses had increased NAbs in serum and synovial fluid after treatment. Taken together, our data show that IA injection of rAAV2- or rAAV5-IGF-I does not incite a clinically detectable inflammatory or cell-mediated immune response and that IA gene therapy using minimally immunogenic vectors represents a clinically relevant tool for treating articular disorders including OA.

Pain Control after Total Knee Arthroplasty: Comparing Intra-Articular Local Anesthetic Injection with Femoral Nerve Block.

Background. Direct intra-articular injection of low doses of local anesthetic (IALA) after closure of the joint capsule remains controversial for pain control after total knee arthroplasty (TKA). Methods. A retrospective study comparing patients receiving IALA with high doses (0.5% bupivacaine 60 mL) of local anesthetics or FNB in addition to intravenous patient-controlled analgesia with opioids for pain management after TKA was conducted. The primary end point was to compare the analgesic efficacy and early ambulation between the two groups. Results. No significant differences between the two groups in pain intensity, cumulative opioid consumption, incidences of opioid-related side effects, the time interval from the end of operation to the first time the patient could walk assisted with a walker postoperatively, and postoperative hospital stay were identified. Three patients in the IALA group but none in the FNB group walked within 12 hours after the end of operation. Summary. IALA with high doses of local anesthetics provides comparable analgesic efficacy as single-shot FNB after TKA and might be associated with earlier ambulation than FNB postoperatively.

Polyethylene-glycol-modified single-walled carbon nanotubes for intra-articular delivery to chondrocytes.

Osteoarthritis (OA) is a common and debilitating degenerative disease of articular joints for which no disease-modifying medical therapy is currently available. Inefficient delivery of pharmacologic agents into cartilage-resident chondrocytes after systemic administration has been a limitation to the development of anti-OA medications. Direct intra-articular injection enables delivery of high concentrations of agents in close proximity to chondrocytes; however, the efficacy of this approach is limited by the fast clearance of small molecules and biomacromolecules after injection into the synovial cavity. Coupling of pharmacologic agents with drug delivery systems able to enhance their residence time and cartilage penetration can enhance the effectiveness of intra-articularly injected anti-OA medications. Herein we describe an efficient intra-articular delivery nanosystem based on single-walled carbon nanotubes (SWCNTs) modified with polyethylene glycol (PEG) chains (PEG-SWCNTs). We show that PEG-SWCNTs are capable to persist in the joint cavity for a prolonged time, enter the cartilage matrix, and deliver gene inhibitors into chondrocytes of both healthy and OA mice. PEG-SWCNT nanoparticles did not elicit systemic or local side effects. Our data suggest that PEG-SWCNTs represent a biocompatible and effective nanocarrier for intra-articular delivery of agents to chondrocytes.