Delivering rhFGF-18 via a bilayer collagen membrane to enhance microfracture treatment of chondral defects in a large animal model.

Augmented microfracture techniques use growth factors, cells, and/or scaffolds to enhance the healing of microfracture-treated cartilage defects. This study investigates the effect of delivering recombinant human fibroblastic growth factor 18 (rhFHF18, Sprifermin) via a collagen membrane on the healing of a chondral defect treated with microfracture in an ovine model. Eight millimeter diameter chondral defects were created in the medial femoral condyle of 40 sheep (n = 5/treatment group). Defects were treated with microfracture alone, microfracture + intra-articular rhFGF-18 or microfracture + rhFGF-18 delivered on a membrane. Outcome measures included mechanical testing, weight bearing, International Cartilage Repair Society repair score, modified O'Driscoll score, qualitative histology, and immunohistochemistry for types I and II collagen. In animals treated with 32 μg rhFGF-18 + membrane and intra-articularly, there was a statistically significant improvement in weight bearing at 2 and 4 weeks post surgery and in the modified O'Driscoll score compared to controls. In addition, repair tissue stained was more strongly stained for type II collagen than for type I collagen. rhFGF-18 delivered via a collagen membrane at the point of surgery potentiates the healing of a microfracture treated cartilage defect.This is the author accepted manuscript. The final version is available via Wiley at http://onlinelibrary.wiley.com/doi/10.1002/jor.22882/abstract

Peripheral blood derived mononuclear cells enhance osteoarthritic human chondrocyte migration.

INTRODUCTION: A major problem in cartilage repair is the lack of chondrogenic cells migrating from healthy tissue into defects. Cartilage is essentially avascular and therefore its healing is not considered to involve mononuclear cells. Peripheral blood derived mononuclear cells (PBMC) offer a readily available autologous cell source for clinical use and therefore this study was designed to evaluate the effects of PBMCs on chondrocytes and cartilage. METHODS: Human primary chondrocytes and cartilage tissue explants were taken from patients undergoing total knee replacement (n = 17). Peripheral blood samples were obtained from healthy volunteers (n = 12) and mononuclear cells were isolated by density-gradient centrifugation. Cell migration and chemokinetic potential were measured using a scratch assay, xCELLigence and CyQuant assay. PCR array and quantitative PCR was used to evaluate mRNA expression of 87 cell motility and/or chondrogenic genes. RESULTS: The chondrocyte migration rate was 2.6 times higher at 3 hour time point (p < 0.0001) and total number of migrating chondrocytes was 9.7 times higher (p < 0.0001) after three day indirect PBMC stimulus and 8.2 times higher (p < 0.0001) after three day direct co-culture with PBMCs. A cartilage explant model confirmed that PBMCs also exert a chemokinetic role on ex vivo tissue. PBMC stimulation was found to significantly upregulate the mRNA levels of 2 chondrogenic genes; collagen type II (COL2A1 600-fold, p < 0.0001) and SRY box 9 (SOX9 30-fold, p < 0.0001) and the mRNA levels of 7 genes central in cell motility and migration were differentially regulated by 24h PBMC stimulation. CONCLUSION: The results support the concept that PBMC treatment enhances chondrocyte migration without suppressing the chondrogenic phenotype possibly via mechanistic pathways involving MMP9 and IGF1. In the future, peripheral blood mononuclear cells could be used as an autologous point-ofcare treatment to attract native chondrocytes from the diseased tissue to aid in cartilage repair.The authors would like to kindly acknowledge the PhD studentship from John Insall Foundation US and thank Dr. Nigel Loveridge for his statistical expertise. Dr. John Wardale acknowledges funding from the Technology Strategy Board and OrthoMimetics and Dr. Roger Brooks acknowledges funding from the National Institute for Health Research.This is the final version of the article. It first appeared from BioMed Central via http://dx.doi.org/10.1186/s13075-015-0709-

Peripheral Blood Mononuclear Cells Enhance Cartilage Repair in in vivo Osteochondral Defect Model.

This study characterized peripheral blood mononuclear cells (PBMC) in terms of their potential in cartilage repair and investigated their ability to improve the healing in a pre-clinical large animal model. Human PBMCs were isolated with gradient centrifugation and adherent PBMC's were evaluated for their ability to differentiate into adipogenic, chondrogenic and osteogenic lineages and also for their expression of musculoskeletal genes. The phenotype of the PBMCs was evaluated using Stro-1, CD34, CD44, CD45, CD90, CD106, CD105, CD146 and CD166 cell surface markers. Osteochondral defects were created in the medial femoral condyle (MFC) of 24 Welsh mountain sheep and evaluated at a six month time point. Four cell treatment groups were evaluated in combination with collagen-GAG-scaffold: (1) MSC alone; (2) MSCs and PBMCs at a ratio of 20:1; (3) MSCs and PBMC at a ratio of 2:1 and (4) PBMCs alone. Samples from the surgical site were evaluated for mechanical properties, ICRS score and histological repair. Fresh PBMC samples were 90% positive for hematopoietic cell surface markers and negative for the MSC antibody panel (<1%, p = 0.006). However, the adherent PBMC population expressed mesenchymal stem cell markers in hypoxic culture and lacked CD34/45 positive cells (<0.2%). This finding demonstrated that the adherent cells had acquired an MSC-like phenotype and transformed in hypoxia from their original hematopoietic lineage. Four key genes in muskuloskeletal biology were significantly upregulated in adherent PBMCs by hypoxia: BMP2 4.2-fold (p = 0.0007), BMP6 10.7-fold (p = 0.0004), GDF5 2.0-fold (p = 0.002) and COL1 5.0-fold (p = 0.046). The monolayer multilineage analysis confirmed the trilineage mesenchymal potential of the adherent PBMCs. PBMC cell therapy was equally good as bone marrow MSC therapy for defects in the ovine large animal model. Our results show that PBMCs support cartilage healing and oxygen tension of the environment was found to have a key effect on the derivation of a novel adherent cell population with an MSC-like phenotype. This study presents a novel and easily attainable point-of-care cell therapy with PBMCs to treat osteochondral defects in the knee avoiding any cell manipulations outside the surgical room.PhD studentship for Niina Hopper was from charitable trust John Insall Foundation US. Dr. John Wardale acknowledges funding from the Technology Strategy Board and industrial partner OrthoMimetics (currently known as Tigenix) and Dr. Roger Brooks acknowledges funding from the National Institute for Health Research for their salaries.This is the final version of the article. It first appeared from PLOS via http://dx.doi.org/10.1371/journal.pone.013393

Bioactive IGF-1 release from collagen-GAG scaffold to enhance cartilage repair in vitro.

Tissue engineering is a promising technique for cartilage repair. Toward this goal, a porous collagen-glycosaminoglycan (CG) scaffold was loaded with different concentrations of insulin-like growth factor-1 (IGF-1) and evaluated as a growth factor delivery device. The biological response was assessed by monitoring the amount of type II collagen and proteoglycan synthesised by the chondrocytes seeded within the scaffolds. IGF-1 release was dependent on the IGF-1 loading concentration used to adsorb IGF-1 onto the CG scaffolds and the amount of IGF-1 released into the media was highest at day 4. This initial IGF-1 release could be modelled using linear regression analysis. Osteoarthritic (OA) chondrocytes seeded within scaffolds containing adsorbed IGF-1 deposited decorin and type II collagen in a dose dependent manner and the highest type II collagen deposition was achieved via loading the scaffold with 50 μg/ml IGF-1. Cells seeded within the IGF-1 loaded scaffolds also deposited more extracellular matrix than the no growth factor control group thus the IGF-1 released from the scaffold remained bioactive and exerted an anabolic effect on OA chondrocytes. The effectiveness of adsorbing IGF-1 onto the scaffold may be due to protection of the molecule from proteolytic digestion allowing a more sustained release of IGF-1 over time compared to adding multiple doses of exogenous growth factor. Incorporating IGF-1 into the CG scaffold provided an initial therapeutic burst release of IGF-1 which is beneficial in initiating ECM deposition and repair in this in vitro model and shows potential for developing this delivery device in vivo.EPSRC and Tigenix UK Limited, Byron House, Cambridge Business Park, Milton Road, Cambridge, are gratefully acknowledged for a studentship for L.M.M.This is the final published version. The final published version is available from Springer at http://link.springer.com/article/10.1007%2Fs10856-014-5325-y

A novel biphasic scaffold supports meniscal tissue repair in ex vivo and in vivo models

The objectives of this study were i) to investigate whether a novel biphasic collagen scaffold supports meniscal repair in an ovine meniscal defect model in vivo, ii) to investigate meniscal cell ingrowth and ECM production in an ex-vivo model and iii) to investigate the effect platelet rich plasma (PRP) on the response of meniscal elements to the scaffold.A novel biphasic scaffold (collagen sponge with 300 mm open pores, reinforced with bands of 150 mm internal strengthening fibres) was implanted into a circular defect in the meniscus of adult sheep with and without PRP. Healing in vivo was monitored by longitudinal gait analysis. At 2 and 26 weeks post surgery durometry, histology and immunohistochemistry was performed on recovered menisci. In vitro, ex vivo ovine meniscus/scaffold models were constructed and the effects of PRP on cell migration and extracellular matrix production monitored by cell tracking, protein extraction and Western blotting.In this study we have demonstrated that a novel biphasic collagen scaffold can support meniscal defect repair in an in vivo model and support the ingrowth of meniscal fibrochondrocyte (MFC) in an ex vivo model. In addition we have demonstrated that the addition of PRP significantly enhances MFC migration and ECM production in vitro and promotes rapid defect healing in vivo.This novel biphasic scaffold supports meniscal healing, which is increased in the presence of PRP, likely due to increased MFC migration

Understanding the influences on successful quality improvement in emergency general surgery: learning from the RCS Chole-QuIC project

Abstract: Background: Acute gallstone disease is the highest volume Emergency General Surgical presentation in the UK. Recent data indicate wide variations in the quality of care provided across the country, with national guidance for care delivery not implemented in most UK hospitals. Against this backdrop, the Royal College of Surgeons of England set up a 13-hospital quality improvement collaborative (Chole-QuIC) to support clinical teams to reduce time to surgery for patients with acute gallstone disease requiring emergency cholecystectomy. Methods: Prospective, mixed-methods process evaluation to answer the following: (1) how was the collaborative delivered by the faculty and received, understood and enacted by the participants; (2) what influenced teams’ ability to improve care for patients requiring emergency cholecystectomy? We collected and analysed a range of data including field notes, ethnographic observations of meetings, and project documentation. Analysis was based on the framework approach, informed by Normalisation Process Theory, and involved the creation of comparative case studies based on hospital performance during the project. Results: Chole-QuIC was delivered as planned and was well received and understood by participants. Four hospitals were identified as highly successful, based upon a substantial increase in the number of patients having surgery in line with national guidance. Conversely, four hospitals were identified as challenged, achieving no significant improvement. The comparative analysis indicate that six inter-related influences appeared most associated with improvement: (1) achieving clarity of purpose amongst site leads and key stakeholders; (2) capacity to lead and effective project support; (3) ideas to action; (4) learning from own and others’ experience; (5) creating additional capacity to do emergency cholecystectomies; and (6) coordinating/managing the patient pathway. Conclusion: Collaborative-based quality improvement is a viable strategy for emergency surgery but success requires the deployment of effective clinical strategies in conjunction with improvement strategies. In particular, achieving clarity of purpose about proposed changes amongst key stakeholders was a vital precursor to improvement, enabling the creation of additional surgical capacity and new pathways to be implemented effectively. Protected time, testing ideas, and the ability to learn quickly from data and experience were associated with greater impact within this cohort

Multicenter Diagnostic Evaluation of OnSite COVID-19 Rapid Test (CTK Biotech) among Symptomatic Individuals in Brazil and the United Kingdom

Evaluating rapid diagnostic tests in diverse populations is essential to improving diagnostic responses as it gives an indication of the accuracy in real-world scenarios. In the case of rapid diagnostic testing within this pandemic, lateral flow tests that meet the minimum requirements for sensitivity and specificity can play a key role in increasing testing capacity, allowing timely clinical management of those infected, and protecting health care systems