Aims. The aim of this study was to determine the rate of indocyanine green (ICG) staining of bone and soft-tissue tumours, as well as the stability and accuracy of ICG fluorescence imaging in detecting tumour residuals during surgery for bone and soft-tissue tumours. Methods. ICG fluorescence imaging was performed during surgery in 34 patients with bone and soft-tissue tumours. ICG was administered intravenously at a dose of 2 mg/kg over a period of 60 minutes on the day prior to surgery. The tumour stain rate and signal-to-background ratio of each tumour were post hoc analyzed. After tumour resection, the tumour bed was scanned to locate sites with fluorescence residuals, which were subsequently inspected and biopsied. Results. The overall tumour stain rate was 88% (30/34 patients), and specific stain rates included 90% for osteosarcomas and 92% for giant cell tumours. For malignant tumours, the overall stain rate was 94%, while it was 82% for benign tumours. The ICG tumour stain was not influenced by different pathologies, such as malignant versus benign pathology, the reception (or lack thereof) of neoadjuvant chemotherapies, the length of time between drug administration and surgery, the number of doses of denosumab for patients with giant cell tumours, or the tumour response to neoadjuvant chemotherapy. The overall accuracy rate of successfully predicting tumour residuals using fluorescence was 49% (23/47 pieces of tissue). The accuracy rate after en bloc resection was significantly lower than that after piecemeal resection (16% vs 71%; p < 0.001). Conclusion. A high percentage of bone and soft-tissue tumours can be stained by ICG and the tumour staining with ICG was stable. This approach can be used in both benign and malignant tumours, regardless of whether neoadjuvant chemotherapy is adopted. The technique is also useful to detect tumour residuals in the wound, especially in patients undergoing piecemeal resection. Cite this article: Bone Joint J 2023;105-B(5):551–558

Aims. Arthroplasty surgery of the knee and hip is performed in two to three million patients annually. Periprosthetic joint infections occur in 4% of these patients. Debridement, antibiotics, and implant retention (DAIR) surgery aimed at cleaning the infected prosthesis often fails, subsequently requiring invasive revision of the complete prosthetic reconstruction. Infection-specific imaging may help to guide DAIR. In this study, we evaluated a bacteria-specific hybrid tracer (. 99m. Tc-UBI. 29-41. -Cy5) and its ability to visualize the bacterial load on femoral implants using clinical-grade image guidance methods. Methods. 99m. Tc-UBI. 29-41. -Cy5 specificity for Stapylococcus aureus was assessed in vitro using fluorescence confocal imaging. Topical administration was used to highlight the location of S. aureus cultured on femoral prostheses using fluorescence imaging and freehand single photon emission CT (fhSPECT) scans. Gamma counting and fhSPECT were used to quantify the bacterial load and monitor cleaning with chlorhexidine. Microbiological culturing helped to relate the imaging findings with the number of (remaining) bacteria. Results. Bacteria could be effectively stained in vitro and on prostheses, irrespective of the presence of biofilm. Infected prostheses revealed bacterial presence on the transition zone between the head and neck, and in the screw hole. Qualitative 2D fluorescence images could be complemented with quantitative 3D fhSPECT scans. Despite thorough chlorhexidine treatments, 28% to 44% of the signal remained present in the locations of the infection that were identified using imaging, which included 500 to 2,000 viable bacteria. Conclusion. The hybrid tracer . 99m. Tc-UBI. 29-41. -Cy5 allowed effective bacterial staining. Qualitative real-time fluorescence guidance could be effectively combined with nuclear imaging that enables quantitative monitoring of the effectiveness of cleaning strategies. Cite this article: Bone Joint Res 2023;12(1):72–79

Aim. Prosthetic joint infections pose a major clinical challenge. Developing novel material surface technologies for orthopedic implants that prevent bacterial adhesion and biofilm formation is essential. Antimicrobial coatings applicable to articulating implant surfaces are limited, due to the articulation mechanics inducing wear, coating degradation, and toxic particle release. Noble metals are known for their antimicrobial activity and high mechanical strength and could be a viable coating alternative for orthopaedic implants [1]. In this study, the potential of thin platinum-based metal alloy coatings was developed, characterized, and tested on cytotoxicity and antibacterial properties. Method. Three platinum-based metal alloy coatings were sputter-coated on medical-grade polished titanium discs. The coatings were characterized using optical topography and scanning electron microscopy with energy dispersive spectroscopy (SEM/EDS). Ion release was measured using inductively coupled plasma optical emission spectrometry (ICP-OES). Cytotoxicity was tested according to ISO10993-5 using mouse fibroblasts (cell lines L929 and 3T3). Antibacterial surface activity, bacterial adhesion, bacterial proliferation, and biofilm formation were tested with gram-positive Staphylococcus aureus ATCC 25923 and gram-negative Escherichia coli ATCC 25922. Colony forming unit (CFU) counts, live-dead fluorescence staining, and SEM-EDS images were used to assess antibacterial activity. Results. Three different platinum-based metal alloys consisting of platinum-iridium, platinum-copper, and platinum-zirconium. The coatings were found 80 nm thick, smooth (roughness average < 60 nm), and non-toxic. The platinum-copper coating showed a CFU reduction larger than one logarithm in adherent bacteria compared to uncoated titanium. The other coatings showed a smaller reduction. This data was confirmed by SEM and live-dead fluorescence images, and accordingly, ICP-OES measurements showed low levels of metal ion release from the coatings. Conclusions. The platinum-copper coating showed low anti-adhesion properties, even with extremely low metal ions released. These platinum-based metal alloy coatings cannot be classified as antimicrobial yet. Further optimization of the coating composition to induce a higher ion release based on the galvanic principle is required and copper looks most promising as the antimicrobial compound of choice. Acknowledgments. This publication is supported by the DARTBAC project (with project number NWA.1292.19.354) of the research program NWA-ORC which is (partly) financed by the Dutch Research Council (NWO); and the AMBITION project (with project number NSP20–1-302), co-funded by the PPP Allowance made available by Health-Holland, Top Sector Life Sciences & Health to ReumaNederland

INTRODUCTION. Trabecular Titanium. ™. (TT) is a novel material with a structure similar to trabecular bone, already used for prosthetic clinical applications. Being the bone-implant interface the weakest point during the initial healing period, the association of TT with a hydrogel enriched with progenitor cells and osteoinductive factors may represent a promising strategy to improve prosthesis osteointegration. In a previous in vitro study we evaluated the ability of an ammidated carboxymethylcellulose hydrogel (CMCA) and of TT enriched with CMCA to support bone marrow mesenchymal stem cells (BMSCs) viability and osteogenic differentiation [1]. The aim of this study was to evaluate in vivo if the association of TT with CMCA enriched with strontium chloride (SrCl. 2. ) and BMSCs could ameliorate TT osteointegration. METHODS. This study combines TT with CMCA, SrCl. 2. and BMSCs. To mimic prosthesis-bone implants, TT discs were seeded with human BMSCs predifferentiated in osteogenic medium, then press-fit into engineered bone. A total of 36 athymic mice were implanted subcutaneously, each animal received 2 constructs as un-seeded TT and TT+CMCA or cell seeded TT+BMSCs and TT+CMCA+BMSCs. After 4, 8 and 12 weeks, osteodeposition, bone mineral density (BMD) and osteointegration were evaluated by fluorescence imaging, micro-CT, SEM, histology and pull-out tests. RESULTS. Micro-CT analysis demonstrated the homogeneity of the engineered bone in all experimental groups, supporting the reproducibility of our novel engineered model. Macroscopic evaluation of explanted constructs after 4 weeks revealed their integration with mice subcutaneous structures. In pull-out biomechanical tests, increases in extraction energy and peak force from 4 to 12 weeks were observed in all the experimental groups, except TT+CMCA. TT+CMCA+BMSCs showed the highest value of peak force and the greatest increase in comparison to samples explanted at 4 weeks. In vivo fluorescence imaging showed osteodeposition activity inside the constructs, observation confirmed by the ex-vivo analyses revealing a higher activity in TT+BMSCs and in TT+CMCA+BMSCs in comparison to acellularized TT samples. SEM evaluation of ECM deposition at the interface between bone scaffolds and TT disks revealed a significant difference between TT+CMCA+BMSCs and the other experimental groups with the former showing an almost complete filling of the space between the integration surfaces already after 4 weeks. In histomorphometric analyses of tissue ingrowth at 8 weeks, TT+BMSCs and TT+CMCA+BMSCs showed a greater tissue ingrowth compared to TT and TT+CMCA samples. DISCUSSION. Several efforts have been made to improve osteointegration with particular attention to critical cases such as implant revision surgeries. The association of porous structures with osteoinductive factors enriched hydrogels and stem cells represents a novel and promising strategy for more effective osteointegration to reduce prosthesis mobilization risks. Our results demonstrate that the association of Trabecular Titanium. ™. with a SrCl. 2. enriched hydrogel and BMSCs increases the production of ECM and may thus represent a valid approach to accelerate prosthesis osteointegration. Further validation of these data will include construct implantation in large animal orthotopic models to better mimic surgical procedures

Objectives. Indocyanine green (ICG) fluorescence angiography is an emerging technique that can provide detailed anatomical information during surgery. The purpose of this study is to determine whether ICG fluorescence angiography can be used to evaluate the blood flow of the rotator cuff tendon in the clinical setting. Methods. Twenty-six patients were evaluated from October 2016 to December 2017. The participants were categorized into three groups based on their diagnoses: the rotator cuff tear group; normal rotator cuff group; and adhesive capsulitis group. After establishing a posterior standard viewing portal, intravenous administration of ICG at 0.2 mg/kg body weight was performed, and fluorescence images were recorded. The time from injection of the drug to the beginning of enhancement of the observed area was measured. The hypovascular area in the rotator cuff was evaluated, and the ratio of the hypovascular area to the anterolateral area of the rotator cuff tendon was calculated (hypovascular area ratio). Results. ICG fluorescence angiography allowed for visualization of blood flow in the rotator cuff in all groups. The adhesive capsulitis group showed significantly earlier enhancement than the other groups. Furthermore, the adhesive capsulitis group had a significantly smaller hypovascular area ratio than the other groups. Conclusion. ICG fluorescence angiography allowed for evaluation of real-time blood flow of the rotator cuff in arthroscopic shoulder surgery. The techniques of ICG fluorescence angiography are simple and easy to observe, observer reliability is high, and it has utility for evaluating blood flow during surgery. Cite this article: N. Doi, T. Izaki, S. Miyake, T. Shibata, T. Ishimatsu, Y. Shibata, T. Yamamoto. Intraoperative evaluation of blood flow for soft tissues in orthopaedic surgery using indocyanine green fluorescence angiography: A pilot study. Bone Joint Res 2019;8:118–125. DOI: 10.1302/2046-3758.83.BJR-2018-0151.R1

One of the most common bacteria in orthopaedic prosthetic infections is Staphylococcus Aureus. Infection causes implant failure due to biofilm production. Biofilms are produced by bacteria once they have adhered to a surface. Nanotopography has major effects on cell behaviour. Our research focuses on bacterial adhesion on nanofabricated materials. We hypothesise that surface nanotopography impacts the differential ability of staphylococci species to adhere via altered metabolomics and may reduce orthopaedic implant infection rate. Bacteria were grown and growth conditions optimised. Polystyrene and titanium (Ti) nanosurfaces were studied. The polystyrene surfaces had different nanopit arrays, while the Ti surfaces expressed different nanowire structures. Adhesion analysis was performed using fluorescence imaging, quantitative PCR and bacterial percentage coverage calculations. Further substitution with ‘heavy’ labelled glucose into growth medium allowed for bacterial metabolomic analysis and identification of any up-regulated metabolites and pathways. Our data demonstrates reduced bacterial adhesion on specific nanopit polystyrene arrays, while nanowired titanium showed increased bacterial adhesion following qPCR (P<0.05) and percentage coverage calculations (P<0.001). Further metabolomic analysis identified significantly increased intensity counts of specific metabolites (Pyruvate, Aspartate, Alanine and Carbamoyl aspartate). Our study shows that by altering nanotopography, bacterial adhesion and therefore biofilm formation can be affected. Specific nanopatterned surfaces may reduce implant infection associated morbidity and mortality. The identification of metabolic pathways involved in adhesion may allow for a targeted approach to biofilm eradication in S. aureus. This is of significant benefit to both the patient and the surgeon, and may well extend far beyond the realms of orthopaedics

The most common bacteria in orthopaedic prosthetic infections are Staphylococcus, namely Staphylococcus Epidermidis (SE) and Staphylococcus Aureus (SA). Infection causes implant failure due to biofilm production. Biofilms are produced by bacteria once they have adhered to a surface. Nanotopography has major effects on cell behaviour. Our research focuses on bacterial adhesion and biofilm formation on nanofabricated materials. Bacteria studied were clinically relevant from an orthopaedic perspective, SA and SE. We hypothesise that that nanosurfaces can modulate bacterial adherence and biofilm formation and may reduce orthopaedic implant infection rate. Isolated bacteria were grown and growth conditions optimised. Bacterial concentrations were calculated by using qPCR. Statistical analysis allowed identification of optimal biofilm growth conditions. These were refined on standard, non-nanopatterned surfaces, and then control and nanopatterned polystyrene (nanopits) and titanium plates (nanowires). Adhesion analysis was performed using fluorescence imaging and quantitative PCR. 4 bacterial strains were isolated and cultured. Growth kinetics based on 24hr cultures allowed isolation of optimal media for biofilm conditions (Dulbecco's Modified Eagle Medium with additional supplements). Highest bacterial concentrations were found following 2hrs incubation with Lysozyme during qPCR. Bacterial concentration significantly increased between 30, 60 and 90 minutes incubation. Differences in percentage coverage on different polysyrene nanosurfaces (nanopits) were noted varying. This was confirmed by qPCR extractions that showed different bacterial concentrations on different nanopatterns. Titanium nanowire surfaces significantly increased bacterial adhesion (P<0.05). Our study cultured and quantified bacterial biofilm and suggests that by altering nanotopography, bacterial adhesion and therefore biofilm formation can be affected. Specific nanopatterned surfaces may reduce implant infection associated morbidity and mortality. Clearly this is of significant benefit to the patient, the surgeon and the NHS, and may well extend far beyond the realms of orthopaedics

Aims

In this investigation, we administered oxidative stress to nucleus pulposus cells (NPCs), recognized DNA-damage-inducible transcript 4 (DDIT4) as a component in intervertebral disc degeneration (IVDD), and devised a hydrogel capable of conveying small interfering RNA (siRNA) to IVDD.

Aims

To investigate the effects of senescent osteocytes on bone homeostasis in the progress of age-related osteoporosis and explore the underlying mechanism.

INTRODUCTION: The natural synovial joints with very low friction and low wear are likely to operate in the adaptive multimode lubrication mechanism, in which various lubrication modes become effective in various daily activities. On the contrary, in the artificial joints composed of ultra-high molecular weight polyethylene (UHMWPE) and metallic or ceramic components, it is difficult to expect the sufficient fluid film formation to prevent the direct contact between rubbing surfaces, and thus considerable wear of UHMWPE occurs. To improve the longevity of joint prostheses by reduction of wear and friction, it is effective to improve the lubrication mode to fluid film lubrication by the application of compliant materials (. 1. ). In this paper, the effectiveness of the compliant artificial cartilage of poly (vinyl alcohol) (PVA) hydrogel of high water content and the existence of the optimum adsorbed films were examined. MATERIALS AND METHODS: The reciprocating friction apparatus was used to investigate the influence of the lubricants containing proteins on the wear properties. The sliding surfaces are composed of PVA hydrogel and itself or a glass plate. PVA hydrogel was prepared by repeated freezing thawing method. The elastic modulus of PVA hydrogel is 1.2 MPa and equivalent water content is 79%. PVA hydrogel stationary upper specimen have elliptical geometry with diameters of 25mm and 40mm. Lubricants as sodium hyaluronate (HA) or saline solutions with or without serum protein were used. For distinction between albumin and γ-globulin in fluorescent observation for adsorbed film formed on glass plate after tests, albumin and γ-globulin were labeled with Rhodamine-B-isothiocyanate and Fluorescein isothiocyanate isomer I, respectively. Next, to conduct in situ observation of adsorbed film formation, the reciprocating apparatus was constructed on the stage in inverted fluorescent microscope. A sliding pair of a spherical reciprocating upper specimen of PVA hydrogel and flat stationary lower specimen of cover glass was used. RESULTS AND DISCUSSION: It is noticed in solutions of single protein that an increase in protein concentration increased wear grade. In contrast, in binary protein solution the minimum wear is found at optimum composition such as total content of 2.1 wt% and A/G ratio of 1/2 or 2/1 (A:albumin, G:γ-globulin). The fluorescence images of adsorbed film on rubbed glass plate showed that the adsorbed films formed a layered structure composed of albumin and γ-globulin. On the contrary, in high wear case for higher protein concentration, the adsorbed film was formed as a heterogeneous separated structure. Furthermore, in situ observation of lubricated conjunction clearly indicated that the formation of layered films or heterogeneous films corresponded to lubricant constituents. In this study, it was shown that the layered adsorbed film formation originated from the optimum composition of proteins in lubricants is effective to maintain low wear and low friction for PVA hydrogel artificial cartilage

INTRODUCTION: The natural synovial joints with very low friction and low wear are likely to operate in the adaptive multimode lubrication mechanism, in which various lubrication modes become effective in various daily activities. On the contrary, in the artificial joints composed of ultra-high molecular weight polyethylene (UHMWPE) and metallic or ceramic components, it is difficult to expect the sufficient fluid film formation to prevent the direct contact between rubbing surfaces, and thus considerable wear of UHMWPE occurs. To improve the longevity of joint prostheses by reduction of wear and friction, it is effective to improve the lubrication mode to fluid film lubrication by the application of compliant materials. 1. In this paper, the effectiveness of the compliant artificial cartilage of poly (vinyl alcohol) (PVA) hydrogel of high water content and the existence of the optimum adsorbed films were examined. MATERIALS AND METHODS: The reciprocating friction apparatus was used to investigate the influence of the lubricants containing proteins on the wear properties. The sliding surfaces are composed of PVA hydrogel and itself or a glass plate. PVA hydrogel was prepared by repeated freezing thawing method. The elastic modulus of PVA hydrogel is 1.2 MPa and equivalent water content is 79%. PVA hydrogel stationary upper specimen have elliptical geometry with diameters of 25mm and 40mm. Lubricants as sodium hyaluronate (HA) or saline solutions with or without serum protein were used. For distinction between albumin and γ-globulin in fluorescent observation for adsorbed film formed on glass plate after tests, albumin and γ-globulin were labeled with Rhodamine-B-isothiocyanate and Fluorescein isothiocyanate isomer I, respectively. Next, to conduct in situ observation of adsorbed film formation, the reciprocating apparatus was constructed on the stage in inverted fluorescent microscope. A sliding pair of a spherical reciprocating upper specimen of PVA hydrogel and flat stationary lower specimen of cover glass was used. RESULTS AND DISCUSSION: It is noticed in solutions of single protein that an increase in protein concentration increased wear grade. In contrast, in binary protein solution the minimum wear is found at optimum composition such as total content of 2.1wt% and A/G ratio of 1/2 or 2/1 (A: albumin, G:γ-globulin). The fluorescence images of adsorbed film on rubbed glass plate showed that the adsorbed films formed a layered structure composed of albumin and γ-globulin. On the contrary, in high wear case for higher protein concentration, the adsorbed film was formed as a heterogeneous separated structure. Furthermore, in situ observation of lubricated conjunction clearly indicated that the formation of layered films or heterogeneous films corresponded to lubricant constituents. In this study, it was shown that the layered adsorbed film formation originated from the optimum composition of proteins in lubricants is effective to maintain low wear and low friction for PVA hydrogel artificial cartilage

Recapitulating tissue elasticity can direct mesenchymal stromal cell (MSC) differentiation; however, it is unclear how substrate elasticity affects MSC metabolism. It is hypothesized MSCs subjected to stiffnesses, atypical of standard tissue culture plastic, display altered metabolic phenotypes during differentiation. In this study, such alterations in MSC metabolic profiles, based on the fluorescence lifetime of NAD(P)H, a critical co-factor in energy production, were monitored using Fluorescence lifetime imaging microscopy (FLIM) as an evaluation tool. Polyacrylamide substrates with varying stiffnesses were fabricated to model the native elasticity of cartilage and bone. MSCs cultured on these substrates exhibited potent alterations in their metabolic status over a 14-day period that were detectable as early as day 3 using FLIM. Overall, soft substrates induced a more glycolytic response after 10 days of culture that persisted at day 14 (as measured by protein-bound NAD(P)H contributions to the lifetime decay). Similarly, by day 10; MSCs on intermediate-stiffness substrates favoured glycolysis. MSCs on stiffer substrates initially displayed a glycolytic phenotype followed by a transition to oxidative phosphorylation by day 10. Staining for mineralised nodules and glycosaminoglycans verified MSCs on stiffer substrates differentiating towards an osteogenic lineage, while MSCs on intermediate substrates showed similarities with differentiated chondrocytes. Overall, it can be concluded that matrix stiffness can induce metabolic perturbations in MSCs for up to 14 days. From this research, ideal culture conditions in which the metabolics of MSCs could be manipulated to promote maximum potency could potentially be defined in the future

Aims

This study aimed to investigate the role and mechanism of meniscal cell lysate (MCL) in fibroblast-like synoviocytes (FLSs) and osteoarthritis (OA).

Aims

To explore the novel molecular mechanisms of histone deacetylase 4 (HDAC4) in chondrocytes via RNA sequencing (RNA-seq) analysis.

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The role of N,N-dimethylformamide (DMF) in diabetes-induced osteoporosis (DM-OS) progression remains unclear. Here, we aimed to explore the effect of DMF on DM-OS development.

Summary Statement. Calcium phosphate (CaP) particles have attracted great interest as transfection reagents, yet little is known about their mechanism of internalisation. We report live cell time-course tracking of CaP particles during internalisation and the influence of Ca:P ratio on transfection efficiency. Introduction. Relatively recent work has seen calcium phosphate (CaP) salts used for the delivery of biological materials into cells in the form of peptides, polymers and DNA sequences. Calcium phosphate salts have a critical safety advantage over other vectors such as viruses in that they pose no risk of pathogenicity due to mutation and show no apparent cytotoxicity. Previous work within the group showed that Ca:P ratio influenced the transfection efficiency, but the fate of the particles on internalisation is yet unknown. The difficulty in tracking the particles can be related to the visual similarity to granulation within the cells. Using a surface modification method that enables the fluorescent labeling of silicon-substituted hydroxyapatite (SiHA) particles, we have tracked the internalisation of the particles to understand their mechanism of entry and how particle composition may influence transfection efficiency. Patients & Methods. SiHA particles were synthesised by the dropwise addition of an aqueous solution of diammonium hydrogen phosphate and silicon tetraacetate to an aqueous solution of calcium nitrate while under mixing and maintained at pH10. The particles were functionalised with thiol groups using (3-mercaptopropyl)trimethoxysilane and dye-labelled with fluorescein-5-maleimide. MC3T3 osteoblast precursor cells were incubated in cell culture media containing labelled particles at a concentration of 0.6μg/mL for 12 hours. Confocal images were obtained with a Zeiss LSM 710 ConfoCor 3 system based around a Zeiss AxioObserverZ1 microscope. Results. DNA binding efficiency between 79 to 94%, the lowest being the CaP sample of new CaP route at Ca/P ratio of 0.33 by SEDS processing, which was 79% and the highest was the HAp SEDS processed sample at 40°C, solvent flowrate of 1 ml/min and antisolvent flowrate of 60 g/min (particle size of 131 nm). From the fluorescence microscopy images, localised regions of particles measuring around 500–1000nm were detected. With a typical SiHA particle size of 50–70nm in length, these regions contain 10's of particles. Discussion/Conclusion. Thiol functionalisation enabled the internalised SiHA to be visually discriminated from the other cellular material with similar morphology and optical contrast as shown in the bright field image. HA particles (Ca:P of 1.67) showed a strong affinity for the cell membrane despite extensive washing with PBS and their higher calcium content may enhance the binding of the DNA to the particle surface, therefore improving transfection efficiency

Aims

The aim of this study was to develop a single-layer hybrid organic-inorganic sol-gel coating that is capable of a controlled antibiotic release for cementless hydroxyapatite (HA)-coated titanium orthopaedic prostheses.

Aims

Extracellular matrix (ECM) and its architecture have a vital role in articular cartilage (AC) structure and function. We hypothesized that a multi-layered chitosan-gelatin (CG) scaffold that resembles ECM, as well as native collagen architecture of AC, will achieve superior chondrogenesis and AC regeneration. We also compared its in vitro and in vivo outcomes with randomly aligned CG scaffold.

Introduction: Tissue engineered scaffolds require vascularization to 1) enhance nutrient exchange and 2) provide cells needed to build new tissue. Cell-seeded scaffolds; bioreactors-- require rapid penetration of vessels or enhanced fluid percolation to keep their contents alive until normal nutrient exchange can be established. Bone fluid flow depends on a pumping system which drives percolation through its own matrix. Recent interest in the pumping mechanism has resulted in bone fluid flow models, which link the pumps to bending of bone by muscle contraction and compression-tension cycles from weight-bearing during locomotion. The present authors have proposed that capillary filtration, the source of the percolating fluid, is sufficiently enhanced by soliton pressure waves in blood driven by the muscle pump during exercise to provide a significant hydraulic pressure component to bone fluid percolating through bone and any bone-implanted scaffold. A proposal and some preliminary results from a pilot project suggesting enhancement of capillary filtration by the muscle pump is presented. Materials and Methods: Optical bone chambers were implanted in adult New Zealand White female rabbits. Chamber construction and implantation were as usual1. At the third week post-op, chamber ends were exposed and weekly intravital microscopy commenced. Transcutaneous electrical stimulation was administered with a ToneATronic® TENS at 85V, 80mA and 2Hz. The stimulator was applied externally over the gastrocnemius muscle. A fluorescence digital image was obtained before 30 minutes of application of transcutaneous electrical nerve stimulation (TENS) after injection of FITC-D70. Blood samples were obtained from an aural vein in the ear opposite that being injected with the fluorescent dye after each injection. Blood concentration of dye was determined with a SPEX Fluoromax-3 spectrofluorometer for both serum (absolute concentration) and whole blood (to detect differences which would make fluorescence in vessels an inaccurate indicator of red blood cell color contamination). For analysis, four vessels were chosen and the average dye concentration profiles before and after 30 minutes of stimulation were obtained. Results: Results are shown in Figure 1. Extravasated dye levels in TENS rabbits were markedly higher than those in controls. Analysis of profiles using an erfc-based diffusion-convection discrimination model2 showed that extravasation was convective. Discussion: These data are consistent with significant contribution to convective percolation of bone fluid through implanted scaffolds by muscle pump-driven extravasating fluid. They do not, however, answer two critical questions: 1) Is the magnitude of this convection a major component of flow through the scaffold? 2) What are the relative contributions of skeletal muscle-generated intravascular pressure solitons and incompressible fluid transmission of bone bending pressure to the convective flow observed? Additional studies with released gastrocnemius muscles are in progress

Objectives

Activation of the leptin pathway is closely correlated with human knee cartilage degeneration. However, the role of the long form of the leptin receptor (Ob-Rb) in cartilage degeneration needs further study. The aim of this study was to determine the effect of increasing the expression of Ob-Rb on chondrocytes using a lentiviral vector containing Ob-Rb.