Objectives

Cellular movement and relocalisation are important for many physiologic properties. Local mesenchymal stem cells (MSCs) from injured tissues and circulating MSCs aid in fracture healing. Cytokines and chemokines such as Stromal cell-derived factor 1(SDF-1) and its receptor chemokine receptor type 4 (CXCR4) play important roles in maintaining mobilisation, trafficking and homing of stem cells from bone marrow to the site of injury. We investigated the differences in migration of MSCs from the femurs of young, adult and ovariectomised (OVX) rats and the effect of CXCR4 over-expression on their migration.

Background

Osteoporosis and bone fractures lead to immobility, chronic pain and high patient care costs. Mesenchymal stem cells (MSCs) from postmenopausal women have a slower growth rate and osteogenic differentiation ability causing lower bone density and reduced fracture healing capacity compared to MSCs from premenopausal women. Cellular movement and relocalisation are necessary for many physiologic properties. Local MSCs from injured tissues and circulating MSCs are involved in fracture healing. Cytokines and chemokines such as SDF-1 and its receptor CXCR4 play important roles in maintaining mobilisation, trafficking and homing of stem cells from bone marrow to the site of injury. This study investigated the effect of CXCR4 over-expression on the migration of MSCs from ovariectomised, normal and young rats.

Introduction

Parathyroid hormone-related peptide (PTHrP) has been shown to be an important regulator of bone remodelling1. The aim of this study was to investigate the effect of the N-terminal domain of PTHrP (1–36) on osteogenic and angiogenic gene expression in human osteoblasts (HOB) and human bone marrow stromal cells (hBMSCs).

PURPOSE OF STUDY

20-70% of patients need blood transfusion postoperatively. There remain safety concerns regarding allogenic blood transfusion. Tranexamic acid (TA) is a synthetic antifibrinolytic agent that has been successfully used to stop bleeding in other specialties. We applied TA topically prior to the wound closure to find out the effect on blood loss as well as need for subsequent blood transfusion. This method of administration is quick, easy, has less systemic side effect and provides a higher concentration at the bleeding site.

We investigated the hypothesis that autologous bone marrow stromal cells (BMSC) sprayed on the surface of acetabular cups would improve bone formation and bone implant contact.

Total hip replacements were implanted in 11 sheep, randomly assigned to receive either acetabular implants sprayed with autologous BMSCs suspended in fibrin (study group) or fibrin only (control group). Sheep were sacrificed after six months and the acetabulum with the implant was retrieved and prepared for undcalcified histology. Implant bone contact in both groups was compared, by microscopically noting the presence or absence of new bone or fibrous tissue along the implant at 35 consecutive points (every 1000 μm). The observers undertaking the histological analysis were blinded.

Significantly increased bone implant contact was noted in the BMSC treated group 30.71% ± 2.95 compared to the control group 5.14% ± 1.67 (p = 0.014). The mean thickness of fibrous tissue in contact with the implant was greater at the periphery 887.21mm ± 158.89 and the dome 902.45mm ± 80.67 of the implant in the control group compared to the BMSC treated group (327.49mm ± 20.38 at the periphery and 739.1 mm ±173.72 at the centre). Conversely direct bone contact with the implant surface was significantly greater around the cups with stem cells.

BMSC sprayed on surface of implants improves bone implant contact. Spraying acetabular cups using stem cells could be used in humans where acetabular bone contact is compromised such as in revision procedures.

Despite advances in total hip arthroplasty, failure of acetabular cup remains a concern. The role of bone marrow stromal cells (BMSCs) to aid osseointegration of orthopaedic implants have been recently studied. We investigated the hypothesis that autologous BMSCs sprayed on the surface of acetabular cups would improve bone formation and bone implant contact.

Total hip replacements were implanted in 11 sheep, randomly assigned to receive either acetabular implants sprayed with autologous BMSCs suspended in fibrin (study group) or fibrin only (control group). Sheep were sacrificed after six months and the acetabulum with the implant was retrieved and prepared for undecalcified histology. Implant bone contact in both groups was compared microscopically, by noting the presence or absence of new bone or fibrous tissue along the implant at 35 consecutive points (every 1000 μm). The observers undertaking the histological analysis were blinded.

Significantly increased bone implant contact was noted in the BMSC treated group 30.71% ± 2.95 compared to the control group 5.14% ± 1.67 (p = 0.014). The mean thickness of fibrous tissue in contact with the implant was greater at the periphery 887.21mm ± 158.89 and the dome 902.45mm ± 80.67 of the implant in the control group compared to the BMSC treated group (327.49mm ± 20.38 at the periphery and 739.1 mm ± 173.72 at the centre). Conversely, direct bone contact with the implant surface was significantly greater around the cups with BMSCs.

Our data demonstrate that BMSC sprayed on surface of acetabular implants improves bone implant contact. Spraying acetabular cups using stem cells could be used in humans where acetabular bone contact is compromised such as in revision procedures.

Introduction: Recent studies have shown that MSCs can be isolated from the peripheral blood of many different species. Hematopoietic stem cell (HSC) mobilization from the bone marrow to the circulating bloodstream can be induced using granulocyte colony stimulating factors (G-CSF). As it has been shown that HSCs and MSCs have positive interactions with each other, it may be possible that G-CSF also promotes the release of circulating peripheral blood MSCs (PBMSCs). The hypothesis of this study was that G-CSF would increase the mobilization of peripheral blood-derived stromal-like cells.

Materials and Methods: Six sheep with normal hematological profiles were given 5& #956;g/kg Neupogen& #63721; (filgrastim, G-CSF) subcutaneously for five days. Pre- and post-G-CSF treatment, blood was taken 4, 12, 24, and 2 weeks post-treatment. PBMSCs were isolated from the blood and cells plated at a cell density of 4.0 x 10e4 nucleated cells/cm2. Fibroblastic colony forming units (CFU-F) were counted 7 and 14 days after initial culture. The cells were tested for their multipotency by treating them with osteogenic, adipogenic, and chondrogenic supplements, and staining with the Von Kossa, Oil Red ‘O,’ and Alcian Blue stains, respectively, to show differentiation down the different lineages.

Results: No CFU-F formation was observed in all blood samples taken before G-CSF therapy (0 CFU-F) after 7 and 14 days in culture. After G-CSF treatment, CFU-Fs were observed in blood samples taken 4, 12, and 336 hours (2 weeks) post-G-CSF. The CFU-F count was highest after 14 days in culture in the blood samples obtained 2 weeks post-G-CSF administration (1.027 ± 30.1353 CFU-F/cm2), compared to the lowest count, which was at 12 hours post-G-CSF treatment (0.064 ± 0.064 CFU-F/cm2). Hematology showed an increase in white blood cell (WBC), neutrophil, and eosinophil counts 24 hours after G-CSF administration. Two weeks post-G-CSF treatment, WBC, neutrophil, lymphocyte, and monocyte counts dropped back to normal range values. The highest number of CFU-F/cm2 were observed at this time. When WBC numbers were correlated with CFU-F counts using Pearson’s correlation co-efficient, the result was 0.523, a significant value (p=0.023) indicating that 27.4% of the WBC counts were related to CFU-F counts and vice versa. When time was accounted for as a third variable using the test for partial correlation coefficients, the co-efficient was found to be −0.0063, and was not significant (p=0.492). Expanded cells were fibroblastic in morphology, and upon differentiation were positive for the Von Kossa, Oil Red ‘O,’ and Alican Blue stains, indicating differentiation down the osteogenic, adipogenic, and chondrogenic lineages, respectively.

Discussion and Conclusions: We have shown that PBMSCs can be isolated after G-CSF administration in sheep, and that the numbers of CFU-F increase after WBC levels have returned to normal. A previous in vitro study proposed that the increased BMSC growth observed when co-cultured with CD45+ HSCs was due to positive interactions between HSCs and MSCs, indicating a possible steady-state balance. PBMSCs may have important future applications in bone tissue regeneration.

Introduction: Aseptic loosening at the bone-implant interface of THA acetabular components is a significant cause of implant failure. This loosening has been attributed either to wear particle-induced osteolysis or to the effects of joint fluid-pressure. It may be possible to prevent the loosening of implants by improving fixation between the bone and implant, or promoting the growth of a biological bony seal, in order to prevent the influx of wear particles or pressurized joint fluid. Additionally in revision implants it is important to promote osseointegration in situations where bone stock may be limited. The hypothesis of this study was spraying autologous BMSCs in fibrin glue onto the surface of HA-coated acetabular components would increase bone formation around the implant and improve bone-implant contact.

Materials and Methods: Bone marrow was aspirated from the iliac crest of six goats, and BMSCs isolated and expanded in vitro. 10 x 10e6 BMSCs were suspended in reconstituted thrombin pre-operatively. A standard posterior approach was used. The acetabular shell was then coated with 2 ml of fibrin glue, with (n=6) or without 10 x 10e6 autologous BMSCs (n=6), and the acetabular component impacted into position. Antibiotic and analgesic prophylaxes were carried out. All animals were weight bearing within 48 hours post-operatively. Walking and ground reaction forces were assessed pre-operatively, as well as 6 and 12 weeks post-operatively. Results were expressed as a percentage of force transmitted through the right leg versus the left leg. After 12 weeks, the acetabulae were retrieved, and processed for histology. The percentage of new bone around the cups was measured within 5 radial zones, using image analysis. Bone-implant contact was also analysed between the new bone and implant surface. Mann Whitney U test was used to show statistical significance.

Results: New bone formation in Zone 5 showed a significant increase in the BMSC group (71.97±10.91%), when compared to the controls (23.85±15.13%, p=0.028). The other zones did not show a significant difference. Overall new bone growth in the BMSC group was 30% greater than the control group (71.42±8.97% and 54.22±16.56%, respectively, p=0.58). Bone-implant contact was significantly improved in the BMSC group (20.03±4.64%), in contrast to the control group (13.71±8.32%, p=0.027). With regards to the force plate analysis, there was no significant difference in loading between groups at both 6 weeks (Controls-79.74±3.63%, BMSCs-59.39±9.33%, p=0.086) and 12 weeks (Controls-86.0%±2.85%, BMSCs-62.33±5.12%, p=0.055).

Discussion and Conclusions: In this study, overall bone growth was greater when cups were treated with BMSCs. Bone-implant contact was significantly improved as well. This study has clinical applications, as using MSCs in fibrin glue promotes a bony seal in contact with the implant which may prevent the migration of particles, or joint fluid, decreasing the likelihood of aseptic loosening of THAs, and improving their longevity. Also, this technique may improve fixation in situations where bone stock is reduced.

Introduction: The survival of massive endoprosthesis replacements is not as successful as conventional joint replacements. The main cause of failure of these implants is aseptic loosening. Bone in-growth onto the implant collar on the shaft of the prosthesis adjacent to the transaction site has been correlated with a decrease in radiolucent lines adjacent to the intramedullary stem and reduced implant loosening. We propose that bone contact and in-growth to the collar may be further enhanced with tissue engineering techniques. The hypothesis of this study was that autologous mesenchymal stem cells (MSCs) suspended within fibrin glue and sprayed onto hydroxyapatite (HA)-coated collars of massive prosthesis will augment bone growth and contact to the implant in an ovine model.

Materials and Methods: MSCs were isolated and expanded in vitro from the iliac crest of six adult sheep. Pre-implantation, 2 x 106 autologous MSCS were suspended in thrombin. During surgery, this mixture was combined with fibrinogen and sprayed onto the proximal and distal HA-coated collars of tibial midshaft prostheses using pressurized air. The implants were cemented into the right hind limb of twelve sheep, six of which received MSCs. Radiographs were taken at 2, 4 and 6 months and bone area within defined regions quantified using image analysis software. After six months, specimens were retrieved and processed for undecalcified histology. Transverse thin sections were prepared through the centre of each collar. Image analysis was used to quantify bone area and contact. Mann Whitney U tests were used for comparative statistical analysis, where p< 0.05 was classified as significant.

Results: Anterior-posterior (AP) radiographs taken at 2, 4, and 6 months showed that animals treated with MSCs produced more bone adjacent to the shaft of the implant. Analysis of bone area on both AP and medio-lateral (ML) radiographs taken after sacrifice showed that stem cell-treated implants encouraged significantly more total bone around the implants at 6 months than the control group (171.94 ± 29.04 mm2, and 87.51 ± 9.81 mm2 bone area, respectively, p = 0.016). Analysis of histological sections shows a significant increase in bone area around midshafts treated with MSCs, compared to the implant controls (53.99 ± 10.64 mm2, and 21.07 ± 7.34 mm2, respectively; p = 0.020). The average surface area contact between the midshaft and bone was almost doubled in the MSC-implant group (19.83 ± 8.73 % contact) than in the control group (8.667 ± 8.667 %, p = 0.196). In the MSC group bone was seen deep within the grooves of the HA coated collar whilst a fibrous soft tissue layer separated the newly formed bone in the control group.

Conclusion: Bone contact and in-growth to massive endoprostheses was significantly improved by spraying the implant with autologous MSCs suspended in fibrin glue. Enhanced fixation using stem cells may help prevent aseptic loosening in these massive implants.