Post-traumatic osteoarthritis (PTOA) is a subset of osteoarthritis, which occurs secondary to traumatic joint injury which is known to cause pathological changes to the osteochondral unit. Articular cartilage degradation is a primary hallmark of OA, and is normally associated with end-stage disease. However, subchondral bone marrow lesions are associated with joint injury, and may represent localized bone microdamage. Changes in the osteochondral unit have been traditionally studied using explant models, of which the femoral-head model is the most common. However, the bone damage caused during harvest can confound studies of microdamage. Thus, we used a novel patellar explant model to study osteochondral tissue dynamics and mechanistic changes in bone-cartilage crosstalk. Firstly, we characterized explants by comparing patella with femoral head models. Then, the patellar explants (n=269) were subjected to either mechanical or inflammatory stimulus. For mechanical stimulus 10% strain was applied at 0.5 and 1 Hz for 10 cycles. We also studied the responses of osteochondral tissues to 10ng/ml of TNF-α or IL-1β for 24hrs. In general the findings showed that patellar explant viability compared extremely well to the femoral head explant. Following IL-1β or TNF-α treatment, MMP13, significantly increased three days post exposure, furthermore we observed a decrease in sulfate glycoaminoglycan (sGAG) content. Bone morphometric analysis showed no significant changes. Contrastingly, mechanical stimulation resulted in a significant decrease sGAG particularly at 0.5Hz, where an increase in MMP13 release 24hrs post stimulation and an upregulation of bone and cartilage matrix degradation markers was observed. Furthermore, mechanical stimulus caused increases in TNF-α, MMP-8, VEGF expression. In summary, this study demonstrates that our novel patella explant model is an excellent system for studying bone-cartilage crosstalk, which responds well to both mechanical and inflammatory stimulus and is thus of great utility in the study of PTOA.
Traditionally, osteoarthritis (OA) has been associated mostly with degradation of cartilage only. More recently, it has been established that other joint tissues, in particular bone, are also centrally involved. However, the link between these two tissues remains unclear. This relationship is particularly evident in post-traumatic OA (PTOA), where bone marrow lesions (BMLs), as well as fluctuating levels of inflammation, are present long before cartilage degradation begins. The process of bone-cartilage crosstalk has been challenging to study due to its multi-tissue complexity. Thus, the use of explant model systems have been crucial in advancing our knowledge. Thus, we developed a novel patellar explant model, to study bone cartilage crosstalk, in particular related to subchondral bone damage, as an alternative to traditional femoral head explants or cylindrical core specimens. The commonly used osteochondral explant models are limited, for our application, since they involve bone damage during harvest. The specifics aim of this study was to validate this novel patellar explant model by using IL-1B to stimulate the inflammatory response and mechanical stimulation to determine the subsequent developments of PTOA. Lewis rats (n=48) were used to obtain patellar and femoral head explants which were harvested under an institutional ethical approval license. Explants were maintained in high glucose media (containing supplements), under sterile culture conditions. Initially, we characterised undamaged patellar explants and compared them with the commonly used femoral head. First, tissue viability was assessed using an assay of metabolic activity and cell damage. Second, we created chemical and mechanical damage in the form of IL-1B treatment, and mechanical stimulation, to replicate damage. Standard biochemical assays, histological assays and microstructural assays were used to evaluate responses. For chemical damage, explants were exposed to 10ng/ml of IL-1B for 24 hours at 0, 1, 3 and 7 days after harvesting. For mechanical damage, tissues were exposed to mechanical compression at 0.5 Hz, 10 % strain for 10 cycles, for 7 days. Contralateral patellae served as controls. In both groups, sGAG, ADAMTS4, and MMP-13 were measured as an assessment of representative cartilage responses while ALP, TRAP and CTSK were assessed as a representative of bone responses. In addition to this, histomorphometric, and immunohistochemical, evaluations of each explant system were also carried out.Introduction and Objective
Materials and Methods
Low back pain (LBP), caused by intervertebral disc (IVD) degeneration represents one of the most significant socioeconomic conditions facing Western economies. Novel regenerative therapies, however, have the potential to restore function and relieve pain. We have previously shown that stimulation of adipose-derived stem cells (ASCs) with growth differentiation factor-6 (GDF6) promotes differentiation to nucleus pulposus (NP) cells of the IVD, offering a potential treatment for LBP. The aims of this study were to i) elucidate GDF6 cell surface receptor profile and signalling pathways to better understand mechanism of action; and (ii) develop a microparticle (MP) delivery system for GDF6 stimulation of ASCs. GDF6 receptor expression by ASCs (N=6) was profiled through western blot, immunofluorescence (IF) and flow cytometry. Signal transduction through Smad1/5/9 and non-Smad pathways following GDF6 (100ng/ml) stimulation was assessed using western blotting and confirmed using pathway specific blockers and type II receptor sub-unit knockdown using CRISPR. Release kinetics of GDF6 from MPs was calculated (BCA assay, ELISAs) and ASC differentiation to NP cells was assessed. BMPR profiling revealed high BMPR2 expression on ASCs. GDF6 stimulation of ASCs resulted in significant increases in Smad1/5/9 and Erk phosphorylation, but not p38 signalling. Blocking GDF6 signalling confirmed differentiation to NP cells required Smad phosphorylation, but not Erk. GDF6 release from MPs was controlled over 14days
Currently, there is a focus on the development of cell based therapies to treat intervertebral disc (IVD) degeneration, particularly for regenerating/repairing the central region, the nucleus pulposus (NP). Recently, we demonstrated that GDF6 promotes NP-like differentiation in mesenchymal stem cells (MSCs). However, bone marrow- (BM-MSCs) and adipose- (Ad-MSCs) showed differential responses to GDF6, with Ad-MSCs adopting a more NP-like phenotype. Here, we investigated GDF6 signalling in BM-MSCs and Ad-MSCs, with the aim to improve future IVD stem cell therapies. GDF6 receptor expression in patient-matched BM-MSCs and Ad-MSCs (N=6) was profiled through western blot and immunocytochemistry (ICC). GDF6 signal transduction was investigated through stimulation with 100 ng ml−1 GDF6 for defined time periods. Subsequently smad1/5/9 phosphorylation and alternative non-smad pathway activation (phospho-p38; phospho-Erk1/2) was analysed (western blot, ELISA). Their role in inducing NP-like gene expression in Ad-MSCs was examined through pathway specific inhibitors.Background
Methods
Stem cell therapy has been suggested as a potential regenerative strategy to treat IVD degeneration and GDF6 has been shown to differentiate adipose-derived stem cells (ASCs) into an NP-like phenotype. However, for clinical translation, a delivery system is required to ensure controlled and sustained GDF6 release. This study aimed to investigate the encapsulation of GDF6 inside novel microparticles (MPs) to control delivery and assess the effect of the released GDF6 on NP-like differentiation of human ASCs. GDF6 release from PLGA-PEG-PLGA MPs over 14 days was determined using BCA and ELISA. The effect of MP loading density on collagen gel formation was assessed through SEM and histological staining. ASCs were cultured in collagen hydrogels for 14 days with GDF6 delivered exogenously or via microspheres. ASC differentiation was assessed by qPCR for NP markers, glycosaminoglycan production (DMMB) and immunohistochemistry.Background
Methods
Signalling by growth differentiation factor 6 (GDF6/BMP13) has been implicated in the development and maintenance of healthy NP cell phenotypes and GDF6 mutations are associated with defective vertebral segmentation in Klippel-Feil syndrome. GDF6 may thus represent a promising biologic for treatment of IVD degeneration. This study aimed to investigate the effect of GDF6 in human NP cells and critical signal transduction pathways involved. BMP receptor expression profile of non-degenerate and degenerate human NP cells was determined through western blot, immunofluorescence and qPCR. Phosphorylation statuses of Smad1/5/9 and non-canonical p38 MAPK and Erk1/2 were assessed in the presence/absence of pathway blockers. NP marker and matrix degrading enzyme gene expression was determined by qPCR following GDF6 stimulation. Glycosaminoglycan and collagen production were assessed through DMMB-assay and histochemical staining.Background
Methods
Hip replacement is a very successful operation and the outcome is usually excellent. There are recognised complications that seem increasingly to give rise to litigation. This paper briefly examines some common scenarios where litigation may be pursued against hip surgeons. With appropriate record keeping, consenting and surgical care, the claim can be successfully defended if not avoided. We hope this short summary will help to highlight some common pitfalls. There is extensive literature available for detailed study.
Clinical success of total knee arthroplasty is correlated with correct orientation of the components. Controversy remains in the orthopaedic community as to whether the intramedullary or extramedullary tibial alignment guide is more accurate in the tibial cut. Is there any difference between intramedullary and extramedullary jigs to achieve better accuracy of the tibial components in total knee replacements? A retrospective study done on 100 patients during the time period 2007 to 2010. The 100 knee replacements were done by the same surgeon, where 50 patients had the intramedullary tibial alignment guide and the other 50 had the extramedullary one. The tibiofemoral angle was measured pre-operatively as well as post operatively, the tibial alignment angle was measured post operatively then the results were statistically analysed using the SPSS. There was no significant difference between both groups regarding the tibial alignment angles. Both techniques proved accurate in producing an acceptable post operative tibial component alignment angle. We recommend orthopaedic surgeons choose either technique knowing that accuracy levels are similar. The debate between intramedullary and extramedullary tibial cutting jigs/guides/ devices continues and most orthopaedic surgeons will use their preferred technique and will continue to achieve good post operative results as we have found in our centre. Our study is rare due to the fact we have a single surgeon performing both techniques, therefore controlling for any surgical experience or operating technique differences.
Supero-medial migration was seen in 27 (49%) of cases, demarcation without migration was seen in 18 cases (33%) and supero-lateral migration was seen in 7 (13%) cases. There were 2 (4%) socket fatigue fractures due to wear. There was 1 (2%) patient with a worn socket and no loosening. Reconstruction was achieved by impaction bone grafting alone in 25 cases, IBG and a block allograft in 9 cases, cement alone in 8 cases and IBG with a rim mesh in 4 cases. In cases where the supero-lateral margin of the socket was covered by host bone, failure always occurred by demarcation alone or in association with supero-medial migration. Rim defects significant enough to require reconstruction were seen in only 4 of these 45 patients (9%). Failure by supero-lateral migration was only seen in the cases of DDH where the socket was left uncovered or where the socket had fractured.
The pattern of socket failure can be reliably predicted from the original post-operative x-rays. Care should be taken to ensure adequate supero-lateral coverage in order that demarcation and migration leave an intact rim for reconstruction.
Recently, there has been a reluctance to perform hip arthrodesis. The number of patients requiring the conversion from hip arthrodesis to arthroplasty has also decreased. We present the functional results following conversion of hip arthrodesis to total hip arthroplasty at a specialist hip centre. 76 patients who underwent conversion of hip arthrodesis to total hip arthroplasty between 1963 and 2000 at the Centre for Hip Surgery, Wrightington Hospital, were included in this retrospective study. 9 patients died of unrelated causes and 7 patients were lost to follow up. The functional scoring was performed using the Merle d’Aubigné and Postel score. The mean age at the time of surgical hip arthrodesis was 16.7 years and at the time of conversion was 48.7 years. Back pain is the most common indication for the conversion. All the patients were pleased with the clinical outcome following conversion to Arthroplasty. 6 patients had postoperative complications. The mean Merle d’Aubigné and Postel score increased from 8.97 to 13.46 at the latest follow-up. The mean wear rate was 0.06 mm/year. Survival of hip arthroplasty was 92.78 % at 18 years.
To determine socket survivorship in DDH based on the severity of hip dysplasia, we carried out a retrospective study of 283 cemented total hip replacements carried out at Wrightington. The hips were classified according to the Crowe and Hartofilakidis classifications. Revision was used as the end point for prosthetic survivorship. The results were analysed statistically using SPSS for Windows The mean age at time of surgery was 42.6 years with a mean follow-up of 15.7 years. The acetabulum was grafted in 46 cases. The commonest cause for revision was aseptic loosening of the acetabular component (88.3%). 254 procedures were carried out through a transtrochanteric approach with a direct lateral approach used for the remaining mildly dysplastic hips. At 10 years 5.3% of dysplastic, 14.8% of low dislocation and 51.1 % of high dislocation hips were revised.. At 10years 6% of Crowe Type1, 8.5% of Type2, 25.5% of Type3 and 39.2% of Type4 hips were revised. At 20 years 24% of dysplastic, 45% of low dislocation and 88% of high dislocation hips were revised. At 20years 27.3% of Crowe Type1, 29.3% of Type2, 63.3% of Type3 and 84.4% of Type4 hips were revised. The 20 year survival of patients less than 50 years of age at the time of surgery was 61% as compared to 92% survival in patients more than 50 years of age. The mean age of patients in the revised group was 35 years as compared to 45 years in the non-revised group.
Patients undergoing total hip replacement (THR) often require further orthopaedic surgery including other primary lower limb joint replacements and revision surgery in their lifetime. We analysed the 10-year data of 552 patients who underwent primary total hip replacement between April 1991 and March 1992 at our institute. Data were available for all patients before the index operation. 77% of patients attended their 5-year review and 67% attended their 10-year review. 233 (42%) had had or subsequently had the opposite hip replaced. 30 patients (5%) had a knee replaced and 19 (3%) had both knees replaced. 4.4% underwent revision surgery.
We present the case of a patient with Rheumatoid Arthritis who underwent a right total hip replacement as a young adult. At the time of surgery there was an intra-operative femoral fracture and the prosthesis and cement breached the cortex of the proximal femur postero-medially. The fracture was detected on the post-operative film and the patient was treated non-operatively until the fracture consolidated. Despite having rheumatoid arthritis our patient went on to an active adult life having a family and she worked full time with this hip replacement. She subsequently required a socket revision at 15 years post index surgery and at the time the femoral component was well fixed, not scratched and left in situ. Currently, the revision socket remains satisfactory, the stem still appears well fixed and clinically the patient is well.
There have been no complications or skin reactions related to this method of skin preparation. There has been no significant difference in the incidence of early post operative wound infection.