Project description:Transcriptome profiling was performed on muscle biopsies from patients immediately before Total Knee Arthroplasty and two hours after TKA and tourniquet application.
Project description:Ischaemic preconditioning is a method of protecting tissue against ischaemia-reperfusion injury. It is an innate protective mechanism that increases a tissue's tolerance to prolonged ischaemia when it is first subjected to short burst of ischaemia and reperfusion. It is thought to provide this protection by increasing the tissue's tolerance to ischaemia, therby reducing oxidative stress, inflammation and apoptosis in the preconditioned tissue. We used microarrays to investigate the genomic response induced by ischaemic preconditioning in muscle biopsies taken from the operative leg of total knee arthroplasty patients in order to gain insight into the ischaemic preconditioning mechanism. Patients undergoing primary knee arthroplasty were randomised to control and treatment (ischaemic preconditioning) groups. Patients in the treatment group received a preconditioning stimulus immediately prior to surgery. The ischaemic preconditioning stimulus consisted of three five-minute periods of tourniquet insufflation on the lower operative limb, interrupted by five minute periods of reperfusion. All patients had a tourniquet applied to the lower limb after the administration of spinal anaesthesia, as per normal protocol for knee arthroplasty surgery. Muscle biopsies were taken from the quadriceps muscle of the operative knee at the immediate onset of surgery (T0) and at 1 hour into surgery (T1). Total RNA was extracted from biospies of four control and four treatment patients and hybridised to the Affymetrix Human U133 2.0 chip.
Project description:To investigate the physiologic responses of whole osteoarthritic synovium to cycle tensile strain. We obtained 3 matched synovial tissue samples from 6 patients undergoing total knee arthroplasty, subjected them to cyclic tensile strain, and then performed gene expression profiling analysis using RNA seq from each cyclic tensile strain protocol.
Project description:These data are from a preclinical model of total knee arthroplasty (TKA) performed in healthy rats. One day after surgery, ipsilateral dorsal root ganglia (DRG) from L3 and L4 were collected for RNA extraction and sequencing.
Project description:OBJECTIVE: To identify the changes in gene expression elicited by miR-27b-3p mimic transfection of human osteoarthritis (OA) primary fibroblast-like synoviocytes (FLS) obtained from patients undergoing knee arthroplasty. Cells were cultured, and treated for 48 h with miR-27b-3p mimic or Cel-miR-39-3p control mimic. Total RNA was then extracted, and mRNA libraries were prepared using Illumina's TruSEQ stranded total RNA library preparation kit, pooled together, and sequenced on Illumina's NextSeq550.
Project description:Analysis of gene expression profiles of ACL-MSCs derived from total knee arthroplasty (TKA) and ACL reconstruction patients. We hypothesized that the proportion of MSCs in ACL samples correlates negatively with donor age, i.e., the proportion of MSCs in ACL samples are higher in younger patients undergoing ACL reconstruction than in older patients undergoing TKA. Results provide additional information on the the effect of age on MSC properties, in particular, the differential regulation of genes encoding components of the extracellular matrix (ECM).
Project description:Background: Endoprosthesis loosening is the major cause of arthroplasty failure. Currently, radiography and histo-pathological classification of the periprosthetic membrane are used retrospectively for diagnosis. Prospective options for early diagnosis and prevention of loosening are not available. In order to approach this challenge, the study presented here aimed to identify molecular biomarkers in single-centre prospectively collected cell and plasma samples from patients. Methods: Four patient cohorts (primary and revision arthroplasty of hip or knee) were defined. Aseptic loosening of implants was assessed by the standardised approach of the Knee Society Total Knee Arthroplasty Roentgenographic Evaluation and Scoring System (KSRESS) which is based on radiolucent lines. Synovial fluid, bone marrow, and blood were collected from a total of 96 patients. Bulk RNA-sequencing (RNA-seq) of mesenchymal stem cells (MSCs) isolated from the bone marrow of patients was performed for 28 samples. The data was analysed by bioinformatic tools. Quantitative real-time PCR served to validate and extend RNA-seq data. Western blots were performed for molecular differentiation. ELISAs were used to quantitate selected protein targets. Results: RNA-seq of MSCs identified glycoprotein non-metastatic melanoma protein B (GPNMB) as significantly upregulated in the cells from revision patients compared to patients with primary implantations. The protein encoded by this gene is plasma membrane-standing; its extracellular domain may be cleaved and secreted. In synovial fluid plasma, the soluble GPNMB protein exhibited increased levels in patients undergoing revision surgery compared to patients subjected to primary arthroplasty. No significant differences were observed in bone marrow plasma and blood plasma. The molecular species detected by Western blotting were similar in the plasma fractions of synovial fluid, bone marrow and blood. Conclusions: The use of GPNMB as a biomarker may be of prime importance for early detection of implant loosening, ahead of radiographic failure demonstration. Translational potential of this article: GPNMB has the potential to act as a prospective and quantitative parameter to assess different stages of implant loosening. Radiography and histo-pathological classification of the periprosthetic membrane, on the contrary, are used retrospectively for diagnosis and are subjective measures, performed mainly at the end of a full loosening process.