Glycoprotein non-metastatic melanoma protein B is a potential biomarker for arthroplasty aseptic loosening stages

Background Endoprosthesis loosening is the major cause of arthroplasty failure. Currently, radiography, histo-pathological classification of the periprosthetic membrane, and microbiological examination are used retrospectively for diagnosis. Prospective options for early diagnosis are not available. The study presented here aimed to identify (prospective) molecular biomarkers of implant loosening in tissue samples from patients. Methods Four patient cohorts (primary and revision arthroplasty due to aseptic loosening 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. Synovial fluid, bone marrow, and blood were collected from 96 patients. Mesenchymal stromal cells (BM-MSCs) were isolated from the bone marrow of patients and cultivated until passage 3. Then, total RNA was isolated. Bulk RNA-sequencing (RNA-seq) was performed for 28 samples. Total gene expression patterns revealed by RNA-seq were compared between cohorts and differentially expressed genes were determined by DESeq2 analysis. Quantitative real-time PCR served to validate and extend RNA-seq data. ELISA assays were used to quantitate selected protein targets in the plasma of different body fluids. Western blots were performed for confirmation of the the molecular identity. Statistical analyses and correlation analyses were performed. Results Glycoprotein non-metastatic melanoma protein B mRNA ( GPNMB ) was significantly upregulated in BM-MSCs derived from revision patients compared to patients with primary implantations. Elevated GPNMB mRNA levels were confirmed with qRT-PCR. In synovial fluid plasma, the 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 or blood plasma. Western blotting detected a major protein band with a molecular weight of about 52.000 g/mol with two different GPNMB antibodies. The molecular weight of this domain detected by Western blotting was similar in the plasma fractions of synovial fluid, bone marrow and blood, indicating similar posttranslationally processed protein species. There was a moderate positive correlation between roentgenographically determined defect length and the relative protein levels of GPNMB in synovial fluid plasma. Conclusion GPNMB could be a potential biomarker for early detection of implant loosening, ahead of current diagnostic procedures.