A quantitative serum biomarker of circulating collagen X effectively correlates with endochondral fracture healing

Abstract

Currently, there are no standardized methods for quantitatively measuring fracture repair. Physicians rely on subjective physical examinations and qualitative evaluation of radiographs to detect mineralized tissue. Since most fractures heal indirectly through a cartilage intermediate, these tools are limited in their diagnostic utility of early repair. Prior to converting to the bone, cartilage undergoes hypertrophic maturation, characterized by the deposition of a provisional collagen X matrix. The objective of this study was to characterize the kinetics of a novel collagen X biomarker relative to other biological measurements of fracture healing using a murine model of endochondral fracture repair in which a closed, mid-shaft tibia fracture was created using the classic drop-weight technique. Serum was collected 5 to 42 days postfracture in male and female mice and compared to uninjured controls (n = 8-12). Collagen X in the serum was quantified using a recently validated ELISA-based bioassay ("Cxm") 1 and compared to genetic and histological markers of fracture healing and inflammation. We found the Cxm biomarker reliably increased from baseline to a statistically unique peak 14 days post-fracture that then resolved to prefracture levels by 3 weeks following injury. The shape and timing of the Cxm curve followed the genetic and histological expression of collagen X but did not show a strong correlation with local inflammatory states. Assessment of fracture healing progress is crucial to making correct and timely clinical decisions for patients. This Cxm bioassay represents a minimally invasive, inexpensive technique that could provide reliable information on the biology of the fracture to significantly improve clinical care.

Methodology, selection, and integration of fracture healing assessments in mice

“…26 markers, often reported in fracture repair studies, include tartrateresistant acid phosphatase (TRAP), receptor activator of nuclear factor kappa beta ligand (RANKL), and c-terminal telopeptide (CTX) reflecting osteoclast activity, alkaline phosphatase, procollagen type 1 N-terminal polypeptide, and osteocalcin reflecting osteoblast activity, CXM and collagen X reflecting chondrocyte activity and transforming growth factor beta, tumor necrosis factor-a (TNF-a), and vascular endothelial growth factor reflecting immune activity and angiogenesis. [29][30][31][32] Assessment of these biomarkers may elucidate how underlying fracture repair processes are altered in various circumstances, manifesting in distinct repair outcomes. [29][30][31][32] Furthermore, recent reviews have investigated the potential of these biomarkers to serve as independent predictors of fracture repair success.…”

“…[29][30][31][32] Assessment of these biomarkers may elucidate how underlying fracture repair processes are altered in various circumstances, manifesting in distinct repair outcomes. [29][30][31][32] Furthermore, recent reviews have investigated the potential of these biomarkers to serve as independent predictors of fracture repair success. [33][34][35] This is of particular interest as reliable correlations could enable earlier clinical identification of nonunion, improving clinical outcomes.…”

Parathyroid hormone promotes cartilage healing after free reduction of mandibular condylar fractures by upregulating Sox9

“…[21][22][23] Collagen X (ColX) is the hallmark of hypertrophic cartilage and is specifically expressed as cartilage converts to bone during development and endochondral fracture healing. 24 A number of studies indicated that PTH has potential for cartilage repair through promoting the proliferation of hypertrophic chondrocytes 25 and inhibiting chondrocyte hypertrophy. [26][27][28] The changes in the expression levels of Sox9, Col2a1, MMP-13, and ColX during the cartilage healing process after mandibular condylar fractures and whether the application of PTH can regulate Sox9 expression and promote condylar cartilage healing need to be studied in more depth.…”

Differences in Fracture Healing Between Female and Male C57BL/6J Mice

“…Some authors report more rapid bone regeneration in male rats and mice compared to females ( Mehta et al, 2011 ; Deng et al, 2020 ). However, there are also studies showing no sex-specific differences in murine fracture healing ( Collier et al, 2020 ; Working et al, 2020 ). In many studies, data generated from female and male mice are pooled together, simply stipulating that there would be no sex-specific difference in the healing process.…”