Age-Related Low Bone Mineral Density in C57BL/6 Mice Is Reflective of Aberrant Bone Morphogenetic Protein-2 Signaling Observed in Human Patients Diagnosed with Osteoporosis

Abstract

Osteoporosis (OP) is a bone disorder characterized by decreased bone mineral density (BMD). Bone Morphogenetic Protein-2 (BMP-2) injections are used to promote bone formation in OP patients. However, patients are unresponsive to BMP-2 while displaying an upregulation of BMP Receptor Type 1a (BMPRIa) and protein kinase CK2α (CK2α). A synthetically produced peptide named casein kinase 2.3 (CK2.3) utilizes the BMP-signaling pathway as it enhances osteogenesis of primary osteoblasts isolated from OP patients, whereas BMP-2 does not. Although shown in OP patients, there is currently no reliable mouse model to study BMP-2 and CK2.3 signaling. In this publication, we show that BMPRIa was required for CK2.3-mediated osteogenesis in C2C12 cells with a CRISPR-Cas9-mediated gene knockout for BMPRIa. We utilized the C57BL/6 (B6) mouse strain as an aging-model to study aberrant BMP-2 signaling, demonstrating that, like OP patients, in 15 and 20-month mice, BMP-2 did not increase bone growth and displayed upregulated BMPRIa and CK2α protein expression. Furthermore, CK2.3 enhanced osteogenesis and decreased osteoclastogenesis in all age groups, whereas BMP-2 only increased mineralization in 6-month mice while increasing osteoclast formation in all age groups. These data demonstrated that aging B6 mice were a reliable model and mimicked data obtained from OP patients.

Methyl-Beta-Cyclodextrin Restores Aberrant Bone Morphogenetic Protein 2-Signaling in Bone Marrow Stromal Cells Obtained from Aged C57BL/6 Mice

“…To ensure B6 mice are a reliable model to investigate aberrant BMP-signaling, the expression of BMPRIa must be further established. We first confirmed that the BMSCs of US 15-month B6 mice overexpress BMPRIa compared to US 6-month mice via western blotting and confocal microscopy, confirming previous data (Figure 1) [67,81]. This overexpression may be caused by aberrant BMPsignaling, and the cells are compensating for the lack of signaling and thereby upregulating BMPRIa expression.…”

“…It was recently demonstrated that primary osteoblasts isolated from OP patients overexpress BMPRIa compared to control patients via immunofluorescent staining and western blotting [67]. Additionally, this overexpression of BMPRIa is also observed in the BMSCs isolated from 15-month mice compared to 6-month mice as illustrated by confocal microscopy [81]. However, the total protein concentration of BMPRIa from whole cell lysates has never been determined and should be explored to confirm this overexpression.…”

“…One model that can be utilized to overcome these obstacles is the C57BL/6 (B6) mouse. These mice display low peak BMD and recent data demonstrates that they are a comparable model to study human OP [81][82][83]. Further, the localization of BMP-2 in the bone marrow stromal cells (BMSCs) within these mice is unknown and requires further attention.…”

Scoping Pleiotropy of CK2 in Musculoskeletal Disorders for a Novel Targeting Approach

“…In 6-to-9-day-old C57BL/6J mice injected with CK2.3 via the calvaria and 8-week-old mice injected via the tail vein, mice showed increased bone formation and increased bone mineral density. At the same time, osteoclast activity appeared to be suppressed, indicating CK2.3 has the potential to alleviate the symptoms of OP by simultaneously enhancing osteoblast activity and inhibiting osteoclast activity, inducing bone growth and repair [174][175][176]. The principle of design of BMPRIa biomimetic peptides CK2.1 and CK2.3 and their effects are summarized in Figure 3.…”