CK2.3 promotes bone formation and inhibits osteoclastogenesis through activation of ERK MAPK signaling pathway
Date
2020
Authors
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Journal ISSN
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Publisher
University of Delaware
Abstract
Bone morphogenetic protein 2 is a member of transforming growth factor beta super family. Bone morphogenetic protein 2 plays an important role in bone formation and skeletal development. Bone morphogenetic protein 2 is showed to be indispensable and required for the process of bone repair and fracture healing. Hence, FDA approved the use of human recombinant bone morphogenetic protein 2 in spinal fusion and open tibia fracture. Nevertheless, the use of human recombinant bone morphogenetic protein 2 has been under scrutiny due to rising concerns about its safety. One of the effects of bone morphogenetic protein 2 is that it also directly enhances the differentiation of osteoclasts. In fact, use of human bone morphogenetic protein 2 is shown to associated with increased bone resorption after spinal fusion. One of the aspects of the research in our laboratory is investigating the bone morphogenetic protein 2 signaling transduction in osteoclastogenesis. Understanding signaling events that lead to osteoclastogenesis will help paving the way for the development of an effective treatment for bone diseases such as osteoporosis. Osteoporosis is a bone disease that affected mostly the older population. It is estimated that one in two women and one in four men 50 or older will experience an osteoporotic fracture in their lifetime. Over 10 million adults in the United States have been diagnosed with osteoporosis. It is a silent disease. Patients do not often display symptoms until a fracture occurs. Current treatments of osteoporosis are divided into two categories: anti-resorptive treatments and anabolic treatments. However, despite having treatments, the mortality rate within a year of fracture has not been reduced and remains unchanged till today. Additionally, the rate of refracture also has not been reduced. Thus, there is a need for a new treatment that could improve the quality of life of patients. ☐ Our laboratory was the first to discover casein kinase 2 (CK2) as an activator of bone morphogenetic protein 2 signaling transduction. Our laboratory designed a novel peptide, CK2.3, to block the interaction between CK2 and BMPRIa and activate bone morphogenetic protein 2 signaling transduction. Our laboratory demonstrated that CK2.3 induced bone formation in vivo. Looking closer, we showed that CK2.3 suppressed osteoclastogenesis. Bone morphogenetic protein 2 is reported as a promoter of osteoclastogenesis. However, as an activator of bone morphogenetic protein 2 signaling transduction, suppression of osteoclastogenesis by CK2.3 was especially surprising. But the mechanism is unknown. Nevertheless, it showed that CK2.3 only activated certain signaling pathways downstream of bone morphogenetic protein 2 to result in the inhibition of osteoclastogenesis. The goal of this study was to elucidate the mechanism of CK2.3 inhibited osteoclastogenesis. Our hypothesis is CK2.3 inhibits RANKL-mediated osteoclastogenesis via upregulation of BMPRIa leading to the activation of the ERK signaling pathway that contribute to the enhancement of BMD in 6-month-old retired breeder mice and OVX rats. ☐ Elucidating the signaling pathways and cellular events down stream of bone morphogenetic protein 2 that lead to the inhibition of osteoclastogenesis will first and foremost contribute to the field of bone research. It contributes to the better understanding of the regulation of osteoclastogenesis by bone morphogenetic protein 2. From there, methods to modulate bone morphogenetic protein 2 signaling transduction can be developed to treat bone diseases such as osteopetrosis and osteoporosis. And such a method was employed in this study using the novel peptide CK2.3. ☐ First, I demonstrated the potential of CK2.3 as a novel treatment for osteoporosis in low bone mineral density 6-month-old retired breeder mice and 4-month-old ovariectomized rats. In both models, CK2.3 improved the trabecular bone mineral density and enhanced bone architecture. Additionally, for the first time I showed that CK2.3 suppressed osteoclastogenesis in vivo. An interesting result for this study was that I showed CK2.3 improved the stiffness of femoral shaft. It suggested that CK2.3 could improved fracture resistance. ☐ After establishing the in vivo effect of CK2.3 on mice and rats, I moved on to establishing the cellular mechanism of CK2.3 inhibited osteoclastogenesis. First, I established and optimized the culture condition of RAW264.7 cells as a model to study the cellular signaling events during osteoclastogenesis. Then, I showed that CK2.3 increased expression of BMPRIa on day 1 in RANKL-induced osteoclastogenesis. Additionally, CK2.3 promoted the activation of ERK on day 1. In fact, inhibition of MEK, an upstream activator of ERK, nullified the inhibitory effect of CK2.3 on RANKL-induced osteoclastogenesis. Even though CK2.3 increased the expression of BMPRIa on day 1, however, activation of SMAD was not observed. Hence, it showed that CK2.3 mediated the non-canonical ERK MAPK signaling pathway rather than the canonical SMAD signaling pathway in RANKL-induced osteoclastogenesis to inhibit it. ☐ To sum up, I demonstrated that CK2.3 could rescue the low bone mineral density phenotype of 6-month-old retired breeder mice and ovariectomized rats. Additionally, I established and optimized the culture condition of RAW264.7 cells for the study of cellular signaling events in osteoclasts. Lastly, I elucidated that CK2.3 upregulated bone morphogenetic protein receptor type 1a to activate ERK during the inhibition of RANKL-induced osteoclastogenesis. The results in this study show the signaling events down stream of bone morphogenetic protein 2 signaling transduction that led to inhibition of RANKL-meditated osteoclastogenesis. The knowledge that was gained from this study would contribute to the better understanding of the regulation of RANKL-induced osteoclastogenesis by bone morphogenetic protein 2 signaling transduction. It would help future researchers in the field of bone research.
Description
Keywords
BMP2, Bone research, CK2.3, Osteoporosis, Peptide treatment, Bone morphogenetic protein 2, Beta super family, Bone repair, Bone fracture healing, Skeletal development, Spinal fusion, Bone diseases, In vivo