Intermittent parathyroid hormone (1–34) application regulates cAMP-response element binding protein activity to promote the proliferation and osteogenic differentiation of bone mesenchymal stromal cells via the cAMP/PKA signaling pathway

摘要

The potential effects of intermittent parathyroid hormone (–) [PTH (–)] administration on bone formation have previously been investigated. A number of studies have suggested that the cyclic adenosine monophosphate/protein kinase A (cAMP/PKA) pathway is associated with PTH-induced osteogenic differentiation. However, the precise signaling pathways and molecular mechanism by which PTH (–) induces the osteogenic differentiation of bone mesenchymal stromal cells (BMSCs) remain elusive. The purpose of the present study was to investigate the mechanism underlying the effect of intermittent PTH (–) application on the proliferation and osteogenic differentiation of BMSCs. BMSCs were randomly divided into four groups, as follows: Osteogenic medium (control group); osteogenic medium and intermittent PTH (–); osteogenic medium and intermittent PTH (–) plus the adenylyl cyclase activator forskolin; and osteogenic medium and intermittent PTH (–) plus the PKA inhibitor H-89. A cell proliferation assay revealed that PTH (–) stimulates BMSC proliferation via the cAMP/PKA pathway. Furthermore, reverse transcription-quantitative polymerase chain reaction, alkaline phosphatase activity testing and cell examination using Alizarin Red S staining demonstrated that PTH (href="#b1-etm-0-0-3177" rid="b1-etm-0-0-3177" class=" bibr popnode tag_hotlink tag_tooltip" id="__tag_536406846">1–href="#b34-etm-0-0-3177" rid="b34-etm-0-0-3177" class=" bibr popnode tag_hotlink tag_tooltip" id="__tag_536406937">34) administration promotes osteogenic differentiation and mineralization, mediated by the cAMP/PKA pathway. Crucially, the results of western blot analyses suggested that PTH (href="#b1-etm-0-0-3177" rid="b1-etm-0-0-3177" class=" bibr popnode tag_hotlink tag_tooltip" id="__tag_536406962">1–href="#b34-etm-0-0-3177" rid="b34-etm-0-0-3177" class=" bibr popnode tag_hotlink tag_tooltip" id="__tag_536406889">34) treatment and, to a greater degree, PTH (href="#b1-etm-0-0-3177" rid="b1-etm-0-0-3177" class=" bibr popnode tag_hotlink tag_tooltip" id="__tag_536406819">1–href="#b34-etm-0-0-3177" rid="b34-etm-0-0-3177" class=" bibr popnode tag_hotlink tag_tooltip" id="__tag_536406835">34) plus forskolin treatment caused an increase in phosphorylated cAMP response element binding protein (p-CREB) expression, but the effect of PTH on p-CREB expression was blocked by H-89. In conclusion, the current study demonstrated that intermittent PTH (href="#b1-etm-0-0-3177" rid="b1-etm-0-0-3177" class=" bibr popnode tag_hotlink tag_tooltip" id="__tag_536406952">1–href="#b34-etm-0-0-3177" rid="b34-etm-0-0-3177" class=" bibr popnode tag_hotlink tag_tooltip" id="__tag_536406823">34) administration regulates downstream proteins, particularly p-CREB, in the cAMP/PKA signaling pathway, to enhance the proliferation, osteogenic differentiation and mineralization of BMSCs.