Polycystin-1 Mediates Mechanical Strain-Induced Osteoblastic Mechanoresponses via Potentiation of Intracellular Calcium and Akt/β-Catenin Pathway

Abstract

Mechanical regulation of bone formation involves a complex biophysical process, yet the underlying mechanisms remain poorly understood. Polycystin-1 (PC1) is postulated to function as a mechanosensory molecule mediating mechanical signal transduction in renal epithelial cells. To investigate the involvement of PC1 in mechanical strain-induced signaling cascades controlling osteogenesis, PKD1 gene was stably silenced in osteoblastic cell line MC3T3-E1 by using lentivirus-mediated shRNA technology. Here, our findings showed that mechanical tensile strain sufficiently enhanced osteogenic gene expressions and osteoblastic proliferation. However, PC1 deficiency resulted in the loss of the ability to sense external mechanical stimuli thereby promoting osteoblastic osteogenesis and proliferation. The signal pathways implicated in this process were intracellular calcium and Akt/β-catenin pathway. The basal levels of intracellular calcium, phospho-Akt, phospho-GSK-3β and nuclear accumulation of active β-catenin were significantly attenuated in PC1 deficient osteoblasts. In addition, PC1 deficiency impaired mechanical strain-induced potentiation of intracellular calcium, and activation of Akt-dependent and Wnt/β-catenin pathways, which was able to be partially reversed by calcium ionophore A23187 treatment. Furthermore, applications of LiCl or A23187 in PC1 deficient osteoblasts could promote osteoblastic differentiation and proliferation under mechanical strain conditions. Therefore, our results demonstrated that osteoblasts require mechanosensory molecule PC1 to adapt to external mechanical tensile strain thereby inducing osteoblastic mechanoresponse, partially through the potentiation of intracellular calcium and downstream Akt/β-catenin signaling pathway.

Physiological mechanisms and therapeutic potential of bone mechanosensing

“…In contrast, exposure of Pkd1 or Pkd2 -deficient osteoblasts to an identical flow stimuli resulted in a significant attenuation of the calcium response curve in the heterozygous cells and complete loss of calcium influx in Pkd1 −/− and Pkd2 −/− osteoblasts. These findings have been confirmed by others [68, 80, 81]. …”

Polycystin-1 interacts with TAZ to stimulate osteoblastogenesis and inhibit adipogenesis

“…This schema proposes that mechanical forces, such as ECM stiffness, flow, and mechanical stretch, activate the PC1/PC2 complex (13,31,50,51,(63)(64)(65)(66), leading to stimulation of intracellular calcium, induction of PC1-CTT cleavage, and TAZ nuclear translocation to enhance Runx2-mediated induction of osteoblast gene transcription and to inhibit PPARγ and adipogenesis (33,39,42,44,46,67). Together this work identifies a new mechanosensing complex and the feasibility of targeting this complex for treating disorders caused by mechanical unloading and ageing.…”

“…For example, interaction of PC1 with other transcription factors (i.e., NFAT and STAT3) has been described in human osteoblasts that regulate cell proliferation/differentiation via induction of Runx2 (63,64). PC1 is also reported to mediate the response to mechanical strain via potentiation of intracellular calcium and Akt/β-catenin pathways in osteoblasts (65).…”

Biglycan mediates suture expansion osteogenesis via potentiation of Wnt/β-catenin signaling

“…Mechanical regulation of bone remodeling involves a complex biophysical process called mechanotransduction, including the perceptions of extracellular mechanical stimuli, their conversion biochemical cascades and ultimately adaptive responses of bone cells (Wang et al, 2014). Anchoring of bone cells to the ECM provides a capability to sense the extracellular biophysical changes (Thompson et al, 2012).…”

“…The immunoblotting was carried out as described before (Wang et al, 2014). Briefly, primary antibody (the antibody against active β-catenin (05-665) or β-actin (04-1116) from Merck Millipore; the antibody against Runx2 (ab76956), biglycan (ab49701), Lamin B1 (ab16048) or osteopontin (OPN) (ab8488) from Abcam; the antibody against β-catenin (610154) from BD Biosciences) was incubated overnight at 4 1C.…”