P2Y2 receptors regulate osteoblast mechanosensitivity during fluid flow.

Area of Science:

• Biochemistry
• Cell Biology
• Biophysics

Background:

• Mechanical stimulation is crucial for osteoblast function and bone formation.
• Osteoblasts release ATP in response to mechanical load, activating purinergic receptors.
• Fluid shear stress (FSS) induces actin stress fiber formation (ASFF) in osteoblasts, potentially via P2Y2 receptor (P2Y2R) activation.

Purpose of the Study:

• To investigate the role of P2Y2R in mediating osteoblast response to FSS.
• To determine if P2Y2R activation affects cell stiffness and mechanosensitivity.
• To elucidate the signaling pathway involved in FSS-induced ASFF and its consequences.

Main Methods:

• Utilized small interfering RNA (siRNA) to suppress P2Y2R expression.
• Measured RhoA GTPase activation, ASFF, and cell stiffness.
• Assessed intracellular calcium ([Ca(2+)]i) response to FSS to determine mechanosensitivity.

Main Results:

• siRNA suppression of P2Y2R attenuated ASFF induced by FSS and ATP.
• RhoA GTPase activation occurred rapidly (within 15 min) upon FSS or ATP onset, mediating ASFF via the ROCK1/LIMK2/cofilin pathway.
• ASFF increased osteoblast cell stiffness, an effect prevented by P2Y2R knockdown.
• Enhanced cell stiffness and ASFF reduced osteoblast mechanosensitivity to subsequent FSS bouts.

Conclusions:

• Osteoblasts regulate their mechanosensitivity to sustained mechanical load.
• P2Y2R activation of the RhoA GTPase pathway leads to ASFF and increased cell stiffness.
• This mechanism allows osteoblasts to modulate their response to continued mechanical stimulation.