Specific Attenuation of Purinergic Signaling during Bortezomib-Induced Peripheral Neuropathy In Vitro

Area of Science:

• Neuroscience
• Toxicology
• Stem Cell Biology

Background:

• Human peripheral neuropathies lack effective models, hindering research.
• Current methods like the PeriTox test use immature neurons and focus on cell death.
• There is a need for models detecting sub-cytotoxic neurotoxic effects.

Purpose of the Study:

• To evaluate mature peripheral neuron cultures for detecting sub-cytotoxic neurotoxicity.
• To identify sensitive functional endpoints for early neurotoxicity detection.
• To establish a model for profiling proteasome inhibitors' neuropathy potential.

Main Methods:

• Generated homogeneous sensory neuron cultures from induced pluripotent stem cells (iPSCs) using a two-step differentiation protocol with neurogenin-1 (NGN1).
• Assessed calcium (Ca2+) signaling responses to P2X3 receptor activation using α,β-methylene ATP.
• Treated neurons with low nanomolar concentrations of proteasome inhibitors (e.g., bortezomib) and analyzed changes in Ca2+ signaling and tubulin organization.

Main Results:

• Mature sensory neurons exhibited robust P2X3 receptor-mediated Ca2+ signaling.
• Low-dose bortezomib specifically attenuated P2X3 signaling, indicating functional cell stress without overt cell death or neurite damage.
• Tubulin reorganization confirmed mild, non-cytotoxic stress responses to various proteasome inhibitors.
• P2X3 signaling was a more sensitive endpoint than cell death or neurite damage detection.

Conclusions:

• Mature iPSC-derived sensory neurons provide a sensitive model for detecting sub-cytotoxic neurotoxicity.
• Altered P2X3 receptor signaling serves as a reliable functional endpoint for assessing neurotoxic potential.
• This model aids in the early profiling of drug-induced peripheral neuropathy, particularly for proteasome inhibitors.