Related Experiment Video
Updated: May 31, 2026
Identification of Intracellular Signaling Events Induced in Viable Cells by Interaction with Neighboring Cells Undergoing Apoptotic Cell Death
Published on: December 27, 2016
Phosphate-induced apoptosis in human peritoneal mesothelial cells in vitro
Jong-Won Park1, Ju-Min Yook, Hye-Myung Ryu
1Department of Internal Medicine, Yeungnam University College of Medicine, Daegu, Korea.
Background:
It has been demonstrated that phosphate uptake through the type III sodium-dependent phosphate co-transporter, Pit-1, induced apoptosis of aortic vascular smooth muscle cells and endothelial cells in vitro. However, the apoptotic effects of high phosphate (HP) level in human peritoneal mesothelial cells (HPMCs) are not known.
Methods:
To examine whether Pit-1 is expressed in HPMCs, we checked the Western blot assay of immunoreactive Pit-1 and the transcription of Pit-1 by reverse transcriptase PCR. We treated several different phosphate concentrations (1-4 mM) and calcium concentrations (1.8 and 2.8 mM) on HPMCs to assess the effects of concentration. MTT, TUNEL assays, and flow cytometry analysis using Annexin V and propidium iodide were performed to identify cell death and apoptosis. Bax and Bcl-2 by Western blot and caspase-3 activity were evaluated by colorimetric assay. In addition, phosphonoformic acid (PFA) and pan-caspase inhibitor, Z-VAD-FMK, were given to prevent phosphate-induced apoptosis.
Results:
Pit-1 expression on HPMCs was demonstrated. Apoptosis in HPMCs significantly increased with a high concentration of phosphate in a dose- and time-dependent manner, and was enhanced in the presence of 2.8 mM calcium. HP concentrations significantly decreased the anti-apoptotic Bcl-2/Bax ratio and increased caspase-3 activity. The treatment with PFA and Z-VAD-FMK prevented cell death by HP.
Conclusion:
Phosphate uptake through Pit-1 induces apoptosis in HPMCs by a caspase-related mechanism.
