Cell Biology – Immunology – Pathology
Kidney International (2002) 61, 525–533; doi:10.1046/j.1523-1755.2002.00163.x
Effects of oxalate on the re-initiation of DNA synthesis in LLC-PK1 cells do not involve p42/44 MAP kinase activation
Sweaty Koul, Lakshmi S Chaturvedi, Avtar Sekhon, Akshay Bhandari, Mani Menon and Hari K Koul
Biochemistry and Molecular Biology Laboratory, Vattikuti Urology Institute, Henry Ford Health Sciences Center, Detroit, Michigan, USA
Correspondence: Hari K. Koul, M.Sc., Ph.D., Biochemistry and Molecular Biology Laboratory, Henry Ford Health Sciences Center, One Ford Place, Suite 2D/33, Detroit, Michigan 48202, USA E-mail: hkoul1@hfhs.org
Received 19 June 2001; Revised 25 September 2001; Accepted 27 September 2001.
Abstract
Effects of oxalate on the re-initiation of DNA synthesis in LLC-PK1 cells do not involve p42/44 MAP kinase activation.
Background
Oxalate interaction with renal epithelial cells results in a program of events that include alterations in gene expression, re-initiation of DNA synthesis, cell growth and apoptosis. Our studies focused on understanding the mechanisms involved in the oxalate-induced re-initiation of the DNA synthesis. The effects of oxalate alone or in combination with epidermal growth factor (EGF), platelet-derived growth factor (PDGF) and insulin were investigated to determine whether oxalate utilized the p42/44 mitogen activated protein (MAP) kinase pathway, which is a common pathway used by a majority of the mitogens.
Methods
LLC-PK1 cells (a renal epithelial cell line of porcine origin) were exposed to oxalate in the presence or absence of three established growth factors, EGF, insulin and PDGF, and of the transcription/translation inhibitors, actinomycin-D and cycloheximide. DNA synthesis was assessed by [3H]-thymidine incorporation. p42/44 MAP kinase activity was assessed by super-shift analysis as well as by immunocomplex kinase assay.
Results
Exposure of growth-arrested LLC-PK1 cells to oxalate resulted in the re-initiation of the DNA synthesis that had been abolished earlier by pretreatment with transcription/translation inhibitors. Oxalate (1 mmol/L), EGF (50 ng/mL) and insulin (100 ng/mL) stimulated DNA synthesis in growth-arrested LLC-PK1 cells, while PDGF (50 ng/mL) had no effect. Effects of EGF and oxalate on DNA synthesis were additive. In contrast, oxalate and insulin had antagonistic effects on DNA synthesis. Additionally, oxalate did not activate the p42/44 MAP kinase pathway while EGF stimulated this pathway.
Conclusions
These findings demonstrate that oxalate does not activate the p42/44 MAP kinase pathway, and the effects of oxalate are mediated by pathways that are distinct from those of EGF, PDGF and insulin.
Keywords:
insulin, EGF, kidney stones, nephrolithiasis, cell proliferation, apoptosis, hyperoxaluria, mitogen pathway, calcium oxalate stones
