High glucose activates the p38 MAPK pathway in cultured human peritoneal mesothelial cells.
BACKGROUND
Peritoneal fibrosis is a serious complication in long-term continuous ambulatory peritoneal dialysis (CAPD) patients, but the underlying mechanism is not well understood. Since high glucose activates the p38 mitogen-activated protein kinase (MAPK) pathway in various kinds of cells, and because mesothelial cells are always exposed to high glucose dialysate, we examined the activity and expression of p38 MAPK members in cultured human peritoneal mesothelial cells (HPMCs) under high glucose conditions.


METHODS
HPMCs were isolated from omentum and subcultured. After serum restriction, HPMCs were exposed to 5.6 mmol/L glucose (low glucose), 5.6 mmol/L glucose + 34.5 mmol/L mannitol (low glucose + mannitol), or 40 mmol/L glucose (high glucose) for 3 minutes to 48 hours with or without SB203580. Reverse transcription-polymerase chain reaction (RT-PCR) and Western blot were performed to determine mRNA and protein expression, respectively.


RESULTS
p38 MAPK and cyclic adenosine monophosphate (cAMP)-responsive element binding protein (CREB) activities and mRNA expressions were significantly increased in HPMCs exposed to high glucose compared to low glucose or low glucose + mannitol after 10 minutes and remain at higher levels to 48 hours (P < 0.05), but total p38 MAPK and CREB protein expressions did not differ. MAPK kinase 3/6 (MKK3/6) activity and mRNA expression were also higher in high glucose cells after 3 minutes (P < 0.05), and fibronectin mRNA expression was significantly increased in HPMCs exposed to high glucose after 2 hours (P < 0.05). In contrast, high glucose significantly inhibited MAPK phosphatase-1 (MKP-1) protein and mRNA expression after 10 minutes (P < 0.05). SB203580 (1 micromol/L) pretreatment for 1 hour significantly reduced high glucose-induced CREB activity and fibronectin mRNA expression by 89% and 75%, respectively (P < 0.05).


CONCLUSIONS
p38 MAPK activity was increased in HPMCs exposed to high glucose, in parallel with increased MKK3/6 activity and decreased MKP-1 expression, resulting in CREB activation. This activated p38 MAPK pathway may play a role in the pathogenesis of peritoneal fibrosis.
Xu
Zhong-Gao
ZG
Departments of Internal Medicine and Surgery, College of Medicine, Institute of Kidney Disease, Yonsei University, Seoul, Korea.
Kim
Kyung Sik
KS
Park
Hyeong Cheon
HC
Choi
Kyu Hun
KH
Lee
Ho Yung
HY
Han
Dae Suk
DS
Kang
Shin-Wook
SW
D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000235:genetics / Q000378:metabolism; D018797:Cell Cycle Proteins; D017362:Cyclic AMP Response Element-Binding Protein / Q000235:genetics / Q000378:metabolism; D054638:Dual Specificity Phosphatase 1; D004790:Enzyme Induction / Q000187:drug effects; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004847:Epithelial Cells / Q000187:drug effects / Q000201:enzymology*; D005353:Fibronectins / Q000096:biosynthesis; D005947:Glucose / Q000494:pharmacology*; D006801:Humans; D007093:Imidazoles / Q000494:pharmacology; D017874:Immediate-Early Proteins / Q000235:genetics / Q000378:metabolism; D048371:MAP Kinase Kinase 3; D048669:MAP Kinase Kinase 6; D020935:MAP Kinase Signaling System / Q000187:drug effects; D020929:Mitogen-Activated Protein Kinase Kinases / Q000235:genetics / Q000378:metabolism; D020928:Mitogen-Activated Protein Kinases / Q000235:genetics / Q000378:metabolism*; D010537:Peritoneum / Q000166:cytology* / Q000201:enzymology; D010749:Phosphoprotein Phosphatases; D054645:Protein Phosphatase 1; D017027:Protein Tyrosine Phosphatases / Q000235:genetics / Q000378:metabolism; D011505:Protein-Tyrosine Kinases / Q000235:genetics / Q000378:metabolism; D011725:Pyridines / Q000494:pharmacology; D012333:RNA, Messenger / Q000032:analysis; D048051:p38 Mitogen-Activated Protein Kinases
D018797:Cell Cycle Proteins; D017362:Cyclic AMP Response Element-Binding Protein; D004791:Enzyme Inhibitors; D005353:Fibronectins; D007093:Imidazoles; D017874:Immediate-Early Proteins; D011725:Pyridines; D012333:RNA, Messenger; D011505:Protein-Tyrosine Kinases; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D020928:Mitogen-Activated Protein Kinases; D048051:p38 Mitogen-Activated Protein Kinases; D048371:MAP Kinase Kinase 3; D048669:MAP Kinase Kinase 6; C482530:MAP2K3 protein, human; C482566:MAP2K6 protein, human; D020929:Mitogen-Activated Protein Kinase Kinases; D010749:Phosphoprotein Phosphatases; D054645:Protein Phosphatase 1; C517019:DUSP1 protein, human; D054638:Dual Specificity Phosphatase 1; D017027:Protein Tyrosine Phosphatases; D005947:Glucose; C093642:SB 203580