In rat renal fibroblasts, mycophenolic acid inhibits proliferation and production of the chemokine CCL2, stimulated by tumour necrosis factor-alpha.
OBJECTIVE
Renal fibroblasts play a pivotal role in the development of tubulointerstitial fibrosis, a condition highly predictive of progression towards end-stage renal disease. The present study investigated the anti-mitogenic and anti-inflammatory effects of an inhibitor of inosine monophosphate dehydrogenase, mycophenolic acid (MPA) and the mechanisms underlying its action in normal rat kidney fibroblasts (49F cells).


METHODS
Proliferation of 49F cells was studied by tetrazole 3-(4, 5-dimethylthiazol-2-yl-)-2,5-diphenyltetrazolium bromide (MTT) test, bromodeoxyuridine incorporation and flow cytometry. The cyclins, tumour suppressor genes and phospho-mitogen-activated protein kinases (MAPKs) were semiquantified by immunoblotting. Apoptosis was measured by quantifying the fragmented DNA and the activity of caspase 3. The monocyte chemokine CCL2 was measured by ELISA. The mRNA expression of CCL2 was measured by real-time PCR.


RESULTS
Mycophenolic acid dose-dependently inhibited steady-state proliferation of 49F cells by up-regulation of p21, p27 and p53, in association with a decrease in cyclins D2 and E. Treatment with MPA also triggered apoptosis of 49F cells by activating the caspase 3 cascade. Furthermore, MPA attenuated tumour necrosis factor-alpha-induced CCL2 expression through down-regulation of p38 MAPK, but not that of ERK1/2 or JNK.


CONCLUSIONS
The anti-mitogenic and anti-inflammatory effects of MPA were mediated by up-regulation of cell cycle inhibitors and pro-apoptotic signals, and by suppression of p38 MAPK pathway respectively. This dual effect of MPA may form the rationale for animal or clinical trials for the treatment of fibrotic renal diseases.
Chang
Hong-Wei
HW
Renal Division, Department of Internal Medicine, National Taiwan University Hospital, College of Medicine, National Taiwan University, Taipei, Taiwan.
Wu
Vin-Cent
VC
Wu
Kwan-Dun
KD
Huang
Hong-Yu
HY
Hsieh
Bor-Shen
BS
Chen
Yung-Ming
YM
D000818:Animals; D000894:Anti-Inflammatory Agents, Non-Steroidal / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects; D015153:Blotting, Western; D053148:Caspase 3 / Q000378:metabolism; D018929:Cell Culture Techniques; D002460:Cell Line; D049109:Cell Proliferation / Q000187:drug effects*; D018932:Chemokine CCL2 / Q000037:antagonists & inhibitors* / Q000096:biosynthesis; D053938:DNA Fragmentation / Q000187:drug effects; D004305:Dose-Response Relationship, Drug; D004797:Enzyme-Linked Immunosorbent Assay; D005347:Fibroblasts / Q000187:drug effects* / Q000276:immunology / Q000473:pathology; D005434:Flow Cytometry; D007668:Kidney / Q000187:drug effects* / Q000276:immunology / Q000473:pathology; D009173:Mycophenolic Acid / Q000494:pharmacology*; D051381:Rats; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014409:Tumor Necrosis Factor-alpha / Q000502:physiology*; D048051:p38 Mitogen-Activated Protein Kinases / Q000037:antagonists & inhibitors
D000894:Anti-Inflammatory Agents, Non-Steroidal; C491766:Ccl2 protein, rat; D018932:Chemokine CCL2; D014409:Tumor Necrosis Factor-alpha; D048051:p38 Mitogen-Activated Protein Kinases; D053148:Caspase 3; D009173:Mycophenolic Acid