Cyclosporine induces different responses in human epithelial, endothelial and fibroblast cell cultures.
BACKGROUND
Nephrotoxicity, accelerated atherosclerosis, and graft vascular disease are common complications of cyclosporine long-term treatment characterized by a wide disruption of organ architecture with increased interstitial areas and accumulation of extracellular matrix (ECM). How cyclosporine induces these changes is not clear, but it is conceivable that they are the sum of changes induced at the cell level.


METHODS
We studied the effects of cyclosporine on human endothelial (HEC), epithelial (HK-2), and fibroblast (MRC5) cells. Cell proliferation was evaluated by cell counting, apoptosis and collagen production by enzyme-linked immunosorbent assay, and nitric oxide by measuring the concentration of nitrite/nitrate in the cell supernatant. (alpha1)I and (alpha2)IV collagen, matrix metalloprotease-9 (MMP9), and tissue inhibitors of metalloprotease-1 (TIMP-1) mRNA levels were measured by reverse transcription-polymerase chain reaction. Proteolytic activity was evaluated by zymography.


RESULTS
Cyclosporine showed a marked antiproliferative and proapoptotic effect on endothelial and epithelial cells. Fibroblast growth was not affected by cyclosporine. Nitric oxide was up-regulated by cyclosporine in epithelial cells and fibroblasts but not in endothelial cells. (alpha1)I and (alpha2)IV collagen synthesis was increased in cyclosporine-treated endothelial and epithelial cells, respectively. Proteolytic activity was increased in endothelial and epithelial cells. TIMP-1 mRNA was up-regulated by cyclosporine in fibroblasts.


CONCLUSIONS
Our results demonstrate that cyclosporine exhibits an antiproliferative effect on endothelial and epithelial cells. This effect is associated with induction of apoptosis probably via nitric oxide up-regulation in epithelial cell cultures. Cyclosporine treatment induces ECM accumulation by increasing collagen synthesis in endothelial and epithelial cells and reducing its degradation by up-regulating TIMP-1 expression in fibroblasts. We conclude that cyclosporine affects cell types differently and that the disruption of organ architecture is the result of multiple effects at the cell level.
Esposito
C
C
Division of Nephrology and Dialysis, IRCCS Policlinico San Matteo, University of Pavia, Italy. ciroe@pop.systemy.it
Fornoni
A
A
Cornacchia
F
F
Bellotti
N
N
Fasoli
G
G
Foschi
A
A
Mazzucchelli
I
I
Mazzullo
T
T
Semeraro
L
L
Dal Canton
A
A
D000199:Actins / Q000235:genetics / Q000378:metabolism; D002196:Capillaries / Q000166:cytology / Q000378:metabolism; D002455:Cell Division / Q000187:drug effects; D002478:Cells, Cultured; D003094:Collagen / Q000235:genetics / Q000378:metabolism; D016572:Cyclosporine / Q000494:pharmacology*; D004730:Endothelium, Vascular / Q000166:cytology* / Q000378:metabolism; D004847:Epithelial Cells / Q000166:cytology / Q000187:drug effects* / Q000201:enzymology; D016326:Extracellular Matrix Proteins / Q000378:metabolism; D005347:Fibroblasts / Q000166:cytology / Q000201:enzymology; D015971:Gene Expression Regulation, Enzymologic / Q000187:drug effects; D006801:Humans; D007166:Immunosuppressive Agents / Q000494:pharmacology*; D016030:Kidney Transplantation / Q000276:immunology; D007684:Kidney Tubules / Q000166:cytology* / Q000378:metabolism; D020780:Matrix Metalloproteinase 9 / Q000235:genetics / Q000378:metabolism; D009569:Nitric Oxide / Q000096:biosynthesis; D012333:RNA, Messenger / Q000032:analysis; D012867:Skin / Q000166:cytology* / Q000378:metabolism; D019715:Tissue Inhibitor of Metalloproteinase-1 / Q000235:genetics / Q000378:metabolism
D000199:Actins; D016326:Extracellular Matrix Proteins; D007166:Immunosuppressive Agents; D012333:RNA, Messenger; D019715:Tissue Inhibitor of Metalloproteinase-1; D009569:Nitric Oxide; D016572:Cyclosporine; D003094:Collagen; D020780:Matrix Metalloproteinase 9