TY - JOUR
T1 - A 3D rotary renal and mesenchymal stem cell culture model unveils cell death mechanisms induced by matrix deficiency and low shear stress
AU - Shimony, Nilly
AU - Avrahami, Idit
AU - Gorodetsky, Raphael
AU - Elkin, Gregory
AU - Tzukert, Keren
AU - Zangi, Lior
AU - Levdansky, Lilia
AU - Krasny, Lina
AU - Haviv, Yosef S.
N1 - Funding Information:
Acknowledgements. This study was supported by the German-Israeli Foundation (grant no. 817/2004), Israeli Science Foundation (grant no. 573/2003) and the Hadassah Medical Organization Women’s Health Fund.
PY - 2008/6
Y1 - 2008/6
N2 - Background. In epithelial and endothelial cells, detachment from the matrix results in anoikis, a form of apoptosis, whereas stromal and cancer cells are often anchorage independent. The classical anoikis model is based on static 3D epithelial cell culture conditions (STCK). Methods. We characterized a new model of renal, stromal and mesenchymal stem cell (MSC) matrix deprivation, based on slow rotation cell culture conditions (ROCK). This model induces anoikis using a low shear stress, laminar flow. The mechanism of cell death was determined via FACS (fluorescence-activated cell sorting) analysis for annexin V and propidium iodide uptake and via DNA laddering. Results. While only renal epithelial cells progressively died in STCK, the ROCK model could induce apoptosis in stromal and transformed cells; cell survival decreased in ROCK versus STCK to 40%, 52%, 62% and 7% in human fibroblast, rat MSC, renal cell carcinoma (RCC) and human melanoma cell lines, respectively. Furthermore, while ROCK induced primarily apoptosis in renal epithelial cells, necrosis was more prevalent in transformed and cancer cells [necrosis/apoptosis ratio of 72.7% in CaKi-1 RCC cells versus 4.3% in MDCK (Madin-Darby canine kidney) cells]. The ROCK-mediated shift to necrosis in RCC cells was further accentuated 3.4-fold by H2O 2-mediated oxidative stress while in adherent HK-2 renal epithelial cells, oxidative stress enhanced apoptosis. ROCK conditions could also unveil a similar pattern in the LZ100 rat MSC line where in ROCK 44% less apoptosis was observed versus STCK and 45% less apoptosis versus monolayer conditions. Apoptosis in response to oxidative stress was also attenuated in the rat MSC line in ROCK, thereby highlighting rat MSC transformation. Conclusions. The ROCK matrix-deficiency cell culture model may provide a valuable insight into the mechanism of renal and MSC cell death in response to matrix deprivation.
AB - Background. In epithelial and endothelial cells, detachment from the matrix results in anoikis, a form of apoptosis, whereas stromal and cancer cells are often anchorage independent. The classical anoikis model is based on static 3D epithelial cell culture conditions (STCK). Methods. We characterized a new model of renal, stromal and mesenchymal stem cell (MSC) matrix deprivation, based on slow rotation cell culture conditions (ROCK). This model induces anoikis using a low shear stress, laminar flow. The mechanism of cell death was determined via FACS (fluorescence-activated cell sorting) analysis for annexin V and propidium iodide uptake and via DNA laddering. Results. While only renal epithelial cells progressively died in STCK, the ROCK model could induce apoptosis in stromal and transformed cells; cell survival decreased in ROCK versus STCK to 40%, 52%, 62% and 7% in human fibroblast, rat MSC, renal cell carcinoma (RCC) and human melanoma cell lines, respectively. Furthermore, while ROCK induced primarily apoptosis in renal epithelial cells, necrosis was more prevalent in transformed and cancer cells [necrosis/apoptosis ratio of 72.7% in CaKi-1 RCC cells versus 4.3% in MDCK (Madin-Darby canine kidney) cells]. The ROCK-mediated shift to necrosis in RCC cells was further accentuated 3.4-fold by H2O 2-mediated oxidative stress while in adherent HK-2 renal epithelial cells, oxidative stress enhanced apoptosis. ROCK conditions could also unveil a similar pattern in the LZ100 rat MSC line where in ROCK 44% less apoptosis was observed versus STCK and 45% less apoptosis versus monolayer conditions. Apoptosis in response to oxidative stress was also attenuated in the rat MSC line in ROCK, thereby highlighting rat MSC transformation. Conclusions. The ROCK matrix-deficiency cell culture model may provide a valuable insight into the mechanism of renal and MSC cell death in response to matrix deprivation.
KW - Anoikis
KW - Mesenchymal stem cells
KW - Necrosis
KW - Renal cancer
KW - Shear stress
UR - http://www.scopus.com/inward/record.url?scp=44449169546&partnerID=8YFLogxK
U2 - 10.1093/ndt/gfn062
DO - 10.1093/ndt/gfn062
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C2 - 18344240
AN - SCOPUS:44449169546
SN - 0931-0509
VL - 23
SP - 2071
EP - 2080
JO - Nephrology Dialysis Transplantation
JF - Nephrology Dialysis Transplantation
IS - 6
ER -