The effects of cysplatin on human adipose tissue derived mesenchymal stromal cells under different oxygen levels

Objective: To evaluate the damaging effects of cisplatin on MMSCs from adipose tissue in a phase of active proliferation and the state of the monolayer, which was exposed at standard (20%) and reduced to 1% and 5% level of oxygen.

Methods: The incubation MMSC with cisplatin was performed on cultures of 2 passage in a state in monolayer and cultures in the active growth phase. Profile surface markers of MMSC determined by flow cytometry. MMSCs viability after incubation with cisplatin was detected by the number of apoptotic and necrotic cells using ANNEXIN V-FITC - PI Kit (Immunotech, France). Standard culture conditions (~ 20% O₂) created in a CO₂ incubator (Sanyo, Japan), 5% O₂ created using multigas incubator (Sanyo, Japan), 1% O₂ - using an airtight chamber (Stemcell Technologies, USA).

Results: Incubation of monolayer MMSC with cisplatin at a concentration of 10 ug/ml for 72 hours leads to death of half of the cells in culture under 20% O₂, 5% O₂ and 1% O₂. Cisplatin increased the fracture of PI⁺-cell, and PI⁺/Ann⁺-cells under all culture conditions. The short-term exposure with cisplatin (24 and 48 hours) did not cause the damaging effect. Effects of cisplatin on the MMSC in the growth phase for 48 hours led to accumulation of Ann⁺-cells and PI⁺/Ann ⁺-cells under all culture conditions. However least damaging effect of cisplatin was observed in culture under hypoxic conditions (1% O₂).

Conclusion: These data suggest that monolayer MMSCs are dying primarily through necrosis, whereas MMSC in the growth phase in response to cisplatin treatment are dying by apoptosis, regardless the oxygen tension.