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Purpose: To investigate the effect of Losartan on articular chondrocyte hypertrophy in a monolayer culture under chemically induced hypoxia with CoCl2.
Materials & Methods: The 2nd passage of human knee chondrocytes was used to determine the cell viability in response to the treatment with Losartan or CoCl2 in the following concentrations 10-4 – 10-12 M. Chondrocytes were seeded in a 96-well plate at the density of 5×103 cells/well. Cell viability was measured after 10 days using a 0,4% Trypan blue stain. After cell viability testing to each reagent (Losartan, CoCl2), the articular chondrocytes were plated in 96-well plate at analogous density and treated every 3rd day with 10-4, 10-6, or 10-9 M concentrations of CoCl2 in the presence or absence of 10-7 M of Losartan. The control wells were treated with a cell culture medium only. Quantity of the following proteins - Collagen X (Col X) and Hypoxia-inducible factor 1 alpha (HIF-1α), were analyzed in cell lysate after 10 days using enzyme-linked immunosorbent assay (ELISA). Differences between independent variables were analyzed by Kruskal-Wallis and Mann-Whitney U-test, while Wilcoxon’s signed-rank test was used for related variables. Data were considered statistically significant when p < 0,05.
Results: Chondrocyte viability increased in response to the treatment with CoCl2 or Losartan at lower concentrations. When cells were treated only with CoCl2, HIF-1α expression was higher compared with treatment in combination with Losartan at the same concentration of CoCl2. Col X expression was elevated when the cells were treated with CoCl2 in combination with Losartan (LC10-9 M). However, Losartan acts as a Col X inhibitor under hypoxic conditions when higher CoCl2 concentrations (LC10-4, LC10-6 M) were used.
Conclusion: Losartan reduced hypoxia in all used CoCl2 concentrations and suppressed hypertrophy of articular chondrocytes at 10-4 and 10-6 M of CoCl2 concentration.
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