Tea tree oil influence on human keratocytes growth and viability

Tea tree oil (TTO) is used in ophthalmology to maintain healthy eyelid skin and to combat parasitic eyelid infections. Keratocytes belong to the structure of the corneal stoma and enable to maintain corneal homeostasis. TTO that reaches the surface of the eye in too high concentration may disturb the functions of these cells. The aim of the study was to test what concentration of TTO is safe for corneal keratocytes in vitro without causing a toxic effect. A normal human keratocytes (HK) cell line was used in the study. Morphology was visualized by light and fluorescence microscopy, cytometric analysis of the cell cycle and cytometric and spectrophotometric viability evaluation were performed. The level of nitric oxide was tested by Griess spectrophotometric method. TTO concentrations exceeding 0.01% significantly reduced cell viability. The IC50 values were on average 0.057%. Increasing TTO concentrations stimulated HK cells to release NOx. The observed values did not exceed 1 μM. The lowest TTO concentration increased the number of HK cells in the G1 phase of the cell cycle. Increasing TTO concentrations (≥0.1%) increased the number of cells in late apoptosis. TTO at concentrations ranging from 0.1% to 0.5% significantly changed cell morphology. Fluorescence analyzes confirmed that TTO at concentrations ≥0.1% induced apoptotic cell death. TTO exerts strong effect on ocular keratocytes depending on applied concentration. Concentrations exceeding 0.1% have a toxic effect on keratocytes, which die mainly by apoptosis. The ocular surface should be protected from excessive exposure to TTO, which may damage corneal stroma cells.