EFFECT OF METHOTREXATE ON CELL-CYCLE AND DNA-SYNTHESIS OF ASTROCYTES IN PRIMARY CULTURE - FLOW CYTOMETRIC STUDIES

Relatively pure population of astrocytes in primary culture were examined by flow cytometry at various time intervals after exposure to 1, 10 and 100-mu-M of methotrexate (MTX). The percentage population of cells in different phases of the cell cycle was determined using propidium iodide (PI) to measure cellular DNA content. DNA synthesis was assessed by measuring the fluorescence intensity of FITC-conjugated antibody to bromodeoxyuridine (BrdUrd). The two parameters were correlated to determine the location of BrdUrd-incorporating cells within the cell cycle. Exposure of astrocytes to MTX caused a consistent increase in number of cells in S-phase which correlated with the increase in BrdUrd incorporation. These studies provide supportive evidence that the astrogliosis seen in MTX encephalopathy may be due to a primary involvement of astrocytes.