MITOCHONDRIAL INJURY PRODUCED BY JANUS GREEN B

In order to see the influence of mitochondrial damages on the cell, timelapsed morphological changes in HeLa cells after staining supravitally with various concentrations of Janus Green B were observed. For light microscopy, NADH dehydrogenase activity employing tetranitroblue tetrazolium was used for the mitochondrial marker and acid phosphatase activity by azo-dye method for the lysosomal marker. Individual specimens were stained with both methods successively. Ultrathin sections prepared from the monolayer cells on cover-slips were examined by an electron-microscope with or without prior histochemical reaction for acid phosphatase.At concentrations of Janus Green B lower than 2.5×10-6M, no changes were observed in any organelles but mitochondria. And the concentration of Janus Green B around 1×10-6M was confirmed to be appropriate for the selective injury of the mitochondria with combination of Janus Green B and ruby laser light, results of which had been reported previously. However, at least from the morphological view points, Janus Green B itself was found to be selectively injurious to mitochondria at lower concentrations. At the concentration around 2.0×10-5M, which was used conventionally for observation of mitochondria, changes in mitochondria were observed immediately after staining for 8 to 15 minutes. With 10-3 M Janus Green B, cells were fixed on the glass surface and the enzyme activities were entirely lost. Cellular degenerative process produced by Janus Green B was discussed, particularly placing emphasis on the mitochondrial changes and on the role of lysosome.