Effects of 42°C hyperthermia on intracellular pH in ovarian carcinoma cells during acute or chronic exposure to low extracellular pH

Abstract Purpose: To determine whether intracellular pH (pH i ) is affected during hyperthermia in substrate-attached cells and whether acute extracellular acidification potentiates the cytotoxicity of hyperthermia via an effect on pH i . Methods and Materials: The pH i was determined in cells attached to extracellular matrix proteins loaded with the flourescent indicator dye BCECF at 37°C and during 42°C hyperthermia at an extracellular pH (pH e ) of 6.7 or 7.3 in cells. Effects on pH i during hyperthermia are compared to effects on clonogenic survival after hyperthermia at pH e 7.3 and 6.7 of cells grown at pH e 7.3, or of cells grown and monitored at pH e 6.7. Results: The results show that pH i values are affected by substrate attachments. Cells attached to extracellular matrix proteins had better signal stability, low dye leakage and evidence of homeostatic regulation of pH i during heating. The net decrease in pH i in cells grown and assayed at pH e = 7.3 during 42°C hyperthermia was 0.28 units and the decrease in low pH adapted cells heated at pH e = 6.7 was 0.14 units. Acute acidification from pH e = 7.3 to pH e = 6.7 at 37°C caused an initial reduction of 0.5-0.8 unit in pH i , but a partial recovery followed during the next 60–90 min. Concurrent 42°C hyperthermia caused the same initial reduction in pH i in acutely acidified cells, but inhibited the partial recovery that occurred during the next 60–90 min at 37°C. After 4 h at 37°C, the net change in pH i in acutely acidified cells was 0.30 pH unit, but at 42°C is 0.63 pH units. The net change in pH i correlated inversely with clonogenic survival. Conclusions: Hyperthermia causes a pH i reduction in cells which was smaller in magnitude by 50% in low pH adapted cells. Hyperthermia inhibited the partial recovery from acute acidification that was observed at 37°C in substrate attached cells, in parallel with a lower subsequent clonogenic survival.