The effects of x-irradiation and hyperthermia on oxygen uptake into intact cells and isolated mitochondria and on membrane fluidity of intact Chinese hamster ovary and HeLa cells and isolated mitochondria and lysosomes under oxic and hypoxic conditions will be studied using electron spin resonance spectroscopy and related methods. The oxygen uptake will be determined using a spin probe which quantitatively changes its spectrum in response to different concentrations of oxygen in solution. Both the environmental concentration and cellular uptake rates will be determined. New methods will be employed to measure oxygen uptake at concentrations producing maximum radioresistance and to measure O2 diffusion in mitochondrial membranes in addition to uptake across the membrane. The study will attempt to differentiate acutely hypoxic from chronically hypoxic cells based on biochemical parameters such as the rate of O2 uptake. The O2 uptake rates following different periods of hypoxia will be perturbed by hyperthermia and x-rays during the reoxygenation processes. Measurement of membrane fluidity will be based on ESR spin-probe methods which label the membranes of cultured mammalian cells with fatty acid spin probes. Conventional ESR (both 9 and 35 GHz) and saturation-transfer ESR will be used for measuring rotational diffusion of lipids and electron electron double resonance (ELDOR) techniques will be used for measuring lateral diffusion of lipids in membranes of intact cells and of isolated mitochondria and lysosomes. Sensitivity of these parameters to hyperthermia and radiation will be altered by agents known to modify cell survival. The determination of the effects of x-rays and heat on O2 uptake of previously hypoxic cells may be important in determining the mechanism of reoxygenation processes in solid tumors. Damage to membrane structure may provide insight into the mechanism of cell killing. The possible correlation between membrane fluidity and O2 uptake will be investigated in detail.

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
Research Project (R01)
Project #
5R01CA037930-03
Application #
3175890
Study Section
Radiation Study Section (RAD)
Project Start
1984-07-01
Project End
1987-06-30
Budget Start
1986-07-01
Budget End
1987-06-30
Support Year
3
Fiscal Year
1986
Total Cost
Indirect Cost
Name
Medical College of Wisconsin
Department
Type
Schools of Medicine
DUNS #
073134603
City
Milwaukee
State
WI
Country
United States
Zip Code
53226
Ankel, E G; Lai, C S; Hopwood, L E et al. (1987) Cytotoxicity of commonly used nitroxide radical spin probes. Life Sci 40:495-8
Lai, C S; Froncisz, W; Hopwood, L E (1987) An evaluation of paramagnetic broadening agents for spin probe studies of intact mammalian cells. Biophys J 52:625-8
Ankel, E; Felix, C C; Kalyanaraman, B (1986) The use of spin label oximetry in the study of photodynamic inactivation of Chinese hamster ovary cells. Photochem Photobiol 44:741-6
Kapiszewska, M; Hopwood, L E (1986) Changes in bleb formation following hyperthermia treatment of Chinese hamster ovary cells. Radiat Res 105:405-12