Tissue oxygenation in a murine SCC VII tumor after X-ray irradiation as determined by EPR spectroscopy

Abstract

Purpose: The goal of this study was to clarify the dynamics of oxygenation (partial pressure of oxygen, pO(2)) in SCC VII murine tumors in mice after X-ray irradiation.

Materials and methods: Changes in pO(2) in tumors were measured by 1.2-GHz electron paramagnetic resonance (EPR) spectroscopy after they were exposed to various doses of irradiation. The pO(2) in tumors was followed for up to six days after irradiation at doses of 0, 5, 10, 15, and 20 Gy. Paramagnetic crystals were used as an oximetry probe and implanted into normal or tumor tissues in mice for prolonged periods.

Results: The pattern of tumor oxygen after a single dose of radiation with the 5-Gy dose was different from those with other doses (10, 15, and 20 Gy). After 5 Gy, pO(2) increased rapidly (P<0.01, Student's t test) and then returned to the level observed before irradiation by 12h (P<0.01). In contrast, after 10, 15, or 20 Gy, pO(2) increased rapidly by 6h after irradiation, continued to increase until at least 24h (P<0.01), and then gradually decreased.

Conclusions: In tumors that received 5 Gy, post-irradiation increases in pO(2) at 4h after irradiation were detected by EPR oximetry (P<0.01) noninvasively.

Fig. 1

Calibration curve of the L-band EPR line-width of LiNc-BuO versus pO₂. LiNc-BuO crystals were suspended in PBS equilibrated with mixtures of oxygen/nitrogen gases of 21, 10, 5, 1, 0.5, and 0 % (in argon) oxygen. The spectra were measured under the following conditions: frequency, 1.2 GHz; incident microwave power, 1 mW; modulation frequency, 90 KHz; modulation amplitude, 0.02 mT; temperature, 35–38°C.

Fig. 2

In vivo L-band EPR spectra of LiNc-BuO implanted in the muscle tissues of mice. Fine LiNc-BuO crystals (about 10 µg) were implanted into normal muscle tissues on the hind leg of a C3H mouse. The EPR spectra shown were from a mouse on day 3 (upper) and 30 d (bottom) after implantation of the crystals. The experimental conditions were as follows: frequency, 1.2 GHz; incident microwave power, 5 mW; modulation frequency, 90 kHz; modulation amplitude, 0.03 mT; temperature, 35–38°C.

Fig. 3

Monitoring pO₂ in the muscle tissues through the change in the line-width of implanted LiNc-BuO. Mean values of pO₂ recorded repetitively from 4 mice were shown. Error bars show the standard errors of the means (SEM).

Fig. 4

In vivo measurement of pO₂ (●) from tumor tissues of mice with SCC VII tumors as a function of tumor growth (□). LiNc-BuO crystals (about 10 µg) were implanted in SCC VII tumors in mice, and the pO₂ values in tumors were measured in the same animal up to 8 d after implantation of the crystal. Mean values of pO₂ and tumor volume obtained from 6 mice are shown. Error bars show SEM.

Fig. 5

The average changes of pO₂ in SCC VII tumors in mice after a single radiation. Changes in pO₂ in individual SCC VII tumors in 6 mice were measured over 6 days after a single dose of radiation, (a) 5 Gy, (b) 10 Gy, (c) 15 Gy, and (d) 20 Gy. Experimental conditions were the same as in Fig. 3, except for the radiation treatment. Each point indicates the mean, and error bars indicate SEM. *, P < 0.05; **, P < 0.01.