Robert S. Olson

Development of a Protocol for Photoradiation Therapy of Malignant Brain Tumors: Part 1: Photosensitization of Normal Brain Tissue with Hematoporphyrin Derivative

The successful application of phototherapy to subcutaneous tumors has suggested that a similar procedure should be developed for treating gliomas. As a result, attempts are being made to determine a set of conditions that would optimize the destruction of tumor cells while minimizing injury to surrounding brain tissue. To initiate this task, we developed a novel assay method to assess the amount of phototoxicity induced in normal brain by light exposure of mice treated with hematoporphyrin derivative (HPD). The application of this procedure demonstrated that a sufficient amount of HPD was retained in brain tissue, even 72 hours after injection, to cause severe cerebral damage in light-treated mice.

Ozone Sensitivity in Aging WI-38 Cells based on Acid Phosphatase Content

Young [16-19 population doubling level (PDL)] and senescing (50-53 PDL) WI-38 cell populations were exposed to 1 ppm ozone for 2 hr and the resultant extracellular and intracellular acid phosphatase concentration was measured. Dose-response curves were also determined for surviving populations of young and old cells after a 1 hr ozone exposure ranging in concentration from 0 to 1.00 ppm. Senescing cells released 8 times more acid phosphatase per million cells than the young cells. Both old and young cells showed a clear dose-response to the 1 hr ozone gradient exposure. However, the older cells demonstrated a consistent 17% average lower survival rate than the young cells. The higher acid hydrolase level in older WI-38 cells is probably related to the lower survival rate observed in the older cells in vitro.

The effect of the laser on cellular respiration

SummaryA frequency of 5300 Å, derived from a frequency-doubled Q-switched neodymium laser was observed to produce progressive injury and death of cells from a culture of newborn rat cerebellum. A subsequent observation that the green laser frequency (but not 6943 Å of the same intensity from a Q-switched ruby laser) could reduce the rate of oxygen consumption of rat brain cell suspensions suggested that the cytochromes may serve as chromophores. This hypothesis was confirmed by a demonstration that cytochromes c+c1 failed to act as hydrogen acceptors following 10 impacts of 1 Mw/cm2 each of the green laser frequency and cytochromes a+a3 showed a similar response when a brain cell suspension was irradiated (200 kw) with frequencies of 6096 and 6013 Å. These data illustrate the principle that laser frequencies which are appropriately matched to the absorption characteristics of target molecules can selectively inhibit specific molecular components in intact cells, if a controlled energy density range is used.

Enzyme inactivation with ultraviolet laser energy (2650 Angstroms).

Inactivation of rat heart lactate dehydrogenase was accomplished by irradiation of the enzyme in solution with a frequency quadrupled neodymium glass laser.

The potential use of laser energy in the management of malaria

Abstract A feasibility study was conducted to illustrate that malarially parasitized erythrocytes from infected ducks and man, when treated with methylene blue and exposed to ruby laser energy, show a wavelength specific photolysis of only the parasitized erythrocytes. On the strength of this preliminary study, continuing investigation is being conducted to evaluate other parameters of laser treatment in the management of malaria in experimental animals. These studies suggest that it is feasible to design a protocol for the management of malaria in human patients.

The effect of laser power at 2650 A on deoxyribonucleic acid

Abstract A study was conducted to determine the effect of laser power at a wavelength of 2650 A on template activity of calf thymus DNA. It was observed that template activity decreased exponentially with increasing U.V. laser exposures and that U.V. laser-treated DNA inhibited the activity of non-irradiated DNA. The optical density of the treated DNA showed less than 7% decrease at 2650 A. Although these results were qualitatively similar to those reported for conventional U.V. effects, there was a suggestion of a lack of reciprocity using the high peak powers of the U.V. laser.

Laser photosensitization and metabolic inhibition of tissue culture cells treated with quinacrine hydrochloride

In a preliminary report (1) it was suggested that nucleolur function could be probed by treatment of tissue culture cells with quinacrine hydrochloride followed by laser microirradiation. Apparently the quinacrine associates selec tively with the nucleolus, thus making this organelle susceptile to selective damage by laser microirradiation. If the laser technique is to be useful for studying nucleolur function, the metabolic effects of the photosensitizing agent (quinacrine) must be elucidated. In addition, since previous studies have suggested shifts ïn absorption characteristics of other amino acridines (2) when bound in vitro, it is important to determine if such shifts occur with quinacrine. This is particularly important in selecting the appropriate wavelengths for microirradiation. In this manuscript we will present data : (a) indicating the quinacrine hydrochloride has only a temporary effect on nucleolur function ; (b) on the in vitro absorption spectrum of quinacrine hydrochloride. Materials and Methods Ce11s utilized were the CMP line of human adenocarcinoma cells (3). For metabolic studies all cells were seeded into Leighton culture tubes, 100,000 cells per tube, and incubated for 48 hours at 37oC in Eagles medium supplemented with lOX fetal calf serum, penicillin (100,000 IU/cc), and streptomycin sulfate (0 .25 mg/1). After 48 hours of growth, the medium was removed and quinacrine 1081

Photoradiation therapy of cutaneous angiosarcomas in mice.

Cutaneous angiosarcoma is a relatively rare but devastating malignant vascular tumor. It has a high incidence of recurrence following conventional therapeutic modalities applied either singly or in combination. The increased vascularity of cutaneous angiosarcomas, facilitating selective uptake and retention of a photosensitizing agent, such as hematoporphyrin derivative (HPD), suggests that these tumors would respond well to photoradiation therapy. To study the feasibility of this treatment modality, transplantable hemangiosarcomas were implanted in B6C3F1 female mice. Within 2.5 to 3.5 hours after intraperitoneal administration of HPD, fluorescence was recorded in the tumor as compared with surrounding normal skin. When these photosensitized tumors were exposed to 70 J/cm2 of laser energy from an argon-pumped dye laser at 630 nm, the tumors showed marked necrosis within 24 hours. In another series, the tumors were initially photosensitized with HPD for 3 hours and then treated with laser energy ranging from 0 to 96 J/cm2. A dual labeling procedure demonstrated a dose-related decrease in DNA synthesis rate in tumors that were exposed to 0 to 30 J/cm2 at 24 hours after treatment. Furthermore, tumor tissue exposed to laser energy in excess of 30 J/cm2 showed no significant cellular DNA synthesis. These data, supported by histologic evidence of tissue destruction, suggest that photoradiation therapy has a great potential as a therapeutic modality for cutaneous angiosarcomas.

MEASUREMENT OF THE CELL MIGRATION INDEX WITH A HENE LASER

Human diploid fibroblasts (WI38) were inoculated into Rose multipurpose culture chambers, using a high population density to develop confluency. After 24 hours, cells from a 1 mm swath were scraped from the center of the chamber. This cleared path was positioned to permit an unobstructed transmission of a 0.9 X 20 mm beam of light from a 1 mW HeNe laser. As cells migrated, at 37 degrees C, into the path of the laser beam the light scatter was recorded, using a photomultiplier tube. Because the amount of light scatter was proportional to the number of cells migrating into the beam, the system measured the migration index of the cells. Slight variations in the design of this device could facilitate data collection during surveys of toxic agents, fertility tests, and delayed hypersensitivity.