Philip S. Thayer

Spin filter culture: the propagation of mammalian cells in suspension.

A spin filter device has been used for the propagation in vitro of cells of mnouse leukemia L1210 to densities approaching 108 cells per milliliter. By manipulation of flow rates, cells may be exposed to exponentially declining, drug concentrations with half-times of 1.5 hours or less, providing a more realistic parallel to in vivo drug treatment than is obtained by other culture methods.

A CURRENT ASSESSMENT OF THE MUTAGENIC AND TERATOGENIC EFFECTS OF CAFFEINE

AbstractIn recent years there have been an increasing number of studies of the mutagenicity and terato-genicity of caffeine in experimental systems ranging from microorganisms to mammals. In the absence of direct human data, experimental or epidemiological, the estimation of the degree of risk to which the human caffeine-consuming population is exposed must be indirect. In this review, an attempt is made to draw pertinent data from the experimental studies of mutagenesis and teratogenesis, in conjunction with studies of caffeine metabolism and distribution in mammalian organisms including man, and to interrelate these factors in terms of mechanism of action and pharmacokinetics.

A COMPARATIVE STUDY OF THE USE OF MICROORGANISMS IN THE SCREENING OF POTENTIAL ANTITUMOR AGENTS

The selection of compounds of possible interest as antitumor agents presents a major problem to those concerned with the chemotherapy of neoplastic disease in man. The ideal method for the selection of such compounds must not only be simple, rapid, inexpensive, and adaptable to large-scale operation, but should detect the maximum number of antitumor agents with a rate of “false positives” sufficiently low not to invalidate the use of the method as a screening procedure. The members of the Subcommittee on Microbiology, Screening Panel, Cancer Chemotherapy National Service Center (CCNSC), National Cancer Institute, Public Health Service, Bethesda, Md., have been engaged in the comparative study of several microbiological assay systems to determine the extent of the usefulness of such methods for the detection of potential antitumor agents. The application of bacterial, fungal, and protozoan bioassay methods to this problem is not a new approach; however, the majority of earlier studies have been based upon the use of a single bacterial (or other) bioassay system for this purp0se.l It should not be expected that any single in vitro system will detect all compounds exhibiting antitumor activity in any one or more diverse in vivo experimental tumor systems; this requires no elaboration. I n the expectation that any in vitro microbiological screening procedure must be based upon multiple bioassay systems covering a variety of diverse metabolic pathways, these collaborative studies were organized to include sixteen microbial

Exposure of four generations of mice to caffeine in drinking water

A population of CD-1 mice has been bred for four generations with continuous ingestion of caffeine. Three caffeine levels (and a control) were used with intakes averaging 4–5, 12–18 and 25–39 mg/kg/day (25–39 mg/kg/day is equivalent to 19–30 cups of coffee in man). At each level 20 pairs of animals were bred in each generation, the F1 matings being divided between brother-sister matings and out-crosses. F2s and F3s were out-crossed. There were no consistent, dose-related effects of caffeine on fertility, age at sexual maturity, mean litter size, weight of offspring at weaning, sex ratio or fetal abnormalities. A few pairs at the highest caffeine level failed to produce litters, but only in the F1 brother-sister and in the F3 matings. Litters in the caffeine-treated series showed a somewhat higher incidence of underweight animals in the F2 and the F3 generations, but not in a dose-related way. A difference in sex ratios (males slightly predominating) in the caffeine-treated groups compared to the control groups is related to an apparent deficit of male births in the control groups rather than to an effect of caffeine. In this study 3824 caffeine-exposed offspring (352 litters) were examined at birth for birth defects and 1365 of these (147 litters) were reexamined at weaning age. The only defect observed was in one litter (of F2 offspring from out-cross parents receiving the middle dose of caffeine) with a digital abnormality. This low frequency of abnormal offspring is consistent with control group experience. Further litters from the same parental pair continued on caffeine showed no abnormalities. These studies in the mouse, thus support the safety of caffeine in relatively high doses during reproduction.

Cigarette Smoke: Charcoal Filters Reduce Components That Inhibit Growth of Cultured Human Cells

Water-soluble components of total cigarette smoke inhibit cell growth and protein synthesis by the KB line of human cells. The cytotoxic components were in both the gas phase and the particulate phase of smoke. Conventional filters of cellulose acetate reduced cytotoxicity of the particulate phase in proportion to the weight of particles trapped, that is, they did not alter the specific activity of the particulate phase. Appropriately designed filters containing activated charcoal granules selectively reduced cytotoxic components in cigarette smoke which would have appeared in both phases, although the reduction, as anticipated, occurred to a greater extent in the gas phase.

Genetic tests in mice of caffeine alone and in combination with mutagens

The possible mutagenicity of caffeine has been studied in mice by the dominant-lethal method, in three experiments. (I) Male mice were given caffeine in drinking water for 8 weeks at 3.6, 13.4, 49, and 122 mg/kg/day (comparable to human consumption of 2.8–95 cups of coffee per day). Subsequent mating of each of six males from each group to five females per week for 8 weeks showed no significant increase in dominantlethal mutations (embryonic deaths) whether expressed as early deaths per pregnant female or as mutation index. Although males consuming the two higher levels of caffeine produced fewer pregnancies, litter sizes of females giving birth were not reduced. (II) Single ip injections of caffeine (15 mg/kg) were given to groups of male mice prior to, subsequent to, and immediately at the time of receiving X-rays (100 r). Each of five males from each group was mated to five females per week for 7 weeks. Embryonic deaths did not show any enhancing effect of caffeine on the mutagenicity produced by the irradiation. (III) Three groups of male mice ingested caffeine in water for 16 weeks at levels of 0, 4 and 13 mg/kg/day. Subgroups of five from each group were given either: no further treatment, a single dose of triethylene melamine at 0.2 mg/kg, or 100 r of X-ray, and mated for 7 weeks as above. Fertility and litter size were not affected by the caffeine pretreatment, nor did it modify the induction of dominant-lethal mutations by triethylene melamine or X-rays. Litter sizes showed no significant preimplantation losses in any experiment. Thus, under the conditions described herein and at the doses employed (higher than human exposure), there was no evidence for the mutagenicity of caffeine or the inhibition of DNA repair mechanisms in these mammalian systems.

Environmental and Human Health Aspects of Commercially Important Surfactants

Seven types of surfactants comprise the majority of those presently used in commercial detergent formulations. These are linear alkylbenzene sulfonates (LAS), alkyl sulfates (AS), alcohol ethoxylates (AE), alkyl phenol ethoxylates (APE), alcohol ethoxy sulfates (AES), alpha olefin sulfonates (AOS) and secondary alkane sulfonates (SAS). LAS surfactants, the mainstay of detergent components, have been in use the longest and their paths of biodegradation are relatively well understood. Environmental levels of methylene blue active substances, the most commonly employed but non-specific measure of anionic surfactant concentration, indicate that LAS are readily biodegradable. Nonionic (AE and APE) and the anionic AES and AS surfactants are also biodegradable with APE degrading somewhat more slowly than the others. Acute toxic effects to aquatic life forms generally occur in adult vertebrates and invertebrates at surfactant concentrations from 1 to 20 mg/L; juvenile and developmental stages show effects at somewhat lower concentrations.

Integrated Chemical and Biological Analysis of Asphalt and Pitch Fumes

Significant increased risks in developing cancer of the lung, upper respiratory tract, and upper gastrointestinal tract, including stomach cancer, have been demonstrated for individuals working 20 or more years in roofing operations (Hammond et al., 1976) . These investigators have also documented trends of increased risk of prostate, bladder, or skin cancer, and leukemia in these workers. The occupational exposures incurred by roofers may be associated with high levels of polynuclear aromatic hydrocarbons (PAHs) arising from the heating and application of petroleum asphalts and coal tar pitch. These findings are consistent with results of other studies that have indicated that excessive occupational exposure to PAHs may be associated with increased mortality from various types of cancer. Examples of these occupations include chimney sweeps (Pott, 1775) and coke oven workers (Lloyd, 1971; Mazumdar et al., 1975; Redmond et al., 1972). Skin cancer mortality may be of borderline significance in many of these occupations only because deaths due to skin cancer are rare. However, skin cancer incidence may be excessive.

COMPARATIVE STUDIES OF DRUG UPTAKE, VIABILITY, AND BIOSYNTHETIC CAPABILITIES OF P388 CELLS TREATED WITH “ACTIVE” OR “INACTIVE” TEREPHTHALANILIDES*

Many substituted terephthalanilides have potent activity against a broad spectrum of transplanted murine leukemias and lymphomas *-9 and against several microorganisms.l~-~~ The interaction between DNA and the terephthalanilides in vitro does not appear to correlate with the therapeutic efficacy of these compounds against mouse leukemia,’4 although the bacteriostatic effect of the terephthalanilides apparently parallels the magnitude of their interaction with DNA and RNA.“ The terephthalanilides, which are cationic at neutral pH, form ionic complexes with many biological components in vitro; e.g. anionic lipids!5.16 protein!7 and nucleic acids.”, 13. 1 4 3 la However, evidence for these ionic complexes in situ has not been obtained. The terephthalanilides are extracted from tissues, tumor cells, nuclei, and mitochondria of tumor cells and E. coli B primarily as ionic drug-lipid complexes.15~ 19-21922 The lipids that are extracted from dog brain and from leukemic cells represent a new class of p h o s p h ~ l i p i d ! ~ ~ ~ ~ ~ ~ The concentration of the extractable drug complexes is generally highest in tissues and tumors that are adversely arfected by terephthalanilide treatment. 19-21, 23-26 Since many terephthalanilides have been synthesized, it has been possible to associate particular structures or combinations thereof with the observed chemotherapy.aq9.11.14 Considering chemotherapeutic activity against L1210 growing in mice, the terephthalanilides may be divided into two broad categories: “active” compounds, which produce an increase in life-span of 125% or more, and “inactive” compounds, which produce less. We have evaluated these structureactivity relationships in terms of drug uptake, viability, and biosynthetic capabilities of P388 leukemia cells treated with structurally related “active” and “ in active” terephthalanilides. A preliminary report of this work has been made by Yesair and colleagues?

The Biologist as Industrial Consultant

BIOLOGISTS have been known to stray from university laboratories into those maintained by private industry. While the character of biological research in the industrial world is relatively familiar, less well-defined is the biologists position in an industrial consultant firm. It may be of interest to biologists in general to learn how their professional colleagues fit into such an organization in terms of their status, contributions, and opportunities for research.