Ozonation of mutagenic and carcinogenic polyaromatic amines and polyaromatic hydrocarbons in water.

The Salmonella-microsome assay for mutagenesis was used to determine the effect of ozone on the mutagenesis of selected carcinogens and mutagens in water. Short periods of ozonation were shown to completely inactivate the mutagenicity of several polyaromatic amine mutagens including acriflavine, proflavine, and beta-naphthylamine. Selected polyaromatic hydrocarbons were also sensitive to ozonation. Kinetic studies revealed that the mutagenicity of benzo(a)pyrene, 3-methylcholanthrene, and 7,12-dimethylbenz(a)anthracene was destroyed after short periods of ozonation. To correlate loss of mutagenicity with loss of carcinogenicity, two polyaromatic hydrocarbons were treated with ozone, extracted from water with hexane, and tested for carcinogenicity in mice. When 7,12-dimethyl-benz(a)anthracene and 3-methyl-cholanthrene were treated with ozone, there was a substantial reduction in carcinogenicity compared to control groups treated with oxygen alone. However, a small number of tumors developed in the group of animals receiving a hexane extract of ozonated 7,12-dimethylbenz(a)anthracene. This activity may be due to breakdown products of 7,12-dimethylbenz(a)anthracene that are not mutagenic.

Ozonation of mutagenic and carcinogenic alkylating agents, pesticides, aflatoxin B1, and benzidine in water.

The effect of ozonation on the mutagenicity of selected chemicals in water was determined. The use of the Salmonella-microsome assay for mutagensis allowed kinetic studies to be performed on the ozonation of all chemicals tested. The results indicate that the mutagenicity of certain pesticides, including captan and Dexon, was inactivated by short periods of ozonation. The mutagenicity of certain alkylating agents including bis(2-chloroethyl)amine and sodium azide was rapidly inactivated by ozonation while other alkylating agents such as beta-propiolactone, propanesultone, and N-methyl-N'-nitro-N-nitrosoguanidine were unaffected by treatment with ozone. The mutagenicity of aflatoxin B1 was rapidly inactivated by treatment with ozone. Three chemicals were shown to be converted to direct mutagens by ozone treatment. Under certain conditions, dimethylhydrazine could be converted to a mutagen that was stable for 3 weeks. A similar chemical, 2-hydroxyethylhydrazine, was converted to an unstable mutagen that was inactive after 24 hr at room temperature. When benzidine was treated with ozone, there was a transient increase in mutagenicity which was lost after longer treatment with ozone.

Selective immunosuppression in mice of natural killer cell activity by ochratoxin A.

Ochratoxin A, a naturally occurring mycotoxin, has recently been shown to cause renal and hepatic carcinomas in mice. In the present studies, the effects of ochratoxin A on immune mechanisms associated with tumor resistance were examined in mice using dose levels similar to those that cause neoplasia. Ochratoxin A was shown to specifically inhibit natural killer (NK) cell activity and increase the growth of transplantable tumor cells without altering T-cell- or macrophage-mediated antitumor activity. In contrast, ochratoxin B, a much less toxic ochratoxin, did not influence immune function. Polyinosinic:polycytidylic induced interferon was markedly reduced in mice following exposure to ochratoxin A although total serum protein levels were slightly increased. Injection of polyinosinic:polycytidylic enhanced NK activity in the presence of ochratoxin A, although the level of enhancement was slightly lower than that produced by the agent in the absence of ochratoxin A. Thus, ochratoxin appears to suppress NK cell activity by inhibiting production of basal interferon. Additionally, these findings suggest a possible role for altered NK cell function in the development of mycotoxin-induced carcinogenesis.

The study of rous sarcoma virus-transformed baby hamster kidney cells using fluorescent probes.

The fluorescent probes pyrene, pyrene butyric acid and N-phenyl 1-naphthylamine have been used to investigate the changes that accompany in vitro transformation of a baby hamster kidney cell line using Rous sarcoma virus. The fluorescent probes which reside in the membrane were used to compare the changes in microviscosity and polarity of the membranes of normal cells with two transformed cell lines. The spectrofluorimetric data indicate that following transformation the probe N-phenyl 1-naphthylamine resides in a more polar environment. However, using the probe pyrene, the yield of excimer indicates decreased mobility of this probe in the membrane of transformed cells. The data also indicate differences between the two transformed cell lines. Laser photolysis was used to study the lifetime of the pyrene probes and the quenching of the pyrene fluorescence in the membrane by several different quenching molecules. The data indicate differences between the three cell lines and suggest that transformation decreases movement within the membrane.

Rat leukocyte interferons: production of an acid-labile interferon after induction with Newcastle disease virus.

Rat leukocytes produce three interferon species after infection with Newcastle disease virus. Two of the three interferon species (22-24,000 and 30,000 daltons) were acid and heat stable, protected rat and mouse cells from virus-mediated cytopathic effect (CPE), and were neutralized by antisera to mouse L-cell (alpha/beta) interferon. The third, 30,000-dalton interferon species was heat and acid labile; protected rat, mouse, human, and bovine cells from virus-mediated CPE; and was neutralized by antisera to mouse L-cell interferon and human leukocyte interferon. Thus, the rat leukocyte interferon system is similar to the human system in the kinetics of interferon production, the production of multiple interferons, heterologous antiviral activity, molecular weight, and in the production of an acid-labile species. The rat leukocyte interferon system therefore represents an important model for examining the normal production and action of acid-labile interferons and their possible role in the regulation of leukocyte function and cellular differentiation.

Comparison of three bioassays for rat interferon.

Compared to the well-characterized murine and human interferons, few studies have been conducted on rat interferons. In the present study, we compared three rat interferon bioassays: a plaque-reduction method, a hemagglutination yield-reduction method, and a method involving reduction in viral cytopathic effect. These methods were evaluated in order to better assay rat interferons and thus facilitate investigations to determine the antiviral, antitumor, and immunoregulatory role of interferon in existing rat model systems. Each method has certain desirable characteristics; the choice of bioassay depends on the specific application required. Overall, the reduction in viral cytopathic effect bioassay combines most of the desired features of a rat interferon bioassay including sensitivity, precision, convenience, rapidity, and economy.

Influenza virus host resistance models in mice and rats: utilization for immune function assessment and immunotoxicology.

Each year influenza viruses are responsible for epidemic respiratory diseases with excess morbidity and mortality. The severity of influenza diseases ranges from mild upper respiratory tract infections to severe lower respiratory tract infections involving pneumonia, bronchiolitis and coincidental bacterial super-infections. The immune response to influenza viruses can be schematically divided into a cascade of non-specific and specific functions. These functions are involved at different well defined time points after infection. We describe in this manuscript three influenza models utilized in our laboratory: (i) a highly virulent influenza virus (influenza A/Hong Kong/8/68 (H3N2) virus) adapted to B6C3F1 mice, (ii) a mouse-adapted influenza A/Port Chalmers/1/73 (H3N2) virus, and (iii) a rat-adapted influenza virus (RAIV) model (influenza A/Port Chalmers/1/73 (H3N2)). This rat-adapted influenza model has been mainly utilized as a model to assess local immunotoxic effects of inhaled environmental pollutants such as phosgene. These host resistance models are also useful for assessing the effect of systemically-induced immunosuppression or immunomodulation by drugs or chemicals on the local pulmonary immune response to influenza virus. The comparison of these different models allowed two major conclusions: (a) viral replication and mortality are two different endpoints and are not necessarily linked (no mortality was observed with Port Chalmers virus in the mouse although the virus replicates to high titers in the lung with a kinetic pattern comparable to the one obtained with Hong Kong virus), (b) mortality, viral replication, and immune function assessment are different endpoints that can be used, depending on the question addressed.

Immunotoxicological investigation using pharmaceutical drugs. In vitro evaluation of immune effects using rodent or human immune cells.

In order to evaluate the relevance of in vitro methods for immunotoxicity assessment, the effects of pharmaceutical drugs on lymphoproliferative and cytotoxic functions of mouse splenocytes and human peripheral blood mononuclear cells (hPBMC) were studied. A comparison of sensitivity of immune cells from different origins to an in vitro exposure to different xenobiotics was performed using non-immunosuppressive (cimetidine and furosemide) and immunosuppressive (azathioprine (AZA), cyclosporine A (CSA), and dexamethasone (DEX)) drugs. For CSA, sensitivity of both rat and mouse splenocytes following in vitro exposure was compared to the one of hPBMC. Immune function tests included lymphoproliferative response to mitogenic lectins (concanavalin A (Con A) and phytohemagglutinin (PHA-P)) or to allogeneic cells (mixed leukocyte response (MLR)) and cytotoxicity assays (cytotoxic-T lymphocyte (CTL) and natural killer (NK) cell-mediated cytolysis). Additionally, to evaluate how well in vitro assays represent the in vivo situation, a comparison of the effect of cyclosporine A on the same immune function tests following in vivo or in vitro exposure was performed. The data obtained show numerous similarities in the effects observed following in vitro exposure of rodent or human cells to the drugs and a very similar sensitivity of rat and mouse cells to CSA in vitro. Discrepancies between human and rodent cells such as lymphoproliferative response to PHA-P following exposure to DEX or sensitivity of CTL-mediated cytolysis to CSA do exist. In vitro assays were very representative of the in vivo situation, both in the rat and in the mouse, following CSA exposure, except for NK cell activity in the rat. These data show the usefulness of in vitro systems for immunotoxicity assessment. They allow direct comparison of rodent and human systems, and could be representative, for drugs altering specifically the immune system like CSA does, of the in vivo situation.

pH-sensitive mutagenic activity in ozone-treated 1,2-dimethylhydrazine in the Salmonella/microsome assay.

Treatment with ozone inactivates the mutagenicity of many carcinogens in aqueous solution. The colon carcinogen, 1,2-dimethylhydrazine (DMH) has been reported an exception; ozone treatment converts dimethylhydrazine from a non-mutagen into a mutagen. In the Salmonella/microsome assay, the mutagenicity of ozone-treated dimethylhydrazine was dependent on pH. The ozonation product was a strong mutagen in acidic solution but was not mutagenic in basic solution. The mutagenicity of the acidic ozonation product was inactivated by raising the pH of the solution. Unlike untreated dimethylhydrazine, its ozonation product in basic solution was not converted to a mutagen in this ozone-low pH system.

Medial temporal lobe amnesia: a case study for nursing.

Maintaining and enhancing cognitive function is a crucial but challenging intervention for patients with memory problems. Research on the medial temporal lobe (MTL) memory system has yielded much information that can guide nurses in planning, evaluating, and performing effective interventions. A patient, Mrs. N, with a diagnosis of anaplastic astrocytoma of the left medial temporal lobe provides an example. Information from research guides assessment of Mrs. N and affords development of specific patient-centered interventions to maintain function, cope, and compensate. Data have been gathered from the patient, relatives, and caregivers to compare with and augment existing research, because few nursing case studies of amnesia involving patients with left medial temporal lobe tumors are available for analysis.

Avoiding evaluation cooptation. Lessons from a renal dialysis evaluation.

An evaluation of the impact of terminating contracts to transport low-income patients for renal dialysis demonstrates how evaluators may become apologists for management and conduct studies too narrow for effective decision making. The evaluation found that patients continued to receive treatment; their death rates had not increased. Changes in their economic and emotional status were not studied; consequently, the evaluators erroneously concluded that the clients had not been seriously harmed. The article reviews the process the evaluation used. The authors conclude that to avoid cooptation, evaluators need independent information available through conducting and analyzing pilot studies and meeting with program constituents. To avoid overly narrow studies, the next level of management should participate in their design. To use the information effectively, evaluators need to see themselves as advisors rather than technicians.

Management, budgeting and the use of resources--a private sector view.

This very brief talk can only scratch at the surface of the way in which we approach these matters within AMI. I would remind you that we are very small in comparison with the national health service, and what is effective for us, may not be effective for you. Our philosophy is to give the manager responsibility, along with authority to act, to control the direction of the organisation centrally, but to delegate downwards the detailed day to day decision making, to measure managerial performance by results, and to emphasise above all the quality of the service we provide to our patients. Our various management structures are pyramid-shaped and identify a responsible leader at each level of the organisation. Our budgeting process enables us to set our financial plans, and our financial control process enables us to measure progress against those plans. Finally, I referred to the vitally important part which our people play translating that management philosophy and those goals and objectives into reality and our efforts to equip those people through training programmes to meet the heavy demands which we place upon them. In summary, we select the right person, train him, provide him with the necessary resources, motivate him and leave him alone to do the job. Whether any of this is of relevance to you in the national health service is for you to judge. My suspicion is that if there is relevance, it is to do with attitudes and expectations and demands and performance, rather than simply in structures and systems and organisational charts.

Enhanced and prolonged pulmonary influenza virus infection following phosgene inhalation.

Animal infectivity models have been important in the demonstration of enhanced susceptibility to viral and bacterial infection as a result of low-level toxicant exposure. This study demonstrated an enhanced and prolonged viral infection using an influenza virus infectivity model in the rat following exposure to the toxicant gas phosgene. Fischer-344 rats exposed to either air or a sublethal concentration of phosgene demonstrated peak pulmonary influenza virus titers 1 d after infection. Virus titers in rats exposed to air declined rapidly falling below detectable levels by 4 d after infection. However, a significantly enhanced and prolonged pulmonary influenza virus infection was observed on d 3 and 4 after infection in rats exposed to phosgene. Virus was cleared below detectable limits on d 5 after infection in animals exposed to phosgene. Thus, inhalation of sublethal concentrations of phosgene resulted in an increased severity of pulmonary influenza virus infection. This study provides a demonstration of the effective use of a rat viral infectivity model to detect the immunotoxicity of inhaled pollutants. This model will allow future studies to focus on the immunological mechanism(s) responsible for the enhanced and prolonged pulmonary influenza virus infection.

Production and characterization of poly(I):poly(C)-induced rat interferons in vitro.

The synthetic polyribonucleotide polyinosinate:polycytidylate [poly(I):poly(C)] was used to induce interferon (IFN) production in a rat leiomyosarcoma cell line. Three separate IFN species were distinguished by antiviral activity on homologous and heterologous cells, interaction with Blue Sepharose CL-6B chromatography matrix, and antibody neutralization studies. Two IFN species, one with a weakly ionic and the other with a hydrophobic interaction with Blue Sepharose, exhibited heterologous antiviral activity, and were acid- and heat-labile. Both had a molecular weight of 30,000 daltons by HPLC-based molecular exclusion chromatography. The weakly ionic species was neutralized only by antiserum to L-cell IFN, whereas the hydrophobic species was neutralized by antiserum to L-cell IFN and antiserum to human leukocyte IFN. The remaining IFN species, with a weakly ionic interaction with Blue Sepharose, was acid- and heat-stable, exhibited no heterologous cell antiviral activity, and was not neutralized by any antisera to IFN. This species had a molecular weight of 39,000 daltons by HPLC-based molecular exclusion chromatography and 34,000 daltons by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. These rat fibroblast IFNs possess greater heterogeneity than that reported or observed for murine or human fibroblast IFNs. These IFNs could thus be classified as follows: one species of rat IFN-beta and two different species of unique acid-labile rat IFNs.

Natural killer activity in Fischer-344 rat lungs as a method to assess pulmonary immunocompetence: immunosuppression by phosgene inhalation.

Phosgene, also known as carbonyl chloride, carbon oxychloride, and chloroformyl chloride, is a toxic air pollutant and a potential occupational health hazard. Studies were initiated (a) to evaluate the measurement of pulmonary natural killer (NK) activity as a method to assess pulmonary immunocompetence, and (b) to determine whether exposure to phosgene resulted in local pulmonary or systemic immune dysfunction. Fischer-344 male rats were exposed either to filtered air or to 1.0 ppm phosgene gas for four hours. The effect of phosgene on lung NK activity was quantified at different times after acute phosgene exposure. Pulmonary NK activity was measured by mincing lung tissue into small pieces prior to incubation with collagenase. Whole-lung homogenate was assayed for NK activity utilizing a 4 hour 51-Cr-release assay with YAC-1 cells as target cells. Acute phosgene exposure resulted in a suppressed pulmonary NK activity on days 1, 2, and 4 after exposure; however, normal levels of biological activity were observed 7 days after exposure. The suppressed NK activity was not restored after removal of adherent cells from the lung homogenate, thus indicating that the effect of phosgene on NK activity was not due to immunosuppression via mobilization of suppressor alveolar macrophages. Pulmonary immunotoxicity was also observed after exposure at 0.5 ppm, while no adverse effects were observed at 0.1 ppm phosgene. Systemic immunotoxic effects were observed for NK activity in the spleen, but not in the peripheral blood. It is thus important in pulmonary immunotoxicology to evaluate systemic immune functions, since secondary effects--distant to the original interaction--may occur with potentially serious consequences. Cells exhibiting natural killer activity comprise a part of the nonspecific innate immunity that is important in defense against both neoplastic and viral diseases. Any perturbation of this important nonspecific immunological mechanism may result in a compromised host more susceptible to infectious and neoplastic disease.

Effect of ozonation on the mutagenicity of carcinogens in aqueous solution.

Ozone is a very strong oxidizing agent that may be used in water purification. The effect of ozonation on the mutagenicity of mutagens and/or carcinogens of diverse chemical structures was evaluated as measured by the Salmonella/microsome assay. The effect of ozonation of 36 mutagens and/or carcinogens has been evaluated and the results of 15 reported here. The mutagenicity of polycyclic aromatic hydrocarbons and aromatic amines was inactivated by ozone treatment, while alkylating agents, nitro aromatics, and nitroso compounds were not affected. Ozonation of hydrazines produced mutagenic intermediates that may be susceptible to base-catalyzed hydrolysis. Therefore, depending on the chemical present, ozonation may be useful in the treatment of waters containing organic carcinogens, including drinking water, waste-water effluents, and other aqueous waste materials containing carcinogens.