Ozone induces glucose intolerance and systemic metabolic effects in young and aged Brown Norway rats.

Air pollutants have been associated with increased diabetes in humans. We hypothesized that ozone would impair glucose homeostasis by altering insulin signaling and/or endoplasmic reticular (ER) stress in young and aged rats. One, 4, 12, and 24 month old Brown Norway (BN) rats were exposed to air or ozone, 0.25 or 1.0 ppm, 6 h/day for 2 days (acute) or 2 d/week for 13 weeks (subchronic). Additionally, 4 month old rats were exposed to air or 1.0 ppm ozone, 6 h/day for 1 or 2 days (time-course). Glucose tolerance tests (GTT) were performed immediately after exposure. Serum and tissue biomarkers were analyzed 18 h after final ozone for acute and subchronic studies, and immediately after each day of exposure in the time-course study. Age-related glucose intolerance and increases in metabolic biomarkers were apparent at baseline. Acute ozone caused hyperglycemia and glucose intolerance in rats of all ages. Ozone-induced glucose intolerance was reduced in rats exposed for 13 weeks. Acute, but not subchronic ozone increased α2-macroglobulin, adiponectin and osteopontin. Time-course analysis indicated glucose intolerance at days 1 and 2 (2>1), and a recovery 18 h post ozone. Leptin increased day 1 and epinephrine at all times after ozone. Ozone tended to decrease phosphorylated insulin receptor substrate-1 in liver and adipose tissues. ER stress appeared to be the consequence of ozone induced acute metabolic impairment since transcriptional markers of ER stress increased only after 2 days of ozone. In conclusion, acute ozone exposure induces marked systemic metabolic impairments in BN rats of all ages, likely through sympathetic stimulation.

A comparison of the allergic responses induced by Penicillium chrysogenum and house dust mite extracts in a mouse model.

UNLABELLED: A report by the Institute of Medicine suggested that more research is needed to better understand mold effects on allergic disease, particularly asthma development. We compared the ability of the fungal Penicillium chrysogenum (PCE) and house dust mite (HDM) extracts to induce allergic responses in BALB/c mice. The extracts were administered by intratracheal aspiration (IA) at several doses (0, 2.5, 5, 10, 20, 40, and 80 µg) four times over a 4-week period. Three days after the last IA exposure, serum and bronchoalveolar lavage fluid (BALF) were collected. The relative allergenicity of the extracts was evaluated based on the lowest dose able to induce a significant response compared to control (0 µg) and the robustness of the response. PCE induced the most robust response at the lowest dose for most endpoints examined: BALF total, macrophage, neutrophil, and eosinophil cell counts, and antigen-specific IgE. Taken together, our data suggest that PCE may induce a more robust allergic and inflammatory response at lower doses than HDM. PRACTICAL IMPLICATIONS: Our data suggest that Penicillium chrysogenum is a robust allergen and may be a more potent allergen source than house dust mite (HDM) in this mouse model. Two critical factors in the development of human allergic disease, exposure levels and sensitization thresholds, are unknown for most allergens including molds/fungi. Human exposure levels are not within the scope of this article. However, the data presented suggest a threshold dose for the induction of allergic responsiveness to P. chrysogenum. Additionally, P. chrysogenum as well as other molds may play an important role in asthma development in our society.

Residual oil fly ash inhalation in guinea pigs: influence of absorbate and glutathione depletion.

Inhaled urban particulate matter (PM) often contains metals that appear to contribute to its toxicity. These particles first make contact with a thin layer of epithelial lining fluid in the respiratory tract. Antioxidants present in this fluid and in cells might be important susceptibility factors in PM toxicity. We investigated the role of ascorbic acid (C) and glutathione (GSH) as determinants of susceptibility to inhaled residual oil fly ash (ROFA) in guinea pigs (male, Hartley). Guinea pigs were divided into four groups, +C+GSH, +C-GSH, -C+GSH, and -C-GSH, and exposed to clean air or ROFA (< 2.5 micron diameter, 19--25 mg/m(3) nose-only for 2.0 h). C and/or GSH were lowered by either feeding C-depleted diet (1 microg C/kg diet, 2 weeks) and/or by ip injection of a mixture of buthionine-S,R-sulfoximine (2.7 mmol/kg body weight) and diethylmaleate (1.2 mmol/kg, 2 h prior). Nasal lavage (NL) and bronchoalveolar lavage (BAL) fluid and cells were examined at 0 h and 24 h postexposure to ROFA. The C-deficient diet lowered C concentrations in BAL fluid and cells and in NL fluid by 90%, and the GSH-depletion regimen lowered both GSH and C in the BAL fluid and cells by 50%. ROFA deposition was calculated at time 0 from lung Ni levels to be 46 microg/g wet lung. In unexposed animals, the combined deficiency of C and GSH modified the cellular composition of cells recovered in lavage fluid, i.e., the increased number of eosinophils and macrophages in BAL fluid. ROFA inhalation increased lung injury in the -C-GSH group only (evidenced by increased BAL protein, LDH and neutrophils, and decreased BAL macrophages). ROFA exposure decreased C in BAL and NL at 0 h, and increased BAL C and GSH (2- to 4-fold above normal) at 24 h in nondepleted guinea pigs, but had no effect on C and GSH in depleted guinea pigs. Combined deficiency of C and GSH resulted in the highest macrophage and eosinophil counts of any group. GSH depletion was associated with increased BAL protein and LDH, increased numbers of BAL macrophages and eosinophils, and decreased rectal body temperatures. We conclude that combined deficiency of C and GSH increased susceptibility to inhaled ROFA; caused unusual BAL cellular changes; resulted in lower antioxidant concentrations in BAL than were observed with single deficiencies. Antioxidant deficiency may explain increased susceptibility to PM in elderly or diseased populations and may have important implications for extrapolating animal toxicity data to humans.

The effect of a vitamin A acetate diet on ultraviolet radiation-induced immune suppression as measured by contact hypersensitivity in mice.

The adverse health effects caused by increased exposure to ultraviolet radiation (UVR) due to deterioration of stratospheric ozone are of major concern. These health effects include sunburn, skin cancer, cataracts and immune suppression. Immune suppression has been associated with the release of cytokines, a defect in antigen presentation, induction of suppressor T cells and suppression of contact hypersensitivity (CH). CH is typically assessed by the mouse ear swelling test (MEST). Previous studies have demonstrated enhanced CH responses with vitamin A acetate (VAA) dietary supplementation assessed by MEST and the local lymph node assay (LLNA). To determine the effect that VAA has on UVR-induced immune suppression, we examined both the induction and elicitation phases of CH using murine models. The MEST was used to evaluate the interaction of UVR and VAA on CH elicitation. However, a positive MEST response requires that the induction phase as well as the elicitation phase of CH be functional. The LLNA was used to evaluate the interaction of UVR and VAA only on CH induction. We tested the hypothesis that mice maintained on a VAA-enriched diet are more resistant to UVR-induced immune suppression (CH) than those maintained on a control diet. Mice were maintained on a VAA-enriched or the control diet for 3 weeks and then exposed to UVR 3 days prior to sensitization with 2,4-dinitrofluorobenzene (DNFB). VAA enhanced the MEST response in both UVR-exposed and non-UVR-exposed mice. The VAA-enriched diet did not significantly alter the LLNA response in either UVR- or non-UVR-exposed mice. However, there was significant suppression in CH by UVR as measured by the LLNA. These results indicate that (1) the VAA-enriched diet does not restore the number of proliferating cells in the CH induction phase of UVR-induced immunosuppression; (2) the immunosuppressive effects of UVR affect the induction phase of CH; and (3) the LLNA should be examined as an alternative to the MEST for measurement of UVR-induced immunosuppression. The data indicate that the VAA-enriched diet enhanced the elicitation response (MEST) but not the earlier induction phase (LLNA). Further studies are necessary to define mechanisms of action, but modulation of cytokines and effects of specific lymphocyte subsets, as well as systemic effects and local modulation at the site of elicitation are possible. Additionally, future studies to evaluate the effect of the VAA-enriched diet when multiple doses of both UVR and DNFB are used would be of interest for both the LLNA and MEST end-points.

Immune responses in farm workers after exposure to Bacillus thuringiensis pesticides.

Although health risks to pesticides containing Bacillus thuringiensis (Bt) have been minimal, the potential allergenicity of these organisms has not been evaluated. Therefore, a health survey was conducted in farm workers before and after exposure to Bt pesticides. Farm workers who picked vegetables that required Bt pesticide spraying were evaluated before the initial spraying operation (n = 48) and 1 and 4 months after (n = 32 and 20, respectively). Two groups of low- (n = 44) and medium- (n = 34) exposure workers not directly exposed to Bt spraying were also assessed. The investigation included questionnaires, nasal/mouth lavages, ventilatory function assessment, and skin tests to indigenous aeroallergens and to a variety of Bt spore and vegetative preparations. To authenticate exposure to the organism present in the commercial preparation, isolates from lavage specimens were tested for Bt genes by DNA-DNA hybridization. Humoral immunoglobulin G (IgG) and immunoglobulin E (IgE) antibody responses to spore and vegetative Bt extracts were assayed. There was no evidence of occupationally related respiratory symptoms. Positive skin-prick tests to several spore extracts were seen chiefly in exposed workers. In particular, there was a significant (p < 0.05) increase in the number of positive skin tests to spore extracts 1 and 4 months after exposure to Bt spray. The number of positive skin test responses was also significantly higher in high (p < 0.05) than in low- or medium-exposure workers. The majority of nasal lavage cultures from exposed workers was positive for the commercial Bt organism, as demonstrated by specific molecular genetic probes. Specific IgE antibodies were present in more high-exposure workers (p < 0.05) than in the low and medium groups. Specific IgG antibodies occurred more in the high (p < 0.05) than in the low-exposure group. Specific IgG and IgE antibodies to vegetative organisms were present in all groups of workers. Exposure to Bt sprays may lead to allergic skin sensitization and induction of IgE and IgG antibodies, or both.

Transurethral injection of silicone microimplants for intrinsic urethral sphincter deficiency.

OBJECTIVE: To assess the short-term efficacy of transurethral injection of silicone microimplants in women with intrinsic sphincter deficiency. METHODS: During January 1995 and December 1996, 32 women (mean age 64.3 years, range 39-85 years) with type III stress incontinence (intrinsic sphincter deficiency) underwent transurethral injection of silicone microimplants under general anesthesia. Twenty-eight had undergone previous continence surgery. Subjective and urodynamic assessments were made at 6 and 12 months after injection to evaluate success and short-term effects. RESULTS: Objective and subjective success rates were 75% and 59% at 6 and 12 months, respectively. Injections of silicone microimplants significantly increased maximum urethral closure pressure (maximum urethral pressure at rest: 34.40+/-16.46 cm H2O, 95% confidence interval [CI] 28.55, 40.25 versus 25.35+/-10.78 cm H2O, 95% CI 21.52, 29.18; P = .027). There were no complications after surgery up to 1 year. CONCLUSION: Transurethral silicone injections were effective in 60% of cases of intrinsic sphincter deficiency, although there was a time-dependent decrease.

Neuropeptide denervation alters both the elicitation and induction phases of contact hypersensitivity in mice.

The effects of permanent disruption of neuropeptide transmission on the induction (i.e., sensitization) and elicitation (i.e., challenge) phases of contact hypersensitivity (CHS) are described. BALB/c mice were chemically denervated of neuropeptide (i.e., tachykinin) containing sensory C fibers by an acute injection of capsaicin (50 mg/kg) on postnatal day (PND) 2 to 3. As young adults (PND 45-60), these mice and their control littermates were sensitized by topical application of 0.1% 2,4-dinitrofluorobenzene (DNFB) or vehicle. Treatment groups generated from this exposure regimen consisted of untreated, controls (O/O), denervated, controls (CAP/O), untreated, sensitized (O/DNFB), and denervated, sensitized (CAP/DNFB). The elicitation phase of CHS was evaluated in these animals by measuring ear thickness in response to a DNFB challenge. In DNFB-sensitized groups, ear thickness was significantly increased over controls but was additionally increased 2.4-fold in CAP/DNFB compared to O/DNFB mice. The induction phase of CHS was next assessed in young adult mice by measuring lymph node cell (LNC) proliferation. For this, mice were sensitized for 3 consecutive days before their draining, auricular nodes were removed. The LNC were dissociated and cultured for 24 h with tritiated thymidine to assess LNC proliferation. As expected, significantly higher numbers of LNC occurred in both DNFB-sensitized groups (CAP/DNFB, O/DNFB) compared to the unsensitized, controls (CAP/O, O/O). However, LNC proliferation in CAP/DNFB was significantly higher than O/DNFB animals. Flow cytometry on similarly exposed mice failed to demonstrate any significant difference in the population of CD4CD8 or CD3CD45R LNC cells from neuropeptide-denervated (CAP/O, CAP/DNFB) mice or their respective treatment mates (O/O, O/DNFB), suggesting that alterations in T or B cell populations did not underlie these changes. Finally, cytokine release from the LNC from these treatment groups was examined. For this, the auricular lymph nodes were removed from animals, 2 to 4 h after the animals were administered a single application of a sensitizing concentration (0.1%) of DNFB or acetone vehicle. LNC, dissociated from these nodes, were cultured for 24 h. The nutrient media was removed from these cultured cells and examined for the release of proinflammatory cytokines, interleukin (IL)-1beta, IL-2 and tumor necrosis factor (TNF)alpha, by ELISA. There were no significant increases in IL-2. However, IL-1beta release was significantly increased in CAP/DNFB mice over O/DNFB by 18-fold and by over 30-fold compare to O/O controls. Levels of TNFalpha were significantly increased in both O/DNFB and CAP/DNFB mice over the nonsensitized controls (O/O, CAP/O). CAP/DNFB values were approximately double that of O/DNFB. There was no significant difference in IL-1beta or TNFalpha release between the nonsensitized controls (O/O, CAP/O). Collectively, these data indicate that neuropeptide denervation by neonatal administration of capsaicin alters both the induction and elicitation phases CHS and may modify sensitivity to chemically induced CHS.

Neuropeptide modulation of chemically induced skin irritation.

This study addresses the hypothesis that the early symptoms of chemically induced skin irritation are neurally mediated. Several approaches were used to affect nerve transmission in adult Balb/c female mice. These included general anesthesia (i.e., sodium pentobarbital), systemic capsaicin treatment, and pretreatment with specific pharmacological antagonists of the neuropeptides substance P (SP) and neurokinin A (NKA). After these treatments, a strongly irritating dose of dinitrofluorobenzene (DNFB) was applied to the ear and its swelling was measured over several postexposure times as an index of tissue irritation. Ear swelling in Nembutal (30 mg/kg)-anesthetized mice was depressed 62 and 76% at 4 and 24 hr postexposure compared to DNFB-treated unanesthetized animals measured at the same time points. Multiple injections of capsaicin (cumulative dose 30 mg/kg) depressed DNFB-ear swelling relative to non-capsaicin, DNFB-treated controls by 15, 40 (ip), and 44 and 43% (sc) at 4 and 24 hr postexposure, respectively. In mice exposed to acute or multiple injections of the SP antagonist CP-96,345 before DNFB application, ear swelling was depressed (relative to DNFB-treated animals) by 64 and 36% (acute, sc, 10 mg/kg) and 91 and 88% (multiple, ip, cumulative 35 mg/kg) at 0.5 and 1 hr postexposure, respectively. Mice exposed to the NKA antagonist, SR 48968, alone and in combination with the SP antagonist CP-96,345 were also examined after DNFB application. Ear swelling was diminished in mice pretreated with the NKA antagonist (1.0 mg/kg) by 17, 24, 34, and 40% at 0.5, 1, 4, and 24 hr postexposure. When used in combination with the SP antagonist, DNFB-induced ear swelling was reduced by 95% compared to unantagonized, DNFB-exposed mice at the 0.5- and 1-hr time points and remained significantly depressed by 33 and 46% at 4 and 24 hr postexposure. Taken in concert, these data suggest that neuropeptides, especially the tachykinins SP and NKA, modulate the early stages of chemically induced skin irritation.

Restrictive lung disease in rats exposed chronically to an urban profile of ozone.

The potential for irreversible lung impairment resulting from life-long ozone (O3) exposure remains uncertain. To address this question, young adult rats (male, F-344) were exposed to a simulated urban profile of O3 for 1, 3, 13, 52, or 78 wk, after which pulmonary function tests were performed. To assess reversibility of effects, cohorts from the 13-, 52-, and 78-wk groups were evaluated, respectively, after an additional 6, 27, and 17 wk of clean air. Static and dynamic lung properties were based on measurements of lung volume apportionment, respiratory system compliance (Crs), DLCO, multibreath N2 washout, and maximum expiratory flow-volume relationships. Electrocardiography was also performed in unanesthetized, restrained rats after 52 and 78 wk, as were determinations of wet and dry lung weights, lung collagen, and associated connective tissue crosslinks. Small (< 10%) but significant reductions in TLC and RV were noted after 13, 52, and 78 wk of O3 exposure. At 13 and 52 wk, N2 washout was enhanced, though at 78 wk it was similar to control. None of these changes appeared progressive with continued O3 exposure. Post exposure to clean air did not completely reverse the reduction in TLC. Additionally, Crs, though not affected during O3 exposure, decreased during the air recovery. No O3-related changes in collagen were apparent, however. Thus, near life-long exposure of F-344 rats to a worse-case, urban profile of O3 appears to have led to a functionally restrictive, i.e. "stiffened," lung without overt fibrosis. Furthermore, certain aspects of the O3-induced effect were not fully reversible.

Dietary vitamin A enhances sensitivity of the local lymph node assay.

Murine assays such as the mouse ear swelling test (MEST) and the local lymph node assay (LLNA) are popular alternatives to guinea pig models for the identification of contact sensitizers, yet there has been concern over the effectiveness of these assays to detect weak and moderate sensitizers. Much work has been done to improve the sensitivity of the MEST, including the addition of a vitamin A acetate (VAA) enriched diet, which increases its sensitivity. Vitamin A acetate has been reported to increase the numbers of Langerhans cells (antigen presenting cells) in the skin, which could in turn enhance the cellular immune response. Because the LLNA relies on tritiated-thymidine incorporation by proliferating T cells during the induction phase, we have studied the potential of the VAA diet to enhance sensitivity of the LLNA. Results indicate that the VAA enriched diet significantly increased the LLNA sensitivity to formalin, eugenol, glutaraldehyde, trimellitic anhydride, and an azo dye at concentrations where no proliferation was observed in mice maintained on the standard diet. Maintenance on a VAA diet for 3 weeks prior to initiating the sensitization procedure was optimal. Thus, incorporation of a VAA diet improves the sensitivity of the LLNA as a quick, objective, and relatively inexpensive screen for detecting moderate and weak contact sensitizers.

Ozone adaptation in rats after chronic exposure to a simulated urban profile of ozone.

Studies in both humans and rats have indicated that certain pulmonary responses induced by exposure to an acute provocative concentration of ozone (O3) will eventually attenuate if the exposure is repeated on a daily basis. This phenomenon is commonly referred to as O3 adaptation. Whether or not a "state" of adaptation develops due to long-term low level O3 exposure is unknown. Two human studies have reported adaptation in subjects living in Los Angeles during periods when ambient O3 concentrations have been relatively high. At present, however, we are not aware of comparable information from rats. This study assessed O3 adaptation in rats following chronic (12 or 18 months) exposure and after a 4-month recovery period. A chronic exposure pattern, similar to that found in an urban area during the summer (0.06 ppm O3 for 13 hr/day, 7 days/week; Monday-Friday, peak to 0.25 ppm O3, over 9 hr), was used. To assess whether adaptation had occurred and/or persisted, awake rats were challenged with high provocative concentrations of O3 for up to 2 hr. During a challenge, rats were monitored for typical O3-induced alterations in spontaneous breathing parameters (e.g., increase in breathing frequency and decrease in tidal volume). Adaptation was defined as attenuation of breathing response during the challenge in rats chronically exposed to O3 as compared to that in "control" rats (chronically exposed to air). Adaptation was found in the rats within 8 hr following the chronic O3 exposure but not after the 4-month recovery period. Spontaneous breathing parameters that were significantly attenuated in the chronically exposed rats were breathing frequency, tidal volume, inspiratory and expiratory times, and maximum expiratory flow. We conclude that rats demonstrated adaptation to O3 after long-term exposure to an urban-type O3 profile and that the adaptation was not seen 4 months postexposure. These results suggest that exposure to environmental O3 in Los Angeles air may have been responsible for the adaptation found in residential subjects.

Evaluation of an azo and two anthraquinone dyes for allergic potential.

Two dye mixtures and the individual component dyes were evaluated for the potential to induce contact or pulmonary hypersensitivity. These dye mixtures were suspect because of anecdotal reports of both pulmonary and contact hypersensitivity in assembly workers, and because the component dyes were structurally related to dyes known to be contact sensitizers. One mixture consisted of disperse blue 3 (DB3) and disperse red 11 (DR11), which are anthraquinones, and the other mixture contained DR11 and solvent red 1 (SR1), an azo dye. Contact hypersensitivity was examined using the local lymph node assay (LLNA) and a modified mouse ear swelling test (MEST). Both the MEST and the LLNA indicated that SR1 has weak contact-sensitizing potential. None of the other individual dye compounds or the two mixtures were identified as contact sensitizers by either method. To evaluate the mixtures as potential pulmonary allergens, guinea pigs were repeatedly exposed by inhalation (300 mg/m3, 6 hr/day) 5 days/week, for 1 week. Weekly exposures were repeated three times with 2 weeks of nonexposure time in between. Guinea pigs were then challenged through the jugular vein using a dye-dimethylsulfoxide mixture. During the challenge, breathing mechanics (dynamic compliance and resistance) were measured in mechanically ventilated animals. Changes in these measurements, indicative of bronchoconstriction, were not observed in animals exposed to either dye mixture, nor were antibodies detected in the sera of exposed animals using individual dye-specific enzyme-linked immunosorbent assays. In conclusion, two methods indicate that SR1 may have contact-sensitizing potential.(ABSTRACT TRUNCATED AT 250 WORDS)

Role of tachykinins in ozone-induced acute lung injury in guinea pigs.

To examine the hypothesis that the acute reversible changes caused by ozone (O3) exposure are mediated by tachykinin release, guinea pigs were depleted of tachykinins by use of repeated capsaicin (CAP) injections before O3 exposure in an attempt to prevent O3-induced functional changes. Unexpectedly, CAP pretreatment caused divergent results in the functional responses to O3. Ventilatory measurements obtained from CAP-pretreated O3-exposed (CAP-O3) animals were exacerbated rather than diminished compared with the effects of O3 alone. Similarly, lavage fluid protein accumulation was enhanced in the CAP-O3 group compared with the O3-exposed group. In better agreement with our initial hypothesis, the CAP-O3 group was less responsive than the O3-exposed animals to histamine aerosol challenge. Additionally, Evans blue dye accumulation, a hallmark of tachykinin release, was increased in O3-exposed animals and was partially blocked in the CAP-O3 group. These data suggest that tachykinin-containing sensory fibers are unlikely to mediate the acute effects of O3 exposure on tidal breathing and lavage fluid protein accumulation but may play a role in causing post-O3 airway hyperreactivity and protein extravasation into the trachea.

Factors that influence the suppression of pulmonary antibacterial defenses in mice exposed to ozone.

Exposure to ozone (O3) has been shown to increase susceptibility of mice to bacterial infection; however, the underlying mechanism has not been well elucidated. This study investigated the effect of O3 exposure on the ability of mice to combat an infectious challenge of Streptococcus zooepidemicus. Following a 3-h exposure to either air, 0.4 ppm O3, or 0.8 ppm O3, 5- and 9-week-old mice received an aerosol infection of bacteria. Intrapulmonary killing of the bacteria was impaired in the O3-exposed mice. The effect was most severe at the higher dose of O3 in the younger mice, and showed good correlation to subsequent mortality assessed over a 20-day period. Alveolar macrophages (AM) from O3-exposed mice had an impaired ability to phagocytose the bacteria. Additionally, prostaglandin E2 (PGE2) levels, which are known to depress AM function, were increased in the bronchoalveolar lavage fluid of the younger mice following exposure to O3, while pretreatment with indomethacin in the drinking water blunted the increased of PGE2 and reduced O3 enhanced mortality from 53 to 33%. The data show that O3 inhalation can reduce the defensive capability of the murine lung and that this is associated with a reduction in AM phagocytosis. The defect is more marked in young mice, suggesting that they may be more susceptible to oxidant exposure. Further studies are required to distinguish between direct toxicity of O3 on the AM and indirect suppression due to modulation of pharmacologic or inflammatory mediators.

Near-lifetime exposure of the rat to a simulated urban profile of nitrogen dioxide: pulmonary function evaluation.

To investigate the potential for up to a near-lifetime exposure to high-ambient levels of nitrogen dioxide (NO2) to induce functional lung damage, groups of rats were exposed to air or a simulated urban profile of NO2 (0.5 ppm background, 1.5 ppm peak) for 1, 3, 13, 52, or 78 weeks. The dynamic, static, and diffusional characteristics of the lung were evaluated postexposure in anesthetized rats. Furthermore, for the 13-, 52-, and 78-week groups, additional animals were tested after a 6-, 26-, or 17-week period in filtered air, respectively. No significant NO2 differences between exposed and control animals were found for the nitrogen washout, compliance, lung volume, or diffusion capacity of carbon monoxide measurements. At 78 weeks, however, a reduction in delta FEF25%, an estimate of convexity in the later portion of the forced expiratory flow volume curve, was observed. Breathing patterns and mechanisms were also assessed postexposure in a parallel group of similarly exposed unanesthetized rats. These rats were examined during a filtered air, 4 and 8% carbon dioxide (CO2) challenge. In the unanesthetized rat, frequency of breathing was significantly decreased and tidal volume, expiratory resistance, and inspiratory and expiratory times tended to increase. For several of these variables, the largest response also occurred at 78 weeks and seemed to be exacerbated by CO2 challenge.(ABSTRACT TRUNCATED AT 250 WORDS)

Effects of inhaled phosgene on rat lung antioxidant systems.

A concentration-response and C x T study were undertaken to determine the effect of phosgene (COCl2) inhalation on pulmonary antioxidant processes as determined by changes in endogenous glutathione (GSH) and antioxidant-associated enzymes (GSH peroxidase, GSH reductase, glucose-6-phosphate dehydrogenase, and superoxide dismutase). Rats were exposed to 0.0, 0.1, 0.25, 0.5, and 1.0 ppm phosgene for 4 hr and 0.25 ppm phosgene for 8 hr. The endpoints were assayed at 0, 1, 2, 3 and 7 days after exposure cessation. The lowest effective concentration was 0.1 ppm phosgene (increases in measured variables from 8 to 35% above control values). At all concentrations, major effects were observed 1 to 2 days after exposure (12 to 159% above control), peaking at 2 to 3 days postexposure (11 to 253% above control), and in some cases were still evident 7 days (10 to 65% above control) after exposure. The C x T study using the same dose (120 ppm-min), but different times and concentration (0.25 ppm for 8 hr and 0.5 ppm for 4 hr), showed a concentration dependence. The peak antioxidant enzyme changes observed for the higher concentration (0.5 ppm) were at least double those observed for the lower concentration (0.25 ppm). These enzyme changes were similar to those reported for the oxidants O3 and NO2. Although the suspected mechanism of initial damage between phosgene and these oxidants is different (acylation vs oxidation) the biological result is similar (i.e., damage, repair, and influx of cells), thus eliciting similar biochemical changes in response to pulmonary injury.(ABSTRACT TRUNCATED AT 250 WORDS)

Effects of 20,25-diazacholesterol on cholesterol synthesis in cultured chick muscle cells: a radiogas chromatographic and mass spectrometric study of the post-squalene sector.

Radiogas chromatography, used in conjunction with mass spectrometry, has been used to analyze the sterol content of cultured chick muscle cells. Seven sterols, plus lanosterol, were detected. These sterols conformed to a linear biosynthetic pathway linking lanosterol and cholesterol. The reaction sequence is: C-14 demethylation, C-4 demethylation, delta 8 leads to delta 5 double bond rearrangement, delta 24 double bond reduction. When chick cells were treated with increasing concentrations of 20,25-diazacholesterol, components of this pathway and aberrant products accumulated. These accumulations suggest that diazacholesterol affects reductases, double bond isomerases and the C-14 demethylation enzymes of sterol biosynthesis.

6-Methylsalicylic Acid Production in Solid Cultures of Penicillium patulum Occurs Only When an Aerial Mycelium Is Present.

When Penicillium patulum was grown on Czapek-Dox agar, 6-methylsalicylic acid was produced as an aerial mycelium was forming. Nutrients were often plentiful in the medium when biosynthesis began. If the formation of an aerial mycelium was prevented by growing the fungus between two sheets of dialysis membrane, no 6-methylsalicylic acid was produced even when nutrients were completely consumed. If the upper sheet of dialysis membrane was stripped off cultures of the latter type, an aerial mycelium formed; concomitantly, 6-methylsalicylic acid biosynthesis was observed. We conclude that 6-methylsalicylic acid was produced only by P. patulum colonies that possessed an aerial mycelium.

Mycophenolic Acid Production by Penicillium brevicompactum on Solid Media.

When grown on Czapek-Dox agar, Penicillium brevicompactum produced mycophenolic acid after a vegetative mycelium had been formed and as aerial hyphae were developing. Nutrients were still plenteous in the agar when the synthesis began. If aerial hyphal development was prevented by placing a dialysis membrane over the growing fungus, no mycophenolic acid was produced. When the dialysis membrane was peeled back and, as a consequence, production of aerial hyphae began, mycophenolic acid biosynthesis was observed. We concluded that mycophenolic acid was produced only by P. brevicompactum colonies that possessed an aerial mycelium.

Radiogas chromatography mass spectrometry in the selected ion monitoring mode.

The value of selected ion monitoring in analyzing biological radio isotope incorporation experiments by radiogas chromatography mass spectrometry is illustrated with reference to the biosynthesis of the mycotoxin mycophenolic acid and the mode of action of the anticholesterolemic drug 20,25-diazacholesterol. It is shown that the increased sensitivity and specificity of the selected ion monitoring mode detector permits straightforward detection and identification of the relatively small cellular pools associated with metabolic intermediates. The computer program RADSIM is described. Problems that still exist in using radiogas gas chromatography mass spectrometry technology to analyse isotope incorporation experiments are discussed.