Impact of Diflubenzuron on Bombus impatiens (Hymenoptera: Apidae) Microcolony Development.

Reliance on the honey bee as a surrogate organism for risk assessment performed on other bees is widely challenged due to differences in phenology, life history, and sensitivity to pesticides between bee species. Consequently, there is a need to develop validated methods for assessing toxicity in non-Apis bees including bumble bees. The usefulness of small-scale, queenless colonies, termed microcolonies, has not been fully investigated for hazard assessment. Using the insect growth regulator diflubenzuron as a reference toxicant, we monitored microcolony development from egg laying to drone emergence using the Eastern bumble bee Bombus impatiens (C.), a non-Apis species native to North America. Microcolonies were monitored following dietary exposure to diflubenzuron (nominal concentrations: 0.1, 1, 10, 100, and 1,000 µg/liter). Microcolony syrup and pollen consumption was significantly reduced by diflubenzuron exposure. Pupal cell production was also significantly decreased at the highest diflubenzuron concentration assessed. Ultimately, diflubenzuron inhibited drone production in a concentration-dependent manner and a 42-d 50% inhibitory concentration (IC50) was determined. None of the dietary concentrations of diflubenzuron tested affected adult worker survival, or average drone weight. These data strengthen the foundation for use of this methodology, and provide valuable information for B. impatiens; however, more work is required to better understand the utility of the bumble bee microcolony model for pesticide hazard assessment.

Development and utilization of a unique in vitro antigen presentation co-culture model for detection of immunomodulating substances.

Current regulatory immunotoxicity studies require the use of animal models. However, evolving regulatory requirements, the need to evaluate large numbers of chemicals efficiently and societal pressures are driving the development and utilization of alternative in vitro methods for identifying potential immunotoxicants. In line with these efforts, we developed a novel in vitro cell-based assay to evaluate effects on antigen presentation - a key step in successful immunization. In this assay, Ch27 B cells acquire and present hen egg lysozyme peptides to antigen-restricted 3A9 T cells, causing them to produce and secrete IL-2. IL-2 levels in the culture medium may be monitored to identify effects of immunotoxicant exposure on antigen uptake, processing or presentation by the Ch27 cells and on antigen recognition and IL-2 production and secretion by the 3A9 cells. IL-2 production was reduced in response to treatment with well-known immunotoxicants cyclosporin A (CYA), dexamethasone (DEX), azathioprine (AZPR), methotrexate (MOT) and benzo(a)pyrene (BAP) but was not affected by treatment with cyclophosphamide (CYPH). A negative control compound mannitol (MANN) altered neither cell viability nor IL-2 levels whereas the lysosomotrophic compound ammonium chloride (AMCL) reduced IL-2 production. This novel in vitro assay of immune function may be suitable for integration into a tiered testing battery for screening and prioritization of potential immunosuppressants.

Dual neurocircuitry dysfunctions in disruptive behavior disorders: emotional responding and response inhibition.

BACKGROUND: To determine the functional integrity of the neural systems involved in emotional responding/regulation and response control/inhibition in youth (age 10-18 years) with disruptive behavioral disorders (DBDs: conduct disorder and/or oppositional defiant disorder) as a function of callous-unemotional (CU) traits. METHOD: Twenty-eight healthy youths and 35 youths with DBD [high CU (HCU), n = 18; low CU (LCU), n = 17] performed the fMRI Affective Stroop task. Participants viewed positive, neutral, and negative images under varying levels of cognitive load. A 3-way ANOVA (group×emotion by task) was conducted on the BOLD response data. RESULTS: Youth with DBD-HCU showed significantly less activation of ventromedial prefrontal cortex (vmPFC) and amygdala in response to negative stimuli, compared to healthy youth and youth with DBD-LCU. vmPFC responsiveness was inversely related to CU symptoms in DBD. Youth with DBD-LCU showed decreased functional connectivity between amygdala and regions including inferior frontal gyrus in response to emotional stimuli. Youth with DBD (LCU and HCU) additionally showed decreased insula responsiveness to high load (incongruent trials) compared to healthy youth. Insula responsiveness was inversely related to ADHD symptoms in DBD. CONCLUSIONS: These data reveal two forms of pathophysiology in DBD. One associated with reduced amygdala and vmPFC responses to negative stimuli and related to increased CU traits. Another associated with reduced insula responses during high load task trials and related to ADHD symptoms. Appropriate treatment will need to be individualized according to the patient's specific pathophysiology.

Lung function changes in mice sensitized to ammonium hexachloroplatinate.

Occupational exposure to halogenated platinum salts can trigger the development of asthma. The risk to the general population that may result from the use of platinum in catalytic converters and its emerging use as a diesel fuel additive is unclear. To investigate pulmonary responses to platinum, we developed a mouse model of platinum hypersensitivity. Mice were sensitized through application of ammonium hexachloroplatinate (AHCP) to the shaved back on days 0, 5 and 19, and to each ear on days 10, 11 and 12. On days 24 and 29, mice were challenged by oropharyngeal aspiration with AHCP in saline. Before and immediately after challenge, pulmonary responses were assessed using whole body plethysmography (WBP). A dose-dependent increase in immediate responses was observed in AHCP-sensitized and challenged mice. On days 26 and 31, changes in ventilatory responses to methacholine (Mch) aerosol were assessed by WBP; dose-dependent increases in Mch responsiveness occurred in sensitized mice. Lymph node cell counts indicate a proliferative response in lymph nodes draining the sites of application. Bronchoalveolar lavage fluid harvested from sensitized mice contained an average of 5% eosinophils compared to less than 0.5% in non-sensitized mice (p < 0.05); significant increases in total serum immunoglobulin E were observed for all sensitized mice. Although a second airway challenge on day 29 affected some results, only one airway challenge was needed to observe changes in lung function.

Development and utilization of an ex vivo bromodeoxyuridine local lymph node assay protocol for assessing potential chemical sensitizers.

The murine local lymph node assay (LLNA) is widely used to identify chemicals that may cause allergic contact dermatitis. Exposure to a dermal sensitizer results in proliferation of local lymph node T cells, which has traditionally been measured by in vivo incorporation of [(3) H]methyl thymidine. A more recent non-isotopic variation of the assay utilizes bromodeoxyuridine (BrdU) incorporation in vivo. To further improve the utility of this assay, we developed an ex vivo BrdU labeling procedure eliminating the need for in vivo injections. The results of this assay correctly identified a strong sensitizer (i.e., trimellitic anhydride) as well as weak/moderate sensitizers (i.e., eugenol, cinnamaldehyde and hexylcinnaminic aldehyde). As anticipated, neither non-sensitizers isopropanol and lactic acid nor the false negative chemical nickel II sulfate hexahydrate induced a positive threshold response in the assay. The results of this assay are in close agreement with those of the in vivo LLNA:BrdU-enzyme-linked immunosorbent assay labeling procedure. We also used the ex vivo BrdU LLNA procedure to evaluate ammonium hexachloroplatinate, ammonium tetrachloroplatinate and cis-diamminedichloroplatinum(II) and the assay correctly identified them as sensitizers based on the calculation of EC2 values. We conclude that this ex vivo BrdU labeling method offers predictive capacity comparable to previously established LLNA protocols while eliminating animal injections and the use of radioisotope. Published 2014. This article is a U.S. Government work and is in the public domain in the USA.

Relative potency based on hepatic enzyme induction predicts immunosuppressive effects of a mixture of PCDDS/PCDFS and PCBS.

The toxic equivalency factor (TEF) approach was employed to compare immunotoxic potency of mixtures containing polychlorinated dibenzo-p-dioxins, polychlorinated dibenzofurans and polychlorinated biphenyls relative to 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), using the antibody response to sheep erythrocytes (SRBC). Mixture-1 (MIX-1) contained TCDD, 1,2,3,7,8-pentachlorodibenzo-p-dioxin (PeCDD), 2,3,7,8-tetrachlorodibenzofuran (TCDF), 1,2,3,7,8-pentachlorodibenzofuran (1-PeCDF), 2,3,4,7,8-pentachlorodibenzofuran (4-PeCDF), and 1,2,3,4,6,7,8,9-octachlorodibenzofuran (OCDF). Mixture-2 (MIX-2) contained MIX-1 and the following PCBs, 3,3',4,4'-tetrachlorobiphenyl (IUPAC No. 77), 3,3',4,4',5-pentachlorobiphenyl (126), 3,3',4,4',5,5N-hexachlorobiphenyl (169), 2,3,3',4,4'-pentachlorobiphenyl (105), 2,3',4,4',5-pentachlorobiphenyl (118), and 2,3,3',4,4',5-hexachlorobiphenyl (156). The mixture compositions were based on relative chemical concentrations in food and human tissues. TCDD equivalents (TEQ) of the mixture were estimated using relative potency factors from hepatic enzyme induction in mice [DeVito, M.J., Diliberto, J.J., Ross, D.G., Menache, M.G., Birnbaum, L.S., 1997. Dose-response relationships for polyhalogenated dioxins and dibenzofurans following subchronic treatment in mice. I .CYP1A1 and CYP1A2 enzyme activity in liver, lung and skin. Toxicol. Appl. Pharmacol. 130, 197-208; DeVito, M.J., Menache, G., Diliberto, J.J., Ross, D.G., Birnbaum L.S., 2000. Dose-response relationships for induction of CYP1A1 and CYP1A2 enzyme activity in liver, lung, and skin in female mice following subchronic exposure to polychlorinated biphenyls. Toxicol. Appl. Pharmacol. 167, 157-172] Female mice received 0, 1.5, 15, 150 or 450 ng TCDD/kg/day or approximately 0, 1.5, 15, 150 or 450 ng TEQ/kg/day of MIX-1 or MIX-2 by gavage 5 days per week for 13 weeks. Mice were immunized 3 days after the last exposure and 4 days later, body, spleen, thymus, and liver weights were measured, and antibody response to SRBCs was observed. Exposure to TCDD, MIX-1, and MIX-2 suppressed the antibody response in a dose-dependent manner. Two-way ANOVA indicated no differences in the response between TCDD and the mixtures for body weight, spleen/body weight and decreased antibody responses. The results support the use of the TEF methodology and suggest that immune suppression by dioxin-like chemicals may be of concern at or near background human exposures.

CYP1A2 is not required for 2,3,7,8-tetrachlorodibenzo-p-dioxin-induced immunosuppression.

One of the most sensitive and reproducible immunotoxic endpoints of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) exposure is suppression of the antibody response to sheep red blood cells (SRBCs) in mice. Immunosuppression occurs in concert with hepatomegaly and associated induction of several hepatic cytochrome P450 enzymes, including CYP1A2 which is responsible for the hepatic sequestration of TCDD. In this study, TCDD-induced immunosuppression was evaluated in C57BL/6N CYP1A2 (+/+) wild-type and compared with that of age-matched CYP1A2 (-/-) knockout and CYP1A2 (+/-) heterozygous female mice. Groups of mice were given a single gavage dose of 0, 0.03, 0.1, 0.3, 1.0, 3.0 or 10.0microg TCDD/kg, followed 7 days later by immunization with SRBCs. Serum was obtained 5 days after immunization and body, spleen, thymus and liver weights were measured. sheep red blood cell (SRBC) antibody titers were determined by an enzyme-linked immunosorbent assay (ELISA). Anti-SRBC titers were suppressed at 1.0, 1.0 and 0.3microg TCDD/kg for CYP1A2 (+/+), CYP1A2 (+/-), and CYP1A2 (-/-) mice, respectively, which indicated a three-fold increase in TCDD-induced immunosuppression for the CYP1A2 (-/-) mice. This increase in TCDD-induced immunosuppression may be due to the inability of CYP1A2 (-/-) mice to sequester TCDD in the liver leading to a higher dose to the immune system. In CYP1A2 (+/+) mice, a dose of 3.0microg TCDD/kg was sufficient to increase the liver weight, while in CYP1A2 (-/-) mice no increase in liver weight was observed. Application of analysis of variance and dose-response modeling approaches indicate that there is little evidence that the immunosuppression dose-response curves, for the three strains, differ in the lower part of the dose-response range. Thus, CYP1A2 is not required for TCDD-induced immunosuppression in the mouse.

The effects of perinatal/juvenile heptachlor exposure on adult immune and reproductive system function in rats.

This study was performed to determine if developmental exposure of rats to heptachlor (H) during the last half of gestation through puberty adversely affects adult functioning of the immune and reproductive systems. Time-bred pregnant female Sprague-Dawley rats were dosed by gavage with H (0, 30, 300, or 3000 microg/kg/day) from gestation day (GD) 12 to postnatal day (PND) 7, followed by direct dosing of the pups with H through PND 42. Separate groups of rats were evaluated with a battery of immune function tests, while other groups of rats were evaluated for reproductive development and function. Additional groups of rats were euthanized at the end of the dosing period for histological analyses of major organ systems. Some dams and PND 7 pups were euthanized; milk, plasma, fat and/or tissues were assayed for H and heptachlor epoxide B (HEB), a major metabolite of H. The amount of H and HEB found in milk, blood, fat, and tissues was proportional to the dose of H administered. There were no effects on the number or survival of pups born to H-exposed dams nor to pups exposed postnatally. There were no effects on the number of treated dams delivering litters or on litter size, nor were there any effects on any of the reproductive end points examined in the F(0) or F(1) rats. There were no effects of H exposure on lymphoid organ weights, splenic natural killer (NK) cell activity, and splenic lymphoproliferative (LP) responses to mitogens and allogeneic cells in a mixed lymphocyte response (MLR) assay at 8 weeks of age. H exposure did not alter delayed or contact hypersensitivity at 10 or 17 weeks of age, respectively. However, the primary IgM antibody response to sheep red blood cells (SRBCs) was suppressed in a dose-dependent manner in males, but not females, at 8 weeks of age. The percentage of B lymphocytes (OX12(+)OX19(-)) in spleen was also reduced in the high-dose males. The anti-SRBC IgM response was reduced only in males exposed to 30 microg H/kg/day in a separate group of rats 21 weeks of age. In these same rats, at 26 weeks of age, the secondary IgG antibody response to SRBCs was suppressed in all of the H-exposed males, but not females. These data indicate that perinatal exposure of male rats to H results in suppression of the primary IgM and secondary IgG anti-SRBC responses. Suppression of these antibody responses persisted for up to 20 weeks after the last exposure to H, at a total exposure of approximately 1500 microg H/kg/rat.

Sensitivity of the SRBC PFC assay versus ELISA for detection of immunosuppression by TCDD and TCDD-like congeners.

The splenic antibody plaque forming cell (PFC) assay is a widely used assay in immunotoxicity testing. A recent revision of the Federal Insecticide, Fungicide and Rodenticide (FIFRA) Immunotoxicity test guidelines by the EPA recommended that either the PFC assay or the sheep red blood cell (SRBC) specific serum IgM enzyme-linked immunosorbent assay (ELISA) be used to assess the primary humoral response to SRBCs. The PFC assay quantifies the number of plasma cells in the spleen producing SRBC-specific antibody, while the ELISA measures SRBC-specific IgM antibody in the serum. Because these two assays measure different endpoints, there is a need for comparison of their sensitivity and reliability. The purpose of this project was to determine if these two assays are equally sensitive to suppression of the SRBC response in B6C3F1 female mice. Female B6C3F1 mice were given a single oral exposure to different doses of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) or four TCDD-like congeners. One week later, two sets of mice were immunized with SRBC. The first set was evaluated for the PFC response and the second for the ELISA response, on day 4 or 5 post-immunization, respectively. The four TCDD-like congeners tested were: 1,2,3,7,8-pentachlorodibenzo-p-dioxin (PeCDD), 1,2,3,4,7-pentachlorodibenzofuran (4PeCDF), 3,3',4,4',5-pentachlorobiphenyl (PCB126) and 2,3',4,4',5-pentachlrorbiphenyl (PCB118). The results were used to generate dose-response curves for the determination of the ED(50) for TCDD and each TCDD-like congener. For all chemicals tested, measuring the level of SRBC-specific IgM antibody by ELISA was more sensitive than the PFC assay to detect immunosuppression, as indicated by lower ED(50) values. These results indicate that the SRBC-specific IgM ELISA is a more sensitive assay for detecting the T-cell mediated immunotoxicity of dioxin-like chemicals in this rodent model.

Screening for personality disorders in a nonclinical population.

Based on the Inventory of Interpersonal Problems (IIP), the IIP-PD and the IIP-C screening scales were developed to distinguish personality disorder (PD) from non-PD and Cluster C from other PD, respectively, in a clinic population. Two studies were conducted to determine (a) validity and reliability of these IIP scales for PD screening in a nonclinical population, (b) specificity of IIP-C for identifying Cluster C, and (c) usefulness of the IIP scales for screening Cluster A. College students were screened using the IIP scales (Study 1, N = 454, Study 2, N = 87). High and low scorers completed PD-related questionnaires in Study 1 and a clinical interview for PD symptomatology in Study 2. Results indicated strong test-retest reliability, internal consistency, and factorial, convergent, and external validity. The scales tapped a common deficit in interpersonal relatedness, with some distinction between externalizing and internalizing dimensions, respectively, and both scales were positively and significantly associated with schizotypal traits. In conclusion, the IIP-PD and IIP-C are useful and valid screening instruments for identifying any versus no PD in nonclinical populations.

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.

Alterations in the developing immune system of the F344 rat after perinatal exposure to 2,3,7,8-tetrachlorodibenzo-p-dioxin: II. Effects on the pup and the adult.

Our recent work showed that in utero 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) exposure produced alterations in fetal and neonatal thymocyte subpopulations. This study was designed to determine the persistence and functional significance of these alterations. One group of timed-bred pregnant F344 rats was dosed with 3.0 mcg TCDD/kg by gavage on gestational day 14 (GD14). The immune function of the perinatally-exposed offspring and age-matched controls were assessed at 14-17 weeks old. Examination of the organ weights and splenic phenotypes showed that TCDD exposure increased the spleen/body weight ratio, decreased the thymus/body weight ratio, and decreased the percentage of splenic CD3+/CD4-CD8- cells in both genders. The delayed-type hypersensitivity (DTH) response to bovine serum albumin (BSA) was suppressed in both the TCDD-exposed males and females. The lymphoproliferative (LP) responses to T-cell and B-cell mitogens and the antibody response to sheep red blood cells were not affected by perinatal TCDD exposure in either gender except for a suppressed LP response to PWM in the females. A second set of timed-pregnant F344 rats was dosed with 0 or 1.0 mcg TCDD/kg on GD14. One day after birth litters were cross-fostered to produce control, placental-only, lactational-only, and placental/lactational exposure groups. The organ weights and thymic and splenic phenotypes of these pups were assayed 1, 2, or 3 weeks post-partum, while the DTH response was assessed in 5-month-old males. Increased liver/body weight ratios, decreased percentages of thymic CD3+/CD4-CD8- cells, and increased percentages of thymic CD3+/CD4-CD8+ cells were seen through 3 weeks old in both genders after TCDD exposure. The severity of the effects was related to the route of exposure (i.e. placental/lactational > lactational > placental). The DTH response to BSA was suppressed in the males receiving both placental and lactational exposure. These results suggest that the immunotoxic effects of perinatal TCDD exposure of rats persist into adulthood and that suppression of the DTH response may represent the most sensitive biomarker for TCDD-induced immunotoxicity in this species.

Opposite effects of 2,2',4,4',5,5'-hexachlorobiphenyl and 2,3,7,8-tetrachlorodibenzo-p-dioxin on the antibody response to sheep erythrocytes in mice.

The effect that cotreatment with 2,2',4,4',5,5'-hexachlorobiphenyl (PCB153) and 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) has on the antibody plaque-forming cell (PFC) response to sheep red blood cells (SRBCs) was determined in female B6C3F1 mice. Groups of eight mice per group were given a single oral dose of PCB153 alone (0, 3.58, 35.8, or 358 mg/kg), TCDD alone (0, 0.1, 1, or 10 micrograms/kg), and all possible combinations of these doses in corn oil 7 days prior to immunization with SRBCs. Separate groups of mice were given phenobarbital (PB) parenterally by intraperitoneal injection at a dosage of 160 mg/kg/day for 3 days. Four days after intravenous immunization, body, spleen, thymus, and liver weights and the PFC response to SRBCs were determined. Exposure to TCDD alone resulted in a dose-related suppression of the PFC response, with significant suppression at 1 and 10 micrograms/kg. In contrast, exposure to PCB153 alone resulted in the enhancement of the PFC response at 358 mg/kg. Combined exposure to 358 mg/ kg PCB153 and TCDD resulted in no change (PCB153 + 0.1 microgram/ kg TCDD) or suppression (PCB153 + 1 or 10 micrograms/kg TCDD) of the PFC response relative to PCB153 alone; however, the PFC response was enhanced (PCB153 + 0.1 microgram/kg TCDD), unaffected (PCB153 + 1 microgram/kg TCDD), or suppressed (PCB153 + 10 micrograms/kg TCDD) relative to corn oil controls. PB did not affect the PFC response to SRBCs, despite a 13-fold induction of hepatic pentoxyresorufin O-dealkylase (PROD) activity. These results suggest that PCB153 enhancement of the PFC response is not related to PROD induction and that it acts as a functional antagonist rather than an aryl hydrocarbon receptor or dispositional antagonist. By enhancing the PFC response to SRBCs, PCB153 raises the "setpoint" response level. Consequently, cotreatment with an immunosuppressive dose of TCDD fails to suppress the PFC response relative to corn oil controls, while clearly suppressing it relative to the appropriate control, PCB153 alone.

Physician-hospital organizations and PHO executives. What lies ahead for the PHO?

The 1996/97 Profile of Physician-Hospital Organizations and PHO Executives, jointly conducted by the American Association of Physician-Hospital Organizations/Integrated Delivery Systems (AAPHO/IDS) and Tyler & Company, takes a closer look at the PHO and trends within the health care system. In examining contracting, enrollees, staff size and experience, compensation, incentives, and projected salary increase for CEOs, the survey found that the PHO is in the midst of transformation into an integrated delivery system. As physicians and hospitals form a team to deliver more efficient and cost-effective health care to patients, they are paving the way to the integrated delivery system. The survey also focuses on the details of the PHO and PHO executives, including an in-depth look at all aspects of compensation for the CEO.

In vitro death of jimpy oligodendrocytes: correlation with onset of DM-20/PLP expression and resistance to oligodendrogliotrophic factors.

Severe hypomyelination in the jimpy (jp) mouse mutation results from premature death of most oligodendrocytes (OCs). We have applied an immunopanning technique to successfully purify oligodendroblasts (OBs) directly from neonatal jp brainstem in order to determine if their death during differentiation into OCs is preventable in culture by diffusible oligodendrogliotrophic factors. No significant differences in the yield (0.9-1.1 x 10(5) cells/brainstem) or viability (approximately 90%) of OB populations from jp and wild-type (wt) littermates were observed, indicating that cell death occurs at a later stage in the mutant lineage. When cultured in a basally defined, insulin-containing medium, wt and jp OBs died 1-2 days later as their differentiation into GalC+ OCs began. Survival was not enhanced by known trophic factors (ciliary neurotrophic factor, leukemia inhibitory factor, neurotrophin-3) for differentiating rat OCs. In medium conditioned by neonatally derived rat or wt mouse astrocytes, however, wt OBs survived terminal OC differentiation, expressing first GalC, then DM-20/PLP on their surface 1-2 days later, before elaborating myelin-like membrane. By contrast, jp OBs in sister cultures survived differentiation initially as well as their normal counterparts did but rapidly died thereafter, beginning at the time when PLP/DM-20 immunoreactivity became detectable on premature wt GalC+ OCs. Additionally under these conditions, there survived a minor population (<5%) of jp cells, including mature OCs, which expressed stunted membranes and DM-20/PLP immunoreactivity in their cytoplasm, and undifferentiated progenitors. This model supports the concept that OC death in jp is effected by an intrinsic program, one mechanistically related to jp PLP/DM-20 gene expression and refractory to trophic cues in the environment.

Comparison of the T cell-independent antibody response of mice and rats exposed to 2,3,7,8-tetrachlorodibenzo-p-dioxin.

2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) is an environmental contaminant that produces adverse effects on the immune system of experimental animals. In this study, the effect that TCDD has on the antibody plaque-forming cell (PFC) response to the T cell-independent (TI) antigen trinitrophenyl-lipopolysaccharide (TNP-LPS) was compared in adult female B6C3F1 mice and F344 rats. Mice or rats were given a single intraperitoneal injection of TCDD at doses ranging from 1 to 30 micrograms/kg, 7 days prior to immunization with TNP-LPS by intravenous injection. Three days later body, spleen, thymus, and liver weights were measured and the PFC response to TNP-LPS was determined. Thymus weights were decreased at 10 and 30 micrograms TCDD/kg, whereas spleen weights were decreased and liver weights increased in mice dosed at 3, 10, and 30 micrograms/kg. Mice dosed at 10 and 30 micrograms TCDD/kg had suppressed PFC responses and serum hemagglutination titers. In rats, thymus weights were decreased and liver weights increased at 3, 10, and 30 micrograms TCDD/kg; however, the PFC response and serum hemagglutination titers to TNP-LPS were suppressed only at 30 micrograms/kg TCDD. TCDD did not affect splenic lymphocyte subsets evaluated by flow cytometry. These results indicate that TCDD suppresses the TI antibody response to TNP-LPS in both B6C3F1 mice and F344 rats, with mice more sensitive to suppression by TCDD than rats.

Effectiveness of erythromycin in the treatment of acute bronchitis.

BACKGROUND: Clinical trials have not shown a consistent benefit of treating bronchitis with antibiotics. Many physicians, however, treat acute bronchitis with antibiotics because of the possibility of Mycoplasma pneumoniae or other pathogens. The objectives of this study were to determine the effectiveness of erythromycin treatment in patients with acute bronchitis and to determine whether a newly developed rapid M pneumoniae antibody test is useful in predicting which patients will respond to therapy. METHODS: We conducted a randomized, double-blind, placebo-controlled clinical trial at three primary care centers in North Carolina. A convenience sample of 140 patients presenting with acute bronchitis were tested for M pneumoniae, 91 of whom were treated with either erythromycin 250 mg four times daily for 10 days or an identical-appearing placebo. RESULTS: Patients treated with erythromycin missed an average of only 0.81 +/- 1.1 days of work compared with 2.16 +/- 3.2 days for placebo-treated patients (P < .02). There were no significant differences in cough, use of cough medicine, general feeling of well-being, or chest congestion between the erythromycin and placebo groups. Twenty-five percent of the patients tested positive for M pneumoniae. There were no differences in response to erythromycin based on whether the patient had a positive test for M pneumoniae. CONCLUSIONS: Erythromycin is effective in significantly reducing lost time from work, but it is not effective in reducing cough or other symptoms in patients with acute bronchitis, regardless of the outcome of the M pneumoniae antibody test.

Repeated high dose oral exposure or continuous subcutaneous infusion of 2-methoxyacetic acid does not suppress humoral immunity in the mouse.

2-Methoxyethanol (ME) has been shown to be immunosuppressive in rats but not mice, with oxidation of ME to 2-methoxyacetic acid (MAA) being a prerequisite for immunosuppression. MAA is more rapidly cleared by mice than rats, consequently this study was designed to determine if increasing the bioavailability of MAA in mice might play a role in this species difference. Female B6C3F1 mice were given MAA by oral multiple daily high doses or by continuous subcutaneous infusion via mini-osmotic pumps. Humoral immunity was evaluated in MAA-exposed mice using the plaque-forming cell (PFC) response to either sheep red blood cells (SRBC) or trinitrophenyl-lipopolysaccharide (TNP-LPS). Female F344 rats were also used to compare the effects of multiple daily MAA exposure on these humoral immune responses. Rats and mice were dosed orally twice a day for 4 days by gavage with MAA at dosages ranging from 40-320 mg/kg/day and 240-1920 mg/kg/day, respectively. All animals were immunized on the first day of dosing and body and lymphoid organ weights and PFC responses to SRBC or TNP-LPS were evaluated 4 days later. While body weights in rats were unaffected, thymus weights were reduced at all dosages of MAA and spleen weights were reduced at 160 or 320 mg/kg/day. PFC responses to SRBC and TNP-LPS were suppressed in rats at dosages of 160 and 320 mg/kg/day. In contrast, thymus weights of mice were reduced only at 960 mg/kg/day or greater, with no effect on spleen or body weights. Furthermore, neither the PFC response to SRBC nor the response to TNP-LPS was suppressed in mice exposed to any oral dosage of MAA. In the continuous infusion study, mice were subcutaneously implanted with mini-osmotic pumps containing MAA which was delivered at 840 mg/kg/day over a 7-day period. Continuous exposure to MAA via mini-osmotic pumps did not suppress the PFC response to either SRBC or TNP-LPS, but rather significantly enhanced the response to TNP-LPS. These results indicate that mice are insensitive to MAA even at the high dosages given as a bolus or continuously over 1 week. The data further support earlier work, which suggested that the observed difference between rats and mice for MAA-induced immunosuppression appears to be unrelated to the availability of MAA to target lymphoid tissue in these rodent species.

Astroglial control of oligodendrocyte survival mediated by PDGF and leukemia inhibitory factor-like protein.

Programmed death and the identification of growth factors delaying this process in the oligodendrocyte lineage suggest that other cell types provide oligodendrogliotrophins. To determine their source, primary cultures of oligodendroblasts immunopurified from postnatal rat cerebrum were used to screen other cultured neural and non-neural cell types for the release of survival factors into a defined insulin-containing medium. In non-conditioned medium, oligodendroblasts died 1-2 days after undergoing terminal differentiation into oligodendrocytes, as defined by the onset of expression of galactocerebroside. In medium conditioned by astrocytes, unlike the other tested cell types, differentiated oligodendrocytes survived for weeks in a mature myelinogenic state. Survival was partially reduced by immunoabsorption of the medium with antibodies to platelet-derived growth factor and abolished by immunoabsorption with antibodies to leukemia inhibitory factor. By the same criterion, survival activity was not attributed to other astrocytic products, ciliary neurotrophic factor and basic fibroblast growth factor. Membrane ultrafiltration analysis indicated the activity corresponded to heat-labile protein smaller (M(r) = 10(-30) x 10(3)) than native rat leukemia inhibitory factor (M(r) = 43 x 10(3)). The astrocytic stimulus was > 4-fold more efficacious than other known oligodendrogliotrophic cytokines, including ciliary neurotrophic factor, neurotrophin-3 and leukemia inhibitory factor itself, tested singly or in combination, and promoted survival additively with these agents. These findings suggest that astrocytes function as paracrine regulators of oligodendroblast and oligodendrocyte survival and that their effect is mediated initially by platelet-derived growth factor and thereafter by a powerful cytokine related to leukemia inhibitory factor.

Immunological effects of 2-methoxyethanol administered dermally or orally to Fischer 344 rats.

Exposure of rats to 2-methoxyethanol (ME) by gavage for 10 consecutive days results in immunotoxicity. To determine whether dermal exposure to ME also induces immunotoxicity, undiluted ME was applied to Fisher 344 male rats at dose levels of 150, 300, 600, 900 or 1200 mg/kg/day on shaved occluded test sites for 4 consecutive days. Decreased thymus weights were produced by all doses of ME, while reductions in spleen weight were observed at doses of 900 mg/kg/day ME or greater. The alterations in these lymphoid organ weights were produced in the absence of loss in body weight. The lymphoproliferative (LP) responses to phytohemagglutinin (PHA) and pokeweed mitogen (PWM) were enhanced at 1200 mg/kg/day ME compared with water controls. Separate groups of rats, employed for the antibody plaque-forming cell (PFC) response to either trinitrophenyl-lipopolysaccharide (TNP-LPS) or sheep red blood cells (SRBC), were exposed dermally to 150, 300 or 600 mg/kg/day ME for 4 consecutive days. A reduction in the PFC response to TNP was observed at 600 mg/kg/day ME, whereas decreases in the PFC response to SRBC were observed at dosages of 300 and 600 mg/kg/day ME. To compare the immunotoxic effects of dermally applied ME to those effects caused by ME administered orally, rats were dosed by gavage with 25, 50, 100 or 200 mg/kg/day ME in distilled water for 4 consecutive days. Reductions in thymus weights were observed at oral dosages ranging from 50-200 mg/kg/day, while spleen weights were reduced in rats dosed at 200 mg/kg/day ME. LP responses to PHA, PWM and Salmonella typhimurium were increased at the 200 mg/kg/day ME dose level. PFC responses to TNP-LPS and SRBC were suppressed at the 50, 100 and 200 mg/kg/day ME dosages. These results indicate that, like oral exposure, dermal exposure to ME compromises the ability of the immune system to mount an effective humoral immune response.