Biological effects of gestational and lactational PCB exposure in neonatal and juvenile C57BL/6 mice.

The purpose of this study was to obtain a better understanding of polychlorinated biphenyl (PCB) immunotoxicity in the developing mouse. Adult female mice were dosed with three subcutaneous injections per week of 50 mg/kg Aroclor 1242 (A1242), Aroclor 1254 (A1254), or corn oil for 2 weeks and then mated with nondosed males. First-litter pups were sacrificed at 7 or 28 days of age. At both ages, the tissue concentration of PCB was significantly higher in both the A1242 and A1254 pups than in oil-treated controls. Seven-day-old pups exposed to A1242 or A1254 had significantly decreased splenic IL-2 production. Alterations in the percentages of T cell subsets compared to controls were observed in A1242-exposed pups; an increased spleen somatic index was noted only in A1254-exposed pups. Twenty-eight-day-old pups exposed to A1254 demonstrated a significant decrease in thymus somatic index, an increase in liver somatic index, a 25% decrease in total circulating T(4), and decreased B cell percentages relative to their controls. Alteration in the percentages of CD3(int) T cells was observed in A1254-exposed 28-day-old pups. A significant increase in 7-ethoxyresorufin- O-deethylase (EROD) and 7-benzoxyresorufin-O-dearylase (BROD) activity was measured at both ages in A1254-exposed pups and in A1242-exposed 28-day-old pups. These data confirm that during gestation and lactation A1242 and A1254 are transferred from dams to pups and that such exposure results in immune-related effects in neonatal (7-day-old) and juvenile (28-day-old) mice. Furthermore, A1254 exposure produces more frequent and pronounced effects than exposure to A1242.

Cloning and sequence analysis of cDNAs encoding the heavy and light chain variable regions of an Ab2beta anti-idiotypic monoclonal antibody possessing an internal image of cocaine.

We report here the cloning and sequence analysis of cDNAs encoding the variable regions of an Ab2beta anti-idiotypic monoclonal antibody (K1-4c, gamma1kappa) that mimics the configuration of cocaine. The Ab2beta specifically binds to the human dopamine transporter as shown by confocal immunofluorescence microscopy. The sequence of the heavy chain complementarity-determining region 3 of K1-4c is strikingly similar to that of a monoclonal antibody (F11.2.32) specific for HIV-1 protease. Three or four amino acids in the epitope recognized by the anti-HIV-1 protease antibody are also present in the third extracellular loop of the dopamine transporter. This epitope is within the conserved region of the known transporters for dopamine, norepinephrine and serotonin in Homo sapiens, Rattus norvegicus, Caenorhabditis elegans and Drosophila melanogaster.

The influence of age and gender on the immune system: a longitudinal study in Labrador Retriever dogs.

While aging studies employing a cross-sectional design have been informative in documenting many age-related alterations in immune function between different age cohorts within a population, longitudinal studies are invaluable for verifying changes at the level of the individual and for defining the precise periods of life during which particular changes occur. In the present study, a battery of immunological parameters were evaluated in a group of Labrador Retrievers as part of a comprehensive longitudinal aging study. Twenty-three dogs (14 females, 9 males; from 4 to 11 years of age) were evaluated annually for total WBC counts, lymphocyte subset distributions, natural killer cell activity and neutrophil phagocytic activity, and biannually for lymphoproliferative activity. An age-related decline in absolute numbers of lymphocytes, T-cells, CD4-cells and CD8-cells was observed in both genders. The distribution of lymphocyte subsets shifted with age, most dramatically in the females; percentages of B-cells declined while those of T-cells increased. Changes in percentages of CD4- and CD8-cells over the 8-year period were not dramatic; in females, percentages of CD8-cells increased significantly in early- to mid-life and then stabilized. Lymphoproliferative responses to mitogens declined over time in both genders. Males demonstrated higher levels of NK cytolytic activity than females; a marginal decline in activity with age was observed. No significant age-related changes in the phagocytic capacity of PMN were observed. These longitudinal findings help to discriminate between those immune parameters which change most dramatically in early-life versus those which either change more dramatically later in life or change gradually over the entire span of life. In addition they identify significant gender differences in several parameters and corroborate our previously published cross-sectional aging data in the same species.

Individual and combined effects of ethanol and cocaine on the human dopamine transporter in neuronal cell lines.

Concurrent use of cocaine and alcohol results in reinforced behavioral consequences, but the molecular mechanisms associated with the co-use of both drugs are not clear. We report here that a 24-h exposure of the human dopamine transporter (hDAT)-transfected mouse neuroblastoma N1E-115 cell line (6C6) to cocaine (1 microM), or ethanol (1%) or both, increased dopamine re-uptake by approximately 25, 29 and 44%, respectively. The same treatment also increased dopamine re-uptake by the hDAT-transfected mouse neuroblastoma Neuro2A cell line (3B7) by approximately 36, 41 and 77%, respectively. However, no increase of dopamine re-uptake was observed in the hDAT-transfected non-neuronal CHO cell line (10E9). These data support the hypothesis that the DAT may be a common neural substrate for cocaine and ethanol in dopaminergic neurons and it may be involved in the psychological effects of both addictions.

Dopamine uptake by mouse neuroblastoma N1E-115 cells stably expressing human dopamine transporter is differentially inhibited by anti-idiotypic ab2beta antibodies mimicking the configuration of cocaine.

We have successfully constructed a mouse neuronal N1E-115 cell line stably expressing a fully functional human dopamine transporter (hDAT). Previous studies in our laboratory have produced several anti-idiotypic Ab2beta antibodies that mimic the configuration of the cocaine molecule at their antigen-combining site. In the present study, we observed that some anti-idiotypic Ab2beta antibodies inhibited dopamine uptake by the hDAT-transfected line. Each antibody showed a different level of inhibition (between 40% and 90%). These findings suggest that the internal images of these antibodies can be used as analog peptides that may compete with cocaine for its binding site on the transporter but not impair dopamine uptake as much as cocaine does.

Exploring the feasibility of an anti-idiotypic cocaine vaccine: analysis of the specificity of anticocaine antibodies (Ab1) capable of inducing Ab2beta anti-idiotypic antibodies.

Conventional vaccination with the cocaine molecule conjugated to a protein carrier is a new approach in the treatment of addiction. Experimentally, this strategy has been shown to alter the pharmacokinetics as well as the psychostimulant effect of a cocaine challenge. The purpose of this study was to investigate whether a more stable and less controversial molecule, an anti-idiotypic antibody, which mimics the configuration of the cocaine molecule (Ab2beta), could be successfully used instead of cocaine. Two cocaine conjugates that presented different areas of the cocaine molecule to the immune system were used to produce monoclonal antibodies specific for cocaine (Ab1). Several anti-idiotypic antibodies were then produced. Four were identified as Ab2beta, or internal images of the antigen; when injected into BALB/c mice, they elicited an anticocaine response. The anticocaine response elicited by one of the four Ab2beta (K1-4c) was sufficient to significantly reduce the level of cocaine that targeted the brain following cocaine challenge, compared with the level of cocaine found in the brain of control animals immunized with irrelevant antibody. In conclusion, the possibility of an anti-idiotypic vaccine seems to be worth pursuing.

Immunological, hematological, and biochemical responses in immature white-footed mice following maternal Aroclor 1254 exposure: A possible bioindicator.

A number of hematological, immunological, and biochemical parameters were measured in Peromyscus leucopus pups born from dams exposed to a single dose (300 mg/kg body weight) of Aroclor 1254. To increase the chances of uncovering even modest consequences of the exposure, in one protocol the pups were weaned at 3 weeks and examined at 6 weeks of age, while in a second protocol the pups were kept with their mother for 4 weeks, at which time they were examined. The older pups showed significant decreases in body weight, ratio of spleen weight to body weight, numbers of peripheral white blood cells and lymphocytes, and number and percentage of monocytes. They also showed significant increases in the stimulation index in response to the mitogen phytohemagglutinin (PHA), percentage of peripheral blood neutrophils and liver EROD induction. Pups sacrificed at 4 weeks of age showed even more significant differences. Their body and liver weights, percentage and number of peripheral blood lymphocytes, and serum antibody titers were significantly lower than those of their controls, while spleen to body weight ratios, percent of neutrophils in their peripheral blood, and liver EROD, PROD, and BROD levels were significantly higher than those of the controls. The primary implication of this work is that white-footed mouse pups could be used as biomonitors of contaminated sites. Females could be captured at the sites and bred in captivity with normal males. The vulnerable parameters identified in this study could then be measured in the resulting offspring and compared with a database collected from normal pups.

Reflections on bioethics: consolidation of the principle of autonomy and legal aspects.

The author highlights the importance of emotions in all ethical reflections. He describes the most common positions of ethicists employing duties and rights as the basis for ethical thought. The author, goes to Freudian theory as viewed by the utilitarians, stating that the 'quest for pleasure' is not necessarily egocentric, especially for adults. For example, the feeling of solidarity emerges 'from the inside out', making irrelevant all the emphasis laid on obedience to duty (from the outside in). The article questions the essence of Kantian theory, based exclusively on 'reason' with disregard for feelings, by establishing what he considers a 'positivist' view of rational thought. It emphasizes the principle of autonomy, which it seen as basically opposing the principles of beneficence and fairness. It is proposed that the latter should be seen as what he calls heteronomy (a concept different from that of the rational ethicists). In theory, autonomy is not assigned to anyone on the basis of an external assessment. Any intervention in individual autonomy must be made (by the intervenor) when it becomes imperative in the defense of social or cultural values. The article distinguishes between ethics and morals) and states that the sole acceptable ethical principle is that ethics (theoretically) has no principle.

Hematology values from clinically healthy Peromyscus leucopus.

Reported herein are hematocrit and total and differential WBCs obtained from 132 clinically healthy male and female white-footed mice (Peromyscus leucopus) of four different age groups: 4, 5-8, 9-20, and >32 wk old. Minimal differences were identified between the age groups. The number and percentage of neutrophils in the 4-wk-old mice were significantly higher than those in the other groups; the relative percentage of lymphocytes in the 4-wk-old mice was significantly lower than in the 9-20-wk-old mice. The only significant gender effects identified were higher numbers of WBCs and lymphocytes in females of the 4-wk-old group and higher hematocrits in males of the 5-8- and >32-wk-old groups.

Antibodies with idiotypic and anti-idiotypic reactivity (epibodies) in conventional immune responses to dinitrophenylated carriers.

A number of monoclonal antibodies were derived from the spleen cells of dinitropheryl (DNP)-immunized mice. Both T-dependent and T-independent carriers were used, and the intensity and length of immunization were varied. It was found that some of the antibodies had only idiotypic (Ab1) reactivity, while others had both idiotypic (Ab1) and anti-idiotypic (Ab2) reactivity. Among the latter antibodies some molecules reacted specifically with DNP and with the combining site of anti-DNP antibodies (epibodies), while others bound DNP and anti-DNP Abs as well as a variety of unrelated antigens (polyreactive antibodies). The proportion of the three types of antibodies (antigen-specific, epibodies and polyreactive antibodies) varied with the nature of the carrier, the intensity of the immunization, and the length of the immunization process. Further characterization of the epibodies, which were predominant in the secondary response to DNP-keyhole limpet haemocyanin (KLH), showed that both Ab1 and Ab2 reactivities were inhibited by both soluble ligands (DNP and anti-DNP), indicating that the specific combining site of the monoclonal antibodies (mAbs) (and/or of the rabbit anti-DNP antibody in the case of Ab2) was involved in both activities. Both Ab1 and Ab2 reactivities were removed by absorption of the mAbs with either immobilized DNP or immobilized rabbit anti-DNP. The mAbs were capable of binding themselves as well as to other mAbs with the same characteristics. The affinity constants of several mAbs for both the DNP and anti-DNP ligands were determined.

[The concept of health]. / O conceito de saúde.

Objections to the present WHO (World Health Organization) definition of HEALTH, as "the state of perfect physical, mental and social well-being", are expressed. It is considered to be anachronistic, first because it aims at perfection which is unaltainelle because of district personality characteristics. As the main support for this idea, the necessary renunciation of part of man's drive to liberty in exchange for the lesser insecurity provided by social life (Freud, Castoriadis and McDougall), is groted. The validity of distinguishing between "soma", "psyche" and "society" is questioned and the concept of the "integrated man", alluding to Pierre Marty and to Freud himself is adapted, and situations are recalled in which the interaction of the three aspects mentioned above is actually evident. Finally, the notion of the quality of life, in accordance with an antipositivistic taken from Bion, point of view, is discussed, and the concept that reality is that of each human being, is adapted. This priority and the proposal to rescue subjectivism which was also observed by Foucault when he studied mental disease, leads to a last criticism of the present definition of health, based exclusively on external, objective evaluations.

The influence of age on the canine immune system.

Immune function was assessed in a group of 47 Labrador Retrievers, ranging in age from 0.8 to 11.5 years, in order to establish baseline data on canine immunosenescence. Natural killer cell activity, lymphocyte subset distributions, antibody production, and mitogen-induced lymphoproliferative responses, all of which have been demonstrated to undergo age-related changes in humans and mice, were chosen as indicators of immune function. Dogs were categorized by age as young (mean 2.4 years), middle-aged (mean 5.8 years), and old (mean 9.1 years). Natural killer cell activity was not affected significantly by age. Lymphocyte subset analysis revealed a significant age-related increase in the percentage of cells staining with a pan T-cell reagent, accompanied by a corresponding increase in the percentage of CD8 cells from youth to middle age. An age-related decrease in the percentage of B-cells was observed concomitant with the increases in T-cell percentages. A gender-related difference in pan T-cell distribution was also observed, with females having a higher percentage than males. Lymphoproliferative responses of both young and middle-aged dogs to the mitogens concanavalin A, phytohemagglutinin, pokeweed mitogen, and staphylococcal enterotoxin B were significantly higher than those of old dogs. In general, the mitogen responses of male dogs were affected more dramatically by age than those of females. A significant age-related decline in in vivo antibody responses to the protein antigen, keyhole limpet hemocyanin, was not observed, although the mean titers of the young dogs were higher than those of the old.

Behavior of the idiotypic network in conventional immune responses. III. Detection and enumeration of cells producing idiotypic and anti-idiotypic antibodies by a spot ELISA technique.

The idiotypic (Ab1) and anti-idiotypic (Ab2) responses of spleen cells of mice immunized with DNP-Ficoll were assayed with the spot ELISA technique in wells coated with DNP-BSA or with affinity-purified rabbit anti-DNP antibodies, respectively. In agreement with results previously obtained with the hemolytic plaque technique, large numbers of Ab1 and Ab2 ELISA spots were found. The cytokinetics of the Ab1 and Ab2 responses were very similar and the peak responses occurred simultaneously. The isotypes of Abs1 and Abs2 were also similar. Both responses were specifically inhibited by soluble DNP-lysine. Similar results were obtained with spleen cells from mice immunized with the T-dependent antigen DNP-KLH and with the T-independent antigen fluorescein-Ficoll. In contrast, no Ab2 response was detected when spleen cells from mice immunized with ovalbumin were assayed, probably because the affinity-purified rabbit anti-ovalbumin antibodies used to coat the wells for the Ab2 assay contained molecules of several specificities, corresponding to the several epitopes of the ovalbumin molecule. The many similarities between the Ab1 and Ab2 responses to the haptens suggest that they reflected two different reactivities of the same antibody molecules.

Effect of time on neonatal immune response to dietary selenium and fat.

The effect of dietary selenium (0.03 mg/kg and 0.31 mg/kg), and fat (5% and 20% corn oil) on antibody formation in the neonatal rat was studied at two different time periods using four diets. The latter were fed to dams throughout pregnancy and lactation and to their pups starting at day 18. A week later, pups received intraperitoneal injections of the test antigen, fluorescein-bovine serum albumin in complete Freund's adjuvant (150 micrograms per 30 g body wt.). On days 7 or 9 post-injection, the pups were sacrificed. The antibody titer, as determined by the quenching of fluorescein by the test sera, and by ELISA, almost doubled from day 7 to day 9 (p < 0.05). There was no significant difference in antibody titers between dietary groups except at day 9 post-injection. Using ELISA, pairwise comparisons indicated that addition of 0.3 mg/kg Se to high fat diet was associated with low antibody titer (p < 0.05).

Behavior of the idiotypic network in conventional immune responses. I. Kinetics of idiotypic and anti-idiotypic antibodies following immunization with T-independent and T-dependent antigens.

A minimal requirement in investigations of the behavior of the idiotypic network during immunization is the ability to quantitate both the idiotypic (Ab1) and anti-idiotypic (Ab2) responses. Quantitation of Ab2 in serum is complicated by the simultaneous presence of Ab1, so that Ab1-Ab2 immune complexes escape detection. In contrast, immune complexes should not complicate the enumeration of Ab2-producing lymphocytes in a hemolytic plaque assay. This study utilizes a procedure that allows detection of Ab2-producing cells in such an assay. The procedure relies upon the insertion of the appropriate antibody (Ab1) into the membrane of indicator SRBC through a covalently attached dipalmitoyl phosphatidylethanolamine (DPPE) tail. When the Ab2 response following murine immunization with DNP-Ficoll was analyzed using such an assay, peak plaque-forming cell (PFC) numbers were found to coincide with peak Ab1 PFC numbers in both the primary and secondary response. In addition, this Ab2 response was found to be T independent. The murine immune response to DNP-HGG demonstrated a peak Ab2 PFC response which followed the peak Ab1 PFC response after both primary and secondary immunization. This Ab2 response appeared to be T dependent. The secondary responses to both DNP-Ficoll and DNP-HGG showed increased levels of Ab2 PFC and decreased levels of Ab1 PFC in comparison to the primary responses to the same antigens, suggesting that immunoregulation may occur within these idiotypic networks.

Behavior of the idiotypic network in conventional immune responses. II. Affinity and heterogeneity of idiotypic and anti-idiotypic antibodies following immunization with T-independent and T-dependent antigens.

The relative affinity and heterogeneity of affinity of idiotypic and anti-idiotypic antibodies in mice immunized with the T-independent antigen DNP-Ficoll and the T-dependent antigen DNP-HGG were measured by a plaque inhibition assay. Idiotypic plaque-forming cells (PFC) were detected by a conventional assay utilizing DNP-coated SRBC. Anti-idiotypic PFC were detected with SRBC coated with affinity-purified anti-DNP antibody of rabbit origin. It was found that both idiotypic and anti-idiotypic antibodies elicited by immunization with the T-independent antigen had lower affinity and were less heterogeneous than the corresponding antibodies originating in mice immunized with the T-dependent antigen. In addition, the affinity and heterogeneity values of the idiotypic antibodies were correlated with the affinity and heterogeneity values of the anti-idiotypic antibodies from the same mice. This finding indicates that idiotypic and anti-idiotypic antibodies mutually regulate each other, thus pointing to internal immunoregulatory effects of the idiotypic network with respect to these parameters.

Radiant heat-induced hyperthermia in mice: in vivo effects on the immune system.

Whole-body hyperthermia (WBH) in mice was induced by 2-4-h exposure to radiant heat resulting in core body temperatures of 38.5-40.4 degrees C, and correlated directly with the magnitude and duration of heat treatment. Two-hour heat treatments in this temperature range did not consistently affect generation of antibody-forming cells in vivo, while 4-h treatments at temperatures greater than or equal to 40 degrees C significantly suppressed the antibody-forming cell response. The capacity of lymphocytes from similarly heated mice to generate antibody-forming cells in vitro was not affected, suggesting that the observed in vivo suppression may be mediated by circulating factors rather than by some heat-induced alteration in the cells themselves. In vivo treatment did not alter T-cell responsiveness to the mitogen Con-A or delayed-type hypersensitivity responses to sheep red blood cells. Quantitative and flow cytometric assessment of splenic and thymic lymphocyte numbers showed that WBH did not alter absolute numbers of lymphocytes but did temporarily change the proportions of lymphocyte subsets. An immediate increase in splenic L3T4+ cells was observed, followed within 18 h by an overall decrease in Lyt 2+ and Thy 1.2+ T-cells. In the thymus the percentages of mature T cells increased. In general, only minimal effects of heat on the immune responses of normal mice could be demonstrated.