Serum TABM produced during anterior chamber-associated immune deviation passively transfers suppression of delayed-type hypersensitivity to primed mice

Injection of soluble protein antigen into the anterior chamber of the eye of primed mice induces anterior chamber-associated immune deviation (ACAID) which is manifested by suppression of delayed-type hypersensitivity (DTH) to the antigen. Recently, we found that ACAID induced in primed mice also results in a rapid rise in serum of soluble T lymphocyte-derived proteins specific for nominal antigen (TABM). Here, we demonstrate that serum TABM induced in primed mice during ACAID will transfer the suppression of DTH to mice primed to the same antigen. Sera from TNP-BSA-primed mice that received an anterior chamber injection of TNP-BSA, but not BSA alone, suppressed the DTH response to TNP when injected into other TNP-BSA-primed mice. Sera absorbed with Sepharose beads conjugated with either anti-TCR C(alpha), anti-TCR C(beta), anti-TABM or TNP-BSA did not contain TNP-specific TABM and did not transfer suppression of DTH. These results suggest that the antigen-specific, TCR C(alphabeta)+ TABM that appear in serum during ACAID are able to confer on or amplify the capacity of sensitized T cells to suppress DTH. We believe this to be the first demonstration of an in vivo immunologic function that is specifically associated with TABM produced in vivo.      

Different modulation by histamine of IL-4 and interferon-gamma (IFN-γ) release according to the phenotype of human Th0, Th1 and Th2 clones

Histamine, an important inflammatory mediator in allergic diseases and asthma, has been reported to have modulator effects on T cells, suggesting that the bronchial microenvironment may regulate the function of resident T cells. We examined the effect of histamine on the release of the Th2-associated cytokines IL-4 and IL-5 and the Th1-associated cytokine IFN-γ by 30 CD4⁺ T cell clones from peripheral blood or bronchial biopsy of one atopic subject. Based on the IL-4/IFN-γ ratio, the clones were ascribed to the Th2 (ratio >1), Th0 (ratio 0.1 and 1) or Th1 (ratio <0.1) phenotype. Histamine inhibited IFN-γ production by Th1-like cells (P<0.02, Kruskall–Wallis), especially from bronchial biopsy, but had no effect on IL-4 release. Regarding Th0 clones, histamine inhibited IL-4 production (P<0.02) in a dose-dependent manner and slightly inhibited IFN-γ production, but had no effect on Th2-like cells. Histamine had a heterogeneous and insignificant effect on IL-5 production. The H₂-receptor antagonist ranitidine completely reversed the inhibition of IL-4 and IFN-γ production, whereas the agonist dimaprit mimicked this effect. In contrast, H₁- and H₃-receptor agonists and antagonists had no significant effect. These data demonstrate that histamine has different effects on IL-4 and IFN-γ release by T helper cells according to their phenotype via H₂-receptors. This study extends the immunomodulatory effects of histamine which may contribute to the perpetuation of airway inflammation in asthma.     

Teaching spiritual care – a grounded theory study among undergraduate nursing educators

Aims and objectives. To explore teachers’ understanding of spirituality and how they prepare undergraduate nursing students to recognise spiritual cues and learn to assess and provide spiritual care. Background. Nursing education addresses patient care in all domains of the person. Systematic teaching and supervision of students to prepare them to assist patients spiritually is an important part of holistic care. However, few role models for spiritual care are seen in clinical practice, and limited research addresses necessary student competencies or how teachers can best facilitate this process. Design. Grounded theory was used to identify teachers’ main concern and develop a substantive grounded theory. Methods. Data collected during semi‐structured interviews at three Norwegian University Colleges in five focus groups with 19 undergraduate nursing teachers were conducted from 2008 to 2009. Data were analysed through constant comparison of transcribed interviews until categories emerged and were saturated. Results. The participants’ main concern was ‘How to help students recognize cues and ways of providing spiritual care’. Participants resolved this by ‘Journeying with Students through their Maturation’. This basic social process has three iterative phases that develop throughout the nursing programme: ‘Raising Student awareness to Recognize the Essence of Spirituality’, ‘Assisting Students to Overcome Personal Barriers’, and ‘Mentoring Students’ Competency in Spiritual Care’. Conclusion. Nursing education should prepare students to recognise and act on spiritual cues. Making spiritual assessment and interventions more visible and explicit throughout nursing programmes, in both classroom and clinical settings, will facilitate student maturation as they learn to integrate theoretical thinking into clinical practice. Relevance to clinical practice. Nursing students need role models who demonstrate spiritual care in the fast‐paced hospital environment as well as in other clinical practice settings. To model spirituality as part of nursing care can assist students to overcome their vulnerability and to safeguard ethical issues and promote patient integrity.     

Fasting induces a large, leptin-dependent increase in the intrinsic action potential frequency of orexigenic arcuate nucleus neuropeptide Y/Agouti-related protein neurons

The neuropeptide Y (NPY)/Agouti-related protein (AgRP) neurons of the hypothalamic arcuate nucleus are thought to promote feeding. Here, we demonstrate that feeding state in vivo, through a leptin-dependent process, induces large and persistent changes in the electrophysiological activity of these neurons as measured extracellularly in vitro. Consistent with an orexigenic role, fasting induced a 4-fold increase in the basal action potential frequency of NPY/AgRP neurons. Leptin, when injected into fasted wild-type mice, induced a dose- and time-dependent decrease in spike frequency, which approached fed levels 2-3 h post treatment. In leptin-deficient (lep(ob)/lep(ob)) and leptin receptor-deficient (lepr(db)/lepr(db)) mice, NPY/AgRP spike frequency was not significantly increased by fasting, and even in mutant mice fed ad libitum, spike frequency was at least as high as in fasted wild-type mice. All recordings included GABA(A) and ionotropic glutamate receptor antagonists, suggesting that expression of this modulation is potentially intrinsic and not synaptically dependent. Recorded neurons were unambiguously identified using NPY-Sapphire transgenic mice. This is a remarkably straightforward example of a very robust in vitro electrophysiogical effect produced by a simple behavioral manipulation, food restriction.     

Disposition of cocaine and its metabolites in human sweat after controlled cocaine administration

BACKGROUND: Sweat testing is a noninvasive technique for monitoring drug exposure in treatment, criminal justice, and employment settings. METHODS: We evaluated cocaine excretion in 9 participants' sweat after they received 3 low doses (75 mg/70 kg) of cocaine HCl subcutaneously within 1 week and, 3 weeks later, 3 high doses (150 mg/70 kg). Six additional participants completed portions of the study. PharmChek sweat patches (n = 1390) were collected throughout a 3-week washout period, reflecting previously self-administered drugs, and during and after controlled dosing. RESULTS: Cocaine was the primary analyte detected with 24% of patches positive at the gas chromatography-mass spectrometry limit of quantification of 2.5 ng/patch and 7% of patches at the proposed Substance Abuse and Mental Health Services Administration cutoff of 25 ng/patch. Ecgonine methyl ester (EME) was detected more often and at generally higher concentrations than benzoylecgonine. In patches containing both metabolites, there was no statistically significant difference in the benzoylecgonine/EME ratio based on length of patch wear. During washout, 2 participants' weekly patches tested positive (> or =25 ng/patch) during the first week; one remained positive during week 2; and none were positive during week 3. Cocaine and EME were detectable within 2 h; benzoylecgonine was not detected until 4-8 h after low doses and slightly sooner after high doses. The majority of drug was excreted within 24 h. Over 70% of weekly patches worn during low doses were positive for cocaine (> or =25 ng/patch), increasing to 100% during high doses. CONCLUSION: Sweat testing is an effective and reliable method of monitoring cocaine exposure.     

Long-term evaluation of T-cell subset changes after effective combination antiretroviral therapy during asymptomatic HIV-infection.

Demonstration of long-lived HIV-reservoirs resistant to the effects of combination antiretroviral therapy raises concern over the ability of treatment to maintain long-term beneficial alterations in T-cell subset composition. To address this issue, we have examined the effect of antiretroviral therapy on T-cell subset change during early HIV-infection in a 2-year prospective open-label trial composed of treatment-naive asymptomatic HIV-infected patients with CD4+ T-cell counts > or =400 cells/microl. Therapy consisted of double (zidovudine and lamivudine) or triple (zidovudine, lamivudine, and ritonavir) combination antiretroviral therapy. Retrospective analysis based on magnitude of viral suppression was used to characterize responder and nonresponder groups. Among responders, long-term antiretroviral therapy maintained a significant increase in numbers of total CD4+, naive CD4+/CD45RA+, and memory CD4+/CD45RO+ T cells. A concomitant significant decrease in numbers of memory CD8+/CD45RO+ and both activated CD8+/HLA-DR+ and CD8+/CD38+ T cells was also maintained. In contrast, long-term antiretroviral therapy among nonresponders led only to a significant increase in the numbers of CD4+ T cells and a significant reduction in numbers of activated CD8+/HLA-DR+ T cells. The long-term ability of antiretroviral therapy during early asymptomatic HIV-infection to maintain reversal of disease-induced T-cell activation and maturation abnormalities continues to support the concept that immunologic advantage is gained by commencing early aggressive antiretroviral therapy. Nevertheless, continued management of T-cell subset recovery is significantly more effective in the presence of completely suppressed viral replication.     

A descending dopamine pathway conserved from basal vertebrates to mammals

Dopamine neurons are classically known to modulate locomotion indirectly through ascending projections to the basal ganglia that project down to brainstem locomotor networks. Their loss in Parkinson’s disease is devastating. In lampreys, we recently showed that brainstem networks also receive direct descending dopaminergic inputs that potentiate locomotor output. Here, we provide evidence that this descending dopaminergic pathway is conserved to higher vertebrates, including mammals. In salamanders, dopamine neurons projecting to the striatum or brainstem locomotor networks were partly intermingled. Stimulation of the dopaminergic region evoked dopamine release in brainstem locomotor networks and concurrent reticulospinal activity. In rats, some dopamine neurons projecting to the striatum also innervated the pedunculopontine nucleus, a known locomotor center, and stimulation of the dopaminergic region evoked pedunculopontine dopamine release in vivo. Finally, we found dopaminergic fibers in the human pedunculopontine nucleus. The conservation of a descending dopaminergic pathway across vertebrates warrants re-evaluating dopamine’s role in locomotion.Dopaminergic neurons represent a vital neuromodulatory component essential for vertebrate motor control, and their loss in neurodegenerative disease is devastating. The meso-diencephalic dopamine (DA) neurons are known to provide ascending projections to the basal ganglia, which, in turn, provide input to cortical structure in mammals but also project caudally to the mesencephalic locomotor region (MLR), a highly conserved structure that controls locomotion in all vertebrates investigated to date (1–7; for review, see ref. 8). A growing body of evidence supports the view that basal ganglia connectivity is highly conserved among vertebrates, from lampreys to mammals (9–11; for review, see ref. 12), with some interspecies differences recently highlighted (13). As such, the homology between DA cell populations remains to be resolved in vertebrates. As a general rule, DA neurons from the meso-diencephalon send projections to the striatum in all vertebrates. In lampreys and teleosts, those neurons are located only in the diencephalon (posterior tuberculum), but in tetrapods and cartilaginous fishes (14) they are located in both the diencephalon and the mesencephalon. An increasing number of authors seem to agree with the hypothesis that at least some of the meso-diencephalic DA neurons located in the diencephalon are homologous in all vertebrates, and thus, homologous to at least a portion of the mammalian substantia nigra pars compacta (SNc)/ventral tegmental area (VTA) (13, 15–19; for review, see ref. 20). Alternatively, it was suggested that the posterior tuberculum DA neurons projecting to the striatum in zebrafish are homologs of the mammalian DA neurons of the A11 group (21). This will be discussed below in light of the results of the present study.In lampreys, only a few meso-diencephalic DA neurons send ascending projections to the striatum (9, 22); the majority of DA neurons send a direct descending projection to the MLR (22, 23), where DA is released and increases locomotor output through D1 receptors (22). These results demonstrate that the descending dopaminergic pathway to the MLR is an important modulator of locomotor output, but it remains to be determined whether this pathway is conserved in higher vertebrates.The existence of a descending dopaminergic pathway that powerfully increases locomotor output has important implications for Parkinson’s disease, which involves the meso-diencephalic DA neurons. A loss of descending dopaminergic projections could play a role in the locomotor deficits systematically observed in that disease. Because of the highly conserved nature of both the dopaminergic system and brainstem locomotor circuitry in vertebrates, we hypothesized that a direct descending dopaminergic pathway to the MLR also exists in higher vertebrates. Previous anatomical (24, 25) and electrophysiological (26) studies in rats support the idea of a descending connection from the SNc to the pedunculopontine nucleus [PPN, considered part of the MLR (2)]. Moreover, dopaminergic terminals were found in the PPN of monkeys (27), but the origin of this projection is still unknown in mammals.Here, we investigated whether the direct descending projection from meso-diencephalic DA neurons to the MLR is present in two tetrapods, the salamander and the rat. Moreover, we supplement our analyses with anatomical data from human brain tissue. Using traditional and virogenetic axonal tracing, immunofluorescence, in vivo voltammetry, and calcium imaging of reticulospinal neurons, we provide anatomical and functional evidence strongly supporting a conserved role for the descending projections of meso-diencephalic DA neurons in the regulation of brainstem locomotor networks across the vertebrate subphylum.     

Thymocytes induced by antigen injection into the anterior chamber activate splenic CD8+ suppressor cells and enhance the antigen-induced production of immunoglobulin G1 antibodies

Injection of antigen into the ocular anterior chamber (AC) of a mouse eye (an immunologically privileged site) induces the activation of immunoregulatory NK1.1+, CD4- CD8-, T-cell receptor (TCR) alphabeta+ thymocytes. These thymocytes transfer the suppression of delayed-type hypersensitivity (DTH) when injected into mice sensitized to the same antigen but do not effect the suppression of DTH. On the other hand, the immunized recipients of these transferred thymocytes produce splenic CD8+ T cells that effect the suppression of DTH. However, it is unclear whether the thymocytes transferred from the AC-injected donor differentiate into and/or activate CD8+ T-splenic suppressor cells. We therefore sought to determine the origin of splenic suppressor cells produced in the recipients of immunoregulatory thymocytes transferred from donors that receive an injection of antigen into the AC. CD45.1+ thymocytes from mice that received an AC injection of 2,4,6-trinitrobenzene sulphonic acid (TNP)-bovine serum albumin (BSA) were transferred to congenic CD45.2+ TNP-BSA-immune recipients. Spleen cells from the recipients were then sorted based on anti-CD45.1 or -CD45.2 antibody binding and assayed for suppressor cells. This was done by the injection of separated spleen cells into the footpad of TNP-BSA-immunized mice, concurrent with the induction of footpad swelling (contact sensitivity) of the footpad elicited by an epicutaneous application of picryl chloride. The systemic distribution of antigen after the injection of antigen into the AC was demonstrated by the injection of fluorescein or 125I-labelled TNP-BSA into the AC. The results demonstrate that (i) splenic CD8+ T-suppressor cells produced in the immunized recipients of immunoregulatory thymocytes are derived from the CD45.2 recipient of the CD45.1+ thymocytes; (ii) the induction of recipient splenic suppressor T cells by the transferred immunoregulatory thymocytes requires that the recipient be immunized to the same antigen as that used to induce immunoregulatory thymocytes; (iii) antigen is introduced to the thymus after an injection of antigen into the AC; (iv) although the transfer of the suppression of DTH by regulatory thymocytes was not dependent on interleukin-4 (IL-4), CD4+ NK1.1- regulatory thymocytes from AC-injected donors enhanced the production of immunoglobulin G1 antibodies to TNP-BSA by an IL-4-dependent mechanism. These observations suggest that the adult thymus plays an active role in the induction and maintenance of anterior chamber-associated immune deviation as manifested by the generation of the suppression of cell-mediated immunity to exogenous antigen and the antigen-induced production of IgG1 antibodies.     

Prehospital Care and New Models of Regionalization

This article summarizes the discussions of the emergency medical services (EMS) breakout session at the June 2010 Academic Emergency Medicine consensus conference “Beyond Regionalization: Integrated Networks of Emergency Care.” The group focused on prehospital issues such as the identification of patients by EMS personnel, protocol‐driven destination selection, bypassing closer nondesignated centers to transport patients directly to more distant designated specialty centers, and the modes of transport to be used as they relate to the regionalization of emergency care. It is our hope that the proposed research agenda will be advanced in a way that begins to rigorously approach the unanswered research questions and that these answers, in turn, will lead to an evidence‐based, cohesive, comprehensive, and more uniform set of guidelines that govern the delivery and practice of prehospital emergency care. ACADEMIC EMERGENCY MEDICINE 2010; 17:1337–1345 © 2010 by the Society for Academic Emergency Medicine     

Developing research criteria to define medical necessity in emergency medical services

“The Neely Conference: Developing Research Criteria to Define Medical Necessity in EMS” convened emergency medical services (EMS) physicians, researchers, administrators, providers, and federal agency representatives to begin the development of a set of uniform triage criteria and outcome measures that could be used to study and evaluate medical necessity among EMS patients. These standardized criteria might be used in research studies examining EMS dispatch and response (e.g., dispatch triage protocols, alternative response configurations), and EMS treatment and transport (e.g., field triage protocols, alternative care destinations). The conference process included review and analysis of the literature, expert judgment, and consensus building. There was general agreement on the following: 1. Any dispatch triage or field triage system that is developed must be designed to offer patients alternatives to EMS, not to refuse care to patients. 2. It is theoretically possible to develop a set of clinical criteria for need. Some groups of patients will clearly need a traditional EMS response and other groups will not, but this has yet to be defined. 3. In addition to clinical criteria, certain social and other nonclinical criteria such as pain or potential abuse may be used to justify a response. 4. Communication barriers, patient age, special needs, and other conditions complicate patient assessment but should not exclude patients from consideration for alternate triage or transport. 5. These research questions are important, and standard sets of outcome measures are needed so that different studies and innovative programs can be compared.