Longitudinal study of pediatric house officers' attitudes toward death and dying.

OBJECTIVE: To investigate pediatric residents' attitudes toward end-of-life issues and their education in dealing with these issues. DESIGN: Exploratory survey. SETTING: Department of Pediatrics at the University of California, Los Angeles, Center for Health Sciences. SUBJECTS: Volunteer sample. A total of 182 of 203 pediatric residents at all levels of training completed anonymous questionnaires. INTERVENTIONS: None. MEASUREMENTS AND MAIN RESULTS: Data on residents' attitudes toward issues of death and dying and the efficacy of educational interventions were collected over a 4-yr period. When entering training, house officers are uncomfortable dealing with death and dying issues (mean, 3.3 of 5; 5 = not comfortable). By the end of their training, these house officers become comfortable dealing with these issues (mean, 2.2; p < .05). During their first 2 yrs of training, house officers report that their medical education is not helping them to deal with the issues of death and dying (mean, 3.3). At the end of their third year of training, residents report that their education is helping them to deal with these issues (mean, 2.5; p < .05). Strikingly, as house officers progress through their residency, they become less comfortable with the idea of administering pain medication to a dying patient, because the pain medication might hasten the patient's death (p < .05). CONCLUSIONS: Pediatric residents may benefit from more formal training in the practical aspects of death and dying issues. Residency education should do more to address these issues systematically for the benefit of both the residents and the patients and family members.

Searching for depolarization-induced genes that modulate synaptic plasticity and neurotrophin-induced genes that mediate neuronal differentiation.

We identify and characterize two classes of immediate-early genes: (i) genes, induced by depolarization in neurons, that play a role in depolarization-induced neuronal plasticity and (ii) genes, induced in neuronal precursors by neurotrophins, that play a causal role in neurotrophin-directed neuronal differentiation. We use rat PC12 pheochromocytoma cells to identify (i) genes preferentially induced by [depolarization or forskolin] versus [Nerve Growth Factor (NGF) or Epidermal Growth Factor (EGF)] and (ii) genes preferentially induced by NGF versus EGF. We describe (i) a collection of genes preferentially induced by depolarization/forskolin in PC12 cells and by kainic acid in vivo, and (ii) a collection of genes preferentially induced by NGF. The synaptotagmin IV gene encodes a synaptic vesicle protein whose level is modulated by depolarization. NGF preferentially induces the urokinase-plasminogen activator receptor in PC12 cells. Antisense oligonucleotide and anti-UPAR antibody experiments demonstrate that NGF-induced UPAR expression is required for NGF-driven PC12 cell differentiation.

rTLE3, a newly identified transducin-like enhancer of split, is induced by depolarization in brain.

The transducin-like enhancers of split are a family of mammalian proteins that share sequence homology with the Drosophila protein Groucho. Using representational difference analysis, we isolated the cDNA for a previously unidentified gene, rTLE3 (rat transducin-like enhancer of split 3), as a sequence induced by depolarization and forskolin, but not by neurotrophins or growth factors, in PC12 pheochromocytoma cells. rTLE3 encodes the protein rTLE3, a 764-amino acid orthologue of mouse and human TLE3. R-esp2, the gene encoding the closest related rat protein, is not induced by any of the four treatments in PC12 cells. rTLE3 and R-esp2 have different patterns of expression in the adult rat CNS and other tissues. After systemic administration of kainic acid, rTLE3 is induced specifically in the dentate gyrus of the hippocampus. We propose that members of the transducin-like enhancer of split family of proteins may have distinct functions in the mature CNS, in addition to their functions during development.

The salt-inducible kinase, SIK, is induced by depolarization in brain.

Membrane depolarization of neurons is thought to lead to changes in gene expression that modulate neuronal plasticity. We used representational difference analysis to identify a group of cDNAs that are induced by membrane depolarization or by forskolin, but not by neurotrophins or growth factors, in PC12 pheochromocytoma cells. One of these genes, SIK (salt-inducible kinase), is a member of the sucrose-nonfermenting 1 protein kinase/AMP-activated protein kinase protein kinase family that was also recently identified from the adrenal gland of rats treated with high-salt diets. SIK mRNA is induced up to eightfold in specific regions of the hippocampus and cortex in rats, following systemic kainic acid administration and seizure induction.

Clinical use of continuous arterial blood gas monitoring in the pediatric intensive care unit.

CONTEXT: Continuous arterial blood gas monitoring is a new technology based on the combination of opto-chemical and fiber-optic detectors that can measure pH, PCO2, PO2, and temperature on a continuous basis via a sensor placed in an artery. OBJECTIVE: To evaluate this technology in pediatric patients who would normally require frequent arterial blood gas sampling. DESIGN: A criterion standard study in which the results of arterial blood gas samples measured by the laboratory analyzer were compared with the sensor readings. SETTING: A pediatric intensive care unit of a tertiary referral center. PATIENTS: Children with severe respiratory failure who required frequent arterial blood gas sampling and who had a 20-gauge arterial line in either a radial or femoral site. RESULTS: Twenty-four patients with a mean age of 6.4 years (range 1-21 years) had a sensor placed. Sensors were in place for a mean of 101 +/- 62 hours. Eighteen patients underwent continuous monitoring for at least 24 hours or until no longer clinically necessary. There were 414 pairs of blood gas samples obtained. The bias/precision for pH was 0.005/0.030; for PCO2, -1.8/6.3 mm Hg; and for PO2, 1.2/24 mm Hg. The correlation (r value) between the sensor readings and the blood gases were pH 0. 960, PCO2 0.927, PO2 0.813 (P <.01 for all values). The bias and precision for PO2 levels < 70 mm Hg were 0.057/9.34 mm Hg. There were no complications from sensor placement. Continuous blood gas monitoring allowed immediate recognition of clinical changes. CONCLUSION: The continuous arterial blood gas sensor is capable of clinically accurate blood gas measurements. This technology provides the clinician with immediate data that can allow rapid interventions in unstable patients.

Nurr1 mRNA expression in neonatal and adult rat brain following kainic acid-induced seizure activity.

Nurr1 is an immediate early gene encoding a member of the steroid-thyroid hormone receptor family. In PC12 cells, Nurr1 is readily induced by membrane depolarization, but not by growth factors. Nurr1 is predominantly expressed in the brain, and is essential to the differentiation of midbrain dopaminergic neurons. However, Nurr1 is also expressed in brain regions unrelated to dopaminergic neurons, e.g., hippocampus and cerebral cortex, and its immediate induction following seizure activity suggests a potential involvement of this transcription factor in modulating gene expression in the nervous system. To investigate the response of Nurr1 to neuronal activation, we analyzed Nurr1 mRNA expression in neonatal and adult rat brain following kainic acid (KA)-induced seizure. In P7 animals, systemic injection of KA increased Nurr1 mRNA levels in a few hilar cells of the dentate gyrus and some pyramidal cells of the CA3 region of the hippocampus. In older animals, Nurr1 induction progressively expanded to all hippocampal regions (P14, P21) and eventually to cortical regions (adult). The increase was rapid and transient in the dentate gyrus, a structure resistant to the neurotoxic effect of KA, and was more prolonged in other regions more susceptible to KA toxicity. Induction of Nurr1 at early postnatal stages and rapid increase in the dentate gyrus following KA-induced seizure, suggest that Nurr1 expression is modulated by neuronal activity. On the other hand, prolonged Nurr1 induction in regions sensitive to KA toxicity indicates a possible involvement of Nurr1 in selective neuronal vulnerability.

Seizure activity induces PIM-1 expression in brain.

We recently identified KID-1, a previously undescribed protein kinase induced by depolarization in PC12 cells and brain (Feldman et al., 1998). KID-1 shares a high degree of sequence homology with PIM-1, a proto-oncogene previously reported to be expressed in hematopoietic and germ cells. We examined PIM-1 expression in stimulated PC12 cells, brains of kainic acid-treated rats, and a number of tissues from untreated rats. We now report that forskolin, but not depolarization or growth factors, induces PIM-1 expression in PC12 cells. PIM-1 is an immediate early gene induced in response to forskolin stimulation. We detect PIM-1 mRNA in a number of unstimulated tissues and at low levels in unstimulated brain. Systemic kainic acid administration to adult rats induces PIM-1 expression in the dentate gyrus region of the hippocampus.

KID-1, a protein kinase induced by depolarization in brain.

Membrane depolarization leads to changes in gene expression that modulate neuronal plasticity. Using representational difference analysis, we have identified a previously undiscovered cDNA, KID-1 (kinase induced by depolarization), that is induced by membrane depolarization or forskolin, but not by neurotrophins or growth factors, in PC12 pheochromocytoma cells. KID-1 is an immediate early gene that shares a high degree of sequence similarity with the family of PIM-1 serine/threonine protein kinases. Recombinant KID-1 fusion protein is able to catalyze both histone phosphorylation and autophosphorylation. KID-1 mRNA is present in a number of unstimulated tissues, including brain. In response to kainic acid and electroconvulsive shock-induced seizures, KID-1 is induced in specific regions of the hippocampus and cortex.

Late development of renal carcinoma in allograft kidney.

The Cincinnati Transplant Tumor Registry recorded 169 cases of renal carcinoma developing in transplant recipients. The great majority of these cases were of primary renal cell carcinoma developing in the recipient native kidneys. Renal carcinoma developing de novo in the renal allograft occurred 17 times, with a maximal interval to clinical development of 85 months after transplantation. The development of multicentric renal cell carcinoma in an allograft 156 months after transplantation is described. The 24-year-old white male recipient with Alport's syndrome received a cadaver renal allograft from a healthy 27-year-old black man who had died of a cerebral hemorrhage in 1977. At 13 years after transplantation the recipient had upper abdominal pain. Ultrasound revealed 2 incidental renal masses and a renal cyst in the allograft. Partial nephrectomy confirmed the presence of multicentric renal carcinoma. The graft was left in situ and immunosuppression was maintained. The recipient continued to do well with no evidence of disease 1 year postoperatively. Deoxyribonucleic acid banding demonstrated that the tumor and recipient blood were of different patterns.

Developmental changes in neuromuscular transmission in the rat diaphragm.

Neuromuscular transmission was studied in diaphragms from rats of three ages, 4-7 days old, 11-12 days old, and adults with the use of an in vitro phrenic nerve-hemidiaphragm preparation. Each hemidiaphragm was stimulated via either muscle or nerve with 1-s stimulus trains at frequencies from 10 to 100 Hz. The patterns of force development obtained in response to the two routes of stimulation were compared for each group. Diaphragms from adults developed maximum force in response to stimulation of approximately 40 Hz with no significant decrease in force at higher frequencies. Within each stimulus train, once peak force was achieved, it was maintained for the remainder of the stimulus and responses to nerve and muscle stimulation were almost identical. In contrast, diaphragms from 4- to 7-day-old rats developed maximum force at approximately 20 Hz; stimulation at greater than or equal to 60 Hz induced significantly less peak force. This decrease in peak force at higher frequencies was significantly larger for nerve than for muscle stimulation. In addition, during each nerve stimulus train diaphragms from 4- to 7-day-old rats were unable to maintain peak force, which decreased at frequencies greater than 20 Hz. The decrease in force reached approximately 50% of peak at stimulation frequencies greater than or equal to 60 Hz. Diaphragms from 11- to 12-day-old rats showed intermediate responses. Based on the responses to phrenic nerve stimulation, we conclude that the neonatal rat diaphragm shows marked neuromuscular transmission failure that is not seen in the adult.(ABSTRACT TRUNCATED AT 250 WORDS)

IgE-mediated chemotaxis of rat basophilic leukemia cells towards specific antigen.

We evaluated chemotactic properties of four sublines of rat basophilic leukemia cells using blindwell Boyden chamber assays. After sensitization with a mouse monoclonal IgE directed against dinitrophenyl (DNP), cells from sublines 2H3-C and 926a underwent chemotaxis toward DNP-bovine serum albumin (BSA) and sublines RBL-1 and 4A did not. Chemotactic responses required specific IgE and were determined by the IgE antigen specificity used for sensitization. The threshold for chemotaxis was on the order of 10(-10) M DNP-BSA. Release of incorporated [3H]-serotonin did not always parallel chemotactic responses, which suggests that chemotaxis and secretion may be two unlinked processes that occur during basophil activation. Our results predict a possible in vivo mechanism whereby specific chemotactic responses of basophils and other FcR epsilon-bearing cells are mediated via specific IgE bound to membrane FcR epsilon.

Effects of aging on cell cooperation and lymphocyte responsiveness to cytokines.

Functional activities and cell cooperation of macrophages (Mphi), T cells, and B cells of young and old Lewis rats were compared. Splenic M phi from young and old rats provided accessory help for T cell mitogenesis and B cell mitogenesis, provided accessory help for generation of PFC, and produced IL 1 equally well as measured in costimulator assays. Splenic T cells of aged Lewis rats, however, were poorly responsive in mitogen assays and did not respond to supplemental IL 2 and antigen with blast transformation and with increased help for B cells to produce PFC. "Old" B cells did not respond in vitro to mitogens with help from M phi and T cells, nor did they respond to B cell helper factor with increased PFC. The data indicate that hyporesponsiveness of the immune system, especially of B cells, in aged rats is due in part to defective reactivity to interleukins and cytokine(s) and to defective cell-cell cooperation.

Inhibition of interleukin synthesis and T cell proliferation by a monoclonal anti-Ia antibody.

The proliferative response in vitro of Lewis rat spleen cells to Con A was inhibited by a monoclonal antibody, OX4, with specificity for a public rat la antigen. Inhibition was dose-dependent and was observed only if the antibody was present during the first 48 hr of culture. OX4 anti-la antibody inhibited production of IL 1 by peritoneal exudate cells activated by lipopolysaccharide and inhibited the production of IL 2 by spleen cells activated by Con A. Addition of exogenous IL 1 to spleen cell cultures inhibited by OX4 anti-la antibody augmented production of IL 2 and returned the proliferative response of T cells to levels observed in Con A-stimulated cultures. We suggest that OX4 anti-la antibody reacts with la+ accessory cells to inhibit production of IL 1, which in turn limits production of IL 2 and finally T cell proliferation.

Development of the orientation of the visuo-tectal map in Xenopus.

Eye rudiments from Xenopus embryos of stage 28 or younger were explanted on to the flank of similar embryos with normal or nasotemporally reversed orientation. After some hours the eyes were retransplanted to an orbit of a stage 32/34 embryo, with either normal or reversed nasotemporal orientation. Later the visuotectal projections through the operated eyes were mapped electrophysiologically. The maps obtained were oriented as if the mapping orientation of the eye had already been determined in the donor orbit before the first transplantation; i.e. if normally oriented in the final host orbit the eye gave a normal map, and if the eye was reversed in the final host orbit it gave a reversed map. In only one out of 25 cases did it seem that the orientation of the map could have been influenced by the orientation of the eye on the flank of the intermediate host, and here the evidence was weak. Two eyes gave reduplicated maps and 4 maps were uninterpretable. It was concluded that the orientation of the map is determined before stage 26 and is not altered by information derived from the flank during stages 26-34.

Membrane phenotype of the rat cytotoxic T lymphocyte.

We have examined rat cytotoxic T lymphocytes for expression of W3/25, OX8, Ia, Thy-1 antigens, and Fc gamma receptors using an effector cell-target cell conjugate formation assay in conjunction with immunofluorescence techniques. Lymph node, spleen, and peritoneal exudate T cells from Lewis rats immunized with allogeneic BN tumor cells specifically bound to and lysed BN tumor targets and BN blast cells, but did not bind or lyse syngeneic Lewis sarcoma cells, Lewis blast cells, or Lou/M blast cells. The numbers of binding and cytotoxic T lymphocytes were greatest in peritoneal exudate cells of immunized rats, less in spleens, and least in lymph nodes. Seventy to 80% of the lymphocytes bound to tumor targets were OX8+ T lymphocytes; less than 12% expressed W3/25, Ia, Thy-1, or Fc gamma R. Moreover, only OX8+ T cells efficiently lysed the target cells to which they were bound. The membrane phenotype of rat cytotoxic T lymphocytes was: OX8+, W3/25-, Ia-, Thy-1, and Fc gamma R-. Monoclonal OX8 antibody did not inhibit target cell binding or subsequent lysis by effector T cells, and there was no diminution of target cell binding or cytotoxic activity when the OX8 antigen was shed from the cell surface before interaction with target cells. There was no preferential association of OX8 antigen at the interface between the effector and target cell. Thus, OX8 antigen marks a subset of rat T lymphocytes that are cytotoxic but the molecule appears not to play a functional role in the cytotoxic process.

Regulation of rat B cell responses by T cell subsets and interleukin 2.

T cell and interleukin 2 (IL 2) regulation of two phases of B cell activation, 3H-TdR uptake and differentiation to antibody-forming cells, were examined. Monoclonal antibody specific for rat kappa light chains (Mab alpha K) stimulated 3H-TdR uptake in vitro in B cell fractions of Lewis spleen cell populations in the presence of T cells and macrophages (M phi); IL 2 reconstituted the response in enriched B cell cultures depleted of T cells and M phi. When IL 2 was added to primary in vitro cultures, spleen cells yielded 100 to 400 anti-SRBC PFC/culture; no PFC were recorded in the absence of IL 2. These results suggested that IL 2 served as a " second signal" for activation in responsive B cell populations. When monoclonal antibody specific for the W3/25+ T cell subset (Mab W3/25) was incorporated into the assay system, both 3H-TdR uptake and PFC responses were inhibited. IL 2 enhancement of B cell responses or responses of reconstituted B cell and T cell fractions was eliminated in the presence of Mab W3/25, indicating that IL 2 mediation of B cell responses was due in part to participation of W3/25+ T cell helper function. In contrast, monoclonal antibody directed to the OX8-bearing T cells (Mab OX8) had no effect on B cell responses. W3/25+ T cells provided helper activity in the generation of a PFC response, whereas OX8+ cells suppressed the antibody response. W3/25+ T cells responded to antigenic stimuli in the presence of IL 2 by undergoing increased blast transformation. OX8+ cells did not exhibit any response. These data define a regulatory network by which T cells, IL 2, and B cells interact to produce in vitro DNA synthesis and antibody formation in activated rat B cells.

Altered levels of mononuclear leukocytes in tumor-bearing rats: decrease of helper T lymphocytes and increase of suppressor cells.

In the spleen and peripheral blood of BN rats with progressive tumors, W3/25+ T helper cells were significantly reduced and OX8+ T suppressor/cytotoxic cells were significantly increased. The ratio of helper to suppressor elements was decreased to 1.6 from a ratio of 3 in normal BN rats without tumors, and this decreased ratio correlated with tumor growth. When tumors were eliminated in vivo by infusion of effector cells (W3/25+ T lymphocytes), the levels of W3/25+ and OX8+ T cells returned to normal and the ratio of helper to suppressor/cytotoxic cells in the spleen and peripheral blood reverted to 3.0 or higher. Macrophages and null cells, T-sIg-, were also elevated in the spleen and peripheral blood of rats bearing expanding tumors and returned to normal levels after cure. Assays of spleen cells for cell-mediated cytotoxicity in rats with large tumors revealed little or no specific cytotoxicity. Cytotoxic activity was high in spleen of rats cured of their neoplasms by infusion of helper cells.

beta-Endorphin enhances lymphocyte proliferative responses.

The opioid peptides alpha- and beta-endorphin and [D-Ala2, Met5]enkephalin were investigated for their effect on the proliferation of resting and activated rat splenic lymphocytes in vitro. beta-Endorphin enhanced the proliferative response of spleen cells to the T cell mitogens concanavalin A and phytohemagglutinin. The effect of beta-endorphin was dose dependent and occurred at peptide concentrations similar to those found in rat plasma. alpha-Endorphin and [D-Ala2, Met5]enkephalin did not affect the proliferative responses to any mitogen tested. Furthermore, the potentiating effect of beta-endorphin was not reversed by treatment with 10 microM naloxone. None of the peptides had any effect on resting, unstimulated spleen cells or on the response to a mixture of lipopolysaccharide and dextran sulfate, which is specifically mitogenic for B lymphocytes. The pharmacological properties of the beta-endorphin potentiation indicate that the effect may be mediated by a nonopiate but beta-endorphin-specific mechanism. These results suggest a possible role for peripheral beta-endorphin and may provide a link between stress and disease susceptibility.