Effects of danofloxacin and tilmicosin on circulating neutrophils in beef heifers.
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Animals↗
Biomedical subjects
D A Frank
Publications and source records attributed to D A Frank.
Potential interaction between celecoxib and warfarin.
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Aged↗
Fusion of the ets transcription factor TEL to Jak2 results in constitutive Jak-Stat signaling.
To study constitutive Janus kinase signaling, chimeric proteins were generated between the pointed domain of the ets transcription factor TEL and the cytosolic tyrosine kinase Jak2. The effects of these proteins on interleukin-3 (IL-3)-dependent proliferation of the hematopoietic cell line, Ba/F3, were studied. Fusion of TEL to the functional kinase (JH1) domain of Jak2 resulted in conversion of Ba/F3 cells to factor-independence. Importantly, fusion of TEL to the Jak2 pseudokinase (JH2) domain or a kinase-inactive Jak2 JH1 domain had no effect on IL-3-dependent proliferation of Ba/F3 cells. Active TEL-Jak2 constructs (consisting of either Jak2 JH1 or Jak2 JH2+JH1 domain fusions) were constitutively tyrosine-phosphorylated but did not affect phosphorylation of endogeneous Jak1, Jak2, or Jak3. TEL-Jak2 activation resulted in the constitutive tyrosine phosphorylation of Stat1, Stat3, and Stat5 as determined by detection of phosphorylation using activation-specific antibodies and by binding of each protein to a preferential GAS sequence in electrophoretic mobility shift assays. Elucidation of signaling events downstream of TEL-Jak2 activation may provide insight into the mechanism of leukemogenesis mediated by this oncogenic fusion protein.
Animals↗
HTLV-I-infected T cells evade the antiproliferative action of IFN-beta.
Human T-cell lymphotropic virus type I (HTLV-I)-infected T-cell clones enter the S-phase of the cell cycle in the absence of exogenous IL-2. The pathway by which HTLV-I activates the host T cell may circumvent normal immunoregulatory mechanisms and thus be important for the pathogenesis of HTLV-I-induced diseases. The early control of viral infections is in part mediated by interferons (IFNs), which possess both antiviral and antiproliferative functions. In order to investigate the antiproliferative effect of IFN-beta on HTLV-I-induced T-cell activation, we generated T-cell clones from patients with HTLV-I-associated myelopathy/tropical spastic paraparesis by single-cell cloning under limiting dilution conditions. Here we demonstrate that HTLV-I-induced T-cell proliferation is resistant to the antiproliferative action of IFN-beta. Moreover, HTLV-I-infected T-cell clones continue to constitutively secrete IFN-gamma in the presence of high doses of IFN-beta. HTLV-I-infected T cells express normal levels of IFNAR1 and are able to respond to IFN-beta by phosphorylation of STAT1 on Tyr701, although they display a relative increase in phosphorylation of the transcriptionally inactive STAT1beta when compared with STAT1alpha. Thus, HTLV-I promotes cell cycle progression in G1 by a mechanism that overcomes inhibitory signals, thereby circumventing an innate immune defense mechanism.
Cell Cycle↗
The functional synergy between IL-12 and IL-2 involves p38 mitogen-activated protein kinase and is associated with the augmentation of STAT serine phosphorylation.
IL-12 and IL-2 can stimulate mitogen- or CD3-activated T cells to proliferate, produce IFN-gamma, and kill tumor cells. The magnitude of these functional responses is greatly augmented when T cells are activated by the combination of IL-12 and IL-2. Although peripheral blood T cells are largely unresponsive to these cytokines without prior activation, a small subset of CD8+ T cells (CD8+CD18bright) is strongly activated by the combination of IL-12 and IL-2. In this report we show that the functional synergy between IL-12 and IL-2 in CD8+CD18bright T cells correlates with the activation of the stress kinases, p38 mitogen-activated protein (MAP) kinase and stress-activated protein kinase (SAPK)/Jun N-terminal kinase, but not with the activation of the extracellular signal-regulated kinases. The functional synergy between IL-2 and IL-12 is also associated with a prominent increase in STAT1 and STAT3 serine phosphorylation over that observed with IL-12 or IL-2 alone. By contrast, STAT tyrosine phosphorylation is not augmented over that seen with either cytokine alone. A specific inhibitor of p38 MAP kinase completely inhibits the serine phosphorylation of STAT1 and STAT3 induced by IL-12 and IL-2 and abrogates the functional synergy between IL-12 and IL-2 without affecting STAT tyrosine phosphorylation. This suggests that p38 MAP kinase may play an important role in regulating STAT serine phosphorylation in response to the combination of IL-12 and IL-2. Furthermore, these findings indicate that the optimal activation of T cells by IL-12 and IL-2 may depend on an interaction between the p38 MAP kinase and Janus kinase/STAT signaling pathways.
Adjuvants, Immunologic↗
IL-2 induces STAT4 activation in primary NK cells and NK cell lines, but not in T cells.
IL-2 exerts potent but distinct functional effects on two critical cell populations of the immune system, T cells and NK cells. Whereas IL-2 leads to proliferation in both cell types, it enhances cytotoxicity primarily in NK cells. In both T cells and NK cells, IL-2 induces the activation of STAT1, STAT3, and STAT5. Given this similarity in intracellular signaling, the mechanism underlying the distinct response to IL-2 in T cells and NK cells is not clear. In this study, we show that in primary NK cells and NK cell lines, in addition to the activation of STAT1 and STAT5, IL-2 induces tyrosine phosphorylation of STAT4, a STAT previously reported to be activated only in response to IL-12 and IFN-alpha. This activation of STAT4 in response to IL-2 is not due to the autocrine production of IL-12 or IFN-alpha. STAT4 activated in response to IL-2 is able to bind to a STAT-binding DNA sequence, suggesting that in NK cells IL-2 is capable of activating target genes through phosphorylation of STAT4. IL-2 induces the activation of Jak2 uniquely in NK cells, which may underlie the ability of IL-2 to activate STAT4 only in these cells. Although the activation of STAT4 in response to IL-2 occurs in primary resting and activated NK cells, it does not occur in primary resting T cells or mitogen-activated T cells. The unique activation of the STAT4-signaling pathway in NK cells may underlie the distinct functional effect of IL-2 on this cell population.
Autocrine Communication↗
Fludarabine-induced immunosuppression is associated with inhibition of STAT1 signaling.
Fludarabine is a nucleoside analog used in the treatment of hematologic malignancies that can induce severe and prolonged immunosuppression. Although it can be incorporated into the DNA of dividing cells, fludarabine is also a potent inhibitor of cells with a low growth fraction, thus it must have other mechanisms of action. STAT1, which is activated in response to many lymphocyte-activating cytokines including the interferons, is essential for cell-mediated immunity, as the absence of this protein is associated with prominent defects in the ability to control viral infections. Here we show that fludarabine, but not the immunosuppressant cyclosporine A, inhibits the cytokine-induced activation of STAT1 and STAT1-dependent gene transcription in normal resting or activated lymphocytes. Fludarabine caused a specific depletion of STAT1 protein (and mRNA) but not of other STATs. This loss of STAT1 was also seen in cells from patients treated with fludarabine in vivo. Brief exposure to fludarabine led to a sustained loss of STAT1, analogous to the prolonged period of immunosuppression induced by exposure to the drug in vivo. Thus, STAT1 may be a useful target in the development of new immunosuppressive and antineoplastic agents.
Alternative Splicing↗
Requirement for Ras/Rac1-mediated p38 and c-Jun N-terminal kinase signaling in Stat3 transcriptional activity induced by the Src oncoprotein.
Signal transducers and activators of transcription (STATs) are transcription factors that mediate normal biologic responses to cytokines and growth factors. However, abnormal activation of certain STAT family members, including Stat3, is increasingly associated with oncogenesis. In fibroblasts expressing the Src oncoprotein, activation of Stat3 induces specific gene expression and is required for cell transformation. Although the Src tyrosine kinase induces constitutive Stat3 phosphorylation on tyrosine, activation of Stat3-mediated gene regulation requires both tyrosine and serine phosphorylation of Stat3. We investigated the signaling pathways underlying the constitutive Stat3 activation in Src oncogenesis. Expression of Ras or Rac1 dominant negative protein blocks Stat3-mediated gene regulation induced by Src in a manner consistent with dependence on p38 and c-Jun N-terminal kinase (JNK). Both of these serine/threonine kinases and Stat3 serine phosphorylation are constitutively induced in Src-transformed fibroblasts. Furthermore, inhibition of p38 and JNK activities suppresses constitutive Stat3 serine phosphorylation and Stat3-mediated gene regulation. In vitro kinase assays with purified full-length Stat3 as the substrate show that both JNK and p38 can phosphorylate Stat3 on serine. Moreover, inhibition of p38 activity and thus of Stat3 serine phosphorylation results in suppression of transformation by v-Src but not v-Ras, consistent with a requirement for Stat3 serine phosphorylation in Src transformation. Our results demonstrate that Ras- and Rac1-mediated p38 and JNK signals are required for Stat3 transcriptional activity induced by the Src oncoprotein. These findings delineate a network of tyrosine and serine/threonine kinase signaling pathways that converge on Stat3 in the context of oncogenesis.
3T3 Cells↗
Level of in utero cocaine exposure and neonatal ultrasound findings.
OBJECTIVE: To assess whether there is an association between the level of in utero cocaine exposure and findings on neonatal cranial ultrasound, controlling for potentially confounding variables. STUDY DESIGN: In a prospective longitudinal study, three cocaine exposure groups were defined by maternal report and infant meconium assay: unexposed, heavier cocaine exposure (>75th percentile self-reported days of use or of meconium benzoylecogonine concentration) or lighter cocaine exposure (all others). Neonatal ultrasounds from 241 well, term infants were read by a single radiologist who was masked to the exposure group. RESULTS: Infants with lighter cocaine exposure did not differ from the unexposed infants on any ultrasound findings. After controlling for infant gender, gestational age, and birth weight z scores and for maternal parity, blood pressure in labor, ethnicity, and use of cigarettes, alcohol, and marijuana during pregnancy, the more heavily cocaine-exposed infants were more likely than the unexposed infants to show subependymal hemorrhage in the caudothalamic groove (covariate adjusted odds ratio: 3.88; 95% confidence interval: 1.45, 10.35). CONCLUSIONS: This is the first study to demonstrate that ultrasound findings suggestive of vascular injury to the neonatal central nervous system are related to the level of prenatal cocaine exposure. Inconsistency in previous research in identifying an association between prenatal cocaine exposure and neonatal cranial ultrasound findings may reflect failure to consider dose effects.
Cocaine↗
Effect of protein intake during training on biochemical and performance variables in sled dogs.
OBJECTIVE: To determine effects of protein intake on blood variables, plasma volume, and maximal oxygen uptake (VO2max) in sled dogs undergoing rigorous training. ANIMALS: 32 Alaskan sled dogs, between 2 and 6 years old. PROCEDURE: Dogs were assigned to 1 of 4 groups on the basis of age, sex, and ability. Isocaloric diets containing 18% (diet A), 23% (diet B), 29% (diet C), or 35% (diet D) of energy as protein were assigned randomly to each group and fed 1 month before and during a 12-week training period. Maximal oxygen uptake was measured at 0 (before training) and 12 weeks. Body weight, protein and energy intake, plasma volume, PCV, hemoglobin concentration, and serum biochemical variables were measured at 0, 8, and 12 weeks. RESULTS: Serum biochemical variables, PCV, and hemoglobin concentration remained within reference ranges for all dogs. Dogs fed diet A had a decrease in VO2max and a greater rate of soft tissue injury throughout training, compared with dogs fed the other diets. At 12 weeks, dogs fed diets C and D had greater serum sodium concentration and hemoglobin concentration than did dogs fed diet A. Dogs fed diet D also had more plasma volume at 12 weeks than did dogs of any other group. CONCLUSIONS AND CLINICAL RELEVANCE: Consumption of a diet with 18% dietary protein on an energy basis (3.0 g of protein/kg of body weight) is insufficient to meet the metabolic requirements of sled dogs in training. For intense interval work, a diet with 35% dietary protein as energy (6.0 g of protein/kg) may provide a performance advantage by promoting an increase in plasma volume.
Animals↗
STAT signaling in the pathogenesis and treatment of cancer.
Exceptional advances have been made recently in our understanding of the signaling pathways that control cellular growth, differentiation, and survival. These processes are regulated by extracellular stimuli such as cytokines, cell-cell interactions, and cell-matrix interactions, which trigger a series of intracellular events culminating in the modulation of specific genes. STATs are a highly homologous group of transcription factors that are activated by various pathways and regulate many of the genes controlling cellular function. STATs are activated by tyrosine phosphorylation and modulated by serine phosphorylation, placing them at a convergence point for numerous intracellular signaling pathways. Given the importance of STATs in the control of normal physiologic processes, it is not surprising that inappropriate activation of these proteins has been found in human malignancies. A number of distinct mechanisms have been elucidated by which STATs are activated inappropriately, including autocrine or paracrine stimulation of normal receptors and increased activity of tyrosine kinases through enhanced expression, mutations, or the presence of activating proteins. Furthermore, inappropriate STAT serine phosphorylation has been found in several tumors as well. The increased understanding of signaling pathways in tumors can be translated into therapeutic strategies that have the potential to be more selective and less toxic than current anti-cancer treatments. Approaches which may be effective include the development of antagonists of receptors that can trigger STAT activation, inhibitors of the tyrosine and serine kinases that phosphorylate and activate STATs, agents that decrease STAT levels or inhibit their recruitment to kinases, and molecules that can prevent the binding of STATs to target DNA sequences. Thus, elucidation of cellular and biochemical processes in tumors has enhanced our understanding of the pathogenesis of malignancies and may provide the basis for significant advances in therapy.
Animals↗
Characterization of a novel subset of CD8(+) T cells that expands in patients receiving interleukin-12.
IL-12 has significant antitumor activity in mice that may be mediated by CD8(+) T cells. We show in this report that repeated subcutaneous injections of IL-12 in patients with cancer resulted in the selective expansion of a subset of peripheral blood CD8(+) T cells. This T cell subset expressed high levels of CD18 and upregulated IL-12 receptor expression after IL-12 treatment in vivo. In normal subjects, these CD3(+)CD8(+)CD18(bright) T cells expressed IL-12 and IL-2 receptors and adhesion/costimulatory molecules to a greater degree than other CD8(+) and CD4(+) T cells. They appeared morphologically as large granular lymphocytes, although they did not express NK cell markers such as CD56. In addition, CD8(+)CD18(bright) T cells were almost exclusively T cell receptor (TCR) alphabeta+, and exhibited a TCR Vbeta repertoire that was strikingly oligoclonal, whereas the Vbeta repertoire of CD18(dim) T cells was polyclonal. Although CD8+CD18(bright) T cells demonstrated little functional responsiveness to IL-12 or IL-2 alone in vitro, they responded to the combination of IL-12+IL-2 with strong IFN-gamma production and proliferation and enhanced non-MHC-restricted cytolytic activity. In contrast, CD18(dim) T cells were not activated by IL-12 or IL-2, alone or in combination. These findings demonstrate that CD8+CD18(bright) T cells are a unique population of peripheral blood lymphocytes with features of both memory and effector cells that are capable of TCR-independent activation through combined stimulation with IL-12+IL-2. As this activation results in IFN-gamma production and enhanced cytolytic activity, these T cells may play a role in innate as well as acquired immunity to tumors and infectious pathogens. Additional studies will be necessary to determine whether CD8+CD18(bright) T cells mediate the antitumor effect of IL-12 or IL-2 administered to cancer patients, and if so, whether maximal activation of these T cells with the combination of IL-12+IL-2 in vivo can augment the clinical effectiveness of these cytokines.
CD8-Positive T-Lymphocytes↗
Neonatal neurobehavioral and neuroanatomic correlates of prenatal cocaine exposure. Problems of dose and confounding.
Complex methodologic challenges face researchers studying the effects of prenatal cocaine exposure on infant outcome. These include unavoidable imprecision in ascertaining the gestational timing and dose of cocaine to which the fetus was exposed and difficulties in identifying and quantifying the confounding, mediating, and moderating variables. Review of research on neonatal behavioral and cranial ultrasound findings following in utero cocaine exposure is used to illustrate these issues. We conclude that there are measurable but not dramatic dose-related effects of prenatal cocaine exposure on infant central nervous system structure and function. The effects of dose of prenatal cocaine exposure on later child development remain to be determined. Such research would be facilitated by a scientific consensus delineating relative doses of prenatal cocaine exposure.
Brain↗
Altered interleukin-12 responsiveness in Th1 and Th2 cells is associated with the differential activation of STAT5 and STAT1.
T-cell activation in response to interleukin-12 (IL-12) is mediated through signaling events that include the tyrosine phosphorylation of STAT4. IL-12 responsiveness and the ability of IL-12 to activate STAT4 is different in T cells induced to differentiate into a Th1 or Th2 phenotype. In this report, we show that STAT5, STAT1alpha, and STAT1beta, in addition to STAT4, are tyrosine phosphorylated in response to IL-12 in phytohemagglutinin (PHA)-activated human T cells. To understand how the activation of these STATs contributes to T-cell IL-12 responsiveness, we analyzed the IL-12-induced activation of STAT5 and STAT1 in T cells stimulated to undergo Th1 or Th2 differentiation. The IL-12-induced tyrosine phosphorylation of STAT5 and STAT1, but not STAT4, is augmented in T cells activated into Th1 cells with PHA + interferon-gamma (IFN-gamma) compared with T cells activated with PHA alone. STAT5 DNA binding induced by IL-12 is also augmented in T cells activated with PHA + IFN-gamma compared with T cells activated with PHA alone, whereas STAT4 DNA binding is not increased. In contrast, the IL-12-induced activation of these STATs is inhibited in T cells activated into Th2 cells with PHA + IL-4. The enhancement of IL-12 signaling by IFN-gamma is not a direct effect of IFN-gamma on T cells, but rather is mediated by IL-12 that is produced by antigen-presenting cells in response to IFN-gamma. This positive autoregulatory effect of IL-12 on the activation of select STATs correlates with an increase in T-cell IFN-gamma production in response to IL-12. These findings suggest that the activation of STAT5 and STAT1 may augment select STAT4-dependent functional responses to IL-12 in Th1 cells.
Adult↗
Semantic development of African-American children prenatally exposed to cocaine.
Semantic content categories were described for the single word, multiple word, and verb relation utterances of 22 African-American 2-year-olds during a 90-min laboratory session. Half of the toddlers had been exposed prenatally to cocaine and half were unexposed, as documented by biological assay in the newborn period. The exposed and unexposed groups were carefully matched on demographic, medical, and proximal caregiving variables. Children's spontaneous utterances were transcribed from audio- and videotapes during the laboratory session and scored for semantic features by a team of reliable coders who were masked to child exposure status. General productive language features (utterance length, verbosity, and intelligibility) were also assessed. To evaluate general language and cognitive skills, the toddlers were evaluated with the Sequenced Inventory of Communicative Development-Revised (SICD-R) and the Bayley Scales of Infant Development (BSID). Although exposed and nonexposed toddlers exhibited similar sequences of semantic development, the exposed toddlers were more restricted and delayed in their semantic representations. No significant group differences were observed, however, for structural features of language (e.g., utterance length, distribution of utterance types) or for children's general language and cognitive functioning as assessed by standardized assessments (i.e., SICD-R, BSID). Thus, a history of prenatal cocaine exposure and associated risk factors (e.g., prenatal exposure to alcohol, diminished birth weight) are related to delays in early semantic development. Proposed diagnostic and treatment strategies are discussed.
Black or African American↗
Blood pressure in children exposed prenatally to cocaine.
Prenatal exposure to cocaine may alter autonomic nervous system function resulting in hypertension. Blood pressure was measured oscillometrically in a convenience sample of cocaine-exposed (n = 32) and nonexposed (n = 23) children, aged 8 to 26 months, by examiners unaware of exposure status. The exposed and nonexposed groups did not differ in systolic pressure (mean +/- SD 102.3 +/- 9.9 mmHg vs 119.1 +/- 19.4). Prenatal exposure to cocaine does not appear to be associated with elevated blood pressure in young children.
Autonomic Nervous System↗
Evolution of a cardiovascular information system.
The competitive nature of our evolving health care system mandates a concomitant increase in the level of sophistication of the cardiovascular information system. The new paradigms in health care also mandate a re-engineering of the process of data collection and analysis. This paper deals with a variety of hardware, software, and human issues encountered at our institution. Topics covered include implementation of service line management, utilization of an interdisciplinary medical informatics model, database conversion, and application development. Special attention will be paid to the development of a system which is a comprehensive information system for clinical and financial management of the cardiovascular patient, not just a quality-of-care database.
Cardiology Service, Hospital↗
B lymphocytes from patients with chronic lymphocytic leukemia contain signal transducer and activator of transcription (STAT) 1 and STAT3 constitutively phosphorylated on serine residues.
To explore the pathogenesis of chronic lymphocytic leukemia (CLL), we examined whether phosphorylation of one or more signal transducer and activator of transcription (STAT) factors was abnormal in cells from CLL patients. No constitutive tyrosine phosphorylation was detected on any STAT in CLL cells. To assess the phosphorylation of serine residues of STAT1 and STAT3 in CLL cells, we raised antibodies that specifically recognize the form of STAT1 phosphorylated on ser-727 and the form of STAT3 phosphorylated on ser-727. We found that in 100% of patients with CLL (n = 32), STAT1 and STAT3 were constitutively phosphorylated on serine. This was in contrast to normal peripheral blood B lymphocytes or CD5+) B cells isolated from tonsils, in which this phosphorylation was absent. Serine phosphorylation of STAT1 and STAT3 was seen occasionally in other leukemias, but it was a universal finding only in CLL. The serine phosphorylation of these STATs was a continuous process, as incubation of CLL cells with the kinase inhibitor H7 led to the dephosphorylation of these serine residues. The STAT serine kinase in CLL cells has not been identified, and appears to be neither mitogen-activated protein kinase nor pp70(s6k). In summary, the constitutive serine phosphorylation of STAT1 and STAT3 is present in all CLL samples tested to date, although the physiologic significance of this modification remains to be determined.
Antibody Formation↗