Biomedical subjects

B Xie

Publications and source records attributed to B Xie.

At least 19 recordsLinked to original sources

PubMed1999

STAT5 interaction with the T cell receptor complex and stimulation of T cell proliferation.

The role of STAT (signal transducer and activator of transcription) proteins in T cell receptor (TCR) signaling was analyzed. STAT5 became immediately and transiently phosphorylated on tyrosine 694 in response to TCR stimulation. Expression of the protein tyrosine kinase Lck, a key signaling protein in the TCR complex, activated DNA binding of transfected STAT5A and STAT5B to specific STAT inducible elements. The role of Lck in STAT5 activation was confirmed in a Lck-deficient T cell line in which the activation of STAT5 by TCR stimulation was abolished. Expression of Lck induced specific interaction of STAT5 with the subunits of the TCR, indicating that STAT5 may be directly involved in TCR signaling. Stimulation of T cell clones and primary T cell lines also induced the association of STAT5 with the TCR complex. Inhibition of STAT5 function by expression of a dominant negative mutant STAT5 reduced antigen-stimulated proliferation of T cells. Thus, TCR stimulation appears to directly activate STAT5, which may participate in the regulation of gene transcription and T cell proliferation during immunological responses.

Animals↗

PubMed1999

Interaction between alpha 5 beta 1 integrin and secreted fibronectin is involved in macrophage differentiation of human HL-60 myeloid leukemia cells.

We examined the role of fibronectin (FN) and FN-binding integrins in macrophage differentiation. Increased FN and alpha5beta1 integrin gene expression was observed in phorbol 12-myristate 13-acetate PMA-treated HL-60 cells and PMA- or macrophage-CSF-treated blood monocytes before the manifestation of macrophage markers. After treatment of HL-60 cells and monocytes, newly synthesized FN was released and deposited on the dishes. An HL-60 cell variant, HL-525, which is deficient in the protein kinase Cbeta (PKC-beta) and resistant to PMA-induced differentiation, failed to express FN after PMA treatment. Transfecting HL-525 cells with a PKC-beta expression plasmid restored PMA-induced FN gene expression and macrophage differentiation. Untreated HL-525 cells (which have a high level of the alpha5beta1 integrin) incubated on FN differentiated into macrophages. The percentage of cells having a macrophage phenotype induced by PMA in HL-60 cells, by FN in HL-525 cells, or by either PMA or macrophage-CSF in monocytes was reduced in the presence of mAbs to FN and alpha5beta1 integrin. The integrin-signaling nonreceptor tyrosine kinase, p72Syk, was activated in PMA-treated HL-60 and FN-treated HL-525 cells. We suggest that macrophage differentiation involves the activation of PKC-beta and expression of extracellular matrix proteins such as FN and the corresponding integrins, alpha5beta1 integrin in particular. The stimulated cells, through the integrins, attach to substrates by binding to the deposited FN. This attachment, in turn, may through integrin signaling activate nonreceptor tyrosine kinases, including p72Syk, and later lead to expression of other genes involved in evoking the macrophage phenotype.

Cell Differentiation↗

PubMed1999

Principles of enzyme thermistor systems: applications to biomedical and other measurements.

This chapter presents an overview of thermistor-based calorimetric measurements. Bioanalytical applications are emphasized from both the chemical and biomedical points of view. The introductory section elucidates the principles involved in the thermometric measurements. The following section describes in detail the evolution of the various versions of enzyme-thermistor devices. Special emphasis is laid on the description of modern "mini" and "miniaturized" versions of enzyme thermistors. Hybrid devices are also introduced in this section. In the sections on applications, the clinical/biomedical areas are dealt with separately, followed by other applications. Mention is also made of miscellaneous applications. A special section is devoted to future developments, wherein novel concepts of telemedicine and home diagnostics are highlighted. The role of communication and information technology in telemedicine is also mentioned. In the concluding sections, an attempt is made to incorporate the most recent references on specific topics based on enzyme-thermistor systems.

Biosensing Techniques↗

PubMed1998

Fibronectin-mediated cell adhesion is required for induction of 92-kDa type IV collagenase/gelatinase (MMP-9) gene expression during macrophage differentiation. The signaling role of protein kinase C-beta.

Induction of the 92-kDa gelatinase (MMP-9) gene expression is associated with macrophage differentiation. In this study, we explored the regulatory mechanisms underlying this differentiation-associated MMP-9 gene expression in human HL-60 myeloid leukemia cells and human peripheral blood monocytes. Phorbol 12-myristate 13-acetate (PMA) markedly induced MMP-9 gene expression in HL-60 cells; the induction closely paralleled the timing and extent of PMA-induced cell adhesion and spreading, a hallmark of macrophage differentiation. Similarly, treatment with PMA or macrophage-colony stimulating factor stimulated adherence and spreading of blood monocytes with a concurrent 7- or 5-fold increase in MMP-9 production, respectively. In protein kinase C (PKC)-beta-deficient HL-60 variant cells (HL-525), PMA failed to induce cell adhesion and MMP-9 gene expression. Transfecting HL-525 cells with a PKC-beta expression plasmid restored PKC-beta levels and PMA inducibility of cell adhesion and spreading as well as MMP-9 gene expression. Induction of cell adhesion and MMP-9 gene expression in HL-60 cells and blood monocytes was strongly inhibited by neutralizing monoclonal antibodies to fibronectin (FN) and its receptor alpha5 beta1 integrin. HL-525 cells, which constitutively display high levels of surface alpha5 beta1 integrin, adhered and spread on immobilized FN with concomitant induction of MMP-9 gene expression. Cytochalasins B and D were each a potent inhibitor of MMP-9 production. Our results suggest that alpha5 beta1 integrin-mediated interaction of immature hematopoietic cells with FN plays a critical role in modulating matrix-degrading activities during macrophage differentiation.

Cell Adhesion↗

PubMed1998

Autocrine regulation of macrophage differentiation and 92-kDa gelatinase production by tumor necrosis factor-alpha via alpha5 beta1 integrin in HL-60 cells.

Tumor necrosis factor-alpha (TNF-alpha) gene is one of the early response genes induced by phorbol 12-myristate 13-acetate (PMA) in human HL-60 myeloid leukemia cells. In the present study, we examined the role of the TNF-alpha autocrine loop in PMA-induced macrophage differentiation and gene expression of 92- and 72-kDa gelatinases (MMP-9 and MMP-2). In HL-60 cells, PMA inhibited cell proliferation and induced cell adhesion and spreading, expression of surface maturation marker OKM1 and phagocytic activity, as well as the expression of both gelatinases, which all characterize the macrophage phenotype. In contrast, TNF-alpha alone was only effective in inhibiting cell proliferation. Blocking the endogenous TNF-alpha activity with neutralizing anti-TNF-alpha antibodies abolished all these PMA-induced events with the exception of MMP-2 gene expression. Since fibronectin (FN)-mediated cell adhesion and spreading are prerequisite for both macrophage differentiation and MMP-9 gene expression in HL-60 cells, we hypothesized that TNF-alpha might be involved in modulating the expression of either the FN or its integrin receptor genes. Whereas PMA substantially enhanced the steady state mRNA and protein levels of both FN and alpha5 beta1 integrins, TNF-alpha alone had little effect on the expression of these genes. However, anti-TNF-alpha antibodies blocked PMA-induced augmentation of both alpha5 and beta1 integrin gene expression without affecting the expression of the FN gene. Our results suggest that TNF-alpha may regulate macrophage differentiation and critical matrix-degrading activities of myeloid progenitor cells in an autocrine manner by augmenting surface levels of the alpha5 beta1 integrin, thus promoting interactions with the extracellular matrix, a key event for maturation and migration of these cells during inflammation.

Autocrine Communication↗

PubMed1998

Biochemical characterization of the Cys138Arg substitution associated with the AB variant form of GM2 gangliosidosis: evidence that Cys138 is required for the recognition of the GM2 activator/GM2 ganglioside complex by beta-hexosaminidase A.

The function of the GM2 activator protein is to act as a substrate-specific cofactor in the hydrolysis of GM2 ganglioside by beta-hexosaminidase A. Mutations in the gene encoding it result in the AB variant form of GM2 gangliosidosis. One such mutation, Cys138 Arg, results in the mutant protein being retained and degraded in the endoplasmic reticulum of mammalian cells. In order to characterize the biochemical effects of this substitution, we expressed the mutant protein in transformed bacteria. We first compared the wild-type protein produced by two bacterial expression methods, one requiring protein refolding, with activator purified from the medium of transfected CHO cells. The "activity" and circular dichroism spectrum (alpha-helical content) of all three proteins were similar, justifying the use of refolded activator from transformed bacteria in structure/function studies. Second, the mutant protein was expressed in both bacterial systems and in each retained approximately 2% of the wild type's specific activity. The presence of even this small amount of activity in the mutant protein coupled with a calculated alpha-helical content nearly identical to the wild type, strongly suggest that no major tertiary or secondary structural changes, respectively, had occurred due to the mutation. However, we demonstrate that its heat stability at 60 degrees C is reduced 14-fold, suggesting some localized change in tertiary structure. The loss of a disulfide loop was confirmed by reacting the mutant protein with Ellman's reagent. A kinetic analysis detected a large increase in the apparent K(m) of beta-hexosaminidase A for the mutant; however, there was no apparent change in Vmax. A fluorescence dequenching assay was used to evaluate the ability of the mutant protein to transport lipids and bind GM2 ganglioside. These assays detected no difference between the wild-type and mutant proteins, indicating that the Cys138 Arg substitution has no effect on these functions. We conclude that the mutation specifically affects a domain in the activator protein that is responsible for the recognition of the activator-GM2 ganglioside complex by beta-hexosaminidase A.

Amino Acid Sequence↗

PubMed1998

Clinical differentiation of narrow QRS complex tachycardias.

Supraventricular tachycardias generally present with narrow QRS complexes and are quite commonly seen in the emergency department. Regular narrow QRS complex tachycardias occur in all age groups and may be associated with minimal symptoms, such as palpitations, or, present with hemodynamic compromise resulting in syncope. While history and physical examination are indispensable, they usually do not lead to a definitive diagnosis. The diagnosis is made by careful analysis of the 12-lead ECG. Therapy is based on hemodynamic assessment and understanding of the tachycardia mechanism.

Diagnosis, Differential↗

PubMed1998

Clinical approach to wide QRS complex tachycardias.

Wide QRS complex tachycardia is a frequently encountered arrhythmia in the emergency department and presents a diagnostic challenge to the emergency physician. The history, physical examination, chest radiograph, and electrocardiogram analysis are important in making the correct diagnosis. The diagnosis of ventricular tachycardia is supported by, 1) a history of prior myocardial infarction or congestive heart failure; 2) a physical examination showing cannon A-waves in the jugular venous pulsation or variable heart sounds; 3) a chest radiograph showing cardiomegaly or evidence of prior cardiac surgery; and 4) characteristic ECG features that include AV dissociation, fusion-capture beats, QRS concordance, or, typical morphologic features in leads V1 and V6. This article presents the diagnostic and therapeutic approaches to wide QRS tachycardias.

Diagnosis, Differential↗

PubMed1998

Permanent cardiac pacing.

Implantation of a permanent pacemaker is the most commonly performed surgical operation involving the heart. The modern cardiac pacemaker is a complex device that can sense and pace in both the atrium and ventricle. It also modulates the pacing rate based on sensed physiologic parameters. This article reviews the fundamental principles of pacemaker technology and provides the emergency physician with approaches to common pacemaker problems.

Electrocardiography↗

PubMed1998

Implantable cardioverter defibrillators.

Implantable cardioverter defibrillators have proven to be an effective therapy for life-threatening ventricular arrhythmias. Given the ever-increasing number of patients who have these devices, increasing numbers of patients are likely to present to emergency departments with defibrillator-related problems. This article discusses normal device function, indications for implantation, and technique of implantation. It also focuses on the evaluation and management of patients with these devices presenting to the emergency department.

Arrhythmias, Cardiac↗

PubMed1998

Spongy left ventricular myocardium in an adult.

"Spongy left ventricular myocardium," or noncompaction of left ventricular myocardium, is a rare disorder of endomyocardial morphogenesis. It is usually seen in the pediatric population and is often associated with other congenital cardiac malformations. We describe an adult with noncompaction of left ventricular myocardium without associated congenital cardiac anomalies.

Adolescent↗

PubMed1997

c-Jun NH2-terminal kinases target the ubiquitination of their associated transcription factors.

Regulatory proteins are often ubiquitinated, depending on their phosphorylation status as well as on their association with ancillary proteins that serve as adapters of the ubiquitination machinery. We previously demonstrated that c-Jun is targeted for ubiquitination by its association with inactive c-Jun NH2-terminal kinase (JNK). Phosphorylation by activated JNK protects c-Jun from ubiquitination, thus by prolonging its half-life. In the study reported here, we determined the ability of JNK to target ubiquitination of its other substrates (Elk1 and activating transcription factor 2 (ATF2)) and associated proteins (ATF2 and JunB). We demonstrate that phosphorylation by JNK protects ATF2, but not Elk1, from JNK-targeted ubiquitination. We also show that association of inactive JNK with JunB or ATF2 is necessary to target them for ubiquitination. Unlike its targeting of c-Jun, JNK requires additional cellular components, yet to be identified, to target the ubiquitination of ATF2. Elk1 is phosphorylated by JNK, but JNK neither associates with nor targets Elk1 for ubiquitination. The implications for the dual role of JNK in the regulation of ubiquitination and stability of c-Jun, ATF2, and JunB in normally growing versus stressed cells are discussed.

3T3 Cells↗

PubMed1997

Evaluation of a miniaturized thermal biosensor for the determination of glucose in whole blood.

A miniaturized thermal biosensor has been evaluated as part of a flow-injection analysis system for the determination of glucose in whole blood. Glucose was determined by measuring the heat evolved when samples containing glucose passed through a small column with immobilized glucose oxidase and catalase. Samples of whole blood (1 microliter) can be measured directly, without any pretreatment. The correlation in the response between the thermal biosensor, the Reflolux S meter (Boehringer Mannheim), the Granutest 100 glucose test kit (Merck Diagnostica) and the Ektachem (Kodak) instrument was evaluated. The influence of the hematocrit value and of possible interferences is reported. The correlation measurements show that the thermal biosensor calibrated with aqueous glucose standards generally gives lower values on blood glucose than the reference methods calibrated for serum or blood measurements. Mean negative biases range from 0.53 to 1.16 mmol/l. Differences in sample treatment clearly complicate comparisons and the proper choice of reference method. There was no influence from substances such as ascorbic acid (0.11 mmol/l), uric acid (0.48 mmol/l), urea (4.3 mmol/l) and acetaminophen (0.17 mmol/l) on the response to 5 mmol/l glucose. The hematocrit value does not influence the glucose determination, for hematocrit values of between 13 and 53%.

Biosensing Techniques↗

PubMed1997

Characterization of the affinity of the G(M2) activator protein for glycolipids by a fluorescence dequenching assay.

The G(M2) activator protein is a substrate specific cofactor for degradation of G(M2) ganglioside by lysosomal beta-hexosaminidase A. Mutations in the gene encoding the activator result in the AB-variant form of G(M2) gangliosidosis. The activator protein contains at least three functional elements; a hydrophobic binding pocket, an oligosaccharide binding site(s), and an area that interacts with hexosaminidase A. In this report a fluorescence dequenching assay specific for only the hydrophobic binding pocket is evaluated and optimized. It is shown that various glycolipids inhibit the transport between liposomes of a self-quenching fluorescent lipid probe, octadecylrhodamine, by the activator protein. The level of inhibition produced by each glycolipid is then used to characterize the oligosaccharide-binding specificity of the activator. The fluorescence dequenching assay is also used to evaluate the functionality of a truncated form of the activator protein. Our results indicate that this simple assay can be used to determine structure-function relationships within the normal or mutant forms of the activator. The data suggest that the C-terminus of the activator is required to produce a functional hydrophobic binding pocket.

Binding Sites↗

PubMed1997

Activation of Stat1 by mutant fibroblast growth-factor receptor in thanatophoric dysplasia type II dwarfism.

The achondroplasia class of chondrodysplasias comprises the most common genetic forms of dwarfism in humans and includes achondroplasia, hypochondroplasia and thanatophoric dysplasia types I and II (TDI and TDII), which are caused by different mutations in a fibroblast growth-factor receptor FGFR3 (ref. 1). The molecular mechanism and the mediators of these FGFR3-related growth abnormalities are not known. Here we show that mutant TDII FGFR3 has a constitutive tyrosine kinase activity which can specifically activate the transcription factor Stat1 (for signal transducer and activator of transcription). Furthermore, expression of TDII FGFR3 induced nuclear translocation of Stat1, expression of the cell-cycle inhibitor p21(WAF1/CIP1), and growth arrest of the cell. Thus, TDII FGFR3 may use Stat1 as a mediator of growth retardation in bone development. Consistent with this, Stat1 activation and increased p21(WAF1/CIP1) expression was found in the cartilage cells from the TDII fetus, but not in those from the normal fetus. Thus, abnormal STAT activation and p21(WAF1/CIP1) expression by the TDII mutant receptor may be responsible for this FGFR3-related bone disease.

Animals↗

PubMed1997

Thermistor-based biosensing.

In this review a universal thermistor-based biosensor system is described with examples from clinical chemistry, bioprocess monitoring and environmental control. The technique is based on the measurement of the small temperature changes associated with enzymatic reactions occurring in a microreactor with immobilized enzyme. The system has good operational stability and a sensitivity that permits measurements down to 1 microM concentrations. Current developments include devices constructed by micromachining for multisensing purposes and miniaturised instrumentation intended for use in portable monitoring. With use of special supports for enzyme immobilisation even untreated whole blood samples can be applied. Another current line of investigation involves hybrid biosensors, such as combinations of electrochemistry and calorimetry into bioelectrocalorimetric devices with interesting new properties.

Alcohols↗

PubMed1997

Detection of proviral DNA of bovine leukaemia virus in cattle by a combination of in-situ hybridization and the polymerase chain reaction.

Bovine leukaemia virus (BLV) proviral DNA was detected in lymphocytes isolated from cattle with persistent lymphocytosis (PL) by the polymerase chain reaction (PCR) and in-situ hybridization (ISH) with a biotinylated pX DNA probe. Many positive cells were observed when short-term culture and a combination of ISH with PCR were used. Immunohistochemical examination of lymphocytes isolated from the lymph node showed that BLV attached mainly to surface immunoglobulins (SIg) of positive B lymphocytes, and to a few tumour-associated antigen (TAA)-, PanT-, and CD8-positive cells and non-CD4 positive cells. Electron microscopical examination revealed colloidal gold particles within the nuclei and cytoplasm of lymphocytes. Lymphoid cells from neoplastic lymph node of enzootic bovine leukosis (EBL) cases gave particularly strong positive signals with the ISH-PCR method. The technique of combined ISH and PCR with a biotinylated pX probe may prove useful in future studies of EBL.

Animals↗

PubMed1997

Direct vasodilator effect of milrinone, an inotropic drug, on arterial coronary bypass grafts. FANZCA .

Milrinone is an inotropic drug with vasodilator activity that has been shown to be useful in increasing cardiac output and decreasing wedge pressure. Despite these advantages, it is unknown whether this drug can be used for the treatment of perioperative spasm of coronary bypass grafts. This study was undertaken to investigate the in vitro vascular effect of milrinone on internal thoracic arteries obtained from patients undergoing coronary artery bypass grafting. The results showed that milrinone produced a potent, concentration-dependent, preventive effect on the norepinephrine-induced contraction of internal thoracic arteries, as well as reversing contraction of internal thoracic arteries by receptor-dependent agents, including the thromboxane A2 mimetic U46619, the vasoconstrictor peptide endothelin-1, and the alpha1-adrenal receptor agonist phenylephrine. The relaxing effect of milrinone was weaker, however, on internal thoracic arteries contracted with 25 mmol/L potassium chloride. Comparison of milrinone with other vasodilators, including papaverine, nitroprusside, and glyceryl trinitrate, showed milrinone to be more potent than papaverine but less potent than nitroprusside and glyceryl trinitrate. The inhibitory effect of milrinone on internal thoracic artery contraction appeared as a reduction in contractile force, not as an increase in the values of concentrations of the agonists causing 50% maximal contraction, which indicates that milrinone exerts its vasodilator effect directly on the smooth muscles, not on the membrane receptors. The results also showed no significant difference in relaxing effect between internal thoracic artery rings with and without endothelium. In conclusion, this study provides experimental evidence that milrinone is a potent, endothelium-independent, direct vasodilator of the human internal thoracic artery and provides the scientific rationale for a future clinical trial with this drug for the perioperative treatment of internal thoracic artery spasm in cardiac surgical patients.

Coronary Artery Bypass↗