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Biomedical subjects

S Zhou

Publications and source records attributed to S Zhou.

At least 397 records · Page 22Linked to original sources
PubMed1996

[Detection of immunoglobulin heavy chain (IgH) gene rearrangement in ANLL by polymerase chain reaction amplification and Southern blot].

Immunoglobulin heavy chain (IgH) gene rearrangement serves as a marker of clonality in B lymphoproliferative malignancies. In order to study the IgH rearranged gene in acute nonlymphoblastic leukemia (ANLL) patients we combine polymerase chain reaction (PCR) with Southern blot to detect 41 ANLL patients and 7 of them (17.1%) were found to have IgH rearrangement by PCR amplification. All these 7 positive cases were confirmed by Southern blot. The sensitivity of this method was 10(-4)-10(-5) level. In 12 patients with complete remission, 3 (25.0%) were found to have IgH rearranged gene. All these 3 cases had clinical relapse within 6 months. Our results show that IgH rearrangement not only may occur in lymphoblastic leukemia of B lineage, but also can be found in ANLL. The mechanism may be that in some ANLL patients, the leukemic transforming event might involve stem cells capable of both B cell and myeloid differentiation or ANLL might differentiate along different lineage with predominant appearance of one or the other subclone in the course of the disease.

Blotting, Southern↗
PubMed1995

The chromo superfamily: new members, duplication of the chromo domain and possible role in delivering transcription regulators to chromatin.

Using computer methods for detecting conserved amino acid sequence motifs, we show that the chromatin organization modifier (chromo) domain that has been previously identified in several proteins involved in transcription down-regulation is present in a much larger group of (putative) chromatin-binding proteins, some of which are positive rather than negative regulators of transcription. The most interesting new members of the chromo superfamily are Drosophila male-specific lethal (MSL-3) protein involved in the X chromosome gene dosage compensation in the males and human retinoblastoma-binding protein RBP-1. We show that the chromo domain is duplicated in several chromatin-binding proteins and use this observation to interpret recent results on chromatin binding obtained with chimeric chromo domain-containing proteins. We hypothesize that the chromo domain may be a vehicle that delivers both positive and negative transcription regulators to the sites of their action on chromatin.

Amino Acid Sequence↗
PubMed1995

The yeast immunophilin Fpr3 is a physiological substrate of the tyrosine-specific phosphoprotein phosphatase Ptp1.

The tyrosine-specific phosphoprotein phosphatase encoded by the Saccharomyces cerevisiae PTP1 gene dephosphorylates artificial substrates in vitro, but little is known about its functions and substrates in vivo. The presence of Ptp1 resulted in dephosphorylation of multiple tyrosine-phosphorylated proteins in yeast expressing a heterologous tyrosine-specific protein kinase, indicating that Ptp1 can dephosphorylate a broad range of substrates in vivo. Correspondingly, several proteins phosphorylated at tyrosine by endogenous protein kinases exhibited a marked increase in tyrosine phosphorylation in ptp1 mutant cells. One of these phosphotyrosyl proteins (p70) was also dephosphorylated in vitro when incubated with recombinant Ptp1. p70 was purified to homogeneity; analysis of four tryptic peptides revealed that p70 is identical to the recently described FPR3 gene product, a nucleolarly localized proline rotamase of the FK506- and rapamycin-binding family. The identity of p70 with Fpr3 was confirmed in the demonstration that the abundance of tyrosine-phosphorylated p70 in ptp1 mutants was strictly correlated with the level of FPR3 expression; immobilized phosphotyrosyl Fpr3 was directly dephosphorylated by recombinant Ptp1. Site-directed mutagenesis demonstrated that the site of tyrosine phosphorylation is Tyr-184, which resides within the nucleolin-like amino-terminal domain of Fpr3. Protein kinase activities from yeast cell extracts can bind to and phosphorylate the immobilized amino-terminal domain of Fpr3 on serine, threonine, and tyrosine. Fpr3 represents the first phosphotyrosyl protein identified in S. cerevisiae that is not itself a protein kinase and is as yet the only known physiological substrate of Ptp1.

Amino Acid Sequence↗
PubMed1995

Identification and characterization of a TFIID-like multiprotein complex from Saccharomyces cerevisiae.

Although the mechanisms of transcriptional regulation by RNA polymerase II are apparently highly conserved from yeast to man, the identification of a yeast TATA-binding protein (TBP)-TBP-associated factor (TAFII) complex comparable to the metazoan TFIID component of the basal transcriptional machinery has remained elusive. Here, we report the isolation of a yeast TBP-TAFII complex which can mediate transcriptional activation by GAL4-VP16 in a highly purified yeast in vitro transcription system. We have cloned and sequenced the genes encoding four of the multiple yeast TAFII proteins comprising the TBP-TAFII multisubunit complex and find that they are similar at the amino acid level to both human and Drosophila TFIID subunits. Using epitope-tagging and immunoprecipitation experiments, we demonstrate that these genes encode bona fide TAF proteins and show that the yeast TBP-TAFII complex is minimally composed of TBP and seven distinct yTAFII proteins ranging in size from M(r) = 150,000 to M(r) = 25,000. In addition, by constructing null alleles of the cloned TAF-encoding genes, we show that normal function of the TAF-encoding genes is essential for yeast cell viability.

Amino Acid Sequence↗
PubMed1995

Molecular cloning and expression of the 32-kDa subunit of human TFIID reveals interactions with VP16 and TFIIB that mediate transcriptional activation.

Transcription factor TFIID consists of TATA binding protein (TBP) and at least eight TBP-associated factors (TAFs). As TAFs are required for activated but not basal transcription, we have proposed that TAFs act as coactivators to mediate signals between activators and the basal transcription machinery. Here we report the cloning, expression, and biochemical characterization of the 32-kDa subunit of human (h) TFIID, termed hTAFII32. We find that hTAFII32 is the human homologue of Drosophila TAFII40. In vitro protein-protein interaction assays reveal that as observed with Drosophila TAFII40, hTAFII32 interacts with the C-terminal 39-amino acid activation domain of the acidic transactivator viral protein 16 (VP16) as well as with the general transcription factor TFIIB. Moreover, a partial recombinant TFIID complex containing hTAFII32 was capable of mediating in vitro transcriptional activation by the VP16 activation domain. These findings indicate that specific activator-coactivator interactions have been conserved between human and Drosophila and provide additional support for the function of these interactions in mediating transcriptional activation.

Amino Acid Sequence↗
PubMed1995

Male-specific lethal 2, a dosage compensation gene of Drosophila, undergoes sex-specific regulation and encodes a protein with a RING finger and a metallothionein-like cysteine cluster.

In Drosophila the equalization of X-linked gene products between males and females, i.e. dosage compensation, is the result of a 2-fold hypertranscription of most of these genes in males. At least four regulatory genes are required for this process. Three of these genes, maleless (mle), male-specific lethal 1 (msl-1) and male-specific lethal 3 (msl-3), have been cloned and their products have been shown to interact and to bind to numerous sites on the X chromosome of males, but not of females. Although binding to the X chromosome is negatively correlated with the function of the master regulatory gene Sex lethal (Sxl), the mechanisms that restrict this binding to males and to the X chromosome are not yet understood. We have cloned the last of the known autosomal genes involved in dosage compensation, male-specific lethal 2 (msl-2), and characterized its product. The encoded protein (MSL-2) consists of 769 amino acid residues and has a RING finger (C3HC4 zinc finger) and a metallothionein-like domain with eight conserved and two non-conserved cysteines. In addition, it contains a positively and a negatively charged amino acid residue cluster and a coiled coil domain that may be involved in protein-protein interactions. Males produce a msl-2 transcript that is shorter than in females, due to differential splicing of an intron of 132 bases in the untranslated leader. Using an antiserum against MSL-2 we have shown that the protein is expressed at a detectable level only in males, where it is physically associated with the X chromosome. Our observations suggest that MSL-2 may be the target of the master regulatory gene Sxl and provide the basic elements of a working hypothesis on the function of MSL-2 in mediating the 2-fold increase in transcription that is characteristic of dosage compensation.

Amino Acid Sequence↗
PubMed1995

Estimating the proportion of immunes in censored samples: a simulation study.

We review currently known results concerning the estimation of an 'immune' or 'cured' proportion, and testing for the presence of immunes, in censored survival data, suggesting that a firm theoretical foundation now exists for analysis. Two types of estimators, parametric and non-parametric, are discussed and compared with respect to their theoretical properties, and, by simulation, with respect to their small sample behaviour. Both estimators have advantages and drawbacks, but together provide powerful tools for the perceptive analysis of survival data with, or even without, immune individuals.

Humans↗
PubMed1995

Electromechanical delay in isometric muscle contractions evoked by voluntary, reflex and electrical stimulation.

Electromechanical delay (EMD) in isometric contractions of knee extensors evoked by voluntary, tendon reflex (TR) and electrical stimulation (ES) was investigated in 21 healthy young subjects. The subject performed voluntary knee extensions with maximum effort (maximal voluntary contraction, MVC), and at 30%, 60% and 80% MVC. Patellar tendon reflexes were evoked with the reflex hammer being dropped from 60 degrees, 75 degrees and 90 degrees positions. In the percutaneous ES evoked contractions, single switches were triggered with pulses of duration 1.0 ms and of intensities 90, 120 and 150 V. Electromyograms of the vastus lateralis and rectus femoris muscles were recorded using surface electrodes. The isometric knee extension force was recorded using a load cell force transducer connected to the subject's lower leg. The major finding of this study was that EMD of the involuntary contractions [e.g. mean 22.1 (SEM 1.32) ms in TR 90 degrees; mean 17.2 (SEM 0.62) ms in ES 150 V] was significantly shorter than that of the voluntary contractions [e.g. mean 38.7 (SEM 1.18) ms in MVC, P < 0.05]. The relationships between EMD, muscle contractile properties and muscle fibre conduction velocity were also investigated. Further study is needed to explain fully the EMD differences found between the voluntary and involuntary contractions.

Adult↗
PubMed1995

Optimization of leaf positions when shaping a radiation field with a multileaf collimator.

Computer-controlled multileaf collimators are receiving increasing acceptance for clinical use in radiation therapy due to their efficiency in shaping radiation fields. Because of the physical leaf width of the multileaf collimator, a smooth field contour prescribed by a physician is approximated by a staircase-shaped field boundary. The point of intersection with the prescribed field contour determines the error of the approximation and may affect the outcome of the treatment. This article provides analytically optimized solutions for leaf positioning based on two different geometric object functions.

Humans↗
PubMed1995

Leukotriene B4 formation: human neutrophil-airway epithelial cell interactions.

Leukotriene B4 (LTB4) is a potent inflammatory mediator involved in the pathogenesis of many pulmonary diseases. Although the neutrophil is the predominant source of LTB4, other cells can also interact with neutrophils and increase LTB4 formation. In this study, we investigated whether human neutrophil-airway epithelial cell interactions can increase LTB4 formation. Neutrophils were cocultured with transformed airway epithelial cells (9HTEo- cells), and LTB4 and leukotriene A4 (LTA4) degradation product release was measured by high-performance liquid chromatography and ultraviolet spectrometry. When stimulated with the calcium ionophore A-23187, neutrophil-9HTEo- cell cocultures released more LTB4 and less LTA4 degradation products in a time- and dose-related manner than did neutrophils alone. This increase in LTB4 release involved the metabolism of neutrophil-derived LTA4 to LTB4 by 9HTEo- cells and was affected by the neutrophil-to-epithelial cell ratio. Enhanced LTB4 release required proximity between neutrophils and 9HTEo- cells but not specific cell-cell adhesion. Our data demonstrate that human neutrophil-airway epithelial cell interactions can increase LTB4 formation through transcellular arachidonic acid metabolism.

Cell Adhesion↗
PubMed1995

Expression and function of the beta-adrenergic receptor coupled-adenylyl cyclase system on human airway epithelial cells.

Beta-adrenergic agonist-mediated activation of the beta receptor coupled-adenylyl cyclase (beta AR-AC) system expressed by human airway epithelial cells alters airway function. However, little is known about the magnitude of expression, subtype, and function of the beta receptor-adenylyl cyclase (beta AR-AC) system in human airway epithelial cells from healthy, nonsmoking subjects. Therefore, we characterized beta AR number and subtype and the cAMP response to isoproterenol (iso) in acutely dissociated human tracheocytes harvested from 22 healthy, nonsmoking adults during fibroptic bronchoscopy. Moreover, because the regulation of beta AR-AC system function in response to beta-agonists or inflammatory mediators released into the airway in asthma is poorly understood, we examined the cAMP response to iso after 30 min exposure of cells to iso or the protein kinase C activator, sn-1,2-dioctanoyl glycerol (diC8). The beta AR-AC system was highly expressed and functional in human airway epithelial cells. Group mean beta AR density (i.e., Bmax), equilibrium dissociation constant (Kd), and the percentage of beta 2AR subtypes assessed by radioligand binding were approximately 8,900 receptors/cell, 45 pM, and approximately 80%, respectively. Mean maximum cAMP production was approximately 42 pmol/10(5) cells, and the mean EC50 of the response to iso was 131 nM. However, Bmax and cAMP responses to iso varied considerably across subjects. For example, Bmax varied ninefold, and the EC50 of the cAMP response varied 39-fold interindividually. The EC50 was inversely related to beta AR density (r = -0.81, p < 0.05), suggesting that sensitivity of the cAMP response to iso was in part dependent on beta AR density. In all experiments, cAMP responses to iso stimulation were markedly desensitized in dose-dependent fashion by 30 min pretreatment with iso or diC8. For example, pretreatment with iso 10 microM or diC8 100 microM reduced maximum cAMP production to 22 and 63% of control values, respectively. These data indicate that: (1) the beta AR-AC system is highly expressed on acutely dissociated airway epithelial cells from normal adult, but beta AR expression and its functional coupling to adenylyl cyclase vary considerably interindividually; and (2) the beta AR-AC system of normal human airway epithelial cells is rapidly desensitized by exposure to beta-adrenergic agonists or activators of PKC.

Adenylyl Cyclases↗
PubMed1995

The distribution of Na+,K(+)-ATPase and 5A11 antigen in apical microvilli of the retinal pigment epithelium is unrelated to alpha-spectrin.

The retinal pigment epithelium was used to study the relationship between the cortical cytoskeleton and two plasma membrane proteins that associate with it. These proteins were the Na+,K(+)-ATPase, an ion pump, and the 5A11 antigen, a member of the immunoglobulin superfamily of receptor proteins. The cytoskeleton was marked by two of its constituents, alpha-spectrin and ankyrin. Ankyrin links the Na+,K(+)-ATPase to spectrin in many cells. The RPE is of interest, because unlike most epithelia it distributes the Na+,K(+)-ATPase to the apical membrane. The development of polarity was studied during chick embryogenesis. On embryonic day 6 (E6), each of these proteins was observed in the apical and lateral plasma membranes. As development proceeded, only the Na+,K(+)-ATPase was removed from the lateral membranes. Beginning on E12, ankyrin, spectrin and 5A11 appeared together in patches along the basal plasma membrane. By E16, these patches coalesced into a uniform distribution along the basal membrane. At the apical pole, alpha-spectrin appeared near the base of the microvilli, but was undetected in the microvilli themselves. This distribution resembled the distribution of alpha-spectrin in the intestine and proximal kidney tubule. By contrast, a pool of ankyrin and 5A11 and nearly all the Na+,K(+)-ATPase appeared in the microvilli. Despite its segregation from alpha-spectrin, the Na+,K(+)-ATPase appeared to associate with a macromolecular complex, as judged by extraction with Triton X-100. Changes in spectrin distribution could not be related to changes in isoform expression, as only one isoform of beta-spectrin was detected by co-immunoprecipitation with alpha-spectrin. By contrast, multiple ankyrin-like peptides could be identified by immunoblotting. These data illustrate some of the unique properties of RPE microvilli. These properties prevent the Na+,K(+)-ATPase from complexing with the alpha-spectrin-based cytoskeleton by sequestering the enzyme into the compartment where its activity is required.

Animals↗
PubMed1995

Airway epithelial cell responses to ozone injury.

The airway epithelial cells is an important target in ozone injury. Once activated, the airway epithelium responds in three phases. The initial, or immediate phase, involves activation of constitutive cells, often through direct covalent interactions including the formation of secondary ozonolysis products--hydroxyhydroperoxides, aldehydes, and hydrogen peroxide. Recently, we found hydroxyhydroperoxides to be potent agonists of bioactive eicosanoid formation by human airway epithelial cells in culture. Other probable immediate events include activation and inactivation of enzymes present on the epithelial surface (e.g., neutral endopeptidase). During the next 2 to 24 hr, or early phase, epithelial cells respond by synthesis and release of chemotactic factors, including chemokines--macrophage inflammatory protein-2, RANTES, and interleukin-8. Infiltrating leukocytes during this period also release elastase, an important agonist of epithelial cell mucus secretion and additional chemokine formation. The third (late) phase of ozone injury is characterized by eosinophil or monocyte infiltration. Cytokine expression leads to alteration of structural protein synthesis, with increases in fibronectin evident by in situ hybridization. Synthesis of epithelial antiproteases, e.g., secretory leukocyte protease inhibitor, may also increase locally 24 to 48 hr after elastase concentrations become excessive. Thus, the epithelium is not merely a passive barrier to ozone injury but has a dynamic role in directing the migration, activating, and then counteracting inflammatory cells. Through these complex interactions, epithelial cells can be viewed as the initiators (alpha) and the receptors (omega) of ozone-induced airway disease.

Animals↗
PubMed1995

Activation of eicosanoid metabolism in human airway epithelial cells by ozonolysis products of membrane fatty acids.

Inhaled ozone can react with a variety of cellular macromolecules within the lung. Recent analyses of the chemistry of ozone reactions with unsaturated fatty acids, which are present in all membranes and in mucus in the airways, indicate that ozonolysis yields one aldehyde and one hydroxyhydroperoxide molecule for each molecule of ozone. The hydroxyhydroperoxide molecule is unstable in aqueous environments, and subsequently yields a second aldehyde and hydrogen peroxide. The structure of common unsaturated fatty acids is such that attack by ozone at the carbon-carbon double bonds will yield 3-, 6-, and 9-carbon saturated and unsaturated aldehydes and hydroxyhydroperoxide. This study examines the effects of ozonolysis products on eicosanoid metabolism in human airway epithelial cells. Eicosanoid biosynthesis is important in a wide array of pathophysiological responses in the airway, and the release of eicosanoids by the epithelial barrier is likely to be significant in diseases induced by environmental factors. Previously, we demonstrated that ozone can increase eicosanoid synthesis from airway epithelial cells exposed in vitro. Human exposures to concentrations of ozone below the current National Ambient Air Quality Standard (0.12 ppm, not to be exceeded for more than one hour once per year) also resulted in increased eicosanoids in bronchoalveolar lavage fluid. To determine whether ozonolysis products could activate eicosanoid release, we exposed human airway epithelial cells to 3-, 6-, and 9-carbon aldehydes, hydroxyhydroperoxides, and hydrogen peroxide. We measured (1) eicosanoid metabolism using high-performance liquid chromatography and radioimmunoassays, and (2) the effects of the aldehydes, hydroxyhydroperoxides, and hydrogen peroxide on cell lysis. Eicosanoid release increased after exposure to aldehyde; release induced by 9-carbon (nonanal) aldehyde was greater than that induced by the 6-carbon (hexanal) or 3-carbon (propanal) aldehydes. Hydroxyhydroperoxides induced greater eicosanoid release than the corresponding aldehydes of equivalent chain length. Again, the longer the aliphatic chain length of the hydroxyhydroperoxide the greater the effect. These effects were noted at concentrations of hydroxyhydroperoxide below those that produce cell lysis, and the time course of the two responses was dissimilar. Because hydroxyhydroperoxides can degrade into an aldehyde and hydrogen peroxide, it is conceivable that the effects observed were attributable to the formation of either hydrogen peroxide or hydrogen peroxide and aldehyde. This mechanism is unlikely, however, because the effects of hydroxyhydroperoxides on eicosanoid release were dependent on chain length, whereas each hydroxyhydroperoxide can produce only one hydrogen peroxide molecule. Although hydrogen peroxide alone also stimulated eicosanoid metabolism, this effect was not augmented when aldehyde and hydrogen peroxide were added together. In addition, the dose of hydroxyhydroperoxide needed to produce an effect (10 to 100 microM) was lower than that of hydrogen peroxide (300 microM). We could not fully evaluate the effects of the unsaturated aldehydes and hydroxyhydroperoxides. Although the 6-carbon and 9-carbon cis-3-aldehydes could be synthesized from the cis-3-alcohols, the resulting aldehydes were not chemically stable. The cis-3-aldehydes were useful for producing the corresponding 1-hydroxy-alkenyl-hydroperoxides of high purity. These results support the method selected for chemical synthesis, but further studies are required to establish proper storage and handling methods before these compounds can be tested in assays of eicosanoid metabolism.

Air Pollutants↗
PubMed1995

[Enzyme-linked immunoadsorbent assays for myelin basic protein and antibodies to myelin basic protein in serum and CSF of patients with diseases of the nervous system].

Simplified enzyme-linked immunoadsorbent assays (ELISA) for myelin basic protein (MBP) and antibodies to myelin basic protein (Anti-MBP) have been used to test 337 patients with diseases of the nervous system including 36 compressive diseases (CMP), 33 multiple sclerosis (MS), 34 cerebrovascular diseases (CVD), 31 inflammatory diseases of central nervous system (ID), 161 epilepsy (EP) and 42 other nervous diseases (OND). Comparison of results among various groups indicates that serum mean MBP values of CMP, MS, CVD, ID and EP groups are significantly higher than those of OND group and normal control (P < 0.01). The serum mean MBP value of 33 acute trauma patients with spine fracture and paraplegia, the majority of CMP group, is the highest compared with MS group (P < 0.05), CVD, ID and EP groups (P < 0.01). CSF mean MBP value of 15 CVD patients is markedly greater than that of OND group (P < 0.05). No statistically significant differences are found in serum MBP values between OND group and normal control, and in serum and CSF Anti-MBP values among six groups by using our method.

Autoantibodies↗