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Biomedical subjects

R Wu

Publications and source records attributed to R Wu.

At least 307 records · Page 17Linked to original sources
PubMed1992

Thioredoxin gene expression is transcriptionally up-regulated by retinol in monkey conducting airway epithelial cells.

Using the differential hybridization technique, a cDNA clone, MT78, was isolated from the cDNA library of retinol-treated monkey tracheobronchial (TBE) epithelial cells. MT78 has a high sequence homology to human thioredoxin. The cDNA insert contains 506 nucleotides which encodes a peptide of 105 amino acids. The deduced peptide contains the highly conserved sequence Cys-Gly-Pro-Cys, found at the active site of all thioredoxins. The expression of the thioredoxin gene is stimulated 8-10 fold by vitamin A (retinol) in monkey TBE cells. The expression is significantly enhanced within 4 h after the vitamin A treatment and concurrent protein synthesis is not required for this enhancement. These results, in conjunction with the nuclear run-on transcriptional assay, support the conclusion that thioredoxin gene is transcriptionally up-regulated by retinol and/or its metabolites.

Amino Acid Sequence↗
PubMed1992

Regulation and interaction of multiple protein factors with the proximal promoter regions of a rice high pI alpha-amylase gene.

The alpha-amylase gene is known to be regulated by the plant hormone gibberellin (GA) in cereal aleurone cells. The accumulation of the mRNA corresponding to a rice high pI alpha-amylase gene, OSamy-c, was stimulated 20-fold by exogenous GA3 in half-seeds lacking embryos. Regulatory regions in the promoter of this high pI sub-family were analyzed. The OSamy-c 5' flanking sequence, spanning positions -231 to +29, was fused upstream of the beta-glucuronidase (GUS) gene coding region. The delivery of this plasmid into rice aleurone cells by the biolistic method resulted in a GA-stimulated synthesis of GUS. Gel retardation assays were performed to study protein-DNA interactions between putative regulatory sequences of OSamy-c and partially purified rice seed extracts. We identified multiple seed-specific protein factors that bind to proximal regions of the OSamy-c promoter between positions -231 and -162. Five different proteins were distinguished based on competitive binding studies. Three protein binding regions were located by footprinting analyses, one of which is located in the conserved sequence also found upstream of other GA-inducible genes. Two protein factors in rice aleurone cells that interact with the putative regulatory sequence do not require GA induction.

Base Sequence↗
PubMed1992

Incidence and risk factors for side effects of spinal anesthesia.

We prospectively studied 952 patients to identify the incidence of hypotension (systolic blood pressure less than 90 mmHg), bradycardia (heart rate less than 50 beats/min), nausea, vomiting, and dysrhythmia during spinal anesthesia. Historical, clinical, and physiologic data were correlated with the incidence of these side effects by univariate and multivariate analysis. Hypotension developed in 314 patients (33%), bradycardia in 125 (13%), nausea in 175 (18%), vomiting in 65 (7%), and dysrhythmia in 20 (2%). Variables conferring increased odds of developing hypotension include peak block height greater than or equal to T5 (odds ratio 3.8, P less than 0.001), age greater than or equal to 40 yr (2.5, P less than 0.001), baseline systolic blood pressure less than 120 mmHg (2.4, P less than 0.001), combination of spinal and general anesthesia (1.9, P = 0.01), spinal puncture at or above the L2-L3 interspace (1.8, P less than 0.001), and addition of phenylephrine to the local anesthetic (1.6, P = 0.02). Variables conferring increased odds of developing bradycardia include a baseline heart rate less than 60 beats/min (odds ratio 4.9, P less than 0.001), ASA physical status classification of 1 versus 3 or 4 (3.5, P less than 0.001), current therapy with beta-adrenergic blocking drugs (2.9, P less than 0.001), and peak block height greater than or equal to T5 (1.7, P = 0.02). Variables conferring increased odds of developing nausea or vomiting include addition of phenylephrine or epinephrine to the local anesthetic (3.0-6.3, P less than or equal to 0.003), peak block height greater than or equal to T5 (odds ratio 3.9, P less than 0.001), use of procaine (2.6-4.4, P less than or equal to 0.003), baseline heart rate greater than or equal to 60 beats/min (2.3, P = 0.03), history of carsickness (2.0, P = 0.01), and development of hypotension during spinal anesthesia (1.7, P = 0.009). Our results indicate that the incidence of side effects during spinal anesthesia may be reduced by 1) minimizing peak block height; 2) using plain solutions of local anesthetics; 3) performing the spinal puncture at or below the L3-L4 interspace; and 4) avoiding the use of procaine in the subarachnoid space.

Adult↗
PubMed1992

Expression of the repA1 gene of IncFII plasmid NR1 is translationally coupled to expression of an overlapping leader peptide.

Examination of a group of mutants of plasmid NR1 that had lost the expression of IncFII plasmid incompatibility (Inc-) revealed a group that had also lost replication proficiency (Rep-). These mutants were obtained from plasmids in which the NR1 replication control region was present in a cointegrate with plasmid pBR322. Whereas the wild-type parental cointegrate plasmid was capable of replicating in a polA host owing to the PolA independence of NR1 replication, the mutants were not able to transform a polA host. Losses of both expression of IncFII plasmid incompatibility and replication proficiency were found to result from the same single base-pair substitution in four independently isolated Inc- Rep- mutants. The mutation inactivates promoter PE for the transcription of RNA-E, a trans-acting repressor of translation of the essential RepA1 replication initiation protein of NR1. Although the loss of RNA-E synthesis had been expected to increase the expression of repA1, the efficiency of translation of repA1 mRNA from these mutants was at least 100-fold lower than that from the wild type, as revealed by repA1-lacZ translational fusions. The PE mutation introduced a stop codon into a 24-amino-acid reading frame that precedes the repA1 gene and terminates just 2 bp downstream from the repA1 start codon. This putative leader peptide was also expressed in a lacZ translational fusion, and its expression was reduced by a factor of 10(4) by the PE mutation. The expression of the leader peptide and the expression of repA1 were regulated by RNA-E. These results suggest that the expression of repA1 is coupled to the translation of the leader peptide and that the repression of repA1 translation by RNA-E may occur via inhibition of the translation of the leader peptide.

Amino Acid Sequence↗
PubMed1992

Characterization of cis-acting elements regulating transcription from the promoter of a constitutively active rice actin gene.

The promoter of the constitutively expressed rice (Oryza sativa) actin 1 gene (Act1) is highly active in transformed rice plants (W. Zhang, D. McElroy, and R. Wu, Plant Cell 3:1150-1160, 1991). A region 834 bp upstream of the Act1 transcription initiation site contains all the regulatory elements necessary for maximal gene expression in transformed rice protoplasts (D. McElroy, W. Zhang, J. Cao, and R. Wu, Plant Cell 2:163-171, 1990). We have constructed a series of Act1 promoter deletions fused to a bacterial beta-glucuronidase reporter sequence (Gus). Transient expression assays in transformed rice protoplasts, as well as transformed maize cells and tissues, identified two distinct cis-acting regulatory elements in the Act1 promoter. A 38-bp poly(dA-dT) region was found to be a positive regulator of Act1 promoter activity. Deletion of the poly(dA-dT) element lowered Gus expression by at least threefold compared with expression produced by the full-length Act1 promoter. By gel retardation and footprinting, we identified a ubiquitous rice protein which specifically recognizes this poly(dA-dT) element in the constitutively active Act1 promoter. A CCCAA pentamer repeat-containing region was found to be a negative regulator of the Act1 promoter in transformed rice protoplasts. Transient expression assays in different maize cells and tissues with use of the Act1 deletion constructs suggested that the CCCAA pentamer repeat region functions in a complex tissue-specific manner. A CCCAA-binding protein was detected only in root extracts.

Actins↗
PubMed1992

Induction of T-cell activation by oxidized low density lipoprotein.

Oxidation and scavenger receptor-mediated uptake of low density lipoprotein (LDL) in intimal macrophages are believed to be key events in the development of atherosclerosis. We report here that oxidized LDL increases DNA synthesis, expression of HLA-DR, and interleukin-2 receptors in T cells. The stimulatory effect of oxidized LDL was not due to a direct effect on T cells but required the presence of monocytes. Oxidized LDL also stimulated the release of interleukin-1 beta from monocytes. The maximal effect of oxidized LDL on T-cell activation and interleukin-1 beta release occurred at a concentration of 1 micrograms/ml. Native LDL also had the capacity to activate T cells, although only at higher concentrations. The stimulatory effect of both native and oxidized LDL was inhibited by superoxide dismutase. Monocytes as well as T cells were found to have the ability to oxidize LDL, suggesting that the stimulatory effect of native LDL may arise as a result of LDL oxidation during incubation with monocytes and T cells. The results suggest that oxidized LDL may activate T cells in atherosclerotic lesions.

Antigens, Surface↗
PubMed1992

Ozone-induced acute tracheobronchial epithelial injury: relationship to granulocyte emigration in the lung.

To investigate the relationship between granulocyte emigration and epithelial injury in specific airway generations of the tracheobronchial tree following short-term ozone exposure, we exposed rhesus monkeys for 8 h to 0.00 (controls) or 0.96 ppm ozone with post-exposure periods of 1, 12, 24, 72, and 168 h in filtered air before necropsy. There were five control and three exposed monkeys for each of the post-exposure times for a total of 20 monkeys. Neutrophils isolated from peripheral blood and labeled with 111In-tropolonate were infused in the cephalic vein in unanesthetized monkeys (except the 1-h group) 4 to 5 h before necropsy. The trachea and microdissected bronchi (fourth and ninth generations) and respiratory bronchioles (fifteenth generation) from the right upper lobe of each monkey were examined by electron microscopy. Labeled neutrophil influx into lung tissue and bronchoalveolar lavage fluid (BALF) was maximal at 12 h and returned to baseline by 24 h after exposure. This was in contrast to total neutrophils (labeled and unlabeled) in BALF, which were significantly elevated through 24 h after exposure but returned to baseline by 72 h. Lavage protein was significantly elevated at 24 h after exposure but was at control levels at all other times. Morphometric observations showed epithelial necrosis at 1 and 12 h in the trachea and bronchioles but continued to be observed in significant numbers at 24 h after exposure in bronchi. A significant increase in the labeling index of epithelial cells was observed at 12 h only in bronchi. Epithelial necrosis and repair was associated with the presence of granulocytes in the epithelium and interstitium of all airway levels. However, eosinophils were maximally increased in the epithelium and interstitium of bronchi at 24 h after exposure when epithelial necrosis was maximal in these airways and when lavage protein was significantly elevated.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1992

An unusual expression of a squamous cell marker, small proline-rich protein gene, in tracheobronchial epithelium: differential regulation and gene mapping.

An unusual expression of a putative squamous cell marker, small proline-rich protein (spr1), in mucociliary epithelial cells of conducting airways was demonstrated in a serum-free culture system. A cDNA clone was isolated from the cDNA library of monkey tracheobronchial epithelial (TBE) cells by differential hybridization. This cDNA clone, MT5, exhibited 98% homology to a DNA sequence obtained from human keratinocytes treated with either UV light or phorbol esters (T. Kartasova et al., 1988, Mol. Cell. Biol. 8:2195-2230). The predicted peptide of MT5 is unusual for its high content of proline (29%), glutamine (18%), and cysteine (9%) and its repeated PKVPEPC units. The level of spr1 mRNA in cultured cells was inhibited more than 90% by vitamin A. In contrast, phorbol 12-myristate 13-acetate (PMA) stimulated the level of spr1 mRNA by 3- to 8-fold. This differential regulation coincided with the effects of these chemicals on the cornification of cultured TBE cells. Using MT5 as a probe, we have localized the tracheal spr1 gene on the human chromosome 1 by a Southern blot analysis using a panel of human-rodent somatic cell hybrid DNAs. The gene was further sublocalized to bands q22-23 by in situ hybridization.

Amino Acid Sequence↗
PubMed1992

Characterization of the mucin differentiation in human lung adenocarcinoma cell lines.

Four human lung adenocarcinoma cell lines were established in serum-free F12 medium supplemented with insulin, transferrin, hydrocortisone, cholera toxin, selenium, epidermal growth factor, bovine hypothalamic extract, and retinoic acid. Histochemical analyses with periodic acid-Schiff with and without diastase treatment (PAS-D technique) and immunocytochemistry with a mucin-specific monoclonal antibody demonstrated that three of the cell lines (CL2, CL3, and NCL2) were capable of mucin production. Biochemical characterizations of mucin produced by adenocarcinoma cells were focused on one of the cell lines, CL2 cells, which showed the most prominent reactivity with mucin-specific monoclonal antibody. Biochemical analysis using the mucin precursors [3H]glucosamine and [14C]serine indicated that CL2 cells can synthesize high-molecular-weight (M(r) greater than 200 kD) glycoprotein molecules that can be immunoprecipitated by this mucin-specific monoclonal antibody. The high-molecular-weight glycoproteins isolated from CL2 cells specifically reacted with mucin-specific monoclonal antibody by Western blot analysis, and composition analyses showed high levels of serine and threonine and a low level of aromatic amino acids, which are similar to human airway mucin. These observations suggest that lung adenocarcinoma CL2 cells cultured in this serum-free medium can retain function of airway mucin synthesis. Cell kinetic studies of these four cell lines showed that the cell line (CL1) without the mucin differentiation had a higher proliferative index and a shorter population doubling time as compared with the other three cell lines (CL2, CL3, and NCL2) with mucin differentiation. Examination of the retinoblastoma protein expressions in these adenocarcinoma cell lines revealed a phosphorylated pattern that correlated inversely with the mucin synthesis status of these cell lines.(ABSTRACT TRUNCATED AT 250 WORDS)

Adenocarcinoma↗
PubMed1992

Regulatory mode of the pac gene expression.

1.65kb HindIII-BgIII DNA fragment containing pac operator was cloned from plasmid pPA4 to pBR322 at HindIII-BamHI sites, the resultant plasmid pPA41 was transformed into E. coli D816 carrying an intact pac operon on its chromosome, then the effect of operator titration was estimated. It was found that the penicillin acylase activity in E. coli D816 (pPA41) cells was higher than that of in E. coli D816 cells. The operators on high copy number plasmid pPA41 competed for the regulatory proteins with the single copy operator on the chromosomal pac operon, thus the expression of the pac gene was enhanced, because the free regulatory proteins were decreased by operator titration. The results of RNA-DNA hybridization showed that the cellular pac mRNA concentration was parallel to the penicillin acylase activity, and the pac mRNA in E. coli D816 (pPA41) cells was much higher than that of in E. coli D816 cells. These results indicated that the expression of pac gene was negatively controlled at transcriptional level.

Biotechnology↗
PubMed1992

Cloning of par region and the effect of par region on the stability of pUC9.

The par region of pSC101 was cloned into pUC9, and pEC302 was obtained. The stability of pEC302 and pUC9 in E. coli HB101 was tested on M9 medium to determine the effect of par region on the stability of plasmid. After 40 generations, pUC9 was almost completely lost, while pEC302 was 100% maintained in host cells.

Biotechnology↗
PubMed1992

Location and trans-action of the pac regulatory gene.

The study on the localization of the pac gene indicated that the regulatory and structural genes were located in a 3.5 kb HindIII-EcoRI fragment. In this paper we report that a series of deletion derivatives of plasmid pPA6 was constructed and the effect of deletions on the pac expression was determined. The results suggested that the pac regulatory gene was located in the SphI-PvuII fragment within the pac structural gene. Then the SphI-PvuII fragment was cloned to the plasmid pTZ18U at the SphI-SmaI sites and the resultant plasmid pPA57 was transformed into the penicillin acylase producing strain E. coli D816 to observe the effect of SphI-PvuII fragment on the chromosomal pac expression. The results demonstrated that the regulatory gene within the SphI-PvuII fragment regulated the pac expression in trans. Computer analysis indicated that there were two ORFs in the SphI-PvuII fragment as possible candidates coding for the regulatory protein. The study on precise localization of the pac regulatory gene is in progress.

Amino Acid Sequence↗
PubMed1992

Characterization of the enantiomers of cis-N-[2-(3,4-dichlorophenyl)ethyl]-N-methyl-2-(1- pyrrolidinyl)cyclohexylamine (BD737 and BD738): novel compounds with high affinity, selectivity and biological efficacy at sigma receptors.

A novel class of compounds with very high affinity and selectivity for sigma receptors has been discovered. BD614 [(+/-)-cis-N-[2-(3,4-dichlorophenyl)ethyl]-N-methyl-2-(1- pyrrolidinyl)cyclohexylamine] and its optically pure 1S,2R-(-)-[BD737] and 1R,2S-(+)-[BD738]enantiomers bound to sigma receptors of guinea pig brain with Ki = 2.0 +/- 0.4, 1.3 +/- 0.3 and 6 +/- 3 nM, respectively. These compounds exhibited little or no affinity for dopamine-D2, kappa opiate or phencyclidine receptors and displayed high biological efficacy in assays of sigma receptor function, ability to produce alterations in motor behavior and inhibition of the muscarinic cholinergic phosphoinositide response. Microinjection of BD614 into the rat red nucleus or substantia nigra produced a dose-dependent alteration in head position and contralateral circling, respectively. BD614, BD737 and BD738 inhibited stimulation of inositol phosphate production by carbachol or oxotremorine-M in a dose-dependent manner. Thus, N-substituted cis-2-(1-pyrrolidinyl)cyclohexylamines may prove useful in studies of sigma receptor structure and function.

Analgesics↗