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Biomedical subjects

Y Xie

Publications and source records attributed to Y Xie.

At least 361 records · Page 20Linked to original sources
PubMed1995

A new reporter system for detection of retroviral infection.

We describe a novel reporter molecule, the murine surface antigen Thy-1, useful for immunoselection and detection of retrovirus-mediated transduction by flow cytometry. A cDNA encoding the murine thy-1 gene was isolated, and cell surface expression of its gene product was demonstrated. The Thy-1 glycoprotein was tested as a cell surface reporter molecule in the context of replication-defective and -competent retroviruses. Cells transduced via murine retroviral vectors carrying the thy-1 and the neomycin phosphotransferase genes express Thy-1 glycoprotein on their surfaces. The Thy-1 marker is potentially useful in gene transfer protocols because selection of transduced cells can be achieved by immunoselection with anti-Thy-1 antibodies shortly after infection with the retroviral vector. In addition, a human immunodeficiency virus type 1 (HIV-1) recombinant expressing Thy-1 is described, which is replication-competent and syncytium-inducing in human peripheral blood mononuclear cells (PBMCs) and immortalized CD4-positive cell lines. Cells infected with this HIV-1 recombinant express Thy-1 on their surfaces and can be detected and purified by fluorescence-activated cell sorting (FACS). Because of these properties, retroviruses expressing this genetic marker can be useful for studies in gene therapy and of the retroviral life-cycle.

Animals↗
PubMed1995

[Quantitative study of myocardial mitochondrial DNA4977 deletion in rheumatic heart disease and its significance].

To better understand the mechaenisms of cardiac function damaged by chronic hypoxemia in rheumatic heart disease, we investigated the level of the 4977 base pair mitochondrial DNA (mtDNA4977) deletion in 16 patients with rheumatic heart disease (RHD) by using polymerase chain reaction (PCR). Myocardial malondialdehyde (SOD) activity was also measured. In control group, mtDNA4977 deletion appeared after 45 years old and reached a maximum of 0.0042%. A higher level of mtDNA4977 deletion (0.02%-1.26%) was found in RHD group. Because mtDNA4977 deletion removes 8 genes coding for subunits of complexes of respiratory chain, its deletion can inhibit oxidative phosphorylation, reduce ATP production and hinder the recovery of cardiac function. In addition, myocardial MDA content increased and SOD activity decreased in the RHD group as compared with the control group. This indicates that the generation of oxygen free radicals enhances and the elimination of free radicals reduces. Oxygen free radical can be an important factor in myocardial mitochondrial DNA injury and lead to mitochondrial DNA deletion.

Adult↗
PubMed1995

[The effects of 2-chloro-4-bromo-alpha-methylcinnamic acid on the neuromuscular transmission of toad].

The effects of 2-chloro-4-bromo-alpha-methylcinnamic acid sodium (SC1001 Na) on neuromuscular transmission were tested by means of intracellular recordings in isolated sciatic nerve-sartorius preparations of toad. The results were: (1) at a concentration of 0.1mmol/L, SC1001 Na had no effect on end-plate potential (EPP); at a concentration of 1 mmol/L, SC1001 Na reduced the amplitude of the EPP significantly, and the paired-pulse facilitation was also decreased; at a concentration of 10mmol/L, SC1001 Na blocked the EPP completely; (2) the 0.1 mmol/L and 1mmol/L groups of drug had no effect on the resting potential of the toad muscle fibers; the resting potential was reduced significantly by SC1001Na at the concentration of 10mmol/L, the action of 10mmol/L drug was irreversible. These results suggest that SC1001 Na may block the neuromuscular transmission of toad, and the blocking effect might be mainly due to a presynaptic mechanism.

Animals↗
PubMed1995

[Urinary calcium excretion in patients with pregnant hypertension syndrome].

We observed the urinary calcium in 103 patients in the third trimester of pregnancy and the relation between the 24 hour calcium excretion and the calcium/creatinine ratio of a single void urine sample. The study population included 75 normal pregnant women and 28 pregnant woman with pregnant hypertension syndrome. The 24 hour urinary calcium excretion in the patients with pregnant hypertension syndrome was significantly lower (0.70 +/- 0.36 mmol/L) than that in normal pregnant women (4.00 +/- 1.90 mml/L) P < 0.0005. The 24 hour calcium excretion correlated well with the calcium to creatinine ratio of a single void morning urine sample in the two groups. We conclude that the urinary calcium excretion can be used as an indicator for pregnant hypertension syndrome. The 24 hour urinary calcium excretion can be estimated from a single void urine sample. Pregnant hypertension syndrome is associated with hypocalciuria, probably due to increased tubular reabsorption of calcium.

Adult↗
PubMed1995

Nucleotide sequence of maize dwarf mosaic virus capsid protein gene and its expression in Escherichia coli.

The 3'-terminal 1,279 nucleotide sequence of maize dwarf mosaic virus (MDMV) genome has been determined. This sequence contains an open reading frame of 1,023 nucleotides and a 3'-non-coding region of 256 nucleotides. The open reading frame includes all of the coding regions for the viral capsid protein (CP) and part of the viral nuclear inclusion protein (NIb). The predicted viral CP consists of 313 amino acid residues with a calculated molecular weight of 35,400. The amino acid sequence of the viral CP derived from MDMV cDNA shows about 47%-54% homology to that of 4 other potyviruses. The viral CP gene was constructed in frame with the lacZ gene in pUC19 plasmid and expressed in E. coli cells. The fusion polypeptide positively reacted in Western blot with an antiserum prepared against the native viral CP.

Amino Acid Sequence↗
PubMed1995

A novel nonviral cytoplasmic gene expression system and its implications in cancer gene therapy.

We recently have developed a unique cytoplasmic transient gene expression system based on cotransfection of target cells with bacteriophage T7 RNA polymerase (RNAP) and plasmid DNA vectors containing a T7 autogene. Because this T7 system is self-initiating, self-maintaining, and requires no cellular factors for transcription, it is therefore likely to function in any mammalian cell with any gene both in vitro and, more importantly, in vivo. In this study we demonstrate that the T7 DNA vector and T7 RNAP could be efficiently codelivered to cultured cells by lipofection. Different target genes were expressed by the T7 system in a wide variety of mammalian cells including several tumor cell lines. Gene expression could be detected in more than 30% of the cells of some tumor cell lines transiently transfected by the T7 vector. Average activity of the reporter enzyme (luciferase) expressed by a transfected cell was relatively constant regardless of the cell line used. When a T7-luciferase vector was directly injected into various tissues of mice without the use of liposomes, luciferase activity could be found in the injected liver, muscle, brain and tail connective tissues. The luciferase levels expressed by the T7 system were found to be up to 200-fold higher, depending upon the injected tissues, than levels achieved with a traditional nuclear gene expression vector. Direct tumor injection with a T7-beta-galactosidase (beta-gal) construct resulted in beta-gal gene expression in tumor cells near the injection sites. In addition, direct injection of the T7 system in mice did not generate detectable quantities of antibodies against the T7 RNAP. These results suggest that this gene expression system may be useful in many different medical applications such as cancer gene therapies and DNA vaccination, where transient but rapid and efficient gene expression is required.

Animals↗
PubMed1995

[The inhibitory effects of immune sera against two recombinant hybrid antigens on the growth of Plasmodium falciparum in vitro].

The inhibitory effects of rabbit immune sera against two recombinant hybrid antigens (C and CAC) of Plasmodium falciparum to the multiplication and development of in vitro cultured P. falciparum parasites were tested. Although both of anti-C and anti-CAC immune sera had inhibitory effect on parasites, the anti-CAC serum showed much greater effect than that of anti-C serum (P < 0.05). The inhibitory capacities in the sera to parasites were enhanced with the increase in the serum concentration as well as with the prolongation of the incubation time of the sera with the parasites. It was shown that after 72 h incubation of the anti-CAC serum with the parasites at the concentration of 1%, 10% and 20%, the inhibition rates were 15%, 54% and 82%, respectively. The immune sera caused the dispersion of parasite cytoplasm, atrophy of parasites, agglutination of free merozoites and degeneration of schizonts, suggesting that the hybrid antigen could produce multi-functional protective antibodies against antigens of Plasmodium falciparum.

Animals↗
PubMed1995

[A potentially new sodium channel blocker: 2-chloro-4-bromo-alpha-methylcinnamic acid].

The effects of 2-Chloro-4-Bromo-alpha-Methylcinnamic acid sodium (SC1001 Na) and some channel antagonists on the action potential (AP) and the resting potential (RP) were tested intracellularly in the sartorius muscle of the toad (Bufo bufo gargarizans). TEA (10mmol/L) and MnCl2 (10 mmol/L) had no effect on the amplitude of the AP, otherwise, TTX (1 mumol/L) blocked the AP completely. SC1001 Na at the concentration of 2mmol/L largely decreased the amplitude of the AP and expanded the duration of the AP (measured at 1/2 peak amplitude), but it had no effect on the RP. The effects of ms group of drug were reversible. Under the pretreatment of the muscle preparation with TEA (10 mmol/L), SC1001 Na still increased the duration of the AP while decreased its amplitude. At the concentration of 10mmol/L, SC1001 Na completely blocked AP and depolarized the RP significantly. These effects were not reversible. As the structure of SC1001 Na is different from that of heterocycloguanidine Na channel blocker, such as TTX, or that of local anesthetics, we infer that SC1001 Na may be a new sodium channel blocker.

Action Potentials↗
PubMed1995

[The establishment of ADBS data bank system for autopsy records].

The ADBS data bank system consists of PC 286 microcomputer, (or 386 and other more advanced machine) VGA color monitor, printer and ADBS V. 2.0 software. According to the international classification of diseases (ICD-9 or ICD-10, SNOMED), pathological diagnoses are coded and put into computer. This system has five functions: search, statistics, edition, print and system restore. For facilitating the operation of this system, all orders are listed as window menu. The statistic results can be shown as tables or figures and put out as disk files or prints. The system is currently an advanced computerized system of histopathological and autopsied records in Chinese.

Autopsy↗
PubMed1994

A unique sodium-hydrogen exchange isoform (NHE-4) of the inner medulla of the rat kidney is induced by hyperosmolarity.

Membrane sodium-hydrogen exchangers (NHEs), found in virtually all cell types, appear to have diverse and essential roles in regulating cellular pH and mediating vectorial transport by epithelial cells. However, the functional and physiological role of the recently cloned isoform NHE-4 remains unknown. Unlike other Na-H exchanger isoforms, NHE-4 transfected into NHE-deficient mutant fibroblasts demonstrated no amiloride-inhibitable sodium uptake, under basal or acid-loaded isoosmotic conditions. By immunoblot analysis, only the NHE-4 transfectants synthesized a 100-kDa protein, which cross-reacted to polyclonal antibody made to an NHE-4 fusion protein. However, when cells were subjected to acute hyperosmolar cell shrinkage conditions, amiloride-sensitive NHE activity was readily detected at 420 mosm, exhibiting maximal activity at 490 mosm. By in situ hybridization, NHE-4 expression in the rat kidney was found to be limited to the inner renal medullary collecting tubules, the region of highest tissue osmolarity fluctuations in the body. We conclude that NHE-4 is an unusual isoform of sodium-hydrogen exchangers that may play a specialized supplementary role in cell volume regulation.

Animals↗
PubMed1994

Nuclear localization of p185neu tyrosine kinase and its association with transcriptional transactivation.

The rat neu protooncogene encodes a 185 kD transmembrane protein (p185neu), which is a member of the epidermal growth factor receptor (EGFr) family. In searching for the signaling transducer of p185neu by using a two-hybrid selection system, we found, surprisingly, that the cytoplasmic domain of p185neu, when fused to the DNA-binding domain of GAL4 (amino acids 1-147), functioned as a transcriptional activator. We subsequently observed nuclear localization of p185neu. Interestingly, nuclear p185neu has a much higher extent of tyrosine phosphorylation than its nonnuclear counterpart. Our results suggest that a transmembrane receptor tyrosine kinase may enter the nucleus and be involved in transcriptional activation. This novel finding unveils a clue in the understanding of the mechanism of receptor tyrosine kinase-mediated signal transduction.

Animals↗
PubMed1994

Effects of the sodium channel blocker tetrodotoxin (TTX) on cellular ion homeostasis in rat brain subjected to complete ischemia.

Anoxic depolarization (AD) and failure of the cellular ion homeostasis are suggested to play a key role in ischemia-induced neuronal death. Recent studies show that the blockade of Na+ influx significantly improved the neuronal outcome. In the present study, we investigated the effects of 10 microM tetrodotoxin (TTX) on ischemia-induced disturbances of ion homeostasis in the isolated perfused rat brain. TTX inhibited the spontaneous EEG activity, delayed the ischemia-induced tissue acidification, and significantly postponed the occurrence of AD by 65%. The [Ca2+]e elevation prior to AD was attenuated from 17.8% to 6% while the increase of the [Na+]e in this period was enhanced (from 2.9% to 7.3%). These findings implied that the ischemia-induced early cellular sodium load and the corresponding shrinkage of the extracellular space was counteracted by TTX. Our results suggest that the Na+ influx via voltage-dependent channels preceding complete breakdown of ion homeostasis is one major factor leading to cell depolarization. The massive Na+ influx coinciding with AD, however, may be mainly via non-selective cation channels or/and receptor-operated channels. Persistent Na+ influx deteriorates neuronal tissue integrity by favouring Ca2+ influx and edema formation. Blockade of ischemia-induced excessive Na+ influx is, therefore, a promising pharmacological approach for stroke treatment.

Animals↗
PubMed1994

Detection of opiate-enhanced increases in DNA damage, HPRT mutants, and the mutation frequency in human HUT-78 cells.

In previous studies we have shown highly significant increases in chromosome damage and sister chromatid exchanges in heroin addicts, particularly when caffeine and metabolic inhibitors are added to the medium. Using human HUT-78 T-cell cultures, we now find direct in vitro evidence of opiate-induced or opiate-promoted mutagenesis via several assay systems. First, with microgel electrophoresis (MGE), we observed graded, dose-dependent, significant increases (P < .0001) in the frequency of comet tails of fragmented DNA when cells were treated with morphine alone (5 x 10(-9) M up to 10(-7) M) or when co-treated with the more potent mutagen, ethylmethanesulfonate (EMS). There were also dose-dependent increases in the lengths and densities of the comet tails observed. These findings were confirmed by a series of MGE experiments in which several days of morphine exposure preceded a 2-hr pulse of EMS. Second, mutant frequency (MF) assays also indicated significant opiate effects. These studies required separate assessment of cloning efficiencies and the frequencies of TG-resistant, HPRT-deficient mutant clones under four test conditions: no treatment, morphine alone for 4 days, morphine plus EMS, and EMS alone. Prior to the treatment phase, aminopterin was used to eliminate background HPRT mutations. The medium was changed after the treatment phase, the cells were allowed to express mutant phenotypes, and then TG was added and resistant mutant clones counted after 16 days. The background MF level for controls and for cells treated with EMS alone were negligible at 5.12 x 10(-8) and 7.25 x 10(-8), respectively. In the cells treated with morphine alone or morphine plus EMS, MF levels increased very significantly (P < .001) by > 100-fold to 5.1 x 10(-6) and 7.0 x 10(-6), respectively. Cloning efficiency also decreased significantly with both morphine-exposed conditions. Preliminary analysis with the single strand conformational polymorphism (SSCP) procedure following 6-thioguanine (TG) selection, also confirmed the occurrence of Exon 3 mutants of the HPRT gene in cells exposed to morphine plus EMS. It appears that brief EMS exposure can be repaired, whereas, if morphine exposure persists through one or more cell cycles, direct or indirect mutagenesis is initiated.

Analysis of Variance↗
PubMed1994

Chimeric evolution of the 2-microns genome in Saccharomyces cerevisiae.

We compared the nucleotide substitution pattern over the entire genome of two unique variants of the 6,300-bp selfish DNA (2 microns) plasmid in Saccharomyces cerevisiae. The DNA sequence of the left-unique region is identical among 2-microns variants, while the right-unique region shows substantial divergence. This chimeric pattern cannot be explained by neutral or Darwinian selection models. We propose that horizontal transmission of the 2-microns plasmid coupled with a directed, polarized gene conversion maintains the DNA sequence of the left-unique region, whereas the right-unique region is subject to random drift and Darwinian selection.

Base Sequence↗
PubMed1994

Cell-extracellular matrix interactions can regulate the switch between growth and differentiation in rat hepatocytes: reciprocal expression of C/EBP alpha and immediate-early growth response transcription factors.

Previous investigations have shown that culture of freshly isolated hepatocytes under conventional conditions, i.e., on dried rat tail collagen in the presence of growth factors, facilitates cell growth but also causes an extensive down-regulation of most liver-specific functions. This dedifferentiation process can be prevented if the cells are cultured on a reconstituted basement membrane gel matrix derived from the Englebreth-Holm-Swarm mouse sarcoma tumor (EHS gel). To gain insight into the mechanisms regulating this response to extracellular matrix, we are analyzing the activities of two families of transcription factors, C/EBP and AP-1, which control the transcription of hepatic and growth-responsive genes, respectively. We demonstrate that isolation of hepatocytes from the normal quiescent rat liver by collagenase perfusion activates the immediate-early growth response program, as indicated by increased expression of c-jun, junB, c-fos, and c-myc mRNAs. Adhesion of these activated cells to dried rat tail collagen augments the elevated levels of these mRNAs for the initial 1 to 2 h postplating; junB and c-myc mRNA levels then drop steeply, with junB returning to normal quiescence and the c-myc level remaining slightly elevated during the 3-day culture period. Levels of c-jun mRNA and AP-1 DNA binding activity, however, remain elevated from the outset, while C/EBP alpha mRNA expression is down-regulated, resulting in a decrease in the steady-state levels of the 42- and 30-kDa C/EBP alpha polypeptides and C/EBP alpha DNA binding activity. In contrast, C/EBP beta mRNA production remains at near-normal hepatic levels for 5 to 8 days of culture, although its DNA binding activity decreases severalfold during this time. Adhesion of hepatocytes to the EHS gel for the same period of time dramatically alters this program: it arrests growth and inhibits AP-1 DNA binding activity and the expression of c-jun, junB, and c-myc mRNAs, but, in addition, it restores C/EBP alpha mRNA and protein as well as C/EBP alpha and C/EBP beta DNA binding activities to the abundant levels present in freshly isolated hepatocytes. These changes are not due merely to growth inhibition, because suppression of hepatocyte proliferation on collagen by epidermal growth factor starvation or addition of transforming growth factor beta does not inhibit AP-1 activity or restore C/EBP alpha DNA binding activity to normal hepatic levels. These data suggest that expression of the normal hepatic phenotype requires that hepatocytes exist in a G0 state of growth arrest, facilitated here by adhesion of cells to the EHS gel, in order to express high levels of hepatic transcription factors such as C/EBP alpha.

Animals↗
PubMed1994

Glucocorticoids regulate Na+/H+ exchange expression and activity in region- and tissue-specific manner.

Na+/H+ exchangers (NHEs) are integral membrane proteins that exchange Na+ for H+ across membranes. Four isoforms have been cloned (NHE-1-4). NHE-3 localizes to the apical domain, and its expression is increased in dexamethasone-treated rats by Northern analysis. This stimulatory effect on expression is region and tissue specific, being present in ileum and proximal colon, but not in jejunum, distal colon, or kidney. The increase in transcript expression in ileum correlates with an increase in protein expression by immunoblotting. Changes in apical Na+/H+ exchange activity, as measured by 22Na uptake into brush-border membrane vesicles, correlate with expression differences, with significant increases observed in ileum and proximal colon. In situ hybridization showed NHE-3 mRNA only in villus and absorptive cells of control rats, the pattern not being altered by dexamethasone treatment. This suggests that dexamethasone does not increase expression by inducing crypt cells to express NHE-3 prematurely. We conclude that glucocorticoids selectively increase intestinal NHE-3 activity in a region-specific manner and that this effect also appears to be tissue specific.

Adrenalectomy↗
PubMed1994

Na+/H+ exchangers, NHE-1 and NHE-3, of rat intestine. Expression and localization.

Na-H exchange (NHE) is one of the major non-nutritive Na absorptive pathways of the intestine and kidney. Of the four NHE isoforms that have been cloned, only one, NHE-3, appears to be epithelial specific. We have examined the regional and cellular expression of NHE-3 in the rat intestine. NHE-3 message in the small intestine was more abundant in the villus fractions of the small intestine than in the crypts. Analysis of NHE-3 mRNA distribution in the gut by in situ hybridization demonstrated epithelial cell specificity, as well as expression preferential to villus cells. NHE-1 message, in contrast, was ubiquitous, with slightly greater expression exhibited in the differentiating crypt and lower villus cells of the small intestine. Isoform-specific NHE-3 fusion protein antibody identified a 97-kD membrane protein in the upper villus cells of the small intestine, which was exclusively localized in the apical membrane. In contrast, antibody previously developed against the COOH-terminal region of human NHE-1 (McSwine, R. L., G. Babnigg, M. W. Musch, E. B. Chang, and M. L. Villereal, manuscript submitted for publication) identified a 110-kD basolateral membrane protein. These data suggest that unlike NHE-1, which probably serves a "housekeeping" function, NHE-3 may be involved in vectorial Na transport by the intestine.

Animals↗