Biomedical subjects

S Zhao

Publications and source records attributed to S Zhao.

At least 379 records · Page 21Linked to original sources

PubMed1997

Construction and identification of a recombinant adenovirus which expresses human interferon-gamma.

An E1-deleted adenoviral vector, pAdl2/RSV-IFN-bpA, harboring human IFN-gamma cDNA, and plasmid pJM17 were cotransfected into a human embryo kidney cell line 293 by the calcium-phosphate precipitation method. Six strains of recombinant adenovirus (rAd) were obtained after 11-day incubation. PCR analysis indicated that all six rAds contained human IFN-gamma cDNA. The biological activity of IFN-gamma was detected in the supernatant of the cultured 293 cells following rAd infection, and it could be blocked by the antibody against IFN-gamma. The B16F10 cells infected with the purified rAd (RSV/IFN-01) didn't show cytopathic effect (CPE), and more biological activity of IFN-gamma was detected in the supernatant of the infected B16F10 cells with increased MOI (multiple of infection) values. Therefore, the recombinant adenovirus constructed in this study can express and secrete human IFN-gamma.

Adenoviruses, Human↗

PubMed1997

[A new subtype of potato class I patatin gene].

Potato class I and class II patatin gene belong to a multigene family with different tissuespecific expression patterns. A class I patatin gene was isolated from a genomic library of Chinese potato cultivar "Dongnong 303" (Solanum tuberosum) by using class II patatin promoter as probe. A 1.8kp DNA sequence was determined, including 1407bp of patatin gene 5' flanking sequence and 363bp of patatin structure gene. Its 5' flanking sequence showed that it is a class I patatin gene. Compared with six reported subtypes of class I patatin genes, there are a large fragment of deletion and insertion in its 5' flanking sequence. Therefore, it might be a new subtype of class I patatin gene.

Base Sequence↗

PubMed1997

Responsiveness of cytosolic free calcium concentration in cultured rat pulmonary arterial smooth muscle cells: confocal microscopic measurement.

Rat pulmonary arterial smooth muscle cells were primarily cultured. alpha-actins in pulmonary arterial smooth muscle cells at day 7 of culture were observed by an immunofluorescence staining method using anti-alpha-actin antibody. Microfluorimetry of Ca2+ signals in fluo-3-loaded single smooth cell at day 7 of culture was performed by a laser-scanned confocal imaging system. The effects of several kinds of Ca(2+)-mobilizing drugs on cytoplasmic Ca2+ concentration ([Ca2+]i) were examined. KCl, a depolarizing agent, and norepinephrine, an alpha-adrenergic agonist, equally increased [Ca2+]i. Angiotensin II, a receptor agonist, and caffeine, a Ca(2+)-induced Ca2+ releaser, elevated [Ca2+]i in the same manner but was more potent than KCl and norepinephrine. Br-A23187, a Ca2+ ionophore, most potently increased [Ca2+]i. The present results suggest that drug receptors on plasma membrane, Ca2+ entry pathways and Ca(2+)-releasing mechanisms act normally, and that our cultured pulmonary arterial smooth muscle cells may be a good model for the study on the essential role of Ca2+ in vasoconstriction.

Adrenergic alpha-Agonists↗

PubMed1997

Changes in tachykinin--ergic nerve terminal densities in rat nasal mucosa during hypersensitivity.

OBJECTIVE: To further carry out a quantitative immunohistochemical study on neuropeptides (NPs) participation in perennial allergic rhinitis (PAR) in nasal mucosa. METHODS: By using light microscopy (routine hematoxylin and eosin staining, Wright's staining and 1% toluidine blue staining) and scanning as well as transmission electron microscopy, an exhaustive immunohistochemical and morphometric study was made on the changes of substance P (SP), neuropeptide A (NKA) and neuropeptide B (NKB) in the nasal mucosa of a toluene-2,4-iso-cyanate (TDI)-induced PAR model of rat. RESULTS: The results showed that the densities of all three peptidergic terminals were significantly increased (P < 0.01) in experimental group (115.72 +/- 28.43 for SP, 39.23 +/- 10.34 for NKA and 37.24 +/- 11.22 for NKB, respectively) as compared with those in control group (49.65 +/- 11.23 for SP, 12.44 +/- 6.21 for NKA and 13.31 +/- 7.60 for NKB, respectively). Increased staining, thickening of peptidergic terminals and enlargement of varicosities were found. Mast cell infiltration in nasal mucosa was in parallel with increase of peptidergic terminals. CONCLUSION: All results support the hypothesis that NP is a newly-recognized but potentially important factor in the pathogenesis of PAR.

Animals↗

PubMed1997

[Computed tomography of the anterior ethmoid sinuses and its adjacent structures for endoscopic sinus surgery].

Forty adult cadavers were examined by computed tomography (CT) imaging with special attention to the pneumatization of the anterior ethmoid sinus for endoscopic sinus surgery. The study of the specimen was done using coronal section and axial section, sagittal reconstructed images were obtained from the data acquired in the axial plane. The cells of anterior ethmoid sinus were classified into four sub-groups, namely, the pre-infundibular cell (appearance rate 87.5%), the lateral infundibular cell (82.7%), the post-infundibular cell (86.3%) and the bulla cell (100.0%), and the anterior ethmoid sinus was classified into two types, namely, intraethmoid type (71.2%) and extraethmoid type (28.8%). The results shows that CT can display the sub-group and the type of the anterior ethmoid sinus and its adjacent structures clearly. The significance of endoscopic sinus surgery was discussed.

Adult↗

PubMed1997

Identification of a novel missense mutation in Wilson's disease gene.

OBJECTIVE: To investigate the allelic heterogeneity of the ATP7B gene in Chinese patients with Wilson's disease (WD). METHODS: Exons of the ATP7B gene from 141 WD patients' DNA were amplified with polymerase chain reaction (PCR) 887-890. Mutations were then screened by single strand conformation polymorphism (SSCP) analysis and further identified by sequencing. RESULTS: The molecular structure of exon 7 of the ATP7B gene from 141 WD patients was analyzed. The same band shift in electrophoretic pattern of 4 cerebral type patients was identified with SSCP and subsequently sequenced. The results showed missense mutation at the second base of the codon as Ser 662 Cys, which is caused by a C to G transversion. CONCLUSIONS: Mutations of the ATP7B gene were investigated for the first time in China and a novel missense mutation was identified in four cases.

Adolescent↗

PubMed1997

[Relation of Lac promotor and the expression of cholera toxin subunit B gene in recombinant Escherichia coli MM2].

Effects of different carbon sources including glucose, lactate and acetate and IPTG induction on the expression of ctb gene, which is on the downstream of lac promotor, in recombinant Escherichia coli MM2 were studied. In medium YC were added 0.048mol/L glucose, 0.102mol/L lactate or 0.167mol/L acetate which separately produce the same energy in the condition of complete oxidization. Addition of glucose largely decreased the expression level of ctb gene because of decrease of pH during culture process. Addition of lactate increased the expression level of ctb gene by 1.15 fold and did not inhibit the growth of MM2 strain. Addition of acetate increasd the expression level of ctb gene by 0.97 fold and inhibited the growth of MM2 strain. Induction by IPTG at different time and different concentration did not increase the expression level of ctb gene, so the lac promotor had no or a little influence upon the expression of ctb gene in recombinant MM2 strain.

Acetates↗

PubMed1997

[Using lyophilization to the preparation of drug paper disks for antimicrobial susceptibility test].

It was found that the paper disks used in the antimicrobial susceptibility test could be prepared by natural drying at 37 degrees C, but the drug distribution between these disks is not so enen as determined by antimicrobial inhibitory test, thus it was difficult to fit the needs of the national standards. Alternatively, we applied the lyophilization technique to the disks preparation and 12 batches, of different antibiotic disks were made out and found that the difference between disks is reduced, and all are meet to the national requirement. It is supposed that the capillary action may play a role in the variation of drug distribution in the natural drying process, and the lyophilization process avoided the capillary action, hence the more even distributed antibiotic disks are produced. The preparation of lyophilized disks is a simple and practical method that could be used in the antimicrobial susceptibility tests in hospital.

Freeze Drying↗

PubMed1997

[Clinical evaluation of polyamine assay in diagnosis of malignant ascites].

Polyamine levels were determined by precolumn derivation RH-HPLC in 68 patients with ascites including 21 malignant ascites, 28 liver cirrhosis and 19 tuberculosis. The total polyamine, especially the contents of cadaverine (CA) and spermidine (SPD) in malignant ascites were significantly higher than that in cirrhosis and tuberculosis. Among them CA levels in malignant ascites and cirrhotic and tubercular ascitic fluid were 2.26 +/- 1.83 nmol.ml-1, 0.75 +/- 0.52 nmol.ml-1 and 0.76 +/- 0.47 nmol.ml-1 respectively, demonstrated the most obvious difference. Using 1.0 nmol.ml-1 as a threshold cut off point to differentiate the malignant ascites from non-malignant ascites, the from non-malignant ascites, the sensitivity, specificity and accuracy reached 76. 2%, 83.0% and 80.9% respectively. The results suggest that polymine level in ascites might be regard as one of the cancer-associated markers.

Ascites↗

PubMed1997

[Lipoprotein(A) distribution among adults and its relation to the incidence of coronary artery disease in Changsha City].

The plasma lipoprotein (a) [Lp(a)] levels were determined among 593 adults in Changsha city, Hunan province. The result showed that the distribution of Lp(a) levels in all the individuals was a skewness which can be normalized by data transformation into logarithm. Plasma Lp (a) concentration > 300 mg.L-1 was 34.8% in 209 subjects with coronary artery disease (CAD) and 17.3% in 305 health controls (P < 0.01). The Lp(a) levels were significantly higher in subjects with CAD than in health controls. Multiple regression stepwise analysis found that Lp(a) levels were positively correlated with CAD. This study indicates that plasma levels of Lp(a) are strongly related to CAD and may be an important independent risk factor for CAD.

Adult↗

PubMed1996

Interaction of a novel sex-dependent, growth hormone-regulated liver nuclear factor with CYP2C12 promoter.

CYP2C12 is a steroid hydroxylase cytochrome P450 whose female-specific expression in adult rat liver is transcriptionally activated by the continuous plasma growth hormone (GH) profile characteristic of adult female rats. DNase I footprinting and gel mobility shift analysis of the 5'-flank of the CYP2C12 gene were carried out to identify cis-acting elements and trans-acting factors that may contribute to the GH-regulated, sex-dependent transcription of this P450 gene. DNase I footprinting analysis revealed sex- and GH-regulated DNase I hypersensitivity sites at the boundaries of several protein binding sites detected along a 1560-nucleotide upstream segment of CYP2C12. Five distinct sites bound a novel continuous GH-regulated nuclear factor, GHNF, which is enriched in adult female and continuous GH-treated male liver nuclear extracts compared to untreated male liver nuclear extracts. Two other footprinted sites correspond to binding sites for the liver transcription factors C/EBP and albumin D element-binding protein and a third to an HNF1 binding site. A specific binding site for GHNF was also found in the 5'-proximal promoter of CYP2C11, an adult male-specific liver P450 gene, suggesting that GHNF may contribute to the down-regulation of that gene by continuous GH. GHNF was distinguished from the nuclear factors that bind to a GH response element upstream of the rat Spi 2.1 gene and is also distinct from the GH-activatable latent cytoplasmic transcription factors STAT 1, STAT 3, and STAT 5. These findings support the hypothesis that continuous GH-activated transcription of CYP2C12 in adult female rat liver (a) involves the activation of a novel GH-regulated nuclear factor which binds to multiple sites along the 5'-flank of this cytochrome P450 gene, and (b) proceeds via a signaling pathway distinct from the GH pulse-activated STAT5 pathway proposed to induce CYP2C11 and other male-expressed liver genes.

Animals↗

PubMed1996

Putative human blue-light photoreceptors hCRY1 and hCRY2 are flavoproteins.

Recently, a human cDNA clone with high sequence homology to the photolyase/blue-light photoreceptor family was identified. The putative protein encoded by this gene exhibited a strikingly high (48% identity) degree of homology to the Drosophila melanogaster (6-4) photolyase [Todo et al. (1996) Science 272, 109-112]. We have now identified a second human gene whose amino acid sequence displays 73% identity to the first one and have named the two genes CRY1 and CRY2, respectively. The corresponding proteins hCRY1 and hCRY2 were purified and characterized as maltose-binding fusion proteins. Similar to other members of the photolyase/blue-light photoreceptor family, both proteins were found to contain FAD and a pterin cofactor. Like the plant blue-light photoreceptors, both hCRY1 and hCRY2 lacked photolyase activity on the cyclobutane pyrimidine dimer and the (6-4) photoproduct. We conclude that these newly discovered members of the photolyase/photoreceptor family are not photolyases and instead may function as blue-light photoreceptors in humans.

Amino Acid Sequence↗

PubMed1996

Involvement of P-glycoprotein in the transmembrane transport of interleukin-2 (IL-2), IL-4, and interferon-gamma in normal human T lymphocytes.

The physiological role of the multidrug resistance P-glycoprotein (P-gp), which is expressed by normal human T lymphocytes, is still largely unknown. To investigate whether or not P-gp is involved in the transport of cytokines, peripheral blood lymphocytes were stimulated with phytohemagglutinin (PHA) in the absence or presence of P-gp inhibitors, and concentrations of cytokines (interleukin-2 [IL-2], IL-4, IL-6, interferon-gamma [IFN-gamma]) in the supernatants of these cultures were quantitated by enzyme-linked immunosorbent assay. P-gp inhibitors included verapamil (Ver), tamoxifen (Tmx), and the P-gp specific monoclonal antibody UIC2. Release of IL-2 was significantly suppressed by these inhibitors at concentrations that were also effective in blocking efflux of Rhodamine-123 from normal T lymphocytes. IL-2 mRNA expression in lymphocytes was not different between PHA control and the cultures with P-gp inhibitors. Ver and Tmx did not interfere with T-cell activation as determined by CD25 and CD69 expression. In a nonhematological model, the P-gp expressing HCT-8 adenocarcinoma cell line, exogenously added IL-2 was shown to exert an inhibitory effect on P-gp mediated Rhodamine-123 efflux. In addition, transepithelial transport of IL-2 by electrophysiologically tight and polarized HCT-8 monolayers was examined. A time-dependent flux of IL-2 across dense monolayers, which was partially inhibited by Ver, was observed. We also investigated whether or not P-gp inhibitors suppressed release of other cytokines produced by activated T cells (IL-4, IL-6, IFN-gamma). Release of IL-4 and IFN-gamma was significantly inhibited by Ver, Tmx, and UIC2; however, release of IL-6 remained unaffected. These data show P-gp mediated transmembrane flux of IL-2 in T lymphocytes and HCT-8 cells. We conclude that P-gp participates in the transport of cytokines (IL-2, IL-4, and IFN-gamma) in normal peripheral T lymphocytes.

ATP Binding Cassette Transporter, Subfamily B, Mem↗

PubMed1996

The novel anthracycline annamycin is not affected by P-glycoprotein-related multidrug resistance: comparison with idarubicin and doxorubicin in HL-60 leukemia cell lines.

A major factor in limiting the efficacy of anthracyclines is overexpression of the MDR1-encoded p-glycoprotein (p-gp). A new analogue less affected by p-gp is annamycin (ANN), an anthracycline antibiotic with high affinity for lipid membranes and significantly more activity than doxorubicin (DOX). We investigated whether ANN was affected by p-gp-mediated multidrug resistance (MDR) by comparing the cellular accumulation and retention of ANN, idarubicin (IDR), and DOX in the p-gp-negative human leukemia cell lines (HL-60S) and its DOX-selected p-gp-positive subline (HL-60/DOX) with and without verapamil (VER). As expected, HL-60/DOX cells showed lower DOX uptake than HL-60S cells; coincubation with VER (10 mmol/L) increased uptake 2.6-fold restoring it to 100% of uptake in HL-60S cells. IDR uptake increased 1.5-fold in the presence of VER, but ANN was not affected. Coincubation with VER increased DOX retention in HL-60/DOX cells 2.8-fold and IDR retention 1.4-fold; unchanged ANN retention indicated that ANN may overcome p-gp. In the cytotoxicity assay to correlate intracellular anthracycline content with antitumor activity, we found ANN to be less potent than DOX and IDR In sensitive cells, ID 50 being the drug concentration that inhibits cell growth by 50% but its resistance index (RI; ID50 resistant cells divided by ID50 sensitive cells) was lower than that of IDR and DOX (2.6 v 40 and 117.5). Coincubation in the presence of VER resulted in 4.5-fold and 2-fold RI decreases of DOX and IDR, respectively, whereas ANN did not change, further confirming ANN's ability to circumvent p-gp-mediated MDR. Confocal microscopy studies of IDR, ANN, and DOX showed higher intracellular drug compartmentalization for DOX in HL-60/DOX cells incubated in the presence of VER. This study provided evidence that, unlike DOX and IDR, ANN is not affected by p-gp-mediated MDR.

ATP Binding Cassette Transporter, Subfamily B, Mem↗

PubMed1996

Differential effects of bFGF on development of the rat retina.

A variety of growth factors can influence the expression of differentiated properties by cell types of the developing retina. One unresolved question has been whether these factors can direct the differentiation pathway of uncommitted precursors or whether they act to help the expression of properties by already committed cells. To address this question we have studied the effects of basic fibroblast growth factor (bFGF) on the differentiation of ganglion cells and rod photoreceptors in explant cultures of embryonic rat retinas. Incubation of retinas in the presence of bFGF accelerated the appearance of differentiated ganglion cells and incubation in the presence of anti-bFGF antibodies delayed the appearance. bFGF had no effect on the appearance of differentiated rod photoreceptors as judged by expression of opsin, although all-trans-retinoic acid did increase the number of cells expressing opsin. bFGF inhibited the formation of rod photoreceptor rosettes suggesting that it does influence some properties of rods or the adjacent Müller glial cells. The results suggest that bFGF can alter the timing of differentiation of retinal ganglion cells but not direct their production from retinal precursors.

Animals↗

PubMed1996

A conformational change in the methyltransferase from Clostridium thermoaceticum facilitates the methyl transfer from (6S)-methyltetrahydrofolate to the corrinoid/iron-sulfur protein in the Acetyl-CoA pathway.

The methyltetrahydrofolate:corrinoid/iron-sulfur protein methyltransferase (MeTr) from Clostridium thermoaceticum catalyzes the methylation of a corrinoid/iron-sulfur protein (C/Fe-SP) by the N5 methyl group of (6S)-methyltetrahydrofolate (CH3-H4folate). This is an important reaction in the reductive acetyl-CoA pathway. The forward and reverse reactions of MeTr have a pH dependence that appears to reflect protonation of a group on the protein [Zhao, S., Roberts, D. L., & Ragsdale, S. W. (1995) Biochemistry 34, 15075-15083]. In the work reported here, fluorescence and rapid reaction kinetics were used to demonstrate that this protonation elicits a rate-limiting conformational change. As the pH was lowered, the emission maximum for intrinsic tryptophan fluorescence underwent a red shift (pK(a) = 5.4) and the emission intensity increased (pK(a) = 5.1). The extrinsic fluorescence probe, 4,4'-bis-1-phenylamino-8-napthalenesulfonate (bis-ANS) was used to report on the conformational change. The bis-ANS fluorescence was strongly enhanced upon binding MeTr. As the pH was decreased, the fluorescence was further enhanced and the emission maximum underwent a 14 nm blue shift (pK(a) = 5.0). By stopped-flow fluorescence studies, it was shown that these fluorescence changes occur at rates similar to the k(cat) for the MeTr reaction and thus reflect catalytically competent events. The combined results indicate that CH3-H4folate binds to a hydrophobic region in MeTr that includes a tryptophan residue(s). MeTr undergoes a pH-dependent conformational change that exposes this region to solvent and facilitates substrate binding.

Acetyl Coenzyme A↗

PubMed1996

Affinity chromatography of regulatory subunits of protein phosphatase-1.

In this study we demonstrate that recombinant rabbit muscle protein phosphatase-1 immobilized on CH-Sepharose is an efficient affinity chromatography support for the isolation of subunits of phosphatase-1. The support was used to isolate the glycogen binding subunit of phosphatase-1 from muscle and nonmuscle rat tissues. Examination of the affinity-purified material from rat heart and liver showed that the major component was a 160-kDa polypeptide on SDS-PAGE. The identity of the purified liver 160-kDa polypeptide as the glycogen binding subunit was confirmed by the demonstration that it is capable of binding to glycogen, and is phosphorylated by the catalytic subunit of PKA. The novel observation was made that the phosphorylation was dependent on the presence of glycogen. Examination of the material from heart, lung, liver, kidney, and brain showed a similar phenomenon. Our studies show that this subunit is widely distributed in tissues. The affinity support was also efficient in the isolation of the NIPP-1 (nuclear inhibitor of protein phosphatase-1) proteins from calf thymus. Examination of heat-treated extracts of rat liver led to the isolation of a novel 19-kDa inhibitory protein which could also be phosphorylated by PKA and may represent the rat liver homolog of calf thymus NIPP-1.

Animals↗

PubMed1996

Polymerase chain reaction for detecting Escherichia coli O157: H7.

Escherichia coli O157:H7 is known as an important cause of hemorrhagic colitis and hemolytic uremic syndrome. Real-time procedures that are sensitive for detecting small populations of this bacterium in food are lacking and needed. An expression library was constructed by ligation of BamHI-EcoRI DNA fragments of E. coli O157:H7 to plasmid vector pUC19 and transformation of recombinant plasmids to E. coli JM109. A clone that contained a specific DNA fragment of E. coli O157:H7 was identified by colony immunoblot assay using monoclonal antibody MAb 4E8C12 that uniquely links to E. coli O157:H7 and a few other serotypes of verotoxin-producing E. coli. The DNA sequence of the clone consisted of 110 bp of 5' region of enterohemorrhagic E. coli (EHEC) eae gene and a 688 bp DNA fragment adjacent to 5' end of the eae gene, including an unknown function gene encoding 156 amino acids. A pair of oligonucleotide primers was synthesized based on the sequence of the 688 bp fragment. The primers were used in a polymerase chain reaction (PCR) to amplify a target DNA of 633 bp. The primers amplified 1 ng of DNA from 67 strains of E. coli O157:H7, two strains of E. coli O157:NM, and 7 of 11 E. coli O55:H7 and O55:NM strains, but not 50 ng of DNA from 34 strains of 29 other E. coli serotypes and 25 strains of 8 other bacterial species. Annealing temperatures from 60 to 63 degrees C could be used for the PCR without loss of specificity. The minimum amount of target DNA detected by the PCR was 5 pg. When a boiling method and GeneReleaser were used, the PCR was able to detect as few as 25 and 38 CFU of E. coli O157:H7, respectively, in 3 h.

Antibodies, Monoclonal↗