Biomedical subjects

L Jiang

Publications and source records attributed to L Jiang.

At least 127 records · Page 7Linked to original sources

PubMed1999

[Loss of heterozygosity and microsatellite instability in laryngeal carcinoma near p16 gene]

OBJECTIVE: To search for the minimal overlap region of tumor suppressor gene in laryngeal carcinoma and discuss the correlation of p16 gene with laryngeal carcinogenesis. METHODS: Five microsatellite polymorphism markers near p16 gene were selected to detect loss of heterozygosity (LOH) and microsatellite instability (MI) in 60 cases of laryngeal squamous cell carcinoma. RESULTS: The frequencies of LOH in 5 markers were less than 23.1%, while the frequencies of MI in 2 markers were higher, with the highest frequency (46.1%) in D9S1752. CONCLUSION: The results suggest that the deletion of p16 gene does not play an important role in the laryngeal carcinogenesis and there may exist a gene around D9S1752 participating in the development of laryngeal carcinoma, which correlates with the mutation of repair gene.

Journal Article↗

PubMed1999

On the importance of being aromatic at an antibody-protein antigen interface: mutagenesis of the extracellular interferon gamma receptor and recognition by the neutralizing antibody A6.

A complex formed between the extracellular human interferon gamma receptor alpha-chain (hIFNgammaR) and the Fab fragment of the neutralizing antibody A6 has been studied by site-directed mutagenesis. Five complementarity determining regions of the A6 antibody interact primarily with the CC' surface loop of the receptor, from Lys47 to Trp56, although contact is also made with residues in the neighbouring F strand, in particular with Trp82. The relative contribution that individual side-chains make to complex stabilization was assessed with 21 receptors mutants, whose affinity for A6 was monitored using a surface plasmon resonance biosensor, as well as by solution-phase competition ELISA. The results reveal two lysine side-chains (Lys47 and Lys52), an asparagine side-chain (Asn53), and two aromatic side-chains (Tyr49 and Trp82) in the receptor that are important for recognition by A6. The role of aromatic side-chains in antibody-antigen recognition is of particular interest, not least in this case because 13 aromatic groups (six Tyr, six Trp and one His) are present at the interface (four in VL, six in VH and three in the receptor), and several are proximal to the charged and polar side-chains of Lys47, Lys52 and Asn53 in the receptor. The results highlight the possibility for aromatic rings to participate in networks of co-operative interactions with not only hydrophobic, but also charged and hydrogen bond donor and acceptor groups, properties that are well suited for creating binding sites for protein epitopes, regardless of the distribution of polar and non-polar surface residues. These findings may contribute, therefore, to an understanding of how surface groups on proteins are captured by the often aromatic-rich hypervariable loops of antibodies, and may be of value for the design of molecules with novel recognition properties.

Antibodies, Monoclonal↗

PubMed1999

Induction of nitric oxide synthase mRNA by shear stress requires intracellular calcium and G-protein signals and is modulated by PI 3 kinase.

We have investigated the signaling pathways by which shear stress induces accumulation of endothelial nitric oxide synthase (eNOS) mRNA in bovine aortic endothelial cells (BAEC). Steady laminar fluid shear stress (20 dyn/cm2) induced a time-dependent increase in eNOS mRNA levels that did not require de novo protein synthesis and was in part transcriptional. Shear responsiveness was conferred on a luciferase reporter by a portion of the eNOS gene promoter encoding the 5'-flanking region between nt -1600 and -779. Shear-mediated induction of eNOS mRNA was abolished by chelation of intracellular calcium ([Ca2+]i) with BAPTA-AM, and inhibited by blockade of calcium entry with SKF96535. In contrast, eNOS mRNA upregulation by shear was potentiated by thapsigargin-mediated depletion of Ca2+i stores. Pertussis toxin (PTX) inhibited both the shear-induced elevation in [Ca2+]i and the subsequent increase in eNOS mRNA, implicating a PTX-sensitive G-protein in both responses. Shear-induced upregulation of eNOS mRNA was unaffected by the calmodulin inhibitor W-7 and by the tyrosine kinase inhibitor herbimycin A, suggesting that neither calmodulin nor tyrosine kinases are required. However, eNOS mRNA upregulation was potentiated by the PI 3-kinase inhibitors wortmannin and LY294002, suggesting that PI 3-kinase inhibits the shear response. Although microtubule integrity is required for the shear-induced regulation of endothelin-1 mRNA and the morphological and cytoskeletal responses to flow, neither microtubule dissolution with nocodazole nor microtubule stabilization with taxol altered shear-induced [Ca2+]i elevation or upregulation of eNOS mRNA. In conclusion, shear stress of BAEC increases eNOS transcriptional rate and upregulates eNOS mRNA levels by a process that requires calmodulin-independent [Ca2+]i signaling and a PTX-sensitive G-protein, is inhibited by PI 3-kinase, and is independent of microtubule integrity and tyrosine kinase activity.

Animals↗

PubMed1999

Electron spin resonance studies on photosensitized formation of hydroxyl radical by C-phycocyanin from Spirulina platensis.

Visible light (>470 nm) irradiation of an oxygen-saturated solution of C-phycocyanin (C-PC) in the presence of the spin trap 5, 5-dimethyl-1-pyrroline-N-oxide (DMPO) gave an ESR spectrum characteristic of the DMPO-hydroxyl radical spin adduct DMPO-OH. The signal intensities of DMPO-OH adduct were enhanced by superoxide dismutase (SOD) and partly inhibited by catalase. It was partly responsible for the production of DMPO-OH that superoxide anion radical (O.-2) dismutated to generate hydrogen peroxide (H2O2) which decomposed ultimately to generate the highly reactive .OH. In addition, it can be concluded that singlet oxygen (1O2) was an important intermediate according to the strong inhibitory action of 1,4-diazabicyclo[2.2.2]octane (DABCO) and histidine on DMPO-OH formation. The experimental results suggest that photodynamic action of C-PC proceed via both type I and type II mechanisms. Furthermore, the decay kinetics of DMPO-OH adduct, the effects of DMPO and C-PC concentrations as well as irradiation time on DMPO-OH adduct formation were also discussed. Concentration of C-PC should be an important factor to influence the ESR signal intensities of DMPO-OH. Therefore, it may be concluded that reasonably lower concentration of C-PC might prolong the duration of photosensitized formation of .OH and might strengthen the photodynamic action.

Cyclic N-Oxides↗

PubMed1999

Comparative studies of Nsc and Fmoc as N(alpha)-protecting groups for SPPS.

2-(4-Nitrophenyl)sulfonylethoxycarbonyl (Nsc) is an alternative base-labile N(alpha)-protecting group to 9-fluorenylmethoxycarbonyl (Fmoc) for amino acids. The UV spectrum of the Nsc group exhibits moderate absorption at 380 nm which is excellent for real-time monitoring of the deprotection process. It also decreases the rearrangement of X-Asp, which can be a serious problem in SPPS.

Amino Acids↗

PubMed1999

Elevated DNase activity and caspase expression in association with apoptosis in failing ischemic sheep left ventricles.

We examined the DNase activity as well as the expression of the caspases in left ventricles of failing sheep hearts and compared it with samples from normal controls. The sheep model of chronic ischemic heart failure was developed by intracoronary microembolisation. Moderate heart failure was defined as a decrease of left ventricular (LV) ejection fraction (EF) to 35% or less and was stable for four weeks after the last embolisation. The failing hearts were harvested from these animals six months later. Bovine pancreas DNase I was used as a standard for the DNase zymograms. In normal sheep left ventricles, a low level of the DNase I-like activity was detected (1.16 +/- 0.17 pg/100 microg protein, n = 20), whereas it was elevated (3.64 +/- 0.49 pg/100 microg, n = 32, p < 0.001) in the ischemic failing LV samples. The identity of the DNase in sheep is yet to be fully characterised. However, we also detected the expression of caspase-2 and caspase-3 by Western blotting. There was a 2.2-fold increased expression of caspase-2 p < 0.001 = and a 2.6-fold increased expression of caspase-3 p < 0.001) in failing left ventricles compared to normal samples.

Animals↗

PubMed1999

Structural features and expression of an alternatively spliced growth factor type I receptor for follitropin signaling in the developing ovary.

The pleiotropic actions of pituitary follitropin (FSH), regulate the expression of many cell cycle genes controlling ovarian follicular development and differentiation. In this study we asked the question whether different receptor motifs are created by the alternative splicing of the single large 80-100 Kb receptor gene. A 1.2 Kb transcript identified from a cDNA library of hormone primed (immature) sheep ovaries, codes for a putative protein lacking the seven transmembrane segment. The receptor of 259 amino acids designated FSH-R3 is derived from a transcript comprising the first eight exons of the Gs coupled larger FSH receptor (R1) spliced to another DNA segment. This event produces a different carboxyl terminus at the junction creating a novel receptor motif with a single membrane spanning domain, assigning it to the growth factor type I receptor family. In transfected cells the expressed receptor localizes on the cell surface and specific antibodies directed against the unique C-terminal portion (residues 242-259) of FSH-R3 demonstrate the presence of the receptor protein in solubilized ovarian and testicular membrane preparations. FSH binding to the transfected cells induced [Ca2+]i identifying coupling of the R3 receptor to calcium signaling pathways. Thus, a growth factor type I receptor for FSH may be implicated in the growth promoting actions of FSH in the ovary. This is the first documentation of alternative splicing of a G protein coupled receptor gene creating a different signaling motif for cellular signaling.

Alternative Splicing↗

PubMed1999

A linear endothelin-1 analogue: solution structure of ET-1[Aib1,3,11,15, Nle7] by nuclear magnetic resonance spectroscopy and molecular modelling.

Two-dimensional nuclear magnetic resonance techniques and a combination of distance geometry and molecular dynamics calculations were utilised to determine the three dimensional solution structure of an ET-1 analogue, ET-1[Aib1,3,11,15, Nle7], in a methanol-d3/water co-solvent. The modelled structure shows that the peptide folds into a consistent alpha-helical conformation between residues Ser4-His16 while the C-terminus prefers no fixed conformation. Our studies confirm that the disulphide links which are normally associated with the endothelin family of neuropeptides are not important for the formation of a helical conformation in solution. This full length, modified, synthetic linear ET-1 analogue plays a vital role towards designing endothelin receptor agonists. Structure activity relationships are discussed in terms of the conformational features of the calculated structure.

Amino Acid Sequence↗

PubMed1999

Real-time visualization of myocardial perfusion and wall thickening in human beings with intravenous ultrasonographic contrast and accelerated intermittent harmonic imaging.

Previous work has demonstrated that at higher peak negative pressures, microbubbles are destroyed by diagnostic ultrasonography. At lower pressures (lower mechanical index), less destruction occurs but enhanced contrast persists. In animals, this lower mechanical index has resulted in enhanced contrast after administration of intravenous microbubbles with intermittent imaging at faster frame rates. We tested whether this accelerated intermittent imaging technique could produce myocardial contrast and detect myocardial perfusion abnormalities in 25 patients (10 with normal wall motion, 15 after myocardial infarction). Three independent reviewers detected persistent myocardial contrast defects within the infarct zone throughout the cardiac cycle in 9 of the 15 patients after acute myocardial infarction; the presence of such defects was predictive of a persistent regional wall motion abnormality at 4-week follow-up. Interobserver agreement on regional contrast enhancement ranged from 88% to 90%. We conclude that accelerated intermittent imaging permits real-time visualization of myocardial blood flow and wall thickening.

Contrast Media↗

PubMed1999

Multiple-quantum HCN-CCH-TOCSY experiment for 13C/15N labeled RNA oligonucleotides.

A multiple-quantum 3D HCN-CCH-TOCSY experiment is presented for the assignment of RNA ribose resonances. The experiment makes use of the chemical shift dispersion of N1 of pyrimidine and N9 of purine to distinguish the ribose spin systems. It provides an alternative approach for the assignment of ribose resonances to the currently used COSY- and TOCSY-type experiments in which either 13C or 1H is utilized to distinguish the different spin systems. Compared to the single-quantum version, the sensitivity of the multiple-quantum HCN-CCH-TOCSY experiment is enhanced on average by a factor of 2 for a 23-mer RNA aptamer complexed with neomycin.

Framycetin↗

PubMed1999

Solution structure of a novel ETB receptor selective agonist ET1-21 [Cys(Acm)1,15, Aib3,11, Leu7] by nuclear magnetic resonance spectroscopy and molecular modelling.

The solution structure of a biologically active modified linear endothelin-1 analogue, ET1-21[Cys(Acm)1,15, Aib3,11, Leu7], has been determined for the first time by two-dimensional nuclear magnetic resonance spectroscopy in a methanol-d3/water solvent mixture. Out of approximately one hundred linear peptide analogues tested by biological assay, this peptide, together with a dozen others, showed significant ETB selective agonist activity. Here we report the solution structure of an ETB selective agonist of a full-length, synthetic linear endothelin analogue. The calculated structures indicate that the peptide adopts an alpha-helical conformation between residues Ser5-His16, whilst both N- and C-termini show no preferred conformation. These results suggest that the disulphide bridges normally associated with endothelin and sarafotoxin peptides may not necessarily be important for either ETB receptor binding activity or the formation of a helical conformation in solution.

Endothelin-1↗

PubMed1999

Functional analysis of a Golgi-localized Kex2p-like protease in tobacco suspension culture cells.

Kex2p is the prototype of a Golgi-resident protease responsible for the processing of prohormones in yeast and mammalian cells. A Kex2p-like pathway was shown to be responsible for processing the fungal KP6 protoxin in transgenic tobacco plants. We previously described a chimeric integral membrane reporter protein that traffics through Golgi to the lytic prevacuole where it was proteolytically processed. As a first step to isolate and clone the Kex2p-like protease in plant cells, we designed and used a similar chimeric reporter protein containing Kex2 cleavage sites to assay the Kex2p-like activity and to determine its substrate specificity in tobacco cells. Here we demonstrate that the Kex2 cleavage sites of the reporter were specifically processed by a protease activity with a substrate specificity characteristic of yeast Kex2p. This Kex2p-like protease in tobacco cells is also a Golgi-resident enzyme. Thus, the reporter protein provides a biochemical marker for studying protein traffic through the Golgi in plant cells. These results additionally should allow the design of synthetic substrates for use in biochemical purification of the plant enzyme.

Amino Acid Sequence↗

PubMed1999

Dietary modulation of intestinal fructose transport and GLUT5 mRNA expression in hypothyroid rat pups.

BACKGROUND: Intestinal fructose transport rates or GLUT5 mRNA levels typically show a two- to threefold increase after weaning in rats allowed to wean normally but can be enhanced precociously by high-fructose diets during early weaning. Developmental increases in serum thyroxine levels coincide with the onset of weaning and have been linked to changes in intestinal sucrase and lactase activities. METHODS: Rat pups were made hypothyroid by giving the dam 0.01% propylthiouracil as drinking water from day 18 of gestation. The hypothyroid pups and age-matched euthyroid control pups were then fed high-fructose or high-glucose solutions by gavage, twice a day starting at 17 days of age for 3 days, and then killed at 20 days of age. RESULTS: Serum thyroxine levels were five times lower in the hypothyroid pups. Rates of intestinal fructose uptake in the proximal and middle small intestine were 2.0 to 2.5 times higher in the hypothyroid and euthyroid pups fed high-fructose solution than in littermates fed high-glucose solution or those allowed to wean normally with the dam. Intestinal glucose uptake also increased in hypothyroid but not in euthyroid pups fed high-fructose or high-glucose solutions. GLUT5 mRNA levels increased in euthyroid and hypothyroid pups fed high fructose and paralleled the increase in fructose uptake. CONCLUSION: During weaning, dietary fructose can precociously enhance intestinal fructose uptake and GLUT5 mRNA expression, independent of developmental increases in serum thyroxine levels. Modest changes in glucose transport rates indicate that nonspecific mechanisms may provide a minor contribution to diet-induced changes in nutrient absorption in hypothyroid pups.

Animals↗

PubMed1999

Computer-aided three-dimensional reconstruction and measurement of the optic canal and intracanalicular structures.

OBJECTIVE: To reconstruct the human optic canal and its inner structures and to provide detailed knowledge of this region for optic nerve decompression. METHODS: Six optic canals and their inner structures were reconstructed using a computer-aided three-dimensional reconstruction system. Quantitative measurement of the canal wall thickness, bony canal transverse area, optic nerve transverse area, dural sheath transverse area, subarachnoid space transverse area, and subarachnoid space volume was done using the computer morphometric analysis system. The detailed spatial relationship among intracanalicular structures was also carefully identified on the three-dimensional models. RESULTS: The thinnest portion of the canal was the middle part of the medial wall (0.45 +/- 0.14 mm) and the narrowest space was in the middle part of the optic canal (the transverse area was 18.21 +/- 1.20 mm2). The volume of subarachnoid space that can be considered the compensatory space for distention incurred by the hemorrhage, optic nerve edema, or hematoma was 21.16 +/- 4.31 mm3. At the cranial opening, the middle part, and the orbital opening, its transverse area was 4.45 +/- 0.46 mm2, 2.68 +/- 0.54 mm2, and 1.23 +/- 0.34 mm2 respectively. CONCLUSIONS: Because the compensatory space was limited, even a tiny amount of blood or swelling of the nerve may cause optic nerve compression. Because the compensatory space for distention gradually decreases from cranial end to orbital end, the middle part and the anterior part of the optic canal and dural sheath are critical in optic nerve decompression.

Adult↗

PubMed1999

Breath ethane as a marker of reactive oxygen species during manipulation of diet and oxygen tension in rats.

Breath ethane, O2 consumption, and CO2 production were analyzed in 24-mo-old female Fischer 344 rats that had been fed continuously ad libitum (AL) or restricted 30% of AL level (DR) diets since 6 wk of age. Rats were placed in a glass chamber that was first flushed with air, then with a gas mixture containing 12% O2. After equilibration, a sample of the outflow was collected in gas sampling bags for subsequent analyses of ethane and CO2. The O2 and CO2 levels were also directly monitored in the outflow of the chamber. O2 consumption and CO2 production increased for DR rats. Hypoxia decreased O2 consumption and CO2 production for the AL-fed and DR rats. These changes reflect changes in metabolic rate due to diet and PO2. A significant decrease in ethane generation was found in DR rats compared with AL-fed rats. Under normoxic conditions, breath ethane decreased from 2.20 to 1.61 pmol ethane/ml CO2. During hypoxia the levels of ethane generation increased, resulting in a DR-associated decrease in ethane from 2.60 to 1.90 pmol ethane/ml CO2. These results support the hypothesis that DR reduces the level of oxidative stress.

Aging↗

PubMed1999

The expression of proUK in Escherichia coli: the vgb promoter replaces IPTG and coexpression of argU compensates for rare codons in a hypoxic induction model.

The expression of the proUK gene was improved by the coexpression of the argU gene cloned in a moderate copy number vector. As the proUK gene contains 2% AGG/AGA codons, which is much higher than the normal frequency in E. coli, about 0.14%-0.21%, the argU gene cloned in a multicopy plasmid was coexpressed with the proUK expression vector in our experiments. In E. coli strain BL21(DE3), IPTG is known to induce the expression of T7 RNA polymerase gene and this enzyme can transcribe the proUK gene under the control of the T7 promoter leading to expression of proUK. To replace IPTG by a cheaper alternative on a large scale, we constructed a plasmid in which the vgb promoter--which is known to be activated by the onset of hypoxic conditions--controls the T7RNA polymerase gene expression. Low oxygen conditions were then used to activate the vgb promoter causing T7RNA polymerase gene expression and finally leading to the expression of proUK as inactive inclusion bodies. Our experiments on a large scale in a bioreactor show that the expression of proUK accounts for about 30% of total protein after about 6 h of anaerobic cultivation, so the presented model represents an economical alternative to IPTG induction.

Bacterial Proteins↗

PubMed1999

[Loss of heterozygosity and microsatellite instability in laryngeal carcinoma near p16 gene].

Gene Deletion↗

PubMed1999

[Gene transfer efficiency enhancement by inactivated Coxsakie virus B1 in primary skeletal muscle cells].

OBJECTIVE: To study the effect of inactivated Coxsackie virus B1(CVB1), which was considered to have high affinity to skeletal muscle cell, on gene transfer efficiency of reporter gene, pCDNA3-LacZ, in primary muscle cells. METHODS: Viruses were purified by polyethylene sedimentation followed by sucrose supercentrifugation. Virus, transferrin-polylysine(TfpL) and LacZ were designed as different concentrations and groups, then positive cells contained beta-galactosidase were calculated as gene transfer efficiency. RESULTS: Inactivated CVB1 made an enhancement of gene transfer efficiency from less than 1% in TfpL+LacZ group to 15-20 per cent in CVB1+TfpL+LacZ group. CONCLUSION: Like adenovirus, inactivated CVB1 can also increase gene transfer efficiency in muscle cells, which provides experimental basis for constructing new vector targeting to muscle cells.

Cells, Cultured↗