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Biomedical subjects

Q Ye

Publications and source records attributed to Q Ye.

At least 109 records · Page 6Linked to original sources
PubMed1998

Mutations of gamma-aminobutyric acid and glycine receptors change alcohol cutoff: evidence for an alcohol receptor?

Alcohols in the homologous series of n-alcohols increase in central nervous system depressant potency with increasing chain length until a "cutoff" is reached, after which further increases in molecular size no longer increase alcohol potency. A similar phenomenon has been observed in the regulation of ligand-gated ion channels by alcohols. Different ligand-gated ion channels exhibit radically different cutoff points, suggesting the existence of discrete alcohol binding pockets of variable size on these membrane proteins. The identification of amino acid residues that determine the alcohol cutoff may, therefore, provide information about the location of alcohol binding sites. Alcohol regulation of the glycine receptor is critically dependent on specific amino acid residues in transmembrane domains 2 and 3 of the alpha subunit. We now demonstrate that these residues in the glycine alpha1 and the gamma-aminobutyric acid rho1 receptors also control alcohol cutoff. By mutation of Ser-267 to Gln, it was possible to decrease the cutoff in the glycine alpha1 receptor, whereas mutation of Ile-307 and/or Trp-328 in the gamma-aminobutyric acid rho1 receptor to smaller residues increased the cutoff. These results support the existence of alcohol binding pockets in these membrane proteins and suggest that the amino acid residues present at these positions can control the size of the alcohol binding cavity.

Alcohols↗
PubMed1998

Neurosteroids act on the GABA(A) receptor at sites on the N-terminal side of the middle of TM2.

Two sets of chimeras between alphaxalone-sensitive GABA(A) receptor alpha2 or beta1 subunits and the alphaxalone-insensitive glycine receptor alpha1 subunit were constructed to determine the structural domains important for the modulatory actions of neuroactive steroids. These data suggest that the site of action for neurosteroids on GABA(A) receptors is not the same as that for volatile anesthetics and ethanol, but is on the N-terminal side of the middle of TM2.

Anesthetics↗
PubMed1998

Enhancement of glycine receptor function by ethanol is inversely correlated with molecular volume at position alpha267.

Glycine and gamma-aminobutyric acid (GABA)A receptors are members of the "superfamily" of ion channels, and are sensitive to allosteric modulation by n-alcohols such as ethanol and butanol. We recently demonstrated that the mutation of Ser-267 to Ile in the alpha1 subunit abolished ethanol regulation of glycine receptors (Gly-R). In the present study, a pair of chimeric receptors was studied, in which a 45-amino acid domain comprising transmembrane domains 2 and 3 was exchanged between the Gly-Ralpha1 and gamma-aminobutyric acid rho1 subunits. Detailed pharmacologic analysis of these chimeras confirmed that this domain of the Gly-R confers enhancement of receptor function by ethanol and butanol. An extensive series of mutations at Ser-267 in the Gly-Ralpha1 subunit was also prepared, and the resulting homomeric receptors were expressed and tested for sensitivity to glycine, and allosteric modulation by alcohols. All of the mutant receptors expressed successfully in Xenopus oocytes. Mutation of Ser-267 to small amino acid residues such as Gly or Ala produced receptors in which glycine responses were potentiated by ethanol. As we have reported previously, the mutant Gly-Ralpha1 (Ser-267 --> Ile) was completely insensitive to ethanol; mutation of Ser-267 to Val had a similar effect. Mutation of Ser-267 to large residues such as His, Cys, or Tyr resulted in inhibition of Gly-R function by ethanol. These results demonstrate that the size of the amino acid residue at position alpha267 plays a crucial role in determining the functional consequences of allosteric modulation of the Gly-R by alcohols.

Animals↗
PubMed1998

The effects of steric mutations on the structure of type III antifreeze protein and its interaction with ice.

The interaction of proteins with ice is poorly understood and difficult to study, partly because ice is transitory and can present many binding surfaces, and partly because structures have been determined for only two ice-binding proteins. This paper focuses on one of these, a 66-residue antifreeze protein (AFP) from eel pout. The high resolution X-ray structure of this fish AFP demonstrated that the proposed ice-binding surface is remarkably flat for such a small protein. The residues on the planar surface thought to be involved in ice binding are restrained by hydrogen bonds or by tight packing of their side-chains. To probe the requirement for a flat binding surface, a conserved alanine in the center of the AFP planar surface was substituted with larger residues. Six alanine replacement mutants (Ala16 > Cys, Thr, Met, Arg, His and Tyr), designed to disrupt the planarity of the surface and sterically block binding to ice, were characterized by X-ray crystallography and compared with the wild-type AFP. In each case, the detail provided by these crystal structures has helped explain the effects of the mutation on antifreeze activity. The substitutions, Ala16 > His and Ala16 > Tyr, were large enough to shield Gln44, one of the putative ice-binding residues, contributing to their very low thermal hysteresis activity. In addition to sterically hindering the putative ice-binding site, the bulkier residues also caused shifts in the putative ice-binding residues owing to the tight packing of side-chains on the planar surface. This unexpected consequence of the mutations helps account for the severely reduced antifreeze activity. One explanation for residual antifreeze activity in some of the mutants lies in the possibility that AFPs have a role in shaping the site on the ice to which they bind. Thus, side-chain dislocations might be partially accommodated by ice that can freeze around them. It is evident that the disruption of the planarity, by introducing larger residues at the center of the proposed ice-binding site, is not the only factor responsible for the loss of antifreeze activity. There are multiple causes including positional change and steric blockage of some putative ice-binding residues.

Alanine↗
PubMed1998

Fluorescence activated cell sorting: a reliable method in tissue engineering of a bioprosthetic heart valve.

BACKGROUND: Techniques of tissue engineering are used to seed human autologous cells in vitro on degradable mesh to create new functional tissue like a bioprosthetic heart valve. A precondition is subsequent seeding of native-valve-analogous pure endothelial and myofibroblast cell lines. The aim of this study is to find a safe method of isolating viable cell lines out of tissues from the operating room. METHODS: Mixed cells from ascending aorta obtained from the operating room were incubated with an endothelial-specific fluorescent marker. The labeled cells were activated and sorted by flow cytometry. Isolated cell lines were cultured and thereafter square sheets of polymeric scaffold were seeded with myofibroblasts, followed by endothelial cells. The created tissue was stained with hematoxylin and eosin, van Gieson stain, and stains for factor VIII and CD34. RESULTS: Control culture samples (n = 25) revealed vital uncontaminated endothelial and myofibroblast cell lines. Microscopy of the seeded meshes (n = 16) demonstrated a tissue-like structure. Van Gieson stain showed production of collagen. Endothelial cells formed a superficial monolayer, demonstrated by factor VIII and CD34; no invasive formation of capillaries was detectable. CONCLUSIONS: These results demonstrate that fluorescence activated cell sorting is a reliable and safe method to gain pure vital autologous cell lines out of human mixed cells for subsequent seeding on degradable mesh and that those cells are active to form new tissue.

Aorta↗
PubMed1998

Oxidation of eugenol to form DNA adducts and 8-hydroxy-2'-deoxyguanosine: role of quinone methide derivative in DNA adduct formation.

We have investigated the activation of eugenol to form DNA adducts and oxidative base damage. Treatment of myeloperoxidase containing HL-60 cells with eugenol, produced a dose-dependent formation of three DNA adducts as detected with P1-enhanced 32P-post-labeling. Incubation of HL-60 cells with the combination of 100 microM eugenol and 100 microM H2O2 potentiated the levels of DNA adduct in HL-60 cells by 14-fold, which suggests peroxidase activation in adduct formation. In vitro activation of eugenol with either horseradish peroxidase or myeloperoxidase and H2O2 produced three DNA adducts that were inhibited by the addition of either ascorbic acid or glutathione, by 66 and 90%, respectively. The DNA adducts formed in HL-60 cells treated with eugenol were the same as those formed by in vitro peroxidase activation. In addition to adduct formation, peroxidase activation of eugenol produced a 2- to 3-fold increase in the level of oxidative base damage. Eugenol quinone methide was prepared by Ag(I)oxide oxidation of eugenol. Peroxidase activation of eugenol gave a product that had the same UV spectrum as eugenol quinone methide, which suggests that it was one of the products. Reaction of eugenol quinone methide with either DNA or deoxyguanosine-3'-phosphate produced two principal adducts (2 and 4). When DNA adduct 2 formed by incubation of eugenol quinone methide with deoxyguanosine-3'-phosphate was compared with DNA 2 adduct formed in HL-60 cells treated with eugenol results demonstrated that they were the same. This suggests that eugenol quinone methide is one of the reactive intermediates leading to DNA adduct formation in cells. Activation of eugenol with 10 microM copper sulfate resulted in the production of one principal (2) and several minor adducts. DNA adduct 2 formed by activation of eugenol with copper sulfate was the same as DNA adduct 2 formed by either peroxidase activation of eugenol or by reactions with eugenol quinone methide, which indicates that the reactive intermediates generated by these activation systems were similar. Copper sulfate produced a 95-fold increase in the level of oxidative base damage, which was significantly inhibited by the addition of either bathocuproinedisulphonic acid or catalase. The formation of oxidative base damage was consistent with a Fenton reaction mechanism. Our results demonstrate that eugenol can be activated to form both DNA adducts and oxidative base damage. We propose that the formation of this DNA damage may contribute to the observed toxic properties of eugenol.

8-Hydroxy-2'-Deoxyguanosine↗
PubMed1998

Propofol and other intravenous anesthetics have sites of action on the gamma-aminobutyric acid type A receptor distinct from that for isoflurane.

Both volatile and intravenous general anesthetics allosterically enhance gamma-aminobutyric acid (GABA)-evoked chloride currents at the GABA type A (GABAA) receptor. Recent work has revealed that two specific amino acid residues within transmembrane domain (TM)2 and TM3 are necessary for positive modulation of GABAA and glycine receptors by the volatile anesthetic enflurane. We now report that mutation of these residues within either GABAA alpha2 (S270 or A291) or beta1 (S265 or M286) subunits resulted in receptors that retain normal or near-normal gating by GABA but are insensitive to clinically relevant concentrations of another inhaled anesthetic, isoflurane. To determine whether receptor modulation by intravenous general anesthetics also was affected by these point mutations, we examined the effects of propofol, etomidate, the barbiturate methohexital, and the steroid alphaxalone on wild-type and mutant GABAA receptors expressed in human embryonic kidney 293 cells. In most cases, these mutations had little or no effect on the actions of these intravenous anesthetics. However, a point mutation in the beta1 subunit (M286W) abolished potentiation of GABA by propofol but did not alter direct activation of the receptor by high concentrations of propofol. These data indicate that the receptor structural requirements for positive modulation by volatile and intravenous general anesthetics may be quite distinct.

Anesthetics, Inhalation↗
PubMed1998

[Effects of exposure to noise in oil-drilling well sites on cochlea in guinea pigs].

OBJECTIVE: To prevent from hearing damage caused by noise in the sites with drilling wells. METHODS: Intensity and spectrum of the noise produced in the sites with drilling wells were measured and analyzed. Measures, such as sound insulation, vibration damping and changing layouts of the drilling platform, were adopted, and electrocochleography (ECochG) and scanning electron microscopy were used to observe the effects of noise in the sites where drilling wells located on the hearing organs in guinea pigs after exposure. RESULTS: There was no significant difference in APN1 latency of ECochG before and after exposure to noise. There were significant difference in hearing threshold values between the groups in a room with sound isolation, in moving status and beside the diesel engine, and no significant difference in those between the groups placed one meter horizontally and vertically away from the diesel engine on the drilling platform. Damage to hairy cell under scanning electron microscope was more severe in the guinea pigs placed beside the diesel engine and at the drilling well platform, with inclination and exfoliation of static pili of outer hairy cells, especially in the second and third lines, and part exfoliation of inner hairy cell. Hairy cells of the guinea pigs in a room with sound insulation and in moving status were basically normal. Inclination of hairy cells only could be seen in the third line. CONCLUSION: Animal experiments indicated that the measures mentioned above are effective and should be applied widely for preventing from hearing damage.

Animals↗
PubMed1998

[The general survey for chondromalacia of 2,743 Chinese population].

OBJECTIVE: To evaluate the distribution of chondromalacia patella in Chinese population. METHODS: A random cluster sampling survey was performed covering 2,743 subjects varied in age, sex and occupation in 1995. RESULTS: The prevalence rate is 36.2%. The occurrence in women was higher than that in men (P < 0.01), while in the age group, 30 to 39 years was the highest being 55.8%. The prevalence rate in soldier being 47.5% was the highest among varied occupations. CONCLUSIONS: Our study is the first survey to be performed in a large number of Chinese population. This investigation may reflect the prevalence rate of chondromalacia patella in China.

Adolescent↗
PubMed1998

[The effects of different instrumentation on scoliosis].

OBJECTIVE: To investigate the effectiveness of the treatment of scoliosis with different surgical methods. METHOD: 125 cases of scoliosis less than 100 degrees treated during 14 years (1984 - 1997) were analyzed. Different operative methods were compared for their advantages and disadvantages, i.e. results and complications 10 months after operation and follow-up. RESULT: Harrington instrumentation was less effective in scoliotic curve correction and had more complications than other procedures. Luque technique needed longer operation time for wire passage at each level and had risk of neurological damage. CD and Zielke method showed 10% - 20% curve correction. No dislodgment of hook and fractured rods was found in the CD group. Anterior release plus posterior instrumentation was superior to other techniques. CONCLUSION: Treatment of scoliosis with anterior release plus posterior CD is practical for children before a new technique without spinal fusion is developed.

Adolescent↗
PubMed1998

Trichloroethanol modulation of recombinant GABAA, glycine and GABA rho 1 receptors.

The actions of 2,2,2,-trichloroethanol were studied on agonist-activated Cl- currents in gamma-aminobutyric acid type A (GABAA), glycine and GABA rho 1 receptors by use of the whole-cell patch-clamp technique. Recombinant wild-type and mutant receptor subunits were transiently expressed in human embryonic kidney (HEK) 293 cells. Trichloroethanol enhanced currents elicited by submaximal (EC20) agonist concentrations at GABAA alpha 2 beta 1 receptors and glycine alpha 1 homomeric receptors in a reversible, concentration-dependent manner. Trichloroethanol, at concentrations of < or = 2 mM, did not significantly alter the magnitude of submaximal GABA currents at GABA rho 1 receptors, whereas higher concentrations inhibited submaximal GABA currents. Recent work has identified residues within putative transmembrane domains 2 and 3 as critical for positive modulation of GABAA and glycine receptors by n-alkanols and volatile ether anesthetics. Submaximal glycine currents at receptors containing either of two specific mutations within the glycine receptor alpha 1 subunit (S267I and A288W) were not enhanced by low concentrations of trichloroethanol and were inhibited by higher concentrations of trichloroethanol. In the GABAA alpha 2 beta 1 receptor, a specific mutation within transmembrane domain 3 of the beta 1 subunit (M286W) also abolished positive modulation by trichloroethanol. Mutations within the GABAA alpha 2 receptor subunit did not alter positive modulation by TCEt, whereas such mutations ablate positive modulation by n-alkanols and volatile anesthetics. In summary, trichloroethanol modulation of GABAA, glycine and GABA rho 1 receptors shares some, but not all, features in common with the requirements for modulation by n-alkanols and volatile anesthetics.

Amino Acid Sequence↗
PubMed1998

A subcutaneous tissue reaction in the early stage to a synthetic auditory ossicle (Bioceram) in rats.

The discs of synthetic auditory ossicle (Bioceram), which are composed of aluminium oxide (Al2O3), were implanted subcutaneously in the interscapular region of 16 rats. The implanted specimens were removed at 1, 3, 7 and 14 days after implantation. The decalcified 6 microns thick sections were stained with H.E. and cell types around the implants were counted microscopically. We found that an acute inflammatory reaction occurred at one day, in which macrophages and neutrophiles predominated, and almost disappeared at about 7 days after implantation. Fibrosis began to be observed at 3 days. During this early stage, foreign body giant cells were found in only one specimen at 3 days. These findings, in comparison with those in the controls, showed that the chemical irritation of Bioceram to the subcutaneous tissue is slight, although the physical and/or chemical irritation of Bioceram lasts continuously and induces fibrosis around the bioimplant. The results so far suggest that Bioceram seems to be a satisfactorily biocompatible material, at least within the extent of 2 weeks.

Aluminum Oxide↗
PubMed1997

Expression of calmodulin and calmodulin binding proteins in rat fibroblasts stably transfected with protein kinase C and oncogenes.

Molecular mechanisms leading to elevated calmodulin (CaM) expression in cancer have not yet been discovered. We have quantitated the levels of transcripts derived from all three CaM genes in a variety of the same origin rat fibroblasts transformed with oncogenes in combination with gene for protein kinase C using Northern blot analysis with three CaM gene specific cDNA probes. Five species of CaM mRNA were detected in all these cells. Surprisingly many of the investigated cell lines exhibited a decreased content of all CaM mRNAs as compared to control cells with CaMI and CaMII transcripts showing the most pronounced alterations. In contrast, CaM protein levels were increased in all these cell lines as determined by a radioimmunoassay. These results suggest that oncogenic up-regulation of CaM synthesis takes place posttranscriptionally. Several CaM binding proteins were found at different concentrations in the studied cell lines depending on the oncogenes used for transformation. However, CaM overexpression does not seem to affect the overall levels of CaM binding proteins.

Animals↗
PubMed1997

Sites of alcohol and volatile anaesthetic action on GABA(A) and glycine receptors.

Volatile anaesthetics have historically been considered to act in a nonspecific manner on the central nervous system. More recent studies, however, have revealed that the receptors for inhibitory neurotransmitters such as gamma-aminobutyric acid (GABA) and glycine are sensitive to clinically relevant concentrations of inhaled anaesthetics. The function of GABA(A) and glycine receptors is enhanced by a number of anaesthetics and alcohols, whereas activity of the related GABA rho1 receptor is reduced. We have used this difference in pharmacology to investigate the molecular basis for modulation of these receptors by anaesthetics and alcohols. By using chimaeric receptor constructs, we have identified a region of 45 amino-acid residues that is both necessary and sufficient for the enhancement of receptor function. Within this region, two specific amino-acid residues in transmembrane domains 2 and 3 are critical for allosteric modulation of both GABA(A) and glycine receptors by alcohols and two volatile anaesthetics. These observations support the idea that anaesthetics exert a specific effect on these ion-channel proteins, and allow for the future testing of specific hypotheses of the action of anaesthetics.

Alanine↗
PubMed1997

Structure and expression of the chicken calmodulin I gene.

The chicken calmodulin I (CaMI) gene has been isolated and characterized on the level of cDNA and genomic DNA. The deduced amino acid (aa) sequence is identical to the one of chicken CaMII which consists of 148 aa. The CaMI gene contains six exons. Its intron/exon organization is identical to that of the chicken CaMII and the CaMI and CaMIII genes of rat and human. Expression of the CaMI gene was detected in all chicken tissues examined, although at varying levels. The gene is transcribed into four mRNAs of 0.8, 1.4, 1.7 and 4.4 kb as determined by Northern blot analysis. Our results demonstrate that the "multigene-one-protein" principle of CaM synthesis is not only applicable to mammals whose CaM is encoded by three different genes, but also to chickens.

Amino Acid Sequence↗
PubMed1997

Effect in supralethally irradiated rats of granulocyte colony-stimulating factor and lisofylline on hematopoietic reconstitution by syngeneic bone marrow or whole organ passenger leukocytes.

We have previously shown the existence of migratory hematopoietic stem cells in adult solid organs. This study demonstrates that granulocyte colony-stimulating factor (G-CSF) and lisofylline, a phosphatidic acid inhibitor that suppresses hematopoiesis-inhibiting cytokines, can enhance the engraftment of organ-based hematopoietic stem cells. When syngeneic heart grafts or liver nonparenchymal cells were transplanted into lethally irradiated (9.5 Gy) Lewis rats, complete hematopoietic reconstitution and animal survival were significantly improved by treating the recipient with G-CSF or, to a lesser extent, with lisofylline. Pretreatment of hepatic nonparenchymal cell donors with G-CSF, but not lisofylline, also resulted in striking improvement of recipient survival which was associated with an augmented subpopulation of donor stem cells. The results suggest that these drugs can be used to enhance the chimerism that we postulate to be the basis of organ allograft acceptance.

Adjuvants, Immunologic↗