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Biomedical subjects

Y Hong

Publications and source records attributed to Y Hong.

At least 19 recordsLinked to original sources
PubMed1997

Actin depolymerizing factor (ADF/cofilin) enhances the rate of filament turnover: implication in actin-based motility.

Actin-binding proteins of the actin depolymerizing factor (ADF)/cofilin family are thought to control actin-based motile processes. ADF1 from Arabidopsis thaliana appears to be a good model that is functionally similar to other members of the family. The function of ADF in actin dynamics has been examined using a combination of physical-chemical methods and actin-based motility assays, under physiological ionic conditions and at pH 7.8. ADF binds the ADP-bound forms of G- or F-actin with an affinity two orders of magnitude higher than the ATP- or ADP-Pi-bound forms. A major property of ADF is its ability to enhance the in vitro turnover rate (treadmilling) of actin filaments to a value comparable to that observed in vivo in motile lamellipodia. ADF increases the rate of propulsion of Listeria monocytogenes in highly diluted, ADF-limited platelet extracts and shortens the actin tails. These effects are mediated by the participation of ADF in actin filament assembly, which results in a change in the kinetic parameters at the two ends of the actin filament. The kinetic effects of ADF are end specific and cannot be accounted for by filament severing. The main functionally relevant effect is a 25-fold increase in the rate of actin dissociation from the pointed ends, while the rate of dissociation from the barbed ends is unchanged. This large increase in the rate-limiting step of the monomer-polymer cycle at steady state is responsible for the increase in the rate of actin-based motile processes. In conclusion, the function of ADF is not to sequester G-actin. ADF uses ATP hydrolysis in actin assembly to enhance filament dynamics.

Actin Cytoskeleton↗
PubMed1997

The bZip dimerization domain of the Epstein-Barr virus BZLF1 (Z) protein mediates lymphoid-specific negative regulation.

The Epstein-Barr virus (EBV) immediate-early (IE) protein, BZLF1 (Z), initiates the switch from latent to lytic infection, Z transactivation of an early viral promoter, BMRF1, is relatively inefficient in lymphoid cells (compared with epithelial cells), unless the other EBV IE protein, BRLF1, is also present. Cellular proteins, including the p65 component of NF-kappa B, have been shown to interact directly with Z in vitro through the bZip dimerization domain and inhibit Z-induced transactivation. Here we precisely define a residue within the bZip dimerization domain of Z (amino acid 200) which is required for interaction in vitro with the p65 component of NF-kappa B, but is not essential for Z homodimerization. In lymphoid cells, a Z mutant which has been altered at amino acid 200 (tyrosine to glutamic acid) transactivates both the early BMRF1 promoter and the immediate-early BZLF1 promoter (Zp) four- to fivefold better than wild-type Z. In contrast, mutation of amino acid 200 does not affect Z transactivator function in epithelial cells. The results suggest that Z function is specifically inhibited by a lymphoid-specific protein(s) through amino acid 200 in the bZip dimerization domain. Modulation of Z's activator function may help to regulate the stringency of viral latency in lymphocytes.

Cell Line↗
PubMed1997

Transactivation of dianthin transgene expression by African cassava mosaic virus AC2.

We have recently described a novel strategy for engineering resistance to African cassava mosaic virus (ACMV) in transgenic Nicotiana benthamiana plants using a virus-inducible promoter to control the expression of a plant ribosome-inactivating protein (RIP) transgene (Y. Hong et al., Virology 220, 119-127, 1996). Here, we have used a potato virus X (PVX) vector to express the ACMV transactivator protein, AC2, in planta. We confirm that amplification of RIP activity in transgenic plants is mediated by AC2; disruption of AC2 expression by either the introduction of an in-frame stop codon or the deletion of 5'-terminal or 3'-terminal coding sequences reduced RIP expression to the basal level associated with PVX-infected plants. AC2 expression from the PVX vector induced necrosis in nontransformed plants as well as in plants containing the RIP transgene, suggesting that the protein can functionally interact with PVX and/or host factors. The potential of this system to provide a direct and sensitive assay to investigate AC2 function in planta is discussed.

Antiviral Agents↗
PubMed1997

Acupressure treatment for prevention of postoperative nausea and vomiting.

Postoperative nausea and vomiting are still common problems after general anesthesia, especially in ambulatory surgery. Drug therapy is often complicated with central nervous system symptoms. We studied a nonpharmacological method of therapy--acupressure--at the Pericardium 6 (P.6) (Nei-Guan) meridian point. Two hundred consecutive healthy patients undergoing a variety of short surgical procedures were included in a randomized, double-blind study: 108 patients were in the acupressure group (Group 1) and 92 patients were in the control group (Group 2). Spherical beads of acupressure bands were placed at the P.6 points in the anterior surface of both forearms in Group 1 patients, while in Group 2 they were placed inappropriately on the posterior surface. The acupressure bands were placed before induction of anesthesia and were removed 6 h postoperatively. They were covered with a soft cotton wrapping to conceal them from the blinded observer who evaluated the patients for presence of nausea and vomiting and checked the order sheet for any antiemetics prescribed. In both groups, the age, gender, height, weight, and type and duration of surgical procedures were all comparable without significant statistical difference. In Group 1, only 25 of 108 patients (23%) had nausea and vomiting as compared to Group 2, in which 38 of 92 patients (41%) had nausea and vomiting (P = 0.0058). We concluded that acupressure at the P.6 (Nei-Guan) point is an effective prophylaxis for postsurgical nausea and vomiting and therefore a good alternative to conventional antiemetic treatment.

Acupressure↗
PubMed1997

Megakaryocyte ploidy and platelet changes in human diabetes and atherosclerosis.

Altered platelet morphology and function have been reported in patients with diabetes. They are likely to be associated with the pathological processes and increased risk of vascular disease seen in these patients. Mean platelet volume (MPV), platelet count, and megakaryocyte (MK) ploidy (DNA content) were measured in (1) nondiabetics with normal coronary arteries, (2) nondiabetics with coronary artery atherosclerosis, (3) diabetics without evidence of vascular complications, and (4) diabetics with vascular disease. The platelet count (+/- SD) was increased in all groups but only significantly in the diabetics with vascular disease (236 +/- 65 versus 250 +/- 54 versus 257 +/- 64 versus 295 +/- 90 [P < or = .05] x 10(9)/L, for groups, I, II, II, and IV, respectively). The MPV was significantly increased in patients with atherosclerosis (7.0 +/- 0.4 versus 8.0 +/- 1.2 [P < or = .05] versus 7.2 +/- 0.9 versus 8.1 +/- 0.9 [P < or = .05] IL). Geometric mean MK ploidy was significantly increased in all groups compared with controls (16 +/- 1.5 versus 18.7 +/- 1.8 [P < or = .05] versus 19.8 +/- 1.6 [P < or = .05] versus 20.1 +/- 2.7 [P < or = .05]). Furthermore, some patients with vascular disease and/or diabetes had a modal ploidy shift from 16 (the normal mammalian modal ploidy) to 32, with a concomitant reduction of MKs in the 8 and 16 ploidy classes. This shift was seen particularly in the diabetics with vascular disease (P = .007). Interleukin-6 (IL-6) levels were measured and were elevated in patients with atherosclerosis; the highest levels were found in the diabetic patients (0.7 +/- 0.9 versus 5.3 +/- 5.5 [P < or = .05] versus 2.5 +/- 2.8 versus 6.7 +/- 5.5 [P < or = .05] ng/L). In the diabetic patients with atherosclerosis, fibrinogen levels were also increased (2.85 +/- 0.76 versus 3.34 +/- 1.32 versus 2.43 +/- 1.50 versus 5.59 +/- 1.72 [P < or = .05] g/L). Furthermore, IL-6 levels correlated with MK ploidy (r = .45, P = .009) and fibrinogen levels (r = .5, P = .0001). This study demonstrates that patients with vascular disease, particularly diabetics, have an altered MK ploidy distribution, showing a shift toward higher ploidy in association with an increased platelet mass (count x volume). Changes in platelets in diabetes probably reflect MK changes, which themselves are a response to systemic change.

Aged↗
PubMed1997

Genetic and environmental architecture of the features of the insulin-resistance syndrome.

The contribution of genetic and environmental effects to the clustering of metabolic factors contained in insulin-resistance syndrome (IRS) is still unclear. To explore the genetic architecture of IRS, we examined a population of elderly twins from the Swedish Adoption/Twin Study of Aging. A sample of 289 pairs of twins (mean age 65.5 years; range 52-86 years), of whom 140 pairs had been reared apart, was studied. The features contained in the IRS consisted of body-mass index (BMI), insulin resistance, triglycerides, HDL cholesterol, and systolic blood pressure. Intraclass correlations, cross-twin correlations, and model-fitting analyses were used to evaluate the relative importance of genetic and environmental influences for variation in and covariation among the components of the syndrome. All of the five principal metabolic components contained in IRS are more or less influenced by a single latent genetic factor, whereas only three of the components (triglycerides, insulin resistance, and HDL cholesterol) are influenced by a latent individual-specific environmental factor. The genetic factor reflected influences of importance to BMI and insulin resistance and to a lesser degree to triglycerides, HDL cholesterol, and systolic blood pressure, whereas the individual-specific environmental factor reflected influences in common to triglycerides and HDL cholesterol and to a lesser degree to insulin resistance. Systolic blood pressure was related to IRS, albeit weakly, only through genetic effects. In conclusion, IRS appears to be influenced by different sets of genetic and environmental mechanisms. The set of genetic influences in common to all the components may initiate the abnormalities underlying IRS.

Aged↗
PubMed1996

RNA polymerase activity catalyzed by a potyvirus-encoded RNA-dependent RNA polymerase.

We have expressed the putative RNA-dependent RNA polymerase encoded by the potyvirus tobacco vein mottling virus (TVMV) in Escherichia coli as a glutathione S-transferase fusion protein. As prepared, the fusion protein possessed the poly(U) polymerase activity that is a hallmark of other picornavirus-encoded polymerases. In addition, this protein was able to utilize full-length TVMV RNA as a template for RNA synthesis. A fusion protein containing a mutation in the highly conserved GDD motif of the polymerase (GDD-->ADD) possessed 7% of the activity of the wild type. Our results confirm that the presumed polymerase encoded by TVMV is in fact an RNA-dependent RNA polymerase and that the GDD motif so widely seen in viral polymerases has an important function in the TVMV protein.

DNA-Directed RNA Polymerases↗
PubMed1996

A glass fiber/diethylaminoethyl double filter binding assay that measures apoptotic internucleosomal DNA fragmentation.

A filter binding assay that measures internucleosomal DNA fragmentation associated with apoptosis is described. The assay is based on a novel principle that consists of using simultaneously two kinds of glass fiber filters to harvest [3H]thymidine-prelabeled cells following their incubation with inducers of apoptosis. One filter, which is neutral, traps intact chromatin and high-molecular-weight DNA. The other filter, which is positively charged with DEAE active groups, traps low-molecular-weight DNA fragments. DNA fragmentation is quantified by measuring the radioactivity retained by each of the filters. The assay was evaluated with the histiocytic lymphoma cell line U937 and the topoisomerase inhibitors camptothecin, etoposide, and doxorubicin. These agents caused a dose-dependent decrease of radioactivity in the neutral filter and a parallel increase of radioactivity in the DEAE filter. Irradiation-induced single strand breaks and topoisomerase-mediated primary DNA damage were not detected by this method. Consistent with the detection of internucleosomal DNA fragmentation, the effects measured by this assay were prevented by the endonuclease inhibitor zinc acetate and by the metabolic inhibitor sodium azide. Results obtained using this assay were validated by observation of DNA ladders on agarose gels and by morphologic examination of apoptotic features. Evaluation of the assay in a mock screen demonstrated that the introduction of the DEAE filter increases the assay sensitivity and eliminates false positives. Thus, this assay may be used in high-throughput screening approaches to discover novel modulators of apoptosis.

Antineoplastic Agents↗
PubMed1996

Resistance to geminivirus infection by virus-induced expression of dianthin in transgenic plants.

Ribosome-inactivating proteins (RIPs) are naturally occurring plant toxins that exhibit antiviral activity against a diverse range of plant and animal viruses. Here, the action of dianthin, a potent RIP isolated from Dianthus caryophyllus, has been exploited to engineer resistance to a plant DNA virus, African cassava mosaic virus (ACMV), in transgenic Nicotiana benthamiana. To achieve this, dianthin has been expressed from the ACMV virion-sense promoter that is transactivated by the product of viral gene AC2. This avoids the need for constitutive expression of the RIP, facilitating the regeneration of phenotypically normal plants, and ensures transgene expression is localized to virus-infected cells. When challenged with ACMV, transgenic plants produce atypical necrotic lesions on inoculated leaves, indicative of dianthin expression, viral DNA accumulation is significantly reduced in these tissues, and plants exhibit attenuated systemic symptoms from which they recover. This phenotype holds for isolates of ACMV but not for other geminiviruses, suggesting that AC2 homologues from the latter are unable to efficiently transactivate the ACMV promoter.

Antiviral Agents↗
PubMed1996

Contribution of peripheral alpha 1A-adrenoceptors to pain induced by formalin or by alpha-methyl-5-hydroxytryptamine plus noradrenaline.

We examined the peripheral adrenergic mechanisms involved in pain induced by alpha-methyl-5-hydroxytryptamine (alpha-methyl-5-HT) plus (+/-)-noradrenaline or prostaglandin E2 and by intraplantar formalin. Agents were injected s.c. into the plantar surface of rats' paws, and the paw lifting and licking response scored. Pain produced by alpha-methyl-5-HT (10 micrograms) plus noradrenaline (10 micrograms) was blocked by pretreatment with the alpha-adrenoceptor antagonists, phentolamine (10 micrograms) and prazocin HCl (alpha 1; 40 micrograms), but not by timolol (beta; 10 micrograms) or idazoxan (alpha 2; 40 micrograms). Phenylepherine, but not clonidine, substituted for noradrenaline to induce pain when combined with alpha-methyl-5-HT. The alpha 1A-adrenoceptor antagonist, WB-4101 (2-(2,6-dimethoxyphenoxyethyl) aminomethyl-1,4-benzodioxane HCl), but not the alpha 1B- adrenoceptor antagonist, chloroethylclonidine, also blocked the pain response produced by alpha-methyl-5-HT plus noradrenaline. Neither of these agents altered pain produced by alpha-methyl-5-HT plus prostaglandin E2 (0.1 microgram). Formalin-induced pain (1%, 50 microliters) was biphasic, and timolol increased the first phase response. The second phase was attenuated by 40% by phentolamine (10 micrograms) injected 10 min before formalin or at the beginning of the second phase; 30 micrograms did not produce a larger effect. Prazosin and WP-4101, but not idazoxan or chloroethylclonidine, also attenuated the second phase. Thus, activation of alpha 1A-adrenoceptors can contribute to pain, but pain induced by alpha-methyl-5-HT plus prostaglandin E2 is independent of adrenergic function, indicating that adrenergic function is not necessary for induction of pain by inflammatory mediators. alpha 1A-Adrenoceptor blockade attenuates pain when administered after development of pain, implying that peripheral adrenergic mechanisms contribute to ongoing maintenance of pain.

Adrenergic alpha-1 Receptor Agonists↗
PubMed1996

Inhibition of protein kinase C suppresses megakaryocytic differentiation and stimulates erythroid differentiation in HEL cells.

The bisindolylmaleimide, GF109203X (2-[1-(3-dimethylaminopropyl)-1H-indol-3-yl]-3-(1H-indol-3-yl)-maleimide ), a highly selective inhibitor of protein kinase C (PKC), was used to test the role of this enzyme in phorbol ester-induced megakaryocytic differentiation of HEL cells. Treatment of these cells with 10 nmol/L phorbol 12-myristate 13-acetate (PMA) for 3 days caused a complete inhibition of proliferation and a threefold increase in the surface expression of glycoprotein (GP) IIIa, a marker of megakaryocytic differentiation that forms part of the fibrinogen receptor complex, GPIIb/IIIa. A similar effect was observed with phorbol 12,13-dibutyrate, but not with the biologically inactive derivative PMA-4-O-methyl ether. The PMA-induced increase in GPIIIa expression was completely inhibited by GF109203X in a dose-dependent manner (IC50 = 0.5 mumol/L), with a maximal effect at 2.5 to 5.0 mumol/L. GF109203X also blocked the inhibitory effect of PMA on cell growth and inhibited PMA-stimulated phosphorylation of the 47-kD PKC substrate, pleckstrin. Incubation of HEL cells with 25 mumol/L hemin for 3 days caused a fourfold to fivefold increase in expression of the erythroid differentiation marker, glycophorin A. In contrast to the inhibitory effect of GF109203X on GPIIIa expression, hemin induction of glycophorin A was enhanced by this compound. Furthermore, GF109203X alone caused a dose-dependent increase in glycophorin A expression, and induced hemoglobinization. Consistent with these changes, Northern blot analysis revealed that GF109203X treatment reduced the steady-state level of GPIIb mRNA and increased those for glycophorin A and gamma-globin. These results suggest that PKC may act as a developmental switch controlling erythroid/megakaryocytic differentiation.

Aphidicolin↗
PubMed1996

Crosslinked poly(1-vinyl-2-pyrrolidinone) as a vitreous substitute.

A hydrogel with a high water content was produced and tested as a possible vitreous substitute. The polymer (poly[1-vinyl-2-pyrrolidinone]) (PVP) was synthesized by free radical bulk polymerization of 1-vinyl-2-pyrrolidinone (VP) in the presence of 0.25% divinyl glycol (DVG) as a crosslinking agent. The fully hydrated polymer, containing about 98% water, was clear, transparent, autoclavable, and easily injected through a small-gauge needle with minimum fragmentation, and without changes in its optical properties. Dynamic mechanical analysis of the hydrogel indicated a covalently crosslinked elastic network both before and after injection. The resilience of hydrogel decreased after being subjected to shear stress during the injection process. A cytotoxicity bioassay of the hydrogel in vitro, using cultured mouse (Balb/ c-3T3) fibroblasts, showed cytostatic but not cytocidal effects. The hydrogel was injected into the vitreous cavity of rabbits and followed up to 4 weeks. The gel was clinically well tolerated, however opacities in the vitreous body were observed following the insertion of the gel. Histopathological examination revealed no adverse reactions to the retina, but the presence of loose polymer particles indicated the possibility of the biodegradation of the polymer. These results suggest the potential use of crosslinked PVP hydrogels as vitreous substitutes, provided that their biodegradation is not significant, a matter that should be further investigated.

3T3 Cells↗
PubMed1996

Expression of three members of the calcium-dependent protein kinase gene family in Arabidopsis thaliana.

Calcium-dependent protein kinases (CDPKs) belong to a unique family of enzymes containing a single polypeptide chain with a kinase domain at the amino terminus and a putative calcium-binding EF hands structure at the carboxyl terminus. From Arabidopsis thaliana, we have cloned three distinct cDNA sequences encoding CDPKs, which were designated as atcdpk6, atcdpk9 and atcdpk19. The full-length cDNA sequences for atcdpk6, atcdpk9 and atcdpk19 encode proteins with a molecular weight of 59343, 55376 and 59947, respectively. Recombinant atCDPK6 and atCDPK9 proteins were fully active as kinases whose activities were induced by Ca2+. Biochemical studies suggested the presence of an autoinhibitory domain in the junction between the kinase domain and the EF hands structure. Serial deletion of the four EF hands of atCDPK6 demonstrated that the integrity of the four EF hands was crucial to the Ca2+ response. All the three atcdpk genes were ubiquitiously expressed in the plant as demonstrated by RNA gel blot experiments. Comparison of the genomic sequences suggested that the three cdpk genes have evolved differently. Using antibodies against atCDPK6 and atCDPK9 for immunohistochemical experiments, CDPKs were found to be expressed in specific cell types in a temporally and developmentally regulated manner.

Amino Acid Sequence↗
PubMed1996

Activation of 5-HT2A receptors potentiates pain produced by inflammatory mediators.

Previous results from our laboratory indicate that serotonin (5-HT) potentiates pain produced by other inflammatory mediators. To characterize the receptor subtype(s) mediating this synergistic effect of 5-HT, selective 5-HT agonists were injected, alone or with noradrenaline (NA) or prostaglandin E2 (PGE2), into the plantar surface of the paws of rats. The behavioural response (favouring, elevation and licking the paw) was recorded using the rating scale developed to quantify formalin-induced pain. The 5-HT1A and 5-HT3 agonists, 8-OH-DPAT and 2-methyl-5-HT, respectively, produced only transient responses by themselves and did not interact with PGE2 or NA. The 5-HT2 agonists, alpha-methyl-5-HT and DOI, also produced transient responses alone, but induced lifting and licking of the injected paw lasting more than 30 min when combined with PGE2 or NA. The lifting and licking response produced by 5-HT plus PGE2 was not altered by intraplantar pretreatment with the 5-HT1A and 5-HT3 antagonists, BMY 7378 and tropisetron, but was attenuated by the 5-HT2A/2C antagonist ketanserin. The pain response produced by alpha-methyl-5-HT plus PGE2 was blocked by pretreatment with the 5-HT2A/2C antagonists ketanserin and ritanserin, and the 5-HT2A antagonist spiperone (MPE50 values 1.4, 7.7 and 0.06 nmol, respectively). The second phase of the response to intraplantar formalin was also attenuated by ketanserin, ritanserin and spiperone (MPE50 values 11.3, 21.8 and 0.23 nmol, respectively). These data imply that 5-HT2A antagonists may be effective peripherally acting analgesics or analgesic adjuncts in pain associated with 5-HT release from platelets, such acute injury and, perhaps, some chronic pain states.

8-Hydroxy-2-(di-n-propylamino)tetralin↗
PubMed1996

Pluripotency and differentiation of embryonic stem cell lines from the medakafish (Oryzias latipes).

Small aquarium fish, like the medaka and zebrafish, offer an excellent opportunity to combine embryological, genetic and molecular analyses of vertebrate development. Pluripotent embryonic stem (ES) cells have enormous potential to study the totipotency and differentiation of cells and provide s bridge linking in vitro manipulations of the genome. In this report we describe the establishment, pluripotency and differentiation of medaka ES-like cell lines (MES). The MES cells exhibit stable growth over 18 months of culture with 100 passages using defined culture conditions in the absence of feeder layer cells. They have a normal karyotype and form colonies of densely packed, alkaline phosphatase-positive cells resembling undifferentiated mouse ES cells. In suspension culture they form embryoid bodies, and under appropriate conditions, differentiate into a variety of cell types.

Alkaline Phosphatase↗
PubMed1996

Identification of plant cytoskeletal, cell cycle-related and polarity-related proteins using Schizosaccharomyces pombe.

The fission yeast Schizosaccharomyces pombe has been used to identify Arabidopsis thaliana proteins that may play a role in cell shape maintenance or cell cycle regulation. An Arabidopsis thaliana cDNA library was constructed in pREP5N vector under the control of the inducible nmt1 promoter and transformed into S. pombe. Expression of the A. thaliana sequences was induced and clones showing severe morphological changes were identified and analysed. Comparison of the sequences of the inserts with the sequence data bases revealed that several cDNAs encode proteins known to play a role in function of the cytoskeleton, the cell cycle and establishment of cell polarity. These include alpha-1, alpha-2, alpha-6 and beta-6 tubulins, myosin heavy chain-like protein, ubiquitin conjugating enzymes UBC9 (E2), 26S protease subunits, Ranbinding protein, myb protein, PRL1 gene product and rho protein. Approximately 30% of the clones encode novel sequences. The results suggest that S. pombe phenotypic screening can be used to identify plant proteins involved in cell shape maintenance and regulation during cell cycle and development.

Actin Depolymerizing Factors↗