[Efficacy of balloon dilatation by fiberoptic bronchoscopy in treating benign bronchial stenosis].
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Adolescent↗
Biomedical subjects
Hong Luo
Publications and source records attributed to Hong Luo.
Induction of Duffy gene (FY) in human endothelial cells and in mouse.
Duffy Blood Group protein is a glycoprotein with seven transmembrane domains that binds to C-X-C and C-C chemokines. The antigen is constitutively expressed in endothelial and epithelial cells of several nonerythroid tissues and in Purkinje cells of the cerebellum. We studied the effect of proinflammatory cytokines on Duffy gene expression in endothelial cells from human umbilical vein (HUVEC) and human pulmonary arteries (HPAEC). Also, we studied the effect of inflammatory agents like bacterial lipopolysaccharide (LPS) on Duffy gene induction in mouse. Reverse transcription-PCR and mRNA blot analyses showed that Duffy mRNA was present in these cells in negligible amounts. However, treatment with tumor necrosis factor-alpha for 6-24h resulted in a 5 to 8-fold increase in Duffy mRNA. On the other hand, treatment with interleukin-1 (IL-1), IL-6 or LPS did not have any effect. Fluorescence microscopy and fluorescence activated cell sorting showed greater expression of Duffy protein in treated cells correlating the increase in mRNA synthesis with an increase in antigen production. In mice, Duffy gene was induced in lungs and brain with LPS treatment indicating that the induction is a physiological event. Vascular endothelial cells may induce Duffy protein to regulate leukocytes and/or chemokine trafficking.
Animals↗
Transient reporter gene (GUS) expression in creeping bentgrass (Agrostis palustris) is affected by in vivo nucleolytic activity.
Leaf and callus tissues of a creeping bentgrass cultivar (Penn A4) had high nuclease activities that degraded exogenously added plasmid DNA. When callus tissue was incubated for 24 h with heparin, spermidine, aurintricarboxylic acid or polyethylene glycol, only heparin and spermidine were effective as in vitro nuclease inhibitors, protecting exogenously added plasmid DNA from degradation. When beta-glucuronidase (GUS) reporter gene activity was evaluated in heparin-treated (0.6%), 14-month old callus following microprojectile bombardment, GUS activity increased 1000-fold compared to equivalent aged untreated Penn A4 callus. Similar enhancement from heparin pretreatment (0.6% or 1.2%) was not observed in 6-month old callus. This is likely due to much higher activities of nuclease in the younger callus.
Aging↗
Dynamic distribution of TTK in HeLa cells: insights from an ultrastructural study.
Entry into mitosis is driven by signaling cascades of mitotic kinases. Our recent studies show that TTK, a kinetochore-associated protein kinase, interacts with CENP-E, a mitotic kinesin located to corona fiber of kinetochore. Using immunoelectron microscopy, here we show that TTK is present at the nuclear pore adjacent complex of interphase HeLa cells. Upon nuclear envelope fragmentation, TTK targets to the outermost region of the developing kinetochores of monoorient chromosome as well as to spindle poles. After stable attachment, throughout chromosome congression, TTK is a constituent of the corona fibers, extending up to 90 nm away from the kinetochore outer plate. Upon metaphase alignment, TTK departs from the kinetochore and migrates toward the centrosomes. Taken together, this evidence strongly supports a model in which TTK functions in spindle checkpoint signaling cascades at both kinetochore and centrosome.
Cell Cycle Proteins↗
Promoter analysis in transient assays using a GUS reporter gene construct in creeping bentgrass (Agrostis palustris).
Transient expression profiles for several chimeric beta-glucuronidase (GUS) gene constructs were determined in tissues (young leaves, mature leaves and roots) of creeping bentgrass (Agrostis palustris, cv. Penn A4) following microprojectile bombardment. The constructs analyzed consisted of the uidA (GUS) reporter gene driven by four different promoters (ubiquitin 3-potato, ubiquitin corn, ubiquitin rice and CaMV 35S). The total number of GUS hits (or transient expression units; TEUs) were determined manually under a dissecting scope after histochemical staining for GUS. Results suggest that the ubiquitin rice promoter is most active in cells of turfgrass, regardless of the developmental stage or tissue-type. The ubiquitin corn promoter was the next best. Of the four promoter used, except for ubiquitin 3-potato, reporter gene activity was dramatically higher in mature leaves compared to young leaves. The relative efficiency of each promoter was about the same in roots and leaves. We have also analyzed uidA (GUS) reporter gene activity following microprojectile bombardment in transient expression assays with callus from two cultivars (Providence or Penn A4) of creeping bentgrass. Differences in the frequency of GUS positive hits were observed between cultivars up to 72 hours post-bombardment. However, this difference between cultivars disappeared after 72 hours post-bombardment. This information describing promoter functionality in bentgrass will be important when designing gene constructs for trait modification and when choosing appropriate cultivars for improvement through gene transfer experiments. This is the first in depth report on organ-specific and developmental gene expression profiles for transgenes in a turfgrass species.
Agrostis↗
[Urinary excretion pattern of 2-thio-thiazolidine-4-carboxylic acid in workers exposed to carbon disulfide].
OBJECTIVE: To study the urinary excretion pattern for of 2-thio-thiazolidine-4-carboxylic acid (TTCA) in workers exposed to CS2, so as to provide experimental basis for working out biomonitoring measures for short-term exposure to CS2. METHODS: Sixty-nine subjects were divided into three groups: (1) fourteen volunteers who had not been exposed to CS2 before were exposed to CS2 for 2 hours, their urine samples were collected and analyzed at different time points; (2) The urine samples of 15 occupational exposure workers were collected on pre-shift, mid-shift, post-shift; (3) The relationship between 8 h time weighted average CS2 exposure concentrations (PC-TWA) and TTCA levels of post-shift urine was studied among 40 workers chronically exposed to CS2. RESULTS: (1) In the 1st group, urine TTCA level reached the peak [(1.03 +/- 0.72) mg/gCr] 4 h after exposure; (2) In the 2nd group, urine TTCA level on pre-shift [(0.37 +/- 0.28) mg/gCr] was lower than that on mid-shift [(1.23 +/- 0.71) mg/gCr, P<0.01] and post-shift [(1.31 +/- 0.78) mg/gCr, P<0.01]; (3) In the 3rd group, there was a linear relationship between the post-shift urine TTCA level and 8 h CS2 exposure concentrations dose (PC-TWA). The regression equation is Y(TTCA mg/gCr)=1.163 6X(CS2 mg/m3)-5.411 6. CONCLUSION: The post-shift urine TTCA levels may be regarded as a bio-monitoring index for workers exposed to CS2.
Adult↗
Direct interaction between nucleolin and hepatitis C virus NS5B.
Hepatitis C virus (HCV) NS5B is an RNA-dependent RNA polymerase (RdRP), a central catalytic enzyme in HCV replication. While studying the subcellular localization of a NS5B mutant lacking the C-terminal membrane-anchoring domain, NS5Bt, we found that expression of the green fluorescent protein (GFP)-fused form was exclusively nucleolar. Interestingly, the distribution of endogenous nucleolin changed greatly in the cells expressing GFP-NS5B, with nucleolin colocalized with GFP-NS5B in perinuclear regions in addition to the nucleolus, suggesting that NS5B retains the ability to bind nucleolin. The interaction between nucleolin and NS5B was demonstrated by GST pull-down assay. GST pull-down assay results indicated that C-terminal region of nucleolin was important for its binding to NS5B. Scanning clustered alanine substitution mutants library of NS5B revealed two sites on NS5B that binds nucleolin. NS5B amino acids 208-214 and 500-506 were both found to be indispensable for the nucleolin binding. We reported that the latter sequence is essential for oligomerization of NS5B, which is a prerequisite for the RdRP activity. C-terminal nucleolin inhibited the NS5B RdRP activity in a dose-dependent manner. Taken together, this indicates the binding ability of nucleolin may be involved in NS5B functions.
Animals↗
Hepatitis C virus (HCV) NS5A binds RNA-dependent RNA polymerase (RdRP) NS5B and modulates RNA-dependent RNA polymerase activity.
Hepatitis C virus (HCV) NS5B is RNA-dependent RNA polymerase (RdRP), the essential catalytic enzyme for HCV replication. Recently, NS5A has been reported to be important for the establishment of HCV replication in vitro by the adaptive mutations, although its role in viral replication remains uncertain. Here we report that purified bacterial recombinant NS5A and NS5B directly interact with each other in vitro, detected by glutathione S-transferase (GST) pull-down assay. Furthermore, complex formation of these proteins transiently coexpressed in mammalian cells was detected by coprecipitation. Using terminally and internally truncated NS5A, two discontinuous regions of NS5A (amino acids 105-162 and 277-334) outside of the adaptive mutations were identified to be independently essential for the binding both in vivo and in vitro (Yamashita, T., Kaneko, S., Shirota, Y., Qin, W., Nomura, T., Kobayashi, K., and Mkyrakami, S. (1998) J. Biol. Chem. 273, 15479-15486). We previously examined the effect of His-NS5A on RdRP activity of the soluble recombinant NS5Bt in vitro (see Yamashita et al. above). Wild NS5A weakly stimulated at first (when less than 0.1 molar ratio to NS5B) and then inhibited the NS5Bt RdRP activity in a dose-dependent manner. The internal deletion mutants defective in NS5B binding exhibited no inhibitory effect, indicating that the NS5B binding is necessary for the inhibition. Taken together, our results support the idea that NS5A modulates HCV replication as a component of replication complex.
Animals↗
Application of FLP/FRT site-specific DNA recombination system in plants.
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DNA Nucleotidyltransferases↗
[Effects of the combination of eicosapentaenoic acid and retinoic acid on the proliferation and differentiation of HL-60 cells].
OBJECTIVE: To study the effect of combination of eicosapentaenoic acid (EPA) and retinoic acid (RA) on the proliferation and differentiation of HL-60 cells and its mechanisms. METHODS: MTT was used for cell proliferation analysis, NBT reduction experiment for cell differentiation, reverse transcriptase polymerase chain reaction (RT-PCR) for retinoblastoma (RB) mRNA expression, and Western blot for RB protein (PRB) expression. RESULTS: The proliferation inhibition rates were 35.74%, 24.38% and 42.75% for RA, EPA and combination of EPA and RA. NBT reduction experiments showed that the differentiation induced by EPA and RA was 5.9 times, and by RA was 2.6 times the capacity of the control. The RB mRNA and PRB expression were not changed by EPA, but significantly decreased by the combination of EPA and RA. Moreover, the dephosphorylation rate of PRB was increased by the treatment with EPA or/and RA. CONCLUSION: The changes of RB expression and PRB phosphorylation may be one of the mechanisms of the synergistic effects of EPA and RA on the HL-60 cell proliferation and differentiation.
Antineoplastic Agents↗
Oligomeric interaction of hepatitis C virus NS5B is critical for catalytic activity of RNA-dependent RNA polymerase.
HCV NS5B is an RNA-dependent RNA polymerase (RdRP), a central catalytic enzyme for HCV replication, which has the "palm and fingers" substructure. We recently identified five novel residues critical for RdRP activity (Qin, W., Yamashita, T., Shirota, Y., Lin, Y., Wei, W., and Murakami, S. (2001) Hepatology 33, 728-737). Among them, GLU-18 and His-502, far from the catalytic center, may be involved in conformational change(s) for RdRP activity as addressed in some palm and fingers enzymes. We examined the possibility that NS5B is oligomerized, and we could detect the interaction between two different tagged NS5B proteins in vitro and transiently expressed in mammalian cells. By scanning 27 clustered and then point alanine substitutions in vivo and in vitro, Glu-18 and His-502 were found to be critical for the homomeric interaction in vivo and in vitro, strongly suggesting a close relationship between the oligomerization and RdRP activity of NS5B. All mutants with substitutions at these two residues failed to bind wild type NS5B, however E18H interacted with H502E in vitro and in vivo. Interestingly, the NS5B protein with E18H or H502E did not exhibit RdRP activity, but a mixture of the two mutant proteins did. These results clearly indicate that two residues of HCV NS5B are critical for the oligomerization that is prerequisite to RdRP activity.
Amino Acids↗