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Biomedical subjects

Hao Yu

Publications and source records attributed to Hao Yu.

54 records · Page 3Linked to original sources

The spike protein of severe acute respiratory syndrome (SARS) is cleaved in virus infected Vero-E6 cells.

Spike protein is one of the major structural proteins of severe acute respiratory syndrome-coronavirus. It is essential for the interaction of the virons with host cell receptors and subsequent fusion of the viral envelop with host cell membrane to allow infection. Some spike proteins of coronavirus, such as MHV, HCoV-OC43, AIBV and BcoV, are proteolytically cleaved into two subunits, S1 and S2. In contrast, TGV, FIPV and HCoV-229E are not. Many studies have shown that the cleavage of spike protein seriously affects its function. In order to investigate the maturation and proteolytic processing of the S protein of SARS CoV, we generated S1 and S2 subunit specific antibodies (Abs) as well as N, E and 3CL protein-specific Abs. Our results showed that the antibodies could efficiently and specifically bind to their corresponding proteins from E.coli expressed or lysate of SARS-CoV infected Vero-E6 cells by Western blot analysis. Furthermore, the anti-S1 and S2 Abs were proved to be capable of binding to SARS CoV under electron microscope observation. When S2 Ab was used to perform immune precipitation with lysate of SARS-CoV infected cells, a cleaved S2 fragment was detected with S2-specific mAb by Western blot analysis. The data demonstrated that the cleavage of S protein was observed in the lysate, indicating that proteolytic processing of S protein is present in host cells.

Animals↗

Signal transducer and activator of transcription 3 (Stat3) expression and activation in rat uterus during early pregnancy.

Signal transducer and activator of transcription 3 (Stat3), a member of the Stat family, is specifically activated during mouse embryo implantation. The aim of this study was to investigate the expression, activation and regulation of Stat3 in rat uterus during early pregnancy, pseudopregnancy, delayed implantation and artificial decidualization. Stat3 mRNA was highly expressed in the luminal epithelium on day 5 and in the luminal epithelium and underlying stromal cells at implantation sites on day 6 of pregnancy. There was a strong level of Stat3 protein expression and phosphorylation in the stromal cells near the lumen and in the luminal epithelium on day 5 of pregnancy, which was similar to day 5 of pseudopregnancy. In the afternoon of day 6, the strong level of Stat3 phosphorylation was detected only in the luminal epithelium. Stat3 was highly expressed and activated in the decidual cells from days 7 to 9 of pregnancy and under artificial decidualization in the present study. Our results suggest that the strong level of Stat3 activation in the luminal epithelium and underlying stromal cells during the pre-implantation period may be important for establishing uterine receptivity as in mice, and the high level of Stat3 expression and activation in decidual cells may play a role during decidualization.

Animals↗

[Vertical mammaplasty using the skin wrinkling suture techniques].

OBJECTIVE: To reduce the incision scar produced by reduction mammaplasty. METHODS: The periareola incision and the vertical incision below the areola were made when performing breast reduction. The incision in the submammary fold was avoided. The breast was suspended and moulded. The redundant skin and the long skin incision below the areola were sutured utilizing the skin wrinkling suture techniques. The wrinkling skin and incision became flat gradually due to the elasticity of the tissue itself. RESULTS: 13 cases have been treated with the techniques mentioned above. All incisions healed primarily and there were no complications such as hematoma, infection and necrosis of skin or areola. Of them, 3 patients with very large and ptotic breasts needed small corrective operations to excise the redundant skin that still existed 6 months postoperatively. The shape of the breasts and the vertical incision scars of the other patients were satisfactory. CONCLUSIONS: The vertical mammaplasty using the skin wrinkling suture techniques can be used in breast reduction of various sizes, owing to the advantages of consistent, good and stable result, simplicity to perform and grasp and no scar in the submammary fold.

Adult↗

Investigation of cytokinin-deficient phenotypes in Arabidopsis by ectopic expression of orchid DSCKX1.

The plant hormone cytokinin plays a major role in regulating plant growth and development. Here we generated cytokinin-reduction Arabidopsis plants by overexpressing a heterologous cytokinin oxidase gene DSCKX1 from Dendrobium orchid. These transgenic plants exhibited reduced biomass, rapid root growth, decreased ability to form roots in vitro, and reduced response to cytokinin in growing calli and roots. Furthermore, the expression of KNAT1, STM, and CycD3 genes was significantly reduced in the transgenic plants, suggesting that cytokinin may function to control the cell cycles and shoot/root development via regulation of these genes.

Arabidopsis↗

A critical role for ATP in the stimulation of retinal guanylyl cyclase by guanylyl cyclase-activating proteins.

It has been believed that retinal guanylyl cyclase (retGC), a key enzyme in the cGMP recovery to the dark state, is solely activated by guanylyl cyclase-activating proteins (GCAPs) in a Ca2+-sensitive manner. However, a question has arisen as to whether the observed GCAP stimulation of retGC is sufficient to account for the cGMP recovery because the stimulated activity measured in vitro is less than the light/GTP-activated cGMP phosphodiesterase activity. Here we report that the retGC activation by GCAPs is larger than previously reported and that a preincubation with adenine nucleotide is essential for the large activation. Under certain conditions, ATP is two times more effective than adenylyl imidodiphosphate (AMP-PNP), a hydrolysis-resistant ATP analog; however, this study mainly used AMP-PNP to focus on the role of adenine nucleotide binding to retGC. When photoreceptor outer segment homogenates are preincubated with AMP-PNP (EC50 = 0.65 +/- 0.20 mM), GCAP2 enhanced the retGC activity 10-13 times over the control rate. Without AMP-PNP, GCAP2 stimulated the control activity only 3-4-fold as in previous reports. The large activation is due to a GCAP2-dependent increase in Vmax without an alteration of retGC affinity for GCAP2 (EC50 = 47.9 +/- 2.7 nM). GCAP1 stimulated retGC activity in a similar fashion but with lower affinity (EC50 = 308 nM). In the AMP-PNP preincubation, low Ca2+ concentrations are not required, and retGC exists as a monomeric form. This large activation is accomplished through enhanced action of GCAPs as shown by Ca2+ inhibition of the activity (IC50 = 178 nM). We propose that retGC is activated by a two-step mechanism: a conformational change by ATP binding to its kinase homology domain under high Ca2+ concentrations that allows large enhancement of GCAP activation under low Ca2+ concentrations.

Adenine↗

Beta1 integrin and alpha-dystroglycan binding sites are localized to different laminin-G-domain-like (LG) modules within the laminin alpha5 chain G domain.

Laminins are a group of extracellular-matrix proteins important in development and disease. They are heterotrimers, and specific domains in the different chains have specialized functions. The G domain of the alpha5 chain has now been produced in transfected mammalian cells as single modules and two tandem arrays, alpha5LG1-3 and alpha5LG4-5 (LG is laminin G domain-like). Using these fragments we produced specific polyclonal antibodies functional in immunoblotting and immunofluorescence studies and in solid-phase assays. Both alpha5LG tandem arrays had physiologically relevant affinities for sulphated ligands such as heparin and sulphatides. Cells adhered to these fragments and acquired a spread morphology when plated on alpha5LG1-3. Binding of integrins alpha3beta1 and alpha6beta1 was localized to the alpha5LG1-3 modules, and alpha-dystroglycan binding was localized to the alpha5LG4-5 modules, thus locating these activities to different LG modules within the laminin alpha5 G domain. However, both these activities were of relatively low affinity, indicating that integrin-mediated cell adhesion to the laminin 10/11 alpha5G domain depends on contributions from the other chains of the heterotrimer and that high-affinity alpha-dystroglycan binding could be dependent on specific Ca(2+)-ion-co-ordinating amino acids absent from alpha5LG4-5.

Animals↗

Opposing roles of integrin alpha6Abeta1 and dystroglycan in laminin-mediated extracellular signal-regulated kinase activation.

Laminin-integrin interactions can in some settings activate the extracellular signal-regulated kinases (ERKs) but the control mechanisms are poorly understood. Herein, we studied ERK activation in response to two laminins isoforms (-1 and -10/11) in two epithelial cell lines. Both cell lines expressed beta1-containing integrins and dystroglycan but lacked integrin alpha6beta4. Antibody perturbation assays showed that both cell lines bound to laminin-10/11 via the alpha3beta1and alpha6beta1 integrins. Although laminin-10/11 was a stronger adhesion complex than laminin-1 for both cell lines, both laminins activated ERK in only one of the two cell lines. The ERK activation was mediated by integrin alpha6beta1 and not by alpha3beta1 or dystroglycan. Instead, we found that dystroglycan-binding domains of both laminin-1 and -10/11 suppressed integrin alpha6beta1-mediated ERK activation. Moreover, the responding cell line expressed the two integrin alpha6 splice variants, alpha6A and alpha6B, whereas the nonresponding cell line expressed only alpha6B. Furthermore, ERK activation was seen in cells transfected with the integrin alpha6A subunit, but not in alpha6B-transfected cells. We conclude that laminin-1 and -10/11 share the ability to induce ERK activation, that this is regulated by integrin alpha6Abeta1, and suggest a novel role for dystroglycan-binding laminin domains as suppressors of this activation.

Cytoskeletal Proteins↗

Functional characterisation of a cytokinin oxidase gene DSCKX1 in Dendrobium orchid.

Cytokinin oxidase plays an important role in the cytokinin regulatory processes. We have cloned a novel putative cytokinin oxidase, DSCKX1 (Dendrobium Sonia cytokinin oxidase), by mRNA differential display from shoot apices of Dendrobium Sonia cultured in the presence of BA. The DSCKX1 gene appears to have three alternative splicing forms and its expression of DSCKX1 was induced in a tissue-specific manner by cytokinins. In transgenic orchid plants overexpressing DSCKX1, the elevated level of cytokinin oxidase activity was accompanied by a reduction of cytokinin content. These plants exhibited slow shoot growth with numerous and long roots in vitro. Their calli also showed decreased capability of shoot formation. Conversly, antisense transgenic plants showed rapid proliferation of shoots and inhibition of root growth combined with a higher endogenous cytokinin content than wild-type plants. Thus DSCKX1 appears to play an important role on cytokinin metabolism and the related developmental programmes in orchid.

Adenine↗

[Cloning, expression, and antibody production of mouse ISP2].

A cDNA encoding mouse implantation serine proteinase 2 ISP2 was amplified from total cDNAs of mouse uterus implantation sites on D4.5 of pregnancy by PCR, and sequenced GenBank accession No. A442918 . DNA sequencing indicated that the ISP2 cDNA had an unreported 204 bp sequence at 3' untranslated region besides the open reading frame encoding 279 amino acid residues, which was identical with literature. In order to obtain recombinant ISP2 rISP2 an expression plasmid pGEX-4T-2/ISP2 was constructed and transformed into E.coli BL21(DE3) strain. Expressed fusion protein GST-ISP2 was purified by SDS-PAGE and digested with thrombin, and the digestion mixture was subjected to SDS-PAGE again to recover rISP2. Rabbits were immunized using rISP2 as immunogen, and the polyclonal anti-ISP2 antisera were obtained. Immunohistochemical analysis using this antisera showed specific and high expression of ISP2 in mouse endometrial gland epithelium in early pregnancy.

Amino Acid Sequence↗

[Expression and purification of recombinant SARS coronavirus spike protein].

A novel coronavirus (SARS-coronavirus, SARS-CoV) was discovered in association with cases of severe acute respiratory syndrome (SARS) recently. The first step in coronavirus infection is binding of the viral spike protein to certain receptor on host cells. The spike protein is the main surface antigen of the coronavirus and there should be antibodies against spike protein in patients serum. Thus, to develop and expression protein fragment from spike protein gene are the purposes of this experiment. Partial spike gene fragments (751-1925 bp, 2005-3410 bp, 1-1925 bp and 32-3659 bp) and its intact gene were cloned into pET32 or pGEX vectors, and transformed into competent Escherichia coli BL21(DE3) (pLysS), respectively. 63, 78, 98, 160 and 164 kD fusion proteins were successfully expressed with amounts of 35%, 34%, 24%, 17% and 5% of total cell protein. The soluble parts of the cell crude extract were then partially purified by GST affinity chromatography.

Blotting, Western↗

The evolution of the State Children's Health Insurance Program (SCHIP) in New York: changing program features and enrollee characteristics.

BACKGROUND: The State Children's Health Insurance Program (SCHIP) has been operating for >5 years. Policy makers are interested in the characteristics of children who have enrolled and changes in the health care needs of enrolled children as programs mature. New York State's SCHIP evolved from a similar statewide health insurance program that was developed in 1991 (Child Health Plus [CHPlus]). Understanding how current SCHIP enrollees differ from early CHPlus enrollees together with how program features changed during the period may shed light on how best to serve the evolving SCHIP population. OBJECTIVE: To 1) describe changes in the characteristics of children enrolled in 1994 CHPlus and 2001 SCHIP; 2) determine if changes in the near-poor, age-eligible population during the time period could account for the evolution of enrollment; and 3) describe changes in the program during the period that could be responsible for the enrollment changes. SETTING: New York State, stratified into 4 regions: New York City, New York City environs, upstate urban counties, and upstate rural counties. DESIGN: Retrospective telephone interviews of parents of 2 cohorts of CHPlus enrollees: 1) children who enrolled in CHPlus in 1993 to 1994 and 2) children who enrolled in New York's SCHIP in 2000 to 2001. The Current Population Survey (CPS) 1992 to 1994 and 1999 to 2001 were used to identify secular trends that could explain differences in the CHPlus and SCHIP enrollees. PROGRAM CHARACTERISTICS: 1994 CHPlus and 2001 SCHIP were similar in design, both limiting eligibility by age, family income, and insurance status. SCHIP 2001 included 1) expansion of eligibility to adolescents 13 to 19 years old; 2) expansion of benefits to include hospitalizations, mental health, and dental benefits; 3) changes in premium contributions; 4) more participating insurance plans, limited to managed care; 5) expansions in marketing and outreach; and 6) a combined enrollment application for SCHIP and several low-income programs including Medicaid. SAMPLE: Cohort 1 included 2126 new CHPlus enrollees 0 to 13 years old who were enrolled for at least 9 months, stratified by geographic region. Cohort 2 included 1100 new SCHIP enrollees 0 to 13 years old who were enrolled for at least 9 months, stratified by geographic region, age, race, and ethnicity. Results were weighted to be representative of statewide CHPlus or SCHIP new enrollees who met the sampling criteria. Samples of age- and income-eligible children from New York State were drawn from the CPS and pooled and reweighted (1992-1994 and 1999-2001) to generate a comparison group of children targeted by CHPlus and SCHIP. MEASURES: Sociodemographic characteristics, race and ethnicity (white non-Hispanic, black non-Hispanic, and Hispanic), prior health insurance, health care access, and first source of information about the program. ANALYSES: Weighted bivariate analyses (comparisons of means and rates) adjusted for the complex sampling design to compare measures between the 2 program cohorts and between the 2 CPS samples. We tested for equivalence by using chi2 statistics. RESULTS: As the program evolved from CHPlus to SCHIP, relatively more black and Hispanic children enrolled (9% to 30% black from 1994 to 2001, and 16% to 48% Hispanic), more New York City residents (46% to 69% from 1994 to 2001), more children with parents who had less than a high school education (10% to 25%), more children from lower income families (59% to 75% below 150% of the federal poverty level), and more children from families with parents not working (7% to 20%) enrolled. These socioeconomic and demographic changes were not reflected in the underlying age- and income-eligible population. A greater proportion of 2001 enrollees were uninsured for some time immediately before enrollment (57% to 76% had an uninsured gap), were insured by Medicaid during the year before enrollment (23% to 48%), and lacked a USC (5% to 14%). Although "word of mouth" was the most common means by which families heard about both programs, a greater proportion of 2001 enrollees learned about SCHIP from marketing or outreach sources. CONCLUSION: As New York programs for the uninsured evolved, more children from minority groups, with lower family incomes and education, and having less baseline access to health care were enrolled. Although changes in the underlying population were relatively small, progressively increased marketing and outreach, particularly in New York City, the introduction of a single application form for SCHIP and Medicaid, and expansions in the benefit package may have accounted, in part, for the large change in the characteristics of enrollees.

Adolescent↗

[The psychological effect of minesweeping infrasonic field].

OBJECTIVE: To investigate the effect of infrasonic field of minesweeper on psychology of minesweeper crews. METHODS: An experimental ship was selected to conduct infrasonic minesweeping, a control ship of the same type was selected to operate likewise with the infrasound generator turned off, and another group of coast-servicemen was chosen as blank control. Attention-span test, digit memory test, two-digital numbers list and profile of mood states (POMS) were used to test the ship crews and coast-servicemen. RESULTS: There were no significant differences in psychological parameters between the two ship crews before the experiment. But the scores of two-digit figures list in ship crews were significant lower than that in coast-servicemen, (31.2 +/- 11.8, 36.4 +/- 14.5 respectively vs 45.8 +/- 13.9, P < 0.05). POMS showed that the scores of anger-hostility, fatigue-inertia, and confusion-bewilderment in both ship crew groups [(15.5 +/- 6.4, 18.3 +/- 6.8), (12.1 +/- 5.0,12.3 +/- 4.9), (11.6 +/- 4.4, 12.5 +/- 4.8), respectively] were higher than those in coast-servicemen (13.9 +/- 7.0, 7.6 +/- 4.1, 8.2 +/- 4.3, respectively, P < 0.05 or 0.01), while vigor-activity in experimental group (15.0 +/- 5.9) was lower than that in coast-servicemen (19.7 +/- 4.7). After the experiment, the scores of experimental crews in digit memory test (14.5 +/- 5.0), vigor-activity (12.2 +/- 5.8), fatigue-inertia (15.8 +/- 6.2) were significantly different from those of control crews (17.3 +/- 4.8, 16.5 +/- 4.6, 11.3 +/- 5.1). CONCLUSIONS: The infrasonic field from minesweeper may injure the crew's psychological health and some neurobehavioral function.

Adult↗

AGAMOUS-LIKE 24, a dosage-dependent mediator of the flowering signals.

The most dramatic phase change in plants is the transition from vegetative to reproductive growth. This flowering process is regulated by several interacting pathways that monitor both the developmental state of the plants and environmental cues such as light and temperature. The flowering-time genes FLOWERING LOCUS T (FT) and SUPPRESSOR OF OVEREXPRESSION OF CO1 (SOC1), together with the floral meristem identity gene LEAFY (LFY), are three essential regulators integrating floral signals from multiple pathways in Arabidopsis thaliana. Part of the crosstalk among these genes is mediated by a putative transcription factor, AGAMOUS-LIKE 24 (AGL24). This gene is gradually activated in shoot apical meristems during the floral transition and later located in the whole zone of both inflorescence and floral meristems. Loss and reduction of AGL24 activity by double-stranded RNA-mediated interference result in late flowering, whereas constitutive overexpression of AGL24 causes precocious flowering. The correlation between the level of AGL24 accumulation and the alteration of flowering time suggests that AGL24 is a dosage-dependent flowering promoter. Analysis of AGL24 expression in various flowering-time mutants shows that it is regulated in several floral inductive pathways. Further genetic analyses of epistasis indicate that AGL24 may act downstream of SOC1 and upstream of LFY.

Arabidopsis↗

Spatial and temporal expression of the orchid floral homeotic gene DOMADS1 is mediated by its upstream regulatory regions.

The orchid floral homeotic gene, DOMADSI, is a marker gene specifically expressed in the transitional shoot apical meristem during floral transition in Dendrobium Madame Thong-In. DOMADSI is not detectable in vegetative tissues except a weak expression in the stem. Its transcript is uniformly localized in both of the inflorescence meristem and floral primordia, and later expressed in almost all of the floral organs. We isolated and sequenced a 3.5 kb DOMADSI promoter fragment upstream of the transcription start site, demonstrating the location of several putative DNA-binding sites, through which MADS-box and class I knox genes may modulate the DOMADSI expression. To gain insight into the molecular basis of the regulation of DOMADS1, deletion analysis of the DOMADSI::beta-glucuronidase (GUS) gene fusions was performed by means of the stable orchid transformation systems. The study shows that the full-length upstream promoter sequence confers the same spatial and temporal GUS staining pattern as that of the distribution of DOMADSI RNA during orchid development. We also identified the distinct cis-acting regulatory regions required for the control of DOMADS1 expression in vegetative and reproductive tissues, as well as the shoot apical meristem during floral transition.

5' Flanking Region↗

Isolation and characterization of the orchid cytokinin oxidase DSCKX1 promoter.

The orchid DSCKX1 is a new member of the cytokinin oxidase gene family, which catalyses the degradation of cytokinins bearing unsaturated isoprenoid side chains. A 3.7 kb fragment upstream of the DSCKX1 coding region was isolated, sequenced and characterized by deletion analysis of DSCKX1::beta-glucuronidase gene fusions using transient orchid and stable Arabidopsis transformation systems. Functional analysis of 5' deletions defined the 5'-upstream region that directs the expression in distinct tissues. Regulatory elements affecting the cytokinin induction of the DSCKX1 gene have also been delineated

5' Flanking Region↗

A novel role of RGS9: inhibition of retinal guanylyl cyclase.

Cyclic GMP plays a key role in retinal phototransduction and its photoreceptor concentration is precisely controlled by the cooperative action of cGMP phosphodiesterase (PDE) and retinal guanylyl cyclase (retGC). However, studies of the relationship between these two systems have focused only on a Ca2+-mediated, indirect connection. This article summarizes our studies strongly suggesting that RGS9-1 is directly involved in the cooperative action of PDE and retGC, and that this ingenious mechanism plays an important role in tuning of cGMP concentration in photoreceptors.

Animals↗

The Characteristics of DCPIPH(2) right curved arrow MV Electron Transport in Rb. sphaeroides 601.

The absorption spectrum and fluorescence emission spectrum of RS601 were found to keep the typical characteristics of those of the purple nonsulfide bacteria Rb. sphaeroides. Under illumination, methyl viologen was reduced by RS601 chromatophores in the presence of DCPIPH(2) as the electron donor, setting up a standard noncyclic electron transport. o-phenanthroline with I(50) of 1.0 mM inhibited the DCPIPH(2) right curved arrow MV electron transport. Antimycin A did not inhibit the DCPIPH(2) right curved arrow MV electron transport and had no I(50). The results suggested that the exact site where methyl viologen accepted electron should locate between the secondary electron acceptor, Q(B), and cyt b, but not at the Q(A) binding site as indicated before. The difference of electron transport between reduced sides of reaction centers of Rb. sphaeroides and Rs. rubrum was discussed.

Journal Article↗

Comparison of Different Effects of Chloroacetates on Electron Transports in PS II and in the Reaction Center of Rb. sphaeroides 601.

Chloroacetates displayed different effects on electron transports in the photosynthetic reaction center of purple bacteria (Rb.sphaeroides 601) and in the photosystem II (PS II) of higher plants. Decays of chlorophyII a fluorescence measured after actinic flashes show that chloroacetates inhibit the electron transport from Q(A)(-) to Q(B) (Q(B)(-)) and the equilibrium between Q(A)(-)Q(B) (Q(B)(-)) and Q(A)Q(B)(-) (Q(B)(2-)), acting on electron transport as well as proton transduction. The study on PSII electron transport indicates another inhibition site of chloroacetate at the oxidation side of PSII. Chloroacetates up to 500 mM have no inhibition on the DCPIPH(2) right curved arrow MV electron transport in the RS601's chromatophores. Dissociation constants of OP and trichloroacetates from RS601 reaction center were assessed to be 1.1x10(-3) M and from Dixon curve respectively. The differences on aspects of structures and functions between RS601 reaction center and PS II were discussed.

Journal Article↗