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Biomedical subjects

X Yang

Publications and source records attributed to X Yang.

At least 523 records · Page 29Linked to original sources
PubMed1999

Ferroxidase activity of ferritin: effects of pH, buffer and Fe(II) and Fe(III) concentrations on Fe(II) autoxidation and ferroxidation.

It is widely accepted that iron deposition in the iron storage protein ferritin in vitro involves Fe(II) oxidation, and that ferritin facilitates this oxidation at a ferroxidase site on the protein. However, these views have recently been questioned, with the protein ferroxidase activity instead being attributed to autoxidation from the buffer alone. Ligand exchange between another protein with ferroxidase activity and ferritin has been proposed as an alternative mechanism for iron incorporation into ferritin. In the present work, a pH stat apparatus is used to eliminate the influence of buffers on iron(II) oxidation. Here we show that the recent experiments questioning the ferroxidase activity of ferritin were flawed by inadequate pH control, that buffers actually retard rather than facilitate iron(II) oxidation, and that horse spleen ferritin has ferroxidase activity when measured under proper experimental conditions. Furthermore, high pH (7.0), a high Fe(II) concentration and the presence of Fe(III) all favour Fe(II) autoxidation in the presence or absence of ferritin.

Animals↗
PubMed1999

Targeted disruption of SMAD3 results in impaired mucosal immunity and diminished T cell responsiveness to TGF-beta.

SMAD3 is one of the intracellular mediators that transduces signals from transforming growth factor-beta (TGF-beta) and activin receptors. We show that SMAD3 mutant mice generated by gene targeting die between 1 and 8 months due to a primary defect in immune function. Symptomatic mice exhibit thymic involution, enlarged lymph nodes, and formation of bacterial abscesses adjacent to mucosal surfaces. Mutant T cells exhibit an activated phenotype in vivo, and are not inhibited by TGF-beta1 in vitro. Mutant neutrophils are also impaired in their chemotactic response toward TGF-beta. Chronic intestinal inflammation is infrequently associated with colonic adenocarcinoma in mice older than 6 months of age. These data suggest that SMAD3 has an important role in TGF-beta-mediated regulation of T cell activation and mucosal immunity, and that the loss of these functions is responsible for chronic infection and the lethality of Smad3-null mice.

Animals↗
PubMed1999

The Megakaryocyte/Platelet-specific enhancer of the alpha2beta1 integrin gene: two tandem AP1 sites and the mitogen-activated protein kinase signaling cascade.

The alpha2beta1 integrin, a collagen receptor on platelets and megakaryocytes, is required for normal platelet function. Transcriptional regulation of the alpha2 integrin gene in cells undergoing megakaryocytic differentiation requires a core promoter between bp -30 and -92, a silencer between bp -92 and -351, and megakaryocytic enhancers in the distal 5' flank. We have now identified a 229-bp region of the distal 5' flank of the alpha2 integrin gene required for high-level enhancer activity in cells with megakaryocytic features. Two tandem AP1 binding sites with dyad symmetry are required for enhancer activity and for DNA-protein complex formation with members of the c-fos/c-jun family. The requirement for AP1 activation suggested a role for the mitogen-activated protein kinase (MAPK) signaling pathway in regulating alpha2 integrin gene expression. Inhibition of the MAP kinase cascade with PD98059, a specific inhibitor of MAPK kinase 1, prevented the expression of the alpha2 integrin subunit in cells induced to become megakaryocytic. We provide a model of megakaryocytic differentiation in which expression of the alpha2 integrin gene requires signaling via the MAP kinase pathway to activate two tandem AP1 binding sites in the alpha2 integrin enhancer.

Blood Platelets↗
PubMed1999

Overexpression of anti-apoptotic gene BAG-1 in human cervical cancer.

Apoptosis is a programmed cell death process in which cells commit suicide under certain environmental conditions. Recent studies suggest that apoptosis is controlled by a variety of cellular genes, and dysregulation of these genes plays an important role in the pathogenesis of human diseases, including cancer. BAG-1 is a novel anti-apoptotic protein isolated by its interaction with another anti-apoptotic protein, Bcl-2. It binds to several hormone receptors and growth factor receptors and modulates their function in apoptosis. However, the role of BAG-1 in the oncogenesis of human cervical cancer has yet to be illustrated. In this study, we examined the expression of BAG-1 in cervical normal and carcinoma cultured cells and tissues. BAG-1 was overexpressed in human cervical carcinoma cell lines and tissues. Overexpression was regulated at the transcriptional level. The increased expression of BAG-1 was correlated with enhanced resistance of cervical carcinoma cells to apoptosis induced by a DNA-damaging reagent. In addition, overexpression of BAG-1 enhanced the resistance of cervical cells to apoptosis. This study provided the first evidence that BAG-1 is upregulated in human cervical cancer and may play an important role in apoptosis and human cervical carcinogenesis.

Apoptosis↗
PubMed1999

Dissecting Fas signaling with an altered-specificity death-domain mutant: requirement of FADD binding for apoptosis but not Jun N-terminal kinase activation.

Fas is a cell surface death receptor that regulates peripheral tolerance and lymphoid homeostasis. In many pathologic conditions, ectopic Fas activation mediates tissue destruction. Several proteins that can bind to the cytoplasmic death domain of Fas have been implicated in Fas signal transduction. Here we show that FADD, which couples Fas to pro-caspase-8, and, Daxx, which couples Fas to the Jun N-terminal kinase pathway, bind independently to the Fas death domain. We have isolated a death domain mutant, termed FasDelta, that selectively binds Daxx but not FADD. In tranfected tissue culture cells, FasDelta activated Jun N-terminal kinase normally but was impaired in cell death induction. These results suggest that FADD and Daxx activate two independent pathways downstream of Fas and confirm the essential role of FADD binding in apoptosis induction.

Adaptor Proteins, Signal Transducing↗
PubMed1999

Brain potentials during mental arithmetic-effects of problem difficulty on event-related brain potentials.

One addend '+' symbol and another addend were presented in sequence to subjects in a monitor, and event-related brain potentials (ERPs) were recorded at the same time to examine the effect of problem difficulty (with or without carrying in solution) on ERPs. After the presentation of the second addend, N1, P1, N2, late positive complex and slow waves were recorded. The P2 amplitude at F3 site for the difficult arithmetic problems between 168 and 184 ms is larger (more positive) than that for easy problems (P < 0.05). The mean latency of P2 at F7 and P3b at F3 and F4 is significantly longer for difficult problems than that for easy ones (P < 0.05). It is suggested that prefrontal activity may be involved in the arithmetic data retrieval process. ERPs is modified to different degrees by changing the difficulty of mental arithmetic.

Adult↗
PubMed1999

IL-10 gene knockout mice show enhanced Th1-like protective immunity and absent granuloma formation following Chlamydia trachomatis lung infection.

We previously reported that higher IL-10 production is correlated with lower IFN-gamma production, weaker delayed hypersensitivity (DTH), and slower organism clearance following chlamydial infection in mice. To assess more directly the role of IL-10, we examined protective immunity and pathological reaction in C57BL/6 IL-10 gene knockout (KO) and wild-type mice. The results showed that in the absence of endogenous IL-10, mice had significantly accelerated chlamydial clearance and developed significantly stronger DTH responses, which could be inhibited by local delivery of rIL-10. Consistent with the enhancement of DTH responses, IL-10 KO mice showed stronger and more persistent CD4 T cell-dependent IFN-gamma production and significant elevation of IL-12 and TNF-alpha production. Additionally, wild-type, but not IL-10 KO, mice showed granuloma formation that was correlated with higher levels of Th2 cytokine (IL-5) production at the later stages of infection. Moreover, chlamydial infection, unlike parasitic protozoan infection, did not induce significant acute toxicity in IL-10 KO mice, which may be due to the low (undetectable) levels of systemic release of proinflammatory cytokines. These results suggest that IL-10 inhibits the priming and expansion of Th1-like T cell responses and that IL-10 plays a role in the fibrotic reaction seen with chlamydial infection.

Animals↗
PubMed1999

Microsatellite instability during the immortalization and transformation of human breast epithelial cells in vitro.

The objective of this study was to determine whether microsatellite instability (MSI) and loss of heterozygosity (LOH) are involved in the immortalization of human breast epithelial cells (HBECs) in vitro and in the early stages of their transformation by benzo[a]pyrene (BP) and 7,12-dimethylbenz[a]anthracene (DMBA). We performed a genome-wide analysis of a total of 466 microsatellite DNA polymorphism loci along the X chromosome and the 22 pairs of human autosomes. MSI was found in the immortalized MCF-10F cells at the following loci: D11S1392 (on chromosome 11p13) and D17S849 (at 17p13.3), D17S796 (at 17p13.1), D17S513 (at 17p13.1), TP53 (at 17p13.1), D17S786 (at 17p13.1), and D17S520 (at 17p12) on chromosome 17. The BP-transformed cells exhibited MSI in the same loci and also in locus D11S912 (at 11q25). The more transformed BP1E cells also exhibited MSI on chromosome 13q12-13 at D13S260 and D13S289, markers known to flank the breast cancer susceptibility gene BRCA2. In the DMBA-transformed D3 and D3-1 cells, MSI was observed at the locus D13S260 in addition to the previously reported locus D16S285 (at 16q12.1). No LOH was observed on any of the chromosomes tested in these cells. These observations led us to conclude that the immortalization and transformation of HBECs may involve defects in mechanisms responsible for the cell's genomic stability, such as DNA replication and DNA mismatch repair.

Breast Neoplasms↗
PubMed1999

Timing of the first cleavage post-insemination affects cryosurvival of in vitro-produced bovine blastocysts.

The time of the first cleavage of bovine zygotes during in vitro culture can affect the rate of development and cell number of the blastocysts. The aim of this work was to study the effect of the timing of first cleavage on the cryosurvival of the resulting blastocysts. Following standard IVM and IVF, zygotes were cultured in modified synthetic oviduct fluid (SOF), with 10% fetal calf serum (FCS) added 48 hr post insemination, in a humidified atmosphere of 5% CO2, 5% O2 and 90% N2. Embryos which cleaved by 24, 27, 30, 33, or 36 hr after insemination (IVF) were harvested and further cultured to the blastocyst stage (day 7 or day 8 post IVF). All developing blastocysts on days 7 and 8 were classified into three groups and were cryopreserved by vitrification. Group A consisted of blastocysts (<150 microm, small blastocysts); group B consisted of expanded or hatching blastocysts (>150 microm, large blastocysts); and group C consisted of morphologically poor quality blastocysts. The vitrification solution consisted of 6.5 M glycerol and 6% bovine serum albumin in PBS (VS3a). Thawed embryos were cultured further and survival was defined as the re-expansion and maintenance of the blastocoel over 24, 48, and 72 hr, respectively. Overall survival and hatching at 72 hr post-thawing was higher in blastocysts formed by day 7 than those formed by day 8 (60% vs. 40% survival; 63% vs. 45% hatching). Large blastocysts from day-7 and day-8 groups survived significantly better than small or poor quality blastocysts (76% vs. 63% and 31%; 72% vs. 30% and 26%, respectively; P < 0.05). Day-7 blastocysts from the 27- and 30-hr cleavage groups survived significantly better than those from the 36-hr group (63% and 66% vs. 25%, P < 0.05). Day-8 blastocysts from later cleaved (30 hr) zygotes had a higher survival than the 27-hr cleavage groups (52% vs. 26%, P < 0.05). These results indicate that the day of blastocyst appearance, developmental stage, and timing of the first cleavage post-insemination can influence the cryosurvival of bovine blastocysts following vitrification.

Animals↗
PubMed1999

Thermotolerance of IVM-derived bovine oocytes and embryos after short-term heat shock.

A series of experiments were designed to study the effect of elevated temperatures on developmental competence of bovine oocytes and embryos produced in vitro. In experiment 1, the effect of heat shock (HS) by a mild elevated temperature (40.5 degrees C) for 0, 30, or 60 min on the viability of in vitro matured (IVM) oocytes was tested following in vitro fertilization (IVF) and culture. No significant difference was observed between the control (39 degrees C) and the heat-treated groups in cleavage, blastocyst formation, or hatching (P > 0.05). In experiment 2, when the HS temperature was increased to 41.5 degrees C, neither the cleavage rate nor blastocyst development was affected by treatment. However, the rate of blastocyst hatching appeared lower in the HS groups (13% in control group vs. 3.9% and 5.6% in 30 min and 60 min, respectively; P < 0.05). When IVM oocytes were treated at 43 degrees C prior to IVF (experiment 3), no difference was detected in blastocyst and expanded blastocyst development following heat treatment for 0, 15, or 30 min, but heat treatment of oocytes for 45 or 60 min significantly reduced blastocyst and expanded blastocyst formation (P < 0.05). In experiment 4, the thermotolerance of day 3 and day 4 bovine IVF embryos were compared. When embryos were pre-treated with a mild elevated temperature (40.5 degrees C) for 1 hr, and then with a higher temperature (43 degrees C) for 1 hr, no improvement in thermotolerance of the embryos was observed as compared to those treated at 43 degrees C alone. However, a higher thermotolerance was observed in day 4 than day 3 embryos. In conclusion, treatment at 43 degrees C, but not 40.5 degrees C or 41.5 degrees C significantly reduced oocyte developmental competence. An increase in thermotolerance was observed from day 3 to day 4 of in vitro embryonic development, which corresponds to the maternal to zygotic transition of gene expression in bovine embryos.

Animals↗
PubMed1999

Cell allocation and chromosomal complement of parthenogenetic and IVF bovine embryos.

Considerable concerns exist regarding the quality of parthenogenetically activated embryos in terms of sufficient numbers of cells comprising the inner cell mass (ICM) and trophectoderm (TE) and the ploidy. Therefore, these two parameters were used to assess the quality of embryos derived from parthenogenetic activation by using calcium ionophore A23187 (CaI) followed by either 6-dimethylaminopurine (6-DMAP, 3.5 hr or 6.5 hr) or cycloheximide (CHX) plus cytochalasin D (CD). The conventional in vitro (IVF) produced embryos served as a control. Double staining of the parthenogenetic blastocysts showed that the total cell number (TC) of embryos from the 6-DMAP 3.5 hr (87.0 +/- 5.3) and CHX+CD (79.0 +/- 6.1) groups was not different (P > 0.05), but was lower than that of control embryos (116.0 +/- 5.8, P < 0.001). The mean ratios of inner cell mass (ICM) and trophectoderm (TE) cells in the 6-DMAP 3.5 hr group (0.57 +/- 0.04) and the control IVF group (0.50 +/- 0. 02) did not differ significantly. Both were higher than those of the CHX+CD group (0.36 +/- 0.02; P < 0.05). Further analysis of chromosomal compositions of developing stage embryos at day four after IVF or parthenogenetic activation demonstrated that prolonged treatment with 6-DMAP for 6.5 hr resulted in a significantly lower percentage of diploid embryos and a significantly higher percentage of abnormal ploidy embryos compared to treatment with 6-DMAP for 3.5 hr or with CHX and IVF. In conclusion, parthenogenetic activation of bovine oocytes with CaI followed by 6-DMAP for 3.5 hr could produce better quality embryos in terms of total cell numbers, the number of cells allocated to the ICM, and the ploidy of embryos.

Adenine↗
PubMed1999

IFN-gamma knockout mice show Th2-associated delayed-type hypersensitivity and the inflammatory cells fail to localize and control chlamydial infection.

Delayed-type hypersensitivity (DTH) has been demonstrated to be a Th1 type immune response which is important in the host defense against infection with intracellular bacteria, including Chlamydia. In the present study, we surprisingly observed that C. trachomatis mouse pneumonitis MoPn-infected IFN-gamma gene knockout (KO) mice mounted strong DTH responses following foopad challenge with inactivated organisms. The DTH responses in IFN-gamma KO mice were associated with Th2 cytokine production and partially blocked by anti-IL-4 monoclonal antibodies. In addition, the inflammatory cells in IFN-gamma KO mice failed to target the cellular sites of chlamydial inclusions in infected tissues and failed to clear the infection. The data, in conjunction with previous studies, suggest that different types (Th1 and Th2 associated) of DTH responses may function differently in host defense against chlamydial infection and that the functional differences in DTH responses may account for the dual role that DTH is speculated to play in chlamydial protective immunity and immunopathology. Moreover, the data suggest that the IFN-gamma KO mouse is a useful model system for studying chlamydial pathogenesis.

Animals↗
PubMed1999

Overtone Transitions of 12CO2 beyond the Venus Bands.

Combination overtone transitions of 12CO2 much weaker than the Venus bands in the near infrared (11 700-14 200 cm-1) have been observed for the first time by using photoacoustic spectroscopy. A total of six bands from two Fermi polyad groups were recorded and both the band origins and relative intensities have been determined and compared with the calculated values. Through a comparison of the relative absorption cross sections within the same type of polyads, intensity shifts have been observed and will be discussed. Copyright 1999 Academic Press.

Journal Article↗
PubMed1999

Posttranscriptional destabilization of the bradykinin B1 receptor messenger RNA: cloning and functional characterization of the 3'-untranslated region.

We showed previously that the inducible bradykinin B1 receptor (BKB1R) gene expression is regulated, in part, through mRNA stabilization. Here we clone the 3'-untranslated region (3'-UTR) of the BKB1R. This region proves to be very short, containing only 14 bases with an alternative polyadenylation signal (AUUAAA) which overlaps with the stop codon. Reverse transcription confirms the presence of this alternative polyadenylation signal. Northern blot shows a single species of BKB1R mRNA of approximately 1.4 kb in agreement with its calculated length. The BKB1R mRNA induced by TNFalpha, phorbol ester, bradykinin, and desArg10-kallidin contain the same 3'-UTR species. To test the role of this region in the regulation of mRNA stability, we generated a chimeric luciferase construct containing the BKB1R 3'-UTR. The mRNA transcribed from the wild-type luciferase gene displayed a half-life of approximately 6 h. The mRNA transcribed from the chimeric construct displayed a half-life of only 1 h. This decrease was also reflected at the level of enzyme activity. Luciferase activity from cells transfected with the chimeric construct was 10 times less than from cells transfected with wild-type luciferase. The data presented provide compelling evidence that the 3'-UTR is participating in the regulation of BKB1R mRNA stability and its ultimate expression.

3' Untranslated Regions↗
PubMed1999

Effect of the G-protein, G alpha(i2), and G alpha(i3) subunit knockdown on bradykinin-induced signal transduction in rat-1 cells.

The bradykinin (BK) B2 receptor (BKB2R) has been shown to interact with the G alpha(q) subunit family. However, it has remained unclear whether this receptor also interacts with the G alpha(i) subunit family. To further resolve this issue, two antisense expression plasmids were generated. In these, the 5'-untranslated regions of rat G alpha(i2) and G alpha(i3) cDNAs were used as specific antisense templates. The plasmids were transfected into Rat-1 cells, which expressed a stably transfected rat BKB2R cDNA and bound BK with a Kd of approximately 3 nM. In these cells, the transfected BKB2R was fully linked to inositol phosphate production, arachidonic acid (ARA) release, and Ca2+ flux. A number of cell lines, each a G alpha(i2) or G alpha(i3) knockdown, were isolated. Of these, two cell lines were chosen for study. One, designated 2-E3, displayed over a 70% decrease in the expression of G alpha(i2) without a change in the expression of G alpha(i3) or G alpha(q). Another, 3-G9, exhibited over a 70% decrease of G alpha(i3) protein without a change in G alpha(i2) or G alpha(q) expression. Knockdown of either G alpha(i2) or G alpha(i3) protein production did not affect the binding of bradykinin. In the G alpha(i2)-depleted 2-E3 cells, BK induced ARA release was reduced by more than 60%. Interestingly, the production of total inositol phosphate in response to BK was also reduced by approximately 35%. However, G alpha(i2) knockdown had no significant effect on BK-induced Ca2+ influx. In the G alpha(i3)-depleted 3-G9 cells, BK-induced ARA release was decreased by over 50%. In this case [Ca2+]i increase in response to BK was reduced by over 50%. This knockdown also resulted in reduced BK-activated total inositol phosphate production. Moreover, cAMP augmented the BK-induced ARA release. Depletions of G alpha(i2) and G alpha(i3) further enhanced this cAMP-dependent BK induction of ARA release. Taken together, these results delineate direct interaction of the BKB2R with both G alpha(i2) and G alpha(i3) subunits in addition to the heretofore described interaction of BKB2R with the G alpha(q) subunit family.

Animals↗
PubMed1999

Preparation of adriamycin magnetic albumin microspheres and their experimental antitumor effects in vitro and in vivo.

The adriamycin magnetic microspheres (ADM-MAMs) were prepared by the heat-stabilized protein methods. Their physico-chemical properties were examined; their cytotoxicities against tumor cells in vitro were assayed by a modified MTT method, and their effects were observed on the implanted gastric tumor in Wistar rats given ADM-MAMs via alimentary canal at the presence of the external magnetic fields. The results showed that the ADM-MAMs were successfully prepared and had cytotoxic effect on tumor cells in vitro similar to the free ADM (P > 0.05). The inhibitory effects of ADM-MAMs on the implanted gastric tumor in vivo were significantly increased as compared with the controls (P < 0.01). Our results suggested that ADM-MAMs were a new type of adriamycin (ADM) preparation and its form alteration did not affect its anticancer effects.

Animals↗
PubMed1999

Experimental study on detached renal tubular epithelial cells in urine of nephropathia epidemic patients.

To elucidate the pathogenesis of acute renal failure (ARF) with nephropathia epidemic (NE), provide experimental evidence for the new therapy to NE and observe the effects of Arg-Gly-Asp (RGD) peptides on adhesion of renal tubular epithelial cell (RTEC), urine specimens of patients were collected under sterile conditions. Detached RTECs were separated, cultured and identified. Hantan Virus antigen was determined by using indirect immunofluorescence method and effects of RGD on adhesion of RTECs was observed by subgroup counting as well as by flow cytometry. This study showed that: (1) sublethal RTECs existed in the urine of NE-ARF patients, which could be cultured in monolayer form; (2) there was NE antigen in RTECs; and (3) adhesion of RTECs could be inhibited by RGD.

Acute Kidney Injury↗