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Biomedical subjects

Y X Zhao

Publications and source records attributed to Y X Zhao.

At least 19 recordsLinked to original sources

Expressed sequence tags from a NaCl-treated Suaeda salsa cDNA library.

Past efforts to improve plant tolerance to osmotic stress have had limited success owing to the genetic complexity of stress responses. The first step towards cataloging and categorizing genetically complex abotic stress responses is the rapid discovery of genes by the large-scale partial sequencing of randomly selected cDNA clones or expressed sequence tags (ESTs). Suaeda salsa, which can survive seawater-level salinity, is a favorite halophytic model for salt tolerant research. We constructed a NaCl-treated cDNA library of Suaeda salsa and sequenced 1048 randomly selected clones, out of which 1016 clones produced readable sequences (773 showed homology to previously identified genes, 227 matched unknown protein coding regions, 16 anomalous sequences or sequences of bacterial origin were excluded from further analysis). By sequence analysis we identified 492 unique clones: 315 showed homology to previously identified genes, 177 matched unknown protein coding regions (101 of which have been found before in other organisms and 76 are completely novel). All our EST data are available on the Internet. We believe that our dbEST and the associated DNA materials will be a useful source to scientists engaging in stress-tolerance study.

Blotting, Northern↗

HAL1 mediate salt adaptation in Arabidopsis thaliana.

The yeast HAL1 gene was introduced into Arabidopsis thaliana by Agrobacterium tumefaciens-mediated transformation with vacuum infiltration under the control of CaMV 35S promoter. Thirty-three individual kanamycin resistant plants were obtained from 75,000 seeds. Southern blotting analysis indicated that HAL1 gene had been integrated into all of the transgenic plants' genomes. The copy number of HAL1 gene in transgenic plants was mostly 1 to 3 by Southern analysis. Phenotypes of transgenic plants have no differences with wild type plants. Several samples of transformants were self-pollinated, and progenies from transformed and non-transformed plants (controls) were evaluated for salt tolerance and gene expression. Measurement of concentrations of intracellular K+ and Na+ showed that transgenic lines were able to retain less Na+ than that of the control under salt stress. Results from different tests indicated the expression of HAL1 gene promotes a higher level of salt tolerance in vivo in the transgenic Arabidopsis plants.

Adaptation, Physiological↗

A fuzzy rule-based approach to recognizing 3-D arm movements.

One particular application of gesture-based systems is to implement a speaking aid for the deaf. For this to happen, it requires a module that can recognize three-dimensional (3-D) arm movements since arm movement is one of the four main attributes characterizing a sign word. In this paper we propose a fuzzy rule-based recognition method. The effectiveness of this system is evaluated by recognizing 3-D arm movements involved in Taiwanese Sign Language (TSL).

Arm↗

[The production and scavenging of reactive oxygen species in plants].

The imposition of environmental stress leads to increased production of reactive oxygen species (ROS) in plant cells, Which can damage proteins, membrane lipids, DNA and other cellular components. Plants have evolved enzymatic and no-enzymatic protection mechanisms that efficiently scavenge ROS. Enzymatic detoxication system includes superoxide dismutase (SOD), ascorbate peroxidase (APX), catalase (CAT) and glutathione peroxidase (GAX); no-enzymatic antioxidants include ascorbic acid, glutathione, mannitol and carotenoids, which occur in high concentrations in plants. The over-expression and accumulation strategies of these antioxidants in plants have been followed up to now, and have gained many transgenic plants showing increased stress tolerance.

Genetic Engineering↗

[Cloning of HAL1 gene and characterization for salt tolerance tomato].

The HAL1 gene was cloned by PCR strategy and confirmed by sequencing. Its open read frame is 879 bp, encoding a peptide of 294 amino acids (32 kD Protein). A chimaeric construct of HAL1 and Npt II (neomycin phosphotransferase II) was constructed and introduced into commercial cultivars of tomato (Zhong SU No. 5: Lycopersicon escullentum) by Agrobacterium tumefacien-mediated gene transformation. Transformants were selected for their ability to grow and root on media containing kanamycin. Transformation was confirmed by analysis of PCR, Southern blot and RT-PCR. The salt tolerance of transgenic tomato is evaluated by comparing the fresh weight, dry weight, Na+, K+ content of transgenic tomato and control tomato. It is concluded that the over-expressing of HAL1 in tomato could enhance the salt tolerance of the transgenic tomato.

Cloning, Molecular↗

Tumor necrosis factor receptor p55-deficient mice respond to acute Yersinia enterocolitica infection with less apoptosis and more effective host resistance.

Tumor necrosis factor (TNF) has generally been regarded as a protective cytokine in host defense against bacterial infections. In the present study, we evaluated the role of TNF in the acute phase of infection by Yersinia enterocolitica by using mice rendered genetically deficient in TNF receptor p55 (TNFRp55(-/-)). Unexpectedly, TNFRp55(-/-) mice showed more effective resistance to the bacteria, reflected in enhanced bacterial clearance and less tissue damage, than did control C57BL/6 mice. C57BL/6 mice showed evidence of extensive apoptosis in the spleen accompanied by a selective decrease in the CD4(+)-T-cell population of splenocytes, whereas TNFRp55(-/-) mice were spared these changes. The splenocytes from TNFRp55(-/-) mice also maintained a robust gamma interferon IFN-gamma response to mitogenic stimulation, while the comparable response in C57BL/6 mice was impaired. In addition, splenocytes harvested from infected mice demonstrated lower production of interleukin-10 IL-10 in TNFRp55(-/-) mice than in C57BL/6 mice. These findings suggest that Yersinia can induce TNFRp55-mediated apoptosis of splenocytes in the acute phase of the infection and that alteration of T-cell-generated cytokines can dramatically alter the early events in host defense against this pathogen.

Acute Disease↗

Secretion of complement components of the alternative pathway (C3 and factor B) by the human alveolar type II epithelial cell line A549.

The complement system participates in the local immune system of the lung. In this study, we investigated the secretion of complement components of the alternative pathway (C3 and factor B) in the alveolar type II epithelial cell line A549. The levels of C3 and factor B protein in the culture medium were determined by enzyme-linked immunosorbent assay (ELISA). The C3 and factor B mRNA expression was assessed by reverse transcription polymerase chain reaction (RT-PCR). The addition of interleukin (IL)-1beta or tumor necrosis factor (TNF)-alpha induced a dose- and time-dependent increase in C3 and factor B secretion. The addition of IL-6 or interferon (IFN)-gamma also induced a weak but significant increase in C3 and factor B secretion. These responses at the protein level were also observed at the mRNA level. Furthermore, the combination of IL-1beta plus TNF-alpha induced a marked increase in C3 and factor B secretion. Similarly, IL-6 and IFN-gamma potently enhanced IL-1beta- or TNF-alpha-induced C3 and factor B secretion, respectively. In this study, we demonstrated C3 and factor B secretion in A549 cells, and showed that the proinflammatory cytokines, IL-1beta, IL-6, TNF-alpha and IFN-gamma, acted as potent inducers of C3 and factor B secretion. It is likely that alveolar type II epithelial cells are the local sites of complement biosynthesis, and that various cytokines act as regulators of this local immune protection system.

Cell Line↗

Tumor necrosis factor receptor p55 controls the severity of arthritis in experimental Yersinia enterocolitica infection.

OBJECTIVE: To dissect the host defense mechanisms in relation to the development of Yersinia-associated arthritis by evaluating the impact of tumor necrosis factor receptor p55 (TNFRp55) deficiency on Yersinia enterocolitica infection. METHODS: TNFRp55-/- and C57BL/6 mice were inoculated intravenously with arthritogenic strain 8081 of Yenterocolitica serotype 0:8. Mice were observed daily for generating survival curves and monitoring arthritis. In subsequent sets of experiments, mice were sacrificed at day 14 after infection for examination of histopathology of joints, bacterial clearance, macrophage microbicidal activity, nitric oxide (NO) production, oxidative burst generation, and cytokine production. RESULTS: There was an 80% mortality rate in TNFRp55-/- mice compared with 25% in the controls at 8 weeks after inoculation with 70 colony-forming units of Y. enterocolitica 0:8. Histologic examination of joint tissues revealed that TNFRp55-/- mice developed more severe arthritis, including cartilage degradation and bony destruction, than controls at day 14 after infection. The more extensive joint pathology in TNFRp55-/- mice was correlated with the higher bacterial load in liver, spleen, and lungs, and with the increased levels of interleukin-10. TNFRp55-/- mice displayed impaired intracellular killing of bacteria by macrophages. This was associated with decreased NO production and impaired oxidative burst activity. CONCLUSION: This study demonstrates that TNF signaling through TNFRp55 controls the severity of Yersinia-induced arthritis and implicates TNF-mediated macrophage microbicidal activity as a central event in this process.

Animals↗

Prepubertal genistein treatment modulates TGF-alpha, EGF and EGF-receptor mRNAs and proteins in the rat mammary gland.

We have previously demonstrated that exposure to genistein early in life protects against chemically-induced mammary cancer in rats. To gain insight into the mechanism of action, we have investigated the expression of the EGF-signaling pathway in the mammary glands of 21 and 50 day old rats treated on days 16, 18, and 20 postpartum with 500 microg genistein/g body weight (B.W.) or an equivalent volume of the vehicle, dimethylsulfoxide (DMSO). This prepubertal genistein treatment up-regulated TGF-alpha and the EGF-receptor (EGFR), but not EGF, in mammary terminal ductal structures at day 21 postpartum. TGF-alpha, EGF and EGFR mRNA levels were similar in 21 day old control- and genistein-treated animals. At day 50 postpartum, mammary glands of genistein treated rats had more lobules and fewer terminal end buds (TEBs) and terminal ducts (TDs), i.e. they were more differentiated. TGF-alpha mRNA levels were down-regulated in TEB of proestrous and estrous females; EGF mRNA levels were down-regulated in TDs of proestrous, but not in estrous females; and EGFR mRNA levels were not altered in 50 day old proestrous or estrous female rats. EGFR immunostaining intensity was decreased in TEBs, but not in the total gland. EGF was increased in TEBs and TDs. TGF-alpha, EGF and EGFR were also observed in the stroma and fat pad, but genistein treatment did not alter the expression of these proteins in those locations. TGF-alpha, but not EGF and EGFR, immunostaining was observed in cell nuclei (not modulated by genistein), suggesting that this growth factor may act directly on nuclear events such as transcription and DNA replication. For comparative purposes, prepubertal diethylstilbestrol treatment was investigated and found to decrease EGFR immunostaining intensity and total IHC staining in all terminal ductal structures. We conclude that prepubertal genistein treatment directly stimulates TGF-alpha and EGFR to enhance mammary gland differentiation. This programs the differentiated cells for a down-regulated EGF-signaling pathway in TEBs and TDs of adult mammary glands. Reduced EGFR expression at time of carcinogen exposure may account for genistein programming against mammary cancer.

Adipose Tissue↗

The dual role of interferon-gamma in experimental Staphylococcus aureus septicaemia versus arthritis.

To evaluate the role of interferon-gamma (IFN-gamma) in Staphylococcus aureus infection, we investigated the effects of supplementation with and neutralization of IFN-gamma during septicaemia and arthritis in a murine model. In vivo administration of IFN-gamma both before and after bacterial inoculation significantly decreased mortality on one hand but enhanced the development of arthritis on the other. Treatment of mice with anti-IFN-gamma monoclonal antibodies (mAb) before and after bacterial inoculation did not significantly influence the survival rate but decreased the frequency and severity of arthritis. The beneficial effect of supplementation with IFN-gamma on septicaemia was correlated to the increased phagocytosis and bacterial clearance from liver and kidneys. The down-regulation of the development of arthritis by anti-IFN-gamma mAb was accompanied by the decreased serum tumour necrosis factor-alpha, interleukin-6 and interleukin-1 beta levels. These results demonstrate a significant role for IFN-gamma in simultaneous protection against septicaemia but promotion for the development of septic arthritis.

Animals↗

V beta 11+ T-lymphocyte expansion by toxic shock syndrome toxin-1 differs in mice bearing H-2q versus H-2b haplotypes.

We have recently demonstrated that toxic shock syndrome toxin-1 (TSST-1) expanded V beta 11+ T lymphocytes contribute to Staphylococcus aureus arthritis and sepsis-induced mortality. Interestingly, V beta 11+ T-cell mediated joint pathology varies in different mouse strains. In this study, we characterized the in vitro pattern of V beta 11+ T-cell expansion by TSST-1 in mice with various genetic backgrounds. Mice expressing major histocompatibility complex (MHC) class II I-E molecules did not expand V beta 11+ T cells upon stimulation with TSST-1. Using B10 congeneic I-E negative mouse strains, we found that the TSST-1-expanded V beta 11+ T cells in B10Q (H-2q) and B10M (H-2f) mice but not in B10B (H-2b) mice. Antigen-presenting cells (APC) from B10Q mice, L cells and lymphoma cell line transfected with a q gene did not restore the deficient V beta 11+ T-cell expansion by TSST-1 in purified T cells from B10B mice. In contrast, I-Ab APC were able to stimulate V beta 11+ T cells from H-2q mice. Furthermore, V beta 11+ T cells in H-2b mice did expand when exposed to staphylococcal enterotoxin A (SEA). These findings suggest that the T-cell repertoire, skewed by clonal deletion and inactivation of self-reactive T cells, accounts for the different magnitude of V beta 11+ T-cell expansion among the different mouse strains.

Animals↗

[Effects of smoking on the lipid peroxides antioxidation and alpha 1-antitrypsin].

OBJECTIVE: Detecting free radicals and alpha 1-antitrypsin (alpha 1-AT) from healthy smoker's blood to study the action of free radicals in smoking induced diseases. METHOD: Thiobarbituric acid method (TBA), RIA, 5-5'-dithio-2-dinitrobenzoicacid (DTNB) method, Ellman method and enzyme linked immunosorbent assay were used to detect the lipid peroxides (LPO), superoxide dismutase (SOD), glutathione peroxidase (GSH-px), glutathione (GSH) and alpha 1-AT. RESULT: In the blood of nonsmokers, LPO was 4.9 +/- 2.5 mumol/L, SOD was 878 +/- 302 ng/g Hb, GSH-px was 120 +/- 35 U, GSH was 3.4 +/- 0.7 mmol/L, alpha 1-AT was 75 +/- 11 mg/L, in the blood of smokers, they were 6.1 +/- 3.0 mumol/L, 780 +/- 282 ng/g Hb, 112 +/- 34 U, 3.1 +/- 0.8 mmol/L, 71 +/- 15 mg/L respectively. The LPO content in the blood of smokers was significant higher than that of nonsmokers (P < 0.01), but the SOD GSH-px GSH and alpha 1-AT contents measured was significant lower (P < 0.05 or P < 0.01). CONCLUSION: The increase of oxidation and decrease of antioxidation, activity of alpha 1-AT and imbalance of protease and antiprotease might play important roleg in smoking induced diseases.

Adult↗

Induction of selective anergy by toxic shock syndrome toxin-1 in CD8+ T cells.

Interferon-gamma (IFN-gamma) mRNA expression of murine splenic CD8+ T cells in response to toxic shock syndrome toxin-1 (TSST-1) was examined in vitro. In primary response to TSST-1, purified CD8+ cells expressed a low frequency of IFN-gamma mRNA and produced very little interleukin-2 (IL-2) compared with unseparated lymphocytes and purified CD4+ cells. The addition of IL-2 to the cell culture enhanced IFN-gamma mRNA expression in the CD8+ population. Upon restimulation with TSST-1, no IL-2 production and hardly any IFN-gamma mRNA expression were observed in the purified CD8+ lymphoblasts. Exogenously added IL-2 did not increase the IFN-gamma mRNA expression in purified CD8+ blasts exposed to TSST-1. In contrast, stimulation with concanavalin A induced a high frequency of IFN-gamma mRNA in purified CD8+ blasts. Moreover, TSST-1 induced strong IFN-gamma mRNA expression in unseparated T lymphoblasts and purified CD4+ populations. Interestingly, a combined immunocytochemistry and in situ hybridization technique showed a high frequency of CD8+ IFN-gamma mRNA-expressing cells in an unseparated blast population. These results demonstrate that TSST-1 induces a selective anergy within the purified CD8- T-cell compartment and that the IFN-gamma mRNA expression in CD8+ T blasts can be achieved by stimulation of unseparated T cells with TSST-1.

Animals↗

Major histocompatibility complex class II region confers susceptibility to Staphylococcus aureus arthritis.

The importance of the MHC class II region for the development of septic arthritis was studied in a murine model of haematogenously induced Staphylococcus aureus arthritis. In the first experiment MHC class II deficient mice (A beta-/-) and their heterozygous (A beta+/-) littermates were intravenously inoculated with a single dose of toxic shock syndrome toxin-1 producing S. aureus LS-1 strain. The results demonstrate that the expression of class II MHC molecules increases the prevalence and severity of arthritis. To analyse the impact of MHC class II haplotypes on the disease onset and progression the authors used congenic C3H.NB, C3H.Q and C3H/HeJ mice in the second set of experiments. The results show that C3H/HeJ mice developed the highest frequency and the most severe course of arthritis compared with C3H.NB and C3H.Q animals. Immunohistochemical analysis of arthritic joints revealed equal number of macrophages, CD4+ and CD8+ lymphocytes in the inflamed synovia in all the congenic mice. In contrast, the number of MHC class II expressing cells was higher in the arthritic joints of C3H/HeJ mice compared with the congenic strains (P < 0.001). Furthermore, serum levels of proarthrtitogenic cytokines, such as tumour necrosis factor and interleukin-6 were higher in C3H/HeJ group. This study indicates that MHC class II expression is necessary for the development of S. aureus arthritis in mice and that different MHC class II haplotypes confer varying susceptibility for development of joint inflammation induced by staphylococci.

Animals↗

In situ hybridization analysis of synovial and systemic cytokine messenger RNA expression in superantigen-mediated Staphylococcus aureus arthritis.

OBJECTIVE: To investigate patterns of synovial and systemic cytokine messenger RNA (mRNA) expression in mice with superantigen-mediated Staphylococcus aureus arthritis. METHODS: Mice were inoculated intravenously with 1 x 10(7) colony-forming units of toxic shock syndrome toxin-1-producing S aureus LS-1. Synovial tissues and spleens were obtained at varying time intervals after bacterial inoculation, and examined for mRNA expression of interleukin-1beta (IL-1beta), IL-4, IL-10, IL-12, tumor necrosis factor alpha (TNFalpha), TNFbeta, interferon-gamma (IFN-gamma), transforming growth factor beta, and perforin, by an in situ hybridization technique. RESULTS: In situ hybridization revealed early synovial up-regulation of TNFalpha and IL-1 beta mRNA expression. Peak frequencies of these proinflammatory cytokines were observed at the second and third week of the infection. Expression of T cell-derived cytokine mRNAs was detected later, and in a relatively low frequency. Notably, induction and peak numbers of Th2 cytokine (IL-4 and IL-10) mRNA expression preceded Th1 cytokine (IFNgamma and TNFbeta) mRNAs. In comparison with synovial tissues, peak spleen cytokine mRNA expression of IL-1beta, TNFalpha, TNFbeta, IL-12, and IFNgamma occurred earlier, but displayed a clearly lower magnitude of expression. CONCLUSION: These findings demonstrate synovial and systemic up-regulation of cytokine mRNA expression during S aureus arthritis, indicating that both monocyte/macrophage and T cell-derived products are involved in the pathogenesis of this disease.

Animals↗

Impact of interferon-gamma receptor deficiency on experimental Staphylococcus aureus septicemia and arthritis.

The role of IFN-gamma in the regulation of host resistance of Staphylococcus aureus was studied using IFN-gamma receptor-deficient (IFN-gamma R-/-) mice in a model of S. aureus-induced septicemia and arthritis. IFN-gamma R-/- mice and wild-type controls were inoculated intravenously with a toxic shock syndrome toxin-1-producing S. aureus LS-1 strain. IFN-gamma R-/- mice displayed significantly more frequent and more severe arthritis compared with wild-type littermates (p < 0.01) throughout the course of infection. Notably, IFN-gamma R-/- mice developed severe sepsis with high mortality early after the inoculation with staphylococci. However, the mortality of wild-type mice became significantly higher at later stages of the disease compared with IFN-gamma R-/- mice (p < 0.05). This differential outcome of sepsis-related mortality was associated with deficiencies of bacterial elimination from blood and parenchymatous organs and correlated well to serum levels of IL-6 and spleen IL-1 beta and TNF-beta mRNA expression. Thus, bacterial growth and proinflammatory cytokines IL-1 beta, TNF-beta, and IL-6 were higher at the early stage of infection in IFN-gamma-/- mice but increased at the later stage in wild-type littermates. Our data indicate that the absence of IFN-gamma R leads to harmful as well as beneficial effects in S. aureus infection, depending on the stage of the disease and the localization of the infection.

Animals↗