Biomedical subjects

X Zhao

Publications and source records attributed to X Zhao.

At least 289 records · Page 16Linked to original sources

PubMed1999

[Relationship between homocysteine-induced apoptosis and teratogenesis in developing avian embryo].

It has been hypothesized that homocysteine(HCY) can induce apoptosis at the organogenesis-stage of chick embryos and which can be the relationship between HCY and congenital defects of neural tube and heart. Exogenous D, L-HCY (4-8 mumol/embryo) was injected into the place near brain and the yolk of the day 4 embryos. Then they were harvested 2 days later and examined by flow cytometry, histopathology, electronic microscope, in situ end labeling of DNA strand break and methylgreen-pyronine double staining. The change of plasma HCY level of day 10 embryos was also inspected by HPLC analysis. The HCY injected at different position of embryos could result in heart and neural tube defects, which included ectopia cardis, hydropericardium, endocardial cushion defect, encephalocele, microcephaly, abnormal flexions and so on. HCY could apparently disturb embryonic cell cycle, suppress the synthesis of DNA and RNA of embryonic cells. Excessive apoptotic cells in brain and heart were observed in HCY\|treatment groups. It was discovered that mitochondrial damage could be described as an early event in cellular apoptosis of chick embryos. After 8 mumol of HCY was placed on the inner shell membrance, there was an obvious increase of plasma HCY concentration with a peak value of 80.58 mumol/L in 120 min, and followed by declining. On the other hand, 5 micrograms of folic acid given simultaneously with HCY could sharply curtail the rise in plasma HCY. It is concluded that HCY is a cytotoxin, inducing over apoptosis of embryos in some areas, which is coincident with the pathologic changes of neural system and heart. These findings show that there is a close relationship among HCY, apoptosis and birth defect in developing chick embryos. Mitochondrial dysmorphosis may be an important early event of apoptosis in embryos after exposure to HCY.

Animals↗

PubMed1999

Characterization of NFkappaB activation by detection of green fluorescent protein-tagged IkappaB degradation in living cells.

Activation of the transcription factor NFkappaB requires rapid degradation of its inhibitor, IkappaBalpha. To facilitate the study of IkappaBalpha degradation, we fused IkappaBalpha protein to enhanced green fluorescent protein to construct IkappaBalpha-enhanced green fluorescent protein (IG). We demonstrated by both flow cytometry and Western blot analysis that the half-life of IG in the presence of human tumor necrosis factor (TNF) alpha is approximately 5 min, which is similar to the half-life of native IkappaBalpha. The degradation coincided with NFkappaB translocation from the cytoplasm to the nucleus and NFkappaB-mediated induction of transcription. Phorbol 12-myristate 13-acetate (PMA), but not forskolin, also induces degradation of IG fusion protein. The half-life of IG in the presence of PMA is approximately 15 min, longer than when induced with TNFalpha. Co-treatment with TNFalpha and PMA did not result in a synergistic effect on IG degradation, although they stimulate different kinases in two different signaling pathways. Degradation of IG was inhibited by mutations at serine residues 32 and 36, which are the target sites of the phosphorylation modification that initiates degradation of IkappaBalpha. We also demonstrated that basal degradation of IG in the presence of cycloheximide is inhibited by such mutations, suggesting that basal degradation of IkappaBalpha also requires phosphorylation as the signal for degradation. Finally, we showed that the rate of TNFalpha-induced degradation of IG remains almost constant throughout the cell cycle, except at the mitotic phase, in which IG degrades more slowly.

DNA-Binding Proteins↗

PubMed1999

A novel form of rhodopsin kinase from chicken retina and pineal gland.

The G protein-coupled receptor kinases (GRKs) are important enzymes in the desensitization of activated G protein-coupled receptors (GPCR). Seven members of the GRK family have been identified to date. Among these enzymes, GRK1 is involved in phototransduction and is the most specialized kinase of the family. GRK1 phosphorylates photoactivated rhodopsin (Rho*), initiating steps in its deactivation. In this study, we found that chicken retina and pineal gland express a novel form of GRK that has sequence features characteristic of GRK1. However, unlike bovine GRK1 which is farnesylated, chicken GRK1 contains a consensus sequence for geranylgeranylation. Peptides corresponding to the C-terminal sequence of chicken GRK1 are geranylgeranylated by a cytosolic extract of chicken liver. Based on results of molecular cloning and immunolocalization, it appears that both rod and cone photoreceptors express this novel GRK1. These data indicate a larger sequence diversity of photoreceptor GRKs than anticipated previously.

Amino Acid Sequence↗

PubMed1999

[Expression of human prp gene in prokaryotic cells using GST fusion protein expression system].

OBJECTIVE: To study the biological features of cell-surface protein PrPc, which is thought to be involved in the prion-associated diseases after converting to a proteinase-resistant isoform PrPSc posttranslationally, and to establish an effective immunologic diagnostic method using PrPc as antigen. METHODS: Amplifying and cloning the human prp gene from lymphocytes of two normal Chinese, after confirmed by DNA sequence analysis, the genes were separately subcloned into a GST-fusion expression plasmid. RESULTS: Sequence analysis showed that one contatined a point mutation that induced the 65th amino acid "Trp" inverting to a stop codon "TAG", whereas the other had the same sequence as the published standard prp gene. Both the standard and the mutated prp genes were separately subcloned into a GST fusion protein expression vector and expressed in the prokaryotic cells effectively. Western blot assay revealed that both of them expressed GST-PrP fusion proteins and could be recognized by PrP specific monoclonal antibody. CONCLUSION: It suggests that human PrP protein can be expressed in the GST fusion protein expression system and the expressed proteins hold good immune-reactivity.

Amino Acid Sequence↗

PubMed1999

[Inhibition of human breast cancer cell line BCap-37 by flavonoid extract of wheat germ in vitro].

Cell growth and proliferation were measured by microculture tetrazolium(MTT) assay, cell colony-forming assay and the synthesis of DNA by 3H-thymidine incorporation. Flavonoid extract of wheat germ resulted to a dose-dependent, time-dependent growth inhibition, reduction of colony and 3H-thymiding incorporation in DNA of human breast cancer cell BCap-37. These findings indicated that the flavonoid extract of wheat germ can inhibit tumor cell growth and proliferation by blocking DNA synthesis in vitro.

Antineoplastic Agents, Phytogenic↗

PubMed1999

[Free-radical theory and the antisenescence effect of fruits and vegetables].

Oxidative damage resulting from free radical is a major cause of ageing and chronic degenerative disease. Epidemiological studies show that frequent consumption of fruit and yellow or green leafy vegetables may prevent many kinds of chronic degenerative diseases associated with the increasing of age. These may be related with the anti-oxidative ingredients in fruit and vegetables. Nevertheless, trials using anti-oxidative nutrient, such as vitamin C, E and beta-carotene supplements, did not get the expected result so far. Thus, it is necessary to strengthen the research on the potential effective anti-oxidative ingredients in fruit and vegetables. On the other hand, the study on the effect of fruit and vegetables on life span and improving life quality are also needed.

Aging↗

PubMed1999

The origin of catecholaminergic nerve fibers in the subdiaphragmatic vagus nerve of rat.

It is known that the vagus nerve contains catecholaminergic fibers. However, the origin of these fibers has not been systematically examined. In this study, we addressed this issue using retrograde tracing from the subdiaphragmatic vagus nerve combined with immunocytochemistry. The cervical and thoracic sympathetic trunk ganglia, the nodose ganglia and the dorsal motor nucleus of the vagus nerve were examined following injection of Fluoro-Gold or cholera toxin horseradish peroxidase conjugate into the trunks of the subdiaphragmatic vagus nerve of rats. Numerous retrogradely labeled neurons were seen in the nodose ganglion and the dorsal motor nucleus of the vagus nerve. Very few labeled neurons were found in the sympathetic ganglia (less than 0.06% of the neurons in either superior cervical ganglion or cervicothoracic ganglion were retrogradely labeled). Double labeling with immunofluoresence for catecholamine synthesizing enzymes revealed that: (1) 92% of all Fluoro-Gold retrogradely labeled tyrosine hydroxylase immunoreactive neurons were found in parasympathetic sources (75% in the dorsal motor nucleus of the vagus nerve and 17% in the nodose ganglia), and only 8% in the cervicothoracic sympathetic ganglia; (2) 12% of the retrogradely labeled catecholaminergic neurons in the dorsal motor nucleus of the vagus nerve were also dopamine-beta-hydroxylase immunopositive neurons; (3) 70% of the retrogradely labeled neurons in the sympathetic ganglia were tyrosine hydroxylase immunopositive and 54% of these catecholaminergic neurons contained dopamine-beta-hydroxylase, while 30% of the retrogradely labeled neurons were non-catecholaminergic neurons. These results indicate that catecholaminergic fibers in the abdominal vagus nerve are primarily dopaminergic and of parasympathetic origin, and that only an extremely small number of these fibers, mostly noradrenergic in nature, arise from postganglionic sympathetic neurons.

Animals↗

PubMed1999

Involvement of conserved, acidic residues in the N-terminal domain of troponin C in calcium-dependent regulation.

We have mutated eight conserved, charged amino acid residues in the N-terminal, regulatory domain of troponin C (TnC) so we could investigate their role in troponin-linked Ca2+ regulation of muscle contraction. These residues surround a hydrophobic pocket in the N-terminal domain of TnC which, when Ca2+ binds to regulatory sites in this domain, is exposed and interacts with the inhibitory region of troponin I (TnI). We constructed three double mutants (E53A/E54A, E60A/E61A, and E85A/D86A) and two single mutants (R44A and R81A) of rabbit fast skeletal muscle troponin C (TnC) in which the charged residues were replaced with neutral alanines. All five of these mutants retained TnC's ability to bind TnI in a Ca2+-dependent manner, to neutralize TnI's inhibition of actomyosin S1 ATPase activity, and to form a ternary complex with TnI and troponin T (TnT). Ternary complexes formed with TnC(R44A) or TnC(R81A) regulated actomyosin S1 ATPase activity normally, with TnI-based inhibition in the absence of Ca2+ and TnT-based activation in the presence of Ca2+. TnC(E53A/E54A) and TnC(E85A/D86A) interacted weakly with TnT, as judged by native gel electrophoresis. Ternary complexes formed with these mutants inhibited actomyosin S1 ATPase activity in both the presence and absence of Ca2+, and did not undergo Ca2+-dependent structural changes in TnI which can be detected by limited chymotryptic digestion. TnC(E60A/E61A) interacted normally with TnT. Its ternary complex showed Ca2+-dependent structural changes in TnI, inhibited actomyosin S1 ATPase in the absence of Ca2+, but did not activate ATPase in the presence of Ca2+. This is the first demonstration that selective mutation of TnC can abolish the activating effect of troponin while its inhibitory function is retained. Our results suggest the existence of an elaborate network of protein-protein interactions formed by TnI, TnT, and the N-terminal domain of TnC, all of which are important in the Ca2+-dependent regulation of muscle contraction.

Actomyosin↗

PubMed1999

Zonula occludens-1 and E-cadherin are coordinately expressed in the mouse uterus with the initiation of implantation and decidualization.

Two-way interactions between the blastocyst trophectoderm and the uterine luminal epithelium are essential for implantation. The key events of this process are cell-cell contact of trophectoderm cells with uterine luminal epithelial cells, controlled invasion of trophoblast cells through the luminal epithelium and the basement membrane, transformation of uterine stromal cells surrounding the blastocyst into decidual cells, and protection of the "semiallogenic" embryo from the mother's immunological responses. Because cell-cell contact between the trophectoderm epithelium and the luminal epithelium is essential for implantation, we investigated the expression of zonula occludens-1 (ZO-1) and E-cadherin, two molecules associated with epithelial cell junctions, in the mouse uterus during the periimplantation period. Preimplantation uterine epithelial cells express both ZO-1 and E-cadherin. With the initiation and progression of implantation, ZO-1 and E-cadherin are expressed in stromal cells of the primary decidual zone (PDZ). As trophoblast invasion progresses, these two molecules are expressed in stroma in advance of the invading trophoblast cells. These results suggest that expression of these adherence and tight junctions molecules in the PDZ serves to function as a permeability barrier to regulate access of immunologically competent maternal cells and/or molecules to the embryo and provide homotypic guidance of trophoblast cells in the endometrium.

Animals↗

PubMed1999

Repression of human fibroblast growth factor 2 by a novel transcription factor.

Here we describe the cloning of the regulator of fibroblast growth factor 2 (FGF-2) transcription (RFT) using a yeast one-hybrid screening with a defined motif in FGF-2 promoter as a target sequence. Overexpression of human RFT (RFT-A) reduces FGF-2 RNA and protein levels in both normal and tumor cell lines. Its splice variants, RFT-A' and RFT-B, have deletions in the putative DNA binding domain and fail to bind FGF-2 promoter and repress FGF-2 gene expression. The ratios of RFT isoforms differ between normal and tumor cells, with the splice variants dominating in tumor cells. Overexpression of RFT-A induces glioma cell death. Our data suggest that regulation of FGF-2 by RFT is important for cellular functions and may be impaired in certain tumors.

Alternative Splicing↗

PubMed1999

E-cadherin mediates aggregation-dependent survival of prostate and mammary epithelial cells through the retinoblastoma cell cycle control pathway.

E-cadherin and the retinoblastoma tumor suppressor (Rb) are traditionally associated with diverse regulatory aspects of cell growth and differentiation. However, we have discovered new evidence, which suggests that these proteins are functionally linked in a physiologic pathway required for cell survival and programmed cell death. Pharmacological activation of protein kinase C (PKC) or inducible overexpression and activation of the alpha isozyme of PKC (PKCalpha) resulted in approximately 60% apoptosis of mammary and prostate epithelial cells. Interestingly, the surviving cells had undergone dramatic aggregation concurrent with increased E-cadherin expression. When aggregation was inhibited by the addition of an E-cadherin-blocking antibody, apoptosis increased synergistically. We hypothesized that survival of the aggregated population was associated with contact-inhibited growth and that apoptosis might result from aberrant growth regulatory signals in non-aggregated, cycling cells. This hypothesis was confirmed by experiments that demonstrated that E-cadherin-dependent aggregation resulted in Rb-mediated G1 arrest and survival. Immunoblot analysis and flow cytometry revealed that hypophosphorylated Rb was present in non-aggregated, S phase cultures concurrent with synergistic cell death. We have also determined that the loss of membrane E-cadherin and subsequent hypophosphorylation of Rb in luminal epithelial cells preceded apoptosis induced by castration. These findings provide compelling evidence that suggests that E-cadherin-mediated aggregation results in Rb activation and G1 arrest that is critical for survival of prostate and mammary epithelial cells. These data also indicate that Rb can initiate a fatal growth signal conflict in non-aggregated, cycling cells when the protein is hypophosphorylated as these epithelial cells enter S phase.

Apoptosis↗

PubMed1999

[The antioxidant activity of wild jujubi, crataegus and grape in vitro].

Two oxidative damaging systems, Fe(2+)-Cys and tert-butyl hydroperoxide, were used to induce the production of malondialdehyde (MDA) in rat liver microsome and erythrocytes. The inhibition rate of wild jujubi, crataegus and grape juice at the concentration of 2.5-3.3 mg/ml to Fe(2+)-Cys system on the production of MDA were 46.2%, 98.3% and 99.1% respectively. The inhibition rate of jujubi (8.3 mg/ml), crataegus (13.9 mg/ml) and grape (55.6 mg/ml) to tert-butyl hydroperoxide were 38.7%, 38.7% and 58.5% respectively. The capability of scavenging O2.- and .OH was measured by ESR technique. It was found that the capability of these juices to eliminate O2.- and .OH generated by the xanthine/xanthine oxidase system and H2O2-FeSO4 system was strong. The elimination rate of jujubi (50 mg/ml), crataegus (50 mg/ml) and grape (134 mg/ml) to O2.- were 84.0%, 85.8% and 74.7% respectively and the rate to .OH were 96.7%, 97.8% and 86.3% respectively. The results indicated that these three fruits bear antioxidant activity.

Animals↗

PubMed1999

Ca2+-dependent interaction of the inhibitory region of troponin I with acidic residues in the N-terminal domain of troponin C.

Ca2+ regulation of vertebrate striated muscle contraction is initiated by conformational changes in the N-terminal, regulatory domain of the Ca2+-binding protein troponin C (TnC), altering the interaction of TnC with the other subunits of troponin complex, TnI and TnT. We have investigated the role of acidic amino acid residues in the N-terminal, regulatory domain of TnC in binding to the inhibitory region (residues 96-116) of TnI. We constructed three double mutants of TnC (E53A/E54A, E60A/E61A and E85A/D86A), in which pairs of acidic amino acid residues were replaced by neutral alanines, and measured their affinities for synthetic inhibitory peptides. These peptides had the same amino acid sequence as TnI segments 95-116, 95-119 or 95-124, except that the natural Phe-100 of TnI was replaced by a tryptophan residue. Significant Ca2+-dependent increases in the affinities of the two longer peptides, but not the shortest one, to TnC could be detected by changes in Trp fluorescence. In the presence of Ca2+, all the mutant TnCs showed about the same affinity as wild-type TnC for the inhibitory peptides. In the presence of Mg2+ and EGTA, the N-terminal, regulatory Ca2+-binding sites of TnC are unoccupied. Under these conditions, the affinity of TnC(E85A/D86A) for inhibitory peptides was about half that of wild-type TnC, while the other two mutants had about the same affinity. These results imply a Ca2+-dependent change in the interaction of TnC Glu-85 and/or Asp-86 with residues (117-124) on the C-terminal side of the inhibitory region of TnI. Since Glu-85 and/or Asp-86 of TnC have also been demonstrated to be involved in Ca2+-dependent regulation through interaction with TnT, this region of TnC must be critical for troponin function.

Amino Acid Sequence↗

PubMed1999

Secretion of 92 kDa gelatinase (MMP-9) by bovine neutrophils.

To understand the process of neutrophil migration into mammary glands during mastitis, secretion of MMP-9 by bovine neutrophils was investigated. The methods of zymograms and RT-PCR were used to measure the secretory MMP-9 activity and its gene transcription, respectively. Both MMP-9 protein and mRNA were found to be expressed constitutively and greatly enhanced by PMA. However, the significant increase in the enzyme activity was found after 1 ng/ml PMA treatment for 1 h, while increasing MMP-9 mRNA transcription was only obvious after 1 ng/ml PMA treatment for 3 h. Moreover, secretion of MMP-9 protein was not inhibited by actinomycin D and cycloheximide. These results suggest that quick MMP-9 protein release from bovine neutrophils by the stimulation of PMA was due to the degranulation. As MMP-9 is an important proteinase in breakdown of the extracellular matrix and can be rapidly secreted by neutrophils through degranulation, it should play a critical role in the recruitment of neutrophils into mammary glands in bovine mastitis.

Animals↗

PubMed1999

Coupling of the cell cycle and myogenesis through the cyclin D1-dependent interaction of MyoD with cdk4.

Proliferating myoblasts express the muscle determination factor, MyoD, throughout the cell cycle in the absence of differentiation. Here we show that a mitogen-sensitive mechanism, involving the direct interaction between MyoD and cdk4, restricts myoblast differentiation to cells that have entered into the G0 phase of the cell cycle under mitogen withdrawal. Interaction between MyoD and cdk4 disrupts MyoD DNA-binding, muscle-specific gene activation and myogenic conversion of 10T1/2 cells independently of cyclin D1 and the CAK activation of cdk4. Forced induction of cyclin D1 in myotubes results in the cytoplasmic to nuclear translocation of cdk4. The specific MyoD-cdk4 interaction in dividing myoblasts, coupled with the cyclin D1-dependent nuclear targeting of cdk4, suggests a mitogen-sensitive mechanism whereby cyclin D1 can regulate MyoD function and the onset of myogenesis by controlling the cellular location of cdk4 rather than the phosphorylation status of MyoD.

Animals↗

PubMed1999

[Study on the chemical components of edible oil fume in kitchen and its genotoxity on Drosophila].

OBJECTIVES: To study the chemical components of the condensate of edible oil fume in kitchen and its genotoxicity on Drosophila. METHODS: Analysis for the chemical components was carried out by gas chromatography and mass spectra (GC/MS) and its genotoxicity was studied by sex linked recessive lethal (SLRL) test in Drosophila. RESULTS: A total of 74 organic compounds were found in samples of condensed oil from the fume in kitchen. It included hydroxylic acids, hydrocarbons, alcohols, esters, aldehydes, ketones, aromatic compounds, and steroids, etc. The total mutagenicity rates in SLRL test induced by the samples at concentrations of 110,320 and 960 mg/L were 0.1732%, 0.4306% and 0.1707% respectively. The sterility rates of the first broods were 2.564%, 2.056% and 2.845% at above 3 concentrations respectively(P < 0.05, as compared with the control). The mutagenicity rate of the second brood at 320 mg/L was 0.530% and that of the third brood at 110 mg/L 0.540%(P < 0.001). CONCLUSION: Some of the compounds in the condensate of edible oil fume were proved to have high recessive lethal effect and genotoxic effect on the reproductive system of Drosophila.

Air Pollutants↗

PubMed1999

[Effects of cooking oil fume condensate on cellular immunity and immunosurveillance in mice].

Rapeseed oil fume, soyabean oil fume and salad oil fume condensate were injected respectively to female Kunming mice. The results showed that, compared to control group, in mice exposed to low level of rapeseed oil fume condensate, the delayed hypersensitivity responses were inhibited, while in mice exposed to high level of that, the delayed hypersensitivity responses and the NK cells activity were inhibited. In mice exposed to high level of soyabean oil fume condensate the delayed hypersensitivity responses, the NK cells activity and CaM activity were inhibited. The results of these parameters were all statistically significantly different from those in control mice. In mice exposed to low level of salad oil fume condensation, the delayed hypersensitivity responses and the NK cells activity were significantly inhibited. The results indicated that cooking oil fume could affect immune system of animals.

Air Pollutants↗

PubMed1999

Mannosylerythritol lipid is a potent inducer of apoptosis and differentiation of mouse melanoma cells in culture.

Malignant melanomas are tumors that are well known to respond poorly to treatment with chemotherapeutic reagents. We report here that mannosylerythritol lipid (MEL), an extracellular glycolipid from yeast, markedly inhibited the growth of mouse melanoma B16 cells in a dose-dependent manner. Exposure of B16 cells to MEL at 10 microM and higher concentrations caused the condensation of chromatin, DNA fragmentation, and sub-G1 arrest, all of which are hallmarks of cells that are undergoing apoptosis. Analysis of the cell cycle also suggested that both the MEL-mediated inhibition of growth and apoptosis were closely associated with growth arrest in the G1 phase. Moreover, MEL exposure stimulated the expression of differentiation markers of melanoma cells, such as tyrosinase activity and the enhanced production of melanin, which is an indication that MEL triggered both apoptotic and cell differentiation programs. Forced expression of Bcl-2 protein in stably transformed B16 cells had a dual effect: it interfered with MEL-induced apoptosis but increased both tyrosinase activity and the production of melanin as compared with these phenomena in vector-transfected MEL-treated control B16 cells. These results provide the first evidence that growth arrest, apoptosis, and the differentiation of mouse malignant melanoma cells can be induced by a microbial extracellular glycolipid.

Animals↗