Complete 1H, 15N, and 13C assignments of an exchangeable apolipoprotein, Locusta migratoria apolipophorin III.
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Animals↗
Biomedical subjects
D Fan
Publications and source records attributed to D Fan.
Reversion of the malignant phenotype of gastric cancer cell SGC7901 by c-erbB-2-specific hammerhead ribozyme.
The c-erbB-2/neu-encoded protein p185 is closely related to the growth and metastasis of adenocarcinoma. We sought to reverse the malignant phenotype of gastric cancer cell line SGC7901 with c-erbB-2-specific ribozyme. We designed the ribozyme and generated the in vitro transcription vectors of the ribozyme and its substrate. In vitro cleavage reaction indicated that the ribozyme catalyzed 79.3% target RNA in 1 hour at 37 degrees C. Then, we generated the eucaryotic expression vectors of the ribozyme and transfected them into SGC7901 cells, which highly express p185. Analyses showed that the c-erbB-2 mRNA and p185 were reduced remarkably in the ribozyme-transfected cells. The growth rate of the ribozyme-transfected cells was much lower than that of the control group. Tumorigenicity was also decreased dramatically in nude mice. The results demonstrated that c-erbB-2-specific ribozyme may inhibit the malignancy of gastric cancer cells SGC7901.
Adenocarcinoma↗
Increasing membrane-bound MCSF does not enhance OPGL-driven osteoclastogenesis from marrow cells.
Macrophage colony-stimulating factor (MCSF) and osteoprotegerin ligand (OPGL), both produced by osteoblasts/stromal cells, are essential factors for osteoclastogenesis. Whether local MCSF levels regulate the amount of osteoclast formation is unclear. Two culture systems, ST-2 and Chinese hamster ovary-membrane-bound MCSF (CHO-mMCSF)-Tet-OFF cells, were used to study the role of mMCSF in osteoclast formation. Cells from bone marrow (BMM) or spleen were cultured with soluble OPGL on glutaraldehyde-fixed cell layers; osteoclasts formed after 7 days. Osteoclast number was proportional to the amount of soluble OPGL added. In contrast, varying mMCSF levels in the ST-2 or CHO-mMCSF-Tet-OFF cell layers, respectively by variable plating or by addition of doxycycline, did not affect BMM osteoclastogenesis: 20-450 U of mMCSF per well generated similar osteoclast numbers. In contrast, spleen cells were resistant to mMCSF: osteoclastogenesis required > or = 250 U per well and further increased as mMCSF rose higher. Our results demonstrate that osteoclast formation in the local bone environment is dominated by OPGL. Increasing mMCSF above basal levels does not further enhance osteoclast formation from BMMs, indicating that mMCSF does not play a dominant regulatory role in the bone marrow.
Animals↗
A case of pyrexia, hepatosplenomegaly, cytopenia and hypergammaglobulinemia.
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Adult↗
[Significance of lung perfusion scanning with technetium labeled macroaggregated albumin and pulmonary function assay for diagnosis of early hepatopulmonary syndrome].
OBJECTIVE: To evaluate the values of pulmonary function assay and dynamic pulmonary perfusion imaging with technetium labeled macroaggregated albumin ((99m)TcMAA) in the early diagnosis of hepatopulmonary syndrome (HPS). METHODS: The pulmonary function assay and (99m)TcMAA scans were performed in 28 patients with HPS, 30 cirrhotic patients (CP) without HPS, and 21 healthy controls (HC). RESULTS: In the patients with HPS, PaO(2), SaO(2) and diffusion capacity for carbon monoxide of lungs (DLco) was significantly lower than that in CP (P<0.01) and HC (P<0.01), and alveolar-arterial gradient [P((A-a))O(2)] was significantly increased (P<0.001). Results from (99m)TcMAA scans showed that the radionuclides were distributed over the spleen, kidney, liver and brain, and the ratios of arterivenous shunt were significantly higher than that in CP (P<0.001) and NC (P<0.001). In cirrhotic patients, DLco significantly decreased (P<0.05), P((A-a))O(2) and shunt ratios increased (P<0.01 and 0.001). CONCLUSIONS: Pulmonary function assay and dynamic pulmonary perfusion imaging with (99m)TcMAA are sensitive methods for diagnosis of the early HPS.
Hepatopulmonary Syndrome↗
Adeno-associated virus vector-mediated triple gene transfer of dopamine synthetic enzymes.
OBJECTIVE: To explore triple gene transfer of dopamine synthetic enzymes with separate adeno-associated virus (AAV) vectors. METHODS: The genes for dopamine synthetic enzymes, tyrosine hydroxylase (TH), aromatic L-amino acid decarboxylase (AADC), and GTP cyclohydrolase I (GCH, an enzyme critical for tetrahydrobiopterin synthesis) were contransduced into 293 cells with separate AAV vectors. Expressions of TH, AADC and GCH were detected by Western blot analysis. Intracellular dopamine level was assayed by high-performance liquid chromatography. RESULTS: TH, AADC and GCH were effectively coexpressed in transduced cells with three separate AAV vectors, AAV-TH, AAV-AADC and AAV-GCH. Furthermore, the coexpression resulted in an effectively spontaneous dopamine production in cotransduced cells. CONCLUSION: The triple transduction of TH, AADC and GCH genes with separate AAV vectors is effective, which might be important to gene therapy for Parkinson's disease.
Animals↗
[The effect of calponin and caldesmon in regulation of the gastrointestinal motility during pathophysiological adaptation].
OBJECTIVE: To investigate the expression of calponin (CaP) and caldesmon (CaD) in the gastrointestinal tract and their effect in regulating gastrointestinal motility during physiological and pathological adaptation. METHODS: Models of chronic gastrointestinal motility hyperfunction in mice were induced by intragastric administration of senna extraction and models of chronic gastrointestinal motility hypofunction in rats were established with carbon tetrachloride induced cirrhosis, CaP and CaD were detected in the gastrointestinal tract of different model groups using SDS-PAGE and Western blot. The color development of Western blots was scanned using densitometric scanning. The relative contents of gastrointestinal CaP and CaD were compared with control animals with different state of gastrointestinal motility. RESULTS: Animal models of abnormal gastrointestinal motility were established in mice and rats. Densitometric quantification of CaP and CaD blots by CP1 and C98 mAbs showed that normal animal colon contained higher amounts of h1-CaP and CaD. In normal mice and rats, the content of CaP and CaD was successively in this order :colon > stomach > small intestine. The content was reduced in mice of chronic gastrointestinal motility hyperfunction, but the expression of CaP and CaD was promoted in rats with cirrhosis and declined to normal level after treatment with L-NAME. CONCLUSION: There is close relation between expression of CaP and CaD and state of gastrointestinal motility. CaP and CaD may inhibit gastrointestinal motility. These suggest that CaP and CaD may play a role in the regulation of gastrointestinal motility during physiological and pathological adaptation.
Animals↗
Blockade of the epidermal growth factor receptor signaling by a novel tyrosine kinase inhibitor leads to apoptosis of endothelial cells and therapy of human pancreatic carcinoma.
We determined whether down-regulation of the epidermal growth factor-receptor (EGF-R) signaling pathway by oral administration of a novel EGF-R tyrosine kinase inhibitor (PKI166) alone or in combination with gemcitabine (administered i.p.) can inhibit growth and metastasis of human pancreatic carcinoma cells implanted into the pancreas of nude mice. Therapy beginning 7 days after orthotopic injection of L3.6pl human pancreatic cancer cells reduced the volume of pancreatic tumors by 59% in mice treated with gemcitabine only, by 45% in those treated with PKI166 only, and by 85% in those given both drugs. The combination therapy also significantly inhibited lymph node and liver metastasis, which led to a significant increase in overall survival. EGF-R activation was significantly blocked by therapy with PKI166 and was associated with significant reduction in tumor cell production of VEGF and IL-8, which in turn correlated with a significant decrease in microvessel density and an increase in apoptotic endothelial cells. Collectively, our results demonstrate that oral administration of an EGF-R tyrosine kinase inhibitor decreased growth and metastasis of human pancreatic cancer growing orthotopically in nude mice and increased survival. The therapeutic effects were mediated in part by inhibition of tumor-induced angiogenesis attributable to a decrease in production of proangiogenic molecules by tumor cells and increased apoptosis of tumor-associated endothelial cells.
Administration, Oral↗
Transcriptional regulation of the expression of macrophage colony stimulating factor.
The regulatory regions for transcriptional control of the MCSF gene are unknown. We examined regulatory control in a 774-bp murine MCSF promoter transfected into MC3T3-E1 osteoblast-like and COS-7 cells. Deletion of upstream sequence from -635 increased basal activity of the promoter by at least four-fold, an increase that was maintained when PU.1, NFkappaB and Egr1/Sp1 consensus sequences were subsequently removed. Mutagenesis identified a suppressor element between -635 and -642 from the transcriptional start site and an oligonucleotide representing this sequence was retarded by nuclear cell protein. TNFalpha (1 ng/ml), PTH (5x10(-8) M), and IL-1alpha (100 pg/ml), which increased MCSF protein secretion, failed to enhance the transcriptional rate of the full-length promoter. TNFalpha was able to stimulate transcription of a heterologous reporter transfected into COS-7 containing multiple copies of the murine MCSF NFkappaB site inserted before a minimal promoter. In contrast, deletion of the same NFkappaB response element increased basal activity in the native promoter. Thus, the NFkappaB sequence may act as a negative regulator in the context of the endogenous promoter. Our results indicate that constitutive transcriptional activity conferred by the MCSF promoter may be damped by a suppressor protein. Transcriptional regulation, however, does not appear to be a major stimulatory mechanism for MCSF secretion.
3T3 Cells↗
Acid pH increases carbonic anhydrase II and calcitonin receptor mRNA expression in mature osteoclasts.
Numerous resorptive stimuli have been shown to enhance osteoclast differentiation, increasing osteoclast numbers and accelerating bone resorption. Currently, there is much less understanding of regulation of mature osteoclast activity. Indeed, there is presently only minimal evidence of changes in gene expression as a mechanism for altering bone resorption. We investigate here, in the mature osteoclast, regulation of 2 genes-carbonic anhydrase II (CAII) and calcitonin receptor (CTR) in response to acidosis, which is known to increase bone resorption. We studied the effect of acid pH on CAII and CTR mRNA expression in mature osteoclasts raised in coculture of ST-2 and primary marrow cells. On day 6 of culture, stromal cells were removed with collagenase, the remaining osteoclasts were incubated overnight, and then exposed to varying pH. RT-PCR was performed on total RNA using primers for CAII, CTR, or glyceraldehyde dehydrogenase phosphate (GAP). Expression of CTR mRNA was increased 2.14 +/- 0.41 and 2.56 +/- 0.45 (P < 0.05)-fold by a 4-hour exposure to pH 6.75 and 6.5, respectively. CAII mRNA was similarly increased 2.18 +/- 0.42 and 2.63 +/- 0.48 (P < 0.05)-fold by pH 6.75 and 6.5, respectively. Increased expression of CAII and CTR mRNA was seen by 2 hours and maximally by 4 hours. Increased expression of CTR and CAII mRNA was not explained by increases in osteoclast numbers: pH 7.4-100 +/- 3.7, 6.75-133 +/- 8.3, 6.5-124 +/- 7.8. These results demonstrate upregulation of two osteoclast genes in response to acidosis, illustrating the ability of the mature osteoclast to respond to resorptive signals with increased functional gene expression.
Animals↗
Moiré technique for spatial coherence measurements of soft-x-ray lasers.
We present a novel moiré diagnostic that allows the time-averaged spatial coherence properties of soft-x-ray lasers to be measured. The technique is an extension of the theoretical work in a recent paper [J. Opt. Soc. Am. A 16, 323 (1999)] and is based on the concept of decompositions of the far-field divergence for partially coherent beams. An appealing feature of the approach is that the spatial coherence width at other planes can be evaluated from the measured one at a given plane on the assumption of shape-invariant beams.
Computer Simulation↗
Site-specific expression of transferrin receptor by human colon cancer cells directly correlates with eradication by antitransferrin recombinant immunotoxin.
We determined the efficacy of HB21(Fv)PE40, a single-chain immunotoxin made by fusing the variable regions of a monoclonal antibody directed at the human transferrin receptor (TfR) with a truncated mutant of Pseudomonas exotoxin (PE), against metastatic human colon carcinoma KM12L4 cells growing in the liver or subcutis of nude mice. Organ-specific modulation of TfR expression was examined by immunohistochemistry and flow cytometry using anti-human CD71 antibody. KM12L4 cells expressed human TfR and were lysed in vitro by HB21(Fv)PE40 but not LMB-7 (a control immunotoxin specific for a Lewis Y-related carbohydrate antigen). KM12L4 cells growing in the liver expressed higher levels of TfR than cells growing s.c. Systemic administration of HB21(Fv)PE40 eliminated KM12L4 liver metastasis, whereas administration of LMB-7 did not. Treatment of mice with HB21(Fv)PE40 only delayed the growth of s.c. tumors. KM12L4 cells recovered from liver metastases, expressed higher levels of TfR, and were more sensitive to lysis by HB21(Fv)PE40 than KM12L4 cells recovered from s.c. tumors. Indeed, collectively, the data show that the expression level of the TfR by human colon cancer cells is modulated by the organ microenvironment which can be advantageous for the use of therapeutic immunotoxins.
Animals↗
[Application of studies on popular words in the systematization of ancient medical books].
Based on the arrangement of Renho Temple edition of Taisu (Extremely Plain), as an example, it can be seen that by correcting the wrong popular words in ancient medical books, the causes of mistakes of ancient editions and solution for annotations of difficult terms of ancient words in the medical books are of significance.
Books↗
Upregulation of drug sensitivity of multidrug-resistant SGC7901/VCR human gastric cancer cells by bax gene transduction.
OBJECTIVE: To investigate the role of bax in a vincristine (VCR)-induced multidrug-resistant (MDR) human gastric cancer cell line, SGC7901/VCR, in which the Bax protein expression level was significantly lower compared with that in parent cells. METHODS: A bax eukaryotic expression vector was constructed and transfected into SGC7901/VCR cells by lipofectamine, and resistant clones were selected by G418. Western blotting detected Bax expression in transfectants. Tetrazolium blue (MTT) assay evaluated the differences in drug sensitivity and cell cycle changes of transfectants were analyzed using flowcytometry (FCM). RESULTS: The bax eukaryotic expression vector was constructed and transfected into SGC7901/VCR cells. Through G418 selection, resistant clones were obtained. Western blotting demonstrated that the expression of Bax protein was markedly increased in bax transduced cells. These cells were more sensitive to adriamycin (ADR) and VCR than mock vector transducted cells. Moreover, bax transfection enhanced ADR-induced apoptosis and VCR-induced G2/M phase arrest of SGC7901/VCR cells. CONCLUSION: Bax was involved in the MDR of SGC7901/VCR cells.
Antineoplastic Agents, Phytogenic↗
[Production of phage-displayed single chain variable fragments of monoclonal antibody MGb1].
OBJECTIVE: To lay a foundation for obtaining a tumor-targeting vehicle for in vivo study on diagnosis and treatment of gastric carcinoma by generating single chain variable fragments (ScFv) of monoclonal antibody MGb1 directed against the cancer. METHODS: mRNA was isolated from MGb1-producing mouse hybridoma cell line, and the variable regions of heavy and light chain cDNAs were amplified separately and assembled into ScFv DNAs with a specially constructed linker DNA by PCR. The ScFv DNAs were ligated into the phagemid vector pCANTAB5E and the ligated sample was transformed into competent E. coli TG1. The transformed cells were infected with M13KO7 helper phage to yield recombinant phage, which display ScFv fragments as a fusion with gene 3 protein on the tips of the phage M13. After two rounds of panning with gastric carcinoma cell line KATO III highly expressing MGb1-binding antigen, the phage clones displayed ScFv fragments of the antibody were selected by enzyme-linked immunosorbent assay (ELISA) from the enriched phages. The affinity of the positive phage clones was detected by competition ELISA. RESULTS: The VH, VL and ScFv DNAs were about 340 bp, 320 bp and 750 bp respectively. 17 phage clones displayed ScFv of MGb1 were selected from 40 enriched phage clones. 4 out of the 17 phage clones could strongly compete with the original hybridoma antibody MGb1 for binding to the antigen expressed on KATOIII cells. CONCLUSION: The phage-displayed ScFv fragments of monoclonal antibody MGb1 are successfully produced by phage antibody technology, which may be useful to widen the range of application of the antibody.
Animals↗
[Effects of antisense human telomerase RNA on growth of human gastric cancer cells].
OBJECTIVE: To observe the inhibitory effects of antisense human telomerase RNA (hTR) on growth of human gastric cancer cells. METHODS: SGC7901 cell line was transfected with antisense hTR expression vector (pBBS-hTR) by lipofectAMINE. The gene transfected cells were cultured in vitro and then inoculated into nude mice. RESULTS: Antisense hTR expression was enhanced and sense hTR expression inhibited in clones containing antisense hTR after hygromycin resistant selection which induced tumor cells apoptosis. The growth rate of the major gene transfected cells in culture and tumorigenesis of the transfected cells in nude mice were significantly reduced in comparison with those of controls. The survival time of nude mice inoculated with the gene transfected cells was markedly prolonged when compared to controls. CONCLUSIONS: The malignant proliferation of gastric cancer cell can be reversed by antisense hTR induced cell apoptosis. Thus antisense hTR may serve as a potential therapeutic approach to the treatment of human cancer.
Animals↗
AMPA receptor protein expression and function in astrocytes cultured from hippocampus.
Glutamate receptors guide the proliferation, migration, and differentiation of glial cells. Here, we characterize AMPA (alpha-amino-3-hydroxy-5-methyl-4-isoxazole-propionic acid) and NMDA receptor protein expression and function and mRNA expression in hippocampal glial cultures. By immunocytochemistry, GluR2 (the subunit that limits the Ca(2+) permeability of AMPA receptors) exhibited prominent labeling in hippocampal glial cultures. Double-labeling of GluR2 with GFAP and with A2B5 revealed GluR2 subunit expression on type-1 and type-2 astrocyte lineage cells. GluR1 subunit expression was more prominent in type-1 than in type-2 astrocytes. To characterize functional properties of glutamate receptors expressed in cultured hippocampal astrocytes, we performed whole-cell patch clamp recording. Application of L-glutamate, AMPA, and kainate, but not NMDA, to small, rounded cells (morphologically identified as type-2 astrocytes) elicited inward currents which were blocked by the AMPA/kainate antagonist 6-cyano-7-nitroquinoxaline-2, 3-dione (CNQX). Cyclothiazide potentiated AMPA- and kainate-elicited currents, indicative of AMPA-preferring receptors. Current voltage analysis indicated that type-2 astrocyte AMPA receptors were electrically linear, indicative of GluR2-containing, Ca(2+)-impermeable AMPA receptors. By Northern blot analysis, GluR1 mRNA was highest in astrocyte cultures from cerebellum and hippocampus and moderate in astrocyte cultures from neocortex and striatum. GluR3 mRNA was detectable in astrocyte cultures from cerebellum and neocortex. GluR2 and NR1 mRNA expression were not detected in astrocytes cultured from any brain region examined. In situ hybridization studies showed wide expression of GluR1 mRNA in cultured astrocytes; GluR2 and GluR3 mRNAs were near background levels. Thus, cultured type-2 astrocytes express functional AMPA receptors in a cell-specific and region-specific manner, consistent with their role in neuronal-glial communication.
Animals↗
Peptide antagonists of the human estrogen receptor.
Estrogen receptor alpha transcriptional activity is regulated by distinct conformational states that are the result of ligand binding. Phage display was used to identify peptides that interact specifically with either estradiol- or tamoxifen-activated estrogen receptor alpha. When these peptides were coexpressed with estrogen receptor alpha in cells, they functioned as ligand-specific antagonists, indicating that estradiol-agonist and tamoxifen-partial agonist activities do not occur by the same mechanism. The ability to regulate estrogen receptor alpha transcriptional activity by targeting sites outside of the ligand-binding pocket has implications for the development of estrogen receptor alpha antagonists for the treatment of tamoxifen-refractory breast cancers.
Amino Acid Sequence↗