Biomedical subjects

Xin Wu

Publications and source records attributed to Xin Wu.

At least 37 records · Page 2Linked to original sources

PubMed2004

Stepwise metastatic human hepatocellular carcinoma cell model system with multiple metastatic potentials established through consecutive in vivo selection and studies on metastatic characteristics.

PURPOSE: To establish a "stepwise metastatic human hepatocellular carcinoma (HCC) cell model system" for in-depth study of the underlying mechanisms of HCC metastasis. METHODS: Using MHCC97- a metastatic human hepatocellular carcinoma (HCC) cell line reported in 1999-as the parent cells, we subsequently established three cell lines (MHCC97-L, HMCC97-H, and HCCLM3) with increasing spontaneous metastatic potential. Now, the fourth cell line with unique multiple metastatic characteristics has been established by six rounds of in vivo selection. RESULTS: This cell line, designated as HCCLM6, is a polygonal epithelial cell with hypotriploid karyotype, the modal chromosomes are 55-58, and marker chromosomal abnormalities include i(1) (q10), i(8)(q10), der (4) t(4;8)(q31;q22), i(X)(q10). The cell population doubling time was 32 h. Fluorescent PCR showed HBV DNA integration in the cellular genome. Thirty-five days after HCCLM6 was injected subcutaneously into BALB/c nude mice, prominent lung metastases occurred in 100% of the recipient animals. When tumor tissue was orthotopically implanted into the liver of nude mouse, widespread loco-regional and pulmonary metastases occurred. Inoculation of this cell into the footpad of nude mice also produced 75% regional lymph node metastasis. Compared with MHCC97-L which was not metastastatic via subcutaneous or footpad inoculation and 40% metastatic via orthotopic inoculation, HCCLM6 had increased expression of matrix metalloproteinase (MMP-2 and MMP-9) and cytokeratin 19 (CK19), and decreased expression of Rb2/p130. The establishment of this new cell line has completed our stepwise metastatic HCC cell mode system, which was characterized by a similar genetic background but with significant differences in spontaneous metastasis behavior. CONCLUSIONS: The study supports the theory that cancer metastasis is a highly selective dynamic process and the cell model system could be a useful platform for the study of HCC metastasis.

Animals↗

PubMed2004

Identification of alternatively spliced mRNA variants related to cancers by genome-wide ESTs alignment.

Several databases have been published to predict alternative splicing of mRNAs by analysing the exon linkage relationship by alignment of expressed sequence tags (ESTs) to the genome sequence; however, little effort has been made to investigate the relationship between cancers and alternative splicing. We developed a program, Alternative Splicing Assembler (ASA), to look for splicing variants of human gene transcripts by genome-wide ESTs alignment. Using ASA, we constructed the biosino alternative splicing database (BASD), which predicted splicing variants for reference sequences from the reference sequence database (RefSeq) and presented them in both graph and text formats. EST clusters that differ from the reference sequences in at least one splicing site were counted as splicing variants. Of 4322 genes screened, 3490 (81%) were observed with at least one alternative splicing variants. To discover the variants associated with cancers, tissue sources of EST sequences were extracted from the UniLib database and ESTs from the same tissue type were counted. These were regarded as the indicators for gene expression level. Using Fisher's exact test, alternative splicing variants, of which EST counts were significantly different between cancer tissues and their counterpart normal tissues, were identified. It was predicted that 2149 variants, or 383 variants after Bonferroni correction, of 26 812 variants were likely tumor-associated. By reverse transcription-PCR, 11 of 13 novel alternative splicing variants and eight of nine variants' tissue specificity were confirmed in hepatocellular carcinoma and in lung cancer. The possible involvement of alternative splicing in cancer is discussed.

Alternative Splicing↗

PubMed2004

Antisense Tcf inhibits the neoplastic growth of liver cancer cells.

PURPOSE: T cell transcription factors are nuclear effectors of the Wnt signaling transduction pathway and play crucial roles in embryonic and malignant development. Our previous study showed increased expression level of Tcf mRNA in liver cancer. In the present paper, antisense Tcf RNA was used to explore the possible therapeutic effect on liver cancer cells by interrupting the abnormal Wnt pathway. METHODS: Antisense expression vectors containing the conserved sequence of Tcf cDNA were constructed and transfected into a human liver cancer cell line SMMC-7721. Tumorigenic potential was determined by cellular growth assay and tumor growth in nude mice. RESULTS: The stable transfection of anti-sense Tcf in SMMC-7721 cells significantly reduced Tcf expression at both mRNA and protein levels compared with parental and mock-transfected cells. Antisense-mediated suppression of Tcf inhibited the in vitro proliferation and in vivo tumor formation ability. Furthermore, the apoptosis rate of antisense transfected cells was significantly higher than that of control, indicating that antisense RNA suppressed malignant growth by induction of apoptosis. CONCLUSION: Our studies demonstrate the critical role of Wnt signaling pathway in the neoplastic growth of liver cancer cells and suggest that inhibition of Tcf activity with antisense Tcf RNA may be a potential new gene therapy method for liver cancer.

Cell Proliferation↗

PubMed2004

Cognitive impairment in PDAPP mice depends on ApoE and ACT-catalyzed amyloid formation.

Biochemical and genetic studies indicate that the inflammatory proteins, apolipoprotein E (ApoE) and alpha(1)-antichymotrypsin (ACT) are important in the pathogenesis of Alzheimer's disease (AD). Using several lines of multiply transgenic/knockout mice we show here that murine ApoE and human ACT separately and synergistically facilitate both diffuse A beta immunoreactive and fibrillar amyloid deposition and thus also promote cognitive impairment in aged PDAPP(V717F) mice. The degree of cognitive impairment is highly correlated with the ApoE- and ACT-dependent hippocampal amyloid burden, with PDAPP mice lacking ApoE and ACT having little amyloid and little learning disability. A analysis of young mice before the onset of amyloid formation shows that steady-state levels of monomeric A beta peptide are unchanged by ApoE or ACT. These data suggest that the process or product of amyloid formation is more critical than monomeric A beta for the neurological decline in AD, and that the risk factors ApoE and ACT participate primarily in disease processes downstream of APP processing.

Aging↗

PubMed2004

A liver X receptor and retinoid X receptor heterodimer mediates apolipoprotein E expression, secretion and cholesterol homeostasis in astrocytes.

Apolipoprotein E (apoE) is an important protein involved in lipoprotein clearance and cholesterol redistribution. ApoE is abundantly expressed in astrocytes in the brain and is closely linked to the pathogenesis of Alzheimer's disease (AD). We report here that small molecule ligands that activate either liver X receptors (LXR) or retinoid X receptor (RXR) lead to a dramatic increase in apoE mRNA and protein expression as well as secretion of apoE in a human astrocytoma cell line (CCF-STTG1 cells). Examination of primary mouse astrocytes also revealed significant induction of apoE mRNA, and protein expression and secretion following incubation with LXR/RXR agonists. Moreover, treatment of mice with a specific synthetic LXR agonist T0901317 resulted in up-regulation of apoE mRNA and protein in both hippocampus and cerebral cortex, indicating that apoE expression in brain can be up-regulated by LXR agonists in vivo. Along with a dramatic induction of ABCA1 cholesterol transporter expression, these ligands effectively mediate cholesterol efflux in both CCF-STTG1 cells and mouse astrocytes in the presence or absence of apolipoprotein AI (apoAI). Our studies provide strong evidence that small molecule LXR/RXR agonists can effectively mediate apoE synthesis and secretion as well as cholesterol homeostasis in astrocytes. LXR/RXR agonists may have significant impact on the pathogenesis of multiple neurological diseases, including AD.

Animals↗

PubMed2004

[Changes in tissue plasminogen activator and plasminogen activator inhibitor during administration of nitroglycerine into internal thoracic artery in dogs with acute myocardial ischemia].

OBJECTIVE: To study the changes in tissue plasminogen activator (t-PA) and plasminogen activator inhibitor (PAI) in a model of acute myocardial infarction reproduced in the dog with administration of nitroglycerin into internal thoracic artery under pressure. METHODS: Sixty healthy cross-breed dogs were randomly divided into experimental and control group with 30 dogs in each group. The animal model of acute myocardial ischemia was reproduced by ligating the left anterior descending coronary artery. Internal thoracic artery in the experimental group was ligated, and a tube was introduced into the proximal end. Nitroglycerine was infused under pressure into the internal thoracic artery in experimental group. The drug was given intravenously in control group. t-PA and PAI were measured before anterior coronary artery ligation and at 0.5, 2 and 6 hours after coronary artery ligation. RESULTS: The t-PA levels in two groups were increased at 0.5 hour after coronary artery ligation, and gradually declined in control group, while no obvious change was found in experimental group. There was significant difference between experimental group and control group at 6 hours after coronary artery ligation(P<0.05). PAI levels were increased after coronary artery ligation, peaking at 6 and 2 hours after coronary artery ligation in both groups. Significant difference in PAI level was observed between two groups at 6 hours after coronary artery ligation(P<0.05). CONCLUSION: Introduction of nitroglycerine through internal thoracic artery under pressure is effective to accelerate release of t-PA from the endothelium while inhibit secretion of PAI, therefore it is useful to modulate the balance of fibrinolysis.

Animals↗

PubMed2003

Nonsteroidal anti-inflammatory drugs can lower amyloidogenic Abeta42 by inhibiting Rho.

A subset of nonsteroidal anti-inflammatory drugs (NSAIDs) has been shown to preferentially reduce the secretion of the highly amyloidogenic, 42-residue amyloid-beta peptide Abeta42. We found that Rho and its effector, Rho-associated kinase, preferentially regulated the amount of Abeta42 produced in vitro and that only those NSAIDs effective as Rho inhibitors lowered Abeta42. Administration of Y-27632, a selective Rock inhibitor, also preferentially lowered brain levels of Abeta42 in a transgenic mouse model of Alzheimer's disease. Thus, the Rho-Rock pathway may regulate amyloid precursor protein processing, and a subset of NSAIDs can reduce Abeta42 through inhibition of Rho activity.

Amides↗

PubMed2003

A specific splicing variant of SVH, a novel human armadillo repeat protein, is up-regulated in hepatocellular carcinomas.

Hepatocellular carcinoma (HCC) is one of the most common malignant tumors with poor prognosis. By representational difference analysis (RDA), a novel human gene designated SVH, up-regulated in the clinical HCC sample, was identified. The deduced SVH protein consisted of 343 amino acids with a transmembrane domain and an armadillo repeat. Northern blot revealed that SVH was expressed in most human adult tissues. Four variants of SVH, SVH-A, -B, -C, and -D, resulting from alternative splicing in the coding region of the SVH transcript, were observed and were all localized in endoplasmic reticulum (ER). Up-regulation of SVH-B, but not the other variants, was evident in about 60% (28 of 46) of HCC samples, detected by quantitative real-time PCR. Human liver cell line QSG-7701, transfected with SVH-B, acquired an accelerated growth rate and tumorigenicity in nude mice, whereas inhibition of SVH-B in hepatoma cell line BEL-7404, using antisense oligodeoxynucleotides, induced apoptosis. It is suggested that the splicing variants of SVH have distinct biological functions, and SVH-B may play an important role in hepatocarcinogenesis.

Alternative Splicing↗

PubMed2003

Allelic imbalance regions on chromosomes 8p, 17p and 19p related to metastasis of hepatocellular carcinoma: comparison between matched primary and metastatic lesions in 22 patients by genome-wide microsatellite analysis.

To understand the molecular mechanisms of metastasis in hepatocellular carcinoma (HCC), it is necessary to identify the accumulating genetic alterations during its progression as well as those responsible for the acquisition of metastatic potential in cancer cells. In our previous study, using comparative genomic hybridization (CGH), we found that loss on chromosome 8p is more frequent in metastatic lesions than in matched primary tumors of HCC. Thus, 8p deletion might contribute to HCC metastasis. To narrow the location of metastasis-related alteration regions, we analyzed 22 primary and matched metastatic lesions of HCC by genome-wide microsatellite analysis. Common regions with high levels of allelic imbalance (AI) were identified on 17p, 8p11-cen, 8p21-23, 4q32-qter, 4q13-23, 16q, and 1p33. Regions with increased AI in metastatic lesions were 8p23.3, 8p11.2, 17p11.2-13.3, 4q21-22, 4q32-qter, 8q24.1, 9p11, 9q31, 11q23.1, 13q14.1-31, 13q32-qter, 16p13.3, 16q13, 16q22, and 19p13.1, and these were considered to be related to the metastasis phenotype. Among them, loss on 8p was again proved to be related to progression and metastasis of HCC, and 8p23.3 and 8p11.2 were two likely regions harboring metastasis-related genes. It was also shown for the first time in HCC that AI of 19p13.1 might also be related to metastatic potential. These results provide some candidate regions for further study to identify putative genes suppressing metastasis of HCC.

Adult↗

PubMed2003

Microglial apolipoprotein E and astroglial apolipoprotein J expression in vitro: opposite effects of lipopolysaccharide.

Apolipoprotein E (apoE) and apoJ are lipid carriers produced in the brain primarily by glial cells. A variety of glial-activating stimuli induce a parallel upregulation of both apolipoproteins expression in vivo and in vitro. To further characterize the cell type and mechanisms by which apoE and apoJ expression are upregulated in activated glia, mixed glial cultures from neonatal rat cortex were treated with the endotoxin lipopolysaccharide (LPS). LPS induced dose-dependent increases in apoJ and decreases in apoE expression and secretion with maximum effects at 1-10 ng/mL and 0.1-1 microg/mL, respectively. Experiments with enriched astroglial and microglial cultures demonstrated that apoE and apoJ expression are predominantly microglial and astroglial, respectively. Given the pivotal role that nuclear factor-kappa B (NF-kappa B) plays in glial activation, we assessed its possible role in mediating apoE and apoJ expression by activated glia. LPS robustly increased NF-kappa B activation in mixed glial cultures. Two NF-kappa B inhibitors, aspirin (10 mM) and MG-132 (0.1 microM), blocked basal apoE and apoJ secretion as well as LPS-induced apoJ secretion. These data demonstrate that glial apoE and apoJ expression are independently regulated by LPS in microglia and astroglia, respectively, and that activated microglia are the predominant source of apoE in mixed glial cultures. The transcription factor NF-kappa B appears to be a critical mediator of LPS-stimulated apoJ expression from astroglia.

Animals↗

PubMed2003

[Study on the risk factors of 100 cases with vulvar dystrophy].

OBJECTIVE: To assess the risk factors for vulvar dystrophy. METHODS: An epidemiological study was carried out. Data on 100 cases with vulvar dystrophy was reviewed and face to face interviewed with a uniform questionnaire including the manner of work, environmental temperature, habit of eating, mood, underwear wearing, autoimmune diseases, marriage, menstrual age, the quantity of menses, orders of pregnancy, and labor trauma of vulvar during delivery, vulvitis and urethritis ect. Univariable analysis and multivariate logistic regression analysis were carried out with 1:1 case-control methodology. RESULTS: Multiple conditional logistic regression analysis showed that vulvar dystrophy was positively associated with hot food (OR = 2.55, 95% CI: 1.24 - 5.25), mood (OR = 4.27, 95% CI: 1.96 - 9.29), order of pregnancy (OR = 3.37, 95% CI: 2.11 - 5.40), vulvitis (OR = 6.74, 95% CI: 2.66 - 17.09) and urethritis (OR = 11.02, 95% CI: 1.01 - 120.19). Vulviitis or urethritis increased 6.74 or 11.02 times the incidence of vulva dystrophy. Anger or nervous state contributed to the incidence of vulva dystrophy (OR = 4.27). Addict to hot food and order of labor also increased risk ratio for 2.55 and 3.37 times, respectively. CONCLUSION: The risk factors of vulvar dystrophy were: addict to hot food, often holding a angry or nervous state, increase of labors, having vulvitis and urethritis.

Diet↗

PubMed2003

[Introduction to genome databases].

A brief introduction to the genome databases GDB, GenoList and Ensembl is given. These databases, mirrored and maintained at the Centre of Bioinformatics, Peking University, provide useful information for genome research.

English Abstract↗

PubMed2002

Prediction and measurement of the interparticle depletion interaction next to a flat wall.

A theoretical and experimental study was performed to investigate the depletion interaction between two colloidal particles next to a solid wall in a solution of nonadsorbing macromolecules. By calculating the change in free volume available to the macromolecules upon approach of the two particles, a relatively simple expression was developed for the interparticle depletion attraction in hard sphere systems as a function of the particle-particle and particle-plate spacing. Perhaps the most useful result obtained from this analysis was that the wall has no effect whenever the ratio of the particle radius to the macromolecule radius is greater than four. (In charged systems, this ratio would apply to the effective particle and macromolecule sizes.) A series of experiments was then performed in which the hydrodynamic force balance (HFB) apparatus was used to measure the shear force needed to separate a colloidal doublet consisting of two particles trapped in a secondary energy well formed by a repulsive electrostatic force and an attractive depletion force. The macromolecules used here were small, nanometer-sized spheres of either silica or polystyrene. Agreement between the measured separation forces and those predicted using the force balance model of J. Y. Walz and A. Sharma (J. Colloid Interface Sci. 168, 485 (1994)) was within a factor of 1.3 using no adjustable parameters and accounting for polydispersity and uncertainty in the macromolecule size. It is shown that this remaining discrepancy could be caused by the Brownian (stochastic) nature of the doublet breakup process.

Journal Article↗

PubMed2002

alpha(4)beta(1) Integrin activation of L-type calcium channels in vascular smooth muscle causes arteriole vasoconstriction.

A pathway for the regulation of vascular tone appears to involve coupling between integrins and extracellular matrix proteins or their fragments and the subsequent modulation of ion movement across the smooth muscle cell membrane. Here, we report that the activation of L-type voltage-activated Ca(2+) channels occurs through a novel interaction of alpha(4)beta(1) integrin with peptides containing the Leu-Asp-Val (LDV) integrin--binding sequence, which is found in the CS-1 region of an alternately spliced fibronectin variant. Experiments were conducted on arterioles isolated from rat skeletal muscle. Arterioles exhibited sustained concentration-dependent vasoconstriction to LDV peptides but not to Leu-Glu-Val (LEV) control peptides. The constriction was associated with increased smooth muscle cell [Ca(2+)](i), as measured by using fura 2. The response could be inhibited with a function-blocking anti--alpha(4) integrin antibody. Removal of the endothelium did not alter the vasoconstrictor response. Further experiments demonstrated that the vasoconstriction was abolished by the L-type Ca(2+) channel inhibitor nifedipine and the Src family kinase inhibitor PP2. In studies of isolated smooth muscle cells using whole-cell patch-clamp methods, the L-type current was enhanced by the LDV but not LEV peptide and was blocked by PP2 or antibodies to alpha(4) integrin. Collectively, these data indicate that activation of alpha(4)beta(1) integrin leads to enhanced influx of Ca(2+) through L-type channels by activating a tyrosine kinase pathway, leading to vasoconstriction. Involvement of integrins in the modulation of vascular tone may be particularly important in vascular responses to mechanical signals, such as pressure and flow, and to tissue injury after damage to the extracellular matrix.

Alternative Splicing↗

PubMed2002

Immunization reverses memory deficits without reducing brain Abeta burden in Alzheimer's disease model.

We have previously shown that chronic treatment with the monoclonal antibody m266, which is specific for amyloid beta-peptide (Abeta), increases plasma concentrations of Abeta and reduces Abeta burden in the PDAPP transgenic mouse model of Alzheimer's disease (AD). We now report that administration of m266 to PDAPP mice can rapidly reverse memory deficits in both an object recognition task and a holeboard learning and memory task, but without altering brain Abeta burden. We also found that an Abeta/antibody complex was present in both the plasma and the cerebrospinal fluid of m266-treated mice. Our data indicate that passive immunization with this anti-Abeta monoclonal antibody can very rapidly reverse memory impairment in certain learning and memory tasks in the PDAPP mouse model of AD, owing perhaps to enhanced peripheral clearance and (or) sequestration of a soluble brain Abeta species.

Alzheimer Disease↗

PubMed2002

Regulation of ion channels by integrins.

Ion channels are regulated by protein phosphorylation and dephosphorylation of serine, threonine, and tyrosine residues. Evidence for regulation of channels by tyrosine phosphorylation comes primarily from investigations of the effects of growth factors, which act through receptor tyrosine kinases. The purpose of the present work is to summarize evidence for the regulation of ion channels by integrins, through their downstream, nonreceptor tyrosine kinases. We review both direct and indirect evidence for this regulation, with particular emphasis on Ca2+-activated K+ and voltage-gated Ca2+ channels. We then discuss the critical roles that cytoskeletal, focal-adhesion, and channel-associated scaffolding proteins may play in localizing nonreceptor tyrosine kinases to the vicinity of ion channels. We conclude by speculating on the physiological significance of these regulatory pathways.

Animals↗

PubMed2002

Association of hTcf-4 gene expression and mutation with clinicopathological characteristics of hepatocellular carcinoma.

AIM: Hepatocellular carcinoma(HCC) is a significant health problem in China. But the molecular mechanisms of HCC remains unclear. APC/beta-Catenin/Tcf signaling pathway, also known as Wnt pathway, plays a critical role in the development and oncogenesis. As little is known about the alteration of human T-cell transcription factor-4 (hTcf-4) gene in HCC, it is of interest to study the expression and mutation of hTcf-4 gene in HCC and the relationship between hTcf-4 gene and progression of HCC. METHODS: Reverse transcription-polymerase chain reaction (RT-PCR) method was used to detect the expression of hTcf-4 mRNA in 32 HCC and para-cancerous tissues and 5 normal liver tissues. PCR-single strand conformation polymorphism (PCR-SSCP) method was used to detect the mutation of hTcf-4 exons 1, 4, 9 and 15 in HCC. The correlation of expression and mutation of the hTcf-4 gene with clinicopathological characteristics of HCC was also analyzed. RESULTS: RT-PCR showed that the expression rate of hTcf-4 mRNA in HCC, para-cancerous tissues and normal liver tissues was 90.6 %, 71.9 % and 80 %, respectively. The gene expression level in tumor was 0.71+/-0.13, much higher than that in para-cancerous liver 0.29+/-0.05 and normal liver 0.26+/-0.05 (P<0.001), although there was no significant difference in gene expression level between para-cancerous tissues and normal liver (P>0.05). Furthermore, hTcf-4 gene expression was closely associated with tumor capsule status and intrahepatic metastasis of HCC. On SSCP, 2 of 32 cases of HCC (6.25 %) displayed characteristic mutational mobility shifts in exon 15 of the hTcf-4 gene. No abnormal shifting bands were observed in para-cancerous tissues. CONCLUSION: The high expression level of hTcf-4 in HCC, especially in tumors with metastasis, suggests that the over-expression of hTcf-4 gene may be closely associated with development and progression of HCC, but the mutation of this gene seemed to play less important role in this respect.

Adolescent↗

PubMed2002

Recombinant human heparin-binding neurite-promoting factor expressed with yeast stimulates neurites outgrowth.

OBJECTIVES: Heparin-binding neurite-promoting factor (HBNF) is a heparin-binding protein primarily found in the brain, which can stimulate neurite outgrowth in vitro. We expressed recombinant human heparin-binding neurite-promoting factor (hrHBNF) using a yeast system, and observed its activity in stimulating neurite outgrowth in vitro. METHODS: cDNA encoding mature human HBNF was amplified from total RNA isolated from an 18-week aborted human fetal brain by RT-PCR method. After amplification, the HBNF cDNA gene was cloned into pPIC9K, a shuttle expression vector for yeast system. The positive clone of expression vector bearing HBNF cDNA gene was obtained by screening. Verified recombinant vector was then used to transform Pichia strain GS115 by electroporation. His(+) transformants were selected on minimal dextrose medium (MD) plates which were histidine free. His(+) yeast recombinants with multi-copy inserts were screened in vivo by their resistance to G418. PCR analysis was used to confirm the integration of the HBNF cDNA gene into the Pichia genome. Secreted expression of hrHBNF protein in culture medium was obtained when the positive clone containing the HBNF cDNA gene was induced by methanol. The hrHBNF product purified by gel chromatography was added to cultured rat pheochromocytoma (PC12) cells to observe its ability to stimulate neurite outgrowth. RESULTS: In the recombinant expression vector, the insert was sequenced to show exactly the sequence encoding human HBNF according to Genbank data. The HBNF cDNA gene was cloned downstream to the alpha-factor, and its open reading frame was in frame with the alpha-factor signal sequence in pPIC9K. SDS-PAGE showed that the molecular weight of the induced expression product was about 18 kDa, consistent with that of human HBNF reported in the literature. The protein product did promote neurite outgrowth in cultured rat pheochromocytoma (PC12) cells. CONCLUSION: Recombinant human heparin-binding neurite-promoting factor can be expressed with a yeast system, and its product possesses the biological activity to promote neurite outgrowth.

Animals↗