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Y Cao

Publications and source records attributed to Y Cao.

At least 145 records · Page 8Linked to original sources
PubMed2000

In-situ immuno-PCR to detect antigens.

In-situ immunoassays do not allow the detection of the minute numbers of target molecules accessible with in-situ PCR. We developed a highly sensitive method, termed in-situ immuno-PCR, in which the DNA marker was linked to target molecules through an antibody-biotin-avidin bridge and amplified by in-situ PCR. Amplified DNA sequences were detected in situ by hybridisation. This technique may be the only one available to detect minute quantities of biological macromolecules such as proteins, carbohydrates, and lipids in intact cells or tissue sections.

Antigens, Viral↗
PubMed2000

Phylogenetic relation of lungfish indicated by the amino acid sequence of myelin DM20.

The cDNA of lungfish Protopterus annectens myelin DM20 was cloned, and the complete amino acid sequence of Protopterusannectens DM20 was deduced. When five possible phylogenetic trees were tested for the DM20 sequences, the maximum likelihood method supported tree 1 [((tetrapods, lungfish), coelacanth), zebrafish, shark] or tree 5 [(tetrapods, lungfish), (coelacanth, zebrafish), shark]. Both tree 1 and tree 5 indicate that lungfish is phylogenetically the closest to tetrapods among the living fishes.

Amino Acid Sequence↗
PubMed2000

Blood-based RT-PCR assays of MN/CA9 or PSMA: clinical application in renal cancer patients.

OBJECTIVES: To report a survey of blood-based RNAs obtained from common groups of control and renal cancer patients for expression of both MN/CA9 and prostate-specific membrane antigen (PSMA) messenger RNAs. METHODS: Reverse transcription polymerase chain reaction (RT-PCR) assays for MN/CA9 and PSMA were performed on RNAs extracted from 81 blood samples (59 patients with renal cancer, 7 with benign tumors, and 15 control volunteers). The results of these assays were statistically analyzed to determine whether a positive result (individually or combined) correlates with any tumor characteristics. RESULTS: Neither MN/CA9 nor PSMA amplification products were detected in the RNAs from peripheral blood samples of the 15 control volunteers and from the 7 patients with benign renal tumor (sensitivity 100%). MN/CA9 alone was detected in 11 (19%) of 59 samples and PSMA alone in 12 (20%) of 59 samples from patients with renal cancer. PSMA positivity was significantly correlated with vascular invasion of the primary tumor. Expression of one or both of these molecular tumor markers was detected in 21 (36%) of 59 renal cancer patients. When combined, the results of the MN/CA9 and PSMA RT-PCR tests were found to be highly associated with vascular invasion in nephrectomy specimens (sensitivity 67%, specificity 77%, odds ratio = 6.89, P = 0.002). CONCLUSIONS: Combination of RT-PCR assays for MN/CA9 and PSMA provides a sensitive blood test for molecular detection of clear cell carcinoma of the kidney and its potential for vascular invasion. Further testing of this assay will be required to evaluate its efficacy in the diagnosis, screening, and follow-up of patients with kidney cancer.

Adult↗
PubMed2000

Block of rat brain recombinant SK channels by tricyclic antidepressants and related compounds.

SK channels are small conductance, Ca(2+)-activated K(+) channels that underlie neuronal slow afterhyperpolarization and mediate spike frequency adaptation. Using the patch clamp technique, we tested the effects of eight clinically relevant psychoactive compounds structurally related to the tricyclic antidepressants, on SK2 subtype channels cloned from rat brain and functionally expressed in the human embryonic kidney cell line, HEK293. Amitriptyline, carbamazepine, chlorpromazine, cyproheptadine, imipramine, tacrine and trifluperazine blocked SK2 channel currents with micromolar affinity. The block was reversible and concentration-dependent. The potency differed according to chemical structure. In contrast, the cognitive enhancer linopirdine was ineffective at blocking these channels. Our results point to a distinct pharmacological profile for SK channels.

Animals↗
PubMed2000

SSA/RO52gene and expressed sequence tags in an 85 kb region of chromosome segment 11p15.5.

Frequent allelic loss in lung cancer has been described in a region on chromosome segment 11p15.5 (LOH11A). The region is approximately 650 kb in size and flanked by the markers D11S988 centromeric and D11S860 telomeric. Clinical and cell biological studies suggest that it contains a gene associated with metastatic tumor spread. One of the genes identified within this region is SSA/Ro52, which has a RING finger domain and may be involved in gene regulation. We studied this gene for mutations using SSCP analysis and for expression using RT-PCR and Western blotting on lung cancer cell lines and tumor-normal tissue pairs. No mutations and no differences in mRNA or protein expression between tumor tissue and normal tissue pairs were identified. We discovered a novel polymorphic site (SSA44C/T) within exon 1 of this gene. Among 141 primary lung cancers, allelic loss was observed in 16% of informative cases. Our analyses excluded SSA/Ro52 as a tumor-suppressor gene in lung cancer and newly defined the centromeric border of the LOH11A region from D11S988 previously to SSA44C/T. This reduced the region of the putative suppressor gene to 460 to 485 kb. A significant difference (p = 0.01) in the frequency of alleles for this polymorphism between Caucasians and African-Americans was observed. The "T" allele frequency was 0.12 in Caucasians and 0.23 in African-Americans. A genomic EcoRI map over 85 kb surrounding the SSA/Ro52 gene was constructed, and 4 expressed sequence tags were identified by sequencing and studied.

Alleles↗
PubMed2000

Repeated administration of low dose ketamine for the treatment of monoarthritic pain in the rat.

The aim of the present study was to observe the effect of repeated subcutaneous (sc) injections of low doses of ketamine for the treatment of acute inflammatory pain in a complete Freund's adjuvant-induced monoarthritic pain model in rats. The results show: (1) sc injection of ketamine at a dose of 2 mg x kg(-1) or 10 mg x kg(-1) produced significant analgesia (P<0.01) in arthritic rats starting from the 2nd week and 3rd week, respectively. (2) Repeated administration of ketamine produced a significant reduction of the circumference of the arthritic ankle (P<0.05 and P<0.01 with different doses). (3) The body weight of the rats was not affected by continuous administration of ketamine for 4 weeks. No abnormal locomotor behavior was observed (It was concerned but not systemically evaluated in this study). The results suggest that repeated sc injection of ketamine for 4 weeks significantly reduce inflammatory pain in monoarthritis without notable aversive side effects.

Analgesics↗
PubMed2000

The cytokine hepatocyte growth factor/scatter factor inhibits apoptosis and enhances DNA repair by a common mechanism involving signaling through phosphatidyl inositol 3' kinase.

Scatter factor (SF) [aka. hepatocyte growth factor (HGF)] (designated HGF/SF) is a multifunctional cytokine that stimulates tumor cell invasion and angiogenesis. We recently reported that HGF/SF protects epithelial and carcinoma cells against cytotoxicity from DNA-damaging agents and that HGF/SF-mediated cytoprotection was associated with up-regulation of the anti-apoptotic protein Bcl-XL in cells exposed to adriamycin. We now report that in addition to blocking apoptosis, HGF/SF markedly enhances the repair of DNA strand breaks caused by adriamycin or gamma radiation. Constitutive expression of Bcl-XL in MDA-MB-453 breast cancer cells not only simulated the HGF/SF-mediated chemoradioresistance, but also enhanced the repair of DNA strand breaks. The ability of HGF/SF to induce both chemoresistance and DNA repair was inhibited by wortmannin, suggesting that these activities of HGF/SF are due, in part, to a phosphatidylinositol-3'-kinase (PI3K) dependent signaling pathway. Consistent with this finding, HGF/SF induced the phosphorylation of c-Akt (protein kinase-B), a PI3K substrate implicated in apoptosis inhibition; and an expression vector encoding a dominant negative kinase inactive Akt partially but significantly inhibited HGF/SF-mediated cell protection and DNA repair. These findings suggest that HGF/SF activates a cell survival and DNA repair pathway that involves signaling through PI3K and c-Akt and stabilization of the expression of Bcl-XL; and they implicate Bcl-XL in the DNA repair process.

Antimutagenic Agents↗
PubMed2000

Impaired endochondral ossification and angiogenesis in mice deficient in membrane-type matrix metalloproteinase I.

Membrane-type matrix metalloproteinase I (MT1-MMP)-deficient mice were found to have severe defects in skeletal development and angiogenesis. The craniofacial, axial, and appendicular skeletons were severely affected, leading to a short and domed skull, marked deceleration of postnatal growth, and death by 3 wk of age. Shortening of bones is a consequence of decreased chondrocyte proliferation in the proliferative zone of the growth plates. Defective vascular invasion of cartilage leads to enlargement of hypertrophic zones of growth plates and delayed formation of secondary ossification centers in long bones. In an in vivo corneal angiogenesis assay, null mice did not have angiogenic response to implanted FGF-2, suggesting that the defect in angiogenesis is not restricted to cartilage alone. In tissues from null mice, activation of latent matrix metalloproteinase 2 was deficient, suggesting that MT1-MMP is essential for its activation in vivo.

Animals↗
PubMed2000

Modeling plasma virus concentration during primary HIV infection.

During primary HIV infection the viral load in plasma increases, reaches a peak, and then declines. Phillips has suggested that the decline is due to a limitation in the number of cells susceptible to HIV infection, while other authors have suggested that the decline in viremia is due to an immune response. Here we address this issue by developing models of primary HIV-1 infection, and by comparing predictions from these models with data from ten anti-retroviral, drug-naive, infected patients. Applying nonlinear least-squares estimation, we find that relatively small variations in parameters are capable of mimicking the highly diverse patterns found in patient viral load data. This approach yields an estimate of 2.5 days for the average lifespan of productively infected cells during primary infection, a value that is consistent with results obtained by drug perturbation experiments. We find that the data from all ten patients are consistent with a target-cell-limited model from the time of initial infection until shortly after the peak in viremia. However, the kinetics of the subsequent fall and recovery in virus concentration in some patients are not consistent with the predictions of the target-cell-limited model. We illustrate that two possible immune response mechanisms, cytotoxic T lymphocyte destruction of infected target cells and cytokine suppression of viral replication, could account for declines in viral load data not predicted by the original target-cell-limited model. We conclude that some additional process, perhaps mediated by CD8+ T cells, is important in at least some patients.

Cytokines↗
PubMed2000

Cortical language lateralization in right handed normal subjects using functional magnetic resonance imaging.

In 95% of right handed individuals the left hemisphere is dominant for speech and language function. The evidence for this is accumulated primarily from clinical populations. We investigated cortical topography of language function and lateralization in a sample of the right handed population using functional magnetic resonance imaging and two lexical-semantic paradigms. Activated cortical language networks were assessed topographically and quantitatively by using a lateralization index. As a group, we observed left hemispheric language dominance. Individually, the lateralization index varied continuously from left hemisphere dominant to bilateral representation. In males, language primarily lateralized to left, and in females, approximately half had left lateralization and the other half had bilateral representation. Our data indicate that a previous view of female bilateral hemispheric dominance for language (McGlone, 1980. Sex differences in human brain asymmetry: a critical survey. Behav Brain Sci 3:215-263; Shaywitz et al., 1995. Sex differences in the functional organization of the brain for language. Nature 373:607-609) simplifies the complexity of cortical language distribution in this population. Analysis of the distribution of the lateralization index in our study allowed us to make this difference in females apparent.

Adult↗
PubMed2000

The COOH-terminal transactivation domain plays a key role in regulating the in vitro and in vivo function of Pax3 homeodomain.

Efficient transcription activation by Pax3 requires binding to a complex DNA sequence element containing binding sites for both the paired domain and the Prd type homeodomain. Previously, we have shown that this requirement is lost in PAX3-FKHR, the product of a t(2;13) chromosomal translocation associated with alveolar rhabdomyosarcoma. In contrast to Pax3, the chimeric PAX3-FKHR, which acts as an oncogene, can efficiently activate a DNA sequence element containing only a homeodomain binding site (TAATAN(2-3)ATTA), despite the presence of an intact Pax3 paired domain. Here, we showed that this alteration in sequence-specific transcription activity was determined in part by the transactivation domain. First, we demonstrated that in intact Pax3, substitution of the Pax3 transactivation domain with an unrelated viral VP16 transactivation domain enabled Pax3 to transactivate homeodomain-specific DNA sequence, as well as to transform fibroblasts. Furthermore, we could abolish the homeodomain-dependent transcription and transforming activities of PAX3-FKHR by replacing its FKHR transactivation domain with Pax3 transactivation domain. Collectively, these results suggested that the transactivation domain influences the DNA binding specificity of Pax3. The translocation process increased the oncogenic potential of Pax3 by removing the inhibitory action of Pax3 transactivation domain on its homeodomain.

3T3 Cells↗
PubMed2000

Expression of fatty acid-CoA ligase 4 during development and in brain.

Fatty acid utilization is initiated by fatty acid-CoA ligase, which converts free fatty acids into fatty acyl-CoA esters. We have cloned previously the human long-chain fatty acid-CoA ligase 4 (FACL4), which is a central enzyme in controlling the free arachidonic acid level in cells and thereby regulating eicosanoid production. We report here the expression of this gene in tissues, particularly in different parts of the brain. We found that FACL4 encoded a 75 kDa enzyme and that there was a modified translation product expressed in the brain. FACL4 was expressed in early stages of development with a significant amount of FACL4 mRNA detected in an E7 mouse embryo. In addition, FACL4 was highly expressed in both adult and newborn mouse brain especially in the granule cells of the dentate gyrus and the pyramidal cell layer of CA1 in hippocampus, and the granular cell layer and Purkinje cells of the cerebellum.

Animals↗
PubMed2000

Influence of growth factors on tissue-engineered pediatric elastic cartilage.

OBJECTIVE: To investigate the influence of growth factors on tissue-engineered pediatric human elastic cartilage relative to potential clinical application. DESIGN: Controlled study. SUBJECTS: Eleven children ranging in age from 5 to 15 years provided auricular elastic cartilage specimens measuring approximately 1 x 1 x 0.2 cm and weighing approximately 100 mg. INTERVENTIONS: Three million chondrocytes were plated into 4 groups of Ham F-12 culture medium: group 1, Ham F-12 culture medium only; no growth factors (control group); group 2, Ham F-12 culture medium and basic fibroblast growth factor; group 3, Ham F-12 culture medium and transforming growth actor beta; and group 4, Ham F-12 culture medium and a combination of both growth factors. At 3 weeks, the cells were harvested and mixed with a copolymer gel of polyethylene glycol and polypropylene oxide (Pluronic F-127). The cell solution was injected subcutaneously into athymic mice. The constructs were harvested at up to 22 weeks of in vivo culture and histologically analyzed. RESULTS: The average number of cells generated in vitro was as follows: group 1, 12 million; group 2, 40 million; group 3, 7 million; and group 4, 35 million. Group 2 in vivo gross specimens were the largest and heaviest. Histologically, the control group and the basic fibroblast growth factor group (groups 1 and 2) exhibited characteristics compatible with normal auricular cartilage; groups 3 and 4 demonstrated cellular disorganization and moderate to severe fibrous tissue infiltration. CONCLUSIONS: Basic fibroblast growth factor demonstrates the greatest positive influence on the in vitro and in vivo growth of engineered pediatric human auricular cartilage. The results suggest that basic fibroblast growth factor has the potential for clinical application in which a goal will be to generate a large volume of tissue-engineered cartilage from a small donor specimen in a short period of time and of a quality similar to native human elastic cartilage.

Adolescent↗
PubMed2000

Internal support of tissue-engineered cartilage.

BACKGROUND: Auricles previously created by tissue engineering in nude mice used a biodegradable internal scaffold to maintain the desired shape of an ear. However, the biodegradable scaffold incited a compromising inflammatory response in subsequent experiments in immunocompetent animals. OBJECTIVE: To test the hypothesis that tissue-engineered autologous cartilage can be bioincorporated with a nonreactive, permanent endoskeletal scaffold. MATERIALS AND METHODS: Auricular elastic cartilage was harvested from Yorkshire swine. The chondrocytes were isolated and suspended into a hydrogel (Pluronic F-127) at a cell concentration of 5 x 10(7) cells/mL. Nonbiodegradable endoskeletal scaffolds were formed with 1 of 5 polymers: (1) high-density polyethylene, (2) soft acrylic, (3) polymethylmethacrylate, (4) extrapurified Silastic, and (5) conventional Silastic. Three groups were studied: (1) a control group using only the 5 polymers, (2) the 5 polymers enveloped by Pluronic F-127 only, and (3) the implants coated with Pluronic F-127 seeded with chondrocytes. All constructs were implanted subdermally; implants containing cells were implanted into the same animal from which the cells had been islolated. The implants were harvested after 8 weeks of in vivo culture and histologically analyzed. RESULTS: Only implants coated by hydrogel plus cells generated healthy new cartilage. With 3 polymers (high-density polyethylene, acrylic, and extrapurified Silastic), the coverage was nearly complete by elastic cartilage, with minimal fibrocartilage and minimal to no inflammatory reaction. The Food and Drug Administration-approved conventional Silastic implants resulted in fragments of fibrous tissue mixed with elastic cartilage plus evidence of chronic inflammation. The polymethylmethacrylate implant was intermediate in the amount of cartilage formed and degree of inflammation. CONCLUSIONS: This pilot technique combining tissue-engineered autologous elastic cartilage with a permanent biocompatible endoskeleton demonstrated success in limiting the inflammatory response to the scaffold, especially to high-density polyethylene, acrylic, and extrapurified Silastic. This model facilitates the potential to generate tissue of intricate shape, such as the human ear, by internal support. Arch Otolaryngol Head Neck Surg. 2000;126:1448-1452

Animals↗
PubMed2000

Serological and histological findings in infection and transmission of GBV-C/HGV to macaques.

Seven healthy macaques were inoculated with the GBV-C/HGV-RNA serum from a non-A-E hepatitis patient. The serology and pathology of the liver in the animals were observed. The results indicated that all inoculated animals were infected with a GBV-C/HGV-RNA viremia and had mildly abnormal alanine transaminase levels during the infectious period. The histology, immuno-histochemistry, and in situ hybridization in the liver tissues of the inoculated animals also showed that there was a very mild hepatitis with the positive antigenic expression and the genome of GBV-C/HGV-NS5 in hepatocytes. The pathological changes in the infected animals appeared to become normal whether or not GBV-C/HGV-RNA viremia persisted. There is a possibility that the mild virulence of the GBV-C/HGV to the host became harmless with time after inoculation. Infection and the transmission of the GBV-C/HGV virus in the macaques provides an appropriate animal model and new information about GBV-C/HGV infection in both humans and animals. It is possible that this virus is a mild and self-limited pathogenic agent to the hepatic cells of primates.

Alanine Transaminase↗
PubMed2000

Flavin-containing monooxygenase isoform specificity for the N-oxidation of tamoxifen determined by product measurement and NADPH oxidation.

The Km value for tamoxifen is 1.2 mM for mouse FMO1 (human FMO1 is not expressed in adults) and 1.4 mM for human FMO3, with no detectable activity being expressed toward tamoxifen by FMO5 from either mouse or human. These data are derived from experiments using 3H-tamoxifen as substrate in which the product, tamoxifen N-oxide, was measured directly. It was not possible to derive meaningful data from the measurement of NADPH consumption because Escherichia coli preparations, in the presence of tamoxifen, regardless of whether the E. coli was expressing an FMO isoform, consumed large amounts of NADPH without the appearance of tamoxifen N-oxide or other discernable product.

Animals↗
PubMed2000

Induced expression of dominant-negative c-jun downregulates NFkappaB and AP-1 target genes and suppresses tumor phenotype in human keratinocytes.

Neoplastically transformed mouse and human keratinocytes elevate transactivation of both activator protein 1 (AP-1) and nuclear factor kappaB (NFkappaB) transcription factors. The present study addresses the question of whether elevated NFkappaB in addition to elevated AP-1-dependent gene expression is necessary for maintaining the tumor cell phenotype. When a tetracycline-regulatable dominant-negative c-jun (TAM67, having a truncated transactivation domain) was expressed in tumorigenic human keratinocytes, AP-1- and NFkappaB- but not p53-dependent reporter activity was inhibited by 40-60%. Tumor phenotype, as measured by anchorage-independent growth, was inhibited by 90%. Neither AP-1/NFkappaB activation nor expression of tumor phenotype was inhibited in TAM67-harboring keratinocytes under noninducing conditions. Electrophoretic mobility shift analysis showed that induction of TAM67 expression slightly increased AP-1- but reduced NFkappaB DNA-binding activity. Immunoprecipitation showed that TAM67 interacted in keratinocyte nuclei with NFkappaB p65, suggesting that inhibition of NFkappaB by TAM67 is mediated by direct protein-protein interactions, possibly producing decreased binding to DNA or inactivating p65. To analyze the putative effector genes that may be targeted by TAM67, expression of genes responsive to AP-1 or NFkappaB was measured by reverse transcriptase-polymerase chain reaction in TAM67 transfectants with or without TAM67 induction. Induction of TAM67 inhibited or reduced the expression of collagenase I, stromelysin I (AP-1 responsive), and interleukins 1 and 6 (NFkappaB responsive). These results indicate that genes controlled by NFkappaB and by AP-1 may be transformation-relevant targets of TAM67 and that TAM67 may inhibit NFkappaB activation through direct interaction with NFkappaB p65. Moreover, the findings provide proof for the principle of using inducible TAM67 as a gene therapy to suppress tumor phenotype in human carcinoma cells.

Cell Adhesion↗