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Biomedical subjects

Z H Lu

Publications and source records attributed to Z H Lu.

At least 19 recordsLinked to original sources
PubMed2005

Precision measurement of the refractive index of air with frequency combs.

We report an experimental method for high-precision refractive index measurement using a Michelson interferometer setup with a femtosecond optical frequency comb as the light source. We demonstrate this technique by measuring the refractive index of air, under different pressures, inside a 29 m multipass cell. Both spectral and temporal interferograms are recorded. The method has a sensitivity of 6.2 x 10(-9) and a standard error of 7.4 x 10(-8) at fixed parameters (pressure and temperature). The possible variation of the density factor from the Lorentz-Lorenz equation is also discussed.

Journal Article↗
PubMed2005

Combinational synthesis of oligonucleotides and assembly fabrication of oligonucleotide array.

In this paper, a simple, reliable and flexible method, which integrated in situ synthesis with the spotting technique, was reported to fabricate oligonucleotide array. Different oligonucleotide sequences are synthesized on their relative code glass slides through combinational chemistry, thus the slides are broken into smaller pieces, in which the same code pieces have the same probe sequences. An oligonucleotide array is fabricated by arbitrarily assembling these different code pieces onto another solid substrate. In principle experimentation, four different sequences of P16 gene were synthesized and a 5 x 5 array including these four sequences and the control black was fabricated. The analysis results indicated that the hybridization fluorescence intensity of the same sequences locating different sets on the array gave the approximate values, and the fluorescence intensity ratio of matched sequence to one middle location base mismatched, two base mismatched, three middle base mismatched is (1.000+/-0.080):(0.4991+/-0.0671):(0.2360+/-0.0044):(0.0493+/-0.0033). Their relative accuracies were from 6.64 to 10.2%. This result might be used to rapidly screen single-nucleotide polymorphisms (SNPs).

Carbocyanines↗
PubMed2005

DNA microarray: a high throughput approach for methylation detection.

We described a DNA microarray-based method combined with bisulphite treatment of DNA and regular PCR to examine hyper-methylation in promoter 1A of APC gene. A set of oligonucleotide probes were designed and immobilized on the aldehyde-coated glass slides for detecting the methylation pattern of 15 selected CpG sites in the region. The methylation status of 30 colorectal tumor samples have been examined by both of methylation-specific PCR (MS-PCR) and the present microarray method. The methylation pattern of the 15 CpG sites for the samples have been obtained with the microarray. A total of 19 samples out of 30 were methylated by microarray, in which five samples cannot be detected by MS-PCR due to the methylated CpG patterns not accordant to the MS-PCR primers. The detecting ratio for methylation of APC gene of colorectal tumor samples increased from 46.7% with MS-PCR to 63.3% with the microarray, which successfully demonstrated that DNA microarray-based method not only can obtained the methylation patterns for the related genes, but also decrease the false-negative results of methylation status by the conventional MS-PCR for the investigated genes.

Adenomatous Polyposis Coli Protein↗
PubMed2004

Diagnostic investigation into the role of Chlamydiae in cases of increased rates of return to oestrus in pigs.

Cervical swabs and serum samples were taken from Swiss herds of sows with high rates of irregular return to oestrus (group A) and from control herds without reproductive problems (group B. The genital tracts of 21 slaughtered sows of group A were also examined. The swabs and genital tracts were screened for Chlamydiae by a new 16S rRNA PCR and the sera by an ELISA for Chlamydiaceae lipopolysaccharide. Chlamydophila (Cp) abortus was isolated from seven of the 65 swabs taken from group A but from none of the 128 swabs taken from group B. Chlamydia suis was present in swabs from both groups A (1.5 per cent) and B (2.3 per cent). In addition, Cp abortus was detected in 33.3 per cent of the genital tracts. Of the 193 sera tested, 61.7 per cent were positive, with no significant difference between group A (52.3 per cent) and group B (66.4 per cent). Chlamydia-like organisms were detected in 28.2 per cent of the swabs from group A and in 22 per cent of those from group B.

Animals↗
PubMed2004

Detection of mycobacteria and chlamydiae in granulomatous inflammation of reptiles: a retrospective study.

A retrospective study on reptile tissues presenting with granulomatous inflammation was performed to detect the possible presence of mycobacteria and chlamydiae in these lesions. Ninety cases including 48 snakes, 27 chelonians, and 15 lizards were selected. Mycobacteria were detected by Ziehl-Neelsen (ZN) staining and a broad-range polymerase chain reaction (PCR) followed by DNA sequencing. To detect chlamydiae, immunohistochemistry with monoclonal antibodies against chlamydial lipopolysaccharide (LPS) and a Chlamydiales order-specific PCR and sequencing were applied. Acid-fast bacilli were found in 14 cases (15.6%) by ZN staining and in 23 cases (25.6%) by PCR. Sequence analysis revealed the presence of Mycobacteria other than Mycobacterium tuberculosis complex (MOTT). Chlamydial LPS antigen was observed within granulomas from five samples (5.6%), whereas the PCR screen revealed 58 positive cases (64.4%). Of these, 9 cases (10%) showed 98-99% similarity to Chlamydophila (Cp.) pneumoniae and 49 cases (54.4%) displayed a high similarity (88-97%) to the newly described "Chlamydia-like" microorganisms Parachlamydia acanthamoebae and Simkania negevensis. Results from this study confirm, on the one hand, that MOTT are probably the most important infectious etiology for granulomatous inflammation in reptiles. On the other hand, they indicate that chlamydia infects reptiles and that Cp. pneumoniae should be considered an etiological agent of granulomatous lesions of reptiles. Because both MOTT and Cp. pneumoniae are human pathogens, the potential of zoonotic transmission from reptiles to humans has to be considered. In contrast, the significance of Chlamydia-like isolates remains completely open, and further studies are needed to evaluate their role.

Animals↗
PubMed2003

An effective method for quantitative evaluation of proteins adsorbed on biomaterial surfaces.

An effective method for the quantitative evaluation of proteins adsorbed on biomaterial surfaces has been developed. First, the kinetic behavior of a range of human fibrinogen (Fib) adsorbed onto polystyrene (PS) films was investigated by using a reflectometry interference spectroscopy setup. The specific molecular number of adsorbed proteins, N(p,) was then defined. According to the definition, the numbers of Fib molecules adsorbed on PS films were calculated. An atomic force microscope (AFM) was used to scan the lateral distribution of the Fib molecules adsorbed on the PS films. From the AFM images, the practical specific molecular numbers were obtained by direct counting of the molecules. In order that the adsorbed number of Fib molecules on a unit area of the PS films could be counted easily, the solution concentration of proteins was reduced to 5 ag/mL (10(-18)g/mL). There was good consistency between the numbers calculated with the formula defined by us and the numbers counted from AFM images. Therefore, the results of the present study prove the validity of our definition of the specific molecular number of adsorbed proteins and the effectiveness of the reflectometry interference spectroscopy-based method for quantitative evaluation of adsorptive proteins.

Adsorption↗
PubMed2002

The binding of ORC2 to chromatin from terminally differentiated cells.

Nuclei from terminally differentiated Xenopus erythrocytes lack essential components of the prereplication complex, including the origin recognition complex (ORC) proteins XORC1 and XORC2. In Xenopus egg extract, these proteins are able to bind erythrocyte chromatin from permeable nuclei, but not from intact nuclei, even though they are able to cross an intact nuclear envelope. In this report we use both permeable and intact erythrocyte nuclei to investigate the role of cyclin-dependent kinase activity in modulating the binding of XORC2 to chromatin. We find that elevating the level of cyclin A-dependent kinase in egg extract prevents the binding of XORC2 to chromatin from permeable nuclei and that kinase inhibition reverses this effect. We also observe a nuclear transport-dependent accumulation of H1 kinase activity within intact nuclei incubated in the extract. However, inhibiting this kinase activity does not facilitate the binding of XORC2 to chromatin, suggesting that other molecules and/or mechanisms exist to prevent association of XORC proteins with replication origins within intact nuclei from terminally differentiated cells.

Active Transport, Cell Nucleus↗
PubMed2001

Copper-induced proteolysis of the CopZ copper chaperone of Enterococcus hirae.

The cop operon is a key element of copper homeostasis in Enterococcus hirae. It encodes two copper ATPases, CopA and CopB, the CopY repressor, and the CopZ metallochaperone. It was previously shown that the transcription of the operon is induced by copper. The concomitant increase in the levels of Cop proteins, particularly the CopB copper export ATPase, allows uncompromised growth of E. hirae in up to 5 mm ambient copper. We here show by Western blotting that the steady-state level of CopZ was increased only up to 0.5 mm copper. At higher copper concentrations, the level of CopZ was decreased and became undetectable at 5 mm media copper. When CopZ was overexpressed from a plasmid, the cells exhibited increased sensitivity to copper and oxidative stress, suggesting that high CopZ expression could become toxic to cells. In wild-type cells, the level of mRNA transcripts from the cop operon remained high in up to 5 mm copper, suggesting that CopZ was proteolyzed. Cell extracts were found to contain a copper-activated proteolytic activity that degraded CopZ in vitro. In this assay, Cu-CopZ was more susceptible to degradation than apo-CopZ. The growth of E. hirae in copper increased the copper-inducible proteolytic activity in extracts. Zymographic studies showed the presence of a copper-dependent protease in crude cell lysates. Thus, copper-stimulated proteolysis plays an important role in the regulation of copper homeostasis in E. hirae.

Bacterial Proteins↗
PubMed2001

Lack of neighborhood effects from a transcriptionally active phosphoglycerate kinase-neo cassette located between the murine beta-major and beta-minor globin genes.

For the treatment of beta-globin gene defects, a homologous recombination-mediated gene correction approach would provide advantages over random integration-based gene therapy strategies. However, "neighborhood effects" from retained selectable marker genes in the targeted locus are among the key issues that must be taken into consideration for any attempt to use this strategy for gene correction. An Ala-to-Ile mutation was created in the beta6 position of the mouse beta-major globin gene (beta(6I)) as a step toward the development of a murine model system that could serve as a platform for therapeutic gene correction studies. The marked beta-major gene can be tracked at the level of DNA, RNA, and protein, allowing investigation of the impact of a retained phosphoglycerate kinase (PGK)-neo cassette located between the mutant beta-major and beta-minor globin genes on expression of these 2 neighboring genes. Although the PGK-neo cassette was expressed at high levels in adult erythroid cells, the abundance of the beta(6I) mRNA was indistinguishable from that of the wild-type counterpart in bone marrow cells. Similarly, the output from the beta-minor globin gene was also normal. Therefore, in this specific location, the retained, transcriptionally active PGK-neo cassette does not disrupt the regulated expression of the adult beta-globin genes. (Blood. 2001;98:65-73)

Animals↗
PubMed2001

Comparison of ganglioside profiles in nuclei and whole cells of NG108-15 and NG-CR72 lines: changes in response to different neuritogenic stimuli.

The plasma and nuclear membranes of neural cells have been shown to express gangliosides to a limited extent before, and at increasing levels during, differentiation. Recent studies employing qualitative cytochemistry have shown that GM1 expression in particular is significantly elevated in both membranes by specific neuritogenic agents. The present study provides a more complete description of ganglioside patterns of the 2 membranes of NG108-15 cells and a mutated form of the latter lacking gangliotetraose gangliosides. Nuclei of wild type NG108-15 cells were found to contain predominantly GM1 and GD1a, whereas whole cells had those in addition to substantial amounts of GM2 and GM3. GM1 and GD1a levels increased 2--3.5-fold in both whole cells and nuclei following axonogenic stimulation, but changed little in response to dendritogenic agents. GM2 expression, limited to the plasma membrane, showed little if any change with axonogenic stimuli but a 1.5--2-fold increase following treatment with dendritogenic agents. GM3 resembled GM2 in being virtually absent from the nuclear membrane, while its presence in the plasma membrane showed only modest change at most with any of the stimuli. The gangliotetraose ganglioside-deficient mutant cell line, NG-CR72, had significantly higher basal levels of GM2 in the plasma membrane compared to wild type NG108-15 cells, and this level increased significantly on treatment with dendritogenic agents. Basal GM3 levels were greatly reduced in the mutant cells and changed little with any of the stimuli. As expected, nuclei of NG-CR72 cells were virtually devoid of gangliosides. These mutant cells were previously shown to extend well defined dendritic neurites but were incapable of forming stable axonal processes. This study thus demonstrates major differences in the ganglioside content of wild type and mutated NG108-15 cells and their nuclei, and in their response to different neuritogenic stimuli.

Animals↗
PubMed2001

Cerebellar neurons lacking complex gangliosides degenerate in the presence of depolarizing levels of potassium.

Mice engineered to lack GM2/GD2 synthase (GalNAc-T), with resultant deficit of GM2, GD2, and all gangliotetraose gangliosides, were originally described as showing a relatively normal phenotype with only a slight reduction in nerve conduction. However, a subsequent study showed that similar animals suffer axonal degeneration, myelination defects, and impaired motor coordination. We have examined the behavior of cerebellar granule neurons from these neonatal knockouts in culture and have found evidence of impaired capacity for Ca2+ regulation. These cells showed relatively normal behavior when grown in the presence of physiological or moderately elevated K+ but gradually degenerated in the presence of high K+. This degeneration in depolarizing medium was accompanied by progressive elevation of intracellular calcium and onset of apoptosis, phenomena not observed with normal cells. No differences were detected in cells from normal vs. heterozygous mice. These findings suggest that neurons from GalNAc-T knockout mice are lacking a calcium regulatory mechanism that is modulated by one or more of the deleted gangliosides, and they support the hypothesis that maintenance of calcium homeostasis is one function of complex gangliosides during, and perhaps subsequent to, neuronal development.

Animals↗
PubMed2001

Mutant NG108-15 cells (NG-CR72) deficient in GM1 synthase respond aberrantly to axonogenic stimuli and are vulnerable to calcium-induced apoptosis: they are rescued with LIGA-20.

The neuroblastoma x glioma NG108-15 hybrid cell line, a widely used model for the study of neuronal differentiation, contains a variety of gangliosides including GM1 and its sialosylated derivative, GD1a. To investigate the role of these a-series gangliotetraose gangliosides in neuritogenesis, we have obtained a mutated subclone of NG108-15 that is deficient in that family of gangliosides. NG108-15 cells were grown in the presence of cholera toxin, which killed the large majority of cells, and from the cholera-resistant survivors we isolated a clone, NG-CR72, that lacks GM1 and GD1a in the plasma and nuclear membranes. GM2 concentration was significantly higher in the plasma membrane. Enzyme assay indicated deficiency of UDP-Gal:GM2 galactosyltransferase (GM1 synthase), which was confirmed by incorporation studies with [3H]sphingosine. These cells resembled wild-type NG108-15 in extending dendritic processes in response to dendritogenic agents (retinoic acid, dibutyryl cAMP) but responded aberrantly to axonogenic stimuli (KCl, ionomycin) by extending unstable neurites that showed the cytoskeletal staining characteristic of dendrites. Moreover, mutant cells treated with the Ca2+ elevating axonogenic agents underwent apoptosis over time, attributed to dysfunction of Ca2+ regulatory mechanisms normally mediated by GM1. Such agents caused dramatic and sustained elevation of intracellular Ca2+ in mutant cells, in contrast to modest and temporary elevation in wild-type cells. Exogenous GM1, inserted into the plasma membrane, had no discernable protective effect on NG-CR72 cells whereas LIGA-20, a membrane-permeant derivative of GM1 that entered both plasma and nuclear membranes, blocked apoptosis, permitted extension of stable neurites, and attenuated the abnormal elevation of intracellular Ca2+.

Apoptosis↗
PubMed2001

AKT-1, -2, and -3 are expressed in both normal and tumor tissues of the lung, breast, prostate, and colon.

PURPOSE: The AKT/PKB kinase controls many of the intracellular processes that are dysregulated in human cancer, including the suppression of apoptosis and anoikis and the induction of cell cycle progression. Three isoforms of AKT have been identified: AKT-1, -2, and -3. Selective up-regulation of AKT-3 RNA expression has been reported in hormone-independent breast and prostate cancer cell lines suggesting that AKT-3 expression may be increased with breast or prostate tumor progression. To determine whether AKT-3 RNA expression is selectively up-regulated in human cancers and whether the patterns of AKT RNA expression may change with tumor development, we examined AKT isoform expression by RT-PCR in human cancer cell lines, primary human cancers, and normal human tissues. EXPERIMENTAL DESIGN: AKT-1, -2, and -3 RNA expression was examined by RT-PCR. Because up-regulated AKT-3 expression has been implicated in human breast and prostate cancer progression, we also examined AKT-3 expression levels by semiquantitative RT-PCR using matched normal/tumor first-strand cDNA pairs from colon, breast, prostate, and lung cancers. RESULTS: Our data reveal that the overwhelming majority of both normal and tumor tissues express all three of the AKT isoforms. Moreover, semiquantitative RT-PCR of matched normal/tumor pairs confirmed similar AKT-3 RNA expression levels in both normal and tumor tissue. CONCLUSIONS: Our data show that both normal and tumor tissues express all three of the AKT isoforms and indicate that tumorigenesis does not involve a dramatic shift in the RNA expression patterns of the three AKT isoforms.

Breast↗
PubMed2001

[ARHI mRNA and protein expression in pancreatic cancers].

OBJECTIVE: To investigate ARHI mRNA and protein expression in pancreatic cancers. METHODS: Fifty-seven paraffin-embedded resected cancer samples and patient-corresponding normal pancreatic tissues were determined by using immunohistochemistry staining and in situ hybridization method. RESULTS: The positive rate of ARHI mRNA and protein expression in normal control pancreatic tissue was 84.2% and 82.5% respectively, and in cancer group was 52.6% both. The positive rate of mRNA and protein expression in cancer group was markedly decreased (P < 0.01). Yet the protein expression correlated with mRNA. Furthermore, positive staining of ARHI protein and mRNA can be observed in duodenal mucus epithelium, fibroblasts, smooth muscle myocytes, and wall of blood vessels. There was no correlation between positive rate and cancer differentiation, or clinical stages. CONCLUSIONS: ARHI protein expression is down-regulated in pancreatic cancer, possibly plays a role as tumor suppress gene, ARHI protein expressed in multiple types of tissues including duodenal mucus epithelium, fibroblasts and so on.

Adult↗
PubMed2000

Endogenous GM1 ganglioside of the plasma membrane promotes neuritogenesis by two mechanisms.

The influence of GM1 on the neuritogenic phase of neuronal differentiation has been highlighted in recent reports showing upregulation of this ganglioside in the plasma and nuclear membranes concomitant with axonogenesis. These changes are accompanied by alterations in Ca2+ flux which constitute an essential component of the signaling mechanism for axon outgrowth. This study examines 2 distinct mechanisms of induced neurite outgrowth involving plasma membrane GM1, as expressed in 3 neuroblastoma cell lines. Growth of Neuro-2a and NG108-15 cells in the presence of neuraminidase (N'ase), an enzyme that increases the cell surface content of GM1, caused prolific outgrowth of neurites which, in the case of Neuro-2a, could be blocked by the B subunit of cholera toxin (Ctx B) which binds specifically to GM1; however, the latter agent applied to NG108-15 cells proved neuritogenic and potentiated the effect of N'ase. With N18 cells, the combination was also neuritogenic as was Ctx B alone, whereas N'ase by itself had no effect. Neurite outgrowth correlated with influx of extracellular Ca2+, determined with fura-2. Treatment of NG108-15 and N18 cells with Ctx B alone caused modest but persistent elevation of intracellular Ca2+ while a more pronounced increase occurred with the combination Ctx B + N'ase. Treatment with N'ase alone also caused modest but prolonged elevation of intracellular Ca2+ in NG108-15 and Neuro-2a but not N18; in the case of Neuro-2a this effect was blocked by Ctx B. Neuro-2a and N18 thus possess 2 distinctly different mechanisms for neuritogenesis based on Ca2+ modulation by plasma membrane GM1, while NG108-15 cells show both capabilities. The neurites stimulated by N'ase + Ctx B treatment of N18 cells were shown to have axonal character, as previously demonstrated for NG108-15 cells stimulated in this manner and for Neuro-2a cells stimulated by N'ase alone.

Cell Membrane↗
PubMed2000

A transient increase in CCK mRNA levels in hippocampus following audiogenic convulsions in audiogenic seizure-prone rats.

AIM: To examine the effects of a single convulsion and multiple convulsions on cholecystokinin (CCK) mRNA expression in hippocampus of audiogenic seizure-prone rats (P77PMC). METHODS: Ringing (electric bell, 100 dB, 60 s) was used to induce convulsions, hippocampal CCK mRNA expression was exhibited by in situ hybridization. RESULTS: 1) The number of CCK mRNA-positive neurons in principal hippocampus of normal rats was 34 +/- 5, which elevated markedly after a single (155 +/- 7, P < 0.01) or multiple convulsions (95 +/- 8, P < 0.01). 2) CCK mRNA levels in multiple consecutive convulsion rats were lower than that in a single convulsion rats (P < 0.01). CONCLUSION: The increased number of CCK mRNA-positive neurons in hippocampus may have important functional consequences in convulsion-associated processes.

Acoustic Stimulation↗
PubMed1999

DNA replication in quiescent cell nuclei: regulation by the nuclear envelope and chromatin structure.

Quiescent nuclei from differentiated somatic cells can reacquire pluripotence, the capacity to replicate, and reinitiate a program of differentiation after transplantation into amphibian eggs. The replication of quiescent nuclei is recapitulated in extracts derived from activated Xenopus eggs; therefore, we have exploited this cell-free system to explore the mechanisms that regulate initiation of replication in nuclei from terminally differentiated Xenopus erythrocytes. We find that these nuclei lack many, if not all, pre-replication complex (pre-RC) proteins. Pre-RC proteins from the extract form a stable association with the chromatin of permeable nuclei, which replicate in this system, but not with the chromatin of intact nuclei, which do not replicate, even though these proteins cross an intact nuclear envelope. During extract incubation, the linker histones H1 and H1(0) are removed from erythrocyte chromatin by nucleoplasmin. We show that H1 removal facilitates the replication of permeable nuclei by increasing the frequency of initiation most likely by promoting the assembly of pre-RCs on chromatin. These data indicate that initiation in erythrocyte nuclei requires the acquisition of pre-RC proteins from egg extract and that pre-RC assembly requires the loss of nuclear envelope integrity and is facilitated by the removal of linker histone H1 from chromatin.

3T3 Cells↗