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Biomedical subjects

G Wang

Publications and source records attributed to G Wang.

At least 73 records · Page 4Linked to original sources
PubMed2002

A cyclin-dependent kinase inhibitor (p21(WAF1/CIP1)) affects thymidine incorporation in human liver cancer cells.

p21(WAF1/CIP1) is a universal cyclin-dependent kinase inhibitor. To investigate the role of p21(WAF1/CIP1) in proliferation of human liver cancer cells, we examined the expression of p53, p21(WAF1/CIP1), cdk2 and cdk4 expression in two human liver cancer cell lines (HepG2 and PLC/PRF/5 cells). The effects of p21(WAF1/CIP1) on [(3)H]thymidine incorporation and cdks were also examined in these cells. HepG2 cells expressed all these proteins with lower level of cdk4. PLC/PRF/5 cells expressed the other proteins except p21(WAF1/CIP1). Transfection of p21(WAF1/CIP1) gene inhibited [(3)H]thymidine incorporation of both cells with different extent. Although the transfection of p21(WAF1/CIP1) did not affect cdk2 and cdk4 expression, it did reduce cdk2 kinase activity by 20%. These results suggest that: (a) p21(WAF1/CIP1) involved in DNA synthesis of human liver cancer cells; (b) p21(WAF1/CIP1) could be a target gene for the treatment of human hepatocellular carcinoma.

Adenosine Triphosphate↗
PubMed2002

Sorption and desorption of Co(II) on alumina: mechanisms and effect of humic substances.

The effects of pH, ionic strength and humic substances on the sorption and desorption of Co(II) on alumina and silica were, respectively investigated by using radiotracer 60Co. The distribution coefficients, the breakthrough curves and the displacement curves were experimentally determined in the batch and the column experiments. The pH and the humic substances influenced the sorption of Co(II) on alumina greatly as compared with the sorption of Co(II) on silica. It was found that the sorption characteristics of Co(II) onto alumina and silica are distinctly different, that the strong chemical bonds are formed between the bare alumina surface and Co(II) and between the coated alumina surface and Co(II), and that a transition from the adsorption to the surface-induced precipitation of Co(II) on the bare alumina surface takes place.

Journal Article↗
PubMed2002

Apical barriers to airway epithelial cell gene transfer with amphotropic retroviral vectors.

Gene transfer to airway epithelia with amphotropic pseudotyped retroviral vectors is inefficient following apical vector application. To better understand this inefficiency, we localized the expression of Pit2, the amphotropic receptor, in polarized human airway epithelia. Pit2 was expressed on both the apical and basolateral surfaces of the cells, suggesting that factors other than receptor abundance may limit apical gene transfer efficiency. Binding studies performed with radiolabeled amphotropic MuLV suggested that the apically applied virus binds to Pit2. Hypothetical barriers to retroviral gene transfer include the apical glycocalyx and other secreted products of epithelia. In this study, we demonstrated that sialic acid, keratan sulfate and collagen type V are present on the apical surface of well-differentiated human airway epithelia. While enzyme treatment reduced the abundance of these components, the treatment also decreased the transepithelial resistance to approximately 35% of the controls, suggesting that the epithelial integrity was impaired. To attain an airway epithelial culture with a modified apical surface and intact epithelial integrity, we utilized 100 mM 2-deoxy-D-glucose, a glycosylation inhibitor, to prevent the glycocalyx from reforming following enzyme treatment. This approach allowed the resistance, but not the apical glycocalyx to recover. Despite this physical modification of the cell surface, the amphotropic retroviral vector failed to transduce airway epithelia following apical application. These results suggest that factors other than apical receptor abundance and the glycocalyx inhibit amphotropic retroviral gene transfer in human airway epithelia.

Cell Polarity↗
PubMed2002

Increased functional expression of transgene in primary human lymphocytes using retroviral vectors modified with IRES and splicing motifs.

Genetic modification of human lymphocytes is being employed in strategies to correct enzyme deficiencies, encode cytokines and to redirect lymphocytes to antigenic targets other than those encoded by their endogenous T cell receptor. However, expression of transgenes in primary lymphocytes is generally low. Reasoning that vector modification may lead to increased transgene expression and subsequent increases in function, we have performed two retroviral vector modifications and report their effect on the functional expression in primary lymphocytes. A chimeric receptor specific for the colon carcinoma-associated antigen, EGP40, was initially incorporated into the retroviral vector LXSN. In this vector, receptor expression is driven by the Moloney murine leukemia virus LTR, and neomycin phosphotransferase expression driven by the SV40 promoter. Replacement of SV40 with an internal ribosomal entry site (IRES) increased the transgene activity of a mouse T cell line and human PBL as judged by increased cytokine release in response to antigen positive target cells. A further increase in transgene function was generated by the additional incorporation of a splice acceptor motif into the construct. Human PBL transduced with vector incorporating both IRES and intron were consistently more effective at lysing antigen positive colorectal carcinoma cells.

Adult↗
PubMed2002

Mechanism of action of methylphenidate: insights from PET imaging studies.

Methylphenidate (MPH) is the most commonly prescribed drug for the treatment of Attention-Deficit/Hyperactivity Disorder (ADHD). We have used Positron Emission Tomography (PET) to investigate the mechanism of action of MPH in the human brain. We have shown (a) that oral MPH reaches peak concentration in the brain 60-90 minutes after its administration, (b) that therapeutic doses of MPH block more than 50% of the dopamine transporters (DAT), and (c) that of the two enantiomers that compose MPH, it is d-threo-methylphenidate (d-MPH) and not l-threo-methylphenidate (l-MPH) that binds to the DAT. We have also shown that therapeutic doses of MPH significantly enhance extracellular dopamine (DA) in the basal ganglia, which is an effect that appears to be modulated by the rate of DA release and that is affected by age (older subjects show less effect). Thus, we postulate (a) that MPH's therapeutic effects are in part due to amplification of DA signals, (b) that variability in responses is in part due to differences in DA tone between subjects, and (c) that MPH's effects are context dependent. Because DA enhances task specific neuronal signaling and decreases noise, we also postulate that MPH-induced increases in DA could improve attention and decrease distractibility; and that since DA modulates motivation, the increases in DA would also enhance the saliency of the task facilitating the 'interest it elicits' and thus improving performance.

Adolescent↗
PubMed2001

[Screening of mouse connexin 31 genomic clone and mapping by fluorescence in situ hybridization].

Connexin 31(Cx31) is one of the human genetic deafness disease genes. To determine the structure and function of Cx31 gene of mouse, the mouse Cx31 gene clone was screened from the genomic library with the probe obtained according to the sequence of the mouse Cx31 gene cDNA. Cx31 gene was mapped to the center of mouse chromosome 4 by fluorescence in situ hybridization. The present study lays a foundation for researches on structure and function of Cx31 gene, as well as establishment of the relative transgenic mice and gene knockout mice.

Animals↗
PubMed2001

PAH and metal mixtures in New Orleans soils and sediments.

The purpose of this study is to determine the degree of PAH contamination and the association of PAHs with inorganic substances in soils and sediments of New Orleans. Bonnet Carré Spillway (BCS) (n = 5) provides modern baseline data, while urban soil samples (CTY) (n = 27) and sediment samples from Bayou St. John (BSJ) (n = 11) provide experimental data for New Orleans. Soil samples were collected from the top 2.5 cm of the surface, air-dried, and sieved (2 mm). Sediments samples were collected with a Wildco-Ekman bottom dredge, air-dried and finely ground. Accelerated solvent extraction (ASE) was used to release PAHs from the samples and analysis was conducted with gas chromatography-mass spectrometry (GC-MS). Metals were extracted using a 5:1 ratio of 1 mol/L nitric acid (room temperature) for soil and sediment samples, shaken for 2 h, centrifuged (1000 x g for 15 min) and filtered. Metal analysis was done by inductively coupled plasma-atomic emission spectrometry (ICP-AES). Mann-Whitney tests show PAH differences (P < 0.001). Ranking of total PAHs is, BSJ sediments (10.3 mg/kg) > CTY soils (3.7 mg/kg) > BCS alluvium (0.28 mg/kg). The sum of the metals are similar for BSJ sediments (698 mg/kg) and CTY soils (679 mg/kg) and significantly lower for BCS (189 mg/kg). Manganese of these samples is similar for each site. For paired samples, Pearson Product Moment Correlation tests reveal that many PAHs are strongly associated with each other at all locations. For BCS alluvium and BSJ sediments, total PAHs are not significantly associated with total metals. For CTY, most pairs of metals are significantly associated, and total soil PAHs are strongly associated with total soil metals (correlation 0.78, P = 4.9 x 10(-4)). The linear model, total soil PAH = 136.3 + 6.25 (total soil metals) forms the basis for a predicted PAH map of New Orleans. Previous empirical research demonstrates an association between soil lead and children's lead exposure. This study indicates that PAHs are part of the soil mixture of accumulated substances and by-products of industrial society that presents exposure potential in cities.

Child↗
PubMed2001

Homogeneous time-resolved fluorescence DNA hybridization assay by DNA-mediated formation of an EDTA-Eu(III)-beta-diketonate ternary complex.

A sensitive homogenous time-resolved fluorescence DNA hybridization assay method based on the formation of an EDTA-Eu(3+)-beta-diketonate ternary complex in the DNA hybrid was developed. The new approach combined the use of two DNA probes whose sequences compose the whole complementary strand to the target DNA, in which one probe was labeled with an EDTA-Eu(3+) complex on the 5'-terminus and the other, labeled with a bidentate beta-diketone on the 3'-terminus. After hybridization of two DNA probes with target DNA, EDTA-Eu(3+) and beta-diketone come close to each other, and an EDTA-Eu(3+)-beta-diketonate ternary complex with a strong and long-lived fluorescence was formed; thus the target DNA was detected sensitively with a detection limit of 6 pM (0.6 fmol per assay) by time-resolved fluorescence measurement. In the absence of the target DNA, due to the poor stability of bidentate beta-diketonate-Eu(3+) complex in very diluted solution, only a small amount of ternary fluorescence complex was formed.

DNA↗
PubMed2001

3-acetylpyridine reduces tongue protrusion force but does not abolish lick rhythm in the rat.

Data from other laboratories suggest that neurons in the inferior olivary nucleus (IO) may play a role in the modulation of rhythmic tongue movements in rats. Because of its known harmful effects on neurons of the IO, it was suspected that administration of the neurotoxin 3-acetylpyridine (3AP) would affect subsequent tongue dynamics during rat licking. In the present study, the task of licking water from a force-transducing disk was investigated in water-restricted rats that received systemic administration of 3AP (12.5, 25, and 50 mg/kg). After recovery from the acute toxic effects of 3AP, tongue dynamics were assessed by measuring lick force, lick rhythm, variability of timing within bursts of licking, and number of licks per 2-min session. At 50 mg/kg, 3AP resulted in: (1) reduced lick force; (2) reduced number of licks; and (3) increased variance in the timing within bursts. Lick rhythm was not significantly affected by any dose of 3AP. All 3AP treatment groups and the vehicle control group displayed slowing of lick rhythm after harmaline challenge. Compared to vehicle controls, rats receiving lower and mid-range doses of 3AP displayed indistinguishable lick behaviors, with one exception--when the lick task was made incrementally more difficult by extending the distance required to make contact with the lick-disk, rats that had received 25 mg/kg 3AP persevered at the task more than all other rats. The various changes in lick dynamics may be due to the detrimental effects of 3AP at the IO, and possibly at the hypoglossal nucleus and other sites.

Animals↗
PubMed2001

Defining the peptide nucleic acids (PNA) length requirement for PNA binding-induced transcription and gene expression.

Induction of gene expression has great potential in the treatment of many human diseases. Peptide nucleic acid (PNA) as a novel DNA-binding reagent provides an ideal system to induce gene-specific expression. In our recent studies, we have demonstrated that PNA bound to double-stranded DNA targets and, therefore, generated single-stranded D-loops and induced transcription of target genes both in vitro and in vivo. Most importantly, we have demonstrated that treatment of cultured human cells with PNAs led to expression of an endogenous target gene. Therefore, the study of the molecular mechanism of PNA binding-induced gene expression will have great implications for the gene therapy of many human diseases. In the current study, we have investigated the PNA length requirement for PNA binding-induced transcription initiation. Using a series of PNAs with different lengths, we have determined that PNAs with lengths of 16 approximately 18 nt induce very high levels of transcription in a HeLa nuclear extract in vitro transcription system. Transfection of the PNA-bound GFP reporter gene plasmid into human normal fibroblast (NF) cells led to a similar result. Gel-mobility shift assays revealed very strong binding affinities of these PNAs. DNA footprinting analysis further demonstrated the specificity of PNAs binding to the targets. These results lead to important understanding of the molecular mechanism of transcription initiation and highly valuable information in PNA design, especially for PNA binding-induced, gene-specific expression.

Base Sequence↗
PubMed2001

Laboratory diagnostic techniques for patients with early Lyme disease associated with erythema migrans: a comparison of different techniques.

Recently, a number of refinements in diagnostic modalities for detection of Borrelia burgdorferi infection have been developed. These include large-volume blood cultures, quantitative polymerase chain reaction (PCR) techniques, and 2-stage serologic testing. In the present study, we compared 6 diagnostic modalities in 47 adult patients who had a clinical diagnosis of erythema migrans. Quantitative PCR on skin biopsy-derived material was the most sensitive diagnostic method (80.9%), followed by 2-stage serologic testing of convalescent-phase samples (66.0%), conventional nested PCR (63.8%), skin culture (51.1%), blood culture (44.7%), and serologic testing of acute-phase samples (40.4%). Results of all assays were negative for 3 patients (6.4%). We conclude that the clinical diagnosis of erythema migrans is highly accurate in an area where B. burgdorferi is endemic if it is made by experienced health care personnel, but some patients with this diagnosis may not have B. burgdorferi infection. No single diagnostic modality is suitable for detection of B. burgdorferi in every patient with erythema migrans.

Biopsy↗
PubMed2001

Immunization against endogenous retroviral tumor-associated antigens.

Endogenous retroviral gene products have been found in some human tumors, and therefore, may serve as antigens for immunotherapy approaches. The murine colorectal carcinoma CT26 and melanoma B16 have recently been found to express the endogenous retroviral gene products gp70 and p15E, respectively, that can serve as antigens recognized by T cells. To date, though, there has been no demonstration of tumor treatment using an endogenous retroviral protein. In this study, we demonstrate that mice immunized with recombinant vaccinia encoding the gp70 H2-L(d)-restricted minimal determinant were protected from CT26 tumor challenge. Splenocytes from mice immunized with vaccinia gp70 specifically secreted IFN-gamma in response to gp70 peptide-pulsed stimulators. Although this strategy could protect against subsequent tumor challenge, it was ineffective against established tumors. Therefore, to investigate the treatment of established CT26 or B16 lung metastases, mice were treated with cultured dendritic cells (DCs) pulsed with gp70 or p15E peptide. Significant inhibition of established lung metastases required immunization with peptide-pulsed DCs pretreated with CD40 ligand that has been demonstrated to increase the T-cell stimulatory activity of DCs. The ability to immunize against endogenous retroviral tumor antigens may have relevance in the induction of antitumor immunity for some human cancers.

Animals↗
PubMed2001

Differential expression of apoptotic protease-activating factor-1 and caspase-3 genes and susceptibility to apoptosis during brain development and after traumatic brain injury.

Neuronal apoptosis plays an essential role in early brain development and contributes to secondary neuronal loss after acute brain injury. Recent studies have provided evidence that neuronal susceptibility to apoptosis induced by traumatic or ischemic injury decreases during brain development. However, the molecular mechanisms responsible for this age-dependent phenomenon remain unclear. Here we demonstrate that, during brain maturation, the potential of the intrinsic apoptotic pathway is progressively reduced and that such repression is associated with downregulation of apoptotic protease-activating factor-1 (Apaf-1) and caspase-3 gene expression. A similar decline in apoptotic susceptibility associated with downregulation of Apaf-1 expression as a function of developmental age was also found in cultured primary rat cortical neurons. Injury-induced cytochrome c-specific cleavage of caspase-9 followed by activation of caspase-3 in mature brain correlated with marked increases in Apaf-1 and caspase-3 mRNA and protein expression. These results suggest that differential expression of Apaf-1 and caspase-3 genes may underlie regulation of apoptotic susceptibility during brain development, as well as after acute injury to mature brain, through the intrinsic pathway of caspase activation.

Aging↗
PubMed2001

Microinjection and growth of bacteria in the cytosol of mammalian host cells.

Most facultative intracellular bacteria replicate in specialized phagosomes after being taken up by mammalian cells. Relatively few intracellular bacteria escape the phagosomal compartment with the help of cytolytic (pore-forming) proteins and replicate in the host cell cytosol. Without such toxins, intracellular bacteria cannot reach this cellular compartment. To circumvent the requirement of an "escape" step, we developed a procedure allowing the efficient direct injection of bacteria into the cytosol of mammalian cells. With this technique, we show that most bacteria, including extracellular bacteria and intracellular pathogens that normally reside in a vacuole, are unable to replicate in the cytosol of the mammalian cells. In contrast, microorganisms that replicate in the cytosol, such as Listeria monocytogenes, Shigella flexneri, and, to some extent, enteroinvasive Escherichia coli, are able to multiply in this cellular compartment after microinjection. Further L. monocytogenes with deletion in its PrfA-regulated hpt gene was found to be impaired in replication when injected into the cytosol. Complementation of the hpt mutation with a plasmid carrying the wild-type hpt gene restored the replication ability in the cytosol. These data indicate that cytosolic intracellular pathogens have evolved specific mechanisms to grow in this compartment of mammalian cells.

Animals↗
PubMed2001

Specific actions of cyanide on membrane potential and voltage-gated ion currents in rostral ventrolateral medulla neurons in rat brainstem slices.

The present study examined specific effects of sodium cyanide (CN) on the membrane potential (MP), spontaneous discharge (SD) and voltage-gated ion current of the identified bulbospinal rostral ventrolateral medulla (RVLM) neuron in the rat pup brainstem slice. 125 microM CN rapidly depolarized MP in the RVLM neuron by 11.6 mV as well as enhanced the SD rate by 300%. In contrast, the same dose of CN immediately hyperpolarized unlabeled, non-RVLM neurons by 4.8 mV. 50 microM CN did not significantly affect voltage-gated Ca(++) or A-type K(+) currents. The same concentration of CN, however, rapidly and reversibly suppressed voltage-gated Na(+) currents and sustained outward K(+) currents in the RVLM neuron by 22.5% and 23%, respectively. Tetraethylammonium could mimic the effect of CN on MP, SD and sustained K(+) current in the RVLM neuron. It is concluded that: (1) like that from the adult rat, the rat pup bulbospinal RVLM neuron can be selectively and rapidly excited by CN; (2) the hypoxia-sensitive, sustained outward K(+) channel may play an important role in the acute hypoxia-induced excitation of the RVLM neurons.

Animals↗
PubMed2001

Meteorological simulations of boundary-layer structure during the 1996 Paso del Norte Ozone Study.

Meteorological simulations centered around the border cities of El Paso and Ciudad Juárez have been performed during an ozone episode that occurred on August 13, 1996 during the 1996 Paso del Norte Ozone Study field campaign. Simulations were performed using the HOTMAC boundary-layer meteorological model using a 1, 2, 4 and 8-km horizontal grid size nested mesh system. Investigation of the vertical structure and evolution of the atmospheric boundary layer for the August 12-14 time period is emphasized in this paper due to its suspected importance in precipitating the ozone episode [Sci Total Environ (2001)]. This period was characterized by a slowly-evolving high pressure system over the region, a persistent upper-level jet at 2500-3500 m above ground level (agl), deep daytime mixed layer heights of 3500 m depth and unusually deep nighttime stable layers extending up to 2500 m above the ground. The fact that the boundary-layer growth stalled on the morning of August 13 relative to that on August 12 has been suggested as a possible reason for the ozone episode on the 13th. In addition, relatively weak surface-level winds were measured on August 13. Using both model results and experimental data we hypothesize explanations for the slower mixed-layer growth on the morning of the 13th and the stronger surface-level winds found on the 12th and 14th.

Air Movements↗
PubMed2001

Low concentration N-methyl-N'-nitro-N-nitrosoguanidine activates DNA polymerase-beta expression via cyclic-AMP-protein kinase A-cAMP response element binding protein pathway.

Ultraviolet light (UV), ionizing-radiation or alkylating agents are known as carcinogens, mostly because of their ability to damage DNA directly. However, they may also play a diverse role in activating the signal pathways and altering the gene expression. We have shown previously that N-methyl-N'-nitro-N-nitrosoguanidine (MNNG) of 0.2 microM can increase the transcription of DNA polymerase-beta gene, which has a cyclic AMP response element (CRE) motif in its promoter. Using the CRE report vector, we show here, such treatment can stimulate the CRE-driven gene expression by approximately 1.5-fold compared with control. Consistent with it, the proportion of ser-133 phosphorylated CRE binding protein (CREB), the related transcription factor was 2.08-fold higher versus control in vero cells after 60 min of MNNG treatment. Although CREB has many putative kinases for its phosphorylation, such as p38 mitogen-activated protein kinase (p38 MAPK), Ca(2+)/calmodulin-dependent protein kinase Pi (CaMK Pi) and protein kinase C (PKC), we found the protein kinase A (PKA) was activated and its activation peaked when cells were treated for 60 min (with arbitrary activity unit of 11.03+/-2.80 and 0.86+/-0.43 in treatment and control, respectively), this phasic character was similar to that of the CREB phosphorylation. We also determined the intracellular cyclic AMP (cAMP) levels and it was found that the cAMP concentration was elevated after 60 min treatment (1.53-fold higher). However, to our surprise, we did not find any accompanying cAMP elevation in cells treated by MNNG for 30 min, in which PKA was activated significantly. These findings, together with other observations, suggest that cAMP-PKA-CREB signaling pathway mediates the low concentration MNNG induced pol-beta expression. In addition to elevated cAMP, there might exist a cAMP-independent PKA activation manner in this course.

Animals↗