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Biomedical subjects

M D Wang

Publications and source records attributed to M D Wang.

At least 19 recordsLinked to original sources
PubMed2006

Advances in molecular classification of renal neoplasms.

Kidney neoplasms are classified by light microscopy using the World Health Organization (WHO) system. The WHO system defines histopathologic tumor subtypes with distinct clinical behavior and underlying genetic mutations. In adults, the common malignant subtypes are variants of renal cell carcinoma (RCC). Histopathologic classification is critical for clinical management of RCC, but is becoming more complex with recognition of novel tumor subtypes, development of procedures yielding small diagnostic biopsies, and emergence of molecular therapies directed at tumor gene activity. Therefore, classification systems based on gene expression are likely to become essential for diagnosis, prognosis and treatment of kidney tumors. Recent DNA microarray studies have shown that clinically relevant renal tumor subtypes are characterized by distinct gene expression profiles, which are useful for discovery of novel diagnostic and prognostic biomarkers. In this review, we summarize the WHO classification system for renal tumors, general applications of microarray technology in cancer research, and specific microarray studies that have advanced knowledge of renal tumor diagnosis, prognosis, therapy and pathobiology.

Carcinoma, Renal Cell↗
PubMed2001

The functional role of a bicuculline-sensitive Ca2+-activated K+ current in rat medial preoptic neurons.

A Ca2+-activated K+ current was identified in neurons from the rat medial preoptic nucleus. Its functional role for the resting potential and for impulse generation was characterised by using the reversible blocking agent bicuculline methiodide. Acutely dissociated neurons were studied by perforated-patch recordings. The effect of bicuculline methiodide was investigated under voltage-clamp conditions to clearly identify the current affected. At membrane potentials > -50 mV, bicuculline methiodide rapidly (< 1 s) and reversibly blocked a steady outward current. Half-saturating concentration was 12 microM. The current amplitude increased with potential in the range -50 to 0 mV. The bicuculline-sensitive current was identified as an apamin-sensitive, Ca2+-dependent K+ current. It was neither affected by the GABAA receptor blocker picrotoxin (100 microM) nor by a changed pipette Cl- concentration, but was affected by substitution of extracellular K+ for Na+. The current was dependent on extracellular Ca2+ and was sensitive to 1 microM apamin but not to 200 nM charybdotoxin. A role for the Ca2+-dependent K+ current in setting the resting potential and controlling spontaneous firing frequency was observed under current-clamp conditions. Bicuculline methiodide (100 microM) induced a positive shift (5 +/- 1 mV; n = 18) of resting potential in all neurons tested. In the majority of spontaneously firing neurons, the firing frequency was reversibly affected, either increased or decreased depending on the cell, by bicuculline methiodide.

Animals↗
PubMed2001

The interaction between ethanol and pregnanolone at induction of anaesthesia investigated with a threshold method in male rats.

1. An anaesthesia threshold was used to investigate the pharmacodynamic and pharmacokinetic interactions between ethanol and pregnanolone in male rats. 2. The criterion to determine threshold doses of pregnanolone was the first burst suppression of 1 s in the EEG. 3. Ethanol (0.5, 1.0, 1.5 and 2.0 g kg(-1)) was injected i.p. 15 min before pregnanolone infusion. Trunk blood, serum, cortex, cerebellum, hippocampus, striatum, brain stem, fat and muscle tissues obtained at criterion were used to determine ethanol (blood) and pregnanolone. Ethanol reduced threshold doses in a dose dependent linear manner. A similar reduction of pregnanolone tissue concentrations was only found in brain stem and striatum. Deviations consisted of larger decreases in serum, cerebellum and hippocampus after 0.5 g kg(-1) ethanol and in cerebellum, cortex and hippocampus after 2.0 g kg(-1) of ethanol. Positive correlations between dose and concentration of pregnanolone was recorded in brain stem, hippocampus, cerebellum and cortex. A kinetic component influenced the concentration in cortex. There was a correlation between dose and serum concentration of pregnanolone only after ethanol. In the muscle 0.5 g kg(-1) ethanol had no influence on pregnanolone concentration. 4. The linear, additive pharmacodynamic interaction could involve the GABA ionophore. A pharmacokinetic interaction was found in cortex. The retained high uptake of pregnanolone in muscle (after 0.5 g kg(-1)) corresponded to losses in other tissues (including serum). The reduced uptake of pregnanolone in cerebellum, cortex and hippocampus (after 2.0 g kg(-1)) was not due to a corresponding change in serum concentration. It was probably due to a reduced blood flow.

Anesthesia↗
PubMed2001

Evaluation and comparison of the pharmacokinetic and pharmacodynamic properties of allopregnanolone and pregnanolone at induction of anaesthesia in the male rat.

We have evaluated and compared the pharmacokinetic and pharmacodynamic properties of allopregnanolone and pregnanolone at induction of anaesthesia in male rats. A threshold method was used, and the first burst suppression period of 1 s or more in the EEG was selected as the end-point after fairly slow infusions. An optimal dose of 4.0 mg kg(-1) min(-1) was noted for both steroids. Brain concentrations were low at low infusion rates, indicating that acute tolerance was not occurring. Significant positive correlations were noted between dose rate and serum concentrations of allopregnanolone (r = 0.94, P<0.001) and pregnanolone (r = 0.88, P<0.001). Such correlations were also seen in striatum, cerebellum, cortex and muscle for both steroids (P<0.01). Despite changing infusion rates, the concentrations of both steroids in brainstem, hippocampus and fat remained stable. Because no correlation between infusion rate and steroid concentration was noted in the brainstem and hippocampus, these two brain areas may be regarded as primary sites of action for allopregnanolone and pregnanolone. Pregnanolone concentrations in the brainstem and hippocampus were significantly higher than those of allopregnanolone, suggesting that allopregnanolone was more potent than pregnanolone in inducing anaesthesia.

Anesthetics, Intravenous↗
PubMed2001

Serotonin 5-HT(1A) receptor mRNA expression in dorsal hippocampus and raphe nuclei after gonadal hormone manipulation in female rats.

Female ovarian steroids influence mood and cognition, an effect presumably mediated by the serotonergic system. A key receptor in this interplay may be the 5-HT(1A) receptor subtype. We gave adult ovariectomized female rats subcutaneous pellets containing different dosages of 17 beta-estradiol alone or in combination with progesterone, or placebo pellets, for 2 weeks. 5-HT(1A) receptor mRNA levels were analyzed by in situ hybridization in the dorsal hippocampus, dorsal and median raphe nuclei, and entorhinal cortex. Estradiol treatment alone reduced 5-HT(1A) gene expression in the dentate gyrus and the CA2 region (17 and 19% decrease, respectively). Estradiol combined with progesterone supplementation increased 5-HT(1A) gene expression versus placebo in the CA1 and CA2 subregions of the dorsal hippocampus (16 and 30% increase, respectively). Concomitantly, 5-HT(1A) mRNA expression was decreased by 13% in the ventrolateral part of the dorsal raphe nuclei, while no changes were found in the median raphe nucleus and entorhinal cortex. Chronic effects of ovarian hormones on 5-HT(1A) receptor mRNA expression appear tissue-specific and involve hippocampal subregions and the raphe nuclei. Modulation of 5-HT(1A) receptor gene expression may be of importance for gonadal steroid effects on mood and cognition.

Animals↗
PubMed2000

gammadelta T-cell-deficient mice do not differ from normal controls in their induction or expression of type 2 dominant responses to exogenous antigen.

The roles that gammadelta T cells play in shaping initial CD4 T cell activation, and sensitivity to development of atopic diseases, remain controversial. Using a genetic knockout model of gammadelta T-cell deficiency, we investigated the role of these cells in initiation of exogenous antigen specific murine cytokine and antibody responses. Given that the most widely distributed and clinically prominent class of allergens are soluble protein antigens, we utilized OVA to examine the role played by gammadelta T cells in shaping the induction and expression of exogenous Ag specific immune responses. Focusing on immunization conditions that stimulate in vivo induction of type 2 dominant immunity, we report that gammadelta deficient and intact C57Bl6 mice exhibit similar OVA-specific responses as indicated by the (i) intensity of initial T-cell activation (ii) the type1 vs. type 2 balance of exogenous Ag specific cytokine synthesis and the (iii) intensity and the relative balance of the resulting IgE vs. IgG(2a) responses in vivo seen in these strains. Taken together, the data are consistent with the hypothesis that gammadelta T cells do not play an essential role in shaping induction of systemic immune responses to soluble exogenous antigen in type 2 dominated responses.

Animals↗
PubMed2000

Stretching of single collapsed DNA molecules.

The elastic response of single plasmid and lambda phage DNA molecules was probed using optical tweezers at concentrations of trivalent cations that provoked DNA condensation in bulk. For uncondensed plasmids, the persistence length, P, decreased with increasing spermidine concentration before reaching a limiting value 40 nm. When condensed plasmids were stretched, two types of behavior were observed: a stick-release pattern and a plateau at approximately 20 pN. These behaviors are attributed to unpacking from a condensed structure, such as coiled DNA. Similarly, condensing concentrations of hexaammine cobalt(III) (CoHex) and spermidine induced extensive changes in the low and high force elasticity of lambda DNA. The high force (5-15 pN) entropic elasticity showed worm-like chain (WLC) behavior, with P two- to fivefold lower than in low monovalent salt. At lower forces, a 14-pN plateau abruptly appeared. This corresponds to an intramolecular attraction of 0.083-0.33 kT/bp, consistent with osmotic stress measurements in bulk condensed DNA. The intramolecular attractive force with CoHex is larger than with spermidine, consistent with the greater efficiency with which CoHex condenses DNA in bulk. The transition from WLC behavior to condensation occurs at an extension about 85% of the contour length, permitting looping and nucleation of condensation. Approximately half as many base pairs are required to nucleate collapse in a stretched chain when CoHex is the condensing agent.

Bacteriophage lambda↗
PubMed2000

The inhibitory effects of allopregnanolone and pregnanolone on the population spike, evoked in the rat hippocampal CA1 stratum pyramidale in vitro, can be blocked selectively by epiallopregnanolone.

The progesterone metabolites allopregnanolone (Allo, 3alpha-hydroxy-5alpha-pregnan-20-one) and pregnanolone (Preg, 3alpha-hydroxy-5beta-pregnan-20-one) enhance the gamma-aminobutyric acid (GABA) action through a distinct site on the GABAA-receptor. Their 3beta-isomers epiallopregnanolone (Epiallo, 3beta-hydroxy-5alpha-pregnan-20-one) and epipregnanolone (Epipreg, 3beta-hydroxy-5beta-pregnan-20-one), do not have these effects on GABAA-receptors. We have studied the interaction between Allo/Preg and their 3beta-isomers on action potentials in rat hippocampal slices in vitro. The Schaffer collaterals were stimulated electrically in CA1 striatum radiatum and the population spike (POPSP) was recorded in stratum pyramidale. A 0.5-nL droplet of drug was applied locally onto stratum oriens-pyramidale via a pressure pipett. Muscimol (Mus) (12.5 fmol), Allo and Preg (6.25 fmol) caused a reversible inhibition of POPSP. On the other hand, 6.25 fmol Epiallo had no significant effect on POPSP compared with the vehicle control. Combined Epiallo and Allo application caused a dose-dependent reduction of the Allo inhibition of POPSP. A full blockage was seen at a molar ratio of 1:1. Epiallo also blocked the Preg inhibition of POPSP, when the two drugs were combined in a molar ratio of 1:1. Epiallo did not block the Mus inhibition of POPSP, when the two drugs were combined at a molar ratio of 1:2. Bath perfusion of 12.5 microM Epiallo blocked the inhibition of 6.25 fmol Allo on POPSP, but not the inhibition caused by 12.5 fmol Mus. Epipreg did not block the inhibition of Allo and Preg on POPSP, when it was combined with the two latter drugs at a molar ratio of 1:1. Our data suggest that the steroid modulation of the GABAA transmitted inhibition of the CA1 pyramidal neurones is selectively and dose dependently blocked by Epiallo, the 3beta-hydroxy-isomer of Allo, but not by Epipreg, the 3beta-hydroxy-isomer of Preg.

Action Potentials↗
PubMed2000

Failure of rIL-12 administration to inhibit established IgE responses in vivo is associated with enhanced IL-4 synthesis by non-B/non-T cells.

Administration of rIL-12 offers a widely successful tactic for preferential induction of type 1 immune responses in vivo. Its use to modulate ongoing cytokine or effector responses has proven to be substantially more difficult. Immediate hypersensitivity is the most common human immunologic disease. Here, rIL-12 was administered to C57Bl/6 and outbred CD1 mice with ongoing ovalbumin (OVA)-specific IgE responses in an attempt to redirect established type 2 cytokine and antibody production. Despite use of a broad range of treatment protocols for >4 months following initial immunization, recall IgE responses were consistently unaffected. rIL-12-treated mice exhibited strong in vivo and in vitro IFN-gamma responses, increased approximately 40-fold relative to controls, but also markedly enhanced (15- to 20-fold) OVA-specific IL-4 production. CD4 T cell function was successfully transformed from a type 2- to a type 1-dominated pattern following long-term IL-12 administration in vivo, as measured by strongly reduced IL-4 and IL-10 responses in antigen-stimulated primary culture, and 5-fold reductions in the frequencies of IL-4- and IL-10-producing OVA-specific CD4 T cells. However, chronically rIL-12-treated mice exhibited increased numbers of non-B/non-T cells that when re-stimulated with specific allergen, produce IL-4 at levels 20-fold higher than did CD4 T cells while IL-13 responses are unaffected. Collectively, the data indicate that even effectively shifting CD4 T cell activation from a type 2- to a type 1-dominated response does not in itself lead to altered effector (IgE) responses upon antigen re-exposure.

Animals↗
PubMed1999

IL-12 administered during Chlamydia psittaci lung infection in mice confers immediate and long-term protection and reduces macrophage inflammatory protein-2 level and neutrophil infiltration in lung tissue.

Protection against infections with the intracellular bacterium Chlamydia spp. requires Th1-polarized CD4+ T cell immunity. In BALB/c mouse lung infections, immediate innate and nascent Chlamydia-specific immune responses following intranasal inoculation of Chlamydia psittaci strain B577 were modulated by 7-day i.p. administration of murine rIL-12, the initiation cytokine for Th1 immunity. Treatment with IL-12 reduced the severity of chlamydial pneumonia, abolished mortality (37.5% in untreated mice), and significantly reduced numbers of chlamydial organisms in lungs. On day 4 after inoculation, the neutrophil:macrophage ratio in bronchointerstitial pneumonias was 1.96 in untreated mice and 0.51 in IL-12-treated mice. This immediate, IL-12-mediated shift in innate inflammatory phenotype was correlated with a significant reduction of lung concentrations of the neutrophil chemoattractant macrophage inflammatory protein (MIP)-2 (putative murine homologue of human IL-8), monocyte chemotactic protein-1, and TNF-alpha; and a reduction in MIP-1alpha and IFN-gamma, at high-dose infection only, and IL-12-independent IL-10 levels. Chlamydia-specific Ab titers and Ig isotype ratios indicated an IL-12-dependent Th1 shift. Recall responses of IL-12-primed mice to secondary chlamydial lung infection eliminated chlamydiae more effectively and generated a lung cytokine profile conducive to perpetuation of the Th1 memory population. These data support the hypothesis that genetic differences in endogenous IL-12 production and response pathways could determine disease outcomes characterized by poor chlamydial clearance and a purulent inflammatory infiltrate vs effective elimination of chlamydiae in a macrophage-dominated response.

Administration, Intranasal↗
PubMed1999

Estimating the persistence length of a worm-like chain molecule from force-extension measurements.

We describe a simple computation of the worm-like chain model and obtain the corresponding force-versus-extension curve. We propose an improvement to the Marko and Siggia interpolation formula of Bustamante et al (Science 1994, 265:1599-1600) that is useful for fitting experimental data. We apply it to the experimental elasticity curve of single DNA molecules. Finally, we present a tool to study the agreement between the worm-like chain model and experiments.

Biophysical Phenomena↗
PubMed1999

Manipulation of single molecules in biology.

The mechanical manipulation of single biological molecules is stimulating new and exciting research in many fields of study, including molecular motor mechanics, biopolymer properties, protein unfolding, receptor-ligand interactions, and more. Some recent highlights include the elucidation of the coupling ratios of myosin and kinesin, the demonstration of oscillatory forces in dynein arms, the determination of the force-velocity relation of RNA polymerase, and the direct mechanical observation of unfolding of single domains of titin and tenascin.

Biopolymers↗
PubMed1998

Force and velocity measured for single molecules of RNA polymerase.

RNA polymerase (RNAP) moves along DNA while carrying out transcription, acting as a molecular motor. Transcriptional velocities for single molecules of Escherichia coli RNAP were measured as progressively larger forces were applied by a feedback-controlled optical trap. The shapes of RNAP force-velocity curves are distinct from those of the motor enzymes myosin or kinesin, and indicate that biochemical steps limiting transcription rates at low loads do not generate movement. Modeling the data suggests that high loads may halt RNAP by promoting a structural change which moves all or part of the enzyme backwards through a comparatively large distance, corresponding to 5 to 10 base pairs. This contrasts with previous models that assumed force acts directly upon a single-base translocation step.

DNA, Bacterial↗
PubMed1998

Interaction between 3 alpha-hydroxy-5 alpha-pregnan-20-one and carbachol in the control of neuronal excitability in hippocampal slices of female rats in defined phases of the oestrus.

The effects of 3 alpha-hydroxy-5 alpha-pregnan-20-one (allopregnanolone) and carbachol on CA1 and dentate gyrus action potentials were studied in hippocampus slices in premature, follicular and luteal phase rats. A 0.5 nL droplet of allopregnanolone (12.5 mumol L-1), carbachol (5 mumol L-1) or a mixed solution of 12.5 mumol L-1 allopregnanolone and 5 mumol L-1 carbachol was applied locally onto the stratum oriens-pyramidale or granular layer. The amplitude of CA1 population spike (POPSP) was reduced by allopregnanolone (-38 +/- 3%) and carbachol (-21 +/- 4%) in the luteal phase slices. The mixture of allopregnanolone and carbachol doubled this inhibition (-77 +/- 6%). The inhibition caused by allopregnanolone and the mixture of allopregnanolone and carbachol in CA1 was significantly larger in the luteal phase than in the follicular phase (P = 0.02 and 0.0002). In the granular layer of the dentate gyrus, these inhibitions showed no significant difference between the phases. Neither in CA1 nor in the dentate gyrus did the carbachol inhibition differ between the phases. Perfusion with 5-10 mumol L-1 carbachol caused an increasing inhibition of the POPSP during the first few minutes. Thereafter the inhibition gradually diminished and was replaced by a facilitation. The local allopregnanolone inhibition was enhanced by simultaneous carbachol perfusion. Picrotoxin (100 mumol L-1) substantially reduced the allopregnanolone but not the carbachol inhibition. Atropine (10 mumol L-1) blocked the carbachol response, but not the allopregnanolone inhibition. Perfusion with a mixed solution of picrotoxin and atropine reduced, but did not block, the inhibition caused by local application of allopregnanolone or by the mixture of allopregnanolone and carbachol. Our data suggest that neuroprogestine modulators of the GABAA-receptor-mediated inhibition may play a significant role in the control of the cholinergic excitation in the hippocampus.

Action Potentials↗
PubMed1997

Immune suppression by lysosomotropic amines and cyclosporine on T-cell responses to minor and major histocompatibility antigens: does synergy exist?

BACKGROUND: Using murine models, we have shown that the lysosomotropic amine, chloroquine, is effective in the prevention of graft-versus-host disease (GVHD) mediated by donor T cells reactive with recipient minor histocompatibility antigens (MiHCs). Because lysosomotropic amines can suppress major histocompatibility complex (MHC) class II antigen presentation, their mechanism of action is potentially different from current immune suppressant drugs used to control GVHD such as cyclosporine. METHODS: We investigated the use of cyclosporine and the lysosomotropic amines chloroquine and hydroxychloroquine in combination for additive or synergistic immunosuppression on T-cell responses in vitro to MiHC and MHC in mice. RESULTS: We found that similar concentrations of chloroquine and hydroxychloroquine suppress the T-cell response to MiHC in mice (C57BL/6 anti-BALB.B) and that lysosomotropic amines in combination with cyclosporine result in synergistic suppression of a proliferative response to MiHC. Similar suppression and synergy appear to be present in an alloreactive response (C57BL/6 anti-BALB/c). Direct inhibition by chloroquine of T-cell proliferative responses induced by anti-CD3epsilon in the absence of antigen-presenting cells is present at higher concentrations than that required to suppress responses to MiHC or MHC. Chloroquine appears to induce decreased T-cell viability at high concentrations. This effect does not appear to be due to decreased T-cell production of interleukin-2 or interferon-gamma. At lower concentrations (<25 microg/ml), chloroquine can also decrease the ability of antigen-presenting cells to stimulate an a C57BL/6 anti-BALB/c T-cell response and can inhibit MHC class II expression after activation with lipopolysaccharide. CONCLUSIONS: Lysosomotropic amines in combination with cyclosporine appear to be synergistic in the suppression of T-cell proliferation to MiHC and MHC. Use of chloroquine in combination with cyclosporine may result in improved control of GVHD.

Animals↗
PubMed1997

Stretching DNA with optical tweezers.

Force-extension (F-x) relationships were measured for single molecules of DNA under a variety of buffer conditions, using an optical trapping interferometer modified to incorporate feedback control. One end of a single DNA molecule was fixed to a coverglass surface by means of a stalled RNA polymerase complex. The other end was linked to a microscopic bead, which was captured and held in an optical trap. The DNA was subsequently stretched by moving the coverglass with respect to the trap using a piezo-driven stage, while the position of the bead was recorded at nanometer-scale resolution. An electronic feedback circuit was activated to prevent bead movement beyond a preset clamping point by modulating the light intensity, altering the trap stiffness dynamically. This arrangement permits rapid determination of the F-x relationship for individual DNA molecules as short as -1 micron with unprecedented accuracy, subjected to both low (approximately 0.1 pN) and high (approximately 50 pN) loads: complete data sets are acquired in under a minute. Experimental F-x relationships were fit over much of their range by entropic elasticity theories based on worm-like chain models. Fits yielded a persistence length, Lp, of approximately 47 nm in a buffer containing 10 mM Na1. Multivalent cations, such as Mg2+ or spermidine 3+, reduced Lp to approximately 40 nm. Although multivalent ions shield most of the negative charges on the DNA backbone, they did not further reduce Lp significantly, suggesting that the intrinsic persistence length remains close to 40 nm. An elasticity theory incorporating both enthalpic and entropic contributions to stiffness fit the experimental results extremely well throughout the full range of extensions and returned an elastic modulus of approximately 1100 pN.

DNA↗
PubMed1997

The regional brain distribution of the neurosteroids pregnenolone and pregnenolone sulfate following intravenous infusion.

We have studied the distribution of the neurosteroids pregnenolone (Pe) and pregnenolone sulfate (PeS) in seven brain regions, and plasma and fat tissues in male adult rats following the intravenous infusion of 14 mg/kg Pe and 18 mg/kg PeS, respectively. After chromatographic separation of steroid sulfate esters and non-conjugated steroids by solid phase octadecyl C18 columns and celite column chromatographic separation of Pe from cross-reacted steroids, the concentrations of Pe and PeS were determined by radioimmunoassay. We found that both Pe and PeS concentrations were significantly increased in plasma, fat and brain compared to the vehicle controls after i.v. infusion of Pe and PeS. In the controls, Pe concentrations were highly correlated within brain regions and between fat and brain regions. Most correlations were lost after Pe and PeS infusions. The content of Pe and PeS was not uniformly distributed in the brain. The hypothalamus contained the highest level of Pe in controls, Pe-infused and PeS-infused rats (12 +/- 3.1, 3500 +/- 180 and 590 +/- 54 ng/g, respectively). The highest concentration of PeS was detected in the hypothalamus (26 +/- 8.2 ng/g) and striatum (17 +/- 4.1 ng/g) in controls, in the hypothalamus (200 +/- 24 ng/g) after PeS infusion as well as in the hypothalamus and medulla oblongata (57 +/- 9.6 and 55 +/- 7.6 ng/g, respectively) after Pe infusion. This study has yielded evidence that PeS injected i.v. can cross the blood-brain barrier without being hydrolysed to the more lipophilic Pe, and can thus be taken up by the brain.

Analysis of Variance↗