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Biomedical subjects

W Ma

Publications and source records attributed to W Ma.

At least 37 records · Page 2Linked to original sources
PubMed2003

Ethanol blocks both basic fibroblast growth factor- and carbachol-mediated neuroepithelial cell expansion with differential effects on carbachol-activated signaling pathways.

We have expanded neuroepithelial cells dissociated from the embryonic rat telencephalon in serum-free defined medium containing basic fibroblast growth factor (bFGF) in order to generate a model neuroepithelium to study the interaction of ethanol with both growth factor- and transmitter-stimulated proliferation. Ethanol blocked proliferation stimulated by bFGF and by carbachol, an agonist at muscarinic acetylcholine receptors, in a dose-dependent manner. In addition, ethanol attenuated autonomous expansion of neuroepithelial cells occurring following withdrawal of bFGF. The latter effect was associated with an increase in the number of apoptotic cells identified by terminal deoxynucleotidyltransferase-mediated dUTP nick end labeling labeling. We studied the effects of ethanol on carbachol-stimulated signaling pathways critical to its proliferative effects. Ethanol significantly reduced carbachol-stimulated Ca(2+) signaling, as well as Erk1/Erk2, Akt and cyclic AMP-response element-binding phosphorylations in a dose-dependent manner. Comparison of the potency of ethanol in attenuating carbachol-stimulated proliferation and signal transduction showed that mitogen-activated protein kinase phosphorylation was less sensitive to ethanol than the other parameters. The results indicate that ethanol's suppression of proliferation induced by carbachol in this model neuroepithelium likely involves multiple signaling pathways. These effects in vitro may help to explain the devastating effects of prenatal ethanol exposure in vivo, which contribute to the fetal alcohol syndrome.

Acetylcholine↗
PubMed2003

Localization and modulation of calcitonin gene-related peptide-receptor component protein-immunoreactive cells in the rat central and peripheral nervous systems.

Calcitonin gene-related peptide (CGRP) is widely distributed in the central and peripheral nervous system. Its highly diverse biological activities are mediated via the G protein-coupled receptor that uniquely requires two accessory proteins for optimal function. CGRP receptor component protein (RCP) is a coupling protein necessary for CGRP-receptor signaling. In this study, we established the anatomical distribution of RCP in the rat central and peripheral nervous system and its relationship to CGRP immunoreactivity. RCP-immunoreactive (IR) perikarya are widely and selectively distributed in the cerebral cortex, septal nuclei, hippocampus, various hypothalamic nuclei, amygdala, nucleus colliculus, periaqueductal gray, parabrachial nuclei, locus coeruleus, cochlear nuclei, dorsal raphe nuclei, the solitary tractus nucleus and gracile nucleus, cerebellar cortex, various brainstem motor nuclei, the spinal dorsal and ventral horns. A sub-population of neurons in the dorsal root ganglia (DRG) and trigeminal ganglia were strongly RCP-IR. Overall, the localization of RCP-IR closely matched with that of CGRP-IR. We also determined whether RCP in DRG and dorsal horn neurons can be modulated by CGRP receptor blockade and pain-related pathological stimuli. The intrathecal injection of the antagonist CGRP(8-37) markedly increased RCP expression in the lumbar DRG and spinal dorsal horn. Carrageenan-induced plantar inflammation produced a dramatic bilateral increase in RCP expression in the dorsal horn while a partial sciatic nerve ligation reduced RCP expression in the ipsilateral superficial dorsal horn. Our data suggest that the distribution of RCP immunoreactivity is closely matched with CGRP immunoreactivity in most of central and peripheral nervous systems. The co-localization of RCP and CGRP in motoneurons and primary sensory neurons suggests that CGRP has an autocrine or paracrine effect on these neurons. Moreover, our data also suggest that RCP expression in DRG and spinal cord can be modulated during CGRP receptor blockade, inflammation or neuropathic pain and this CGRP receptor-associated protein is dynamically regulated.

Animals↗
PubMed2003

Cyclooxygenase 2 in infiltrating inflammatory cells in injured nerve is universally up-regulated following various types of peripheral nerve injury.

We previously reported the up-regulation of cyclooxygenase 2 (COX2) in injured sciatic nerve of rats with partial sciatic nerve ligation (PSNL) and the reversal of PSNL-elicited tactile allodynia by local injection of the COX inhibitor ketorolac [Eur J Neurosci 15 (2002) 1037]. We further asked whether COX2 up-regulation in injured nerve is a universal phenomenon following various types of nerve injury. In the current study, we observed that abundant COX2 immunoreactive (IR) cell profiles appeared in injured nerves of rats following spinal nerve ligation (SNL), chronic constriction injury (CCI) and complete sciatic nerve transection. Most COX2-IR cells were identified as infiltrating macrophages. Partial injury induced greater COX2 up-regulation than complete injury. COX2 up-regulation reached a peak at 2-4 weeks, evidently declined by 3 months and disappeared by 7 months postlesion. These findings suggest that up-regulation of COX2 in injured nerve is a common event during the initial several months after nerve injury. We observed that local ketorolac-elicited anti-allodynia was closely associated with the abundance of COX2-IR cells in injured nerve, varying with the type of injury and time after injury. The anti-allodynia lasted the longest when local ketorolac was given 2-4 weeks after PSNL, CCI and SNL. The duration of local ketorolac's anti-allodynia was the longest in CCI rats, which also exhibited the most abundant COX2 up-regulation. Local ketorolac's anti-allodynia lasted much shorter when given 2-3 months after lesion. Local ketorolac failed to induce anti-allodynia 7 months after lesion, a time when COX2-IR cells completely disappeared from the injured nerve except a few cells at the injury site. Our data strongly suggest that during the initial several months after nerve injury, peripherally over-produced prostaglandins play an important role in the maintenance of neuropathic pain.

Animals↗
PubMed2003

Intraplantar injection of a cyclooxygenase inhibitor ketorolac reduces immunoreactivities of substance P, calcitonin gene-related peptide, and dynorphin in the dorsal horn of rats with nerve injury or inflammation.

We previously reported that partial sciatic nerve ligation (PSNL) dramatically up-regulates cyclooxygenase 2 (COX2) in injured sciatic nerve, and local injection of the COX inhibitor, ketorolac, reverses tactile allodynia and suppresses increased phosphorylation of the transcription factor cAMP responsive element binding protein [Eur J Neurosci 15 (2002) 1037]. These findings suggest that peripheral prostaglandins (PGs) are over-produced and contribute to the central plasticity and the maintenance of neuropathic pain after nerve injury. PGs, particularly PGE2, are well known to facilitate the release of the pro-nociceptive neuropeptide substance P (SP) and calcitonin gene-related peptide (CGRP) from primary sensory afferents. Thus, suppressing peripheral PG over-production may inhibit the release of these two neuropeptides from primary afferents and thereby increase the content of these neuropeptides remaining in afferent terminals in the dorsal horn. In this study we tested this hypothesis by examining the immunoreactivities of SP and CGRP in the dorsal horn of PSNL rats intraplantarly injected with saline and ketorolac. Four weeks after PSNL, SP- and CGRP-immunoreactivities (IR) in the ipsilateral dorsal horn were not significantly different from the contralateral side. Five days following intraplantar injection of ketorolac, CGRP- and SP-IR in the ipsilateral and contralateral dorsal horn were dramatically reduced compared with saline-injected PSNL rats. Local ketorolac also suppressed PSNL-induced increase in dynorphin-IR in dorsal horn neurons. Since abundant production of PGs during inflammation is well documented, we further examined the effect of intraplantar ketorolac on neuropeptide expression in the dorsal horn following carrageenan inflammation. We observed that co-administration of ketorolac with carrageenan in the hindpaw also reduced SP- and dynorphin-IR in the ipsilateral and contralateral dorsal horn. These findings are in contrast to our hypothesis, suggesting that peripherally over-produced PGs following nerve injury and inflammation possibly contribute to the production of SP and CGRP in primary sensory neurons, to the up-regulation of dynorphin in the dorsal horn neurons, and finally to the mechanisms of neuropathic and inflammation pain.

Animals↗
PubMed2003

Long-term growth arrest of PUVA-treated fibroblasts in G2/M in the absence of p16(INK4a) p21(CIP1) or p53.

Premature aging of the skin is a prominent side-effect of psoralen photoactivation, a therapy used for different skin disorders. Recently, we demonstrated that treatment of fibroblasts with 8-methoxypsoralen and ultraviolet A irradiation resulted in growth arrest with morphological and functional changes reminiscent of replicative senescence. To further elucidate the underlying molecular mechanisms, we analysed the cell-cycle phases of the growth-arrested fibroblasts. After PUVA treatment, fibroblasts arrested in G2/M, in contrast to spontaneously senesced fibroblasts arresting in a cell-cycle phase with many features similar to G1. To address the role of the cell-cycle controlling genes p16(INK4a), p21(CIP1) and p53, we analysed the expression of these genes. p16(INK4a), p21(CIP1) and p53 protein levels increased substantially with different time kinetics in growth-arrested fibroblasts. Because p16(INK4a), p21(CIP1) and p53 are involved in replicative senescence, we applied the PUVA regimen to fibroblasts deficient in either of these genes. p16(INK4a), p21(CIP1) and p53 null mutant fibroblast strains underwent growth arrest with a senescent phenotype similar to wild-type human fibroblasts. Based on these results, we propose that redundant or alternate pathways are involved in the response of dermal fibroblasts to PUVA treatment resulting in a phenocopy of replicative senescence in vitro.

Cell Division↗
PubMed2003

Localization of centromere proteins and their association with chromosomes and microtubules during meiotic maturation in pig oocytes.

Centromere proteins (CENPs) are required for the attachment of microtubules to chromosomes. However, their structure and mechanism of action are not well understood, especially in mammalian meiosis. The present study was conducted to examine (i). whether a human nuclear centromere autoantibody can be used to localize the CENPs in pig oocytes and (ii). the dynamics of CENPs and their association with microtubules and chromosomes during meiosis in pigs. Oocytes at various stages were double-labelled for CENPs, chromosomes or microtubules and examined by confocal fluorescence microscopy. Quantification of tubulin and CENPs in the oocytes was determined by immunoblotting. CENPs were detected in all oocytes from germinal vesicle (GV) to metaphase II (MII) stages. The changes in the location were associated with chromosome movement and spindle formation. Tubulin was detected in the oocytes from GV to MII stages and no differences in content were observed. Two major CENPs at 80 kDa (CENP-B) and 50 kDa (CENP-D) were also found in the oocytes by the autoantibody and its content was significantly lower in the oocytes at GV stage compared with oocytes at other stages. These results indicate that the autoantibody used in this study can be used to detect CENPs in the kinetochores, and the proteins are expressed in pig oocytes at all stages during meiosis. As the localization of CENPs is associated with spindle formation and chromosome movement, CENPs may participate in cell cycle changes during meiosis in mammals.

Analysis of Variance↗
PubMed2002

Acute exposure of toluene transiently potentiates p42/44 mitogen-activated protein kinase (MAPK) activity in cultured rat cortical astrocytes.

It has been shown that the inhalation of toluene in rats can cause neuronal apoptosis in the central nervous system. However, the cellular and molecular effects of toluene directly on astrocytes are relatively unknown. We used primary cultures of astrocytes isolated from the neonatal rat cortex as a model to study the toluene effects on cell outcome and associated signal transduction pathways using immunostaining and Western blotting. We observed that acute toluene exposure significantly induced caspase-dependent cell apoptosis and transiently stimulated the activation of p42/44 mitogen-activated protein kinase (MAPK) in the primary astrocytes. Interestingly, the inhibition of the p42/44 MAPK signaling cascade by PD98059 in conjunction with the toluene treatment evoked more cellular apoptosis than toluene alone, suggesting that the toluene-induced transient MAPK activation may play a role in promoting cell survival during the toluene exposure.

Animals↗
PubMed2002

X-linked Charcot-Marie-Tooth disease caused by a novel point mutation in the connexin-32 gene.

We report the clinical and electrophysiological findings of a patient with X-linked Charcot-Marie-Tooth disease and a novel point mutation in the connexin-32 gene. A 31-year-old man presented with a 5 year history of progressive imbalance and distal weakness in his legs. Electrophysiological studies confirmed an asymmetric, predominantly axonal sensorimotor neuropathy with some demyelinating features. Genetic testing revealed a G/A transition (Ala40Thr) in a conserved transmembrane region of the connexin-32 gene.

Adult↗
PubMed2002

Amyloid beta peptide induces tau phosphorylation and loss of cholinergic neurons in rat primary septal cultures.

The neuropathological features associated with Alzheimer's disease (AD) brain include the presence of extracellular neuritic plaques composed of amyloid beta protein (Abeta), intracellular neurofibrillary tangles containing phosphorylated tau protein and the loss of basal forebrain cholinergic neurons which innervate regions such as the hippocampus and the cortex. Studies of the pathological changes that characterize AD and several other lines of evidence indicate that Abeta accumulation in vivo may initiate phosphorylation of tau protein, which by disrupting neuronal network may trigger the process of neurodegeneration observed in AD brains. However, the underlying cause of degeneration of the basal forebrain cholinergic neurons and their association, if any, to Abeta peptides or phosphorylated tau remains mostly unknown. In the present study, using rat primary septal cultures, we have shown that aggregated Abeta peptides, in a time (18-96 h)- and concentration (0.7-60 microM)-dependent manner, induce toxicity and decrease choline acetyltransferase enzyme activity in cultured neurons. Using immunocytochemistry and immunoblotting, we have also demonstrated that Abeta treatment can significantly increase the phosphorylation of tau protein in septal cultures. At the cellular level, hyperphosphorylated tau is mostly apparent in the somatodendritic compartment of the neurons. Abeta peptide (10 microM), in addition to tau phosphorylation, also activates mitogen-activated protein kinase and glycogen synthase kinase-3beta, the two kinases which are known to be involved in the formation of hyperphosphorylated tau in the AD brain. Exposure to specific inhibitors of the mitogen-activated protein kinase (i.e. PD98059) or glycogen synthase kinase-3beta (i.e. LiCl) attenuated the hyperphosphorylation of the tau protein in cultured neurons. Given the evidence that tau phosphorylation can induce cell loss by disrupting neuronal cytoskeleton, it is likely that aggregated Abeta peptide triggers degeneration of septal neurons, including those expressing the cholinergic phenotype, by phosphorylation of the tau protein activated by mitogen-activated protein kinase and glycogen synthase kinase-3beta. These results, taken together, suggest that cultured septal cholinergic neurons are vulnerable to Abeta-mediated toxicity and tau phosphorylation may play an important role in Abeta-induced neurodegeneration.

Amyloid beta-Peptides↗
PubMed2002

Expression of calcitonin gene-related peptide, substance P and protein kinase C in cultured dorsal root ganglion neurons following chronic exposure to mu, delta and kappa opiates.

The mechanisms involved in morphine tolerance are poorly understood. It was reported by our group that calcitonin gene-related peptide (CGRP)-like immunoreactivity (IR) was increased in the spinal dorsal horn during morphine tolerance [Ménard et al. (1996) J. Neurosci. 16, 2342-2351]. More recently, we observed that it was possible to mimic these results in cultured dorsal root ganglion (DRG) neurons allowing for more detailed mechanistic studies [Ma et al. (2000) Neuroscience 99, 529-539]. The aim of the present series of experiments was to further validate the DRG cell culture model by establishing which subtypes of opioid receptors are involved in the induction of CGRP in cultured rat DRG neurons, and to examine the signaling pathway possibly involved in the induction of CGRP-like IR following repeated opiate treatments. Other neuropeptides known to be expressed in DRG neurons, such as substance P (SP), neuropeptide Y (NPY) and galanin, were investigated to assess specificity. Following treatment with any of the three opioid agonists (mu, DAMGO; delta, DPDPE; kappa, U50488H), the number of CGRP- and SP-IR cultured DRG neurons increased significantly, and in a concentration-dependent manner, with the effects of kappa agonist being less pronounced. NPY and galanin were not affected.Double-immunofluorescence staining showed that the three opioid receptors were co-localized with both CGRP- and SP-like IR.Protein kinase C (PKC)-like IR was found to be significantly increased following a repetitive treatment with DAMGO. Double-immunofluorescence staining showed the co-localization of PKCalpha with CGRP- and SP-IR in cultured DRG neurons. Moreover, a combined treatment with DAMGO and a PKC inhibitor (chelerythrine chloride or Gö 6976) was able to block the effects of the opioid on increased CGRP-like IR. These data suggest that the three opioid receptors may be involved in the induction of CGRP and SP observed following chronic exposure to opiates, and that PKC probably plays a role in the signaling pathway leading to the up-regulation of these neuropeptides. These findings further validate the DRG cell culture as a suitable model to study intracellular pathways that govern changes seen following repeated opioid treatments possibly leading to opioid tolerance.

3,4-Dichloro-N-methyl-N-(2-(1-pyrrolidinyl)-cycloh↗
PubMed2002

Treatment of antibiotics wastewater utilizing successive hydrolysis, denitrification and nitrification.

A process consisting of anaerobic hydrolysis, denitrification, and oxidation/nitrification was proposed for simultaneous removal of carbon and nitrogen from terramycin crystallizing mother solution (TCMS), and its performance was investigated by treating diluted TCMS in a lab-scale continuous flow column system. Direct denitrification-nitrification of diluted TCMS produced a significant residual of nitrate and nitrite, which disappeared 44 days after startup of anaerobic hydrolysis column. The electron donors available to denitrification were increased by 6 times after diluted TCMS was treated in the hydrolysis column under an HRT of 2.5 h or longer. The reaction rates of organics decomposition, nitrification, and denitrification were also significantly increased due to reduction of terramycin and decomposition of complicated molecules to small molecules during anaerobic hydrolysis. The specific denitrification rate and nitrification rate increased from 0.033 d(-1) and 0.01 d(-1) to 0.045 d(-1) and 0.021 d(-1) respectively after diluted TCMS (dilution ratio: 1:4) was hydrolyzed in anaerobic hydrolysis column at a hydraulic retention time (HRT) of 4 h.

Anti-Bacterial Agents↗
PubMed2001

Alpha-lipoic acid protects rat cortical neurons against cell death induced by amyloid and hydrogen peroxide through the Akt signalling pathway.

Substantial evidence suggests that the accumulation of beta-amyloid (Abeta)-derived peptides contributes to the aetiology of Alzheimer's disease (AD) by stimulating formation of free radicals. Thus, the antioxidant alpha-lipoate, which is able to cross the blood-brain barrier, would seem an ideal substance in the treatment of AD. We have investigated the potential effectiveness of alpha-lipoic acid (LA) against cytotoxicity induced by Abeta peptide (31-35) (30 microM) and hydrogen peroxide (H(2)O(2)) (100 microM) with the cellular 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-tetrazolium bromide (MTT) reduction and fluorescence dye propidium iodide assays in primary neurons of rat cerebral cortex. We found that treatment with LA protected cortical neurons against cytotoxicity induced by Abeta or H(2)O(2). In addition, LA-induced increase in the level of Akt in the neurons was observed by Western blot. The LA-induced neuroprotection and Akt increase were attenuated by pre-treatment with the phosphatidylinositol 3-kinase inhibitor, LY294002 (50 microM). Our data suggest that the neuroprotective effects of the antioxidant LA are partly mediated through activation of the PKB/Akt signaling pathway.

Alzheimer Disease↗
PubMed2001

Regulation of NMDA receptors by cyclin-dependent kinase-5.

Members of the N-methyl-d-aspartate (NMDA) class of glutamate receptors (NMDARs) are critical for development, synaptic transmission, learning and memory; they are targets of pathological disorders in the central nervous system. NMDARs are phosphorylated by both serine/threonine and tyrosine kinases. Here, we demonstrate that cyclin dependent kinase-5 (Cdk5) associates with and phosphorylates NR2A subunits at Ser-1232 in vitro and in intact cells. Moreover, we show that roscovitine, a selective Cdk5 inhibitor, blocks both long-term potentiation induction and NMDA-evoked currents in rat CA1 hippocampal neurons. These results suggest that Cdk5 plays a key role in synaptic transmission and plasticity through its up-regulation of NMDARs.

Animals↗
PubMed2001

Identification of a prostate-specific G-protein coupled receptor in prostate cancer.

Membrane receptors coupled to heterotrimeric G-proteins play an essential role in the transmission of signals from the extracellular environment to the cytoplasm of the cell. A wide variety of external stimuli, including neurotransmitters, hormones, phospholipids, photons, odorants, taste ligands, and growth factors, can activate specific members of the G-protein coupled receptors (GPCRs). Besides essential functions in fully differentiated cells and tissues, GPCRs are also involved in embryogenesis, tissue regeneration, cell growth stimulation, and cell proliferation. In this study, we identified a novel prostate-specific G-protein coupled receptor that interacts with Galpha(12) in our yeast two-hybrid assays. The expression of the receptor protein is highly restricted to human prostate tissues using multiple-tissue Northern blot analysis, and tissue expression array. Furthermore, the expression of prostate-specific receptor is increased significantly in prostate tumors in comparison with the matched normal prostate tissues using PCR and Southern blot analysis, suggesting a potential role of this tissue-specific G-protein coupled receptor in prostate cancer development.

Amino Acid Sequence↗
PubMed2001

Analysis of trace amino acid neurotransmitters in hypothalamus of rats after exhausting exercise using microdialysis.

A simple but effective coupling of microdialysis and capillary electrophoresis with laser induced fluorescence detection technique was applied to analysis of amino acid neurotransmitters in the hypothalamus of rats after acute exhausting exercise. The separation of amino acids was achieved using an uncoated fused-silica capillary (57 cm x 75 microm I.D.) with a buffer of 10 mM disodium tetraborate at pH 10 and an applied voltage of 12.5 kV. The detection limit was 10(-10) M for each amino acid. It is sufficiently sensitive and rapid for the determination of amino acids in a 5-microl Microdialysate. In comparison to pre-exercise, a significant increase in the levels of six hypothalamic amino acids (arginine, glycine, lysine, glutamic acid, alanine, gamma-amino-n-butyric acid) was found after exercise. These results demonstrate that the increase of metabolic amino acids in the hypothalamus of rats can be induced by exhausting exercise and suggests that amino acid neurotransmitters may play functional roles in the central effects of exercise.

Amino Acids↗
PubMed2001

Estrogen protects against beta-amyloid-induced neurotoxicity in rat hippocampal neurons by activation of Akt.

The cellular mechanisms underlying the neuroprotective effects of estrogen are only beginning to be elucidated. Here we examined the role of protein kinase B (Akt) activation in 17beta-estradiol (E2) inhibition of beta-amyloid peptide (31-35) (Abeta31-35)-induced neurotoxicity in cultured rat hippocampal neurons. Abeta31-35 (25-30 betaM) significantly decreased the total number of microtubule associated protein-2 positive cells (MAP2+). This decrease was significantly reversed by pre-treatment with 100 nM E2. Further, 100 nM E2 alone significantly increased the total number of protein kinase B and microtubule associated protein-2 positive cells compared with controls. Such E2-induced increases were inhibited by LY294002 (20 microM), a specific PI3-K inhibitor, as well as by tamoxifen, an estrogen receptor antagonist/selective estrogen receptor modulator. These results indicate that the neuroprotective effects of E2 may be mediated at least in part via estrogen receptor-mediated protein kinase B activation.

Alzheimer Disease↗
PubMed2001

Gene expression analysis of an H(2)O(2)-resistant lens epithelial cell line.

Gene expression patterns were examined in lens epithelial cells conditioned to grow in 125 microM hydrogen peroxide in order to define the protective mechanisms that may be involved in survival during oxidative stress. RNA was extracted from normal and hydrogen peroxide-resistant alphaTN4 mouse lens epithelial cells. Gene expression was evaluated using Differential Display (DD) and RT-PCR. Upregulation of mRNAs for antioxidant and cellular defense enzymes was observed. The highest elevation detected was a 14-fold increase in catalase in the hydrogen peroxide-resistant cells. Glutathione peroxidase, ferritin, and alphaB-crystallin were upregulated 2-fold, and reticulocalbin was upregulated 6-fold in the resistant cells. alphaA-crystallin was downregulated 5-fold, while aldose reductase and mitochondrial gene products were unchanged. Thus, in the alphaTN4 mouse lens cell line, long-term exposure to high levels of hydrogen peroxide elicited an upregulation of transcripts for enzymes involved in hydrogen peroxide degradation, metal binding, and chaperone function. Since mitochondrial gene transcription is sensitive to hydrogen peroxide, the presence of normal levels of mitochondrial transcripts, in this study, demonstrates the effectiveness of the antioxidant defense systems.

Animals↗