Biomedical subjects

Ming Fan

Publications and source records attributed to Ming Fan.

At least 37 records · Page 2Linked to original sources

PubMed2005

Co-activation of ERK, NF-kappaB, and GADD45beta in response to ionizing radiation.

NF-kappaB has been well documented to play a critical role in signaling cell stress reactions. The extracellular signal-regulated kinase (ERK) regulates cell proliferation and survival. GADD45beta is a primary cell cycle element responsive to NF-kappaB activation in anti-apoptotic responses. The present study provides evidence demonstrating that NK-kappaB, ERK and GADD45beta are co-activated by ionizing radiation (IR) in a pattern of mutually dependence to increase cell survival. Stress conditions generated in human breast cancer MCF-7 cells by the administration of a single exposure of 5 Gy IR resulted in the activation of ERK but not p38 or JNK, along with an enhancement of the NF-kappaB transactivation and GADD45beta expression. Overexpression of dominant negative Erk (DN-Erk) or pre-exposure to ERK inhibitor PD98059 inhibited NF-kappaB. Transfection of dominant negative mutant IkappaB that blocks NF-kappaB nuclear translocation, inhibited ERK activity and GADD45beta expression and increased cell radiosensitivity. Interaction of p65 and ERK was visualized in living MCF-7 cells by bimolecular fluorescence complementation analysis. Antisense inhibition of GADD45beta strikingly blocked IR-induced NF-kappaB and ERK but not p38 and JNK. Overall, these results demonstrate a possibility that NF-kappaB, ERK, and GADD45beta are able to coordinate in a loop-like signaling network to defend cells against the cytotoxicity induced by ionizing radiation.

Active Transport, Cell Nucleus↗

PubMed2005

Modeling Pb sorption to microporous amorphous oxides as discrete particles and coatings.

Hydrous amorphous Al (HAO), Fe (HFO), and Mn (HMO) oxides are ubiquitous in the subsurface as both discrete particles and coatings and exhibit a high affinity for heavy metal contaminants. To assess risks associated with heavy metals, such as Pb, to the surrounding environment and manage remedial activities requires accurate mechanistic models with well-defined transport parameters that represent sorption processes. Experiments were conducted to evaluate Pb sorption to microporous Al, Fe, and Mn oxides, as well as to montmorillonite and HAO-coated montmorillonite. Intraparticle diffusion, a natural attenuating process, was observed to be the rate-limiting mechanism in the sorption process, where best-fit surface diffusivities ranged from 10(-18) to 10(-15) cm(2) s(-1). Specifically, diffusivities of Pb sorption to discrete aluminum oxide, aluminum oxide-coated montmorillonite, and montmorillonite indicated substrate surface characteristics influence metal mobility where diffusivity increased as affinity decreased. Furthermore, the diffusivity for aluminum oxide-coated montmorillonite was consistent with the concentrations of the individual minerals present and their associated particle size distributions. These results suggest that diffusivities for other coated systems can be predicted, and that oxide coatings and montmorillonite are effective sinks for heavy metal ions.

Journal Article↗

PubMed2005

Underlying mechanism of hypoxic preconditioning decreasing apoptosis induced by anoxia in cultured hippocampal neurons.

It is known that hypoxic preconditioning (HP, a brief period of sublethal hypoxia) provides neuroprotection against subsequent severe anoxia, but the mechanisms of this increased tolerance have not been fully elucidated. A hypoxic preconditioning model was established by exposing a 4-day hippocampal culture to 1% O(2) for 20 min/day for 8 days. The preconditioning significantly decreased the number of apoptotic neurons at reoxygenation 24 h after 4 h of severe anoxia (0% O(2)). Further study demonstrated that the degradation of mitochondrial membrane potential (MMP) was greatly inhibited and the expression of B-cell lymphoma protein-2 (Bcl-2) was increased considerably after severe anoxia in the HP groups. These results indicate that the increased anoxic tolerance, which is induced by HP in cultured hippocampal cells, may be correlated with Bcl-2 overexpression and enhanced stability of MMP, which ultimately reduces apoptosis 24 h after reoxygenation.

Animals↗

PubMed2005

Effects of hypoxia on the proliferation and differentiation of NSCs.

Oxygen is vital to nearly all forms of life on Earth via its role in energy homeostasis and other cell functions. Until recently, the effects of oxygen on the proliferation and differentiation of neural stem cells (NSCs) have been largely ignored. Some studies have been carried out on the basis of the fact that NSCs exists within a "physiological hypoxic" environment at 1 to 5% O2 in both embryonic and adult brains. The results showed that hypoxia could promote the growth of NSCs and maintain its survival in vitro. In vivo studies also showed that ischemia/hypoxia increased the number of endogenous NSCs in the subventricular zone and dentate gyrus. In addition, hypoxia could influence the differentiation of NSCs. More neurons, especially more doparminergic neurons, were produced under hypoxic condition. The effects of hypoxia on the other kind of stem cell were briefly introduced as additional evidence. The mechanism of these responses might be primarily involved in the hypoxic inducible factor-1 (HIF-1) signal pathway. The present review summarizes recent works on the role of hypoxia in the proliferation and differentiation of NSCs both in vitro and in vivo, and the mechanism involved in HIF-1 signaling pathway behind this response was also discussed.

Animals↗

PubMed2005

[Repair of peripheral nerve gap with the use of tissue engineering scaffold complex].

OBJECTIVE: To study the feasibility to repair the peripheral nerve gap with tissue engineering scaffold complex that is composed of medical biodegradable material agarose hydrogel and nerve growth factor (NGF). METHODS: Chitosan tube containing agarose hydrogel and NGF was transplanted to bridge a 10 mm gap of injured sciatic nerve in rat. Chitosan duct without agarose hydrogel and NGF was used as negative control, while autograft nerve as positive control. Sixteen weeks after operation, the regeneration of nerve fiber was observed with morphological and immunohistochemistrical methods. RESULT: The number and diameter of regenerating nerve fibers bridged by the scaffold complex of agarose hydrogel and NGF were better than negative control group (P < 0.01) and reached the level of autograft nerve group. CONCLUSIONS: The new type of tissue engineering scaffold complex of agarose hydrogel and NGF may provide a microenvironment, as well as autograft nerve, to promote nerve regeneration. This technique may benefit patients with nerve injury in the future.

Absorbable Implants↗

PubMed2004

Crystallographic analysis of murine constitutive androstane receptor ligand-binding domain complexed with 5alpha-androst-16-en-3alpha-ol.

The constitutive androstane receptor (CAR) is a member of the nuclear receptor superfamily. In contrast to classical nuclear receptors, which possess small-molecule ligand-inducible activity, CAR exhibits constitutive transcriptional activity in the apparent absence of ligand. CAR is among the most important transcription factors; it coordinately regulates the expression of microsomal cytochrome P450 genes and other drug-metabolizing enzymes. The murine CAR ligand-binding domain (LBD) was coexpressed with the steroid receptor coactivator protein (SRC-1) receptor-interacting domain (RID) in Escherichia coli. The mCAR LBD subunit was purified away from SRC-1 by affinity, anion-exchange and size-exclusion chromatography, crystallized with androstenol and the structure of the complex determined by molecular replacement.

Androstenols↗

PubMed2004

Identification of a novel alternative splicing form of human netrin-4 and analyzing the expression patterns in adult rat brain.

Netrins are a family of secreted proteins that function as tropic cues directing axon growth and cell migration during neural development. Beta-netrin or netrin-4 (NTN4), the fourth member of this family, was previously reported by Koch (M. Koch, J.R. Murrell, D.D. Hunter, P.F. Olson, W. Jin, D.R. Keene, W.J. Brunken, R.E. Burgeson. A novel member of the netrin family, beta-netrin, shares homology with the beta chain of laminin: identification, expression, and functional characterization. J. Cell Biol. 151 (2000) 221-234) and Yin (Y. Yin, J.R. Sanes, J.H. Miner, Identification and expression of mouse netrin-4. Mech. Dev. 96 (2000) 115-119), respectively. However, whether there is another isoform of netrin-4 and the expression patterns have not been fully determined. Here we report the cloning of a novel isoform of human netrin-4 using rapid amplification of complementary DNA (cDNA) ends (RACE) technique and analysis on the netrin-4 mRNA expression in adult rat brain using in situ hybridization. The new isoform lacks the signal peptide region and might represent a truncated form at the Laminin-N terminal domain of human netrin-4. Examination of the expression pattern of netrin-4 mRNA in adult rat brain revealed that the netrin-4 mRNA was widely expressed in many regions of the brain. High levels of netrin-4 mRNA was found in the pyramidal cell layer of the cerebral cortex, prepiriform cortex, amygdaloid nuclei, pyramidal layer of hippocampus, Purkinje's cells, medial cerebellar nucleus and interposed cerebellar nucleus, medial nucleus of the trapezoid body and mitral cell layer of the olfactory bulb. Moderate signals were present in frontal cortex, several thalamic and hypothalamic nuclei. The widespread expression of netrin-4 in adult rat brain indicates that netrin-4 is not only important for axon guidance during embryonic development, but also critical to neuronal plasticity in adult stage.

Alternative Splicing↗

PubMed2004

Screening of functional antidotes of RNA aptamers against bovine thrombin.

A specific RNA aptamer (T705) against bovine thrombin had been obtained after seven rounds of SELEX (systematic evolution of ligands by exponential enrichment) selection from a random RNA library previously. In order to further investigate the relationship between the structure and function of this aptamer, three truncated RNA aptamers, T705a, T705b and T705c, were designed according to the secondary structure of T705 RNA. Our results showed that T705c keeping the precise stem-loop structure but lacking most of the stem region sequence of T705 could inhibit clot formation in vitro in the same way as its parental form. We also report here that single-stranded DNA (ssDNA) antisense oligonucleotides, c' and c'-22, which were complementary to different portions of T705c could act as efficient antidotes reversing the inhibitory activity of T705. It is demonstrated for the first time that ssDNA antisense oligonucleotides are potential antidotes of RNA aptamers and this may be an effective, rapid strategy to find antidotes of RNA aptamers which would be of important usefulness in basic research and drug screening.

Animals↗

PubMed2004

Involvement of increased stability of mitochondrial membrane potential and overexpression of Bcl-2 in enhanced anoxic tolerance induced by hypoxic preconditioning in cultured hypothalamic neurons.

The effects of hypoxic preconditioning (HP) on changes in mitochondrial membrane potential (MMP) and Bcl-2 expression in cultured hypothalamic neurons after severe anoxia were investigated. In the HP group, hypothalamic neurons, after a 4-day culture, were preconditioned daily under a hypoxic condition (1% O(2), 10 min) for 8 days; subsequently, the HP neurons and those in the control group (similarly cultured, but without HP) were exposed to 6 h of severe anoxia (0% O(2)). The preconditioned neurons had a higher survival rate and a lower lactate dehydrogenase leakage, compared with the control group. Although HP did not prevent the degradation of MMP during severe hypoxia, preconditioned neurons exhibited a higher level of MMP than that of the control group. Increased expression of Bcl-2 was also observed in the preconditioned hypothalamic neurons. These results suggest that HP enhances the hypoxic tolerance of hypothalamic neurons, and the underlying mechanisms may be related to the increased stability of MMP and the overexpression of Bcl-2 induced by HP.

Animals↗

PubMed2004

Prokaryotic expression, polyclonal antibody preparation, and sub-cellular localization analysis of Na+, K+-ATPase beta2 subunit.

Na+, K+-ATPase beta2 subunit (NKA1b2) is not only a regulator of Na+, K+-ATPase, but also functions in the interaction between neuron and glia cells as a Ca2+-dependent adhesion molecule. To further study the function of NKA1b2, the anti-NKA1b2 polyclonal antibody was prepared to recognize the outer-membrane carboxyl portion segment of NKA1b2. The coding region for amino acids 190-290 at the carboxyl portion of NKA1b2 (NKA1b2-CP) was sub-cloned into the vector pGEX-4T-2 and introduced into the Escherichia coli BL21(DE3) cell for efficient soluble expression. The amino acid sequence of expressed protein was determined using mass spectrometry following Mascot analysis. After purification, GST-NKA-beta2-CP was used to immunize the adult rabbits following standard protocols. The produced antiserum could detect the NKA1b2 protein expressed not only in the prokaryotic cells (E. coli) but also in the eukaryotic cells (COS7) transfected with NKA1b2 expression vector (pEGFP-NKA1b2). Furthermore, the antiserum was used for determining the localization of NKA1b2 in primary culture of neonatal rat neurons using immunohistochemical technique. Results demonstrated that NKA1b2 was localized both in the cytoplasm and cellular membrane. The preparation of anti-NKA-beta2-CP polyclonal antibody will facilitate further functional study on NKA1b2.

Amino Acid Sequence↗

PubMed2004

Comparison of the time to onset of action on myocardial ischaemia following intravenous administration of isosorbide dinitrate and 5-isosorbide mononitrate in Chinese patients.

OBJECTIVE: The onset of action of intravenous isosorbide dinitrate (CAS 87-33-2, ISDN) and intravenous 5-isosorbide mononitrate (CAS 16051-77-7, 5-ISMN) were compared by measurement of the indicators of perfusion to ischaemic myocardium. METHODS: Twenty-five patients with coronary heart disease were randomly allocated to receive intravenous ISDN or 5-ISMN. The extent of myocardial ischaemia before infusion and at 3, 15 and 45 min after commencement of infusion was evaluated using 99mTc-MIBI myocardium tomography imaging and electrocardiograms. RESULTS: The perfusion defects were significantly reduced or resolved in 11 patients (84.1%) receiving ISDN and 2 patients (15.38%) receiving 5-ISMN at 3 min. At 15 min the improvement was significantly greater in the ISDN group than in the 5-ISMN group. The improvements of 99mTc-MIBI myocardial uptake ratio and electrocardiograms were statistically significant in the ISDN group at 3 min and 15 min compared to pre-infusion. Although a significant improvement appeared at 15 min in the 5-ISMN group, it was significantly less than that observed in the ISDN group (p < 0.05). After 45 min, there were improvements in ischaemia in both groups with the difference compared to pre-infusion being significant, and there was no statistically significant difference between the ISDN and 5-ISMN group. CONCLUSION: In patients with coronary heart disease with ischaemic episodes the onset of therapeutic activity was more rapid with intravenous ISDN compared to 5-ISMN. ISDN should be the preferred intravenous nitrate for acute ischaemic episodes where a rapid onset of therapeutic action is desired.

Acute Disease↗

PubMed2004

The cloning and preliminarily functional analysis of the human neurotrimin gene.

Proteins of the immunoglobulin superfamily (IgSF) are involved in a variety of specific cell-cell interactions in the developing nervous system. To identify and characterize new members of this protein family in human nervous system, we screen the human fetal brain cDNA library and isolate a full-length cDNA clone which contains a 1032 bp open reading frame encoding a protein of 344 amino acids. Sequence analysis reveals that it is a glycoprotein comprised of three C2-like immunoglobulin domains and is anchored to the plasma membrane via a post-translationally attached glycosyl-phosphatidylinositol (GPI) moiety. The protein shows high sequence similarity to the rat Ntm (97%), so we term it human neurotrimin (NTM). Northern blot analysis reveals that (HUMAN)NTM has three different transcripts with the length of 3.2 kb, 4.0 kb and 9.0 kb respectively. It has a wider expression pattern than that of (RAT) Ntm. Notably, the expression of NTM in fetal brain is higher than that in mature brain and is stronger in nervous tumors than that in normal brain tissues. We insert an HA epitope tag between the third Ig-like domain of NTM and the site of GPI attachment, then construct it into the eukaryotic expression vector pcDNA3.1+/Zeocin. The pcDNA3.1-HA-NTM is transfected into the Chinese Hamster Ovary (CHO) cells. The results demonstrate that HA-NTM is expressed on the surface of CHO cells and could strengthen the aggregation of CHO-NTM cells.

Amino Acid Sequence↗

PubMed2003

[Glutamate-neuroexcitotoxicity protective study of monoclonal antibody against human NMDA receptor key subunit].

OBJECTIVE: To study if monoclonal antibody against human NMDA receptor key subunit (NR1) may protect neurons from excitotoxicity. METHODS: We cultured primary hippocampal neurons from 10 newborn rats and made the glutamate excitotoxicity model, examined the ratio of surviving neuron (Trypan blue dye staining) and LDH assay to study the protective effects of mAbN1, There were 2 plates in parallel per group, the experiments were repeated 4 times. RESULTS: 0.3 micro mol/L mAbN1 could protect cultured hippocampal neurons from neuroexcitotoxicity induced by 500 micro mol/L glutamate with increased survival rate 35%. Epitope peptide could block the protection of mAbN1, which suggests that the protection of mAbN1 is specific. CONCLUSION: mAbN1 is a novel NMDAR blocker with in vitro neuroprotective activity which may provide a basis for antibody therapy of neuroexcitotoxicity.

Animals↗

PubMed2003

[Exogenous rhCNTF inhibits myoblast differentiation of skeletal muscle of adult human in vitro].

To determine the role of recombinant human ciliary neurotrophic factor (rhCNTF) in myogenesis, we observed the effects of rhCNTF (0 10 ng/ml) on myoblast differentiation of adult human in vitro. The results showed that compared with the control group, the groups of 2.5 10 ng/ml rhCNTF treatment significantly inhibited myoblast differentiation (P<0.01), and the inhibition was dose-dependent and reversible. Western blots also indicated that compared with the control group, the expression of myogenin and p21, markers of myoblast differentiation phase, was significantly reduced (P<0.01), while Myf5 and desmin, markers of myoblast proliferative phase, significantly increased (P<0.01) in the groups of 2.5 10 ng/ml rhCNTF treatment. These findings demonstrate that exogenous rhCNTF can reversibly inhibit differentiation but permits proliferation of adult human myoblasts in vitro.

Adult↗

PubMed2003

SamCluster: an integrated scheme for automatic discovery of sample classes using gene expression profile.

MOTIVATION: Feature (gene) selection can dramatically improve the accuracy of gene expression profile based sample class prediction. Many statistical methods for feature (gene) selection such as stepwise optimization and Monte Carlo simulation have been developed for tissue sample classification. In contrast to class prediction, few statistical and computational methods for feature selection have been applied to clustering algorithms for pattern discovery. RESULTS: An integrated scheme and corresponding program SamCluster for automatic discovery of sample classes based on gene expression profile is presented in this report. The scheme incorporates the feature selection algorithms based on the calculation of CV (coefficient of variation) and t-test into hierarchical clustering and proceeds as follows. At first, the genes with their CV greater than the pre-specified threshold are selected for cluster analysis, which results in two putative sample classes. Then, significantly differentially expressed genes in the two putative sample classes with p-values < or = 0.01, 0.05, or 0.1 from t-test are selected for further cluster analysis. The above processes were iterated until the two stable sample classes were found. Finally, the consensus sample classes are constructed from the putative classes that are derived from the different CV thresholds, and the best putative sample classes that have the minimum distance between the consensus classes and the putative classes are identified. To evaluate the performance of the feature selection for cluster analysis, the proposed scheme was applied to four expression datasets COLON, LEUKEMIA72, LEUKEMIA38, and OVARIAN. The results show that there are only 5, 1, 0, and 0 samples that have been misclassified, respectively. We conclude that the proposed scheme, SamCluster, is an efficient method for discovery of sample classes using gene expression profile. AVAILABILITY: The related program SamCluster is available upon request or from the web page http://www.sph.uth.tmc.edu:8052/hgc/Downloads.asp.

Algorithms↗

PubMed2003

Single-stranded DNA aptamers that bind differentiated but not parental cells: subtractive systematic evolution of ligands by exponential enrichment.

In this paper, single-stranded (ss)DNA aptamers with capability to distinguish differentiated PC12 cells from normal PC12 cells were selected by subtractive systematic evolution of ligands by exponential enrichment (SELEX) method. Before each round of selection, randomized ssDNAs were incubated with regular PC12 cells to eliminate those that recognize the common cellular components of both differentiated and undifferentiated PC12 cells. After six rounds of cell-based selection, both of individual aptamers and aptamers of the sixth round pool were found binding to differentiated PC12 cells, but not to the parental PC12 cells. The aptamers of the starting pool showed no such binding. Sequence analysis illustrated that the amount of G content in central random region of these aptamers was much higher than that of the starting pool, which would be expected to be average. The aptamers obtained from this method were also able to identify differentiated PC12 cells from a mixture of both normal and differentiated cells. The results indicate that subtractive SELEX is a useful tool in finding ligands to specific biological markers that distinguish a subtype of cells from cells of homologous origin, such as carcinoma cells among normal epithelial tissues. Both these aptamers and their markers may play important roles in basic research and clinical diagnosis.

Animals↗

PubMed2003

Identification of an endogenous ligand that activates pregnane X receptor-mediated sterol clearance.

The nuclear receptor PXR (pregnane X receptor) is a broad-specificity sensor that recognizes a wide variety of synthetic drugs and xenobiotic agents. On activation by these compounds, PXR coordinately induces a network of transporters, cytochrome P450 enzymes, and other genes that effectively clear xenobiotics from the liver and intestine. Like PXR, the majority of its target genes also possess a broad specificity for exogenous compounds. Thus, PXR is both a sensor and effector in a well integrated and generalized pathway for chemical immunity. Although it is clear that PXR responds to numerous foreign compounds, it is unclear whether it possesses an endogenous ligand. To address this issue, we noted that there is substantial overlap in the substrate specificities of PXR and its critical CYP3A target gene. This prompted us to ask whether endogenous CYP3A substrates also serve as PXR ligands. We demonstrate that 5beta-cholestane-3alpha,7alpha,12alpha-triol (triol), a cholesterol-derived CYP3A substrate, is a potent PXR agonist that effectively induces cyp3a expression in mice. This defines a critical salvage pathway that can be autoinduced to minimize triol accumulation. In contrast, triol can accumulate to very high levels in humans, and unlike mice, these people develop the severe clinical manifestations of cerebrotendinous xanthomatosis. The reason for these dramatic species differences has remained unclear. We now demonstrate that triol fails to activate human PXR or induce the CYP3A-salvage pathway. This explains why humans are more susceptible to sterol accumulation and suggests that synthetic ligands for human PXR could be used to treat cerebrotendinous xanthomatosis and other disorders of cholesterol excess.

Anesthetics↗

PubMed2003

Characterization of adenosine receptor(s) involved in adenosine-induced bronchoconstriction in an allergic mouse model.

We recently reported that adenosine caused bronchoconstriction and enhanced airway inflammation in an allergic mouse model. In this study, we further report the characterization of the subtype of adenosine receptor(s) involved in bronchoconstriction. 5'-(N-ethylcarboxamido)adenosine (NECA), a nonselective adenosine agonist, elicited bronchoconstriction in a dose-dependent manner. Little effects of N(6)-cyclopentyladenosine (A(1)-selective agonist) and 2-p-(2-carboxyethyl)phenethylamino-5'-N-ethylcarboxamidoadenosine (A(2A)-selective agonist) compared with NECA were observed in this model. 2-Chloro-N(6)-(3-iodobenzyl)-9-[5-(methylcarbamoyl)-beta-d-ribofuranosyl]adenosine, an A(3)-selective receptor agonist, produced a dose-dependent bronchoconstrictor response, which was blocked by selective A(3) antagonist 2,3-diethyl-4,5-dipropyl-6-phenylpyridine-3-thiocarboxylate-5-carboxylate (MRS1523). However, MRS1523 only partially inhibited NECA-induced bronchoconstriction. Neither selective A(1) nor A(2A) antagonists affected NECA-induced bronchoconstriction. Enprofylline, a relatively selective A(2B) receptor antagonist, blocked partly NECA-induced bronchoconstriction. Furthermore, a combination of enprofylline and MRS1523 completely abolished NECA-induced bronchoconstrictor response. Using RT-PCR, we found that all four adenosine receptor subtypes are expressed in control lungs. Allergen sensitization and challenge significantly increased transcript levels of the A(2B) and A(3) receptors, whereas the A(1) receptor message decreased. No change in transcript levels of A(2A) receptors was observed after allergen sensitization and challenge. These findings suggest that A(2B) and A(3) adenosine receptors play an important role in adenosine-induced bronchoconstriction in our allergic mouse model. Finally, whether the airway effects of the receptor agonists/antagonists are direct or indirect needs further investigations.

Adenosine↗