Biomedical subjects

Lei Zhang

Publications and source records attributed to Lei Zhang.

At least 181 records · Page 10Linked to original sources

PubMed2005

BRAF mediates RET/PTC-induced mitogen-activated protein kinase activation in thyroid cells: functional support for requirement of the RET/PTC-RAS-BRAF pathway in papillary thyroid carcinogenesis.

In human papillary thyroid cancers (PTCs), mutations of RET/PTC, NTRK, RAS, or BRAF are found in about two thirds of cases with practically no overlap, providing genetic evidence that constitutive signaling along RET-RAS-BRAF-MAPK is key to their development. The requirement for BRAF in RET/PTC-mediated MAPK activation and gene expression has not been tested functionally. There are three RAF isoforms: ARAF, BRAF, and CRAF. Compared with the others, ARAF is a much weaker stimulator of MAPK. To determine the key RAF isoform mediating RET/PTC-induced ERK phosphorylation, we stably transfected doxycycline-inducible RET/PTC3-expressing thyroid PCCL3 cells with small interfering RNA vectors to induce selective knockdown of BRAF or CRAF. Conditional RET/PTC3 expression induced comparable ERK phosphorylation in CRAF knockdown and control cells but negligible ERK phosphorylation in BRAF knockdown cells. Selective knockdown of BRAF prevented RET/PTC-dependent down-regulation of the sodium iodide symporter, a gene that confers key biological effects of RET/PTC in PTCs. Moreover, microarray analysis revealed numerous RET/PTC-regulated genes showing requirement of BRAF for appropriate expression. These data indicate that BRAF is required for RET/PTC-induced MAPK activation in thyroid cells and support the notion that BRAF inactivation may be an attractive target for PTCs.

Animals↗

PubMed2005

Kinase inhibitor recognition by use of a multivariable QSAR model.

We have applied a retrosynthetic program to determine the scaffold and R-group chemical space seen within a library of known kinase inhibitors and non-kinase drug-like molecules. Comparison of the differences quickly revealed that kinase inhibitors are distinct in several chemical fragment and physical properties. We then applied these descriptors in a multivariable quantitative structure-activity relationship (QSAR) model with the goal to distinguish kinase inhibitors from non-kinase drug-like molecules. This model is heuristic in that it was trained over a dataset of 258 known kinase inhibitors and 230 non-kinase drug molecules. The final model recognized 98% of the training set as being kinase inhibitors and had a false positive rate of 15%. This trait for false positives was accepted out of a desire to maintain diversity and not miss possible good kinase inhibitors for screening. The model was validated by reserving a portion of the datasets as test sets, which were not included in the QSAR model building stage. This was done repetitively for different percentiles of the total dataset population. It was seen that model recognition and false positive were only slightly damaged well down to a 70% reserve (30% dataset used for QSAR model training while 70% used for reserve test set). Beyond 70%, the QSAR models were inconsistent, signifying that the training sets were inadequately diverse to represent the greater reserve test sets. We applied this model to evaluate the commercial kinase libraries available from Asinex, BioFocus, ChemDiv and LifeChemicals to facilitate purchase decisions for compounds for HTS for lead compounds. We observed that there are significant differences in populations of recognizable kinase inhibitors across the vendors analyzed, with BioFocus showing the greatest population of kinase like molecules.

Computational Biology↗

PubMed2005

Laser-induced fluorescence spectra of 4-methylcyclohexoxy radical and perdeuterated cyclohexoxy radical and direct kinetic studies of their reactions with O2.

The laser-induced fluorescence (LIF) excitation spectra of the 4-methylcyclohexoxy and d11-cyclohexoxy radicals have been measured for the first time. LIF intensity was used as a probe in direct kinetic studies of the reaction of O(2) with trans-4-methylcyclohexoxy and d11-cyclohexoxy radicals from 228 to 301 K. Measured rate constants near room temperature are uniformly higher than the Arrhenius fit to the lower-temperature data, which can be explained by the regeneration of cyclic alkoxy radicals from the product of their beta-scission and the effect of O(2) concentration on the extent of regeneration. The Arrhenius expressions obtained over more limited ranges were k(O2) = (1.4(+8)(-1)) x 10(-13) exp[(-810 +/- 400)/T] cm(3) molecule(-1) s(-1) for trans-4-methylcyclohexoxy (228-292 K) and k(O2) = (3.7(+4)(-1)) x 10(-14) exp )[(-760 +/- 400) /T] cm(3) molecule(-1) s(-1) for d11-cyclohexoxy (228-267 K) independent of pressure in the range 50-90 Torr. The room-temperature rate constant for the reaction of trans-4-methylcyclohexoxy radical with O2 (obtained from the Arrhenius fit) is consistent with the commonly recommended value, but the observed activation energy is approximately 3 times larger than the recommended value of 0.4 kcal/mol and half the value previously found for the reaction of normal cyclohexoxy radical with O2.

Cyclohexanes↗

PubMed2005

Restoration of immunity in lymphopenic individuals with cancer by vaccination and adoptive T-cell transfer.

Immunodeficiency is a barrier to successful vaccination in individuals with cancer and chronic infection. We performed a randomized phase 1/2 study in lymphopenic individuals after high-dose chemotherapy and autologous hematopoietic stem cell transplantation for myeloma. Combination immunotherapy consisting of a single early post-transplant infusion of in vivo vaccine-primed and ex vivo costimulated autologous T cells followed by post-transplant booster immunizations improved the severe immunodeficiency associated with high-dose chemotherapy and led to the induction of clinically relevant immunity in adults within a month after transplantation. Immune assays showed accelerated restoration of CD4 T-cell numbers and function. Early T-cell infusions also resulted in significantly improved T-cell proliferation in response to antigens that were not contained in the vaccine, as assessed by responses to staphylococcal enterotoxin B and cytomegalovirus antigens (P < 0.05). In the setting of lymphopenia, combined vaccine therapy and adoptive T-cell transfer fosters the development of enhanced memory T-cell responses.

Adoptive Transfer↗

PubMed2005

Identification of gene markers based on well validated and subcategorized stressed animals for potential clinical applications in PTSD.

Post-traumatic stress disorder (PTSD) is a complex mental disorder that can develop in response to traumatic experiences. The molecular mechanisms underlying the pathology of PTSD are poorly understood, and this lack of knowledge hampers our ability to find superior therapeutic approaches to the treatment of this disorder. There are two main reasons for our lack of study in this area: here is no sufficiently validated animal model and lack of large-scale studies for the search of underlying molecular mechanisms. Thus, to promote research on PTSD (especially its molecular mechanisms) and to set molecular basis for searching novel medications of this disorder, large-scale, genome-wide interrogation of a significant amount of genes based upon a well validated animal model is demanded. We hypothesize that a significant number of genes are involved in PTSD. It is only with a large number of these genes identified in specific samples of PTSD-related population, and then it is possible for a sufficient understanding of the pathology at the molecular level of a PTSD, as well as for enhancing the PTSD's therapeutic and preventative strategies. Two prerequisites are needed for testing this hypothesis: (1) relative pure samples from a well validated animal model; and (2) genome-wide screening of PTSD molecular targets. For the animal model, we suggest to use the predator-exposure paradigm, in which rats are exposed to a predator, this model has previously been evaluated behaviorally well emulated the clinical symptoms of PTSD. For a better stringency, three criteria can be used to further validate this animal model: analogous (similarity of behavior), predictive (predictability of drug response) and biological mechanism (e.g., electrophysiological and pathological change in amygdala). For large-scale molecular target screening, the new microarray technology, which can profile expression of tens of thousands genes simultaneously, is the method of choice. The validity and practicability of this hypothesis and the strategy for its testing have been supported by our preliminary laboratory data.

Animals↗

PubMed2005

Activation of bone morphogenetic protein-6 gene transcription in MCF-7 cells by estrogen.

BACKGROUND: Bone morphogenetic protein-6 (BMP-6) is closely correlated with tumor differentiation and skeletal metastasis. Estrogen is considered as a stimulant for the initiation and promotion of breast cancer. Previous studies demonstrated that 17beta-estadiol (E2) can selectively increase the expression of BMP-6. This experiment is designed to detect the molecular mechanism of estrogen activating BMP-6 gene transcription in human estrogen receptor positive (ER+) breast cancer cell line MCF-7. METHODS: After the treatment of MCF-7 cells with E2 at different concentrations (10(-11) mol/L, 10(-9) mol/L, 10(-7) mol/L), the BMP-6 expression level was examined through real-time polymerase chain reaction. Through restriction enzyme digestion, human BMP-6 1.2 kb long promoter, BMP-6 0.7 kb long promoter was cloned into pGL-3 basic vector; after the treatment with 10(-7) mol/L E2, luciferase activities of the two promoters were detected. Site-directed mutagenesis was performed to obtain the mutant forms of estrogen response element half-site (1/2 ERE) element and Sp1 sites in the BMP-6 promoter, the activities of these mutant form promoters were detected following the methods mentioned above. Chromatin immunoprecipitation (ChIP) assay was also used to confirm the binding of estrogen receptor alpha (ERalpha) on BMP-6 promoter in the presence of E2. RESULTS: E2 dose dependently increased BMP-6 mRNA expression in human ER+ breast cancer cell line MCF-7. At a dose of 10(-7) mol/L E2, human BMP-6 1.2 kb promoter activity was increased by 90% compared with the control group treated with ethanol (P < 0.05). Both the 1/2 ERE response element mutant form and the Sp1 site mutant form of the BMP-6 promoter abolished the activation of the BMP-6 promoter's response to E2. Through ChIP assay, the binding of ERalpha on 1/2 ERE response element in BMP-6 promoter was further validated. CONCLUSION: Estrogen induces BMP-6 expression in human ER+ breast cancer cell line MCF-7 through its receptor ERalpha binding on 1/2 ERE element in the BMP-6 promoter.

Bone Morphogenetic Protein 6↗

PubMed2005

AFM and STM characterization of thiol and thiophene functionalized SWNTs: pitfalls in the use of chemical markers to determine the extent of sidewall functionalization in SWNTs.

Thiol- and thiophene-functionalized SWNTs prepared via the reaction of a substituted amine with fluoronanotubes show similar levels of sidewall functionalization, however, the use of Au nanoparticles as chemical markers for AFM gives misleading results for substituent distribution since STM shows the thiol substituents grouped in bands while the thiophene substituents uniformly distributed along the SWNTs.

Journal Article↗

PubMed2005

Photosynthetic responses of C3 and C4 species to seasonal water variability and competition.

This study examined the impacts of seasonal water variability and interspecific competition on the photosynthetic characteristics of a C3 (Leymus chinensis) and a C4 (Chloris virgata) grass species. Plants received the same amount of water but in three seasonal patterns, i.e. the one-peak model (more water in the summer than in the spring and autumn), the two-peak model (more water in the spring and autumn than in the summer), and the average model (water evenly distributed over the growing season). The effects of water variability on the photosynthetic characteristics of the C3 and C4 species were dependent on season. There were significant differences in the photosynthetic characteristics of the C4 species in the summer and the C3 species in the autumn among the three water treatments. Interspecific competition exerted negative impacts on the C3 species in August and September but had no effects on the C4 species in any of the four measuring dates. The relative competitive capability of the two species was not altered by water availability. The assimilation rate, the maximum quantum yield of net CO2 assimilation, and the maximum rate of carboxylation of the C3 species were 13-56%, 5-11%, and 11-48% greater, respectively, in a monoculture than in a mixture in August and September. The results demonstrated that the photosynthetic characteristics of the C3 and C4 species were affected by water availability, but the effects varied considerably with season.

Carbon↗

PubMed2005

Dissociation in the neural basis underlying Chinese tone and vowel production.

Neuropsychologists have debated over whether the processing of segmental and suprasegmental units involves different neural mechanisms. Focusing on the production of Chinese lexical tones (suprasegmental units) and vowels (segmental units), this study used the adaptation paradigm to investigate a possible neural dissociation for tone and vowel production. Ten native Chinese speakers were asked to name Chinese characters and pinyin (Romanized phonetic system for Chinese language) that varied in terms of tones and vowels. fMRI results showed significant differences in the right inferior frontal gyrus between tone and vowel production (more activation for tones than for vowels). Brain asymmetry analysis further showed that tone production was less left-lateralized than vowel production, although both showed left-hemisphere dominance.

Adult↗

PubMed2005

Functions of the Per/ARNT/Sim domains of the hypoxia-inducible factor.

The heterodimeric transcription factor hypoxia-inducible factor (HIF) plays an important role in the progression of a number of processes in which O2 availability is compromised and, as such, has become an increasingly attractive therapeutic target. Although tremendous progress has been made in recent years in unraveling the mechanisms underlying O2-dependent regulation of HIF through its O2-dependent degradation domain and C-terminal transactivation domain, our understanding of the contributions of other structural elements, particularly the Per/ARNT/Sim (PAS)-A and PAS-B domains, to the activity of HIF is incomplete. Using insights derived from the recently determined solution structures of the HIF PAS-B domains as a starting point, we have explored the function(s) of the HIF-2alpha PAS domains via mutational analysis. In contrast to recent models, our data reveal that both PAS domains of the HIF-alpha subunit are necessary for heterodimer formation but are not required to mediate other HIF functions in which PAS domains have been implicated. Because disruption of individual PAS domains compromise HIF function independent of the mechanism of HIF induction, these data demonstrate the potential utility of targeting these domains for therapeutic applications.

Animals↗

PubMed2005

Expression of heparanase mRNA in anti-sense oligonucleotide-transfected human esophageal cancer EC9706 cells.

AIM: To investigate the effects of anti-sense oligonucleotides (ASODNs) on mRNA expression of heparanase in human esophageal cancer EC9706 cells. METHODS: One non-sense oligonucleotide (N-ODN) and five ASODNs against different heparanase mRNA sites were transfected into EC9706 cells, then the expression of heparanase mRNA in EC9706 cells was studied by in situ hybridization. RESULTS: The expression of heparanase mRNA could be inhibited by ASODNs. There was no significant difference among five ASODNs (P>0.05), but there was a significant difference between ASODNs and N-ODN or non-transfected group (ASODN1: 2.25+/-0.25, ASODN2: 2.21+/-0.23, ASODN3: 2.23+/-0.23, ASODN4: 2.25+/-0.24 vs N-ODN: 3.47+/-2.80 or non- transfected group: 3.51+/-2.93 respectively, P<0.05). CONCLUSION: The expression of heparanase mRNA in EC9706 cells can be inhibited by ASODNs in vivo, and heparanase ASODNs can inhibit metastasis of esophageal squamous cell carcinoma or other tumors by inhibiting the expression of heparanase.

Cell Line, Tumor↗

PubMed2005

A colloidal Au monolayer modulates the conformation and orientation of a protein at the electrode/solution interface.

The orientation and conformation of adsorbed cytochrome c (cyt c) at the interface between an electrode modified with colloidal Au and a solution were studied by electrochemical, spectroscopic, and spectroelectrochemical techniques. The results indicate that the colloidal Au monolayer formed via preformation of an organic self-assembled monolayer (SAM) can increase the electronic coupling between the SAM and cyt c in the same manner as bifunctional molecular bridges, one functional group of which is bound to the electrode surface while the other interacts with the protein surface. The approach of cyt c to the modified electrode/solution interface can be assisted by strong interactions of the intrinsic charge of colloidal particles with cyt c, while the heme pocket remains almost unchanged due to the screening effect of the negatively charged field created by the intrinsic charge. The conformational changes of cyt c induced by its adsorption at a bare glassy carbon electrode/solution interface and the effect of the electric field on the ligation state of the heme can be avoided at the colloidal-Au-modified electrode/solution interface. Finally, a possible model for the adsorption orientation of cyt c at the colloidal-Au-modified electrode/solution interface is proposed.

Electrochemistry↗

PubMed2005

Saururus cernuus lignans--potent small molecule inhibitors of hypoxia-inducible factor-1.

Hypoxia-inducible factor-1 (HIF-1) represents an important tumor-selective therapeutic target for solid tumors. In search of novel small molecule HIF-1 inhibitors, 5400 natural product-rich extracts from plants, marine organisms, and microbes were examined for HIF-1 inhibitory activities using a cell-based reporter assay. Bioassay-guided fractionation and isolation, followed by structure elucidation, yielded three potent natural product-derived HIF-1 inhibitors and two structurally related inactive compounds. In a T47D cell-based reporter assay, manassantin B1, manassantin A, and 4-O-methylsaucerneol inhibited hypoxia-induced HIF-1 activation with IC50 values of 3, 3, and 20 nM, respectively. All three compounds are relatively hypoxia-specific inhibitors of HIF-1 activation, in comparison to other stimuli. The hypoxic induction of HIF-1 target genes CDKN1A, VEGF, and GLUT-1 were also inhibited. These compounds inhibit HIF-1 by blocking hypoxia-induced nuclear HIF-1alpha protein accumulation without affecting HIF-1alpha mRNA levels. In addition, preliminary structure-activity studies suggest specific structural requirements for this class of HIF-1 inhibitors.

Cell Line, Tumor↗

PubMed2005

Stepholidine protects against H2O2 neurotoxicity in rat cortical neurons by activation of Akt.

The fundamental pathological process(es) associated with schizophrenia (SZ) remain(s) uncertain, but multiple lines of evidence suggest that this condition is associated with excessive stimulation of striatal dopamine (DA) D2 receptors, deficient stimulation of medial prefrontal cortex (mPFC) D1 receptors as well as neuronal apoptosis. Unlike typical antipsychotics, stepholidine (SPD), which is isolated from the Chinese herb stephania, has D1 and D2 dual properties and regulates neuronal cell differentiation and proliferation. It is unknown, however, whether it possesses a neuroprotective property. Here, we report that SPD prevented neuronal cell death from H2O2 exposure and increased the levels of phosphorylated Akt (pAkt), a serine/threonine protein kinase. The SPD-induced neuroprotection and activation of Akt were blocked by LY294002, a PI3-K inhibitor, suggesting that the anti-apoptotic action of SPD is mediated via the PI3-K/Akt signaling pathway. Thus, as a survival or anti-apoptotic factor for neuronal cells, SPD may contribute to the therapeutic action of SPD in SZ treatment.

2,3,4,5-Tetrahydro-7,8-dihydroxy-1-phenyl-1H-3-ben↗

PubMed2005

Carbachol induces burst firing of dopamine cells in the ventral tegmental area by promoting calcium entry through L-type channels in the rat.

Enhanced activity of the central dopamine system has been implicated in many psychiatric disorders including schizophrenia and addiction. Besides terminal mechanisms that boost dopamine levels at the synapse, the cell body of dopamine cells enhances terminal dopamine concentration through encoding action potentials in bursts. This paper presents evidence that burst firing of dopamine cells in the ventral tegmental area was under cholinergic control using nystatin-perforated patch clamp recording from slice preparations. The non-selective cholinergic agonist carbachol excited the majority of recorded neurones, an action that was not affected by blocking glutamate and GABA ionotropic receptors. Twenty per cent of dopamine cells responded to carbachol with robust bursting, an effect mediated by both muscarinic and nicotinic cholinoceptors postsynaptically. Burst firing induced as such was completely dependent on calcium entry as it could be blocked by cadmium and more specifically the L-type blocker nifedipine. In the presence of the sodium channel blocker tetrodotoxin, carbachol induced membrane potential oscillation that had similar kinetics and frequency as burst firing cycles and could also be blocked by cadmium and nifedipine. Direct activation of the L-type channel with Bay K8644 induced strong bursting which could be blocked by nifedipine but not by depleting internal calcium stores. These results indicate that carbachol increases calcium entry into the postsynaptic cell through L-type channels to generate calcium-dependent membrane potential oscillation and burst firing. This could establish the L-type channel as a target for modulating the function of the central dopamine system in disease conditions.

3-Pyridinecarboxylic acid, 1,4-dihydro-2,6-dimethy↗

PubMed2005

Electrochemical behavior of a covalently modified glassy carbon electrode with aspartic acid and its use for voltammetric differentiation of dopamine and ascorbic acid.

Aspartic acid was covalently grafted on to a glassy carbon electrode (GCE) by amine cation radical formation in the electrooxidation of the amino-containing compound. X-ray photoelectron spectroscopic (XPS) measurement and cyclic voltammetric experiments proved the aspartic acid was immobilized as a monolayer on the GCE. Electron transfer to Fe(CN)6(4-) in solution of different pH was studied by cyclic voltammetry. Changes in solution pH resulted in the variation of the charge state of the terminal group; surface pK(a) values were estimated on the basis of these results. Because of electrostatic interactions between the negatively charged groups on the electrode surface and dopamine (DA) and ascorbic acid (AA), the modified electrode was used for electrochemical differentiation between DA and AA. The peak current for DA at the modified electrode was greatly enhanced and that for AA was significantly reduced, which enabled determination of DA in the presence of AA. The differential pulse voltammetric (DPV) peak current was linearly dependent on DA concentration over the range 1.8 x 10(-6)-4.6 x 10(-4) mol L(-1) with slope (nA micromol(-1) L) and intercept (nA) of 47.6 and 49.2, respectively. The detection limit (3delta) was 1.2 x 10(-6) mol L(-1). The high selectivity and sensitivity for dopamine was attributed to charge discrimination and analyte accumulation. The modified electrode has been used for determination of DA in samples, in the presence of AA, with satisfactory results.

Ascorbic Acid↗

PubMed2005

Bilirubin removal from human plasma by Cibacron Blue F3GA using immobilized microporous affinity membranous capillary method.

A novel affinity sorbent system for direct bilirubin removal from human plasma was developed. These new adsorbents comprise Cibacron Blue F3GA as the specific ligand, and microporous membranous poly(tetrafluoroethylene) capillary (modified by coating with a hydrophilic layer of poly(vinyl alcohol) after activation) as the carrier matrix. The affinity adsorbents carrying 126.5 micromol Cibacron Blue F3GA/g polymer was then used to remove bilirubin in a flow-injection system. Non-specific adsorption on the poly(vinyl alcohol) coated capillary remains low, and higher affinity adsorption capacity, of up to 76.2 mg/g polymer was obtained after dye immobilization. The bilirubin adsorption capacity of the affinity capillary decreased with increase in the recirculation rate of plasma. The adsorption capacity increased with increase the temperature while decreased with increase the ionic strength. The maximum adsorption was only observed in neutral solution (pH 6-7). The adsorption isotherm fitted the Langmuir model well. These new adsorbents have higher velocity of mass transfer, better adsorption capacity, less fouling, longer service life and good reusability. The results of blood tests suggested the dye affinity capillary has good blood compatibility.

Adsorption↗

PubMed2005

Microvascular flow routes in muscle controlled by vasoconstrictors.

Vasoconstrictors can either increase or decrease metabolism of the constant flow pump-perfused rat hindlimb. In addition, there is indirect evidence from vascular casts, surface fluorometry, dye entrapment studies, and fluorescent microsphere mapping of flow that this may be due to redistribution of flow between putatively nutritive and non-nutritive routes within muscle. In the present study, we used two methods in an attempt to identify perfused nutritive and non-nutritive vessels in muscle sections: (i) a combination of perfusion fixation with glutaraldehyde and post-perfusion Griffonia simplicifolia lectin and (ii) perfusion with rhodamine-dextran70 (lysine fixable) and post-fixation with formaldehyde. Perfusions involved vehicle only (control, a mix of nutritive and non-nutritive flow), 15 nM angiotensin II (AII) to increase, or 1 microM serotonin (5-HT) to decrease nutritive flow. Microscopic examination of muscle sections following AII showed an increase in perfused capillaries with fewer areas of under-perfusion, relative to control. In contrast, 5-HT caused a marked decrease in perfused capillaries relative to control and evidence that flow was carried by connective tissue vessels that on average were of greater diameter and were more sparsely distributed than capillaries. It is concluded that vasoconstrictors that alter hindlimb metabolism do so by intra-muscle redistribution between capillaries (nutritive) and non-nutritive (connective tissue) vessels within each muscle.

Angiotensin II↗