Biomedical subjects

Li Li

Publications and source records attributed to Li Li.

At least 469 records · Page 26Linked to original sources

PubMed2003

Internal tandem duplication mutation of FLT3 blocks myeloid differentiation through suppression of C/EBPalpha expression.

Constitutively activating mutations of FMS-like tyrosine kinase 3 (FLT3) occur in approximately one third of patients with acute myeloid leukemia (AML) and are associated with poor prognosis. Altered FLT3 signaling leads to antiapoptotic and proliferative signaling pathways. We recently showed that these mutations can also contribute to the differentiation arrest that characterizes leukemia. In this report we investigated the mechanism by which internal tandem duplication (ITD) mutation of FLT3 signaling blocks differentiation. Normally, myeloid differentiation requires the induction of CCAAT/enhancer-binding protein alpha (C/EBPalpha) and PU.1 expression. Expression of both genes was repressed by FLT3/ITD signaling in 32Dcl3 (32D) cells and this repression was overcome by treatment with a FLT3 inhibitor, allowing differentiation to proceed. We also observed increased expression of C/EBPalpha and PU.1 accompanied by signs of differentiation in 2 of 3 primary AML samples from patients with FLT3/ITD mutations receiving a FLT3 inhibitor, CEP-701, as part of a clinical trial. Forced expression of C/EBPalpha was also able to overcome FLT3/ITD-mediated differentiation block, further proving the importance of C/EBPalpha in this process.

Blotting, Northern↗

PubMed2003

Inclusion complexation and solubilization of paclitaxel by bridged bis(beta-cyclodextrin)s containing a tetraethylenepentaamino spacer.

A novel water-soluble paclitaxel complex has been prepared by inclusion complexation with bridged bis(beta-cyclodextrin)s and characterized by means of (1)H NMR, SEM, powder X-ray diffraction patterns, TG-DTA, DSC, FT-IR, and 2D NOESY. The cyclodextrins were able to solubilize paclitaxel to levels as high as 2 mg/mL. Furthermore, the cytotoxicity of the novel complexes was assessed using a K562 leukemia cell line which indicated that drug concentrations of 10 pg/mL elicited an inhibitory effect.

Antineoplastic Agents↗

PubMed2003

AFLP-based genetic linkage maps of the pacific oyster Crassostrea gigas Thunberg.

Amplified fragment length polymorphisms (AFLPs) were used for genome mapping in the Pacific oyster Crassostrea gigas Thunberg. Seventeen selected primer combinations produced 1106 peaks, of which 384 (34.7%) were polymorphic in a backcross family. Among the polymorphic markers, 349 were segregating through either the female or the male parent. Chi -square analysis indicated that 255 (73.1%) of the markers segregated in a Mendelian ratio, and 94 (26.9%) showed significant ( P < 0.05) segregation distortion. Separate genetic linkage maps were constructed for the female and male parents. The female framework map consisted of 119 markers in 11 linkage groups, spanning 1030.7 cM, with an average interval of 9.5 cM per marker. The male map contained 96 markers in 10 linkage groups, covering 758.4 cM, with 8.8 cM per marker. The estimated genome length of the Pacific oyster was 1258 cM for the female and 933 cM for the male, and the observed coverage was 82.0% for the female map and 81.3% for the male map. Most distorted markers were deficient for homozygotes and closely linked to each other on the genetic map, suggesting the presence of major recessive deleterious genes in the Pacific oyster.

Animals↗

PubMed2003

The potentiation role of hepatopoietin on activator protein-1 is dependent on its sulfhydryl oxidase activity.

Hepatopoietin (HPO) is a novel hepatotrophic growth factor that stimulates hepatocyte proliferation by two pathways. In the first, intracellular HPO specifically modulates the activator protein-1 (AP-1) pathway through JAB1 (Jun activation domain-binding protein 1), whereas in the second, extracellular HPO triggers the mitogen-activated protein kinase pathway by binding its specific receptor on the cell surface. In this report we demonstrate that HPO is a flavin-linked sulfhydryl oxidase, and the invariant CXXC (Cys-Xaa-Xaa-Cys) motif in HPO is essential for the enzyme activity of HPO but not for its dimerization nor for its binding ability with JAB1. Two intramolecular disulfides were identified in HPO by mass spectrometry, one of which is formed by the redox CXXC cysteine residues. HPO site-directed mutants (Cys/Ser) at active sites, which lost sulfhydryl oxidase activity, could not increase c-Jun phosphorylation and failed to potentiate JAB1-mediated AP-1 activation. However, the mutants still have mitogenic stimulation and mitogen-activated protein kinase activation effects on HepG2 cells. Thus, it can be concluded that the potentiation role of HPO on AP-1 is dependent on its sulfhydryl oxidase activity.

Amino Acid Sequence↗

PubMed2003

Identification of an alternatively processed nicotinic receptor alpha7 subunit RNA in mouse brain.

The data in this report describe the discovery and characterization of a previously unidentified alternatively processed RNA for the neuronal nicotinic receptor alpha7 subunit. The unique transcript contains an extra exon that arises from alternative splicing of intron nine of the alpha7 subunit RNA. The alpha7 subunit protein resulting from this alternatively processed RNA is truncated shortly after transmembrane domain three. The variant protein also has a predicted amino acid substitution in the large N-terminal domain as a consequence of a non-templated nucleotide substitution present in the variant alpha7 subunit RNA. The mechanism responsible for the nucleotide substitution is not known. Initial characterization of the variant alpha7 subunit suggests that it is expressed in mouse brain in a pattern similar to the standard alpha7 subunit although at reduced levels. The variant alpha7 subunit was also found to act as a dominant-negative effecter of normal alpha7 subunit function.

Alternative Splicing↗

PubMed2003

Shock-induced arrhythmogenesis is enhanced by 2,3-butanedione monoxime compared with cytochalasin D.

Investigation of the mechanisms of arrhythmia genesis and maintenance has benefited from the use of optical mapping techniques that employ excitation-contraction uncouplers. We investigated the effects of the excitation-contraction uncouplers 2,3-butanedione monoxime (BDM) and cytochalasin D (Cyto D) on the induction and maintenance of arrhythmia by electric shocks. Electrical activity was optically mapped from anterior epicardium of rabbit hearts (n = 9) during shocks (-100 V, 8 ms) applied from a ventricular lead at various phases of action potential duration (APD). Restitution curves were obtained using S1-S2 protocol and measurement of APD values at 70% of repolarization. Compared with Cyto D, BDM significantly shortened APD at 90% of repolarization, although no significant difference in dispersion of repolarization was observed. Wavelength was also shortened with BDM. In general, shock-induced arrhythmias with BDM and Cyto D were ventricular tachycardic in nature. With respect to shock-induced sustained arrhythmias, the vulnerable window was wider and the incidence was higher with BDM than with Cyto D. There was also a difference in the morphology of ventricular tachycardia (VT) between the two agents. The arrhythmias with BDM usually resembled monomorphic VT, especially those that lasted >30 s. In contrast, arrhythmias with Cyto D more resembled polymorphic VT. However, the average number of phase singularities increased under Cyto D vs. BDM, whereas no significant difference in the dominant frequency of shock-induced sustained arrhythmia was observed. BDM reduced the slope of the restitution curve compared with Cyto D, but duration of arrhythmia under BDM was significantly increased compared with Cyto D. In conclusion, BDM increased arrhythmia genesis and maintenance relative to Cyto D.

Action Potentials↗

PubMed2003

Analysis of the complete genome of the tick-borne flavivirus Omsk hemorrhagic fever virus.

Omsk hemorrhagic fever virus (OHF) is a tick-borne flavivirus endemic to Western Siberia. This virus is the only known tick-borne flavivirus to cause hemorrhagic disease in humans in the absence of encephalitis. OHF virus circulates within a small, defined niche in which other tick-borne complex flaviviruses are also present. The objectives of this study were to genetically classify OHF virus based on its complete genome and to identify genetic determinants that might be involved in tissue tropism and viral replication leading to the disease state caused by this virus. The OHF virus genome was sequenced and phylogenetic analysis demonstrated that OHF virus falls within the tick-borne encephalitis serocomplex of flaviviruses, yet is distinct from other members of the complex, including those closely associated geographically. OHF is also distinct from Alkhurma (ALK) and Kyasanur forest disease (KFD) viruses, both of which cause disease that includes hemorrhagic and encephalitic manifestations. Several amino acid residues were found to be distinct among OHF, KFD, and ALK viruses; these residues include E-76, which is closely associated with the viral envelope protein fusion peptide. In addition, variation between the viral 5'-untranslated region of OHF and other tick-borne flaviviruses suggests potential variability in viral replication. These data demonstrate that OHF is a unique virus among the tick-borne flaviviruses and also provide insight to viral biodiversity and tropism.

5' Untranslated Regions↗

PubMed2003

Inclusion complexation behavior of dyestuff guest molecules by a bridged bis(cyclomaltoheptaose)[bis(beta-cyclodextrin)] with a pyromellitic acid diamide tether.

A novel bridged bis(beta-cyclodextrin) with a pyromellitic acid 2,5-diamide tether (2) has been synthesized by reaction of 6(I)-(2-aminoethyleneamino)-6-deoxycyclomaltoheptaose [mono 6-(2-aminoethyleneamino)-6-deoxy-beta-cyclodextrin] with 1,2,4,5-benzenetetracarboxylic dianhydride. Its inclusion complexation behavior with some representative dyestuffs, i.e., Acridine Red (AR), Rhodamine B (RhB), Neutral Red (NR), Brilliant Green (BG), was studied by using UV-absorption, fluorescence, and 2D NMR spectroscopy. Fluorescence titrations have been performed at 25 degrees C in pH 7.2 buffer solution to calculate the binding constants of resulting complexes. These results obtained indicated that bis(beta-cyclodextrin) 2 exhibits the strongly enhanced binding ability with all dye molecules examined compared with natural cyclodextrins. The binding modes of 2 with dye molecules have been deduced by 2D NMR experiments to establish the correlations between molecular conformations and binding constants of inclusion complexation. It is found that the improved binding ability and molecular selectivity of 2 could be attributed to double-cavity cooperative inclusion interaction and the size/shape matching between the host and guest.

Amides↗

PubMed2003

Reconstitution of CD40 and CD80 in dendritic cells generated from blasts of patients with acute myeloid leukemia.

Acute myeloid leukemia (AML) is a clonal disease of hematopoiesis with poor clinical outcome despite recent improvements in chemotherapy and stem cell transplantation regimens. Immunotherapy with dendritic cells (DCs) eliciting specific T cell responses to leukemia-associated antigens (LAAs) might be a therapeutic option. DCs must express HLA class I/II molecules and the costimulatory molecules CD40, CD80 and CD86 to effectively activate T cells for the subsequent lysis of leukemic blasts. The expression of these antigens on DCs generated from 15 AML patients (AML-DCs) and on DCs generated from 15 healthy volunteers (HV-DCs) was analyzed by FACS. All DCs displayed the typical morphology and tested negative for B, T and NK cell markers. The sustained mRNA expression of LAAs such as PRAME, RHAMM or WT-1 proved that the AML-DCs originated from AML blasts. Compared with AML blasts, the expression of CD40, CD80, CD86 and HLA-DR was upregulated during DC culture to a median of 80-98% on AML-DCs. HLA-ABC was preserved on AML-DCs (median 95%). Expression of CD40, CD80 and CD83 remained lower on AML-DCs than on HV-DCs. AML-DCs express at least one LAA and strongly express HLA and costimulatory molecules, the prerequisites for eliciting T cell responses. AML-DCs may play a role in vaccine-based immunotherapies for AML patients.

Acute Disease↗

PubMed2003

A long-range Shh enhancer regulates expression in the developing limb and fin and is associated with preaxial polydactyly.

Unequivocal identification of the full composition of a gene is made difficult by the cryptic nature of regulatory elements. Regulatory elements are notoriously difficult to locate and may reside at considerable distances from the transcription units on which they operate and, moreover, may be incorporated into the structure of neighbouring genes. The importance of regulatory mutations as the basis of human abnormalities remains obscure. Here, we show that the chromosome 7q36 associated preaxial polydactyly, a frequently observed congenital limb malformation, results from point mutations in a Shh regulatory element. Shh, normally expressed in the ZPA posteriorly in the limb bud, is expressed in an additional ectopic site at the anterior margin in mouse models of PPD. Our investigations into the basis of the ectopic Shh expression identified the enhancer element that drives normal Shh expression in the ZPA. The regulator, designated ZRS, lies within intron 5 of the Lmbr1 gene 1 Mb from the target gene Shh. The ZRS drives the early spatio-temporal expression pattern in the limb of tetrapods. Despite the morphological differences between limbs and fins, an equivalent regulatory element is found in fish. The ZRS contains point mutations that segregate with polydactyly in four unrelated families with PPD and in the Hx mouse mutant. Thus point mutations residing in long-range regulatory elements are capable of causing congenital abnormalities, and possess the capacity to modify gene activity such that a novel gamut of abnormalities is detected.

Animals↗

PubMed2003

Overexpression of leukocyte-type 12-lipoxygenase promotes W256 tumor cell survival by enhancing alphavbeta5 expression.

The metabolism of arachidonic acid (AA) leads to the generation of biologically active metabolites that have been implicated in cell growth and proliferation, as well as survival and apoptosis. We have previously demonstrated that rat Walker 256 (W256) carcinosarcoma cells express the platelet-type 12-lipoxygenase (12-LOX) and synthesize 12(S)- and 15(S)-HETE as their major LOX metabolites. Here we show that Walker 256 cells also express leukocyte-type 12-LOX and that its overexpression in these cells significantly extends their survival and delays apoptosis when cells are cultured under serum-free conditions. Under serum-free conditions, the expression of leukocyte-type 12-LOX is upregulated. 12-LOX-transfected W256 cells had a more spread morphology in culture compared with wild-type or mock-transfected cells. Examination of W256 cells showed that the cells expressed a number of integrins on their surface. Overexpression of 12-LOX enhanced the surface expression and focal adhesion localization of integrin alphavbeta5, while not affecting other integrins. Also, the 12-LOX-transfected W256 cells exhibited higher levels of microfilament content. Treatment of cells with monoclonal antibody to alphavbeta5 or cytochalasin B (a microfilament-disrupting agent), but not antibodies to other integrin receptors, resulted in significant apoptosis, characterized by rapid rounding up and detachment from the substratum. These results show that the 12-LOX pathway is a regulator of cell survival and apoptosis, by affecting the expression and localization of the alphavbeta5 integrin and actin microfilaments in Walker 256 cells.

Actins↗

PubMed2003

ZDOCK predictions for the CAPRI challenge.

The CAPRI Challenge is a blind test of protein-protein-docking algorithms that predict the complex structure from the crystal structures of the interacting proteins. We participated in both rounds of this blind test and submitted predictions for all seven targets, relying mainly on our Fast Fourier Transform based algorithm ZDOCK that combines shape complementarity, desolvation, and electrostatics. Our group made good predictions for three targets and had at least some success with three others. Implications of the treatment of prior biological information as well as contributions of manual inspection to docking predictions are also discussed.

Algorithms↗

PubMed2003

ZDOCK: an initial-stage protein-docking algorithm.

The development of scoring functions is of great importance to protein docking. Here we present a new scoring function for the initial stage of unbound docking. It combines our recently developed pairwise shape complementarity with desolvation and electrostatics. We compare this scoring function with three other functions on a large benchmark of 49 nonredundant test cases and show its superior performance, especially for the antibody-antigen category of test cases. For 44 test cases (90% of the benchmark), we can retain at least one near-native structure within the top 2000 predictions at the 6 degrees rotational sampling density, with an average of 52 near-native structures per test case. The remaining five difficult test cases can be explained by a combination of poor binding affinity, large backbone conformational changes, and our algorithm's strong tendency for identifying large concave binding pockets. All four scoring functions have been integrated into our Fast Fourier Transform based docking algorithm ZDOCK, which is freely available to academic users at http://zlab.bu.edu/~ rong/dock.

Algorithms↗

PubMed2003

Protein tyrosine nitration in the mitochondria from diabetic mouse heart. Implications to dysfunctional mitochondria in diabetes.

Oxidative stress has been implicated in dysfunctional mitochondria in diabetes. Tyrosine nitration of mitochondrial proteins was observed under conditions of oxidative stress. We hypothesize that nitration of mitochondrial proteins is a common mechanism by which oxidative stress causes dysfunctional mitochondria. The putative mechanism of nitration in a diabetic model of oxidative stress and functional changes of nitrated proteins were studied in this work. As a source of mitochondria, alloxan-susceptible and alloxan-resistant mice were used. These inbred strains are distinguished by the differential ability to detoxify free radicals. A proteomic approach revealed significant similarity between patterns of tyrosine-nitrated proteins generated in the heart mitochondria under different in vitro and in vivo conditions of oxidative stress. This observation points to a common nitrating species, which may derive from different nitrating pathways in vivo and may be responsible for the majority of nitrotyrosine formed. Functional studies show that protein nitration has an adverse effect on protein function and that protection against nitration protects functional properties of proteins. Because proteins that undergo nitration are involved in major mitochondrial functions, such as energy production, antioxidant defense, and apoptosis, we concluded that tyrosine nitration of mitochondrial proteins may lead to dysfunctional mitochondria in diabetes.

Alloxan↗

PubMed2003

Bone marrow-derived stem cells initiate pancreatic regeneration.

We show that transplantation of adult bone marrow-derived cells expressing c-kit reduces hyperglycemia in mice with streptozotocin-induced pancreatic damage. Although quantitative analysis of the pancreas revealed a low frequency of donor insulin-positive cells, these cells were not present at the onset of blood glucose reduction. Instead, the majority of transplanted cells were localized to ductal and islet structures, and their presence was accompanied by a proliferation of recipient pancreatic cells that resulted in insulin production. The capacity of transplanted bone marrow-derived stem cells to initiate endogenous pancreatic tissue regeneration represents a previously unrecognized means by which these cells can contribute to the restoration of organ function.

Animals↗

PubMed2003

T-linker-specific ligation PCR (T-linker PCR): an advanced PCR technique for chromosome walking or for isolation of tagged DNA ends.

Dozens of PCR-based methods are available for chromosome walking from a known sequence to an unknown region. These methods are of three types: inverse PCR, ligation-mediated PCR and randomly primed PCR. However, none of them has been generally applied for this purpose, because they are either difficult or inefficient. Here we describe a simple and efficient PCR strategy--T-linker-specific ligation PCR (T-linker PCR) for gene or chromosome walking. The strategy amplifies the template molecules in three steps. First, genomic DNA is digested with 3' overhang enzymes. Secondly, primed by a specific primer, a strand of the target molecule is replicated by Taq DNA polymerase and a single A tail is generated on the 3' unknown end of the target molecule, and then a 3' overhang-T linker (named T-linker) is specifically ligated onto the target. Thirdly, the target is amplified by two rounds of nested PCR with specific primers and T-linker primers. T-linker PCR significantly improves the existing PCR methods for walking because it uses specific T/A ligation instead of arbitrary ligation or random annealing. To show the feasibility and efficiency of T-linker PCR, we have exploited this method to identify vector DNA or T-DNA insertions in transgenic plants.

Arabidopsis↗

PubMed2003

Major kinetic traps for the oxidative folding of leech carboxypeptidase inhibitor.

The leech carboxypeptidase inhibitor (LCI) is a 66-amino acid protein, containing four disulfides that stabilize its structure. This polypeptide represents an excellent model for the study and understanding of the diversity of folding pathways in small, cysteine-rich proteins. The pathway of oxidative folding of LCI has been elucidated in this work, using structural and kinetic analysis of the folding intermediates trapped by acid quenching. Reduced and denatured LCI refolds through a rapid, sequential flow of one- and two-disulfide intermediates and reaches a rate-limiting step in which a mixture of three major three-disulfide species and a heterogeneous population of non-native four-disulfide (scrambled) isomers coexist. The three three-disulfide intermediates have been identified as major kinetic traps along the folding pathway of LCI, and their disulfide structures have been elucidated in this work. Two of them contain only native disulfide pairings, and one contains one native and two non-native disulfide bonds. The coexistence of three-disulfide kinetic traps adopting native disulfide bonds together with a significant proportion of fully oxidized scrambled isomers shows that the folding pathway of LCI features properties exhibited by both the bovine pancreatic trypsin inhibitor and hirudin, two diverse models with extreme folding characteristics. The results further demonstrate the large diversity of disulfide folding pathways.

Amino Acid Sequence↗

PubMed2003

Cloning and characterization of a novel gene which encodes a protein interacting with the mitosis-associated kinase-like protein NTKL.

NTKL is an evolutionarily conserved kinase-like protein. The cell-cycle-dependent centrosomal localization of NTKL suggested that it was involved in centrosome-related cellular function. The mouse NTKL protein is highly homologous with human NTKL. A novel mouse protein was identified as an NTKL-binding protein (NTKL-BP1) by yeast two-hybrid screening, and the full-length cDNA was amplified based on the result of a sequence data analysis cloning strategy. The full-length cDNA sequence of the NTKL-BP1 gene consists of 2,537 bp, which encode 368 amino acids. A database search revealed that homologues of NTKL-BP1 exist in different organisms, including Arabidopsis thaliana, Drosophila melanogaster, Plasmodium falciparum, Geobacter metallireducens, Anopheles gambiae and human. It suggests that NTKL-BP1 is an evolutionarily conserved protein. The expression of NTKL-BP1 was observed in multiple normal mouse tissues. The interaction of the two proteins was confirmed by co-immunoprecipitation. Moreover, immunofluorescent staining indicated that NTKL and NTKL-BP1 were all localized in the cytoplasm.

Amino Acid Sequence↗