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Biomedical subjects

L Wu

Publications and source records attributed to L Wu.

At least 73 records · Page 4Linked to original sources
PubMed2001

Adaptation to blockade of human immunodeficiency virus type 1 entry imposed by the anti-CCR5 monoclonal antibody 2D7.

The second extracellular loop (ECL2) domain of CC-chemokine receptor 5 (CCR5) has been proposed as a specific target site for therapeutic agents aimed at blocking CCR5-dependent entry by human immunodeficiency virus type I (HIV-1). We have adapted two CCR5-using HIV-1 isolates, prototypic JR-CSF, and a primary isolate, 11-121, to replicate in vitro in the presence of high concentrations of a monoclonal antibody (MAb 2D7) specific for the CCR5 ECL2 domain. The 75% inhibitory concentrations (IC(75)) for the two 2D7-adapted isolates were approximately 100-fold higher than those for corresponding control isolates passaged without the MAb. Adapted isolates did not acquire the ability to use CXCR4, CCR3, or CCR1. Env clones derived from MAb 2D7-adapted JR-CSF showed several gp120 mutations that were not found in any of the control JR-CSF clones. The in vitro observations suggest that CCR5-using HIV-1 strains might also be able to adapt in vivo to evade an ECL2-blocking therapeutic agent.

Adaptation, Biological↗
PubMed2001

Utilization of a peptide lead for the discovery of a novel PTP1B-binding motif.

Examination of the PTP1B inhibitory potency of an extensive series of phosphotyrosyl (pTyr) mimetics (Xxx) expressed in the EGFr-derived hexapeptide platform Ac-Asp-Ala-Asp-Xxx-Leu-amide previously led to the finding of high inhibitory potency when Xxx = 4-(phosphonodifluoromethyl)phenylalanyl (F2Pmp) (K(i) = 0.2 microM) and when Xxx = 3-carboxy-4-carboxymethyloxyphenylalanyl (K(i) = 3.6 microM). In the first instance, further work led from the F2Pmp-containing peptide to monomeric inhibitor, 6-(phosphonodifluoromethyl)-2-naphthoic acid (K(i) = 22 microM), and to the pseudo-dipeptide mimetic, N-[6-(phosphonodifluoromethyl)-2-naphthoyl]-glutamic acid (K(i) = 12 microM). In the current study, a similar approach was applied to the 3-carboxy-4-carboxymethyloxyphenylalanyl-containing peptide, which led to the preparation of monomeric 5-carboxy-6-carboxymethyloxy-2-naphthoic acid (K(i) = 900 microM). However, contrary to expectations based on the aforementioned F2Pmp work, incorporation of this putative pTyr mimetic into the pseudo-dipeptide, N-[5-carboxy-6-carboxymethyloxy-2-naphthoyl]-glutamic acid, resulted in a substantial loss of binding affinity. A reevaluation of binding orientation for 5-carboxy-6-carboxymethyloxy-2-naphthoic acid was therefore undertaken, which indicated a 180 degrees reversal of the binding orientation within the PTP1B catalytic site. In the new orientation, the naphthyl 2-carboxyl group, and not the o-carboxy carboxymethyloxy groups, mimics a phosphoryl group. Indeed, when 5-carboxy-2-naphthoic acid itself was examined at neutral pH for inhibitory potency, it was found to have K(i) = 31 +/- 7 microM, which is lower than parent 5-carboxy-6-carboxymethyloxy-2-naphthoic acid. In this fashion, 5-carboxy-2-naphthoic acid (or more appropriately, 6-carboxy-1-naphthoic acid) has been identified as a novel PTP1B binding motif.

Catalytic Domain↗
PubMed2001

Cleavage of the Bloom's syndrome gene product during apoptosis by caspase-3 results in an impaired interaction with topoisomerase IIIalpha.

In higher eukaryotes, the integration of signals triggered in response to certain types of stress can result in programmed cell death. Central to these events is the sequential activation of a cascade of proteinases known as caspases. The final activated effector caspases of this cascade digest a number of cellular proteins, in some cases increasing their enzymatic activity, in others destroying their function. Of the proteins shown to be targets for caspase-mediated proteolysis, a surprisingly large proportion are proteins involved in the signalling or repair of DNA damage. Here we investigate whether BLM, the product of the gene mutated in Bloom's syndrome, a human autosomal disease characterised by cancer predisposition and sunlight sensitivity, is cleaved during apoptosis. BLM interacts with topoisomerase IIIalpha and has been proposed to play an important role in maintaining genomic integrity through its roles in DNA repair and replication. We show that BLM is cleaved during apoptosis by caspase-3 and reveal that the main cleavage site is located at the junction between the N-terminal and central helicase domains of BLM. Proteolytic cleavage by caspase-3 produces a 120 kDa fragment, which contains the intact helicase domain and three smaller fragments, the relative amounts of which depend on time of incubation with caspase-3. The 120 kDa fragment retains the helicase activity of the intact BLM protein. However, its interaction with topoisomerase IIIalpha is severely impaired. Since the BLM-topoisomerase interaction is believed to be necessary for many of the replication and recombination functions of BLM, we suggest that caspase-3 cleavage of BLM could alter the localisation and/or function of BLM and that these changes may be important in the process of apoptosis.

Adenosine Triphosphatases↗
PubMed2001

Characterization of thymus-seeding precursor cells from mouse bone marrow.

The nature of the cells that seed the thymus of an irradiated recipient after intravenous (IV) transfer of bone marrow (BM) cells was investigated using 2 approaches. First, direct entry of a small number of donor BM cells into the thymus was tracked using a Ly-5 marker. Second, secondary IV transfer of the seeded thymus cells into a secondary recipient was used as an assay for precursor activity. A range of cell types was found to enter the recipient thymus initially, including B-lineage cells and myeloid cells, but T precursors were undetectable by flow cytometry over the first few days. Although all cells initially entering the thymus proliferated, no sustained thymus reconstitution was seen until day 4, when recognizable T-lineage precursors began to appear. The secondary transfer assays revealed the presence of lymphoid precursors in the recipient thymus, including T, NKT, NK, and B precursor activity, with a notable early burst of B-lineage generative capacity. There was no evidence of sustained myeloid precursor or multipotent stem cell activity, even though these were seen if BM cells were injected directly into the recipient thymus rather than introduced into the bloodstream. It is concluded that even though many cell types may initially enter an irradiated thymus, the thymus acts as a sieve, allowing lymphoid precursors, but not multipotent stem cells, to seed the environmental niches that permit selected precursor cell development and thymus reconstitution. (Blood. 2001;98:696-704)

Animals↗
PubMed2001

Mature dendritic cells prime functionally superior melan-A-specific CD8+ lymphocytes as compared with nonprofessional APC.

Priming of melan-A(26/27-35)-specific CTL occurs only in a fraction of late stage melanoma patients, whereas during the early stages of the disease and in healthy volunteers, melan-A CTL have functional and phenotypic markers consistent with a naive phenotype. To study the requirements for expansion of naive melan-A CTL from healthy donors, we set up an in vitro priming protocol and, using tetramer assays, we demonstrate that the activity and phenotype of the expanded melan-A CTL are profoundly influenced by the type of APC used. Priming by nonprofessional APC leads to expansion of melan-A CTL with reduced cytolytic activity and low level of IFN-gamma secretion. In contrast, mature dendritic cells (DC) expand cytolytic and IFN-gamma-producing melan-A CTL. Priming by mature DC is also efficient at low peptide concentration and requires only one round of stimulation. Finally, we observed that a significant fraction of CD45RO(+) melan-A CTL primed by mature DC expresses high levels of the homing receptor CD62L, whereas CTL primed by nonprofessional APC express CD62L in lower percentages and at lower levels. These results suggest that suboptimal priming by nonprofessional APC could account for the presence in vivo of dysfunctional cells and strongly support the immunotherapeutic use of mature DC for expansion of effector and memory Ag-specific CTL.

Antigen-Presenting Cells↗
PubMed2001

The distribution and expression of the Bloom's syndrome gene product in normal and neoplastic human cells.

Bloom's syndrome (BS) is an autosomal recessive disorder associated with a predisposition to cancers of all types. Cells from BS sufferers display extreme genomic instability. The BS gene product, BLM, is a 159 kDa DNA helicase enzyme belonging to the RecQ family. Here, we have analysed the distribution of BLM in normal and tumour tissues from humans using a recently characterized, specific monoclonal antibody. BLM was found to be localized to nuclei in normal lymphoid tissues, but was largely absent from other normal tissues analysed with the exception of the proliferating compartment of certain tissues. In contrast, expression of BLM was observed in a variety of tumours of both lymphoid and epithelial origin. A strong correlation was observed between expression of BLM and the proliferative status of cells, as determined by staining for markers of cell proliferation (PCNA and Ki67). We conclude that BLM is a proliferation marker in normal and neoplastic cells in vivo, and, as a consequence, is expressed at a higher level in tumours than in normal quiescent tissues.

Adenosine Triphosphatases↗
PubMed2001

Induction of human matrix metalloproteinase-12 gene transcriptional activity by GM-CSF requires the AP-1 binding site in human U937 monocytic cells.

Matrix metalloproteinase-12 (MMP-12) is critical for the migration of monocytes/macrophages into inflammatory sites through the basement membranes. We previously reported that MMP-12 expression was initially induced by granulocyte macrophage colony-stimulating factor (GM-CSF) in human peripheral blood monocytes and U937 monocytic cells. To further elucidate the molecular mechanism for the regulation of MMP-12 expression by GM-CSF in monocytes, we determined the sequence requirements for the MMP-12 gene transcriptional response of U937 monocytic cells to GM-CSF by using luciferase reporter and electrophoretic mobility shift assays. A series of 5'-deletion and site-directed mutation of the human MMP-12 promoter demonstrated that an AP-1 site spanning the -81 to -75-bp region is critical for the induction of MMP-12 promoter activity by GM-CSF. The electrophoretic mobility shift assay revealed that AP-1 binding activity was increased by GM-CSF treatment and that the AP-1 complex induced by GM-CSF consisted of multiple Jun and Fos isoforms. These results indicate that MMP-12 expression in U937 monocytes was initially induced by GM-CSF through the AP-1 binding activity.

5' Untranslated Regions↗
PubMed2001

Tumor necrosis factor receptor-associated factor (TRAF)2 represses the T helper cell type 2 response through interaction with NFAT-interacting protein (NIP45).

Recently we have identified a novel protein NIP45 (nuclear factor of activated T cells [NFAT]-interacting protein) which substantially augments interleukin (IL)-4 gene transcription. The provision of NIP45 together with NFAT and the T helper cell type 2 (Th2)-specific transcription factor c-Maf to cells normally refractory to IL-4 production, such as B cells or Th1 clones, results in substantial IL-4 secretion to levels that approximate those produced by primary Th2 cells. In studies designed to further our understanding of NIP45 activity, we have uncovered a novel facet of IL-4 gene regulation. We present evidence that members of the tumor necrosis factor receptor-associated factor (TRAF) family of proteins, generally known to function as adapter proteins that transduce signals from the tumor necrosis factor receptor superfamily, contribute to the repression of IL-4 gene transcription and that this effect is mediated through their interaction with NIP45.

Animals↗
PubMed2001

Patients with pseudomyxoma peritonei associated with disseminated peritoneal adenomucinosis have a significantly more favorable prognosis than patients with peritoneal mucinous carcinomatosis.

BACKGROUND: Pseudomyxoma peritonei (PMP) is a poorly understood condition characterized by disseminated intraperitoneal mucinous tumors, often with mucinous ascites. The term PMP has been applied historically as a pathologic diagnostic term to both benign and malignant mucinous neoplasms that produce abundant extracellular mucin, resulting in a variable and poorly predictable prognosis. A recent study reported a pathologic classification that separated patients into prognostically distinct groups, but the follow-up was relatively short. METHODS: Long-term follow-up data were analyzed for a previously reported series of 109 patients with PMP to examine the prognostic utility of a pathologic classification system that divided patients into three groups: disseminated peritoneal adenomucinosis (DPAM), peritoneal mucinous carcinomatosis (PMCA), and peritoneal mucinous carcinomatosis with intermediate or discordant features (PMCA-I/D). Patients whose tumors were classified 25 DPAM (n = 65 patients) had disease that was characterized by histologically bland to low-grade adenomatous mucinous epithelium associated with abundant extracellular mucin and fibrosis, often with an identifiable appendiceal mucinous adenoma that was the source of the peritoneal lesions. Patients whose tumors were classified 25 PMCA (n = 30 patients) had disease that was characterized by peritoneal lesions that displayed the cytologic and architectural features of mucinous carcinoma associated with extracellular mucin, often with an identifiable invasive mucinous adenocarcinoma of the gastrointestinal tract. Patients whose tumors were classified 25 PMCA-I (n = 11 patients) had peritoneal lesions that combined the features of DPAM and PMCA derived from well differentiated mucinous adenocarcinomas associated with adenomas. Patients whose tumors were classified 25 PMCA-D (n = 3 patients) had markedly atypical appendiceal adenomas associated with peritoneal lesions similar to PMCA. RESULTS: Patients with DPAM had 5-year and 10-year survival rates of 75% and 68%, respectively (mean follow-up, 96 months; median follow-up, 104 months). Patients with PMCA and PMCA-I/D had a significantly worse prognosis, with 5-year and 10-year survival rates, respectively, of 50% and 21% for PMCA-I/D (mean follow-up, 58 months; median follow-up, 51 months) and 14% and 3% for PMCA (mean follow-up, 27 months; median follow-up, 16 months; P = 0.0001). CONCLUSIONS: The term PMP should be used only as a clinical descriptor for patients who have the syndrome of mucinous ascites accompanied by a characteristic distribution of peritoneal mucinous tumors with the pathologic features of DPAM. DPAM should be used as a pathologic diagnostic term for patients with the bland peritoneal mucinous tumors associated with ruptured appendiceal mucinous adenomas and PMP. These patients should not be diagnosed with carcinoma, because they have disease that is distinct pathologically and prognostically from PMCA.

Adenocarcinoma, Mucinous↗
PubMed2001

CD8 T cells specific for human immunodeficiency virus, Epstein-Barr virus, and cytomegalovirus lack molecules for homing to lymphoid sites of infection.

CD8 T cells are classified as naïve, effector, or memory cells on the basis of CD45RA, CD62L, and CCR7 expression. Sequential engagement of cell-surface CD62L and CCR7 receptors is required for efficient trafficking to lymphoid tissue by means of high endothelial venules. Naïve CD8 T cells are CCR7(+)CD62L(+) CD45RA(+), whereas long-term memory cells are CCR7(+)CD62L(+)CD45RA(-). Effector cytotoxic T cells are thought to be CCR7(-)CD45RA(+). The distribution of CD8 subsets and cytolytic protein expression in healthy donors and donors seropositive for human immunodeficiency virus (HIV) were compared. In HIV-infected subjects, CCR7(-) CD8 T cells expanded at the expense of naïve and long-term memory cells. In both healthy donors and HIV-infected donors, CCR7(+) CD8 T cells were uniformly negative for perforin. In all subsets, perforin and granzyme A were not coordinately expressed, with perforin expression being more tightly regulated. The properties of CD8 T cells specific for cytomegalovirus, Epstein-Barr virus (EBV), and HIV were studied by staining with major histocompatibility complex peptide tetramers. Antigen-specific cells for chronic infections with these viruses were uniformly CCR7(-) and predominantly CD62L(-). In 2 HIV-seropositive donors, 3- to 4-fold fewer EBV-tetramer-positive cells were present in lymph nodes compared with blood. Antigen-specific CD8 T cells are therefore preferentially excluded from lymphoid sites, even when infection is primarily in lymphoid tissue. This may protect lymphoid tissues from immunopathological changes but compromise immune defense against viruses, such as HIV and EBV, that target lymphocytes. HIV-specific CD8 T cells do not express CD45RA, whereas EBV- and CMV-specific CD8 T cells are heterogeneous in CD45RA(+) expression. Lack of CD45RA expression may indicate incomplete differentiation of HIV-specific CD8 T cells to cytotoxic T cells.

CD8-Positive T-Lymphocytes↗
PubMed2001

Memory T cells constitute a subset of the human CD8+CD45RA+ pool with distinct phenotypic and migratory characteristics.

Using HLA class I-viral epitope tetramers to monitor herpes virus-specific CD8(+) T cell responses in humans, we have shown that a significant fraction of responding cells revert from a CD45RO(+) to a CD45RA(+) state after priming. All tetramer-binding CD45RA(+) cells, regardless of epitope specificity, expressed a phenotype LFA-1(high)CCR7(low) that was stable for at least 10 years in infectious mononucleosis patients and indefinitely in asymptomatic carriers. CD8(+)CD45RA(+)LFA-1(high) cells were not present in cord blood but in adults account for up to 50% of CD8(+)CD45RA(+) cells. These CD45RA(+)LFA-1(high) cells have significantly shorter telomeres than CD45RA(+)LFA-1(low) cells, suggesting that the latter represent a naive population, while the former are memory cells. CD45RA(+) memory cells are a stable population of noncycling cells, but on stimulation they are potent producers of IFN-gamma, while naive CD8(+) cells produce only IL-2. The chemokine receptor profile and migratory potential of CD45RA(+) memory cells is very similar to CD45RO(+) cells but different to naive CD8 cells. In accord with this, CD45RA(+) memory cells were significantly underrepresented in lymph nodes, but account for virtually all CD8(+)CD45RA(+) T cells in peripheral tissues of the same individuals.

Apoptosis↗
PubMed2001

A multiple imputation method for missing covariates in non-linear mixed-effects models with application to HIV dynamics.

We propose a three-step multiple imputation method, implemented by Gibbs sampler, for estimating parameters in non-linear mixed-effects models with missing covariates. Estimates obtained by the proposed multiple imputation method are compared to those obtained by the mean-value imputation method and the complete-case method through simulations. We find that the proposed multiple imputation method offers smaller biases and smaller mean-squared errors for the estimates of covariate coefficients compared to other two methods. We apply the three missing data methods to modelling HIV viral dynamics from an AIDS clinical trial. We believe that the results from the proposed multiple imputation method are more reliable than that from the other two commonly used methods.

Antiretroviral Therapy, Highly Active↗
PubMed2001

Subspecialization of CXCR5+ T cells: B helper activity is focused in a germinal center-localized subset of CXCR5+ T cells.

The T helper (Th) cell pool is composed of specialized cells with heterogeneous effector functions. Apart from Th1 and 2 cells, CXCR5+ T cells have been suggested to be another type of effector T cell specialized for B cell help. We show here that CXCR5+ T cells are heterogeneous, and we identify subsets of CXCR5+ CD4 T cells that differ in function and microenvironmental localization in secondary lymphoid tissues. CD57+CXCR5 T cells, hereafter termed germinal center Th (GC-Th) cells, are localized only in GCs, lack CCR7, and are highly responsive to the follicular chemokine B lymphocyte chemoattractant but not to the T cell zone EBI1-ligand chemokine. Importantly, GC-Th cells are much more efficient than CD57-CXCR5+ T cells or CXCR5- T cells in inducing antibody production from B cells. Consistent with their function, GC-Th cells produce elevated levels of interleukin 10 upon stimulation which, with other cytokines and costimulatory molecules, may help confer their B cell helper activity. Our results demonstrate that CXCR5+ T cells are functionally heterogeneous and that the GC-Th cells, a small subset of CXCR5+ T cells, are the key helpers for B cell differentiation and antibody production in lymphoid tissues.

B-Lymphocytes↗
PubMed2001

Study on the abundance of CFCs varying with the latitude at the bottom of the troposphere in the southern hemisphere.

As the main ozone depleting compounds, the abundance of CFCs in the troposphere and stratosphere has been focused on all the while. However, there are few reports concerning on the CFCs' latitudinal dependence. Relying on the longterm monitoring that has been performed in Shanghai from 1997 and Cooperating with the Chinese Third Antarctic Inland Ice-sheet Traversal Party, we did intensive sampling in the southern hemisphere from the beginning of November to the end of December in 1998 and then got the corresponding concentrations of CFC-12, CFC-11, and CFC-113 according to the latitude. The data were studied inductively in accordance with the geographical location and meteorological condition of every sampling point. In summary, the average concentrations of CFCs (refer to CFC-11, CFC-12, and CFC-113) in equatorial area are the highest in the southern hemisphere; however, the concentrations in prevailing west-wind belt and polar east-wind belt are much lower and at the same levels, which should be a perfect implication of the mean background in the southern hemisphere. Being related to the special polar circumfluence, the concentrations of CFCs on the glacier reveal a special rise. Compared with the mean background of the southern hemisphere that is represented by the data of the prevailing west-wind belt and Polar east-wind belt, the concentrations in Shanghai bear the same level.

Air Movements↗
PubMed2001

Unique subpopulations of CD56+ NK and NK-T peripheral blood lymphocytes identified by chemokine receptor expression repertoire.

CD56, an adhesion molecule closely related to neural cell adhesion molecule, is an immunophenotypic marker for several unique populations of PBLS: Although CD56(+) cells derive from multiple lymphocyte lineages, they share a role in immunosurveillance and antitumor responses. We have studied the chemokine receptor expression patterns and functional migratory responses of three distinct CD56(+) populations from human peripheral blood. NK-T cells were found to differ greatly from NK cells, and CD16(+) NK cells from CD16(-) NK cells. CD16(+) NK cells were the predominant population responding to IL-8 and fractalkine, whereas NK-T cells were the predominant population responding to the CCR5 ligand macrophage-inflammatory protein-1beta. CD16(-) NK cells were the only CD56(+) population that uniformly expressed trafficking molecules necessary for homing into secondary lymphoid organs through high endothelial venule. These findings describe a diverse population of cells that may have trafficking patterns entirely different from each other, and from other lymphocyte types.

Adult↗
PubMed2001

Synthesis and structure-activity relationships of naphthamides as dopamine D3 receptor ligands.

A series of naphthamides were synthesized, and the affinities of these compounds were determined for dopamine D2 and D3 receptors using radioligand binding techniques. The naphthamide compounds that were prepared include N-(1-alkylpiperidin-4-yl)-4-bromo-1-methoxy-2-naphthamides (1-6), (S)-N-(1-alkylpyrrolidin-3-yl)-4-bromo-1-methoxy-2-naphthamides (7-12), (R)-N-(1-alkylpyrrolidin-3-yl)-4-bromo-1-methoxy-2-naphthamides (13-18), (S)-N-(1-alkyl-2-pyrrolidinylmethyl)-4-bromo-1-methoxy-2-naphthamides (19-25), (R)-N-(1-alkyl-2-pyrrolidinylmethyl)-4-bromo-1-methoxy-2-naphthamides (26-31), and N-(9-alkyl-9-azabicyclo[3.3.1]nonan-3beta-yl)-4-bromo-1-methoxy-2-naphthamides (32, 33). The results of in vitro radioligand binding studies indicated that the majority of the naphthamide analogues bound with high affinity at both the D2 and D3 dopamine receptor subtypes and most of the compounds demonstrated some selectivity for the dopamine D3 dopamine receptor subtype. These results demonstrated that both the structure of the central amine moiety (piperidine, pyrrolidine, and 9-azabicyclo[3.3.1]nonane) ring and the N-(alkyl) substitution on the amine significantly effects the binding affinity at D2 and D3 dopamine receptors. The bulkiness of the N-(1-alkyl) substituent was found to (a) have no effect on pharmacologic selectivity, (b) increase the affinity at D3 receptors, or (c) decrease the affinity at D2 receptors. The most potent analogue in this series was (S)-N-(1-cycloheptylpyrrolidin-3-yl)-4-bromo-1-methoxy-2-naphthamide (10), which had equilibrium dissociation (K(i)) values of 1.8 and 0.2 nM for D2 and D3 receptors, respectively. The most selective analogue was (R)-N-(1-cycloheptyl-2-pyrrolidinylmethyl)-4-bromo-1-methoxy-2-naphthamide (30), which had K(i) values of 62.8 and 2.4 nM for D2 and D3 receptors, respectively. Radioligand binding results for sigma receptors indicated that the structure of the amine moiety and the N-(1-alkyl) substitutions also significantly influence the affinity and selectivity of these compounds at the sigma1 and sigma2 sigma receptor subtypes. The two naphthamides containing a 9-azabicyclo[3.3.1]nonan-3beta-yl central ring were found to be selective for sigma2 receptors.

Animals↗