Biomedical subjects

Yali Chen

Publications and source records attributed to Yali Chen.

At least 19 recordsLinked to original sources

PubMed2006

Internalization of CD40 regulates its signal transduction in vascular endothelial cells.

The CD40 ligand (CD40L)-CD40 dyad can ignite proinflammatory and procoagulatory activities of the vascular endothelium in the pathogenesis and progression of atherosclerosis. Besides being expressed on the activated CD4(+) T cell surface (mCD40L), the majority of circulating CD40L reservoir (sCD40L) in plasma is released from stimulated platelets. It remains debatable which form of CD40L triggers endothelial inflammation. Here, we demonstrate that the agonistic antibody of CD40 (G28.5), which mimics the action of sCD40L, induces rapid endocytosis of CD40 independent of TRAF2/3/6 binding while CD40L expressed on the surface of HEK293A cells captures CD40 at the cell conjunction. Forced internalization of CD40 by constitutively active mutant of Rab5 preemptively activates NF-kappaB pathway, suggesting that CD40 was able to form an intracellular signal complex in the early endosomes. Internalized CD40 exhibits different patterns of TRAF2/3/6 recruitment and Akt phosphorylation from the membrane anchored CD40 complex. Finally, mCD40L but not sCD40L induces the upregulation of proinflammatory cytokines and cell adhesion factors in the primary human vascular endothelial cells in vitro, although both forms of CD40L activate NF-kappaB pathway. These results therefore may help understand the molecular mechanism of CD40L signaling that contributes to the pathophysiology of atherosclerosis.

Animals↗

PubMed2006

Anti-CD3 and nasal proinsulin combination therapy enhances remission from recent-onset autoimmune diabetes by inducing Tregs.

Safe induction of autoantigen-specific long-term tolerance is the "holy grail" for the treatment of autoimmune diseases. In animal models of type 1 diabetes, oral or i.n. immunization with islet antigens induces Tregs that are capable of bystander suppression. However, such interventions are only effective early in the prediabetic phase. Here, we demonstrate that a novel combination treatment with anti-CD3epsilon-specific antibody and i.n. proinsulin peptide can reverse recent-onset diabetes in 2 murine diabetes models with much higher efficacy than with monotherapy with anti-CD3 or antigen alone. In vivo, expansion of CD25(+)Foxp3(+) and insulin-specific Tregs producing IL-10, TGF-beta, and IL-4 was strongly enhanced. These cells could transfer dominant tolerance to immunocompetent recent-onset diabetic recipients and suppressed heterologous autoaggressive CD8 responses. Thus, combining a systemic immune modulator with antigen-specific Treg induction is more efficacious in reverting diabetes. Since Tregs act site-specifically, this strategy should also be expected to reduce the potential for systemic side effects.

Administration, Intranasal↗

PubMed2006

Drusen complement components C3a and C5a promote choroidal neovascularization.

Age-related macular degeneration (AMD) is the leading cause of irreversible blindness in industrialized nations, affecting 30-50 million people worldwide. The earliest clinical hallmark of AMD is the presence of drusen, extracellular deposits that accumulate beneath the retinal pigmented epithelium. Although drusen nearly always precede and increase the risk of choroidal neovascularization (CNV), the late vision-threatening stage of AMD, it is unknown whether drusen contribute to the development of CNV. Both in patients with AMD and in a recently described mouse model of AMD, early subretinal pigmented epithelium deposition of complement components C3 and C5 occurs, suggesting a contributing role for these inflammatory proteins in the development of AMD. Here we provide evidence that bioactive fragments of these complement components (C3a and C5a) are present in drusen of patients with AMD, and that C3a and C5a induce VEGF expression in vitro and in vivo. Further, we demonstrate that C3a and C5a are generated early in the course of laser-induced CNV, an accelerated model of neovascular AMD driven by VEGF and recruitment of leukocytes into the choroid. We also show that genetic ablation of receptors for C3a or C5a reduces VEGF expression, leukocyte recruitment, and CNV formation after laser injury, and that antibody-mediated neutralization of C3a or C5a or pharmacological blockade of their receptors also reduces CNV. Collectively, these findings establish a mechanistic basis for the clinical observation that drusen predispose to CNV, revealing a role for immunological phenomena in angiogenesis and providing therapeutic targets for AMD.

Aged, 80 and over↗

PubMed2006

AFM study of the plastic deformation of polysynthetically-twinned (PST) TiAl crystals in soft orientation.

PST TiAl samples with a nominal composition of Ti52Al48 were deformed at room temperature with compression axis inclined to the lamellar interfaces by 45 degrees and one of the side surface normal directions set to be (112). The deformation structures on the free surfaces of the deformed samples were investigated using Atomic Force Microscope (AFM). It was found that in-plane shear (shear in planes parallel to lamellar interfaces) is the dominant deformation mode in all gamma domains and most of the deformation traces on the free surfaces are parallel to lamellar interfaces. Out-of-plane shear (shear in planes inclined to lamellar interfaces) also occurs but contributes much less to the macroscopic strain. This selective activation of deformation modes leads to a highly anisotropic deformation behavior in PST crystals with this orientation.

Aluminum↗

PubMed2006

Clinical features of autosomal dominant retinitis pigmentosa associated with a Rhodopsin mutation.

INTRODUCTION: Retinitis pigmentosa (RP) describes a group of inherited disorders characterised by progressive retinal dysfunction, cell loss and atrophy of retinal tissue. RP demonstrates considerable clinical and genetic heterogeneity, with wide variations in disease severity, progression, and gene involvement. We studied a large family with RP to determine the pattern of inheritance and identify the disease-causing mutation, and then to describe the phenotypic presentation of this family. MATERIALS AND METHODS: Ophthalmic examination was performed on 46 family members to identify affected individuals and to characterise the disease phenotype. Family pedigree was obtained. Some family members also had fundus photographs, fluorescein angiography, and/or optical coherence tomography (OCT) analysis performed. Genetic linkage was performed using short tandem repeat (STR) polymorphic markers encompassing the known loci for autosomal dominant RP. Finally, DNA sequencing was performed to identify the mutation present in this family. RESULTS: Clinical features included nyctalopia, constriction of visual fields and eventual loss of central vision. Sequence analysis revealed a G-to-T nucleotide change in the Rhodopsin gene, predicting a Gly-51-Val substitution. CONCLUSIONS: This large multi-generation family demonstrates the phenotypic variability of a previously identified autosomal dominant mutation of the Rhodopsin gene.

Adolescent↗

PubMed2006

A novel locus for X-linked retinitis pigmentosa.

INTRODUCTION: Retinitis pigmentosa (RP) is the most prevalent group of inherited retinopathies and demonstrates considerable clinical and genetic heterogeneity, with wide variations in disease severity, progression, and gene involvement. We studied a large family with RP to determine the pattern of inheritance and to identify the disease-causing gene/locus. MATERIALS AND METHODS: Ophthalmic examination was performed on 35 family members to identify affected individuals and carriers and to characterise the disease phenotype. Genetic linkage analysis was performed using short tandem repeat (STR) polymorphic markers encompassing the known loci for Xlinked RP (xlRP) including RP2, RP3, RP6, RP23, and RP24. Mutation screening was performed by direct sequencing of PCR-amplified genomic DNA of the RP2 and RPGR genes of the affected individuals. RESULTS: A highly penetrant, X-linked form of RP was observed in this family. Age of onset was from 5 to 8 years and visual acuity ranged from 20/25 in children to light perception in older adults. Linkage analysis and direct sequencing showed that no known loci/genes were associated with the phenotype in this kindred. CONCLUSION: A novel disease gene locus/loci is responsible for the xlRP phenotype in this family.

Adolescent↗

PubMed2005

Processivity factor of KSHV contains a nuclear localization signal and binding domains for transporting viral DNA polymerase into the nucleus.

Kaposi's sarcoma-associated human herpesvirus (KSHV) encodes a processivity factor (PF-8, ORF59) that forms homodimers and binds to viral DNA polymerase (Pol-8, ORF9). PF-8 is essential for stabilizing Pol-8 on template DNA so that Pol-8 can incorporate nucleotides continuously. Here, the intracellular interaction of these two viral proteins was examined by confocal immunofluorescence microscopy. When individually expressed, PF-8 was observed exclusively in the nucleus, whereas Pol-8 was found only in the cytoplasm. However, when co-expressed, Pol-8 was co-translocated with PF-8 into the nucleus. Mutational analysis revealed that PF-8 contains a nuclear localization signal (NLS) as well as domains located at the N-terminus and the C-proximal regions that are required for Pol-8 binding. This study suggests that the mechanism that enables PF-8 to transport Pol-8 into the nucleus is the first critical step required for Pol-8 and PF-8 to function processively in KSHV DNA synthesis.

Amino Acid Sequence↗

PubMed2005

Loss of ER retention and sequestration of the wild-type ELOVL4 by Stargardt disease dominant negative mutants.

PURPOSE: Mutations in ELOVL4, a member of the fatty acid elongase (ELO) family, are responsible for autosomal dominant Stargardt-like macular degeneration. The specific role of ELOVL4 in photoreceptors and the degenerative events induced by dominant ELOVL4 mutations are not well understood. As a first step to identifying possible mechanisms contributing to cellular dysfunction, we transfected HEK293 and COS cells with fluorescent-labeled wild-type and mutant ELOVL4 constructs. Effects of mutant ELOVL4 on interaction with wild-type protein were examined in this in vitro model. METHODS: Wild-type and mutant ELOVL4 proteins including ELOVL4 truncation (270X, a truncated ELOVL4 protein at amino acid position 270) and ELOVL4 5 bp deletion (5bp-del) and ELOVL4 (5A, substituting the ER retention signal, KAKGD, with a five alanine amino acid tract) were expressed as EGFP or DsRed fusion proteins. Cellular localization of these proteins was examined by fluorescence microscopy. ELOVL4 protein aggregates were measured by co-immunoprecipitation and by sucrose gradient centrifugation followed by immunodetection with western blots. To study cellular status of cells expressing mutant ELOVL4 proteins, transfected cells were examined for upregulation of Bip and CHOP, markers for the unfolded protein response (UPR) by western blotting. RESULTS: ELOVL4 mutants were not retained within the ER but were rather mislocalized and formed aggregates. Importantly, when cotransfected with wild-type ELOVL4, the mutants bound to and sequestered the wild-type protein into the aggregates. Expression of ELOVL4 mutants also induced UPR as evidenced by Bip and CHOP expression. CONCLUSIONS: Using this in vitro cell system, we have identified alterations in wild-type ELOVL4 protein localization, aggregate formation, and the induction of cellular stress by the ELOVL4 mutants. We propose that "inactivation" of the wild-type ELOVL4 protein through sequestration to a non-ER compartment by ELOVL4 mutants may play a role in cellular dysfunction.

Animals↗

PubMed2005

Assessment of a systematic expression profiling approach in ENU-induced mouse mutant lines.

Comparative genomewide expression profiling is a powerful tool in the effort to annotate the mouse genome with biological function. The systematic analysis of RNA expression data of mouse lines from the Munich ENU mutagenesis screen might support the understanding of the molecular biology of such mutants and provide new insights into mammalian gene function. In a direct comparison of DNA microarray experiments of individual versus pooled RNA samples of organs from ENU-induced mouse mutants, we provide evidence that individual RNA samples may outperform pools in some aspects. Genes with high biological variability in their expression levels (noisy genes) are identified as false positives in pooled samples. Evidence suggests that highly stringent housing conditions and standardized procedures for the isolation of organs significantly reduce biological variability in gene expression profiling experiments. Data on wild-type individuals demonstrate the positive effect of controlling variables such as social status, food intake before organ sampling, and stress with regard to reproducibility of gene expression patterns. Analyses of several organs from various ENU-induced mutant lines in general show low numbers of differentially expressed genes. We demonstrate the feasibility to detect transcriptionally affected organs employing RNA expression profiling as a tool for molecular phenotyping.

Animals↗

PubMed2005

Receptor for advanced glycation end products and its ligands: a journey from the complications of diabetes to its pathogenesis.

Many studies have suggested that the expression of RAGE (receptor for advanced glycation end products) is upregulated in human tissues susceptible to the long-term complications of diabetes. From the kidneys to the macrovessels of the aorta, RAGE expression is upregulated in a diverse array of cell types, from glomerular epithelial cells (podocytes) to endothelial cells, vascular smooth muscle cells, and inflammatory mononuclear phagocytes and lymphocytes. Although RAGE was first described as a receptor for advanced glycation end products (AGEs), the key finding that RAGE was also a signaling receptor for proinflammatory S100/calgranulins and amphoterin, led to the premise that even in euglycemia, ligand-RAGE interaction propagated inflammatory mechanisms linked to chronic cellular perturbation and tissue injury. Indeed, such considerations suggested that RAGE might even participate in the pathogenesis of type 1 diabetes. Our studies have shown that pharmacological and/or genetic deletion/mutation of the receptor attenuates the development of hyperglycemia in NOD mice; in mice with myriad complications of diabetes, interruption of ligand-RAGE interaction prevents or delays the chronic complications of the disease in both macro- and microvessel structures. Taken together, these findings suggest that RAGE is "at the right place and time" to contribute to the pathogenesis of diabetes and it complications. Studies are in progress to test the premise that antagonism of this interaction is a logical strategy for the prevention and treatment of diabetes.

Animals↗

PubMed2004

Spontaneous mutations in digestive tract of old mice show tissue-specific patterns of genomic instability.

In an attempt to evaluate the possible role of mutations in the age-dependent increase of tumor incidence, we studied the mutational burden that accumulates in the aging process in different parts of the digestive tract in mice. The mutations were monitored in lacZ genes integrated in the mouse genome. The digestive tract was divided into the esophagus, stomach, proximal, medial, and distal part of the small intestine, and the colon. Epithelial tissues were separated from these tissues with the exception of the esophagus, in which case the whole tissue was examined. At a young age, the mutant frequencies as well as the molecular nature of the mutations were similar among the tissues examined. In old age, on the other hand, mutant frequencies were elevated to different degrees among the tissues; they were high in the small intestine and colon, intermediate in the stomach, and low in the esophagus. The molecular characteristics of the mutations also revealed distinct tissue-specificity; there were elevated rates of a small deletion mutation in the esophagus, G:C to T:A transversion in the proximal small intestine, and multiple mutations in the distal small intestine and colon. The results indicate that different parts of the digestive tract suffer from different kinds of mutational stress in the aging process. The nature of the multiple mutations suggests the presence of a mutator phenotype based on an imbalance in deoxyribonucleotide pools.

Aging↗

PubMed2004

Tat-dependent repression of human immunodeficiency virus type 1 long terminal repeat promoter activity by fusion of cellular transcription factors.

Transcription initiation from HIV-1 long terminal repeat (LTR) promoter requires the virally encoded transactivator, Tat, and several cellular co-factors to accomplish the Tat-dependent processive transcription elongation. Individual cellular transcription activators, LBP-1b and Oct-1, on the other hand, have been shown to inhibit LTR promoter activities probably via competitive binding against TFIID to the TATA-box in LTR promoter. To explore the genetic interference strategies against the viral replication, we took advantage of the existence of the bipartite DNA binding domains and the repression domains of LBP-1b and Oct-1 factors to generate a chimeric transcription repressor. Our results indicated that the fusion protein of LBP-1b and Oct-1 exhibited higher DNA binding affinity to the viral promoter than the individual factors, and little interference with the host cell gene expression due to its anticipated rare cognate DNA sites in the host cell genome. Moreover, the chimera exerted increased Tat-dependent repression of transcription initiation at the LTR promoter both in vitro and in vivo compared to LBP-1b, Oct-1 or combination of LBP-1b and Oct-1. These results might provide the lead in generating a therapeutic reagent useful to suppress HIV-1 replication.

DNA-Binding Proteins↗

PubMed2004

Blockade of late stages of autoimmune diabetes by inhibition of the receptor for advanced glycation end products.

Ligation of the receptor for advanced glycation end products (RAGE) occurs during inflammation. Engagement of RAGE results in enhanced expression of addressins and it is therefore, not surprising that previous studies have shown a role of RAGE/ligand interactions in immune responses including cell/cell contact but the role of RAGE in spontaneous autoimmunity has not been clearly defined. To study the role of RAGE/ligand interactions in autoimmune diabetes, we tested the ability of soluble RAGE, a scavenger of RAGE ligands, in late stages of diabetes development in the NOD mouse-disease transferred with diabetogenic T cells and recurrent disease in NOD/scid recipients of syngeneic islet transplants. RAGE expression was detected on CD4(+), CD8(+), and B cells from diabetic mice and transferred to NOD/scid recipients. RAGE and its ligand, S100B, were found in the islets of NOD/scid mice that developed diabetes. Treatment of recipient NOD/scid mice with soluble RAGE prevented transfer of diabetes and delayed recurrent disease in syngeneic islet transplants. RAGE blockade was associated with increased expression of IL-10 and TGF-beta in the islets from protected mice. RAGE blockade reduced the transfer of disease with enriched T cells, but had no effect when diabetes was transferred with the activated CD4(+) T cell clone, BDC2.5. We conclude that RAGE/ligand interactions are involved in the differentiation of T cells to a mature pathogenic phenotype during the late stages of the development of diabetes.

Animals↗

PubMed2004

Papilin, a novel component of basement membranes, in relation to ADAMTS metalloproteases and ECM development.

Papilins are homologous, secreted extracellular matrix proteins which share a common order of protein domains. They occur widely, from nematodes to man, and can differ in the number of repeats of a given type of domain. Within one species the number of repeats can vary by differential RNA splicing. A distinctly conserved cassette of domains at the amino-end of papilins is homologous with a cassette of protein domains at the carboxyl-end of the ADAMTS subgroup of secreted, matrix-associated metalloproteases. Papilins primarily occur in basement membranes. Papilins interact with several extracellular matrix components and ADAMTS enzymes. Papilins are essential for embryonic development of Drosophila melanogaster and Caenorhabditis elegans.

Animals↗

PubMed2004

Analysis of mouse germ-cell transcriptome at different stages of spermatogenesis by SAGE: biological significance.

The transcriptomes of mouse type A spermatogonia (Spga), pachytene spermatocytes (Spcy), and round spermatids (Sptd) were determined by sequencing the respective SAGE (Serial Analysis of Gene Expression) libraries. A total of 444,015 tags derived from one Spga, two Spcy, and one Sptd library were analyzed, and 34,619 different species of transcripts were identified, 5279 of which were novel. Results indicated the germ-cell transcriptome comprises of more than 30,000 transcripts. Virtual subtraction showed that cell-specific transcripts constitute 12-19.5% of the transcriptome. Components of the protein biosynthetic machinery are highly expressed in Spga. In Spcy transcription factors are abundantly expressed while transcripts encoding proteins involved in chromosome remodeling and testis-specific transcripts are prominent in Sptd. The databases generated by this work provide very useful resources for cellular localization of genes in silico. They are also extremely useful as sources for identification of splice variants of genes in germ cells.

Animals↗

PubMed2004

Properties of radicals formed by the irradiation of wool fibers.

Wool fibers of different sample conditions were irradiated in different atmospheres by (60)Co gamma-rays and were studied by electron spin resonance method (ESR). It was found that a large percentage of the alpha-carbon radicals of polymer main chain were more long-lived radicals. The ESR measurements of irradiated cortex samples of the wool fibers proved that most radicals from the cortex were long-lived ones. Low water content (as low as 27.5%) in the reaction system did not greatly affect the radical formation, but higher water contents would reduce the radical concentrations dramatically and accelerate their decaying process. The results will be of help in property modification of wool products by radiation graft copolymerization.

Animals↗

PubMed2003

[Study on the properties of methyl parathion hydrolase from Pseudomonas sp. WBC-3].

A methyl parathion degradation enzyme, or methyl parathion hydrolase (MPH, EC 3.1.8.1), locating in the soluble intracellular fraction of Pseudomonas sp. WBC-3, was purified 49.1-fold to homogeneity by one-step ion exchange chromatography. The physical and chemical properties of the purified MPH were studied. The purified MPH displayed relatively broad optimal temperature around 40 degrees. The activity of MPH was affected by pH and the optimal pH was 11.0. Cd2+ and Fe2+ could enhance the catalytic efficiency of MPH while Hg2+, Zn2+, Al3+ and Bi3+ showed inhibition effect. With methyl parathion as the optimal substrate, the Km was 0.0807mmol/L and the kcat was 2.1 x 10(6) min(-1). In addition, the comparison of native and subunit molecular weights of MPH suggested that this enzyme was a monomer of approximate 34kD.

Enzyme Stability↗

PubMed2002

Generation and in vitro differentiation of a spermatogonial cell line.

Spermatogenesis is the process by which spermatogonial stem cells divide and differentiate to produce sperm. In vitro sperm production has been difficult to achieve because of the lack of a culture system to maintain viable spermatogonia for long periods of time. Here we report the in vitro generation of spermatocytes and spermatids from telomerase-immortalized mouse type A spermatogonial cells in the presence of stem cell factor. This differentiation can occur in the absence of supportive cells. The immortalized spermatogonial cell line may serve as a powerful tool in elucidating the molecular mechanisms of spermatogenesis. Furthermore, through genomic modification and transplantation techniques, this male germ cell line may be used to generate transgenic mice and to develop germ cell gene therapy.

Acrosin↗