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Biomedical subjects

Ying Lin

Publications and source records attributed to Ying Lin.

At least 37 records · Page 2Linked to original sources
PubMed2005

[Static pressure induces vascular smooth muscle cells proliferation and apoptosis].

OBJECTIVE: To study the role of static pressure on proliferation and apoptosis of the vascular smooth muscle cells. METHODS: Vascular smooth muscle cell line A10 were cultured at normal atmosphere, 80 mmHg, 100 mmHg, 200 mmHg static pressure respectively. Cell viability was determined by MTT assay, cell cycle and apoptosis rate were analyzed by flow cytometer. RESULTS: After exposure to lower static pressure (100 mmHg) for 48 hour, cell viability and proliferation index (PI) of A10 cell significantly increased, the percentage of A10 cell in G(0)/G(1) phase decreased significantly and the apoptosis rate showed no difference as compared with those exposed to atmospheric pressures. However after exposure of A10 cell to higher static pressure (200 mmHg) for 48 hours, cell viability and proliferation index (PI) significantly decreased, the percentage of A10 cell in G(0)/G(1) phase increased significantly, apoptosis rate significantly increased, as compared with cells exposed to atmospheric pressure. CONCLUSION: Lower static pressure can facilitate vascular smooth muscle cells proliferation, while higher pressure can induce cell apoptosis.

Animals↗
PubMed2005

Prostaglandin E2 induces FOXP3 gene expression and T regulatory cell function in human CD4+ T cells.

Naturally occurring CD4+CD25+ regulatory T cells (T reg) are pivotal in suppressing immune responses and maintaining tolerance. The identification of molecules controlling T reg differentiation and function is important in understanding host immune responses in malignancy and autoimmunity. In this study we show that PGE2 enhances the in vitro inhibitory function of human purified CD4+CD25+ T reg cells. Moreover, PGE2 induces a regulatory phenotype in CD4+CD25- T cells. PGE2-treated T cell-mediated inhibition of anti-CD3-stimulated lymphocyte proliferation did not require cell contact. Phenotypic analysis revealed that PGE2 diminished CD25 expression in both CD4+CD25dim T cells and CD4+CD25bright T reg cells. PGE2 exposure induced the T reg cell-specific transcription factor forkhead/winged helix transcription factor gene (FOXP3) in CD4+CD25- T cells and significantly up-regulated its expression in CD4+CD25+ T reg cells. Similarly, 24-h incubation with supernatants from cyclooxygenase-2-overexpressing lung cancer cells that secrete high levels of PGE2 significantly induced FOXP3 in CD4+CD25- T cells. Finally, PGE2 up-regulated FOXP3 at both mRNA and protein levels and enhanced FOXP3 promoter activity. This is the first report indicating that PGE2 can modulate FOXP3 expression and T reg function in human lymphocytes.

Adjuvants, Immunologic↗
PubMed2005

[Angiogenic effect of interleukin-8 in breast cancer and its association with estrogen receptor].

OBJECTIVE: To explore the angiogenic effect of interleukin-8 (IL-8) in breast cancer and its association with estrogen receptor (ER). METHODS: The supernatants of culture liquid of breast cancer cells of different lines with high expression of Il-8 (MDA-MB-231 and MDA-MB-157), moderate expression of Il-8 (SKBr-3), or low expression of Il-8 (T47D and ZR75-1) were collected. These different conditioned media and human umbilical cord vein endothelial cells (HUVECs) were used in cell migration test to calculate the number of migrating HUVECs. Neutralizing antibody of IL-8 was added into above supernatants to observe the change in HUVECs migration. Mouse-tail collagen was added into the 12-well plate and then solidified, HUVECs were added thereon, and different supernatants were used as culture fluid so as to observe the angiogenesis of HUVECs on the collagen. HUVECs were cultured in the supernatants of culture liquid of different breast cells and CyQUANT dye was added at different time points so as to observe the proliferation of HUVECs by CCD imaging system. The supernatants of the culture fluid of the MDA-MB-231, SKBr-3, or T47D cells with or without fibroblast growth factor (FGF)-2 were injected subcutaneously into mice. Five days later the mice were killed to strip the skin to observe the angiogenesis microscopically. Supernatant chip test was made to measure the concentrations of IL-8 in different supernatants. Estrogen receptor (ER)-alpha and pN1481 Luc containing IL-8 promoter or p Luc0 not containing IL-8 promoter were transfected into different human breast cancer cells. Dual-luciferase assay was performed to study the regulation of IL-8 level by ER. RESULTS: The numbers of migrating HUVECs cultured in the supernatants of MDA-MB-231 cells, SKB-Br-3 cells, and T47D cells were 7800 +/- 368, 6510 +/- 419, and 3470 +/- 297 respectively (P < 0.05). After the addition of IL-8 neutralizing antibody, the number of migrating HUVECs cultured in the supernatant of MDA-MB-231 cells was 4700 +/- 233 and 4900 +/- 328 respectively, both significantly lower than those before the addition (both P < 0.05). In comparison with those cultured in the supernatants of the breast cancer cells expressing lower level of IL-8, the HUVECs cultured in the supernatants of the breast cancer cells expressing higher level of IL-8 tended to form more microangioid structure and proliferate more rapidly (P < 0.05). The skin of the mice injected with the supernatants of the breast cancer cells expressing higher level of IL-8 showed more blood vessel formation. Transient transfection test showed that the IL-8 level of the MDA-MB-231 cells transfected with ER was decreased by 8.8 folds compared with that of the ER negative MDA-MB-231 cells. Dual-luciferase assay showed that the activity of IL-8 promoter was significantly down-regulated by ERalpha (r = 0.856, P < 0.05). CONCLUSIONS: IL-8 is the key factor involved in angiogenesis of human breast cancer cell. The IL-8 level in human breast cancer cells is negatively correlated with ER status. Exogenous ER may down-regulate the expression of IL-8 in breast cancer cell.

Animals↗
PubMed2005

The adipocyte as an important target cell for Trypanosoma cruzi infection.

Adipose tissue plays an active role in normal metabolic homeostasis as well as in the development of human disease. Beyond its obvious role as a depot for triglycerides, adipose tissue controls energy expenditure through secretion of several factors. Little attention has been given to the role of adipocytes in the pathogenesis of Chagas disease and the associated metabolic alterations. Our previous studies have indicated that hyperglycemia significantly increases parasitemia and mortality in mice infected with Trypanosoma cruzi. We determined the consequences of adipocyte infection in vitro and in vivo. Cultured 3T3-L1 adipocytes can be infected with high efficiency. Electron micrographs of infected cells revealed a large number of intracellular parasites that cluster around lipid droplets. Furthermore, infected adipocytes exhibited changes in expression levels of a number of different adipocyte-specific or adipocyte-enriched proteins. The adipocyte is therefore an important target cell during acute Chagas disease. Infection of adipocytes by T. cruzi profoundly influences the pattern of adipokines. During chronic infection, adipocytes may represent an important long-term reservoir for parasites from which relapse of infection can occur. We have demonstrated that acute infection has a unique metabolic profile with a high degree of local inflammation in adipose tissue, hypoadiponectinemia, hypoglycemia, and hypoinsulinemia but with relatively normal glucose disposal during an oral glucose tolerance test.

3T3-L1 Cells↗
PubMed2005

Adipocyte-derived collagen VI affects early mammary tumor progression in vivo, demonstrating a critical interaction in the tumor/stroma microenvironment.

The interactions of transformed cells with the surrounding stromal cells are of importance for tumor progression and metastasis. The relevance of adipocyte-derived factors to breast cancer cell survival and growth is well established. However, it remains unknown which specific adipocyte-derived factors are most critical in this process. Collagen VI is abundantly expressed in adipocytes. Collagen(-/-) mice in the background of the mouse mammary tumor virus/polyoma virus middle T oncogene (MMTV-PyMT) mammary cancer model demonstrate dramatically reduced rates of early hyperplasia and primary tumor growth. Collagen VI promotes its growth-stimulatory and pro-survival effects in part by signaling through the NG2/chondroitin sulfate proteoglycan receptor expressed on the surface of malignant ductal epithelial cells to sequentially activate Akt and beta-catenin and stabilize cyclin D1. Levels of the carboxyterminal domain of collagen VIalpha3, a proteolytic product of the full-length molecule, are dramatically upregulated in murine and human breast cancer lesions. The same fragment exerts potent growth-stimulatory effects on MCF-7 cells in vitro. Therefore, adipocytes play a vital role in defining the ECM environment for normal and tumor-derived ductal epithelial cells and contribute significantly to tumor growth at early stages through secretion and processing of collagen VI.

Adipocytes↗
PubMed2005

Synthetic peptides derived from SARS coronavirus S protein with diagnostic and therapeutic potential.

The spike (S) protein of severe acute respiratory syndrome coronavirus (SARS-CoV) is an important viral structural protein. Based on bioinformatics analysis, 10 antigenic peptides derived from the S protein sequence were selected and synthesized. The antigenicity and immunoreactivity of all the peptides were tested in vivo and in vitro. Four peptides (P6, P8, P9 and P10) which contain B cell epitopes of the S protein were identified, and P8 peptide was confirmed in vivo to have a potential in serological diagnosis. By using a syncytia formation model, we tested the neutralization ability of all 10 peptides and their corresponding antibodies. It is interesting to find that P8 and P9 peptides inhibited syncytia formation, suggesting that the P8 and P9 spanning regions may provide a good target for anti-SARS-CoV drug design. Our data suggest that we have identified peptides derived from the S protein of SARS-CoV, which are useful for SARS treatment and diagnosis.

Amino Acid Sequence↗
PubMed2004

A draft sequence for the genome of the domesticated silkworm (Bombyx mori).

We report a draft sequence for the genome of the domesticated silkworm (Bombyx mori), covering 90.9% of all known silkworm genes. Our estimated gene count is 18,510, which exceeds the 13,379 genes reported for Drosophila melanogaster. Comparative analyses to fruitfly, mosquito, spider, and butterfly reveal both similarities and differences in gene content.

Algorithms↗
PubMed2004

Postsynaptic density protein-95 regulates NMDA channel gating and surface expression.

NMDA receptors (NMDARs) colocalize with postsynaptic density protein-95 (PSD-95), a multivalent synaptic scaffolding protein and core component of the postsynaptic density, at excitatory synapses. Although much is known about the identity and properties of scaffolding proteins, little is known about their actions on NMDAR function. Here we show that association of PSD-95 with NMDARs modulates channel gating and surface expression. PSD-95 increases the number of functional channels at the cell surface and channel opening rate of NMDARs, with little or no change in conductance, reversal potential, or mean open time. We show further that PSD-95 increases NMDAR surface expression by increasing the rate of channel insertion and decreasing the rate of channel internalization. The PDZ (PSD-95, discs large, zona occludens-1) binding motif at the distal end of the NR2 C-terminal tail is critical to the actions of PSD-95 on NMDAR function and surface expression. Given that activity bi-directionally modifies synaptic levels of PSD-95, our findings suggest a novel mechanism for activity-dependent regulation of NMDARs at central synapses.

Amino Acid Motifs↗
PubMed2004

The hyperglycemia-induced inflammatory response in adipocytes: the role of reactive oxygen species.

Hyperglycemia is a major independent risk factor for diabetic macrovascular disease. The consequences of exposure of endothelial cells to hyperglycemia are well established. However, little is known about how adipocytes respond to both acute as well as chronic exposure to physiological levels of hyperglycemia. Here, we analyze adipocytes exposed to hyperglycemia both in vitro as well as in vivo. Comparing cells differentiated at 4 mm to cells differentiated at 25 mm glucose (the standard differentiation protocol) reveals severe insulin resistance in cells exposed to 25 mm glucose. A global assessment of transcriptional changes shows an up-regulation of a number of mitochondrial proteins. Exposure to hyperglycemia is associated with a significant induction of reactive oxygen species (ROS), both in vitro as well as in vivo in adipocytes isolated from streptozotocin-treated hyperglycemic mice. Furthermore, hyperglycemia for a few hours in a clamped setting will trigger the induction of a pro-inflammatory response in adipose tissue from rats that can effectively be reduced by co-infusion of N-acetylcysteine (NAC). ROS levels in 3T3-L1 adipocytes can be reduced significantly with pharmacological agents that lower the mitochondrial membrane potential, or by overexpression of uncoupling protein 1 or superoxide dismutase. In parallel with ROS, interleukin-6 secretion from adipocytes is significantly reduced. On the other hand, treatments that lead to a hyperpolarization of the mitochondrial membrane, such as overexpression of the mitochondrial dicarboxylate carrier result in increased ROS formation and decreased insulin sensitivity, even under normoglycemic conditions. Combined, these results highlight the importance ROS production in adipocytes and the associated insulin resistance and inflammatory response.

3T3-L1 Cells↗
PubMed2004

Characterization of the 3a protein of SARS-associated coronavirus in infected vero E6 cells and SARS patients.

Proteomics was used to identify a protein encoded by ORF 3a in a SARS-associated coronavirus (SARS-CoV). Immuno-blotting revealed that interchain disulfide bonds might be formed between this protein and the spike protein. ELISA indicated that sera from SARS patients have significant positive reactions with synthesized peptides derived from the 3a protein. These results are concordant with that of a spike protein-derived peptide. A tendency exists for co-mutation between the 3a protein and the spike protein of SARS-CoV isolates, suggesting that the function of the 3a protein correlates with the spike protein. Taken together, the 3a protein might be tightly correlated to the spike protein in the SARS-CoV functions. The 3a protein may serve as a new clinical marker or drug target for SARS treatment.

Animals↗
PubMed2004

Adipocyte differentiation induces dynamic changes in NF-kappaB expression and activity.

The adipocyte exerts an important role in energy homeostasis, both as depot for energy-rich triglycerides and as a source for metabolic hormones. Adipocytes also contribute to inflammation and the innate immune response. Although it can be physiologically beneficial to combine these two functions in a single cell type under some circumstances, the proinflammatory signals emanating from adipocytes in the obese state can have local and systemic effects that promote atherosclerosis and insulin resistance. The transcriptional machinery in the adipocyte that mediates these pro-inflammatory responses has remained poorly characterized to date. In particular, no information is currently available on the NF-kappaB family of transcription factors. Here, we show that adipogenesis is associated with changes in amount and subunit composition of the NF-kappaB complexes. NF-kappaB subunits p65 (RelA), p68 (RelB), and IkappaB are upregulated during fat cell differentiation. Correspondingly, basal NF-kappaB nuclear gel shift and luciferase reporter assays are induced in parallel during differentiation. Surprisingly, endotoxin sensitivity of the classical NF-kappaB pathway is substantially delayed and attenuated despite increased overall inflammatory response in the mature adipocyte, as judged by induction of IL-6 and TNF-alpha. As a reflection of the constitutively elevated NF-kappaB activity in the mature adipocyte, adipocytes (but not preadipocytes) exert a strong inflammatory stimulus on macrophages in vitro, suggesting a cross talk between adipocytes and interstitial macrophages in adipose tissue in vivo. These effects are mediated by a secretory product of adipocytes that is unlikely to be IL-6 or TNF-alpha.

3T3 Cells↗
PubMed2004

Identification of interleukin-8 as estrogen receptor-regulated factor involved in breast cancer invasion and angiogenesis by protein arrays.

With the goal of identifying key factors involved in human breast cancer progression, we applied human cytokine antibody arrays we have developed to screen cytokine expression levels in human breast cancer cell lines and identified interleukin (IL)-8 as a key factor involved in breast cancer invasion and angiogenesis. Elevated expression of IL-8 in breast cancer cells was associated with breast cancer invasiveness and angiogenesis. Neutralization of antibody against IL-8 specifically blocked IL-8-mediated tumor cell invasion and angiogenesis. Furthermore, IL-8 levels in human breast cancer cells were closely related to estrogen receptor (ER) status. ER positive cells expressed low levels of IL-8 whereas ER negative cells expressed high levels of IL-8. Expression of exogenous ERalpha substantially inhibited IL-8 expression. Our findings raise intriguing questions regarding the role of IL-8 in the development and progression of human breast cancer in association with ER status.

Breast Neoplasms↗
PubMed2004

Cyclooxygenase-2-dependent expression of angiogenic CXC chemokines ENA-78/CXC Ligand (CXCL) 5 and interleukin-8/CXCL8 in human non-small cell lung cancer.

Elevated tumor cyclooxygenase (COX)-2 activity plays a multifaceted role in non-small cell lung cancer (NSCLC). To elucidate the role of COX-2 in the in vitro and in vivo expression of two known NSCLC angiogenic peptides, CXC ligand (CXCL) 8 and CXCL5, we studied two COX-2 gene-modified NSCLC cell lines, A549 and H157. COX-2 overexpression enhanced the in vitro expression of both CXCL8 and CXCL5. In contrast, specific COX-2 inhibition decreased the production of both peptides as well as nuclear translocation of nuclear factor kappaB. In a severe combined immunodeficient mouse model of human NSCLC, the enhanced tumor growth of COX-2-overexpressing tumors was inhibited by neutralizing anti-CXCL5 and anti-CXCL8 antisera. We conclude that COX-2 contributes to the progression of NSCLC tumorigenesis by enhancing the expression of angiogenic chemokines CXCL8 and CXCL5.

Active Transport, Cell Nucleus↗
PubMed2004

Enhanced protein profiling arrays with ELISA-based amplification for high-throughput molecular changes of tumor patients' plasma.

PURPOSE: The purpose of this study is to develop a high-throughput approach to detect protein expression from hundreds and thousands of samples and to apply this technology to profile circulating angiogenic factor protein levels in patients with gynecological tumors. EXPERIMENTAL DESIGN: Analytes containing a mixture of protein are immobilized onto antibody-coated surface of support in array format. The presence of protein in analytes is detected with biotin-labeled antibody coupled with an enhanced chemiluminescence or fluorescence detection system. The exact amount of protein can be quantitatively measured. The expression levels of five angiogenic factors (angiogenin, interleukin 8, vascular endothelial growth factor, platelet-derived growth factor, and epidermal growth factor) from 157 samples were quantitatively measured using this novel protein array technology and were statistically analyzed. The expression patterns of angiogenic factors were analyzed using two-way hierarchical cluster analysis approach. RESULTS: A novel protein array technology, which can simultaneously and quantitatively measure few protein levels from hundreds and thousands of samples was developed. Only minute amounts of sample are required for the assay. This approach also features high sensitivity and specificity. Using this novel protein array approach, we analyzed the plasma expression levels of five angiogenic factors in 137 patients diagnosed with a tumor and 20 controls. Statistical analysis reveals different expression levels of angiogenic factors between patients and controls. Cluster analysis suggests a possible classification of normal subjects from patients. CONCLUSIONS: Enhanced protein profiling arrays provide a high-throughput and sensitive system to detect one or few protein from hundreds and thousands of samples. Such an approach should have broad application in biomedical discovery.

Adolescent↗
PubMed2004

Mining single nucleotide polymorphisms from EST data of silkworm, Bombyx mori, inbred strain Dazao.

We made use of 81,635 expressed sequence tags (ESTs) derived from 12 different cDNA libraries of the silkworm, Bombyx mori, inbred strain Dazao (P50), to identify high-quality candidate single nucleotide polymorphisms (SNPs). By PHRAP assembling, 12,980 contigs containing 11,537 contigs assembled by more than one read were obtained, and 101 candidate SNPs and 27 single base insertions/deletions were identified from 117 contigs assembled from 1576 high-quality reads base-called with PHRED and screened on the basis of the neighborhood quality standard (NQS). Simultaneously, we also predicted 40 SNPs in coding regions (cSNPs), of which 26 were predicted to lead to amino acid non-synonymous variations and 14 synonymous substitutions. Also, the 1.66:1 ratio of transition/transversion is different from that of other insects. As the first SNP analysis of a Lepidoptera, B. mori, the single nucleotide polymorphic density is estimated to be 1.3 x 10(-3) by sequence diversity. This analysis shows that expressed sequences from multiple libraries may provide an abundant source of comparative reads to mine for cSNPs from the silkworm genome.

Amino Acid Substitution↗
PubMed2004

The spike protein of severe acute respiratory syndrome (SARS) is cleaved in virus infected Vero-E6 cells.

Spike protein is one of the major structural proteins of severe acute respiratory syndrome-coronavirus. It is essential for the interaction of the virons with host cell receptors and subsequent fusion of the viral envelop with host cell membrane to allow infection. Some spike proteins of coronavirus, such as MHV, HCoV-OC43, AIBV and BcoV, are proteolytically cleaved into two subunits, S1 and S2. In contrast, TGV, FIPV and HCoV-229E are not. Many studies have shown that the cleavage of spike protein seriously affects its function. In order to investigate the maturation and proteolytic processing of the S protein of SARS CoV, we generated S1 and S2 subunit specific antibodies (Abs) as well as N, E and 3CL protein-specific Abs. Our results showed that the antibodies could efficiently and specifically bind to their corresponding proteins from E.coli expressed or lysate of SARS-CoV infected Vero-E6 cells by Western blot analysis. Furthermore, the anti-S1 and S2 Abs were proved to be capable of binding to SARS CoV under electron microscope observation. When S2 Ab was used to perform immune precipitation with lysate of SARS-CoV infected cells, a cleaved S2 fragment was detected with S2-specific mAb by Western blot analysis. The data demonstrated that the cleavage of S protein was observed in the lysate, indicating that proteolytic processing of S protein is present in host cells.

Animals↗
PubMed2004

Expression of intracellular domain of epidermal growth factor receptor and generation of its monoclonal antibody.

To prepare monoclonal antibody specific to epidermal growth factor receptor (EGFR) intracellular domain, its gene was amplified from total RNA of A431 cell by RT-PCR. Then the gene was cloned into prokaryotic vector pET30a(+). The recombinant plasmid was transformed into E. coli BL21 (DE3) strain for protein expression. Recombinant protein was induced with IPTG and purified using Ni2+-NTA agarose. Then the anti-EGFR monoclonal antibody (mAb) was prepared with classical hybridoma technique. Positive clones were selected using indirect enzyme-linked immunoabsorbent assay (ELISA). Totally 4 hybridoma clones were obtained and these mAbs were IgG1 (3 clones) and IgG2a (1 clone), respectively. Their light chains were all kappa chains. Western blotting analysis and confocal immunofluorescence assays demonstrated that mAbs could specifically recognize EGFR expressing on A431 carcinoma cell line. The mAbs will be useful in the study of EGFR-mediated signal transduction.

Animals↗