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Sue-Hwa Lin

Publications and source records attributed to Sue-Hwa Lin.

At least 19 recordsLinked to original sources
PubMed2005

SRC-3 is required for prostate cancer cell proliferation and survival.

Prostate cancer is the most common cancer in men in America. Currently, steroid receptor coactivators have been proposed to mediate the development and progression of prostate cancer, at times in a steroid-independent manner. Steroid receptor coactivator-3 (SRC-3, p/CIP, AIB1, ACTR, RAC3, and TRAM-1) is a member of the p160 family of coactivators for nuclear hormone receptors including the androgen receptor. SRC-3 is frequently amplified or overexpressed in a number of cancers. However, the role of SRC-3 in cancer cell proliferation and survival is still poorly understood. In this study, we show that SRC-3 is overexpressed in prostate cancer patients and its overexpression correlates with prostate cancer proliferation and is inversely correlated with apoptosis. Consistent with patient data, we have observed that reduction of SRC-3 expression by small interfering RNA decreases proliferation, delays the G1-S transition, and increases cell apoptosis of different prostate cancer cell lines. Furthermore, with decreased SRC-3 expression, proliferating cell nuclear antigen and Bcl-2 expression, as well as bromodeoxyuridine incorporation in prostate cancer cells are reduced. Finally, knockdown of SRC-3 with inducible short hairpin RNA expression in prostate cancer cells decreased tumor growth in nude mice. Taken together, these findings indicate that SRC-3 is an important regulator of prostate cancer proliferation and survival.

Acetyltransferases↗
PubMed2005

16-kDa prolactin down-regulates inducible nitric oxide synthase expression through inhibition of the signal transducer and activator of transcription 1/IFN regulatory factor-1 pathway.

Angiogenesis plays a key role in promoting tumorigenesis and metastasis. Several antiangiogenic factors have been shown to inhibit tumor growth in animal models. Understanding their mechanism of action would allow for better therapeutic application. 16-kDa prolactin (PRL), a NH2-terminal natural breakdown fragment of the intact 23-kDa PRL, exerts potent antiangiogenic and antitumor activities. The signaling mechanism involved in 16-kDa PRL action in endothelial cells remains unclear. One of the actions of 16-kDa PRL is to attenuate the production of nitric oxide (NO) through the inhibition of inducible NO synthase (iNOS) expression in endothelial cells. To delineate the signaling mechanism from 16-kDa PRL, we examined the effect of 16-kDa PRL on interleukin IL-1beta-inducible iNOS expression, which is regulated by two parallel pathways, one involving IFN regulatory factor 1 (IRF-1) and the other nuclear factor-kappaB (NF-kappaB). Our studies showed that 16-kDa PRL specifically blocked IRF-1 but not NF-kappaB signaling to the iNOS promoter. We found that IL-1beta regulated IRF-1 gene expression through stimulation of p38 mitogen-activated protein kinase (MAPK), which mediated signal transducer and activator of transcription 1 (Stat1) serine phosphorylation and Stat1 nuclear translocation to activate the IRF-1 promoter. 16-kDa PRL effectively inhibited IL-1beta-inducible p38 MAPK phosphorylation, resulting in blocking Stat1 serine phosphorylation, its subsequent nuclear translocation and activation of the Stat1 target gene IRF-1. Thus, 16-kDa PRL inhibits the p38 MAPK/Stat1/IRF-1 pathway to attenuate iNOS/NO production in endothelial cells.

Animals↗
PubMed2005

Insulin acutely decreases hepatic fatty acid synthase activity.

Insulin is viewed as a positive regulator of fatty acid synthesis by increasing fatty acid synthase (FAS) mRNA transcription. We uncover a new mechanism by which insulin acutely reduces hepatic FAS activity by inducing phosphorylation of the carcinoembryonic antigen-related cell adhesion molecule 1 (CEACAM1) and its interaction with FAS. Ceacam1 null mice (Cc1(-/-)) show loss of insulin's ability to acutely decrease hepatic FAS activity. Moreover, adenoviral delivery of wild-type, but not the phosphorylation-defective Ceacam1 mutant, restores the acute effect of insulin on FAS activity in Cc1(-/-) primary hepatocytes. Failure of insulin to acutely reduce hepatic FAS activity in hyperinsulinemic mice, including L-SACC1 transgenics with liver inactivation of CEACAM1, and Ob/Ob obese mice, suggests that the acute effect of insulin on FAS activity depends on the prior insulinemic state. We propose that this mechanism acts to reduce hepatic lipogenesis incurred by insulin pulses during refeeding.

Animals↗
PubMed2005

Expression of recombinant MDA-BF-1 with a kinase recognition site and a 7-histidine tag for receptor binding and purification.

Prostate cancer metastasizes predominantly to bone, where it induces osteoblastic lesions. Paracrine factors secreted by the metastatic cancer cells are thought to mediate these events. We previously isolated a novel bone metastasis-related factor (MDA-BF-1) from bone marrow aspirate samples from patients with prostate cancer and bone metastasis, and found that this factor stimulated osteoblast differentiation, possibly by interacting with a receptor on the osteoblasts. Identifying this putative MDA-BF-1 receptor biochemically requires the expression of MDA-BF-1 for receptor binding assays and for the preparation of a ligand-affinity column. We tagged MDA-BF-1 with a peptide containing a protein kinase A phosphorylation site plus a 7-histidine sequence to facilitate the labeling of MDA-BF-1 for receptor binding assay and the binding of MDA-BF-1 to an immobilized metal affinity column. The recombinant MDA-BF-1 protein (MDA-BF1-kinase-his) was expressed in Sf9 cells using a baculovirus expression system. About 0.8 mg of purified MDA-BF1-kinase-his protein was obtained from 4 x 10(8) Sf9 cells. MDA-BF1-kinase-his can be phosphorylated by PKA with a specific activity around 10(5)cpm/mug protein. Receptor binding assays using this (32)P-labeled MDA-BF-1 showed that MDA-BF-1 bound to membranes prepared from Saos-2, an osteosarcoma cell line, and C2C12, a mouse pluripotent mesenchymal precursor cell line that can be induced to become osteoblast by BMP-2. In contrast, MDA-BF-1 did not bind to membranes from PC-3 human prostate cancer cells or HEK293 human embryonic kidney cells. These observations suggest that the MDA-BF-1 receptor is expressed in cells of osteoblastic lineage. In addition to its use as a ligand for receptor binding assays, a ligand affinity column can be prepared by binding MDA-BF1-kinase-his to an IMAC for the purification of MDA-BF-1 receptor.

Animals↗
PubMed2005

Osteoblasts in prostate cancer metastasis to bone.

Metastasis to bone is common in lung, kidney, breast and prostate cancers. However, prostate cancer is unique in that bone is often the only clinically detectable site of metastasis, and the resulting tumours tend to be osteoblastic (bone forming) rather than osteolytic (bone lysing). The interaction between host cells and metastatic cancer cells is an important component of organ-specific cancer progression. How can this knowledge lead to the development of more effective therapies?

Bone Neoplasms↗
PubMed2004

Identification of a novel protein, LYRIC, localized to tight junctions of polarized epithelial cells.

Tight junctions (TJ) are multiprotein complexes that function to regulate paracellular transport of molecules through epithelial and endothelial cell layers. Many new tight junction-associated proteins have been identified in the past few years, and their functional roles and interactions have just begun to be elucidated. In this paper, we describe a novel protein LYsine-RIch CEACAM1 co-isolated (LYRIC) that is widely expressed and highly conserved between species. LYRIC has no conserved domains that would indicate function and does not appear to be a member of a larger protein family. Data from analysis of rat and human tissue sections and cell lines show that LYRIC colocalizes with tight junction proteins ZO-1 and occludin in polarized epithelial cells, suggesting that LYRIC is part of the tight junction complex. LYRIC dissociates from ZO-1 when junctional complexes are disrupted, and as tight junctions reform, ZO-1 relocalizes before LYRIC. These results suggest that LYRIC is most likely not a structural component required for TJ formation, but rather is recruited during the maturation of the tight junction complex.

Animals↗
PubMed2004

16K-prolactin inhibits activation of endothelial nitric oxide synthase, intracellular calcium mobilization, and endothelium-dependent vasorelaxation.

Activation of endothelial nitric oxide synthase (eNOS) and subsequent nitric oxide production (NO) are events that mediate the effect of important angiogenic, vasopermeability, and vasorelaxation factors, including vascular endothelial growth factor (VEGF), bradykinin (BK), and acetylcholine (ACh). The N-terminal 16-kDa fragment of prolactin (16K-PRL) acts on endothelial cells to inhibit angiogenesis both in vivo and in vitro. Here, we show that 16K-PRL inhibits VEGF-induced eNOS activation in endothelial cells. Inhibition of eNOS activation may mediate the antiangiogenic properties of 16K-PRL, because the NO donor (Z)-1-[2-(2-aminoethyl)- N-(2-ammonio-ethyl)amino]diazen-1-ium-1,2-diolate (DETANONOate) prevented 16K-PRL from blocking the VEGF-induced proliferation of endothelial cells. In addition, 16K-PRL inhibited eNOS activation by BK and blocked the BK-evoked transient increase in intracellular Ca(2+) in endothelial cells. This finding suggests that 16K-PRL interferes with the mobilization of intracellular Ca(2+), thereby inhibiting the Ca(2+)-dependent activation of eNOS. Blockage of eNOS activation can lead to inhibition of vasodilation. Consistently, 16K-PRL inhibited BK-induced relaxation of coronary vessels in isolated perfused guinea pig hearts. Moreover, 16K-PRL inhibited eNOS activation induced by ACh, and this action resulted in the inhibition of both ACh-evoked relaxation of coronary vessels in isolated perfused rat hearts and ACh-induced, endothelium-dependent relaxation of rat aortic segments. In conclusion, 16K-PRL can block the Ca(2+)-mediated activation of eNOS by three different vasoactive substances, and this action results in the inhibition of both angiogenesis and vasorelaxation.

Animals↗
PubMed2004

Two variable regions in carcinoembryonic antigen-related cell adhesion molecule1 N-terminal domains located in or next to monoclonal antibody and adhesion epitopes show evidence of recombination in rat but not in human.

In this paper, we have characterized the structure, evolutionary origin, and function of rat and human carcinoembryonic antigen-related cell adhesion molecule1 (CEACAM1) multifunctional Ig-like cell adhesion proteins that are expressed by many epithelial tissues. Restriction enzyme digestion reverse transcriptase-PCR analysis identified three cDNAs encoding novel CEACAM1 N-domains. Comparative sequence analysis showed that human and rat CEACAM1 N-domains segregated into two groups differing in similarity to rat CEACAM1(a)-4L and human CEACAM1. Sequence variability analysis indicated that both human and rat N-domains possessed two variable regions, and one contained a major adhesive epitope. Recombination analysis showed that the group of rat but not human N-domains with high sequence similarity was derived at least in part by recombination. Binding assays revealed that three monoclonal antibodies with strong reactivity for the CEACAM1(a)-4L N-domain showed no reactivity with CEACAM1(b)-4S, an allele with a different N-domain sequence. CEACAM1(b)-4S displayed adhesive activity efficiently blocked by a synthetic peptide corresponding to the adhesive epitope in CEACAM1(a)-4L. Blocking analysis also showed that the adhesive epitope for rat CEACAM1 was located downstream from the equivalent human and mouse epitopes. Glycosylation analysis demonstrated O-linked sugars on rat CEACAM1(b)-4S from COS-1 cells. However, this was not the alteration responsible for the lack of monoclonal antibody reactivity. When considered together with previous studies, our findings suggest an inverse relationship between functionality and amino acid sequence similarity to CEACAM1. Like IgG, the N-domain of CEACAM1 appears to tolerate 10-15% sequence diversification without loss of function but begins to show either altered specificity or diminished functionality at higher levels.

Amino Acid Sequence↗
PubMed2004

Identification of Sp2 as a transcriptional repressor of carcinoembryonic antigen-related cell adhesion molecule 1 in tumorigenesis.

Down-regulation of carcinoembryonic antigen-related cell adhesion molecule 1 (CEACAM1) tumor suppressor gene expression is common in several malignancies including prostate, colon, and breast cancer. The mechanism that mediates this down-regulation is not known. Here, we report that down-regulation of CEACAM1 expression in prostate cancer cells occurs primarily at the transcriptional level and is mediated by Sp2, a member of the Sp family of transcription factors. Sp2 binds to the CEACAM1 promoter in vitro and in vivo, and transient overexpression of Sp2 down-regulates endogenous CEACAM1 expression in normal prostate epithelial cells. Sp2 appears to repress CEACAM1 gene expression by recruiting histone deacetylase activity to the CEACAM1 promoter. In human prostate cancer specimens, Sp2 expression is high in prostate cancer cells but low in normal prostate epithelial cells and is inversely correlated with CEACAM1 expression. Our studies show that transcriptional repression by Sp2 represents one mechanism by which CEACAM1 tumor suppressor gene is down-regulated in prostate cancer.

Acetyltransferases↗
PubMed2004

Inhibition of prostate tumor growth by overexpression of NudC, a microtubule motor-associated protein.

Microtubules play a central role in coordinating various cellular functions that are orchestrated by their interaction with molecular motors. Anticancer drugs that target microtubule dynamics have been shown to be effective in cancer treatment. However, the effect of microtubule motor-associated molecules on cancer cell proliferation is not clear. Here, we investigated the role of NudC, a nuclear movement protein associated with the microtubule motor dynein, on prostate tumorigenesis. Recombinant adenovirus expressing NudC (Ad-NudC) was used to examine the effects of NudC on the tumorigenicity of prostate cancer cells. Expression of NudC in LNCaP cells inhibited their anchorage-independent growth in a soft agar colony assay. Expression of NudC in DU145 or PC-3 cells inhibited tumor growth in a subcutaneous xenograft model. At the cellular level, expression of NudC in DU145 and PC-3 cells inhibited cell proliferation at 48 h after Ad-NudC infection. FACS analysis of cell cycle distribution showed that 50-60% of Ad-NudC-infected PC-3 cells have a G2/M-phase DNA content compared to about 16-19% in Ad-Luciferase (Ad-Luc)-infected control cells, suggesting that NudC overexpression resulted in aberrant cell cycle progression. Immunofluorescence microscopy revealed a significant increase in cells with a single enlarged nucleus and cells exhibiting multiple nuclei, along with a concomitant increase in cell size in Ad-NudC-infected cells. These results suggest that NudC overexpression led to a block in cell division of prostate cancer cells, and that Ad-NudC may provide a new anticancer drug approach targeting the function of a microtubule motor-associated protein.

Adenoviridae↗
PubMed2004

Gene selection for oligonucleotide array: an approach using PM probe level data.

MOTIVATION: Analysis of oligonucleotide array data, especially to select genes of interest, is a highly challenging task because of the large volume of information and various experimental factors. Moreover, interaction effect (i.e. expression changes depend on probe effects) complicates the analysis because current methods often use an additive model to analyze data. We propose an approach to address these issues with the aim of producing a more reliable selection of differentially expressed genes. The approach uses the rank for normalization, employs the percentile-range to measure expression variation, and applies various filters to monitor expression changes. RESULTS: We compare our approach with MAS and Dchip models. A data set from an angiogenesis study is used for illustration. Results show that our approach performs better than other methods either in identification of the positive control gene or in PCR confirmatory tests. In addition, the invariant set of genes in our approach provides an efficient way for normalization.

Algorithms↗
PubMed2004

Up-regulation of MDA-BF-1, a secreted isoform of ErbB3, in metastatic prostate cancer cells and activated osteoblasts in bone marrow.

Prostate cancer (PCa) has a propensity to metastasize to bone. The identification of molecules that mediate the biological interactions between PCa cells and the bone environment is crucial to the understanding of PCa bone metastasis. The present study has used protein purification to identify bone metastasis-related factors present in bone marrow aspirates from PCa patients with bone metastasis. MDA-BF-1 was the first bone metastasis factor to be identified and is a secreted isoform of the ErbB3 growth factor receptor. To determine which cell types in PCa bone metastases express MDA-BF-1, MDA-BF-1 expression was studied in both primary and metastatic PCa cells, using an antibody to the extracellular domain (Ab10) and another to the cytoplasmic domain (RTJ.2) of ErbB3 to distinguish MDA-BF-1 from p180-ErbB3. It was found that epithelial cells in primary PCa did not express MDA-BF-1. In contrast, epithelial cells in 41 of 45 PCa metastases (18 of 19 lymph node metastases and 23 of 26 bone metastases), and activated osteoblasts in bone metastases, expressed MDA-BF-1. In addition, newly formed bone matrices adjacent to activated osteoblasts were also immunopositive for MDA-BF-1, suggesting that activated osteoblasts secrete MDA-BF-1. These observations indicate that MDA-BF-1 is up-regulated in metastatic PCa and raise the interesting possibility that MDA-BF-1 may play a role in the metastasis and progression of PCa, particularly in bone.

Antibodies↗
PubMed2004

CEACAM1 modulates epidermal growth factor receptor--mediated cell proliferation.

Phosphorylation of the cell adhesion protein CEACAM1 increases insulin sensitivity and decreases insulin-dependent mitogenesis in vivo. Here we show that CEACAM1 is a substrate of the EGFR and that upon being phosphorylated, CEACAM1 reduces EGFR-mediated growth of transfected Cos-7 and MCF-7 cells in response to EGF. Using transgenic mice overexpressing a phosphorylation-defective CEACAM1 mutant in liver (L-SACC1), we show that the effect of CEACAM1 on EGF-dependent cell proliferation is mediated by its ability to bind to and sequester Shc, thus uncoupling EGFR signaling from the ras/MAPK pathway. In L-SACC1 mice, we also show that impaired CEACAM1 phosphorylation leads to ligand-independent increase of EGFR-mediated cell proliferation. This appears to be secondary to visceral obesity and the metabolic syndrome, with increased levels of output of free fatty acids and heparin-binding EGF-like growth factor from the adipose tissue of the mice. Thus, L-SACC1 mice provide a model for the mechanistic link between increased cell proliferation in states of impaired metabolism and visceral obesity.

Animals↗
PubMed2003

Role for NudC, a dynein-associated nuclear movement protein, in mitosis and cytokinesis.

NudC, a nuclear movement protein that associates with dynein, was originally cloned as a mitogen-inducible early growth response gene. NudC forms a biochemical complex with components of the dynein/dynactin complex and is suggested to play a role in translocation of nuclei in proliferating neuronal progenitors as well as in migrating neurons in culture. Here, we show that NudC plays multiple roles in mitosis and cytokinesis in cultured mammalian cells. Altering NudC levels by either small interfering RNA-mediated gene silencing or adenovirus-mediated overexpression resulted in multinucleated cells and cells with persistent intercellular connections and disorganized midzone and midbody matrix. These phenotypes suggest a failure in cytokinesis in NudC altered cells. Further, a key mitotic enzyme, polo-like kinase, is mislocalized from the centrosomes and the midbody in NudC altered cells. Gene silencing of nud-1, the Caenorhabditis elegans ortholog of NudC, led to a loss of midzone microtubules and the rapid regression of the cleavage furrow, which resulted in one-celled embryos containing two nuclei. The loss of midzone microtubule organization owing to silencing of the NudC/nud-1 gene in two systems, coupled with the loss of Plk1 from mitotic structures in mammalian cells, provide clues to the cytokinesis defect and the multinucleation phenotype. Our findings suggest that NudC functions in mitosis and cytokinesis, in part by regulating microtubule organization at the midzone and midbody.

Active Transport, Cell Nucleus↗
PubMed2003

Antitumor activity of the 16-kDa prolactin fragment in prostate cancer.

The 16-kDa prolactin (PRL), derived from the proteolytic cleavage of wild-type 23-kDa PRL, has been shown to have antiangiogenic activity. Such an antiangiogenic activity may have an effect on tumor growth in vivo. Here we examined the effect of 23-kDa and 16-kDa PRL on tumor growth, and the potential of using recombinant 16-kDa human PRL for prostate cancer therapy. The effects of 23-kDa PRL and 16-kDa PRL on the tumorigenicity of prostate cancer cells in vivo were studied. Using an adenovirus transfer vector to achieve high efficiency 23-kDa and 16-kDa PRL transfection in DU145 and PC-3 human prostate carcinoma cell lines, we demonstrated that expression of 16-kDa PRL in the prostate cancer cells markedly reduced their ability to form tumors in a xenograft animal model. These studies established that the 16-kDa PRL has antitumor activity in vivo, presumably as a result of its antiangiogenic effect. Interestingly, 23-kDa PRL showed a weak and transient suppression of prostate tumor growth. The weak antitumor activity of 23-kDa PRL may be because of the production of 16-kDa PRL from 23-kDa PRL by the tumor cells. Thus, the apparent effect of 23-kDa PRL on the growth of DU145 and PC-3 cells in vivo may result from the combined effects of 23-kDa PRL and 16-kDa PRL. These results suggest that the 16-kDa PRL has potential as a treatment agent in prostate cancer.

Adenoviridae↗
PubMed2003

Expression and purification of the angiogenesis inhibitor 16-kDa prolactin fragment from insect cells.

The 16-kDa fragment of prolactin (16-kDa PRL), derived from proteolytic cleavage of 23-kDa PRL, was shown to have antiangiogenic activity. Previous studies have shown that recombinant 16-kDa PRL produced from bacteria often contained endotoxins, which are cytotoxic to endothelial cells, and varied in its biological activity due to changes in its refolding from inclusion bodies. These problems limited the use of recombinant 16-kDa PRL. To improve the generation of recombinant 16-kDa PRL, we expressed 16-kDa PRL in Sf9 insect cells using a baculoviral expression system. The signal sequence of the human PRL gene and codons for seven histidines were added to the N- and C-termini, respectively, of the 16-kDa PRL cDNA construct. Recombinant 16-kDa PRL was detected in both the cell pellet and the medium. About 0.28 mg purified protein was isolated from the cell pellet of 4 x 10(7) infected cells using nickel affinity chromatography. Sixteen kilodalton PRL was posttranslationally modified with apparent molecular weights of 16 and 18 kDa on SDS-PAGE. The level of 18-kDa protein was significantly reduced after digestion with peptidyl-N-glycosidase, suggesting that the heterogeneity was due to glycosylation of 16-kDa PRL. N-terminal sequence analysis confirmed the fact that both proteins were human 16-kDa PRL and the signal sequences were cleaved at the same position as that of human PRL. Consistent with its role as an angiogenesis inhibitor, purified recombinant 16-kDa PRL inhibits the proliferation of endothelial cells with a potency similar to that previously reported for the protein generated in Escherichia coli. This 16-kDa PRL expressed in Sf9 cells is a useful reagent for functional studies and for the purification and identification of its receptor.

Angiogenesis Inhibitors↗