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Sample records for chemoattractant protein-1 expression

  1. Angiotensin II induces monocyte chemoattractant protein-1 gene expression in rat vascular smooth muscle cells.

    PubMed

    Chen, X L; Tummala, P E; Olbrych, M T; Alexander, R W; Medford, R M

    1998-11-01

    Monocyte infiltration into the vessel wall, a key initial step in the process of atherosclerosis, is mediated in part by monocyte chemoattractant protein-1 (MCP-1). Hypertension, particularly in the presence of an activated renin-angiotensin system, is a major risk factor for the development of atherosclerosis. To investigate a potential molecular basis for a link between hypertension and atherosclerosis, we studied the effects of angiotensin II (Ang II) on MCP-1 gene expression in rat aortic smooth muscle cells. Rat smooth muscle cells treated with Ang II exhibited a dose-dependent increase in MCP-1 mRNA accumulation that was prevented by the AT1 receptor antagonist losartan. Ang II also activated MCP-1 gene transcription. Inhibition of NADH/NADPH oxidase, which generates superoxide and H2O2, with diphenylene iodonium or apocynin decreased Ang II-induced MCP-1 mRNA accumulation. Induction of MCP-1 gene expression by Ang II was inhibited by catalase, suggesting a second messenger role for H2O2. The tyrosine kinase inhibitor genistein and the mitogen-activated protein kinase kinase inhibitor PD098059 inhibited Ang II-induced MCP-1 gene expression, consistent with a mitogen-activated protein kinase-dependent signaling mechanism. Ang II may thus promote atherogenesis by direct activation of MCP-1 gene expression in vascular smooth muscle cells.

  2. Renal expression of monocyte chemoattractant protein-1 in lupus autoimmune mice.

    PubMed

    Zoja, C; Liu, X H; Donadelli, R; Abbate, M; Testa, D; Corna, D; Taraboletti, G; Vecchi, A; Dong, Q G; Rollins, B J; Bertani, T; Remuzzi, G

    1997-05-01

    Mononuclear cell infiltration in glomeruli and renal interstitium is a prominent feature of some types of glomerulonephritis, including lupus nephritis. The mechanism(s) underlying monocyte influx into the kidney is not fully understood. Recently, monocyte chemoattractant protein-1 (MCP-1) has been identified as a chemotactic factor involved in the recruitment of monocytes/macrophages in the glomeruli of rats with mesangioproliferative as well as anti-glomerular basement membrane glomerulonephritis. In the study presented here, renal MCP-1 mRNA expression in New Zealand Black x New Zealand White (NZB/W) F1 mice, a model of genetically determined immune complex disease that mimics systemic lupus in humans, was investigated. Northern blot analysis revealed a single 0.7 kb MCP-1 transcript of very low intensity in kidneys from 2-month-old NZB/W mice that had not yet developed proteinuria nor renal damage. Message levels, which increased markedly with the progression of nephritis and in association with mononuclear cell infiltration, were 10- and 15- fold higher in 8-10-month-old mice than in 2-month-old mice. By in situ hybridization, increased expression of MCP-1 mRNA was demonstrated in glomeruli and, even more striking, in tubular epithelial cells. Western blot analysis demonstrated increased expression of MCP-1 protein in kidneys of 10-month-old NZB/W mice, consistent with MCP-1 mRNA data. When NZB/W mice were treated with cyclophosphamide up to 12 months of age, expression of MCP-1 in the renal tissue remained low, the influx of inflammatory cells did not appear, and glomerular and tubular structures remained well preserved. These data suggest that elevated MCP-1 might act as a signal for inflammatory cells to infiltrate the kidney in lupus nephritis.

  3. Enhanced expression and production of monocyte chemoattractant protein-1 in myocarditis

    PubMed Central

    Fuse, K; Kodama, M; Hanawa, H; Okura, Y; Ito, M; Shiono, T; Maruyama, S; Hirono, S; Kato, K; Watanabe, K; Aizawa, Y

    2001-01-01

    Monocyte chemoattractant protein-1 (MCP-1) is a member of the C-C chemokine family that has been shown to play a major role in the migration of monocytes and T cells to an inflammatory focus. To clarify the role of MCP-1 in the pathogenesis of myocarditis, we have examined the expression of MCP-1 in rat hearts with experimental autoimmune myocarditis (EAM), and have also measured serum levels of MCP-1 in patients with histology-proven acute myocarditis. Lewis rats were immunized with cardiac myosin and were killed 9, 12, 15, 18, 21, 24, 27, 30, 33, 36, 42 and 56 days after immunization. Large mononuclear cells in the myocardial interstitium were stained with an anti-MCP-1 antibody. mRNA of MCP-1 increased in the hearts of EAM rats from days 15–27 as shown by quantitative reverse transcription-polymerase chain reaction. Serum MCP-1 levels of the rats with EAM were significantly elevated from days 15–24. In the clinical study, serum levels of MCP-1 in 24 patients with acute myocarditis at the time of admission (165·2 ± 55·8 pg/ml) were significantly (P = 0·0301) elevated compared with those of 20 healthy volunteers (61·8 ± 10·7 pg/ml). Serum MCP-1 levels of 8 fatal cases (371·8 ± 145·2 pg/ml) were significantly (P = 0·0058) higher than those of 16 cases who survived (65·5 ± 12·8 pg/ml). In conclusions, MCP-1 may play an important role in the pathogenesis of human acute myocarditis as well as in the progression of rat EAM. PMID:11472393

  4. Suppression of lipin-1 expression increases monocyte chemoattractant protein-1 expression in 3T3-L1 adipocytes

    SciTech Connect

    Takahashi, Nobuhiko; Hiranaka, Natsumi; Suzuki, Takeshi; Yui, Tomoo; Akanuma, Masayasu; Oka, Kazuya; Kanazawa, Kaoru; Yoshida, Mika; Naito, Sumiyoshi; Fujiya, Mikihiro; Kohgo, Yutaka

    2011-11-11

    Highlights: Black-Right-Pointing-Pointer Lipin-1 affects lipid metabolism, adipocyte differentiation, and transcription. Black-Right-Pointing-Pointer Adipose lipin-1 expression is reduced in obesity. Black-Right-Pointing-Pointer Lipin-1 depletion using siRNA in 3T3-L1 adipocytes increased MCP-1 expression. Black-Right-Pointing-Pointer Lipin-1 is involved in adipose inflammation. -- Abstract: Lipin-1 plays a crucial role in the regulation of lipid metabolism and cell differentiation in adipocytes. Expression of adipose lipin-1 is reduced in obesity, and metabolic syndrome. However, the significance of this reduction remains unclear. This study investigated if and how reduced lipin-1 expression affected metabolism. We assessed mRNA expression levels of various genes related to adipocyte metabolism in lipin-1-depleted 3T3-L1 adipocytes by introducing its specific small interfering RNA. In lipin-1-depleted adipocytes, mRNA and protein expression levels of monocyte chemoattractant protein-1 (MCP-1) were significantly increased, although the other genes tested were not altered. The conditioned media from the cells promoted monocyte chemotaxis. The increase in MCP-1 expression was prevented by treatment with quinazoline or salicylate, inhibitors of nuclear factor-{kappa}B activation. Because MCP-1 is related to adipose inflammation and systemic insulin resistance, these results suggest that a reduction in adipose lipin-1 in obesity may exacerbate adipose inflammation and metabolism.

  5. Hypoxia reduces constitutive and TNF-{alpha}-induced expression of monocyte chemoattractant protein-1 in human proximal renal tubular cells

    SciTech Connect

    Li Xuan; Kimura, Hideki . E-mail: hkimura@fmsrsa.fukui-med.ac.jp; Hirota, Kiichi; Sugimoto, Hidehiro; Yoshida, Haruyoshi

    2005-10-07

    Chronic hypoxia has been reported to be associated with macrophage infiltration in progressive forms of kidney disease. Here, we investigated the regulatory effects of hypoxia on constitutive and TNF-{alpha}-stimulated expression of monocyte chemoattractant protein-1 (MCP-1) in cultured human proximal renal tubular cells (HPTECs). Hypoxia reduced constitutive MCP-1 expression at the mRNA and protein levels in a time-dependent fashion for up to 48 h. Hypoxia also inhibited MCP-1 up-regulation by TNF-{alpha}. Treatment with actinomycin D showed that hypoxic down-regulation of MCP-1 expression resulted mainly from a decrease in the transcription but not the mRNA stability. Immunoblot and immunofluorescence analyses revealed that treatment with hypoxia or an iron chelator, desferrioxamine, induced nuclear accumulation of hypoxia-inducible factor-1{alpha} (HIF-1{alpha}) in HPTECs. Desferrioxamine mimicked hypoxia in the reduction of MCP-1 expression. However, overexpression of a dominant negative form of HIF-1{alpha} did not abolish the hypoxia-induced reduction of MCP-1 expression in HPTECs. These results suggest that hypoxia is an important negative regulator of monocyte chemotaxis to the renal inflamed interstitium, by reducing MCP-1 expression partly via hypoxia-activated signals other than the HIF-1 pathway.

  6. Fibrinogen up-regulates the expression of monocyte chemoattractant protein 1 in human saphenous vein endothelial cells.

    PubMed Central

    Harley, S L; Powell, J T

    1999-01-01

    High concentrations of fibrinogen in plasma have been associated with an increased risk of saphenous vein graft pathology. We have investigated the ability of fibrinogen to up-regulate the expression of monocyte chemoattractant protein 1 (MCP-1) in cultured human saphenous vein endothelial cells (HSVEC) isolated from saphenous vein. Increasing concentrations of fibrinogen (0-4 microM) stimulated a 20-fold increase in MCP-1 secretion within 4 h. Incubation of HSVEC with 2 microM fibrinogen for 4 h also caused a 2-fold increase in the MCP-1-to-glyceraldehyde-3-phosphate dehydrogenase mRNA ratio. The fibrinogen-mediated MCP-1 secretion fell to basal levels after preincubation of HSVEC with the complex of fibrinogen fragments D and E but remained unchanged after preincubation of HSVEC with either fibrinogen fragment E, s-ICAM-1 or the pentapeptide GRGDV. In contrast, fibrinogen fragment D acted as a potent inhibitor of fibrinogen-mediated MCP-1 secretion. Labelled fibrinogen fragment D bound to HSVEC with a K(d) of 6.5 microM. These findings indicate that fibrinogen, at physiological concentrations, uses an epitope on the fibrinogen D domain to bind to a receptor on HSVEC to up-regulate MCP-1 expression and secretion. This receptor seems to be distinct from intercellular adhesion molecule 1 and the integrins previously recognized as fibrinogen receptors. PMID:10417339

  7. Effects of Tumor Necrosis Factor-α on Podocyte Expression of Monocyte Chemoattractant Protein-1 and in Diabetic Nephropathy

    PubMed Central

    Chung, Choon Hee; Fan, Jingyi; Lee, Eun Young; Kang, Jeong Suk; Lee, Seung Joo; Pyagay, Petr E.; Khoury, Charbel C.; Yeo, Tet-Kin; Khayat, Mark F.; Wang, Amy; Chen, Sheldon

    2015-01-01

    Background/Aims Tumor necrosis factor (TNF)-α is believed to play a role in diabetic kidney disease. This study explores the specific effects of TNF-α with regard to nephropathy-relevant parameters in the podocyte. Methods Cultured mouse podocytes were treated with recombinant TNF-α and assayed for production of monocyte chemoattractant protein-1 (MCP-1) by enzyme-linked immunosorbent assay (ELISA). TNF-α signaling of MCP-1 was elucidated by antibodies against TNF receptor (TNFR) 1 or TNFR2 or inhibitors of nuclear factor-kappaB (NF-κB), phosphatidylinositol 3-kinase (PI3K) or Akt. In vivo studies were done on male db/m and type 2 diabetic db/db mice. Levels of TNF-α and MCP-1 were measured by RT-qPCR and ELISA in the urine, kidney and plasma of the two cohorts and correlated with albuminuria. Results Podocytes treated with TNF-α showed a robust increase (∼900%) in the secretion of MCP-1, induced in a dose- and time-dependent manner. Signaling of MCP-1 expression occurred through TNFR2, which was inducible by TNF-α ligand, but did not depend on TNFR1. TNF-α then proceeded via the NF-κB and the PI3K/Akt systems, based on the effectiveness of the inhibitors of those pathways. For in vivo relevance to diabetic kidney disease, TNF-α and MCP-1 levels were found to be elevated in the urine of db/db mice but not in the plasma. Conclusion TNF-α potently stimulates podocytes to produce MCP-1, utilizing the TNFR2 receptor and the NF-κB and PI3K/Akt pathways. Both TNF-α and MCP-1 levels were increased in the urine of diabetic db/db mice, correlating with the severity of diabetic albuminuria. PMID:25852733

  8. Molecular cloning and functional expression of two monocyte chemoattractant protein 1 receptors reveals alternative splicing of the carboxyl-terminal tails.

    PubMed Central

    Charo, I F; Myers, S J; Herman, A; Franci, C; Connolly, A J; Coughlin, S R

    1994-01-01

    Monocyte chemoattractant protein 1 (MCP-1) is a member of the chemokine family of cytokines that mediate leukocyte chemotaxis. The potent and specific activation of monocytes by MCP-1 may mediate the monocytic infiltration of tissues in atherosclerosis and other inflammatory diseases. We have isolated cDNAs that encode two MCP-1-specific receptors with alternatively spliced carboxyl tails. Expression of the receptors in Xenopus oocytes conferred robust mobilization of intracellular calcium in response to nanomolar concentrations of MCP-1 but not to related chemokines. The MCP-1 receptors are most closely related to the receptor for the chemokines macrophage inflammatory protein 1 alpha and RANTES (regulated on activation, normal T expressed and secreted). The identification of the MCP-1 receptor and cloning of two distinct isoforms provide powerful tools for understanding the specificity and signaling mechanisms of this important chemokine. Images PMID:8146186

  9. Coxsackievirus group B type 3 infection upregulates expression of monocyte chemoattractant protein 1 in cardiac myocytes, which leads to enhanced migration of mononuclear cells in viral myocarditis.

    PubMed

    Shen, Yan; Xu, Wei; Chu, Yi-Wei; Wang, Ying; Liu, Quan-Sheng; Xiong, Si-Dong

    2004-11-01

    Coxsackievirus group B type 3 (CVB3) is an important cause of viral myocarditis. The infiltration of mononuclear cells into the myocardial tissue is one of the key events in viral myocarditis. Immediately after CVB3 infects the heart, the expression of chemokine(s) by infected myocardial cells may be the first trigger for inflammatory infiltration and immune response. However, it is unknown whether CVB3 can induce the chemokine expression in cardiac myocytes. Monocyte chemoattractant protein 1 (MCP-1) is a potent chemokine that stimulates the migration of mononuclear cells. The objective of the present study was to investigate the effect of CVB3 infection on MCP-1 expression in murine cardiac myocytes and the role of MCP-1 in migration of mononuclear cells in viral myocarditis. Our results showed that the expression of MCP-1 was significantly increased in cardiac myocytes after wild-type CVB3 infection in a time- and dose-dependent manner, which resulted in enhanced migration of mononuclear cells in mice with viral myocarditis. The migration of mononuclear cells was partially abolished by antibodies specific for MCP-1 in vivo and in vitro. Administration of anti-MCP-1 antibody prevented infiltration of mononuclear cells bearing the MCP-1 receptor CCR2 in mice with viral myocarditis. Infection by UV-irradiated CVB3 induced rapid and transient expression of MCP-1 in cardiac myocytes. In conclusion, our results indicate that CVB3 infection stimulates the expression of MCP-1 in myocardial cells, which subsequently leads to migration of mononuclear cells in viral myocarditis. PMID:15507642

  10. Bone morphogenetic protein-7 inhibits constitutive and interleukin-1 beta-induced monocyte chemoattractant protein-1 expression in human mesangial cells: role for JNK/AP-1 pathway.

    PubMed

    Lee, Myung-Ja; Yang, Chul Woo; Jin, Dong Chan; Chang, Yoon Sik; Bang, Byung Kee; Kim, Yong-Soo

    2003-03-01

    Bone morphogenetic protein-7 (BMP-7), which belongs to the TGF-beta superfamily, has been shown to reduce macrophage infiltration and tissue injury in animal models of inflammatory renal disease. To explore the mechanism involved in the anti-inflammatory effect, we investigated the effect of BMP-7 on monocyte chemoattractant protein-1 (MCP-1) expression in cultured human mesangial cells. BMP- 7 significantly inhibited constitutive and IL-1 beta-induced MCP-1 protein production and MCP-1 mRNA expression by mesangial cells in a time- and concentration-dependent manner. BMP-7 also inhibited IL-1 beta-induced monocyte chemotactic activity released from the mesangial cells. We examined the role of transcription factors NF-kappa B and AP-1 in BMP-7 inhibition of IL-1 beta-induced MCP-1 expression. IL-1 beta increased NF-kappa B and AP-1 activity and both transcription factors mediated IL-1 beta-induced MCP-1 expression in mesangial cells. BMP-7 inhibited IL-1 beta-induced AP-1 activity in a concentration-dependent manner. In contrast, IL-1 beta-induced NF-kappa B activity and I kappa B alpha degradation were not affected by BMP-7. Furthermore, IL-1 beta-induced phosphorylation of c-Jun N-terminal kinase was inhibited by BMP-7. These data suggest that BMP-7 inhibits constitutive and IL-1 beta-induced MCP-1 expression in human mesangial cells partly by inhibiting c-Jun N-terminal kinase activity and subsequent AP-1 activity, and provide new insight into the therapeutic potential of BMP-7 in the inflammatory renal diseases.

  11. Monocyte Chemoattractant Protein 1 (MCP-1) in Obesity and Diabetes

    PubMed Central

    Panee, Jun

    2012-01-01

    Monocyte chemoattractant protein-1 (MCP-1) is the first discovered and most extensively studied CC chemokine, and the amount of studies on its role in the etiologies of obesity- and diabetes-related diseases have increased exponentially during the past 2 decades. This review attempted to provide a panoramic perspective of the history, regulatory mechanisms, functions, and therapeutic strategies of this chemokine. The highlights of this review include the roles of MCP-1 in the development of obesity, diabetes, cardiovascular diseases, insulitis, diabetic nephropathy, and diabetic retinopathy. Therapies that specifically or non-specifically inhibit MCP-1 overproduction have been summarized. PMID:22766373

  12. The detection and localization of monocyte chemoattractant protein-1 (MCP-1) in human ovarian cancer.

    PubMed Central

    Negus, R P; Stamp, G W; Relf, M G; Burke, F; Malik, S T; Bernasconi, S; Allavena, P; Sozzani, S; Mantovani, A; Balkwill, F R

    1995-01-01

    Chemokines may control the macrophage infiltrate found in many solid tumors. In human ovarian cancer, in situ hybridization detected mRNA for the macrophage chemokine monocyte chemoattractant protein-1 (MCP-1) in 16/17 serous carcinomas, 4/4 mucinous carcinomas, 2/2 endometrioid carcinomas, and 1/3 borderline tumors. In serous tumors, mRNA expression mainly localized to the epithelial areas, as did immunoreactive MCP-1 protein. In the other tumors, both stromal and epithelial expression were seen. All tumors contained variable numbers of cells positive for the macrophage marker CD68. MCP-1 mRNA was also detected in the stroma of 5/5 normal ovaries. RT-PCR demonstrated mRNA for MCP-1 in 7/7 serous carcinomas and 6/6 ovarian cancer cell lines. MCP-1 protein was detected by ELISA in ascites from patients with ovarian cancer (mean 4.28 ng/ml) and was produced primarily by the cancer cells. Human MCP-1 protein was also detected in culture supernatants from cell lines and in ascites from human ovarian tumor xenografts which induce a peritoneal monocytosis in nude mice. We conclude that the macrophage chemoattractant MCP-1 is produced by epithelial ovarian cancer and that the tumor cells themselves are probably a major source. MCP-1 may contribute to the accumulation of tumor-associated macrophages, which may subsequently influence tumor behavior. Images PMID:7738202

  13. Cilostazol prevents remnant lipoprotein particle-induced monocyte adhesion to endothelial cells by suppression of adhesion molecules and monocyte chemoattractant protein-1 expression via lectin-like receptor for oxidized low-density lipoprotein receptor activation.

    PubMed

    Park, So Youn; Lee, Jeong Hyun; Kim, Yong Ki; Kim, Chi Dae; Rhim, Byung Yong; Lee, Won Suk; Hong, Ki Whan

    2005-03-01

    This study shows cilostazol effect to prevent remnant lipoprotein particle (RLP)-induced monocyte adhesion to human umbilical vein endothelial cells (HUVECs). Upon incubation of HUVECs with RLP (50 microg/ml), adherent monocytes significantly increased by 3.3-fold with increased cell surface expression of vascular cell adhesion molecule-1 (VCAM-1), intercellular adhesion molecule-1, E-selectin, and monocyte chemoattractant protein-1 (MCP-1). Cilostazol ( approximately 1-100 microM) concentration dependently repressed these variables as did (E)3-[(4-t-butylphenyl)sulfonyl]-2-propenenitrile (BAY 11-7085) (10 microM), a specific nuclear factor-kappaB (NF-kappaB) inhibitor. Cilostazol effects were significantly antagonized by iberiotoxin (1 microM), a maxi-K channel blocker. RLP significantly increased expression of lectin-like receptor for oxidized low-density lipoprotein (LDL) (LOX-1) receptor protein. Upon transfection with antisense LOX-1 oligodeoxynucleotide (As-LOX-1), LOX-1 receptor expression was reduced, whereas HUVECs with sense LOX-1 oligodeoxynucleotide did express high LOX-1 receptor. RLP-stimulated superoxide and tumor necrosis factor-alpha levels were significantly lowered with decreased expression of VCAM-1 and MCP-1 by transfection with As-LOX-1 as did polyinosinic acid (10 microg/ml, a LOX-1 receptor inhibitor). RLP significantly degraded inhibitory kappaBalpha in the cytoplasm and activated nuclear factor-kappaB (NF-kappaB) p65 in the nucleus of HUVECs with increased luciferase activity of NF-kappaB, all of which were reversed by cilostazol (10 microM), BAY 11-7085, and polyinosinic acid. Together, cilostazol suppresses RLP-stimulated increased monocyte adhesion to HUVECs by suppression of LOX-1 receptor-coupled NF-kappaB-dependent nuclear transcription via mediation of the maxi-K channel opening.

  14. Arctigenin suppresses transforming growth factor-β1-induced expression of monocyte chemoattractant protein-1 and the subsequent epithelial-mesenchymal transition through reactive oxygen species-dependent ERK/NF-κB signaling pathway in renal tubular epithelial cells.

    PubMed

    Li, A; Wang, J; Zhu, D; Zhang, X; Pan, R; Wang, R

    2015-01-01

    Transforming growth factor-β1 (TGF-β1) induces expression of the proinflammatory and profibrotic cytokine monocyte chemoattractant protein-1 (MCP-1) in tubular epithelial cells (TECs) and thereby contributes to the tubular epithelial-mesenchymal transition (EMT), which in turn leads to the progression of tubulointerstitial inflammation into tubulointerstitial fibrosis. Exactly how TGF-β1 causes MCP-1 overexpression and subsequent EMT is not well understood. Using human tubular epithelial cultures, we found that TGF-β1 upregulated the expression of reduced nicotinamide adenine dinucleotide phosphate oxidases 2 and 4 and their regulatory subunits, inducing the production of reactive oxygen species. These reactive species activated a signaling pathway mediated by extracellular signal-regulated kinase (ERK1/2) and nuclear factor-κB (NF-κB), which upregulated expression of MCP-1. Incubating cultures with TGF-β1 was sufficient to induce hallmarks of EMT, such as downregulation of epithelial marker proteins (E-cadherin and zonula occludens-1), induction of mesenchymal marker proteins (α-smooth muscle actin, fibronectin, and vimentin), and elevated cell migration and invasion in an EMT-like manner. Overexpressing MCP-1 in cells exposed to TGF-β1 exacerbated these EMT-like changes. Pretreating cells with the antioxidant and anti-inflammatory compound arctigenin (ATG) protected them against these TGF-β1-induced EMT-like changes; the compound worked by inhibiting the ROS/ERK1/2/NF-κB pathway to decrease MCP-1 upregulation. These findings suggest ATG as a new therapeutic candidate to inhibit or even reverse tubular EMT-like changes during progression to tubulointerstitial fibrosis, and they provide the first clues to how ATG may work.

  15. Arctigenin suppresses transforming growth factor-β1-induced expression of monocyte chemoattractant protein-1 and the subsequent epithelial-mesenchymal transition through reactive oxygen species-dependent ERK/NF-κB signaling pathway in renal tubular epithelial cells.

    PubMed

    Li, A; Wang, J; Zhu, D; Zhang, X; Pan, R; Wang, R

    2015-01-01

    Transforming growth factor-β1 (TGF-β1) induces expression of the proinflammatory and profibrotic cytokine monocyte chemoattractant protein-1 (MCP-1) in tubular epithelial cells (TECs) and thereby contributes to the tubular epithelial-mesenchymal transition (EMT), which in turn leads to the progression of tubulointerstitial inflammation into tubulointerstitial fibrosis. Exactly how TGF-β1 causes MCP-1 overexpression and subsequent EMT is not well understood. Using human tubular epithelial cultures, we found that TGF-β1 upregulated the expression of reduced nicotinamide adenine dinucleotide phosphate oxidases 2 and 4 and their regulatory subunits, inducing the production of reactive oxygen species. These reactive species activated a signaling pathway mediated by extracellular signal-regulated kinase (ERK1/2) and nuclear factor-κB (NF-κB), which upregulated expression of MCP-1. Incubating cultures with TGF-β1 was sufficient to induce hallmarks of EMT, such as downregulation of epithelial marker proteins (E-cadherin and zonula occludens-1), induction of mesenchymal marker proteins (α-smooth muscle actin, fibronectin, and vimentin), and elevated cell migration and invasion in an EMT-like manner. Overexpressing MCP-1 in cells exposed to TGF-β1 exacerbated these EMT-like changes. Pretreating cells with the antioxidant and anti-inflammatory compound arctigenin (ATG) protected them against these TGF-β1-induced EMT-like changes; the compound worked by inhibiting the ROS/ERK1/2/NF-κB pathway to decrease MCP-1 upregulation. These findings suggest ATG as a new therapeutic candidate to inhibit or even reverse tubular EMT-like changes during progression to tubulointerstitial fibrosis, and they provide the first clues to how ATG may work. PMID:25968940

  16. Monocyte chemoattractant protein-1 serum levels in ovarian cancer patients

    PubMed Central

    Hefler, L; Tempfer, C; Heinze, G; Mayerhofer, K; Breitenecker, G; Leodolter, S; Reinthaller, A; Kainz, C

    1999-01-01

    The chemokine monocyte chemoattractant protein (MCP)-1 is an important mediator of monocyte infiltration in various solid tumours of epithelial origin. The aim of the present study was to evaluate the role of MCP-1 in the natural history of ovarian cancer and to determine its value as differentiation marker and prognostic marker regarding disease free and overall survival. This retrospective study comprises 86 patients with ovarian cancer, 48 with primary ovarian cancer and 38 with recurrent ovarian cancer, 67 patients with benign ovarian cysts and 42 healthy women. Median serum levels in patients with primary ovarian cancer, recurrent ovarian cancer, benign ovarian cysts and in healthy women were 535.6 (range 129.6–1200) pg ml–1, 427.3 (range 193.4–1101) pg ml–1, 371.2 (range 222–986.8) pg ml–1 and 318.7 (range 241.3–681.4) pg ml–1 respectively (Mann–Whitney U-test, P < 0.001). Univariate logistic regression models revealed a significant influence of MCP-1 serum levels on the odds of presenting with primary ovarian cancer versus benign cysts and versus healthy women respectively (univariate logistic regression, P < 0.001 and P < 0.001 respectively). In a multivariate logistic regression model considering MCP-1 and CA 125 serum levels simultaneously, both MCP-1 and CA 125 revealed statistical significance on the odds of presenting with primary ovarian cancer versus benign cysts (multivariate logistic regression, P = 0.05 and P < 0.001 respectively). In ovarian cancer patients, MCP-1 serum levels showed a statistically significant correlation with histological grade (Mann–Whitney U-test, P = 0.02) and age at the time of diagnosis (Mann–Whitney U-test, P = 0.03). Elevated MCP-1 serum levels prior to therapy were not associated with disease-free and overall survival (log-rank test, P = 0.2 and P = 0.7 respectively). In summary these data indicate that MCP-1 might play a functional role in the natural history of ovarian cancer and might serve as

  17. Monocyte chemoattractant protein 1 and interleukin-8 production in mononuclear cells stimulated by oral microorganisms.

    PubMed Central

    Jiang, Y; Russell, T R; Graves, D T; Cheng, H; Nong, S H; Levitz, S M

    1996-01-01

    Chemokines are a family of low-molecular-weight proinflammatory cytokines that stimulate recruitment of leukocytes. The chemokines interleukin-8 (IL-8) and monocyte chemoattractant protein 1 (MCP-1) are relatively specific chemoattractants for neutrophils and monocytes, respectively. Chemokine expression contributes to the presence of different leukocyte populations observed in normal and pathologic states. In the present studies, peripheral blood mononuclear cells (PBMC) were stimulated by microbes (Candida albicans, Streptococcus mutans, Porphyromonas gingivalis, and Actinobacillus actinomycetemcomitans) selected based upon their importance as oral pathogens. IL-8 and MCP-1 gene expression and protein release were determined by Northern blot (RNA blot) analysis and enzyme-linked immunosorbent assay. C. albicans, P. gingivalis, and A. actinomycetemcomitans induced high levels of production of both MCP-1 and IL-8. S. mutans was a strong inducer of MCP-1, but it did not stimulate significant production of IL-8. C. albicans, S. mutans, and A. actinomycetemcomitans were 500 to 5,000 times more potent than P. gingivalis in terms of MCP-1 production. In general, the microbe-to-PBMC ratios required for maximum gene expression of MCP-1 were lower than those for IL-8. However, for actual protein release of MCP-1 versus IL-8, differences in the effects of various microbe concentrations were observed only for A. actinomycetemcomitans. These results demonstrate that different oral pathogens induce specific dose-dependent patterns of chemokine gene expression and release. Such patterns may help explain the immunopathology of oral infections, particularly with regard to inflammatory leukocyte recruitment. PMID:8890191

  18. Nicotinamide downregulates gene expression of interleukin-6, interleukin-10, monocyte chemoattractant protein-1, and tumour necrosis factor-α gene expression in HaCaT keratinocytes after ultraviolet B irradiation.

    PubMed

    Monfrecola, G; Gaudiello, F; Cirillo, T; Fabbrocini, G; Balato, A; Lembo, S

    2013-03-01

    Ultraviolet (UV) radiation has profound effects on human skin, causing sunburn, inflammation, cellular-tissue injury, cell death, and skin cancer. Most of these effects are mediated by a number of cytokines produced by keratinocytes. In this study we investigated whether nicotinamide (NCT), the amide form of vitamin B3, might have a protective function in reducing the expression of interleukin (IL)-1β, IL-6, IL-8, IL-10, monocyte chemoattractant protein (MCP)-1 and tumour necrosis factor (TNF)-α in UV-irradiated keratinocytes. HaCaT cells were treated with UVB in the presence or absence of NCT, and cytokine mRNA levels were examined by quantitative real-time PCR. NCT significantly downregulated IL-6, IL-10, MCP-1 and TNF-α mRNA expression, whereas it did not exert any significant effect on IL-1β or IL-8 expression. Because of its ability to decrease these cytokine mediators after UV exposure, NCT is a possible therapy to improve or prevent conditions induced or aggravated by UV light.

  19. The Potential Role of Monocyte Chemoattractant Protein-1 for Major Depressive Disorder

    PubMed Central

    2014-01-01

    The immune hypothesis of major depressive disorder (MDD) fits well with the supposed interaction between genetic and environmental factors in disorders with a complicated etiopathogenesis. It has been suggested that infectious diseases are associated with MDD in that cytokines may play a critical role as a key modulator in the transition between infection and the development of MDD. It has been also suggested that antidepressants have immunomodulatory effects on some cytokines and cytokine receptors, although the exact mechanism has not yet been fully elucidated. Among cytokines, monocyte chemoattractant protein-1 (MCP-1) is especially well known and has attracted considerable interest owing to its immunomodulatory functions. MCP-1 is expressed in highly regionalized neuronal areas in the brain, leading to kind of modulation of neuronal activity and neuroendocrine functions commonly seen in patients with MDD. Additionally, it is involved in the control of other cytokines that have been consistently proposed as associated with the development of MDD. It also has a possible role in the neurodegenerative process of a number of central nervous system (CNS) diseases. Hence, this paper draws from the perspective of immunology to offer several suggestions about the role of MPC-1 in the development of MDD. PMID:25110491

  20. Monocyte chemoattractant protein-1 affects migration of hippocampal neural progenitors following status epilepticus in rats

    PubMed Central

    2013-01-01

    Background Epilepsy is a common brain disorder characterized by a chronic predisposition to generate spontaneous seizures. The mechanisms for epilepsy formation remain unknown. A growing body of evidence suggests the involvement of inflammatory processes in epileptogenesis. In the present study, we investigated the involvement of monocyte chemoattractant protein-1 (MCP-1) in aberrant migration of hippocampal progenitors in rats after the insult of status epilepticus (SE). Methods SE was induced with pilocarpine in Sprague–Dawley rats. Transcriptional expression of MCP-1 in the dentate gyrus (DG) was measured using quantitative real-time PCR. From 1 to 28 days after SE, the temporal profiles of MCP-1 protein expression in DG were evaluated using enzyme-linked immunosorbent assay. Chemokine (C-C motif) receptor 2 (CCR2) expression in doublecortin-positive neuronal progenitors was examined using double-labeling immunohistochemistry. The involvement of MCP-1/CCR2 signaling in aberrant neuronal progenitor migration in the epileptic hippocampus was assessed in the SE rats using a CCR2 antagonist, RS102895, and the ectopic migration of neuronal progenitors was determined using Prox1/doublecortin double immunostaining. Results After SE, MCP-1 gene was significantly upregulated and its corresponding protein expression in the DG was significantly increased on days 1 and 3. Some hilar ectopic progenitor cells of SE rats expressed the MCP-1 receptor, CCR2. Notably, the ectopic migration of neuronal progenitors into hilus was attenuated by a blockade of the MCP-1/CCR2 interaction with a selective CCR2 inhibitor, RS102895. Conclusions An increase in dentate MCP-1 is associated with seizure-induced aberrant migration of neuronal progenitors through the interaction with CCR2. The upregulation of MCP-1 after an insult of SE may play a role in the generation of epilepsy. PMID:23339567

  1. Phyllostachys edulis Compounds Inhibit Palmitic Acid-Induced Monocyte Chemoattractant Protein 1 (MCP-1) Production

    PubMed Central

    Higa, Jason K.; Liang, Zhibin; Williams, Philip G.; Panee, Jun

    2012-01-01

    Background Phyllostachys edulis Carriere (Poaceae) is a bamboo species that is part of the traditional Chinese medicine pharmacopoeia. Compounds and extracts from this species have shown potential applications towards several diseases. One of many complications found in obesity and diabetes is the link between elevated circulatory free fatty acids (FFAs) and chronic inflammation. This study aims to present a possible application of P. edulis extract in relieving inflammation caused by FFAs. Monocyte chemoattractant protein 1 (MCP-1/CCL2) is a pro-inflammatory cytokine implicated in chronic inflammation. Nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) and activator protein 1 (AP-1) are transcription factors activated in response to inflammatory stimuli, and upregulate pro-inflammatory cytokines such as MCP-1. This study examines the effect of P. edulis extract on cellular production of MCP-1 and on the NF-κB and AP-1 pathways in response to treatment with palmitic acid (PA), a FFA. Methodology/Principal Findings MCP-1 protein was measured by cytometric bead assay. NF-κB and AP-1 nuclear localization was detected by colorimetric DNA-binding ELISA. Relative MCP-1 mRNA was measured by real-time quantitative PCR. Murine cells were treated with PA to induce inflammation. PA increased expression of MCP-1 mRNA and protein, and increased nuclear localization of NF-κB and AP-1. Adding bamboo extract (BEX) inhibited the effects of PA, reduced MCP-1 production, and inhibited nuclear translocation of NF-κB and AP-1 subunits. Compounds isolated from BEX inhibited MCP-1 secretion with different potencies. Conclusions/Significance PA induced MCP-1 production in murine adipose, muscle, and liver cells. BEX ameliorated PA-induced production of MCP-1 by inhibiting nuclear translocation of NF-κB and AP-1. Two O-methylated flavones were isolated from BEX with functional effects on MCP-1 production. These results may represent a possible therapeutic

  2. Monocyte chemoattractant protein 1 (MCP-1/CCL2) contributes to thymus atrophy in acute myeloid leukemia.

    PubMed

    Driss, Virginie; Quesnel, Bruno; Brinster, Carine

    2015-02-01

    Recent studies on acute myelogenous leukemia (AML) patients have revealed the existence of T-cell immunodeficiencies, characterized by peripheral T lymphocytes that are unable to interact with blasts, reduced thymic emigrants and oligoclonal restricted repertoires. These observations suggest that there is a profound thymic dysregulation, which is difficult to study in AML patients. Using the C1498 AML mouse model, we demonstrated that leukemia development was associated with thymus atrophy, which was defined by abnormal organ weight and reduced cellularity. In addition, we observed a dramatic loss of peripheral CD4(+) and CD8(+) T-cell numbers with increased frequencies of CD4(+) FoxP3(+) regulatory and activated/memory T cells. Investigating the mechanisms leading to this atrophy, we observed a significant accumulation of the monocyte chemoattractant protein 1 (MCP-1/CCL2) in thymi of leukemic mice. Treatment of AML-bearing animals with a blocking anti-CCL2 antibody revealed a lower tumor burden, augmented antileukemic T-cell responses, and improved survival rate compared to nontreated mice. These results were not observed when neutralization of CCL2 was performed in thymectomized mice. Altogether, we show that the CCL2 protein participates in thymic atrophy in AML mice, and this could have important implications for future immunotherapeutic strategies.

  3. Endogenous monocyte chemoattractant protein-1 recruits monocytes in the zymosan peritonitis model.

    PubMed

    Ajuebor, M N; Flower, R J; Hannon, R; Christie, M; Bowers, K; Verity, A; Perretti, M

    1998-01-01

    The role of monocyte chemoattractant protein-1 (MCP-1) in the recruitment of blood-derived monocytes in a model of zymosan peritoneal inflammation was investigated. After zymosan injection (1 mg) a rapid influx of polymorphonuclear leukocytes (PMN) and monocytes into the peritoneal cavity associated with mouse MCP-1 (JE) gene activation and protein secretion in the exudates occurred. MCP-1 production (maximal at 4 h) preceded the accumulation of monocytes (F4/80-positive cells, maximally recovered between 16 and 24 h). Treatment of mice with a single injection of anti-mouse MCP-1 antibody inhibited 16-h monocyte accumulation by approximately 40%, however, a significant decrease in the number of PMN was also measured. Finally, intraperitoneal injection of murine recombinant MCP-1 (1 microg) produced a selective accumulation of monocytes (F4/80-positive cells) into the peritoneal cavity. In conclusion, we show the novel existence of a strict relationship between MCP-1 production and leukocyte accumulation in this model of acute inflammation.

  4. Proteolytic activation of monocyte chemoattractant protein-1 by plasmin underlies excitotoxic neurodegeneration in mice.

    PubMed

    Sheehan, John J; Zhou, Chun; Gravanis, Iordanis; Rogove, Andrew D; Wu, Yan-Ping; Bogenhagen, Daniel F; Tsirka, Stella E

    2007-02-14

    Exposure of neurons to high concentrations of excitatory neurotransmitters causes them to undergo excitotoxic death via multiple synergistic injury mechanisms. One of these mechanisms involves actions undertaken locally by microglia, the CNS-resident macrophages. Mice deficient in the serine protease plasmin exhibit decreased microglial migration to the site of excitatory neurotransmitter release and are resistant to excitotoxic neurodegeneration. Microglial chemotaxis can be signaled by the chemokine monocyte chemoattractant protein-1 (MCP-1)/CCL2 (CC chemokine ligand 2). We show here that mice genetically deficient for MCP-1 phenocopy plasminogen deficiency by displaying decreased microglial recruitment and resisting excitotoxic neurodegeneration. Connecting these pathways, we demonstrate that MCP-1 undergoes a proteolytic processing step mediated by plasmin. The processing, which consists of removal of the C terminus of MCP-1, enhances the potency of MCP-1 in in vitro migration assays. Finally, we show that infusion of the cleaved form of MCP-1 into the CNS restores microglial recruitment and excitotoxicity in plasminogen-deficient mice. These findings identify MCP-1 as a key downstream effector in the excitotoxic pathway triggered by plasmin and identify plasmin as an extracellular chemokine activator. Finally, our results provide a mechanism that explains the resistance of plasminogen-deficient mice to excitotoxicity.

  5. EP2/EP4 signalling inhibits monocyte chemoattractant protein-1 production induced by interleukin 1ß in synovial fibroblasts

    PubMed Central

    Largo, R; Diez-Ortego, I; Sanchez-Pernaute, O; Lopez-Armada, M; Alvarez-Soria, M; Egido, J; Herrero-Beaumont, G

    2004-01-01

    Background: Besides its proinflammatory properties, prostaglandin E2 (PGE2) acts as a regulator of the expression of inducible genes. Inhibition of PGE2 synthesis might thus result in a paradoxical deleterious effect on inflammation. Objective: To examine the effect of PGE2 on monocyte chemoattractant protein-1 (MCP-1) expression in cultured synovial fibroblasts (SF) stimulated with interleukin (IL)1ß. Methods: MCP-1 expression was assessed in SF stimulated with IL1ß in the presence of PGE2 or different NSAIDs by RT-PCR or northern blot and immunocytochemistry. Expression of cyclo-oxygenase (COX) isoforms was studied by western blot techniques. The role of PGE2 receptors (EP) in PGE2 action was assessed employing EP receptor subtype-specific agonists. Results: PGE2 significantly inhibited IL1ß induced MCP-1 expression in SF in a dose dependent manner. IL1ß increased COX-2 and did not alter COX-1 synthesis in SF. 11-Deoxy-PGE1, an EP2/EP4 agonist, reproduced PGE2 action on MCP-1 expression. Butaprost, a selective EP2 agonist, was less potent than PGE2. Sulprostone, an EP1/EP3 agonist, had no effect on IL1ß induced MCP-1 expression. Inhibition of endogenous PGE2 synthesis by NSAIDs further enhanced MCP-1 mRNA expression in IL1ß stimulated SF, an effect prevented by addition of exogenous PGE2. Conclusion: Activation of EP2/EP4 receptors down regulates the expression of MCP-1 in IL1ß stimulated SF, while PGE2 pharmacological inhibition cuts off this signalling pathway and results in a superinduction of MCP-1 expression. The data suggest that NSAIDs may intercept a natural regulatory circuit controlling the magnitude of inflammation, which questions their continuous administration in inflammatory joint diseases. PMID:15361371

  6. Loss of monocyte chemoattractant protein-1 alters macrophage polarization and reduces NFκB activation in the foreign body response.

    PubMed

    Moore, Laura Beth; Sawyer, Andrew J; Charokopos, Antonios; Skokos, Eleni A; Kyriakides, Themis R

    2015-01-01

    Implantation of biomaterials elicits a foreign body response characterized by fusion of macrophages to form foreign body giant cells and fibrotic encapsulation. Studies of the macrophage polarization involved in this response have suggested that alternative (M2) activation is associated with more favorable outcomes. Here we investigated this process in vivo by implanting mixed cellulose ester filters or polydimethylsiloxane disks in the peritoneal cavity of wild-type (WT) and monocyte chemoattractant protein-1 (MCP-1) knockout mice. We analyzed classical (M1) and alternative (M2) gene expression via quantitative polymerase chain reaction, immunohistochemistry and enzyme-linked immunosorbent assay in both non-adherent cells isolated by lavage and implant-adherent cells. Our results show that macrophages undergo unique activation that displays features of both M1 and M2 polarization including induction of tumor necrosis factor α (TNF), which induces the expression and nuclear translocation of p50 and RelA determined by immunofluorescence and Western blot. Both processes were compromised in fusion-deficient MCP-1 KO macrophages in vitro and in vivo. Furthermore, inclusion of BAY 11-7028, an inhibitor of NFκB activation, reduced nuclear translocation of RelA and fusion in WT macrophages. Our studies suggest that peritoneal implants elicit a unique macrophage polarization phenotype leading to induction of TNF and activation of the NFκB pathway. PMID:25242651

  7. Loss of monocyte chemoattractant protein-1 alters macrophage polarization and reduces NFκB activation in the foreign body response.

    PubMed

    Moore, Laura Beth; Sawyer, Andrew J; Charokopos, Antonios; Skokos, Eleni A; Kyriakides, Themis R

    2015-01-01

    Implantation of biomaterials elicits a foreign body response characterized by fusion of macrophages to form foreign body giant cells and fibrotic encapsulation. Studies of the macrophage polarization involved in this response have suggested that alternative (M2) activation is associated with more favorable outcomes. Here we investigated this process in vivo by implanting mixed cellulose ester filters or polydimethylsiloxane disks in the peritoneal cavity of wild-type (WT) and monocyte chemoattractant protein-1 (MCP-1) knockout mice. We analyzed classical (M1) and alternative (M2) gene expression via quantitative polymerase chain reaction, immunohistochemistry and enzyme-linked immunosorbent assay in both non-adherent cells isolated by lavage and implant-adherent cells. Our results show that macrophages undergo unique activation that displays features of both M1 and M2 polarization including induction of tumor necrosis factor α (TNF), which induces the expression and nuclear translocation of p50 and RelA determined by immunofluorescence and Western blot. Both processes were compromised in fusion-deficient MCP-1 KO macrophages in vitro and in vivo. Furthermore, inclusion of BAY 11-7028, an inhibitor of NFκB activation, reduced nuclear translocation of RelA and fusion in WT macrophages. Our studies suggest that peritoneal implants elicit a unique macrophage polarization phenotype leading to induction of TNF and activation of the NFκB pathway.

  8. Duffy antigen receptor for chemokines (Darc) polymorphism regulates circulating concentrations of monocyte chemoattractant protein-1 and other inflammatory mediators

    PubMed Central

    Schnabel, Renate B.; Baumert, Jens; Barbalic, Maja; Dupuis, Josée; Ellinor, Patrick T.; Durda, Peter; Dehghan, Abbas; Bis, Joshua C.; Illig, Thomas; Morrison, Alanna C.; Jenny, Nancy S.; Keaney, John F.; Gieger, Christian; Tilley, Cathy; Yamamoto, Jennifer F.; Khuseyinova, Natalie; Heiss, Gerardo; Doyle, Margaret; Blankenberg, Stefan; Herder, Christian; Walston, Jeremy D.; Zhu, Yanyan; Vasan, Ramachandran S.; Klopp, Norman; Boerwinkle, Eric; Larson, Martin G.; Psaty, Bruce M.; Peters, Annette; Ballantyne, Christie M.; Witteman, Jacqueline C. M.

    2010-01-01

    To identify the genetic basis of circulating concentrations of monocyte chemoattractant protein-1 (MCP-1), we conducted genome-wide association analyses for MCP-1 in 3 independent cohorts (n = 9598). The strongest association was for serum MCP-1 with a nonsynonymous polymorphism, rs12075 (Asp42Gly) in DARC, the gene for Duffy antigen receptor for chemokines, a known vascular reservoir of proinflammatory cytokines (minor allele frequency, 45.6%; P < 1.0 * 10−323). This association was supported by family-based genetic linkage at a locus encompassing the DARC gene (genome-wide P = 8.0 * 10−13). Asp42Gly accounted for approximately 20% of the variability in serum MCP-1 concentrations and also was associated with serum concentrations of interleukin-8 and RANTES. While exploring a lack of association between this polymorphism and EDTA plasma MCP-1 concentrations (P = .82), we determined that both clotting and exogenous heparan sulfate (unfractionated heparin) released substantial amounts of MCP-1 from Darc. Quantitative immunoflow cytometry failed to identify meaningful Asp42Gly-associated differences in Darc expression, suggesting that a functional change is responsible for the differential cytokine binding. We conclude that Asp42Gly is a major regulator of erythrocyte Darc-mediated cytokine binding and thereby the circulating concentrations of several proinflammatory cytokines. We have also identified for the first time 2 mechanisms for the release of reservoir chemokines with possible clinical implications. PMID:20040767

  9. Serum monocyte chemoattractant protein-1 is a biomarker in patients with diabetes and periodontitis

    PubMed Central

    Radhakrishnan, Preethi; Srikanth, Padma; Seshadri, Krishna G.; Barani, Ramya; Samanta, Maitreya

    2014-01-01

    Introduction: The role of serum Monocyte Chemoattractant Protein-1 (MCP-1) as a biomarker of periodontitis is well documented; however, its role in diabetic patients with periodontitis is unknown. Aim: This study was conducted to determine the presence and concentration of serum MCP-1 in diabetic patients with and without periodontitis and correlate it glycemic status with periodontitis. Materials and Methods: Adult diabetic patients were enrolled and grouped into group I, II, and III based on their glycemic status and serum MCP-1 estimated by ELISA. Linear regression and correlation tests were performed using R statistical software, Medcalc software to observe correlation between the serum MCP-1 and glycated hemoglobin level among different groups. Results: Serum samples obtained from 37 patients tested positive for MCP-1. Mean serum MCP-1 concentration was highest (482.3 pg/ml) in group III, lowest (149.3 pg/ml) in group I, and intermediate 398.8 pg/ml in group II. Correlation and regression analysis was done between HbA1c and serum MCP-1. A significant positive correlation (P < 0.001) was observed. Serum MCP-1 increased by 37.278 pg/ml for every 1% rise in HbA1c, and the levels were raised in group II and group III than in group I irrespective of their glycemic status. With an HbA1c range of 6.5-6.9% (group II), the serum MCP-1 values cluster around 380-410 pg/ml. Elevated levels of serum MCP-1 (>500 pg/ml) in three subjects corresponded to HbA1c values more than 12.2% (group III). Conclusion: To our knowledge, this is the first study to document serum MCP-1 levels in diabetic patients with periodontitis. Glycemic status influences serum MCP-1, and lack of glycemic control contributes to increased serum MCP-1 levels. Serum MCP-1 may thus serve as a biomarker of inflammation and disease progression in diabetes with periodontitis. PMID:25143907

  10. Identification of serum monocyte chemoattractant protein-1 and prolactin as potential tumor markers in hepatocellular carcinoma.

    PubMed

    Wang, Who-Whong; Ang, Soo Fan; Kumar, Rajneesh; Heah, Charmain; Utama, Andi; Tania, Navessa Padma; Li, Huihua; Tan, Sze Huey; Poo, Desmond; Choo, Su Pin; Chow, Wan Cheng; Tan, Chee Kiat; Toh, Han Chong

    2013-01-01

    Early diagnosis of hepatocellullar carcinoma (HCC) remains a challenge. The current practice of serum alpha-fetoprotein (AFP) measurement is inadequate. Here we utilized a proteomic approach to identify novel serum biomarkers for distinguishing HCC patients from non-cancer controls. We profiled the serum proteins in a group of 58 resectable HCC patients and 11 non-HCC chronic hepatitis B (HBV) carrier samples from the Singapore General Hospital (SGH) using the RayBio® L-Series 507 Antibody Array and found 113 serum markers that were significantly modulated between HCC and control groups. Selected potential biomarkers from this list were quantified using a multiplex sandwich enzyme-linked immunosorbent assay (ELISA) array in an expanded SGH cohort (126 resectable HCC patients and 115 non-HCC chronic HBV carriers (NC group)), confirming that serum prolactin and monocyte chemoattractant protein-1 (MCP-1) were significantly upregulated in HCC patients. This finding of serum MCP-1 elevation in HCC patients was validated in a separate cohort of serum samples from the Mochtar Riady Institute for Nanotechnology, Indonesia (98 resectable HCC, 101 chronic hepatitis B patients and 100 asymptomatic HBV/HCV carriers) by sandwich ELISA. MCP-1 and prolactin levels were found to correlate with AFP, while MCP-1 also correlated with disease stage. Subsequent receiver operating characteristic (ROC) analysis of AFP, prolactin and MCP-1 in the SGH cohort and comparing their area under the ROC curve (AUC) indicated that neither prolactin nor MCP-1 on their own performed better than AFP. However, the combination of AFP+MCP-1 (AUC, 0.974) had significantly superior discriminative ability than AFP alone (AUC, 0.942; p<0.001). In conclusion, prolactin and MCP-1 are overexpressed in HCC and are conveniently quantifiable in patients' sera by ELISA. MCP-1 appears to be a promising complementary biomarker for HCC diagnosis and this MCP-1+AFP model should be further evaluated as potential

  11. Induction of the Gene Encoding Macrophage Chemoattractant Protein 1 by Orientia tsutsugamushi in Human Endothelial Cells Involves Activation of Transcription Factor Activator Protein 1

    PubMed Central

    Cho, Nam-Hyuk; Seong, Seung-Yong; Huh, Myung-Sook; Kim, Na-Hyun; Choi, Myung-sik; Kim, Ik-sang

    2002-01-01

    Human macrophage chemoattractant protein 1 (MCP-1) is a potent mediator of macrophage migration and therefore plays an essential role in early events of inflammation. In endothelial cells, at least three independent pathways regulate MCP-1 expression by NF-κB and AP-1. Orientia tsutsugamushi causes vasculitis in humans by replicating inside macrophages and endothelial cells. In the present study, we investigated the cis-acting and trans-acting elements involved in O. tsutsugamushi-induced MCP-1 gene expression in human umbilical vein endothelial cells (HUVEC). Although NF-κB activation was observed in HUVEC infected with O. tsutsugamushi, inhibition of NF-κB activation did not affect the MCP-1 expression. However, treatment of HUVEC with extracellular signal-regulated kinase (ERK) kinase inhibitor or p38 mitogen-activated protein kinase (MAPK) inhibitor suppressed expression of MCP-1 mRNA concomitant with downregulation of activator protein 1 (AP-1) activation. Deletion of triphorbol acetate response elements (TRE) at position −69 to −63 of MCP-1 gene abolished inducible promoter activity. Deletion of TRE at position −69 to −63−96 to −90 or deletion of NF-κB-binding site at position −69 to −63−88 to −79 did not affect the inducibility of promoter. Site-directed mutagenesis of the NF-κB binding sites at positions −2640 to −2632, −2612 to −2603 in the enhancer region, or the AP-1 biding site at position −2276 to −2270 decreased the inducible activity of the promoter. Taken together, AP-1 activation by both the ERK pathway and the p38 MAPK pathway as well as their binding to TRE at position −69 to −63 in proximal promoter and TRE at position −2276 to −2270 in enhancer region is altogether essential in induction of MCP-1 mRNA in HUVEC infected with O. tsutsugamushi. Although NF-κB activation is not essential per se, the κB site in the enhancer region is important in MCP-1 induction of HUVEC. This discrepancy in the

  12. Up-regulation of endothelial monocyte chemoattractant protein-1 by coplanar PCB77 is caveolin-1-dependent

    SciTech Connect

    Majkova, Zuzana; Smart, Eric; Toborek, Michal; Hennig, Bernhard

    2009-05-15

    Atherosclerosis, the primary cause of heart disease and stroke is initiated in the vascular endothelium, and risk factors for its development include environmental exposure to persistent organic pollutants. Caveolae are membrane microdomains involved in regulation of many signaling pathways, and in particular in endothelial cells. We tested the hypothesis that intact caveolae are required for coplanar PCB77-induced up-regulation of monocyte chemoattractant protein-1 (MCP-1), an endothelium-derived chemokine that attracts monocytes into sub-endothelial space in early stages of the atherosclerosis development. Atherosclerosis-prone LDL-R{sup -/-} mice (control) or caveolin-1{sup -/-}/LDL-R{sup -/-} mice were treated with PCB77. PCB77 induced aortic mRNA expression and plasma protein levels of MCP-1 in control, but not caveolin-1{sup -/-}/LDL-R{sup -/-} mice. To study the mechanism of this effect, primary endothelial cells were used. PCB77 increased MCP-1 levels in endothelial cells in a time- and concentration-dependent manner. This effect was abolished by caveolin-1 silencing using siRNA. Also, MCP-1 up-regulation by PCB77 was prevented by inhibiting p38 and c-Jun N-terminal kinase (JNK), but not ERK1/2, suggesting regulatory functions via p38 and JNK MAPK pathways. Finally, pre-treatment of endothelial cells with the aryl hydrocarbon receptor (AhR) inhibitor {alpha}-naphthoflavone ({alpha}-NF) partially blocked MCP-1 up-regulation. Thus, our data demonstrate that coplanar PCB77 can induce MCP-1 expression by endothelial cells and that this effect is mediated by AhR, as well as p 38 and JNK MAPK pathways. Intact caveolae are required for these processes both in vivo and in vitro. This further supports a key role for caveolae in vascular inflammation induced by persistent organic pollutants.

  13. Monocyte chemoattractant protein-1-induced CCR2B receptor desensitization mediated by the G protein-coupled receptor kinase 2

    PubMed Central

    Aragay, A. M.; Mellado, M.; Frade, J. M. R.; Martin, A. M.; Jimenez-Sainz, M. C.; Martinez-A, C.; Mayor, F.

    1998-01-01

    Monocyte chemoattractant protein 1 (MCP-1) is a member of the chemokine cytokine family, whose physiological function is mediated by binding to the CCR2 and CCR4 receptors, which are members of the G protein-coupled receptor family. MCP-1 plays a critical role in both activation and migration of leukocytes. Rapid chemokine receptor desensitization is very likely essential for accurate chemotaxis. In this report, we show that MCP-1 binding to the CCR2 receptor in Mono Mac 1 cells promotes the rapid desensitization of MCP-1-induced calcium flux responses. This desensitization correlates with the Ser/Thr phosphorylation of the receptor and with the transient translocation of the G protein-coupled receptor kinase 2 (GRK2, also called β-adrenergic kinase 1 or βARK1) to the membrane. We also demonstrate that GRK2 and the uncoupling protein β-arrestin associate with the receptor, forming a macromolecular complex shortly after MCP-1 binding. Calcium flux responses to MCP-1 in HEK293 cells expressing the CCR2B receptor were also markedly reduced upon cotransfection with GRK2 or the homologous kinase GRK3. Nevertheless, expression of the GRK2 dominant-negative mutant βARK-K220R did not affect the initial calcium response, but favored receptor response to a subsequent challenge by agonists. The modulation of the CCR2B receptor by GRK2 suggests an important role for this kinase in the regulation of monocyte and lymphocyte response to chemokines. PMID:9501202

  14. Enhanced production of the chemotactic cytokines interleukin-8 and monocyte chemoattractant protein-1 in human abdominal aortic aneurysms.

    PubMed Central

    Koch, A. E.; Kunkel, S. L.; Pearce, W. H.; Shah, M. R.; Parikh, D.; Evanoff, H. L.; Haines, G. K.; Burdick, M. D.; Strieter, R. M.

    1993-01-01

    Inflammatory leukocytes play a central role in the pathogenesis of human atherosclerotic disease, from early atherogenesis to the late stages of atherosclerosis, such as aneurysm formation. We have shown previously that human abdominal aortic aneurysms are characterized by the presence of numerous chronic inflammatory cells throughout the vessel wall (Am J Pathol 1990, 137: 1199-1213). The signals that attract lymphocytes and monocytes into the aortic wall in aneurysmal disease remain to be precisely defined. We have studied the production of the chemotactic cytokines interleukin-8 (IL-8) and monocyte chemoattractant protein-1 (MCP-1) by aortic tissues obtained from 47 subjects. We compared the antigenic production of these cytokines by explants of: 1) human abdominal aneurysmal tissue, 2) occlusive (atherosclerotic) aortas, and 3) normal aortas. IL-8, which is chemotactic for neutrophils, lymphocytes, and endothelial cells was liberated in greater quantities by abdominal aortic aneurysms than by occlusive or normal aortas. Using immunohistochemistry, macrophages, and to a lesser degree endothelial cells, were found to be positive for the expression of antigenic IL-8. Similarly, MCP-1, a potent chemotactic cytokine for monocytes/macrophages, was released by explants from abdominal aortic aneurysms in greater quantities than by explants from occlusive or normal aortas. Using immunohistochemistry, the predominant MCP-1 antigen-positive cells were macrophages and to a lesser extent smooth muscle cells. Our results indicate that human abdominal aortic aneurysms produce IL-8 and MCP-1, both of which may serve to recruit additional inflammatory cells into the abdominal aortic wall, hence perpetuating the inflammatory reaction that may result in the pathology of vessel wall destruction and aortic aneurysm formation. Images Figure 2 Figure 3 Figure 4 Figure 5 PMID:8494046

  15. Cardiac overexpression of monocyte chemoattractant protein-1 in transgenic mice prevents cardiac dysfunction and remodeling after myocardial infarction.

    PubMed

    Morimoto, Hajime; Takahashi, Masafumi; Izawa, Atsushi; Ise, Hirohiko; Hongo, Minoru; Kolattukudy, Pappachan E; Ikeda, Uichi

    2006-10-13

    Myocardial infarction (MI) is accompanied by inflammatory responses that lead to the recruitment of leukocytes and subsequent myocardial damage, healing, and scar formation. Because monocyte chemoattractant protein-1 (MCP-1) (also known as CCL2) regulates monocytic inflammatory responses, we investigated the effect of cardiac MCP-1 overexpression on left ventricular (LV) dysfunction and remodeling in a murine MI model. Transgenic mice expressing the mouse JE-MCP-1 gene under the control of the alpha-cardiac myosin heavy chain promoter (MHC/MCP-1 mice) were used for this purpose. MHC/MCP-1 mice had reduced infarct area and scar formation and improved LV dysfunction after MI. These mice also showed induction of macrophage infiltration and neovascularization; however, few bone marrow-derived endothelial cells were detected in MHC/MCP-1 mice whose bone marrow was replaced with that of Tie2/LacZ transgenic mice. Flow cytometry analysis showed no increase in endothelial progenitor cells (CD34+/Flk-1+ cells) in MHC/MCP-1 mice. Marked myocardial interleukin (IL)-6 secretion, STAT3 activation, and LV hypertrophy were observed after MI in MHC/MCP-1 mice. Furthermore, cardiac myofibroblasts accumulated after MI in MHC/MCP-1 mice. In vitro experiments revealed that a combination of IL-6 with MCP-1 synergistically stimulated and sustained STAT3 activation in cardiomyocytes. MCP-1, IL-6, and hypoxia directly promoted the differentiation of cardiac fibroblasts into myofibroblasts. Our results suggest that cardiac overexpression of MCP-1 induced macrophage infiltration, neovascularization, myocardial IL-6 secretion, and accumulation of cardiac myofibroblasts, thereby resulting in the prevention of LV dysfunction and remodeling after MI. They also provide a new insight into the role of cardiac MCP-1 in the pathophysiology of MI. PMID:16990567

  16. Investigation of the functional role played by the chemokine monocyte chemoattractant protein-1 in interleukin-1-induced murine peritonitis

    PubMed Central

    Ajuebor, Maureen N; Gibbs, Linda; Flower, Roderick J; Das, Anuk M; Perretti, Mauro

    1998-01-01

    Intraperitoneal (i.p.) injection of murine recombinant IL-1β (mrIL-1β) produced a dose-dependent (0.5–50 ng) and time-related (0.5–2 h) secretion of murine monocyte chemoattractant protein-1 (mMCP-1; 3–4 ng per cavity) in the lavage fluids. MCP-1 mRNA could also be detected in the cell pellets by reverse transcriptase-polymerase chain reaction (RT-PCR).MCP-1 levels were reduced by more than 90% by co-administration of IL-1 receptor antagonist (10 μg) (n=6, P<0.05). In contrast, an IL-1 mutant with low affinity for IL-1 receptor type I, termed yIL-1βΔ4 (50 ng), produced only a modest release of the chemokine. Treatment of mice with dexamethasone (DEX) (∼1 mg kg−1 s.c.) reduced mrIL-1β-induced mMCP-1 gene expression (apparent total inhibition) and protein release in the lavage fluids (∼40% reduction; n=10; P<0.05). Drastic reductions in the numbers of residential macrophages or mast cells did not modify the levels of mMCP-1 recovered in the lavage fluids.Injection of mrIL-1β produced neutrophil accumulation into the peritoneal cavities (maximal at 4 h with 1.42±0.15×106 cells per mouse). Co-injection of a specific polyclonal antibody against mMCP-1 reduced this process by more than 50% (n=6; P<0.05). In conclusion, we studied the mechanisms leading to the specific release of the CC chemokine mMCP-1 after in vivo administration of mrIL-1β. PMID:9786504

  17. Pattern recognition of monocyte chemoattractant protein-1 (MCP-1) in whole blood samples using new platforms based on nanostructured materials.

    PubMed

    Stefan-van Staden, Raluca-Ioana; Gugoasa, Livia Alexandra; Biris, Alexandru Radu

    2015-09-28

    Four stochastic microsensors based on nanostructured materials (graphene, maltodextrin (MD), and diamond) integrated in miniaturized platforms were proposed. Monocyte chemoattractant protein-1 (MCP-1) is a pro-inflammatory cytokine whose main function is to regulate cell trafficking. It is correlated with the incidence of cardiovascular diseases and obesity, and was used as the model analyte in this study. The screening of whole blood samples for MCP-1 can be done for concentrations ranging from 10(-12) to 10(-8) g mL(-1). The method was used for both qualitative and quantitative assessments of MCP-1 in whole blood samples. The lowest quantification limits for the assay of MCP-1 (1 pg mL(-1)) were reached when the microsensors based on protoporphyrin IX/Graphene-Au-3 and on MD/Graphene were employed in the platform design. PMID:26183340

  18. Pattern recognition of monocyte chemoattractant protein-1 (MCP-1) in whole blood samples using new platforms based on nanostructured materials

    NASA Astrophysics Data System (ADS)

    Stefan-van Staden, Raluca-Ioana; Gugoasa, Livia Alexandra; Biris, Alexandru Radu

    2015-09-01

    Four stochastic microsensors based on nanostructured materials (graphene, maltodextrin (MD), and diamond) integrated in miniaturized platforms were proposed. Monocyte chemoattractant protein-1 (MCP-1) is a pro-inflammatory cytokine whose main function is to regulate cell trafficking. It is correlated with the incidence of cardiovascular diseases and obesity, and was used as the model analyte in this study. The screening of whole blood samples for MCP-1 can be done for concentrations ranging from 10-12 to 10-8 g mL-1. The method was used for both qualitative and quantitative assessments of MCP-1 in whole blood samples. The lowest quantification limits for the assay of MCP-1 (1 pg mL-1) were reached when the microsensors based on protoporphyrin IX/Graphene-Au-3 and on MD/Graphene were employed in the platform design.

  19. Biocompatibility of heparin-grafted hemodialysis membranes: impact on monocyte chemoattractant protein-1 circulating level and oxidative status.

    PubMed

    Morena, Marion; Jaussent, Isabelle; Chalabi, Lotfi; Bargnoux, Anne-Sophie; Dupuy, Anne-Marie; Badiou, Stéphanie; Rakic, Claire; Thomas, Michel; Canaud, Bernard; Cristol, Jean-Paul; Michel, Françoise

    2010-10-01

    This prospective observational study aimed at evaluating efficacy and biocompatibility performances of the new heparin-coated Evodial dialyzers with/without systemic heparin reduction. After a 4-week wash-out period with reference polysulfone F70S dialyzers, 6 hemodialysis patients were sequentially dialyzed with Evodial, F70S, and Evodial dialyzers using 30% heparin reduction, each period of treatment was 4 weeks. Removal rates (RR) (urea, creatinine, and β2-microglobulin), dialysis dose, and instantaneous clearances (urea and creatinine) were measured as well as inflammatory (C-reactive protein, fibrinogen, interleukin 6, tumor necrosis factor α, and monocyte chemoattractant protein-1) and oxidative stress (OS) (superoxide anion, homocysteine, and isoprostanes) parameters at the end of each study period. Patients treated with Evodial or F70S dialyzers for 4 weeks presented comparable dialysis efficacy parameters including urea and creatinine RR, dialysis dose and instantaneous clearances. By contrast, a significantly lower but reasonably good β2-microglobulin RR was achieved with Evodial dialyzers. Regarding biocompatibility, no significant difference was observed with inflammation and OS except for postdialysis monocyte chemoattractant protein-1 which significantly decreased with Evodial dialyzers. Thirty percent heparinization reduction with Evodial dialyzers did not induce any change in inflammation but led to an improvement in OS as demonstrated by a decrease in postdialysis superoxide production and predialysis homocysteine and isoprostane. This bioactive dialyzer together with heparin dose reduction represents a good trade-off between efficacy and biocompatibility performance (improvement in OS with a weak decrease in efficacy) and its use is encouraging for hemodialysis patients not only in reducing OS but also in improving patient comorbid conditions due to lesser heparin side effects.

  20. Antagonists of monocyte chemoattractant protein 1 identified by modification of functionally critical NH2-terminal residues

    PubMed Central

    1995-01-01

    Monocyte chemoattractant protein (MCP)-1 analogues were designed to determine the role of the NH2-terminal region in structure and function. The NH2-terminal residue was important for function and receptor binding, as it could not be deleted or extended. However the NH2-terminal pyroglutamate residue of the wild type was not essential as it could be replaced by several other noncyclic amino acids without loss of activity. Residues 7-10 were essential for receptor desensitization, but were not sufficient for function, and the integrity of residues 1-6 were required for functional activity. A peptide corresponding to MCP-1, 1-10 lacked detectable receptor-binding activities, indicating that residues 1-10 are essential for MCP-1 function, but that other residues are also involved. Several truncated analogues, including 8-76, 9-76, and 10-76, desensitized MCP-1-induced Ca2+ induction, but were not significantly active. These analogues were antagonists of MCP-1 activity with the most potent being the 9-76 analogue (IC50 = 20 nM) The 9-76 specifically bound to MCP-1 receptors with a Kd of 8.3 nM, which was three-fold higher than MCP-1 (Kd 2.8 nM). The 9-76 analogue desensitized the Ca2+ response to MCP-1 and MCP- 3, but not to other CC chemokines, suggesting that it is MCP receptor specific. The availability of these compounds will be helpful in evaluating MCP receptor antagonists as anti-inflammatory therapeutics. PMID:7836918

  1. Urine Monocyte Chemoattractant Protein-1 and Lupus Nephritis Disease Activity: Preliminary Report of a Prospective Longitudinal Study

    PubMed Central

    Alharazy, Sabah; Kong, Norella C. T.; Mohd, Marlyn; Shah, Shamsul A.; Ba'in, Arbaiyah; Abdul Gafor, Abdul Halim

    2015-01-01

    Objective. This longitudinal study aimed to determine the urine monocyte chemoattractant protein-1 (uMCP-1) levels in patients with biopsy-proven lupus nephritis (LN) at various stages of renal disease activity and to compare them to current standard markers. Methods. Patients with LN—active or inactive—had their uMCP-1 levels and standard disease activity markers measured at baseline and 2 and 4 months. Urinary parameters, renal function test, serological markers, and renal SLE disease activity index-2K (renal SLEDAI-2K) were analyzed to determine their associations with uMCP-1. Results. A hundred patients completed the study. At each visit, uMCP-1 levels (pg/mg creatinine) were significantly higher in the active group especially with relapses and were significantly associated with proteinuria and renal SLEDAI-2K. Receiver operating characteristic (ROC) curves showed that uMCP-1 was a potential biomarker for LN. Whereas multiple logistic regression analysis showed that only proteinuria and serum albumin and not uMCP-1 were independent predictors of LN activity. Conclusion. uMCP-1 was increased in active LN. Although uMCP-1 was not an independent predictor for LN activity, it could serve as an adjunctive marker when the clinical diagnosis of LN especially early relapse remains uncertain. Larger and longer studies are indicated. PMID:26246906

  2. The therapeutic effect of monocyte chemoattractant protein-1 delivered by an electrospun scaffold for hyperglycemia and nephrotic disorders.

    PubMed

    Yong, Cai; Wang, Zhengxin; Zhang, Xing; Shi, Xiaomin; Ni, Zhijia; Fu, Hong; Ding, Guoshan; Fu, Zhiren; Yin, Hao

    2014-01-01

    Here, we investigated in diabetic mice the therapeutic effect of monocyte chemoattractant protein-1 (MCP-1), locally delivered by an electrospun scaffold, on transplanted islets. This therapeutic scheme is expected to exert a synergistic effect to ameliorate hyperglycemia and its associated nephrotic disorders. The cumulative amount of MCP-1 released from the scaffold in vitro within a 3-week window was 267.77 ± 32.18 ng, without a compromise in bioactivity. After 8 weeks following the transplantation, the islet population stimulated by MCP-1 was 35.14%± 7.23% larger than the non-stimulated islet population. Moreover, MCP-1 increased concentrations of blood insulin and C-peptide 2 by 49.83%± 5.29% and 43.49%± 9.21%, respectively. Consequently, the blood glucose concentration in the MCP-1 group was significantly lower than that in the control group at week 2 post-surgery. MCP-1 also enhanced the tolerance of sudden oral glucose challenge. The rapid decrease of blood creatinine, urine creatinine, and blood urea nitrogen suggested that the recovery of renal functions compromised by hyperglycemia could also be attributed to MCP-1. Our study shed new light on a synergistic strategy to alleviate hyperglycemia and nephrotic disorders in diabetic patients. PMID:24600221

  3. Host-Derived Smooth Muscle Cells Accumulate in Cardiac Allografts: Role of Inflammation and Monocyte Chemoattractant Protein 1

    PubMed Central

    Bojakowski, Krzysztof; Soin, Joanna; Nozynski, Jerzy; Zakliczynski, Michal; Gaciong, Zbigniew; Zembala, Marian; Söderberg-Nauclér, Cecilia

    2009-01-01

    Transplant arteriosclerosis is characterized by inflammation and intimal thickening caused by accumulation of smooth muscle cells (SMCs) both from donor and recipient. We assessed the relationship between clinical factors and the presence of host-derived SMCs in 124 myocardial biopsies from 26 consecutive patients who received hearts from opposite-sex donors. Clinical and demographic information was obtained from the patients' medical records. Host-derived SMCs accounted for 3.35±2.3% of cells in arterioles (range, 0.08–12.51%). As shown by linear regression analysis, an increased number of SMCs was associated with rejection grade (mean, 1.41±1.03, p = 0.034) and the number of leukocytes (19.1±12.7 per 20 high-power fields, p = 0.01). The accumulation of host-derived SMCs was associated with an increased number of leukocytes in the allografts. In vitro, monocyte chemoattractant protein 1 (MCP-1) released from leukocytes was crucial for SMC migration. After heart allotransplantion, mice treated with MCP-1-specific antibodies had significantly fewer host-derived SMCs in the grafts than mice treated with isotypic antibody controls. We conclude that the number of host-derived SMCs in human cardiac allografts is associated with the rejection grade and that MCP-1 may play pivotal role in recruiting host-derived SMCs into cardiac allografts. PMID:19142231

  4. Increased interleukin-8 and monocyte chemoattractant protein-1 concentrations in mechanically ventilated preterm infants with pulmonary hemorrhage.

    PubMed

    Baier, R John; Loggins, John; Kruger, Thomas E

    2002-08-01

    Pulmonary hemorrhage (PH) is a serious complication causing acute respiratory distress in the premature infant, and it is associated with significant mortality and morbidity. The role of inflammatory mediators in this condition is largely undefined. Serial tracheal aspirates (TA) were obtained at intervals from 65 mechanically ventilated infants with birth weights less than 1,250 g during the first 21 days of life. Clinically significant PH developed in 15 infants. TA concentrations of interleukin-8 (IL-8) and monocyte chemoattractant protein-1 (MCP-1) were determined by enzyme-linked immunosorbent assay (ELISA).PH was associated with an increased risk of death, bronchopulmonary dysplasia, intraventricular hemorrhage, and prolonged need for mechanical ventilation and supplemental oxygen. TA aspirate concentrations of IL-8 and MCP-1 (P = 0.001, ANOVA) were significantly increased in infants with PH compared to infants who did not develop this condition. TA cytokine concentrations were also significantly increased in infants who developed bronchopulmonary dysplasia (BPD). Peak TA concentrations of IL-8 and MCP-1 were significantly higher in infants with poor outcome (BPD or death). TA MCP-1 but not IL-8 concentrations were significantly higher in infants who were oxygen-dependent at 36 weeks postconceptional age. These data suggest a pathogenic role for IL-8 and MCP-1 in the development of adverse pulmonary outcome in preterm infants with clinically significant PH.

  5. Hepatocyte growth factor induction of macrophage chemoattractant protein-1 and osteophyte-inducing factors in osteoarthritis.

    PubMed

    Dankbar, Berno; Neugebauer, Katja; Wunrau, Christina; Tibesku, Carsten O; Skwara, Adrian; Pap, Thomas; Fuchs-Winkelmann, Susanne

    2007-05-01

    In osteoarthritis (OA), hepatocyte growth factor (HGF) is supposed to play a role in cartilage repair. Because the development of osteophytes is a major characteristic of OA and thought to be part of an attempted repair process, the purpose of this study was to determine whether HGF may be involved in osteophyte formation. HGF levels in synovial fluids from 41 patients assessed by enzyme immunosorbant assay were correlated with disease severity and osteophyte formation, evaluated by anteroposterior weight-bearing radiographs. Detection of HGF, c-Met, and CD68 in cartilage and synovial tissues was assessed by immunohistochemistry. Effects of HGF on the secretion of TGF-beta1 and BMP-2 by chondrocytes, fibroblast-like synovial cells (FLS), and macrophages as well as HGF-induced secretion of MCP-1 by FLS and chondrocytes were determined by ELISA. HGF was detected in all synovial fluids and concentrations correlated highly with disease severity and osteophyte formation (p < 0.001). Immunohistochemistry revealed weak synovial staining for HGF, whereas increasing numbers of HGF expressing chondrocytes were detected depending on disease severity. In addition, an increased number of macrophages in synovial specimens was observed, which was likewise severity dependent. In a series of subsequent in vitro studies, HGF remarkable induced MCP-1 secretion by FLS in a dose-dependent manner. No effect on TGF-beta1 and BMP-2 secretion by FLS and chondrocytes was evident upon HGF stimulation, whereas secretion of these growth factors by PMA-differentiated THP-1 cells was significantly increased by HGF. The results indicate that HGF may facilitate osteophyte development by promoting MCP-1-mediated entry of monocytes/macrophages into the OA-affected joint and/or by stimulating macrophage-derived growth factors.

  6. Inhibitory effects of C-type natriuretic peptide on the differentiation of cardiac fibroblasts, and secretion of monocyte chemoattractant protein-1 and plasminogen activator inhibitor-1.

    PubMed

    Li, Zhi-Qiang; Liu, Ying-Long; Li, Gang; Li, Bin; Liu, Yang; Li, Xiao-Feng; Liu, Ai-Jun

    2015-01-01

    The present study aimed to investigate the effect of C-type natriuretic peptide (CNP) on the function of cardiac fibroblasts (CFs). Western blotting was used to investigate the expression of myofibroblast marker proteins: α-smooth muscle actin (α-SMA), extra domain-A fibronectin, collagen I and collagen III, and the activity of extracellular signal-regulated kinase 1/2 (ERK1/2). Immunofluorescence was used to examine the morphological changes; a transwell assay was used to analyze migration, and reverse transcription-quantitative polymerase chain reaction and ELISA were employed to determine the mRNA expression and protein secretion of monocyte chemoattractant protein-1 (MCP-1) and plasminogen activator inhibitor-1 (PAI-1). The results demonstrated that CNP significantly reduced the protein expression of α-SMA, fibronectin, collagen I and collagen III, and suppressed the migratory ability of CFs. Additionally, the mRNA and protein expression of MCP-1 and PAI-1 was inhibited under the CNP treatment; and this effect was mediated by the inhibition of the ERK1/2 activity. In conclusion, CNP inhibited cardiac fibroblast differentiation and migration, and reduced the secretion of MCP-1 and PAI-1, which demonstrates novel mechanisms to explain the antifibrotic effect of CNP.

  7. Radiation-Induced Thymidine Phosphorylase Upregulation in Rectal Cancer Is Mediated by Tumor-Associated Macrophages by Monocyte Chemoattractant Protein-1 From Cancer Cells

    SciTech Connect

    Kim, Tae-Dong; Li Ge; Song, Kyoung-Sub; Kim, Jin-Man; Kim, Jun-Sang; Kim, Jong-Seok; Yun, Eun-Jin; Park, Jong-Il; Park, Hae-Duck; Hwang, Byung-Doo; Lim, Kyu Yoon, Wan-Hee

    2009-03-01

    Purpose: The mechanisms of thymidine phosphorylase (TP) regulation induced by radiation therapy (XRT) in various tumors are poorly understood. We investigated the effect and mechanisms of preoperative XRT on TP expression in rectal cancer tissues. Methods and Materials: TP expression and CD68 and monocyte chemoattractant protein-1 (MCP-1) levels in rectal cancer tissues and cancer cell lines were evaluated before and after XRT in Western blotting, immunohistochemistry, enzyme-linked immunoassay, and reverse transcription-polymerase chain reaction studies. Isolated peripheral blood monocytes were used in the study of chemotaxis under the influence of MCP-1 released by irradiated colon cancer cells. Results: Expression of TP was significantly elevated by 9 Gy of XRT in most rectal cancer tissues but not by higher doses of XRT. In keeping with the close correlation of the increase in both TP expression and the number of tumor-associated macrophages (TAMs), anti-TP immunoreactivity was found in the CD68-positive TAMs and not the neoplastic cells. Expression of MCP-1 was increased in most cases after XRT, and this increase was strongly correlated with TP expression. However, this increase in MCP-1 expression occurred in tumor cells and not stromal cells. The XRT upregulated MCP-1 mRNA and also triggered the release of MCP-1 protein from cultured colon cancer cells. The supernatant of irradiated colon cancer cells showed strong chemotactic activity for monocyte migration, but this activity was completely abolished by neutralizing antibody. Conclusions: Use of XRT induces MCP-1 expression in cancer cells, which causes circulating monocytes to be recruited into TAMs, which then upregulate TP expression in rectal cancer tissues.

  8. Urinary monocyte chemoattractant protein-1 and hepcidin and early diabetic nephropathy lesions in type 1 diabetes mellitus

    PubMed Central

    Fufaa, Gudeta D.; Weil, E. Jennifer; Nelson, Robert G.; Hanson, Robert L.; Knowler, William C.; Rovin, Brad H.; Wu, Haifeng; Klein, Jon B.; Mifflin, Theodore E.; Feldman, Harold I.; Vasan, Ramachandran S.; Kimmel, Paul L.; Kusek, John W.; Mauer, Michael; Zinman, Bernard; Donnelly, Sandra; Canada, Toronto; Gardiner, Robert; Suissa, Samy; Drummond, Keith; Goodyer, Paul; Sinaiko, Alan; Strand, Trudy; Gubler, Marie Claire; Klein, Ronald

    2015-01-01

    Background Urinary monocyte chemoattractant protein-1 (MCP-1) and hepcidin are potential biomarkers of renal inflammation. We examined their association with development of diabetic nephropathy (DN) lesions in normotensive normoalbuminuric subjects with type 1 diabetes (T1D) from the Renin-Angiotensin System Study. Methods Biomarker concentrations were measured in baseline urine samples from 224 subjects who underwent kidney biopsies at baseline and after 5 years. Fifty-eight urine samples below the limit of quantitation (LOQ, 28.8 pg/mL) of the MCP-1 assay were assigned concentrations of LOQ/√2 for analysis. Relationships between ln(MCP-1/Cr) or ln(hepcidin/Cr) and morphometric variables were assessed by sex using multiple linear regression after adjustment for age, T1D duration, HbA1c, mean arterial pressure, albumin excretion rate (AER) and glomerular filtration rate (GFR). In models that examined changes in morphometric variables, the baseline morphometric value was also included. Results Baseline mean age was 24.6 years, mean duration of T1D 11.2 years, median AER 6.4 µg/min and mean iohexol GFR 129 mL/min/1.73 m2. No associations were found between hepcidin/Cr and morphometric variables. Higher MCP-1/Cr was associated with higher interstitial fractional volume at baseline and after 5 years in women (baseline partial r = 0.244, P = 0.024; 5-year partial r = 0.299, P = 0.005), but not in men (baseline partial r = −0.049, P = 0.678; 5-year partial r = 0.026, P = 0.830). MCP-1 was not associated with glomerular lesions in either sex. Conclusions Elevated urinary MCP-1 concentration measured before clinical findings of DN in women with T1D was associated with changes in kidney interstitial volume, suggesting that inflammatory processes may be involved in the pathogenesis of early interstitial changes in DN. PMID:25648911

  9. Urine Monocyte Chemoattractant Protein-1 Is an Independent Predictive Factor of Hospital Readmission and Survival in Cirrhosis

    PubMed Central

    Graupera, Isabel; Solà, Elsa; Fabrellas, Núria; Moreira, Rebeca; Solé, Cristina; Huelin, Patricia; de la Prada, Gloria; Pose, Elisa; Ariza, Xavier; Risso, Alessandro; Albertos, Sonia; Morales-Ruiz, Manuel; Jiménez, Wladimiro; Ginès, Pere

    2016-01-01

    MCP-1 (monocyte chemoattractant protein-1) is a proinflammatory cytokine involved in chemotaxis of monocytes. In several diseases, such as acute coronary syndromes and heart failure, elevated MCP-1 levels have been associated with poor outcomes. Little is known about MCP-1 in cirrhosis. AIM: To investigate the relationship between MCP-1 and outcome in decompensated cirrhosis. METHODS: Prospective study of 218 patients discharged from hospital after an admission for complications of cirrhosis. Urine and plasma levels of MCP-1 and other urine proinflammatroy biomarkers: osteopontin(OPN), trefoil-factor3 and liver-fatty-acid-binding protein were measured at admission. Urine non-inflammatory mediators cystatin-C, β2microglobulin and albumin were measured as control biomarkers. The relationship between these biomarkers and the 3-month hospital readmission, complications of cirrhosis, and mortality were assessed. RESULTS: 69 patients(32%) had at least one readmission during the 3-month period of follow-up and 30 patients died(14%). Urine MCP-1 and OPN levels, were associated with 3-month probability of readmission (0.85 (0.27–2.1) and 2003 (705–4586) ug/g creat vs 0.47 (0.2–1.1) and 1188 (512–2958) ug/g creat, in patients with and without readmission, respectively; p<0.05; median (IQR)). Furthermore, urine levels of MCP-1 were significantly associated with mortality (1.01 (1–3.6) vs 0.5 (0.2–1.1) μg/g creat, in dead and alive patients at 3 months; p<0.05). Patients with higher levels of urine MCP-1 (above percentile 75th) had higher probability of development of hepatic encephalopathy, bacterial infections or AKI. Urine MCP-1 was an independent predictive factor of hospital readmission and combined end-point of readmission or dead at 3 months. Plasma levels of MCP-1 did not correlated with outcomes. CONCLUSION: Urine, but not plasma, MCP-1 levels are associated with hospital readmission, development of complications of cirrhosis, and mortality. These

  10. Relationships of single nucleotide polymorphisms of monocyte chemoattractant protein 1 and chemokine receptor 2 with susceptibility and clinicopathologic characteristics of neoplasia of uterine cervix in Taiwan women.

    PubMed

    Wu, Hsin-Hung; Lee, Tsung-Hsien; Tee, Yi-Torng; Chen, Shiuan-Chih; Yang, Shun-Fa; Lee, Shu-Kuei; Ko, Jiunn-Liang; Wang, Po-Hui

    2013-10-01

    Few studies reported the implication of single nucleotide polymorphisms (SNPs) of monocyte chemoattractant protein 1 (MCP-1) and its receptor chemokine receptor 2 (CCR-2) in clinical significance of cancer of uterine cervix. We hypothesized that SNPs of MCP-1 and CCR-2 may affect the expression of these genes and then proteins. Therefore, we investigated the influence of the gene polymorphisms of MCP-1 and CCR-2 on the susceptibility and clinicopathologic characteristics of cervical neoplasia in Taiwan women. We recruited 86 patients with invasive cancer and 61 with high-grade dysplasia and 253 control women and selected 1 MCP-1 SNP rs1024611 (-2518G/A) and 1 CCR-2 SNP rs1799864 (190G/A; V64I) to determine their genotypes distribution using polymerase chain reaction-restriction fragment length polymorphism. In comparison to normal individuals with homozygotes GG in MCP-2 SNP, women with GA or AA carried a 2.01 odds ratio of developing cervical cancer. Nevertheless, it was not demonstrated in CCR-2 SNP. Furthermore, women with mutant homozygote (AA) of MCP-1 SNP increased the risk of deep stromal invasion, large tumor diameter, and parametrium invasion of cervical cancer, when compared to those with wild homozygote GG or heterozygote GA. However, women with mutant homozygotes (AA) of CCR-2 SNP did not increase the risk of poor clinicopathologic characteristics. In conclusion, MCP-1 SNP may be correlated with the development, deep stromal invasion, large tumor diameter, and parametrium invasion of cervical cancer but not with cancer recurrence or survival of Taiwan women patients with cancer. However, the SNP of its receptor, CCR-2, is not implicated in cervical cancer.

  11. Urinary monocyte chemoattractant protein-1 (MCP-1) and connective tissue growth factor (CCN2) as prognostic markers for progression of diabetic nephropathy.

    PubMed

    Tam, Frederick W K; Riser, Bruce L; Meeran, Karim; Rambow, JoAnn; Pusey, Charles D; Frankel, Andrew H

    2009-07-01

    Profibrotic growth factors and inflammatory chemokines have been implicated in the pathogenesis of diabetic nephropathy (DN). However, measurement of urinary monocyte chemoattractant protein-1 (MCP-1) and connective tissue growth factor (CCN2) as prognostic markers has not previously been reported, and neither have two such molecules in urine been examined in a single study of DN. In this prospective observational study, 43 adult diabetic patients were studied, 40 were followed up for 6years. Urinary MCP-1/creatinine ratios were found to be significantly higher in patients with macroalbuminuria (3.3- and 2.1-fold higher (p<0.01) than normoalbuminuric and microalbuminuric patients, respectively). CCN2 exhibited a pattern different from that of urinary MCP-1. Urinary CCN2/creatinine ratios were greatly elevated in both microalbuminuric and macroalbuminuric patients (125- and 74-fold higher than normoalbuminuric patients, respectively, p<0.01 and p<0.05, respectively). Further, urinary CCN2, but not MCP-1, correlated with progression of microalbuminuria (R=0.49, p<0.05). In contrast, MCP-1, but not CCN2, correlated with the rate of eGFR decline for all patients (R=0.61, p<0.0001), reflective of its predictive value in patients with macroalbuminuria, but not for patients with microalbuminuria or normoalbuminuria. In conclusion, increased urinary CCN2 is associated with the early progression of DN, whereas MCP-1 is associated with later stage disease. PMID:19409809

  12. Urinary monocyte chemoattractant protein 1 and alpha 1 acid glycoprotein as biomarkers of renal disease activity in juvenile-onset systemic lupus erythematosus.

    PubMed

    Watson, L; Midgley, A; Pilkington, C; Tullus, K; Marks, Sd; Holt, Rcl; Jones, Ca; Beresford, Mw

    2012-04-01

    A higher proportion of patients with juvenile-onset systemic lupus erythematosus (JSLE) will have renal involvement compared with adult-onset disease, some progressing to renal failure in adulthood. Histological examination is the gold standard for diagnosing lupus nephritis (LN), but its invasive nature limits routine use. Using cross-sectional cohort analysis, we aimed to determine whether urinary concentrations of monocyte chemoattractant protein-1 (MCP1), alpha-1-acid glycoprotein (AGP) and interferon-inducible protein 10 (IP10) are biomarkers of active LN. Sixty JSLE patients recruited to the UK JSLE Cohort Study were categorized according to the British Isles Lupus Assessment Group (BILAG) activity index. Patients with active renal JSLE (n = 8; renal BILAG score A, B) had significantly higher urinary MCP1 concentrations than patients with inactive renal disease (n = 52; renal BILAG score C, D, E; 582 pg/mg creatinine [Cr], 207 pg/mg Cr; p = 0.018) or healthy controls (n = 23; 117 pg/mg Cr; p = 0.005). Urinary AGP concentration was significantly elevated in patients with active renal disease compared with inactive renal disease (1517 ng/mg Cr, 485 ng/mg Cr; p = 0.027) or healthy controls (313 ng/mg Cr; p = 0.013). Urinary IP10 concentration was not significantly different between groups, but did strongly correlate with uMCP and uAGP levels (rho = 0.38, p = 0.009; rho = 0.33, p = 0.021). Urinary MCP1 and AGP are biomarkers of LN, providing insight into its pathophysiology. Longitudinal studies are warranted.

  13. Mutant monocyte chemoattractant protein 1 protein attenuates migration of and inflammatory cytokine release by macrophages exposed to orthopedic implant wear particles

    PubMed Central

    Yao, Zhenyu; Keeney, Michael; Lin, Tzu-Hua; Pajarinen, Jukka; Barcay, Katherine; Waters, Heather; Egashira, Kensuke; Yang, Fan; Goodman, Stuart

    2014-01-01

    Wear particles generated from total joint replacements can stimulate macrophages to release chemokines, such as monocyte chemoattractant protein 1 (MCP-1), which is the most important chemokine regulating systemic and local cell trafficking and infiltration of monocyte/macrophages in chronic inflammation. One possible strategy to curtail the adverse events associated with wear particles is to mitigate migration and activation of monocyte/macrophages. The purpose of this study is to modulate the adverse effects of particulate biomaterials and inflammatory stimuli such as endotoxin by interfering with the biological effects of the chemokine MCP-1. In the current study, the function of MCP-1 was inhibited by the mutant MCP-1 protein called 7ND, which blocks its receptor, the C–C chemokine receptor type 2 (CCR2) on macrophages. Addition of 7ND decreased MCP-1-induced migration of THP-1 cells in cell migration experiments in a dose-dependent manner. Conditioned media from murine macrophages exposed to clinically relevant polymethylmethacrylate (PMMA) particles with/without endotoxin [lipopolysaccharide (LPS)] had a chemotactic effect on human macrophages, which was decreased dramatically by 7ND. 7ND demonstrated no adverse effects on the viability of macrophages, and the capability of mesenchymal stem cells (MSCs) to form bone at the doses tested. Finally, proinflammatory cytokine production was mitigated when macrophages were exposed to PMMA particles with/without LPS in the presence of 7ND. Our studies confirm that the MCP-1 mutant protein 7ND can decrease macrophage migration and inflammatory cytokine release without adverse effects at the doses tested. Local delivery of 7ND at the implant site may provide a therapeutic strategy to diminish particle-associated periprosthetic inflammation and osteolysis. PMID:24123855

  14. Regulatory roles of tumor necrosis factor-alpha and interleukin-1 beta in monocyte chemoattractant protein-1-mediated pulmonary granuloma formation in the rat.

    PubMed Central

    Flory, C. M.; Jones, M. L.; Miller, B. F.; Warren, J. S.

    1995-01-01

    Intravenous infusion of particulate yeast cell wall glucan into rats results in the synchronous development of angiocentric pulmonary granulomas that are composed almost entirely of monocytes and macrophages. Previous studies indicate that locally produced monocyte chemoattractant protein-1 (MCP-1) is required for full granuloma development. Because tumor necrosis factor-alpha (TNF-alpha) and interleukin 1 (IL-1) can induce MCP-1 production in a variety of cell types, we sought to determine their potential regulatory roles in this model. A single infusion of anti-TNF-alpha antibody at the time of glucan infusion (time 0) markedly reduced MCP-1 mRNA levels at 1 and 6 hours but not at later time points; there was no effect on granuloma size or number measured at 48 hours. When multiple infusions of anti-TNF-alpha antibody were administered over a 23-hour period (0 to 23 hours), MCP-1 mRNA was reduced through 24 hours, there was a significant reduction in peak bronchoalveolar lavage fluid MCP-1 activity at 48 hours, and there were marked reductions in granuloma size and number at 48 hours. Similar results were observed in animals that received infusions of anti-IL-1 beta. Infusion of anti-IL-1 beta at time 0 resulted in moderate reductions in MCP-1 mRNA at 1 and 6 hours and had no effect on granuloma size or number measured at 48 hours. When multiple infusions of anti-IL-1 beta were administered over a 23-hour period (0 to 23 hours), MCP-1 mRNA was reduced through 24 hours, there was a moderate reduction in peak bronchoalveolar lavage fluid MCP-1 activity at 48 hours, and there were marked reductions in granuloma size and number at 48 hours. A single infusion of anti-TNF-alpha and anti-IL-1 beta together at time 0 resulted in marked reductions in whole lung MCP-1 and mRNA at 1 and 6 hours, but not at 24 hours. Multiple combined infusions of anti-TNF-alpha and anti-IL-1 beta over a 23-hour period resulted in additive reductions in MCP-1 mRNA through 24 hours

  15. Gut Microbiota in Type 2 Diabetes Individuals and Correlation with Monocyte Chemoattractant Protein1 and Interferon Gamma from Patients Attending a Tertiary Care Centre in Chennai, India

    PubMed Central

    Pushpanathan, Premalatha; Srikanth, Padma; Seshadri, Krishna G.; Selvarajan, Sribal; Pitani, Ravi Shankar; Kumar, Thomas David; Janarthanan, R.

    2016-01-01

    Background: Type 2 diabetes mellitus (T2DM) and obesity are associated with changes in gut microbiota and characterized by chronic low-grade inflammation. Monocyte chemoattractant protein-1 (MCP-1) and interferon gamma (IFNγ) are proinflammatory cytokines which play an important role in the development of T2DM. We undertook this study to analyze the gut microbiota of T2DM and nondiabetic subjects and to determine the profile of MCP 1 and IFNγ in the same subjects attending a tertiary care center in Chennai, Tamil Nadu, India. Methods: The study included 30 subjects with clinical details. Stool and blood samples were collected from all the subjects. DNA was extracted from fecal samples and polymerase chain reaction was done using fusion primers. Metagenomic analysis was performed using ion torrent sequencing. The reads obtained were in FASTA format and reported as operational taxonomic units. Human MCP 1 and IFNγ enzyme linked immunosorbent assay (ELISA) were performed for 23 serum samples. Results: The study consisted of 30 subjects; 17 were T2DM and 13 were nondiabetics. The gut microbiota among T2DM consisted predominantly of Gram negative bacteria; Escherichia and Prevotella, when compared with the nondiabetic group with predominantly Gram positive organisms suchas Faecalibacterium, Eubacterium, and Bifidobacterium. The mean MCP-1 values in the diabetic group were 232.8 pg/ml and in the nondiabetic group 170.84 pg/ml. IFNγ (mean 385.5 pg/ml) was raised in glycated hemoglobin (HbA1c) group of 6.5–7.5% which was statistically significant. Association of Escherichia with T2DM and association of Bifidobacteria in the nondiabetics were also statistically significant. Conclusion: Escherichia counts were elevated in T2DM with HbA1c of 6.5–8.5% which was statistically significant suggesting that lipopolysaccharides present in the cell wall of Gram-negative bacteria may be responsible for low-grade inflammation as evidenced by elevated MCP-1 and IFNγ levels in T

  16. Plasma Levels of Monocyte Chemoattractant Protein-1, n-Terminal Fragment of Brain Natriuretic Peptide and Calcidiol Are Independently Associated with the Complexity of Coronary Artery Disease

    PubMed Central

    Martín-Reyes, Roberto; Franco-Peláez, Juan Antonio; Lorenzo, Óscar; González-Casaus, María Luisa; Pello, Ana María; Aceña, Álvaro; Carda, Rocío; Martín-Ventura, José Luis; Blanco-Colio, Luis; Martín-Mariscal, María Luisa; Martínez-Milla, Juan; Villa-Bellosta, Ricardo; Piñero, Antonio; Navarro, Felipe; Egido, Jesús; Tuñón, José

    2016-01-01

    Background and Objectives We investigated the relationship of the Syntax Score (SS) and coronary artery calcification (CAC), with plasma levels of biomarkers related to cardiovascular damage and mineral metabolism, as there is sparse information in this field. Methods We studied 270 patients with coronary disease that had an acute coronary syndrome (ACS) six months before. Calcidiol, fibroblast growth factor-23, parathormone, phosphate and monocyte chemoattractant protein-1 [MCP-1], high-sensitivity C-reactive protein, galectin-3, and N-terminal pro-brain natriuretic peptide [NT-proBNP] levels, among other biomarkers, were determined. CAC was assessed by coronary angiogram as low-grade (0–1) and high-grade (2–3) calcification, measured with a semiquantitative scale ranging from 0 (none) to 3 (severe). For the SS study patients were divided in SS<14 and SS≥14. Multivariate linear and logistic regression analyses were performed. Results MCP-1 predicted independently the SS (RC = 1.73 [95%CI = 0.08–3.39]; p = 0.040), along with NT-proBNP (RC = 0.17 [95%CI = 0.05–0.28]; p = 0.004), male sex (RC = 4.15 [95%CI = 1.47–6.83]; p = 0.003), age (RC = 0.13 [95%CI = 0.02–0.24]; p = 0.020), hypertension (RC = 3.64, [95%CI = 0.77–6.50]; p = 0.013), hyperlipidemia (RC = 2.78, [95%CI = 0.28–5.29]; p = 0.030), and statins (RC = 6.12 [95%CI = 1.28–10.96]; p = 0.013). Low calcidiol predicted high-grade calcification independently (OR = 0.57 [95% CI = 0.36–0.90]; p = 0.013) along with ST-elevation myocardial infarction (OR = 0.38 [95%CI = 0.19–0.78]; p = 0.006), diabetes (OR = 2.35 [95%CI = 1.11–4.98]; p = 0.028) and age (OR = 1.37 [95%CI = 1.18–1.59]; p<0.001). During follow-up (1.79 [0.94–2.86] years), 27 patients developed ACS, stroke, or transient ischemic attack. A combined score using SS and CAC predicted independently the development of the outcome. Conclusions MCP-1 and NT-proBNP are independent predictors of SS, while low calcidiol plasma levels

  17. Nicotinic α7 receptor inhibits the acylation stimulating protein-induced production of monocyte chemoattractant protein-1 and keratinocyte-derived chemokine in adipocytes by modulating the p38 kinase and nuclear factor-κB signaling pathways

    PubMed Central

    Jiao, Zhou-Yang; Wu, Jing; Liu, Chao; Wen, Bing; Zhao, Wen-Zeng; Du, Xin-Ling

    2016-01-01

    Obesity is associated with chronic low-grade inflammation, which is characterized by increased infiltration of macrophages into adipose tissue. Acylation stimulating protein (ASP) is an adipokine derived from the immune complement system, which constitutes a link between adipocytes and macrophages, and is involved in energy homeostasis and inflammation. The purpose of the present study was to preliminarily investigate in vitro, whether functional α7nAChR in adipocytes may suppress ASP-induced inflammation and determine the possible signaling mechanism. Studies have reported associations between the expression of α7 nicotinic acetylcholine receptor (α7nAChR) and obesity, insulin resistance and diabetes. Additionally, α7nAChRs are important peripheral mediators of chronic inflammation, which is a key contributor to health problems in obesity. The primary aim of the present study was to evaluate the impact of exogenous ASP and α7nAChR on macrophage infiltration in adipose tissue and to examine the potential underlying molecular mechanism. Western blot analysis revealed that recombinant ASP increased the expression levels of monocyte chemoattractant protein-1 (MCP-1) and keratinocyte-derived chemokine (KC) by 3T3-L1 adipocytes. However, nicotine significantly inhibited the production of ASP-induced cytokines via the stimulation of α7nAChR. It was also found that α7nAChR inhibited the ASP-induced activation of p38 kinase and nuclear factor-κB (NF-κB), and the production of MCP-1 and KC. These data indicated that α7nAChR caused the inhibition of ASP-induced activation of p38 kinase and NF-κB to inhibit the production of MCP-1 and KC. PMID:27572255

  18. Relationship between vitamin D and IL-23, IL-17 and macrophage chemoattractant protein-1 as markers of fibrosis in hepatitis C virus Egyptians

    PubMed Central

    El Husseiny, Noha M; Fahmy, Hala M; Mohamed, Waleed A; Amin, Hisham H

    2012-01-01

    AIM: To assess vitamin D in hepatitis C patients and its relationship to interleukin (IL)-23, IL-17, and macrophage chemoattractant protein-1 (MCP-1). METHODS: The study was conducted on 50 Egyptian hepatitis C virus (HCV) genotype number IV-infected patients and 25 age- and gender-matched healthy subjects. Venous blood samples were obtained. Samples were allowed to clot and sera were separated by centrifugation and stored at -20 °C. A 25 hydroxy vitamin D assay was carried out using solid phase RIA. A 1,25 dihydroxy vitamin D assay was carried out using a commercial kit purchased from Incstar Corporation. IL-17 and -23 and MCP-1 were assayed by an enzyme immunoassay. Quantitative and qualitative polymerase chain reaction for HCV virus were done by TaqMan technology. Only HCV genotype IV-infected subjects were included in the study. The mean ± SD were determined, a t-test for comparison of means of different parameters was used. Correlation analysis was done using Pearson’s correlation. Differences among different groups were determined using the Kruskal-Wallis test. RESULTS: The mean vitamin D level in HCV patients (group I) was 15 ± 5.2 ng/mL while in control (group II) was 39.7 ± 10.8. For active vitamin D in group I as 16.6 ± 4.8 ng/mL while in group II was 41.9 ± 7.9. IL-23 was 154 ± 97.8 in group I and 6.7 ± 2.17 in group II. IL-17 was 70.7 ± 72.5 in cases and 1.2 ± 0.4 in control. MCP-1 was 1582 ± 794.4 in group I and 216.1 ± 5.38 in group II. Vitamin D deficiency affected 72% of HCV-infected patients and 0% of the control group. Vitamin D insufficiency existed in 28% of HCV-infected patients and 12% of the control group. One hundred percent of the cirrhotic patients and 40% of non cirrhotic HCV-infected patients had vitamin D deficiency. IL-23, IL-17, and MCP-1 were markedly increased in HCV-infected patients in comparison to controls.A significant negative correlation between vitamin D and IL-17 and -23 and MCP-1 was detected

  19. Hydrogen sulfide suppresses oxidized low-density lipoprotein (ox-LDL)-stimulated monocyte chemoattractant protein 1 generation from macrophages via the nuclear factor κB (NF-κB) pathway.

    PubMed

    Du, Junbao; Huang, Yaqian; Yan, Hui; Zhang, Qiaoli; Zhao, Manman; Zhu, Mingzhu; Liu, Jia; Chen, Stella X; Bu, Dingfang; Tang, Chaoshu; Jin, Hongfang

    2014-04-01

    This study was designed to examine the role of hydrogen sulfide (H2S) in the generation of oxidized low-density lipoprotein (ox-LDL)-stimulated monocyte chemoattractant protein 1 (MCP-1) from macrophages and possible mechanisms. THP-1 cells and RAW macrophages were pretreated with sodium hydrosulfide (NaHS) and hexyl acrylate and then treated with ox-LDL. The results showed that ox-LDL treatment down-regulated the H2S/cystathionine-β-synthase pathway, with increased MCP-1 protein and mRNA expression in both THP-1 cells and RAW macrophages. Hexyl acrylate promoted ox-LDL-induced inflammation, whereas the H2S donor NaHS inhibited it. NaHS markedly suppressed NF-κB p65 phosphorylation, nuclear translocation, DNA binding activity, and recruitment to the MCP-1 promoter in ox-LDL-treated macrophages. Furthermore, NaHS decreased the ratio of free thiol groups in p65, whereas the thiol reductant DTT reversed the inhibiting effect of H2S on the p65 DNA binding activity. Most importantly, site-specific mutation of cysteine 38 to serine in p65 abolished the effect of H2S on the sulfhydration of NF-κB and ox-LDL-induced NF-κB activation. These results suggested that endogenous H2S inhibited ox-LDL-induced macrophage inflammation by suppressing NF-κB p65 phosphorylation, nuclear translocation, DNA binding activity, and recruitment to the MCP-1 promoter. The sulfhydration of free thiol group on cysteine 38 in p65 served as a molecular mechanism by which H2S inhibited NF-κB pathway activation in ox-LDL-induced macrophage inflammation.

  20. Hydrogen Sulfide Suppresses Oxidized Low-density Lipoprotein (Ox-LDL)-stimulated Monocyte Chemoattractant Protein 1 generation from Macrophages via the Nuclear Factor κB (NF-κB) Pathway*

    PubMed Central

    Du, Junbao; Huang, Yaqian; Yan, Hui; Zhang, Qiaoli; Zhao, Manman; Zhu, Mingzhu; Liu, Jia; Chen, Stella X.; Bu, Dingfang; Tang, Chaoshu; Jin, Hongfang

    2014-01-01

    This study was designed to examine the role of hydrogen sulfide (H2S) in the generation of oxidized low-density lipoprotein (ox-LDL)-stimulated monocyte chemoattractant protein 1 (MCP-1) from macrophages and possible mechanisms. THP-1 cells and RAW macrophages were pretreated with sodium hydrosulfide (NaHS) and hexyl acrylate and then treated with ox-LDL. The results showed that ox-LDL treatment down-regulated the H2S/cystathionine-β-synthase pathway, with increased MCP-1 protein and mRNA expression in both THP-1 cells and RAW macrophages. Hexyl acrylate promoted ox-LDL-induced inflammation, whereas the H2S donor NaHS inhibited it. NaHS markedly suppressed NF-κB p65 phosphorylation, nuclear translocation, DNA binding activity, and recruitment to the MCP-1 promoter in ox-LDL-treated macrophages. Furthermore, NaHS decreased the ratio of free thiol groups in p65, whereas the thiol reductant DTT reversed the inhibiting effect of H2S on the p65 DNA binding activity. Most importantly, site-specific mutation of cysteine 38 to serine in p65 abolished the effect of H2S on the sulfhydration of NF-κB and ox-LDL-induced NF-κB activation. These results suggested that endogenous H2S inhibited ox-LDL-induced macrophage inflammation by suppressing NF-κB p65 phosphorylation, nuclear translocation, DNA binding activity, and recruitment to the MCP-1 promoter. The sulfhydration of free thiol group on cysteine 38 in p65 served as a molecular mechanism by which H2S inhibited NF-κB pathway activation in ox-LDL-induced macrophage inflammation. PMID:24550391

  1. Expression of T lymphocyte chemoattractants and activation markers in vernal keratoconjunctivitis

    PubMed Central

    El-Asrar, A M Abu; Struyf, S; Al-Kharashi, S A; Missotten, L; Van Damme, J; Geboes, K

    2002-01-01

    Background/aims: T lymphocytes are present in increased numbers in the conjunctiva of patients with vernal keratoconjunctivitis (VKC) and their activation has a central role in the pathogenesis of the chronic allergic inflammatory reactions seen in VKC. The aims of this study were to examine the expression of three recently described potent T lymphocyte chemoattractants, PARC (pulmonary and activation regulated chemokine), macrophage derived chemokine (MDC), and I-309, the MDC receptor CCR4, and T lymphocyte activation markers, CD25, CD26, CD62L, CD71, and CD30, and to correlate them with the counts of CD3+ T lymphocytes in the conjunctiva of patients with VKC. Method: Conjunctival biopsy specimens from 11 patients with active VKC, and eight control subjects were studied by immunohistochemical techniques using a panel of monoclonal and polyclonal antibodies directed against PARC, MDC, I-309, CCR4, CD25, CD26, CD62L, CD71, and CD30. The numbers of positively stained cells were counted. The phenotype of inflammatory cells expressing chemokines was examined by double immunohistochemistry. Results: In the normal conjunctiva, vascular endothelial cells in the upper substantia propria showed weak immunoreactivity for CD26. There was no immunoreactivity for the other antibodies. VKC specimens showed inflammatory cells expressing PARC, MDC, and I-309. The numbers of PARC+ inflammatory cells were higher than the numbers of MDC+ and I-309+ inflammatory cells and the mean values of the three groups differed significantly (17.0 (SD 10.1); 9.5 (9.9), and 4.3 (7.9), respectively, p = 0.0117, ANOVA). The numbers of PARC+ inflammatory cells had the strongest correlation with the numbers of CD3+ T lymphocytes. Few CCR4+ inflammatory cells were observed in only three specimens. Double immunohistochemistry revealed that all inflammatory cells expressing chemokines were CD68+ monocytes/macrophages. The numbers of CD25+ T lymphocytes were higher than the numbers of CD26+, CD62L+, CD71

  2. Role of the monocyte chemoattractant protein-1/C-C chemokine receptor 2 signaling pathway in transient receptor potential vanilloid type 1 ablation-induced renal injury in salt-sensitive hypertension

    PubMed Central

    Zhu, Mingjun; Xu, Hui; Cui, Lin; Liu, Weihong; Wang, Xiaoxiao; Shen, Si; Wang, Donna H

    2015-01-01

    Our recent studies indicate that the transient receptor potential vanilloid type 1 (TRPV1) channel may act as a potential regulator of monocyte/macrophage recruitment to reduce renal injury in salt-sensitive hypertension. This study tests the hypothesis that deletion of TRPV1 exaggerates salt-sensitive hypertension-induced renal injury due to enhanced inflammatory responses via monocyte chemoattractant protein-1 (MCP-1)/C-C chemokine receptor 2 (CCR2)-dependent pathways. Wild type (WT) and TRPV1-null mutant (TRPV1−/−) mice were subjected to uninephrectomy and deoxycorticosterone acetate (DOCA)-salt treatment for four weeks with or without the selective CCR2 antagonist, RS504393. DOCA-salt treatment increased systolic blood pressure (SBP) to the same degree in both strains, but increased urinary excretion of albumin and 8-isoprostane and decreased creatinine clearance with greater magnitude in TRPV1−/− mice compared to WT mice. DOCA-salt treatment also caused renal glomerulosclerosis, tubulointerstitial injury, collagen deposition, monocyte/macrophage infiltration, proinflammatory cytokine and chemokine production, and NF-κB activation in greater degree in TRPV1−/− mice compared to WT mice. Blockade of the CCR2 with RS504393 (4 mg/kg/day) had no effect on SBP in DOCA-salt-treated WT or TRPV1−/− mice compared to their respective controls. However, treatment with RS504393 ameliorated renal dysfunction and morphological damage, and prevented the increase in monocyte/macrophage infiltration, cytokine/chemokine production, and NF-κB activity in both DOCA-salt hypertensive strains with a greater effect in DOCA-salt-treated TRPV1−/− mice compared to DOCA-salt-treated WT mice. No differences in CCR2 protein expression in kidney were found between DOCA-salt-treated WT and TRPV1−/− mice with or without RS504393 treatment. Our studies for the first time indicate that deletion of TRPV1 aggravated renal injury in salt-sensitive hypertension via

  3. High-fat diet enhances and monocyte chemoattractant protein-1 deficiency reduces bone loss in mice with pulmonary metastases of Lewis lung carcinoma

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Bone is adversely affected by metastasis and metastasis-associated complications. Obesity is a risk factor for both bone and cancer. Adipose tissue is an endocrine organ that produces pro-inflammatory adipokines, such as monocyte chemotactic protein-1 (MCP-1), that contribute to obesity and obesit...

  4. Differential regulation of dentin matrix protein 1 expression during odontogenesis.

    PubMed

    Lu, Yongbo; Zhang, Shubin; Xie, Yixia; Pi, Yuli; Feng, Jian Q

    2005-01-01

    Dentin matrix protein 1 (DMP1) is highly expressed in mineralized tooth and bone. Both in vitro and in vivo data show that DMP1 is critical for mineralization and tooth morphogenesis (growth and development). In this study, we studied Dmp1 gene regulation. The in vitro transient transfection assay identified two important DNA fragments, the 2.4- and 9.6-kb promoter regions. We next generated and analyzed transgenic mice bearing the beta-galactosidase (lacZ) reporter gene driven by the 2.4- or 9.6-kb promoter with the complete 4-kb intron 1. The 9.6-kb Dmp1-lacZ mice conferred a DMP1 expression pattern in odontoblasts identical to that in the endogenous Dmp1 gene. This is reflected by lacZ expression in Dmp1-lacZ knock-in mice during all stages of odontogenesis. In contrast, the 2.4-kb Dmp1-lacZ mice display activity in odontoblast cells only at the early stage of odontogenesis. Thus, we propose that different transcription factors regulate early or later cis-regulatory domains of the Dmp1 promoter, which gives rise to the unique spatial and temporal expression pattern of Dmp1 gene at different stages of tooth development.

  5. Cloning and expression of a gene encoding a protein obtained from earthworm secretion that is a chemoattractant for garter snakes.

    PubMed

    Liu, W; Wang, D; Chen, P; Halpern, M

    1997-10-24

    The protein ES20, derived from earthworm shock secretion, is a vomeronasally mediated chemoattractant for garter snakes (Jiang, X. C., Inouchi, J., Wang, D., and Halpern, M. (1990) J. Biol. Chem. 265, 8736-8744). Based on its 15-residue N-terminal amino acid sequence, degenerative oligodeoxynucleotide probes were synthesized and used to screen a cDNA library that was constructed in sense orientation using a Uni-ZAPTM XR vector and XL1-Blue MRF' host. A gene was cloned from a polymerase chain reaction as well as from the cDNA library. A combination of the forward degenerative primer and T7 primer was used to obtain gene-specific DNA fragments, from which probes were synthesized and successfully used in screening the cDNA library. The ES20 gene is about 700 base pairs long and encodes 208 amino residues. The ES20 gene was excised from a recombinant plasmid pSK-ES20, ligated to pQE30 expression vector, and transformed into Escherichia coli strain JM109. The selected recombinant plasmids were transformed into expression host cell, E. coli M15[pREP4]. Three transformants were selected, induced with isopropyl-1-thio-beta-D-galactopyranoside for fusion gene expression and an expressed 20-kDa fusion protein purified under denaturing conditions. This protein was refolded and gave a positive reaction against ES20-specific polyclonal antibodies. The fusion protein that had not been denatured remained as an aggregate and was an active chemoattractant for garter snakes. PMID:9341189

  6. L-cystathionine inhibits oxidized low density lipoprotein-induced THP-1-derived macrophage inflammatory cytokine monocyte chemoattractant protein-1 generation via the NF-κB pathway.

    PubMed

    Zhu, Mingzhu; Du, Junbao; Liu, Angie Dong; Holmberg, Lukas; Chen, Selena Y; Bu, Dingfang; Tang, Chaoshu; Jin, Hongfang

    2015-05-28

    This study aimed to explore whether and how L-cystathionine had any regulatory effect on the inflammatory response in THP-1-derived macrophages cultured in vitro under oxidized low-density lipoprotein (ox-LDL) stimulation. The human monocyte line THP-1 cell was cultured in vitro and differentiated into macrophages after 24 hours of PMA induction. Macrophages were pretreated with L-cystathionine and then treated with ox-LDL. The results showed that compared with the controls, ox-LDL stimulation significantly upregulated the expression of THP-1-derived macrophage MCP-1 by enhancing NF-κB p65 phosphorylation, nuclear translocation and DNA binding with the MCP-1 promoter. Compared with the ox-LDL group, 0.3 mmol/L and 1.0 mmol/L L-cystathionine significantly inhibited the expression of THP-1-derived macrophage MCP-1. Mechanistically, 0.3 mmol/L and 1.0 mmol/L L-cystathionine suppressed phosphorylation and nuclear translocation of the NF-κB p65 protein, as well as the DNA binding activity and DNA binding level of NF-κB with the MCP-1 promoter, which resulted in a reduced THP-1-derived macrophage MCP-1 generation. This study suggests that L-cystathionine could inhibit the expression of MCP-1 in THP-1-derived macrophages induced by ox-LDL via inhibition of NF-κB p65 phosphorylation, nuclear translocation, and binding of the MCP-1 promoter sequence after entry into the nucleus.

  7. Yersinia enterocolitica Invasin Protein Triggers Differential Production of Interleukin-1, Interleukin-8, Monocyte Chemoattractant Protein 1, Granulocyte-Macrophage Colony-Stimulating Factor, and Tumor Necrosis Factor Alpha in Epithelial Cells: Implications for Understanding the Early Cytokine Network in Yersinia Infections

    PubMed Central

    Kampik, Daniel; Schulte, Ralf; Autenrieth, Ingo B.

    2000-01-01

    Yersinia enterocolitica infection of epithelial cells results in interleukin-8 (IL-8) mRNA expression. Herein we demonstrate that besides IL-8, increased mRNA levels of five other cytokines, IL-1α, IL-1β, monocyte chemoattractant protein 1 (MCP-1), granulocyte-macrophage colony-stimulating factor (GM-CSF), and tumor necrosis factor alpha (TNF-α), can be detected upon infection of HeLa cells with Yersinia. Yersinia-triggered cytokine production was not affected by blocking phosphatidylinositol-3-phosphate kinase with wortmannin, which inhibited bacterial invasion. Comparable cytokine mRNA responses were triggered by Escherichia coli expressing Yersinia inv, while no response was triggered by an inv-deficient Yersinia mutant. Moreover, cytokine responses were independent from metabolic activity of the bacteria, as killed bacterial cells were sufficient for triggering cytokine responses in HeLa cells. Semiquantitative reverse transcription-PCR analysis was used to assess the kinetics of cytokine mRNA expression in infected HeLa cells. IL-8, IL-1α, IL-1β, MCP-1, GM-CSF, and TNF-α mRNA expression increased within 1 h postinfection, reached a maximum after 3 to 4 h, and then declined to preinfection levels within 3 h. IL-8, MCP-1, and GM-CSF were secreted by HeLa cells, whereas IL-1α and IL-1β were not secreted and thus were found exclusively intracellularly. TNF-α protein could not be detected in cell lysates or supernatants. Stimulation of HeLa cells with IL-1α was followed by increased IL-8 mRNA expression, whereas stimulation with IL-8 did not induce cytokine production. Likewise, MCP-1 and GM-CSF did not induce significant cytokine responses in HeLa cells. Our results implicate that the initial host response to Yersinia infection might be sustained by IL-8, MCP-1, and GM-CSF produced by epithelial cells. PMID:10768935

  8. Increased expression of monocyte chemotactic protein-1 during active hepatic fibrogenesis: correlation with monocyte infiltration.

    PubMed Central

    Marra, F.; DeFranco, R.; Grappone, C.; Milani, S.; Pastacaldi, S.; Pinzani, M.; Romanelli, R. G.; Laffi, G.; Gentilini, P.

    1998-01-01

    Monocyte chemotactic protein (MCP)-1 is a chemoattractant and activator for circulating monocytes and T lymphocytes. We investigated MCP-1 protein and gene expression during chronic liver disease at different stages, using immunohistochemistry and in situ hybridization, respectively. In normal liver, a modest expression of MCP-1 was confined to few peri-sinusoidal cells and to bile duct epithelial cells. During chronic hepatitis, MCP-1 immunostaining and gene expression were evident in the inflammatory infiltrate of the portal tract. In tissue from patients with active cirrhosis, MCP-1 expression was clearly up-regulated and was present in the portal tract, in the epithelial cells of regenerating bile ducts, and in the active septa surrounding regenerating nodules. A combination of in situ hybridization for MCP-1 and immunohistochemistry showed that activated stellate cells and monocyte/macrophages contribute to MCP-1 expression in vivo together with bile duct epithelial cells. Comparison of serial sections of liver biopsies from patients with various degrees of necro-inflammatory activity showed that infiltration of the portal tracts with monocytes/macrophages is directly correlated with the expression of MCP-1. These data expand previous in vitro studies showing that secretion of MCP-1 may contribute to the formation and maintenance of the inflammatory infiltrate observed during chronic liver disease. Images Figure 1 Figure 2 Figure 3 Figure 4 PMID:9466568

  9. Human Cementum Protein 1 induces expression of bone and cementum proteins by human gingival fibroblasts

    SciTech Connect

    Carmona-Rodriguez, Bruno; Alvarez-Perez, Marco Antonio; Narayanan, A. Sampath; Zeichner-David, Margarita; Reyes-Gasga, Jose; Molina-Guarneros, Juan; Garcia-Hernandez, Ana Lilia; Suarez-Franco, Jose Luis; Chavarria, Ivet Gil; Villarreal-Ramirez, Eduardo; Arzate, Higinio . E-mail: harzate@servidor.unam.mx

    2007-07-06

    We recently presented evidence showing that a human cementoblastoma-derived protein, named Cementum Protein 1 (CEMP1) may play a role as a local regulator of cementoblast differentiation and cementum-matrix mineralization. This protein was shown to be expressed by cementoblasts and progenitor cells localized in the periodontal ligament. In this study we demonstrate that transfection of CEMP1 into human gingival fibroblasts (HGF) induces mineralization and expression of bone and cementum-matrix proteins. The transfected HGF cells had higher alkaline phosphatase activity and proliferation rate and they expressed genes for alkaline phosphatase, bone sialoprotein, osteocalcin, osteopontin, the transcription factor Runx2/Cbfa1, and cementum attachment protein (CAP). They also produced biological-type hydroxyapatite. These findings indicate that the CEMP1 might participate in differentiation and mineralization of nonosteogenic cells, and that it might have a potential function in cementum and bone formation.

  10. Regulation of pathogenesis-related protein-1a gene expression in tobacco.

    PubMed

    Uknes, S; Dincher, S; Friedrich, L; Negrotto, D; Williams, S; Thompson-Taylor, H; Potter, S; Ward, E; Ryals, J

    1993-02-01

    Pathogenesis-related protein-1a (PR-1a) is a protein of unknown function that is strongly induced during the onset of systemic acquired resistance (SAR) in tobacco. The expression of PR-1a is under complex regulation that is controlled at least partially by the rate of transcription. In this study, we demonstrated that 661 bp of 5' flanking DNA was sufficient to impart tobacco mosaic virus and salicylic acid inducibility to a reporter gene. The PR-1a promoter did not respond significantly to treatments with either auxin or cytokinin. Experiments with the protein synthesis inhibitor cycloheximide indicated that protein synthesis is required for salicylate-dependent mRNA accumulation. At flowering, the PR-1a gene was expressed primarily in the mesophyll and epidermal tissues of the leaf blade and the sepals of the flower. Several artifacts, most importantly ectopic expression in pollen, were associated with the use of the beta-glucuronidase reporter gene.

  11. Organ-selective regulation of vascular adhesion protein-1 expression in man.

    PubMed

    Arvilommi, A M; Salmi, M; Jalkanen, S

    1997-07-01

    Vascular adhesion protein-1 (VAP-1) is an endothelial molecule which mediates lymphocyte binding to endothelium in peripheral lymph nodes and at certain sites of inflammation. The expression of VAP-1 in vivo is strongly up-regulated in inflamed tissues, such as gut and skin. The purpose of this work was to examine the factors responsible for this induction of VAP-1. Since the expression of VAP-1 could not be induced in cultured endothelial cells with a large panel of mediators, we used an organ culture technique for the investigation of the regulation of VAP-1 expression in a more physiological micromilieu. Indeed, we found that the expression of endothelial VAP-1 could be up-regulated in human tonsillar tissue with interleukin (IL)-1, IL-4, tumor necrosis factor (TNF-alpha), interferon (IFN)-gamma and lipopolysaccharide, whereas histamine, thrombin, dibutyryl cAMP, N-formyl-Met-Leu-Phe (fMLP) and phorbol 12-myristate 13-acetate (PMA) had no effect. The induced VAP-1 protein was similar in molecular weight to the non-induced VAP-1, suggesting that VAP-1 synthesized de novo carries appropriate carbohydrate moieties. In contrast to tonsil organ culture, similar inductions performed with human appendix showed no up-regulation of VAP-1 expression, indicating that the regulation of VAP-1 expression exhibits organ-selective characteristics. Furthermore, in these tissues the smooth muscle cells, which constitutively express VAP-1, could not be stimulated to alter their level of expression of this molecule. In conclusion, the expression of VAP-1 can be markedly up-regulated with several mediators in tonsil but not in appendix organ culture, whereas cultured endothelial cells cannot be induced to express VAP-1. These results indicate that the expression of VAP-1 is regulated in a tissue- and cell type-selective manner, and a correct micromilieu is required for the up-regulation to occur. PMID:9247594

  12. Regulatory Effects of Macrophage Inflammatory Protein 1α/CCL3 on the Development of Immunity to Cryptococcus neoformans Depend on Expression of Early Inflammatory Cytokines

    PubMed Central

    Olszewski, Michal A.; Huffnagle, Gary B.; Traynor, Timothy R.; McDonald, Roderick A.; Cook, Donald N.; Toews, Galen B.

    2001-01-01

    Macrophage inflammatory protein 1α (MIP-1α)/CCL3 prevents the development of eosinophilic pneumonia (EP) driven by a nonprotective T2-type immunity during infection with a highly virulent strain of Cryptococcus neoformans. The present study evaluated the interaction of MIP-1α with other innate immune system cytokines by comparing the immune responses that followed pulmonary infections with high- (C. neoformans 145A) and low (C. neoformans 52D)-virulence strains. In contrast to what was found for C. neoformans 145A infection, lack of MIP-1α in C. neoformans 52D infection did not cause the development of EP. C. neoformans 52D induced tumor necrosis factor alpha (TNF-α), gamma interferon (IFN-γ), and MCP-1 in the lungs of infected wild-type (WT) and MIP-1α knockout (KO) mice by day 7 postinfection. Both WT and MIP-1α KO mice subsequently cleared this infection. Thus, the robust expression of early inflammatory cytokines in C. neoformans 52D-infected mice promoted the development of protective immunity even in the absence of MIP-1α. Alternatively, C. neoformans 145A-infected WT and MIP-1α KO mice had diminished TNF-α, IFN-γ, and macrophage chemoattractant protein 1 (MCP-1) responses, indicating that virulent C. neoformans 145A evaded early innate host defenses. However C. neoformans 145A-infected WT mice had an early induction of MIP-1α and subsequently did not develop EP. In contrast, C. neoformans 145A-infected MIP-1α KO mice developed EP and had increased C. neoformans dissemination into the brain by day 35. We conclude that, in the absence of other innate immune response effector molecules, MIP-1α is crucial to prevent the development of EP and to control C. neoformans dissemination to the brain. PMID:11553568

  13. A mitochondrial uncoupling artifact can be caused by expression of uncoupling protein 1 in yeast.

    PubMed Central

    Stuart, J A; Harper, J A; Brindle, K M; Jekabsons, M B; Brand, M D

    2001-01-01

    Uncoupling protein 1 (UCP1) from mouse was expressed in yeast and the specific (GDP-inhibitable) and artifactual (GDP-insensitive) effects on mitochondrial uncoupling were assessed. UCP1 provides a GDP-inhibitable model system to help interpret the uncoupling effects of high expression in yeast of other members of the mitochondrial carrier protein family, such as the UCP1 homologues UCP2 and UCP3. Yeast expressing UCP1 at modest levels (approx. 1 microg/mg of mitochondrial protein) showed no growth defect, normal rates of chemically uncoupled respiration and an increased non-phosphorylating proton conductance that was completely GDP-sensitive. The catalytic-centre activity of UCP1 in these yeast mitochondria was similar to that in mammalian brown-adipose-tissue mitochondria. However, yeast expressing UCP1 at higher levels (approx. 11 microg/mg of mitochondrial protein) showed a growth defect. Their mitochondria had depressed chemically uncoupled respiration rates and an increased proton conductance that was partly GDP-insensitive. Thus, although UCP1 shows native behaviour at modest levels of expression in yeast, higher levels (or rates) of expression can lead to an uncoupling that is not a physiological property of the native protein and is therefore artifactual. This observation might be important in the interpretation of results from experiments in which the functions of UCP1 homologues are verified by their ability to uncouple yeast mitochondria. PMID:11389685

  14. Yes-associated protein 1 is widely expressed in human brain tumors and promotes glioblastoma growth.

    PubMed

    Orr, Brent A; Bai, Haibo; Odia, Yazmin; Jain, Deepali; Anders, Robert A; Eberhart, Charles G

    2011-07-01

    The hippo pathway and its downstream mediator yes-associated protein 1 (YAP1) regulate mammalian organ size in part through modulating progenitor cell numbers. YAP1 has also been implicated as an oncogene in multiple human cancers. Currently, little is known about the expression of YAP1 either in normal human brain tissue or in central nervous system neoplasms. We used immunohistochemistry to evaluate nuclear YAP1 expression in the fetal and normal adult human brains and in 264 brain tumors. YAP1 was expressed in fetal and adult brain regions known to harbor neural progenitor cells, but there was little YAP1 immunoreactivity in the adult cerebral cortex. YAP1 protein was also readily detected in the nuclei of human brain tumors. In medulloblastoma, the expression varied between histologic subtypes and was most prominent in nodular/desmoplastic tumors. In gliomas, it was frequently expressed in infiltrating astrocytomas and oligodendrogliomas but rarely in pilocytic astrocytomas. Using a loss-of-function approach, we show that YAP1 promoted growth of glioblastoma cell lines in vitro. High levels of YAP1 messenger RNA expression were associated with aggressive molecular subsets of glioblastoma and with a nonsignificant trend toward reduced mean survival in human astrocytoma patients. These findings suggest that YAP1 may play an important role in normal human brain development and that it could represent a new target in human brain tumors.

  15. X-ray Structure of Engineered Human Aortic Preferentially Expressed Protein-1 (APEG-1)

    SciTech Connect

    Manjasetty,B.; Niesen, F.; Scheich, C.; Roske, Y.; Goetz, F.; Behlke, J.; Sievert, V.; Heinemann, U.; Buessow, K.

    2005-01-01

    Arterial smooth muscle cells (SMC) are essential for the formation and function of the cardiovascular system. Abnormalities in their growth can cause a wide range of human disorders such as atherosclerosis, the principal cause for heart failure, thus the leading cause for deaths in the western world. The molecular mechanisms that regulate SMC growth and differentiation are unclear partly due to the lack of specific markers and defined in vitro differentiation systems. The recently discovered Aortic Preferentially Expressed Protein-1 (APEG-1) may serve as a sensitive marker for vascular SMC differentiation. APEG-1 is expressed in differentiated vascular SMC in vivo and was found to be down-regulated rapidly in de-differentiated vascular SMC in vitro and in injured arteries in vivo.

  16. The Expression Level and Prognostic Value of Y-Box Binding Protein-1 in Rectal Cancer

    PubMed Central

    Zhang, Yu; Zhao, Ping-Wu; Feng, Gang; Xie, Gang; Wang, An-Qun; Yang, Yong-Hong; Wang, Dong; Du, Xiao-Bo

    2015-01-01

    The aims of this study were to simultaneously evaluate the expression of Y-box binding protein-1 (YB-1) in non-neoplastic rectal tissue and rectal cancer tissue, and to collect clinical follow-up data for individual patients. Additionally, we aimed to investigate the developmental functions and prognostic value of YB-1 in rectal cancer. We performed immunohistochemical studies to examine YB-1 expression in tissue samples from 80 patients with rectal cancer, 30 patients with rectal tubular adenoma, and 30 patients with rectitis. The mean YB-1 histological scores for rectal cancer, rectal tubular adenoma, and rectitis tissue specimens were 205.5, 164.3, and 137.7, respectively. Shorter disease-free and overall survival times were found in patients with rectal cancer who had higher YB-1 expression than in those with lower expression (38.2 months vs. 52.4 months, P = 0.013; and 44.4 months vs. 57.3 months, P = 0.008, respectively). Our results indicate that YB-1 expression is higher in rectal cancer tissue than in rectal tubular adenoma and rectitis tissue and that it may be an independent prognostic factor for rectal cancer. PMID:25790262

  17. SIRT1 Suppresses Activator Protein-1 Transcriptional Activity and Cyclooxygenase-2 Expression in Macrophages*

    PubMed Central

    Zhang, Ran; Chen, Hou-Zao; Liu, Jin-Jing; Jia, Yu-Yan; Zhang, Zhu-Qin; Yang, Rui-Feng; Zhang, Yuan; Xu, Jing; Wei, Yu-Sheng; Liu, De-Pei; Liang, Chih-Chuan

    2010-01-01

    SIRT1 (Sirtuin type 1), a mammalian orthologue of yeast SIR2 (silent information regulator 2), has been shown to mediate a variety of calorie restriction (CR)-induced physiological events, such as cell fate regulation via deacetylation of the substrate proteins. However, whether SIRT1 deacetylates activator protein-1 (AP-1) to influence its transcriptional activity and target gene expression is still unknown. Here we demonstrate that SIRT1 directly interacts with the basic leucine zipper domains of c-Fos and c-Jun, the major components of AP-1, by which SIRT1 suppressed the transcriptional activity of AP-1. This process requires the deacetylase activity of SIRT1. Notably, SIRT1 reduced the expression of COX-2, a typical AP-1 target gene, and decreased prostaglandin E2 (PGE2) production of peritoneal macrophages (pMΦs). pMΦs with SIRT1 overexpression displayed improved phagocytosis and tumoricidal functions, which are associated with depressed PGE2. Furthermore, SIRT1 protein level was up-regulated in CR mouse pMΦs, whereas elevated SIRT1 decreased COX-2 expression and improved PGE2-related macrophage functions that were reversed following inhibition of SIRT1 deacetylase activity. Thus, our results indicate that SIRT1 may be a mediator of CR-induced macrophage regulation, and its deacetylase activity contributes to the inhibition of AP-1 transcriptional activity and COX-2 expression leading to amelioration of macrophage function. PMID:20042607

  18. High expression of cellular retinol binding protein-1 in lung adenocarcinoma is associated with poor prognosis

    PubMed Central

    Doldo, Elena; Costanza, Gaetana; Ferlosio, Amedeo; Pompeo, Eugenio; Agostinelli, Sara; Bellezza, Guido; Mazzaglia, Donatella; Giunta, Alessandro; Sidoni, Angelo; Orlandi, Augusto

    2015-01-01

    Purpose Adenocarcinoma, the most common non-small cell lung cancer is a leading cause of death worldwide, with a low overall survival (OS) despite increasing attempts to achieve an early diagnosis and accomplish surgical and multimodality treatment strategies. Cellular retinol binding protein-1 (CRBP-1) regulates retinol bioavailability and cell differentiation, but its role in lung cancerogenesis remains uncertain. Experimental design CRBP-1 expression, clinical outcome and other prognostic factors were investigated in 167 lung adenocarcinoma patients. CRBP-1 expression was evaluated by immunohistochemistry of tissue microarray sections, gene copy number analysis and tumor methylation specific PCR. Effects of CRBP-1 expression on proliferation/apoptosis gene array, protein and transcripts were investigated in transfected A549 lung adenocarcinoma cells. Results CRBP-1High expression was observed in 62.3% of adenocarcinomas and correlated with increased tumor grade and reduced OS as an independent prognostic factor. CRBP-1 gene copy gain also associated with tumor CRBP-1High status and dedifferentiation. CRBP-1-transfected (CRBP-1+) A549 grew more than CRBP-1− A549 cells. At >1μM concentrations, all trans-retinoic acid and retinol reduced viability more in CRBP-1+ than in CRBP-1− A549 cells. CRBP-1+ A549 cells showed up-regulated RARα/ RXRα and proliferative and transcriptional genes including pAkt, pEGFR, pErk1/2, creb1 and c-jun, whereas RARβ and p53 were strongly down-regulated; pAkt/pErk/ pEGFR inhibitors counteracted proliferative advantage and increased RARα/RXRα, c-jun and CD44 expression in CRBP-1+ A549 cells. Conclusion CRBP-1High expression in lung adenocarcinoma correlated with increased tumor grade and reduced OS, likely through increased Akt/Erk/EGFR-mediated cell proliferation and differentiation. CRBP-1High expression can be considered an additional marker of poor prognosis in lung adenocarcinoma patients. PMID:26807202

  19. High-level expression and characterization of a glycosylated human cementum protein 1 with lectin activity.

    PubMed

    Romo-Arévalo, Enrique; Arzate, Higinio; Montoya-Ayala, Gonzalo; Rodríguez-Romero, Adela

    2016-01-01

    This work aims to contribute to the knowledge of human cementum protein 1 (CEMP1), its conformational characteristics and influence during the biomineralization process. The results revealed that hrCEMP1 expressed in Pichia pastoris is a 2.4% glycosylated, thermostable protein which possesses a molecular mass of 28,770 Da. The circular dichroism spectrum indicated a secondary structure content of 28.6% of alpha-helix, 9.9% of beta-sheet and 61.5% of random-coil forms. Biological activity assays demonstrated that hrCEMP1 nucleates and regulates hydroxyapatite crystal growth. Hereby, it is demonstrated for the first time that CEMP1 has a (C-type) lectin-like activity and specifically recognizes mannopyranoside. The information produced by this biochemical and structural characterization may contribute to understand more fully the biological functions of CEMP1.

  20. Expression of activator protein-1 (AP-1) family members in breast cancer

    PubMed Central

    2013-01-01

    Background The activator protein-1 (AP-1) transcription factor is believed to be important in tumorigenesis and altered AP-1 activity was associated with cell transformation. We aimed to assess the potential role of AP-1 family members as novel biomarkers in breast cancer. Methods We studied the expression of AP-1 members at the mRNA level in 72 primary breast tumors and 37 adjacent non-tumor tissues and evaluated its correlation with clinicopathological parameters including estrogen receptor (ER), progesterone receptor (PR) and HER2/neu status. Expression levels of Ubiquitin C (UBC) were used for normalization. Protein expression of AP-1 members was assessed using Western blot analysis in a subset of tumors. We used student’s t-test, one-way ANOVA, logistic regression and Pearson’s correlation coefficient for statistical analyses. Results We found significant differences in the expression of AP-1 family members between tumor and adjacent non-tumor tissues for all AP-1 family members except Fos B. Fra-1, Fra-2, Jun-B and Jun-D mRNA levels were significantly higher in tumors compared to adjacent non-tumor tissues (p < 0.001), whilst c-Fos and c-Jun mRNA levels were significantly lower in tumors compared with adjacent non-tumor tissues (p < 0.001). In addition, Jun-B overexpression had outstanding discrimination ability to differentiate tumor tissues from adjacent non-tumor tissues as determined by ROC curve analysis. Moreover, Fra-1 was significantly overexpressed in the tumors biochemically classified as ERα negative (p = 0.012) and PR negative (p = 0.037). Interestingly, Fra-1 expression was significantly higher in triple-negative tumors compared with luminal carcinomas (p = 0.01). Conclusions Expression levels of Fra-1 and Jun-B might be possible biomarkers for prognosis of breast cancer. PMID:24073962

  1. Leukocyte trafficking-associated vascular adhesion protein 1 is expressed and functionally active in atherosclerotic plaques

    PubMed Central

    Silvola, Johanna M. U.; Virtanen, Helena; Siitonen, Riikka; Hellberg, Sanna; Liljenbäck, Heidi; Metsälä, Olli; Ståhle, Mia; Saanijoki, Tiina; Käkelä, Meeri; Hakovirta, Harri; Ylä-Herttuala, Seppo; Saukko, Pekka; Jauhiainen, Matti; Veres, Tibor Z.; Jalkanen, Sirpa; Knuuti, Juhani; Saraste, Antti; Roivainen, Anne

    2016-01-01

    Given the important role of inflammation and the potential association of the leukocyte trafficking-associated adhesion molecule vascular adhesion protein 1 (VAP-1) with atherosclerosis, this study examined whether functional VAP-1 is expressed in atherosclerotic lesions and, if so, whether it could be targeted by positron emission tomography (PET). First, immunohistochemistry revealed that VAP-1 localized to endothelial cells of intra-plaque neovessels in human carotid endarterectomy samples from patients with recent ischemic symptoms. In low-density lipoprotein receptor-deficient mice expressing only apolipoprotein B100 (LDLR−/−ApoB100/100), VAP-1 was expressed on endothelial cells lining inflamed atherosclerotic lesions; normal vessel walls in aortas of C57BL/6N control mice were VAP-1-negative. Second, we discovered that the focal uptake of VAP-1 targeting sialic acid-binding immunoglobulin-like lectin 9 based PET tracer [68Ga]DOTA-Siglec-9 in atherosclerotic plaques was associated with the density of activated macrophages (r = 0.58, P = 0.022). As a final point, we found that the inhibition of VAP-1 activity with small molecule LJP1586 decreased the density of macrophages in inflamed atherosclerotic plaques in mice. Our results suggest for the first time VAP-1 as a potential imaging target for inflamed atherosclerotic plaques, and corroborate VAP-1 inhibition as a therapeutic approach in the treatment of atherosclerosis. PMID:27731409

  2. Identification, characterization, and expression of dentin matrix protein 1 gene in Xenopus laevis.

    PubMed

    Yonekura, Tomoko; Homma, Hiromi; Sakurai, Atsuo; Moriguchi, Mitsuko; Miake, Yasuo; Toyosawa, Satoru; Shintani, Seikou

    2013-12-01

    Dentin matrix protein 1 (DMP1) is an acidic extracellular matrix protein expressed mainly in bone and dentin, and is a member of the small integrin-binding ligand N-linked glycoprotein (SIBLING) family. The DMP1 gene, however, appears to evolve rapidly in comparison with other SIBLING genes, even though such functionally important molecules usually evolve more slowly than less important ones. The purpose of this study was to identify and characterize an ortholog of the DMP1 gene in an amphibian (Xenopus laevis; X. laevis) to clarify molecular evolutionary alterations in DMP1 associated with calcified tissues in tetrapods. Furthermore, we analyzed the mRNA expression of this gene to elucidate its functional change in bone and developing tooth germ in comparison with amniote DMP1s. The similarities of the deduced amino acid sequence of X. laevis DMP1 to that of the corresponding amniote proteins were low, although they did share several unique features specific to DMP1 and have similar properties. Expression of X. laevis DMP1 mRNA was predominant in osteocytes and odontoblasts, but only transiently observed in ameloblasts, as in amniotes. These results suggest that DMP1 has conserved several functions during tetrapod evolution. This indicates that continuity of biochemical properties has been more important in maintaining DMP1 functionality than that of the sequence of amino acid residues, which has undergone change over the course of molecular evolution.

  3. Expression of monocyte chemotactic protein-1 by monocytes and endothelial cells exposed to thrombin.

    PubMed Central

    Colotta, F.; Sciacca, F. L.; Sironi, M.; Luini, W.; Rabiet, M. J.; Mantovani, A.

    1994-01-01

    Thrombin, in addition to being a key enzyme in hemostasis, affects a series of endothelial and leukocyte functions and thus may be involved in the regulation of inflammatory reactions. Because leukocyte recruitment and activation are important events in inflammatory and thrombotic processes, in this study we have examined the possibility that thrombin induces the production of a cytokine chemotactic for mononuclear phagocytes. Human peripheral blood mononuclear cells (PBMC) exposed in vitro to thrombin expressed transcripts of monocyte chemotactic protein-1 (MCP-1; alternative acronyms: JE, monocyte chemotactic and activating factor, tumor-derived chemotactic factor). Thrombin was two- to threefold less effective than endotoxin in inducing MCP-1 transcripts in PBMC. Among circulating mononuclear cells, monocytes were identified as the cells expressing MCP-1 in response to thrombin. Monocytes expressed thrombin receptor transcripts. Boiling, hirudin, antithrombin III, and mutation of the catalytic site serine 205 into alanine) blocked the capacity of thrombin to induce MCP-1 expression. The thrombin receptor-activating peptide mimicked the effect of thrombin in inducing MCP-1 expression. Induction of MCP-1 transcript by thrombin was not reduced by blocking interleukin-1 and tumor necrosis factor, suggesting that these mediators are not involved in thrombin-induced expression of MCP-1. In addition to monocytes, endothelial cells (EC) also expressed MCP-1 in response to thrombin, although at lower levels compared with monocytes. Actinomycin D experiments indicated that induction of MCP-1 by thrombin in PBMC and EC was gene transcription dependent. The inhibition of protein synthesis blocked thrombin-induced MCP-1 expression in PBMC, whereas it superinduced both constitutive and thrombin-inducible expression of MCP-1 in EC, indicating different mechanisms of regulation of this gene in mononuclear phagocytes versus endothelial cells. Thrombin stimulated mononuclear

  4. Constitutive Nuclear Expression of Dentin Matrix Protein 1 Fails to Rescue the Dmp1-null Phenotype*

    PubMed Central

    Lin, Shuxian; Zhang, Qi; Cao, Zhengguo; Lu, Yongbo; Zhang, Hua; Yan, Kevin; Liu, Ying; McKee, Marc D.; Qin, Chunlin; Chen, Zhi; Feng, Jian Q.

    2014-01-01

    Dentin matrix protein 1 (DMP1) plays multiple roles in bone, tooth, phosphate homeostasis, kidney, salivary gland, reproductive cycles, and the development of cancer. In vitro studies have indicated two different biological mechanisms: 1) as a matrix protein, DMP1 interacts with αvβ3 integrin and activates MAP kinase signaling; and 2) DMP1 serves as a transcription co-factor. In vivo studies have demonstrated its key role in osteocytes. This study attempted to determine whether DMP1 functions as a transcription co-factor and regulates osteoblast functions. For gene expression comparisons using adenovirus constructs, we targeted the expression of DMP1 either to the nucleus only by replacing the endogenous signal peptide with a nuclear localization signal (NLS) sequence (referred to as NLSDMP1) or to the extracellular matrix as the WT type (referred to as SPDMP1) in MC3T3 osteoblasts. High levels of DMP1 in either form greatly increased osteogenic gene expression in an identical manner. However, the targeted NLSDMP1 transgene driven by a 3.6-kb rat Col 1α1 promoter in the nucleus of osteoblasts and osteocytes failed to rescue the phenotyope of Dmp1-null mice, whereas the SPDMP1 transgene rescued the rickets defect. These studies support the notion that DMP1 functions as an extracellular matrix protein, rather than as a transcription co-factor in vivo. We also show that DMP1 continues its expression in osteoblasts during postnatal development and that the deletion of Dmp1 leads to an increase in osteoblast proliferation. However, poor mineralization in the metaphysis indicates a critical role for DMP1 in both osteoblasts and osteocytes. PMID:24917674

  5. Heterochromatin Protein 1 Binding Protein 3 Expression as a Candidate Marker of Intrinsic 5-Fluorouracil Resistance

    PubMed Central

    HADAC, JAMIE N.; MILLER, DEVON D.; GRIMES, IAN C.; CLIPSON, LINDA; NEWTON, MICHAEL A.; SCHELMAN, WILLIAM R.; HALBERG, RICHARD B.

    2016-01-01

    Background Despite receiving post-operative 5-fluorouracil (5-FU)-based chemotherapy, approximately 50% of patients with stage IIIC colon cancer experience recurrence. Currently, no molecular signature can predict response to 5-FU. Materials and Methods Mouse models of colon cancer have been developed and characterized. Individual tumors in these mice can be longitudinally monitored and assessed to identify differences between those that are responsive and those that are resistant to therapy. Gene expression was analyzed in serial biopsies that were collected before and after treatment with 5-FU. Colon tumors had heterogeneous responses to treatment with 5-FU. Microarray analysis of pretreatment biopsies revealed that Hp1bp3, a gene encoding heterochromatin protein 1 binding protein 3, was differentially expressed between sensitive and resistant tumors. Conclusion Using mouse models of human colorectal cancer, Hp1bp3 was identified as a candidate marker of intrinsic 5-FU resistance and may represent a potential biomarker for patient stratification or a target of clinical importance. PMID:26976970

  6. A comparative approach to understanding tissue-specific expression of uncoupling protein 1 expression in adipose tissue.

    PubMed

    Shore, Andrew; Emes, Richard D; Wessely, Frank; Kemp, Paul; Cillo, Clemente; D'Armiento, Maria; Hoggard, Nigel; Lomax, Michael A

    2012-01-01

    The thermoregulatory function of brown adipose tissue (BAT) is due to the tissue-specific expression of uncoupling protein 1 (UCP1) which is thought to have evolved in early mammals. We report that a CpG island close to the UCP1 transcription start site is highly conserved in all 29 vertebrates examined apart from the mouse and xenopus. Using methylation sensitive restriction digest and bisulfite mapping we show that the CpG island in both the bovine and human is largely un-methylated and is not related to differences in UCP1 expression between white and BAT. Tissue-specific expression of UCP1 has been proposed to be regulated by a conserved 5' distal enhancer which has been reported to be absent in marsupials. We demonstrate that the enhancer, is also absent in five eutherians as well as marsupials, monotremes, amphibians, and fish, is present in pigs despite UCP1 having become a pseudogene, and that absence of the enhancer element does not relate to BAT-specific UCP1 expression. We identify an additional putative 5' regulatory unit which is conserved in 14 eutherian species but absent in other eutherians and vertebrates, but again unrelated to UCP1 expression. We conclude that despite clear evidence of conservation of regulatory elements in the UCP1 5' untranslated region, this does not appear to be related to species or tissues-specific expression of UCP1. PMID:23293654

  7. High mobility group protein 1: A collaborator in nucleosome dynamics and estrogen-responsive gene expression

    PubMed Central

    Scovell, William M

    2016-01-01

    High mobility group protein 1 (HMGB1) is a multifunctional protein that interacts with DNA and chromatin to influence the regulation of transcription, DNA replication and repair and recombination. We show that HMGB1 alters the structure and stability of the canonical nucleosome (N) in a nonenzymatic, adenosine triphosphate-independent manner. As a result, the canonical nucleosome is converted to two stable, physically distinct nucleosome conformers. Although estrogen receptor (ER) does not bind to its consensus estrogen response element within a nucleosome, HMGB1 restructures the nucleosome to facilitate strong ER binding. The isolated HMGB1-restructured nucleosomes (N’ and N’’) remain stable and exhibit a number of characteristics that are distinctly different from the canonical nucleosome. These findings complement previous studies that showed (1) HMGB1 stimulates in vivo transcriptional activation at estrogen response elements and (2) knock down of HMGB1 expression by siRNA precipitously reduced transcriptional activation. The findings indicate that a major facet of the mechanism of HMGB1 action involves a restructuring of aspects of the nucleosome that appear to relax structural constraints within the nucleosome. The findings are extended to reveal the differences between ER and the other steroid hormone receptors. A working proposal outlines mechanisms that highlight the multiple facets that HMGB1 may utilize in restructuring the nucleosome. PMID:27247709

  8. mRNA expression and protein localization of dentin matrix protein 1 during dental root formation.

    PubMed

    Toyosawa, S; Okabayashi, K; Komori, T; Ijuhin, N

    2004-01-01

    Dentin matrix protein 1 (DMP1) is an acidic phosphoprotein. DMP1 was initially detected in dentin and later in other mineralized tissues including cementum and bone, but the DMP1 expression pattern in tooth is still controversial. To determine the precise localization of DMP1 messenger RNA (mRNA) and the protein in the tooth, we performed in situ hybridization and immunohistochemical analyses using rat molars and incisors during various stages of root formation. During root dentin formation of molars, DMP1 mRNA was detected in root odontoblasts in parallel with mineralization of the dentin. However, the level of DMP1 mRNA expression in root odontoblasts decreased near the coronal part and was absent in coronal odontoblasts. DMP1 protein was localized along dentinal tubules and their branches in mineralized root dentin, and the distribution of DMP1 shifted from the end of dentinal tubules to the base of the tubules as dentin formation progressed. During the formation of the acellular cementum, DMP1 mRNA was detected in cementoblasts lining the acellular cementum where its protein was localized. During the formation of the cellular cementum, DMP1 mRNA was detected in cementocytes embedded in the cellular cementum but not in cementoblasts, and its protein was localized in the pericellular cementum of cementocytes including their processes. During dentin formation of incisors, DMP1 mRNA was detected in odontoblasts on the cementum-related dentin, where its protein was localized along dentinal tubules near the mineralization front. The localization of DMP1 mRNA and protein in dentin and cementum was related to their mineralization, suggesting that one of the functions of DMP1 may be involved in the mineralization of dentin and cementum during root formation. PMID:14751569

  9. Porcine dentin matrix protein 1: gene structure, cDNA sequence, and expression in teeth.

    PubMed

    Kim, Jung-Wook; Yamakoshi, Yasuo; Iwata, Takanori; Hu, Yuan Yuan; Zhang, Hengmin; Hu, Jan C-C; Simmer, James P

    2006-02-01

    Dentin matrix protein 1 (DMP1) is an acidic non-collagenous protein that is necessary for the proper biomineralization of bone, cartilage, cementum, dentin, and enamel. Dentin matrix protein 1 is highly phosphorylated and potentially glycosylated, but there is no experimental data identifying which specific amino acids are modified. For the purpose of facilitating the characterization of DMP1 from pig, which has the advantage of large developing teeth for obtaining protein in quantity and extensive structural information concerning other tooth matrix proteins, we characterized the porcine DMP1 cDNA and gene structure, raised anti-peptide immunoglobulins that are specific for porcine DMP1, and detected DMP1 protein in porcine tooth extracts and histological sections. Porcine DMP1 has 510 amino acids, including a 16-amino acid signal peptide. The deduced molecular weight of the secreted, unmodified protein is 53.5 kDa. The protein has 93 serines and 12 threonines in the appropriate context for phosphorylation, and four asparagines in a context suitable for glycosylation. Dentin matrix protein 1 protein bands with apparent molecular weights between 30 and 45 kDa were observed in partially purified dentin extracts. In developing teeth, immunohistochemistry localized DMP1 in odontoblasts and the dentinal tubules of mineralized dentin and in ameloblasts, but not in the enamel matrix.

  10. Leukocyte chemoattractant activity of diacylglycerol

    SciTech Connect

    Wright, T.M.; Hoffman, R.D.; Nishijima, J.; Shin, H.S.

    1986-03-05

    Phosphatidylinositol breakdown with the generation of 1,2-diacylglycerol (1,2-DG) and inositol phosphates occurs in response to receptor mediated stimulation of lymphocytes and polymorphonuclear neutrophils (PMN). In the authors attempt to demonstrate the direct role of 1,2-DG in cell migration, they have found 1,2 dioctanoyl glycerol (1,2-C8DG) to be a chemoattractant for 6C3HED, a mouse thymic lymphoma, and human peripheral blood PMN's. The chemoattractant activity for both cell types was observed at concentrations from 0.5 to 10mM in an under agarose assay. The maximum effect of 1,2-C8DG on 6C3HED cells was similar to that of 1mM lysophosphatidylcholine and the maximum effect of 1,2-C8DG on PMN's was similar to that of 10/sup -7/M f-met-leu-phe. Other 1,2-DG's with acyl chains ranging from 6 to 18 carbons in length and 1-oleoyl-2-acetyl-glycerol were also chemoattractants for 6C3HED, although their activities were less than 1,2-C8DG. In addition, phorbol myristate acetate (PMA), another activator of protein kinase C, was a chemoattractant for 6C3HED and human PMN's. PMA was more potent than 1,2-C8DG for both 6C3HED and PMN's with chemoattractant activity in the range of 30nM to 1..mu..M. These studies support the direct role of 1,2-DG in the transduction of chemotactic stimuli in leukocytes and further suggest that the formation of diacylglycerol represents a common step in the migratory responses of lymphoid and myeloid cells.

  11. Allosteric Modulation of Chemoattractant Receptors

    PubMed Central

    Allegretti, Marcello; Cesta, Maria Candida; Locati, Massimo

    2016-01-01

    Chemoattractants control selective leukocyte homing via interactions with a dedicated family of related G protein-coupled receptor (GPCR). Emerging evidence indicates that the signaling activity of these receptors, as for other GPCR, is influenced by allosteric modulators, which interact with the receptor in a binding site distinct from the binding site of the agonist and modulate the receptor signaling activity in response to the orthosteric ligand. Allosteric modulators have a number of potential advantages over orthosteric agonists/antagonists as therapeutic agents and offer unprecedented opportunities to identify extremely selective drug leads. Here, we resume evidence of allosterism in the context of chemoattractant receptors, discussing in particular its functional impact on functional selectivity and probe/concentration dependence of orthosteric ligands activities. PMID:27199992

  12. Histone Deacetylase Inhibitors Activate Tristetraprolin Expression through Induction of Early Growth Response Protein 1 (EGR1) in Colorectal Cancer Cells

    PubMed Central

    Sobolewski, Cyril; Sanduja, Sandhya; Blanco, Fernando F.; Hu, Liangyan; Dixon, Dan A.

    2015-01-01

    The RNA-binding protein tristetraprolin (TTP) promotes rapid decay of mRNAs bearing 3' UTR AU-rich elements (ARE). In many cancer types, loss of TTP expression is observed allowing for stabilization of ARE-mRNAs and their pathologic overexpression. Here we demonstrate that histone deacetylase (HDAC) inhibitors (Trichostatin A, SAHA and sodium butyrate) promote TTP expression in colorectal cancer cells (HCA-7, HCT-116, Moser and SW480 cells) and cervix carcinoma cells (HeLa). We found that HDAC inhibitors-induced TTP expression, promote the decay of COX-2 mRNA, and inhibit cancer cell proliferation. HDAC inhibitors were found to promote TTP transcription through activation of the transcription factor Early Growth Response protein 1 (EGR1). Altogether, our findings indicate that loss of TTP in tumors occurs through silencing of EGR1 and suggests a therapeutic approach to rescue TTP expression in colorectal cancer. PMID:26343742

  13. Low expression of nucleus accumbens-associated protein 1 predicts poor prognosis for patients with pancreatic ductal adenocarcinoma.

    PubMed

    Nishi, Takeshi; Maruyama, Riruke; Urano, Takeshi; Nakayama, Naomi; Kawabata, Yasunari; Yano, Seiji; Yoshida, Manabu; Nakayama, Kentaro; Miyazaki, Kohji; Takenaga, Keizo; Tanaka, Tsuneo; Tajima, Yoshitsugu

    2012-12-01

    Nucleus accumbens-associated protein 1 (NAC1) is overexpressed in various carcinomas including ovarian, cervical, breast, and pancreatic carcinomas. High expression of NAC1 is considered to have adverse effects on prognosis through negative regulation of growth arrest and DNA-damage-inducible 45-γ interacting protein 1 (GADD45GIP1) in ovarian and cervical carcinomas. In the present study, the expression of NAC1 in pancreatic ductal adenocarcinoma (PDA) was measured using immunohistochemistry and computer-assisted image analysis in order to investigate its correlation with various clinicopathological parameters and prognosis. Patients with low-NAC1 PDA had worse overall survival (P = 0.0010) and a shorter disease-free survival (P = 0.0036) than patients with high-NAC1 PDA. This was a clinical effect opposite to that reported in ovarian and cervical carcinomas. Furthermore, knockdown of NAC1 in pancreatic carcinoma cell lines did not increase expression of the GADD45GIP1 protein. These results indicate that the gene(s) regulated by NAC1 vary depending on the types of carcinoma or originating tissue, and that low expression of NAC1 predicts poor prognosis for patients with PDA.

  14. Age-dependent increase in the expression of antioxidant-like protein-1 in the gerbil hippocampus

    PubMed Central

    Park, Jin-A; Park, Joon Ha; Ahn, Ji Hyeon; Kim, Jong-Dai; Won, Moo-Ho; Lee, Choong-Hyun

    2016-01-01

    Antioxidant-like protein-1 (AOP-1) reduces the intracellular level of reactive oxygen species. In the present study, the age-related change in AOP-1 expression in the hippocampus among young, adult and aged gerbils was compared using western blot analysis and immunohistochemistry. The results demonstrated that the protein expression of AOP-1 was gradually and significantly increased in the hippocampus during the normal aging process. In addition, the age-dependent increase in AOP-1 immunoreactivity was also observed in pyramidal neurons of the hippocampus proper; however, in the dentate gyrus, AOP-1 immunoreactivity was not altered during the normal aging process. These results indicated that the expression of AOP-1 is significantly increased in the hippocampus proper, but not in the dentate gyrus, during the normal aging process. PMID:27511601

  15. Biotype expression and insecticide response of Bemisia tabaci chemosensory protein-1.

    PubMed

    Liu, Guo Xia; Xuan, Ning; Chu, Dong; Xie, Hong Yan; Fan, Zhong Xue; Bi, Yu Ping; Picimbon, Jean-François; Qin, Yu Chuan; Zhong, Su Ting; Li, Yao Fa; Gao, Zhan Lin; Pan, Wen Liang; Wang, Guo Ying; Rajashekar, Balaji

    2014-03-01

    Chemosensory proteins (CSPs) are a group of small soluble proteins found so far exclusively in arthropod species. These proteins act in chemical communication and perception. In this study, a gene encoding the Type 1 CSP (BtabCSP1) from the agricultural pest Bemisia tabaci (whitefly) was analyzed to understand sequence variation and expression specificity in different biotypes. Sequence analysis of BtabCSP1 showed significant differences between the two genetically characterized biotypes, B and Q. The B-biotype had a larger number of BtabCSP1 mutations than the Q-biotype. Similar to most other CSPs, BtabCSP1 was more expressed in the head than in the rest of the body. One-step RT-PCR and qPCR analysis on total messenger RNA showed that biotype-Q had higher BtabCSP1 expression levels than biotype-B. Females from a mixed field-population had high levels of BtabCSP1 expression. The interaction of BtabCSP1 with the insecticide thiamethoxam was investigated by analyzing the BtabCSP1 expression levels following exposure to the neonicotinoid, thiamethoxam, in a time/dose-response study. Insecticide exposure increased BtabCSP1 expression (up to tenfold) at 4 and 24 h following 50 or 100 g/ml treatments. PMID:24478049

  16. Melatonin decreases breast cancer metastasis by modulating Rho-associated kinase protein-1 expression

    PubMed Central

    Borin, Thaiz Ferraz; Arbab, Ali Syed; Gelaleti, Gabriela Bottaro; Ferreira, Lívia Carvalho; Moschetta, Marina Gobbe; Jardim-Perassi, Bruna Victorasso; Iskander, ASM; Varma, Nadimpalli Ravi S.; Shankar, Adarsh; Coimbra, Verena Benedick; Fabri, Vanessa Alves; de Oliveira, Juliana Garcia; de Campos Zuccari, Debora Aparecida Pires

    2016-01-01

    The occurrence of metastasis, an important breast cancer prognostic factor, depends on cell migration/invasion mechanisms, which can be controlled by regulatory and effector molecules such as Rho-associated kinase protein (ROCK-1). Increased expression of this protein promotes tumor growth and metastasis, which can be restricted by ROCK-1 inhibitors. Melatonin has shown oncostatic, antimetastatic, and anti-angiogenic effects and can modulate ROCK-1 expression. Metastatic and nonmetastatic breast cancer cell lines were treated with melatonin as well as with specific ROCK-1 inhibitor (Y27632). Cell viability, cell migration/invasion, and ROCK-1 gene expression and protein expression were determined in vitro. In vivo lung metastasis study was performed using female athymic nude mice treated with either melatonin or Y27832 for 2 and 5 wk. The metastases were evaluated by X-ray computed tomography and single photon emission computed tomography (SPECT) and by immunohistochemistry for ROCK-1 and cytokeratin proteins. Melatonin and Y27632 treatments reduced cell viability and invasion/migration of both cell lines and decreased ROCK-1 gene expression in metastatic cells and protein expression in nonmetastatic cell line. The numbers of ‘hot’ spots (lung metastasis) identified by SPECT images were significantly lower in treated groups. ROCK-1 protein expression also was decreased in metastatic foci of treated groups. Melatonin has shown to be effective in controlling metastatic breast cancer in vitro and in vivo, not only via inhibition of the proliferation of tumor cells but also through direct antagonism of metastatic mechanism of cells rendered by ROCK-1 inhibition. When Y27632 was used, the effects were similar to those found with melatonin treatment. PMID:26292662

  17. Melatonin decreases breast cancer metastasis by modulating Rho-associated kinase protein-1 expression.

    PubMed

    Borin, Thaiz Ferraz; Arbab, Ali Syed; Gelaleti, Gabriela Bottaro; Ferreira, Lívia Carvalho; Moschetta, Marina Gobbe; Jardim-Perassi, Bruna Victorasso; Iskander, A S M; Varma, Nadimpalli Ravi S; Shankar, Adarsh; Coimbra, Verena Benedick; Fabri, Vanessa Alves; de Oliveira, Juliana Garcia; Zuccari, Debora Aparecida Pires de Campos

    2016-01-01

    The occurrence of metastasis, an important breast cancer prognostic factor, depends on cell migration/invasion mechanisms, which can be controlled by regulatory and effector molecules such as Rho-associated kinase protein (ROCK-1). Increased expression of this protein promotes tumor growth and metastasis, which can be restricted by ROCK-1 inhibitors. Melatonin has shown oncostatic, antimetastatic, and anti-angiogenic effects and can modulate ROCK-1 expression. Metastatic and nonmetastatic breast cancer cell lines were treated with melatonin as well as with specific ROCK-1 inhibitor (Y27632). Cell viability, cell migration/invasion, and ROCK-1 gene expression and protein expression were determined in vitro. In vivo lung metastasis study was performed using female athymic nude mice treated with either melatonin or Y27832 for 2 and 5 wk. The metastases were evaluated by X-ray computed tomography and single photon emission computed tomography (SPECT) and by immunohistochemistry for ROCK-1 and cytokeratin proteins. Melatonin and Y27632 treatments reduced cell viability and invasion/migration of both cell lines and decreased ROCK-1 gene expression in metastatic cells and protein expression in nonmetastatic cell line. The numbers of 'hot' spots (lung metastasis) identified by SPECT images were significantly lower in treated groups. ROCK-1 protein expression also was decreased in metastatic foci of treated groups. Melatonin has shown to be effective in controlling metastatic breast cancer in vitro and in vivo, not only via inhibition of the proliferation of tumor cells but also through direct antagonism of metastatic mechanism of cells rendered by ROCK-1 inhibition. When Y27632 was used, the effects were similar to those found with melatonin treatment. PMID:26292662

  18. eIF4E binding protein 1 expression is associated with clinical survival outcomes in colorectal cancer

    PubMed Central

    Chao, Min-Wu; Wang, Li-Ting; Lai, Chin-Yu; Yang, Xiao-Ming; Cheng, Ya-Wen; Lee, Kuo-Hsiung

    2015-01-01

    eIF4E binding protein 1 (4E-BP1), is critical for cap-dependent and cap-independent translation. This study is the first to demonstrate that 4E-BP1 expression correlates with colorectal cancer (CRC) progression. Compared to its expression in normal colon epithelial cells, 4E-BP1 was upregulated in CRC cell lines and was detected in patient tumor tissues. Furthermore, high 4E-BP1 expression was statistically associated with poor prognosis. Hypoxia has been considered as an obstacle for cancer therapeutics. Our previous data showed that YXM110, a cryptopleurine derivative, exhibited anticancer activity via 4E-BP1 depletion. Here, we investigated whether YXM110 could inhibit protein synthesis under hypoxia. 4E-BP1 expression was notably decreased by YXM110 under hypoxic conditions, implying that cap-independent translation could be suppressed by YXM110. Moreover, YXM110 repressed hypoxia-inducible factor 1α (HIF-1α) expression, which resulted in decreased downstream vascular endothelial growth factor (VEGF) expression. These observations highlight 4E-BP1 as a useful biomarker and therapeutic target, indicating that YXM110 could be a potent CRC therapeutic drug. PMID:26204490

  19. First Evidence of Bone Morphogenetic Protein 1 Expression and Activity in Sheep Ovarian Follicles1

    PubMed Central

    Canty-Laird, Elizabeth; Carré, Gwenn-Aël; Mandon-Pépin, Béatrice; Kadler, Karl E.; Fabre, Stéphane

    2010-01-01

    Bone morphogenetic protein (BMP) 1 is a vertebrate metalloproteinase of the astacin family. BMP1 plays a key role in regulating the formation of the extracellular matrix (ECM), particularly by processing the C-propeptide of fibrillar procollagens. BMP1 also promotes BMP signaling by releasing BMP signaling molecules from complexes with the BMP-antagonist chordin. As a result of BMP1′s dual role in both ECM formation and BMP signaling, we hypothesized that BMP1 could play a role in ovarian physiology. Using the sheep ovary as a model system, we showed that BMP1 was expressed in the ovary throughout early fetal stages to adulthood. Furthermore, in adult ovaries, BMP1 was expressed along with chordin, BMP4, and twisted gastrulation, which together form an extracellular regulatory complex for BMP signaling. Within ovine ovaries, immunohistochemical localization demonstrated that BMP1 was present in granulosa cells at all stages of follicular development, from primordial to large antral follicles, and that the levels of BMP1 were not affected by the final follicle selection mechanism. In cultured granulosa cells, BMP1 expression was not affected by gonadotropins, but BMP4 and activin A had opposing effects on the levels of BMP1 mRNA. BMP1 appeared to be secreted into the follicular fluid of antral follicles, where it is able to exert procollagen C-proteinase and chordinase activities. Interestingly, BMP1 activity in follicular fluid decreased with follicular growth. PMID:20357269

  20. Oxidored-nitro domain-containing protein 1 expression is associated with the progression of hepatocellular carcinoma

    PubMed Central

    LI, DENG-QING; QIU, MING; NIE, XIN-MIN; GUI, RONG; HUANG, MIN-ZHU

    2016-01-01

    Hepatocarcinogenesis is a stepwise process during which multiple genes are altered. Understanding the molecular mechanisms that induce hepatocarcinogenesis may improve the screening, prevention and treatment of patients with hepatocellular carcinoma (HCC). In recent years, the oxidored-nitro domain-containing protein 1 (NOR1) gene has been identified to have an important role in the development of HCC in vitro experiments. The current study aimed to examine the expression of NOR1 mRNA and protein expression in specimens of normal liver, hepatitis, cirrhosis and HCC, together representing the process of HCC development. Furthermore, the association between NOR1 expression and clinicopathological parameters of HCC patients was analyzed. Tissue microarrays containing the specimens of human normal liver, hepatitis, cirrhosis and HCC were purchased, and in situ hybridization and immunohistochemistry were used to detect the expression of NOR1 mRNA and protein expression, respectively. It was revealed that the positive rate of NOR1 protein and mRNA expression in the specimens of hepatitis and cirrhosis were not significantly different from that in the normal liver samples. However, the specimens of HCC exhibited an increased positive rate of NOR1 protein and mRNA expression in comparison with the normal liver samples. In addition, a higher positive rate of NOR1 protein expression was observed in HCC patients with a poor pathological differentiation grade and high tumor node metastasis (TNM) stage. In conclusion, the present study provides evidence, for the first time, of the increased expression of NOR1 in human HCC tissues, and its correlation with the pathological stage and TNM status. These findings indicate that NOR1 may be involved in the progression of HCC and it could be employed as a predictive biomarker in HCC development. PMID:27123053

  1. Reduced expression of plasma membrane calcium ATPase 2 and collapsin response mediator protein 1 promotes death of spinal cord neurons.

    PubMed

    Kurnellas, M P; Li, H; Jain, M R; Giraud, S N; Nicot, A B; Ratnayake, A; Heary, R F; Elkabes, S

    2010-09-01

    The mechanisms underlying neuronal pathology and death in the spinal cord (SC) during inflammation remain elusive. We previously showed the important role of plasma membrane calcium ATPases (PMCAs) in the survival of SC neurons, in vitro. We also postulated that a decrease in PMCA2 expression could cause neuronal death during experimental autoimmune encephalomyelitis (EAE), an animal model of multiple sclerosis. The current studies were undertaken to define the specific contribution of PMCA2 to degeneration of SC neurons, the effectors downstream to PMCA2 mediating neuronal death and the triggers that reduce PMCA2 expression. We report that knockdown of PMCA2 in SC neurons decreases collapsin response mediator protein 1 (CRMP1) levels. This is followed by cell death. Silencing of CRMP1 expression also leads to neuronal loss. Kainic acid reduces both PMCA2 and CRMP1 levels and induces neuronal death. Administration of an alpha-amino-3-hydroxyl-5-methyl-4-isoxazole-propionate (AMPA)/kainate receptor antagonist, at onset or peak of EAE, restores the decreased PMCA2 and CRMP1 levels to control values and ameliorates clinical deficits. Thus, our data link the reduction in PMCA2 expression with perturbations in the expression of CRMP1 and the ensuing death of SC neurons. This represents an additional mechanism underlying AMPA/kainate receptor-mediated excitotoxicity with relevance to neurodegeneration in EAE. PMID:20489728

  2. Interleukin-1β stimulates macrophage inflammatory protein-1α and -1β expression in human neuronal cells (NT2-N)

    PubMed Central

    Guo, Chang-Jiang; Douglas, Steven D.; Lai, Jian-Ping; Pleasure, David E.; Li, Yuan; Williams, Marge; Bannerman, Peter; Song, Li; Ho, Wen-Zhe

    2014-01-01

    Chemokines are important mediators in immune responses and inflammatory processes of neuroimmunologic and infectious diseases. Although chemokines are expressed predominantly by cells of the immune system, neurons also express chemokines and chemokine receptors. We report herein that human neuronal cells (NT2-N) produce macrophage inflammatory protein-1α and -1β (MIP-1α and MIP-1β), which could be enhanced by interleukin (IL)-1β at both mRNA and protein levels. The addition of supernatants from human peripheral blood monocyte-derived macrophage (MDM) cultures induced MIP-1β mRNA expression in NT2-N cells. Anti-IL-1β antibody removed most, but not all, of the MDM culture supernatant-induced MIP-1β mRNA expression in NT2-N cells, suggesting that IL-1β in the MDM culture supernatants is a major factor in the induction of MIP-1β expression. Investigation of the mechanism(s) responsible for IL-1β-induced MIP-1α and -1β expression demonstrated that IL-1β activated nuclear factor kappa B (NF-κB) promoter-directed luciferase activity in NT2-N cells. Caffeic acid phenethyl ester, a potent and specific inhibitor of activation of NF-κB, not only blocked IL-1β-induced activation of the NF-κB promoter but also decreased IL-1β-induced MIP-1α and -1β expression in NT2-N cells. These data suggest that NF-κB is at least partially involved in the IL-1β-mediated action on MIP-1α and -1β in NT2-N cells. IL-1β-mediated up-regulation of β-chemokine expression may have important implications in the immuno-pathogenesis of inflammatory diseases in the CNS. PMID:12603824

  3. Expression of ribosome-binding protein 1 correlates with shorter survival in Her-2 positive breast cancer

    PubMed Central

    Liang, Xiaoshuan; Sun, Shanshan; Zhang, Xianyu; Wu, Hao; Tao, Weiyang; Liu, Tong; Wei, Wei; Geng, Jingshu; Pang, Da

    2015-01-01

    The aim of this study is to investigate the expression of ribosome-binding protein 1 (RRBP1) in invasive breast cancer and to analyze its relationship to clinical features and prognosis. RRBP1 expression was studied using real-time quantitative PCR and western blotting using pair-matched breast samples and immunohistochemical staining using a tissue microarray. Then the correlation between RRBP1 expression and clinicopathologic features was analyzed. RRBP1 mRNA and protein expression were significantly increased in breast cancer tissues compared with normal tissues. The protein level of RRBP1 is proved to be positively related to histological grade (P = 0.02), molecular subtype (P = 0.048) and status of Her-2 (P = 0.026) and P53 (P = 0.015). We performed a grade-stratified analysis of all patients according to the level of RRBP1 expression and found that RRBP1 overexpression highly affected overall survival in patients with early-stage (I and II) tumors (P = 0.042). Furthermore, Her-2 positive patients with negative RRBP1 expression had longer overall survival rates than those with positive RRBP1 expression (P = 0.031). Using multivariate analysis, it was determined that lymph node metastasis (LNM, P = 0.002) and RRBP1 expression (P = 0.005) were independent prognosis factors for overall survival. RRBP1 is a valuable prognostic factor in Her-2-positive breast cancer patients, indicating that RRBP1 is a potentially important target for the prediction of prognosis. PMID:25845758

  4. Curcumin inhibits srebp-2 expression in activated hepatic stellate cells in vitro by reducing the activity of specificity protein-1.

    PubMed

    Kang, Qiaohua; Chen, Anping

    2009-12-01

    Elevated levels of cholesterol/low-density lipoprotein (LDL) are a risk factor for the development of nonalcoholic steatohepatitis and its associated hepatic fibrosis. However, underlying mechanisms remain elusive. We previously reported that curcumin induced gene expression of peroxisome proliferator-activated receptor (PPAR)-gamma and stimulated its activity, leading to the inhibition of the activation of hepatic stellate cells (HSCs), the major effector cells during hepatic fibrogenesis. We recently showed that curcumin suppressed gene expression of LDL receptor in activated HSCs in vitro by repressing gene expression of the transcription factor sterol regulatory element binding protein-2 (SREBP-2), leading to the reduction in the level of intracellular cholesterol in HSCs and to the attenuation of the stimulatory effects of LDL on HSCs activation. The current study aimed at exploring molecular mechanisms by which curcumin inhibits srebp-2 expression in HSCs. Promoter deletion assays, mutagenesis assays, and EMSAs localize a specificity protein-1 (SP-1) binding GC-box in the srebp-2 promoter, which is responsible for enhancing the promoter activity and responding to curcumin in HSCs. Curcumin suppresses gene expression of SP-1 and reduces its trans-activation activity, which are mediated by the activation of PPARgamma. The inhibitory effect of curcumin on SP-1 binding to the GC-box is confirmed by chromatin immuno-precipitation. In summary, our results demonstrate that curcumin inhibits srebp-2 expression in cultured HSCs by activating PPARgamma and reducing the SP-1 activity, leading to the repression of ldlr expression. These results provide novel insights into molecular mechanisms by which curcumin inhibits LDL-induced HSC activation.

  5. Leukocyte chemoattractant receptors in human disease pathogenesis.

    PubMed

    Zabel, Brian A; Rott, Alena; Butcher, Eugene C

    2015-01-01

    Combinations of leukocyte attractant ligands and cognate heptahelical receptors specify the systemic recruitment of circulating cells by triggering integrin-dependent adhesion to endothelial cells, supporting extravasation, and directing specific intratissue localization via gradient-driven chemotaxis. Chemoattractant receptors also control leukocyte egress from lymphoid organs and peripheral tissues. In this article, we summarize the fundamental mechanics of leukocyte trafficking, from the evolution of multistep models of leukocyte recruitment and navigation to the regulation of chemoattractant availability and function by atypical heptahelical receptors. To provide a more complete picture of the migratory circuits involved in leukocyte trafficking, we integrate a number of nonchemokine chemoattractant receptors into our discussion. Leukocyte chemoattractant receptors play key roles in the pathogenesis of autoimmune diseases, allergy, inflammatory disorders, and cancer. We review recent advances in our understanding of chemoattractant receptors in disease pathogenesis, with a focus on genome-wide association studies in humans and the translational implications of mechanistic studies in animal disease models.

  6. Short communication: Altered expression of specificity protein 1 impairs milk fat synthesis in goat mammary epithelial cells.

    PubMed

    Zhu, J J; Luo, J; Xu, H F; Wang, H; Loor, J J

    2016-06-01

    Specificity protein 1 (encoded by SP1) is a novel transcription factor important for the regulation of lipid metabolism and the normal function of various hormones in model organisms. Its potential role, if any, on ruminant milk fat is unknown. Despite the lower expression of the lipolysis-related gene ATGL (by 44 and 37% respectively), both the adenoviral overexpression and the silencing of SP1 [via short interfering (si)RNA] markedly reduced cellular triacylglycerol (TAG) content (by 28 and 25%, respectively), at least in part by decreasing the expression of DGAT1 (-36% in adenovirus treatment) and DGAT2 (-81 and -87%, respectively) that are involved in TAG synthesis. Consistent with the markedly lower expression of genes related to lipid droplet formation and secretion (TIP47 by 19 and 32%, and ADFP by 25 and 25%, respectively), cellular lipid droplet content was also decreased sharply, by 9 and 8.5%, respectively, after adenoviral overexpression of SP1 or its silencing via siRNA. Overall, the results underscored a potentially important role of SP1 in maintaining milk-fat droplet synthesis in goat mammary epithelial cells. PMID:26995134

  7. Novel nuclear protein ALC-INTERACTING PROTEIN1 is expressed in vascular and mesocarp cells in Arabidopsis.

    PubMed

    Wang, Fang; Shi, Dong-Qiao; Liu, Jie; Yang, Wei-Cai

    2008-07-01

    Pod shattering is an agronomical trait that is a result of the coordinated action of cell differentiation and separation. In Arabidopsis, pod shattering is controlled by a complex genetic network in which ALCATRAZ (ALC), a member of the basic helix-loop-helix family, is critical for cell separation during fruit dehiscence. Herein, we report the identification of ALC-INTERACTING PROTEIN1 (ACI1) via the yeast two-hybrid screen. ACI1 encodes a nuclear protein with a lysine-rich domain and a C-terminal serine-rich domain. ACI1 is mainly expressed in the vascular system throughout the plant and mesocarp of the valve in siliques. Our data showed that ACI1 interacts strongly with the N-terminal portion of ALC in yeast cells and in plant cells in the nucleus as demonstrated by bimolecular fluorescence complementation assay. Both ACI1 and ALC share an overlapping expression pattern, suggesting that they likely function together in planta. However, no detectable phenotype was found in plants with reduced ACI1 expression by RNA interference technology, suggesting that ACI1 may be redundant. Taken together, these data indicate that ALC may interact with ACI1 and its homologs to control cell separation during fruit dehiscence in Arabidopsis. PMID:18713402

  8. Short communication: Altered expression of specificity protein 1 impairs milk fat synthesis in goat mammary epithelial cells.

    PubMed

    Zhu, J J; Luo, J; Xu, H F; Wang, H; Loor, J J

    2016-06-01

    Specificity protein 1 (encoded by SP1) is a novel transcription factor important for the regulation of lipid metabolism and the normal function of various hormones in model organisms. Its potential role, if any, on ruminant milk fat is unknown. Despite the lower expression of the lipolysis-related gene ATGL (by 44 and 37% respectively), both the adenoviral overexpression and the silencing of SP1 [via short interfering (si)RNA] markedly reduced cellular triacylglycerol (TAG) content (by 28 and 25%, respectively), at least in part by decreasing the expression of DGAT1 (-36% in adenovirus treatment) and DGAT2 (-81 and -87%, respectively) that are involved in TAG synthesis. Consistent with the markedly lower expression of genes related to lipid droplet formation and secretion (TIP47 by 19 and 32%, and ADFP by 25 and 25%, respectively), cellular lipid droplet content was also decreased sharply, by 9 and 8.5%, respectively, after adenoviral overexpression of SP1 or its silencing via siRNA. Overall, the results underscored a potentially important role of SP1 in maintaining milk-fat droplet synthesis in goat mammary epithelial cells.

  9. Association between p53-binding protein 1 expression and genomic instability in oncocytic follicular adenoma of the thyroid.

    PubMed

    Mussazhanova, Zhanna; Akazawa, Yuko; Matsuda, Katsuya; Shichijo, Kazuko; Miura, Shiro; Otsubo, Ryota; Oikawa, Masahiro; Yoshiura, Koh-Ichiro; Mitsutake, Norisato; Rogounovitch, Tatiana; Saenko, Vladimir; Kozykenova, Zhanna; Zhetpisbaev, Bekbolat; Shabdarbaeva, Dariya; Sayakenov, Nurlan; Amantayev, Bakanay; Kondo, Hisayoshi; Ito, Masahiro; Nakashima, Masahiro

    2016-05-31

    Oncocytic follicular adenomas (FAs) of the thyroid are neoplasms of follicular cell origin that are predominantly composed of large polygonal cells with eosinophilic and granular cytoplasm. However, the pathological characteristics of these tumors are largely unexplored. Both the initiation and progression of cancer can be caused by an accumulation of genetic mutations that can induce genomic instability. Thus, the aim of this study was to evaluate the extent of genomic instability in oncocytic FA. As the presence of p53-binding protein 1 (53BP1) in nuclear foci has been found to reflect DNA double-strand breaks that are triggered by various stresses, the immunofluorescence expression pattern of 53BP-1 was assessed in oncocytic and conventional FA. The association with the degree of DNA copy number aberration (CNA) was also evaluated using array-based comparative genomic hybridization. Data from this study demonstrated increased 53BP1 expression (i.e., "unstable" expression) in nuclear foci of oncocytic FA and a higher incidence of CNAs compared with conventional FA. There was also a particular focus on the amplification of chromosome 1p36 in oncocytic FA, which includes the locus for Tumor protein 73, a member of the p53 family implicated as a factor in the development of malignancies. Further evaluations revealed that unstable 53BP1 expression had a significant positive correlation with the levels of expression of Tumor protein 73. These data suggest a higher level of genomic instability in oncocytic FA compared with conventional FA, and a possible relationship between oncocytic FA and abnormal amplification of Tumor protein 73. PMID:26935218

  10. Mast Cell Chemotaxis – Chemoattractants and Signaling Pathways

    PubMed Central

    Halova, Ivana; Draberova, Lubica; Draber, Petr

    2012-01-01

    Migration of mast cells is essential for their recruitment within target tissues where they play an important role in innate and adaptive immune responses. These processes rely on the ability of mast cells to recognize appropriate chemotactic stimuli and react to them by a chemotactic response. Another level of intercellular communication is attained by production of chemoattractants by activated mast cells, which results in accumulation of mast cells and other hematopoietic cells at the sites of inflammation. Mast cells express numerous surface receptors for various ligands with properties of potent chemoattractants. They include the stem cell factor (SCF) recognized by c-Kit, antigen, which binds to immunoglobulin E (IgE) anchored to the high affinity IgE receptor (FcεRI), highly cytokinergic (HC) IgE recognized by FcεRI, lipid mediator sphingosine-1-phosphate (S1P), which binds to G protein-coupled receptors (GPCRs). Other large groups of chemoattractants are eicosanoids [prostaglandin E2 and D2, leukotriene (LT) B4, LTD4, and LTC4, and others] and chemokines (CC, CXC, C, and CX3C), which also bind to various GPCRs. Further noteworthy chemoattractants are isoforms of transforming growth factor (TGF) β1–3, which are sensitively recognized by TGF-β serine/threonine type I and II β receptors, adenosine, C1q, C3a, and C5a components of the complement, 5-hydroxytryptamine, neuroendocrine peptide catestatin, tumor necrosis factor-α, and others. Here we discuss the major types of chemoattractants recognized by mast cells, their target receptors, as well as signaling pathways they utilize. We also briefly deal with methods used for studies of mast cell chemotaxis and with ways of how these studies profited from the results obtained in other cellular systems. PMID:22654878

  11. Expression of uncoupling protein 1 in skeletal muscle decreases muscle energy efficiency and affects thermoregulation and substrate oxidation.

    PubMed

    Klaus, Susanne; Rudolph, Bettina; Dohrmann, Cord; Wehr, Roland

    2005-04-14

    Skeletal muscle uncoupling by ectopic expression of mitochondrial uncoupling protein 1 (UCP1) has been shown to result in a lean phenotype in mice characterized by increased energy expenditure (EE), resistance to diet-induced obesity, and improved glucose tolerance. Here, we investigated in detail the effect of ectopic UCP1 expression in skeletal muscle on thermoregulation and energy homeostasis in HSA-mUCP1 transgenic mice. Thermoneutrality was determined to be approximately 30 degrees C for both wild-type (WT) and transgenic mice. EE, body temperature (Tb), activity, and respiratory quotient (RQ) were then measured over 24 h at ambient temperatures (Ta) of 30, 22, and 5 degrees C. HSA-mUCP1 transgenic mice showed increased activity-related EE and heat loss but similar basal metabolic rate compared with WT. Tb at resting periods was progressively decreased with declining Ta in HSA-mUCP1 transgenic mice but not in WT. Compared with WT littermates, the transgenic HSA-mUCP1 mice displayed increased RQ levels during night time, indicative of increased overall glucose oxidation, and failed to decrease their RQ levels with declining Ta. Thus increased EE caused by skeletal muscle uncoupling is clearly due to a decreased muscle energy efficiency during activity combined with increased glucose oxidation and a compromised thermoregulation associated with increased overall heat loss. At Tas below thermoneutrality, this puts increasing energy demands on the animals, whereas at thermoneutrality most differences in energy metabolism are not apparent any more.

  12. Expression of SPARC like protein 1 (SPARCL1), extracellular matrix-associated protein is down regulated in gastric adenocarcinoma

    PubMed Central

    Jakharia, Aniruddha; Borkakoty, Biswajyoti

    2016-01-01

    Background SPARC-like protein 1 (SPARCL1/Hevin), a member of the SPARC family is defined by the presence of a highly acidic domain-I, a follistatin-like domain, and an extracellular calcium (EC) binding domain. SPARCL1 has been shown to be down-regulated in many types of cancer and may serve as a negative regulator of cell growth and proliferation. Methods Both tumor and adjacent normal tissue were collected from patients with gastric adenocarcinoma. Monoclonal antibody developed against recombinant SPARCL1 was used to analyze the expression of SPARCL1 by immunohisto chemical and western blotting (WB) analysis. Results The expression of SPARCL1 was found to be significantly lower or negligible in gastric adenocarcinoma tissues in nearly all of the cases in comparison with adjacent normal tissue. This comparison was found to be independent of the patient’s age, sex, and stage of cancer. Conclusions We postulate that down regulation of SPARCL1 may be related to inactivation of its tumor suppressor functions and might play an important role in the development of gastric adenocarcinoma. PMID:27034797

  13. Coronavirus nonstructural protein 1: common and distinct functions in the regulation of host and viral gene expression

    PubMed Central

    Narayanan, Krishna; Ramirez, Sydney I.; Lokugamage, Kumari G.; Makino, Shinji

    2014-01-01

    The recent emergence of two highly pathogenic human coronaviruses (CoVs), severe acute respiratory syndrome CoV and Middle East respiratory syndrome CoV, has ignited a strong interest in the identification of viral factors that determine the virulence and pathogenesis of CoVs. The nonstructural protein 1 (nsp1) of CoVs has attracted considerable attention in this regard as a potential virulence factor and a target for CoV vaccine development because of accumulating evidence that point to its role in the downregulation of host innate immune responses to CoV infection. Studies have revealed both functional conservation and mechanistic divergence among the nsp1 of different mammalian CoVs in perturbing host gene expression and antiviral responses. This review summarizes the current knowledge about the biological functions of CoV nsp1 that provides an insight into the novel strategies utilized by this viral protein to modulate host and viral gene expression during CoV infection. PMID:25432065

  14. Visualizing the beta interferon response in mice during infection with influenza A viruses expressing or lacking nonstructural protein 1.

    PubMed

    Kallfass, Carsten; Lienenklaus, Stefan; Weiss, Siegfried; Staeheli, Peter

    2013-06-01

    The innate host defense against influenza virus is largely dependent on the type I interferon (IFN) system. However, surprisingly little is known about the cellular source of IFN in the infected lung. To clarify this question, we employed a reporter mouse that contains the firefly luciferase gene in place of the IFN-β-coding region. IFN-β-producing cells were identified either by simultaneous immunostaining of lungs for luciferase and cellular markers or by generating conditional reporter mice that express luciferase exclusively in defined cell types. Two different strains of influenza A virus were employed that either do or do not code for nonstructural protein 1 (NS1), which strongly suppresses innate immune responses of infected cells. We found that epithelial cells and lung macrophages, which represent the prime host cells for influenza viruses, showed vigorous IFN-β responses which, however, were severely reduced and delayed if the infecting virus was able to produce NS1. Interestingly, CD11c(+) cell populations that were either expressing or lacking macrophage markers produced the bulk of IFN-β at 48 h after infection with wild-type influenza A virus. Our results demonstrate that the virus-encoded IFN-antagonistic factor NS1 disarms specifically epithelial cells and lung macrophages, which otherwise would serve as main mediators of the early response against infection by influenza virus.

  15. Secondary-structure characterization by far-UV CD of highly purified uncoupling protein 1 expressed in yeast.

    PubMed Central

    Douette, Pierre; Navet, Rachel; Bouillenne, Fabrice; Brans, Alain; Sluse-Goffart, Claudine; Matagne, André; Sluse, Francis E

    2004-01-01

    The rat UCP1 (uncoupling protein 1) is a mitochondrial inner-membrane carrier involved in energy dissipation and heat production. We expressed UCP1 carrying a His6 epitope at its C-terminus in Saccharomyces cerevisiae mitochondria. The recombinant-tagged UCP1 was purified by immobilized metal-ion affinity chromatography to homogeneity (>95%). This made it suitable for subsequent biophysical characterization. Fluorescence resonance energy transfer experiments showed that n-dodecyl-beta-D-maltoside-solubilized UCP1-His6 retained its PN (purine nucleotide)-binding capacity. The far-UV CD spectrum of the functional protein clearly indicated the predominance of alpha-helices in the UCP1 secondary structure. The UCP1 secondary structure exhibited an alpha-helical degree of approx. 68%, which is at least 25% higher than the previously reported estimations based on computational predictions. Moreover, the helical content remained unchanged in free and PN-loaded UCP1. A homology model of the first repeat of UCP1, built on the basis of X-ray-solved close parent, the ADP/ATP carrier, strengthened the CD experimental results. Our experimental and computational results indicate that (i) alpha-helices are the major component of UCP1 secondary structure; (ii) PN-binding mechanism does not involve significant secondary-structure rearrangement; and (iii) UCP1 shares similar secondary-structure characteristics with the ADP/ATP carrier, at least for the first repeat. PMID:14766012

  16. von Hippel–Lindau binding protein 1-mediated degradation of integrase affects HIV-1 gene expression at a postintegration step

    PubMed Central

    Mousnier, Aurélie; Kubat, Nicole; Massias-Simon, Aurélie; Ségéral, Emmanuel; Rain, Jean-Christophe; Benarous, Richard; Emiliani, Stéphane; Dargemont, Catherine

    2007-01-01

    HIV-1 integrase, the viral enzyme responsible for provirus integration into the host genome, can be actively degraded by the ubiquitin–proteasome pathway. Here, we identify von Hippel–Lindau binding protein 1(VBP1), a subunit of the prefoldin chaperone, as an integrase cellular binding protein that bridges interaction between integrase and the cullin2 (Cul2)-based von Hippel–Lindau (VHL) ubiquitin ligase. We demonstrate that VBP1 and Cul2/VHL are required for proper HIV-1 expression at a step between integrase-dependent proviral integration into the host genome and transcription of viral genes. Using both an siRNA approach and Cul2/VHL mutant cells, we show that VBP1 and the Cul2/VHL ligase cooperate in the efficient polyubiquitylation of integrase and its subsequent proteasome-mediated degradation. Results presented here support a role for integrase degradation by the prefoldin–VHL–proteasome pathway in the integration–transcription transition of the viral replication cycle. PMID:17698809

  17. The shift from low to high non-structural protein 1 expression in rotavirus-infected MA-104 cells.

    PubMed

    Martínez-Álvarez, Laura; Piña-Vázquez, Carolina; Zarco, Wilbert; Padilla-Noriega, Luis

    2013-06-01

    A hallmark of group/species A rotavirus (RVA) replication in MA-104 cells is the logarithmic increase in viral mRNAs that occurs four-12 h post-infection. Viral protein synthesis typically lags closely behind mRNA synthesis but continues after mRNA levels plateau. However, RVA non-structural protein 1 (NSP1) is present at very low levels throughout viral replication despite showing robust protein synthesis. NSP1 has the contrasting properties of being susceptible to proteasomal degradation, but being stabilised against proteasomal degradation by viral proteins and/or viral mRNAs. We aimed to determine the kinetics of the accumulation and intracellular distribution of NSP1 in MA-104 cells infected with rhesus rotavirus (RRV). NSP1 preferentially localises to the perinuclear region of the cytoplasm of infected cells, forming abundant granules that are heterogeneous in size. Late in infection, large NSP1 granules predominate, coincident with a shift from low to high NSP1 expression levels. Our results indicate that rotavirus NSP1 is a late viral protein in MA-104 cells infected with RRV, presumably as a result of altered protein turnover.

  18. Regulation of Human CYP2C9 Expression by Electrophilic Stress Involves Activator Protein 1 Activation and DNA Looping

    PubMed Central

    Makia, Ngome L.; Surapureddi, Sailesh; Monostory, Katalin; Prough, Russell A.

    2014-01-01

    Cytochrome P450 (CYP)2C9 and CYP2C19 are important human enzymes that metabolize therapeutic drugs, environmental chemicals, and physiologically important endogenous compounds. Initial studies using primary human hepatocytes showed induction of both the CYP2C9 and CYP2C19 genes by tert-butylhydroquinone (tBHQ). As a pro-oxidant, tBHQ regulates the expression of cytoprotective genes by activation of redox-sensing transcription factors, such as the nuclear factor E2-related factor 2 (Nrf2) and members of the activator protein 1 (AP-1) family of proteins. The promoter region of CYP2C9 contains two putative AP-1 sites (TGAGTCA) at positions −2201 and −1930, which are also highly conserved in CYP2C19. The CYP2C9 promoter is activated by ectopic expression of cFos and JunD, whereas Nrf2 had no effect. Using specific kinase inhibitors for mitogen-activated protein kinase, we showed that extracellular signal-regulated kinase and Jun N-terminal kinase are essential for tBHQ-induced expression of CYP2C9. Electrophoretic mobility shift assays demonstrate that cFos distinctly interacts with the distal AP-1 site and JunD with the proximal site. Because cFos regulates target genes as heterodimers with Jun proteins, we hypothesized that DNA looping might be required to bring the distal and proximal AP-1 sites together to activate the CYP2C9 promoter. Chromosome conformation capture analyses confirmed the formation of a DNA loop in the CYP2C9 promoter, possibly allowing interaction between cFos at the distal site and JunD at the proximal site to activate CYP2C9 transcription in response to electrophiles. These results indicate that oxidative stress generated by exposure to electrophilic xenobiotics and metabolites induces the expression of CYP2C9 and CYP2C19 in human hepatocytes. PMID:24830941

  19. Anti-inflammatory actions of Chemoattractant Receptor-homologous molecule expressed on Th2 by the antagonist MK-7246 in a novel rat model of Alternaria alternata elicited pulmonary inflammation.

    PubMed

    Gil, Malgorzata A; Caniga, Michael; Woodhouse, Janice D; Eckman, Joseph; Lee, Hyun-Hee; Salmon, Michael; Naber, John; Hamilton, Valerie T; Sevilla, Raquel S; Bettano, Kimberly; Klappenbach, Joel; Moy, Lily; Correll, Craig C; Gervais, Francois G; Siliphaivanh, Phieng; Zhang, Weisheng; Zhang-Hoover, Jie; McLeod, Robbie L; Cicmil, Milenko

    2014-11-15

    Alternaria alternata is a fungal allergen linked to the development of severe asthma in humans. In view of the clinical relationship between A. alternata and asthma, we sought to investigate the allergic activity of this antigen after direct application to the lungs of Brown Norway rats. Here we demonstrate that a single intratracheal instillation of A. alternata induces dose and time dependent eosinophil influx, edema and Type 2 helper cell cytokine production in the lungs of BN rats. We established the temporal profile of eosinophilic infiltration and cytokine production, such as Interleukin-5 and Interleukin-13, following A. alternata challenge. These responses were comparable to Ovalbumin induced models of asthma and resulted in peak inflammatory responses 48h following a single challenge, eliminating the need for multiple sensitizations and challenges. The initial perivascular and peribronchiolar inflammation preceded alveolar inflammation, progressing to a more sub-acute inflammatory response with notable epithelial cell hypertrophy. To limit the effects of an A. alternata inflammatory response, MK-7246 was utilized as it is an antagonist for Chemoattractant Receptor-homologous molecule expressed in Th2 cells. In a dose-dependent manner, MK-7246 decreased eosinophil influx and Th2 cytokine production following the A. alternata challenge. Furthermore, therapeutic administration of corticosteroids resulted in a dose-dependent decrease in eosinophil influx and Th2 cytokine production. Reproducible asthma-related outcomes and amenability to pharmacological intervention by mechanisms relevant to asthma demonstrate that an A. alternata induced pulmonary inflammation in BN rats is a valuable preclinical pharmacodynamic in vivo model for evaluating the pharmacological inhibitors of allergic pulmonary inflammation.

  20. Cellular retinol-binding protein-1 is transiently expressed in granulation tissue fibroblasts and differentially expressed in fibroblasts cultured from different organs.

    PubMed Central

    Xu, G.; Redard, M.; Gabbiani, G.; Neuville, P.

    1997-01-01

    We have reported that cellular retinol-binding protein-1 (CRBP-1) is transiently expressed by arterial smooth muscle cells during experimental intimal repair (P. Neuville, A. Geinoz, G. Benzonana, M. Redard, F. Gabbiani, P. Ropraz, G. Gabbiani: Am J Pathol 1997, 150:509-521). We have examined here the expression of CRBP-1 during wound healing after a full-thickness rat skin wound. CRBP-1 was transiently expressed by a significant proportion of fibroblastic cells including myofibroblasts. Expression started 4 days after wounding, reached a maximum at 12 days, and persisted up to 30 days when a scar was formed. After wound closure, most CRBP-1-containing fibroblastic cells underwent apoptosis. We have further investigated CRBP-1 expression in rat fibroblasts cultured from different organs. CRBP-1 was abundant in lung and heart fibroblasts and was detected in decreasing amounts in muscle, tendon, subcutaneous tissue, and granulation tissue fibroblasts. Dermis fibroblasts contained no detectable levels of CRBP-1. All-trans retinoic acid and transforming growth factor-beta1 inhibited cell proliferation and increased CRBP-1 expression in fibroblastic populations except dermis fibroblasts. We demonstrate that during granulation tissue formation a subpopulation of fibroblastic cells express CRBP-1 de novo. We also demonstrate that CRBP-1 expression by fibroblasts is regulated in vitro by retinoic acid and transforming growth factor-beta1. Our results suggest that CRBP-1 and possibly retinoic acid play a role in the evolution of granulation tissue. Images Figure 1 Figure 2 Figure 3 Figure 4 Figure 7 PMID:9403724

  1. Monocyte chemotactic protein-1 expression as a prognosic biomarker in patients with solid tumor: a meta analysis

    PubMed Central

    Wang, Hong; Zhang, Qiongwen; Kong, Hongyu; Zeng, Yunhui; Hao, Meiqin; Yu, Ting; Peng, Jing; Xu, Zhao; Chen, Jingquan; Shi, Huashan

    2014-01-01

    Purpose: A great deal of studies have been performed on the prognostic value of monocyte chemotactic protein-1 (MCP-1) in solid tumors in recent years. However, no consistent outcomes are reported. Therefore, the prognostic value of MCP-1 still remains controversial in patients with solid tumors. Here we aimed to evaluate the prognostic value of MCP-1 expression for patients with solid tumors. Methods: Comprehensive literature was selected from PUBMED and EMBASE and clinical studies which reported analysis of survival data about MCP-1 in solid tumors were included. Stata 11.0 was used for performing a meta-analysis on evaluating the relation between MCP-1 and clinical staging, overall survival (OS) and disease free survival (DFS). Results: Eleven studies with a total of 1324 patients with solid tumors were included into our meta-analysis. The result showed that high concentration of MCP-1 was related to a worse OS (HR = 1.95, 95% CI 1.32-2.88). The subgroup analysis on different location of tumors showed that high concentration of MCP-1 meant bad prognosis in patients with digestive cancer (HR = 2.66, 95% CI 1.44-4.91) and urogenital cancer (HR = 2.23, 95% CI 1.61-3.10), even head and neck cancer (HR = 1.99, 95% CI 0.95-4.18) other than respiratory cancer (HR = 1.10, 95% CI 0.39-3.11). Another subgroup analysed on different sites of cancer and indicated a poor prognosis on adenocarcinoma (HR = 2.10, 95% CI 1.63-2.69). Conclusions: Our findings suggest that MCP-1 can be regarded as a poor prognostic maker for solid tumors and may represent important new therapeutic targets. PMID:25120764

  2. A Chemoattractant-mediated Gi-coupled Pathway Activates Adenylyl Cyclase in Human Neutrophils

    PubMed Central

    Mahadeo, Dana C.; Janka-Junttila, Mirkka; Smoot, Rory L.; Roselova, Pavla

    2007-01-01

    Neutrophils and Dictyostelium use conserved signal transduction pathways to decipher chemoattractant gradients and migrate directionally. In both cell types, addition of chemoattractants stimulates the production of cAMP, which has been suggested to regulate chemotaxis. We set out to define the mechanism by which chemoattractants increase cAMP levels in human neutrophils. We show that chemoattractants elicit a rapid and transient activation of adenylyl cyclase (AC). This activation is sensitive to pertussis toxin treatment but independent of phosphoinositide-3 kinase activity and an intact cytoskeleton. Remarkably, and in sharp contrast to Gαs-mediated activation, chemoattractant-induced AC activation is lost in cell lysates. Of the nine, differentially regulated transmembrane AC isoforms in the human genome, we find that isoforms III, IV, VII, and IX are expressed in human neutrophils. We conclude that the signal transduction cascade used by chemoattractants to activate AC is conserved in Dictyostelium and human neutrophils and is markedly different from the canonical Gαs-meditated pathway. PMID:17135293

  3. Hydrogen peroxide activates activator protein-1 and mitogen-activated protein kinases in pancreatic stellate cells.

    PubMed

    Kikuta, Kazuhiro; Masamune, Atsushi; Satoh, Masahiro; Suzuki, Noriaki; Satoh, Kennichi; Shimosegawa, Tooru

    2006-10-01

    Activated pancreatic stellate cells (PSCs) are implicated in the pathogenesis of pancreatic inflammation and fibrosis, where oxidative stress is thought to play a key role. Reactive oxygen species such as hydrogen peroxide (H(2)O(2)) may act as a second messenger to mediate the actions of growth factors and cytokines. But the role of reactive oxygen species in the activation and regulation of cell functions in PSCs remains largely unknown. We here examined the effects of H(2)O(2) on the activation of signal transduction pathways and cell functions in PSCs. PSCs were isolated from the pancreas of male Wistar rats, and used in their culture-activated, myofibroblast-like phenotype unless otherwise stated. Activation of transcription factors was examined by electrophoretic mobility shift assay and luciferase assay. Activation of mitogen-activated protein (MAP) kinases was assessed by Western blotting using anti-phosphospecific antibodies. The effects of H(2)O(2) on proliferation, alpha(1)(I)procollagen gene expression, and monocyte chemoattractant protein-1 production were evaluated. The effect of H(2)O(2) on the transformation of freshly isolated PSCs in culture was also assessed. H(2)O(2) at non-cytotoxic concentrations (up to 100 microM) induced oxidative stress in PSCs. H(2)O(2) activated activator protein-1, but not nuclear factor kappaB. In addition, H(2)O(2) activated three classes of MAP kinases: extracellular signal-regulated kinase, c-Jun N-terminal kinase, and p38 MAP kinase. H(2)O(2) induced alpha(1)(I)procollagen gene expression but did not induce proliferation or monocyte chemoattractant protein-1 production. H(2)O(2) did not initiate the transformation of freshly isolated PSCs to myofibroblast-like phenotype. Specific activation of these signal transduction pathways and collagen gene expression by H(2)O(2) may play a role in the pathogenesis of pancreatic fibrosis.

  4. Expression of uncoupling proteins-1, -2 and -3 mRNA is induced by an adenocarcinoma-derived lipid-mobilizing factor

    PubMed Central

    Bing, C; Russell, S T; Beckett, E E; Collins, P; Taylor, S; Barraclough, R; Tisdale, M J; Williams, G

    2002-01-01

    The abnormalities of lipid metabolism observed in cancer cachexia may be induced by a lipid-mobilizing factor produced by adenocarcinomas. The specific molecules and metabolic pathways that mediate the actions of lipid-mobilizing factor are not known. The mitochondrial uncoupling proteins-1, -2 and -3 are suggested to play essential roles in energy dissipation and disposal of excess lipid. Here, we studied the effects of lipid-mobilizing factor on the expression of uncoupling proteins-1, -2 and -3 in normal mice. Lipid-mobilizing factor isolated from the urine of cancer patients was injected intravenously into mice over a 52-h period, while vehicle was similarly given to controls. Lipid-mobilizing factor caused significant reductions in body weight (−10%, P=0.03) and fat mass (−20%, P<0.01) accompanied by a marked decrease in plasma leptin (−59%, P<0.01) and heavy lipid deposition in the liver. In brown adipose tissue, uncoupling protein-1 mRNA levels were elevated in lipid-mobilizing factor-treated mice (+96%, P<0.01), as were uncoupling proteins-2 and -3 (+57% and +37%, both P<0.05). Lipid-mobilizing factor increased uncoupling protein-2 mRNA in both skeletal muscle (+146%, P<0.05) and liver (+142%, P=0.03). The protein levels of uncoupling protein-1 in brown adipose tissue and uncoupling protein-2 in liver were also increased with lipid-mobilizing factor administration (+49% and +67%, both P=0.02). Upregulation by lipid-mobilizing factor of uncoupling proteins-1, -2 and -3 in brown adipose tissue, and of uncoupling protein-2 in skeletal muscle and liver, suggests that these uncoupling proteins may serve to utilize excess lipid mobilized during fat catabolism in cancer cachexia. British Journal of Cancer (2002) 86, 612–618. DOI: 10.1038/sj/bjc/6600101 www.bjcancer.com © 2002 Cancer Research UK PMID:11870545

  5. Low Density Lipoprotein-Receptor Related Protein 1 Is Differentially Expressed by Neuronal and Glial Populations in the Developing and Mature Mouse Central Nervous System

    PubMed Central

    Auderset, Loic; Cullen, Carlie L.; Young, Kaylene M.

    2016-01-01

    The low density lipoprotein-receptor related protein 1 (LRP1) is a large endocytic cell surface receptor that is known to interact with a variety of ligands, intracellular adaptor proteins and other cell surface receptors to regulate cellular behaviours ranging from proliferation to cell fate specification, migration, axon guidance, and lipid metabolism. A number of studies have demonstrated that LRP1 is expressed in the brain, yet it is unclear which central nervous system cell types express LRP1 during development and in adulthood. Herein we undertake a detailed study of LRP1 expression within the mouse brain and spinal cord, examining a number of developmental stages ranging from embryonic day 13.5 to postnatal day 60. We report that LRP1 expression in the brain peaks during postnatal development. On a cellular level, LRP1 is expressed by radial glia, neuroblasts, microglia, oligodendrocyte progenitor cells (OPCs), astrocytes and neurons, with the exception of parvalbumin+ interneurons in the cortex. Most cell populations exhibit stable expression of LRP1 throughout development; however, the proportion of OPCs that express LRP1 increases significantly from ~69% at E15.5 to ~99% in adulthood. We also report that LRP1 expression is rapidly lost as OPCs differentiate, and is absent from all oligodendrocytes, including newborn oligodendrocytes. While LRP1 function has been primarily examined in mature neurons, these expression data suggest it plays a more critical role in glial cell regulation–where expression levels are much higher. PMID:27280679

  6. Distal Interleukin-1β (IL-1β) Response Element of Human Matrix Metalloproteinase-13 (MMP-13) Binds Activator Protein 1 (AP-1) Transcription Factors and Regulates Gene Expression*

    PubMed Central

    Schmucker, Adam C.; Wright, Jason B.; Cole, Michael D.; Brinckerhoff, Constance E.

    2012-01-01

    The collagenase matrix metalloproteinase-13 (MMP-13) plays an important role in the destruction of cartilage in arthritic joints. MMP-13 expression is strongly up-regulated in arthritis, largely because of stimulation by inflammatory cytokines such as IL-1β. Treatment of chondrocytes with IL-1β induces transcription of MMP-13 in vitro. IL-1β signaling converges upon the activator protein-1 transcription factors, which have been shown to be required for IL-1β-induced MMP-13 gene expression. Using chromatin immunoprecipitation (ChIP), we detected activator protein-1 binding within an evolutionarily conserved DNA sequence ∼20 kb 5′ relative to the MMP-13 transcription start site (TSS). Also using ChIP, we detected histone modifications and binding of RNA polymerase II within this conserved region, all of which are consistent with transcriptional activation. Chromosome conformation capture indicates that chromosome looping brings this region in close proximity with the MMP-13 TSS. Finally, a luciferase reporter construct driven by a component of the conserved region demonstrated an expression pattern similar to that of endogenous MMP-13. These data suggest that a conserved region at 20 kb upstream from the MMP-13 TSS includes a distal transcriptional response element of MMP-13, which contributes to MMP-13 gene expression. PMID:22102411

  7. Selenium-binding protein 1 in head and neck cancer is low-expression and associates with the prognosis of nasopharyngeal carcinoma

    PubMed Central

    Chen, Fasheng; Chen, Chen; Qu, Yangang; Xiang, Hua; Ai, Qingxiu; Yang, Fei; Tan, Xueping; Zhou, Yi; Jiang, Guang; Zhang, Zixiong

    2016-01-01

    Abstract Background: Selenium-binding protein 1 (SELENBP1) expression is reduced markedly in many types of cancers and low SELENBP1 expression levels are associated with poor patient prognosis. Methods: SELENBP1 gene expression in head and neck squamous cell carcinoma (HNSCC) was analyzed with GEO dataset and characteristics of SELENBP1 expression in paraffin embedded tissue were summarized. Expression of SELENBP1 in nasopharyngeal carcinoma (NPC), laryngeal cancer, oral cancer, tonsil cancer, hypopharyngeal cancer and normal tissues were detected using immunohistochemistry, at last, 99 NPC patients were followed up more than 5 years and were analyzed the prognostic significance of SELENBP1. Results: Analysis of GEO dataset concluded that SELENBP1 gene expression in HNSCC was lower than that in normal tissue (P < 0.01), but there was no significant difference of SELENBP1 gene expression in different T-stage and N-stage (P > 0.05). Analysis of pathological section concluded that SELENBP1 in the majority of HNSCC is low expression and in cancer nests is lower expression than surrounding normal tissue, even associated with the malignant degree of tumor. Further study indicated the low SELENBP1 expression group of patients with NPC accompanied by poor overall survival and has significantly different comparing with the high expression group. Conclusion: SELENBP1 expression was down-regulated in HNSCC, but has no associated with T-stage and N-stage of tumor. Low expression of SELENBP1 in patients with NPC has poor over survival, so SELENBP1 could be a novel biomarker for predicting prognosis. PMID:27583873

  8. Extracellular galectin-3 induces MMP9 expression by activating p38 MAPK pathway via lysosome-associated membrane protein-1 (LAMP1).

    PubMed

    Dange, Manohar C; Agarwal, Akhil Kumar; Kalraiya, Rajiv D

    2015-06-01

    Matrix metalloproteinases (MMPs) play a key role in matrix remodelling and thus invasion and metastasis. Extracellular galectin-3 has been shown to induce MMP9 secretion. Here, we demonstrate that galectin-3 induces MMP9 at transcript level and it is dependent on the surface levels of poly-N-acetyllactosamine (polyLacNAc). By employing signalling pathway inhibitors, MMP9 expression was shown to be induced via p38 MAP-kinase pathway. Using clones of melanoma cells expressing shRNAs to lysosome-associated membrane protein-1 (LAMP1), a major carrier of polyLacNAc, surface LAMP1 was demonstrated to serve as one of the key mediators of galectin-3-induced MMP9 expression via p38 MAPK pathway.

  9. Klf10 regulates odontoblast differentiation and mineralization via promoting expression of dentin matrix protein 1 and dentin sialophosphoprotein genes

    PubMed Central

    Chen, Zhuo; Li, Wentong; Wang, Han; Wan, Chunyan; Luo, Daoshu; Deng, Shuli

    2016-01-01

    Klf10, a member of the Krüppel-like family of transcription factors, is critical for osteoblast differentiation, bone formation and mineralization. However, whether Klf10 is involved in odontoblastic differentiation and tooth development has not been determined. In this study, we investigate the expression patterns of Klf10 during murine tooth development in vivo and its role in odontoblastic differentiation in vitro. Klf10 protein was expressed in the enamel organ and the underlying mesenchyme, ameloblasts and odontoblasts at early and later stages of murine molar formation. Furthermore, the expression of Klf10, Dmp1, Dspp and Runx2 was significantly elevated during the process of mouse dental papilla mesenchymal differentiation and mineralization. The overexpression of Klf10 induced dental papilla mesenchymal cell differentiation and mineralization as detected by alkaline phosphatase staining and alizarin red S assay. Klf10 additionally up-regulated the expression of odontoblastic differentiation marker genes Dmp1, Dspp and Runx2 in mouse dental papilla mesenchymal cells. The molecular mechanism of Klf10 in controlling Dmp1 and Dspp expression is thus to activate their regulatory regions in a dosage-dependent manner. Our results suggest that Klf10 is involved in tooth development and promotes odontoblastic differentiation via the up-regulation of Dmp1 and Dspp transcription. PMID:26310138

  10. Osteoblast-derived WNT-induced secreted protein 1 increases VCAM-1 expression and enhances prostate cancer metastasis by down-regulating miR-126.

    PubMed

    Tai, Huai-Ching; Chang, An-Chen; Yu, Hong-Jeng; Huang, Chao-Yuan; Tsai, Yu-Chieh; Lai, Yu-Wei; Sun, Hui-Lung; Tang, Chih-Hsin; Wang, Shih-Wei

    2014-09-15

    Bone metastases of prostate cancer (PCa) may cause intractable pain. Wnt-1-induced secreted protein 1 (WISP-1) belongs to the CCN family (CTGF/CYR61/NOV) that plays a key role in bone formation. We found that osteoblast-conditioned medium (OBCM) stimulates migration and vascular adhesion molecule-1 (VCAM)-1 expression in human PCa (PC3 and DU145) cells. Osteoblast transfection with WISP-1 shRNA reduced OBCM-mediated PCa migration and VCAM-1 expression. Stimulation of PCa with OBCM or WISP-1 elevated focal adhesion kinase (FAK) and p38 phosphorylation. Either FAK and p38 inhibitors or siRNA abolished osteoblast-derived WISP-1-induced migration and VCAM-1 expression. Osteoblast-derived WISP-1 inhibited miR-126 expression. Moreover, miR-216 mimic reversed the WISP-1-enhanced migration and VCAM-1 expression. This study suggests that osteoblast-derived WISP-1 promotes migration and VCAM-1 expression in human PCa cells by down-regulating miR-126 expression via αvβ1 integrin, FAK, and p38 signaling pathways. Thus, WISP-1 may be a new molecular therapeutic target in PCa bone metastasis.

  11. Corticotropin-releasing hormone stimulates mitotic kinesin-like protein 1 expression via a PLC/PKC-dependent signaling pathway in hippocampal neurons.

    PubMed

    Sheng, Hui; Xu, Yongjun; Chen, Yanming; Zhang, Yanmin; Ni, Xin

    2012-10-15

    Corticotropin-releasing hormone (CRH) has been shown to modulate dendritic development in hippocampus. Mitotic kinesin-like protein 1 (MKLP1) plays key roles in dendritic differentiation. In the present study, we examined the effects of CRH on MKLP1 expression in cultured hippocampal neurons and determine subsequent signaling pathways involved. CRH dose-dependently increased MKLP1 mRNA and protein expression. This effect can be reversed by CRHR1 antagonist but not by CRHR2 antagonist. CRHR1 knockdown impaired this effect of CRH. CRH stimulated GTP-bound Gαs protein and phosphorylated phospholipase C (PLC)-β3 expression, which were blocked by CRHR1 antagonist. Transfection of GP antagonist-2A, an inhibitory peptide of Gαq protein, blocked CRH-induced phosphorylated PLC-β3 expression. PLC and PKC inhibitors completely blocked whereas adenylyl cyclase (AC) and PKA inhibitors did not affect CRH-induced MKLP1 expression. Our results indicate that CRH act on CRHR1 to induce MKLP1 expression via PLC/PKC signaling pathway. CRH may regulate MKLP1 expression, thereby modulating dendritic development.

  12. Momilactione B inhibits protein kinase A signaling and reduces tyrosinase-related proteins 1 and 2 expression in melanocytes.

    PubMed

    Lee, Ji Hae; Cho, Boram; Jun, Hee-jin; Seo, Woo-Duck; Kim, Dong-Woo; Cho, Kang-Jin; Lee, Sung-Joon

    2012-05-01

    Momilactone B (MB) is a terpenoid phytoalexin present in rice bran that exhibits several biological activities. MB reduced the melanin content in B16 melanocytes melanin content and inhibited tyrosinase activities. Using transcriptome analysis, the genes involved in protein kinase A (PKA) signaling were found to be markedly altered. B16 cells stimulated with MB had decreased concentrations of cAMP protein kinase A activity, and cAMP-response element-binding protein which is a key transcription factor for microphthalmia-associated transcription factor (MITF) expression. Accordingly, the expression of MITF and its target genes, which are essential for melanogenesis, were reduced. MB thus exhibits anti-melanogenic effects by repressing tyrosinase enzyme activity and inhibiting the PKA signaling pathway which, in turn, decreases melanogenic gene expression.

  13. Hepatic PCSK9 expression is regulated by nutritional status via insulin and sterol regulatory element-binding protein 1c.

    PubMed

    Costet, Philippe; Cariou, Bertrand; Lambert, Gilles; Lalanne, Florent; Lardeux, Bernard; Jarnoux, Anne-Laure; Grefhorst, Aldo; Staels, Bart; Krempf, Michel

    2006-03-10

    Familial autosomal dominant hypercholesterolemia is associated with high risk for cardiovascular accidents and is related to mutations in the low density lipoprotein receptor or its ligand apolipoprotein B (apoB). Mutations in a third gene, proprotein convertase subtilisin kexin 9 (PCSK9), were recently associated to this disease. PCSK9 acts as a natural inhibitor of the low density lipoprotein receptor pathway, and both genes are regulated by depletion of cholesterol cell content and statins, via sterol regulatory element-binding protein (SREBP). Here we investigated the regulation of PCSK9 gene expression during nutritional changes. We showed that PCSK9 mRNA quantity is decreased by 73% in mice after 24 h of fasting, leading to a 2-fold decrease in protein level. In contrast PCSK9 expression was restored upon high carbohydrate refeeding. PCSK9 mRNA increased by 4-5-fold in presence of insulin in rodent primary hepatocytes, whereas glucose had no effect. Moreover, insulin up-regulated hepatic PCSK9 expression in vivo during a hyperinsulinemic-euglycemic clamp in mice. Adenoviral mediated overexpression of a dominant or negative form of SREBP-1c confirmed the implication of this transcription factor in insulin-mediated stimulation of PCSK9 expression. Liver X receptor agonist T0901317 also regulated PCSK9 expression via this same pathway (a 2-fold increase in PCSK9 mRNA of primary hepatocytes cultured for 24 h in presence of 1 microm T0901317). As our last investigation, we isolated PCSK9 proximal promoter and verified the functionality of a SREBP-1c responsive element located from 335 bp to 355 bp upstream of the ATG. Together, these results show that PCSK9 expression is regulated by nutritional status and insulinemia. PMID:16407292

  14. Nitric oxide increases Wnt-induced secreted protein-1 (WISP-1/CCN4) expression and function in colitis.

    PubMed

    Wang, Hongying; Zhang, Rui; Wen, Shoubin; McCafferty, Donna-Marie; Beck, Paul L; MacNaughton, Wallace K

    2009-04-01

    Nitric oxide (NO) derived from the inducible NO synthase (iNOS) is an important and complex mediator of inflammation in the intestine. Wnt-inducible secreted protein (WISP)-1 (CCN4), a member of the connective tissue growth factor family, is involved in tissue repair. We sought to determine the relationship between iNOS and WISP-1 in colitis. By analyzing human colonic biopsy samples, we showed that the expression of mRNA for both iNOS and WISP-1 was significantly higher in ulcerative colitis samples compared with control tissue. The upregulation of WISP-1 was positively correlated with iNOS expression in two models of colitis, induced by intrarectal trinitrobenzenesulfonic acid (TNBS) or occurring spontaneously in IL-10 deficient mice. Loss of iNOS, studied using iNOS(-/-) mice in both TNBS-induced and IL-10(-/-) colitis models, significantly attenuated the colitis-related WISP-1 increase. In human colonic epithelial cell lines, the NO donor, DETA-NONOate, elevated WISP-1 mRNA and protein expression through a beta-catenin and CREB-dependent, but Wnt-1-independent, pathway. In addition, NO-induced WISP-1 directly induced secretion of soluble collagen in colonic fibroblast cells. NO increases WISP-1 expression both in vitro and in vivo, suggesting a new role for iNOS and NO in colitis.

  15. Expression of Iron Regulatory Protein 1 Is Regulated not only by HIF-1 but also pCREB under Hypoxia

    PubMed Central

    Luo, Qian-Qian; Qian, Zhong-Ming; Zhou, Yu-Fu; Zhang, Meng-Wan; Wang, Dang; Zhu, Li; Ke, Ya

    2016-01-01

    The inconsistent of responses of IRP1 and HIF-1 alpha to hypoxia and the similar tendencies in the changes of IRP1 and pCREB contents led us to hypothesize that pCREB might be involved in the regulation of IRP1 under hypoxia. Here, we investigated the role of pCREB in IRP1 expression in HepG2 cells under hypoxia using quantitative PCR, western blot, immunofluorescence, electrophoretic mobility shift assay (EMSA) and chromatin immunoprecipitation (ChIP). We demonstrated that 1) Hypoxia increased pCREB levels inside of the nucleus; 2) Putative CREs were found in the IRP1 gene; 3) Nuclear extracts of HepG2 cells treated with hypoxia could bind to CRE1 and CRE3, and 100-fold competitor of putative CREs could abolish the binding activity to varying degrees; 4) pCREB was found in the CRE1 and CRE3 DNA-protein complexes of EMSA; 5) CRE1 and CRE3 binding activity of IRP1 depended on CREB activation but not on HIF-1; 6) Increased IRP1 expression under hypoxia could be prevented by LY294002; 7) ChIP assays demonstrated that pCREB binds to IRP1 promoter; and 8) HIF-1 and/or HIF-2 siRNA had no effect on the expression of pCREB and IRP1 proteins in cells treated with hypoxia for 8 hours. Our findings evidenced for the involvement of pCREB in IRP1 expression and revealed a dominant role of PI3K/Akt pathway in CREB activation under hypoxia and also suggested that dual-regulation of IRP1 expression by HIF-1 and pCERB or other transcription factor(s) under hypoxia might be a common mechanism in most if not all of hypoxia-inducible genes. PMID:27766034

  16. Serum amyloid A chemoattracts immature dendritic cells and indirectly provokes monocyte chemotaxis by induction of cooperating CC and CXC chemokines.

    PubMed

    Gouwy, Mieke; De Buck, Mieke; Pörtner, Noëmie; Opdenakker, Ghislain; Proost, Paul; Struyf, Sofie; Van Damme, Jo

    2015-01-01

    Serum amyloid A (SAA) is an acute phase protein that is upregulated in inflammatory diseases and chemoattracts monocytes, lymphocytes, and granulocytes via its G protein-coupled receptor formyl peptide receptor like 1/formyl peptide receptor 2 (FPRL1/FPR2). Here, we demonstrated that the SAA1α isoform also chemoattracts monocyte-derived immature dendritic cells (DCs) in the Boyden and μ-slide chemotaxis assay and that its chemotactic activity for monocytes and DCs was indirectly mediated via rapid chemokine induction. Indeed, SAA1 induced significant amounts (≥5 ng/mL) of macrophage inflammatory protein-1α/CC chemokine ligand 3 (MIP-1α/CCL3) and interleukin-8/CXC chemokine ligand 8 (IL-8/CXCL8) in monocytes and DCs in a dose-dependent manner within 3 h. However, SAA1 also directly activated monocytes and DCs for signaling and chemotaxis without chemokine interference. SAA1-induced monocyte migration was nevertheless significantly prevented (60-80% inhibition) in the constant presence of desensitizing exogenous MIP-1α/CCL3, neutralizing anti-MIP-1α/CCL3 antibody, or a combination of CC chemokine receptor 1 (CCR1) and CCR5 antagonists, indicating that this endogenously produced CC chemokine was indirectly contributing to SAA1-mediated chemotaxis. Further, anti-IL-8/CXCL8 antibody neutralized SAA1-induced monocyte migration, suggesting that endogenous IL-8/CXCL8 acted in concert with MIP-1α/CCL3. This explained why SAA1 failed to synergize with exogenously added MIP-1α/CCL3 or stromal cell-derived factor-1α (SDF-1α)/CXCL12 in monocyte and DC chemotaxis. In addition to direct leukocyte activation, SAA1 induces a chemotactic cascade mediated by expression of cooperating chemokines to prolong leukocyte recruitment to the inflammatory site.

  17. Caveolin-1 Increases Proinflammatory Chemoattractants and Blood–Retinal Barrier Breakdown but Decreases Leukocyte Recruitment in Inflammation

    PubMed Central

    Li, Xiaoman; Gu, Xiaowu; Boyce, Timothy M.; Zheng, Min; Reagan, Alaina M.; Qi, Hui; Mandal, Nawajes; Cohen, Alex W.; Callegan, Michelle C.; Carr, Daniel J. J.; Elliott, Michael H.

    2014-01-01

    Purpose. Caveolin-1 (Cav-1), the signature protein of caveolae, modulates inflammatory responses, and innate immunity. However, Cav-1′s role in retinal inflammation has not been rigorously tested. In this study, we examined the effect of Cav-1 ablation on the sensitivity of the retina to inflammation. Methods. Cav-1 knockout (KO) mice were challenged by intravitreal injection of lipopolysaccharide (LPS) and inflammatory cell recruitment was assessed by flow cytometry and immunohistochemistry. Leukostasis was assessed in retinal flatmounts after perfusion with FITC-labeled Concanavalin A (FITC-ConA). Chemoattractants were measured by multiplex immunoassays. Blood–retinal barrier (BRB) breakdown was assessed quantitatively by a FITC-dextran permeability assay. The ratio of extravascular to total immune cells was determined by CD45 immunohistochemistry of retinal flatmounts. Results. Inflammatory challenge resulted in significant blunting of proinflammatory cytokine (monocyte chemoattractant protein-1 [MCP-1/CCL2], CXCL1/KC, IL-6, and IL-1β) responses as well as reduced inflammatory BRB breakdown in Cav-1 KO retinas. Paradoxically, Cav-1 deficiency resulted in significantly increased recruitment of immune cells compared with controls as well as increased leukostasis. A similar ratio of extravascular/total leukocytes were found in Cav-1 KO and wild-type (WT) retinas suggesting that Cav-1 deficient leukocytes were as competent to extravasate as those from WT mice. We found increased levels of circulating immune cells in naïve (not challenged with LPS) Cav-1 KO mice compared with controls. Conclusions. Caveolin-1 paradoxically modulates inflammatory signaling and leukocyte infiltration through distinct mechanisms. We hypothesize that Cav-1 expression may enhance inflammatory signaling while at the same time supporting the physical properties of the BRB. PMID:25159208

  18. Controlled Pseudopod Extension of Human Neutrophils Stimulated with Different Chemoattractants

    PubMed Central

    Zhelev, Doncho V.; Alteraifi, Abdullatif M.; Chodniewicz, David

    2004-01-01

    The formation of pseudopods and lamellae after ligation of chemoattractant sensitive G-protein coupled receptors (GPCRs) is essential for chemotaxis. Here, pseudopod extension was stimulated with chemoattractant delivered from a micropipet. The chemoattractant diffusion and convection mass transport were considered, and it is shown that when the delivery of chemoattractant was limited by diffusion there was a strong chemoattractant gradient along the cell surface. The diffusion-limited delivery of chemoattractant from a micropipet allowed for maintaining an almost constant chemoattractant concentration at the leading edge of single pseudopods during their growth. In these conditions, the rate of pseudopod extension was dependent on the concentration of chemoattractant in the pipet delivering chemoattractant. The pseudopod extension induced using micropipets was oscillatory even in the presence of a constant delivery of chemoattractant. This oscillatory pseudopod extension was controlled by activated RhoA and its downstream effector kinase ROCK and was abolished after the inhibition of RhoA activation with Clostridium botulinium C3 exoenzyme (C3) or the blocking of ROCK activation with Y-27632. The ability of the micropipet assay to establish a well-defined chemoattractant distribution around the activated cell over a wide range of molecular weights of the used chemoattractants allowed for comparison of the effect of chemoattractant stimulation on the dynamics of pseudopod growth. Pseudopod growth was stimulated using N-formylated peptide (N-formyl-methionyl-leucyl-phenylalanine (fMLP)), platelet activating factor (PAF), leukotriene B4 (LTB4), C5a anaphylotoxin (C5a), and interleukin-8 (IL-8), which represent the typical ligands for G-protein coupled chemotactic receptors. The dependence of the rate of pseudopod extension on the concentration of these chemoattractants and their equimolar mixture was measured and shown to be similar for all chemoattractants. The

  19. Regulation of Peroxisome Proliferator-Activated Receptor γ Expression by Adipocyte Differentiation and Determination Factor 1/Sterol Regulatory Element Binding Protein 1: Implications for Adipocyte Differentiation and Metabolism

    PubMed Central

    Fajas, Lluis; Schoonjans, Kristina; Gelman, Laurent; Kim, Jae B.; Najib, Jamila; Martin, Genevieve; Fruchart, Jean-Charles; Briggs, Michael; Spiegelman, Bruce M.; Auwerx, Johan

    1999-01-01

    Peroxisome proliferator-activated receptor γ (PPARγ) is a nuclear receptor implicated in adipocyte differentiation and insulin sensitivity. We investigated whether PPARγ expression is dependent on the activity of adipocyte differentiation and determination factor 1/sterol regulatory element binding protein 1 (ADD-1/SREBP-1), another transcription factor associated with both adipocyte differentiation and cholesterol homeostasis. Ectopic expression of ADD-1/SREBP-1 in 3T3-L1 and HepG2 cells induced endogenous PPARγ mRNA levels. The related transcription factor SREBP-2 likewise induced PPARγ expression. In addition, cholesterol depletion, a condition known to result in proteolytic activation of transcription factors of the SREBP family, induced PPARγ expression and improved PPRE-driven transcription. The effect of the SREBPs on PPARγ expression was mediated through the PPARγ1 and -3 promoters. Both promoters contain a consensus E-box motif that mediates the regulation of the PPARγ gene by ADD-1/SREBP-1 and SREBP-2. These results suggest that PPARγ expression can be controlled by the SREBP family of transcription factors and demonstrate new interactions between transcription factors that can regulate different pathways of lipid metabolism. PMID:10409739

  20. A genetic polymorphism affects the risk and prognosis of renal cell carcinoma: association with follistatin-like protein 1 expression

    PubMed Central

    Liu, Yan; Han, Xue; Yu, Yongwei; Ding, Yibo; Ni, Chong; Liu, Wenbin; Hou, Xiaomei; Li, Zixiong; Hou, Jianguo; Shen, Dan; Yin, Jianhua; Zhang, Hongwei; Thompson, Timothy C.; Tan, Xiaojie; Cao, Guangwen

    2016-01-01

    Few single nucleotide polymorphisms (SNPs) associated with the risk of renal cell carcinoma (RCC) have been identified, yet genetic predisposition contributes significantly to this malignancy. We previously showed that follistatin-like 1 (FSTL1) was significantly down-regulated in clear cell RCC (ccRCC), in particular metastatic ccRCC. In the present study, we systemically investigated the associations of the 6 SNPs within FSTL1-coding genomic region with RCC risk and postoperative prognosis. Age- and gender-matched case-control study (417 vs 855) indicated that rs1259293 variant genotype CC was significantly associated with an increased risk of RCC, with an odds ratio of 2.004 (95% confidence internal [CI] = 1.190–3.375). Multivariate Cox regression analysis in 309 of 417 cases showed that rs1259293 genotype (CC vs TT + CT) independently predicted an unfavorable prognosis, with a hazard ratio of 2.531 (95% CI = 1.052–6.086). Expression of FSTL1 was significantly higher in adjacent renal tissues than in tumors, and significantly higher in the tissues with rs1259293 TT genotype than in those with rs1259293 TC+CC genotypes. rs1259293 C allele might generate a CTCF binding site that blocks trans-activation of FSTL1 expression. Our results indicate that rs1259293 is associated with an increased risk and unfavorable postoperative prognosis of RCC, possibly by down-regulating FSTL1 expression in renal tissues. PMID:27225192

  1. Glutamate regulates kainate-binding protein expression in cultured chick Bergmann glia through an activator protein-1 binding site.

    PubMed

    Aguirre, A; López, T; López-Bayghen, E; Ortega, A

    2000-12-15

    The expression of the chick kainate-binding protein, a member of the ionotropic glutamate receptor family, is restricted to the cerebellum, specifically to Bergmann glia. Glutamate induces a membrane to nuclei signaling involved in gene expression regulation. Exposure of cultured chick Bergmann glia cells to glutamate leads to an increase in kainate binding protein and mRNA levels, suggesting a transcriptional level of regulation. The 5' proximal region of the chick kainate binding gene was cloned and transfected 4into Bergmann glia cells. Three main regulatory regions could be defined, a minimal promoter region, a negative regulatory region, and interestingly, a glutamate-responsive element. Deletion of this element abolishes the agonist effect. Moreover, electrophoretic mobility shift assays, cotransfection experiments, and site-directed mutagenesis clearly suggest that the glutamate effect is mediated through an AP-1 site by a Fos/Jun heterodimer. The present results favor the notion of a functional role of kainate-binding protein in glutamatergic cerebellar neurotransmission.

  2. PI3K/AKT signaling modulates transcriptional expression of EWS/FLI1 through specificity protein 1

    PubMed Central

    Giorgi, Chiara; Boro, Aleksandar; Rechfeld, Florian; Lopez-Garcia, Laura A.; Gierisch, Maria E.; Schäfer, Beat W.; Niggli, Felix K.

    2015-01-01

    Ewing sarcoma (ES) is the second most frequent bone cancer in childhood and is characterized by the presence of the balanced translocation t(11;22)(q24;q12) in more than 85% of cases, generating a dysregulated transcription factor EWS/FLI1. This fusion protein is an essential oncogenic component of ES development which is necessary for tumor cell maintenance and represents an attractive therapeutic target. To search for modulators of EWS/FLI1 activity we screened a library of 153 targeted compounds and identified inhibitors of the PI3K pathway to directly modulate EWS/FLI1 transcription. Surprisingly, treatment of four different ES cell lines with BEZ235 resulted in down regulation of EWS/FLI1 mRNA and protein by ∼50% with subsequent modulation of target gene expression. Analysis of the EWS/FLI1 promoter region (−2239/+67) using various deletion constructs identified two 14bp minimal elements as being important for EWS/FLI1 transcription. We identified SP1 as modulator of EWS/FLI1 gene expression and demonstrated direct binding to one of these regions in the EWS/FLI1 promoter by EMSA and ChIP experiments. These results provide the first insights on the transcriptional regulation of EWS/FLI1, an area that has not been investigated so far, and offer an additional molecular explanation for the known sensitivity of ES cell lines to PI3K inhibition. PMID:26336820

  3. Sodium butyrate up-regulates cathelicidin gene expression via activator protein-1 and histone acetylation at the promoter region in a human lung epithelial cell line, EBC-1.

    PubMed

    Kida, Yutaka; Shimizu, Takashi; Kuwano, Koichi

    2006-05-01

    The antimicrobial protein cathelicidin is considered to play an important role in the defense mechanisms against bacterial infection. Recent studies show that sodium butyrate induces cathelicidin gene expression in human colonic, gastric and hepatic cells. However, little is known about the precise regulatory mechanisms underlying sodium butyrate-induced cathelicidin gene expression. In this study, we examined the regulatory mechanisms involved in sodium butyrate-induced cathelicidin gene expression using a human lung epithelial cell line, EBC-1. Our results indicate that sodium butyrate induces both cathelicidin mRNA and protein expression. Moreover, deletion or mutation of a putative activator protein-1 (AP-1) binding site in the cathelicidin gene promoter abrogated the response to sodium butyrate stimulation. Three different mitogen-activated protein (MAP) kinase inhibitors suppressed sodium butyrate-induced transactivation of the cathelicidin promoter. Electrophoretic mobility shift assays (EMSA) showed that nuclear extracts prepared from sodium butyrate-stimulated EBC-1 cells generated specific binding to probe including a putative AP-1 binding site in the cathelicidin gene promoter. Furthermore, chromatin immunoprecipitation (ChIP) assays demonstrated that sodium butyrate augmented histone acetylation of the cathelicidin promoter in EBC-1 cells. Therefore, these results indicate that AP-1 and histone acetylation of the cathelicidin promoter play a critical role in the regulation of inducible cathelicidin gene expression in EBC-1 cells stimulated with sodium butyrate.

  4. Differential Expression of the Activator Protein 1 Transcription Factor Regulates Interleukin-1ß Induction of Interleukin 6 in the Developing Enterocyte

    PubMed Central

    Cahill, Catherine M.; Tam, Bosco; Rajanala, Susruthi; Rogers, Jack T.; Walker, W. Allan

    2016-01-01

    The innate immune response is characterized by activation of transcription factors, nuclear factor kappa B and activator protein-1 and their downstream targets, the pro-inflammatory cytokines including interleukin 1β and interleukin 6. Normal development of this response in the intestine is critical to survival of the human neonate and delays can cause the onset of devastating inflammatory diseases such as necrotizing enterocolitis. Previous studies have addressed the role of nuclear factor kappa B in the development of the innate immune response in the enterocyte, however despite its central role in the control of multiple pro-inflammatory cytokine genes, little is known on the role of Activator Protein 1 in this response in the enterocyte. Here we show that the canonical Activator Protein 1 members, cJun and cFos and their upstream kinases JNK and p38 play an essential role in the regulation of interleukin 6 in the immature enterocyte. Our data supports a model whereby the cFos/cJun heterodimer and the more potent cJun homodimer downstream of JNK are replaced by less efficient JunD containing dimers, contributing to the decreased responsiveness to interleukin 1β and decreased interleukin 6 secretion observed in the mature enterocyte. The tissue specific expression of JunB in colonocytes and colon derived tissues together with its ability to repress Interleukin-1β induction of an Interleukin-6 gene reporter in the NCM-460 colonocyte suggests that induction of JunB containing dimers may offer an attractive therapeutic strategy for the control of IL-6 secretion during inflammatory episodes in this area of the intestine PMID:26799482

  5. An in vivo assay for chemoattractant activity.

    PubMed

    Zetter, B R; Rasmussen, N; Brown, L

    1985-09-01

    We have devised an implantable device for the study of leukocyte chemoattraction. The device consists of a 0.25-mm thick patch of Dacron fabric coupled to a disc of ethylene vinyl acetate copolymer. Such polymers can release biologically active molecules at a constant rate for at least 18 days. Attracted cells invade and are trapped within the Dacron fabric. Upon removal from the host, the fabric patches are sectioned and stained to reveal the distribution of attracted cells. Distinct patterns of cellular accumulation can be seen for different chemoattractant molecules. These include the attraction of eosinophils by histamine, monocytes by tuftsin, and mast cells by glycyl-histidyl-lysine. Maximal accumulation of specific cell types occurs at postimplantation days 1 to 2 for neutrophils, days 3 to 5 for monocytes, and days 5 to 6 for macrophages and eosinophils. Control polymers fail to cause significant leukocyte accumulation, indicating that neither the polymer nor the Dacron fabric provokes an inflammatory response. PMID:3162062

  6. The transcription factor Mesp1 interacts with cAMP-responsive element binding protein 1 (Creb1) and coactivates Ets variant 2 (Etv2) gene expression.

    PubMed

    Shi, Xiaozhong; Zirbes, Katie M; Rasmussen, Tara L; Ferdous, Anwarul; Garry, Mary G; Koyano-Nakagawa, Naoko; Garry, Daniel J

    2015-04-10

    Mesoderm posterior 1 (Mesp1) is well recognized for its role in cardiac development, although it is expressed broadly in mesodermal lineages. We have previously demonstrated important roles for Mesp1 and Ets variant 2 (Etv2) during lineage specification, but their relationship has not been defined. This study reveals that Mesp1 binds to the proximal promoter and transactivates Etv2 gene expression via the CRE motif. We also demonstrate the protein-protein interaction between Mesp1 and cAMP-responsive element binding protein 1 (Creb1) in vitro and in vivo. Utilizing transgenesis, lineage tracing, flow cytometry, and immunostaining technologies, we define the lineage relationship between Mesp1- and Etv2-expressing cell populations. We observe that the majority of Etv2-EYFP(+) cells are derived from Mesp1-Cre(+) cells in both the embryo and yolk sac. Furthermore, we observe that the conditional deletion of Etv2, using a Mesp1-Cre transgenic strategy, results in vascular and hematopoietic defects similar to those observed in the global deletion of Etv2 and that it has embryonic lethality by embryonic day 9.5. In summary, our study supports the hypothesis that Mesp1 is a direct upstream transactivator of Etv2 during embryogenesis and that Creb1 is an important cofactor of Mesp1 in the transcriptional regulation of Etv2 gene expression. PMID:25694434

  7. Human papillomavirus 16 E2 interacts with neuregulin receptor degradation protein 1 affecting ErbB-3 expression in vitro and in clinical samples of cervical lesions.

    PubMed

    Paolini, Francesca; Curzio, Gianfranca; Melucci, Elisa; Terrenato, Irene; Antoniani, Barbara; Carosi, Mariantonia; Mottolese, Marcella; Vici, Patrizia; Mariani, Luciano; Venuti, Aldo

    2016-05-01

    The ErbB tyrosine kinase receptors play a key role in regulating many cellular functions and human papillomaviruses (HPVs) may interact with transductional pathway of different growth factor receptors. Here, these interactions were analysed in W12 cell line carrying HPV 16 genome and in clinical samples. W12 cells, in which HPV16 becomes integrated during passages, were utilised to detect viral and ErbB family expression at early (W12E) and late passages (W12G). Interestingly, a strong reduction of ErbB-3 expression was observed in W12G. Loss of the E2 and E5 viral genes occurs in W12G and this may affect ErbB-3 receptor expression. E2 and E5 rescue experiments demonstrated that only E2 gene was able to restore ErbB-3 expression. E2 is a transcriptional factor but the expression levels of ErbB3 were unaffected and ErbB-3 promoter did not show any consensus sequence for E2, thus E2 may interact in another way with ErbB3. Indeed, HPV 16 E2 can modulate ErbB-3 by interacting with the ubiquitin ligase neuregulin receptor degradation protein 1 (Nrdp-1) that is involved in the regulation of this receptor, via ubiquitination and degradation. E2 co-immunoprecipitated in a complex with Nrdp-1 leading to hypothesise an involvement of this interaction in ErbB-3 regulation. In addition, 90% of the clinical samples with integrated virus and E2 loss showed no or low ErbB-3 positivity, confirming in vitro results. In conclusion, the new discovered interaction of HPV-16 E2 with Nrdp-1 may affect ErbB-3 expression.

  8. Expression Patterns of the Wnt Pathway Inhibitors Dickkopf3 and Secreted Frizzled-Related Proteins 1 and 4 in Endometrial Endometrioid Adenocarcinoma

    PubMed Central

    Eskander, Ramez N.; Ali, Shamshad; Dellinger, Thanh; Lankes, Heather A.; Randall, Leslie M.; Ramirez, Nilsa C.; Monk, Bradley J.; Walker, Joan L.; Eisenhauer, Eric; Hoang, Bang H.

    2016-01-01

    Objective The aim of the study was to determine the differential expression patterns of the wingless-type (Wnt) pathway inhibitors Dkk3 (Dickkopf 3), SFRP1 (secreted frizzled-related protein 1), and SFRP4 in normal müllerian tissue and endometrial endometrioid adenocarcinoma specimens. Methods Messenger RNA (mRNA) and protein levels of the Wnt pathway inhibitors Dkk3, SFRP1, and SFRP4 were evaluated by real-time reverse transcription–polymerase chain reaction and Western blot analysis. A total of 87 human tissue specimens were obtained from 60 women who participated in Gynecologic Oncology Group protocol 210. Twenty-seven normal müllerian tissues, 32 early-stage, and 28 advanced-stage endometrial endometrioid cancer specimens were analyzed. Results Median age for this cohort was 60 years, with median body mass index of 32 kg/m2. There was a difference in Dkk3 protein expression between normal müllerian tissues and primary endometrial endometrioid adenocarcinoma samples (P = 0.05). There was down-regulation of Dkk3, SFRP1, and SFRP4 mRNA expression in patients with high-grade disease (P = 0.08, 0.06, and 0.05, respectfully). Furthermore, a decrease in SFRP1 and SFPR4 mRNA expression was noted in patients with a diagnosis of locoregional and distant disease recurrence. Lastly, a trend toward decreased progression-free survival in patients with low Dkk3, SFRP1, and SFRP4 mRNA expression levels was noted. Conclusions Wnt pathway inhibitor (Dkk3, sFRP1, and/or sFRP4) expression was down-regulated in patients with high-grade disease and was associated with locoregional and distant disease recurrence. Despite sample size (power) limitations, these results support previous preclinical studies and may suggest a therapeutic role for Wnt signaling in endometrial cancer. PMID:26397159

  9. Human papillomavirus 16 E2 interacts with neuregulin receptor degradation protein 1 affecting ErbB-3 expression in vitro and in clinical samples of cervical lesions.

    PubMed

    Paolini, Francesca; Curzio, Gianfranca; Melucci, Elisa; Terrenato, Irene; Antoniani, Barbara; Carosi, Mariantonia; Mottolese, Marcella; Vici, Patrizia; Mariani, Luciano; Venuti, Aldo

    2016-05-01

    The ErbB tyrosine kinase receptors play a key role in regulating many cellular functions and human papillomaviruses (HPVs) may interact with transductional pathway of different growth factor receptors. Here, these interactions were analysed in W12 cell line carrying HPV 16 genome and in clinical samples. W12 cells, in which HPV16 becomes integrated during passages, were utilised to detect viral and ErbB family expression at early (W12E) and late passages (W12G). Interestingly, a strong reduction of ErbB-3 expression was observed in W12G. Loss of the E2 and E5 viral genes occurs in W12G and this may affect ErbB-3 receptor expression. E2 and E5 rescue experiments demonstrated that only E2 gene was able to restore ErbB-3 expression. E2 is a transcriptional factor but the expression levels of ErbB3 were unaffected and ErbB-3 promoter did not show any consensus sequence for E2, thus E2 may interact in another way with ErbB3. Indeed, HPV 16 E2 can modulate ErbB-3 by interacting with the ubiquitin ligase neuregulin receptor degradation protein 1 (Nrdp-1) that is involved in the regulation of this receptor, via ubiquitination and degradation. E2 co-immunoprecipitated in a complex with Nrdp-1 leading to hypothesise an involvement of this interaction in ErbB-3 regulation. In addition, 90% of the clinical samples with integrated virus and E2 loss showed no or low ErbB-3 positivity, confirming in vitro results. In conclusion, the new discovered interaction of HPV-16 E2 with Nrdp-1 may affect ErbB-3 expression. PMID:26963794

  10. Suppression of tumor necrosis factor receptor-associated protein 1 expression induces inhibition of cell proliferation and tumor growth in human esophageal cancer cells.

    PubMed

    Tian, Xin; Ma, Ping; Sui, Cheng-Guang; Meng, Fan-Dong; Li, Yan; Fu, Li-Ye; Jiang, Tao; Wang, Yang; Jiang, You-Hong

    2014-06-01

    Tumor necrosis factor receptor-associated protein 1 (TRAP1) is a molecular chaperone involved in multidrug resistance and antiapoptosis in some human tumors, but its regulatory mechanisms have not been revealed in esophageal squamous cell carcinoma (ESCC). In this study, 138 specimens of ESCC were analyzed. TRAP1 was overexpressed in ESCC, particularly in poorly differentiated tumors. To further explore the molecular regulatory mechanism, we constructed specific small interfering RNA-expressing vectors targeting Trap1, and knocked down Trap1 expression in the esophageal cancer cell lines ECA109 and EC9706. Knockdown of Trap1 induced increases in reactive oxygen species and mitochondrial depolarization, which have been proposed as critical regulators of apoptosis. The cell cycle was arrested in G2/M phase, and in vitro inhibition of cell proliferation was confirmed with 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-tetrazolium bromide and bromodeoxyuridine assays. Furthermore, re-expression of TRAP1 in Trap1 small interfering RNA-transfected ESCC cells restored cell proliferation and cell apoptosis. Bioluminescence of subcutaneously xenografted ESCC tumor cells demonstrated significant inhibition of in vivo tumor growth by Trap1 knockdown. This study shows that TRAP1 was overexpressed in most patients with ESCC, and caused an increase in antiapoptosis potency. TRAP1 may be regarded as a target in ESCC biotherapy.

  11. Deregulation of Fragile X-related protein 1 by the lipodystrophic lamin A p.R482W mutation elicits a myogenic gene expression program in preadipocytes.

    PubMed

    Oldenburg, Anja R; Delbarre, Erwan; Thiede, Bernd; Vigouroux, Corinne; Collas, Philippe

    2014-03-01

    The nuclear lamina is implicated in the regulation of various nuclear functions. Several laminopathy-causing mutations in the LMNA gene, notably the p.R482W substitution linked to familial partial lipodystrophy type 2 (FPLD2), are clustered in the immunoglobulin fold of lamin A. We report a functional association between lamin A and fragile X-related protein 1 (FXR1P), a protein of the fragile X-related family involved in fragile X syndrome. Searching for proteins differentially interacting with the immunoglobulin fold of wild-type and R482W mutant lamin A, we identify FXR1P as a novel component of the lamin A protein network. The p.R482W mutation abrogates interaction of FXR1P with lamin A. Fibroblasts from FPLD2 patients display elevated levels of FXR1P and delocalized FXR1P. In human adipocyte progenitors, deregulation of lamin A expression leads to FXR1P up-regulation, impairment of adipogenic differentiation and induction of myogenin expression. FXR1P overexpression also stimulates a myogenic gene expression program in these cells. Our results demonstrate a cross-talk between proteins hitherto implicated in two distinct mesodermal pathologies. We propose a model where the FPLD2 lamin A p.R482W mutation elicits, through up-regulation of FXR1P, a remodeling of an adipogenic differentiation program into a myogenic program.

  12. Piperine activates human pregnane X receptor to induce the expression of cytochrome P450 3A4 and multidrug resistance protein 1

    PubMed Central

    Wang, Yue-Ming; Lin, Wenwei; Chai, Sergio C.; Wu, Jing; Ong, Su Sien; Schuetz, Erin G.; Chen, Taosheng

    2013-01-01

    Activation of the pregnane X receptor (PXR) and subsequently its target genes, including those encoding drug transporters and metabolizing enzymes, while playing substantial roles in xenobiotics detoxification, might cause undesired drug-drug interactions. Recently, an increased awareness has been given to dietary components for potential induction of diet-drug interactions through activation of PXR. Here, we studied, whether piperine (PIP), a major component extracted from the widely-used daily spice black pepper, could induce PXR-mediated expression of cytochrome P450 3A4 (CYP3A4) and multidrug resistance protein 1 (MDR1). Our results showed that PIP activated human PXR (hPXR)-mediated CYP3A4 and MDR1 expression in human hepatocytes, intestine cells, and a mouse model; PIP activated hPXR by recruiting its coactivator SRC-1 in both cellular and cell-free systems; PIP bound to the hPXR ligand binding domain in a competitive ligand binding assay in vitro. The dichotomous effects of PIP on induction of CYP3A4 and MDR1 expression observed here and inhibition of their activity reported elsewhere challenges the potential use of PIP as a bioavailability enhancer and suggests that cautions should be taken for PIP consumption during drug treatment in patients, particularly those who favor daily pepper spice or rely on certain pepper remedies. PMID:23707768

  13. Bi-phasic expression of Heterochromatin Protein 1 (HP1) during breast cancer progression: Potential roles of HP1 and chromatin structure in tumorigenesis

    PubMed Central

    Lee, Young-Ho; Ann, David K.

    2015-01-01

    Epigenetics in cancer prognosis and therapy is gaining recognition in recent years. Breast cancer is a genetic disease harboring numerous genetic mutations, including tumor suppressor BRCA1 and BRCA2 mutations. However, the functions of BRCA1 in cancer cells are also altered by non-genetic mechanisms, including DNA methylation and chromatin structure. Therefore, identification of epigenetic markers for breast cancer is very important for early diagnosis and effective therapy. This review focuses on recent findings on the roles of Heterochromatin protein 1 (HP1) in BRCA1 functions and breast cancer progression. We previously showed that BRCA1 function and breast cancer progression are frequently associated with HP1 expression level and potentially with chromatin structure. Herein, we suggest that bi-phasic expression of HP1 during breast cancer progression indicates dual roles of HP1 in tumorigenesis. Exploiting differential HP1 expression in tumors could lead to effective cancer therapy. Re-setting the chromatin structure may be a critical step for high-efficiency cancer therapy for many breast cancer patients. PMID:26082944

  14. Progesterone from the Cumulus Cells Is the Sperm Chemoattractant Secreted by the Rabbit Oocyte Cumulus Complex

    PubMed Central

    Guidobaldi, Héctor Alejandro; Teves, María Eugenia; Uñates, Diego Rafael; Anastasía, Agustín; Giojalas, Laura Cecilia

    2008-01-01

    Sperm chemotaxis in mammals have been identified towards several female sources as follicular fluid (FF), oviduct fluid, and conditioned medium from the cumulus oophorus (CU) and the oocyte (O). Though several substances were confirmed as sperm chemoattractant, Progesterone (P) seems to be the best chemoattractant candidate, because: 1) spermatozoa express a cell surface P receptor, 2) capacitated spermatozoa are chemotactically attracted in vitro by gradients of low quantities of P; 3) the CU cells produce and secrete P after ovulation; 4) a gradient of P may be kept stable along the CU; and 5) the most probable site for sperm chemotaxis in vivo could be near and/or inside the CU. The aim of this study was to verify whether P is the sperm chemoattractant secreted by the rabbit oocyte-cumulus complex (OCC) in the rabbit, as a mammalian animal model. By means of videomicroscopy and computer image analysis we observed that only the CU are a stable source of sperm attractants. The CU produce and secrete P since the hormone was localized inside these cells by immunocytochemistry and in the conditioned medium by enzyme immunoassay. In addition, rabbit spermatozoa express a cell surface P receptor detected by western blot and localized over the acrosomal region by immunocytochemistry. To confirm that P is the sperm chemoattractant secreted by the CU, the sperm chemotactic response towards the OCC conditioned medium was inhibited by three different approaches: P from the OCC conditioned medium was removed with an anti-P antibody, the attractant gradient of the OCC conditioned medium was disrupted by a P counter gradient, and the sperm P receptor was blocked with a specific antibody. We concluded that only the CU but not the oocyte secretes P, and the latter chemoattract spermatozoa by means of a cell surface receptor. Our findings may be of interest in assisted reproduction procedures in humans, animals of economic importance and endangered species. PMID:18725941

  15. Mathematics of Experimentally Generated Chemoattractant Gradients.

    PubMed

    Postma, Marten; van Haastert, Peter J M

    2016-01-01

    Many eukaryotic cells move in the direction of a chemical gradient. Several assays have been developed to measure this chemotactic response, but no complete mathematical models of the spatial and temporal gradients are available to describe the fundamental principles of chemotaxis. Here we provide analytical solutions for the gradients formed by release of chemoattractant from a point source by passive diffusion or forced flow (micropipettes) and gradients formed by laminar diffusion in a Zigmond chamber. The results show that gradients delivered with a micropipette are formed nearly instantaneously, are very steep close to the pipette, and have a steepness that is strongly dependent on the distance from the pipette. In contrast, gradients in a Zigmond chamber are formed more slowly, are nearly independent of the distance from the source, and resemble the temporal and spatial properties of the natural cAMP wave that Dictyostelium cells experience during cell aggregation. PMID:27271915

  16. Mathematics of Experimentally Generated Chemoattractant Gradients.

    PubMed

    Postma, Marten; van Haastert, Peter J M

    2016-01-01

    Many eukaryotic cells move in the direction of a chemical gradient. Several assays have been developed to measure this chemotactic response, but no complete mathematical models of the spatial and temporal gradients are available to describe the fundamental principles of chemotaxis. Here we provide analytical solutions for the gradients formed by release of chemoattractant from a point source by passive diffusion or forced flow (micropipettes) and gradients formed by laminar diffusion in a Zigmond chamber. The results show that gradients delivered with a micropipette are formed nearly instantaneously, are very steep close to the pipette, and have a steepness that is strongly dependent on the distance from the pipette. In contrast, gradients in a Zigmond chamber are formed more slowly, are nearly independent of the distance from the source, and resemble the temporal and spatial properties of the natural cAMP wave that Dictyostelium cells experience during cell aggregation.

  17. Chromodomain Helicase/ATPase DNA-Binding Protein 1-Like Gene (CHD1L) Expression and Implications for Invasion and Metastasis of Breast Cancer

    PubMed Central

    Mu, Qing-Jie; Li, Hong-Li; Yao, Yuan; Liu, Shi-Chao; Yin, Chong-Gao; Ma, Xue-Zhen

    2015-01-01

    Background Chromodomain helicase/ATPase DNA-binding protein 1-like gene (CHD1L), also known as ALC1 (amplified in liver cancer 1 gene), is a new oncogene amplified in many solid tumors. Whether this gene plays a role in invasion and metastasis of breast cancer is unknown. Methods Immunohistochemistry was performed to detect the expression of CHD1L in patients with invasive ductal carcinoma and normal mammary glands. Chemotaxis, wound healing, and Transwell invasion assays were also performed to examine cell migration and invasion. Western blot analysis was conducted to detect the expression of CHD1L, MMP-2, MMP-9, pAkt/Akt, pARK5/ARK5, and pmTOR/mTOR. Moreover, ELISA was carried out to detect the expression levels of MMP-2 and MMP-9. Nude mice xenograft model was used to detect the invasion and metastasis of breast cancer cell lines. Results CHD1L overexpression was observed in 112 of 268 patients (41.8%). This overexpression was associated with lymph node metastasis (P = 0.008), tumor differentiation (P = 0.020), distant metastasis (P = 0.026), MMP-2 (P = 0.035), and MMP-9 expression (P = 0.022). In the cell experiment, reduction of CHD1L inhibited the invasion and metastasis of breast cancer cells by mediating MMP-2 and MMP-9 expression. CHD1L knockdown via siRNA suppressed EGF-induced pAkt, pARK5, and pmTOR. This knockdown inhibited the metastasis of breast cancer cells into the lungs of SCID mice. Conclusions CHD1L promoted the invasion and metastasis of breast cancer cells via the PI3K/Akt/ARK5/mTOR/MMP signaling pathway. This study identified CHD1L as a potential anti-metastasis target for therapeutic intervention in breast cancer. PMID:26599012

  18. Does the Clearance of Inhaled (99m)Tc-Sestamibi Correlate with Multidrug Resistance Protein 1 Expression in the Human Lung?

    PubMed

    Mohan, Hosahalli K; Routledge, Thomas; Cane, Paul; Livieratos, Lefteris; Ballinger, James R; Peters, Adrien M

    2016-09-01

    Purpose To examine the relation between the lung elimination rate of inhaled technetium 99m ((99m)Tc)-sestamibi and immunohistochemical expression of bronchopulmonary multidrug resistance protein 1 (MRP1) and permeability glycoprotein (P-gp) and assess the repeatability of the inhaled (99m)Tc-sestamibi clearance technique. Materials and Methods (99m)Tc-sestamibi is a known substrate for P-gp and MRP1, which are established cellular drug efflux transporters. The elimination rate of (99m)Tc-sestamibi from the lungs after inhalation as an aerosol has been hypothesized to be regulated by expression of these transporters. Institutional ethics committee approval was received for this prospective study. Written informed consent was obtained from all participants. The clearance of inhaled (99m)Tc-sestamibi from the lungs of 13 patients due to undergo surgery for primary lung cancer (five of 13) or spontaneous pneumothorax (eight of 13) was estimated after dynamic imaging of the lungs during a period of 40 minutes. The time taken to clear 50% of inhaled sestamibi (T1/2) was compared with a semiquantitative immunohistochemical assessment (grade 0-3) of MRP1 and P-gp expression in the lung by using parametric and nonparametric tests. The study was repeated in five participants to assess the repeatability of the technique by using a Bland Altman analysis method. Results MRP1 expression was seen in 12 of 13 patients, while P-gp expression was seen in only two. The mean (99m)Tc-sestamibi elimination rate was faster in patients (n = 6) with low levels of MRP1 expression (grade 0-1) and mean T1/2 of 105 minutes ± 20 (standard deviation), compared with those with higher levels of MRP1 expression (grade 2-3, n = 7) and mean T1/2 of 149 minutes ± 28 (P = .008). Bland-Altman analysis revealed excellent agreement between test and retest values. Conclusion Inhaled (99m)Tc-sestamibi clearance study is a repeatable technique demonstrating significant correlation with MRP1 expression in

  19. Low p53 Binding Protein 1 (53BP1) Expression Is Associated With Increased Local Recurrence in Breast Cancer Patients Treated With Breast-Conserving Surgery and Radiotherapy

    SciTech Connect

    Neboori, Hanmanth J.R.; Haffty, Bruce G.; Wu Hao; Yang Qifeng; Aly, Amal; Goyal, Sharad; Schiff, Devora; Moran, Meena S.; Golhar, Ryan; Chen Chunxia; Moore, Dirk; and others

    2012-08-01

    Purpose: To investigate whether the expression of p53 binding protein 1 (53BP1) has prognostic significance in a cohort of early-stage breast cancer patients treated with breast-conserving surgery and radiotherapy (BCS+RT). Methods and Materials: A tissue microarray of early-stage breast cancer treated with BCS+RT from a cohort of 514 women was assayed for 53BP1, estrogen receptor, progesterone receptor, and HER2 expression by immunohistochemistry. Through log-rank tests and univariate and multivariate models, the staining profile of each tumor was correlated with clinical endpoints, including ipsilateral breast recurrence-free survival (IBRFS), distant metastasis-free survival (DMFS), cause-specific survival (CSS), recurrence-free survival (RFS), and overall survival (OS). Results: Of the 477 (93%) evaluable tumors, 63 (13%) were scored as low. Low expression of 53BP1 was associated with worse outcomes for all endpoints studied, including 10-year IBRFS (76.8% vs. 90.5%; P=.01), OS (66.4% vs. 81.7%; P=.02), CSS (66.0% vs. 87.4%; P<.01), DMFS (55.9% vs. 87.0%; P<.01), and RFS (45.2% vs. 80.6%; P<.01). Multivariate analysis incorporating various clinico-pathologic markers and 53BP1 expression found that 53BP1 expression was again an independent predictor of all endpoints (IBRFS: P=.0254; OS: P=.0094; CSS: P=.0033; DMFS: P=.0006; RFS: P=.0002). Low 53BP1 expression was also found to correlate with triple-negative (TN) phenotype (P<.01). Furthermore, in subset analysis of all TN breast cancer, negative 53BP1 expression trended for lower IBRFS (72.3% vs. 93.9%; P=.0361) and was significant for worse DMFS (48.2% vs. 86.8%; P=.0035) and RFS (37.8% vs. 83.7%; P=.0014). Conclusion: Our data indicate that low 53BP1 expression is an independent prognostic indicator for local relapse among other endpoints in early-stage breast cancer and TN breast cancer patients treated with BCS+RT. These results should be verified in larger cohorts of patients to validate their clinical

  20. Piperine activates human pregnane X receptor to induce the expression of cytochrome P450 3A4 and multidrug resistance protein 1

    SciTech Connect

    Wang, Yue-Ming; Lin, Wenwei; Chai, Sergio C.; Wu, Jing; Ong, Su Sien; Schuetz, Erin G.; Chen, Taosheng

    2013-10-01

    Activation of the pregnane X receptor (PXR) and subsequently its target genes, including those encoding drug transporters and metabolizing enzymes, while playing substantial roles in xenobiotic detoxification, might cause undesired drug-drug interactions. Recently, an increased awareness has been given to dietary components for potential induction of diet–drug interactions through activation of PXR. Here, we studied, whether piperine (PIP), a major component extracted from the widely-used daily spice black pepper, could induce PXR-mediated expression of cytochrome P450 3A4 (CYP3A4) and multidrug resistance protein 1 (MDR1). Our results showed that PIP activated human PXR (hPXR)-mediated CYP3A4 and MDR1 expression in human hepatocytes, intestine cells, and a mouse model; PIP activated hPXR by recruiting its coactivator SRC-1 in both cellular and cell-free systems; PIP bound to the hPXR ligand binding domain in a competitive ligand binding assay in vitro. The dichotomous effects of PIP on induction of CYP3A4 and MDR1 expression observed here and inhibition of their activity reported elsewhere challenges the potential use of PIP as a bioavailability enhancer and suggests that caution should be taken in PIP consumption during drug treatment in patients, particularly those who favor daily pepper spice or rely on certain pepper remedies. - Highlights: • Piperine induces PXR-mediated CYP3A4 and MDR1 expression. • Piperine activates PXR by binding to PXR and recruiting coactivator SRC-1. • Piperine induces PXR activation in vivo. • Caution should be taken in piperine consumption during drug treatment.

  1. Serological Conservation of Parasite-Infected Erythrocytes Predicts Plasmodium falciparum Erythrocyte Membrane Protein 1 Gene Expression but Not Severity of Childhood Malaria.

    PubMed

    Warimwe, George M; Abdi, Abdirahman I; Muthui, Michelle; Fegan, Gregory; Musyoki, Jennifer N; Marsh, Kevin; Bull, Peter C

    2016-05-01

    Plasmodium falciparum erythrocyte membrane protein 1 (PfEMP1), expressed on P. falciparum-infected erythrocytes, is a major family of clonally variant targets of naturally acquired immunity to malaria. Previous studies have demonstrated that in areas where malaria is endemic, antibodies to infected erythrocytes from children with severe malaria tend to be more seroprevalent than antibodies to infected erythrocytes from children with nonsevere malaria. These data have led to a working hypothesis that PfEMP1 variants associated with parasite virulence are relatively conserved in structure. However, the longevity of such serologically conserved variants in the parasite population is unknown. Here, using infected erythrocytes from recently sampled clinical P. falciparum samples, we measured serological conservation using pools of antibodies in sera that had been sampled 10 to 12 years earlier. The serological conservation of infected erythrocytes strongly correlated with the expression of specific PfEMP1 subsets previously found to be associated with severe malaria. However, we found no association between serological conservation per se and disease severity within these data. This contrasts with the simple hypothesis that P. falciparum isolates with a serologically conserved group of PfEMP1 variants cause severe malaria. The data are instead consistent with periodic turnover of the immunodominant epitopes of PfEMP1 associated with severe malaria.

  2. La Autoantigen Induces Ribosome Binding Protein 1 (RRBP1) Expression through Internal Ribosome Entry Site (IRES)-Mediated Translation during Cellular Stress Condition

    PubMed Central

    Gao, Wenqing; Li, Qi; Zhu, Ruiyu; Jin, Jian

    2016-01-01

    The function of ribosome binding protein 1 (RRBP1) is regulating the transportation and secretion of some intracellular proteins in mammalian cells. Transcription of RRBP1 is induced by various cytokines. However, few studies focused on the process of RRPB1 mRNA translation. The RRBP1 mRNA has a long 5′ untranslated region that potentially formed a stable secondary structure. In this study, we show that the 5′ UTR of RRBP1 mRNA contains an internal ribosome entry site (IRES). Moreover, the RRBP1 expression is induced by chemotherapeutic drug paclitaxel or adriamycin in human hepatocellular carcinoma cells and accompanied with the increased expression of La autoantigen (La), which binds to RRBP1 IRES element and facilitates translation initiation. Interestingly, we found IRES-mediated RRBP1 translation is also activated during serum-starvation condition which can induce cytoplasmic localization of La. After mapping the entire RRBP1 5′ UTR, we determine the core IRES activity is located between nt-237 and -58. Furthermore, two apical GARR loops within the functional RRBP1 IRES elements may be important for La binding. These results strongly suggest an important role for IRES-dependent translation of RRBP1 mRNA in hepatocellular carcinoma cells during cellular stress conditions. PMID:27447629

  3. La Autoantigen Induces Ribosome Binding Protein 1 (RRBP1) Expression through Internal Ribosome Entry Site (IRES)-Mediated Translation during Cellular Stress Condition.

    PubMed

    Gao, Wenqing; Li, Qi; Zhu, Ruiyu; Jin, Jian

    2016-01-01

    The function of ribosome binding protein 1 (RRBP1) is regulating the transportation and secretion of some intracellular proteins in mammalian cells. Transcription of RRBP1 is induced by various cytokines. However, few studies focused on the process of RRPB1 mRNA translation. The RRBP1 mRNA has a long 5' untranslated region that potentially formed a stable secondary structure. In this study, we show that the 5' UTR of RRBP1 mRNA contains an internal ribosome entry site (IRES). Moreover, the RRBP1 expression is induced by chemotherapeutic drug paclitaxel or adriamycin in human hepatocellular carcinoma cells and accompanied with the increased expression of La autoantigen (La), which binds to RRBP1 IRES element and facilitates translation initiation. Interestingly, we found IRES-mediated RRBP1 translation is also activated during serum-starvation condition which can induce cytoplasmic localization of La. After mapping the entire RRBP1 5' UTR, we determine the core IRES activity is located between nt-237 and -58. Furthermore, two apical GARR loops within the functional RRBP1 IRES elements may be important for La binding. These results strongly suggest an important role for IRES-dependent translation of RRBP1 mRNA in hepatocellular carcinoma cells during cellular stress conditions. PMID:27447629

  4. Effects of tibolone and its metabolites on prolactin and insulin-like growth factor binding protein-1 expression in human endometrial stromal cells.

    PubMed

    Guzel, Elif; Buchwalder, Lynn; Basar, Murat; Kayisli, Umit; Ocak, Nehir; Bozkurt, Idil; Lockwood, Charles J; Schatz, Frederick

    2015-05-01

    The effects of the postmenopausal replacement steroid tibolone and its 3α-, 3β-OH and Δ-4 tibolone metabolites were evaluated on progesterone receptor-mediated classic decidualization markers insulin-like growth factor binding protein-1 (IGFBP-1) and prolactin expression in human endometrial stromal cells (HESCs). Supernatants of conditioned medium or erxtracted RNA from experimental cell incubations of confluent HESCs were subjected to ELISAs, Western blot analysis and RT/PCR, and results were statisically assesed. Over 21 days, specific ELISAs observed linear increases in secreted IGFBP-1 and prolactin levels elicited by tibolone and its metabolites. Cultured HESCs were refractory to E2 and dexamethasone, whereas tibolone and each metabolite exceeded medroxyprogesterone acetate in significantly elevating IGFBP-1 and prolactin output. Anti-progestins eliminated IGFBP-1 and prolactin induction by tibolone and its metabolites. Immunoblotting and RT/PCR confirmed ELISA results. These observations of IGFBP-1 and prolactin expression: (a) indicate the relevance of cultured HESCs in evaluating the chronic effects of tibolone administration to women; (b) are consistent with PR-mediated endometrial atrophy and protection against endometrial bleeding despite the persistence of circulating ER-binding, but not PR-binding metabolites following tibolone administration to women.

  5. cAMP-Response Element-Binding 3-Like Protein 1 (CREB3L1) is Required for Decidualization and its Expression is Decreased in Women with Endometriosis.

    PubMed

    Ahn, J I; Yoo, J-Y; Kim, T H; Kim, Y I; Ferguson, S D; Fazleabas, A T; Young, S L; Lessey, B A; Ahn, J Y; Lim, J M; Jeong, J-W

    2016-01-01

    Endometriosis is a major cause of infertility and pelvic pain, affecting more than 10% of reproductive-aged women. Progesterone resistance has been observed in the endometrium of women with this disease, as evidenced by alterations in progesterone-responsive gene and protein expression. cAMPResponse Element-Binding 3-like protein 1 (Creb3l1) has previously been identified as a progesterone receptor (PR) target gene in mouse uterus via high density DNA microarray analysis. However, CREB3L1 function has not been studied in the context of endometriosis and uterine biology. In this study, we validated progesterone (P4) regulation of Creb3l1 in the uteri of wild-type and progesterone receptor knockout (PRKO) mice. Furthermore, we observed that CREB3L1 expression was significantly higher in secretory phase human endometrium compared to proliferative phase and that CREB3L1 expression was significantly decreased in the endometrium of women with endometriosis. Lastly, by transfecting CREB3L1 siRNA into cultured human endometrial stromal cells (hESCs) prior to hormonal induction of in vitro decidualization, we showed that CREB3L1 is required for the decidualization process. Interestingly, phosphorylation of ERK1/2, critical factor for decidualization, was also significantly reduced in CREB3L1-silenced hESCs. It is known that hESCs from patients with endometriosis show impaired decidualization and that dysregulation of the P4-PR signaling axis is linked to a variety of endometrial diseases including infertility and endometriosis. Therefore, these results suggest that CREB3L1 is required for decidualization in mice and humans and may be linked to the pathogenesis of endometriosis in a P4-dependent manner. PMID:26917262

  6. Constitutive expression of a COOH-terminal leucine mutant of lysosome-associated membrane protein-1 causes its exclusive localization in low density intracellular vesicles.

    PubMed

    Akasaki, Kenji; Shiotsu, Keiko; Michihara, Akihiro; Ide, Norie; Wada, Ikuo

    2014-07-01

    Lysosome-associated membrane protein-1 (LAMP-1) is a type I transmembrane protein with a short cytoplasmic tail that possesses a lysosome-targeting signal of GYQTI(382)-COOH. Wild-type (WT)-LAMP-1 was exclusively localized in high density lysosomes, and efficiency of LAMP-1's transport to lysosomes depends on its COOH-terminal amino acid residue. Among many different COOH-terminal amino acid substitution mutants of LAMP-1, a leucine-substituted mutant (I382L) displays the most efficient targeting to late endosomes and lysosomes [Akasaki et al. (2010) J. Biochem. 148: , 669-679]. In this study, we generated two human hepatoma cell lines (HepG2 cell lines) that stably express WT-LAMP-1 and I382L, and compared their intracellular distributions. The subcellular fractionation study using Percoll density gradient centrifugation revealed that WT-LAMP-1 had preferential localization in the high density secondary lysosomes where endogenous human LAMP-1 was enriched. In contrast, a major portion of I382L was located in a low density fraction. The low density fraction also contained approximately 80% of endogenous human LAMP-1 and significant amounts of endogenous β-glucuronidase and LAMP-2, which probably represents occurrence of low density lysosomes in the I382L-expressing cells. Double immunofluorescence microscopic analyses distinguished I382L-containing intracellular vesicles from endogenous LAMP-1-containing lysosomes and early endosomes. Altogether, constitutive expression of I382L causes its aberrant intracellular localization and generation of low density lysosomes, indicating that the COOH-terminal isoleucine is critical for normal localization of LAMP-1 in the dense lysosomes.

  7. Tyrosinase-related protein 1 mRNA expression in lymph node metastases predicts overall survival in high-risk melanoma patients

    PubMed Central

    El Hajj, P; Journe, F; Wiedig, M; Laios, I; Salès, F; Galibert, M-D; Van Kempen, L C; Spatz, A; Badran, B; Larsimont, D; Awada, A; Ghanem, G

    2013-01-01

    Background: Clinical outcome of high-risk melanoma patients is not reliably predicted from histopathological analyses of primary tumours and is often adjusted during disease progression. Our study aimed at extending our previous findings in skin metastases to evaluate the prognostic value of tyrosinase-related protein 1 (TYRP1) in lymph node metastases of stages III and IV melanoma patients. Methods: TYRP1 mRNA expression in 104 lymph node metastases was quantified by real-time PCR and normalised to S100 calcium-binding protein B (S100B) mRNA expression to correct for tumour load. TYRP1/S100B ratios were calculated and median was used as cutoff value. TYRP1/S100B mRNA values were correlated to clinical follow-up and histopathological characteristics of the primary lesion. Results: A high TYRP1/S100B mRNA ratio significantly correlated with reduced disease-free (DFS) and overall survival (OS; Cox regression analysis, P=0.005 and 0.01, respectively), increased Breslow thickness (Spearman's rho test, P<0.001) and the presence of ulceration (Mann–Whitney test, P=0.02) of the primaries. Moreover, high TYRP1/S100B was of better prognostic value (lower P-value) for OS than Breslow thickness and ulceration. Finally, it was well conserved during disease progression with respect to high/low TYRP1 groups. Conclusion: High TYRP1/S100B mRNA expression in lymph node metastases from melanoma patients is associated with unfavourable clinical outcome. Its evaluation in lymph node metastases may refine initial prognosis for metastatic patients, may define prognosis for those with unknown or non-evaluable primary lesions and may allow different management of the two groups of patients. PMID:23519055

  8. Impact of Concanavalin-A-Mediated Cytoskeleton Disruption on Low-Density Lipoprotein Receptor-Related Protein-1 Internalization and Cell Surface Expression in Glioblastomas

    PubMed Central

    Nanni, Samuel Burke; Pratt, Jonathan; Beauchemin, David; Haidara, Khadidja; Annabi, Borhane

    2016-01-01

    The low-density lipoprotein receptor-related protein 1 (LRP-1) is a multiligand endocytic receptor, which plays a pivotal role in controlling cytoskeleton dynamics during cancer cell migration. Its rapid endocytosis further allows efficient clearance of extracellular ligands. Concanavalin-A (ConA) is a lectin used to trigger in vitro physiological cellular processes, including cytokines secretion, nitric oxide production, and T-lymphocytes activation. Given that ConA exerts part of its effects through cytoskeleton remodeling, we questioned whether it affected LRP-1 expression, intracellular trafficking, and cell surface function in grade IV U87 glioblastoma cells. Using flow cytometry and confocal microscopy, we found that loss of the cell surface 600-kDa mature form of LRP-1 occurs upon ConA treatment. Consequently, internalization of the physiological α2-macroglobulin and the synthetic angiopep-2 ligands of LRP-1 was also decreased. Silencing of known mediators of ConA, such as the membrane type-1 matrix metalloproteinase, and the Toll-like receptors (TLR)-2 and TLR-6 was unable to rescue ConA-mediated LRP-1 expression decrease, implying that the loss of LRP-1 was independent of cell surface relayed signaling. The ConA-mediated reduction in LRP-1 expression was emulated by the actin cytoskeleton-disrupting agent cytochalasin-D, but not by the microtubule inhibitor nocodazole, and required both lysosomal- and ubiquitin-proteasome system-mediated degradation. Our study implies that actin cytoskeleton integrity is required for proper LRP-1 cell surface functions and that impaired trafficking leads to specialized compartmentation and degradation. Our data also strengthen the biomarker role of cell surface LRP-1 functions in the vectorized transport of therapeutic angiopep bioconjugates into brain cancer cells. PMID:27226736

  9. Studying Chemoattractant Signal Transduction Dynamics in Dictyostelium by BRET.

    PubMed

    Islam, A F M Tariqul; Stepanski, Branden M; Charest, Pascale G

    2016-01-01

    Understanding the dynamics of chemoattractant signaling is key to our understanding of the mechanisms underlying the directed migration of cells, including that of neutrophils to sites of infections and of cancer cells during metastasis. A model frequently used for deciphering chemoattractant signal transduction is the social amoeba Dictyostelium discoideum. However, the methods available to quantitatively measure chemotactic signaling are limited. Here, we describe a protocol to quantitatively study chemoattractant signal transduction in Dictyostelium by monitoring protein-protein interactions and conformational changes using Bioluminescence Resonance Energy Transfer (BRET). PMID:27271894

  10. Assessment of the prognostic impact of the Epstein–Barr virus-encoded latent membrane protein-1 expression in Hodgkin's disease

    PubMed Central

    Glavina-Durdov, M; Jakic-Razumovic, J; Capkun, V; Murray, P

    2001-01-01

    We have examined expression of the Epstein–Barr virus (EBV) latent membrane protein-1 (LMP1) in the malignant Hodgkin and Reed–Sternberg (HRS) cells of Hodgkin's disease (HD) and its impact on response to treatment and on survival. Paraffin tissue from 100 adult immunocompetent patients with HD were analysed using immunohistochemistry to identify LMP1 expression. According to the Rye classification, 8% of patients had lymphocyte predominance (LP) subtype, 48% had nodular sclerosis (NS) disease, 37% were of the mixed cellularity (MC) subtype and 7% were of the lymphocyte depletion (LD) subtype. During the five year follow-up period 27 patients died and 74 patients achieved a complete remission. Patients with LD subtype were older (P = 0.03), less frequently achieved complete remission (P = 0.01), had shorter disease-free survival (P = 0.01) and overall survival (P = 0.002) compared with the other subtypes of HD. LMP1 expression was found in the tumour cells of 26% of cases of HD. LMP1 expression was less common in NS disease than in the other subtypes (P = 0.05), whereas an association between MC subtype and LMP1 expression was not found (P = 0.22). Using the log-rank test there were no differences in overall survival or disease-free survival based on EBV status either when all patients were analysed or when LD and LP subtypes were excluded. However, the presence of EBV was associated with significantly longer disease-free survival in the subgroup of patients ≤ 30 years old (P = 0.02) and in those patients ≤ 34 years old (P = 0.05). In contrast, there was a trend for shorter disease-free survival for EBV-positive patients in the subgroup > 35 years old, but this difference was not statistically significant (P = 0.11). A similar trend was observed in patients > 50 years old. Analysis of the impact of LMP1 expression in patients who had different stage and B symptoms status showed that expression of EBV was associated with longer disease-free survival

  11. Polymorphism, Expression of Natural Resistance-associated Macrophage Protein 1 Encoding Gene (NRAMP1) and Its Association with Immune Traits in Pigs.

    PubMed

    Ding, Xiaoling; Zhang, Xiaodong; Yang, Yong; Ding, Yueyun; Xue, Weiwei; Meng, Yun; Zhu, Weihua; Yin, Zongjun

    2014-08-01

    Natural resistance-associated macrophage protein 1 encoding gene (NRAMP1) plays an important role in immune response against intracellular pathogens. To evaluate the effects of NRAMP1 gene on immune capacity in pigs, tissue expression of NRAMP1 mRNA was observed by real time quantitative polymerase chain reaction (PCR), and the results revealed NRAMP1 expressed widely in nine tissues. One single nucleotide polymorphism (SNP) (ENSSSCG00000025058: g.130 C>T) in exon1 and one SNP (ENSSSCG00000025058: g.657 A>G) in intron1 region of porcine NRAMP1 gene were demonstrated by DNA sequencing and PCR-RFLP analysis. A further analysis of SNP genotypes associated with immune traits including contain of white blood cell (WBC), granulocyte, lymphocyte, monocyte (MO), rate of cytotoxin in monocyte (MC) and CD4/CD8 T lymphocyte subpopulations in blood was carried out in four pig populations including Large White and three Chinese indigenous breeds (Wannan Black, Huai pig and Wei pig). The results showed that the SNP (ENSSSCG00000025058: g.130 C>T) was significantly associated with level of WBC % (p = 0.031), MO% (p = 0.024), MC% (p = 0.013) and CD4(-)CD8(+) T lymphocyte (p = 0.023). The other SNP (ENSSSCG00000025058: g.657 A>G) was significantly associated with the level of MO% (p = 0.012), MC% (p = 0.019) and CD4(-)CD8(+) T lymphocyte (p = 0.037). These results indicate that the NRAMP1 gene can be regarded as a molecular marker for genetic selection of disease susceptibility in pig breeding. PMID:25083114

  12. Effect of multiple cysteine substitutions on the functionality of human multidrug resistance protein 1 expressed in human embryonic kidney 293 cells: identification of residues essential for function.

    PubMed

    Qin, Lei; Tam, Shui-Pang; Deeley, Roger G

    2012-07-01

    Multidrug resistance protein 1 (MRP1) is a broad-specificity membrane transporter belonging to the C branch of the ATP binding cassette (ABC) superfamily. MRP1 confers resistance to various chemotherapeutic drugs and transports a wide range of conjugated organic anions. Several ABCC proteins, including MRP1, are unusual among ABC transporters in having a third membrane-spanning domain (MSD), MSD0, at their N termini. MRP1 lacking this additional MSD (ΔMRP1) is able to traffic to the plasma membrane of mammalian cells and to transport a number of well characterized substrates. A cysteineless (cysless) ΔMRP1 has been expressed in yeast and reported to be functional. However, we found that trafficking of such a construct in human cells was severely compromised, and, even when expressed in insect Sf21 cells, the protein had extremely low transport activity. Therefore, we have systematically examined the effects of substituting cysteines in the four domains of ΔMRP1, initially with alanine. These studies allowed us to identify five cysteines that cannot be replaced with alanine without inactivating the protein. Substitution of two of these residues with alternative amino acids has allowed us to produce an almost cysless form of ΔMRP1 that traffics to the plasma membrane and transports leukotriene C(4), 17β-estradiol 17-β-D-glucuronide, and estrone-3-sulfate with kinetic characteristics similar to those of the wild-type protein. The distribution of the remaining Cys residues is such that the protein will provide a useful template for a variety of cysteine based mutagenesis studies.

  13. Increased levels of reactive oxygen species and expression of a cytoplasmic aconitase/iron regulatory protein 1 homolog during the early response of maize pulvini to gravistimulation.

    PubMed

    Clore, A M; Doore, S M; Tinnirello, S M N

    2008-01-01

    The maize (Zea mays L.) stem pulvinus is a disc of tissue located apical to each node that functions to return a tipped stem to a more upright position via increased cell elongation on its lower side. We investigated the possibility that reactive oxygen species (ROS) and hydrogen peroxide (H2O2), in particular, are involved in the gravitropic response of the pulvinus prior to initiation of the growth response by employing the cytochemical stain 3,3'-diaminobenzidine (DAB). DAB polymers were found in the bundle sheath cells of gravistimulated pulvini in association with amyloplasts after 1 min of gravistimulation, and the signal spread throughout the cytosol of these cells by 30 min. Furthermore, treatment of maize stem explants containing pulvini with 1 mm H2O2 on their upper sides caused reversal of bending polarity. Similar, though less dramatic, results were obtained via application of 1 mm ascorbic acid to the lower side of the explants. In addition, we determined that a maize cytoplasmic aconitase/iron regulatory protein 1 (IRP1) homolog is up-regulated in the pulvinus bundle sheath cells after gravistimulation using suppressive subtractive hybridization PCR (SSH PCR), real-time RT-PCR and in situ hybridization. Although we do not yet know the role of the IRP1 homolog in the pulvinus, the protein is known to be a redox sensor in other systems. Collectively, our results point to an increase in ROS quite early in the gravitropic signalling pathway and its possible role in determining the direction of bending of the pulvini. We speculate that an ROS burst may serve to link the physical phenomenon of amyloplast sedimentation to the changes in cellular biochemistry and gene expression that facilitate directional growth. PMID:18004982

  14. Insulin-like growth factor-binding protein-1 (IGFBP-1) in goldfish, Carassius auratus: molecular cloning, tissue expression, and mRNA expression responses to periprandial changes and cadmium exposure.

    PubMed

    Chen, Wenbo; Zhang, Zhen; Dong, Haiyan; Yan, Fangfang

    2016-06-01

    In this study, the cDNA encoding insulin-like growth factor-binding protein-1 (IGFBP-1) was cloned from the liver of goldfish (Carassius auratus). The obtained goldfish IGFBP-1 cDNA sequence was 1037 bp in length and had an open reading frame of 789 bp encoding a predicted polypeptide of 262 amino acid residues. IGFBP-1 transcript was detected in all tested central nervous and peripheral tissues. The relatively higher levels of IGFBP-1 mRNA were observed in the liver, gill, kidney, heart, spleen, fat and testis, while the lower levels were found in all different regions of brain, muscle and intestine. In the skin, IGFBP-1 mRNA expression level was extremely low. The IGFBP-1 mRNA expression level in liver was significantly elevated after feeding. With cadmium exposure for 24 h, IGFBP-1 mRNA expression levels in spleen and liver were significantly increased at different cadmium concentrations ranging from 0.5 to 10 ppm. The results in this study provided the data regarding molecular characteristics and expression patterns of IGFBP-1 in goldfish and showed that the expression of IGFBP-1 mRNA might be associated with metabolic status and heavy metal stress and regulated by metabolic factors and cadmium in fish. PMID:26753895

  15. Oxygen radicals as second messengers for expression of the monocyte chemoattractant protein, JE/MCP-1, and the monocyte colony-stimulating factor, CSF-1, in response to tumor necrosis factor-alpha and immunoglobulin G. Evidence for involvement of reduced nicotinamide adenine dinucleotide phosphate (NADPH)-dependent oxidase.

    PubMed

    Satriano, J A; Shuldiner, M; Hora, K; Xing, Y; Shan, Z; Schlondorff, D

    1993-09-01

    The potential involvement of reactive oxygen species in the expression of genes involved in immune response was examined in mesangial cells. Tumor necrosis factor (TNF-alpha) and aggregated (aggr.) IgG increased mRNA levels for the monocyte chemoattractant protein, JE/MCP-1, and the colony-stimulating factor, CSF-1. Scavengers for free radicals such as di- and tetra-methylthiourea (DMTU and TMTU) attenuated the increase in mRNA levels in response to TNF-alpha and aggr. IgG. Generation of superoxide anion by xanthine oxidase and hypoxanthine increased mRNA levels of these genes, but exogenous H2O2 did not. Addition of NADPH to activate a membrane-bound NADPH-oxidase generated superoxide and caused a dose-dependent increase in mRNA levels and further enhanced the stimulation by TNF-alpha or aggr. IgG. An inhibitor of NADPH-dependent oxidase 4'-hydroxy-3'-methoxy-acetophenone attenuated the rise in mRNA levels in response to TNF-alpha and aggr. IgG. By nuclear run-on experiments TNF-alpha, aggr. IgG and NADPH increased the transcription rates for JE/MCP-1 and CSF-1, effects inhibited by TMTU. We conclude that generation of reactive oxygen species, possibly by NADPH-dependent oxidase, are involved in the induction of the JE/MCP-1 and CSF-1 genes by TNF-alpha and IgG complexes. The concerted expression of leukocyte-directed cytokines represents a general response to tissue injury.

  16. The Neutrophil's Eye-View: Inference and Visualisation of the Chemoattractant Field Driving Cell Chemotaxis In Vivo

    PubMed Central

    Kadirkamanathan, Visakan; Anderson, Sean R.; Billings, Stephen A.; Zhang, Xiliang; Holmes, Geoffrey R.; Reyes-Aldasoro, Constantino C.; Elks, Philip M.; Renshaw, Stephen A.

    2012-01-01

    As we begin to understand the signals that drive chemotaxis in vivo, it is becoming clear that there is a complex interplay of chemotactic factors, which changes over time as the inflammatory response evolves. New animal models such as transgenic lines of zebrafish, which are near transparent and where the neutrophils express a green fluorescent protein, have the potential to greatly increase our understanding of the chemotactic process under conditions of wounding and infection from video microscopy data. Measurement of the chemoattractants over space (and their evolution over time) is a key objective for understanding the signals driving neutrophil chemotaxis. However, it is not possible to measure and visualise the most important contributors to in vivo chemotaxis, and in fact the understanding of the main contributors at any particular time is incomplete. The key insight that we make in this investigation is that the neutrophils themselves are sensing the underlying field that is driving their action and we can use the observations of neutrophil movement to infer the hidden net chemoattractant field by use of a novel computational framework. We apply the methodology to multiple in vivo neutrophil recruitment data sets to demonstrate this new technique and find that the method provides consistent estimates of the chemoattractant field across the majority of experiments. The framework that we derive represents an important new methodology for cell biologists investigating the signalling processes driving cell chemotaxis, which we label the neutrophils eye-view of the chemoattractant field. PMID:22563379

  17. Gene expression profile of zeitlupe/lov kelch protein1 T-DNA insertion mutants in Arabidopsis thaliana: Downregulation of auxin-inducible genes in hypocotyls

    PubMed Central

    Saitoh, Aya; Takase, Tomoyuki; Kitaki, Hiroyuki; Miyazaki, Yuji; Kiyosue, Tomohiro

    2015-01-01

    Elongation of hypocotyl cells has been studied as a model for elucidating the contribution of cellular expansion to plant organ growth. ZEITLUPE (ZTL) or LOV KELCH PROTEIN1 (LKP1) is a positive regulator of warmth-induced hypocotyl elongation under white light in Arabidopsis, although the molecular mechanisms by which it promotes hypocotyl cell elongation remain unknown. Microarray analysis showed that 134 genes were upregulated and 204 genes including 15 auxin-inducible genes were downregulated in the seedlings of 2 ztl T-DNA insertion mutants grown under warm conditions with continuous white light. Application of a polar auxin transport inhibitor, an auxin antagonist or an auxin biosynthesis inhibitor inhibited hypocotyl elongation of control seedlings to the level observed with the ztl mutant. Our data suggest the involvement of auxin and auxin-inducible genes in ZTL-mediated hypocotyl elongation. PMID:26237185

  18. Progranulin Is a Chemoattractant for Microglia and Stimulates Their Endocytic Activity

    PubMed Central

    Pickford, Fiona; Marcus, Jacob; Camargo, Luiz Miguel; Xiao, Qiurong; Graham, Danielle; Mo, Jan-Rung; Burkhardt, Matthew; Kulkarni, Vinayak; Crispino, Jamie; Hering, Heike; Hutton, Michael

    2011-01-01

    Mutations resulting in progranulin haploinsufficiency cause disease in patients with a subset of frontotemporal lobar degeneration; however, the biological functions of progranulin in the brain remain unknown. To address this subject, the present study initially assessed changes in gene expression and cytokine secretion in rat primary cortical neurons treated with progranulin. Molecular pathways enriched in the progranulin gene set included cell adhesion and cell motility pathways and pathways involved in growth and development. Secretion of cytokines and several chemokines linked to chemoattraction but not inflammation were also increased from progranulin-treated primary neurons. Therefore, whether progranulin is involved in recruitment of immune cells in the brain was investigated. Localized lentiviral expression of progranulin in C57BL/6 mice resulted in an increase of Iba1-positive microglia around the injection site. Moreover, progranulin alone was sufficient to promote migration of primary mouse microglia in vitro. Primary microglia and C4B8 cells demonstrated more endocytosis of amyloid β1-42 when treated with progranulin. These data demonstrate that progranulin acts as a chemoattractant in the brain to recruit or activate microglia and can increase endocytosis of extracellular peptides such as amyloid β. PMID:21224065

  19. Sperm navigation along helical paths in 3D chemoattractant landscapes.

    PubMed

    Jikeli, Jan F; Alvarez, Luis; Friedrich, Benjamin M; Wilson, Laurence G; Pascal, René; Colin, Remy; Pichlo, Magdalena; Rennhack, Andreas; Brenker, Christoph; Kaupp, U Benjamin

    2015-01-01

    Sperm require a sense of direction to locate the egg for fertilization. They follow gradients of chemical and physical cues provided by the egg or the oviduct. However, the principles underlying three-dimensional (3D) navigation in chemical landscapes are unknown. Here using holographic microscopy and optochemical techniques, we track sea urchin sperm navigating in 3D chemoattractant gradients. Sperm sense gradients on two timescales, which produces two different steering responses. A periodic component, resulting from the helical swimming, gradually aligns the helix towards the gradient. When incremental path corrections fail and sperm get off course, a sharp turning manoeuvre puts sperm back on track. Turning results from an 'off' Ca(2+) response signifying a chemoattractant stimulation decrease and, thereby, a drop in cyclic GMP concentration and membrane voltage. These findings highlight the computational sophistication by which sperm sample gradients for deterministic klinotaxis. We provide a conceptual and technical framework for studying microswimmers in 3D chemical landscapes. PMID:26278469

  20. Sperm navigation along helical paths in 3D chemoattractant landscapes

    NASA Astrophysics Data System (ADS)

    Jikeli, Jan F.; Alvarez, Luis; Friedrich, Benjamin M.; Wilson, Laurence G.; Pascal, René; Colin, Remy; Pichlo, Magdalena; Rennhack, Andreas; Brenker, Christoph; Kaupp, U. Benjamin

    2015-08-01

    Sperm require a sense of direction to locate the egg for fertilization. They follow gradients of chemical and physical cues provided by the egg or the oviduct. However, the principles underlying three-dimensional (3D) navigation in chemical landscapes are unknown. Here using holographic microscopy and optochemical techniques, we track sea urchin sperm navigating in 3D chemoattractant gradients. Sperm sense gradients on two timescales, which produces two different steering responses. A periodic component, resulting from the helical swimming, gradually aligns the helix towards the gradient. When incremental path corrections fail and sperm get off course, a sharp turning manoeuvre puts sperm back on track. Turning results from an `off' Ca2+ response signifying a chemoattractant stimulation decrease and, thereby, a drop in cyclic GMP concentration and membrane voltage. These findings highlight the computational sophistication by which sperm sample gradients for deterministic klinotaxis. We provide a conceptual and technical framework for studying microswimmers in 3D chemical landscapes.

  1. Screening and formulation of chemoattractant coatings for artificial reef structures.

    PubMed

    Lee, Han Seong; Sidharthan, M; Shim, Cheol Soo; Kim, Young Do; Lim, Chi Young; Ko, J W; Han, Man Deuk; Rang, Maeng Joo; Bim, Lee Sae; Cho, Hwan Sung; Shin, H W

    2008-07-01

    This study was carried out to augment the colonization of marine benthic communities on artificial reef structure. Increasing marine pollution along with various natural hazards cause severe damages to marine algae and associated fauna. In recent years, artificial reefs have been deployed in coastal regions of several parts of the world in order to increase the marine productivity. They are mainly built with concrete materials, however their leachates have considerable impacts on algae. Therefore to increase the algal colonization five chemoattractants such as ferrous sulfate, zinc oxide, ammonium nitrate, sodium phosphate and ferrous lactate were screened against spores of a fouling alga, Ulva pertusa. FeSO4 / ZnO (8:2) and ferrous lactate coatings showed the highest spore attachment with 52 +/- 5.2 cm2 and 79.5 +/- 10.2 cm2 spores respectively (p<0.01). Furthermore using these chemoattractants, coating formulations were made and their performances were investigated at East coast (Ayajin harbor) and South coast (Meejo harbor) of Korea. A maximum fouling coverage (with green algae 25%, red algae 11.3% and brown algae 63.7%) was estimated from ferrous lactate coatings (p<0.01). Different composition of coating formulations and their chemoattractive properties were evaluated. PMID:19195405

  2. Screening and formulation of chemoattractant coatings for artificial reef structures.

    PubMed

    Lee, Han Seong; Sidharthan, M; Shim, Cheol Soo; Kim, Young Do; Lim, Chi Young; Ko, J W; Han, Man Deuk; Rang, Maeng Joo; Bim, Lee Sae; Cho, Hwan Sung; Shin, H W

    2008-07-01

    This study was carried out to augment the colonization of marine benthic communities on artificial reef structure. Increasing marine pollution along with various natural hazards cause severe damages to marine algae and associated fauna. In recent years, artificial reefs have been deployed in coastal regions of several parts of the world in order to increase the marine productivity. They are mainly built with concrete materials, however their leachates have considerable impacts on algae. Therefore to increase the algal colonization five chemoattractants such as ferrous sulfate, zinc oxide, ammonium nitrate, sodium phosphate and ferrous lactate were screened against spores of a fouling alga, Ulva pertusa. FeSO4 / ZnO (8:2) and ferrous lactate coatings showed the highest spore attachment with 52 +/- 5.2 cm2 and 79.5 +/- 10.2 cm2 spores respectively (p<0.01). Furthermore using these chemoattractants, coating formulations were made and their performances were investigated at East coast (Ayajin harbor) and South coast (Meejo harbor) of Korea. A maximum fouling coverage (with green algae 25%, red algae 11.3% and brown algae 63.7%) was estimated from ferrous lactate coatings (p<0.01). Different composition of coating formulations and their chemoattractive properties were evaluated.

  3. Characterization of a chemoattractant for endothelium induced by angiogenesis effectors.

    PubMed

    Raju, K S; Alessandri, G; Gullino, P M

    1984-04-01

    The mechanism of neovascularization was further explored by the use of chemically defined angiogenesis effectors. The vascularization of the rabbit cornea was selected as an experimental approach that permits comparison of one cornea treated by the angiogenesis effector with the contralateral cornea of the same subject treated by the same molecule deprived of angiogenic capacity. Under these conditions, we observed that neovascularization was initiated by the appearance of a chemoattractant for the bovine capillary endothelium only in the cornea treated by the angiogenesis effector. The chemoattractant was purified about 150-fold by a single-step procedure, using gelatin:Sepharose affinity chromatography. Chemoattraction resulted from the combined effect of a chemotactic factor(s) and an activating factor(s). The association of the two enhanced 5- to 8-fold the motility of the capillary endothelium in a concentration-dependent manner with optimum at 0.2 mg/ml. The activating factor(s) does not have chemotactic capacity, but without it, chemotaxis is reduced to about one half. The chemotactic complex was present in the cornea regardless of the nature of the angiogenesis effector used as the triggering device. Heat and proteases eliminated chemotaxis and destroyed the chemotactic complex. Thus, neovascularization may be triggered by effectors able to induce in the cornea proteins, normally not present, that influence angiogenesis via mobilization of capillary endothelium. PMID:6200213

  4. Key role of regulated upon activation normal T-cell expressed and secreted, nonstructural protein1 and myeloperoxidase in cytokine storm induced by influenza virus PR-8 (A/H1N1) infection in A549 bronchial epithelial cells.

    PubMed

    Phung, Thuy Thi Bich; Sugamata, Ryuichi; Uno, Kazuko; Aratani, Yasuaki; Ozato, Keiko; Kawachi, Shoji; Thanh Nguyen, Liem; Nakayama, Toshinori; Suzuki, Kazuo

    2011-12-01

    Influenza virus infection causes severe respiratory disease such as that due to avian influenza (H5N1). Influenza A viruses proliferate in human epithelial cells, which produce inflammatory cytokines/chemokines as a "cytokine storm" attenuated with the viral nonstructural protein 1 (NS1). Cytokine/chemokine production in A549 epithelial cells infected with influenza A/H1N1 virus (PR-8) or nonstructural protein 1 (NS1) plasmid was examined in vitro. Because tumor necrosis factor-α (TNF-α) and regulated upon activation normal T-cell expressed and secreted (RANTES) are predominantly produced from cells infected with PR-8 virus, the effects of mRNA knockdown of these cytokines were investigated. Small interfering (si)TNF-α down-regulated RANTES expression and secretion of RANTES, interleukin (IL)-8, and monocyte chemotactic protein-1 (MCP-1). In addition, siRANTES suppressed interferon (IFN)-γ expression and secretion of RANTES, IL-8, and MCP-1, suggesting that TNF-α stimulates production of RANTES, IL-8, MCP-1, and IFN-γ, and RANTES also increased IL-8, MCP-1, and IFN-γ. Furthermore, administration of TNF-α promoted increased secretion of RANTES, IL-8, and MCP-1. Administration of RANTES enhanced IL-6, IL-8, and MCP-1 production without PR-8 infection. These results strongly suggest that, as an initial step, TNF-α regulates RANTES production, followed by increase of IL-6, IL-8, and MCP-1 and IFNs concentrations. At a later stage, cells transfected with viral NS1 plasmid showed production of a large amount of IL-8 and MCP-1 in the presence of the H(2)O(2)-myeloperoxidse (MPO) system, suggesting that NS1 of PR-8 may induce a "cytokine storm" from epithelial cells in the presence of an H(2)O(2)-MPO system.

  5. Elevated Dengue Virus Nonstructural Protein 1 Serum Levels and Altered Toll-Like Receptor 4 Expression, Nitric Oxide, and Tumor Necrosis Factor Alpha Production in Dengue Hemorrhagic Fever Patients

    PubMed Central

    Carvalho, Denise Maciel; Garcia, Fernanda Gonçalves; Terra, Ana Paula Sarreta; Lopes Tosta, Ana Cristina; Silva, Luciana de Almeida; Castellano, Lúcio Roberto; Silva Teixeira, David Nascimento

    2014-01-01

    Background. During dengue virus (DV) infection, monocytes produce tumor necrosis factor alpha (TNF-α) and nitric oxide (NO) which might be critical to immunopathogenesis. Since intensity of DV replication may determine clinical outcomes, it is important to know the effects of viral nonstructural protein 1 (NS1) on innate immune parameters of infected patients. The present study investigates the relationships between dengue virus nonstructural protein 1 (NS1) serum levels and innate immune response (TLR4 expression and TNF-α/NO production) of DV infected patients presenting different clinical outcomes. Methodology/Principal Findings. We evaluated NO, NS1 serum levels (ELISA), TNF-α production by peripheral blood mononuclear cells (PBMCs), and TLR4 expression on CD14+ cells from 37 dengue patients and 20 healthy controls. Early in infection, increased expression of TLR4 in monocytes of patients with dengue fever (DF) was detected compared to patients with dengue hemorrhagic fever (DHF). Moreover, PBMCs of DHF patients showed higher NS1 and lower NO serum levels during the acute febrile phase and a reduced response to TLR4 stimulation by LPS (with a reduced TNF-α production) when compared to DF patients. Conclusions/Significance. During DV infection in humans, some innate immune parameters change, depending on the NS1 serum levels, and phase and severity of the disease which may contribute to development of different clinical outcomes. PMID:25580138

  6. Expression of Estrogen Receptor Coactivator Proline-, Glutamic Acid- and Leucine-Rich Protein 1 within Paraspinal Muscles in Adolescents with Idiopathic Scoliosis

    PubMed Central

    Skibinska, Izabela; Tomaszewski, Marek; Andrusiewicz, Miroslaw; Urbaniak, Paulina; Czarnecka-Klos, Roza; Shadi, Milud; Kotwicki, Tomasz; Kotwicka, Malgorzata

    2016-01-01

    Purpose The aim of this study was to detect and assess the estrogen receptor (ESR) coactivator PELP1 expression within human paraspinal skeletal muscles in patients suffering from idiopathic scoliosis. Methods During surgical correction of scoliosis the muscle biopsies harvested in 29 females. Presence of PELP1, ESR1 and ESR2 genes transcripts was studied using RT-qPCR technique while immunohistochemistry and western blot methods were used to detect the PEPL1 protein presence. Results PELP1 expression in deep paraspinal muscles revealed higher than in superficial back muscles (p = 0.005). Positive immunohistochemical staining for PELP1 was observed in the nuclei of the paraspinal muscle cells. Western blot revealed PELP1 protein in all samples. No significant difference in PELP1 expression between the convex and the concave scoliosis side (p>0.05) was found. In deep paraspinal back muscles, a significant correlation between the PELP1 expression level on the concave side and the Cobb angle (r = 0.4; p<0.05) was noted as well as between the PELP1 and ESR1 expression level (r = 0.7; p<0.05) while no correlation between PELP1 and ESR2 expression level was found. Conclusion To our knowledge, three techniques for the first time demonstrated the presence of the PELP1 in paraspinal muscles of patients with idiopathic scoliosis. The PELP1 potential regulatory impact on back muscle function is to be further investigated. PMID:27045366

  7. Two Types of Assays for Detecting Frog Sperm Chemoattraction

    PubMed Central

    Burnett, Lindsey A.; Tholl, Nathan; Chandler, Douglas E.

    2011-01-01

    Sperm chemoattraction in invertebrates can be sufficiently robust that one can place a pipette containing the attractive peptide into a sperm suspension and microscopically visualize sperm accumulation around the pipette1. Sperm chemoattraction in vertebrates such as frogs, rodents and humans is more difficult to detect and requires quantitative assays. Such assays are of two major types - assays that quantitate sperm movement to a source of chemoattractant, so-called sperm accumulation assays, and assays that actually track the swimming trajectories of individual sperm. Sperm accumulation assays are relatively rapid allowing tens or hundreds of assays to be done in a single day, thereby allowing dose response curves and time courses to be carried out relatively rapidly. These types of assays have been used extensively to characterize many well established chemoattraction systems - for example, neutrophil chemotaxis to bacterial peptides and sperm chemotaxis to follicular fluid. Sperm tracking assays can be more labor intensive but offer additional data on how chemoattractancts actually alter the swimming paths that sperm take. This type of assay is needed to demonstrate the orientation of sperm movement relative to the chemoattrractant gradient axis and to visualize characteristic turns or changes in orientation that bring the sperm closer to the egg. Here we describe methods used for each of these two types of assays. The sperm accumulation assay utilized is called a "two-chamber" assay. Amphibian sperm are placed in a tissue culture plate insert with a polycarbonate filter floor having 12 μm diameter pores. Inserts with sperm are placed into tissue culture plate wells containing buffer and a chemoatttractant carefully pipetted into the bottom well where the floor meets the wall (see Fig. 1). After incubation, the top insert containing the sperm reservoir is carefully removed, and sperm in the bottom chamber that have passed through the membrane are removed

  8. Enhanced Tolerance of Transgenic Potato Plants Over-Expressing Non-specific Lipid Transfer Protein-1 (StnsLTP1) against Multiple Abiotic Stresses.

    PubMed

    Gangadhar, Baniekal H; Sajeesh, Kappachery; Venkatesh, Jelli; Baskar, Venkidasamy; Abhinandan, Kumar; Yu, Jae W; Prasad, Ram; Mishra, Raghvendra K

    2016-01-01

    Abiotic stresses such as heat, drought, and salinity are major environmental constraints that limit potato (Solanum tuberosum L.) production worldwide. Previously, we found a potential thermo-tolerance gene, named StnsLTP1 from potato using yeast functional screening. Here, we report the functional characterization of StnsLTP1 and its role in multiple abiotic stresses in potato plants. Computational analysis of StnsLTP1 with other plant LTPs showed eight conserved cysteine residues, and four α-helices stabilized by four disulfide bridges. Expression analysis of StnsLTP1 gene showed differential expression under heat, water-deficit and salt stresses. Transgenic potato lines over-expressing StnsLTP1 gene displayed enhanced cell membrane integrity under stress conditions, as indicated by reduced membrane lipid per-oxidation, and hydrogen peroxide content relative to untransformed (UT) control plants. In addition, transgenic lines over-expressing StLTP1 also exhibited increased antioxidant enzyme activity with enhanced accumulation of ascorbates, and up-regulation of stress-related genes including StAPX, StCAT, StSOD, StHsfA3, StHSP70, and StsHSP20 compared with the UT plants. These results suggests that StnsLTP1 transgenic plants acquired improved tolerance to multiple abiotic stresses through enhanced activation of antioxidative defense mechanisms via cyclic scavenging of reactive oxygen species and regulated expression of stress-related genes. PMID:27597854

  9. Enhanced Tolerance of Transgenic Potato Plants Over-Expressing Non-specific Lipid Transfer Protein-1 (StnsLTP1) against Multiple Abiotic Stresses

    PubMed Central

    Gangadhar, Baniekal H.; Sajeesh, Kappachery; Venkatesh, Jelli; Baskar, Venkidasamy; Abhinandan, Kumar; Yu, Jae W.; Prasad, Ram; Mishra, Raghvendra K.

    2016-01-01

    Abiotic stresses such as heat, drought, and salinity are major environmental constraints that limit potato (Solanum tuberosum L.) production worldwide. Previously, we found a potential thermo-tolerance gene, named StnsLTP1 from potato using yeast functional screening. Here, we report the functional characterization of StnsLTP1 and its role in multiple abiotic stresses in potato plants. Computational analysis of StnsLTP1 with other plant LTPs showed eight conserved cysteine residues, and four α-helices stabilized by four disulfide bridges. Expression analysis of StnsLTP1 gene showed differential expression under heat, water-deficit and salt stresses. Transgenic potato lines over-expressing StnsLTP1 gene displayed enhanced cell membrane integrity under stress conditions, as indicated by reduced membrane lipid per-oxidation, and hydrogen peroxide content relative to untransformed (UT) control plants. In addition, transgenic lines over-expressing StLTP1 also exhibited increased antioxidant enzyme activity with enhanced accumulation of ascorbates, and up-regulation of stress-related genes including StAPX, StCAT, StSOD, StHsfA3, StHSP70, and StsHSP20 compared with the UT plants. These results suggests that StnsLTP1 transgenic plants acquired improved tolerance to multiple abiotic stresses through enhanced activation of antioxidative defense mechanisms via cyclic scavenging of reactive oxygen species and regulated expression of stress-related genes. PMID:27597854

  10. Enhanced Tolerance of Transgenic Potato Plants Over-Expressing Non-specific Lipid Transfer Protein-1 (StnsLTP1) against Multiple Abiotic Stresses

    PubMed Central

    Gangadhar, Baniekal H.; Sajeesh, Kappachery; Venkatesh, Jelli; Baskar, Venkidasamy; Abhinandan, Kumar; Yu, Jae W.; Prasad, Ram; Mishra, Raghvendra K.

    2016-01-01

    Abiotic stresses such as heat, drought, and salinity are major environmental constraints that limit potato (Solanum tuberosum L.) production worldwide. Previously, we found a potential thermo-tolerance gene, named StnsLTP1 from potato using yeast functional screening. Here, we report the functional characterization of StnsLTP1 and its role in multiple abiotic stresses in potato plants. Computational analysis of StnsLTP1 with other plant LTPs showed eight conserved cysteine residues, and four α-helices stabilized by four disulfide bridges. Expression analysis of StnsLTP1 gene showed differential expression under heat, water-deficit and salt stresses. Transgenic potato lines over-expressing StnsLTP1 gene displayed enhanced cell membrane integrity under stress conditions, as indicated by reduced membrane lipid per-oxidation, and hydrogen peroxide content relative to untransformed (UT) control plants. In addition, transgenic lines over-expressing StLTP1 also exhibited increased antioxidant enzyme activity with enhanced accumulation of ascorbates, and up-regulation of stress-related genes including StAPX, StCAT, StSOD, StHsfA3, StHSP70, and StsHSP20 compared with the UT plants. These results suggests that StnsLTP1 transgenic plants acquired improved tolerance to multiple abiotic stresses through enhanced activation of antioxidative defense mechanisms via cyclic scavenging of reactive oxygen species and regulated expression of stress-related genes.

  11. Enhanced Tolerance of Transgenic Potato Plants Over-Expressing Non-specific Lipid Transfer Protein-1 (StnsLTP1) against Multiple Abiotic Stresses.

    PubMed

    Gangadhar, Baniekal H; Sajeesh, Kappachery; Venkatesh, Jelli; Baskar, Venkidasamy; Abhinandan, Kumar; Yu, Jae W; Prasad, Ram; Mishra, Raghvendra K

    2016-01-01

    Abiotic stresses such as heat, drought, and salinity are major environmental constraints that limit potato (Solanum tuberosum L.) production worldwide. Previously, we found a potential thermo-tolerance gene, named StnsLTP1 from potato using yeast functional screening. Here, we report the functional characterization of StnsLTP1 and its role in multiple abiotic stresses in potato plants. Computational analysis of StnsLTP1 with other plant LTPs showed eight conserved cysteine residues, and four α-helices stabilized by four disulfide bridges. Expression analysis of StnsLTP1 gene showed differential expression under heat, water-deficit and salt stresses. Transgenic potato lines over-expressing StnsLTP1 gene displayed enhanced cell membrane integrity under stress conditions, as indicated by reduced membrane lipid per-oxidation, and hydrogen peroxide content relative to untransformed (UT) control plants. In addition, transgenic lines over-expressing StLTP1 also exhibited increased antioxidant enzyme activity with enhanced accumulation of ascorbates, and up-regulation of stress-related genes including StAPX, StCAT, StSOD, StHsfA3, StHSP70, and StsHSP20 compared with the UT plants. These results suggests that StnsLTP1 transgenic plants acquired improved tolerance to multiple abiotic stresses through enhanced activation of antioxidative defense mechanisms via cyclic scavenging of reactive oxygen species and regulated expression of stress-related genes.

  12. Expression of Tax-interacting protein 1 (TIP-1) facilitates angiogenesis and tumor formation of human glioblastoma cells in nude mice

    PubMed Central

    Han, Miaojun; Wang, Hailun; Zhang, Hua-Tang; Han, Zhaozhong

    2012-01-01

    Glioblastoma is the most common and fatal type of primary brain tumors featured with hyperplastic blood vessels. Here, we performed meta-analyses of published data and established a correlation between high TIP-1 expression levels and the poor prognosis of glioblastoma patients. Next, we explored the biological relevance of TIP-1 expression in the pathogenesis of glioblastoma. By using orthotopic and heterotopic mouse models of human glioblastomas, this study has characterized TIP-1 as one contributing factor to the tumor-driven angiogenesis. In vitro and in vivo functional assays, along with biochemical analyses with microarrays and antibody arrays, have demonstrated that TIP-1 utilizes multiple pathways including modulating fibronectin gene expression and uPA protein secretion, to establish or maintain a pro-angiogenic microenvironment within human glioblastoma. In conclusion, this work supports the hypothesis that TIP-1 represents a novel prognostic biomarker and a therapeutic target of human glioblastoma. PMID:23010083

  13. Macrophage-like tumor cells as tools to study chemoattractive activity.

    PubMed

    Terheggen, P; Van Loveren, H; Den Otter, W

    1985-12-01

    Macrophage-like tumor cells can be obtained in large quantities as rather homogeneous populations, making these cells useful for chemotaxis assays. Therefore, macrophage-like cells J774A, WEHI-3, P388D1, IC-21, and NCTC 1469, all of murine origin, and U937 of human origin, were tested for chemotactic activity to a number of chemoattractive agents, such as casein, an N-formyl tetrapeptide (N-formyl-L-norleucyl-L-leucyl-L-phenylalanyl-L-tyrosine), and culture supernatants of murine SL2 lymphoma cells. J774A and WEHI-3 macrophage-like cells of murine (BALB/c) origin expressed the strongest chemotactic activity to casein and N-formyl tetrapeptide, respectively. The results show that: very standardized chemotaxis assays can be performed using these cell lines; these assays require appropriate cell line-stimulus combinations; there are substantial differences among cell lines as to sensitivity to various chemoattractive substances; macrophage cell lines and functional mutants may be helpful for the study of receptors for chemotaxins and the study of transducer signals for chemotaxis. PMID:3864910

  14. Extracellular galectin-3 counteracts adhesion and exhibits chemoattraction in Helicobacter pylori-infected gastric cancer cells.

    PubMed

    Subhash, Vinod Vijay; Ling, Samantha Shi Min; Ho, Bow

    2016-08-01

    Galectin-3 (Gal-3) is a β-galactoside lectin that is upregulated and rapidly secreted by gastric epithelial cells in response to Helicobacter pylori infection. An earlier study reported the involvement of H. pylori cytotoxin-associated gene A (cagA) in the expression of intracellular Gal-3. However, the role of extracellular Gal-3 and its functional significance in H. pylori-infected cells remains uncharacterized. Data presented here demonstrate secretion of Gal-3 is an initial host response event in gastric epithelial cells during H. pylori infection and is independent of CagA. Previously, Gal-3 was shown to bind to H. pylori LPS. The present study elaborates the significance of this binding, as extracellular recombinant Gal-3 (rGal-3) was shown to inhibit the adhesion of H. pylori to the gastric epithelial cells. Interestingly, a decrease in H. pylori adhesion to host cells also resulted in a decrease in apoptosis. Furthermore, the study also demonstrated a chemoattractant role of extracellular rGal-3 in the recruitment of THP-1 monocytes. This study outlines the previously unidentified roles of extracellular Gal-3 where it acts as a negative regulator of H. pylori adhesion and apoptosis in gastric epithelial cells, and as a chemoattractant to THP-1 monocytes. Our findings could contribute to the better understanding of how Gal-3 acts as a modulator under H. pylori-induced pathological conditions. PMID:27283429

  15. Phosphorylation of chemoattractant receptors regulates chemotaxis, actin reorganization and signal relay.

    PubMed

    Brzostowski, Joseph A; Sawai, Satoshi; Rozov, Orr; Liao, Xin-Hua; Imoto, Daisuke; Parent, Carole A; Kimmel, Alan R

    2013-10-15

    Migratory cells, including mammalian leukocytes and Dictyostelium, use G-protein-coupled receptor (GPCR) signaling to regulate MAPK/ERK, PI3K, TORC2/AKT, adenylyl cyclase and actin polymerization, which collectively direct chemotaxis. Upon ligand binding, mammalian GPCRs are phosphorylated at cytoplasmic residues, uncoupling G-protein pathways, but activating other pathways. However, connections between GPCR phosphorylation and chemotaxis are unclear. In developing Dictyostelium, secreted cAMP serves as a chemoattractant, with extracellular cAMP propagated as oscillating waves to ensure directional migratory signals. cAMP oscillations derive from transient excitatory responses of adenylyl cyclase, which then rapidly adapts. We have studied chemotactic signaling in Dictyostelium that express non-phosphorylatable cAMP receptors and show through chemotaxis modeling, single-cell FRET imaging, pure and chimeric population wavelet quantification, biochemical analyses and TIRF microscopy, that receptor phosphorylation is required to regulate adenylyl cyclase adaptation, long-range oscillatory cAMP wave production and cytoskeletal actin response. Phosphorylation defects thus promote hyperactive actin polymerization at the cell periphery, misdirected pseudopodia and the loss of directional chemotaxis. Our data indicate that chemoattractant receptor phosphorylation is required to co-regulate essential pathways for migratory cell polarization and chemotaxis. Our results significantly extend the understanding of the function of GPCR phosphorylation, providing strong evidence that this evolutionarily conserved mechanism is required in a signal attenuation pathway that is necessary to maintain persistent directional movement of Dictyostelium, neutrophils and other migratory cells.

  16. The aqueous extract of Hibiscus sabdariffa calices modulates the production of monocyte chemoattractant protein-1 in humans.

    PubMed

    Beltrán-Debón, R; Alonso-Villaverde, C; Aragonès, G; Rodríguez-Medina, I; Rull, A; Micol, V; Segura-Carretero, A; Fernández-Gutiérrez, A; Camps, J; Joven, J

    2010-03-01

    Diet supplementation and/or modulation is an important strategy to significantly improve human health. The search of plants as additional sources of bioactive phenolic compounds is relevant in this context. The aqueous extract of Hibiscus sabdariffa is rich in anthocyanins and other phenolic compounds including hydroxycitric and chlorogenic acids. Using this extract we have shown an effective protection of cultured peripheral blood mononuclear cells from the cellular death induced by H(2)O(2) and a significant role in the production of inflammatory cytokines. In vitro, the extract promotes the production of IL-6 and IL-8 and decreases the concentration of MCP-1 in supernatants in a dose-dependent manner. In humans, the ingestion of an acute dose of the extract (10g) was well tolerated and decreased plasma MCP-1 concentrations significantly without further effects on other cytokines. This effect was not due to a concomitant increase in the antioxidant capacity of plasma. Instead, its mechanisms probably involve a direct inhibition of inflammatory and/or metabolic pathways responsible for MCP-1 production, and may be relevant in inflammatory and chronic conditions in which the role of MCP-1 is well established. If beneficial effects are confirmed in patients, Hibiscus sabdariffa could be considered a valuable traditional herbal medicine for the treatment of chronic inflammatory diseases with the advantage of being devoid of caloric value or potential alcohol toxicity. PMID:19765963

  17. Expression of a gene for a protein similar to HIV-1 Tat binding protein 1 (TBP1) in floral organs of Brassica rapa.

    PubMed

    Kitashiba, H; Toriyama, K

    1997-08-01

    A cDNA for a protein similar to human immunodeficiency virus Tat binding protein was isolated from an anther cDNA library of Brassica rapa. RNA in situ analysis in flower buds showed that the gene for this cDNA was specifically expressed in the tapetum and middle layer of anthers and pollen.

  18. Involvement of miR-326 in chemotherapy resistance of breast cancer through modulating expression of multidrug resistance-associated protein 1.

    PubMed

    Liang, Zhongxing; Wu, Hui; Xia, James; Li, Yuhua; Zhang, Yawei; Huang, Ke; Wagar, Nicholas; Yoon, Younghyoun; Cho, Heidi T; Scala, Stefania; Shim, Hyunsuk

    2010-03-15

    Multidrug resistance-associated protein (MRP-1/ABCC1) transports a wide range of therapeutic agents and may play a critical role in the development of multidrug resistance (MDR) in tumor cells. However, the regulation of MRP-1 remains controversial. To explore whether miRNAs are involved in the regulation of MRP-1 expression and modulate the sensitivity of tumor cells to chemotherapeutic agents, we analyzed miRNA expression levels in VP-16-resistant MDR cell line, MCF-7/VP, in comparison with its parent cell line, MCF-7, using a miRNA microarray. MCF-7/VP overexpressed MRP-1 mRNA and protein not MDR-1 and BCRP. miR-326 was downregulated in MCF-7/VP compared to MCF-7. Additionally, miR-326 was downregulated in a panel of advanced breast cancer tissues and consistent reversely with expression levels of MRP-1. Furthermore, the elevated levels of miR-326 in the mimics-transfected VP-16-resistant cell line, MCF-7/VP, downregulated MRP-1 expression and sensitized these cells to VP-16 and doxorubicin. These findings demonstrate for the first time the involvement of miRNAs in multidrug resistance mediated by MRP-1 and suggest that miR-326 may be an efficient agent for preventing and reversing MDR in tumor cells.

  19. Role of NF-kappa B in cell survival and transcription of latent membrane protein 1-expressing or Epstein-Barr virus latency III-infected cells.

    PubMed

    Cahir-McFarland, Ellen D; Carter, Kara; Rosenwald, Andreas; Giltnane, Jena M; Henrickson, Sarah E; Staudt, Louis M; Kieff, Elliott

    2004-04-01

    Epstein-Barr virus (EBV) latency III infection converts B lymphocytes into lymphoblastoid cell lines (LCLs) by expressing EBV nuclear and membrane proteins, EBNAs, and latent membrane proteins (LMPs), which regulate transcription through Notch and tumor necrosis factor receptor pathways. The role of NF-kappa B in LMP1 and overall EBV latency III transcriptional effects was investigated by treating LCLs with BAY11-7082 (BAY11). BAY11 rapidly and irreversibly inhibited NF-kappa B, decreased mitochondrial membrane potential, induced apoptosis, and altered LCL gene expression. BAY11 effects were similar to those of an NF-kappa B inhibitor, Delta N-I kappa B alpha, in effecting decreased JNK1 expression and in microarray analyses. More than 80% of array elements that decreased with Delta N-I kappa B alpha expression decreased with BAY11 treatment. Newly identified NF-kappa B-induced, LMP1-induced, and EBV-induced genes included pleckstrin, Jun-B, c-FLIP, CIP4, and I kappa B epsilon. Of 776 significantly changed array elements, 134 were fourfold upregulated in EBV latency III, and 74 were fourfold upregulated with LMP1 expression alone, whereas only 28 were more than fourfold downregulated by EBV latency III. EBV latency III-regulated gene products mediate cell migration (EBI2, CCR7, RGS1, RANTES, MIP1 alpha, MIP1 beta, CXCR5, and RGS13), antigen presentation (major histocompatibility complex proteins and JAW1), mitogen-activated protein kinase pathway (DUSP5 and p62Dok), and interferon (IFN) signaling (IFN-gamma R alpha, IRF-4, and STAT1). Comparison of EBV latency III LCL gene expression to immunoglobulin M (IgM)-stimulated B cells, germinal-center B cells, and germinal-center-derived lymphomas clustered LCLs with IgM-stimulated B cells separately from germinal-center cells or germinal-center lymphoma cells. Expression of IRF-2, AIM1, ASK1, SNF2L2, and components of IFN signaling pathways further distinguished EBV latency III-infected B cells from IgM-stimulated or

  20. The rescue of dentin matrix protein 1 (DMP1)-deficient tooth defects by the transgenic expression of dentin sialophosphoprotein (DSPP) indicates that DSPP is a downstream effector molecule of DMP1 in dentinogenesis.

    PubMed

    Gibson, Monica Prasad; Zhu, Qinglin; Wang, Suzhen; Liu, Qilin; Liu, Ying; Wang, Xiaofang; Yuan, Baozhi; Ruest, L Bruno; Feng, Jian Q; D'Souza, Rena N; Qin, Chunlin; Lu, Yongbo

    2013-03-01

    Dentin matrix protein 1 (DMP1) and dentin sialophosphoprotein (DSPP) are essential for the formation of dentin. Previous in vitro studies have indicated that DMP1 might regulate the expression of DSPP during dentinogenesis. To examine whether DMP1 controls dentinogenesis through the regulation of DSPP in vivo, we cross-bred transgenic mice expressing normal DSPP driven by a 3.6-kb rat Col1a1 promoter with Dmp1 KO mice to generate mice expressing the DSPP transgene in the Dmp1 KO genetic background (referred to as "Dmp1 KO/DSPP Tg mice"). We used morphological, histological, and biochemical techniques to characterize the dentin and alveolar bone of Dmp1 KO/DSPP Tg mice compared with Dmp1 KO and wild-type mice. Our analyses showed that the expression of endogenous DSPP was remarkably reduced in the Dmp1 KO mice. Furthermore, the transgenic expression of DSPP rescued the tooth and alveolar bone defects of the Dmp1 KO mice. In addition, our in vitro analyses showed that DMP1 and its 57-kDa C-terminal fragment significantly up-regulated the Dspp promoter activities in a mesenchymal cell line. In contrast, the expression of DMP1 was not altered in the Dspp KO mice. These results provide strong evidence that DSPP is a downstream effector molecule that mediates the roles of DMP1 in dentinogenesis.

  1. Regulation of sterol regulatory-element binding protein 1 gene expression in liver: role of insulin and protein kinase B/cAkt.

    PubMed Central

    Fleischmann, M; Iynedjian, P B

    2000-01-01

    Insulin stimulates the transcription of the sterol regulatory- element binding protein (SREBP) 1/ADD1 gene in liver. Hepatocytes in primary culture were used to delineate the insulin signalling pathway for induction of SREBP1 gene expression. The inhibitors of phosphoinositide 3-kinase (PI 3-kinase), wortmannin and LY 294002, abolished the insulin-dependent increase in SREBP1 mRNA, whereas the inhibitor of the mitogen- activated protein kinase cascade, PD 98059, was without effect. To investigate the role of protein kinase B (PKB)/cAkt downstream of PI 3-kinase, hepatocytes were transduced with an adenovirus encoding a PKB--oestrogen receptor fusion protein. The PKB activity of this recombinant protein was rapidly activated in hepatocytes challenged with 4-hydroxytamoxifen (OHT), as was endogenous PKB in hepatocytes challenged with insulin. The addition of OHT to transduced hepatocytes resulted in accumulation of SREBP1 mRNA, with a time-course and magnitude similar to the effect of insulin in non-transduced cells. The level of SREBP1 mRNA was not increased by OHT in hepatocytes expressing a mutant form of the recombinant protein whose PKB activity was not activated by OHT. Thus acute activation of PKB is sufficient to induce SREBP1 mRNA accumulation in primary hepatocytes, and might be the major signalling event by which insulin induces SREBP1 gene expression in the liver. PMID:10861205

  2. Intraspecific and interspecific chemoattraction inBiomphalaria glabrata andHelisoma trivolvis (Gastropoda: Planorbidae).

    PubMed

    Marcopoulos, A A; Fried, B

    1994-10-01

    A Petri dish bioassay previously used to examine food preferences in planorbid snails was used to study intraspecific and interspecific chemoattraction inBiomphalaria glabrata (albino strain, M-line) andHelisoma trivolvis (Colorado strain) snails.B. glabrata snails showed significant intraspecific chemoattraction in the absence of visual cues and snail thigmotaxis.H. trivolvis snails also showed significant intraspecific chemoattraction. Interspecific chemoattraction between these species occurred in the bioassay, suggesting that the chemoattractants were not species specific. Artificial spring water conditioned by aqueous excretory-secretory products (snail-conditioned water) ofB. glabrata elicited significant intraspecific chemoattraction. However, lipophilic excretory-secretory products ofB. glabrata elicited significant chemorepulsion. Repellant factors in the lipophilic fraction were not characterized. PMID:24241838

  3. Reversal of phenol and naphthalene effects on ciliate chemoattraction

    SciTech Connect

    Berk, S.G.; Ting, R.S.; Roberts, R.O. ); Mills, B.A. ); Stewart, K.C. )

    1990-02-01

    In an effort to address research needs in the area of rapid screening tests for aquatic toxicology, the authors engaged in a study of pollutant effects on protozoan chemoattraction. Among pollutants tested were metals and hydrocarbons. To ascertain whether inhibition observed after brief exposures to certain concentrations of the pollutants were irreversible, they examined the possibility of nullifying the inhibitory effect by removing protozoa from the toxicants after short exposures. Earlier work showed that inhibitory effects of metals could be removed, and they report here the nullification and reversibility of effects of phenol and naphthalene on certain ciliates.

  4. Honokiol reverses alcoholic fatty liver by inhibiting the maturation of sterol regulatory element binding protein-1c and the expression of its downstream lipogenesis genes

    SciTech Connect

    Yin Huquan; Kim, Youn-Chul; Chung, Young-Suk; Kim, Young-Chul; Shin, Young-Kee; Lee, Byung-Hoon

    2009-04-01

    Ethanol induces hepatic steatosis via a complex mechanism that is not well understood. Among the variety of molecules that have been proposed to participate in this mechanism, the sterol regulatory element (SRE)-binding proteins (SREBPs) have been identified as attractive targets for therapeutic intervention. In the present study, we evaluated the effects of honokiol on alcoholic steatosis and investigated its possible effect on the inhibition of SREBP-1c maturation. In in vitro studies, H4IIEC3 rat hepatoma cells developed increased lipid droplets when exposed to ethanol, but co-treatment with honokiol reversed this effect. Honokiol inhibited the maturation of SREBP-1c and its translocation to the nucleus, the binding of nSREBP-1c to SRE or SRE-related sequences of its lipogenic target genes, and the expression of genes for fatty acid synthesis. In contrast, magnolol, a structural isomer of honokiol, had no effect on nSREBP-1c levels. Male Wistar rats fed with a standard Lieber-DeCarli ethanol diet for 4 weeks exhibited increased hepatic triglyceride and decreased hepatic glutathione levels, with concomitantly increased serum alanine aminotransferase and TNF-{alpha} levels. Daily administration of honokiol (10 mg/kg body weight) by gavage during the final 2 weeks of ethanol treatment completely reversed these effects on hepatotoxicity markers, including hepatic triglyceride, hepatic glutathione, and serum TNF-{alpha}, with efficacious abrogation of fat accumulation in the liver. Inhibition of SREBP-1c protein maturation and of the expression of Srebf1c and its target genes for hepatic lipogenesis were also observed in vivo. A chromatin immunoprecipitation assay demonstrated inhibition of specific binding of SREBP-1c to the Fas promoter by honokiol in vivo. These results demonstrate that honokiol has the potential to ameliorate alcoholic steatosis by blocking fatty acid synthesis regulated by SREBP-1c.

  5. Transport of antimony salts by Arabidopsis thaliana protoplasts over-expressing the human multidrug resistance-associated protein 1 (MRP1/ABCC1).

    PubMed

    Gayet, Landry; Picault, Nathalie; Cazalé, Anne-Claire; Beyly, Audrey; Lucas, Philippe; Jacquet, Hélène; Suso, Henri-Pierre; Vavasseur, Alain; Peltier, Gilles; Forestier, Cyrille

    2006-12-22

    ABC transporters from the multidrug resistance-associated protein (MRP) subfamily are glutathione S-conjugate pumps exhibiting a broad substrate specificity illustrated by numerous xenobiotics, such as anticancer drugs, herbicides, pesticides and heavy metals. The engineering of MRP transporters into plants might be interesting either to reduce the quantity of xenobiotics taken up by the plant in the context of "safe-food" strategies or, conversely, in the development of phytoremediation strategies in which xenobiotics are sequestered in the vacuolar compartment. In this report, we obtained Arabidopsis transgenic plants overexpressing human MRP1. In these plants, expression of MRP1 did not increase plant resistance to antimony salts (Sb(III)), a classical glutathione-conjugate substrate of MRP1. However, the transporter was fully translated in roots and shoots, and targeted to the plasma membrane. In order to investigate the functionality of MRP1 in Arabidopsis, mesophyll cell protoplasts (MCPs) were isolated from transgenic plants and transport activities were measured by using calcein or Sb(III) as substrates. Expression of MRP1 at the plasma membrane was correlated with an increase in the MCPs resistance to Sb(III) and a limitation of the metalloid content in the protoplasts due to an improvement in Sb(III) efflux. Moreover, Sb(III) transport was sensitive to classical inhibitors of the human MRP1, such as MK571 or glibenclamide. These results demonstrate that a human ABC transporter can be functionally introduced in Arabidopsis, which might be useful, with the help of stronger promoters, to reduce the accumulation of xenobiotics in plants, such as heavy metals from multi-contaminated soils.

  6. Capsaicin attenuates palmitate-induced expression of macrophage inflammatory protein 1 and interleukin 8 by increasing palmitate oxidation and reducing c-Jun activation in THP-1 (human acute monocytic leukemia cell) cells.

    PubMed

    Choi, Sung-E; Kim, Tae Ho; Yi, Sang-A; Hwang, Yun Cheong; Hwang, Won Sun; Choe, Sun Jung; Han, Seung Jin; Kim, Hae Jin; Kim, Dae Jung; Kang, Yup; Lee, Kwan-Woo

    2011-06-01

    Capsaicin, a spicy component of hot peppers, has been shown to improve inflammatory disease and obesity. In this study, we tested the hypothesis that the anti-inflammatory activity of capsaicin can be used to improve free fatty acid (FFA)-induced inflammation by reducing gene expression of macrophage inflammatory protein 1 (MIP-1) and interleukin 8 (IL-8) in THP-1 (human acute monocytic leukemia cell) macrophages. To investigate whether capsaicin ameliorates palmitate-induced MIP-1 and IL-8 gene expressions, we treated THP-1 cells with palmitate in the presence or absence of capsaicin and measured MIP-1 and IL-8 by real-time polymerase chain reaction. To elucidate the mechanism by which capsaicin effects on palmitate-induced MIP-1 and IL-8 gene expressions, we performed immunoblotting with stress kinase-related antibodies and measured palmitate oxidation and palmitate oxidation-related gene expression. Palmitate and stearate but not the unsaturated FFA oleate significantly increased MIP-1 and IL-8 expressions in THP-1 macrophages. Treatment with capsaicin or FFA oxidation stimulators inhibited palmitate-induced MIP-1 and IL-8 expressions in THP-1 macrophages. Capsaicin increased the gene expression of carnitine palmitoyltransferase 1 and the β-oxidation of palmitate. Furthermore, capsaicin significantly reduced palmitate-stimulated activation of c-Jun N-terminal kinase, c-Jun, and p38. Our data suggest that the attenuation of palmitate-induced MIP-1 and IL-8 gene expressions by capsaicin is associated with reduced activation of c-Jun N-terminal kinase, c-Jun, and p38 and preserved β-oxidation activity.

  7. Contracting C2C12 myotubes release CCL2 in an NF-κB-dependent manner to induce monocyte chemoattraction.

    PubMed

    Miyatake, Shouta; Bilan, Philip J; Pillon, Nicolas J; Klip, Amira

    2016-01-15

    Muscle inflammation following exercise is characterized by expression of inflammatory cytokines and chemokines. Exercise also increases muscle macrophages derived from circulating monocytes. However, it is unknown whether muscle cells themselves attract circulating monocytes, or what is the underlying mechanism. We used an in vitro system of electrical stimulation (ES) causing C2C12 myotube contraction to explore whether monocyte chemoattraction ensues and investigated the mediating chemoattractants. Conditioned medium from ES-contracted myotubes caused robust chemoattraction of THP-1 monocytes across Boyden chambers. Following ES, expression of several known monocyte chemokines [C-C motif ligand 2 (CCL2) and C-X-C motif ligand (CXCL)1, -2, and -5] was elevated, but of these, only recombinant CCL2 effectively reproduced monocyte migration. Electrically stimulated myotubes secreted CCL2, and neutralization of CCL2 in conditioned medium or antagonizing the CCL2 receptor (CCR2) in THP-1 monocytes inhibited ES-induced monocyte migration. N-benzyl-p-toluene sulfonamide (BTS), a myosin II-ATPase inhibitor, prevented ES-induced myotube contraction but not CCL2 gene expression and secretion. The membrane-permeant calcium chelator BAPTA-AM reduced ES-induced CCL2 secretion. Hence, electrical depolarization, rather than mechanical contraction, drives the rise in CCL2, with partial calcium input. ES activated the NF-κB pathway; NF-κB inhibitors reduced ES-induced CCL2 gene expression and secretion and repressed ES-induced THP-1 chemoattraction. Thus, electrically stimulated myotubes chemoattract monocytes through NF-κB-regulated CCL2 secretion. PMID:26554595

  8. Ketamine inhibits tumor necrosis factor-{alpha} and interleukin-6 gene expressions in lipopolysaccharide-stimulated macrophages through suppression of toll-like receptor 4-mediated c-Jun N-terminal kinase phosphorylation and activator protein-1 activation

    SciTech Connect

    Wu, G.-J.; Chen, T.-L.; Ueng, Y.-F.; Chen, R.-M.

    2008-04-01

    Our previous study showed that ketamine, an intravenous anesthetic agent, has anti-inflammatory effects. In this study, we further evaluated the effects of ketamine on the regulation of tumor necrosis factor-{alpha} (TNF-{alpha}) and interlukin-6 (IL-6) gene expressions and its possible signal-transducing mechanisms in lipopolysaccharide (LPS)-activated macrophages. Exposure of macrophages to 1, 10, and 100 {mu}M ketamine, 100 ng/ml LPS, or a combination of ketamine and LPS for 1, 6, and 24 h was not cytotoxic to macrophages. A concentration of 1000 {mu}M of ketamine alone or in combined treatment with LPS caused significant cell death. Administration of LPS increased cellular TNF-{alpha} and IL-6 protein levels in concentration- and time-dependent manners. Meanwhile, treatment with ketamine concentration- and time-dependently alleviated the enhanced effects. LPS induced TNF-{alpha} and IL-6 mRNA syntheses. Administration of ketamine at a therapeutic concentration (100 {mu}M) significantly inhibited LPS-induced TNF-{alpha} and IL-6 mRNA expressions. Application of toll-like receptor 4 (TLR4) small interfering (si)RNA into macrophages decreased cellular TLR4 levels. Co-treatment of macrophages with ketamine and TLR4 siRNA decreased the LPS-induced TNF-{alpha} and IL-6 productions more than alone administration of TLR4 siRNA. LPS stimulated phosphorylation of c-Jun N-terminal kinase and translocation of c-Jun and c-Fos from the cytoplasm to nuclei. However, administration of ketamine significantly decreased LPS-induced activation of c-Jun N-terminal kinase and translocation of c-Jun and c-Fos. LPS increased the binding of nuclear extracts to activator protein-1 consensus DNA oligonucleotides. Administration of ketamine significantly ameliorated LPS-induced DNA binding activity of activator protein-1. Therefore, a clinically relevant concentration of ketamine can inhibit TNF-{alpha} and IL-6 gene expressions in LPS-activated macrophages. The suppressive mechanisms

  9. Special AT-rich sequence-binding protein-1 participates in the maintenance of breast cancer stem cells through regulation of the Notch signaling pathway and expression of Snail1 and Twist1

    PubMed Central

    SUN, ZHENGKUI; ZHANG, CHAO; ZOU, XUESEN; JIANG, GUIXIANG; XU, ZONGQUAN; LI, WENTING; XIE, HUI

    2015-01-01

    The stem cell populations in cancerous tissues and cell lines vary widely and are often associated with aggressive cases of breast cancer. Despite research on the topic, the mechanism underlying the regulation of the breast cancer stem cell (BCSC) population within tumors remains to be fully elucidated. To investigate the function of special AT-rich sequence-binding protein-1 (SATB1) in the maintenance of the BCSC population, SATB1 was overexpressed with lentivirus in MCF-7 cells or knocked down with shRNA-lentivirus in BT-549 cells. The effects of SATB1 overexpression or knockdown on mammosphere formation, the size of the of BCSC population, cell invasion and tumorigenesis were investigated. Activation of the Notch signaling pathway and expression of Snail1 and Twist1 were also examined in the cells. Overexpression of SATB1 in MCF-7 cells was observed to increase mammosphere formation, the size of the BCSC population, cell invasion and tumorigenesis, accompanied by an increase in the activation of Notch signaling and expression levels of Snail1 and Twist1. Conversely, knockdown of SATB1 in BT-549 cells produced the opposite effects. The results indicated that expression of SATB1 may increase the size of the BCSC population via the activation of the Notch signaling pathway and by increasing expression levels of Snail1 and Twist1. PMID:25586771

  10. Bacterial chemoattraction towards jasmonate plays a role in the entry of Dickeya dadantii through wounded tissues.

    PubMed

    Antunez-Lamas, Maria; Cabrera, Ezequiel; Lopez-Solanilla, Emilia; Solano, Roberto; González-Melendi, Pablo; Chico, Jose Manuel; Toth, Ian; Birch, Paul; Pritchard, Leighton; Prichard, Leighton; Liu, Hui; Rodriguez-Palenzuela, Pablo

    2009-11-01

    Jasmonate is a key signalling compound in plant defence that is synthesized in wounded tissues. In this work, we have found that this molecule is also a strong chemoattractant for the phythopathogenic bacteria Dickeya dadantii (ex-Erwinia chysanthemi). Jasmonic acid induced the expression of a subset of bacterial genes possibly involved in virulence/survival in the plant apoplast and bacterial cells pre-treated with jasmonate showed increased virulence in chicory and Saintpaulia leaves. We also showed that tissue wounding induced bacterial spread through the leaf surface. Moreover, the jasmonate-deficient aos1 Arabidopsis thaliana mutant was more resistant to bacterial invasion by D. dadantii than wild-type plants. These results are consistent with the hypothesis that sensing jasmonic acid by this bacterium helps the pathogen to ingress inside plant tissues. PMID:19818025

  11. Mycoplasma pulmonis possesses a novel chemoattractant for B lymphocytes.

    PubMed Central

    Ross, S E; Simecka, J W; Gambill, G P; Davis, J K; Cassell, G H

    1992-01-01

    Mycoplasma pulmonis causes chronic murine respiratory mycoplasmosis, which is characterized by extensive peribronchial and perivascular infiltration of mononuclear cells, including B lymphocytes. B-lymphocyte recruitment into sites of inflammation is presently poorly understood but must involve directed chemotaxis of these cells in response to some external recruitment stimulus. In these studies, picogram amounts of M. pulmonis membrane protein were found to possess potent chemoattractant activity for resting rat B lymphocytes. This report is the first description of a bacterially derived chemoattractant for B lymphocytes and offers a unique opportunity to study regulation of B-lymphocyte recruitment to a site of chronic pulmonary inflammation. Furthermore, M. pulmonis membrane activation of fresh rat serum was found to produce a potent stimulus for recruitment of peritoneal and alveolar macrophages. M. pulmonis-mediated recruitment of lymphocytes and macrophages may play a significant role in the pathogenesis of murine respiratory mycoplasmosis, a role in which organisms on the bronchiolar epithelial surfaces may release proteins which can directly or indirectly promote chemotaxis of inflammatory cells from the circulation. PMID:1730502

  12. Over-expression of sterol-regulatory-element-binding protein-1c (SREBP1c) in rat pancreatic islets induces lipogenesis and decreases glucose-stimulated insulin release: modulation by 5-aminoimidazole-4-carboxamide ribonucleoside (AICAR).

    PubMed Central

    Diraison, Frédérique; Parton, Laura; Ferré, Pascal; Foufelle, Fabienne; Briscoe, Celia P; Leclerc, Isabelle; Rutter, Guy A

    2004-01-01

    Accumulation of intracellular lipid by pancreatic islet beta-cells has been proposed to inhibit normal glucose-regulated insulin secretion ('glucolipotoxicity'). In the present study, we determine whether over-expression in rat islets of the lipogenic transcription factor SREBP1c (sterol-regulatory-element-binding protein-1c) affects insulin release, and whether changes in islet lipid content may be reversed by activation of AMPK (AMP-activated protein kinase). Infection with an adenovirus encoding the constitutively active nuclear fragment of SREBP1c resulted in expression of the protein in approx. 20% of islet cell nuclei, with a preference for beta-cells at the islet periphery. Real-time PCR (TaqMan) analysis showed that SREBP1c up-regulated the expression of FAS (fatty acid synthase; 6-fold), acetyl-CoA carboxylase-1 (2-fold), as well as peroxisomal-proliferator-activated receptor-gamma (7-fold), uncoupling protein-2 (1.4-fold) and Bcl2 (B-cell lymphocytic-leukaemia proto-oncogene 2; 1.3-fold). By contrast, levels of pre-proinsulin, pancreatic duodenal homeobox-1, glucokinase and GLUT2 (glucose transporter isoform-2) mRNAs were unaltered. SREBP1c-transduced islets displayed a 3-fold increase in triacylglycerol content, decreased glucose oxidation and ATP levels, and a profound inhibition of glucose-, but not depolarisation-, induced insulin secretion. Culture of islets with the AMPK activator 5-amino-4-imidazolecarboxamide riboside decreased the expression of the endogenous SREBP1c and FAS genes, and reversed the effect of over-expressing active SREBP1c on FAS mRNA levels and cellular triacylglycerol content. We conclude that SREBP1c over-expression, even when confined to a subset of beta-cells, leads to defective insulin secretion from islets and may contribute to some forms of Type II diabetes. PMID:14690455

  13. MOLECULAR CLONING, EXPRESSION PATTERN OF MULTIDRUG RESISTANCE ASSOCIATED PROTEIN 1 (MRP1, ABCC1) GENE, AND THE SYNERGISTIC EFFECTS OF VERAPAMIL ON TOXICITY OF TWO INSECTICIDES IN THE BIRD CHERRY-OAT APHID.

    PubMed

    Kang, Xin-Le; Zhang, Meng; Wang, Kang; Qiao, Xian-Feng; Chen, Mao-Hua

    2016-05-01

    The ATP-binding cassette (ABC) transporters are important transmembrane proteins encoded by a supergene family. The majority of ABC proteins are primary active transporters that bind and hydrolyze ATP to mediate the efflux of a diverse range of substrates across lipid membranes. In this study, we cloned and characterized a putative multidrug resistance associated protein 1 (MRP1) from Rhopalosiphum padi encoded by ABCC1. Structural analysis showed that this protein has structural features typical of the ABC transporter family. Phylogenetic analysis indicated that the amino acid sequence was highly similar that of the corresponding protein from Acyrthosiphon pisum. Real-time quantitative polymerase chain reaction (PCR) analysis showed that ABCC1 was expressed throughout all R. padi developmental stages, with the highest level of expression in the fourth larval instar. We also examined ABCC1 expression in four different tissue types and found that it was most highly expressed in the midgut. Exposing R. padi to imidacloprid and chlorpyrifos increased ABCC1 expression. Furthermore, ABCC1 expression was higher in the imidacloprid-resistant (IR) and chlorpyrifos-resistant (CR) strains than in an insecticide-susceptible strain (SS) of R. padi. Exposing R. padi to verapamil in combination with insecticides significantly increased the toxicity of the insecticides. The respective synergy factor of CR and IR R. padi strain was 1.33 and 1.26, which was lower than that (2.72 and 1.64, respectively) of the SS. Our results clarify the biological function of ABCC1 in R. padi, particularly its role in insecticide resistance, and suggest novel strategies for pest management that use ABC transporter inhibitors to increase the effectiveness of insecticides.

  14. Dietary green tea extract lowers plasma and hepatic triglycerides and decreases the expression of sterol regulatory element-binding protein-1c mRNA and its responsive genes in fructose-fed, ovariectomized rats.

    PubMed

    Shrestha, Sudeep; Ehlers, Sarah J; Lee, Ji-Young; Fernandez, Maria-Luz; Koo, Sung I

    2009-04-01

    The objective of this study was to determine whether green tea (GT) inhibits the expression of genes regulating hepatic lipogenesis and intestinal lipid transport in fructose-fed ovariectomized (OX) rats. OX rats were assigned to: 1) a control group (S) fed the AIN-93G diet with corn starch as the major carbohydrate source; 2) another control group (F) fed the same diet but containing fructose at 60% as the major carbohydrate source; 3) a group fed the F diet but containing 0.5% GT; and 4) a group fed the F diet containing 1% GT. At 6 wk, plasma and liver triglyceride (TG) and cholesterol and expression of liver sterol regulatory element-binding protein-1c (SREBP-1c) and selected genes involved in lipogenesis and lipid transport were measured. Fructose elevated plasma TG and cholesterol compared with the S group. GT at 0.5 and 1.0% markedly lowered plasma and liver TG. Fructose increased the expression of SREBP-1c, fatty acid synthase, and stearoyl-CoA desaturase 1 mRNA in the liver, whereas GT decreased the expression of these lipogenic genes. Similarly, fructose increased the abundance of hepatic 3-hydroxy-3-methyl-glutaryl-CoA reductase mRNA, whereas GT significantly decreased its expression. GT did not alter the expression of scavenger receptor class B, type 1, microsomal TG transfer protein, and apobec 1 in the liver and intestine. The results suggest that the lipid-lowering effect of GT is mediated partly by its inhibition of hepatic lipogenesis involving SREBP-1c and its responsive genes without affecting lipoprotein assembly.

  15. Enhanced glutathione depletion, protein adduct formation, and cytotoxicity following exposure to 4-hydroxy-2-nonenal (HNE) in cells expressing human multidrug resistance protein-1 (MRP1) together with human glutathione S-transferase-M1 (GSTM1).

    PubMed

    Rudd, Lisa P; Kabler, Sandra L; Morrow, Charles S; Townsend, Alan J

    2011-11-15

    4-Hydroxy-2-nonenal (HNE) is one of the most reactive products of lipid peroxidation and has both cytotoxic and genotoxic effects in cells. Several enzymatic pathways have been reported to detoxify HNE, including conjugation by glutathione-S-transferases (GSTs). Removal of the resulting HNE-glutathione conjugate (HNE-SG) by an efflux transporter may be required for complete detoxification. We investigated the effect of expression of GSTM1 and/or the ABC efflux transporter protein, multidrug-resistance protein-1 (MRP1), on HNE-induced cellular toxicity. Stably transfected MCF7 cell lines were used to examine the effect of GSTM1 and/or MRP1 expression on HNE-induced cytotoxicity, GSH depletion, and HNE-protein adduct formation. Co-expression in the MCF7 cell line of GSTM1 with MRP1 resulted in a 2.3-fold sensitization to HNE cytotoxicity (0.44-fold IC(50) value relative to control) rather than the expected protection. Expression of either GSTM1 or MRP1 alone also resulted in slight sensitization to HNE cytotoxicity (0.79-fold and 0.71-fold decreases in IC(50) values, respectively). Co-expression of GSTM1 and MRP1 strongly enhanced the formation of HNE-protein adducts relative to the non-expressing control cell line, whereas expression of either MRP1 alone or GSTM1 alone yielded similarly low levels of HNE-protein adducts to that of the control cell line. Glutathione (GSH) levels were reduced by 10-20% in either the control cell line or the MCF7/GSTM1 cell line with the same HNE exposure for 60min. However, HNE induced >80% depletion of GSH in cells expressing MRP1 alone. Co-expression of both MRP1 and GSTM1 caused slightly greater GSH depletion, consistent with the greater protein adduct formation and cytotoxicity in this cell line. Since expression of GSTM1 or MRP1 alone did not strongly sensitize cells to HNE, or result in greater HNE-protein adducts than in the control cell line, these results indicate that MRP1 and GSTM1 collaborate to enhance HNE-protein adduct

  16. The viral non-structural protein 1 alpha (Nsp1α) inhibits p53 apoptosis activity by increasing murine double minute 2 (mdm2) expression in porcine reproductive and respiratory syndrome virus (PRRSV) early-infected cells.

    PubMed

    Wang, Xiaodu; Shao, Chunyan; Wang, Luyan; Li, Qunjing; Song, Houhui; Fang, Weihuan

    2016-02-29

    Apoptosis is one of the most important mechanisms of pathogenesis in porcine reproductive and respiratory syndrome virus (PRRSV)-infected cells. The tumor suppressor p53 plays a critical role in apoptotic induction in viral infections. In the present study, we found that p53 activity was inhibited at the early stage of PRRSV infection in both the highly pathogenic (HP) and lowly pathogenic (LP) PRRSV isolates. Bax expression showed a similar change pattern to that of p53. Murine double minute 2 (mdm2) expressed higher in PRRSV-infected cells than in uninfected cells at the early stage of infection and promoted p53 degradation. We show for the first time that the non-structural protein 1 alpha (Nsp1α) of PRRSV is a negative regulator of p53 activity through increasing mdm2 expression and p53 ubiquitination, while p53 is inhibitory to PRRSV replication at the early stage of infection. We conclude that PRRSV manipulates the host factors mdm2 and p53 via its Nsp1α for increased replication at the early stage of infection. These provide a novel perspective to understand the interaction between apoptosis and replication of PRRSV.

  17. Confined housing system increased abdominal and subcutaneous fat deposition and gene expressions of carbohydrate response element-binding protein and sterol regulatory element-binding protein 1 in chicken.

    PubMed

    Li, Q; Zhao, X L; Gilbert, E R; Liu, Y P; Wang, Y; Qiu, M H; Zhu, Q

    2015-01-01

    Free-range production system is increasingly being used in poultry breeding and feed production in many countries. The objective of the current experiment was to evaluate the effects of different raising systems on fat-related traits and mRNA levels of liver lipogenesis genes in Erlang Mountainous chicken. Each of 10 birds (91 day old) from caged, indoor-floor housed, and free-range housing systems was slaughtered, and fat-related traits, live body weight (BW), subcutaneous fat thickness (SFT), abdominal fat weight (AFW), abdominal fat percentage (AFP), and intramuscular fat content were determined. The mRNA levels of liver X receptor α, carbohydrate response element-binding protein (ChREBP), sterol regulatory element-binding protein-1 (SREBP1), and fatty acid synthase were detected. The caged chicken exhibited significantly higher BW, SFT, and AFW than those of free-ranged chicken (P < 0.05). All the 4 genes had a similar expression pattern, and they showed the highest level in caged chicken, while the lowest level was found in free-ranged chicken. Association analysis indicated that there were significant (P < 0.05) or highly significant (P < 0.01) positive correlations between the mRNA levels of ChREBP, SREBP1, and fat traits of SFT, AFW, and AFP. Thus, we deduced that increased fat deposition in caged chicken was probably induced by increased gene expression of ChREBP and SREBP1 in the liver. PMID:25730060

  18. Confined housing system increased abdominal and subcutaneous fat deposition and gene expressions of carbohydrate response element-binding protein and sterol regulatory element-binding protein 1 in chicken.

    PubMed

    Li, Q; Zhao, X L; Gilbert, E R; Liu, Y P; Wang, Y; Qiu, M H; Zhu, Q

    2015-02-06

    Free-range production system is increasingly being used in poultry breeding and feed production in many countries. The objective of the current experiment was to evaluate the effects of different raising systems on fat-related traits and mRNA levels of liver lipogenesis genes in Erlang Mountainous chicken. Each of 10 birds (91 day old) from caged, indoor-floor housed, and free-range housing systems was slaughtered, and fat-related traits, live body weight (BW), subcutaneous fat thickness (SFT), abdominal fat weight (AFW), abdominal fat percentage (AFP), and intramuscular fat content were determined. The mRNA levels of liver X receptor α, carbohydrate response element-binding protein (ChREBP), sterol regulatory element-binding protein-1 (SREBP1), and fatty acid synthase were detected. The caged chicken exhibited significantly higher BW, SFT, and AFW than those of free-ranged chicken (P < 0.05). All the 4 genes had a similar expression pattern, and they showed the highest level in caged chicken, while the lowest level was found in free-ranged chicken. Association analysis indicated that there were significant (P < 0.05) or highly significant (P < 0.01) positive correlations between the mRNA levels of ChREBP, SREBP1, and fat traits of SFT, AFW, and AFP. Thus, we deduced that increased fat deposition in caged chicken was probably induced by increased gene expression of ChREBP and SREBP1 in the liver.

  19. The group A streptococcal collagen-like protein-1, Scl1, mediates biofilm formation by targeting the extra domain A-containing variant of cellular fibronectin expressed in wounded tissue.

    PubMed

    Oliver-Kozup, Heaven; Martin, Karen H; Schwegler-Berry, Diane; Green, Brett J; Betts, Courtney; Shinde, Arti V; Van De Water, Livingston; Lukomski, Slawomir

    2013-02-01

    Wounds are known to serve as portals of entry for group A Streptococcus (GAS). Subsequent tissue colonization is mediated by interactions between GAS surface proteins and host extracellular matrix components. We recently reported that the streptococcal collagen-like protein-1, Scl1, selectively binds the cellular form of fibronectin (cFn) and also contributes to GAS biofilm formation on abiotic surfaces. One structural feature of cFn, which is predominantly expressed in response to tissue injury, is the presence of a spliced variant containing extra domain A (EDA/EIIIA). We now report that GAS biofilm formation is mediated by the Scl1 interaction with EDA-containing cFn. Recombinant Scl1 proteins that bound cFn also bound recombinant EDA within the C-C' loop region recognized by the α(9)β(1) integrin. The extracellular 2-D matrix derived from human dermal fibroblasts supports GAS adherence and biofilm formation. Altogether, this work identifies and characterizes a novel molecular mechanism by which GAS utilizes Scl1 to specifically target an extracellular matrix component that is predominantly expressed at the site of injury in order to secure host tissue colonization.

  20. Bone marrow-derived mesenchymal stem cells attenuate acute liver injury and regulate the expression of fibrinogen-like-protein 1 and signal transducer and activator of transcription 3.

    PubMed

    Zou, Zhuolin; Cai, Yijing; Chen, Yi; Chen, Si; Liu, Liyuan; Shen, Zhonghai; Zhang, Sainan; Xu, Lanman; Chen, Yongping

    2015-08-01

    In recent years, bone marrow-derived mesenchymal stem cells (BMSCs) have been demonstrated to exert extensive therapeutic effects on acute liver injury; however, the underlying mechanisms of these effects have remained to be elucidated. The present study focused on the potential anti-apoptotic and pro-regenerative effects of BMSCs in D-galactosamine (D-Gal) and lipopolysaccharide (LPS)-induced acute liver injury in rats. An experimental rat acute liver injury model was established by intraperitoneal injection of D-Gal (400 mg/kg) and LPS (80 μg/kg). BMSCs and an identical volume of saline were administered via the caudal vein 2 h after the D-Gal and LPS challenge. Subsequently, the serum samples were collected to detect the levels of alanine aminotransferase and aspartate aminotransferase. Hematoxylin and eosin staining, terminal deoxynucleotidyl transferase-mediated nick-end labeling assay and immunohistochemical staining were performed to determine apoptosis, regeneration and histological changes of liver sections. Western blotting and reverse transcription-quantitative polymerase chain reaction were performed to detect the protein and mRNA expression levels of fibrinogen-like-protein 1 (FGL1), phosphorylated signal transducer and activator of transcription 3 (p-STAT3), STAT3 and B-cell lymphoma 2 (Bcl-2) and Bcl-2 associated X protein (Bax) in liver tissue samples. The results indicated that intravenous transplantation of BMSCs significantly decreased the levels of alanine aminotransferase and aspartate aminotransferase, and reduced hepatocellular necrosis and inflammatory cell infiltration. Additionally, a terminal deoxynucleotidyl transferase-mediated nick-end labeling assay and immunohistochemical staining revealed that BMSC treatment reduced hepatocyte apoptosis and enhanced liver regeneration. Furthermore, Bcl-2 expression was increased, whilst the protein expression of Bax was reduced. The expression of FGL1 and p-STAT3 were elevated concurrently with the

  1. Switching of chemoattractant receptors programs development and morphogenesis in Dictyostelium: receptor subtypes activate common responses at different agonist concentrations.

    PubMed

    Kim, J Y; Borleis, J A; Devreotes, P N

    1998-05-01

    One of the common functional features among G-protein coupled receptors is the occurrence of multiple subtypes involved in similar signal transduction events. The cAMP chemoattractant receptor family of Dictyostelium discoideum is composed of four receptors (cAR1-cAR4), which are expressed sequentially throughout the developmental transition from a unicellular to a multicellular organism. The receptors differ in affinity for cAMP and in the sequences of their C-terminal domains. In this study, we constitutively expressed cAR1, cAR2, and cAR3 as well as a series of chimeric and mutant receptors and assessed the capacity of each to mediate chemotaxis, activation of adenylyl cyclase and actin polymerization, and rescue the developmental defect of car1-/car3- cells. We found that various receptors and mutants sense different concentration ranges of cAMP but all can mediate identical responses during the aggregation stage of development. The responses displayed very similar kinetics, suggesting no major differences in regulatory properties attributable to the C-terminal domains. We speculate that switching of receptor subtypes during development enables the organism to respond to the changing concentrations of the chemoattractant and thereby program morphogenesis appropriately. PMID:9578623

  2. Disorganized muscle protein-1 (DIM-1) of filarial parasite Brugia malayi: cDNA cloning, expression, purification, structural modeling and its potential as vaccine candidate for human filarial infection.

    PubMed

    Kushwaha, Vikas; Kumar, Vikash; Verma, Shiv K; Sharma, Rolee; Siddiqi, M I; Murthy, P K

    2014-03-26

    We have recently identified disorganized muscle protein-1 (DIM-1) in one of the proinflammatory fractions of the human filaria Brugia malayi adult worm. The present study was undertaken to characterize B. malayi DIM-1 (DIM-1bm) and explore its vaccine potential. In this study we cloned and expressed the DIM-1bm gene, investigated its sequence homology with other nematodes, constructed in silico structural model, purified the recombinant DIM-1bm (rDIM-1bm) protein, and studied the effect of immunization with rDIM-1bm on the establishment of B. malayi infection in Mastomys coucha. DIM-1bm showed similarity with DIM-1 of Caenorhabditis elegans, Ascaris suum and Loa loa. Structural modeling revealed three immunoglobulin domains in DIM-1bm indicating that it is a member of immunoglobulin superfamily (IgSF) and 'blastn' results showed that DIM-1bm coding sequence (CDS) have almost no homology with human and mouse nucleotide sequences. Immunization with rDIM-1bm partially protected M. coucha against establishment of infection as inferred by a low recovery of microfilariae (37-64%) and parasite burden (∼50%). The enhanced activity of macrophages, and IFN-γ and NO responses, and elevated levels of specific IgG, IgG1, IgG2a and IgG2b correlated with parasitological findings. This is the first report on cloning, expression, structural modeling and purification of rDIM-1bm and its ability to partially prevent establishment of B. malayi infection. DIM-1bm's almost complete lack of homology with the human counterpart makes it an attractive protein for exploring its vaccine potential. PMID:24513011

  3. Restoring host-microbe homeostasis via selective chemoattraction of Tregs.

    PubMed

    Garlet, G P; Sfeir, C S; Little, S R

    2014-09-01

    The disruption of host-microbe homeostasis at the site of periodontal disease is considered a key factor for disease initiation and progress. While the downstream mechanisms responsible for the tissue damage per se are relatively well-known (involving various patterns of immune response operating toward periodontal tissue destruction), we are only beginning to understand the complexity of host-microbe interactions in the periodontal environment. Unfortunately, most of the research has been focused on the disruption of host-microbe homeostasis instead of focusing on the factors responsible for maintaining homeostasis. In this context, regulatory T-cells (Tregs) comprise a CD4+FOXp3 +T-cell subset with a unique ability to regulate other leukocyte functions to avoid excessive immune activation and its pathological consequences. Tregs act as critical determinants of host-microbe homeostasis, as well as determinants of a balanced host response after the disruption of host-microbe homeostasis by pathogens. In periodontitis, Tregs play a protective role, with their natural recruitment being responsible for conversion of active into inactive lesions. With controlled-release technology, it is now possible to achieve a selective chemoattraction of Tregs to periodontal tissues, attenuating experimental periodontitis evolution due to the local control of inflammatory immune response and the generation of a pro-reparative environment.

  4. Dynamic Distribution of Chemoattractant Receptors in Living Cells During Chemotaxis and Persistent Stimulation

    PubMed Central

    Xiao, Zhan; Zhang, Ning; Murphy, Douglas B.; Devreotes, Peter N.

    1997-01-01

    While the localization of chemoattractant receptors on randomly oriented cells has been previously studied by immunohistochemistry, the instantaneous distribution of receptors on living cells undergoing directed migration has not been determined. To do this, we replaced cAR1, the primary cAMP receptor of Dictyostelium, with a cAR1-green fluorescence protein fusion construct. We found that this chimeric protein is functionally indistinguishable from wild-type cAR1. By time-lapse imaging of single cells, we observed that the receptors remained evenly distributed on the cell surface and all of its projections during chemotaxis involving turns and reversals of polarity directed by repositioning of a chemoattractant-filled micropipet. Thus, cell polarization cannot result from a gradient-induced asymmetric distribution of chemoattractant receptors. Some newly extended pseudopods at migration fronts showed a transient drop in fluorescence signals, suggesting that the flow of receptors into these zones may slightly lag behind the protrusion process. Challenge with a uniform increase in chemoattractant, sufficient to cause a dramatic decrease in the affinity of surface binding sites and cell desensitization, also did not significantly alter the distribution profile. Hence, the induced reduction in binding activity and cellular sensitivity cannot be due to receptor relocalization. The chimeric receptors were able to “cap” rapidly during treatment with Con A, suggesting that they are mobile in the plane of the cell membrane. This capping was not influenced by pretreatment with chemoattractant. PMID:9334341

  5. Merozoite surface protein-1 of Plasmodium yoelii fused via an oligosaccharide moiety of cholera toxin B subunit glycoprotein expressed in yeast induced protective immunity against lethal malaria infection in mice.

    PubMed

    Miyata, Takeshi; Harakuni, Tetsuya; Taira, Toki; Matsuzaki, Goro; Arakawa, Takeshi

    2012-01-20

    Methylotrophic yeast (Pichia pastoris) secreted cholera toxin B subunit (CTB) predominantly as a biologically active pentamer (PpCTB) with identical ganglioside binding affinity profiles to that of choleragenoid. Unlike choleragenoid, however, the PpCTB did not induce a footpad edema response in mice. Of the two potential glycosylation sites (NIT(4-6) and NKT(90-92)) for this protein, a N-linked oligosaccharide was identified at Asn4. The oligosaccharide, presumed to extend from the lateral circumference of the CTB pentamer ring structure, was exploited as a site-specific anchoring scaffold for the C-terminal 19-kDa merozoite surface protein-1 (MSP1-19) of the rodent malaria parasite, Plasmodium yoelii. Conjugation of MSP1-19 to PpCTB via its oligosaccharide moiety induced higher protective efficacy against lethal parasite infection than conjugation directly to the PpCTB protein body in both intranasal and subcutaneous immunization regimes. Such increased protection was potentially due to the higher antigen loading capacity of CTB achieved when the antigen was linked to the extended branches of the oligosaccharide. This might have allowed the antigen to reside in more spacious molecular environment with less steric hindrance between the constituent molecules of the fusion complex. PMID:22119928

  6. Local modulation of chemoattractant concentrations by single cells: dissection using a bulk-surface computational model

    PubMed Central

    Nolan, M.

    2016-01-01

    Chemoattractant gradients are usually considered in terms of sources and sinks that are independent of the chemotactic cell. However, recent interest has focused on ‘self-generated’ gradients, in which cell populations create their own local gradients as they move. Here, we consider the interplay between chemoattractants and single cells. To achieve this, we extend a recently developed computational model to incorporate breakdown of extracellular attractants by membrane-bound enzymes. Model equations are parametrized, using the published estimates from Dictyostelium cells chemotaxing towards cyclic AMP. We find that individual cells can substantially modulate their local attractant field under physiologically appropriate conditions of attractant and enzymes. This means the attractant concentration perceived by receptors can be a small fraction of the ambient concentration. This allows efficient chemotaxis in chemoattractant concentrations that would be saturating without local breakdown. Similar interactions in which cells locally mould a stimulus could function in many types of directed cell motility, including haptotaxis, durotaxis and even electrotaxis. PMID:27708760

  7. Effect of steady and unsteady flow on chemoattractant plume formation and sperm taxis

    NASA Astrophysics Data System (ADS)

    Bell, Allison F.; Crimaldi, John P.

    2015-08-01

    The formation of chemoattractant plumes around benthic invertebrate eggs in steady and unsteady shear flows is investigated for a range of shear rates, and the ability of sperm to navigate within these plumes is assessed using several chemotactic strategies. Although many of the details of sperm taxis remain uncertain, we investigate the role of basic processes using a toy model in two dimensions. Search strategies in 2D are intrinsically less complex than 3D, but many of the basic components are similar, and the simplified geometry permits an understanding and identification of the key factors of navigation tactics. Numerical simulations are used to model the advection and diffusion of the chemoattractant within the different flows, using three different sperm swimming behaviors. A Monte-Carlo approach is then used to determine the probability of a sperm reaching an egg for a range of flow conditions, initial conditions, and swimming behaviors. The spatial structure of chemoattractant plumes at the scale of the gametes is also investigated. Success rates for locating an egg decrease monotonically with increasing shear rates, and a definitive hierarchical ordering of the tested swimming strategies is identified. A conceptual framework to study and identify important aspects of this fundamental process to support further studies is provided.

  8. Motility of Pseudomonas aeruginosa in saturated granular media as affected by chemoattractant.

    PubMed

    Chen, Jiazhou; Jin, Yan

    2011-09-25

    To examine and quantify the effects of glass beads and chemoattractant on bacterial motility in granular media, we examined the motile behavior of P. aeruginosa in a saturated granular medium and quantified the effects of glass beads and the presence of a chemoattractant. By recording individual cell trajectories in microfluidic channels under a high-speed confocal microscope, we directly measured the cell's run direction and corresponding run-length, speed and turn angle. Bacterial run speed increased in the presence of chemoattractant in both aqueous and granular media. But it decreased in glass-beads compared to in aqueous media due to the restricted pore geometry and interactions between bacteria and grain surfaces. Notably, the relatively higher frequency distribution at turn angles of 170° decreased dramatically, while the smaller peak at 70° increased and became dominant on a bimodal distribution, showing more bacteria changed directions at smaller turn angles rather than reverse their swimming directions. Additionally, the presence of glass beads also decreased the chemotactic velocity and random motility by similar proportions due to the restrictive geometry and the interactions between bacteria and glass beads surface. Our study indicates that the swimming parameters measured from aqueous media cannot be directly adopted in models for predicting bacteria travel in granular media.

  9. Lipopolysaccharide Decreases Single Immunoglobulin Interleukin-1 Receptor-related Molecule (SIGIRR) Expression by Suppressing Specificity Protein 1 (Sp1) via the Toll-like Receptor 4 (TLR4)-p38 Pathway in Monocytes and Neutrophils*

    PubMed Central

    Ueno-Shuto, Keiko; Kato, Kosuke; Tasaki, Yukihiro; Sato, Miki; Sato, Keizo; Uchida, Yuji; Sakai, Hiromichi; Ono, Tomomi; Suico, Mary Ann; Mitsutake, Kazunori; Tokutomi, Naofumi; Kai, Hirofumi; Shuto, Tsuyoshi

    2014-01-01

    Single immunoglobulin interleukin-1 receptor-related molecule (SIGIRR) is one of the immunoglobulin-like membrane proteins that is crucial for negative regulation of toll-like receptor 4 (TLR4) and interleukin-1 receptor. Despite the importance of understanding its expression and function, knowledge is limited on the regulatory mechanism in the epithelial tissues, such as the liver, lung, and gut, where its predominant expression is originally described. Here, we found expression of SIGIRR in non-epithelial innate immune cells, including primary peripheral blood monocytes, polymorphonuclear neutrophils, monocytic RAW264 cells, and neutrophilic-differentiated HL-60 cells. Consistent with previous findings in epithelial tissues, SIGIRR gene and protein expression were also down-regulated by LPS treatment in a time-dependent manner in primary blood monocytes and polymorphonuclear neutrophils. A reduction was also observed in RAW264 and differentiated HL-60 cells. Notably, exogenous introduction of the dominant negative form of TLR4 and siRNA of p38 resulted in inhibition of LPS-induced SIGIRR down-regulation, whereas treatment with p38 activator anisomycin showed a dose-dependent decrease in SIGIRR expression, suggesting TLR4-p38 signal as a critical pathway for LPS-induced SIGIRR down-regulation. Finally, reporter gene and chromatin immunoprecipitation assays demonstrated that Sp1 is a key factor that directly binds to the proximal promoter of SIGIRR gene and consequently regulates basal SIGIRR expression, which is negatively regulated by the LPS-dependent TLR4-p38 pathway. In summary, the data precisely demonstrate how LPS down-regulates SIGIRR expression and provide a role of LPS signal that counteracts Sp1-dependent basal promoter activation of SIGIRR gene via TLR4-p38 pathway in non-epithelial innate immune cells. PMID:24821721

  10. Mutation in and lack of expression of tyrosinase-related protein-1 (TRP-1) in melanocytes from an individual with brown oculocutaneous albinism: a new subtype of albinism classified as "OCA3".

    PubMed Central

    Boissy, R. E.; Zhao, H.; Oetting, W. S.; Austin, L. M.; Wildenberg, S. C.; Boissy, Y. L.; Zhao, Y.; Sturm, R. A.; Hearing, V. J.; King, R. A.; Nordlund, J. J.

    1996-01-01

    Most types of human oculocutaneous albinism (OCA) result from mutations in the gene for tyrosinase (OCA1) or the P protein (OCA2), although other types of OCA have been described but have not been mapped to specific loci. Melanocytes were cultured from an African-American with OCA, who exhibited the phenotype of Brown OCA, and his normal fraternal twin. Melanocytes cultured from the patient with OCA and the normal twin appeared brown versus black, respectively. Melanocytes from both the patient with OCA and the normal twin demonstrated equal amounts of NP-40-soluble melanin; however, melanocytes from the patient with OCA contained only 7% of the amount of insoluble melanin found from the normal twin. Tyrosinase- related protein-1 (TRP-1) was not detected in the OCA melanocytes by use of various anti-TRP-1 probes. Furthermore, transcripts for TRP-1 were absent in cultured OCA melanocytes. The affected twin was homozygous for a single-bp deletion in exon 6, removing an A in codon 368 and leading to a premature stop at codon 384. Tyrosine hydroxylase activity of the OCA melanocytes was comparable to controls when assayed in cell lysates but was only 30% of controls when assayed in intact cells. We conclude that this mutation of the human TRP-1 gene affects its interaction with tyrosinase, resulting in dysregulation of tyrosinase activity, promotes the synthesis of brown versus black melanin, and is responsible for a third genetic type of OCA in humans, which we classify as "OCA3." Images Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 Figure 6 PMID:8651291

  11. Mutation in and lack of expression of tyrosinase-related protein-1 (TRP-1) in melanocytes from an individual with brown oculocutaneous albinism: A new subtype of albinism classified as {open_quotes}OCA3{close_quotes}

    SciTech Connect

    Boissy, R.E.; Zhao, H.; Austin, L.M.; Boissy, Y.L.; Zhao, Y.

    1996-06-01

    Most types of human oculocutaneous albinism (OCA) result from mutations in the gene for tyrosinase (OCA1) or the P protein (OCA2), although other types of OCA have been described but have not been mapped to specific loci. Melanocytes were cultured from an African-American with OCA, who exhibited the phenotype of Brown OCA, and his normal fraternal twin. Melanocytes cultured from the patient with OCA and the normal twin appeared brown versus black, respectively. Melanocytes from both the patient with OCA and the normal twin demonstrated equal amounts of NP-40-soluble melanin; however, melanocytes from the patient with OCA contained only 7% of the amount of insoluble melanin found from the normal twin. Tyrosinase-related protein-1 (TRP-1) was not detected in the OCA melanocytes by use of various anti-TRP-1 probes. Furthermore, transcripts for TRP-1 were absent in cultured OCA melanocytes. The affected twin was homozygous for a single-bp deletion in exon 6, removing an A in codon 368 and leading to a premature stop at codon 384. Tyrosine hydroxylase activity of the OCA melanocytes was comparable to controls when assayed in cell lysates but was only 30% of controls when assayed in intact cells. We conclude that this mutation of the human TRP-1 gene affects its interaction with tyrosinase, resulting in dysregulation of tyrosinase activity, promotes the synthesis of brown versus black melanin, and is responsible for a third genetic type of OCA in humans, which we classify as {open_quotes}OCA3.{close_quotes} 69 refs., 7 figs., 3 tabs.

  12. L-Arogenate is a chemoattractant which can be utilized as the sole source of carbon and nitrogen by Pseudomonas aeruginosa

    SciTech Connect

    Fischer, R.S.; Song, Jian; Gu, Wei; Jensen, R.A.

    1997-02-01

    L-Arogenate is a commonplace amino acid in nature in consideration of its role as a ubiquitous precursor of L-phenylalanine and/or L-tyrosine. However, the questions of whether it serves as a chemoattractant molecule and whether it can serve as a substrate for catabolism have never been studied. We found that Pseudomonas aeruginosa recognizes L-arogenate as a chemoattractant molecule which can be utilized as a source of both carbon and nitrogen. Mutants lacking expression of either cyclohexadienyl dehydratase or phenylalanine hydroxylase exhibited highly reduced growth rates when utilizing L-arogenate as a nitrogen source. Utilization of L-arogenate as a source of either carbon or nitrogen was dependent upon {sub S}{sup 54}, as revealed by the use of an rpoN null mutant. The evidence suggests that catabolism of L-arogenate proceeds via alternative pathways which converge at 4-hydroxyphenylpyruvate. In one pathway, prephenate formed in the periplasm by deamination of L-arogenate is converted to 4-hydroxyphenylpyruvate by cyclohexadienyl dehydrogenase. The second route depends upon the sequential action of periplasmic cyclohexadienyl dehydratase, phenylalanine hydroxylase, and aromatic aminotransferase. 32 refs., 5 figs., 4 tabs.

  13. The uncoupling protein 1 gene, UCP1, is expressed in mammalian islet cells and associated with acute insulin response to glucose in African American families from the IRAS Family Study

    PubMed Central

    Sale, Michèle M; Hsu, Fang-Chi; Palmer, Nicholette D; Gordon, Candace J; Keene, Keith L; Borgerink, Hermina M; Sharma, Arun J; Bergman, Richard N; Taylor, Kent D; Saad, Mohammed F; Norris, Jill M

    2007-01-01

    Background Variants of uncoupling protein genes UCP1 and UCP2 have been associated with a range of traits. We wished to evaluate contributions of known UCP1 and UCP2 variants to metabolic traits in the Insulin Resistance and Atherosclerosis (IRAS) Family Study. Methods We genotyped five promoter or coding single nucleotide polymorphisms (SNPs) in 239 African American (AA) participants and 583 Hispanic participants from San Antonio (SA) and San Luis Valley. Generalized estimating equations using a sandwich estimator of the variance and exchangeable correlation to account for familial correlation were computed for the test of genotypic association, and dominant, additive and recessive models. Tests were adjusted for age, gender and BMI (glucose homeostasis and lipid traits), or age and gender (obesity traits), and empirical P-values estimated using a gene dropping approach. Results UCP1 A-3826G was associated with AIRg in AA (P = 0.006) and approached significance in Hispanic families (P = 0.054); and with HDL-C levels in SA families (P = 0.0004). Although UCP1 expression is reported to be restricted to adipose tissue, RT-PCR indicated that UCP1 is expressed in human pancreas and MIN-6 cells, and immunohistochemistry demonstrated co-localization of UCP1 protein with insulin in human islets. UCP2 A55V was associated with waist circumference (P = 0.045) in AA, and BMI in SA (P = 0.018); and UCP2 G-866A with waist-to-hip ratio in AA (P = 0.016). Conclusion This study suggests a functional variant of UCP1 contributes to the variance of AIRg in an AA population; the plausibility of this unexpected association is supported by the novel finding that UCP1 is expressed in islets. PMID:17397545

  14. Cut-like Homeobox 1 (CUX1) Regulates Expression of the Fat Mass and Obesity-associated and Retinitis Pigmentosa GTPase Regulator-interacting Protein-1-like (RPGRIP1L) Genes and Coordinates Leptin Receptor Signaling*

    PubMed Central

    Stratigopoulos, George; LeDuc, Charles A.; Cremona, Maria L.; Chung, Wendy K.; Leibel, Rudolph L.

    2011-01-01

    The first intron of FTO contains common single nucleotide polymorphisms associated with body weight and adiposity in humans. In an effort to identify the molecular basis for this association, we discovered that FTO and RPGRIP1L (a ciliary gene located in close proximity to the transcriptional start site of FTO) are regulated by isoforms P200 and P110 of the transcription factor, CUX1. This regulation occurs via a single AATAAATA regulatory site (conserved in the mouse) within the FTO intronic region associated with adiposity in humans. Single nucleotide polymorphism rs8050136 (located in this regulatory site) affects binding affinities of P200 and P110. Promoter-probe analysis revealed that binding of P200 to this site represses FTO, whereas binding of P110 increases transcriptional activity from the FTO as well as RPGRIP1L minimal promoters. Reduced expression of Fto or Rpgrip1l affects leptin receptor isoform b trafficking and leptin signaling in N41 mouse hypothalamic or N2a neuroblastoma cells in vitro. Leptin receptor clusters in the vicinity of the cilium of arcuate hypothalamic neurons in C57BL/6J mice treated with leptin, but not in fasted mice, suggesting a potentially important role of the cilium in leptin signaling that is, in part, regulated by FTO and RPGRIP1L. Decreased Fto/Rpgrip1l expression in the arcuate hypothalamus coincides with decreased nuclear enzymatic activity of a protease (cathepsin L) that has been shown to cleave full-length CUX1 (P200) to P110. P200 disrupts (whereas P110 promotes) leptin receptor isoform b clustering in the vicinity of the cilium in vitro. Clustering of the receptor coincides with increased leptin signaling as reflected in protein levels of phosphorylated Stat3 (p-Stat3). Association of the FTO locus with adiposity in humans may reflect functional consequences of A/C alleles at rs8050136. The obesity-risk (A) allele shows reduced affinity for the FTO and RPGRIP1L transcriptional activator P110, leading to the

  15. Structure of human monocyte chemoattractant protein 4 (MCP-4/CCL13)

    SciTech Connect

    Barinka, Cyril; Prahl, Adam; Lubkowski, Jacek

    2008-04-02

    Monocyte chemoattractant proteins (MCPs) belong to the CC chemokine family and are involved in many (patho)physiological processes characterized by mononuclear cell infiltration, including tissue remodeling, atherosclerosis and cancer metastasis. Here, the crystal structure of human monocyte chemoattractant protein 4 (MCP-4) refined at 1.70 {angstrom} resolution is reported with crystallographic values R = 0.180 and R{sub free} = 0.212. The overall MCP-4 fold reveals the typical tertiary features of the CC chemokine family. A central three-stranded antiparallel {beta}-sheet is C-terminally flanked by an overlaying {alpha}-helix, while the N-terminal part of the molecule forms an extended loop that is anchored to the rest of the molecule via two disulfide bridges, Cys11-Cys35 and Cys12-Cys51. The crystal packing suggests the existence of MCP-4 dimers with a dimerization interface similar to those previously reported for the X-ray structures of MCP-1 and MCP-2.

  16. The chemoattractant potential of ginsenosides in the ginseng - Pythium irregulare pathosystem.

    PubMed

    Ivanov, Dimitre A; Georgakopoulos, Jorge R C; Bernards, Mark A

    2016-02-01

    Ginsenosides produced by ginseng (Panax quinquefolius L.) are mildly fungitoxic saponins; however, exposure of the ginseng root pathogen Pythium irregulare Buisman to ginsenosides enhances its growth in a dose dependent manner, leading to speculation that ginsenosides may function as chemoattractants and/or growth regulators in the context of the ginseng - P. irregulare pathosystem. In the present work, it was demonstrated that the treatment of ginseng plants with a relatively high dose of ginsenosides by dipping their roots into a solution of ginsenosides prior to planting results in delayed infection by P. irregulare in pot experiments, as monitored by non-invasive chlorophyll fluorescence imaging. In an attempt to determine whether this observation results from a protective effect of the ginsenosides, or from a modification of P. irregulare growth habit in response to ginsenosides present in the soil, standard in vitro disk diffusion assays were conducted. Here, exposure of P. irregulare to crude ginsenosides or pure ginsenoside Rb1, resulted in delayed hyphal progression, while enhancing aerial hyphae build-up around ginsenoside-treated disks. By contrast, assays with pure ginsenoside F2 resulted in clear zones of inhibition around treated disks. While it remains unclear whether ginsenosides act as chemoattractants for P. irregulare in vivo, the results here suggest that these saponins serve to alter the growth habit of this organism, both in vivo and in vitro.

  17. Positive feedback may cause the biphasic response observed in the chemoattractant-induced response of Dictyostelium cells*

    PubMed Central

    Yang, Liu; Iglesias, Pablo A.

    2006-01-01

    After stimulation by chemoattractant, Dictyostelium cells exhibit a rapid response. The concentrations of several intracellular proteins rise rapidly reaching their maximum levels approximately 5–10 seconds, after which they return to prestimulus levels. This response, which is found in many other chemotaxing cells, is an example of a step disturbance rejection, a process known to biologists as perfect adaptation. Unlike other cells, however, the initial first peak observed in the chemoattractant-induced response of Dictyostelium cells is then followed by a slower, smaller phase peaking approximately one to two minutes after the stimulus. Until recently, the nature of this biphasic response has been poorly understood. Moreover, the origin for the second phase is unknown. In this paper we conjecture the existence of a feedback path between the response and stimulus. Using a mathematical model of the chemoattractant-induced response in cells, and standard tools from control engineering, we show that positive feedback may elicit this second peak. PMID:17401451

  18. Early stimulation and late inhibition of peroxisome proliferator-activated receptor gamma (PPAR gamma) gene expression by transforming growth factor beta in human aortic smooth muscle cells: role of early growth-response factor-1 (Egr-1), activator protein 1 (AP1) and Smads.

    PubMed Central

    Fu, Mingui; Zhang, Jifeng; Lin, Yimin; Zhu, Xiaojun; Zhao, Luning; Ahmad, Mushtaq; Ehrengruber, Markus U; Chen, Yuqing E

    2003-01-01

    Transforming growth factor beta (TGF beta) and peroxisome proliferator-activated receptor gamma (PPAR gamma) play major roles in the development of vascular diseases. It has been documented that PPAR gamma activation inhibits the TGF beta signal pathway in vascular smooth muscle cells (VSMC). Here we examined whether TGF beta can regulate PPAR gamma expression. Northern blot analyses revealed that both TGF beta 1 and 2 exert a biphasic effect (early stimulation and late repression) on PPAR gamma gene expression in VSMC. TGF beta rapidly and transiently induced early growth-response factor-1 (Egr-1) expression through the mitogen-activated protein kinase/extracellular signal-regulated kinase kinase 1 (MEK1)/ERK-mediated pathway. Inhibition of MEK1/ERK by PD98059 not only abrogated the induction of Egr-1 but also abolished the rapid and transient induction of PPAR gamma by TGF beta. Furthermore, overexpression of NAB2, a repressor of Egr-1 activation, also blocked the induction of PPAR gamma by TGF beta in VSMC, suggesting that Egr-1 mediates the rapid and transient induction of PPAR gamma by TGF beta. With regard to the TGF beta repression of PPAR gamma expression, activator protein 1 (AP1) and Smad3/4 dramatically inhibited the PPAR gamma promoter activity in transient-transfection studies. In contrast, adenovirus-mediated overexpression of a dominant-negative form of c-Jun partially rescued the TGF beta-induced PPAR gamma repression in VSMC. Taken together, our data demonstrate that Egr-1, AP1 and Smad are part components of the TGF beta signal transduction pathway that regulates PPAR gamma expression. PMID:12457461

  19. Hydrogen peroxide induces murine macrophage chemokine gene transcription via extracellular signal-regulated kinase- and cyclic adenosine 5'-monophosphate (cAMP)-dependent pathways: involvement of NF-kappa B, activator protein 1, and cAMP response element binding protein.

    PubMed

    Jaramillo, Maritza; Olivier, Martin

    2002-12-15

    Hydrogen peroxide (H(2)O(2)) has been shown to act as a second messenger that activates chemokine expression. In the present study, we investigated the mechanisms underlying this cellular regulation in the murine macrophage cell line B10R. We report that H(2)O(2) increases mRNA expression of various chemokines, macrophage-inflammatory protein (MIP)-1alpha/CC chemokine ligand (CCL)3, MIP-1beta/CCL4, MIP-2/CXC chemokine ligand 2, and monocyte chemoattractant protein-1/CCL2, by activating the extracellular signal-regulated kinase (ERK) pathway and the nuclear translocation of the transcription factors NF-kappaB, AP-1, and CREB. Blockage of the ERK pathway with specific inhibitors against mitogen-activated protein kinase kinase 1/2 and ERK1/ERK2 completely abolished both the H(2)O(2)-mediated chemokine up-regulation and the activation of all NF studied. Similarly, selective inhibition of cAMP and NF-kappaB strongly down-regulated the induction of all chemokine transcripts as well as CREB and NF-kappaB activation, respectively. Of interest, we detected a significant decrease of NF-kappaB, AP-1, and CREB DNA binding activities by reciprocal competition for these binding sites when either specific cold oligonucleotides (NF-kappaB, AP-1, and CREB) or Abs against various transcription factor subunits (p50, p65, c-Fos, Jun B, c-Jun, and CREB-1) were added. These findings indicate that cooperation between ERK- and cAMP-dependent pathways seems to be required to achieve the formation of an essential transcriptional factor complex for maximal H(2)O(2)-dependent chemokine modulation. Finally, experiments performed with actinomycin D suggest that H(2)O(2)-mediated MIP-1beta mRNA up-regulation results from transcriptional control, whereas that of MIP-1alpha, MIP-2, and monocyte chemoattractant protein-1 is due to both gene transcription activation and mRNA posttranscriptional stabilization.

  20. X-box-binding protein 1-modified neural stem cells for treatment of Parkinson's disease.

    PubMed

    Si, Lihui; Xu, Tianmin; Wang, Fengzhang; Liu, Qun; Cui, Manhua

    2012-04-01

    X-box-binding protein 1-transfected neural stem cells were transplanted into the right lateral ventricles of rats with rotenone-induced Parkinson's disease. The survival capacities and differentiation rates of cells expressing the dopaminergic marker tyrosine hydroxylase were higher in X-box-binding protein 1-transfected neural stem cells compared to non-transfected cells. Moreover, dopamine and 3,4-dihydroxyphenylacetic acid levels in the substantia nigra were significantly increased, α-synuclein expression was decreased, and neurological behaviors were significantly ameliorated in rats following transplantation of X-box-binding protein 1-transfected neural stem cells. These results indicate that transplantation of X-box-binding protein 1-transfected neural stem cells can promote stem cell survival and differentiation into dopaminergic neurons, increase dopamine and 3,4-dihydroxyphenylacetic acid levels, reduce α-synuclein aggregation in the substantia nigra, and improve the symptoms of Parkinson's disease in rats.

  1. Identification of formyl peptides from Listeria monocytogenes and Staphylococcus aureus as potent chemoattractants for mouse neutrophils 1

    PubMed Central

    Southgate, Erica L.; He, Rong L.; Gao, Ji-Liang; Murphy, Philip M.; Nanamori, Masakatsu; Ye, Richard D.

    2009-01-01

    The prototypic formyl peptide N-formyl-Met-Leu-Phe (fMLF) is a major chemoattractant found in Escherichia coli culture supernatants and a potent agonist at human formyl peptide receptor (FPR) 1. Consistent with this, fMLF induces bactericidal functions in human neutrophils at nanomolar concentrations. However, it is a much less potent agonist for mouse FPR (mFPR) 1 and mouse neutrophils, requiring micromolar concentrations for cell activation. To determine whether other bacteria produce more potent agonists for mFPR1, we examined formyl peptides from Listeria monocytogenes and Staphylococcus aureus for their abilities to activate mouse neutrophils. A pentapeptide (N-formyl-Met-Ile-Val-Ile-Leu (fMIVIL)) from L. monocytogenes and a tetrapeptide (N-formyl-Met-Ile-Phe-Leu (fMIFL)) from S. aureus were found to induce mouse neutrophil chemotaxis at 1-10 nM and superoxide production at 10-100 nM, similar to the potency of fMLF on human neutrophils. Using transfected cell lines expressing mFPR1 and mFPR2, which are major forms of FPRs in mouse neutrophils, we found that mFPR1 is responsible for the high potency of fMIVIL and fMIFL. In comparison, activation of mFPR2 requires micromolar concentrations of the two peptides. Genetic deletion of mfpr1 resulted in abrogation of neutrophil superoxide production and degranulation in response to fMIVIL and fMIFL, further demonstrating that mFPR1 is the primary receptor for detection of these formyl peptides. In conclusion, the formyl peptides from L. monocytogenes and S. aureus are 100-fold more potent than fMLF in activating mouse neutrophils. The ability of mFPR1 to detect bacterially derived formyl peptides indicates that this important host defense mechanism is conserved in mice. PMID:18606697

  2. Slime mold solves maze in one pass, assisted by gradient of chemo-attractants.

    PubMed

    Adamatzky, Andrew

    2012-06-01

    Plasmodium of Physarum polycephalum is a large cell, visible by unaided eye, which exhibits sophisticated patterns of foraging behaviour. The plasmodium's behaviour is well interpreted in terms of computation, where data are spatially extended configurations of nutrients and obstacles, and results of computation are networks of protoplasmic tubes formed by the plasmodium. In laboratory experiments and numerical simulation we show that if plasmodium of P. polycephalum is inoculated in a maze's peripheral channel and an oat flake (source of attractants) in a the maze's central chamber then the plasmodium grows toward target oat flake and connects the flake with the site of original inoculation with a pronounced protoplasmic tube. The protoplasmic tube represents a path in the maze. The plasmodium solves maze in one pass because it is assisted by a gradient of chemo-attractants propagating from the target oat flake.

  3. Expression patterns of cytokines and chemokines genes in human hepatoma cells.

    PubMed

    Shin, Eui-Cheol; Choi, Youn-Hee; Kim, Ji Su; Kim, Se Jong; Park, Jeon Han

    2002-10-01

    Various cytokines and chemokines play a role in carcinogenesis. However, no study has previously been undertaken to investigate comprehensively the expressions of cytokines and chemokines in hepatoma cells. In this study, we determined which cytokines and chemokines are expressed in hepatoma cells. Recently, it was reported that the expressions of several chemokines could be increased by Fas stimulus in many normal and cancer cells. Therefore, we also investigated whether chemokines expression is regulated by Fas ligation. To address this issue, we performed RNase protection assays upon 13 cytokines and 8 chemokines genes in 10 human hepatoma cell lines, comprising 8 hepatitis B virus (HBV)-associated hepatoma cell lines. Transforming growth factor-beta2 (TGF-beta2) was found to be expressed in 8 HBV-associated hepatoma cell lines, and to be potently expressed in 5 cell lines; however, the mRNA expressions of interleukin-10 (IL-10), IL-12, interferon-gamma(IFN-gamma) and tumor necrosis factor-alpha(TNF-alpha) were not detected in any cell lines examined. Among the chemokines investigated in this study, IL-8 was expressed by 8 HBV- associated hepatoma cell lines, and monocyte chemoattractant protein-1 (MCP-1) by 7 HBV-associated hepatoma cell lines. However, the mRNA expressions of macrophage inflammatory protein-1alpha(MIP-1alpha), MIP-1beta, interferon-inducible protein-10 (IP-10), RANTES, lymphotactin and I-309 were either very weak or undetectable. Fas ligation did not increase chemokines expression in hepatoma cells. Conclusively, TGF-beta2, IL-8 and MCP-1 were overexpressed in HBV-associated hepatoma cells, and the expressions of chemokines were not increased by Fas ligation in human hepatoma cells.

  4. Complex chemoattractive and chemorepellent Kit signals revealed by direct imaging of murine mast cells in microfluidic gradient chambers†

    PubMed Central

    Shamloo, Amir; Manchandia, Milan; Ferreira, Meghaan; Mani, Maheswaran; Nguyen, Christopher; Jahn, Thomas; Weinberg, Kenneth

    2014-01-01

    Besides its cooperating effects on stem cell proliferation and survival, Kit ligand (KL) is a potent chemotactic protein. While transwell assays permit studies of the frequency of migrating cells, the lack of direct visualization precludes dynamic chemotaxis studies. In response, we utilize microfluidic chambers that enable direct observation of murine bone marrow-derived mast cells (BMMC) within stable KL gradients. Using this system, individual Kit+ BMMC were quantitatively analyzed for migration speed and directionality during KL-induced chemotaxis. Our results indicated a minimum activating threshold of ~3 ng ml−1 for chemoattraction. Analysis of cells at KL concentrations below 3 ng ml−1 revealed a paradoxical chemorepulsion, which has not been described previously. Unlike chemoattraction, which occurred continuously after an initial time lag, chemorepulsion occurred only during the first 90 minutes of observation. Both chemoattraction and chemorepulsion required the action of G-protein coupled receptors (GPCR), as treatment with pertussis toxin abrogated directed migration. These results differ from previous studies of GPCR-mediated chemotaxis, where chemorepulsion occurred at high ligand concentrations. These data indicate that Kit-mediated chemotaxis is more complex than previously understood, with the involvement of GPCRs in addition to the Kit receptor tyrosine kinase and the presence of both chemoattractive and chemorepellent phases. PMID:23835699

  5. AUXIN BINDING PROTEIN1: The Outsider

    PubMed Central

    Sauer, Michael; Kleine-Vehn, Jürgen

    2011-01-01

    AUXIN BINDING PROTEIN1 (ABP1) is one of the first characterized proteins that bind auxin and has been implied as a receptor for a number of auxin responses. Early studies characterized its auxin binding properties and focused on rapid electrophysiological and cell expansion responses, while subsequent work indicated a role in cell cycle and cell division control. Very recently, ABP1 has been ascribed a role in modulating endocytic events at the plasma membrane and RHO OF PLANTS-mediated cytoskeletal rearrangements during asymmetric cell expansion. The exact molecular function of ABP1 is still unresolved, but its main activity apparently lies in influencing events at the plasma membrane. This review aims to connect the novel findings with the more classical literature on ABP1 and to point out the many open questions that still separate us from a comprehensive model of ABP1 action, almost 40 years after the first reports of its existence. PMID:21719690

  6. Macrophage Migration Inhibitor Factor Upregulates MCP-1 Expression in an Autocrine Manner in Hepatocytes during Acute Mouse Liver Injury

    PubMed Central

    Xie, Jieshi; Yang, Le; Tian, Lei; Li, Weiyang; Yang, Lin; Li, Liying

    2016-01-01

    Macrophage migration inhibitor factor (MIF), a multipotent innate immune mediator, is an upstream component of the inflammatory cascade in diseases such as liver disease. Monocyte chemoattractant protein-1 (MCP-1), a highly representative chemokine, is critical in liver disease pathogenesis. We investigated the role of MIF in regulating hepatocytic MCP-1 expression. MIF and MCP-1 expression were characterized by immunochemistry, RT-PCR, ELISA, and immunoblotting in CCl4-treated mouse liver and isolated hepatocytes. MIF was primarily distributed in hepatocytes, and its expression increased upon acute liver injury. Its expression was also increased in injured hepatocytes, induced by LPS or CCl4, which mimic liver injury in vitro. MIF was expressed earlier than MCP-1, strongly inducing hepatocytic MCP-1 expression. Moreover, the increase in MCP-1 expression induced by MIF was inhibited by CD74- or CD44-specific siRNAs and SB203580, a p38 MAPK inhibitor. Further, CD74 or CD44 deficiency effectively inhibited MIF-induced p38 activation. MIF inhibitor ISO-1 reduced MCP-1 expression and p38 phosphorylation in CCl4-treated mouse liver. Our results showed that MIF regulates MCP-1 expression in hepatocytes of injured liver via CD74, CD44, and p38 MAPK in an autocrine manner, providing compelling information on the role of MIF in liver injury, and implying a new regulatory mechanism for liver inflammation. PMID:27273604

  7. Blunted activation of NF-{kappa}B and NF-{kappa}B-dependent gene expression by geranylgeranylacetone: Involvement of unfolded protein response

    SciTech Connect

    Hayakawa, Kunihiro; Hiramatsu, Nobuhiko; Okamura, Maro; Yao, Jian; Paton, Adrienne W.; Paton, James C.; Kitamura, Masanori

    2008-01-04

    Geranylgeranylacetone (GGA), an anti-ulcer agent, has anti-inflammatory potential against experimental colitis and ischemia-induced renal inflammation. However, molecular mechanisms involved in its anti-inflammatory effects are largely unknown. We found that, in glomerular mesangial cells, GGA blocked activation of nuclear factor-{kappa}B and consequent induction of monocyte chemoattractant protein 1 (MCP-1) by inflammatory cytokines. It was inversely correlated with induction of unfolded protein response (UPR) evidenced by expression of 78 kDa glucose-regulated protein (GRP78) and suppression of endoplasmic reticulum stress-responsive alkaline phosphatase. Various inducers of UPR including tunicamycin, thapsigargin, A23187, 2-deoxyglucose, dithiothreitol, and AB{sub 5} subtilase cytotoxin reproduced the suppressive effects of GGA. Furthermore, attenuation of UPR by stable transfection with GRP78 diminished the anti-inflammatory effects of GGA. These results disclosed a novel, UPR-dependent mechanism underlying the anti-inflammatory potential of GGA.

  8. Early Cytokine and Chemokine Gene Expression during Pseudomonas aeruginosa Corneal Infection in Mice

    PubMed Central

    Kernacki, Karen A.; Goebel, Dennis J.; Poosch, Michael S.; Hazlett, Linda D.

    1998-01-01

    Using a multiprobe RNase protection assay, we examined cytokine and chemokine mRNAs that were expressed after corneal infection with Pseudomonas aeruginosa in mice. Cytokines that were upregulated included interleukin-1α (IL-1α) and -1β, IL-1 receptor antagonist, IL-6, IL-11, granulocyte colony-stimulating factor, granulocyte-macrophage colony-stimulating factor, macrophage colony-stimulating factor, stem cell factor, lymphotoxin β, transforming growth factor β1, and tumor necrosis factor alpha. Chemokine transcripts that were upregulated included Eotaxin; gamma-interferon-inducible protein 10; monocyte chemoattractant protein 1; macrophage inflammatory proteins 1α, 1β, and 2; and RANTES. Peak expression of these cytokines and chemokines was observed between 1 and 3 days after infection. These responses returned to or approached baseline preinfection levels by 7 days after ocular challenge. Identification of the various cytokines and chemokines upregulated during corneal infection provides important information relevant to unraveling the pathogenesis induced by this bacterium and provides hope that specific molecules can be targeted for therapy. PMID:9423885

  9. Self-Generated Chemoattractant Gradients: Attractant Depletion Extends the Range and Robustness of Chemotaxis

    PubMed Central

    Tweedy, Luke; Knecht, David A.; Mackay, Gillian M.; Insall, Robert H.

    2016-01-01

    Chemotaxis is fundamentally important, but the sources of gradients in vivo are rarely well understood. Here, we analyse self-generated chemotaxis, in which cells respond to gradients they have made themselves by breaking down globally available attractants, using both computational simulations and experiments. We show that chemoattractant degradation creates steep local gradients. This leads to surprising results, in particular the existence of a leading population of cells that moves highly directionally, while cells behind this group are undirected. This leading cell population is denser than those following, especially at high attractant concentrations. The local gradient moves with the leading cells as they interact with their surroundings, giving directed movement that is unusually robust and can operate over long distances. Even when gradients are applied from external sources, attractant breakdown greatly changes cells' responses and increases robustness. We also consider alternative mechanisms for directional decision-making and show that they do not predict the features of population migration we observe experimentally. Our findings provide useful diagnostics to allow identification of self-generated gradients and suggest that self-generated chemotaxis is unexpectedly universal in biology and medicine. PMID:26981861

  10. Self-Generated Chemoattractant Gradients: Attractant Depletion Extends the Range and Robustness of Chemotaxis.

    PubMed

    Tweedy, Luke; Knecht, David A; Mackay, Gillian M; Insall, Robert H

    2016-03-01

    Chemotaxis is fundamentally important, but the sources of gradients in vivo are rarely well understood. Here, we analyse self-generated chemotaxis, in which cells respond to gradients they have made themselves by breaking down globally available attractants, using both computational simulations and experiments. We show that chemoattractant degradation creates steep local gradients. This leads to surprising results, in particular the existence of a leading population of cells that moves highly directionally, while cells behind this group are undirected. This leading cell population is denser than those following, especially at high attractant concentrations. The local gradient moves with the leading cells as they interact with their surroundings, giving directed movement that is unusually robust and can operate over long distances. Even when gradients are applied from external sources, attractant breakdown greatly changes cells' responses and increases robustness. We also consider alternative mechanisms for directional decision-making and show that they do not predict the features of population migration we observe experimentally. Our findings provide useful diagnostics to allow identification of self-generated gradients and suggest that self-generated chemotaxis is unexpectedly universal in biology and medicine.

  11. Structure Activity Relationships of Monocyte Chemoattractant Proteins in Complex with a Blocking Antibody

    SciTech Connect

    Reid,C.; Rushe, M.; Jarpe, M.; Van Vlijmen, H.; Dolinski, B.; Qian, F.; Cachero, T.; Cuervo, H.; Yanachkova, M.; et al.

    2006-01-01

    Monocyte chemoattractant proteins (MCPs) are cytokines that direct immune cells bearing appropriate receptors to sites of inflammation or injury and are therefore attractive therapeutic targets for inhibitory molecules. 11K2 is a blocking mouse monoclonal antibody active against several human and murine MCPs. A 2.5 Angstroms structure of the Fab fragment of this antibody in complex with human MCP-1 has been solved. The Fab blocks CCR2 receptor binding to MCP-1 through an adjacent but distinct binding site. The orientation of the Fab indicates that a single MCP-1 dimer will bind two 11K2 antibodies. Several key residues on the antibody and on human MCPs were predicted to be involved in antibody selectivity. Mutational analysis of these residues confirms their involvement in the antibody- chemokine interaction. In addition to mutations that decreased or disrupted binding, one antibody mutation resulted in a 70-fold increase in affinity for human MCP-2. A key residue missing in human MCP-3, a chemokine not recognized by the antibody, was identified and engineering the preferred residue into the chemokine conferred binding to the antibody.

  12. Complete amino acid sequence of a human monocyte chemoattractant, a putative mediator of cellular immune reactions.

    PubMed Central

    Robinson, E A; Yoshimura, T; Leonard, E J; Tanaka, S; Griffin, P R; Shabanowitz, J; Hunt, D F; Appella, E

    1989-01-01

    In a study of the structural basis for leukocyte specificity of chemoattractants, we determined the complete amino acid sequence of human glioma-derived monocyte chemotactic factor (GDCF-2), a peptide that attracts human monocytes but not neutrophils. The choice of a tumor cell product for analysis was dictated by its relative abundance and an amino acid composition indistinguishable from that of lymphocyte-derived chemotactic factor (LDCF), the agonist thought to account for monocyte accumulation in cellular immune reactions. By a combination of Edman degradation and mass spectrometry, it was established that GDCF-2 comprises 76 amino acid residues, commencing at the N terminus with pyroglutamic acid. The peptide contains four half-cystines, at positions 11, 12, 36, and 52, which create a pair of loops, clustered at the disulfide bridges. The relative positions of the half-cystines are almost identical to those of monocyte-derived neutrophil chemotactic factor (MDNCF), a peptide of similar mass but with only 24% sequence identity to GDCF. Thus, GDCF and MDNCF have a similar gross secondary structure because of the loops formed by the clustered disulfides, and their different leukocyte specificities are most likely determined by the large differences in primary sequence. PMID:2648385

  13. A Knockout Mutation of a Constitutive GPCR in Tetrahymena Decreases Both G-Protein Activity and Chemoattraction

    PubMed Central

    Lampert, Thomas J.; Coleman, Kevin D.; Hennessey, Todd M.

    2011-01-01

    Although G-protein coupled receptors (GPCRs) are a common element in many chemosensory transduction pathways in eukaryotic cells, no GPCR or regulated G-protein activity has yet been shown in any ciliate. To study the possible role for a GPCR in the chemoresponses of the ciliate Tetrahymena, we have generated a number of macronuclear gene knockouts of putative GPCRs found in the Tetrahymena Genome database. One of these knockout mutants, called G6, is a complete knockout of a gene that we call GPCR6 (TTHERM_00925490). Based on sequence comparisons, the Gpcr6p protein belongs to the Rhodopsin Family of GPCRs. Notably, Gpcr6p shares highest amino acid sequence homologies to GPCRs from Paramecium and several plants. One of the phenotypes of the G6 mutant is a decreased responsiveness to the depolarizing ions Ba2+ and K+, suggesting a decrease in basal excitability (decrease in Ca2+ channel activity). The other major phenotype of G6 is a loss of chemoattraction to lysophosphatidic acid (LPA) and proteose peptone (PP), two known chemoattractants in Tetrahymena. Using microsomal [35S]GTPγS binding assays, we found that wild-type (CU427) have a prominent basal G-protein activity. This activity is decreased to the same level by pertussis toxin (a G-protein inhibitor), addition of chemoattractants, or the G6 mutant. Since the basal G-protein activity is decreased by the GPCR6 knockout, it is likely that this gene codes for a constitutively active GPCR in Tetrahymena. We propose that chemoattractants like LPA and PP cause attraction in Tetrahymena by decreasing the basal G-protein stimulating activity of Gpcr6p. This leads to decreased excitability in wild-type and longer runs of smooth forward swimming (less interrupted by direction changes) towards the attractant. Therefore, these attractants may work as inverse agonists through the constitutively active Gpcr6p coupled to a pertussis-sensitive G-protein. PMID:22140501

  14. Gene expression profiling in spleens of deoxynivalenol-exposed mice: immediate early genes as primary targets.

    PubMed

    Kinser, Shawn; Jia, Qunshan; Li, Maioxing; Laughter, Ashley; Cornwell, Paul; Corton, J Christopher; Pestka, James

    2004-09-24

    Exposure to the trichothecene mycotoxin deoxynivalenol (DON) alters immune functions in vitro and in vivo. To gain further insight into DON's immunotoxic effects, microarrays were used to determine how acute exposure to this mycotoxin modulates gene expression profiles in murine spleen. B6C3F1 mice were treated orally with 25mg/kg body weight DON, and 2h later spleens were collected for macroarray analysis. Following normalization using a local linear regression model, expression of 116 out of 1176 genes was significantly altered compared to average expression levels in all treatment groups. When genes were arranged into an ontology tree to facilitate comparison of expression profiles between treatment groups, DON was found primarily to modulate genes associated with immunity, inflammation, and chemotaxis. Real-time polymerase chain reaction was used to confirm modulation for selected genes. DON was found to induce the cytokines interleukin (IL)-1alpha, IL-1beta, IL-6 and IL-11. In analogous fashion, DON upregulated expression of the chemokines macrophage inhibitory protein-2 (MIP-2), cytokine-induced chemoattractant protein-1 (CINC-1), monocyte chemoattractant protein (MCP)-1, MCP-3, and cytokine-responsive gene-2 (CRG-2). c-Fos, Fra-, c-Jun, and JunB, components of the activator protein-1 (AP-1) transcription factor complex, were induced by DON as well as another transcription factor, NR4A1. Four hydrolases were found to be upregulated by DON, including mitogen-activated protein kinase phosphatase 1 (MKP1), catalytic subunit beta isoform (CnAbeta), protein tyrosine phosphatase receptor type J (Ptprj), and protein tyrosine phosphatase nonreceptor type 8 (Ptpn8), whereas three other hydrolases, microsomal epoxide hydrolase (Eph) 1, histidine triad nucleotide binding protein (Hint), and proteosome subunit beta type 8 (Psmb8) were significantly decreased by the toxin. Finally, cysteine-rich protein 61 (CRP61) and heat-shock protein 40 (Hsp40), genes associated with

  15. Identification of a Chemoattractant G-Protein-Coupled Receptor for Folic Acid that Controls Both Chemotaxis and Phagocytosis.

    PubMed

    Pan, Miao; Xu, Xuehua; Chen, Yong; Jin, Tian

    2016-02-22

    Eukaryotic phagocytes search and destroy invading microorganisms via chemotaxis and phagocytosis. The social amoeba Dictyostelium discoideum is a professional phagocyte that chases bacteria through chemotaxis and engulfs them as food via phagocytosis. G-protein-coupled receptors (GPCRs) are known for detecting chemoattractants and directing cell migration, but their roles in phagocytosis are not clear. Here, we developed a quantitative phosphoproteomic technique to discover signaling components. Using this approach, we discovered the long sought after folic acid receptor, fAR1, in D. discoideum. We showed that the seven-transmembrane receptor fAR1 is required for folic acid-mediated signaling events. Significantly, we discovered that fAR1 is essential for both chemotaxis and phagocytosis of bacteria, thereby representing a chemoattractant GPCR that mediates not only chasing but also ingesting bacteria. We revealed that a phagocyte is able to internalize particles via a chemoattractant-mediated engulfment process. We propose that mammalian phagocytes may also use this mechanism to engulf and ingest bacterial pathogens.

  16. Identification of a Chemoattractant G-Protein-Coupled Receptor for Folic Acid that Controls Both Chemotaxis and Phagocytosis.

    PubMed

    Pan, Miao; Xu, Xuehua; Chen, Yong; Jin, Tian

    2016-02-22

    Eukaryotic phagocytes search and destroy invading microorganisms via chemotaxis and phagocytosis. The social amoeba Dictyostelium discoideum is a professional phagocyte that chases bacteria through chemotaxis and engulfs them as food via phagocytosis. G-protein-coupled receptors (GPCRs) are known for detecting chemoattractants and directing cell migration, but their roles in phagocytosis are not clear. Here, we developed a quantitative phosphoproteomic technique to discover signaling components. Using this approach, we discovered the long sought after folic acid receptor, fAR1, in D. discoideum. We showed that the seven-transmembrane receptor fAR1 is required for folic acid-mediated signaling events. Significantly, we discovered that fAR1 is essential for both chemotaxis and phagocytosis of bacteria, thereby representing a chemoattractant GPCR that mediates not only chasing but also ingesting bacteria. We revealed that a phagocyte is able to internalize particles via a chemoattractant-mediated engulfment process. We propose that mammalian phagocytes may also use this mechanism to engulf and ingest bacterial pathogens. PMID:26906738

  17. Low density lipoprotein receptor related protein 1 variant interacts with saturated fatty acids in Puerto Ricans

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Low density lipoprotein related receptor protein 1 (LRP1) is a multi-functional endocytic receptor that is highly expressed in adipocytes and the hypothalamus. Animal models and in vitro studies support a role for LRP1 in adipocyte metabolism and leptin signaling, but genetic polymorphisms have not ...

  18. PTPRT regulates the interaction of Syntaxin-binding protein 1 with Syntaxin 1 through dephosphorylation of specific tyrosine residue

    SciTech Connect

    Lim, So-Hee; Moon, Jeonghee; Lee, Myungkyu; Lee, Jae-Ran

    2013-09-13

    Highlights: •PTPRT is a brain-specific, expressed, protein tyrosine phosphatase. •PTPRT regulated the interaction of Syntaxin-binding protein 1 with Syntaxin 1. •PTPRT dephosphorylated the specific tyrosine residue of Syntaxin-binding protein 1. •Dephosphorylation of Syntaxin-binding protein 1 enhanced the interaction with Syntaxin 1. •PTPRT appears to regulate the fusion of synaptic vesicle through dephosphorylation. -- Abstract: PTPRT (protein tyrosine phosphatase receptor T), a brain-specific tyrosine phosphatase, has been found to regulate synaptic formation and development of hippocampal neurons, but its regulation mechanism is not yet fully understood. Here, Syntaxin-binding protein 1, a key component of synaptic vesicle fusion machinery, was identified as a possible interaction partner and an endogenous substrate of PTPRT. PTPRT interacted with Syntaxin-binding protein 1 in rat synaptosome, and co-localized with Syntaxin-binding protein 1 in cultured hippocampal neurons. PTPRT dephosphorylated tyrosine 145 located around the linker between domain 1 and 2 of Syntaxin-binding protein 1. Syntaxin-binding protein 1 directly binds to Syntaxin 1, a t-SNARE (soluble N-ethylmaleimide-sensitive factor attachment protein receptor) protein, and plays a role as catalysts of SNARE complex formation. Syntaxin-binding protein 1 mutant mimicking non-phosphorylation (Y145F) enhanced the interaction with Syntaxin 1 compared to wild type, and therefore, dephosphorylation of Syntaxin-binding protein 1 appeared to be important for SNARE-complex formation. In conclusion, PTPRT could regulate the interaction of Syntaxin-binding protein 1 with Syntaxin 1, and as a result, the synaptic vesicle fusion appeared to be controlled through dephosphorylation of Syntaxin-binding protein 1.

  19. Iron Regulatory Protein-1 Protects against Mitoferrin-1-deficient Porphyria*

    PubMed Central

    Chung, Jacky; Anderson, Sheila A.; Gwynn, Babette; Deck, Kathryn M.; Chen, Michael J.; Langer, Nathaniel B.; Shaw, George C.; Huston, Nicholas C.; Boyer, Leah F.; Datta, Sumon; Paradkar, Prasad N.; Li, Liangtao; Wei, Zong; Lambert, Amy J.; Sahr, Kenneth; Wittig, Johannes G.; Chen, Wen; Lu, Wange; Galy, Bruno; Schlaeger, Thorsten M.; Hentze, Matthias W.; Ward, Diane M.; Kaplan, Jerry; Eisenstein, Richard S.; Peters, Luanne L.; Paw, Barry H.

    2014-01-01

    Mitochondrial iron is essential for the biosynthesis of heme and iron-sulfur ([Fe-S]) clusters in mammalian cells. In developing erythrocytes, iron is imported into the mitochondria by MFRN1 (mitoferrin-1, SLC25A37). Although loss of MFRN1 in zebrafish and mice leads to profound anemia, mutant animals showed no overt signs of porphyria, suggesting that mitochondrial iron deficiency does not result in an accumulation of protoporphyrins. Here, we developed a gene trap model to provide in vitro and in vivo evidence that iron regulatory protein-1 (IRP1) inhibits protoporphyrin accumulation. Mfrn1+/gt;Irp1−/− erythroid cells exhibit a significant increase in protoporphyrin levels. IRP1 attenuates protoporphyrin biosynthesis by binding to the 5′-iron response element (IRE) of alas2 mRNA, inhibiting its translation. Ectopic expression of alas2 harboring a mutant IRE, preventing IRP1 binding, in Mfrn1gt/gt cells mimics Irp1 deficiency. Together, our data support a model whereby impaired mitochondrial [Fe-S] cluster biogenesis in Mfrn1gt/gt cells results in elevated IRP1 RNA-binding that attenuates ALAS2 mRNA translation and protoporphyrin accumulation. PMID:24509859

  20. Iron regulatory protein-1 protects against mitoferrin-1-deficient porphyria.

    PubMed

    Chung, Jacky; Anderson, Sheila A; Gwynn, Babette; Deck, Kathryn M; Chen, Michael J; Langer, Nathaniel B; Shaw, George C; Huston, Nicholas C; Boyer, Leah F; Datta, Sumon; Paradkar, Prasad N; Li, Liangtao; Wei, Zong; Lambert, Amy J; Sahr, Kenneth; Wittig, Johannes G; Chen, Wen; Lu, Wange; Galy, Bruno; Schlaeger, Thorsten M; Hentze, Matthias W; Ward, Diane M; Kaplan, Jerry; Eisenstein, Richard S; Peters, Luanne L; Paw, Barry H

    2014-03-14

    Mitochondrial iron is essential for the biosynthesis of heme and iron-sulfur ([Fe-S]) clusters in mammalian cells. In developing erythrocytes, iron is imported into the mitochondria by MFRN1 (mitoferrin-1, SLC25A37). Although loss of MFRN1 in zebrafish and mice leads to profound anemia, mutant animals showed no overt signs of porphyria, suggesting that mitochondrial iron deficiency does not result in an accumulation of protoporphyrins. Here, we developed a gene trap model to provide in vitro and in vivo evidence that iron regulatory protein-1 (IRP1) inhibits protoporphyrin accumulation. Mfrn1(+/gt);Irp1(-/-) erythroid cells exhibit a significant increase in protoporphyrin levels. IRP1 attenuates protoporphyrin biosynthesis by binding to the 5'-iron response element (IRE) of alas2 mRNA, inhibiting its translation. Ectopic expression of alas2 harboring a mutant IRE, preventing IRP1 binding, in Mfrn1(gt/gt) cells mimics Irp1 deficiency. Together, our data support a model whereby impaired mitochondrial [Fe-S] cluster biogenesis in Mfrn1(gt/gt) cells results in elevated IRP1 RNA-binding that attenuates ALAS2 mRNA translation and protoporphyrin accumulation.

  1. Glycosylation of Dentin Matrix Protein 1 is critical for osteogenesis

    PubMed Central

    Sun, Yao; Weng, Yuteng; Zhang, Chenyang; Liu, Yi; Kang, Chen; Liu, Zhongshuang; Jing, Bo; Zhang, Qi; Wang, Zuolin

    2015-01-01

    Proteoglycans play important roles in regulating osteogenesis. Dentin matrix protein 1 (DMP1) is a highly expressed bone extracellular matrix protein that regulates both bone development and phosphate metabolism. After glycosylation, an N-terminal fragment of DMP1 protein was identified as a new proteoglycan (DMP1-PG) in bone matrix. In vitro investigations showed that Ser89 is the key glycosylation site in mouse DMP1. However, the specific role of DMP1 glycosylation is still not understood. In this study, a mutant DMP1 mouse model was developed in which the glycosylation site S89 was substituted with G89 (S89G-DMP1). The glycosylation level of DMP1 was down-regulated in the bone matrix of S89G-DMP1 mice. Compared with wild type mice, the long bones of S89G-DMP1 mice showed developmental changes, including the speed of bone remodeling and mineralization, the morphology and activities of osteocytes, and activities of both osteoblasts and osteoclasts. These findings indicate that glycosylation of DMP1 is a key posttranslational modification process during development and that DMP1-PG functions as an indispensable proteoglycan in osteogenesis. PMID:26634432

  2. Glycosylation of Dentin Matrix Protein 1 is critical for osteogenesis.

    PubMed

    Sun, Yao; Weng, Yuteng; Zhang, Chenyang; Liu, Yi; Kang, Chen; Liu, Zhongshuang; Jing, Bo; Zhang, Qi; Wang, Zuolin

    2015-12-04

    Proteoglycans play important roles in regulating osteogenesis. Dentin matrix protein 1 (DMP1) is a highly expressed bone extracellular matrix protein that regulates both bone development and phosphate metabolism. After glycosylation, an N-terminal fragment of DMP1 protein was identified as a new proteoglycan (DMP1-PG) in bone matrix. In vitro investigations showed that Ser(89) is the key glycosylation site in mouse DMP1. However, the specific role of DMP1 glycosylation is still not understood. In this study, a mutant DMP1 mouse model was developed in which the glycosylation site S(89) was substituted with G(89) (S89G-DMP1). The glycosylation level of DMP1 was down-regulated in the bone matrix of S89G-DMP1 mice. Compared with wild type mice, the long bones of S89G-DMP1 mice showed developmental changes, including the speed of bone remodeling and mineralization, the morphology and activities of osteocytes, and activities of both osteoblasts and osteoclasts. These findings indicate that glycosylation of DMP1 is a key posttranslational modification process during development and that DMP1-PG functions as an indispensable proteoglycan in osteogenesis.

  3. [Production of recombinant human interleukin-38 and its inhibitory effect on the expression of proinflammatory cytokines in THP-1 cells].

    PubMed

    Yuan, X L; Li, Y; Pan, X H; Zhou, M; Gao, Q Y; Li, M C

    2016-01-01

    Interleukin (IL)-38 is the latest member of the IL-1 cytokine family. However, as a result of lacking efficient method to generate relatively large quantity of IL-38, its precise functions are poorly understood. In the present study, the cloning, expression, purification, and activity analysis of recombinant human IL-38 was described. Human IL-38 cDNA was cloned into the prokaryotic expression vector pET-44. The recombinant IL-38 containing a C-hexahistidine tag was expressed in Escherichia coli BL21 (DE3) which induced by isopropyl-β-D-thiogalactoside. The expressed fusion protein was purified by Ni-NTA affinity chromatography. IL-38 protein was largely found in the soluble fraction. The purified IL-38 appeared a single band on SDS-PAGE, the yield of IL-38 was 4 mg from 1 L of bacterial culture, and the purity was more than 98% with low endotoxin level (<0.1 EU/μg). Western blotting confirmed the identity of the purified protein. Activity analysis showed that IL-38 can inhibit effectively the expression of proinflammatory cytokines, such as tumor necrosis factor-α, IL-1β, IL-17, and monocyte chemoattractant protein-1 in lipopolysaccharide-activated THP-1 cells. The production and characterization of biologically active IL-38 will be beneficial for its potential role in clinical applications. PMID:27414784

  4. Folic acid is a potent chemoattractant of free-living amoebae in a new and amazing species of protist, Vahlkampfia sp.

    PubMed

    Maeda, Yasuo; Mayanagi, Taira; Amagai, Aiko

    2009-03-01

    Folic acid (folate; vitamin Bc) is well recognized as essential for the proper metabolism of the essential amino acid methionine as well as for the synthesis of adenine and thymine. A folate deficiency has been Implicated in a wide variety of disorders from Alzheimer's disease to depression and neural tube defects. In the cellular slime molds, including Dictyostelium, vegetative growth-phase cells are known to chemotactically move toward folate that is secreted by bacterial food sources such as Escherichia coli. Intracellular folate signal transductlon, including G proteins, Ca(2+)channels, and the PIP3 pathway, has been reported in D. discoideum. To our surprise, the genuine chemoattractant(s) of free-living protozoan amoebae have remained to be determined, possibly because of lack of a pertinent method for assaying chemotaxis. We recently isolated a primitive free-living amoeba from the soil of Costa Rica and identified it as a new species of the genus Vahlkampfia belonging to Subclass Gymnamoebia, which includes Entamoeba and Acanthamoeba. The amoebae can grow and multiply quite rapidly, engulfing nearby bacteria such as E. coli. Importantly, we have demonstrated here using a quite simple but finely designed chemotaxis assay that the Vahlkampfia amoebae exhibit chemotaxis toward higher folate concentrations. Riboflavin and cyanocobalamin were also found to serve as positive chemoattractants. Among these chemoattractants, folate is of particular importance because its function seems to be evolutionarily conserved as a potent chemoattractant of amoeboid cells in a wide range of organisms as well as in the Protista and cellular slime molds.

  5. Microemboli alter the acute stress response and cause prolonged expression of MCP-1 in the hippocampus.

    PubMed

    Nemeth, Christina L; Neigh, Gretchen N

    2015-04-01

    Microvascular ischemia is linked to cardiovascular disease pathology, as well as alterations in mood and cognition. Ischemia activates the hypothalamic-pituitary-adrenal (HPA) axis and through chronic activation, alters HPA axis function. Dysregulation of the HPA axis can lead to the chronic release of glucocorticoids, a hyper-inflammatory cerebral response, cell damage, and changes in behavior. Although the interactions between injury and HPA axis activity have been established in global ischemia, HPA-related repercussions of diffuse ischemic damage and subsequent inflammation have not been assessed. The current study used a rat model of microsphere embolism (ME) ischemia to test the hypothesis that microvascular ischemia would lead to long term alterations in HPA axis function and inflammatory activity. Furthermore, given the pro-inflammatory nature of chronic stress, we assessed the implications of chronic stress for gene expression of inflammatory factors and key components of the glucocorticoid receptor response, following microvascular ischemia. Results indicated that ME altered the response to an acute stress fourteen days following ME injury and increased hippocampal expression of monocyte chemoattractant protein 1 (Mcp-1) as long as 4 weeks following ME injury, without concomitant effects on gene expression of the glucocorticoid receptor or its co-chaperones. Furthermore, no exacerbative effects of chronic stress exposure were observed following ME injury beyond the effects of ME injury alone. Together, these results indicate that ME injury is sufficient to alter both HPA axis activity and cerebral inflammation for a prolonged period of time following injury.

  6. Shwachman-Diamond syndrome neutrophils have altered chemoattractant-induced F-actin polymerization and polarization characteristics.

    PubMed

    Orelio, Claudia; Kuijpers, Taco W

    2009-03-01

    Shwachman-Diamond syndrome is a hereditary disorder characterized by pancreatic insufficiency and bone marrow failure. Most Shwachman-Diamond syndrome patients have mutations in the SBDS gene located at chromosome 7 and suffer from recurrent infections, due to neutropenia in combination with impaired neutrophil chemotaxis. Currently, the role of the actin cytoskeleton in Shwachman-Diamond syndrome neutrophils has not been investigated. Therefore, we performed immunofluorescence for SBDS and F-actin on human neutrophilic cells. Additionally, we examined in control neutrophils and cells from genetically defined Shwachman-Diamond syndrome patients F-actin polymerization and cytoskeletal polarization characteristics upon chemoattractant stimulation. These studies showed that SBDS and F-actin co-localize in neutrophilic cells and that F-actin polymerization and depolymerization characteristics are altered in Shwachman-Diamond syndrome neutrophils as compared to control neutrophils in response to both fMLP and C5a. Moreover, F-actin cytoskeletal polarization is delayed in Shwachman-Diamond syndrome neutrophils. Thus, Shwachman-Diamond syndrome neutrophils have aberrant chemoattractant-induced F-actin properties which might contribute to the impaired neutrophil chemotaxis.

  7. A double-edged sword: the role of VEGF in wound repair and chemoattraction of opportunist pathogens.

    PubMed

    Birkenhauer, Eric; Neethirajan, Suresh

    2015-01-01

    Wound healing is a complex process essential to repairing damaged tissues and preventing infection. Skin is the first line of defense, a chief physical barrier to microbe entry. Wound healing is a physical rebuilding process, but at the same time it is an inflammatory event. In turn, molecules for wound repair are secreted by fibroblasts and others present at the wound site. Vascular endothelial growth factor (VEGF) is a critical cytokine that exhibits chemoattractant properties, recruiting other immune cells to the site. Although generally beneficial, VEGF may also act as a chemoattractant for invading microorganisms, such as Pseudomonas aeruginosa. P. aeruginosa is problematic during wound infection due to its propensity to form biofilms and exhibit heightened antimicrobial resistance. Here, we explored the influence of VEGF gradients (in a microfluidic device wound model) on the motility and chemotactic properties of P. aeruginosa. At lower concentrations, VEGF had little effect on motility, but as the maximal concentration within the gradient increased, P. aeruginosa cells exhibited directed movement along the gradient. Our data provide evidence that while beneficial, VEGF, in excess, may aid colonization by P. aeruginosa. This highlights the necessity for the efficient resolution of inflammation. Understanding the dynamics of wound colonization may lead to new/enhanced therapeutics to hasten recovery.

  8. Identification and characterization of merozoite surface protein 1 epitope

    PubMed Central

    Singh, Satarudra Prakash; Mishra, Bhartendu Nath

    2009-01-01

    Malaria is an important tropical infection which urgently requires intervention of an effective vaccine. Antigenic variations of the parasite and allelic diversity of the host are main problems in the development of an effective malaria vaccine. Cytotoxic T lymphocytes (CTL) directed against Plasmodium falciparum­derived antigens are shown to play an important role for the protection against malaria. The merozoite surface protein 1 (MSP1) is expressed in all the four life-cycle stages of Plasmodium falciparum and did not find any sequence similarity to human and mouse reference proteins. MSP1 is a known target of the immune response and a single CTL epitope binding to the HLA­A*0201 is available for merozoite form. Here, we report the results from the computational characterization of MSP1, precursor (1720 residue) and screening of highest scoring potential CTL epitopes for 1712 overlapping peptides binding to thirty four HLA class­I alleles and twelve HLA class­I supertypes (5 HLA­A and 7 HLA­B) using bioinformatics tools. Supertypes are the clustered groups of HLA class­I molecules, representing a sets of molecules that share largely overlapping peptide binding specificity. The prediction results for MSP1 as adhesin and adhesin-like in terms of probability is 1.0. Results also show that MSP1 has orthologs to other related species as well as having non allergenicity and single transmembrane properties demonstrating its suitability as a vaccine candidate. The predicted peptides are expected to be useful in the design of multi-epitope vaccines without compromising the human population coverage. PMID:20011145

  9. Molecular basis of cellular localization of poly C binding protein 1 in neuronal cells

    SciTech Connect

    Berry, Andrea M.; Flock, Kelly E.; Loh, Horace H.; Ko, Jane L. . E-mail: kojane@shu.edu

    2006-11-03

    Poly C binding protein 1 (PCBP) is involved in the transcriptional regulation of neuronal mu-opioid receptor gene. In this study, we examined the molecular basis of PCBP cellular/nuclear localization in neuronal cells using EGFP fusion protein. PCBP, containing three KH domains and a variable domain, distributed in cytoplasm and nucleus with a preferential nuclear expression. Domain-deletional analyses suggested the requirement of variable and KH3 domains for strong PCBP nuclear expression. Within the nucleus, a low nucleolar PCBP expression was observed, and PCBP variable domain contributed to this restricted nucleolar expression. Furthermore, the punctate nuclear pattern of PCBP was correlated to its single-stranded (ss) DNA binding ability, with both requiring cooperativity of at least three sequential domains. Collectively, certain PCBP domains thus govern its nuclear distribution and transcriptional regulatory activity in the nucleus of neurons, whereas the low nucleolar expression implicates the disengagement of PCBP in the ribosomal RNA synthesis.

  10. Redirection of doublecortin-positive cell migration by over-expression of the chemokines MCP-1, MIP-1α and GRO-α in the adult rat brain.

    PubMed

    Tang, S K; Knobloch, R A; Maucksch, C; Connor, B

    2014-02-28

    Inflammation-induced chemoattraction plays a major role in adult subventricular zone (SVZ)-derived precursor cell migration following neural cell loss, in particular through the release of chemokines by activated microglia and macrophages. We previously demonstrated that monocyte chemotactic protein-1 (MCP-1) (chemokine (c-c motif) ligand (CCL)2), macrophage inflammatory protein-1α (MIP-1α) (CCL3) and growth regulatory protein-α (GRO-α) (chemokine (c-x-c motif) ligand (CXCL)1) are up-regulated following neural cell loss in the adult striatum and act as potent chemoattractants for SVZ-derived precursor cells in vitro. Based on these observations, the current study aimed to examine the individual effect of MCP-1, MIP-1α and GRO-α on the migration of adult SVZ-derived neural precursor cells in vivo. To address this without the confounding effects of injury-induced chemotactic cues, adeno-associated viral (AAV)2-mediated in vivo gene transfer was used to ectopically express either MCP-1, MIP-1α or GRO-α, or the control red fluorescent protein (RFP) in the normal adult rat striatum. The extent of doublecortin (Dcx)-positive cell recruitment from the SVZ into the striatal parenchyma was then determined at 4 and 8weeks following AAV2 injection. Ectopic expression either of MCP-1 or MIP-1α in the normal adult rat brain significantly increased the number of Dcx-positive cells and the extent of their migration into the striatum at both 4 and 8weeks after vector injection but did not promote either precursor cell proliferation or neural differentiation. In contrast, while over-expression of GRO-α 4weeks after vector injection induced a significant increase in Dcx-positive cell migration compared to control, this effect was reduced to control levels by 8weeks post injection. Further, direct comparison between MCP-1, MIP-1α and GRO-α at both 4 and 8weeks post vector injection indicated that GRO-α may have a reduced effect in inducing Dcx-positive cell migration

  11. NMAAP1 Expressed in BCG-Activated Macrophage Promotes M1 Macrophage Polarization.

    PubMed

    Liu, Qihui; Tian, Yuan; Zhao, Xiangfeng; Jing, Haifeng; Xie, Qi; Li, Peng; Li, Dong; Yan, Dongmei; Zhu, Xun

    2015-10-01

    Macrophages are divided into two subpopulations: classically activated macrophages (M1) and alternatively activated macrophages (M2). BCG (Bacilli Calmette-GuC)rin) activates disabled naC/ve macrophages to M1 macrophages, which act as inflammatory, microbicidal and tumoricidal cells through cell-cell contact and/or the release of soluble factors. Various transcription factors and signaling pathways are involved in the regulation of macrophage activation and polarization. We discovered that BCG-activated macrophages (BAM) expressed a new molecule, and we named it Novel Macrophage Activated Associated Protein 1 (NMAAP1). The current study found that the overexpression of NMAAP1 in macrophages results in M1 polarization with increased expression levels of M1 genes, such as inducible nitric oxide synthase (iNOS), tumor necrosis factor alpha (TNF-N1), Interleukin 6 (IL-6), Interleukin 12 (IL-12), Monocyte chemoattractant protein-1 (MCP-1) and Interleukin-1 beta (IL-1N2), and decreased expression of some M2 genes, such as Kruppel-like factor 4 (KLF4) and suppressor of cytokine signaling 1 (SOCS1), but not other M2 genes, including arginase-1 (Arg-1), Interleukin (IL-10), transforming growth factor beta (TGF-N2) and found in inflammatory zone 1 (Fizz1). Moreover, NMAAP1 overexpression in the RAW264.7 cell line increased cytotoxicity against MCA207 tumor cells, which depends on increased inflammatory cytokines rather than cell-cell contact. NMAAP1 also substantially enhanced the phagocytic ability of macrophages, which implies that NMAAP1 promoted macrophage adhesive and clearance activities. Our results indicate that NMAAP1 is an essential molecule that modulates macrophages phenotype and plays an important role in macrophage tumoricidal functions.

  12. Regulated expression of platelet factor 4 in human monocytes--role of PARs as a quantitatively important monocyte activation pathway.

    PubMed

    Schaffner, Andreas; Rhyn, Petra; Schoedon, Gabriela; Schaer, Dominik J

    2005-07-01

    Human mononuclear phagocytes have recently been shown to express constitutively and even more so, upon stimulation with bacteria, fungi, lipopolysaccharide (LPS), zymosan, or thrombin platelet basic protein (PBP). This CXC chemokine as well as platelet factor 4 (PF4), which is located genomically at a short distance from the PBP, were previously considered to be specific markers for the megakaryocyte cell lineage. Both chemokines have signaling and antimicrobial activity. In the present studies, transcriptional and expressional regulation of PF4 and related chemokines was studied in human monocytes. As shown by quantitative mRNA analysis, Western blots, radioimmunoprecipitation of cell extracts, and immunofluorescence and quantitatively with enzyme-linked immunosorbent assay, human monocytes express PF4 in the same order of magnitude as the known, regulated CXC chemokine interleukin (IL)-8. Expression of PF4 is up-regulated at the mRNA and protein level by thrombin and mediated by proteinase-activated receptors (PARs), resulting in a 32- to 128-fold higher mRNA level and leading to an up-to-sixfold increase of the peptide concentration in monocyte culture supernatants. Thrombin and the synthetic ligand of PAR-1 and PAR-2, SFLLRN, also induced comparable increases in the levels of mRNA for PBP, IL-8, regulated on activation, normal T expressed and secreted (RANTES), monocyte chemoattractant protein-1, and macrophage-inflammatory protein-1alpha and increased synthesis of these chemokines as shown by immunofluorescence or a quantitative immunobead-based method. The induction of increased mRNA levels for all chemokines by SFLLRN was unsurpassed by LPS, zymosan, interferon-gamma (IFN-gamma), tumor necrosis factor alpha (TNF-alpha), and IL-1. Activation of monocytes through PARs represents an alternate activation mechanism, independent from IFN-gamma, TNF-alpha, or other signaling pathways. PMID:15788441

  13. Hepatitis B virus X protein mediates yes-associated protein 1 upregulation in hepatocellular carcinoma

    PubMed Central

    Wu, Yuzhuo; Zhang, Junhe; Zhang, Huaihong; Zhai, Yufeng

    2016-01-01

    Hepatitis B virus (HBV) X protein (HBx) is implicated in the development of hepatocellular carcinoma (HCC). Yes-associated protein 1 (YAP) is an important proto-oncogene, which is a downstream effector molecule in the Hippo signaling pathway. The aim of the present study was to investigate the association between HBx expression in HCC samples and YAP expression in the Hippo pathway. A total of 20 pathologically confirmed HCC samples, 20 corresponding adjacent non-tumor liver tissues and 5 normal liver tissue samples were collected. The expression of HBx and YAP in the tissues was analyzed by quantitative reverse transcription-polymerase chain reaction and western blot analysis. The intensity and location of YAP expression were analyzed by immunohistochemistry. YAP mRNA and protein expression levels in HCC samples infected with HBV were significantly higher than those of normal liver tissues. Furthermore, YAP expression was positively correlated with HBx expression in HBV-positive HCC samples. Immunohistochemical staining revealed that YAP was predominantly expressed in the nuclei in HBV-positive HCC tissues. YAP expression was significantly decreased in the normal liver tissue and corresponding adjacent liver tissue when compared with the HCC tissues and by contrast to HCC tissues, YAP was predominantly located in the cytoplasm. In conclusion, these results indicate that the YAP gene is a key driver of HBx-induced liver cancer. Therefore, YAP may present a novel target in the treatment of HBV-associated HCC. PMID:27602122

  14. Hepatitis B virus X protein mediates yes-associated protein 1 upregulation in hepatocellular carcinoma

    PubMed Central

    Wu, Yuzhuo; Zhang, Junhe; Zhang, Huaihong; Zhai, Yufeng

    2016-01-01

    Hepatitis B virus (HBV) X protein (HBx) is implicated in the development of hepatocellular carcinoma (HCC). Yes-associated protein 1 (YAP) is an important proto-oncogene, which is a downstream effector molecule in the Hippo signaling pathway. The aim of the present study was to investigate the association between HBx expression in HCC samples and YAP expression in the Hippo pathway. A total of 20 pathologically confirmed HCC samples, 20 corresponding adjacent non-tumor liver tissues and 5 normal liver tissue samples were collected. The expression of HBx and YAP in the tissues was analyzed by quantitative reverse transcription-polymerase chain reaction and western blot analysis. The intensity and location of YAP expression were analyzed by immunohistochemistry. YAP mRNA and protein expression levels in HCC samples infected with HBV were significantly higher than those of normal liver tissues. Furthermore, YAP expression was positively correlated with HBx expression in HBV-positive HCC samples. Immunohistochemical staining revealed that YAP was predominantly expressed in the nuclei in HBV-positive HCC tissues. YAP expression was significantly decreased in the normal liver tissue and corresponding adjacent liver tissue when compared with the HCC tissues and by contrast to HCC tissues, YAP was predominantly located in the cytoplasm. In conclusion, these results indicate that the YAP gene is a key driver of HBx-induced liver cancer. Therefore, YAP may present a novel target in the treatment of HBV-associated HCC.

  15. Astragalus membranaceus inhibits peritoneal fibrosis via monocyte chemoattractant protein (MCP)-1 and the transforming growth factor-β1 (TGF-β1) pathway in rats submitted to peritoneal dialysis.

    PubMed

    Li, Zhenghong; Zhang, Lu; He, Weiming; Zhu, Changle; Yang, Jinsong; Sheng, Meixiao

    2014-07-22

    Inflammation and transforming growth factor-β1 (TGF-β1) contribute to the development of peritoneal fibrosis (PF), which is associated with peritoneal dialysis (PD). Astragalus membranaceus (Astragalus) has anti-inflammatory and anti-fibrotic effects in many diseases. The goal of this study was to determine the anti-fibrotic effects of Astragalus on the PF response to PD. A rat model of PD was induced using standard PD fluid, and PF was verified by HE and Masson's staining, as well as through the expression of fibroblast surface protein (FSP) and collagen III. The expression levels of monocyte chemoattractant protein (MCP)-1, F4/80 (macrophage/monocyte marker in rat), TGF-β1 and the downstream proteins phospho-SMAD 2/3 in dialyzed peritoneal tissue treated with or without Astragalus was evaluated using immunohistochemistry analysis. Overall correlations between MCP-1 and TGF-β1 staining were analyzed using both the Spearman and Pearson methods. The results showed that Astragalus could inhibit the recruitment and activation of monocytes/macrophages, thereby reducing the production of TGF-β1 in the dialyzed peritoneal membrane. PF was also significantly decreased following treatment with Astragalus. MCP-1 expression had a strong positive correlation with TGF-β1 sensitivity, suggesting that the anti-fibrotic function of Astragalus was mediated by MCP-1 and the TGF-β1 pathway. Our results indicate that Astragalus could be a useful agent against PD-induced PF.

  16. Neural regeneration protein is a novel chemoattractive and neuronal survival-promoting factor

    SciTech Connect

    Gorba, Thorsten; Bradoo, Privahini; Antonic, Ana; Marvin, Keith; Liu, Dong-Xu; Lobie, Peter E.; Reymann, Klaus G.; Gluckman, Peter D.; Sieg, Frank . E-mail: fsieg@neurenpharma.com

    2006-10-01

    Neurogenesis and neuronal migration are the prerequisites for the development of the central nervous system. We have identified a novel rodent gene encoding for a neural regeneration protein (NRP) with an activity spectrum similar to the chemokine stromal-derived factor (SDF)-1, but with much greater potency. The Nrp gene is encoded as a forward frameshift to the hypothetical alkylated DNA repair protein AlkB. The predicted protein sequence of NRP contains domains with homology to survival-promoting peptide (SPP) and the trefoil protein TFF-1. The Nrp gene is first expressed in neural stem cells and expression continues in glial lineages. Recombinant NRP and NRP-derived peptides possess biological activities including induction of neural migration and proliferation, promotion of neuronal survival, enhancement of neurite outgrowth and promotion of neuronal differentiation from neural stem cells. NRP exerts its effect on neuronal survival by phosphorylation of the ERK1/2 and Akt kinases, whereas NRP stimulation of neural migration depends solely on p44/42 MAP kinase activity. Taken together, the expression profile of Nrp, the existence in its predicted protein structure of domains with similarities to known neuroprotective and migration-inducing factors and the high potency of NRP-derived synthetic peptides acting in femtomolar concentrations suggest it to be a novel gene of relevance in cellular and developmental neurobiology.

  17. Secreted frizzled-related protein 1 regulates adipose tissue expansion and is dysregulated in severe obesity

    PubMed Central

    Lagathu, Claire; Christodoulides, Constantinos; Tan, Chong Yew; Virtue, Sam; Laudes, Matthias; Campbell, Mark; Ishikawa, Ko; Ortega, Francisco; Tinahones, Francisco J.; Fernández-Real, Jose-Manuel; Orešič, Matej; Sethi, Jaswinder K.; Vidal-Puig, Antonio

    2014-01-01

    Aim The Wnt/β-catenin signalling network offers potential targets to diagnose and uncouple obesity from its metabolic complications. Here we investigate the role of the Wnt antagonist, secreted Frizzled related protein 1 (SFRP1) in promoting adipogenesis in vitro and adipose tissue expansion in vivo. Methods We use a combination of human and murine, in vivo and in vitro models of adipogenesis, adipose tissue expansion and obesity-related metabolic syndrome to profile the involvement of SFRP1. Results Secreted Frizzled related protein 1 (SFRP1) is expressed in both murine and human mature adipocytes. The expression of SFRP1 is induced during in vitro adipogenesis and SFRP1 is preferentially expressed in mature adipocytes in human adipose tissue. Constitutive ectopic expression of SFRP1 is proadipogenic and inhibits the Wnt/β-catenin signalling pathway. In vivo endogenous levels of adipose SFRP1 are regulated in line with proadipogenic states. However, in longitudinal studies of high fat diet-fed mice we observed a dynamic temporal but biphasic regulation of endogenous SFRP1. In agreement with this profile we observed that SFRP1 expression in human tissues peaks in patients with mild obesity and gradually falls in morbidly obese subjects. Conclusions Our results suggest that SFRP1 is an endogenous modulator of Wnt/β-catenin signalling and participates in the paracrine regulation of human adipogenesis. The reduced adipose expression of SFRP1 in morbid obesity and its knock-on effect to prevent further adipose tissue expansion may contribute to the development of metabolic complications in these individuals. PMID:20514047

  18. Effect of the feeding system on the fatty acid composition, expression of the Δ9-desaturase, Peroxisome Proliferator-Activated Receptor Alpha, Gamma, and Sterol Regulatory Element Binding Protein 1 genes in the semitendinous muscle of light lambs of the Rasa Aragonesa breed

    PubMed Central

    2010-01-01

    Background Conjugated linoleic acids (CLAs) are receiving increasing attention because of their beneficial effects on human health, with milk and meat products derived from ruminants as important sources of CLA in the human diet. SCD gene is responsible for some of the variation in CLA concentration in adipose tissues, and PPARγ, PPARα and SREBP1 genes are regulator of SCD gene. The aim of this work was to evaluate the effect of the feeding system on fatty acid composition, CLA content and relative gene expression of Δ9-desaturase (SCD), Peroxisome Proliferator-Activated Receptor Gamma (PPARγ), Peroxisome Proliferator-Activated Receptor Alpha, (PPARα) and Sterol Regulatory Element Binding Protein (SREBP1) in Rasa Aragonesa light lambs in semitendinous muscle. Forty-four single-born male lambs were used to evaluate the effect of the feeding system, varying on an intensity gradient according to the use of concentrates: 1. grazing alfalfa, 2. grazing alfalfa with a supplement for lambs, 3. indoor lambs with grazing ewes and 4. drylot. Results Both grazing systems resulted in a higher concentration of vaccenic acid (VA), CLA, CLA/VA acid ratio, and a lower oleic content, oleic acid (C18:1)/stearic acid (C18:0) ratio, PUFA n-6/n-3 ratio and SCD expression compared to other diets. In addition feeding system affected the fatty acid composition and SCD expression, possibly due to CLA concentration or the PUFA n-6/n-3 ratio. Both expression of the SCD gene and the feeding system were important factors affecting CLA concentration in the animal's semitendinous muscle. PPARγ, PPARα and SREBP1 expression seemed to be unaffected by the feeding system. Although no significant results were found, PPARγ, PPARα and SREBP1 showed similar expression pattern as SCD. Moreover, the correlation results between SCD expression and PPARγ (p < 0.01), as well as SREBP1 (p < 0.01) expression, may suggest that these genes were affecting SCD expression in a different way. Conclusions

  19. MCP-1 expressed by osteoclasts stimulates osteoclastogenesis in an autocrine/paracrine manner

    SciTech Connect

    Miyamoto, Kana; Ninomiya, Ken; Sonoda, Koh-Hei; Miyauchi, Yoshiteru; Hoshi, Hiroko; Iwasaki, Ryotaro; Miyamoto, Hiroya; and others

    2009-06-05

    Monocyte chemoattractant protein-1 (MCP-1) is a chemokine that plays a critical role in the recruitment and activation of leukocytes. Here, we describe that multinuclear osteoclast formation was significantly inhibited in cells derived from MCP-1-deficient mice. MCP-1 has been implicated in the regulation of osteoclast cell-cell fusion; however defects of multinuclear osteoclast formation in the cells from mice deficient in DC-STAMP, a seven transmembrane receptor essential for osteoclast cell-cell fusion, was not rescued by recombinant MCP-1. The lack of MCP-1 in osteoclasts resulted in a down-regulation of DC-STAMP, NFATc1, and cathepsin K, all of which were highly expressed in normal osteoclasts, suggesting that osteoclast differentiation was inhibited in MCP-1-deficient cells. MCP-1 alone did not induce osteoclastogenesis, however, the inhibition of osteoclastogenesis in MCP-1-deficient cells was restored by addition of recombinant MCP-1, indicating that osteoclastogenesis was regulated in an autocrine/paracrine manner by MCP-1 under the stimulation of RANKL in osteoclasts.

  20. Expression of pro-inflammatory cytokines and the risk of intracranial aneurysm.

    PubMed

    Zhang, Hai-Feng; Zhao, Ming-Guang; Liang, Guo-Biao; Song, Zhen-Quan; Li, Zhi-Qing

    2013-12-01

    Intracranial aneurysm (IA) lingers as a potentially devastating clinical problem, in which inflammation acts as a critical contributor to the pathogenesis of this disease. Cytokines play a major role in regulating inflammation. The aim of this study was to gain insight in the inflammatory response in IA by assessing plasma cytokine profiles. Plasma levels of 10 cytokines were quantified by multiplex protein arrays in 66 patients with IA and 78 healthy controls. Results showed that plasma level of interleukin 1 beta (IL-1β) was 2.4-fold higher in patients than in controls (p < 0.05). The level of monocyte chemoattractant protein-1 (MCP-1) was 2.8-fold higher in patient than in controls (p < 0.01). The level of tumor necrosis factor-alpha (TNF-α) was 2.1-fold higher in cases than in controls (p < 0.001). When comparing the expression of cytokines in IA patients with different characteristics, cases with ruptured aneurysm revealed increased level of MCP-1 than those with unruptured aneurysm (p < 0.05), whereas cases with multiple numbers of aneurysms demonstrated higher levels of MCP-1 and TNF-α than those with single aneurysm (p < 0.05 and p < 0.05, respectively). These data indicated that IL-1β, MCP-1, and TNF-α were associated with increased risk of IA and may affect the development of this disease.

  1. Astaxanthin Inhibits Expression of Retinal Oxidative Stress and Inflammatory Mediators in Streptozotocin-Induced Diabetic Rats

    PubMed Central

    Yeh, Po-Ting; Huang, Hsin-Wei; Yang, Chung-May; Yang, Wei-Shiung; Yang, Chang-Hao

    2016-01-01

    Purpose We evaluated whether orally administered astaxanthin (AST) protects against oxidative damage in the ocular tissues of streptozotocin (STZ)-induced diabetic rats. Methods and Results Fifty 6-week-old female Wistar rats were randomly assigned to receive an injection of STZ to induce diabetes (n = 40) or to remain uninduced (n = 10). The diabetic rats were randomly selected into four groups and they were separately administered normal saline, 0.6 mg/kg AST, 3 mg/kg AST, or 0.5 mg/kg lutein daily for eight weeks. Retinal functions of each group were evaluated by electroretinography. The expression of oxidative stress and inflammatory mediators in the ocular tissues was then assessed by immunohistochemistry, western blot analysis, ELISA, RT-PCR, and electrophoretic mobility shift assay (EMSA). Retinal functions were preserved by AST and lutein in different levels. Ocular tissues from AST- and lutein-treated rats had significantly reduced levels of oxidative stress mediators (8-hydroxy-2'-deoxyguanosine, nitrotyrosine, and acrolein) and inflammatory mediators (intercellular adhesion molecule-1, monocyte chemoattractant protein-1, and fractalkine), increased levels of antioxidant enzymes (heme oxygenase-1 and peroxiredoxin), and reduced activity of the transcription factor nuclear factor-kappaB (NF-κB). Conclusion The xanthophyll carotenoids AST and lutein have neuroprotective effects and reduce ocular oxidative stress, and inflammation in the STZ diabetic rat model, which may be mediated by downregulation of NF-κB activity. PMID:26765843

  2. In Patients with Coronary Artery Disease and Type 2 Diabetes, SIRT1 Expression in Circulating Mononuclear Cells Is Associated with Levels of Inflammatory Cytokines but Not with Coronary Lesions

    PubMed Central

    Li, Yuanmin; Ni, Jing; Guo, Rong; Li, Weiming

    2016-01-01

    While SIRT1 is significantly associated with atherosclerosis and diabetic complications, its relevance to coronary lesions in patients with coronary artery disease and type 2 diabetes has not been specifically investigated. Thus, we assessed SIRT1 expression in peripheral blood mononuclear cells in these patients. We found that SIRT1 expression did not significantly correlate with syntax scores from coronary angiography (p > 0.05). Notably, plasma levels of the inflammatory cytokines tumor necrosis factor-α, monocyte chemoattractant protein-1, and high-sensitivity C-reactive protein were markedly higher in diabetic patients (p < 0.05). In addition, SIRT1 expression was negatively correlated with levels of these cytokines, as well as that of interleukin-6 (p < 0.05). In summary, the data indicate that SIRT1 expression in peripheral blood mononuclear cells is significantly correlated with inflammatory cytokines levels in patients with coronary artery disease and type 2 diabetes but not with the severity of coronary lesions. PMID:27123454

  3. Self-organization of chemoattractant waves in Dictyostelium depends on F-actin and cell–substrate adhesion

    PubMed Central

    Fukujin, Fumihito; Nakajima, Akihiko; Shimada, Nao; Sawai, Satoshi

    2016-01-01

    In the social amoeba Dictyostelium discoideum, travelling waves of extracellular cyclic adenosine monophosphate (cAMP) self-organize in cell populations and direct aggregation of individual cells to form multicellular fruiting bodies. In contrast to the large body of studies that addressed how movement of cells is determined by spatial and temporal cues encoded in the dynamic cAMP gradients, how cell mechanics affect the formation of a self-generated chemoattractant field has received less attention. Here, we show, by live cell imaging analysis, that the periodicity of the synchronized cAMP waves increases in cells treated with the actin inhibitor latrunculin. Detail analysis of the extracellular cAMP-induced transients of cytosolic cAMP (cAMP relay response) in well-isolated cells demonstrated that their amplitude and duration were markedly reduced in latrunculin-treated cells. Similarly, in cells strongly adhered to a poly-l-lysine-coated surface, the response was suppressed, and the periodicity of the population-level oscillations was markedly lengthened. Our results suggest that cortical F-actin is dispensable for the basic low amplitude relay response but essential for its full amplification and that this enhanced response is necessary to establish high-frequency signalling centres. The observed F-actin dependence may prevent aggregation centres from establishing in microenvironments that are incompatible with cell migration. PMID:27358278

  4. Self-organization of chemoattractant waves in Dictyostelium depends on F-actin and cell-substrate adhesion.

    PubMed

    Fukujin, Fumihito; Nakajima, Akihiko; Shimada, Nao; Sawai, Satoshi

    2016-06-01

    In the social amoeba Dictyostelium discoideum, travelling waves of extracellular cyclic adenosine monophosphate (cAMP) self-organize in cell populations and direct aggregation of individual cells to form multicellular fruiting bodies. In contrast to the large body of studies that addressed how movement of cells is determined by spatial and temporal cues encoded in the dynamic cAMP gradients, how cell mechanics affect the formation of a self-generated chemoattractant field has received less attention. Here, we show, by live cell imaging analysis, that the periodicity of the synchronized cAMP waves increases in cells treated with the actin inhibitor latrunculin. Detail analysis of the extracellular cAMP-induced transients of cytosolic cAMP (cAMP relay response) in well-isolated cells demonstrated that their amplitude and duration were markedly reduced in latrunculin-treated cells. Similarly, in cells strongly adhered to a poly-l-lysine-coated surface, the response was suppressed, and the periodicity of the population-level oscillations was markedly lengthened. Our results suggest that cortical F-actin is dispensable for the basic low amplitude relay response but essential for its full amplification and that this enhanced response is necessary to establish high-frequency signalling centres. The observed F-actin dependence may prevent aggregation centres from establishing in microenvironments that are incompatible with cell migration. PMID:27358278

  5. Chemotaxis can take plant-parasitic nematodes to the source of a chemo-attractant via the shortest possible routes.

    PubMed

    Reynolds, Andy M; Dutta, Tushar K; Curtis, Rosane H C; Powers, Stephen J; Gaur, Hari S; Kerry, Brian R

    2011-04-01

    It has long been recognized that chemotaxis is the primary means by which nematodes locate host plants. Nonetheless, chemotaxis has received scant attention. We show that chemotaxis is predicted to take nematodes to a source of a chemo-attractant via the shortest possible routes through the labyrinth of air-filled or water-filled channels within a soil through which the attractant diffuses. There are just two provisos: (i) all of the channels through which the attractant diffuses are accessible to the nematodes and (ii) nematodes can resolve all chemical gradients no matter how small. Previously, this remarkable consequence of chemotaxis had gone unnoticed. The predictions are supported by experimental studies of the movement patterns of the root-knot nematodes Meloidogyne incognita and Meloidogyne graminicola in modified Y-chamber olfactometers filled with Pluronic gel. By providing two routes to a source of the attractant, one long and one short, our experiments, the first to demonstrate the routes taken by nematodes to plant roots, serve to test our predictions. Our data show that nematodes take the most direct route to their preferred hosts (as predicted) but often take the longest route towards poor hosts. We hypothesize that a complex of repellent and attractant chemicals influences the interaction between nematodes and their hosts.

  6. Double-Stranded RNA-Induced Activation of Activating Protein-1 Promoter Is Differentially Regulated by the Non-structural Protein 1 of Avian Influenza A Viruses

    PubMed Central

    Zohari, Siamak; Belák, Sándor; Berg, Mikael

    2012-01-01

    Abstract Non-structural protein 1 (NS1) of influenza A viruses is a multifunctional protein that antagonizes the host immune response by interfering with several host signaling pathways. Based on putative amino acid sequences, NS1 proteins are categorized into two gene pools, allele A and allele B. Here we identified that allele A NS1 proteins of H6N8 and H4N6 are able to inhibit double-stranded RNA (dsRNA)-induced activating protein-1 (AP-1) promoter in cultured cell lines (human A549 and mink lung cells). Allele B NS1 proteins from corresponding subtypes of influenza A viruses are weak in this inhibition, despite significant levels of expression of each NS1 protein in human A549 cells. Furthermore, the capability to inhibit AP-1 promoter was mapped in the effector domain, since RNA binding domain alone lost its ability to inhibit this promoter activation. Chimeric forms of NS1 protein, composed of either RNA binding domain of allele A or B and effector domain of allele A or B, showed comparable inhibition to that of their wild-type NS1 proteins, or to the effector domain of corresponding NS1 proteins. Both alleles A and B NS1 proteins of H6N8 and H4N6 were expressed to significant levels, and were localized predominantly in the nucleus of human A549 cells. These results underscore the importance of the effector domain in inhibiting AP-1 promoter activation, and the biological function of the effector domain in stabilizing the RNA binding domain. Further, we revealed the versatile nature of NS1 in inhibiting the AP-1 transcription factor, in a manner dependent on allele type. Comprehensive studies, focusing on the molecular mechanisms behind this differential inhibition, may facilitate exploration of the zoonotic and pathogenic potential of influenza A viruses. PMID:22239235

  7. Caprine arthritis encephalitis virus dysregulates the expression of cytokines in macrophages.

    PubMed Central

    Lechner, F; Machado, J; Bertoni, G; Seow, H F; Dobbelaere, D A; Peterhans, E

    1997-01-01

    Caprine arthritis encephalitis virus (CAEV) is a lentivirus of goats that leads to chronic mononuclear infiltration of various tissues, in particular, the radiocarpal joints. Cells of the monocyte/macrophage lineage are the major host cells of CAEV in vivo. We have shown that infection of cultured goat macrophages with CAEV results in an alteration of cytokine expression in vitro. Constitutive expression of interleukin 8 (IL-8) and monocyte chemoattractant protein 1 (MCP-1) was increased in infected macrophages, whereas transforming growth factor beta1 (TGF-beta1) mRNA was down-regulated. When macrophages were infected with a CAEV clone lacking the trans-acting nuclear regulatory gene tat, IL-8 and MCP-1 were also increased. No significant differences from cells infected with the wild-type clone were observed, suggesting that Tat is not required for the increased expression of IL-8 and MCP-1 in infected macrophages. Furthermore, infection with CAEV led to an altered pattern of cytokine expression in response to lipopolysaccharide (LPS), heat-killed Listeria monocytogenes plus gamma interferon, or fixed cells of Staphylococcus aureus Cowan I. In infected macrophages, tumor necrosis factor alpha, IL-1beta, IL-6, and IL-12 p40 mRNA expression was reduced in response to all stimuli tested whereas changes in expression of granulocyte-macrophage colony-stimulating factor depended on the stimulating agent. Electrophoretic mobility shift assays demonstrated that, in contrast to effects of human immunodeficiency virus infection of macrophages, CAEV infection had no effect on the level of constitutive nuclear factor-kappaB (NF-kappaB) activity or on the level of LPS-stimulated NF-kappaB activity, suggesting that NF-kappaB is not involved in altered regulation of cytokine expression in CAEV-infected cells. In contrast, activator protein 1 (AP-1) binding activity was decreased in infected macrophages. These data show that CAEV infection may result in a dysregulation of

  8. Chemoattraction and penetration of Echinostoma trivolvis and E. caproni cercariae in the presence of Biomphalaria glabrata, Helisoma trivolvis, and Lymnaea elodes dialysate.

    PubMed

    Fried, B; Frazer, B A; Reddy, A

    1997-01-01

    A petri-dish bioassay was used to study the chemoattraction and penetration of the cercariae of Echinostoma trivolvis and E. caproni in the presence of snail dialysates from Helisoma trivolvis (Pennsylvania and Colorado strains). Biomphalaria glabrata, and Lynmaea elodes. Significant chemoattraction was seen with E. trivolvis cercariae in the presence of all snail dialysates released from nonperforated dialysis sacs with a molecular-weight exclusion of 12,000. Under the same conditions, E. caproni was significantly attracted to B. glabrata and H. trivolvis (CO strain) but not to L. elodes or H. trivolvis (PA strain). Dialysis sacs were perforated with needles to allow the release of snail substances of all molecular weights into the bioassay. Cercariae of both species were significantly attracted to all snail dialysates released from perforated sacs. Moreover, cercariae entered these sacs and penetrated the snails, and 24 h later the percentage of cysts per snail species ranged from 70% to 83% for E. trivolvis and from 73% to 93% for E. caproni. Dialysates released from intact sacs were extracted in choloroform-methanol (2:1) to obtain hydrophilic and lipophilic fractions. When these extracts were placed on agar plugs in the bioassay, the lipophilic fraction, but not the hydrophilic fraction, was mainly chemoattractive. PMID:9039703

  9. MELANOCYTIC GALECTIN-3 IS ASSOCIATED WITH TYROSINASE RELATED PROTEIN-1 AND PIGMENT BIOSYNTHESIS

    PubMed Central

    Chalupa, Allison; Koshoffer, Amy; Galan, Emily; Yu, Lan; Liu, Fu-Tong; Boissy, Raymond E.

    2014-01-01

    Galectin-3 is a family member of the carbohydrate binding proteins widely expressed by many cell types and exhibits multiple cellular functions. We demonstrate that melanocytes express galectin-3, which is predominantly localized to the cell body peripherally along the Golgi zone. Downregulation of galectin-3 in human melanocytes using shRNA technology resulted in reduction of both melanin synthesis and expression/activity of Tyrp-1. In the cell body, galectin-3 co-localizes with melanosome destined cargo, specifically tyrosinase and tyrosinase-related protein-1. We studied melanocytes cultured from patients with forms of Hermansky-Pudlak syndrome containing defects in trafficking steps governed by BLOC-2 (HPS5), BLOC-3 (HPS1) and adaptin-3 (HPS2). We found galectin-3 expression mimicked the defective expression of the tyrosinase cargo in dendrites of HPS-5 melanocytes, but was not altered in HPS1 or HPS2 melanocytes. In addition, galectin-3 co-localized predominantly with the HPS-5 component of BLOC-2 in normal human melanocytes. These data indicate that galectin-3 is a regulatory component in melanin synthesis affecting the expression of Tyrp-1. PMID:25054620

  10. A Critical Role for Cysteine 57 in the Biological Functions of Selenium Binding Protein-1.

    PubMed

    Ying, Qi; Ansong, Emmanuel; Diamond, Alan M; Yang, Wancai

    2015-11-18

    The concentration of selenium-binding protein1 (SBP1) is often lower in tumors than in the corresponding tissue and lower levels have been associated with poor clinical outcomes. SBP1 binds tightly selenium although what role selenium plays in its biological functions remains unknown. Previous studies indicated that cysteine 57 is the most likely candidate amino acid for selenium binding. In order to investigate the role of cysteine 57 in SBP1, this amino acid was altered to a glycine and the mutated protein was expressed in human cancer cells. The SBP1 half-life, as well as the cellular response to selenite cytotoxicity, was altered by this change. The ectopic expression of SBP1(GLY) also caused mitochondrial damage in HCT116 cells. Taken together, these results indicated that cysteine 57 is a critical determinant of SBP1 function and may play a significant role in mitochondrial function.

  11. A Critical Role for Cysteine 57 in the Biological Functions of Selenium Binding Protein-1.

    PubMed

    Ying, Qi; Ansong, Emmanuel; Diamond, Alan M; Yang, Wancai

    2015-01-01

    The concentration of selenium-binding protein1 (SBP1) is often lower in tumors than in the corresponding tissue and lower levels have been associated with poor clinical outcomes. SBP1 binds tightly selenium although what role selenium plays in its biological functions remains unknown. Previous studies indicated that cysteine 57 is the most likely candidate amino acid for selenium binding. In order to investigate the role of cysteine 57 in SBP1, this amino acid was altered to a glycine and the mutated protein was expressed in human cancer cells. The SBP1 half-life, as well as the cellular response to selenite cytotoxicity, was altered by this change. The ectopic expression of SBP1(GLY) also caused mitochondrial damage in HCT116 cells. Taken together, these results indicated that cysteine 57 is a critical determinant of SBP1 function and may play a significant role in mitochondrial function. PMID:26593911

  12. Tubulin polymerization promoting protein 1 (TPPP1): A DNA-damage induced microtubule regulatory gene.

    PubMed

    Schofield, Alice; Bernard, Ora

    2013-11-01

    The eukaryotic cell cycle relies heavily on the mechanical forces vested by the dynamic rearrangement of the microtubule (MT) network. Tubulin Polymerization promoting Protein 1 (TPPP1) alters MT dynamics by driving MT polymerization as well as stabilization, via increasing MT acetylation. It increases MT rigidity, which results in reduced cell proliferation through downregulation of G1/S-phase and mitosis to G1-phase cell cycle transitioning. In this communication, we provide further evidence that TPPP1 may be an important regulator of genomic homeostasis. Our preliminary data show that long-term TPPP1 overexpression reduces cell viability via induction of apoptotic cell death pathways. Moreover, induction of DNA-damage results in increased TPPP1 expression, which is inhibited in the absence of expression of the tumor suppressor p53.

  13. Far upstream element-binding protein 1 is a prognostic biomarker and promotes nasopharyngeal carcinoma progression

    PubMed Central

    Liu, Z-H; Hu, J-L; Liang, J-Z; Zhou, A-J; Li, M-Z; Yan, S-M; Zhang, X; Gao, S; Chen, L; Zhong, Q; Zeng, M-S

    2015-01-01

    Nasopharyngeal carcinoma (NPC) is a malignant epithelial tumor with tremendous invasion and metastasis capacities, and it has a high incidence in southeast Asia and southern China. Previous studies identified that far upstream element-binding protein 1 (FBP1), a transcriptional regulator of c-Myc that is one of the most frequently aberrantly expressed oncogenes in various human cancers, including NPC, is an important biomarker for many cancers. Our study aimed to investigate the expression and function of FBP1 in human NPC. Quantitative real-time RT-PCR (qRT-PCR), western blot and immunohistochemical staining (IHC) were performed in NPC cells and biopsies. Furthermore, the effect of FBP1 knockdown on cell proliferation, colony formation, side population tests and tumorigenesis in nude mice were measured by MTT, clonogenicity analysis, flow cytometry and a xenograft model, respectively. The results showed that the mRNA and protein levels of FBP1, which are positively correlated with c-Myc expression, were substantially higher in NPC than that in nasopharyngeal epithelial cells. IHC revealed that the patients with high FBP1 expression had a significantly poorer prognosis compared with the patients with low expression (P=0.020). In univariate analysis, high FBP1 and c-Myc expression predicted poorer overall survival (OS) and poorer progression-free survival. Multivariate analysis indicated that high FBP1 and c-Myc expression were independent prognostic markers. Knockdown of FBP1 reduced cell proliferation, clonogenicity and the ratio of side populations, as well as tumorigenesis in nude mice. These data indicate that FBP1 expression, which is closely correlated with c-Myc expression, is an independent prognostic factor and promotes NPC progression. Our results suggest that FBP1 can not only serve as a useful prognostic biomarker for NPC but also as a potential therapeutic target for NPC patients. PMID:26469968

  14. Phytoestrogens mediated anti-inflammatory effect through suppression of IRF-1 and pSTAT1 expressions in lipopolysaccharide-activated microglia.

    PubMed

    Jantaratnotai, Nattinee; Utaisincharoen, Pongsak; Sanvarinda, Pimtip; Thampithak, Anusorn; Sanvarinda, Yupin

    2013-10-01

    Microglial activation has been implicated in various neurological disorders, including Alzheimer's disease, Parkinson's disease, multiple sclerosis, and HIV encephalopathy. Phytoestrogens have been shown to be neuroprotective in neurotoxicity models; however, their effect on microglia has not been well established. In the current study, we report that the soy phytoestrogens, genistein, daidzein, and coumestrol, decreased nitric oxide (NO) production induced by lipopolysaccharide (LPS) in the rat microglial cell line (HAPI). The levels of inducible NO synthase (iNOS) mRNA and protein expression were also reduced. Transcription factors known to govern iNOS expression including interferon regulatory factor-1 (IRF-1) and phosphorylated STAT1 were down regulated. These observations explain, at least in part, the inhibitory effect of phytoestrogens on NO production. The levels of monocyte chemoattractant protein-1 and interleukin-6 mRNA, proinflammatory chemokine and cytokine associated with various neurological disorders, were also reduced following LPS stimulation when HAPI cells were pretreated with phytoestrogens. Hence, genistein, daidzein, and coumestrol could serve as anti-inflammatory agents and may have beneficial effects in the treatment of neurodegenerative diseases.

  15. Differential flux of macrophage inflammatory protein-2 and cytokine-induced neutrophil chemoattractant from the lung after intrapulmonary delivery.

    PubMed

    Zamjahn, John B; Quinton, Lee J; Mack, Justin C; Frevert, Charles W; Nelson, Steve; Bagby, Gregory J

    2011-10-01

    Previously we showed that cytokine-induced neutrophil chemoattractant (CINC), but not macrophage inflammatory protein-2 (MIP-2), is detected in plasma after intratracheal challenge with LPS or the particular chemokines. To further understand the differences between CINC and MIP-2 flux from the lung, we attempted to detect the two chemokines in isolated erythrocytes and leukocytes in rats after intratracheal LPS challenge. In response to intratracheal LPS, we found both CINC and MIP-2 in isolated erythrocytes and leukocytes, suggesting that MIP-2 produced in the LPS-challenged lung entered the circulation like CINC. To assess the relative flux of CINC and MIP-2 from the intra-alveolar compartment into the blood, experiments were performed in rats implanted with vascular catheters in which both chemokines were either injected intratracheally (5 μg) or infused intravenously (20 ng/min) and subsequently measured in plasma or with the cellular elements. Both chemokines appeared in the blood following intratracheal injection, with CINC detected in plasma and cells but MIP-2 only detected in the cellular fraction of blood. Infusion of both chemokines allowed detection of MIP-2 and CINC in plasma and with the cellular elements, which allowed us to calculate clearance for each chemokine and to assess CINC and MIP-2 rates of appearance (Ra) following intratracheal injection. On the basis of plasma and whole blood clearance, CINC Ra was more than sevenfold and fourfold higher, respectively, than MIP-2 Ra. This analysis indicates that differences exist in the rate of flux of CINC and MIP-2 across the epithelial/endothelial barrier of the lung, despite similar molecular size.

  16. Angiopoietin-like protein 1 suppresses SLUG to inhibit cancer cell motility.

    PubMed

    Kuo, Tsang-Chih; Tan, Ching-Ting; Chang, Yi-Wen; Hong, Chih-Chen; Lee, Wei-Jiunn; Chen, Min-Wei; Jeng, Yung-Ming; Chiou, Jean; Yu, Pei; Chen, Pai-Sheng; Wang, Ming-Yang; Hsiao, Michael; Su, Jen-Liang; Kuo, Min-Liang

    2013-03-01

    Angiopoietin-like protein 1 (ANGPTL1) is a potent regulator of angiogenesis. Growing evidence suggests that ANGPTL family proteins not only target endothelial cells but also affect tumor cell behavior. In a screen of 102 patients with lung cancer, we found that ANGPTL1 expression was inversely correlated with invasion, lymph node metastasis, and poor clinical outcomes. ANGPTL1 suppressed the migratory, invasive, and metastatic capabilities of lung and breast cancer cell lines in vitro and reduced metastasis in mice injected with cancer cell lines overexpressing ANGPTL1. Ectopic expression of ANGPTL1 suppressed the epithelial-to-mesenchymal transition (EMT) by reducing the expression of the zinc-finger protein SLUG. A microRNA screen revealed that ANGPTL1 suppressed SLUG by inducing expression of miR-630 in an integrin α(1)β(1)/FAK/ERK/SP1 pathway-dependent manner. These results demonstrate that ANGPTL1 represses lung cancer cell motility by abrogating the expression of the EMT mediator SLUG.

  17. Express

    Integrated Risk Information System (IRIS)

    Express ; CASRN 101200 - 48 - 0 Human health assessment information on a chemical substance is included in the IRIS database only after a comprehensive review of toxicity data , as outlined in the IRIS assessment development process . Sections I ( Health Hazard Assessments for Noncarcinogenic Effect

  18. Nonstructural Protein 1 of Influenza A Virus Interacts with Human Guanylate-Binding Protein 1 to Antagonize Antiviral Activity

    PubMed Central

    Yan, Wenjun; Wei, Jianchao; Shao, Donghua; Deng, Xufang; Wang, Shaohui; Li, Beibei; Tong, Guangzhi; Ma, Zhiyong

    2013-01-01

    Human guanylate-binding protein 1 (hGBP1) is an interferon-inducible protein involved in the host immune response against viral infection. In response to infection by influenza A virus (IAV), hGBP1 transcript and protein were significantly upregulated. Overexpression of hGBP1 inhibited IAV replication in a dose-dependent manner in vitro. The lysine residue at position 51 (K51) of hGBP1 was essential for inhibition of IAV replication. Mutation of K51 resulted in an hGBP1 that was unable to inhibit IAV replication. The viral nonstructural protein 1 (NS1) was found to interact directly with hGBP1. K51 of hGBP1 and a region between residues 123 and 144 in NS1 were demonstrated to be essential for the interaction between NS1 and hGBP1. Binding of NS1 to hGBP1 resulted in a significant reduction in both GTPase activity and the anti-IAV activity of hGBP1. These findings indicated that hGBP1 contributed to the host immune response against IAV replication and that hGBP1-mediated antiviral activity was antagonized by NS1 via binding to hGBP1. PMID:23405236

  19. Nonstructural protein 1 of influenza A virus interacts with human guanylate-binding protein 1 to antagonize antiviral activity.

    PubMed

    Zhu, Zixiang; Shi, Zixue; Yan, Wenjun; Wei, Jianchao; Shao, Donghua; Deng, Xufang; Wang, Shaohui; Li, Beibei; Tong, Guangzhi; Ma, Zhiyong

    2013-01-01

    Human guanylate-binding protein 1 (hGBP1) is an interferon-inducible protein involved in the host immune response against viral infection. In response to infection by influenza A virus (IAV), hGBP1 transcript and protein were significantly upregulated. Overexpression of hGBP1 inhibited IAV replication in a dose-dependent manner in vitro. The lysine residue at position 51 (K51) of hGBP1 was essential for inhibition of IAV replication. Mutation of K51 resulted in an hGBP1 that was unable to inhibit IAV replication. The viral nonstructural protein 1 (NS1) was found to interact directly with hGBP1. K51 of hGBP1 and a region between residues 123 and 144 in NS1 were demonstrated to be essential for the interaction between NS1 and hGBP1. Binding of NS1 to hGBP1 resulted in a significant reduction in both GTPase activity and the anti-IAV activity of hGBP1. These findings indicated that hGBP1 contributed to the host immune response against IAV replication and that hGBP1-mediated antiviral activity was antagonized by NS1 via binding to hGBP1.

  20. Inhibitory Effect of 3-(4-Hydroxyphenyl)-1-(thiophen-2-yl) prop-2-en-1-one, a Chalcone Derivative on MCP-1 Expression in Macrophages via Inhibition of ROS and Akt Signaling

    PubMed Central

    Kim, Mi Jin; Kadayat, Taraman; Um, Yeon Ji; Jeong, Tae Cheon; Lee, Eung-Seok; Park, Pil-Hoon

    2015-01-01

    Chalcones (1,3-diaryl-2-propen-1-ones), a subfamily of flavonoid, are widely known to possess potent anti-inflammatory and anti-oxidant properties. In this study, we investigated the effect of 3-(4-Hydroxyphenyl)-1-(thio3-(4-Hydroxyphenyl phen-2-yl)prop-2-en-1-one (TI-I-175), a synthetic chalcone derivative, on endotoxin-induced expression of monocyte chemoattractant protein-1 (MCP-1), one of the key chemokines that regulates migration and infiltration of immune cells, and its potential mechanisms. TII-175 potently inhibited MCP-1 mRNA expression stimulated by lipopolysaccharide (LPS) in RAW 264.7 macrophages without significant effect on cell viability. Treatment of cells with TI-I-175 markedly prevented LPS-induced transcriptional activation of activator protein-1 (AP-1) as measured by luciferase reporter assay, while nuclear factor-κB (NF-κB) activity was not inhibited by TI-I-175, implying that TI-I-175 suppressed MCP-1 expression probably via regulation of AP-1. In addition, TI-I-175 treatment significantly inhibited LPS-induced Akt phosphorylation and led to a significant decrease in reactive oxygen species (ROS) production by LPS, which act as up-stream signaling events required for AP-1 activation in RAW 264.7 macrophages. Taken together, these results indicate that TI-I-175 suppresses MCP-1 gene expression in LPS-stimulated RAW 264.7 macrophages via suppression of ROS production and Akt activation. PMID:25767679

  1. Obesity and inflammation: reduced cytokine expression due to resveratrol in a human in vitro model of inflamed adipose tissue

    PubMed Central

    Zagotta, Ivana; Dimova, Elitsa Y.; Debatin, Klaus-Michael; Wabitsch, Martin; Kietzmann, Thomas; Fischer-Posovszky, Pamela

    2015-01-01

    Obesity is associated with an inflammatory status and linked with a number of pathophysiological complications among them cardiovascular disease, type 2 diabetes mellitus, or the metabolic syndrome. Resveratrol was proposed to improve obesity-related inflammatory problems, but the effect of resveratrol on cytokine expression in obesity is not completely understood. In this study, we used an in vitro model of human adipose tissue inflammation to examine the effects of resveratrol on the production of the inflammatory cytokines interleukin 6 (IL-6), IL-8, and monocyte chemoattractant protein 1 (MCP-1). We found that resveratrol reduced IL-6, IL-8, and MCP-1 levels in a concentration-dependent manner in adipocytes under inflammatory conditions. Further experiments showed that the action of resveratrol was mainly due to its NFκB inhibitory potential. Thus, our data support the concept that resveratrol can alleviate obesity-induced up-regulation of inflammatory cytokines providing a new insight toward novel treatment options in obesity. PMID:25926797

  2. Localization of Vascular Adhesion Protein-1 (VAP-1) in the Human Eye

    PubMed Central

    Almulki, Lama; Noda, Kousuke; Nakao, Shintaro; Hisatomi, Toshio; Thomas, Kennard L.; Hafezi-Moghadam, Ali

    2009-01-01

    Recently we showed a critical role for Vascular Adhesion Protein-1 (VAP-1) in rodents during acute ocular inflammation, angiogenesis, and diabetic retinal leukostasis. However, the expression of VAP-1 in the human eye is unknown. VAP-1 localization was investigated by immunohistochemistry. Five μm thick sections were generated from human ocular tissues embedded in paraffin. Sections were incubated overnight with primary mAbs against VAP-1 (5μg/ml), smooth muscle actin (1μg/ml), CD31 or isotype-matched IgG at 4°C. Subsequently, a secondary mAb was used for 30min at room temperature, followed by Dako Envision + HRP (AEC) System for signal detection. The stained sections were examined using light microscopy and the signal intensity was quantified by two masked evaluators and graded into 4 discrete categories. In all examined ocular tissues, VAP-1 staining was confined to the vasculature. VAP-1 labeling showed the highest intensity in both arteries and veins of neuronal tissues; retina, and optic nerve, and the lowest intensity in the iris vasculature (p<0.05). Scleral and choroidal vessels showed moderate staining for VAP-1. VAP-1 intensity was significantly higher in the arteries compared to veins (p<0.05). Furthermore, VAP-1 staining in arteries co-localized with both CD31 and smooth muscle actin (sm-actin) staining, suggesting expression of VAP-1 in endothelial cells, smooth muscle cells or potentially pericytes. In conclusion, Immunohistochemistry reveals constitutive expression of VAP-1 in human ocular tissues. VAP-1 expression is exclusive to the vasculature with arteries showing significantly higher expression than veins. Furthermore, VAP-1 expression in the ocular vasculature is heterogeneous, with the vessels of the optic nerve and the retina showing highest expressions. These results characterize VAP-1 expression in human ocular tissues. PMID:19761765

  3. Small Molecule Inhibitors Targeting Activator Protein 1 (AP-1)

    PubMed Central

    2015-01-01

    Activator protein 1 (AP-1) is a pivotal transcription factor that regulates a wide range of cellular processes including proliferation, apoptosis, differentiation, survival, cell migration, and transformation. Accumulating evidence supports that AP-1 plays an important role in several severe disorders including cancer, fibrosis, and organ injury, as well as inflammatory disorders such as asthma, psoriasis, and rheumatoid arthritis. AP-1 has emerged as an actively pursued drug discovery target over the past decade. Excitingly, a selective AP-1 inhibitor T-5224 (51) has been investigated in phase II human clinical trials. Nevertheless, no effective AP-1 inhibitors have yet been approved for clinical use. Despite significant advances achieved in understanding AP-1 biology and function, as well as the identification of small molecules modulating AP-1 associated signaling pathways, medicinal chemistry efforts remain an urgent need to yield selective and efficacious AP-1 inhibitors as a viable therapeutic strategy for human diseases. PMID:24831826

  4. Structural studies of human glioma pathogenesis-related protein 1

    SciTech Connect

    Asojo, Oluwatoyin A.; Koski, Raymond A.; Bonafé, Nathalie

    2011-10-01

    Structural analysis of a truncated soluble domain of human glioma pathogenesis-related protein 1, a membrane protein implicated in the proliferation of aggressive brain cancer, is presented. Human glioma pathogenesis-related protein 1 (GLIPR1) is a membrane protein that is highly upregulated in brain cancers but is barely detectable in normal brain tissue. GLIPR1 is composed of a signal peptide that directs its secretion, a conserved cysteine-rich CAP (cysteine-rich secretory proteins, antigen 5 and pathogenesis-related 1 proteins) domain and a transmembrane domain. GLIPR1 is currently being investigated as a candidate for prostate cancer gene therapy and for glioblastoma targeted therapy. Crystal structures of a truncated soluble domain of the human GLIPR1 protein (sGLIPR1) solved by molecular replacement using a truncated polyalanine search model of the CAP domain of stecrisp, a snake-venom cysteine-rich secretory protein (CRISP), are presented. The correct molecular-replacement solution could only be obtained by removing all loops from the search model. The native structure was refined to 1.85 Å resolution and that of a Zn{sup 2+} complex was refined to 2.2 Å resolution. The latter structure revealed that the putative binding cavity coordinates Zn{sup 2+} similarly to snake-venom CRISPs, which are involved in Zn{sup 2+}-dependent mechanisms of inflammatory modulation. Both sGLIPR1 structures have extensive flexible loop/turn regions and unique charge distributions that were not observed in any of the previously reported CAP protein structures. A model is also proposed for the structure of full-length membrane-bound GLIPR1.

  5. Functional Expression of TWEAK and the Receptor Fn14 in Human Malignant Ovarian Tumors: Possible Implication for Ovarian Tumor Intervention

    PubMed Central

    Zhu, Jing; Ding, Chuanwei; Xu, Hai-bo; Qiu, Lihua; Di, Wen

    2013-01-01

    The aim of this current study was to investigate the expression of the tumor necrosis factor (TNF)-like weak inducer of apoptosis (TWEAK) and its receptor fibroblast growth factor-inducible 14 (Fn14) in human malignant ovarian tumors, and test TWEAK’s potential role on tumor progression in cell models in-vitro. Using immunohistochemistry (IHC), we found that TWEAK and its receptor Fn14 were expressed in human malignant ovarian tumors, but not in normal ovarian tissues or in borderline/benign epithelial ovarian tumors. High levels of TWEAK expression was detected in the majority of malignant tumors (36 out of 41, 87.80%). Similarly, 35 out of 41 (85.37%) malignant ovarian tumors were Fn14 positive. In these malignant ovarian tumors, however, TWEAK/Fn14 expression was not corrected with patients’ clinical subtype/stages or pathological features. In vitro, we demonstrated that TWEAK only inhibited ovarian cancer HO-8910PM cell proliferation in combination with tumor necrosis factor-α (TNF-α), whereas either TWEAK or TNF-α alone didn’t affect HO-8910PM cell growth. TWEAK promoted TNF-α production in cultured THP-1 macrophages. Meanwhile, conditioned media from TWEAK-activated macrophages inhibited cultured HO-8910PM cell proliferation and invasion. Further, TWEAK increased monocyte chemoattractant protein-1 (MCP-1) production in cultured HO-8910PM cells to possibly recruit macrophages. Our results suggest that TWEAK/Fn14, by activating macrophages, could be ovarian tumor suppressors. The unique expression of TWEAK/Fn14 in malignant tumors indicates that it might be detected as a malignant ovarian tumor marker. PMID:23469193

  6. Epilepsy, Behavioral Abnormalities, and Physiological Comorbidities in Syntaxin-Binding Protein 1 (STXBP1) Mutant Zebrafish

    PubMed Central

    Grone, Brian P.; Marchese, Maria; Hamling, Kyla R.; Kumar, Maneesh G.; Krasniak, Christopher S.; Sicca, Federico; Santorelli, Filippo M.; Patel, Manisha; Baraban, Scott C.

    2016-01-01

    Mutations in the synaptic machinery gene syntaxin-binding protein 1, STXBP1 (also known as MUNC18-1), are linked to childhood epilepsies and other neurodevelopmental disorders. Zebrafish STXBP1 homologs (stxbp1a and stxbp1b) have highly conserved sequence and are prominently expressed in the larval zebrafish brain. To understand the functions of stxbp1a and stxbp1b, we generated loss-of-function mutations using CRISPR/Cas9 gene editing and studied brain electrical activity, behavior, development, heart physiology, metabolism, and survival in larval zebrafish. Homozygous stxbp1a mutants exhibited a profound lack of movement, low electrical brain activity, low heart rate, decreased glucose and mitochondrial metabolism, and early fatality compared to controls. On the other hand, homozygous stxbp1b mutants had spontaneous electrographic seizures, and reduced locomotor activity response to a movement-inducing “dark-flash” visual stimulus, despite showing normal metabolism, heart rate, survival, and baseline locomotor activity. Our findings in these newly generated mutant lines of zebrafish suggest that zebrafish recapitulate clinical phenotypes associated with human syntaxin-binding protein 1 mutations. PMID:26963117

  7. The investigation on the role of mitochondrial fusion protein 1 in the development of myopia

    PubMed Central

    Cai, Yun-Lin; Zou, Yun-Chun; Lei, Jia-Hong; Zeng, Guan-Peng; Wang, Ying

    2016-01-01

    Purpose: The aim of this study is to preliminarily investigate the expression of mitochondrial fusion protein 1 (MFN1) in a lens-induced animal myopia (LIM) model and to explore the relationship between MFN1 and the visual development. Materials and Methods: MFN1 gene expression in guinea pigs was examined during the development of minus LIM, 15 tri-colored guinea pigs were obtained, and one eye of each pig was randomly selected and treated with −7.00D lenses. Ocular refraction and axial length were collected before intervention and 1, 2, and 3 weeks after intervention. After the refraction and axial length measurements at 1, 2, and 3 weeks of lens intervention, five guinea pigs were randomly selected. MFN1 expression in the retina of both eyes was tested by immunohistochemistry technique. Results: MFN1-positive cells could be observed in the retina of both eyes. The positive cells in the LIM eyes were staining deeper, and much more positive cells could be observed. Furthermore, MFN1-positive expression could be seen mainly in ganglion cells after 1 week of minus lens intervention, and with time extension, more and more positive cells appeared in the rod-cone cell and bipolar cell layer, and this phenomenon could not be found in the normal control eyes. Conclusion: This study suggested that MFN1 might be correlated to the development of myopia. PMID:27609161

  8. Vascular adhesion protein-1 promotes liver inflammation and drives hepatic fibrosis

    PubMed Central

    Weston, Chris J.; Shepherd, Emma L.; Claridge, Lee C.; Rantakari, Pia; Curbishley, Stuart M.; Tomlinson, Jeremy W.; Hubscher, Stefan G.; Reynolds, Gary M.; Aalto, Kristiina; Anstee, Quentin M.; Jalkanen, Sirpa; Salmi, Marko; Smith, David J.; Day, Christopher P.; Adams, David H.

    2014-01-01

    Nonalcoholic fatty liver disease (NAFLD) encompasses a range of manifestations, including steatosis and cirrhosis. Progressive disease is characterized by hepatic leukocyte accumulation in the form of steatohepatitis. The adhesion molecule vascular adhesion protein-1 (VAP-1) is a membrane-bound amine oxidase that promotes leukocyte recruitment to the liver, and the soluble form (sVAP-1) accounts for most circulating monoamine oxidase activity, has insulin-like effects, and can initiate oxidative stress. Here, we determined that hepatic VAP-1 expression is increased in patients with chronic liver disease and that serum sVAP-1 levels are elevated in patients with NAFLD compared with those in control individuals. In 4 murine hepatic injury models, an absence or blockade of functional VAP-1 reduced inflammatory cell recruitment to the liver and attenuated fibrosis. Moreover, disease was reduced in animals expressing a catalytically inactive form of VAP-1, implicating enzyme activity in the disease pathogenesis. Within the liver, hepatic stromal cells expressed functional VAP-1, and evaluation of cultured cells revealed that sVAP-1 promotes leukocyte migration through catalytic generation of ROS, which depended on VAP-1 enzyme activity. VAP-1 enhanced stromal cell spreading and wound closure and modulated expression of profibrotic genes. Together, these results link the amine oxidase activity of VAP-1 with hepatic inflammation and fibrosis and suggest that targeting VAP-1 has therapeutic potential for NAFLD and other chronic fibrotic liver diseases. PMID:25562318

  9. Phytanic acid, a novel activator of uncoupling protein-1 gene transcription and brown adipocyte differentiation.

    PubMed Central

    Schlüter, Agatha; Barberá, Maria José; Iglesias, Roser; Giralt, Marta; Villarroya, Francesc

    2002-01-01

    Phytanic acid (3,7,11,15-tetramethylhexadecanoic acid) is a phytol-derived branched-chain fatty acid present in dietary products. Phytanic acid increased uncoupling protein-1 (UCP1) mRNA expression in brown adipocytes differentiated in culture. Phytanic acid induced the expression of the UCP1 gene promoter, which was enhanced by co-transfection with a retinoid X receptor (RXR) expression vector but not with other expression vectors driving peroxisome proliferator-activated receptor (PPAR)alpha, PPARgamma or a form of RXR devoid of ligand-dependent sensitivity. The effect of phytanic acid on the UCP1 gene required the 5' enhancer region of the gene and the effects of phytanic acid were mediated in an additive manner by three binding sites for RXR. Moreover, phytanic acid activates brown adipocyte differentiation: long-term exposure of brown preadipocytes to phytanic acid promoted the acquisition of the brown adipocyte morphology and caused a co-ordinate induction of the mRNAs for gene markers of brown adipocyte differentiation, such as UCP1, adipocyte lipid-binding protein aP2, lipoprotein lipase, the glucose transporter GLUT4 or subunit II of cytochrome c oxidase. In conclusion, phytanic acid is a natural product of phytol metabolism that activates brown adipocyte thermogenic function. It constitutes a potential nutritional signal linking dietary status to adaptive thermogenesis. PMID:11829740

  10. The multidrug resistance protein 1: a functionally important activation marker for murine Th1 cells.

    PubMed

    Prechtl, S; Roellinghoff, M; Scheper, R; Cole, S P; Deeley, R G; Lohoff, M

    2000-01-15

    Previously, we described the expression of an energy-dependent pump in resting murine Th2 (but not resting Th1) cells which extruded the fluorescent dye Fluo-3. After stimulation with Ag and APCs, Th1 cells also expressed this pump. Furthermore, expression of the murine multidrug resistance protein 1 (mrp1) correlated with the presence of the pump. In this study, we report that Fluo-3 is indeed transported by murine mrp1 or its human ortholog MRP1, as revealed by transfection of HEK 293 cells with mrp1 or MRP1 cDNA. Like antigenic activation, IL-2 dose-dependently enhanced the Fluo-3-extruding activity in murine Th1 cells. Although TNF-alpha and IL-12 by themselves only weakly enhanced Fluo-3 extrusion, each of them did so in strong synergism with IL-2. An Ab directed against mrp1 was used to quantify the expression of mrp1 protein in T cells at the single-cell level. Like the Fluo-3 pump, mrp1 protein expression was enhanced by IL-2. Immunohistochemical studies using confocal laser microscopy indicated that mrp1 is localized mainly at the plasma membrane. In addition, protein expression of mrp1 was induced in Vbeta8+CD4+ T cells 12 h after in vivo application of Staphylococcal enterotoxin B. Finally, mrp1 was functionally relevant during the activation process of Th1 cells, because T cell activation could be suppressed by exposure of cells to the mrp1 inhibitor MK571. Thus, we present mrp1 as a novel, functionally important activation marker for Th1 cells and short-term in vivo activated CD4+ T cells, whereas its expression seems to be constitutive in Th2 cells.

  11. G-protein coupled receptor-associated sorting protein 1 (GASP-1), a ubiquitous tumor marker.

    PubMed

    Zheng, Xiaoyi; Chang, Frank; Zhang, Xinmin; Rothman, Vicki L; Tuszynski, George P

    2012-08-01

    Using an innovative "2-D high performance liquid electrophoresis" (2-D HPLE) technology we identified that a specific fragment of G-protein coupled receptor-associated sorting protein 1 (GASP-1) was present in the sera of breast cancer patients and was over-expressed in early and late stage breast tumors (Tuszynski, G.P. et al., 2011). In this study we further investigated the significance of GASP-1 as a tumor marker by investigating the expression GASP-1 in different kinds of tumors as well as in the sera of patients with various cancers. Over expression of GASP-1 was detected in brain, pancreatic, and breast cancers as compared to their respective normal tissues as assessed by immunohistochemical staining of tissue arrays using a "peptide specific" GASP-1 antibody. We found that across these cancers, GASP-1 was expressed approximately 10 fold more in the cancer as compared to normal tissue. The increase in GASP-1 expression was also seen in hyperplastic and inflammatory lesions of breast and pancreatic cancers as compared to normal tissue. GASP-1 was primarily expressed in the tumor epithelium of the epithelial-derived cancers and in the transformed glial cells of the brain tumors. Using a sensitive "competitive ELISA" for GASP-1, we found that sera from patients with brain, liver, breast and lung cancers expressed 4-7 fold more GASP-1 peptide than sera from normal healthy individuals. These studies identify GASP-1 as a potential new serum and tumor biomarker for several cancers and suggest that GASP-1 may be a novel target for development of cancer therapeutics. PMID:22483848

  12. Prostate Androgen-Regulated Mucin-Like protein 1: A Novel Regulator of Progesterone Metabolism

    PubMed Central

    Park, Ji Yeon; Jang, Hyein; Curry, Thomas E.; Sakamoto, Aiko

    2013-01-01

    The LH surge reprograms preovulatory follicular cells to become terminally differentiated luteal cells which produce high levels of progesterone and become resistant to apoptosis. PARM1 (prostate androgen regulated mucin-like protein 1) has been implicated in cell differentiation and cell survival in nonovarian cells, but little is known about PARM1 in the ovary. This study demonstrated that the LH surge induced a dramatic increase in Parm1 expression in periovulatory follicles and newly forming CL in both cycling and immature rat models. We further demonstrated that hCG increases Parm1 expression in granulosa cell cultures. The in vitro up-regulation of Parm1 expression was mediated by hCG-activated multiple signaling pathways and transcriptional activation of this gene. Parm1 knockdown increased the viability of cultured granulosa cells but resulted in a decrease in progesterone levels. The inhibitory effect of Parm1 silencing on progesterone was reversed by adenoviral mediated add-back expression of Parm1. Parm1 silencing had little effect on the expression of genes involved in progesterone biosynthesis and metabolism such as Scarb1, Ldlr, Vldlr, Scp2, Star, Cyp11a1, Hsd3b, and Srd5a1, while decreasing the expression of Akr1c3. Analyses of culture media steroid levels revealed that Parm1 knockdown had no effect on pregnenolone levels, while resulting in time-dependent decreases in progesterone and 20α-dihydroprogesterone and accelerated accumulation of 5α-pregnanediol. This study revealed that the up-regulation of Parm1 expression promotes progesterone and 20α-dihydroprogesterone accumulation in luteinizing granulosa cells by inhibiting progesterone catabolism to 5α-pregnanediol. PARM1 contributes to ovulation and/or luteal function by acting as a novel regulator of progesterone metabolism. PMID:24085821

  13. Restoration of Brain Acid Soluble Protein 1 Inhibits Proliferation and Migration of Thyroid Cancer Cells

    PubMed Central

    Guo, Run-Sheng; Yu, Yue; Chen, Jun; Chen, Yue-Yu; Shen, Na; Qiu, Ming

    2016-01-01

    Background: Brain acid soluble protein 1 (BASP1) is identified as a novel potential tumor suppressor in several cancers. However, its role in thyroid cancer has not been investigated yet. In the present study, the antitumor activities of BASP1 against the growth and migration of thyroid cancer cells were evaluated. Methods: BASP1 expression in thyroid cancer tissues and normal tissues were examined by immunohistochemical staining and the association between its expression and prognosis was analyzed. pcDNA-BASP1 carrying full length of BASP1 cDNA was constructed to restore the expression of BASP1 in thyroid cancer cell lines (BHT-101 and KMH-2). The cell proliferation in vitro and in vivo was evaluated by WST-1 assay and xenograft tumor models, respectively. Cell cycle distribution after transfection was analyzed using flow cytometry. Cell apoptosis after transfection was examined by annexin V/propidium iodide assay. The migration was examined using transwell assay. Results: BASP1 expression was abundant in normal tissues while it is significantly decreased in cancer tissues (P = 0.000). pcDNA-BASP1 restored the expression of BASP1 and significantly inhibited the growth of BHT-101 and KMH-2 cells as well as xenograft tumors in nude mice (P = 0.000). pcDNA-BASP1 induced G1 arrest and apoptosis in BHT-101 and KMH-2 cells. In addition, pcDNA-BASP1 significantly inhibited the cell migration. Conclusions: Downregulation of BASP1 expression may play a role in the tumorigenesis of thyroid cancer. Restoration of BASP1 expression exerted extensive antitumor activities against growth and migration of thyroid cancer cells, which suggested that BASP1 gene might act as a potential therapeutic agent for the treatment of thyroid cancer. PMID:27270539

  14. In Vivo Detection of Vascular Adhesion Protein-1 in Experimental Inflammation

    PubMed Central

    Jaakkola, Kimmo; Nikula, Tuomo; Holopainen, Riikka; Vähäsilta, Tommi; Matikainen, Marja-Terttu; Laukkanen, Marja-Leena; Huupponen, Risto; Halkola, Lauri; Nieminen, Lauri; Hiltunen, Jukka; Parviainen, Sakari; Clark, Michael R.; Knuuti, Juhani; Savunen, Timo; Kääpä, Pekka; Voipio-Pulkki, Liisa Maria; Jalkanen, Sirpa

    2000-01-01

    Vascular adhesion protein-1 (VAP-1) is an inflammation-inducible endothelial glycoprotein which mediates leukocyte-endothelial cell interactions. To study the pathogenetic significance of VAP-1 in inflammatory disorders, an in vivo immunodetection method was used to detect the regulation of luminally expressed VAP-1 in experimental skin and joint inflammation in the pig and dog. Moreover, VAP-1 was studied as a potential target to localize inflammation by radioimmunoscintigraphy. Up-regulation of VAP-1 in experimental dermatitis and arthritis could be visualized by specifically targeted immunoscintigraphy. Moreover, the translocation of VAP-1 to the functional position on the endothelial surface was only seen in inflamed tissues. These results suggest that VAP-1 is both an optimal candidate for anti-adhesive therapy and a potential target molecule for imaging inflammation. PMID:10934150

  15. Amyloid-beta peptide binds to microtubule-associated protein 1B (MAP1B).

    PubMed

    Gevorkian, Goar; Gonzalez-Noriega, Alfonso; Acero, Gonzalo; Ordoñez, Jorge; Michalak, Colette; Munguia, Maria Elena; Govezensky, Tzipe; Cribbs, David H; Manoutcharian, Karen

    2008-05-01

    Extracellular and intraneuronal formation of amyloid-beta aggregates have been demonstrated to be involved in the pathogenesis of Alzheimer's disease. However, the precise mechanism of amyloid-beta neurotoxicity is not completely understood. Previous studies suggest that binding of amyloid-beta to a number of targets have deleterious effects on cellular functions. In the present study we have shown for the first time that amyloid-beta 1-42 bound to a peptide comprising the microtubule binding domain of the heavy chain of microtubule-associated protein 1B by the screening of a human brain cDNA library expressed on M13 phage. This interaction may explain, in part, the loss of neuronal cytoskeletal integrity, impairment of microtubule-dependent transport and synaptic dysfunction observed previously in Alzheimer's disease.

  16. AMYLOID-β PEPTIDE BINDS TO MICROTUBULE-ASSOCIATED PROTEIN 1B (MAP1B)

    PubMed Central

    Gevorkian, Goar; Gonzalez-Noriega, Alfonso; Acero, Gonzalo; Ordoñez, Jorge; Michalak, Colette; Munguia, Maria Elena; Govezensky, Tzipe; Cribbs, David H.; Manoutcharian, Karen

    2008-01-01

    Extracellular and intraneuronal formation of amyloid-beta aggregates have been demonstrated to be involved in the pathogenesis of Alzheimer’s disease. However, the precise mechanism of amyloid-beta neurotoxicity is not completely understood. Previous studies suggest that binding of amyloid-beta to a number of targets have deleterious effects on cellular functions. In the present study we have shown for the first time that amyloid-beta 1-42 bound to a peptide comprising the microtubule binding domain of the heavy chain of microtubule-associated protein 1B by the screening of a human brain cDNA library expressed on M13 phage. This interaction may explain, in part, the loss of neuronal cytoskeletal integrity, impairment of microtubule-dependent transport and synaptic dysfunction observed previously in Alzheimer’s disease. PMID:18079022

  17. Phosphatidylethanolamine binding protein 1 in vacular endothelial cell autophagy and atherosclerosis

    PubMed Central

    Wang, Li; Li, HaiYing; Zhang, JinFeng; Lu, Wei; Zhao, Jing; Su, Le; Zhao, BaoXiang; Zhang, Yun; Zhang, ShangLi; Miao, JunYing

    2013-01-01

    We previously found that phosphatidylcholine-specific phospholipase C (PC-PLC) was a key inducing element of atherosclerosis, and might negatively regulate human umbilical vein endothelial cell (HUVEC) autophagy. To further investigate the mechanism of PC-PLC action, we initially identified phosphatidylethanolamine binding protein 1 (PEBP1) as a binding partner of PC-PLC by using mass spectrometry (MS, MALDI-TOF/TOF). We found that PEBP1 positively regulated PC-PLC activity in HUVECs, and inhibition of PC-PLC by its inhibitor D609 suppressed PEBP1 expression dramatically. Moreover, both PC-PLC and PEBP1 negatively regulated HUVEC autophagy independently of mammalian target of rapamycin (mTOR). Furthermore, the PEBP1 level was elevated during the development of atherosclerosis, while D609 significantly decreased the upregulated PEBP1 level in apoE−/− mice. PMID:23959677

  18. Modulation of Kaposi's Sarcoma-Associated Herpesvirus Interleukin-6 Function by Hypoxia-Upregulated Protein 1

    PubMed Central

    Giffin, Louise; Yan, Feng; Major, M. Ben

    2014-01-01

    ABSTRACT Kaposi's sarcoma-associated herpesvirus (KSHV, also called human herpesvirus 8) is linked to the development of Kaposi's sarcoma (KS), primary effusion lymphoma (PEL), and multicentric Castleman's disease (MCD). KSHV expresses several proteins that modulate host cell signaling pathways. One of these proteins is viral interleukin-6 (vIL-6), which is a homolog of human IL-6 (hIL-6). vIL-6 is able to prevent apoptosis and promote proinflammatory signaling, angiogenesis, and cell proliferation. Although it can be secreted, vIL-6 is mainly an intracellular protein that is retained in the endoplasmic reticulum (ER). We performed affinity purification and mass spectrometry to identify novel vIL-6 binding partners and found that a cellular ER chaperone, hypoxia-upregulated protein 1 (HYOU1), interacts with vIL-6. Immunohistochemical staining reveals that both PEL and KS tumor tissues express significant amounts of HYOU1. We also show that HYOU1 increases endogenous vIL-6 protein levels and that HYOU1 facilitates vIL-6-induced JAK/STAT signaling, migration, and survival in endothelial cells. Furthermore, our data suggest that HYOU1 also modulates vIL-6's ability to induce CCL2, a chemokine involved in cell migration. Finally, we investigated the impact of HYOU1 on cellular hIL-6 signaling. Collectively, our data indicate that HYOU1 is important for vIL-6 function and may play a role in the pathogenesis of KSHV-associated cancers. IMPORTANCE KSHV vIL-6 is detectable in all KSHV-associated malignancies and promotes tumorigenesis and inflammation. We identified a cellular protein, called hypoxia-upregulated protein 1 (HYOU1), that interacts with KSHV vIL-6 and is present in KSHV-infected tumors. Our data suggest that HYOU1 facilitates the vIL-6-induced signaling, migration, and survival of endothelial cells. PMID:24920810

  19. The impact of Cysteine-Rich Intestinal Protein 1 (CRIP1) in human breast cancer

    PubMed Central

    2013-01-01

    Background CRIP1 (cysteine-rich intestinal protein 1) has been found in several tumor types, its prognostic impact and its role in cellular processes, particularly in breast cancer, are still unclear. Methods To elucidate the prognostic impact of CRIP1, we analyzed tissues from 113 primary invasive ductal breast carcinomas using immunohistochemistry. For the functional characterization of CRIP1, its endogenous expression was transiently downregulated in T47D and BT474 breast cancer cells and the effects analyzed by immunoblotting, WST-1 proliferation assay and invasion assay. Results We found a significant correlation between CRIP1 and HER2 (human epidermal growth factor receptor 2) expression levels (p = 0.016) in tumor tissues. In Kaplan Meier analyses, CRIP1 expression was significantly associated with the distant metastases-free survival of patients, revealing a better prognosis for high CRIP1 expression (p = 0.039). Moreover, in multivariate survival analyses, the expression of CRIP1 was an independent negative prognostic factor, along with the positive prognosticators nodal status and tumor size (p = 0.029). CRIP1 knockdown in the T47D and BT474 breast cancer cell lines led to the increased phosphorylation of MAPK and Akt, to the reduced phosphorylation of cdc2, and to a significantly elevated cell proliferation in vitro (p < 0.001). These results indicate that reduced CRIP1 levels may increase cell proliferation and activate cell growth. In addition, CRIP1 knockdown increased cell invasion in vitro. Conclusions Because the lack of CRIP1 expression in breast cancer tissue is significantly associated with a worse prognosis for patients and low endogenous CRIP1 levels in vitro increased the malignant potential of breast cancer cells, we hypothesize that CRIP1 may act as a tumor suppressor in proliferation and invasion processes. Therefore, CRIP1 may be an independent prognostic marker with significant predictive power for use in breast cancer

  20. Overexpression of vascular adhesion protein-1 is associated with poor prognosis of astrocytomas.

    PubMed

    Kostoro, Joanna; Chang, Shu-Jyuan; Clark Lai, Yen-Chang; Wu, Chun-Chieh; Chai, Chee-Yin; Kwan, Aij-Lie

    2016-06-01

    Vascular adhesion protein-1 (VAP-1) is one of the endothelial adhesion molecules that is believed to play a role in tumor progression and metastasis, supporting cancer cell extravasation. Very few studies have been performed on analyzing the contribution of VAP-1 in brain tumor. Astrocytomas are the most common type of brain tumors, which are classified by World Health Organization (WHO) into four grades according to the degree of malignancy. This study was designed to investigate VAP-1 expression level in different astrocytoma grades and its correlation with clinicopathological features as well as prognosis of astrocytoma patients. Eighty-seven patients with different grades of astrocytoma (WHO Grade I-Grade IV) were enrolled in this study. The expression of VAP-1 was assayed by immunohistochemistry. The correlation between VAP-1 expression and clinicopathological features was evaluated by Chi-square test, and overall survival was analyzed by Kaplan-Meier method. Cox regression analysis was applied to analyze the independent influence of each parameter on overall survival. The expression level of VAP-1 was significantly higher in diffuse astrocytoma than those of pilocytic astrocytoma (p < 0.0001). In the subgroup analysis, upregulated VAP-1 expression was frequently found in older age patients (≥50 years). The VAP-1 expression was found to be significantly correlated with the overall survival (p = 0.0002). There was a statistical correlation between VAP-1(high) tumors in diffuse astrocytoma and VAP-1(low) tumors in pilocytic astrocytoma (p < 0.0001). Multivariate Cox analysis indicated VAP-1 was an independent predictive marker for poorer prognosis (p = 0.0036). Therefore, VAP-1 could be a promising prognostic biomarker in astrocytoma.

  1. Revisiting Apoplastic Auxin Signaling Mediated by AUXIN BINDING PROTEIN 1

    PubMed Central

    Feng, Mingxiao; Kim, Jae-Yean

    2015-01-01

    It has been suggested that AUXIN BINDING PROTEIN 1 (ABP1) functions as an apoplastic auxin receptor, and is known to be involved in the post-transcriptional process, and largely independent of the already well-known SKP-cullin-F-box-transport inhibitor response (TIR1) /auxin signaling F-box (AFB) (SCFTIR1/AFB) pathway. In the past 10 years, several key components downstream of ABP1 have been reported. After perceiving the auxin signal, ABP1 interacts, directly or indirectly, with plasma membrane (PM)-localized transmembrane proteins, transmembrane kinase (TMK) or SPIKE1 (SPK1), or other unidentified proteins, which transfer the signal into the cell to the Rho of plants (ROP). ROPs interact with their effectors, such as the ROP interactive CRIB motif-containing protein (RIC), to regulate the endocytosis/exocytosis of the auxin efflux carrier PIN-FORMED (PIN) proteins to mediate polar auxin transport across the PM. Additionally, ABP1 is a negative regulator of the traditional SCFTIR1/AFB auxin signaling pathway. However, Gao et al. (2015) very recently reported that ABP1 is not a key component in auxin signaling, and the famous abp1-1 and abp1-5 mutant Arabidopsis lines are being called into question because of possible additional mutantion sites, making it necessary to reevaluate ABP1. In this review, we will provide a brief overview of the history of ABP1 research. PMID:26467289

  2. Revisiting Apoplastic Auxin Signaling Mediated by AUXIN BINDING PROTEIN 1.

    PubMed

    Feng, Mingxiao; Kim, Jae-Yean

    2015-10-01

    It has been suggested that AUXIN BINDING PROTEIN 1 (ABP1) functions as an apoplastic auxin receptor, and is known to be involved in the post-transcriptional process, and largely independent of the already well-known SKP-cullin-F-box-transport inhibitor response (TIR1) /auxin signaling F-box (AFB) (SCF(TIR1/AFB)) pathway. In the past 10 years, several key components downstream of ABP1 have been reported. After perceiving the auxin signal, ABP1 interacts, directly or indirectly, with plasma membrane (PM)-localized transmembrane proteins, transmembrane kinase (TMK) or SPIKE1 (SPK1), or other unidentified proteins, which transfer the signal into the cell to the Rho of plants (ROP). ROPs interact with their effectors, such as the ROP interactive CRIB motif-containing protein (RIC), to regulate the endocytosis/exocytosis of the auxin efflux carrier PIN-FORMED (PIN) proteins to mediate polar auxin transport across the PM. Additionally, ABP1 is a negative regulator of the traditional SCF(TIR1/AFB) auxin signaling pathway. However, Gao et al. (2015) very recently reported that ABP1 is not a key component in auxin signaling, and the famous abp1-1 and abp1-5 mutant Arabidopsis lines are being called into question because of possible additional mutantion sites, making it necessary to reevaluate ABP1. In this review, we will provide a brief overview of the history of ABP1 research.

  3. Immunohistochemical localization of dentin matrix protein 1 in human dentin.

    PubMed

    Orsini, G; Ruggeri, A; Mazzoni, A; Nato, F; Falconi, M; Putignano, A; Di Lenarda, R; Nanci, A; Breschi, L

    2008-01-01

    Dentin matrix protein 1 (DMP1) is a non-collagenous matrix protein with a recognized role in the formation of mineralized tissues such as dentin. The aim of this study was to analyze the distribution of DMP1 in human dentin by means of immunofluorescence and high-resolution immunogold labeling. Fully developed, sound human dentin specimens were submitted to fluorescence labeling and post-embedding immunolabeling techniques with a rabbit polyclonal antihuman DMP1 antibody followed by corresponding fluorochrome-conjugated or gold-conjugated secondary antibodies. Both immunofluorescence and immunogold labeling showed an intense labeling associated with the peritubular dentin. In addition, at the ultrastructural level, there was also a moderate and diffuse immunoreaction over intertubular dentin, and a weak labeling within predentin which increased in density towards the mineralization front. This study suggests that in adult human teeth, like in rodents, DMP1 is prevalently concentrated at the level of peritubular dentin and this feature is preserved also in fully developed-teeth. These data are consistent with what has been observed in rodents and suggest that DMP1 plays a role in maintenance of the dentin tubular space.

  4. The Bone Morphogenetic Protein 1/Tolloid-like Metalloproteinases

    PubMed Central

    Hopkins, Delana R.; Keles, Sunduz; Greenspan, Daniel S.

    2009-01-01

    A decade ago, bone morphogenetic protein 1 (BMP1) was shown to provide the activity necessary for proteolytic removal of the C-propeptides of procollagens I–III: precursors of the major fibrillar collagens. Subsequent studies have shown BMP1 to be the prototype of a small group of extracellular metalloproteinases that play manifold roles in regulating formation of the extracellular matrix (ECM). Soon after initial cloning of BMP1, genetic studies showed the related Drosophila proteinase Tolloid (TLD) to be necessary for formation of the dorsal-ventral axis in early embryogenesis. It is now clear that the BMP1/TLD-like proteinases, conserved in species ranging from Drosophila to humans, act in dorsal-ventral patterning via activation of transforming growth factor β (TGFβ)-like proteins BMP2, BMP4 (vertebrates) and decapentaplegic (arthropods). More recently, it has become apparent that the BMP1/TLD-like proteinases are key activators of a broader subset of the TGFβ superfamily of proteins, with implications that these proteinases may be key in orchestrating formation of ECM with growth factor activation and BMP signaling in morphogenetic processes. PMID:17560775

  5. The Zinc Finger Transcription Factor ZXDC Activates CCL2 Gene Expression by Opposing BCL6-mediated Repression

    PubMed Central

    Ramsey, Jon E.; Fontes, Joseph D.

    2013-01-01

    The zinc finger X-linked duplicated (ZXD) family of transcription factors has been implicated in regulating transcription of major histocompatibility complex class II genes in antigen presenting cells; roles beyond this function are not yet known. The expression of one gene in this family, ZXD family zinc finger C (ZXDC), is enriched in myeloid lineages and therefore we hypothesized that ZXDC may regulate myeloid-specific gene expression. Here we demonstrate that ZXDC regulates genes involved in myeloid cell differentiation and inflammation. Overexpression of the larger isoform of ZXDC, ZXDC1, activates expression of monocyte-specific markers of differentiation and synergizes with phorbol 12-myristate 13-acetate (which causes differentiation) in the human leukemic monoblast cell line U937. To identify additional gene targets of ZXDC1, we performed gene expression profiling which revealed multiple inflammatory gene clusters regulated by ZXDC1. Using a combination of approaches we show that ZXDC1 activates transcription of a gene within one of the regulated clusters, chemokine (C-C motif) ligand 2 (CCL2; monocyte chemoattractant protein 1; MCP1) via a previously defined distal regulatory element. Further, ZXDC1-dependent up-regulation of the gene involves eviction of the transcriptional repressor B-cell CLL/lymphoma 6 (BCL6), a factor known to be important in resolving inflammatory responses, from this region of the promoter. Collectively, our data show that ZXDC1 is a regulator in the process of myeloid function and that ZXDC1 is responsible for Ccl2 gene de-repression by BCL6. PMID:23954399

  6. The Effect of Post-Resistance Exercise Amino Acids on Plasma MCP-1 and CCR2 Expression

    PubMed Central

    Wells, Adam J.; Hoffman, Jay R.; Jajtner, Adam R.; Varanoske, Alyssa N.; Church, David D.; Gonzalez, Adam M.; Townsend, Jeremy R.; Boone, Carleigh H.; Baker, Kayla M.; Beyer, Kyle S.; Mangine, Gerald T.; Oliveira, Leonardo P.; Fukuda, David H.; Stout, Jeffrey R.

    2016-01-01

    The recruitment and infiltration of classical monocytes into damaged muscle is critical for optimal tissue remodeling. This study examined the effects of an amino acid supplement on classical monocyte recruitment following an acute bout of lower body resistance exercise. Ten resistance-trained men (24.7 ± 3.4 years; 90.1 ± 11.3 kg; 176.0 ± 4.9 cm) ingested supplement (SUPP) or placebo (PL) immediately post-exercise in a randomized, cross-over design. Blood samples were obtained at baseline (BL), immediately (IP), 30-min (30P), 1-h (1H), 2-h (2H), and 5-h (5H) post-exercise to assess plasma concentrations of monocyte chemoattractant protein 1 (MCP-1), myoglobin, cortisol and insulin concentrations; and expressions of C-C chemokine receptor-2 (CCR2), and macrophage-1 antigen (CD11b) on classical monocytes. Magnitude-based inferences were used to provide inferences on the true effects of SUPP compared to PL. Changes in myoglobin, cortisol, and insulin concentrations were similar between treatments. Compared to PL, plasma MCP-1 was “very likely greater” (98.1% likelihood effect) in SUPP at 2H. CCR2 expression was “likely greater” at IP (84.9% likelihood effect), “likely greater” at 1H (87.7% likelihood effect), “very likely greater” at 2H (97.0% likelihood effect), and “likely greater” at 5H (90.1% likelihood effect) in SUPP, compared to PL. Ingestion of SUPP did not influence CD11b expression. Ingestion of an amino acid supplement immediately post-exercise appears to help maintain plasma MCP-1 concentrations and augment CCR2 expression in resistance trained men. PMID:27384580

  7. Nifedipine, a calcium channel blocker, inhibits advanced glycation end product (AGE)-elicited mesangial cell damage by suppressing AGE receptor (RAGE) expression via peroxisome proliferator-activated receptor-gamma activation

    SciTech Connect

    Matsui, Takanori; Yamagishi, Sho-ichi; Takeuchi, Masayoshi; Ueda, Seiji; Fukami, Kei; Okuda, Seiya

    2009-07-24

    The interaction between advanced glycation end products (AGE) and their receptor RAGE mediates the progressive alteration in renal architecture and loss of renal function in diabetic nephropathy. Oxidative stress generation and inflammation also play a central role in diabetic nephropathy. This study investigated whether and how nifedipine, a calcium channel blocker (CCB), blocked the AGE-elicited mesangial cell damage in vitro. Nifedipine, but not amlodipine, a control CCB, down-regulated RAGE mRNA levels and subsequently reduced reactive oxygen species (ROS) generation in AGE-exposed mesangial cells. AGE increased mRNA levels of vascular cell adhesion molecule-1 (VCAM-1) and induced monocyte chemoattractant protein-1 (MCP-1) production in mesangial cells, both of which were prevented by the treatment with nifedipine, but not amlodipine. The beneficial effects of nifedipine on AGE-exposed mesangial cells were blocked by the simultaneous treatment of GW9662, an inhibitor of peroxisome proliferator-activated receptor-{gamma} (PPAR-{gamma}). Although nifedipine did not affect expression levels of PPAR-{gamma}, it increased the PPAR-{gamma} transcriptional activity in mesangial cells. Our present study provides a unique beneficial aspect of nifedipine on diabetic nephropathy; it could work as an anti-inflammatory agent against AGE by suppressing RAGE expression in cultured mesangial cells via PPAR-{gamma} activation.

  8. The multidrug resistance-associated protein 1 transports methoxychlor and protects the seminiferous epithelium from injury.

    PubMed

    Tribull, Tiffany E; Bruner, Richard H; Bain, Lisa J

    2003-04-30

    We examined the ability of the multidrug resistance-associated protein 1 (MRP1/ABCC1) to transport pesticides, as this transporter mediates the cellular efflux of a variety of xenobiotics, typically as glucuronide, sulfate, or glutathione conjugates. NIH3T3 cells stably expressing MRP1 were 3.37-fold more resistant to the toxicity of fenitrothion, 3.12-fold more resistant to chlorpropham, and 2.5-fold more resistant to methoxychlor, a pesticide with estrogenic and anti-androgenic metabolites. The cells expressing MRP1 also eliminated methoxychlor two times more rapidly than their mock-transfected counterparts. We then examined whether mrp1 expression could alter the toxicity of methoxychlor in vivo using male FVB/mrp1 knockout mice (FVB/mrp1-/-). Both control and knockout mice were fed 25 mg/kg methoxychlor in honey for 39 days, and its effects on testicular morphology were examined. Methoxychlor treatment did not significantly affect testicular morphology in the FVB mice, but markedly reduced the number of developing spermatocytes in the FVB/mrp1-/- mice. These results suggest that MRPI may play a role in protecting the seminiferous tubules from methoxychlor-induced damage.

  9. Distribution of the SynDIG4/proline-rich transmembrane protein 1 in rat brain.

    PubMed

    Kirk, Lyndsey M; Ti, Shu W; Bishop, Hannah I; Orozco-Llamas, Mayra; Pham, Michelle; Trimmer, James S; Díaz, Elva

    2016-08-01

    The modulation of AMPA receptor (AMPAR) content at synapses is thought to be an underlying molecular mechanism of memory and learning. AMPAR content at synapses is highly plastic and is regulated by numerous AMPAR accessory transmembrane proteins such as TARPs, cornichons, and CKAMPs. SynDIG (synapse differentiation-induced gene) defines a family of four genes (SynDIG1-4) expressed in distinct and overlapping patterns in the brain. SynDIG1 was previously identified as a novel transmembrane AMPAR-associated protein that regulates synaptic strength. The related protein SynDIG4 [also known as Prrt1 (proline-rich transmembrane protein 1)] has recently been identified as a component of AMPAR complexes. In this study, we show that SynDIG1 and SynDIG4 have distinct yet overlapping patterns of expression in the central nervous system, with SynDIG4 having especially prominent expression in the hippocampus and particularly within CA1. In contrast to SynDIG1 and other traditional AMPAR auxiliary subunits, SynDIG4 is de-enriched at the postsynaptic density and colocalizes with extrasynaptic GluA1 puncta in primary dissociated neuron culture. These results indicate that, although SynDIG4 shares sequence similarity with SynDIG1, it might act through a unique mechanism as an auxiliary factor for extrasynaptic GluA1-containing AMPARs. J. Comp. Neurol. 524:2266-2280, 2016. © 2015 Wiley Periodicals, Inc.

  10. Depletion of Paraspeckle Protein 1 Enhances Methyl Methanesulfonate-Induced Apoptosis through Mitotic Catastrophe.

    PubMed

    Gao, Xiangjing; Zhang, Guanglin; Shan, Shigang; Shang, Yunlong; Chi, Linfeng; Li, Hongjuan; Cao, Yifei; Zhu, Xinqiang; Zhang, Meibian; Yang, Jun

    2016-01-01

    Previously, we have shown that paraspeckle protein 1 (PSPC1), a protein component of paraspeckles that was involved in cisplatin-induced DNA damage response (DDR), probably functions at the G1/S checkpoint. In the current study, we further examined the role of PSPC1 in another DNA-damaging agent, methyl methanesulfonate (MMS)-induced DDR, in particular, focusing on MMS-induced apoptosis in HeLa cells. First, it was found that MMS treatment induced the expression of PSPC1. While MMS treatment alone can induce apoptosis, depletion of PSPC1 expression using siRNA significantly increased the level of apoptosis following MMS exposure. In contrast, overexpressing PSPC1 decreased the number of apoptotic cells. Interestingly, morphological observation revealed that many of the MMS-treated PSPC1-knockdown cells contained two or more nuclei, indicating the occurrence of mitotic catastrophe. Cell cycle analysis further showed that depletion of PSPC1 caused more cells entering the G2/M phase, a prerequisite of mitosis catastrophe. On the other hand, over-expressing PSPC1 led to more cells accumulating in the G1/S phase. Taken together, these observations suggest an important role for PSPC1 in MMS-induced DDR, and in particular, depletion of PSPC1 can enhance MMS-induced apoptosis through mitotic catastrophe.

  11. Vitamin D represses dentin matrix protein 1 in cementoblasts and osteocytes.

    PubMed

    Nociti, F H; Foster, B L; Tran, A B; Dunn, D; Presland, R B; Wang, L; Bhattacharyya, N; Collins, M T; Somerman, M J

    2014-02-01

    Calcium and phosphorus homeostasis is achieved by interplay among hormones, including 1,25(OH)2D3 (1,25D), parathyroid hormone, and fibroblast growth factor 23 (FGF23), and their interactions with other proteins. For example, mutations in dentin matrix protein 1 (DMP-1) result in increased FGF23 and hypophosphatemic rickets. 1,25D is reported to modulate FGF23; thus, we hypothesized that 1,25D may be involved in modulating DMP-1 in an intermediary step. Murine cementoblasts (OCCM-30) and osteocyte-like cells (MLO-Y4 and MLO-A5), known to express DMP-1, were used to analyze effects of 1,25D on DMP-1 expression in vitro. DMP-1 mRNA levels decreased by 50% (p < .05) in the presence of 1,25D in all cell types, while use of a vitamin D receptor (VDR) agonist (EB1089) and antagonist (23S,25S)-DLAM-2P confirmed that VDR pathway activation was required for this response. Further analysis showed that histone deacetylase recruitment was necessary, but neither protein kinase A nor C pathways were required. In conclusion, our results support the hypothesis that 1,25D regulates DMP-1 expression through a VDR-dependent mechanism, possibly contributing to local changes in bone/tooth mineral homeostasis. PMID:24334408

  12. Evaluation of salivary gland protein 1 antibodies in patients with primary and secondary Sjogren's syndrome.

    PubMed

    Shen, Long; Kapsogeorgou, Efstathia K; Yu, Meixing; Suresh, Lakshmanan; Malyavantham, Kishore; Tzioufas, Anthanasios G; Ambrus, Julian L

    2014-11-01

    Sjogren's syndrome (SS) has been associated with the expression of anti-Ro and anti-La antibodies. Anti-salivary gland protein 1 (SP1) antibodies have recently been identified in patients with SS. The current work involved a cross sectional study to determine whether anti-SP1 antibodies were identified in particular subgroups of patients with SS. The results of this study revealed that anti-SP1 antibodies were present in the sera of 52% of SS patients while anti-Ro/anti-La was present in 63% of patients. 19% of patients had anti-SP1 without anti-Ro/anti-La. Patients with SS and lymphoma expressed anti-Ro, anti-La and anti-SP1 together. In SS associated with RA, 50% had antibodies anti-SP1 while 40% had anti-Ro/anti-La. In conclusion, anti-SP1 antibodies are commonly seen in both primary and secondary SS and rarely in normal controls. Future studies are needed to determine the roles and timing of expression of anti-SP1 antibodies in Sjogren's syndrome.

  13. Depletion of Paraspeckle Protein 1 Enhances Methyl Methanesulfonate-Induced Apoptosis through Mitotic Catastrophe

    PubMed Central

    Gao, Xiangjing; Zhang, Guanglin; Shan, Shigang; Shang, Yunlong; Chi, Linfeng; Li, Hongjuan; Cao, Yifei; Zhu, Xinqiang; Zhang, Meibian; Yang, Jun

    2016-01-01

    Previously, we have shown that paraspeckle protein 1 (PSPC1), a protein component of paraspeckles that was involved in cisplatin-induced DNA damage response (DDR), probably functions at the G1/S checkpoint. In the current study, we further examined the role of PSPC1 in another DNA-damaging agent, methyl methanesulfonate (MMS)-induced DDR, in particular, focusing on MMS-induced apoptosis in HeLa cells. First, it was found that MMS treatment induced the expression of PSPC1. While MMS treatment alone can induce apoptosis, depletion of PSPC1 expression using siRNA significantly increased the level of apoptosis following MMS exposure. In contrast, overexpressing PSPC1 decreased the number of apoptotic cells. Interestingly, morphological observation revealed that many of the MMS-treated PSPC1-knockdown cells contained two or more nuclei, indicating the occurrence of mitotic catastrophe. Cell cycle analysis further showed that depletion of PSPC1 caused more cells entering the G2/M phase, a prerequisite of mitosis catastrophe. On the other hand, over-expressing PSPC1 led to more cells accumulating in the G1/S phase. Taken together, these observations suggest an important role for PSPC1 in MMS-induced DDR, and in particular, depletion of PSPC1 can enhance MMS-induced apoptosis through mitotic catastrophe. PMID:26785254

  14. Human heterochromatin protein 1 isoforms regulate androgen receptor signaling in prostate cancer.

    PubMed

    Itsumi, Momoe; Shiota, Masaki; Yokomizo, Akira; Kashiwagi, Eiji; Takeuchi, Ario; Tatsugami, Katsunori; Inokuchi, Junichi; Song, Yoohyun; Uchiumi, Takeshi; Naito, Seiji

    2013-06-01

    Androgen receptor (AR) signaling is critical for the tumorigenesis and development of prostate cancer, as well as the progression to castration-resistant prostate cancer. We previously showed that the heterochromatin protein 1 (HP1) β isoform plays a critical role in transactivation of AR signaling as an AR coactivator that promotes prostate cancer cell proliferation. However, the roles of other HP1 isoforms, HP1α and HP1γ, in AR expression and prostate cancer remain unclear. Here, we found that knockdown of HP1γ, but not HP1α, reduced AR expression and cell proliferation by inducing cell cycle arrest at G1 phase in LNCaP cells. Conversely, overexpression of full-length HP1α and its C-terminal deletion mutant increased AR expression and cell growth, whereas overexpression of HP1γ had no effect. Similarly, HP1α overexpression promoted 22Rv1 cell growth, whereas HP1γ knockdown reduced the proliferation of CxR cells, a castration-resistant LNCaP derivative. Taken together, HP1 isoforms distinctly augment AR signaling and cell growth in prostate cancer. Therefore, silencing of HP1β and HP1γ may be a promising therapeutic strategy for treatment of prostate cancer.

  15. Nicotinic α7 receptor inhibits the acylation stimulating protein‑induced production of monocyte chemoattractant protein‑1 and keratinocyte‑derived chemokine in adipocytes by modulating the p38 kinase and nuclear factor‑κB signaling pathways.

    PubMed

    Jiao, Zhou-Yang; Wu, Jing; Liu, Chao; Wen, Bing; Zhao, Wen-Zeng; Du, Xin-Ling

    2016-10-01

    Obesity is associated with chronic low‑grade inflammation, which is characterized by increased infiltration of macrophages into adipose tissue. Acylation stimulating protein (ASP) is an adipokine derived from the immune complement system, which constitutes a link between adipocytes and macrophages, and is involved in energy homeostasis and inflammation. The purpose of the present study was to preliminarily investigate in vitro, whether functional α7nAChR in adipocytes may suppress ASP‑induced inflammation and determine the possible signaling mechanism. Studies have reported associations between the expression of α7 nicotinic acetylcholine receptor (α7nAChR) and obesity, insulin resistance and diabetes. Additionally, α7nAChRs are important peripheral mediators of chronic inflammation, which is a key contributor to health problems in obesity. The primary aim of the present study was to evaluate the impact of exogenous ASP and α7nAChR on macrophage infiltration in adipose tissue and to examine the potential underlying molecular mechanism. Western blot analysis revealed that recombinant ASP increased the expression levels of monocyte chemoattractant protein‑1 (MCP‑1) and keratinocyte‑derived chemokine (KC) by 3T3‑L1 adipocytes. However, nicotine significantly inhibited the production of ASP‑induced cytokines via the stimulation of α7nAChR. It was also found that α7nAChR inhibited the ASP‑induced activation of p38 kinase and nuclear factor‑κB (NF‑κB), and the production of MCP‑1 and KC. These data indicated that α7nAChR caused the inhibition of ASP‑induced activation of p38 kinase and NF‑κB to inhibit the production of MCP‑1 and KC. PMID:27572255

  16. Nicotinic α7 receptor inhibits the acylation stimulating protein‑induced production of monocyte chemoattractant protein‑1 and keratinocyte‑derived chemokine in adipocytes by modulating the p38 kinase and nuclear factor‑κB signaling pathways.

    PubMed

    Jiao, Zhou-Yang; Wu, Jing; Liu, Chao; Wen, Bing; Zhao, Wen-Zeng; Du, Xin-Ling

    2016-10-01

    Obesity is associated with chronic low‑grade inflammation, which is characterized by increased infiltration of macrophages into adipose tissue. Acylation stimulating protein (ASP) is an adipokine derived from the immune complement system, which constitutes a link between adipocytes and macrophages, and is involved in energy homeostasis and inflammation. The purpose of the present study was to preliminarily investigate in vitro, whether functional α7nAChR in adipocytes may suppress ASP‑induced inflammation and determine the possible signaling mechanism. Studies have reported associations between the expression of α7 nicotinic acetylcholine receptor (α7nAChR) and obesity, insulin resistance and diabetes. Additionally, α7nAChRs are important peripheral mediators of chronic inflammation, which is a key contributor to health problems in obesity. The primary aim of the present study was to evaluate the impact of exogenous ASP and α7nAChR on macrophage infiltration in adipose tissue and to examine the potential underlying molecular mechanism. Western blot analysis revealed that recombinant ASP increased the expression levels of monocyte chemoattractant protein‑1 (MCP‑1) and keratinocyte‑derived chemokine (KC) by 3T3‑L1 adipocytes. However, nicotine significantly inhibited the production of ASP‑induced cytokines via the stimulation of α7nAChR. It was also found that α7nAChR inhibited the ASP‑induced activation of p38 kinase and nuclear factor‑κB (NF‑κB), and the production of MCP‑1 and KC. These data indicated that α7nAChR caused the inhibition of ASP‑induced activation of p38 kinase and NF‑κB to inhibit the production of MCP‑1 and KC.

  17. Allurin, a 21 kD sperm chemoattractant, is rapidly released from the outermost jelly layer of the Xenopus egg by diffusion and medium convection.

    PubMed

    Xiang, Xueyu; Kittelson, Andrew; Olson, John; Bieber, Allan; Chandler, Douglas

    2005-03-01

    Allurin, a 21 kD protein from Xenopus laevis egg jelly, has been demonstrated to attract sperm by video microscopy and by quantitative chemotaxis chamber assays. Here, we use immunocytochemistry to demonstrate that this sperm chemoattractant is located in the outermost layer of egg jelly (J3) and is rapidly released into the surrounding medium. SDS-PAGE analysis and Western blotting confirm the appearance of allurin in the medium within 1.5 min and separation of proteins in the medium by anion exchange FPLC, shows that nearly half of the allurin released over a 12-hr period is discharged in the first 5 min. The kinetics of allurin release from J3 and its appearance in the medium were quantitatively accounted for, by computer simulation of mathematical diffusion and convection models. Comparison of simulation data to quantitative measurements of allurin appearance in the medium suggests that allurin, although larger than most chemoattractants, is effectively dispersed by a combination of diffusion and medium mixing at the jelly surface during spawning. Our model further predicts that the innermost jelly layer, J1, is less permeable to allurin than the other layers, allowing it to act as a "reflector" to speed up allurin discharge. PMID:15625699

  18. Crystallization of an engineered RUN domain of Rab6-interacting protein 1/DENND5

    SciTech Connect

    Fernandes, Humberto; Franklin, Edward; Khan, Amir R.

    2011-08-29

    Effectors of the Rab small GTPases are large multi-domain proteins which have proved difficult to express in soluble form in Escherichia coli. Generally, effectors are recruited to a distinct subcellular compartment by active (GTP-bound) Rabs, which are linked to membranes by one or two prenylated Cys residues at their C-termini. Following recruitment via their Rab-binding domain (RBD), effectors carry out various aspects of vesicle formation, transport, tethering and fusion through their other domains. Previously, successful purification of the RUN-PLAT tandem domains (residues 683-1061) of the 1263-residue Rab6-interacting protein 1 (R6IP1) required co-expression with Rab6, as attempts to solubly express the effector alone were unsuccessful. R6IP1 is also known as DENN domain-containing protein 5 (DENND5) and is expressed as two isoforms, R6IP1A/B (DENND5A/B), which differ by 24 amino acids at the N-terminus. Here, a deletion in R6IP1 was engineered to enable soluble expression and to improve the quality of the crystals grown in complex with Rab6. A large 23-residue loop linking two -helices in the RUN1 domain was removed and replaced with a short linker. This loop resides on the opposite face to the Rab6-binding site and is not conserved in the RUN-domain family. In contrast to wild-type R6IP1-Rab6 crystals, which took several weeks to grow to full size, the engineered R6IP1 (RPdel)-Rab6 crystals could be grown in a matter of days.

  19. Sequential, ordered acquisition of antibodies to Plasmodium falciparum erythrocyte membrane protein 1 domains.

    PubMed

    Cham, Gerald K K; Turner, Louise; Lusingu, John; Vestergaard, Lasse; Mmbando, Bruno P; Kurtis, Jonathan D; Jensen, Anja T R; Salanti, Ali; Lavstsen, Thomas; Theander, Thor G

    2009-09-01

    The binding of erythrocytes infected with mature blood stage parasites to the vascular bed is key to the pathogenesis of malignant malaria. The binding is mediated by members of Plasmodium falciparum erythrocyte membrane protein 1 (PfEMP1) family. PfEMP1s can be divided into groups, and it has previously been suggested that parasites expressing group A or B/A PfEMP1s are most pathogenic. To test the hypothesis that the first malaria infections in infants and young children are dominated by parasites expressing A and B/A PfEMP1s, we measured the plasma Ab level against 48 recombinant PfEMP1 domains of different groupings in 1342 individuals living in five African villages characterized by markedly different malaria transmission. We show that children progressively acquire a broader repertoire of anti-PfEMP1 Abs, but that the rate of expansion is governed by transmission intensity. However, independently of transmission intensity, Abs are first acquired to particular Duffy binding ligand-like domains belonging to group A or B/A PfEMP1s. The results support the view that anti-PfEMP1 Ab responses effectively structure the expenditure of the repertoire of PfEMP1 maintained by the parasite. Parasites expressing certain group A and B/A PfEMP1s are responded to first by individuals with limited previous exposure, and the resulting Abs reduce the fitness and pathogenicity of these parasites during subsequent infections. This allows parasites expressing less pathogenic PFEMP1s to dominate during later infections. The identification of PfEMP1 domains expressed by parasites causing disease in infants and young children is important for development of vaccines protecting against severe malaria.

  20. Nickel decreases cellular iron level and converts cytosolic aconitase to iron-regulatory protein 1 in A549 cells

    SciTech Connect

    Chen Haobin; Davidson, Todd; Singleton, Steven; Garrick, Michael D.; Costa, Max . E-mail: costam@env.med.nyu.edu

    2005-08-15

    Nickel (Ni) compounds are well-established carcinogens and are known to initiate a hypoxic response in cells via the stabilization and transactivation of hypoxia-inducible factor-1 alpha (HIF-1{alpha}). This change may be the consequence of nickel's interference with the function of several Fe(II)-dependent enzymes. In this study, the effects of soluble nickel exposure on cellular iron homeostasis were investigated. Nickel treatment decreased both mitochondrial and cytosolic aconitase (c-aconitase) activity in A549 cells. Cytosolic aconitase was converted to iron-regulatory protein 1, a form critical for the regulation of cellular iron homeostasis. The increased activity of iron-regulatory protein 1 after nickel exposure stabilized and increased transferrin receptor (Tfr) mRNA and antagonized the iron-induced ferritin light chain protein synthesis. The decrease of aconitase activity after nickel treatment reflected neither direct interference with aconitase function nor obstruction of [4Fe-4S] cluster reconstitution by nickel. Exposure of A549 cells to soluble nickel decreased total cellular iron by about 40%, a decrease that likely caused the observed decrease in aconitase activity and the increase of iron-regulatory protein 1 activity. Iron treatment reversed the effect of nickel on cytosolic aconitase and iron-regulatory protein 1. To assess the mechanism for the observed effects, human embryonic kidney (HEK) cells over expressing divalent metal transporter-1 (DMT1) were compared to A549 cells expressing only endogenous transporters for inhibition of iron uptake by nickel. The inhibition data suggest that nickel can enter via DMT1 and compete with iron for entry into the cell. This disturbance of cellular iron homeostasis by nickel may have a great impact on the ability of the cell to regulate a variety of cell functions, as well as create a state of hypoxia in cells under normal oxygen tension. These effects may be very important in how nickel exerts phenotypic

  1. Lysophosphatidic Acid Enhanced the Angiogenic Capability of Human Chondrocytes by Regulating Gi/NF-kB-Dependent Angiogenic Factor Expression

    PubMed Central

    Chen, Kai-Hua; Hong, Chang-Zern; Chang, Pey-Jium; Hsu, Hung-Chih

    2014-01-01

    Lysophosphatidic acid (LPA) has been found to mediate myeloid differentiation, stimulate osteogenesis, alter cell proliferation and migration, and inhibit apoptosis in chondrocytes. The effect of LPA on the angiogenic capability of chondrocytes is not clear. This study aimed to investigate its effect on the angiogenic capability of human chondrocytes and the underlying mechanism of these effects. Human chondrocyte cell line, CHON-001, commercialized human chondrocytes (HC) derived from normal human articular cartilage, and human vascular endothelial cells (HUVECs) were used as cell models in this study. The angiogenic capability of chondrocytes was determined by capillary tube formation, monolayer permeability, cell migration, and cell proliferation. An angiogenesis protein array kit was used to evaluate the secretion of angiogenic factors in conditioned medium. Angiogenin, insulin-like growth factor-binding protein 1 (IGFBP-1), interleukin (IL)-8, monocyte chemoattractant protein-1 (MCP-1), matrix metalloproteinase (MMP)-9, and vascular endothelial growth factor (VEGF) mRNA and protein expressions were evaluated by Q-RT-PCR and EIA, respectively. LPA receptor (LPAR) expression was determined by RT-PCR. Signaling pathways were clarified using inhibitors, Western blot analysis, and reporter assays. The LPA treatment promoted the angiogenic capability of CHON-001 cells and HC, resulting in enhanced HUVEC capillary tube formation, monolayer permeability, migration, and cell growth. Angiogenin, IGFBP-1, IL-8, MCP-1, MMP-9, and VEGF mRNA and protein expressions were significantly enhanced in LPA-treated chondrocytes. LPA2, 3, 4 and 6 were expressed in CHON-001 and HC cells. Pretreatment with the Gi/o type G protein inhibitor, pertussis toxin (PTX), and the NF-kB inhibitor, PDTC, significantly inhibited LPA-induced angiogenin, IGFBP-1, IL-8, MCP-1, MMP-9, and VEGF expressions in chondrocytes. The PTX pretreatment also inhibited LPA-mediated NF-kB activation, suggesting

  2. Guanylate-Binding Protein-1 protects ovarian cancer cell lines but not breast cancer cell lines from killing by paclitaxel.

    PubMed

    Tipton, Aaron R; Nyabuto, Geoffrey O; Trendel, Jill A; Mazur, Travis M; Wilson, John P; Wadi, Suzan; Justinger, Jacob S; Moore, Garret L; Nguyen, Peter T; Vestal, Deborah J

    2016-09-30

    Forced expression of the cytokine-induced large GTPase, human Guanylate-Binding Protein-1 (hGBP-1), in ovarian cancer cell lines increases resistance to paclitaxel. Elevated hGBP-1 RNA in ovarian tumors correlates with shorter recurrence-free survival. In contract, hGBP-1 is part of a gene signature predicting improved prognosis in all subtypes of breast cancers. hGBP-1 does not confer paclitaxel resistance on MCF-7 and TMX2-28 breast cancer cells. Expression of the isotype of the hGBP-1-interacting protein, PIM1, which may contribute to paclitaxel resistance when associated with hGBP-1, is different in breast and ovarian cancer cell lines. Breast cancer cell lines express the 44 kDa isoform of PIM-1, and ovarian cancer cell lines express the 33 kDa isoform. PMID:27590579

  3. Developmental regulation of collagenase-3 mRNA in normal, differentiating osteoblasts through the activator protein-1 and the runt domain binding sites

    NASA Technical Reports Server (NTRS)

    Winchester, S. K.; Selvamurugan, N.; D'Alonzo, R. C.; Partridge, N. C.

    2000-01-01

    Collagenase-3 mRNA is initially detectable when osteoblasts cease proliferation, increasing during differentiation and mineralization. We showed that this developmental expression is due to an increase in collagenase-3 gene transcription. Mutation of either the activator protein-1 or the runt domain binding site decreased collagenase-3 promoter activity, demonstrating that these sites are responsible for collagenase-3 gene transcription. The activator protein-1 and runt domain binding sites bind members of the activator protein-1 and core-binding factor family of transcription factors, respectively. We identified core-binding factor a1 binding to the runt domain binding site and JunD in addition to a Fos-related antigen binding to the activator protein-1 site. Overexpression of both c-Fos and c-Jun in osteoblasts or core-binding factor a1 increased collagenase-3 promoter activity. Furthermore, overexpression of c-Fos, c-Jun, and core-binding factor a1 synergistically increased collagenase-3 promoter activity. Mutation of either the activator protein-1 or the runt domain binding site resulted in the inability of c-Fos and c-Jun or core-binding factor a1 to increase collagenase-3 promoter activity, suggesting that there is cooperative interaction between the sites and the proteins. Overexpression of Fra-2 and JunD repressed core-binding factor a1-induced collagenase-3 promoter activity. Our results suggest that members of the activator protein-1 and core-binding factor families, binding to the activator protein-1 and runt domain binding sites are responsible for the developmental regulation of collagenase-3 gene expression in osteoblasts.

  4. Wnt-induced secreted protein-1 is a prohypertrophic and profibrotic growth factor.

    PubMed

    Colston, J T; de la Rosa, S D; Koehler, M; Gonzales, K; Mestril, R; Freeman, G L; Bailey, S R; Chandrasekar, B

    2007-09-01

    Wnt1-induced secreted protein-1 (WISP-1) is a member of the cysteine-rich 61, connective tissue growth factor, and nephroblastoma overexpressed (CCN) family of growth factors and is expressed in the heart at low basal levels. The purpose of this study was to investigate whether WISP-1 is upregulated in postinfarct myocardium and whether WISP-1 exerts prohypertrophic and mitogenic effects stimulating myocyte hypertrophy, cardiac fibroblast (CF) proliferation, and collagen expression. Male C57Bl/6 (25 g) mice underwent permanent occlusion of the left anterior descending coronary artery. mRNA and protein levels were analyzed by Northern and Western blot analyses. Cardiomyocyte hypertrophy was quantified by protein and DNA synthesis. CF proliferation was quantified by CyQuant assay, and soluble collagen release by Sircol assay. A time-dependent increase in WISP-1 expression was detected in vivo in the noninfarct zone of the left ventricle, which peaked at 24 h (3.1-fold, P < 0.01). Similarly, biglycan expression was increased by 3.71-fold (P < 0.01). IL-1beta and TNF-alpha expression preceded WISP-1 expression in vivo and stimulated WISP-1 expression in neonatal rat ventricular myocytes in vitro. WISP-1-induced cardiomyocyte hypertrophy was evidenced by increased protein (2.78-fold), but not DNA synthesis, and enhanced Akt phosphorylation and activity. Treatment of primary CF with WISP-1 significantly stimulated proliferation at 48 h (6,966 +/- 264 vs. 5,476 +/- 307 cells/well, P < 0.01) and enhanced collagen release by 72 h (18.4 +/- 3.1 vs. 8.4 +/- 1.0 ng/cell, P < 0.01). Our results demonstrate for the first time that WISP-1 and biglycan are upregulated in the noninfarcted myocardium in vivo, suggesting a positive amplification of WISP-1 signaling. WISP-1 stimulates cardiomyocyte hypertrophy, fibroblast proliferation, and ECM expression in vitro. These results suggest that WISP-1 may play a critical role in post-myocardial infarction remodeling.

  5. Inhibitory effects of omega-3 fatty acids on early brain injury after subarachnoid hemorrhage in rats: Possible involvement of G protein-coupled receptor 120/β-arrestin2/TGF-β activated kinase-1 binding protein-1 signaling pathway.

    PubMed

    Yin, Jia; Li, Haiying; Meng, Chengjie; Chen, Dongdong; Chen, Zhouqing; Wang, Yibin; Wang, Zhong; Chen, Gang

    2016-06-01

    Omega-3 fatty acids have been reported to improve neuron functions during aging and in patients affected by mild cognitive impairment, and mediate potent anti-inflammatory via G protein-coupled receptor 120 (GPR120) signal pathway. Neuron dysfunction and inflammatory response also contributed to the progression of subarachnoid hemorrhage (SAH)-induced early brain injury (EBI). This study was to examine the effects of omega-3 fatty acids on SAH-induced EBI. Two weeks before SAH, 30% Omega-3 fatty acids was administered by oral gavage at 1g/kg body weight once every 24h. Specific siRNA for GPR120 was exploited. Terminal deoxynucleotidyl transferase dUTP nick end labeling, fluoro-Jade B staining, and neurobehavioral scores and brain water content test showed that omega-3 fatty acids effectively suppressed SAH-induced brain cell apoptosis and neuronal degradation, behavioral impairment, and brain edema. Western blot, immunoprecipitation, and electrophoretic mobility shift assays results showed that omega-3 fatty acids effectively suppressed SAH-induced elevation of inflammatory factors, including cyclooxygenase-2, monocyte chemoattractant protein-1, and inducible nitric oxide synthase. In addition, omega-3 fatty acids could inhibit phosphorylation of transforming growth factor β activated kinase-1 (TAK1), MEK4, c-Jun N-terminal kinase, and IkappaB kinase as well as activation of nuclear factor kappa B through regulating GPR120/β-arrestin2/TAK1 binding protein-1 pathway. Furthermore, siRNA-induced GPR120 silencing blocked the protective effects of omega-3 fatty acids. Here, we show that stimulation of GPR120 with omega-3 fatty acids pretreatment causes anti-apoptosis and anti-inflammatory effects via β-arrestin2/TAK1 binding protein-1/TAK1 pathway in the brains of SAH rats. Fish omega-3 fatty acids as part of a daily diet may reduce EBI in an experimental rat model of SAH.

  6. Inhibitory effects of omega-3 fatty acids on early brain injury after subarachnoid hemorrhage in rats: Possible involvement of G protein-coupled receptor 120/β-arrestin2/TGF-β activated kinase-1 binding protein-1 signaling pathway.

    PubMed

    Yin, Jia; Li, Haiying; Meng, Chengjie; Chen, Dongdong; Chen, Zhouqing; Wang, Yibin; Wang, Zhong; Chen, Gang

    2016-06-01

    Omega-3 fatty acids have been reported to improve neuron functions during aging and in patients affected by mild cognitive impairment, and mediate potent anti-inflammatory via G protein-coupled receptor 120 (GPR120) signal pathway. Neuron dysfunction and inflammatory response also contributed to the progression of subarachnoid hemorrhage (SAH)-induced early brain injury (EBI). This study was to examine the effects of omega-3 fatty acids on SAH-induced EBI. Two weeks before SAH, 30% Omega-3 fatty acids was administered by oral gavage at 1g/kg body weight once every 24h. Specific siRNA for GPR120 was exploited. Terminal deoxynucleotidyl transferase dUTP nick end labeling, fluoro-Jade B staining, and neurobehavioral scores and brain water content test showed that omega-3 fatty acids effectively suppressed SAH-induced brain cell apoptosis and neuronal degradation, behavioral impairment, and brain edema. Western blot, immunoprecipitation, and electrophoretic mobility shift assays results showed that omega-3 fatty acids effectively suppressed SAH-induced elevation of inflammatory factors, including cyclooxygenase-2, monocyte chemoattractant protein-1, and inducible nitric oxide synthase. In addition, omega-3 fatty acids could inhibit phosphorylation of transforming growth factor β activated kinase-1 (TAK1), MEK4, c-Jun N-terminal kinase, and IkappaB kinase as well as activation of nuclear factor kappa B through regulating GPR120/β-arrestin2/TAK1 binding protein-1 pathway. Furthermore, siRNA-induced GPR120 silencing blocked the protective effects of omega-3 fatty acids. Here, we show that stimulation of GPR120 with omega-3 fatty acids pretreatment causes anti-apoptosis and anti-inflammatory effects via β-arrestin2/TAK1 binding protein-1/TAK1 pathway in the brains of SAH rats. Fish omega-3 fatty acids as part of a daily diet may reduce EBI in an experimental rat model of SAH. PMID:27000704

  7. Lipoprotein receptor-related protein 1 variants and dietary fatty acids: meta-analysis of European origin and African American studies

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Low-density lipoprotein-related receptor protein 1 (LRP1) is a multi-functional endocytic receptor and signaling molecule that is expressed in adipose and the hypothalamus. Evidence for a role of LRP1 in adiposity is accumulating from animal and in vitro models, but data from human studies are limit...

  8. Regulatory effect of cytokine-induced neutrophil chemoattractant, epithelial neutrophil-activating peptide 78 and pyrrolidine dithiocarbamate on pulmonary neutrophil aggregation mediated by nuclear factor-κB in lipopolysaccharide-induced acute respiratory distress syndrome mice

    PubMed Central

    Wang, Hongman; Zhao, Jiping; Xue, Guansheng; Wang, Junfei; Wu, Jinxiang; Wang, Donghui; Dong, Liang

    2016-01-01

    In the present study, the regulatory effect of cytokine-induced neutrophil chemoattractant (CINC) and epithelial neutrophil-activating peptide 78 (ENA-78) on pulmonary neutrophil (PMN) accumulation in lipopolysaccharide (LPS)-induced acute respiratory distress syndrome (ARDS) mice, and the therapeutic effect of pyrrolidine dithiocarbamate (PDTC), was investigated. BALB/c mice were divided into control, LPS and PDTC + LPS groups using a random number table. The phosphorylation of nuclear factor-κB (NF-κB) was detected using a western blot, and the mRNA expression levels of CINC were evaluated using reverse transcription-quantitative polymerase chain reaction. The expression of NF-κB, CINC and ENA-78 was detected using immunohistochemistry. The production of interleukin (IL)-8 and IL-10 in serum and broncho-alveolar lavage fluid (BALF) was analyzed using an enzyme-linked immunosorbent assay. The total number of leukocytes and proportion of PMNs in BALF was also determined. Following injection with LPS (20 mg/kg), the expression levels of p-NF-κB, CINC and ENA-78 were increased in lung tissue, and the expression levels of IL-8, IL-10 and the number of PMNs increased in serum and BALF. However, in comparison with the LPS group, the degree of lung injury was reduced in ARDS mice that were treated with PDTC. In addition, the expression level of p-NF-κB and the production of chemokines in lung tissue decreased in ARDS mice that were treated with PDTC, and the number of PMNs in BALF also decreased. In conclusion, the results of the present study suggest that the LPS-induced phosphorylation of NF-κB may result in the synthesis and release of CINC and ENA-78, which induce the accumulation of PMNs in the lung. Therefore, PDTC may be used to reduce the production of chemokines and cytokines, thereby decreasing the activation of PMNs in lung tissue and reducing the damage of lung tissue in ARDS. PMID:27602092

  9. Regulatory effect of cytokine-induced neutrophil chemoattractant, epithelial neutrophil-activating peptide 78 and pyrrolidine dithiocarbamate on pulmonary neutrophil aggregation mediated by nuclear factor-κB in lipopolysaccharide-induced acute respiratory distress syndrome mice

    PubMed Central

    Wang, Hongman; Zhao, Jiping; Xue, Guansheng; Wang, Junfei; Wu, Jinxiang; Wang, Donghui; Dong, Liang

    2016-01-01

    In the present study, the regulatory effect of cytokine-induced neutrophil chemoattractant (CINC) and epithelial neutrophil-activating peptide 78 (ENA-78) on pulmonary neutrophil (PMN) accumulation in lipopolysaccharide (LPS)-induced acute respiratory distress syndrome (ARDS) mice, and the therapeutic effect of pyrrolidine dithiocarbamate (PDTC), was investigated. BALB/c mice were divided into control, LPS and PDTC + LPS groups using a random number table. The phosphorylation of nuclear factor-κB (NF-κB) was detected using a western blot, and the mRNA expression levels of CINC were evaluated using reverse transcription-quantitative polymerase chain reaction. The expression of NF-κB, CINC and ENA-78 was detected using immunohistochemistry. The production of interleukin (IL)-8 and IL-10 in serum and broncho-alveolar lavage fluid (BALF) was analyzed using an enzyme-linked immunosorbent assay. The total number of leukocytes and proportion of PMNs in BALF was also determined. Following injection with LPS (20 mg/kg), the expression levels of p-NF-κB, CINC and ENA-78 were increased in lung tissue, and the expression levels of IL-8, IL-10 and the number of PMNs increased in serum and BALF. However, in comparison with the LPS group, the degree of lung injury was reduced in ARDS mice that were treated with PDTC. In addition, the expression level of p-NF-κB and the production of chemokines in lung tissue decreased in ARDS mice that were treated with PDTC, and the number of PMNs in BALF also decreased. In conclusion, the results of the present study suggest that the LPS-induced phosphorylation of NF-κB may result in the synthesis and release of CINC and ENA-78, which induce the accumulation of PMNs in the lung. Therefore, PDTC may be used to reduce the production of chemokines and cytokines, thereby decreasing the activation of PMNs in lung tissue and reducing the damage of lung tissue in ARDS.

  10. Dorsal root ganglion myeloid zinc finger protein 1 contributes to neuropathic pain after peripheral nerve trauma.

    PubMed

    Li, Zhisong; Gu, Xiyao; Sun, Linlin; Wu, Shaogen; Liang, Lingli; Cao, Jing; Lutz, Brianna Marie; Bekker, Alex; Zhang, Wei; Tao, Yuan-Xiang

    2015-04-01

    Peripheral nerve injury-induced changes in gene transcription and translation in primary sensory neurons of the dorsal root ganglion (DRG) are considered to contribute to neuropathic pain genesis. Transcription factors control gene expression. Peripheral nerve injury increases the expression of myeloid zinc finger protein 1 (MZF1), a transcription factor, and promotes its binding to the voltage-gated potassium 1.2 (Kv1.2) antisense (AS) RNA gene in the injured DRG. However, whether DRG MZF1 participates in neuropathic pain is still unknown. Here, we report that blocking the nerve injury-induced increase of DRG MZF1 through microinjection of MZF1 siRNA into the injured DRG attenuated the initiation and maintenance of mechanical, cold, and thermal pain hypersensitivities in rats with chronic constriction injury (CCI) of the sciatic nerve, without affecting locomotor functions and basal responses to acute mechanical, heat, and cold stimuli. Mimicking the nerve injury-induced increase of DRG MZF1 through microinjection of recombinant adeno-associated virus 5 expressing full-length MZF1 into the DRG produced significant mechanical, cold, and thermal pain hypersensitivities in naive rats. Mechanistically, MZF1 participated in CCI-induced reductions in Kv1.2 mRNA and protein and total Kv current and the CCI-induced increase in neuronal excitability through MZF1-triggered Kv1.2 AS RNA expression in the injured DRG neurons. MZF1 is likely an endogenous trigger of neuropathic pain and might serve as a potential target for preventing and treating this disorder. PMID:25630025

  11. Female-Specific Induction of Rat Pituitary Dentin Matrix Protein-1 by GnRH

    PubMed Central

    Kucka, Marek; Bjelobaba, Ivana; Clokie, Samuel J. H.; Klein, David C.

    2013-01-01

    Hypothalamic GnRH is the primary regulator of reproduction in vertebrates, acting via the G protein-coupled GnRH receptor (GnRHR) in pituitary gonadotrophs to control synthesis and release of gonadotropins. To identify elements of the GnRHR-coupled gene network, GnRH was applied in a pulsatile manner for 6 hours to a mixed population of perifused pituitary cells from cycling females, mRNA was extracted, and RNA sequencing analysis was performed. This revealed 83 candidate-regulated genes, including a large number coding for secreted proteins. Most notably, GnRH induces a greater than 600-fold increase in expression of dentin matrix protein-1 (Dmp1), one of five members of the small integrin-binding ligand N-linked glycoprotein gene family. The Dmp1 response is mediated by the GnRHR, not elicited by other hypothalamic releasing factors, and is approximately 20-fold smaller in adult male pituitary cells. The sex-dependent Dmp1 response is established during the peripubertal period and independent of the developmental pattern of Gnrhr expression. In vitro, GnRH-induced expression of this gene is coupled with release of DMP1 in extracellular medium through the regulated secretory pathway. In vivo, pituitary Dmp1 expression in identified gonadotrophs is elevated after ovulation. Cell signaling studies revealed that the GnRH induction of Dmp1 is mediated by the protein kinase C signaling pathway and reflects opposing roles of ERK1/2 and p38 MAPK; in addition, the response is facilitated by progesterone. These results establish that DMP1 is a novel secretory protein of female rat gonadotrophs, the synthesis and release of which are controlled by the hypothalamus through the GnRHR signaling pathway. This advance raises intriguing questions about the intrapituitary and downstream effects of this new player in GnRH signaling. PMID:24085820

  12. Yes-Associated Protein 1 Is Activated and Functions as an Oncogene in Meningiomas

    PubMed Central

    Baia, Gilson S.; Caballero, Otavia L.; Orr, Brent A.; Lal, Anita; Ho, Janelle S.Y.; Cowdrey, Cynthia; Tihan, Tarik; Mawrin, Christian; Riggins, Gregory J.

    2015-01-01

    The Hippo signaling pathway is functionally conserved in Drosophila melanogaster and mammals, and its proposed function is to control tissue homeostasis by regulating cell proliferation and apoptosis. The core components are composed of a kinase cascade that culminates with the phosphorylation and inhibition of Yes-associated protein 1 (YAP1). Phospho-YAP1 is retained in the cytoplasm. In the absence of Hippo signaling, YAP1 translocates to the nucleus, associates with co-activators TEAD1-4, and functions as a transcriptional factor promoting the expression of key target genes. Components of the Hippo pathway are mutated in human cancers, and deregulation of this pathway plays a role in tumorigenesis. Loss of the NF2 tumor suppressor gene is the most common genetic alteration in meningiomas, and the NF2 gene product, Merlin, acts upstream of the Hippo pathway. Here, we show that primary meningioma tumors have high nuclear expression of YAP1. In meningioma cells, Merlin expression is associated with phosphorylation of YAP1. Using an siRNA transient knockdown of YAP1 in NF2-mutant meningioma cells, we show that suppression of YAP1 impaired cell proliferation and migration. Conversely, YAP1 overexpression led to a strong augment of cell proliferation and anchorage-independent growth and restriction of cisplatin-induced apoptosis. In addition, expression of YAP1 in nontransformed arachnoidal cells led to the development of tumors in nude mice. Together, these findings suggest that in meningiomas, deregulation of the Hippo pathway is largely observed in primary tumors and that YAP1 functions as an oncogene promoting meningioma tumorigenesis. PMID:22618028

  13. Effects of obesity on severity of colitis and cytokine expression in mouse mesenteric fat. Potential role of adiponectin receptor 1.

    PubMed

    Sideri, Aristea; Stavrakis, Dimitris; Bowe, Collin; Shih, David Q; Fleshner, Phillip; Arsenescu, Violeta; Arsenescu, Razvan; Turner, Jerrold R; Pothoulakis, Charalabos; Karagiannides, Iordanes

    2015-04-01

    In inflammatory bowel disease (IBD), obesity is associated with worsening of the course of disease. Here, we examined the role of obesity in the development of colitis and studied mesenteric fat-epithelial cell interactions in patients with IBD. We combined the diet-induce obesity with the trinitrobenzene sulfonic acid (TNBS) colitis mouse model to create groups with obesity, colitis, and their combination. Changes in the mesenteric fat and intestine were assessed by histology, myeloperoxidase assay, and cytokine mRNA expression by real-time PCR. Medium from human mesenteric fat and cultured preadipocytes was obtained from obese patients and those with IBD. Histological analysis showed inflammatory cell infiltrate and increased histological damage in the intestine and mesenteric fat of obese mice with colitis compared with all other groups. Obesity also increased the expression of proinflammatory cytokines including IL-1β, TNF-α, monocyte chemoattractant protein 1, and keratinocyte-derived chemokine, while it decreased the TNBS-induced increases in IL-2 and IFN-γ in mesenteric adipose and intestinal tissues. Human mesenteric fat isolated from obese patients and those with and IBD demonstrated differential release of adipokines and growth factors compared with controls. Fat-conditioned media reduced adiponectin receptor 1 (AdipoR1) expression in human NCM460 colonic epithelial cells. AdipoR1 intracolonic silencing in mice exacerbated TNBS-induced colitis. In conclusion, obesity worsens the outcome of experimental colitis, and obesity- and IBD-associated changes in adipose tissue promote differential mediator release in mesenteric fat that modulates colonocyte responses and may affect the course of colitis. Our results also suggest an important role for AdipoR1 for the fat-intestinal axis in the regulation of inflammation during colitis.

  14. Apoptosis signal-regulating kinase 1 is mediated in TNF-α-induced CCL2 expression in human synovial fibroblasts.

    PubMed

    Tsou, Hsi-Kai; Chen, Hsien-Te; Chang, Chia-Hao; Yang, Wan-Yu; Tang, Chih-Hsin

    2012-11-01

    Tumor necrosis factor-α (TNF-α), a pro-inflammatory cytokine with a critical role in osteoarthritis (OA), was primarily produced by monocytes/macrophages and plays a crucial role in the inflammatory response. Here, we investigated the intracellular signaling pathways involved in TNF-α-induced monocyte chemoattractant protein 1 (MCP-1)/CCL2 expression in human synovial fibroblast cells. Stimulation of synovial fibroblasts (OASF) with TNF-α induced concentration- and time-dependent increases in CCL2 expression. TNF-α-mediated CCL2 production was attenuated by TNFR1 monoclonal antibody (Ab). Pretreatment with an apoptosis signal-regulating kinase 1 (ASK1) inhibitor (thioredoxin), JNK inhibitor (SP600125), p38 inhibitor (SB203580), or AP-1 inhibitor (curcumin or tanshinone IIA) also blocked the potentiating action of TNF-α. Stimulation of cells with TNF-α enhanced ASK1, JNK, and p38 activation. Treatment of OASF with TNF-α also increased the accumulation of phosphorylated c-Jun in the nucleus, AP-1-luciferase activity, and c-Jun binding to the AP-1 element on the CCL2 promoter. TNF-α-mediated AP-1-luciferase activity and c-Jun binding to the AP-1 element were inhibited by TNFR1 Ab, thioredoxin, SP600125, and SB203580. Our results suggest that the interaction between TNF-α and TNFR1 increases CCL2 expression in human synovial fibroblasts via the ASK1, JNK/p38, c-Jun, and AP-1 signaling pathway. PMID:22711527

  15. D-Psicose inhibits the expression of MCP-1 induced by high-glucose stimulation in HUVECs.

    PubMed

    Murao, Koji; Yu, Xiao; Cao, Wen M; Imachi, Hitomi; Chen, Ke; Muraoka, Tomie; Kitanaka, Noriko; Li, Junhun; Ahmed, Rania A M; Matsumoto, Kensuke; Nishiuchi, Takamasa; Tokuda, Masaaki; Ishida, Toshihiko

    2007-07-26

    Monocyte chemoattractant protein-1 (MCP-1) is a 76-amino-acid chemokine thought to be the major chemotactic factor for monocytes. MCP-1 is found in macrophage-rich areas of atherosclerotic lesions. Recent report indicates that MCP-1 is induced by glucose-stimulation, raising the important link between diabetes mellitus and atherosclerosis. One of the rare sugars, d-psicose (d-ribo-2-hexulose) is present in small quantities in commercial carbohydrate complexes, however the physiological functions of d-psicose have not been evaluated. In this study, we examined the effects of d-psicose on MCP-1 expression in human umbilical vein endothelial cells (HUVECs). Results showed that MCP-1 mRNA and protein were stimulated following exposure to 22.4 mM glucose. Transcriptional activity of MCP-1 promoter paralleled endogenous expression of the gene and this activity was dependent on the dose of d-glucose. d-Psicose inhibited these effects. Next we used inhibitors of selected signal transduction pathways to show that high-glucose (HG) stimulated MCP-1 promoter activity was sensitive to p38-Mitogen-Activated Protein Kinase (p38-MAPK) pathway inhibitor. As expected, a dominant-negative p38-MAPK abolished the stimulatory effect of HG on the promoter activity. To incubate the cells with HG and d-psicose reduced the activation of p38-MAPK. Together, these results indicate that the d-psicose suppression of HG induced MCP-1 expression is mediated in part by inhibition of the p38-MAPK pathway and raise the possibility that d-psicose may be of therapeutic value in the treatment of diseases such as atherosclerosis.

  16. Molecular cloning and subcellular localization of Tektin2-binding protein 1 (Ccdc 172) in rat spermatozoa.

    PubMed

    Yamaguchi, Airi; Kaneko, Takane; Inai, Tetsuichiro; Iida, Hiroshi

    2014-04-01

    Tektins (TEKTs) are composed of a family of filament-forming proteins localized in cilia and flagella. Five types of mammalian TEKTs have been reported, all of which have been verified to be present in sperm flagella. TEKT2, which is indispensable for sperm structure, mobility, and fertilization, was present at the periphery of the outer dense fiber (ODF) in the sperm flagella. By yeast two-hybrid screening, we intended to isolate flagellar proteins that could interact with TEKT2, which resulted in the isolation of novel two genes from the mouse testis library, referred as a TEKT2-binding protein 1 (TEKT2BP1) and -protein 2 (TEKT2BP2). In this study, we characterized TEKT2BP1, which is registered as a coiled-coil domain-containing protein 172 (Ccdc172) in the latest database. RT-PCR analysis indicated that TEKT2BP1 was predominantly expressed in rat testis and that its expression was increased after 3 weeks of postnatal development. Immunocytochemical studies discovered that TEKT2BP1 localized in the middle piece of rat spermatozoa, predominantly concentrated at the mitochondria sheath of the flagella. We hypothesize that the TEKT2-TEKT2BP1 complex might be involved in the structural linkage between the ODF and mitochondria in the middle piece of the sperm flagella.

  17. Molecular cloning and subcellular localization of Tektin2-binding protein 1 (Ccdc 172) in rat spermatozoa.

    PubMed

    Yamaguchi, Airi; Kaneko, Takane; Inai, Tetsuichiro; Iida, Hiroshi

    2014-04-01

    Tektins (TEKTs) are composed of a family of filament-forming proteins localized in cilia and flagella. Five types of mammalian TEKTs have been reported, all of which have been verified to be present in sperm flagella. TEKT2, which is indispensable for sperm structure, mobility, and fertilization, was present at the periphery of the outer dense fiber (ODF) in the sperm flagella. By yeast two-hybrid screening, we intended to isolate flagellar proteins that could interact with TEKT2, which resulted in the isolation of novel two genes from the mouse testis library, referred as a TEKT2-binding protein 1 (TEKT2BP1) and -protein 2 (TEKT2BP2). In this study, we characterized TEKT2BP1, which is registered as a coiled-coil domain-containing protein 172 (Ccdc172) in the latest database. RT-PCR analysis indicated that TEKT2BP1 was predominantly expressed in rat testis and that its expression was increased after 3 weeks of postnatal development. Immunocytochemical studies discovered that TEKT2BP1 localized in the middle piece of rat spermatozoa, predominantly concentrated at the mitochondria sheath of the flagella. We hypothesize that the TEKT2-TEKT2BP1 complex might be involved in the structural linkage between the ODF and mitochondria in the middle piece of the sperm flagella. PMID:24394471

  18. Sex-Specific Protection of Osteoarthritis by Deleting Cartilage Acid Protein 1

    PubMed Central

    Ge, Xianpeng; Ritter, Susan Y.; Tsang, Kelly; Shi, Ruirui; Takei, Kohtaro; Aliprantis, Antonios O.

    2016-01-01

    Cartilage acidic protein 1 (CRTAC1) was recently identified as an elevated protein in the synovial fluid of patients with osteoarthritis (OA) by a proteomic analysis. This gene is also upregulated in both human and mouse OA by transcriptomic analysis. The objective of this study was to characterize the expression and function of CRTAC1 in OA. Here, we first confirm the increase of CRTAC1 in cartilage biopsies from OA patients undergoing joint replacement by real-time PCR and immunohistochemistry. Furthermore, we report that proinflammatory cytokines interleukin-1beta and tumor necrosis factor alpha upregulate CRTAC1 expression in primary human articular chondrocytes and synovial fibroblasts. Genetic deletion of Crtac1 in mice significantly inhibited cartilage degradation, osteophyte formation and gait abnormalities of post-traumatic OA in female, but not male, animals undergoing the destabilization of medial meniscus (DMM) surgery. Taken together, CRTAC1 is upregulated in the osteoarthritic joint and directly induced in chondrocytes and synovial fibroblasts by pro-inflammatory cytokines. This molecule is necessary for the progression of OA in female mice after DMM surgery and thus represents a potential therapy for this prevalent disease, especially for women who demonstrate higher rates and more severe OA. PMID:27415616

  19. Molecular cloning and characterization of Babesia orientalis rhoptry-associated protein 1.

    PubMed

    Yu, Qian; He, Lan; Zhang, Wen-Jie; Cheng, Jian-Xi; Hu, Jin-Fang; Miao, Xiao-Yan; Huang, Yuan; Fan, Li-Zhe; Khan, Muhammad Kasib; Zhou, Yan-Qin; Hu, Min; Zhao, Jun-Long

    2014-10-15

    The rhoptry-associated protein 1 (RAP-1) gene of Babesia orientalis was obtained from a cDNA expression library by immunoscreening with B. orientalis-infected water buffalo sera. The nucleotide sequence of the cDNA was 1732 bp with an open reading frame (ORF) of 1434 bp, encoding a polypeptide of 478 amino acid residues with a predicted size of 52.5 kDa. The ORF was cloned into a pGEX-KG plasmid and subsequently expressed as a GST-fusion protein. The recombinant RAP-1 of B. orientalis (rBoRAP-1) was purified and evaluated as an antigen using Western blotting. The native BoRAP-1 was recognized by the antibodies raised in rabbits against rBoRAP-1. Strong immunofluorescence signals were observed in erythrocytes infected with B. orientalis. Phylogentic analysis revealed that B. orientalis fell into a Babesia clade and most closely related to Babesia bovis and Babesia ovis, which was similar to the previous reported trees based on 18S rRNA and HSP70 genes. The present study suggests that the BoRAP-1 might be a potential diagnostic antigen, and the RAP-1 genes can aid in the classification of Babesia and Theileria species.

  20. PDX-1 (pancreatic/duodenal homeobox-1 protein 1).

    PubMed

    Pedica, F; Beccari, S; Pedron, S; Montagna, L; Piccoli, P; Doglioni, C; Chilosi, M

    2014-12-01

    The homeodomain-containing transcription factor pancreatic duodenal homeobox 1 (PDX-1) plays a key role in pancreatic development and β-cell function. It is a major regulator of transcription in pancreatic cells, and transactivates the insulin gene by binding to a specific DNA motif in its promoter region. Glucose also regulates insulin gene transcription through PDX-1. It has been shown that PDX-1 is required for maintaining pancreatic islet functions by activating gene expression and has a dual role in pancreatic development. It initially contributes to pancreatic formation during embryogenesis and subsequently regulates the pancreatic islet cell physiology in mature islet cells. Because of this key role in the embryologic development of the pancreas, PDX-1 expression has been investigated in pancreatic cancer cell lines and human tumors. Moreover, a few reports have described expression of PDX-1 in other human neoplasms and have investigated its potential role in differential diagnosis, but data on normal human tissues are lacking. Understanding the molecular mechanisms of pancreas formation, and especially the function of PDX-1, may contribute to the improved treatment and prevention of debilitating diseases such as diabetes, insulinomas and pancreatic carcinomas. Nevertheless, further studies are needed concerning its possible application in routine practice.

  1. Overexpression of iron regulatory protein 1 suppresses growth of tumor xenografts.

    PubMed

    Chen, Guohua; Fillebeen, Carine; Wang, Jian; Pantopoulos, Kostas

    2007-04-01

    Iron is essential for proliferation of normal and neoplastic cells. Cellular iron uptake, utilization and storage are regulated by transcriptional and post-transcriptional mechanisms. We hypothesized that the disruption of iron homeostasis may modulate the growth properties of cancer cells. To address this, we employed H1299 lung cancer cells engineered for tetracycline-inducible overexpression of the post-transcriptional regulator iron regulatory protein 1 (IRP1). The induction of IRP1 (wild-type or the constitutive IRP1(C437S) mutant) did not affect the proliferation of the cells in culture, and only modestly reduced their efficiency to form colonies in soft agar. However, IRP1 dramatically impaired the capacity of the cells to form solid tumor xenografts in nude mice. Tumors derived from IRP1-transfectants were <20% in size compared to those from parent cells. IRP1 coordinately controls the expression of transferrin receptor 1 (TfR1) and ferritin by binding to iron-responsive elements (IREs) within their mRNAs. Biochemical analysis revealed high expression of epitope-tagged IRP1 in tumor tissue, which was associated with a profound increase in IRE-binding activity. As expected, this response misregulated iron metabolism by increasing TfR1 levels. Surprisingly, IRP1 failed to suppress ferritin expression and did not affect the levels of the iron transporter ferroportin. Our results show that the overexpression of IRP1 is associated with an apparent tumor suppressor phenotype and provide a direct regulatory link between the IRE/IRP system and cancer.

  2. Induction of the neural cell adhesion molecule and neuronal aggregation by osteogenic protein 1.

    PubMed Central

    Perides, G; Safran, R M; Rueger, D C; Charness, M E

    1992-01-01

    The neural cell adhesion molecule (N-CAM) plays a fundamental role in nervous system development and regeneration, yet the regulation of the expression of N-CAM in different brain regions has remained poorly understood. Osteogenic protein 1 (OP-1) is a member of the transforming growth factor beta superfamily that is expressed in the nervous system. Treatment of the neuroblastoma-glioma hybrid cell line NG108-15 for 1-4 days with recombinant human OP-1 (hOP-1) induced alterations in cell shape, formation of epithelioid sheets, and aggregation of cells into multilayered clusters. Immunofluorescence studies and Western blots demonstrated a striking differential induction of the three N-CAM isoforms in hOP-1-treated cells. hOP-1 caused a 6-fold up-regulation of the 140-kDa N-CAM, the isoform showing the highest constitutive expression, and a 29-fold up-regulation of the 180-kDa isoform. The 120-kDa isoform was not detected in control NG108-15 cells but was readily identified in hOP-1-treated cells. Incubation of NG108-15 cells with an antisense N-CAM oligonucleotide reduced the induction of N-CAM by hOP-1 and decreased the formation of multilayered cell aggregates. Anti-N-CAM monoclonal antibodies also diminished the formation of multilayered cell aggregates by hOP-1 and decreased cell-cell adhesion when hOP-1-treated NG108-15 cells were dispersed and replated. Thus, hOP-1 produces morphologic changes in NG108-15 cells, at least in part, by inducing N-CAM. These observations suggest that OP-1 or a homologue may participate in the regulation of N-CAM during nervous system development and regeneration. Images PMID:1438217

  3. Dentin Matrix Protein-1 Isoforms Promote Differential Cell Attachment and Migration*S⃞

    PubMed Central

    von Marschall, Zofia; Fisher, Larry W.

    2008-01-01

    Dentin matrix protein-1 (DMP1), bone sialoprotein (BSP), and osteopontin (OPN) are three SIBLINGs (small integrin-binding ligand, N-linked glycoproteins) co-expressed/secreted by skeletal and active ductal epithelial cells. Although etiological mechanisms remain unclear, DMP1 is the only one of these three genes currently known to have mutations resulting in human disease, and yet it remains the least studied. All three contain the highly conserved integrin-binding tripeptide, RGD, and experiments comparing the cell attachment and haptotactic migration-enhancing properties of DMP1 to BSP and OPN were performed using human skeletal (MG63 and primary dental pulp cells) and salivary gland (HSG) cells. Mutation of any SIBLING's RGD destroyed all attachment and migration activity. Using itsαVβ5 integrin, HSG cells attached to BSP but not to DMP1 or OPN. However, HSG cells could not migrate onto BSP in a modified Boyden chamber assay. Expression of αVβ3 integrin enhanced HSG attachment to DMP1 and OPN and promoted haptotactic migration onto all three proteins. Interchanging the first four coding exons or the conserved amino acids adjacent to the RGD of DMP1 with corresponding sequences of BSP did not enhance the ability of DMP1 to bindαVβ5. For αVβ3-expressing cells, intact DMP1, its BMP1-cleaved C-terminal fragment, and exon six lacking all post-translational modifications worked equally well but the proteoglycan isoform of DMP1 had greatly reduced ability for cell attachment and migration. The sequence specificity of the proposed BMP1-cleavage site of DMP1 was verified by mutation analysis. Direct comparison of the three proteins showed that cells discriminate among these SIBLINGs and among DMP1 isoforms. PMID:18819913

  4. Targeted disruption of fibrinogen like protein-1 accelerates hepatocellular carcinoma development

    SciTech Connect

    Nayeb-Hashemi, Hamed; Desai, Anal; Demchev, Valeriy; Bronson, Roderick T.; Hornick, Jason L.; Cohen, David E.; Ukomadu, Chinweike

    2015-09-18

    Fibrinogen like protein-1 (Fgl1) is a predominantly liver expressed protein that has been implicated as both a hepatoprotectant and a hepatocyte mitogen. Fgl1 expression is decreased in hepatocellular carcinoma (HCC) and its loss correlates with a poorly differentiated phenotype. To better elucidate the role of Fgl1 in hepatocarcinogenesis, we treated mice wild type or null for Fgl1 with diethyl nitrosamine and monitored for incidence of hepatocellular cancer. We find that mice lacking Fgl1 develop HCC at more than twice the rate of wild type mice. We show that hepatocellular cancers from Fgl1 null mice are molecularly distinct from those of the wild type mice. In tumors from Fgl1 null mice there is enhanced activation of Akt and downstream targets of the mammalian target of rapamycin (mTOR). In addition, there is paradoxical up regulation of putative hepatocellular cancer tumor suppressors; tripartite motif-containing protein 35 (Trim35) and tumor necrosis factor super family 10b (Tnfrsf10b). Taken together, these findings suggest that Fgl1 acts as a tumor suppressor in hepatocellular cancer through an Akt dependent mechanism and supports its role as a potential therapeutic target in HCC. - Highlights: • Fgl1 knockout mice (Fgl1KO) are more prone to carcinogen-induced liver cancer compared to wild type (WT) mates. • Tumors from the Fgl1KO are molecularly distinct with enhanced Akt and mTOR activity in comparison with Fgl1WT tumors. • Tumors from the Fgl1KO have enhanced expression of Trim35 and Tnfrsf10b, putative HCC tumor suppressors.

  5. Role of two single nucleotide polymorphisms in secreted frizzled related protein 1 and bladder cancer risk.

    PubMed

    Rogler, Anja; Hoja, Sabine; Socher, Eileen; Nolte, Elke; Wach, Sven; Wieland, Wolf; Hofstädter, Ferdinand; Goebell, Peter J; Wullich, Bernd; Hartmann, Arndt; Stoehr, Robert

    2013-01-01

    In this study, we determined the genotype distribution of two single nucleotide polymorphisms (SNPs) in secreted frizzled related protein 1 (SFRP1), rs3242 and rs921142, in a Caucasian bladder cancer case-control study. Allelic variants of the SNPs were determined using restriction fragment length polymorphism (RFLP) analysis and partly verified by sequencing analysis. Overall, DNA from 188 consecutive and 215 early-onset bladder cancer patients (≤45 years) as well as from 332 controls was investigated. Potential microRNA binding sites were determined for rs3242, and microRNA expression was analysed in cell lines and tumour specimens. We observed a remarkable distribution difference in rs3242 between bladder cancer patients and healthy controls (p=0.05). Additionally, we found a significant difference in genotype distribution (p=0.032), resulting from the difference of early-onset patients and the control group (p=0.007). The risk allele T showed increased frequency in the early-onset patient group (p=0.002). Genotype-dependent differences of microRNA binding capacity were predicted in SFRP1 mRNA for two microRNAs. Hsa-miR-3646 showed strong expression in cell lines and tumour tissue, whereas hsa-miR-603 exhibited weak expression. The rs921142 SNP showed no significant association with bladder cancer risk. This is the first study to describe an association of the SFRP1 SNP rs3242 and bladder cancer risk as well as the influence of rs3242 on genotype-dependent microRNA capacity on SFRP1 mRNA. The onset of bladder seems to be associated with the increased occurrence of the T-allele in rs3242. PMID:24133576

  6. Role of two single nucleotide polymorphisms in secreted frizzled related protein 1 and bladder cancer risk

    PubMed Central

    Rogler, Anja; Hoja, Sabine; Socher, Eileen; Nolte, Elke; Wach, Sven; Wieland, Wolf; Hofstädter, Ferdinand; Goebell, Peter J; Wullich, Bernd; Hartmann, Arndt; Stoehr, Robert

    2013-01-01

    In this study, we determined the genotype distribution of two single nucleotide polymorphisms (SNPs) in secreted frizzled related protein 1 (SFRP1), rs3242 and rs921142, in a Caucasian bladder cancer case-control study. Allelic variants of the SNPs were determined using restriction fragment length polymorphism (RFLP) analysis and partly verified by sequencing analysis. Overall, DNA from 188 consecutive and 215 early-onset bladder cancer patients (≤45 years) as well as from 332 controls was investigated. Potential microRNA binding sites were determined for rs3242, and microRNA expression was analysed in cell lines and tumour specimens. We observed a remarkable distribution difference in rs3242 between bladder cancer patients and healthy controls (p=0.05). Additionally, we found a significant difference in genotype distribution (p=0.032), resulting from the difference of early-onset patients and the control group (p=0.007). The risk allele T showed increased frequency in the early-onset patient group (p=0.002). Genotype-dependent differences of microRNA binding capacity were predicted in SFRP1 mRNA for two microRNAs. Hsa-miR-3646 showed strong expression in cell lines and tumour tissue, whereas hsa-miR-603 exhibited weak expression. The rs921142 SNP showed no significant association with bladder cancer risk. This is the first study to describe an association of the SFRP1 SNP rs3242 and bladder cancer risk as well as the influence of rs3242 on genotype-dependent microRNA capacity on SFRP1 mRNA. The onset of bladder seems to be associated with the increased occurrence of the T-allele in rs3242. PMID:24133576

  7. A role for tyrosinase-related protein 1 in 4-tert-butylphenol-induced toxicity in melanocytes: Implications for vitiligo.

    PubMed

    Manga, Prashiela; Sheyn, David; Yang, Fan; Sarangarajan, Rangaprasad; Boissy, Raymond E

    2006-11-01

    Vitiligo presents with depigmented cutaneous lesions following localized melanocyte death. Multiple factors contribute to cell death, including genetically determined susceptibility to trauma, and environmental factors, such as exposure to 4-tert-butylphenol (4-TBP). We demonstrate that 4-TBP induces oxidative stress that is more readily overcome by melanocytes from normally pigmented individuals than from two individuals with vitiligo. The antioxidant catalase selectively and significantly reduced death of melanocytes derived from two individuals with vitiligo, indicating a role for oxidative stress in vitiligo pathogenesis. In normal melanocytes, oxidative stress results in reduced expression of microphthalmia-associated transcription factor (MITF). Melanocyte-stimulating hormone-induced expression of MITF protein caused increased sensitivity to 4-TBP, whereas sensitivity of melanomas correlated with MITF expression. MITF stimulates melanin synthesis by up-regulating expression of melanogenic enzymes such as tyrosinase-related protein-1 (Tyrp1). Although melanin content per se did not affect sensitivity to 4-TBP, expression of Tyrp1 significantly increased sensitivity. Melanocytes and melanomas that express functional Tyrp1 were significantly more sensitive to 4-TBP than Tyrp1-null cells. Thus, normal melanocytes respond to 4-TBP by reducing expression of MITF and Tyrp1. We hypothesize that melanocytes in vitiligo demonstrate reduced ability to withstand oxidative stress due, partly, to a disruption in MITF regulation of Tyrp1.

  8. Influenza virus non-structural protein 1 (NS1) disrupts interferon signaling.

    PubMed

    Jia, Danlin; Rahbar, Ramtin; Chan, Renee W Y; Lee, Suki M Y; Chan, Michael C W; Wang, Ben Xuhao; Baker, Darren P; Sun, Bing; Peiris, J S Malik; Nicholls, John M; Fish, Eleanor N

    2010-01-01

    Type I interferons (IFNs) function as the first line of defense against viral infections by modulating cell growth, establishing an antiviral state and influencing the activation of various immune cells. Viruses such as influenza have developed mechanisms to evade this defense mechanism and during infection with influenza A viruses, the non-structural protein 1 (NS1) encoded by the virus genome suppresses induction of IFNs-α/β. Here we show that expression of avian H5N1 NS1 in HeLa cells leads to a block in IFN signaling. H5N1 NS1 reduces IFN-inducible tyrosine phosphorylation of STAT1, STAT2 and STAT3 and inhibits the nuclear translocation of phospho-STAT2 and the formation of IFN-inducible STAT1:1-, STAT1:3- and STAT3:3- DNA complexes. Inhibition of IFN-inducible STAT signaling by NS1 in HeLa cells is, in part, a consequence of NS1-mediated inhibition of expression of the IFN receptor subunit, IFNAR1. In support of this NS1-mediated inhibition, we observed a reduction in expression of ifnar1 in ex vivo human non-tumor lung tissues infected with H5N1 and H1N1 viruses. Moreover, H1N1 and H5N1 virus infection of human monocyte-derived macrophages led to inhibition of both ifnar1 and ifnar2 expression. In addition, NS1 expression induces up-regulation of the JAK/STAT inhibitors, SOCS1 and SOCS3. By contrast, treatment of ex vivo human lung tissues with IFN-α results in the up-regulation of a number of IFN-stimulated genes and inhibits both H5N1 and H1N1 virus replication. The data suggest that NS1 can directly interfere with IFN signaling to enhance viral replication, but that treatment with IFN can nevertheless override these inhibitory effects to block H5N1 and H1N1 virus infections.

  9. Influenza Virus Non-Structural Protein 1 (NS1) Disrupts Interferon Signaling

    PubMed Central

    Jia, Danlin; Rahbar, Ramtin; Chan, Renee W. Y.; Lee, Suki M. Y.; Chan, Michael C. W.; Wang, Ben Xuhao; Baker, Darren P.; Sun, Bing; Peiris, J. S. Malik; Nicholls, John M.; Fish, Eleanor N.

    2010-01-01

    Type I interferons (IFNs) function as the first line of defense against viral infections by modulating cell growth, establishing an antiviral state and influencing the activation of various immune cells. Viruses such as influenza have developed mechanisms to evade this defense mechanism and during infection with influenza A viruses, the non-structural protein 1 (NS1) encoded by the virus genome suppresses induction of IFNs-α/β. Here we show that expression of avian H5N1 NS1 in HeLa cells leads to a block in IFN signaling. H5N1 NS1 reduces IFN-inducible tyrosine phosphorylation of STAT1, STAT2 and STAT3 and inhibits the nuclear translocation of phospho-STAT2 and the formation of IFN-inducible STAT1:1-, STAT1:3- and STAT3:3- DNA complexes. Inhibition of IFN-inducible STAT signaling by NS1 in HeLa cells is, in part, a consequence of NS1-mediated inhibition of expression of the IFN receptor subunit, IFNAR1. In support of this NS1-mediated inhibition, we observed a reduction in expression of ifnar1 in ex vivo human non-tumor lung tissues infected with H5N1 and H1N1 viruses. Moreover, H1N1 and H5N1 virus infection of human monocyte-derived macrophages led to inhibition of both ifnar1 and ifnar2 expression. In addition, NS1 expression induces up-regulation of the JAK/STAT inhibitors, SOCS1 and SOCS3. By contrast, treatment of ex vivo human lung tissues with IFN-α results in the up-regulation of a number of IFN-stimulated genes and inhibits both H5N1 and H1N1 virus replication. The data suggest that NS1 can directly interfere with IFN signaling to enhance viral replication, but that treatment with IFN can nevertheless override these inhibitory effects to block H5N1 and H1N1 virus infections. PMID:21085662

  10. Far Upstream Element-Binding Protein 1 Binds the 3' Untranslated Region of PKD2 and Suppresses Its Translation.

    PubMed

    Zheng, Wang; Shen, Fan; Hu, Ruikun; Roy, Birbickram; Yang, JungWoo; Wang, Qian; Zhang, Fan; King, Jennifer C; Sergi, Consolato; Liu, Song-Mei; Cordat, Emmanuelle; Tang, Jingfeng; Cao, Ying; Ali, Declan; Chen, Xing-Zhen

    2016-09-01

    Autosomal dominant polycystic kidney disease pathogenesis can be recapitulated in animal models by gene mutations in or dosage alterations of polycystic kidney disease 1 (PKD1) or PKD2, demonstrating that too much and too little PKD1/PKD2 are both pathogenic. Gene dosage manipulation has become an appealing approach by which to compensate for loss or gain of gene function, but the mechanisms controlling PKD2 expression remain incompletely characterized. In this study, using cultured mammalian cells and dual-luciferase assays, we found that the 3' untranslated region (3'UTR) of PKD2 mRNA inhibits luciferase protein expression. We then identified nucleotides 691-1044, which we called 3FI, as the 3'UTR fragment necessary for repressing the expression of luciferase or PKD2 in this system. Using a pull-down assay and mass spectrometry we identified far upstream element-binding protein 1 (FUBP1) as a 3FI-binding protein. In vitro overexpression of FUBP1 inhibited the expression of PKD2 protein but not mRNA. In embryonic zebrafish, FUBP1 knockdown (KD) by morpholino injection increased PKD2 expression and alleviated fish tail curling caused by morpholino-mediated KD of PKD2. Conversely, FUBP1 overexpression by mRNA injection significantly increased pronephric cyst occurrence and tail curling in zebrafish embryos. Furthermore, FUBP1 binds directly to eukaryotic translation initiation factor 4E-binding protein 1, indicating a link to the translation initiation complex. These results show that FUBP1 binds 3FI in the PKD2 3'UTR to inhibit PKD2 translation, regulating zebrafish disease phenotypes associated with PKD2 KD.

  11. Far Upstream Element-Binding Protein 1 Binds the 3' Untranslated Region of PKD2 and Suppresses Its Translation.

    PubMed

    Zheng, Wang; Shen, Fan; Hu, Ruikun; Roy, Birbickram; Yang, JungWoo; Wang, Qian; Zhang, Fan; King, Jennifer C; Sergi, Consolato; Liu, Song-Mei; Cordat, Emmanuelle; Tang, Jingfeng; Cao, Ying; Ali, Declan; Chen, Xing-Zhen

    2016-09-01

    Autosomal dominant polycystic kidney disease pathogenesis can be recapitulated in animal models by gene mutations in or dosage alterations of polycystic kidney disease 1 (PKD1) or PKD2, demonstrating that too much and too little PKD1/PKD2 are both pathogenic. Gene dosage manipulation has become an appealing approach by which to compensate for loss or gain of gene function, but the mechanisms controlling PKD2 expression remain incompletely characterized. In this study, using cultured mammalian cells and dual-luciferase assays, we found that the 3' untranslated region (3'UTR) of PKD2 mRNA inhibits luciferase protein expression. We then identified nucleotides 691-1044, which we called 3FI, as the 3'UTR fragment necessary for repressing the expression of luciferase or PKD2 in this system. Using a pull-down assay and mass spectrometry we identified far upstream element-binding protein 1 (FUBP1) as a 3FI-binding protein. In vitro overexpression of FUBP1 inhibited the expression of PKD2 protein but not mRNA. In embryonic zebrafish, FUBP1 knockdown (KD) by morpholino injection increased PKD2 expression and alleviated fish tail curling caused by morpholino-mediated KD of PKD2. Conversely, FUBP1 overexpression by mRNA injection significantly increased pronephric cyst occurrence and tail curling in zebrafish embryos. Furthermore, FUBP1 binds directly to eukaryotic translation initiation factor 4E-binding protein 1, indicating a link to the translation initiation complex. These results show that FUBP1 binds 3FI in the PKD2 3'UTR to inhibit PKD2 translation, regulating zebrafish disease phenotypes associated with PKD2 KD. PMID:26839368

  12. Preferential expression of functional IL-17R in glioma stem cells: potential role in self-renewal

    PubMed Central

    Parajuli, Prahlad; Anand, Rohit; Mandalaparty, Chandramouli; Suryadevara, Raviteja; Sriranga, Preethi U.; Michelhaugh, Sharon K.; Cazacu, Simona; Finniss, Susan; Thakur, Archana; Lum, Lawrence G.; Schalk, Dana; Brodie, Chaya; Mittal, Sandeep

    2016-01-01

    Gliomas are the most common primary brain tumor and one of the most lethal solid tumors. Mechanistic studies into identification of novel biomarkers are needed to develop new therapeutic strategies for this deadly disease. The objective for this study was to explore the potential direct impact of IL-17−IL-17R interaction in gliomas. Immunohistochemistry and flow cytometry analysis of 12 tumor samples obtained from patients with high grade gliomas revealed that a considerable population (2–19%) of cells in all malignant gliomas expressed IL-17RA, with remarkable co-expression of the glioma stem cell (GSC) markers CD133, Nestin, and Sox2. IL-17 enhanced the self-renewal of GSCs as determined by proliferation and Matrigel® colony assays. IL-17 also induced cytokine/chemokine (IL-6, IL-8, interferon-γ-inducible protein [IP-10], and monocyte chemoattractant protein-1 [MCP-1]) secretion in GSCs, which were differentially blocked by antibodies against IL-17R and IL-6R. Western blot analysis showed that IL-17 modulated the activity of signal transducer and activator of transcription 3 (STAT3), nuclear factor κ-light-chain-enhancer of activated B cells (NF-κB), glycogen synthase kinase-3β (GSK-3β) and β-catenin in GSCs. While IL-17R-mediated secretion of IL-6 and IL-8 were significantly blocked by inhibitors of NF-κB and STAT3; NF-κB inhibitor was more potent than STAT3 inhibitor in blocking IL-17-induced MCP-1 secretion. Overall, our results suggest that IL-17–IL-17R interaction in GSCs induces an autocrine/paracrine cytokine feedback loop, which may provide an important signaling component for maintenance/self-renewal of GSCs via constitutive activation of both NF-κB and STAT3. The results also strongly implicate IL-17R as an important functional biomarker for therapeutic targeting of GSCs. PMID:26755664

  13. Behavioral analysis of the huntingtin-associated protein 1 ortholog trak-1 in Caenorhabditis elegans.

    PubMed

    Norflus, Fran; Bu, Jingnan; Guyton, Evon; Gutekunst, Claire-Anne

    2016-09-01

    The precise role of huntingtin-associated protein 1 (HAP1) is not known, but studies have shown that it is important for early development and survival. A Caenorhabditis elegans ortholog of HAP1, T27A3.1 (also called trak-1), has been found and is expressed in a subset of neurons. Potential behavioral functions of three knockout lines of T27A3.1 were examined. From its suspected role in mice we hypothesize that T27A3.1 might be involved in egg hatching and early growth, mechanosensation, chemosensation, sensitivity to osmolarity, and synaptic transmission. Our studies show that the knockout worms are significantly different from the wild-type (WT) worms only in the synaptic transmission test, which was measured by adding aldicarb, an acetylcholinesterase inhibitor. The change in function was determined by measuring the number of worms paralyzed. However, when the T27A3.1 worms were tested for egg hatching and early growth, mechanosensation, chemosensation, and sensitivity to osmolarity, there were no significant differences between the knockout and WT worms. © 2016 Wiley Periodicals, Inc. PMID:27319755

  14. NAP-1, Nucleosome assembly protein 1, a histone chaperone involved in Drosophila telomeres.

    PubMed

    López-Panadès, Elisenda; Casacuberta, Elena

    2016-03-01

    Telomere elongation is a function that all eukaryote cells must accomplish in order to guarantee, first, the stability of the end of the chromosomes and second, to protect the genetic information from the inevitable terminal erosion. The targeted transposition of the telomere transposons HeT-A, TART and TAHRE perform this function in Drosophila, while the telomerase mechanism elongates the telomeres in most eukaryotes. In order to integrate telomere maintenance together with cell cycle and metabolism, different components of the cell interact, regulate, and control the proteins involved in telomere elongation. Different partners of the telomerase mechanism have already been described, but in contrast, very few proteins have been related with assisting the telomere transposons of Drosophila. Here, we describe for the first time, the implication of NAP-1 (Nucleosome assembly protein 1), a histone chaperone that has been involved in nuclear transport, transcription regulation, and chromatin remodeling, in telomere biology. We find that Nap-1 and HeT-A Gag, one of the major components of the Drosophila telomeres, are part of the same protein complex. We also demonstrate that their close interaction is necessary to guarantee telomere stability in dividing cells. We further show that NAP-1 regulates the transcription of the HeT-A retrotransposon, pointing to a positive regulatory role of NAP-1 in telomere expression. All these results facilitate the understanding of the transposon telomere maintenance mechanism, as well as the integration of telomere biology with the rest of the cell metabolism.

  15. Regulation of muscle contraction by Drebrin-like protein 1 probed by atomic force microscopy

    NASA Astrophysics Data System (ADS)

    Garces, Renata; Butkevich, Eugenia; Platen, Mitja; Schmidt, Christoph F.; Biophysics Team

    Sarcomeres are the fundamental contractile units of striated muscle cells. They are composed of a variety of structural and regulatory proteins functioning in a precisely orchestrated fashion to enable coordinated force generation in striated muscles. Recently, we have identified a C. elegans drebrin-like protein 1 (DBN-1) as a novel sarcomere component, which stabilizes actin filaments during muscle contraction. To further characterize the function of DBN-1 in muscle cells, we generated a new dbn-1 loss-of-function allele. Absence of DBN-1 resulted in a unique worm movement phenotype, characterized by hyper-bending. It is not clear yet if DBN-1 acts to enhance or reduce the capacity for contraction. We present here an experimental mechanical study on C. elegans muscle mechanics. We measured the stiffness of the worm by indenting living C. eleganswith a micron-sized sphere adhered to the cantilever of an atomic force microscope (AFM). Modeling the worm as a pressurized elastic shell allows us to monitor the axial tension in the muscle through the measured stiffness. We compared responses of wild-type and mutant C. elegans in which DBN-1 is not expressed..

  16. VIP1 is very important/interesting protein 1 regulating touch responses of Arabidopsis

    PubMed Central

    Tsugama, Daisuke; Liu, Shenkui; Takano, Tetsuo

    2016-01-01

    ABSTRACT VIP1 (VIRE2-INTERACTING PROTEIN 1) is a bZIP transcription factor in Arabidopsis thaliana. VIP1 and its close homologs (i.e., Arabidopsis group I bZIP proteins) are present in the cytoplasm under steady conditions, but are transiently localized to the nucleus when cells are exposed to hypo-osmotic conditions, which mimic mechanical stimuli such as touch. Recently we have reported that overexpression of a repression domain-fused form of VIP1 represses the expression of some touch-responsive genes, changes structures and/or local auxin responses of the root cap cells, and enhances the touch-induced root waving. This raises the possibility that VIP1 suppresses touch-induced responses. VIP1 should be useful to further characterize touch responses of plants. Here we discuss 2 seemingly interesting perspectives about VIP1: (1) What factors are involved in regulating the nuclear localization of VIP1?; (2) What can be done to further characterize the physiological functions of VIP1 and other Arabidopsis group I bZIP proteins? PMID:27171129

  17. Obesity-associated mouse adipose stem cell secretion of monocyte chemotactic protein-1.

    PubMed

    Zhou, Hui Ren; Kim, Eun-Kyoung; Kim, Hyojung; Claycombe, Kate J

    2007-11-01

    Studies showed that monocyte chemotactic protein-1 (MCP-1) concentrations are increased in obesity. In our current study, we demonstrate that plasma MCP-1 level in leptin-deficient ob/ob mice is significantly higher than in lean mice. Furthermore, we determined that basal adipose tissue MCP-1 mRNA levels are significantly higher in ob/ob mice compared with lean mice. To determine the mechanisms underlying obesity-associated increases in plasma and adipose tissue MCP-1 levels, we determined adipose tissue cell type sources of MCP-1 production. Our data show that adipose tissue stem cells (CD34(+)), macrophages (F4/80(+)), and stromal vascular fraction (SVF) cells express significantly higher levels of MCP-1 compared with adipocytes under both basal and lipopolysaccharide (LPS)-stimulated conditions. Furthermore, basal and LPS-induced MCP-1 secretion levels were the same for both adipose F4/80(+) and CD34(+) cells, whereas adipose CD34(+) cells have twofold higher cell numbers (30% of total SVF cells) compared with F4/80(+) macrophages (15%). Our data also show that CD34(+) cells from visceral adipose tissue depots secrete significantly higher levels of MCP-1 ex vivo when compared with CD34(+) cells from subcutaneous adipose tissue depots. Taken together, our data suggest that adipose CD34(+) stem cells may play an important role in obesity-associated increases in plasma MCP-1 levels.

  18. Adherens junctional associated protein-1: a novel 1p36 tumor suppressor candidate in gliomas (Review).

    PubMed

    Zeng, Liang; Fee, Brian E; Rivas, Miriam V; Lin, James; Adamson, David Cory

    2014-07-01

    In a broad range of human cancers 1p36 has been a mutational hotspot which strongly suggests that the loss of tumor suppressor activity maps to this genomic region during tumorigenesis. Adherens junctional associated protein-1 (AJAP1; also known as Shrew1) was initially discovered as a novel transmembrane protein of adherent junctions in epithelial cells. Gene profiling showed AJAP1 on 1p36 is frequently lost or epigenetically silenced. AJAP1 may affect cell motility, migration, invasion and proliferation by unclear mechanisms. AJAP1 may be translocated to the nucleus, via its interaction with β-catenin complexes, where it can regulate gene transcription, then possibly have a potent impact on cell cycling and apoptosis. Significantly, loss of AJAP1 expression predicts poor clinical outcome of patients with malignant gliomas such as GBM and it may serve as a promising tumor suppressor-related target. In this review, we summarize and discuss current knowledge that may identify AJAP1 as a tumor suppressor in gliomas.

  19. Blockade of Vascular Adhesion Protein-1 Inhibits Lymphocyte Infiltration in Rat Liver Allograft Rejection

    PubMed Central

    Martelius, Timi; Salaspuro, Ville; Salmi, Marko; Krogerus, Leena; Höckerstedt, Krister; Jalkanen, Sirpa; Lautenschlager, Irmeli

    2004-01-01

    Vascular adhesion protein-1 (VAP-1) has been shown to mediate lymphocyte adhesion to endothelia at sites of inflammation, but its functional role in vivo has not been tested in any rodent model. Here we report the effects of VAP-1 blockade on rat liver allograft rejection. BN recipients of PVG liver allografts (known to develop acute rejection by day 7) were treated with 2 mg/kg anti-VAP-1 (a new anti-rat VAP-1 mAb 174–5) or isotype-matched irrelevant antibody (NS1) every other day (n = 6/group) and one group with anti-VAP-1 2 mg/kg daily (n = 7). On day 7, samples were collected for transplant aspiration cytology, histology, and immunohistochemistry. Lymphocyte infiltration to the graft was clearly affected by VAP-blockade. The total inflammation, mainly the number of active lymphoid cells, in transplant aspiration cytology was significantly decreased in animals treated with anti-VAP-1 (4.7 ± 1.0 and 2.4 ± 1.0 corrected increment units, respectively) compared to control (6.6 ± 1.0) (P < 0.05). In histology, the intensity of portal inflammation was significantly decreased (P < 0.05). The amount of T cells expressing activation markers diminished. This is the first demonstration in any prolonged in vivo model that VAP-1 plays an important role in lymphocyte infiltration to sites of inflammation, and, in particular, liver allograft rejection. PMID:15579442

  20. The role of high mobility group box chromosomal protein 1 in rheumatoid arthritis.

    PubMed

    Chen, Yu; Sun, Wei; Gao, Rongfen; Su, Yuying; Umehara, Hisanori; Dong, Lingli; Gong, Feili

    2013-10-01

    High mobility group box chromosomal protein 1 (HMGB1) is a ubiquitous highly conserved single polypeptide in all mammal eukaryotic cells. HMGB1 exists mainly within the nucleus and acts as a DNA chaperone. When passively released from necrotic cells or actively secreted into the extracellular milieu in response to appropriate signal stimulation, HMGB1 binds to related cell signal transduction receptors, such as RAGE, TLR2, TLR4 and TLR9, and becomes a proinflammatory cytokine that participates in the development and progression of many diseases, such as arthritis, acute lung injury, graft rejection immune response, ischaemia reperfusion injury and autoimmune liver damage. Only a small amount of HMGB1 release occurs during apoptosis, which undergoes oxidative modification on Cys106 and delivers tolerogenic signals to suppress immune activity. This review focuses on the important role of HMGB1 in the pathogenesis of RA, mainly manifested as the aberrant expression of HMGB1 in the serum, SF and synovial tissues; overexpression of signal transduction receptors; abnormal regulation of osteoclastogenesis and bone remodelling leading to the destruction of cartilage and bones. Intervention with HMGB1 may ameliorate the pathogenic conditions and attenuate disease progression of RA. Therefore administration of an HMGB1 inhibitor may represent a promising clinical approach for the treatment of RA.

  1. Activator protein 1 (Fos/Jun) functions in inflammatory bone and skin disease.

    PubMed

    Zenz, Rainer; Eferl, Robert; Scheinecker, Clemens; Redlich, Kurt; Smolen, Josef; Schonthaler, Helia B; Kenner, Lukas; Tschachler, Erwin; Wagner, Erwin F

    2008-01-01

    Activator protein 1 (AP-1) (Fos/Jun) is a transcriptional regulator composed of members of the Fos and Jun families of DNA binding proteins. The functions of AP-1 were initially studied in mouse development as well as in the whole organism through conventional transgenic approaches, but also by gene targeting using knockout strategies. The importance of AP-1 proteins in disease pathways including the inflammatory response became fully apparent through conditional mutagenesis in mice, in particular when employing gene inactivation in a tissue-specific and inducible fashion. Besides the well-documented roles of Fos and Jun proteins in oncogenesis, where these genes can function both as tumor promoters or tumor suppressors, AP-1 proteins are being recognized as regulators of bone and immune cells, a research area termed osteoimmunology. In the present article, we review recent data regarding the functions of AP-1 as a regulator of cytokine expression and an important modulator in inflammatory diseases such as rheumatoid arthritis, psoriasis and psoriatic arthritis. These new data provide a better molecular understanding of disease pathways and should pave the road for the discovery of new targets for therapeutic applications.

  2. Heterochromatin protein 1α: a hallmark of cell proliferation relevant to clinical oncology

    PubMed Central

    De Koning, Leanne; Savignoni, Alexia; Boumendil, Charlène; Rehman, Haniya; Asselain, Bernard; Sastre-Garau, Xavier; Almouzni, Geneviève

    2009-01-01

    Mammalian cells contain three closely related heterochromatin protein 1 (HP1) isoforms, HP1α, β and γ, which, by analogy to their unique counterpart in Schizosaccharomyces pombe, have been implicated in gene silencing, genome stability and chromosome segregation. However, the individual importance of each isoform during normal cell cycle and disease has remained an unresolved issue. Here, we reveal that HP1α shows a proliferation-dependent regulation, which neither HP1β nor γ display. During transient cell cycle exit, the HP1α mRNA and protein levels diminish. Transient depletion of HP1α, but not HP1β or γ, in tumoural and primary human cells leads to defects in chromosome segregation. Notably, analysis of an annotated collection of samples derived from carcinomas reveals an overexpression of HP1α mRNA and protein, which correlates with clinical data and disease outcome. Our results unveil a specific expression pattern for the HP1α isoform, suggesting a unique function related to cell division and tumour growth. The overexpression of HP1α constitutes a new example of a potential epigenetic contribution to tumourigenesis that is of clinical interest for cancer prognosis. PMID:20049717

  3. The Epigenetic Regulator CXXC Finger Protein 1 is Essential for Murine Hematopoiesis

    PubMed Central

    Dobrota, Erika; Tate, Courtney; Lee, Jeong-Heon; Khan, Shehnaz; Haneline, Laura; HogenEsch, Harm; Skalnik, David G.

    2014-01-01

    CXXC finger protein 1 (Cfp1), encoded by the Cxxc1 gene, binds to DNA sequences containing an unmethylated CpG dinucleotide and is an epigenetic regulator of both cytosine and histone methylation. Cxxc1-null mouse embryos fail to gastrulate, and Cxxc1-null embryonic stem cells are viable but cannot differentiate, suggesting that Cfp1 is required for chromatin remodeling associated with stem cell differentiation and embryogenesis. Mice homozygous for a conditional Cxxc1 deletion allele and carrying the inducible Mx1-Cre transgene were generated to assess Cfp1 function in adult animals. Induction of Cre expression in adult animals led to Cfp1 depletion in hematopoietic cells, a failure of hematopoiesis with a nearly complete loss of lineage-committed progenitors and mature cells, elevated levels of apoptosis, and death within two weeks. A similar pathology resulted following transplantation of conditional Cxxc1 bone marrow cells into wild type recipients, demonstrating this phenotype is intrinsic to Cfp1 function within bone marrow cells. Remarkably, the Lin−Sca-1+c-Kit+ population of cells in the bone marrow, which is enriched for hematopoietic stem cells and multi-potential progenitor cells, persists and expands in the absence of Cfp1 during this time frame. Thus, Cfp1 is necessary for hematopoietic stem and multi-potential progenitor cell function and for the developmental potential of differentiating hematopoietic cells. PMID:25470594

  4. DNA Methyltransferase protein synthesis is reduced in CXXC finger protein 1-deficient embryonic stem cells.

    PubMed

    Butler, Jill S; Palam, Lakshmi R; Tate, Courtney M; Sanford, Jeremy R; Wek, Ronald C; Skalnik, David G

    2009-05-01

    CXXC finger protein 1 (CFP1) binds to unmethylated CpG dinucleotides and is required for embryogenesis. CFP1 is also a component of the Setd1A and Setd1B histone H3K4 methyltransferase complexes. Murine embryonic stem (ES) cells lacking CFP1 fail to differentiate, and exhibit a 70% reduction in global genomic cytosine methylation and a 50% reduction in DNA methyltransferase (DNMT1) protein and activity. This study investigated the underlying mechanism for reduced DNMT1 expression in CFP1-deficient ES cells. DNMT1 transcript levels were significantly elevated in ES cells lacking CFP1, despite the observed reduction in DNMT1 protein levels. To address the posttranscriptional mechanisms by which CFP1 regulates DNMT1 protein activity, pulse/chase analyses were carried out, demonstrating a modest reduction in DNMT1 protein half-life in CFP1-deficient ES cells. Additionally, global protein synthesis was decreased in ES cells lacking CFP1, contributing to a reduction in the synthesis of DNMT1 protein. ES cells lacking CFP1 were found to contain elevated levels of phosphorylated eIF2alpha, and an accompanying reduction in translation initiation as revealed by a lower level of polyribosomes. These results reveal a novel role for CFP1 in the regulation of translation initiation, and indicate that loss of CFP1 function leads to decreased DNMT1 protein synthesis and half-life. PMID:19388845

  5. Novel alternatively spliced endoplasmic reticulum retention signal in the cytoplasmic loop of Proteolipid Protein-1.

    PubMed

    Southwood, Cherie; Olson, Kevin; Wu, Chia-Yen; Gow, Alexander

    2007-02-15

    Increased awareness about the importance of protein folding and trafficking to the etiology of gain-of-function diseases has driven extensive efforts to understand the cell and molecular biology underlying the life cycle of normal secretory pathway proteins and the detrimental effects of abnormal proteins. In this regard, the quality-control machinery in the endoplasmic reticulum (ER) has emerged as a major mechanism by which cells ensure that secreted and transmembrane proteins either adopt stable secondary, tertiary, and quaternary structures or are retained in the ER and degraded. Here we examine cellular and molecular aspects of ER retention in transfected fibroblasts expressing missense mutations in the Proteolipid Protein-1 (PLP1) gene that cause mild or severe forms of neurodegenerative disease in humans. Mild mutations cause protein retention in the ER that is partially dependent on the presence of a cytoplasmically exposed heptapeptide, KGRGSRG. In contrast, retention associated with severe mutations occurs independently of this peptide. Accordingly, the function of this novel heptapeptide has a significant impact on pathogenesis and provides new insight into the functions of the two splice isoforms encoded by the PLP1 gene, PLP1 and DM-20. PMID:17171701

  6. Fibrinogen-like protein 1, a hepatocyte derived protein is an acute phase reactant

    SciTech Connect

    Liu Zhilin; Ukomadu, Chinweike

    2008-01-25

    Fibrinogen-like protein 1 (FGL1) is a hepatocyte derived protein that is upregulated in regenerating rodent livers following partial hepatectomy. It has been implicated as a mitogen for liver cell proliferation. In this study, we show that recombinant human IL-6 induces FGL1 expression in Hep G2 cells in a pattern similar to those of acute phase reactants. Following induction of acute inflammation in rats by subcutaneous injection of turpentine oil, serum FGL1 levels are also enhanced. Although, a recent report suggests that FGL1 associates almost exclusively with the fibrin matrix, we report here that approximately 20% of the total plasma FGL1 remains free. The enhancement of FGL1 levels in vitro by IL-6 and its induction after turpentine oil injection suggest that it is an acute phase reactant. Its presence in bound and free forms in the blood also implies biological roles that extend beyond the proposed autocrine effect it has on hepatocytes during regeneration.

  7. The role of vitamin D3 upregulated protein 1 in thioacetamide-induced mouse hepatotoxicity

    SciTech Connect

    Kwon, Hyo-Jung; Lim, Jong-Hwan; Han, Jong-Tak; Lee, Sae-Bhom; Yoon, Won-Kee; Nam, Ki-Hoan; Choi, In-Pyo; Kim, Dae-Yong; Won, Young-Suk; Kim, Hyoung-Chin

    2010-11-01

    Thioacetamide (TA) is a commonly used drug that can trigger acute hepatic failure (AHF) through generation of oxidative stress. Vitamin D3 upregulated protein 1 (VDUP1) is an endogenous inhibitor of thioredoxin, a ubiquitous thiol oxidoreductase, that regulates cellular redox status. In this study, we investigated the role of VDUP1 in AHF using a TA-induced liver injury model. VDUP1 knockout (KO) and wild-type (WT) mice were subjected to a single intraperitoneal TA injection, and various parameters of hepatic injury were assessed. VDUP1 KO mice displayed a significantly higher survival rate, lower serum alanine aminotransferase and aspartate aminotransferase levels, and less hepatic damage, compared to WT mice. In addition, induction of apoptosis was decreased in VDUP1 KO mice, with the alteration of caspase-3 and -9 activities, Bax-to-Bcl-2 expression ratios, and mitogen activated protein kinase (MAPK) signaling pathway. Importantly, analysis of TA bioactivation revealed lower plasma clearance of TA and covalent binding of [{sup 14}C]TA to liver macromolecules in VDUP1 KO mice. Furthermore, the level of oxidative stress was significantly less in VDUP1 KO mice than in their WT counterparts, as evident from lipid peroxidation assay. These results collectively indicate that VDUP1 deficiency protects against TA-induced acute liver injury via lower bioactivation of TA and antioxidant effects.

  8. Oligouridylate Binding Protein 1b Plays an Integral Role in Plant Heat Stress Tolerance.

    PubMed

    Nguyen, Cam Chau; Nakaminami, Kentaro; Matsui, Akihiro; Kobayashi, Shuhei; Kurihara, Yukio; Toyooka, Kiminori; Tanaka, Maho; Seki, Motoaki

    2016-01-01

    Stress granules (SGs), which are formed in the plant cytoplasm under stress conditions, are transient dynamic sites (particles) for mRNA storage. SGs are actively involved in protecting mRNAs from degradation. Oligouridylate binding protein 1b (UBP1b) is a component of SGs. The formation of microscopically visible cytoplasmic foci, referred to as UBP1b SG, was induced by heat treatment in UBP1b-overexpressing Arabidopsis plants (UBP1b-ox). A detailed understanding of the function of UBP1b, however, is still not clear. UBP1b-ox plants displayed increased heat tolerance, relative to control plants, while ubp1b mutants were more sensitive to heat stress than control plants. Microarray analysis identified 117 genes whose expression was heat-inducible and higher in the UBP1b-ox plants. RNA decay analysis was performed using cordycepin, a transcriptional inhibitor. In order to determine if those genes serve as targets of UBP1b, the rate of RNA degradation of a DnaJ heat shock protein and a stress-associated protein (AtSAP3) in UBP1b-ox plants was slower than in control plants; indicating that the mRNAs of these genes were protected within the UBP1b SG granule. Collectively, these data demonstrate that UBP1b plays an integral role in heat stress tolerance in plants. PMID:27379136

  9. β-lapachone suppresses the proliferation of human malignant melanoma cells by targeting specificity protein 1.

    PubMed

    Bang, Woong; Jeon, Young-Joo; Cho, Jin Hyoung; Lee, Ra Ham; Park, Seon-Min; Shin, Jae-Cheon; Choi, Nag-Jin; Choi, Yung Hyun; Cho, Jung-Jae; Seo, Jae-Min; Lee, Seung-Yeop; Shim, Jung-Hyun; Chae, Jung-Il

    2016-02-01

    β-lapachone (β-lap), a novel natural quinone derived from the bark of the Pink trumpet tree (Tabebuia avellanedae) has been demonstrated to have anticancer activity. In this study, we investigated whether β-lap exhibits anti-proliferative effects on two human malignant melanoma (HMM) cell lines, G361 and SK-MEL-28. The effects of β-lap on the HMM cell lines were investigated using 3-(4,5-dimethylthiazol-2-yl)‑5-(3-carboxymethoxyphenyl)‑2-(4-sulfophenyl-2H-tetrazolium (MTS) assay, 4',6-diamidino-2-phenylindole (DAPI) staining, Annexin V and Dead cell assay, mitochondrial membrane potential (MMP) assay and western blot analysis. We demonstrated that β-lap significantly induced apoptosis and suppressed cell viability in the HMM cells. Intriguingly, the transcription factor specificity protein 1 (Sp1) was significantly downregulated by β-lap in a dose- and time-dependent manner. Furthermore, β-lap modulated the protein expression level of the Sp1 regulatory genes including cell cycle regulatory proteins and apoptosis-associated proteins. Taken together, our findings indicated that β-lap modulates Sp1 transactivation and induces apoptotic cell death through the regulation of cell cycle- and apoptosis-associated proteins. Thus, β-lap may be used as a promising anticancer drug for cancer prevention and may improve the clinical outcome of patients with cancer. PMID:26718788

  10. Role of Yes-associated protein 1 in gliomas: pathologic and therapeutic aspects.

    PubMed

    Liu, Yong-Chang; Wang, Yan-zhou

    2015-04-01

    The activation of proline-rich phosphoprotein Yes-associated protein 1 (YAP1) possesses a possible link between stem/progenitor cells, organ size, and cancer. YAP1 has been indicated as an oncoprotein, and overexpression of YAP1 is reported in many human brain tumors, including infiltrating gliomas. During normal brain development, the neurofibromatosis 2 (NF2) protein suppresses YAP1 activity in neural progenitor cells to promote guidepost cell differentiation, but loss of NF2 causes elevating YAP1 activity in midline neural progenitors, which disrupts guidepost formation. Overexpression of endogenous CD44 (cancer stem cell marker) promotes phosphorylation/inactivation of NF2, and upregulates YAP1 expression and leads to cancer cell resistance in glioblastoma. The hippo pathway is also related to the YAP1 action. However, the mechanism of YAP1 action in glioma is still far from clear understanding. Advances in clinical management based on an improved understanding of the function of YAP1 may help to serve as a molecular target in glioma therapeutics. Knockdown of YAP1 by shRNA technology has been shown to reduce glioma in vitro; however, clinical implications are still under investigation. YAP1 can be used as a diagnostic marker for gliomas to monitor the disease status and may help to evaluate its treatment effects. More functional experiments are needed to support the direct roles of YAP1 on gliomas at molecular and cellular levels.

  11. Oligouridylate Binding Protein 1b Plays an Integral Role in Plant Heat Stress Tolerance

    PubMed Central

    Nguyen, Cam Chau; Nakaminami, Kentaro; Matsui, Akihiro; Kobayashi, Shuhei; Kurihara, Yukio; Toyooka, Kiminori; Tanaka, Maho; Seki, Motoaki

    2016-01-01

    Stress granules (SGs), which are formed in the plant cytoplasm under stress conditions, are transient dynamic sites (particles) for mRNA storage. SGs are actively involved in protecting mRNAs from degradation. Oligouridylate binding protein 1b (UBP1b) is a component of SGs. The formation of microscopically visible cytoplasmic foci, referred to as UBP1b SG, was induced by heat treatment in UBP1b-overexpressing Arabidopsis plants (UBP1b-ox). A detailed understanding of the function of UBP1b, however, is still not clear. UBP1b-ox plants displayed increased heat tolerance, relative to control plants, while ubp1b mutants were more sensitive to heat stress than control plants. Microarray analysis identified 117 genes whose expression was heat-inducible and higher in the UBP1b-ox plants. RNA decay analysis was performed using cordycepin, a transcriptional inhibitor. In order to determine if those genes serve as targets of UBP1b, the rate of RNA degradation of a DnaJ heat shock protein and a stress-associated protein (AtSAP3) in UBP1b-ox plants was slower than in control plants; indicating that the mRNAs of these genes were protected within the UBP1b SG granule. Collectively, these data demonstrate that UBP1b plays an integral role in heat stress tolerance in plants. PMID:27379136

  12. Honokiol ameliorates endothelial dysfunction through suppression of PTX3 expression, a key mediator of IKK/IκB/NF-κB, in atherosclerotic cell model

    PubMed Central

    Qiu, Ling; Xu, Rong; Wang, Siyang; Li, Shuijun; Sheng, Hongguang; Wu, Jiaxi; Qu, Yi

    2015-01-01

    Pentraxin 3 (PTX3) was identified as a marker of the inflammatory response and overexpressed in various tissues and cells related to cardiovascular disease. Honokiol, an active component isolated from the Chinese medicinal herb Magnolia officinalis, was shown to have a variety of pharmacological activities. In the present study, we aimed to investigate the effects of honokiol on palmitic acid (PA)-induced dysfunction of human umbilical vein endothelial cells (HUVECs) and to elucidate potential regulatory mechanisms in this atherosclerotic cell model. Our results showed that PA significantly accelerated the expression of PTX3 in HUVECs through the IκB kinase (IKK)/IκB/nuclear factor-κB (NF-κB) pathway, reduced cell viability, induced cell apoptosis and triggered the inflammatory response. Knockdown of PTX3 supported cell growth and prevented apoptosis by blocking PA-inducted nitric oxide (NO) overproduction. Honokiol significantly suppressed the overexpression of PTX3 in PA-inducted HUVECs by inhibiting IκB phosphorylation and the expression of two NF-κB subunits (p50 and p65) in the IKK/IκB/NF-κB signaling pathway. Furthermore, honokiol reduced endothelial cell injury and apoptosis by regulating the expression of inducible NO synthase and endothelial NO synthase, as well as the generation of NO. Honokiol showed an anti-inflammatory effect in PA-inducted HUVECs by significantly inhibiting the generation of interleukin-6 (IL-6), IL-8 and monocyte chemoattractant protein-1. In summary, honokiol repaired endothelial dysfunction by suppressing PTX3 overexpression in an atherosclerotic cell model. PTX3 may be a potential therapeutic target for atherosclerosis. PMID:26138903

  13. Lactones from Ligusticum chuanxiong Hort. reduces atherosclerotic lesions in apoE-deficient mice via inhibiting over expression of NF-kB-dependent adhesion molecules.

    PubMed

    Xiao, Yang; Wang, Ying-Chao; Li, Lai-Lai; Jin, Ye-Cheng; Sironi, Luigi; Wang, Yi; Wang, Yi

    2014-06-01

    The present study aims to investigate the anti-atherosclerotic effects of lactones extracted from Ligusticum chuanxiong Hort (LLC) in apoE-deficient mice (ApoE(-/-) mice) and proclaim its underlying mechanisms. Expression of endothelial adhesion molecules and NF-κB around the atherosclerotic lesions was detected by immunohistochemistry (IHC). To further validate the mechanism, effect of LLC on the secretion of ICAM-1 and VCAM-1 of human umbilical vein endothelial cells (HUVECs) induced by tumor necrosis factor α (TNF-α) was measured by ELISA. And the activation of NF-κB was detected by western blot. Mice treated with LLC showed significant reduction in lesion sizes of thoracic segments of the aorta (p<0.01). Meanwhile, LLC treatments lead to decreases of serum TG, TC and LDL-C contents, respectively. LLC also decreased the expression of CD31, intercellular adhesion molecule-1 (ICAM-1), monocyte chemoattractant protein-1 (MCP-1) and nuclear factor-kappa B (NF-κB) in the atherosclerotic plaque. Moreover, LLC at 3.125-25 μg/mL can dose-dependently attenuate the expression of ICAM-1 and VCAM-1 in TNF-α stimulated HUVECs. Western blot result indicated LLC inhibited activation of NF-κB. These results suggested that LLC could ameliorate atherosclerosis in ApoE(-/-) mice. The mechanism of action of LLC on anti-atherosclerotic effect may be attributed to the suppression of the production of NF-κB-dependent adhesion molecules. PMID:24594239

  14. Honokiol ameliorates endothelial dysfunction through suppression of PTX3 expression, a key mediator of IKK/IκB/NF-κB, in atherosclerotic cell model.

    PubMed

    Qiu, Ling; Xu, Rong; Wang, Siyang; Li, Shuijun; Sheng, Hongguang; Wu, Jiaxi; Qu, Yi

    2015-01-01

    Pentraxin 3 (PTX3) was identified as a marker of the inflammatory response and overexpressed in various tissues and cells related to cardiovascular disease. Honokiol, an active component isolated from the Chinese medicinal herb Magnolia officinalis, was shown to have a variety of pharmacological activities. In the present study, we aimed to investigate the effects of honokiol on palmitic acid (PA)-induced dysfunction of human umbilical vein endothelial cells (HUVECs) and to elucidate potential regulatory mechanisms in this atherosclerotic cell model. Our results showed that PA significantly accelerated the expression of PTX3 in HUVECs through the IκB kinase (IKK)/IκB/nuclear factor-κB (NF-κB) pathway, reduced cell viability, induced cell apoptosis and triggered the inflammatory response. Knockdown of PTX3 supported cell growth and prevented apoptosis by blocking PA-inducted nitric oxide (NO) overproduction. Honokiol significantly suppressed the overexpression of PTX3 in PA-inducted HUVECs by inhibiting IκB phosphorylation and the expression of two NF-κB subunits (p50 and p65) in the IKK/IκB/NF-κB signaling pathway. Furthermore, honokiol reduced endothelial cell injury and apoptosis by regulating the expression of inducible NO synthase and endothelial NO synthase, as well as the generation of NO. Honokiol showed an anti-inflammatory effect in PA-inducted HUVECs by significantly inhibiting the generation of interleukin-6 (IL-6), IL-8 and monocyte chemoattractant protein-1. In summary, honokiol repaired endothelial dysfunction by suppressing PTX3 overexpression in an atherosclerotic cell model. PTX3 may be a potential therapeutic target for atherosclerosis. PMID:26138903

  15. Dectin-1-mediated Signaling Leads to Characteristic Gene Expressions and Cytokine Secretion via Spleen Tyrosine Kinase (Syk) in Rat Mast Cells*

    PubMed Central

    Kimura, Yukihiro; Chihara, Kazuyasu; Honjoh, Chisato; Takeuchi, Kenji; Yamauchi, Shota; Yoshiki, Hatsumi; Fujieda, Shigeharu; Sada, Kiyonao

    2014-01-01

    Dectin-1 recognizes β-glucan and plays important roles for the antifungal immunity through the activation of spleen tyrosine kinase (Syk) in dendritic cells or macrophages. Recently, expression of Dectin-1 was also identified in human and mouse mast cells, although its physiological roles were largely unknown. In this report, rat mast cell line RBL-2H3 was analyzed to investigate the molecular mechanism of Dectin-1-mediated activation and responses of mast cells. Treatment of cells with Dectin-1-specific agonist curdlan induced tyrosine phosphorylation of cellular proteins and the interaction of Dectin-1 with the Src homology 2 domain of Syk. These responses depended on tyrosine phosphorylation of the hemi-immunoreceptor tyrosine-based activation motif in the cytoplasmic tail of Dectin-1, whereas they were independent of the γ-subunit of high-affinity IgE receptor. DNA microarray and real-time PCR analyses showed that Dectin-1-mediated signaling stimulated gene expression of transcription factor Nfkbiz and inflammatory cytokines, such as monocyte chemoattractant protein-1, IL-3, IL-4, IL-13, and tumor necrosis factor (TNF)-α. The response was abrogated by pretreatment with Syk inhibitor R406. These results suggest that Syk is critical for Dectin-1-mediated activation of mast cells, although the signaling differs from that triggered by FcϵRI activation. In addition, these gene expressions induced by curdlan stimulation were specifically observed in mast cells, suggesting that Dectin-1-mediated signaling of mast cells offers new insight into the antifungal immunity. PMID:25246527

  16. Inflammation Promotes Expression of Stemness-Related Properties in HBV-Related Hepatocellular Carcinoma.

    PubMed

    Chang, Te-Sheng; Chen, Chi-Long; Wu, Yu-Chih; Liu, Jun-Jen; Kuo, Yung Che; Lee, Kam-Fai; Lin, Sin-Yi; Lin, Sey-En; Tung, Shui-Yi; Kuo, Liang-Mou; Tsai, Ying-Huang; Huang, Yen-Hua

    2016-01-01

    The expression of cancer stemness is believed to reduce the efficacy of current therapies against hepatocellular carcinoma (HCC). Understanding of the stemness-regulating signaling pathways incurred by a specific etiology can facilitate the development of novel targets for individualized therapy against HCC. Niche environments, such as virus-induced inflammation, may play a crucial role. However, the mechanisms linking inflammation and stemness expression in HCC remain unclear. Here we demonstrated the distinct role of inflammatory mediators in expressions of stemness-related properties involving the pluripotent octamer-binding transcription factor 4 (OCT4) in cell migration and drug resistance of hepatitis B virus-related HCC (HBV-HCC). We observed positive immunorecognition for macrophage chemoattractant protein 1 (MCP-1)/CD68 and OCT4/NANOG in HBV-HCC tissues. The inflammation-conditioned medium (inflamed-CM) generated by lipopolysaccharide-stimulated U937 human leukemia cells significantly increased the mRNA and protein levels of OCT4/NANOG preferentially in HBV-active (HBV+HBsAg+) HCC cells. The inflamed-CM also increased the side population (SP) cell percentage, green fluorescent protein (GFP)-positive cell population, and luciferase activity of OCT4 promoter-GFP/luciferase in HBV-active HCC cells. Furthermore, the inflamed-CM upregulated the expressions of insulin-like growth factor-I (IGF-I)/IGF-I receptor (IGF-IR) and activated IGF-IR/Akt signaling in HBV-HCC. The IGF-IR phosphorylation inhibitor picropodophyllin (PPP) suppressed inflamed-CM-induced OCT4 and NANOG levels in HBV+HBsAg+ Hep3B cells. Forced expression of OCT4 significantly increased the secondary sphere formation and cell migration, and reduced susceptibility of HBV-HCC cells to cisplatin, bleomycin, and doxorubicin. Taking together, our results show that niche inflammatory mediators play critical roles in inducing the expression of stemness-related properties involving IGF-IR activation, and

  17. Huntingtin-interacting protein 1 is overexpressed in prostate and colon cancer and is critical for cellular survival

    PubMed Central

    Rao, Dinesh S.; Hyun, Teresa S.; Kumar, Priti D.; Mizukami, Ikuko F.; Rubin, Mark A.; Lucas, Peter C.; Sanda, Martin G.; Ross, Theodora S.

    2002-01-01

    Huntingtin-interacting protein 1 (HIP1) is a cofactor in clathrin-mediated vesicle trafficking. It was first implicated in cancer biology as part of a chromosomal translocation in leukemia. Here we report that HIP1 is expressed in prostate and colon tumor cells, but not in corresponding benign epithelia. The relationship between HIP1 expression in primary prostate cancer and clinical outcomes was evaluated with tissue microarrays. HIP1 expression was significantly associated with prostate cancer progression and metastasis. Conversely, primary prostate cancers lacking HIP1 expression consistently showed no progression after radical prostatectomy. In addition, the expression of HIP1 was elevated in prostate tumors from the transgenic mouse model of prostate cancer (TRAMP). At the molecular level, expression of a dominant negative mutant of HIP1 led to caspase-9–dependent apoptosis, suggesting that HIP1 is a cellular survival factor. Thus, HIP1 may play a role in tumorigenesis by allowing the survival of precancerous or cancerous cells. HIP1 might accomplish this via regulation of clathrin-mediated trafficking, a fundamental cellular pathway that has not previously been associated with tumorigenesis. HIP1 represents a putative prognostic factor for prostate cancer and a potential therapy target in prostate as well as colon cancers. PMID:12163454

  18. Cannabinoid Receptor–Interacting Protein 1a Modulates CB1 Receptor Signaling and Regulation

    PubMed Central

    Smith, Tricia H.; Blume, Lawrence C.; Straiker, Alex; Cox, Jordan O.; David, Bethany G.; McVoy, Julie R. Secor; Sayers, Katherine W.; Poklis, Justin L.; Abdullah, Rehab A.; Egertová, Michaela; Chen, Ching-Kang; Mackie, Ken; Elphick, Maurice R.; Howlett, Allyn C.

    2015-01-01

    Cannabinoid CB1 receptors (CB1Rs) mediate the presynaptic effects of endocannabinoids in the central nervous system (CNS) and most behavioral effects of exogenous cannabinoids. Cannabinoid receptor–interacting protein 1a (CRIP1a) binds to the CB1R C-terminus and can attenuate constitutive CB1R-mediated inhibition of Ca2+ channel activity. We now demonstrate cellular colocalization of CRIP1a at neuronal elements in the CNS and show that CRIP1a inhibits both constitutive and agonist-stimulated CB1R-mediated guanine nucleotide–binding regulatory protein (G-protein) activity. Stable overexpression of CRIP1a in human embryonic kidney (HEK)-293 cells stably expressing CB1Rs (CB1-HEK), or in N18TG2 cells endogenously expressing CB1Rs, decreased CB1R-mediated G-protein activation (measured by agonist-stimulated [35S]GTPγS (guanylyl-5′-[O-thio]-triphosphate) binding) in both cell lines and attenuated inverse agonism by rimonabant in CB1-HEK cells. Conversely, small-interfering RNA–mediated knockdown of CRIP1a in N18TG2 cells enhanced CB1R-mediated G-protein activation. These effects were not attributable to differences in CB1R expression or endocannabinoid tone because CB1R levels did not differ between cell lines varying in CRIP1a expression, and endocannabinoid levels were undetectable (CB1-HEK) or unchanged (N18TG2) by CRIP1a overexpression. In CB1-HEK cells, 4-hour pretreatment with cannabinoid agonists downregulated CB1Rs and desensitized agonist-stimulated [35S]GTPγS binding. CRIP1a overexpression attenuated CB1R downregulation without altering CB1R desensitization. Finally, in cultured autaptic hippocampal neurons, CRIP1a overexpression attenuated both depolarization-induced suppression of excitation and inhibition of excitatory synaptic activity induced by exogenous application of cannabinoid but not by adenosine A1 agonists. These results confirm that CRIP1a inhibits constitutive CB1R activity and demonstrate that CRIP1a can also inhibit agonist

  19. Cannabinoid receptor-interacting protein 1a modulates CB1 receptor signaling and regulation.

    PubMed

    Smith, Tricia H; Blume, Lawrence C; Straiker, Alex; Cox, Jordan O; David, Bethany G; McVoy, Julie R Secor; Sayers, Katherine W; Poklis, Justin L; Abdullah, Rehab A; Egertová, Michaela; Chen, Ching-Kang; Mackie, Ken; Elphick, Maurice R; Howlett, Allyn C; Selley, Dana E

    2015-04-01

    Cannabinoid CB1 receptors (CB1Rs) mediate the presynaptic effects of endocannabinoids in the central nervous system (CNS) and most behavioral effects of exogenous cannabinoids. Cannabinoid receptor-interacting protein 1a (CRIP1a) binds to the CB1R C-terminus and can attenuate constitutive CB1R-mediated inhibition of Ca(2+) channel activity. We now demonstrate cellular colocalization of CRIP1a at neuronal elements in the CNS and show that CRIP1a inhibits both constitutive and agonist-stimulated CB1R-mediated guanine nucleotide-binding regulatory protein (G-protein) activity. Stable overexpression of CRIP1a in human embryonic kidney (HEK)-293 cells stably expressing CB1Rs (CB1-HEK), or in N18TG2 cells endogenously expressing CB1Rs, decreased CB1R-mediated G-protein activation (measured by agonist-stimulated [(35)S]GTPγS (guanylyl-5'-[O-thio]-triphosphate) binding) in both cell lines and attenuated inverse agonism by rimonabant in CB1-HEK cells. Conversely, small-interfering RNA-mediated knockdown of CRIP1a in N18TG2 cells enhanced CB1R-mediated G-protein activation. These effects were not attributable to differences in CB1R expression or endocannabinoid tone because CB1R levels did not differ between cell lines varying in CRIP1a expression, and endocannabinoid levels were undetectable (CB1-HEK) or unchanged (N18TG2) by CRIP1a overexpression. In CB1-HEK cells, 4-hour pretreatment with cannabinoid agonists downregulated CB1Rs and desensitized agonist-stimulated [(35)S]GTPγS binding. CRIP1a overexpression attenuated CB1R downregulation without altering CB1R desensitization. Finally, in cultured autaptic hippocampal neurons, CRIP1a overexpression attenuated both depolarization-induced suppression of excitation and inhibition of excitatory synaptic activity induced by exogenous application of cannabinoid but not by adenosine A1 agonists. These results confirm that CRIP1a inhibits constitutive CB1R activity and demonstrate that CRIP1a can also inhibit agonist

  20. Convergent Signaling Pathways Controlled by LRP1 (Receptor-related Protein 1) Cytoplasmic and Extracellular Domains Limit Cellular Cholesterol Accumulation.

    PubMed

    El Asmar, Zeina; Terrand, Jérome; Jenty, Marion; Host, Lionel; Mlih, Mohamed; Zerr, Aurélie; Justiniano, Hélène; Matz, Rachel L; Boudier, Christian; Scholler, Estelle; Garnier, Jean-Marie; Bertaccini, Diego; Thiersé, Danièle; Schaeffer, Christine; Van Dorsselaer, Alain; Herz, Joachim; Bruban, Véronique; Boucher, Philippe

    2016-03-01

    The low density lipoprotein receptor-related protein 1 (LRP1) is a ubiquitously expressed cell surface receptor that protects from intracellular cholesterol accumulation. However, the underlying mechanisms are unknown. Here we show that the extracellular (α) chain of LRP1 mediates TGFβ-induced enhancement of Wnt5a, which limits intracellular cholesterol accumulation by inhibiting cholesterol biosynthesis and by promoting cholesterol export. Moreover, we demonstrate that the cytoplasmic (β) chain of LRP1 suffices to limit cholesterol accumulation in LRP1(-/-) cells. Through binding of Erk2 to the second of its carboxyl-terminal NPXY motifs, LRP1 β-chain positively regulates the expression of ATP binding cassette transporter A1 (ABCA1) and of neutral cholesterol ester hydrolase (NCEH1). These results highlight the unexpected functions of LRP1 and the canonical Wnt5a pathway and new therapeutic potential in cholesterol-associated disorders including cardiovascular diseases.

  1. Identification of Human Fumarylacetoacetate Hydrolase Domain-containing Protein 1 (FAHD1) as a Novel Mitochondrial Acylpyruvase*

    PubMed Central

    Pircher, Haymo; Straganz, Grit D.; Ehehalt, Daniela; Morrow, Geneviève; Tanguay, Robert M.; Jansen-Dürr, Pidder

    2011-01-01

    The human fumarylacetoacetate hydrolase (FAH) domain-containing protein 1 (FAHD1) is part of the FAH protein superfamily, but its enzymatic function is unknown. In the quest for a putative enzymatic function of FAHD1, we found that FAHD1 exhibits acylpyruvase activity, demonstrated by the hydrolysis of acetylpyruvate and fumarylpyruvate in vitro, whereas several structurally related compounds were not hydrolyzed as efficiently. Conserved amino acids Asp-102 and Arg-106 of FAHD1 were found important for its catalytic activity, and Mg2+ was required for maximal enzyme activity. FAHD1 was found expressed in all tested murine tissues, with highest expression in liver and kidney. FAHD1 was also found in several human cell lines, where it localized to mitochondria. In summary, the current work identified mammalian FAHD1 as a novel mitochondrial enzyme with acylpyruvate hydrolase activity. PMID:21878618

  2. Nickel decreases cellular iron level and converts cytosolic aconitase to iron-regulatory protein 1 in A549 cells.

    PubMed

    Chen, Haobin; Davidson, Todd; Singleton, Steven; Garrick, Michael D; Costa, Max

    2005-08-15

    Nickel (Ni) compounds are well-established carcinogens and are known to initiate a hypoxic response in cells via the stabilization and transactivation of hypoxia-inducible factor-1 alpha (HIF-1alpha). This change may be the consequence of nickel's interference with the function of several Fe(II)-dependent enzymes. In this study, the effects of soluble nickel exposure on cellular iron homeostasis were investigated. Nickel treatment decreased both mitochondrial and cytosolic aconitase (c-aconitase) activity in A549 cells. Cytosolic aconitase was converted to iron-regulatory protein 1, a form critical for the regulation of cellular iron homeostasis. The increased activity of iron-regulatory protein 1 after nickel exposure stabilized and increased transferrin receptor (Tfr) mRNA and antagonized the iron-induced ferritin light chain protein synthesis. The decrease of aconitase activity after nickel treatment reflected neither direct interference with aconitase function nor obstruction of [4Fe-4S] cluster reconstitution by nickel. Exposure of A549 cells to soluble nickel decreased total cellular iron by about 40%, a decrease that likely caused the observed decrease in aconitase activity and the increase of iron-regulatory protein 1 activity. Iron treatment reversed the effect of nickel on cytosolic aconitase and iron-regulatory protein 1. To assess the mechanism for the observed effects, human embryonic kidney (HEK) cells over expressing divalent metal transporter-1 (DMT1) were compared to A549 cells expressing only endogenous transporters for inhibition of iron uptake by nickel. The inhibition data suggest that nickel can enter via DMT1 and compete with iron for entry into the cell. This disturbance of cellular iron homeostasis by nickel may have a great impact on the ability of the cell to regulate a variety of cell functions, as well as create a state of hypoxia in cells under normal oxygen tension. These effects may be very important in how nickel exerts phenotypic

  3. Myc Promoter-Binding Protein-1 (MBP-1) Is a Novel Potential Prognostic Marker in Invasive Ductal Breast Carcinoma

    PubMed Central

    Contino, Flavia; Mazzarella, Claudia; Sbacchi, Silvia; Roz, Elena; Lupo, Carmelo; Perconti, Giovanni; Giallongo, Agata; Migliorini, Paola; Marrazzo, Antonio; Feo, Salvatore

    2010-01-01

    Background Alpha-enolase is a glycolytic enzyme that catalyses the formation of phosphoenolpyruvate in the cell cytoplasm. α-Enolase and the predominantly nuclear Myc promoter-binding protein-1 (MBP-1) originate from a single gene through the alternative use of translational starting sites. MBP-1 binds to the P2 c-myc promoter and competes with TATA-box binding protein (TBP) to suppress gene transcription. Although several studies have shown an antiproliferative effect of MBP-1 overexpression on several human cancer cells, to date detailed observations of α-enolase and MBP-1 relative expression in primary tumors versus normal tissues and their correlation with clinicopathological features have not been undertaken. Methodology and Findings We analyzed α-enolase and MBP-1 expression in normal breast epithelium and primary invasive ductal breast carcinoma (IDC) from 177 patients by Western blot and immunohistochemical analyses, using highly specific anti-α-enolase monoclonal antibodies. A significant increase in the expression of cytoplasmic α-enolase was observed in 98% of the tumors analysed, compared to normal tissues. Nuclear MBP-1 was found in almost all the normal tissues while its expression was retained in only 35% of the tumors. Statistically significant associations were observed among the nuclear expression of MBP-1 and ErbB2 status, Ki-67 expression, node status and tumor grade. Furthermore MBP-1 expression was associated with good survival of patients with IDC. Conclusions MBP-1 functions in repressing c-myc gene expression and the results presented indicate that the loss of nuclear MBP-1 expression in a large number of IDC may be a critical step in the development and progression of breast cancer and a predictor of adverse outcome. Nuclear MBP-1 appears to be a novel and valuable histochemical marker with potential prognostic value in breast cancer. PMID:20886042

  4. Effect of telmisartan on the expression of adiponectin receptors and nicotinamide adenine dinucleotide phosphate oxidase in the heart and aorta in type 2 diabetic rats

    PubMed Central

    2012-01-01

    Background Diabetic cardiovascular disease is associated with decreased adiponectin and increased oxidative stress. This study investigated the effect of telmisartan on the expression of adiponectin receptor 2 (adipoR2) and nicotinamide adenine dinucleotide phosphate (NADPH) oxidase subunits in the heart and the expression of adiponectin receptor 1 (adipoR1) in aorta in type 2 diabetic rats. Methods Type 2 diabetes was induced by high-fat and high-sugar diet and intraperitoneal injection of a low dose of streptozotocin (STZ). Heart function, adipoR2, p22phox, NOX4, glucose transporter 4(GLUT4), monocyte chemoattractant protein-1(MCP-1) and connective tissue growth factor (CTGF)in the heart, and adipoR1, MCP-1 and nuclear factor kappa B (NF-κB) in aorta were analyzed in controls and diabetic rats treated with or without telmisartan (5mg/kg/d) by gavage for 12 weeks. Results Heart function, plasma and myocardial adiponectin levels, the expression of myocardial adipoR2 and GLUT4 were significantly decreased in diabetic rats (P <0.05). The expression of myocardial p22phox, NOX4, MCP-1, and CTGF was significantly increased in diabetic rats (P <0.05). The expression of adipoR1 was decreased and the expression of MCP-1 and NF-κB was increased in the abdominal aorta in diabetic rats (P <0.05). Telmisartan treatment significantly attenuated these changes in diabetic rats (P <0.05). Conclusions Our results suggest that telmisartan upregulates the expression of myocardial adiponectin, its receptor 2 and GLUT4. Simultaneously, it downregulates the expression of myocardial p22phox, NOX4, MCP-1, and CTGF, contributing so to the improvement of heart function in diabetic rats. Telmisartan also induces a protective role on the vascular system by upregulating the expression of adipoR1 and downregulating the expression of MCP-1 and NF-κB in the abdominal aorta in diabetic rats. PMID:22873349

  5. CCR2 and CXCR3 agonistic chemokines are differently expressed and regulated in human alveolar epithelial cells type II

    PubMed Central

    Pechkovsky, Dmitri V; Goldmann, Torsten; Ludwig, Corinna; Prasse, Antje; Vollmer, Ekkehard; Müller-Quernheim, Joachim; Zissel, Gernot

    2005-01-01

    The attraction of leukocytes from circulation to inflamed lungs depends on the activation of both the leukocytes and the resident cells within the lung. In this study we determined gene expression and secretion patterns for monocyte chemoattractant protein-1 (MCP-1/CCL2) and T-cell specific CXCR3 agonistic chemokines (Mig/CXCL9, IP-10/CXCL10, and I-TAC/CXCL11) in TNF-α-, IFN-γ-, and IL-1β-stimulated human alveolar epithelial cells type II (AEC-II). AEC-II constitutively expressed high level of CCL2 mRNA in vitro and in situ , and released CCL2 protein in vitro . Treatment of AEC-II with proinflammatory cytokines up-regulated both CCL2 mRNA expression and release of immunoreactive CCL2, whereas IFN-γ had no effect on CCL2 release. In contrast, CXCR3 agonistic chemokines were not detected in freshly isolated AEC-II or in non-stimulated epithelial like cell line A549. IFN-γ, alone or in combination with IL-1β and TNF-α resulted in an increase in CXCL10, CXCL11, and CXCL9 mRNA expression and generation of CXCL10 protein by AEC-II or A549 cells. CXCL10 gene expression and secretion were induced in dose-dependent manner after cytokine-stimulation of AEC-II with an order of potency IFN-γ>>IL-1β ≥ TNF-α. Additionally, we localized the CCL2 and CXCL10 mRNAs in human lung tissue explants by in situ hybridization, and demonstrated the selective effects of cytokines and dexamethasone on CCL2 and CXCL10 expression. These data suggest that the regulation of the CCL2 and CXCL10 expression exhibit significant differences in their mechanisms, and also demonstrate that the alveolar epithelium contributes to the cytokine milieu of the lung, with the ability to respond to locally generated cytokines and to produce potent mediators of the local inflammatory response. PMID:16033640

  6. [Preparation and detection of anti-influenza A virus polymerase basic protein 1 polyclonal antibody].

    PubMed

    Qin, Yujie; Zhang, Tinghong; Ye, Xin

    2016-01-01

    Influenza A virus is an enveloped virus that belongs to the Orthomyxoviridae family. It has 8 negative RNA segments that encode 16 viral proteins. The viral polymerase consists of 3 proteins (PB 1, PB2 and PA) which plays an important role in the transcription and replication of the influenza A virus. Polymerase basic protein 1 (PB 1) is a critical member of viral polymerase complex. In order to further study the function of PB1, we need to prepare the PB1 antibody with good quality. Therefore, we amplified PB1 conserved region (nt1648-2265) by PCR and cloned it into pET-30a vector, and transformed into Escherichia coli BL2 1. The expression of His tagged PB 1 protein was induced by IPTG, and His-PB 1 proteins were purified by Ni-NTA resin. For preparation of PB 1 protein antiserum, rabbits were immunized with His-PB 1 fusion protein 3 times. Then the titer of PB 1 polyclonal antibody was measured by indirect ELISA. The antibody was purified by membrane affinity purification and subjected to immunoblotting analysis. Data showed that PB1 antibody can recognize PB 1 protein from WSN virus infected or pCMV FLAG-PB 1 transfected cells. Meanwhile, PB 1 antibody can also recognize specifically other subtype strains of influenza A virus such as H9N2 and H3N2. PB 1 polyclonal antibody we generated will be a useful tool to study the biological function of PB1. PMID:27363203

  7. Role of Plasmodium vivax Duffy-binding protein 1 in invasion of Duffy-null Africans

    PubMed Central

    Gunalan, Karthigayan; Lo, Eugenia; Hostetler, Jessica B.; Yewhalaw, Delenasaw; Mu, Jianbing; Neafsey, Daniel E.; Yan, Guiyun; Miller, Louis H.

    2016-01-01

    The ability of the malaria parasite Plasmodium vivax to invade erythrocytes is dependent on the expression of the Duffy blood group antigen on erythrocytes. Consequently, Africans who are null for the Duffy antigen are not susceptible to P. vivax infections. Recently, P. vivax infections in Duffy-null Africans have been documented, raising the possibility that P. vivax, a virulent pathogen in other parts of the world, may expand malarial disease in Africa. P. vivax binds the Duffy blood group antigen through its Duffy-binding protein 1 (DBP1). To determine if mutations in DBP1 resulted in the ability of P. vivax to bind Duffy-null erythrocytes, we analyzed P. vivax parasites obtained from two Duffy-null individuals living in Ethiopia where Duffy-null and -positive Africans live side-by-side. We determined that, although the DBP1s from these parasites contained unique sequences, they failed to bind Duffy-null erythrocytes, indicating that mutations in DBP1 did not account for the ability of P. vivax to infect Duffy-null Africans. However, an unusual DNA expansion of DBP1 (three and eight copies) in the two Duffy-null P. vivax infections suggests that an expansion of DBP1 may have been selected to allow low-affinity binding to another receptor on Duffy-null erythrocytes. Indeed, we show that Salvador (Sal) I P. vivax infects Squirrel monkeys independently of DBP1 binding to Squirrel monkey erythrocytes. We conclude that P. vivax Sal I and perhaps P. vivax in Duffy-null patients may have adapted to use new ligand–receptor pairs for invasion. PMID:27190089

  8. Lithium decreases the effects of neuronal calcium sensor protein 1 in pedunculopontine neurons.

    PubMed

    D'Onofrio, Stasia; Urbano, Francisco J; Messias, Erick; Garcia-Rill, Edgar

    2016-03-01

    Human postmortem studies reported increased expression of neuronal calcium sensor protein 1 (NCS-1) in the brains of some bipolar disorder patients, and reduced or aberrant gamma band activity is present in the same disorder. Bipolar disorder is characterized by sleep dysregulation, suggesting a role for the reticular activating system (RAS). Lithium (Li(+)) has been shown to effectively treat the mood disturbances in bipolar disorder patients and was proposed to act by inhibiting the interaction betweenNCS-1 and inositol 1,4,5-triphosphate receptor protein (InsP3R).NCS-1 is known to enhance the activity of InsP3R, and of Ca(2+)-mediated gamma oscillatory activity in the pedunculopontine nucleus (PPN), part of theRAS This study aimed to determine the nature of some of the intracellular mechanisms of Li(+)on ratPPNcells and to identify the interaction between Li(+)andNCS-1. Since Li(+)has been shown to act by inhibiting the enhancing effects ofNCS-1, we tested the hypothesis that Li(+)would reduced the effects of overexpression ofNCS-1 and prevent the downregulation of gamma band activity. Li(+)decreased gamma oscillation frequency and amplitude by downregulating Ca(2+)channel activity, whereasNCS-1 reduced the effect of Li(+)on Ca(2+)currents. These effects were mediated by a G-protein overinhibition of Ca(2+)currents. These results suggest that Li(+)affected intracellular pathways involving the activation of voltage-gated Ca(2+)channels mediated by an intracellular mechanism involving voltage-dependent activation of G proteins, thereby normalizing gamma band oscillations mediated by P/Q-type calcium channels modulated byNCS-1.

  9. Controlled release of recombinant human cementum protein 1 from electrospun multiphasic scaffold for cementum regeneration.

    PubMed

    Chen, Xiaofeng; Liu, Yu; Miao, Leiying; Wang, Yangyang; Ren, Shuangshuang; Yang, Xuebin; Hu, Yong; Sun, Weibin

    2016-01-01

    Periodontitis is a major cause for tooth loss, which affects about 15% of the adult population. Cementum regeneration has been the crux of constructing the periodontal complex. Cementum protein 1 (CEMP1) is a cementum-specific protein that can induce cementogenic differentiation. In this study, poly(ethylene glycol) (PEG)-stabilized amorphous calcium phosphate (ACP) nanoparticles were prepared by wet-chemical method and then loaded with recombinant human CEMP1 (rhCEMP1) for controlled release. An electrospun multiphasic scaffold constituted of poly(ε-caprolactone) (PCL), type I collagen (COL), and rhCEMP1/ACP was fabricated. The effects of rhCEMP1/ACP/PCL/COL scaffold on the attachment proliferation, osteogenic, and cementogenic differentiations of human periodontal ligament cells, (PDLCs) were systematically investigated. A critical size defect rat model was introduced to evaluate the effect of tissue regeneration of the scaffolds in vivo. The results showed that PEG-stabilized ACP nanoparticles formed a core-shell structure with sustained release of rhCEMP1 for up to 4 weeks. rhCEMP1/ACP/PCL/COL scaffold could suppress PDLCs proliferation behavior and upregulate the expression of cementoblastic markers including CEMP1 and cementum attachment protein while downregulating osteoblastic markers including osteocalcin and osteopontin when it was cocultured with PDLCs in vitro for 7 days. Histology analysis of cementum after being implanted with the scaffold in rats for 8 weeks showed that there was cementum-like tissue formation but little bone formation. These results indicated the potential of using electrospun multiphasic scaffolds for controlled release of rhCEMP1 for promoting cementum regeneration in reconstruction of the periodontal complex. PMID:27471382

  10. Role of Plasmodium vivax Duffy-binding protein 1 in invasion of Duffy-null Africans.

    PubMed

    Gunalan, Karthigayan; Lo, Eugenia; Hostetler, Jessica B; Yewhalaw, Delenasaw; Mu, Jianbing; Neafsey, Daniel E; Yan, Guiyun; Miller, Louis H

    2016-05-31

    The ability of the malaria parasite Plasmodium vivax to invade erythrocytes is dependent on the expression of the Duffy blood group antigen on erythrocytes. Consequently, Africans who are null for the Duffy antigen are not susceptible to P. vivax infections. Recently, P. vivax infections in Duffy-null Africans have been documented, raising the possibility that P. vivax, a virulent pathogen in other parts of the world, may expand malarial disease in Africa. P. vivax binds the Duffy blood group antigen through its Duffy-binding protein 1 (DBP1). To determine if mutations in DBP1 resulted in the ability of P. vivax to bind Duffy-null erythrocytes, we analyzed P. vivax parasites obtained from two Duffy-null individuals living in Ethiopia where Duffy-null and -positive Africans live side-by-side. We determined that, although the DBP1s from these parasites contained unique sequences, they failed to bind Duffy-null erythrocytes, indicating that mutations in DBP1 did not account for the ability of P. vivax to infect Duffy-null Africans. However, an unusual DNA expansion of DBP1 (three and eight copies) in the two Duffy-null P. vivax infections suggests that an expansion of DBP1 may have been selected to allow low-affinity binding to another receptor on Duffy-null erythrocytes. Indeed, we show that Salvador (Sal) I P. vivax infects Squirrel monkeys independently of DBP1 binding to Squirrel monkey erythrocytes. We conclude that P. vivax Sal I and perhaps P. vivax in Duffy-null patients may have adapted to use new ligand-receptor pairs for invasion.

  11. Controlled release of recombinant human cementum protein 1 from electrospun multiphasic scaffold for cementum regeneration

    PubMed Central

    Chen, Xiaofeng; Liu, Yu; Miao, Leiying; Wang, Yangyang; Ren, Shuangshuang; Yang, Xuebin; Hu, Yong; Sun, Weibin

    2016-01-01

    Periodontitis is a major cause for tooth loss, which affects about 15% of the adult population. Cementum regeneration has been the crux of constructing the periodontal complex. Cementum protein 1 (CEMP1) is a cementum-specific protein that can induce cementogenic differentiation. In this study, poly(ethylene glycol) (PEG)-stabilized amorphous calcium phosphate (ACP) nanoparticles were prepared by wet-chemical method and then loaded with recombinant human CEMP1 (rhCEMP1) for controlled release. An electrospun multiphasic scaffold constituted of poly(ε-caprolactone) (PCL), type I collagen (COL), and rhCEMP1/ACP was fabricated. The effects of rhCEMP1/ACP/PCL/COL scaffold on the attachment proliferation, osteogenic, and cementogenic differentiations of human periodontal ligament cells, (PDLCs) were systematically investigated. A critical size defect rat model was introduced to evaluate the effect of tissue regeneration of the scaffolds in vivo. The results showed that PEG-stabilized ACP nanoparticles formed a core-shell structure with sustained release of rhCEMP1 for up to 4 weeks. rhCEMP1/ACP/PCL/COL scaffold could suppress PDLCs proliferation behavior and upregulate the expression of cementoblastic markers including CEMP1 and cementum attachment protein while downregulating osteoblastic markers including osteocalcin and osteopontin when it was cocultured with PDLCs in vitro for 7 days. Histology analysis of cementum after being implanted with the scaffold in rats for 8 weeks showed that there was cementum-like tissue formation but little bone formation. These results indicated the potential of using electrospun multiphasic scaffolds for controlled release of rhCEMP1 for promoting cementum regeneration in reconstruction of the periodontal complex. PMID:27471382

  12. Inhibition of Protein Synthesis by Y Box-Binding Protein 1 Blocks Oncogenic Cell Transformation†

    PubMed Central

    Bader, Andreas G.; Vogt, Peter K.

    2005-01-01

    The multifunctional Y box-binding protein 1 (YB-1) is transcriptionally repressed by the oncogenic phosphoinositide 3-kinase (PI3K) pathway (with P3K as an oncogenic homolog of the catalytic subunit) and, when reexpressed with the retroviral vector RCAS, interferes with P3K- and Akt-induced transformation of chicken embryo fibroblasts. Retrovirally expressed YB-1 binds to the cap of mRNAs and inhibits cap-dependent and cap-independent translation. To determine the requirements for the inhibitory role of YB-1 in P3K-induced transformation, we conducted a mutational analysis, measuring YB-1-induced interference with transformation, subcellular localization, cap binding, mRNA binding, homodimerization, and inhibition of translation. The results show that (i) interference with transformation requires RNA binding and a C-terminal domain that is distinct from the cytoplasmic retention domain, (ii) interference with transformation is tightly correlated with inhibition of translation, and (iii) masking of mRNAs by YB-1 is not sufficient to block transformation or to inhibit translation. We identified a noncanonical nuclear localization signal (NLS) in the C-terminal half of YB-1. A mutant lacking the NLS retains its ability to interfere with transformation, indicating that a nuclear function is not required. These results suggest that YB-1 interferes with P3K-induced transformation by a specific inhibition of translation through its RNA-binding domain and a region in the C-terminal domain. Potential functions of the C-terminal region are discussed. PMID:15743808

  13. Role of CC chemokine CCL6/C10 as a monocyte chemoattractant in a murine acute peritonitis.

    PubMed Central

    LaFleur, Andrew M; Lukacs, Nicholas W; Kunkel, Steven L; Matsukawa, Akihiro

    2004-01-01

    The aim of this study was to determine the role of CC chemokine CCL6/C10 in acute inflammation. Intraperitoneal injection of thioglycollate increased peritoneal CCL6, which peaked at 4 h and remained elevated at 48 h. Neutralization of CCL6 significantly inhibited the macrophage infiltration (34-48% reduction), but not other cell types, without decreasing the other CC chemokines known to attract monocytes/macrophages. CCL6 was expressed in peripheral eosinophils and elicited macrophages, but not in elicited neutrophils. Peritoneal CCL6 level was not decreased in granulocyte-depleted mice where eosinophil influx was significantly impaired. Thus, CCL6 appears to contribute to the macrophage infiltration that is independent of other CC chemokines. Eosinophils pre-store CCL6, but do not release CCL6 in the peritoneum in this model of inflammation. PMID:15770051

  14. Heterochromatin protein 1 (HP1) connects the FACT histone chaperone complex to the phosphorylated CTD of RNA polymerase II

    PubMed Central

    Kwon, So Hee; Florens, Laurence; Swanson, Selene K.; Washburn, Michael P.; Abmayr, Susan M.; Workman, Jerry L.

    2010-01-01

    Heterochromatin protein 1 (HP1) is well known as a silencing protein found at pericentric heterochromatin. Most eukaryotes have at least three isoforms of HP1 that play differential roles in heterochromatin and euchromatin. In addition to its role in heterochromatin, HP1 proteins have been shown to function in transcription elongation. To gain insights into the transcription functions of HP1, we sought to identify novel HP1-interacting proteins. Biochemical and proteomic approaches revealed that HP1 interacts with the histone chaperone complex FACT (facilitates chromatin transcription). HP1c interacts with the SSRP1 (structure-specific recognition protein 1) subunit and the intact FACT complex. Moreover, HP1c guides the recruitment of FACT to active genes and links FACT to active forms of RNA polymerase II. The absence of HP1c partially impairs the recruitment of FACT into heat-shock loci and causes a defect in heat-shock gene expression. Thus, HP1c functions to recruit the FACT complex to RNA polymerase II. PMID:20889714

  15. Molecular pathobiology of human cervical high-grade lesions: paracrine STAT3 activation in tumor-instructed myeloid cells drives local MMP-9 expression.

    PubMed

    Schröer, Nadine; Pahne, Jennifer; Walch, Barbara; Wickenhauser, Claudia; Smola, Sigrun

    2011-01-01

    In many tumors, the switch from precancerous lesions to malignancy critically relies on expression of the matrix-metalloprotease MMP-9, which is predominantly provided by infiltrating inflammatory cells. Our study defines a novel molecular cascade, how human neoplastic cells instruct tumor-associated myeloid cells to produce MMP-9. In biopsies of human papillomavirus-associated precancerous cervical intraepithelial neoplasia (CIN III lesions), we show broad activation of the transcription factor STAT3 and coexpression of MMP-9 in perivascular inflammatory cells. For the first time, we establish a causative link between tumor-mediated paracrine STAT3 activation and MMP-9 production by human tumor-instructed monocytes, whereas NF-κB activation is dispensable for this response. Our data provide evidence that STAT3 does not directly induce MMP-9 but first leads to a strong production of the monocyte chemoattractant protein-1 (CCL2) in the nanogram range. In a second phase, autocrine stimulation of the CCR2 receptor in the tumor-instructed monocytes amplifies MMP-9 expression via intracellular Ca(2+) signaling. These findings elucidate a critical mechanism in the molecular pathobiology of cervical carcinogenesis at the switch to malignancy. Particularly in tumors, which are associated with infectious agents, STAT3-driven inflammation may be pivotal to promote carcinogenesis, while at the same time limit NF-κB-dependent immune responses and thus rejection of the infected preneoplastic cells. The molecular cascade defined in this study provides the basis for a rational design of future adjuvant therapies of cervical precancerous lesions.

  16. Longevity is associated with increased vascular resistance to high glucose-induced oxidative stress and inflammatory gene expression in Peromyscus leucopus.

    PubMed

    Labinskyy, Nazar; Mukhopadhyay, Partha; Toth, Janos; Szalai, Gabor; Veres, Monika; Losonczy, Gyorgy; Pinto, John T; Pacher, Pal; Ballabh, Praveen; Podlutsky, Andrej; Austad, Steven N; Csiszar, Anna; Ungvari, Zoltan

    2009-04-01

    Vascular aging is characterized by increased oxidative stress and proinflammatory phenotypic alterations. Metabolic stress, such as hyperglycemia in diabetes, is known to increase the production of ROS and promote inflammatory gene expression, accelerating vascular aging. The oxidative stress hypothesis of aging predicts that vascular cells of long-lived species exhibit lower steady-state production of ROS and/or superior resistance to the prooxidant effects of metabolic stress. We tested this hypothesis using two taxonomically related rodents, the white-footed mouse (Peromyscus leucopus) and the house mouse (Mus musculus), which show a more than twofold difference in maximum lifespan potential (8.2 and 3.5 yr, respectively). We compared interspecies differences in steady-state and high glucose (HG; 30 mmol/l)-induced production of O(2)(*-) and H(2)O(2), endothelial function, mitochondrial ROS generation, and inflammatory gene expression in cultured aortic segments. In P. leucopus aortas, steady-state endothelial O(2)(*-) and H(2)O(2) production and ROS generation by mitochondria were less than in M. musculus vessels. Furthermore, vessels of P. leucopus were more resistant to the prooxidant effects of HG. Primary fibroblasts from P. leucopus also exhibited less steady-state and HG-induced ROS production than M. musculus cells. In M. musculus arteries, HG elicited significant upregulation of inflammatory markers (TNF-alpha, IL-6, ICAM-1, VCAM, and monocyte chemoattractant protein-1). In contrast, the proinflammatory effects of HG were blunted in P. leucopus vessels. Thus, increased life span potential in P. leucopus is associated with decreased cellular ROS generation and increased resistance to prooxidant and proinflammatory effects of metabolic stress, which accord with predictions of the oxidative stress hypothesis of aging.

  17. PPAR Agonist-Induced Reduction of Mcp1 in Atherosclerotic Plaques of Obese, Insulin-Resistant Mice Depends on Adiponectin-Induced Irak3 Expression

    PubMed Central

    Arnould, Thierry; Tsatsanis, Christos; Holvoet, Paul

    2013-01-01

    Synthetic peroxisome proliferator-activated receptor (PPAR) agonists are used to treat dyslipidemia and insulin resistance. In this study, we examined molecular mechanisms that explain differential effects of a PPARα agonist (fenofibrate) and a PPARγ agonist (rosiglitazone) on macrophages during obesity-induced atherogenesis. Twelve-week-old mice with combined leptin and LDL-receptor deficiency (DKO) were treated with fenofibrate, rosiglitazone or placebo for 12 weeks. Only rosiglitazone improved adipocyte function, restored insulin sensitivity, and inhibited atherosclerosis by decreasing lipid-loaded macrophages. In addition, it increased interleukin-1 receptor-associated kinase-3 (Irak3) and decreased monocyte chemoattractant protein-1 (Mcp1) expressions, indicative of a switch from M1 to M2 macrophages. The differences between fenofibrate and rosiglitazone were independent of Pparγ expression. In bone marrow-derived macrophages (BMDM), we identified the rosiglitazone-associated increase in adiponectin as cause of the increase in Irak3. Interestingly, the deletion of Irak3 in BMDM (IRAK3−/− BMDM) resulted in activation of the canonical NFκB signaling pathway and increased Mcp1 protein secretion. Rosiglitazone could not decrease the elevated Mcp1 secretion in IRAK3−/− BMDM directly and fenofibrate even increased the secretion, possibly due to increased mitochondrial reactive oxygen species production. Furthermore, aortic extracts of high-fat insulin-resistant LDL-receptor deficient mice, with lower adiponectin and Irak3 and higher Mcp1, showed accelerated atherosclerosis. In aggregate, our results emphasize an interaction between PPAR agonist-mediated increase in adiponectin and macrophage-associated Irak3 in the protection against atherosclerosis by PPAR agonists. PMID:23620818

  18. Coupled Rapid Cell and Lattice Boltzmann Models to Simulate Hydrodynamics of Bacterial Transport in Response to Chemoattractant Gradients in Confined Domains

    NASA Astrophysics Data System (ADS)

    McKay, Cameron; Nguyen, Hoa; Basagaoglu, Hakan; Carpenter, Alexander; Succi, Sauro; Healy, Frank

    2015-11-01

    The Rapid Cell (RC) model was developed to simulate motility and adaptation dynamics of flagellar bacterial chemotaxis in environments with spatiotemporal variations in chemoattractant gradients. RC is best suited to motility studies in unbounded domains within non-fluid environments; this limits its use as a simulation tool. In this study, we eliminate these constraints by dynamically coupling RC with the colloidal lattice-Boltzmann (LB) model, a versatile tool for simulating transport of particles (e.g., surrogate bacteria) of distinct shapes and finite sizes in transient flow fields in geometrically complex microchannels. This was accomplished by tracking positions of chemoreceptor clusters on the particle surface that vary with particle angular and translational velocities, and by including additional forces and torques due to particles tumbling and to running motions in particle force-balance and torque-balance equations. The coupled RC-LB model successfully simulated transport of multiple particles in confined domains with single- or multi-attractant sources in a variety of settings. The coupled RC-LB model represents a first step toward development of a new modeling capability to simulate chemotaxis-driven bacterial transport in a network of geometrically irregular flow channels typically observed in tumor vasculature in the context of targeted drug delivery.

  19. Lymphocyte binding to vascular endothelium in inflamed skin revisited: a central role for vascular adhesion protein-1 (VAP-1).

    PubMed

    Arvilommi, A M; Salmi, M; Kalimo, K; Jalkanen, S

    1996-04-01

    The binding of leukocytes to vascular endothelium and their migration into tissues is mediated by adhesion molecules on the endothelial cells and leukocytes. Vascular adhesion protein-1 (VAP-1) is a 170-180/90-kDa endothelial molecule expressed most prominently in high endothelial venules in peripheral lymph node (PLN) type lymphatic tissues. VAP-1 mediates lymphocyte binding to PLN, tonsil and synovium. The expression of VAP-1 is induced in inflammatory diseases such as arthritis and gut inflammation. We examined the expression, structure and function of VAP-1 in normal and inflamed skin and compared it to those of other adhesion molecules implicated in skin homing. In psoriasis lichen ruber planus, pemphigoid and allergic lesions, VAP-1 was markedly upregulated. The expression of VAP-1 was also increased in biopsies of healthy skin of the patients. The VAP-1 molecule induced in skin is decorated with abundant sialic acids. VAP-1 inflamed skin is functional, since inhibition with anti-VAP-1 monoclonal antibodies caused a 60% reduction in lymphocytes adhesion to vascular endothelium. Antibodies against E-selectin, which has been regarded as the major vascular addressin directing cutaneous lymphocyte traffic, and, surprisingly, against peripheral lymph node addressin (PNAd), caused inhibitions of 30% and 60%, respectively, in the frozen section adhesion assay. These findings suggest important roles also for VAP-1 and PNAd in lymphocyte homing into inflamed skin. PMID:8625974

  20. Activator protein-1 (AP-1) and response to pathogen infection in the Hong Kong oyster (Crassostrea hongkongensis).

    PubMed

    Xiang, Zhiming; Qu, Fufa; Li, Jun; Qi, Lin; Yang, Zhang; Kong, Xiaoyu; Yu, Ziniu

    2014-01-01

    Growing evidence suggests that the transcription factor activator protein-1 (AP-1), a downstream target of mitogen-activated protein kinase (MAPK) signaling, plays a major role in stimulating the synthesis of immune effector molecules during innate immune responses. We have characterized ChAP-1, an AP-1-like protein in Crassostrea hongkongensis that is a member of the AP-1 family of proteins. ChAP-1 is composed of 290 amino acid residues with a Jun and bZIP domain at the N- and C-termini, respectively, a structure similar to that of known Ap-1 proteins. ChAP-1 mRNA is expressed in several tissues analyzed, with highest expression in the mantle. Expression of ChAP-1 increases in response to Vibrio alginolyticus, Salmo haemolyticus or Salmo cerevisiae infection and, despite the location of GFP-tagged full-length ChAP-1 protein in the cytoplasm, ChAP-1 activates the transcription of an L8G5-luc reporter gene, and its over-expression can also activate the AP-1-Luc reporter gene in HEK293T cells.

  1. Activator Protein-1 Regulation of Murine Aldehyde Dehydrogenase 1a1

    PubMed Central

    Makia, N. L.; Amunom, I.; Falkner, K. C.; Conklin, D. J.; Surapureddi, S.; Goldstein, J. A.

    2012-01-01

    Previously we demonstrated that aldehyde dehydrogenase (ALDH) 1a1 is the major ALDH expressed in mouse liver and is an effective catalyst in metabolism of lipid aldehydes. Quantitative real-time polymerase chain reaction analysis revealed a ≈2.5- to 3-fold induction of the hepatic ALDH1A1 mRNA in mice administered either acrolein (5 mg/kg acrolein p.o.) or butylated hydroxylanisole (BHA) (0.45% in the diet) and of cytosolic NAD+-dependent ALDH activity. We observed ≈2-fold increases in ALDH1A1 mRNA levels in both Nrf2(+/+) and Nrf2(−/−) mice treated with BHA compared with controls, suggesting that BHA-induced expression is independent of nuclear factor E2-related factor 2 (Nrf2). The levels of activator protein-1 (AP-1) mRNA and protein, as well as the amount of phosphorylated c-Jun were significantly increased in mouse liver or Hepa1c1c7 cells treated with either BHA or acrolein. With use of luciferase reporters containing the 5′-flanking sequence of Aldh1a1 (−1963/+27), overexpression of c-Jun resulted in an ≈4-fold induction in luciferase activity, suggesting that c-Jun transactivates the Aldh1a1 promoter as a homodimer and not as a c-Jun/c-Fos heterodimer. Promoter deletion and mutagenesis analyses demonstrated that the AP-1 site at position −758 and possibly −1069 relative to the transcription start site was responsible for c-Jun-mediated transactivation. Electrophoretic mobility shift assay analysis with antibodies against c-Jun and c-Fos showed that c-Jun binds to the proximal AP-1 site at position −758 but not at −1069. Recruitment of c-Jun to this proximal AP-1 site by BHA was confirmed by chromatin immunoprecipitation analysis, indicating that recruitment of c-Jun to the mouse Aldh1a1 gene promoter results in increased transcription. This mode of regulation of an ALDH has not been described before. PMID:22740640

  2. The mitochondrial fission factor dynamin-related protein 1 modulates T-cell receptor signalling at the immune synapse

    PubMed Central

    Baixauli, Francesc; Martín-Cófreces, Noa B; Morlino, Giulia; Carrasco, Yolanda R; Calabia-Linares, Carmen; Veiga, Esteban; Serrador, Juan M; Sánchez-Madrid, Francisco

    2011-01-01

    During antigen-specific T-cell activation, mitochondria mobilize towards the vicinity of the immune synapse. We show here that the mitochondrial fission factor dynamin-related protein 1 (Drp1) docks at mitochondria, regulating their positioning and activity near the actin-rich ring of the peripheral supramolecular activation cluster (pSMAC) of the immune synapse. Mitochondrial redistribution in response to T-cell receptor engagement was abolished by Drp1 silencing, expression of the phosphomimetic mutant Drp1S637D and the Drp1-specific inhibitor mdivi-1. Moreover, Drp1 knockdown enhanced mitochondrial depolarization and T-cell receptor signal strength, but decreased myosin phosphorylation, ATP production and T-cell receptor assembly at the central supramolecular activation cluster (cSMAC). Our results indicate that Drp1-dependent mitochondrial positioning and activity controls T-cell activation by fuelling central supramolecular activation cluster assembly at the immune synapse. PMID:21326213

  3. Low Molecular Weight Hyaluronan-Pulsed Human Dendritic Cells Showed Increased Migration Capacity and Induced Resistance to Tumor Chemoattraction

    PubMed Central

    Rizzo, Manglio; Bayo, Juan; Piccioni, Flavia; Malvicini, Mariana; Fiore, Esteban; Peixoto, Estanislao; García, Mariana G.; Aquino, Jorge B.; Gonzalez Campaña, Ariel; Podestá, Gustavo; Terres, Marcelo; Andriani, Oscar; Alaniz, Laura; Mazzolini, Guillermo

    2014-01-01

    We have shown that ex vivo pre-conditioning of bone marrow-derived dendritic cells (DC) with low molecular weight hyaluronan (LMW HA) induces antitumor immunity against colorectal carcinoma (CRC) in mice. In the present study we investigated the effects of LMW HA priming on human-tumor-pulsed monocytes-derived dendritic cells (DC/TL) obtained from healthy donors and patients with CRC. LMW HA treatment resulted in an improved maturation state of DC/TL and an enhanced mixed leucocyte reaction activity in vivo. Importantly, pre-conditioning of DC/TL with LMW HA increased their ability to migrate and reduced their attraction to human tumor derived supernatants. These effects were associated with increased CCR7 expression levels in DC. Indeed, a significant increase in migratory response toward CCL21 was observed in LMW HA primed tumor-pulsed monocyte-derived dendritic cells (DC/TL/LMW HA) when compared to LWM HA untreated cells (DC/TL). Moreover, LMW HA priming modulated other mechanisms implicated in DC migration toward lymph nodes such as the metalloproteinase activity. Furthermore, it also resulted in a significant reduction in DC migratory capacity toward tumor supernatant and IL8 in vitro. Consistently, LMW HA dramatically enhanced in vivo DC recruitment to tumor-regional lymph nodes and reduced DC migration toward tumor tissue. This study shows that LMW HA –a poorly immunogenic molecule- represents a promising candidate to improve human DC maturation protocols in the context of DC-based vaccines development, due to its ability to enhance their immunogenic properties as well as their migratory capacity toward lymph nodes instead of tumors. PMID:25238610

  4. Inhibitory action of levocetirizine on the production of eosinophil chemoattractants RANTES and eotaxin in vitro and in vivo.

    PubMed

    Kanei, Ayako; Asano, Kazuhito; Kanai, Ken-Ichi; Furuta, Atsuko; Sasaki, Keigo; Suzaki, Harumi

    2014-01-01

    Eosinophils are well known to play essential roles in the development and maintenance of allergic diseases. However, the influence of histamine H1 receptor antagonists on eosinophil functions, especially chemokine production, are not well-defined. Therefore, in the present study, we examined the influence of histamine H1 receptor antagonist on chemokine production by eosinophils through the use of levocetirizine in vitro and in vivo. Eosinophils prepared from mice were stimulated with specific antigens in the presence of different concentrations of levocetirizine. After 24 h, regulated on activation normal T cell expressed and secreted (RANTES) and eotaxin levels in culture supernatants were measured by enzyme-linked immunosorbent assay (ELISA). Patients with Japanese cedar pollinosis were treated with 5 mg levocetirizine once a day for four weeks during the pollen season (February 2012 to April 2012). RANTES and eotaxin levels in nasal secretions were also examined by ELISA. The addition of levocetirizine to eosinophil cultures caused a dose-dependent decrease in the ability of cells to produce RANTES and eotaxin in response to antigen stimulation, and the minimum concentration that caused a significant decrease was 0.05 μM. Although cetirizine also exerted suppressive effects on the production of RANTES and eotaxin by eosinophils, the minimum concentration that caused significant suppression was 0.15 μM, which was three-times higher than that of levocetirizine. Oral administration of levocetirizine for four weeks also reduced RANTES and eotaxin levels in nasal secretions from patients with pollinosis, along with attenuation of clinical symptoms. The ability of levocetirizine to reduce RANTES and eotaxin levels may account, at least in part, for the clinical efficacy of the agent for allergic disorders, including allergic rhinitis. PMID:24982238

  5. Epigenetic inactivation of the canonical Wnt antagonist secreted frizzled-related protein 1 in hepatocellular carcinoma cells.

    PubMed

    Wu, Y; Li, J; Sun, C Y; Zhou, Y; Zhao, Y F; Zhang, S J

    2012-01-01

    Secreted Frizzled-related protein 1 (sFRP1), as one of most important Wnt antagonists, is frequently silenced by promoter hypermethylation in many types of tumor, including hepatocellular carcinoma (HCC). In this study, we aimed to investigate whether restoration of sFRP1 affected HCC metastatic behavior. sFRP1 mRNA expression and promoter methylation in HCC tissues and cell lines were examined using RT-PCR and methylation-specific PCR (MS-PCR), respectively. sFRP1 protein expression was assessed by Western Blot. We generated stable HCC cell line restoration of sFRP1 in HepG2 cells, which naturally do not express detectable sFRP1 mRNA. The effects of exogenous sFRP1 on HepG2 cell invasion were investigated using trans-well assay. Also the effects of sFRP1 re-expression on the β-catenin/T-cell factor-dependent transcription activity was measured by luciferase assay.sFRP1 promoter methylation was frequently observed in HCC tissues (60%) and cell lines (75%). All samples with sFRP1 methylation showed down-regulation of sFRP1 expression in HCC cell lines. Demethylation treatment with 5-aza-20-deoxycytidine in HCC cells restored sFRP1 expression. Restoration of sFRP1 substantially impaired the invasive potentials of HepG2 cells. Moreover, exogenous sFRP1 caused significant decrease of β-catenin/T-cell factor-dependent transcription activity.These findings demonstrate that sFRP1 silencing due to promoter hypermethylation is a major event during tumorigenesis. sFRP1 is also a negative modulator of canonical Wnt signaling, which could contribute to metastasis in HCC progression, thus providing a possible therapeutic strategy against HCC. PMID:22296502

  6. The CXCL12γ Chemokine Displays Unprecedented Structural and Functional Properties that Make It a Paradigm of Chemoattractant Proteins

    PubMed Central

    Rueda, Patricia; Balabanian, Karl; Lagane, Bernard; Staropoli, Isabelle; Chow, Ken; Levoye, Angelique; Laguri, Cedric; Sadir, Rabia; Delaunay, Thierry; Izquierdo, Elena; Pablos, Jose Luis; Lendinez, Elena; Caruz, Antonio; Franco, Diego; Baleux, Françoise; Lortat-Jacob, Hugues; Arenzana-Seisdedos, Fernando

    2008-01-01

    The CXCL12γ chemokine arises by alternative splicing from Cxcl12, an essential gene during development. This protein binds CXCR4 and displays an exceptional degree of conservation (99%) in mammals. CXCL12γ is formed by a protein core shared by all CXCL12 isoforms, extended by a highly cationic carboxy-terminal (C-ter) domain that encompass four overlapped BBXB heparan sulfate (HS)-binding motifs. We hypothesize that this unusual domain could critically determine the biological properties of CXCL12γ through its interaction to, and regulation by extracellular glycosaminoglycans (GAG) and HS in particular. By both RT-PCR and immunohistochemistry, we mapped the localization of CXCL12γ both in mouse and human tissues, where it showed discrete differential expression. As an unprecedented feature among chemokines, the secreted CXCL12γ strongly interacted with cell membrane GAG, thus remaining mostly adsorbed on the plasmatic membrane upon secretion. Affinity chromatography and surface plasmon resonance allowed us to determine for CXCL12γ one of the higher affinity for HS (Kd = 0.9 nM) ever reported for a protein. This property relies in the presence of four canonical HS-binding sites located at the C-ter domain but requires the collaboration of a HS-binding site located in the core of the protein. Interestingly, and despite reduced agonist potency on CXCR4, the sustained binding of CXCL12γ to HS enabled it to promote in vivo intraperitoneal leukocyte accumulation and angiogenesis in matrigel plugs with much higher efficiency than CXCL12α. In good agreement, mutant CXCL12γ chemokines selectively devoid of HS-binding capacity failed to promote in vivo significant cell recruitment. We conclude that CXCL12γ features unique structural and functional properties among chemokines which rely on the presence of a distinctive C-ter domain. The unsurpassed capacity to bind to HS on the extracellular matrix would make CXCL12γ the paradigm of haptotactic proteins, which

  7. Differential distribution of Y-box-binding protein 1 and cold shock domain protein A in developing and adult human brain.

    PubMed

    Bernstein, Hans-Gert; Lindquist, Jonathan A; Keilhoff, Gerburg; Dobrowolny, Henrik; Brandt, Sabine; Steiner, Johann; Bogerts, Bernhard; Mertens, Peter R

    2015-07-01

    The two cold shock domain containing proteins, Y-box-binding protein-1 and cold shock domain protein A were immunolocalized in developing and adult human brain. With the exception of a small population of hypothalamic astrocytes, brain Y-box-binding protein-1 was predominantly found in multiple neurons in the mature human CNS, which might be related to its involvement in neurotransmission and other neuron-associated functions. Cold shock domain protein A was typically observed in astrocytes, oligodendrocytes, choroid plexus epithelia and nerve fibers. However, in circumscribed brain regions as hypothalamus, habenula, and cerebellum, this protein was also expressed in neurons. In the prenatal brain, both proteins were found to be abundantly expressed in radial glial cells, neuroblasts and neurons, which might be an anatomical correlate of the proposed roles of both proteins in cell proliferation and differentiation. In addition, Y-box-binding protein-1 was identified in cultured, lipopolysaccharide-stimulated microglial cells, which underscores its putative role as a mediator in immune and inflammatory processes.

  8. Role of WNT1-inducible-signaling pathway protein 1 in etoposide resistance in lung adenocarcinoma A549 cells

    PubMed Central

    Xu, Yunhua; Lu, Shun

    2015-01-01

    Object: The aim of this study was to explore the role of WNT1-inducible-signaling Pathway Protein 1 (WISP-1) in etoposide resistance in lung adenocarcinoma A549 cells. Methods: WISP-1 overexpression A549 lung adenocarcinoma cell was established. After exposure to ultraviolet (UV) and etoposide, cell viability and apoptosis were evaluated. Moreover, western-blot was employed to examine the expression of apoptotic pathway proteins. In addition, a nude mice tumor model was established to examine the effect of WISP-1 overexpression in vivo and TUNEL staining was used to assess cell apoptosis of tumor tissue. Results: WISP-1 overexpression significantly increased cell viability and decreased cell apoptosis after treatment with UV and etoposide. Decreased expression of Bad and Bax and increased expression of Bcl-2 was found after etoposide treatment in WISP-1 overexpressed cells. A significantly increasing of tumor volume in WISP-1 overexpressed group was found and TUNEL staining revealed that decreased cell apoptosis in WISP-1 overexpressed group. Conclusion: Our results demonstrated that WISP-1 may have a facilitating role in etoposide resistance through increasing cell viability and decreasing cell apoptosis. PMID:26628978

  9. Loss of Selenium-Binding Protein 1 Decreases Sensitivity to Clastogens and Intracellular Selenium Content in HeLa Cells

    PubMed Central

    Zhao, Changhui; Zeng, Huawei; Wu, Ryan T. Y.; Cheng, Wen-Hsing

    2016-01-01

    Selenium-binding protein 1 (SBP1) is not a selenoprotein but structurally binds selenium. Loss of SBP1 during carcinogenesis usually predicts poor prognosis. Because genome instability is a hallmark of cancer, we hypothesize that SBP1 sequesters cellular selenium and sensitizes cancer cells to DNA-damaging agents. To test this hypothesis, we knocked down SBP1 expression in HeLa cervical cancer cells by employing a short hairpin RNA (shRNA) approach. Reduced sensitivity to hydrogen peroxide, paraquat and camptothecin, reactive oxygen species content, and intracellular retention of selenium after selenomethionine treatment were observed in SBP1 shRNA HeLa cells. Results from Western analyses showed that treatment of HeLa cells with selenomethionine resulted in increased SBP1 protein expression in a dose-dependent manner. Knockdown of SBP1 rendered HeLa cells increased expression of glutathione peroxidase-1 but not glutathione peroxidase-4 protein levels and accelerated migration from a wound. Altogether, SBP1 retains supplemental selenium and sensitizes HeLa cancer cells to clastogens, suggesting a new cancer treatment strategy by sequestering selenium through SBP1. PMID:27404728

  10. Ovarian structure protein 1: A sensitive molecular biomarker of gonadal intersex in female Japanese medaka after androgen exposure.

    PubMed

    Abdel-Moneim, Ahmed; Mahapatra, Cecon T; Hatef, Azadeh; Sepúlveda, Maria S

    2015-09-01

    Intersex in gonochoristic fish can be induced after exposure to androgens and estrogens. The main objective of the present study was to identify biomarkers that would be predictive of intersex in Japanese medaka (Oryzias latipes) after exposure to synthetic hormones. First a gene was identified, ovarian structure protein 1 (osp1), with strong female-specific expression during gonadal differentiation. The authors hypothesized that osp1 expression would decrease to male levels in females after the exposure of larvae (15-25 d postfertilization [dpf]) to 17β-trenbolone (TRB; 5 ng/L) and would increase to female levels in males exposed to 17α-ethinylestradiol (EE2; 5 ng/L) and that gonadal intersex would be induced later in life (60 dpf). Tissue distribution and cellular localization of OSP1 was investigated using Western blot and immunohistochemistry. The results indicate that this exposure regime delays testicular maturation in males and development of ovarian intersex in females. Although decreased osp1 expression in females exposed to TRB correlated to changes in ovarian phenotype, up-regulation of osp1 was not observed in males exposed to EE2. In addition, OSP1 was only observed in ovaries and localized in the cytoplasm and follicular layer of immature and mature oocytes. The authors conclude that osp1 is a promising biomarker of androgen exposure and gonadal intersex in female medaka. PMID:25914295

  11. Cloning, characterization and sub-cellular localization of gamma subunit of T-complex protein-1 (chaperonin) from Leishmania donovani.

    PubMed

    Bhaskar; Kumari, Neeti; Goyal, Neena

    2012-12-01

    T-complex protein-1 (TCP1) complex, a chaperonin class of protein, ubiquitous in all genera of life, is involved in intracellular assembly and folding of various proteins. The gamma subunit of TCP1 complex (TCP1γ), plays a pivotal role in the folding and assembly of cytoskeleton protein(s) as an individual or complexed with other subunits. Here, we report for the first time cloning, characterization and