Science.gov

Sample records for osteopontin specifically bound

  1. SUBTYPE-SPECIFIC REGENERATION OF RETINAL GANGLION CELLS FOLLOWING AXOTOMY: EFFECTS OF OSTEOPONTIN AND MTOR SIGNALING

    PubMed Central

    Duan, Xin; Qiao, Mu; Bei, Fengfeng; Kim, In-Jung; He, Zhigang; Sanes, Joshua R.

    2015-01-01

    SUMMARY In mammals, few retinal ganglion cells (RGCs) survive following axotomy and even fewer regenerate axons. This could reflect differential extrinsic influences or the existence of subpopulations that vary in their responses to injury. We tested these alternatives by comparing responses of molecularly distinct subsets of mouse RGCs to axotomy. Survival rates varied dramatically among subtypes, with alpha-RGCs (αRGCs) surviving preferentially. Among survivors, αRGCs accounted for nearly all regeneration following down-regulation of PTEN, which activates the mTOR pathway. αRGCs have uniquely high mTOR signaling levels among RGCs and also selectively express osteopontin (OPN) and receptors for the growth factor, insulin-like growth factor 1 (IGF-1). Administration of OPN plus IGF-1 promotes regeneration as effectively as down-regulation of PTEN; however, regeneration is still confined to αRGCs. Our results reveal dramatic subtype-specific differences in the ability of RGCs to survive and regenerate following injury, and they identify promising agents for promoting axonal regeneration. PMID:25754821

  2. Osteopontin in Immune-mediated Diseases.

    PubMed

    Rittling, S R; Singh, R

    2015-12-01

    Since its initial identification as one of the genes most highly upregulated upon T-cell activation, osteopontin (or Eta-1, as it was designated then) has been demonstrated to have many roles in the regulation of the immune response on multiple levels. It contributes to the development of immune-mediated and inflammatory diseases, and it regulates the host response to infection. In some cases, the mechanisms of these effects have been elucidated, while other mechanistic functions of the protein remain obscure. The protein itself makes these analyses complex, since it binds to a series of different integrins, and in addition to its classically secreted form, an intracellular form of osteopontin has been identified, which participates in several aspects of immune regulation. In this review, we focus on the role of osteopontin in a series of immune-related diseases, particularly those where significant advances have been made in recent years: multiple sclerosis, rheumatoid arthritis, lupus and related diseases, Sjögren's disease, colitis, and 1 area of inflammatory pathology, alcoholic and nonalcoholic liver diseases. A recurring theme in these diseases is a link between osteopontin and pathogenic T cells, particularly T helper 17 cells, where osteopontin produced by dendritic cells supports IL-17 expression, contributing to pathology. In addition, a role for osteopontin in B-cell differentiation is becoming clear. In general, osteopontin contributes to pathology in these diseases, but there are examples where it has a protective role; deciphering the mechanisms underlying these differences and the specific receptors for osteopontin will be a research challenge for the future. Aside from its newly discovered role in the development of Sjögren's disease, the role of osteopontin in inflammatory conditions in the oral cavity is still poorly understood. Elucidation of this role will be of interest. PMID:26341976

  3. Class-specific Error Bounds for Ensemble Classifiers

    SciTech Connect

    Prenger, R; Lemmond, T; Varshney, K; Chen, B; Hanley, W

    2009-10-06

    The generalization error, or probability of misclassification, of ensemble classifiers has been shown to be bounded above by a function of the mean correlation between the constituent (i.e., base) classifiers and their average strength. This bound suggests that increasing the strength and/or decreasing the correlation of an ensemble's base classifiers may yield improved performance under the assumption of equal error costs. However, this and other existing bounds do not directly address application spaces in which error costs are inherently unequal. For applications involving binary classification, Receiver Operating Characteristic (ROC) curves, performance curves that explicitly trade off false alarms and missed detections, are often utilized to support decision making. To address performance optimization in this context, we have developed a lower bound for the entire ROC curve that can be expressed in terms of the class-specific strength and correlation of the base classifiers. We present empirical analyses demonstrating the efficacy of these bounds in predicting relative classifier performance. In addition, we specify performance regions of the ROC curve that are naturally delineated by the class-specific strengths of the base classifiers and show that each of these regions can be associated with a unique set of guidelines for performance optimization of binary classifiers within unequal error cost regimes.

  4. BION web server: predicting non-specifically bound surface ions

    PubMed Central

    Alexov, Emil

    2013-01-01

    Motivation: Ions are essential component of the cell and frequently are found bound to various macromolecules, in particular to proteins. A binding of an ion to a protein greatly affects protein’s biophysical characteristics and needs to be taken into account in any modeling approach. However, ion’s bounded positions cannot be easily revealed experimentally, especially if they are loosely bound to macromolecular surface. Results: Here, we report a web server, the BION web server, which addresses the demand for tools of predicting surface bound ions, for which specific interactions are not crucial; thus, they are difficult to predict. The BION is easy to use web server that requires only coordinate file to be inputted, and the user is provided with various, but easy to navigate, options. The coordinate file with predicted bound ions is displayed on the output and is available for download. Availability: http://compbio.clemson.edu/bion_server/ Supplementary information: Supplementary data are available at Bioinformatics online. Contact: ealexov@clemson.edu PMID:23380591

  5. Expression and distribution of osteopontin in human tissues: widespread association with luminal epithelial surfaces.

    PubMed Central

    Brown, L F; Berse, B; Van de Water, L; Papadopoulos-Sergiou, A; Perruzzi, C A; Manseau, E J; Dvorak, H F; Senger, D R

    1992-01-01

    Osteopontin, a glycoprotein with a glycine-arginine-glycine-aspartate-serine (GRGDS) cell-binding domain, has been described in bone and is also known to be expressed in other organs, particularly kidney. The goal of the present work was to define the distribution of osteopontin synthesis and deposition in a wide variety of normal adult human tissues using a multifaceted approach that included immunohistochemistry, in situ hybridization, and Northern analysis. Immunohistochemical studies have revealed the unexpected finding that osteopontin is deposited as a prominent layer at the luminal surfaces of specific populations of epithelial cells of the gastrointestinal tract, gall bladder, pancreas, urinary and reproductive tracts, lung, breast, salivary glands, and sweat glands. Northern analyses identified gallbladder as a major site of osteopontin gene transcription comparable in magnitude with that of kidney, and immunoblotting identified osteopontin in bile. In situ hybridization localized osteopontin gene transcripts predominantly to the epithelium of a variety of organs as well as to ganglion cells of bowel wall. Osteopontin of epithelial cell origin, like bone-derived osteopontin, promoted GRGDS-dependent cell spreading in attachment assays. We postulate that osteopontin secreted by epithelium binds integrins on luminal surfaces. Collectively, these findings suggest an important role for osteopontin on many luminal epithelial surfaces communicating with the external environment. Images PMID:1421573

  6. Curcumin Veto the Effects of Osteopontin (OPN) Specific Inhibitor on Leukemic Stem Cell Colony Forming Potential via Promotion of OPN Overexpression

    PubMed Central

    Mohammadi, Saeed; Ghaffari, Seyed H.; Shaiegan, Mojgan; Nikogoftar Zarif, Mahin; Nikbakht, Mohsen; Alimoghaddam, Kamran; Ghavamzadeh, Ardeshir

    2016-01-01

    Background: Acute myeloid leukemia (AML) is an immunophenotypically heterogeneous malignant disease, in which CD34 positivity is associated with poor prognosis. Osteopontin (OPN) plays different roles in physiologic and pathologic conditions like: survival, metastasis and cell protection from cytotoxic and apoptotic stimuli. Due to anti-apoptotic effect of OPN in normal and malignant cells, silencing of OPN leads to elevation of sensitivity towards chemotherapeutic agents and attenuates cancer cells migration and invasion. Therefore, the aim of this study was to evaluate OPN roles in modulating curcumin-mediated growth inhibitory on leukemic stem cells (LSCs) colony forming potential and survival in AML cell lines and primary CD34+/CD38- bone marrow-derived AML cells. Materials and Methods: Primary human CD34+/CD38- cells were isolated from bone marrow mononuclear cells of 10 AML patients at initial state of diagnosis, using a CD34 Multi sort kit. The growth inhibitory effects of curcumin (CUR) were evaluated by MTT and colony-formation assays. Apoptosis was analyzed by 7AAD assay in CD34+ KG-1, U937 cell lines and primary isolated cells. Short interfering RNA (siRNA) against OPN was used for OPN silencing in both cell lines and primary AML cells. Then, transfected cells were incubated with/without curcumin. The change in OPN gene expression was examined by Real-time PCR. Results: CUR inhibited proliferation and induced apoptosis in both KG-1 and U937 cells and also primary isolated AML cells. OPN silencing by siRNA increased the susceptibility of KG-1, U937 and primary CD34+/CD38- AML cells to apoptosis. Moreover, soft agar colony assays revealed that silencing of OPN with siRNA significantly decreased colony numbers in LSCs compared with the non-targeting group. Furthermore, CD34+/CD38- populations as a main LSCs compartment through OPN overexpression towards CUR treatment might be nullified the inhibitory effects of OPN siRNA on their survival and colony forming

  7. Control of Rate-Bounded Hybrid Systems with Liveness Specifications

    NASA Technical Reports Server (NTRS)

    Heymann, Michael; Lin, Feng; Meyer, George

    1998-01-01

    In the present paper we examine the control problem for a class of composite hybrid machines (CHMs) that consist of concurrent operation (employing synchronous composition) of elementary hybrid machines (EHMs), that allows both signal sharing and event synchronization. A controller can then be coupled with the plant by means of synchronous composition. We confine our attention to controllers that interact with the system only through event synchronization. We present an initial investigation of synthesis of liveness controllers for hybrid machines. To this end we define open hybrid machines as systems that can interact with the environment through event synchronization and can be therefor be "driven" to their marked configuration by user (controller). Liveness specifications must be associated with timing constraints. We may require that for a specified time limit, every run reach a marked configuration within that time limit. Alternatively, a more relaxed specification may be that, for some (unspecified) global time bound, every run of the system reach a marked configuration within that time bound. Finally, the least restrictive liveness requirement is that every run reach a marked configuration within a finite time limit (but we do not insist on the existence of a global time bound for all runs).

  8. Increased serum osteopontin levels in autistic children: relation to the disease severity.

    PubMed

    Al-ayadhi, Laila Y; Mostafa, Gehan A

    2011-10-01

    Autoimmunity to brain may play an etiopathogenic role in autism. Osteopontin is a pro-inflammatory cytokine that has been shown to play an important role in various autoimmune neuroinflammatory diseases. Osteopontin induces IL-17 production by T-helper 17 lymphocytes, the key players in the pathogenesis of autoimmune disorders. Anti-osteopontin treatment reduces the clinical severity of some autoimmune neuroinflammatory diseases by reducing IL-17 production. We are the first to measure serum osteopontin levels, by ELISA, in 42 autistic children in comparison to 42 healthy-matched children. The relationship between serum osteopontin levels and the severity of autism, which was assessed by using the Childhood Autism Rating Scale (CARS), was also studied. Autistic children had significantly higher serum osteopontin levels than healthy controls (P<0.001). Increased serum osteopontin levels were found in 80.95% (34/42) of autistic children. Children with severe autism had significantly higher serum osteopontin levels than patients with mild to moderate autism (P=0.02). Moreover, serum osteopontin levels of autistic patients had significant positive correlations with CARS (P=0.007). In conclusions, serum osteopontin levels were increased in many autistic children and they were significantly correlated to the severity of autism. Further wide-scale studies are warranted to shed light on the etiopathogenic role of osteopontin in autism and to investigate its relation to IL-17 and brain-specific auto-antibodies, which are indicators of autoimmunity, in these patients. The therapeutic role of anti-osteopontin antibodies in amelioration of autistic manifestations should also be studied. PMID:21521652

  9. Osteopontin involvement in integrin-mediated cell signaling and regulation of expression of alkaline phosphatase during early differentiation of UMR cells.

    PubMed

    Liu, Y K; Uemura, T; Nemoto, A; Yabe, T; Fujii, N; Ushida, T; Tateishi, T

    1997-12-22

    To clarify the function of osteopontin in osteoblast differentiation, we have examined the signal transduction pathway in an osteoblastic cell line (UMR106-6) bound to osteopontin, fibronectin, vitronectin and collagen type I surfaces. This was done by investigating the production and autophosphorylation of focal adhesion kinase (FAK) and the expression of alkaline phosphatase (ALP) at the transcription level. Results suggest that osteopontin was not only responsible for the autophosphorylation of FAK but regulated the expression of ALP, which was strongly correlated with FAK activity. These results suggest that osteopontin might act as a trigger in the early differentiation of osteoblasts. PMID:9450560

  10. Regulation of the pro-inflammatory cytokine osteopontin by GIP in adipocytes - A role for the transcription factor NFAT and phosphodiesterase 3B

    SciTech Connect

    Omar, Bilal; Banke, Elin; Guirguis, Emilia; Aakesson, Lina; Manganiello, Vincent; Lyssenko, Valeriya; Groop, Leif; Gomez, Maria F.; Degerman, Eva

    2012-09-07

    Highlights: Black-Right-Pointing-Pointer GIP stimulates lipogenesis and osteopontin expression in primary adipocytes. Black-Right-Pointing-Pointer GIP-induced osteopontin expression is NFAT-dependent. Black-Right-Pointing-Pointer Osteopontin expression is PDE3-dependent. Black-Right-Pointing-Pointer Osteopontin expression is increased in PDE3B KO mice. -- Abstract: The incretin - glucose-dependent insulinotropic polypeptide (GIP) - and the pro-inflammatory cytokine osteopontin are known to have important roles in the regulation of adipose tissue functions. In this work we show that GIP stimulates lipogenesis and osteopontin expression in primary adipocytes. The GIP-induced increase in osteopontin expression was inhibited by the NFAT (the transcription factor nuclear factor of activated T-cells) inhibitor A-285222. Also, the NFAT kinase glycogen synthase kinase (GSK) 3 was upregulated by GIP. To test whether cAMP might be involved in GIP-mediated effects on osteopontin a number of strategies were used. Thus, the {beta}3-adrenergic receptor agonist CL316,243 stimulated osteopontin expression, an effects which was mimicked by OPC3911, a specific inhibitor of phosphodiesterase 3. Furthermore, treatment of phosphodiesterase 3B knock-out mice with CL316,243 resulted in a dramatic upregulation of osteopontin in adipose tissue which was not the case in wild-type mice. In summary, we delineate mechanisms by which GIP stimulates osteopontin in adipocytes. Given the established link between osteopontin and insulin resistance, our data suggest that GIP by stimulating osteopontin expression, also could promote insulin resistance in adipocytes.

  11. Scaling of the Specific Heat of Bounded ^4He Films.

    NASA Astrophysics Data System (ADS)

    Mehta, S.; Gasparini, F. M.

    1998-03-01

    We report new measurements of the specific heat of a thick helium film confined between two silicon wafers(S. Mehta, W.Y. Yu, A. Petrou, J. Lipa, D. Bishop and F.M. Gasparini, Czechoslovak J. Phys. 46, 133(1996).). The latest data are for a 0.048μm thick film, which extend our earlier measurements on similar films(S. Mehta and F.M. Gasparini, Phys. Rev. Lett. 78, 2596(1997).) by a factor of 2. The latest experimental cell has the new oxide pattern, and has allowed us to make measurements into the superfluid region. When these data are analyzed to test predictions of correlation-length scaling(M.E. Fisher in Critical Phenomenon, Proc. 51^st) Enrico Fermi Summer School, Varenna, Italy, ed. M.S. Green (Academic Press, NY, 1971)., they collapse well onto the earlier data for ν=0.6705(L.S. Goldner and G. Ahlers, Phys. Rev. B45, 13129(1992).) both above and below T_λ. Some issues regarding scaling still remain near the heat capacity maximum. The experimental techniques used to obtain these data will also be discussed.

  12. Osteopontin: A non-invasive parameter of portal hypertension and prognostic marker of cirrhosis

    PubMed Central

    Bruha, Radan; Jachymova, Marie; Petrtyl, Jaromir; Dvorak, Karel; Lenicek, Martin; Urbanek, Petr; Svestka, Tomislav; Vitek, Libor

    2016-01-01

    AIM: To investigate the relationship between osteopontin plasma concentrations and the severity of portal hypertension and to assess osteopontin prognostic value. METHODS: A cohort of 154 patients with confirmed liver cirrhosis (112 ethylic, 108 men, age 34-72 years) were enrolled in the study. Hepatic venous pressure gradient (HVPG) measurement and laboratory and ultrasound examinations were carried out for all patients. HVPG was measured using a standard catheterization method with the balloon wedge technique. Osteopontin was measured using the enzyme-linked immunosorbent assay (ELISA) method in plasma. Patients were followed up with a specific focus on mortality. The control group consisted of 137 healthy age- and sex- matched individuals. RESULTS: The mean value of HVPG was 16.18 ± 5.6 mmHg. Compared to controls, the plasma levels of osteopontin in cirrhotic patients were significantly higher (P < 0.001). The plasma levels of osteopontin were positively related to HVPG (P = 0.0022, r = 0.25) and differed among the individual Child-Pugh groups of patients. The cut-off value of 80 ng/mL osteopontin distinguished patients with significant portal hypertension (HVPG above 10 mmHg) at 75% sensitivity and 63% specificity. The mean follow-up of patients was 3.7 ± 2.6 years. The probability of cumulative survival was 39% for patients with HVPG > 10 mmHg and 65% for those with HVPG ≤ 10 mmHg (P = 0.0086, odds ratio (OR), 2.92, 95% confidence interval (CI): 1.09-7.76). Osteopontin showed a similar prognostic value to HVPG. Patients with osteopontin values above 80 ng/mL had significantly lower cumulative survival compared to those with osteopontin ≤ 80 ng/mL (37% vs 56%, P = 0.00035; OR = 2.23, 95%CI: 1.06-4.68). CONCLUSION: Osteopontin is a non-invasive parameter of portal hypertension that distinguishes patients with clinically significant portal hypertension. It is a strong prognostic factor for survival. PMID:27022226

  13. Autoantibodies against complement C1q specifically target C1q bound on early apoptotic cells.

    PubMed

    Bigler, Cornelia; Schaller, Monica; Perahud, Iryna; Osthoff, Michael; Trendelenburg, Marten

    2009-09-01

    Autoantibodies against complement C1q (anti-C1q) are frequently found in patients with systemic lupus erythematosus (SLE). They strongly correlate with the occurrence of severe lupus nephritis, suggesting a pathogenic role in SLE. Because anti-C1q are known to recognize a neoepitope on bound C1q, but not on fluid-phase C1q, the aim of this study was to clarify the origin of anti-C1q by determining the mechanism that renders C1q antigenic. We investigated anti-C1q from serum and purified total IgG of patients with SLE and hypocomplementemic urticarial vasculitis as well as two monoclonal human anti-C1q Fab from a SLE patient generated by phage display. Binding characteristics, such as their ability to recognize C1q bound on different classes of Igs, on immune complexes, and on cells undergoing apoptosis, were analyzed. Interestingly, anti-C1q did not bind to C1q bound on Igs or immune complexes. Neither did we observe specific binding of anti-C1q to C1q bound on late apoptotic/necrotic cells when compared with binding in the absence of C1q. However, as shown by FACS analysis and confocal microscopy, anti-C1q specifically targeted C1q bound on early apoptotic cells. Anti-C1q were found to specifically target C1q bound on cells undergoing apoptosis. Our observations suggest that early apoptotic cells are a major target of the autoimmune response in SLE and provide a direct link between human SLE, apoptosis, and C1q. PMID:19648280

  14. Lateral diffusion of specific antibodies bound to lipid monolayers on alkylated substrates.

    PubMed Central

    Subramaniam, S; Seul, M; McConnell, H M

    1986-01-01

    We have measured the lateral mobility of fluoresceinated monoclonal IgG antibodies bound specifically to a spin label lipid hapten in phospholipid monolayers supported on alkylated silicon oxide surfaces. Dimyristoyl phosphatidylcholine and dipalmitoyl phosphatidylcholine monolayers containing 5 mol% of the lipid hapten were transferred by conventional Langmuir-Blodgett techniques onto substrates alkylated with hydrocarbon chains containing 10, 16, and 18 carbon atoms. We show that the diffusion of the bound antibodies depends on their lateral density, the composition of the lipid monolayer, and the nature of lipid coupling to hydrocarbon chains on the alkylated substrate. Antibody diffusion coefficients at low antibody densities are within a factor of 2 of those displayed by the lipid hapten in the absence of the bound antibody. High antibody densities result in reduced antibody mobility, but the lateral diffusion of unbound lipids is unaffected. Images PMID:3006037

  15. Gradients of substrate-bound laminin orient axonal specification of neurons

    PubMed Central

    Dertinger, Stephan K. W.; Jiang, Xingyu; Li, Zhiying; Murthy, Venkatesh N.; Whitesides, George M.

    2002-01-01

    Little is known about the influence of substrate-bound gradients on neuronal development, since it has been difficult to fabricate gradients over the distances typically required for biological studies (a few hundred micrometers). This article demonstrates a generally applicable technique for the fabrication of substrate-bound gradients of proteins with complex shapes, using laminar flows in microchannels. Gradients that range from pure laminin to pure BSA were formed in solution by using a network of microchannels, and these proteins were allowed to adsorb onto a homogeneous layer of poly-l-lysine. Rat hippocampal neurons were cultivated on these substrate-bound gradients. Analysis of optical images of these neurons showed that axon specification is oriented in the direction of increasing surface density of laminin. Linear gradients in laminin adsorbed from a gradient in solution having a slope of ∇[laminin] > about 0.06 μg (ml⋅μm)−1 (defined by dividing the change of concentration of laminin in solution over the distance of the gradient) orient axon specification, whereas those with ∇[laminin] < about 0.06 μg (ml⋅μm)−1 have no effect. PMID:12237407

  16. Rare attributes in finite universe: Hypotheses testing specification and exact randomized upper confidence bounds

    SciTech Connect

    Wright, T.

    1993-03-01

    When attributes are rare and few or none are observed in the selected sample from a finite universe, sampling statisticians are increasingly being challenged to use whatever methods are available to declare with high probability or confidence that the universe is near or completely attribute-free. This is especially true when the attribute is undesirable. Approximations such as those based on normal theory are frequently inadequate with rare attributes. For simple random sampling without replacement, an appropriate probability distribution for statistical inference is the hypergeometric distribution. But even with the hypergeometric distribution, the investigator is limited from making claims of attribute-free with high confidence unless the sample size is quite large using nonrandomized techniques. In the hypergeometric setting with rare attributes, exact randomized tests of hypothesis a,re investigated to determine the effect on power of how one specifies the null hypothesis. In particular, specifying the null hypothesis as zero attributes does not always yield maximum possible power. We also consider the hypothesis specification question under complex sampling designs including stratified random sampling and two-stage cluster sampling (one case involves random selection at first stage and another case involves probability proportional to size without replacement selection at first stage). Also under simple random sampling, this article defines and presents a simple algorithm for the construction of exact ``randomized`` upper confidence bounds which permit one to possibly report tighter bounds than those exact bounds obtained using ``nonrandomized`` methods.

  17. Rare attributes in finite universe: Hypotheses testing specification and exact randomized upper confidence bounds

    SciTech Connect

    Wright, T.

    1993-03-01

    When attributes are rare and few or none are observed in the selected sample from a finite universe, sampling statisticians are increasingly being challenged to use whatever methods are available to declare with high probability or confidence that the universe is near or completely attribute-free. This is especially true when the attribute is undesirable. Approximations such as those based on normal theory are frequently inadequate with rare attributes. For simple random sampling without replacement, an appropriate probability distribution for statistical inference is the hypergeometric distribution. But even with the hypergeometric distribution, the investigator is limited from making claims of attribute-free with high confidence unless the sample size is quite large using nonrandomized techniques. In the hypergeometric setting with rare attributes, exact randomized tests of hypothesis a,re investigated to determine the effect on power of how one specifies the null hypothesis. In particular, specifying the null hypothesis as zero attributes does not always yield maximum possible power. We also consider the hypothesis specification question under complex sampling designs including stratified random sampling and two-stage cluster sampling (one case involves random selection at first stage and another case involves probability proportional to size without replacement selection at first stage). Also under simple random sampling, this article defines and presents a simple algorithm for the construction of exact randomized'' upper confidence bounds which permit one to possibly report tighter bounds than those exact bounds obtained using nonrandomized'' methods.

  18. Osteopontin Deficiency Increases Bone Fragility but Preserves Bone Mass

    PubMed Central

    Thurner, Philipp J.; Chen, Carol G.; Ionova-Martin, Sophi; Sun, Luling; Harman, Adam; Porter, Alexandra; Ager, Joel W.; Ritchie, Robert O.; Alliston, Tamara

    2010-01-01

    The ability of bone to resist catastrophic failure is critically dependent upon the material properties of bone matrix, a composite of hydroxyapatite, collagen type I, and noncollagenous proteins. These properties include elastic modulus, hardness, and fracture toughness. Like other aspects of bone quality, matrix material properties are biologically-defined and can be disrupted in skeletal disease. While mineral and collagen have been investigated in greater detail, the contribution of noncollagenous proteins such as osteopontin to bone matrix material properties remains unclear. Several roles have been ascribed to osteopontin in bone, many of which have the potential to impact material properties. To elucidate the role of osteopontin in bone quality, we evaluated the structure, composition, and material properties of bone from osteopontin-deficient mice and wild-type littermates at several length scales. Most importantly, the results show that osteopontin deficiency causes a 30% decrease in fracture toughness, suggesting an important role for OPN in preventing crack propagation. This significant decline in fracture toughness is independent of changes in whole bone mass, structure, or matrix porosity. Using nanoindentation and quantitative backscattered electron imaging to evaluate osteopontin-deficient bone matrix at the micrometer level, we observed a significant reduction in elastic modulus and increased variability in calcium concentration. Matrix heterogeneity was also apparent at the ultrastructural level. In conclusion, we find that osteopontin is essential for the fracture toughness of bone, and reduced toughness in osteopontin-deficient bone may be related to the increased matrix heterogeneity observed at the micro-scale. By exploring the effects of osteopontin-deficiency on bone matrix material properties, composition and organization, this study suggests that reduced fracture toughness is one mechanism by which loss of noncollagenous proteins contribute

  19. Substrate specificity of mitochondrial intermediate peptidase analysed by a support-bound peptide library

    PubMed Central

    Marcondes, M.F.M.; Alves, F.M.; Assis, D.M.; Hirata, I.Y.; Juliano, L.; Oliveira, V.; Juliano, M.A.

    2015-01-01

    The substrate specificity of recombinant human mitochondrial intermediate peptidase (hMIP) using a synthetic support-bound FRET peptide library is presented. The collected fluorescent beads, which contained the hydrolysed peptides generated by hMIP, were sequenced by Edman degradation. The results showed that this peptidase presents a remarkable preference for polar uncharged residues at P1 and P1′ substrate positions: Ser = Gln > Thr at P1 and Ser > Thr at P1′. Non-polar residues were frequent at the substrate P3, P2, P2′ and P3′ positions. Analysis of the predicted MIP processing sites in imported mitochondrial matrix proteins shows these cleavages indeed occur between polar uncharged residues. Previous analysis of these processing sites indicated the importance of positions far from the MIP cleavage site, namely the presence of a hydrophobic residue (Phe or Leu) at P8 and a polar uncharged residue (Ser or Thr) at P5. To evaluate this, additional kinetic analyses were carried out, using fluorogenic substrates synthesized based on the processing sites attributed to MIP. The results described here underscore the importance of the P1 and P1′ substrate positions for the hydrolytic activity of hMIP. The information presented in this work will help in the design of new substrate-based inhibitors for this peptidase. PMID:26082885

  20. Substrate specificity of mitochondrial intermediate peptidase analysed by a support-bound peptide library.

    PubMed

    Marcondes, M F M; Alves, F M; Assis, D M; Hirata, I Y; Juliano, L; Oliveira, V; Juliano, M A

    2015-01-01

    The substrate specificity of recombinant human mitochondrial intermediate peptidase (hMIP) using a synthetic support-bound FRET peptide library is presented. The collected fluorescent beads, which contained the hydrolysed peptides generated by hMIP, were sequenced by Edman degradation. The results showed that this peptidase presents a remarkable preference for polar uncharged residues at P1 and P1' substrate positions: Ser = Gln > Thr at P1 and Ser > Thr at P1'. Non-polar residues were frequent at the substrate P3, P2, P2' and P3' positions. Analysis of the predicted MIP processing sites in imported mitochondrial matrix proteins shows these cleavages indeed occur between polar uncharged residues. Previous analysis of these processing sites indicated the importance of positions far from the MIP cleavage site, namely the presence of a hydrophobic residue (Phe or Leu) at P8 and a polar uncharged residue (Ser or Thr) at P5. To evaluate this, additional kinetic analyses were carried out, using fluorogenic substrates synthesized based on the processing sites attributed to MIP. The results described here underscore the importance of the P1 and P1' substrate positions for the hydrolytic activity of hMIP. The information presented in this work will help in the design of new substrate-based inhibitors for this peptidase. PMID:26082885

  1. Posttranslational modifications of bovine osteopontin: identification of twenty-eight phosphorylation and three O-glycosylation sites.

    PubMed Central

    Sørensen, E. S.; Højrup, P.; Petersen, T. E.

    1995-01-01

    Osteopontin (OPN) is a multiphosphorylated glycoprotein found in bone and other normal and malignant tissues, as well as in the physiological fluids urine and milk. The present study demonstrates that bovine milk osteopontin is phosphorylated at 27 serine residues and 1 threonine residue. Phosphoamino acids were identified by a combination of amino acid analysis, sequence analysis of S-ethylcysteine-derivatized phosphopeptides, and mass spectrometric analysis. Twenty-five phosphoserines and one phosphothreonine were located in Ser/Thr-X-Glu/Ser(P)/Asp motifs, and two phosphoserines were found in the sequence Ser-X-X-Glu/Ser(P). These sequence motifs are identical with the recognition sequences of mammary gland casein kinase and casein kinase II, respectively. Examination of the phosphorylation pattern revealed that the phosphorylations were clustered in groups of approximately three spanned by unphosphorylated regions of 11-32 amino acids. This pattern is probably of importance in the multiple functions of OPN involving interaction with Ca2+ and inorganic calcium salts. Furthermore, three O-glycosylated threonines (Thr 115, Thr 124, and Thr 129) have been identified in a threonine- and proline-rich region of the protein. Three putative N-glycosylation sites (Asn 63, Asn 85, and Asn 193) are present in bovine osteopontin, but sequence and mass spectrometric analysis showed that none of these asparagines were glycosylated in bovine mammary gland osteopontin. Alignment analysis showed that the majority of the phosphorylation sites in bovine osteopontin as well as all three O-glycosylation sites were conserved in other mammalian sequences. This conservation of serines, even in otherwise less well-conserved regions of the protein, indicates that the phosphorylation of osteopontin at specific sites is essential for the function of the protein. PMID:8535240

  2. Quantification of ferritin bound iron in human serum using species-specific isotope dilution mass spectrometry.

    PubMed

    Ren, Yao; Walczyk, Thomas

    2014-09-01

    Ferritin is a hollow sphere protein composed of 24 subunits that can store up to 4500 iron atoms in its inner cavity. It is mainly found in the liver and spleen but also in serum at trace levels. Serum ferritin is considered as the best single indicator in assessing body iron stores except liver or bone marrow biopsy. However, it is confounded by other disease conditions. Ferritin bound iron (FBI) and ferritin saturation have been suggested as more robust biomarkers. The current techniques for FBI determination are limited by low antibody specificity, low instrument sensitivity and possible analyte losses during sample preparation. The need for a highly sensitive and reliable method is widely recognized. Here we describe a novel technique to detect serum FBI using species-specific isotope dilution mass spectrometry (SS-IDMS). [(57)Fe]-ferritin was produced by biosynthesis and in vitro labeling with the (57)Fe spike in the form of [(57)Fe]-citrate after cell lysis and heat treatment. [(57)Fe]-ferritin for sample spiking was further purified by fast liquid protein chromatography. Serum ferritin and added [(57)Fe]-ferritin were separated from other iron species by ultrafiltration followed by isotopic analysis of FBI using negative thermal ionization mass spectrometry. Repeatability of our assay is 8% with an absolute detection limit of 18 ng FBI in the sample. As compared to other speciation techniques, SS-IDMS offers maximum control over sample losses and species conversion during analysis. The described technique may therefore serve as a reference technique for clinical applications of FBI as a new biomarker for assessing body iron status. PMID:25008269

  3. Crystal Structure of Human Thymine DNA Glycosylase Bound to DNA Elucidates Sequence-Specific Mismatch Recognition

    SciTech Connect

    Maiti, A.; Morgan, M.T.; Pozharski, E.; Drohat, A.C.

    2009-05-19

    Cytosine methylation at CpG dinucleotides produces m{sup 5}CpG, an epigenetic modification that is important for transcriptional regulation and genomic stability in vertebrate cells. However, m{sup 5}C deamination yields mutagenic G{center_dot}T mispairs, which are implicated in genetic disease, cancer, and aging. Human thymine DNA glycosylase (hTDG) removes T from G{center_dot}T mispairs, producing an abasic (or AP) site, and follow-on base excision repair proteins restore the G{center_dot}C pair. hTDG is inactive against normal A{center_dot}T pairs, and is most effective for G{center_dot}T mispairs and other damage located in a CpG context. The molecular basis of these important catalytic properties has remained unknown. Here, we report a crystal structure of hTDG (catalytic domain, hTDG{sup cat}) in complex with abasic DNA, at 2.8 {angstrom} resolution. Surprisingly, the enzyme crystallized in a 2:1 complex with DNA, one subunit bound at the abasic site, as anticipated, and the other at an undamaged (nonspecific) site. Isothermal titration calorimetry and electrophoretic mobility-shift experiments indicate that hTDG and hTDG{sup cat} can bind abasic DNA with 1:1 or 2:1 stoichiometry. Kinetics experiments show that the 1:1 complex is sufficient for full catalytic (base excision) activity, suggesting that the 2:1 complex, if adopted in vivo, might be important for some other activity of hTDG, perhaps binding interactions with other proteins. Our structure reveals interactions that promote the stringent specificity for guanine versus adenine as the pairing partner of the target base and interactions that likely confer CpG sequence specificity. We find striking differences between hTDG and its prokaryotic ortholog (MUG), despite the relatively high (32%) sequence identity.

  4. Intranasal Osteopontin for Rodent Germinal Matrix Hemorrhage.

    PubMed

    Malaguit, Jay; Casel, Darlene; Dixon, Brandon; Doycheva, Desislava; Tang, Jiping; Zhang, John H; Lekic, Tim

    2016-01-01

    Germinal matrix hemorrhage (GMH) is the most common and devastating neurological problem of premature infants. Current treatment is largely ineffective and GMH has been nonpreventable. Osteopontin (OPN) is an endogenous protein that has been shown to be neuroprotective, however, it has not been tested in GMH. P7 neonatal rats were subjected to stereotactic ganglionic eminence collagenase infusion. Groups were as follows: (1) sham, (2) GMH + vehicle, (3) GMH + intranasal OPN. Seventy-two hours later, the animals were evaluated using righting reflex, blood-brain barrier (BBB) permeability by Evans blue dye leakage, brain water content, and hemoglobin assay. Intranasal OPN improved outcomes after GMH by attenuation of brain swelling, BBB function, re-bleeding, and neurological outcomes. OPN may play an important role in enhancing neuroprotective brain signaling following GMH. These observed effects may offer novel possibilities for therapy in this patient population. PMID:26463952

  5. Effect of osteopontin on the initial adhesion of dental bacteria.

    PubMed

    Schlafer, Sebastian; Meyer, Rikke L; Sutherland, Duncan S; Städler, Brigitte

    2012-12-28

    Bacterial biofilms are involved in numerous infections of the human body, including dental caries. While conventional therapy of biofilm diseases aims at eradication and mechanical removal of the biofilms, recent therapeutic approaches target the mechanisms of biofilm formation and bacterial adhesion in particular. The effect of bovine milk osteopontin, a highly phosphorylated whey protein, on adhesion of Streptococcus mitis, Streptococcus sanguinis, and Actinomyces naeslundii, three prominent colonizers in dental biofilms, to saliva-coated surfaces was investigated. While adhesion of A. naeslundii was not affected by osteopontin, a strong, dose-dependent reduction in the number of adhering S. mitis was shown. No difference in bacterial adhesion was observed for caseinoglycomacropeptide, another phosphorylated milk protein. Osteopontin did not affect bacterial viability, but changed bacterial surface hydrophobicity, and may be suggested to prevent the adhesins of S. mitis from interacting with their salivary receptors. The antiadhesive effect of osteopontin may be useful for caries prevention. PMID:23167781

  6. Specificity of neomycin analogues bound to the packaging region of human immunodeficiency virus type 1 RNA.

    PubMed

    McPike, Mark P; Goodisman, Jerry; Dabrowiak, James C

    2004-04-15

    -neo must be directed away from the binding pocket when these analogues are bivalently bound to RNA. This study showed how variations in the structure of aminoglycosides lead to different binding specificity to part of the packaging region of HIV-1. Such knowledge will be important in design of drugs to target this region. PMID:15051052

  7. Effects of microgravity on urinary osteopontin

    NASA Technical Reports Server (NTRS)

    Hoyer, J. R.; Pietrzyk, R. A.; Liu, H.; Whitson, P. A.

    1999-01-01

    Increased risk of renal stone formation during space flight has been linked primarily to increased calcium excretion from bone demineralization induced by space flight. Other factors contributing to increased risk include increased urinary calcium oxalate supersaturation, while urinary citrate, magnesium and volume are all decreased. The aim of this study was to increase the predictive value of stone risk profiles for crew members during space flight by evaluating the excretion of urinary protein inhibitors of calcium crystallization so that more comprehensive stone risk profiles could relate mineral saturation to the concentrations of inhibitor proteins. Levels of urinary osteopontin (uropontin) are reported in a series of 14 astronauts studied before, during, and after space flights. During space flight, a compensatory increase in uropontin excretion was not observed. However, the uropontin excretion of a majority of astronauts was increased during the period after space flight and was maximal at 2 wk after landing. The downward shift in the molecular size of uropontin observed in samples obtained during space flight was shown to result from storage at ambient temperature during flight, rather than an effect of microgravity on uropontin synthesis.

  8. Role of Osteopontin in Liver Diseases

    PubMed Central

    Wen, Yankai; Jeong, Seogsong; Xia, Qiang; Kong, Xiaoni

    2016-01-01

    Osteopontin (OPN), a multifunctional protein, is involved in numerous pathological conditions including inflammation, immunity, angiogenesis, fibrogenesis and carcinogenesis in various tissues. Extensive studies have elucidated the critical role of OPN in cell signaling such as regulation of cell proliferation, migration, inflammation, fibrosis and tumor progression. In the liver, OPN interacts with integrins, CD44, vimentin and MyD88 signaling, thereby induces infiltration, migration, invasion and metastasis of cells. OPN is highlighted as a chemoattractant for macrophages and neutrophils during injury in inflammatory liver diseases. OPN activates hepatic stellate cells (HSCs) to exert an enhancer in fibrogenesis. The role of OPN in hepatocellular carcinoma (HCC) has also generated significant interests, especially with regards to its role as a diagnostic and prognostic factor. Interestingly, OPN acts an opposing role in liver repair under different pathological conditions. This review summarizes the current understanding of OPN in liver diseases. Further understanding of the pathophysiological role of OPN in cellular interactions and molecular mechanisms associated with hepatic inflammation, fibrosis and cancer may contribute to the development of novel strategies for clinical diagnosis, monitoring and therapy of liver diseases. PMID:27570486

  9. Calcium-phosphate-osteopontin particles for caries control.

    PubMed

    Schlafer, Sebastian; Birkedal, Henrik; Olsen, Jakob; Skovgaard, Jonas; Sutherland, Duncan S; Wejse, Peter L; Nyvad, Bente; Meyer, Rikke L

    2016-01-01

    Caries is caused by acid production in biofilms on dental surfaces. Preventing caries therefore involves control of microorganisms and/or the acid produced. Here, calcium-phosphate-osteopontin particles are presented as a new approach to caries control. The particles are made by co-precipitation and designed to bind to bacteria in biofilms, impede biofilm build-up without killing the microflora, and release phosphate ions to buffer bacterial acid production if the pH decreases below 6. Analysis of biofilm formation and pH in a five-species biofilm model for dental caries showed that treatment with particles or pure osteopontin led to less biofilm formation compared to untreated controls or biofilms treated with osteopontin-free particles. The anti-biofilm effect can thus be ascribed to osteopontin. The particles also led to a slower acidification of the biofilm after exposure to glucose, and the pH always remained above 5.5. Hence, calcium-phosphate-osteopontin particles show potential for applications in caries control. PMID:26923119

  10. MicroRNA-127-5p regulates osteopontin expression and osteopontin-mediated proliferation of human chondrocytes

    PubMed Central

    Tu, Min; Li, Yusheng; Zeng, Chao; Deng, Zhenhan; Gao, Shuguang; Xiao, Wenfeng; Luo, Wei; Jiang, Wei; Li, Liangjun; Lei, Guanghua

    2016-01-01

    The aim of this study was to determine the specific microRNA (miRNA) that regulates expression of osteopontin (OPN) in osteoarthritis (OA). The potential regulatory miRNAs for OPN messenger RNA (mRNA) were predicted by miRNA prediction programs. Among eight potential regulatory miRNAs, miR-220b, miR-513a-3p and miR-548n increased, while miR-181a, miR-181b, miR-181c, miR-181d and miR-127-5p decreased in OA patients. miRNA-127-5p mimics suppressed OPN production as well as the activity of a reporter construct containing the 3′-UTR of human OPN mRNA. In addition, mutation of miR-127-5p binding site in the 3′-UTR of OPN mRNA abolished miR-127-5p-mediated repression of reporter activity. Conversely, treatment with miR-127-5p inhibitor increased reporter activity and OPN production. Interestingly, miR-127-5p inhibited proliferation of chondrocytes through OPN. In conclusion, miRNA-127-5p is an important regulator of OPN in human chondrocytes and may contribute to the development of OA. PMID:27126955

  11. N-Glycan structures of an osteopontin from human bone.

    PubMed

    Masuda, K; Takahashi, N; Tsukamoto, Y; Honma, H; Kohri, K

    2000-02-24

    N-Glycan structures of osteopontin (a bone matrix protein) from human bone (lumbar vertabrate) are reported in detail. Asn-linked glycan portion was released from 100 microg of osteopontin by digestion with glycoamidase A (from sweet almond), and the reducing ends of the N-glycans were reductively aminated with 2-aminopyridine. The derivatized N-glycans were separated and structurally identified by a multidimensional mapping technique on HPLC columns. Two major N-glycan structures were also confirmed by mass spectrometry. The proposed structures are shown below. The result should permit future comparison with the N-glycan structures of osteopontins obtained from other sources (kidney tissues, macrophages, urinary stones, human milk, etc.). PMID:10679288

  12. Inhibition of hydroxyapatite formation by osteopontin phosphopeptides.

    PubMed Central

    Pampena, David A; Robertson, Karen A; Litvinova, Olga; Lajoie, Gilles; Goldberg, Harvey A; Hunter, Graeme K

    2004-01-01

    Osteopontin (OPN) is an acidic phosphoglycoprotein that is believed to function in the prevention of soft tissue calcification. In vitro studies have shown that OPN can inhibit the formation of hydroxyapatite (HA) and other biologically relevant crystal phases, and that this inhibitory activity requires phosphorylation of the protein; however, it is not known which phosphorylated residues are involved. We have synthesized peptides corresponding to four phosphoserine-containing sequences in rat OPN: OPN7-17, containing phosphoserines 10 and 11; OPN41-52, containing phosphoserines 46 and 47; OPN248-264, containing phosphoserines 250, 257 and 262; and OPN290-301, containing phosphoserines 295-297. The abilities of these peptides to inhibit de novo HA formation were determined using a constant-composition autotitration assay. All four OPN phosphopeptides caused a dose-dependent increase in nucleation lag time, but did not significantly affect subsequent formation of the crystals. However, OPN41-52 (inhibitory constant 73.5 min/microM) and OPN290-301 (72.2 min/microM) were approx. 4 times more potent inhibitors than OPN7-17 (19.7 min/microM) and OPN247-264 (16.3 min/microM). 'Scrambling' the amino acid sequence of OPN290-301 resulted in decreased potency (45.6 min/microM), whereas omission of the phosphate groups from this peptide caused a greater decrease (5.20 min/microM). These findings have identified phosphorylated sequences that are important for the ability of rat bone OPN to inhibit HA crystal formation, and suggest that negative-charge density is an important factor in this activity. PMID:14678013

  13. Simulation of detachment of specifically bound particles from surfaces by shear flow.

    PubMed Central

    Kuo, S C; Hammer, D A; Lauffenburger, D A

    1997-01-01

    The receptor-mediated adhesion of cells to ligand-coated surfaces is important in many physiological and biotechnological processes. Previously, we measured the detachment of antibody-coated spheres from counter-antibody- and protein A-coated substrates using a radial-flow detachment assay and were able to relate mechanical adhesion strength to chemical binding affinity (Kuo and Lauffenburger, Biophys. J. 65:2191-2200 (1993)). In this paper, we use "adhesive dynamics" to simulate the detachment of antibody-coated hard spheres from a ligand-coated substrate. We modeled the antibody-ligand (either counter-antibody or protein A) bonds as adhesive springs. In the simulation as in the experiments, beads attach to the substrate under static conditions. Flow is then initiated, and detachment is measured by the significant displacement of previously bound particles. The model can simulate the effects of many parameters on cell detachment, including hydrodynamic stresses, receptor number, ligand density, reaction rates between receptor and ligand, and stiffness and reactive compliance of the adhesive springs. The simulations are compared with experimental detachment data, thus relating measured bead adhesion strength to molecular properties of the adhesion molecules. The simulations accurately recreated the logarithmic dependence of adhesion strength on affinity of receptor-ligand recognition, which was seen in experiments and predicted by analytic theory. In addition, we find the value of the reactive compliance, the parameter which relates the strain of a bond to its rate of breakage, that gives the best match between theory and experiment to be 0.01. Finally, we analyzed the effect of varying either the forward or reverse rate constants as different ways to achieve the same affinity, and showed that adhesion strength depends uniquely on the equilibrium affinity, not on the kinetics of binding. Given that attachment is independent of affinity, detachment and attachment

  14. Wogonin suppresses osteopontin expression in adipocytes by activating PPARα

    PubMed Central

    Zhang, Ye-min; Li, Ming-xin; Tang, Zhao; Wang, Chang-hua

    2015-01-01

    Aim: Wogonin (5,7-dihydroxy-8-methoxyflavone), a major bioactive compound of the flavonoid family, is commonly extracted from the traditional Chinese medicine Scutellaria baicalensis and possesses antioxidant and anti-inflammatory activities and is assumed to have anti-diabetes function. Indeed, a current study has shown that it can possibly treat metabolic disorders such as those found in db/db mice. However, the underlying molecular mechanism remains largely unclear. The aim of this study was to investigate the impact of wogonin on osteopontin (OPN) expression in adipose tissue from type 1 diabetic mice and in 3T3-L1 adipocytes. Methods: Type 1 diabetes was induced by streptozotocin (STZ) injection. 3T3-L1 preadipocytes were converted to 3T3-L1 adipocytes through treatment with insulin, dexamethasone, and 3-isobutyl-1-methylxanthine (IBMX). Western blot analysis and RT-PCR were performed to detect protein expression and mRNA levels, respectively. Results: Wogonin treatment suppressed the increase in serum OPN levels and reduced OPN expression in adipose tissue from STZ-induced type 1 diabetic mice. Administration of wogonin enhanced PPARα expression and activity. Silencing of PPARα diminished the inhibitory effects of wogonin on OPN expression in 3T3-L1 adipocytes. Furthermore, the levels of c-Fos and phosphorylated c-Jun were reduced in wogonin-treated adipose tissue and 3T3-L1 adipocytes. In addition, wogonin treatment dramatically mitigated p38 MAPK phosphorylation. Pharmacological inhibition of p38 MAPK by its specific inhibitor SB203580 increased PPARα activity and decreased OPN expression. Conclusion: Our results suggest that wogonin downregulated OPN expression in adipocytes through the inhibition of p38 MAPK and the sequential activation of the PPARα pathway. Given the adverse effects of high OPN levels on metabolism, our results provide evidence for the potential administration of wogonin as a treatment for diabetes. PMID:26073326

  15. N-Acetyl-beta-D-glucopyranosylamine 6-phosphate is a specific inhibitor of glycogen-bound protein phosphatase 1.

    PubMed Central

    Board, M

    1997-01-01

    Previous work has shown that the C-1-substituted glucose-analogue N-acetyl-beta-D-glucopyranosylamine (1-GlcNAc) is a competitive inhibitor of glycogen phosphorylase (GP) and stimulates the inactivation of this enzyme by GP phosphatase. In addition to its effects on GP, 1-GlcNAc also prevents the glucose-led activation of glycogen synthase (GS) in whole hepatocytes. Such an effect on GS was thought to be due to the formation of 1-GlcNAc-6-P by the action of glucokinase within the hepatocyte [Board, Bollen, Stalmans, Kim, Fleet and Johnson (1995) Biochem. J. 311, 845-852]. To investigate this possibility further, a pure preparation of 1-GlcNAc-6-P was synthesized. The effects of the phosphorylated glucose analogue on the activity of protein phosphatase 1 (PP1), the enzyme responsible for dephosphorylation and activation of GS, are reported. During the present study, 1-GlcNAc-6-P inhibited the activity of the glycogen-bound form of PP1, affecting both the GSb phosphatase and GPa phosphatase activities. A level of 50% inhibition of GSb phosphatase activity was achieved with 85 microM 1-GlcNAc-6-P in the absence of Glc-6-P and with 135 microM in the presence of 10 mM Glc-6-P. At either Glc-6-P concentration, 500 microM 1-GlcNAc-6-P completely inhibited activity. The Glc-6-P stimulation of the GPa phosphatase activity of PP1 was negated by 1-GlcNAc-6-P but there was no inhibition of the basal rate in the absence of Glc-6-P. 1-GlcNAc-6-P inhibition was specific for the glycogen-bound form of PP1 and did not inhibit the GSb phosphatase activity of the cytosolic form of the enzyme. The present work explains our previous observations on the inactivating effects on GS of incubating whole hepatocytes with 1-GlcNAc. These observations have their basis in the inhibition of glycogen-bound PP1 by 1-GlcNAc-6-P. A novel inhibitor of PP1, specific for the glycogen-bound form of the enzyme, is presented. PMID:9371733

  16. Quantitation of specific antibodies bound to feline leukemia virus in the plasma of pet cats.

    PubMed

    Snyder, H W; Singhal, M C; Yoshida, L H; Jones, F R

    1985-08-01

    A method is described for determining levels of circulating immune complexes (CIC) composed of feline leukemia virus (FeLV) antigens and corresponding antibodies in plasma of persistently-infected pet cats. The procedure is based on the ability of high-titered heterologous anti-FeLV serum to chase cat anti-FeLV IgG from dissociated CIC by successfully competing for binding of free antigen. The eluted cat antibody is then collected and quantitated. In a study of cats in the process of clearing persistent FeLV infections, measured levels of FeLV-specific CIC correlated well with fluctuating levels of free FeLV antigen and antibody. The Raji cell assay for CIC in those cats was of comparatively little value in following the clearance of the virus, presumably because that assay does not distinguish between CIC containing viral and those containing non-viral antigens. The method described can be adapted to studies of specific immune complexes associated with a variety of syndromes, provided that the antigen eliciting the immune response is known. PMID:2995795

  17. Osteopontin facilitates angiogenesis, accumulation of osteoclasts, and resorption in ectopic bone.

    PubMed

    Asou, Y; Rittling, S R; Yoshitake, H; Tsuji, K; Shinomiya, K; Nifuji, A; Denhardt, D T; Noda, M

    2001-03-01

    Osteoclastic bone resorption requires a number of complex steps that are under the control of local regulatory molecules. Osteopontin is expressed in osteoclasts and is also present in bone matrix; however, its biological function has not been fully understood. To elucidate the role of osteopontin in the process of osteoclastic bone resorption, we conducted ectopic bone implantation experiments using wild-type and osteopontin knockout mouse. In the wild-type group, bone discs from calvariae implanted ectopically in muscle were resorbed, and their mass was reduced by 25% within 4 weeks. In contrast, the mass of the bone discs from calvariae of osteopontin knockout mice was reduced by only 5% when implanted in osteopontin knockout mice. Histological analyses indicated that the number of osteoclasts associated with the implanted bones was reduced in the osteopontin knockout mice. As osteopontin deficiency does not suppress osteoclastogenesis per se, we further examined vascularization immunohistologically and found that the number of vessels containing CD31-positive endothelial cells around the bone discs implanted in muscle was reduced in the osteopontin knockout mice. Furthermore, sc implantation assays indicated that the length and branching points of the newly formed vasculatures associated with the bone discs were also reduced in the absence of osteopontin. In this assay, tartrate-resistant acid phosphatase-positive area of the bone discs was also reduced in the osteopontin knockout mice, indicating further the link between the osteopontin-dependent vascularization and osteoclast accumulation. The bone resorption defect could be rescued by topical administration of recombinant osteopontin to the bones implanted in muscle. These observations indicate that osteopontin is required for efficient vascularization by the hemangiogenic endothelial cells and subsequent osteoclastic resorption of bones. PMID:11181551

  18. Hepcidin, the hormone of iron metabolism, is bound specifically to alpha-2-macroglobulin in blood.

    PubMed

    Peslova, Gabriela; Petrak, Jiri; Kuzelova, Katerina; Hrdy, Ivan; Halada, Petr; Kuchel, Philip W; Soe-Lin, Shan; Ponka, Prem; Sutak, Robert; Becker, Erika; Huang, Michael Li-Hsuan; Suryo Rahmanto, Yohan; Richardson, Des R; Vyoral, Daniel

    2009-06-11

    Hepcidin is a major regulator of iron metabolism. Hepcidin-based therapeutics/diagnostics could play roles in hematology in the future, and thus, hepcidin transport is crucial to understand. In this study, we identify alpha2-macroglobulin (alpha2-M) as the specific hepcidin-binding molecule in blood. Interaction of 125I-hepcidin with alpha2-M was identified using fractionation of plasma proteins followed by native gradient polyacrylamide gel electrophoresis and mass spectrometry. Hepcidin binding to nonactivated alpha2-M displays high affinity (Kd 177 +/- 27 nM), whereas hepcidin binding to albumin was nonspecific and displayed nonsaturable kinetics. Surprisingly, the interaction of hepcidin with activated alpha2-M exhibited a classical sigmoidal binding curve demonstrating cooperative binding of 4 high-affinity (Kd 0.3 microM) hepcidin-binding sites. This property probably enables efficient sequestration of hepcidin and its subsequent release or inactivation that may be important for its effector functions. Because alpha2-M rapidly targets ligands to cells via receptor-mediated endocytosis, the binding of hepcidin to alpha2-M may influence its functions. In fact, the alpha2-M-hepcidin complex decreased ferroportin expression in J774 cells more effectively than hepcidin alone. The demonstration that alpha2-M is the hepcidin transporter could lead to better understanding of hepcidin physiology, methods for its sensitive measurement and the development of novel drugs for the treatment of iron-related diseases. PMID:19380872

  19. Quantification of ferritin-bound iron in plant samples by isotope tagging and species-specific isotope dilution mass spectrometry.

    PubMed

    Hoppler, Matthias; Zeder, Christophe; Walczyk, Thomas

    2009-09-01

    Ferritin is nature's predominant iron storage protein. The molecule consists of a hollow protein shell composed of 24 subunits which is capable of storing up to 4500 iron atoms per molecule. Recently, this protein has been identified as a target molecule for increasing iron content in plant staple foods in order to combat dietary iron deficiency, a major public health problem in developing countries. Here, we present a novel technique for quantification of ferritin-bound iron in edible plant seeds using species-specific isotope dilution mass spectrometry (IDMS) by means of a biosynthetically produced (57)Fe-labeled ferritin spike and negative thermal ionization mass spectrometry (NTIMS). Native plant ferritin and added spike ferritin were extracted in 20 mM Tris buffer (pH 7.4) and separated by anion exchange chromatography (DEAE Sepharose), followed by isotopic analysis by thermal ionization mass spectrometry. The chosen IDMS approach was critically evaluated by assessing the (i) efficiency of analyte extraction, (ii) identical behavior of spike and analyte, and (iii) potential iron isotope exchange with natural iron. Repeatabilities that can be achieved are on the order of <5% RSD for quintuplicate analyses at an absolute detection limit of 60 ng of ferritin-bound iron for plant seeds. Studies in six different legumes revealed ferritin-iron contents ranging from 15% of total iron in red kidney beans up to 69% in lentils. PMID:19653660

  20. Association of the osteopontin rs1126616 polymorphism and a higher serum osteopontin level with lupus nephritis

    PubMed Central

    SALIMI, SAEEDEH; NOORA, MEHRANGIZ; NABIZADEH, SIMA; REZAEI, MAHNAZ; SHAHRAKI, HOSSAIN; MILAD, MOHAMMADOO-KHORASSANI; NAGHAVI, ANOOSH; FARAJIAN-MASHHADI, FARZANEH; ZAKERI, ZAHRA; SANDOUGHI, MAHNAZ

    2016-01-01

    Osteopontin (OPN) is a chemokine-like glycoprotein that has a prominent role in regulating inflammation and immunity. OPN polymorphisms and elevated OPN levels are associated with systemic lupus erythematosus (SLE) in several populations. The aim of present study was to evaluate the association between the OPN rs1126616 polymorphism and OPN level with SLE susceptibility. A total of 163 SLE patients and 180 age-, gender- and ethnically matched controls were genotyped for the rs1126616 polymorphism by the polymerase chain reaction-restriction fragment length polymorphism method. Serum OPN levels were assayed by the enzyme-linked immunosorbent assay. There was no association between the OPN rs1126616 C/T polymorphism and SLE. The frequency of the OPN rs1126616 CT genotype was significantly higher in SLE patients with nephritis compared to SLE patients without nephritis and controls. Additionally, the frequency of TT genotypes was higher in SLE patients with nephritis compared to controls. The serum OPN levels were significantly higher in SLE patients compared to controls (50.6±22 vs. 35.6±15.8 ng/ml, P<0.001). Increased serum OPN levels were observed in SLE patients with lupus nephritis and joint symptoms. There was no correlation between OPN levels and the OPN rs1126616 polymorphism. The present data suggest that the CT and TT genotypes of the OPN rs1126616 polymorphism could be a risk factor for lupus nephritis. The OPN level is associated with SLE and certain SLE manifestations. However, there was no association between the OPN rs1126616 C/T polymorphism and SLE susceptibility. PMID:26998275

  1. Normal rat kidney cells secrete both phosphorylated and nonphosphorylated forms of osteopontin showing different physiological properties

    SciTech Connect

    Nemir, M.; DeVouge, M.W.; Mukherjee, B.B. )

    1989-10-25

    We have reported previously that the 69-kDa major phosphoprotein, secreted by normal rat kidney (NRK) cells, is osteopontin, a glycosylated bone matrix protein. Here we show that this 69-kDa osteopontin is secreted by NRK cells in both phosphorylated (pp69) and nonphosphorylated (np69) forms, with estimated isoelectric points of 3.8 and 4.5, respectively. Electrophoretic analysis of radioiodinated cell surface proteins immunoprecipitated with an anti-69-kDa osteopontin serum, demonstrates that the 69-kDa osteopontin is also present on the cell surface, but only its phosphorylated form (pp69) shows such cell surface association. Because osteopontin mediates cell adhesion and spreading, and contains an Arg-Gly-Asp-Ser cell-binding sequence, our observations strongly suggest that the cell surface localization of pp69 osteopontin is receptor-mediated, and the modification by phosphorylation may be crucial for its receptor binding activity. We also report that antisera directed against either fibronectin or 69-kDa osteopontin co-immunoprecipitate both np69 osteopontin and fibronectin as a heat-dissociable complex. In contrast, pp69 osteopontin does not co-precipitate with fibronectin. Furthermore, compared to NRK cells, vanadyl sulfate-treated NRK cells which acquire a reversible transformed phenotype, including anchorage-independent growth, show increased levels of pp69 on the cell surface, concomitant with significantly decreased levels of pp69 and elevated levels of np69 in the conditioned media. The data presented here establish transformation sensitivity of NRK cell-secreted osteopontin with respect to its secretion and cell surface localization, and demonstrate that phosphorylated and nonphosphorylated forms of osteopontin have different physiological properties, which may regulate the functional roles of this extracellular matrix protein.

  2. Mapping proteoglycan-bound water in cartilage: improved specificity of matrix assessment using multiexponential transverse relaxation analysis

    PubMed Central

    Reiter, David A.; Roque, Remigio A.; Lin, Ping-Chang; Irrechukwu, Onyi; Doty, Stephen; Longo, Dan; Pleshko, Nancy; Spencer, Richard G.

    2012-01-01

    Association of MR parameters with cartilage matrix components remains an area of ongoing investigation. Multiexponential analysis of non-localized transverse relaxation data has previously been used to quantify water compartments associated with matrix macromolecules in cartilage. We extend this to mapping the proteoglycan-bound water fraction (wPG) in cartilage, using mature and young bovine nasal cartilage model systems, towards the goal of matrix component-specific imaging. wPG from mature and young bovine nasal cartilage was 0.3±0.04 and 0.22±0.06, respectively, in agreement with biochemically-derived proteoglycan content and proteoglycan-to-water weight ratios. Fourier transform infrared imaging spectroscopic-derived proteoglycan maps normalized by water content (IR-PGww) showed spatial correspondence with wPG maps. Extensive simulation analysis demonstrated that the accuracy and precision of our determination of wPG was within 2%, which is substantially smaller than the observed tissue differences. Our results demonstrate the feasibility of performing imaging-based multiexponential analysis of transverse relaxation data to map proteoglycan in cartilage. PMID:21264931

  3. Loop Shaping Control Design for a Supersonic Propulsion System Model Using Quantitative Feedback Theory (QFT) Specifications and Bounds

    NASA Technical Reports Server (NTRS)

    Connolly, Joseph W.; Kopasakis, George

    2010-01-01

    This paper covers the propulsion system component modeling and controls development of an integrated mixed compression inlet and turbojet engine that will be used for an overall vehicle Aero-Propulso-Servo-Elastic (APSE) model. Using previously created nonlinear component-level propulsion system models, a linear integrated propulsion system model and loop shaping control design have been developed. The design includes both inlet normal shock position control and jet engine rotor speed control for a potential supersonic commercial transport. A preliminary investigation of the impacts of the aero-elastic effects on the incoming flow field to the propulsion system are discussed, however, the focus here is on developing a methodology for the propulsion controls design that prevents unstart in the inlet and minimizes the thrust oscillation experienced by the vehicle. Quantitative Feedback Theory (QFT) specifications and bounds, and aspects of classical loop shaping are used in the control design process. Model uncertainty is incorporated in the design to address possible error in the system identification mapping of the nonlinear component models into the integrated linear model.

  4. Osteopontin as a marker of weight loss in lung cancer.

    PubMed

    Karadag, Fisun; Gulen, Sule T; Karul, Aslihan B; Kilicarslan, Naciye; Ceylan, Emel; Kuman, Nilgun K; Cildag, Orhan

    2011-12-01

    Although the role of osteopontin (OPN) in tumorigenesis and invasiveness is well-known, its role in systemic consequences of lung cancer has not been studied yet. The objective of the current study was to assess the value of osteopontin as a marker of weight loss in relation to systemic inflammation in non-small cell lung cancer (NSCLC) patients. A total of 63 male NSCLC patients (stage III and IV) and 25 age and sex-matched controls were included. The NSCLC patients were further divided into subgroups depending on whether they had > 5% weight loss in the last 6 months or not. Serum OPN and TNF-α concentrations were measured by ELISA using commercially available kits. Serum C-reactive protein (CRP) concentration was measured by the turbidimetric method. OPN (p = 0.001) and CRP (p < 0.001) concentrations were significantly higher in lung cancer patients compared to controls whereas TNF-α concentrations were similar in cancer and control groups (p = 0.063). There were 33 NSCLC patients (52.4%) with weight loss. Serum OPN concentration was found to be higher in this weight-losing group (p = 0.042). CRP concentration was also higher in the weight-losing group but the difference was not statistically significant (p = 0.246). TNF-α concentrations were similar in both subgroups (p = 0.094). In correlation tests, there was a positive correlation between OPN and CRP (r = 0.299, p = 0.044), but no correlation was detected between OPN and TNF-α (r = − 0.009, p = 0.930). A negative correlation was detected between OPN and BMI (r = − 0.246, p = 0.048). In addition to being an indicator of systemic inflammation in lung cancer patients, osteopontin may also be an indicator of weight loss. PMID:22017168

  5. Functional atomic force microscopy investigation of osteopontin affinity for silicon stabilized tricalcium phosphate bioceramic surfaces.

    PubMed

    Pietak, Alexis M; Sayer, Michael

    2006-01-01

    Resorbable silicon stabilized tricalcium phosphate (Si-TCP)-based bioceramics are characterized from a biological perspective by measuring the intermolecular interaction force between osteopontin (OPN) protein and the material surface using atomic force microscopy (AFM). OPN protein was covalently bound to silicon nitride AFM tips and adsorption and adhesion forces were measured in an electrolyte with a composition similar to that of physiological fluids. A strong relationship exists between the adhesion force of OPN on the material surface, the number of adherent osteoclasts (OC) and the resorption of the material. OPN adhesion is strongest on hydroxyapatite (HA) surfaces, or in samples that induce a HA-like surface through a precipitation reaction in electrolytic media. It is proposed that the increased biological response of the Si-TCP phase can be attributed in part to its reactivity in a physiological electrolyte, which involves a rapid conversion to a calcium deficient HA phase with a corresponding increase in the adhesion strength of OPN to the material, with a consequentially higher OC resorption response. PMID:16011845

  6. Gender-Specific Models of Work-Bound Korean Adolescents' Social Supports and Career Adaptability on Subsequent Job Satisfaction

    ERIC Educational Resources Information Center

    Han, Hyojung; Rojewski, Jay W.

    2015-01-01

    A Korean national database, the High School Graduates Occupational Mobility Survey, was used to examine the influence of perceived social supports (family and school) and career adaptability on the subsequent job satisfaction of work-bound adolescents 4 months after their transition from high school to work. Structural equation modeling analysis…

  7. Inhibition of Cellular Adhesion by Immunological Targeting of Osteopontin Neoepitopes Generated through Matrix Metalloproteinase and Thrombin Cleavage

    PubMed Central

    Jürets, Alexander; Le Bras, Marie; Staffler, Günther; Stein, Gesine; Leitner, Lukas; Neuhofer, Angelika; Tardelli, Matteo; Turkof, Edvin; Zeyda, Maximilian; Stulnig, Thomas M.

    2016-01-01

    Osteopontin (OPN), a secreted protein involved in inflammatory processes and cancer, induces cell adhesion, migration, and activation of inflammatory pathways in various cell types. Cells bind OPN via integrins at a canonical RGD region in the full length form as well as to a contiguous cryptic site that some have shown is unmasked upon thrombin or matrix metalloproteinase cleavage. Thus, the adhesive capacity of osteopontin is enhanced by proteolytic cleavage that may occur in inflammatory conditions such as obesity, atherosclerosis, rheumatoid arthritis, tumor growth and metastasis. Our aim was to inhibit cellular adhesion to recombinant truncated proteins that correspond to the N-terminal cleavage products of thrombin- or matrix metalloproteinase-cleaved OPN in vitro. We specifically targeted the cryptic integrin binding site with monoclonal antibodies and antisera induced by peptide immunization of mice. HEK 293 cells adhered markedly stronger to truncated OPN proteins than to full length OPN. Without affecting cell binding to the full length form, the raised monoclonal antibodies specifically impeded cellular adhesion to the OPN fragments. Moreover, we show that the peptides used for immunization were able to induce antisera, which impeded adhesion either to all OPN forms, including the full-length form, or selectively to the corresponding truncated recombinant proteins. In conclusion, we developed immunological tools to selectively target functional properties of protease-cleaved OPN forms, which could find applications in treatment and prevention of various inflammatory diseases and cancers. PMID:26840958

  8. Osteopontin-A Master Regulator of Epithelial-Mesenchymal Transition.

    PubMed

    Kothari, Anai N; Arffa, Matthew L; Chang, Victor; Blackwell, Robert H; Syn, Wing-Kin; Zhang, Jiwang; Mi, Zhiyong; Kuo, Paul C

    2016-01-01

    Osteopontin (OPN) plays an important functional role in both physiologic and pathologic states. OPN is implicated in the progression of fibrosis, cancer, and metastatic disease in several organ systems. The epithelial-mesenchymal transition (EMT), first described in embryology, is increasingly being recognized as a significant contributor to fibrotic phenotypes and tumor progression. Several well-established transcription factors regulate EMT and are conserved across tissue types and organ systems, including TWIST, zinc finger E-box-binding homeobox (ZEB), and SNAIL-family members. Recent literature points to an important relationship between OPN and EMT, implicating OPN as a key regulatory component of EMT programs. In this review, OPN's interplay with traditional EMT activators, both directly and indirectly, will be discussed. Also, OPN's ability to restructure the tissue and tumor microenvironment to indirectly modify EMT will be reviewed. Together, these diverse pathways demonstrate that OPN is able to modulate EMT and provide new targets for directing therapeutics. PMID:27023622

  9. Tumorigenic properties of alternative osteopontin isoforms in mesothelioma

    SciTech Connect

    Ivanov, Sergey V.; Ivanova, Alla V.; Goparaju, Chandra M.V.; Chen, Yuanbin; Beck, Amanda; Pass, Harvey I.

    2009-05-08

    Osteopontin (SPP1) is an inflammatory cytokine that we previously characterized as a diagnostic marker in patients with asbestos-induced malignant mesothelioma (MM). While SPP1 shows both pro- and anti-tumorigenic biological effects, little is known about the molecular basis of these activities. In this study, we demonstrate that while healthy pleura possesses all three differentially spliced SPP1 isoforms (A-C), in clinical MM specimens isoform A is markedly up-regulated and predominant. To provide a clue to possible functions of the SPP1 isoforms we next performed their functional evaluation via transient expression in MM cell lines. As a result, we report that isoforms A-C demonstrate different activities in cell proliferation, wound closure, and invasion assays. These findings suggest different functions for SPP1 isoforms and underline pro-tumorigenic properties of isoforms A and B.

  10. IgE-dependent humoral immune response in Echinococcus multilocularis infection: circulating and basophil-bound specific IgE against Echinococcus antigens in patients with alveolar echinococcosis.

    PubMed Central

    Vuitton, D A; Bresson-Hadni, S; Lenys, D; Flausse, F; Liance, M; Wattre, P; Miguet, J P; Capron, A

    1988-01-01

    Clinical symptoms of immediate-type hypersensitivity (ITH) and specific IgE against Echinococcus granulosus antigens are frequently present in patients with hydatid cysts. In alveolar echinococcosis (AE) due to E. multilocularis, clinical manifestations related to ITH have never been reported. The IgE-dependent humoral immune response was evaluated in 30 patients with AE. Circulating specific IgE (sIgE) were determined with two different methods of radio-allergo-sorbent test. Serum sIgE were determined sequentially in 18 patients over 15 months. Specific IgE bound to circulating basophils were assessed with two tests in vitro, measuring specific degranulation and histamine release. The respective abilities of E. granulosus and E. multilocularis antigens to reveal bound and circulating IgE antibodies were also assayed. Despite the absence of clinical symptoms of ITH and the frequent lack of circulating sIgE, an immunological response involving IgE was always present in human AE: basophil-bound sIgE were revealed in every patient by histamine release and degranulation tests; these tests were constantly negative in control subjects. Echinococcus granulosus extracts were more effective for detecting circulating sIgE; however E. multilocularis antigenic preparation induced a histamine release significantly higher than E. granulosus extracts. These results suggest that IgE-dependent humoral immune response could play a role in the host-parasite relationship in AE. Moreover, the sensitivity of the tests used to detect basophil-bound sIgE was higher than that of the usual serological tests, and the basophil degranulation test could be used to confirm diagnosis of AE in endemic countries. PMID:2450708

  11. Plants increase arsenic in solution but decrease the non-specifically bound fraction in the rhizosphere of an alkaline, naturally rich soil.

    PubMed

    Obeidy, Carole; Bravin, Matthieu N; Bouchardon, Jean-Luc; Conord, Cyrille; Moutte, Jacques; Guy, Bernard; Faure, Olivier

    2016-04-01

    We aimed at determining the major physical-chemical processes that drive arsenic (As) dynamic in the rhizosphere of four species (Holcus lanatus, Dittrichia viscosa, Lotus corniculatus, Plantago lanceolata) tested for phytostabilization. Experiments were performed with an alkaline soil naturally rich in As. Composition of the soil solution of planted and unplanted pots was monitored every 15 days for 90 days, with a focus on the evolution of As concentrations in solution and in the non-specifically bound (i.e. easily exchangeable) fraction. The four species similarly increased As concentration in solution, but decreased As concentration in the non-specifically bound fraction. The major part (60%) of As desorbed from the non-specifically bound fraction in planted pots was likely redistributed on the less available fractions of As on the solid phase. A second part (35%) of desorbed As was taken up by plants. The minor part (5%) of desorbed As supplied As increase in solution. To conclude, plants induced a substantial redistribution of As on the less available fractions in the rhizosphere, as expected in phytostabilization strategies. Plants however concomitantly increased As concentration in the rhizosphere solution which may contribute to As transfer through plant uptake and leaching. PMID:26707185

  12. Early and persistent up-regulated expression of renal cortical osteopontin in experimental hydronephrosis.

    PubMed Central

    Diamond, J. R.; Kees-Folts, D.; Ricardo, S. D.; Pruznak, A.; Eufemio, M.

    1995-01-01

    The mechanical disturbance after unilateral ureteral obstruction (UUO) is a nonimmune stimulus that is capable of eliciting a florid macrophage infiltration of the kidney and subsequent post-inflammatory renal scarring. Osteopontin has potential chemoattractant activity and, for this reason, we delineated the kinetics of its expression in the renal cortex of rats with UUO. Whole body X-irradiation and reversal of UUO were utilized as interventional maneuvers to give additional pathobiological insight into this protein's role in the response of the kidneys to ureteral obstruction. Increased osteopontin mRNA levels in obstructed kidneys versus contralateral unobstructed specimens were evident as early as 4 hours after UUO and steadily increased at 12, 24, 48, and 96 hours after UUO. Both X-irradiation and reversal of UUO failed to significantly modulate renal cortical osteopontin mRNA expression at all of the above time points. Paralleling the increments in renal cortical osteopontin mRNA levels were significant elevations in the cortical renal interstitial macrophage number, which was significantly diminished by previous X-irradiation but not reversal of UUO. Focal labeling of osteopontin was noted in both tubular and Bowman's capsular epithelium in obstructed kidneys as early as 4 hours after UUO, whereas, in the contralateral unobstructed specimens, there was only faint staining in Bowman's capsule. By 96 hours after UUO, obstructed kidneys exhibited intense, diffuse staining for osteopontin in both tubules and Bowman's capsule. Osteopontin's immunolocalization was not modulated by X-irradiation or reversal of UUO. These data support the contention that osteopontin is involved in the accumulation of macrophages within the peritubular and periglomerular interstitium in the obstructed renal cortex. Images Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 Figure 6 Figure 7 PMID:7778684

  13. Osteopontin stimulates gelsolin-associated phosphoinositide levels and phosphatidylinositol triphosphate-hydroxyl kinase.

    PubMed Central

    Chellaiah, M; Hruska, K

    1996-01-01

    Based on previous studies demonstrating activation of phosphatidylinositol 3-hydroxyl kinase (PI3-kinase) and stimulation of a change in cell shape, we examined the effect of osteopontin on the association of phospholipids with gelsolin, an actin-capping/severing protein. Osteopontin stimulated a rapid increase in phosphatidylinositol bisphosphate and phosphatidylinositol triphosphate levels associated with gelsolin in Triton-soluble fractions of cell lysates. The increased levels of phosphatidylinositol triphosphate associated with gelsolin were due to stimulation of PI3-kinase activity associated with gelsolin in the Triton-soluble fractions, and they were blocked by the PI3-kinase inhibitor wortmannin. Osteopontin stimulated translocation of PI3-kinase from the Triton-insoluble to Triton-soluble gelsolin. Osteopontin also decreased Triton-soluble gelsolin/actin complexes consistent with actin uncapping, and increased F-actin levels, which were also blocked by wortmannin. The osteopontin effects were mediated through binding to the alpha(v)beta 3 integrin. Taken together, our studies indicate that osteopontin/alpha(v)beta 3-mediated changes in gelsolin-associated phosphoinositide levels and PI3-kinase activity are related to stimulation of F-actin formation in osteoclasts. Images PMID:8744948

  14. Nanoscale patterning of membrane-bound proteins formed through curvature-induced partitioning of phase-specific receptor lipids.

    PubMed

    Ogunyankin, Maria O; Huber, Dale L; Sasaki, Darryl Y; Longo, Marjorie L

    2013-05-21

    This work describes a technique for forming high-density arrays and patterns of membrane-bound proteins through binding to a curvature-organized compositional pattern of metal-chelating lipids (Cu(2+)-DOIDA or Cu(2+)-DSIDA). In this bottom-up approach, the underlying support is an e-beam formed, square lattice pattern of hemispheres. This curvature pattern sorts Cu(2+)-DOIDA to the 200 nm hemispherical lattice sites of a 600 nm × 600 nm unit cell in Ld - Lo phase separated lipid multibilayers. Binding of histidine-tagged green fluorescent protein (His-GFP) creates a high density array of His-GFP-bound pixels localized to the square lattice sites. In comparison, the negative pixel pattern is created by sorting Cu(2+)-DSIDA in Ld - Lβ' phase separated lipid multibilayers to the flat grid between the lattice sites followed by binding to His-GFP. Lattice defects in the His-GFP pattern lead to interesting features such as pattern circularity. We also observe defect-free arrays of His-GFP that demonstrate perfect arrays can be formed by this method suggesting the possibility of using this approach for the localization of various active molecules to form protein, DNA, or optically active molecular arrays. PMID:23642033

  15. Different Poses for Ligand and Chaperone in Inhibitor Bound Hsp90 and GRP94: Implications for Paralog-specific Drug Design

    PubMed Central

    Immormino, Robert M.; Metzger, Louis E.; Reardon, Patrick N.; Dollins, D. Eric; Blagg, Brian S.J.; Gewirth, Daniel T.

    2009-01-01

    Hsp90 chaperones contain an N-terminal ATP binding site that has been effectively targeted by competitive inhibitors. Despite the myriad of inhibitors, none to date have been designed to bind specifically to just one of the four mammalian hsp90 paralogs, which are cytoplasmic Hsp90α and β, ER GRP94, and mitochondrial Trap-1. Given that each of the hsp90 paralogs is responsible for chaperoning a distinct set of client proteins, specific targeting of one hsp90 paralog may result in higher efficacy and therapeutic control. Specific inhibitors may also help elucidate the biochemical roles of each hsp90 paralog. Here we present side by side comparisons of the structures of yeast Hsp90 and mammalian GRP94, bound to the pan-hsp90 inhibitors Geldanamycin and Radamide. These structures reveal paralog specific differences in the Hsp90 and GRP94 conformations in response to Geldanamycin binding. We also report significant variation in the pose and disparate binding affinities for the Geldanamycin-Radicicol chimera Radamide when bound to the two paralogs, which may be exploited in the design of paralog-specific inhibitors. PMID:19361515

  16. Increased osteopontin and liver stiffness measurement by transient elastography in biliary atresia

    PubMed Central

    Honsawek, Sittisak; Chayanupatkul, Maneerat; Chongsrisawat, Voranush; Vejchapipat, Paisarn; Poovorawan, Yong

    2010-01-01

    AIM: To analyze plasma osteopontin levels and liver stiffness using transient elastography in postoperative biliary atresia (BA) children compared with healthy controls. METHODS: Thirty children with postoperative BA and 10 normal controls were enrolled. The patients were categorized into two groups according to their jaundice status. Plasma levels of osteopontin were determined using commercially available enzyme-linked immunosorbent assay. Liver stiffness was measured by using transient elastography (Fibroscan). Ten validated Fibroscan measurements were performed in each patient and control with the result expressed in kilopascals (kPa). RESULTS: Plasma osteopontin was significantly elevated in BA children compared with that of healthy controls (47.0 ± 56.4 ng/mL vs 15.1 ± 15.0 ng/mL, P = 0.01). The liver stiffness measurement was markedly elevated in the patients with BA compared with that of controls (26.9 ± 24.6 kPa vs 3.9 ± 0.7 kPa, P = 0.001). Subgroup analysis showed that the BA patients with jaundice had more pronounced plasma osteopontin levels than those without jaundice (87.1 ± 61.6 ng/mL vs 11.9 ± 6.1 ng/mL, P = 0.001). Furthermore, the mean liver stiffness was significantly greater in the jaundiced BA patients compared with non-jaundiced patients (47.7 ± 21.8 kPa vs 8.7 ± 3.0 kPa, P = 0.001). Additionally, plasma osteopontin was positively related to serum total bilirubin (r = 0.64, P < 0.001). There was also a correlation between plasma osteopontin and liver stiffness values (r = 0.60, P < 0.001). CONCLUSION: High plasma osteopontin positively correlated with degree of hepatic fibrosis and could be used as a biochemical parameter reflecting disease severity in postoperative BA children. PMID:21086566

  17. Altered specificity of single-chain antibody fragments bound to pandemic H1N1-2009 influenza virus after conversion of the phage-bound to the soluble form

    PubMed Central

    2012-01-01

    Background In 2009, a novel influenza A/H1N1 virus (H1N1pdm) quickly spread worldwide and co-circulated with then-existing seasonal H1N1 virus (sH1N1). Distinguishing between these 2 viruses was necessary to better characterize the epidemiological properties of the emergent virus, including transmission patterns, pathogenesis, and anti-influenza drug resistance. This situation prompted us to develop a point-of-care virus differentiation system before entering the 2009–2010 influenza season. Aiming to establish H1N1pdm-specific detection tools rapidly, we employed phage display libraries to select H1N1pdm-specific single-chain variable fragments (scFvs). Findings Human single-fold scFv libraries (Tomlinson I + J) underwent selection for the ability to bind H1N1pdm virus particles. Three rounds of panning brought 1152 phage-bound scFvs, of which 58 clones reacted with H1N1pdm specifically or preferentially over sH1N1 in an enzyme-linked immunosorbent assay (ELISA). After conversion of the scFvs to soluble form, 7 clones demonstrating high/stable expression were finally obtained. However, all the soluble scFvs except No. 29 were found to have lost their specificity/preference for H1N1pdm in ELISA. The specificity/preference of No. 29 was also confirmed by immunofluorescence assay and immunoprecipitation, and the viral nucleoprotein was identified by ELISA as its target protein. The change in specificity associated with scFv conversion from phage-bound to soluble form could be due to loss of phage scaffold pIII protein, which likely provides structural support for the scFv antigen-binding site. It is also possible that the similar antigenic properties of H1N1pdm and sH1N1 led to the observed alterations in scFv specificity. Discussion Using a phage display library, we obtained 7 soluble scFv clones reactive against H1N1pdm; however, only 1 showed specificity/preference toward H1N1pdm. Our results confirmed that using phage display libraries was highly

  18. Developmental patterns of free and protein-bound biotin during maturation and germination of seeds of Pisum sativum: characterization of a novel seed-specific biotinylated protein.

    PubMed

    Duval, M; Job, C; Alban, C; Douce, R; Job, D

    1994-04-01

    Mature dry pea seeds contain three major biotinylated proteins. Two of these of subunit molecular mass about 75 kDa and 200 kDa are associated with 3-methylcrotonyl-CoA carboxylase (EC 6.4.1.4) and acetyl-CoA carboxylase activities (EC 6.4.1.2) respectively. The third does not exhibit any of the biotin-dependent carboxylase activities found in higher organisms and represents the major part of the total protein-bound biotin in the seeds. This novel protein has been purified from a whole pea seed extract. Because in SDS/polyacrylamide gels the protein migrates with an apparent molecular mass of about 65 kDa, it is referred to as SBP65, for 65 kDa seed biotinylated protein. The molecular mass of native SBP65 is greater than 400 kDa, suggesting that the native protein assumes a polymeric structure, resulting from the association of six to eight identical subunits. The results of CNBr cleavage experiments suggest that biotin is covalently bound to the protein. The stoichiometry is 1 mol of biotin per 1 mol of 65 kDa polypeptide. The temporal and spatial pattern of expression of SBP65 is described. SBP65 is specifically expressed in the seeds, being absent from leaf, root, stem, pod and flower tissues of pea plants. The level of SBP65 increases dramatically during seed development. The protein is not detectable in very young seeds. Its accumulation pattern parallels that for storage proteins, being maximally expressed in the mature dry seeds. SBP65 disappears at a very high rate during seed germination. The level of free biotin has also been evaluated for various organs of pea plants. In all proliferating tissues examined (young developing seeds, leaf, root, stem, pod and flower tissues), free biotin is in excess of protein-bound biotin. Only in the mature dry seeds is protein-bound biotin (i.e. that bound to SBP65) in excess of free biotin. These temporal expression patterns, and the strict organ specificity for expression of SBP65, are discussed with regard to the

  19. RANKL, osteopontin, and osteoclast homeostasis in a hyperocclusion mouse model

    SciTech Connect

    Walker, Cameron G.; Ito, Yoshihiro; Dangaria, Smit; Luan, Xianghong; Diekwisch, Thomas G.H.

    2009-10-21

    The biological mechanisms that maintain the position of teeth in their sockets establish a dynamic equilibrium between bone resorption and apposition. In order to reveal some of the dynamics involved in the tissue responses towards occlusal forces on periodontal ligament (PDL) and alveolar bone homeostasis, we developed the first mouse model of hyperocclusion. Swiss-Webster mice were kept in hyperocclusion for 0, 3, 6, and 9 d. Morphological and histological changes in the periodontium were assessed using micro-computed tomography (micro-CT) and ground sections with fluorescent detection of vital dye labels. Sections were stained for tartrate-resistant acid phosphatase, and the expression of receptor activator of nuclear factor-{kappa}B ligand (RANKL) and osteopontin (OPN) was analyzed by immunohistochemistry and real-time polymerase chain reaction (PCR). Traumatic occlusion resulted in enamel surface abrasion, inhibition of alveolar bone apposition, significant formation of osteoclasts at 3, 6 and 9 d, and upregulation of OPN and RANKL. Data from this study suggest that both OPN and RANKL contribute to the stimulation of bone resorption in the hyperocclusive state. In addition, we propose that the inhibition of alveolar bone apposition by occlusal forces is an important mechanism for the control of occlusal height that might work in synergy with RANKL-induced bone resorption to maintain normal occlusion.

  20. Sympathetic control of bone mass regulated by osteopontin

    PubMed Central

    Nagao, Masashi; Feinstein, Timothy N.; Ezura, Yoichi; Hayata, Tadayoshi; Notomi, Takuya; Saita, Yoshitomo; Hanyu, Ryo; Hemmi, Hiroaki; Izu, Yayoi; Takeda, Shu; Wang, Kathryn; Rittling, Susan; Nakamoto, Tetsuya; Kaneko, Kazuo; Kurosawa, Hisashi; Karsenty, Gerard; Denhardt, David T.; Vilardaga, Jean-Pierre; Noda, Masaki

    2011-01-01

    The sympathetic nervous system suppresses bone mass by mechanisms that remain incompletely elucidated. Using cell-based and murine genetics approaches, we show that this activity of the sympathetic nervous system requires osteopontin (OPN), a cytokine and one of the major members of the noncollagenous extracellular matrix proteins of bone. In this work, we found that the stimulation of the sympathetic tone by isoproterenol increased the level of OPN expression in the plasma and bone and that mice lacking OPN (OPN-KO) suppressed the isoproterenol-induced bone loss by preventing reduced osteoblastic and enhanced osteoclastic activities. In addition, we found that OPN is necessary for changes in the expression of genes related to bone resorption and bone formation that are induced by activation of the sympathetic tone. At the cellular level, we showed that intracellular OPN modulated the capacity of the β2-adrenergic receptor to generate cAMP with a corresponding modulation of cAMP-response element binding (CREB) phosphorylation and associated transcriptional events inside the cell. Our results indicate that OPN plays a critical role in sympathetic tone regulation of bone mass and that this OPN regulation is taking place through modulation of the β2-adrenergic receptor/cAMP signaling system. PMID:21990347

  1. Osteopontin Deficiency Alters Biliary Homeostasis and Protects against Gallstone Formation

    PubMed Central

    Lin, Jing; Shao, Wei-qing; Chen, Zong-you; Zhu, Wen-wei; Lu, Lu; Cai, Duan; Qin, Lun-xiu; Jia, Hu-liang; Lu, Ming; Chen, Jin-hong

    2016-01-01

    The precipitation of excess biliary cholesterol as solid crystals is a prerequisite for cholesterol gallstone formation, which occurs due to disturbed biliary homeostasis. Biliary homeostasis is regulated by an elaborate network of genes in hepatocytes. If unmanaged, the cholesterol crystals will aggregate, fuse and form gallstones. We have previously observed that the levels of osteopontin (OPN) in bile and gallbladder were reduced in gallstone patients. However, the role and mechanism for hepatic OPN in cholesterol gallstone formation is undetermined. In this study, we found that the expression of hepatic OPN was increased in gallstone patients compared with gallstone-free counterparts. Then, we observed that OPN-deficient mice were less vulnerable to cholesterol gallstone formation than wild type mice. Further mechanistic studies revealed that this protective effect was associated with alterations of bile composition and was caused by the increased hepatic CYP7A1 expression and the reduced expression of hepatic SHP, ATP8B1, SR-B1 and SREBP-2. Finally, the correlations between the expression of hepatic OPN and the expression of these hepatic genes were validated in gallstone patients. Taken together, our findings reveal that hepatic OPN contributes to cholesterol gallstone formation by regulating biliary metabolism and might be developed as a therapeutic target for gallstone treatments. PMID:27484115

  2. Methods to Identify Chromatin-Bound Protein Complexes: From Genome-Wide to Locus-Specific Approaches.

    PubMed

    Massie, Charles E

    2016-01-01

    High-throughput sequencing approaches coupled with functional genomics experiments have facilitated a rapid growth in our understanding of chromatin biology, from genome-wide maps of transcription factor binding and histone modifications to insights into higher order chromatin organization under specific cellular conditions. However in most cases these methods require a prior knowledge of the system of interest (e.g., targets for immunoprecipitation or modulation) and therefore are limited in their utility to identify novel components of pathways or for the study of uncharacterized pathways. Several orthologous proteomics approaches have been developed recently that bridge this gap, allowing the identification of protein complexes globally or at specific genomic loci. In this chapter the relative advantages of each approach will be explored and a detailed protocol given for DNA pull-down of a specific androgen receptor (AR) genomic target. PMID:27246338

  3. Age dependent increase in the levels of osteopontin inhibits skeletal muscle regeneration.

    PubMed

    Paliwal, Preeti; Pishesha, Novalia; Wijaya, Denny; Conboy, Irina M

    2012-08-01

    Skeletal muscle regeneration following injury is accompanied by rapid infiltration of macrophages, which play a positive role in muscle repair. Increased chronic inflammation inhibits the regeneration of dystrophic muscle, but the properties of inflammatory cells are not well understood in the context of normal muscle aging. This work uncovers pronounced age-specific changes in the expression of osteopontin (OPN) in CD11b+ macrophages present in the injured old muscle as well as in the blood serum of old injured mice and in the basement membrane surrounding old injured muscle fibers. Furthermore, young CD11b+ macrophages enhance regenerative capacity of old muscle stem cells even when old myofibers and old sera are present; and neutralization of OPN similarly rejuvenates the myogenic responses of old satellite cells in vitro and notably, in vivo. This study highlights potential mechanisms by which age related inflammatory responses become counter-productive for muscle regeneration and suggests new strategies for enhancing muscle repair in the old. PMID:22915705

  4. Down-regulation of osteopontin attenuates breast tumour progression in vivo.

    PubMed

    Chakraborty, Goutam; Jain, Shalini; Patil, Tushar V; Kundu, Gopal C

    2008-12-01

    Development of breast tumour malignancies results in enhanced expression of various oncogenic molecules. Elevated expression of osteopontin (OPN) in higher grades of breast carcinoma correlates with enhanced expressions of several oncogenic molecules (urokinase-type plasminogen activator [uPA], matrix metalloproteinase-2/-9 [MMP-2 and -9]) and increased angiogenic potential of breast carcinoma. In this study, using in vitro and multiple in vivo models, we have demonstrated that silencing of OPN by its specific small interfering RNA (siRNA) down-regulates the expressions of oncogenic molecules such as uPA, MMP-2 and -9 resulting in inhibition of in vitro cell motility and in vivo tumourigenicity in mice. Moreover our results demonstrated that OPN-/- mice showed slower progression of tumour growth in breast cancer model as compared to wild-type mice. Furthermore, the data showed that injection of carcinogenic compound, pristane (2, 6,10,14-tetramethylpen-tadecane) induces breast tumour progression leading to enhanced expression of OPN and other oncogenic molecules in mammary fat pad of nude- and wild-type mice but not in OPN-/- mice. However, intratumoural injection of OPN siRNA to pristane-induced tumour significantly suppressed these effects. Our data revealed that knocking down of OPN effectively curb breast cancer progression and further suggested that developing of OPN-based therapeutics might be an emerging approach for the next generation of breast cancer management. PMID:18266970

  5. Physical Uncertainty Bounds (PUB)

    SciTech Connect

    Vaughan, Diane Elizabeth; Preston, Dean L.

    2015-03-19

    This paper introduces and motivates the need for a new methodology for determining upper bounds on the uncertainties in simulations of engineered systems due to limited fidelity in the composite continuum-level physics models needed to simulate the systems. We show that traditional uncertainty quantification methods provide, at best, a lower bound on this uncertainty. We propose to obtain bounds on the simulation uncertainties by first determining bounds on the physical quantities or processes relevant to system performance. By bounding these physics processes, as opposed to carrying out statistical analyses of the parameter sets of specific physics models or simply switching out the available physics models, one can obtain upper bounds on the uncertainties in simulated quantities of interest.

  6. Analysis of endogenous peptides bound by soluble MHC class I molecules: a novel approach for identifying tumor-specific antigens.

    PubMed

    Barnea, Eilon; Beer, Ilan; Patoka, Renana; Ziv, Tamar; Kessler, Ofra; Tzehoval, Esther; Eisenbach, Lea; Zavazava, Nicholas; Admon, Arie

    2002-01-01

    The Human MHC Project aims at comprehensive cataloging of peptides presented within the context of different human leukocyte antigens (HLA) expressed by cells of various tissue origins, both in health and in disease. Of major interest are peptides presented on cancer cells, which include peptides derived from tumor antigens that are of interest for immunotherapy. Here, HLA-restricted tumor-specific antigens were identified by transfecting human breast, ovarian and prostate tumor cell lines with truncated genes of HLA-A2 and HLA-B7. Soluble HLA secreted by these cell lines were purified by affinity chromatography and analyzed by nano-capillary electrospray ionization-tandem mass spectrometry. Typically, a large peptide pool was recovered and sequenced including peptides derived from MAGE-B2 and mucin and other new tumor-derived antigens that may serve as potential candidates for immunotherapy. PMID:11782012

  7. Importance of Phosphorylation for Osteopontin Regulation of Biomineralization

    PubMed Central

    Gericke, A.; Qin, C.; Spevak, L.; Fujimoto, Y.; Butler, W. T.; Sørensen, E. S.; Boskey, A. L.

    2006-01-01

    Previous in vitro and in vivo studies demonstrated that osteopontin (OPN) is an inhibitor of the formation and growth of hydroxyapatite (HA) and other biominerals. The present study tests the hypotheses that the interaction of OPN with HA is determined by the extent of protein phosphorylation and that this interaction regulates the mineralization process. Bone OPN as previously reported inhibited HA formation and HA-seeded growth in a gelatin-gel system. A transglutaminase-linked OPN polymer had similar effects. Recombinant, nonphosphorylated OPN and chemically dephosphorylated OPN, had no effect on HA formation or growth in this system. In contrast, highly phosphorylated milk OPN (mOPN) promoted HA formation. The mOPN stabilized the conversion of amorphous calcium phosphate (a noncrystalline constituent of milk) to HA, whereas bone OPN had a lesser effect on this conversion. Mixtures of OPN and osteocalcin known to form a complex in vitro, unexpectedly promoted HA formation. To test the hypothesis that small alterations in protein conformation caused by phosphorylation account for the differences in the observed ability of OPN to interact with HA, the conformation of bone OPN and mOPN in the presence and absence of crystalline HA was determined by attenuated total reflection (ATR) infrared (IR) spectroscopy. Both proteins exhibited a predominantly random coil structure, which was unaffected by the addition of Ca2+. Binding to HA did not alter the secondary structure of bone OPN, but induced a small increase of β-sheet (few percent) in mOPN. These data taken together suggest that the phosphorylation of OPN is an important factor in regulating the OPN-mediated mineralization process. PMID:16007483

  8. Osteopontin Facilitates Ultraviolet B-induced Squamous Cell Carcinoma Development

    PubMed Central

    Chang, Pi-Ling; Hsieh, Yu-Hua; Wang, Chao-Cheng; Juliana, M. Margaret; Tsuruta, Yuko; Timares, Laura; Elmets, Craig; Ho, Kang-Jey

    2014-01-01

    Background Osteopontin (OPN) is a matricellular glycoprotein that is markedly expressed in cutaneous squamous cell carcinomas (cSCCs) and in actinic keratoses implicating its role in photocarcinogenesis. Objective To determine whether OPN facilitates the development of cSCC and its function. Methods cSCCs development was compared between wild-type (WT) and OPN-null mice subjected to UVB irradiation for 43 weeks. UVB-induced OPN expression was determined by Western blot, immunoprecipitation, ELISA, and semi-quantitative RT-PCR. Epidermal layer and TUNEL analyses assessed if OPN mediates UVB-induced epidermal hyperplasia or suppresses UVB-induced apoptosis of basal keratinocytes, respectively. In vitro experiments determined whether OPN enhances cell survival of UVB-induced apoptosis and its potential mechanisms. Immunohistochemical analyses of epidermis assessed the expression of CD44 and focal adhesion kinase (FAK), molecules that mediate OPN survival function. Results Compared to female WT mice, OPN-null mice did not develop cSCCs. UVB irradiation stimulated OPN protein expression in the dorsal skin by 11 h and remains high at 24 to 48h.OPN did not mediate UVB-induced epidermal hyperplasia; instead, it protected basal keratinocytes from undergoing apoptosis upon UVB exposure. Likewise, the addition of OPN suppressed UVB-induced OPN-null cSCC cell apoptosis, the activation of caspase-9 activity, and increased phosphorylation of FAK at Y397. Furthermore, the expression of CD44 and FAK in WT mice epidermis was greater than that of OPN-null mice prior to and during early acute UVB exposure. Conclusion These data support the hypothesis that chronic UVB-induced OPN expression protects the survival of initiated basal keratinocytes and, consequently, facilitates cSCC develop. PMID:24888687

  9. Role of Osteopontin in Psoriasis: An Immunohistochemical Study

    PubMed Central

    Abdel-Mawla, M Yousry; El-Kasheshy, Kamal Ahmed; Ghonemy, Soheir; Al Balat, Walid; Elsayed, Amira Ahmed

    2016-01-01

    Background: Osteopontin (OPN) has been postulated to have a role in several T-helper (Th) 1 and Th 17-mediated diseases including psoriasis (PS), through multiple mechanisms sharing in the onset and worsening of PS, OPN shares in induction of keratinocyte proliferation through inhibiting keratinocyte apoptosis, OPN acts as a proinflammatory agent that participates in the upregulation of Th cell lineages, among which are the Th 1 and Th 17 cells. Aims and Objectives: The aim of this study was to explore the possible role of OPN in the pathogenesis of PS. Materials and Methods: This case–control study was carried out on 18 patients of chronic plaque PS (mean age 37.61 ± 14.48) and a control group of 18 apparently healthy volunteers (mean age 41.11 ± 11.02 years). Severity of PS was assessed using the PS area and severity index score. Two skin biopsies were taken from psoriatic patients. The first was taken from the lesional skin and the other from a counter apparently healthy site. Results: Our results showed statistically significant differences in the expression of OPN, between lesional and nonlesional skin as well as between nonlesional skin and control group (P ≤ 0.001). In addition, there was a significant difference in the expression of OPN, between control and lesional group. Conclusions: OPN involvement in PS enlarges the list of cytokines able to stimulate the inflammatory response in this disease, anti-OPN antibodies, may eventually become a useful therapeutic approach in PS. PMID:27293251

  10. Recombinant Osteopontin in Cerebral Vasospasm After Subarachnoid Hemorrhage

    PubMed Central

    Suzuki, Hidenori; Hasegawa, Yu; Chen, Wanqiu; Kanamaru, Kenji; Zhang, John H.

    2010-01-01

    Objective Osteopontin (OPN), a pleiotropic extracellular matrix glycoprotein, has been reported to be protective against ischemic lesions, but effects of OPN on vascular functions have not been investigated. The aim of this study was to assess whether recombinant OPN (r-OPN) could prevent cerebral vasospasm after subarachnoid hemorrhage (SAH) in rats. Methods r-OPN was administered intraventricularly to rats undergoing SAH by the endovascular perforation, and its protective effects were evaluated by measuring the diameter of cerebral arteries and neurobehavioral testing. Western blotting and immunofluorescence were performed to explore the underlying mechanisms. An integrin receptor antagonist GRGDSP or mitogen-activated protein kinase (MAPK) phosphatase (MKP)-1 small interfering RNA (siRNA) was also administered to r-OPN-treated SAH rats, and those effects were evaluated. Results Pre-SAH administration of r-OPN prevented vasospasm and neurological impairments at 24–72 hours post-SAH. r-OPN enhanced an endogenous MAPK inhibitor, MKP-1, and suppressed the phosphorylation of MAPKs, caldesmon and heat shock protein 27 in the spastic cerebral arteries at 24 hours post-SAH. Immunofluorescence revealed that MKP-1 was induced in the arterial smooth muscle layer. GRGDSP prevented r-OPN-induced MKP-1 upregulation, and MKP-1 siRNA abolished both MAPK inactivation and anti-vasospastic effects by r-OPN. Post-SAH r-OPN treatment also prevented vasospasm. Interpretation r-OPN induced MKP-1 in the spastic cerebral arteries via binding to L-arginyl-glycyl-L-aspartate-dependent integrin receptors and prevented vasospasm after SAH. Therapeutic induction of MKP-1 may be a novel approach for the prevention and treatment of cerebral vasospasm. PMID:21031580

  11. Expression of osteopontin and CD44 molecule in papillary renal cell tumors.

    PubMed

    Matusan, Koviljka; Dordevic, Gordana; Mozetic, Vladimir; Lucin, Ksenija

    2005-01-01

    The aim of the study was to analyze the expression of CD44 adhesion molecule and its ligand osteopontin in papillary renal cell tumors, and to assess the possible prognostic significance of CD44 and osteopontin expression in papillary renal cell carcinomas. The expression of the standard and v6 exon containing isoforms of CD44 molecule, as well as of its ligand osteopontin, was immunohistochemically evaluated in 43 papillary renal cell tumors, which included 5 adenomas and 38 carcinomas. In order to assess their prognostic significance, the results obtained in papillary renal cell carcinomas were compared to usual clinicopathological parameters such as tumor size, histological grade, pathological stage, and Ki-67 proliferation index. Normal renal tissue was negative for CD44s and v6 isoforms, while the expression of osteopontin was found in distal tubular epithelial cells in the form of cytoplasmic granular positivity. CD44s and v6 isoforms were upregulated in 22 (58%) and 12 (32%) out of 38 carcinomas, respectively. Among all clinicopathological parameters examined, we only found significant association of CD44s-positive carcinomas with lower pathological stage (p=0.026). Papillary renal cell adenomas were generally negative for CD44s, except for focal positivity found in one sample. The osteopontin protein was detected in all adenomas and all papillary renal cell carcinomas, except one. Our results show constitutive expression of osteopontin in papillary renal tumors, including papillary renal cell adenomas. The upregulation of CD44s and v6 isoforms, although found in a considerable number of papillary renal cell carcinomas, does not appear to have any prognostic value in this type of renal cancer. PMID:15999156

  12. Preparation, characterization, and immunogenicity of conjugates composed of the O-specific polysaccharide of Shigella dysenteriae type 1 (Shiga's bacillus) bound to tetanus toxoid.

    PubMed Central

    Chu, C Y; Liu, B K; Watson, D; Szu, S S; Bryla, D; Shiloach, J; Schneerson, R; Robbins, J B

    1991-01-01

    The background for developing conjugate vaccines for shigellosis composed of the O-specific polysaccharide (O-SP) bound to a protein is described elsewhere (C. Y. Chu, R. Schneerson, and J. B. Robbins, submitted for publication). Briefly, there is direct evidence for type (lipopolysaccharide [LPS])-specific protection after infection with the wild type or with attenuated strains of shigellae. Prospective studies of Israeli armed forces recruits show a correlation between preexisting serum immunoglobulin G (IgG) LPS antibodies and resistance to shigellosis (D. Cohen, M. S. Green, C. Block, R. Slephon, and I. Ofek, J. Clin. Microbiol. 29:386-389, 1991). In order to elicit IgG LPS-specific antibodies to Shigella dysenteriae type 1, the O-SP of this pathogen was purified and bound to tetanus toxoid (TT) by three schemes. The most immunogenic used a modification of a published method (C. Y. Chu, R. Schneerson, J. B. Robbins, and S. C. Rastogi, Infect. Immun. 40:245-256, 1983). The resultant O-SP-TT conjugates were stable and elicited high levels of IgG O-SP antibodies and booster responses in young mice when injected subcutaneously in saline at 1/10 the proposed human dose. Adsorption onto alum or concurrent administration with monophosphoryl lipid A enhanced both the IgG and IgM antibody responses to the O-SP of the conjugate; both the nonadsorbed and adsorbed conjugates elicited higher rises of IgG than of IgM antibodies. Clinical evaluations of S. dysenteriae type 1 O-SP-TT conjugates are planned. Images PMID:1937803

  13. Structure of xylose reductase bound to NAD+ and the basis for single and dual co-substrate specificity in family 2 aldo-keto reductases.

    PubMed Central

    Kavanagh, Kathryn L; Klimacek, Mario; Nidetzky, Bernd; Wilson, David K

    2003-01-01

    The co-ordinates reported have been submitted to the Protein Data Bank under accession number 1MI3. Xylose reductase (XR; AKR2B5) is an unusual member of aldo-keto reductase superfamily, because it is one of the few able to efficiently utilize both NADPH and NADH as co-substrates in converting xylose into xylitol. In order to better understand the basis for this dual specificity, we have determined the crystal structure of XR from the yeast Candida tenuis in complex with NAD(+) to 1.80 A resolution (where 1 A=0.1 nm) with a crystallographic R -factor of 18.3%. A comparison of the NAD(+)- and the previously determined NADP(+)-bound forms of XR reveals that XR has the ability to change the conformation of two loops. To accommodate both the presence and absence of the 2'-phosphate, the enzyme is able to adopt different conformations for several different side chains on these loops, including Asn(276), which makes alternative hydrogen-bonding interactions with the adenosine ribose. Also critical is the presence of Glu(227) on a short rigid helix, which makes hydrogen bonds to both the 2'- and 3'-hydroxy groups of the adenosine ribose. In addition to changes in hydrogen-bonding of the adenosine, the ribose unmistakably adopts a 3'- endo conformation rather than the 2'- endo conformation seen in the NADP(+)-bound form. These results underscore the importance of tight adenosine binding for efficient use of either NADH or NADPH as a co-substrate in aldo-keto reductases. The dual specificity found in XR is also an important consideration in designing a high-flux xylose metabolic pathway, which may be improved with an enzyme specific for NADH. PMID:12733986

  14. Sliding of Proteins Non-specifically Bound to DNA: Brownian Dynamics Studies with Coarse-Grained Protein and DNA Models

    PubMed Central

    Ando, Tadashi; Skolnick, Jeffrey

    2014-01-01

    DNA binding proteins efficiently search for their cognitive sites on long genomic DNA by combining 3D diffusion and 1D diffusion (sliding) along the DNA. Recent experimental results and theoretical analyses revealed that the proteins show a rotation-coupled sliding along DNA helical pitch. Here, we performed Brownian dynamics simulations using newly developed coarse-grained protein and DNA models for evaluating how hydrodynamic interactions between the protein and DNA molecules, binding affinity of the protein to DNA, and DNA fluctuations affect the one dimensional diffusion of the protein on the DNA. Our results indicate that intermolecular hydrodynamic interactions reduce 1D diffusivity by 30%. On the other hand, structural fluctuations of DNA give rise to steric collisions between the CG-proteins and DNA, resulting in faster 1D sliding of the protein. Proteins with low binding affinities consistent with experimental estimates of non-specific DNA binding show hopping along the CG-DNA. This hopping significantly increases sliding speed. These simulation studies provide additional insights into the mechanism of how DNA binding proteins find their target sites on the genome. PMID:25504215

  15. Max-flow segmentation of the left ventricle by recovering subject-specific distributions via a bound of the Bhattacharyya measure.

    PubMed

    Ben Ayed, Ismail; Chen, Hua-Mei; Punithakumar, Kumaradevan; Ross, Ian; Li, Shuo

    2012-01-01

    This study investigates fast detection of the left ventricle (LV) endo- and epicardium boundaries in a cardiac magnetic resonance (MR) sequence following the optimization of two original discrete cost functions, each containing global intensity and geometry constraints based on the Bhattacharyya similarity. The cost functions and the corresponding max-flow optimization built upon an original bound of the Bhattacharyya measure yield competitive results in nearly real-time. Within each frame, the algorithm seeks the LV cavity and myocardium regions consistent with subject-specific model distributions learned from the first frame in the sequence. Based on global rather than pixel-wise information, the proposed formulation relaxes the need of a large training set and optimization with respect to geometric transformations. Different from related active contour methods, it does not require a large number of iterative updates of the segmentation and the corresponding computationally onerous kernel density estimates (KDEs). The algorithm requires very few iterations and KDEs to converge. Furthermore, the proposed bound can be used for several other applications and, therefore, can lead to segmentation algorithms which share the flexibility of active contours and computational advantages of max-flow optimization. Quantitative evaluations over 2280 images acquired from 20 subjects demonstrated that the results correlate well with independent manual segmentations by an expert. Moreover, comparisons with a related recent active contour method showed that the proposed framework brings significant improvements in regard to accuracy and computational efficiency. PMID:21705264

  16. Osteopontin Fragments with Intact Thrombin-Sensitive Site Circulate in Cervical Cancer Patients

    PubMed Central

    Lim, Pak-Leong; Cheung, Tak-Hong; Wong, Raymond R. Y.; Yim, So-Fan; Ng, Margaret H. L.; Tam, Frankie C. H.; Chung, Tony K. H.; Wong, Yick-Fu

    2016-01-01

    We investigated whether circulating osteopontin (OPN) could be used as a biomarker for cervical cancer. We employed a monoclonal antibody (mAb 659) specific for the unique and intact thrombin-sensitive site in OPN using an inhibition ELISA. We found significantly higher levels of OPN in 33 cervical cancer patients in both the plasma (mean +/- SD, 612 +/- 106 ng/mL) and serum (424 +/- 121 ng/mL) compared to healthy subjects [409 +/- 56 ng/mL, from 31 plasma samples (P < 0.0001), and 314 +/- 98 ng/mL, from 32 serum samples (P = 0.0002), respectively]. Similar results were obtained when the plasma from a bigger group (147 individuals) of cervical cancer patients (560 +/- 211 ng/mL) were compared with the same plasma samples of the healthy individuals (P = 0.0014). More significantly, the OPN level was highest in stage III-IV disease (614 +/- 210 ng/mL, from 52 individuals; P = 0.0001) and least and non-discriminatory in stage I (473 +/- 110 ng/mL, from 40 individuals; P = 0.5318). No such discrimination was found when a mAb of a different specificity (mAb 446) was used in a similar inhibition ELISA to compare the two groups in the first study; a commercial capture ELISA also failed. The possibility that the target epitope recognized by the antibody probe in these assays was absent from the circulating OPN due to protein truncation was supported by gel fractionation of the OPN found in patients’ plasma: 60–64 kDa fragments were found instead of the presumably full-length OPN (68 kDa) seen in healthy people. How these fragments are generated and what possible role they play in cancer biology remain interesting questions. PMID:27494141

  17. The distal enhancer implicated in the developmental regulation of the tyrosine aminotransferase gene is bound by liver-specific and ubiquitous factors.

    PubMed Central

    Nitsch, D; Schütz, G

    1993-01-01

    Tyrosine aminotransferase gene expression is confined to parenchymal cells of the liver, is inducible by glucocorticoids and glucagon, and is repressed by insulin. Three enhancers control this tissue-specific and hormone-dependent activity, one of which, located at -11 kb, is implicated in establishing an active expression domain. We have studied in detail this important regulatory element and have identified a 221-bp fragment containing critical enhancer sequences which stimulated the heterologous thymidine kinase promoter more than 100-fold in hepatoma cells. Within this region, we have characterized two essential liver-specific enhancer domains, one of which was bound by proteins of the hepatocyte nuclear factor 3 (HNF3) family. Analyses with the dedifferentiated hepatoma cell line HTC suggested that HNF3 alpha and/or -gamma, but not HNF3 beta, are involved in activating the tyrosine aminotransferase gene via the -11-kb enhancer. Genomic footprinting and in vitro protein-DNA binding studies documented cell-type-specific binding of ubiquitous factors to the second essential enhancer domain, which by itself stimulated the thymidine kinase promoter preferentially in hepatoma cells. These results will allow further characterization of the role of these enhancer sequences in developmental activation of the tyrosine aminotransferase gene. Images PMID:8101632

  18. Role of osteopontin in hepatic neutrophil infiltration during alcoholic steatohepatitis

    SciTech Connect

    Apte, Udayan M.; Banerjee, Atrayee; McRee, Rachel; Wellberg, Elizabeth; Ramaiah, Shashi K. . E-mail: sramaiah@cvm.tamu.edu

    2005-08-22

    Alcoholic liver disease (ALD) is a major complication of heavy alcohol (EtOH) drinking and is characterized by three progressive stages of pathology: steatosis, steatohepatitis, and fibrosis/cirrhosis. Alcoholic steatosis (AS) is the initial stage of ALD and consists of fat accumulation in the liver accompanied by minimal liver injury. AS is known to render the hepatocytes increasingly sensitive to toxicants such as bacterial endotoxin (LPS). Alcoholic steatohepatitis (ASH), the second and rate-limiting step in the progression of ALD, is characterized by hepatic fat accumulation, neutrophil infiltration, and neutrophil-mediated parenchymal injury. However, the pathogenesis of ASH is poorly defined. It has been theorized that the pathogenesis of ASH involves interaction of increased circulating levels of LPS with hepatocytes being rendered highly sensitive to LPS due to heavy EtOH consumption. We hypothesize that osteopontin (OPN), a matricellular protein (MCP), plays an important role in the hepatic neutrophil recruitment due to its enhanced expression during the early phase of ALD (AS and ASH). To study the role of OPN in the pathogenesis of ASH, we induced AS in male Sprague-Dawley rats by feeding EtOH-containing Lieber-DeCarli liquid diet for 6 weeks. AS rats experienced extensive fat accumulation and minimal liver injury. Moderate induction in OPN was observed in AS group. ASH was induced by feeding male Sprague-Dawley rats EtOH-containing Lieber-DeCarli liquid diet for 6 weeks followed by LPS injection. The ASH rats had substantial neutrophil infiltration, coagulative oncotic necrosis, and developed higher liver injury. Significant increases in the hepatic and circulating levels of OPN was observed in the ASH rats. Higher levels of the active, thrombin-cleaved form of OPN in the liver in ASH group correlated remarkably with hepatic neutrophil infiltration. Finally, correlative studies between OPN and hepatic neutrophil infiltration was corroborated in a simple

  19. [Determination of osteopontin at trace levels by non-gel sieving capillary electrophoresis].

    PubMed

    Zhao, Jingshan; Wen, Jinkun; Han, Mei

    2005-09-01

    A method of non-gel sieving capillary electrophoresis (NGSCE) was established to determine osteopontin at trace levels. The capillary used was uncoated fused silica with a size of 57 cm x 75 microm i. d. and an effective length of 50 cm. The electrode buffer was a 150 mmol/L boric acid-borate buffer containing 30 g/L polyethylene glycol 20000 (pH 10.0). Other conditions were as follows: separation voltage 23 kV; detection wavelength 214 nm; pressure of injecting sample 3.4 kPa (0.5 psi) x 5 s; and column temperature 25 degrees C. The NGSCE method had excellent linearity with correlation coefficient of 0. 996, and reproducibility with the relative standard deviation of migration time of osteopontin less than 5% . The recovery was 95% and better, the sensitivity was 0. 079 g/L. Osteopontin secreted by vascular smooth muscle cells was determined by the NGSCE method at different times after serum withdrawal, and the results were in agreement with those of Western blot method. The results indicate that NGSCE is a simple and rapid method of determining osteopontin at trace levels. This method only needs a micro-amount of sample and is easily automated. PMID:16350797

  20. Dynamic light scattering study of inhibition of nucleation and growth of hydroxyapatite crystals by osteopontin.

    PubMed

    de Bruyn, John R; Goiko, Maria; Mozaffari, Maryam; Bator, Daniel; Dauphinee, Ron L; Liao, Yinyin; Flemming, Roberta L; Bramble, Michael S; Hunter, Graeme K; Goldberg, Harvey A

    2013-01-01

    We study the effect of isoforms of osteopontin (OPN) on the nucleation and growth of crystals from a supersaturated solution of calcium and phosphate ions. Dynamic light scattering is used to monitor the size of the precipitating particles and to provide information about their concentration. At the ion concentrations studied, immediate precipitation was observed in control experiments with no osteopontin in the solution, and the size of the precipitating particles increased steadily with time. The precipitate was identified as hydroxyapatite by X-ray diffraction. Addition of native osteopontin (nOPN) extracted from rat bone caused a delay in the onset of precipitation and reduced the number of particles that formed, but the few particles that did form grew to a larger size than in the absence of the protein. Recombinant osteopontin (rOPN), which lacks phosphorylation, caused no delay in initial calcium phosphate precipitation but severely slowed crystal growth, suggesting that rOPN inhibits growth but not nucleation. rOPN treated with protein kinase CK2 to phosphorylate the molecule (p-rOPN) produced an effect similar to that of nOPN, but at higher protein concentrations and to a lesser extent. These results suggest that phosphorylations are critical to OPN's ability to inhibit nucleation, whereas the growth of the hydroxyapatite crystals is effectively controlled by the highly acidic OPN polypeptide. This work also demonstrates that dynamic light scattering can be a powerful tool for delineating the mechanism of protein modulation of mineral formation. PMID:23457612

  1. Osteopontin That Is Elevated in the Airways during COPD Impairs the Antibacterial Activity of Common Innate Antibiotics

    PubMed Central

    Mori, Michiko; Keenan, Paul; Mörgelin, Matthias; Erjefält, Jonas S.; Herwald, Heiko; Egesten, Arne; Kasetty, Gopinath

    2016-01-01

    Bacterial infections of the respiratory tract contribute to exacerbations and disease progression in chronic obstructive pulmonary disease (COPD). There is also an increased risk of invasive pneumococcal disease in COPD. The underlying mechanisms are not fully understood but include impaired mucociliary clearance and structural remodeling of the airways. In addition, antimicrobial proteins that are constitutively expressed or induced during inflammatory conditions are an important part of the airway innate host defense. In the present study, we show that osteopontin (OPN), a multifunctional glycoprotein that is highly upregulated in the airways of COPD patients co-localizes with several antimicrobial proteins expressed in the airways. In vitro, OPN bound lactoferrin, secretory leukocyte peptidase inhibitor (SLPI), midkine, human beta defensin-3 (hBD-3), and thymic stromal lymphopoietin (TSLP) but showed low or no affinity for lysozyme and LL-37. Binding of OPN impaired the antibacterial activity against the important bacterial pathogens Streptococcus pneumoniae and Pseudomonas aeruginosa. Interestingly, OPN reduced lysozyme-induced killing of S. pneumoniae, a finding that could be explained by binding of OPN to the bacterial surface, thereby shielding the bacteria. A fragment of OPN generated by elastase of P. aeruginosa retained some inhibitory effect. Some antimicrobial proteins have additional functions. However, the muramidase-activity of lysozyme and the protease inhibitory function of SLPI were not affected by OPN. Taken together, OPN can contribute to the impairment of innate host defense by interfering with the function of antimicrobial proteins, thus increasing the vulnerability to acquire infections during COPD. PMID:26731746

  2. A CD153+CD4+ T follicular cell population with cell-senescence features plays a crucial role in lupus pathogenesis via osteopontin production.

    PubMed

    Tahir, Suhail; Fukushima, Yuji; Sakamoto, Keiko; Sato, Kyosuke; Fujita, Harumi; Inoue, Joe; Uede, Toshimitsu; Hamazaki, Yoko; Hattori, Masakazu; Minato, Nagahiro

    2015-06-15

    Immune aging results in diminished adaptive immunity and increased risk for autoimmunity. We previously reported a unique PD-1(+) CD44(high)CD4(+) T cell population that increases with age in normal mice. In this study, we indicate that the age-dependent PD-1(+) CD44(high)CD4(+) T cells develop as unique T follicular (TF) cells in a B cell-dependent manner and consist of two subpopulations, as follows: CD153(+) cells preferentially secreting abundant osteopontin on TCR stimulation and CD153(-) cells that are apparently TCR anergic. These unique TF cells with essentially similar features increase much earlier and are accumulated in the spontaneous germinal centers (GCs) in lupus-prone female BWF1 (f-BWF1) mice. These TF cells showed characteristic cell-senescence features and developed in association with extensive CD4(+) T cell proliferation in vivo, suggesting replicative senescence. Although the CD153(+) TF cells were defective in proliferation capacity, they were quite stable and specifically responded to self GC-B cells to secret abundant osteopontin, which inhibited B cell receptor-induced GC-B cell apoptosis in f-BWF1 mice. Transfer of CD153(+) PD-1(+) CD4(+) T cells promoted the growth of spontaneous GCs, whereas administration of anti-osteopontin Ab suppressed GC enlargement and anti-nuclear Ab production and ameliorated clinical lupus nephritis of f-BWF1 mice. Current results suggest that senescent CD153(+) TF cells generated as a consequence of extensive endogenous CD4(+) T cell proliferation play an essential, if not sufficient, role in lupus pathogenesis in lupus-prone genetic background and may also contribute to an increased autoimmunity risk with age. PMID:25972477

  3. Actinide(IV) Deposits on Bone: Potential Role of the Osteopontin-Thorium Complex.

    PubMed

    Creff, Gaëlle; Safi, Samir; Roques, Jérôme; Michel, Hervé; Jeanson, Aurélie; Solari, Pier-Lorenzo; Basset, Christian; Simoni, Eric; Vidaud, Claude; Den Auwer, Christophe

    2016-01-01

    In case of a nuclear event, contamination (broad or limited) of the population or of specific workers might occur. In such a senario, the fate of actinide contaminants may be of first concern, in particular with regard to human target organs like the skeleton. To improve our understanding of the toxicological processes that might take place, a mechanistic approach is necessary. For instance, ∼50% of Pu(IV) is known from biokinetic data to accumulate in bone, but the underlining mechanisms are almost unknown. In this context, and to obtain a better description of the toxicological mechanisms associated with actinides(IV), we have undertaken the investigation, on a molecular scale, of the interaction of thorium(IV) with osteopontin (OPN) a hyperphosphorylated protein involved in bone turnover. Thorium is taken here as a simple model for actinide(IV) chemistry. In addition, we have selected a phosphorylated hexapeptide (His-pSer-Asp-Glu-pSer-Asp-Glu-Val) that is representative of the peptidic sequence involved in the bone interaction. For both the protein and the biomimetic peptide, we have determined the local environment of Th(IV) within the bioactinidic complex, combining isothermal titration calorimetry, attenuated total reflectance Fourier transform infrared spectroscopy, theoretical calculations with density functional theory, and extended X-ray absorption fine structure spectroscopy at the Th LIII edge. The results demonstrate a predominance of interaction of metal with the phosphate groups and confirmed the previous physiological studies that have highlighted a high affinity of Th(IV) for the bone matrix. Data are further compared with those of the uranyl case, representing the actinyl(V) and actinyl(VI) species. Last, our approach shows the importance of developing simplified systems [Th(IV)-peptide] that can serve as models for more biologically relevant systems. PMID:26684435

  4. Identification of amino acid residues that determine the substrate specificity of mammalian membrane-bound front-end fatty acid desaturases.

    PubMed

    Watanabe, Kenshi; Ohno, Makoto; Taguchi, Masahiro; Kawamoto, Seiji; Ono, Kazuhisa; Aki, Tsunehiro

    2016-01-01

    Membrane-bound desaturases are physiologically and industrially important enzymes that are involved in the production of diverse fatty acids such as polyunsaturated fatty acids and their derivatives. Here, we identified amino acid residues that determine the substrate specificity of rat Δ6 desaturase (D6d) acting on linoleoyl-CoA by comparing its amino acid sequence with that of Δ5 desaturase (D5d), which converts dihomo-γ-linolenoyl-CoA. The N-terminal cytochrome b5-like domain was excluded as a determinant by domain swapping analysis. Substitution of eight amino acid residues (Ser209, Asn211, Arg216, Ser235, Leu236, Trp244, Gln245, and Val344) of D6d with the corresponding residues of D5d by site-directed mutagenesis switched the substrate specificity from linoleoyl-CoA to dihomo-γ-linolenoyl-CoA. In addition, replacement of Leu323 of D6d with Phe323 on the basis of the amino acid sequence of zebra fish Δ5/6 bifunctional desaturase was found to render D6d bifunctional. Homology modeling of D6d using recent crystal structure data of human stearoyl-CoA (Δ9) desaturase revealed that Arg216, Trp244, Gln245, and Leu323 are located near the substrate-binding pocket. To our knowledge, this is the first report on the structural basis of the substrate specificity of a mammalian front-end fatty acid desaturase, which will aid in efficient production of value-added fatty acids. PMID:26590171

  5. Osteopontin: A Novel Cytokine Involved in the Regulation of Mycobacterium avium subsp. paratuberculosis Infection in Periparturient Dairy Cattle

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Osteopontin (Opn), an important mediator of the cell-mediated immune response, enhances the host immune response against mycobacterial infections. Infections caused by the intracellular bacterium, Mycobacterium avium subsp. paratuberculosis (MAP), have a devastating impact on the dairy industry. ...

  6. Osteopontin Immunoreactivity in the Ileum and Ileoceccal Lymph Node of Dairy Cows Naturally Infected with Mycobacterium avium subsp. paratuberculosis

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Osteopontin (Opn), a highly acidic glycoprotein, promotes cellular adhesion and recruitment and has been shown to be upregulated in the granulomas of mycobacterial infections. Johne’s disease, caused by Mycobacterium avium subsp. paratuberculosis (MAP), is associated with granulomatous enteritis. ...

  7. Anti-osteopontin monoclonal antibody prevents ovariectomy-induced osteoporosis in mice by promotion of osteoclast apoptosis

    SciTech Connect

    Zhang, Bo; Dai, Jianxin; Wang, Huaqing; Wei, Huafeng; Zhao, Jian; Guo, Yajun; and others

    2014-09-26

    Highlight: • We first report that anti-osteopontin mAb could protect osteoporosis in mice. • Anti-osteopontin mAb could promote the osteoclast apoptosis. • Targeting osteopontin might have therapeutic potentials for osteoporosis. - Abstract: Osteopontin (OPN) is abundant in mineralized tissues and has long been implicated in bone remodeling. However, the therapeutic effect of targeting OPN in bone loss diseases and the underlying molecular mechanism remain largely unknown. Here, we reported that anti-OPN mAb (23C3) could protect against ovariectomy-induced osteoporosis in mice, demonstrated by microcomputed tomography analysis and histopathology evaluation. In vitro assay showed that 23C3 mAb reduced osteoclasts (OCs)-mediated bone resorption through promotion of mature OC apoptosis. Thus, the study has important implications for understanding the role of OPN in OC bone resorption and survival, and OPN antagonists may have therapeutic potential for osteoporosis and other osteopenic diseases.

  8. Humanized-Single Domain Antibodies (VH/VHH) that Bound Specifically to Naja kaouthia Phospholipase A2 and Neutralized the Enzymatic Activity

    PubMed Central

    Chavanayarn, Charnwit; Thanongsaksrikul, Jeeraphong; Thueng-in, Kanyarat; Bangphoomi, Kunan; Sookrung, Nitat; Chaicumpa, Wanpen

    2012-01-01

    Naja kaouthia (monocled cobra) venom contains many isoforms of secreted phospholipase A2 (sPLA2). The PLA2 exerts several pharmacologic and toxic effects in the snake bitten subject, dependent or independent on the enzymatic activity. N. kaouthia venom appeared in two protein profiles, P3 and P5, after fractionating the venom by ion exchange column chromatography. In this study, phage clones displaying humanized-camel single domain antibodies (VH/VHH) that bound specifically to the P3 and P5 were selected from a humanized-camel VH/VHH phage display library. Two phagemid transfected E. coli clones (P3-1 and P3-3) produced humanized-VHH, while another clone (P3-7) produced humanized-VH. At the optimal venom:antibody ratio, the VH/VHH purified from the E. coli homogenates neutralized PLA2 enzyme activity comparable to the horse immune serum against the N. kaouthia holo-venom. Homology modeling and molecular docking revealed that the VH/VHH covered the areas around the PLA2 catalytic groove and inserted their Complementarity Determining Regions (CDRs) into the enzymatic cleft. It is envisaged that the VH/VHH would ameliorate/abrogate the principal toxicity of the venom PLA2 (membrane phospholipid catabolism leading to cellular and subcellular membrane damage which consequently causes hemolysis, hemorrhage, and dermo-/myo-necrosis), if they were used for passive immunotherapy of the cobra bitten victim. The speculation needs further investigations. PMID:22852068

  9. Outward Bound.

    ERIC Educational Resources Information Center

    Outward Bound, Inc., Andover, MA.

    The Outward Bound concept was developed in Germany and Great Britain with the saving of human life as the ultimate goal. Courses are designed to help students discover their true physical and mental limits through development of skills including emergency medical aid, firefighting, search and rescue, mountaineering, and sailing. Five Outward Bound…

  10. Universal bounds on current fluctuations

    NASA Astrophysics Data System (ADS)

    Pietzonka, Patrick; Barato, Andre C.; Seifert, Udo

    2016-05-01

    For current fluctuations in nonequilibrium steady states of Markovian processes, we derive four different universal bounds valid beyond the Gaussian regime. Different variants of these bounds apply to either the entropy change or any individual current, e.g., the rate of substrate consumption in a chemical reaction or the electron current in an electronic device. The bounds vary with respect to their degree of universality and tightness. A universal parabolic bound on the generating function of an arbitrary current depends solely on the average entropy production. A second, stronger bound requires knowledge both of the thermodynamic forces that drive the system and of the topology of the network of states. These two bounds are conjectures based on extensive numerics. An exponential bound that depends only on the average entropy production and the average number of transitions per time is rigorously proved. This bound has no obvious relation to the parabolic bound but it is typically tighter further away from equilibrium. An asymptotic bound that depends on the specific transition rates and becomes tight for large fluctuations is also derived. This bound allows for the prediction of the asymptotic growth of the generating function. Even though our results are restricted to networks with a finite number of states, we show that the parabolic bound is also valid for three paradigmatic examples of driven diffusive systems for which the generating function can be calculated using the additivity principle. Our bounds provide a general class of constraints for nonequilibrium systems.

  11. Thrombin Cleavage of Osteopontin Modulates Its Activities in Human Cells In Vitro and Mouse Experimental Autoimmune Encephalomyelitis In Vivo.

    PubMed

    Boggio, Elena; Dianzani, Chiara; Gigliotti, Casimiro Luca; Soluri, Maria Felicia; Clemente, Nausicaa; Cappellano, Giuseppe; Toth, Erika; Raineri, Davide; Ferrara, Benedetta; Comi, Cristoforo; Dianzani, Umberto; Chiocchetti, Annalisa

    2016-01-01

    Osteopontin is a proinflammatory cytokine and plays a pathogenetic role in multiple sclerosis and its animal model, experimental autoimmune encephalomyelitis (EAE), by recruiting autoreactive T cells into the central nervous system. Osteopontin functions are modulated by thrombin cleavage generating N- and C-terminal fragment, whose individual roles are only partly known. Published data are difficult to compare since they have been obtained with heterogeneous approaches. Interestingly, thrombin cleavage of osteopontin unmasks a cryptic domain of interaction with α 4 β 1 integrin that is the main adhesion molecule involved in lymphocyte transmigration to the brain and is the target for natalizumab, the most potent drug preventing relapses. We produced recombinant osteopontin and its N- and C-terminal fragments in an eukaryotic system in order to allow their posttranslational modifications. We investigated, in vitro, their effect on human cells and in vivo in EAE. We found that the osteopontin cleavage plays a key role in the function of this cytokine and that the two fragments exert distinct effects both in vitro and in vivo. These findings suggest that drugs targeting each fragment may be used to fine-tune the pathological effects of osteopontin in several diseases. PMID:27478856

  12. Thrombin Cleavage of Osteopontin Modulates Its Activities in Human Cells In Vitro and Mouse Experimental Autoimmune Encephalomyelitis In Vivo

    PubMed Central

    Boggio, Elena; Gigliotti, Casimiro Luca; Soluri, Maria Felicia; Clemente, Nausicaa; Toth, Erika; Raineri, Davide; Ferrara, Benedetta; Chiocchetti, Annalisa

    2016-01-01

    Osteopontin is a proinflammatory cytokine and plays a pathogenetic role in multiple sclerosis and its animal model, experimental autoimmune encephalomyelitis (EAE), by recruiting autoreactive T cells into the central nervous system. Osteopontin functions are modulated by thrombin cleavage generating N- and C-terminal fragment, whose individual roles are only partly known. Published data are difficult to compare since they have been obtained with heterogeneous approaches. Interestingly, thrombin cleavage of osteopontin unmasks a cryptic domain of interaction with α4β1 integrin that is the main adhesion molecule involved in lymphocyte transmigration to the brain and is the target for natalizumab, the most potent drug preventing relapses. We produced recombinant osteopontin and its N- and C-terminal fragments in an eukaryotic system in order to allow their posttranslational modifications. We investigated, in vitro, their effect on human cells and in vivo in EAE. We found that the osteopontin cleavage plays a key role in the function of this cytokine and that the two fragments exert distinct effects both in vitro and in vivo. These findings suggest that drugs targeting each fragment may be used to fine-tune the pathological effects of osteopontin in several diseases. PMID:27478856

  13. Fractional diffusion on bounded domains

    DOE PAGESBeta

    Defterli, Ozlem; D'Elia, Marta; Du, Qiang; Gunzburger, Max Donald; Lehoucq, Richard B.; Meerschaert, Mark M.

    2015-03-13

    We found that the mathematically correct specification of a fractional differential equation on a bounded domain requires specification of appropriate boundary conditions, or their fractional analogue. In this paper we discuss the application of nonlocal diffusion theory to specify well-posed fractional diffusion equations on bounded domains.

  14. Osteopontin induces {beta}-catenin signaling through activation of Akt in prostate cancer cells

    SciTech Connect

    Robertson, Brian W.; Chellaiah, Meenakshi A.

    2010-01-01

    Secretion of osteopontin (OPN) by cancer cells is a known mediator of tumorigenesis and cancer progression in both experimental and clinical studies. Our work demonstrates that OPN can activate Akt, an important step in cancer progression. Both ILK and PI3K are integral proteins in the OPN/Akt pathway, as inhibition of either kinase leads to a loss of OPN-mediated Akt activation. Subsequent to OPN-induced Akt activation, we observe inactivation of GSK-3{beta}, a regulator of {beta}-catenin. Osteopontin stimulation leads to an overall increase in {beta}-catenin protein levels with a resultant transfer of {beta}-catenin to the nucleus. Through the nuclear import of {beta}-catenin, OPN increases both the transcription and protein levels of MMP-7 and CD44, which are known TCF/LEF transcription targets. This work describes an important aspect of cancer progression induced by OPN.

  15. Humanized-single domain antibodies (VH/VHH) that bound specifically to Naja kaouthia phospholipase A2 and neutralized the enzymatic activity.

    PubMed

    Chavanayarn, Charnwit; Thanongsaksrikul, Jeeraphong; Thueng-In, Kanyarat; Bangphoomi, Kunan; Sookrung, Nitat; Chaicumpa, Wanpen

    2012-07-01

    Naja kaouthia (monocled cobra) venom contains many isoforms of secreted phospholipase A2 (sPLA(2)). The PLA(2) exerts several pharmacologic and toxic effects in the snake bitten subject, dependent or independent on the enzymatic activity. N. kaouthia venom appeared in two protein profiles, P3 and P5, after fractionating the venom by ion exchange column chromatography. In this study, phage clones displaying humanized-camel single domain antibodies (VH/V(H)H) that bound specifically to the P3 and P5 were selected from a humanized-camel VH/V(H)H phage display library. Two phagemid transfected E. coli clones (P3-1 and P3-3) produced humanized-V(H)H, while another clone (P3-7) produced humanized-VH. At the optimal venom:antibody ratio, the VH/V(H)H purified from the E. coli homogenates neutralized PLA(2) enzyme activity comparable to the horse immune serum against the N. kaouthia holo-venom. Homology modeling and molecular docking revealed that the VH/V(H)H covered the areas around the PLA(2) catalytic groove and inserted their Complementarity Determining Regions (CDRs) into the enzymatic cleft. It is envisaged that the VH/V(H)H would ameliorate/abrogate the principal toxicity of the venom PLA(2) (membrane phospholipid catabolism leading to cellular and subcellular membrane damage which consequently causes hemolysis, hemorrhage, and dermo-/myo-necrosis), if they were used for passive immunotherapy of the cobra bitten victim. The speculation needs further investigations. PMID:22852068

  16. Biochemical Studies and Ligand-bound Structures of Biphenyl Dehydrogenase from Pandoraea pnomenusa Strain B-356 Reveal a Basis for Broad Specificity of the Enzyme*

    PubMed Central

    Dhindwal, Sonali; Patil, Dipak N.; Mohammadi, Mahmood; Sylvestre, Michel; Tomar, Shailly; Kumar, Pravindra

    2011-01-01

    Biphenyl dehydrogenase, a member of short-chain dehydrogenase/reductase enzymes, catalyzes the second step of the biphenyl/polychlorinated biphenyls catabolic pathway in bacteria. To understand the molecular basis for the broad substrate specificity of Pandoraea pnomenusa strain B-356 biphenyl dehydrogenase (BphBB-356), the crystal structures of the apo-enzyme, the binary complex with NAD+, and the ternary complexes with NAD+-2,3-dihydroxybiphenyl and NAD+-4,4′-dihydroxybiphenyl were determined at 2.2-, 2.5-, 2.4-, and 2.1-Å resolutions, respectively. A crystal structure representing an intermediate state of the enzyme was also obtained in which the substrate binding loop was ordered as compared with the apo and binary forms but it was displaced significantly with respect to the ternary structures. These five structures reveal that the substrate binding loop is highly mobile and that its conformation changes during ligand binding, starting from a disorganized loop in the apo state to a well organized loop structure in the ligand-bound form. Conformational changes are induced during ligand binding; forming a well defined cavity to accommodate a wide variety of substrates. This explains the biochemical data that shows BphBB-356 converts the dihydrodiol metabolites of 3,3′-dichlorobiphenyl, 2,4,4′-trichlorobiphenyl, and 2,6-dichlorobiphenyl to their respective dihydroxy metabolites. For the first time, a combination of structural, biochemical, and molecular docking studies of BphBB-356 elucidate the unique ability of the enzyme to transform the cis-dihydrodiols of double meta-, para-, and ortho-substituted chlorobiphenyls. PMID:21880718

  17. Association of osteopontin polymorphism with cancer risk: a meta-analysis

    PubMed Central

    Yang, Gang; Peng, Xiaoxing; Guo, Pengju; Yang, Ge

    2015-01-01

    To investigate the association of osteopontin gene -443 C>T, -156 G>GG, and -1748 A>G polymorphisms with cancer risk. The Medline, PubMed, PUBMED, EMBASE and Web of Science databases were searched. Meta-analyses were conducted using RevMan 5.2 software. After searching and evaluating the included papers, total 10 documents involved in -443 C>T, 8 papers involved in four articles involved in -156 G>GG and -1748 A>G were included into this meta analysis. There were no significant differences in genotype osteopontin -443 C>T distribution between cancer cases and control (OR=0.98, 95% CI=0.68-1.40, P=0.90; OR=0.90, 95% CI=0.60-1.35, P=0.62; OR=0.98, 95% CI=0.59-1.64, P=0.94; OR=0.87, 95% CI=0.60-1.25, P=0.44, respectively). Meanwhile, no association between osteopontin -1748 A>G polymorphism and tumors under all genetic models. (OR=0.73, 95% CI=0.54-1.00, P=0.05; OR=0.95, 95% CI=0.82-1.10, P=0.48; OR=1.31, 95% CI=0.95-1.81, P=0.10; OR=0.90, 95% CI=0.77-1.06, P=0.20, respectively). However, osteopontin -156 G>GG polymorphism is only partly related to the tumor risk. (GGGG+GGG vs GG model, OR=1.21, 95% CI=1.01-1.46, P=0.04; GGG vs GG model: OR=1.19, 95% CI=1.05-1.35, P=0.008, respectively) osteopontin gene polymorphisms, -443 C>T and -1748 A>G was not associated with cancer risk, but partly associated to tumor risk for -156 G>GG gene polymorphism. PMID:26885018

  18. Osteopontin knockdown in the kidneys of hyperoxaluric rats leads to reduction in renal calcium oxalate crystal deposition.

    PubMed

    Tsuji, Hidenori; Shimizu, Nobutaka; Nozawa, Masahiro; Umekawa, Tohru; Yoshimura, Kazuhiro; De Velasco, Marco A; Uemura, Hirotsugu; Khan, Saeed R

    2014-06-01

    Osteopontin (OPN) expression is increased in kidneys of rats with ethylene glycol (EG) induced hyperoxaluria and calcium oxalate (CaOx) nephrolithiasis. The aim of this study is to clarify the effect of OPN knockdown by in vivo transfection of OPN siRNA on deposition of CaOx crystals in the kidneys. Hyperoxaluria was induced in 6-week-old male Sprague-Dawley rats by administering 1.5% EG in drinking water for 2 weeks. Four groups of six rats each were studied: Group A, untreated animals (tap water); Group B, administering 1.5% EG; Group C, 1.5% EG with in vivo transfection of OPN siRNA; Group D, 1.5% EG with in vivo transfection of negative control siRNA. OPN siRNA transfections were performed on day 1 and 8 by renal sub-capsular injection. Rats were killed at day 15 and kidneys were removed. Extent of crystal deposition was determined by measuring renal calcium concentrations and counting renal crystal deposits. OPN siRNA transfection resulted in significant reduction in expression of OPN mRNA as well as protein in group C compared to group B. Reduction in OPN expression was associated with significant decrease in crystal deposition in group C compared to group B. Specific suppression of OPN mRNA expression in kidneys of hyperoxaluric rats leads to a decrease in OPN production and simultaneously inhibits renal crystal deposition. PMID:24619192

  19. Osteopontin knockdown in the kidneys of hyperoxaluric rats leads to reduction in renal calcium oxalate crystal deposition

    PubMed Central

    Shimizu, Nobutaka; Nozawa, Masahiro; Umekawa, Tohru; Yoshimura, Kazuhiro; De Velasco, Marco A.; Uemura, Hirotsugu; Khan, Saeed R.

    2016-01-01

    Osteopontin (OPN) expression is increased in kidneys of rats with ethylene glycol (EG) induced hyperoxaluria and calcium oxalate (CaOx) nephrolithiasis. The aim of this study is to clarify the effect of OPN knockdown by in vivo transfection of OPN siRNA on deposition of CaOx crystals in the kidneys. Hyperoxaluria was induced in 6-week-old male Sprague–Dawley rats by administering 1.5 % EG in drinking water for 2 weeks. Four groups of six rats each were studied: Group A, untreated animals (tap water); Group B, administering 1.5 % EG; Group C, 1.5 % EG with in vivo transfection of OPN siRNA; Group D, 1.5 % EG with in vivo transfection of negative control siRNA. OPN siRNA transfections were performed on day 1 and 8 by renal sub-capsular injection. Rats were killed at day 15 and kidneys were removed. Extent of crystal deposition was determined by measuring renal calcium concentrations and counting renal crystal deposits. OPN siRNA transfection resulted in significant reduction in expression of OPN mRNA as well as protein in group C compared to group B. Reduction in OPN expression was associated with significant decrease in crystal deposition in group C compared to group B. Specific suppression of OPN mRNA expression in kidneys of hyperoxaluric rats leads to a decrease in OPN production and simultaneously inhibits renal crystal deposition. PMID:24619192

  20. Osteopontin modulates inflammation, mucin production, and gene expression signatures after inhalation of asbestos in a murine model of fibrosis.

    PubMed

    Sabo-Attwood, Tara; Ramos-Nino, Maria E; Eugenia-Ariza, Maria; Macpherson, Maximilian B; Butnor, Kelly J; Vacek, Pamela C; McGee, Sean P; Clark, Jessica C; Steele, Chad; Mossman, Brooke T

    2011-05-01

    Inflammation and lung remodeling are hallmarks of asbestos-induced fibrosis, but the molecular mechanisms that control these events are unclear. Using laser capture microdissection (LCM) of distal bronchioles in a murine asbestos inhalation model, we show that osteopontin (OPN) is up-regulated by bronchiolar epithelial cells after chrysotile asbestos exposures. In contrast to OPN wild-type mice (OPN(+/+)) inhaling asbestos, OPN null mice (OPN(-/-)) exposed to asbestos showed less eosinophilia in bronchoalveolar lavage fluids, diminished lung inflammation, and decreased mucin production. Bronchoalveolar lavage fluid concentrations of inflammatory cytokines (IL-1β, IL-4, IL-6, IL-12 subunit p40, MIP1α, MIP1β, and eotaxin) also were significantly less in asbestos-exposed OPN(-/-) mice. Microarrays performed on lung tissues from asbestos-exposed OPN(+/+) and OPN(-/-) mice showed that OPN modulated the expression of a number of genes (Col1a2, Timp1, Tnc, Eln, and Col3a1) linked to fibrosis via initiation and cross talk between IL-1β and epidermal growth factor receptor-related signaling pathways. Novel targets of OPN identified include genes involved in cell signaling, immune system/defense, extracellular matrix remodeling, and cell cycle regulation. Although it is unclear whether the present findings are specific to chrysotile asbestos or would be observed after inhalation of other fibers in general, these results highlight new potential mechanisms and therapeutic targets for asbestosis and other diseases (asthma, smoking-related interstitial lung diseases) linked to OPN overexpression. PMID:21514415

  1. Osteopontin, E-cadherin, and β-catenin expression as prognostic biomarkers in patients with radically resected gastric cancer.

    PubMed

    Di Bartolomeo, Maria; Pietrantonio, Filippo; Pellegrinelli, Alessandro; Martinetti, Antonia; Mariani, Luigi; Daidone, Maria Grazia; Bajetta, Emilio; Pelosi, Giuseppe; de Braud, Filippo; Floriani, Irene; Miceli, Rosalba

    2016-04-01

    A correlation between osteopontin, E-cadherin, β-catenin, and cyclooxygenase 2 overexpression and poor clinicopathological features and prognosis has been previously suggested in gastric cancer. This translational study was aimed at assessing the correlation of these immunohistochemical biomarkers with outcome in patients with radically resected gastric cancer. We analyzed osteopontin, E-cadherin, β-catenin, and cyclooxygenase 2 expression by immunohistochemistry in 346 primary gastric tumor tissue samples from patients enrolled in the ITACA-S trial. This phase III study randomized patients with radically resected gastric cancer to receive adjuvant chemotherapy with either 5-fluorouracil and leucovorin or a sequential regimen of infusional 5-fluorouracil and leucovorin plus irinotecan followed by cisplatin and docetaxel. High expression of osteopontin was correlated with high histological grade, diffuse histotype, and peritoneal relapse, but not with TNM stage. Moreover, osteopontin overexpression was associated with higher risk of tumor recurrence and metastases, and was an independent prognostic factor for both relapse-free and overall survival of gastric cancer patients following adjuvant chemotherapy. Abnormal E-cadherin expression and abnormal β-catenin expression were correlated with more advanced disease stage, and as a consequence, with poor outcome. Our results suggest that osteopontin overexpression is a valuable independent predictor of tumor recurrence and survival in patients with radically resected gastric cancer. PMID:25862567

  2. Genomic organization of the human osteopontin gene: Exclusion of the locus from a causative role in the pathogenesis of dentinogenesis imperfecta type II.

    SciTech Connect

    Crosby, A.H.; Edwards, S.J.; Murray, J.C.

    1995-05-01

    Osteopontin (SPP1) is the principal phosphorylated glycoprotein of bone that is also expressed in a limited number of other tissues including dentine. In the current investigation the authors report the genomic organization of the SPP1 gene, which comprises seven exons, six of which contain coding sequence. The splice sites for exon donor and acceptor positions are in close agreement with previously published consensus sequences. Comparison of the human gene with its murine and bovine counterparts revealed a highly homologous organization. A highly informative short tandem repeat polymorphism isolated at the SPP1 locus showed no recombination with the autosomal dominant disorder dentinogenesis imperfecta type II. Nevertheless, sequencing of each exon in individuals affected by this disorder failed to reveal any disease-specific mutations. 25 refs., 2 figs., 2 tabs.

  3. Expression of Osteopontin in Oral Squamous Cell Carcinoma and its Surgical Margins-An Immunohistochemical Study

    PubMed Central

    Narasimhan, Malathi; Thiyagarajan, Muthukumar; Munuswamy, Balu David; Jayamani, Logeswari

    2015-01-01

    Introduction Despite the advances in the treatment modalities offered for oral squamous cell carcinoma. The recurrence rate of it still remains quite high. Early detection of recurrence will improve the outcome and the survival of the patient. Osteopontin, a transformation–related phosphorylated protein in epithelial cells has been closely related with tumourigenesis. This study was undertaken to explore the potential of OPN as a tumour marker of recurrence in OSCC. Aim To analyse the expression of Osteopontin (OPN) in Oral Squamous Cell Carcinoma (OSCC), patient matched tumour free surgical margins and normal oral mucosa and to correlate with local & loco regional recurrence. Materials and Methods Twenty cases each of formalin fixed paraffin embedded blocks of histopathologically diagnosed cases of OSCC, patient matched tumour free surgical margins and normal oral mucosal tissues were obtained from the archives of the Oral Pathology & Microbiology Department, Faculty of Dental Sciences, SRU and Govt. Arignar Anna Memorial Cancer Hospital, Kancheepuram. Immunohistochemical analysis was performed with an antibody to Osteopontin protein. Patients with secondary tumours and those treated with chemotherapy and radiotherapy were excluded from this study. Results The expression of OPN was elevated in 95% of tumours & 55% of histologically tumour free margin samples. There was negative OPN expression in normal mucosal samples. The result of the study was statistically analysed using Pearson chi-square test and was found to be statistically significant. Conclusion OPN can be used as a diagnostic marker in Oral Squamous Cell Carcinoma. In the tumour free surgical margins, elevated levels of OPN may predict a significantly increased risk of recurrence. PMID:26675878

  4. Post-translational modification of osteopontin: Effects on in vitro hydroxyapatite formation and growth

    SciTech Connect

    Boskey, Adele L.; Christensen, Brian; Taleb, Hayat; Sorensen, Esben S.

    2012-03-09

    Highlights: Black-Right-Pointing-Pointer Thrombin-cleaved fragments of milk-osteopontin effect hydroxyapatite formation differently. Black-Right-Pointing-Pointer N- and C-terminal fragments promoted hydroxyapatite formation and growth. Black-Right-Pointing-Pointer A central fragment inhibited hydroxyapatite formation and growth. Black-Right-Pointing-Pointer Binding to collagen or hydroxyapatite seed crystals modified these effects. -- Abstract: The manuscript tests the hypothesis that posttranslational modification of the SIBLING family of proteins in general and osteopontin in particular modify the abilities of these proteins to regulate in vitro hydroxyapatite (HA) formation. Osteopontin has diverse effects on hydroxyapatite (HA) mineral crystallite formation and growth depending on the extent of phosphorylation. We hypothesized that different regions of full-length OPN would also have distinct effects on the mineralization process. Thrombin fragmentation of milk OPN (mOPN) was used to test this hypothesis. Three fragments were tested in a de novo HA formation assay; an N-terminal fragment (aa 1-147), a central fragment (aa 148-204) denoted SKK-fragment and a C-terminal fragment (aa 205-262). Compared to intact mOPN the C- and N-terminal fragments behaved comparably, promoting HA formation and growth, but the central SKK-fragment acted as a mineralization inhibitor. In a seeded growth experiment all fragments inhibited mineral proliferation, but the SKK-fragment was the most effective inhibitor. These effects, seen in HA-formation and seeded growth assays in a gelatin gel system and in a pH-stat experiment were lost when the protein or fragments were dephosphorylated. Effects of the fully phosphorylated protein and fragments were also altered in the presence of fibrillar collagen. The diverse effects can be explained in terms of the intrinsically disordered nature of OPN and its fragments which enable them to interact with their multiple partners.

  5. Proteins of rough microsomal membranes related to ribosome binding. II. Cross-linking of bound ribosomes to specific membrane proteins exposed at the binding sites

    PubMed Central

    1978-01-01

    Two proteins (ribophorins I and II), which are integral components of rough microsomal membranes and appear to be related to the bound ribosomes, were shown to be exposed on the surface of rat liver rough microsomes (RM) and to be in close proximity to the bound ribosomes. Both proteins were labeled when intact RM were incubated with a lactoperoxidase iodinating system, but only ribophorin I was digested during mild trypsinization of intact RM. Ribophorin II (63,000 daltons) was only proteolyzed when the luminal face of the microsomal vesicles was made accessible to trypsin by the addition of sublytical detergent concentrations. Only 30--40% of the bound ribosomes were released during trypsinization on intact RM, but ribosome release was almost complete in the presence of low detergent concentrations. Very low glutaraldehyde concentrations (0.005--0.02%) led to the preferential cross-linking of large ribosomal subunits of bound ribosomes to the microsomal membranes. This cross-linking prevented the release of subunits caused by puromycin in media of high ionic strength, but not the incorporation of [3H]puromycin into nascent polypeptide chains. SDS- acrylamide gel electrophoresis of cross-linked samples a preferential reduction in the intensity of the bands representing the ribophorins and the formation of aggregates which did not penetrate into the gels. At low methyl-4-mercaptobutyrimidate (MMB) concentrations (0.26 mg/ml) only 30% of the ribosomes were cross-linked to the microsomal membranes, as shown by the puromycin-KCl test, but membranes could still be solubilized with 1% DOC. This allowed the isolation of the ribophorins together with the sedimentable ribosomes, as was shown by electrophoresis of the sediments after disruption of the cross-links by reduction. Experiments with RM which contained only inactive ribosomes showed that the presence of nascent chains was not necessary for the reversible cross-linking of ribosomes to the membranes. These

  6. NMR investigation of the role of osteocalcin and osteopontin at the organic-inorganic interface in bone

    PubMed Central

    Nikel, Ondřej; Laurencin, Danielle; McCallum, Scott A.; Gundberg, Caren M.; Vashishth, Deepak

    2014-01-01

    Mechanical resilience of bone tissue decreases with age. The ability to comprehensively probe and understand bone properties could help alleviate this problem. One important aspect of bone quality which has recently been made evident is the presence of dilatational bands formed by osteocalcin (OC) and osteopontin (OPN), which contribute to fracture toughness. However, experimental evidence of the structural role of these two proteins at the organic-mineral interface in bone is still needed. Solid state nuclear magnetic resonance (SSNMR) is emerging as a useful technique in probing molecular level aspects of bone. Here, we present the first SSNMR study of bone tissue from genetically modified mice lacking OC and/or OPN. Probing the mineral phase, the organic matrix and their interface revealed that despite the absence of OC and OPN, the organic matrix and mineral were well preserved, and the overall exposure of collagen to hydroxyapatite (HA) nanoparticles was hardly affected. However, the proximity to HA surface was slightly increased for a number bone components including less abundant amino acids like lysine, suggesting that this is how the tissue compensates for the lack of OC and OPN. Taken together, the NMR data supports the recently proposed model, in which the contribution of OC - OPN to fracture toughness is related to their presence at the extrafibrillar organic-mineral interfaces, where they reinforce the network of mineralized fibrils and form dilatational bands. In effort towards understanding further the structural role of individual amino acids of low abundance in bone, we then explored the possibility of specific 13C enrichment of mouse bone, and report the first SSNMR spectra of 97% 13C lysine-enriched tissues. Results show that such isotopic enrichment allows valuable molecular-level structural information to be extracted, and sheds light on post-translational modifications undergone by specific amino acids in vivo. PMID:24128197

  7. Osteopontin Expression During the Periparturient Period in Dairy Cows Naturally Infected with Mycobacterium avium subsp. paratuberculosis Infection

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Investigation of the role of osteopontin (Opn) in Johne’s disease is of interest based upon its ability to influence cytokine expression and to improve host defense against mycobacterial infections. The objective of this study was to characterize Opn expression and secretion by peripheral blood mono...

  8. Correlation Between Tumor Growth Delay and Expression of Cancer and Host VEGF, VEGFR2, and Osteopontin in Response to Radiotherapy

    SciTech Connect

    Solberg, Timothy D.; Nearman, Jessica; Mullins, John; Li Sicong; Baranowska-Kortylewicz, Janina

    2008-11-01

    Purpose: To determine the late effects of radiotherapy (RT) on vascular endothelial growth factor (VEGF), VEGF receptor-2 (VEGFR2), and osteopontin (OPN) expression in cancer and stromal cells. Methods and Materials: LS174T xenografted athymic mice were used as a tumor model. Radiation was delivered in two equivalent fractionation schemes: 5 x 7 Gy and 1 x 20 Gy, the latter at two dose rates. Results: Tumor growth arrest was similar in all treatment groups, with the exception of a better response of small-size tumors in the 5 x 7-Gy group. The host VEGF and OPN levels were directly proportional to the tumor doubling time and were independent of the fractionation scheme. The host and cancer cell VEGFR2 levels in tumor were also directly related to the tumor response to RT. Conclusion: Upregulated VEGFR2 in cancer cells suggest paracrine signaling in the VEGFR2 pathway of cancer cells as the factor contributing to RT failure. The transient activation of the host VEGF/VEGFR2 pathway in tumor supports the model of angiogenic regeneration and suggests that radiation-induced upregulation of VEGF, VEGFR2, and downstream proteins might contribute to RT failure by escalating the rate of vascular repair. Coexpression of host OPN and VEGF, two factors closely associated with angiogenesis, indicate that OPN can serve as a surrogate marker of tumor recovery after RT. Taken together, these results strongly support the notion that to achieve optimal therapeutic outcomes, the scheduling of RT and antiangiogenic therapies will require patient-specific post-treatment monitoring of the VEGF/VEGFR2 pathway and that tumor-associated OPN can serve as an indicator of tumor regrowth.

  9. Potential roles of osteopontin and αVβ3 integrin in the development of coronary artery restenosis after angioplasty

    PubMed Central

    Panda, Dibyendu; Kundu, Gopal C.; Lee, Benjamin I.; Peri, Alessandro; Fohl, David; Chackalaparampil, Isaac; Mukherjee, Barid B.; Li, Xiao D.; Mukherjee, Diane C.; Seides, Stuart; Rosenberg, Joel; Stark, Karen; Mukherjee, Anil B.

    1997-01-01

    Angioplasty procedures are increasingly used to reestablish blood flow in blocked atherosclerotic coronary arteries. A serious complication of these procedures is reocclusion (restenosis), which occurs in 30–50% of patients. Migration of coronary artery smooth muscle cells (CASMCs) to the site of injury caused by angioplasty and subsequent proliferation are suggested mechanisms of reocclusion. Using both cultured human CASMCs and coronary atherectomy tissues, we studied the roles of osteopontin (OPN) and one of its receptors, αvβ3 integrin, in the pathogenesis of coronary restenosis. We also measured the plasma levels of OPN before and after angioplasty and determined the effect of exogenous OPN on CASMC migration, extracellular matrix invasion, and proliferation. We found that cultured CASMCs during log phase of growth and smooth muscle cell layer of the coronary atherosclerotic tissues of patients express both OPN mRNA and protein at a significantly elevated level compared with controls. Interestingly, whereas the baseline plasma OPN levels in control samples were virtually undetectable, those in patient plasma were remarkably high. We also found that interaction of OPN with αvβ3 integrin, expressed on CASMCs, causes migration, extracellular matrix invasion, and proliferation. These effects were abolished when OPN or αvβ3 integrin gene expression in CASMCs was inhibited by specific antisense S-oligonucleotide treatment or OPN-αvβ3 interaction was blocked by treatment of CASMCs with antibodies against OPN or αvβ3 integrin. Our results demonstrate that OPN and αvβ3 integrin play critical roles in regulating cellular functions deemed essential for restenosis. In addition, these results raise the possibility that transient inhibition of OPN gene expression or blocking of OPN-αvβ3 interaction may provide a therapeutic approach to preventing restenosis. PMID:9256478

  10. Increased Expression of Osteopontin in Retinal Degeneration Induced by Blue Light-Emitting Diode Exposure in Mice.

    PubMed

    Chang, Seung Wook; Kim, Hyung Il; Kim, Gyu Hyun; Park, Su Jin; Kim, In-Beom

    2016-01-01

    Osteopontin (OPN) is a multifunctional adhesive glycoprotein that is implicated in a variety of pro-inflammatory as well as neuroprotective and repair-promoting effects in the brain. As a first step towards understanding the role of OPN in retinal degeneration (RD), we examined changes in OPN expression in a mouse model of RD induced by exposure to a blue light-emitting diode (LED). RD was induced in BALB/c mice by exposure to a blue LED (460 nm) for 2 h. Apoptotic cell death was evaluated by terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) assay. In order to investigate changes in OPN in RD, western blotting and immunohistochemistry were performed. Anti-OPN labeling was compared to that of anti-glial fibrillary acidic protein (GFAP), which is a commonly used marker for retinal injury or stress including inflammation. OPN expression in RD retinas markedly increased at 24 h after exposure, was sustained through 72 h, and subsided at 120 h. Increased OPN expression was observed co-localized with microglial cells in the outer nuclear layer (ONL), outer plexiform layer (OPL), and subretinal space. Expression was restricted to the central retina in which photoreceptor cell death occurred. Interestingly, OPN expression in the ONL/OPL was closely associated with microglia, whereas most of the OPN plaques observed in the subretinal space were not. Immunogold electron microscopy demonstrated that OPN was distributed throughout the cytoplasm of microglia and in nearby fragments of degenerating photoreceptors. In addition, we found that OPN was induced more acutely and with greater region specificity than GFAP. These results indicate that OPN may be a more useful marker for retinal injury or stress, and furthermore act as a microglial pro-inflammatory mediator and a phagocytosis-inducing opsonin in the subretinal space. Taken together, our data suggest that OPN plays an important role in the pathogenesis of RD. PMID:27504084

  11. Prevention of neointimal hyperplasia in balloon-injured rat carotid artery via small interference RNA mediated downregulation of osteopontin gene.

    PubMed

    Xu, Jian; Sun, Yingxian; Wang, Tairan; Liu, Guinan

    2013-05-01

    The aim of the present study was to take osteopontin (OPN) as molecular target to study its effects on injured intima model of carotid artery in rat using perivascular transfer of OPN-small interference RNA (siRNA). OPN mRNA in cultured VSMCs was quantified by real-time RT-PCR, and OPN-siRNA-002 was determined as the most sensitive sequence and used as transfected siRNA in the subsequent animal experiments. We established rat carotid arterial intima-injured model with balloon-injured method, and then perivascularly transfected OPN-siRNA-002 to study the role of OPN-siRNA in regulating several related genes including proliferating cell nuclear antigen (PCNA), transforming growth factor β1(TGF-β1), matrix metalloproteinase-2 (MMP-2), and matrix metalloproteinase-14 (MMP-14), as well as its role in neointimal formation. OPN mRNA and protein decreased about 50 % with corresponding decrease in intima thickness after transfecting with specific OPN-siRNA-002 compared with Pluronic control group and OPN-SCR-siRNA group on each time point (n = 6, p < 0.001), and this inhibiting effects persisted up to 14 days after balloon injury. PCNA, TGF-β1, MMP-2, and MMP-14 mRNA and protein correlated directly with the respective levels of OPN, suggesting its functions via regulating these downstream factors (n = 6, p < 0.001). OPN may be a potential target gene in reducing the risk for arterial restenosis after vascular intervention. PMID:23467880

  12. Increased Expression of Osteopontin in Retinal Degeneration Induced by Blue Light-Emitting Diode Exposure in Mice

    PubMed Central

    Chang, Seung Wook; Kim, Hyung Il; Kim, Gyu Hyun; Park, Su Jin; Kim, In-Beom

    2016-01-01

    Osteopontin (OPN) is a multifunctional adhesive glycoprotein that is implicated in a variety of pro-inflammatory as well as neuroprotective and repair-promoting effects in the brain. As a first step towards understanding the role of OPN in retinal degeneration (RD), we examined changes in OPN expression in a mouse model of RD induced by exposure to a blue light-emitting diode (LED). RD was induced in BALB/c mice by exposure to a blue LED (460 nm) for 2 h. Apoptotic cell death was evaluated by terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) assay. In order to investigate changes in OPN in RD, western blotting and immunohistochemistry were performed. Anti-OPN labeling was compared to that of anti-glial fibrillary acidic protein (GFAP), which is a commonly used marker for retinal injury or stress including inflammation. OPN expression in RD retinas markedly increased at 24 h after exposure, was sustained through 72 h, and subsided at 120 h. Increased OPN expression was observed co-localized with microglial cells in the outer nuclear layer (ONL), outer plexiform layer (OPL), and subretinal space. Expression was restricted to the central retina in which photoreceptor cell death occurred. Interestingly, OPN expression in the ONL/OPL was closely associated with microglia, whereas most of the OPN plaques observed in the subretinal space were not. Immunogold electron microscopy demonstrated that OPN was distributed throughout the cytoplasm of microglia and in nearby fragments of degenerating photoreceptors. In addition, we found that OPN was induced more acutely and with greater region specificity than GFAP. These results indicate that OPN may be a more useful marker for retinal injury or stress, and furthermore act as a microglial pro-inflammatory mediator and a phagocytosis-inducing opsonin in the subretinal space. Taken together, our data suggest that OPN plays an important role in the pathogenesis of RD. PMID:27504084

  13. Comparative immunogenicity of conjugates composed of Escherichia coli O111 O-specific polysaccharide, prepared by treatment with acetic acid or hydrazine, bound to tetanus toxoid by two synthetic schemes.

    PubMed Central

    Gupta, R K; Egan, W; Bryla, D A; Robbins, J B; Szu, S C

    1995-01-01

    Escherichia coli O111, of various H types and virulence factors, causes enteritis throughout the world, especially in young children. This O type is found rarely in healthy individuals. Serum antibodies to the O-specific polysaccharide of O111 lipopolysaccharide (LPS) protect mice and dogs against infection with this E. coli serotype. The O111 O-specific polysaccharide is composed of a pentasaccharide repeat unit with two colitoses bound to the C-3 and C-6 of glucose in a trisaccharide backbone; this structure is identical to that of Salmonella adelaide (O35), another enteric pathogen. Nonpyrogenic O111 O-specific polysaccharide was prepared by treatment of its LPS with acetic acid (O-SP) or the organic base hydrazine (DeA-LPS). The O-SP had a reduced concentration of colitose. These products were derivatized with adipic acid dihydrazide (ADH) or thiolated with N-succinimidyl-3(2-pyridyldithio) propionate (SPDP). The four derivatives were covalently bound to tetanus toxoid (TT) by carbodiimide-mediated condensation or with SPDP to form conjugates. Immunization of BALB/c and general-purpose mice by a clinically acceptable route showed that DeA-LPS-TTADH, of the four conjugates, elicited the highest level of LPS antibodies. Possible reasons to explain this differential immunogenicity between the four conjugates are discussed. PMID:7542631

  14. Nano-biosensor development for bacterial detection during human kidney infection: use of glycoconjugate-specific antibody-bound gold NanoWire arrays (GNWA).

    PubMed

    Basu, Manju; Seggerson, Sara; Henshaw, Joshua; Jiang, Juan; del A Cordona, Rocio; Lefave, Clare; Boyle, Patrick J; Miller, Albert; Pugia, Michael; Basu, Subhash

    2004-01-01

    Infectious disease, commonly caused by bacterial pathogens, is now the world's leading cause of premature death and third overall cause behind cardiovascular disease and cancer. Urinary Tract Infection (UTI), caused by E. coli bacteria, is a very common bacterial infection, a majority in women (85%) and may result in severe kidney failure if not detected quickly. Among hundreds of strains the bacteria, E. coli 0157:H7, is emerging as the most aggressive one because of its capability to produce a toxin causing hemolytic uremic syndrome (HUS) resulting in death, especially in children. In the present study, a project has been undertaken for developing a rapid method for UTI detection in very low bacteria concentration, applying current knowledge of nano-technology. Experiments have been designed for the development of biosensors using nano-fabricated structures coated with elements such as gold that have affinity for biomolecules. A biosensor is a device in which a biological sensing element is either intimately connected to or integrated within a transducer. The basic principle for the detection procedure of the infection is partly based on the enzyme-linked immunosorbent assay system. Anti-E. coli antibody-bound Gold Nanowire Arrays (GNWA) prepared on anodized porous alumina template is used for the primary step followed by binding of the bacteria containing specimen. An alkaline phosphatase-conjugated second antibody is then added to the system and the resultant binding determined by both electrochemical and optical measurements. Various kinds of GNWA templates were used in order to determine the one with the best affinity for antibody binding. In addition, an efficient method for enhanced antibody binding has been developed with the covalent immobilization of an organic linker Dithiobissuccinimidylundecanoate (DSU) on the GNWA surface. Studies have also been conducted to optimize the antibody-binding conditions to the linker-attached GNWA surfaces for their

  15. Intracellular osteopontin stabilizes TRAF3 to positively regulate innate antiviral response

    PubMed Central

    Zhao, Kai; Zhang, Meng; Zhang, Lei; Wang, Peng; Song, Guanhua; Liu, Bingyu; Wu, Haifeng; Yin, Zhinan; Gao, Chengjiang

    2016-01-01

    Osteopontin (OPN) is a multifunctional protein involved in both innate immunity and adaptive immunity. However, the function of OPN, especially the intracellular form OPN (iOPN) on innate antiviral immune response remains elusive. Here, we demonstrated that iOPN is an essential positive regulator to protect the host from virus infection. OPN deficiency or knockdown significantly attenuated virus-induced IRF3 activation, IFN-β production and antiviral response. Consistently, OPN-deficient mice were more susceptible to VSV infection than WT mice. Mechanistically, iOPN was found to interact with tumor necrosis factor receptor (TNFR)-associated factor 3 (TRAF3) and inhibit Triad3A-mediated K48-linked polyubiquitination and degradation of TRAF3 through the C-terminal fragment of iOPN. Therefore, our findings delineated a new function for iOPN to act as a positive regulator in innate antiviral immunity through stabilization of TRAF3. PMID:27026194

  16. The Osteopontin Transgenic Mouse is a New Model for Sjögren’s Syndrome

    PubMed Central

    Husain-Krautter, Sehba; Kramer, Jill M.; Li, Wentian; Guo, Benchang; Rothstein, Thomas L.

    2015-01-01

    Osteopontin (Opn) is a cytokine involved in both physiological and pathological processes, and is elevated in many autoimmune diseases. Sjögren’s syndrome (SS) is an autoimmune disease with a strong female predilection characterized by lymphocytic infiltration of exocrine glands. We hypothesized Opn contributes to SS pathogenesis. We examined an established SS model, and found increased Opn locally and systemically. Next, we examined Opn transgenic (Opn Tg) mice for evidence of SS. Opn Tg animals exhibited lymphocytic infiltration of salivary and lacrimal glands, and Opn co-localized with the infiltrates. Moreover, saliva production was reduced, and SS autoantibodies were observed in the serum of these mice. Finally, female Opn Tg mice showed more severe disease compared to males. Taken together, these data support a role for Opn in SS pathogenesis. We identify a new model of spontaneous SS that recapitulates the human disease in terms of sex predilection, histopathology, salivary deficits, and autoantibodies. PMID:25572532

  17. The osteopontin transgenic mouse is a new model for Sjögren's syndrome.

    PubMed

    Husain-Krautter, Sehba; Kramer, Jill M; Li, Wentian; Guo, Benchang; Rothstein, Thomas L

    2015-03-01

    Osteopontin (Opn) is a cytokine involved in both physiological and pathological processes, and is elevated in many autoimmune diseases. Sjögren's syndrome (SS) is an autoimmune disease with a strong female predilection characterized by lymphocytic infiltration of exocrine glands. We hypothesized that Opn contributes to SS pathogenesis. We examined an established SS model and found increased Opn locally and systemically. Next, we examined Opn transgenic (Opn Tg) mice for evidence of SS. Opn Tg animals exhibited lymphocytic infiltration of salivary and lacrimal glands, and Opn co-localized with the infiltrates. Moreover, saliva production was reduced, and SS autoantibodies were observed in the serum of these mice. Finally, female Opn Tg mice showed more severe disease compared to males. Taken together, these data support a role for Opn in SS pathogenesis. We identify a new model of spontaneous SS that recapitulates the human disease in terms of sex predilection, histopathology, salivary deficits, and autoantibodies. PMID:25572532

  18. Parthenolide Induces Apoptosis in Committed Progenitor AML Cell line U937 via Reduction in Osteopontin

    PubMed Central

    Zahedpanah, Mahdi; Shaiegan, Mojgan; Ghaffari, Seyed Hamidollah; Nikbakht, Mohsen; Nikugoftar, Mahin; Mohammadi, Saeed

    2016-01-01

    Background: Interfering with cell proliferation and survival is a critical role for antineoplastic drugs leading to cell death through induction of apoptosis. Alternative treatments with herbal extracts offer insights into acute myeloid leukemia (AML) therapy. Parthenolide (PTL), an extract from feverfew, induces apoptosis in primary human leukemia stem cells (LSCs) and bulk leukemic cell populations. Osteopontin (OPN) preserves cell viability in response to anticancer agents and its receptors could be utilized for therapeutic targeting of cancer cells. Methods: U937 cells were cultured in RPMI 1640 with concentrations of 2, 4, 6, 8, and 10 µM PTL for 20-24 hours for MTT assays. Apoptosis assays were performed with Annexin V-Alexa Fluor-488/PI as Annexin V+/PI- and Annexin V+/PI+ to measure early and late apoptosis, respectively. Quantitative real-time PCR was used to measure OPN gene expression using the 2-ΔΔCt method. The PTL–treated cells were stained with FITC-CD38 antibody for flow cytometry analyses. Data were compared using one-way analysis of variance (ANOVA) by SPSS 19. Results: Parthenolide inhibited growth of U937 cells with IC25 and IC50 values of 4 and 5.8 µM, respectively. Death induction with PTL was apoptotic. Flow cytometry showed a significant decrease in the percentage of CD38+ U937 cells in response to PTL. Osteopontin gene expression decreased in response to PTL. Conclusion: PTL induced apoptosis and reduced OPN gene expression in U937 cells.

  19. Osteopontin Involves Cisplatin Resistance and Poor Prognosis in Oral Squamous Cell Carcinoma

    PubMed Central

    Luo, Sheng-Dean; Chen, Yi-Ju; Liu, Chien-Ting; Rau, Kun-Ming; Chen, Yi-Ching; Tsai, Hsin-Ting; Chen, Chang-Han; Chiu, Tai-Jan

    2015-01-01

    Background. Osteopontin (OPN) is a multifunctional cytokine involved in cell survival, migration, and adhesion. However, its role in chemosensitivity in locally advanced oral squamous cell carcinoma (OSCC) in humans has not yet been investigated. Methods. We enrolled 121 patients with locally advanced stage IVA/B OSCC receiving cisplatin-based IC followed by CCRT from January 1, 2006, through January 1, 2012. Immunohistochemistry was used to assess OPN expression in OSCC patients' biopsy specimens from paraffin blocks before treatment. In addition, MTT/colony formation assay was used to estimate the influence of OPN in an oral cancer cell line treated with cisplatin. Results. Of the 121 patients, 94 had positive OPN findings and 52 responded to IC followed by CCRT. Positive osteopontin immunostaining also correlated significantly with positive N status/TNM stage/male gender and smoking. Univariate analyses showed that patients whose tumors had a low expression of OPN were more likely to respond to chemotherapy and have a significantly better OS than those whose tumors had a high expression of OPN. Multivariate analysis revealed that prolonged survival was independently predicted for patients with stage IVA disease, negative lymph nodes, and negative expressions of OPN and for those who received chemotherapy with Docetaxel/cisplatin/fluorouracil (TPF). An oral cancer line stimulated with OPN exhibited a dose-dependent resistance to cisplatin treatment. Conversely, endogenous OPN depletion by OPN-mediated shRNA increased sensitivity to cisplatin. Conclusions. A positive expression of OPN predicts a poor response and survival in patients with locally advanced stage IVA/B OSCC treated with cisplatin-based IC followed by CCRT. PMID:26491674

  20. Perinatal exposure to low-dose DE-71 increases serum thyroid hormones and gonadal osteopontin gene expression

    PubMed Central

    Blake, Charles A; McCoy, George L; Hui, Yvonne Y; LaVoie, Holly A

    2014-01-01

    Polybrominated diphenyl ethers (PBDEs) are flame retardants that have been widely used in manufacturing. They are major household and environmental contaminants that bioaccumulate. Humans are exposed primarily through dust inhalation and dietary ingestion of animal products. In animal studies, high doses of penta-brominated diphenyl ethers (penta-BDEs) in the mg/kg body weight (BW) range negatively impact brain development, behavior, memory, circulating thyroid hormone concentrations, the reproductive system and bone development. We investigated the effects of ingestion of a relatively low dose of the penta-BDE mixture DE-71 by pregnant and lactating rats on reproductive and thyroid parameters of the F1 offspring. F0 mothers received 60 μg/kg BW of DE-71 or vehicle daily by gavage from Day 1.5 of pregnancy through lactation (except the day of parturition). F1 pups were sacrificed at 21 d of age or outbred at approximately 80 d of age. Bred F1 females were sacrificed at Day 14.5 of pregnancy or at five months of age. Bred F1 males were sacrificed at five months of age. DE-71 treatment of the mothers affected the F1 females as evidenced by lower body weights at 80 d and five months of age, elevated serum T3 and T4 concentrations at Day 14.5 of pregnancy and increased thyroid gland weight and ovarian osteopontin mRNA at five months of age. Perinatal DE-71 exposure also increased testicular osteopontin mRNA in 21-day-old F1 males. Utilizing a granulosa cell in vitro model, we demonstrated that DE-71 activated the rat osteopontin gene promoter. Our results are the first to demonstrate that PBDEs increase rodent circulating T3 and T4 concentrations and gonadal osteopontin mRNA, and activate the osteopontin gene promoter. These changes may have clinical implications as others have shown associations between human exposure to PBDEs and subclinical hyperthyroidism, and overexpression of ovarian osteopontin has been associated with ovarian cancer. PMID:21367881

  1. Membrane-bound Dictyostelium myosin heavy chain kinase: a developmentally regulated substrate-specific member of the protein kinase C family.

    PubMed Central

    Ravid, S; Spudich, J A

    1992-01-01

    A cDNA clone corresponding to the Dictyostelium myosin heavy chain kinase (MHCK) gene was isolated using antibodies specific to the purified enzyme. Sequence analysis of the cDNA revealed that the Dictyostelium MHCK possesses all of the domains characteristic of members of the protein kinase C family. The amino-terminal region of the MHCK contains the cysteine-rich motif with an internal duplication that is present in all known protein kinase C species. This domain precedes sequences that are highly homologous to protein kinase catalytic domains. The carboxyl-terminal region contains a cluster of 23 serine and threonine residues that may represent the autophosphorylation domain of the Dictyostelium MHCK. These results, along with previous studies that indicate that this enzyme has very restrictive substrate specificity, incorporates approximately 20 mol of phosphate per mol of kinase through an autophosphorylation reaction, and is expressed only during development, suggest that the Dictyostelium MHCK is a distinct member of the protein kinase C family and imply that this kinase family, which may include members with very specific cellular functions, may be even more heterogeneous than previously thought. Images PMID:1321427

  2. Heterosubtypic Antibodies to Influenza A Virus Have Limited Activity against Cell-Bound Virus but Are Not Impaired by Strain-Specific Serum Antibodies

    PubMed Central

    Wyrzucki, Arkadiusz; Bianchi, Matteo; Kohler, Ines; Steck, Marco

    2014-01-01

    ABSTRACT The majority of influenza virus-specific antibodies elicited by vaccination or natural infection are effective only against the eliciting or closely related viruses. Rare stem-specific heterosubtypic monoclonal antibodies (hMAbs) can neutralize multiple strains and subtypes by preventing hemagglutinin (HA)-mediated fusion of the viral membrane with the endosomal membrane. The epitopes recognized by these hMAbs are therefore considered promising targets for the development of pan-influenza virus vaccines. Here, we report the isolation of a novel human HA stem-reactive monoclonal antibody, hMAb 1.12, with exceptionally broad neutralizing activity encompassing viruses from 15 distinct HA subtypes. Using MAb 1.12 and two other monoclonal antibodies, we demonstrate that neutralization by hMAbs is virtually irreversible but becomes severely impaired following virus attachment to cells. In contrast, no interference by human anti-influenza virus serum antibodies was found, indicating that apically binding antibodies do not impair access to the membrane-proximal heterosubtypic epitopes. Our findings therefore encourage development of new vaccine concepts aiming at the induction of stem-specific heterosubtypic antibodies, as we provide support for their effectiveness in individuals previously exposed to influenza virus. IMPORTANCE The influenza A virus hemagglutinin (HA) can easily accommodate changes in its antigenic structures to escape preexisting immunity. This variability restricts the breadth and long-term efficacy of influenza vaccines. Only a few heterosubtypic antibodies (hMAbs), i.e., antibodies that can neutralize more than one subtype of influenza A virus, have been identified. The molecular interactions between these heterosubtypic antibodies and hemagglutinin are well characterized, yet little is known about the functional properties of these antibodies. Using a new, extraordinarily broad hMAb, we show that virus neutralization by hMAbs is virtually

  3. Effect of gingival application of melatonin on alkaline and acid phosphatase, osteopontin and osteocalcin in patients with diabetes and periodontal disease

    PubMed Central

    López-Valverde, Antonio; Gómez-de-Diego, Rafel; Arias-Santiago, Salvador; de Vicente-Jiménez, Joaquín

    2013-01-01

    Objectives: To assess the effect of topical application of melatonin to the gingiva on salivary fluid concentrations of acid phosphatase, alkaline phosphatase, osteopontin, and osteocalcin. Study Design: Cross-sectional study of 30 patients with diabetes and periodontal disease and 30 healthy subjects. Diabetic patients were treated with topical application of melatonin (1% orabase cream formula) once daily for 20 days and controls with a placebo formulation. Results: Before treatment with melatonin, diabetic patients showed significantly higher mean salivary levels of alkaline and acid phosphatase, osteopontin and osteocalcin than healthy subjects (P < 0.01). After treatment with melatonin, there was a statistically significant decrease of the gingival index (15.84± 10.3 vs 5.6 ± 5.1) and pocket depth (28.3 ± 19.5 vs 11.9 ± 9.0) (P < 0.001). Also, use of melatonin was associated with a significant reduction of the four biomarkers. Changes of salivary acid phosphatase and osteopontin correlated significantly with changes in the gingival index, whereas changes of alkaline phosphatase and osteopontin correlated significantly with changes in the pocket depth. Conclusions: Treatment with topical melatonin was associated with an improvement in the gingival index and pocket depth, a reduction in salivary concentrations of acid phosphatase, alkaline phosphatase, osteopontin and osteocalcin. Key words:Melatonin, diabetes mellitus, alkaline phosphatase, acid phosphatase, osteopontin, osteocalcin. PMID:23524437

  4. U2504 Determines the Species Specificity of the A-site Cleft Antibiotics: The sStructures of Tiamulin, Homoharringtonine and Bruceantin Bound to the Ribosome

    SciTech Connect

    Gurel, G.; Blaha, G; Moore, P; Steitz,

    2009-01-01

    Structures have been obtained for the complexes that tiamulin, homoharringtonine, and bruceantin form with the large ribosomal subunit of Haloarcula marismortui at resolutions ranging from 2.65 to 3.2 {angstrom}. They show that all these inhibitors block protein synthesis by competing with the amino acid side chains of incoming aminoacyl-tRNAs for binding in the A-site cleft in the peptidyl-transferase center, which is universally conserved. In addition, these structures support the hypothesis that the species specificity exhibited by the A-site cleft inhibitors is determined by the interactions they make, or fail to make, with a single nucleotide, U2504 (Escherichia coli). In the ribosome, the position of U2504 is controlled by its interactions with neighboring nucleotides, whose identities vary among kingdoms.

  5. U2504 Determines the Species Specificity of the A-Site Cleft Antibiotics: The Structures of Tiamulin, Homoharringtonine, and Bruceantin Bound to the Ribosome

    SciTech Connect

    Gürel, Güliz; Blaha, Gregor; Moore, Peter B.; Steitz, Thomas A.; Yale

    2009-06-30

    Structures have been obtained for the complexes that tiamulin, homoharringtonine, and bruceantin form with the large ribosomal subunit of Haloarcula marismortui at resolutions ranging from 2.65 to 3.2 {angstrom}. They show that all these inhibitors block protein synthesis by competing with the amino acid side chains of incoming aminoacyl-tRNAs for binding in the Asite cleft in the peptidyl-transferase center, which is universally conserved. In addition, these structures support the hypothesis that the species specificity exhibited by the A-site cleft inhibitors is determined by the interactions they make, or fail to make, with a single nucleotide, U2504 (Escherichia coli). In the ribosome, the position of U2504 is controlled by its interactions with neighboring nucleotides, whose identities vary among kingdoms.

  6. Identification and characterization of the type-IVA cyclic AMP-specific phosphodiesterase RD1 as a membrane-bound protein expressed in cerebellum.

    PubMed

    Shakur, Y; Wilson, M; Pooley, L; Lobban, M; Griffiths, S L; Campbell, A M; Beattie, J; Daly, C; Houslay, M D

    1995-03-15

    An antiserum was generated against a dodecapeptide whose sequence is found at the C-terminus of a cyclic AMP (cAMP)-specific, type-IVA phosphodiesterase encoded by the rat 'dunc-like' cyclic AMP phosphodiesterase (RD1) cDNA. This antiserum identified a single approximately 73 kDa protein species upon immunoblotting of cerebellum homogenates. This species co-migrated upon SDS/PAGE with a single immunoreactive species observed in COS cells transfected with the cDNA for RD1. Native RD1 in cerebellum was found to be predominantly (approximately 93%) membrane-associated and could be found in isolated synaptosome populations, in particular those enriched in post-synaptic densities. Fractionation of lysed synaptosomes on sucrose density gradients identified RD1 as co-migrating with the plasma membrane marker 5'-nucleotidase. Laser scanning confocal and digital deconvolution immunofluorescence studies done on intact COS cells transfected with RD1 cDNA showed RD1 to be predominantly localized to plasma membranes but also associated with the Golgi apparatus and intracellular vesicles. RD1-specific antisera immunoprecipitated phosphodiesterase activity from solubilized cerebellum membranes. This activity had the characteristics expected of the type-IV cAMP phosphodiesterase RD1 in that it was cAMP specific, exhibited a low Km cAMP of 2.3 microM, high sensitivity to inhibition by 4-[3-(cyclopentoxyl)-4-methoxyphenyl]-2-pyrrolidone (rolipram) (Ki approximately 0.7 microM) and was unaffected by Ca2+/calmodulin and low concentrations of cyclic GMP. The phosphodiesterase activities of RD1 solubilized from both cerebellum and transfected COS cell membranes showed identical first-order thermal denaturation kinetics at 50 degrees C. Native RD1 from cerebellum was shown to be an integral protein in that it was solubilized using the non-ionic detergent Triton X-100 but not by either re-homogenization or high NaCl concentrations. The observation that hydroxylamine was unable to cause

  7. Ophiophagus hannah Venom: Proteome, Components Bound by Naja kaouthia Antivenin and Neutralization by N. kaouthia Neurotoxin-Specific Human ScFv

    PubMed Central

    Danpaiboon, Witchuda; Reamtong, Onrapak; Sookrung, Nitat; Seesuay, Watee; Sakolvaree, Yuwaporn; Thanongsaksrikul, Jeeraphong; Dong-din-on, Fonthip; Srimanote, Potjanee; Thueng-in, Kanyarat; Chaicumpa, Wanpen

    2014-01-01

    Venomous snakebites are an important health problem in tropical and subtropical countries. King cobra (Ophiophagus hannah) is the largest venomous snake found in South and Southeast Asia. In this study, the O. hannah venom proteome and the venom components cross-reactive to N. kaouthia monospecific antivenin were studied. O. hannah venom consisted of 14 different protein families, including three finger toxins, phospholipases, cysteine-rich secretory proteins, cobra venom factor, muscarinic toxin, L-amino acid oxidase, hypothetical proteins, low cysteine protein, phosphodiesterase, proteases, vespryn toxin, Kunitz, growth factor activators and others (coagulation factor, endonuclease, 5’-nucleotidase). N. kaouthia antivenin recognized several functionally different O. hannah venom proteins and mediated paratherapeutic efficacy by rescuing the O. hannah envenomed mice from lethality. An engineered human ScFv specific to N. kaouthia long neurotoxin (NkLN-HuScFv) cross-neutralized the O. hannah venom and extricated the O. hannah envenomed mice from death in a dose escalation manner. Homology modeling and molecular docking revealed that NkLN-HuScFv interacted with residues in loops 2 and 3 of the neurotoxins of both snake species, which are important for neuronal acetylcholine receptor binding. The data of this study are useful for snakebite treatment when and where the polyspecific antivenin is not available. Because the supply of horse-derived antivenin is limited and the preparation may cause some adverse effects in recipients, a cocktail of recombinant human ScFvs for various toxic venom components shared by different venomous snakes, exemplified by the in vitro produced NkLN-HuScFv in this study, should contribute to a possible future route for an improved alternative to the antivenins. PMID:24828754

  8. Ophiophagus hannah venom: proteome, components bound by Naja kaouthia antivenin and neutralization by N. kaouthia neurotoxin-specific human ScFv.

    PubMed

    Danpaiboon, Witchuda; Reamtong, Onrapak; Sookrung, Nitat; Seesuay, Watee; Sakolvaree, Yuwaporn; Thanongsaksrikul, Jeeraphong; Dong-din-on, Fonthip; Srimanote, Potjanee; Thueng-in, Kanyarat; Chaicumpa, Wanpen

    2014-05-01

    Venomous snakebites are an important health problem in tropical and subtropical countries. King cobra (Ophiophagus hannah) is the largest venomous snake found in South and Southeast Asia. In this study, the O. hannah venom proteome and the venom components cross-reactive to N. kaouthia monospecific antivenin were studied. O. hannah venom consisted of 14 different protein families, including three finger toxins, phospholipases, cysteine-rich secretory proteins, cobra venom factor, muscarinic toxin, L-amino acid oxidase, hypothetical proteins, low cysteine protein, phosphodiesterase, proteases, vespryn toxin, Kunitz, growth factor activators and others (coagulation factor, endonuclease, 5'-nucleotidase). N. kaouthia antivenin recognized several functionally different O. hannah venom proteins and mediated paratherapeutic efficacy by rescuing the O. hannah envenomed mice from lethality. An engineered human ScFv specific to N. kaouthia long neurotoxin (NkLN-HuScFv) cross-neutralized the O. hannah venom and extricated the O. hannah envenomed mice from death in a dose escalation manner. Homology modeling and molecular docking revealed that NkLN-HuScFv interacted with residues in loops 2 and 3 of the neurotoxins of both snake species, which are important for neuronal acetylcholine receptor binding. The data of this study are useful for snakebite treatment when and where the polyspecific antivenin is not available. Because the supply of horse-derived antivenin is limited and the preparation may cause some adverse effects in recipients, a cocktail of recombinant human ScFvs for various toxic venom components shared by different venomous snakes, exemplified by the in vitro produced NkLN-HuScFv in this study, should contribute to a possible future route for an improved alternative to the antivenins. PMID:24828754

  9. Antibodies from patients with heparin-induced thrombocytopenia/thrombosis are specific for platelet factor 4 complexed with heparin or bound to endothelial cells.

    PubMed Central

    Visentin, G P; Ford, S E; Scott, J P; Aster, R H

    1994-01-01

    Heparin-induced thrombocytopenia/thrombosis (HITP) is thought to be mediated by immunoglobulins that activate platelets in the presence of pharmacologic concentrations of heparin, but the molecular basis for this relatively common and often serious complication of heparin therapy has not been established. We found that plasma from each of 12 patients with HITP contained high titer (> or = 1:200) antibodies that reacted with immobilized complexes of heparin and platelet factor 4 (PF4), a heparin-binding protein contained in platelet alpha-granules. Recombinant human PF4 behaved similarly to PF4 isolated from platelets in this assay system. Complexes formed at an apparent heparin/PF4 molecular ratio of approximately 1:2 (fresh heparin) and approximately 1:12 (outdated heparin) were most effective in binding antibody. Immune complexes consisting of PF4, heparin, and antibody reacted with resting platelets; this interaction was inhibited by a monoclonal antibody specific for the Fc gamma RII receptor and by excess heparin. Human umbilical vein endothelial cells, known to express heparin-like glycosaminoglycan molecules on their surface, were recognized by antibody in the presence of PF4 alone; this reaction was inhibited by excess heparin, but not by anti-Fc gamma RII. Antibodies reactive with heparin/PF4 were not found in normal plasma, but IgG and IgM antibodies were detected at dilutions of 1:10 (IgG) and 1:50 (IgM) in 3 of 50 patients (6%) with other types of immune thrombocytopenia. These findings indicate that antibodies associated with HITP react with PF4 complexed with heparin in solution or with glycosaminoglycan molecules on the surface of endothelial cells and provide the basis for a new hypothesis to explain the development of thrombocytopenia with thrombosis or disseminated intravascular coagulation in patients sensitive to heparin. PMID:8282825

  10. Potential Prepore Trimer Formation by the Bacillus thuringiensis Mosquito-specific Toxin: MOLECULAR INSIGHTS INTO A CRITICAL PREREQUISITE OF MEMBRANE-BOUND MONOMERS.

    PubMed

    Sriwimol, Wilaiwan; Aroonkesorn, Aratee; Sakdee, Somsri; Kanchanawarin, Chalermpol; Uchihashi, Takayuki; Ando, Toshio; Angsuthanasombat, Chanan

    2015-08-21

    The insecticidal feature of the three-domain Cry δ-endotoxins from Bacillus thuringiensis is generally attributed to their capability to form oligomeric pores, causing lysis of target larval midgut cells. However, the molecular description of their oligomerization process has not been clearly defined. Here a stable prepore of the 65-kDa trypsin-activated Cry4Ba mosquito-specific toxin was established through membrane-mimetic environments by forming an ∼200-kDa octyl-β-D-glucoside micelle-induced trimer. The SDS-resistant trimer caused cytolysis to Sf9 insect cells expressing Aedes-mALP (a Cry4Ba receptor) and was more effective than a toxin monomer in membrane perturbation of calcein-loaded liposomes. A three-dimensional model of toxin trimer obtained by negative-stain EM in combination with single-particle reconstruction at ∼5 nm resolution showed a propeller-shaped structure with 3-fold symmetry. Fitting the three-dimensional reconstructed EM map with a 100-ns molecular dynamics-simulated Cry4Ba structure interacting with an octyl-β-D-glucoside micelle showed relative positioning of individual domains in the context of the trimeric complex with a major protrusion from the pore-forming domain. Moreover, high-speed atomic force microscopy imaging at nanometer resolution and a subsecond frame rate demonstrated conformational transitions from a propeller-like to a globularly shaped trimer upon lipid membrane interactions, implying prepore-to-pore conversion. Real-time trimeric arrangement of monomers associated with L-α-dimyristoylphosphatidylcholine/3-[(3-cholamidopropyl)dimethylammonio]-2-hydroxy-1-propanesulfonic acid bicelle membranes was also envisaged by successive high-speed atomic force microscopy imaging, depicting interactions among three individual subunits toward trimer formation. Together, our data provide the first pivotal insights into the structural requirement of membrane-induced conformational changes of Cry4Ba toxin monomers for the

  11. Bound states and the Bekenstein bound

    SciTech Connect

    Bousso, Raphael

    2003-10-16

    We explore the validity of the generalized Bekenstein bound, S<= pi M a. We define the entropy S as the logarithm of the number of states which have energy eigenvalue below M and are localized to a flat space region of width alpha. If boundary conditions that localize field modes are imposed by fiat, then the bound encounters well-known difficulties with negative Casimir energy and large species number, as well as novel problems arising only in the generalized form. In realistic systems, however, finite-size effects contribute additional energy. We study two different models for estimating such contributions. Our analysis suggests that the bound is both valid and nontrivial if interactions are properly included, so that the entropy S counts the bound states of interacting fields.

  12. Osteopontin promotes epithelial-mesenchymal transition of hepatocellular carcinoma through regulating vimentin

    PubMed Central

    Zhang, Xiaofei; Gao, Xiaomei; Wei, Jinwang; Sheng, Yuanyuan; Zheng, Yan; Yu, Jian; Xie, Lu; Qin, Yi; Qiao, Peng; Zhou, Chuang; Yu, Xinxin; Jia, Huliang; Ren, Ning; Zhou, Haijun; Ye, Qinghai; Qin, Lunxiu

    2016-01-01

    Our previous studies have found that osteopontin (OPN) is a promoter for hepatocellular carcinoma (HCC) progression. However, the molecular mechanism by which OPN enhances HCC metastasis remains elusive. Epithelial-mesenchymal transition (EMT) of cancer cells plays a pivotal role in promoting metastatic process. In this study, we demonstrated that OPN promotes HCC metastasis by inducing an EMT-like, more aggressive cellular phenotype in vitro and in vivo. Furthermore, OPN was identified to interact with vimentin by reciprocal OPN and vimentin immunoprecipitation as well as co-immunofluorescence examination. By using deletion mutants, we found that the residues between 246 and 406 in vimentin are required for binding to OPN. Importantly, OPN significantly increased vimentin stability through inhibition of its protein degradation. Knockdown of vimentin neutralized the EMT induced by OPN both in vitro and in vivo. Moreover, a significant correlation between OPN and vimentin levels was found in clinical HCC specimens and their combination had a worse prognosis with shorter overall survival (OS) and time to recurrence (TTR). In multivariate analysis, OPN and their combination were demonstrated to be independent prognostic indicators for OS and TTR of HCC patients. Collectively, this study indicates that OPN can induce EMT of HCC cells through increasing vimentin stability, which provides more in-depth understanding about the molecular mechanisms of OPN in promoting HCC metastasis and opens tantalizing therapeutic possibilities in HCC. PMID:26824421

  13. Osteopontin splice variants expression is involved on docetaxel resistance in PC3 prostate cancer cells.

    PubMed

    Nakamura, K D M; Tilli, T M; Wanderley, J L; Palumbo, A; Mattos, R M; Ferreira, A C; Klumb, C E; Nasciutti, L E; Gimba, E R

    2016-02-01

    Osteopontin (OPN) is a phosphoprotein that activates several aspects of tumor progression. Alternative splicing of the OPN primary transcript generates three splicing isoforms, OPNa, OPNb and OPNc. In this report, we investigated some cellular mechanisms by which OPN splice variants could mediate PC3 prostate cancer (PCa) cell survival and growth in response to docetaxel (DXT)-induced cell death. Cell survival before and after DXT treatment was analyzed by phase-contrast microscopy and crystal-violet staining assays. Quantitative real-time PCR and immunocytochemical staining assays were used to evaluate the putative involvement of epithelial-mesenchymal transition (EMT) and OPN isoforms on mediating PC3 cell survival. Upon DXT treatment, PC3 cells overexpressing OPNb or OPNc isoforms showed higher cell densities, compared to cells overexpressing OPNa and controls. Notably, cells overexpressing OPNb or OPNc isoforms showed a downregulated pattern of EMT epithelial cell markers, while mesenchymal markers were mostly upregulated in these experimental conditions. We concluded that OPNc or OPNb overexpression in PC3 cells can mediate resistance and cell survival features in response to DXT-induced cell death. Our data also provide evidence the EMT program could be one of the molecular mechanisms mediating survival in OPNb- or OPNc-overexpressing cells in response to DXT treatment. These data could further contribute to a better understanding of the mechanisms by which PCa cells acquire resistance to DXT treatment. PMID:26404131

  14. SOX9 is a novel cancer stem cell marker surrogated by osteopontin in human hepatocellular carcinoma

    PubMed Central

    Kawai, Takayuki; Yasuchika, Kentaro; Ishii, Takamichi; Miyauchi, Yuya; Kojima, Hidenobu; Yamaoka, Ryoya; Katayama, Hokahiro; Yoshitoshi, Elena Yukie; Ogiso, Satoshi; Kita, Sadahiko; Yasuda, Katsutaro; Fukumitsu, Ken; Komori, Junji; Hatano, Etsuro; Kawaguchi, Yoshiya; Uemoto, Shinji

    2016-01-01

    The current lack of cancer stem cell (CSC) markers that are easily evaluated by blood samples prevents the establishment of new therapeutic strategies in hepatocellular carcinoma (HCC). Herein, we examined whether sex determining region Y-box 9 (SOX9) represents a new CSC marker, and whether osteopontin (OPN) can be used as a surrogate marker of SOX9 in HCC. In HCC cell lines transfected with a SOX9 promoter-driven enhanced green fluorescence protein gene, FACS-isolated SOX9+ cells were capable of self-renewal and differentiation into SOX9− cells, and displayed high proliferation capacity in vitro. Xenotransplantation experiments revealed that SOX9+ cells reproduced, differentiated into SOX9− cells, and generated tumors at a high frequency in vivo. Moreover, SOX9+ cells were found to be involved in epithelial-mesenchymal transition (EMT) and activation of TGFb/Smad signaling. Gain/loss of function experiments showed that SOX9 regulates Wnt/beta-catenin signaling, including cyclin D1 and OPN. Immunohistochemistry of 166 HCC surgical specimens and serum OPN measurements showed that compared to SOX9− patients, SOX9+ patients had significantly poorer recurrence-free survival, stronger venous invasion, and higher serum OPN levels. In conclusion, SOX9 is a novel HCC-CSC marker regulating the Wnt/beta-catenin pathway and its downstream target, OPN. OPN is a useful surrogate marker of SOX9 in HCC. PMID:27457505

  15. Dynamic Light Scattering Analysis of the Effect of Phosphorylated Osteopontin Peptides on Mineral Formation

    NASA Astrophysics Data System (ADS)

    Mozaffari, Maryam; Goiko, Maria; de Bruyn, John; Goldberg, Harvey

    2015-03-01

    Biomineralization is the process by which living organisms synthesize minerals. Osteopontin (OPN), a mineral-associated protein, has been shown to be a potent inhibitor of mineral formation, a process that is dependent on phosphorylation. To gain a better understanding of the mechanism of inhibition, dynamic light scattering (DLS) was used to monitor the initial stages of nucleation, providing information about the size and relative concentration of the growing crystals as a function of time. DLS was used to investigate the effect of phosphorylated (P3, pOPAR) and non-phosphorylated (P0, OPAR) OPN peptides on the formation and growth of hydroxyapatite (HA) crystals from supersaturated solutions of calcium and phosphate ions. The non-phosphorylated P0 had a limited effect on HA nucleation and growth, while its thrice-phosphorylated isoform, P3, was a potent inhibitor of HA nucleation. The aspartic acid-rich OPAR was found to moderately inhibit nucleation but not growth, while its singly-phosphorylated isoform, pOPAR, inhibited HA nucleation more effectively, with some effect on HA crystal growth. The order of the inhibitory potential of these peptides was pOPAR>OPAR>P3>P0. This work confirms that highly acidic and phosphorylated peptides can inhibit the nucleation of HA more effectively.

  16. Progressive Secondary Neurodegeneration and Microcalcification Co-Occur in Osteopontin-Deficient Mice

    PubMed Central

    Maetzler, Walter; Berg, Daniela; Funke, Claudia; Sandmann, Freya; Stünitz, Holger; Maetzler, Corina; Nitsch, Cordula

    2010-01-01

    In the brain, osteopontin (OPN) may function in a variety of pathological conditions, including neurodegeneration, microcalcification, and inflammation. In this study, we addressed the role of OPN in primary and secondary neurodegeneration, microcalcification, and inflammation after an excitotoxic lesion by examining OPN knock-out (KO) mice. Two, four, and ten weeks after injection of the glutamate analogue ibotenate into the corticostriatal boundary, the brains of 12 mice per survival time and strain were evaluated. OPN was detectable in neuron-shaped cells, in microglia, and at the surface of dense calcium deposits. At this primary lesion site, although the glial reaction was attenuated in OPN-KO mice, lesion size and presence of microcalcification were comparable between OPN-KO and wild-type mice. In contrast, secondary neurodegeneration at the thalamus was more prominent in OPN-KO mice, and this difference increased over time. This was paralleled by a dramatic rise in the regional extent of dense microcalcification. Despite these differences, the numbers of glial cells did not significantly differ between the two strains. This study demonstrates for the first time a genetic model with co-occurrence of neurodegeneration and microcalcification, mediated by the lack of OPN, and suggests a basic involvement of OPN action in these conditions. In the case of secondary retrograde or transneuronal degeneration, OPN may have a protective role as intracellular actor. PMID:20522649

  17. RANKL, Osteopontin, and Osteoclast Homeostasis in a Hyper-Occlusion Mouse Model

    SciTech Connect

    Walker, Cameron G.; Ito, Yoshihiro; Dangaria, Smit; Luan, Xianghong; Diekwisch, Thomas G.H.

    2010-11-15

    The biological mechanisms that maintain the position of teeth in their sockets establish a dynamic equilibrium between bone resorption and apposition. In order to reveal some of the dynamics involved in the tissue responses towards occlusal forces on periodontal ligament (PDL) and alveolar bone homeostasis, we developed the first mouse model of hyperocclusion. Swiss-Webster mice were kept in hyperocclusion for 0, 3, 6, and 9 d. Morphological and histological changes in the periodontium were assessed using micro-computed tomography (micro-CT) and ground sections with fluorescent detection of vital dye labels. Sections were stained for tartrate-resistant acid phosphatase, and the expression of receptor activator of nuclear factor-{kappa}B ligand (RANKL) and osteopontin (OPN) was analyzed by immunohistochemistry and real-time polymerase chain reaction (PCR). Traumatic occlusion resulted in enamel surface abrasion, inhibition of alveolar bone apposition, significant formation of osteoclasts at 3, 6 and 9 d, and upregulation of OPN and RANKL. Data from this study suggest that both OPN and RANKL contribute to the stimulation of bone resorption in the hyperocclusive state. In addition, we propose that the inhibition of alveolar bone apposition by occlusal forces is an important mechanism for the control of occlusal height that might work in synergy with RANKL-induced bone resorption to maintain normal occlusion.

  18. Identification of Transglutaminase Reactive Residues in Human Osteopontin and Their Role in Polymerization

    PubMed Central

    Christensen, Brian; Zachariae, Elias D.; Scavenius, Carsten; Thybo, Morten; Callesen, Morten M.; Kløverpris, Søren; Oxvig, Claus; Enghild, Jan J.; Sørensen, Esben S.

    2014-01-01

    Osteopontin (OPN) is a highly posttranslationally modified protein present in several tissues where it is implicated in numerous physiological processes. OPN primarily exerts its functions through interaction with integrins via the Arg-Gly-Asp and Ser-Val-Val-Tyr-Gly-Leu-Arg sequences located in the N-terminal part of the protein. OPN can be polymerized by the cross-linking enzyme transglutaminase 2 (TG2), and polymerization has been shown to enhance the biological activity of OPN. However, little is known about the reactivity and location of the glutamine and lysine residues involved in the TG2-mediated modification of OPN. Here we show that TG2 catalyses the incorporation of 5-(Biotinamido)pentylamine at glutamines in both the N- and C-terminal parts of OPN, whereas TG2 primarily incorporated the glutamine-donor peptide biotinyl-TVQQEL-OH into the C-terminal part of OPN. By mass spectrometric analyses we identified Gln34, Gln42, Gln193 and Gln248 as the major TG2 reactive glutamines in OPN. The distribution of reactive Gln and Lys residues in OPN proved to be important, as the full-length protein but not the physiologically highly active integrin-binding N-terminal part of OPN were able to polymerize in a TG2-mediated reaction. Collectively, these data provide important new molecular knowledge about the mechanism of OPN polymerization. PMID:25419572

  19. Osteopontin Promoter Polymorphism is Associated with Increased Carotid Intima-Media Thickness

    PubMed Central

    de las Fuentes, Lisa; Gu, C. Charles; Mathews, Santhosh J.; Reagan, Joann L.; Ruthmann, Nicholas P.; Waggoner, Alan D.; Lai, Chung-Fang; Towler, Dwight A.; Dávila-Román, Víctor G.

    2008-01-01

    Background Osteopontin (OPN)-transgenic mice exhibit increased carotid artery intima-media thickness (CIMT), smooth muscle cell proliferation, and atheroma formation. Methods An association of the human T-66G promoter variant with CIMT was examined in Caucasian adults grouped according to metabolic syndrome (MetS) criteria: Present (+MetS, n=70) and Absent (−MetS, n=70). Results The G allele frequency was 22%. For the entire cohort, the G group (TG and GG) was associated with significantly lower age/gender-adjusted CIMT compared to the TT group (p=0.008); similar analysis by MetS group found a significant difference only in the −MetS group (p=0.018). Stepwise multivariable regression showed that after age and waist circumference, the T-66G variant was predictive of CIMT (p=0.007). These data suggest that in a normoglycemic environment, human vascular OPN gene expression contributes to arterial structure, an effect diminished in dysmetabolic states. Conclusions Humans with the OPN −66 TT genotype, particularly those without MetS, exhibit thicker CIMT. PMID:18406574

  20. Serum Osteopontin as a Novel Biomarker for Muscle Regeneration in Duchenne Muscular Dystrophy.

    PubMed

    Kuraoka, Mutsuki; Kimura, En; Nagata, Tetsuya; Okada, Takashi; Aoki, Yoshitsugu; Tachimori, Hisateru; Yonemoto, Naohiro; Imamura, Michihiro; Takeda, Shin'ichi

    2016-05-01

    Duchenne muscular dystrophy is a lethal X-linked muscle disorder. We have already reported that osteopontin (OPN), an inflammatory cytokine and myogenic factor, is expressed in the early dystrophic phase in canine X-linked muscular dystrophy in Japan, a dystrophic dog model. To further explore the possibility of OPN as a new biomarker for disease activity in Duchenne muscular dystrophy, we monitored serum OPN levels in dystrophic and wild-type dogs at different ages and compared the levels to other serum markers, such as serum creatine kinase, matrix metalloproteinase-9, and tissue inhibitor of metalloproteinase-1. Serum OPN levels in the dystrophic dogs were significantly elevated compared with those in wild-type dogs before and 1 hour after a cesarean section birth and at the age of 3 months. The serum OPN level was significantly correlated with the phenotypic severity of dystrophic dogs at the period corresponding to the onset of muscle weakness, whereas other serum markers including creatine kinase were not. Immunohistologically, OPN was up-regulated in infiltrating macrophages and developmental myosin heavy chain-positive regenerating muscle fibers in the dystrophic dogs, whereas serum OPN was highly elevated. OPN expression was also observed during the synergic muscle regeneration process induced by cardiotoxin injection. In conclusion, OPN is a promising biomarker for muscle regeneration in dystrophic dogs and can be applicable to boys with Duchenne muscular dystrophy. PMID:26963343

  1. Association Between Osteopontin Gene Polymorphisms and Cerebral Palsy in a Chinese Population.

    PubMed

    Shang, Qing; Zhou, Chongchen; Liu, Dongzhi; Li, Wenxia; Chen, Mingjie; Xu, Yiran; Wang, Fei; Bi, Dan; Zhang, Xiaoli; Zhao, Xinzhi; Wang, Lei; Zhu, Changlian; Xing, Qinghe

    2016-06-01

    Cerebral palsy (CP) is a neurological disorder affecting movement and posture that develops as a complication of prenatal, perinatal, and postnatal brain injury. Such non-progressive brain injury is often accompanied by neonatal encephalopathy and inflammation. The widely expressed soluble cytokine osteopontin (OPN) plays an important role in inflammation and neurological protection. Therefore, it is of great interest to study the relationship between CP and genetic variants of OPN. To explore the genetic association between OPN gene single nucleotide polymorphisms (SNPs) and CP in the Chinese Han population, five SNPs (rs2853744, rs2853749, rs11728697, rs4754, and rs1126616) were genotyped among 715 CP patients and 658 healthy controls using the MassArray platform. Statistical analysis was performed using the online SHEsis program, and Bonferroni correction was applied as necessary. We found an association between rs1126616 and global CP (corrected allelic P = 0.0006 and genotypic P = 0.0011 after Bonferroni correction). The other SNPs were not statistically associated with CP or any of its subgroups. By testing a relatively large sample size, our study demonstrates that the OPN gene SNP rs1126616 is statistically associated with CP. We suspect that the OPN gene might be a susceptibility factor for CP. PMID:27114095

  2. Osteopontin deficiency reduces kidney damage from hypercholesterolemia in Apolipoprotein E-deficient mice

    PubMed Central

    Pei, Zouwei; Okura, Takafumi; Nagao, Tomoaki; Enomoto, Daijiro; Kukida, Masayoshi; Tanino, Akiko; Miyoshi, Ken-ichi; Kurata, Mie; Higaki, Jitsuo

    2016-01-01

    Hypercholesterolemia is a well-established risk factor for kidney injury, which can lead to chronic kidney disease (CKD). Osteopontin (OPN) has been implicated in the pathology of several renal conditions. This study was to evaluate the effects of OPN on hypercholesterolemia induced renal dysfunction. Eight-week-old male mice were divided into 4 groups: apolipoprotein E knockout (ApoE−/−) and ApoE/OPN knockout (ApoE−/−/OPN−/−) mice fed a normal diet (ND) or high cholesterol diet (HD). After 4 weeks, Periodic acid-Schiff (PAS) and oil red O staining revealed excessive lipid deposition in the glomeruli of ApoE−/−HD mice, however, significantly suppressed in ApoE−/−/OPN−/−HD mice. Lectin-like oxidized low-density lipoprotein receptor-1 (LOX-1) expression was lower in the glomeruli of ApoE−/−/OPN−/−HD mice than ApoE−/−HD mice. In vitro study, primary mesangial cells were incubated with recombinant mouse OPN (rmOPN). RmOPN induced LOX-1 mRNA and protein expression in primary mesangial cells. Pre-treatment with an ERK inhibitor suppressed the LOX-1 gene expression induced by rmOPN. These results indicate that OPN contributes to kidney damage in hypercholesterolemia and suggest that inhibition of OPN may provide a potential therapeutic target for the prevention of hypercholesterolemia. PMID:27353458

  3. Osteopontin deficiency reduces kidney damage from hypercholesterolemia in Apolipoprotein E-deficient mice.

    PubMed

    Pei, Zouwei; Okura, Takafumi; Nagao, Tomoaki; Enomoto, Daijiro; Kukida, Masayoshi; Tanino, Akiko; Miyoshi, Ken-Ichi; Kurata, Mie; Higaki, Jitsuo

    2016-01-01

    Hypercholesterolemia is a well-established risk factor for kidney injury, which can lead to chronic kidney disease (CKD). Osteopontin (OPN) has been implicated in the pathology of several renal conditions. This study was to evaluate the effects of OPN on hypercholesterolemia induced renal dysfunction. Eight-week-old male mice were divided into 4 groups: apolipoprotein E knockout (ApoE(-/-)) and ApoE/OPN knockout (ApoE(-/-)/OPN(-/-)) mice fed a normal diet (ND) or high cholesterol diet (HD). After 4 weeks, Periodic acid-Schiff (PAS) and oil red O staining revealed excessive lipid deposition in the glomeruli of ApoE(-/-)HD mice, however, significantly suppressed in ApoE(-/-)/OPN(-/-)HD mice. Lectin-like oxidized low-density lipoprotein receptor-1 (LOX-1) expression was lower in the glomeruli of ApoE(-/-)/OPN(-/-)HD mice than ApoE(-/-)HD mice. In vitro study, primary mesangial cells were incubated with recombinant mouse OPN (rmOPN). RmOPN induced LOX-1 mRNA and protein expression in primary mesangial cells. Pre-treatment with an ERK inhibitor suppressed the LOX-1 gene expression induced by rmOPN. These results indicate that OPN contributes to kidney damage in hypercholesterolemia and suggest that inhibition of OPN may provide a potential therapeutic target for the prevention of hypercholesterolemia. PMID:27353458

  4. Osteopontin ablation ameliorates muscular dystrophy by shifting macrophages to a pro-regenerative phenotype.

    PubMed

    Capote, Joana; Kramerova, Irina; Martinez, Leonel; Vetrone, Sylvia; Barton, Elisabeth R; Sweeney, H Lee; Miceli, M Carrie; Spencer, Melissa J

    2016-04-25

    In the degenerative disease Duchenne muscular dystrophy, inflammatory cells enter muscles in response to repetitive muscle damage. Immune factors are required for muscle regeneration, but chronic inflammation creates a profibrotic milieu that exacerbates disease progression. Osteopontin (OPN) is an immunomodulator highly expressed in dystrophic muscles. Ablation of OPN correlates with reduced fibrosis and improved muscle strength as well as reduced natural killer T (NKT) cell counts. Here, we demonstrate that the improved dystrophic phenotype observed with OPN ablation does not result from reductions in NKT cells. OPN ablation skews macrophage polarization toward a pro-regenerative phenotype by reducing M1 and M2a and increasing M2c subsets. These changes are associated with increased expression of pro-regenerative factors insulin-like growth factor 1, leukemia inhibitory factor, and urokinase-type plasminogen activator. Furthermore, altered macrophage polarization correlated with increases in muscle weight and muscle fiber diameter, resulting in long-term improvements in muscle strength and function in mdx mice. These findings suggest that OPN ablation promotes muscle repair via macrophage secretion of pro-myogenic growth factors. PMID:27091452

  5. Correlation between Serum Osteopontin and miR-181a Levels in Allergic Rhinitis Children

    PubMed Central

    Liu, Wenlong; Zeng, Qingxiang; Luo, Renzhong

    2016-01-01

    Background. Osteopontin (OPN) has been proved to be associated with allergic airway inflammation. However, the roles of OPN and its regulation in childhood allergic rhinitis (AR) are poorly understood. Objective. This study aims to evaluate the expression of OPN and miR-181a in children with AR and their association with Th1/Th2 immune response. Methods. Children who suffered from AR were included along with control subjects. Serum was collected to examine the level of OPN and Th1/Th2 cytokines by enzyme-linked immunosorbent assay (ELISA) and the level of miR-181a by quantitative polymerase chain reaction (qPCR). Results. Children with AR had significantly higher serum levels of OPN and lower serum levels of miR-181a than healthy controls. Furthermore, serum levels of OPN were positively correlated with Th2 cytokine and negatively correlated with Th1 cytokine. On the contrary, miR-181a level had a negative correlation with IL-4/IL-5 and positive correlation with IFN-γ/IL-12. More importantly, there was also significant negative correlation between OPN and miR-181a. Conclusion. The OPN protein and miR-181a levels may serve as predictors of disease severity in childhood AR and appear to be promising targets for modulating AR. PMID:27199509

  6. Osteopontin deficiency does not prevent but promotes alcoholic neutrophilic hepatitis in mice

    PubMed Central

    Lazaro, Raul; Wu, Raymond; Lee, Sunyoung; Zhu, Nian-Ling; Chen, Chia-Lin; French, Samuel W.; Xu, Jun; Machida, Keigo; Tsukamoto, Hidekazu

    2014-01-01

    Alcoholic hepatitis (AH) is a distinct spectrum of alcoholic liver disease (ALD) with intense neutrophilic (PMN) inflammation and high mortality. Although a recent study implicates osteopontin (SPP1) in AH, SPP1 is also shown to have protective effects on experimental ALD. To address this unsettled question, we examined the effects of SPP1 deficiency in male mice given 40% calories derived from ad libitum consumption of the Western diet high in cholesterol and saturated fat (HCFD) and the rest from intragastric feeding (iG) of alcohol diet without or with weekly alcohol binge. Weekly binge in this new hybrid feeding model shifts chronic ASH with macrophage inflammation and perisinusoidal and pericelluar fibrosis to AH in 57% (15/26) of the mice, accompanied by inductions of chemokines (Spp1, Cxcl1, Il-17a), progenitor genes (Cd133, Cd24, Nanog, Epcam), PMN infiltration, and clinical features of AH such as hypoalbuminemia, bilirubinemia, and splenomegaly. SPP1 deficiency does not reduce the AH incidence and inductions of progenitor and fibrogenic genes but rather enhances the Il-17a induction and PMN infiltration in some mice. Further, in the absence of SPP1, chronic ASH mice without weekly binge begin to develop AH. In conclusion, these results suggest SPP1 has a protective rather than causal role for experimental AH reproduced in our model. PMID:25132354

  7. Intracellular Osteopontin inhibits toll-like receptor signaling and impedes liver carcinogenesis.

    PubMed

    Fan, Xiaoyu; He, Chunyan; Jing, Wei; Zhou, Xuyu; Chen, Rui; Cao, Lei; Zhu, Minhui; Jia, Rongjie; Wang, Hao; Guo, Yajun; Zhao, Jian

    2015-01-01

    Osteopontin (OPN) has been implicated widely in tumor growth and metastasis, but the range of its contributions is not yet fully understood. In this study, we show that genetic ablation of Opn in mice sensitizes them to diethylnitrosamine (DEN)-induced hepatocarcinogenesis. Opn-deficient mice (Opn(-/-) mice) exhibited enhanced production of proinflammatory cytokines and compensatory proliferation. Administering OPN antibody or recombinant OPN protein to wild-type or Opn(-/-) mice-derived macrophages, respectively, had little effect on cytokine production. In contrast, overexpression of intracellular OPN (iOPN) in Opn-deficient macrophages strongly suppressed production of proinflammatory cytokines. In addition, we found that iOPN was able to interact with the pivotal Toll-like receptor (TLR) signaling protein MyD88 in macrophages after stimulation with cellular debris, thereby disrupting TLR signaling in macrophages. Our results indicated that iOPN was capable of functioning as an endogenous negative regulator of TLR-mediated immune responses, acting to ameliorate production of proinflammatory cytokines and curtail DEN-induced hepatocarcinogenesis. Together, our results expand the important role of OPN in inflammation-associated cancers and deepen its relevance for novel treatment strategies in liver cancer. PMID:25398438

  8. Osteopontin is an alpha motor neuron marker in the mouse spinal cord.

    PubMed

    Misawa, Hidemi; Hara, Mayumi; Tanabe, Shogo; Niikura, Mamiko; Moriwaki, Yasuhiro; Okuda, Takashi

    2012-04-01

    Motor neurons (MNs) are designated as alpha/gamma and fast/slow based on their target sites and the types of muscle fibers innervated; however, few molecular markers that distinguish between these subtypes are available. Here we report that osteopontin (OPN) is a selective marker of alpha MNs in the mouse spinal cord. OPN was detected in approximately 70% of postnatal choline acetyltransferase (ChAT)-positive MNs with relatively large somas, but not in those with smaller somas. OPN+/ChAT+ MNs were also positive for NeuN, an alpha MN marker, but were negative for Err3, a gamma MN marker. The size distribution of OPN+/ChAT+ cells was nearly identical to that of NeuN+/ChAT+ alpha MNs. Group Ia proprioceptive terminals immunoreactive for vesicular glutamate transporter-1 were selectively detected on the OPN+/ChAT+ cells. OPN staining was also detected at motor axon terminals at neuromuscular junctions, where the OPN+ terminals were positive or negative for SV2A, a marker distinguishing fast/slow motor endplates. Finally, retrograde labeling following intramuscular injection of fast blue indicated that OPN is expressed in both fast and slow MNs. Collectively, our findings show that OPN is an alpha MN marker present in both the soma and the endplates of alpha MNs in the postnatal mouse spinal cord. PMID:22420030

  9. Abnormal osteopontin and matrix extracellular phosphoglycoprotein localization, and odontoblast differentiation, in X-linked hypophosphatemic teeth.

    PubMed

    Salmon, B; Bardet, C; Coyac, B R; Baroukh, B; Naji, J; Rowe, P S; Opsahl Vital, S; Linglart, A; Mckee, M D; Chaussain, C

    2014-08-01

    Mutations in phosphate-regulating gene (PHEX) lead to X-linked hypophosphatemic rickets (XLH), a genetic disease characterized by impaired mineralization in bones and teeth. In human XLH tooth dentin, calcospherites that would normally merge as part of the mineralization process are separated by unmineralized interglobular spaces where fragments of matrix proteins accumulate. Here, we immunolocalized osteopontin (OPN) in human XLH teeth, in a three-dimensional XLH human dental pulp stem cell-collagen scaffold culture model and in a rat tooth injury repair model treated with acidic serine- and aspartate-rich motif peptides (ASARM). In parallel, matrix extracellular phosphoglycoprotein (MEPE) immunolocalization and alkaline phosphatase (ALP) activity were assessed in XLH teeth. OPN was expressed by odontoblasts in the XLH models, and localized to the abnormal calcospherites of XLH tooth dentin. In addition, ALP activity and MEPE localization were abnormal in human XLH teeth, with MEPE showing an accumulation in the unmineralized interglobular spaces in dentin. Furthermore, XLH odontoblasts failed to form a well-polarized odontoblast layer. These data suggest that both MEPE and OPN are involved in impaired tooth mineralization associated with XLH, possibly through different effects on the mineralization process. PMID:25158186

  10. Prognostic Value of Osteopontin Splice Variant-c Expression in Breast Cancers: A Meta-Analysis

    PubMed Central

    Hao, Chengcheng; Wang, Zhiyan; Gu, Yanan; Jiang, Wen G.

    2016-01-01

    Objectives. Osteopontin (OPN) is overexpressed in breast cancers, while its clinical and prognostic significance remained unclear. This study aimed to assess the prognostic value of OPN, especially its splice variants, in breast cancers. Methods. Data were extracted from eligible studies concerning the OPN and OPN-c expression in breast cancer patients and were used to calculate the association between OPN/OPN-c and survival. Two reviewer teams independently screened the literatures according to the inclusion and exclusion criteria based on quality evaluation. Following the processes of data extraction, assessment, and transformation, meta-analysis was carried out via RevMan 5.3 software. Results. A total of ten studies involving 1,567 patients were included. The results demonstrated that high level OPN indicated a poor outcome in the OS (HR = 2.22, 95% CI: 1.23–4.00, and P = 0.008; random-effects model) with heterogeneity (I2 = 62%) of breast cancer patients. High level OPN-c appeared to be more significantly associated with poor survival (HR = 2.14, 95% CI: 1.51–3.04, and P < 0.0001; fixed-effects model) with undetected heterogeneity (I2 = 0%). Conclusions. Our analyses indicated that both OPN and OPN-c could be considered as prognostic markers for breast cancers. The high level of OPN-c was suggested to be more reliably associated with poor survival in breast cancer patients. PMID:27462610

  11. Resveratrol improves bone repair by modulation of bone morphogenetic proteins and osteopontin gene expression in rats.

    PubMed

    Casarin, R C; Casati, M Z; Pimentel, S P; Cirano, F R; Algayer, M; Pires, P R; Ghiraldini, B; Duarte, P M; Ribeiro, F V

    2014-07-01

    This study investigated the effect of resveratrol on bone healing and its influence on the gene expression of osteogenic markers. Two calvarial defects were created and one screw-shaped titanium implant was inserted in the tibia of rats that were assigned to daily administration of placebo (control group, n=15) or 10mg/kg of resveratrol (RESV group, n=15) for 30 days. The animals were then sacrificed. One of the calvarial defects was processed for histomorphometric analysis and the tissue relative to the other was collected for mRNA quantification of bone morphogenetic protein (BMP)-2, BMP-7, osteopontin (OPN), bone sialoprotein (BSP), osteoprotegrin (OPG), and receptor activator of NF-κB ligand (RANKL). Implants were removed by applying a counter-torque force. Histomorphometric analysis revealed higher remaining defect in the calvarial defects of the control group than the RESV group (P=0.026). Resveratrol increased the counter-torque values of implant removal when compared to control therapy (P=0.031). Gene expression analysis showed a higher expression of BMP-2 (P=0.011), BMP-7 (P=0.049), and OPN (P=0.002) genes in the RESV group than in the control group. In conclusion, resveratrol improved the repair of critical-sized bone defects and the biomechanical retention of implants. Indeed, this natural agent may up-regulate the gene expression of important osteogenic markers. PMID:24530035

  12. The role of osteopontin in D-galactosamine-induced liver injury in genetically obese mice

    SciTech Connect

    Kwon, Hyo-Jung; Won, Young-Suk; Yoon, Won-Kee; Nam, Ki-Hoan; Kim, Dae-Yong; Kim, Hyoung-Chin

    2010-02-01

    Various epidemiological studies have shown that obesity increases the risk of liver disease, but the precise mechanisms through which this occurs are poorly understood. In the present study, we hypothesized that osteopontin (OPN), an extracellular matrix and proinflammatory cytokine, has an important role in making obese mice more susceptible to inflammatory liver injury. After exposure of genetically obese ob/ob and db/db mice to a single dose of D-galactosamine (GalN), the plasma liver enzyme levels, histology and expression levels of cytokines and OPN were evaluated. The ob/ob and db/db mice, which were more sensitive to GalN-induced inflammatory liver injury compared with wild-type mice, had significantly higher plasma and hepatic OPN expression levels. Increased OPN expression was mainly found in hepatocytes and inflammatory cells and was correlated with markedly up-regulated interleukin (IL)-12 and IL-18 levels. Furthermore, pretreatment with a neutralizing OPN (nOPN) antibody attenuated the GalN-induced inflammatory liver injury in ob/ob and db/db mice, which was accompanied by significantly reduced macrophages recruitment and IL-12 and IL-18 productions. Taken together, these results suggest that up-regulated OPN expression is a contributing factor to increased susceptibility of genetically obese mice to GalN-induced liver injury by promoting inflammation and modulating immune response.

  13. A matrix lower bound

    SciTech Connect

    Grcar, Joseph F.

    2002-02-04

    A matrix lower bound is defined that generalizes ideas apparently due to S. Banach and J. von Neumann. The matrix lower bound has a natural interpretation in functional analysis, and it satisfies many of the properties that von Neumann stated for it in a restricted case. Applications for the matrix lower bound are demonstrated in several areas. In linear algebra, the matrix lower bound of a full rank matrix equals the distance to the set of rank-deficient matrices. In numerical analysis, the ratio of the matrix norm to the matrix lower bound is a condition number for all consistent systems of linear equations. In optimization theory, the matrix lower bound suggests an identity for a class of min-max problems. In real analysis, a recursive construction that depends on the matrix lower bound shows that the level sets of continuously differential functions lie asymptotically near those of their tangents.

  14. Increased Expression of Osteopontin in the Degenerating Striatum of Rats Treated with Mitochondrial Toxin 3-Nitropropionic Acid: A Light and Electron Microscopy Study

    PubMed Central

    Kim, Hong-Lim; Lee, Mun-Yong; Shin, Yoo-Jin; Song, Doo-Won; Park, Jieun; Chang, Byung-Soo; Lee, Jong-Hwan

    2015-01-01

    The mycotoxin 3-nitropropionic acid (3NP) is an irreversible inhibitor that induces neuronal damage by inhibiting mitochondrial complex II. Neurodegeneration induced by 3NP, which is preferentially induced in the striatum, is caused by an excess influx and accumulation of calcium in mitochondria. Osteopontin (OPN) is a glycosylated phosphoprotein and plays a role in the regulation of calcium precipitation in the injured brain. The present study was designed to examine whether induction of OPN protein is implicated in the pathogenesis of 3NP-induced striatal neurodegeneration. We observed overlapping regional expression of OPN, the neurodegeneration marker Fluoro-Jade B, and the microglial marker ionized calcium-binding adaptor molecule 1 (Iba1) in the 3NP-lesioned striatum. OPN expression was closely associated with the mitochondrial marker NADH dehydrogenase (ubiquinone) flavoprotein 2 in the damaged striatum. In addition, immunoelectron microscopy demonstrated that OPN protein was specifically localized to the inner membrane and matrix of the mitochondria in degenerating striatal neurons, and cell fragments containing OPN-labeled mitochondria were also present within activated brain macrophages. Thus, our study revealed that OPN expression is associated with mitochondrial dysfunction produced by 3NP-induced alteration of mitochondrial calcium homeostasis, suggesting that OPN is involved in the pathogenesis of striatal degeneration by 3NP administration. PMID:26633905

  15. Increased Expression of Osteopontin in the Degenerating Striatum of Rats Treated with Mitochondrial Toxin 3-Nitropropionic Acid: A Light and Electron Microscopy Study.

    PubMed

    Kim, Hong-Lim; Lee, Mun-Yong; Shin, Yoo-Jin; Song, Doo-Won; Park, Jieun; Chang, Byung-Soo; Lee, Jong-Hwan

    2015-10-29

    The mycotoxin 3-nitropropionic acid (3NP) is an irreversible inhibitor that induces neuronal damage by inhibiting mitochondrial complex II. Neurodegeneration induced by 3NP, which is preferentially induced in the striatum, is caused by an excess influx and accumulation of calcium in mitochondria. Osteopontin (OPN) is a glycosylated phosphoprotein and plays a role in the regulation of calcium precipitation in the injured brain. The present study was designed to examine whether induction of OPN protein is implicated in the pathogenesis of 3NP-induced striatal neurodegeneration. We observed overlapping regional expression of OPN, the neurodegeneration marker Fluoro-Jade B, and the microglial marker ionized calcium-binding adaptor molecule 1 (Iba1) in the 3NP-lesioned striatum. OPN expression was closely associated with the mitochondrial marker NADH dehydrogenase (ubiquinone) flavoprotein 2 in the damaged striatum. In addition, immunoelectron microscopy demonstrated that OPN protein was specifically localized to the inner membrane and matrix of the mitochondria in degenerating striatal neurons, and cell fragments containing OPN-labeled mitochondria were also present within activated brain macrophages. Thus, our study revealed that OPN expression is associated with mitochondrial dysfunction produced by 3NP-induced alteration of mitochondrial calcium homeostasis, suggesting that OPN is involved in the pathogenesis of striatal degeneration by 3NP administration. PMID:26633905

  16. Antisense targeting of TGF-{beta}1 augments BMP-induced upregulation of osteopontin, type I collagen and Cbfa1 in human Saos-2 cells

    SciTech Connect

    Shen, Zhong-Jian . E-mail: zshen2@wisc.edu; Kook Kim, Sang; Youn Jun, Do; Park, Wan; Ho Kim, Young; Malter, James S.; Jo Moon, Byung . E-mail: bjmoon@mail.knu.ac.kr

    2007-04-15

    Despite commonalities in signal transduction in osteoblasts from different species, the role of TGF-{beta}1 on bone formation remains elusive. In particular, the role of autocrine TGF-{beta}1 on human osteoblasts is largely unknown. Here we show the effect of TGF-{beta}1 knock-down on the proliferation and differentiation of osteoblasts induced by BMP2. Treatment with antisense TGF-{beta}1 moderately increased the rate of cell proliferation, which was completely reversed by the exogenous addition of TGF-{beta}1. Notably, TGF-{beta}1 blockade significantly enhanced BMP2-induced upregulation of mRNAs encoding osteopontin, type I collagen and Cbfa1, which was suppressed by exogenous TGF-{beta}1. Moreover, TGF-{beta}1 knock-down increased BMP2-induced phosphorylation of Smad1/5 as well as their nuclear import, which paralleled a reduction of inhibitory Smad6. These data suggest autocrine TGF-{beta}1 antagonizes BMP signaling through modulation of inducible Smad6 and the activity of BMP specific Smad1/5.

  17. Osteopontin, a chemotactic protein with cytokine-like properties, is up-regulated in muscle injury caused by Bothrops lanceolatus (fer-de-lance) snake venom.

    PubMed

    Barbosa-Souza, Valéria; Contin, Daniel Kiss; Filho, Waldemar Bonventi; de Araújo, Albetiza Lôbo; Irazusta, Silvia Pierre; da Cruz-Höfling, Maria Alice

    2011-10-01

    Osteopontin (OPN) is a chemotactic, adhesive protein whose receptors include some integrins and matrix proteins known to have role in inflammatory and repair processes. We examined the time course of OPN expression at acute and chronic stages after intramuscular injection of Bothrops lanceolatus venom in rats. Additionally, we examined the expression of CD68 (a marker for phagocytic macrophages) and the myogenic factors, myoD and myogenin. There was a biphasic upregulation of OPN (6-48 h and 3-14 days post-venom), i.e., during acute inflammation and myogenic cell proliferation and differentiation phases. OPN was detected in CD68 + macrophages, fibroblasts, normal and damaged myofibers, myoblasts and myotubes. Myogenin was expressed in the cytoplasm (atypical pattern) and nucleus of myoblasts and myotubes from 18 h to 7 days, after which it was expressed only in nuclei. Macrophage numbers, OPN and myogenin expression were still elevated at 7, 14 and 7 days. At 3 days, when OPN achieved the peak, some clusters of myoblasts were within regions of intense collagen deposition. Fibrosis may represent limitation for repairing processes and may explain the small diameter of regenerated fibers at 21 days post-venom. The expression of OPN in the course of venom-induced damage and regeneration suggests stages-specific mediation role along the whole process. PMID:21839764

  18. NF-κB and Matrix-Dependent Regulation of Osteopontin Promoter Activity in Allylamine-Activated Vascular Smooth Muscle Cells

    PubMed Central

    Williams, E. Spencer; Wilson, Emily; Ramos, Kenneth S.

    2012-01-01

    Repeated cycles of oxidative injury by allylamine in vivo induce a proliferative rat vascular (aortic) smooth muscle cell (vSMC) phenotype characterized by matrix-dependent enhancement of mitogenic sensitivity, changes in cell surface integrin expression, and osteopontin (opn) overexpression. Here, we show that constitutive and mitogen-stimulated NF-κB DNA binding activity is enhanced in allylamine vSMCs. Matrix-specific changes in cellular Rel protein expression were observed in allylamine vSMCs. The NF-κB DNA binding element located at −1943 in the 5′-UTR strongly inhibited opn promoter activity in allylamine vSMCs, and this response was regulated by the extracellular matrix. Constitutive increases in opn promoter activity were only seen when allylamine cells were seeded on a fibronectin substrate, and this response was independent of the NF-κB DNA binding sequence within the regulatory region. Thus, NF-κB functions as a critical regulator of the allylamine-induced proliferative phenotype in vSMCs. PMID:22315656

  19. Immunomodulation Induced by Stem Cell Mobilization and Harvesting in Healthy Donors: Increased Systemic Osteopontin Levels after Treatment with Granulocyte Colony-Stimulating Factor

    PubMed Central

    Melve, Guro Kristin; Ersvaer, Elisabeth; Akkök, Çiğdem Akalın; Ahmed, Aymen Bushra; Kristoffersen, Einar K.; Hervig, Tor; Bruserud, Øystein

    2016-01-01

    Peripheral blood stem cells from healthy donors mobilized by granulocyte colony-stimulating factor (G-CSF) and harvested by leukapheresis are commonly used for allogeneic stem cell transplantation. The frequency of severe graft versus host disease is similar for patients receiving peripheral blood and bone marrow allografts, even though the blood grafts contain more T cells, indicating mobilization-related immunoregulatory effects. The regulatory phosphoprotein osteopontin was quantified in plasma samples from healthy donors before G-CSF treatment, after four days of treatment immediately before and after leukapheresis, and 18–24 h after apheresis. Myeloma patients received chemotherapy, combined with G-CSF, for stem cell mobilization and plasma samples were prepared immediately before, immediately after, and 18–24 h after leukapheresis. G-CSF treatment of healthy stem cell donors increased plasma osteopontin levels, and a further increase was seen immediately after leukapheresis. The pre-apheresis levels were also increased in myeloma patients compared to healthy individuals. Finally, in vivo G-CSF exposure did not alter T cell expression of osteopontin ligand CD44, and in vitro osteopontin exposure induced only small increases in anti-CD3- and anti-CD28-stimulated T cell proliferation. G-CSF treatment, followed by leukapheresis, can increase systemic osteopontin levels, and this effect may contribute to the immunomodulatory effects of G-CSF treatment. PMID:27447610

  20. Immunomodulation Induced by Stem Cell Mobilization and Harvesting in Healthy Donors: Increased Systemic Osteopontin Levels after Treatment with Granulocyte Colony-Stimulating Factor.

    PubMed

    Melve, Guro Kristin; Ersvaer, Elisabeth; Akkök, Çiğdem Akalın; Ahmed, Aymen Bushra; Kristoffersen, Einar K; Hervig, Tor; Bruserud, Øystein

    2016-01-01

    Peripheral blood stem cells from healthy donors mobilized by granulocyte colony-stimulating factor (G-CSF) and harvested by leukapheresis are commonly used for allogeneic stem cell transplantation. The frequency of severe graft versus host disease is similar for patients receiving peripheral blood and bone marrow allografts, even though the blood grafts contain more T cells, indicating mobilization-related immunoregulatory effects. The regulatory phosphoprotein osteopontin was quantified in plasma samples from healthy donors before G-CSF treatment, after four days of treatment immediately before and after leukapheresis, and 18-24 h after apheresis. Myeloma patients received chemotherapy, combined with G-CSF, for stem cell mobilization and plasma samples were prepared immediately before, immediately after, and 18-24 h after leukapheresis. G-CSF treatment of healthy stem cell donors increased plasma osteopontin levels, and a further increase was seen immediately after leukapheresis. The pre-apheresis levels were also increased in myeloma patients compared to healthy individuals. Finally, in vivo G-CSF exposure did not alter T cell expression of osteopontin ligand CD44, and in vitro osteopontin exposure induced only small increases in anti-CD3- and anti-CD28-stimulated T cell proliferation. G-CSF treatment, followed by leukapheresis, can increase systemic osteopontin levels, and this effect may contribute to the immunomodulatory effects of G-CSF treatment. PMID:27447610

  1. Asymptotic entropy bounds

    NASA Astrophysics Data System (ADS)

    Bousso, Raphael

    2016-07-01

    We show that known entropy bounds constrain the information carried off by radiation to null infinity. We consider distant, planar null hypersurfaces in asymptotically flat spacetime. Their focusing and area loss can be computed perturbatively on a Minkowski background, yielding entropy bounds in terms of the energy flux of the outgoing radiation. In the asymptotic limit, we obtain boundary versions of the quantum null energy condition, of the generalized Second Law, and of the quantum Bousso bound.

  2. Transversely bounded DFB lasers. [bounded distributed-feedback lasers

    NASA Technical Reports Server (NTRS)

    Elachi, C.; Evans, G.; Yeh, C.

    1975-01-01

    Bounded distributed-feedback (DFB) lasers are studied in detail. Threshold gain and field distribution for a number of configurations are derived and analyzed. More specifically, the thin-film guide, fiber, diffusion guide, and hollow channel with inhomogeneous-cladding DFB lasers are considered. Optimum points exist and must be used in DFB laser design. Different-modes feedback and the effects of the transverse boundaries are included. A number of applications are also discussed.

  3. Regulation of osteosarcoma cell invasion through osteopontin modification by miR-4262.

    PubMed

    Song, Kun; Liu, Ning; Yang, Yan; Qiu, Xue

    2016-05-01

    Osteopontin (OPN) is a phosphorylated glycoprotein that plays a critical role in the invasion of osteosarcoma (OS), the most common primary malignant bone tumor. Since microRNAs (miRNAs) have been well documented as key players in the tumorigenesis, cancer cell growth, and metastases, determination of the involved miRNAs that may regulate OPN-mediated OS cell invasion appears to be one important question in the current understanding and therapeutic strategies for OS. Here, we found that the levels of miR-4262 were significantly decreased and the levels of OPN were significantly increased in OS specimens, compared to the paired adjacent non-tumor tissue. Moreover, miR-4262 and OPN inversely correlated in OS specimens. The 5-year survival of the patients with lower miR-4262 levels in the resected OS was worse than that of patients with high miR-4262 levels. Bioinformatics analyses showed that miR-4262 targeted the 3'-UTR of OPN mRNA to inhibit its translation, which was proved by luciferase reporter assay. Furthermore, miR-4262 overexpression inhibited OPN-mediated cell invasion, while miR-4262 depletion increased OPN-mediated cell invasion in OS cells, in both a transwell cell invasion assay and a scratch wound healing assay. Together, our data suggest that suppression of miR-4262 in OS cells may promote OPN-mediated cancer invasion, highlighting miR-4262 as an intriguing therapeutic target to prevent OS metastases. PMID:26634745

  4. Circulating Osteopontin and Prediction of Hepatocellular Carcinoma Development in a Large European Population.

    PubMed

    Duarte-Salles, Talita; Misra, Sandeep; Stepien, Magdalena; Plymoth, Amelie; Muller, David; Overvad, Kim; Olsen, Anja; Tjønneland, Anne; Baglietto, Laura; Severi, Gianluca; Boutron-Ruault, Marie-Christine; Turzanski-Fortner, Renee; Kaaks, Rudolf; Boeing, Heiner; Aleksandrova, Krasimira; Trichopoulou, Antonia; Lagiou, Pagona; Bamia, Christina; Pala, Valeria; Palli, Domenico; Mattiello, Amalia; Tumino, Rosario; Naccarati, Alessio; Bueno-de-Mesquita, H B As; Peeters, Petra H; Weiderpass, Elisabete; Quirós, J Ramón; Agudo, Antonio; Sánchez-Cantalejo, Emilio; Ardanaz, Eva; Gavrila, Diana; Dorronsoro, Miren; Werner, Mårten; Hemmingsson, Oskar; Ohlsson, Bodil; Sjöberg, Klas; Wareham, Nicholas J; Khaw, Kay-Tee; Bradbury, Kathryn E; Gunter, Marc J; Cross, Amanda J; Riboli, Elio; Jenab, Mazda; Hainaut, Pierre; Beretta, Laura

    2016-09-01

    We previously identified osteopontin (OPN) as a promising marker for the early detection of hepatocellular carcinoma (HCC). In this study, we investigated the association between prediagnostic circulating OPN levels and HCC incidence in a large population-based cohort. A nested case-control study was conducted within the European Prospective Investigation into Cancer and Nutrition (EPIC) cohort. During a mean follow-up of 4.8 years, 100 HCC cases were identified. Each case was matched to two controls and OPN levels were measured in baseline plasma samples. Viral hepatitis, liver function, and α-fetoprotein (AFP) tests were also conducted. Conditional logistic regression models were used to calculate multivariable odds ratio (OR) and 95% confidence intervals (95% CI) for OPN levels in relation to HCC. Receiver operating characteristics curves were constructed to determine the discriminatory accuracy of OPN alone or in combination with other liver biomarkers in the prediction of HCC. OPN levels were positively associated with HCC risk (per 10% increment, ORmultivariable = 1.30; 95% CI, 1.14-1.48). The association was stronger among cases diagnosed within 2 years of follow-up. Adding liver function tests to OPN improved the discriminatory performance for subjects who developed HCC (AUC = 0.86). For cases diagnosed within 2 years, the combination of OPN and AFP was best able to predict HCC risk (AUC = 0.88). The best predictive model for HCC in this low-risk population is OPN in combination with liver function tests. Within 2 years of diagnosis, the combination of OPN and AFP best predicted HCC development, suggesting that measuring OPN and AFP could identify high-risk groups independently of a liver disease diagnosis. Cancer Prev Res; 9(9); 758-65. ©2016 AACR. PMID:27339170

  5. Role of plasma osteopontin as a biomarker in locally advanced breast cancer

    PubMed Central

    Anborgh, Pieter H; Caria, Laura BR; Chambers, Ann F; Tuck, Alan B; Stitt, Larry W; Brackstone, Muriel

    2015-01-01

    Osteopontin (OPN), a malignancy-associated secreted phosphoprotein, is a prognostic plasma biomarker for survival in metastatic breast cancer patients. We evaluated the role of OPN in Locally Advanced Breast Cancer (LABC) patients in predicting response to neoadjuvant chemotherapy and association with survival. Fifty-three patients with non-metastatic LABC were enrolled in this study and monitored serially for plasma OPN levels by ELISA during neoadjuvant chemotherapy prior to surgery. For fifty patients who had baseline OPN levels available for analysis, the median baseline OPN level was 63.6 ng/ml. Median patient follow up was 45 months and thirteen patients died from metastatic disease. Patients with baseline OPN levels ≥ 63.6 ng/ml were significantly more likely to die of their disease than those with baseline OPN < 63.6 ng/mL (Hazard Ratio = 3.4; 95% confidence interval 1.4-11.3; P = 0.011), and overall, baseline OPN level was significantly associated with survival (P = 0.002). There was little support for value of serial OPN determination in monitoring response to therapy in this patient population. Although the percentage of patients with baseline OPN levels < 63.6 ng/ml was higher in patients with pathological complete response than in those with no response, the difference was not statistically significant (64% and 14%, respectively (P = 0.066)). Thus, baseline plasma OPN level is a prognostic biomarker in this group of LABC patients, and could also be helpful in identifying LABC patients who will respond to neoadjuvant chemotherapy. Our results call for validation of our findings in large prospective trial data sets. PMID:26064440

  6. Novel Roles for Osteopontin and Clusterin in Peripheral Motor and Sensory Axon Regeneration

    PubMed Central

    Mi, Ruifa; Connor, Emmalynn; Reed, Nicole; Vyas, Alka; Alspalter, Manula; Coppola, Giovanni; Geschwind, Daniel H.; Brushart, Thomas M.

    2014-01-01

    Previous studies demonstrated that Schwann cells (SCs) express distinct motor and sensory phenotypes, which impact the ability of these pathways to selectively support regenerating neurons. In the present study, unbiased microarray analysis was used to examine differential gene expression in denervated motor and sensory pathways in rats. Several genes that were significantly upregulated in either denervated sensory or motor pathways were identified and two secreted factors were selected for further analysis: osteopontin (OPN) and clusterin (CLU) which were upregulated in denervated motor and sensory pathways, respectively. Sciatic nerve transection induced upregulation of OPN and CLU and expression of both returned to baseline levels with ensuing regeneration. In vitro analysis using exogenously applied OPN induced outgrowth of motor but not sensory neurons. CLU, however, induced outgrowth of sensory neurons, but not motor neurons. To assess the functional importance of OPN and CLU, peripheral nerve regeneration was examined in OPN and CLU−/− mice. When compared with OPN+/+ mice, motor neuron regeneration was reduced in OPN−/− mice. Impaired regeneration through OPN−/− peripheral nerves grafted into OPN+/+ mice indicated that loss of OPN in SCs was responsible for reduced motor regeneration. Sensory neuron regeneration was impaired in CLU−/− mice following sciatic nerve crush and impaired regeneration nerve fibers through CLU−/− nerve grafts transplanted into CLU+/+ mice indicated that reduced sensory regeneration is likely due to SC-derived CLU. Together, these studies suggest unique roles for SC-derived OPN and CLU in regeneration of peripheral motor and sensory axons. PMID:24478351

  7. Na+/H+ Exchanger Isoform 1-Induced Osteopontin Expression Facilitates Cardiomyocyte Hypertrophy

    PubMed Central

    Mohamed, Iman A.; Gadeau, Alain-Pierre; Fliegel, Larry; Lopaschuk, Gary; Mlih, Mohamed; Abdulrahman, Nabeel; Fillmore, Natasha; Mraiche, Fatima

    2015-01-01

    Enhanced expression and activity of the Na+/H+ exchanger isoform 1 (NHE1) has been implicated in cardiomyocyte hypertrophy in various experimental models. The upregulation of NHE1 was correlated with an increase in osteopontin (OPN) expression in models of cardiac hypertrophy (CH), and the mechanism for this remains to be delineated. To determine whether the expression of active NHE1-induces OPN and contributes to the hypertrophic response in vitro, cardiomyocytes were infected with the active form of the NHE1 adenovirus or transfected with OPN silencing RNA (siRNA-OPN) and characterized for cardiomyocyte hypertrophy. Expression of NHE1 in cardiomyocytes resulted in a significant increase in cardiomyocyte hypertrophy markers: cell surface area, protein content, ANP mRNA and expression of phosphorylated-GATA4. NHE1 activity was also significantly increased in cardiomyocytes expressing active NHE1. Interestingly, transfection of cardiomyocytes with siRNA-OPN significantly abolished the NHE1-induced cardiomyocyte hypertrophy. siRNA-OPN also significantly reduced the activity of NHE1 in cardiomyocytes expressing NHE1 (68.5±0.24%; P<0.05), confirming the role of OPN in the NHE1-induced hypertrophic response. The hypertrophic response facilitated by NHE1-induced OPN occurred independent of the extracellular-signal-regulated kinases and Akt, but required p90-ribosomal S6 kinase (RSK). The ability of OPN to facilitate the NHE1-induced hypertrophic response identifies OPN as a potential therapeutic target to reverse the hypertrophic effect induced by the expression of active NHE1. PMID:25884410

  8. Targeting osteopontin suppresses glioblastoma stem-like cell character and tumorigenicity in vivo.

    PubMed

    Lamour, Virginie; Henry, Aurélie; Kroonen, Jérôme; Nokin, Marie-Julie; von Marschall, Zofia; Fisher, Larry W; Chau, Tieu-Lan; Chariot, Alain; Sanson, Marc; Delattre, Jean-Yves; Turtoi, Andrei; Peulen, Olivier; Rogister, Bernard; Castronovo, Vincent; Bellahcène, Akeila

    2015-09-01

    Osteopontin (OPN) is a secreted protein involved in most aspects of tumor progression and metastasis development. Elevated OPN expression has been reported in multiple types of cancer including glioblastoma (GBM), the highest grade and most aggressive brain tumor. GBMs contain a subpopulation of glioma-initiating cells (GICs) implicated in progression, therapeutic resistance and recurrence. We have previously demonstrated that OPN silencing inhibited GBM cell growth in vitro and in vivo. Moreover, activation of CD44 signaling upon OPN ligation has been recently implicated in the acquisition of a stem cell phenotype by GBM cells. The present study is aimed to explore OPN autocrine function using shRNA silencing strategy in GICs enriched from GBM cell lines and a human primary GBM grown in EGF and bFGF defined medium. The removal of these growth factors and addition of serum induced a significant loss of OPN expression in GICs. We showed that OPN-silenced GICs were unable to grow as spheres and this capacity was restored by exogenous OPN. Importantly, the expression of Sox2, Oct3/4 and Nanog, key stemness transcription factors, was significantly decreased in GICs upon OPN targeting. We identified Akt/mTOR/p70S6K as the main signaling pathway triggered following OPN-mediated EGFR activation in GICs. Finally, in an orthotopic xenograft mouse model, the tumorigenic potential of U87-MG sphere cells was completely abrogated upon OPN silencing. Our demonstration of endogenous OPN major regulatory effects on GICs stemness phenotype and tumorigenicity implies a greater role than anticipated for OPN in GBM pathogenesis from initiation and progression to probable recurrence. PMID:25620078

  9. Osteopontin in cerebrospinal fluid as diagnostic biomarker for central nervous system lymphoma.

    PubMed

    Strehlow, Felicitas; Bauer, Sandra; Martus, Peter; Weller, Michael; Roth, Patrick; Schlegel, Uwe; Seidel, Sabine; Scheibenbogen, Carmen; Korfel, Agnieszka; Kreher, Stephan

    2016-08-01

    Central nervous system lymphoma (CNSL) is diagnostically challenging. The identification of reliable and easy to measure biomarkers is desirable to facilitate diagnosis. Here, we evaluated the value of cerebrospinal fluid (CSF) osteopontin (OPN) as a diagnostic biomarker for CNSL. OPN concentrations in CSF from 37 patients with CNSL (29 with primary CNSL and 8 with secondary CNS involvement of systemic lymphoma) and 36 controls [6 patients with inflammatory CNS disease other than multiple sclerosis (MS), 8 with MS, 9 with glioblastoma (GBM) and 13 healthy controls] were determined using an enzyme-linked immunosorbent assay. Non-parametric tests and receiver operating characteristic (ROC) curves were performed for determination of diagnostic accuracy. Median CSF OPN level in all CNSL patients was 620 ng/mL and higher than in patients with inflammatory CNS disease (356 ng/mL); P < .05, MS (163 ng/mL); P < .01, GBM (41 ng/mL); P < .01, or healthy controls (319 ng/mL); P < .01. The area under the ROC curve was 0.865 [95 % confidence interval (CI) 0.745-0.985] for differentiating CNSL and patients with inflammatory CNS disease; 0.956 (95 % CI 0.898-1.000) for CNSL and MS patients; 0.988 (95 % CI 0.964-1.000) for CNSL and GBM patients, and 0.915 (95 % CI 0.834-0.996) for CNSL patients and healthy controls. In multivariate analysis, high CSF OPN level was associated with shorter progression-free (HR 1.61, 95 % CI 1.13-2.31; P = .009) and overall survival (HR 1.52, 95 % CI 1.04-2.21; P = .029). CSF OPN is a potential biomarker in CNSL. PMID:27294357

  10. Osteopontin Expression in Acute Immune Response Mediates Hippocampal Synaptogenesis and Adaptive Outcome Following Cortical Brain Injury

    PubMed Central

    Chan, Julie L.; Reeves, Thomas M.; Phillips, Linda L.

    2014-01-01

    Traumatic brain injury (TBI) produces axotomy, deafferentation and reactive synaptogenesis. Inflammation influences synaptic repair, and the novel brain cytokine osteopontin (OPN) has potential to support axon regeneration through exposure of its integrin receptor binding sites. This study explored whether OPN secretion and proteolysis by matrix metalloproteinases (MMPs) mediate the initial degenerative phase of synaptogenesis, targeting reactive neuroglia to affect successful repair. Adult rats received unilateral entorhinal cortex lesion (UEC) modeling adaptive synaptic plasticity. Over the first week postinjury, hippocampal OPN protein and mRNA were assayed and histology performed. At 1–2d, OPN protein increased up to 51 fold, and was localized within activated, mobilized glia. OPN transcript also increased over 50 fold, predominantly within reactive microglia. OPN fragments known to be derived from MMP proteolysis were elevated at 1d, consistent with prior reports of UEC glial activation and enzyme production. Postinjury minocycline immunosuppression attenuated MMP-9 gelatinase activity, which was correlated with reduction of neutrophil gelatinase-associated lipocalin (LCN2) expression, and reduced OPN fragment generation. The antibiotic also attenuated removal of synapsin-1 positive axons from the deafferented zone. OPN KO mice subjected to UEC had similar reduction of hippocampal MMP-9 activity, as well as lower synapsin-1 breakdown over the deafferented zone. MAP1B and N-cadherin, surrogates of cytoarchitecture and synaptic adhesion, were not affected. OPN KO mice with UEC exhibited time dependent cognitive deficits during the synaptogenic phase of recovery. This study demonstrates that OPN can mediate immune response during TBI synaptic repair, positively influencing synapse reorganization and functional recovery. PMID:25151457

  11. Plasma Osteopontin Levels and Adverse Cardiovascular Outcomes in the PEACE Trial

    PubMed Central

    Austin, Erin E.; Gersh, Bernard J.; Solak, Nusret; Rizvi, Syed A.; Bailey, Kent R.; Kullo, Iftikhar J.

    2016-01-01

    Osteopontin (OPN) is a secreted glycophosphoprotein that has a role in inflammation, immune response and calcification. We hypothesized that plasma OPN levels are associated with adverse cardiovascular outcomes in patients with stable coronary artery disease (CAD) and preserved ejection fraction (EF) enrolled in the PEACE trial. We measured plasma OPN levels at baseline in 3567 CAD patients (mean age 64.5 ± 8.1 years, 81% men) by a sandwich chemiluminescent assay (coefficient of variation = 4.1%). OPN levels were natural log (Ln) transformed prior to analyses. We assessed whether Ln OPN levels were associated with the composite primary endpoint of cardiovascular death, non-fatal myocardial infarction and hospitalization for heart failure using multiple event multivariable Cox proportional hazards regression. Adjustment was performed for: (a) age and sex; (b) additional potential confounders; and (c) a parsimonious set of statistically significant 10 variates. During a median follow-up of 4.8 years, 416 adverse cardiovascular outcomes occurred in 366 patients. Ln OPN was significantly associated with the primary endpoint; HR (95% CI) = 1.56 (1.27, 1.92); P <0.001, and remained significant after adjustment for age and sex [1.31 (1.06, 1.61); P = 0.01] and after adjustment for relevant covariates [1.24 (1.01, 1.52); P = 0.04]. In a secondary analysis of the individual event types, Ln OPN was significantly associated with incident hospitalization for heart failure: HR (95% CI) = 2.04 (1.44, 2.89); P <0.001, even after adjustment for age, sex and additional relevant covariates. In conclusion, in patients with stable CAD and preserved EF on optimal medical therapy, plasma OPN levels were independently associated with the composite incident endpoint of adverse cardiovascular outcomes as well as incident hospitalization for heart failure. PMID:27284698

  12. Distribution of alkaline phosphatase, osteopontin, RANK ligand and osteoprotegerin in calcified human carotid atheroma.

    PubMed

    Higgins, Catherine L; Isbilir, Salim; Basto, Pamela; Chen, Iou Yih; Vaduganathan, Muthiah; Vaduganathan, Periyanan; Reardon, Michael J; Lawrie, Gerald; Peterson, Leif; Morrisett, Joel D

    2015-10-01

    Ectopic vascular calcification is a significant component of atherosclerotic disease. Osteopontin (OPN), Osteoprotegerin (OPG), Receptor Activator of NFκB Ligand (RANKL), and alkaline phosphatase (ALP) are each thought to play central roles in the calcification or demineralization of atherosclerotic lesions. Abnormalities in the balance of these proteins may lead to perturbations in bone remodeling and arterial calcification. The purpose of this study was to measure the distribution of these proteins in human carotid lesions and to elucidate possible mechanism(s) whereby they control the deposition or depletion of arterial calcification. Thirty-three patients who had undergone carotid endarterectomy (CEA) within the previous 18 months and 11 control patients were enrolled. CEA specimens were analyzed by EBCT for calcification content in terms of Agatston (AGAT) and Volume scores. CEA specimens were then cut into 5 mm segments which were homogenized and extracted. Extracts were analyzed for tissue levels of calcium, phosphorus, ALP, OPN, RANKL, and OPG. Fasting blood samples were analyzed for the same components. In CEA tissue segments, the calcification levels (CHA AGAT) were inversely associated with the levels of OPG (r = -0.432/-0.579, p < 0.05) and positively associated with the levels of RANKL (r = 0.332/0.415, p < 0.05). In turn, the tissue levels of OPG were associated with homologous serum levels of OPG (r = 0.820/0.389, p < 0.001), and the tissue levels of RANKL were associated with the serum levels of homologous RANKL (r = 0.739/0.666, p < 0.0001). This study suggests that serum levels of OPG and RANKL may be useful biomarkers for estimating the degree of calcification in carotid atherosclerotic lesions. PMID:26307009

  13. Osteopontin and adiponectin: how far are they related in the complexity of psoriasis?

    PubMed

    Kadry, D; Hegazy, R A; Rashed, L

    2013-12-01

    Increasing attention has been drawn towards the involvement of both osteopontin (OPN) and adiponectin in psoriasis. The relationship between them has been studied before in the context of essential hypertension. To our knowledge, whether a relation between them exists in cases of psoriasis and the metabolic status in such patients have not been investigated. We aimed to verify their possible roles and relations in psoriasis and its metabolic associations. 35 patients with psoriasis vulgaris and 35 controls were included. Patients were clinically assessed by PASI and investigated for the presence of metabolic syndrome (MetS) and/or its components. Plasma levels of OPN and adiponectin were measured using ELISA. On comparing psoriatics to controls, patients showed significantly elevated levels of OPN (90.474 ± 21.22 vs 34.709 ± 13.95 ng/mL) and significantly depressed levels of adiponectin (4,586 ± 1.187 vs 5,905 ± 1.374 ng/mL), (p < 0.001). Strong negative correlation between plasma OPN and adiponectin was detected in patients (r = -0.912, p < 0.001), but not in controls. OPN elevation was related to diabetes mellitus, insulin resistance, and MetS. Adiponectin depression was related to body mass index, and MetS. This study demonstrates for the first time a significant correlation between OPN and adiponectin in psoriasis, hypothesized to be mostly attributed to the inflammatory milieu of psoriasis and MetS as well as the enhanced renin-angiotensin-aldosterone system previously documented in psoriasis. Adjuvant therapies aiming at modulating levels of OPN and adiponectin are speculated to add benefit in psoriasis treatment and protecting against its metabolic risks. PMID:23884541

  14. Epigenetic regulation of glucose-stimulated osteopontin (OPN) expression in diabetic kidney.

    PubMed

    Cai, Mengyin; Bompada, Pradeep; Atac, David; Laakso, Markku; Groop, Leif; De Marinis, Yang

    2016-01-01

    Diabetes nephropathy (DN) is the leading cause of end stage renal disease and it affects up to 40% of diabetic patients. In addition to hyperglycemia, genetic factors are thought to contribute to the development of DN, but few if any genetic factors have been convincingly linked to DN. Other possible mechanisms may involve epigenetic regulation of glucose-stimulated gene activity which was suggested to explain long-term effects of poor glycemic control on risk of diabetic complications, often referred to as metabolic memory. Osteopontin (OPN) is one of the genes upregulated in kidneys from diabetic mouse models as well as humans with DN, and suggested to play an important role in the pathogenesis of DN. In this study, we demonstrated that OPN gene expression is upregulated in the kidneys of a hyperglycemia diabetes mouse model SUR1-E1506K, and glucose-stimulated OPN gene expression is strongly associated with increases in activating histone marks H3K9ac, H3K4me1 and H3K4me3 and decrease in inactivating mark H3K27me3 in the promoter region of OPN gene. These findings were replicated in human mesangial cells treated with high glucose. Further proof for the involvement of histone acetylation and methylation in glucose-induced changes in OPN gene expression was obtained by manipulating histone modifications thereby OPN gene expression by histone deacetylase (HDAC) inhibitor trichostatin A and histone methyltransferase (HMT) inhibitor MM-102. We conclude that glucose is a potent inducer of histone acetylation and methylation, which in turn leads to upregulation of OPN gene expression. Treatment targeting histone marks may therefore represent an alternative method to protect kidneys from deleterious effects of glucose. PMID:26592666

  15. Protective effects of recombinant osteopontin on early brain injury after subarachnoid hemorrhage in rats

    PubMed Central

    Suzuki, Hidenori; Ayer, Robert; Sugawara, Takashi; Chen, Wanqiu; Sozen, Takumi; Hasegawa, Yu; Kanamaru, Kenji; Zhang, John H.

    2009-01-01

    Objective Accumulated evidence suggests that the primary cause of poor outcome after subarachnoid hemorrhage (SAH) is not only cerebral arterial narrowing, but also early brain injury (EBI). Our objective was to determine the effect of recombinant osteopontin (r-OPN), a pleiotropic extracellular matrix glycoprotein, on post-SAH EBI in rats. Design Controlled in vivo laboratory study. Setting Animal research laboratory. Subjects One hundred seventy-seven male adult Sprague-Dawley rats, 300–370g. Interventions The endovascular perforation model of SAH was produced. SAH or sham-operated rats were treated with an equal volume (1μL) of pre-SAH intracerebroventricular administration of two dosages (0.02 and 0.1μg) of r-OPN, albumin or vehicle. Body weight, neurological scores, brain edema and blood-brain barrier (BBB) disruption were evaluated, and Western blot analyses were performed to determine the effect of r-OPN on matrix metalloproteinase (MMP)-9, substrates of MMP-9 (zona occludens [ZO]-1, laminin), tissue inhibitor of MMP (TIMP)-1, inflammation (interleukin-1β), and nuclear factor (NF)-κ B signaling pathways. Measurements and Main Results Treatment with r-OPN prevented a significant loss in body weight, neurological impairment, brain edema, and BBB disruption after SAH. These effects were associated with the deactivation of NF-κB activity, inhibition of MMP-9 induction, the maintenance of TIMP-1, and the consequent preservation of the cerebral microvessel basal lamina protein laminin, and the tight junction protein ZO-1. Conclusions These results demonstrate that r-OPN treatment is effective for post-SAH EBI. PMID:19851092

  16. Osteopontin is involved in the formation of malignant pleural effusion in lung cancer.

    PubMed

    Cui, Ri; Takahashi, Fumiyuki; Ohashi, Rina; Yoshioka, Masakata; Gu, Tao; Tajima, Ken; Unnoura, Takeshi; Iwakami, Shinichiro; Hirama, Michihiro; Ishiwata, Toshiji; Iwase, Akihiko; Takahashi, Kazuhisa

    2009-03-01

    Malignant pleural effusion (MPE) is associated with advanced-stage lung cancer and is a poor prognostic sign for these patients. Osteopontin (OPN) is a multifunctional cytokine that is involved in the tumor progression and angiogenesis of lung cancer cells. The purpose of this study is to investigate and provide evidence for the role of OPN in the formation of MPE associated with lung cancer. In this study, we established an OPN knockdown murine lung cancer cell line, 3LL cells, utilizing the small interfering RNA (siRNA) technique. To reveal the effect of OPN on the formation of MPE associated with lung cancer, we directly injected OPN knockdown 3LL cells, 3LL/OPN siRNA, or control cells, 3LL/control siRNA, into the pleural space of C57BL/6 mice. OPN knockdown significantly reduced the formation of MPE, but did not inhibit in vivo tumor growth of 3LL cells in mice. Vascular endothelial growth factor (VEGF) concentration in MPE was markedly decreased in the 3LL/OPN siRNA in comparison with that of the 3LL/control siRNA. In vitro, recombinant OPN protein enhanced VEGF secretion from human umbilical vein endothelial cell (HUVEC) or human mesothelial cell line, Met5A cells, in a concentration-dependent manner. These results suggest that OPN is positively involved in the formation of MPE of lung cancer presumably by promoting VEGF secretion from vascular endothelial cells or mesothelial cells. OPN could be an effective target molecule for reducing MPE in lung cancer patients. PMID:18752867

  17. Osteopontin Neutralization Abrogates the Liver Progenitor Cell Response and Fibrogenesis in Mice

    PubMed Central

    Coombes, J; Swiderska-Syn, M; Dollé, L; Reid, D; Eksteen, B; Claridge, L; Briones-Orta, MA; Shetty, S; Oo, YH; Riva, A; Chokshi, S; Papa, S; Mi, Z; Kuo, PC; Williams, R; Canbay, A; Adams, DH; Diehl, AM; van Grunsven, LA; Choi, SS; Syn, WK

    2015-01-01

    Background Chronic liver injury triggers a progenitor-cell repair-response, and liver fibrosis occurs when repair becomes de-regulated. Previously, we reported that reactivation of the Hedgehog (Hh) pathway promotes fibrogenic liver-repair. Osteopontin (OPN) is a Hh-target, and a cytokine that is highly upregulated in fibrotic tissues, and regulates stem-cell fate. Thus, we hypothesized that OPN may modulate liver progenitor-cell response, and thereby, modulate fibrotic outcomes. We further evaluated the impact of OPN-neutralization on murine liver fibrosis. Methods Liver progenitors (603B and BMOL) were treated with OPN-neutralizing aptamers in the presence or absence of TGF–β, to determine if (and how) OPN modulates liver progenitor function. Effects of OPN-neutralization (using OPN-aptamers or OPN-neutralizing antibodies) on liver progenitor-cell response and fibrogenesis were assessed in three models of liver fibrosis (carbon tetrachloride, methionine-choline deficient diet, 3, 5,-diethoxycarbonyl-1,4-dihydrocollidine diet) by qRTPCR, Sirius-Red staining, hydroxyproline assay, and semi-quantitative double-immunohistochemistry. Finally, OPN expression and liver progenitor response were corroborated in liver tissues obtained from patients with chronic liver disease. Results OPN is over-expressed by liver progenitors in humans and mice. In cultured progenitors, OPN enhances viability and wound-healing by modulating TGF-β signaling. In vivo, OPN-neutralization attenuates the liver progenitor-cell response, reverses epithelial-mesenchymal-transition in Sox9+ cells, and abrogates liver fibrogenesis. Conclusions OPN upregulation during liver injury is a conserved repair-response, and influences liver progenitor-cell function. OPN-neutralization abrogates the liver progenitor-cell response and fibrogenesis in mouse models of liver fibrosis. PMID:24902765

  18. Regulation of Erk1/2 activation by osteopontin in PC3 human prostate cancer cells

    PubMed Central

    2010-01-01

    Background Osteopontin (OPN) has been shown to play many roles in the progression of cancer. We have recently demonstrated the activation of Akt by OPN. Integrin-linked kinase and PI3-kinase are integral proteins in OPN/AKT pathway in PC3 cells. To investigate the role of the extracellular receptors in OPN signaling, we have examined the spatio-temporal regulation of CD44 and integrin αvβ3 receptor in OPN-induced Akt activation in PC3 cells. Results Here, our studies demonstrate that OPN can activate Akt either through the αVβ3 integrin or the CD44 cell surface receptor. Members of the Mitogen Activated Protein Kinase (MAPK) family have been shown to be up-regulated in a variety of human cancers and have been implicated in the metastatic behavior. Our studies have demonstrated an increase in the phosphorylation of c-Raf at Ser259 and Ser338 in PC3 cells over-expressing OPN. This increase matches up with the Erk1/2 phosphorylation at Thr202/204 and activation. However, the inhibition of Akt activity augments the phosphorylation state of ERK1/2 to two to three fold with a concomitant reduction in the phosphorylation state of c-Raf at Ser259. Conclusions Regulation c-Raf phosphorylation at Ser259 has a role in the anti-apoptotic pathways mediated by Akt or Raf/MEK/ERK proteins. OPN may have dual effects in the activation of Erk1/2. We propose this based on the observations that while OPN activates c-Raf and Erk1/2; it also acts to inhibit c-Raf and Erk1/2 activation through Akt pathway. Our observations suggest that the activation of c-Raf-ERK cascade may promote cell cycle arrest in prostate cancer cells and OPN signaling has a role in the anti-apoptotic mechanism. PMID:20868520

  19. Osteopontin in relation to Prognosis following Coronary Artery Bypass Graft Surgery.

    PubMed

    Sbarouni, Eftihia; Georgiadou, Panagiota; Chatzikyriakou, Sofia; Analitis, Antonis; Chaidaroglou, Antigoni; Degiannis, Demitris; Voudris, Vassilis

    2016-01-01

    Cardiovascular events may occur even after complete revascularization in patients with coronary artery disease. We measured preoperative osteopontin (OPN) levels in 131 consecutive patients (66.5 ± 10 years old, 117 men and 14 women) with left ventricular ejection fraction of 50.7 ± 9.2% and low logistic EuroScore (3.5 ± 3.2%) undergoing elective Coronary Artery Bypass Grafting (CABG) surgery. Patients were prospectively followed up for a median of 12 months (range 11-24). The primary study endpoint was the composite of cardiovascular death, nonfatal myocardial infarction, need for repeat revascularization, and hospitalization for cardiovascular events. Pre-op OPN plasma levels were 77.9 (49.5, 150.9). Patients with prior acute myocardial infarction (AMI) had significantly higher OPN levels compared to those without [131.5 (52.2, 219) versus 73.3 (45.1, 125), p = 0.007]. OPN levels were positively related to EuroScore (r = 0.2, p = 0.031). Pre-op OPN levels did not differ between patients who had a major adverse event during follow-up compared to those with no event (p = 0.209) and had no effect on the hazard of future adverse cardiac events [HR (95% CI): 1.48 (0.43-4.99), p = 0.527]. The history of AMI was associated with increased risk of subsequent cardiovascular events at follow-up (p = 0.02). OPN is associated with preoperative risk assessment prior to low-risk CABG but did not independently predict outcome. PMID:27597799

  20. Osteopontin in relation to Prognosis following Coronary Artery Bypass Graft Surgery

    PubMed Central

    Sbarouni, Eftihia; Chatzikyriakou, Sofia; Analitis, Antonis; Chaidaroglou, Antigoni; Degiannis, Demitris; Voudris, Vassilis

    2016-01-01

    Cardiovascular events may occur even after complete revascularization in patients with coronary artery disease. We measured preoperative osteopontin (OPN) levels in 131 consecutive patients (66.5 ± 10 years old, 117 men and 14 women) with left ventricular ejection fraction of 50.7 ± 9.2% and low logistic EuroScore (3.5 ± 3.2%) undergoing elective Coronary Artery Bypass Grafting (CABG) surgery. Patients were prospectively followed up for a median of 12 months (range 11–24). The primary study endpoint was the composite of cardiovascular death, nonfatal myocardial infarction, need for repeat revascularization, and hospitalization for cardiovascular events. Pre-op OPN plasma levels were 77.9 (49.5, 150.9). Patients with prior acute myocardial infarction (AMI) had significantly higher OPN levels compared to those without [131.5 (52.2, 219) versus 73.3 (45.1, 125), p = 0.007]. OPN levels were positively related to EuroScore (r = 0.2, p = 0.031). Pre-op OPN levels did not differ between patients who had a major adverse event during follow-up compared to those with no event (p = 0.209) and had no effect on the hazard of future adverse cardiac events [HR (95% CI): 1.48 (0.43–4.99), p = 0.527]. The history of AMI was associated with increased risk of subsequent cardiovascular events at follow-up (p = 0.02). OPN is associated with preoperative risk assessment prior to low-risk CABG but did not independently predict outcome. PMID:27597799

  1. Osteopontin is increased in the cerebrospinal fluid of patients with Alzheimer's disease and its levels correlate with cognitive decline.

    PubMed

    Comi, Cristoforo; Carecchio, Miryam; Chiocchetti, Annalisa; Nicola, Stefania; Galimberti, Daniela; Fenoglio, Chiara; Cappellano, Giuseppe; Monaco, Francesco; Scarpini, Elio; Dianzani, Umberto

    2010-01-01

    Inflammation is believed to play a role in Alzheimer's disease (AD). Osteopontin (OPN) is a molecule involved in macrophage recruitment and activation and implicated in neurodegeneration. In order to elucidate the role of OPN in AD, we evaluated its levels in serum and cerebrospinal fluid (CSF) of 67 AD patients, 46 frontotemporal dementia (FTD) patients, and 69 controls. We found that OPN levels: i) are significantly increased in the CSF of AD patients; ii) correlate with MMSE score; and iii) are higher in the early disease phases ( 2 years). These findings support a role of OPN in AD pathogenesis. PMID:20308780

  2. Bounding Species Distribution Models

    NASA Technical Reports Server (NTRS)

    Stohlgren, Thomas J.; Jarnevich, Cahterine S.; Morisette, Jeffrey T.; Esaias, Wayne E.

    2011-01-01

    Species distribution models are increasing in popularity for mapping suitable habitat for species of management concern. Many investigators now recognize that extrapolations of these models with geographic information systems (GIS) might be sensitive to the environmental bounds of the data used in their development, yet there is no recommended best practice for "clamping" model extrapolations. We relied on two commonly used modeling approaches: classification and regression tree (CART) and maximum entropy (Maxent) models, and we tested a simple alteration of the model extrapolations, bounding extrapolations to the maximum and minimum values of primary environmental predictors, to provide a more realistic map of suitable habitat of hybridized Africanized honey bees in the southwestern United States. Findings suggest that multiple models of bounding, and the most conservative bounding of species distribution models, like those presented here, should probably replace the unbounded or loosely bounded techniques currently used [Current Zoology 57 (5): 642-647, 2011].

  3. Bounding species distribution models

    USGS Publications Warehouse

    Stohlgren, T.J.; Jarnevich, C.S.; Esaias, W.E.; Morisette, J.T.

    2011-01-01

    Species distribution models are increasing in popularity for mapping suitable habitat for species of management concern. Many investigators now recognize that extrapolations of these models with geographic information systems (GIS) might be sensitive to the environmental bounds of the data used in their development, yet there is no recommended best practice for "clamping" model extrapolations. We relied on two commonly used modeling approaches: classification and regression tree (CART) and maximum entropy (Maxent) models, and we tested a simple alteration of the model extrapolations, bounding extrapolations to the maximum and minimum values of primary environmental predictors, to provide a more realistic map of suitable habitat of hybridized Africanized honey bees in the southwestern United States. Findings suggest that multiple models of bounding, and the most conservative bounding of species distribution models, like those presented here, should probably replace the unbounded or loosely bounded techniques currently used. ?? 2011 Current Zoology.

  4. Causality and Tsirelson's bounds

    SciTech Connect

    Buhrman, H.; Massar, S.

    2005-11-15

    We study the properties of no-signaling correlations that cannot be reproduced by local measurements on entangled quantum states. We say that such correlations violate Tsirelson bounds. We show that if these correlations are obtained by some reversible unitary quantum evolution U, then U cannot be written in the product form U{sub A}xU{sub B}. This implies that U can be used for signaling and for entanglement generation. This result is completely general and in fact can be viewed as a characterization of Tsirelson bounds. We then show how this result can be used as a tool to study Tsirelson bounds and we illustrate this by rederiving the Tsirelson bound of 2{radical}(2) for the Clauser-Horn-Shimony-Holt inequality, and by deriving a new Tsirelson bound for qutrits.

  5. Bound infragravity waves

    NASA Astrophysics Data System (ADS)

    Okihiro, Michele; Guza, R. T.; Seymour, R. J.

    1992-07-01

    Model predictions of bound (i.e., nonlinearly forced by and coupled to wave groups) infragravity wave energy are compared with about 2 years of observations in 8- to 13-m depths at Imperial Beach, California, and Barbers Point, Hawaii. Frequency-directional spectra of free waves at sea and swell frequencies, estimated with a small array of four pressure sensors, are used to predict the bound wave spectra below 0.04 Hz. The predicted total bound wave energy is always less than the observed infragravity energy, and the underprediction increases with increasing water depth and especially with decreasing swell energy. At most half, and usually much less, of the observed infragravity energy is bound. Bound wave spectra are also predicted with data from a single wave gage in 183-m depth at Point Conception, California, and the assumption of unidirectional sea and swell. Even with energetic swell, less than 10% of the total observed infragravity energy in 183-m depth is bound. Free waves, either leaky or edge waves, are more energetic than bound waves at both the shallow and deep sites. The low level of infragravity energy observed in 183-m depth compared with 8- to 13-m depths, with similarly moderate sea and swell energy, suggests that leaky (and very high-mode edge) waves contribute less than 10% of the infragravity energy in 8-13 m. Most of the free infragravity energy in shallow water is refractively trapped and does not reach deep water.

  6. Osteopontin-stimulated apoptosis in cardiac myocytes involves oxidative stress and mitochondrial death pathway: role of a pro-apoptotic protein BIK.

    PubMed

    Dalal, Suman; Zha, Qinqin; Singh, Mahipal; Singh, Krishna

    2016-07-01

    Increased osteopontin (OPN) expression in the heart, specifically in myocytes, associates with increased myocyte apoptosis and myocardial dysfunction. Recently, we provided evidence that OPN interacts with CD44 receptor, and induces myocyte apoptosis via the involvement of endoplasmic reticulum stress and mitochondrial death pathways. Here we tested the hypothesis that OPN induces oxidative stress in myocytes and the heart via the involvement of mitochondria and NADPH oxidase-4 (NOX-4). Treatment of adult rat ventricular myocytes (ARVMs) with OPN (20 nM) increased oxidative stress as analyzed by protein carbonylation, and intracellular reactive oxygen species (ROS) levels as analyzed by ROS detection kit and dichlorohydrofluorescein diacetate staining. Pretreatment with NAC (antioxidant), apocynin (NOX inhibitor), MnTBAP (superoxide dismutase mimetic), and mitochondrial KATP channel blockers (glibenclamide and 5-hydroxydecanoate) decreased OPN-stimulated ROS production, cytosolic cytochrome c levels, and apoptosis. OPN increased NOX-4 expression, while decreasing SOD-2 expression. OPN decreased mitochondrial membrane potential as measured by JC-1 staining, and induced mitochondrial abnormalities including swelling and reorganization of cristae as observed using transmission electron microscopy. OPN increased expression of BIK, a pro-apoptotic protein involved in reorganization of mitochondrial cristae. Expression of dominant-negative BIK decreased OPN-stimulated apoptosis. In vivo, OPN expression in cardiac myocyte-specific manner associated with increased protein carbonylation, and expression of NOX-4 and BIK. Thus, OPN induces oxidative stress via the involvement of mitochondria and NOX-4. It may affect mitochondrial morphology and integrity, at least in part, via the involvement of BIK. PMID:27262843

  7. Osteopontin Is Essential for Type I Collagen Secretion in Reparative Dentin.

    PubMed

    Saito, K; Nakatomi, M; Ida-Yonemochi, H; Ohshima, H

    2016-08-01

    Osteopontin (OPN) is a highly phosphorylated glycoprotein that is a prominent component of the mineralized extracellular matrix of bone. The secretion of OPN by immunocompetent cells plays a role in the differentiation of odontoblast-like cells during pulpal healing following tooth transplantation. This study aimed to clarify the role of OPN during reparative dentinogenesis. A groove-shaped cavity was prepared on the mesial surface of the upper first molars of wild-type (WT) and Opn knockout (KO) mice, and the samples were collected at intervals of 1 to 14 d. The demineralized sections were processed for immunohistochemistry for Ki67, nestin, OPN, dentin sialoprotein (DSP), integrin αvβ3, and type I collagen; in situ hybridization for Opn, col1a1, and dentin sialophosphoprotein (Dspp); and apoptosis assay. For the loss and gain of function experiments, an in vitro culture assay for evaluating dentin-pulp complex regeneration was performed. On day 1 in WT mice, odontoblasts beneath the affected dentin lost nestin immunoreactivity. On day 3, the expression of Opn was recognized at the mesial dental pulp, and OPN was deposited along the predentin-dentin border. Nestin-positive newly differentiated odontoblast-like cells expressed both Dspp and col1a1 and showed positive immunoreactivity for integrin αvβ3, DSP, and type I collagen. Until day 14, reparative dentin formation continued next to the preexisting dentin at the mesial coronal pulp. In contrast, there was no reparative dentin in the Opn KO mice where nestin- and DSP-positive newly differentiated odontoblast-like cells lacked immunoreaction for type I collagen. The in vitro organ culture demonstrated that the administration of recombinant OPN rescued the type I collagen secretion by odontoblast-like cells in the Opn KO mice. The results suggested that the deposition of OPN at the calcification front is essential for the type I collagen secretion by newly differentiated odontoblast-like cells to form

  8. Synthesis, characterization, and clinical evaluation of conjugate vaccines composed of the O-specific polysaccharides of Shigella dysenteriae type 1, Shigella flexneri type 2a, and Shigella sonnei (Plesiomonas shigelloides) bound to bacterial toxoids.

    PubMed Central

    Taylor, D N; Trofa, A C; Sadoff, J; Chu, C; Bryla, D; Shiloach, J; Cohen, D; Ashkenazi, S; Lerman, Y; Egan, W

    1993-01-01

    The theoretic basis for developing conjugate vaccines, to induce immunoglobulin G (IgG) lipopolysaccharide (LPS) antibodies for the prevention of shigellosis, has been described (J. B. Robbins, C.-Y. Chu, and R. Schneerson, Clin. Infect. Dis. 15:346-361, 1992). The O-specific polysaccharides (O-SPs) of Shigella dysenteriae type 1, S. flexneri type 2a, and S. sonnei were covalently bound to carrier proteins. Alone, the O-SPs were not immunogenic in mice. Conjugates of these O-SPs, injected into young outbred mice subcutaneously as saline solutions containing 2.5 micrograms of saccharide, elicited serum IgG and IgM antibodies with booster responses; adsorption onto alum enhanced their immunogenicity. Injection of 25 micrograms of these conjugates into adult volunteers elicited mild local reactions only. Each conjugate induced a significant rise of the geometric mean serum IgG, IgM, and IgA LPS antibody levels. A second injection 6 weeks later did not elicit booster responses, and adsorption of the conjugates onto alum did not enhance their immunogenicity. Conjugate-induced levels of IgA, but not IgG or IgM, declined to preimmunization levels at day 56. The levels of postimmunization antibodies of the three immunoglobulin classes were similar to or higher than those of recruits in the Israel Defense Force following shigellosis caused by S. flexneri type 2a or S. sonnei. These data provide the basis for evaluating these conjugates to prevent shigellosis. PMID:8359890

  9. Structure of the Escherichia coli Antitoxin MqsA (YgiT/b3021) Bound to Its Gene Promoter Reveals Extensive Domain Rearrangements and the Specificity of Transcriptional Regulation

    SciTech Connect

    B Brown; T Wood; W Peti; R Page

    2011-12-31

    Bacterial cultures, especially biofilms, produce a small number of persister cells, a genetically identical subpopulation of wild type cells that are metabolically dormant, exhibit multidrug tolerance, and are highly enriched in bacterial toxins. The gene most highly up-regulated in Escherichia coli persisters is mqsR, a ribonuclease toxin that, along with mqsA, forms a novel toxin-antitoxin (TA) system. Like all known TA systems, both the MqsR-MqsA complex and MqsA alone regulate their own transcription. Despite the importance of TA systems in persistence and biofilms, very little is known about how TA modules, and antitoxins in particular, bind and recognize DNA at a molecular level. Here, we report the crystal structure of MqsA bound to a 26-bp fragment from the mqsRA promoter. We show that MqsA binds DNA predominantly via its C-terminal helix-turn-helix domain, with direct binding of recognition helix residues Asn{sup 97} and Arg{sup 010} to the DNA major groove. Unexpectedly, the structure also revealed that the MqsA N-terminal domain interacts with the DNA phosphate backbone. This results in a more than 105{sup o} rotation of the N-terminal domains between the free and complexed states, an unprecedented rearrangement for an antitoxin. The structure also shows that MqsA bends the DNA by more than 55{sup o} in order to achieve symmetrical binding. Finally, using a combination of biochemical and NMR studies, we show that the DNA sequence specificity of MqsA is mediated by direct readout.

  10. Osteopontin Immunoreactivity in Peripheral Blood Mononuclear Cells, Ileum, and Ileocecal Lymph Node of Dairy Cows Naturally Infected with Mycobacterium avium subsp. paratuberculosis

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Osteopontin (Opn), a highly acidic glycoprotein, plays an early role in initiating the innate immune response to mycobacterial infections by promoting cellular adhesion and recruitment of inflammatory cells from the peripheral blood. The formation of granulomas at the site of Mycobacterium avium s...

  11. Osteopontin in Spontaneous Germinal Centers Inhibits Apoptotic Cell Engulfment and Promotes Anti-Nuclear Antibody Production in Lupus-Prone Mice.

    PubMed

    Sakamoto, Keiko; Fukushima, Yuji; Ito, Koyu; Matsuda, Michiyuki; Nagata, Shigekazu; Minato, Nagahiro; Hattori, Masakazu

    2016-09-15

    Disposal of apoptotic cells is important for tissue homeostasis. Defects in this process in immune tissues may lead to breakdown of self-tolerance against intracellular molecules, including nuclear components. Development of diverse anti-nuclear Abs (ANAs) is a hallmark of lupus, which may arise, in part, due to impaired apoptotic cell clearance. In this work, we demonstrate that spontaneous germinal centers (GCs) in lupus-prone mice contain significantly elevated levels of unengulfed apoptotic cells, which are otherwise swiftly engulfed by tingible body macrophages. We indicate that osteopontin (OPN) secreted by CD153(+) senescence-associated T cells, which selectively accumulate in the GCs of lupus-prone mice, interferes with phagocytosis of apoptotic cells specifically captured via MFG-E8. OPN induced diffuse and prolonged Rac1 activation in phagocytes via integrin αvβ3 and inhibited the dissolution of phagocytic actin cup, causing defective apoptotic cell engulfment. In wild-type B6 mice, administration of TLR7 ligand also caused spontaneous GC reactions with increasing unengulfed apoptotic cells and ANA production, whereas B6 mice deficient for Spp1 encoding OPN showed less apoptotic cells and developed significantly reduced ANAs in response to TLR7 ligand. Our results suggest that OPN secreted by follicular CD153(+) senescence-associated T cells in GCs promotes a continuous supply of intracellular autoantigens via apoptotic cells, thus playing a key role in the progression of the autoreactive GC reaction and leading to pathogenic autoantibody production in lupus-prone mice. PMID:27534552

  12. Higher Matrix Stiffness Upregulates Osteopontin Expression in Hepatocellular Carcinoma Cells Mediated by Integrin β1/GSK3β/β-Catenin Signaling Pathway

    PubMed Central

    You, Yang; Zheng, Qiongdan; Dong, Yinying; Wang, Yaohui; Zhang, Lan; Xue, Tongchun; Xie, Xiaoying; Hu, Chao; Wang, Zhiming; Chen, Rongxin; Wang, Yanhong; Cui, Jiefeng; Ren, Zhenggang

    2015-01-01

    Increased stromal stiffness is associated with hepatocellular carcinoma (HCC) development and progression. However, the molecular mechanism by which matrix stiffness stimuli modulate HCC progress is largely unknown. In this study, we explored whether matrix stiffness-mediated effects on osteopontin (OPN) expression occur in HCC cells. We used a previously reported in vitro culture system with tunable matrix stiffness and found that OPN expression was remarkably upregulated in HCC cells with increasing matrix stiffness. Furthermore, the phosphorylation level of GSK3β and the expression of nuclear β-catenin were also elevated, indicating that GSK3β/β-catenin pathway might be involved in OPN regulation. Knock-down analysis of integrin β1 showed that OPN expression and p-GSK3β level were downregulated in HCC cells grown on high stiffness substrate compared with controls. Simultaneously, inhibition of GSK-3β led to accumulation of β-catenin in the cytoplasm and its enhanced nuclear translocation, further triggered the rescue of OPN expression, suggesting that the integrin β1/GSK-3β/β-catenin pathway is specifically activated for matrix stiffness-mediated OPN upregulation in HCC cells. Tissue microarray analysis confirmed that OPN expression was positively correlated with the expression of LOX and COL1. Taken together, high matrix stiffness upregulated OPN expression in HCC cells via the integrin β1/GSK-3β/β-catenin signaling pathway. It highlights a new insight into a pathway involving physical mechanical signal and biochemical signal molecules which contributes to OPN expression in HCC cells. PMID:26280346

  13. Bounding the Bogoliubov coefficients

    SciTech Connect

    Boonserm, Petarpa; Visser, Matt

    2008-11-15

    While over the last century or more considerable effort has been put into the problem of finding approximate solutions for wave equations in general, and quantum mechanical problems in particular, it appears that as yet relatively little work seems to have been put into the complementary problem of establishing rigourous bounds on the exact solutions. We have in mind either bounds on parametric amplification and the related quantum phenomenon of particle production (as encoded in the Bogoliubov coefficients), or bounds on transmission and reflection coefficients. Modifying and streamlining an approach developed by one of the present authors [M. Visser, Phys. Rev. A 59 (1999) 427-438, (arXiv:quant-ph/9901030)], we investigate this question by developing a formal but exact solution for the appropriate second-order linear ODE in terms of a time-ordered exponential of 2x2 matrices, then relating the Bogoliubov coefficients to certain invariants of this matrix. By bounding the matrix in an appropriate manner, we can thereby bound the Bogoliubov coefficients.

  14. Validation of EMP bounds

    SciTech Connect

    Warne, L.K.; Merewether, K.O.; Chen, K.C.; Jorgenson, R.E.; Morris, M.E.; Solberg, J.E.; Lewis, J.G.; Derr, W.

    1996-07-01

    Test data on canonical weapon-like fixtures are used to validate previously developed analytical bounding results. The test fixtures were constructed to simulate (but be slightly worse than) weapon ports of entry but have known geometries (and electrical points of contact). The exterior of the test fixtures exhibited exterior resonant enhancement of the incident fields at the ports of entry with magnitudes equal to those of weapon geometries. The interior consisted of loaded transmission lines adjusted to maximize received energy or voltage but incorporating practical weapon geometrical constraints. New analytical results are also presented for bounding the energies associated with multiple bolt joints and for bounding the exterior resonant enhancement of the exciting fields.

  15. Osteopontin is associated with nuclear factor {kappa}B gene expression during tail-suspension-induced bone loss

    SciTech Connect

    Ishijima, Muneaki; Ezura, Yoichi . E-mail: ezura.mph@mril.tmd.ac.jp; Tsuji, Kunikazu

    2006-10-01

    Osteoporosis due to unloading-induced bone loss is a critical issue in the modern aging society. Although the mechanisms underlying this phenomenon are largely unknown, osteopontin (OPN) is one of the critical mediators required for unloading-induced bone loss [M. Ishijima, S.R. Rittling, T. Yamashita, K. Tsuji, H. Kurosawa, A. Nifuji, D.T. Denhardt, and M. Noda, Enhancement of osteoclastic bone resorption and suppression of osteoblastic bone formation in response to reduced mechanical stress do not occur in the absence of osteopontin, J Exp Med, 193 (2001) 399-404]. To clarify the molecular bases for OPN actions, we carried out microarray analyses on the genes expressed in the femoral bone marrow cells in wild type and OPN-/- mice. The removal of the mechanical load induced bone loss in wild type, but not in OPN-/- mice, as previously reported. Expression analysis of 9586 cDNAs on a microarray system revealed that OPN deficiency blocked tail-suspension-induced expression of ten genes (group A). This observation was confirmed based on semi-quantitative RT-PCR analyses. On the other hand, expression of four genes (group B) was not altered by tail suspension in wild type but was enhanced in OPN-deficient mice. NF-{kappa}B p105 subunit gene (Nfkb1) was found in group A and Bax in group B. p53 gene expression was upregulated by tail suspension in wild type mice, but it was no longer observed in OPN-/- mice. These data indicate that OPN acts to mediate mechanical stress signaling upstream to the genes encoding apoptosis-related molecules, and its action is associated with alteration of the genes.

  16. Osteopontin promoter polymorphisms at locus -443 are associated with metastasis and poor prognosis of human intrahepatic cholangiocarcinoma in Chinese population

    PubMed Central

    Zhao, Xiang-Qian; Ma, Huan-Xian; Su, Mao-Sheng; He, Lei

    2014-01-01

    Purpose: Osteopontin (OPN) is known to be a secreted adhesive glycoprotein. Role of OPN in human intrahepatic cholangiocarcinoma (ICC) has not been well understood. This study explored whether genetic variations in the osteopontin gene are associated with ICC risk, progression and metastasis. Material and methods: 260 patients with stages I to IV between 2008 and 2013 were recruited in this study and same number healthy persons were used as control. OPN-66 T/G, -156 G/GG and -443 C/T variants were genotyped using DNA from blood lymphocytes. Chi-square test and a Fisher’s exact test were used to analyze the genotype distribution between healthy subjects and patients, and further its distribution among TNM stages and incidence metastasis in patients. Results: For the variant at nt- 443 (CC), there was a significant difference between the number of patients with stage IV and those with all other stages of ICC (P < 0.01). Patients with -443 (CC) variant had significant higher incidence of lymph and distant metastasis development compared to other genotypes. For the variant at nt- 443 (CT), there was a significant difference between the number of ICC patients with stage III + IV and those with stage I + II (P < 0.01). The survival rates for ICC patients with the C/C genotype were significantly lower than for patients with the other two genotypes (C/T, T/T). Conclusion: OPN -443 C/T polymorphism is a potential predictive marker of metastasis and poor prognosis in ICC patients. PMID:25400775

  17. Computing Graphical Confidence Bounds

    NASA Technical Reports Server (NTRS)

    Mezzacappa, M. A.

    1983-01-01

    Approximation for graphical confidence bounds is simple enough to run on programmable calculator. Approximation is used in lieu of numerical tables not always available, and exact calculations, which often require rather sizable computer resources. Approximation verified for collection of up to 50 data points. Method used to analyze tile-strength data on Space Shuttle thermal-protection system.

  18. Petawatt laser absorption bounded

    PubMed Central

    Levy, Matthew C.; Wilks, Scott C.; Tabak, Max; Libby, Stephen B.; Baring, Matthew G.

    2014-01-01

    The interaction of petawatt (1015 W) lasers with solid matter forms the basis for advanced scientific applications such as table-top particle accelerators, ultrafast imaging systems and laser fusion. Key metrics for these applications relate to absorption, yet conditions in this regime are so nonlinear that it is often impossible to know the fraction of absorbed light f, and even the range of f is unknown. Here using a relativistic Rankine-Hugoniot-like analysis, we show for the first time that f exhibits a theoretical maximum and minimum. These bounds constrain nonlinear absorption mechanisms across the petawatt regime, forbidding high absorption values at low laser power and low absorption values at high laser power. For applications needing to circumvent the absorption bounds, these results will accelerate a shift from solid targets, towards structured and multilayer targets, and lead the development of new materials. PMID:24938656

  19. Petawatt laser absorption bounded

    NASA Astrophysics Data System (ADS)

    Levy, Matthew C.; Wilks, Scott C.; Tabak, Max; Libby, Stephen B.; Baring, Matthew G.

    2014-06-01

    The interaction of petawatt (1015 W) lasers with solid matter forms the basis for advanced scientific applications such as table-top particle accelerators, ultrafast imaging systems and laser fusion. Key metrics for these applications relate to absorption, yet conditions in this regime are so nonlinear that it is often impossible to know the fraction of absorbed light f, and even the range of f is unknown. Here using a relativistic Rankine-Hugoniot-like analysis, we show for the first time that f exhibits a theoretical maximum and minimum. These bounds constrain nonlinear absorption mechanisms across the petawatt regime, forbidding high absorption values at low laser power and low absorption values at high laser power. For applications needing to circumvent the absorption bounds, these results will accelerate a shift from solid targets, towards structured and multilayer targets, and lead the development of new materials.

  20. Diagnostic Performance of Alpha-Fetoprotein, Protein Induced by Vitamin K Absence, Osteopontin, Dickkopf-1 and Its Combinations for Hepatocellular Carcinoma

    PubMed Central

    Jang, Eun Sun; Jeong, Sook-Hyang; Kim, Jin-Wook; Choi, Yun Suk; Leissner, Philippe; Brechot, Christian

    2016-01-01

    Background & Aims Alpha-fetoprotein (AFP) is the most widely used serum biomarker for hepatocellular carcinoma (HCC), despite its limitations. As complementary biomarkers, protein induced by vitamin K absence (PIVKA-II), osteopontin (OPN), and Dickkopf-1 (DKK-1) have been proposed. This study aimed to perform a head-to-head comparison of the diagnostic performance of AFP, PIVKA-II, OPN and DKK-1 as single or in combination to seek the best biomarker or panel, and to investigate the clinical factors affecting their performance. Methods Using 401 stored plasma samples obtained from 208 HCC patients and 193 liver cirrhosis control patients, plasma AFP, PIVKA-II, OPN and DKK-1 levels were measured by ELISA, and receiver operating characteristic curve analyses were performed for each biomarker and for every combination of two to four markers. Results Of the four biomarkers, AFP showed the highest area under the curve (0.786). The sensitivity and specificity for each single biomarker was 62% and 90.2% (AFP>20 ng/mL), 51.0% and 91.2% (PIVKA-II>10 ng/mL), 46.2% and 80.3% (OPN>100 ng/mL), and 50.0% and 80.8% (DKK-1>500 pg/mL), respectively. Among the combinations of two biomarkers, AFP>20 ng/mL or DKK-1>500 pg/mL showed the best diagnostic performance (sensitivity 78.4%, specificity 72.5%). Triple or quadruple combination did not improve the diagnostic performance further. The patient’s age, etiology and tumor invasiveness of HCC affected the performance of each marker. Conclusions AFP was the most useful single biomarker for HCC diagnosis, and the combined measurement of AFP and DKK-1 could maximize the diagnostic yield. Clinical decision should be based on the consideration of various factors affecting the diagnostic performance of each biomarker. Efforts to seek novel HCC biomarkers should be continued. PMID:26986465

  1. CD133+ and Nestin+ Glioma Stem-Like Cells Reside Around CD31+ Arterioles in Niches that Express SDF-1α, CXCR4, Osteopontin and Cathepsin K.

    PubMed

    Hira, Vashendriya V V; Ploegmakers, Kimberley J; Grevers, Frederieke; Verbovšek, Urška; Silvestre-Roig, Carlos; Aronica, Eleonora; Tigchelaar, Wikky; Turnšek, Tamara Lah; Molenaar, Remco J; Van Noorden, Cornelis J F

    2015-07-01

    Poor survival of high-grade glioma is at least partly caused by glioma stem-like cells (GSLCs) that are resistant to therapy. GSLCs reside in niches in close vicinity of endothelium. The aim of the present study was to characterize proteins that may be functional in the GSLC niche by performing immunohistochemistry on serial cryostat sections of human high-grade glioma samples. We have found nine niches in five out of five high-grade glioma samples that were all surrounding arterioles with CD31+ endothelial cells and containing cellular structures that were CD133+ and nestin+. All nine niches expressed stromal-derived factor-1α (SDF-1α), its receptor C-X-C chemokine receptor type 4 (CXCR4), osteopontin and cathepsin K. SDF-1α plays a role in homing of CXCR4+ stem cells and leukocytes, whereas osteopontin and cathepsin K promote migration of cancer cells and leukocytes. Leukocyte-related markers, such as CD68, macrophage matrix metalloprotease-9, CD177 and neutrophil elastase were often but not always detected in the niches. We suggest that SDF-1α is involved in homing of CXCR4+ GSLCs and leukocytes and that cathepsin K and osteopontin are involved in the migration of GSLCs out of the niches. PMID:25809793

  2. Bound charges and currents

    NASA Astrophysics Data System (ADS)

    Herczyński, Andrzej

    2013-03-01

    Bound charges and currents are among the conceptually challenging topics in advanced courses on electricity and magnetism. It may be tempting for students to believe that they are merely computational tools for calculating electric and magnetic fields in matter, particularly because they are usually introduced through abstract manipulation of integral identities, with the physical interpretation provided a posteriori. Yet these charges and currents are no less real than free charges and currents and can be measured experimentally. A simpler and more direct approach to introducing this topic, suggested by the ideas in the classic book by Purcell and emphasizing the physical origin of these phenomena, is proposed.

  3. Monoclonal antibodies specific to a Ca2(+)-bound form of lipocortin I distinguish its Ca2(+)-dependent phospholipid-binding ability from its ability to inhibit phospholipase A2.

    PubMed

    Hayashi, H; Owada, M K; Sonobe, S; Domae, K; Yamanouchi, T; Kakunaga, T; Kitajima, Y; Yaoita, H

    1990-08-01

    Lipocortin I, a Ca2(+)-and phospholipid-binding protein without EF-hand structures, has many biological effects in vitro. Its actual role in vivo, however is unknown. We obtained and characterized five monoclonal antibodies to lipocortin I. Two of these monoclonal antibodies (L2 and L4-MAbs) reacted with the Ca(+)-bound form of lipocortin I, but not with the Ca2(+)-free form, both in vivo and in vitro. Lipocortin I required greater than or equal to 10 microM-Ca2+ to bind the two antibodies, and this Ca2+ requirement was not affected by phosphatidylserine. L2-MAb abolished the phospholipase A2 inhibitory activity of lipocortin I and inhibited its binding to Escherichia coli membranes and to phosphatidylserine in vitro. L4-MAb abolished the phospholipase A2 inhibitory activity of lipocortin I, but did not affect its binding to E. coli membranes or to phosphatidylserine. These findings indicated that the inhibition of phospholipase A2 by lipocortin I was not simply due to removal or capping of the substrates in E. coli membranes. Furthermore, an immunofluorescence study using L2-MAb showed the actual existence of Ca2(+)-bound form of lipocortin I in vivo. PMID:2143895

  4. Monoclonal antibodies specific to a Ca2(+)-bound form of lipocortin I distinguish its Ca2(+)-dependent phospholipid-binding ability from its ability to inhibit phospholipase A2.

    PubMed Central

    Hayashi, H; Owada, M K; Sonobe, S; Domae, K; Yamanouchi, T; Kakunaga, T; Kitajima, Y; Yaoita, H

    1990-01-01

    Lipocortin I, a Ca2(+)-and phospholipid-binding protein without EF-hand structures, has many biological effects in vitro. Its actual role in vivo, however is unknown. We obtained and characterized five monoclonal antibodies to lipocortin I. Two of these monoclonal antibodies (L2 and L4-MAbs) reacted with the Ca(+)-bound form of lipocortin I, but not with the Ca2(+)-free form, both in vivo and in vitro. Lipocortin I required greater than or equal to 10 microM-Ca2+ to bind the two antibodies, and this Ca2+ requirement was not affected by phosphatidylserine. L2-MAb abolished the phospholipase A2 inhibitory activity of lipocortin I and inhibited its binding to Escherichia coli membranes and to phosphatidylserine in vitro. L4-MAb abolished the phospholipase A2 inhibitory activity of lipocortin I, but did not affect its binding to E. coli membranes or to phosphatidylserine. These findings indicated that the inhibition of phospholipase A2 by lipocortin I was not simply due to removal or capping of the substrates in E. coli membranes. Furthermore, an immunofluorescence study using L2-MAb showed the actual existence of Ca2(+)-bound form of lipocortin I in vivo. Images Fig. 1. Fig. 2. Fig. 3. Fig. 4. Fig. 6. Fig. 7. Fig. 8. PMID:2143895

  5. Binding patterns of bovine seminal plasma proteins A1/A2, 30 kDa and osteopontin on ejaculated sperm before and after incubation with isthmic and ampullary oviductal fluid.

    PubMed

    Souza, Carlos Eduardo A; Moura, Arlindo A; Monaco, Elisa; Killian, Gary J

    2008-04-01

    Previous studies from our laboratory have reported empirical associations between bovine seminal plasma protein(s) (BSP) A1/A2 and 30 kDa and osteopontin (OPN) in accessory sex gland fluid and bull fertility. These BSP and OPN are believed to bind to sperm at ejaculation and to remain bound until sperm reach the oviduct. The objective of the present study was to evaluate the topographical distribution of BSP A1/A2, 30 kDa and OPN binding on: (1) bovine ejaculated sperm; (2) ejaculated sperm incubated with isthmic oviductal fluid (ODF); (3) ejaculated sperm+isthmic ODF incubated in ampullary ODF. From each of these media, aliquots of sperm for BSP and OPN were processed for immunocytochemistry and analysis by laser scanning confocal microscopy. Isthmic and ampullary ODF was collected from indwelling catheters and used as pools from three cows in the non-luteal phase of the estrous cycle. Anti-BSP A1/A2 was detected bound to the midpiece, post-equatorial and equatorial segments and acrosome of sperm after ejaculation and after incubation with isthmic and ampullary ODF. The BSP A1/A2 fluorescence was more concentrated on the midpiece and increased as acrosome-intact sperm came in contact with ODF. As compared with acrosome-intact sperm, non-intact acrosome intact sperm had 39 and 68% reductions of acrosome fluorescence and 36% and 90% increases of post-equatorial fluorescence after contact with isthmic and ampullary ODF (P<0.05). Anti-BSP 30 kDa was more intense on the midpiece than on post-equatorial, equatorial and acrosome regions of sperm after ejaculation and contact with ODF. However, equatorial fluorescence was 141% and 89% more intense and acrosome stainning was 80% and 76% less (P<0.05) in non-intact acrosome sperm than in acrosome intact cells, during all ODF incubations. Anti-OPN was identified on the acrosome of ejaculated sperm, but with less fluorescence (P<0.05) on the post-equatorial segment and midpiece. Incubation of sperm with isthmic ODF increased

  6. Transactivation of human osteopontin promoter by human T-cell leukemia virus type 1-encoded Tax protein.

    PubMed

    Zhang, Jing; Yamada, Osamu; Matsushita, Yoshihisa; Chagan-Yasutan, Haorile; Hattori, Toshio

    2010-06-01

    Osteopontin (OPN) is a cytokine that contributes substantially to the growth and metastasis in a wide spectrum of malignancies. We report here that OPN gene is transactivated by Tax protein of human T-cell leukemia virus type 1 (HTLV-1). Northern blot showed enhanced OPN gene expression in cells stably expressing Tax. Co-expression of Tax increased the reporter gene expression directed by OPN promoter. Tax-induced OPN activation was abrogated by treatment with LY294002 (PI3K inhibitor) or co-transfection with AKT siRNA, suggesting PI3K/AKT pathway is involved in Tax-mediated transactivation. Reporter assay with deletion mutants showed that the 5'-partial sequence between -765 and -660 of the OPN promoter is the region responsive to Tax, and further, disrupting the AP-1 site within this region abolished the OPN induction by Tax, indicating that Tax activation of OPN promoter is likely mediated by AP-1 site. This study suggests that OPN is one of the downstream mediators of aberrantly activated PI3K/AKT signaling by Tax, which may partially contribute to HTLV-1-associated leukemogenesis. PMID:19767100

  7. Elevation of osteopontin levels in brain tumor cells reduces burden and promotes survival through the inhibition of cell dispersal

    PubMed Central

    Morrow, Jay; Barone, Tara A.; Hoffer, Alan; Lock, Jeffrey; DeChant, Anne; Mangla, Saisho; Plunkett, Robert J.; Miller, Robert H.

    2010-01-01

    Osteopontin (OPN) is a pleotrophic molecule that has been associated with multiple disorders of the central nervous system (CNS). Its roles in CNS malignancy are unclear but suggest that higher levels of OPN expression correlate with increased tumor grade and increased migratory capacity of tumor cells. In this study OPN cDNA was cloned into a retroviral vector and used to infect F98 Fischer rat-derived glioma cells and U87 human-derived glioblastoma multiforme (GBM) cells in vitro. Cells expressing high levels of OPN migrated less distance than control cells in vitro. This effect was not RGD mediated, but was reversed in the presence of c-Jun N-terminal kinase (JNK) inhibitor suggesting that JNK1 is an essential component of a negative feedback loop affecting OPN activated signaling cascades. Implantation of tumor cells expressing high levels of OPN into adult Fischer rats and nude rats resulted in morphologically distinct tumors and prolonged host survival relative to controls. We propose that local produced, high level OPN expression limits the malignant character of glioma cells and that the downstream mechanisms involved represent pathways that may have therapeutic value in the treatment of human CNS malignancy. PMID:17928956

  8. Genetic deletion of osteopontin in TRAMP mice skews prostate carcinogenesis from adenocarcinoma to aggressive human-like neuroendocrine cancers

    PubMed Central

    Mauri, Giorgio; Jachetti, Elena; Comuzzi, Barbara; Dugo, Matteo; Arioli, Ivano; Miotti, Silvia; Sangaletti, Sabina; Di Carlo, Emma; Tripodo, Claudio; Colombo, Mario P.

    2016-01-01

    Osteopontin (OPN) is a secreted glycoprotein, that belongs to the non-structural extracellular matrix (ECM), and its over expression in human prostate cancer has been associated with disease progression, androgen independence and metastatic ability. Nevertheless, the pathophysiology of OPN in prostate tumorigenesis has never been studied. We crossed TRansgenic Adenocarcinoma of the Mouse Prostate (TRAMP) mice with OPN deficient (OPN−/−) mice and followed tumor onset and progression in these double mutants. Ultrasound examination detected the early onset of a rapidly growing, homogeneous and spherical tumor in about 60% of OPN−/− TRAMP mice. Such neoplasms seldom occurred in parental TRAMP mice otherwise prone to adenocarcinomas and were characterized for being androgen receptor negative, highly proliferative and endowed with neuroendocrine (NE) features. Gene expression profiling showed up-regulation of genes involved in tumor progression, cell cycle and neuronal differentiation in OPN-deficient versus wild type TRAMP tumors. Down-regulated genes included key genes of TGFa pathway, including SMAD3 and Filamin, which were confirmed at the protein level. Furthermore, NE genes and particularly those characterizing early prostatic lesions of OPN-deficient mice were found to correlate with those of human prostate NE tumours. These data underscore a novel role of OPN in the early stages of prostate cancer growth, protecting against the development of aggressive NE tumors. PMID:26700622

  9. Genetic variations in the osteopontin promoters T-443C and G-156GG increase carotid intima–media thickness

    PubMed Central

    Yueniwati, Yuyun; Yurina, Valentina; Sobah, Nurus; Rahayu, Endang

    2016-01-01

    Carotid intima–media thickness (CIMT) is a clear predictor of atherosclerosis. The increase of CIMT is affected by mutations in the osteopontin (OPN) promoters. The purpose of this study was to examine genetic variations in OPN promoters T-443C and G-156GG, identified in Javanese children with ischemic stroke parents, and to investigate their relationship with the increase of CIMT. A case–control analytic study was performed on 20 case and 12 control samples. Case samples were Javanese children aged between 10 to 21 years with ischemic stroke parents. Control samples were children with healthy parents. Mutations of T-443C and G-156GG were determined by employing polymerase chain reaction. Results of sequencing were analyzed using CLC Main Workbench 6.0. CIMT was defined using ultrasound. Genetic variations of T-443C were identified in six samples. Likewise, genetic variations of G-156GG were identified in six samples. Genetic variations in the OPN promoters T-443C and G-156GG were not potential risk factors in an increase of CIMT (P=0.654 and P=0.654). This study proves that genetic variations could be identified at the points of T-443C and G-156GG in children with ischemic stroke parents. Although statistically insignificant, the tendency to increase CIMT occurs in children with genetic variations. Children with ischemic stroke parents have thicker CIMT than children of healthy parents. PMID:27274305

  10. Osteopontin Is Expressed in the Mouse Uterus during Early Pregnancy and Promotes Mouse Blastocyst Attachment and Invasion In Vitro

    PubMed Central

    Qi, Qian-Rong; Xie, Qing-Zhen; Liu, Xue-Li; Zhou, Yun

    2014-01-01

    Embryo implantation into the maternal uterus is a decisive step for successful mammalian pregnancy. Osteopontin (OPN) is a member of the small integrin-binding ligand N-linked glycoprotein family and participates in cell adhesion and invasion. In this study, we showed that Opn mRNA levels are up-regulated in the mouse uterus on day 4 and at the implantation sites on days 5 and 8 of pregnancy. Immunohistochemistry localized the OPN protein to the glandular epithelium on day 4 and to the decidual zone on day 8 of pregnancy. OPN mRNA and proteins are induced by in vivo and in vitro decidualization. OPN expression in the endometrial stromal cells is regulated by progesterone, a key regulator during decidualization. As a secreted protein, the protein level of OPN in the uterine cavity is enriched on day 4, and in vitro embryo culturing has indicated that OPN can facilitate blastocyst hatching and adhesion. Knockdown of OPN attenuates the adhesion and invasion of blastocysts in mouse endometrial stromal cells by suppressing the expression and enzymatic activity of matrix metalloproteinase-9 in the trophoblast. Our data indicated that OPN expression in the mouse uterus during early pregnancy is essential for blastocyst hatching and adhesion and that the knockdown of OPN in mouse endometrial stroma cells could lead to a restrained in vitro trophoblast invasion. PMID:25133541

  11. Osteopontin and latent-TGF β binding-protein 2 as potential diagnostic markers for HBV-related hepatocellular carcinoma.

    PubMed

    da Costa, Andre Nogueira; Plymoth, Amelie; Santos-Silva, Daniela; Ortiz-Cuaran, Sandra; Camey, Suzy; Guilloreau, Paule; Sangrajrang, Suleeporn; Khuhaprema, Thiravud; Mendy, Maimuna; Lesi, Olufunmilayo A; Chang, Hee-Kyung; Oh, Jin-Kyoung; Lee, Duk-Hee; Shin, Hai-Rim; Kirk, Gregory D; Merle, Philippe; Beretta, Laura; Hainaut, Pierre

    2015-01-01

    Chronic Hepatitis B (HB) is the main risk factor for chronic liver disease (CLD) and hepatocellular carcinoma (HCC) in many low-resource countries, where diagnosis is constrained by lack of clinical, histopathological and biomarker resources. We have used proteomics to detect plasma biomarkers that outperform α-Fetoprotein (AFP), the most widely used biomarker for HCC diagnosis in low-resource contexts. Deep-plasma proteome analysis was performed in HCC patients, patients with CLD and in HB-carrier controls from Thailand (South-East Asia) and The Gambia (West-Africa). Mass spectrometry profiling identified latent-transforming growth factor β binding-protein 2 (LTBP2) and Osteopontin (OPN) as being significantly elevated in HCC versus CLD and controls. These two proteins were further analyzed by ELISA in a total of 684 plasma samples, including 183 HCC, 274 CLD and 227 asymptomatic controls. When combined, LTBP2 and OPN showed an area under the receiver operating curve of 0.85 in distinguishing HCC from CLD in subjects with AFP <20 ng/mL. In a prospective cohort of 115 CLD patients from Korea, increased plasma levels of LTBP2 and/or OPN were detected in plasma collected over 2 years prior to diagnosis in 21 subjects who developed HCC. Thus, the combination of LTBP2 and OPN outperformed AFP for diagnosis and prediction of HCC and may therefore improve biomarker-based detection of HBV-related HCC. PMID:24803312

  12. Osteopontin reduced hypoxia-ischemia neonatal brain injury by suppression of apoptosis in a rat pup model

    PubMed Central

    Chen, Wanqiu; Ma, Qingyi; Suzuki, Hidenori; Hartman, Richard; Tang, Jiping; Zhang, John H.

    2011-01-01

    Background and Purpose Osteopontin (OPN) is neuroprotective in ischemic brain injuries in adult experimental models, therefore, we hypothesized that OPN would provide neuroprotection and improve long term neurological function in the immature brain after hypoxic-ischemic (HI) injury. Methods HI was induced by unilateral ligation of the right carotid artery followed by hypoxia (8% O2 for 2h) in postnatal day 7 rats. OPN (0.03 µg or 0.1 µg) was injected intracerebroventricularly at 1h post HI. Temporal expression of endogenous OPN was evaluated in the normal rat brain at the age of day 0, 4, 7, 11, 14, and 21, and in the ipsilateral hemisphere following HI. The effects of OPN were evaluated using TTC staining, apoptotic cell death assay, and cleaved caspase-3 expression. Neurological function was assessed by Morris water maze test. Results Endogenous OPN expression in the brain was the highest at the age of day 0, with continuous reduction till the age of day 21 during development. After HI injury, endogenous OPN expression was increased and peaked at 48h. Exogenous OPN decreased infarct volume and improved neurological outcomes 7 weeks after HI injury. OPN-induced neuroprotection was blocked by an integrin antagonist. Conclusions OPN-induced neuroprotection was associated with cleaved-caspase-3 inhibition and antiapoptotic cell death. OPN treatment improved long-term neurological function against neonatal HI brain injury. PMID:21273567

  13. Osteopontin Expression in the Brain Triggers Localized Inflammation and Cell Death When Immune Cells Are Activated by Pertussis Toxin

    PubMed Central

    Marcondes, Maria Cecilia Garibaldi; Ojakian, Ryan; Bortell, Nikki; Flynn, Claudia; Conti, Bruno; Fox, Howard S.

    2014-01-01

    Upregulation of osteopontin (OPN) is a characteristic of central nervous system pathologies. However, the role of OPN in inflammation is still controversial, since it can both prevent cell death and induce the migration of potentially damaging inflammatory cells. To understand the role of OPN in inflammation and cell survival, we expressed OPN, utilizing an adenoviral vector, in the caudoputamen of mice deficient in OPN, using beta-galactosidase- (β-gal-) expressing vector as control. The tissue pathology and the expression of proinflammatory genes were compared in both treatments. Interestingly, inflammatory infiltrate was only found when the OPN-vector was combined with a peripheral treatment with pertussis toxin (Ptx), which activated peripheral cells to express the OPN receptor CD44v6. Relative to β-gal, OPN increased the levels of inflammatory markers, including IL13Rα1, CXCR3, and CD40L. In Ptx-treated OPN KOs, apoptotic TUNEL+ cells surrounding the OPN expression site increased, compared to β-gal. Together, these results show that local OPN expression combined with a peripheral inflammatory stimulus, such as Ptx, may be implicated in the development of brain inflammation and induction of cell death, by driving a molecular pattern characteristic of cytotoxicity. These are characteristics of inflammatory pathologies of the CNS in which OPN upregulation is a hallmark. PMID:25525298

  14. Genetic deletion of osteopontin in TRAMP mice skews prostate carcinogenesis from adenocarcinoma to aggressive human-like neuroendocrine cancers.

    PubMed

    Mauri, Giorgio; Jachetti, Elena; Comuzzi, Barbara; Dugo, Matteo; Arioli, Ivano; Miotti, Silvia; Sangaletti, Sabina; Di Carlo, Emma; Tripodo, Claudio; Colombo, Mario P

    2016-01-26

    Osteopontin (OPN) is a secreted glycoprotein, that belongs to the non-structural extracellular matrix (ECM), and its over expression in human prostate cancer has been associated with disease progression, androgen independence and metastatic ability. Nevertheless, the pathophysiology of OPN in prostate tumorigenesis has never been studied. We crossed TRansgenic Adenocarcinoma of the Mouse Prostate (TRAMP) mice with OPN deficient (OPN-/-) mice and followed tumor onset and progression in these double mutants. Ultrasound examination detected the early onset of a rapidly growing, homogeneous and spherical tumor in about 60% of OPN-/- TRAMP mice. Such neoplasms seldom occurred in parental TRAMP mice otherwise prone to adenocarcinomas and were characterized for being androgen receptor negative, highly proliferative and endowed with neuroendocrine (NE) features. Gene expression profiling showed up-regulation of genes involved in tumor progression, cell cycle and neuronal differentiation in OPN-deficient versus wild type TRAMP tumors. Down-regulated genes included key genes of TGFa pathway, including SMAD3 and Filamin, which were confirmed at the protein level. Furthermore, NE genes and particularly those characterizing early prostatic lesions of OPN-deficient mice were found to correlate with those of human prostate NE tumours. These data underscore a novel role of OPN in the early stages of prostate cancer growth, protecting against the development of aggressive NE tumors. PMID:26700622

  15. Bounded link prediction in very large networks

    NASA Astrophysics Data System (ADS)

    Cui, Wei; Pu, Cunlai; Xu, Zhongqi; Cai, Shimin; Yang, Jian; Michaelson, Andrew

    2016-09-01

    Evaluating link prediction methods is a hard task in very large complex networks due to the prohibitive computational cost. However, if we consider the lower bound of node pairs' similarity scores, this task can be greatly optimized. In this paper, we study CN index in the bounded link prediction framework, which is applicable to enormous heterogeneous networks. Specifically, we propose a fast algorithm based on the parallel computing scheme to obtain all node pairs with CN values larger than the lower bound. Furthermore, we propose a general measurement, called self-predictability, to quantify the performance of similarity indices in link prediction, which can also indicate the link predictability of networks with respect to given similarity indices.

  16. Primary-backup protocols: Lower bounds and optimal implementations

    NASA Technical Reports Server (NTRS)

    Budhiraja, Navin; Marzullo, Keith; Schneider, Fred B.; Toueg, Sam

    1992-01-01

    A formal specification of primary-backup is presented. We then prove lower bounds on the degree of replication, failover time, and worst-case response time to client requests assuming different failure models. Finally, we outline primary-backup protocols and indicate which of our lower bounds are tight.

  17. Petawatt laser absorption bounded

    NASA Astrophysics Data System (ADS)

    Levy, Matthew; Wilks, Scott; Tabak, Max; Libby, Stephen; Baring, Matthew

    2014-10-01

    The interaction of petawatt (1015 W) lasers with solid matter forms the basis for advanced scientific applications such as table-top relativistic particle accelerators, ultrafast charged particle imaging systems and fast ignition inertial confinement fusion. Key metrics for these applications relate to absorption, yet conditions in this regime are so nonlinear that it is often impossible to know the fraction of absorbed light f, and even the range of f is unknown. In this presentation, using a relativistic Rankine-Hugoniot-like analysis, we show how to derive the theoretical maximum and minimum of f. These boundaries constrain nonlinear absorption mechanisms across the petawatt regime, forbidding high absorption values at low laser power and low absorption values at high laser power. Close agreement is shown with several dozens of published experimental data points and simulation results, helping to confirm the theory. For applications needing to circumvent the absorption bounds, these results will accelerate a shift from solid targets, towards structured and multilayer targets, and lead the development of new materials.

  18. Schistosome-induced cholangiocyte proliferation and osteopontin secretion correlate with fibrosis and portal hypertension in human and murine schistosomiasis mansoni

    PubMed Central

    Pereira, Thiago A.; Syn, Wing-Kin; Machado, Mariana V.; Vidigal, Paula V.; Resende, Vivian; Voieta, Izabela; Xie, Guanhua; Otoni, Alba; Souza, Márcia M.; Santos, Elisângela T.; Chan, Isaac S.; Trindade, Guilherme V.M.; Choi, Steve S.; Witek, Rafal P.; Pereira, Fausto E.; Secor, William E.; Andrade, Zilton A.; Lambertucci, José Roberto

    2015-01-01

    Schistosomiasis is a major cause of portal hypertension worldwide. It associates with portal fibrosis that develops during chronic infection. The mechanisms by which the pathogen evokes these host responses remain unclear. We evaluated the hypothesis that schistosome eggs release factors that directly stimulate liver cells to produce osteopontin (OPN), a pro-fibrogenic protein that stimulates hepatic stellate cells to become myofibroblasts. We also investigated the utility of OPN as a biomarker of fibrosis and/or severity of portal hypertension. Cultured cholangiocytes, Kupffer cells and hepatic stellate cells were treated with soluble egg antigen (SEA); OPN production was quantified by quantitative reverse transcriptase polymerase chain reaction (qRTPCR) and ELISA; cell proliferation was assessed by BrdU (5-bromo-2'-deoxyuridine). Mice were infected with Schistosoma mansoni for 6 or 16 weeks to cause early or advanced fibrosis. Liver OPN was evaluated by qRTPCR and immunohistochemistry (IHC) and correlated with liver fibrosis and serum OPN. Livers from patients with schistosomiasis mansoni (early fibrosis n=15; advanced fibrosis n=72) or healthy adults (n=22) were immunostained for OPN and fibrosis markers. Results were correlated with plasma OPN levels and splenic vein pressures. SEA-induced cholangiocyte proliferation and OPN secretion (P<0.001 compared with controls). Cholangiocytes were OPN (+) in Schistosoma-infected mice and humans. Liver and serum OPN levels correlated with fibrosis stage (mice: r=0.861; human r=0.672, P=0.0001) and myofibroblast accumulation (mice: r=0.800; human: r=0.761, P=0.0001). Numbers of OPN (+) bile ductules strongly correlated with splenic vein pressure (r=0.778; P=0.001). S. mansoni egg antigens stimulate cholangiocyte proliferation and OPN secretion. OPN levels in liver and blood correlate with fibrosis stage and portal hypertension severity. PMID:26201095

  19. The Osteopontin Level in Liver, Adipose Tissue and Serum Is Correlated with Fibrosis in Patients with Alcoholic Liver Disease

    PubMed Central

    Voican, Cosmin S.; Anty, Rodolphe; Saint-Paul, Marie-Christine; Rosenthal-Allieri, Maria-Alessandra; Agostini, Hélène; Njike, Micheline; Barri-Ova, Nadége; Naveau, Sylvie; Le Marchand-Brustel, Yannick; Veillon, Pascal; Calès, Paul; Perlemuter, Gabriel; Tran, Albert; Gual, Philippe

    2012-01-01

    Background Osteopontin (OPN) plays an important role in the progression of chronic liver diseases. We aimed to quantify the liver, adipose tissue and serum levels of OPN in heavy alcohol drinkers and to compare them with the histological severity of hepatic inflammation and fibrosis. Methodology/Principal Findings OPN was evaluated in the serum of a retrospective and prospective group of 109 and 95 heavy alcohol drinkers, respectively, in the liver of 34 patients from the retrospective group, and in the liver and adipose tissue from an additional group of 38 heavy alcohol drinkers. Serum levels of OPN increased slightly with hepatic inflammation and progressively with the severity of hepatic fibrosis. Hepatic OPN expression correlated with hepatic inflammation, fibrosis, TGFβ expression, neutrophils accumulation and with the serum OPN level. Interestingly, adipose tissue OPN expression also correlated with hepatic fibrosis even after 7 days of alcohol abstinence. The elevated serum OPN level was an independent risk factor in estimating significant (F≥2) fibrosis in a model combining alkaline phosphatase, albumin, hemoglobin, OPN and FibroMeter® levels. OPN had an area under the receiving operator curve that estimated significant fibrosis of 0.89 and 0.88 in the retrospective and prospective groups, respectively. OPN, Hyaluronate (AUROC: 0.88), total Cytokeratin 18 (AUROC: 0.83) and FibroMeter® (AUROC: 0.90) estimated significance to the same extent in the retrospective group. Finally, the serum OPN levels also correlated with hepatic fibrosis and estimated significant (F≥2) fibrosis in 86 patients with chronic hepatitis C, which suggested that its elevated level could be a general response to chronic liver injury. Conclusion/Significance OPN increased in the liver, adipose tissue and serum with liver fibrosis in alcoholic patients. Further, OPN is a new relevant biomarker for significant liver fibrosis. OPN could thus be an important actor in the

  20. Role of hepatitis C virus induced osteopontin in epithelial to mesenchymal transition, migration and invasion of hepatocytes.

    PubMed

    Iqbal, Jawed; McRae, Steven; Mai, Thi; Banaudha, Krishna; Sarkar-Dutta, Mehuli; Waris, Gulam

    2014-01-01

    Osteopontin (OPN) is a secreted phosphoprotein which has been linked to tumor progression and metastasis in a variety of cancers including hepatocellular carcinoma (HCC). Previous studies have shown that OPN is upregulated during liver injury and inflammation. However, the role of OPN in hepatitis C virus (HCV)-induced liver disease pathogenesis is not known. In this study, we determined the induction of OPN, and then investigated the effect of secreted forms of OPN in epithelial to mesenchymal transition (EMT), migration and invasion of hepatocytes. We show the induction of OPN mRNA and protein expression by HCV-infection. Our results also demonstrate the processing of precursor OPN (75 kDa) into 55 kDa, 42 kDa and 36 kDa forms of OPN in HCV-infected cells. Furthermore, we show the binding of secreted OPN to integrin αVβ3 and CD44 at the cell surface, leading to the activation of downstream cellular kinases such as focal adhesion kinase (FAK), Src, and Akt. Importantly, our results show the reduced expression of epithelial marker (E-cadherin) and induction of mesenchymal marker (N-cadherin) in HCV-infected cells. We also show the migration and invasion of HCV-infected cells using wound healing assay and matrigel coated Boyden chamber. In addition, we demonstrate the activation of above EMT markers, and the critical players involved in OPN-mediated cell signaling cascade using primary human hepatocytes infected with Japanese fulminant hepatitis (JFH)-1 HCV. Taken together, these studies suggest a potential role of OPN in inducing chronic liver disease and HCC associated with chronic HCV infection. PMID:24498111

  1. Osteopontin (OPN) Is an Important Protein to Mediate Improvements in the Biocompatibility of C Ion-Implanted Silicone Rubber

    PubMed Central

    Zhang, Yi-ming; Shen, Li-ru; Lei, Ze-yuan; Zhang, Zhi-qing; Cao, Cong; Fan, Dong-li

    2014-01-01

    Medical device implants are drawing increasing amounts of interest from modern medical practitioners. However, this attention is not evenly spread across all such devices; most of these implantable devices can cause adverse reactions such as inflammation, fibrosis, thrombosis, and infection. In this work, the biocompatibility of silicone rubber (SR) was improved through carbon (C) ion implantation. Scanning electron microscopy (SEM), atomic force microscopy (AFM), X-ray photoelectron spectroscopy (XPS), and X-ray diffraction (XRD) results confirmed that these newly generated carbon-implanted silicone rubbers (C-SRs) had large, irregular peaks and deep valleys on their surfaces. The water contact angle of the SR surface decreased significantly after C ion implantation. C ion implantation also changed the surface charge distribution, silicone oxygen rate, and chemical-element distribution of SR to favor cell attachment. The dermal fibroblasts cultured on the surface C-SR grew faster and showed more typical fibroblastic shapes. The expression levels of major adhesion proteins, including talin-1, zyxin, and vinculin, were significantly higher in dermal fibroblasts cultured on C-SR coated plates than in dermal fibroblasts cultured on SR. Those same dermal fibroblasts on C-SRs showed more pronounced adhesion and migration abilities. Osteopontin (OPN), a critical extracellular matrix (ECM) protein, was up-regulated and secreted from dermal fibroblasts cultured on C-SR. Matrix metalloproteinase-9 (MMP-9) activity was also increased. These cells were highly mobile and were able to adhere to surfaces, but these abilities were inhibited by the monoclonal antibody against OPN, or by shRNA-mediated MMP-9 knockdown. Together, these results suggest that C ion implantation significantly improves SR biocompatibility, and that OPN is important to promote cell adhesion to the C-SR surface. PMID:24911051

  2. Osteopontin enhances HIV replication and is increased in the brain and cerebrospinal fluid of HIV-infected individuals

    PubMed Central

    Islam, Tanzeem; Adams, Robert; Nerle, Sujata; Kamara, Masiray; Eger, Caitlin; Marder, Karen; Cohen, Bruce; Schifitto, Giovanni; McArthur, Justin C.; Sacktor, Ned; Pardo, Carlos A.

    2012-01-01

    Despite effective and widely available suppressive anti-HIV therapy, the prevalence of mild neurocognitive dysfunction continues to increase. HIV-associated neurocognitive disorder (HAND) is a multifactorial disease with sustained central nervous system inflammation and immune activation as prominent features. Inflammatory macrophages, HIV-infected and uninfected, play a central role in the development of HIV dementia. There is a critical need to identify biomarkers and to better understand the molecular mechanisms leading to cognitive dysfunction in HAND. In this regard, we identified through a subtractive hybridization strategy osteopontin (OPN, SPP1, gene) an inflammatory marker, as an upregulated gene in HIV-infected primary human monocyte-derived macrophages. Knockdown of OPN in primary macrophages resulted in a threefold decrease in HIV-1 replication. Ectopic expression of OPN in the TZM-bl cell line significantly enhanced HIV infectivity and replication. A significant increase in the degradation of the NF-κB inhibitor, IκBα and an increase in the nuclear-to-cytoplasmic ratio of NF-κB were found in HIV-infected cells expressing OPN compared to controls. Moreover, mutation of the NF-κB binding domain in the HIV-LTR abrogated enhanced promoter activity stimulated by OPN. Interestingly, compared to cerebrospinal fluid from normal and multiple sclerosis controls, OPN levels were significantly higher in HIV-infected individuals both with and without neurocognitive disorder. OPN levels were highest in HIV-infected individuals with moderate to severe cognitive impairment. Moreover, OPN was significantly elevated in brain tissue from HIV-infected individuals with cognitive disorder versus those without impairment. Collectively, these data suggest that OPN stimulates HIV-1 replication and that high levels of OPN are present in the CNS compartment of HIV-infected individuals, reflecting ongoing inflammatory processes at this site despite anti-HIV therapy. PMID

  3. Effect of otoconial proteins fetuin A, osteopontin, and otoconin 90 on the nucleation and growth of calcite

    SciTech Connect

    Hong, Mina; Moreland, K. Trent; Chen, Jiajun; Teng, Henry H.; Thalmann, Ruediger; De Yoreo, James J.

    2014-10-30

    Here, we investigated the roles of three proteins associated with the formation of otoconia including fetuin A, osteopontin (OPN), and otoconin 90 (OC90). In situ atomic force microscopy (AFM) studies of the effects of these proteins on the growth of atomic steps on calcite surfaces were performed to obtain insight into their effects on the growth kinetics. We also used scanning electron microscopy to examine the effects of these proteins on crystal morphology. All three proteins were found to be potent inhibitors of calcite growth, although fetuin A promoted growth at concentrations below about 40 nM and only became an inhibitor at higher concentrations. We then used in situ optical microscopy to observe calcite nucleation on films of these proteins adsorbed onto mica surfaces. By measuring the calcite nucleation rate as a function of supersaturation, the value of the interfacial energy that controls the free energy barrier to heterogeneous nucleation was determined for each protein. OPN and OC90 films led to significantly reduced interfacial energies as compared to the value for homogeneous calcite nucleation in bulk solution. The value for fetuin A was equal to that for bulk solution within experimental error. Zeta potential measurements showed all of the proteins possessed negative surface charge and varied in magnitude according to sequence fetuin A > OC90 > OPN. In addition, the interfacial energies exhibited an inverse scaling with the zeta potential. In analogy to previous measurements on polysaccharide films, this scaling indicates the differences between the proteins arise from the effect of protein surface charge on the solution–substrate interfacial energy.

  4. Effect of otoconial proteins fetuin A, osteopontin, and otoconin 90 on the nucleation and growth of calcite

    DOE PAGESBeta

    Hong, Mina; Moreland, K. Trent; Chen, Jiajun; Teng, Henry H.; Thalmann, Ruediger; De Yoreo, James J.

    2014-10-30

    Here, we investigated the roles of three proteins associated with the formation of otoconia including fetuin A, osteopontin (OPN), and otoconin 90 (OC90). In situ atomic force microscopy (AFM) studies of the effects of these proteins on the growth of atomic steps on calcite surfaces were performed to obtain insight into their effects on the growth kinetics. We also used scanning electron microscopy to examine the effects of these proteins on crystal morphology. All three proteins were found to be potent inhibitors of calcite growth, although fetuin A promoted growth at concentrations below about 40 nM and only became anmore » inhibitor at higher concentrations. We then used in situ optical microscopy to observe calcite nucleation on films of these proteins adsorbed onto mica surfaces. By measuring the calcite nucleation rate as a function of supersaturation, the value of the interfacial energy that controls the free energy barrier to heterogeneous nucleation was determined for each protein. OPN and OC90 films led to significantly reduced interfacial energies as compared to the value for homogeneous calcite nucleation in bulk solution. The value for fetuin A was equal to that for bulk solution within experimental error. Zeta potential measurements showed all of the proteins possessed negative surface charge and varied in magnitude according to sequence fetuin A > OC90 > OPN. In addition, the interfacial energies exhibited an inverse scaling with the zeta potential. In analogy to previous measurements on polysaccharide films, this scaling indicates the differences between the proteins arise from the effect of protein surface charge on the solution–substrate interfacial energy.« less

  5. Mechanisms of Osteopontin-Induced Stabilization of Blood-Brain Barrier Disruption After Subarachnoid Hemorrhage in Rats

    PubMed Central

    Suzuki, Hidenori; Hasegawa, Yu; Kanamaru, Kenji; Zhang, John H.

    2010-01-01

    Background and Purpose Osteopontin (OPN) is an inducible multifunctional extracellular matrix protein that may be protective against blood-brain barrier (BBB) disruption after subarachnoid hemorrhage (SAH). However, the protective mechanisms remain unclear. Methods We produced the endovascular perforation model of SAH in rats and studied the time course of OPN induction in brains using Western blotting and immunofluorescence (n=50). Then, 34 rats were randomly assigned to sham (n=3), sham+OPN small interfering RNA (siRNA; n=3), SAH+negative control siRNA (n=14) and SAH+OPN siRNA (n=14) groups, and 109 rats to sham+vehicle (n=17), sham+recombinant OPN (r-OPN; n=17), SAH+vehicle (n=33), SAH+r-OPN (n=31) and SAH+r-OPN+GRGDSP (L-arginyl-glycyl-L-aspartate motif-containing hexapeptide; n=11) groups. Effects of OPN siRNA or r-OPN on BBB disruption and the related proteins were studied. Results OPN was significantly induced in the reactive astrocytes and capillary endothelial cells, peaking at 72 hours post-SAH, during the recovery phase of BBB disruption. Blockage of endogenous OPN induction exacerbated BBB disruption associated with reduction of angiopoietin-1 and mitogen-activated protein kinase (MAPK) phosphatase-1 (an endogenous MAPK inhibitor), activation of MAPKs and induction of vascular endothelial growth factor (VEGF)-A at 72 hours post-SAH, while r-OPN treatment improved it associated with MKP-1 induction, MAPK inactivation and VEGF-A reduction, which was blocked by GRGDSP at 24 hours post-SAH. VEGF-B and angiopoietin-2 levels were unchanged. Conclusions OPN may increase MKP-1 that inactivates MAPKs, upstream and downstream of VEGF-A, via binding to L-arginyl-glycyl-L-aspartate-dependent integrin receptors, suggesting a novel mechanism of OPN-induced post-SAH BBB protection. PMID:20616319

  6. Thrombin-cleaved Fragments of Osteopontin Are Overexpressed in Malignant Glial Tumors and Provide a Molecular Niche with Survival Advantage*

    PubMed Central

    Yamaguchi, Yasuto; Shao, Zhifei; Sharif, Shadi; Du, Xiao-Yan; Myles, Timothy; Merchant, Milton; Harsh, Griffith; Glantz, Michael; Recht, Lawrence; Morser, John; Leung, Lawrence L. K.

    2013-01-01

    Osteopontin (OPN), which is highly expressed in malignant glioblastoma (GBM), possesses inflammatory activity modulated by proteolytic cleavage by thrombin and plasma carboxypeptidase B2 (CPB2) at a highly conserved cleavage site. Full-length OPN (OPN-FL) was elevated in cerebrospinal fluid (CSF) samples from all cancer patients compared with noncancer patients. However, thrombin-cleaved OPN (OPN-R) and thrombin/CPB2-double-cleaved OPN (OPN-L) levels were markedly increased in GBM and non-GBM gliomas compared with systemic cancer and noncancer patients. Cleaved OPN constituted ∼23 and ∼31% of the total OPN in the GBM and non-GBM CSF samples, respectively. OPN-R was also elevated in GBM tissues. Thrombin-antithrombin levels were highly correlated with cleaved OPN, but not OPN-FL, suggesting that the cleaved OPN fragments resulted from increased thrombin and CPB2 in this extracellular compartment. Levels of VEGF and CCL4 were increased in CSF of GBM and correlated with the levels of cleaved OPN. GBM cell lines were more adherent to OPN-R and OPN-L than OPN-FL. Adhesion to OPN altered gene expression, in particular genes involved with cellular processes, cell cycle regulation, death, and inflammation. OPN and its cleaved forms promoted motility of U-87 MG cells and conferred resistance to apoptosis. Although functional mutation of the RGD motif in OPN largely abolished these functions, OPNRAA-R regained significant cell binding and signaling function, suggesting that the SVVYGLR motif in OPN-R may substitute for the RGD motif if the latter becomes inaccessible. OPN cleavage contributes to GBM development by allowing more cells to bind in niches where they acquire anti-apoptotic properties. PMID:23204518

  7. Roles of osteopontin and matrix metalloproteinase-7 in occurrence, progression, and prognosis of nonsmall cell lung cancer

    PubMed Central

    Sun, Ying; Li, Dan; Lv, Xiao-Hong; Hua, Shu-Cheng; Han, Ji-Chang; Xu, Feng; Li, Xian-Dong

    2015-01-01

    Background: This study detected osteopontin (OPN) and matrix metalloproteinase-7 (MMP-7) expressions to explore the roles of OPN and MMP-7 in the occurrence, progression, and prognosis of nonsmall cell lung cancer (NSCLC). Materials and Methods: A retrospective study was conducted on NSCLC tissues (n = 152; case group) and adjacent nonneoplastic lung parenchyma (adjacent to tumor >5 cm; n = 152; control group) collected from 152 NSCLC patients. The protein expressions of OPN and MMP-7 were detected by immunohistochemistry. OPN and MMP-7 messenger RNA (mRNA) expressions were detected by reverse transcription polymerase chain reaction (RT-PCR). Results: The protein and mRNA expressions of OPN and MMP-7 in NSCLC tissues were evidently higher than those in adjacent nonneoplastic lung parenchyma (all P < 0.05). OPN protein and mRNA expression were associated with the degree of differentiation, tumor node metastasis (TNM) staging, and lymph node metastasis in NSCLC (all P < 0.05). MMP-7 protein expression was associated with TNM staging and lymph node metastasis (both P < 0.05) while MMP-7 mRNA expression was associated with the degree of differentiation, TNM staging, and lymph node metastasis (all P < 0.05). A significantly positive relativity was revealed between OPN expression and MMP-7 expression (protein: r = 0.789, P < 0.001; mRNA: r = 0.377, P < 0.001). Lymph node metastasis, TNM staging, OPN, and MMP-7 protein expressions were independent risk factors for the prognosis of NSCLC (all P < 0.05). Conclusion: High MMP-7 and OPN protein expressions are closely related to the occurrence, progression, and prognosis of NSCLC, and can be served as unfavorable prognostic factors for NSCLC. PMID:26958047

  8. Rat Urinary Osteopontin and Neutrophil Gelatinase-Associated Lipocalin Improve Certainty of Detecting Drug-Induced Kidney Injury.

    PubMed

    Phillips, Jonathan A; Holder, Daniel J; Ennulat, Daniela; Gautier, Jean-Charles; Sauer, John-Michael; Yang, Yi; McDuffie, Eric; Sonee, Manisha; Gu, Yi-Zhong; Troth, Sean P; Lynch, Karen; Hamlin, Diane; Peters, David G; Brees, Dominique; Walker, Elizabeth G

    2016-06-01

    Traditional kidney biomarkers are insensitive indicators of acute kidney injury, with meaningful changes occurring late in the course of injury. The aim of this work was to demonstrate the diagnostic potential of urinary osteopontin (OPN) and neutrophil gelatinase-associated lipocalin (NGAL) for drug-induced kidney injury (DIKI) in rats using data from a recent regulatory qualification submission of translational DIKI biomarkers and to compare performance of NGAL and OPN to five previously qualified DIKI urinary biomarkers. Data were compiled from 15 studies of 11 different pharmaceuticals contributed by Critical Path Institute's Predictive Safety Testing Consortium (PSTC) Nephrotoxicity Working Group (NWG). Rats were given doses known to cause DIKI or other target organ toxicity, and urinary levels of the candidate biomarkers were assessed relative to kidney histopathology and serum creatinine (sCr) and blood urea nitrogen (BUN).OPN and NGAL outperformed sCr and BUN in identifying DIKI manifested as renal tubular epithelial degeneration or necrosis. In addition, urinary OPN and NGAL, when used with sCr and BUN, increased the ability to detect renal tubular epithelial degeneration or necrosis. NGAL and OPN had comparable or improved performance relative to Kim-1, clusterin, albumin, total protein, and beta-2 microglobulin. Given these data, both urinary OPN and NGAL are appropriate for use with current methods for assessing nephrotoxicity to identify and monitor DIKI in regulatory toxicology studies in rats. These data also support exploratory use of urinary OPN and NGAL in safety monitoring strategies of early clinical trials to aid in the assurance of patient safety. PMID:27026710

  9. Effect of Otoconial Proteins Fetuin A, Osteopontin, and Otoconin 90 on the Nucleation and Growth of Calcite

    PubMed Central

    2015-01-01

    We investigated the roles of three proteins associated with the formation of otoconia including fetuin A, osteopontin (OPN), and otoconin 90 (OC90). In situ atomic force microscopy (AFM) studies of the effects of these proteins on the growth of atomic steps on calcite surfaces were performed to obtain insight into their effects on the growth kinetics. We also used scanning electron microscopy to examine the effects of these proteins on crystal morphology. All three proteins were found to be potent inhibitors of calcite growth, although fetuin A promoted growth at concentrations below about 40 nM and only became an inhibitor at higher concentrations. We then used in situ optical microscopy to observe calcite nucleation on films of these proteins adsorbed onto mica surfaces. By measuring the calcite nucleation rate as a function of supersaturation, the value of the interfacial energy that controls the free energy barrier to heterogeneous nucleation was determined for each protein. OPN and OC90 films led to significantly reduced interfacial energies as compared to the value for homogeneous calcite nucleation in bulk solution. The value for fetuin A was equal to that for bulk solution within experimental error. Zeta potential measurements showed all of the proteins possessed negative surface charge and varied in magnitude according to sequence fetuin A > OC90 > OPN. In addition, the interfacial energies exhibited an inverse scaling with the zeta potential. In analogy to previous measurements on polysaccharide films, this scaling indicates the differences between the proteins arise from the effect of protein surface charge on the solution–substrate interfacial energy. PMID:25709560

  10. Bounds for Asian basket options

    NASA Astrophysics Data System (ADS)

    Deelstra, Griselda; Diallo, Ibrahima; Vanmaele, Michèle

    2008-09-01

    In this paper we propose pricing bounds for European-style discrete arithmetic Asian basket options in a Black and Scholes framework. We start from methods used for basket options and Asian options. First, we use the general approach for deriving upper and lower bounds for stop-loss premia of sums of non-independent random variables as in Kaas et al. [Upper and lower bounds for sums of random variables, Insurance Math. Econom. 27 (2000) 151-168] or Dhaene et al. [The concept of comonotonicity in actuarial science and finance: theory, Insurance Math. Econom. 31(1) (2002) 3-33]. We generalize the methods in Deelstra et al. [Pricing of arithmetic basket options by conditioning, Insurance Math. Econom. 34 (2004) 55-57] and Vanmaele et al. [Bounds for the price of discrete sampled arithmetic Asian options, J. Comput. Appl. Math. 185(1) (2006) 51-90]. Afterwards we show how to derive an analytical closed-form expression for a lower bound in the non-comonotonic case. Finally, we derive upper bounds for Asian basket options by applying techniques as in Thompson [Fast narrow bounds on the value of Asian options, Working Paper, University of Cambridge, 1999] and Lord [Partially exact and bounded approximations for arithmetic Asian options, J. Comput. Finance 10 (2) (2006) 1-52]. Numerical results are included and on the basis of our numerical tests, we explain which method we recommend depending on moneyness and time-to-maturity.

  11. Northwest Outward Bound Instructor's Manual.

    ERIC Educational Resources Information Center

    Northwest Outward Bound School, Portland, OR.

    Instructor responsibilities, procedures for completing activities safely, and instructional methods and techniques are outlined to assist instructors in the Northwest Outward Bound School (Portland, Oregon) as they strive for teaching excellence. Information is organized into six chapters addressing: history and philosophy of Outward Bound; course…

  12. Isocurvature bounds on axions revisited

    SciTech Connect

    Beltran, Maria; Garcia-Bellido, Juan; Lesgourgues, Julien

    2007-05-15

    The axion is one of the best motivated candidates for particle dark matter. We study and update the constraints imposed by the recent CMB and LSS experiments on the mass of axions produced by the misalignment mechanism, as a function of both the inflationary scale and the reheating temperature. Under some particular although not unconventional assumptions, the axion induces isocurvature perturbations with an amplitude too large to be compatible with observations. Specifically, for inflation taking place at intermediate energy scales, we derive some restrictive limits which can only be evaded by assuming an efficient reheating mechanism, with T{sub rh}>10{sup 11} GeV. Chaotic inflation with a quadratic potential is still compatible with the axion scenario, provided that the Peccei-Quinn scale f{sub a} is close to 10{sup 10} or 10{sup 11} GeV. Isocurvature bounds eliminate the possibility of a larger f{sub a} and a small misalignment angle. We find that isocurvature constraints on the axion scenario must be taken into account whenever the scale of inflation is above 10{sup 12} GeV; below this scale, axionic isocurvature modes are too small to be probed by current observations.

  13. Saturating the holographic entropy bound

    SciTech Connect

    Bousso, Raphael; Freivogel, Ben; Leichenauer, Stefan

    2010-10-15

    The covariant entropy bound states that the entropy, S, of matter on a light sheet cannot exceed a quarter of its initial area, A, in Planck units. The gravitational entropy of black holes saturates this inequality. The entropy of matter systems, however, falls short of saturating the bound in known examples. This puzzling gap has led to speculation that a much stronger bound, S < or approx. A{sup 3/4}, may hold true. In this note, we exhibit light sheets whose entropy exceeds A{sup 3/4} by arbitrarily large factors. In open Friedmann-Robertson-Walker universes, such light sheets contain the entropy visible in the sky; in the limit of early curvature domination, the covariant bound can be saturated but not violated. As a corollary, we find that the maximum observable matter and radiation entropy in universes with positive (negative) cosmological constant is of order {Lambda}{sup -1} ({Lambda}{sup -2}), and not |{Lambda}|{sup -3/4} as had hitherto been believed. Our results strengthen the evidence for the covariant entropy bound, while showing that the stronger bound S < or approx. A{sup 3/4} is not universally valid. We conjecture that the stronger bound does hold for static, weakly gravitating systems.

  14. Regulation of collagenase-3 and osteocalcin gene expression by collagen and osteopontin in differentiating MC3T3-E1 cells

    NASA Technical Reports Server (NTRS)

    D'Alonzo, Richard C.; Kowalski, Aaron J.; Denhardt, David T.; Nickols, G. Allen; Partridge, Nicola C.

    2002-01-01

    Both collagenase-3 and osteocalcin mRNAs are expressed maximally during the later stages of osteoblast differentiation. Here, we demonstrate that collagenase-3 mRNA expression in differentiating MC3T3-E1 cells is dependent upon the presence of ascorbic acid, is inhibited in the presence of the collagen synthesis inhibitor, 3,4-dehydroproline, and is stimulated by growth on collagen in the absence of ascorbic acid. Transient transfection studies show that collagenase-3 promoter activity increases during cell differentiation and requires the presence of ascorbic acid. Additionally, we show that, in differentiating MC3T3-E1 cells, collagenase-3 gene expression increases in the presence of an anti-osteopontin monoclonal antibody that binds near the RGD motif of this protein, whereas osteocalcin expression is inhibited. Furthermore, an RGD peptidomimetic compound, designed to block interaction of ligands to the alpha(v) integrin subunit, increases osteocalcin expression and inhibits collagenase-3 expression, suggesting that the RGD peptidomimetic initiates certain alpha(v) integrin signaling in osteoblastic cells. Overall, these studies demonstrate that stimulation of collagenase-3 expression during osteoblast differentiation requires synthesis of a collagenous matrix and that osteopontin and alpha(v) integrins exert divergent regulation of collagenase-3 and osteocalcin expression during osteoblast differentiation.

  15. Toll-like receptors 2 and 4 impair insulin-mediated brain activity by interleukin-6 and osteopontin and alter sleep architecture.

    PubMed

    Sartorius, Tina; Lutz, Stefan Z; Hoene, Miriam; Waak, Jens; Peter, Andreas; Weigert, Cora; Rammensee, Hans-Georg; Kahle, Philipp J; Häring, Hans-Ulrich; Hennige, Anita M

    2012-05-01

    Impaired insulin action in the brain represents an early step in the progression toward type 2 diabetes, and elevated levels of saturated free fatty acids are known to impair insulin action in prediabetic subjects. One potential mediator that links fatty acids to inflammation and insulin resistance is the Toll-like receptor (TLR) family. Therefore, C3H/HeJ/TLR2-KO (TLR2/4-deficient) mice were fed a high-fat diet (HFD), and insulin action in the brain as well as cortical and locomotor activity was analyzed by using telemetric implants. TLR2/4-deficient mice were protected from HFD-induced glucose intolerance and insulin resistance in the brain and displayed an improvement in cortical and locomotor activity that was not observed in C3H/HeJ mice. Sleep recordings revealed a 42% increase in rapid eye movement sleep in the deficient mice during daytime, and these mice spent 41% more time awake during the night period. Treatment of control mice with a neutralizing IL-6 antibody improved insulin action in the brain as well as cortical activity and diminished osteopontin protein to levels of the TLR2/4-deficient mice. Together, our data suggest that the lack of functional TLR2/4 protects mice from a fat-mediated impairment in insulin action, brain activity, locomotion, and sleep architecture by an IL-6/osteopontin-dependent mechanism. PMID:22278939

  16. Osteopontin (OPN/SPP1) isoforms collectively enhance tumor cell invasion and dissemination in esophageal adenocarcinoma

    PubMed Central

    Lin, Jules; Myers, Amy L.; Wang, Zhuwen; Nancarrow, Derek J.; Ferrer-Torres, Daysha; Handlogten, Amy; Leverenz, Kimmy; Bao, Julia; Thomas, Dafydd G.; Wang, Thomas D.; Orringer, Mark B.; Reddy, Rishindra M.; Chang, Andrew C.; Beer, David G.; Lin, Lin

    2015-01-01

    Esophageal adenocarcinoma (EAC) is often diagnosed at an advanced stage, thus understanding the molecular basis for EAC invasion and metastasis is critical. Here we report that SPP1/OPN was highly overexpressed in primary EACs and intracellularly localized to tumor cells. We further demonstrate that all known OPN isoforms (OPNa, b, c, 4 and 5) were frequently co-overexpressed in primary EACs. Distinct pro-invasion and dissemination phenotypes of isoform-specific OPNb and OPNc stable transfectants were observed. Expression of OPNb significantly enhanced cell migration and adhesion to laminin. In contrast, OPNc cells showed significantly decreased cell migration yet increased cell detachment. Enhanced invasion, both in vitro and in vivo, was observed for OPNb- but not OPNc-expressing cells. Inhibition of RGD integrins, one family of OPN receptors, attenuated OPNb cell migration, abrogated OPNb cell adhesion and significantly reduced OPNb cell clonogenic survival but did not affect OPNc phenotypes, indicating that OPNb but not OPNc acts through integrin-dependent signaling. Differential expression of vimentin, E-cadherin and β-catenin in OPN stable cells may account for the variation in cell adhesion and detachment between these isoforms. We conclude that while all OPN isoforms are frequently co-overexpressed in primary EACs, isoforms OPNb and OPNc enhance invasion and dissemination through collective yet distinct mechanisms. PMID:26068949

  17. Tsirelson's bound from a generalized data processing inequality

    NASA Astrophysics Data System (ADS)

    Dahlsten, Oscar C. O.; Lercher, Daniel; Renner, Renato

    2012-06-01

    The strength of quantum correlations is bounded from above by Tsirelson's bound. We establish a connection between this bound and the fact that correlations between two systems cannot increase under local operations, a property known as the data processing inequality (DPI). More specifically, we consider arbitrary convex probabilistic theories. These can be equipped with an entropy measure that naturally generalizes the von Neumann entropy, as shown recently in Short and Wehner (2010 New J. Phys. 12 033023) and Barnum et al (2010 New J. Phys. 12 033024). We prove that if the DPI holds with respect to this generalized entropy measure then the underlying theory necessarily respects Tsirelson's bound. We, moreover, generalize this statement to any entropy measure satisfying certain minimal requirements. A consequence of our result is that not all the entropic relations used for deriving Tsirelson's bound via information causality in Pawlowski et al (2009 Nature 461 1101-4) are necessary.

  18. Bounding the elliptic Mahler measure

    NASA Astrophysics Data System (ADS)

    Pinner, Christopher

    1998-11-01

    We give a simple inequality relating the elliptic Mahler measure of a polynomial to the traditional Mahler measure (via the length of the polynomial). These bounds are essentially sharp. We also give the corresponding result for polynomials in several variables.

  19. Crystal Structures of Beryllium Fluoride-Free and Beryllium Fluoride-Bound CheY in Complex with the Conserved C-Terminal Peptide of CheZ Reveal Dual Binding Modes Specific to CheY Conformation

    SciTech Connect

    Guhaniyogi,J.; Robinson, V.; Stock, A.

    2006-01-01

    Chemotaxis, the environment-specific swimming behavior of a bacterial cell is controlled by flagellar rotation. The steady-state level of the phosphorylated or activated form of the response regulator CheY dictates the direction of flagellar rotation. CheY phosphorylation is regulated by a fine equilibrium of three phosphotransfer activities: phosphorylation by the kinase CheA, its auto-dephosphorylation and dephosphorylation by its phosphatase CheZ. Efficient dephosphorylation of CheY by CheZ requires two spatially distinct protein-protein contacts: tethering of the two proteins to each other and formation of an active site for dephosphorylation. The former involves interaction of phosphorylated CheY with the small highly conserved C-terminal helix of CheZ (CheZ{sub C}), an indispensable structural component of the functional CheZ protein. To understand how the CheZ{sub C} helix, representing less than 10% of the full-length protein, ascertains molecular specificity of binding to CheY, we have determined crystal structures of CheY in complex with a synthetic peptide corresponding to 15 C-terminal residues of CheZ (CheZ{sub 200-214}) at resolutions ranging from 2.0 Angstroms to 2.3 Angstroms. These structures provide a detailed view of the CheZC peptide interaction both in the presence and absence of the phosphoryl analog, BeF{sub 3}{sup -}. Our studies reveal that two different modes of binding the CheZ{sub 200-214} peptide are dictated by the conformational state of CheY in the complex. Our structures suggest that the CheZ{sub C} helix binds to a 'meta-active' conformation of inactive CheY and it does so in an orientation that is distinct from the one in which it binds activated CheY. Our dual binding mode hypothesis provides implications for reverse information flow in CheY and extends previous observations on inherent resilience in CheY-like signaling domains.

  20. In photosynthetic reaction centers, the free energy difference for electron transfer between quinones bound at the primary and secondary quinone-binding sites governs the observed secondary site specificity.

    PubMed Central

    Giangiacomo, K M; Dutton, P L

    1989-01-01

    The secondary quinone-binding site (QB site) of bacterial reaction centers from Rhodobacter sphaeroides is generally regarded to be highly specific for its native ubiquinone-10 molecule. We demonstrate here that this is a misconception rooted in the kinetic methods used to assay for occupancy of a quinone in the QB site. We show that observance of occupancy of the QB site, revealed by kinetic assay, is sensitive to the free-energy difference for electron transfer between the quinone at the primary quinone-binding site (QA site) and the QB site (-delta G0e-). For many of the compounds previously tested for binding at the QB site, the -delta G0e- between QA and QB is too small to permit detection of the functional quinone in the QB site. With an increased -delta G0e- achieved by replacing the native ubiquinone-10 at the QA site with lower-potential quinones or by testing higher-potential QB candidates, it is shown that the QB site binds and functions with the unsubstituted 1,4-benzoquinone, 1,4-naphthoquinone, and 9,10-phenanthraquinone, as well as with their various substituted forms. Moreover, quinones with the ortho-carbonyl configuration appear to function in a similar manner to quinones with the para-carbonyl configuration. PMID:2649889

  1. Human osteoclast and giant cell differentiation: the apparent switch from nonspecific esterase to tartrate resistant acid phosphatase activity coincides with the in situ expression of osteopontin mRNA.

    PubMed

    Connor, J R; Dodds, R A; James, I E; Gowen, M

    1995-12-01

    Animal model and in vitro cultures suggest that osteoclasts and cells of the mononuclear phagocyte system share a common precursor. However, the human osteoclast precursor has not been positively identified. We attempted to identify the precursor in situ by using a number of osteoclast- and macrophage-selective markers, together with the expression of osteopontin mRNA, previously shown to be abundant in human osteoclasts. Sections of osteophytic bone and a panel of inflammatory connective tissues were processed for in situ hybridization; serial sections were analyzed for tartrate-resistant acid phosphatase (TRAP) and nonspecific esterase (NSE) activity, selective cytochemical markers for the osteoclast and cells of the macrophage/monocyte lineage, respectively. The murine anti-human osteoclast monoclonal antibodies 23C6 (vitronectin receptor) and C35 (osteoclast-selective) were used to further identify the osteoclast phenotype. We compared osteoclasts, giant cells, and their respective putative mononuclear precursors. At resorption sites within osteophytic bone, osteopontin mRNA was expressed in osteoclasts and a distinct population of TRAP+, NSE- mononuclear cells. Adjacent clusters of mononuclear cells were TRAP- and NSE+ or were active for both enzymes; these cells demonstrated variable expression of osteopontin mRNA. In the inflammatory connective tissues, abundant macrophage-like cells (NSE+/TRAP-) did not express osteopontin mRNA. However, TRAP+ mononuclear cells observed among clusters of NSE+ cells did express osteopontin mRNA. At these sites, clusters of putative macrophage polykaryons removing fragments of bone debris were observed. These giant cells and associated mononuclear cells were NSE- and distinctly TRAP+, and expressed osteopontin mRNA, C35, and 23C6 (human osteoclast) reactivity. Therefore, cells involved in the remodeling (resorption) of bone or the removal of bone debris, together with their immediate precursors, switch from being NSE

  2. Hybrid Homology Modeling and Mutational Analysis of Cytochrome P450C24A1 (CYP24A1) of the Vitamin D Pathway: Insights into Substrate Specificity and Membrane Bound Structure-Function

    PubMed Central

    Annalora, Andrew J.; Bobrovnikova-Marjon, Ekaterina; Serda, Rita; Pastuszyn, Andrzej; Graham, Sandra E.; Marcus, Craig B.; Omdahl, John L.

    2007-01-01

    Cytochrome P450C24A1 (CYP24A1), a peripheral inner mitochondrial membrane hemoprotein and candidate oncogene, regulates the side-chain metabolism and biological function of vitamin D and many of its related analog drugs. Rational mutational analysis of rat CYP24A1 based on hybrid (2C5/BM-3) homology modeling and affinity labeling studies clarified the role of key domains (N-terminus, A', A, and F-helices, β3a strand, & β5 hairpin) in substrate binding and catalysis. The scope of our study was limited by an inability to purify stable mutant enzyme targeting soluble domains (B', G, and I-helices) and suggested greater conformational flexibility among CYP24A1's membrane-associated domains. The most notable mutants developed by modeling were V391T and I500A, which displayed defective binding function and profound metabolic defects for 25-hydroxylated vitamin D3 substrates similar to a non-functional F-helix mutant (F249T) that we previously reported. Val-391 (β3a strand) and Ile-500 (β5 hairpin) are modeled to interact with Phe-249 (F-helix) in a hydrophobic cluster that directs substrate binding events through interactions with the vitamin D cis-triene moiety. Prior affinity labeling studies identified an amino-terminal residue (Ser-57) as a putative active-site residue that interacts with the 3β-OH group of the vitamin D A-ring. Studies with 3-epi and 3-deoxy-1,25(OH)2D3 analogs confirmed interactions between the 3β-OH group and Ser-57 effect substrate recognition and trafficking while establishing that the trans conformation of A-ring hydroxyl groups (1α & 3β) is obligate for high-affinity binding to rat CYP24A1. Our work suggests that CYP24A1's amphipathic nature allows for monotopic membrane insertion, whereby a pw2d-like substrate access channel is formed to shuttle secosteroid substrate from the membrane to the active-site. We hypothesize that CYP24A1 has evolved a unique amino-terminal membrane binding motif that contributes to substrate specificity and

  3. Bound polarons in semiconductor nanostructures

    NASA Astrophysics Data System (ADS)

    Woggon, U.; Miller, D.; Kalina, F.; Gerlach, B.; Kayser, D.; Leonardi, K.; Hommel, D.

    2003-01-01

    Bound polarons are discrete, confined electronic states, spatially localized due to a local potential V(r) but sharing a common phonon state of the surrounding crystal. We study the energy states of polarons bound in a potential and determine the local optical absorption spectrum up to first-order time-dependent perturbation theory with respect to the electron-photon interaction. The model is applied to describe the optical properties of submonolayer CdSe insertions epitaxially grown between ZnSe layers. As a typical signature of bound polarons we found excited-state energies equidistantly separated by the LO phonon energy and with optical transition probabilities determined by the anisotropies in V(r).

  4. Unitarity bound for gluon shadowing

    SciTech Connect

    Kopeliovich, B. Z.; Levin, E.; Potashnikova, I. K.; Schmidt, Ivan

    2009-06-15

    Although at small Bjorken x gluons originated from different nucleons in a nucleus overlap in the longitudinal direction, most of them are still well separated in the transverse plane and therefore cannot fuse. For this reason the gluon density in nuclei cannot drop at small x below a certain bottom bound, which we evaluated in a model independent manner assuming the maximal strength of gluon fusion. We also calculated gluon shadowing in the saturated regime using the Balitsky-Kovchegov equation and found the nuclear ratio to be well above the unitarity bound. The recently updated analysis of parton distributions in nuclei, including BNL Relativistic Heavy Ion Collider (RHIC) data on high-p{sub T} hadron production at forward rapidities, led to strong gluon shadowing. Such strong shadowing and therefore the interpretation of the nuclear modification of the p{sub T} spectra in dA collisions at RHIC seem to be inconsistent with this unitarity bound.

  5. Bounds for nonlocality distillation protocols

    SciTech Connect

    Forster, Manuel

    2011-06-15

    Nonlocality can be quantified by the violation of a Bell inequality. Since this violation may be amplified by local operations, an alternative measure has been proposed--distillable nonlocality. The alternative measure is difficult to calculate exactly due to the double exponential growth of the parameter space. In this paper, we give a way to bound the distillable nonlocality of a resource by the solutions to a related optimization problem. Our upper bounds are exponentially easier to compute than the exact value and are shown to be meaningful in general and tight in some cases.

  6. Osteopontin Upregulation in Rotavirus-Induced Murine Biliary Atresia Requires Replicating Virus But is Not Necessary for Development of Biliary Atresia

    PubMed Central

    Hertel, Paula M.; Crawford, Sue E.; Finegold, Milton J.; Estes, Mary K.

    2011-01-01

    Biliary atresia (BA) is a progressive fibro-inflammatory pediatric liver disease in which osteopontin (OPN), a glycoprotein with inflammatory and fibrogenic activity, may play a pathogenic role. The current studies were conducted in a mouse model of rotavirus-induced BA to test the hypotheses that live but not inactivated rotavirus causes antigenemia, upregulation of hepatic OPN expression, and induction of BA and fibrosis; and that OPN is necessary for development of BA. Prolonged or transient antigenemia developed in mice inoculated with live or inactivated virus, respectively, but only live virus upregulated hepatic OPN and caused BA and fibrosis. OPN was expressed in intra- and extrahepatic bile ducts in healthy mice and in mice with BA. OPN-deficient mice, similarly to WT mice, developed BA. Together, these data show that live but not inactivated rotavirus causes upregulation of hepatic OPN expression and BA but that OPN is not necessary for development of BA. PMID:21742364

  7. Spinal but not cortical microglia acquire an atypical phenotype with high VEGF, galectin-3 and osteopontin, and blunted inflammatory responses in ALS rats.

    PubMed

    Nikodemova, Maria; Small, Alissa L; Smith, Stephanie M C; Mitchell, Gordon S; Watters, Jyoti J

    2014-09-01

    Activation of microglia, CNS resident immune cells, is a pathological hallmark of amyotrophic lateral sclerosis (ALS), a neurodegenerative disorder affecting motor neurons. Despite evidence that microglia contribute to disease progression, the exact role of these cells in ALS pathology remains unknown. We immunomagnetically isolated microglia from different CNS regions of SOD1(G93A) rats at three different points in disease progression: presymptomatic, symptom onset and end-stage. We observed no differences in microglial number or phenotype in presymptomatic rats compared to wild-type controls. Although after disease onset there was no macrophage infiltration, there were significant increases in microglial numbers in the spinal cord, but not cortex. At disease end-stage, microglia were characterized by high expression of galectin-3, osteopontin and VEGF, and concomitant downregulated expression of TNFα, IL-6, BDNF and arginase-1. Flow cytometry revealed the presence of at least two phenotypically distinct microglial populations in the spinal cord. Immunohistochemistry showed that galectin-3/osteopontin positive microglia were restricted to the ventral horns of the spinal cord, regions with severe motor neuron degeneration. End-stage SOD1(G93A) microglia from the cortex, a less affected region, displayed similar gene expression profiles to microglia from wild-type rats, and displayed normal responses to systemic inflammation induced by LPS. On the other hand, end-stage SOD1(G93A) spinal microglia had blunted responses to systemic LPS suggesting that in addition to their phenotypic changes, they may also be functionally impaired. Thus, after disease onset, microglia acquired unique characteristics that do not conform to typical M1 (inflammatory) or M2 (anti-inflammatory) phenotypes. This transformation was observed only in the most affected CNS regions, suggesting that overexpression of mutated hSOD1 is not sufficient to trigger these changes in microglia. These

  8. Symmetry and boundness of four-particle coulomb systems

    SciTech Connect

    Rebane, T. K.

    2012-04-15

    The problem of boundness of a{sup +}b{sup +}c{sup -}d{sup -} four-particle Coulomb systems (quadrions) is studied versus the masses of the particles involved. Inequalities that make it possible to deduce that, if some reference quadrions form a bound state, the same is true for a large number of quadrions formed by particles having various masses were derived. A compendium of calculations for energies of reference systems that possess various symmetries [positronium molecules (e{sup +}e{sup +}e{sup -}e{sup -}) and quadrions of the a{sup +}b{sup +}b{sup -}b{sup -}, a{sup +}b{sup +}a{sup --}, and a{sup +}a{sup +}b{sup -}c{sup -} types] is given, and groups of bound asymmetric quadrions corresponding to them are determined. An inequality for kinetic energies of particles that makes it possible to find out, by using asymmetric reference systems, whether specific quadrions are bound is obtained. It is shown that the boundness of many quadrions is ensured by the boundness of respective three-particle systems. The entire body of the present results permits proving that, of the total number of 406 quadrions containing electrons, muons, pions, kaons, protons, deuterons, and tritons and their antiparticles, 227 quadrions are bound.

  9. Loosely-Bound Diatomic Molecules.

    ERIC Educational Resources Information Center

    Balfour, W. J.

    1979-01-01

    Discusses concept of covalent bonding as related to homonuclear diatomic molecules. Article draws attention to the existence of bound rare gas and alkaline earth diatomic molecules. Summarizes their molecular parameters and offers spectroscopic data. Strength and variation with distance of interatomic attractive forces is given. (Author/SA)

  10. Teacher Education in Outward Bound.

    ERIC Educational Resources Information Center

    Robinson, Richard A.

    A series of Outward Bound programs and experiences was planned for El Paso County, Colorado, school teachers to increase their awareness of their personal characteristics, especially those that might enhance learning on the part of their students. Part of the planning for the program involved a survey of county high school teachers, counselors,…