An enhanceosome containing the Jun B/Fra-2 heterodimer and the HMG-I(Y) architectural protein controls HPV 18 transcription.

PubMed

Bouallaga, I; Massicard, S; Yaniv, M; Thierry, F

2000-11-01

Recent studies have reported new mechanisms that mediate the transcriptional synergy of strong tissue-specific enhancers, involving the cooperative assembly of higher-order nucleoprotein complexes called enhanceosomes. Here we show that the HPV18 enhancer, which controls the epithelial-specific transcription of the E6 and E7 transforming genes, exhibits characteristic features of these structures. We used deletion experiments to show that a core enhancer element cooperates, in a specific helical phasing, with distant essential factors binding to the ends of the enhancer. This core sequence, binding a Jun B/Fra-2 heterodimer, cooperatively recruits the architectural protein HMG-I(Y) in a nucleoprotein complex, where they interact with each other. Therefore, in HeLa cells, HPV18 transcription seems to depend upon the assembly of an enhanceosome containing multiple cellular factors recruited by a core sequence interacting with AP1 and HMG-I(Y).

Unphysiologically high magnesium concentrations support chondrocyte proliferation and redifferentiation.

PubMed

Feyerabend, Frank; Witte, Frank; Kammal, Michael; Willumeit, Regine

2006-12-01

The effect of unphysiologically high extracellular magnesium concentrations on chondrocytes, induced by the supplementation of magnesium sulfate, was studied using a 3-phase tissue engineering model. The experiments showed that chondrocyte proliferation and redifferentiation, on the gene and protein expression level, are enhanced. A negative influence was found during chondrogenesis where an inhibition of extracellular matrix formation was observed. In addition, a direct impact on chondrocyte metabolism, elevated magnesium concentrations also affected growth factor effectiveness by consecutive influences during chondrogenesis. All observations were dosage dependent. The results of this study indicate that magnesium may be a useful tool for cartilage tissue engineering.

Imaging analyses of coagulation-dependent initiation of fibrinolysis on activated platelets and its modification by thrombin-activatable fibrinolysis inhibitor.

PubMed

Brzoska, Tomasz; Suzuki, Yuko; Sano, Hideto; Suzuki, Seiichirou; Tomczyk, Martyna; Tanaka, Hiroki; Urano, Tetsumei

2017-04-03

Using intravital confocal microscopy, we observed previously that the process of platelet phosphatidylserine (PS) exposure, fibrin formation and lysine binding site-dependent plasminogen (plg) accumulation took place only in the centre of thrombi, not at their periphery. These findings prompted us to analyse the spatiotemporal regulatory mechanisms underlying coagulation and fibrinolysis. We analysed the fibrin network formation and the subsequent lysis in an in vitro experiment using diluted platelet-rich plasma supplemented with fluorescently labelled coagulation and fibrinolytic factors, using confocal laser scanning microscopy. The structure of the fibrin network formed by supplemented tissue factor was uneven and denser at the sites of coagulation initiation regions (CIRs) on PS-exposed platelets. When tissue-type plasminogen activator (tPA; 7.5 nM) was supplemented, labelled plg (50 nM) as well as tPA accumulated at CIRs, from where fibrinolysis started and gradually expanded to the peripheries. The lysis time at CIRs and their peripheries (50 µm from the CIR) were 27.9 ± 6.6 and 44.4 ± 9.7 minutes (mean ± SD, n=50 from five independent experiments) after the addition of tissue factor, respectively. Recombinant human soluble thrombomodulin (TMα; 2.0 nM) attenuated the CIR-dependent plg accumulation and strongly delayed fibrinolysis at CIRs. A carboxypeptidase inhibitor dose-dependently enhanced the CIR-dependent fibrinolysis initiation, and at 20 µM it completely abrogated the TMα-induced delay of fibrinolysis. Our findings are the first to directly present crosstalk between coagulation and fibrinolysis, which takes place on activated platelets' surface and is further controlled by thrombin-activatable fibrinolysis inhibitor (TAFI).

Insights into insect wing origin provided by functional analysis of vestigial in the red flour beetle, Tribolium castaneum.

PubMed

Clark-Hachtel, Courtney M; Linz, David M; Tomoyasu, Yoshinori

2013-10-15

Despite accumulating efforts to unveil the origin of insect wings, it remains one of the principal mysteries in evolution. Currently, there are two prominent models regarding insect wing origin: one connecting the origin to the paranotal lobe and the other to the proximodorsal leg branch (exite). However, neither hypothesis has been able to surpass the other. To approach this conundrum, we focused our analysis on vestigial (vg), a critical wing gene initially identified in Drosophila. Our investigation in Tribolium (Coleoptera) has revealed that, despite the well-accepted view of vg as an essential wing gene, there are two groups of vg-dependent tissues in the "wingless" first thoracic segment (T1). We show that one of these tissues, the carinated margin, also depends on other factors essential for wing development (such as Wingless signal and apterous), and has nubbin enhancer activity. In addition, our homeotic mutant analysis shows that wing transformation in T1 originates from both the carinated margin and the other vg-dependent tissue, the pleural structures (trochantin and epimeron). Intriguingly, these two tissues may actually be homologous to the two proposed wing origins (paranotal lobes and exite bearing proximal leg segments). Therefore, our findings suggest that the vg-dependent tissues in T1 could be wing serial homologs present in a more ancestral state, thus providing compelling functional evidence for the dual origin of insect wings.

The angiogenic factor CCN1 promotes adhesion and migration of circulating CD34+ progenitor cells: potential role in angiogenesis and endothelial regeneration.

PubMed

Grote, Karsten; Salguero, Gustavo; Ballmaier, Matthias; Dangers, Marc; Drexler, Helmut; Schieffer, Bernhard

2007-08-01

Tissue regeneration involves the formation of new blood vessels regulated by angiogenic factors. We reported recently that the expression of the angiogenic factor CCN1 is up-regulated under various pathophysiologic conditions within the cardiovascular system. Because CD34+ progenitor cells participate in cardiovascular tissue regeneration, we investigated whether CCN1-detected for the first time in human plasma-promotes the recruitment of CD34+ progenitor cells to endothelial cells, thereby enhancing endothelial proliferation and neovascularization. In this study, we demonstrated that CCN1 and supernatants from CCN1-stimulated human CD34+ progenitor cells promoted proliferation of endothelial cells and angiogenesis in vitro and in vivo. In addition, CCN1 induced migration and transendothelial migration of CD34+ cells and the release of multiple growth factors, chemokines, and matrix metalloproteinase-9 (MMP-9) from these cells. Moreover, the CCN1-specific integrins alpha(M)beta(2) and alpha(V)beta(3) are expressed on CD34+ cells and CCN1 stimulated integrin-dependent signaling. Furthermore, integrin antagonists (RGD-peptides) suppressed both binding of CCN1 to CD34+ cells and CCN1-induced adhesion of CD34+ cells to endothelial cells. These data suggest that CCN1 promotes integrin-dependent recruitment of CD34+ progenitor cells to endothelial cells, which may contribute to paracrine effects on angiogenesis and tissue regeneration.

ChIP-seq Identification of Weakly Conserved Heart Enhancers

PubMed Central

Blow, Matthew J.; McCulley, David J.; Li, Zirong; Zhang, Tao; Akiyama, Jennifer A.; Holt, Amy; Plajzer-Frick, Ingrid; Shoukry, Malak; Wright, Crystal; Chen, Feng; Afzal, Veena; Bristow, James; Ren, Bing; Black, Brian L.; Rubin, Edward M.; Visel, Axel; Pennacchio, Len A.

2011-01-01

Accurate control of tissue-specific gene expression plays a pivotal role in heart development, but few cardiac transcriptional enhancers have thus far been identified. Extreme non-coding sequence conservation successfully predicts enhancers active in many tissues, but fails to identify substantial numbers of heart enhancers. Here we used ChIP-seq with the enhancer-associated protein p300 from mouse embryonic day 11.5 heart tissue to identify over three thousand candidate heart enhancers genome-wide. Compared to other tissues studied at this time-point, most candidate heart enhancers are less deeply conserved in vertebrate evolution. Nevertheless, the testing of 130 candidate regions in a transgenic mouse assay revealed that most of them reproducibly function as enhancers active in the heart, irrespective of their degree of evolutionary constraint. These results provide evidence for a large population of poorly conserved heart enhancers and suggest that the evolutionary constraint of embryonic enhancers can vary depending on tissue type. PMID:20729851

Targeting the NO/superoxide ratio in adipose tissue: relevance to obesity and diabetes management.

PubMed

Jankovic, Aleksandra; Korac, Aleksandra; Buzadzic, Biljana; Stancic, Ana; Otasevic, Vesna; Ferdinandy, Péter; Daiber, Andreas; Korac, Bato

2017-06-01

Insulin sensitivity and metabolic homeostasis depend on the capacity of adipose tissue to take up and utilize excess glucose and fatty acids. The key aspects that determine the fuel-buffering capacity of adipose tissue depend on the physiological levels of the small redox molecule, nitric oxide (NO). In addition to impairment of NO synthesis, excessive formation of the superoxide anion (О 2 •- ) in adipose tissue may be an important interfering factor diverting the signalling of NO and other reactive oxygen and nitrogen species in obesity, resulting in metabolic dysfunction of adipose tissue over time. Besides its role in relief from superoxide burst, enhanced NO signalling may be responsible for the therapeutic benefits of different superoxide dismutase mimetics, in obesity and experimental diabetes models. This review summarizes the role of NO in adipose tissue and highlights the effects of NO/О 2 •- ratio 'teetering' as a promising pharmacological target in the metabolic syndrome. This article is part of a themed section on Redox Biology and Oxidative Stress in Health and Disease. To view the other articles in this section visit http://onlinelibrary.wiley.com/doi/10.1111/bph.v174.12/issuetoc. © 2016 The British Pharmacological Society.

Hard tissue remodeling using biofabricated coralline biomaterials.

PubMed

Vago, Razi; Plotquin, Daniel; Bunin, Alex; Sinelnikov, Igor; Atar, Dan; Itzhak, David

2002-01-04

Biotechnical and biomedical approaches were combined in an attempt to identify potential uses of biofabricated marine carbonate materials in biomedical applications, particularly as biomatrices for remodeling bone and cartilage tissue. After grafting, it is desirable for bone ingrowth to proceed as quickly as possible because the strength of the implanted region depends on a good mechanical bond forming between the implant and surrounding regions in the body. Ingrowth can take place as a result of growth of tissue and cells into the implanted porous material, or it may be promoted by transplanting cells seeded onto such a material. The rate at which ingrowth occurs is dependent on many factors, including pore size and the interconnectivity of the implanted structure. In vivo graftings into osteochondral defects demonstrated that our biofabricated porous material is highly biocompatible with cartilage and bone tissue. The biofabricated matrix was well incorporated into the biphasic osteochondral area. Resorption was followed by bone and cartilage formation, and after 4 months, the biomaterial had been replaced by new tissue. Ossification was induced and enhanced without introduction of additional factors. We believe that this is the first time that such biofabricated materials have been used for biomedical purposes. In face of the obvious environmental disadvantages of harvesting from limited natural resources, we propose the use of bioengineered coralline and other materials such as those cultured by our group under field and laboratory conditions as a possible biomatrix for hard tissue remodeling.

Generating favorable growth factor and protease release profiles to enable extracellular matrix accumulation within an in vitro tissue engineering environment.

PubMed

Zhang, Xiaoqing; Battiston, Kyle G; Labow, Rosalind S; Simmons, Craig A; Santerre, J Paul

2017-05-01

Tissue engineering (particularly for the case of load-bearing cardiovascular and connective tissues) requires the ability to promote the production and accumulation of extracellular matrix (ECM) components (e.g., collagen, glycosaminoglycan and elastin). Although different approaches have been attempted in order to enhance ECM accumulation in tissue engineered constructs, studies of underlying signalling mechanisms that influence ECM deposition and degradation during tissue remodelling and regeneration in multi-cellular culture systems have been limited. The current study investigated vascular smooth muscle cell (VSMC)-monocyte co-culture systems using different VSMC:monocyte ratios, within a degradable polyurethane scaffold, to assess their influence on ECM generation and degradation processes, and to elucidate relevant signalling molecules involved in this in vitro vascular tissue engineering system. It was found that a desired release profile of growth factors (e.g. insulin growth factor-1 (IGF-1)) and hydrolytic proteases (e.g. matrix-metalloproteinases 2, 9, 13 and 14 (MMP2, MMP9, MMP13 and MMP14)), could be achieved in co-culture systems, yielding an accumulation of ECM (specifically for 2:1 and 4:1 VSMC:monocyte culture systems). This study has significant implications for the tissue engineering field (including vascular tissue engineering), not only because it identified important cytokines and proteases that control ECM accumulation/degradation within synthetic tissue engineering scaffolds, but also because the established culture systems could be applied to improve the development of different types of tissue constructs. Sufficient extracellular matrix accumulation within cardiovascular and connective tissue engineered constructs is a prerequisite for their appropriate function in vivo. This study established co-culture systems with tissue specific cells (vascular smooth muscle cells (VSMCs)) and defined ratios of immune cells (monocytes) to investigate extracellular matrix (ECM) generation and degradation processes, revealing important mechanisms underlying ECM turnover during vascular tissue regeneration/remodelling. A specific growth factor (IGF-1), as well as hydrolytic proteases (e.g. MMP2, MMP9, MMP13 and MMP14), were identified as playing important roles in these processes. ECM accumulation was found to be dependent on achieving a desired release profile of these ECM-promoting and ECM-degrading factors within the multi-cellular microenvironment. The findings enhance our understanding of ECM deposition and degradation during in vitro tissue engineering and would be applicable to the repair or regeneration of a variety of tissues. Copyright © 2017 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

Matriptase activation connects tissue factor-dependent coagulation initiation to epithelial proteolysis and signaling.

PubMed

Le Gall, Sylvain M; Szabo, Roman; Lee, Melody; Kirchhofer, Daniel; Craik, Charles S; Bugge, Thomas H; Camerer, Eric

2016-06-23

The coagulation cascade is designed to sense tissue injury by physical separation of the membrane-anchored cofactor tissue factor (TF) from inactive precursors of coagulation proteases circulating in plasma. Once TF on epithelial and other extravascular cells is exposed to plasma, sequential activation of coagulation proteases coordinates hemostasis and contributes to host defense and tissue repair. Membrane-anchored serine proteases (MASPs) play critical roles in the development and homeostasis of epithelial barrier tissues; how MASPs are activated in mature epithelia is unknown. We here report that proteases of the extrinsic pathway of blood coagulation transactivate the MASP matriptase, thus connecting coagulation initiation to epithelial proteolysis and signaling. Exposure of TF-expressing cells to factors (F) VIIa and Xa triggered the conversion of latent pro-matriptase to an active protease, which in turn cleaved the pericellular substrates protease-activated receptor-2 (PAR2) and pro-urokinase. An activation pathway-selective PAR2 mutant resistant to direct cleavage by TF:FVIIa and FXa was activated by these proteases when cells co-expressed pro-matriptase, and matriptase transactivation was necessary for efficient cleavage and activation of wild-type PAR2 by physiological concentrations of TF:FVIIa and FXa. The coagulation initiation complex induced rapid and prolonged enhancement of the barrier function of epithelial monolayers that was dependent on matriptase transactivation and PAR2 signaling. These observations suggest that the coagulation cascade engages matriptase to help coordinate epithelial defense and repair programs after injury or infection, and that matriptase may contribute to TF-driven pathogenesis in cancer and inflammation.

Hippo, TGF-β, and Src-MAPK pathways regulate transcription of the upd3 cytokine in Drosophila enterocytes upon bacterial infection.

PubMed

Houtz, Philip; Bonfini, Alessandro; Liu, Xi; Revah, Jonathan; Guillou, Aurélien; Poidevin, Mickael; Hens, Korneel; Huang, Hsin-Yi; Deplancke, Bart; Tsai, Yu-Chen; Buchon, Nicolas

2017-11-01

Cytokine signaling is responsible for coordinating conserved epithelial regeneration and immune responses in the digestive tract. In the Drosophila midgut, Upd3 is a major cytokine, which is induced in enterocytes (EC) and enteroblasts (EB) upon oral infection, and initiates intestinal stem cell (ISC) dependent tissue repair. To date, the genetic network directing upd3 transcription remains largely uncharacterized. Here, we have identified the key infection-responsive enhancers of the upd3 gene and show that distinct enhancers respond to various stresses. Furthermore, through functional genetic screening, bioinformatic analyses and yeast one-hybrid screening, we determined that the transcription factors Scalloped (Sd), Mothers against dpp (Mad), and D-Fos are principal regulators of upd3 expression. Our study demonstrates that upd3 transcription in the gut is regulated by the activation of multiple pathways, including the Hippo, TGF-β/Dpp, and Src, as well as p38-dependent MAPK pathways. Thus, these essential pathways, which are known to control ISC proliferation cell-autonomously, are also activated in ECs to promote tissue turnover the regulation of upd3 transcription.

Distinct modes of gene regulation by a cell-specific transcriptional activator.

PubMed

Sengupta, Tanushri; Cohet, Nathalie; Morlé, François; Bieker, James J

2009-03-17

The architectural layout of a eukaryotic RNA polymerase II core promoter plays a role in general transcriptional activation. However, its role in tissue-specific expression is not known. For example, differing modes of its recognition by general transcription machinery can provide an additional layer of control within which a single tissue-restricted transcription factor may operate. Erythroid Kruppel-like factor (EKLF) is a hematopoietic-specific transcription factor that is critical for the activation of subset of erythroid genes. We find that EKLF interacts with TATA binding protein-associated factor 9 (TAF9), which leads to important consequences for expression of adult beta-globin. First, TAF9 functionally supports EKLF activity by enhancing its ability to activate the beta-globin gene. Second, TAF9 interacts with a conserved beta-globin downstream promoter element, and ablation of this interaction by beta-thalassemia-causing mutations decreases its promoter activity and disables superactivation. Third, depletion of EKLF prevents recruitment of TAF9 to the beta-globin promoter, whereas depletion of TAF9 drastically impairs beta-promoter activity. However, a TAF9-independent mode of EKLF transcriptional activation is exhibited by the alpha-hemoglobin-stabilizing protein (AHSP) gene, which does not contain a discernable downstream promoter element. In this case, TAF9 does not enhance EKLF activity and depletion of TAF9 has no effect on AHSP promoter activation. These studies demonstrate that EKLF directs different modes of tissue-specific transcriptional activation depending on the architecture of its target core promoter.

A Novel mouse model of enhanced proteostasis: Full-length human heat shock factor 1 transgenic mice

DOE Office of Scientific and Technical Information (OSTI.GOV)

Pierce, Anson, E-mail: piercea2@uthscsa.edu; Barshop Institute for Longevity and Aging Studies, The University of Texas Health Science Center at San Antonio, San Antonio, Texas, 78229; The Department of Veteran's Affairs, South Texas Veterans Health Care System, San Antonio, Texas, 78284

2010-11-05

Research highlights: {yields} Development of mouse overexpressing native human HSF1 in all tissues including CNS. {yields} HSF1 overexpression enhances heat shock response at whole-animal and cellular level. {yields} HSF1 overexpression protects from polyglutamine toxicity and favors aggresomes. {yields} HSF1 overexpression enhances proteostasis at the whole-animal and cellular level. -- Abstract: The heat shock response (HSR) is controlled by the master transcriptional regulator heat shock factor 1 (HSF1). HSF1 maintains proteostasis and resistance to stress through production of heat shock proteins (HSPs). No transgenic model exists that overexpresses HSF1 in tissues of the central nervous system (CNS). We generated a transgenicmore » mouse overexpressing full-length non-mutant HSF1 and observed a 2-4-fold increase in HSF1 mRNA and protein expression in all tissues studied of HSF1 transgenic (HSF1{sup +/0}) mice compared to wild type (WT) littermates, including several regions of the CNS. Basal expression of HSP70 and 90 showed only mild tissue-specific changes; however, in response to forced exercise, the skeletal muscle HSR was more elevated in HSF1{sup +/0} mice compared to WT littermates and in fibroblasts following heat shock, as indicated by levels of inducible HSP70 mRNA and protein. HSF1{sup +/0} cells elicited a significantly more robust HSR in response to expression of the 82 repeat polyglutamine-YFP fusion construct (Q82YFP) and maintained proteasome-dependent processing of Q82YFP compared to WT fibroblasts. Overexpression of HSF1 was associated with fewer, but larger Q82YFP aggregates resembling aggresomes in HSF1{sup +/0} cells, and increased viability. Therefore, our data demonstrate that tissues and cells from mice overexpressing full-length non-mutant HSF1 exhibit enhanced proteostasis.« less

Enhanced nutrient transport improves the depth-dependent properties of tri-layered engineered cartilage constructs with zonal co-culture of chondrocytes and MSCs.

PubMed

Kim, Minwook; Farrell, Megan J; Steinberg, David R; Burdick, Jason A; Mauck, Robert L

2017-08-01

Biomimetic design in cartilage tissue engineering is a challenge given the complexity of the native tissue. While numerous studies have generated constructs with near-native bulk properties, recapitulating the depth-dependent features of native tissue remains a challenge. Furthermore, limitations in nutrient transport and matrix accumulation in engineered constructs hinders maturation within the central core of large constructs. To overcome these limitations, we fabricated tri-layered constructs that recapitulate the depth-dependent cellular organization and functional properties of native tissue using zonally derived chondrocytes co-cultured with MSCs. We also introduced porous hollow fibers (HFs) and HFs/cotton threads to enhance nutrient transport. Our results showed that tri-layered constructs with depth-dependent organization and properties could be fabricated. The addition of HFs or HFs/threads improved matrix accumulation in the central core region. With HF/threads, the local modulus in the deep region of tri-layered constructs nearly matched that of native tissue, though the properties in the central regions remained lower. These constructs reproduced the zonal organization and depth-dependent properties of native tissue, and demonstrate that a layer-by-layer fabrication scheme holds promise for the biomimetic repair of focal cartilage defects. Articular cartilage is a highly organized tissue driven by zonal heterogeneity of cells, extracellular matrix proteins and fibril orientations, resulting in depth-dependent mechanical properties. Therefore, the recapitulation of the functional properties of native cartilage in a tissue engineered construct requires such a biomimetic design of the morphological organization, and this has remained a challenge in cartilage tissue engineering. This study demonstrates that a layer-by-layer fabrication scheme, including co-cultures of zone-specific articular CHs and MSCs, can reproduce the depth-dependent characteristics and mechanical properties of native cartilage while minimizing the need for large numbers of chondrocytes. In addition, introduction of a porous hollow fiber (combined with a cotton thread) enhanced nutrient transport and depth-dependent properties of the tri-layered construct. Such a tri-layered construct may provide critical advantages for focal cartilage repair. These constructs hold promise for restoring native tissue structure and function, and may be beneficial in terms of zone-to-zone integration with adjacent host tissue and providing more appropriate strain transfer after implantation. Copyright © 2017 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

ΔNp63 promotes stem cell activity in mammary gland development and basal-like breast cancer by enhancing Fzd7 expression and Wnt signaling

PubMed Central

Chakrabarti, Rumela; Wei, Yong; Hwang, Julie; Hang, Xiang; Blanco, Mario Andres; Choudhury, Abrar; Tiede, Benjamin; Romano, Rose-Anne; DeCoste, Christina; Mercatali, Laura; Ibrahim, Toni; Amadori, Dino; Kannan, Nagarajan; Eaves, Connie J; Sinha, Satrajit; Kang, Yibin

2014-01-01

Emerging evidence suggests that cancer is populated and maintained by tumor initiating cells (TICs) with stem-like properties similar to that of adult tissue stem cells. Despite recent advances, the molecular regulatory mechanisms that may be shared between normal and malignant stem cells remain poorly understood. Here we show that the ΔNp63 isoform of the Trp63 transcription factor promotes normal mammary stem cell (MaSC) activity by increasing the expression of the Wnt receptor Fzd7, thereby enhancing Wnt signaling. Importantly, Fzd7-dependent enhancement of Wnt signaling by ΔNp63 also governs tumor initiating activity of the basal subtype of breast cancer. These findings establish ΔNp63 as a key regulator of stem cells in both normal and malignant mammary tissues and provide direct evidence that breast cancer TICs and normal MaSCs share common regulatory mechanisms. PMID:25241036

Insights into insect wing origin provided by functional analysis of vestigial in the red flour beetle, Tribolium castaneum

PubMed Central

Clark-Hachtel, Courtney M.; Linz, David M.; Tomoyasu, Yoshinori

2013-01-01

Despite accumulating efforts to unveil the origin of insect wings, it remains one of the principal mysteries in evolution. Currently, there are two prominent models regarding insect wing origin: one connecting the origin to the paranotal lobe and the other to the proximodorsal leg branch (exite). However, neither hypothesis has been able to surpass the other. To approach this conundrum, we focused our analysis on vestigial (vg), a critical wing gene initially identified in Drosophila. Our investigation in Tribolium (Coleoptera) has revealed that, despite the well-accepted view of vg as an essential wing gene, there are two groups of vg-dependent tissues in the “wingless” first thoracic segment (T1). We show that one of these tissues, the carinated margin, also depends on other factors essential for wing development (such as Wingless signal and apterous), and has nubbin enhancer activity. In addition, our homeotic mutant analysis shows that wing transformation in T1 originates from both the carinated margin and the other vg-dependent tissue, the pleural structures (trochantin and epimeron). Intriguingly, these two tissues may actually be homologous to the two proposed wing origins (paranotal lobes and exite bearing proximal leg segments). Therefore, our findings suggest that the vg-dependent tissues in T1 could be wing serial homologs present in a more ancestral state, thus providing compelling functional evidence for the dual origin of insect wings. PMID:24085843

Multiple roles of the coagulation protease cascade during virus infection.

PubMed

Antoniak, Silvio; Mackman, Nigel

2014-04-24

The coagulation cascade is activated during viral infections. This response may be part of the host defense system to limit spread of the pathogen. However, excessive activation of the coagulation cascade can be deleterious. In fact, inhibition of the tissue factor/factor VIIa complex reduced mortality in a monkey model of Ebola hemorrhagic fever. Other studies showed that incorporation of tissue factor into the envelope of herpes simplex virus increases infection of endothelial cells and mice. Furthermore, binding of factor X to adenovirus serotype 5 enhances infection of hepatocytes but also increases the activation of the innate immune response to the virus. Coagulation proteases activate protease-activated receptors (PARs). Interestingly, we and others found that PAR1 and PAR2 modulate the immune response to viral infection. For instance, PAR1 positively regulates TLR3-dependent expression of the antiviral protein interferon β, whereas PAR2 negatively regulates expression during coxsackievirus group B infection. These studies indicate that the coagulation cascade plays multiple roles during viral infections.

Uncaria rhynchophylla induces angiogenesis in vitro and in vivo.

PubMed

Choi, Do-Young; Huh, Jeong-Eun; Lee, Jae-Dong; Cho, Eun-Mi; Baek, Yong-Hyeon; Yang, Ha-Ru; Cho, Yoon-Je; Kim, Kang-Il; Kim, Deog-Yoon; Park, Dong-Suk

2005-12-01

Angiogenesis consists of the proliferation, migration, and differentiation of endothelial cells, and angiogenic factors and matrix protein interactions modulate this process. The aim of this study was to determine the angiogenic properties of Uncaria rhynchophylla. Uncaria rhynchophylla significantly enhanced human umbilical vein endothelial cells (HUVECs) proliferation in a dose-dependent manner. Neutralization of vascular endothelial growth factor (VEGF) or basic fibroblast growth factor (bFGF) by monoclonal antibody suppressed the Uncaria rhynchophylla stimulatory effect on proliferation. In addition, Uncaria rhynchophylla significantly increased chemotactic-migration on gelatin and tubular structures on Matrigel of HUVECs in a dose-dependent manner. Interestingly, Uncaria rhynchophylla dose-dependently increased VEGF, and bFGF gene expression and protein secretion of HUVEC. The angiogenic activity of Uncaria rhynchophylla was confirmed using an in vivo Matrigel angiogenesis model, showing promotion of blood vessel formation. These results suggest that Uncaria rhynchophylla could potentially used to accelerate vascular wound healing or to promote the growth of collateral blood vessel in ischemic tissues.

Insulin signalling mechanisms for triacylglycerol storage.

PubMed

Czech, M P; Tencerova, M; Pedersen, D J; Aouadi, M

2013-05-01

Insulin signalling is uniquely required for storing energy as fat in humans. While de novo synthesis of fatty acids and triacylglycerol occurs mostly in liver, adipose tissue is the primary site for triacylglycerol storage. Insulin signalling mechanisms in adipose tissue that stimulate hydrolysis of circulating triacylglycerol, uptake of the released fatty acids and their conversion to triacylglycerol are poorly understood. New findings include (1) activation of DNA-dependent protein kinase to stimulate upstream stimulatory factor (USF)1/USF2 heterodimers, enhancing the lipogenic transcription factor sterol regulatory element binding protein 1c (SREBP1c); (2) stimulation of fatty acid synthase through AMP kinase modulation; (3) mobilisation of lipid droplet proteins to promote retention of triacylglycerol; and (4) upregulation of a novel carbohydrate response element binding protein β isoform that potently stimulates transcription of lipogenic enzymes. Additionally, insulin signalling through mammalian target of rapamycin to activate transcription and processing of SREBP1c described in liver may apply to adipose tissue. Paradoxically, insulin resistance in obesity and type 2 diabetes is associated with increased triacylglycerol synthesis in liver, while it is decreased in adipose tissue. This and other mysteries about insulin signalling and insulin resistance in adipose tissue make this topic especially fertile for future research.

Enzymatically oxidized phospholipids restore thrombin generation in coagulation factor deficiencies.

PubMed

Slatter, David A; Percy, Charles L; Allen-Redpath, Keith; Gajsiewicz, Joshua M; Brooks, Nick J; Clayton, Aled; Tyrrell, Victoria J; Rosas, Marcela; Lauder, Sarah N; Watson, Andrew; Dul, Maria; Garcia-Diaz, Yoel; Aldrovandi, Maceler; Heurich, Meike; Hall, Judith; Morrissey, James H; Lacroix-Desmazes, Sebastien; Delignat, Sandrine; Jenkins, P Vincent; Collins, Peter W; O'Donnell, Valerie B

2018-03-22

Hemostatic defects are treated using coagulation factors; however, clot formation also requires a procoagulant phospholipid (PL) surface. Here, we show that innate immune cell-derived enzymatically oxidized phospholipids (eoxPL) termed hydroxyeicosatetraenoic acid-phospholipids (HETE-PLs) restore hemostasis in human and murine conditions of pathological bleeding. HETE-PLs abolished blood loss in murine hemophilia A and enhanced coagulation in factor VIII- (FVIII-), FIX-, and FX-deficient human plasma . HETE-PLs were decreased in platelets from patients after cardiopulmonary bypass (CPB). To explore molecular mechanisms, the ability of eoxPL to stimulate individual isolated coagulation factor/cofactor complexes was tested in vitro. Extrinsic tenase (FVIIa/tissue factor [TF]), intrinsic tenase (FVIIIa/FIXa), and prothrombinase (FVa/FXa) all were enhanced by both HETE-PEs and HETE-PCs, suggesting a common mechanism involving the fatty acid moiety. In plasma, 9-, 15-, and 12-HETE-PLs were more effective than 5-, 11-, or 8-HETE-PLs, indicating positional isomer specificity. Coagulation was enhanced at lower lipid/factor ratios, consistent with a more concentrated area for protein binding. Surface plasmon resonance confirmed binding of FII and FX to HETE-PEs. HETE-PEs increased membrane curvature and thickness, but not surface charge or homogeneity, possibly suggesting increased accessibility to cations/factors. In summary, innate immune-derived eoxPL enhance calcium-dependent coagulation factor function, and their potential utility in bleeding disorders is proposed.

Enzymatically oxidized phospholipids restore thrombin generation in coagulation factor deficiencies

PubMed Central

Slatter, David A.; Percy, Charles L.; Allen-Redpath, Keith; Gajsiewicz, Joshua M.; Brooks, Nick J.; Tyrrell, Victoria J.; Lauder, Sarah N.; Watson, Andrew; Dul, Maria; Garcia-Diaz, Yoel; Aldrovandi, Maceler; Heurich, Meike; Hall, Judith; Lacroix-Desmazes, Sebastien; Delignat, Sandrine; Jenkins, P. Vincent; Collins, Peter W.; O’Donnell, Valerie B.

2018-01-01

Hemostatic defects are treated using coagulation factors; however, clot formation also requires a procoagulant phospholipid (PL) surface. Here, we show that innate immune cell–derived enzymatically oxidized phospholipids (eoxPL) termed hydroxyeicosatetraenoic acid–phospholipids (HETE-PLs) restore hemostasis in human and murine conditions of pathological bleeding. HETE-PLs abolished blood loss in murine hemophilia A and enhanced coagulation in factor VIII- (FVIII-), FIX-, and FX-deficient human plasma . HETE-PLs were decreased in platelets from patients after cardiopulmonary bypass (CPB). To explore molecular mechanisms, the ability of eoxPL to stimulate individual isolated coagulation factor/cofactor complexes was tested in vitro. Extrinsic tenase (FVIIa/tissue factor [TF]), intrinsic tenase (FVIIIa/FIXa), and prothrombinase (FVa/FXa) all were enhanced by both HETE-PEs and HETE-PCs, suggesting a common mechanism involving the fatty acid moiety. In plasma, 9-, 15-, and 12-HETE-PLs were more effective than 5-, 11-, or 8-HETE-PLs, indicating positional isomer specificity. Coagulation was enhanced at lower lipid/factor ratios, consistent with a more concentrated area for protein binding. Surface plasmon resonance confirmed binding of FII and FX to HETE-PEs. HETE-PEs increased membrane curvature and thickness, but not surface charge or homogeneity, possibly suggesting increased accessibility to cations/factors. In summary, innate immune-derived eoxPL enhance calcium-dependent coagulation factor function, and their potential utility in bleeding disorders is proposed. PMID:29563336

Requirement of GM2 ganglioside activator for phospholipase D activation

PubMed Central

Nakamura, Shun-ichi; Akisue, Toshihiro; Jinnai, Hitoshi; Hitomi, Tomohiro; Sarkar, Sukumar; Miwa, Noriko; Okada, Taro; Yoshida, Kimihisa; Kuroda, Shun’ichi; Kikkawa, Ushio; Nishizuka, Yasutomi

1998-01-01

Sequence analysis of a heat-stable protein necessary for the activation of ADP ribosylation factor-dependent phospholipase D (PLD) reveals that this protein has a structure highly homologous to the previously known GM2 ganglioside activator whose deficiency results in the AB-variant of GM2 gangliosidosis. The heat-stable activator protein indeed has the capacity to enhance enzymatic conversion of GM2 to GM3 ganglioside that is catalyzed by β-hexosaminidase A. Inversely, GM2 ganglioside activator purified separately from tissues as described earlier [Conzelmann, E. & Sandhoff, K. (1987) Methods Enzymol. 138, 792–815] stimulates ADP ribosylation factor-dependent PLD in a dose-dependent manner. At higher concentrations of ammonium sulfate, the PLD activator protein apparently substitutes for protein kinase C and phosphatidylinositol 4,5-bisphosphate, both of which are known as effective stimulators of the PLD reaction. The mechanism of action of the heat-stable PLD activator protein remains unknown. PMID:9770472

Expansion and redifferentiation of chondrocytes from osteoarthritic cartilage: cells for human cartilage tissue engineering.

PubMed

Hsieh-Bonassera, Nancy D; Wu, Iwen; Lin, Jonathan K; Schumacher, Barbara L; Chen, Albert C; Masuda, Koichi; Bugbee, William D; Sah, Robert L

2009-11-01

To determine if selected culture conditions enhance the expansion and redifferentiation of chondrocytes isolated from human osteoarthritic cartilage with yields appropriate for creation of constructs for treatment of joint-scale cartilage defects, damage, or osteoarthritis. Chondrocytes isolated from osteoarthritic cartilage were analyzed to determine the effects of medium supplement on cell expansion in monolayer and then cell redifferentiation in alginate beads. Expansion was assessed as cell number estimated from DNA, growth rate, and day of maximal growth. Redifferentiation was evaluated quantitatively from proteoglycan and collagen type II content, and qualitatively by histology and immunohistochemistry. Using either serum or a growth factor cocktail (TFP: transforming growth factor beta1, fibroblast growth factor 2, and platelet-derived growth factor type bb), cell growth rate in monolayer was increased to 5.5x that of corresponding conditions without TFP, and cell number increased 100-fold within 17 days. In subsequent alginate bead culture with human serum or transforming growth factor beta1 and insulin-transferrin-selenium-linoleic acid-bovine serum albumin, redifferentiation was enhanced with increased proteoglycan and collagen type II production. Effects of human serum were dose dependent, and 5% or higher induced formation of chondron-like structures with abundant proteoglycan-rich matrix. Chondrocytes from osteoarthritic cartilage can be stimulated to undergo 100-fold expansion and then redifferentiation, suggesting that they may be useful as a cell source for joint-scale cartilage tissue engineering.

Pigment epithelium-derived factor stimulates skeletal muscle glycolytic activity through NADPH oxidase-dependent reactive oxygen species production.

PubMed

Carnagarin, Revathy; Carlessi, Rodrigo; Newsholme, Philip; Dharmarajan, Arun M; Dass, Crispin R

2016-09-01

Pigment epithelium-derived factor is a multifunctional serpin implicated in insulin resistance in metabolic disorders. Recent evidence suggests that exposure of peripheral tissues such as skeletal muscle to PEDF has profound metabolic consequences with predisposition towards chronic conditions such as obesity, type 2 diabetes, metabolic syndrome and polycystic ovarian syndrome. Chronic inflammation shifts muscle metabolism towards increased glycolysis and decreased oxidative metabolism. In the present study, we demonstrate a novel effect of PEDF on cellular metabolism in mouse cell line (C2C12) and human primary skeletal muscle cells. PEDF addition to skeletal muscle cells induced enhanced phospholipase A2 activity. This was accompanied with increased production of reactive oxygen species in a nicotinamide adenine dinucleotide phosphate (NADPH) oxidase-dependent manner that triggered a shift towards a more glycolytic phenotype. Extracellular flux analysis and glucose consumption assays demonstrated that PEDF treatment resulted in enhanced glycolysis but did not change mitochondrial respiration. Our results demonstrate that skeletal muscle cells express a PEDF-inducible oxidant generating system that enhances glycolysis but is sensitive to antioxidants and NADPH oxidase inhibition. Copyright © 2016 Elsevier Ltd. All rights reserved.

The prospective opportunities offered by magnetic scaffolds for bone tissue engineering: a review

PubMed Central

ORTOLANI, ALESSANDRO; BIANCHI, MICHELE; MOSCA, MASSIMILIANO; CARAVELLI, SILVIO; FUIANO, MARIO; MARCACCI, MAURILIO; RUSSO, ALESSANDRO

2016-01-01

Magnetic scaffolds are becoming increasingly attractive in tissue engineering, due to their ability to enhance bone tissue formation by attracting soluble factors, such as growth factors, hormones and polypeptides, directly to the implantation site, as well as their potential to improve the fixation and stability of the implant. Moreover, there is increasing evidence that the synergistic effects of magnetic scaffolds and magnetic fields can promote bone repair and regeneration. In this manuscript we review the recent innovations in bone tissue engineering that exploit magnetic biomaterials combined with static magnetic fields to enhance bone cell adhesion and proliferation, and thus bone tissue growth. PMID:28217659

Tissue engineering of bladder using vascular endothelial growth factor gene-modified endothelial progenitor cells.

PubMed

Chen, Bai-Song; Xie, Hua; Zhang, Sheng-Li; Geng, Hong-Quan; Zhou, Jun-Mei; Pan, Jun; Chen, Fang

2011-12-01

This study assessed the use of vascular endothelial growth factor (VEGF) gene-modified endothelial progenitor cells (EPCs) seeded onto bladder acellular matrix grafts (BAMGs), to enhance the blood supply in tissue-engineered bladders in a porcine model. Autologous porcine peripheral EPCs were isolated, cultured, expanded, characterized, and modified with the VEGF gene using an adenovirus vector. The expression of VEGF was examined using reverse transcriptase polymerase chain reaction (RT-PCR) and an enzyme-linked immunosorbent assay (ELISA). VEGF gene modified EPCs were seeded onto BAMG and cultured for 3 days before implantation into pigs for bladder tissue engineering. A partial bladder cystectomy was performed in 12 pigs. The experimental group (6 pigs) received VEGF gene-modified EPC-seeded BAMG. The control group (6 pigs) received BAMG without seeded EPCs. The resulting tissue-engineered bladders were subject to a general and histological analysis. Microvessel density (MVD) was assessed using immunohistochemistry. The ex vivo transfection efficiency of EPCs was greater than 60%-70% when concentrated adenovirus was used. The genetically modified cells expressed both VEGF and green fluorescent protein (GFP). Scanning electron microscopy (SEM) and Masson's trichrome staining of cross sections of the cultured cells seeded to BAMG showed cell attachment and proliferation on the surface of the BAMG. Histological examination revealed bladder regeneration in a time-dependent fashion. Significant increases in MVD were observed in the experimental group, in comparison with the control group. VEGF-modified EPCs significantly enhanced neovascularization, compared with BAMG alone. These results indicate that EPCs, combined with VEGF gene therapy, may be a suitable approach for increasing blood supply in the tissue engineering of bladders. Thus, a useful strategy to achieve a tissue-engineered bladder is indicated.

Enhanced bioactive scaffolds for bone tissue regeneration

NASA Astrophysics Data System (ADS)

Karnik, Sonali

Bone injuries are commonly termed as fractures and they vary in their severity and causes. If the fracture is severe and there is loss of bone, implant surgery is prescribed. The response to the implant depends on the patient's physiology and implant material. Sometimes, the compromised physiology and undesired implant reactions lead to post-surgical complications. [4, 5, 20, 28] Efforts have been directed towards the development of efficient implant materials to tackle the problem of post-surgical implant failure. [ 15, 19, 24, 28, 32]. The field of tissue engineering and regenerative medicine involves the use of cells to form a new tissue on bio-absorbable or inert scaffolds. [2, 32] One of the applications of this field is to regenerate the damaged or lost bone by using stem cells or osteoprogenitor cells on scaffolds that can integrate in the host tissue without causing any harmful side effects. [2, 32] A variety of natural, synthetic materials and their combinations have been used to regenerate the damaged bone tissue. [2, 19, 30, 32, 43]. Growth factors have been supplied to progenitor cells to trigger a sequence of metabolic pathways leading to cellular proliferation, differentiation and to enhance their functionality. [56, 57] The challenge persists to supply these proteins, in the range of nano or even picograms, and in a sustained fashion over a period of time. A delivery system has yet to be developed that would mimic the body's inherent mechanism of delivering the growth factor molecules in the required amount to the target organ or tissue. Titanium is the most preferred metal for orthopedic and orthodontic implants. [28, 46, 48] Even though it has better osteogenic properties as compared to other metals and alloys, it still has drawbacks like poor integration into the surrounding host tissue leading to bone resorption and implant failure. [20, 28, 35] It also faces the problem of postsurgical infections that contributes to the implant failure. [26, 37]. The focus of this dissertation was to design and develop novel implant materials for coating titanium to improve its biological properties. These natural and/or semi-synthetic materials improved cellular adhesion, biological response to the scaffolds and prevented growth of bacteria when they were enhanced with growth factor and anti-infective loaded nanotubes. The implant materials showed promise when tested in vitro for cell proliferation, differentiation and bacterial growth inhibition.

Infiltration of plasma rich in growth factors enhances in vivo angiogenesis and improves reperfusion and tissue remodeling after severe hind limb ischemia.

PubMed

Anitua, Eduardo; Pelacho, Beatriz; Prado, Roberto; Aguirre, José Javier; Sánchez, Mikel; Padilla, Sabino; Aranguren, Xabier L; Abizanda, Gloria; Collantes, María; Hernandez, Milagros; Perez-Ruiz, Ana; Peñuelas, Ivan; Orive, Gorka; Prosper, Felipe

2015-03-28

PRGF is a platelet concentrate within a plasma suspension that forms an in situ-generated fibrin-matrix delivery system, releasing multiple growth factors and other bioactive molecules that play key roles in tissue regeneration. This study was aimed at exploring the angiogenic and myogenic effects of PRGF on in vitro endothelial cells (HUVEC) and skeletal myoblasts (hSkMb) as well as on in vivo mouse subcutaneously implanted matrigel and on limb muscles after a severe ischemia. Human PRGF was prepared and characterized. Both proliferative and anti-apoptotic responses to PRGF were assessed in vitro in HUVEC and hSkMb. In vivo murine matrigel plug assay was conducted to determine the angiogenic capacity of PRGF, whereas in vivo ischemic hind limb model was carried out to demonstrate PRGF-driven vascular and myogenic regeneration. Primary HUVEC and hSkMb incubated with PRGF showed a dose dependent proliferative and anti-apoptotic effect and the PRGF matrigel plugs triggered an early and significant sustained angiogenesis compared with the control group. Moreover, mice treated with PRGF intramuscular infiltrations displayed a substantial reperfusion enhancement at day 28 associated with a fibrotic tissue reduction. These findings suggest that PRGF-induced angiogenesis is functionally effective at expanding the perfusion capacity of the new vasculature and attenuating the endogenous tissue fibrosis after a severe-induced skeletal muscle ischemia. Copyright © 2015 Elsevier B.V. All rights reserved.

Customized biomaterials to augment chondrocyte gene therapy.

PubMed

Aguilar, Izath Nizeet; Trippel, Stephen; Shi, Shuiliang; Bonassar, Lawrence J

2017-04-15

A persistent challenge in enhancing gene therapy is the transient availability of the target gene product. This is particularly true in tissue engineering applications. The transient exposure of cells to the product could be insufficient to promote tissue regeneration. Here we report the development of a new material engineered to have a high affinity for a therapeutic gene product. We focus on insulin-like growth factor-I (IGF-I) for its highly anabolic effects on many tissues such as spinal cord, heart, brain and cartilage. One of the ways that tissues store IGF-I is through a group of insulin like growth factor binding proteins (IGFBPs), such as IGFBP-5. We grafted the IGF-I binding peptide sequence from IGFBP-5 onto alginate in order to retain the endogenous IGF-I produced by transfected chondrocytes. This novel material bound IGF-I and released the growth factor for at least 30days in culture. We found that this binding enhanced the biosynthesis of transfected cells up to 19-fold. These data demonstrate the coordinated engineering of cell behavior and material chemistry to greatly enhance extracellular matrix synthesis and tissue assembly, and can serve as a template for the enhanced performance of other therapeutic proteins. The present manuscript focuses on the enhancement of chondrocyte gene therapy through the modification of scaffold materials to enhance the retention of targeted gene products. This study combined tissue engineering and gene therapy, where customized biomaterials augmented the action of IGF-I by enhancing the retention of protein produced by transfection of the IGF-I gene. This approach enabled tuning of binding of IGF-I to alginate, which increased GAG and HYPRO production by transfected chondrocytes. To our knowledge, peptide-based modification of materials to augment growth factor-targeted gene therapy has not been reported previously. Copyright © 2017 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

Identification of activated enhancers and linked transcription factors in breast, prostate, and kidney tumors by tracing enhancer networks using epigenetic traits.

PubMed

Rhie, Suhn Kyong; Guo, Yu; Tak, Yu Gyoung; Yao, Lijing; Shen, Hui; Coetzee, Gerhard A; Laird, Peter W; Farnham, Peggy J

2016-01-01

Although technological advances now allow increased tumor profiling, a detailed understanding of the mechanisms leading to the development of different cancers remains elusive. Our approach toward understanding the molecular events that lead to cancer is to characterize changes in transcriptional regulatory networks between normal and tumor tissue. Because enhancer activity is thought to be critical in regulating cell fate decisions, we have focused our studies on distal regulatory elements and transcription factors that bind to these elements. Using DNA methylation data, we identified more than 25,000 enhancers that are differentially activated in breast, prostate, and kidney tumor tissues, as compared to normal tissues. We then developed an analytical approach called Tracing Enhancer Networks using Epigenetic Traits that correlates DNA methylation levels at enhancers with gene expression to identify more than 800,000 genome-wide links from enhancers to genes and from genes to enhancers. We found more than 1200 transcription factors to be involved in these tumor-specific enhancer networks. We further characterized several transcription factors linked to a large number of enhancers in each tumor type, including GATA3 in non-basal breast tumors, HOXC6 and DLX1 in prostate tumors, and ZNF395 in kidney tumors. We showed that HOXC6 and DLX1 are associated with different clusters of prostate tumor-specific enhancers and confer distinct transcriptomic changes upon knockdown in C42B prostate cancer cells. We also discovered de novo motifs enriched in enhancers linked to ZNF395 in kidney tumors. Our studies characterized tumor-specific enhancers and revealed key transcription factors involved in enhancer networks for specific tumor types and subgroups. Our findings, which include a large set of identified enhancers and transcription factors linked to those enhancers in breast, prostate, and kidney cancers, will facilitate understanding of enhancer networks and mechanisms leading to the development of these cancers.

Cardiac tissue enriched factors serum response factor and GATA-4 are mutual coregulators

NASA Technical Reports Server (NTRS)

Belaguli, N. S.; Sepulveda, J. L.; Nigam, V.; Charron, F.; Nemer, M.; Schwartz, R. J.

2000-01-01

Combinatorial interaction among cardiac tissue-restricted enriched transcription factors may facilitate the expression of cardiac tissue-restricted genes. Here we show that the MADS box factor serum response factor (SRF) cooperates with the zinc finger protein GATA-4 to synergistically activate numerous myogenic and nonmyogenic serum response element (SRE)-dependent promoters in CV1 fibroblasts. In the absence of GATA binding sites, synergistic activation depends on binding of SRF to the proximal CArG box sequence in the cardiac and skeletal alpha-actin promoter. GATA-4's C-terminal activation domain is obligatory for synergistic coactivation with SRF, and its N-terminal domain and first zinc finger are inhibitory. SRF and GATA-4 physically associate both in vivo and in vitro through their MADS box and the second zinc finger domains as determined by protein A pullout assays and by in vivo one-hybrid transfection assays using Gal4 fusion proteins. Other cardiovascular tissue-restricted GATA factors, such as GATA-5 and GATA-6, were equivalent to GATA-4 in coactivating SRE-dependent targets. Thus, interaction between the MADS box and C4 zinc finger proteins, a novel regulatory paradigm, mediates activation of SRF-dependent gene expression.

Propagation of thrombosis by neutrophils and extracellular nucleosome networks

PubMed Central

Pfeiler, Susanne; Stark, Konstantin; Massberg, Steffen; Engelmann, Bernd

2017-01-01

Neutrophils, early mediators of the innate immune defense, are recruited to developing thrombi in different types of thrombosis. They amplify intravascular coagulation by stimulating the tissue factor-dependent extrinsic pathway via inactivation of endogenous anticoagulants, enhancing factor XII activation or decreasing plasmin generation. Neutrophil-dependent prothrombotic mechanisms are supported by the externalization of decondensed nucleosomes and granule proteins that together form neutrophil extracellular traps. These traps, either in intact or fragmented form, are causally involved in various forms of experimental thrombosis as first indicated by their role in the enhancement of both microvascular thrombosis during bacterial infection and carotid artery thrombosis. Neutrophil extracellular traps can be induced by interactions of neutrophils with activated platelets; vice versa, these traps enhance adhesion of platelets via von Willebrand factor. Neutrophil-induced microvascular thrombus formation can restrict the dissemination and survival of blood-borne bacteria and thereby sustain intravascular immunity. Dysregulation of this innate immune pathway may support sepsis-associated coagulopathies. Notably, neutrophils and extracellular nucleosomes, together with platelets, critically promote fibrin formation during flow restriction-induced deep vein thrombosis. Neutrophil extracellular traps/extracellular nucleosomes are increased in thrombi and in the blood of patients with different vaso-occlusive pathologies and could be therapeutically targeted for the prevention of thrombosis. Thus, during infections and in response to blood vessel damage, neutrophils and externalized nucleosomes are major promoters of intravascular blood coagulation and thrombosis. PMID:27927771

Hypoxia-inducible Factor-2α-dependent Hypoxic Induction of Wnt10b Expression in Adipogenic Cells*

PubMed Central

Park, Young-Kwon; Park, Bongju; Lee, Seongyeol; Choi, Kang; Moon, Yunwon; Park, Hyunsung

2013-01-01

Adipocyte hyperplasia and hypertrophy in obesity can lead to many changes in adipose tissue, such as hypoxia, metabolic dysregulation, and enhanced secretion of cytokines. In this study, hypoxia increased the expression of Wnt10b in both human and mouse adipogenic cells, but not in hypoxia-inducible factor (HIF)-2α-deficient adipogenic cells. Chromatin immunoprecipitation analysis revealed that HIF-2α, but not HIF-1α, bound to the Wnt10b enhancer region as well as upstream of the Wnt1 gene, which is encoded by an antisense strand of the Wnt10b gene. Hypoxia-conditioned medium (H-CM) induced phosphorylation of lipoprotein-receptor-related protein 6 as well as β-catenin-dependent gene expression in normoxic cells, which suggests that H-CM contains canonical Wnt signals. Furthermore, adipogenesis of both human mesenchymal stem cells and mouse preadipocytes was inhibited by H-CM even under normoxic conditions. These results suggest that O2 concentration gradients influence the formation of Wnt ligand gradients, which are involved in the regulation of pluripotency, cell proliferation, and cell differentiation. PMID:23900840

FXIa and platelet polyphosphate as therapeutic targets during human blood clotting on collagen/tissue factor surfaces under flow

PubMed Central

Zhu, Shu; Travers, Richard J.; Morrissey, James H.

2015-01-01

Factor XIIa (FXIIa) and factor XIa (FXIa) contribute to thrombosis in animal models, whereas platelet-derived polyphosphate (polyP) may potentiate contact or thrombin-feedback pathways. The significance of these mediators in human blood under thrombotic flow conditions on tissue factor (TF) –bearing surfaces remains inadequately resolved. Human blood (corn trypsin inhibitor treated [4 μg/mL]) was tested by microfluidic assay for clotting on collagen/TF at TF surface concentration ([TF]wall) from ∼0.1 to 2 molecules per μm2. Anti-FXI antibodies (14E11 and O1A6) or polyP-binding protein (PPXbd) were used to block FXIIa-dependent FXI activation, FXIa-dependent factor IX (FIX) activation, or platelet-derived polyP, respectively. Fibrin formation was sensitive to 14E11 at 0 to 0.1 molecules per µm2 and sensitive to O1A6 at 0 to 0.2 molecules per µm2. However, neither antibody reduced fibrin generation at ∼2 molecules per µm2 when the extrinsic pathway became dominant. Interestingly, PPXbd reduced fibrin generation at low [TF]wall (0.1 molecules per µm2) but not at zero or high [TF]wall, suggesting a role for polyP distinct from FXIIa activation and requiring low extrinsic pathway participation. Regardless of [TF]wall, PPXbd enhanced fibrin sensitivity to tissue plasminogen activator and promoted clot retraction during fibrinolysis concomitant with an observed PPXbd-mediated reduction of fibrin fiber diameter. This is the first detection of endogenous polyP function in human blood under thrombotic flow conditions. When triggered by low [TF]wall, thrombosis may be druggable by contact pathway inhibition, although thrombolytic susceptibility may benefit from polyP antagonism regardless of [TF]wall. PMID:26136249

Gender discrimination in the influence of hyperglycemia and hyperosmolarity on rat aortic tissue responses to insulin.

PubMed

Wong, Nikki L; Achike, Francis I

2010-08-09

Hyperglycaemia initiates endothelial dysfunction causing diabetic macro- and micro-vasculopathy, the main causes of morbidity and mortality in diabetes mellitus. The vasculopathy exhibits gender peculiarities. We therefore explored gender differences in comparing the effects of hyperglycaemia (50 mM) per se with its hyperosmolar (50 mM) effects on vascular tissue responses to insulin. Endothelium-intact or denuded thoracic aortic rings from age-matched male and female Sprague-Dawley rats were incubated for 10 min or 6 h (acute versus chronic exposure) in normal, hyperglycaemic or hyperosmolar Krebs solution. Relaxant responses to insulin (6.9x10(-7)-6.9x10(-5) M) of the phenylephrine-contracted tissues were recorded. Endothelium denudation in both genders inhibited relaxation to insulin in all conditions, more significantly in female than in male tissues, suggesting the female response to insulin is more endothelium-dependent than the male. Acutely and chronically exposed normoglycemic endothelium-intact or -denuded tissues responded similarly to insulin. Chronic hyperglycemic or hyperosmolar exposure did not alter the endothelium-denuded tissue responses to insulin, whereas the responses of the endothelium-intact male and female hyperosmolar, and male hyperglycemic tissues were enhanced. The results show that insulin exerts an endothelium-dependent and independent relaxation with the female tissue responses more endothelium-dependent than the male. The data also suggest that hyperosmolarity per se enhances aortic tissue relaxant responses to insulin whereas hyperglycemia per se inhibits the same and more so in female than male tissues. These effects are endothelium-dependent. Copyright (c) 2010 Elsevier B.V. All rights reserved.

Fibrin, γ’-fibrinogen, and trans-clot pressure gradient control hemostatic clot growth during human blood flow over a collagen/tissue factor wound

PubMed Central

Muthard, Ryan W.; Welsh, John D.; Brass, Lawrence F.; Diamond, Scott L.

2015-01-01

SUMMARY Objective Biological and physical factors interact to modulate blood response in a wounded vessel, resulting in a hemostatic clot or an occlusive thrombus. Flow and pressure differential (ΔP) across the wound from the lumen to the extravascular compartment may impact hemostasis and the observed core/shell architecture. We examined physical and biological factors responsible for regulating thrombin mediated clot growth. Approach and Results Using factor XIIa-inhibited human whole blood perfused in a microfluidic device over collagen/tissue factor at controlled wall shear rate and ΔP, we found thrombin to be highly localized in the P-selectin+ core of hemostatic clots. Increasing ΔP from 9 to 29 mm-Hg (wall shear rate = 400 s−1) reduced P-selectin+ core size and total clot size due to enhanced extravasation of thrombin. Blockade of fibrin polymerization with 5 mM GPRP dysregulated hemostasis by enhancing both P-selectin+ core size and clot size at 400 s−1 (20 mm-Hg). For whole blood flow (no GPRP), the thickness of the P-selectin-negative shell was reduced under arterial conditions (2000 s−1, 20 mm-Hg). Consistent with the antithrombin-1 activity of fibrin implicated with GPRP, anti-γ’-fibrinogen antibody enhanced core-localized thrombin, core size, and overall clot size, especially at venous (100 s−1) but not arterial wall shear rates (2000 s−1). Pathological shear (15,000 s−1) and GPRP synergized to exacerbate clot growth. Conclusions Hemostatic clotting was dependent on core-localized thrombin that (1) triggered platelet P-selectin display and (2) was highly regulated by fibrin and the trans-clot ΔP. Also, γ’-fibrinogen had a role in venous but not arterial conditions. PMID:25614284

Inhibition of thrombin action ameliorates insulin resistance in type 2 diabetic db/db mice.

PubMed

Mihara, Masatomo; Aihara, Ken-ichi; Ikeda, Yasumasa; Yoshida, Sumiko; Kinouchi, Mizuho; Kurahashi, Kiyoe; Fujinaka, Yuichi; Akaike, Masashi; Matsumoto, Toshio

2010-02-01

The binding of thrombin to its receptor stimulates inflammatory cytokines including IL-6 and monocyte chemoattractant protein-1 (MCP-1); both are associated with the development of insulin resistance. Because increased adiposity enhanced the expression of coagulation factor VII that stimulates the coagulation pathway in adipose tissue, we tested whether the inhibition of thrombin action ameliorates insulin resistance in obese diabetic (Lpr(-/-):db/db) mice. The 4-wk administration of argatroban, a selective thrombin inhibitor, reduced fasting plasma glucose and ameliorated insulin resistance in these mice. It also reduced adipocyte size and macrophage infiltration into adipose tissue. The aberrant gene expression of MCP-1, IL-6, adiponectin, and factor VII and suppressed insulin receptor substrate-1-Akt signaling in adipose tissue of db/db mice were reversed by argatroban treatment. These results demonstrate that increased adiposity enhances the production of thrombin in adipose tissue by stimulating factor VII expression and suggest that increased thrombin activity in adipose tissue plays an important role in the development of insulin resistance via enhancing MCP-1 production, leading to macrophage infiltration and insulin receptor substrate-1-Akt pathway inactivation.

Minimally invasive esthetic ridge preservation with growth-factor enhanced bone matrix.

PubMed

Nevins, Marc L; Said, Sherif

2017-12-28

Extraction socket preservation procedures are critical to successful esthetic implant therapy. Conventional surgical approaches are technique sensitive and often result in alteration of the soft tissue architecture, which then requires additional corrective surgical procedures. This case series report presents the ability of flapless surgical techniques combined with a growth factor-enhanced bone matrix to provide esthetic ridge preservation at the time of extraction for compromised sockets. When considering esthetic dental implant therapy, preservation, or further enhancement of the available tissue support at the time of tooth extraction may provide an improved esthetic outcome with reduced postoperative sequelae and decreased treatment duration. Advances in minimally invasive surgical techniques combined with recombinant growth factor technology offer an alternative for bone reconstruction while maintaining the gingival architecture for enhanced esthetic outcome. The combination of freeze-dried bone allograft (FDBA) and rhPDGF-BB (platelet-derived growth factor-BB) provides a growth-factor enhanced matrix to induce bone and soft tissue healing. The use of a growth-factor enhanced matrix is an option for minimally invasive ridge preservation procedures for sites with advanced bone loss. Further studies including randomized clinical trials are needed to better understand the extent and limits of these procedures. The use of minimally invasive techniques with growth factors for esthetic ridge preservation reduces patient morbidity associated with more invasive approaches and increases the predictability for enhanced patient outcomes. By reducing the need for autogenous bone grafts the use of this technology is favorable for patient acceptance and ease of treatment process for esthetic dental implant therapy. © 2017 Wiley Periodicals, Inc.

Inherent Contrast in Magnetic Resonance Imaging and the Potential for Contrast Enhancement

PubMed Central

Brasch, Robert C.

1985-01-01

Magnetic resonance (MR) imaging is emerging as a powerful new diagnostic tool valued for its apparent lack of adverse effects. The excellent inherent contrast between biologic tissues and fluids afforded by MR imaging is one of the foremost characteristics of this technique and depends on physicochemical properties such as hydrogen density and T1 and T2 relaxation rates, on magnetic field strength and on operator-chosen factors for acquiring the MR imaging signal. Pharmaceutical contrast-enhancing agents shorten the MR imaging process and improve sensitivity and diagnostic accuracy. ImagesFigure 1.Figure 2.Figure 3.Figure 4.Figure 5.Figure 6.Figure 8.Figure 9.Figure 10.Figure 11. PMID:2992172

Synergism between a half-site and an imperfect estrogen-responsive element, and cooperation with COUP-TFI are required for estrogen receptor (ER) to achieve a maximal estrogen-stimulation of rainbow trout ER gene.

PubMed

Petit, F G; Métivier, R; Valotaire, Y; Pakdel, F

1999-01-01

In all oviparous, liver represents one of the main E2-target tissues where estrogen receptor (ER) constitutes the key mediator of estrogen action. The rainbow trout estrogen receptor (rtER) gene expression is markedly up-regulated by estrogens and the sequences responsible for this autoregulation have been located in a 0.2 kb upstream transcription start site within - 40/- 248 enhancer region. Absence of interference with steroid hormone receptors and tissue-specific factors and a conserved basal transcriptional machinery between yeast and higher eukaryotes, make yeast a simple assay system that will enable determination of important cis-acting regulatory sequences within rtER gene promoter and identification of transcription factors implicated in the regulation of this gene. Deletion analysis allowed to show a synergistic effect between an imperfect estrogen-responsive element (ERE) and a consensus half-ERE to achieve a high hormone-dependent transcriptional activation of the rtER gene promoter in the presence of stably expressed rtER. As in mammalian cells, here we observed a positive regulation of the rtER gene promoter by the chicken ovalbumin upstream promoter-transcription factor I (COUP-TFI) through enhancing autoregulation. Using a point mutation COUP-TFI mutant unable to bind DNA demonstrates that enhancement of rtER gene autoregulation requires the interaction of COUP-TFI to the DNA. Moreover, this enhancement of transcriptional activation by COUP-TFI requires specifically the AF-1 transactivation function of ER and can be observed in the presence of E2 or 4-hydroxytamoxifen but not ICI 164384. Thus, this paper describes the reconstitution of a hormone-responsive transcription unit in yeast in which the regulation of rtER gene promoter could be enhanced by the participation of cis-elements and/or trans-acting factors, such as ER itself or COUP-TF.

Delivery of Brain-Derived Neurotrophic Factor by 3D Biocompatible Polymeric Scaffolds for Neural Tissue Engineering and Neuronal Regeneration.

PubMed

Limongi, T; Rocchi, A; Cesca, F; Tan, H; Miele, E; Giugni, A; Orlando, M; Perrone Donnorso, M; Perozziello, G; Benfenati, Fabio; Di Fabrizio, Enzo

2018-03-29

Biopolymers are increasingly employed for neuroscience applications as scaffolds to drive and promote neural regrowth, thanks to their ability to mediate the upload and subsequent release of active molecules and drugs. Synthetic degradable polymers are characterized by different responses ranging from tunable distension or shrinkage to total dissolution, depending on the function they are designed for. In this paper we present a biocompatible microfabricated poly-ε-caprolactone (PCL) scaffold for primary neuron growth and maturation that has been optimized for the in vitro controlled release of brain-derived neurotrophic factor (BDNF). We demonstrate that the designed morphology confers to these devices an enhanced drug delivery capability with respect to monolithic unstructured supports. After incubation with BDNF, micropillared PCL devices progressively release the neurotrophin over 21 days in vitro. Moreover, the bioactivity of released BDNF is confirmed using primary neuronal cultures, where it mediates a consistent activation of BDNF signaling cascades, increased synaptic density, and neuronal survival. These results provide the proof-of-principle on the fabrication process of micropatterned PCL devices, which represent a promising therapeutic option to enhance neuronal regeneration after lesion and for neural tissue engineering and prosthetics.

The human tissue-biomaterial interface: a role for PPARγ-dependent glucocorticoid receptor activation in regulating the CD163+ M2 macrophage phenotype.

PubMed

Bullers, Samuel J; Baker, Simon C; Ingham, Eileen; Southgate, Jennifer

2014-09-01

In vivo studies of implanted acellular biological scaffolds in experimental animals have shown constructive remodeling mediated by anti-inflammatory macrophages. Little is known about the human macrophage response to such biomaterials, or the nature of the signaling mechanisms that govern the macrophage phenotype in this environment. The cellular events at the interface of a tissue and implanted decellularized biomaterial were examined by establishing a novel ex vivo tissue culture model in which surgically excised human urinary tract tissue was combined with porcine acellular bladder matrix (PABM). Evaluation of the tissue-biomaterial interface showed a time-dependent infiltration of the biomaterial by CD68(+) CD80(-) macrophages. The migration of CD68(+) cells from the tissue to the interface was accompanied by maturation to a CD163(hi) phenotype, suggesting that factor(s) associated with the biomaterial or the wound edge was/were responsible for the active recruitment and polarization of local macrophages. Glucocorticoid receptor (GR) and peroxisome proliferator activated receptor gamma (PPARγ) signaling was investigated as candidate pathways for integrating inflammatory responses; both showed intense nuclear labeling in interface macrophages. GR and PPARγ activation polarized peripheral blood-derived macrophages from a default M1 (CD80(+)) toward an M2 (CD163(+)) phenotype, but PPARγ signaling predominated, as its antagonism blocked any GR-mediated effect. Seeding on PABM was effective at polarizing peripheral blood-derived macrophages from a default CD80(+) phenotype on glass to a CD80(-) phenotype, with intense nuclear localization of PPARγ. These results endorse in vivo observations that the infiltration of decellularized biological scaffolds, exemplified here by PABM, is pioneered by macrophages. Thus, it appears that natural factors present in PABM are involved in the active recruitment and polarization of macrophages to a CD163(+) phenotype, with activation of PPARγ identified as the candidate pathway. The harnessing of these natural matrix-associated factors may be useful in enhancing the integration of synthetic and other natural biomaterials by polarizing macrophage activation toward an M2 regulatory phenotype.

Dose Enhancement near Metal Interfaces in Synthetic Diamond Based X-ray Dosimeters

NASA Astrophysics Data System (ADS)

Alamoudi, Dalal

Diamond is an attractive material for medical dosimetry due to its radiation hardness, fast response, chemical resilience, small sensitive volume, high spatial resolution, near-tissue equivalence, and energy and dose rate independence. These properties make diamond a promising material for medical dosimetry compared to other semiconductor detector materials and wider radiation detection applications. This study is focused on one of the important factors to consider in the radiation detector; the influence of dose enhancement on the photocurrent performance at metallic interfaces in synthetic diamond radiation dosimeters with carbon based electrodes as a function of bias voltages. Monte Carlo (MC) simulations with BEAMnrc code were carried out to simulate the dose enhancement factor (DEF) and compared against the equivalent photocurrent ratio from experimental investigation. MC simulations show that the sensitive region for the absorbed dose distribution covers a few micrometers distances from the interface. Experimentally, two single crystal (SC) and one polycrystalline (PC) samples with carbon based electrodes were used. The samples were each mounted inside a tissue equivalent encapsulation design in order to minimize fluence perturbations. Copper, Gold and Lead have been investigated experimentally as generators of photoelectrons using 50 kVp and 100 kVp X-rays relevant for medical dosimetry. The results show enhancement in the detectors' photocurrent performance when different metals are butted up to the diamond detector. The variation in the photocurrent ratio measurements depends on the type of diamond samples, their electrode fabrication and the applied bias voltages indicating that the dose enhancement from diamond-metal interface modifies the electronic performance of the detector.

The tumour suppressor CDKN2A/p16INK4a regulates adipogenesis and bone marrow-dependent development of perivascular adipose tissue

PubMed Central

Wouters, Kristiaan; Deleye, Yann; Hannou, Sarah A; Vanhoutte, Jonathan; Maréchal, Xavier; Coisne, Augustin; Tagzirt, Madjid; Derudas, Bruno; Bouchaert, Emmanuel; Duhem, Christian; Vallez, Emmanuelle; Schalkwijk, Casper G; Pattou, François; Montaigne, David; Staels, Bart; Paumelle, Réjane

2017-01-01

The genomic CDKN2A/B locus, encoding p16INK4a among others, is linked to an increased risk for cardiovascular disease and type 2 diabetes. Obesity is a risk factor for both cardiovascular disease and type 2 diabetes. p16INK4a is a cell cycle regulator and tumour suppressor. Whether it plays a role in adipose tissue formation is unknown. p16INK4a knock-down in 3T3/L1 preadipocytes or p16INK4a deficiency in mouse embryonic fibroblasts enhanced adipogenesis, suggesting a role for p16INK4a in adipose tissue formation. p16INK4a-deficient mice developed more epicardial adipose tissue in response to the adipogenic peroxisome proliferator activated receptor gamma agonist rosiglitazone. Additionally, adipose tissue around the aorta from p16INK4a-deficient mice displayed enhanced rosiglitazone-induced gene expression of adipogenic markers and stem cell antigen, a marker of bone marrow-derived precursor cells. Mice transplanted with p16INK4a-deficient bone marrow had more epicardial adipose tissue compared to controls when fed a high-fat diet. In humans, p16INK4a gene expression was enriched in epicardial adipose tissue compared to other adipose tissue depots. Moreover, epicardial adipose tissue from obese humans displayed increased expression of stem cell antigen compared to lean controls, supporting a bone marrow origin of epicardial adipose tissue. These results show that p16INK4a modulates epicardial adipose tissue development, providing a potential mechanistic link between the genetic association of the CDKN2A/B locus and cardiovascular disease risk. PMID:28868898

Growth enhancement by embryonic fibroblasts upon cotransplantation of noncommitted pig embryonic tissues with fully committed organs.

PubMed

Cohen, Sivan; Tchorsh-Yutsis, Dalit; Aronovich, Anna; Tal, Orna; Eventov-Friedman, Smadar; Katchman, Helena; Klionsky, Yael; Shezen, Elias; Reisner, Yair

2010-05-27

We recently defined the optimal gestational time windows for the transplantation of several embryonic tissues. We showed that the liver and kidney obtained from E28 pig embryos can grow and differentiate normally after transplantation, whereas 1 week earlier in gestation, these tissues develop into teratoma-like structures or fibrotic mass. In this study, we investigated whether cotransplantation of E28 with E21 tissue could control its tumorogenic potential, or alternatively whether the stem cells derived from the earlier tissue contribute to the growth of the more committed one. Pig embryonic precursors from E21 and E28 gestational age were transplanted alone or together, into nonobese diabetic/severe combined immunodeficiency mice, and their growth and differentiation was evaluated by immunohistology. In situ analysis, based on sex disparity between the E21 and E28 tissues, was used to identify the tissue source. In some experiments, mouse embryonic fibroblasts (MEF) were cotransplanted with E28 liver, and their effect was evaluated. E28 tissues could not abrogate the propensity of the cells within the undifferentiated tissue to form teratoma-like structures. However, E21 kidney or liver tissue markedly enhanced the growth and function of E28 kidney, liver, and heart grafts. Moreover, similar growth enhancement was observed on coimplantation of E28 liver tissue with MEF or on infusion of MEF culture medium, indicating that this enhancement is likely mediated through soluble factors secreted by the fibroblasts. Our results suggest a novel approach for the enhancement of growth and differentiation of transplanted embryonic tissues by the use of soluble factors secreted by embryonic fibroblasts.

Cartilage constructs engineered from chondrocytes overexpressing IGF-I improve the repair of osteochondral defects in a rabbit model.

PubMed

Madry, H; Kaul, G; Zurakowski, D; Vunjak-Novakovic, G; Cucchiarini, M

2013-04-16

Tissue engineering combined with gene therapy is a promising approach for promoting articular cartilage repair. Here, we tested the hypothesis that engineered cartilage with chondrocytes overexpressing a human insulin-like growth factor I (IGF-I) gene can enhance the repair of osteochondral defects, in a manner dependent on the duration of cultivation. Genetically modified chondrocytes were cultured on biodegradable polyglycolic acid scaffolds in dynamic flow rotating bioreactors for either 10 or 28 d. The resulting cartilaginous constructs were implanted into osteochondral defects in rabbit knee joints. After 28 weeks of in vivo implantation, immunoreactivity to ß-gal was detectable in the repair tissue of defects that received lacZ constructs. Engineered cartilaginous constructs based on IGF-I-overexpressing chondrocytes markedly improved osteochondral repair compared with control (lacZ) constructs. Moreover, IGF-I constructs cultivated for 28 d in vitro significantly promoted osteochondral repair vis-à-vis similar constructs cultivated for 10 d, leading to significantly decreased osteoarthritic changes in the cartilage adjacent to the defects. Hence, the combination of spatially defined overexpression of human IGF-I within a tissue-engineered construct and prolonged bioreactor cultivation resulted in most enhanced articular cartilage repair and reduction of osteoarthritic changes in the cartilage adjacent to the defect. Such genetically enhanced tissue engineering provides a versatile tool to evaluate potential therapeutic genes in vivo and to improve our comprehension of the development of the repair tissue within articular cartilage defects. Insights gained with additional exploration using this model may lead to more effective treatment options for acute cartilage defects.

CARTILAGE CONSTRUCTS ENGINEERED FROM CHONDROCYTES OVEREXPRESSING IGF-I IMPROVE THE REPAIR OF OSTEOCHONDRAL DEFECTS IN A RABBIT MODEL

PubMed Central

Madry, Henning; Kaul, Gunter; Zurakowski, David; Vunjak-Novakovic, Gordana; Cucchiarini, Magali

2015-01-01

Tissue engineering combined with gene therapy is a promising approach for promoting articular cartilage repair. Here, we tested the hypothesis that engineered cartilage with chondrocytes over expressing a human insulin-like growth factor I (IGF-I) gene can enhance the repair of osteochondral defects, in a manner dependent on the duration of cultivation. Genetically modified chondrocytes were cultured on biodegradable polyglycolic acid scaffolds in dynamic flow rotating bioreactors for either 10 or 28 d. The resulting cartilaginous constructs were implanted into osteochondral defects in rabbit knee joints. After 28 weeks of in vivo implantation, immunoreactivity to ß-gal was detectable in the repair tissue of defects that received lacZ constructs. Engineered cartilaginous constructs based on IGF-I-over expressing chondrocytes markedly improved osteochondral repair compared with control (lacZ) constructs. Moreover, IGF-I constructs cultivated for 28 d in vitro significantly promoted osteochondral repair vis-à-vis similar constructs cultivated for 10 d, leading to significantly decreased osteoarthritic changes in the cartilage adjacent to the defects. Hence, the combination of spatially defined overexpression of human IGF-I within a tissue-engineered construct and prolonged bioreactor cultivation resulted in most enhanced articular cartilage repair and reduction of osteoarthritic changes in the cartilage adjacent to the defect. Such genetically enhanced tissue engineering provides a versatile tool to evaluate potential therapeutic genes in vivo and to improve our comprehension of the development of the repair tissue within articular cartilage defects. Insights gained with additional exploration using this model may lead to more effective treatment options for acute cartilage defects. PMID:23588785

Tissue mimicking simulations for temporal enhanced ultrasound-based tissue typing

NASA Astrophysics Data System (ADS)

Bayat, Sharareh; Imani, Farhad; Gerardo, Carlos D.; Nir, Guy; Azizi, Shekoofeh; Yan, Pingkun; Tahmasebi, Amir; Wilson, Storey; Iczkowski, Kenneth A.; Lucia, M. Scott; Goldenberg, Larry; Salcudean, Septimiu E.; Mousavi, Parvin; Abolmaesumi, Purang

2017-03-01

Temporal enhanced ultrasound (TeUS) is an imaging approach where a sequence of temporal ultrasound data is acquired and analyzed for tissue typing. Previously, in a series of in vivo and ex vivo studies we have demonstrated that, this approach is effective for detecting prostate and breast cancers. Evidences derived from our experiments suggest that both ultrasound-signal related factors such as induced heat and tissue-related factors such as the distribution and micro-vibration of scatterers lead to tissue typing information in TeUS. In this work, we simulate mechanical micro-vibrations of scatterers in tissue-mimicking phantoms that have various scatterer densities reflecting benign and cancerous tissue structures. Finite element modeling (FEM) is used for this purpose where the vertexes are scatterers representing cell nuclei. The initial positions of scatterers are determined by the distribution of nuclei segmented from actual digital histology scans of prostate cancer patients. Subsequently, we generate ultrasound images of the simulated tissue structure using the Field II package resulting in a temporal enhanced ultrasound. We demonstrate that the micro-vibrations of scatterers are captured by temporal ultrasound data and this information can be exploited for tissue typing.

Time- and dose-related interactions between glucocorticoid and cyclic adenosine 3',5'-monophosphate on CCAAT/enhancer-binding protein-dependent insulin-like growth factor I expression by osteoblasts

NASA Technical Reports Server (NTRS)

McCarthy, T. L.; Ji, C.; Chen, Y.; Kim, K.; Centrella, M.

2000-01-01

Glucocorticoid has complex effects on osteoblasts. Several of these changes appear to be related to steroid concentration, duration of exposure, or specific effects on growth factor expression or activity within bone. One important bone growth factor, insulin-like growth factor I (IGF-I), is induced in osteoblasts by hormones such as PGE2 that increase intracellular cAMP levels. In this way, PGE2 activates transcription factor CCAAT/enhancer-binding protein-delta (C/EBPdelta) and enhances its binding to a specific control element found in exon 1 in the IGF-I gene. Our current studies show that preexposure to glucocorticoid enhanced C/EBPdelta and C/EBPbeta expression by osteoblasts and thereby potentiated IGF-I gene promoter activation in response to PGE2. Importantly, this directly contrasts with inhibitory effects on IGF-I expression that result from sustained or pharmacologically high levels of glucocorticoid exposure. Consistent with the stimulatory effect of IGF-I on bone protein synthesis, pretreatment with glucocorticoid sensitized osteoblasts to PGE2, and in this context significantly enhanced new collagen and noncollagen protein synthesis. Therefore, pharmacological levels of glucocorticoid may reduce IGF-I expression by osteoblasts and cause osteopenic disease, whereas physiological transient increases in glucocorticoid may permit or amplify the effectiveness of hormones that regulate skeletal tissue integrity. These events appear to converge on the important role of C/EBPdelta and C/EBPbeta on IGF-I expression by osteoblasts.

Enhanced HIV-1 replication in ex vivo ectocervical tissues from post-menopausal women correlates with increased inflammatory responses.

PubMed

Rollenhagen, C; Asin, S N

2011-11-01

Knowledge about early innate immune responses at the mucosal surfaces of the female genital tract is important in understanding the pathogenesis of heterosexual transmission of human immunodeficiency virus type-1 (HIV-1). As estradiol decreases inflammatory responses, we postulated that an estradiol-deficient state such as post-menopause could enhance expression of inflammatory factors that stimulate HIV-1 replication. We compare HIV-1 integration, transcription, and viral p24 release levels among ectocervical tissues obtained from pre- and post-menopausal donors. We detected enhanced HIV-1 p24 release levels in post- compared with pre-menopausal tissues (P<0.0001), but saw no difference in HIV-1 integration. Overall, 100% of post-menopausal tissues exhibited levels of HIV-1 transcription above background compared with only 60% of pre-menopausal tissues. Increased HIV-1 transcription was associated with enhanced interleukin (IL)-1β, IL-6, monocyte chemotactic protein-1, growth-regulated oncogene-α, and interferon-γ-inducible protein-10 expression. Neutralization and nuclear factor-κB-targeting small-interfering RNA experiments both decreased HIV-1 transcription, suggesting that the early inflammatory response may facilitate HIV-1 replication in ex vivo ectocervical tissues from post-menopausal women.

EGFR Ligands Drive Multipotential Stromal Cells to Produce Multiple Growth Factors and Cytokines via Early Growth Response-1

PubMed Central

Kerpedjieva, Svetoslava S.; Kim, Duk Soo; Barbeau, Dominique J.

2012-01-01

Cell therapy with adult bone marrow multipotential stromal cells/mesenchymal stem cells (MSCs) presents a promising approach to promote wound healing and tissue regeneration. The strong paracrine capability of various growth factors and cytokines is a key mechanism of MSC-mediated wound healing and tissue regeneration, and the goal of this study is to understand the underlying mechanism that supports the strong paracrine machineries in MSCs. Microarray database analyses revealed that early growth response-1 (EGR1) is highly expressed in MSCs. Our previous studies showed that epidermal growth factor (EGF) treatment induces growth factor production in MSCs in vitro. Since EGF strongly upregulates EGR1, we hypothesized that EGF receptor (EGFR)–EGR1 signaling plays a pivotal role in MSC paracrine activity. EGF treatment upregulated the gene expression of growth factors and cytokines, including EGFR ligands in a protein kinase C (PKC)- and/or mitogen-activated protein kinase–extracellular-signal-regulated kinase-dependent manner, and it was reversed by shRNA against EGR1. PKC activator phorbol 12-myristate 13-acetate enhanced EGFR tyrosyl phosphorylation and upregulated the gene expression of growth factors and cytokines in a heparin-binding EGF-like growth factor (HBEGF) inhibitor CRM197 sensitive manner, indicating an involvement of autocrined HBEGF in the downstream of PKC signaling. Moreover, stimulation with growth factors and cytokines induced the expression of EGFR ligands, presumably via EGR1 upregulation. These data indicate EGR1 as a convergence point of multiple signaling pathways, which in turn augments the production of multiple growth factors and cytokines by enhancing the autocrine signaling with EGFR ligands. PMID:22316125

EGFR ligands drive multipotential stromal cells to produce multiple growth factors and cytokines via early growth response-1.

PubMed

Kerpedjieva, Svetoslava S; Kim, Duk Soo; Barbeau, Dominique J; Tamama, Kenichi

2012-09-01

Cell therapy with adult bone marrow multipotential stromal cells/mesenchymal stem cells (MSCs) presents a promising approach to promote wound healing and tissue regeneration. The strong paracrine capability of various growth factors and cytokines is a key mechanism of MSC-mediated wound healing and tissue regeneration, and the goal of this study is to understand the underlying mechanism that supports the strong paracrine machineries in MSCs. Microarray database analyses revealed that early growth response-1 (EGR1) is highly expressed in MSCs. Our previous studies showed that epidermal growth factor (EGF) treatment induces growth factor production in MSCs in vitro. Since EGF strongly upregulates EGR1, we hypothesized that EGF receptor (EGFR)-EGR1 signaling plays a pivotal role in MSC paracrine activity. EGF treatment upregulated the gene expression of growth factors and cytokines, including EGFR ligands in a protein kinase C (PKC)- and/or mitogen-activated protein kinase-extracellular-signal-regulated kinase-dependent manner, and it was reversed by shRNA against EGR1. PKC activator phorbol 12-myristate 13-acetate enhanced EGFR tyrosyl phosphorylation and upregulated the gene expression of growth factors and cytokines in a heparin-binding EGF-like growth factor (HBEGF) inhibitor CRM197 sensitive manner, indicating an involvement of autocrined HBEGF in the downstream of PKC signaling. Moreover, stimulation with growth factors and cytokines induced the expression of EGFR ligands, presumably via EGR1 upregulation. These data indicate EGR1 as a convergence point of multiple signaling pathways, which in turn augments the production of multiple growth factors and cytokines by enhancing the autocrine signaling with EGFR ligands.

Vitamin D modulates tissue factor and protease-activated receptor 2 expression in vascular smooth muscle cells.

PubMed

Martinez-Moreno, Julio M; Herencia, Carmen; Montes de Oca, Addy; Muñoz-Castañeda, Juan R; Rodríguez-Ortiz, M Encarnación; Díaz-Tocados, Juan M; Peralbo-Santaella, Esther; Camargo, Antonio; Canalejo, Antonio; Rodriguez, Mariano; Velasco-Gimena, Francisco; Almaden, Yolanda

2016-03-01

Clinical and epidemiologic studies reveal an association between vitamin D deficiency and increased risk of cardiovascular disease. Because vascular smooth muscle cell (VSMC)-derived tissue factor (TF) is suggested to be critical for arterial thrombosis, we investigated whether the vitamin D molecules calcitriol and paricalcitol could reduce the expression of TF induced by the proinflammatory cytokine TNF-α in human aortic VSMCs. We found that, compared with controls, incubation with TNF-α increased TF expression and procoagulant activity in a NF-κB-dependent manner, as deduced from the increased nuclear translocation of nuclear factor κ-light-chain-enhancer of activated B cells protein 65 (p65-NF-κB) and direct interaction of NF-κB to the TF promoter. This was accompanied by the up-regulation of TF signaling mediator protease-activated receptor 2 (PAR-2) expression and by the down-regulation of vitamin D receptor expression in a miR-346-dependent way. However, addition of calcitriol or paricalcitol blunted the TNF-α-induced TF expression and activity (2.01 ± 0.24 and 1.32 ± 0.14 vs. 3.02 ± 0.39 pmol/mg protein, P < 0.05), which was associated with down-regulation of NF-κB signaling and PAR-2 expression, as well as with restored levels of vitamin D receptor and enhanced expression of TF pathway inhibitor. Our data suggest that inflammation promotes a prothrombotic state through the up-regulation of TF function in VSMCs and that the beneficial cardiovascular effects of vitamin D may be partially due to decreases in TF expression and its activity in VSMCs. © FASEB.

General equation for the differential pathlength factor of the frontal human head depending on wavelength and age.

PubMed

Scholkmann, Felix; Wolf, Martin

2013-10-01

Continuous-wave near-infrared spectroscopy and near-infrared imaging enable the measurement of relative concentration changes in oxy- and deoxyhemoglobin and thus hemodynamics and oxygenation. The accuracy of determined changes depends mainly on the modeling of the light transport through the probed tissue. Due to the highly scattering nature of tissue, the light path is longer than the source-detector separation (d). This is incorporated in modeling by multiplying d by a differential pathlength factor (DPF) which depends on several factors such as wavelength, age of the subject, and type of tissue. In the present work, we derive a general DPF equation for the frontal human head, incorporating dependency on wavelength and age, based on published data. We validated the equation using different data sets of experimentally determined DPFs from six independent studies.

Computational Identification of Tissue-Specific Splicing Regulatory Elements in Human Genes from RNA-Seq Data.

PubMed

Badr, Eman; ElHefnawi, Mahmoud; Heath, Lenwood S

2016-01-01

Alternative splicing is a vital process for regulating gene expression and promoting proteomic diversity. It plays a key role in tissue-specific expressed genes. This specificity is mainly regulated by splicing factors that bind to specific sequences called splicing regulatory elements (SREs). Here, we report a genome-wide analysis to study alternative splicing on multiple tissues, including brain, heart, liver, and muscle. We propose a pipeline to identify differential exons across tissues and hence tissue-specific SREs. In our pipeline, we utilize the DEXSeq package along with our previously reported algorithms. Utilizing the publicly available RNA-Seq data set from the Human BodyMap project, we identified 28,100 differentially used exons across the four tissues. We identified tissue-specific exonic splicing enhancers that overlap with various previously published experimental and computational databases. A complicated exonic enhancer regulatory network was revealed, where multiple exonic enhancers were found across multiple tissues while some were found only in specific tissues. Putative combinatorial exonic enhancers and silencers were discovered as well, which may be responsible for exon inclusion or exclusion across tissues. Some of the exonic enhancers are found to be co-occurring with multiple exonic silencers and vice versa, which demonstrates a complicated relationship between tissue-specific exonic enhancers and silencers.

Intestinal Master Transcription Factor CDX2 Controls Chromatin Access for Partner Transcription Factor Binding

PubMed Central

Verzi, Michael P.; Shin, Hyunjin; San Roman, Adrianna K.

2013-01-01

Tissue-specific gene expression requires modulation of nucleosomes, allowing transcription factors to occupy cis elements that are accessible only in selected tissues. Master transcription factors control cell-specific genes and define cellular identities, but it is unclear if they possess special abilities to regulate cell-specific chromatin and if such abilities might underlie lineage determination and maintenance. One prevailing view is that several transcription factors enable chromatin access in combination. The homeodomain protein CDX2 specifies the embryonic intestinal epithelium, through unknown mechanisms, and partners with transcription factors such as HNF4A in the adult intestine. We examined enhancer chromatin and gene expression following Cdx2 or Hnf4a excision in mouse intestines. HNF4A loss did not affect CDX2 binding or chromatin, whereas CDX2 depletion modified chromatin significantly at CDX2-bound enhancers, disrupted HNF4A occupancy, and abrogated expression of neighboring genes. Thus, CDX2 maintains transcription-permissive chromatin, illustrating a powerful and dominant effect on enhancer configuration in an adult tissue. Similar, hierarchical control of cell-specific chromatin states is probably a general property of master transcription factors. PMID:23129810

Tissue-Specific Regulation of Chromatin Insulator Function

PubMed Central

Matzat, Leah H.; Dale, Ryan K.; Moshkovich, Nellie; Lei, Elissa P.

2012-01-01

Chromatin insulators organize the genome into distinct transcriptional domains and contribute to cell type–specific chromatin organization. However, factors regulating tissue-specific insulator function have not yet been discovered. Here we identify the RNA recognition motif-containing protein Shep as a direct interactor of two individual components of the gypsy insulator complex in Drosophila. Mutation of shep improves gypsy-dependent enhancer blocking, indicating a role as a negative regulator of insulator activity. Unlike ubiquitously expressed core gypsy insulator proteins, Shep is highly expressed in the central nervous system (CNS) with lower expression in other tissues. We developed a novel, quantitative tissue-specific barrier assay to demonstrate that Shep functions as a negative regulator of insulator activity in the CNS but not in muscle tissue. Additionally, mutation of shep alters insulator complex nuclear localization in the CNS but has no effect in other tissues. Consistent with negative regulatory activity, ChIP–seq analysis of Shep in a CNS-derived cell line indicates substantial genome-wide colocalization with a single gypsy insulator component but limited overlap with intact insulator complexes. Taken together, these data reveal a novel, tissue-specific mode of regulation of a chromatin insulator. PMID:23209434

Dose-dependent collagen cross-linking of rabbit scleral tissue by blue light and riboflavin treatment probed by dynamic shear rheology.

PubMed

Schuldt, Carsten; Karl, Anett; Körber, Nicole; Koch, Christian; Liu, Qing; Fritsch, Anatol W; Reichenbach, Andreas; Wiedemann, Peter; Käs, Josef A; Francke, Mike; Iseli, Hans Peter

2015-08-01

To determine the visco-elastic properties of isolated rabbit scleral tissue and dose-dependent biomechanical and morphological changes after collagen cross-linking by riboflavin/blue light treatment. Scleral patches from 87 adult albino rabbit eyes were examined by dynamic shear rheology. Scleral patches were treated by riboflavin and different intensities of blue light (450 nm), and the impact on the visco-elastic properties was determined by various rheological test regimes. The relative elastic modulus was calculated from non-treated and corresponding treated scleral patches, and treatments with different blue light intensities were compared. Shear rheology enables us to study the material properties of scleral tissue within physiological relevant parameters. Cross-linking treatment increased the viscous as well as the elastic modulus and changed the ratio of the elastic versus viscous proportion in scleral tissue. Constant riboflavin application combined with different blue light intensities from 12 mW/cm(2) up to 100 mW/cm(2) increased the relative elastic modulus of scleral tissue by factors up to 1.8. Further enhancement of the applied light intensity caused a decline of the relative elastic modulus. This might be due to destructive changes of the collagen bundle structure at larger light intensities, as observed by histological examination. Collagen cross-linking by riboflavin/blue light application increases the biomechanical stiffness of the sclera in a dose-dependent manner up to certain light intensities. Therefore, this treatment might be a suitable therapeutic approach to stabilize the biomechanical properties of scleral tissue in cases of pathological eye expansion. © 2014 Acta Ophthalmologica Scandinavica Foundation. Published by John Wiley & Sons Ltd.

Differential roles of tissue factor and phosphatidylserine in activation of coagulation.

PubMed

Spronk, Henri M H; ten Cate, Hugo; van der Meijden, Paola E J

2014-05-01

It has been suggested that the main physiological trigger of coagulation, tissue factor, possesses limited procoagulant activity and occurs in an inactive or so-called encrypted state. For the conversion of encrypted into decrypted tissue factor with sufficient procoagulant activity, four distinct models have been proposed: 1; dimer formation, 2; lipid rafts, 3; disulfide bonds, and 4; phosphatidylserine exposure. Pro and cons can be given for each of these mechanisms of tissue factor encryption/decryption, however, it seems most likely that two or more mechanisms act together in activating the procoagulant activity. The exposure of phosphatidylserine in the outer layer of cell membranes supports coagulation through enhanced formation of the tenase (factors IXa, VIIIa and X) and prothrombinase (factors Xa, Va and prothrombin) complexes. The proposed role for phosphatidylserine in decryption of tissue factor could contribute to the correct orientation of the tissue factor - factor VII complex. Overall, the contribution of both tissue factor and phosphatidylserine to coagulation seems distinct with tissue factor being the physiological activator and phosphatidylserine the driving force of propagation of coagulation. Copyright © 2014 Elsevier Ltd. All rights reserved.

Anomalously Fast Diffusion of Targeted Carbon Nanotubes in Cellular Spheroids.

PubMed

Wang, Yichun; Bahng, Joong Hwan; Che, Quantong; Han, Jishu; Kotov, Nicholas A

2015-08-25

Understanding transport of carbon nanotubes (CNTs) and other nanocarriers within tissues is essential for biomedical imaging and drug delivery using these carriers. Compared to traditional cell cultures in animal studies, three-dimensional tissue replicas approach the complexity of the actual organs and enable high temporal and spatial resolution of the carrier permeation. We investigated diffusional transport of CNTs in highly uniform spheroids of hepatocellular carcinoma and found that apparent diffusion coefficients of CNTs in these tissue replicas are anomalously high and comparable to diffusion rates of similarly charged molecules with molecular weights 10000× lower. Moreover, diffusivity of CNTs in tissues is enhanced after functionalization with transforming growth factor β1. This unexpected trend contradicts predictions of the Stokes-Einstein equation and previously obtained empirical dependences of diffusivity on molecular mass for permeants in gas, liquid, solid or gel. It is attributed to the planar diffusion (gliding) of CNTs along cellular membranes reducing effective dimensionality of diffusional space. These findings indicate that nanotubes and potentially similar nanostructures are capable of fast and deep permeation into the tissue, which is often difficult to realize with anticancer agents.

Identification and Validation of Novel Hedgehog-Responsive Enhancers Predicted by Computational Analysis of Ci/Gli Binding Site Density

PubMed Central

Richards, Neil; Parker, David S.; Johnson, Lisa A.; Allen, Benjamin L.; Barolo, Scott; Gumucio, Deborah L.

2015-01-01

The Hedgehog (Hh) signaling pathway directs a multitude of cellular responses during embryogenesis and adult tissue homeostasis. Stimulation of the pathway results in activation of Hh target genes by the transcription factor Ci/Gli, which binds to specific motifs in genomic enhancers. In Drosophila, only a few enhancers (patched, decapentaplegic, wingless, stripe, knot, hairy, orthodenticle) have been shown by in vivo functional assays to depend on direct Ci/Gli regulation. All but one (orthodenticle) contain more than one Ci/Gli site, prompting us to directly test whether homotypic clustering of Ci/Gli binding sites is sufficient to define a Hh-regulated enhancer. We therefore developed a computational algorithm to identify Ci/Gli clusters that are enriched over random expectation, within a given region of the genome. Candidate genomic regions containing Ci/Gli clusters were functionally tested in chicken neural tube electroporation assays and in transgenic flies. Of the 22 Ci/Gli clusters tested, seven novel enhancers (and the previously known patched enhancer) were identified as Hh-responsive and Ci/Gli-dependent in one or both of these assays, including: Cuticular protein 100A (Cpr100A); invected (inv), which encodes an engrailed-related transcription factor expressed at the anterior/posterior wing disc boundary; roadkill (rdx), the fly homolog of vertebrate Spop; the segment polarity gene gooseberry (gsb); and two previously untested regions of the Hh receptor-encoding patched (ptc) gene. We conclude that homotypic Ci/Gli clustering is not sufficient information to ensure Hh-responsiveness; however, it can provide a clue for enhancer recognition within putative Hedgehog target gene loci. PMID:26710299

Bioactive Nano-Fibrous Scaffolds for Bone and Cartilage Tissue Engineering

NASA Astrophysics Data System (ADS)

Feng, Kai

Scaffolds that can mimic the structural features of natural extracellular matrix and can deliver biomolecules in a controlled fashion may provide cells with a favorable microenvironment to facilitate tissue regeneration. Biodegradable nanofibrous scaffolds with interconnected pore network have previously been developed in our laboratory to mimic collagen matrix and advantageously support both bone and cartilage regeneration. This dissertation project aims to expand both the structural complexity and the biomolecule delivery capacity of such biomimetic scaffolds for tissue engineering. We first developed a nanofibrous scaffold that can release an antibiotic (doxycycline) with a tunable release rate and a tunable dosage, which was demonstrated to be able to inhibit bacterial growth over a prolonged time period. We then developed a nanofibrous tissue-engineciing scaffold that can release basic fibroblast growth factor (bFGF) in a spatially and temporally controlled fashion. In a mouse subcutaneous implantation model, the bFGF-releasing scaffold was shown to enhance cell penetration, tissue ingrowth and angiogenesis. It was also found that both the dose and the release rate of bFGF play roles in the biologic function of the scaffold. After that, we developed a nanofibrous PLLA scaffold that can release both bone morphogenetic protein 7 (BMP-7) and platelet-derived growth factor (PDGF) with distinct dosages and release kinetics. It was demonstrated that BMP-7 and PDGF could synergistically enhance bone regeneration using a mouse ectopic bone formation model and a rat periodontal fenestration defect regeneration model. The regeneration outcome was dependent on the dosage, the ratio and the release kinetics of the two growth factors. Last, we developed an anisotropic composite scaffold with an upper layer mimicking the superficial zone of cartilage and a lower layer mimicking the middle zone of cartilage. The thin superficial layer was fabricated using an electrospinning technique to support a more parallel ECM orientation to the cartilage surface. The lower layer was fabricated using a phase-separation technique to support a more isotropic ECM distribution. Human bone marrow-derived mesenchymal stem cells (hMSCs) were seeded on this complex scaffold and cultured under chondrogenic conditions. The results showed that the composite scaffold was indeed able to support anisotropic cartilage tissue structure formation.

Activation of Peroxisome Proliferator-activated Receptor γ (PPARγ) and CD36 Protein Expression

PubMed Central

Yang, Xiaoxiao; Zhang, Wenwen; Chen, Yuanli; Li, Yan; Sun, Lei; Liu, Ying; Liu, Mengyang; Yu, Miao; Li, Xiaoju; Han, Jihong; Duan, Yajun

2016-01-01

Progesterone or its analog, one of components of hormone replacement therapy, may attenuate the cardioprotective effects of estrogen. However, the underlying mechanisms have not been fully elucidated. Expression of CD36, a receptor for oxidized LDL (oxLDL) that enhances macrophage/foam cell formation, is activated by the transcription factor peroxisome proliferator-activated receptor γ (PPARγ). CD36 also functions as a fatty acid transporter to influence fatty acid metabolism and the pathophysiological status of several diseases. In this study, we determined that progesterone induced macrophage CD36 expression, which is related to progesterone receptor (PR) activity. Progesterone enhanced cellular oxLDL uptake in a CD36-dependent manner. Mechanistically, progesterone increased PPARγ expression and PPARγ promoter activity in a PR-dependent manner and the binding of PR with the progesterone response element in the PPARγ promoter. Specific deletion of macrophage PPARγ (MφPPARγ KO) expression in mice abolished progesterone-induced macrophage CD36 expression and cellular oxLDL accumulation. We also determined that, associated with gestation and increased serum progesterone levels, CD36 and PPARγ expression in mouse adipose tissue, skeletal muscle, and peritoneal macrophages were substantially activated. Taken together, our study demonstrates that progesterone can play dual pathophysiological roles by activating PPARγ expression, in which progesterone increases macrophage CD36 expression and oxLDL accumulation, a negative effect on atherosclerosis, and enhances the PPARγ-CD36 pathway in adipose tissue and skeletal muscle, a protective effect on pregnancy. PMID:27226602

Comparative Biology of Decellularized Lung Matrix: Implications of Species Mismatch in Regenerative Medicine

PubMed Central

Balestrini, Jenna L.; Gard, Ashley L.; Gerhold, Kristin A.; Wilcox, Elise C.; Liu, Angela; Schwan, Jonas; Le, Andrew V.; Baevova, Pavlina; Dimitrievska, Sashka; Zhao, Liping; Sundaram, Sumati; Sun, Huanxing; Rittié, Laure; Dyal, Rachel; Broekelmann, Tom J.; Mecham, Robert P.; Schwartz, Martin A.; Niklason, Laura E.; White, Eric S.

2016-01-01

Lung engineering is a promising technology, relying on re-seeding of either human or xenographic decellularized matrices with patient-derived pulmonary cells. Little is known about the species-specificity of decellularization in various models of lung regeneration, or if species dependent cell-matrix interactions exist within these systems. Therefore decellularized scaffolds were produced from rat, pig, primate and human lungs, and assessed by measuring residual DNA, mechanical properties, and key matrix proteins (collagen, elastin, glycosaminoglycans). To study intrinsic matrix biologic cues, human endothelial cells were seeded onto acellular slices and analyzed for markers of cell health and inflammation. Despite similar levels of collagen after decellularization, human and primate lungs were stiffer, contained more elastin, and retained fewer glycosaminoglycans than pig or rat lung scaffolds. Human endothelial cells seeded onto human and primate lung tissue demonstrated less expression of vascular cell adhesion molecule and activation of nuclear factor-κB compared to those seeded onto rodent or porcine tissue. Adhesion of endothelial cells was markedly enhanced on human and primate tissues. Our work suggests that species-dependent biologic cues intrinsic to lung extracellular matrix could have profound effects on attempts at lung regeneration. PMID:27344365

Tissue factor-dependent coagulation activation by heme: A thromboelastometry study.

PubMed

de Souza, Gleice Regina; Hounkpe, Bidossessi Wilfried; Fiusa, Maiara Marx Luz; Colella, Marina Pereira; Annichino-Bizzacchi, Joyce M; Traina, Fabiola; Costa, Fernando Ferreira; De Paula, Erich Vinicius

2017-01-01

Heme has been characterized as potent trigger of inflammation. In hemostasis, although heme has been shown to both induce and inhibit different compartments of hemostasis, its net effect on the hemostatic balance, and the biological relevance of these effects remain to be determined. Herein we evaluated the effect of heme on hemostasis using a global assay able to generate clinically relevant data in several other complex hemostatic diseases. Citrated whole blood samples from healthy participants were stimulated by heme or vehicle and incubated for 4h at 37°C. Rotational thromboelastometry was immediately performed. The participation of tissue factor in coagulation activation was evaluated using inhibitory antibody. Heme was able of inducing ex vivo coagulation activation in whole blood, affecting predominantly parameters associated with the initial phases of clot formation. This activation effect was at least partially dependent on hematopoietic tissue factor, since the effects of heme were partially abrogated by the inhibition of human tissue factor. In conclusion, using a global hemostasis assay, our study confirmed that heme is able to activate coagulation in whole blood, in a tissue factor-dependent way. These findings could explain the disturbance in hemostatic balance observed in conditions associated with the release of heme such as sickle cell disease.

Investigation of scattering coefficients and anisotropy factors of human cancerous and normal prostate tissues using Mie theory

NASA Astrophysics Data System (ADS)

Pu, Yang; Chen, Jun; Wang, Wubao

2014-02-01

The scattering coefficient, μs, the anisotropy factor, g, the scattering phase function, p(θ), and the angular dependence of scattering intensity distributions of human cancerous and normal prostate tissues were systematically investigated as a function of wavelength, scattering angle and scattering particle size using Mie theory and experimental parameters. The Matlab-based codes using Mie theory for both spherical and cylindrical models were developed and applied for studying the light propagation and the key scattering properties of the prostate tissues. The optical and structural parameters of tissue such as the index of refraction of cytoplasm, size of nuclei, and the diameter of the nucleoli for cancerous and normal human prostate tissues obtained from the previous biological, biomedical and bio-optic studies were used for Mie theory simulation and calculation. The wavelength dependence of scattering coefficient and anisotropy factor were investigated in the wide spectral range from 300 nm to 1200 nm. The scattering particle size dependence of μs, g, and scattering angular distributions were studied for cancerous and normal prostate tissues. The results show that cancerous prostate tissue containing larger size scattering particles has more contribution to the forward scattering in comparison with the normal prostate tissue. In addition to the conventional simulation model that approximately considers the scattering particle as sphere, the cylinder model which is more suitable for fiber-like tissue frame components such as collagen and elastin was used for developing a computation code to study angular dependence of scattering in prostate tissues. To the best of our knowledge, this is the first study to deal with both spherical and cylindrical scattering particles in prostate tissues.

hCG-dependent regulation of angiogenic factors in human granulosa lutein cells.

PubMed

Phan, B; Rakenius, A; Pietrowski, D; Bettendorf, H; Keck, C; Herr, D

2006-07-01

As prerequisite for development and maintenance of many diseases angiogenesis is of particular interest in medicine. Pathologic angiogenesis takes place in chronic arthritis, collagen diseases, arteriosclerosis, retinopathy associated with diabetes, and particularly in cancers. However, angiogenesis as a physiological process regularly occurs in the ovary. After ovulation the corpus luteum is formed by rapid vascularization of initially avascular granulosa lutein cell tissue. This process is regulated by gonadotropic hormones. In order to gain further insights in the regulatory mechanisms of angiogenesis in the ovary, we investigated these mechanisms in cell culture of human granulosa lutein cells. In particular, we determined the expression and production of several angiogenic factors including tissue inhibitor of matrix metalloproteinases-1 (TIMP-1), Leptin, connective tissue growth factor (CTGF), meningioma-associated complimentary DNA (Mac25), basic fibroblast growth factor (bFGF), and Midkine. In addition, we showed that human chorionic gonadotropin (hCG) has distinct effects on their expression and production. hCG enhances the expression and production of TIMP-1, whereas it downregulates the expression of CTGF and Mac25. Furthermore it decreases the expression of Leptin. Our results provide evidence that hCG determines growth and development of the corpus luteum by mediating angiogenic pathways in human granulosa lutein cells. Hence we describe a further approach to understand the regulation of angiogenesis in the ovary.

Exposure of T lymphocytes to leflunomide but not to dexamethasone favors the production by monocytic cells of interleukin-1 receptor antagonist and the tissue-inhibitor of metalloproteinases-1 over that of interleukin-1beta and metalloproteinases.

PubMed

Déage, V; Burger, D; Dayer, J M

1998-12-01

On direct cell-cell contact, stimulated T lymphocytes potently trigger the production of pro-inflammatory factors such as interleukin-1beta (IL-1beta) and matrix metalloproteinases (MMP-1 and MMP-9), as well as anti-inflammatory factors such as IL-1 receptor antagonist (IL-1Ra) and the tissue inhibitor of metalloproteinases (TIMP-1) in peripheral blood monocytes and the monocytic cell line THP-1. Such mechanisms might play an important part in many inflammatory diseases where tissue destruction occurs. To assess whether anti-inflammatory agents such as dexamethasone (DEX) and leflunomide (LF) would affect contact-activation of monocytic cells, T lymphocytes were stimulated by PMA and PHA in the presence or absence of increasing concentrations of drug. LF and DEX (10- 4 M) inhibited the ability of stimulated T lymphocytes to activate monocytic cells by 66-97% and 43-70%, respectively, depending on the readout product. Upon contact with T lymphocytes stimulated in the presence of 10- 5 M LF, the molar ratio of IL-1Ra/IL-1beta and TIMP-1/MMP-1 produced by THP-1 cells was enhanced 3.6- and 1.9-fold, respectively, whereas it was enhanced only 1.3- and 1.4-fold upon contact with T lymphocytes stimulated in the presence of 10- 4 M DEX. Therefore, LF tends to favor the inhibition of pro-inflammatory and matrix-destructive factors over that of anti-inflammatory factors and metalloproteinase inhibitors, thus interfering with both inflammation and tissue destruction. These experiments indicate that LF and DEX have the potential to affect the capacity of stimulated T lymphocytes to activate, on direct cell-cell contact, monocytic cells. Furthermore, flow cytometric analysis revealed that surface molecules of T lymphocytes that were partially involved in contact-signaling of monocytes (i.e., CD69 and CD11) were not modulated by either LF or DEX, suggesting that factors which remain to be identified were mainly involved in the activation of monocytes on direct cell-cell contact.

Combined chemical and structural signals of biomaterials synergistically activate cell-cell communications for improving tissue regeneration.

PubMed

Xu, Yachen; Peng, Jinliang; Dong, Xin; Xu, Yuhong; Li, Haiyan; Chang, Jiang

2017-06-01

Biomaterials are only used as carriers of cells in the conventional tissue engineering. Considering the multi-cell environment and active cell-biomaterial interactions in tissue regeneration process, in this study, structural signals of aligned electrospun nanofibers and chemical signals of bioglass (BG) ionic products in cell culture medium are simultaneously applied to activate fibroblast-endothelial co-cultured cells in order to obtain an improved skin tissue engineering construct. Results demonstrate that the combined biomaterial signals synergistically activate fibroblast-endothelial co-culture skin tissue engineering constructs through promotion of paracrine effects and stimulation of gap junctional communication between cells, which results in enhanced vascularization and extracellular matrix protein synthesis in the constructs. Structural signals of aligned electrospun nanofibers play an important role in stimulating both of paracrine and gap junctional communication while chemical signals of BG ionic products mainly enhance paracrine effects. In vivo experiments reveal that the activated skin tissue engineering constructs significantly enhance wound healing as compared to control. This study indicates the advantages of synergistic effects between different bioactive signals of biomaterials can be taken to activate communication between different types of cells for obtaining tissue engineering constructs with improved functions. Tissue engineering can regenerate or replace tissue or organs through combining cells, biomaterials and growth factors. Normally, for repairing a specific tissue, only one type of cells, one kind of biomaterials, and specific growth factors are used to support cell growth. In this study, we proposed a novel tissue engineering approach by simply using co-cultured cells and combined biomaterial signals. Using a skin tissue engineering model, we successfully proved that the combined biomaterial signals such as surface nanostructures and bioactive ions could synergistically stimulate the cell-cell communication in co-culture system through paracrine effects and gap junction activation, and regulated expression of growth factors and extracellular matrix proteins, resulting in an activated tissue engineering constructs that significantly enhanced skin regeneration. Copyright © 2017 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

In vivo bioresponses to silk proteins.

PubMed

Thurber, Amy E; Omenetto, Fiorenzo G; Kaplan, David L

2015-12-01

Silks are appealing materials for numerous biomedical applications involving drug delivery, tissue engineering, or implantable devices, because of their tunable mechanical properties and wide range of physical structures. In addition to the functionalities needed for specific clinical applications, a key factor necessary for clinical success for any implanted material is appropriate interactions with the body in vivo. This review summarizes our current understanding of the in vivo biological responses to silks, including degradation, the immune and inflammatory response, and tissue remodeling with particular attention to vascularization. While we focus in this review on silkworm silk fibroin protein due to the large quantity of in vivo data thanks to its widespread use in medical materials and consumer products, spider silk information is also included if available. Silk proteins are degraded in the body on a time course that is dependent on the method of silk fabrication and can range from hours to years. Silk protein typically induces a mild inflammatory response that decreases within a few weeks of implantation. The response involves recruitment and activation of macrophages and may include activation of a mild foreign body response with the formation of multinuclear giant cells, depending on the material format and location of implantation. The number of immune cells present decreases with time and granulation tissue, if formed, is replaced by endogenous, not fibrous, tissue. Importantly, silk materials have not been demonstrated to induce mineralization, except when used in calcified tissues. Due to its ability to be degraded, silk can be remodeled in the body allowing for vascularization and tissue ingrowth with eventual complete replacement by native tissue. The degree of remodeling, tissue ingrowth, or other specific cell behaviors can be modulated with addition of growth or other signaling factors. Silk can also be combined with numerous other materials including proteins, synthetic polymers, and ceramics to enhance its characteristics for a particular function. Overall, the diverse array of silk materials shows excellent bioresponses in vivo with low immunogenicity and the ability to be remodeled and replaced by native tissue making it suitable for numerous clinical applications. Copyright © 2015 Elsevier Ltd. All rights reserved.

Increased avidity for Dpp/BMP2 maintains the proliferation of progenitors-like cells in the Drosophila eye.

PubMed

Neto, Marta; Aguilar-Hidalgo, Daniel; Casares, Fernando

2016-10-01

During organ development, the progenitor state is transient, and depends on specific combinations of transcription factors and extracellular signals. Not surprisingly, abnormal maintenance of progenitor transcription factors may lead to tissue overgrowth, and the concurrence of signals from the local environment is often critical to trigger this overgrowth. Therefore, identifying specific combinations of transcription factors/signals promoting -or opposing- proliferation in progenitors is essential to understand normal development and disease. We have investigated this issue using the Drosophila eye as model. Transcription factors hth and tsh are transiently expressed in eye progenitors causing the expansion of the progenitor pool. However, if their co-expression is maintained experimentally, cell proliferation continues and differentiation is halted. Here we show that Hth+Tsh-induced tissue overgrowth requires the BMP2 Dpp and the abnormal hyperactivation of its pathway. Rather than using autocrine Dpp expression, Hth+Tsh cells increase their avidity for Dpp, produced locally, by upregulating extracellular matrix components. During normal development, Dpp represses hth and tsh ensuring that the progenitor state is transient. However, cells in which Hth+Tsh expression is forcibly maintained use Dpp to enhance their proliferation. Copyright © 2016 Elsevier Inc. All rights reserved.

Enhanced angiogenic effect of adipose-derived stromal cell spheroid with low-level light therapy in hindlimb ischemia mice

NASA Astrophysics Data System (ADS)

Park, In-Su; Ahn, Jin-Chul; Chung, Phil-Sang

2014-02-01

Adipose-derived stromal cells (ASCs) are attractive cell source for tissue engineering. However, one obstacle to this approach is that the transplanted ASC population can decline rapidly in the recipient tissue. The aim of this study was to investigate the effects of low-level laser therapy (LLLT) on transplanted human ASCs (hASCs) spheroid in a hindlimb ischemia animal model. LLLT, hASCs spheroid and hASCs spheroid transplantation with LLLT (spheroid + LLLT) were applied to the ischemic hindlimbs in athymic mice. The survival, differentiation and secretion of vascular endothelial growth (VEGF) of spheroid ASCs were evaluated by immunohistochemistry. The spheroid + LLLT group enhanced the tissue regeneration, including angiogenesis, compared with other groups. The spheroid contributed tissue regeneration via differentiation and secretion of growth factors. In the spheroid + LLLT group, the survival of spheroid hASCs was increased by the decreased apoptosis of spheroid hASCs in the ischemic hindlimb. The secretion of growth factors was stimulated in the spheroid + LLLT group compared with the ASCs group and spheroid group. These data suggest that LLLT is an effective biostimulator of spheroid hASCs in tissue regeneration that enhances the survival of ASCs and stimulates the secretion of growth factors in the ischemic hindlimb.

PPARβ/δ ameliorates fructose-induced insulin resistance in adipocytes by preventing Nrf2 activation.

PubMed

Barroso, Emma; Rodríguez-Rodríguez, Rosalía; Chacón, Matilde R; Maymó-Masip, Elsa; Ferrer, Laura; Salvadó, Laia; Salmerón, Emilio; Wabistch, Martin; Palomer, Xavier; Vendrell, Joan; Wahli, Walter; Vázquez-Carrera, Manuel

2015-05-01

We studied whether PPARβ/δ deficiency modifies the effects of high fructose intake (30% fructose in drinking water) on glucose tolerance and adipose tissue dysfunction, focusing on the CD36-dependent pathway that enhances adipose tissue inflammation and impairs insulin signaling. Fructose intake for 8 weeks significantly increased body and liver weight, and hepatic triglyceride accumulation in PPARβ/δ-deficient mice but not in wild-type mice. Feeding PPARβ/δ-deficient mice with fructose exacerbated glucose intolerance and led to macrophage infiltration, inflammation, enhanced mRNA and protein levels of CD36, and activation of the JNK pathway in white adipose tissue compared to those of water-fed PPARβ/δ-deficient mice. Cultured adipocytes exposed to fructose also exhibited increased CD36 protein levels and this increase was prevented by the PPARβ/δ activator GW501516. Interestingly, the levels of the nuclear factor E2-related factor 2 (Nrf2), a transcription factor reported to up-regulate Cd36 expression and to impair insulin signaling, were increased in fructose-exposed adipocytes whereas co-incubation with GW501516 abolished this increase. In agreement with Nrf2 playing a role in the fructose-induced CD36 protein level increases, the Nrf2 inhibitor trigonelline prevented the increase and the reduction in insulin-stimulated AKT phosphorylation caused by fructose in adipocytes. Protein levels of the well-known Nrf2 target gene quinone oxidoreductase 1 (Nqo1) were increased in water-fed PPARβ/δ-null mice, suggesting that PPARβ/δ deficiency increases Nrf2 activity; and this increase was exacerbated in fructose-fed PPARβ/δ-deficient mice. These findings indicate that the combination of high fructose intake and PPARβ/δ deficiency increases CD36 protein levels via Nrf2, a process that promotes chronic inflammation and insulin resistance in adipose tissue. Copyright © 2015 Elsevier B.V. All rights reserved.

FXIa and platelet polyphosphate as therapeutic targets during human blood clotting on collagen/tissue factor surfaces under flow.

PubMed

Zhu, Shu; Travers, Richard J; Morrissey, James H; Diamond, Scott L

2015-09-17

Factor XIIa (FXIIa) and factor XIa (FXIa) contribute to thrombosis in animal models, whereas platelet-derived polyphosphate (polyP) may potentiate contact or thrombin-feedback pathways. The significance of these mediators in human blood under thrombotic flow conditions on tissue factor (TF) -bearing surfaces remains inadequately resolved. Human blood (corn trypsin inhibitor treated [4 μg/mL]) was tested by microfluidic assay for clotting on collagen/TF at TF surface concentration ([TF]wall) from ∼0.1 to 2 molecules per μm(2). Anti-FXI antibodies (14E11 and O1A6) or polyP-binding protein (PPXbd) were used to block FXIIa-dependent FXI activation, FXIa-dependent factor IX (FIX) activation, or platelet-derived polyP, respectively. Fibrin formation was sensitive to 14E11 at 0 to 0.1 molecules per µm(2) and sensitive to O1A6 at 0 to 0.2 molecules per µm(2). However, neither antibody reduced fibrin generation at ∼2 molecules per µm(2) when the extrinsic pathway became dominant. Interestingly, PPXbd reduced fibrin generation at low [TF]wall (0.1 molecules per µm(2)) but not at zero or high [TF]wall, suggesting a role for polyP distinct from FXIIa activation and requiring low extrinsic pathway participation. Regardless of [TF]wall, PPXbd enhanced fibrin sensitivity to tissue plasminogen activator and promoted clot retraction during fibrinolysis concomitant with an observed PPXbd-mediated reduction of fibrin fiber diameter. This is the first detection of endogenous polyP function in human blood under thrombotic flow conditions. When triggered by low [TF]wall, thrombosis may be druggable by contact pathway inhibition, although thrombolytic susceptibility may benefit from polyP antagonism regardless of [TF]wall. © 2015 by The American Society of Hematology.

Sequential Delivery of BMP-2 and IGF-1 Using a Chitosan Gel with gelatin Microspheres Enhances Early osteoblastic Differentiation

DTIC Science & Technology

2012-01-18

Vunjak- Novakovic G, Freed LE. Growth factors for sequential cellular de- and re-differentiation in tissue engineering. Biochem Biophys Res Commun...2002;294(1):149–54. [35] Martin I, Suetterlin R, Baschong W, Heberer M, Vunjak- Novakovic G, Freed LE. Enhanced cartilage tissue engineering by sequential

Growth Hormone and Reproduction: A Review of Endocrine and Autocrine/Paracrine Interactions

PubMed Central

Hull, Kerry L.; Harvey, Steve

2014-01-01

The somatotropic axis, consisting of growth hormone (GH), hepatic insulin-like growth factor I (IGF-I), and assorted releasing factors, regulates growth and body composition. Axiomatically, since optimal body composition enhances reproductive function, general somatic actions of GH modulate reproductive function. A growing body of evidence supports the hypothesis that GH also modulates reproduction directly, exerting both gonadotropin-dependent and gonadotropin-independent actions in both males and females. Moreover, recent studies indicate GH produced within reproductive tissues differs from pituitary GH in terms of secretion and action. Accordingly, GH is increasingly used as a fertility adjunct in males and females, both humans and nonhumans. This review reconsiders reproductive actions of GH in vertebrates in respect to these new conceptual developments. PMID:25580121

Negative regulatory role of PI3-kinase in TNF-induced tumor necrosis.

PubMed

Matschurat, Susanne; Blum, Sabine; Mitnacht-Kraus, Rita; Dijkman, Henry B P M; Kanal, Levent; De Waal, Robert M W; Clauss, Matthias

2003-10-20

Tissue factor is the prime initiator of blood coagulation. Expression of tissue factor in tumor endothelial cells leads to thrombus formation, occlusion of vessels and development of hemorrhagic infarctions in the tumor tissue, often followed by regression of the tumor. Tumor cells produce endogenous vascular endothelial growth factor (VEGF), which sensitizes endothelial cells for systemically administered tumor necrosis factor alpha (TNF alpha) and synergistically enhances the TNF-induced expression of tissue factor. We have analyzed the pathways involved in the induction of tissue factor in human umbilical cord vein endothelial cells (HUVECs) after combined stimulation with TNF and VEGF. By using specific low molecular weight inhibitors, we demonstrated that protein kinase C (PKC), p44/42 and p38 mitogen-activated protein (MAP) kinases, and stress-activated protein kinase (JNK) are essentially involved in the induction of tissue factor. In contrast, the application of wortmannin, an inhibitor of phosphatidylinositol 3 (PI3)-kinase, led to strongly enhanced expression of tissue factor in TNF- and VEGF-treated cells, implicating a negative regulatory role for PI3-kinase. In vivo, the application of wortmannin promoted the formation of TNF-induced hemorrhages and intratumoral necroses in murine meth A tumors. The co-injection of wortmannin lowered the effective dose of applied TNF. Therefore, it is conceivable that the treatment of TNF-sensitive tumors with a combination of TNF and wortmannin will ensure the selective damage of the tumor endothelium and minimize the risk of systemic toxicity of TNF. TNF-treatment in combination with specific inhibition of PI3-kinase is a novel concept in anti-cancer therapy. Copyright 2003 Wiley-Liss, Inc.

Significance of biological resource collection and tumor tissue bank creation.

PubMed

Yu, Ying-Yan; Zhu, Zheng-Gang

2010-01-15

Progress in the molecular oncology of gastrointestinal carcinomas depends on high quality cancer tissues for research. Recent acceleration on new technological platforms as well as the "omics" revolution increases the demands on tissues and peripheral blood for research at the DNA, mRNA and protein levels. Tissue bank creation emerges as a priority. Tumor tissue banks are facilities that are organized to collect, store and distribute samples of tumor and normal tissue for further use in basic and translational cancer research. The samples are generally obtained immediately after excision, prior to fixation, to ensure optimal preservation of proteins and nucleic acids. It is possible for surgeons or pathologists to collect fresh tissue prospectively during their routine dissection procedures. Most tissue banks are "project-driven" tumor banks, which are specialized collections of tumor samples on which their research is based. Systematic collection of all available tumor tissue is much rarer. High quality tissue banks need the collaboration of clinicians and basic scientists, but also the informed consent of patients and ethical approval. Through the standard operation procedure, snap frozen fresh tissue collection, storage and quality control for cryopreserved tissues are the pivotal factors on tissue bank construction and maintaining. The purpose of the tissue bank creation is enhancing the quality and speed on both the basic and translational research on gastrointestinal cancer. The quality assurance and quality control are handled based on reviewing HE staining slides or touch imprint cytology by pathologists.

The tumor secretory factor ZAG promotes white adipose tissue browning and energy wasting.

PubMed

Elattar, Sawsan; Dimri, Manali; Satyanarayana, Ande

2018-03-23

Cachexia is a complex tissue-wasting syndrome characterized by inflammation, hypermetabolism, increased energy expenditure, and anorexia. Browning of white adipose tissue (WAT) is one of the significant factors that contribute to energy wasting in cachexia. By utilizing a cell implantation model, we demonstrate here that the lipid mobilizing factor zinc-α 2 -glycoprotein (ZAG) induces WAT browning in mice. Increased circulating levels of ZAG not only induced lipolysis in adipose tissues but also caused robust browning in WAT. Stimulating WAT progenitors with ZAG recombinant protein or expression of ZAG in mouse embryonic fibroblasts (MEFs) strongly enhanced brown-like differentiation. At the molecular level, ZAG stimulated peroxisome proliferator-activated receptor γ (PPARγ) and early B cell factor 2 expression and promoted their recruitment to the PR/SET domain 16 (Prdm16) promoter, leading to enhanced expression of Prdm16, which determines brown cell fate. In brown adipose tissue, ZAG stimulated the expression of PPARγ and PPARγ coactivator 1α and promoted recruitment of PPARγ to the uncoupling protein 1 (Ucp1) promoter, leading to increased expression of Ucp1. Overall, our results reveal a novel function of ZAG in WAT browning and highlight the targeting of ZAG as a potential therapeutic application in humans with cachexia.-Elattar, S., Dimri, M., Satyanarayana, A. The tumor secretory factor ZAG promotes white adipose tissue browning and energy wasting.

Reciprocal inhibition of p53 and nuclear factor-kappaB transcriptional activities determines cell survival or death in neurons.

PubMed

Culmsee, Carsten; Siewe, Jan; Junker, Vera; Retiounskaia, Marina; Schwarz, Stephanie; Camandola, Simonetta; El-Metainy, Shahira; Behnke, Hagen; Mattson, Mark P; Krieglstein, Josef

2003-09-17

The tumor suppressor and transcription factor p53 is a key modulator of cellular stress responses, and activation of p53 precedes apoptosis in many cell types. Controversial reports exist on the role of the transcription factor nuclear factor-kappaB (NF-kappaB) in p53-mediated apoptosis, depending on the cell type and experimental conditions. Therefore, we sought to elucidate the role of NF-kappaB in p53-mediated neuron death. In cultured neurons DNA damaging compounds induced activation of p53, whereas NF-kappaB activity declined significantly. The p53 inhibitor pifithrin-alpha (PFT) preserved NF-kappaB activity and protected neurons against apoptosis. Immunoprecipitation experiments revealed enhanced p53 binding to the transcriptional cofactor p300 after induction of DNA damage, whereas binding of p300 to NF-kappaB was reduced. In contrast, PFT blocked the interaction of p53 with the cofactor, whereas NF-kappaB binding to p300 was enhanced. Most interestingly, similar results were observed after oxygen glucose deprivation in cultured neurons and in ischemic brain tissue. Ischemia-induced repression of NF-kappaB activity was prevented and brain damage was reduced by the p53 inhibitor PFT in a dose-dependent manner. It is concluded that a balanced competitive interaction of p53 and NF-kappaB with the transcriptional cofactor p300 exists in neurons. Exposure of neurons to lethal stress activates p53 and disrupts NF-kappaB binding to p300, thereby blocking NF-kappaB-mediated survival signaling. Inhibitors of p53 provide pronounced neuroprotective effects because they block p53-mediated induction of cell death and concomitantly enhance NF-kappaB-induced survival signaling.

Active medulloblastoma enhancers reveal subgroup-specific cellular origins

PubMed Central

Lin, Charles Y.; Erkek, Serap; Tong, Yiai; Yin, Linlin; Federation, Alexander J.; Zapatka, Marc; Haldipur, Parthiv; Kawauchi, Daisuke; Risch, Thomas; Warnatz, Hans-Jörg; Worst, Barbara C.; Ju, Bensheng; Orr, Brent A.; Zeid, Rhamy; Polaski, Donald R.; Segura-Wang, Maia; Waszak, Sebastian M.; Jones, David T.W.; Kool, Marcel; Hovestadt, Volker; Buchhalter, Ivo; Sieber, Laura; Johann, Pascal; Chavez, Lukas; Gröschel, Stefan; Ryzhova, Marina; Korshunov, Andrey; Chen, Wenbiao; Chizhikov, Victor V.; Millen, Kathleen J.; Amstislavskiy, Vyacheslav; Lehrach, Hans; Yaspo, Marie-Laure; Eils, Roland; Lichter, Peter; Korbel, Jan O.; Pfister, Stefan M.; Bradner, James E.; Northcott, Paul A.

2016-01-01

Summary Medulloblastoma is a highly malignant paediatric brain tumour, often inflicting devastating consequences on the developing child. Genomic studies have revealed four distinct molecular subgroups with divergent biology and clinical behaviour. An understanding of the regulatory circuitry governing the transcriptional landscapes of medulloblastoma subgroups, and how this relates to their respective developmental origins, is lacking. Using H3K27ac and BRD4 ChIP-Seq, coupled with tissue-matched DNA methylation and transcriptome data, we describe the active cis-regulatory landscape across 28 primary medulloblastoma specimens. Analysis of differentially regulated enhancers and super-enhancers reinforced inter-subgroup heterogeneity and revealed novel, clinically relevant insights into medulloblastoma biology. Computational reconstruction of core regulatory circuitry identified a master set of transcription factors, validated by ChIP-Seq, that are responsible for subgroup divergence and implicate candidate cells-of-origin for Group 4. Our integrated analysis of enhancer elements in a large series of primary tumour samples reveals insights into cis-regulatory architecture, unrecognized dependencies, and cellular origins. PMID:26814967

Activated Monocytes Enhance Platelet-Driven Contraction of Blood Clots via Tissue Factor Expression.

PubMed

Peshkova, Alina D; Le Minh, Giang; Tutwiler, Valerie; Andrianova, Izabella A; Weisel, John W; Litvinov, Rustem I

2017-07-11

Platelet-driven reduction in blood clot volume (clot contraction or retraction) has been implicated to play a role in hemostasis and thrombosis. Although these processes are often linked with inflammation, the role of inflammatory cells in contraction of blood clots and thrombi has not been investigated. The aim of this work was to study the influence of activated monocytes on clot contraction. The effects of monocytes were evaluated using a quantitative optical tracking methodology to follow volume changes in a blood clot formed in vitro. When a physiologically relevant number of isolated human monocytes pre-activated with phorbol-12-myristate-13-acetate (PMA) were added back into whole blood, the extent and rate of clot contraction were increased compared to addition of non-activated cells. Inhibition of tissue factor expression or its inactivation on the surface of PMA-treated monocytes reduced the extent and rate of clot contraction back to control levels with non-activated monocytes. On the contrary, addition of tissue factor enhanced clot contraction, mimicking the effects of tissue factor expressed on the activated monocytes. These data suggest that the inflammatory cells through their expression of tissue factor can directly affect hemostasis and thrombosis by modulating the size and density of intra- and extravascular clots and thrombi.

Enhanced angiogenic effect of adipose-derived stromal cell spheroid with low-level light therapy in hind limb ischemia mice.

PubMed

Park, In-Su; Chung, Phil-Sang; Ahn, Jin Chul

2014-11-01

The aim of this study was to investigate the effects of low-level laser therapy (LLLT) on transplanted human adipose-derived mesenchymal stem cells (hASCs) spheroid in a hind limb ischemia animal model. LLLT, hASCs spheroid and hASCs spheroid transplantation with LLLT (spheroid + LLLT) were applied to the ischemic hind limbs in athymic mice. The survival, differentiation and secretion of vascular endothelial growth factor (VEGF), basic fibroblast growth factor (FGF), and hepatocyte growth factor (HGF) of the spheroid ASCs were evaluated by immunohistochemistry and western blots. Spheroid + LLLT group had enhanced the tissue regeneration, including angiogenesis, compared with the ASC group. The spheroid ASCs contributed to tissue regeneration via differentiation and secretion of growth factors. In the spheroid + LLLT group, the survival of spheroid hASCs increased with a concomitant decrease in apoptosis of spheroid hASCs in the ischemic hind limb. The secretion of growth factors was stimulated in the spheroid + LLLT group compared with the ASCs and spheroid group. These data suggested that LLLT is an effective biostimulator of spheroid hASCs in tissue regeneration that enhanced the survival of ASCs and stimulated the secretion of growth factors in the ischemic hind limb. Copyright © 2014 Elsevier Ltd. All rights reserved.

Bone morphogenetic protein-4 strongly potentiates growth factor-induced proliferation of mammary epithelial cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Montesano, Roberto; Sarkoezi, Rita; Schramek, Herbert

2008-09-12

Bone morphogenetic proteins (BMPs) are multifunctional cytokines that elicit pleiotropic effects on biological processes such as cell proliferation, cell differentiation and tissue morphogenesis. With respect to cell proliferation, BMPs can exert either mitogenic or anti-mitogenic activities, depending on the target cells and their context. Here, we report that in low-density cultures of immortalized mammary epithelial cells, BMP-4 did not stimulate cell proliferation by itself. However, when added in combination with suboptimal concentrations of fibroblast growth factor (FGF)-2, FGF-7, FGF-10, epidermal growth factor (EGF) or hepatocyte growth factor (HGF), BMP-4 potently enhanced growth factor-induced cell proliferation. These results reveal a hithertomore » unsuspected interplay between BMP-4 and growth factors in the regulation of mammary epithelial cell proliferation. We suggest that the ability of BMP-4 to potentiate the mitogenic activity of multiple growth factors may contribute to mammary gland ductal morphogenesis as well as to breast cancer progression.« less

The role of the endocrine system in feeding-induced tissue-specific circadian entrainment.

PubMed

Sato, Miho; Murakami, Mariko; Node, Koichi; Matsumura, Ritsuko; Akashi, Makoto

2014-07-24

The circadian clock is entrained to environmental cycles by external cue-mediated phase adjustment. Although the light input pathway has been well defined, the mechanism of feeding-induced phase resetting remains unclear. The tissue-specific sensitivity of peripheral entrainment to feeding suggests the involvement of multiple pathways, including humoral and neuronal signals. Previous in vitro studies with cultured cells indicate that endocrine factors may function as entrainment cues for peripheral clocks. However, blood-borne factors that are well characterized in actual feeding-induced resetting have yet to be identified. Here, we report that insulin may be involved in feeding-induced tissue-type-dependent entrainment in vivo. In ex vivo culture experiments, insulin-induced phase shift in peripheral clocks was dependent on tissue type, which was consistent with tissue-specific insulin sensitivity, and peripheral entrainment in insulin-sensitive tissues involved PI3K- and MAPK-mediated signaling pathways. These results suggest that insulin may be an immediate early factor in feeding-mediated tissue-specific entrainment. Copyright © 2014 The Authors. Published by Elsevier Inc. All rights reserved.

Impact of static magnetic fields on human myoblast cell cultures.

PubMed

Stern-Straeter, Jens; Bonaterra, Gabriel Alejandro; Kassner, Stefan S; Faber, Anne; Sauter, Alexander; Schulz, Johannes D; Hörmann, Karl; Kinscherf, Ralf; Goessler, Ulrich Reinhart

2011-12-01

Treatment of skeletal muscle loss due to trauma or tumor ablation therapy still lacks a suitable clinical approach. Creation of functional muscle tissue in vitro using the differentiation potential of human satellite cells (myoblasts) is a promising new research field called tissue engineering. Strong differentiation stimuli, which can induce formation of myofibers after cell expansion, have to be identified and evaluated in order to create sufficient amounts of neo-tissue. The objective of this study was to determine the influence of static magnetic fields (SMF) on human satellite cell cultures as one of the preferred stem cell sources in skeletal muscle tissue engineering. Experiments were performed using human satellite cells with and without SMF stimulation after incubation with a culture medium containing low [differentiation medium (DM)] or high [growth medium (GM)] concentrations of growth factors. Proliferation analysis using the alamarBlue assay revealed no significant influence of SMF on cell division. Real-time RT-PCR of the following marker genes was investigated: myogenic factor 5 (MYF5), myogenic differentiation antigen 1 (MYOD1), myogenin (MYOG), skeletal muscle α1 actin (ACTA1), and embryonic (MYH3), perinatal (MYH8) and adult (MYH1) skeletal muscle myosin heavy chain. We detected an influence on marker gene expression by SMF in terms of a down-regulation of the marker genes in cell cultures treated with SMF and DM, but not in cell cultures treated with SMF and GM. Immunocytochemical investigations using antibodies directed against the differentiation markers confirmed the gene expression results and showed an enhancement of maturation after stimulation with GM and SMF. Additional calculation of the fusion index also revealed an increase in myotube formation in cell cultures treated with SMF and GM. Our findings show that the effect of SMF on the process of differentiation depends on the growth factor concentration in the culture medium in human satellite cultures. SMF alone enhances the maturation of human satellite cells treated with GM, but not satellite cells that were additionally stimulated with serum cessation. Therefore, further investigations are necessary before consideration of SMF for skeletal muscle tissue engineering approaches.

Injectable biomimetic liquid crystalline scaffolds enhance muscle stem cell transplantation

PubMed Central

Sleep, Eduard; McClendon, Mark T.; Preslar, Adam T.; Chen, Charlotte H.; Sangji, M. Hussain; Pérez, Charles M. Rubert; Haynes, Russell D.; Meade, Thomas J.; Blau, Helen M.; Stupp, Samuel I.

2017-01-01

Muscle stem cells are a potent cell population dedicated to efficacious skeletal muscle regeneration, but their therapeutic utility is currently limited by mode of delivery. We developed a cell delivery strategy based on a supramolecular liquid crystal formed by peptide amphiphiles (PAs) that encapsulates cells and growth factors within a muscle-like unidirectionally ordered environment of nanofibers. The stiffness of the PA scaffolds, dependent on amino acid sequence, was found to determine the macroscopic degree of cell alignment templated by the nanofibers in vitro. Furthermore, these PA scaffolds support myogenic progenitor cell survival and proliferation and they can be optimized to induce cell differentiation and maturation. We engineered an in vivo delivery system to assemble scaffolds by injection of a PA solution that enabled coalignment of scaffold nanofibers with endogenous myofibers. These scaffolds locally retained growth factors, displayed degradation rates matching the time course of muscle tissue regeneration, and markedly enhanced the engraftment of muscle stem cells in injured and noninjured muscles in mice. PMID:28874575

Enhancer Activation Requires Trans-Recruitment of a Mega Transcription Factor Complex

PubMed Central

Liu, Zhijie; Merkurjev, Daria; Yang, Feng; Li, Wenbo; Oh, Soohwan; Friedman, Meyer J.; Song, Xiaoyuan; Zhang, Feng; Ma, Qi; Ohgi, Kenneth; Krones, Anna; Rosenfeld, Michael G.

2014-01-01

Summary Enhancers provide critical information directing cell-type specific transcriptional programs, regulated by binding of signal-dependent transcription factors and their associated cofactors. Here we report that the most strongly activated estrogen (E2)-responsive enhancers are characterized by trans-recruitment and in situ assembly of a large 1-2 MDa complex of diverse DNA-binding transcription factors by ERα at ERE-containing enhancers. We refer to enhancers recruiting these factors as mega transcription factor-bound in trans (MegaTrans) enhancers. The MegaTrans complex is a signature of the most potent functional enhancers and is required for activation of enhancer RNA transcription and recruitment of coactivators, including p300 and Med1. The MegaTrans complex functions, in part, by recruiting specific enzymatic machinery, exemplified by DNA-dependent protein kinase. Thus, MegaTrans-containing enhancers represent a cohort of functional enhancers that mediate a broad and important transcriptional program and provide a molecular explanation for transcription factor clustering and hotspots noted in the genome. PMID:25303530

Estrogen signaling in colorectal carcinoma microenvironment: expression of ERbeta1, AIB-1, and TIF-2 is upregulated in cancer-associated myofibroblasts and correlates with disease progression.

PubMed

Tzelepi, Vassiliki; Grivas, Petros; Kefalopoulou, Zinovia; Kalofonos, Haralabos; Varakis, John N; Melachrinou, Maria; Sotiropoulou-Bonikou, Georgia

2009-04-01

Epidemiological and molecular data suggest the involvement of estrogen signaling in colorectal tissue, mediated mainly through estrogen receptor beta (ERbeta). Estrogens may mediate their effects in epithelial cells indirectly by acting on stromal cells. Expression of ERalpha, ERbeta1, and the ER coregulators, amplified in breast cancer-1 (AIB-1) and transcriptional intermediary factor 2 (TIF-2), was evaluated in myofibroblasts of 107 colorectal carcinomas, 77 paired samples of normal mucosa, and 29 adenomas by immunohistochemistry. Double immunostaining with a-SMA was used to identify the myofibroblasts of normal tissue, adenomas, and cancer microenvironment. ERalpha was not expressed in stromal cells. Nuclear expression of ERbeta1, AIB-1, and TIF-2 in myofibroblasts gradually increased from normal mucosa, through adenomas, to carcinomas. Cytoplasmic ERbeta1 and TIF-2 expression was enhanced in carcinomas compared to normal mucosa and adenomas. Enhanced nuclear and cytoplasmic ERbeta1 expression and elevated nuclear AIB-1 expression were more frequently noted in myofibroblasts of carcinomas of advanced stage. ERbeta1 expression in cancer-associated myofibroblasts correlated to AIB-1 and TIF-2 expression. None of the markers correlated with patients' prognosis. Our findings imply that ERbeta1-dependent (genomic and non-genomic) and ER-coregulator-dependent (AIB-1, TIF-2) signal transductions in myofibroblasts may be involved in the initiation and progression of colorectal carcinomas.

Febuxostat protects rats against lipopolysaccharide-induced lung inflammation in a dose-dependent manner.

PubMed

Fahmi, Alaa N A; Shehatou, George S G; Shebl, Abdelhadi M; Salem, Hatem A

2016-03-01

The aim of the present work was to investigate possible protective effects of febuxostat, a highly potent xanthine oxidase inhibitor, against acute lung injury (ALI) induced by lipopolysaccharide (LPS) in rats. Male Sprague Dawley rats were randomly divided into six groups, as follows: (i) vehicle control group; (ii) and (iii) febuxostat 10 and febuxostat 15 groups, drug-treated controls; (iv) LPS group, receiving an intraperitoneal injection of LPS (7.5 mg/kg); (v) and (vi) febuxostat 10-LPS and febuxostat 15-LPS groups, receiving oral treatment of febuxostat (10 and 15 mg/kg/day, respectively) for 7 days before LPS. After 18 h administration of LPS, blood was collected for C-reactive protein (CRP) measurement. Bronchoalveolar lavage fluid (BALF) was examined for leukocyte infiltration, lactate dehydrogenase (LDH) activity, protein content, and total nitrate/nitrite. Lung weight gain was determined, and lung tissue homogenate was prepared and evaluated for oxidative stress. Tumor necrosis factor-α (TNF-α) was assessed in BALF and lung homogenate. Moreover, histological changes of lung tissues were evaluated. LPS elicited lung injury characterized by increased lung water content (by 1.2 fold), leukocyte infiltration (by 13 fold), inflammation and oxidative stress (indicated by increased malondialdehyde (MDA), by 3.4 fold), and reduced superoxide dismutase (SOD) activity (by 34 %). Febuxostat dose-dependently decreased LPS-induced lung edema and elevations in BALF protein content, infiltration of leukocytes, and LDH activity. Moreover, the elevated levels of TNF-α in BALF and lung tissue of LPS-treated rats were attenuated by febuxostat pretreatment. Febuxostat also displayed a potent antioxidant activity by decreasing lung tissue levels of MDA and enhancing SOD activity. Histological analysis of lung tissue further demonstrated that febuxostat dose-dependently reversed LPS-induced histopathological changes. These findings demonstrate a significant dose-dependent protection by febuxostat against LPS-induced lung inflammation in rats.

Tissue factor-expressing monocytes inhibit fibrinolysis through a TAFI-mediated mechanism, and make clots resistant to heparins

PubMed Central

Semeraro, Fabrizio; Ammollo, Concetta T.; Semeraro, Nicola; Colucci, Mario

2009-01-01

Background Thrombin is the main activator of the fibrinolysis inhibitor TAFI (thrombin activatable fibrinolysis inhibitor) and heightened clotting activation is believed to impair fibrinolysis through the increase of thrombin activatable fibrinolysis inhibitor activation. However, the enhancement of thrombin generation by soluble tissue factor was reported to have no effect on plasma fibrinolysis and it is not known whether the same is true for cell-associated tissue factor. The aim of this study was to evaluate the effect of tissue factor-expressing monocytes on plasma fibrinolysis in vitro. Design and Methods Tissue factor expression by human blood mononuclear cells (MNC) and monocytes was induced by LPS stimulation. Fibrinolysis was spectrophotometrically evaluated by measuring the lysis time of plasma clots containing LPS-stimulated or control cells and a low concentration of exogenous tissue plasminogen activator. Results LPS-stimulated MNC (LPS-MNC) prolonged fibrinolysis time as compared to unstimulated MNC (C-MNC) in contact-inhibited but not in normal citrated plasma. A significantly prolonged lysis time was observed using as few as 30 activated cells/μL. Fibrinolysis was also impaired when clots were generated on adherent LPS-stimulated monocytes. The antifibrinolytic effect of LPS-MNC or LPS-monocytes was abolished by an anti-tissue factor antibody, by an antibody preventing thrombin-mediated thrombin activatable fibrinolysis inhibitor activation, and by a TAFIa inhibitor (PTCI). Assays of thrombin and TAFIa in contact-inhibited plasma confirmed the greater generation of these enzymes in the presence of LPS-MNC. Finally, the profibrinolytic effect of unfractionated heparin and enoxaparin was markedly lower (~50%) in the presence of LPS-MNC than in the presence of a thromboplastin preparation displaying an identical tissue factor activity. Conclusions Our data indicate that LPS-stimulated monocytes inhibit fibrinolysis through a tissue factor-mediated enhancement of thrombin activatable fibrinolysis inhibitor activation and make clots resistant to the profibrinolytic activity of heparins, thus providing an additional mechanism whereby tissue factor-expressing monocytes/macrophages may favor fibrin accumulation and diminish the antithrombotic efficacy of heparins. PMID:19377079

Activation of Peroxisome Proliferator-activated Receptor γ (PPARγ) and CD36 Protein Expression: THE DUAL PATHOPHYSIOLOGICAL ROLES OF PROGESTERONE.

PubMed

Yang, Xiaoxiao; Zhang, Wenwen; Chen, Yuanli; Li, Yan; Sun, Lei; Liu, Ying; Liu, Mengyang; Yu, Miao; Li, Xiaoju; Han, Jihong; Duan, Yajun

2016-07-15

Progesterone or its analog, one of components of hormone replacement therapy, may attenuate the cardioprotective effects of estrogen. However, the underlying mechanisms have not been fully elucidated. Expression of CD36, a receptor for oxidized LDL (oxLDL) that enhances macrophage/foam cell formation, is activated by the transcription factor peroxisome proliferator-activated receptor γ (PPARγ). CD36 also functions as a fatty acid transporter to influence fatty acid metabolism and the pathophysiological status of several diseases. In this study, we determined that progesterone induced macrophage CD36 expression, which is related to progesterone receptor (PR) activity. Progesterone enhanced cellular oxLDL uptake in a CD36-dependent manner. Mechanistically, progesterone increased PPARγ expression and PPARγ promoter activity in a PR-dependent manner and the binding of PR with the progesterone response element in the PPARγ promoter. Specific deletion of macrophage PPARγ (MφPPARγ KO) expression in mice abolished progesterone-induced macrophage CD36 expression and cellular oxLDL accumulation. We also determined that, associated with gestation and increased serum progesterone levels, CD36 and PPARγ expression in mouse adipose tissue, skeletal muscle, and peritoneal macrophages were substantially activated. Taken together, our study demonstrates that progesterone can play dual pathophysiological roles by activating PPARγ expression, in which progesterone increases macrophage CD36 expression and oxLDL accumulation, a negative effect on atherosclerosis, and enhances the PPARγ-CD36 pathway in adipose tissue and skeletal muscle, a protective effect on pregnancy. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

Arctigenin Efficiently Enhanced Sedentary Mice Treadmill Endurance

PubMed Central

Chen, Jing; Yu, Liang; Hu, Lihong; Jiang, Hualiang; Shen, Xu

2011-01-01

Physical inactivity is considered as one of the potential risk factors for the development of type 2 diabetes and other metabolic diseases, while endurance exercise training could enhance fat oxidation that is associated with insulin sensitivity improvement in obesity. AMP-activated protein kinase (AMPK) as an energy sensor plays pivotal roles in the regulation of energy homeostasis, and its activation could improve glucose uptake, promote mitochondrial biogenesis and increase glycolysis. Recent research has even suggested that AMPK activation contributed to endurance enhancement without exercise. Here we report that the natural product arctigenin from the traditional herb Arctium lappa L. (Compositae) strongly increased AMPK phosphorylation and subsequently up-regulated its downstream pathway in both H9C2 and C2C12 cells. It was discovered that arctigenin phosphorylated AMPK via calmodulin-dependent protein kinase kinase (CaMKK) and serine/threonine kinase 11(LKB1)-dependent pathways. Mice treadmill based in vivo assay further indicated that administration of arctigenin improved efficiently mice endurance as reflected by the increased fatigue time and distance, and potently enhanced mitochondrial biogenesis and fatty acid oxidation (FAO) related genes expression in muscle tissues. Our results thus suggested that arctigenin might be used as a potential lead compound for the discovery of the agents with mimic exercise training effects to treat metabolic diseases. PMID:21887385

Remote limb ischemic conditioning enhances motor learning in healthy humans

PubMed Central

Cherry-Allen, Kendra M.; Gidday, Jeff M.; Lee, Jin-Moo; Hershey, Tamara

2015-01-01

Brief bouts of sublethal ischemia have been shown to protect exposed tissue (ischemic conditioning) and tissues at remote sites (remote ischemic conditioning) against subsequent ischemic challenges. Given that the mechanisms of this protective phenomenon are multifactorial and epigenetic, we postulated that remote limb ischemic conditioning (RLIC) might enhance mechanisms responsible for neural plasticity, and thereby facilitate learning. Specifically, we hypothesized that conditioning of the nervous system with RLIC, achieved through brief repetitive limb ischemia prior to training, would facilitate the neurophysiological processes of learning, thus making training more effective and more long-lasting. Eighteen healthy adults participated in this study; nine were randomly allocated to RLIC and nine to sham conditioning. All subjects underwent seven consecutive weekday sessions and 2-wk and 4-wk follow-up sessions. We found that RLIC resulted in significantly greater motor learning and longer retention of motor performance gains in healthy adults. Changes in motor performance do not appear to be due to a generalized increase in muscle activation or muscle strength and were not associated with changes in serum brain-derived neurotrophic factor (BDNF) concentration. Of note, RLIC did not enhance cognitive learning on a hippocampus-dependent task. While future research is needed to establish optimal conditioning and training parameters, this inexpensive, clinically feasible paradigm might ultimately be implemented to enhance motor learning in individuals undergoing neuromuscular rehabilitation for brain injury and other pathological conditions. PMID:25867743

Insulin antagonises pigment epithelium-derived factor (PEDF)-induced modulation of lineage commitment of myocytes and heterotrophic ossification.

PubMed

Carnagarin, Revathy; Elahy, Mina; Dharmarajan, Arun M; Dass, Crispin R

2017-12-16

Extensive bone defects arising as a result of trauma, infection and tumour resection and other bone pathologies necessitates the identification of effective strategies in the form of tissue engineering, gene therapy and osteoinductive agents to enhance the bone repair process. PEDF is a multifunctional glycoprotein which plays an important role in regulating osteoblastic differentiation and bone formation. PEDF treatment of mice and human skeletal myocytes at physiological concentration inhibited myogenic differentiation and activated Erk1/2 MAPK- dependent osteogenic transdifferentiation of myocytes. In mice, insulin, a promoter of bone regeneration, attenuated PEDF-induced expression of osteogenic markers such as osteocalcin, alkaline phosphatase and mineralisation for bone formation in the muscle and surrounding adipose tissue. These results provide new insights into the molecular aspects of the antagonising effect of insulin on PEDF-dependent modulation of the differentiation commitment of musculoskeletal environment into osteogenesis, and suggest that PEDF may be developed as an effective clinical therapy for bone regeneration as its heterotopic ossification can be controlled via co-administration of insulin. Copyright © 2017 Elsevier B.V. All rights reserved.

The enhanced anti-tissue adhesive effect of injectable pluronic-HA hydrogel by poly(γ-glutamic acid).

PubMed

Kim, Manse; Hwang, Youngmin; Tae, Giyoong

2016-12-01

The stability of tissue barrier in physiological condition is a key factor to isolate the damaged site from adjacent tissue for anti-tissue adhesion. Although pluronic or pluronic-hyaluronic acid (HA) hydrogel as an injectable formulation can prevent tissue adhesion at the injection site, the anti-tissue adhesion effect is limited due to its poor stability. Herein, we prepared tissue barrier formulations composed of pluronic F127 (F127) and HA mixture (F127-HA) and the effect of the addition of poly(γ-glutamic acid) (PGA) was characterized. All of F127, HA, and F127-HA mixture showed the poor in vitro residence stability less than 3 days. However, by adding PGA into F127-HA mixture, their stability was significantly enhanced by the control of the molecular weight and concentration of PGA. Thus, F127-HA with 10wt% PGA (2000kDa) showed the long-term stability over 10 days. Similarly, the enhanced stability of F127-HA with PGA resulted in the enhanced and excellent in vivo anti-tissue adhesion effect, evidenced by histological analysis and grading of tissue adhesion. Therefore, F127-HA containing PGA could be applied as an efficient injectable tissue barrier for anti-tissue adhesion. Copyright © 2016 Elsevier B.V. All rights reserved.

Inhibition of TGF-β Signaling in SHED Enhances Endothelial Differentiation.

PubMed

Xu, J G; Gong, T; Wang, Y Y; Zou, T; Heng, B C; Yang, Y Q; Zhang, C F

2018-02-01

Low efficiency of deriving endothelial cells (ECs) from adult stem cells hampers their utilization in tissue engineering studies. The purpose of this study was to investigate whether suppression of transforming growth factor beta (TGF-β) signaling could enhance the differentiation efficiency of dental pulp-derived stem cells into ECs. We initially used vascular endothelial growth factor A (VEGF-A) to stimulate 2 dental pulp-derived stem cells (dental pulp stem cells and stem cells from human exfoliated deciduous teeth [SHED]) and compared their differentiation capacity into ECs. We further evaluated whether the vascular endothelial growth factor receptor I (VEGF-RI)-specific ligand placental growth factor-1 (PlGF-1) could mediate endothelial differentiation. Finally, we investigated whether the TGF-β signaling inhibitor SB-431542 could enhance the inductive effect of VEGF-A on endothelial differentiation, as well as the underlying mechanisms involved. ECs differentiated from dental pulp-derived stem cells exhibited the typical phenotypes of primary ECs, with SHED possessing a higher endothelial differentiation potential than dental pulp stem cells. VEGFR1-specific ligand-PLGF exerted a negligible effect on SHED-ECs differentiation. Compared with VEGF-A alone, the combination of VEGF-A and SB-431542 significantly enhanced the endothelial differentiation of SHED. The presence of SB-431542 inhibited the phosphorylation of Suppressor of Mothers Against Decapentaplegic 2/3 (SMAD2/3), allowing for VEGF-A-dependent phosphorylation and upregulation of VEGFR2. Our results indicate that the combination of VEGF-A and SB-431542 could enhance the differentiation of dental pulp-derived stem cells into endothelial cells, and this process is mediated through enhancement of VEGF-A-VEGFR2 signaling and concomitant inhibition of TGF-β-SMAD2/3 signaling.

AAV vector encoding human VEGF165-transduced pectineus muscular flaps increase the formation of new tissue through induction of angiogenesis in an in vivo chamber for tissue engineering: A technique to enhance tissue and vessels in microsurgically engineered tissue.

PubMed

Moimas, Silvia; Manasseri, Benedetto; Cuccia, Giuseppe; Stagno d'Alcontres, Francesco; Geuna, Stefano; Pattarini, Lucia; Zentilin, Lorena; Giacca, Mauro; Colonna, Michele R

2015-01-01

In regenerative medicine, new approaches are required for the creation of tissue substitutes, and the interplay between different research areas, such as tissue engineering, microsurgery and gene therapy, is mandatory. In this article, we report a modification of a published model of tissue engineering, based on an arterio-venous loop enveloped in a cross-linked collagen-glycosaminoglycan template, which acts as an isolated chamber for angiogenesis and new tissue formation. In order to foster tissue formation within the chamber, which entails on the development of new vessels, we wondered whether we might combine tissue engineering with a gene therapy approach. Based on the well-described tropism of adeno-associated viral vectors for post-mitotic tissues, a muscular flap was harvested from the pectineus muscle, inserted into the chamber and transduced by either AAV vector encoding human VEGF165 or AAV vector expressing the reporter gene β-galactosidase, as a control. Histological analysis of the specimens showed that muscle transduction by AAV vector encoding human VEGF165 resulted in enhanced tissue formation, with a significant increase in the number of arterioles within the chamber in comparison with the previously published model. Pectineus muscular flap, transduced by adeno-associated viral vectors, acted as a source of the proangiogenic factor vascular endothelial growth factor, thus inducing a consistent enhancement of vessel growth into the newly formed tissue within the chamber. In conclusion, our present findings combine three different research fields such as microsurgery, tissue engineering and gene therapy, suggesting and showing the feasibility of a mixed approach for regenerative medicine.

Integrated microarray and ChIP analysis identifies multiple Foxa2 dependent target genes in the notochord.

PubMed

Tamplin, Owen J; Cox, Brian J; Rossant, Janet

2011-12-15

The node and notochord are key tissues required for patterning of the vertebrate body plan. Understanding the gene regulatory network that drives their formation and function is therefore important. Foxa2 is a key transcription factor at the top of this genetic hierarchy and finding its targets will help us to better understand node and notochord development. We performed an extensive microarray-based gene expression screen using sorted embryonic notochord cells to identify early notochord-enriched genes. We validated their specificity to the node and notochord by whole mount in situ hybridization. This provides the largest available resource of notochord-expressed genes, and therefore candidate Foxa2 target genes in the notochord. Using existing Foxa2 ChIP-seq data from adult liver, we were able to identify a set of genes expressed in the notochord that had associated regions of Foxa2-bound chromatin. Given that Foxa2 is a pioneer transcription factor, we reasoned that these sites might represent notochord-specific enhancers. Candidate Foxa2-bound regions were tested for notochord specific enhancer function in a zebrafish reporter assay and 7 novel notochord enhancers were identified. Importantly, sequence conservation or predictive models could not have readily identified these regions. Mutation of putative Foxa2 binding elements in two of these novel enhancers abrogated reporter expression and confirmed their Foxa2 dependence. The combination of highly specific gene expression profiling and genome-wide ChIP analysis is a powerful means of understanding developmental pathways, even for small cell populations such as the notochord. Copyright © 2011 Elsevier Inc. All rights reserved.

Concentration rather than dose defines the local brain toxicity of agents that are effectively distributed by convection-enhanced delivery.

PubMed

Zhang, Rong; Saito, Ryuta; Mano, Yui; Kanamori, Masayuki; Sonoda, Yukihiko; Kumabe, Toshihiro; Tominaga, Teiji

2014-01-30

Convection-enhanced delivery (CED) has been developed as a potentially effective drug-delivery strategy into the central nervous system. In contrast to systemic intravenous administration, local delivery achieves high concentration and prolonged retention in the local tissue, with increased chance of local toxicity, especially with toxic agents such as chemotherapeutic agents. Therefore, the factors that affect local toxicity should be extensively studied. With the assumption that concentration-oriented evaluation of toxicity is important for local CED, we evaluated the appearance of local toxicity among different agents after delivery with CED and studied if it is dose dependent or concentration dependent. Local toxicity profile of chemotherapeutic agents delivered via CED indicates BCNU was dose-dependent, whereas that of ACNU was concentration-dependent. On the other hand, local toxicity for doxorubicin, which is not distributed effectively by CED, was dose-dependent. Local toxicity for PLD, which is extensively distributed by CED, was concentration-dependent. Traditional evaluation of drug induced toxicity was dose-oriented. This is true for systemic intravascular delivery. However, with local CED, toxicity of several drugs exacerbated in concentration-dependent manner. From our study, local toxicity of drugs that are likely to distribute effectively tended to be concentration-dependent. Concentration rather than dose may be more important for the toxicity of agents that are effectively distributed by CED. Concentration-oriented evaluation of toxicity is more important for CED. Copyright © 2013 Elsevier B.V. All rights reserved.

Experimental evaluation of electrical conductivity imaging of anisotropic brain tissues using a combination of diffusion tensor imaging and magnetic resonance electrical impedance tomography

DOE Office of Scientific and Technical Information (OSTI.GOV)

Sajib, Saurav Z. K.; Jeong, Woo Chul; Oh, Tong In

Anisotropy of biological tissues is a low-frequency phenomenon that is associated with the function and structure of cell membranes. Imaging of anisotropic conductivity has potential for the analysis of interactions between electromagnetic fields and biological systems, such as the prediction of current pathways in electrical stimulation therapy. To improve application to the clinical environment, precise approaches are required to understand the exact responses inside the human body subjected to the stimulated currents. In this study, we experimentally evaluate the anisotropic conductivity tensor distribution of canine brain tissues, using a recently developed diffusion tensor-magnetic resonance electrical impedance tomography method. At lowmore » frequency, electrical conductivity of the biological tissues can be expressed as a product of the mobility and concentration of ions in the extracellular space. From diffusion tensor images of the brain, we can obtain directional information on diffusive movements of water molecules, which correspond to the mobility of ions. The position dependent scale factor, which provides information on ion concentration, was successfully calculated from the magnetic flux density, to obtain the equivalent conductivity tensor. By combining the information from both techniques, we can finally reconstruct the anisotropic conductivity tensor images of brain tissues. The reconstructed conductivity images better demonstrate the enhanced signal intensity in strongly anisotropic brain regions, compared with those resulting from previous methods using a global scale factor.« less

Evolution of Nova-Dependent Splicing Regulation in the Brain

PubMed Central

Živin, Marko; Darnell, Robert B

2007-01-01

A large number of alternative exons are spliced with tissue-specific patterns, but little is known about how such patterns have evolved. Here, we study the conservation of the neuron-specific splicing factors Nova1 and Nova2 and of the alternatively spliced exons they regulate in mouse brain. Whereas Nova RNA binding domains are 94% identical across vertebrate species, Nova-dependent splicing silencer and enhancer elements (YCAY clusters) show much greater divergence, as less than 50% of mouse YCAY clusters are conserved at orthologous positions in the zebrafish genome. To study the relation between the evolution of tissue-specific splicing and YCAY clusters, we compared the brain-specific splicing of Nova-regulated exons in zebrafish, chicken, and mouse. The presence of YCAY clusters in lower vertebrates invariably predicted conservation of brain-specific splicing across species, whereas their absence in lower vertebrates correlated with a loss of alternative splicing. We hypothesize that evolution of Nova-regulated splicing in higher vertebrates proceeds mainly through changes in cis-acting elements, that tissue-specific splicing might in some cases evolve in a single step corresponding to evolution of a YCAY cluster, and that the conservation level of YCAY clusters relates to the functions encoded by the regulated RNAs. PMID:17937501

Antisense oligonucleotide–mediated MDM4 exon 6 skipping impairs tumor growth

PubMed Central

Dewaele, Michael; Tabaglio, Tommaso; Willekens, Karen; Bezzi, Marco; Teo, Shun Xie; Low, Diana H.P.; Koh, Cheryl M.; Rambow, Florian; Fiers, Mark; Rogiers, Aljosja; Radaelli, Enrico; Al-Haddawi, Muthafar; Tan, Soo Yong; Hermans, Els; Amant, Frederic; Yan, Hualong; Lakshmanan, Manikandan; Koumar, Ratnacaram Chandrahas; Lim, Soon Thye; Derheimer, Frederick A.; Campbell, Robert M.; Bonday, Zahid; Tergaonkar, Vinay; Shackleton, Mark; Blattner, Christine; Marine, Jean-Christophe; Guccione, Ernesto

2015-01-01

MDM4 is a promising target for cancer therapy, as it is undetectable in most normal adult tissues but often upregulated in cancer cells to dampen p53 tumor-suppressor function. The mechanisms that underlie MDM4 upregulation in cancer cells are largely unknown. Here, we have shown that this key oncogenic event mainly depends on a specific alternative splicing switch. We determined that while a nonsense-mediated, decay-targeted isoform of MDM4 (MDM4-S) is produced in normal adult tissues as a result of exon 6 skipping, enhanced exon 6 inclusion leads to expression of full-length MDM4 in a large number of human cancers. Although this alternative splicing event is likely regulated by multiple splicing factors, we identified the SRSF3 oncoprotein as a key enhancer of exon 6 inclusion. In multiple human melanoma cell lines and in melanoma patient–derived xenograft (PDX) mouse models, antisense oligonucleotide–mediated (ASO-mediated) skipping of exon 6 decreased MDM4 abundance, inhibited melanoma growth, and enhanced sensitivity to MAPK-targeting therapeutics. Additionally, ASO-based MDM4 targeting reduced diffuse large B cell lymphoma PDX growth. As full-length MDM4 is enhanced in multiple human tumors, our data indicate that this strategy is applicable to a wide range of tumor types. We conclude that enhanced MDM4 exon 6 inclusion is a common oncogenic event and has potential as a clinically compatible therapeutic target. PMID:26595814

Dynamic reciprocity in cell-scaffold interactions.

PubMed

Mauney, Joshua R; Adam, Rosalyn M

2015-03-01

Tissue engineering in urology has shown considerable promise. However, there is still much to understand, particularly regarding the interactions between scaffolds and their host environment, how these interactions regulate regeneration and how they may be enhanced for optimal tissue repair. In this review, we discuss the concept of dynamic reciprocity as applied to tissue engineering, i.e. how bi-directional signaling between implanted scaffolds and host tissues such as the bladder drives the process of constructive remodeling to ensure successful graft integration and tissue repair. The impact of scaffold content and configuration, the contribution of endogenous and exogenous bioactive factors, the influence of the host immune response and the functional interaction with mechanical stimulation are all considered. In addition, the temporal relationships of host tissue ingrowth, bioactive factor mobilization, scaffold degradation and immune cell infiltration, as well as the reciprocal signaling between discrete cell types and scaffolds are discussed. Improved understanding of these aspects of tissue repair will identify opportunities for optimization of repair that could be exploited to enhance regenerative medicine strategies for urology in future studies. Copyright © 2014 Elsevier B.V. All rights reserved.

IL-33 promotes an innate immune pathway of intestinal tissue protection dependent on amphiregulin-EGFR interactions.

PubMed

Monticelli, Laurel A; Osborne, Lisa C; Noti, Mario; Tran, Sara V; Zaiss, Dietmar M W; Artis, David

2015-08-25

The barrier surfaces of the skin, lung, and intestine are constantly exposed to environmental stimuli that can result in inflammation and tissue damage. Interleukin (IL)-33-dependent group 2 innate lymphoid cells (ILC2s) are enriched at barrier surfaces and have been implicated in promoting inflammation; however, the mechanisms underlying the tissue-protective roles of IL-33 or ILC2s at surfaces such as the intestine remain poorly defined. Here we demonstrate that, following activation with IL-33, expression of the growth factor amphiregulin (AREG) is a dominant functional signature of gut-associated ILC2s. In the context of a murine model of intestinal damage and inflammation, the frequency and number of AREG-expressing ILC2s increases following intestinal injury and genetic disruption of the endogenous AREG-epidermal growth factor receptor (EGFR) pathway exacerbated disease. Administration of exogenous AREG limited intestinal inflammation and decreased disease severity in both lymphocyte-sufficient and lymphocyte-deficient mice, revealing a previously unrecognized innate immune mechanism of intestinal tissue protection. Furthermore, treatment with IL-33 or transfer of ILC2s ameliorated intestinal disease severity in an AREG-dependent manner. Collectively, these data reveal a critical feedback loop in which cytokine cues from damaged epithelia activate innate immune cells to express growth factors essential for ILC-dependent restoration of epithelial barrier function and maintenance of tissue homeostasis.

Compound parabolic concentrator probe for efficient light collection in spectroscopy of biological tissue

NASA Astrophysics Data System (ADS)

Tanaka, Kazunori; Pacheco, Marcos T. T.; Brennan, James F., III; Itzkan, Irving; Berger, Andrew J.; Dasari, Ramachandra R.; Feld, Michael S.

1996-02-01

We describe a compound parabolic concentrator (CPC)-based probe for enhanced signal collection in the spectroscopy of biological tissues. Theoretical considerations governing signal enhancement compared with conventional collection methods are given. A ray-tracing program was used to analyze the throughput of CPC's with shape deviations and surface imperfections. A modified CPC shape with 99% throughput was discovered. A 4.4-mm-long CPC was manufactured and incorporated into an optical fiber-based near-infrared Raman spectrometer system. For human tissue samples, light collection was enhanced by a factor of 7 compared with collection with 0.29-NA optical fibers.

Biochemical and molecular mechanisms for the association between obesity, chronic inflammation, and breast cancer.

PubMed

Rose, David P; Vona-Davis, Linda

2014-01-01

Upper body obesity is a risk factor for postmenopausal breast cancer and is related to an aggressive tumor phenotype and a poor prognosis regardless of menopausal status. After the menopause, the major mechanism for the association with disease risk is elevated estrogen production by adipose tissue, due to a high level of aromatase activity: these hormone-dependent tumors express both estrogen and progesterone receptors. Other important biological factors of risk include leptin and adiponectin, adipokines with opposing endocrine and paracrine activities, and obesity-related hyperinsulinemia. Chronic inflammation of the breast adipose tissue, which occurs in some obese women and is indicated by the accumulation of macrophages around dead adipocytes ("crown-like structures"), rather than adiposity per se, may prove to be the pathological lesion responsible for both local aromatase induction, and enhanced invasiveness and metastatic capacity through biological mechanisms that involve leptin, tumor necrosis factor-α, and insulin. A causal association between obesity in premenopausal women and breast cell epithelial-mesenchymal transition, perhaps with the participation of the Wnt signaling pathway, and aggressive hormone-independent breast cancer is suggested by a number of experimental and clinical studies. © 2013 International Union of Biochemistry and Molecular Biology.

Dose in bone and tissue near bone-tissue interface from electron beam.

PubMed

Shiu, A S; Hogstrom, K R

1991-08-01

This work has quantitatively studied the variation of dose both within bone and in unit density tissue near bone-tissue interfaces. Dose upstream of a bone-tissue interface is increased because of an increase in the backscattered electrons from the bone. The magnitude of this effect was measured using a thin parallel-plate ionization chamber upstream of a polymethyl methacrylate (PMMA)-hard bone interface. The electron backscatter factor (EBF) increased rapidly with bone thickness until a full EBF was achieved. This occurred at approximately 3.5 mm at 2 MeV and 6 mm at 13.1 MeV. The full EBF at the interface ranged from approximately 1.018 at 13.1 MeV to 1.05 at 2 MeV. It was also observed that the EBF had a dependence on the energy spectrum at the interface. The penetration of the backscattered electrons in the upstream direction of PMMA was also measured. The dose penetration fell off rapidly in the upstream direction of the interface. Dose enhancement to unit density tissue in bone was measured for an electron beam by placing thermoluminescent dosimeters (TLDs) in a PMMA-bone-PMMA phantom. The maximum dose enhancement in bone was approximately 7% of the maximum dose in water. However, the pencil-beam algorithm of Hogstrom et al. predicted an increase of only 1%, primarily owing to the inverse-square correction. Film was also used to measure the dose enhancement in bone. The film plane was aligned either perpendicular or parallel to the central axis of the beam. The film data indicated that the maximum dose enhancement in bone was approximately 8% for the former film alignment (which was similarly predicted by the TLD measurements) and 13% for the latter film alignment. These results confirm that the X ray film is not suitable to be irritated "edge on" in an inhomogeneous phantom without making perturbation corrections resulting from the film acting as a long narrow inhomogeneous cavity within the bone. In addition, the results give the radiotherapist a basis for clinical judgment when electron beams are used to treat lesions behind bone or near bony structures. We feel these data enhance the ability to recognize the shortcomings of the current dose calculation algorithm used clinically.

25-Hydroxycholesterol promotes fibroblast-mediated tissue remodeling through NF-κB dependent pathway

DOE Office of Scientific and Technical Information (OSTI.GOV)

Ichikawa, Tomohiro; Sugiura, Hisatoshi, E-mail: sugiura@rm.med.tohoku.ac.jp; Koarai, Akira

2013-05-01

Abnormal structural alterations termed remodeling, including fibrosis and alveolar wall destruction, are important features of the pathophysiology of chronic airway diseases such as chronic obstructive pulmonary disease (COPD) and asthma. 25-hydroxycholesterol (25-HC) is enzymatically produced by cholesterol 25-hydorxylase (CH25H) in macrophages and is reported to be involved in the formation of arteriosclerosis. We previously demonstrated that the expression of CH25H and production of 25HC were increased in the lungs of COPD. However, the role of 25-HC in lung tissue remodeling is unknown. In this study, we investigated the effect of 25-HC on fibroblast-mediated tissue remodeling using human fetal lung fibroblastsmore » (HFL-1) in vitro. 25-HC significantly augmented α-smooth muscle actin (SMA) (P<0.001) and collagen I (P<0.001) expression in HFL-1. 25-HC also significantly enhanced the release and activation of matrix metallaoproteinase (MMP)-2 (P<0.001) and MMP-9 (P<0.001) without any significant effect on the production of tissue inhibitor of metalloproteinase (TIMP)-1 and TIMP-2. 25-HC stimulated transforming growth factor (TGF)-β{sub 1} production (P<0.01) and a neutralizing anti-TGF-β antibody restored these 25-HC-augmented pro-fibrotic responses. 25-HC significantly promoted the translocation of nuclear factor (NF)-κB p65 into the nuclei (P<0.01), but not phospholylated-c-jun, a complex of activator protein-1. Pharmacological inhibition of NF-κB restored the 25-HC-augmented pro-fibrotic responses and TGF-β{sub 1} release. These results suggest that 25-HC could contribute to fibroblast-mediated lung tissue remodeling by promoting myofibroblast differentiation and the excessive release of extracellular matrix protein and MMPs via an NF-κB-TGF-β dependent pathway.« less

Effects of insulin, triiodothyronine and fat soluble vitamins on adipocyte differentiation and LPL gene expression in the stromal-vascular cells of red sea bream, Pagrus major.

PubMed

Oku, Hiromi; Tokuda, Masaharu; Okumura, Takuji; Umino, Tetsuya

2006-07-01

Various kinds of hormones including insulin, triiodothyronine (T(3)) and fat-soluble vitamins have been proposed as mediators of adipocyte differentiation in mammals. To investigate the factors which are responsible for fish adipocyte differentiation, we developed a serum-free culture system of stromal-vascular cells of red sea bream adipose tissue and examined the effects of bovine insulin, T(3), and fat-soluble vitamins (all-trans retinoic acid, retinyl acetate and 1,25-dihydroxyvitamin D(3)) on the differentiation-linked expression of the lipoprotein lipase (LPL) gene. As assessed by the increase in LPL gene expression after 3 day cultivation, like in mammalian adipocytes, insulin enhanced the adipocyte differentiation in a concentration-dependent manner. During 2 week cultivation, bovine insulin promoted lipid accumulation in differentiating adipocytes concentration-dependently until the terminal differentiation. These results indicate that the differentiation of fish adipocytes is inducible by insulin alone. T(3) alone had no effect but enhanced the differentiation-linked LPL gene expression in the presence of insulin. Fat-soluble vitamins, unlike in mammalian adipocytes, did not show any significant effects. The method developed in this study should be of interest for the characterization of factors involved in fish adipocyte differentiation.

Macroporous Hydrogel Scaffolds for Three-Dimensional Cell Culture and Tissue Engineering.

PubMed

Fan, Changjiang; Wang, Dong-An

2017-10-01

Hydrogels have been promising candidate scaffolds for cell delivery and tissue engineering due to their tissue-like physical properties and capability for homogeneous cell loading. However, the encapsulated cells are generally entrapped and constrained in the submicron- or nanosized gel networks, seriously limiting cell growth and tissue formation. Meanwhile, the spatially confined settlement inhibits attachment and spreading of anchorage-dependent cells, leading to their apoptosis. In recent years, macroporous hydrogels have attracted increasing attention in use as cell delivery vehicles and tissue engineering scaffolds. The introduction of macropores within gel scaffolds not only improves their permeability for better nutrient transport but also creates space/interface for cell adhesion, proliferation, and extracellular matrix deposition. Herein, we will first review the development of macroporous gel scaffolds and outline the impact of macropores on cell behaviors. In the first part, the advantages and challenges of hydrogels as three-dimensional (3D) cell culture scaffolds will be described. In the second part, the fabrication of various macroporous hydrogels will be presented. Third, the enhancement of cell activities within macroporous gel scaffolds will be discussed. Finally, several crucial factors that are envisaged to propel the improvement of macroporous gel scaffolds are proposed for 3D cell culture and tissue engineering.

Selective tissue factor/factor VIIa Inhibitor, ER-410660, and its prodrug, E5539, have anti-venous and anti-arterial thrombotic effects with a low risk of bleeding.

PubMed

Nagakura, Tadashi; Tabata, Kimiyo; Kira, Kazunobu; Hirota, Shinsuke; Clark, Richard; Matsuura, Fumiyoshi; Hiyoshi, Hironobu

2013-08-01

Many anticoagulant drugs target factors common to both the intrinsic and extrinsic coagulation pathways, which may lead to bleeding complications. Since the tissue factor (TF)/factor VIIa complex is associated with thrombosis onset and specifically activates the extrinsic coagulation pathway, compounds that inhibit this complex may provide therapeutic and/or prophylactic benefits with a decreased risk of bleeding. The in vitro enzyme profile and anticoagulation selectivity of the TF/VIIa complex inhibitor, ER-410660, and its prodrug E5539 were assessed using enzyme inhibitory and plasma clotting assays. In vivo effects of ER-410660 and E5539 were determined using a TF-induced, thrombin generation rhesus monkey model; a stasis-induced, venous thrombosis rat model; a photochemically induced, arterial thrombosis rat model; and a rat tail-cut bleeding model. ER-410660 selectively prolonged prothrombin time, but had a less potent anticoagulant effect on the intrinsic pathway. It also exhibited a dose-dependent inhibitory effect on thrombin generation caused by TF-injection in the rhesus monkey model. ER-410660 also reduced venous thrombus weights in the TF-administered, stasis-induced, venous thrombosis rat model and prolonged the occlusion time induced by arterial thrombus formation after vascular injury. The compound was capable of doubling the total bleeding time in the rat tail-cut model, albeit with a considerably higher dose compared to the effective dose in the venous and arterial thrombosis models. Moreover, E5539, an orally available ER-410660 prodrug, reduced the thrombin-anti-thrombin complex levels, induced by TF-injection, in a dose-dependent manner. Selective TF/VIIa inhibitors have potential as novel anticoagulants with a lower propensity for enhancing bleeding. Copyright © 2013 Elsevier Ltd. All rights reserved.

Ubiquitin fusion expression and tissue-dependent targeting of hG-CSF in transgenic tobacco

PubMed Central

2011-01-01

Background Human granulocyte colony-stimulating factor (hG-CSF) is an important human cytokine which has been widely used in oncology and infection protection. To satisfy clinical needs, expression of recombinant hG-CSF has been studied in several organisms, including rice cell suspension culture and transient expression in tobacco leaves, but there was no published report on its expression in stably transformed plants which can serve as a more economical expression platform with potential industrial application. Results In this study, hG-CSF expression was investigated in transgenic tobacco leaves and seeds in which the accumulation of hG-CSF could be enhanced through fusion with ubiquitin by up to 7 fold in leaves and 2 fold in seeds, leading to an accumulation level of 2.5 mg/g total soluble protein (TSP) in leaves and 1.3 mg/g TSP in seeds, relative to hG-CSF expressed without a fusion partner. Immunoblot analysis showed that ubiquitin was processed from the final protein product, and ubiquitination was up-regulated in all transgenic plants analyzed. Driven by CaMV 35S promoter and phaseolin signal peptide, hG-CSF was observed to be secreted into apoplast in leaves but deposited in protein storage vacuole (PSV) in seeds, indicating that targeting of the hG-CSF was tissue-dependent in transgenic tobacco. Bioactivity assay showed that hG-CSF expressed in both seeds and leaves was bioactive to support the proliferation of NFS-60 cells. Conclusions In this study, the expression of bioactive hG-CSF in transgenic plants was improved through ubiquitin fusion strategy, demonstrating that protein expression can be enhanced in both plant leaves and seeds through fusion with ubiquitin and providing a typical case of tissue-dependent expression of recombinant protein in transgenic plants. PMID:21985646

Experimentally determined spectral optimization for dedicated breast computed tomography.

PubMed

Prionas, Nicolas D; Huang, Shih-Ying; Boone, John M

2011-02-01

The current study aimed to experimentally identify the optimal technique factors (x-ray tube potential and added filtration material/thickness) to maximize soft-tissue contrast, microcalcification contrast, and iodine contrast enhancement using cadaveric breast specimens imaged with dedicated breast computed tomography (bCT). Secondarily, the study aimed to evaluate the accuracy of phantom materials as tissue surrogates and to characterize the change in accuracy with varying bCT technique factors. A cadaveric breast specimen was acquired under appropriate approval and scanned using a prototype bCT scanner. Inserted into the specimen were cylindrical inserts of polyethylene, water, iodine contrast medium (iodixanol, 2.5 mg/ml), and calcium hydroxyapatite (100 mg/ml). Six x-ray tube potentials (50, 60, 70, 80, 90, and 100 kVp) and three different filters (0.2 mm Cu, 1.5 mm Al, and 0.2 mm Sn) were tested. For each set of technique factors, the intensity (linear attenuation coefficient) and noise were measured within six regions of interest (ROIs): Glandular tissue, adipose tissue, polyethylene, water, iodine contrast medium, and calcium hydroxyapatite. Dose-normalized contrast to noise ratio (CNRD) was measured for pairwise comparisons among the six ROIs. Regression models were used to estimate the effect of tube potential and added filtration on intensity, noise, and CNRD. Iodine contrast enhancement was maximized using 60 kVp and 0.2 mm Cu. Microcalcification contrast and soft-tissue contrast were maximized at 60 kVp. The 0.2 mm Cu filter achieved significantly higher CNRD for iodine contrast enhancement than the other two filters (p = 0.01), but microcalcification contrast and soft-tissue contrast were similar using the copper and aluminum filters. The average percent difference in linear attenuation coefficient, across all tube potentials, for polyethylene versus adipose tissue was 1.8%, 1.7%, and 1.3% for 0.2 mm Cu, 1.5 mm Al, and 0.2 mm Sn, respectively. For water versus glandular tissue, the average percent difference was 2.7%, 3.9%, and 4.2% for the three filter types. Contrast-enhanced bCT, using injected iodine contrast medium, may be optimized for maximum contrast of enhancing lesions at 60 kVp with 0.2 mm Cu filtration. Soft-tissue contrast and microcalcification contrast may also benefit from lower tube potentials (60 kVp). The linear attenuation coefficients of water and polyethylene slightly overestimate the values of their corresponding tissues, but the reported differences may serve as guidance for dosimetry and quality assurance using tissue equivalent phantoms.

Iloprost up-regulates vascular endothelial growth factor expression in human dental pulp cells in vitro and enhances pulpal blood flow in vivo.

PubMed

Limjeerajarus, Chalida Nakalekha; Osathanon, Thanaphum; Manokawinchoke, Jeeranan; Pavasant, Prasit

2014-07-01

Prostacyclin (PGI2) is a biomolecule capable of enhancing angiogenesis and cellular proliferation. We investigated the influence of a PGI2 analogue (iloprost) on dental pulp revascularization in vitro and in vivo by using human dental pulp cells (HDPCs) and a rat tooth injury model, respectively. Iloprost stimulated the human dental pulp cell mRNA expression of vascular endothelial growth factor (VEGF), fibroblast growth factor-2 (FGF-2), and platelet-derived growth factor (PDGF) in a significant dose-dependent manner. This mRNA up-regulation was significantly inhibited by pretreatment with a PGI2 receptor antagonist and forskolin (a protein kinase A activator). In contrast, a protein kinase A inhibitor significantly enhanced the iloprost-induced mRNA expression of VEGF, FGF-2, and PDGF. Pretreatment with a fibroblast growth factor receptor inhibitor attenuated the VEGF, FGF-2, and PDGF mRNA expression, indicating opposing regulatory mechanisms. The effect of iloprost on the dental pulp was investigated in vivo by using a rat molar pulp injury model. The iloprost-treated group exhibited a significant increase in pulpal blood flow at 72 hours compared with control. The present study indicates that iloprost may be a candidate agent to promote neovascularization in dental pulp tissue, suggesting the potential clinical use of iloprost in vital pulp therapy. Copyright © 2014 American Association of Endodontists. Published by Elsevier Inc. All rights reserved.

Time-dependent photon migration imaging

NASA Astrophysics Data System (ADS)

Sevick, Eva M.; Wang, NaiGuang; Chance, Britton

1992-02-01

Recently, the application of both time- and frequency-resolved fluorescence techniques for the determination of photon migration characteristics in strongly scattering media has been used to characterize the optical properties in strongly scattering media. Specifically, Chance and coworkers have utilized measurement of photon migration characteristics to determine tissue hemoglobin absorbance and ultimately oxygenation status in homogeneous tissues. In this study, we present simulation results and experimental measurements for both techniques to show the capacity of time-dependent photon migration characteristics to image optically obscure absorbers located in strongly scattering media. The applications of time-dependent photon imaging in the biomedical community include imaging of light absorbing hematomas, tumors, hypoxic tissue volumes, and other tissue abnormalities. Herein, we show that the time-resolved parameter of mean photon path length, , and the frequency- resolved parameter of phase-shift, (theta) , can be used similarly to obtain three dimensional information of absorber position from two-dimensional measurements. Finally, we show that unlike imaging techniques that monitor the intensity of light without regard to the migration characteristics, the resolution of time-dependent photon migration measurements is enhanced by tissue scattering, further potentiating their use for biomedical imaging.

Priming Dental Pulp Stem Cells With Fibroblast Growth Factor-2 Increases Angiogenesis of Implanted Tissue-Engineered Constructs Through Hepatocyte Growth Factor and Vascular Endothelial Growth Factor Secretion

PubMed Central

Gorin, Caroline; Rochefort, Gael Y.; Bascetin, Rumeyza; Ying, Hanru; Lesieur, Julie; Sadoine, Jérémy; Beckouche, Nathan; Berndt, Sarah; Novais, Anita; Lesage, Matthieu; Hosten, Benoit; Vercellino, Laetitia; Merlet, Pascal; Le-Denmat, Dominique; Marchiol, Carmen; Letourneur, Didier; Nicoletti, Antonino; Vital, Sibylle Opsahl; Poliard, Anne; Salmon, Benjamin; Germain, Stéphane

2016-01-01

Tissue engineering strategies based on implanting cellularized biomaterials are promising therapeutic approaches for the reconstruction of large tissue defects. A major hurdle for the reliable establishment of such therapeutic approaches is the lack of rapid blood perfusion of the tissue construct to provide oxygen and nutrients. Numerous sources of mesenchymal stem cells (MSCs) displaying angiogenic potential have been characterized in the past years, including the adult dental pulp. Establishment of efficient strategies for improving angiogenesis in tissue constructs is nevertheless still an important challenge. Hypoxia was proposed as a priming treatment owing to its capacity to enhance the angiogenic potential of stem cells through vascular endothelial growth factor (VEGF) release. The present study aimed to characterize additional key factors regulating the angiogenic capacity of such MSCs, namely, dental pulp stem cells derived from deciduous teeth (SHED). We identified fibroblast growth factor-2 (FGF-2) as a potent inducer of the release of VEGF and hepatocyte growth factor (HGF) by SHED. We found that FGF-2 limited hypoxia-induced downregulation of HGF release. Using three-dimensional culture models of angiogenesis, we demonstrated that VEGF and HGF were both responsible for the high angiogenic potential of SHED through direct targeting of endothelial cells. In addition, FGF-2 treatment increased the fraction of Stro-1+/CD146+ progenitor cells. We then applied in vitro FGF-2 priming to SHED before encapsulation in hydrogels and in vivo subcutaneous implantation. Our results showed that FGF-2 priming is more efficient than hypoxia at increasing SHED-induced vascularization compared with nonprimed controls. Altogether, these data demonstrate that FGF-2 priming enhances the angiogenic potential of SHED through the secretion of both HGF and VEGF. Significance The results from the present study show that fibroblast growth factor-2 (FGF-2) priming is more efficient than hypoxia at increasing dental pulp stem cells derived from deciduous teeth (SHED)-induced vascularization compared with nonprimed controls. Together, these data demonstrate that FGF-2 priming enhances the angiogenic potential of SHED through the secretion of both hepatocyte growth factor and vascular endothelial growth factor. PMID:26798059

Kinetic Model Facilitates Analysis of Fibrin Generation and Its Modulation by Clotting Factors: Implications for Hemostasis-Enhancing Therapies

DTIC Science & Technology

2014-01-01

facilitates analysis of fibrin generation and its modulation by clotting factors : implications for hemostasis-enhancing therapies† Alexander Y...investigate the ability of fibrinogen and a recently proposed prothrombin complex concentrate composition, PCC-AT (a combination of the clotting factors II...kinetics. Moreover, the model qualitatively predicted the impact of tissue factor and tPA/tenecteplase level variations on the fibrin output. In the

A coherent transcriptional feed-forward motif model for mediating auxin-sensitive PIN3 expression during lateral root development

PubMed Central

Chen, Qian; Liu, Yang; Maere, Steven; Lee, Eunkyoung; Van Isterdael, Gert; Xie, Zidian; Xuan, Wei; Lucas, Jessica; Vassileva, Valya; Kitakura, Saeko; Marhavý, Peter; Wabnik, Krzysztof; Geldner, Niko; Benková, Eva; Le, Jie; Fukaki, Hidehiro; Grotewold, Erich; Li, Chuanyou; Friml, Jiří; Sack, Fred; Beeckman, Tom; Vanneste, Steffen

2015-01-01

Multiple plant developmental processes, such as lateral root development, depend on auxin distribution patterns that are in part generated by the PIN-formed family of auxin-efflux transporters. Here we propose that AUXIN RESPONSE FACTOR7 (ARF7) and the ARF7-regulated FOUR LIPS/MYB124 (FLP) transcription factors jointly form a coherent feed-forward motif that mediates the auxin-responsive PIN3 transcription in planta to steer the early steps of lateral root formation. This regulatory mechanism might endow the PIN3 circuitry with a temporal ‘memory' of auxin stimuli, potentially maintaining and enhancing the robustness of the auxin flux directionality during lateral root development. The cooperative action between canonical auxin signalling and other transcription factors might constitute a general mechanism by which transcriptional auxin-sensitivity can be regulated at a tissue-specific level. PMID:26578065

High cancer-specific expression of mesothelin (MSLN) is attributable to an upstream enhancer containing a transcription enhancer factor dependent MCAT motif.

PubMed

Hucl, Tomas; Brody, Jonathan R; Gallmeier, Eike; Iacobuzio-Donahue, Christine A; Farrance, Iain K; Kern, Scott E

2007-10-01

Identification of genes with cancer-specific overexpression offers the potential to efficiently discover cancer-specific activities in an unbiased manner. We apply this paradigm to study mesothelin (MSLN) overexpression, a nearly ubiquitous, diagnostically and therapeutically useful characteristic of pancreatic cancer. We identified an 18-bp upstream enhancer, termed CanScript, strongly activating transcription from an otherwise weak tissue-nonspecific promoter and operating selectively in cells having aberrantly elevated cancer-specific MSLN transcription. Introducing mutations into CanScript showed two functionally distinct sites: an Sp1-like site and an MCAT element. Gel retardation and chromatin immunoprecipitation assays showed the MCAT element to be bound by transcription enhancer factor (TEF)-1 (TEAD1) in vitro and in vivo. The presence of TEF-1 was required for MSLN protein overexpression as determined by TEF-1 knockdown experiments. The cancer specificity seemed to be provided by a putative limiting cofactor of TEF-1 that could be outcompeted by exogenous TEF-1 only in a MSLN-overexpressing cell line. A CanScript concatemer offered enhanced activity. These results identify a TEF family member as a major regulator of MSLN overexpression, a fundamental characteristic of pancreatic and other cancers, perhaps due to an upstream and highly frequent aberrant cellular activity. The CanScript sequence represents a modular element for cancer-specific targeting, potentially suitable for nearly a third of human malignancies.

Anti-inflammatory and anti-fibrinolytic effects of thrombomodulin alfa through carboxypeptidase B2 in the presence of thrombin.

PubMed

Tawara, Shunsuke; Sakai, Takumi; Matsuzaki, Osamu

2016-11-01

Thrombomodulin (TM) alfa, a recombinant human soluble TM, enhances activation of pro-carboxypeptidase B2 (pro-CPB2) by thrombin. Activated pro-CPB2 (CPB2) exerts anti-inflammatory and anti-fibrinolytic activities. Therefore, TM alfa may also have anti-inflammatory and anti-fibrinolytic effects through CPB2. However, these effects of TM alfa have not been elucidated. In the present study, we investigated the effects of TM alfa on inactivation of complement component C5a as an anti-inflammatory effect and prolongation of clot lysis time as an anti-fibrinolytic effect via CPB2 in vitro. CPB2 activity and tissue factor-induced thrombin generation was examined by a chromogenic assay. C5a inactivation was evaluated by C-terminal cleavage of C5a and inhibition of C5a-induced human neutrophil migration. Clot lysis time prolongation was examined by a tissue-type plasminogen activator-induced clot lysis assay. CPB2 activity in human plasma was increased by TM alfa and thrombin in a concentration-dependent manner. TM alfa inhibited tissue factor-induced thrombin generation and enhanced pro-CPB2 activation in human plasma simultaneously. The mass spectrum of C5a treated with TM alfa, thrombin, and pro-CPB2 was decreased at 156m/z, indicating that TM alfa enhanced the processing of C5a to C-terminal-cleaved C5a, an inactive form of C5a. C5a-induced human neutrophil migration was decreased after C5a treatment with TM alfa, thrombin, and pro-CPB2. TM alfa prolonged the clot lysis time in human plasma, and this effect was completely abolished by addition of a CPB2 inhibitor. TM alfa exerts anti-inflammatory and anti-fibrinolytic effects through CPB2 in the presence of thrombin in vitro. Copyright © 2016 Elsevier Ltd. All rights reserved.

[Diabetic retinopathy: pathogenesis and therapeutic implications].

PubMed

Pelikánová, Terezie

Diabetic retinopathy (DR) develops in patients with both type 1 and type 2 diabetes and is the major cause of vision loss and blindness in the working population. The main risk factor of DR is hyperglycemia accompanied by enhanced mitochondrial production of reactive oxygen species and oxidative stress, formation of advanced glycation end products (AGE) and hexosamines, increase in polyol metabolism of glucose. The severity of vascular injury depends on the individual genetic background and is modified by other epigenetic, metabolic and haemodynamic factors, including hypertension, dyslipidemia and oxidative stress. In diabetes, damage to the retina occurs in the vasculature (endothelial cells and pericytes), neurons and glia, pigment epithelial cells and infiltrating immunocompetent cells: monocytes, granulocytes, lymfocytes. These activated cells change the production pattern of a number of mediators such as growth factors, proinflammatory cytokines, vasoactive molecules, coagulation factors and adhesion molecules resulting in increased blood flow, increased capillary permeability, proliferation of extracellular matrix and thickening of basal membranes, altered cell turnover (apoptosis, proliferation, hypertrophy), procoagulant and proaggregant pattern, and finally in angiogenesis and tissue remodelling. Brain, liver, adipose tissue, GUT, skeletal muscle and other tissues could be another source of mediators. Therapeutic approaches used for patients with or at risk for diabetic retinopathy include drug therapy to reduce modifiable risk factors, laser photocoagulation, intravitreous administration of anti-VEGF agents/steroids and intraocular surgery. Screening plays an important role in early detection and intervention to prevent the progression of diabetic retinopathy. Described insights into pathophysiological mechanisms responsible for DR, could help in the development of more targeted approach for prevention and treatment of diabetic retinopathy. anti-VEGF - diabetic dyslipidemia - growth factors - hyperglycemia - hypertension - microRNA - oxidative stress - retinopathy.

Multiplex, quantitative cellular analysis in large tissue volumes with clearing-enhanced 3D microscopy (Ce3D)

PubMed Central

Li, Weizhe; Germain, Ronald N.

2017-01-01

Organ homeostasis, cellular differentiation, signal relay, and in situ function all depend on the spatial organization of cells in complex tissues. For this reason, comprehensive, high-resolution mapping of cell positioning, phenotypic identity, and functional state in the context of macroscale tissue structure is critical to a deeper understanding of diverse biological processes. Here we report an easy to use method, clearing-enhanced 3D (Ce3D), which generates excellent tissue transparency for most organs, preserves cellular morphology and protein fluorescence, and is robustly compatible with antibody-based immunolabeling. This enhanced signal quality and capacity for extensive probe multiplexing permits quantitative analysis of distinct, highly intermixed cell populations in intact Ce3D-treated tissues via 3D histo-cytometry. We use this technology to demonstrate large-volume, high-resolution microscopy of diverse cell types in lymphoid and nonlymphoid organs, as well as to perform quantitative analysis of the composition and tissue distribution of multiple cell populations in lymphoid tissues. Combined with histo-cytometry, Ce3D provides a comprehensive strategy for volumetric quantitative imaging and analysis that bridges the gap between conventional section imaging and disassociation-based techniques. PMID:28808033

An extended model based on the modified Nernst-Planck equation for describing transdermal iontophoresis of weak electrolytes.

PubMed

Imanidis, Georgios; Luetolf, Peter

2006-07-01

An extended model for iontophoretic enhancement of transdermal drug permeation under constant voltage is described based on the previously modified Nernst-Planck equation, which included the effect of convective solvent flow. This model resulted in an analytical expression for the enhancement factor as a function of applied voltage, convective flow velocity due to electroosmosis, ratio of lipid to aqueous pathway passive permeability, and weighted average net ionic valence of the permeant in the aqueous epidermis domain. The shift of pH in the epidermis compared to bulk caused by the electrical double layer at the lipid-aqueous domain interface was evaluated using the Poisson-Boltzmann equation. This was solved numerically for representative surface charge densities and yielded pH differences between bulk and epidermal aqueous domain between 0.05 and 0.4 pH units. The developed model was used to analyze the experimental enhancement of an amphoteric weak electrolyte measured in vitro using human cadaver epidermis and a voltage of 250 mV at different pH values. Parameter values characterizing the involved factors were determined that yielded the experimental enhancement factors and passive permeability coefficients at all pH values. The model provided a very good agreement between experimental and calculated enhancement and passive permeability. The deduced parameters showed (i) that the pH shift in the aqueous permeation pathway had a notable effect on the ionic valence and the partitioning of the drug in this domain for a high surface charge density and depending on the pK(a) and pI of the drug in relation to the bulk pH; (ii) the magnitude and the direction of convective transport due to electroosmosis typically reflected the density and sign, respectively, of surface charge of the tissue and its effect on enhancement was substantial for bulk pH values differing from the pI of epidermal tissue; (iii) the aqueous pathway predominantly determined passive permeability of the studied compound despite its measurable lipophilicity and therefore the lipid pathway did not notably affect enhancement. Hence, the proposed model can provide a good quantitative insight into the interplay between different phenomena and permeant properties influencing iontophoresis and can potentially be used as a predictive tool of the process.

Sphingosine 1-phosphate receptor 3 regulates recruitment of anti-inflammatory monocytes to microvessels during implant arteriogenesis

PubMed Central

Awojoodu, Anthony O.; Ogle, Molly E.; Sefcik, Lauren S.; Bowers, Daniel T.; Martin, Kyle; Brayman, Kenneth L.; Lynch, Kevin R.; Peirce-Cottler, Shayn M.; Botchwey, Edward

2013-01-01

Endothelial cells play significant roles in conditioning tissues after injury by the production and secretion of angiocrine factors. At least two distinct subsets of monocytes, CD45+CD11b+Gr1+Ly6C+ inflammatory and CD45+CD11b+Gr1−Ly6C− anti-inflammatory monocytes, respond differentially to these angiocrine factors and promote pathogen/debris clearance and arteriogenesis/tissue regeneration, respectively. We demonstrate here that local sphingosine 1-phosphate receptor 3 (S1P3) agonism recruits anti-inflammatory monocytes to remodeling vessels. Poly(lactic-co-glycolic acid) thin films were used to deliver FTY720, an S1P1/3 agonist, to inflamed and ischemic tissues, which resulted in a reduction in proinflammatory cytokine secretion and an increase in regenerative cytokine secretion. The altered balance of cytokine secretion results in preferential recruitment of anti-inflammatory monocytes from circulation. The chemotaxis of these cells, which express more S1P3 than inflammatory monocytes, toward SDF-1α was also enhanced with FTY720 treatment, but not in S1P3 knockout cells. FTY720 delivery enhanced arteriolar diameter expansion and increased length density of the local vasculature. This work establishes a role for S1P receptor signaling in the local conditioning of tissues by angiocrine factors that preferentially recruit regenerative monocytes that can enhance healing outcomes, tissue regeneration, and biomaterial implant functionality. PMID:23918395

Chronic stress sensitizes rats to pancreatitis induced by cerulein: role of TNF-α.

PubMed

Binker, Marcelo-G; Binker-Cosen, Andres-A; Richards, Daniel; Gaisano, Herbert-Y; de Cosen, Rodica-H; Cosen-Binker, Laura-I

2010-11-28

To investigate chronic stress as a susceptibility factor for developing pancreatitis, as well as tumor necrosis factor-α (TNF-α) as a putative sensitizer. Rat pancreatic acini were used to analyze the influence of TNF-α on submaximal (50 pmol/L) cholecystokinin (CCK) stimulation. Chronic restraint (4 h every day for 21 d) was used to evaluate the effects of submaximal (0.2 μg/kg per hour) cerulein stimulation on chronically stressed rats. In vitro exposure of pancreatic acini to TNF-α disorganized the actin cytoskeleton. This was further increased by TNF-α/CCK treatment, which additionally reduced amylase secretion, and increased trypsin and nuclear factor-κB activities in a protein-kinase-C δ and ε-dependent manner. TNF-α/CCK also enhanced caspases' activity and lactate dehydrogenase release, induced ATP loss, and augmented the ADP/ATP ratio. In vivo, rats under chronic restraint exhibited elevated serum and pancreatic TNF-α levels. Serum, pancreatic, and lung inflammatory parameters, as well as caspases'activity in pancreatic and lung tissue, were substantially enhanced in stressed/cerulein-treated rats, which also experienced tissues' ATP loss and greater ADP/ATP ratios. Histological examination revealed that stressed/cerulein-treated animals developed abundant pancreatic and lung edema, hemorrhage and leukocyte infiltrate, and pancreatic necrosis. Pancreatitis severity was greatly decreased by treating animals with an anti-TNF-α-antibody, which diminished all inflammatory parameters, histopathological scores, and apoptotic/necrotic markers in stressed/cerulein-treated rats. In rats, chronic stress increases susceptibility for developing pancreatitis, which involves TNF-α sensitization of pancreatic acinar cells to undergo injury by physiological cerulein stimulation.

The small molecule AUTEN-99 (autophagy enhancer-99) prevents the progression of neurodegenerative symptoms

PubMed Central

Kovács, Tibor; Billes, Viktor; Komlós, Marcell; Hotzi, Bernadette; Manzéger, Anna; Tarnóci, Anna; Papp, Diána; Szikszai, Fanni; Szinyákovics, Janka; Rácz, Ákos; Noszál, Béla; Veszelka, Szilvia; Walter, Fruzsina R.; Deli, Mária A.; Hackler, Laszlo; Alfoldi, Robert; Huzian, Orsolya; Puskas, Laszlo G.; Liliom, Hanna; Tárnok, Krisztián; Schlett, Katalin; Borsy, Adrienn; Welker, Ervin; Kovács, Attila L.; Pádár, Zsolt; Erdős, Attila; Legradi, Adam; Bjelik, Annamaria; Gulya, Károly; Gulyás, Balázs; Vellai, Tibor

2017-01-01

Autophagy functions as a main route for the degradation of superfluous and damaged constituents of the cytoplasm. Defects in autophagy are implicated in the development of various age-dependent degenerative disorders such as cancer, neurodegeneration and tissue atrophy, and in accelerated aging. To promote basal levels of the process in pathological settings, we previously screened a small molecule library for novel autophagy-enhancing factors that inhibit the myotubularin-related phosphatase MTMR14/Jumpy, a negative regulator of autophagic membrane formation. Here we identify AUTEN-99 (autophagy enhancer-99), which activates autophagy in cell cultures and animal models. AUTEN-99 appears to effectively penetrate through the blood-brain barrier, and impedes the progression of neurodegenerative symptoms in Drosophila models of Parkinson’s and Huntington’s diseases. Furthermore, the molecule increases the survival of isolated neurons under normal and oxidative stress-induced conditions. Thus, AUTEN-99 serves as a potent neuroprotective drug candidate for preventing and treating diverse neurodegenerative pathologies, and may promote healthy aging. PMID:28205624

The small molecule AUTEN-99 (autophagy enhancer-99) prevents the progression of neurodegenerative symptoms.

PubMed

Kovács, Tibor; Billes, Viktor; Komlós, Marcell; Hotzi, Bernadette; Manzéger, Anna; Tarnóci, Anna; Papp, Diána; Szikszai, Fanni; Szinyákovics, Janka; Rácz, Ákos; Noszál, Béla; Veszelka, Szilvia; Walter, Fruzsina R; Deli, Mária A; Hackler, Laszlo; Alfoldi, Robert; Huzian, Orsolya; Puskas, Laszlo G; Liliom, Hanna; Tárnok, Krisztián; Schlett, Katalin; Borsy, Adrienn; Welker, Ervin; Kovács, Attila L; Pádár, Zsolt; Erdős, Attila; Legradi, Adam; Bjelik, Annamaria; Gulya, Károly; Gulyás, Balázs; Vellai, Tibor

2017-02-16

Autophagy functions as a main route for the degradation of superfluous and damaged constituents of the cytoplasm. Defects in autophagy are implicated in the development of various age-dependent degenerative disorders such as cancer, neurodegeneration and tissue atrophy, and in accelerated aging. To promote basal levels of the process in pathological settings, we previously screened a small molecule library for novel autophagy-enhancing factors that inhibit the myotubularin-related phosphatase MTMR14/Jumpy, a negative regulator of autophagic membrane formation. Here we identify AUTEN-99 (autophagy enhancer-99), which activates autophagy in cell cultures and animal models. AUTEN-99 appears to effectively penetrate through the blood-brain barrier, and impedes the progression of neurodegenerative symptoms in Drosophila models of Parkinson's and Huntington's diseases. Furthermore, the molecule increases the survival of isolated neurons under normal and oxidative stress-induced conditions. Thus, AUTEN-99 serves as a potent neuroprotective drug candidate for preventing and treating diverse neurodegenerative pathologies, and may promote healthy aging.

Contact area affects frequency-dependent responses to vibration in the peripheral vascular and sensorineural systems.

PubMed

Krajnak, Kristine; Miller, G R; Waugh, Stacey

2018-01-01

Repetitive exposure to hand-transmitted vibration is associated with development of peripheral vascular and sensorineural dysfunctions. These disorders and symptoms associated with it are referred to as hand-arm vibration syndrome (HAVS). Although the symptoms of the disorder have been well characterized, the etiology and contribution of various exposure factors to development of the dysfunctions are not well understood. Previous studies performed using a rat-tail model of vibration demonstrated that vascular and peripheral nervous system adverse effects of vibration are frequency-dependent, with vibration frequencies at or near the resonant frequency producing the most severe injury. However, in these investigations, the amplitude of the exposed tissue was greater than amplitude typically noted in human fingers. To determine how contact with vibrating source and amplitude of the biodynamic response of the tissue affects the risk of injury occurring, this study compared the influence of frequency using different levels of restraint to assess how maintaining contact of the tail with vibrating source affects the transmission of vibration. Data demonstrated that for the most part, increasing the contact of the tail with the platform by restraining it with additional straps resulted in an enhancement in transmission of vibration signal and elevation in factors associated with vascular and peripheral nerve injury. In addition, there were also frequency-dependent effects, with exposure at 250 Hz generating greater effects than vibration at 62.5 Hz. These observations are consistent with studies in humans demonstrating that greater contact and exposure to frequencies near the resonant frequency pose the highest risk for generating peripheral vascular and sensorineural dysfunction.

IRF7 in the Australian black flying fox, Pteropus alecto: evidence for a unique expression pattern and functional conservation.

PubMed

Zhou, Peng; Cowled, Chris; Mansell, Ashley; Monaghan, Paul; Green, Diane; Wu, Lijun; Shi, Zhengli; Wang, Lin-Fa; Baker, Michelle L

2014-01-01

As the only flying mammal, bats harbor a number of emerging and re-emerging viruses, many of which cause severe diseases in humans and other mammals yet result in no clinical symptoms in bats. As the master regulator of the interferon (IFN)-dependent immune response, IFN regulatory factor 7 (IRF7) plays a central role in innate antiviral immunity. To explore the role of bat IRF7 in the regulation of the IFN response, we performed sequence and functional analysis of IRF7 from the pteropid bat, Pteropus alecto. Our results demonstrate that bat IRF7 retains the ability to bind to MyD88 and activate the IFN response despite unique changes in the MyD88 binding domain. We also demonstrate that bat IRF7 has a unique expression pattern across both immune and non-immune related tissues and is inducible by double-strand RNA. The broad tissue distribution of IRF7 may provide bats with an enhanced ability to rapidly activate the IFN response in a wider range of tissues compared to other mammals. The importance of IRF7 in antiviral activity against the bat reovirus, Pulau virus was confirmed by siRNA knockdown of IRF7 in bat cells resulting in enhanced viral replication. Our results highlight the importance of IRF7 in innate antiviral immunity in bats.

IRF7 in the Australian Black Flying Fox, Pteropus alecto: Evidence for a Unique Expression Pattern and Functional Conservation

PubMed Central

Zhou, Peng; Cowled, Chris; Mansell, Ashley; Monaghan, Paul; Green, Diane; Wu, Lijun; Shi, Zhengli; Wang, Lin-Fa; Baker, Michelle L.

2014-01-01

As the only flying mammal, bats harbor a number of emerging and re-emerging viruses, many of which cause severe diseases in humans and other mammals yet result in no clinical symptoms in bats. As the master regulator of the interferon (IFN)-dependent immune response, IFN regulatory factor 7 (IRF7) plays a central role in innate antiviral immunity. To explore the role of bat IRF7 in the regulation of the IFN response, we performed sequence and functional analysis of IRF7 from the pteropid bat, Pteropus alecto. Our results demonstrate that bat IRF7 retains the ability to bind to MyD88 and activate the IFN response despite unique changes in the MyD88 binding domain. We also demonstrate that bat IRF7 has a unique expression pattern across both immune and non-immune related tissues and is inducible by double-strand RNA. The broad tissue distribution of IRF7 may provide bats with an enhanced ability to rapidly activate the IFN response in a wider range of tissues compared to other mammals. The importance of IRF7 in antiviral activity against the bat reovirus, Pulau virus was confirmed by siRNA knockdown of IRF7 in bat cells resulting in enhanced viral replication. Our results highlight the importance of IRF7 in innate antiviral immunity in bats. PMID:25100081

Developing functional musculoskeletal tissues through hypoxia and lysyl oxidase-induced collagen cross-linking

PubMed Central

Makris, Eleftherios A.; Responte, Donald J.; Hu, Jerry C.; Athanasiou, Kyriacos A.

2014-01-01

The inability to recapitulate native tissue biomechanics, especially tensile properties, hinders progress in regenerative medicine. To address this problem, strategies have focused on enhancing collagen production. However, manipulating collagen cross-links, ubiquitous throughout all tissues and conferring mechanical integrity, has been underinvestigated. A series of studies examined the effects of lysyl oxidase (LOX), the enzyme responsible for the formation of collagen cross-links. Hypoxia-induced endogenous LOX was applied in multiple musculoskeletal tissues (i.e., cartilage, meniscus, tendons, ligaments). Results of these studies showed that both native and engineered tissues are enhanced by invoking a mechanism of hypoxia-induced pyridinoline (PYR) cross-links via intermediaries like LOX. Hypoxia was shown to enhance PYR cross-linking 1.4- to 6.4-fold and, concomitantly, to increase the tensile properties of collagen-rich tissues 1.3- to 2.2-fold. Direct administration of exogenous LOX was applied in native cartilage and neocartilage generated using a scaffold-free, self-assembling process of primary chondrocytes. Exogenous LOX was found to enhance native tissue tensile properties 1.9-fold. LOX concentration- and time-dependent increases in PYR content (∼16-fold compared with controls) and tensile properties (approximately fivefold compared with controls) of neocartilage were also detected, resulting in properties on par with native tissue. Finally, in vivo subcutaneous implantation of LOX-treated neocartilage in nude mice promoted further maturation of the neotissue, enhancing tensile and PYR content approximately threefold and 14-fold, respectively, compared with in vitro controls. Collectively, these results provide the first report, to our knowledge, of endogenous (hypoxia-induced) and exogenous LOX applications for promoting collagen cross-linking and improving the tensile properties of a spectrum of native and engineered tissues both in vitro and in vivo. PMID:25349395

A spectrophotometer-based diffusivity assay reveals that diffusion hindrance of small molecules in extracellular matrix gels used in 3D cultures is dominated by viscous effects.

PubMed

Galgoczy, Roland; Pastor, Isabel; Colom, Adai; Giménez, Alicia; Mas, Francesc; Alcaraz, Jordi

2014-08-01

The design of 3D culture studies remains challenging due to the limited understanding of extracellular matrix (ECM)-dependent hindered diffusion and the lack of simple diffusivity assays. To address these limitations, we set up a cost-effective diffusivity assay based on a Transwell plate and the spectrophotometer of a Microplate Reader, which are readily accessible to cell biology groups. The spectrophotometer-based assay was used to assess the apparent diffusivity D of FITC-dextrans with molecular weight (4-70kDa) spanning the physiological range of signaling factors in a panel of acellular ECM gels including Matrigel, fibrin and type I collagen. Despite their technical differences, D data exhibited ∼15% relative difference with respect to FRAP measurements. Our results revealed that diffusion hindrance of small particles is controlled by the enhanced viscosity of the ECM gel in conformance with the Stokes-Einstein equation rather than by geometrical factors. Moreover, we provided a strong rationale that the enhanced ECM viscosity is largely contributed to by unassembled ECM macromolecules. We also reported that gels with the lowest D exhibited diffusion hindrance closest to the large physiologic hindrance of brain tissue, which has a typical pore size much smaller than ECM gels. Conversely, sparse gels (≤1mg/ml), which are extensively used in 3D cultures, failed to reproduce the hindered diffusion of tissues, thereby supporting that dense (but not sparse) ECM gels are suitable tissue surrogates in terms of macromolecular transport. Finally, the consequences of reduced diffusivity in terms of optimizing the design of 3D culture experiments were addressed in detail. Copyright © 2014 Elsevier B.V. All rights reserved.

Epithelial atrophy in oral submucous fibrosis is mediated by copper (II) and arecoline of areca nut

PubMed Central

Khan, Imran; Pant, Ila; Narra, Sivakrishna; Radhesh, Rekha; Ranganathan, Kannan; Rao, Somanahalli Girish; Kondaiah, Paturu

2015-01-01

Exposure of oral cavity to areca nut is associated with several pathological conditions including oral submucous fibrosis (OSF). Histopathologically OSF is characterized by epithelial atrophy, chronic inflammation, juxtaepithelial hyalinization, leading to fibrosis of submucosal tissue and affects 0.5% of the population in the Indian subcontinent. As the molecular mechanisms leading to atrophied epithelium and fibrosis are poorly understood, we studied areca nut actions on human keratinocyte and gingival fibroblast cells. Areca nut water extract (ANW) was cytotoxic to epithelial cells and had a pro-proliferative effect on fibroblasts. This opposite effect of ANW on epithelial and fibroblast cells was intriguing but reflects the OSF histopathology such as epithelial atrophy and proliferation of fibroblasts. We demonstrate that the pro-proliferative effects of ANW on fibroblasts are dependent on insulin-like growth factor signalling while the cytotoxic effects on keratinocytes are dependent on the generation of reactive oxygen species. Treatment of keratinocytes with arecoline which is a component of ANW along with copper resulted in enhanced cytotoxicity which becomes comparable to IC50 of ANW. Furthermore, studies using cyclic voltammetry, mass spectrometry and plasmid cleavage assay suggested that the presence of arecoline increases oxidation reduction potential of copper leading to enhanced cleavage of DNA which could generate an apoptotic response. Terminal deoxynucleotidyl transferase dUTP Nick End Labeling assay and Ki-67 index of OSF tissue sections suggested epithelial apoptosis, which could be responsible for the atrophy of OSF epithelium. PMID:26248978

Quantitative contrast-enhanced ultrasound imaging: a review of sources of variability

PubMed Central

Tang, M.-X.; Mulvana, H.; Gauthier, T.; Lim, A. K. P.; Cosgrove, D. O.; Eckersley, R. J.; Stride, E.

2011-01-01

Ultrasound provides a valuable tool for medical diagnosis offering real-time imaging with excellent spatial resolution and low cost. The advent of microbubble contrast agents has provided the additional ability to obtain essential quantitative information relating to tissue vascularity, tissue perfusion and even endothelial wall function. This technique has shown great promise for diagnosis and monitoring in a wide range of clinical conditions such as cardiovascular diseases and cancer, with considerable potential benefits in terms of patient care. A key challenge of this technique, however, is the existence of significant variations in the imaging results, and the lack of understanding regarding their origin. The aim of this paper is to review the potential sources of variability in the quantification of tissue perfusion based on microbubble contrast-enhanced ultrasound images. These are divided into the following three categories: (i) factors relating to the scanner setting, which include transmission power, transmission focal depth, dynamic range, signal gain and transmission frequency, (ii) factors relating to the patient, which include body physical differences, physiological interaction of body with bubbles, propagation and attenuation through tissue, and tissue motion, and (iii) factors relating to the microbubbles, which include the type of bubbles and their stability, preparation and injection and dosage. It has been shown that the factors in all the three categories can significantly affect the imaging results and contribute to the variations observed. How these factors influence quantitative imaging is explained and possible methods for reducing such variations are discussed. PMID:22866229

Expansion and delivery of adipose-derived mesenchymal stem cells on three microcarriers for soft tissue regeneration.

PubMed

Zhou, Yalei; Yan, Zhiwei; Zhang, Hongmei; Lu, Wei; Liu, Shiyu; Huang, Xinhui; Luo, Hailang; Jin, Yan

2011-12-01

Cell/microcarrier combinations can be injected to repair tissue defects, but whether currently available microcarriers can be utilized to repair different tissue defects remains unknown. Here, we compared the suitability of fabricated micronized acellular dermal matrix (MADM), micronized small intestinal submucosa (MSIS), and gelatin microspheres as expansion and delivery scaffolds for adipose-derived mesenchymal stem cells (ADSCs). The results of MTS assay, scanning electron microscopy (SEM), and flow cytometry suggested that the three microcarriers all have good biocompatibility. Quantitative polymerase chain reaction revealed enhanced epidermal growth factor, vascular endothelial growth factor, basal fibroblast growth factor, and transforming growth factor-β expression levels after ADSCs had been cultured on MADM or MSIS for 5 days. After culturing ADSCs on microcarriers in osteogenic medium for 7 days, the expression levels of bone formation-related genes were enhanced. ADSC/microcarrier treatment accelerated wound closure. The ADSC/MADM and ADSC/MSIS combinations retained more of the original implant volume at 1 month postimplantation than ADSC/gelatin microspheres combination in soft-tissue augmentation studies. All implants displayed fibroblast and capillary vessel infiltrations; but ectopic bone formation did not occur, and the calvarial defect repair results were unfavorable. Our study demonstrates the potential utility of these microcarriers not only as a cell-culture substrate but also as a cell-transplantation vehicle for skin regeneration and soft-tissue reconstruction.

Nitric Oxide and ERK mediates regulation of cellular processes by Ecdysterone

DOE Office of Scientific and Technical Information (OSTI.GOV)

Omanakuttan, Athira; Bose, Chinchu; Pandurangan, Nanjan

The complex process of wound healing is a major problem associated with diabetes, venous or arterial disease, old age and infection. A wide range of pharmacological effects including anabolic, anti-diabetic and hepato-protective activities have been attributed to Ecdysterone. In earlier studies, Ecdysterone has been shown to modulate eNOS and iNOS expression in diabetic animals and activate osteogenic differentiation through the Extracellular-signal-Regulated Kinase (ERK) pathway in periodontal ligament stem cells. However, in the wound healing process, Ecdysterone has only been shown to enhance granulation tissue formation in rabbits. There have been no studies to date, which elucidate the molecular mechanism underlyingmore » the complex cellular process involved in wound healing. The present study, demonstrates a novel interaction between the phytosteroid Ecdysterone and Nitric Oxide Synthase (NOS), in an Epidermal Growth Factor Receptor (EGFR)-dependent manner, thereby promoting cell proliferation, cell spreading and cell migration. These observations were further supported by the 4-amino-5-methylamino- 2′ ,7′ -difluorofluorescein diacetate (DAF FM) fluorescence assay which indicated that Ecdysterone activates NOS resulting in increased Nitric Oxide (NO) production. Additionally, studies with inhibitors of both the EGFR and ERK, demonstrated that Ecdysterone activates NOS through modulation of EGFR and ERK. These results clearly demonstrate, for the first time, that Ecdysterone enhances Nitric Oxide production and modulates complex cellular processes by activating ERK1/2 through the EGF pathway. - Highlights: • Ecdysterone significantly enhances cell migration in a dose dependent manner. • Ecdysterone augments cell spreading during the initial phase of cell migration through actin cytoskeletal rearrangement. • Ecdysterone enhances cell proliferation in a nitric oxide dependent manner. • Ecdysterone enhances nitric oxide production via activation of EGFR and phosphorylation of ERK.« less

Regulation of the human ascorbate transporter SVCT2 exon 1b gene by zinc-finger transcription factors

PubMed Central

Qiao, Huan; May, James M.

2011-01-01

The sodium-dependent vitamin C transporter (SVCT) 2 is crucial for ascorbate uptake in metabolically active and specialized tissues. The present study focused on the gene regulation of the SVCT2 exon 1b, which is ubiquitously expressed in human and mouse tissues. Although the human SVCT2 exon 1b promoter doesn’t contain a classical TATA-box, we found that it does contain a functional initiator (Inr) that binds YY1 and interacts with upstream Sp1/Sp3 elements in the proximal promoter region. These elements in turn play a critical role in regulating YY1-mediated transcription of the exon 1b gene. Formation of YY1/Sp complexes on the promoter is required for its optional function. YY1 with Sp1 or Sp3 synergistically enhanced exon 1b promoter activity as well as the endogenous SVCT2 protein expression. Further, in addition to Sp1/Sp3 both EGR-1 and -2 were detected in the protein complexes that bound the three GC boxes bearing overlapping binding sites for EGR/WT1 and Sp1/3. The EGR family factors, WT1 and MAZ were found to differentially regulate exon 1b promoter activity. These results show that differential occupancy of transcription factors on the GC-rich consensus sequences in SVCT2 exon 1b promoter contributes to the regulation of cell and tissue expression of SVCT2. PMID:21335086

Fibroblast growth factor-2 stimulates adipogenic differentiation of human adipose-derived stem cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kakudo, Natsuko; Shimotsuma, Ayuko; Kusumoto, Kenji

2007-07-27

Adipose-derived stem cells (ASCs) have demonstrated a capacity for differentiating into a variety of lineages, including bone, cartilage, or fat, depending on the inducing stimuli and specific growth and factors. It is acknowledged that fibroblast growth factor-2 (FGF-2) promotes chondrogenic and inhibits osteogenic differentiation of ASCs, but thorough investigations of its effects on adipogenic differentiation are lacking. In this study, we demonstrate at the cellular and molecular levels the effect of FGF-2 on adipogenic differentiation of ASCs, as induced by an adipogenic hormonal cocktail consisting of 3-isobutyl-1-methylxanthine (IBMX), dexamethasone, insulin, and indomethacin. FGF-2 significantly enhances the adipogenic differentiation of humanmore » ASCs. Furthermore, in cultures receiving FGF-2 before adipogenic induction, mRNA expression of peroxisome proliferator-activated receptor {gamma}2 (PPAR{gamma}2), a key transcription factor in adipogenesis, was upregulated. The results of FGF-2 supplementation suggest the potential applications of FGF-2 and ASCs in adipose tissue regeneration.« less

Influence of Desorption Conditions on Analyte Sensitivity and Internal Energy in Discrete Tissue or Whole Body Imaging by IR-MALDESI

NASA Astrophysics Data System (ADS)

Rosen, Elias P.; Bokhart, Mark T.; Ghashghaei, H. Troy; Muddiman, David C.

2015-06-01

Analyte signal in a laser desorption/postionization scheme such as infrared matrix-assisted laser desorption electrospray ionization (IR-MALDESI) is strongly coupled to the degree of overlap between the desorbed plume of neutral material from a sample and an orthogonal electrospray. In this work, we systematically examine the effect of desorption conditions on IR-MALDESI response to pharmaceutical drugs and endogenous lipids in biological tissue using a design of experiments approach. Optimized desorption conditions have then been used to conduct an untargeted lipidomic analysis of whole body sagittal sections of neonate mouse. IR-MALDESI response to a wide range of lipid classes has been demonstrated, with enhanced lipid coverage received by varying the laser wavelength used for mass spectrometry imaging (MSI). Targeted MS2 imaging (MS2I) of an analyte, cocaine, deposited beneath whole body sections allowed determination of tissue-specific ion response factors, and CID fragments of cocaine were monitored to comment on wavelength-dependent internal energy deposition based on the "survival yield" method.

Hypoxia as a target for tissue specific gene therapy.

PubMed

Rhim, Taiyoun; Lee, Dong Yun; Lee, Minhyung

2013-12-10

Hypoxia is a hallmark of various ischemic diseases such as ischemic heart disease, ischemic limb, ischemic stroke, and solid tumors. Gene therapies for these diseases have been developed with various therapeutic genes including growth factors, anti-apoptotic genes, and toxins. However, non-specific expression of these therapeutic genes may induce dangerous side effects in the normal tissues. To avoid the side effects, gene expression should be tightly regulated in an oxygen concentration dependent manner. The hypoxia inducible promoters and enhancers have been evaluated as a transcriptional regulation tool for hypoxia inducible gene therapy. The hypoxia inducible UTRs were also used in gene therapy for spinal cord injury as a translational regulation strategy. In addition to transcriptional and translational regulations, post-translational regulation strategies have been developed using the HIF-1α ODD domain. Hypoxia inducible transcriptional, translational, and post-translational regulations are useful for tissue specific gene therapy of ischemic diseases. In this review, hypoxia inducible gene expression systems are discussed and their applications are introduced. Copyright © 2013 Elsevier B.V. All rights reserved.

Antigen Masking During Fixation and Embedding, Dissected

PubMed Central

Scalia, Carla Rossana; Boi, Giovanna; Bolognesi, Maddalena Maria; Riva, Lorella; Manzoni, Marco; DeSmedt, Linde; Bosisio, Francesca Maria; Ronchi, Susanna; Leone, Biagio Eugenio; Cattoretti, Giorgio

2016-01-01

Antigen masking in routinely processed tissue is a poorly understood process caused by multiple factors. We sought to dissect the effect on antigenicity of each step of processing by using frozen sections as proxies of the whole tissue. An equivalent extent of antigen masking occurs across variable fixation times at room temperature. Most antigens benefit from longer fixation times (>24 hr) for optimal detection after antigen retrieval (AR; for example, Ki-67, bcl-2, ER). The transfer to a graded alcohol series results in an enhanced staining effect, reproduced by treating the sections with detergents, possibly because of a better access of the polymeric immunohistochemical detection system to tissue structures. A second round of masking occurs upon entering the clearing agent, mostly at the paraffin embedding step. This may depend on the non-freezable water removal. AR fully reverses the masking due both to the fixation time and the paraffin embedding. AR itself destroys some epitopes which do not survive routine processing. Processed frozen sections are a tool to investigate fixation and processing requirements for antigens in routine specimens. PMID:27798289

Determination of the methylation status of MGMT in different regions within glioblastoma multiforme.

PubMed

Hamilton, Mark G; Roldán, Gloria; Magliocco, Anthony; McIntyre, John B; Parney, Ian; Easaw, Jacob C

2011-04-01

Epigenetic silencing of the MGMT gene through promoter methylation correlates with improved survival in Glioblastoma Multiforme (GBM) patients receiving concurrent chemoradiotherapy. Although the clinical benefit is primarily seen in patients with methylated MGMT promoter, some unmethylated patients also respond to Temozolomide. One possible explanation may be intratumoral heterogeneity. This study was designed to assess the methylation status of the MGMT promoter in different areas of GBM and determine if methylation status varied depending on the fixation technique (paraffin-embedding versus fresh frozen) used to store tissue. Using intraoperative navigation, biopsies were obtained from three distinct regions: the enhancing outer area, the non-enhancing inner core, and an area immediately outside the enhancing region. Only patients with GBM were included for evaluation and analysis. Samples taken from each area were divided with half stored by flash freezing and the other half stored using paraffin fixation. Methylation Specific-PCR (MS-PCR) was used for analysis of MGMT promoter methylation. Thirteen patients were included. Ten were male with a median age of 62 years. In each patient, samples were taken from the enhancing rim and the necrotic centre. However, it was not considered safe or feasible to obtain samples from the area immediately adjacent to the enhancing tumor rim in one case. All patients were homogeneous for methylation status throughout their tumor and tissue taken adjacent to it when frozen tissue was used. However, four patients had discrepancies in the MGMT promoter status between the frozen and paraffin-embedded blocks and one patient was not homogeneous within the tumor when paraffin-embedded tissue was used. MGMT promoter methylation status was homogeneous in all GBM tumors. Our observation that methylation status varied depending if the DNA was extracted from paraffin-embedded versus frozen tissue is concerning. Although the reason for this is unclear, we postulate that the timing from resection to fixation or the process of fixation itself may potentially alter methylation status in paraffin-embedded tumors.

BRD4 localization to lineage-specific enhancers is associated with a distinct transcription factor repertoire

PubMed Central

Najafova, Zeynab; Tirado-Magallanes, Roberto; Subramaniam, Malayannan; Hossan, Tareq; Schmidt, Geske; Nagarajan, Sankari; Baumgart, Simon J.; Mishra, Vivek Kumar; Bedi, Upasana; Hesse, Eric; Knapp, Stefan; Hawse, John R.; Johnsen, Steven A.

2017-01-01

Proper temporal epigenetic regulation of gene expression is essential for cell fate determination and tissue development. The Bromodomain-containing Protein-4 (BRD4) was previously shown to control the transcription of defined subsets of genes in various cell systems. In this study we examined the role of BRD4 in promoting lineage-specific gene expression and show that BRD4 is essential for osteoblast differentiation. Genome-wide analyses demonstrate that BRD4 is recruited to the transcriptional start site of differentiation-induced genes. Unexpectedly, while promoter-proximal BRD4 occupancy correlated with gene expression, genes which displayed moderate expression and promoter-proximal BRD4 occupancy were most highly regulated and sensitive to BRD4 inhibition. Therefore, we examined distal BRD4 occupancy and uncovered a specific co-localization of BRD4 with the transcription factors C/EBPb, TEAD1, FOSL2 and JUND at putative osteoblast-specific enhancers. These findings reveal the intricacies of lineage specification and provide new insight into the context-dependent functions of BRD4. PMID:27651452

BRD4 localization to lineage-specific enhancers is associated with a distinct transcription factor repertoire.

PubMed

Najafova, Zeynab; Tirado-Magallanes, Roberto; Subramaniam, Malayannan; Hossan, Tareq; Schmidt, Geske; Nagarajan, Sankari; Baumgart, Simon J; Mishra, Vivek Kumar; Bedi, Upasana; Hesse, Eric; Knapp, Stefan; Hawse, John R; Johnsen, Steven A

2017-01-09

Proper temporal epigenetic regulation of gene expression is essential for cell fate determination and tissue development. The Bromodomain-containing Protein-4 (BRD4) was previously shown to control the transcription of defined subsets of genes in various cell systems. In this study we examined the role of BRD4 in promoting lineage-specific gene expression and show that BRD4 is essential for osteoblast differentiation. Genome-wide analyses demonstrate that BRD4 is recruited to the transcriptional start site of differentiation-induced genes. Unexpectedly, while promoter-proximal BRD4 occupancy correlated with gene expression, genes which displayed moderate expression and promoter-proximal BRD4 occupancy were most highly regulated and sensitive to BRD4 inhibition. Therefore, we examined distal BRD4 occupancy and uncovered a specific co-localization of BRD4 with the transcription factors C/EBPb, TEAD1, FOSL2 and JUND at putative osteoblast-specific enhancers. These findings reveal the intricacies of lineage specification and provide new insight into the context-dependent functions of BRD4. © The Author(s) 2016. Published by Oxford University Press on behalf of Nucleic Acids Research.

Targeting the Tissue Factor-Factor VIIa Signaling Pathway to Enhance Activity of mTOR Inhibitors in the Treatment of Breast Cancer

DTIC Science & Technology

2009-09-01

Salzberg M, Ostapenko V, Illiger HJ, Behringer D, Bardy -Bouxin N, Boni J , Kong S, Cincotta M, and Moore L. Phase II study of temsirolimus (CCI-779), a ...factor interaction results in a tissue factor cytoplasmic domain- independent activation of protein synthesis, p70, and p90 S6 kinase phosphorylation. J ...The mTOR Pathway in Breast Cancer. J Mammary Gland Biol Neoplasia 2006; 11: 53-61. 23. Guba M, Yezhelyev, Eichhorn ME, Schmid G, Ischenko, Papyan A

ATF3 activates Stat3 phosphorylation through inhibition of p53 expression in skin cancer cells.

PubMed

Hao, Zhen-Feng; Ao, Jun-Hong; Zhang, Jie; Su, You-Ming; Yang, Rong-Ya

2013-01-01

ATF3, a member of the ATF/CREB family of transcription factors, has been found to be selectively induced by calcineurin/NFAT inhibition and to enhance keratinocyte tumor formation, although the precise role of ATF3 in human skin cancer and possible mechanisms remain unknown. In this study, clinical analysis of 30 skin cancer patients and 30 normal donors revealed that ATF3 was accumulated in skin cancer tissues. Functional assays demonstrated that ATF3 significantly promoted skin cancer cell proliferation. Mechanically, ATF3 activated Stat3 phosphorylation in skin cancer cell through regulation of p53 expression. Moreover, the promotion effect of ATF3 on skin cancer cell proliferation was dependent on the p53-Stat3 signaling cascade. Together, the results indicate that ATF3 might promote skin cancer cell proliferation and enhance skin keratinocyte tumor development through inhibiting p53 expression and then activating Stat3 phosphorylation.

Optimal temperature control of tissue embedded with gold nanoparticles for enhanced thermal therapy based on two-energy equation model.

PubMed

Wang, Shen-Ling; Qi, Hong; Ren, Ya-Tao; Chen, Qin; Ruan, Li-Ming

2018-05-01

Thermal therapy is a very promising method for cancer treatment, which can be combined with chemotherapy, radiotherapy and other programs for enhanced cancer treatment. In order to get a better effect of thermal therapy in clinical applications, optimal internal temperature distribution of the tissue embedded with gold nanoparticles (GNPs) for enhanced thermal therapy was investigated in present research. The Monte Carlo method was applied to calculate the heat generation of the tissue embedded with GNPs irradiated by continuous laser. To have a better insight into the physical problem of heat transfer in tissues, the two-energy equation was employed to calculate the temperature distribution of the tissue in the process of GNPs enhanced therapy. The Arrhenius equation was applied to evaluate the degree of permanent thermal damage. A parametric study was performed to investigate the influence factors on the tissue internal temperature distribution, such as incident light intensity, the GNPs volume fraction, the periodic heating and cooling time, and the incident light position. It was found that period heating and cooling strategy can effectively avoid overheating of skin surface and heat damage of healthy tissue. Lower GNPs volume fraction will be better for the heat source distribution. Furthermore, the ring heating strategy is superior to the central heating strategy in the treatment effect. All the analysis provides theoretical guidance for optimal temperature control of tissue embedded with GNP for enhanced thermal therapy. Copyright © 2018 Elsevier Ltd. All rights reserved.

Trophic factors from adipose tissue-derived multi-lineage progenitor cells promote cytodifferentiation of periodontal ligament cells.

PubMed

Sawada, Keigo; Takedachi, Masahide; Yamamoto, Satomi; Morimoto, Chiaki; Ozasa, Masao; Iwayama, Tomoaki; Lee, Chun Man; Okura, Hanayuki; Matsuyama, Akifumi; Kitamura, Masahiro; Murakami, Shinya

2015-08-14

Stem and progenitor cells are currently being investigated for their applicability in cell-based therapy for periodontal tissue regeneration. We recently demonstrated that the transplantation of adipose tissue-derived multi-lineage progenitor cells (ADMPCs) enhances periodontal tissue regeneration in beagle dogs. However, the molecular mechanisms by which transplanted ADMPCs induce periodontal tissue regeneration remain to be elucidated. In this study, trophic factors released by ADMPCs were examined for their paracrine effects on human periodontal ligament cell (HPDL) function. ADMPC conditioned medium (ADMPC-CM) up-regulated osteoblastic gene expression, alkaline phosphatase activity and calcified nodule formation in HPDLs, but did not significantly affect their proliferative response. ADMPCs secreted a number of growth factors, including insulin-like growth factor binding protein 6 (IGFBP6), hepatocyte growth factor and vascular endothelial growth factor. Among these, IGFBP6 was most highly expressed. Interestingly, the positive effects of ADMPC-CM on HPDL differentiation were significantly suppressed by transfecting ADMPCs with IGFBP6 siRNA. Our results suggest that ADMPCs transplanted into a defect in periodontal tissue release trophic factors that can stimulate the differentiation of HPDLs to mineralized tissue-forming cells, such as osteoblasts and cementoblasts. IGFBP6 may play crucial roles in ADMPC-induced periodontal regeneration. Copyright © 2015 Elsevier Inc. All rights reserved.

Low level laser therapy increases angiogenesis in a model of ischemic skin flap in rats mediated by VEGF, HIF-1α and MMP-2*

PubMed Central

Cury, Vivian; Moretti, Ana Iochabel Soares; Assis, Lívia; Bossini, Paulo; de Souza Crusca, Jaqueline; Neto, Carlos Benatti; Fangel, Renan; de Souza, Heraldo Possolo; Hamblin, Michael R; Parizotto, Nivaldo Antonio

2013-01-01

It is known that low level laser therapy is able to improve skin flap viability by increasing angiogenesis. However, the mechanism for new blood vessel formation is not completely understood. Here, we investigated the effects of 660 nm and 780 nm lasers at fluences of 30 and 40 J/cm2 on three important mediators activated during angiogenesis. Sixty male Wistar rats were used and randomly divided into five groups with twelve animals each. Groups were distributed as follows: skin flap surgery non-irradiated group as a control; skin flap surgery irradiated with 660 nm laser at a fluence of 30 or 40 J/cm2 and skin flap surgery irradiated with 780 nm laser at a fluence of 30 or 40 J/cm2. The random skin flap was performed measuring 10 × 4 cm, with a plastic sheet interposed between the flap and the donor site. Laser irradiation was performed on 24 points covering the flap and surrounding skin immediately after the surgery and for 7 consecutive days thereafter. Tissues were collected, and the number of vessels, angiogenesis markers (vascular endothelial growth factor, VEGF and hypoxia inducible factor, HIF-1α) and a tissue remodeling marker (matrix metalloproteinase, MMP-2) were analyzed. LLLT increased an angiogenesis, HIF-1α and VEGF expression and decrease MMP-2 activity. These phenomena were dependent on the fluences, and wavelengths used. In this study we showed that LLLT may improve the healing of skin flaps by enhancing the amount of new vessels formed in the tissue. Both 660 nm and 780 nm lasers were able to modulate VEGF secretion, MMP-2 activity and HIF-1α expression in a dose dependent manner. PMID:23831843

Radiation Exposure Enhances Hepatocyte Proliferation in Neonatal Mice but not in Adult Mice.

PubMed

Shang, Yi; Sawa, Yurika; Blyth, Benjamin J; Tsuruoka, Chizuru; Nogawa, Hiroyuki; Shimada, Yoshiya; Kakinuma, Shizuko

2017-08-01

There is a natural tendency to expect that irradiation of an infant organ prior to development-related expansion will result in a higher risk of developing cancer than that of fully-developed adult tissue, and this has generally been observed. However, if tissues also vary in their initial responses to radiation depending on age, the interplay between tissue- and age-dependent risk would potentially be quite complex. We have previously shown opposing age-dependent induction of apoptosis for the intestinal epithelium and hematopoietic cells in mice, but such data are not yet available for the liver. Here, we have examined markers of DNA damage, initiation of DNA damage responses, cell cycle arrest, apoptosis and proliferation, as well as gene expression, in the B6C3F1 mouse liver over the hours and days after irradiation of mice at 1 or 7 weeks of age. We found that induction and resolution of radiation-induced DNA damage is not accompanied by significant changes in these cellular end points in the adult liver, while in infant hepatocytes modest induction of p53 accumulation and p21-mediated cell cycle arrest in a small fraction of damaged cells was overshadowed by a further stimulation of proliferation over the relatively high levels already found in the neonatal liver. We observed distinct expression of genes that regulate cell division between the ages, which may contribute to the differential responses. These data suggest that the growth factor signaling environment of the infant liver may mediate radiation-induced proliferation and increased liver cancer risk after irradiation during early life.

Exogenous transforming growth factor-β1 enhances smooth muscle differentiation in embryonic mouse jejunal explants.

PubMed

Coletta, Riccardo; Roberts, Neil A; Randles, Michael J; Morabito, Antonino; Woolf, Adrian S

2017-01-13

An ex vivo experimental strategy that replicates in vivo intestinal development would in theory provide an accessible setting with which to study normal and dysmorphic gut biology. The current authors recently described a system in which mouse embryonic jejunal segments were explanted onto semipermeable platforms and fed with chemically defined serum-free media. Over 3 days in organ culture, explants formed villi and they began to undergo spontaneous peristalsis. As defined in the current study, the wall of the explanted gut failed to form a robust longitudinal smooth muscle (SM) layer as it would do in vivo over the same time period. Given the role of transforming growth factor β1 (TGFβ1) in SM differentiation in other organs, it was hypothesized that exogenous TGFβ1 would enhance SM differentiation in these explants. In vivo, TGFβ receptors I and II were both detected in embryonic longitudinal jejunal SM cells and, in organ culture, exogenous TGFβ1 induced robust differentiation of longitudinal SM. Microarray profiling showed that TGFβ1 increased SM specific transcripts in a dose dependent manner. TGFβ1 proteins were detected in amniotic fluid at a time when the intestine was physiologically herniated. By analogy with the requirement for exogenous TGFβ1 for SM differentiation in organ culture, the TGFβ1 protein that was demonstrated to be present in the amniotic fluid may enhance intestinal development when it is physiologically herniated in early gestation. Future studies of embryonic intestinal cultures should include TGFβ1 in the defined media to produce a more faithful model of in vivo muscle differentiation. Copyright © 2017 The Authors Journal of Tissue Engineering and Regenerative Medicine Published by John Wiley & Sons, Ltd. Copyright © 2017 The Authors Journal of Tissue Engineering and Regenerative Medicine Published by John Wiley & Sons, Ltd.

Priming Dental Pulp Stem Cells With Fibroblast Growth Factor-2 Increases Angiogenesis of Implanted Tissue-Engineered Constructs Through Hepatocyte Growth Factor and Vascular Endothelial Growth Factor Secretion.

PubMed

Gorin, Caroline; Rochefort, Gael Y; Bascetin, Rumeyza; Ying, Hanru; Lesieur, Julie; Sadoine, Jérémy; Beckouche, Nathan; Berndt, Sarah; Novais, Anita; Lesage, Matthieu; Hosten, Benoit; Vercellino, Laetitia; Merlet, Pascal; Le-Denmat, Dominique; Marchiol, Carmen; Letourneur, Didier; Nicoletti, Antonino; Vital, Sibylle Opsahl; Poliard, Anne; Salmon, Benjamin; Muller, Laurent; Chaussain, Catherine; Germain, Stéphane

2016-03-01

Tissue engineering strategies based on implanting cellularized biomaterials are promising therapeutic approaches for the reconstruction of large tissue defects. A major hurdle for the reliable establishment of such therapeutic approaches is the lack of rapid blood perfusion of the tissue construct to provide oxygen and nutrients. Numerous sources of mesenchymal stem cells (MSCs) displaying angiogenic potential have been characterized in the past years, including the adult dental pulp. Establishment of efficient strategies for improving angiogenesis in tissue constructs is nevertheless still an important challenge. Hypoxia was proposed as a priming treatment owing to its capacity to enhance the angiogenic potential of stem cells through vascular endothelial growth factor (VEGF) release. The present study aimed to characterize additional key factors regulating the angiogenic capacity of such MSCs, namely, dental pulp stem cells derived from deciduous teeth (SHED). We identified fibroblast growth factor-2 (FGF-2) as a potent inducer of the release of VEGF and hepatocyte growth factor (HGF) by SHED. We found that FGF-2 limited hypoxia-induced downregulation of HGF release. Using three-dimensional culture models of angiogenesis, we demonstrated that VEGF and HGF were both responsible for the high angiogenic potential of SHED through direct targeting of endothelial cells. In addition, FGF-2 treatment increased the fraction of Stro-1+/CD146+ progenitor cells. We then applied in vitro FGF-2 priming to SHED before encapsulation in hydrogels and in vivo subcutaneous implantation. Our results showed that FGF-2 priming is more efficient than hypoxia at increasing SHED-induced vascularization compared with nonprimed controls. Altogether, these data demonstrate that FGF-2 priming enhances the angiogenic potential of SHED through the secretion of both HGF and VEGF. ©AlphaMed Press.

Antitumor Activity of Emodin against Pancreatic Cancer Depends on Its Dual Role: Promotion of Apoptosis and Suppression of Angiogenesis

PubMed Central

Chen, Kang-Jie; Tong, Hong-Fei; Wang, Zhao-Hong; Ni, Zhong-Lin; Liu, Hai-Bin; Guo, Hong-Chun; Liu, Dian-Lei

2012-01-01

Background Emodin has been showed to induce apoptosis of pancreatic cancer cells and inhibit tumor growth in our previous studies. This study was designed to investigate whether emodin could inhibit the angiogenesis of pancreatic cancer tissues and its mechanism. Methodology/Principal Finding In accordance with our previous study, emodin inhibited pancreatic cancer cell growth, induced apoptosis, and enhanced the anti-tumor effect of gemcitabine on pancreatic caner cells in vitro and in vivo by inhibiting the activity of NF-κB. Here, for the first time, we demonstrated that emodin inhibited tumor angiogenesis in vitro and in implanted pancreatic cancer tissues, decreased the expression of angiogenesis-associated factors (NF-κB and its regulated factors VEGF, MMP-2, MMP-9, and eNOS), and reduced eNOS phosphorylation, as evidenced by both immunohistochemistry and western blot analysis of implanted tumors. In addition, we found that emodin had no effect on VEGFR expression in vivo. Conclusions/Significance Our results suggested that emodin has potential anti-tumor effect on pancreatic cancer via its dual role in the promotion of apoptosis and suppression of angiogenesis, probably through regulating the expression of NF-κB and NF-κB-regulated angiogenesis-associated factors. PMID:22876305

Chronic Ethanol consumption modulates growth factor release, mucosal cytokine production and microRNA expression in nonhuman primates

PubMed Central

Asquith, Mark; Pasala, Sumana; Engelmann, Flora; Haberthur, Kristen; Meyer, Christine; Park, Byung; Grant, Kathleen A.; Messaoudi, Ilhem

2013-01-01

BACKGROUND Chronic alcohol consumption has been associated with enhanced susceptibility to both systemic and mucosal infections. However, the exact mechanisms underlying this enhanced susceptibility remain incompletely understood. METHODS Using a nonhuman primate model of ethanol self-administration, we examined the impact of chronic alcohol exposure on immune homeostasis, cytokine and growth factor production in peripheral blood, lung and intestinal mucosa following twelve months of chronic ethanol exposure. RESULTS Ethanol exposure inhibited activation-induced production of growth factors HGF, G-CSF and VEGF by peripheral blood mononuclear cells (PBMC). Moreover, ethanol significantly reduced the frequency of colonic Th1 and Th17 cells in a dose-dependent manner. In contrast, we did not observe differences in lymphocyte frequency or soluble factor production in the lung of ethanol-consuming animals. To uncover mechanisms underlying reduced growth factor and Th1/Th17 cytokine production, we compared expression levels of microRNAs in PBMC and intestinal mucosa. Our analysis revealed ethanol-dependent upregulation of distinct microRNAs in affected tissues (miR-181a and miR-221 in PBMC; miR-155 in colon). Moreover, we were able to detect reduced expression of the transcription factors STAT3 and ARNT, which regulate expression of VEGF, G-CSF and HGF and contain targets for these microRNAs. To confirm and extend these observations, PBMC were transfected with either mimics or antagomirs of miR181 and 221and protein levels of the transcription factors and growth factors were determined. Transfection of microRNA mimics led to a reduction in both STAT-3/ARNT as well as VEGF/HGF/G-CSF levels. The opposite outcome was observed when microRNA antagomirs were transfected CONCLUSION Chronic ethanol consumption significantly disrupts both peripheral and mucosal immune homeostasis, and this dysregulation may be mediated by changes in microRNA expression. PMID:24329418

Experimental forensic and bioanthropological aspects of soft tissue taphonomy: 1. Factors influencing postmortem tissue desiccation rate.

PubMed

Aturaliya, S; Lukasewycz, A

1999-09-01

Euthanized rats' carcasses were exposed in an environmental chamber to multiple variables including: (1) position, (2) enveloping clothing, and (3) soil interment in an effort to determine the individual variables' effect on postmortem rate of body and visceral organ water loss. Results indicated that body water loss was enhanced by a horizontal position versus vertical, probably because of wider spread of bacteria- and enzyme-laden abdominal fluid secondary to diaphragm digestion with consequent greater tissue digestion and liquefaction. Clothing also accelerated the desiccation rate. Desiccation was about equally as effective by soil interment as by air exposure, though simulating windy conditions by tripling the air flow rate resulted in much more rapid desiccation in the air-exposed specimen. These studies suggest that the single most important factor influencing postmortem body water loss rate is the environment at the skin surface that acts to enhance or impair water removal from the skin surface and thus influences the water concentration gradient between the skin and underlying deeper tissues.

Regulation of the angiopoietin-2 gene by hCG in ovarian cancer cell line OVCAR-3.

PubMed

Pietrowski, D; Wiehle, P; Sator, M; Just, A; Keck, C

2010-05-01

Angiogenesis is a crucial step in growing tissues including many tumors. It is regulated by pro- and antiangiogenic factors including the family of angiopoietins and their corresponding receptors. In previous work we have shown that in human ovarian cells the expression of angiopoietin 2 (ANG2) is regulated by human chorionic gonadotropin (hCG). To better understand the mechanisms of hCG-dependent regulation of the ANG2-gene we have now investigated upstream regulatory active elements of the ANG2-promoter in the ovarian carcinoma cell line OVCAR-3. We cloned several ANG2-promoter-fragments of different lengths into a luciferase reporter-gene-vector and analyzed the corresponding ANG2 expression before and after hCG stimulation. We identified regions of the ANG2-promoter between 1 048 bp and 613 bp upstream of the transcriptional start site where hCG-dependent pathways promote a significant downregulation of gene expression. By sequence analysis of this area we found several potential binding sites for transcription factors that are involved in regulation of ANG2-expression, vascular development and ovarian function. These encompass the forkhead family transcription factors FOXC2 and FOXO1 as well as the CCAAT/enhancer binding protein family (C/EBP). In conclusion, we have demonstrated that the regulation of ANG2-expression in ovarian cancer cells is hCG-dependent and we suggest that forkhead transcription factor and C/EBP-dependent pathways are involved in the regulation of ANG2-expression in ovarian cancer cells. Georg Thieme Verlag KG Stuttgart-New York.

A comparative study of tissue factor and kaolin on blood coagulation assays using rotational thromboelastometry and thromboelastography.

PubMed

Peng, Henry T; Grodecki, Richard; Rizoli, Sandro; Shek, Pang N

2016-01-01

Rotational thromboelastometry (ROTEM) and thromboelastography (TEG) have been increasingly used to diagnose acute coagulopathy and guide blood transfusion. The tests are routinely performed using different triggering activators such as tissue factor and kaolin, which activate different pathways yielding different results. To optimize the global blood coagulation assays using ROTEM and TEG, we conducted a comparative study on the activation methods employing tissue factor and kaolin at different concentrations as well as standard reagents as recommended by the manufacturer of each device. Key parameter values were obtained at various assay conditions to evaluate and compare coagulation and fibrinolysis profiles of citrated whole blood collected from healthy volunteers. It was found that tissue factor reduced ROTEM clotting time and TEG R, and increased ROTEM clot formation time and TEG K in a concentration-dependent manner. In addition, tissue factor affected ROTEM alpha angle, and maximum clot firmness, especially in the absence of kaolin activation, whereas both ROTEM and TEG clot lysis (LI30, CL30, and LY30) remained unaffected. Moreover, kaolin reduced ROTEM clotting time and TEG R and K, but to a lesser extent than tissue factor, in-tem and ex-tem. Correlations in all corresponding parameters between ROTEM and TEG were observed, when the same activators were used in the assays compared with lesser correlations between standard kaolin TEG and ROTEM (INTEM/EXTEM). The two types of viscoelastic point-of-care devices provide different results, depending on the triggering reagent used to perform the assay. Optimal assay condition was obtained to reduce assay time and improve assay accuracy.

Connective tissue growth factor is expressed in bone marrow stromal cells and promotes interleukin-7-dependent B lymphopoiesis.

PubMed

Cheung, Laurence C; Strickland, Deborah H; Howlett, Meegan; Ford, Jette; Charles, Adrian K; Lyons, Karen M; Brigstock, David R; Goldschmeding, Roel; Cole, Catherine H; Alexander, Warren S; Kees, Ursula R

2014-07-01

Hematopoiesis occurs in a complex bone marrow microenvironment in which bone marrow stromal cells provide critical support to the process through direct cell contact and indirectly through the secretion of cytokines and growth factors. We report that connective tissue growth factor (Ctgf, also known as Ccn2) is highly expressed in murine bone marrow stromal cells. In contrast, connective tissue growth factor is barely detectable in unfractionated adult bone marrow cells. While connective tissue growth factor has been implicated in hematopoietic malignancies, and is known to play critical roles in skeletogenesis and regulation of bone marrow stromal cells, its role in hematopoiesis has not been described. Here we demonstrate that the absence of connective tissue growth factor in mice results in impaired hematopoiesis. Using a chimeric fetal liver transplantation model, we show that absence of connective tissue growth factor has an impact on B-cell development, in particular from pro-B to more mature stages, which is linked to a requirement for connective tissue growth factor in bone marrow stromal cells. Using in vitro culture systems, we demonstrate that connective tissue growth factor potentiates B-cell proliferation and promotes pro-B to pre-B differentiation in the presence of interleukin-7. This study provides a better understanding of the functions of connective tissue growth factor within the bone marrow, showing the dual regulatory role of the growth factor in skeletogenesis and in stage-specific B lymphopoiesis. Copyright© Ferrata Storti Foundation.

Connective tissue growth factor is expressed in bone marrow stromal cells and promotes interleukin-7-dependent B lymphopoiesis

PubMed Central

Cheung, Laurence C.; Strickland, Deborah H.; Howlett, Meegan; Ford, Jette; Charles, Adrian K.; Lyons, Karen M.; Brigstock, David R.; Goldschmeding, Roel; Cole, Catherine H.; Alexander, Warren S.; Kees, Ursula R.

2014-01-01

Hematopoiesis occurs in a complex bone marrow microenvironment in which bone marrow stromal cells provide critical support to the process through direct cell contact and indirectly through the secretion of cytokines and growth factors. We report that connective tissue growth factor (Ctgf, also known as Ccn2) is highly expressed in murine bone marrow stromal cells. In contrast, connective tissue growth factor is barely detectable in unfractionated adult bone marrow cells. While connective tissue growth factor has been implicated in hematopoietic malignancies, and is known to play critical roles in skeletogenesis and regulation of bone marrow stromal cells, its role in hematopoiesis has not been described. Here we demonstrate that the absence of connective tissue growth factor in mice results in impaired hematopoiesis. Using a chimeric fetal liver transplantation model, we show that absence of connective tissue growth factor has an impact on B-cell development, in particular from pro-B to more mature stages, which is linked to a requirement for connective tissue growth factor in bone marrow stromal cells. Using in vitro culture systems, we demonstrate that connective tissue growth factor potentiates B-cell proliferation and promotes pro-B to pre-B differentiation in the presence of interleukin-7. This study provides a better understanding of the functions of connective tissue growth factor within the bone marrow, showing the dual regulatory role of the growth factor in skeletogenesis and in stage-specific B lymphopoiesis. PMID:24727816

Dose of radiation enhancement, using silver nanoparticles in a human tissue equivalent gel dosimeter.

PubMed

Hassan, Muhammad; Waheed, Muhammad Mohsin; Anjum, Muhammad Naeem

2016-01-01

To quantify the radiation dose enhancement in a human tissue-equivalent polymer gel impregnated with silver nanoparticles. The case-control study was conducted at the Bahawalpur Institute of Nuclear Medicine and Oncology, Bahawalpur, Pakistan, in January 2014. Silver nanoparticles used in this study were prepared by wet chemical method. Polymer gel was prepared by known quantity of gelatine, methacrylic acid, ascorbic acid, copper sulphate pentahydrate, hydroquinone and water. Different concentrations of silver nanoparticles were added to the gel during its cooling process. The gel was cooled in six plastic vials of 50ml each. Two vials were used as a control sample while four vials were impregnated with silver nanoparticles. After 22 hours, the vials were irradiated with gamma rays by aCobalt-60 unit. Radiation enhancement was assessed by taking magnetic resonance images of the vials. The images were analysed using Image J software. The dose enhancement factor was 24.17% and 40.49% for 5Gy and 10Gy dose respectively. The dose enhancement factor for the gel impregnated with 0.10mM silver nanoparticles was 32.88% and 51.98% for 5Gy and 10Gy dose respectively. The impregnation of a tissue-equivalent gel with silver nanoparticles resulted in dose enhancement and this effect was magnified up to a certain level with the increase in concentration of silver nanoparticles.

Monocyte B7 and Sialyl Lewis X modulates the efficacy of IL-10 down-regulation of LPS-induced monocyte tissue factor in whole blood.

PubMed

Warnes, G; Biggerstaff, J P; Francis, J L

1998-07-01

Recent studies have investigated the use of anti-inflammatory cytokine, interleukin 10 (IL-10) to control the development of disseminated intravascular coagulation (DIC) in sepsis by down-regulation of monocyte tissue factor (MTF) induced by lipopolysaccharide (LPS) in the initial phase of the disease. In vitro and in vivo human studies have shown that a minimal (<1 h) delay in IL-10 treatment significantly reduces the cytokines ability to inhibit LPS-induced MTF expression and the end products of coagulation. In this whole blood in vitro study we investigated the role of lymphocyte and platelet interactions with monocytes to up-regulate MTF expression in the presence of IL-10 in the initial phase of exposure to LPS. Individual blockade of monocyte B7 or platelet P-selectin significantly (35%) reduced MTF expression (P<0.05). IL-10 showed a dose-dependent inhibition of LPS (0.1 microg/ml) induced MTF expression, with 56% inhibition at 1 ng/ml, maximizing at 5 ng/ml IL-10 (75%; P<0.05). Simultaneous exposure to LPS and IL-10 (1 ng/ml) or addition of IL-10 1 h after LPS, with individual B7 and P-selectin blockade significantly enhanced the inhibition of MTF expression by IL-10 (P<0.05). We conclude that the efficacy of IL-10 to control DIC could be enhanced by a simultaneous B7 and P-selectin blockade.

Mineral concentration dependent modulation of mechanical properties of bone-inspired bionanocomposite scaffold

NASA Astrophysics Data System (ADS)

Biswas, Abhijit; Ovaert, Timothy C.; Slaboch, Constance; Zhao, He; Bayer, Ilker S.; Biris, Alexandru S.; Wang, Tao

2011-07-01

We demonstrate tunable mechanical properties of bone-inspired bionanocomposite scaffolds while maintaining the required viscoelasticity. Mechanical properties such as hardness and elastic modulus of the bionanocomposite scaffolds were controlled by varying mineral concentrations of the bioscaffold. In particular, higher calcium and oxygen contents in the bioscaffold resulted in a significant enhancement in hardness and modulus of the bionanocomposite. Moreover, the phosphorous content appeared to be a determining factor in the hardness and mechanical properties of the bionanocomposites. These results open up the possibility of designing new engineered biocompatible nanoscaffolds with desired and tunable biomimetic functions and biomechanical properties with significant potential for advanced bone tissue engineering platforms and bone substitutes.

Nebivolol dilates human penile arteries and reverses erectile dysfunction in diabetic rats through enhancement of nitric oxide signaling.

PubMed

Angulo, Javier; Wright, Harold M; Cuevas, Pedro; González-Corrochano, Rocío; Fernández, Argentina; Cuevas, Begoña; La Fuente, José M; Gupta, Sandeep; Sáenz de Tejada, Iñigo

2010-08-01

Traditional beta-blockers have sometimes been associated with erectile dysfunction (ED). Nebivolol is a cardioselective β(1)-adrenoceptor antagonist that promotes vasodilation through a nitric oxide (NO)-dependent mechanism. We evaluated the effects of nebivolol on the NO/cyclic guanosine monophosphate (cGMP) signaling pathway, on erectile function and dysfunction, and in human penile vascular tissues. Erectile response to cavernosal nerve electrical stimulation in control and diabetes-induced ED rats were evaluated, along with serum nitrite/nitrate (NOx) concentration and plasma/tissue cGMP levels. Endothelium-dependent and sildenafil-induced relaxation of isolated human corpus cavernosum (HCC) and human penile resistance arteries (HPRA) were also determined. The effects of nebivolol on erectile function and dysfunction and on NO/cGMP-mediated responses. Treatment with nebivolol significantly potentiated erectile response in control rats, regardless of its effects on blood pressure. Nebivolol increased NOx and plasma cGMP by 3-fold and 2.75-fold, respectively, and significantly augmented the elevation of plasma cGMP produced by sildenafil. Nebivolol enhanced endothelium-dependent and sildenafil-induced relaxations of HCC tissue, and produced endothelium-dependent vasodilation of HPRA. Nebivolol, but not atenolol, significantly improved erectile response in diabetic rats (51.6%, 53.2%, and 87.1% of response at 3 Hz in nondiabetic rats, for vehicle-treated, atenolol-treated, and nebivolol-treated diabetic rats, respectively); after sildenafil administration, ED was completely reversed in nebivolol-treated diabetic rats (69.6% and 112% for diabetic rats treated with sildenafil and nebivolol plus sildenafil, respectively). Accordingly, nebivolol restored systemic NOx levels and cGMP content in penile tissue from these animals. Nebivolol in vivo activated the NO/cGMP pathway, enhanced erectile response and reversed ED in diabetic rats. Moreover, nebivolol in vitro potentiated NO/cGMP-mediated relaxation of human erectile tissues. These effects may account for the low incidence of ED in nebivolol-treated hypertensive patients. Nebivolol therefore may have utility in the treatment of ED, particularly ED associated with diabetes. © 2010 International Society for Sexual Medicine.

Controlled Release Strategies for Bone, Cartilage, and Osteochondral Engineering—Part I: Recapitulation of Native Tissue Healing and Variables for the Design of Delivery Systems

PubMed Central

Santo, Vítor E.; Mano, João F.; Reis, Rui L.

2013-01-01

The potential of growth factors to stimulate tissue healing through the enhancement of cell proliferation, migration, and differentiation is undeniable. However, critical parameters on the design of adequate carriers, such as uncontrolled spatiotemporal presence of bioactive factors, inadequate release profiles, and supraphysiological dosages of growth factors, have impaired the translation of these systems onto clinical practice. This review describes the healing cascades for bone, cartilage, and osteochondral interface, highlighting the role of specific growth factors for triggering the reactions leading to tissue regeneration. Critical criteria on the design of carriers for controlled release of bioactive factors are also reported, focusing on the need to provide a spatiotemporal control over the delivery and presentation of these molecules. PMID:23268651

Color Swapping to Enhance Breast Cancer Digital Images Qualities Using Stain Normalization

NASA Astrophysics Data System (ADS)

Muhimmah, Izzati; Puspasari Wijaya, Dhina; Indrayanti

2017-03-01

Histopathology is the disease diagnosis by means of the visual examination of tissues under the microscope. The virtually transparent tissue sections were prepared using a number of colored histochemical stains bound selectively to the cellular components. A variation of colors comes to be a problem in histopathology based upon the microscope lighting for the range of factors. This research aimed to investigate an image enhancement by applying a nonlinear mapping approach to stain normalization and histogram equalization for contrast enhancement. Validation was carried out in 59 datasets with 96.6% accordance and expert justification.

Evaluation of the Effects of Platelet-Rich Plasma (PRP) Therapy Involved in the Healing of Sports-Related Soft Tissue Injuries

PubMed Central

Middleton, Kellie K.; Barro, Victor; Muller, Bart; Terada, Satosha; Fu, Freddie H.

2012-01-01

Abstract Musculoskeletal injuries are the most common cause of severe long-term pain and physical disability, and affect hundreds of millions of people around the world. One of the most popular methods used to biologically enhance healing in the fields of orthopaedic surgery and sports medicine includes the use of autologous blood products, namely, platelet rich plasma (PRP). PRP is an autologous concentration of human platelets to supra-physiologic levels. At baseline levels, platelets function as a natural reservoir for growth factors including platelet-derived growth factor (PDGF), epidermal growth factor (EGF), transforming growth factor-beta 1 (TGF-β1), vascular endothelial growth factor (VEGF), basic fibroblast growth factor (FGF), hepatocyte growth factor (HGF), and insulin-like growth factor (IGF-I). PRP is commonly used in orthopaedic practice to augment healing in sports-related injuries of skeletal muscle, tendons, and ligaments. Despite its pervasive use, the clinical efficacy of PrP therapy and varying mechanisms of action have yet to be established. Basic science research has revealed that PRP exerts is effects through many downstream events secondary to release of growth factors and other bioactive factors from its alpha granules. These effects may vary depending on the location of injury and the concentration of important growth factors involved in various soft tissue healing responses. This review focuses on the effects of PrP and its associated bioactive factors as elucidated in basic science research. Current findings in PRP basic science research, which have shed light on its proposed mechanisms of action, have opened doors for future areas of PrP research. PMID:23576936

Elevated plasma factor VIII enhances venous thrombus formation in rabbits: contribution of factor XI, von Willebrand factor and tissue factor.

PubMed

Sugita, Chihiro; Yamashita, Atsushi; Matsuura, Yunosuke; Iwakiri, Takashi; Okuyama, Nozomi; Matsuda, Shuntaro; Matsumoto, Tomoko; Inoue, Osamu; Harada, Aya; Kitazawa, Takehisa; Hattori, Kunihiro; Shima, Midori; Asada, Yujiro

2013-07-01

Elevated plasma levels of factor VIII (FVIII) are associated with increased risk of deep venous thrombosis. The aim of this study is to elucidate how elevated FVIII levels affect venous thrombus formation and propagation in vivo. We examined rabbit plasma FVIII activity, plasma thrombin generation, whole blood coagulation, platelet aggregation and venous wall thrombogenicity before and one hour after an intravenous infusion of recombinant human FVIII (rFVIII). Venous thrombus induced by the endothelial denudation of rabbit jugular veins was histologically assessed. Thrombus propagation was evaluated as indocyanine green fluorescence intensity. Argatroban, a thrombin inhibitor, and neutralised antibodies for tissue factor (TF), factor XI (FXI), and von Willebrand factor (VWF) were infused before or after thrombus induction to investigate their effects on venous thrombus formation or propagation. Recombinant FVIII (100 IU/kg) increased rabbit plasma FVIII activity two-fold and significantly enhanced whole blood coagulation and total plasma thrombin generation, but did not affect initial thrombin generation time, platelet aggregation and venous wall thrombogenicity. The rFVIII infusion also increased the size of venous thrombus 1 hour after thrombus induction. Argatroban and the antibodies for TF, FXI or VWF inhibited such enhanced thrombus formation and all except TF suppressed thrombus propagation. In conclusion, elevated plasma FVIII levels enhance venous thrombus formation and propagation. Excess thrombin generation by FXI and VWF-mediated FVIII recruitment appear to contribute to the growth of FVIII-driven venous thrombus.

Diabetes impairs endothelium-dependent relaxation of human penile vascular tissues mediated by NO and EDHF.

PubMed

Angulo, Javier; Cuevas, Pedro; Fernández, Argentina; Gabancho, Sonia; Allona, Antonio; Martín-Morales, Antonio; Moncada, Ignacio; Videla, Sebastián; Sáenz de Tejada, Iñigo

2003-12-26

Standard treatments for erectile dysfunction (ED) (i.e., PDE5 inhibitors) are less effective in diabetic patients for unknown reasons. Endothelium-dependent relaxation (EDR) of human corpus cavernosum (HCC) depends on nitric oxide (NO), while in human penile resistance arteries (HPRA) endothelium-derived hyperpolarizing factor (EDHF) and NO participate. Here we show that diabetes significantly reduced EDR induced by acetylcholine (ACh) in HCC and HPRA. Relaxation attributed to EDHF was also impaired in HPRA from diabetic patients. The PDE5 inhibitor, sildenafil (10nM), reversed diabetes-induced endothelial dysfunction in HCC, but not in HPRA. Calcium dobesilate (DOBE; 10 microM) fully reversed diabetes-induced endothelial dysfunction in HPRA by specifically potentiating the EDHF-mediated component of EDR. Impairment by diabetes of NO and EDHF-dependent responses precluded the complete recovery of endothelial function in HPRA by sildenafil. This could explain the poor clinical response to PDE5 inhibitors of diabetic men with ED and suggests that a pharmacological approach that combines enhancement of NO/cGMP and EDHF pathways could be necessary to treat ED in many diabetic men.

Local application of periodontal ligament stromal cells promotes soft tissue regeneration.

PubMed

Baik, H S; Park, J; Lee, K J; Chung, C

2014-09-01

To test the potential stimulatory effect of local application of periodontal ligament (PDL) stromal cells on soft tissue regeneration. Fluorescently labeled PDL cells outgrown from extracted human premolars or phosphate-buffered saline were locally injected to the cutaneous wounds created on mice. Soft tissue regeneration was evaluated for 14 days using photographs and histomorphometry. PDL cell engraftment was tracked with confocal microscopy. To detect the paracrine effect of the PDL cells on soft tissue regeneration, PDL cell-conditioned medium (CM) was evaluated for the concentration of secretory factors, transforming growth factor-beta 1 (TGFβ1). The effect of PDL CM on the proliferation and migration of dermal fibroblast and keratinocyte was tested using MTT assay and migration assay. The application of PDL cells significantly promoted soft tissue regeneration compared with the application of PBS. Self-replicating PDL cells were engrafted into the hair follicles of the host tissue. Dermal fibroblast proliferation and keratinocyte migration were significantly enhanced by the treatment with PDL CM. Physiologically significant amount of TGFβ1 was secreted from PDL cells into the CM. Local injection of PDL cells promoted soft tissue regeneration in part by the enhancement of fibroblast proliferation and keratinocyte migration through a paracrine mechanism. © 2013 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Development of a 3D co-culture model using human stem ...

EPA Pesticide Factsheets

Morphogenetic tissue fusion is a critical and complex event in embryonic development and failure of this event leads to birth defects, such as cleft palate. Palatal fusion requires adhesion and subsequent dissolution of the medial epithelial layer of the mesenchymal palatal shelves, and is regulated by the growth factors EGF and TGFβ, and others, although the complete regulatory mechanism is not understood. Three dimensional (3D) organotypic models allow us to mimic the native architecture of human tissue to facilitate the study of tissue dynamics and their responses to developmental toxicants. Our goal was to develop and characterize a spheroidal model of palatal fusion to investigate the mechanisms regulating fusion with exposure to growth factors and chemicals in the ToxCast program known to disrupt this event. We present a spheroidal model using human umbilical-derived mesenchymal stem cells (hMSC) spheroid cores cultured for 13 days and then coated with MaxGel™ basement membrane and a layer of human progenitor epithelial keratinocytes (hPEK) (hMSC+hPEK spheroids). We characterized the growth, differentiation, proliferation and fusion activity of the model. Spheroid diameter was dependent on hMSC seeding density, size of the seeding wells, time in culture, and type of medium. hMSC spheroid growth was enhanced with osteogenic differentiation medium. Alkaline phosphatase activity in the hMSC spheroid, indicating osteogenic differentiation, increased in inte

PRRX2 as a novel TGF-β-induced factor enhances invasion and migration in mammary epithelial cell and correlates with poor prognosis in breast cancer.

PubMed

Juang, Yu-Lin; Jeng, Yung-Ming; Chen, Chi-Long; Lien, Huang-Chun

2016-12-01

TGF-β and cancer progression share a multifaceted relationship. Despite the knowledge of TGF-β biology in the development of cancer, several factors that mediate the cancer-promoting role of TGF-β continue to be identified. This study aimed to identify and characterise novel factors potentially related to TGF-β-mediated tumour aggression in breast cells. We treated the human mammary epithelial cell line MCF10A with TGF-β and identified TGF-β-dependent upregulation of PRRX2, the gene encoding paired-related homeobox 2 transcription factor. Overexpression of PRRX2 enhanced migration, invasion and anchorage-independent growth of MCF10A cells and induced partial epithelial mesenchymal transition (EMT), as determined by partial fibroblastoid morphology of cells, upregulation of EMT markers and partially disrupted acinar structure in a three-dimensional culture. We further identified PLAT, the gene encoding tissue-type plasminogen activator (tPA), as the highest differentially expressed gene in PRRX2-overexpressing MCF10A cells, and demonstrated direct binding and transactivation of the PLAT promoter by PRRX2. Furthermore, PLAT knockdown inhibited PRRX2-mediated enhanced migration and invasion, suggesting that tPA may mediate PRRX2-induced migration and invasion. Finally, the significant correlation of PRRX2 expression with poor survival in 118 primary breast tumour samples (P = 0.027) and the increased PRRX2 expression in metaplastic breast carcinoma samples, which is pathogenetically related to EMT, validated the biological importance of PRRX2-enhanced migration and invasion and PRRX2-induced EMT. Thus, our data suggest that upregulation of PRRX2 may be a mechanism contributing to TGF-β-induced invasion and EMT in breast cancer. © 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.

Unexpected hypoxia-dependent erythropoietin secretion during experimental conditions not affecting tissue oxygen supply/demand ratio.

PubMed

Bozzini, C E; Barceló, A C; Conti, M I; Martínez, M P; Lezón, C E; Bozzini, C; Alippi, R M

1997-02-01

Although a great deal of evidence supports the hypothesis that plasma erythropoietin (EPO) levels of mammals are related to the oxygen supply to the tissues relative to their oxygen needs, several observation millitate against its inherent simplicity. This study presents our results obtained from in vivo experiments that suggest that hypoxia-dependent EPO production can be altered by conditions which apparently do not modify the tissue oxygen supply/demand ratio. Hypoxia-dependent EPO production rate (EPO-PR), derived from plasma EPO titers and plasma EPO half-lives, were estimated in both transfused-polycythemic and normocythemic mouse models subjected to different treatments. From calculations of the O2 carrying capacity of blood and body O2 consumption, it was assumed that the tissue supply/demand ratios were similar in both experimental and control mice of the same model at the time of induction of EPO production. The following observations were worth noting: (1) EPO-PRs in transfused polycythemic mice whose erythropoietic rates were stimulated by intermittent exposure to hypobaria (0.5 atm, 18 hr/day x 3 weeks), phenylhydrazine administration (40 mg/kg at weekly intervals x 3 weeks) or repeated rh-EPO injections (1500 U/kg 3 times a week x 3 weeks) before transfusion were more than five times high than in comparabily polycythemic mice whose erythropoietic rates were not stimulated previously; and (2) EPO-PR in response to hypobaric hypoxia was 2.08 times normal in normocythemic mice with cyclophosphamide (100 mg/kg) induced depression of erythropoiesis, and 0.33 times normal in normocythemic mice with rh-EPO (400 U/kg x 2) induced enhancement of erythropoiesis. Although the results obtained in polycythemic mice are difficult to explain, those from normocythemic mice suggest the existence of a feedback mechanism between EPO-responsive cells and EPO-producing cells. Both demonstrate the existence of experimental conditions in which modulation of the hypoxia-dependent expression of the EPO gene appears to occur. This modulation would be dependent on factors other than oxygen.

Hypoxic preconditioning potentiates the trophic effects of mesenchymal stem cells on co-cultured human primary hepatocytes.

PubMed

Qin, Harry H; Filippi, Céline; Sun, Song; Lehec, Sharon; Dhawan, Anil; Hughes, Robin D

2015-12-01

Mesenchymal stem/stromal cells (MSCs) improve the metabolic function of co-cultured hepatocytes. The present study aimed to further enhance the trophic effects of co-culture with hepatocytes using hypoxic preconditioning (HPc) of the MSCs and also to investigate the underlying molecular mechanisms involved. Human adipose tissue-derived MSCs were subjected to hypoxia (2 % O2; HPc) or normoxia (20 % O2) for 24 h and then co-cultured with isolated human hepatocytes. Assays of metabolic function and apoptosis were performed to investigate the hepatotrophic and anti-apoptotic effects of co-culture. Indirect co-cultures and co-culture with MSC-conditioned medium investigated the role of paracrine factors in the hepatotrophic effects of co-culture. Reactive oxygen species (ROS) activity was antagonised with N-acetylcysteine to investigate whether HPc potentiated the effects of MSCs by intracellular ROS-dependent mechanisms. Tumour necrosis factor (TNF)-α, transforming growth factor (TGF)-β1, and extracellular collagen production was determined and CASP9 and BAX/BCL-2 signalling pathways analysed to investigate the role of soluble factors, extracellular matrix deposition, and apoptosis-associated gene signalling in the effects of co-culture. HPc potentiated the hepatotrophic and anti-apoptotic effects of co-culture by ROS-dependent mechanisms. There was increased MSC TGF-β1 production, and enhanced MSC deposition of extracellular collagen, with reduced synthesis of TNF-α, as well as a downregulation of the expression of pro-apoptotic CASP9, BAX, BID and BLK genes and upregulated expression of anti-apoptotic BCL-2 in hepatocytes. HPc potentiated the trophic and anti-apoptotic effects of MSCs on hepatocytes via mechanisms including intracellular ROS, autocrine TGF-β, extracellular collagen and caspase and BAX/BCL-2 signalling pathways.

Chitosan-aluminum monostearate composite sponge dressing containing asiaticoside for wound healing and angiogenesis promotion in chronic wound.

PubMed

Phaechamud, Thawatchai; Yodkhum, Kotchamon; Charoenteeraboon, Juree; Tabata, Yasuhiko

2015-05-01

There are many factors that delay healing in chronic wounds including lowering level of growth factors and increasing exudate level comprising high amount of tissue destructive enzymes. Asiaticoside possesses interesting wound healing and angiogenic activities that are employed to stimulate tissue regeneration in wound healing application. This study attempted to develop chitosan-aluminum monostearate (Alst) composite sponge containing asiaticoside for use as an absorbent medical dressing in chronic wound. N-methyl-2-pyrrolidone (NMP) was used to enhance homogeneity of asiaticoside in the polymer composite matrix. The sponge dressings were prepared by lyophilization and dehydrothermal treatment (DHT). Functional group interaction, crystallinity, and morphology of the prepared sponges were investigated using FT-IR, PXRD, and SEM, respectively. Physicochemical properties, porosity, hydrophilic/hydrophobic properties and mechanical property, were evaluated. Wound dressing properties, water vapor transmission rate (WVTR), fluid absorbency, oxygen permeation (OP), and bio-adhesive property, were investigated. In vitro asiaticoside release study was conducted using immersion method. Cytotoxicity was studied in normal human dermal fibroblast (NHDF) and normal human epidermal keratinocyte (NHEK). Angiogenic activity of asiaticoside was evaluated using chick-chorioallantoic membrane (CAM) assay. FT-IR and PXRD results revealed the amidation after DHT to enhance the crystallinity of the prepared sponges. The prepared sponges had high porosity comprising high Alst-loaded amount that exhibited more compact structure. Alst enhanced hydrophobicity therefore it reduced the fluid absorption and WVTR together with bio-adhesion of the prepared sponge dressings. Porosity of all sponges was more than 85% therefore resulting in their high OP. Enhancing hydrophobicity of the material by Alst and more homogeneity caused by NMP eventually retarded the asiaticoside release for 7 days. The sponge extractions were non-toxic to the cells moreover they promoted NHDF and NHEK cell proliferation. Asiaticoside and asiaticoside-contained dressings exhibited dose-dependent angiogenic activity in CAM model. Copyright © 2015 Elsevier B.V. All rights reserved.

Enhanced migration of murine fibroblast-like 3T3-L1 preadipocytes on type I collagen-coated dish is reversed by silibinin treatment.

PubMed

Liu, Xiaoling; Xu, Qian; Liu, Weiwei; Yao, Guodong; Zhao, Yeli; Xu, Fanxing; Hayashi, Toshihiko; Fujisaki, Hitomi; Hattori, Shunji; Tashiro, Shin-Ichi; Onodera, Satoshi; Yamato, Masayuki; Ikejima, Takashi

2018-04-01

Migration of fibroblast-like preadipocytes is important for the development of adipose tissue, whereas excessive migration is often responsible for impaired adipose tissue related with obesity and fibrotic diseases. Type I collagen (collagen I) is the most abundant component of extracellular matrix and has been shown to regulate fibroblast migration in vitro, but its role in adipose tissue is not known. Silibinin is a bioactive natural flavonoid with antioxidant and antimetastasis activities. In this study, we found that type I collagen coating promoted the proliferation and migration of murine 3T3-L1 preadipocytes in a dose-dependent manner, implying that collagen I could be an extracellular signal. Regarding the mechanisms of collagen I-stimulated 3T3-L1 migration, we found that NF-κB p65 is activated, including the increased nuclear translocation of NF-κB p65 as well as the upregulation of NF-κB p65 phosphorylation and acetylation, accompanied by the increased expressions of proinflammatory factors and the generation of reactive oxygen species (ROS). Reduction of collagen I-enhanced migration of cells by treatment with silibinin was associated with suppression of NF-κB p65 activity and ROS generation, and negatively correlated with the increasing sirt1 expression. Taken together, the enhanced migration of 3T3-L1 cells induced on collagen I-coated dish is mediated by the activation of NF-κB p65 function and ROS generation that can be alleviated with silibinin by upregulation of sirt1, leading to the repression of NF-κB p65 function and ROS generation.

Endothelial connexin 32 regulates tissue factor expression induced by inflammatory stimulation and direct cell-cell interaction with activated cells.

PubMed

Okamoto, Takayuki; Akita, Nobuyuki; Hayashi, Tatsuya; Shimaoka, Motomu; Suzuki, Koji

2014-10-01

Endothelial cell (EC) interacts with adjacent EC through gap junction, and abnormal expression or function of Cxs is associated with cardiovascular diseases. In patients with endothelial dysfunction, the up-regulation of tissue factor (TF) expression promotes the pathogenic activation of blood coagulation, however the relationship between gap junctions and TF expression in ECs remains uncharacterized. ECs express the gap junction (GJ) proteins connexin32 (Cx32), Cx37, Cx40 and Cx43. We investigated the role of endothelial gap junctions, particularly Cx32, in modulating TF expression during vascular inflammation. Human umbilical vein endothelial cells (HUVECs) were stimulated with tumor necrosis factor-α (TNF-α) and TF activity was assessed in the presence of GJ blockers and an inhibitory anti-Cx32 monoclonal antibody. Treatment with GJ blockers and anti-Cx32 monoclonal antibody enhanced the TNF-α-induced TF activity and mRNA expression in HUVECs. TNF-α-activated effector HUVECs or mouse MS-1 cells were co-cultured with non-stimulated acceptor HUVECs and TF expression in acceptor HUVECs was detected. Effector EC induced TF expression in adjacent acceptor HUVECs through direct cell-cell interaction. Cell-cell interaction induced TF expression was reduced by anti-intercellular adhesion molecule-1 (ICAM1) monoclonal antibody. Soluble ICAM1-Fc fusion protein promotes TF expression. GJ blockers and anti-Cx32 monoclonal antibody enhanced TF expression induced by cell-cell interaction and ICAM1-Fc treatment. Blockade of endothelial Cx32 increased TF expression induced by TNF-α stimulation and cell-cell interaction which was at least partly dependent upon ICAM1. These results suggest that direct Cx32-mediated interaction modulates TF expression in ECs during vascular inflammation. Copyright © 2014 Elsevier Ireland Ltd. All rights reserved.

Digital holography of intracellular dynamics to probe tissue physiology.

PubMed

Merrill, Daniel; An, Ran; Turek, John; Nolte, David D

2015-01-01

Digital holography provides improved capabilities for imaging through dense tissue. Using a short-coherence source, the digital hologram recorded from backscattered light performs laser ranging that maintains fidelity of information acquired from depths much greater than possible by traditional imaging techniques. Biodynamic imaging (BDI) is a developing technology for live-tissue imaging of up to a millimeter in depth that uses the hologram intensity fluctuations as label-free image contrast and can study tissue behavior in native microenvironments. In this paper BDI is used to investigate the change in adhesion-dependent tissue response in 3D cultures. The results show that increasing density of cellular adhesions slows motion inside tissue and alters the response to cytoskeletal drugs. A clear signature of membrane fluctuations was observed in mid-frequencies (0.1-1 Hz) and was enhanced by the application of cytochalasin-D that degrades the actin cortex inside the cell membrane. This enhancement feature is only observed in tissues that have formed adhesions, because cell pellets initially do not show this signature, but develop this signature only after incubation enables adhesions to form.

Gypenoside XLIX, a naturally occurring gynosaponin, PPAR-alpha dependently inhibits LPS-induced tissue factor expression and activity in human THP-1 monocytic cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Huang, Tom Hsun-Wei; Van Hoan Tran; Roufogalis, Basil D.

2007-01-01

Tissue factor (TF) is involved not only in the progression of atherosclerosis and other cardiovascular diseases, but is also associated with tumor growth, metastasis, and angiogenesis and hence may be an attractive target for directed cancer therapeutics. Gynostemma pentaphyllum (GP) is widely used in the treatment of various cardiovascular diseases including atherosclerosis, as well as cancers. Gypenoside (Gyp) XLIX, a dammarane-type glycoside, is one of the prominent components in GP. We have recently reported Gyp XLIX to be a potent peroxisome proliferator-activated receptor (PPAR)-alpha activator. Here we demonstrate that Gyp XLIX (0-300 {mu}M) concentration dependently inhibited TF promoter activity aftermore » induction by the inflammatory stimulus lipopolysaccharide (LPS) in human monocytic THP-1 cells transfected with promoter reporter constructs pTF-LUC. Furthermore, Gyp XLIX inhibited LPS-induced TF mRNA and protein overexpression in THP-1 monocyte cells. Its inhibition of LPS-induced TF hyperactivity was further confirmed by chromogenic enzyme activity assay. The activities of Gyp XLIX reported in this study were similar to those of Wy-14643, a potent synthetic PPAR-alpha activator. Furthermore, the Gyp XLIX-induced inhibitory effect on TF luciferase activity was completely abolished in the presence of the PPAR-alpha selective antagonist MK-886. The present findings suggest that Gyp XLIX inhibits LPS-induced TF overexpression and enhancement of its activity in human THP-1 monocytic cells via PPAR-alpha-dependent pathways. The data provide new insights into the basis of the use of the traditional Chinese herbal medicine G. pentaphyllum for the treatment of cardiovascular and inflammatory diseases, as well as cancers.« less

Silencing of cytosolic NADP+-dependent isocitrate dehydrogenase gene enhances ethanol-induced toxicity in HepG2 cells.

PubMed

Yang, Eun Sun; Lee, Su-Min; Park, Jeen-Woo

2010-07-01

It has been shown that acute and chronic alcohol administrations increase the production of reactive oxygen species, lower cellular antioxidant levels and enhance oxidative stress in many tissues. We recently reported that cytosolic NADP(+)-dependent isocitrate dehydrogenase (IDPc) functions as an antioxidant enzyme by supplying NADPH to the cytosol. Upon exposure to ethanol, IDPc was susceptible to the loss of its enzyme activity in HepG2 cells. Transfection of HepG2 cells with an IDPc small interfering RNA noticeably downregulated IDPc and enhanced the cells' vulnerability to ethanol-induced cytotoxicity. Our results suggest that suppressing the expression of IDPc enhances ethanol-induced toxicity in HepG2 cells by further disruption of the cellular redox status.

The effect of dose enhancement near metal interfaces on synthetic diamond based X-ray dosimeters

NASA Astrophysics Data System (ADS)

Alamoudi, D.; Lohstroh, A.; Albarakaty, H.

2017-11-01

This study investigates the effects of dose enhancement on the photocurrent performance at metallic interfaces in synthetic diamond detectors based X-ray dosimeters as a function of bias voltages. Monte Carlo (MC) simulations with the BEAMnrc code were carried out to simulate the dose enhancement factor (DEF) and compared against the equivalent photocurrent ratio from experimental investigations. The MC simulation results show that the sensitive region for the absorbed dose distribution covers a few micrometers distances from the interface. Experimentally, two single crystals (SC) and one polycrystalline (PC) synthetic diamond samples were fabricated into detectors with carbon based electrodes by boron and carbon ion implantation. Subsequently; the samples were each mounted inside a tissue equivalent encapsulation to minimize unintended fluence perturbation. Dose enhancement was generated by placing copper, lead or gold near the active volume of the detectors using 50 kVp and 100 kVp X-rays relevant for medical dosimetry. The results show enhancement in the detectors' photocurrent performance when different metals are butted up to the diamond bulk as expected. The variation in the photocurrent measurement depends on the type of diamond samples, their electrodes' fabrication and the applied bias voltages indicating that the dose enhancement near the detector may modify their electronic performance.

Diode laser soft-tissue surgery: advancements aimed at consistent cutting, improved clinical outcomes.

PubMed

Romanos, Georgios E

2013-01-01

Laser dentistry and soft-tissue surgery, in particular, have become widely adopted in recent years. Significant cost reductions for dental lasers and the increasing popularity of CADCAM, among other factors, have contributed to a substantial increase in the installed base of dental lasers, especially soft-tissue lasers. New development in soft-tissue surgery, based on the modern understanding of laser-tissue interactions and contact soft-tissue surgery mechanisms, will bring a higher quality and consistency level to laser soft-tissue surgery. Recently introduced diode-laser technology enables enhanced control of side effects that result from tissue overheating and may improve soft-tissue surgical outcomes.

Trophic factors from adipose tissue-derived multi-lineage progenitor cells promote cytodifferentiation of periodontal ligament cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Sawada, Keigo; Takedachi, Masahide, E-mail: takedati@dent.osaka-u.ac.jp; Yamamoto, Satomi

Stem and progenitor cells are currently being investigated for their applicability in cell-based therapy for periodontal tissue regeneration. We recently demonstrated that the transplantation of adipose tissue-derived multi-lineage progenitor cells (ADMPCs) enhances periodontal tissue regeneration in beagle dogs. However, the molecular mechanisms by which transplanted ADMPCs induce periodontal tissue regeneration remain to be elucidated. In this study, trophic factors released by ADMPCs were examined for their paracrine effects on human periodontal ligament cell (HPDL) function. ADMPC conditioned medium (ADMPC-CM) up-regulated osteoblastic gene expression, alkaline phosphatase activity and calcified nodule formation in HPDLs, but did not significantly affect their proliferative response.more » ADMPCs secreted a number of growth factors, including insulin-like growth factor binding protein 6 (IGFBP6), hepatocyte growth factor and vascular endothelial growth factor. Among these, IGFBP6 was most highly expressed. Interestingly, the positive effects of ADMPC-CM on HPDL differentiation were significantly suppressed by transfecting ADMPCs with IGFBP6 siRNA. Our results suggest that ADMPCs transplanted into a defect in periodontal tissue release trophic factors that can stimulate the differentiation of HPDLs to mineralized tissue-forming cells, such as osteoblasts and cementoblasts. IGFBP6 may play crucial roles in ADMPC-induced periodontal regeneration. - Highlights: • ADMPC-derived humoral factors stimulate cytodifferentiation of HPDLs. • ADMPCs secret growth factors including IGFBP6, VEGF and HGF. • IGFBP6 is involved in the promotion effect of ADMPC-CM on HPDL cytodifferentiation.« less

Autophagy regulates tissue overgrowth in a context-dependent manner.

PubMed

Pérez, E; Das, G; Bergmann, A; Baehrecke, E H

2015-06-01

Autophagy is a catabolic process that has been implicated both as a tumor suppressor and in tumor progression. Here, we investigate this dichotomy in cancer biology by studying the influence of altered autophagy in Drosophila models of tissue overgrowth. We find that the impact of altered autophagy depends on both genotype and cell type. As previously observed in mammals, decreased autophagy suppresses Ras-induced eye epithelial overgrowth. In contrast, autophagy restricts epithelial overgrowth in a Notch-dependent eye model. Even though decreased autophagy did not influence Hippo pathway-triggered overgrowth, activation of autophagy strongly suppresses this eye epithelial overgrowth. Surprisingly, activation of autophagy enhanced Hippo pathway-driven overgrowth in glia cells. These results indicate that autophagy has different influences on tissue growth in distinct contexts, and highlight the importance of understanding the influence of autophagy on growth to augment a rationale therapeutic strategy.

[Oxygen-dependent processes in the blood cells from children with arterial hypertension concomitant with biliary dyskinesia].

PubMed

Mikashinovich, Z I; Nagornaia, G Iu; Kovalenko, T D; Zvereva, E A

2011-02-01

Age individuality is characterized by an imbalance of the molecular mechanisms of antioxidant defense in adolescents with arterial hypertension and biliary dyskinesia, as documented by an enzyme imbalance of the first line of antioxidant defense and H2O, accumulation, by a substantial increase in glutathione peroxidase activity, and by inhibition of the activity of glutathione-dependent enzymes. The considerable rise of 2,3-diphosphoglycerate suggests tissue hypoxia. With this, enhanced neutrophil elastase activity causes damage to the structural components of vascular wall connective tissue, resulting in the development of endothelial dysfunction.

Targeting Tissue Factor-Factor VIIa Signaling Pathway to Enhance Activity of mTOR Inhibitors in the Treatment of Breast Cancer

DTIC Science & Technology

2010-03-01

Salzberg M, Ostapenko V, Illiger HJ, Behringer D, Bardy -Bouxin N, Boni J , Kong S, Cincotta M, and Moore L. Phase II study of temsirolimus (CCI-779), a novel...interaction results in a tissue factor cytoplasmic domain- independent activation of protein synthesis, p70, and p90 S6 kinase phosphorylation. J ...mTOR Pathway in Breast Cancer. J Mammary Gland Biol Neoplasia 2006; 11: 53-61. 23. Guba M, Yezhelyev, Eichhorn ME, Schmid G, Ischenko, Papyan A

Spatial regulation of controlled bioactive factor delivery for bone tissue engineering

PubMed Central

Samorezov, Julia E.; Alsberg, Eben

2015-01-01

Limitations of current treatment options for critical size bone defects create a significant clinical need for tissue engineered bone strategies. This review describes how control over the spatiotemporal delivery of growth factors, nucleic acids, and drugs and small molecules may aid in recapitulating signals present in bone development and healing, regenerating interfaces of bone with other connective tissues, and enhancing vascularization of tissue engineered bone. State-of-the-art technologies used to create spatially controlled patterns of bioactive factors on the surfaces of materials, to build up 3D materials with patterns of signal presentation within their bulk, and to pattern bioactive factor delivery after scaffold fabrication are presented, highlighting their applications in bone tissue engineering. As these techniques improve in areas such as spatial resolution and speed of patterning, they will continue to grow in value as model systems for understanding cell responses to spatially regulated bioactive factor signal presentation in vitro, and as strategies to investigate the capacity of the defined spatial arrangement of these signals to drive bone regeneration in vivo. PMID:25445719

PASSIVE CAVITATION DETECTION DURING PULSED HIFU EXPOSURES OF EX VIVO TISSUES AND IN VIVO MOUSE PANCREATIC TUMORS

PubMed Central

Li, Tong; Chen, Hong; Khokhlova, Tatiana; Wang, Yak-Nam; Kreider, Wayne; He, Xuemei; Hwang, Joo Ha

2014-01-01

Pulsed high-intensity focused ultrasound (pHIFU) has been demonstrated to enhance vascular permeability, disrupt tumor barriers and enhance drug penetration into tumor tissue through acoustic cavitation. Monitoring of cavitation activity during pHIFU treatments and knowing the ultrasound pressure levels sufficient to reliably induce cavitation in a given tissue are therefore very important. Here, three metrics of cavitation activity induced by pHIFU and evaluated by confocal passive cavitation detection were introduced: cavitation probability, cavitation persistence and the level of the broadband acoustic emissions. These metrics were used to characterize cavitation activity in several ex vivo tissue types (bovine tongue and liver and porcine adipose tissue and kidney) and gel phantoms (polyacrylamide and agarose) at varying peak-rarefactional focal pressures (1–12 MPa) during the following pHIFU protocol: frequency 1.1 MHz, pulse duration 1 ms, pulse repetition frequency 1 Hz. To evaluate the relevance of the measurements in ex vivo tissue, cavitation metrics were also investigated and compared in the ex vivo and in vivo murine pancreatic tumors that develop spontaneously in transgenic KPC mice and closely recapitulate human disease in their morphology. The cavitation threshold, defined at 50 % cavitation probability, was found to vary broadly among the investigated tissues (within 2.5–10 MPa), depending mostly on the water-lipid ratio that characterizes the tissue composition. Cavitation persistence and the intensity of broadband emissions depended both on tissue structure and lipid concentration. Both the cavitation threshold and broadband noise emission level were similar between ex vivo and in vivo pancreatic tumor tissue. The largest difference between in vivo and ex vivo settings was found in the pattern of cavitation occurrence throughout pHIFU exposure: it was sporadic in vivo, but ex vivo it decreased rapidly and stopped over the first few pulses. Cavitation activity depended on the interplay between the destruction and circulation of cavitation nuclei, which are not only used up by HIFU treatment but also replenished or carried away by circulation in vivo. These findings are important for treatment planning and optimization in pHIFU-induced drug delivery, in particular for pancreatic tumors. PMID:24613635

Hydrodynamics of stratified epithelium: Steady state and linearized dynamics

NASA Astrophysics Data System (ADS)

Yeh, Wei-Ting; Chen, Hsuan-Yi

2016-05-01

A theoretical model for stratified epithelium is presented. The viscoelastic properties of the tissue are assumed to be dependent on the spatial distribution of proliferative and differentiated cells. Based on this assumption, a hydrodynamic description of tissue dynamics at the long-wavelength, long-time limit is developed, and the analysis reveals important insights into the dynamics of an epithelium close to its steady state. When the proliferative cells occupy a thin region close to the basal membrane, the relaxation rate towards the steady state is enhanced by cell division and cell apoptosis. On the other hand, when the region where proliferative cells reside becomes sufficiently thick, a flow induced by cell apoptosis close to the apical surface enhances small perturbations. This destabilizing mechanism is general for continuous self-renewal multilayered tissues; it could be related to the origin of certain tissue morphology, tumor growth, and the development pattern.

Hypoxia Inducible Factor-Stabilizing Bioactive Glasses for Directing Mesenchymal Stem Cell Behavior

PubMed Central

Azevedo, Maria M.; Tsigkou, Olga; Nair, Rekha; Jones, Julian R.; Jell, Gavin

2015-01-01

Oxygen tension is a known regulator of mesenchymal stem cell (MSC) plasticity, differentiation, proliferation, and recruitment to sites of injury. Materials capable of affecting the MSC oxygen-sensing pathway, independently of the environmental oxygen pressure, are therefore of immense interest to the tissue engineering (TE) and regenerative medicine community. In this study, we describe the evaluation of the effect of hypoxia inducible factor (HIF)-stabilizing bioactive glasses (BGs) on human MSCs. The dissolution products from these hypoxia-mimicking BGs stabilized HIF-1α in a concentration-dependent manner, altered cell proliferation and metabolism, and upregulated a number of genes involved in the hypoxic response (HIF1A, HIF2A, and VHL), MSC survival (SAG and BCL2), extracellular matrix remodeling (MMP1), and angiogenesis (VEGF and PDGF). These HIF-stabilizing materials can therefore be used to improve MSC survival and enhance regeneration in a number of TE strategies. PMID:25167933

Antiobesity Effect of Exopolysaccharides Isolated from Kefir Grains.

PubMed

Lim, Juha; Kale, Madhuvanti; Kim, Dong-Hyeon; Kim, Hong-Seok; Chon, Jung-Whan; Seo, Kun-Ho; Lee, Hyeon Gyu; Yokoyama, Wallace; Kim, Hyunsook

2017-11-22

Physiological properties of water-soluble exopolysaccharides (EPS) and residues after EPS removal (Res) from the probiotic kefir were determined in high-fat (HF) diet-fed C57BL/6J mice. EPS solutions showed rheological properties and lower viscosity compared to those of β-glucan (BG). EPS significantly suppressed the adipogenesis of 3T3-L1 preadipocytes in a dose-dependent manner. Mice were fed HF diets containing 5% EPS, 5% BG, 8% Res, or 5% microcrystalline cellulose (control) for 4 weeks. Compared with the control, EPS supplementation significantly reduced HF diet-induced body weight gain, adipose tissue weight, and plasma very-low-density lipoprotein cholesterol concentration (P < 0.05). Res and BG significantly reduced body weight gain; however, reduction in adipose tissue weight was not statistically significant, suggesting that the antiobesity effect of EPS occurs due to viscosity and an additional factor. EPS supplementation significantly enhanced abundance of Akkermansia spp. in feces. These data indicate that EPS shows significant antiobesity effects possibly via intestinal microbiota alterations.

A universal formula for the field enhancement factor

NASA Astrophysics Data System (ADS)

Biswas, Debabrata

2018-04-01

The field enhancement factor (FEF) is an important quantity in field emission calculations since the tunneling electron current depends very sensitively on its magnitude. The exact dependence of FEF on the emitter height h, the radius of curvature at the apex Ra, as well as the shape of the emitter base are still largely unknown. In this work, a universal formula for the field enhancement factor is derived for a single emitter. It depends on the ratio h/Ra and has the form γ a = ( 2 h / R a ) / [ α 1 ln ( 4 h / R a ) - α 2 ] , where α1 and α2 depend on the charge distribution on the emitter. Numerical results show that a simpler form γ a = ( 2 h / R a ) / [ ln ( 4 h / R a ) - α ] is equally valid with α depending on the emitter-base. Thus, for the hyperboloid, conical, and ellipsoid emitters, the value of α is 0, 0.88, and 2, while for the cylindrical base, α ≃ 2.6.

delta(9)-Tetrahydrocannabinol-dependent mice undergoing withdrawal display impaired spatial memory.

PubMed

Wise, Laura E; Varvel, Stephen A; Selley, Dana E; Wiebelhaus, Jason M; Long, Kelly A; Middleton, Lisa S; Sim-Selley, Laura J; Lichtman, Aron H

2011-10-01

Cannabis users display a constellation of withdrawal symptoms upon drug discontinuation, including sleep disturbances, irritability, and possibly memory deficits. In cannabinoid-dependent rodents, the CB(1) antagonist rimonabant precipitates somatic withdrawal and enhances forskolin-stimulated adenylyl cyclase activity in cerebellum, an effect opposite that of acutely administered ∆(9)-tetrahydrocannabinol (THC), the primary constituent in cannabis. Here, we tested whether THC-dependent mice undergoing rimonabant-precipitated withdrawal display short-term spatial memory deficits, as assessed in the Morris water maze. We also evaluated whether rimonabant would precipitate adenylyl cyclase superactivation in hippocampal and cerebellar tissue from THC-dependent mice. Rimonabant significantly impaired spatial memory of THC-dependent mice at lower doses than those necessary to precipitate somatic withdrawal behavior. In contrast, maze performance was near perfect in the cued task, suggesting sensorimotor function and motivational factors were unperturbed by the withdrawal state. Finally, rimonabant increased adenylyl cyclase activity in cerebellar, but not in hippocampal, membranes. The memory disruptive effects of THC undergo tolerance following repeated dosing, while the withdrawal state leads to a rebound deficit in memory. These results establish spatial memory impairment as a particularly sensitive component of cannabinoid withdrawal, an effect that may be mediated through compensatory changes in the cerebellum.

Enhancing nerve regeneration in the peripheral nervous system using polymeric scaffolds, stem cell engineering and nanoparticle delivery system

NASA Astrophysics Data System (ADS)

Sharma, Anup Dutt

Peripheral nerve regeneration is a complex biological process responsible for regrowth of neural tissue following a nerve injury. The main objective of this project was to enhance peripheral nerve regeneration using interdisciplinary approaches involving polymeric scaffolds, stem cell therapy, drug delivery and high content screening. Biocompatible and biodegradable polymeric materials such as poly (lactic acid) were used for engineering conduits with micropatterns capable of providing mechanical support and orientation to the regenerating axons and polyanhydrides for fabricating nano/microparticles for localized delivery of neurotrophic growth factors and cytokines at the site of injury. Transdifferentiated bone marrow stromal cells or mesenchymal stem cells (MSCs) were used as cellular replacements for lost native Schwann cells (SCs) at the injured nerve tissue. MSCs that have been transdifferentiated into an SC-like phenotype were tested as a substitute for the myelinating SCs. Also, genetically modified MSCs were engineered to hypersecrete brain- derived neurotrophic factor (BDNF) and glial cell line-derived neurotrophic factor (GDNF) to secrete therapeutic factors which Schwann cell secrete. To further enhance the regeneration, nerve growth factor (NGF) and interleukin-4 (IL4) releasing polyanhydrides nano/microparticles were fabricated and characterized in vitro for their efficacy. Synergistic use of these proposed techniques was used for fabricating a multifunctional nerve regeneration conduit which can be used as an efficient tool for enhancing peripheral nerve regeneration.

Inflammation-Dependent IL18 Signaling Restricts Hepatocellular Carcinoma Growth by Enhancing the Accumulation and Activity of Tumor-Infiltrating Lymphocytes.

PubMed

Markowitz, Geoffrey J; Yang, Pengyuan; Fu, Jing; Michelotti, Gregory A; Chen, Rui; Sui, Jianhua; Yang, Bin; Qin, Wen-Hao; Zhang, Zheng; Wang, Fu-Sheng; Diehl, Anna Mae; Li, Qi-Jing; Wang, Hongyang; Wang, Xiao-Fan

2016-04-15

Chronic inflammation in liver tissue is an underlying cause of hepatocellular carcinoma. High levels of inflammatory cytokine IL18 in the circulation of patients with hepatocellular carcinoma correlates with poor prognosis. However, conflicting results have been reported for IL18 in hepatocellular carcinoma development and progression. In this study, we used tissue specimens from hepatocellular carcinoma patients and clinically relevant mouse models of hepatocellular carcinoma to evaluate IL18 expression and function. In a mouse model of liver fibrosis that recapitulates a tumor-promoting microenvironment, global deletion of the IL18 receptor IL18R1 enhanced tumor growth and burden. Similarly, in a carcinogen-induced model of liver tumorigenesis, IL18R1 deletion increased tumor burden. Mechanistically, we found that IL18 exerted inflammation-dependent tumor-suppressive effects largely by promoting the differentiation, activity, and survival of tumor-infiltrating T cells. Finally, differences in the expression of IL18 in tumor tissue versus nontumor tissue were more predictive of patient outcome than overall tissue expression. Taken together, our findings resolve a long-standing contradiction regarding a tumor-suppressive role for IL18 in established hepatocellular carcinoma and provide a mechanistic explanation for the complex relationship between its expression pattern and hepatocellular carcinoma prognosis. Cancer Res; 76(8); 2394-405. ©2016 AACR. ©2016 American Association for Cancer Research.

Macrophage Control of Phagocytosed Mycobacteria Is Increased by Factors Secreted by Alveolar Epithelial Cells through Nitric Oxide Independent Mechanisms

PubMed Central

Freidl, Raphaela; Fernández, Carmen

2014-01-01

Tissue-resident macrophages are heterogeneous with tissue-specific and niche-specific functions. Thus, simplified models of macrophage activation do not explain the extent of heterogeneity seen in vivo. We focus here on the respiratory tract and ask whether factors secreted by alveolar epithelial cells (AEC) can influence the functionality of resident pulmonary macrophages (PuM). We have previously reported that factors secreted by AEC increase control of intracellular growth of BCG in macrophages. In the current study, we also aimed to investigate possible mechanisms by which AEC-derived factors increase intracellular control of BCG in both primary murine interstitial macrophages, and bone marrow-derived macrophages and characterize further the effect of these factors on macrophage differentiation. We show that; a) in contrast to other macrophage types, IFN-γ did not increase intracellular growth control of Mycobacterium bovis, Bacillus Calmette-Guérin (BCG) by interstitial pulmonary macrophages although the same macrophages could be activated by factors secreted by AEC; b) the lack of response of pulmonary macrophages to IFN-γ was apparently regulated by suppressor of cytokine signaling (SOCS)1; c) AEC-derived factors did not induce pro-inflammatory pathways induced by IFN-γ e.g. expression of inducible nitric oxide synthase (iNOS), secretion of nitric oxide (NO), or IL-12, d) in contrast to IFN-γ, intracellular bacterial destruction induced by AEC-derived factors was not dependent on iNOS transcription and NO production. Collectively, our data show that PuM were restricted in inflammatory responses mediated by IFN-γ through SOCS1 and that factors secreted by AEC- enhanced the microbicidal capacities of macrophages by iNOS independent mechanisms. PMID:25089618

Effects of Structural Properties of Electrospun TiO2 Nano-fiber Meshes on their Osteogenic Potential

PubMed Central

Wang, Xiaokun; Gittens, Rolando A.; Song, Rosemary; Tannenbaum, Rina; Olivares-Navarrete, Rene; Schwartz, Zvi; Chen, Haifeng; Boyan, Barbara D.

2011-01-01

Ideal outcomes in the field of tissue engineering and regenerative medicine involve biomaterials that can enhance cell differentiation and production of local factors for natural tissue regeneration without the use of systemic drugs. Biomaterials typically used in tissue engineering applications include polymeric scaffolds that mimic the 3-D structural environment of the native tissue, but these are often functionalized with proteins or small peptides to improve their biological performance. For bone applications, titanium (Ti) implants, or more appropriately the titania (TiO2) passive oxide layer formed on their surface, have been shown to enhance osteoblast differentiation in vitro and to promote osseointegration in vivo. In this study we evaluated the effect on osteoblast differentiation of pure TiO2 nano-fiber meshes with different surface micro-roughness and nano-fiber diameters, prepared by the electrospinning method. MG63 cells were seeded on TiO2 meshes, and cell number, differentiation markers and local factor production were analyzed. The results showed that cells grew throughout the entire surfaces and with similar morphology in all groups. Cell number was sensitive to surface micro-roughness, whereas cell differentiation and local factor production was regulated by both surface roughness and nano-fiber diameter. These results indicate that scaffold structural cues alone can be used to drive cell differentiation and create an osteogenic environment without the use of exogenous factors. PMID:22075122

Chronic ethanol consumption modulates growth factor release, mucosal cytokine production, and microRNA expression in nonhuman primates.

PubMed

Asquith, Mark; Pasala, Sumana; Engelmann, Flora; Haberthur, Kristen; Meyer, Christine; Park, Byung; Grant, Kathleen A; Messaoudi, Ilhem

2014-04-01

Chronic alcohol consumption has been associated with enhanced susceptibility to both systemic and mucosal infections. However, the exact mechanisms underlying this enhanced susceptibility remain incompletely understood. Using a nonhuman primate model of ethanol (EtOH) self-administration, we examined the impact of chronic alcohol exposure on immune homeostasis, cytokine, and growth factor production in peripheral blood, lung, and intestinal mucosa following 12 months of chronic EtOH exposure. EtOH exposure inhibited activation-induced production of growth factors hepatocyte growth factor (HGF), granulocyte colony-stimulating factor (G-CSF), and vascular-endothelial growth factor (VEGF) by peripheral blood mononuclear cells (PBMC). Moreover, EtOH significantly reduced the frequency of colonic Th1 and Th17 cells in a dose-dependent manner. In contrast, we did not observe differences in lymphocyte frequency or soluble factor production in the lung of EtOH-consuming animals. To uncover mechanisms underlying reduced growth factor and Th1/Th17 cytokine production, we compared expression levels of microRNAs in PBMC and intestinal mucosa. Our analysis revealed EtOH-dependent up-regulation of distinct microRNAs in affected tissues (miR-181a and miR-221 in PBMC; miR-155 in colon). Moreover, we were able to detect reduced expression of the transcription factors STAT3 and ARNT, which regulate expression of VEGF, G-CSF, and HGF and contain targets for these microRNAs. To confirm and extend these observations, PBMC were transfected with either mimics or antagomirs of miR-181 and miR-221, and protein levels of the transcription factors and growth factors were determined. Transfection of microRNA mimics led to a reduction in both STAT3/ARNT as well as VEGF/HGF/G-CSF levels. The opposite outcome was observed when microRNA antagomirs were transfected. Chronic EtOH consumption significantly disrupts both peripheral and mucosal immune homeostasis, and this dysregulation may be mediated by changes in microRNA expression. Copyright © 2013 by the Research Society on Alcoholism.

Enhanced Growth and Hepatic Differentiation of Fetal Liver Epithelial Cells through Combinational and Temporal Adjustment of Soluble Factors

PubMed Central

Qian, Lichuan; Krause, Diane S.; Saltzman, W. Mark

2012-01-01

Fetal liver epithelial cells (FLEC) are valuable for liver cell therapy and tissue engineering, but methods for culture and characterization of these cells are not well developed. This work explores the influence of multiple soluble factors on FLEC, with the long-term goal of developing an optimal culture system to generate functional liver tissue. Our comparative analysis suggests hepatocyte growth factor (HGF) is required throughout the culture period. In the presence of HGF, addition of oncostatin M (OSM) at culture initiation results in concurrent growth and maturation, while constant presence of protective agents like ascorbic acid enhances cell survival. Study observations led to the development of a culture medium that provided optimal growth and hepatic differentiation conditions. FLEC expansion was observed to be ~2 fold of that under standard conditions, albumin secretion rate was 2 – 3 times greater than maximal values obtained with other media, and the highest level of glycogen accumulation among all conditions was observed with the developed medium. Our findings serve to advance culture methods for liver progenitors in cell therapy and tissue engineering applications. PMID:21922669

Internal quantum efficiency enhancement of GaInN/GaN quantum-well structures using Ag nanoparticles

DOE Office of Scientific and Technical Information (OSTI.GOV)

Iida, Daisuke; Department of Photonics Engineering, Technical University of Denmark, 2800 Lyngby; Faculty of Science and Technology, Meijo University, 1-501 Shiogamaguchi Tempaku, 468-8502 Nagoya

2015-09-15

We report internal quantum efficiency enhancement of thin p-GaN green quantum-well structure using self-assembled Ag nanoparticles. Temperature dependent photoluminescence measurements are conducted to determine the internal quantum efficiency. The impact of excitation power density on the enhancement factor is investigated. We obtain an internal quantum efficiency enhancement by a factor of 2.3 at 756 W/cm{sup 2}, and a factor of 8.1 at 1 W/cm{sup 2}. A Purcell enhancement up to a factor of 26 is estimated by fitting the experimental results to a theoretical model for the efficiency enhancement factor.

Delivery of growth factors for tissue regeneration and wound healing.

PubMed

Koria, Piyush

2012-06-01

Growth factors are soluble secreted proteins capable of affecting a variety of cellular processes important for tissue regeneration. Consequently, the self-healing capacity of patients can be augmented by artificially enhancing one or more processes important for healing through the application of growth factors. However, their application in clinics remains limited due to lack of robust delivery systems and biomaterial carriers. Interestingly, all clinically approved therapies involving growth factors utilize some sort of a biomaterial carrier for growth factor delivery. This suggests that biomaterial delivery systems are extremely important for successful usage of growth factors in regenerative medicine. This review outlines the role of growth factors in tissue regeneration, and their application in both pre-clinical animal models of regeneration and clinical trials is discussed. Additionally, current status of biomaterial substrates and sophisticated delivery systems such as nanoparticles for delivery of exogenous growth factors and peptides in humans are reviewed. Finally, issues and possible future research directions for growth factor therapy in regenerative medicine are discussed.

Uric Acid Secretion from Adipose Tissue and Its Increase in Obesity*

PubMed Central

Tsushima, Yu; Nishizawa, Hitoshi; Tochino, Yoshihiro; Nakatsuji, Hideaki; Sekimoto, Ryohei; Nagao, Hirofumi; Shirakura, Takashi; Kato, Kenta; Imaizumi, Keiichiro; Takahashi, Hiroyuki; Tamura, Mizuho; Maeda, Norikazu; Funahashi, Tohru; Shimomura, Iichiro

2013-01-01

Obesity is often accompanied by hyperuricemia. However, purine metabolism in various tissues, especially regarding uric acid production, has not been fully elucidated. Here we report, using mouse models, that adipose tissue could produce and secrete uric acid through xanthine oxidoreductase (XOR) and that the production was enhanced in obesity. Plasma uric acid was elevated in obese mice and attenuated by administration of the XOR inhibitor febuxostat. Adipose tissue was one of major organs that had abundant expression and activities of XOR, and adipose tissues in obese mice had higher XOR activities than those in control mice. 3T3-L1 and mouse primary mature adipocytes produced and secreted uric acid into culture medium. The secretion was inhibited by febuxostat in a dose-dependent manner or by gene knockdown of XOR. Surgical ischemia in adipose tissue increased local uric acid production and secretion via XOR, with a subsequent increase in circulating uric acid levels. Uric acid secretion from whole adipose tissue was increased in obese mice, and uric acid secretion from 3T3-L1 adipocytes was increased under hypoxia. Our results suggest that purine catabolism in adipose tissue could be enhanced in obesity. PMID:23913681

The transcription factor Grainy head primes epithelial enhancers for spatiotemporal activation by displacing nucleosomes.

PubMed

Jacobs, Jelle; Atkins, Mardelle; Davie, Kristofer; Imrichova, Hana; Romanelli, Lucia; Christiaens, Valerie; Hulselmans, Gert; Potier, Delphine; Wouters, Jasper; Taskiran, Ibrahim I; Paciello, Giulia; González-Blas, Carmen B; Koldere, Duygu; Aibar, Sara; Halder, Georg; Aerts, Stein

2018-06-04

Transcriptional enhancers function as docking platforms for combinations of transcription factors (TFs) to control gene expression. How enhancer sequences determine nucleosome occupancy, TF recruitment and transcriptional activation in vivo remains unclear. Using ATAC-seq across a panel of Drosophila inbred strains, we found that SNPs affecting binding sites of the TF Grainy head (Grh) causally determine the accessibility of epithelial enhancers. We show that deletion and ectopic expression of Grh cause loss and gain of DNA accessibility, respectively. However, although Grh binding is necessary for enhancer accessibility, it is insufficient to activate enhancers. Finally, we show that human Grh homologs-GRHL1, GRHL2 and GRHL3-function similarly. We conclude that Grh binding is necessary and sufficient for the opening of epithelial enhancers but not for their activation. Our data support a model positing that complex spatiotemporal expression patterns are controlled by regulatory hierarchies in which pioneer factors, such as Grh, establish tissue-specific accessible chromatin landscapes upon which other factors can act.

Prediction of oxygen distribution in aortic valve leaflet considering diffusion and convection.

PubMed

Wang, Ling; Korossis, Sotirios; Fisher, John; Ingham, Eileen; Jin, Zhongmin

2011-07-01

Oxygen supply and transport is an important consideration in the development of tissue engineered constructs. Previous studies from our group have focused on the effect of tissue thickness on the oxygen diffusion within a three-dimensional aortic valve leaflet model, and highlighted the necessity for additional transport mechanisms such as oxygen convection. The aims of this study were to investigate the effect of interstitial fluid flow within the aortic valve leaflet, induced by the cyclic loading of the leaflet, on oxygen transport. Indentation testing and finite element modelings were employed to derive the biphasic properties of the leaflet tissue. The biphasic properties were subsequently used in the computational modeling of oxygen convection in the leaflet, which was based on the effective interstitial fluid velocity and the tissue deformation. Subsequently, the oxygen profile was predicted within the valve leaflet model by solving the diffusion and convection equation simultaneously utilizing the finite difference method. The compression modulus (E) and hydraulic permeability were determined by adapting a finite element model to the experimental indentation test on valvular tissue, E = 0.05MPa, and k =2.0 mm4/Ns. Finite element model of oxygen convection in valvular tissue incorporating the predicted biphasic properties was developed and the interstitial fluid flow rate was calculated falling in range of 0.025-0.25 mm/s depending on the tissue depth. Oxygen distribution within valvular tissue was predicted using one-dimensional oxygen diffusion model taking into consider the interstitial fluid effect. It was found that convection did enhance the oxygen transport in valvular tissue by up to 68% increase in the minimum oxygen tension within the tissue, depending on the strain level of the tissue as reaction of the magnitude and frequencies of the cardiac loading. The effective interstitial fluid velocity was found to play an important role in enhancing the oxygen transport within the valve leaflet. Such an understanding is important in the development of valvular tissue engineered constructs.

Tissue factor transcription driven by Egr-1 is a critical mechanism of murine pulmonary fibrin deposition in hypoxia

PubMed Central

Yan, Shi-Fang; Zou, Yu Shan; Gao, Yun; Zhai, Chao; Mackman, Nigel; Lee, Stephen L.; Milbrandt, Jeffrey; Pinsky, David; Kisiel, Walter; Stern, David

1998-01-01

Local hypoxemia and stasis trigger thrombosis. We have demonstrated previously that in a murine model of normobaric hypoxia pulmonary fibrin deposition is a result of expression of tissue factor, especially in oxygen-deprived mononuclear phagocytes (MPs). We now show that transcription factor early-growth-response gene product (Egr-1) is rapidly activated in hypoxia, both in vitro and in vivo, and is responsible for transcription and expression of tissue factor in hypoxic lung. MPs and HeLa cells subjected to hypoxia (pO2 ≈13 torr) had increased levels of tissue factor transcripts (≈18-fold) and an increased rate of transcription (≈15-fold), based on nuclear run-on analysis. Gel-shift analysis of nuclear extracts from hypoxic MPs and HeLa cells demonstrated increased DNA-binding activity at the serum response region (SRR; −111/+14 bp) of the tissue factor promoter at Egr-1 motifs. Using 32P-labeled Egr consensus oligonucleotide, we observed induction of DNA-binding activity in nuclear extracts from hypoxic lung and HeLa cells because of activation of Egr-1, by means of supershift analysis. Transient transfection of HeLa cells with chimeric plasmids containing wild-type or mutant SRR from the tissue factor promoter showed that intact Sp1 sites are necessary for basal promoter activity, whereas the integrity of Egr-1 sites was required for hypoxia-enhanced expression. A central role for Egr-1 in hypoxia-mediated tissue factor expression was confirmed by experiments with homozygous Egr-1 null mice; wild-type mice subjected to oxygen deprivation expressed tissue factor and showed fibrin deposition, but hypoxic homozygous Egr-1 null mice displayed neither tissue factor nor fibrin. These data delineate a novel biology for hypoxia-induced fibrin deposition, in which oxygen deprivation-induced activation of Egr-1, resulting in expression of tissue factor, has an unexpected and central role. PMID:9653181

Approaches to improve angiogenesis in tissue-engineered skin.

PubMed

Sahota, Parbinder S; Burn, J Lance; Brown, Nicola J; MacNeil, Sheila

2004-01-01

A problem with tissue-engineered skin is clinical failure due to delays in vascularization. The aim of this study was to explore a number of simple strategies to improve angiogenesis/vascularization using a tissue-engineered model of skin to which small vessel human dermal microvascular endothelial cells were added. For the majority of these studies, a modified Guirguis chamber was used, which allowed the investigation of several variables within the same experiment using the same human dermis; cell type, angiogenic growth factors, the influence of keratinocytes and fibroblasts, mechanical penetration of the human dermis, the site of endothelial cell addition, and the influence of hypoxia were all examined. A qualitative scoring system was used to assess the impact of these factors on the penetration of endothelial cells throughout the dermis. Similar results were achieved using freshly isolated small vessel human dermal microvascular endothelial cells or an endothelial cell line and a minimum cell seeding density was identified. Cell penetration was not influenced by the addition of angiogenic growth factors (vascular endothelial growth factor and basic fibroblast growth factor); similarly, including epidermal keratinocytes or dermal fibroblasts did not encourage endothelial cell entry, and neither did mechanical introduction of holes throughout the dermis. Two factors were identified that significantly enhanced endothelial cell penetration into the dermis: hypoxia and the site of endothelial cell addition. Endothelial cells added from the papillary surface entered into the dermis much more effectively than when cells were added to the reticular surface of the dermis. We conclude that this model is valuable in improving our understanding of how to enhance vascularization of tissue-engineered grafts.

Multivalent conjugates of basic fibroblast growth factor enhance in vitro proliferation and migration of endothelial cells.

PubMed

Zbinden, Aline; Browne, Shane; Altiok, Eda I; Svedlund, Felicia L; Jackson, Wesley M; Healy, Kevin E

2018-05-01

Growth factors hold great promise for regenerative therapies. However, their clinical use has been halted by poor efficacy and rapid clearance from tissue, necessitating the delivery of extremely high doses to achieve clinical effectiveness which has raised safety concerns. Thus, strategies to either enhance growth factor activity at low doses or to increase their residence time within target tissues are necessary for clinical success. In this study, we generated multivalent conjugates (MVCs) of basic fibroblast growth factor (bFGF), a key growth factor involved in angiogenesis and wound healing, to hyaluronic acid (HyA) polymer chains. Multivalent bFGF conjugates (mvbFGF) were fabricated with minimal non-specific interaction observed between bFGF and the HyA chain. The hydrodynamic radii of mvbFGF ranged from ∼50 to ∼75 nm for conjugation ratios of bFGF to HyA chains at low (10 : 1) and high (30 : 1) feed ratios, respectively. The mvbFGF demonstrated enhanced bioactivity compared to unconjugated bFGF in assays of cell proliferation and migration, processes critical to angiogenesis and tissue regeneration. The 30 : 1 mvbFGF outperformed the 10 : 1 conjugate, which could be due to either FGF receptor clustering or interference with receptor mediated internalization and signal deactivation. This study simultaneously investigated the role of both protein to polymer ratio and multivalent conjugate size on their bioactivity, and determined that increasing the protein-to-polymer ratio and conjugate size resulted in greater cell bioactivity.

Super-lncRNAs: identification of lncRNAs that target super-enhancers via RNA:DNA:DNA triplex formation.

PubMed

Soibam, Benjamin

2017-11-01

Super-enhancers are characterized by high levels of Mediator binding and are major contributors to the expression of their associated genes. They exhibit high levels of local chromatin interactions and a higher order of local chromatin organization. On the other hand, lncRNAs can localize to specific DNA sites by forming a RNA:DNA:DNA triplex, which in turn can contribute to local chromatin organization. In this paper, we characterize a new class of lncRNAs called super-lncRNAs that target super-enhancers and which can contribute to the local chromatin organization of the super-enhancers. Using a logistic regression model based on the number of RNA:DNA:DNA triplex sites a lncRNA forms within the super-enhancer, we identify 442 unique super-lncRNA transcripts in 27 different human cell and tissue types; 70% of these super-lncRNAs were tissue restricted. They primarily harbor a single triplex-forming repeat domain, which forms an RNA:DNA:DNA triplex with multiple anchor DNA sites (originating from transposable elements) within the super-enhancers. Super-lncRNAs can be grouped into 17 different clusters based on the tissue or cell lines they target. Super-lncRNAs in a particular cluster share common short structural motifs and their corresponding super-enhancer targets are associated with gene ontology terms pertaining to the tissue or cell line. Super-lncRNAs may use these structural motifs to recruit and transport necessary regulators (such as transcription factors and Mediator complexes) to super-enhancers, influence chromatin organization, and act as spatial amplifiers for key tissue-specific genes associated with super-enhancers. © 2017 Soibam; Published by Cold Spring Harbor Laboratory Press for the RNA Society.

Response functions for computing absorbed dose to skeletal tissues from photon irradiation--an update.

PubMed

Johnson, Perry B; Bahadori, Amir A; Eckerman, Keith F; Lee, Choonsik; Bolch, Wesley E

2011-04-21

A comprehensive set of photon fluence-to-dose response functions (DRFs) is presented for two radiosensitive skeletal tissues-active and total shallow marrow-within 15 and 32 bone sites, respectively, of the ICRP reference adult male. The functions were developed using fractional skeletal masses and associated electron-absorbed fractions as reported for the UF hybrid adult male phantom, which in turn is based upon micro-CT images of trabecular spongiosa taken from a 40 year male cadaver. The new DRFs expand upon both the original set of seven functions produced in 1985, and a 2007 update calculated under the assumption of secondary electron escape from spongiosa. In this study, it is assumed that photon irradiation of the skeleton will yield charged particle equilibrium across all spongiosa regions at energies exceeding 200 keV. Kerma coefficients for active marrow, inactive marrow, trabecular bone and spongiosa at higher energies are calculated using the DRF algorithm setting the electron-absorbed fraction for self-irradiation to unity. By comparing kerma coefficients and DRF functions, dose enhancement factors and mass energy-absorption coefficient (MEAC) ratios for active marrow to spongiosa were derived. These MEAC ratios compared well with those provided by the NIST Physical Reference Data Library (mean difference of 0.8%), and the dose enhancement factors for active marrow compared favorably with values calculated in the well-known study published by King and Spiers (1985 Br. J. Radiol. 58 345-56) (mean absolute difference of 1.9 percentage points). Additionally, dose enhancement factors for active marrow were shown to correlate well with the shallow marrow volume fraction (R(2) = 0.91). Dose enhancement factors for the total shallow marrow were also calculated for 32 bone sites representing the first such derivation for this target tissue.

Synthesis of and in vitro and in vivo evaluation of a novel TGF-β1-SF-CS three-dimensional scaffold for bone tissue engineering.

PubMed

Tong, Shuang; Xu, Da-Peng; Liu, Zi-Mei; Du, Yang; Wang, Xu-Kai

2016-08-01

The role of transforming growth factor-β1 (TGF-β1) in normal human fracture healing has been previously demonstrated. The objective of the present study was to examine the biocompatibility of TGF-β1-silk fibroin-chitosan (TGF-β1-SF-CS) three-dimensional (3D) scaffolds in order to construct an ideal scaffold for bone tissue engineering. We added TGF-β1 directly to the SF-CS scaffold to construct a 3D scaffold for the first time, to the best of our knowledge, and performed evaluations to determine whether it may have potential applications as a growth factor delivery device. Bone marrow-derived mesenchymal stem cells (BMSCs) were seeded on the TGF-β1-SF-CS scaffolds and the silk fibroin-chitosan (SF-CS) scaffolds. On the TGF-β1‑SF-CS and the SF-CS scaffolds, the cell adhesion rate increased in a time‑dependent manner. Using a Cell Counting Kit-8 (CCK-8) assay and analyzing the alkaline phosphatase (ALP) expression proved that TGF-β1 significantly enhanced the growth and proliferation of BMSCs on the SF-CS scaffolds in a time-dependent manner. To examine the in vivo biocompatibility and osteogenesis of the TGF-β1‑SF-CS scaffolds, the TGF-β1-SF-CS scaffolds and the SF-CS scaffolds were implanted in rabbit mandibles and studied histologically and microradiographically. The 3D computed tomography (CT) scan and histological examinations of the samples showed that the TGF-β1-SF-CS scaffolds exhibited good biocompatibility and extensive osteoconductivity with the host bone after 8 weeks. Moreover, the introduction of TGF-β1 to the SF-CS scaffolds markedly enhanced the efficiency of new bone formation, and this was confirmed using bone mineral density (BMD) and biomechanical evaluation, particularly at 8 weeks after implantation. We demonstrated that the TGF-β1‑SF-CS scaffolds possessed as good biocompatibility and osteogenesis as the hybrid ones. Taken together, these findings indicate that the TGF-β1-SF-CS scaffolds fulfilled the basic requirements of bone tissue engineering, and have the potential to be applied in orthopedic, reconstructive and maxillofacial surgery. Thus, TGF-β1-SF-CS composite scaffolds represent a promising, novel type of scaffold for use in bone tissue engineering.

Heterogeneous engineered cartilage growth results from gradients of media-supplemented active TGF-β and is ameliorated by the alternative supplementation of latent TGF-β.

PubMed

Albro, Michael B; Nims, Robert J; Durney, Krista M; Cigan, Alexander D; Shim, Jay J; Vunjak-Novakovic, Gordana; Hung, Clark T; Ateshian, Gerard A

2016-01-01

Transforming growth factor beta (TGF-β) has become one of the most widely utilized mediators of engineered cartilage growth. It is typically exogenously supplemented in the culture medium in its active form, with the expectation that it will readily transport into tissue constructs through passive diffusion and influence cellular biosynthesis uniformly. The results of this investigation advance three novel concepts regarding the role of TGF-β in cartilage tissue engineering that have important implications for tissue development. First, through the experimental and computational analysis of TGF-β concentration distributions, we demonstrate that, contrary to conventional expectations, media-supplemented exogenous active TGF-β exhibits a pronounced concentration gradient in tissue constructs, resulting from a combination of high-affinity binding interactions and a high cellular internalization rate. These gradients are sustained throughout the entire culture duration, leading to highly heterogeneous tissue growth; biochemical and histological measurements support that while biochemical content is enhanced up to 4-fold at the construct periphery, enhancements are entirely absent beyond 1 mm from the construct surface. Second, construct-encapsulated chondrocytes continuously secrete large amounts of endogenous TGF-β in its latent form, a portion of which undergoes cell-mediated activation and enhances biosynthesis uniformly throughout the tissue. Finally, motivated by these prior insights, we demonstrate that the alternative supplementation of additional exogenous latent TGF-β enhances biosynthesis uniformly throughout tissue constructs, leading to enhanced but homogeneous tissue growth. This novel demonstration suggests that latent TGF-β supplementation may be utilized as an important tool for the translational engineering of large cartilage constructs that will be required to repair the large osteoarthritic defects observed clinically. Copyright © 2015. Published by Elsevier Ltd.

Heterogeneous engineered cartilage growth results from gradients of media-supplemented active TGF-β and is ameliorated by the alternative supplementation of latent TGF-β

PubMed Central

Durney, Krista M.; Cigan, Alexander D.; Shim, Jay J.; Vunjak-Novakovic, Gordana; Hung, Clark T.; Ateshian, Gerard A.

2016-01-01

Transforming growth factor beta (TGF-β) has become one of the most widely utilized mediators of engineered cartilage growth. It is typically exogenously supplemented in the culture medium in its active form, with the expectation that it will readily transport into tissue constructs through passive diffusion and influence cellular biosynthesis uniformly. The results of this investigation advance three novel concepts regarding the role of TGF-β in cartilage tissue engineering that have important implications for tissue development. First, through the experimental and computational analysis of TGF-β concentration distributions, we demonstrate that, contrary to conventional expectations, media-supplemented exogenous active TGF-β exhibits a pronounced concentration gradient in tissue constructs, resulting from a combination of high-affinity binding interactions and a high cellular internalization rate. These gradients are sustained throughout the entire culture duration, leading to highly heterogeneous tissue growth; biochemical and histological measurements support that while biochemical content is enhanced up to 4-fold at the construct periphery, enhancements are entirely absent beyond 1 mm from the construct surface. Second, construct-encapsulated chondrocytes continuously secrete large amounts of endogenous TGF-β in its latent form, a portion of which undergoes cell-mediated activation and enhances biosynthesis uniformly throughout the tissue. Finally, motivated by these prior insights, we demonstrate that the alternative supplementation of additional exogenous latent TGF-β enhances biosynthesis uniformly throughout tissue constructs, leading to enhanced but homogeneous tissue growth. This novel demonstration suggests that latent TGF-β supplementation may be utilized as an important tool for the translational engineering of large cartilage constructs that will be required to repair the large osteoarthritic defects observed clinically. PMID:26599624

Effect of cigarette smoke on monocyte procoagulant activity: Focus on platelet-derived brain-derived neurotrophic factor (BDNF).

PubMed

Amadio, Patrizia; Baldassarre, Damiano; Sandrini, Leonardo; Weksler, Babette B; Tremoli, Elena; Barbieri, Silvia S

2017-01-01

Cigarette smoke (CS) activates platelets, promotes vascular dysfunction, and enhances Tissue Factor (TF) expression in blood monocytes favoring pro-thrombotic states. Brain-derived neurotrophic factor (BDNF), a member of the family of neurotrophins involved in survival, growth, and maturation of neurons, is released by activated platelets (APLTs) and plays a role in the cardiovascular system. The effect of CS on circulating levels of BDNF is controversial and the function of circulating BDNF in atherothrombosis is not fully understood. Here, we have shown that human platelets, treated with an aqueous extract of CS (CSE), released BDNF in a dose-dependent manner. In addition, incubation of human monocytes with BDNF or with the supernatant of platelets activated with CSE increased TF activity by a Tropomyosin receptor kinase B (TrkB)-dependent mechanism. Finally, comparing serum and plasma samples of 12 male never smokers (NS) and 29 male active smokers (AS) we observed a significant increase in microparticle-associated TF activity (MP-TF) as well as BDNF in AS, while in serum, BDNF behaved oppositely. Taken together these findings suggest that platelet-derived BDNF is involved in the regulation of TF activity and that CS plays a role in this pathway by favoring a pro-atherothrombotic state.

Potential application of metal nanoparticles for dosimetric systems: Concepts and perspectives

NASA Astrophysics Data System (ADS)

Guidelli, Eder José; Baffa, Oswaldo

2014-11-01

Metallic nanoparticles increase the delivered dose and consequently enhance tissue radio sensitization during radiation therapy of cancer. The Dose Enhancement Factor (DEF) corresponds to the ratio between the dose deposited on a tissue containing nanoparticles, and the dose deposited on a tissue without nanoparticles. In this sense, we have used electron spin resonance spectroscopy (ESR) to investigate how silver and gold nanoparticles affect the dose deposition in alanine dosimeters, which act as a surrogate of soft tissue. Besides optimizing radiation absorption by the dosimeter, the optical properties of these metal nanoparticles could also improve light emission from materials employed as radiation detectors. Therefore, we have also examined how the plasmonic properties of noble metal nanoparticles could enhance radiation detection using optically stimulated luminescence (OSL) dosimetry. This work will show results on how the use of gold and silver nanoparticles are beneficial for the ESR and OSL dosimetric techniques, and will describe the difficulties we have been facing, the challenges to overcome, and the perspectives.

Extracellular Vesicles Present in Human Ovarian Tumor Microenvironments Induce a Phosphatidylserine Dependent Arrest in the T Cell Signaling Cascade

PubMed Central

Kelleher, Raymond J.; Balu-Iyer, Sathy; Loyall, Jenni; Sacca, Anthony J.; Shenoy, Gautam N.; Peng, Peng; Iyer, Vandana; Fathallah, Anas M.; Berenson, Charles S.; Wallace, Paul K.; Tario, Joseph; Odunsi, Kunle; Bankert, Richard B.

2015-01-01

The identification of immunosuppressive factors within human tumor microenvironments, and the ability to block these factors, would be expected to enhance patients’ anti-tumor immune responses. We previously established that an unidentified factor, or factors, present in ovarian tumor ascites fluids reversibly inhibited the activation of T cells by arresting the T cell signaling cascade. Ultracentrifugation of the tumor ascites fluid has now revealed a pellet that contains small extracellular vesicles (EV) with an average diameter of 80nm. The T cell arrest was determined to be causally linked to phosphatidylserine (PS) that is present on the outer leaflet of the vesicle bilayer, as a depletion of PS expressing EV or a blockade of PS with anti-PS antibody significantly inhibits the vesicle induced signaling arrest. The inhibitory EV were also isolated from solid tumor tissues. The presence of immune suppressive vesicles in the microenvironments of ovarian tumors and our ability to block their inhibition of T cell function represent a potential therapeutic target for patients with ovarian cancer. PMID:26112921

The POU proteins Brn-2 and Oct-6 share important functions in Schwann cell development.

PubMed

Jaegle, Martine; Ghazvini, Mehrnaz; Mandemakers, Wim; Piirsoo, Marko; Driegen, Siska; Levavasseur, Francoise; Raghoenath, Smiriti; Grosveld, Frank; Meijer, Dies

2003-06-01

The genetic hierarchy that controls myelination of peripheral nerves by Schwann cells includes the POU domain Oct-6/Scip/Tst-1and the zinc-finger Krox-20/Egr2 transcription factors. These pivotal transcription factors act to control the onset of myelination during development and tissue regeneration in adults following damage. In this report we demonstrate the involvement of a third transcription factor, the POU domain factor Brn-2. We show that Schwann cells express Brn-2 in a developmental profile similar to that of Oct-6 and that Brn-2 gene activation does not depend on Oct-6. Overexpression of Brn-2 in Oct-6-deficient Schwann cells, under control of the Oct-6 Schwann cell enhancer (SCE), results in partial rescue of the developmental delay phenotype, whereas compound disruption of both Brn-2 and Oct-6 results in a much more severe phenotype. Together these data strongly indicate that Brn-2 function largely overlaps with that of Oct-6 in driving the transition from promyelinating to myelinating Schwann cells.

Human NRDRB1, an alternatively spliced isoform of NADP(H)-dependent retinol dehydrogenase/reductase enhanced enzymatic activity of benzil.

PubMed

Yan, Yinxia; Song, Xuhong; Liu, Gefei; Su, Zhongjing; Du, Yongming; Sui, Xuxia; Chang, Xiaolan; Huang, Dongyang

2012-01-01

Human NRDRB1, a 226 amino acid alternatively spliced isoform of the NADP(H)- dependent retinol dehydrogenase/reductase (NRDR), lacks the complete coding region of exon 3, but preserves all the important functional motifs for NRDR catalytic activity. Nevertheless, its tissue distribution and physiological function remain to be elucidated. Expression of NRDRB1 and NRDR in cells and tissues was analyzed by semi-quantitative polymerase chain reaction (PCR) and western blot. NRDRB1 was expressed as a His(6) fusion protein and subjected to kinetics assays. Recombinant NRDRB1 had 1.2 to 8.6 fold higher k(cat)/K(m) values than recombinant NRDR, depending on the substrate. NRDRB1 catalyzed the NADPH-dependent reduction of α-dicarbonyl compounds, such as isatin, 9,10-phenanthrenequinone, and especially benzil. The significantly high catalytic activity and the relatively high expression in human liver of NRDRB1 conferred cellular resistance to benzil-induced cell toxicity and over-expression of NRDRB1 in low expressing Ec109 cells significantly enhanced cell tolerance toward benzil. Based on its substrate specificity, catalytic activity and relatively high expression in human liver tissue, our results suggest that NRDRB1, an alternatively spliced isoform of NRDR in vivo functions better than NRDR as a dicarbonyl reductase for xenobiotics containing reactive carbonyls. Our study is the first reporting this phenomenon of the enzymes involved in biochemical reactions. Copyright © 2012 S. Karger AG, Basel.

Perfluorocarbon Enhanced Glasgow Oxygen Level Dependent (GOLD) Magnetic Resonance Metabolic Imaging Identifies the Penumbra Following Acute Ischemic Stroke

PubMed Central

Deuchar, Graeme A; Brennan, David; Holmes, William M; Shaw, Martin; Macrae, I Mhairi; Santosh, Celestine

2018-01-01

The ability to identify metabolically active and potentially salvageable ischaemic penumbra is crucial for improving treatment decisions in acute stroke patients. Our solution involves two complementary novel MRI techniques (Glasgow Oxygen Level Dependant (GOLD) Metabolic Imaging), which when combined with a perfluorocarbon (PFC) based oxygen carrier and hyperoxia can identify penumbra due to dynamic changes related to continued metabolism within this tissue compartment. Our aims were (i) to investigate whether PFC offers similar enhancement of the second technique (Lactate Change) as previously demonstrated for the T2*OC technique (ii) to demonstrate both GOLD metabolic imaging techniques working concurrently to identify penumbra, following administration of Oxycyte® (O-PFC) with hyperoxia. Methods: An established rat stroke model was utilised. Part-1: Following either saline or PFC, magnetic resonance spectroscopy was applied to investigate the effect of hyperoxia on lactate change in presumed penumbra. Part-2; rats received O-PFC prior to T2*OC (technique 1) and MR spectroscopic imaging, which was used to identify regions of tissue lactate change (technique 2) in response to hyperoxia. In order to validate the techniques, imaging was followed by [14C]2-deoxyglucose autoradiography to correlate tissue metabolic status to areas identified as penumbra. Results: Part-1: PFC+hyperoxia resulted in an enhanced reduction of lactate in the penumbra when compared to saline+hyperoxia. Part-2: Regions of brain tissue identified as potential penumbra by both GOLD metabolic imaging techniques utilising O-PFC, demonstrated maintained glucose metabolism as compared to adjacent core tissue. Conclusion: For the first time in vivo, enhancement of both GOLD metabolic imaging techniques has been demonstrated following intravenous O-PFC+hyperoxia to identify ischaemic penumbra. We have also presented preliminary evidence of the potential therapeutic benefit offered by O-PFC. These unique theranostic applications would enable treatment based on metabolic status of the brain tissue, independent of time from stroke onset, leading to increased uptake and safer use of currently available treatment options. PMID:29556351

Electrical Polarization of Titanium Surfaces for the Enhancement of Osteoblast Differentiation

PubMed Central

Gittens, Rolando A.; Olivares-Navarrete, Rene; Rettew, Robert; Butera, Robert J.; Alamgir, Faisal M.; Boyan, Barbara D.; Schwartz, Zvi

2014-01-01

Electrical stimulation has been used clinically to promote bone regeneration in cases of fractures with delayed union or nonunion, with several in vitro and in vivo reports suggesting its beneficial effects on bone formation. However, the use of electrical stimulation of titanium (Ti) implants to enhance osseointegration is less understood, in part because of the few in vitro models that attempt to represent the in vivo environment. In this article, the design of a new in vitro system that allows direct electrical stimulation of osteoblasts through their Ti substrates without the flow of exogenous currents through the media is presented, and the effect of applied electrical polarization on osteoblast differentiation and local factor production was evaluated. A custom-made polycarbonate tissue culture plate was designed to allow electrical connections directly underneath Ti disks placed inside the wells, which were supplied with electrical polarization ranging from 100 to 500 mV to stimulate MG63 osteoblasts. Our results show that electrical polarization applied directly through Ti substrates on which the cells are growing in the absence of applied electrical currents may increase osteoblast differentiation and local factor production in a voltage-dependent manner. PMID:23996899

Gastric Inhibitory Peptide Controls Adipose Insulin Sensitivity via Activation of cAMP-response Element-binding Protein and p110β Isoform of Phosphatidylinositol 3-Kinase*

PubMed Central

Mohammad, Sameer; Ramos, Lavoisier S.; Buck, Jochen; Levin, Lonny R.; Rubino, Francesco; McGraw, Timothy E.

2011-01-01

Gastric inhibitory peptide (GIP) is an incretin hormone secreted in response to food intake. The best known function of GIP is to enhance glucose-dependent insulin secretion from pancreatic β-cells. Extra-pancreatic effects of GIP primarily occur in adipose tissues. Here, we demonstrate that GIP increases insulin-dependent translocation of the Glut4 glucose transporter to the plasma membrane and exclusion of FoxO1 transcription factor from the nucleus in adipocytes, establishing that GIP has a general effect on insulin action in adipocytes. Stimulation of adipocytes with GIP alone has no effect on these processes. Using pharmacologic and molecular genetic approaches, we show that the effect of GIP on adipocyte insulin sensitivity requires activation of both the cAMP/protein kinase A/CREB signaling module and p110β phosphoinositol-3′ kinase, establishing a novel signal transduction pathway modulating insulin action in adipocytes. This insulin-sensitizing effect is specific for GIP because isoproterenol, which elevates adipocyte cAMP and activates PKA/CREB signaling, does not affect adipocyte insulin sensitivity. The insulin-sensitizing activity points to a more central role for GIP in intestinal regulation of peripheral tissue metabolism, an emerging feature of inter-organ communication in the control of metabolism. PMID:22027830

Peri-implant esthetics assessment and management

PubMed Central

Balasubramaniam, Aarthi S.; Raja, Sunitha V.; Thomas, Libby John

2013-01-01

Providing an esthetic restoration in the anterior region of the mouth has been the basis of peri-implant esthetics. To achieve optimal esthetics, in implant supported restorations, various patient and tooth related factors have to be taken into consideration. Peri-implant plastic surgery has been adopted to improve the soft tissue and hard tissue profiles, during and after implant placement. The various factors and the procedures related to enhancement of peri-implant esthetics have been discussed in this review article. PMID:23878557

Tissue-engineered cartilage: the crossroads of biomaterials, cells and stimulating factors.

PubMed

Bhardwaj, Nandana; Devi, Dipali; Mandal, Biman B

2015-02-01

Damage to cartilage represents one of the most challenging tasks of musculoskeletal therapeutics due to its limited propensity for healing and regenerative capabilities. Lack of current treatments to restore cartilage tissue function has prompted research in this rapidly emerging field of tissue regeneration of functional cartilage tissue substitutes. The development of cartilaginous tissue largely depends on the combination of appropriate biomaterials, cell source, and stimulating factors. Over the years, various biomaterials have been utilized for cartilage repair, but outcomes are far from achieving native cartilage architecture and function. This highlights the need for exploration of suitable biomaterials and stimulating factors for cartilage regeneration. With these perspectives, we aim to present an overview of cartilage tissue engineering with recent progress, development, and major steps taken toward the generation of functional cartilage tissue. In this review, we have discussed the advances and problems in tissue engineering of cartilage with strong emphasis on the utilization of natural polymeric biomaterials, various cell sources, and stimulating factors such as biophysical stimuli, mechanical stimuli, dynamic culture, and growth factors used so far in cartilage regeneration. Finally, we have focused on clinical trials, recent innovations, and future prospects related to cartilage engineering. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Re-engineering the Pancreas Tumor Microenvironment: A “Regenerative Program” Hacked

PubMed Central

Evan, Gerard I.; Hah, Nasun; Littlewood, Trevor D.; Sodir, Nicole M.; Vidal, Tania Campos; Downes, Michael; Evans, Ronald M.

2017-01-01

The “hallmarks” of pancreatic ductal adenocarcinoma (PDAC) include proliferative, invasive and metastatic tumor cells and an associated dense desmoplasia comprised of fibroblasts, pancreatic stellate cells, extracellular matrix and immune cells. The oncogenically-activated pancreatic epithelium and its associated stroma are obligatorily interdependent, with the resulting inflammatory and immune-suppressive microenvironment contributing greatly to the evolution and maintenance of PDAC. The peculiar pancreas-specific tumor phenotype is a consequence of oncogenes hacking the resident pancreas regenerative program, a tissue specific repair mechanism regulated by discrete super enhancer networks. Defined as genomic regions containing clusters of multiple enhancers, super enhancers play pivotal roles in cell/tissue specification, identity and maintenance. Hence, interfering with such super enhancer driven repair networks should exert a disproportionately disruptive effect on tumor versus normal pancreatic tissue. Novel drugs that directly or indirectly inhibit processes regulating epigenetic status and integrity, including those driven by histone deacetylases, histone methyltransferase and hydroxylases, DNA methyltransferases, various metabolic enzymes, and bromodomain and extra-terminal motif proteins (BETs) have shown the feasibility of disrupting super enhancer-dependent transcription in treating multiple tumor types, including PDAC. The idea that pancreatic adenocarcinomas rely on embedded super enhancer transcriptional mechanism suggests a vulnerability that can be potentially targeted as novel therapies for this intractable disease. PMID:28373363

Host‐related factors explaining interindividual variability of carotenoid bioavailability and tissue concentrations in humans

PubMed Central

Desmarchelier, Charles; Dragsted, Lars O.; Nielsen, Charlotte S.; Stahl, Wilhelm; Rühl, Ralph; Keijer, Jaap; Borel, Patrick

2017-01-01

Carotenoid dietary intake and their endogenous levels have been associated with a decreased risk of several chronic diseases. There are indications that carotenoid bioavailability depends, in addition to the food matrix, on host factors. These include diseases (e.g. colitis), life‐style habits (e.g. smoking), gender and age, as well as genetic variations including single nucleotide polymorphisms that govern carotenoid metabolism. These are expected to explain interindividual differences that contribute to carotenoid uptake, distribution, metabolism and excretion, and therefore possibly also their association with disease risk. For instance, digestion enzymes fostering micellization (PNLIP, CES), expression of uptake/efflux transporters (SR‐BI, CD36, NPC1L1), cleavage enzymes (BCO1/2), intracellular transporters (FABP2), secretion into chylomicrons (APOB, MTTP), carotenoid metabolism in the blood and liver (LPL, APO C/E, LDLR), and distribution to target tissues such as adipose tissue or macula (GSTP1, StARD3) depend on the activity of these proteins. In addition, human microbiota, e.g. via altering bile‐acid concentrations, may play a role in carotenoid bioavailability. In order to comprehend individual, variable responses to these compounds, an improved knowledge on intra‐/interindividual factors determining carotenoid bioavailability, including tissue distribution, is required. Here, we highlight the current knowledge on factors that may explain such intra‐/interindividual differences. PMID:28101967

Hemoglobin enhances tissue factor expression on human malignant cells.

PubMed

Siddiqui, F A; Amirkhosravi, A; Amaya, M; Meyer, T; Biggerstaff, J; Desai, H; Francis, J L

2001-04-01

Tissue Factor (TF) is a transmembrane glycoprotein that complexes with factor VII/activated factor VII to initiate blood coagulation. TF may be expressed on the surface of various cells including monocytes and endothelial cells. Over-expression of TF in human tumor cell lines promotes metastasis. We recently showed that hemoglobin (Hb) forms a specific complex with TF purified from human malignant melanoma cells and enhances its procoagulant activity (PCA). To further study this interaction, we examined the effect of Hb on the expression of TF on human malignant (TF+) cells and KG1 myeloid leukemia (TF-) cells. Human melanoma A375 and J82 bladder carcinoma cells, which express TF at moderate and relatively high levels, respectively, were incubated with varying concentrations (0-1.5 mg/ml) of Hb. After washing, cells were analyzed for Hb binding and TF expression using flow cytometry and confocal microscopy. Hb bound to the cells in a concentration-dependent manner, and increased both TF expression and PCA. The human A375 malignant melanoma cells incubated with Hb (1 mg/ml) expressed up to six times more TF antigen than cells without Hb. This increase in TF expression and PCA of intact cells incubated with Hb was significantly inhibited by cycloheximide at a concentration of 10 microg/ml (P < 0.01). An increase in total cellular TF antigen content was demonstrated by specific immunoassay. In contrast, Hb (5 mg/ml) did not induce TF expression and PCA on KG1 cells as determined by flow cytometry and TF (FXAA) activity. We conclude that Hb specifically binds to TF-bearing malignant cells and increases their PCA. This effect seems to be at least partly due to de novo synthesis of TF and increased surface expression. However, the exact mechanism by which Hb binds and upregulates TF expression remains to be determined.

Stromal Tissue Rigidity Promotes Mesenchymal Stem Cell-Mediated Corneal Wound Healing Through the Transforming Growth Factor β Signaling Pathway.

PubMed

Yang, Yun-Hsiang; Hsieh, Ting-Lieh; Ji, Andrea Tung-Qian; Hsu, Wei-Tse; Liu, Chia-Yu; Lee, Oscar Kuang-Sheng; Ho, Jennifer Hui-Chun

2016-10-01

The healing of a corneal epithelial defect is essential for preventing infectious corneal ulcers and subsequent blindness. We previously demonstrated that mesenchymal stem cells (MSCs) in the corneal stroma, through a paracrine mechanism, yield a more favorable therapeutic benefit for corneal wound re-epithelialization than do MSCs in the corneal epithelium. In this study, MSCs were grown on a matrix with the rigidity of the physiological human vitreous (1 kPa), corneal epithelium (8 kPa), or corneal stroma (25 kPa) for investigating the role of corneal tissue rigidity in MSC functions regarding re-epithelialization promotion. MSC growth on a 25-kPa dish significantly promoted the wound healing of human corneal epithelial (HCE-T) cells. Among growth factors contributing to corneal epithelial wound healing, corneal stromal rigidity selectively enhanced transforming growth factor-beta (TGF-β) secretion from MSCs. Inhibitors of TGF-β pan receptor, TGF-β receptor 1, and Smad2 dose dependently abrogated MSC-mediated HCE-T wound healing. Furthermore, MSCs growth on a matrix with corneal stromal rigidity enhanced the ability of themselves to promote corneal re-epithelialization by activating matrix metalloproteinase (MMP) expression and integrin β1 production in HCE-T cells through TGF-β signaling pathway activation. Smad2 activation resulted in the upregulation of MMP-2 and -13 expression in HCE-T cells, whereas integrin β1 production favored a Smad2-independent TGF-β pathway. Altogether, we conclude that corneal stromal rigidity is a critical factor for MSC-induced promotion of corneal re-epithelialization. The activation of the TGF-β signaling pathway, which maintains the balance between integrin and MMP expression, in HCE-T cells is the major pathway responsible for MSC-mediated wound healing. Stem Cells 2016;34:2525-2535. © 2016 AlphaMed Press.

Peroxisome Proliferator Activated Receptor-α/Hypoxia Inducible Factor-1α Interplay Sustains Carbonic Anhydrase IX and Apoliprotein E Expression in Breast Cancer Stem Cells

PubMed Central

Papi, Alessio; Storci, Gianluca; Guarnieri, Tiziana; De Carolis, Sabrina; Bertoni, Sara; Avenia, Nicola; Sanguinetti, Alessandro; Sidoni, Angelo; Santini, Donatella; Ceccarelli, Claudio; Taffurelli, Mario; Orlandi, Marina; Bonafé, Massimiliano

2013-01-01

Aims Cancer stem cell biology is tightly connected to the regulation of the pro-inflammatory cytokine network. The concept of cancer stem cells “inflammatory addiction” leads to envisage the potential role of anti-inflammatory molecules as new anti-cancer targets. Here we report on the relationship between nuclear receptors activity and the modulation of the pro-inflammatory phenotype in breast cancer stem cells. Methods Breast cancer stem cells were expanded as mammospheres from normal and tumor human breast tissues and from tumorigenic (MCF7) and non tumorigenic (MCF10) human breast cell lines. Mammospheres were exposed to the supernatant of breast tumor and normal mammary gland tissue fibroblasts. Results In mammospheres exposed to the breast tumor fibroblasts supernatant, autocrine tumor necrosis factor-α signalling engenders the functional interplay between peroxisome proliferator activated receptor-α and hypoxia inducible factor-1α (PPARα/HIF1α). The two proteins promote mammospheres formation and enhance each other expression via miRNA130b/miRNA17-5p-dependent mechanism which is antagonized by PPARγ. Further, the PPARα/HIF1α interplay regulates the expression of the pro-inflammatory cytokine interleukin-6, the hypoxia survival factor carbonic anhydrase IX and the plasma lipid carrier apolipoprotein E. Conclusion Our data demonstrate the importance of exploring the role of nuclear receptors (PPARα/PPARγ) in the regulation of pro-inflammatory pathways, with the aim to thwart breast cancer stem cells functioning. PMID:23372804

Cholesteryl group- and acryloyl group-bearing pullulan nanogel to deliver BMP2 and FGF18 for bone tissue engineering.

PubMed

Fujioka-Kobayashi, Masako; Ota, Masato S; Shimoda, Asako; Nakahama, Ken-ichi; Akiyoshi, Kazunari; Miyamoto, Youji; Iseki, Sachiko

2012-10-01

To create a drug delivery system that allows the controlled release of proteins, such as growth factors, over a long-term period, cholesteryl group- and acryloyl group-bearing pullulan (CHPOA) nanogels were aggregated to form fast-degradable hydrogels (CHPOA/hydrogels) by cross-linking with thiol-bearing polyethylene glycol. The gold standard of clinical bone reconstruction therapy with a physiologically active material is treatment with recombinant human bone morphogenetic protein 2 (BMP2); however, this approach has limitations, such as inflammation, poor cost-efficiency, and varying interindividual susceptibility. In this study, two distinct growth factors, BMP2 and recombinant human fibroblast growth factor 18 (FGF18), were applied to a critical-size skull bone defect for bone repair by the CHPOA/hydrogel system. The CHPOA-FGF18/hydrogel displayed identical results to the control CHPOA-PBS/hydrogel, and the CHPOA-BMP2/hydrogel treatment imperfectly induced bone repair. By contrast, the CHPOA-FGF18 + BMP2/hydrogel treatment strongly enhanced and stabilized the BMP2-dependent bone repair, inducing osteoprogenitor cell infiltration inside and around the hydrogel. This report indicates that the CHPOA/hydrogel system can successfully deliver two different proteins to the bone defect to induce effective bone repair. The combination of the CHPOA/hydrogel system with the growth factors FGF18 and BMP2 might be a step towards efficient bone tissue engineering. Copyright © 2012 Elsevier Ltd. All rights reserved.

Cocoa-enriched diet enhances antioxidant enzyme activity and modulates lymphocyte composition in thymus from young rats.

PubMed

Ramiro-Puig, Emma; Urpí-Sardà, Mireia; Pérez-Cano, Francisco J; Franch, Angels; Castellote, Cristina; Andrés-Lacueva, Cristina; Izquierdo-Pulido, Maria; Castell, Margarida

2007-08-08

Cocoa is a rich source of flavonoids, mainly (-)-epicatechin, (+)-catechin, and procyanidins. This article reports the effect of continuous cocoa intake on antioxidant capacity in plasma and tissues, including lymphoid organs and liver, from young rats. Weaned Wistar rats received natural cocoa (4% or 10% food intake) for three weeks, corresponding to their infancy. Flavonoid absorption was confirmed through the quantification of epicatechin metabolites in urine. Total antioxidant capacity (TAC) and the activity of antioxidant enzymes, superoxide dismutase (SOD) and catalase, were examined. Cocoa intake enhanced TAC in all tissues especially in thymus. Moreover, thymus SOD and catalase activities were also dose-dependently increased by cocoa. It was also analyzed whether the enhanced antioxidant system in thymus could influence its cellular composition. An increase in the percentage of thymocytes in advanced development stage was found. In summary, cocoa diet enhances thymus antioxidant defenses and influences thymocyte differentiation.

Biomimetic hybrid porous scaffolds immobilized with platelet derived growth factor-BB promote cellularization and vascularization in tissue engineering.

PubMed

Murali, Ragothaman; Ponrasu, Thangavel; Cheirmadurai, Kalirajan; Thanikaivelan, Palanisamy

2016-02-01

Development of hybrid scaffolds with synergistic combination of growth factor is a promising approach to promote early in vivo wound repair and tissue regeneration. Here, we show the rapid wound healing in Wistar albino rats using biomimetic collagen-poly(dialdehyde) guar gum based hybrid porous scaffolds covalently immobilized with platelet derived growth factor-BB. The immobilized platelet derived growth factor in the hybrid scaffolds not only enhance the total protein, collagen, hexosamine, and uronic acid contents in the granulation tissue but also provide stronger tissues. The wound closure analysis reveal that the complete epithelialization period is 15.4 ± 0.9 days for collagen-poly(dialdehyde) guar gum-platelet derived growth factor hybrid scaffolds, whereas it is significantly higher for control, collagen, collagen- poly(dialdehyde) guar gum and povidine-iodine treated groups. Further, the histological evaluation shows that the immobilized platelet derived growth factor in the hybrid scaffolds induced a more robust cellular and vascular response in the implanted site. Hence, we demonstrate that the collagen-poly(dialdehyde) guar gum hybrid scaffolds loaded with platelet derived growth factor stimulates chemotactic effects in the implanted site to promote rapid tissue regeneration and wound repair without the assistance of antibacterial agents. © 2015 Wiley Periodicals, Inc.

Arabidopsis JACKDAW and MAGPIE zinc finger proteins delimit asymmetric cell division and stabilize tissue boundaries by restricting SHORT-ROOT action

PubMed Central

Welch, David; Hassan, Hala; Blilou, Ikram; Immink, Richard; Heidstra, Renze; Scheres, Ben

2007-01-01

In the Arabidopsis root, the SHORT-ROOT transcription factor moves outward to the ground tissue from its site of transcription in the stele and is required for the specification of the endodermis and the stem cell organizing quiescent center cells. In addition, SHORT-ROOT and the downstream transcription factor SCARECROW control an oriented cell division in ground tissue stem cell daughters. Here, we show that the JACKDAW and MAGPIE genes, which encode members of a plant-specific family of zinc finger proteins, act in a SHR-dependent feed-forward loop to regulate the range of action of SHORT-ROOT and SCARECROW. JACKDAW expression is initiated independent of SHORT-ROOT and regulates the SCARECROW expression domain outside the stele, while MAGPIE expression depends on SHORT-ROOT and SCARECROW. We provide evidence that JACKDAW and MAGPIE regulate tissue boundaries and asymmetric cell division and can control SHORT-ROOT and SCARECROW activity in a transcriptional and protein interaction network. PMID:17785527

Recent advances in gene-enhanced bone tissue engineering.

PubMed

Betz, Volker M; Kochanek, Stefan; Rammelt, Stefan; Müller, Peter E; Betz, Oliver B; Messmer, Carolin

2018-03-30

The loss of bone tissue represents a critical clinical condition that is frequently faced by surgeons. Substantial progress has been made in the area of bone research, providing insight into the biology of bone under physiological and pathological conditions, as well as tools for the stimulation of bone regeneration. The present review discusses recent advances in the field of gene-enhanced bone tissue engineering. Gene transfer strategies have emerged as highly effective tissue engineering approaches for supporting the repair of the musculoskeletal system. By contrast to treatment with recombinant proteins, genetically engineered cells can release growth factors at the site of injury over extended periods of time. Of particular interest are the expedited technologies that can be applied during a single surgical procedure in a cost-effective manner, allowing translation from bench to bedside. Several promising methods based on the intra-operative genetic manipulation of autologous cells or tissue fragments have been developed in preclinical studies. Moreover, gene therapy for bone regeneration has entered the clinical stage with clinical trials for the repair of alveolar bone. Current trends in gene-enhanced bone engineering are also discussed with respect to the movement of the field towards expedited, translational approaches. It is possible that gene-enhanced bone tissue engineering will become a clinical reality within the next few years. Copyright © 2018 John Wiley & Sons, Ltd.

Enhanced somatic embryogenesis in Theobroma cacao using the homologous BABY BOOM transcription factor.

PubMed

Florez, Sergio L; Erwin, Rachel L; Maximova, Siela N; Guiltinan, Mark J; Curtis, Wayne R

2015-05-16

Theobroma cacao, the chocolate tree, is an important economic crop in East Africa, South East Asia, and South and Central America. Propagation of elite varieties has been achieved through somatic embryogenesis (SE) but low efficiencies and genotype dependence still presents a significant limitation for its propagation at commercial scales. Manipulation of transcription factors has been used to enhance the formation of SEs in several other plant species. This work describes the use of the transcription factor Baby Boom (BBM) to promote the transition of somatic cacao cells from the vegetative to embryonic state. An ortholog of the Arabidopsis thaliana BBM gene (AtBBM) was characterized in T. cacao (TcBBM). TcBBM expression was observed throughout embryo development and was expressed at higher levels during SE as compared to zygotic embryogenesis (ZE). TcBBM overexpression in A. thaliana and T. cacao led to phenotypes associated with SE that did not require exogenous hormones. While transient ectopic expression of TcBBM provided only moderate enhancements in embryogenic potential, constitutive overexpression dramatically increased SE proliferation but also appeared to inhibit subsequent development. Our work provides validation that TcBBM is an ortholog to AtBBM and has a specific role in both somatic and zygotic embryogenesis. Furthermore, our studies revealed that TcBBM transcript levels could serve as a biomarker for embryogenesis in cacao tissue. Results from transient expression of TcBBM provide confirmation that transcription factors can be used to enhance SE without compromising plant development and avoiding GMO plant production. This strategy could compliment a hormone-based method of reprogramming somatic cells and lead to more precise manipulation of SE at the regulatory level of transcription factors. The technology would benefit the propagation of elite varieties with low regeneration potential as well as the production of transgenic plants, which similarly requires somatic cell reprogramming.

Adipose Tissue Angiogenesis: Impact on Obesity and Type-2 Diabetes

PubMed Central

Corvera, Silvia; Gealekman, Olga

2013-01-01

The growth and function of tissues is critically dependent on their vascularization. Adipose tissue is capable of expanding many-fold during adulthood, therefore requiring the formation of new vasculature to supply growing and proliferating adipocytes. The expansion of the vasculature in adipose tissue occurs through angiogenesis, where new blood vessels develop from those pre-existing within the tissue. Inappropriate angiogenesis may underlie adipose tissue dysfunction in obesity, which in turn increases type-2 diabetes risk. In addition, genetic and developmental factors involved in vascular patterning may define the size and expandability of diverse adipose tissue depots, which are also associated with type-2 diabetes risk. Moreover, the adipose tissue vasculature appears to be the niche for pre-adipocyte precursors, and factors that affect angiogenesis may directly impact the generation of new adipocytes. Here we review recent advances on the basic mechanisms of angiogenesis, and on the role of angiogenesis in adipose tissue development and obesity. A substantial amount of data point to a deficit in adipose tissue angiogenesis as a contributing factor to insulin resistance and metabolic disease in obesity. These emerging findings support the concept of the adipose tissue vasculature as a source of new targets for metabolic disease therapies. PMID:23770388

Rat Strains Bred for Low and High Aerobic Running Capacity do not Differ in Their Survival Time to Hemorrhage

DTIC Science & Technology

2009-12-01

23. Ensunsa JL, Symons JD, Lanoue L, Schrader HR, Keen CL. Reducing arginase activity via dietary manganese deficiency enhances endothelium- dependent ...maximal oxygen consumption (Vo2max) was 12% greater during normoxia and 20% greater during hypoxia in the HCR vs LCR (12). This enhanced Vo2max...was ∼ 40% greater in HCR vs LCR due to both increased oxygen delivery-- resulting from an enhanced stroke volume-- and oxygen tissue transfer (13

Heparanase enhances the generation of activated factor X in the presence of tissue factor and activated factor VII.

PubMed

Nadir, Yona; Brenner, Benjamin; Fux, Liat; Shafat, Itay; Attias, Judith; Vlodavsky, Israel

2010-11-01

Heparanase is an endo-β-D-glucuronidase dominantly involved in tumor metastasis and angiogenesis. Recently, we demonstrated that heparanase is involved in the regulation of the hemostatic system. Our hypothesis was that heparanase is directly involved in activation of the coagulation cascade. Activated factor X and thrombin were studied using chromogenic assays, immunoblotting and thromboelastography. Heparanase levels were measured by enzyme-linked immunosorbent assay. A potential direct interaction between tissue factor and heparanase was studied by co-immunoprecipitation and far-western assays. Interestingly, addition of heparanase to tissue factor and activated factor VII resulted in a 3- to 4-fold increase in activation of the coagulation cascade as shown by increased activated factor X and thrombin production. Culture medium of human embryonic kidney 293 cells over-expressing heparanase and its derivatives increased activated factor X levels in a non-enzymatic manner. When heparanase was added to pooled normal plasma, a 7- to 8-fold increase in activated factor X level was observed. Subsequently, we searched for clinical data supporting this newly identified role of heparanase. Plasma samples from 35 patients with acute leukemia at presentation and 20 healthy donors were studied for heparanase and activated factor X levels. A strong positive correlation was found between plasma heparanase and activated factor X levels (r=0.735, P=0.001). Unfractionated heparin and an inhibitor of activated factor X abolished the effect of heparanase, while tissue factor pathway inhibitor and tissue factor pathway inhibitor-2 only attenuated the procoagulant effect. Using co-immunoprecipitation and far-western analyses it was shown that heparanase interacts directly with tissue factor. Overall, our results support the notion that heparanase is a potential modulator of blood hemostasis, and suggest a novel mechanism by which heparanase increases the generation of activated factor X in the presence of tissue factor and activated factor VII.

Sonodynamic therapy--a review of the synergistic effects of drugs and ultrasound.

PubMed

Rosenthal, Ionel; Sostaric, Joe Z; Riesz, Peter

2004-09-01

Sonodynamic therapy, the ultrasound dependent enhancement of cytotoxic activities of certain compounds (sonosensitizers) in studies with cells in vitro and in tumor bearing animals, is reviewed. The attractive features of this modality for cancer treatment emerges from the ability to focus the ultrasound energy on malignancy sites buried deep in tissues and to locally activate a preloaded sonosensitizer. Possible mechanisms of sonodynamic therapy include generation of sonosensitizer derived radicals which initiate chain peroxidation of membrane lipids via peroxyl and/or alkoxyl radicals, the physical destabilization of the cell membrane by the sonosensitizer thereby rendering the cell more susceptible to shear forces or ultrasound enhanced drug transport across the cell membrane (sonoporation). Evidence against the role of singlet oxygen in sonodynamic therapy is discussed. The mechanism of sonodynamic therapy is probably not governed by a universal mechanism, but may be influenced by multiple factors including the nature of the biological model, the sonosensitizer and the ultrasound parameters. The current review emphasizes the effect of ultrasound induced free radicals in sonodynamic therapy.

IGF-1-induced MMP-11 expression promotes the proliferation and invasion of gastric cancer cells through the JAK1/STAT3 signaling pathway.

PubMed

Su, Chao; Wang, Wenchang; Wang, Cunchuan

2018-05-01

The present study aimed to investigate the association between insulin-like growth factor-1 (IGF-1) and matrix metalloproteinase-11 (MMP-11) expression in gastric cancer (GC) and the underlying mechanisms in SGC-7901 cells. Reverse transcription-quantitative polymerase chain reaction analysis revealed that the expression of IGF-1 and MMP-11 was significantly upregulated in GC tissues compared with normal gastric tissue. Furthermore, IGF-1 significantly and dose-dependently promoted MMP-11. Western blotting revealed that the addition of IGF-1 to SGC-7901 cells led to an evident enhancement in signal transducer and activator of transcription 3 (STAT3), IGF-1R and Janus kinase 1 (JAK1) phosphorylation at 20 and 40 min. A decrease in the extent of the elevated expression of MMP-11 and the enhanced phosphorylation of STAT3, JAK1 and IGF-1 receptor (IGF-1R) induced by IGF-1 in SGC-7901 cells were observed following treatment with NT157 (an IGF-1R inhibitor). Furthermore, piceatannol (a JAK1 inhibitor) or small interfering RNA against STAT3 reduced the extent of the increased expression of MMP-11 induced by IGF-1 in SGC-7901 cells. Piceatannol treatment induced the dose-dependent decline in the enhancement of STAT3 phosphorylation induced by IGF-1, indicating that the JAK1/STAT3 pathway may be implicated in the elevated expression of MMP-11 induced by IGF-1 in SGC-7901 cells. Finally, IGF-1 treatment significantly promoted the proliferation and invasion of SGC-7901 cells, which was inhibited following NT157, piceatannol or si-STAT3 treatment. The present study therefore demonstrated that IGF-1-induced MMP-11 may have facilitated the proliferation and invasion of SGC-7901 cells via the JAK1/STAT3 pathway.

Collagen-binding VEGF mimetic peptide: Structure, matrix interaction, and endothelial cell activation

NASA Astrophysics Data System (ADS)

Chan, Tania R.

Long term survival of artificial tissue constructs depends greatly on proper vascularization. In nature, differentiation of endothelial cells and formation of vasculature are directed by dynamic spatio-temporal cues in the extracellular matrix that are difficult to reproduce in vitro. In this dissertation, we present a novel bifunctional peptide that mimics matrix-bound vascular endothelial growth factor (VEGF), which can be used to encode spatially controlled angiogenic signals in collagen-based scaffolds. The peptide, QKCMP, contains a collagen mimetic domain (CMP) that binds to type I collagen by a unique triple helix hybridization mechanism and a VEGF mimetic domain (QK) with pro-angiogenic activity. We demonstrate QKCMP's ability to hybridize with native and heat denatured collagens through a series of binding studies on collagen and gelatin substrates. Circular dichroism experiments show that the peptide retains the triple helical structure vital for collagen binding, and surface plasmon resonance study confirms the molecular interaction between the peptide and collagen strands. Cell culture studies demonstrate QKCMP's ability to induce endothelial cell morphogenesis and network formation as a matrix-bound factor in 2D and 3D collagen scaffolds. We also show that the peptide can be used to spatially modify collagen-based substrates to promote localized endothelial cell activation and network formation. To probe the biological events that govern these angiogenic cellular responses, we investigated the cell signaling pathways activated by collagen-bound QKCMP and determined short and long-term endothelial cell response profiles for p38, ERK1/2, and Akt signal transduction cascades. Finally, we present our efforts to translate the peptide's in vitro bioactivity to an in vivo burn injury animal model. When implanted at the wound site, QKCMP functionalized biodegradable hydrogels induce enhanced neovascularization in the granulation tissue. The results show QKCMP's efficacy as a matrix-bound angiogenic factor that directs endothelial cell proliferation and migration. These findings suggest that QKCMP can be used to enhance microvasculature formation during wound healing as well as to promote spatially controlled microvasculature for tissue engineering applications.

Action Mechanism of Fibroblast Growth Factor-2 (FGF-2) in the Promotion of Periodontal Regeneration in Beagle Dogs

PubMed Central

Nagayasu-Tanaka, Toshie; Anzai, Jun; Takaki, Shu; Shiraishi, Noriko; Terashima, Akio; Asano, Taiji; Nozaki, Takenori; Kitamura, Masahiro; Murakami, Shinya

2015-01-01

Fibroblast growth factor-2 (FGF-2) enhances the formation of new alveolar bone, cementum, and periodontal ligament (PDL) in periodontal defect models. However, the mechanism through which FGF-2 acts in periodontal regeneration in vivo has not been fully clarified yet. To reveal the action mechanism, the formation of regenerated tissue and gene expression at the early phase were analyzed in a beagle dog 3-wall periodontal defect model. FGF-2 (0.3%) or the vehicle (hydroxypropyl cellulose) only were topically applied to the defect in FGF-2 and control groups, respectively. Then, the amount of regenerated tissues and the number of proliferating cells at 3, 7, 14, and 28 days and the number of blood vessels at 7 days were quantitated histologically. Additionally, the expression of osteogenic genes in the regenerated tissue was evaluated by real-time PCR at 7 and 14 days. Compared with the control, cell proliferation around the existing bone and PDL, connective tissue formation on the root surface, and new bone formation in the defect at 7 days were significantly promoted by FGF-2. Additionally, the number of blood vessels at 7 days was increased by FGF-2 treatment. At 28 days, new cementum and PDL were extended by FGF-2. Moreover, FGF-2 increased the expression of bone morphogenetic protein 2 (BMP-2) and osteoblast differentiation markers (osterix, alkaline phosphatase, and osteocalcin) in the regenerated tissue. We revealed the facilitatory mechanisms of FGF-2 in periodontal regeneration in vivo. First, the proliferation of fibroblastic cells derived from bone marrow and PDL was accelerated and enhanced by FGF-2. Second, angiogenesis was enhanced by FGF-2 treatment. Finally, osteoblastic differentiation and bone formation, at least in part due to BMP-2 production, were rapidly induced by FGF-2. Therefore, these multifaceted effects of FGF-2 promote new tissue formation at the early regeneration phase, leading to enhanced formation of new bone, cementum, and PDL. PMID:26120833

A method for converting dose-to-medium to dose-to-tissue in Monte Carlo studies of gold nanoparticle-enhanced radiotherapy

NASA Astrophysics Data System (ADS)

Koger, B.; Kirkby, C.

2016-03-01

Gold nanoparticles (GNPs) have shown potential in recent years as a means of therapeutic dose enhancement in radiation therapy. However, a major challenge in moving towards clinical implementation is the exact characterisation of the dose enhancement they provide. Monte Carlo studies attempt to explore this property, but they often face computational limitations when examining macroscopic scenarios. In this study, a method of converting dose from macroscopic simulations, where the medium is defined as a mixture containing both gold and tissue components, to a mean dose-to-tissue on a microscopic scale was established. Monte Carlo simulations were run for both explicitly-modeled GNPs in tissue and a homogeneous mixture of tissue and gold. A dose ratio was obtained for the conversion of dose scored in a mixture medium to dose-to-tissue in each case. Dose ratios varied from 0.69 to 1.04 for photon sources and 0.97 to 1.03 for electron sources. The dose ratio is highly dependent on the source energy as well as GNP diameter and concentration, though this effect is less pronounced for electron sources. By appropriately weighting the monoenergetic dose ratios obtained, the dose ratio for any arbitrary spectrum can be determined. This allows complex scenarios to be modeled accurately without explicitly simulating each individual GNP.

Subchondral pre-solidified chitosan/blood implants elicit reproducible early osteochondral wound-repair responses including neutrophil and stromal cell chemotaxis, bone resorption and repair, enhanced repair tissue integration and delayed matrix deposition

PubMed Central

2013-01-01

Background In this study we evaluated a novel approach to guide the bone marrow-driven articular cartilage repair response in skeletally aged rabbits. We hypothesized that dispersed chitosan particles implanted close to the bone marrow degrade in situ in a molecular mass-dependent manner, and attract more stromal cells to the site in aged rabbits compared to the blood clot in untreated controls. Methods Three microdrill hole defects, 1.4 mm diameter and 2 mm deep, were created in both knee trochlea of 30 month-old New Zealand White rabbits. Each of 3 isotonic chitosan solutions (150, 40, 10 kDa, 80% degree of deaceylation, with fluorescent chitosan tracer) was mixed with autologous rabbit whole blood, clotted with Tissue Factor to form cylindrical implants, and press-fit in drill holes in the left knee while contralateral holes received Tissue Factor or no treatment. At day 1 or day 21 post-operative, defects were analyzed by micro-computed tomography, histomorphometry and stereology for bone and soft tissue repair. Results All 3 implants filled the top of defects at day 1 and were partly degraded in situ at 21 days post-operative. All implants attracted neutrophils, osteoclasts and abundant bone marrow-derived stromal cells, stimulated bone resorption followed by new woven bone repair (bone remodeling) and promoted repair tissue-bone integration. 150 kDa chitosan implant was less degraded, and elicited more apoptotic neutrophils and bone resorption than 10 kDa chitosan implant. Drilled controls elicited a poorly integrated fibrous or fibrocartilaginous tissue. Conclusions Pre-solidified implants elicit stromal cells and vigorous bone plate remodeling through a phase involving neutrophil chemotaxis. Pre-solidified chitosan implants are tunable by molecular mass, and could be beneficial for augmented marrow stimulation therapy if the recruited stromal cells can progress to bone and cartilage repair. PMID:23324433

c-MYC independent nuclear reprogramming favors cardiogenic potential of induced pluripotent stem cells

PubMed Central

Martinez-Fernandez, Almudena; Nelson, Timothy J.; Ikeda, Yasuhiro; Terzic, Andre

2010-01-01

Induced pluripotent stem cell (iPS) technology has launched a new platform in regenerative medicine aimed at deriving unlimited replacement tissue from autologous sources through somatic cell reprogramming using stemness factor sets. In this way, authentic cardiomyocytes have been obtained from iPS and recently demonstrated in proof-of-principle studies to repair infarcted heart. Optimizing the cardiogenic potential of iPS progeny would ensure a maximized yield of bioengineered cardiac tissue. Here, we reprogrammed fibroblasts in the presence or absence of c-MYC to determine if the acquired cardiogenicity is sensitive to the method of nuclear reprogramming. Using lentiviral constructs that expressed stemness factors SOX2, OCT4, and KLF4 with or without c-MYC, iPS clones generated through fibroblast reprogramming demonstrated indistinguishable characteristics for 5 days of differentiation with similar cell morphology, growth rates, and chimeric embryo integration. However, 4-factor c-MYC dependent nuclear reprogramming produced iPS progeny that consistently prolonged the expression of pluripotent Oct-4 and Fgf4 genes and repressed cardiac differentiation. In contrast, 3-factor c-MYC-less iPS clones efficiently up-regulated pre-cardiac (CXCR4, Flk-1, and Mesp1/2) and cardiac (Nkx2.5, Mef2c, and Myocardin) gene expression patterns. In fact, 3-factor iPS progeny demonstrated early and robust cardiogenesis during in vitro differentiation with consistent beating activity, sarcomere maturation, and rhythmical intracellular calcium dynamics. Thus, nuclear reprogramming independent of c-MYC enhances production of pluripotent stem cells with innate cardiogenic potential. PMID:20221419

Targeting p35/Cdk5 signalling via CIP-peptide promotes angiogenesis in hypoxia.

PubMed

Bosutti, Alessandra; Qi, Jie; Pennucci, Roberta; Bolton, David; Matou, Sabine; Ali, Kamela; Tsai, Li-Huei; Krupinski, Jerzy; Petcu, Eugene B; Montaner, Joan; Al Baradie, Raid; Caccuri, Francesca; Caruso, Arnaldo; Alessandri, Giulio; Kumar, Shant; Rodriguez, Cristina; Martinez-Gonzalez, Jose; Slevin, Mark

2013-01-01

Cyclin-dependent kinase-5 (Cdk5) is over-expressed in both neurons and microvessels in hypoxic regions of stroke tissue and has a significant pathological role following hyper-phosphorylation leading to calpain-induced cell death. Here, we have identified a critical role of Cdk5 in cytoskeleton/focal dynamics, wherein its activator, p35, redistributes along actin microfilaments of spreading cells co-localising with p(Tyr15)Cdk5, talin/integrin beta-1 at the lamellipodia in polarising cells. Cdk5 inhibition (roscovitine) resulted in actin-cytoskeleton disorganisation, prevention of protein co-localization and inhibition of movement. Cells expressing Cdk5 (D144N) kinase mutant, were unable to spread, migrate and form tube-like structures or sprouts, while Cdk5 wild-type over-expression showed enhanced motility and angiogenesis in vitro, which was maintained during hypoxia. Gene microarray studies demonstrated myocyte enhancer factor (MEF2C) as a substrate for Cdk5-mediated angiogenesis in vitro. MEF2C showed nuclear co-immunoprecipitation with Cdk5 and almost complete inhibition of differentiation and sprout formation following siRNA knock-down. In hypoxia, insertion of Cdk5/p25-inhibitory peptide (CIP) vector preserved and enhanced in vitro angiogenesis. These results demonstrate the existence of critical and complementary signalling pathways through Cdk5 and p35, and through which coordination is a required factor for successful angiogenesis in sustained hypoxic condition.

Exchange enhancement of the electron g-factor in a two-dimensional semimetal in HgTe quantum wells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Bovkun, L. S., E-mail: bovkun@ipmras.ru; Krishtopenko, S. S.; Zholudev, M. S.

The exchange enhancement of the electron g-factor in perpendicular magnetic fields to 12 T in HgTe/CdHgTe quantum wells 20 nm wide with a semimetal band structure is studied. The electron effective mass and g-factor at the Fermi level are determined by analyzing the temperature dependence of the amplitude of Shubnikov–de Haas oscillation in weak fields and near odd Landau-level filling factors ν ≤ 9. The experimental values are compared with theoretical calculations performed in the one-electron approximation using the eight-band kp Hamiltonian. The found dependence of g-factor enhancement on the electron concentration is explained by changes in the contributions ofmore » hole- and electron-like states to exchange corrections to the Landau-level energies in the conduction band.« less

Hypoxia enhances the interaction between pancreatic stellate cells and cancer cells via increased secretion of connective tissue growth factor.

PubMed

Eguchi, Daiki; Ikenaga, Naoki; Ohuchida, Kenoki; Kozono, Shingo; Cui, Lin; Fujiwara, Kenji; Fujino, Minoru; Ohtsuka, Takao; Mizumoto, Kazuhiro; Tanaka, Masao

2013-05-01

Pancreatic cancer (PC), a hypovascular tumor, thrives under hypoxic conditions. Pancreatic stellate cells (PSCs) promote PC progression by secreting soluble factors, but their functions in hypoxia are poorly understood. This study aimed to clarify the effects of hypoxic conditions on the interaction between PC cells and PSCs. We isolated human PSCs from fresh pancreatic ductal adenocarcinomas and analyzed functional differences in PSCs between normoxia (21% O2) and hypoxia (1% O2), including expression of various factors related to tumor-stromal interactions. We particularly analyzed effects on PC invasiveness of an overexpressed molecule-connective tissue growth factor (CTGF)-in PSCs under hypoxic conditions, using RNA interference techniques. Conditioned media from hypoxic PSCs enhanced PC cell invasiveness more intensely than that from normoxic PSCs (P < 0.01). When co-cultured with PSCs, PC cell invasion was more enhanced under hypoxia than under normoxia (P < 0.05). Among various soluble factors, which were related to invasiveness, CTGF was one of the overexpressed molecules in hypoxic PSCs. A higher level of CTGF expression was also found in supernatant of hypoxic PSCs than in supernatant of normoxic PSCs. PC cell invasiveness was reduced by CTGF knockdown in hypoxic PSCs co-cultured with PC cells (P < 0.05). Hypoxia induces PSCs' secretion of CTGF, leading to enhancement of PC invasiveness. CTGF derived from hypoxia-stimulated PSCs may be a new therapeutic target for pancreatic cancer. Copyright © 2013 Elsevier Inc. All rights reserved.

Angiogenesis in calcium phosphate scaffolds by inorganic copper ion release.

PubMed

Barralet, Jake; Gbureck, Uwe; Habibovic, Pamela; Vorndran, Elke; Gerard, Catherine; Doillon, Charles J

2009-07-01

Angiogenesis in a tissue-engineered device may be induced by incorporating growth factors (e.g., vascular endothelial growth factor [VEGF]), genetically modified cells, and=or vascular cells. It represents an important process during the formation and repair of tissue and is essential for nourishment and supply of reparative and immunological cells. Inorganic angiogenic factors, such as copper ions, are therefore of interest in the fields of regenerative medicine and tissue engineering due to their low cost, higher stability, and potentially greater safety compared with recombinant proteins or genetic engineering approaches. The purpose of this study was to compare tissue responses to 3D printed macroporous bioceramic scaffolds implanted in mice that had been loaded with either VEGF or copper sulfate. These factors were spatially localized at the end of a single macropore some 7 mm from the surface of the scaffold. Controls without angiogenic factors exhibited only poor tissue growth within the blocks; in contrast, low doses of copper sulfate led to the formation of microvessels oriented along the macropore axis. Further, wound tissue ingrowth was particularly sensitive to the quantity of copper sulfate and was enhanced at specific concentrations or in combination with VEGF. The potential to accelerate and guide angiogenesis and wound healing by copper ion release without the expense of inductive protein(s) is highly attractive in the area of tissue-engineered bone and offers significant future potential in the field of regenerative biomaterials.

Decreasing CNPY2 Expression Diminishes Colorectal Tumor Growth and Development through Activation of p53 Pathway.

PubMed

Yan, Ping; Gong, Hui; Zhai, Xiaoyan; Feng, Yi; Wu, Jun; He, Sheng; Guo, Jian; Wang, Xiaoxia; Guo, Rui; Xie, Jun; Li, Ren-Ke

2016-04-01

Neovascularization drives tumor development, and angiogenic factors are important neovascularization initiators. We recently identified the secreted angiogenic factor CNPY2, but its involvement in cancer has not been explored. Herein, we investigate CNPY2's role in human colorectal cancer (CRC) development. Tumor samples were obtained from CRC patients undergoing surgery. Canopy 2 (CNPY2) expression was analyzed in tumor and adjacent normal tissue. Stable lines of human HCT116 cells expressing CNPY2 shRNA or control shRNA were established. To determine CNPY2's effects on tumor xenografts in vivo, human CNPY2 shRNA HCT116 cells and controls were injected into nude mice, separately. Cellular apoptosis, growth, and angiogenesis in the xenografts were evaluated. CNPY2 expression was significantly higher in CRC tissues. CNPY2 knockdown in HCT116 cells inhibited growth and migration and promoted apoptosis. In xenografts, CNPY2 knockdown prevented tumor growth and angiogenesis and promoted apoptosis. Knockdown of CNPY2 in the HCT116 CRC cell line reversibly increased p53 activity. The p53 activation increased cyclin-dependent kinase inhibitor p21 and decreased cyclin-dependent kinase 2, thereby inhibiting tumor cell growth, inducing cell apoptosis, and reducing angiogenesis both in vitro and in vivo. CNPY2 may play a critical role in CRC development by enhancing cell proliferation, migration, and angiogenesis and by inhibiting apoptosis through negative regulation of the p53 pathway. Therefore, CNPY2 may represent a novel CRC therapeutic target and prognostic indicator. Copyright © 2016 American Society for Investigative Pathology. Published by Elsevier Inc. All rights reserved.

Aspirin Enhances Osteogenic Potential of Periodontal Ligament Stem Cells (PDLSCs) and Modulates the Expression Profile of Growth Factor-Associated Genes in PDLSCs.

PubMed

Abd Rahman, Fazliny; Mohd Ali, Johari; Abdullah, Mariam; Abu Kasim, Noor Hayaty; Musa, Sabri

2016-07-01

This study investigates the effects of aspirin (ASA) on the proliferative capacity, osteogenic potential, and expression of growth factor-associated genes in periodontal ligament stem cells (PDLSCs). Mesenchymal stem cells (MSCs) from PDL tissue were isolated from human premolars (n = 3). The MSCs' identity was confirmed by immunophenotyping and trilineage differentiation assays. Cell proliferation activity was assessed through 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay. Polymerase chain reaction array was used to profile the expression of 84 growth factor-associated genes. Pathway analysis was used to identify the biologic functions and canonic pathways activated by ASA treatment. The osteogenic potential was evaluated through mineralization assay. ASA at 1,000 μM enhances osteogenic potential of PDLSCs. Using a fold change (FC) of 2.0 as a threshold value, the gene expression analyses indicated that 19 genes were differentially expressed, which includes 12 upregulated and seven downregulated genes. Fibroblast growth factor 9 (FGF9), vascular endothelial growth factor A (VEGFA), interleukin-2, bone morphogenetic protein-10, VEGFC, and 2 (FGF2) were markedly upregulated (FC range, 6 to 15), whereas pleotropin, FGF5, brain-derived neurotrophic factor, and Dickkopf WNT signaling pathway inhibitor 1 were markedly downregulated (FC 32). Of the 84 growth factor-associated genes screened, 35 showed high cycle threshold values (≥35). ASA modulates the expression of growth factor-associated genes and enhances osteogenic potential in PDLSCs. ASA upregulated the expression of genes that could activate biologic functions and canonic pathways related to cell proliferation, human embryonic stem cell pluripotency, tissue regeneration, and differentiation. These findings suggest that ASA enhances PDLSC function and may be useful in regenerative dentistry applications, particularly in the areas of periodontal health and regeneration.

Connective tissue growth factor/CCN2-null mouse embryonic fibroblasts retain intact transforming growth factor-{beta} responsiveness

DOE Office of Scientific and Technical Information (OSTI.GOV)

Mori, Yasuji; Hinchcliff, Monique; Wu, Minghua

2008-03-10

Background: The matricellular protein connective tissue growth factor (CCN2) has been implicated in pathological fibrosis, but its physiologic role remains elusive. In vitro, transforming growth factor-{beta} (TGF-{beta}) induces CCN2 expression in mesenchymal cells. Because CCN2 can enhance profibrotic responses elicited by TGF-{beta}, it has been proposed that CCN2 functions as an essential downstream signaling mediator for TGF-{beta}. To explore this notion, we characterized TGF-{beta}-induced activation of fibroblasts from CCN2-null (CCN2{sup -/-}) mouse embryos. Methods: The regulation of CCN2 expression was examined in vivo in a model of fibrosis induced by bleomycin. Cellular TGF-{beta} signal transduction and regulation of collagen genemore » expression were examined in CCN2{sup -/-} MEFs by immunohistochemistry, Northern, Western and RT-PCR analysis, immunocytochemistry and transient transfection assays. Results: Bleomycin-induced skin fibrosis in the mouse was associated with substantial CCN2 up-regulation in lesional fibroblasts. Whereas in vitro proliferation rate of CCN2{sup -/-} MEFs was markedly reduced compared to wild type MEFs, TGF-{beta}-induced activation of the Smad pathways, including Smad2 phosphorylation, Smad2/3 and Smad4 nuclear accumulation and Smad-dependent transcriptional responses, were unaffected by loss of CCN2. The stimulation of COL1A2 and fibronectin mRNA expression and promoter activity, and of corresponding protein levels, showed comparable time and dose-response in wild type and CCN2{sup -/-} MEFs, whereas stimulation of alpha smooth muscle actin and myofibroblast transdifferentiation showed subtle impairment in MEFs lacking CCN2. Conclusion: Whereas endogenous CCN2 plays a role in regulation of proliferation and TGF-{beta}-induced myofibroblast transdifferentiation, it appears to be dispensable for Smad-dependent stimulation of collagen and extracellular matrix synthesis in murine embryonic fibroblasts.« less

Analysis of the hypoxia-sensing pathway in Drosophila melanogaster

PubMed Central

Arquier, Nathalie; Vigne, Paul; Duplan, Eric; Hsu, Tien; Therond, Pascal P.; Frelin, Christian; D'Angelo, Gisela

2005-01-01

The mechanism by which hypoxia induces gene transcription involves the inhibition of HIF-1α (hypoxia-inducible factor-1 α subunit) PHD (prolyl hydroxylase) activity, which prevents the VHL (von Hippel-Lindau)-dependent targeting of HIF-1α to the ubiquitin/proteasome pathway. HIF-1α thus accumulates and promotes gene transcription. In the present study, first we provide direct biochemical evidence for the presence of a conserved hypoxic signalling pathway in Drosophila melanogaster. An assay for 2-oxoglutarate-dependent dioxygenases was developed using Drosophila embryonic and larval homogenates as a source of enzyme. Drosophila PHD has a low substrate specificity and hydroxylates key proline residues in the ODD (oxygen-dependent degradation) domains of human HIF-1α and Similar, the Drosophila homologue of HIF-1α. The enzyme promotes human and Drosophila [35S]VHL binding to GST (glutathione S-transferase)–ODD-domain fusion protein. Hydroxylation is enhanced by proteasomal inhibitors and was ascertained using an anti-hydroxyproline antibody. Secondly, by using transgenic flies expressing a fusion protein that combined an ODD domain and the green fluorescent protein (ODD–GFP), we analysed the hypoxic cascade in different embryonic and larval tissues. Hypoxic accumulation of the reporter protein was observed in the whole tracheal tree, but not in the ectoderm. Hypoxic stabilization of ODD–GFP in the ectoderm was restored by inducing VHL expression in these cells. These results show that Drosophila tissues exhibit different sensitivities to hypoxia. PMID:16176182

Sucrose-induced anthocyanin accumulation in vegetative tissue of Petunia plants requires anthocyanin regulatory transcription factors.

PubMed

Ai, Trinh Ngoc; Naing, Aung Htay; Arun, Muthukrishnan; Lim, Sun-Hyung; Kim, Chang Kil

2016-11-01

The effects of three different sucrose concentrations on plant growth and anthocyanin accumulation were examined in non-transgenic (NT) and transgenic (T 2 ) specimens of the Petunia hybrida cultivar 'Mirage rose' that carried the anthocyanin regulatory transcription factors B-Peru+mPAP1 or RsMYB1. Anthocyanin accumulation was not observed in NT plants in any treatments, whereas a range of anthocyanin accumulation was observed in transgenic plants. The anthocyanin content detected in transgenic plants expressing the anthocyanin regulatory transcription factors (B-Peru+mPAP1 or RsMYB1) was higher than that in NT plants. In addition, increasing sucrose concentration strongly enhanced anthocyanin content as shown by quantitative real-time polymerase chain reaction (qRT-PCR) analysis, wherein increased concentrations of sucrose enhanced transcript levels of the transcription factors that are responsible for the induction of biosynthetic genes involved in anthocyanin synthesis; this pattern was not observed in NT plants. In addition, sucrose affected plant growth, although the effects were different between NT and transgenic plants. Taken together, the application of sucrose could enhance anthocyanin production in vegetative tissue of transgenic Petunia carrying anthocyanin regulatory transcription factors, and this study provides insights about interactive effects of sucrose and transcription factors in anthocyanin biosynthesis in the transgenic plant. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

Double emulsion electrospun nanofibers as a growth factor delivery vehicle for salivary gland regeneration

NASA Astrophysics Data System (ADS)

Foraida, Zahraa I.; Sharikova, Anna; Peerzada, Lubna N.; Khmaladze, Alexander; Larsen, Melinda; Castracane, James

2017-08-01

Sustained delivery of growth factors, proteins, drugs and other biologically active molecules is necessary for tissue engineering applications. Electrospun fibers are attractive tissue engineering scaffolds as they partially mimic the topography of the extracellular matrix (ECM). However, they do not provide continuous nourishment to the tissue. In search of a biomimetic scaffold for salivary gland tissue regeneration, we previously developed a blend nanofiber scaffold composed of the protein elastin and the synthetic polymer polylactic-co-glycolic acid (PLGA). The nanofiber scaffold promoted in vivo-like salivary epithelial cell tissue organization and apicobasal polarization. However, in order to enhance the salivary cell proliferation and biomimetic character of the scaffold, sustained growth factor delivery is needed. The composite nanofiber scaffold was optimized to act as a growth factor delivery system using epidermal growth factor (EGF) as a model protein. The nanofiber/EGF hybrid nanofibers were synthesized by double emulsion electrospinning where EGF is emulsified within a water/oil/water (w/o/w) double emulsion system. Successful incorporation of EGF was confirmed using Raman spectroscopy. EGF release profile was characterized using enzyme-linked immunosorbent assay (ELIZA) of the EGF content. Double emulsion electrospinning resulted in slower release of EGF. We demonstrated the potential of the proposed double emulsion electrospun nanofiber scaffold for the delivery of growth factors and/or drugs for tissue engineering and pharmaceutical applications.

Pax2 regulates a fadd-dependent molecular switch that drives tissue fusion during eye development.

PubMed

Viringipurampeer, Ishaq A; Ferreira, Todd; DeMaria, Shannon; Yoon, Jookyung J; Shan, Xianghong; Moosajee, Mariya; Gregory-Evans, Kevin; Ngai, John; Gregory-Evans, Cheryl Y

2012-05-15

Tissue fusion is an essential morphogenetic mechanism in development, playing a fundamental role in developing neural tube, palate and the optic fissure. Disruption of genes associated with the tissue fusion can lead to congenital malformations, such as spina bifida, cleft lip/palate and ocular coloboma. For instance, the Pax2 transcription factor is required for optic fissure closure, although the mechanism of Pax2 action leading to tissue fusion remains elusive. This lack of information defining how transcription factors drive tissue morphogenesis at the cellular level is hampering new treatments options. Through loss- and gain-of-function analysis, we now establish that pax2 in combination with vax2 directly regulate the fas-associated death domain (fadd) gene. In the presence of fadd, cell proliferation is restricted in the developing eye through a caspase-dependent pathway. However, the loss of fadd results in a proliferation defect and concomitant activation of the necroptosis pathway through RIP1/RIP3 activity, leading to an abnormal open fissure. Inhibition of RIP1 with the small molecule drug necrostatin-1 rescues the pax2 eye fusion defect, thereby overcoming the underlying genetic defect. Thus, fadd has an essential physiological function in protecting the developing optic fissure neuroepithelium from RIP3-dependent necroptosis. This study demonstrates the molecular hierarchies that regulate a cellular switch between proliferation and the apoptotic and necroptotic cell death pathways, which in combination drive tissue morphogenesis. Furthermore, our data suggest that future therapeutic strategies may be based on small molecule drugs that can bypass the gene defects causing common congenital tissue fusion defects.

Effect of needle insertion speed on tissue injury, stress, and backflow distribution for convection-enhanced delivery in the rat brain.

PubMed

Casanova, Fernando; Carney, Paul R; Sarntinoranont, Malisa

2014-01-01

Flow back along a needle track (backflow) can be a problem during direct infusion, e.g. convection-enhanced delivery (CED), of drugs into soft tissues such as brain. In this study, the effect of needle insertion speed on local tissue injury and backflow was evaluated in vivo in the rat brain. Needles were introduced at three insertion speeds (0.2, 2, and 10 mm/s) followed by CED of Evans blue albumin (EBA) tracer. Holes left in tissue slices were used to reconstruct penetration damage. These measurements were also input into a hyperelastic model to estimate radial stress at the needle-tissue interface (pre-stress) before infusion. Fast insertion speeds were found to produce more tissue bleeding and disruption; average hole area at 10 mm/s was 1.87-fold the area at 0.2 mm/s. Hole measurements also differed at two fixation time points after needle retraction, 10 and 25 min, indicating that pre-stresses are influenced by time-dependent tissue swelling. Calculated pre-stresses were compressive (0 to 485 Pa) and varied along the length of the needle with smaller average values within white matter (116 Pa) than gray matter (301 Pa) regions. Average pre-stress at 0.2 mm/s (351.7 Pa) was calculated to be 1.46-fold the value at 10 mm/s. For CED backflow experiments (0.5, 1, and 2 µL/min), measured EBA backflow increased as much as 2.46-fold between 10 and 0.2 mm/s insertion speeds. Thus, insertion rate-dependent damage and changes in pre-stress were found to directly contribute to the extent of backflow, with slower insertion resulting in less damage and improved targeting.

Formation of proteoglycan and collagen-rich scaffold-free stiff cartilaginous tissue using two-step culture methods with combinations of growth factors.

PubMed

Miyazaki, Tatsuya; Miyauchi, Satoshi; Matsuzaka, Satoshi; Yamagishi, Chie; Kobayashi, Kohei

2010-05-01

Tissue-engineered cartilage may be expected to serve as an alternative to autologous chondrocyte transplantation treatment. Several methods for producing cartilaginous tissue have been reported. In this study, we describe the production of scaffold-free stiff cartilaginous tissue of pig and human, using allogeneic serum and growth factors. The tissue was formed in a mold using chondrocytes recovered from alginate bead culture and maintained in a medium with transforming growth factor-beta and several other additives. In the case of porcine tissue, the tear strength of the tissue and the contents of proteoglycan (PG) and collagen per unit of DNA increased dose-dependently with transforming growth factor-beta. The length of culture was significantly and positively correlated with thickness, tear strength, and PG and collagen contents. Tear strength showed positive high correlations with both PG and collagen contents. A positive correlation was also seen between PG content and collagen content. Similar results were obtained with human cartilaginous tissue formed from chondrocytes expanded in monolayer culture. Further, an in vivo pilot study using pig articular cartilage defect model demonstrated that the cartilaginous tissue was well integrated with surrounding tissue at 13 weeks after the implantation. In conclusion, we successfully produced implantable scaffold-free stiff cartilaginous tissue, which characterized high PG and collagen contents.

Anterior Gradient 2 (AGR2) Induced Epidermal Growth Factor Receptor (EGFR) Signaling Is Essential for Murine Pancreatitis-Associated Tissue Regeneration

PubMed Central

Wodziak, Dariusz; Dong, Aiwen; Basin, Michael F.; Lowe, Anson W.

2016-01-01

A recently published study identified Anterior Gradient 2 (AGR2) as a regulator of EGFR signaling by promoting receptor presentation from the endoplasmic reticulum to the cell surface. AGR2 also promotes tissue regeneration in amphibians and fish. Whether AGR2-induced EGFR signaling is essential for tissue regeneration in higher vertebrates was evaluated using a well-characterized murine model for pancreatitis. The impact of AGR2 expression and EGFR signaling on tissue regeneration was evaluated using the caerulein-induced pancreatitis mouse model. EGFR signaling and cell proliferation were examined in the context of the AGR2-/- null mouse or with the EGFR-specific tyrosine kinase inhibitor, AG1478. In addition, the Hippo signaling coactivator YAP1 was evaluated in the context of AGR2 expression during pancreatitis. Pancreatitis-induced AGR2 expression enabled EGFR translocation to the plasma membrane, the initiation of cell signaling, and cell proliferation. EGFR signaling and tissue regeneration were partially inhibited by the tyrosine kinase inhibitor AG1478, but absent in the AGR2-/- null mouse. AG1478-treated and AGR2-/- null mice with pancreatitis died whereas all wild-type controls recovered. YAP1 activation was also dependent on pancreatitis-induced AGR2 expression. AGR2-induced EGFR signaling was essential for tissue regeneration and recovery from pancreatitis. The results establish tissue regeneration as a major function of AGR2-induced EGFR signaling in adult higher vertebrates. Enhanced AGR2 expression and EGFR signaling are also universally present in human pancreatic cancer, which support a linkage between tissue injury, regeneration, and cancer pathogenesis. PMID:27764193

Diffusion Limitation and Hyperoxic Enhancement of Oxygen Consumption in Zooxanthellate Sea Anemones, Zoanthids, and Corals.

PubMed

Shick, J M

1990-08-01

Depending on their size and morphology, anthozoan polyps and colonies may be diffusion-limited in their oxygen consumption, even under well-stirred, air-saturated conditions. This is indicated by an enhancement of oxygen consumption under steady-state hyperoxic conditions that simulate the levels of O2 produced photosynthetically by zooxanthellae in the hosts' tissues. Such hyperoxia in the tissues of zooxanthellate species negates the effect of the diffusive boundary layer, and increases the rate of oxygen consumption; thus, in many cases, the rate of respiration measured under normoxia in the dark may not be representative of the rate during the day when the zooxanthellae are photosynthesizing and when the supply of oxygen for respiration is in the tissues themselves, not from the environment. These results have implications in respirometric methodology and in calculating the rate of gross photosynthesis in energetic studies. The activity of cytochrome c oxidase is higher in aposymbiotic than in zooxanthellate specimens of the sea anemone Aiptasia pulchella, and this may indicate a compensation for the relative hypoxia in the tissues of the former, enhancing the delivery of oxygen to the mitochondria from the environment.

Delivery of Alginate Scaffold Releasing Two Trophic Factors for Spinal Cord Injury Repair

PubMed Central

Grulova, I.; Slovinska, L.; Blaško, J.; Devaux, S.; Wisztorski, M.; Salzet, M.; Fournier, I.; Kryukov, O.; Cohen, S.; Cizkova, D.

2015-01-01

Spinal cord injury (SCI) has been implicated in neural cell loss and consequently functional motor and sensory impairment. In this study, we propose an alginate -based neurobridge enriched with/without trophic growth factors (GFs) that can be utilized as a therapeutic approach for spinal cord repair. The bioavailability of key GFs, such as Epidermal Growth factor (EGF) and basic Fibroblast Growth Factor (bFGF) released from injected alginate biomaterial to the central lesion site significantly enhanced the sparing of spinal cord tissue and increased the number of surviving neurons (choline acetyltransferase positive motoneurons) and sensory fibres. In addition, we document enhanced outgrowth of corticospinal tract axons and presence of blood vessels at the central lesion. Tissue proteomics was performed at 3, 7 and 10 days after SCI in rats indicated the presence of anti-inflammatory factors in segments above the central lesion site, whereas in segments below, neurite outgrowth factors, inflammatory cytokines and chondroitin sulfate proteoglycan of the lectican protein family were overexpressed. Collectively, based on our data, we confirm that functional recovery was significantly improved in SCI groups receiving alginate scaffold with affinity-bound growth factors (ALG +GFs), compared to SCI animals without biomaterial treatment. PMID:26348665

Invited review: mesenchymal progenitor cells in intramuscular connective tissue development.

PubMed

Miao, Z G; Zhang, L P; Fu, X; Yang, Q Y; Zhu, M J; Dodson, M V; Du, M

2016-01-01

The abundance and cross-linking of intramuscular connective tissue contributes to the background toughness of meat, and is thus undesirable. Connective tissue is mainly synthesized by intramuscular fibroblasts. Myocytes, adipocytes and fibroblasts are derived from a common pool of progenitor cells during the early embryonic development. It appears that multipotent mesenchymal stem cells first diverge into either myogenic or non-myogenic lineages; non-myogenic mesenchymal progenitors then develop into the stromal-vascular fraction of skeletal muscle wherein adipocytes, fibroblasts and derived mesenchymal progenitors reside. Because non-myogenic mesenchymal progenitors mainly undergo adipogenic or fibrogenic differentiation during muscle development, strengthening progenitor proliferation enhances the potential for both intramuscular adipogenesis and fibrogenesis, leading to the elevation of both marbling and connective tissue content in the resulting meat product. Furthermore, given the bipotent developmental potential of progenitor cells, enhancing their conversion to adipogenesis reduces fibrogenesis, which likely results in the overall improvement of marbling (more intramuscular adipocytes) and tenderness (less connective tissue) of meat. Fibrogenesis is mainly regulated by the transforming growth factor (TGF) β signaling pathway and its regulatory cascade. In addition, extracellular matrix, a part of the intramuscular connective tissue, provides a niche environment for regulating myogenic differentiation of satellite cells and muscle growth. Despite rapid progress, many questions remain in the role of extracellular matrix on muscle development, and factors determining the early differentiation of myogenic, adipogenic and fibrogenic cells, which warrant further studies.

Low-Intensity Extracorporeal Shock Wave Therapy Enhances Brain-Derived Neurotrophic Factor Expression through PERK/ATF4 Signaling Pathway.

PubMed

Wang, Bohan; Ning, Hongxiu; Reed-Maldonado, Amanda B; Zhou, Jun; Ruan, Yajun; Zhou, Tie; Wang, Hsun Shuan; Oh, Byung Seok; Banie, Lia; Lin, Guiting; Lue, Tom F

2017-02-16

Low-intensity extracorporeal shock wave therapy (Li-ESWT) is used in the treatment of erectile dysfunction, but its mechanisms are not well understood. Previously, we found that Li-ESWT increased the expression of brain-derived neurotrophic factor (BDNF). Here we assessed the underlying signaling pathways in Schwann cells in vitro and in penis tissue in vivo after nerve injury. The result indicated that BDNF were significantly increased by the Li-ESWT after nerve injury, as well as the expression of BDNF in Schwann cells (SCs, RT4-D6P2T) in vitro. Li-ESWT activated the protein kinase RNA-like endoplasmic reticulum (ER) kinase (PERK) pathway by increasing the phosphorylation levels of PERK and eukaryotic initiation factor 2a (eIF2α), and enhanced activating transcription factor 4 (ATF4) in an energy-dependent manner. In addition, GSK2656157-an inhibitor of PERK-effectively inhibited the effect of Li-ESWT on the phosphorylation of PERK, eIF2α, and the expression of ATF4. Furthermore, silencing ATF4 dramatically attenuated the effect of Li-ESWT on the expression of BDNF, but had no effect on hypoxia-inducible factor (HIF)1α or glial cell-derived neurotrophic factor (GDNF) in Schwann cells. In conclusion, our findings shed new light on the underlying mechanisms by which Li-ESWT may stimulate the expression of BDNF through activation of PERK/ATF4 signaling pathway. This information may help to refine the use of Li-ESWT to further improve its clinical efficacy.

Low-Intensity Extracorporeal Shock Wave Therapy Enhances Brain-Derived Neurotrophic Factor Expression through PERK/ATF4 Signaling Pathway

PubMed Central

Wang, Bohan; Ning, Hongxiu; Reed-Maldonado, Amanda B.; Zhou, Jun; Ruan, Yajun; Zhou, Tie; Wang, Hsun Shuan; Oh, Byung Seok; Banie, Lia; Lin, Guiting; Lue, Tom F.

2017-01-01

Low-intensity extracorporeal shock wave therapy (Li-ESWT) is used in the treatment of erectile dysfunction, but its mechanisms are not well understood. Previously, we found that Li-ESWT increased the expression of brain-derived neurotrophic factor (BDNF). Here we assessed the underlying signaling pathways in Schwann cells in vitro and in penis tissue in vivo after nerve injury. The result indicated that BDNF were significantly increased by the Li-ESWT after nerve injury, as well as the expression of BDNF in Schwann cells (SCs, RT4-D6P2T) in vitro. Li-ESWT activated the protein kinase RNA-like endoplasmic reticulum (ER) kinase (PERK) pathway by increasing the phosphorylation levels of PERK and eukaryotic initiation factor 2a (eIF2α), and enhanced activating transcription factor 4 (ATF4) in an energy-dependent manner. In addition, GSK2656157—an inhibitor of PERK—effectively inhibited the effect of Li-ESWT on the phosphorylation of PERK, eIF2α, and the expression of ATF4. Furthermore, silencing ATF4 dramatically attenuated the effect of Li-ESWT on the expression of BDNF, but had no effect on hypoxia-inducible factor (HIF)1α or glial cell-derived neurotrophic factor (GDNF) in Schwann cells. In conclusion, our findings shed new light on the underlying mechanisms by which Li-ESWT may stimulate the expression of BDNF through activation of PERK/ATF4 signaling pathway. This information may help to refine the use of Li-ESWT to further improve its clinical efficacy. PMID:28212323

Neurotrophins and tonsillar hypertrophy in children with obstructive sleep apnea.

PubMed

Goldbart, Aviv D; Mager, Edward; Veling, Maria C; Goldman, Julie L; Kheirandish-Gozal, Leila; Serpero, Laura D; Piedimonte, Giovanni; Gozal, David

2007-10-01

Enlarged adenotonsillar tissue (AT) is a major determinant of obstructive sleep apnea (OSA) severity in children; however, mechanisms of AT proliferation are poorly understood. We hypothesized that early exposure to respiratory syncytial virus (RSV) may modify AT proliferation through up-regulation of nerve growth factor (NGF)-neurokinin 1 (NK1) receptor dependent pathways. AT harvested from 34 children with OSA and 25 children with recurrent tonsillitis (RI) were examined for mRNA expression of multiple growth factors and their receptors. In addition, NK1 receptor expression and location, and substance P tissue concentrations were compared in AT from OSA and RI children. NGF mRNA and its high-affinity tyrosine kinase receptor (trkA) expression were selectively increased in OSA (p<0.001). NK1 receptor mRNA and protein expression were also enhanced in OSA (p<0.01), and substance P concentrations in OSA patients were higher than in RI (p<0.0001). AT from OSA children exhibit distinct differences in the expression of NGF and trkA receptors, NK1 receptors, and substance P. The homology between these changes and those observed in the lower airways following RSV infection suggests that RSV may have induced neuro-immunomodulatory changes within AT, predisposing them to increased proliferation, and ultimately contribute to emergence of OSA.

Neurotrophins and Tonsillar Hypertrophy in Children With Obstructive Sleep Apnea

PubMed Central

GOLDBART, AVIV D.; MAGER, EDWARD; VELING, MARIA C.; GOLDMAN, JULIE L.; KHEIRANDISH-GOZAL, LEILA; SERPERO, LAURA D.; PIEDIMONTE, GIOVANNI; GOZAL, DAVID

2013-01-01

Enlarged adenotonsillar tissue (AT) is a major determinant of obstructive sleep apnea (OSA) severity in children; however, mechanisms of AT proliferation are poorly understood. We hypothesized that early exposure to respiratory syncytial virus (RSV) may modify AT proliferation through up-regulation of nerve growth factor (NGF)-neurokinin 1 (NK1) receptor dependent pathways. AT harvested from 34 children with OSA and 25 children with recurrent tonsillitis (RI) were examined for mRNA expression of multiple growth factors and their receptors. In addition, NK1 receptor expression and location, and substance P tissue concentrations were compared in AT from OSA and RI children. NGF mRNA and its high-affinity tyrosine kinase receptor (trkA) expression were selectively increased in OSA (p < 0.001). NK1 receptor mRNA and protein expression were also enhanced in OSA (p < 0.01), and substance P concentrations in OSA patients were higher than in RI (p < 0.0001). AT from OSA children exhibit distinct differences in the expression of NGF and trkA receptors, NK1 receptors, and substance P. The homology between these changes and those observed in the lower airways following RSV infection suggests that RSV may have induced neuro-immunomodulatory changes within AT, predisposing them to increased proliferation, and ultimately contribute to emergence of OSA. PMID:17667845

How electromagnetic fields can influence adult stem cells: positive and negative impacts.

PubMed

Maziarz, Aleksandra; Kocan, Beata; Bester, Mariusz; Budzik, Sylwia; Cholewa, Marian; Ochiya, Takahiro; Banas, Agnieszka

2016-04-18

The electromagnetic field (EMF) has a great impact on our body. It has been successfully used in physiotherapy for the treatment of bone disorders and osteoarthritis, as well as for cartilage regeneration or pain reduction. Recently, EMFs have also been applied in in vitro experiments on cell/stem cell cultures. Stem cells reside in almost all tissues within the human body, where they exhibit various potential. These cells are of great importance because they control homeostasis, regeneration, and healing. Nevertheless, stem cells when become cancer stem cells, may influence the pathological condition. In this article we review the current knowledge on the effects of EMFs on human adult stem cell biology, such as proliferation, the cell cycle, or differentiation. We present the characteristics of the EMFs used in miscellaneous assays. Most research has so far been performed during osteogenic and chondrogenic differentiation of mesenchymal stem cells. It has been demonstrated that the effects of EMF stimulation depend on the intensity and frequency of the EMF and the time of exposure to it. However, other factors may affect these processes, such as growth factors, reactive oxygen species, and so forth. Exploration of this research area may enhance the development of EMF-based technologies used in medical applications and thereby improve stem cell-based therapy and tissue engineering.

Macrophages redirect phagocytosis by non-professional phagocytes and influence inflammation.

PubMed

Han, Claudia Z; Juncadella, Ignacio J; Kinchen, Jason M; Buckley, Monica W; Klibanov, Alexander L; Dryden, Kelly; Onengut-Gumuscu, Suna; Erdbrügger, Uta; Turner, Stephen D; Shim, Yun M; Tung, Kenneth S; Ravichandran, Kodi S

2016-11-24

Professional phagocytes (such as macrophages) and non-professional phagocytes (such as epithelial cells) clear billions of apoptotic cells and particles on a daily basis. Although professional and non-professional macrophages reside in proximity in most tissues, whether they communicate with each other during cell clearance, and how this might affect inflammation, is not known. Here we show that macrophages, through the release of a soluble growth factor and microvesicles, alter the type of particles engulfed by non-professional phagocytes and influence their inflammatory response. During phagocytosis of apoptotic cells or in response to inflammation-associated cytokines, macrophages released insulin-like growth factor 1 (IGF-1). The binding of IGF-1 to its receptor on non-professional phagocytes redirected their phagocytosis, such that uptake of larger apoptotic cells was reduced whereas engulfment of microvesicles was increased. IGF-1 did not alter engulfment by macrophages. Macrophages also released microvesicles, whose uptake by epithelial cells was enhanced by IGF-1 and led to decreased inflammatory responses by epithelial cells. Consistent with these observations, deletion of IGF-1 receptor in airway epithelial cells led to exacerbated lung inflammation after allergen exposure. These genetic and functional studies reveal that IGF-1- and microvesicle-dependent communication between macrophages and epithelial cells can critically influence the magnitude of tissue inflammation in vivo.

Comparative analysis of human tissue interactomes reveals factors leading to tissue-specific manifestation of hereditary diseases.

PubMed

Barshir, Ruth; Shwartz, Omer; Smoly, Ilan Y; Yeger-Lotem, Esti

2014-06-01

An open question in human genetics is what underlies the tissue-specific manifestation of hereditary diseases, which are caused by genomic aberrations that are present in cells across the human body. Here we analyzed this phenomenon for over 300 hereditary diseases by using comparative network analysis. We created an extensive resource of protein expression and interactions in 16 main human tissues, by integrating recent data of gene and protein expression across tissues with data of protein-protein interactions (PPIs). The resulting tissue interaction networks (interactomes) shared a large fraction of their proteins and PPIs, and only a small fraction of them were tissue-specific. Applying this resource to hereditary diseases, we first show that most of the disease-causing genes are widely expressed across tissues, yet, enigmatically, cause disease phenotypes in few tissues only. Upon testing for factors that could lead to tissue-specific vulnerability, we find that disease-causing genes tend to have elevated transcript levels and increased number of tissue-specific PPIs in their disease tissues compared to unaffected tissues. We demonstrate through several examples that these tissue-specific PPIs can highlight disease mechanisms, and thus, owing to their small number, provide a powerful filter for interrogating disease etiologies. As two thirds of the hereditary diseases are associated with these factors, comparative tissue analysis offers a meaningful and efficient framework for enhancing the understanding of the molecular basis of hereditary diseases.

Numerical Study on Focusing of Ultrasounds in Microbubble-enhanced HIFU

NASA Astrophysics Data System (ADS)

Matsumoto, Yoichiro; Okita, Kohei; Takagi, Shu

2011-11-01

The injection of microbubbles into the target tissue enhances tissue heating in High-Intensity Focused Ultrasound therapy, via inertial cavitation. The control of the inertial cavitation is required to achieve the efficient tissue ablation. Microbubbles between a transducer and a target disturb the ultrasound propagation depending on the conditions. A method to clear such microbubbles has been proposed by Kajiyama et al. [Physics Procedia 3 (2010) 305-314]. In the method, the irradiation of intense ultrasounds with a burst waveform fragmentize microbubbles in the pathways before the irradiation of ultrasounds for tissue heating. The vitro experiment using a gel containing microbubbles has showed that the method enables to heat the target correctly by controlling the microbubble distribution. Following the experiment, we simulate the focusing of ultrasounds through a mixture containing microbubbles with considering the size and number density distributions in space. The numerical simulation shows that the movement of the heating region from the transducer side to the target by controlling the microbubble distributions. The numerical results elucidate well the experimental ones.

Estrogen receptor 1 (ESR1) regulates VEGFA in adipose tissue.

PubMed

Fatima, L A; Campello, R S; Santos, R de Souza; Freitas, H S; Frank, A P; Machado, U F; Clegg, D J

2017-12-01

Vascular endothelial growth factor A (VEGFA) is a key factor in the regulation of angiogenesis in adipose tissue. Poor vascularization during adipose tissue proliferation causes fibrosis and local inflammation, and is associated with insulin resistance. It is known that 17-beta estradiol (E2) regulates adipose tissue function and VEGFA expression in other tissues; however, the ability of E2 to regulate VEGFA in adipose tissue is currently unknown. In this study, we showed that, in 3T3-L1 cells, E2 and the estrogen receptor 1 (ESR1) agonist PPT induced VEGFA expression, while ESR1 antagonist (MPP), and selective knockdown of ESR1 using siRNA decreased VEGFA and prevented the ability of E2 to modulate its expression. Additionally, we found that E2 and PPT induced the binding of hypoxia inducible factor 1 alpha subunit (HIF1A) in the VEGFA gene promoter. We further found that VEGFA expression was lower in inguinal and gonadal white adipose tissues of ESR1 total body knockout female mice compared to wild type mice. In conclusion, our data provide evidence of an important role for E2/ESR1 in modulating adipose tissue VEGFA, which is potentially important to enhance angiogenesis, reduce inflammation and improve adipose tissue function.

Passive cavitation detection during pulsed HIFU exposures of ex vivo tissues and in vivo mouse pancreatic tumors.

PubMed

Li, Tong; Chen, Hong; Khokhlova, Tatiana; Wang, Yak-Nam; Kreider, Wayne; He, Xuemei; Hwang, Joo Ha

2014-07-01

Pulsed high-intensity focused ultrasound (pHIFU) has been shown to enhance vascular permeability, disrupt tumor barriers and enhance drug penetration into tumor tissue through acoustic cavitation. Monitoring of cavitation activity during pHIFU treatments and knowing the ultrasound pressure levels sufficient to reliably induce cavitation in a given tissue are therefore very important. Here, three metrics of cavitation activity induced by pHIFU and evaluated by confocal passive cavitation detection were introduced: cavitation probability, cavitation persistence and the level of the broadband acoustic emissions. These metrics were used to characterize cavitation activity in several ex vivo tissue types (bovine tongue and liver and porcine adipose tissue and kidney) and gel phantoms (polyacrylamide and agarose) at varying peak-rare factional focal pressures (1-12 MPa) during the following pHIFU protocol: frequency 1.1 MHz, pulse duration 1 ms and pulse repetition frequency 1 Hz. To evaluate the relevance of the measurements in ex vivo tissue, cavitation metrics were also investigated and compared in the ex vivo and in vivo murine pancreatic tumors that develop spontaneously in transgenic KrasLSL.G12 D/+; p53 R172 H/+; PdxCretg/+ (KPC) mice and closely re-capitulate human disease in their morphology. The cavitation threshold, defined at 50% cavitation probability, was found to vary broadly among the investigated tissues (within 2.5-10 MPa), depending mostly on the water-lipid ratio that characterizes the tissue composition. Cavitation persistence and the intensity of broadband emissions depended both on tissue structure and lipid concentration. Both the cavitation threshold and broadband noise emission level were similar between ex vivo and in vivo pancreatic tumor tissue. The largest difference between in vivo and ex vivo settings was found in the pattern of cavitation occurrence throughout pHIFU exposure: it was sporadic in vivo, but it decreased rapidly and stopped over the first few pulses ex vivo. Cavitation activity depended on the interplay between the destruction and circulation of cavitation nuclei, which are not only used up by HIFU treatment but also replenished or carried away by circulation in vivo. These findings are important for treatment planning and optimization in pHIFU-induced drug delivery, in particular for pancreatic tumors. Published by Elsevier Inc.

Calcium dobesilate potentiates endothelium-derived hyperpolarizing factor-mediated relaxation of human penile resistance arteries

PubMed Central

Angulo, Javier; Cuevas, Pedro; Fernández, Argentina; Gabancho, Sonia; Videla, Sebastián; Tejada, Iñigo Sáenz de

2003-01-01

We have evaluated the participation of endothelium-derived hyperpolarizing factor (EDHF) in the endothelium-dependent relaxation of isolated human penile resistance arteries (HPRA) and human corpus cavernosum (HCC) strips. In addition, the effect of the angioprotective agent, calcium dobesilate (DOBE), on the endothelium-dependent relaxation of these tissues was investigated. Combined inhibition of nitric oxide synthase (NOS) and cyclooxygenase (COX) nearly abolished the endothelium-dependent relaxation to acetylcholine (ACh) in HCC, while 60% relaxation of HPRA was observed under these conditions. Endothelium-dependent relaxation of HPRA resistant to NOS and COX inhibition was prevented by raising the extracellular concentration of K+ (35 mM) or by blocking Ca2+-activated K+ channels, with apamin (APA; 100 nM) and charybdotoxin (CTX; 100 nM), suggesting the involvement of EDHF in these responses. Endothelium-dependent relaxation to ACh was markedly enhanced by DOBE (10 μM) in HPRA but not in HCC. The potentiating effects of DOBE on ACh-induced responses in HPRA, remained after NOS and COX inhibition, were reduced by inhibition of cytochrome P450 oxygenase with miconazole (0.3 mM) and were abolished by high K+ or a combination of APA and CTX. In vivo, DOBE (10 mg kg−1 i.v.) significantly potentiated the erectile responses to cavernosal nerve stimulation in male rats. EDHF plays an important role in the endothelium-dependent relaxation of HPRA but not in HCC. DOBE significantly improves endothelium-dependent relaxation of HPRA mediated by EDHF and potentiates erectile responses in vivo. Thus, EDHF becomes a new therapeutic target for the treatment of erectile dysfunction (ED) and DOBE could be considered a candidate for oral therapy for ED. PMID:12813009

The effects of deformation, ischemia, and reperfusion on the development of muscle damage during prolonged loading.

PubMed

Loerakker, S; Manders, E; Strijkers, G J; Nicolay, K; Baaijens, F P T; Bader, D L; Oomens, C W J

2011-10-01

Deep tissue injury (DTI) is a severe form of pressure ulcer where tissue damage starts in deep tissues underneath intact skin. In the present study, the contributions of deformation, ischemia, and reperfusion to skeletal muscle damage development were examined in a rat model during a 6-h period. Magnetic resonance imaging (MRI) was used to study perfusion (contrast-enhanced MRI) and tissue integrity (T2-weighted MRI). The levels of tissue deformation were estimated using finite element models. Complete ischemia caused a gradual homogeneous increase in T2 (∼20% during the 6-h period). The effect of reperfusion on T2 was highly variable, depending on the anatomical location. In experiments involving deformation, inevitably associated with partial ischemia, a variable T2 increase (17-66% during the 6-h period) was observed reflecting the significant variation in deformation (with two-dimensional strain energies of 0.60-1.51 J/mm) and ischemia (50.8-99.8% of the leg) between experiments. These results imply that deformation, ischemia, and reperfusion all contribute to the damage process during prolonged loading, although their importance varies with time. The critical deformation threshold and period of ischemia that cause muscle damage will certainly vary between individuals. These variations are related to intrinsic factors, such as pathological state, which partly explain the individual susceptibility to the development of DTI and highlight the need for regular assessments of individual subjects.

The influence of thiazolidinediones on adipogenesis in vitro and in vivo: potential modifiers of intramuscular adipose tissue deposition in meat animals.

PubMed

Hausman, G J; Poulos, S P; Pringle, T D; Azain, M J

2008-04-01

Thiazolidinediones (TZD) are insulin sensitizing agents currently used for the treatment of type 2 diabetes and are widely used as adipogenic agents because they are ligands of peroxisome proliferator-activated receptor gamma (PPARgamma), a key adipogenic transcription factor. In vivo and in vitro studies of TZD as potential modifiers of intramuscular or marbling adipogenesis are reviewed. Thiazolidinedione-induced adipogenesis has been reported in numerous cell culture systems, including rodent, human, bovine, and porcine adipose tissue stromal-vascular (S-V) cell cultures. Studies of porcine S-V cell cultures derived from semitendinosus muscle show that TZD can potentially modify intramuscular or marbling adipogenesis. Preadipocyte recruitment was TZD-dependent in muscle S-V cultures but TZD-independent in adipose S-V cultures. There appear to be differences between adipocytes in muscle and subcutaneous adipose tissue, reminiscent of differences observed in adipocytes from different adipose tissue depots. Troglitazone, a TZD, induces marbling adipogenesis without inhibiting myogenesis when cells are grown on laminin precoated culture dishes. Additionally, troglitazone treatment does not increase lipid content in porcine adipose tissue or muscle S-V cell cultures. Thiazolidinedione treatment increases lipid content of muscle in rodents and humans; however, rosiglitazone treatment for 49 d in pigs did not influence muscle lipid content and meat quality, but several significant changes in muscle fatty acid composition were observed. Although timing of treatment with TZD needs to be optimized, evidence suggests these compounds may enhance marbling deposition in swine.

Activated PAR-2 regulates pancreatic cancer progression through ILK/HIF-α-induced TGF-α expression and MEK/VEGF-A-mediated angiogenesis.

PubMed

Chang, Li-Hsun; Pan, Shiow-Lin; Lai, Chin-Yu; Tsai, An-Chi; Teng, Che-Ming

2013-08-01

Tissue factor initiates the process of thrombosis and activates cell signaling through protease-activated receptor-2 (PAR-2). The aim of this study was to investigate the pathological role of PAR-2 signaling in pancreatic cancer. We first demonstrated that activated PAR-2 up-regulated the protein expression of both hypoxia-inducible factor-1α (HIF-1α) and HIF-2α, resulting in enhanced transcription of transforming growth factor-α (TGF-α). Down-regulation of HIFs-α by siRNA or YC-1, an HIF inhibitor, resulted in depleted levels of TGF-α protein. Furthermore, PAR-2, through integrin-linked kinase (ILK) signaling, including the p-AKT, promoted HIF protein expression. Diminishing ILK by siRNA decreased the levels of PAR-2-induced p-AKT, HIFs-α, and TGF-α; our results suggest that ILK is involved in the PAR-2-mediated TGF-α via an HIF-α-dependent pathway. Furthermore, the culture medium from PAR-2-treated pancreatic cancer cells enhanced human umbilical vein endothelial cell proliferation and tube formation, which was blocked by the MEK inhibitor, PD98059. We also found that activated PAR-2 enhanced tumor angiogenesis through the release of vascular endothelial growth factor-A (VEGF-A) from cancer cells, independent of the ILK/HIFs-α pathways. Consistent with microarray analysis, activated PAR-2 induced TGF-A and VEGF-A gene expression. In conclusion, the activation of PAR-2 signaling induced human pancreatic cancer progression through the induction of TGF-α expression by ILK/HIFs-α, as well as through MEK/VEGF-A-mediated angiogenesis, and it plays a role in the interaction between cancer progression and cancer-related thrombosis. Copyright © 2013 American Society for Investigative Pathology. Published by Elsevier Inc. All rights reserved.

Angiotensin II Receptor Antagonism Reduces Transforming Growth Factor Beta and Smad Signaling in Thoracic Aortic Aneurysm

PubMed Central

Nataatmadja, Maria; West, Jennifer; Prabowo, Sulistiana; West, Malcolm

2013-01-01

ABSTRACT Background The expression of transforming growth factor beta (TGF-β) and Smad3 regulates extracellular matrix homeostasis and inflammation in aortic aneurysms. The expression of Smad3 depends on signaling by angiotensin II (AngII) receptor pathways through TGF-β receptor–dependent and –independent pathways. Methods To determine the expression of AngII type 1 (AT1R) and type 2 receptors (AT2R), TGF-β, and Smad3 in thoracic aortic aneurysms, we performed immunohistochemistry testing on tissue and cultured cells derived from subjects with Marfan syndrome (MFS) and bicuspid aortic valve (BAV) malformation and from normal aortas of subjects who were organ donors. Results MFS and BAV aneurysm tissue showed enhanced accumulation of TGF-β and Smad3 in vascular smooth muscle cells (VSMCs) and in inflammatory cells in the subintimal layer and tunica media. The normal aortic wall exhibited minimal TGF-β and Smad3 staining. Cultured VSMCs from MFS and BAV samples showed nuclear Smad3 and strong cytoplasmic TGF-β expression in the cytoplasmic vesicles. In control cells, Smad3 was located mainly in the cytoplasm, and weak cytoplasmic TGF-β was distributed with a pattern similar to that of the aneurysm-derived cells. Compared to normal aorta cells, AT1R and AT2R expression was increased in both aneurysm types. Treatment of cultured VSMCs with the AT1R antagonist losartan caused both reduced TGF-β vesicle localization and nuclear expression of Smad3. Conclusions Increased TGF-β and Smad3 expression in aneurysm tissue and cultured VSMCs is consistent with aberrant TGF-β expression and the activation of Smad3 signaling. Losartan-mediated reduction in TGF-β expression and the cytoplasmic localization of Smad3 support a role for AT1R antagonism in the inhibition of aneurysm progression. PMID:23532685

Growth of Murine Splenic Tissue Is Suppressed by Lymphotoxin β-Receptor Signaling (LTβR) Originating from Splenic and Non-Splenic Tissues.

PubMed

Milićević, Novica M; Nohroudi, Klaus; Schmidt, Friederike; Schmidt, Hendrik; Ringer, Cornelia; Sorensen, Grith Lykke; Milićević, Živana; Westermann, Jürgen

2016-01-01

Development and maintenance of secondary lymphoid organs such as lymph nodes and spleen essentially depend on lymphotoxin β-receptor (LTβR) signaling. It is unclear, however, by which molecular mechanism their size is limited. Here, we investigate whether the LTβR pathway is also growth suppressing. By using splenic tissue transplantation it is possible to analyze a potential contribution of LTβR signaling inside and outside of the implanted tissue. We show that LTβR signaling within the endogenous spleen and within non-splenic tissues both significantly suppressed the regeneration of implanted splenic tissue. The suppressive activity positively correlated with the total number of LTβR expressing cells in the animal (regenerate weights of 115 ± 8 mg in LTβR deficient recipients and of 12 ± 9 mg in wild-type recipients), affected also developed splenic tissue, and was induced but not executed via LTβR signaling. Two-dimensional differential gel electrophoresis and subsequent mass spectrometry of stromal splenic tissue was applied to screen for potential factors mediating the LTβR dependent suppressive activity. Thus, LTβR dependent growth suppression is involved in regulating the size of secondary lymphoid organs, and might be therapeutically used to eradicate tertiary lymphoid tissues during autoimmune diseases.

Growth of Murine Splenic Tissue Is Suppressed by Lymphotoxin β-Receptor Signaling (LTβR) Originating from Splenic and Non-Splenic Tissues

PubMed Central

Schmidt, Friederike; Schmidt, Hendrik; Ringer, Cornelia; Sorensen, Grith Lykke; Milićević, Živana; Westermann, Jürgen

2016-01-01

Development and maintenance of secondary lymphoid organs such as lymph nodes and spleen essentially depend on lymphotoxin β-receptor (LTβR) signaling. It is unclear, however, by which molecular mechanism their size is limited. Here, we investigate whether the LTβR pathway is also growth suppressing. By using splenic tissue transplantation it is possible to analyze a potential contribution of LTβR signaling inside and outside of the implanted tissue. We show that LTβR signaling within the endogenous spleen and within non-splenic tissues both significantly suppressed the regeneration of implanted splenic tissue. The suppressive activity positively correlated with the total number of LTβR expressing cells in the animal (regenerate weights of 115 ± 8 mg in LTβR deficient recipients and of 12 ± 9 mg in wild-type recipients), affected also developed splenic tissue, and was induced but not executed via LTβR signaling. Two-dimensional differential gel electrophoresis and subsequent mass spectrometry of stromal splenic tissue was applied to screen for potential factors mediating the LTβR dependent suppressive activity. Thus, LTβR dependent growth suppression is involved in regulating the size of secondary lymphoid organs, and might be therapeutically used to eradicate tertiary lymphoid tissues during autoimmune diseases. PMID:27936003

Bioglass Activated Skin Tissue Engineering Constructs for Wound Healing.

PubMed

Yu, Hongfei; Peng, Jinliang; Xu, Yuhong; Chang, Jiang; Li, Haiyan

2016-01-13

Wound healing is a complicated process, and fibroblast is a major cell type that participates in the process. Recent studies have shown that bioglass (BG) can stimulate fibroblasts to secrete a multitude of growth factors that are critical for wound healing. Therefore, we hypothesize that BG can stimulate fibroblasts to have a higher bioactivity by secreting more bioactive growth factors and proteins as compared to untreated fibroblasts, and we aim to construct a bioactive skin tissue engineering graft for wound healing by using BG activated fibroblast sheet. Thus, the effects of BG on fibroblast behaviors were studied, and the bioactive skin tissue engineering grafts containing BG activated fibroblasts were applied to repair the full skin lesions on nude mouse. Results showed that BG stimulated fibroblasts to express some critical growth factors and important proteins including vascular endothelial growth factor, basic fibroblast growth factor, epidermal growth factor, collagen I, and fibronectin. In vivo results revealed that fibroblasts in the bioactive skin tissue engineering grafts migrated into wound bed, and the migration ability of fibroblasts was stimulated by BG. In addition, the bioactive BG activated fibroblast skin tissue engineering grafts could largely increase the blood vessel formation, enhance the production of collagen I, and stimulate the differentiation of fibroblasts into myofibroblasts in the wound site, which would finally accelerate wound healing. This study demonstrates that the BG activated skin tissue engineering grafts contain more critical growth factors and extracellular matrix proteins that are beneficial for wound healing as compared to untreated fibroblast cell sheets.

* Central Growth Factor Loaded Depots in Bone Tissue Engineering Scaffolds for Enhanced Cell Attraction.

PubMed

Quade, Mandy; Knaack, Sven; Akkineni, Ashwini Rahul; Gabrielyan, Anastasia; Lode, Anja; Rösen-Wolff, Angela; Gelinsky, Michael

2017-08-01

Tissue engineering, the application of stem and progenitor cells in combination with an engineered extracellular matrix, is a promising strategy for bone regeneration. However, its success is limited by the lack of vascularization after implantation. The concept of in situ tissue engineering envisages the recruitment of cells necessary for tissue regeneration from the host environment foregoing ex vivo cell seeding of the scaffold. In this study, we developed a novel scaffold system for enhanced cell attraction, which is based on biomimetic mineralized collagen scaffolds equipped with a central biopolymer depot loaded with chemotactic agents. In humid milieu, as after implantation, the signaling factors are expected to slowly diffuse out of the central depot forming a gradient that stimulates directed cell migration toward the scaffold center. Heparin, hyaluronic acid, and alginate have been shown to be capable of depot formation. By using vascular endothelial growth factor (VEGF) as model factor, it was demonstrated that the release kinetics can be adjusted by varying the depot composition. While alginate and hyaluronic acid are able to reduce the initial burst and prolong the release of VEGF, the addition of heparin led to a much stronger retention that resulted in an almost linear release over 28 days. The biological activity of released VEGF was proven for all variants using an endothelial cell proliferation assay. Furthermore, migration experiments with endothelial cells revealed a relationship between the degree of VEGF retention and migration distance: cells invaded deepest in scaffolds containing a heparin-based depot indicating that the formation of a steep gradient is crucial for cell attraction. In conclusion, this novel in situ tissue engineering approach, specifically designed to recruit and accommodate endogenous cells upon implantation, appeared highly promising to stimulate cell invasion, which in turn would promote vascularization and finally new bone formation.

Histone-Targeted Nucleic Acid Delivery for Tissue Regenerative Applications

NASA Astrophysics Data System (ADS)

Munsell, Erik V.

Nucleic acid delivery has garnered significant attention as an innovative therapeutic approach for treating a wide variety of diseases. However, the design of non-viral delivery systems that negotiate efficient intracellular trafficking and nuclear entry represents a significant challenge. Overcoming these hurdles requires a combination of well-controlled materials approaches with techniques to understand and direct cellular delivery. Recent investigations have highlighted the roles histone tail sequences play in directing nuclear delivery and retention, as well as activating DNA transcription. We established the ability to recapitulate these natural histone tail activities within non-viral gene nanocarriers, driving gene transfer/expression by enabling effective navigation to the nucleus via retrograde vesicular trafficking. A unique finding of this histone-targeted approach was that nanocarriers gained enhanced access to the nucleus during mitosis. The work described in this dissertation builds off of these fundamental insights to facilitate the translation of this histone-targeted delivery approach toward regenerative medicine applications. During native tissue repair, actively proliferating mesenchymal stem cells (MSCs) respond to a complex series of growth factor signals that direct their differentiation. Accordingly, the investigations in this work focused on utilizing the histone-targeted nanocarriers to enhance osteogenic growth factor gene transfer in dividing MSCs leading to augmented MSC chondrogenic differentiation, an essential first step in skeletal tissue repair. Concurrently, additional studies focused on optimizing the histone-targeted nanocarrier design strategy to enable improved plasmid DNA (pDNA) binding stability and tunable harnessing of native cellular processing pathways for enhanced gene transfer. Overall, the work presented herein demonstrated substantial increases in growth factor expression following histone-targeted gene transfer. This enhanced expression enabled more robust levels of chondrogenesis in MSCs than treatments with equivalent amounts of recombinant growth factor protein. Additionally, nanocarrier design optimization provided effective pDNA condensation and controllable interactions with native histone effectors. Importantly, these optimized nanocarriers conferred stable nanoplex formation and maintained transfection efficiency under physiologically relevant conditions. Taken together, these advances may help drive the clinical translation of histone-targeted nucleic acid delivery strategies for the regeneration of damaged tissue following traumatic injury.

Loss of Sirt3 accelerates arterial thrombosis by increasing formation of neutrophil extracellular traps and plasma tissue factor activity

PubMed Central

Gaul, Daniel S; Weber, Julien; van Tits, Lambertus J; Sluka, Susanna; Pasterk, Lisa; Reiner, Martin F; Calatayud, Natacha; Lohmann, Christine; Klingenberg, Roland; Pahla, Jürgen; Vdovenko, Daria; Tanner, Felix C; Camici, Giovanni G; Eriksson, Urs; Auwerx, Johan; Mach, François; Windecker, Stephan; Rodondi, Nicolas; Lüscher, Thomas F; Winnik, Stephan; Matter, Christian M

2018-01-01

Abstract Aims Sirtuin 3 (Sirt3) is a mitochondrial, nicotinamide adenine dinucleotide (NAD+)-dependent deacetylase that reduces oxidative stress by activation of superoxide dismutase 2 (SOD2). Oxidative stress enhances arterial thrombosis. This study investigated the effects of genetic Sirt3 deletion on arterial thrombosis in mice in an inflammatory setting and assessed the clinical relevance of these findings in patients with ST-elevation myocardial infarction (STEMI). Methods and results Using a laser-induced carotid thrombosis model with lipopolysaccharide (LPS) challenge, in vivo time to thrombotic occlusion in Sirt3−/− mice (n = 6) was reduced by half compared to Sirt3+/+ wild-type (n = 8, P < 0.01) controls. Ex vivo analyses of whole blood using rotational thromboelastometry revealed accelerated clot formation and increased clot stability in Sirt3−/− compared to wild-type blood. rotational thromboelastometry of cell-depleted plasma showed accelerated clotting initiation in Sirt3−/− mice, whereas overall clot formation and firmness remained unaffected. Ex vivo LPS-induced neutrophil extracellular trap formation was increased in Sirt3−/− bone marrow-derived neutrophils. Plasma tissue factor (TF) levels and activity were elevated in Sirt3−/− mice, whereas plasma levels of other coagulation factors and TF expression in arterial walls remained unchanged. SOD2 expression in bone marrow -derived Sirt3−/− neutrophils was reduced. In STEMI patients, transcriptional levels of Sirt3 and its target SOD2 were lower in CD14+ leukocytes compared with healthy donors (n = 10 each, P < 0.01). Conclusions Sirt3 loss-of-function enhances experimental thrombosis in vivo via an increase of neutrophil extracellular traps and elevation of TF suggesting thrombo-protective effects of endogenous Sirt3. Acute coronary thrombosis in STEMI patients is associated with lower expression levels of SIRT3 and SOD2 in CD14+ leukocytes. Therefore, enhancing SIRT3 activity by pan-sirtuin activating NAD+-boosters may provide a novel therapeutic target to prevent or treat thrombotic arterial occlusion in myocardial infarction or stroke. PMID:29444200

Backscattering enhancement factor dependence of a Laguerre-Gaussian laser beam propagating on the location path in the atmosphere on optical turbulence intensity

NASA Astrophysics Data System (ADS)

Rytchkov, D. S.

2017-11-01

The paper presents the results of a study of the backscattering enhancement factor (BSE) dependence of vortex LaguerreGaussian beams propagating on monostatic location paths in the atmosphere on optical turbulence intensity. The numeric simulation split-step method of laser beam propagation was used to obtain BSE factor values of a laser beam propagated on monostatic location path in the turbulent atmosphere and reflected from a diffuse target. It is shown that BSE factor of the averaged intensity of a backscattered vortex laser beam of any topological charge is less than BSE factor values of backscattered Gaussian beam in arbitrary turbulent conditions.

Interleukin-4 but not interleukin-10 inhibits the production of leukemia inhibitory factor by rheumatoid synovium and synoviocytes.

PubMed

Dechanet, J; Taupin, J L; Chomarat, P; Rissoan, M C; Moreau, J F; Banchereau, J; Miossec, P

1994-12-01

The expression of the proinflammatory cytokine leukemia inhibitory factor (LIF) has been reported in the cartilage and synovium of rheumatoid arthritis (RA) patients. Here, we show that high levels of LIF were constitutively produced by cultures of synovium pieces. Low levels of LIF were produced spontaneously by isolated synoviocytes, but interleukin (IL)-1 beta caused a fourfold enhancement of this secretion. The anti-inflammatory cytokine IL-4 reduced the production of LIF by synovium pieces by 75%, as observed earlier with IL-6, IL-1 beta and tumor necrosis factor (TNF)-alpha. IL-4 had a direct effect since it inhibited LIF production by unstimulated and IL-1 beta- or TNF-alpha-stimulated synoviocytes. Conversely, IL-4 enhanced the production of IL-6, which shares with LIF biological activities and receptor components. The inhibitory effect of IL-4 was dose dependent and was reversed using a blocking anti-IL-4 receptor antibody. Similar inhibitory action of IL-4 on LIF production was observed on synovium pieces from patients with osteoarthritis and on normal synoviocytes. IL-10, another anti-inflammatory cytokine acting on monocytes, had no effect on LIF production by either synovium pieces or isolated synoviocytes. Thus, the production of LIF by synovium tissue was inhibited by IL-4 through both a direct effect on synoviocytes and an indirect effect by inhibition of the production of LIF-inducing cytokines.

Resistin facilitates VEGF-C-associated lymphangiogenesis by inhibiting miR-186 in human chondrosarcoma cells.

PubMed

Su, Chen-Ming; Tang, Chih-Hsin; Chi, Meng-Ju; Lin, Chih-Yang; Fong, Yi-Chin; Liu, Yueh-Ching; Chen, Wei-Cheng; Wang, Shih-Wei

2018-05-03

Chondrosarcoma is a common primary malignant tumor of the bone that can metastasize through the vascular system to other organs. A key step in the metastatic process, lymphangiogenesis, involves vascular endothelial growth factor-C (VEGF-C). However, the effects of lymphangiogenesis in chondrosarcoma metastasis remain to be clarified. Accumulating evidence shows that resistin, a cytokine secreted from adipocytes and monocytes, also promotes tumor pathogenesis. Notably, chondrosarcoma can easily metastasize. In this study, we demonstrate that resistin enhances VEGF-C expression and lymphatic endothelial cells (LECs)-associated lymphangiogenesis in human chondrosarcoma cells. We also show that resistin triggers VEGF-C-dependent lymphangiogenesis via the c-Src signaling pathway and down-regulating micro RNA (miR)-186. Overexpression of resistin in chondrosarcoma cells significantly enhanced VEGF-C production and LECs-associated lymphangiogenesis in vitro and tumor-related lymphangiogenesis in vivo. Resistin levels were positively correlated with VEGF-C-dependent lymphangiogenesis via the down-regulation of miR-186 expression in clinical samples from chondrosarcoma tissue. This study is the first to evaluate the mechanism underlying resistin-induced promotion of LECs-associated lymphangiogenesis via the upregulation of VEGF-C expression in human chondrosarcomas. We suggest that resistin may represent a molecular target in VEGF-C-associated tumor lymphangiogenesis in chondrosarcoma metastasis. Copyright © 2018 Elsevier Inc. All rights reserved.

Continued clearance of apoptotic cells critically depends on the phagocyte Ucp2 protein.

PubMed

Park, Daeho; Han, Claudia Z; Elliott, Michael R; Kinchen, Jason M; Trampont, Paul C; Das, Soumita; Collins, Sheila; Lysiak, Jeffrey J; Hoehn, Kyle L; Ravichandran, Kodi S

2011-08-21

Rapid and efficient removal of apoptotic cells by phagocytes is important during development, tissue homeostasis and in immune responses. Efficient clearance depends on the capacity of a single phagocyte to ingest multiple apoptotic cells successively, and to process the corpse-derived cellular material. However, the factors that influence continued clearance by phagocytes are not known. Here we show that the mitochondrial membrane potential of the phagocyte critically controls engulfment capacity, with lower potential enhancing engulfment and vice versa. The mitochondrial membrane protein Ucp2, which acts to lower the mitochondrial membrane potential, was upregulated in phagocytes engulfing apoptotic cells. Loss of Ucp2 reduced phagocytic capacity, whereas Ucp2 overexpression enhanced engulfment. Mutational and pharmacological studies indicated a direct role for Ucp2-mediated mitochondrial function in phagocytosis. Macrophages from Ucp2-deficient mice were impaired in phagocytosis in vitro, and Ucp2-deficient mice showed profound in vivo defects in clearing dying cells in the thymus and testes. Collectively, these data indicate that mitochondrial membrane potential and Ucp2 are key molecular determinants of apoptotic cell clearance. As Ucp2 is linked to metabolic diseases and atherosclerosis, this newly discovered role for Ucp2 in apoptotic cell clearance has implications for the complex aetiology and pathogenesis of these diseases.

Enhanced migration of tissue inhibitor of metalloproteinase overexpressing hepatoma cells is attributed to gelatinases: Relevance to intracellular signaling pathways

PubMed Central

Roeb, Elke; Bosserhoff, Anja-Katrin; Hamacher, Sabine; Jansen, Bettina; Dahmen, Judith; Wagner, Sandra; Matern, Siegfried

2005-01-01

AIM: To study the effect of gelatinases (especially MMP-9) on migration of tissue inhibitor of metalloproteinase (TIMP-1) overexpressing hepatoma cells. METHODS: Wild type HepG2 cells, cells stably transfected with TIMP-1 and TIMP-1 antagonist (MMP-9-H401A, a catalytically inactive matrix metalloproteinase (MMP) which still binds and neutralizes TIMP-1) were incubated in Boyden chambers either with or without Galardin (a synthetic inhibitor of MMP-1, -2, -3, -8, -9) or a specific inhibitor of gelatinases. RESULTS: Compared to wild type HepG2 cells, the cells overexpressing TIMP-1 showed 115% migration (P<0.05) and the cells overexpressing MMP-9-H401A showed 62% migration (P<0.01). Galardin reduced cell migration dose dependently in all cases. The gelatinase inhibitor reduced migration in TIMP-1 overexpressing cells predominantly. Furthermore, we examined intracellular signal transduction pathways of TIMP-1-dependent HepG2 cells. TIMP-1 deactivates cell signaling pathways of MMP-2 and MMP-9 involving p38 mitogen-activated protein kinase. Specific blockade of the ERK pathway suppresses gelatinase expression either in the presence or absence of TIMP-1. CONCLUSION: Overexpressing functional TIMP-1- enhanced migration of HepG2-TIMP-1 cells depends on enhanced MMP-activity, especially MMP-9. PMID:15754388

Delivering heparin-binding insulin-like growth factor 1 with self-assembling peptide hydrogels.

PubMed

Florine, Emily M; Miller, Rachel E; Liebesny, Paul H; Mroszczyk, Keri A; Lee, Richard T; Patwari, Parth; Grodzinsky, Alan J

2015-02-01

Heparin-binding insulin-like growth factor 1 (HB-IGF-1) is a fusion protein of IGF-1 with the HB domain of heparin-binding epidermal growth factor-like growth factor. A single dose of HB-IGF-1 has been shown to bind specifically to cartilage and to promote sustained upregulation of proteoglycan synthesis in cartilage explants. Achieving strong integration between native cartilage and tissue-engineered cartilage remains challenging. We hypothesize that if a growth factor delivered by the tissue engineering scaffold could stimulate enhanced matrix synthesis by both the cells within the scaffold and the adjacent native cartilage, integration could be enhanced. In this work, we investigated methods for adsorbing HB-IGF-1 to self-assembling peptide hydrogels to deliver the growth factor to encapsulated chondrocytes and cartilage explants cultured with growth factor-loaded hydrogels. We tested multiple methods for adsorbing HB-IGF-1 in self-assembling peptide hydrogels, including adsorption prior to peptide assembly, following peptide assembly, and with/without heparan sulfate (HS, a potential linker between peptide molecules and HB-IGF-1). We found that HB-IGF-1 and HS were retained in the peptide for all tested conditions. A subset of these conditions was then studied for their ability to stimulate increased matrix production by gel-encapsulated chondrocytes and by chondrocytes within adjacent native cartilage. Adsorbing HB-IGF-1 or IGF-1 prior to peptide assembly was found to stimulate increased sulfated glycosaminoglycan per DNA and hydroxyproline content of chondrocyte-seeded hydrogels compared with basal controls at day 10. Cartilage explants cultured adjacent to functionalized hydrogels had increased proteoglycan synthesis at day 10 when HB-IGF-1 was adsorbed, but not IGF-1. We conclude that delivery of HB-IGF-1 to focal defects in cartilage using self-assembling peptide hydrogels is a promising technique that could aid cartilage repair via enhanced matrix production and integration with native tissue.

The orphan nuclear receptor Nur77 regulates decidual prolactin expression in human endometrial stromal cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Jiang, Yue; Hu, Yali; Zhao, Jing

2011-01-14

Research highlights: {yields} Decidually produced PRL plays a key role during pregnancy. {yields} Overexpression of Nur77 increased PRL mRNA expression and enhanced decidual PRL promoter activity. {yields} Knockdown of Nur77 decreased decidual PRL secretion induced by 8-Br-cAMP and MPA. {yields} Nur77 is a novel transcription factor that plays an active role in decidual prolactin expression. -- Abstract: Prolactin (PRL) is synthesized and released by several extrapituitary tissues, including decidualized stromal cells. Despite the important role of decidual PRL during pregnancy, little is understood about the factors involved in the proper regulation of decidual PRL expression. Here we present evidence thatmore » the transcription factor Nur77 plays an active role in decidual prolactin expression in human endometrial stromal cells (hESCs). Nur77 mRNA expression in hESCs was significantly increased after decidualization stimulated by 8-Br-cAMP and medroxyprogesterone acetate (MPA). Adenovirus-mediated overexpression of Nur77 in hESCs markedly increased PRL mRNA expression and enhanced decidual PRL promoter (dPRL/-332Luc) activity in a concentration-dependent manner. Furthermore, knockdown of Nur77 in hESCs significantly decreased decidual PRL promoter activation and substantially attenuated PRL mRNA expression and PRL secretion (P < 0.01) induced by 8-Br-cAMP and MPA. These results demonstrate that Nur77 is a novel transcription factor that contributes significantly to the regulation of prolactin gene expression in human endometrial stromal cells.« less

Re-engineering the Pancreas Tumor Microenvironment: A "Regenerative Program" Hacked.

PubMed

Evan, Gerard I; Hah, Nasun; Littlewood, Trevor D; Sodir, Nicole M; Campos, Tania; Downes, Michael; Evans, Ronald M

2017-04-01

The "hallmarks" of pancreatic ductal adenocarcinoma (PDAC) include proliferative, invasive, and metastatic tumor cells and an associated dense desmoplasia comprised of fibroblasts, pancreatic stellate cells, extracellular matrix, and immune cells. The oncogenically activated pancreatic epithelium and its associated stroma are obligatorily interdependent, with the resulting inflammatory and immunosuppressive microenvironment contributing greatly to the evolution and maintenance of PDAC. The peculiar pancreas-specific tumor phenotype is a consequence of oncogenes hacking the resident pancreas regenerative program, a tissue-specific repair mechanism regulated by discrete super enhancer networks. Defined as genomic regions containing clusters of multiple enhancers, super enhancers play pivotal roles in cell/tissue specification, identity, and maintenance. Hence, interfering with such super enhancer-driven repair networks should exert a disproportionately disruptive effect on tumor versus normal pancreatic tissue. Novel drugs that directly or indirectly inhibit processes regulating epigenetic status and integrity, including those driven by histone deacetylases, histone methyltransferase and hydroxylases, DNA methyltransferases, various metabolic enzymes, and bromodomain and extraterminal motif proteins, have shown the feasibility of disrupting super enhancer-dependent transcription in treating multiple tumor types, including PDAC. The idea that pancreatic adenocarcinomas rely on embedded super enhancer transcriptional mechanisms suggests a vulnerability that can be potentially targeted as novel therapies for this intractable disease. Clin Cancer Res; 23(7); 1647-55. ©2017 AACR See all articles in this CCR Focus section, "Pancreatic Cancer: Challenge and Inspiration." ©2017 American Association for Cancer Research.

Gut memories do not fade: epigenetic regulation of lasting gut homing receptor expression in CD4+ memory T cells.

PubMed

Szilagyi, B A; Triebus, J; Kressler, C; de Almeida, M; Tierling, S; Durek, P; Mardahl, M; Szilagyi, A; Floess, S; Huehn, J; Syrbe, U; Walter, J; Polansky, J K; Hamann, A

2017-11-01

The concept of a "topographical memory" in lymphocytes implies a stable expression of homing receptors mediating trafficking of lymphocytes back to the tissue of initial activation. However, a significant plasticity of the gut-homing receptor α 4 β 7 was found in CD8 + T cells, questioning the concept. We now demonstrate that α 4 β 7 expression in murine CD4 + memory T cells is, in contrast, imprinted and remains stable in the absence of the inducing factor retinoic acid (RA) or other stimuli from mucosal environments. Repetitive rounds of RA treatment enhanced the stability of de novo induced α 4 β 7 . A novel enhancer element in the murine Itga4 locus was identified that showed, correlating to stability, selective DNA demethylation in mucosa-seeking memory cells and methylation-dependent transcriptional activity in a reporter gene assay. This implies that epigenetic mechanisms contribute to the stabilization of α 4 β 7 expression. Analogous DNA methylation patterns could be observed in the human ITGA4 locus, suggesting that its epigenetic regulation is conserved between mice and men. These data prove that mucosa-specific homing mediated by α 4 β 7 is imprinted in CD4 + memory T cells, reinstating the validity of the concept of "topographical memory" for mucosal tissues, and imply a critical role of epigenetic mechanisms.

Micafungin Enhances the Human Macrophage Response to Candida albicans through β-Glucan Exposure.

PubMed

Guirao-Abad, José Pedro; Sánchez-Fresneda, Ruth; Machado, Francisco; Argüelles, Juan Carlos; Martínez-Esparza, María

2018-05-01

Micafungin belongs to the antifungal family of echinocandins, which act as noncompetitive inhibitors of the fungal cell wall β-1,3-d-glucan synthase. Since Candida albicans is the most prevalent pathogenic fungus in humans, we study the involvement of micafungin in the modulation of the inflammatory response developed by human tissue macrophages against C. albicans The MIC for micafungin was 0.016 μg/ml on the C. albicans SC5314 standard strain. Micafungin induced a drastic reduction in the number of exponential SC5314 viable cells, with the fungicidal effect being dependent on the cellular metabolic activity. Notably, micafungin also caused a structural remodelling of the cell wall, leading to exposure of the β-glucan and chitin content on the external surface. At the higher doses used (0.05 μg/ml), the antifungal also induced the blowing up of budding yeasts. In addition, preincubation with micafungin before exposure to human tissue macrophages enhanced the secretion of tumor necrosis factor alpha (TNF-α), interleukin-17A (IL-17A), and IL-10 cytokines. Our results strongly suggest that in C. albicans treatment with micafungin, in addition to having the expected toxic antifungal effect, it potentiates the immune response, improving the interaction and activation of human macrophages, probably through the unmasking of β-glucans on the cell wall surface. Copyright © 2018 American Society for Microbiology.

Environmentally Clean Mitigation of Undesirable Plant Life Using Lasers

DOE Office of Scientific and Technical Information (OSTI.GOV)

Rubenchik, A M; McGrann, T J; Yamamoto, R M

This concept comprises a method for environmentally clean destruction of undesirable plant life using visible or infrared radiation. We believe that during the blossom stage, plant life is very sensitive to electromagnetic radiation, with an enhanced sensitivity to specific spectral ranges. Small doses of irradiation can arrest further plant growth, cause flower destruction or promote plant death. Surrounding plants, which are not in the blossoming stage, should not be affected. Our proposed mechanism to initiate this effect is radiation produced by a laser. Tender parts of the blossom possess enhanced absorptivity in some spectral ranges. This absorption can increase themore » local tissue temperature by several degrees, which is sufficient to induce bio-tissue damage. In some instances, the radiation may actually stimulate plant growth, as an alternative for use in increased crop production. This would be dependent on factors such as plant type, the wavelength of the laser radiation being used and the amount of the radiation dose. Practical, economically viable realization of this concept is possible today with the advent of high efficiency, compact and powerful laser diodes. The laser diodes provide an efficient, environmentally clean source of radiation at a variety of power levels and radiation wavelengths. Figure 1 shows the overall concept, with the laser diodes mounted on a movable platform, traversing and directing the laser radiation over a field of opium poppies.« less

Is NAA reduction in normal contralateral cerebral tissue in stroke patients dependent on underlying risk factors?

PubMed

Walker, P M; Ben Salem, D; Giroud, M; Brunotte, F

2006-05-01

This retrospective study investigated the dependence of N-acetyl aspartate (NAA) ratios on risk factors for cerebral vasculopathy such as sex, age, hypertension, diabetes mellitus, carotid stenosis, and dyslipidaemia, which may have affected brain vessels and induced metabolic brain abnormalities prior to stroke. We hypothesise that in stroke patients metabolic alterations in the apparently normal contralateral brain are dependent on the presence or not of such risk factors. Fifty nine patients (31 male, 28 female: 58.8+/-16.1 years old) with cortical middle cerebral artery (MCA) territory infarction were included. Long echo time chemical shift imaging spectroscopy was carried out on a Siemens 1.5 T Magnetom Vision scanner using a multi-voxel PRESS technique. Metabolite ratios (NAA/choline, NAA/creatine, lactate/choline, etc) were studied using uni- and multivariate analyses with respect to common risk factors. The influence of age, stroke lesion size, and time since stroke was studied using a linear regression approach. Age, sex, and hypertension all appeared to individually influence metabolite ratios, although only hypertension was significant after multivariate analysis. In both basal ganglia and periventricular white matter regions in apparently normal contralateral brain, the NAA/choline ratio was significantly lower in hypertensive (1.37+/-0.16 and 1.50+/-0.19, respectively) than in normotensive patients (1.72+/-0.19 and 1.85+/-0.15, respectively). Regarding MCA infarction, contralateral tissue remote from the lesion behaves abnormally in the presence of hypertension, the NAA ratios in hypertensive patients being significantly lower. These data suggest that hypertension may compromise the use of contralateral tissue data as a reference for comparison with ischaemic tissue.

Virulence arsenal of the most pathogenic species among the Gram-positive anaerobic cocci, Finegoldia magna.

PubMed

Boyanova, Lyudmila; Markovska, Rumyana; Mitov, Ivan

2016-12-01

This review focuses on the virulence arsenal of the most pathogenic species among Gram positive anaerobic cocci, Finegoldia magna according to recently published data from 2012 to 2016. Virulence factors like sortase dependent pili and F. magna adhesion factor (FAF) facilitate the start of the infection. Albumin binding protein (PAB) enhances F. magna survival. FAF, subtilisin-like extracellular serine protease (SufA) and superantigen protein L protect the bacteria from factors of innate defense system. SufA, capsule and tissue-destroying enzymes provide a deep penetration or spread of the infections and the protein L is associated with infection severity. Biofilm production results in infection chronification and complicated treatment as well as to persistence of multi-species biofilms. Resistance rates to quinolones (13.0->70%) and clindamycin (0-40.0%) are important, and resistance to penicillins (<4%), chloramphenicol (7.0%) and metronidazole (<7%) has been reported. F. magna should not be overlooked when present in monoinfections or mixed infections in humans. Copyright © 2016 Elsevier Ltd. All rights reserved.

Adipose Tissue Plasticity During Catch-Up Fat Driven by Thrifty Metabolism

PubMed Central

Summermatter, Serge; Marcelino, Helena; Arsenijevic, Denis; Buchala, Antony; Aprikian, Olivier; Assimacopoulos-Jeannet, Françoise; Seydoux, Josiane; Montani, Jean-Pierre; Solinas, Giovanni; Dulloo, Abdul G.

2009-01-01

OBJECTIVE Catch-up growth, a risk factor for later type 2 diabetes, is characterized by hyperinsulinemia, accelerated body-fat recovery (catch-up fat), and enhanced glucose utilization in adipose tissue. Our objective was to characterize the determinants of enhanced glucose utilization in adipose tissue during catch-up fat. RESEARCH DESIGN AND METHODS White adipose tissue morphometry, lipogenic capacity, fatty acid composition, insulin signaling, in vivo glucose homeostasis, and insulinemic response to glucose were assessed in a rat model of semistarvation-refeeding. This model is characterized by glucose redistribution from skeletal muscle to adipose tissue during catch-up fat that results solely from suppressed thermogenesis (i.e., without hyperphagia). RESULTS Adipose tissue recovery during the dynamic phase of catch-up fat is accompanied by increased adipocyte number with smaller diameter, increased expression of genes for adipogenesis and de novo lipogenesis, increased fatty acid synthase activity, increased proportion of saturated fatty acids in triglyceride (storage) fraction but not in phospholipid (membrane) fraction, and no impairment in insulin signaling. Furthermore, it is shown that hyperinsulinemia and enhanced adipose tissue de novo lipogenesis occur concomitantly and are very early events in catch-up fat. CONCLUSIONS These findings suggest that increased adipose tissue insulin stimulation and consequential increase in intracellular glucose flux play an important role in initiating catch-up fat. Once activated, the machinery for lipogenesis and adipogenesis contribute to sustain an increased insulin-stimulated glucose flux toward fat storage. Such adipose tissue plasticity could play an active role in the thrifty metabolism that underlies glucose redistribution from skeletal muscle to adipose tissue. PMID:19602538

Collapse of proteostasis represents an early molecular event in Caenorhabditis elegans aging.

PubMed

Ben-Zvi, Anat; Miller, Elizabeth A; Morimoto, Richard I

2009-09-01

Protein damage contributes prominently to cellular aging. To address whether this occurs at a specific period during aging or accumulates gradually, we monitored the biochemical, cellular, and physiological properties of folding sensors expressed in different tissues of C. elegans. We observed the age-dependent misfolding and loss of function of diverse proteins harboring temperature-sensitive missense mutations in all somatic tissues at the permissive condition. This widespread failure in proteostasis occurs rapidly at an early stage of adulthood, and coincides with a severely reduced activation of the cytoprotective heat shock response and the unfolded protein response. Enhancing stress responsive factors HSF-1 or DAF-16 suppresses misfolding of these metastable folding sensors and restores the ability of the cell to maintain a functional proteome. This suggests that a compromise in the regulation of proteostatic stress responses occurs early in adulthood and tips the balance between the load of damaged proteins and the proteostasis machinery. We propose that the collapse of proteostasis represents an early molecular event of aging that amplifies protein damage in age-associated diseases of protein conformation.

Surgical stapling device-tissue interactions: what surgeons need to know to improve patient outcomes.

PubMed

Chekan, Edward; Whelan, Richard L

2014-01-01

The introduction of both new surgical devices and reengineered existing devices leads to modifications in the way traditional tasks are carried out and allows for the development of new surgical techniques. Each new device has benefits and limitations in regards to tissue interactions that, if known, allow for optimal use. However, most surgeons are unaware of these attributes and, therefore, new device introduction creates a "knowledge gap" that is potentially dangerous. The goal of this review is to present a framework for the study of device- tissue interactions and to initiate the process of "filling in" the knowledge gap via the available literature. Surgical staplers, which are continually being developed, are the focus of this piece. The integrity of the staple line, which depends on adequate tissue compression, is the primary factor in creating a stable anastomosis. This review focuses on published studies that evaluated the creation of stable anastomoses in bariatric, thoracic, and colorectal procedures. Understanding how staplers interact with target tissues is key to improving patient outcomes. It is clear from this review that each tissue type presents unique challenges. The thickness of each tissue varies as do the intrinsic biomechanical properties that determine the ideal compressive force and prefiring compression time for each tissue type. The correct staple height will vary depending on these tissue-specific properties and the tissue pathology. These studies reinforce the universal theme that compression, staple height, tissue thickness, tissue compressibility, and tissue type must all be considered by the surgeon prior to choosing a stapler and cartridge. The surgeon's experience, therefore, is a critical factor. Educational programs need to be established to inform and update surgeons on the characteristics of each stapler. It is hoped that the framework presented in this review will facilitate this process.

Effect of long-term treatment with urocortin on the activity of somatic angiotensin-converting enzyme in spontaneously hypertensive rats.

PubMed

Yang, Cui; Liu, Xiuxia; Li, Shengnan

2010-02-01

Our previous acute study on urocortin (Ucn) demonstrated that Ucn altered serum and tissue angiotensin-converting enzyme (ACE) activity in rats. Therefore, the present investigation was designed to explore the effect of long-term treatment with Ucn on somatic ACE (sACE) and other components of the renin-angiotensin system (RAS). After 8 weeks of intravenous administration of Ucn in spontaneously hypertensive rats (SHR), serum and tissue sACE, angiotensin II (Ang II), nitric oxide (NO), Ang-(1-7), and tissue chymase activities were evaluated. RT-PCR analysis was performed to determine the quantity of tissue sACE mRNA. Serum sACE activity was reduced by Ucn, although tissue sACE activity and tissue sACE mRNA were elevated. Chymase activity was observed to be enhanced by Ucn, whereas the ACE inhibitor enalapril failed to influence chymase. Serum and tissue Ang II activity was reduced, but NO and Ang-(1-7) production was increased in a concentration-dependent manner after Ucn treatment. Meanwhile, a significant decrease of the systolic blood pressure (SBP) was observed after the long-term Ucn administration, and there was a significant positive correlation (r2 = 0.6993) between serum ACE activity and SBP. Pretreatment with the corticotropin-releasing factor (CRF) blocker astressin and the extracellular signal-regulated kinase 1/2 (ERK1/2) pathway blocker PD98059 abolished these effects of Ucn. Our findings further support the hypothesis that the changes of sACE activity and the production of other RAS components may play roles in the vasodilatory property of Ucn via the activation of the ERK1/2 pathway.

Thymosin β4 promotes endothelial progenitor cell angiogenesis via a vascular endothelial growth factor‑dependent mechanism.

PubMed

Zhao, Yanbo; Song, Jiale; Bi, Xukun; Gao, Jing; Shen, Zhida; Zhu, Junhui; Fu, Guosheng

2018-06-20

Endothelial progenitor cells (EPCs) are a promising cell source for tissue repair and regeneration, predominantly through angiogenesis promotion. Paracrine functions serve a pivotal role in EPC‑mediated angiogenesis, which may be impaired by various cardiovascular risk factors. Therefore, it is important to identify a solution that optimizes the paracrine function of EPCs. Thymosin β4 (Tβ4) is a peptide with the potential to promote tissue regeneration and wound healing. A previous study demonstrated that Tβ4 enhances the EPC‑mediated angiogenesis of the ischemic myocardium. In the present study, whether Tβ4 improved angiogenesis by enhancing the paracrine effects of EPCs was investigated. A tube formation assay was used to assess the effect of angiogenesis, and the paracrine effects were measured using an ELISA kit. The results indicated that Tβ4 improved the paracrine effects of EPCs, evidenced by an increase in the expression of vascular endothelial growth factor (VEGF). EPC‑conditioned medium (EPC‑CM) significantly promoted human umbilical vein endothelial cell angiogenesis in vitro, which was further enhanced by pretreatment with Tβ4. The effect of Tβ4 pretreated EPC‑CM on angiogenesis was abolished by VEGF neutralizing antibody in vitro, indicating that increased VEGF secretion had a pivotal role in Tβ4‑mediated EPC angiogenesis. Furthermore, transplantation of EPCs pretreated with Tβ4 into infarcted rat hearts resulted in significantly higher VEGF expression in the border zone, compared with EPC transplantation alone. To further investigate whether the Akt/eNOS pathway was involved in Tβ4‑induced VEGF secretion in EPCs, the expression levels of VEGF in EPC‑CM were significantly decreased following knockdown of Akt or eNOS by small interfering RNA transfection. In conclusion, Tβ4 significantly increased angiogenesis by enhancing the paracrine effects of EPCs, evidenced by the increased expression of VEGF. The RAC‑α serine/threonine‑protein kinase/endothelial nitric oxide synthase signal transduction pathway was involved in the regulation of Tβ4‑induced VEGF secretion in EPCs. Further studies are required to investigate the long‑term prognosis of patients with coronary heart disease following Tβ4‑pretreated EPC transplantation.

Nutrient/TOR-dependent regulation of RNA polymerase III controls tissue and organismal growth in Drosophila

PubMed Central

Marshall, Lynne; Rideout, Elizabeth J; Grewal, Savraj S

2012-01-01

The nutrient/target-of-rapamycin (TOR) pathway has emerged as a key regulator of tissue and organismal growth in metazoans. The signalling components of the nutrient/TOR pathway are well defined; however, the downstream effectors are less understood. Here, we show that the control of RNA polymerase (Pol) III-dependent transcription is an essential target of TOR in Drosophila. We find that TOR activity controls Pol III in growing larvae via inhibition of the repressor Maf1 and, in part, via the transcription factor Drosophila Myc (dMyc). Moreover, we show that loss of the Pol III factor, Brf, leads to reduced tissue and organismal growth and prevents TOR-induced cellular growth. TOR activity in the larval fat body, a tissue equivalent to vertebrate fat or liver, couples nutrition to insulin release from the brain. Accordingly, we find that fat-specific loss of Brf phenocopies nutrient limitation and TOR inhibition, leading to decreased systemic insulin signalling and reduced organismal growth. Thus, stimulation of Pol III is a key downstream effector of TOR in the control of cellular and systemic growth. PMID:22367393

Insulin-like Growth Factor-I and Slow, Bi-directional Perfusion Enhance the Formation of Tissue-Engineered Cardiac Grafts

PubMed Central

Cheng, Mingyu; Moretti, Matteo; Engelmayr, George C.

2009-01-01

Biochemical and mechanical signals enabling cardiac regeneration can be elucidated using in vitro tissue-engineering models. We hypothesized that insulin-like growth factor-I (IGF) and slow, bi-directional perfusion could act independently and interactively to enhance the survival, differentiation, and contractile performance of tissue-engineered cardiac grafts. Heart cells were cultured on three-dimensional porous scaffolds in medium with or without supplemental IGF and in the presence or absence of slow, bi-directional perfusion that enhanced transport and provided shear stress. Structural, molecular, and electrophysiologic properties of the resulting grafts were quantified on culture day 8. IGF had independent, beneficial effects on apoptosis (p < 0.01), cellular viability (p < 0.01), contractile amplitude (p < 0.01), and excitation threshold (p < 0.01). Perfusion independently affected the four aforementioned parameters and also increased amounts of cardiac troponin-I (p < 0.01), connexin-43 (p < 0.05), and total protein (p < 0.01) in the grafts. Interactive effects of IGF and perfusion on apoptosis were also present (p < 0.01). Myofibrillogenesis and spontaneous contractility were present only in grafts cultured with perfusion, although contractility was inducible by electrical field stimulation of grafts from all groups. Our findings demonstrate that multi-factorial stimulation of tissue-engineered cardiac grafts using IGF and perfusion resulted in independent and interactive effects on heart cell survival, differentiation, and contractility. PMID:18759675

A mutation within the SH2 domain of slp-76 regulates the tissue distribution and cytokine production of iNKT cells in mice.

PubMed

Danzer, Claudia; Koller, Anna; Baier, Julia; Arnold, Harald; Giessler, Claudia; Opoka, Robert; Schmidt, Stephanie; Willers, Maike; Mihai, Sidonia; Parsch, Hans; Wirtz, Stefan; Daniel, Christoph; Reinhold, Annegret; Engelmann, Swen; Kliche, Stefanie; Bogdan, Christian; Hoebe, Kasper; Mattner, Jochen

2016-09-01

TCR ligation is critical for the selection, activation, and integrin expression of T lymphocytes. Here, we explored the role of the TCR adaptor protein slp-76 on iNKT-cell biology. Compared to B6 controls, slp-76(ace/ace) mice carrying a missense mutation (Thr428Ile) within the SH2-domain of slp-76 showed an increase in iNKT cells in the thymus and lymph nodes, but a decrease in iNKT cells in spleens and livers, along with reduced ADAP expression and cytokine response. A comparable reduction in iNKT cells was observed in the livers and spleens of ADAP-deficient mice. Like ADAP(-/-) iNKT cells, slp-76(ace/ace) iNKT cells were characterized by enhanced CD11b expression, correlating with an impaired induction of the TCR immediate-early gene Nur77 and a decreased adhesion to ICAM-1. Furthermore, CD11b-intrinsic effects inhibited cytokine release, concanavalin A-mediated inflammation, and iNKT-cell accumulation in the liver. Unlike B6 and ADAP(-/-) mice, the expression of the transcription factors Id3 and PLZF was reduced, whereas NP-1-expression was enhanced in slp-76(ace/ace) mice. Blockade of NP-1 decreased the recovery of iNKT cells from peripheral lymph nodes, identifying NP-1 as an iNKT-cell-specific adhesion factor. Thus, slp-76 contributes to the regulation of the tissue distribution, PLZF, and cytokine expression of iNKT cells via ADAP-dependent and -independent mechanisms. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Impact of Tissue Factor Localization on Blood Clot Structure and Resistance under Venous Shear.

PubMed

Govindarajan, Vijay; Zhu, Shu; Li, Ruizhi; Lu, Yichen; Diamond, Scott L; Reifman, Jaques; Mitrophanov, Alexander Y

2018-02-27

The structure and growth of a blood clot depend on the localization of tissue factor (TF), which can trigger clotting during the hemostatic process or promote thrombosis when exposed to blood under pathological conditions. We sought to understand how the growth, structure, and mechanical properties of clots under flow are shaped by the simultaneously varying TF surface density and its exposure area. We used an eight-channel microfluidic device equipped with a 20- or 100-μm-long collagen surface patterned with lipidated TF of surface densities ∼0.1 and ∼2 molecules/μm 2 . Human whole blood was perfused at venous shear, and clot growth was continually measured. Using our recently developed computational model of clot formation, we performed simulations to gain insights into the clot's structure and its resistance to blood flow. An increase in TF exposure area resulted not only in accelerated bulk platelet, thrombin, and fibrin accumulation, but also in increased height of the platelet mass and increased clot resistance to flow. Moreover, increasing the TF surface density or exposure area enhanced platelet deposition by approximately twofold, and thrombin and fibrin generation by greater than threefold, thereby increasing both clot size and its viscous resistance. Finally, TF effects on blood flow occlusion were more pronounced for the longer thrombogenic surface than for the shorter one. Our results suggest that TF surface density and its exposure area can independently enhance both the clot's occlusivity and its resistance to blood flow. These findings provide, to our knowledge, new insights into how TF affects thrombus growth in time and space under flow. Copyright © 2018 Biophysical Society. All rights reserved.

Surfactants, not size or zeta-potential influence blood-brain barrier passage of polymeric nanoparticles.

PubMed

Voigt, Nadine; Henrich-Noack, Petra; Kockentiedt, Sarah; Hintz, Werner; Tomas, Jürgen; Sabel, Bernhard A

2014-05-01

Nanoparticles (NP) can deliver drugs across the blood-brain barrier (BBB), but little is known which of the factors surfactant, size and zeta-potential are essential for allowing BBB passage. To this end we designed purpose-built fluorescent polybutylcyanoacrylate (PBCA) NP and imaged the NP's passage over the blood-retina barrier - which is a model of the BBB - in live animals. Rats received intravenous injections of fluorescent PBCA-NP fabricated by mini-emulsion polymerisation to obtain various NP's compositions that varied in surfactants (non-ionic, anionic, cationic), size (67-464nm) and zeta-potential. Real-time imaging of retinal blood vessels and retinal tissue was carried out with in vivo confocal neuroimaging (ICON) before, during and after NP's injection. Successful BBB passage with subsequent cellular labelling was achieved if NP were fabricated with non-ionic surfactants or cationic stabilizers but not when anionic compounds were added. NP's size and charge had no influence on BBB passage and cell labelling. This transport was not caused by an unspecific opening of the BBB because control experiments with injections of unlabelled NP and fluorescent dye (to test a "door-opener" effect) did not lead to parenchymal labelling. Thus, neither NP's size nor chemo-electric charge, but particle surface is the key factor determining BBB passage. This result has important implications for NP engineering in medicine: depending on the surfactant, NP can serve one of two opposite functions: while non-ionic tensides enhance brain up-take, addition of anionic tensides prevents it. NP can now be designed to specifically enhance drug delivery to the brain or, alternatively, to prevent brain penetration so to reduce unwanted psychoactive effects of drugs or prevent environmental nanoparticles from entering tissue of the central nervous system. Copyright © 2014 Elsevier B.V. All rights reserved.

Effects of topographical and mechanical property alterations induced by oxygen plasma modification on stem cell behavior.

PubMed

Yang, Yong; Kulangara, Karina; Lam, Ruby T S; Dharmawan, Rena; Leong, Kam W

2012-10-23

Polymeric substrates intended for cell culture and tissue engineering are often surface-modified to facilitate cell attachment of most anchorage-dependent cell types. The modification alters the surface chemistry and possibly topography. However, scant attention has been paid to other surface property alterations. In studying oxygen plasma treatment of polydimethylsiloxane (PDMS), we show that oxygen plasma treatment alters the surface chemistry and, consequently, the topography and elasticity of PDMS at the nanoscale level. The elasticity factor has the predominant effect, compared with the chemical and topographical factors, on cell adhesions of human mesenchymal stem cells (hMSCs). The enhanced focal adhesions favor cell spreading and osteogenesis of hMSCs. Given the prevalent use of PDMS in biomedical device construction and cell culture experiments, this study highlights the importance of understanding how oxygen plasma treatment would impact subsequent cell-substrate interactions. It helps explain inconsistency in the literature and guides preparation of PDMS-based biomedical devices in the future.

Perspectives on the Role and Relevance of Copper in Cardiac Disease.

PubMed

Medeiros, Denis M

2017-03-01

Cardiac hypertrophy as a result of dietary copper deficiency has been studied for 40 plus years and is the subject of this review. While connective tissue anomalies occur, a hallmark pathology is cardiac hypertrophy, increased mitochondrial biogenesis, with disruptive cristae, vacuolization of mitochondria, and deposition of lipid droplets. Electrocardiogram abnormalities have been demonstrated along with biochemical changes especially as it relates to the copper-containing enzyme cytochrome c oxidase. The master controller of mitochondrial biogenesis, PGC1-α expression and protein, along with other proteins and transcriptional factors that play a role are upregulated. Nitric oxide, vascular endothelial growth factor, and cytochrome c oxidase all may enhance the upregulation of mitochondrial biogenesis. Marginal copper intakes reveal similar pathologies in the absence of cardiac hypertrophy. Reversibility of the copper-deficient rat heart with a copper-replete diet has resulted in mixed results, depending on both the animal model used and temporal relationships. New information has revealed that copper supplementation may rescue cardiac hypertrophy induced by pressure overload.

Ononitol monohydrate enhances PRDM16 & UCP-1 expression, mitochondrial biogenesis and insulin sensitivity via STAT6 and LTB4R in maturing adipocytes.

PubMed

Subash-Babu, P; Alshatwi, Ali A

2018-03-01

Ononitol monohydrate (OMH), a glycoside was originally isolated from Cassia tora (Linn.). Glycosides regulate lipid metabolism but scientific validation desired. Hence, we aimed to evaluate the effect of OMH on enhancing mitochondrial potential, mitochondrial biogenesis, upregulate the expression of brown adipogenesis specific genes in maturing adipocytes. In addition, we observed the inter-relation between adipocyte and T-lymphocyte; whether, OMH treated adipocyte-condition medium stimulate T-cell chemokine linked with insulin resistance. In a dose dependent manner OMH treated to preadipocyte significantly inhibited maturation and enhanced mitochondrial biogenesis, it was confirmed by Oil red 'O and Nile red stain without inducing cytotoxicity. The mRNA levels of adipocyte browning related genes such as, PR domain containing 16 (PRDM16), peroxisome proliferator activated receptor gamma coactivator 1 alpha (PPARγC1α) and uncoupling protein-1 (UCP-1) have been significantly upregulated. In addition, adipogenic transcription factors [such as proliferator activated receptor γ (PPARγ), CCAAT/enhancer binding protein (C/EBPα) and sterol regulatory element binding protein-1c (SREBP-1c)] and adipogenic genes were significantly down-regulated by treatment with OMH when compared to control cells. Protein expression levels of adiponectin have been increased; leptin, C/EBPα and leukotriene B4 receptor (LTB4R) were down regulated by OMH in mature adipocytes. In addition, adipocyte condition medium and OMH treated T-lymphocyte, significantly increased insulin signaling pathway related mRNAs, such as interlukin-4 (IL-4), signal transducer and activator of transcription 6 (STAT 6 ) and decreased leukotriene B4 (LTB 4 ). The present findings suggest that OMH increased browning factors in differentiating and maturing preadipocyte also decreased adipose tissue inflammation as well as the enhanced insulin signaling. Copyright © 2018. Published by Elsevier Masson SAS.

Ischemia postconditioning and mesenchymal stem cells engraftment synergistically attenuate ischemia reperfusion-induced lung injury in rats.

PubMed

Chen, Shuchen; Chen, Liangwan; Wu, Xiaonan; Lin, Jiangbo; Fang, Jun; Chen, Xiangqi; Wei, Shijin; Xu, Jianxin; Gao, Qin; Kang, Mingqiang

2012-11-01

It has been reported that ischemic postconditioning (IPO) or mesenchymal stem cell (MSC) engraftment could protect organs from ischemia/reperfusion (I/R) injury. We investigated the synergetic effects of combined treatment on lung injury induced by I/R. Adult Sprague-Dawley rats were randomly assigned to one of the following groups: sham-operated control, I/R, IPO, MSC engraftment, and IPO plus MSC engraftment. Lung injury was assessed by arterial blood gas analysis, the wet/dry lung weight ratio, superoxide dismutase level, malondialdehyde content, myeloperoxidase activity, and tissue histologic changes. Cytokine expression was detected using real-time polymerase chain reaction, Western blotting, and enzyme-linked immunosorbent assay. Cell apoptosis was determined by terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate-biotin nick end assay and annexin V staining. MSC engraftment or IPO alone markedly attenuated the lung wet/dry weight ratio, malondialdehyde and myeloperoxidase production, and lung pathologic injury and enhanced arterial partial oxygen pressure, superoxide dismutase content, inhibited pro-inflammatory cytokine levels, and decreased cell apoptosis in lung tissue, compared with the I/R group. In contrast, IPO pretreatment enhanced the protective effects of MSC on I/R-induced lung injury compared with treatment alone. Moreover, in the combined treatment group, the number of MSC engraftments in the lung tissue was increased, associated with enhanced survival of MSCs compared with MSC treatment alone. Additional investigation showed that IPO treatment increased expression of vascular endothelial growth factor and stromal cell-derived factor-1 in I/R lung tissue. IPO might contribute to the homing and survival of transplanted MSCs and enhance their therapeutic effects through improvement of the microenvironment of I/R injury. Copyright © 2012 Elsevier Inc. All rights reserved.

Enhanced field emission properties of tilted graphene nanoribbons on aggregated TiO{sub 2} nanotube arrays

DOE Office of Scientific and Technical Information (OSTI.GOV)

Hung, Shang-Chao, E-mail: schung99@gmail.com; Chen, Yu-Jyun

2016-07-15

Highlights: • Graphene nanoribbons (GNBs) slanted on aggregate TiO{sub 2} nanotube (A-TNTs) as field-emitters. • Turn-on electric field and field enhancement factor β are dependent on the substrate morphology. • Various quantities of GNRs are deposited on top of A-TNTs (GNRs/A-TNTs) with different morphologies. • With an increase of GNBs compositions, the specimens' turn-on electric field is reduced to 2.8 V/μm. • The field enhancement factor increased rapidly to about 1964 with the addition of GNRs. - Abstract: Graphene nanoribbons (GNRs) slanted on aggregate TiO{sub 2} nanotube arrays (A-TNTs) with various compositions as field-emitters are reported. The morphology, crystalline structure,more » and composition of the as-obtained specimens were characterized by field-emission scanning electron microscopy (FE-SEM), X-ray diffraction (XRD) and Raman spectrometry. The dependence of the turn-on electric field and the field enhancement factor β on substrate morphology was studied. An increase of GNRs reduces the specimens’ turn-on electric field to 2.8 V/μm and the field enhancement factor increased rapidly to about 1964 with the addition of GNRs. Results show a strong dependence of the field emission on GNR composition aligned with the gradient on the top of the A-TNT substrate. Enhanced FE properties of the modified TNTs can be mainly attributed to their improved electrical properties and rougher surface morphology.« less

Limb-Enhancer Genie: An accessible resource of accurate enhancer predictions in the developing limb

DOE PAGES

Monti, Remo; Barozzi, Iros; Osterwalder, Marco; ...

2017-08-21

Epigenomic mapping of enhancer-associated chromatin modifications facilitates the genome-wide discovery of tissue-specific enhancers in vivo. However, reliance on single chromatin marks leads to high rates of false-positive predictions. More sophisticated, integrative methods have been described, but commonly suffer from limited accessibility to the resulting predictions and reduced biological interpretability. Here we present the Limb-Enhancer Genie (LEG), a collection of highly accurate, genome-wide predictions of enhancers in the developing limb, available through a user-friendly online interface. We predict limb enhancers using a combination of > 50 published limb-specific datasets and clusters of evolutionarily conserved transcription factor binding sites, taking advantage ofmore » the patterns observed at previously in vivo validated elements. By combining different statistical models, our approach outperforms current state-of-the-art methods and provides interpretable measures of feature importance. Our results indicate that including a previously unappreciated score that quantifies tissue-specific nuclease accessibility significantly improves prediction performance. We demonstrate the utility of our approach through in vivo validation of newly predicted elements. Moreover, we describe general features that can guide the type of datasets to include when predicting tissue-specific enhancers genome-wide, while providing an accessible resource to the general biological community and facilitating the functional interpretation of genetic studies of limb malformations.« less

Enhancement factor in low-coherence enhanced backscattering and its applications for characterizing experimental skin carcinogenesis

NASA Astrophysics Data System (ADS)

Liu, Jingjing; Xu, Zhengbin; Song, Qinghai; Konger, Raymond L.; Kim, Young L.

2010-05-01

We experimentally study potential mechanisms by which the enhancement factor in low-coherence enhanced backscattering (LEBS) can probe subtle variations in radial intensity distribution in weakly scattering media. We use enhanced backscattering of light by implementing either (1) low spatial coherence illumination or (2) multiple spatially independent detections using a microlens array under spatially coherent illumination. We show that the enhancement factor in these configurations is a measure of the integrated intensity within the localized coherence or detection area, which can exhibit strong dependence on small perturbations in scattering properties. To further evaluate the utility of the LEBS enhancement factor, we use a well-established animal model of cutaneous two-stage chemical carcinogenesis. In this pilot study, we demonstrate that the LEBS enhancement factor can be substantially altered at a stage of preneoplasia. Our animal result supports the idea that early carcinogenesis can cause subtle alterations in the scattering properties that can be captured by the LEBS enhancement factor. Thus, the LEBS enhancement factor has the potential as an easily measurable biomarker in skin carcinogenesis.

Image enhancement based on in vivo hyperspectral gastroscopic images: a case study

NASA Astrophysics Data System (ADS)

Gu, Xiaozhou; Han, Zhimin; Yao, Liqing; Zhong, Yunshi; Shi, Qiang; Fu, Ye; Liu, Changsheng; Wang, Xiguang; Xie, Tianyu

2016-10-01

Hyperspectral imaging (HSI) has been recognized as a powerful tool for noninvasive disease detection in the gastrointestinal field. However, most of the studies on HSI in this field have involved ex vivo biopsies or resected tissues. We proposed an image enhancement method based on in vivo hyperspectral gastroscopic images. First, we developed a flexible gastroscopy system capable of obtaining in vivo hyperspectral images of different types of stomach disease mucosa. Then, depending on a specific object, an appropriate band selection algorithm based on dependence of information was employed to determine a subset of spectral bands that would yield useful spatial information. Finally, these bands were assigned to be the color components of an enhanced image of the object. A gastric ulcer case study demonstrated that our method yields higher color tone contrast, which enhanced the displays of the gastric ulcer regions, and that it will be valuable in clinical applications.

Release of oxygen radicals by articular chondrocytes: a study of luminol-dependent chemiluminescence and hydrogen peroxide secretion.

PubMed

Rathakrishnan, C; Tiku, K; Raghavan, A; Tiku, M L

1992-10-01

We previously established that normal articular chondrocytes, like macrophages, express class II major histocompatibility antigens, present antigen, and induce mixed and autologous lymphocyte stimulation. In a recent study using the trapped indicator 2',7'-dichlorofluorescein diacetate, we were able to measure levels of intracellular hydrogen peroxide within normal articular chondrocytes (J Immunol 245:690-696, 1990). In the present study, we utilized the technique of chemiluminescence and the biochemical method of quantitating hydrogen peroxide release to measure the production of reactive oxygen intermediates by articular chondrocytes. Chondrocytes, in suspension or adherent to coverslips, showed luminol-dependent chemiluminescence that was dependent on the number and viability of cells. There was a dose-dependent increase in chemiluminescence in response to soluble stimuli, such as phorbol myristate acetate (PMA), concanavalin A (ConA), and f-Met-Leu-Phe (FMLP). Azide inhibited chemiluminescence, suggesting that the light emission in chondrocytes is myeloperoxidase dependent. The antioxidant, catalase, inhibited chemiluminescence but superoxide dismutase had no effect, suggesting that luminol-dependent chemiluminescence in chondrocytes mostly measured hydrogen peroxide. Chemiluminescence was also observed in fragments of live cartilage tissue, indicating that chondrocytes that are cartilage matrix bound can generate the respiratory burst response. Using the scopoletin oxidation assay, we confirmed the release of increasing amounts of hydrogen peroxide by chondrocytes exposed to interleukin-1, rabbit interferon, and tumor necrosis factor alpha. Tumor necrosis factor alpha had both priming and enhancing effects on reactive oxygen intermediate production by chondrocytes. Reactive oxygen intermediates have been shown to play a significant role in matrix degradation. We suggest that reactive oxygen intermediates produced by chondrocytes play an important role in the degradation of matrix in arthritis.

Multimodal analysis of pretreated biomass species highlights generic markers of lignocellulose recalcitrance.

PubMed

Herbaut, Mickaël; Zoghlami, Aya; Habrant, Anouck; Falourd, Xavier; Foucat, Loïc; Chabbert, Brigitte; Paës, Gabriel

2018-01-01

Biomass recalcitrance to enzymatic hydrolysis has been assigned to several structural and chemical factors. However, their relative importance remains challenging to evaluate. Three representative biomass species (wheat straw, poplar and miscanthus) were submitted to four standard pretreatments (dilute acid, hot water, ionic liquid and sodium chlorite) in order to generate a set of contrasted samples. A large array of techniques, including wet chemistry analysis, porosity measurements using NMR spectroscopy, electron and fluorescence microscopy, were used in order to determine possible generic factors of biomass recalcitrance. The pretreatment conditions selected allowed obtaining samples displaying different susceptibility to enzymatic hydrolysis (from 3 up to 98% of the initial glucose content released after 96 h of saccharification). Generic correlation coefficients were calculated between the measured chemical and structural features and the final saccharification rates. Increases in porosity displayed overall strong positive correlations with saccharification efficiency, but different porosity ranges were concerned depending on the considered biomass. Lignin-related factors displayed highly negative coefficients for all biomasses. Lignin content, which is likely involved in the correlations observed for porosity, was less detrimental to enzymatic hydrolysis than lignin composition. Lignin influence was highlighted by the strong negative correlation with fluorescence intensity which mainly originates from monolignols in mature tissues. Our results provide a better understanding of the factors responsible for biomass recalcitrance that can reasonably be considered as generic. The correlations with specific porosity ranges are biomass species-dependent, meaning that enzymes cocktails with fitted enzyme size are likely to be needed to optimise saccharification depending on the biomass origin. Lignin composition, which probably influences its structure, is the most important parameter to overcome to enhance enzymes access to the polysaccharides. Accordingly, fluorescence intensity was found to be a rapid and simple method to assess recalcitrance after pretreatment.

A low-protein diet induces body weight loss and browning of subcutaneous white adipose tissue through enhanced expression of hepatic fibroblast growth factor 21 (FGF21).

PubMed

Pérez-Martí, Albert; Garcia-Guasch, Maite; Tresserra-Rimbau, Anna; Carrilho-Do-Rosário, Alexandra; Estruch, Ramon; Salas-Salvadó, Jordi; Martínez-González, Miguel Ángel; Lamuela-Raventós, Rosa; Marrero, Pedro F; Haro, Diego; Relat, Joana

2017-08-01

Fibroblast growth factor 21 (FGF21) is considered a promising therapeutic candidate for the treatment of obesity. Since FGF21 production is regulated by various nutritional factors, we analyze the impact of low protein intake on circulating levels of this growth hormone in mice and in a sub cohort of the PREDIMED (Prevención con Dieta Mediterránea) trial. We also describe the role of hepatic FGF21 in metabolic adaptation to a low-protein diet (LPD). We fed control and liver-specific Fgf21 knockout (LFgf21KO) mice a LPD. This diet increased FGF21 production by inducing its overexpression in liver, and this correlated with a body weight decrease without changes in food intake. The LPD also caused FGF21-dependent browning in subcutaneous white adipose tissue (scWAT), as indicated by an increase in the expression of uncoupling protein 1 (UCP1). In a subgroup of 78 individuals from the PREDIMED trial, we observed an inverse correlation between protein intake and circulating FGF21 levels. Our results reinforce the involvement of FGF21 in coordinating energy homeostasis under a range of nutritional conditions. Moreover, here we describe an approach to increase the endogenous production of FGF21, which if demonstrated functional in humans, could generate a treatment for obesity. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

RESPONSE FUNCTIONS FOR COMPUTING ABSORBED DOSE TO SKELETAL TISSUES FROM PHOTON IRRADIATION – AN UPDATE

PubMed Central

Johnson, Perry; Bahadori, Amir; Eckerman, Keith; Lee, Choonsik; Bolch, Wesley E.

2014-01-01

A comprehensive set of photon fluence-to-dose response functions (DRFs) are presented for two radiosensitive skeletal tissues – active and total shallow marrow – within 15 and 32 bones sites, respectively, of the ICRP reference adult male. The functions were developed using fractional skeletal masses and associated electron absorbed fractions as reported for the UF hybrid adult male phantom, which in turn is based upon microCT images of trabecular spongiosa taken from a 40-year male cadaver. The new DRFs expand upon both the original set of seven functions produced in 1985, as well as a 2007 update calculated under the assumption of secondary electron escape from spongiosa. In the present study, it is assumed that photon irradiation of the skeleton will yield charged particle equilibrium across all spongiosa regions at energies exceeding 200 keV. Kerma factors for active marrow, inactive marrow, trabecular bone, and spongiosa at higher energies are calculated using the DRF algorithm setting the electron absorbed fraction for self-irradiation to unity. By comparing kerma factors and DRF functions, dose enhancement factors and mass energy-absorption coefficient (MEAC) ratios for active marrow to spongiosa were derived. These MEAC ratios compared well with those provided by the NIST Physical Reference Data Library (mean difference of 0.8%), and the dose enhancement factors for active marrow compared favorably with values calculated in the well-known study published by King and Spiers (1985) (mean absolute difference of 1.9 percentage points). Additionally, dose enhancement factors for active marrow were shown to correlate well with the shallow marrow volume fraction (R2 = 0.91). Dose enhancement factors for the total shallow marrow were also calculated for 32 bone sites PMID:21427484

Keratinocyte Growth Factor Administration Attenuates Murine Pulmonary Mycobacterium tuberculosis Infection through Granulocyte-Macrophage Colony-stimulating Factor (GM-CSF)-dependent Macrophage Activation and Phagolysosome Fusion*

PubMed Central

Pasula, Rajamouli; Azad, Abul K.; Gardner, Jason C.; Schlesinger, Larry S.; McCormack, Francis X.

2015-01-01

Augmentation of innate immune defenses is an appealing adjunctive strategy for treatment of pulmonary Mycobacterium tuberculosis infections, especially those caused by drug-resistant strains. The effect of intranasal administration of keratinocyte growth factor (KGF), an epithelial mitogen and differentiation factor, on M. tuberculosis infection in mice was tested in prophylaxis, treatment, and rescue scenarios. Infection of C57BL6 mice with M. tuberculosis resulted in inoculum size-dependent weight loss and mortality. A single dose of KGF given 1 day prior to infection with 105 M. tuberculosis bacilli prevented weight loss and enhanced pulmonary mycobacterial clearance (compared with saline-pretreated mice) for up to 28 days. Similar effects were seen when KGF was delivered intranasally every third day for 15 days, but weight loss and bacillary growth resumed when KGF was withdrawn. For mice with a well established M. tuberculosis infection, KGF given every 3 days beginning on day 15 postinoculation was associated with reversal of weight loss and an increase in M. tuberculosis clearance. In in vitro co-culture experiments, M. tuberculosis-infected macrophages exposed to conditioned medium from KGF-treated alveolar type II cell (MLE-15) monolayers exhibited enhanced GM-CSF-dependent killing through mechanisms that included promotion of phagolysosome fusion and induction of nitric oxide. Alveolar macrophages from KGF-treated mice also exhibited enhanced GM-CSF-dependent phagolysosomal fusion. These results provide evidence that administration of KGF promotes M. tuberculosis clearance through GM-CSF-dependent mechanisms and enhances host defense against M. tuberculosis infection. PMID:25605711

Enhancement of High-Density Lipoprotein Cholesterol Functions by Encapsulation of Policosanol Exerts Anti-Senescence and Tissue Regeneration Effects Via Improvement of Anti-Glycation, Anti-Apoptosis, and Cholesteryl Ester Transfer Inhibition.

PubMed

Lim, So-Mang; Yoo, Jeong-Ah; Lee, Eun-Young; Cho, Kyung-Hyun

2016-02-01

Consumption of policosanol (PCO), a refined mixture of sugar cane wax alcohols, can elevate serum levels of high-density lipoprotein cholesterol (HDL-C), although the molecular mechanism is still unknown. To investigate the mechanism of action responsible for the anti-senescence effects of PCO on lipoprotein metabolism and HDL functionality, we synthesized reconstituted HDL (rHDL) containing PCO. Encapsulation of PCO by rHDL (PCO-rHDL) enhanced anti-oxidant activity against cupric ion-mediated low-density lipoprotein (LDL) oxidation. PCO-rHDL (final concentration, 9 μM PCO) showed more potent anti-oxidant activity than vitamin C treatment (final concentration, 100 μM). PCO-rHDL inhibited fructose-mediated glycation, which is a major pathological mechanism of diabetic complications, in a dose-dependent manner. PCO also showed cytoprotective effects in monocytes and macrophages with less triggering of apoptotic processes and reactive oxygen species (ROS) production in the presence of hydrogen peroxide (H2O2). PCO-rHDL strongly inhibited uptake of acetylated LDL into macrophages, which is an initial atherosclerotic process. Surprisingly, PCO-rHDL inhibited human serum cholesteryl ester transfer protein (CETP) activity by up to 47% (final concentration, 10 μM PCO). Subcutaneous injection of PCO-rHDL dose-dependently enhanced tissue regeneration activity by 2.4-fold and 3.6-fold compared to that of the phosphate-buffered saline (PBS) control. In conclusion, PCO in HDL showed potent anti-oxidant, anti-glycation, and CETP inhibitory activities along with tissue regenerative activity, especially upon incorporation into HDL. These results suggest that PCO can enhance functionality of HDL in serum to exert anti-senescence and longevity effects.

RNA helicase MOV10 functions as a co-factor of HIV-1 Rev to facilitate Rev/RRE-dependent nuclear export of viral mRNAs

DOE Office of Scientific and Technical Information (OSTI.GOV)

Huang, Feng; Zhang, Junsong; Zhang, Yijun

Human immunodeficiency virus type 1 (HIV-1) exploits multiple host factors during its replication. The REV/RRE-dependent nuclear export of unspliced/partially spliced viral transcripts needs the assistance of host proteins. Recent studies have shown that MOV10 overexpression inhibited HIV-1 replication at various steps. However, the endogenous MOV10 was required in certain step(s) of HIV-1 replication. In this report, we found that MOV10 potently enhances the nuclear export of viral mRNAs and subsequently increases the expression of Gag protein and other late products through affecting the Rev/RRE axis. The co-immunoprecipitation analysis indicated that MOV10 interacts with Rev in an RNA-independent manner. The DEAG-boxmore » of MOV10 was required for the enhancement of Rev/RRE-dependent nuclear export and the DEAG-box mutant showed a dominant-negative activity. Our data propose that HIV-1 utilizes the anti-viral factor MOV10 to function as a co-factor of Rev and demonstrate the complicated effects of MOV10 on HIV-1 life cycle. - Highlights: • MOV10 can function as a co-factor of HIV-1 Rev. • MOV10 facilitates Rev/RRE-dependent transport of viral mRNAs. • MOV10 interacts with Rev in an RNA-independent manner. • The DEAG-box of MOV10 is required for the enhancement of Rev/RRE-dependent export.« less

Genome-wide DNA methylation measurements in prostate tissues uncovers novel prostate cancer diagnostic biomarkers and transcription factor binding patterns.

PubMed

Kirby, Marie K; Ramaker, Ryne C; Roberts, Brian S; Lasseigne, Brittany N; Gunther, David S; Burwell, Todd C; Davis, Nicholas S; Gulzar, Zulfiqar G; Absher, Devin M; Cooper, Sara J; Brooks, James D; Myers, Richard M

2017-04-17

Current diagnostic tools for prostate cancer lack specificity and sensitivity for detecting very early lesions. DNA methylation is a stable genomic modification that is detectable in peripheral patient fluids such as urine and blood plasma that could serve as a non-invasive diagnostic biomarker for prostate cancer. We measured genome-wide DNA methylation patterns in 73 clinically annotated fresh-frozen prostate cancers and 63 benign-adjacent prostate tissues using the Illumina Infinium HumanMethylation450 BeadChip array. We overlaid the most significantly differentially methylated sites in the genome with transcription factor binding sites measured by the Encyclopedia of DNA Elements consortium. We used logistic regression and receiver operating characteristic curves to assess the performance of candidate diagnostic models. We identified methylation patterns that have a high predictive power for distinguishing malignant prostate tissue from benign-adjacent prostate tissue, and these methylation signatures were validated using data from The Cancer Genome Atlas Project. Furthermore, by overlaying ENCODE transcription factor binding data, we observed an enrichment of enhancer of zeste homolog 2 binding in gene regulatory regions with higher DNA methylation in malignant prostate tissues. DNA methylation patterns are greatly altered in prostate cancer tissue in comparison to benign-adjacent tissue. We have discovered patterns of DNA methylation marks that can distinguish prostate cancers with high specificity and sensitivity in multiple patient tissue cohorts, and we have identified transcription factors binding in these differentially methylated regions that may play important roles in prostate cancer development.

Self-production of tissue factor-coagulation factor VII complex by ovarian cancer cells.

PubMed

Yokota, N; Koizume, S; Miyagi, E; Hirahara, F; Nakamura, Y; Kikuchi, K; Ruf, W; Sakuma, Y; Tsuchiya, E; Miyagi, Y

2009-12-15

Thromboembolic events are a major complication in ovarian cancer patients. Tissue factor (TF) is frequently overexpressed in ovarian cancer tissue and correlates with intravascular thrombosis. TF binds to coagulation factor VII (fVII), changing it to its active form, fVIIa. This leads to activation of the extrinsic coagulation cascade. fVII is produced by the liver and believed to be supplied from blood plasma at the site of coagulation. However, we recently showed that ovarian cancer cells express fVII transcripts under normoxia and that this transcription is inducible under hypoxia. These findings led us to hypothesise that ovarian cancer cells are intrinsically associated with TF-fVIIa coagulation activity, which could result in thrombosis. In this study, we examined whether ectopically expressed fVII could cause thrombosis by means of immunohistochemistry, RT-PCR, western blotting and flow cytometry. Ectopic fVII expression occurs frequently in ovarian cancers, particularly in clear cell carcinoma. We further showed that ovarian cancer cells express TF-fVIIa on the cell surface under normoxia and that this procoagulant activity is enhanced by hypoxic stimuli. Moreover, we showed that ovarian cancer cells secrete microparticles (MPs) with TF-fVIIa activity. Production of this procoagulant secretion is enhanced under hypoxia. These results raise the possibility that cancer cell-derived TF-fVIIa could cause thrombotic events in ovarian cancer patients.

Application of platelet-rich plasma and platelet-rich fibrin in fat grafting: basic science and literature review.

PubMed

Liao, Han-Tsung; Marra, Kacey G; Rubin, J Peter

2014-08-01

Due to the natural properties of fat, fat grafting remains a popular procedure for soft tissue volume augmentation and reconstruction. However, clinical outcome varies and is technique dependent. Platelet-rich plasma (PRP) contains α-granules, from which multiple growth factors such as platelet-derived growth factor, transforming growth factor-β, vascular endothelial growth factor, and epidermal growth factor can be released after activation. In recent years, the scope of PRP therapies has extended from bone regeneration, wound healing, and healing of musculoskeletal injuries, to enhancement of fat graft survival. In this review, we focus on the definition of PRP, the different PRP preparation and activation methods, and growth factor concentrations. In addition, we discuss possible mechanisms for the role of PRP in fat grafting by reviewing in vitro studies with adipose-derived stem cells, preadipocytes, and adipocytes, and preclinical and clinical research. We also review platelet-rich fibrin, a so-called second generation PRP, and its slow-releasing biology and effects on fat grafts compared to PRP in both animal and clinical research. Finally, we provide a general foundation on which to critically evaluate earlier studies, discuss the limitations of previous research, and direct plans for future experiments to improve the optimal effects of PRP in fat grafting.

Fifty Years of Research in ARDS. Cell-based Therapy for Acute Respiratory Distress Syndrome. Biology and Potential Therapeutic Value.

PubMed

Laffey, John G; Matthay, Michael A

2017-08-01

On the basis of several preclinical studies, cell-based therapy has emerged as a potential new therapeutic for acute respiratory distress syndrome (ARDS). Of the various cell-based therapy options, mesenchymal stem/stromal cells (MSCs) from bone marrow, adipose tissue, and umbilical cord have the most experimental data to support their potential efficacy for lung injury from both infectious and noninfectious causes. Mechanistically, MSCs exert their beneficial effects by release of paracrine factors, microvesicles, and transfer of mitochondria, all of which have antiinflammatory and pro-resolving effects on injured lung endothelium and alveolar epithelium, including enhancing the resolution of pulmonary edema by up-regulating sodium-dependent alveolar fluid clearance. MSCs also have antimicrobial effects mediated by release of antimicrobial factors and by up-regulating monocyte/macrophage phagocytosis. Phase 2a clinical trials to establish safety in ARDS are in progress, and two phase 1 trials did not report any serious adverse events. Several issues need further study, including: determining the optimal methods for large-scale production, reconstitution of cryopreserved cells for clinical use, defining cell potency assays, and determining the therapeutic potential of conditioned media derived from MSCs. Because ARDS is a heterogeneous syndrome, targeting MSCs to patients with ARDS with a more hyperinflammatory endotype may further enhance their potential for efficacy.

Effect of hypoxia on tissue factor pathway inhibitor expression in breast cancer.

PubMed

Cui, X Y; Tinholt, M; Stavik, B; Dahm, A E A; Kanse, S; Jin, Y; Seidl, S; Sahlberg, K K; Iversen, N; Skretting, G; Sandset, P M

2016-02-01

ESSENTIALS: A hypoxic microenvironment is a common feature of tumors that may influence activation of coagulation. MCF-7 and SK-BR-3 breast cancer cells and breast cancer tissue samples were used. The results showed transcriptional repression of tissue factor pathway inhibitor expression in hypoxia. Hypoxia-inducible factor 1α may be a target for the therapy of cancer-related coagulation and thrombosis. Activation of coagulation is a common finding in patients with cancer, and is associated with an increased risk of venous thrombosis. As a hypoxic microenvironment is a common feature of solid tumors, we investigated the role of hypoxia in the regulation of tissue factor (TF) pathway inhibitor (TFPI) expression in breast cancer. To explore the transcriptional regulation of TFPI by hypoxia-inducible factor (HIF)-1α in breast cancer cells and their correlation in breast cancer tissues. MCF-7 and SK-BR-3 breast cancer cells were cultured in 1% oxygen or treated with cobalt chloride (CoCl2 ) to mimic hypoxia. Time-dependent and dose-dependent downregulation of TFPI mRNA (quantitative RT-PCR) and of free TFPI protein (ELISA) were observed in hypoxia. Western blotting showed parallel increases in the levels of HIF-1α protein and TF. HIF-1α inhibitor abolished or attenuated the hypoxia-induced downregulation of TFPI. Luciferase reporter assay showed that both hypoxia and HIF-1α overexpression caused strong repression of TFPI promoter activity. Subsequent chromatin immunoprecipitation and mutagenesis analysis demonstrated a functional hypoxia response element within the TFPI promoter, located at -1065 to -1060 relative to the transcriptional start point. In breast cancer tissue samples, gene expression analyses showed a positive correlation between the mRNA expression of TFPI and that of HIF-1α. This study demonstrates that HIF-1α is involved in the transcriptional regulation of the TFPI gene, and suggests that a hypoxic microenvironment inside a breast tumor may induce a procoagulant state in breast cancer patients. © 2015 International Society on Thrombosis and Haemostasis.

Development of a platform for co-registered ultrasound and MR contrast imaging in vivo

NASA Astrophysics Data System (ADS)

Chandrana, Chaitanya; Bevan, Peter; Hudson, John; Pang, Ian; Burns, Peter; Plewes, Donald; Chopra, Rajiv

2011-02-01

Imaging of the microvasculature is often performed using contrast agents in combination with either ultrasound (US) or magnetic resonance (MR) imaging. Contrast agents are used to enhance medical imaging by highlighting microvascular properties and function. Dynamic signal changes arising from the passage of contrast agents through the microvasculature can be used to characterize different pathologies; however, comparisons across modalities are difficult due to differences in the interactions of contrast agents with the microvasculature. Better knowledge of the relationship of contrast enhancement patterns with both modalities could enable better characterization of tissue microvasculature. We developed a co-registration platform for multi-modal US and MR imaging using clinical imaging systems in order to study the relationship between US and MR contrast enhancement. A preliminary validation study was performed in phantoms to determine the registration accuracy of the platform. In phantoms, the in-plane registration accuracy was measured to be 0.2 ± 0.2 and 0.3 ± 0.2 mm, in the lateral and axial directions, respectively. The out-of-plane registration accuracy was estimated to be 0.5 mm ±0.1. Co-registered US and MR imaging was performed in a rabbit model to evaluate contrast kinetics in different tissue types after bolus injections of US and MR contrast agents. The arrival time of the contrast agent in the plane of imaging was relatively similar for both modalities. We studied three different tissue types: muscle, large vessels and fat. In US, the temporal kinetics of signal enhancement were not strongly dependent on tissue type. In MR, however, due to the different amounts of agent extravasation in each tissue type, tissue-specific contrast kinetics were observed. This study demonstrates the feasibility of performing in vivo co-registered contrast US and MR imaging to study the relationships of the enhancement patterns with each modality.

Experimental investigations on intracavity sonography. Part 2: Alteration of imaging by artificial alterations in the wall of isolated porcine urinary bladders.

PubMed

Jaeger, N; Vahlensieck, W

1986-01-01

Because the determination of the depth of urinary bladder tumors by means of intracavity sonography depends on several factors (tumor size, reflection behavior of the tumor etc.), we checked the imaging of this diagnostic technique in the isolated porcine urinary bladder under various experimental conditions. Different tissues of defined size were fixed on the inner or outer surface of the bladder wall; both the bladder mucosa and the foreign tissue were damaged thermally or by incision. The importance of a limited depth of sound penetration or of a sound shadow depending on the characteristics of the tissue under investigation was revealed; tissue types could not be distinguished unequivocally by the reflection pattern; above all, a sonographic diagnosis of the tumor was not possible in the presence of histo-pathologically detectable tissue changes due to thermal damage.

In Vivo Determination of the Complex Elastic Moduli of Cetacean Head Tissue

DTIC Science & Technology

2013-09-30

of an ultrasonic Doppler vibration measurement system called NVMS developed at Georgia Tech iii. Algorithms have been developed to enable the...magnitude and phase of vibration to be determined as a function of range (tissue depth) along the ultrasonic beam. By measuring the differential phase of...The frequency dependence of the propagation speed is then used to determine the shear loss factor. The elastic properties of tissue phantoms

Combined effects of chemical priming and mechanical stimulation on mesenchymal stem cell differentiation on nanofiber scaffolds

PubMed Central

Subramony, Siddarth D.; Su, Amanda; Yeager, Keith; Lu, Helen H.

2014-01-01

Functional tissue engineering of connective tissues such as the anterior cruciate ligament (ACL) remains a significant clinical challenge, largely due to the need for mechanically competent scaffold systems for grafting, as well as a reliable cell source for tissue formation. We have designed an aligned, polylactide-co-glycolide (PLGA) nanofiber-based scaffold with physiologically relevant mechanical properties for ligament regeneration. The objective of this study is to identify optimal tissue engineering strategies for fibroblastic induction of human mesenchymal stem cells (hMSC), testing the hypothesis that basic fibroblast growth factor (bFGF) priming coupled with tensile loading will enhance hMSC-mediated ligament regeneration. It was observed that compared to the unloaded, as well as growth factor-primed but unloaded controls, bFGF stimulation followed by physiologically relevant tensile loading enhanced hMSC proliferation, collagen production and subsequent differentiation into ligament fibroblast-like cells, upregulating the expression of types I and III collagen, as well as tenasin-C and tenomodulin. The results of this study suggest that bFGF priming increases cell proliferation, while mechanical stimulation of the hMSCs on the aligned nanofiber scaffold promotes fibroblastic induction of these cells. In addition to demonstrating the potential of nanofiber scaffolds for hMSC-mediated functional ligament tissue engineering, this study yields new insights into the interactive effects of chemical and mechanical stimuli on stem cell differentiation. PMID:24267271

Endothelial Progenitor Cells=EPC=Elemental Pernicious Complexity

PubMed Central

Ushio-Fukai, Masuko

2011-01-01

Abstract Endothelial progenitor cells (EPCs) represent a heterogeneous population of cells with a pro-angiogenic potential that are derived not only from bone marrow but also from other tissues. Depending on the model and cell type used, the pro-angiogenic effect is a consequence of direct vascular integration, the paracrine release of growth factors and cytokines, or complex interactions with other cellular components like monocytes or platelets. The pro-angiogenic potential of EPCs is dependent on the particular type of EPC studied and modulated by the risk and life style factors of the patient as well as by local factors determining the homing to diseased tissue and the EPC proteome. In this Forum on EPCs these aspects will be covered in individual review articles, which are accompanied by two original research studies on the role of NADPH oxidases for EPC mobilization and the impact of organic nitrates on EPCs. Antioxid. Redox Signal. 15, 911–914. PMID:21128729

Hypersalinity Acclimation Increases the Toxicity of the Insecticide Phorate in Coho Salmon (Oncorhynchus kisutch)

PubMed Central

Lavado, Ramon; Maryoung, Lindley A.; Schlenk, Daniel

2012-01-01

Previous studies in euryhaline fish have shown that acclimation to hypersaline environments enhances the toxicity of thioether organophosphate and carbamate pesticides. To better understand the potential mechanism of enhanced toxicity, the effects of the organophosphate insecticide phorate were evaluated in coho salmon (Oncorhynchus kisutch) maintained in freshwater (<0.5 g/L salinity) and 32 g/L salinity. The observed 96-h LC50 in freshwater fish (67.34 ± 3.41 μg/L) was significantly reduced to 2.07 ± 0.16 μg/L in hypersaline-acclimated fish. Because organophosphates often require bioactivation to elicit toxicity through acetylcholinesterase (AChE) inhibition, the in vitro biotransformation of phorate was evaluated in coho salmon maintained in different salinities in liver, gills, and olfactory tissues. Phorate sulfoxide was the predominant metabolite in each tissue but rates of formation diminished in a salinity-dependent manner. In contrast, formation of phorate-oxon (gill; olfactory tissues), phorate sulfone (liver), and phorate-oxon sulfoxide (liver; olfactory tissues) was significantly enhanced in fish acclimated to higher salinities. From previous studies, it was expected that phorate and phorate sulfoxide would be less potent AChE inhibitors than phorate-oxon, with phorate-oxon sulfoxide being the most potent of the compounds tested. This trend was confirmed in this study. In summary, these results suggest that differential expression and/or catalytic activities of Phase I enzymes may be involved to enhance phorate oxidative metabolism and subsequent toxicity of phorate to coho salmon under hypersaline conditions. The outcome may be enhanced fish susceptibility to anticholineterase oxon sulfoxides. PMID:21488666

Micrometer scale guidance of mesenchymal stem cells to form structurally oriented large-scale tissue engineered cartilage.

PubMed

Chou, Chih-Ling; Rivera, Alexander L; Williams, Valencia; Welter, Jean F; Mansour, Joseph M; Drazba, Judith A; Sakai, Takao; Baskaran, Harihara

2017-09-15

Current clinical methods to treat articular cartilage lesions provide temporary relief of the symptoms but fail to permanently restore the damaged tissue. Tissue engineering, using mesenchymal stem cells (MSCs) combined with scaffolds and bioactive factors, is viewed as a promising method for repairing cartilage injuries. However, current tissue engineered constructs display inferior mechanical properties compared to native articular cartilage, which could be attributed to the lack of structural organization of the extracellular matrix (ECM) of these engineered constructs in comparison to the highly oriented structure of articular cartilage ECM. We previously showed that we can guide MSCs undergoing chondrogenesis to align using microscale guidance channels on the surface of a two-dimensional (2-D) collagen scaffold, which resulted in the deposition of aligned ECM within the channels and enhanced mechanical properties of the constructs. In this study, we developed a technique to roll 2-D collagen scaffolds containing MSCs within guidance channels in order to produce a large-scale, three-dimensional (3-D) tissue engineered cartilage constructs with enhanced mechanical properties compared to current constructs. After rolling the MSC-scaffold constructs into a 3-D cylindrical structure, the constructs were cultured for 21days under chondrogenic culture conditions. The microstructure architecture and mechanical properties of the constructs were evaluated using imaging and compressive testing. Histology and immunohistochemistry of the constructs showed extensive glycosaminoglycan (GAG) and collagen type II deposition. Second harmonic generation imaging and Picrosirius red staining indicated alignment of neo-collagen fibers within the guidance channels of the constructs. Mechanical testing indicated that constructs containing the guidance channels displayed enhanced compressive properties compared to control constructs without these channels. In conclusion, using a novel roll-up method, we have developed large scale MSC based tissue-engineered cartilage that shows microscale structural organization and enhanced compressive properties compared to current tissue engineered constructs. Tissue engineered cartilage constructs made with human mesenchymal stem cells (hMSCs), scaffolds and bioactive factors are a promising solution to treat cartilage defects. A major disadvantage of these constructs is their inferior mechanical properties compared to the native tissue, which is likely due to the lack of structural organization of the extracellular matrix of the engineered constructs. In this study, we developed three-dimensional (3-D) cartilage constructs from rectangular scaffold sheets containing hMSCs in micro-guidance channels and characterized their mechanical properties and metabolic requirements. The work led to a novel roll-up method to embed 2-D microscale structures in 3-D constructs. Further, micro-guidance channels incorporated within the 3-D cartilage constructs led to the production of aligned cell-produced matrix and enhanced mechanical function. Copyright © 2017 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

Characterization of (/sup 3/H)forskolin binding sites in the iris-ciliary body of the albino rabbit

DOE Office of Scientific and Technical Information (OSTI.GOV)

Goldman, M.E.; Mallorga, P.; Pettibone, D.J.

1988-01-01

(/sup 3/H)forskolin binding sites were identified using membranes prepared from the iris-ciliary body of adult, albino rabbits. Scatchard analysis of saturation binding experiments demonstrated that (/sup 3/H)forskolin bound to a single population of high affinity sites. The K/sub d/ and B/sub max/ values were 8.7 +- 0.9 nM and 119.0 +- 30.9 fmolmg prot. using membranes prepared from frozen tissue and 17.0 +- 6.2 nM and 184.4 +- 47.2 fmolmg prot. using fresh tissue. The binding of (/sup 3/H)forskolin was magnesium-dependent. The B/sub max/ was enhanced by sodium fluoride and Gpp(NH)p, a nonhydrolyzable guanine nucleotide analog. Forskolin was the mostmore » potent inhibitor of (/sup 3/H)forskolin binding; two commercially-available analogs were weaker inhibitors. In an adenylate cyclase assay, there was the same rank order of potency to enhance enzyme activity. Based upon binding affinities, magnesium-dependence, sensitivity to sodium fluoride and Gpp(NH)p, rank order of potencies of analogs and correlation of binding with adenylate cyclase activity, these studies suggest that the (/sup 3/H)forskolin binding site in the iris-ciliary body is similar to the binding site in other tissues« less

TH-E-BRD-01: Innovation in (gold) Nanoparticle-Enhanced Therapy

DOE Office of Scientific and Technical Information (OSTI.GOV)

Krishnan, S; Chithrani, B; Berbeco, R

2014-06-15

Radiation therapy relies on the concept of delivering high dose to tumor volumes whilst simultaneously aiming to minimize irradiation of healthy tissue. Gold and other metallic nanoparticles (GNPs) have the potential to greatly enhance dose depositions in their close proximity. While it was originally thought that this effect would only be significant for kV photon beams, it has been shown that GNPs also enhance dose and increase cell killing and survival fraction for MV photons as well as protons. GNPs have been shown to be preferentially taken up in tumors, depending on the GNP properties either internalized in the tumormore » cells or clustering in the tumor vasculature. Therefore GNPs offer an intriguing additional option to target the tumor while sparing healthy tissue. While a growing amount of research shows GNP induced enhancement factors in the order of 1.5 and higher, GNPs have not yet entered into clinical routine. In this symposium we will have three presentations discussing the current status of GNP based research, the potential to include GNPs in radiation therapy and the limitations and problems to use GNPs in the clinic. Physical and biological underpinnings of radiosensitization with gold nano particles An evolving body of recent literature alludes to the potential to sensitize tumors to radiation therapy using metallic nanoparticles. In preclinical studies, the techniques that hold promise for eventual clinical deployment are nanoparticle-assisted radiation dose enhancement and hyperthermic radiosensitization. To understand the underlying nanoparticle-radiation interactions, computational techniques offer an explanation for and predict the biophysical consequences at a nano-/meso-scopic scale. Nonetheless, there are persisting gaps in knowledge relating to the molecular mechanism of action of these radiosensitization approaches — some of these issues will be addressed. Since the literature relating to the diverse disciplines involved in these efforts spans across multiple specialties (clinical radiation oncology, radiation physics, radiation biology, nanotechnology, material science, biomedical engineering, pharmacology, chemistry, and tumor biology) and numerous specialty journals, there is no single compilation of extant research in this arena or forum for merging analogous concepts and paradigms. This symposium will provide such a venue — my presentation will start with familiarizing the audience with the potential applications of metallic nanoparticles in radiation therapy using specific illustrative examples and begin to explore ways to understand the underlying mechanisms of the effects observed. Biological effects of Gold nanoparticles in radiation therapy Gold nanoparticles (GNP) have been investigated as platforms to carry drugs or radio-sensitizing agents to tumors due to the biocompatibility of gold and relative ease of conjugation with therapeutic and targeting moieties. Recently, there has been interest in exploiting the physical properties of gold, specifically the high atomic number, to enhance radiation therapy. When irradiated, gold atoms will produce low energy electrons, depositing energy within a short distance. The ratio of dose deposited in the presence of the GNP to the dose deposited in the absence of GNP is referred to as the dose enhancement factor (DEF). This factor has been shown to depend on the concentration of GNP and the energy of the incident photons. The physics of this process, preliminary in vitro and in vivo experiments and future directions for this nascent field are described in this presentation. Gold Nanoparticles for improved therapeutic outcome in radiation therapy The application of nanoparticles (NPs) for improved therapeutics is at the forefront of cancer nanotechnology. Among other NP systems, gold nanoparticles (GNPs) are extensively used due to its impressive ability to act as both an anticancer drug carrier in chemotherapy and as a dose enhancer in radiotherapy. Cellular uptake of GNPs was dependent on their size. Among GNPs of diameter between 14–74 nm, GNPs of size 50 nm has the highest uptake. Radiosensitization was dependent on the size of the GNPs as well. GNPs of size 50-nm showed the highest radiosensitization enhancement factor compared to GNPs of 14 and 74 nm for lower- (105 kVp) and higher- (6 MVp) energy photons. GNPs used in those studies were predominantly localized in the cell cytoplasm. However, the therapeutic response can be further enhanced if NPs can be effectively targeted into the nucleus. Here, we present an effective strategy for designing a GNP-peptide complex for nuclear targeting. Two peptides were conjugated onto a GNP: One peptide enhanced the uptake while the other peptide enhanced the nuclear delivery. The nuclear-targeted cells displayed a fourfold increase in the therapeutic response when treated with radiation as compared to untargeted ones. DNA double-strand breaks were quantified using radiation-induced foci of γ- H2AX and 53BP1, and a modest increase in the number of foci per nucleus was observed in irradiated cell populations with internalized GNPs. This research will establish a more aggressive NP-based treatment approach for improved outcome in cancer therapy. Learning Objectives: Introduce radiosensitization concepts of metallic nanoparticle and provide the theoretical basis Provide an overview over the size and coating dependence for GNP uptake in cells Provide a compilation of the extant, multi-discipline research on metallic nanoparticles Understand the prospects for future studies and innovations and the potential for applications of metallic nanoparticles in radiation therapy.« less

Electric field enhancement due to a saw-tooth asperity in a channel and implications on microscale gas breakdown

NASA Astrophysics Data System (ADS)

Venkattraman, Ayyaswamy

2014-10-01

The electric field enhancement due to an isolated saw-tooth asperity in an infinite channel is considered with the goal of providing some inputs to the choice of field enhancement factors used to describe microscale gas breakdown. The Schwarz-Christoffel transformation is used to map the interior of the channel to the upper half of the transformed plane. The expression for the electric field in the transformed plane is then used to determine the electric field distribution in the channel as well as field enhancement near the asperity. The effective field enhancement factor is determined and its dependence on operating and geometrical parameters is studied. While the effective field enhancement factor depends only weakly on the height of the asperity in comparison to the channel, it is influenced significantly by the base angles of the asperity. Due to the strong dependence of field emission current density on electric field, the effective field enhancement factor (βeff) is shown to vary rapidly with the applied electric field irrespective of the geometrical parameters. This variation is included in the analysis of microscale gas breakdown and compared with results obtained using a constant βeff as is done traditionally. Even though results for a varying βeff may be approximately reproduced using an equivalent constant βeff independent of E-field, it might be important for a range of operating conditions. This is confirmed by extracting βeff from experimental data for breakdown in argon microgaps with plane-parallel cathodes and comparing its dependence on the E-field. While the use of two-dimensional asperities is shown to be a minor disadvantage of the proposed approach in its current form, it can potentially help in developing predictive capabilities as opposed to treating βeff as a curve-fitting parameter.

Loss of pericyte smoothened activity in mice with genetic deficiency of leptin.

PubMed

Xie, Guanhua; Swiderska-Syn, Marzena; Jewell, Mark L; Machado, Mariana Verdelho; Michelotti, Gregory A; Premont, Richard T; Diehl, Anna Mae

2017-04-20

Obesity is associated with multiple diseases, but it is unclear how obesity promotes progressive tissue damage. Recovery from injury requires repair, an energy-expensive process that is coupled to energy availability at the cellular level. The satiety factor, leptin, is a key component of the sensor that matches cellular energy utilization to available energy supplies. Leptin deficiency signals energy depletion, whereas activating the Hedgehog pathway drives energy-consuming activities. Tissue repair is impaired in mice that are obese due to genetic leptin deficiency. Tissue repair is also blocked and obesity enhanced by inhibiting Hedgehog activity. We evaluated the hypothesis that loss of leptin silences Hedgehog signaling in pericytes, multipotent leptin-target cells that regulate a variety of responses that are often defective in obesity, including tissue repair and adipocyte differentiation. We found that pericytes from liver and white adipose tissue require leptin to maintain expression of the Hedgehog co-receptor, Smoothened, which controls the activities of Hedgehog-regulated Gli transcription factors that orchestrate gene expression programs that dictate pericyte fate. Smoothened suppression prevents liver pericytes from being reprogrammed into myofibroblasts, but stimulates adipose-derived pericytes to become white adipocytes. Progressive Hedgehog pathway decay promotes senescence in leptin-deficient liver pericytes, which, in turn, generate paracrine signals that cause neighboring hepatocytes to become fatty and less proliferative, enhancing vulnerability to liver damage. Leptin-responsive pericytes evaluate energy availability to inform tissue construction by modulating Hedgehog pathway activity and thus, are at the root of progressive obesity-related tissue pathology. Leptin deficiency inhibits Hedgehog signaling in pericytes to trigger a pericytopathy that promotes both adiposity and obesity-related tissue damage.

Stem Cell-based Tissue Engineering Approaches for Musculoskeletal Regeneration

PubMed Central

Brown, Patrick T.; Handorf, Andrew M.; Jeon, Won Bae; Li, Wan-Ju

2014-01-01

The field of regenerative medicine and tissue engineering is an ever evolving field that holds promise in treating numerous musculoskeletal diseases and injuries. An important impetus in the development of the field was the discovery and implementation of stem cells. The utilization of mesenchymal stem cells, and later embryonic and induced pluripotent stem cells, opens new arenas for tissue engineering and presents the potential of developing stem cell-based therapies for disease treatment. Multipotent and pluripotent stem cells can produce various lineage tissues, and allow for derivation of a tissue that may be comprised of multiple cell types. As the field grows, the combination of biomaterial scaffolds and bioreactors provides methods to create an environment for stem cells that better represent their microenvironment for new tissue formation. As technologies for the fabrication of biomaterial scaffolds advance, the ability of scaffolds to modulate stem cell behavior advances as well. The composition of scaffolds could be of natural or synthetic materials and could be tailored to enhance cell self-renewal and/or direct cell fates. In addition to biomaterial scaffolds, studies of tissue development and cellular microenvironments have determined other factors, such as growth factors and oxygen tension, that are crucial to the regulation of stem cell activity. The overarching goal of stem cell-based tissue engineering research is to precisely control differentiation of stem cells in culture. In this article, we review current developments in tissue engineering, focusing on several stem cell sources, induction factors including growth factors, oxygen tension, biomaterials, and mechanical stimulation, and the internal and external regulatory mechanisms that govern proliferation and differentiation. PMID:23432679

Intracrine Androgens Enhance Decidualization and Modulate Expression of Human Endometrial Receptivity Genes

PubMed Central

Gibson, Douglas A.; Simitsidellis, Ioannis; Cousins, Fiona L.; Critchley, Hilary O. D.; Saunders, Philippa T. K.

2016-01-01

The endometrium is a complex, steroid-dependent tissue that undergoes dynamic cyclical remodelling. Transformation of stromal fibroblasts (ESC) into specialised secretory cells (decidualization) is fundamental to the establishment of a receptive endometrial microenvironment which can support and maintain pregnancy. Androgen receptors (AR) are present in ESC; in other tissues local metabolism of ovarian and adrenal-derived androgens regulate AR-dependent gene expression. We hypothesised that altered expression/activity of androgen biosynthetic enzymes would regulate tissue availability of bioactive androgens and the process of decidualization. Primary human ESC were treated in vitro for 1–8 days with progesterone and cAMP (decidualized) in the presence or absence of the AR antagonist flutamide. Time and treatment-dependent changes in genes essential for a) intra-tissue biosynthesis of androgens (5α-reductase/SRD5A1, aldo-keto reductase family 1 member C3/AKR1C3), b) establishment of endometrial decidualization (IGFBP1, prolactin) and c) endometrial receptivity (SPP1, MAOA, EDNRB) were measured. Decidualization of ESC resulted in significant time-dependent changes in expression of AKR1C3 and SRD5A1 and secretion of T/DHT. Addition of flutamide significantly reduced secretion of IGFBP1 and prolactin and altered the expression of endometrial receptivity markers. Intracrine biosynthesis of endometrial androgens during decidualization may play a key role in endometrial receptivity and offer a novel target for fertility treatment. PMID:26817618

Intracrine Androgens Enhance Decidualization and Modulate Expression of Human Endometrial Receptivity Genes.

PubMed

Gibson, Douglas A; Simitsidellis, Ioannis; Cousins, Fiona L; Critchley, Hilary O D; Saunders, Philippa T K

2016-01-28

The endometrium is a complex, steroid-dependent tissue that undergoes dynamic cyclical remodelling. Transformation of stromal fibroblasts (ESC) into specialised secretory cells (decidualization) is fundamental to the establishment of a receptive endometrial microenvironment which can support and maintain pregnancy. Androgen receptors (AR) are present in ESC; in other tissues local metabolism of ovarian and adrenal-derived androgens regulate AR-dependent gene expression. We hypothesised that altered expression/activity of androgen biosynthetic enzymes would regulate tissue availability of bioactive androgens and the process of decidualization. Primary human ESC were treated in vitro for 1-8 days with progesterone and cAMP (decidualized) in the presence or absence of the AR antagonist flutamide. Time and treatment-dependent changes in genes essential for a) intra-tissue biosynthesis of androgens (5α-reductase/SRD5A1, aldo-keto reductase family 1 member C3/AKR1C3), b) establishment of endometrial decidualization (IGFBP1, prolactin) and c) endometrial receptivity (SPP1, MAOA, EDNRB) were measured. Decidualization of ESC resulted in significant time-dependent changes in expression of AKR1C3 and SRD5A1 and secretion of T/DHT. Addition of flutamide significantly reduced secretion of IGFBP1 and prolactin and altered the expression of endometrial receptivity markers. Intracrine biosynthesis of endometrial androgens during decidualization may play a key role in endometrial receptivity and offer a novel target for fertility treatment.

Transforming growth factor-beta stimulates wound healing and modulates extracellular matrix gene expression in pig skin. I. Excisional wound model.

PubMed

Quaglino, D; Nanney, L B; Kennedy, R; Davidson, J M

1990-09-01

The effect of transforming growth factor-beta 1 (TGF-beta 1) on matrix gene expression has been investigated during the process of wound repair, where the formation of new connective tissue represents a critical step in restoring tissue integrity. Split-thickness excisional wounds in the pig were studied by in situ hybridization in order to obtain subjective findings on the activity and location of cells involved in matrix gene expression after the administration of recombinant TGF-beta 1. Data focus on the stimulatory role of this growth factor in granulation tissue formation, on the enhanced mRNA content of collagen types I and III, fibronectin, TGF-beta 1 itself, and on the reduction in stromelysin mRNA, suggesting that increased matrix formation measured after treatment with TGF-beta 1 is due to fibroplasia regulated by the abundance of mRNAs for several different structural, matrix proteins as well as inhibition of proteolytic phenomena elicited by metalloproteinases. These studies reveal elastin mRNA early in the repair process, and elastin mRNA expression is enhanced by administration of TGF-beta 1. Moreover, we show that TGF-beta 1 was auto-stimulating in wounds, accounting, at least in part, for the persistent effects of single doses of this multipotential cytokine.

Incorporating Platelet-Rich Plasma into Electrospun Scaffolds for Tissue Engineering Applications

PubMed Central

Wolfe, Patricia S.; Ericksen, Jeffery J.; Simpson, David G.; Bowlin, Gary L.

2011-01-01

Platelet-rich plasma (PRP) therapy has seen a recent spike in clinical interest due to the potential that the highly concentrated platelet solutions hold for stimulating tissue repair and regeneration. The aim of this study was to incorporate PRP into a number of electrospun materials to determine how growth factors are eluted from the structures, and what effect the presence of these factors has on enhancing electrospun scaffold bioactivity. PRP underwent a freeze-thaw-freeze process to lyse platelets, followed by lyophilization to create a powdered preparation rich in growth factors (PRGF), which was subsequently added to the electrospinning process. Release of protein from scaffolds over time was quantified, along with the quantification of human macrophage and adipose-derived stem cell (ADSC) chemotaxis and proliferation. Protein assays demonstrated a sustained release of protein from PRGF-containing scaffolds at up to 35 days in culture. Scaffold bioactivity was enhanced as ADSCs demonstrated increased proliferation in the presence of PRGF, whereas macrophages demonstrated increased chemotaxis to PRGF. In conclusion, the work performed in this study demonstrated that the incorporation of PRGF into electrospun structures has a significant positive influence on the bioactivity of the scaffolds, and may prove beneficial in a number of tissue engineering applications. PMID:21679135

Tissue factor is an angiogenic-specific receptor for factor VII-targeted immunotherapy and photodynamic therapy.

PubMed

Hu, Zhiwei; Cheng, Jijun; Xu, Jie; Ruf, Wolfram; Lockwood, Charles J

2017-02-01

Identification of target molecules specific for angiogenic vascular endothelial cells (VEC), the inner layer of pathological neovasculature, is critical for discovery and development of neovascular-targeting therapy for angiogenesis-dependent human diseases, notably cancer, macular degeneration and endometriosis, in which vascular endothelial growth factor (VEGF) plays a central pathophysiological role. Using VEGF-stimulated vascular endothelial cells (VECs) isolated from microvessels, venous and arterial blood vessels as in vitro angiogenic models and unstimulated VECs as a quiescent VEC model, we examined the expression of tissue factor (TF), a membrane-bound receptor on the angiogenic VEC models compared with quiescent VEC controls. We found that TF is specifically expressed on angiogenic VECs in a time-dependent manner in microvessels, venous and arterial vessels. TF-targeted therapeutic agents, including factor VII (fVII)-IgG1 Fc and fVII-conjugated photosensitizer, can selectively bind angiogenic VECs, but not the quiescent VECs. Moreover, fVII-targeted photodynamic therapy can selectively and completely eradicate angiogenic VECs. We conclude that TF is an angiogenic-specific receptor and the target molecule for fVII-targeted therapeutics. This study supports clinical trials of TF-targeted therapeutics for the treatment of angiogenesis-dependent diseases such as cancer, macular degeneration and endometriosis.

Tissue Cancellation in Dual Energy Mammography Using a Calibration Phantom Customized for Direct Mapping.

PubMed

Han, Seokmin; Kang, Dong-Goo

2014-01-01

An easily implementable tissue cancellation method for dual energy mammography is proposed to reduce anatomical noise and enhance lesion visibility. For dual energy calibration, the images of an imaging object are directly mapped onto the images of a customized calibration phantom. Each pixel pair of the low and high energy images of the imaging object was compared to pixel pairs of the low and high energy images of the calibration phantom. The correspondence was measured by absolute difference between the pixel values of imaged object and those of the calibration phantom. Then the closest pixel pair of the calibration phantom images is marked and selected. After the calibration using direct mapping, the regions with lesion yielded different thickness from the background tissues. Taking advantage of the different thickness, the visibility of cancerous lesions was enhanced with increased contrast-to-noise ratio, depending on the size of lesion and breast thickness. However, some tissues near the edge of imaged object still remained after tissue cancellation. These remaining residuals seem to occur due to the heel effect, scattering, nonparallel X-ray beam geometry and Poisson distribution of photons. To improve its performance further, scattering and the heel effect should be compensated.

Adhesive protein interactions with chitosan: consequences for valve endothelial cell growth on tissue-engineering materials.

PubMed

Cuy, Janet L; Beckstead, Benjamin L; Brown, Chad D; Hoffman, Allan S; Giachelli, Cecilia M

2003-11-01

Stable endothelialization of a tissue-engineered heart valve is essential for proper valve function, although adhesive characteristics of the native valve endothelial cell (VEC) have rarely been explored. This research evaluated VEC adhesive qualities and attempted to enhance VEC growth on the biopolymer chitosan, a novel tissue-engineering scaffold material with promising biological and chemical properties. Aortic VEC cultures were isolated and found to preferentially adhere to fibronectin, collagen types IV and I over laminin and osteopontin in a dose-dependent manner. Seeding of VEC onto comparison substrates revealed VEC growth and morphology to be preferential in the order: tissue culture polystyrene > gelatin, poly(DL-lactide-co-glycolide), chitosan > poly(hydroxy alkanoate). Adhesive protein precoating of chitosan did not significantly enhance VEC growth, despite equivalent protein adsorption as to polystyrene. Initial cell adhesion to protein-precoated chitosan, however, was higher than for polystyrene. Composite chitosan/collagen type IV films were investigated as an alternative to simple protein precoatings, and were shown to improve VEC growth and morphology over chitosan alone. These findings suggest potential manipulation of chitosan properties to improve amenability to valve tissue-engineering applications. Copyright 2003 Wiley Periodicals, Inc.

Honeybee product therapeutic as stem cells homing for ovary failure.

PubMed

Safitri, Erma; Widiyatno, Thomas V; Prasetyo, R Heru

2016-11-01

Complexity of the method of isolation, cultivation in vitro and the expensive cost of transplantation process of stem cells, it would require an innovation to homing and differentiation of stem cells and increase folliculogenesis. The stem cells homing was achieved through the provision of food or beverages derived from natural materials like honeybee product. Through honeybee product, there will be homing of stem cells and accompany with the sources from the body itself will take place in regeneration of the ovary. Female rats model of degenerative ovary was obtained through food fasting but still have drinking water for 5 days. It caused malnutrition and damage of the ovarian tissue. The administration of 50% honeybee product (T1) was performed for 10 consecutive days, while the positive control group (T0+) was fasted and not given honeybee product and the negative control (T0-) not fasted and without honeybee product. Observations were taken for homing of stem cells, raised of folliculogenesis, differentiation of stem cells, and regeneration of the ovarian tissue using routine H&E staining. Homing of stem cells shown the vascular endothelial growth factor and granulocyte colony-stimulating factor expression; enhancement of folliculogenesis was indicated by an increase of follicle dee Graaf count; enhancement of differentiation of stem cells was indicated by growth differentiation factor-9 expression; and regeneration of ovarian tissue indicated by intact ovarian tissue with growing follicles. Honeybee product can be induced endogenous stem cells in regeneration of ovary failure due to malnutrition.

Wound healing potential of adipose tissue stem cell extract

DOE Office of Scientific and Technical Information (OSTI.GOV)

Na, You Kyung; Ban, Jae-Jun; Lee, Mijung

Adipose tissue stem cells (ATSCs) are considered as a promising source in the field of cell therapy and regenerative medicine. In addition to direct cell replacement using stem cells, intercellular molecule exchange by stem cell secretory factors showed beneficial effects by reducing tissue damage and augmentation of endogenous repair. Delayed cutaneous wound healing is implicated in many conditions such as diabetes, aging, stress and alcohol consumption. However, the effects of cell-free extract of ATSCs (ATSC-Ex) containing secretome on wound healing process have not been investigated. In this study, ATSC-Ex was topically applied on the cutaneous wound and healing speed wasmore » examined. As a result, wound closure was much faster in the cell-free extract treated wound than control wound at 4, 6, 8 days after application of ATSC-Ex. Dermal fibroblast proliferation, migration and extracellular matrix (ECM) production are critical aspects of wound healing, and the effects of ATSC-Ex on human dermal fibroblast (HDF) was examined. ATSC-Ex augmented HDF proliferation in a dose-dependent manner and migration ability was enhanced by extract treatment. Representative ECM proteins, collagen type I and matrix metalloproteinase-1, are significantly up-regulated by treatment of ATSC-Ex. Our results suggest that the ATSC-Ex have improving effect of wound healing and can be the potential therapeutic candidate for cutaneous wound healing. - Highlights: • Topical application of ATSC-Ex results in faster wound closure than normal wound in vivo. • ATSC-Ex enhances dermal fibroblast proliferation, migration and extracellular matrix production. • This study suggests that ATSC-Ex is an effective source to augment wound healing.« less

GABAA-current rundown of temporal lobe epilepsy is associated with repetitive activation of GABAA “phasic” receptors

PubMed Central

Palma, Eleonora; Roseti, Cristina; Maiolino, Francesca; Fucile, Sergio; Martinello, Katiuscia; Mazzuferi, Manuela; Aronica, Eleonora; Manfredi, Mario; Esposito, Vincenzo; Cantore, Gianpaolo; Miledi, Ricardo; Simonato, Michele; Eusebi, Fabrizio

2007-01-01

A study was made of the “rundown” of GABAA receptors, microtransplanted to Xenopus oocytes from surgically resected brain tissues of patients afflicted with drug-resistant human mesial temporal lobe epilepsy (mTLE). Cell membranes, isolated from mTLE neocortex specimens, were injected into frog oocytes that rapidly incorporated functional GABAA receptors. Upon repetitive activation with GABA (1 mM), “epileptic” GABAA receptors exhibited a GABAA-current (IGABA) rundown that was significantly enhanced by Zn2+ (≤250 μM), and practically abolished by the high-affinity GABAA receptor inverse agonist SR95531 (gabazine; 2.5–25 μM). Conversely, IGABA generated by “control” GABAA receptors microtransplanted from nonepileptic temporal lobe, lesional TLE, or authoptic disease-free tissues remained stable during repetitive stimulation, even in oocytes treated with Zn2+. We conclude that rundown of mTLE epileptic receptors depends on the presence of “phasic GABAA receptors” that have low sensitivity to antagonism by Zn2+. Additionally, we found that GABAA receptors, microtransplanted from the cerebral cortex of adult rats exhibiting recurrent seizures, caused by pilocarpine-induced status epilepticus, showed greater rundown than control tissue, an event also occurring in patch-clamped rat pyramidal neurons. Rundown of epileptic rat receptors resembled that of human mTLE receptors, being enhanced by Zn2+ (40 μM) and sensitive to the antiepileptic agent levetiracetam, the neurotrophin brain-derived neurotrophic factor, and the phosphatase blocker okadaic acid. Our findings point to the rundown of GABAA receptors as a hallmark of TLE and suggest that modulating tonic and phasic mTLE GABAA receptor activity may represent a useful therapeutic approach to the disease. PMID:18083839

High-level secretion of tissue factor-rich extracellular vesicles from ovarian cancer cells mediated by filamin-A and protease-activated receptors.

PubMed

Koizume, Shiro; Ito, Shin; Yoshioka, Yusuke; Kanayama, Tomohiko; Nakamura, Yoshiyasu; Yoshihara, Mitsuyo; Yamada, Roppei; Ochiya, Takahiro; Ruf, Wolfram; Miyagi, Etsuko; Hirahara, Fumiki; Miyagi, Yohei

2016-01-01

Thromboembolic events occur frequently in ovarian cancer patients. Tissue factor (TF) is often overexpressed in tumours, including ovarian clear-cell carcinoma (CCC), a subtype with a generally poor prognosis. TF-coagulation factor VII (fVII) complexes on the cell surface activate downstream coagulation mechanisms. Moreover, cancer cells secrete extracellular vesicles (EVs), which act as vehicles for TF. We therefore examined the characteristics of EVs produced by ovarian cancer cells of various histological subtypes. CCC cells secreted high levels of TF within EVs, while the high-TF expressing breast cancer cell line MDA-MB-231 shed fewer TF-positive EVs. We also found that CCC tumours with hypoxic tissue areas synthesised TF and fVII in vivo, rendering the blood of xenograft mice bearing these tumours hypercoagulable compared with mice bearing MDA-MB-231 tumours. Incorporation of TF into EVs and secretion of EVs from CCC cells exposed to hypoxia were both dependent on the actin-binding protein, filamin-A (filA). Furthermore, production of these EVs was dependent on different protease-activated receptors (PARs) on the cell surface. These results show that CCC cells could produce large numbers of TF-positive EVs dependent upon filA and PARs. This phenomenon may be the mechanism underlying the increased incidence of venous thromboembolism in ovarian cancer patients.

[Gigantism with low serum level of growth hormone: a case report].

PubMed

Ran, X; Zhang, L; Xiong, P; Zhao, T; Tong, N; Li, X

2001-12-01

Gigantism with low or normal basal concentrations of growth hormone (GH) is a rare condition, possibly due to abnormal GH secretory patterns, enhanced tissue sensitivity to GH, or the existence of an unidentified growth promoting factor. Here we report an 11 year-old female case of gigantism with a normal pituitary gland. Her height was 181 cm, body weight 77 kg, and bone age 11.1 years. Her basal serum GH levels were lower than 1 ng/ml. The levels of T3, T4, FT3, FT4, TSH, E2, LH, FSH, PRL, PTC and ACTH were normal. Serum GH response to insulin-induced hypoglycemia or arginine stimulation tests was blunted. In this case, non-pulsatile GH secretion and enhanced tissue sensitivity to GH may induce hypersecretion of IGF-1 and the existence of an unidentified growth promoting factor or biologically active anti-GH receptor antibodies may cause clinical gigantism.

Blood Flow Changes in Subsynovial Connective Tissue on Contrast-Enhanced Ultrasonography in Patients With Carpal Tunnel Syndrome Before and After Surgical Decompression.

PubMed

Motomiya, Makoto; Funakoshi, Tadanao; Ishizaka, Kinya; Nishida, Mutsumi; Matsui, Yuichiro; Iwasaki, Norimasa

2017-11-24

Although qualitative alteration of the subsynovial connective tissue in the carpal tunnel is considered to be one of the most important factors in the pathophysiologic mechanisms of carpal tunnel syndrome (CTS), little information is available about the microcirculation in the subsynovial connective tissue in patients with CTS. The aims of this study were to use contrast-enhanced ultrasonography (US) to evaluate blood flow in the subsynovial connective tissue proximal to the carpal tunnel in patients with CTS before and after carpal tunnel release. The study included 15 volunteers and 12 patients with CTS. The blood flow in the subsynovial connective tissue and the median nerve was evaluated preoperatively and at 1, 2, and 3 months postoperatively using contrast-enhanced US. The blood flow in the subsynovial connective tissue was higher in the patients with CTS than in the volunteers. In the patients with CTS, there was a significant correlation between the blood flow in the subsynovial connective tissue and the median nerve (P = .01). The blood flow in both the subsynovial connective tissue and the median nerve increased markedly after carpal tunnel release. Our results suggest that increased blood flow in the subsynovial connective tissue may play a role in the alteration of the microcirculation within the median nerve related to the pathophysiologic mechanisms of CTS. The increase in the blood flow in the subsynovial connective tissue during the early postoperative period may contribute to the changes in intraneural circulation, and these changes may lead to neural recovery. © 2017 by the American Institute of Ultrasound in Medicine.

Adult Stem Cell Based Enhancement of Nerve Conduit for Peripheral Nerve Repair

DTIC Science & Technology

2016-10-01

isolated stem cells from the injured tissue site that have wound healing promoting activities. In this application, we propose to use these cells, which may...Regeneration, Nanofiber, Neurotrophic Factor, Tissue Engineering, Multifunctional 3. ACCOMPLISHMENTS:  The major goals of this project are...have been stored in cell bank . Objective 9: Purchase reagents and materials for methacrylation of ECM hydrogel  All necessary reagents and

Cathepsin K in Lymphangioleiomyomatosis: LAM Cell-Fibroblast Interactions Enhance Protease Activity by Extracellular Acidification.

PubMed

Dongre, Arundhati; Clements, Debbie; Fisher, Andrew J; Johnson, Simon R

2017-08-01

Lymphangioleiomyomatosis (LAM) is a rare disease in which LAM cells and fibroblasts form lung nodules and it is hypothesized that LAM nodule-derived proteases cause cyst formation and tissue damage. On protease gene expression profiling in whole lung tissue, cathepsin K gene expression was 40-fold overexpressed in LAM compared with control lung tissue (P ≤ 0.0001). Immunohistochemistry confirmed cathepsin K protein was expressed in LAM but not control lungs. Cathepsin K gene expression and protein and protease activity were detected in LAM-associated fibroblasts but not the LAM cell line 621-101. In lung nodules, cathepsin K immunoreactivity predominantly co-localized with LAM-associated fibroblasts. In vitro, fibroblast extracellular cathepsin K activity was minimal at pH 7.5 but significantly enhanced at pH 7 and 6. 621-101 cells reduced extracellular pH with acidification dependent on 621-101 mechanistic target of rapamycin activity and net hydrogen ion exporters, particularly sodium bicarbonate co-transporters and carbonic anhydrases, which were also expressed in LAM lung tissue. In LAM cell-fibroblast co-cultures, acidification paralleled cathepsin K activity, and both were reduced by sodium bicarbonate co-transporter (P ≤ 0.0001) and carbonic anhydrase inhibitors (P = 0.0021). Our findings suggest that cathepsin K activity is dependent on LAM cell-fibroblast interactions, and inhibitors of extracellular acidification may be potential therapies for LAM. Copyright © 2017 American Society for Investigative Pathology. Published by Elsevier Inc. All rights reserved.

The neurobiology and addiction potential of anabolic androgenic steroids and the effects of growth hormone.

PubMed

Grönbladh, Alfhild; Nylander, Erik; Hallberg, Mathias

2016-09-01

Anabolic androgenic steroids (AAS) are substances that mimic the hormone testosterone, and primarily act via the androgen receptor. In addition to their physiological effect on muscle tissue and growth, research from the last decade has shown that AAS have a pronounced impact on the central nervous system. A large number of studies have demonstrated that AAS affect the mesolimbic reward system in the brain. However, whether the direct effects of AAS on endorphins, dopamine, serotonin and GABA etc. and on the corresponding and related systems lead to dependence needs to be further elucidated. According to recent studies, the prevalence of AAS dependence among AAS users has been estimated to be approximately 30%, and polysubstance use, of both pharmaceutical drugs and narcotics, within this group is common. The present review primarily discusses AAS in the context of addiction and dependence, and further addresses the issue of using multiple substances, i.e. stimulants and opiates in combination with AAS. In addition, aspects of the treatment of AAS dependence, the connection between AAS abuse and cognition, and AAS-induced neurotoxicity are presented. Currently, performance enhancing drugs are frequently used in combination with AAS. Therefore, a large section on growth hormone and insulin-like growth factor is also included. Copyright © 2016. Published by Elsevier Inc.

Hypoxia inducible factor-1 mediates the expression of the immune checkpoint HLA-G in glioma cells through hypoxia response element located in exon 2.

PubMed

Yaghi, Layale; Poras, Isabelle; Simoes, Renata T; Donadi, Eduardo A; Tost, Jörg; Daunay, Antoine; de Almeida, Bibiana Sgorla; Carosella, Edgardo D; Moreau, Philippe

2016-09-27

HLA-G is an immune checkpoint molecule with specific relevance in cancer immunotherapy. It was first identified in cytotrophoblasts, protecting the fetus from maternal rejection. HLA-G tissue expression is very restricted but induced in numerous malignant tumors such as glioblastoma, contributing to their immune escape. Hypoxia occurs during placenta and tumor development and was shown to activate HLA-G. We aimed to elucidate the mechanisms of HLA-G activation under conditions combining hypoxia-mimicking treatment and 5-aza-2'deoxycytidine, a DNA demethylating agent used in anti-cancer therapy which also induces HLA-G. Both treatments enhanced the amount of HLA-G mRNA and protein in HLA-G negative U251MG glioma cells. Electrophoretic Mobility Shift Assays and luciferase reporter gene assays revealed that HLA-G upregulation depends on Hypoxia Inducible Factor-1 (HIF-1) and a hypoxia responsive element (HRE) located in exon 2. A polymorphic HRE at -966 bp in the 5'UT region may modulate the magnitude of the response mediated by the exon 2 HRE. We suggest that therapeutic strategies should take into account that HLA-G expression in response to hypoxic tumor environment is dependent on HLA-G gene polymorphism and DNA methylation state at the HLA-G locus.

Ischemia/Reperfusion

PubMed Central

Kalogeris, Theodore; Baines, Christopher P.; Krenz, Maike; Korthuis, Ronald J.

2017-01-01

Ischemic disorders, such as myocardial infarction, stroke, and peripheral vascular disease, are the most common causes of debilitating disease and death in westernized cultures. The extent of tissue injury relates directly to the extent of blood flow reduction and to the length of the ischemic period, which influence the levels to which cellular ATP and intracellular pH are reduced. By impairing ATPase-dependent ion transport, ischemia causes intracellular and mitochondrial calcium levels to increase (calcium overload). Cell volume regulatory mechanisms are also disrupted by the lack of ATP, which can induce lysis of organelle and plasma membranes. Reperfusion, although required to salvage oxygen-starved tissues, produces paradoxical tissue responses that fuel the production of reactive oxygen species (oxygen paradox), sequestration of proinflammatory immunocytes in ischemic tissues, endoplasmic reticulum stress, and development of postischemic capillary no-reflow, which amplify tissue injury. These pathologic events culminate in opening of mitochondrial permeability transition pores as a common end-effector of ischemia/reperfusion (I/R)-induced cell lysis and death. Emerging concepts include the influence of the intestinal microbiome, fetal programming, epigenetic changes, and microparticles in the pathogenesis of I/R. The overall goal of this review is to describe these and other mechanisms that contribute to I/R injury. Because so many different deleterious events participate in I/R, it is clear that therapeutic approaches will be effective only when multiple pathologic processes are targeted. In addition, the translational significance of I/R research will be enhanced by much wider use of animal models that incorporate the complicating effects of risk factors for cardiovascular disease. PMID:28135002

An injectable elastin-based gene delivery platform for dose-dependent modulation of angiogenesis and inflammation for critical limb ischemia.

PubMed

Dash, Biraja C; Thomas, Dilip; Monaghan, Michael; Carroll, Oliver; Chen, Xizhe; Woodhouse, Kimberly; O'Brien, Timothy; Pandit, Abhay

2015-10-01

Critical limb ischemia is a major clinical problem. Despite rigorous treatment regimes, there has been only modest success in reducing the rate of amputations in affected patients. Reduced level of blood flow and enhanced inflammation are the two major pathophysiological changes that occur in the ischemic tissue. The objective of this study was to develop a controlled dual gene delivery system capable of delivering therapeutic plasmid eNOS and IL-10 in a temporal manner. In order to deliver multiple therapeutic genes, an elastin-like polypeptide (ELP) based injectable system was designed. The injectable system was comprised of hollow spheres and an in situ-forming gel scaffold of elastin-like polypeptide capable of carrying gene complexes, with an extended manner release profile. In addition, the ELP based injectable system was used to deliver human eNOS and IL-10 therapeutic genes in vivo. A subcutaneous dose response study showed enhanced blood vessel density in the treatment groups of eNOS (20 μg) and IL-10 (10 μg)/eNOS (20 μg) and reduced inflammation with IL-10 (10 μg) alone. Next, we carried out a hind-limb ischemia model comparing the efficacy of the following interventions; Saline; IL-10, eNOS and IL-10/eNOS. The selected dose of eNOS, exhibited enhanced angiogenesis. IL-10 treatment groups showed reduction in the level of inflammatory cells. Furthermore, we demonstrated that eNOS up-regulated major proangiogenic growth factors such as vascular endothelial growth factors, platelet derived growth factor B, and fibroblast growth factor 1, which may explain the mechanism of this approach. These factors help in formation of a stable vascular network. Thus, ELP injectable system mediating non-viral delivery of human IL10-eNOS is a promising therapy towards treating limb ischemia. Copyright © 2015 Elsevier Ltd. All rights reserved.

Characterization of the Influence of Semen-Derived Enhancer of Virus Infection on the Interaction of HIV-1 with Female Reproductive Tract Tissues

PubMed Central

Allen, Shannon A.; Carias, Ann M.; Anderson, Meegan R.; Okocha, Eneniziaogochukwu A.; Benning, Lorie; McRaven, Michael D.; Kelley, Z L.; Lurain, John; Veazey, Ronald S.

2015-01-01

ABSTRACT The majority of human immunodeficiency virus type 1 (HIV-1) transmission events occur in women when semen harboring infectious virus is deposited onto the mucosal barriers of the vaginal, ectocervical, and endocervical epithelia. Seminal factors such as semen-derived enhancer of virus infection (SEVI) fibrils were previously shown to greatly enhance the infectivity of HIV-1 in cell culture systems. However, when SEVI is intravaginally applied to living animals, there is no effect on vaginal transmission. To define how SEVI might function in the context of sexual transmission, we applied HIV-1 and SEVI to intact human and rhesus macaque reproductive tract tissues to determine how it influences virus interactions with these barriers. We show that SEVI binds HIV-1 and sequesters most virions to the luminal surface of the stratified squamous epithelium, significantly reducing the number of virions that penetrated the tissue. In the simple columnar epithelium, SEVI was no longer fibrillar in structure and was detached from virions but allowed significantly deeper epithelial virus penetration. These observations reveal that the action of SEVI in intact tissues is very different in the anatomical context of sexual transmission and begin to explain the lack of stimulation of infection observed in the highly relevant mucosal transmission model. IMPORTANCE The most common mode of HIV-1 transmission in women occurs via genital exposure to the semen of HIV-infected men. A productive infection requires the virus to penetrate female reproductive tract epithelial barriers to infect underlying target cells. Certain factors identified within semen, termed semen-derived enhancers of virus infection (SEVI), have been shown to significantly enhance HIV-1 infectivity in cell culture. However, when applied to the genital tracts of living female macaques, SEVI did not enhance virus transmission. Here we show that SEVI functions very differently in the context of intact mucosal tissues. SEVI decreases HIV-1 penetration of squamous epithelial barriers in humans and macaques. At the mucus-coated columnar epithelial barrier, the HIV-1/SEVI interaction is disrupted. These observations suggest that SEVI may not play a significant stimulatory role in the efficiency of male-to-female sexual transmission of HIV. PMID:25740984

Epigenetics in adipose tissue, obesity, weight loss and diabetes

USDA-ARS?s Scientific Manuscript database

Given the role that the diet and other environmental factors play in the development of obesity and type 2 diabetes, the implication of different epigenetic processes is being investigated. Although it is well known that the environmental factors can cause cell type-dependent epigenetic changes, inc...

Low-oxygen tensions found in Salmonella-infected gut tissue boost Salmonella replication in macrophages by impairing antimicrobial activity and augmenting Salmonella virulence.

PubMed

Jennewein, Jonas; Matuszak, Jasmin; Walter, Steffi; Felmy, Boas; Gendera, Kathrin; Schatz, Valentin; Nowottny, Monika; Liebsch, Gregor; Hensel, Michael; Hardt, Wolf-Dietrich; Gerlach, Roman G; Jantsch, Jonathan

2015-12-01

In Salmonella infection, the Salmonella pathogenicity island-2 (SPI-2)-encoded type three secretion system (T3SS2) is of key importance for systemic disease and survival in host cells. For instance, in the streptomycin-pretreated mouse model SPI-2-dependent Salmonella replication in lamina propria CD11c(-)CXCR1(-) monocytic phagocytes/macrophages (MΦ) is required for the development of colitis. In addition, containment of intracellular Salmonella in the gut critically depends on the antimicrobial effects of the phagocyte NADPH oxidase (PHOX), and possibly type 2 nitric oxide synthase (NOS2). For both antimicrobial enzyme complexes, oxygen is an essential substrate. However, the amount of available oxygen upon enteroinvasive Salmonella infection in the gut tissue and its impact on Salmonella-MΦ interactions was unknown. Therefore, we measured the gut tissue oxygen levels in a model of Salmonella enterocolitis using luminescence two-dimensional in vivo oxygen imaging. We found that gut tissue oxygen levels dropped from ∼78 Torr (∼11% O2) to values of ∼16 Torr (∼2% O2) during infection. Because in vivo virulence of Salmonella depends on the Salmonella survival in MΦ, Salmonella-MΦ interaction was analysed under such low oxygen values. These experiments revealed an increased intracellular replication and survival of wild-type and t3ss2 non-expressing Salmonella. These findings were paralleled by blunted nitric oxide and reactive oxygen species (ROS) production and reduced Salmonella ROS perception. In addition, hypoxia enhanced SPI-2 transcription and translocation of SPI-2-encoded virulence protein. Neither pharmacological blockade of PHOX and NOS2 nor impairment of T3SS2 virulence function alone mimicked the effect of hypoxia on Salmonella replication under normoxic conditions. However, if t3ss2 non-expressing Salmonella were used, hypoxia did not further enhance Salmonella recovery in a PHOX and NOS2-deficient situation. Hence, these data suggest that hypoxia-induced impairment of antimicrobial activity and Salmonella virulence cooperate to allow for enhanced Salmonella replication in MΦ. © 2015 John Wiley & Sons Ltd.

Normal breast tissue DNA methylation differences at regulatory elements are associated with the cancer risk factor age.

PubMed

Johnson, Kevin C; Houseman, E Andres; King, Jessica E; Christensen, Brock C

2017-07-10

The underlying biological mechanisms through which epidemiologically defined breast cancer risk factors contribute to disease risk remain poorly understood. Identification of the molecular changes associated with cancer risk factors in normal tissues may aid in determining the earliest events of carcinogenesis and informing cancer prevention strategies. Here we investigated the impact cancer risk factors have on the normal breast epigenome by analyzing DNA methylation genome-wide (Infinium 450 K array) in cancer-free women from the Susan G. Komen Tissue Bank (n = 100). We tested the relation of established breast cancer risk factors, age, body mass index, parity, and family history of disease, with DNA methylation adjusting for potential variation in cell-type proportions. We identified 787 cytosine-guanine dinucleotide (CpG) sites that demonstrated significant associations (Q value <0.01) with subject age. Notably, DNA methylation was not strongly associated with the other evaluated breast cancer risk factors. Age-related DNA methylation changes are primarily increases in methylation enriched at breast epithelial cell enhancer regions (P = 7.1E-20), and binding sites of chromatin remodelers (MYC and CTCF). We validated the age-related associations in two independent populations, using normal breast tissue samples (n = 18) and samples of normal tissue adjacent to tumor tissue (n = 97). The genomic regions classified as age-related were more likely to be regions altered in both pre-invasive (n = 40, P = 3.0E-03) and invasive breast tumors (n = 731, P = 1.1E-13). DNA methylation changes with age occur at regulatory regions, and are further exacerbated in cancer, suggesting that age influences breast cancer risk in part through its contribution to epigenetic dysregulation in normal breast tissue.

Gene Expression of Tissue-Specific Molecules in Ex vivo Dermacentor variabilis (Acari: Ixodidae) During Rickettsial Exposure

PubMed Central

SUNYAKUMTHORN, PIYANATE; PETCHAMPAI, NATTHIDA; GRASPERGE, BRITTON J.; KEARNEY, MICHAEL T.; SONENSHINE, DANIEL E.; MACALUSO, KEVIN R.

2014-01-01

Ticks serve as both vectors and the reservoir hosts capable of transmitting spotted fever group Rickettsia by horizontal and vertical transmission. Persistent maintenance of Rickettsia species in tick populations is dependent on the specificity of the tick and Rickettsia relationship that limits vertical transmission of particular Rickettsia species, suggesting host-derived mechanisms of control. Tick-derived molecules are differentially expressed in a tissue-specific manner in response to rickettsial infection; however, little is known about tick response to specific rickettsial species. To test the hypothesis that tissue-specific tick-derived molecules are uniquely responsive to rickettsial infection, a bioassay to characterize the tick tissue-specific response to different rickettsial species was used. Whole organs of Dermacentor variabilis (Say) were exposed to either Rickettsia montanensis or Rickettsia amblyommii, two Rickettsia species common, or absent, in field-collected D. variabilis, respectively, for 1 and 12 h and harvested for quantitative real time-polymerase chain reaction assays of putative immune-like tick-derived factors. The results indicated that tick genes are differently expressed in a temporal and tissue-specific manner. Genes encoding glutathione S-transferase 1 (dvgst1) and Kunitz protease inhibitor (dvkpi) were highly expressed in midgut, and rickettsial exposure downregulated the expression of both genes. Two other genes encoding glutathione S-transferase 2 (dvgst2) and β-thymosin (dvβ-thy) were highly expressed in ovary, with dvβ-thy expression significantly downregulated in ovaries exposed to R. montanensis, but not R. amblyommii, at 12-h postexposure, suggesting a selective response. Deciphering the tissue-specific molecular interactions between tick and Rickettsia will enhance our understanding of the key mechanisms that mediate rickettsial infection in ticks. PMID:24180114

Understanding Mechanobiology: Physical Therapists as a Force in Mechanotherapy and Musculoskeletal Regenerative Rehabilitation

PubMed Central

Thompson, William R.; Scott, Alexander; Loghmani, M. Terry; Ward, Samuel R.

2016-01-01

Achieving functional restoration of diseased or injured tissues is the ultimate goal of both regenerative medicine approaches and physical therapy interventions. Proper integration and healing of the surrogate cells, tissues, or organs introduced using regenerative medicine techniques are often dependent on the co-introduction of therapeutic physical stimuli. Thus, regenerative rehabilitation represents a collaborative approach whereby rehabilitation specialists, basic scientists, physicians, and surgeons work closely to enhance tissue restoration by creating tailored rehabilitation treatments. One of the primary treatment regimens that physical therapists use to promote tissue healing is the introduction of mechanical forces, or mechanotherapies. These mechanotherapies in regenerative rehabilitation activate specific biological responses in musculoskeletal tissues to enhance the integration, healing, and restorative capacity of implanted cells, tissues, or synthetic scaffolds. To become future leaders in the field of regenerative rehabilitation, physical therapists must understand the principles of mechanobiology and how mechanotherapies augment tissue responses. This perspective article provides an overview of mechanotherapy and discusses how mechanical signals are transmitted at the tissue, cellular, and molecular levels. The synergistic effects of physical interventions and pharmacological agents also are discussed. The goals are to highlight the critical importance of mechanical signals on biological tissue healing and to emphasize the need for collaboration within the field of regenerative rehabilitation. As this field continues to emerge, physical therapists are poised to provide a critical contribution by integrating mechanotherapies with regenerative medicine to restore musculoskeletal function. PMID:26637643

Role of neurotrophins in the development and function of neural circuits that regulate energy homeostasis.

PubMed

Fargali, Samira; Sadahiro, Masato; Jiang, Cheng; Frick, Amy L; Indall, Tricia; Cogliani, Valeria; Welagen, Jelle; Lin, Wei-Jye; Salton, Stephen R

2012-11-01

Members of the neurotrophin family, including nerve growth factor, brain-derived neurotrophic factor, neurotrophin-3, and neurotrophin-4/5, and other neurotrophic growth factors such as ciliary neurotrophic factor and artemin, regulate peripheral and central nervous system development and function. A subset of the neurotrophin-dependent pathways in the hypothalamus, brainstem, and spinal cord, and those that project via the sympathetic nervous system to peripheral metabolic tissues including brown and white adipose tissue, muscle and liver, regulate feeding, energy storage, and energy expenditure. We briefly review the role that neurotrophic growth factors play in energy balance, as regulators of neuronal survival and differentiation, neurogenesis, and circuit formation and function, and as inducers of critical gene products that control energy homeostasis.

Monte Carlo modeling of light-tissue interactions in narrow band imaging.

PubMed

Le, Du V N; Wang, Quanzeng; Ramella-Roman, Jessica C; Pfefer, T Joshua

2013-01-01

Light-tissue interactions that influence vascular contrast enhancement in narrow band imaging (NBI) have not been the subject of extensive theoretical study. In order to elucidate relevant mechanisms in a systematic and quantitative manner we have developed and validated a Monte Carlo model of NBI and used it to study the effect of device and tissue parameters, specifically, imaging wavelength (415 versus 540 nm) and vessel diameter and depth. Simulations provided quantitative predictions of contrast-including up to 125% improvement in small, superficial vessel contrast for 415 over 540 nm. Our findings indicated that absorption rather than scattering-the mechanism often cited in prior studies-was the dominant factor behind spectral variations in vessel depth-selectivity. Narrow-band images of a tissue-simulating phantom showed good agreement in terms of trends and quantitative values. Numerical modeling represents a powerful tool for elucidating the factors that affect the performance of spectral imaging approaches such as NBI.

Energy absorption buildup factors, exposure buildup factors and Kerma for optically stimulated luminescence materials and their tissue equivalence for radiation dosimetry

NASA Astrophysics Data System (ADS)

Singh, Vishwanath P.; Badiger, N. M.

2014-11-01

Optically stimulated luminescence (OSL) materials are sensitive dosimetric materials used for precise and accurate dose measurement for low-energy ionizing radiation. Low dose measurement capability with improved sensitivity makes these dosimeters very useful for diagnostic imaging, personnel monitoring and environmental radiation dosimetry. Gamma ray energy absorption buildup factors and exposure build factors were computed for OSL materials using the five-parameter Geometric Progression (G-P) fitting method in the energy range 0.015-15 MeV for penetration depths up to 40 mean free path. The computed energy absorption buildup factor and exposure buildup factor values were studied as a function of penetration depth and incident photon energy. Effective atomic numbers and Kerma relative to air of the selected OSL materials and tissue equivalence were computed and compared with that of water, PMMA and ICRU standard tissues. The buildup factors and kerma relative to air were found dependent upon effective atomic numbers. Buildup factors determined in the present work should be useful in radiation dosimetry, medical diagnostics and therapy, space dosimetry, accident dosimetry and personnel monitoring.

Protein disulfide isomerase acts as an injury response signal that enhances fibrin generation via tissue factor activation

PubMed Central

Reinhardt, Christoph; von Brühl, Marie-Luise; Manukyan, Davit; Grahl, Lenka; Lorenz, Michael; Altmann, Berid; Dlugai, Silke; Hess, Sonja; Konrad, Ildiko; Orschiedt, Lena; Mackman, Nigel; Ruddock, Lloyd; Massberg, Steffen; Engelmann, Bernd

2008-01-01

The activation of initiator protein tissue factor (TF) is likely to be a crucial step in the blood coagulation process, which leads to fibrin formation. The stimuli responsible for inducing TF activation are largely undefined. Here we show that the oxidoreductase protein disulfide isomerase (PDI) directly promotes TF-dependent fibrin production during thrombus formation in vivo. After endothelial denudation of mouse carotid arteries, PDI was released at the injury site from adherent platelets and disrupted vessel wall cells. Inhibition of PDI decreased TF-triggered fibrin formation in different in vivo murine models of thrombus formation, as determined by intravital fluorescence microscopy. PDI infusion increased — and, under conditions of decreased platelet adhesion, PDI inhibition reduced — fibrin generation at the injury site, indicating that PDI can directly initiate blood coagulation. In vitro, human platelet–secreted PDI contributed to the activation of cryptic TF on microvesicles (microparticles). Mass spectrometry analyses indicated that part of the extracellular cysteine 209 of TF was constitutively glutathionylated. Mixed disulfide formation contributed to maintaining TF in a state of low functionality. We propose that reduced PDI activates TF by isomerization of a mixed disulfide and a free thiol to an intramolecular disulfide. Our findings suggest that disulfide isomerases can act as injury response signals that trigger the activation of fibrin formation following vessel injury. PMID:18274674

Protein disulfide isomerase acts as an injury response signal that enhances fibrin generation via tissue factor activation.

PubMed

Reinhardt, Christoph; von Brühl, Marie-Luise; Manukyan, Davit; Grahl, Lenka; Lorenz, Michael; Altmann, Berid; Dlugai, Silke; Hess, Sonja; Konrad, Ildiko; Orschiedt, Lena; Mackman, Nigel; Ruddock, Lloyd; Massberg, Steffen; Engelmann, Bernd

2008-03-01

The activation of initiator protein tissue factor (TF) is likely to be a crucial step in the blood coagulation process, which leads to fibrin formation. The stimuli responsible for inducing TF activation are largely undefined. Here we show that the oxidoreductase protein disulfide isomerase (PDI) directly promotes TF-dependent fibrin production during thrombus formation in vivo. After endothelial denudation of mouse carotid arteries, PDI was released at the injury site from adherent platelets and disrupted vessel wall cells. Inhibition of PDI decreased TF-triggered fibrin formation in different in vivo murine models of thrombus formation, as determined by intravital fluorescence microscopy. PDI infusion increased - and, under conditions of decreased platelet adhesion, PDI inhibition reduced - fibrin generation at the injury site, indicating that PDI can directly initiate blood coagulation. In vitro, human platelet-secreted PDI contributed to the activation of cryptic TF on microvesicles (microparticles). Mass spectrometry analyses indicated that part of the extracellular cysteine 209 of TF was constitutively glutathionylated. Mixed disulfide formation contributed to maintaining TF in a state of low functionality. We propose that reduced PDI activates TF by isomerization of a mixed disulfide and a free thiol to an intramolecular disulfide. Our findings suggest that disulfide isomerases can act as injury response signals that trigger the activation of fibrin formation following vessel injury.

The stem cell factor (SCF)/c-KIT signalling in testis and prostate cancer.

PubMed

Cardoso, Henrique J; Figueira, Marília I; Socorro, Sílvia

2017-12-01

The stem cell factor (SCF) is a cytokine that specifically binds the tyrosine kinase receptor c-KIT. The SCF/c-KIT interaction leads to receptor dimerization, activation of kinase activity and initiation of several signal transduction pathways that control cell proliferation, apoptosis, differentiation and migration in several tissues. The activity of SCF/c-KIT system is linked with the phosphatidylinositol 3-kinase (PI3-K), the Src, the Janus kinase/signal transducers and activators of transcription (JAK/STAT), the phospholipase-C (PLC-γ) and the mitogen-activated protein kinase (MAPK) pathways. Moreover, it has been reported that cancer cases display an overactivation of c-KIT due to the presence of gain-of-function mutations or receptor overexpression, which renders c-KIT a tempting target for cancer treatment. In the case of male cancers the most documented activated pathways are the PI3-K and Src, both enhancing abnormal cell proliferation. It is also known that the Src activity in prostate cancer cases depends on the presence of tr-KIT, the cytoplasmic truncated variant of c-KIT that is specifically expressed in tumour tissues and, thus, a very interesting target for drug development. The present review provides an overview of the signalling pathways activated by SCF/c-KIT and discusses the potential application of c-KIT inhibitors for treatment of testicular and prostatic cancers.

Immunohistochemical detection of active transforming growth factor-beta in situ using engineered tissue

NASA Technical Reports Server (NTRS)

Barcellos-Hoff, M. H.; Ehrhart, E. J.; Kalia, M.; Jirtle, R.; Flanders, K.; Tsang, M. L.; Chatterjee, A. (Principal Investigator)

1995-01-01

The biological activity of transforming growth factor-beta 1 (TGF-beta) is governed by dissociation from its latent complex. Immunohistochemical discrimination of active and latent TGF-beta could provide insight into TGF-beta activation in physiological and pathological processes. However, evaluation of immunoreactivity specificity in situ has been hindered by the lack of tissue in which TGF-beta status is known. To provide in situ analysis of antibodies to differentiate between these functional forms, we used xenografts of human tumor cells modified by transfection to overexpress latent TGF-beta or constitutively active TGF-beta. This comparison revealed that, whereas most antibodies did not differentiate between TGF-beta activation status, the immunoreactivity of some antibodies was activation dependent. Two widely used peptide antibodies to the amino-terminus of TGF-beta, LC(1-30) and CC(1-30) showed marked preferential immunoreactivity with active TGF-beta versus latent TGF-beta in cryosections. However, in formalin-fixed, paraffin-embedded tissue, discrimination of active TGF-beta by CC(1-30) was lost and immunoreactivity was distinctly extracellular, as previously reported for this antibody. Similar processing-dependent extracellular localization was found with a neutralizing antibody raised to recombinant TGF-beta. Antigen retrieval recovered cell-associated immunoreactivity of both antibodies. Two antibodies to peptides 78-109 showed mild to moderate preferential immunoreactivity with active TGF-beta only in paraffin sections. LC(1-30) was the only antibody tested that discriminated active from latent TGF-beta in both frozen and paraffin-embedded tissue. Thus, in situ discrimination of active versus latent TGF-beta depends on both the antibody and tissue preparation. We propose that tissues engineered to express a specific form of a given protein provide a physiological setting in which to evaluate antibody reactivity with specific functional forms of a protein.

The Good the Bad and the Ugly of Glycosaminoglycans in Tissue Engineering Applications

PubMed Central

Ayerst, Bethanie I.; Merry, Catherine L.R.; Day, Anthony J.

2017-01-01

High sulfation, low cost, and the status of heparin as an already FDA- and EMA- approved product, mean that its inclusion in tissue engineering (TE) strategies is becoming increasingly popular. However, the use of heparin may represent a naïve approach. This is because tissue formation is a highly orchestrated process, involving the temporal expression of numerous growth factors and complex signaling networks. While heparin may enhance the retention and activity of certain growth factors under particular conditions, its binding ‘promiscuity’ means that it may also inhibit other factors that, for example, play an important role in tissue maintenance and repair. Within this review we focus on articular cartilage, highlighting the complexities and highly regulated processes that are involved in its formation, and the challenges that exist in trying to effectively engineer this tissue. Here we discuss the opportunities that glycosaminoglycans (GAGs) may provide in advancing this important area of regenerative medicine, placing emphasis on the need to move away from the common use of heparin, and instead focus research towards the utility of specific GAG preparations that are able to modulate the activity of growth factors in a more controlled and defined manner, with less off-target effects. PMID:28608822

Mitochondrial remodeling in adipose tissue associated with obesity and treatment with rosiglitazone

PubMed Central

Wilson-Fritch, Leanne; Nicoloro, Sarah; Chouinard, My; Lazar, Mitchell A.; Chui, Patricia C.; Leszyk, John; Straubhaar, Juerg; Czech, Michael P.; Corvera, Silvia

2004-01-01

Adipose tissue plays a central role in the control of energy homeostasis through the storage and turnover of triglycerides and through the secretion of factors that affect satiety and fuel utilization. Agents that enhance insulin sensitivity, such as rosiglitazone, appear to exert their therapeutic effect through adipose tissue, but the precise mechanisms of their actions are unclear. Rosiglitazone changes the morphological features and protein profiles of mitochondria in 3T3-L1 adipocytes. To examine the relevance of these effects in vivo, we studied white adipocytes from ob/ob mice during the development of obesity and after treatment with rosiglitazone. The levels of approximately 50% of gene transcripts encoding mitochondrial proteins were decreased with the onset of obesity. About half of those genes were upregulated after treatment with rosiglitazone, and this was accompanied by an increase in mitochondrial mass and changes in mitochondrial structure. Functionally, adipocytes from rosiglitazone-treated mice displayed markedly enhanced oxygen consumption and significantly increased palmitate oxidation. These data reveal mitochondrial remodeling and increased energy expenditure in white fat in response to rosiglitazone treatment in vivo and suggest that enhanced lipid utilization in this tissue may affect whole-body energy homeostasis and insulin sensitivity. PMID:15520860

The significance of cell-related challenges in the clinical application of tissue engineering.

PubMed

Almela, Thafar; Brook, Ian M; Moharamzadeh, Keyvan

2016-12-01

Tissue engineering is increasingly being recognized as a new approach that could alleviate the burden of tissue damage currently managed with transplants or synthetic devices. Making this novel approach available in the future for patients who would potentially benefit is largely dependent on understanding and addressing all those factors that impede the translation of this technology to the clinic. Cell-associated factors in particular raise many challenges, including those related to cell sources, up- and downstream techniques, preservation, and the creation of in vitro microenvironments that enable cells to grow and function as far as possible as they would in vivo. This article highlights the main confounding issues associated with cells in tissue engineering and how these issues may hinder the advancement of therapeutic tissue engineering. © 2016 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 104A: 3157-3163, 2016. © 2016 Wiley Periodicals, Inc.

Comprehensive evaluation of disease- and trait-specific enrichment for eight functional elements among GWAS-identified variants.

PubMed

Markunas, Christina A; Johnson, Eric O; Hancock, Dana B

2017-07-01

Genome-wide association study (GWAS)-identified variants are enriched for functional elements. However, we have limited knowledge of how functional enrichment may differ by disease/trait and tissue type. We tested a broad set of eight functional elements for enrichment among GWAS-identified SNPs (p < 5×10 -8 ) from the NHGRI-EBI Catalog across seven disease/trait categories: cancer, cardiovascular disease, diabetes, autoimmune disease, psychiatric disease, neurological disease, and anthropometric traits. SNPs were annotated using HaploReg for the eight functional elements across any tissue: DNase sites, expression quantitative trait loci (eQTL), sequence conservation, enhancers, promoters, missense variants, sequence motifs, and protein binding sites. In addition, tissue-specific annotations were considered for brain vs. blood. Disease/trait SNPs were compared to a control set of 4809 SNPs matched to the GWAS SNPs (N = 1639) on allele frequency, gene density, distance to nearest gene, and linkage disequilibrium at ~3:1 ratio. Enrichment analyses were conducted using logistic regression, with Bonferroni correction. Overall, a significant enrichment was observed for all functional elements, except sequence motifs. Missense SNPs showed the strongest magnitude of enrichment. eQTLs were the only functional element significantly enriched across all diseases/traits. Magnitudes of enrichment were generally similar across diseases/traits, where enrichment was statistically significant. Blood vs. brain tissue effects on enrichment were dependent on disease/trait and functional element (e.g., cardiovascular disease: eQTLs P TissueDifference  = 1.28 × 10 -6 vs. enhancers P TissueDifference  = 0.94). Identifying disease/trait-relevant functional elements and tissue types could provide new insight into the underlying biology, by guiding a priori GWAS analyses (e.g., brain enhancer elements for psychiatric disease) or facilitating post hoc interpretation.

Molecular stress-inducing compounds increase osteoclast formation in a heat shock factor 1 protein-dependent manner.

PubMed

Chai, Ryan C; Kouspou, Michelle M; Lang, Benjamin J; Nguyen, Chau H; van der Kraan, A Gabrielle J; Vieusseux, Jessica L; Lim, Reece C; Gillespie, Matthew T; Benjamin, Ivor J; Quinn, Julian M W; Price, John T

2014-05-09

Many anticancer therapeutic agents cause bone loss, which increases the risk of fractures that severely reduce quality of life. Thus, in drug development, it is critical to identify and understand such effects. Anticancer therapeutic and HSP90 inhibitor 17-(allylamino)-17-demethoxygeldanamycin (17-AAG) causes bone loss by increasing osteoclast formation, but the mechanism underlying this is not understood. 17-AAG activates heat shock factor 1 (Hsf1), the master transcriptional regulator of heat shock/cell stress responses, which may be involved in this negative action of 17-AAG upon bone. Using mouse bone marrow and RAW264.7 osteoclast differentiation models we found that HSP90 inhibitors that induced a heat shock response also enhanced osteoclast formation, whereas HSP90 inhibitors that did not (including coumermycin A1 and novobiocin) did not affect osteoclast formation. Pharmacological inhibition or shRNAmir knockdown of Hsf1 in RAW264.7 cells as well as the use of Hsf1 null mouse bone marrow cells demonstrated that 17-AAG-enhanced osteoclast formation was Hsf1-dependent. Moreover, ectopic overexpression of Hsf1 enhanced 17-AAG effects upon osteoclast formation. Consistent with these findings, protein levels of the essential osteoclast transcription factor microphthalmia-associated transcription factor were increased by 17-AAG in an Hsf1-dependent manner. In addition to HSP90 inhibitors, we also identified that other agents that induced cellular stress, such as ethanol, doxorubicin, and methotrexate, also directly increased osteoclast formation, potentially in an Hsf1-dependent manner. These results, therefore, indicate that cellular stress can enhance osteoclast differentiation via Hsf1-dependent mechanisms and may significantly contribute to pathological and therapeutic related bone loss.

Silencing of ATF2 inhibits growth of pancreatic cancer cells and enhances sensitivity to chemotherapy.

PubMed

Li, Mu; Wu, Xingda; Liu, Ning; Li, Xiaoying; Meng, Fanbin; Song, Shaowei

2017-06-01

Pancreatic cancer is one of the leading causes of cancer-related death worldwide. Activating transcription factor 2 (ATF2) is a multifunctional transcription factor, and is implicated in tumor progress, yet its role in pancreatic cancer remains unclear. In the present study, the level of ATF2 in pancreatic cancer tissues and the adjacent non-tumorous tissues was detected by quantitative real-time PCR and Western blot. The roles of ATF2 in the proliferation, cell cycle, and apoptosis of pancreatic cancer cells were investigated through ATF2 silencing, and the effect of ATF2 shRNA on the sensitivity of pancreatic cancer cells to gemcitabine, an anti-tumor drug, was explored. The results of our study showed that the ATF2 level in the pancreatic cancer tissues was higher than that in the adjacent non-tumorous tissues. Silencing of ATF2 was found to inhibit proliferation, arrest cell cycle at G1 phase and induce apoptosis in pancreatic cancer cells. Moreover, ATF2 silencing enhanced gemcitabine-induced growth-inhibition and apoptosis-induction effects in pancreatic cancer cells. In summary, silencing of ATF2 inhibited the growth of pancreatic cancer cells and enhanced the anti-tumor effects of gemcitabine, suggesting that ATF2 plays a pro-survival role in pancreatic cancer. Our results also propose that a high level of ATF2 may serve as a potential biomarker of pancreatic cancer, and that ATF2 may become a potential target for anti-tumor therapy. © 2017 International Federation for Cell Biology.

Enalapril and captopril enhance glutathione-dependent antioxidant defenses in mouse tissues.

PubMed

de Cavanagh, E M; Inserra, F; Ferder, L; Fraga, C G

2000-03-01

The effect of enalapril and captopril on total glutathione content (GSSG + GSH) and selenium-dependent glutathione peroxidase (Se-GPx) and glutathione reductase (GSSG-Rd) activities was investigated in mouse tissues. CF-1 mice (4-mo-old females) received water containing enalapril (20 mg/l) or captopril (50 mg/l) for 11 wk. Enalapril increased GSSG + GSH content (P < 0.05) in erythrocytes (147%), brain (112%), and lung (67%), and captopril increased GSSG + GSH content in erythrocytes (190%) and brain (132%). Enalapril enhanced Se-GPx activity in kidney cortex (42%) and kidney medulla (23%) and captopril in kidney cortex (30%). GSSG-Rd activity was enhanced by enalapril in erythrocytes (21%), brain (21%), liver (18%), and kidney cortex (53%) and by captopril in erythrocytes (25%), brain (19%), and liver (34%). In vitro erythrocyte oxidant stress was evaluated by thiobarbituric acid-reactive substances (TBARS) production (control 365 +/- 11, enalapril 221 +/- 26, captopril 206 +/- 17 nmol TBARS x g Hb(-1) x h(-1); both P < 0.05 vs. control) and phenylhydrazine-induced methemoglobin (MetHb) formation (control 66.5 +/- 3.5, enalapril 52.9 +/- 0.4, captopril: 56.4 +/- 2.9 micromol MetHb/g Hb; both P < 0.05 vs. control). Both angiotensin-converting enzyme inhibitor treatments were associated with increased nitric oxide production, as assessed by plasma NO-(3) + NO-(2) level determination (control 9.22 +/- 0.64, enalapril 13.7 +/- 1.9, captopril 17.3 +/- 3.0 micromol NO-(3) + NO-(2)/l plasma; both P < 0.05 vs. control). These findings support our previous reports on the enalapril- and captopril-induced enhancement of endogenous antioxidant defenses and include new data on glutathione-dependent defenses, thus furthering current knowledge on the association of ACE inhibition and antioxidants.

MGMT DNA repair gene promoter/enhancer haplotypes alter transcription factor binding and gene expression.

PubMed

Xu, Meixiang; Cross, Courtney E; Speidel, Jordan T; Abdel-Rahman, Sherif Z

2016-10-01

The O 6 -methylguanine-DNA methyltransferase (MGMT) protein removes O 6 -alkyl-guanine adducts from DNA. MGMT expression can thus alter the sensitivity of cells and tissues to environmental and chemotherapeutic alkylating agents. Previously, we defined the haplotype structure encompassing single nucleotide polymorphisms (SNPs) in the MGMT promoter/enhancer (P/E) region and found that haplotypes, rather than individual SNPs, alter MGMT promoter activity. The exact mechanism(s) by which these haplotypes exert their effect on MGMT promoter activity is currently unknown, but we noted that many of the SNPs comprising the MGMT P/E haplotypes are located within or in close proximity to putative transcription factor binding sites. Thus, these haplotypes could potentially affect transcription factor binding and, subsequently, alter MGMT promoter activity. In this study, we test the hypothesis that MGMT P/E haplotypes affect MGMT promoter activity by altering transcription factor (TF) binding to the P/E region. We used a promoter binding TF profiling array and a reporter assay to evaluate the effect of different P/E haplotypes on TF binding and MGMT expression, respectively. Our data revealed a significant difference in TF binding profiles between the different haplotypes evaluated. We identified TFs that consistently showed significant haplotype-dependent binding alterations (p ≤ 0.01) and revealed their role in regulating MGMT expression using siRNAs and a dual-luciferase reporter assay system. The data generated support our hypothesis that promoter haplotypes alter the binding of TFs to the MGMT P/E and, subsequently, affect their regulatory function on MGMT promoter activity and expression level.

Loss of function of fatty acid desturase7 in tomato enhances basal aphid resistance in a salicylate-dependent manner

USDA-ARS?s Scientific Manuscript database

Jasmonic acid (JA) and its derivatives mediate induced resistance against caterpillars and other herbivores that cause tissue disruption. Far less is known about the role of jasmonates in plant interactions with phloem-feeding insects such as aphids. This study compared responses in tomato (Solanu...

Enhanced Flexible Tubular Microelectrode with Conducting Polymer for Multi-Functional Implantable Tissue-Machine Interface

NASA Astrophysics Data System (ADS)

Tian, Hong-Chang; Liu, Jing-Quan; Kang, Xiao-Yang; Tang, Long-Jun; Wang, Ming-Hao; Ji, Bo-Wen; Yang, Bin; Wang, Xiao-Lin; Chen, Xiang; Yang, Chun-Sheng

2016-05-01

Implantable biomedical microdevices enable the restoration of body function and improvement of health condition. As the interface between artificial machines and natural tissue, various kinds of microelectrodes with high density and tiny size were developed to undertake precise and complex medical tasks through electrical stimulation and electrophysiological recording. However, if only the electrical interaction existed between electrodes and muscle or nerve tissue without nutrition factor delivery, it would eventually lead to a significant symptom of denervation-induced skeletal muscle atrophy. In this paper, we developed a novel flexible tubular microelectrode integrated with fluidic drug delivery channel for dynamic tissue implant. First, the whole microelectrode was made of biocompatible polymers, which could avoid the drawbacks of the stiff microelectrodes that are easy to be broken and damage tissue. Moreover, the microelectrode sites were circumferentially distributed on the surface of polymer microtube in three dimensions, which would be beneficial to the spatial selectivity. Finally, the in vivo results confirmed that our implantable tubular microelectrodes were suitable for dynamic electrophysiological recording and simultaneous fluidic drug delivery, and the electrode performance was further enhanced by the conducting polymer modification.

Enhanced Flexible Tubular Microelectrode with Conducting Polymer for Multi-Functional Implantable Tissue-Machine Interface.

PubMed

Tian, Hong-Chang; Liu, Jing-Quan; Kang, Xiao-Yang; Tang, Long-Jun; Wang, Ming-Hao; Ji, Bo-Wen; Yang, Bin; Wang, Xiao-Lin; Chen, Xiang; Yang, Chun-Sheng

2016-05-27

Implantable biomedical microdevices enable the restoration of body function and improvement of health condition. As the interface between artificial machines and natural tissue, various kinds of microelectrodes with high density and tiny size were developed to undertake precise and complex medical tasks through electrical stimulation and electrophysiological recording. However, if only the electrical interaction existed between electrodes and muscle or nerve tissue without nutrition factor delivery, it would eventually lead to a significant symptom of denervation-induced skeletal muscle atrophy. In this paper, we developed a novel flexible tubular microelectrode integrated with fluidic drug delivery channel for dynamic tissue implant. First, the whole microelectrode was made of biocompatible polymers, which could avoid the drawbacks of the stiff microelectrodes that are easy to be broken and damage tissue. Moreover, the microelectrode sites were circumferentially distributed on the surface of polymer microtube in three dimensions, which would be beneficial to the spatial selectivity. Finally, the in vivo results confirmed that our implantable tubular microelectrodes were suitable for dynamic electrophysiological recording and simultaneous fluidic drug delivery, and the electrode performance was further enhanced by the conducting polymer modification.

[MORPHOLOGICAL CHANGES OF THE LIVER IN OBTURATION JAUNDICE, CAUSED BY CHOLEDOCHOLITHIASIS, DEPENDING ON ITS DURATION].

PubMed

Sipliviy, V A; Yevtushenko, D V; Naumova, O V; Andreyeshchev, S A; Yevtushenko, A V

2016-02-01

Abstract The results of surgical treatment of 184 patients for obturation jaundice, caused by choledocholithiasis, were analyzed. Morphological changes of the liver were studied in 20 patients. There were three groups of patients delineated, depending on the obturation jaundice duration: up to 7 days, from 8 to 14 days, more than 15 days, and also a group of patients after the bile outflow restoration. The obturation jaundice occurrence in choledocholithiasis is accompanied by significant morphological changes in the liver, severity of which is enhancing while the obturation jaundice persistence increasing. While persistence of obturation jaundice through 8 days and more the connective tissue volume is enhancing, a relative volume of hepatocytes is reducing and a stromal-parenchymatous index is increasing. The bile outflow restoration secures significant reduction of intensity of alterative and inflammatory changes in hepatic parenchyma, as well as activation of reparative processes in the tissue. In cholangitis, caused by P. aeruginosa and E. coli, according to morphological investigations data, in the liver a diffuse purulent cholangitis on background of chronic changes in accordance to duration of the obturation jaundice persists.

The Probiotic Mixture VSL#3 Accelerates Gastric Ulcer Healing by Stimulating Vascular Endothelial Growth Factor

PubMed Central

Dharmani, Poonam; De Simone, Claudio; Chadee, Kris

2013-01-01

Studies assessing the effect and mechanism of probiotics on diseases of the upper gastrointestinal tract (GI) including gastric ulcers are limited despite extensive work and promising results of this therapeutic option for other GI diseases. In this study, we investigated the mechanisms by which the probiotic mixture VSL#3 (a mixture of eight probiotic bacteria including Lactobacilli, Bifidobacteria and Streptococcus species) heals acetic acid induced gastric ulcer in rats. VSL#3 was administered orally at low (6×109 bacteria) or high (1.2×1010 bacteria) dosages from day 3 after ulcer induction for 14 consecutive days. VSL#3 treatments significantly enhanced gastric ulcer healing in a dose-dependent manner. To assess the mechanism(s) whereby VSL#3 exerted its protective effects, we quantified the gene expression of several pro-inflammatory cytokines, protein and expression of stomach mucin-Muc5ac, regulatory cytokine-IL-10, COX-2 and various growth factors. Of all the components examined, only expression and protein production of VEGF was increased 332-fold on day 7 in the ulcerated tissues of animals treated with VSL#3. Predictably, animals treated with VEGF neutralizing antibody significantly delayed gastric ulcer healing in VSL#3 treated animals. This is the first report to demonstrate high efficacy of the probiotic mixture VSL#3 in enhancing gastric ulcer healing. Probiotic efficacy was effective at higher concentrations of VSL#3 by specifically increasing the expression and production of angiogenesis promoting growth factors, primarily VEGF. PMID:23484048

Enhanced second-harmonic-generation detection of collagen by means of optical wavefront shaping

NASA Astrophysics Data System (ADS)

Thompson, Jonathan V.; Throckmorton, Graham A.; Hokr, Brett H.; Yakovlev, Vladislav V.

2016-03-01

Second-harmonic generation (SHG) has proven to be an effective method to both image and detect structural variations in fibrillar collagen. The ability to detect these differences is especially useful in studying diseases like cancer and fibrosis.1 SHG techniques have historically been limited by their ability to penetrate and image through strongly scattering tissues. Recently, optical wavefront shaping has enabled light to be focused through highly scattering media such as biological tissue.2-4 This technology also enables us to examine the dependence of second harmonic generation on the spatial phase of the pump laser. Here, we demonstrate that wavefront shaping can be used to enhance the generation of second harmonic light from collagen fibrils even when scattering is low or non-existent.

A Cyclin Dependent Kinase Regulatory Subunit (CKS) Gene of Pigeonpea Imparts Abiotic Stress Tolerance and Regulates Plant Growth and Development in Arabidopsis

PubMed Central

Tamirisa, Srinath; Vudem, Dashavantha R.; Khareedu, Venkateswara R.

2017-01-01

Frequent climatic changes in conjunction with other extreme environmental factors are known to affect growth, development and productivity of diverse crop plants. Pigeonpea, a major grain legume of the semiarid tropics, endowed with an excellent deep-root system, is known as one of the important drought tolerant crop plants. Cyclin dependent kinases (CDKs) are core cell cycle regulators and play important role in different aspects of plant growth and development. The cyclin-dependent kinase regulatory subunit gene (CKS) was isolated from the cDNA library of pigeonpea plants subjected to drought stress. Pigeonpea CKS (CcCKS) gene expression was detected in both the root and leaf tissues of pigeonpea and was upregulated by polyethylene glycol (PEG), mannitol, NaCl and abscisic acid (ABA) treatments. The overexpression of CcCKS gene in Arabidopsis significantly enhanced tolerance of transgenics to drought and salt stresses as evidenced by different physiological parameters. Under stress conditions, transgenics showed higher biomass, decreased rate of water loss, decreased MDA levels, higher free proline contents, and glutathione levels. Moreover, under stress conditions transgenics exhibited lower stomatal conductance, lower transpiration, and higher photosynthetic rates. However, under normal conditions, CcCKS-transgenics displayed decreased plant growth rate, increased cell size and decreased stomatal number compared to those of wild-type plants. Real-time polymerase chain reaction revealed that CcCKS could regulate the expression of both ABA-dependent and ABA-independent genes associated with abiotic stress tolerance as well as plant growth and development. As such, the CcCKS seems promising and might serve as a potential candidate gene for enhancing the abiotic stress tolerance of crop plants. PMID:28239388

A Cyclin Dependent Kinase Regulatory Subunit (CKS) Gene of Pigeonpea Imparts Abiotic Stress Tolerance and Regulates Plant Growth and Development in Arabidopsis.

PubMed

Tamirisa, Srinath; Vudem, Dashavantha R; Khareedu, Venkateswara R

2017-01-01

Frequent climatic changes in conjunction with other extreme environmental factors are known to affect growth, development and productivity of diverse crop plants. Pigeonpea, a major grain legume of the semiarid tropics, endowed with an excellent deep-root system, is known as one of the important drought tolerant crop plants. Cyclin dependent kinases (CDKs) are core cell cycle regulators and play important role in different aspects of plant growth and development. The cyclin-dependent kinase regulatory subunit gene ( CKS ) was isolated from the cDNA library of pigeonpea plants subjected to drought stress. Pigeonpea CKS ( CcCKS ) gene expression was detected in both the root and leaf tissues of pigeonpea and was upregulated by polyethylene glycol (PEG), mannitol, NaCl and abscisic acid (ABA) treatments. The overexpression of CcCKS gene in Arabidopsis significantly enhanced tolerance of transgenics to drought and salt stresses as evidenced by different physiological parameters. Under stress conditions, transgenics showed higher biomass, decreased rate of water loss, decreased MDA levels, higher free proline contents, and glutathione levels. Moreover, under stress conditions transgenics exhibited lower stomatal conductance, lower transpiration, and higher photosynthetic rates. However, under normal conditions, CcCKS -transgenics displayed decreased plant growth rate, increased cell size and decreased stomatal number compared to those of wild-type plants. Real-time polymerase chain reaction revealed that Cc CKS could regulate the expression of both ABA-dependent and ABA-independent genes associated with abiotic stress tolerance as well as plant growth and development. As such, the CcCKS seems promising and might serve as a potential candidate gene for enhancing the abiotic stress tolerance of crop plants.

Activated Tissue-Resident Mesenchymal Stromal Cells Regulate Natural Killer Cell Immune and Tissue-Regenerative Function.

PubMed

Petri, Robert Michael; Hackel, Alexander; Hahnel, Katrin; Dumitru, Claudia Alexandra; Bruderek, Kirsten; Flohe, Stefanie B; Paschen, Annette; Lang, Stephan; Brandau, Sven

2017-09-12

The interaction of mesenchymal stromal cells (MSCs) with natural killer (NK) cells is traditionally thought of as a static inhibitory model, whereby resting MSCs inhibit NK cell effector function. Here, we use a dynamic in vitro system of poly(I:C) stimulation to model the interaction of NK cells and tissue-resident MSCs in the context of infection or tissue injury. The experiments suggest a time-dependent system of regulation and feedback, where, at early time points, activated MSCs secrete type I interferon to enhance NK cell effector function, while at later time points TGF-β and IL-6 limit NK cell effector function and terminate inflammatory responses by induction of a regulatory senescent-like NK cell phenotype. Importantly, feedback of these regulatory NK cells to MSCs promotes survival, proliferation, and pro-angiogenic properties. Our data provide additional insight into the interaction of stromal cells and innate immune cells and suggest a model of time-dependent MSC polarization and licensing. Copyright © 2017 The Authors. Published by Elsevier Inc. All rights reserved.

Lineage-specific enhancers activate self-renewal genes in macrophages and embryonic stem cells

PubMed Central

Soucie, Erinn L.; Weng, Ziming; Geirsdóttir, Laufey; Molawi, Kaaweh; Maurizio, Julien; Fenouil, Romain; Mossadegh-Keller, Noushine; Gimenez, Gregory; VanHille, Laurent; Beniazza, Meryam; Favret, Jeremy; Berruyer, Carole; Perrin, Pierre; Hacohen, Nir; Andrau, J.-C.; Ferrier, Pierre; Dubreuil, Patrice; Sidow, Arend; Sieweke, Michael H.

2016-01-01

Differentiated macrophages can self-renew in tissues and expand long-term in culture, but the gene regulatory mechanisms that accomplish self-renewal in the differentiated state have remained unknown. Here we show that in mice, the transcription factors MafB and c-Maf repress a macrophage-specific enhancer repertoire associated with a gene network controlling self-renewal. Single cell analysis revealed that, in vivo, proliferating resident macrophages can access this network by transient down-regulation of Maf transcription factors. The network also controls embryonic stem cell self-renewal but is associated with distinct embryonic stem cell-specific enhancers. This indicates that distinct lineage-specific enhancer platforms regulate a shared network of genes that control self-renewal potential in both stem and mature cells. PMID:26797145

Connective tissue fibroblasts and Tcf4 regulate myogenesis

PubMed Central

Mathew, Sam J.; Hansen, Jody M.; Merrell, Allyson J.; Murphy, Malea M.; Lawson, Jennifer A.; Hutcheson, David A.; Hansen, Mark S.; Angus-Hill, Melinda; Kardon, Gabrielle

2011-01-01

Muscle and its connective tissue are intimately linked in the embryo and in the adult, suggesting that interactions between these tissues are crucial for their development. However, the study of muscle connective tissue has been hindered by the lack of molecular markers and genetic reagents to label connective tissue fibroblasts. Here, we show that the transcription factor Tcf4 (transcription factor 7-like 2; Tcf7l2) is strongly expressed in connective tissue fibroblasts and that Tcf4GFPCre mice allow genetic manipulation of these fibroblasts. Using this new reagent, we find that connective tissue fibroblasts critically regulate two aspects of myogenesis: muscle fiber type development and maturation. Fibroblasts promote (via Tcf4-dependent signals) slow myogenesis by stimulating the expression of slow myosin heavy chain. Also, fibroblasts promote the switch from fetal to adult muscle by repressing (via Tcf4-dependent signals) the expression of developmental embryonic myosin and promoting (via a Tcf4-independent mechanism) the formation of large multinucleate myofibers. In addition, our analysis of Tcf4 function unexpectedly reveals a novel mechanism of intrinsic regulation of muscle fiber type development. Unlike other intrinsic regulators of fiber type, low levels of Tcf4 in myogenic cells promote both slow and fast myogenesis, thereby promoting overall maturation of muscle fiber type. Thus, we have identified novel extrinsic and intrinsic mechanisms regulating myogenesis. Most significantly, our data demonstrate for the first time that connective tissue is important not only for adult muscle structure and function, but is a vital component of the niche within which muscle progenitors reside and is a critical regulator of myogenesis. PMID:21177349

Enhanced Recovery Pathway in Microvascular Autologous Tissue-Based Breast Reconstruction: Should It Become the Standard of Care?

PubMed

Kaoutzanis, Christodoulos; Ganesh Kumar, Nishant; O'Neill, Dillon; Wormer, Blair; Winocour, Julian; Layliev, John; McEvoy, Matthew; King, Adam; Braun, Stephane A; Higdon, K Kye

2018-04-01

Enhanced recovery pathway programs have demonstrated improved perioperative care and shorter length of hospital stay in several surgical disciplines. The purpose of this study was to compare outcomes of patients undergoing autologous tissue-based breast reconstruction before and after the implementation of an enhanced recovery pathway program. The authors retrospectively reviewed consecutive patients who underwent autologous tissue-based breast reconstruction performed by two surgeons before and after the implementation of the enhanced recovery pathway at a university center over a 3-year period. Patient demographics, perioperative data, and 45-day postoperative outcomes were compared between the traditional standard of care (pre-enhanced recovery pathway) and enhanced recovery pathway patients. Multivariate logistic regression was performed to identify risk factors for length of hospital stay. Cost analysis was performed. Between April of 2014 and January of 2017, 100 consecutive women were identified, with 50 women in each group. Both groups had similar demographics, comorbidities, and reconstruction types. Postoperatively, the enhanced recovery pathway cohort used significantly less opiate and more acetaminophen compared with the traditional standard of care cohort. Median length of stay was shorter in the enhanced recovery pathway cohort, which resulted in an extrapolated $279,258 savings from freeing up inpatient beds and increase in overall contribution margins of $189,342. Participation in an enhanced recovery pathway program and lower total morphine-equivalent use were independent predictors for decreased length of hospital stay. Overall 45-day major complication rates, partial flap loss rates, emergency room visits, hospital readmissions, and unplanned reoperations were similar between the two groups. Enhanced recovery pathway program implementation should be considered as the standard approach for perioperative care in autologous tissue-based breast reconstruction because it does not affect morbidity and is associated with accelerated recovery with reduced postoperative opiate use and decreased length of hospital stay, leading to downstream health care cost savings. Therapeutic, III.

Multispectral image enhancement for H&E stained pathological tissue specimens

NASA Astrophysics Data System (ADS)

Bautista, Pinky A.; Abe, Tokiya; Yamaguchi, Masahiro; Ohyama, Nagaaki; Yagi, Yukako

2008-03-01

The presence of a liver disease such as cirrhosis can be determined by examining the proliferation of collagen fiber from a tissue slide stained with special stain such as the Masson's trichrome(MT) stain. Collagen fiber and smooth muscle, which are both stained the same in an H&E stained slide, are stained blue and pink respectively in an MT-stained slide. In this paper we show that with multispectral imaging the difference between collagen fiber and smooth muscle can be visualized even from an H&E stained image. In the method M KL bases are derived using the spectral data of those H&E stained tissue components which can be easily differentiated from each other, i.e. nucleus, cytoplasm, red blood cells, etc. and based on the spectral residual error of fiber weighting factors are determined to enhance spectral features at certain wavelengths. Results of our experiment demonstrate the capability of multispectral imaging and its advantage compared to the conventional RGB imaging systems to delineate tissue structures with subtle colorimetric difference.

Survival of residual neutrophils and accelerated myelopoiesis limit the efficacy of antibody-mediated depletion of Ly-6G+ cells in tumor-bearing mice.

PubMed

Moses, Katrin; Klein, Johanna C; Männ, Linda; Klingberg, Anika; Gunzer, Matthias; Brandau, Sven

2016-06-01

Expansion of Ly-6G(+) myeloid cells has been reported in most murine cancer models. However, divergent findings exist regarding the role and effect of these cells on host immunity and tumor progression. Antibody-mediated depletion of Ly-6G(+) cells is a common technique to assess the in vivo relevance of these cells. Interpretation of results crucially depends on the efficacy and course of depletion. We established murine head and neck cancer models and analyzed the efficacy of antibody-mediated depletion by flow cytometry, conventional histology, and intravital imaging with a novel Ly-6G-transgenic mouse model. The first phase of depletion was characterized by effective elimination of Ly-6G(+) cells from the peripheral blood. Nevertheless, viable, resistant cells were found to reside in the tumor tissue and spleen. This peripheral depletion phase was associated with high systemic levels of granulocyte colony-stimulating factor and KC and enhanced splenic production of Ly-6G(+) cells. Even under sustained treatment with either αGr-1 or αLy-6G antibodies, peripheral blood depletion ended after approximately 1 wk and was followed by reappearance of immature Ly-6G(+) cells with an immunoregulatory phenotype. Reappearance of these depletion-resistant immature cells was enhanced in tumor-bearing, compared with naïve, control mice. Collectively, our data suggest that depletion of Ly-6G(+) myeloid cells in tumor-bearing mice is counteracted by the persistence of intratumoral cells, enhanced extramedullary granulopoiesis, and accelerated reappearance of immature cells. Hence, extensive monitoring of in vivo kinetics and tissue distribution of Ly-6G(+) cells is required in depletion studies. © Society for Leukocyte Biology.

Effects of pergolide mesylate on transduction efficiency of PEP-1-catalase protein

DOE Office of Scientific and Technical Information (OSTI.GOV)

Sohn, Eun Jeong; Kim, Dae Won; Kim, Young Nam

2011-03-18

Research highlights: {yields} We studied effects of pergolide mesylate (PM) on in vitro and in vivo transduction of PEP-1-catalase. {yields} PEP-1-catatase inhibited 12-O-tetradecanoylphorbol-13-acetate (TPA)-induced inflammation. {yields} PM enhanced the transduction of PEP-1-catalase into HaCaT cells and skin tissue. {yields} PM increased anti-inflammatory activity of PEP-1-catalase. {yields} PM stimulated therapeutic action of anti-oxidant enzyme catalase in oxidative-related diseases. -- Abstract: The low transduction efficiency of various proteins is an obstacle to their therapeutic application. However, protein transduction domains (PTDs) are well-known for a highly effective tool for exogenous protein delivery to cells. We examined the effects of pergolide mesylate (PM) onmore » the transduction of PEP-1-catalase into HaCaT human keratinocytes and mice skin and on the anti-inflammatory activity of PEP-1-catatase against 12-O-tetradecanoylphorbol-13-acetate (TPA)-induced inflammation using Western blot and histological analysis. PM enhanced the time- and dose-dependent transduction of PEP-1-catalase into HaCaT cells without affecting the cellular toxicity. In a mouse edema model, PEP-1-catalase inhibited the increased expressions of inflammatory mediators and cytokines such as cyclooxygenase-2, inducible nitric oxide synthase, interleukin-6 and -1{beta}, and tumor necrosis factor-{alpha} induced by TPA. On the other hand, PM alone failed to exert any significant anti-inflammatory effects. However, the anti-inflammatory effect of co-treatment with PEP-1-catalase and PM was more potent than that of PEP-1-catalase alone. Our results indicate that PM may enhance the delivery of PTDs fusion therapeutic proteins to target cells and tissues and has potential to increase their therapeutic effects of such drugs against various diseases.« less

Alignment of cellular motility forces with tissue flow as a mechanism for efficient wound healing

PubMed Central

Basan, Markus; Elgeti, Jens; Hannezo, Edouard; Rappel, Wouter-Jan; Levine, Herbert

2013-01-01

Recent experiments have shown that spreading epithelial sheets exhibit a long-range coordination of motility forces that leads to a buildup of tension in the tissue, which may enhance cell division and the speed of wound healing. Furthermore, the edges of these epithelial sheets commonly show finger-like protrusions whereas the bulk often displays spontaneous swirls of motile cells. To explain these experimental observations, we propose a simple flocking-type mechanism, in which cells tend to align their motility forces with their velocity. Implementing this idea in a mechanical tissue simulation, the proposed model gives rise to efficient spreading and can explain the experimentally observed long-range alignment of motility forces in highly disordered patterns, as well as the buildup of tensile stress throughout the tissue. Our model also qualitatively reproduces the dependence of swirl size and swirl velocity on cell density reported in experiments and exhibits an undulation instability at the edge of the spreading tissue commonly observed in vivo. Finally, we study the dependence of colony spreading speed on important physical and biological parameters and derive simple scaling relations that show that coordination of motility forces leads to an improvement of the wound healing process for realistic tissue parameters. PMID:23345440

An Ultraconserved Brain-specific Enhancer within ADGRL3 (LPHN3) Underpins ADHD Susceptibility

PubMed Central

Martinez, Ariel F.; Abe, Yu; Hong, Sungkook; Molyneux, Kevin; Yarnell, David; Löhr, Heiko; Driever, Wolfgang; Acosta, Maria T.; Arcos-Burgos, Mauricio; Muenke, Maximilian

2016-01-01

BACKGROUND Genetic factors predispose to attention deficit/hyperactivity disorder (ADHD). Previous studies have reported linkage and association to ADHD of gene variants within ADGRL3. In this study, we functionally analyzed non-coding variants in this gene as likely pathological contributors. METHODS In silico, in vitro and in vivo approaches were used to identify and characterize evolutionary conserved elements within the ADGRL3 linkage region (~207 Kb). Family-based genetic analyses on 838 individuals (372 affected and 466 unaffected) identified ADHD-associated SNPs harbored in some of these conserved elements. Luciferase assays and zebrafish GFP transgenesis tested conserved elements for transcriptional enhancer activity. Electromobility shift assays were used to verify transcription factor binding disruption by ADHD risk alleles. RESULTS An ultraconserved element was discovered (ECR47) that functions as a transcriptional enhancer. A three-variant ADHD risk haplotype in ECR47, formed by rs17226398, rs56038622 and rs2271338, reduced enhancer activity by 40% in neuroblastoma and astrocytoma cells (PBonferroni<0.0001). This enhancer also drove GFP expression in the zebrafish brain in a tissue-specific manner, sharing aspects of endogenous ADGRL3 expression. The rs2271338 risk allele disrupts binding of YY1, an important factor in the development and function of the central nervous system. Expression quantitative trait loci analysis of post-mortem human brain tissues revealed an association between rs2271338 and reduced ADGRL3 expression in the thalamus. CONCLUSIONS These results uncover the first functional evidence of common non-coding variants with potential implications for the pathology of ADHD. PMID:27692237

Macrophages: development and tissue specialization.

PubMed

Varol, Chen; Mildner, Alexander; Jung, Steffen

2015-01-01

Macrophages are myeloid immune cells that are strategically positioned throughout the body tissues, where they ingest and degrade dead cells, debris, and foreign material and orchestrate inflammatory processes. Here we review two major recent paradigm shifts in our understanding of tissue macrophage biology. The first is the realization that most tissue-resident macrophages are established prenatally and maintained through adulthood by longevity and self-renewal. Their generation and maintenance are thus independent from ongoing hematopoiesis, although the cells can be complemented by adult monocyte-derived macrophages. Second, aside from being immune sentinels, tissue macrophages form integral components of their host tissue. This entails their specialization in response to local environmental cues to contribute to the development and specific function of their tissue of residence. Factors that govern tissue macrophage specialization are emerging. Moreover, tissue specialization is reflected in discrete gene expression profiles of macrophages, as well as epigenetic signatures reporting actual and potential enhancer usage.

Is there any difference in Amide and NOE CEST effects between white and gray matter at 7 T?

NASA Astrophysics Data System (ADS)

Khlebnikov, Vitaliy; Siero, Jeroen C. W.; Wijnen, Jannie; Visser, Fredy; Luijten, Peter R.; Klomp, Dennis W. J.; Hoogduin, Hans

2016-11-01

Measurement of Chemical Exchange Saturation Transfer (CEST) is providing tissue physiology dependent contrast, e.g. by looking at Amide and NOE (Nuclear Overhauser Enhancement) effects. CEST is unique in providing quantitative metabolite information at high imaging resolution. However, direct comparison of Amide and NOE effects between different tissues may result in wrong conclusions on the metabolite concentration due to the additional contributors to the observed CEST contrast, such as water content (WC) and water T1 relaxation (T1w). For instance, there are multiple contradictory reports in the literature on Amide and NOE effects in white matter (WM) and gray matter (GM) at 7 T. This study shows that at 7 T, tissue water T1 relaxation is a stronger contributor to CEST contrasts than WC. After water T1 correction, there was no difference in Amide effects between WM and GM, whereas WM/GM contrast was enhanced for NOE effects.

Above threshold spectral dependence of linewidth enhancement factor, optical duration and linear chirp of quantum dot lasers.

PubMed

Kim, Jimyung; Delfyett, Peter J

2009-12-07

The spectral dependence of the linewidth enhancement factor above threshold is experimentally observed from a quantum dot Fabry-Pérot semiconductor laser. The linewidth enhancement factor is found to be reduced when the quantum dot laser operates approximately 10 nm offset to either side of the gain peak. It becomes significantly reduced on the anti-Stokes side as compared to the Stokes side. It is also found that the temporal duration of the optical pulses generated from quantum dot mode-locked lasers is shorter when the laser operates away from the gain peak. In addition, less linear chirp is impressed on the pulse train generated from the anti-Stokes side whereas the pulses generated from the gain peak and Stokes side possess a large linear chirp. These experimental results imply that enhanced performance characteristics of quantum dot lasers can be achieved by operating on the anti-Stokes side, approximately 10 nm away from the gain peak.

Estimation of pairwise sequence similarity of mammalian enhancers with word neighbourhood counts.

PubMed

Göke, Jonathan; Schulz, Marcel H; Lasserre, Julia; Vingron, Martin

2012-03-01

The identity of cells and tissues is to a large degree governed by transcriptional regulation. A major part is accomplished by the combinatorial binding of transcription factors at regulatory sequences, such as enhancers. Even though binding of transcription factors is sequence-specific, estimating the sequence similarity of two functionally similar enhancers is very difficult. However, a similarity measure for regulatory sequences is crucial to detect and understand functional similarities between two enhancers and will facilitate large-scale analyses like clustering, prediction and classification of genome-wide datasets. We present the standardized alignment-free sequence similarity measure N2, a flexible framework that is defined for word neighbourhoods. We explore the usefulness of adding reverse complement words as well as words including mismatches into the neighbourhood. On simulated enhancer sequences as well as functional enhancers in mouse development, N2 is shown to outperform previous alignment-free measures. N2 is flexible, faster than competing methods and less susceptible to single sequence noise and the occurrence of repetitive sequences. Experiments on the mouse enhancers reveal that enhancers active in different tissues can be separated by pairwise comparison using N2. N2 represents an improvement over previous alignment-free similarity measures without compromising speed, which makes it a good candidate for large-scale sequence comparison of regulatory sequences. The software is part of the open-source C++ library SeqAn (www.seqan.de) and a compiled version can be downloaded at http://www.seqan.de/projects/alf.html. Supplementary data are available at Bioinformatics online.

AGED DOMINANT NEGATIVE p38α MAPK MICE ARE RESISTANT TO AGE-DEPENDENT DECLINE IN ADULT-NEUROGENESIS AND CONTEXT DISCRIMINATION FEAR CONDITIONING

PubMed Central

Cortez, IbDanelo; Bulavin, Dmitry V.; Wu, Ping; McGrath, Erica L; Cunningham, Kathryn A; Wakamiya, Maki; Papaconstantinou, John; Dineley, Kelly T

2018-01-01

A major aspect of mammalian aging is the decline in functional competence of many self-renewing cell types, including adult-born neuronal precursors. Since age-related senescence of self-renewal occurs simultaneously with chronic up-regulation of the p38MAPKalpha (p38α) signaling pathway, we used the dominant negative mouse model for attenuated p38α activity (DN-p38αAF/+ ) in which Thr180 and Tyr182 are mutated (T→A/Y→F) to prevent phosphorylation activation (DN-p38αAF/+) and kinase activity. As a result, aged DN-p38αAF/+ mice are resistant to age-dependent decline in proliferation and regeneration of several peripheral tissue progenitors when compared to wild-type littermates. Aging is the major risk factor for non-inherited forms of Alzheimer’s disease (AD); environmental and genetic risk factors that accelerate the senescence phenotype are thought to contribute to an individual’s relative risk. In the present study, we evaluated aged DN-p38αAF/+ and wildtype littermates in a series of behavioral paradigms to test if p38α mutant mice exhibit altered baseline abnormalities in neurological reflexes, locomotion, anxiety-like behavior, and age-dependent cognitive decline. While aged DN-p38αAF/+ and wildtype littermates appear equal in all tested baseline neurological and behavioral parameters, DN-p38αAF/+ exhibit superior context discrimination fear conditioning. Context discrimination is a cognitive task that is supported by proliferation and differentiation of adult-born neurons in the dentate gyrus of the hippocampus. Consistent with enhanced context discrimination in aged DN-p38αAF/+, we discovered enhanced production of adult-born neurons in the dentate gyrus of DN-p38αAF/+ mice compared to wildtype littermates. Our findings support the notion that p38α inhibition has therapeutic utility in aging diseases that affect cognition, such as AD. PMID:27765672

Aged dominant negative p38α MAPK mice are resistant to age-dependent decline in adult-neurogenesis and context discrimination fear conditioning.

PubMed

Cortez, IbDanelo; Bulavin, Dmitry V; Wu, Ping; McGrath, Erica L; Cunningham, Kathryn A; Wakamiya, Maki; Papaconstantinou, John; Dineley, Kelly T

2017-03-30

A major aspect of mammalian aging is the decline in functional competence of many self-renewing cell types, including adult-born neuronal precursors. Since age-related senescence of self-renewal occurs simultaneously with chronic up-regulation of the p38MAPKalpha (p38α) signaling pathway, we used the dominant negative mouse model for attenuated p38α activity (DN-p38α AF/+ ) in which Thr180 and Tyr182 are mutated (T→A/Y→F) to prevent phosphorylation activation (DN-p38α AF/+ ) and kinase activity. As a result, aged DN-p38α AF/+ mice are resistant to age-dependent decline in proliferation and regeneration of several peripheral tissue progenitors when compared to wild-type littermates. Aging is the major risk factor for non-inherited forms of Alzheimer's disease (AD); environmental and genetic risk factors that accelerate the senescence phenotype are thought to contribute to an individual's relative risk. In the present study, we evaluated aged DN-p38α AF/+ and wildtype littermates in a series of behavioral paradigms to test if p38α mutant mice exhibit altered baseline abnormalities in neurological reflexes, locomotion, anxiety-like behavior, and age-dependent cognitive decline. While aged DN-p38α AF/+ and wildtype littermates appear equal in all tested baseline neurological and behavioral parameters, DN-p38α AF/+ exhibit superior context discrimination fear conditioning. Context discrimination is a cognitive task that is supported by proliferation and differentiation of adult-born neurons in the dentate gyrus of the hippocampus. Consistent with enhanced context discrimination in aged DN-p38α AF/+ , we discovered enhanced production of adult-born neurons in the dentate gyrus of DN-p38α AF/+ mice compared to wildtype littermates. Our findings support the notion that p38α inhibition has therapeutic utility in aging diseases that affect cognition, such as AD. Copyright © 2016 Elsevier B.V. All rights reserved.

A Polarization-Dependent Normal Incident Quantum Cascade Detector Enhanced Via Metamaterial Resonators.

PubMed

Wang, Lei; Zhai, Shen-Qiang; Wang, Feng-Jiao; Liu, Jun-Qi; Liu, Shu-Man; Zhuo, Ning; Zhang, Chuan-Jin; Wang, Li-Jun; Liu, Feng-Qi; Wang, Zhan-Guo

2016-12-01

The design, fabrication, and characterization of a polarization-dependent normal incident quantum cascade detector coupled via complementary split-ring metamaterial resonators in the infrared regime are presented. The metamaterial structure is designed through three-dimensional finite-difference time-domain method and fabricated on the top metal contact, which forms a double-metal waveguide together with the metallic ground plane. With normal incidence, significant enhancements of photocurrent response are obtained at the metamaterial resonances compared with the 45° polished edge coupling device. The photocurrent response enhancements exhibit clearly polarization dependence, and the largest response enhancement factor of 165% is gained for the incident light polarized parallel to the split-ring gap.

RIP1 regulates TNF-α-mediated lymphangiogenesis and lymphatic metastasis in gallbladder cancer by modulating the NF-κB-VEGF-C pathway

PubMed Central

Lin, Bin; Hong, Hai-Jie; Zhu, Si-Yuan; Jiang, Lei; Wang, Xiao-Qian; Tang, Nan-Hong; She, Fei-Fei; Chen, Yan-Ling

2018-01-01

Background Tumor necrosis factor alpha (TNF-α) enhances lymphangiogenesis in gallbladder carcinoma (GBC) via activation of nuclear factor (NF-κB)-dependent vascular endothelial growth factor-C (VEGF-C). Receptor-interacting protein 1 (RIP1) is a multifunctional protein in the TNF-α signaling pathway and is highly expressed in GBC. However, whether RIP1 participates in the signaling pathway of TNF-α-mediated VEGF-C expression that enhances lymphangiogenesis in GBC remains unclear. Methods The RIP1 protein levels in the GBC-SD and NOZ cells upon stimulation with increasing concentrations of TNF-α as indicated was examined using Western blot. Lentiviral RIP1 shRNA and siIκBα were constructed and transduced respectively them into NOZ and GBC-SD cells, and then PcDNA3.1-RIP1 vectors was transduced into siRIP1 cell lines to reverse RIP1 expression. The protein expression of RIP1, inhibitor of NF-κB alpha (IκBα), p-IκBα, TAK1, NF-κB essential modulator were examined through immunoblotting or immunoprecipitation. Moreover, VEGF-C mRNA levels were measured by quantitative real-time polymerase chain reaction, VEGF-C protein levels were measured by immunoblotting and enzyme-linked immunosorbent assay, and VEGF-C promoter and NF-κB activities were quantified using a dual luciferase reporter assay. The association of NF-κB with the VEGF-C promoter was analysed by chromatin immunoprecipitation assay. A three-dimensional coculture method and orthotopic transplantation nude mice model were used to evaluate lymphatic tube-forming and metastasis ability in GBC cells. The expression of RIP1 protein, TNF-α protein and lymphatic vessels in human GBC tissues was examined by immunohistochemistry, and the dependence between RIP1 protein with TNF-α protein and lymphatic vessel density was analysed. Results TNF-α dose- and time-dependently increased RIP1 protein expression in the GBC-SD and NOZ cells of GBC, and the strongest effect was observed with a concentration of 50 ng/ml. RIP1 is fundamental for TNF-α-mediated NF-κB activation in GBC cells and can regulate TNF-α-mediated VEGF-C expression at the protein and transcriptional levels through the NF-κB pathway. RIP1 can regulate TNF-α-mediated lymphatic tube formation and metastasis in GBC cells both in vitro and vivo. The average optical density of RIP1 was linearly related to that of TNF-α protein and the lymphatic vessel density in GBC tissues. Conclusion We conclude that RIP1 regulates TNF-α-mediated lymphangiogenesis and lymph node metastasis in GBC by modulating the NF-κB-VEGF-C pathway. PMID:29844685

RIP1 regulates TNF-α-mediated lymphangiogenesis and lymphatic metastasis in gallbladder cancer by modulating the NF-κB-VEGF-C pathway.

PubMed

Li, Cheng-Zong; Jiang, Xiao-Jie; Lin, Bin; Hong, Hai-Jie; Zhu, Si-Yuan; Jiang, Lei; Wang, Xiao-Qian; Tang, Nan-Hong; She, Fei-Fei; Chen, Yan-Ling

2018-01-01

Tumor necrosis factor alpha (TNF-α) enhances lymphangiogenesis in gallbladder carcinoma (GBC) via activation of nuclear factor (NF-κB)-dependent vascular endothelial growth factor-C (VEGF-C). Receptor-interacting protein 1 (RIP1) is a multifunctional protein in the TNF-α signaling pathway and is highly expressed in GBC. However, whether RIP1 participates in the signaling pathway of TNF-α-mediated VEGF-C expression that enhances lymphangiogenesis in GBC remains unclear. The RIP1 protein levels in the GBC-SD and NOZ cells upon stimulation with increasing concentrations of TNF-α as indicated was examined using Western blot. Lentiviral RIP1 shRNA and siIκBα were constructed and transduced respectively them into NOZ and GBC-SD cells, and then PcDNA3.1-RIP1 vectors was transduced into siRIP1 cell lines to reverse RIP1 expression. The protein expression of RIP1, inhibitor of NF-κB alpha (IκBα), p-IκBα, TAK1, NF-κB essential modulator were examined through immunoblotting or immunoprecipitation. Moreover, VEGF-C mRNA levels were measured by quantitative real-time polymerase chain reaction, VEGF-C protein levels were measured by immunoblotting and enzyme-linked immunosorbent assay, and VEGF-C promoter and NF-κB activities were quantified using a dual luciferase reporter assay. The association of NF-κB with the VEGF-C promoter was analysed by chromatin immunoprecipitation assay. A three-dimensional coculture method and orthotopic transplantation nude mice model were used to evaluate lymphatic tube-forming and metastasis ability in GBC cells. The expression of RIP1 protein, TNF-α protein and lymphatic vessels in human GBC tissues was examined by immunohistochemistry, and the dependence between RIP1 protein with TNF-α protein and lymphatic vessel density was analysed. TNF-α dose- and time-dependently increased RIP1 protein expression in the GBC-SD and NOZ cells of GBC, and the strongest effect was observed with a concentration of 50 ng/ml. RIP1 is fundamental for TNF-α-mediated NF-κB activation in GBC cells and can regulate TNF-α-mediated VEGF-C expression at the protein and transcriptional levels through the NF-κB pathway. RIP1 can regulate TNF-α-mediated lymphatic tube formation and metastasis in GBC cells both in vitro and vivo. The average optical density of RIP1 was linearly related to that of TNF-α protein and the lymphatic vessel density in GBC tissues. We conclude that RIP1 regulates TNF-α-mediated lymphangiogenesis and lymph node metastasis in GBC by modulating the NF-κB-VEGF-C pathway.

Influence of the environmental factors on the intensity of the oxygen, ammonium, and phosphate metabolism in the agar-containing seaweed Ahnfeltia tobuchiensis (Ahnfeltiales, Rhodophyta)

NASA Astrophysics Data System (ADS)

Cherbadgy, I. I.; Sabitova, L. I.

2011-02-01

A complex study of the influence of various environmental factors on the rate of the oxygen (MO 2), ammonium (MNH 4), and phosphate (MPO 4) metabolism in Ahnfeltia tobuchiensis has been carried out in situ in the Izmena Bay of Kunashir Island. The following environmental factors have been included into the investigation: the photosynthetically active radiation (PAR); the ammonium (NH4); the phosphate (PO4); and the tissue content of carbon (C), nitrogen (N), phosphorus (P), and chlorophyll a (Chl). The population of agar-containing seaweed A. tobuchiensis forms a layer with a thickness up to 0.5 m, which occupies about 23.3 km2; the population's biomass is equal to 125000 tons. The quantitative assessment of the organic matter production and nutrient consumption during the oxygen metabolism (MO 2) has been carried out for the whole population. It has been shown that the daily rate depends on the PAR intensity, the seawater concentrations of PO4 and NH4, and the tissue content of N and P ( r 2 = 0.78, p < 0.001). The daily NH4 consumption averages 0.21 μmol/(gDW h) and depends on the NH4 and O2 concentrations in the seawater and on the C and Chl a content in the algal tissues ( r 2 = 0.64, p < 0.001). The daily PO4 consumption averages 0.01 μmol/(gDW h) and depends on the NH4 concentration in the seawater and on the P content in the algal tissues ( r 2 = 0.40, p < 0.001).

Characteristics of the somatotropic axis in insulin dependent diabetes mellitus.

PubMed

Mercado, M; Baumann, G

1995-01-01

Growth hormone (GH) plays an important role in glucose homeostasis in both healthy subjects and patients with diabetes. Patients with poorly controlled insulin-dependent diabetes mellitus (IDDM) have high basal and integrated serum GH concentrations, as well as an enhanced GH response to several secretagogues. Yet, these patients have impaired generation of insulin-like growth factor-I (IGF-I). These abnormalities tend to return to normal as an adequate metabolic control is achieved. In view of this hormonal profile, IDDM has been considered a state of relative GH resistance. Studies in experimental animals with streptozotocin-induced diabetes have shown a decreased binding of radiolabeled GH to liver membranes. More recently, adults and children with IDDM have been found to have low levels of the high affinity growth hormone binding protein (GHBP), which represents the extracellular portion of the GH receptor, and is thought to reflect GH receptor tissue concentrations. The abnormalities in the GH/IGF-I axis have been implicated in the worsening of metabolic control that occurs in some patients, as well as in the development of microvascular complications, particularly retinopathy.

Exosomes of human placenta-derived mesenchymal stem cells stimulate angiogenesis.

PubMed

Komaki, Motohiro; Numata, Yuri; Morioka, Chikako; Honda, Izumi; Tooi, Masayuki; Yokoyama, Naoki; Ayame, Hirohito; Iwasaki, Kengo; Taki, Atsuko; Oshima, Noriko; Morita, Ikuo

2017-10-03

The therapeutic potential of mesenchymal stem cells (MSCs) may be attributed partly to humoral factors such as growth factors, cytokines, and chemokines. Human term placental tissue-derived MSCs (PlaMSCs), or conditioned medium left over from cultures of these cells, have been reported to enhance angiogenesis. Recently, the exosome, which can transport a diverse suite of macromolecules, has gained attention as a novel intercellular communication tool. However, the potential role of the exosome in PlaMSC therapeutic action is not well understood. The purpose of this study was to evaluate PlaMSC-derived exosome angiogenesis promotion in vitro and in vivo. MSCs were isolated from human term placental tissue by enzymatic digestion. Conditioned medium was collected after 48-h incubation in serum-free medium (PlaMSC-CM). Angiogenic factors present in PlaMSC-CM were screened by a growth factor array. Exosomes were prepared by ultracentrifugation of PlaMSC-CM, and confirmed by transmission electron microscopy, dynamic light scattering, and western blot analyses. The proangiogenic activity of PlaMSC-derived exosomes (PlaMSC-exo) was assessed using an endothelial tube formation assay, a cell migration assay, and reverse transcription-PCR analysis. The in-vivo angiogenic activity of PlaMSC-exo was evaluated using a murine auricle ischemic injury model. PlaMSC-CM contained both angiogenic and angiostatic factors, which enhanced endothelial tube formation. PlaMSC-exo were incorporated into endothelial cells; these exosomes stimulated both endothelial tube formation and migration, and enhanced angiogenesis-related gene expression. Laser Doppler blood flow analysis showed that PlaMSC-exo infusion also enhanced angiogenesis in an in-vivo murine auricle ischemic injury model. PlaMSC-exo enhanced angiogenesis in vitro and in vivo, suggesting that exosomes play a role in the proangiogenic activity of PlaMSCs. PlaMSC-exo may be a novel therapeutic approach for treating ischemic diseases.

Interleukin 6 downregulates p53 expression and activity by stimulating ribosome biogenesis: a new pathway connecting inflammation to cancer

PubMed Central

Brighenti, E; Calabrese, C; Liguori, G; Giannone, F A; Trerè, D; Montanaro, L; Derenzini, M

2014-01-01

Chronic inflammation is an established risk factor for the onset of cancer, and the inflammatory cytokine IL-6 has a role in tumorigenesis by enhancing proliferation and hindering apoptosis. As factors stimulating proliferation also downregulate p53 expression by enhancing ribosome biogenesis, we hypothesized that IL-6 may cause similar changes in inflamed tissues, thus activating a mechanism that favors neoplastic transformation. Here, we showed that IL-6 downregulated the expression and activity of p53 in transformed and untransformed human cell lines. This was the consequence of IL-6-dependent stimulation of c-MYC mRNA translation, which was responsible for the upregulation of rRNA transcription. The enhanced rRNA transcription stimulated the MDM2-mediated proteasomal degradation of p53, by reducing the availability of ribosome proteins for MDM2 binding. The p53 downregulation induced the acquisition of cellular phenotypic changes characteristic of epithelial–mesenchymal transition, such as a reduced level of E-cadherin expression, increased cell invasiveness and a decreased response to cytotoxic stresses. We found that these changes also occurred in colon epithelial cells of patients with ulcerative colitis, a very representative example of chronic inflammation at high risk for tumor development. Histochemical and immunohistochemical analysis of colon biopsy samples showed an upregulation of ribosome biogenesis, a reduced expression of p53, together with a focal reduction or absence of E-cadherin expression in chronic colitis in comparison with normal mucosa samples. These changes disappeared after treatment with anti-inflammatory drugs. Taken together, the present results highlight a new mechanism that may link chronic inflammation to cancer, based on p53 downregulation, which is activated by the enhancement of rRNA transcription upon IL-6 exposure. PMID:24531714

Histidine-rich stabilized polyplexes for cMet-directed tumor-targeted gene transfer

NASA Astrophysics Data System (ADS)

Kos, Petra; Lächelt, Ulrich; Herrmann, Annika; Mickler, Frauke Martina; Döblinger, Markus; He, Dongsheng; Krhač Levačić, Ana; Morys, Stephan; Bräuchle, Christoph; Wagner, Ernst

2015-03-01

Overexpression of the hepatocyte growth factor receptor/c-Met proto oncogene on the surface of a variety of tumor cells gives an opportunity to specifically target cancerous tissues. Herein, we report the first use of c-Met as receptor for non-viral tumor-targeted gene delivery. Sequence-defined oligomers comprising the c-Met binding peptide ligand cMBP2 for targeting, a monodisperse polyethylene glycol (PEG) for polyplex surface shielding, and various cationic (oligoethanamino) amide cores containing terminal cysteines for redox-sensitive polyplex stabilization, were assembled by solid-phase supported syntheses. The resulting oligomers exhibited a greatly enhanced cellular uptake and gene transfer over non-targeted control sequences, confirming the efficacy and target-specificity of the formed polyplexes. Implementation of endosomal escape-promoting histidines in the cationic core was required for gene expression without additional endosomolytic agent. The histidine-enriched polyplexes demonstrated stability in serum as well as receptor-specific gene transfer in vivo upon intratumoral injection. The co-formulation with an analogous PEG-free cationic oligomer led to a further compaction of pDNA polyplexes with an obvious change of shape as demonstrated by transmission electron microscopy. Such compaction was critically required for efficient intravenous gene delivery which resulted in greatly enhanced, cMBP2 ligand-dependent gene expression in the distant tumor.Overexpression of the hepatocyte growth factor receptor/c-Met proto oncogene on the surface of a variety of tumor cells gives an opportunity to specifically target cancerous tissues. Herein, we report the first use of c-Met as receptor for non-viral tumor-targeted gene delivery. Sequence-defined oligomers comprising the c-Met binding peptide ligand cMBP2 for targeting, a monodisperse polyethylene glycol (PEG) for polyplex surface shielding, and various cationic (oligoethanamino) amide cores containing terminal cysteines for redox-sensitive polyplex stabilization, were assembled by solid-phase supported syntheses. The resulting oligomers exhibited a greatly enhanced cellular uptake and gene transfer over non-targeted control sequences, confirming the efficacy and target-specificity of the formed polyplexes. Implementation of endosomal escape-promoting histidines in the cationic core was required for gene expression without additional endosomolytic agent. The histidine-enriched polyplexes demonstrated stability in serum as well as receptor-specific gene transfer in vivo upon intratumoral injection. The co-formulation with an analogous PEG-free cationic oligomer led to a further compaction of pDNA polyplexes with an obvious change of shape as demonstrated by transmission electron microscopy. Such compaction was critically required for efficient intravenous gene delivery which resulted in greatly enhanced, cMBP2 ligand-dependent gene expression in the distant tumor. Electronic supplementary information (ESI) available. See DOI: 10.1039/c4nr06556e

Exendin-4, a glucagon-like peptide-1 analogue accelerates healing of chronic gastric ulcer in diabetic rats

PubMed Central

Chen, Yen-Cheng; Ho, Ching-Chun; Yi, Chih-Hsun; Liu, Xiu-Zhu; Cheng, Tzu-Ting

2017-01-01

Background Diabetes mellitus is an independent risk factor for impaired healing of peptic ulcers, and there are currently no supplementary therapeutics other than the standard antipeptic medicine to improve the ulcer healing in diabetes. This study examined the potential pleiotropic effect of a glucagon-like peptide (Glp)-1 analogue exendin (Ex)-4 on the regeneration of gastric ulcer in streptozotocin-induced diabetic rats. Methods and results Chronic ulcer was created in rat stomach by submucosal injection of acetic acid and peri-ulcer tissues were analyzed 7 days after operation. Ulcer wound healing was impaired in diabetic rats with suppressed tissue expression of eNOS and enhanced levels of pro-inflammatory reactions. Treatment with intraperitoneal injection of Ex4 (0.5 μg/kg/d) significantly reduced the area of gastric ulcer without changing blood glucose level. Ex-4 restored the expression of pro-angiogenic factors, and attenuated the generation of regional inflammation and superoxide anions. The improvement of ulcer healing was associated with increased expression of MMP-2 and formation of granulation tissue in the peri-ulcer area. Conclusion Administration of Ex4 may induce pro-angiogenic, anti-inflammatory and anti-oxidative reactions in the peri-ulcer tissue of diabetic rats that eventually enhances tissue granulation and closure of ulcerative wounds. Our results support the potential clinical application of Glp-1 analogues as supplementary hypoglycemic agents in the antipeptic ulcer medication in diabetes. PMID:29095895

DOE Office of Scientific and Technical Information (OSTI.GOV)

Yang, Tao; Li, Xiao-Na; Li, Xing-Guang

Highlights: • DNAJC6 is up-regulated in hepatocellular carcinoma tissues. • DNAJC6 promotes hepatocellular carcinoma cell proliferation and invasion. • DNAJC6 induces epithelial–mesenchymal transition by activating transforming growth factor β signaling. - Abstract: Epithelial–mesenchymal transition (EMT) is a developmental program, which is associated with hepatocellular carcinoma (HCC) development and progression. DNAJC6 (DNA/HSP40 homolog subfamily C member 6) encodes auxilin, which is responsible for juvenile Parkinsonism with phenotypic variability. However, the role of DNAJC6 in HCC development and progression is limited. Here, we report that DNAJC6 is up-regulated in HCC tissues and up-regulation of DNAJC6 expression predicts poor outcome in patients withmore » HCC. Furthermore, overexpression of DNAJC6 enhances the ability for acquisition of mesenchymal traits, enhanced cell proliferation and invasion. DNAJC6 positively regulated expression of EMT-related transcription factor, also activating transforming growth factor β (TGF-β) pathway to contribute to EMT. Our findings demonstrated an important function of DNAJC6 in the progression of HCC by induction of EMT, and they implicate DNAJC6 as a marker of poor outcome in HCC.« less

Emulating Native Periosteum Cell Population and Subsequent Paracrine Factor Production To Promote Tissue Engineered Periosteum-Mediated Allograft Healing

PubMed Central

Hoffman, Michael D.

2015-01-01

Emulating autograft healing within the context of decellularized bone allografts has immediate clinical applications in the treatment of critical-sized bone defects. The periosteum, a thin, osteogenic tissue that surrounds bone, houses a heterogeneous population of stem cells and osteoprogenitors. There is evidence that periosteum-cell derived paracrine factors, specifically vascular endothelial growth factor (VEGF) and bone morphogenetic protein 2 (BMP2), orchestrate autograft healing through host cell recruitment and subsequent tissue elaboration. In previous work, we demonstrated that the use of poly(ethylene glycol) (PEG) hydrogels as a tissue engineered (T.E.) periosteum to localize mesenchymal stem cells (MSCs) to the surface of decellularized bone enhances allograft healing and integration. Herein, we utilize a mixed population of 50:50 MSCs and osteoprogenitor cells to better mimic native periosteum cell population and paracrine factor production to further promote allograft healing. This mixed cell population was localized to the surface of decellularized allografts within degradable hydrogels and shown to expedite allograft healing. Specifically, bone callus formation and biomechanical graft-host integration are increased as compared to unmodified allografts. These results demonstrate the dual importance of periosteum-mediated paracrine factors orchestrating host cell recruitment as well as new bone formation while developing clinically translatable strategies for allograft healing and integration. PMID:25818449

Time-dependent change of blood flow in the prostate treated with high-intensity focused ultrasound.

PubMed

Shoji, Sunao; Tonooka, Akiko; Hashimoto, Akio; Nakamoto, Masahiko; Tomonaga, Tetsuro; Nakano, Mayura; Sato, Haruhiro; Terachi, Toshiro; Koike, Junki; Uchida, Toyoaki

2014-09-01

Avascular areas on contrast-enhanced magnetic resonance imaging have been considered to be areas of localized prostate cancer successfully treated by high-intensity focused ultrasound. However, the optimal timing of magnetic resonance imaging has not been discussed. The thermal effect of high-intensity focused ultrasound is degraded by regional prostatic blood flow. Conversely, the mechanical effect of high-intensity focused ultrasound (cavitation) is not affected by blood flow, and can induce vessel damage. In this series, the longitudinal change of blood flow on contrast-enhanced magnetic resonance imaging was observed from postoperative day 1 to postoperative day 14 in 10 patients treated with high-intensity focused ultrasound. The median rates of increase in the non-enhanced volume of the whole gland, transition zone and peripheral zone from postoperative day 1 to postoperative day 14 were 36%, 39%, and 34%, respectively. In another pathological analysis of the prostate tissue of 17 patients immediately after high-intensity focused ultrasound without neoadjuvant hormonal therapy, we observed diffuse coagulative degeneration and partial non-coagulative prostate tissue around arteries with vascular endothelial cell detachment. These observations on contrast-enhanced magnetic resonance imaging support a time-dependent change of the blood flow in the prostate treated with high-intensity focused ultrasound. Additionally, our pathological findings support the longitudinal changes of these magnetic resonance imaging findings. Further large-scale studies will investigate the most appropriate timing of contrast-enhanced magnetic resonance imaging for evaluation of the effectiveness of high-intensity focused ultrasound for localized prostate cancer. © 2014 The Japanese Urological Association.

Surgical stapling device–tissue interactions: what surgeons need to know to improve patient outcomes

PubMed Central

Chekan, Edward; Whelan, Richard L

2014-01-01

The introduction of both new surgical devices and reengineered existing devices leads to modifications in the way traditional tasks are carried out and allows for the development of new surgical techniques. Each new device has benefits and limitations in regards to tissue interactions that, if known, allow for optimal use. However, most surgeons are unaware of these attributes and, therefore, new device introduction creates a “knowledge gap” that is potentially dangerous. The goal of this review is to present a framework for the study of device– tissue interactions and to initiate the process of “filling in” the knowledge gap via the available literature. Surgical staplers, which are continually being developed, are the focus of this piece. The integrity of the staple line, which depends on adequate tissue compression, is the primary factor in creating a stable anastomosis. This review focuses on published studies that evaluated the creation of stable anastomoses in bariatric, thoracic, and colorectal procedures. Understanding how staplers interact with target tissues is key to improving patient outcomes. It is clear from this review that each tissue type presents unique challenges. The thickness of each tissue varies as do the intrinsic biomechanical properties that determine the ideal compressive force and prefiring compression time for each tissue type. The correct staple height will vary depending on these tissue-specific properties and the tissue pathology. These studies reinforce the universal theme that compression, staple height, tissue thickness, tissue compressibility, and tissue type must all be considered by the surgeon prior to choosing a stapler and cartridge. The surgeon’s experience, therefore, is a critical factor. Educational programs need to be established to inform and update surgeons on the characteristics of each stapler. It is hoped that the framework presented in this review will facilitate this process. PMID:25246812

Advances in biologic augmentation for rotator cuff repair

PubMed Central

Patel, Sahishnu; Gualtieri, Anthony P.; Lu, Helen H.; Levine, William N.

2016-01-01

Rotator cuff tear is a very common shoulder injury that often necessitates surgical intervention for repair. Despite advances in surgical techniques for rotator cuff repair, there is a high incidence of failure after surgery because of poor healing capacity attributed to many factors. The complexity of tendon-to-bone integration inherently presents a challenge for repair because of a large biomechanical mismatch between the tendon and bone and insufficient regeneration of native tissue, leading to the formation of fibrovascular scar tissue. Therefore, various biological augmentation approaches have been investigated to improve rotator cuff repair healing. This review highlights recent advances in three fundamental approaches for biological augmentation for functional and integrative tendon–bone repair. First, the exploration, application, and delivery of growth factors to improve regeneration of native tissue is discussed. Second, applications of stem cell and other cell-based therapies to replenish damaged tissue for better healing is covered. Finally, this review will highlight the development and applications of compatible biomaterials to both better recapitulate the tendon–bone interface and improve delivery of biological factors for enhanced integrative repair. PMID:27750374

Keap1-knockdown decreases fasting-induced fatty liver via altered lipid metabolism and decreased fatty acid mobilization from adipose tissue.

PubMed

Xu, Jialin; Donepudi, Ajay C; Moscovitz, Jamie E; Slitt, Angela L

2013-01-01

The purpose of this study was to determine whether Nrf2 activation, via Keap1-knockdown (Keap1-KD), regulates lipid metabolism and mobilization induced by food deprivation (e.g. fasting). Male C57BL/6 (WT) and Keap1-KD mice were either fed ad libitum or food deprived for 24 hours. After fasting, WT mice exhibited a marked increase in hepatic lipid accumulation, but Keap1-KD mice had an attenuated increase of lipid accumulation, along with reduced expression of lipogenic genes (acetyl-coA carboxylase, stearoyl-CoA desaturase-1, and fatty acid synthase) and reduced expression of genes related to fatty acid transport, such as fatty acid translocase/CD36 (CD36) and Fatty acid transport protein (FATP) 2, which may attribute to the reduced induction of Peroxisome proliferator-activated receptor (Ppar) α signaling in the liver. Additionally, enhanced Nrf2 activity by Keap1-KD increased AMP-activated protein kinase (AMPK) phosphorylation in liver. In white adipose tissue, enhanced Nrf2 activity did not change the lipolysis rate by fasting, but reduced expression of fatty acid transporters--CD36 and FATP1, via a PPARα-dependent mechanism, which impaired fatty acid transport from white adipose tissue to periphery circulation system, and resulted in increased white adipose tissue fatty acid content. Moreover, enhanced Nrf2 activity increased glucose tolerance and Akt phosphorylation levels upon insulin administration, suggesting Nrf2 signaling pathway plays a key role in regulating insulin signaling and enhanced insulin sensitivity in skeletal muscle. Enhanced Nrf2 activity via Keap1-KD decreased fasting-induced steatosis, pointing to an important function of Nrf2 on lipid metabolism under the condition of nutrient deprivation.

Keap1-Knockdown Decreases Fasting-Induced Fatty Liver via Altered Lipid Metabolism and Decreased Fatty Acid Mobilization from Adipose Tissue

PubMed Central

Xu, Jialin; Donepudi, Ajay C.; Moscovitz, Jamie E.; Slitt, Angela L.

2013-01-01

Aims The purpose of this study was to determine whether Nrf2 activation, via Keap1-knockdown (Keap1-KD), regulates lipid metabolism and mobilization induced by food deprivation (e.g. fasting). Methods and Results Male C57BL/6 (WT) and Keap1-KD mice were either fed ad libitum or food deprived for 24 hours. After fasting, WT mice exhibited a marked increase in hepatic lipid accumulation, but Keap1-KD mice had an attenuated increase of lipid accumulation, along with reduced expression of lipogenic genes (acetyl-coA carboxylase, stearoyl-CoA desaturase-1, and fatty acid synthase) and reduced expression of genes related to fatty acid transport, such as fatty acid translocase/CD36 (CD36) and Fatty acid transport protein (FATP) 2, which may attribute to the reduced induction of Peroxisome proliferator-activated receptor (Ppar) α signaling in the liver. Additionally, enhanced Nrf2 activity by Keap1-KD increased AMP-activated protein kinase (AMPK) phosphorylation in liver. In white adipose tissue, enhanced Nrf2 activity did not change the lipolysis rate by fasting, but reduced expression of fatty acid transporters — CD36 and FATP1, via a PPARα-dependent mechanism, which impaired fatty acid transport from white adipose tissue to periphery circulation system, and resulted in increased white adipose tissue fatty acid content. Moreover, enhanced Nrf2 activity increased glucose tolerance and Akt phosphorylation levels upon insulin administration, suggesting Nrf2 signaling pathway plays a key role in regulating insulin signaling and enhanced insulin sensitivity in skeletal muscle. Conclusion Enhanced Nrf2 activity via Keap1-KD decreased fasting-induced steatosis, pointing to an important function of Nrf2 on lipid metabolism under the condition of nutrient deprivation. PMID:24224011

Active spice-derived components can inhibit inflammatory responses of adipose tissue in obesity by suppressing inflammatory actions of macrophages and release of monocyte chemoattractant protein-1 from adipocytes.

PubMed

Woo, Hae-Mi; Kang, Ji-Hye; Kawada, Teruo; Yoo, Hoon; Sung, Mi-Kyung; Yu, Rina

2007-02-13

Inflammation plays a key role in obesity-related pathologies such as cardiovascular disease, type II diabetes, and several types of cancer. Obesity-induced inflammation entails the enhancement of the recruitment of macrophages into adipose tissue and the release of various proinflammatory proteins from fat tissue. Therefore, the modulation of inflammatory responses in obesity may be useful for preventing or ameliorating obesity-related pathologies. Some spice-derived components, which are naturally occurring phytochemicals, elicit antiobesity and antiinflammatory properties. In this study, we investigated whether active spice-derived components can be applied to the suppression of obesity-induced inflammatory responses. Mesenteric adipose tissue was isolated from obese mice fed a high-fat diet and cultured to prepare an adipose tissue-conditioned medium. Raw 264.7 macrophages were treated with the adipose tissue-conditioned medium with or without active spice-derived components (i.e., diallyl disulfide, allyl isothiocyanate, piperine, zingerone and curcumin). Chemotaxis assay was performed to measure the degree of macrophage migration. Macrophage activation was estimated by measuring tumor necrosis factor-alpha (TNF-alpha), nitric oxide, and monocyte chemoattractant protein-1 (MCP-1) concentrations. The active spice-derived components markedly suppressed the migration of macrophages induced by the mesenteric adipose tissue-conditioned medium in a dose-dependent manner. Among the active spice-derived components studied, allyl isothiocyanate, zingerone, and curcumin significantly inhibited the cellular production of proinflammatory mediators such as TNF-alpha and nitric oxide, and significantly inhibited the release of MCP-1 from 3T3-L1 adipocytes. Our findings suggest that the spice-derived components can suppress obesity-induced inflammatory responses by suppressing adipose tissue macrophage accumulation or activation and inhibiting MCP-1 release from adipocytes. These spice-derived components may have a potential to improve chronic inflammatory conditions in obesity.

Transcription factors ETF, E2F, and SP-1 are involved in cytokine-independent proliferation of murine hepatocytes.

PubMed

Zellmer, Sebastian; Schmidt-Heck, Wolfgang; Godoy, Patricio; Weng, Honglei; Meyer, Christoph; Lehmann, Thomas; Sparna, Titus; Schormann, Wiebke; Hammad, Seddik; Kreutz, Clemens; Timmer, Jens; von Weizsäcker, Fritz; Thürmann, Petra A; Merfort, Irmgard; Guthke, Reinhard; Dooley, Steven; Hengstler, Jan G; Gebhardt, Rolf

2010-12-01

The cellular basis of liver regeneration has been intensely investigated for many years. However, the mechanisms initiating hepatocyte "plasticity" and priming for proliferation are not yet fully clear. We investigated alterations in gene expression patterns during the first 72 hours of C57BL/6N mouse hepatocyte culture on collagen monolayers (CM), which display a high basal frequency of proliferation in the absence of cytokines. Although many metabolic genes were down-regulated, genes related to mitogen-activated protein kinase (MAPK) signaling and cell cycle were up-regulated. The latter genes showed an overrepresentation of transcription factor binding sites (TFBS) for ETF (TEA domain family member 2), E2F1 (E2F transcription factor 1), and SP-1 (Sp1 transcription factor) (P < 0.001), all depending on MAPK signaling. Time-dependent increase of ERK1/2 phosphorylation occurred during the first 48 hours (and beyond) in the absence of cytokines, accompanied by an enhanced bromodeoxyuridine labeling index of 20%. The MEK inhibitor PD98059 blunted these effects indicating MAPK signaling as major trigger for this cytokine-independent proliferative response. In line with these in vitro findings, liver tissue of mice challenged with CCl(4) displayed hepatocytes with intense p-ERK1/2 staining and nuclear SP-1 and E2F1 expression. Furthermore, differentially expressed genes in mice after partial hepatectomy contained overrepresented TFBS for ETF, E2F1, and SP-1 and displayed increased expression of E2F1. Cultivation of murine hepatocytes on CM primes cells for proliferation through cytokine-independent activation of MAPK signaling. The transcription factors ETF, E2F1, and SP-1 seem to play a pronounced role in mediating proliferation-dependent differential gene expression. Similar events, but on a shorter time-scale, occur very early after liver damage in vivo. Copyright © 2010 American Association for the Study of Liver Diseases.

Effect of misonidazole or metronidazole pretreatment on the response of the RIF-1 mouse sarcoma to melphalan, cyclophosphamide, chlorambucil and CCNU.

PubMed Central

Twentyman, P.; Workman, P.

1982-01-01

The effect has been studied of adding either misonidazole (MISO) or metronidazole (METRO) to cytotoxic drug treatment of C3H mice bearing the RIF-1 sarcoma. The nitroimidazoles were injected 30 min before the cytotoxic drugs at a dose of 2 . 5 mmol/kg. Both clonogenic-cell survival and growth delay were measured as indicators of tumour response and depression in WBC count and acute lethality were used to indicate normal-tissue response. For melphalan, neither pretreatment agent produced any change in tumor response. For cyclophosphamide, no change was produced by METRO but a minimal increase in tumour response occurred with MISO. An enhancement of cell killing by CCNU was seen with MISO pretreatment, but there was no increase in tumour growth delay. METRO, however, did not enhance tumour response by either endpoint. WBC depression by CCNU was not enhanced by MISO pretreatment, and there was no significant reduction in the acute LD50. This indicates a therapeutic advantage from the addition of MISO to CCNU in this model system. For chlorambucil, considerable enhancement of tumour response followed either MISO or METRO pretreatment (dose-modifying factors of 2 . 0 and 1 . 4 respectively). However, the modification by MISO of normal-tissue response to chlorambucil was also enhanced by about a factor of 2, with no therapeutic gain. PMID:7073938

Adipose tissue NAD+ biology in obesity and insulin resistance: From mechanism to therapy.

PubMed

Yamaguchi, Shintaro; Yoshino, Jun

2017-05-01

Nicotinamide adenine dinucleotide (NAD + ) biosynthetic pathway, mediated by nicotinamide phosphoribosyltransferase (NAMPT), a key NAD + biosynthetic enzyme, plays a pivotal role in controlling many biological processes, such as metabolism, circadian rhythm, inflammation, and aging. Over the past decade, NAMPT-mediated NAD + biosynthesis, together with its key downstream mediator, namely the NAD + -dependent protein deacetylase SIRT1, has been demonstrated to regulate glucose and lipid metabolism in a tissue-dependent manner. These discoveries have provided novel mechanistic and therapeutic insights into obesity and its metabolic complications, such as insulin resistance, an important risk factor for developing type 2 diabetes and cardiovascular disease. This review will focus on the importance of adipose tissue NAMPT-mediated NAD + biosynthesis and SIRT1 in the pathophysiology of obesity and insulin resistance. We will also critically explore translational and clinical aspects of adipose tissue NAD + biology. © 2017 WILEY Periodicals, Inc.

The perlecan-interacting growth factor progranulin regulates ubiquitination, sorting, and lysosomal degradation of sortilin.

PubMed

Tanimoto, Ryuta; Palladino, Chiara; Xu, Shi-Qiong; Buraschi, Simone; Neill, Thomas; Gomella, Leonard G; Peiper, Stephen C; Belfiore, Antonino; Iozzo, Renato V; Morrione, Andrea

2017-12-01

Despite extensive clinical and experimental studies over the past decades, the pathogenesis and progression to the castration-resistant stage of prostate cancer remains largely unknown. Progranulin, a secreted growth factor, strongly binds the heparin-sulfate proteoglycan perlecan, and counteracts its biological activity. We established that progranulin acts as an autocrine growth factor and promotes prostate cancer cell motility, invasion, and anchorage-independent growth. Progranulin was overexpressed in prostate cancer tissues vis-à-vis non-neoplastic tissues supporting the hypothesis that progranulin may play a key role in prostate cancer progression. However, progranulin's mode of action is not well understood and proteins regulating progranulin signaling have not been identified. Sortilin, a single-pass type I transmembrane protein of the Vps10 family, binds progranulin in neurons and targets progranulin for lysosomal degradation. Significantly, in DU145 and PC3 cells, we detected very low levels of sortilin associated with high levels of progranulin production and enhanced motility. Restoring sortilin expression decreased progranulin levels, inhibited motility and anchorage-independent growth and destabilized Akt. These results demonstrated a critical role for sortilin in regulating progranulin and suggest that sortilin loss may contribute to prostate cancer progression. Here, we provide the novel observation that progranulin downregulated sortilin protein levels independent of transcription. Progranulin induced sortilin ubiquitination, internalization via clathrin-dependent endocytosis and sorting into early endosomes for lysosomal degradation. Collectively, these results constitute a regulatory feed-back mechanism whereby sortilin downregulation ensures sustained progranulin-mediated oncogenesis. Copyright © 2017. Published by Elsevier B.V.

CONNECTIVE TISSUE GROWTH FACTOR IS A TARGET OF NOTCH SIGNALING IN CELLS OF THE OSTEOBLASTIC LINEAGE

PubMed Central

Canalis, Ernesto; Zanotti, Stefano; Smerdel-Ramoya, Anna

2014-01-01

Connective tissue growth factor (Ctgf) or CCN2 is a protein synthesized by osteoblasts necessary for skeletal homeostasis, although its overexpression inhibits osteogenic signals and bone formation. Ctgf is induced by bone morphogenetic proteins, transforming growth factor β and Wnt; and in the present studies, we explored whether Notch regulated Ctgf expression in osteoblasts. We employed RosaNotch mice, where the Notch intracellular domain (NICD) is expressed following the excision of a STOP cassette, placed between the Rosa26 promoter and NICD. Notch was activated by transduction of adenoviral vectors expressing Cre recombinase (Ad-CMV-Cre). Notch induced Ctgf mRNA levels in a time dependent manner and increased Ctgf heterogeneous nuclear RNA. Notch also destabilized Ctgf mRNA shortening its half-life from 13 h to 3 h. The effect of Notch on Ctgf expression was lost following Rbpjκ downregulation, demonstrating that it was mediated by Notch canonical signaling. However, downregulation of the classic Notch target genes Hes1, Hey1 and Hey2 did not modify the effect of Notch on Ctgf expression. Wild type osteoblasts exposed to immobilized Delta-like 1 displayed enhanced Notch signaling and increased Ctgf expression. In addition to the effects of Notch in vitro, Notch induced Ctgf in vivo, and calvariae and femurs from RosaNotch mice mated with transgenics expressing the Cre recombinase in cells of the osteoblastic lineage exhibited increased expression of Ctgf. In conclusion, Ctgf is a target of Notch canonical signaling in osteoblasts, and may act in concert with Notch to regulate skeletal homeostasis. PMID:24792956

Association Between Fungal Contamination and Eye Bank-Prepared Endothelial Keratoplasty Tissue: Temperature-Dependent Risk Factors and Antifungal Supplementation of Optisol-Gentamicin and Streptomycin.

PubMed

Brothers, Kimberly M; Shanks, Robert M Q; Hurlbert, Susan; Kowalski, Regis P; Tu, Elmer Y

2017-11-01

Fungal contamination and infection from donor tissues processed for endothelial keratoplasty is a growing concern, prompting analysis of donor tissues after processing. To determine whether eyebank-processed endothelial keratoplasty tissue is at higher risk of contamination than unprocessed tissue and to model eyebank processing with regard to room temperature exposure on Candida growth in optisol-gentamicin and streptomycin (GS) with and without antifungal supplementation. An examination of the 2013 Eversight Eyebank Study follow-up database for risk factors associated with post-keratoplasty infection identified an increased risk of positive fungal rim culture results in tissue processed for endothelial keratoplasty vs unprocessed tissue. Processing steps at room temperature were hypothesized as a potential risk factor for promotion of fungal growth between these 2 processes. Candida albicans, Candida glabrata, and Candida parapsilosis endophthalmitis isolates were each inoculated into optisol-GS and subjected to 2 different room temperature incubation regimens reflective of current corneal tissue handling protocols. Eversight Eyebank Study outcomes and measures were follow-up inquiries from 6592 corneal transplants. Efficacy study outcomes and measures were fungal colony-forming units from inoculated vials of optisol-GS taken at 2 different processing temperatures. Donor rim culture results were 3 times more likely to be positive for fungi in endothelial keratoplasty-processed eyes (1.14%) than for other uses (0.37%) (difference, 0.77%; 95% CI, 0.17-.1.37) (P = .009). In vitro, increased room temperature incubation of optisol-GS increased growth of Candida species over time. The addition of caspofungin and voriconazole decreased growth of Candida in a species-dependent manner. Detectable Candida growth in donor rim cultures, associated with a higher rate of post keratoplasty infection, is seen in endothelial keratoplasty tissue vs other uses at the time of transplantation, likely owing in part to eyebank preparation processes extending the time of tissue warming. Reduced room temperature incubation and the addition of antifungal agents decreased growth of Candida species in optisol-GS and should be further explored to reduce the risk of infection.

Effects of elemental composition on the incorporation of dietary nitrogen and carbon isotopic signatures in an omnivorous songbird

DOE Office of Scientific and Technical Information (OSTI.GOV)

Pearson, Scott, F.; Levey, Douglas, J.; Greenberg, Catheryn, H.

2003-02-28

Pearson, S.F., D.J. Levey, C.H. Greenberg, and C.M. del Rio. 2003. Effects of elemental composition on the incorporation of dietary nitrogen and carbon isotopic signatures in an omnivorous songbird. Oecologia. 135:516-523. The use of stable isotopes to infer diet requires quantifying the relationship between diet and tissues and, in particular, knowing of how quickly isotopes turnover in different tissues and how isotopic concentrations of different food components change (discriminate) when incorporated into consumer tissues. We used feeding trials with wild-caught yellow-rumped warblers (Dendroica coronata) to determine d15N and d13C turnover rates for blood, d15N and d13C diet-tissue discrimination factors, andmore » diet-tissue relationships for blood and feathers. After 3 weeks on a common diet, 36 warblers were assigned to one of four diets differing in the relative proportion of fruit and insects. Plasma half-life estimates ranged from 0.4 to 0.7 days for d13C and from 0.5 to 1.7 days for d15N. Half-life did not differ among diets. Whole blood half-life for d13C ranged from 3.9 to 6.1 days. Yellow-rumped warbler tissues were enriched relative to diet by 1.7.3.6% for nitrogen isotopes and by 1.2 to 4.3% for carbon isotopes, depending on tissue and diet. Consistent with previous studies, feathers were the most enriched and whole blood and plasma were the least enriched or, in the case of carbon, slightly depleted relative to diet. In general, tissues were more enriched relative to diet for birds with high percentages of insects. For all tissues, carbon and nitrogen isotope discrimination factors increased with carbon and nitrogen concentrations of diets. The isotopic signature of plasma increased linearly with the sum of the isotopic signature of the diet and the discrimination factor. Because the isotopic signature of tissues depends on both elemental concentration and isotopic signature of the diet, attempts to reconstruct diet from stable isotope signatures require mixing models that incorporate elemental concentration.« less

Actin depolymerization enhances adipogenic differentiation in human stromal stem cells.

PubMed

Chen, Li; Hu, Huimin; Qiu, Weimin; Shi, Kaikai; Kassem, Moustapha

2018-05-01

Human stromal stem cells (hMSCs) differentiate into adipocytes that play a role in skeletal tissue homeostasis and whole body energy metabolism. During adipocyte differentiation, hMSCs exhibit significant changes in cell morphology suggesting changes in cytoskeletal organization. Here, we examined the effect of direct modulation of actin microfilament dynamics on adipocyte differentiation. Stabilizing actin filaments in hMSCs by siRNA-mediated knock down of the two main actin depolymerizing factors (ADFs): Cofilin 1 (CFL1) and Destrin (DSTN) or treating the cells by Phalloidin reduced adipocyte differentiation as evidenced by decreased number of mature adipocytes and decreased adipocyte specific gene expression (ADIPOQ, LPL, PPARG, FABP4). In contrast, disruption of actin cytoskeleton by Cytochalasin D enhanced adipocyte differentiation. Follow up studies revealed that the effects of CFL1 on adipocyte differentiation depended on the activity of LIM domain kinase 1 (LIMK1) which is the major upstream kinase of CFL1. Inhibiting LIMK by its specific chemical inhibitor LIMKi inhibited the phosphorylation of CFL1 and actin polymerization, and enhanced the adipocyte differentiation. Moreover, treating hMSCs by Cytochalasin D inhibited ERK and Smad2 signaling and this was associated with enhanced adipocyte differentiation. On the other hand, Phalloidin enhanced ERK and Smad2 signaling, but inhibited adipocyte differentiation which was rescued by ERK specific chemical inhibitor U0126. Our data provide a link between restructuring of hMSCs cytoskeleton and hMSCs lineage commitment and differentiation. Copyright © 2018 Elsevier B.V. All rights reserved.

Idiopathic pulmonary fibrosis fibroblasts become resistant to Fas ligand-dependent apoptosis via the alteration of decoy receptor 3.

PubMed

Im, Jintaek; Kim, Kyutae; Hergert, Polla; Nho, Richard Seonghun

2016-09-01

Idiopathic pulmonary fibrosis (IPF) is an irreversible lethal lung disease with an unknown etiology. IPF patients' lung fibroblasts express inappropriately high Akt activity, protecting them in response to an apoptosis-inducing type I collagen matrix. FasL, a ligand for Fas, is known to be increased in the lung tissues of patients with IPF, implicated with the progression of IPF. Expression of Decoy Receptor3 (DcR3), which binds to FasL, thereby subsequently suppressing the FasL-Fas-dependent apoptotic pathway, is frequently altered in various human disease. However, the role of DcR3 in IPF fibroblasts in regulating their viability has not been examined. We found that enhanced DcR3 expression exists in the majority of IPF fibroblasts on collagen matrices, resulting in the protection of IPF fibroblasts from FasL-induced apoptosis. Abnormally high Akt activity suppresses GSK-3β function, thereby accumulating the nuclear factor of activated T-cells cytoplasmic 1 (NFATc1) in the nucleus, increasing DcR3 expression in IPF fibroblasts. This alteration protects IPF cells from FasL-induced apoptosis on collagen. However, the inhibition of Akt or NFATc1 decreases DcR3 mRNA and protein levels, which sensitizes IPF fibroblasts to FasL-mediated apoptosis. Furthermore, enhanced DcR3 and NFATc1 expression is mainly present in myofibroblasts in the fibroblastic foci of lung tissues derived from IPF patients. Our results showed that when IPF cells interact with collagen matrix, aberrantly activated Akt increases DcR3 expression via GSK-3β-NFATc1 and protects IPF cells from the FasL-dependent apoptotic pathway. These findings suggest that the inhibition of DcR3 function may be an effective approach for sensitizing IPF fibroblasts in response to FasL, limiting the progression of lung fibrosis. Copyright © 2016 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd. Copyright © 2016 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

Tissue Engineering Using Transfected Growth-Factor Genes

NASA Technical Reports Server (NTRS)

Madry, Henning; Langer, Robert S.; Freed, Lisa E.; Trippel, Stephen; Vunjak-Novakovic, Gordana

2005-01-01

A method of growing bioengineered tissues includes, as a major component, the use of mammalian cells that have been transfected with genes for secretion of regulator and growth-factor substances. In a typical application, one either seeds the cells onto an artificial matrix made of a synthetic or natural biocompatible material, or else one cultures the cells until they secrete a desired amount of an extracellular matrix. If such a bioengineered tissue construct is to be used for surgical replacement of injured tissue, then the cells should preferably be the patient s own cells or, if not, at least cells matched to the patient s cells according to a human-leucocyteantigen (HLA) test. The bioengineered tissue construct is typically implanted in the patient's injured natural tissue, wherein the growth-factor genes enhance metabolic functions that promote the in vitro development of functional tissue constructs and their integration with native tissues. If the matrix is biodegradable, then one of the results of metabolism could be absorption of the matrix and replacement of the matrix with tissue formed at least partly by the transfected cells. The method was developed for articular chondrocytes but can (at least in principle) be extended to a variety of cell types and biocompatible matrix materials, including ones that have been exploited in prior tissue-engineering methods. Examples of cell types include chondrocytes, hepatocytes, islet cells, nerve cells, muscle cells, other organ cells, bone- and cartilage-forming cells, epithelial and endothelial cells, connective- tissue stem cells, mesodermal stem cells, and cells of the liver and the pancreas. Cells can be obtained from cell-line cultures, biopsies, and tissue banks. Genes, molecules, or nucleic acids that secrete factors that influence the growth of cells, the production of extracellular matrix material, and other cell functions can be inserted in cells by any of a variety of standard transfection techniques.

Therapeutically Targeting the Inflammasome Product in a Chimeric Model of Endometriosis-Related Surgical Adhesions.

PubMed

Stocks, Meredith M; Crispens, Marta A; Ding, Tianbing; Mokshagundam, Shilpa; Bruner-Tran, Kaylon L; Osteen, Kevin G

2017-08-01

Development of adhesions commonly occurs in association with surgery for endometriosis. Even in the absence of surgery, women with endometriosis appear to be at an enhanced risk of developing adhesions. In the current study, we utilized a chimeric mouse model of experimental endometriosis in order to examine the role of inflammasome activation in the development of postsurgical adhesions. Mice were randomized to receive peritoneal injections of human endometrial tissue fragments or endometrial tissue conditioned media (CM) from women with or without endometriosis 16 hours after ovariectomy and placement of an estradiol-releasing silastic capsule. A subset of mice receiving CM was also treated with interleukin (IL) 1 receptor antagonist (IL-1ra). Our studies demonstrate that peritoneal injection of endometrial tissue fragments near the time of surgery resulted in extensive adhesive disease regardless of tissue origin. However, adhesion scores were significantly higher in mice receiving CM from tissues acquired from patients with endometriosis compared to control tissue CM ( P = .0001). Cytokine bead array analysis of endometrial CM revealed enhanced expression of IL-1β from patients with endometriosis compared to controls ( P < .01). Finally, the ability of human tissue CM to promote adhesive disease was dramatically reduced in mice cotreated with IL-1ra ( P < .0001). Our data implicate enhanced expression of IL-1β in women with endometriosis as a potential causal factor in their increased susceptibility of developing postsurgical adhesions. Thus, targeting inflammasome activation may be an effective strategy for the prevention of surgical adhesions in patients with endometriosis.

Enhancement of ultraweak photon emission with 3 MHz ultrasonic irradiation on transplanted tumor tissues of mice.

PubMed

Kim, Hongbae; Ahn, Saeyoung; Kim, Jungdae; Soh, Kwang-Sup

2008-07-01

We investigated photon emissions of various bio-samples which were induced by ultrasonic stimulation. It has been reported that ultrasonic stimulations induced the thermal excitation of the bio-tissues. After ultrasonic stimulation, any measurement of photon radiation in the visible spectral range has not been carried out yet. The instruments consisted of electronic devices for an ultrasonic generator of the frequency 3 MHz and a photomultiplier tube (PMT) system counting photons from bio-tissues. The transplanted tumor tissues of mice were prepared for the experiments and their liver and spleen tissues were also used for the controls. It was found that the continuous ultrasonic stimulations with the electrical power 2300 mW induced ultraweak photon emissions from the tumor tissues. The number of induced photon was dependent of the type of the tissues and the stimulation time intervals. The level of photon emission was increased from the mouse tumor exposed to the ultrasonic stimulations, and the changes were discriminated from those of the spleens and livers.

Monitoring of coagulation factor therapy in patients with von Willebrand disease type 3 using a microchip flow chamber system.

PubMed

Ågren, Anna; Holmström, Margareta; Schmidt, David E; Hosokawa, Kazuya; Blombäck, Margareta; Hjemdahl, Paul

2017-01-05

Patients with type 3 von Willebrand disease (VWD-3) have no measurable levels of VW factor (VWF) and usually require treatment with VWF-FVIII concentrate to prevent and/or stop bleeding. Even though the patients are treated prophylactically, they may experience bleeding symptoms. The aim of this study was to evaluate the effect of VWF-FVIII concentrate treatment in VWD-3 patients with the Total Thrombus Analysis System (T-TAS ® ), which measures thrombus formation under flow conditions. Coagulation profiles of 10 VWD-3 patients were analysed using T-TAS before and 30 minutes after VWF-FVIII concentrate (Haemate ® ) injection. Results were compared to VWF- and FVIII activity in plasma, and results with thromboelastometry and ristocetin-activated platelet impedance aggregometry (Multiplate ® ) in whole blood. For comparison, 10 healthy controls were also analysed with T-TAS. A median dose of 27 (range 15-35) IU/kg of VWF-FVIII concentrate increased VWF- and FVIII activity as expected. T-TAS thrombus formation was enhanced when a tissue factor/collagen-coated flow chamber was used at low shear, but treatment effects at high shear using a collagen-coated flow chamber were minimal. Whole blood coagulation assessed by thromboelastometry was normal and did not change (p > 0.05) but ristocetin-induced platelet aggregation improved (p < 0.001). In conclusion, T-TAS detects effects of VWF-FVIII concentrate treatment on coagulation-dependent thrombus formation at low shear, but minor effects are observed on platelet-dependent thrombus formation at high shear. The poor prediction of bleeding by conventional laboratory monitoring in VWD-3 patients might be related to insufficient restoration of platelet-dependent thrombus formation.

Long-distance communication and signal amplification in systemic acquired resistance

PubMed Central

Shah, Jyoti; Zeier, Jürgen

2013-01-01

Systemic acquired resistance (SAR) is an inducible defense mechanism in plants that confers enhanced resistance against a variety of pathogens. SAR is activated in the uninfected systemic (distal) organs in response to a prior (primary) infection elsewhere in the plant. SAR is associated with the activation of salicylic acid (SA) signaling and the priming of defense responses for robust activation in response to subsequent infections. The activation of SAR requires communication by the primary infected tissues with the distal organs. The vasculature functions as a conduit for the translocation of factors that facilitate long-distance intra-plant communication. In recent years, several metabolites putatively involved in long-distance signaling have been identified. These include the methyl ester of SA (MeSA), the abietane diterpenoid dehydroabietinal (DA), the dicarboxylic acid azelaic acid (AzA), and a glycerol-3-phosphate (G3P)-dependent factor. Long-distance signaling by some of these metabolites also requires the lipid-transfer protein DIR1 (DEFECTIVE IN INDUCED RESISTANCE 1). The relative contribution of these factors in long-distance signaling is likely influenced by environmental conditions, for example light. In the systemic leaves, the AGD2-LIKE DEFENSE RESPONSE PROTEIN1 (ALD1)-dependent production of the lysine catabolite pipecolic acid (Pip), FLAVIN-DEPENDENT MONOOXYGENASE1 (FMO1) signaling, as well as SA synthesis and downstream signaling are required for the activation of SAR. This review summarizes the involvement and interaction between long-distance SAR signals and details the recently discovered role of Pip in defense amplification and priming that allows plants to acquire immunity at the systemic level. Recent advances in SA signaling and perception are also highlighted. PMID:23440336

Blue light alters miR167 expression and microRNA-targeted auxin response factor genes in Arabidopsis thaliana plants.

PubMed

Pashkovskiy, Pavel P; Kartashov, Alexander V; Zlobin, Ilya E; Pogosyan, Sergei I; Kuznetsov, Vladimir V

2016-07-01

The effect of blue LED (450 nm) on the photomorphogenesis of Arabidopsis thaliana Col-0 plants and the transcript levels of several genes, including miRNAs, photoreceptors and auxin response factors (ARF) was investigated. It was observed that blue light accelerated the generative development, reduced the rosette leaf number, significantly reduced the leaf area, dry biomass and led to the disruption of conductive tissue formation. The blue LED differentially influenced the transcript levels of several phytochromes (PHY a, b, c, d, and e), cryptochromes (CRY 1 and 2) and phototropins (PHOT 1 and 2). At the same time, the blue LED significantly increased miR167 expression compared to a fluorescent lamp or white LEDs. This increase likely resulted in the enhanced transcription of the auxin response factor genes ARF4 and ARF8, which are regulated by this miRNA. These findings support the hypothesis that the effects of blue light on A. thaliana are mediated by auxin signalling pathway involving miRNA-dependent regulation of ARF gene expression. Copyright © 2016 Elsevier Masson SAS. All rights reserved.

Juvenile Hormone Analogues, Methoprene and Fenoxycarb Dose-Dependently Enhance Certain Enzyme Activities in the Silkworm Bombyx Mori (L)

PubMed Central

Mamatha, Devi M.; Kanji, Vijaya K.; Cohly, Hari H.P.; Rao, M. Rajeswara

2008-01-01

Use of Juvenile Hormone Analogues (JHA) in sericulture practices has been shown to boost good cocoon yield; their effect has been determined to be dose-dependent. We studied the impact of low doses of JHA compounds such as methoprene and fenoxycarb on selected key enzymatic activities of the silkworm Bombyx mori. Methoprene and fenoxycarb at doses of 1.0 μg and 3.0fg/larvae/48 hours showed enhancement of the 5th instar B. mori larval muscle and silkgland protease, aspartate aminotransaminase (AAT) and alanine aminotransaminase (ALAT), adenosine triphosphate synthase (ATPase) and cytochrome-c-oxidase (CCO) activity levels, indicating an upsurge in the overall oxidative metabolism of the B.mori larval tissues. PMID:18678927

Collection efficiency of a single optical fiber in turbid media.

PubMed

Bargo, Paulo R; Prahl, Scott A; Jacques, Steven L

2003-06-01

If a single optical fiber is used for both delivery and collection of light, two major factors affect the measurement of collected light: (1) the light transport in the medium that describes the amount of light that returns to the fiber and (2) the light coupling to the optical fiber that depends on the angular distribution of photons entering the fiber. We focus on the importance of the latter factor and describe how the efficiency of the coupling depends on the optical properties of the medium. For highly scattering tissues, the efficiency is well predicted by the numerical aperture (NA) of the fiber. For lower scattering, such as in soft tissues, photons arrive at the fiber from deeper depths, and the coupling efficiency could increase twofold to threefold above that predicted by the NA.

Nanoparticle-mediated transcriptional modification enhances neuronal differentiation of human neural stem cells following transplantation in rat brain.

PubMed

Li, Xiaowei; Tzeng, Stephany Y; Liu, Xiaoyan; Tammia, Markus; Cheng, Yu-Hao; Rolfe, Andrew; Sun, Dong; Zhang, Ning; Green, Jordan J; Wen, Xuejun; Mao, Hai-Quan

2016-04-01

Strategies to enhance survival and direct the differentiation of stem cells in vivo following transplantation in tissue repair site are critical to realizing the potential of stem cell-based therapies. Here we demonstrated an effective approach to promote neuronal differentiation and maturation of human fetal tissue-derived neural stem cells (hNSCs) in a brain lesion site of a rat traumatic brain injury model using biodegradable nanoparticle-mediated transfection method to deliver key transcriptional factor neurogenin-2 to hNSCs when transplanted with a tailored hyaluronic acid (HA) hydrogel, generating larger number of more mature neurons engrafted to the host brain tissue than non-transfected cells. The nanoparticle-mediated transcription activation method together with an HA hydrogel delivery matrix provides a translatable approach for stem cell-based regenerative therapy. Copyright © 2016 Elsevier Ltd. All rights reserved.

Wound healing potential of adipose tissue stem cell extract.

PubMed

Na, You Kyung; Ban, Jae-Jun; Lee, Mijung; Im, Wooseok; Kim, Manho

2017-03-25

Adipose tissue stem cells (ATSCs) are considered as a promising source in the field of cell therapy and regenerative medicine. In addition to direct cell replacement using stem cells, intercellular molecule exchange by stem cell secretory factors showed beneficial effects by reducing tissue damage and augmentation of endogenous repair. Delayed cutaneous wound healing is implicated in many conditions such as diabetes, aging, stress and alcohol consumption. However, the effects of cell-free extract of ATSCs (ATSC-Ex) containing secretome on wound healing process have not been investigated. In this study, ATSC-Ex was topically applied on the cutaneous wound and healing speed was examined. As a result, wound closure was much faster in the cell-free extract treated wound than control wound at 4, 6, 8 days after application of ATSC-Ex. Dermal fibroblast proliferation, migration and extracellular matrix (ECM) production are critical aspects of wound healing, and the effects of ATSC-Ex on human dermal fibroblast (HDF) was examined. ATSC-Ex augmented HDF proliferation in a dose-dependent manner and migration ability was enhanced by extract treatment. Representative ECM proteins, collagen type I and matrix metalloproteinase-1, are significantly up-regulated by treatment of ATSC-Ex. Our results suggest that the ATSC-Ex have improving effect of wound healing and can be the potential therapeutic candidate for cutaneous wound healing. Copyright © 2017 Elsevier Inc. All rights reserved.

A probable risk factor of female breast cancer: study on benign and malignant breast tissue samples.

PubMed

Rehman, Sohaila; Husnain, Syed M

2014-01-01

The study reports enhanced Fe, Cu, and Zn contents in breast tissues, a probable risk factor of breast cancer in females. Forty-one formalin-fixed breast tissues were analyzed using atomic absorption spectrophotometry. Twenty malignant, six adjacent to malignant and 15 benign tissues samples were investigated. The malignant tissues samples were of grade 11 and type invasive ductal carcinoma. The quantitative comparison between the elemental levels measured in the two types of specimen (benign and malignant) tissues (removed after surgery) suggests significant elevation of these metals (Fe, Cu, and Zn) in the malignant tissue. The specimens were collected just after mastectomy of women aged 19 to 59 years from the hospitals of Islamabad and Rawalpindi, Pakistan. Most of the patients belong to urban areas of Pakistan. Findings of study depict that these elements have a promising role in the initiation and development of carcinoma as consistent pattern of elevation for Fe, Cu, and Zn was observed. The results showed the excessive accumulation of Fe (229 ± 121 mg/L) in malignant breast tissue samples of patients (p < 0.05) to that in benign tissues samples (49.1 ± 11.4 mg/L). Findings indicated that excess accumulation of iron in malignant tissues can be a risk factor of breast cancer. In order to validate our method of analysis, certified reference material muscle tissue lyophilized (IAEA) MA-M-2/TM was analyzed for metal studied. Determined concentrations were quite in good agreement with certified levels. Asymmetric concentration distribution for Fe, Cu, and Zn was observed in both malignant and benign tissue samples.

Intraocular tissue ablation using an optical fibre to deliver the 5th harmonic of a Nd:YAG

NASA Astrophysics Data System (ADS)

Miller, Joseph; Yu, Xiaobo; Yu, Paula K.; Cringle, Stephen J.; Yu, Dao-Yi

2009-02-01

We report the evaluation of a system which delivers the 5th harmonic of an Nd:YAG (213nm) via optical fibre to ocular tissue sites. The 213nm beam is concentrated, using a hollow glass taper, prior to launch into 200 μm or 600 μm core diameter silica/silica optical fibre. The fibre tip was tapered to enhance the fluence delivered. An operating window of fluence values that could be delivered via 330 - 1100mm lengths of optical fibre was determined. The lower value of 0.2J/cm2 determined by the ablation threshold of the tissue and the upper value of 1.3J/cm2 by the launch, transmission and tip characteristics of the optical fibre. The fluence output decreased as a function of both transmitted pulse energy and number of pulses transmitted. Fresh retinal tissue was cleanly ablated with minimal damage to the surrounding tissue. Lesions were generated using 1, 3 and 10 pulses with fluences from 0.2 to 1.0J/cm2. The lesion depth demonstrated clear dose dependence. Lesions generated in ex vivo preparations of human trabecular meshwork in a fluid environment also demonstrated dose dependence, 50 pulses being sufficient to create a hole within the trabecular meshwork extending to Schlemm's canal. The dose dependence of the ablation depth combined with the ability of this technique to create a conduit through to Schlemm's canal demonstrates the potential of this technique for ophthalmological applications requiring precise and controlled intraocular tissue removal and has potential applications in the treatment and management of glaucoma.

Negative effects of a high tumour necrosis factor-α concentration on human gingival mesenchymal stem cell trophism: the use of natural compounds as modulatory agents.

PubMed

Giacomelli, Chiara; Natali, Letizia; Nisi, Marco; De Leo, Marinella; Daniele, Simona; Costa, Barbara; Graziani, Filippo; Gabriele, Mario; Braca, Alessandra; Trincavelli, M Letizia; Martini, Claudia

2018-05-11

Adult mesenchymal stem cells (MSCs) play a crucial role in the maintenance of tissue homeostasis and in regenerative processes. Among the different MSC types, the gingiva-derived mesenchymal stem cells (GMSCs) have arisen as a promising tool to promote the repair of damaged tissues secreting trophic mediators that affect different types of cells involved in regenerative processes. Tumour necrosis factor (TNF)-α is one of the key mediators of inflammation that could affect tissue regenerative processes and modify the MSC properties in in-vitro applications. To date, no data have been reported on the effects of TNF-α on GMSC trophic activities and how its modulation with anti-inflammatory agents from natural sources could modulate the GMSC properties. GMSCs were isolated and characterized from healthy subjects. The effects of TNF-α were evaluated on GMSCs and on the well-being of endothelial cells. The secretion of cytokines was measured and related to the modification of GMSC-endothelial cell communication using a conditioned-medium method. The ability to modify the inflammatory response was evaluated in the presence of Ribes nigrum bud extract (RBE). TNF-α differently affected GMSC proliferation and the expression of inflammatory-related proteins (interleukin (IL)-6, IL-10, transforming growth factor (TGF)-β, and cyclooxygenase (COX)-2) dependent on its concentration. A high TNF-α concentration decreased the GMSC viability and impaired the positive cross-talk between GMSCs and endothelial cells, probably by enhancing the amount of pro-inflammatory cytokines in the GMSC secretome. RBE restored the beneficial effects of GMSCs on endothelial viability and motility under inflammatory conditions. A high TNF-α concentration decreased the well-being of GMSCs, modifying their trophic activities and decreasing endothelial cell healing. These data highlight the importance of controlling TNF-α concentrations to maintain the trophic activity of GMSCs. Furthermore, the use of natural anti-inflammatory agents restored the regenerative properties of GMSCs on endothelial cells, opening the way to the use and development of natural extracts in wound healing, periodontal regeneration, and tissue-engineering applications that use MSCs.

Heparin functionalization increases retention of TGF-β2 and GDF5 on biphasic silk fibroin scaffolds for tendon/ligament-to-bone tissue engineering.

PubMed

Font Tellado, Sònia; Chiera, Silvia; Bonani, Walter; Poh, Patrina S P; Migliaresi, Claudio; Motta, Antonella; Balmayor, Elizabeth R; van Griensven, Martijn

2018-05-01

The tendon/ligament-to-bone transition (enthesis) is a highly specialized interphase tissue with structural gradients of extracellular matrix composition, collagen molecule alignment and mineralization. These structural features are essential for enthesis function, but are often not regenerated after injury. Tissue engineering is a promising strategy for enthesis repair. Engineering of complex tissue interphases such as the enthesis is likely to require a combination of biophysical, biological and chemical cues to achieve functional tissue regeneration. In this study, we cultured human primary adipose-derived mesenchymal stem cells (AdMCs) on biphasic silk fibroin scaffolds with integrated anisotropic (tendon/ligament-like) and isotropic (bone/cartilage like) pore alignment. We functionalized those scaffolds with heparin and explored their ability to deliver transforming growth factor β2 (TGF-β2) and growth/differentiation factor 5 (GDF5). Heparin functionalization increased the amount of TGF-β2 and GDF5 remaining attached to the scaffold matrix and resulted in biological effects at low growth factor doses. We analyzed the combined impact of pore alignment and growth factors on AdMSCs. TGF-β2 and pore anisotropy synergistically increased the expression of tendon/ligament markers and collagen I protein content. In addition, the combined delivery of TGF-β2 and GDF5 enhanced the expression of cartilage markers and collagen II protein content on substrates with isotropic porosity, whereas enthesis markers were enhanced in areas of mixed anisotropic/isotropic porosity. Altogether, the data obtained in this study improves current understanding on the combined effects of biological and structural cues on stem cell fate and presents a promising strategy for tendon/ligament-to-bone regeneration. Regeneration of the tendon/ligament-to-bone interphase (enthesis) is of significance in the repair of ruptured tendons/ligaments to bone to improve implant integration and clinical outcome. This study proposes a novel approach for enthesis regeneration based on a biomimetic and integrated tendon/ligament-to-bone construct, stem cells and heparin-based delivery of growth factors. We show that heparin can keep growth factors local and biologically active at low doses, which is critical to avoid supraphysiological doses and associated side effects. In addition, we identify synergistic effects of biological (growth factors) and structural (pore alignment) cues on stem cells. These results improve current understanding on the combined impact of biological and structural cues on the multi-lineage differentiation capacity of stem cells for regenerating complex tissue interphases. Copyright © 2018 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

Possible Mechanism for Denervation Effect on Wound Healing

DTIC Science & Technology

1989-05-17

under investigation is the regenerating limb of the axolotl , in which growth is strictly dependent on unknown factors from peripheral nerves. The...hypothesis that nerves contribute transferrin to cells of the regenerating tissues. Before experiments of this nature can be undertaken, axolotl transferrin...other tissues from axolotls . During the first year of this project, transferrin was purified from axolotls and antisera against it were generated in

[Tumor cells transfer between the patient and laboratory animal as a basic methodological approach to the study of cancerogenesis and identification of biomarkers].

PubMed

Klos, D; Stašek, M; Loveček, M; Skalický, P; Vrba, R; Aujeský, R; Havlík, R; Neoral, Č; Varanashi, L; Hajdúch, M; Vrbková, J; Džubák, P

The investigation of prognostic and predictive factors for early diagnosis of tumors, their surveillance and monitoring of the impact of therapeutic modalities using hybrid laboratory models in vitro/in vivo is an experimental approach with a significant potential. It is preconditioned by the preparation of in vivo tumor models, which may face a number of potential technical difficulties. The assessment of technical success of grafting and xenotransplantation based on the type of the tumor or cell line is important for the preparation of these models and their further use for proteomic and genomic analyses. Surgically harvested gastrointestinal tract tumor tissue was processed or stable cancer cell lines were cultivated; the viability was assessed, and subsequently the cells were inoculated subcutaneously to SCID mice with an individual duration of tumor growth, followed by its extraction. We analysed 140 specimens of tumor tissue including 17 specimens of esophageal cancer (viability 13/successful inoculations 0), 13 tumors of the cardia (11/0), 39 gastric tumors (24/4), 47 pancreatic tumors (34/1) and 24 specimens of colorectal cancer (22/9). 3 specimens were excluded due to histological absence of the tumor (complete remission after neoadjuvant therapy in 2 cases of esophageal carcinoma, 1 case of chronic pancreatitis). We observed successful inoculation in 17 of 28 tumor cell lines. The probability of successful grafting to the mice model in tumors of the esophagus, stomach and pancreas is significantly lower in comparison with colorectal carcinoma and cell lines generated tumors. The success rate is enhanced upon preservation of viability of the harvested tumor tissue, which depends on the sequence of clinical and laboratory algorithms with a high level of cooperation.Key words: proteomic analysis - xenotransplantation - prognostic and predictive factors - gastrointestinal tract tumors.

Mechanisms of fenthion activation in rainbow trout (Oncorhynchus mykiss) acclimated to hypersaline environments

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lavado, Ramon; Rimoldi, John M.; Schlenk, Daniel

2009-03-01

Previous studies in rainbow trout have shown that acclimation to hypersaline environments enhances the toxicity to thioether organophosphate and carbamate pesticides. In order to determine the role of biotransformation in this process, the metabolism of the thioether organophosphate biocide, fenthion was evaluated in microsomes from gills, liver and olfactory tissues in rainbow trout (Oncorhynchus mykiss) acclimated to freshwater and 17 per mille salinity. Hypersalinity acclimation increased the formation of fenoxon and fenoxon sulfoxide from fenthion in liver microsomes from rainbow trout, but not in gills or in olfactory tissues. NADPH-dependent and independent hydrolysis was observed in all tissues, but onlymore » NADPH-dependent fenthion cleavage was differentially modulated by hypersalinity in liver (inhibited) and gills (induced). Enantiomers of fenthion sulfoxide (65% and 35% R- and S-fenthion sulfoxide, respectively) were formed in liver and gills. The predominant pathway of fenthion activation in freshwater appears to be initiated through initial formation of fenoxon which may be subsequently converted to the most toxic metabolite fenoxon R-sulfoxide. However, in hypersaline conditions both fenoxon and fenthion sulfoxide formation may precede fenoxon sulfoxide formation. Stereochemical evaluation of sulfoxide formation, cytochrome P450 inhibition studies with ketoconazole and immunoblots indicated that CYP3A27 was primarily involved in the enhancement of fenthion activation in hypersaline-acclimated fish with limited contribution of FMO to initial sulfoxidation.« less

Simultaneous expression of tissue factor and tissue factor pathway inhibitor by human monocytes. A potential mechanism for localized control of blood coagulation

PubMed Central

1994-01-01

Cells of monocytic lineage can initiate extravascular fibrin deposition via expression of blood coagulation mediators. This report is about experiments on three mechanisms with the potential to modulate monocyte- initiated coagulation. Monocyte procoagulant activity was examined as a function of lipid cofactor, protein cofactor, and specific inhibitor expression during short-term culture in vitro. Lipid cofactor activity was measured as the initial rate of factor X activation by intrinsic- pathway components, the assembly of which depends on this cofactor. Lipid cofactor activity levels changed by < 30% during 48-h culture. Protein cofactor, i.e., tissue factor (TF) antigen was measured by enzyme immunoassay. It increased from 461 pg/ml to a maximum value of 3,550 pg/ml at 24 h and remained at 70% of this value. Specific TF activity, measured as factor VII-dependent factor X activation rate, decreased from 54 to 18 nM FXa/min between 24 and 48 h. TF activity did not correlate well with either lipid cofactor or TF protein levels. In contrast, the decrease in TF activity coincided in time with maximal expression of tissue factor pathway inhibitor (TFPI) mRNA, which was determined using reverse transcriptase polymerase chain reaction (RT- PCR), and with maximal TFPI protein levels measured by immunoassay. The number of mRNA copies coding for TFPI and TF in freshly isolated blood monocytes were 46 and 20 copies/cells, respectively. These values increased to 220 and 63 copies/cell during short-term cell culture in the presence of endotoxin. Results demonstrate concomitant expression by monocytes of genes coding for both the essential protein cofactor and the specific inhibitor of the extrinsic coagulation pathway. Together with functional and antigenic analyses, they also imply that the initiation of blood clotting by extravascular monocyte/macrophages can be modulated locally by TFPI independently of plasma sources of the inhibitor. PMID:8195712

Francisella DnaK Inhibits Tissue-nonspecific Alkaline Phosphatase*

PubMed Central

Arulanandam, Bernard P.; Chetty, Senthilnath Lakshmana; Yu, Jieh-Juen; Leonard, Sean; Klose, Karl; Seshu, Janakiram; Cap, Andrew; Valdes, James J.; Chambers, James P.

2012-01-01

Following pulmonary infection with Francisella tularensis, we observed an unexpected but significant reduction of alkaline phosphatase, an enzyme normally up-regulated following inflammation. However, no reduction was observed in mice infected with a closely related Gram-negative pneumonic organism (Klebsiella pneumoniae) suggesting the inhibition may be Francisella-specific. In similar fashion to in vivo observations, addition of Francisella lysate to exogenous alkaline phosphatase (tissue-nonspecific isozyme) was inhibitory. Partial purification and subsequent proteomic analysis indicated the inhibitory factor to be the heat shock protein DnaK. Incubation with increasing amounts of anti-DnaK antibody reduced the inhibitory effect in a dose-dependent manner. Furthermore, DnaK contains an adenosine triphosphate binding domain at its N terminus, and addition of adenosine triphosphate enhances dissociation of DnaK with its target protein, e.g. alkaline phosphatase. Addition of adenosine triphosphate resulted in decreased DnaK co-immunoprecipitated with alkaline phosphatase as well as reduction of Francisella-mediated alkaline phosphatase inhibition further supporting the binding of Francisella DnaK to alkaline phosphatase. Release of DnaK via secretion and/or bacterial cell lysis into the extracellular milieu and inhibition of plasma alkaline phosphatase could promote an orchestrated, inflammatory response advantageous to Francisella. PMID:22923614

Francisella DnaK inhibits tissue-nonspecific alkaline phosphatase.

PubMed

Arulanandam, Bernard P; Chetty, Senthilnath Lakshmana; Yu, Jieh-Juen; Leonard, Sean; Klose, Karl; Seshu, Janakiram; Cap, Andrew; Valdes, James J; Chambers, James P

2012-10-26

Following pulmonary infection with Francisella tularensis, we observed an unexpected but significant reduction of alkaline phosphatase, an enzyme normally up-regulated following inflammation. However, no reduction was observed in mice infected with a closely related gram-negative pneumonic organism (Klebsiella pneumoniae) suggesting the inhibition may be Francisella-specific. In similar fashion to in vivo observations, addition of Francisella lysate to exogenous alkaline phosphatase (tissue-nonspecific isozyme) was inhibitory. Partial purification and subsequent proteomic analysis indicated the inhibitory factor to be the heat shock protein DnaK. Incubation with increasing amounts of anti-DnaK antibody reduced the inhibitory effect in a dose-dependent manner. Furthermore, DnaK contains an adenosine triphosphate binding domain at its N terminus, and addition of adenosine triphosphate enhances dissociation of DnaK with its target protein, e.g. alkaline phosphatase. Addition of adenosine triphosphate resulted in decreased DnaK co-immunoprecipitated with alkaline phosphatase as well as reduction of Francisella-mediated alkaline phosphatase inhibition further supporting the binding of Francisella DnaK to alkaline phosphatase. Release of DnaK via secretion and/or bacterial cell lysis into the extracellular milieu and inhibition of plasma alkaline phosphatase could promote an orchestrated, inflammatory response advantageous to Francisella.

Myogenic transcription factors regulate pro-metastatic miR-182.

PubMed

Dodd, R D; Sachdeva, M; Mito, J K; Eward, W C; Brigman, B E; Ma, Y; Dodd, L; Kim, Y; Lev, D; Kirsch, D G

2016-04-07

Approximately 30% of patients with soft-tissue sarcoma die from pulmonary metastases. The mechanisms that drive sarcoma metastasis are not well understood. Recently, we identified miR-182 as a driver of sarcoma metastasis in a primary mouse model of soft-tissue sarcoma. We also observed elevated miR-182 in a subset of primary human sarcomas that metastasized to the lungs. Here, we show that myogenic differentiation factors regulate miR-182 levels to contribute to metastasis in mouse models. We find that MyoD directly binds the miR-182 promoter to increase miR-182 expression. Furthermore, mechanistic studies revealed that Pax7 can promote sarcoma metastasis in vivo through MyoD-dependent regulation of pro-metastatic miR-182. Taken together, these results suggest that sarcoma metastasis can be partially controlled through Pax7/MyoD-dependent activation of miR-182 and provide insight into the role that myogenic transcription factors have in sarcoma progression.

Autophagy enhances NFκB activity in specific tissue macrophages by sequestering A20 to boost antifungal immunity

PubMed Central

Kanayama, Masashi; Inoue, Makoto; Danzaki, Keiko; Hammer, Gianna; He, You-Wen; Shinohara, Mari L.

2014-01-01

Immune responses must be well restrained in a steady state to avoid excessive inflammation. However, such restraints are quickly removed to exert anti-microbial responses. Here, we report a role of autophagy in an early host anti-fungal response by enhancing NFκB activity through A20 sequestration. Enhancement of NFκB activation is achieved by autophagic depletion of A20, an NFκB inhibitor, in F4/80hi macrophages in the spleen, peritoneum, and kidney. We show that p62, an autophagic adaptor protein, captures A20 to sequester it in the autophagosome. This allows the macrophages to release chemokines to recruit neutrophils. Indeed, mice lacking autophagy in myeloid cells show higher susceptibility to Candida albicans infection due to impairment in neutrophil recruitment. Thus, at least in the specific aforementioned tissues, autophagy appears to break A20-dependent suppression in F4/80hi macrophages, which express abundant A20 and contribute to the initiation of efficient innate immune responses. PMID:25609235

Strategies for Controlled Delivery of Growth Factors and Cells for Bone Regeneration

PubMed Central

Vo, Tiffany N.; Kasper, F. Kurtis; Mikos, Antonios G.

2012-01-01

The controlled delivery of growth factors and cells within biomaterial carriers can enhance and accelerate functional bone formation. The carrier system can be designed with preprogrammed release kinetics to deliver bioactive molecules in a localized, spatiotemporal manner most similar to the natural wound healing process. The carrier can also act as an extracellular matrix-mimicking substrate for promoting osteoprogenitor cellular infiltration and proliferation for integrative tissue repair. This review discusses the role of various regenerative factors involved in bone healing and their appropriate combinations with different delivery systems for augmenting bone regeneration. The general requirements of protein, cell and gene therapy are described, with elaboration on how the selection of materials, configurations and processing affects growth factor and cell delivery and regenerative efficacy in both in vitro and in vivo applications for bone tissue engineering. PMID:22342771

Dose- and tissue-specific interaction of monoterpenes with the gibberellin-mediated release of potato tuber bud dormancy, sprout growth and induction of α-amylases and β-amylases.

PubMed

Rentzsch, Sonja; Podzimska, Dagmara; Voegele, Antje; Imbeck, Madeleine; Müller, Kerstin; Linkies, Ada; Leubner-Metzger, Gerhard

2012-01-01

Gibberellins (GA) are involved in bud dormancy release in several species. We show here that GA-treatment released bud dormancy, initiated bud sprouting and promoted sprout growth of excised potato tuber bud discs ('eyes'). Monoterpenes from peppermint oil (PMO) and S-(+)-carvone (CAR) interact with the GA-mediated bud dormancy release in a hormesis-type response: low monoterpene concentrations enhance dormancy release and the initiation of bud sprouting, whereas high concentrations inhibit it. PMO and CAR did, however, not affect sprout growth rate after its onset. We further show that GA-induced dormancy release is associated with tissue-specific regulation of α- and β-amylases. Molecular phylogenetic analysis shows that potato α-amylases cluster into two distinct groups: α-AMY1 and α-AMY2. GA-treatment induced transcript accumulation of members of both α-amylase groups, as well as α- and β-amylase enzyme activity in sprout and 'sub-eye' tissues. In sprouts, CAR interacts with the GA-mediated accumulation of α-amylase transcripts in an α-AMY2-specific and dose-dependent manner. Low CAR concentrations enhance the accumulation of α-AMY2-type α-amylase transcripts, but do not affect the α-AMY1-type transcripts. Low CAR concentrations also enhance the accumulation of α- and β-amylase enzyme activity in sprouts, but not in 'sub-eye' tissues. In contrast, high CAR concentrations have no appreciable effect in sprouts on the enzyme activities and the α-amylase transcript abundances of either group. The dose-dependent effects on the enzyme activities and the α-AMY2-type α-amylase transcripts in sprouts are specific for CAR but not for PMO. Different monoterpenes therefore may have specific targets for their interaction with hormone signalling pathways.

Towards an Optimal Gradient-dependent Energy Functional of the PZ-SIC Form

DOE PAGES

Jónsson, Elvar Örn; Lehtola, Susi; Jónsson, Hannes

2015-06-01

Results of Perdew–Zunger self-interaction corrected (PZ-SIC) density functional theory calculations of the atomization energy of 35 molecules are compared to those of high-level quantum chemistry calculations. While the PBE functional, which is commonly used in calculations of condensed matter, is known to predict on average too high atomization energy (overbinding of the molecules), the application of PZ-SIC gives a large overcorrection and leads to significant underestimation of the atomization energy. The exchange enhancement factor that is optimal for the generalized gradient approximation within the Kohn-Sham (KS) approach may not be optimal for the self-interaction corrected functional. The PBEsol functional, wheremore » the exchange enhancement factor was optimized for solids, gives poor results for molecules in KS but turns out to work better than PBE in PZ-SIC calculations. The exchange enhancement is weaker in PBEsol and the functional is closer to the local density approximation. Furthermore, the drop in the exchange enhancement factor for increasing reduced gradient in the PW91 functional gives more accurate results than the plateaued enhancement in the PBE functional. A step towards an optimal exchange enhancement factor for a gradient dependent functional of the PZ-SIC form is taken by constructing an exchange enhancement factor that mimics PBEsol for small values of the reduced gradient, and PW91 for large values. The average atomization energy is then in closer agreement with the high-level quantum chemistry calculations, but the variance is still large, the F 2 molecule being a notable outlier.« less

Signal enhancement ratio (SER) quantified from breast DCE-MRI and breast cancer risk

NASA Astrophysics Data System (ADS)

Wu, Shandong; Kurland, Brenda F.; Berg, Wendie A.; Zuley, Margarita L.; Jankowitz, Rachel C.; Sumkin, Jules; Gur, David

2015-03-01

Breast magnetic resonance imaging (MRI) is recommended as an adjunct to mammography for women who are considered at elevated risk of developing breast cancer. As a key component of breast MRI, dynamic contrast-enhanced MRI (DCE-MRI) uses a contrast agent to provide high intensity contrast between breast tissues, making it sensitive to tissue composition and vascularity. Breast DCE-MRI characterizes certain physiologic properties of breast tissue that are potentially related to breast cancer risk. Studies have shown that increased background parenchymal enhancement (BPE), which is the contrast enhancement occurring in normal cancer-unaffected breast tissues in post-contrast sequences, predicts increased breast cancer risk. Signal enhancement ratio (SER) computed from pre-contrast and post-contrast sequences in DCE-MRI measures change in signal intensity due to contrast uptake over time and is a measure of contrast enhancement kinetics. SER quantified in breast tumor has been shown potential as a biomarker for characterizing tumor response to treatments. In this work we investigated the relationship between quantitative measures of SER and breast cancer risk. A pilot retrospective case-control study was performed using a cohort of 102 women, consisting of 51 women who had diagnosed with unilateral breast cancer and 51 matched controls (by age and MRI date) with a unilateral biopsy-proven benign lesion. SER was quantified using fully-automated computerized algorithms and three SER-derived quantitative volume measures were compared between the cancer cases and controls using logistic regression analysis. Our preliminary results showed that SER is associated with breast cancer risk, after adjustment for the Breast Imaging Reporting and Data System (BI-RADS)-based mammographic breast density measures. This pilot study indicated that SER has potential for use as a risk factor for breast cancer risk assessment in women at elevated risk of developing breast cancer.

Capsaicin induces browning of white adipose tissue and counters obesity by activating TRPV1 channel‐dependent mechanisms

PubMed Central

Baskaran, Padmamalini; Krishnan, Vivek; Ren, Jun

2016-01-01

Background and Purpose The growing epidemic of obesity and metabolic diseases necessitates the development of novel strategies to prevent and treat such diseases. Current research suggests that browning of white adipose tissue (WAT) promotes energy expenditure to counter obesity. Recent research suggests that activation of the TRPV1 channels counters obesity. However, the mechanism by which activation of TRPV1 channels counters obesity still remains unclear. Experimental Approach We evaluated the effect of dietary capsaicin to induce a browning program in WAT by activating TRPV1 channels to prevent diet‐induced obesity using wild‐type and TRPV1−/− mouse models. We performed experiments using preadipocytes and fat pads from these mice. Key Results Capsaicin stimulated the expression of brown fat‐specific thermogenic uncoupling protein‐1 and bone morphogenetic protein‐8b in WAT. Capsaicin triggered browning of WAT by promoting sirtuin‐1 expression and activity via TRPV1 channel‐dependent elevation of intracellular Ca2 + and phosphorylation of Ca2 +/calmodulin‐activated protein kinase II and AMP‐activated kinase. Capsaicin increased the expression of PPARγ 1 coactivator α and enhanced metabolic and ambulatory activity. Further, capsaicin stimulated sirtuin‐1‐dependent deacetylation of PPARγ and the transcription factor PRDM‐16 and facilitated PPARγ–PRDM‐16 interaction to induce browning of WAT. Dietary capsaicin did not protect TRPV1−/− mice from obesity. Conclusions and Interpretations Our results show for the first time that activation of TRPV1 channels by dietary capsaicin triggers browning of WAT to counteract obesity. Our results suggest that activation of TRPV1 channels is a promising strategy to counter obesity. PMID:27174467

Different wound healing properties of dermis, adipose, and gingiva mesenchymal stromal cells.

PubMed

Boink, Mireille A; van den Broek, Lenie J; Roffel, Sanne; Nazmi, Kamran; Bolscher, Jan G M; Gefen, Amit; Veerman, Enno C I; Gibbs, Susan

2016-01-01

Oral wounds heal faster and with better scar quality than skin wounds. Deep skin wounds where adipose tissue is exposed, have a greater risk of forming hypertrophic scars. Differences in wound healing and final scar quality might be related to differences in mesenchymal stromal cells (MSC) and their ability to respond to intrinsic (autocrine) and extrinsic signals, such as human salivary histatin, epidermal growth factor, and transforming growth factor beta1. Dermis-, adipose-, and gingiva-derived MSC were compared for their regenerative potential with regards to proliferation, migration, and matrix contraction. Proliferation was assessed by cell counting and migration using a scratch wound assay. Matrix contraction and alpha smooth muscle actin was assessed in MSC populated collagen gels, and also in skin and gingival full thickness tissue engineered equivalents (reconstructed epithelium on MSC populated matrix). Compared to skin-derived MSC, gingiva MSC showed greater proliferation and migration capacity, and less matrix contraction in full thickness tissue equivalents, which may partly explain the superior oral wound healing. Epidermal keratinocytes were required for enhanced adipose MSC matrix contraction and alpha smooth muscle actin expression, and may therefore contribute to adverse scarring in deep cutaneous wounds. Histatin enhanced migration without influencing proliferation or matrix contraction in all three MSC, indicating that salivary peptides may have a beneficial effect on wound closure in general. Transforming growth factor beta1 enhanced contraction and alpha smooth muscle actin expression in all three MSC types when incorporated into collagen gels. Understanding the mechanisms responsible for the superior oral wound healing will aid us to develop advanced strategies for optimal skin regeneration, wound healing and scar formation. © 2015 by the Wound Healing Society.

Dimensionality and noise in energy selective x-ray imaging

PubMed Central

Alvarez, Robert E.

2013-01-01

Purpose: To develop and test a method to quantify the effect of dimensionality on the noise in energy selective x-ray imaging. Methods: The Cramèr-Rao lower bound (CRLB), a universal lower limit of the covariance of any unbiased estimator, is used to quantify the noise. It is shown that increasing dimensionality always increases, or at best leaves the same, the variance. An analytic formula for the increase in variance in an energy selective x-ray system is derived. The formula is used to gain insight into the dependence of the increase in variance on the properties of the additional basis functions, the measurement noise covariance, and the source spectrum. The formula is also used with computer simulations to quantify the dependence of the additional variance on these factors. Simulated images of an object with three materials are used to demonstrate the trade-off of increased information with dimensionality and noise. The images are computed from energy selective data with a maximum likelihood estimator. Results: The increase in variance depends most importantly on the dimension and on the properties of the additional basis functions. With the attenuation coefficients of cortical bone, soft tissue, and adipose tissue as the basis functions, the increase in variance of the bone component from two to three dimensions is 1.4 × 103. With the soft tissue component, it is 2.7 × 104. If the attenuation coefficient of a high atomic number contrast agent is used as the third basis function, there is only a slight increase in the variance from two to three basis functions, 1.03 and 7.4 for the bone and soft tissue components, respectively. The changes in spectrum shape with beam hardening also have a substantial effect. They increase the variance by a factor of approximately 200 for the bone component and 220 for the soft tissue component as the soft tissue object thickness increases from 1 to 30 cm. Decreasing the energy resolution of the detectors increases the variance of the bone component markedly with three dimension processing, approximately a factor of 25 as the resolution decreases from 100 to 3 bins. The increase with two dimension processing for adipose tissue is a factor of two and with the contrast agent as the third material for two or three dimensions is also a factor of two for both components. The simulated images show that a maximum likelihood estimator can be used to process energy selective x-ray data to produce images with noise close to the CRLB. Conclusions: The method presented can be used to compute the effects of the object attenuation coefficients and the x-ray system properties on the relationship of dimensionality and noise in energy selective x-ray imaging systems. PMID:24320442

Anti-Tumor Effect of Steamed Codonopsis lanceolata in H22 Tumor-Bearing Mice and Its Possible Mechanism

PubMed Central

Li, Wei; Xu, Qi; He, Yu-Fang; Liu, Ying; Yang, Shu-Bao; Wang, Zi; Zhang, Jing; Zhao, Li-Chun

2015-01-01

Although previous studies confirmed that steaming and the fermentation process could significantly improve the cognitive-enhancement and neuroprotective effects of Codonopsis lanceolata, the anti-tumor efficacy of steamed C. lanceolata (SCL) and what mechanisms are involved remain largely unknown. The present study was designed to evaluate the anti-tumor effect in vivo of SCL in H22 tumor-bearing mice. The results clearly indicated that SCL could not only inhibit the tumor growth, but also prolong the survival time of H22 tumor-bearing mice. Besides, the serum levels of cytokines, such as interferon gamma (IFN-γ), tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6) and interleukin-2 (IL-2), were enhanced by SCL administration. The observations of Hoechst 33258 staining demonstrated that SCL was able to induce tumor cell apoptosis. Finally, immunohistochemical analysis revealed that SCL treatment significantly increased Bax expression and decreased Bcl-2 and vascular endothelial growth factor (VEGF) expression of H22 tumor tissues in a dose-dependent manner. Moreover, LC/MS analysis of SCL indicated that it mainly contained lobetyolin and six saponins. Taken all together, the findings in the present study clearly demonstrated that SCL inhibited the H22 tumor growth in vivo at least partly via improving the immune functions, inducing apoptosis and inhibiting angiogenesis. PMID:26426041

Calreticulin Enhances Porcine Wound Repair by Diverse Biological Effects

PubMed Central

Nanney, Lillian B.; Woodrell, Christopher D.; Greives, Mathew R.; Cardwell, Nancy L.; Pollins, Alonda C.; Bancroft, Tara A.; Chesser, Adrianne; Michalak, Marek; Rahman, Mohammad; Siebert, John W.; Gold, Leslie I.

2008-01-01

Extracellular functions of the endoplasmic reticulum chaperone protein calreticulin (CRT) are emerging. Here we show novel roles for exogenous CRT in both cutaneous wound healing and diverse processes associated with repair. Compared with platelet-derived growth factor-BB-treated controls, topical application of CRT to porcine excisional wounds enhanced the rate of wound re-epithelialization. In both normal and steroid-impaired pigs, CRT increased granulation tissue formation. Immunohistochemical analyses of the wounds 5 and 10 days after injury revealed marked up-regulation of transforming growth factor-β3 (a key regulator of wound healing), a threefold increase in macrophage influx, and an increase in the cellular proliferation of basal keratinocytes of the new epidermis and of cells of the neodermis. In vitro studies confirmed that CRT induced a greater than twofold increase in the cellular proliferation of primary human keratinocytes, fibroblasts, and microvascular endothelial cells (with 100 pg/ml, 100 ng/ml, and 1.0 pg/ml, respectively). Moreover, using a scratch plate assay, CRT maximally induced the cellular migration of keratinocytes and fibroblasts (with 10 pg/ml and 1 ng/ml, respectively). In addition, CRT induced concentration-dependent migration of keratinocytes, fibroblasts macrophages, and monocytes in chamber assays. These in vitro bioactivities provide mechanistic support for the positive biological effects of CRT observed on both the epidermis and dermis of wounds in vivo, underscoring a significant role for CRT in the repair of cutaneous wounds. PMID:18753412

Lineage-specific enhancers activate self-renewal genes in macrophages and embryonic stem cells.

PubMed

Soucie, Erinn L; Weng, Ziming; Geirsdóttir, Laufey; Molawi, Kaaweh; Maurizio, Julien; Fenouil, Romain; Mossadegh-Keller, Noushine; Gimenez, Gregory; VanHille, Laurent; Beniazza, Meryam; Favret, Jeremy; Berruyer, Carole; Perrin, Pierre; Hacohen, Nir; Andrau, J-C; Ferrier, Pierre; Dubreuil, Patrice; Sidow, Arend; Sieweke, Michael H

2016-02-12

Differentiated macrophages can self-renew in tissues and expand long term in culture, but the gene regulatory mechanisms that accomplish self-renewal in the differentiated state have remained unknown. Here we show that in mice, the transcription factors MafB and c-Maf repress a macrophage-specific enhancer repertoire associated with a gene network that controls self-renewal. Single-cell analysis revealed that, in vivo, proliferating resident macrophages can access this network by transient down-regulation of Maf transcription factors. The network also controls embryonic stem cell self-renewal but is associated with distinct embryonic stem cell-specific enhancers. This indicates that distinct lineage-specific enhancer platforms regulate a shared network of genes that control self-renewal potential in both stem and mature cells. Copyright © 2016, American Association for the Advancement of Science.

Platelet-Rich Plasma

PubMed Central

Cole, Brian J.; Seroyer, Shane T.; Filardo, Giuseppe; Bajaj, Sarvottam; Fortier, Lisa A.

2010-01-01

Context: Platelet-rich plasma (PRP) may affect soft tissue healing via growth factors released after platelet degranulation. Because of this potential benefit, clinicians have begun to inject PRP for the treatment of tendon, ligament, muscle, and cartilage injuries and early osteoarthritis. Evidence Acquisition: A PubMed search was performed for studies relating to PRP, growth factors, and soft tissue injuries from 1990 to 2010. Relevant references from these studies were also retrieved. Results: Soft tissue injury is a major source of disability that may often be complicated by prolonged and incomplete recovery. Numerous growth factors may potentiate the healing and regeneration of tendons and ligaments. The potential benefits of biologically enhanced healing processes have led to a recent interest in the use of PRP in orthopaedic sports medicine. There has been widespread anecdotal use of PRP for muscle strains, tendinopathy, and ligament injuries and as a surgical adjuvant to rotator cuff repair, anterior cruciate ligament reconstruction, and meniscal or labral repairs. Although the fascination with this emerging technology has led to a dramatic increase in its use, scientific data supporting this use are still in their infancy. Conclusions: The literature is replete with studies on the basic science of growth factors and their relation to the maintenance, proliferation, and regeneration of various tissues and tissue-derived cells. Despite the promising results of several animal studies, well-controlled human studies are lacking. PMID:23015939

Tissue-Engineered Nanofibrous Nerve Grafts for Enhancing the Rate of Nerve Regeneration

DTIC Science & Technology

2015-10-01

structured nanofibrous biodegradable nerve graft system that present ECM protein, neurotrophic factor, and pre-seeded with bone marrow stromal cells in...nanofibrous biodegradable nerve graft system that present extracellular matrix (ECM) protein, nerve growth factor, and pre-seeded with bone marrow stromal...proposed novel structured nanofibrous biodegradable grafts will provide the micro environment, bioactivity, transport features and mechanics ideal for

Proteomic and transcriptomic investigation of acne vulgaris microcystic and papular lesions: Insights in the understanding of its pathophysiology.

PubMed

Quanico, Jusal; Gimeno, Jean-Pascal; Nadal-Wollbold, Florence; Casas, Christiane; Alvarez-Georges, Sandrine; Redoulès, Daniel; Schmitt, Anne-Marie; Fournier, Isabelle; Salzet, Michel

2017-03-01

The pathogenesis of acne vulgaris involves several phases including androgen-dependent hyper-seborrhea, colonization by Propionibacterium acnes, and inflammation. Recent investigations have shown that in fact P. acnes provokes the activation of the inflammasome present in macrophages and dendritic cells. This signaling pathway leads to excessive production of interleukin IL-1β, a proinflammatory cytokine. Nevertheless, these well-studied phenomena in acne fail to elucidate the mechanisms responsible for the appearance of different lesions. We investigate response pathways for specific acne lesions such as microcysts and papules using shot-gun proteomic followed by systemic biology and transcriptomic approaches. Results show that most of the proteins identified as differentially expressed between the normal and acne tissue biopsies associated with the immune system response were identified as highly or exclusively expressed in the papule biopsies. They were also expressed in microcysts, but in lower amounts compared to those in papules. These results are supported by the identification of CAMP factor protein produced by P. acnes in microcysts, indicating its enhanced proliferation in this type of lesion CONCLUSIONS: As CAMP factor protein was not detected in papule biopsies, we can see a clear delineation in the stages of progression of acne pathogenesis, which begins with a hyphenated inflammatory response in the papule stage, followed by imbalance of lipid production, which in turn triggers the enhanced proliferation of P. acnes. We demonstrate that expression inflammation varies across the two types of lesions, suggesting different pathways enhanced as a function of the progression of P. acnes. Copyright © 2016 Elsevier B.V. All rights reserved.

NF-Y coassociates with FOS at promoters, enhancers, repetitive elements, and inactive chromatin regions, and is stereo-positioned with growth-controlling transcription factors

PubMed Central

Fleming, Joseph D.; Pavesi, Giulio; Benatti, Paolo; Imbriano, Carol; Mantovani, Roberto; Struhl, Kevin

2013-01-01

NF-Y, a trimeric transcription factor (TF) composed of two histone-like subunits (NF-YB and NF-YC) and a sequence-specific subunit (NF-YA), binds to the CCAAT motif, a common promoter element. Genome-wide mapping reveals 5000–15,000 NF-Y binding sites depending on the cell type, with the NF-YA and NF-YB subunits binding asymmetrically with respect to the CCAAT motif. Despite being characterized as a proximal promoter TF, only 25% of NF-Y sites map to promoters. A comparable number of NF-Y sites are located at enhancers, many of which are tissue specific, and nearly half of the NF-Y sites are in select subclasses of HERV LTR repeats. Unlike most TFs, NF-Y can access its target DNA motif in inactive (nonmodified) or polycomb-repressed chromatin domains. Unexpectedly, NF-Y extensively colocalizes with FOS in all genomic contexts, and this often occurs in the absence of JUN and the AP-1 motif. NF-Y also coassociates with a select cluster of growth-controlling and oncogenic TFs, consistent with the abundance of CCAAT motifs in the promoters of genes overexpressed in cancer. Interestingly, NF-Y and several growth-controlling TFs bind in a stereo-specific manner, suggesting a mechanism for cooperative action at promoters and enhancers. Our results indicate that NF-Y is not merely a commonly used proximal promoter TF, but rather performs a more diverse set of biological functions, many of which are likely to involve coassociation with FOS. PMID:23595228

Enhanced Inflammation without Impairment of Insulin Signaling in the Visceral Adipose Tissue of 5α-Dihydrotestosterone-Induced Animal Model of Polycystic Ovary Syndrome.

PubMed

Milutinović, Danijela Vojnović; Nikolić, Marina; Veličković, Nataša; Djordjevic, Ana; Bursać, Biljana; Nestorov, Jelena; Teofilović, Ana; Antić, Ivana Božić; Macut, Jelica Bjekić; Zidane, Abdulbaset Shirif; Matić, Gordana; Macut, Djuro

2017-09-01

Polycystic ovary syndrome is a heterogeneous endocrine and metabolic disorder associated with abdominal obesity, dyslipidemia and insulin resistance. Since abdominal obesity is characterized by low-grade inflammation, the aim of the study was to investigate whether visceral adipose tissue inflammation linked to abdominal obesity and dyslipidemia could lead to impaired insulin sensitivity in the animal model of polycystic ovary syndrome.Female Wistar rats were treated with nonaromatizable 5α-dihydrotestosterone pellets in order to induce reproductive and metabolic characteristics of polycystic ovary syndrome. Glucose, triglycerides, non-esterified fatty acids and insulin were determined in blood plasma. Visceral adipose tissue inflammation was evaluated by the nuclear factor kappa B intracellular distribution, macrophage migration inhibitory factor protein level, as well as TNFα, IL6 and IL1β mRNA levels. Insulin sensitivity was assessed by intraperitoneal glucose tolerance test and homeostasis model assessment index, and through analysis of insulin signaling pathway in the visceral adipose tissue.Dihydrotestosterone treatment led to increased body weight, abdominal obesity and elevated triglycerides and non-esterified fatty acids, which were accompanied by the activation of nuclear factor kappa B and increase in macrophage migration inhibitory factor, IL6 and IL1β levels in the visceral adipose tissue. In parallel, insulin sensitivity was affected in 5α-dihydrotestosterone-treated animals only at the systemic and not at the level of visceral adipose tissue.The results showed that abdominal obesity and dyslipidemia in the animal model of polycystic ovary syndrome were accompanied with low-grade inflammation in the visceral adipose tissue. However, these metabolic disturbances did not result in decreased tissue insulin sensitivity. © Georg Thieme Verlag KG Stuttgart · New York.

[Study of changes in the enzyme-salt composition affecting the permeability of ocular tissues under infrasound phonophoresis].

PubMed

Filatov, V V

2005-01-01

This paper deals with the study of infrasound phonophoresis-induced changes in biochemical factors, which affect the permeability of eyeball tissues. During 10 days, the rabbit right eye was exposed to an infrasound in the changing pressure mode at 4 Hz and 173 dB for 10 minutes every day. The left eye remained control. After finishing a series of studies, the animals were slaughtered, the eyes were enucleated and prepared into individual tissues. Changes in sodium-potassium composition were investigated in the first series. By causing a reduction in the cellular content of K+, infrasound exposure was found to cause a decrease in membranous potential and activation Na-channel, as confirmed by the elevated intracellular levels of Na+. This in turn enhances ocular tissue permeability for drugs without damaging the structure of a cell membrane. Changes in the activity of the following enzymes: beta-glucosidase, cathepsin D, and hy- aluronidase. Infrasound was ascertained to enhance the activity of beta-glucosidase, which accounts for the lower levels of glucose in ocular tissues and points to the activation and acceleration of biochemical processes in the tissues. At the same time the increased concentrations of cathepsin D and hyaluronidase found in ocular tissues were responsible for a temporary reduction in the viscosity of hyaluronic acid, which promotes resolution of opacities, adhesions or scars and increased tissue permeability.

Measurements of the linewidth enhancement factor of mid-infrared quantum cascade lasers by different optical feedback techniques

DOE Office of Scientific and Technical Information (OSTI.GOV)

Jumpertz, L., E-mail: louise.jumpertz@telecom-paristech.fr; MirSense, 8 avenue de la Vauve, F-91120 Palaiseau; Michel, F.

2016-01-15

Precise knowledge of the linewidth enhancement factor of a semiconductor laser under actual operating conditions is of prime importance since this parameter dictates various phenomena such as linewidth broadening or optical nonlinearities enhancement. The above-threshold linewidth enhancement factor of a mid-infrared quantum cascade laser structure operated at 10{sup ∘}C is determined experimentally using two different methods based on optical feedback. Both Fabry-Perot and distributed feedback quantum cascade lasers based on the same active area design are studied, the former by following the wavelength shift as a function of the feedback strength and the latter by self-mixing interferometry. The results aremore » consistent and unveil a clear pump current dependence of the linewidth enhancement factor, with values ranging from 0.8 to about 3.« less

Ethylene-enhanced catabolism of ( sup 14 C)indole-3-acetic acid to indole-3-carboxylic acid in citrus leaf tissues. [Citrus sinensis

DOE Office of Scientific and Technical Information (OSTI.GOV)

Sagee, O.; Riov, J.; Goren, J.

Exogenous ({sup 14}C)indole-3-acetic acid (IAA) is conjugated in citrus (Citrus sinensis) leaf tissues to one major substance which has been identified as indole-3-acetylaspartic acid (IAAsp). Ethylene pretreatment enhanced the catabolism of ({sup 14}C)IAA to indole-3-carboxylic acid (ICA), which accumulated as glucose esters (ICGlu). Increased formation of ICGlu by ethylene was accompanied by a concomitant decrease in IAAsp formation. IAAsp and ICGlu were identified by combined gas chromatography-mass spectrometry. Formation of ICGlu was dependent on the concentration of ethylene and the duration of the ethylene pretreatment. It is suggested that the catabolism of IAA to ICA may be one of themore » mechanisms by which ethylene endogenous IAA levels.« less

Regulation of the Anaphase-promoting Complex–Separase Cascade by Transforming Growth Factor-β Modulates Mitotic Progression in Bone Marrow Stromal Cells

PubMed Central

Fujita, Takeo; Epperly, Michael W.; Zou, Hui; Greenberger, Joel S.

2008-01-01

Alteration of the tumor microenvironment by aberrant stromal cells influences many aspects of cell biology, including differentiation of stem cells and tumor metastasis. The role of transforming growth factor (TGF)-β signaling in stromal cells of the tissue microenvironment is critical to both pathways. We examined murine marrow stromal cells with deletion of Smad3 and found that they have an altered cell cycle profile, with a higher fraction of cells in G2/M phase. Deletion of Smad3 significantly abrogates TGF-β signaling and suppresses phosphorylation of CDC27–anaphase-promoting complex (APC) during mitosis, thereby resulting in elevated cyclin-dependent kinase (CDK)1 activity via increased levels of cyclin B. Enhanced CDK1 activity due to deregulation of APC leads in turn to hyperphosphorylation of separase, impeding chromatid separation. A residue Ser1126Ala mutation in separase specifically abolished separase hyperphosphorylation in Smad3-deficient cells. The present results unveil a new function for the TGF-β pathway in the regulation of APC to mediate chromatid separation during mitosis. PMID:18843049

Merlin negative regulation by miR-146a promotes cell transformation

DOE Office of Scientific and Technical Information (OSTI.GOV)

Pérez-García, Erick I.; Meza-Sosa, Karla F.; López-Sevilla, Yaxem

2015-12-25

Inactivation of the tumor suppressor Merlin, by deleterious mutations or by protein degradation via sustained growth factor receptor signaling-mediated mechanisms, results in cell transformation and tumor development. In addition to these mechanisms, here we show that, miRNA-dependent negative regulation of Merlin protein levels also promotes cell transformation. We provide experimental evidences showing that miR-146a negatively regulates Merlin protein levels through its interaction with an evolutionary conserved sequence in the 3´ untranslated region of the NF2 mRNA. Merlin downregulation by miR-146a in A549 lung epithelial cells resulted in enhanced cell proliferation, migration and tissue invasion. Accordingly, stable miR-146a-transfectant cells formed tumorsmore » with metastatic capacity in vivo. Together our results uncover miRNAs as yet another negative mechanism controlling Merlin tumor suppressor functions.« less

The Crosstalk of RAS with the TGF-β Family During Carcinoma Progression and its Implications for Targeted Cancer Therapy

PubMed Central

Grusch, M.; Petz, M.; Metzner, T.; Öztürk, D.; Schneller, D.; Mikulits, W.

2010-01-01

Both RAS and transforming growth factor (TGF)-β signaling cascades are central in tumorigenesis and show synergisms depending on tumor stage and tissue context. In this review we focus on the interaction of RAS subeffector proteins with signaling components of the TGF-β family including those of TGF-βs, activins and bone morphogenic proteins. Compelling evidence indicates that RAS signaling is essentially involved in the switch from tumor-suppressive to tumor-promoting functions of the TGF-β family leading to enhanced cancer growth and metastatic dissemination of primary tumors. Thus, the interface of these signaling cascades is considered as a promising target for the development of novel cancer therapeutics. The current pharmacological anti-cancer concepts combating the molecular cooperation between RAS and TGF-β family signaling during carcinoma progression are critically discussed. PMID:20718708

Non-thermal dielectric barrier discharge plasma induces angiogenesis through reactive oxygen species.

PubMed

Arjunan, Krishna P; Clyne, Alisa Morss

2011-01-01

Vascularization plays a key role in processes such as wound healing and tissue engineering. Non-thermal plasma, which primarily produces reactive oxygen species (ROS), recently emerged as an efficient tool in medical applications. Liquids and endothelial cells were treated with a non-thermal dielectric barrier discharge plasma. Plasma treatment of phosphate buffered saline (PBS) and serum-free medium increased ROS concentration in a dose-dependent manner, with a higher concentration in serum-free medium. ROS concentration in cells peaked 1 hour after treatment. 4.2 J/cm(2) increased cell proliferation, 2D and 3D migration, as well as tube formation. A fibroblast growth factor-2 (FGF-2) neutralizing antibody and ROS scavengers for hydrogen peroxide and hydroxyl radicals abrogated these angiogenic effects. Non-thermal plasma may be a potential tool for applying ROS in precise doses to enhance vascularization.

Placenta Growth Factor in Diabetic Wound Healing

PubMed Central

Cianfarani, Francesca; Zambruno, Giovanna; Brogelli, Laura; Sera, Francesco; Lacal, Pedro Miguel; Pesce, Maurizio; Capogrossi, Maurizio C.; Failla, Cristina Maria; Napolitano, Monica; Odorisio, Teresa

2006-01-01

Reduced microcirculation and diminished expression of growth factors contribute to wound healing impairment in diabetes. Placenta growth factor (PlGF), an angiogenic mediator promoting pathophysiological neovascularization, is expressed during cutaneous wound healing and improves wound closure by enhancing angiogenesis. By using streptozotocin-induced diabetic mice, we here demonstrate that PlGF induction is strongly reduced in diabetic wounds. Diabetic transgenic mice overexpressing PlGF in the skin displayed accelerated wound closure compared with diabetic wild-type littermates. Moreover, diabetic wound treatment with an adenovirus vector expressing the human PlGF gene (AdCMV.PlGF) significantly accelerated the healing process compared with wounds treated with a control vector. The analysis of treated wounds showed that PlGF gene transfer improved granulation tissue formation, maturation, and vascularization, as well as monocytes/macrophages local recruitment. Platelet-derived growth factor, fibroblast growth factor-2, and vascular endothelial growth factor mRNA levels were increased in AdCMV.PlGF-treated wounds, possibly enhancing PlGF-mediated effects. Finally, PlGF treatment stimulated cultured dermal fibroblast migration, pointing to a direct role of PlGF in accelerating granulation tissue maturation. In conclusion, our data indicate that reduced PlGF expression contributes to impaired wound healing in diabetes and that PlGF gene transfer to diabetic wounds exerts therapeutic activity by promoting different aspects of the repair process. PMID:17003476

Notochordal cells in the adult intervertebral disc: new perspective on an old question.

PubMed

Risbud, Makarand V; Shapiro, Irving M

2011-01-01

The intervertebral disc is a tissue positioned between each of the vertebrae that accommodates applied biomechanical forces to the spine. The central compartment of the disc contains the nucleus pulposus (NP) which is enclosed by the annulus fibrosus and the endplate cartilage.The NP is derived from the notochord, a rod-like structure of mesodermal origin. Development of the notochord is tightly regulated by interactive transcription factors and target genes. Since a number of these molecules are unique they have be used for cell lineage and fate mapping studies of tissues of the intervertebral disc. These studies have shown that in a number of species including human, NP tissue retains notochordal cells throughout life. In the adult NP, there are present both large and small notochordal cells, as well as a progenitor cell population which can differentiate along the mesengenic pathway. Since tissue renewal in the intervertebral disc is dependent on the ability of these cells to commit to the NP lineage and undergo terminal differentiation, studies have been performed to assess which signaling pathways may regulate these activities. The notch signaling pathway is active in the intervertebral disc and is responsive to hypoxia, probably through HIF-1a. From a disease viewpoint, it is hypothesized that an oxemic shift, possibly mediated by alterations in the vascular supply to the tissues of the disc would be expected to lead to a failure in notochordal progenitor cell activation and a decrease in the number of differentiated cells. In turn, this would lead to decrements in function and enhancement of the effect of agents that are known to promote disc degeneration.

Response functions for computing absorbed dose to skeletal tissues from photon irradiation—an update

NASA Astrophysics Data System (ADS)

Johnson, Perry B.; Bahadori, Amir A.; Eckerman, Keith F.; Lee, Choonsik; Bolch, Wesley E.

2011-04-01

A comprehensive set of photon fluence-to-dose response functions (DRFs) is presented for two radiosensitive skeletal tissues—active and total shallow marrow—within 15 and 32 bone sites, respectively, of the ICRP reference adult male. The functions were developed using fractional skeletal masses and associated electron-absorbed fractions as reported for the UF hybrid adult male phantom, which in turn is based upon micro-CT images of trabecular spongiosa taken from a 40 year male cadaver. The new DRFs expand upon both the original set of seven functions produced in 1985, and a 2007 update calculated under the assumption of secondary electron escape from spongiosa. In this study, it is assumed that photon irradiation of the skeleton will yield charged particle equilibrium across all spongiosa regions at energies exceeding 200 keV. Kerma coefficients for active marrow, inactive marrow, trabecular bone and spongiosa at higher energies are calculated using the DRF algorithm setting the electron-absorbed fraction for self-irradiation to unity. By comparing kerma coefficients and DRF functions, dose enhancement factors and mass energy-absorption coefficient (MEAC) ratios for active marrow to spongiosa were derived. These MEAC ratios compared well with those provided by the NIST Physical Reference Data Library (mean difference of 0.8%), and the dose enhancement factors for active marrow compared favorably with values calculated in the well-known study published by King and Spiers (1985 Br. J. Radiol. 58 345-56) (mean absolute difference of 1.9 percentage points). Additionally, dose enhancement factors for active marrow were shown to correlate well with the shallow marrow volume fraction (R2 = 0.91). Dose enhancement factors for the total shallow marrow were also calculated for 32 bone sites representing the first such derivation for this target tissue.

Dual-energy contrast-enhanced digital mammography (DE-CEDM): optimization on digital subtraction with practical x-ray low/high-energy spectra

NASA Astrophysics Data System (ADS)

Chen, Biao; Jing, Zhenxue; Smith, Andrew P.; Parikh, Samir; Parisky, Yuri

2006-03-01

Dual-energy contrast enhanced digital mammography (DE-CEDM), which is based upon the digital subtraction of low/high-energy image pairs acquired before/after the administration of contrast agents, may provide physicians physiologic and morphologic information of breast lesions and help characterize their probability of malignancy. This paper proposes to use only one pair of post-contrast low / high-energy images to obtain digitally subtracted dual-energy contrast-enhanced images with an optimal weighting factor deduced from simulated characteristics of the imaging chain. Based upon our previous CEDM framework, quantitative characteristics of the materials and imaging components in the x-ray imaging chain, including x-ray tube (tungsten) spectrum, filters, breast tissues / lesions, contrast agents (non-ionized iodine solution), and selenium detector, were systemically modeled. Using the base-material (polyethylene-PMMA) decomposition method based on entrance low / high-energy x-ray spectra and breast thickness, the optimal weighting factor was calculated to cancel the contrast between fatty and glandular tissues while enhancing the contrast of iodized lesions. By contrast, previous work determined the optimal weighting factor through either a calibration step or through acquisition of a pre-contrast low/high-energy image pair. Computer simulations were conducted to determine weighting factors, lesions' contrast signal values, and dose levels as functions of x-ray techniques and breast thicknesses. Phantom and clinical feasibility studies were performed on a modified Selenia full field digital mammography system to verify the proposed method and computer-simulated results. The resultant conclusions from the computer simulations and phantom/clinical feasibility studies will be used in the upcoming clinical study.

The fibrinolytic mechanism of defibrotide: effect of defibrotide on plasmin activity.

PubMed

Echart, Cinara L; Graziadio, Barbara; Somaini, Simona; Ferro, Laura I; Richardson, Paul G; Fareed, Jawed; Iacobelli, Massimo

2009-12-01

Fibrinolytic activity has been shown to be reduced in many vascular diseases, including hepatic veno-occlusive disease after stem cell transplantation, a microangiopathy characterized by sinusoidal endothelial cell injury. Defibrotide is a polydisperse oligonucleotide with antithrombotic, profibrinolytic, anti-ischemic, and antiadhesive properties. Numerous clinical studies have shown promising activity of defibrotide in the treatment and prevention of veno-occlusive disease, with minimal toxicity. In corollary laboratory studies, defibrotide has been shown to decrease plasminogen activator inhibitor-1, increase tissue plasminogen activator levels, and increase overall plasma fibrinolytic activity in patients. Plasmin, a potent and nonspecific serine protease, plays a pivotal role in fibrinolysis by virtue of its ability to effectively degrade fibrin clots. In this study, defibrotide increases the activity of plasmin in hydrolyzing its substrate in a dose-dependent and length-dependent manner. Similar concentration-dependent effects of defibrotide were observed when plasmin was generated by tissue plasminogen activator or urokinase activation of plasminogen. In contrast, defibrotide had no direct effect on the activation of plasminogen to plasmin. Defibrotide was also able to enhance the activity of plasmin in degrading fibrin clot formed from fibrinogen, plasminogen, and thrombin. This effect was also concentration-dependent and directly correlated with the enzymatic activity of plasmin. This study therefore demonstrates that defibrotide is capable of enhancing the activity of plasmin and so contributes to its fibrinolytic activity. Taken together, these results support the effect of defibrotide in restoring the fibrinolytic vascular phenotype, in microangiopathic conditions such as veno-occlusive disease.

Mesenchymal Stem Cells From Bone Marrow, Adipose Tissue, and Lung Tissue Differentially Mitigate Lung and Distal Organ Damage in Experimental Acute Respiratory Distress Syndrome.

PubMed

Silva, Johnatas D; Lopes-Pacheco, Miquéias; Paz, Ana H R; Cruz, Fernanda F; Melo, Elga B; de Oliveira, Milena V; Xisto, Débora G; Capelozzi, Vera L; Morales, Marcelo M; Pelosi, Paolo; Cirne-Lima, Elizabeth; Rocco, Patricia R M

2018-02-01

Mesenchymal stem cells-based therapies have shown promising effects in experimental acute respiratory distress syndrome. Different mesenchymal stem cells sources may result in diverse effects in respiratory diseases; however, there is no information regarding the best source of mesenchymal stem cells to treat pulmonary acute respiratory distress syndrome. We tested the hypothesis that mesenchymal stem cells derived from bone marrow, adipose tissue, and lung tissue would lead to different beneficial effects on lung and distal organ damage in experimental pulmonary acute respiratory distress syndrome. Animal study and primary cell culture. Laboratory investigation. Seventy-five Wistar rats. Wistar rats received saline (control) or Escherichia coli lipopolysaccharide (acute respiratory distress syndrome) intratracheally. On day 2, acute respiratory distress syndrome animals were further randomized to receive saline or bone marrow, adipose tissue, or lung tissue mesenchymal stem cells (1 × 10 cells) IV. Lung mechanics, histology, and protein levels of inflammatory mediators and growth factors were analyzed 5 days after mesenchymal stem cells administration. RAW 264.7 cells (a macrophage cell line) were incubated with lipopolysaccharide followed by coculture or not with bone marrow, adipose tissue, and lung tissue mesenchymal stem cells (10 cells/mL medium). Regardless of mesenchymal stem cells source, cells administration improved lung function and reduced alveolar collapse, tissue cellularity, collagen, and elastic fiber content in lung tissue, as well as decreased apoptotic cell counts in liver. Bone marrow and adipose tissue mesenchymal stem cells administration also reduced levels of tumor necrosis factor-α, interleukin-1β, keratinocyte-derived chemokine, transforming growth factor-β, and vascular endothelial growth factor, as well as apoptotic cell counts in lung and kidney, while increasing expression of keratinocyte growth factor in lung tissue. Additionally, mesenchymal stem cells differently modulated the secretion of biomarkers by macrophages depending on their source. Mesenchymal stem cells from different sources led to variable responses in lungs and distal organs. Bone marrow and adipose tissue mesenchymal stem cells yielded greater beneficial effects than lung tissue mesenchymal stem cells. These findings may be regarded as promising in clinical trials.

The Effect of Insulin and Insulin-Like Growth Factors on Hippocampus- and Amygdala-Dependent Long-Term Memory Formation

ERIC Educational Resources Information Center

Stern, Sarah A.; Chen, Dillon Y.; Alberini, Cristina M.

2014-01-01

Recent work has reported that the insulin-like growth factor 2 (IGF2) promotes memory enhancement. Furthermore, impaired insulin or IGF1 functions have been suggested to play a role in the pathogenesis of neurodegeneration and cognitive impairments, hence implicating the insulin/IGF system as an important target for cognitive enhancement and/or…

Connective tissue growth factor enhances the migration of gastric cancer through downregulation of E-cadherin via the NF-κB pathway.

PubMed

Mao, Zhengfa; Ma, Xiaoyan; Rong, Yefei; Cui, Lei; Wang, Xuqing; Wu, Wenchuan; Zhang, Jianxin; Jin, Dayong

2011-01-01

Local invasion and distant metastasis are difficult problems for surgical intervention and treatment in gastric cancer. Connective tissue growth factor (CTGF/CCN2) was considered to have an important role in this process. In this study, we demonstrated that expression of CTGF was significantly upregulated in clinical tissue samples of gastric carcinoma (GC) samples. Forced expression of CTGF in AGS GC cells promoted their migration in culture and significantly increased tumor metastasis in nude mice, whereas RNA interference-mediated knockdown of CTGF in GC cells significantly inhibited cell migration in vitro. We disclose that CTGF downregulated the expression of E-cadherin through activation of the nuclear factor-κappa B (NF-κB) pathway. The effects of CTGF in GC cells were abolished by dominant negative IκappaB. Collectively, these data reported here demonstrate CTGF could modulate the NF-κappaB pathway and perhaps be a promising therapeutic target for gastric cancer invasion and metastasis. © 2010 Japanese Cancer Association.

Enhancing the Migration Ability of Mesenchymal Stromal Cells by Targeting the SDF-1/CXCR4 Axis

PubMed Central

Marquez-Curtis, Leah A.

2013-01-01

Mesenchymal stromal cells (MSCs) are currently being investigated in numerous clinical trials of tissue repair and various immunological disorders based on their ability to secrete trophic factors and to modulate inflammatory responses. MSCs have been shown to migrate to sites of injury and inflammation in response to soluble mediators including the chemokine stromal cell-derived factor-(SDF-)1, but during in vitro culture expansion MSCs lose surface expression of key homing receptors particularly of the SDF-1 receptor, CXCR4. Here we review studies on enhancement of SDF-1-directed migration of MSCs with the premise that their improved recruitment could translate to therapeutic benefits. We describe our studies on approaches to increase the CXCR4 expression in in vitro-expanded cord blood-derived MSCs, namely, transfection, using the commercial liposomal reagent IBAfect, chemical treatment with the histone deacetylase inhibitor valproic acid, and exposure to recombinant complement component C1q. These methodologies will be presented in the context of other cell targeting and delivery strategies that exploit pathways involved in MSC migration. Taken together, these findings indicate that MSCs can be manipulated in vitro to enhance their in vivo recruitment and efficacy for tissue repair. PMID:24381939

Estrogen-Dependent Proteolytic Cleavage of Semaphorin 4D and Plexin-B1 Enhances Semaphorin 4D-Induced Apoptosis during Postnatal Vaginal Remodeling in Pubescent Mice

PubMed Central

Yoshida, Kenji; Ueyama, Takashi; Miyajima, Masayasu; Negishi, Takayuki; Kawasaki, Takahiko; Takamatsu, Hyota; Kikutani, Hitoshi; Kumanogoh, Atsushi; Yukawa, Kazunori

2014-01-01

Around the fifth week after birth, the vaginal cavity in female mouse pups opens to the overlaying skin. This postnatal tissue remodeling of the genital tract occurs during puberty, and it largely depends upon hormonally induced apoptosis that mainly occurs in the epithelium at the lower part of the mouse vaginal cavity. Previously, we showed that most BALB/c mice lacking the class IV Semaphorin (Sema4D) develop imperforate vagina and hydrometrocolpos; therefore, we reasoned that the absence of Sema4D-induced apoptosis in vaginal epithelial cells may cause the imperforate vagina. Sema4D signals via the Plexin-B1 receptor; nevertheless detailed mechanisms mediating this hormonally triggered apoptosis are not fully documented. To investigate the estrogen-dependent control of Sema4D signaling during the apoptosis responsible for mouse vaginal opening, we examined structural and functional modulation of Sema4D, Plexin-B1, and signaling molecules by analyzing both wild-type and Sema4D−/− mice with or without ovariectomy. Both the release of soluble Sema4D and the conversion of Plexin-B1 by proteolytic processing in vaginal tissue peaked 5 weeks after birth of wild-type BALB/c mice at the time of vaginal opening. Estrogen supplementation of ovariectomized wild-type mice revealed that both the release of soluble Sema4D and the conversion of Plexin-B1 into an active form were estrogen-dependent and concordant with apoptosis. Estrogen supplementation of ovariectomized Sema4D−/− mice did not induce massive vaginal apoptosis in 5-week-old mice; therefore, Sema4D may be an essential apoptosis-inducing ligand that acts downstream of estrogen action in vaginal epithelium during this postnatal tissue remodeling. Analysis of ovariectomized mice also indicated that Sema4D contributed to estrogen-dependent dephosphorylation of Akt and ERK at the time of vaginal opening. Based on our results, we propose that apoptosis in vaginal epithelium during postnatal vaginal opening is induced by enhanced Sema4D signaling that is caused by estrogen-dependent structural changes of Sema4D and Plexin-B1. PMID:24841081

Oil biosynthesis in a basal angiosperm: transcriptome analysis of Persea Americana mesocarp.

PubMed

Kilaru, Aruna; Cao, Xia; Dabbs, Parker B; Sung, Ha-Jung; Rahman, Md Mahbubur; Thrower, Nicholas; Zynda, Greg; Podicheti, Ram; Ibarra-Laclette, Enrique; Herrera-Estrella, Luis; Mockaitis, Keithanne; Ohlrogge, John B

2015-08-16

The mechanism by which plants synthesize and store high amounts of triacylglycerols (TAG) in tissues other than seeds is not well understood. The comprehension of controls for carbon partitioning and oil accumulation in nonseed tissues is essential to generate oil-rich biomass in perennial bioenergy crops. Persea americana (avocado), a basal angiosperm with unique features that are ancestral to most flowering plants, stores ~ 70 % TAG per dry weight in its mesocarp, a nonseed tissue. Transcriptome analyses of select pathways, from generation of pyruvate and leading up to TAG accumulation, in mesocarp tissues of avocado was conducted and compared with that of oil-rich monocot (oil palm) and dicot (rapeseed and castor) tissues to identify tissue- and species-specific regulation and biosynthesis of TAG in plants. RNA-Seq analyses of select lipid metabolic pathways of avocado mesocarp revealed patterns similar to that of other oil-rich species. However, only some predominant orthologs of the fatty acid biosynthetic pathway genes in this basal angiosperm were similar to those of monocots and dicots. The accumulation of TAG, rich in oleic acid, was associated with higher transcript levels for a putative stearoyl-ACP desaturase and endoplasmic reticulum (ER)-associated acyl-CoA synthetases, during fruit development. Gene expression levels for enzymes involved in terminal steps to TAG biosynthesis in the ER further indicated that both acyl-CoA-dependent and -independent mechanisms might play a role in TAG assembly, depending on the developmental stage of the fruit. Furthermore, in addition to the expression of an ortholog of WRINKLED1 (WRI1), a regulator of fatty acid biosynthesis, high transcript levels for WRI2-like and WRI3-like suggest a role for additional transcription factors in nonseed oil accumulation. Plastid pyruvate necessary for fatty acid synthesis is likely driven by the upregulation of genes involved in glycolysis and transport of its intermediates. Together, a comparative transcriptome analyses for storage oil biosynthesis in diverse plants and tissues suggested that several distinct and conserved features in this basal angiosperm species might contribute towards its rich TAG content. Our work represents a comprehensive transcriptome resource for a basal angiosperm species and provides insight into their lipid metabolism in mesocarp tissues. Furthermore, comparison of the transcriptome of oil-rich mesocarp of avocado, with oil-rich seed and nonseed tissues of monocot and dicot species, revealed lipid gene orthologs that are highly conserved during evolution. The orthologs that are distinctively expressed in oil-rich mesocarp tissues of this basal angiosperm, such as WRI2, ER-associated acyl-CoA synthetases, and lipid-droplet associated proteins were also identified. This study provides a foundation for future investigations to increase oil-content and has implications for metabolic engineering to enhance storage oil content in nonseed tissues of diverse species.