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Sample records for factor confers stromal

  1. Immune suppressor factor confers stromal cell line with enhanced supporting activity for hematopoietic stem cells

    SciTech Connect

    Nakajima, Hideaki . E-mail: hnakajim@ims.u-tokyo.ac.jp; Shibata, Fumi; Fukuchi, Yumi; Goto-Koshino, Yuko; Ito, Miyuki; Urano, Atsushi; Nakahata, Tatsutoshi; Aburatani, Hiroyuki; Kitamura, Toshio

    2006-02-03

    Immune suppressor factor (ISF) is a subunit of the vacuolar ATPase proton pump. We earlier identified a short form of ISF (ShIF) as a stroma-derived factor that supports cytokine-independent growth of mutant Ba/F3 cells. Here, we report that ISF/ShIF supports self-renewal and expansion of primary hematopoietic stem cells (HSCs). Co-culture of murine bone marrow cells with a stromal cell line overexpressing ISF or ShIF (MS10/ISF or MS10/ShIF) not only enhanced their colony-forming activity and the numbers of long-term culture initiating cells, but also maintained the competitive repopulating activity of HSC. This stem cell supporting activity depended on the proton-transfer function of ISF/ShIF. Gene expression analysis of ISF/ShIF-transfected cell lines revealed down-regulation of secreted frizzled-related protein-1 and tissue inhibitor of metalloproteinase-3, and the restoration of their expressions in MS10/ISF cells partially reversed its enhanced LTC-IC supporting activity to a normal level. These results suggest that ISF/ShIF confers stromal cells with enhanced supporting activities for HSCs by modulating Wnt-activity and the extracellular matrix.

  2. Human placental eXpanded (PLX) mesenchymal-like adherent stromal cells confer neuroprotection to nerve growth factor (NGF)-differentiated PC12 cells exposed to ischemia by secretion of IL-6 and VEGF.

    PubMed

    Lahiani, Adi; Zahavi, Efrat; Netzer, Nir; Ofir, Racheli; Pinzur, Lena; Raveh, Shani; Arien-Zakay, Hadar; Yavin, Ephraim; Lazarovici, Philip

    2015-02-01

    Mesenchymal stem cells are potent candidates in stroke therapy due to their ability to secrete protective anti-inflammatory cytokines and growth factors. We investigated the neuroprotective effects of human placental mesenchymal-like adherent stromal cells (PLX) using an established ischemic model of nerve growth factor (NGF)-differentiated pheochromocytoma PC12 cells exposed to oxygen and glucose deprivation (OGD) followed by reperfusion. Under optimal conditions, 2 × 10⁵ PLX cells, added in a trans-well system, conferred 30-60% neuroprotection to PC12 cells subjected to ischemic insult. PC12 cell death, measured by LDH release, was reduced by PLX cells or by conditioned medium derived from PLX cells exposed to ischemia, suggesting the active release of factorial components. Since neuroprotection is a prominent function of the cytokine IL-6 and the angiogenic factor VEGF165, we measured their secretion using selective ELISA of the cells under ischemic or normoxic conditions. IL-6 and VEGF165 secretion by co-culture of PC12 and PLX cells was significantly higher under ischemic compared to normoxic conditions. Exogenous supplementation of 10 ng/ml each of IL-6 and VEGF165 to insulted PC12 cells conferred neuroprotection, reminiscent of the neuroprotective effect of PLX cells or their conditioned medium. Growth factors as well as co-culture conditioned medium effects were reduced by 70% and 20% upon pretreatment with 240 ng/ml Semaxanib (anti VEGF165) and/or 400 ng/ml neutralizing anti IL-6 antibody, respectively. Therefore, PLX-induced neuroprotection in ischemic PC12 cells may be partially explained by IL-6 and VEGF165 secretion. These findings may also account for the therapeutic effects seen in clinical trials after treatment with these cells.

  3. Stromal cell derived factor-1 (SDF-1) targeting reperfusion reduces myocardial infarction in isolated rat hearts.

    PubMed

    Jang, Young-Ho; Kim, June-Hong; Ban, Changill; Ahn, Kyohan; Cheong, Jae-Hun; Kim, Hyung-Hoi; Kim, Jung-Soo; Park, Yong-Hyun; Kim, Jun; Chun, Kook-Jin; Lee, Gyeong-Ho; Kim, Miju; Kim, Cheolmin; Xu, Zhelong

    2012-10-01

    Recent studies have shown that stromal cell derived factor-1 (SDF-1), first known as a cytokine involved in recruiting stem cells into injured organs, confers myocardial protection in myocardial infarction, which is not dependent on stem cell recruitment but related with modulation of ischemia-reperfusion (I/R) injury. However, the effect of SDF has been studied only in a preischemic exposure model, which is not clinically relevant if SDF is to be used as a therapeutic agent. Our study was aimed at evaluating whether or not SDF-1 confers cardioprotection during the reperfusion period. Hearts from SD rats were isolated and perfused with the Langendorff system. Proximal left coronary artery ligation, reperfusion, and SDF perfusion in KH buffer was done according to study protocol. Area of necrosis (AN) relative to area at risk (AR) was the primary endpoint of the study. Significant reduction of AN/AR by SDF in an almost dose-dependent manner was noted during both the preischemic exposure and reperfusion periods. In particular, infusion of a high concentration of SDF (25 nM/L) resulted in a dramatic reduction of infarct size, which was greater than that achieved with ischemic pre- or postconditioning. SDF perfusion during reperfusion was associated with a similar significant reduction of infarct size as preischemic SDF exposure. Further studies are warranted to assess the potential of SDF as a therapeutic agent for reducing I/R injury in clinical practice.

  4. Factors affecting mesenchymal stromal cells yield from bone marrow aspiration.

    PubMed

    Li, Jing; Wong, Wilfred Hing-Sang; Chan, Shing; Chim, James Chor-San; Cheung, Kenneth Man-Chee; Lee, Tsz-Leung; Au, Wing-Yan; Ha, Shau-Yin; Lie, Albert Kwok-Wei; Lau, Yu-Lung; Liang, Raymond Hin-Suen; Chan, Godfrey Chi-Fung

    2011-03-01

    This study was to investigate the variables in bone marrow harvesting procedure and individual donor factors which can potentially affect the yield of mesenchymal stromal cells (MSC). WE DETERMINED THE YIELD OF MSC FROM BONE MARROW UNDER DIFFERENT CLINICAL CONDITIONS BY COMPARING THE MSC COLONY NUMBERS FROM: (1) donors of different ages; (2) healthy donors and patients with leukemia; (3) bone marrow aspirated at different time points during marrow harvesting; (4) bone marrow harvested by different needles. During the process of harvesting, the number of MSC significantly decreased with increase number of aspiration, from 675/ml at the initial decreased to 60/ml after 100 ml bone marrow aspirated, and 50/ml after 200 ml bone marrow aspirated. The number of MSC retrieved from leukemia patients (99/ml bone marrow) was significantly lower than that of healthy donors (708/ml bone marrow). However, there was no significant difference in growth rate. There was no significant age-related difference of MSC yielded from donors <55 years. And there was no significant difference in MSC number between the samples from single end-holed needle and those from multiple-side-hole needle. The optimal bone marrow samples for MSC collection should be obtained earlier in the process of harvesting procedure. Bone marrow from donors <55 years was equally good as MSC sources. The autologous MSC from leukemia patients can be utilized for in-vitro MSC expansion.

  5. Patient factors influencing the concentration of stromal vascular fraction (SVF) for adipose-derived stromal cell (ASC) therapy in dogs.

    PubMed

    Astor, Donniel E; Hoelzler, Michael G; Harman, Robert; Bastian, Richard P

    2013-07-01

    The objective of this study was to determine whether patient factors influence the concentration of the stromal vascular fraction (SVF) in fat for adipose-derived stromal cell (ASC) therapy in dogs. A total of 1265 dogs underwent adipose collection surgeries by veterinarians for processing by the Vet-Stem laboratory and data on cell counts and patient factors were collected. Body condition score (BCS) and breed size did not significantly affect the viable cells per gram (VCPG) of adipose tissue that represents the viable SVF. Age significantly affected the VCPG, with dogs in age quartile 1 having a significantly higher VCPG than those in quartile 2 (P = 0.003) and quartile 4 (P = 0.002). Adipose tissue collected at the falciform location had significantly fewer VCPG than tissue collected at the thoracic wall and inguinal locations (P < 0.001). When the interaction of gender and location was evaluated, there were significantly fewer VCPG in tissue collected at the falciform location than at the thoracic wall and inguinal locations in female spayed dogs (P < 0.001) and male neutered dogs (P < 0.001), but not in female intact dogs (P = 0.743) or male intact dogs (P = 0.208). It was concluded that specific patient factors should be taken into consideration in order to obtain the maximal yield of VCPG from an adipose collection procedure.

  6. Multiple CD11c+ cells collaboratively express IL-1β to modulate stromal vascular endothelial growth factor and lymph node vascular-stromal growth.

    PubMed

    Benahmed, Fairouz; Chyou, Susan; Dasoveanu, Dragos; Chen, Jingfeng; Kumar, Varsha; Iwakura, Yoichiro; Lu, Theresa T

    2014-05-01

    Lymphadenopathy in autoimmune and other lymphoproliferative diseases is in part characterized by immunoblasts and vascular proliferation. The lymph node vasculature, along with the nonvascular stromal compartment, supports lymphocyte function, and targeting vascular-stromal expansion in inflamed nodes may modulate lymphocyte function in disease. CD11c(+) cells are essential for vascular-stromal proliferation and the upregulation of vascular endothelial growth factor (VEGF) needed for vascular proliferation. However, targetable CD11c(+) cell-derived molecular mediators, the identity of relevant CD11c(+) cells, and whether CD11c(+) cells directly stimulate VEGF-expressing stromal cells are poorly understood. In this study we show that CD11c(+) CD11b(+) CCR2-dependent monocytes and CCR7-dependent dendritic cells express IL-1β. IL-1β blockade, IL-1β deficiency in radiosensitive cells, and CCR2/CCR7 double deficiency but not single deficiency all attenuate immunization-induced vascular-stromal proliferation. gp38(+) stromal fibroblastic reticular cells (FRCs) that express VEGF are enriched for Thy1(+) cells and partially overlap with CCL21-expressing FRCs, and FRC VEGF is attenuated with IL-1β deficiency or blockade. IL-1β localizes to the outer borders of the T zone, where VEGF-expressing cells are also enriched. Ex vivo, CD11b(+) cells enriched for IL-1β(+) cells can directly induce cultured gp38(+)Thy1(+) FRCs to upregulate VEGF. Taken together, these results suggest a mechanism whereby multiple recruited CD11c(+) populations express IL-1β and directly modulate FRC function to help promote the initiation of vascular-stromal growth in stimulated lymph nodes. These data provide new insight into how CD11c(+) cells regulate the lymph node vascular-stromal compartment, add to the evolving understanding of functional stromal subsets, and suggest a possible utility for IL-1β blockade in preventing inflammatory lymph node growth.

  7. Prostaglandin E2 regulates macrophage colony stimulating factor secretion by human bone marrow stromal cells.

    PubMed

    Besse, A; Trimoreau, F; Faucher, J L; Praloran, V; Denizot, Y

    1999-07-08

    Bone marrow stromal cells regulate marrow haematopoiesis by secreting growth factors such as macrophage colony stimulating factor (M-CSF) that regulates the proliferation, differentiation and several functions of cells of the mononuclear-phagocytic lineage. By using a specific ELISA we found that their constitutive secretion of M-CSF is enhanced by tumour necrosis factor-alpha (TNF-alpha). The lipid mediator prostaglandin E2 (PGE2) markedly reduces in a time- and dose-dependent manner the constitutive and TNF-alpha-induced M-CSF synthesis by bone marrow stromal cells. In contrast, other lipid mediators such as 12-HETE, 15-HETE, leukotriene B4, leukotriene C4 and lipoxin A4 have no effect. EP2/EP4 selective agonists (11-deoxy PGE1 and 1-OH PGE1) and EP2 agonist (19-OH PGE2) inhibit M-CSF synthesis by bone marrow stromal cells while an EP1/EP3 agonist (sulprostone) has no effect. Stimulation with PGE2 induces an increase of intracellular cAMP levels in bone marrow stromal cells. cAMP elevating agents (forskolin and cholera toxin) mimic the PGE2-induced inhibition of M-CSF production. In conclusion, PGE2 is a potent regulator of M-CSF production by human bone marrow stromal cells, its effects being mediated via cAMP and PGE receptor EP2/EP4 subtypes.

  8. Modulation of Mammary Stromal Cell Lactate Dynamics by Ambient Glucose and Epithelial Factors.

    PubMed

    Tobar, Nicolas; Porras, Omar; Smith, Patricio C; Barros, L Felipe; Martínez, Jorge

    2017-01-01

    Hyperglycemia is a risk factor for a variety of human cancers. Increased access to glucose and that tumor metabolize glucose by a glycolytic process even in the presence of oxygen (Warburg effect), provide a framework to analyze a particular set of metabolic adaptation mechanisms that may explain this phenomenon. In the present work, using a mammary stromal cell line derived from healthy tissue that was subjected to a long-term culture in low (5 mM) or high (25 mM) glucose, we analyzed kinetic parameters of lactate transport using a FRET biosensor. Our results indicate that the glucose pre-culture and soluble epithelial factors constitute a stimulus for lactate stromal production, factors that also modify the kinetic parameters and the monocarboxylate transporters expression in stromal cells. We also observed a vectorial flux of lactate from stroma to epithelial cells in a co-culture setting and found that the uptake of lactate by epithelial cells correlates with the degree of malignancy. Glucose preconditioning of the stromal cell stimulated epithelial motility. Our findings suggest that lactate generated by stromal cells in the high glucose condition stimulate epithelial migration. Overall, our results support the notion that glucose not only provides a substrate for tumor nutrition but also behaves as a signal promoting malignancy. J. Cell. Physiol. 232: 136-144, 2017. © 2016 Wiley Periodicals, Inc.

  9. Ultrastructural localization of stem cell factor in canine marrow-derived stromal cells.

    PubMed

    Huss, R; Hong, D S; Beckham, C; Kimball, L; Myerson, D H; Storb, R; Deeg, H J

    1995-01-01

    Stromal cell lines derived from canine long-term bone marrow cultures (LTBMC) were characterized regarding the expression of growth factors and especially the localization of stem cell factor (SCF) (c-kit ligand). One cell line (DO64) was immortalized by transformation with a retroviral vector containing the open reading frames (ORFs) E6 and E7 of the human papilloma virus type 16 (HPV-16). Transfection did not change cellular characteristics but rendered the cell line more independent from culture conditions. The transformed line DO64 consisted mainly of fibroblast-like cells. In addition, some cells showed endothelial and some smooth-muscle cell features. Stromal cells expressed a broad spectrum of surface markers, including low levels of major histocompatibility-complex (MHC) class-II antigens. A new murine monoclonal antibody (MAb), RG7.6 (IgG1), specific for canine SCF, recognized the majority of fibroblast-like stromal cells. The staining pattern for SCF showed perinuclear and intracytoplasmic dense areas. Immunoelectron microscopy revealed the localization of SCF in secretory vesicles, the perivesicular cytoplasm, and bound to the cytoplasmatic membrane. RNA analysis showed that stromal cells transcribed, in addition to SCF, messages for granulocyte colony-stimulating factor (G-CSF), granulocyte-monocyte CSF (GM-CSF), interleukin-6 (IL-6), and transforming growth factor-beta (TGF-beta). In summary, we have established and characterized canine marrow-derived stromal cell lines, and using the new MAb RG7.6, we have localized SCF to cytoplasmatic vesicles as well as the membrane of stromal cells.

  10. Identification of Factors Produced and Secreted by Mesenchymal Stromal Cells with the SILAC Method.

    PubMed

    Rocha, Beatriz; Calamia, Valentina; Blanco, Francisco J; Ruiz-Romero, Cristina

    2016-01-01

    Mesenchymal stromal cells (MSCs) secrete a large variety of proteins and factors, which shape the secretome. These proteins participate in multiple cellular functions, including the promotion of regenerative processes in the damaged tissue. Secretomes derived from either undifferentiated MSCs or these cells undergoing osteogenic, chondrogenic, or adipogenic differentiation have been characterized using different liquid chromatography tandem mass spectrometry (LC-MS/MS)-based quantitative proteomic approaches. In this chapter, we describe the use of the Stable Isotope Labeling by Amino Acids in Cell culture (SILAC) strategy for the identification and relative quantification of the mesenchymal stromal cell secretome, specifically during chondrogenesis.

  11. Stromal-epithelial interaction study: The effect of corneal epithelial cells on growth factor expression in stromal cells using organotypic culture model.

    PubMed

    Kobayashi, Takeshi; Shiraishi, Atsushi; Hara, Yuko; Kadota, Yuko; Yang, Lujun; Inoue, Tomoyuki; Shirakata, Yuji; Ohashi, Yuichi

    2015-06-01

    Interactions between stromal and epithelial cells play important roles in the development, homeostasis, and pathological conditions of the cornea. Soluble cytokines are critical factors in stromal-epithelial interactions, and growth factors secreted from corneal stromal cells contribute to the regulation of proliferation and differentiation of corneal epithelial cells (CECs). However, the manner in which the expression of growth factors is regulated in stromal cells has not been completely determined. To study stromal-epithelial cell interactions, we used an organotypic culture model. Human or rabbit CECs (HCECs or RCECs) were cultured on amniotic membranes placed on human corneal fibroblasts (HCFs) embedded in a collagen gel. The properties of the organotypic culture were examined by hematoxylin-eosin staining and immunofluorescence. In the organotypic culture, HCECs or RCECs were stratified into two-three layers after five days and five-seven layers after nine days. However, stratification was not observed when the HCECs were seeded on a collagen gel without fibroblasts. K3/K12 were expressed on day 9. The HCF-embedded collagen gels were collected on days 3, 5, or 9 after seeding the RCECs, and mRNA expression of growth factors FGF7, HGF, NGF, EGF, TGF-α, SCF, TGF-β1, TGF-β2, and TGF-β3 were quantified by real-time PCR. mRNA expression of the growth factors in HCFs cultured with RCECs were compared with those cultured without RCECs, as well as in monolayer cultures. mRNA expression of TGF-α was markedly increased in HCFs cultured with RCECs. However, mRNA expression of the TGF-β family was suppressed in HCFs cultured with RCECs. Principal component analysis revealed that mRNA expression of the growth factors in HCFs were generally similar when they were cultured with RCECs. In organotypic cultures, the morphological changes in the CECs and the expression patterns of the growth factors in the stromal cells clearly demonstrated stromal-epithelial cell

  12. Neuromodulatory loop mediated by nerve growth factor and interleukin 6 in thymic stromal cell cultures.

    PubMed Central

    Screpanti, I; Meco, D; Scarpa, S; Morrone, S; Frati, L; Gulino, A; Modesti, A

    1992-01-01

    Neural crest cell derivatives have been suggested to be involved in thymus development. We established nonlymphoid thymic stromal cell cultures capable of supporting T-cell differentiation. In these nonlymphoid cell cultures, we identified cells with phenotypic and biochemical markers specific for neuronal cells. Neurofilament mRNA and 68- and 160-kDa neurofilament proteins, as well as 74-kDa synapsin I isoform, were expressed in many of the cultured cells. For example, neurofilament immunoreactivity was detected in 20-30% of the cells. To see whether thymic neuronal-like cells were involved in a neural differentiation pathway, we investigated the effect of nerve growth factor (NGF) and interleukin 6 (IL-6), two known neurotrophic factors. The expression of the above-described neural markers was enhanced by NGF and IL-6, which we report to be produced in an autocrine way by thymic stromal cell cultures. Finally, we found that IL-6 gene expression in these cell cultures was enhanced by NGF. Evidence is thus offered of a neuromodulatory loop within the thymic stromal cell population supported by local production of NGF and IL-6 and involving neural cell elements. Interestingly, IL-6, which is known to be implicated in thymocyte differentiation, also displays a neuromodulatory activity on thymic stromal cells, suggesting a multivalent role for this cytokine within the thymus. Images PMID:1373490

  13. Bioengineered implantable scaffolds as a tool to study stromal-derived factors in metastatic cancer models.

    PubMed

    Bersani, Francesca; Lee, Jungwoo; Yu, Min; Morris, Robert; Desai, Rushil; Ramaswamy, Sridhar; Toner, Mehmet; Haber, Daniel A; Parekkadan, Biju

    2014-12-15

    Modeling the hematogenous spread of cancer cells to distant organs poses one of the greatest challenges in the study of human metastasis. Both tumor cell-intrinsic properties as well as interactions with reactive stromal cells contribute to this process, but identification of relevant stromal signals has been hampered by the lack of models allowing characterization of the metastatic niche. Here, we describe an implantable bioengineered scaffold, amenable to in vivo imaging, ex vivo manipulation, and serial transplantation for the continuous study of human metastasis in mice. Orthotopic or systemic inoculation of tagged human cancer cells into the mouse leads to the release of circulating tumor cells into the vasculature, which seed the scaffold, initiating a metastatic tumor focus. Mouse stromal cells can be readily recovered and profiled, revealing differential expression of cytokines, such as IL1β, from tumor-bearing versus unseeded scaffolds. Finally, this platform can be used to test the effect of drugs on suppressing initiation of metastatic lesions. This generalizable model to study cancer metastasis may thus identify key stromal-derived factors with important implications for basic and translational cancer research.

  14. Epigenetic Alterations Affecting Transcription Factors and Signaling Pathways in Stromal Cells of Endometriosis

    PubMed Central

    Yotova, Iveta; Hsu, Emily; Do, Catherine; Gaba, Aulona; Sczabolcs, Matthias; Dekan, Sabine; Kenner, Lukas; Wenzl, Rene; Tycko, Benjamin

    2017-01-01

    Endometriosis is characterized by growth of endometrial-like tissue outside the uterine cavity. Since its pathogenesis may involve epigenetic changes, we used Illumina 450K Methylation Beadchips to profile CpG methylation in endometriosis stromal cells compared to stromal cells from normal endometrium. We validated and extended the Beadchip data using bisulfite sequencing (bis-seq), and analyzed differential methylation (DM) at the CpG-level and by an element-level classification for groups of CpGs in chromatin domains. Genes found to have DM included examples encoding transporters (SLC22A23), signaling components (BDNF, DAPK1, ROR1, and WNT5A) and transcription factors (GATA family, HAND2, HOXA cluster, NR5A1, OSR2, TBX3). Intriguingly, among the TF genes with DM we also found JAZF1, a proto-oncogene affected by chromosomal translocations in endometrial stromal tumors. Using RNA-Seq we identified a subset of the DM genes showing differential expression (DE), with the likelihood of DE increasing with the extent of the DM and its location in enhancer elements. Supporting functional relevance, treatment of stromal cells with the hypomethylating drug 5aza-dC led to activation of DAPK1 and SLC22A23 and repression of HAND2, JAZF1, OSR2, and ROR1 mRNA expression. We found that global 5hmC is decreased in endometriotic versus normal epithelial but not stroma cells, and for JAZF1 and BDNF examined by oxidative bis-seq, found that when 5hmC is detected, patterns of 5hmC paralleled those of 5mC. Together with prior studies, these results define a consistent epigenetic signature in endometriosis stromal cells and nominate specific transcriptional and signaling pathways as therapeutic targets. PMID:28125717

  15. Epigenetic Alterations Affecting Transcription Factors and Signaling Pathways in Stromal Cells of Endometriosis.

    PubMed

    Yotova, Iveta; Hsu, Emily; Do, Catherine; Gaba, Aulona; Sczabolcs, Matthias; Dekan, Sabine; Kenner, Lukas; Wenzl, Rene; Tycko, Benjamin

    2017-01-01

    Endometriosis is characterized by growth of endometrial-like tissue outside the uterine cavity. Since its pathogenesis may involve epigenetic changes, we used Illumina 450K Methylation Beadchips to profile CpG methylation in endometriosis stromal cells compared to stromal cells from normal endometrium. We validated and extended the Beadchip data using bisulfite sequencing (bis-seq), and analyzed differential methylation (DM) at the CpG-level and by an element-level classification for groups of CpGs in chromatin domains. Genes found to have DM included examples encoding transporters (SLC22A23), signaling components (BDNF, DAPK1, ROR1, and WNT5A) and transcription factors (GATA family, HAND2, HOXA cluster, NR5A1, OSR2, TBX3). Intriguingly, among the TF genes with DM we also found JAZF1, a proto-oncogene affected by chromosomal translocations in endometrial stromal tumors. Using RNA-Seq we identified a subset of the DM genes showing differential expression (DE), with the likelihood of DE increasing with the extent of the DM and its location in enhancer elements. Supporting functional relevance, treatment of stromal cells with the hypomethylating drug 5aza-dC led to activation of DAPK1 and SLC22A23 and repression of HAND2, JAZF1, OSR2, and ROR1 mRNA expression. We found that global 5hmC is decreased in endometriotic versus normal epithelial but not stroma cells, and for JAZF1 and BDNF examined by oxidative bis-seq, found that when 5hmC is detected, patterns of 5hmC paralleled those of 5mC. Together with prior studies, these results define a consistent epigenetic signature in endometriosis stromal cells and nominate specific transcriptional and signaling pathways as therapeutic targets.

  16. The Influence of Stromal Transforming Growth Factor-Beta Receptor Signaling on Mouse Mammary Neoplasia

    DTIC Science & Technology

    2004-08-01

    and -P3) are members of a family of peptide growth factors that include inhibins, bone morphogenic proteins (BMPs) and growth and differentiation...DNIIR) in the mammary epithelium and in stromal fibroblasts resulted in precocious lobuloalveolar development and increased lateral branching...necessary for proper ductal development during puberty . It has been suggested that TGF-P regulates pubertal mammary development through the epithelium and

  17. Highly aligned stromal collagen is a negative prognostic factor following pancreatic ductal adenocarcinoma resection

    PubMed Central

    Drifka, Cole R.; Loeffler, Agnes G.; Mathewson, Kara; Keikhosravi, Adib; Eickhoff, Jens C.; Liu, Yuming; Weber, Sharon M.

    2016-01-01

    Risk factors for pancreatic ductal adenocarcinoma (PDAC) progression after surgery are unclear, and additional prognostic factors are needed to inform treatment regimens and therapeutic targets. PDAC is characterized by advanced sclerosis of the extracellular matrix, and interactions between cancer cells, fibrillar collagen, and other stromal components play an integral role in progression. Changes in stromal collagen alignment have been shown to modulate cancer cell behavior and have important clinical value in other cancer types, but little is known about its role in PDAC and prognostic value. We hypothesized that the alignment of collagen is associated with PDAC patient survival. To address this, pathology-confirmed tissues from 114 PDAC patients that underwent curative-intent surgery were retrospectively imaged with Second Harmonic Generation (SHG) microscopy, quantified with fiber segmentation algorithms, and correlated to patient survival. The same tissue regions were analyzed for epithelial-to-mesenchymal (EMT), α-SMA, and syndecan-1 using complimentary immunohistostaining and visualization techniques. Significant inter-tumoral variation in collagen alignment was found, and notably high collagen alignment was observed in 12% of the patient cohort. Stratification of patients according to collagen alignment revealed that high alignment is an independent negative factor following PDAC resection (p = 0.0153, multivariate). We also found that epithelial expression of EMT and the stromal expression of α-SMA and syndecan-1 were positively correlated with collagen alignment. In summary, stromal collagen alignment may provide additional, clinically-relevant information about PDAC tumors and underscores the importance of stroma-cancer interactions. PMID:27776346

  18. Paracrine Factors Produced by Bone Marrow Stromal Cells Induce Apoptosis and Neuroendocrine Differentiation in Prostate Cancer Cells

    PubMed Central

    Zhang, Chu; Soori, Mehrnoosh; Miles, Fayth; Sikes, Robert A.; Carson, Daniel D.; Chung, Leland L.W.; Farach-Carson, Mary C.

    2010-01-01

    Background Preferential bony metastasis of human prostate cancer (PCa) cells contributes to disease mortality and morbidity. Local factors in bone stromal extracellular matrix microenvironment affect tumor growth through paracrine interactions between tumor and stromal cells. Methods Using co-culture and medium transfer, we used several methods to assess interactions between PCa and bone stromal cells using three PCa cell lines: PC3, LNCaP, and the LNCaP derivative, C4-2B. Results Co-culture of LNCaP and C4-2B cells with bone marrow stromal cell lines, HS27a and HS5, decreased cell number, as did culture with conditioned medium (CM) harvested from these two cell lines suggesting a soluble paracrine factor was responsible. PC3 cell growth was unaffected. CM harvested from bone stromal cell lines triggered apoptosis in LNCaP and C4-2B cell lines, but not in PC3 cells. Surviving C4-2B cells grown in bone stromal cell CM over several days were growth arrested, suggesting presence of a growth inhibitor. Apoptosis induced by CM was dose-dependent. Flow cytometry demonstrated that over a five day culture period in stromal cell CM, LNCaP and C4-2B cell lines, but not PC3 cells, underwent greater apoptosis than parallel cultures in SF medium. The LNCaP and C4-2B cells showed morphology and biomarker expression consistent with transdifferentiation towards a neuroendocrine phenotype after exposure to stromal cell CM. Conclusions The reactive bone stromal microenvironment initially is hostile to PCa cells producing widespread apoptosis. Activation of transdifferentiation in a subset of apoptotic resistant cells may support phenotypic adaptation during disease progression in bone, eventually favoring lethal disease. PMID:20665531

  19. Outcome and Prognostic Factors in Endometrial Stromal Tumors: A Rare Cancer Network Study

    SciTech Connect

    Schick, Ulrike; Bolukbasi, Yasmin; Thariat, Juliette; Abdah-Bortnyak, Roxolyana; Kuten, Abraham; Igdem, Sefik; Caglar, Hale; Ozsaran, Zeynep; Loessl, Kristina; Schleicher, Ursula; Zwahlen, Daniel; Villette, Sylviane; Vees, Hansjoerg

    2012-04-01

    Purpose: To provide further understanding regarding outcome and prognostic factors of endometrial stromal tumors (EST). Methods and Materials: A retrospective analysis was performed on the records of 59 women diagnosed with EST and treated with curative intent between 1983 and 2007 in the framework of the Rare Cancer Network. Results: Endometrial stromal sarcomas (ESS) were found in 44% and undifferentiated ESS (UES) in 49% of the cases. In 7% the grading was unclear. Of the total number of patients, 33 had Stage I, 4 Stage II, 20 Stage III, and 1 presented with Stage IVB disease. Adjuvant chemotherapy was administered to 12 patients, all with UES. External-beam radiotherapy (RT) was administered postoperatively to 48 women. The median follow-up was 41.4 months. The 5-year overall survival (OS) rate was 96.2% and 64.8% for ESS and UES, respectively, with a corresponding 5-year disease-free survival (DFS) rate of 49.4% and 43.4%, respectively. On multivariate analysis, adjuvant RT was an independent prognostic factor for OS (p = 0.007) and DFS (p = 0.013). Locoregional control, DFS, and OS were significantly associated with age ({<=}60 vs. >60 years), grade (ESS vs. UES), and International Federation of Gynecology and Obstetrics stage (I-II vs. III-IV). Positive lymph node staging had an impact on OS (p < 0.001). Conclusion: The prognosis of ESS differed from that of UES. Endometrial stromal sarcomas had an excellent 5-year OS, whereas the OS in UES was rather low. However, half of ESS patients had a relapse. For this reason, adjuvant treatment such as RT should be considered even in low-grade tumors. Multicenter randomized studies are still warranted to establish clear guidelines.

  20. 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

  1. Stromal expression of vascular endothelial growth factor correlates with tumor grade and microvessel density in mammary phyllodes tumors: a multicenter study of 185 cases.

    PubMed

    Tse, Gary M K; Lui, Philip C W; Lee, C Soon; Kung, Fred Y l; Scolyer, Richard A; Law, Bonita K B; Lau, Tai-Shing; Karim, Rooshdiya; Putti, Thomas C

    2004-09-01

    A retrospective review of 185 mammary phyllodes tumors (105 benign, 51 borderline, 29 malignant) from 4 centers was performed by immunohistochemistry to investigate the expression of vascular endothelial growth factor in the epithelial and stromal cells of mammary phyllodes tumors. The correlation of vascular endothelial growth factor with tumor grade, stromal cell nuclear pleomorphism, cellularity, mitotic rate, margin histomorphology, and the stromal microvessel density was evaluated. Vascular endothelial growth factor expression was found in the epithelium in 29% and in the stromal cells in 31% of cases. There was significant increase of vascular endothelial growth factor expression in the stromal cells with increasing degree of malignancy, but not the epithelium. Microvessel density in the stroma also showed significant correlation with tumor malignancy, and a correlation was shown with the stromal vascular endothelial growth factor expression. Statistical overlap of stromal vascular endothelial growth factor and microvessel density in predicting malignancy suggests that angiogenesis may be an effector mechanism for vascular endothelial growth factor. Assessment of stromal VEGF may be useful as an adjunctive diagnostic criterion in the histologic assessment of malignancy in phyllodes tumors.

  2. Role of stromal cell-derived factor 1α pathway in bone metastatic prostate cancer

    PubMed Central

    Gupta, Nisha; Duda, Dan G.

    2016-01-01

    Abstract Metastatic prostate cancer is one of the leading causes of cancer-related death in men. The primary site of metastasis from prostate cancers is the bone. During the last decade, multiple studies have pointed to the role of the stromal cell-derived factor 1 alpha (SDF1α)/CXCR4 axis in the metastatic spread of the disease, but the mechanisms that underlie this effect are still incompletely understood. In this review, we summarize the current understanding of the role of the SDF1α/CXCR4 pathway in bone metastatic prostate cancer. We also discuss the therapeutic potential of disrupting the interaction between prostate tumor cells and bone environment with focus on the SDF1α pathway. PMID:27533927

  3. Insulin-like Growth Factor (IGF) system and gastrointestinal stromal tumours (GIST): present and future.

    PubMed

    Nannini, Margherita; Biasco, Guido; Astolfi, Annalisa; Urbini, Milena; Pantaleo, Maria A

    2014-02-01

    In the last decades, the concept that Insulin-like Growth Factor (IGF) axis plays a key role in several steps of tumorigenesis, cancer growth and metastasis has been widely documented. The aberration of the IGF system has been described in many kinds of tumours, providing several lines of evidence in support of IGF receptor type 1 (IGF1R) as molecular target in cancer treatment. Gastrointestinal stromal tumors (GIST) are the most common mesenchymal tumor of the gastrointestinal tract, commonly characterized in most cases by KIT and PDGFRA gain mutations. Beyond to the well recognized KIT and PDGFRA gain mutations, in the last years other molecular aberrations have been investigated. Recently, several lines of evidence about the involvement of the IGF system in GIST have been accumulated. The aim of this review is to report all current data about the IGF system involvement in GIST, focusing on the current clinical implication and future perspectives.

  4. Paracrine Engineering of Human Explant-Derived Cardiac Stem Cells to Over-Express Stromal-Cell Derived Factor 1α Enhances Myocardial Repair.

    PubMed

    Tilokee, Everad L; Latham, Nicholas; Jackson, Robyn; Mayfield, Audrey E; Ye, Bin; Mount, Seth; Lam, Buu-Khanh; Suuronen, Erik J; Ruel, Marc; Stewart, Duncan J; Davis, Darryl R

    2016-07-01

    First generation cardiac stem cell products provide indirect cardiac repair but variably produce key cardioprotective cytokines, such as stromal-cell derived factor 1α, which opens the prospect of maximizing up-front paracrine-mediated repair. The mesenchymal subpopulation within explant derived human cardiac stem cells underwent lentiviral mediated gene transfer of stromal-cell derived factor 1α. Unlike previous unsuccessful attempts to increase efficacy by boosting the paracrine signature of cardiac stem cells, cytokine profiling revealed that stromal-cell derived factor 1α over-expression prevented lv-mediated "loss of cytokines" through autocrine stimulation of CXCR4+ cardiac stem cells. Stromal-cell derived factor 1α enhanced angiogenesis and stem cell recruitment while priming cardiac stem cells to readily adopt a cardiac identity. As compared to injection with unmodified cardiac stem cells, transplant of stromal-cell derived factor 1α enhanced cells into immunodeficient mice improved myocardial function and angiogenesis while reducing scarring. Increases in myocardial stromal-cell derived factor 1α content paralleled reductions in myocyte apoptosis but did not influence long-term engraftment or the fate of transplanted cells. Transplantation of stromal-cell derived factor 1α transduced cardiac stem cells increased the generation of new myocytes, recruitment of bone marrow cells, new myocyte/vessel formation and the salvage of reversibly damaged myocardium to enhance cardiac repair after experimental infarction. Stem Cells 2016;34:1826-1835. © 2016 AlphaMed Press.

  5. Human Adenomyosis Endometrium Stromal Cells Secreting More Nerve Growth Factor: Impact and Effect.

    PubMed

    Li, Yan; Zou, Shien; Xia, Xian; Zhang, Shaofen

    2015-09-01

    Abnormal expression of nerve growth factor (NGF) was found in adenomyosis (AM). We collected AM foci from patients and eutopic endometrium from non-AM controls. Endometrium stromal cells (ESCs) were cultured. Different levels of 17β-estradiol, tumor necrosis factor (TNF), CoCl2, and H2O2 were added to the culture system separately, then the expression level of NGF in ESCs was detected. After adding different levels of NGF, the proliferation and apoptosis of ESCs and aromatase expression were detected. We found that 17β-estradiol promoted NGF production in AM ESCs but not in control ESCs; TNF promoted NGF production in both AM and control ESCs; and CoCl2 inhibited NGF production in control ESCs, but had no effect in AM ESCs. Nerve growth factor promoted the proliferation and synthesis of aromatase in AM ESCs. In conclusion, locally increased estrogen levels and inflammation may cause increased NGF production in the uterus of patients with AM. Nerve growth factor stimulated the proliferation and increased aromatase expression of ESCs from AM foci, suggesting NGF might contribute to the pathology and etiology of AM.

  6. Stromal Cell-Derived Factor-1 Alpha is Cardioprotective After Myocardial Infarction

    PubMed Central

    Saxena, Ankur; Fish, Jason E.; White, Michael D.; Yu, Sangho; Smyth, James WP; Shaw, Robin M.; DiMaio, J. Michael; Srivastava, Deepak

    2009-01-01

    Background Heart disease is a leading cause of mortality throughout the world. Tissue damage from vascular occlusive events results in the replacement of contractile myocardium by nonfunctional scar tissue. The potential of new technologies to regenerate damaged myocardium is significant, although cell-based therapies must overcome several technical barriers. One possible cell-independent alternative is the direct administration of small proteins to damaged myocardium. Methods and Results Here we show that the secreted signaling protein stromal cell-derived factor-1 alpha (SDF-1α), which activates the cell-survival factor protein kinase B (PKB/Akt) via the G-protein-coupled receptor CXCR4, protected tissue after an acute ischemic event in mice and activated Akt within endothelial cells and myocytes of the heart. Significantly better cardiac function than in control mice was evident as early as 24 hours post-infarction as well as at 3, 14 and 28 days post-infarction. Prolonged survival of hypoxic myocardium was followed by an increase in levels of vascular endothelial growth factor (VEGF) protein and neo-angiogenesis. Consistent with improved cardiac function, mice exposed to SDF-1α demonstrated significantly decreased scar formation than control mice. Conclusions These findings suggest that SDF-1α may serve a tissue-protective and regenerative role for solid organs suffering a hypoxic insult. PMID:18427137

  7. Clinico-pathological characteristics and prognostic factors of gastrointestinal stromal tumors among a Chinese population

    PubMed Central

    Li, Jiehua; Zhang, Haitian; Chen, Zhibai; Su, Ka

    2015-01-01

    Gastrointestinal stromal tumors (GISTs) are the most common primary mesenchymal tumors of the digestive tract. GISTs include a group of heterogeneous tumors with different morphology, biologic behavior, and genetic characteristics, so their epidemiology, clinico-pathological features and prognosis is distinct in different countries. The objective of this study is to analyze clinico-pathological characteristics and prognostic factors of GISTs among Chinese population. We investigated 112 GIST patients were diagnosed between July 2008 and January 2013 at the First Affiliated Hospital of Guangxi Medical University. Histologic evaluation and immunohistochemistry analysis was performed on paraffin-embedded tissue from the 112 GISTs. Overall survival analysis was carried out using the Kaplan-Meier method and the log-rank test. Multivariate analysis was performed according to Cox’s proportional hazards model. Three and 5-year OS rates were 71.4 and 58.6% respectively. Univariate analysis showed that the following factors were significant in predicting OS: tumor site, tumor size, metastasis, resection margin status, cell type, invasion of adjacent organ, invasion of smooth muscle, mitotic rate, P53 and adjuvant therapy with imatinib (P<0.05). Multivariate analysis showed that tumor size, metastasis, resection margin status, mitotic rate, P53 and adjuvant therapy with imatinib were independent prognostic factors associated with OS. This may aid in the prediction of clinical evolution and guide treatments in patients with GIST in China. PMID:26884871

  8. Genetic factors conferring metastasis in osteosarcoma.

    PubMed

    Maximov, Vadim V; Aqeilan, Rami I

    2016-07-01

    Osteosarcoma (OS) is a deadly bone malignancy affecting mostly children and adolescents. OS has outstandingly complex genetic alterations likely due to p53-independent genomic instability. Based on analysis of recent published research we claim existence of various genetic mechanisms of osteosarcomagenesis conferring great variability to different OS properties including metastatic potential. We also propose a model explaining how diverse genetic mechanisms occur and providing a framework for future research. P53-independent preexisting genomic instability, which precedes and frequently causes TP53 genetic alterations, is central in our model. In addition, our analyses reveal a possible cooperation between aberrantly activated HIF-1α and AP-1 genetic pathways in OS metastasis. We also review the involvement of noncoding RNA genes in OS metastasis.

  9. Hepatocyte Growth Factor Is Required for Mesenchymal Stromal Cell Protection Against Bleomycin-Induced Pulmonary Fibrosis

    PubMed Central

    Cahill, Emer F.; Kennelly, Helen; Carty, Fiona; Mahon, Bernard P.

    2016-01-01

    The incidence of idiopathic pulmonary fibrosis is on the rise and existing treatments have failed to halt or reverse disease progression. Mesenchymal stromal cells (MSCs) have potent cytoprotective effects, can promote tissue repair, and have demonstrated efficacy in a range of fibrotic lung diseases; however, the exact mechanisms of action remain to be elucidated. Chemical antagonists and short hairpin RNA knockdown were used to identify the mechanisms of action used by MSCs in promoting wound healing, proliferation, and inhibiting apoptosis. Using the bleomycin induced fibrosis model, the protective effects of early or late MSC administration were examined. The role for hepatocyte growth factor (HGF) in MSC protection against bleomycin lung injury was examined using HGF knockdown MSC. Terminal deoxynucleotidyl transferase (TdT) dUTP nick-end labeling assay was performed on ex vivo lung sections to examine the effects of MSC on apoptosis. MSC conditioned media (CM) enhanced wound closure and inhibited apoptosis of pulmonary cells in vitro. HGF was required for MSC CM enhancement of epithelial cell proliferation and inhibition of apoptosis. In contrast, MSC required COX-2 for CM to inhibit fibroblast proliferation. In a murine model, early administration of MSC protected against bleomycin induced lung fibrosis and correlated with reduced levels of the proinflammatory cytokine interleukin-1β, reduced levels of apoptosis, and significantly increased levels of HGF. These protective effects were in part mediated by MSC derived HGF as HGF knockdown MSC were unable to protect against fibrosis in vivo. These findings delineate the mechanisms of MSC protection in a preclinical model of fibrotic lung disease. Significance The mechanisms used by mesenchymal stromal cells (MSCs) in mediating protective effects in chronic models of lung disease are not understood and remain to be elucidated. These findings from in vitro studies highlight an important role for the MSC

  10. A murine stromal cell line promotes the proliferation of the human factor-dependent leukemic cell line UT-7.

    PubMed

    Auffray, I; Dubart, A; Izac, B; Vainchenker, W; Coulombel, L

    1994-05-01

    In long-term human bone marrow cultures, stromal cells of human origin are usually used on the assumption that human primitive progenitor cells do not respond to cytokines produced by stromal cells from other species. There is accumulating evidence, however, that murine stromal cells also promote maintenance and differentiation of very primitive human stem cells, which suggests the existence of novel stromal activities that cross species barriers. In this study, we show that a murine bone marrow-derived stromal cell line, MS-5, allows the proliferation of the human leukemic cell line UT-7. The long-term growth of UT-7 is usually supported only by human interleukin-3 (IL-3), granulocyte-macrophage colony-stimulating factor (GM-CSF), or erythropoietin (Epo). None of these three cytokines was involved in the observed effect, since murine GM-CSF and IL-3 do not act on human cells and MS-5 cells do not produce Epo. Soluble stem cell factor (SCF) induced UT-7 cell proliferation. However, S1/S1 mutant fibroblasts also supported UT-7 cell growth and anti-c-kit antibodies only partially abolished UT-7 cell proliferative response to MS-5 cells. These observations excluded a major role of SCF in this system. MS-5-derived growth-promoting activity was diffusible, but attempts to grow UT-7 cells in high levels of known soluble murine stromal-derived cytokines active on human cells showed no or minimal response, suggesting that MS-5's proliferative effect was not mediated by known cytokines. Finally, involvement of an autocrine loop of activation induced by MS-5 was excluded: RT-PCR analysis did not detect increased transcripts for GM-CSF, IL-3, IL-6, SCF, or Epo in UT-7 cells cocultured for 2 to 6 days with MS-5. In addition, UT-7 cell proliferation on MS-5 was not inhibited by neutralizing antibodies against the human GM-CSF receptor or the human IL-6 receptor alpha chain. Whether UT-7 cell proliferation triggered by MS-5 reflects the existence of novel stromal cytokines or

  11. Adipose-derived stromal cells overexpressing vascular endothelial growth factor accelerate mouse excisional wound healing.

    PubMed

    Nauta, Allison; Seidel, Catharina; Deveza, Lorenzo; Montoro, Daniel; Grova, Monica; Ko, Sae Hee; Hyun, Jeong; Gurtner, Geoffrey C; Longaker, Michael T; Yang, Fan

    2013-02-01

    Angiogenesis is essential to wound repair, and vascular endothelial growth factor (VEGF) is a potent factor to stimulate angiogenesis. Here, we examine the potential of VEGF-overexpressing adipose-derived stromal cells (ASCs) for accelerating wound healing using nonviral, biodegradable polymeric vectors. Mouse ASCs were transfected with DNA plasmid encoding VEGF or green fluorescent protein (GFP) using biodegradable poly (β-amino) esters (PBAE). Cells transfected using Lipofectamine 2000, a commercially available transfection reagent, were included as controls. ASCs transfected using PBAEs showed enhanced transfection efficiency and 12-15-fold higher VEGF production compared with cells transfected using Lipofectamine 2000 (*P < 0.05). When transplanted into a mouse wild-type excisional wound model, VEGF-overexpressing ASCs led to significantly accelerated wound healing, with full wound closure observed at 8 days compared to 10-12 days in groups treated with ASCs alone or saline control (*P < 0.05). Histology and polarized microscopy showed increased collagen deposition and more mature collagen fibers in the dermis of wound beds treated using PBAE/VEGF-modified ASCs than ASCs alone. Our results demonstrate the efficacy of using nonviral-engineered ASCs to accelerate wound healing, which may provide an alternative therapy for treating many diseases in which wound healing is impaired.

  12. Adipose-derived Stromal Cells Overexpressing Vascular Endothelial Growth Factor Accelerate Mouse Excisional Wound Healing

    PubMed Central

    Nauta, Allison; Seidel, Catharina; Deveza, Lorenzo; Montoro, Daniel; Grova, Monica; Ko, Sae Hee; Hyun, Jeong; Gurtner, Geoffrey C; Longaker, Michael T; Yang, Fan

    2013-01-01

    Angiogenesis is essential to wound repair, and vascular endothelial growth factor (VEGF) is a potent factor to stimulate angiogenesis. Here, we examine the potential of VEGF-overexpressing adipose-derived stromal cells (ASCs) for accelerating wound healing using nonviral, biodegradable polymeric vectors. Mouse ASCs were transfected with DNA plasmid encoding VEGF or green fluorescent protein (GFP) using biodegradable poly (β-amino) esters (PBAE). Cells transfected using Lipofectamine 2000, a commercially available transfection reagent, were included as controls. ASCs transfected using PBAEs showed enhanced transfection efficiency and 12–15-fold higher VEGF production compared with cells transfected using Lipofectamine 2000 (*P < 0.05). When transplanted into a mouse wild-type excisional wound model, VEGF-overexpressing ASCs led to significantly accelerated wound healing, with full wound closure observed at 8 days compared to 10–12 days in groups treated with ASCs alone or saline control (*P < 0.05). Histology and polarized microscopy showed increased collagen deposition and more mature collagen fibers in the dermis of wound beds treated using PBAE/VEGF-modified ASCs than ASCs alone. Our results demonstrate the efficacy of using nonviral-engineered ASCs to accelerate wound healing, which may provide an alternative therapy for treating many diseases in which wound healing is impaired. PMID:23164936

  13. Control of human endometrial stromal cell motility by PDGF-BB, HB-EGF and trophoblast-secreted factors.

    PubMed

    Schwenke, Maren; Knöfler, Martin; Velicky, Philipp; Weimar, Charlotte H E; Kruse, Michelle; Samalecos, Annemarie; Wolf, Anja; Macklon, Nick S; Bamberger, Ana-Maria; Gellersen, Birgit

    2013-01-01

    Human implantation involves extensive tissue remodeling at the fetal-maternal interface. It is becoming increasingly evident that not only trophoblast, but also decidualizing endometrial stromal cells are inherently motile and invasive, and likely contribute to the highly dynamic processes at the implantation site. The present study was undertaken to further characterize the mechanisms involved in the regulation of endometrial stromal cell motility and to identify trophoblast-derived factors that modulate migration. Among local growth factors known to be present at the time of implantation, heparin-binding epidermal growth factor-like growth factor (HB-EGF) triggered chemotaxis (directed locomotion), whereas platelet-derived growth factor (PDGF)-BB elicited both chemotaxis and chemokinesis (non-directed locomotion) of endometrial stromal cells. Supernatants of the trophoblast cell line AC-1M88 and of first trimester villous explant cultures stimulated chemotaxis but not chemokinesis. Proteome profiling for cytokines and angiogenesis factors revealed neither PDGF-BB nor HB-EGF in conditioned media from trophoblast cells or villous explants, while placental growth factor, vascular endothelial growth factor and PDGF-AA were identified as prominent secretory products. Among these, only PDGF-AA triggered endometrial stromal cell chemotaxis. Neutralization of PDGF-AA in trophoblast conditioned media, however, did not diminish chemoattractant activity, suggesting the presence of additional trophoblast-derived chemotactic factors. Pathway inhibitor studies revealed ERK1/2, PI3 kinase/Akt and p38 signaling as relevant for chemotactic motility, whereas chemokinesis depended primarily on PI3 kinase/Akt activation. Both chemotaxis and chemokinesis were stimulated upon inhibition of Rho-associated, coiled-coil containing protein kinase. The chemotactic response to trophoblast secretions was not blunted by inhibition of isolated signaling cascades, indicating activation of

  14. Stromal-derived factor 1 signalling regulates radial and tangential migration in the developing cerebral cortex.

    PubMed

    Liapi, Anastasia; Pritchett, James; Jones, Owen; Fujii, Nobutaka; Parnavelas, John G; Nadarajah, Bagirathy

    2008-01-01

    Stromal-derived factor 1 (SDF-1), a known chemoattractant, and its receptor CXCR4 are widely expressed in the developing and adult cerebral cortex. Recent studies have highlighted potential roles for SDF-1 during early cortical development. In view of the current findings, our histological analysis has revealed a distinct pattern of SDF-1 expression in the developing cerebral cortex at a time when cell proliferation and migration are at peak. To determine the role of chemokine signalling during early cortical development, embryonic rat brain slices were exposed to a medium containing secreted SDF-1 to perturb the endogenous levels of chemokine. Alternatively, brain slices were treated with 40 muM of T140 or AMD3100, known antagonists of CXCR4. Using these experimental approaches, we demonstrate that chemokine signalling is imperative for the maintenance of the early cortical plate. In addition, we provide evidence that both neurogenesis and radial migration are concomitantly regulated by this signalling system. Conversely, interneurons, although not dependent on SDF-1 signalling to transgress the telencephalic boundary, require the chemokine to maintain their tangential migration. Collectively, our results demonstrate that SDF-1 with its distinct pattern of expression is essential and uniquely positioned to regulate key developmental events that underlie the formation of the cerebral cortex.

  15. Platelet-derived growth factors for GMP-compliant propagation of mesenchymal stromal cells.

    PubMed

    Schallmoser, Katharina; Rohde, Eva; Bartmann, Christina; Obenauf, Anna C; Reinisch, Andreas; Strunk, Dirk

    2009-01-01

    Stem cell-based therapies are a promising prospect for regenerative medicine. Particularly, human multipotent mesenchymal stromal cells (MSCs) are currently in focus regarding their regenerative and immune modulating capacities. An increasing number of clinical trials investigating MSC efficiency and safety are ongoing. Ex vivo propagation of human MSCs is considered to be a prerequisite for MSC therapy. The to date standard use of fetal bovine serum in cell culture bears risks including xenoimmunization and transmission of pathogens. Alternatively, human platelet-derived growth factors have been efficiently implemented into routine MSC expansion protocols. In compliance with good manufacturing practice we established an effective time- and resource-saving procedure for MSC propagation in an animal serum-free system. Bone marrow was seeded without manipulation directly in pooled human platelet lysate (pHPL) and L-glutamine supplemented minimum essential medium without antibiotics. Clinical scale expanded MSCs were harvested already after primary culture. MSC quality, identity, purity and function were assessed according to a defined panel of release criteria and comparative genomic hybridization was used to determine genomic stability. Because various potential risks of MSCs have recently been reported, further research is required to prove efficiency and long-term safety of human MSCs for cell therapy.

  16. THE ROLE OF STROMAL DERIVED FACTOR-1 – CXCR7 AXIS IN DEVELOPMENT AND CANCER

    PubMed Central

    Maksym, Radoslaw B.; Tarnowski, Maciej; Grymula, Katarzyna; Tarnowska, Joanna; Wysoczynski, Marcin; Liu, Riu; Czerny, Boguslaw; Ratajczak, Janina; Kucia, Magda; Ratajczak, Mariusz Z.

    2009-01-01

    Cancer metastasis is a major clinical problem that contributes to unsuccessful therapy. Augmenting evidence indicates that metastasizing cancer cells employ several mechanisms that are involved in developmental trafficking of normal stem cells. Stromal-derived factor-1 (SDF-1) is an important α-chemokine that binds to the G-protein-coupled seven-transmembrane span CXCR4. The SDF-1-CXCR4 axis regulates trafficking of normal and malignant cells. SDF-1 is an important chemoattractant for a variety of cells including hematopoietic stem/progenitor cells. For many years, it was believed that CXCR4 was the only receptor for SDF-1. However, several reports recently provided evidence that SDF-1 also binds to another seven-transmembrane span receptor called CXCR7, sharing this receptor with another chemokine family member called Interferon-inducible T-cell chemoattractant (I-TAC). Thus, with CXCR7 identified as a new receptor for SDF-1, the role of the SDF-1-CXCR4 axis in regulating several biological processes becomes more complex. Based on the available literature, this review addresses the biological significance of SDF-1’s interaction with CXCR7, which may act as a kind of decoy or signaling receptor depending on cell type. Augmenting evidence suggests that CXCR7 is involved in several aspects of tumorogenesis and could become an important target for new anti-metastatic and anti-cancer drugs. PMID:19835865

  17. Stromal cell-derived factor-1 potentiates bone morphogenetic protein-2 induced bone formation.

    PubMed

    Higashino, Kosaku; Viggeswarapu, Manjula; Bargouti, Maggie; Liu, Hui; Titus, Louisa; Boden, Scott D

    2011-02-01

    The mechanisms driving bone marrow stem cell mobilization are poorly understood. A recent murine study found that circulating bone marrow-derived osteoprogenitor cells (MOPCs) were recruited to the site of recombinant human bone morphogenetic protein-2 (BMP-2)-induced bone formation. Stromal cell-derived factor-1α (SDF-1α) and its cellular receptor CXCR4 have been shown to mediate the homing of stem cells to injured tissues. We hypothesized that chemokines, such as SDF-1, are also involved with mobilization of bone marrow cells. The CD45(-) fraction is a major source of MOPCs. In this report we determined that the addition of BMP-2 or SDF-1 to collagen implants increased the number of MOPCs in the peripheral blood. BMP-2-induced mobilization was blocked by CXCR4 antibody, confirming the role of SDF-1 in mobilization. We determined for the first time that addition of SDF-1 to implants containing BMP-2 enhances mobilization, homing of MOPCs to the implant, and ectopic bone formation induced by suboptimal BMP-2 doses. These results suggest that SDF-1 increases the number of osteoprogenitor cells that are mobilized from the bone marrow and then home to the implant. Thus, addition of SDF-1 to BMP-2 may improve the efficiency of BMPs in vivo, making their routine use for orthopaedic applications more affordable and available to more patients.

  18. Lack of transforming growth factor-β signaling promotes collective cancer cell invasion through tumor-stromal crosstalk

    PubMed Central

    2012-01-01

    Introduction Transforming growth factor beta (TGF-β) has a dual role during tumor progression, initially as a suppressor and then as a promoter. Epithelial TGF-β signaling regulates fibroblast recruitment and activation. Concurrently, TGF-β signaling in stromal fibroblasts suppresses tumorigenesis in adjacent epithelia, while its ablation potentiates tumor formation. Much is known about the contribution of TGF-β signaling to tumorigenesis, yet the role of TGF-β in epithelial-stromal migration during tumor progression is poorly understood. We hypothesize that TGF-β is a critical regulator of tumor-stromal interactions that promote mammary tumor cell migration and invasion. Methods Fluorescently labeled murine mammary carcinoma cells, isolated from either MMTV-PyVmT transforming growth factor-beta receptor II knockout (TβRII KO) or TβRIIfl/fl control mice, were combined with mammary fibroblasts and xenografted onto the chicken embryo chorioallantoic membrane. These combinatorial xenografts were used as a model to study epithelial-stromal crosstalk. Intravital imaging of migration was monitored ex ovo, and metastasis was investigated in ovo. Epithelial RNA from in ovo tumors was isolated by laser capture microdissection and analyzed to identify gene expression changes in response to TGF-β signaling loss. Results Intravital microscopy of xenografts revealed that mammary fibroblasts promoted two migratory phenotypes dependent on epithelial TGF-β signaling: single cell/strand migration or collective migration. At epithelial-stromal boundaries, single cell/strand migration of TβRIIfl/fl carcinoma cells was characterized by expression of α-smooth muscle actin and vimentin, while collective migration of TβRII KO carcinoma cells was identified by E-cadherin+/p120+/β-catenin+ clusters. TβRII KO tumors also exhibited a twofold greater metastasis than TβRIIfl/fl tumors, attributed to enhanced extravasation ability. In TβRII KO tumor epithelium compared with T

  19. Cloning, expression and identification of an isoform of human stromal cell derived factor-1α

    PubMed Central

    LIANG, YIN-KU; PING, WEI; BIAN, LIU-JIAO

    2015-01-01

    Human stromal cell derived factor-1α (hSDF-1α), a chemotactic factor of stem cells, regulates inflammation, promotes the mobilization of stem cells and induces angiogenesis following ischemia. Six SDF-1 isoforms, SDF-1α, SDF-1β, SDF-1γ, SDF-1δ, SDF-1ε and SDF-1ϕ, which all contain a signal peptide at the N-terminus, have been reported. In the present study a special isoform of hSDF-1α is described that does not contain the N-terminal signal peptide sequence. The hSDF-1α gene was cloned with the recombinant plasmid pCMV-SPORT6-hSDF1 as the template, and the prokaryotic expression vector pET15b-hSDF-1α was constructed. This hSDF-1α was successfully expressed as an inclusion body in Escherichia coli BL21(DE3). The recombinant hSDF-1α was refolded in vitro and separated by cation exchange chromatography. Following these two steps the purity of the hSDF-1α was able to reach >85%. The recombinant hSDF-1α was then purified by size-exclusion chromatography. SDS-PAGE analysis demonstrated that the purity of the hSDF-1α was >95%, which meets almost all the requirements of a protein experiment. Chemotactic activity of the recombinant hSDF-1α was analyzed by Transwell migration assay and it was found that the recombinant hSDF-1α was able to stimulate THP-1 cell migration. These data suggest that the procedure of producing recombinant hSDF-1α proteins with chemotactic activity was feasible and the N-terminal signal peptide of hSDF-1α has little effect on the chemotactic activity of hSDF-1α. PMID:26136888

  20. Patterns of deregulation of insulin growth factor signalling pathway in paediatric and adult gastrointestinal stromal tumours.

    PubMed

    Italiano, Antoine; Chen, Junwei; Zhang, Lei; Hajdu, Mihai; Singer, Samuel; DeMatteo, Ronald P; Antonescu, Cristina R

    2012-11-01

    Data regarding the patterns and the mechanisms of deregulation of the insulin growth factor (IGF) pathway in adult and paediatric gastrointestinal stromal tumours (GISTs) are limited. We investigated the expression profiling of the genes encoding the main components of the IGF signalling pathway in 131 GISTs (106 adults, 21 paediatric and four young adults) and 25 other soft-tissue sarcomas (STS) using an Affymetrix U133A platform. IGF2 was investigated for loss of imprinting (LOI) whereas IGF1R was analysed for copy number aberration and mutation. IGF2 was the most highly overexpressed gene of the IGF pathway in GIST. IGF2 expression was also significantly higher than in other STS. IGF2 expression was correlated to the age onset and mutational status of GIST. Indeed, IGF2 expression was significantly higher in the 'adult' group than in the 'paediatric' and 'young adult' groups. Among adult GIST, IGF2 expression was higher in tumours lacking Homo sapiens v-kit Hardy-Zuckerman 4 feline sarcoma viral oncogene homolog (KIT) or alpha-type platelet-derived growth factor receptor (PDGFRA) mutations in comparison with mutated cases. A trend for a higher expression of IGF2 in resistant GIST in comparison to responsive GIST was also found. Overexpression of IGF2 was not related to LOI. Conversely, the expression of the IGF1R gene was significantly higher in the paediatric group than in the adult group. No copy number gains or mutations of IGF1R were observed. The IGF pathway is deregulated in GIST with distinct patterns according to age onset and mutational status. The IGF pathway may represent a therapeutic target in patients with primary or secondary resistance to imatinib. Copyright © 2012 Elsevier Ltd. All rights reserved.

  1. Genistein promotes DNA demethylation of the steroidogenic factor 1 (SF-1) promoter in endometrial stromal cells

    SciTech Connect

    Matsukura, Hiroshi; Aisaki, Ken-ichi; Igarashi, Katsuhide; Matsushima, Yuko; Kanno, Jun; Muramatsu, Masaaki; Sudo, Katsuko; Sato, Noriko

    2011-08-26

    Highlights: {yields} Genistein (GEN) is a phytoestrogen found in soy products. {yields} GEN demethylated/unsilenced the steroidogenic factor 1 gene in endometrial tissue. {yields} GEN thus altered mRNA expression in uteri of ovariectomized (OVX) mice. {yields} A high-resolution melting assay was used to screen for epigenetic change. {yields} We isolated an endometrial cell clone that was epigenetically modulated by GEN. -- Abstract: It has recently been demonstrated that genistein (GEN), a phytoestrogen in soy products, is an epigenetic modulator in various types of cells; but its effect on endometrium has not yet been determined. We investigated the effects of GEN on mouse uterine cells, in vivo and in vitro. Oral administration of GEN for 1 week induced mild proliferation of the endometrium in ovariectomized (OVX) mice, which was accompanied by the induction of steroidogenic factor 1 (SF-1) gene expression. GEN administration induced demethylation of multiple CpG sites in the SF-1 promoter; these sites are extensively methylated and thus silenced in normal endometrium. The GEN-mediated promoter demethylation occurred predominantly on the luminal side, as opposed to myometrium side, indicating that the epigenetic change was mainly shown in regenerated cells. Primary cultures of endometrial stromal cell colonies were screened for GEN-mediated alterations of DNA methylation by a high-resolution melting (HRM) method. One out of 20 colony-forming cell clones showed GEN-induced demethylation of SF-1. This clone exhibited a high proliferation capacity with continuous colony formation activity through multiple serial clonings. We propose that only a portion of endometrial cells are capable of receiving epigenetic modulation by GEN.

  2. Stromal Cell-Derived Factor-1 Alpha Is Decreased in Women With Migraine With Aura.

    PubMed

    Liman, Thomas G; Neeb, Lars; Rosinski, Jana; Reuter, Uwe; Endres, Matthias

    2016-09-01

    Endothelial dysfunction may contribute to the pathophysiology of migraine with aura. Stromal cell-derived factor-1 alpha (SDF-1α) is involved in the maintenance of endothelial integrity via mobilization of vascular stem cells. We sought to determine whether SDF-1α levels are decreased in women with MA. In this post hoc analysis of a case-cohort study, levels of SDF-1α were determined by enzyme-linked immunosorbent assay. Endothelial function was assessed using peripheral arterial tonometry. Arterial stiffness was assessed by fingertip tonometry derived and heart-rate-adjusted augmentation index (AI). Twenty-eight women with MA and 27 age-matched healthy women were included in this study. Levels of SDF-1α were significantly lower in women with MA compared to age- and risk factor-matched healthy women (1763 ± 281 vs 2013 ± 263 pg/mL, P = 0.006). SDF-1α levels were positively correlated with AI in healthy women (r = 0.49, P = 0.009), but not in women with MA (r = 0.05, P = 0.78). SDF-1α levels were negatively correlated with CD144-positive endothelial microparticles (EMP; r = -0.31, P = .02), and activated CD62E-positive EMP (r = -0.35, P = .01). Levels of SDF-1α are decreased in women with MA and are associated with EMPs as a surrogate marker of endothelial dysfunction. This might contribute to the pathophysiology and vascular risk in MA, but evidence from larger prospective studies is warranted. © 2016 American Headache Society.

  3. Stromal cell–derived factor 2 is critical for Hsp90-dependent eNOS activation

    PubMed Central

    Siragusa, Mauro; Fröhlich, Florian; Park, Eon Joo; Schleicher, Michael; Walther, Tobias C.; Sessa, William C.

    2016-01-01

    Endothelial nitric oxide synthase (eNOS) catalyzes the conversion of l-arginine and molecular oxygen into l-citrulline and nitric oxide (NO), a gaseous second messenger that influences cardiovascular physiology and disease. Several mechanisms regulate eNOS activity and function, including phosphorylation at Ser and Thr residues and protein-protein interactions. Combining a tandem affinity purification approach and mass spectrometry, we identified stromal cell–derived factor 2 (SDF2) as a component of the eNOS macromolecular complex in endothelial cells. SDF2 knockdown impaired agonist-stimulated NO synthesis and decreased the phosphorylation of eNOS at Ser1177, a key event required for maximal activation of eNOS. Conversely, SDF2 overexpression dose-dependently increased NO synthesis through a mechanism involving Akt and calcium (induced with ionomycin), which increased the phosphorylation of Ser1177 in eNOS. NO synthesis by iNOS (inducible NOS) and nNOS (neuronal NOS) was also enhanced upon SDF2 overexpression. We found that SDF2 was a client protein of the chaperone protein Hsp90, interacting preferentially with the M domain of Hsp90, which is the same domain that binds to eNOS. In endothelial cells exposed to vascular endothelial growth factor (VEGF), SDF2 was required for the binding of Hsp90 and calmodulin to eNOS, resulting in eNOS phosphorylation and activation. Thus, our data describe a function for SDF2 as a component of the Hsp90-eNOS complex that is critical for signal transduction in endothelial cells. PMID:26286023

  4. Stromal cell-derived factor 2 is critical for Hsp90-dependent eNOS activation.

    PubMed

    Siragusa, Mauro; Fröhlich, Florian; Park, Eon Joo; Schleicher, Michael; Walther, Tobias C; Sessa, William C

    2015-08-18

    Endothelial nitric oxide synthase (eNOS) catalyzes the conversion of l-arginine and molecular oxygen into l-citrulline and nitric oxide (NO), a gaseous second messenger that influences cardiovascular physiology and disease. Several mechanisms regulate eNOS activity and function, including phosphorylation at Ser and Thr residues and protein-protein interactions. Combining a tandem affinity purification approach and mass spectrometry, we identified stromal cell-derived factor 2 (SDF2) as a component of the eNOS macromolecular complex in endothelial cells. SDF2 knockdown impaired agonist-stimulated NO synthesis and decreased the phosphorylation of eNOS at Ser(1177), a key event required for maximal activation of eNOS. Conversely, SDF2 overexpression dose-dependently increased NO synthesis through a mechanism involving Akt and calcium (induced with ionomycin), which increased the phosphorylation of Ser(1177) in eNOS. NO synthesis by iNOS (inducible NOS) and nNOS (neuronal NOS) was also enhanced upon SDF2 overexpression. We found that SDF2 was a client protein of the chaperone protein Hsp90, interacting preferentially with the M domain of Hsp90, which is the same domain that binds to eNOS. In endothelial cells exposed to vascular endothelial growth factor (VEGF), SDF2 was required for the binding of Hsp90 and calmodulin to eNOS, resulting in eNOS phosphorylation and activation. Thus, our data describe a function for SDF2 as a component of the Hsp90-eNOS complex that is critical for signal transduction in endothelial cells. Copyright © 2015, American Association for the Advancement of Science.

  5. Using polarization-sensitive optical coherence tomography to identify tumor stromal fibrosis and increase tumor biopsy yield (Conference Presentation)

    NASA Astrophysics Data System (ADS)

    Hariri, Lida P.; Adams, David C.; Miller, Alyssa J.; Mino-Kenudson, Mari; Suter, Melissa J.

    2016-03-01

    Tissue biopsy is the principal method used to diagnose tumors in a variety of organ systems. It is essential to maximize tumor yield in biopsy specimens for both clinical diagnostic and research purposes. This is particularly important in tumors where additional tissue is needed for molecular analysis to identify patients who would benefit from mutation-specific targeted therapy, such as in lung carcinomas. Inadvertent sampling of fibrotic stroma within tumor nodules contaminates biopsies, decreases tumor yield, and can impede diagnosis. The ability to assess tumor composition and guide biopsy site selection in real time is likely to improve diagnostic yield. Polarization sensitive OCT (PS-OCT) measures birefringence in organized tissues, such as collagen, and could be used to distinguish tumor from fibrosis. In this study, PS-OCT was obtained in 65 lung nodule samples from surgical resection specimens containing varying ratios of tumor and fibrosis. PS-OCT was obtained with either a custom-built helical scanning catheter (0.8 or 1.6mm in diameter) or a dual-axis bench top scanner. Strong birefringence was observed in nodules containing dense fibrosis, with no birefringence in adjacent regions of tumor. Tumors admixed with early, loosely-organized collagen demonstrated mild-to-moderate birefringence, and tumors with little collagen content showed little to no birefringent signal. PS-OCT provides significant insights into tumor nodule composition, and has potential to differentiate tumor from stromal fibrosis during biopsy site selection to increase diagnostic tumor yield.

  6. Patterns of Deregulation of Insulin Growth Factor Signaling Pathway in Pediatric and Adult Gastrointestinal Stromal Tumors

    PubMed Central

    Italiano, Antoine; Chen, Junwei; Zhang, Lei; Hajdu, Mihai; Singer, Samuel; DeMatteo, Ronald P; Antonescu, Cristina R.

    2013-01-01

    Background Data regarding the patterns and the mechanisms of deregulation of the insulin growth factor (IGF) pathway in adult and pediatric gastrointestinal stromal tumors (GISTs) are limited. Methods We investigated the expression profiling of the genes encoding the main components of the IGF signaling pathway in 131 GISTs (106 adult, 21 pediatric and 4 young adult) and 25 other soft-tissue sarcomas (STS) using an Affymetrix U133A platform. IGF2 was investigated for loss of imprinting (LOI) whereas IGF1R was analyzed for copy number aberration and mutation. Results IGF2 was the most highly overexpressed gene of the IGF pathway in GIST. IGF2 expression was also significantly higher than in other STS. IGF2 expression was correlated to the age onset and mutational status of GIST. Indeed, IGF2 expression was significantly higher in the “adult” group than in the “pediatric” and “young adult” groups. Among adult GIST, IGF2 expression was higher in tumors lacking KIT or PDGFRA mutations in comparison with mutated cases. A trend for a higher expression of IGF2 in resistant GIST in comparison to responsive GIST was also found. Overexpression of IGF2 was not related to LOI. Conversely, the expression of the IGF1R gene was significantly higher in the pediatric group than in the adult group. No copy number gains or mutations of IGF1R were observed. Conclusion The IGF pathway is deregulated in GIST with distinct patterns according to age onset and mutational status. The IGF pathway may represent a therapeutic target in patients with primary or secondary resistance to imatinib. PMID:22770876

  7. Tumor necrosis factor superfamily 14 (LIGHT) controls thymic stromal lymphopoietin to drive pulmonary fibrosis.

    PubMed

    Herro, Rana; Da Silva Antunes, Ricardo; Aguilera, Amelia Roman; Tamada, Koji; Croft, Michael

    2015-09-01

    Pulmonary fibrosis is characterized by excessive accumulation of collagen and α-smooth muscle actin in the lung. The key molecules that promote these phenotypes are of clinical interest. Thymic stromal lymphopoietin (TSLP) has been found at high levels in patients with asthma and idiopathic pulmonary fibrosis, and TSLP has been proposed as a primary driver of lung fibrotic disease. We asked whether tumor necrosis factor superfamily protein 14 (TNFSF14) (aka LIGHT) controls TSLP production to initiate fibrosis. Expression of TSLP and initiation of pulmonary fibrosis induced by bleomycin were assessed in mice deficient in LIGHT. The ability of recombinant LIGHT, given intratracheally to naive mice, to promote TSLP and fibrosis was also determined. Genetic deletion of LIGHT abolished lung TSLP expression driven by bleomycin, accompanied by near-complete absence of accumulation of lung collagen and α-smooth muscle actin. Furthermore, recombinant LIGHT administered in vivo induced lung expression of TSLP in the absence of other inflammatory stimuli, and strikingly reproduced the primary features of bleomycin-driven disease in a TSLP-dependent manner. Blockade of LIGHT binding to either of its receptors, herpes virus entry mediator and lymphotoxin beta receptor, inhibited clinical symptoms of pulmonary fibrosis, and correspondingly both receptors were found on human bronchial epithelial cells, a primary source of TSLP. Moreover, LIGHT induced TSLP directly in human bronchial epithelial cells and synergized with IL-13 and TGF-β in vivo to promote TSLP in the lungs and drive fibrosis. These results show that LIGHT is a profibrogenic cytokine that may be a key driver of TSLP production during the initiation and development of lung fibrotic disease. Copyright © 2015 American Academy of Allergy, Asthma & Immunology. Published by Elsevier Inc. All rights reserved.

  8. Structure and chromosomal localization of the human stromal cell-derived factor 1 (SDF1) gene

    SciTech Connect

    Shirozu, Michio; Takano, Toru; Tada, Hideaki; Honjo, Tasuku

    1995-08-10

    Stromal cell-derived factors 1{alpha} and 1{beta} are small cytokines belonging to the intercrine CXC subfamily and originally isolated from a murine bone-marrow stroma cell line by the signal sequence trap method. cDNA and genomic clones of human SDF1{alpha} and SDF1{beta} (SDF1A and SDF1B) were isolated and characterized. cDNAs of SDF1{alpha} and SDF1{beta} encode proteins of 89 and 93 amino acids, respectively. SDF1{alpha} and SDF1{beta} sequences are more than 92% identical to those of the human counterparts. The genomic structure of the SDF1 gene revealed that human SDF1{alpha} and SDF1{beta} are encoded by a single gene and arise by alternative splicing. SDF1{alpha} and SDF1{beta} are encoded by 3 and 4 exons, respectively. Ubiquitous expression of the SDF1 gene, except in blood cells, was consistent with the presence of the GC-rich sequence in the 5{prime}-flanking region of the SDF1 gene, as is often the case in the {open_quotes}housekeeping{close_quotes} genes. Although genes encoding other members of the intercrine family are localized on chromosome 4q or 17q, the human SDF1 gene was mapped to chromosome 10q by fluorescence in situ hybridization. Strong evolutionary conservation and unique chromosomal localization of the SDF1 gene suggest that SDF1{alpha} and SDF1{beta} may have important functions distinct from those of other members of the intercrine family. 37 refs., 5 figs.

  9. Sustained expression of coagulation factor IX by modified cord blood-derived mesenchymal stromal cells.

    PubMed

    Dodd, Megan; Marquez-Curtis, Leah; Janowska-Wieczorek, Anna; Hortelano, Gonzalo

    2014-01-01

    Hemophilia B patients are subject to frequent and spontaneous bleeding caused by a deficiency of clotting factor IX (FIX). Mesenchymal stromal cells (MSCs) have been used in cellular therapies as a result of their immunomodulatory properties, the ability to home to sites of injury and their amenability to various ex vivo modifications, including lentiviral-mediated gene transfer. MSCs were isolated from human umbilical cord blood and differentiated into adipogenic, chondrogenic and osteogenic lineages. A lentiviral DNA vector containing the human FIX gene was generated using traditional restriction enzyme digest and ligation techniques to generate viable replication-incompetent lentiviral particles that were used to transduce MSCs. Quantitative measurement of FIX expression was conducted using an enzyme-linked immunosorbent assay. The over-expression of FIX was sustained in vitro at levels > 4 µg/10(6) cells/24 h and FIX coagulant activity was > 2.5 mIU/10(6) cells/24 h for the 6-week duration of study. Lentiviral modification of cells with a multiplicity of infection of 10 did not adversely affect the potential of cord blood (CB) MSCs to differentiate to adipocytes, chondrocytes and osteoblastic cells, and the expression of functional FIX was sustained after differentiation and was similar to that in nondifferentiated cells. Modification of human CB MSCs with a lentiviral vector resulted in sustained high FIX expression in vitro after differentiation to adipogenic, chondrogenic and osteoblastic cells. These modified MSCs could have applications in cellular therapies for hemophilia B. Copyright © 2014 John Wiley & Sons, Ltd.

  10. Stromal-Derived Factor-1α (CXCL12) Levels Increase in Periodontal Disease

    PubMed Central

    Havens, Aaron M.; Chiu, Evonne; Taba, Mario; Wang, Jincheng; Shiozawa, Yusuke; Jung, Younghun; Taichman, L. Susan; D'Silva, Nisha J.; Gopalakrishnan, R.; Wang, CunYu; Giannobile, William V.; Taichman, Russell S.

    2008-01-01

    Background The CXC chemokine receptor 4 (CXCR4) and its ligand, stromal cell–derived factor-1 (SDF-1α or CXC chemokine ligand 12) are involved in the trafficking of leukocytes into and out of extravascular tissues. The purpose of this study was to determine whether SDF-1α secreted by host cells plays a role in recruiting inflammatory cells into the periodontia during local inflammation. Methods SDF-1α levels were determined by enzyme-linked immunosorbent assay in gingival crevicular fluid (GCF) of 24 individuals with periodontitis versus healthy individuals in tissue biopsies and in a preclinical rat model of Porphyromonas gingivalis lipopolysaccharide–induced experimental bone loss. Neutrophil chemotaxis assays were also used to evaluate whether SDF-1α plays a role in the recruitment of host cells at periodontal lesions. Results Subjects with periodontal disease had higher levels of SDF-1α in their GCF compared to healthy subjects. Subjects with periodontal disease who underwent mechanical therapy demonstrated decreased levels of SDF-1α. Immunohistologic staining showed that SDF-1α and CXCR4 levels were elevated in samples obtained from periodontally compromised individuals. Similar results were observed in the rodent model. Neutrophil migration was enhanced in the presence of SDF-1α, mimicking immune cell migration in periodontal lesions. Conclusions SDF-1α may be involved in the immune defense pathway activated during periodontal disease. Upon the development of diseased tissues, SDF-1α levels increase and may recruit host defensive cells into sites of inflammation. These studies suggest that SDF-1α may be a useful biomarker for the identification of periodontal disease progression. PMID:18454663

  11. Stromal cell-derived factor-1 (SDF-1) as a target in liver diseases.

    PubMed

    Liepelt, Anke; Tacke, Frank

    2016-08-01

    The chemokine stromal cell-derived factor-1 (SDF-1) or CXCL12 is constitutively expressed in healthy liver. However, its expression increases following acute or chronic liver injury. Liver sinusoidal endothelial cells (LSEC), hepatic stellate cells (HSC), and malignant hepatocytes are important sources of SDF-1/CXCL12 in liver diseases. CXCL12 is able to activate two chemokine receptors with different downstream signaling pathways, CXCR4 and CXCR7. CXCR7 expression is relevant on LSEC, while HSC, mesenchymal stem cells, and tumor cells mainly respond via CXCR4. Here, we summarize recent developments in the field of liver diseases involving this chemokine and its receptors. SDF-1-dependent signaling contributes to modulating acute liver injury and subsequent tissue regeneration. By activating HSC and recruiting mesenchymal cells from bone marrow, CXCL12 can promote liver fibrosis progression, while CXCL12-CXCR7 interactions endorse proregenerative responses in chronic injury. Moreover, the SDF-1 pathway is linked to development of hepatocellular carcinoma (HCC) by promoting tumor growth, angiogenesis, and HCC metastasis. High hepatic CXCR4 expression has been suggested as a biomarker indicating poor prognosis of HCC patients. Tumor-infiltrating myeloid-derived suppressor cells (MDSC) also express CXCR4 and migrate toward CXCL12. Thus CXCL12 inhibition might not only directly block HCC growth but also modulate the tumor microenvironment (angiogenesis, MDSC), thereby sensitizing HCC patients to conventional or emerging novel cancer therapies (e.g., sorafenib, regorafenib, nivolumab, pembrolizumab). We herein summarize the current knowledge on the complex interplay between CXCL12 and CXCR4/CXCR7 in liver diseases and discuss approaches on the therapeutic targeting of these axes in hepatitis, fibrosis, and liver cancer. Copyright © 2016 the American Physiological Society.

  12. Xenopus laevis Stromal cell-derived factor 1: conservation of structure and function during vertebrate development.

    PubMed

    Braun, Mike; Wunderlin, Markus; Spieth, Kathrin; Knöchel, Walter; Gierschik, Peter; Moepps, Barbara

    2002-03-01

    Transmembrane signaling of the CXC chemokine stromal cell-derived factor-1 (SDF-1) is mediated by CXCR4, a G protein-coupled receptor initially identified in leukocytes and shown to serve as a coreceptor for the entry of HIV into lymphocytes. Characterization of SDF-1- and CXCR4-deficient mice has revealed that SDF-1 and CXCR4 are of vital developmental importance. To study the role of the SDF-1/CXCR4-chemokine/receptor system as a regulator of vertebrate development, we isolated and characterized a cDNA encoding SDF-1 of the lower vertebrate Xenopus laevis (xSDF-1). Recombinant xSDF-1 was produced in insect cells, purified, and functionally characterized. Although xSDF-1 is only 64-66% identical with its mammalian counterparts, it is indistinguishable from human (h)SDF-1alpha in terms of activating both X. laevis CXCR4 and hCXCR4. Thus, both xSDF-1 and hSDF-1alpha promoted CXCR4-mediated activation of heterotrimeric G(i2) in a cell-free system and induced release of intracellular calcium ions in and chemotaxis of intact lymphoblastic cells. Analysis of the time course of xSDF-1 mRNA expression during Xenopus embryogenesis revealed a tightly coordinated regulation of xSDF-1 and X. laevis CXCR4. xSDF-1 mRNA was specifically detected in the developing CNS, incipient sensory organs, and the embryonic heart. In Xenopus, CXCR4 mRNA appears to be absent from the heart anlage, but present in neural crest cells. This observation suggests that xSDF-1 expressed in the heart anlage may attract cardiac neural crest cells expressing CXCR4 to migrate to the primordial heart to regulate both septation of the cardiac outflow tract and differentiation of the myocardium during early heart development.

  13. Postacute stromal cell-derived factor-1α expression promotes neurovascular recovery in ischemic mice.

    PubMed

    Li, Yaning; Huang, Jun; He, Xiaosong; Tang, Guanghui; Tang, Yao-Hui; Liu, Yanqun; Lin, Xiaojie; Lu, Yifan; Yang, Guo-Yuan; Wang, Yongting

    2014-06-01

    Acute interventions of stroke are often challenged by a narrow treatment window. In this study, we explore treatments in the postacute phase of stroke with wider windows of opportunity. We investigated the effects of stromal cell-derived factor (SDF-1α) in neurovascular recovery during the postacute phase and downstream signaling pathways, underlying SDF-1α-mediated neurovascular recovery. Adult male Institute of Cancer Research (ICR) mice underwent middle cerebral artery occlusion. One week after middle cerebral artery occlusion, the animals received stereotactic injection of adenoassociated virus (AAV) carrying SDF-1α gene as treatment or AAV-green fluorescent protein as control and were monitored for 5 weeks. Neurobehavioral outcomes were evaluated, and brain atrophy was measured. Neurogenesis and angiogenesis were examined. The proliferation and migration of neural progenitor cells were evaluated. Downstream pathways of SDF-1α were investigated. Inflammatory response was monitored. Neurobehavioral outcomes were improved, and brain atrophy was greatly reduced for ≤5 weeks in AAV-SDF-1α groups when compared with the control. SDF-1 receptor CXCR4 was upregulated and colocalized with neural and endothelial progenitor cells. The number of nestin(+) and doublecortin(+)/bromodeoxyuridine(+) cells in the subventricular zone, doublecortin(+) and neuron(+)/bromodeoxyuridine(+) cells in the perifocal region, and cluster of differentiation (CD)31(+) and bromodeoxyuridine(+)/CD31(+) microvessels are also significantly increased in AAV-SDF-1α groups. Administration of CXCR4 antagonist AMD3100 eliminated the beneficial effects of SDF-1α. SDF-1α/CXCR4 interaction activated AKT, extracellular signal-regulated kinases (ERK), and P38 mitogen-activated protein kinase (MAPK) signaling pathways but not the c-Jun N-terminal kinase (JNK) pathway. SDF-1α promoted neurogenesis and angiogenesis during the postacute phase of ischemia without eliciting an inflammatory response

  14. Continuous delivery of stromal cell-derived factor-1 from alginate scaffolds accelerates wound healing.

    PubMed

    Rabbany, Sina Y; Pastore, Joseph; Yamamoto, Masaya; Miller, Tim; Rafii, Shahin; Aras, Rahul; Penn, Marc

    2010-01-01

    Proper wound diagnosis and management is an increasingly important clinical challenge and is a large and growing unmet need. Pressure ulcers, hard-to-heal wounds, and problematic surgical incisions are emerging at increasing frequencies. At present, the wound-healing industry is experiencing a paradigm shift towards innovative treatments that exploit nanotechnology, biomaterials, and biologics. Our study utilized an alginate hydrogel patch to deliver stromal cell-derived factor-1 (SDF-1), a naturally occurring chemokine that is rapidly overexpressed in response to tissue injury, to assess the potential effects SDF-1 therapy on wound closure rates and scar formation. Alginate patches were loaded with either purified recombinant human SDF-1 protein or plasmid expressing SDF-1 and the kinetics of SDF-1 release were measured both in vitro and in vivo in mice. Our studies demonstrate that although SDF-1 plasmid- and protein-loaded patches were able to release therapeutic product over hours to days, SDF-1 protein was released faster (in vivo K(d) 0.55 days) than SDF-1 plasmid (in vivo K(d) 3.67 days). We hypothesized that chronic SDF-1 delivery would be more effective in accelerating the rate of dermal wound closure in Yorkshire pigs with acute surgical wounds, a model that closely mimics human wound healing. Wounds treated with SDF-1 protein (n = 10) and plasmid (n = 6) loaded patches healed faster than sham (n = 4) or control (n = 4). At day 9, SDF-1-treated wounds significantly accelerated wound closure (55.0 +/- 14.3% healed) compared to nontreated controls (8.2 +/- 6.0%, p < 0.05). Furthermore, 38% of SDF-1-treated wounds were fully healed at day 9 (vs. none in controls) with very little evidence of scarring. These data suggest that patch-mediated SDF-1 delivery may ultimately provide a novel therapy for accelerating healing and reducing scarring in clinical wounds.

  15. Effect of laparoscopic ovarian drilling on vascular endothelial growth factor and ovarian stromal blood flow using 3-dimensional power Doppler.

    PubMed

    El Behery, Manal M; Diab, Abdalla E; Mowafy, Hala; Ebrahiem, Moustafa A; Shehata, Amal E

    2011-02-01

    To determine, by using 3-dimensional power Doppler ultrasonography, the effect of laparoscopic ovarian drilling (LOD) on the serum level of vascular endothelial growth factor (VEGF) and ovarian stromal blood flow changes in polycystic ovary syndrome (PCOS). A prospective controlled clinical study was conducted on 26 clomiphene-resistant women with PCOS who were scheduled for LOD and a control group of 22 fertile regularly menstruating women. VEGF and 3 ovarian Doppler indices-vascularization index, flow index, and vascularization flow index-were measured and compared between the 2 groups, and before and after LOD in the PCOS group. Serum VEGF and the Doppler indices of ovarian stromal blood flow were significantly higher in the PCOS group than in the control group. Serum VEGF and the ovarian stromal blood flow Doppler indices were significantly reduced in the PCOS group after LOD. Increased vascularity in PCOS demonstrated by Doppler blood flow measurements might be explained by the high level of VEGF. LOD reduced ovarian vascularization and serum VEGF. Copyright © 2010 International Federation of Gynecology and Obstetrics. Published by Elsevier Ireland Ltd. All rights reserved.

  16. Modulation properties of factors released by bone marrow stromal cells on activated microglia: an in vitro study

    PubMed Central

    Cizkova, Dasa; Devaux, Stéphanie; Le Marrec-Croq, Françoise; Franck, Julien; Slovinska, Lucia; Blasko, Juraj; Rosocha, Jan; Spakova, Timea; Lefebvre, Christophe; Fournier, Isabelle; Salzet, Michel

    2014-01-01

    In the present paper we develop a new non-cell based (cell-free) therapeutic approach applied to BV2 microglial cells and spinal cord derived primary microglia (PM) using conditioned media from rat bone marrow stromal cells (BMSCs-CM). First we collected conditioned media (CM) from either naive or injured rat spinal cord tissue (SCI-CM, inflammatory stimulation agent) and from rat bone marrow stromal cells (BMSCs-CM, therapeutic immunomodulation agent). They were both subsequently checked for the presence of chemokines and growth, neurotrophic and neural migration factors using proteomics analysis. The data clearly showed that rat BMSCs-CM contain in vitro growth factors, neural migration factors, osteogenic factors, differentiating factors and immunomodulators, whereas SCI-CM contain chemokines, chemoattractant factors and neurotrophic factors. Afterwards we determined whether the BMSCs-CM affect chemotactic activity, NO production, morphological and pro-apoptotic changes of either BV2 or PM cells once activated with SCI-CM. Our results confirm the anti-migratory and NO-inhibitory effects of BMSCs-CM on SCI-CM-activated microglia with higher impact on primary microglia. The cytotoxic effect of BMSCs-CM occurred only on SCI-CM-stimulated BV2 cells and PM, not on naive BV2 cells, nor on PM. Taken together, the molecular cocktail found in BMSCs-CM is favorable for immunomodulatory properties. PMID:25524416

  17. Modeling extracellular matrix (ECM) alterations in ovarian cancer by multiphoton excited fabrication of stromal models (Conference Presentation)

    NASA Astrophysics Data System (ADS)

    Campagnola, Paul J.; Ajeti, Visar; Lara, Jorge; Eliceiri, Kevin W.; Patankar, Mansh

    2016-04-01

    A profound remodeling of the extracellular matrix (ECM) occurs in human ovarian cancer but it unknown how this affects tumor growth, where this understanding could lead to better diagnostics and therapeutic approaches. We investigate the role of these ECM alterations by using multiphoton excited (MPE) polymerization to fabricate biomimetic models to investigate operative cell-matrix interactions in invasion/metastasis. First, we create nano/microstructured gradients mimicking the basal lamina to study adhesion/migration dynamics of ovarian cancer cells of differing metastatic potential. We find a strong haptotactic response that depends on both contact guidance and ECM binding cues. While we found enhanced migration for more invasive cells, the specifics of alignment and directed migration also depend on cell polarity. We further use MPE fabrication to create collagen scaffolds with complex, 3D submicron morphology. The stromal scaffold designs are derived directly from "blueprints" based on SHG images of normal, high risk, and malignant ovarian tissues. The models are seeded with different cancer cell lines and this allows decoupling of the roles of cell characteristics (metastatic potential) and ECM structure and composition (normal vs cancer) on adhesion/migration dynamics. We found the malignant stroma structure promotes enhanced migration and proliferation and also cytoskeletal alignment. Creating synthetic models based on fibers patterns further allows decoupling the topographic roles of the fibers themselves vs their alignment within the tissue. These models cannot be synthesized by other conventional fabrication methods and we suggest the MPE image-based fabrication method will enable a variety of studies in cancer biology.

  18. Transforming growth factor-{beta} inhibits CCAAT/enhancer-binding protein expression and PPAR{gamma} activity in unloaded bone marrow stromal cells

    SciTech Connect

    Ahdjoudj, S.; Kaabeche, K.; Holy, X.; Fromigue, O.; Modrowski, D.; Zerath, E.; Marie, P.J. . E-mail: pierre.marie@larib.inserm.fr

    2005-02-01

    The molecular mechanisms regulating the adipogenic differentiation of bone marrow stromal cells in vivo remain largely unknown. In this study, we investigated the regulatory effects of transforming growth factor beta-2 (TGF-{beta}2) on transcription factors involved in adipogenic differentiation induced by hind limb suspension in rat bone marrow stromal cells in vivo. Time course real-time quantitative reverse-transcription polymerase chain reaction (RT-PCR) analysis of gene expression showed that skeletal unloading progressively increases the expression of CCAAT/enhancer-binding protein (C/EBP){alpha} and C/EBP{beta} {alpha} at 5 days in bone marrow stromal cells resulting in increased peroxisome proliferator-activated receptor {gamma} (PPAR{gamma}2) transcripts at 7 days. TGF-{beta}2 administration in unloaded rats corrected the rise in C/EBP{alpha} and C/EBP{beta} transcripts induced by unloading in bone marrow stromal cells. This resulted in inhibition of PPAR{gamma}2 expression that was associated with increased Runx2 expression. Additionally, the inhibition of C/EBP{alpha} and C/EBP{beta} expression by TGF-{beta}2 was associated with increased PPAR{gamma} serine phosphorylation in bone marrow stromal cells, a mechanism that inhibits PPAR{gamma} transactivating activity. The sequential inhibitory effect of TGF-{beta}2 on C/EBP{alpha}, C/EBP{beta}, and PPAR{gamma}2 resulted in reduced LPL expression and abolition of bone marrow stromal cell adipogenic differentiation, which contributed to prevent bone loss induced by skeletal unloading. We conclude that TGF-{beta}2 inhibits the excessive adipogenic differentiation of bone marrow stromal cells induced by skeletal unloading by inhibiting C/EBP{alpha}, C/EBP{beta}, and PPAR{gamma} expression and activity, which provides a sequential mechanism by which TGF-{beta}2 regulates adipogenic differentiation of bone marrow stromal cells in vivo.

  19. Alkali-induced corneal stromal melting prevention by a novel platelet-activating factor receptor antagonist.

    PubMed

    He, Jiucheng; Bazan, Nicolas G; Bazan, Haydee E P

    2006-01-01

    To evaluate the effect of LAU0901 (2,4,6-trimethyl-1,4-dihydropyridine-3,5-dicarboxylic acid ester), a novel platelet-activating factor (PAF) receptor antagonist, on a rabbit model of severe corneal alkali injury. Adult New Zealand albino rabbits were anesthetized and the right eyes were injured with 2N sodium hydroxide for 60 seconds using a 12-mm plastic well, then rinsed. After the injury, 10 rabbits were treated topically with LAU0901 every 2 hours 4 times per day and received a subconjunctival injection of 200 microL of LAU0901 once per week and 10 rabbits were treated with vehicle the same way. Over the course of 4 weeks, the corneas were examined daily by slitlamp microscopy and corneal ulcers were graded with a clinical scoring system. Ten additional rabbits were treated as described but 1 rabbit from each group was killed at 1, 3, 7, 14, or 21 days after injury. The corneas were processed for histopathologic and immunofluorescence examination. Persistent epithelial defects were present in both groups from day 5 postinjury, but from day 9 through day 25, the average clinical scores of both epithelial defects and stromal ulcerations in the vehicle-treated eyes were significantly higher than those in the LAU0901-treated eyes (P<.01). By day 28, 90% of the eyes in the vehicle-treated group perforated, while only 20% of the eyes in the LAU0901-treated group developed deep ulceration and none were perforated. Histologic examination showed that the corneas treated with LAU0901 for 4 weeks were completely reepithelialized, with fewer inflammatory polymorphonuclear leukocytes and more repair fibroblasts (myofibroblasts) in the stroma as compared with those treated with vehicle. LAU0901 inhibits corneal ulceration and perforation in a severe alkali-burn model in the rabbit. In the cornea, PAF is a strong inflammatory mediator, a chemotactic to inflammatory polymorphonuclear leukocytes, and an inducer of several proteases that degrade the extracellular matrix

  20. Hematopoiesis on cellulose ester membranes. XI. Induction of new bone and a hematopoietic microenvironment by matrix factors secreted by marrow stromal cells.

    PubMed

    Knospe, W H; Husseini, S G; Fried, W

    1989-07-01

    Cellulose ester membranes (CEM) were coated with stromal cells from bone marrow (BM) or bone and implanted intraperitoneally (IP) in CAF1 mice for intervals of 1 to 6 months. Previous studies indicated that matrix factors [glycoproteins (GPs), proteoglycans (PGs), and glycosaminoglycans (GAGs)] were secreted by the regenerating stromal cells and adsorbed by the CEM. After 1 to 6 months, the CEMs were removed, scraped free of adherent cells, and irradiated in vitro with 40 Gy. The scraped and irradiated CEMs were then reimplanted IP or subcutaneously (SC) for periods of 1 to 6 months in secondary syngeneic murine hosts. They were then removed for histologic study. CEMs reimplanted in SC sites developed bone and hematopoiesis as early as 1 month after implantation. Maximum hematopoiesis and bone formation was observed after 3 months. CEMs coated during the initial implantation with bone-derived stromal cells contained more bone and hematopoietic cells than did CEMs coated with marrow-derived stromal cells after SC implementation. Neither the CEMs coated with bone stromal cells nor those coated with marrow stromal cells developed new bone or trilineal hematopoiesis after being implanted IP. A few CEMs contained small foci of granulopoiesis only. We conclude that noncellular matrix substances deposited on CEMs by bone, and to a lesser degree by marrow cells, can induce prestromal cells in the SC tissues to produce a microenvironment suitable for trilineal hematopoiesis.

  1. Plexiform Fibrohistiocytic Tumor on the Ear: Case Report and Immunohistochemical Investigation of Stromal Factor

    PubMed Central

    Shido, Kosuke; Fujimura, Taku; Kakizaki, Aya; Furudate, Sadanori; Asano, Masayuki; Aiba, Setsuya

    2016-01-01

    Plexiform fibrohistiocytic tumor (PFT) is a rare mesenchymal neoplasm of intermediate malignant potential with a high local recurrence rate. In this report, we describe a case of PFT on the ear, which showed a dense deposition of periostin (POSTN) in the stromal areas of the tumor. In addition, dense infiltration of CD163+CD206– tumor-associated macrophages (TAMs) was detected in the same areas as POSTN. Since POSTN was previously reported to possess immunomodulatory effects on TAMs, our present report suggested the significance of the POSTN/TAMs axis in the progression of PFT. PMID:27293390

  2. Tumour necrosis factor-alpha (TNFα) stimulates the growth of human bone marrow stromal cells

    PubMed Central

    Rougier, F.; Cornu, E.; Gachard, N.; Praloran, V.

    1997-01-01

    This study reports that TNF-α is a potent mitogen for human bone marrow sternal cells in vitro (assessed by [3H]-thymidine incorporation into DNA and cell counts). In contrast, cytokines such as IL-1α, IL-1β, IL-2, IL-3, IL-4, IL-6, LIF, SCF, M-CSF, G-CSF and GM-CSF had no effect. The effect of TNF-α on the growth of human bone marrow stromal cells could be of importance during inflammatory processes which take place in the marrow, for example marrow fibrosis. PMID:18472825

  3. Stromal Cell-Derived Factor 1 Polymorphism in Retinal Vein Occlusion

    PubMed Central

    Szigeti, Andrea; Ecsedy, Mónika; Schneider, Miklós; Lénárt, Lilla; Lesch, Balázs; Nagy, Zoltán Zsolt

    2016-01-01

    Background Stromal cell-derived factor 1 (SDF1) has crucial role in the regulation of angiogenesis and ocular neovascularisation (NV). The purpose of this study was to evaluate the association between SDF1-3’G(801)A polymorphism and NV complications of retinal vein occlusion (RVO). Methods 130 patients with RVO (median age: 69.0, range 35–93 years; male/female– 58/72; 55 patients had central RVO, 75 patients had branch RVO) were enrolled in this study. In the RVO group, 40 (30.8%) patients were diagnosed with NV complications of RVO and 90 (69.2%) patients without NVs. The median follow up period was 40.3 months (range: 18–57 months). The SDF1-3’G(801)A polymorphism was detected by PCR-RFLP. Allelic prevalence was related to reference values obtained in the control group consisted of 125 randomly selected, age and gender matched, unrelated volunteers (median age: 68.0, range 36–95 years; male/female– 53/72). Statistical analysis of the allele and genotype differences between groups (RVO patients vs controls; RVO patients with NV vs RVO patients without NV) was determined by chi-squared test. P value of <0.05 was considered statistically significant. Results Hardy-Weinberg criteria was fulfilled in all groups. The SDF1-3’G(801)A allele and genotype frequencies of RVO patients were similar to controls (SDF1-3’A allele: 22.3% vs 20.8%; SDF1-3’(801)AA: 5.4% vs 4.8%, SDF1-3’(801)GG: 60.8% vs 63.2%). The frequency of SDF1-3’(801)AA and SDF1-3’(801)GA genotypes, as well as the SDF1-3’(801)A allele frequency were higher in RVO patients with NV versus in patients without NV complication (SDF1-3’(801)AA+AG genotypes: 57.5% vs 31.1%, p = 0.008; SDF1-3’(801)A allele: 35.0% vs 16.7%, p = 0.002) or versus controls (SDF1-3’(801)AA+AG genotypes 57.5% vs 36.8%, p = 0.021; SDF1-3’(801)A allele: 35.0% vs 20.8% p = 0.01). Carrying of SDF1-3’(801)A allele increased the risk of neovascularisation complications of RVO by 2.69 (OR, 95% CI = 1.47–4

  4. Growth differentiation factor 6 derived from mesenchymal stem/stromal cells reduces age-related functional deterioration in multiple tissues

    PubMed Central

    Hisamatsu, Daisuke; Ohno-Oishi, Michiko; Nakamura, Shiho; Mabuchi, Yo; Naka-Kaneda, Hayato

    2016-01-01

    The senescence-associated secretory phenotype (SASP) has attracted attention as a mechanism that connects cellular senescence to tissue dysfunction, and specific SASP factors have been identified as systemic pro-aging factors. However, little is known about the age-dependent changes in the secretory properties of stem cells. Young, but not old, mesenchymal stem/stromal cells (MSCs) are a well-known source of critical regenerative factors, but the identity of these factors remains elusive. In this study, we identified growth differentiation factor 6 (Gdf6; also known as Bmp13 and CDMP-2) as a regenerative factor secreted from young MSCs. The expression of specific secretory factors, including Gdf6, was regulated by the microRNA (miRNA) miR-17, whose expression declined with age. Upregulation of Gdf6 restored the osteogenic capacity of old MSCs in vitro and exerted positive effects in vivo on aging-associated pathologies such as reduced lymphopoiesis, insufficient muscle repair, reduced numbers of neural progenitors in the brain, and chronic inflammation. Our results suggest that manipulation of miRNA could enable control of the SASP, and that regenerative factors derived from certain types of young cells could be used to treat geriatric diseases. PMID:27311402

  5. The role of p38 mitogen-activated protein kinase in serum-induced leukemia inhibitory factor secretion by bone marrow stromal cells from pediatric myelodysplastic syndromes.

    PubMed

    da Costa, Simone V; Roela, Rosimeire A; Junqueira, Mara Souza; Arantes, Camila; Brentani, M Mitzi

    2010-04-01

    Stromal cells from pediatric myelodysplastic syndromes (MDS) and acute myeloid leukemia (AML) associated with MDS (MDS-AML) present high expression of leukemia inhibitor factor (LIF). We demonstrated using mitogen-activated protein kinase (MAPK) inhibitors that in stromal cells from pediatric MDS and MDS-AML, p38MAPK was critical in serum-induced secretion of LIF. The serum induction of phosphorylated p38MAPK form was observed only in stromal cells from healthy children, whereas in MDS and MDS-AML basal levels were maintained suggesting constitutive p38MAPK activation. Our study suggested the possible importance in pediatric MDS of p38MAPK signaling pathway which may be a future therapeutic target.

  6. Stromal cell-derived factor-1 promotes migration of cells from the upper rhombic lip in cerebellar development.

    PubMed

    Yu, Tao; Huang, Hai; Li, Hui-Fang

    2010-10-01

    During cerebellar development, the chemokine stromal cell-derived factor-1 alpha (SDF-1 alpha) has been shown to play an important role in recruiting cells from the upper rhombic lip (URL) and external granule cell layer (EGL). However, its function in cerebellar development is still poorly understood. Our results have demonstrated that SDF-1 is necessary for EGL development, and URL cells stream to the SDF-1 source in vitro. Results of embryonic URL explant assays and transwell assays indicated that SDF-1 induces neural cell migration from the URL region in chemotactic and chemokinetic responses. The time-lapse results showed that the migration speed of granule cell progenitors out of the URL was accelerated by the addition of recombinant SDF-1 alpha. Collectively, our study shows that SDF-1 increases the motility of URL cells in the absence of a gradient and promotes the migration of granule cell progenitors during cerebellar development.

  7. Thymic stromal lymphopoietin induction by skin irritation is independent of tumour necrosis factor-α, but supported by interleukin-1.

    PubMed

    Kumari, V; Babina, M; Hazzan, T; Worm, M

    2015-04-01

    Thymic stromal lymphopoietin (TSLP) is an extensively studied cytokine linked to the pathogenesis of allergic diseases, but the inherent activities behind TSLP expression are not well defined. To explore the conditions favourable to TSLP induction outside of a typically allergic set-up and determine the associated mechanisms, and to assess whether TSLP is similarly controlled in murine and human skin. A combination of primary keratinocytes, skin explants/epidermal sheets and in vivo strategies was employed. The skin of wild-type and tumour necrosis factor knockout (TNF-/-) mice was subjected to acute irritation. Cells and specimens were stimulated with a range of TSLP inducers in the presence or absence of neutralizing antibodies. TSLP was quantitated by quantitative reverse-transcriptase polymerase chain reaction, enzyme-linked immunosorbent assay and immunohistochemistry. In addition to cytokines, skin irritation brought about by various causes (e.g. shaving, scratching and chemical perturbation) elicited uniformly high-level production of TSLP, which entered the circulatory system. Despite the potency of TNF-α as an in vitro TSLP inducer, the use of TNF-/- mice revealed that this mechanism was completely independent of endogenous TNF-α. Conversely, irritation-elicited TSLP depended on interleukin (IL)-1, which had a more pronounced influence in human skin than in murine skin. Murine and human skin differed considerably regarding TSLP regulation. Thymic stromal lymphopoietin is a general responder to disrupted skin homeostasis and may have a role in triggering the alarm system of the skin. TSLP induction is rapid, transient and driven by a mechanism that does not involve TNF-α, but partially relies on the evolutionarily ancient IL-1 system. The irritated skin secretes TSLP into the circulatory system. TSLP regulation varies between species. © 2014 British Association of Dermatologists.

  8. Distinct Stromal Cell Factor Combinations Can Separately Control Hematopoietic Stem Cell Survival, Proliferation, and Self-Renewal

    PubMed Central

    Wohrer, Stefan; Knapp, David J.H.F.; Copley, Michael R.; Benz, Claudia; Kent, David G.; Rowe, Keegan; Babovic, Sonja; Mader, Heidi; Oostendorp, Robert A.J.; Eaves, Connie J.

    2014-01-01

    Summary Hematopoietic stem cells (HSCs) are identified by their ability to sustain prolonged blood cell production in vivo, although recent evidence suggests that durable self-renewal (DSR) is shared by HSC subtypes with distinct self-perpetuating differentiation programs. Net expansions of DSR-HSCs occur in vivo, but molecularly defined conditions that support similar responses in vitro are lacking. We hypothesized that this might require a combination of factors that differentially promote HSC viability, proliferation, and self-renewal. We now demonstrate that HSC survival and maintenance of DSR potential are variably supported by different Steel factor (SF)-containing cocktails with similar HSC-mitogenic activities. In addition, stromal cells produce other factors, including nerve growth factor and collagen 1, that can antagonize the apoptosis of initially quiescent adult HSCs and, in combination with SF and interleukin-11, produce >15-fold net expansions of DSR-HSCs ex vivo within 7 days. These findings point to the molecular basis of HSC control and expansion. PMID:24910437

  9. Stromal-Derived Factor-1α Correlates With Circulating Endothelial Progenitor Cells and With Acute Lesion Volume in Stroke Patients

    PubMed Central

    Bogoslovsky, Tanya; Spatz, Maria; Chaudhry, Aneeka; Maric, Dragan; Luby, Marie; Frank, Joseph; Warach, Steven

    2016-01-01

    Background and Purpose Endothelial progenitor cells (EPC) are important participants of neovascularization and are mobilized through signaling with stromal-derived factor (SDF-1α), vascular endothelial growth factor (VEGF), granulocyte colony-stimulating factor, and stem cell factor. The association between EPC levels and these growth factors (GF) in acute stroke has not been previously established. We aimed to determine the association between EPC and these GF, and to elucidate a relationship between these GF and stroke severity in acute stroke patients. Methods Seventeen patients were selected from 175 patients with imaging-confirmed acute ischemic stroke. EPC were quantified using CD34, CD133, and VEGF-R2 markers. Plasma VEGF, SDF-1α, granulocyte colony-stimulating factor, and stem cell factor were determined by enzyme-linked immunosorbent assay on days 1 and 3, and brain MRI was performed at baseline and on days 1 and 5 after the stroke onset. Results Levels of SDF-1α strongly (r=0.6) correlated with the numbers of EPC subsets CD133+VEFG-R2+ (P<0.004), CD34+VEGF-R2+ (P<0.01), and CD34+CD133+VEGF-R2+ (P<0.01) on day 1. Stem cell factor strongly (r=0.5) correlated with CD133+VEGF-R2+ (P<0.05). SDF-1α moderately inversely (r<−0.49) correlated with baseline diffusion-weighted imaging lesion volumes (P<0.04). Median levels of SDF-1α (1561 pg/mL) increased (P<0.04) on day 3 compared to day 1 (1379 pg/mL). Similarly, VEGF at day 3 (95 pg/mL) increased (P<0.03) compared to day 1 (64 pg/mL). Conclusions These results indicate that SDF-1α and stem cell factor correlate with an increase in EPC early in ischemic stroke patients. PMID:21257825

  10. Enhanced wound healing by topical administration of mesenchymal stem cells transfected with stromal cell-derived factor-1.

    PubMed

    Nakamura, Yoko; Ishikawa, Hidefumi; Kawai, Katsuya; Tabata, Yasuhiko; Suzuki, Shigehiko

    2013-12-01

    The objective of this study was to investigate the ability of mesenchymal stem cells (MSC) genetically engineered with stromal cell-derived factor-1 (SDF-1) to heal skin wounds. When transfected with SDF-1 plasmid DNA, MSC which were isolated from the bone marrow of rats, secreted SDF-1 for 7 days. In vitro cell migration assay revealed that the SDF-1-engineered MSC (SDF-MSC) enhanced the migration of MSC and dermal fibroblasts to a significantly greater extent than MSC. The SDF-MSC secreted vascular endothelial growth factor, hepatocyte growth factor, and interleukin 6 at a significantly high level. A skin defect model of rats was prepared and MSC and SDF-MSC were applied to the wound to evaluate wound healing in terms of wound size and histological examinations. The wound size decreased significantly faster with SDF-MSC treatment than with MSC and PBS treatments. The length of the neoepithelium and the number of blood vessels newly formed were significantly larger. A cell-tracing experiment with fluorescently labeled cells demonstrated that the percent survival of SDF-MSC in the tissue treated was significantly high compared with that of MSC. It was concluded that SDF-1 genetic engineering is a promising way to promote the wound healing activity of MSC for a skin defect. Copyright © 2013 Elsevier Ltd. All rights reserved.

  11. Insulin-like growth factor 1 receptor is a potential therapeutic target for gastrointestinal stromal tumors

    PubMed Central

    Tarn, Chi; Rink, Lori; Merkel, Erin; Flieder, Douglas; Pathak, Harsh; Koumbi, Daphne; Testa, Joseph R.; Eisenberg, Burton; von Mehren, Margaret; Godwin, Andrew K.

    2008-01-01

    A subset of gastrointestinal stromal tumors (GISTs) lack gain-of-function mutations in c-KIT and PDGFRα. These so-called wild-type (WT) GISTs tend to be less responsive to imatinib-based therapies and have a poor prognosis. We identified amplification of IGF1R in a SNP analysis of GIST and thus studied its potential as a therapeutic target in WT and mutant GIST. Expression of IGF1R and downstream effectors in clinical GIST samples was examined by using immunoblots and immunohistochemistry. The roles of IGF1R signaling in GIST and viability were analyzed by using NVP-AEW541, an inhibitor of IGF1R, alone and in combination with imatinib, or via siRNA silencing of IGF1R. IGF1R was strongly overexpressed, and IGF1R amplification was detected at a significantly higher frequency in WT GISTs, including a pediatric WT GIST, compared with mutant GISTs (P = 0.0173 and P = 0.0163, respectively). Inhibition of IGF1R activity in vitro with NVP-AEW541 or down-regulation of expression with siIGF1R led to cytotoxicity and induced apoptosis in GIST cell lines via AKT and MAPK signaling. Combination of NVP-AEW541 and imatinib in GIST cell lines induced a strong cytotoxicity response. Our results reveal that IGF1R is amplified and the protein is overexpressed in WT and pediatric GISTs. We also demonstrate that the aberrant expression of IGF1R may be associated with oncogenesis in WT GISTs and suggest an alternative and/or complementary therapeutic regimen in the clinical management of all GISTs, especially in a subset of tumors that respond less favorably to imatinib-based therapy. PMID:18550829

  12. Different expression patterns of growth factors in rat fetuses with spina bifida aperta after in utero mesenchymal stromal cell transplantation.

    PubMed

    Li, Hui; Miao, Jianing; Zhao, Guifeng; Wu, Di; Liu, Bo; Wei, Xiaowei; Cao, Songying; Gu, Hui; Zhang, Yi; Wang, Lili; Fan, Yang; Yuan, Zhengwei

    2014-03-01

    In a previous study, we successfully devised a prenatal surgical approach and transplanted mesenchymal stromal cells (MSCs) to fetal rat spinal column to treat retinoic acid-induced neural tube defects in rat. Our results show that MSCs survived, migrated and differentiated into neural lineage cells. We intended to study various growth factor expressions in rat fetal spinal cords with spina bifida aperta after in utero MSC transplantation and the effect of in vivo growth factor introduction for prenatal spina bifida treatment. Pregnant rats were treated with retinoic acid on embryonic day 10 and then received fetal surgery for MSC transplantation and/or lentiviral epidermal growth factor (EGF) injection on embryonic day 16; various growth factor expression in spinal cords from embryonic day 20 fetuses were analyzed by means of quantitative reverse transcriptase-polymerase chain reaction. Terminal deoxynucleotidyl transferase dUTP nick end labeling analysis was performed to observe spinal tissue apoptosis. Growth factor expression was dysregulated in spinal cords with spina bifida. After MSC transplantation, we observed significantly increased expression of EGF, fibroblast growth factor (FGF)-8, FGF-2 and FGF-20 in the MSC transplantation group compared with blank injection; Furthermore, EGF expression positively correlated with surviving MSC amounts. Expression of other growth factors was not significantly different. In vivo EGF introduction reduced spinal tissue apoptosis. Our results suggest that intrinsic EGF and FGF-2, FGF-8 and FGF-20 might affect the in vivo fate of transplanted MSCs in a fetal rat spina bifida model. In vivo EGF introduction together with MSC transplantation might serve as a new strategy for prenatal spina bifida treatment. Copyright © 2014 International Society for Cellular Therapy. All rights reserved.

  13. Interferon-γ differentially modulates the impact of tumor necrosis factor-α on human endometrial stromal cells.

    PubMed

    Spratte, Julia; Oemus, Anne; Zygmunt, Marek; Fluhr, Herbert

    2015-09-01

    The pro-inflammatory T helper (Th)-1 cytokines, tumor necrosis factor-α (TNF-α) and interferon-γ (IFN-γ), are immunological factors relevant at the feto-maternal interface and involved in the pathophysiology of implantation disorders. The synergistic action of the two cytokines has been described with regard to apoptotic cell death and inflammatory responses in different cell types, but little is known regarding the human endometrium. Therefore, we examined the interaction of TNF-α and IFN-γ in human endometrial stromal cells (ESCs). ESCs were isolated from specimens obtained during hysterectomy and decidualized in vitro. Cells were incubated with TNF-α, IFN-γ or signaling-inhibitor. Insulin-like growth factor binding protein (IGFBP)-1, prolactin (PRL), leukemia inhibitory factor (LIF), interleukin (IL)-6, IL-8, regulated on activation normal T-cell expressed and secreted protein (RANTES) and monocyte chemotactic protein (MCP)-1 were measured using ELISA and real-time RT-PCR. Nuclear factor of transcription (NF)-κB and its inhibitor (IκBα) were analyzed by in-cell western assay and transcription factor assay. TNF-α inhibited and IFN-γ did not affect the decidualization of ESCs. In contrast, IFN-gamma differentially modulated the stimulating effect of TNF-alpha on cytokines by enhancing IL-6, RANTES and MCP-1 and attenuating LIF mRNA expression. These effects were time- and dose-dependent. IFN-γ had no impact on the initial activation of NF-κB signaling. Histone-deacetylase activity was involved in the modulating effect of IFN-γ on RANTES secretion. These observations showed a distinct pattern of interaction of the Th-1 cytokines, TNF-α and IFN-γ in the human endometrium, which could play an important role in the pathophysiology of implantation disorders.

  14. Requirement of soluble factors produced by bone marrow stromal cells on the growth of novel established human myeloma cell line.

    PubMed

    Aikawa, Shingo; Hatta, Yoshihiro; Tanaka, Megumi; Kaneita, Yoshitaka; Yasukawa, Kiyotaka; Sawada, Umihiko; Horie, Takashi; Tsuboi, Isao; Aizawa, Shin

    2003-03-01

    The growth of myeloma cells is believed to be mediated by functional interactions between tumor cells and the marrow environment involving the action of several cytokines. We report on the establishment and characterization of a new human myeloma cell line (TAB1) that can be long-term maintained in the presence of conditioned medium of bone marrow stromal cells (BMCM) and a BMCM independent variant, C2-2. Both cell lines have plasma cell morphology and express plasma cell antigens (CD38, PCA-1 and immunoglobulin kappa light chain). In the absence of BMCM, TAB1 cells undergoing apoptosis were observed. Among the adherent molecules tested, these cells expressed VLA-4, ICAM-1 and H-CAM, but not VLA-5, suggesting that these were mostly immature plasmacytes. Introduction with exogenous IL-6 and/or GM-CSF, which were detected in BMCM, partially supported the proliferation of TAB1 cells. Treatment with anti-IL-6 antibody partially inhibited the proliferation of TAB1 cells cultured with BMCM. These findings strongly suggest that TAB1 required at least two or more factors on their growth in vitro; IL-6 was one of the factors necessary for cell growth. Further studies are required to clarify the precise molecules which support TAB1 cell growth in combination with IL-6, however, TAB1 and its variant C2-2 cells may offer an attractive model to unravel novel molecular mechanisms involved in bone marrow stroma-dependent growth of myeloma cells.

  15. The membrane-tethered transcription factor ANAC089 serves as redox-dependent suppressor of stromal ascorbate peroxidase gene expression.

    PubMed

    Klein, Peter; Seidel, Thorsten; Stöcker, Benedikt; Dietz, Karl-Josef

    2012-01-01

    The stromal ascorbate peroxidase (sAPX) functions as central element of the chloroplast antioxidant defense system. Its expression is under retrograde control of chloroplast signals including redox- and reactive oxygen species-linked cues. The sAPX promoter of Arabidopsis thaliana was dissected in transient reporter assays using mesophyll protoplasts. The study revealed regulatory elements up to -1868 upstream of the start codon. By yeast-one-hybrid screening, the transcription factor ANAC089 was identified to bind to the promoter fragment 2 (-1262 to -1646 bp upstream of translational initiation). Upon mutation of the cis-acting element CACG, binding of ANAC089 was abolished. Expression of a fused fluorescent protein version and comparison with known endomembrane markers localized ANAC089 to the trans-Golgi network and the ER. The transcription factor was released upon treatment with reducing agents and targeted to the nucleus. Transactivation assays using wild type and mutated versions of the promoter showed a partial suppression of reporter expression. The data indicate that ANAC089 functions in a negative retrograde loop, lowering sAPX expression if the cell encounters a highly reducing condition. This conclusion was supported by reciprocal transcript accumulation of ANAC089 and sAPX during acclimation to low, normal, and high light conditions.

  16. Soluble Receptor for Advanced Glycation End Products Improves Stromal Cell–Derived Factor-1 Activity in Model Diabetic Environments

    PubMed Central

    Olekson, Melissa Przyborowski; Faulknor, Renea A.; Hsia, Henry C.; Schmidt, Ann Marie; Berthiaume, François

    2016-01-01

    Objective: In diabetes, hyperglycemia causes the accumulation of advanced glycation end products (AGEs) that trigger reactive oxygen species (ROS) generation through binding the receptor for AGEs (RAGE). Because exogenous growth factors have had little success in enhancing chronic wound healing, we investigated whether hyperglycemia-induced AGEs interfere with cellular responses to extracellular signals. We used stromal cell–derived factor-1 (SDF-1), an angiogenic chemokine also known to promote stem cell recruitment in skin wounds. Approach: Human leukemia-60 (HL-60) cells and mouse peripheral blood mononuclear cells (PBMCs), which express the SDF-1 receptor CXCR-4, were incubated for 24 h in medium supplemented with 25 mM d-glucose. Soluble RAGE (sRAGE) was used to block RAGE activation. Response to SDF-1 was measured in cellular migration and ROS assays. A diabetic murine excisional wound model measured SDF-1 liposome and sRAGE activity in vivo. Results: Hyperglycemia led to significant accumulation of AGEs, decreased SDF-1–directed migration, and elevated baseline ROS levels; it suppressed the ROS spike normally triggered by SDF-1. sRAGE decreased the ROS baseline and restored both the SDF-1–mediated spike and cell migration. Topically applied sRAGE alone promoted healing and enhanced the effect of exogenous SDF-1 on diabetic murine wounds. Innovation: While there is interest in using growth factors to improve wound healing, this strategy is largely ineffective in diabetic wounds. We show that sRAGE may restore signaling, thus potentiating the effect of exogenously applied growth factors. Conclusion: Blocking RAGE with sRAGE restores SDF-1–mediated cellular responses in hyperglycemic environments and may potentiate the effectiveness of SDF-1 applied in vivo. PMID:28078186

  17. Enhanced trophic factor secretion by mesenchymal stem/stromal cells with Glycine-Histidine-Lysine (GHK)-modified alginate hydrogels

    PubMed Central

    Jose, Soumia; Hughbanks, Marissa L.; Binder, Bernard Y.K.; Ingavle, Ganesh C.; Leach, J. Kent

    2014-01-01

    Recombinant proteins and cytokines are under broad preclinical and clinical investigation to promote angiogenesis, but their success is limited by ineffective delivery, lack of long-term stability, and excessive cost. Mesenchymal stem/stromal cells (MSC) secrete bioactive trophic factors, and thus, may provide an effective alternative to address these challenges. Glycine-Histidine-Lysine (GHK) is a peptide fragment of osteonectin (SPARC), a matricellular protein with reported proangiogenic potential. We examined the capacity of GHK to upregulate secretion of proangiogenic factors from human MSC in culture and when covalently coupled to alginate hydrogels. GHK had no apparent cytotoxic effects on MSC in culture over a wide range of concentrations. We detected a dose-dependent increase in vascular endothelial growth factor (VEGF) concentration in media conditioned by GHK-treated MSC, which increased endothelial cell proliferation, migration, and tubule formation. We covalently coupled GHK to alginate using carbodiimide chemistry, and human MSC were entrapped in alginate hydrogels to assess VEGF secretion. Similar to monolayer culture, MSC responded to GHK-modified gels by secreting increased concentrations of VEGF and basic fibroblast growth factor (bFGF) compared to unmodified gels. The pre-treatment of MSC with antibodies to α6 and β1 integrins prior to entrapment in GHK-modified gels abrogated VEGF secretion, suggesting that the proangiogenic response of MSC was integrin-mediated. These data demonstrate that the proangiogenic potential of MSC can be significantly increased by the presentation of GHK with a biodegradable carrier, therefore increasing their clinical potential when used for tissue repair. PMID:24468583

  18. Cell Injury-Induced Release of Fibroblast Growth Factor 2: Relevance to Intracerebral Mesenchymal Stromal Cell Transplantations

    PubMed Central

    Vinodkumar, Deepti; McGrogan, Michael; Bates, Damien

    2015-01-01

    Beneficial effects of intracerebral transplantation of mesenchymal stromal cells (MSC) and their derivatives are believed to be mediated mostly by factors produced by engrafted cells. However, the mesenchymal cell engraftment rate is low, and the majority of grafted cells disappear within a short post-transplantation period. Here, we hypothesize that dying transplanted cells can affect surrounding tissues by releasing their active intracellular components. To elucidate the type, amounts, and potency of these putative intracellular factors, freeze/thaw extracts of MSC or their derivatives were tested in enzyme-linked immunosorbent assays and bioassays. We found that fibroblast growth factor (FGF)2 and FGF1, but not vascular endothelial growth factor and monocyte chemoattractant protein 1 levels were high in extracts despite being low in conditioned media. Extracts induced concentration-dependent proliferation of rat cortical neural progenitor cells and human umbilical vein endothelial cells; these proliferative responses were specifically blocked by FGF2-neutralizing antibody. In the neuropoiesis assay with rat cortical cells, both MSC extracts and killed cells induced expression of nestin, but not astrocyte differentiation. However, suspensions of killed cells strongly potentiated the astrogenic effects of live MSC. In transplantation-relevant MSC injury models (peripheral blood cell-mediated cytotoxicity and high cell density plating), MSC death coincided with the release of intracellular FGF2. The data showed that MSC contain a major depot of active FGF2 that is released upon cell injury and is capable of acutely stimulating neuropoiesis and angiogenesis. We therefore propose that both dying and surviving grafted MSC contribute to tissue regeneration. PMID:25873141

  19. Carbon nanotubes functionalized with fibroblast growth factor accelerate proliferation of bone marrow-derived stromal cells and bone formation

    NASA Astrophysics Data System (ADS)

    Hirata, Eri; Ménard-Moyon, Cécilia; Venturelli, Enrica; Takita, Hiroko; Watari, Fumio; Bianco, Alberto; Yokoyama, Atsuro

    2013-11-01

    Multi-walled carbon nanotubes (MWCNTs) were functionalized with fibroblast growth factor (FGF) and the advantages of their use as scaffolds for bone augmentation were evaluated in vitro and in vivo. The activity of FGF was assessed by measuring the effect on the proliferation of rat bone marrow stromal cells (RBMSCs). The presence of FGF enhanced the proliferation of RBMSCs and the FGF covalently conjugated to the nanotubes (FGF-CNT) showed the same effect as FGF alone. In addition, FGF-CNT coated sponges were implanted between the parietal bone and the periosteum of rats and the formation of new bone was investigated. At day 14 after implantation, a larger amount of newly formed bone was clearly observed in most pores of FGF-CNT coated sponges. These findings indicated that MWCNTs accelerated new bone formation in response to FGF, as well as the integration of particles into new bone during its formation. Scaffolds coated with FGF-CNT could be considered as promising novel substituting materials for bone regeneration in future tissue engineering applications.

  20. Connective tissue growth factor regulates adipocyte differentiation of mesenchymal stromal cells and facilitates leukemia bone marrow engraftment

    PubMed Central

    Battula, V. Lokesh; Chen, Ye; Cabreira, Maria da Graca; Ruvolo, Vivian; Wang, Zhiqiang; Ma, Wencai; Konoplev, Sergej; Shpall, Elizabeth; Lyons, Karen; Strunk, Dirk; Bueso-Ramos, Carlos; Davis, Richard Eric; Konopleva, Marina

    2013-01-01

    Mesenchymal stromal cells (MSCs) are a major component of the leukemia bone marrow (BM) microenvironment. Connective tissue growth factor (CTGF) is highly expressed in MSCs, but its role in the BM stroma is unknown. Therefore, we knocked down (KD) CTGF expression in human BM-derived MSCs by CTGF short hairpin RNA. CTGF KD MSCs exhibited fivefold lower proliferation compared with control MSCs and had markedly fewer S-phase cells. CTGF KD MSCs differentiated into adipocytes at a sixfold higher rate than controls in vitro and in vivo. To study the effect of CTGF on engraftment of leukemia cells into BM, an in vivo model of humanized extramedullary BM (EXM-BM) was developed in NOD/SCID/IL-2rgnull mice. Transplanted Nalm-6 or Molm-13 human leukemia cells engrafted at a threefold higher rate in adipocyte-rich CTGF KD MSC-derived EXM-BM than in control EXM-BM. Leptin was found to be highly expressed in CTGF KD EXM-BM and in BM samples of patients with acute myeloid and acute lymphoblastic leukemia, whereas it was not expressed in normal controls. Given the established role of the leptin receptor in leukemia cells, the data suggest an important role of CTGF in MSC differentiation into adipocytes and of leptin in homing and progression of leukemia. PMID:23741006

  1. Compound K attenuates stromal cell-derived growth factor 1 (SDF-1)-induced migration of C6 glioma cells

    PubMed Central

    Kim, Hyuck; Roh, Hyo Sun; Kim, Jai Eun; Park, Sun Dong; Park, Won Hwan

    2016-01-01

    BACKGROUND/OBJECTIVES Stromal cell-derived growth factor 1 (SDF-1), also known as chemokine ligand 12, and chemokine receptor type 4 are involved in cancer cell migration. Compound K (CK), a metabolite of protopanaxadiol-type ginsenoside by gut microbiota, is reported to have therapeutic potential in cancer therapy. However, the inhibitory effect of CK on SDF-1 pathway-induced migration of glioma has not yet been established. MATERIALS/METHODS Cytotoxicity of CK in C6 glioma cells was determined using an EZ-Cytox cell viability assay kit. Cell migration was tested using the wound healing and Boyden chamber assay. Phosphorylation levels of protein kinase C (PKC)α and extracellular signal-regulated kinase (ERK) were measured by western blot assay, and matrix metallopeptidases (MMP) were measured by gelatin-zymography analysis. RESULTS CK significantly reduced the phosphorylation of PKCα and ERK1/2, expression of MMP9 and MMP2, and inhibited the migration of C6 glioma cells under SDF-1-stimulated conditions. CONCLUSIONS CK is a cell migration inhibitor that inhibits C6 glioma cell migration by regulating its downstream signaling molecules including PKCα, ERK1/2, and MMPs. PMID:27247721

  2. Stromal cell-derived factor-1 mediates changes of bone marrow stem cells during the bone repair process.

    PubMed

    Okada, Kiyotaka; Kawao, Naoyuki; Yano, Masato; Tamura, Yukinori; Kurashimo, Shinzi; Okumoto, Katsumi; Kojima, Kotarou; Kaji, Hiroshi

    2016-01-01

    Osteoblasts, osteoclasts, chondrocytes, and macrophages that participate in the bone repair process are derived from hematopoietic stem cells (HSCs) and mesenchymal stem cells (MSCs). However, the roles of these stem cells during the repair of injured bone tissue are still unclear. In the present study, we examined the effects of bone defect on HSCs and MSCs in bone marrow and spleen in 75 mice and its mechanism. We analyzed the HSC and MSC populations in these tissues of a mouse with femoral bone damage by using flow cytometry. The number of HSCs in the bone marrow of mice with damaged femurs was significantly lower than the number of these cells in the bone marrow of the contralateral intact femurs on day 2 after injury. Meanwhile, the number of MSCs in the bone marrow of mice with damaged femurs was significantly higher than that of the contralateral femurs. Both intraperitoneal administration of AMD3100, a C-X-C chemokine receptor 4 (CXCR4) antagonist, and local treatment with an anti-stromal cell-derived factor-1 (SDF-1) antibody blunted the observed decrease in HSC and increase in MSC populations within the bone marrow of injured femurs. In conclusion, the present study revealed that there is a concurrent decrease and increase in the numbers of HSCs and MSCs, respectively, in the bone marrow during repair of mouse femoral bone damage. Furthermore, the SDF-1/CXCR4 system was implicated as contributing to the changes in these stem cell populations upon bone injury.

  3. Fibronectin-Alginate microcapsules improve cell viability and protein secretion of encapsulated Factor IX-engineered human mesenchymal stromal cells.

    PubMed

    Sayyar, Bahareh; Dodd, Megan; Marquez-Curtis, Leah; Janowska-Wieczorek, Anna; Hortelano, Gonzalo

    2015-01-01

    Continuous delivery of proteins by engineered cells encapsu-lated in biocompatible polymeric microcapsules is of considerable therapeutic potential. However, this technology has not lived up to expectations due to inadequate cell--matrix interactions and subsequent cell death. In this study we hypoth-esize that the presence of fibronectin in an alginate matrix may enhance the viability and functionality of encapsulated human cord blood-derived mesenchymal stromal cells (MSCs) expressing the human Factor IX (FIX) gene. MSCs were encapsulated in alginate-PLL microcapsules containing 10, 100, or 500 μg/ml fibronectin to ameliorate cell survival. MSCs in microcapsules with 100 and 500 μg/ml fibronectin demonstrated improved cell viability and proliferation and higher FIX secretion compared to MSCs in non-supplemented microcapsules. In contrast, 10 μg/ml fibronectin did not significantly affect the viability and protein secretion from the encapsulated cells. Differentiation studies demonstrated osteogenic (but not chondrogenic or adipogenic) differentiation capability and efficient FIX secretion of the enclosed MSCs in the fibronectin-alginate suspension culture. Thus, the use of recombinant MSCs encapsulated in fibronectin-alginate microcapsules in basal or osteogenic cultures may be of practical use in the treatment of hemophilia B.

  4. Stromal cell-derived factor 1α (SDF-1α): A marker of disease burden in patients with atrial fibrillation.

    PubMed

    Li, Dana; Bjørnager, Louise; Langkilde, Anne; Andersen, Ove; Jøns, Christian; Agner, Bue F R; Dixen, Ulrik; Landex, Nadia L

    2016-01-01

    Stromal cell-derived factor 1a (SDF-1α), is a chemokine and is able to home hematopoietic progenitor cells to injured areas of heart tissue for structural repair. Previous studies have found increased levels of SDF-1α in several cardiac diseases, but only few studies have investigated SDF-1α in patients with atrial fibrillation (AF). We aimed to test SDF-1α in a large cohort of patients with AF and its role as a prognostic marker. Between January 1st 2008 to December 1st 2012, 290 patients with ECG documented AF were enrolled from the in- and outpatient clinics at the Department of Cardiology, Hvidovre Hospital, University of Copenhagen, Hvidovre, Denmark. Plasma levels of SDF-1α were measured using ELISA technique. Clinical data were registered and patient follow-up was conducted. Patients with permanent AF had significantly higher SDF-1α levels (2199.5 pg/ml) than the patients with paroxysmal AF (1982.0 pg/ml) and persistent AF (1906.0 pg/ml), p < 0.0005. Higher SDF-1α level was associated with longer time spent in the hospital per readmission, p < 0.05. In AF patients, a higher SDF-1α level was found in patients with a more progressive state of arrhythmia and was associated with longer hospitalizations. These findings suggest that SDF-1α could prove valuable in risk stratification and evaluating the disease burden in AF patients.

  5. Involvement of phosphatidylinositol 3-kinase in stromal cell-derived factor-1 alpha-induced lymphocyte polarization and chemotaxis.

    PubMed

    Vicente-Manzanares, M; Rey, M; Jones, D R; Sancho, D; Mellado, M; Rodriguez-Frade, J M; del Pozo, M A; Yáñez-Mó, M; de Ana, A M; Martínez-A, C; Mérida, I; Sánchez-Madrid, F

    1999-10-01

    The role of phosphatidylinositol 3-kinase (PI3-kinase), an important enzyme involved in signal transduction events, has been studied in the polarization and chemotaxis of lymphocytes induced by the chemokine stromal cell-derived factor-1 alpha (SDF-1 alpha). This chemokine was able to directly activate p85/p110 PI3-kinase in whole human PBL and to induce the association of PI3-kinase to the SDF-1 alpha receptor, CXCR4, in a pertussis toxin-sensitive manner. Two unrelated chemical inhibitors of PI3-kinase, wortmannin and Ly294002, prevented ICAM-3 and ERM protein moesin polarization as well as the chemotaxis of PBL in response to SDF-1 alpha. However, they did not interfere with the reorganization of either tubulin or the actin cytoskeleton. Moreover, the transient expression of a dominant negative form of the PI3-kinase 85-kDa regulatory subunit in the constitutively polarized Peer T cell line inhibited ICAM-3 polarization and markedly reduced SDF-1 alpha-induced chemotaxis. Conversely, overexpression of a constitutively activated mutant of the PI3-kinase 110-kDa catalytic subunit in the round-shaped PM-1 T cell line induced ICAM-3 polarization. These results underline the role of PI3-kinase in the regulation of lymphocyte polarization and motility and indicate that PI3-kinase plays a selective role in the regulation of adhesion and ERM proteins redistribution in the plasma membrane of lymphocytes.

  6. Stromal cell–derived factor-1 and hematopoietic cell homing in an adult zebrafish model of hematopoietic cell transplantation

    PubMed Central

    Glass, Tiffany J.; Patrinostro, Xiaobai; Tolar, Jakub; Bowman, Teresa V.; Zon, Leonard I.; Blazar, Bruce R.

    2011-01-01

    In mammals, stromal cell–derived factor-1 (SDF-1) promotes hematopoietic cell mobilization and migration. Although the zebrafish, Danio rerio, is an emerging model for studying hematopoietic cell transplantation (HCT), the role of SDF-1 in the adult zebrafish has yet to be determined. We sought to characterize sdf-1 expression and function in the adult zebrafish in the context of HCT. In situ hybridization of adult zebrafish organs shows sdf-1 expression in kidney tubules, gills, and skin. Radiation up-regulates sdf-1 expression in kidney to nearly 4-fold after 40 Gy. Assays indicate that zebrafish hematopoietic cells migrate toward sdf-1, with a migration ratio approaching 1.5 in vitro. A sdf-1a:DsRed2 transgenic zebrafish allows in vivo detection of sdf-1a expression in the adult zebrafish. Matings with transgenic reporters localized sdf-1a expression to the putative hematopoietic cell niche in proximal and distal renal tubules and collecting ducts. Importantly, transplant of hematopoietic cells into myelosuppressed recipients indicated migration of hematopoietic cells to sdf-1a–expressing sites in the kidney and skin. We conclude that sdf-1 expression and function in the adult zebrafish have important similarities to mammals, and this sdf-1 transgenic vertebrate will be useful in characterizing the hematopoietic cell niche and its interactions with hematopoietic cells. PMID:21622651

  7. Tumor necrosis factor improves vascularization in osteogenic grafts engineered with human adipose-derived stem/stromal cells.

    PubMed

    Hutton, Daphne L; Kondragunta, Renu; Moore, Erika M; Hung, Ben P; Jia, Xiaofeng; Grayson, Warren L

    2014-01-01

    The innate immune response following bone injury plays an important role in promoting cellular recruitment, revascularization, and other repair mechanisms. Tumor necrosis factor-α (TNF) is a prominent pro-inflammatory cytokine in this cascade, and has been previously shown to improve bone formation and angiogenesis in a dose- and timing-dependent manner. This ability to positively impact both osteogenesis and vascular growth may benefit bone tissue engineering, as vasculature is essential to maintaining cell viability in large grafts after implantation. Here, we investigated the effects of exogenous TNF on the induction of adipose-derived stem/stromal cells (ASCs) to engineer pre-vascularized osteogenic tissue in vitro with respect to dose, timing, and co-stimulation with other inflammatory mediators. We found that acute (2-day), low-dose exposure to TNF promoted vascularization, whereas higher doses and continuous exposure inhibited vascular growth. Co-stimulation with platelet-derived growth factor (PDGF), another key factor released following bone injury, increased vascular network formation synergistically with TNF. ASC-seeded grafts were then cultured within polycaprolactone-fibrin composite scaffolds and implanted in nude rats for 2 weeks, resulting in further tissue maturation and increased angiogenic ingrowth in TNF-treated grafts. VEGF-A expression levels were significantly higher in TNF-treated grafts immediately prior to implantation, indicating a long-term pro-angiogenic effect. These findings demonstrate that TNF has the potential to promote vasculogenesis in engineered osteogenic grafts both in vitro and in vivo. Thus, modulation and/or recapitulation of the immune response following bone injury may be a beneficial strategy for bone tissue engineering.

  8. Increased Migration of Human Mesenchymal Stromal Cells by Autocrine Motility Factor (AMF) Resulted in Enhanced Recruitment towards Hepatocellular Carcinoma

    PubMed Central

    Aquino, Jorge B.; Malvicini, Mariana; Rizzo, Manglio; Peixoto, Estanislao; Andriani, Oscar; Alaniz, Laura; Piccioni, Flavia; Bolontrade, Marcela; Podhajcer, Osvaldo

    2014-01-01

    Background and Aims Several reports described the migration of human mesenchymal stromal cells (MSCs) towards tumor-released factors. Autocrine motility factor (AMF) is produced by several tumors including hepatocellular carcinoma (HCC). The aim of this study was to analyze AMF involvement on MSC migration towards human HCC. Methods Production of AMF by HCC tumors was evaluated by western analysis. The effects of AMF on MSCs from different sources (bone marrow, adipose tissue and perivascular cells from umbilical cord) were analyzed using in vitro migration assay; metalloproteinase 2 (MMP2) activity and expression of critical genes were studied by zymography and qRT-PCR, respectively. To assess AMF involvement on the in vivo MSC migration, noninvasive fluorescence imaging was performed. To test the effect of AMF-primed MSCs on tumor development, in vitro proliferation and spheroids growth and in vivo tumor volume were evaluated. Results AMF produced by HCC was found to induce migration of different MSCs in vitro and to enhance their MMP2 activity. Stimulation of MSCs with recombinant AMF (rAMF) also induced the in vitro adhesion to endothelial cells in coincidence with changes in the expression levels of MMP3, AMF receptor, caveolin-1, and -2 and GDI-2. Importantly, stimulation of MSCs with rAMF increased the in vivo migration of MSCs towards experimental HCC tumors. AMF-priming of MSCs did not induce a pro-tumorigenic effect on HCC cells neither in vivo nor in vitro. Conclusion AMF plays a role in MSC recruitment towards HCC. However, its ability to increase MSC migration to HCC for therapeutic purposes merits further evaluation. PMID:24736611

  9. Identification of Pathways Mediating Growth Differentiation Factor5-Induced Tenogenic Differentiation in Human Bone Marrow Stromal Cells.

    PubMed

    Tan, Sik-Loo; Ahmad, Tunku Sara; Ng, Wuey-Min; Azlina, Amir Abbas; Azhar, Mahmood Merican; Selvaratnam, Lakshmi; Kamarul, Tunku

    2015-01-01

    To date, the molecular signalling mechanisms which regulate growth factors-induced MSCs tenogenic differentiation remain largely unknown. Therefore, a study to determine the global gene expression profile of tenogenic differentiation in human bone marrow stromal cells (hMSCs) using growth differentiation factor 5 (GDF5) was conducted. Microarray analyses were conducted on hMSCs cultures supplemented with 100 ng/ml of GDF5 and compared to undifferentiated hMSCs and adult tenocytes. Results of QuantiGene® Plex assay support the use and interpretation of the inferred gene expression profiles and pathways information. From the 27,216 genes assessed, 873 genes (3.21% of the overall human transcriptome) were significantly altered during the tenogenic differentiation process (corrected p<0.05). The genes identified as potentially associated with tenogenic differentiation were ARHGAP29, CCL2, integrin alpha 8 and neurofilament medium polypeptides. These genes, were mainly associated with cytoskeleton reorganization (stress fibers formation) signaling. Pathway analysis demonstrated the potential molecular pathways involved in tenogenic differentiation were: cytoskeleton reorganization related i.e. keratin filament signaling and activin A signaling; cell adhesion related i.e. chemokine and adhesion signaling; and extracellular matrix related i.e. arachidonic acid production signaling. Further investigation using atomic force microscopy and confocal laser scanning microscopy demonstrated apparent cytoskeleton reorganization in GDF5-induced hMSCs suggesting that cytoskeleton reorganization signaling is an important event involved in tenogenic differentiation. Besides, a reduced nucleostemin expression observed suggested a lower cell proliferation rate in hMSCs undergoing tenogenic differentiation. Understanding and elucidating the tenogenic differentiation signalling pathways are important for future optimization of tenogenic hMSCs for functional tendon cell-based therapy and

  10. Identification of Pathways Mediating Growth Differentiation Factor5-Induced Tenogenic Differentiation in Human Bone Marrow Stromal Cells

    PubMed Central

    Tan, Sik-Loo; Ahmad, Tunku Sara; Ng, Wuey-Min; Azlina, Amir Abbas; Azhar, Mahmood Merican; Selvaratnam, Lakshmi; Kamarul, Tunku

    2015-01-01

    To date, the molecular signalling mechanisms which regulate growth factors-induced MSCs tenogenic differentiation remain largely unknown. Therefore, a study to determine the global gene expression profile of tenogenic differentiation in human bone marrow stromal cells (hMSCs) using growth differentiation factor 5 (GDF5) was conducted. Microarray analyses were conducted on hMSCs cultures supplemented with 100 ng/ml of GDF5 and compared to undifferentiated hMSCs and adult tenocytes. Results of QuantiGene® Plex assay support the use and interpretation of the inferred gene expression profiles and pathways information. From the 27,216 genes assessed, 873 genes (3.21% of the overall human transcriptome) were significantly altered during the tenogenic differentiation process (corrected p<0.05). The genes identified as potentially associated with tenogenic differentiation were ARHGAP29, CCL2, integrin alpha 8 and neurofilament medium polypeptides. These genes, were mainly associated with cytoskeleton reorganization (stress fibers formation) signaling. Pathway analysis demonstrated the potential molecular pathways involved in tenogenic differentiation were: cytoskeleton reorganization related i.e. keratin filament signaling and activin A signaling; cell adhesion related i.e. chemokine and adhesion signaling; and extracellular matrix related i.e. arachidonic acid production signaling. Further investigation using atomic force microscopy and confocal laser scanning microscopy demonstrated apparent cytoskeleton reorganization in GDF5-induced hMSCs suggesting that cytoskeleton reorganization signaling is an important event involved in tenogenic differentiation. Besides, a reduced nucleostemin expression observed suggested a lower cell proliferation rate in hMSCs undergoing tenogenic differentiation. Understanding and elucidating the tenogenic differentiation signalling pathways are important for future optimization of tenogenic hMSCs for functional tendon cell-based therapy and

  11. Effect of stromal-cell-derived factor 1 on stem-cell homing and tissue regeneration in ischaemic cardiomyopathy.

    PubMed

    Askari, Arman T; Unzek, Samuel; Popovic, Zoran B; Goldman, Corey K; Forudi, Farhad; Kiedrowski, Matthew; Rovner, Aleksandr; Ellis, Stephen G; Thomas, James D; DiCorleto, Paul E; Topol, Eric J; Penn, Marc S

    2003-08-30

    Myocardial regeneration via stem-cell mobilisation at the time of myocardial infarction is known to occur, although the mechanism for stem-cell homing to infarcted tissue subsequently and whether this approach can be used for treatment of ischaemic cardiomyopathy are unknown. We investigated these issues in a Lewis rat model (ligation of the left anterior descending artery) of ischaemic cardiomyopathy. We studied the effects of stem-cell mobilisation by use of granulocyte colony-stimulating factor (filgrastim) with or without transplantation of syngeneic cells. Shortening fraction and myocardial strain by tissue doppler imaging were quantified by echocardiography. Stem-cell mobilisation with filgrastim alone did not lead to engraftment of bone-marrow-derived cells. Stromal-cell-derived factor 1 (SDF-1), required for stem-cell homing to bone marrow, was upregulated immediately after myocardial infarction and downregulated within 7 days. 8 weeks after myocardial infarction, transplantation into the peri-infarct zone of syngeneic cardiac fibroblasts stably transfected to express SDF-1 induced homing of CD117-positive stem cells to injured myocardium after filgrastim administration (control vs SDF-1-expressing cardiac fibroblasts mean 7.2 [SD 3.4] vs 33.2 [6.0] cells/mm2, n=4 per group, p<0.02) resulting in greater left-ventricular mass (1.24 [0.29] vs 1.57 [0.27] g) and better cardiac function (shortening fraction 9.2 [4.9] vs 17.2 [4.2]%, n=8 per group, p<0.05). These findings show that SDF-1 is sufficient to induce therapeutic stem-cell homing to injured myocardium and suggest a strategy for directed stem-cell engraftment into injured tissues. Our findings also indicate that therapeutic strategies focused on stem-cell mobilisation for regeneration of myocardial tissue must be initiated within days of myocardial infarction unless signalling for stem-cell homing is re-established.

  12. Computational protein design to reengineer stromal cell-derived factor-1α generates an effective and translatable angiogenic polypeptide analog.

    PubMed

    Hiesinger, William; Perez-Aguilar, Jose Manuel; Atluri, Pavan; Marotta, Nicole A; Frederick, John R; Fitzpatrick, J Raymond; McCormick, Ryan C; Muenzer, Jeffrey R; Yang, Elaine C; Levit, Rebecca D; Yuan, Li-Jun; Macarthur, John W; Saven, Jeffery G; Woo, Y Joseph

    2011-09-13

    Experimentally, exogenous administration of recombinant stromal cell-derived factor-1α (SDF) enhances neovasculogenesis and cardiac function after myocardial infarction. Smaller analogs of SDF may provide translational advantages including enhanced stability and function, ease of synthesis, lower cost, and potential modulated delivery via engineered biomaterials. In this study, computational protein design was used to create a more efficient evolution of the native SDF protein. Protein structure modeling was used to engineer an SDF polypeptide analog (engineered SDF analog [ESA]) that splices the N-terminus (activation and binding) and C-terminus (extracellular stabilization) with a diproline segment designed to limit the conformational flexibility of the peptide backbone and retain the relative orientation of these segments observed in the native structure of SDF. Endothelial progenitor cells (EPCs) in ESA gradient, assayed by Boyden chamber, showed significantly increased migration compared with both SDF and control gradients. EPC receptor activation was evaluated by quantification of phosphorylated AKT, and cells treated with ESA yielded significantly greater phosphorylated AKT levels than SDF and control cells. Angiogenic growth factor assays revealed a distinct increase in angiopoietin-1 expression in the ESA- and SDF-treated hearts. In addition, CD-1 mice (n=30) underwent ligation of the left anterior descending coronary artery and peri-infarct intramyocardial injection of ESA, SDF-1α, or saline. At 2 weeks, echocardiography demonstrated a significant gain in ejection fraction, cardiac output, stroke volume, and fractional area change in mice treated with ESA compared with controls. Compared with native SDF, a novel engineered SDF polypeptide analog (ESA) more efficiently induces EPC migration and improves post-myocardial infarction cardiac function and thus offers a more clinically translatable neovasculogenic therapy.

  13. Stromal derived factor-1 regulates bone morphogenetic protein 2-induced osteogenic differentiation of primary mesenchymal stem cells

    PubMed Central

    Hosogane, Naobumi; Huang, Zhiping; Rawlins, Bernard A.; Liu, Xia; Boachie-Adjei, Oheneba; Boskey, Adele L.; Zhu, Wei

    2010-01-01

    Stromal derived factor-1 (SDF-1) is a chemokine signaling molecule that binds to its transmembrane receptor CXC chemokine receptor-4 (CXCR4). While we previously detected that SDF-1 was co-required with bone morphogenetic protein 2 (BMP2) for differentiating mesenchymal C2C12 cells into osteoblastic cells, it is unknown whether SDF-1 is similarly involved in the osteogenic differentiation of mesenchymal stem cells (MSCs). Therefore, here we examined the role of SDF-1 signaling during BMP2-induced osteogenic differentiation of primary MSCs that were derived from human and mouse bone marrow. Our data showed that blocking of the SDF-1/CXCR4 signal axis or adding SDF-1 protein to MSCs significantly affected BMP2-induced alkaline phosphatase (ALP) activity and osteocalcin (OCN) synthesis, markers of preosteoblasts and mature osteoblasts, respectively. Moreover, disrupting the SDF-1 signaling impaired bone nodule mineralization during terminal differentiation of MSCs. Furthermore, we detected that blocking of the SDF-1 signaling inhibited the BMP2-induced early expression of Runt-related factor-2 (Runx2) and osterix (Osx), two “master” regulators of osteogenesis, and the SDF-1 effect was mediated via intracellular Smad and Erk activation. In conclusion, our results demonstrated a regulatory role of SDF-1 in BMP2-induced osteogenic differentiation of MSCs, as perturbing the SDF-1 signaling affected the differentiation of MSCs towards osteoblastic cells in response to BMP2 stimulation. These data provide novel insights into molecular mechanisms underlying MSC osteogenesis, and will contribute to the development of MSC therapies for enhancing bone formation and regeneration in broad orthopaedic situations. PMID:20362069

  14. Stromal platelet-derived growth factor receptor α (PDGFRα) provides a therapeutic target independent of tumor cell PDGFRα expression in lung cancer xenografts

    PubMed Central

    Gerber, David E.; Gupta, Puja; Dellinger, Michael T.; Toombs, Jason E.; Peyton, Michael; Duignan, Inga; Malaby, Jennifer; Bailey, Timothy; Burns, Colleen; Brekken, Rolf A.; Loizos, Nick

    2012-01-01

    In lung cancer, platelet-derived growth factor receptor α (PDGFRα) is expressed frequently by tumor-associated stromal cells and by cancer cells in a subset of tumors. We sought to determine the effect of targeting stromal PDGFRα in preclinical lung tumor xenograft models (human tumor, mouse stroma). Effects of anti-human (IMC-3G3) and anti-mouse (1E10) PDGFRα mAbs on proliferation and PDGFRα signaling were evaluated in lung cancer cell lines and mouse fibroblasts. Therapy studies were performed using established PDGFRα-positive H1703 cells and PDGFRα-negative Calu-6, H1993, and A549 subcutaneous tumors in immunocompromised mice treated with vehicle, anti-PDGFRα mAbs, chemotherapy, or combination therapy. Tumors were analyzed for growth and levels of growth factors. IMC-3G3 inhibited PDGFRα activation and the growth of H1703 cells in vitro and tumor growth in vivo, but had no effect on PDGFRα-negative cell lines or mouse fibroblasts. 1E10 inhibited growth and PDGFRα activation of mouse fibroblasts, but had no effect on human cancer cell lines in vitro. In vivo, 1E10-targeted inhibition of murine PDGFRα reduced tumor growth as single-agent therapy in Calu-6 cells and enhanced the effect of chemotherapy in xenografts derived from A549 cells. We also identified that low expression cancer cell expression of VEGF-A and elevated expression of PDGF-AA were associated with response to stromal PDGFRα targeting. We conclude that stromal PDGFRα inhibition represents a means for enhancing control of lung cancer growth in some cases, independent of tumor cell PDGFRα expression. PMID:22933705

  15. Morphologic, molecular and microenvironment factors associated with stromal invasion in breast ductal carcinoma in situ: Role of myoepithelial cells.

    PubMed

    Aguiar, Fernando N; Cirqueira, Cinthya S; Bacchi, Carlos E; Carvalho, Filomena M

    2015-01-01

    Ductal carcinoma in situ is the last step preceding invasive ductal carcinoma in breast carcinogenesis. We investigated the role of myoepithelial cells and epithelium characteristics as predictors of the risk of stromal invasion. We selected 236 cases with initial diagnosis of DCIS followed by surgical ressection distributed in groups 1 (without invasion) and 2 (with invasive carcinoma). The risk of stromal invasion after a DCIS diagnosis in biopsy was associated to triple-negative profile and loss of CD10 expression by myoepithelial cells, and inversely associated with CK5/6 expression by neoplastic cells and high expression of Smooth Muscle Myosin Heavy Chain (SMMHC) by myoepithelial cells. A combination of characteristics of epithelial and myoepithelial cells in DCIS in biopsy specimens is related to the risk of stromal invasion.

  16. The Interferon-Inducible Host Factor Bone Marrow Stromal Antigen 2/Tetherin Restricts Virion Release, but Is It Actually a Viral Restriction Factor?

    PubMed Central

    Andrew, Amy

    2011-01-01

    Viruses face a variety of obstacles when infecting a new host. The past few years have brought exciting new insights into the function of restriction factors, which form part of the host's innate immune system. One of the most recently identified restriction factors is bone marrow stromal antigen 2 (BST-2)/tetherin. BST-2 is an interferon-inducible gene whose expression dramatically reduces the release of viruses from infected cells. This effect of BST-2 is not specific to human immunodeficiency virus but affects a broad range of enveloped viruses. Since the identification of BST-2 as a restriction factor in 2008, much progress has been made in understanding the molecular properties and functional characteristics of this host factor. The goal of this review was to provide an update on our current understanding of the role of BST-2 in regulating virus release and to discuss its role in controlling virus spread during productive infection with special emphasis on human immunodeficiency virus-1. PMID:21166593

  17. The outcome and predictive factors of sunitinib therapy in advanced gastrointestinal stromal tumors (GIST) after imatinib failure - one institution study

    PubMed Central

    2012-01-01

    Background Gastrointestinal stromal tumors (GIST) mutational status is recognized factor related to the results of tyrosine kinase inhibitors therapy such as imatinib (IM) or sunitinib (SU). Arterial hypertension (AH) is common adverse event related to SU, reported as predictive factor in renal cell carcinoma. The aim of the study was to analyze the outcomes and factors predicting results of SU therapy in inoperable/metastatic CD117(+) GIST patients after IM failure. Methods We identified 137 consecutive patients with advanced inoperable/metastatic GIST treated in one center with SU (2nd line treatment). Median follow-up time was 23 months. Additionally, in 39 patients there were analyzed selected constitutive single nucleotide polymorphisms (SNPs) of VEGFA and VEGFR2 genes. Results One year progression-free survival (PFS; calculated from the start of SU) rate was 42% and median PFS was 43 weeks. The estimated overall survival (OS, calculated both from start of SU or IM) was 74 weeks and 51 months, respectively. One-year PFS was 65% (median 74 weeks) in 55 patients with AH vs. 22% (median 17 weeks) in patients without AH. Patients with primary tumors carrying mutations in KIT exon 9 or wild-type had substantially better 1-year PFS (68% and 57%; median 65.5 and 50.5 weeks, respectively) than patients having tumors with KIT exon 11 or PDGFRA mutations (34% and 15%; median 36.8 and 9 weeks, respectively). We identified two independent factors with significant impact on PFS and OS in univariate and multivariate analysis: primary tumor genotype and presence of AH. The most common adverse events during therapy were: fatigue, AH, hypothyroidism, hand and foot syndrome, mucositis, skin reactions, dyspepsia, and diarrhea. Two deaths were assessed as related to tumor rupture caused by reaction to SU therapy. The presence of C-allele in rs833061 and the T-allele in rs3025039 polymorphism of VEGFA were associated with significantly higher risk of hypothyroidism (OR: 10.0 p = 0

  18. Bone marrow stromal cell paracrine factors direct osteo/odontogenic differentiation of dental pulp cells.

    PubMed

    Al-Sharabi, Niyaz; Xue, Ying; Fujio, Masahito; Ueda, Minoru; Gjerde, Cecilie; Mustafa, Kamal; Fristad, Inge

    2014-11-01

    Growth factors play an important role in osteo/odontogenic differentiation of human dental pulp cells (hDPCs). The aim of this in vitro study was to compare the biological effects of recombinant human growth differentiation factor 5 (rhGDF-5) alone and a cocktail of soluble growth factors (conditioned medium) released from human bone marrow mesenchymal stem cells (hBMMSCs) on the morphology, proliferation and osteo/odontogenic differentiation potential of hDPCs. Passage 4 hDPCs were harvested for culture in four different media: (a) DMEM with 10% FBS, (b) odontogenic induction medium (OM), (c) OM plus 500 ng/mL rhGDF-5, and (d) OM plus conditioned medium (CM). Morphological changes at 48 and 120 h were determined by crystal violet staining. The proliferation rates at 3, 24, 48, and 120 h were assayed by MTT. Using real-time reverse transcription-polymerase chain reaction (RT-PCR), the mRNA levels of dentin sialophosphoprotein (DSPP), dentin matrix protein 1 (DMP1), collagen type I (Col 1), Runt-related transcription factor 2 (Cbfa1/Runx2), alkaline phosphatase (ALP), osteocalcin (OC), β3 tubulin (TUBB3), glial cell-derived neurotrophic factor (GDNF), angiopoietin-1 (Ang1), and vascular endothelial growth factor A (VEGFA), were determined at 2, 5, and 9 days. Protein expression of dental sialoprotein (DSP), DMP1, OC, and TUBB3 was recorded at 5 days, using western blot and immunocytochemistry. The effect of the different culture media on mineralization was determined by ALP staining at day 5 and Alizarin red S staining at days 7 and 14. In response to the different culture media, the shape of the hDPCs varied from spindled to polygonal and cuboidal. CM inhibited the cellular proliferation rate, while rhGDF-5 had no effect at early time points, but promoted cellular proliferation at 120 h of culture. In the CM group, the mRNA levels of Cbfa1/Runx2, Col 1, ALP, VEGFA, Ang1, and TUBB3 decreased and the levels of GDNF and OC increased. The mRNA levels of DSPP

  19. Circulating Stromal Cell-Derived Factor 1α Levels in Heart Failure: A Matter of Proper Sampling

    PubMed Central

    Baerts, Lesley; Waumans, Yannick; Brandt, Inger; Jungraithmayr, Wolfgang; Van der Veken, Pieter; Vanderheyden, Marc; De Meester, Ingrid

    2015-01-01

    Background The chemokine Stromal cell-derived factor 1α (SDF1α, CXCL12) is currently under investigation as a biomarker for various cardiac diseases. The correct interpretation of SDF1α levels is complicated by the occurrence of truncated forms that possess an altered biological activity. Methodology We studied the immunoreactivities of SDF1α forms and evaluated the effect of adding a DPP4 inhibitor in sampling tubes on measured SDF1α levels. Using optimized sampling, we measured DPP4 activity and SDF1α levels in patients with varying degrees of heart failure. Results The immunoreactivities of SDF1α and its degradation products were determined with three immunoassays. A one hour incubation of SDF1α with DPP4 at 37°C resulted in 2/3 loss of immunoreactivity in each of the assays. Incubation with serum gave a similar result. Using appropriate sampling, SDF1α levels were found to be significantly higher in those heart failure patients with a severe loss of left ventricular function. DPP4 activity in serum was not altered in the heart failure population. However, the DPP4 activity was found to be significantly decreased in patients with high SDF1α levels Conclusions We propose that all samples for SDF1α analysis should be collected in the presence of at least a DPP4 inhibitor. In doing so, we found higher SDF1α levels in subgroups of patients with heart failure. Our work supports the need for further research on the clinical relevance of SDF1α levels in cardiac disease. PMID:26544044

  20. Defining an optimal stromal derived factor-1 presentation for effective recruitment of mesenchymal stem cells in 3D.

    PubMed

    Iannone, Maria; Ventre, Maurizio; Pagano, Gemma; Giannoni, Paolo; Quarto, Rodolfo; Netti, Paolo Antonio

    2014-11-01

    In "situ" tissue engineering is a promising approach in regenerative medicine, envisaging to potentiate the physiological tissue repair processes by recruiting the host's own cellular progenitors at the lesion site by means of bioactive materials. Despite numerous works focused the attention in characterizing novel chemoattractant molecules, only few studied the optimal way to present signal in the microenvironment, in order to recruit cells more effectively. In this work, we have analyzed the effects of gradients of stromal derived factor-1 (SDF-1) on the migratory behavior of human mesenchymal stem cells (MSCs). We have characterized the expression of the chemokine-associated receptor, CXCR4, using cytofluorimetric and real-time PCR analyses. Gradients of SDF-1 were created in 3D collagen gels in a chemotaxis chamber. Migration parameters were evaluated using different chemoattractant concentrations. Our results show that cell motion is strongly affected by the spatio-temporal features of SDF-1 gradients. In particular, we demonstrated that the presence of SDF-1 not only influences cell motility but alters the cell state in terms of SDF-1 receptor expression and productions, thus modifying the way cells perceive the signal itself. Our observations highlight the importance of a correct stimulation of MSCs by means of SDF-1 in order to implement on effective cell recruitment. Our results could be useful for the creation of a "cell instructive material" that is capable to communicate with the cells and control and direct tissue regeneration. Biotechnol. Bioeng. 2014;111: 2303-2316. © 2014 Wiley Periodicals, Inc. © 2014 Wiley Periodicals, Inc.

  1. Stromal cell-derived factor-1 promotes human adipose tissue-derived stem cell survival and chronic wound healing

    PubMed Central

    LI, QIANG; GUO, YANPING; CHEN, FEIFEI; LIU, JING; JIN, PEISHENG

    2016-01-01

    Adipose tissue-derived stem cells (ADSCs) hold great potential for the stem cell-based therapy of cutaneous wound healing. Stromal cell-derived factor-1 (SDF-1) activates CXC chemokine receptor (CXCR)4+ and CXCR7+ cells and plays an important role in wound healing. Increasing evidence suggests a critical role for SDF-1 in cell apoptosis and the survival of mesenchymal stem cells. However, the function of SDF-1 in the apoptosis and wound healing ability of ADSCs is not well understood. The aim of this study was to analyze the effect of SDF-1 on the apoptosis and therapeutic effect of ADSCs in cutaneous chronic wounds in vitro and in vivos. By flow cytometric analysis, it was found that hypoxia and serum free promoted the apoptosis of ADSCs. When pretreated with SDF-1, the apoptosis of ADSCs induced by hypoxia and serum depletion was partly recovered. Furthermore, in vivo experiments established that the post-implantation cell survival and chronic wound healing ability of ADSCs were increased following pretreatment with SDF-1 in a diabetic mouse model of chronic wound healing. To explore the potential mechanism underlying the effect of SDF-1 on ADSC apoptosis, western blot analysis was employed and the results indicate that SDF-1 may protect against cell apoptosis in hypoxic and serum-free conditions through activation of the caspase signaling pathway in ADSCs. This study provides evidence that SDF-1 pretreatment can increase the therapeutic effect of ADSCs in cutaneous chronic wounds in vitro and in vivo. PMID:27347016

  2. Plasmid-based transient human stromal cell-derived factor-1 gene transfer improves cardiac function in chronic heart failure

    PubMed Central

    Sundararaman, S; Miller, T J; Pastore, J M; Kiedrowski, M; Aras, R; Penn, M S

    2011-01-01

    We previously demonstrated that transient stromal cell-derived factor-1 alpha (SDF-1) improved cardiac function when delivered via cell therapy in ischemic cardiomyopathy at a time remote from acute myocardial infarction (MI) rats. We hypothesized that non-viral gene transfer of naked plasmid DNA-expressing hSDF-1 could similarly improve cardiac function. To optimize plasmid delivery, we tested SDF-1 and luciferase plasmids driven by the cytomegalovirus (CMV) promoter with (pCMVe) or without (pCMV) translational enhancers or α myosin heavy chain (pMHC) promoter in a rodent model of heart failure. In vivo expression of pCMVe was 10-fold greater than pCMV and pMHC expression and continued over 30 days. We directly injected rat hearts with SDF-1 plasmid 1 month after MI and assessed heart function. At 4 weeks after plasmid injection, we observed a 35.97 and 32.65% decline in fractional shortening (FS) in control (saline) animals and pMHC-hSDF1 animals, respectively, which was sustained to 8 weeks. In contrast, we observed a significant 24.97% increase in animals injected with the pCMVe-hSDF1 vector. Immunohistochemistry of cardiac tissue revealed a significant increase in vessel density in the hSDF-1-treated animals compared with control animals. Increasing SDF-1 expression promoted angiogenesis and improved cardiac function in rats with ischemic heart failure along with evidence of scar remodeling with a trend toward decreased myocardial fibrosis. These data demonstrate that stand-alone non-viral hSDF-1 gene transfer is a strategy for improving cardiac function in ischemic cardiomyopathy. PMID:21472007

  3. Mathematically-Engineered Stromal Cell-Derived Factor 1alpha Stem Cell Cytokine Analogue Enhances Mechanical Properties of Infarcted Myocardium

    PubMed Central

    Jr, John W. MacArthur; Trubelja, Alen; Shudo, Yasuhiro; Hsiao, Philip; Fairman, Alex; Yang, Elaine; Hiesinger, William; Atluri, Pavan; Woo, Y Joseph

    2014-01-01

    Background The biomechanical response to a myocardial infarction consists of ventricular remodeling that leads to dilation, loss of contractile function, abnormal stress patterns and ultimately heart failure. We hypothesized that intramyocardial injection of our previously designed pro-angiogenic chemokine, an engineered stromal cell derived factor 1alpha analogue(ESA), improves mechanical properties of the heart post-infarction. Methods Male rats (n=54) underwent either sham surgery (n=17) with no coronary artery ligation or ligation of the LAD (n=37). Rats in the MI group were then randomized to receive either saline (0.1cc, n=18) or ESA (6μg/kg, n=19) injected into the myocardium at 4 predetermined spots around the borderzone. Echocardiograms were performed preoperatively and before the terminal surgery. After 4 weeks the hearts were explanted and longitudinally sectioned. Uniaxial tensile testing was completed using an Instron 5543 Microtester. Optical strain was evaluated utilizing custom image acquisition software, Digi-Velpo, and analyzed in MATLAB. Results Compared to the saline control group at 4 weeks, the ESA injected hearts had higher ejection fractions (71.8% ± 9.0 vs. 55.3% ± 12.6, p= 0.0004) smaller end-diastolic left ventricular internal dimensions (0.686cm ± 0.110 vs. 0.763cm ± 0.160, p= 0.04), higher cardiac output (36ml/min ± 11.6 vs. 26.9ml/min ± 7.3, p= 0.05) and the tensile modulus was lower(251kPa ± 56 vs. 301kPa ± 81, p= 0.04). The tensile modulus for the sham group was 195kPa ± 56, indicating ESA injection results in a less stiff ventricle. Conclusions Direct injection of ESA alters the biomechanical response to MI, improving the mechanical properties in the post-infarct heart. PMID:23244259

  4. Central Actions of the Chemokine Stromal Cell-Derived Factor-1 Contribute to Neurohumoral Excitation in Heart Failure Rats

    PubMed Central

    Wei, Shun-Guang; Zhang, Zhi-Hua; Yu, Yang; Weiss, Robert M.; Felder, Robert B.

    2012-01-01

    The ample expression of chemokines and their receptors by neurons in the brain suggests that they play a functional role beyond the coordination of inflammatory and immune responses. Growing evidence implicates brain chemokines in the regulation of neuronal activity and neurohormonal release. This study examined the potential role of brain chemokines in regulating hemodynamic, sympathetic and neuroendocrine mechanisms in rats with ischemia-induced heart failure (HF). Immunohistochemical analysis revealed that the chemokine stromal cell-derived factor-1 (SDF-1)/CXCL12 was highly expressed in the hypothalamic paraventricular (PVN) and subfornical organ, and that SDF-1 expression was significantly increased in HF rats compared with sham-operated (SHAM) control rats. Intracerebroventricular (ICV) injection of SDF-1 induced substantial and long-lasting increases in blood pressure (BP), heart rate (HR) and renal sympathetic nerve activity (RSNA) in both SHAM and HF rats, but responses were exaggerated in HF rats. Bilateral microinjection of SDF-1 into the PVN also elicited exaggerated increases in BP, HR and RSNA in the HF rats. A 4-hour ICV infusion of SDF-1 increased plasma levels of arginine vasopressin (AVP), adrenocorticotropic hormone (ACTH) and norepinephrine (NE) in normal rats, responses that were prevented by pretreatment with ICV SDF-1 short-hairpin RNA (shRNA). ICV administration of SDF-1 shRNA also reduced plasma AVP, ACTH and NE levels in HF rats. These data suggest that the chemokine SDF-1, acting within the brain, plays an important role in regulating sympathetic drive, neuroendocrine release, and hemodynamic function in normal and pathophysiological conditions, and so may contribute to the neural and humoral activation in HF. PMID:22493069

  5. The CXC Chemokine Receptor 4 Ligands Ubiquitin and Stromal Cell-derived Factor-1α Function through Distinct Receptor Interactions*

    PubMed Central

    Saini, Vikas; Staren, Daniel M.; Ziarek, Joshua J.; Nashaat, Zayd N.; Campbell, Edward M.; Volkman, Brian F.; Marchese, Adriano; Majetschak, Matthias

    2011-01-01

    Recently, we identified extracellular ubiquitin as an endogenous CXC chemokine receptor (CXCR) 4 agonist. However, the receptor selectivity and molecular basis of the CXCR4 agonist activity of ubiquitin are unknown, and functional consequences of CXCR4 activation with ubiquitin are poorly defined. Here, we provide evidence that ubiquitin and the cognate CXCR4 ligand stromal cell-derived factor (SDF)-1α do not share CXCR7 as a receptor. We further demonstrate that ubiquitin does not utilize the typical two-site binding mechanism of chemokine-receptor interactions, in which the receptor N terminus is important for ligand binding. CXCR4 activation with ubiquitin and SDF-1α lead to similar Gαi-responses and to a comparable magnitude of phosphorylation of ERK-1/2, p90 ribosomal S6 kinase-l and Akt, although phosphorylations occur more transiently after activation with ubiquitin. Despite the similarity of signal transduction events after activation of CXCR4 with both ligands, ubiquitin possesses weaker chemotactic activity than SDF-lα in cell migration assays and does not interfere with productive entry of HIV-1 into P4.R5 multinuclear activation of galactosidase indicator cells. Unlike SDF-1α, ubiquitin lacks interactions with an N-terminal CXCR4 peptide in NMR spectroscopy experiments. Binding and signaling studies in the presence of antibodies against the N terminus and extracellular loops 2/3 of CXCR4 confirm that the ubiquitin CXCR4 interaction is independent of the N-terminal receptor domain, whereas blockade of extracellular loops 2/3 prevents receptor binding and activation. Our findings define ubiquitin as a CXCR4 agonist, which does not interfere with productive cellular entry of HIV-1, and provide new mechanistic insights into interactions between CXCR4 and its natural ligands. PMID:21757744

  6. Dermal fibroblast expression of stromal cell-derived factor-1 (SDF-1) promotes epidermal keratinocyte proliferation in normal and diseased skin.

    PubMed

    Quan, Chunji; Cho, Moon Kyun; Shao, Yuan; Mianecki, Laurel E; Liao, Eric; Perry, Daniel; Quan, Taihao

    2015-12-01

    Stromal cells provide a crucial microenvironment for overlying epithelium. Here we investigated the expression and function of a stromal cell-specific protein, stromal cell-derived factor-1 (SDF-1), in normal human skin and in the tissues of diseased skin. Immunohistology and laser capture microdissection (LCM)-coupled quantitative real-time RT-PCR revealed that SDF-1 is constitutively and predominantly expressed in dermal stromal cells in normal human skin in vivo. To our surprise, an extremely high level of SDF-1 transcription was observed in the dermis of normal human skin in vivo, evidenced by much higher mRNA expression level than type I collagen, the most abundant and highly expressed protein in human skin. SDF-1 was also upregulated in the tissues of many human skin disorders including psoriasis, basal cell carcinoma (BCC), and squamous cell carcinoma (SCC). Double immunostaining for SDF-1 and HSP47 (heat shock protein 47), a marker of fibroblasts, revealed that fibroblasts were the major source of stroma-cell-derived SDF-1 in both normal and diseased skin. Functionally, SDF-1 activates the ERK (extracellular-signal-regulated kinases) pathway and functions as a mitogen to stimulate epidermal keratinocyte proliferation. Both overexpression of SDF-1 in dermal fibroblasts and treatment with rhSDF-1 to the skin equivalent cultures significantly increased the number of keratinocyte layers and epidermal thickness. Conversely, the stimulative function of SDF-1 on keratinocyte proliferation was nearly completely eliminated by interfering with CXCR4, a specific receptor of SDF-1, or by knock-down of SDF-1 in fibroblasts. Our data reveal that extremely high levels of SDF-1 provide a crucial microenvironment for epidermal keratinocyte proliferation in both physiologic and pathologic skin conditions.

  7. Safety of repeated transplantations of neurotrophic factors-secreting human mesenchymal stromal stem cells

    PubMed Central

    2014-01-01

    Background Therapies based on mesenchymal stem cells (MSC) have been shown to have potential benefit in several clinical studies. We have shown that, using a medium-based approach, MSC can be induced to secrete elevated levels of neurotropic factors, which have been shown to have protective effects in animal models of neurodegenerative diseases. These cells, designated MSC-NTF cells (Neurotrophic factor-secreting MSC, also known as NurOwn™) derived from the patient's own bone marrow, have been recently used for Phase I/II and Phase IIa clinical studies in patients with Amyotrophic Lateral Sclerosis (ALS). In these studies, ALS patients were subjected to a single administration of autologous MSC-NTF cells. The data from these studies indicate that the single administration of MSC-NTF cells is safe and well tolerated. In a recently published case report, it was shown that repeated MSC-NTF injections in an ALS patient treated on a compassionate basis were safe and well tolerated [Muscle Nerve 49:455-457, 2014]. Methods In the current study we studied the toxicity and tolerability of three consecutive intramuscular injections (IM) of cryopreserved human MSC-NTF cells in C57BL/B6 mice to investigate the effect of repeated administration of these cells. Results Monitoring of clinical signs and immune reactions showed that repeated injections of the cells did not lead to any serious adverse events. Pathology, histology and blood biochemistry parameters tested were found to be within normal ranges with no sign of tumor formation. Conclusions Based on these results we conclude that repeated injections of human MSC-NTF are well tolerated in mice. The results of this study suggest that if the outcomes of additional clinical studies point to the need for repeated treatments, such option can be considered safe. PMID:25097724

  8. Peripheral Blood-Derived Mesenchymal Stromal Cells Promote Angiogenesis via Paracrine Stimulation of Vascular Endothelial Growth Factor Secretion in the Equine Model

    PubMed Central

    Bussche, Leen

    2014-01-01

    Mesenchymal stromal cells (MSCs) have received much attention as a potential treatment of ischemic diseases, including ischemic tissue injury and cardiac failure. The beneficial effects of MSCs are thought to be mediated by their ability to provide proangiogenic factors, creating a favorable microenvironment that results in neovascularization and tissue regeneration. To study this in more detail and to explore the potential of the horse as a valuable translational model, the objectives of the present study were to examine the presence of angiogenic stimulating factors in the conditioned medium (CM) of peripheral blood-derived equine mesenchymal stromal cells (PB-MSCs) and to study their in vitro effect on angiogenesis-related endothelial cell (EC) behavior, including proliferation and vessel formation. Our salient findings were that CM from PB-MSCs contained significant levels of several proangiogenic factors. Furthermore, we found that CM could induce angiogenesis in equine vascular ECs and confirmed that endothelin-1, insulin growth factor binding protein 2, interleukin-8, and platelet-derived growth factor-AA, but not urokinase-type plasminogen activator, were responsible for this enhanced EC network formation by increasing the expression level of vascular endothelial growth factor-A, an important angiogenesis stimulator. PMID:25313202

  9. Human decidual stromal cells secrete soluble pro-apoptotic factors during decidualization in a cAMP-dependent manner.

    PubMed

    Leno-Durán, E; Ruiz-Magaña, M J; Muñoz-Fernández, R; Requena, F; Olivares, E G; Ruiz-Ruiz, C

    2014-10-10

    Is there a relationship between decidualization and apoptosis of decidual stromal cells (DSC)? Decidualization triggers the secretion of soluble factors that induce apoptosis in DSC. The differentiation and apoptosis of DSC during decidualization of the receptive decidua are crucial processes for the controlled invasion of trophoblasts in normal pregnancy. Most DSC regress in a time-dependent manner, and their removal is important to provide space for the embryo to grow. However, the mechanism that controls DSC death is poorly understood. The apoptotic response of DSC was analyzed after exposure to different exogenous agents and during decidualization. The apoptotic potential of decidualized DSC supernatants and prolactin (PRL) was also evaluated. DSC lines were established from samples of decidua from first trimester pregnancies. Apoptosis was assayed by flow cytometry. PRL production, as a marker of decidualization, was determined by enzyme-linked immunosorbent assay. DSCs were resistant to a variety of apoptosis-inducing substances. Nevertheless, DSC underwent apoptosis during decidualization in culture, with cAMP being essential for both apoptosis and differentiation. In addition, culture supernatants from decidualized DSC induced apoptosis in undifferentiated DSC, although paradoxically these supernatants decreased the spontaneous apoptosis of decidual lymphocytes. Exogenously added PRL did not induce apoptosis in DSC and an antibody that neutralized the PRL receptor did not decrease the apoptosis induced by supernatants. Further studies are needed to examine the involvement of other soluble factors secreted by decidualized DSC in the induction of apoptosis. The present results indicate that apoptosis of DSC occurs in parallel to differentiation, in response to decidualization signals, with soluble factors secreted by decidualized DSC being responsible for triggering cell death. These studies are relevant in the understanding of how the regression of decidua

  10. Granulocyte colony-stimulating factor inhibits CXCR4/SDF-1α signaling and overcomes stromal-mediated drug resistance in the HL-60 cell line.

    PubMed

    Sheng, Xianfu; Zhong, Hua; Wan, Haixia; Zhong, Jihua; Chen, Fangyuan

    2016-07-01

    Combining cytarabine, aclarubicin and granulocyte colony-stimulating factor (G-CSF) has demonstrated marked efficacy in the treatment of elderly and relapsed/refractory patients with acute myeloid leukemia (AML); however, the role of G-CSF remains poorly understood. The present study aimed to investigate the ability of G-CSF to overcome stromal-mediated drug resistance and the underlying molecular mechanism. Two types of co-culture models were established in the HS-5 human bone marrow/stromal and HL-60 human promyelocytic leukemia cell lines, in order to imitate the interactions between stromal and leukemia cells in vitro, which is mediated by the stromal cell-derived factor (SDF)-1α signaling axis. In the present study, HL-60 cells were attracted and adhered to HS-5 cells using migration assay and flow cytometry, respectively; however, these interactions were inhibited by treatment with G-CSF and/or the C-X-C chemokine receptor type 4 (CXCR4) antagonist, AMD3100. Co-culture with HS-5 cells, including direct and indirect contact, protected HL-60 cells against spontaneous apoptosis or drug-induced apoptosis; however, these protective effects were disrupted by treatment with G-CSF and/or AMD3100. Notably, G-CSF and/or AMD3100 did not alter cell viability or apoptosis when HL-60 cells were cultured with medium alone. In addition, G-CSF significantly reduced the expression levels of surface CXCR4 protein, total CXCR4 protein and CXCR4 mRNA, and significantly upregulated the expression of microRNA (miR)-146a. Conversely, AMD3100 significantly reduced surface CXCR4 expression levels, but not the total CXCR4, CXCR4 mRNA or miR-146a expression levels. The results of the present study suggested that interfering with the CXCR4/SDF-1α signaling axis via G-CSF inhibited the migration and adhesion of HL-60 cells to HS-5 cells and eliminated HS5 cell-mediated protective effects. Furthermore, G-CSF administration reduced CXCR4 expression levels by upregulating the expression of

  11. Effect of stromal-cell-derived factor 1 on stem-cell homing and tissue regeneration in ischaemic cardiomyopathy

    NASA Technical Reports Server (NTRS)

    Askari, Arman T.; Unzek, Samuel; Popovic, Zoran B.; Goldman, Corey K.; Forudi, Farhad; Kiedrowski, Matthew; Rovner, Aleksandr; Ellis, Stephen G.; Thomas, James D.; DiCorleto, Paul E.; hide

    2003-01-01

    BACKGROUND: Myocardial regeneration via stem-cell mobilisation at the time of myocardial infarction is known to occur, although the mechanism for stem-cell homing to infarcted tissue subsequently and whether this approach can be used for treatment of ischaemic cardiomyopathy are unknown. We investigated these issues in a Lewis rat model (ligation of the left anterior descending artery) of ischaemic cardiomyopathy. METHODS: We studied the effects of stem-cell mobilisation by use of granulocyte colony-stimulating factor (filgrastim) with or without transplantation of syngeneic cells. Shortening fraction and myocardial strain by tissue doppler imaging were quantified by echocardiography. FINDINGS: Stem-cell mobilisation with filgrastim alone did not lead to engraftment of bone-marrow-derived cells. Stromal-cell-derived factor 1 (SDF-1), required for stem-cell homing to bone marrow, was upregulated immediately after myocardial infarction and downregulated within 7 days. 8 weeks after myocardial infarction, transplantation into the peri-infarct zone of syngeneic cardiac fibroblasts stably transfected to express SDF-1 induced homing of CD117-positive stem cells to injured myocardium after filgrastim administration (control vs SDF-1-expressing cardiac fibroblasts mean 7.2 [SD 3.4] vs 33.2 [6.0] cells/mm2, n=4 per group, p<0.02) resulting in greater left-ventricular mass (1.24 [0.29] vs 1.57 [0.27] g) and better cardiac function (shortening fraction 9.2 [4.9] vs 17.2 [4.2]%, n=8 per group, p<0.05). INTERPRETATION: These findings show that SDF-1 is sufficient to induce therapeutic stem-cell homing to injured myocardium and suggest a strategy for directed stem-cell engraftment into injured tissues. Our findings also indicate that therapeutic strategies focused on stem-cell mobilisation for regeneration of myocardial tissue must be initiated within days of myocardial infarction unless signalling for stem-cell homing is re-established.

  12. Effect of stromal-cell-derived factor 1 on stem-cell homing and tissue regeneration in ischaemic cardiomyopathy

    NASA Technical Reports Server (NTRS)

    Askari, Arman T.; Unzek, Samuel; Popovic, Zoran B.; Goldman, Corey K.; Forudi, Farhad; Kiedrowski, Matthew; Rovner, Aleksandr; Ellis, Stephen G.; Thomas, James D.; DiCorleto, Paul E.; Topol, Eric J.; Penn, Marc S.

    2003-01-01

    BACKGROUND: Myocardial regeneration via stem-cell mobilisation at the time of myocardial infarction is known to occur, although the mechanism for stem-cell homing to infarcted tissue subsequently and whether this approach can be used for treatment of ischaemic cardiomyopathy are unknown. We investigated these issues in a Lewis rat model (ligation of the left anterior descending artery) of ischaemic cardiomyopathy. METHODS: We studied the effects of stem-cell mobilisation by use of granulocyte colony-stimulating factor (filgrastim) with or without transplantation of syngeneic cells. Shortening fraction and myocardial strain by tissue doppler imaging were quantified by echocardiography. FINDINGS: Stem-cell mobilisation with filgrastim alone did not lead to engraftment of bone-marrow-derived cells. Stromal-cell-derived factor 1 (SDF-1), required for stem-cell homing to bone marrow, was upregulated immediately after myocardial infarction and downregulated within 7 days. 8 weeks after myocardial infarction, transplantation into the peri-infarct zone of syngeneic cardiac fibroblasts stably transfected to express SDF-1 induced homing of CD117-positive stem cells to injured myocardium after filgrastim administration (control vs SDF-1-expressing cardiac fibroblasts mean 7.2 [SD 3.4] vs 33.2 [6.0] cells/mm2, n=4 per group, p<0.02) resulting in greater left-ventricular mass (1.24 [0.29] vs 1.57 [0.27] g) and better cardiac function (shortening fraction 9.2 [4.9] vs 17.2 [4.2]%, n=8 per group, p<0.05). INTERPRETATION: These findings show that SDF-1 is sufficient to induce therapeutic stem-cell homing to injured myocardium and suggest a strategy for directed stem-cell engraftment into injured tissues. Our findings also indicate that therapeutic strategies focused on stem-cell mobilisation for regeneration of myocardial tissue must be initiated within days of myocardial infarction unless signalling for stem-cell homing is re-established.

  13. Thermosensitive chitosan-based hydrogels releasing stromal cell derived factor-1 alpha recruit MSC for corneal epithelium regeneration.

    PubMed

    Tang, Qiaomei; Luo, Chenqi; Lu, Bing; Fu, Qiuli; Yin, Houfa; Qin, Zhenwei; Lyu, Danni; Zhang, Lifang; Fang, Zhi; Zhu, Yanan; Yao, Ke

    2017-10-01

    Corneal epithelium integrity depends on continuous self-renewing of epithelium and connections between adjacent cells or between the cells and the basement membrane. Self-renewing epithelium cells mainly arise from the continuous proliferation and differentiation of the basal layer and limbal stem cells. The aim of the present study was to generate a bioactive, thermosensitive chitosan-gelatin hydrogel (CHI hydrogel) by incorporating exogenous recombinant human stromal cell-derived factor-1 alpha (SDF-1 alpha) for corneal epithelium regeneration. The exogenous SDF-1 alpha could enhance the stem cells proliferation, chemotaxis and migration, and the expression levels of related genes were significantly elevated in LESCs and mesenchymal stem cells (MSCs) in vitro. Moreover, the MSCs promoted the proliferation and maintained the corneal fate of the LESCs. The rat alkali injury model was used for in vivo study. The injured eyes were covered with CHI hydrogel alone or rhSDF-1 alpha-loaded CHI hydrogel. All rats were followed for 13days. Histological examination showed that the SDF-1 alpha/CHI hydrogel complex group had a nearly normal thickness; moreover, it was also found that this group could upregulate the expression of some genes and had more ΔNp63-positive cells. The SDF-1 alpha/CHI hydrogel complex group had a more tightly arranged epithelium compared with the control group using transmission electron microscopy (TEM). The mechanism for this may have involved the activation of stem cell homing and the secretion of growth factors via the SDF-1/CXCR4 chemokine axis. Therefore, SDF-1 alpha/CHI hydrogel complexes could provide a new idea for the clinical application. The clarity of cornea is important for normal vision. The loss or dysfunction of LESCs leads to the impairment of corneal epithelium. The complete regeneration of corneal epithelium has not been achieved. Our study demonstrated that the incorporation of rhSDF-1 alpha with CHI hydrogel accelerated corneal

  14. Chemokines, macrophage inflammatory protein-2 and stromal cell-derived factor-1{alpha}, suppress amyloid {beta}-induced neurotoxicity

    SciTech Connect

    Raman, Dayanidhi; Milatovic, Snjezana-Zaja; Milatovic, Dejan; Fan, Guo-Huang; Richmond, Ann

    2011-11-15

    Alzheimer's disease (AD) is characterized by a progressive cognitive decline and accumulation of neurotoxic oligomeric peptides amyloid-{beta} (A{beta}). Although the molecular events are not entirely known, it has become evident that inflammation, environmental and other risk factors may play a causal, disruptive and/or protective role in the development of AD. The present study investigated the ability of the chemokines, macrophage inflammatory protein-2 (MIP-2) and stromal cell-derived factor-1{alpha} (SDF-1{alpha}), the respective ligands for chemokine receptors CXCR2 and CXCR4, to suppress A{beta}-induced neurotoxicity in vitro and in vivo. Pretreatment with MIP-2 or SDF-1{alpha} significantly protected neurons from A{beta}-induced dendritic regression and apoptosis in vitro through activation of Akt, ERK1/2 and maintenance of metalloproteinase ADAM17 especially with SDF-1{alpha}. Intra-cerebroventricular (ICV) injection of A{beta} led to reduction in dendritic length and spine density of pyramidal neurons in the CA1 area of the hippocampus and increased oxidative damage 24 h following the exposure. The A{beta}-induced morphometric changes of neurons and increase in biomarkers of oxidative damage, F{sub 2}-isoprostanes, were significantly inhibited by pretreatment with the chemokines MIP-2 or SDF-1{alpha}. Additionally, MIP-2 or SDF-1{alpha} was able to suppress the aberrant mislocalization of p21-activated kinase (PAK), one of the proteins involved in the maintenance of dendritic spines. Furthermore, MIP-2 also protected neurons against A{beta} neurotoxicity in CXCR2-/- mice, potentially through observed up regulation of CXCR1 mRNA. Understanding the neuroprotective potential of chemokines is crucial in defining the role for their employment during the early stages of neurodegeneration. -- Research highlights: Black-Right-Pointing-Pointer Neuroprotective ability of the chemokines MIP2 and CXCL12 against A{beta} toxicity. Black-Right-Pointing-Pointer MIP-2 or

  15. Low Intensity Pulsed Ultrasound Enhanced Mesenchymal Stem Cell Recruitment through Stromal Derived Factor-1 Signaling in Fracture Healing

    PubMed Central

    Wei, Fang-Yuan; Leung, Kwok-Sui; Li, Gang; Qin, Jianghui; Chow, Simon Kwoon-Ho; Huang, Shuo; Sun, Ming-Hui; Qin, Ling; Cheung, Wing-Hoi

    2014-01-01

    Low intensity pulsed ultrasound (LIPUS) has been proven effective in promoting fracture healing but the underlying mechanisms are not fully depicted. We examined the effect of LIPUS on the recruitment of mesenchymal stem cells (MSCs) and the pivotal role of stromal cell-derived factor-1/C-X-C chemokine receptor type 4 (SDF-1/CXCR4) pathway in response to LIPUS stimulation, which are essential factors in bone fracture healing. For in vitro study, isolated rat MSCs were divided into control or LIPUS group. LIPUS treatment was given 20 minutes/day at 37°C for 3 days. Control group received sham LIPUS treatment. After treatment, intracellular CXCR4 mRNA, SDF-1 mRNA and secreted SDF-1 protein levels were quantified, and MSCs migration was evaluated with or without blocking SDF-1/CXCR4 pathway by AMD3100. For in vivo study, fractured 8-week-old young rats received intracardiac administration of MSCs were assigned to LIPUS treatment, LIPUS+AMD3100 treatment or vehicle control group. The migration of transplanted MSC to the fracture site was investigated by ex vivo fluorescent imaging. SDF-1 protein levels at fracture site and in serum were examined. Fracture healing parameters, including callus morphology, micro-architecture of the callus and biomechanical properties of the healing bone were investigated. The in vitro results showed that LIPUS upregulated SDF-1 and CXCR4 expressions in MSCs, and elevated SDF-1 protein level in the conditioned medium. MSCs migration was promoted by LIPUS and partially inhibited by AMD3100. In vivo study demonstrated that LIPUS promoted MSCs migration to the fracture site, which was associated with an increase of local and serum SDF-1 level, the changes in callus formation, and the improvement of callus microarchitecture and mechanical properties; whereas the blockade of SDF-1/CXCR4 signaling attenuated the LIPUS effects on the fractured bones. These results suggested SDF-1 mediated MSCs migration might be one of the crucial mechanisms

  16. Baicalein reduces endometriosis by suppressing the viability of human endometrial stromal cells through the nuclear factor-κB pathway in vitro.

    PubMed

    Jin, Zhixing; Huang, Jianqin; Zhu, Zhiling

    2017-10-01

    The aim of the present study was to evaluate the effects of baicalein on human endometrial stromal cells in vitro. Ectopic endometrium samples were obtained from 6 female patients with ovarian endometriosis who underwent laparoscopic surgical procedures from July to September 2015. After culturing the cells, immunocytochemistry was performed to verify the purity and homogeneity of the endometrial stromal cells, and a Cell Counting Kit-8 assay was used to evaluate cell viability. In addition, cell cycle progression was analyzed using flow cytometry, and the effects of baicalein on the expression of B-cell lymphoma 2 (Bcl-2), Bcl-2-associated X protein (Bax), proliferating cell nuclear antigen (PCNA) and cyclin D1 in endometrial stromal cells were evaluated using western blot analysis. The related signaling pathways were also investigated by incubating cells with inhibitors of signaling pathways, prior to adding 40 µM baicalein for 48 h, followed by analysis of cell viability using a Cell Counting Kit-8 assay. The results indicated that treatment with baicalein significantly induced a dose-dependent decrease (P<0.05) in the viability of human endometrial stromal cells, which was abolished by inhibition of the nuclear factor (NF)-κB signaling pathway. However, baicalein treatment did not induce a time-dependent decrease in viability, as cell viabilities between the 24, 48 and 72 h treatment groups did not differ significantly. The number of cells in the G0/G1 phase significantly increased following treatment with baicalein (P<0.05), while the number of cells in the S and G2/M phases significantly decreased (P<0.05). Baicalein-treated cells also exhibited significantly reduced expression of Bcl-2, PCNA and cyclin D1 compared with control cells (P<0.05). These results suggested that baicalein may suppress the viability of human endometrial stromal cells through the NF-κB signaling pathway in vitro, and may induce apoptosis and promote cell cycle arrest at the G0/G1

  17. A murine uterine transcriptome, responsive to steroid receptor coactivator-2, reveals transcription factor 23 as essential for decidualization of human endometrial stromal cells.

    PubMed

    Kommagani, Ramakrishna; Szwarc, Maria M; Kovanci, Ertug; Creighton, Chad J; O'Malley, Bert W; Demayo, Francesco J; Lydon, John P

    2014-04-01

    Recent data from human and mouse studies strongly support an indispensable role for steroid receptor coactivator-2 (SRC-2)-a member of the p160/SRC family of coregulators-in progesterone-dependent endometrial stromal cell decidualization, an essential cellular transformation process that regulates invasion of the developing embryo into the maternal compartment. To identify the key progesterone-induced transcriptional changes that are dependent on SRC-2 and required for endometrial decidualization, we performed comparative genome-wide transcriptional profiling of endometrial tissue RNA from ovariectomized SRC-2(flox/flox) (SRC-2(f/f) [control]) and PR(cre/+)/SRC-2(flox/flox) (SRC-2(d/d) [SRC-2-depleted]) mice, acutely treated with vehicle or progesterone. Although data mining revealed that only a small subset of the total progesterone-dependent transcriptional changes is dependent on SRC-2 (∼13%), key genes previously reported to mediate progesterone-driven endometrial stromal cell decidualization are present within this subset. Along with providing a more detailed molecular portrait of the decidual transcriptional program governed by SRC-2, the degree of functional diversity of these progesterone mediators underscores the pleiotropic regulatory role of SRC-2 in this tissue. To showcase the utility of this powerful informational resource to uncover novel signaling paradigms, we stratified the total SRC-2-dependent subset of progesterone-induced transcriptional changes in terms of novel gene expression and identified transcription factor 23 (Tcf23), a basic-helix-loop-helix transcription factor, as a new progesterone-induced target gene that requires SRC-2 for full induction. Importantly, using primary human endometrial stromal cells in culture, we demonstrate that TCF23 function is essential for progesterone-dependent decidualization, providing crucial translational support for this transcription factor as a new decidual mediator of progesterone action.

  18. Transforming growth factor β-activated kinase 1 negatively regulates interleukin-1α-induced stromal-derived factor-1 expression in vascular smooth muscle cells

    SciTech Connect

    Yang, Bin; Li, Wei; Zheng, Qichang; Qin, Tao; Wang, Kun; Li, Jinjin; Guo, Bing; Yu, Qihong; Wu, Yuzhe; Gao, Yang; Cheng, Xiang; Hu, Shaobo; Kumar, Stanley Naveen; Liu, Sanguang; Song, Zifang

    2015-07-17

    Stromal-derived Factor-1 (SDF-1) derived from vascular smooth muscle cells (VSMCs) contributes to vascular repair and remodeling in various vascular diseases. In this study, the mechanism underlying regulation of SDF-1 expression by interleukin-1α (IL-1α) was investigated in primary rat VSMCs. We found IL-1α promotes SDF-1 expression by up-regulating CCAAT-enhancer-binding protein β (C/EBPβ) in an IκB kinase β (IKKβ) signaling-dependent manner. Moreover, IL-1α-induced expression of C/EBPβ and SDF-1 was significantly potentiated by knockdown of transforming growth factor β-activated kinase 1 (TAK1), an upstream activator of IKKβ signaling. In addition, we also demonstrated that TAK1/p38 mitogen-activated protein kinase (p38 MAPK) signaling exerted negative effect on IL-1α-induced expression of C/EBPβ and SDF-1 through counteracting ROS-dependent up-regulation of nuclear factor erythroid 2-related factor 2 (NRF2). In conclusion, TAK1 acts as an important regulator of IL-1α-induced SDF-1 expression in VSMCs, and modulating activity of TAK1 may serve as a potential strategy for modulating vascular repair and remodeling. - Highlights: • IL-1α induces IKKβ signaling-dependent SDF-1 expression by up-regulating C/EBPβ. • Activation of TAK1 by IL-1α negatively regulates C/EBPβ-dependent SDF-1 expression. • IL-1α-induced TAK1/p38 MAPK signaling counteracts ROS-dependent SDF-1 expression. • TAK1 counteracts IL-1α-induced SDF-1 expression by attenuating NRF2 up-regulation.

  19. [Culturing and characterization of human gingival mesenchymal stem cells and their chemotactic responses to stromal cell-derived factor-1].

    PubMed

    Du, Lingqian; Yang, Pishan; Ge, Shaohua

    2015-06-01

    To investigate the expression of chemokine stromal cell-derived factor-1 (SDF-1) receptor CXCR4 in human gingival mesenchymal stem cells (GMSCs) and the migration potential of GMSCs stimulated with SDF-1. Human GMSCs were isolated by single-cell cloning method. Their cell surface markers were characterized by flow cytometry, and the rate of colony formation was evaluated. Differentiation assay was used to detect the differentiation potential of GMSCs. The expression of chemokine SDF-l receptor CXCR4 in GMSCs was detected by immunocytochemical staining. The chemotactic effect of SDF-1 on GMSCs was detected using a 24-multiwell Transwell cell culture chamber. The number of net migrated cells was counted in different microscope fields. Human GMSCs possessed high self-renewal potential and formed single-cell colonies cultured in vitro. GMSCs expressed mesenchymal stem cells-associated markers CD44, CD73, CD90, CD105, and CD166, and the expression of hemopoietic stem cell surface markers CD14, CD34, and CD45 was negative. GMSCs differentiated into osteoblasts and adipocytes under defined culture conditions. The colony forming unit-fibroblastic for GMSCs was 21.4%/±2.8%. Immunocytochemical staining demonstrated that GMSCs expressed chemokine SDF-1 receptor CXCR4. The number of GMSCs migrating at concentrations of 100 ng.mL-1 and 200 ng.mL-1 of SDF-l in the Transwell cell culture chamber was significantly higher than that of the negative control (189.3±4.4, 164.6±4.9 cells/field vs. 47.8±2.5 cells/field, P<0.01). Treatment with the CXCR4 neutralizing antibody, an antagonist for CXCR4, significantly reduced the migratory effect compared with the negative controls (29.0±2.4 cells/field vs. 47.8±2.5 cells/field, P<0.01). Human GMSCs express chemokine SDF-l receptor CXCR4. SDF-1 may participate in regulating chemotaxis of human GMSCs. Results suggest that the migration induced by SDF-1 is mediated by CXCR4.

  20. Mathematically engineered stromal cell-derived factor-1α stem cell cytokine analog enhances mechanical properties of infarcted myocardium.

    PubMed

    MacArthur, John W; Trubelja, Alen; Shudo, Yasuhiro; Hsiao, Philip; Fairman, Alexander S; Yang, Elaine; Hiesinger, William; Sarver, Joseph J; Atluri, Pavan; Woo, Y Joseph

    2013-01-01

    The biomechanical response to a myocardial infarction consists of ventricular remodeling that leads to dilatation, loss of contractile function, abnormal stress patterns, and ultimately heart failure. We hypothesized that intramyocardial injection of our previously designed pro-angiogenic chemokine, an engineered stromal cell-derived factor-1α analog (ESA), improves mechanical properties of the heart after infarction. Male rats (n = 54) underwent either sham surgery (n = 17) with no coronary artery ligation or ligation of the left anterior descending artery (n = 37). The rats in the myocardial infarction group were then randomized to receive either saline (0.1 mL, n = 18) or ESA (6 μg/kg, n = 19) injected into the myocardium at 4 predetermined spots around the border zone. Echocardiograms were performed preoperatively and before the terminal surgery. After 4 weeks, the hearts were explanted and longitudinally sectioned. Uniaxial tensile testing was completed using an Instron 5543 Microtester. Optical strain was evaluated using custom image acquisition software, Digi-Velpo, and analyzed in MATLAB. Compared with the saline control group at 4 weeks, the ESA-injected hearts had a greater ejection fraction (71.8% ± 9.0% vs 55.3% ± 12.6%, P = .0004), smaller end-diastolic left ventricular internal dimension (0.686 ± 0.110 cm vs 0.763 ± 0.160 cm, P = .04), greater cardiac output (36 ± 11.6 mL/min vs 26.9 ± 7.3 mL/min, P = .05), and a lower tensile modulus (251 ± 56 kPa vs 301 ± 81 kPa, P = .04). The tensile modulus for the sham group was 195 ± 56 kPa, indicating ESA injection results in a less stiff ventricle. Direct injection of ESA alters the biomechanical response to myocardial infarction, improving the mechanical properties in the postinfarct heart. Copyright © 2013 The American Association for Thoracic Surgery. Published by Mosby, Inc. All rights reserved.

  1. Primordial germ cell migration in the yellowtail kingfish (Seriola lalandi) and identification of stromal cell-derived factor 1.

    PubMed

    Fernández, J A; Bubner, E J; Takeuchi, Y; Yoshizaki, G; Wang, T; Cummins, S F; Elizur, A

    2015-03-01

    Primordial germ cells (PGCs) are progenitors of the germ cell lineage, giving rise to either spermatogonia or oogonia after the completion of gonadal differentiation. Currently, there is little information on the mechanism of PGCs migration leading to the formation of the primordial gonad in perciform fish. Yellowtail kingfish (Seriola lalandi) (YTK) (order Perciforms) inhabit tropical and temperate waters in the southern hemisphere. Fundamental details into the molecular basis of larval development in this species can be easily studied in Australia, as they are commercially cultured and readily available. In this study, histological analysis of YTK larvae revealed critical time points for the migration of PGCs to the genital ridge, resulting in the subsequent development of the primordial gonad. In YTK larvae at 3, 5, 7 and 10 days post hatch (DPH), PGCs were not yet enclosed by somatic cells, indicating the primordial gonad had not yet started to form. While at 15, 18 and 20 DPH PGCs had already settled at the genital ridge and started to become enclosed by somatic cells indicating the primordial gonad had started to develop. A higher number of PGCs were observed in the larvae at 15 and 18 DPH indicating PGCs proliferation, which corresponds with them becoming enclosed by the somatic cells. Directional migration of PGCs toward the genital ridge is a critical event in the subsequent development of a gonad. In zebrafish, mouse and chicken, stromal-cell derived factor (SDF1) signalling is one of the key molecules for PGC migration. We subsequently isolated from YTK the SDF1 (Slal-SDF1) gene, which encodes for a 98-residue precursor protein with a signal peptide at the N-terminus. There is spatial conservation between fish species of four cysteine residues at positions C9, C11, C34 and C49, expected to form disulphide bonds and stabilize the SDF structure. In YTK, Slal-SDF1 gene expression analyses shows that this gene is expressed in larvae from 1 to 22 DPH and

  2. 50 Years in the sun of Bürgenstock--on the success factors of a famous conference.

    PubMed

    Müller, Klaus

    2015-04-20

    The secret of success: This year the famous "Bürgenstock Conference" will take place for the 50th time. This conference has become internationally one of the, if not the, most highly regarded conference in chemistry, chemical biology, and physical chemistry. What are the success factors of this conference? These as well as a number of perhaps more hidden figures and facts are discussed. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  3. Stromal derived factor‐1 and granulocyte‐colony stimulating factor treatment improves regeneration of Pax7−/− mice skeletal muscles

    PubMed Central

    Kowalski, Kamil; Archacki, Rafał; Archacka, Karolina; Stremińska, Władysława; Paciorek, Anna; Gołąbek, Magdalena; Ciemerych, Maria A.

    2015-01-01

    Abstract Background The skeletal muscle has the ability to regenerate after injury. This process is mediated mainly by the muscle specific stem cells, that is, satellite cells. In case of extensive damage or under pathological conditions, such as muscular dystrophy, the process of muscle reconstruction does not occur properly. The aim of our study was to test whether mobilized stem cells, other than satellite cells, could participate in skeletal muscle reconstruction. Methods Experiments were performed on wild‐type mice and mice lacking the functional Pax7 gene, that is, characterized by the very limited satellite cell population. Gastrocnemius mice muscles were injured by cardiotoxin injection, and then the animals were treated by stromal derived factor‐1 (Sdf‐1) with or without granulocyte‐colony stimulating factor (G‐CSF) for 4 days. The muscles were subjected to thorough assessment of the tissue regeneration process using histological and in vitro methods, as well as evaluation of myogenic factors' expression at the transcript and protein levels. Results Stromal derived factor‐1 alone and Sdf‐1 in combination with G‐CSF significantly improved the regeneration of Pax7−/− skeletal muscles. The Sdf‐1 and G‐CSF treatment caused an increase in the number of mononucleated cells associated with muscle fibres. Further analysis showed that Sdf‐1 and G‐CSF treatment led to the rise in the number of CD34+ and Cxcr4+ cells and expression of Cxcr7. Conclusions Stromal derived factor‐1 and G‐CSF stimulated regeneration of the skeletal muscles deficient in satellite cells. We suggest that mobilized CD34+, Cxcr4+, and Cxcr7+ cells can efficiently participate in the skeletal muscle reconstruction and compensate for the lack of satellite cells. PMID:27239402

  4. Piloting Vertical Flight Aircraft: A Conference on Flying Qualities and Human Factors

    NASA Technical Reports Server (NTRS)

    Blanken, Christopher L. (Editor); Whalley, Matthew S. (Editor)

    1993-01-01

    This document contains papers from a specialists' meeting entitled 'Piloting Vertical Flight Aircraft: A Conference on Flying Qualities and Human Factors.' Vertical flight aircraft, including helicopters and a variety of Vertical Takeoff and Landing (VTOL) concepts, place unique requirements on human perception, control, and performance for the conduct of their design missions. The intent of this conference was to examine, for these vehicles, advances in: (1) design of flight control systems for ADS-33C standards; (2) assessment of human factors influences of cockpit displays and operational procedures; (3) development of VTOL design and operational criteria; and (4) development of theoretical methods or models for predicting pilot/vehicle performance and mission suitability. A secondary goal of the conference was to provide an initial venue for enhanced interaction between human factors and handling qualities specialists.

  5. Nerve growth factor is involved in the supportive effect by bone marrow--derived stromal cells of the factor-dependent human cell line UT-7.

    PubMed

    Auffray, I; Chevalier, S; Froger, J; Izac, B; Vainchenker, W; Gascan, H; Coulombel, L

    1996-09-01

    We previously demonstrated that murine MS-5 and SI/SI4 cell lines induce the proliferation of human factor-dependent UT-7 cells in the absence of normally required human cytokines and also stimulate the differentiation of CD34+/CD38-LTC-ICs. We report in this study that the effect of MS-5 cells on UT-7 cells can be completely explained by the synergistic action of nerve growth factor (NGF) and stem cell factor (SCF) produced by these murine stromal cells. Purified murine NGF was able to support short-term clone formation and long-term growth of UT-7 cells in suspension cultures as efficiently as rhu-granulocyte-macrophage colony-stimulating factor. NGF action was mediated through the TrkA receptor, in which messenger RNA (mRNA) was easily detected in UT-7 cells by Northern blot. MS-5 cells strongly expressed NGF mRNA in Northern blot, and direct implication of MS-5-derived NGF in the induction of UT-7 cells proliferation was demonstrated in inhibition assays with an anti-NGF monoclonal antibody (MoAb) that neutralized by 84% +/- 4.1% (n = 5) UT-7 clone formation. However, NGF did not act alone, and several arguments demonstrated the synergistic action of MS-5-derived SCF: (1) an anti-c-kit partially inhibited UT-7 cells clone formation in coculture assays, (2) SCF and NGF synergized in an H3-TdR incorporation assay, and (3) the stimulatory effect of 10x-concentrated MS-5 supernatant was completely inhibited by an anti-c-kit but not by an anti-NGF, and levels of soluble NGF (1.2 ng/mL) detected by enzyme-linked immunosorbent assay in 10x supernatant of MS-5 cells cultures were below the biologically active concentrations. In contrast, although MS-5 cells also promoted the differentiation of very primitive CD34+/CD38- human stem cells both in colony assays and long-term cultures, we could not incriminate MS-5-derived NGF in the observed effect: an anti-NGF MoAb did not inhibit the synergistic effect of MS-5 cells in colony assays or long-term cultures nor did soluble

  6. Gastric stromal tumor.

    PubMed

    Ovali, Gülgün Yilmaz; Tarhan, Serdar; Serter, Selim; Pabuşçu, Yüksel

    2005-06-01

    Gastric stromal tumors are rare neoplasms of the stomach. In this report we present a gastric stromal tumor with an exophytic growth pattern, and describe magnetic resonance imaging and endoscopic ultrasonography findings.

  7. Growth Factor-Activated Stem Cell Circuits and Stromal Signals Cooperatively Accelerate Non-Integrated iPSC Reprogramming of Human Myeloid Progenitors

    PubMed Central

    Park, Tea Soon; Huo, Jeffrey S.; Peters, Ann; Talbot, C. Conover; Verma, Karan; Zimmerlin, Ludovic; Kaplan, Ian M.; Zambidis, Elias T.

    2012-01-01

    Nonviral conversion of skin or blood cells into clinically useful human induced pluripotent stem cells (hiPSC) occurs in only rare fractions (∼0.001%–0.5%) of donor cells transfected with non-integrating reprogramming factors. Pluripotency induction of developmentally immature stem-progenitors is generally more efficient than differentiated somatic cell targets. However, the nature of augmented progenitor reprogramming remains obscure, and its potential has not been fully explored for improving the extremely slow pace of non-integrated reprogramming. Here, we report highly optimized four-factor reprogramming of lineage-committed cord blood (CB) myeloid progenitors with bulk efficiencies of ∼50% in purified episome-expressing cells. Lineage-committed CD33+CD45+CD34− myeloid cells and not primitive hematopoietic stem-progenitors were the main targets of a rapid and nearly complete non-integrated reprogramming. The efficient conversion of mature myeloid populations into NANOG+TRA-1-81+ hiPSC was mediated by synergies between hematopoietic growth factor (GF), stromal activation signals, and episomal Yamanaka factor expression. Using a modular bioinformatics approach, we demonstrated that efficient myeloid reprogramming correlated not to increased proliferation or endogenous Core factor expressions, but to poised expression of GF-activated transcriptional circuits that commonly regulate plasticity in both hematopoietic progenitors and embryonic stem cells (ESC). Factor-driven conversion of myeloid progenitors to a high-fidelity pluripotent state was further accelerated by soluble and contact-dependent stromal signals that included an implied and unexpected role for Toll receptor-NFκB signaling. These data provide a paradigm for understanding the augmented reprogramming capacity of somatic progenitors, and reveal that efficient induced pluripotency in other cell types may also require extrinsic activation of a molecular framework that commonly regulates self

  8. The non-alcoholic fraction of beer increases stromal cell derived factor 1 and the number of circulating endothelial progenitor cells in high cardiovascular risk subjects: a randomized clinical trial.

    PubMed

    Chiva-Blanch, Gemma; Condines, Ximena; Magraner, Emma; Roth, Irene; Valderas-Martínez, Palmira; Arranz, Sara; Casas, Rosa; Martínez-Huélamo, Miriam; Vallverdú-Queralt, Anna; Quifer-Rada, Paola; Lamuela-Raventos, Rosa M; Estruch, Ramon

    2014-04-01

    Moderate alcohol consumption is associated with a decrease in cardiovascular risk, but fermented beverages seem to confer greater cardiovascular protection due to their polyphenolic content. Circulating endothelial progenitor cells (EPC) are bone-marrow-derived stem cells with the ability to repair and maintain endothelial integrity and function and are considered as a surrogate marker of vascular function and cumulative cardiovascular risk. Nevertheless, no study has been carried out on the effects of moderate beer consumption on the number of circulating EPC in high cardiovascular risk patients. To compare the effects of moderate consumption of beer, non-alcoholic beer and gin on the number of circulating EPC and EPC-mobilizing factors. In this crossover trial, 33 men at high cardiovascular risk were randomized to receive beer (30 g alcohol/d), the equivalent amount of polyphenols in the form of non-alcoholic beer, or gin (30 g alcohol/d) for 4 weeks. Diet and physical exercise were carefully monitored. The number of circulating EPC and EPC-mobilizing factors were determined at baseline and after each intervention. After the beer and non-alcoholic beer interventions, the number of circulating EPC significantly increased by 8 and 5 units, respectively, while no significant differences were observed after the gin period. In correlation, stromal cell derived factor 1 increased significantly after the non-alcoholic and the beer interventions. The non-alcoholic fraction of beer increases the number of circulating EPC in peripheral blood from high cardiovascular risk subjects. http://www.controlled-trials.com/ISRCTN95345245 ISRCTN95345245. Copyright © 2014 Elsevier Ireland Ltd. All rights reserved.

  9. Granulocyte-Colony-Stimulating Factor Stimulation of Bone Marrow Mesenchymal Stromal Cells Promotes CD34+ Cell Migration Via a Matrix Metalloproteinase-2-Dependent Mechanism

    PubMed Central

    Ponte, Adriana López; Ribeiro-Fleury, Tatiana; Chabot, Valérie; Gouilleux, Fabrice; Langonné, Alain; Hérault, Olivier; Charbord, Pierre

    2012-01-01

    Human hematopoietic stem/progenitor cells (HSPCs) can be mobilized into the circulation using granulocyte-colony stimulating factor (G-CSF), for graft collection in view of hematopoietic transplantation. This process has been related to bone marrow (BM) release of serine proteases and of the matrix metalloproteinase-9 (MMP-9). Yet, the role of these mediators in HSC egress from their niches remains questionable, because they are produced by nonstromal cells (mainly neutrophils and monocytes/macrophages) that are not a part of the niche. We show here that the G-CSF receptor (G-CSFR) is expressed by human BM mesenchymal stromal/stem cells (MSCs), and that G-CSF prestimulation of MSCs enhances the in vitro trans-stromal migration of CD34+ cells. Zymography analysis indicates that pro-MMP-2 (but not pro-MMP-9) is expressed in MSCs, and that G-CSF treatment increases its expression and induces its activation at the cell membrane. We further demonstrate that G-CSF-stimulated migration depends on G-CSFR expression and is mediated by a mechanism that involves MMPs. These results suggest a molecular model whereby G-CSF infusion may drive, by the direct action on MSCs, HSPC egress from BM niches via synthesis and activation of MMPs. In this model, MMP-2 instead of MMP-9 is implicated, which constitutes a major difference with mouse mobilization models. PMID:22651889

  10. The plasmid-encoded regulator activates factors conferring lysozyme resistance on enteropathogenic Escherichia coli strains.

    PubMed

    Salinger, Nina; Kokona, Bashkim; Fairman, Robert; Okeke, Iruka N

    2009-01-01

    We demonstrate that enhanced lysozyme resistance of enteropathogenic Escherichia coli requires the plasmid-encoded regulator, Per, and is mediated by factors outside the locus for enterocyte effacement. EspC, a Per-activated serine protease autotransporter protein, conferred enhanced resistance on nonpathogenic E. coli, and a second Per-regulated, espC-independent lysozyme resistance mechanism was identified.

  11. The Plasmid-Encoded Regulator Activates Factors Conferring Lysozyme Resistance on Enteropathogenic Escherichia coli Strains▿

    PubMed Central

    Salinger, Nina; Kokona, Bashkim; Fairman, Robert; Okeke, Iruka N.

    2009-01-01

    We demonstrate that enhanced lysozyme resistance of enteropathogenic Escherichia coli requires the plasmid-encoded regulator, Per, and is mediated by factors outside the locus for enterocyte effacement. EspC, a Per-activated serine protease autotransporter protein, conferred enhanced resistance on nonpathogenic E. coli, and a second Per-regulated, espC-independent lysozyme resistance mechanism was identified. PMID:18997020

  12. Transforming growth factor-β synthesized by stromal cells and cancer cells participates in bone resorption induced by oral squamous cell carcinoma

    SciTech Connect

    Nakamura, Ryosuke; Kayamori, Kou; Oue, Erika; Sakamoto, Kei; Harada, Kiyoshi; Yamaguchi, Akira

    2015-03-20

    Transforming growth factor beta (TGF-β) plays a significant role in the regulation of the tumor microenvironment. To explore the role of TGF-β in oral cancer-induced bone destruction, we investigated the immunohistochemical localization of TGF-β and phosphorylated Smad2 (p-Smad2) in 12 surgical specimens of oral squamous cell carcinoma (OSCC). These studies revealed TGF-β and p-Smad2 expression in cancer cells in all tested cases. Several fibroblasts located between cancer nests and resorbing bone expressed TGF-β in 10 out of 12 cases and p-Smad2 in 11 out of 12 cases. Some osteoclasts also exhibited p ∼ Smad2 expression. The OSCC cell line, HSC3, and the bone marrow-derived fibroblastic cell line, ST2, synthesized substantial levels of TGF-β. Culture media derived from HSC3 cells could stimulate Tgf-β1 mRNA expression in ST2 cells. Recombinant TGF-β1 could stimulate osteoclast formation induced by receptor activator of nuclear factor kappa-B ligand (RANKL) in RAW264 cells. TGF-β1 could upregulate the expression of p-Smad2 in RAW264 cells, and this action was suppressed by the addition of a neutralizing antibody against TGF-β or by SB431542. Transplantation of HSC3 cells onto the calvarial region of athymic mice caused bone destruction, associated with the expression of TGF-β and p-Smad2 in both cancer cells and stromal cells. The bone destruction was substantially inhibited by the administration of SB431542. The present study demonstrated that TGF-β synthesized by both cancer cells and stromal cells participates in the OSCC-induced bone destruction. - Highlights: • Cancer cell, fibroblastic cells, and osteoclasts at bone resorbing area by oral cancer exhibited TGF-β and p-Smad2. • TGF-β1 stimulated osteoclastogenesis induced by RAKL in RAW264 cell. • Xenograft model of oral cancer-induced bone resorption was substantially inhibited by SB431542. • TGF-β synthesized by both cancer cells and stromal cells participates in the OSCC

  13. Implication of Tumor Microenvironment in Chemoresistance: Tumor-Associated Stromal Cells Protect Tumor Cells from Cell Death

    PubMed Central

    Castells, Magali; Thibault, Benoît; Delord, Jean-Pierre; Couderc, Bettina

    2012-01-01

    Tumor development principally occurs following the accumulation of genetic and epigenetic alterations in tumor cells. These changes pave the way for the transformation of chemosensitive cells to chemoresistant ones by influencing the uptake, metabolism, or export of drugs at the cellular level. Numerous reports have revealed the complexity of tumors and their microenvironment with tumor cells located within a heterogeneous population of stromal cells. These stromal cells (fibroblasts, endothelial or mesothelial cells, adipocytes or adipose tissue-derived stromal cells, immune cells and bone marrow-derived stem cells) could be involved in the chemoresistance that is acquired by tumor cells via several mechanisms: (i) cell–cell and cell–matrix interactions influencing the cancer cell sensitivity to apoptosis; (ii) local release of soluble factors promoting survival and tumor growth (crosstalk between stromal and tumor cells); (iii) direct cell-cell interactions with tumor cells (crosstalk or oncologic trogocytosis); (iv) generation of specific niches within the tumor microenvironment that facilitate the acquisition of drug resistance; or (v) conversion of the cancer cells to cancer-initiating cells or cancer stem cells. This review will focus on the implication of each member of the heterogeneous population of stromal cells in conferring resistance to cytotoxins and physiological mediators of cell death. PMID:22949815

  14. Conference Planning.

    ERIC Educational Resources Information Center

    Burke, W. Warner, Ed.; Beckhard, Richard, Ed.

    This book, written to instruct in the use of a conference as a medium of social intercourse, is divided into four sections. Section I, which contains five articles, deals with factors to be considered in planning a conference. Specific techniques one can employ to improve a conference and several different techniques for evaluating the…

  15. Transforming growth factor-β synthesized by stromal cells and cancer cells participates in bone resorption induced by oral squamous cell carcinoma.

    PubMed

    Nakamura, Ryosuke; Kayamori, Kou; Oue, Erika; Sakamoto, Kei; Harada, Kiyoshi; Yamaguchi, Akira

    2015-03-20

    Transforming growth factor beta (TGF-β) plays a significant role in the regulation of the tumor microenvironment. To explore the role of TGF-β in oral cancer-induced bone destruction, we investigated the immunohistochemical localization of TGF-β and phosphorylated Smad2 (p-Smad2) in 12 surgical specimens of oral squamous cell carcinoma (OSCC). These studies revealed TGF-β and p-Smad2 expression in cancer cells in all tested cases. Several fibroblasts located between cancer nests and resorbing bone expressed TGF-β in 10 out of 12 cases and p-Smad2 in 11 out of 12 cases. Some osteoclasts also exhibited p ∼ Smad2 expression. The OSCC cell line, HSC3, and the bone marrow-derived fibroblastic cell line, ST2, synthesized substantial levels of TGF-β. Culture media derived from HSC3 cells could stimulate Tgf-β1 mRNA expression in ST2 cells. Recombinant TGF-β1 could stimulate osteoclast formation induced by receptor activator of nuclear factor kappa-B ligand (RANKL) in RAW264 cells. TGF-β1 could upregulate the expression of p-Smad2 in RAW264 cells, and this action was suppressed by the addition of a neutralizing antibody against TGF-β or by SB431542. Transplantation of HSC3 cells onto the calvarial region of athymic mice caused bone destruction, associated with the expression of TGF-β and p-Smad2 in both cancer cells and stromal cells. The bone destruction was substantially inhibited by the administration of SB431542. The present study demonstrated that TGF-β synthesized by both cancer cells and stromal cells participates in the OSCC-induced bone destruction.

  16. Platelet-Derived Growth Factor BB Enhances Osteogenesis of Adipose-Derived But Not Bone Marrow-Derived Mesenchymal Stromal/Stem Cells.

    PubMed

    Hung, Ben P; Hutton, Daphne L; Kozielski, Kristen L; Bishop, Corey J; Naved, Bilal; Green, Jordan J; Caplan, Arnold I; Gimble, Jeffrey M; Dorafshar, Amir H; Grayson, Warren L

    2015-09-01

    Tissue engineering using mesenchymal stem cells (MSCs) holds great promise for regenerating critically sized bone defects. While the bone marrow-derived MSC is the most widely studied stromal/stem cell type for this application, its rarity within bone marrow and painful isolation procedure have motivated investigation of alternative cell sources. Adipose-derived stromal/stem cells (ASCs) are more abundant and more easily procured; furthermore, they also possess robust osteogenic potency. While these two cell types are widely considered very similar, there is a growing appreciation of possible innate differences in their biology and response to growth factors. In particular, reports indicate that their osteogenic response to platelet-derived growth factor BB (PDGF-BB) is markedly different: MSCs responded negatively or not at all to PDGF-BB while ASCs exhibited enhanced mineralization in response to physiological concentrations of PDGF-BB. In this study, we directly tested whether a fundamental difference existed between the osteogenic responses of MSCs and ASCs to PDGF-BB. MSCs and ASCs cultured under identical osteogenic conditions responded disparately to 20 ng/ml of PDGF-BB: MSCs exhibited no difference in mineralization while ASCs produced more calcium per cell. siRNA-mediated knockdown of PDGFRβ within ASCs abolished their ability to respond to PDGF-BB. Gene expression was also different; MSCs generally downregulated and ASCs generally upregulated osteogenic genes in response to PDGF-BB. ASCs transduced to produce PDGF-BB resulted in more regenerated bone within a critically sized murine calvarial defect compared to control ASCs, indicating PDGF-BB used specifically in conjunction with ASCs might enhance tissue engineering approaches for bone regeneration.

  17. Sustained Release of Engineered Stromal Cell–Derived Factor 1-α From Injectable Hydrogels Effectively Recruits Endothelial Progenitor Cells and Preserves Ventricular Function After Myocardial Infarction

    PubMed Central

    MacArthur, John W.; Purcell, Brendan P.; Shudo, Yasuhiro; Cohen, Jeffrey E.; Fairman, Alex; Trubelja, Alen; Patel, Jay; Hsiao, Philip; Yang, Elaine; Lloyd, Kelsey; Hiesinger, William; Atluri, Pavan; Burdick, Jason A.; Woo, Y. Joseph

    2014-01-01

    Background Exogenously delivered chemokines have enabled neovasculogenic myocardial repair in models of ischemic cardiomyopathy; however, these molecules have short half-lives in vivo. In this study, we hypothesized that the sustained delivery of a synthetic analog of stromal cell–derived factor 1-α (engineered stromal cell–derived factor analog [ESA]) induces continuous homing of endothelial progenitor cells and improves left ventricular function in a rat model of myocardial infarction. Methods and Results Our previously designed ESA peptide was synthesized by the addition of a fluorophore tag for tracking. Hyaluronic acid was chemically modified with hydroxyethyl methacrylate to form hydrolytically degradable hydrogels through free-radical–initiated crosslinking. ESA was encapsulated in hyaluronic acid hydrogels during gel formation, and then ESA release, along with gel degradation, was monitored for more than 4 weeks in vitro. Chemotactic properties of the eluted ESA were assessed at multiple time points using rat endothelial progenitor cells in a transwell migration assay. Finally, adult male Wistar rats (n=33) underwent permanent ligation of the left anterior descending (LAD) coronary artery, and 100 μL of saline, hydrogel alone, or hydrogel+25 μg ESA was injected into the borderzone. ESA fluorescence was monitored in animals for more than 4 weeks, after which vasculogenic, geometric, and functional parameters were assessed to determine the therapeutic benefit of each treatment group. ESA release was sustained for 4 weeks in vitro, remained active, and enhanced endothelial progenitor cell chemotaxis. In addition, ESA was detected in the rat heart >3 weeks when delivered within the hydrogels and significantly improved vascularity, ventricular geometry, ejection fraction, cardiac output, and contractility compared with controls. Conclusions We have developed a hydrogel delivery system that sustains the release of a bioactive endothelial progenitor cell

  18. Pathological complete response in advanced gastric stromal tumor after imatinib mesylate therapy: a case report

    PubMed Central

    2011-01-01

    Introduction Gastrointestinal stromal tumors are a rare neoplasm exhibiting, in most cases, mutations of c-kit. Imatinib mesylate is the standard treatment for patients who have advanced gastrointestinal stromal tumors. Although the response rate in patients treated with imatinib mesylate in prospective clinical studies is above 50%, a complete response is very rare. We report the case of a patient with a gastric gastrointestinal stromal tumor who had a pathological complete response after neoadjuvant treatment with imatinib mesylate. Case presentation We report the case of a 54-year-old Arab woman with a gastrointestinal stromal tumor who had a pathological complete response after neoadjuvant treatment with imatinib mesylate. Conclusion The pathological examination of our patient documented a complete pathological response after imatinib therapy. Recently, it has been confirmed that the kinase genotype of KIT and platelet-derived growth factor receptor α can accurately predict a good response to imatinib mesylate therapy. We propose that this patient had a mutation conferring high sensitivity to imatinib mesylate. PMID:21600007

  19. The role of environmental factors in medically unexplained symptoms and related syndromes: conference summary and recommendations.

    PubMed Central

    Kipen, Howard M; Fiedler, Nancy

    2002-01-01

    This monograph of peer-reviewed articles is based on presentations at the conference "Environmental Factors in Medically Unexplained Physical Symptoms and Related Syndromes" held 10-12 January 2001 in Piscataway, New Jersey, USA. The purpose of the conference was to determine research priorities for elucidating the role of environmental factors in medically unexplained symptoms and symptom syndromes. These include conditions such as chronic fatigue syndrome, multiple chemical sensitivities, sick building syndrome, Gulf War illness, and the like. Approximately 1 1/2 days were devoted to plenary talks and 1 day was devoted to break-out sessions to discuss epidemiologic, psychosocial, and experimental research. Recommendations were made for a series of epidemiologic, psychosocial, and experimental research approaches, with acknowledgment that nosology issues are clearly fundamental to advancing understanding of these conditions. PMID:12194891

  20. Stromal Cell-Derived Factor-1β Potentiates Bone Morphogenetic Protein-2-Stimulated Osteoinduction of Genetically Engineered Bone Marrow-Derived Mesenchymal Stem Cells In Vitro

    PubMed Central

    Herberg, Samuel; Fulzele, Sadanand; Yang, Nianlan; Shi, Xingming; Hess, Matthew; Periyasamy-Thandavan, Sudharsan; Hamrick, Mark W.; Isales, Carlos M.

    2013-01-01

    Skeletal injuries are among the most prevalent clinical problems and bone marrow-derived mesenchymal stem/stromal cells (BMSCs) have successfully been used for the treatment thereof. Stromal cell-derived factor-1 (SDF-1; CXCL12) is a member of the CXC chemokine family with multiple splice variants. The two most abundant variants, SDF-1α and SDF-1β, share identical amino acid sequences, except for four additional amino acids at the C-terminus of SDF-1β, which may mediate surface stabilization via glycosaminoglycans and protect SDF-1β from proteolytic cleavage, rendering it twice as potent as SDF-1α. Increasing evidence suggests that SDF-1 is involved in bone formation through regulation of recruitment, engraftment, proliferation, and differentiation of stem/progenitor cells. The underlying molecular mechanisms, however, have not yet been fully elucidated. In this study, we tested the hypothesis that SDF-1β can potentiate bone morphogenetic protein-2 (BMP-2)-stimulated osteogenic differentiation and chemotaxis of BMSCs in vitro. Utilizing retrovirus-mediated gene transfer to generate novel Tet-Off-SDF-1β BMSCs, we found that conditional SDF-1β expression is tightly regulated by doxycycline in a dose-dependent and temporal fashion, leading to significantly increased SDF-1β mRNA and protein levels. In addition, SDF-1β was found to enhance BMP-2-stimulated mineralization, mRNA and protein expression of key osteogenic markers, and regulate BMP-2 signal transduction via extracellular signal-regulated kinases 1/2 (Erk1/2) phosphorylation in genetically engineered BMSCs in vitro. We also showed that SDF-1β promotes the migratory response of CXC chemokine receptor 4 (CXCR4)-expressing BMSCs in vitro. Taken together, these data support that SDF-1β can play an important role in BMP-2-stimulated osteogenic differentiation of BMSCs and may exert its biological activity in both an autocrine and paracrine fashion. PMID:22779446

  1. Gastrointestinal stromal tumor

    PubMed Central

    Yue, Changjun

    2012-01-01

    Gastrointestinal stromal tumor has received a lot of attention over the last 10 years due to its unique biologic behavior, clinicopathological features, molecular mechanisms, and treatment implications. GIST is the most common mesenchymal neoplasm in the gastrointestinal tract and has emerged from a poorly understood and treatment resistant neoplasm to a well-defined tumor entity since the discovery of particular molecular abnormalities, KIT and PDGFRA gene mutations. The understanding of GIST biology at the molecular level promised the development of novel treatment modalities. Diagnosis of GIST depends on the integrity of histology, immunohistochemistry and molecular analysis. The risk assessment of the tumor behavior relies heavily on pathological evaluation and significantly impacts clinical management. In this review, historic review, epidemiology, pathogenesis and genetics, diagnosis, role of molecular analysis, prognostic factor and treatment strategies have been discussed. PMID:22943011

  2. Dexamethasone Regulates EphA5, a Potential Inhibitory Factor with Osteogenic Capability of Human Bone Marrow Stromal Cells

    PubMed Central

    Yamada, Tsuyoshi; Yoshii, Toshitaka; Yasuda, Hiroaki; Okawa, Atsushi; Sotome, Shinichi

    2016-01-01

    We previously demonstrated the importance of quality management procedures for the handling of human bone marrow stromal cells (hBMSCs) and provided evidence for the existence of osteogenic inhibitor molecules in BMSCs. One candidate inhibitor is the ephrin type-A receptor 5 (EphA5), which is expressed in hBMSCs and upregulated during long-term culture. In this study, forced expression of EphA5 diminished the expression of osteoblast phenotypic markers. Downregulation of endogenous EphA5 by dexamethasone treatment promoted osteoblast marker expression. EphA5 could be involved in the normal growth regulation of BMSCs and could be a potential marker for replicative senescence. Although Eph forward signaling stimulated by ephrin-B-Fc promoted the expression of ALP mRNA in BMSCs, exogenous addition of EphA5-Fc did not affect the ALP level. The mechanism underlying the silencing of EphA5 in early cultures remains unclear. EphA5 promoter was barely methylated in hBMSCs while histone deacetylation could partially suppress EphA5 expression in early-passage cultures. In repeatedly passaged cultures, the upregulation of EphA5 independent of methylation could competitively inhibit osteogenic signal transduction pathways such as EphB forward signaling. Elucidation of the potential inhibitory function of EphA5 in hBMSCs may provide an alternative approach for lineage differentiation in cell therapy strategies and regenerative medicine. PMID:27057165

  3. Influence of Factors of Cryopreservation and Hypothermic Storage on Survival and Functional Parameters of Multipotent Stromal Cells of Placental Origin

    PubMed Central

    Pogozhykh, Olena; Mueller, Thomas; Prokopyuk, Olga

    2015-01-01

    Human placenta is a highly perspective source of multipotent stromal cells (MSCs) both for the purposes of patient specific auto-banking and allogeneic application in regenerative medicine. Implementation of new GMP standards into clinical practice enforces the search for relevant methods of cryopreservation and short-term hypothermic storage of placental MSCs. In this paper we analyze the effect of different temperature regimes and individual components of cryoprotective media on viability, metabolic and culture properties of placental MSCs. We demonstrate (I) the possibility of short-term hypothermic storage of these cells; (II) determine DMSO and propanediol as the most appropriate cryoprotective agents; (III) show the possibility of application of volume expanders (plasma substituting solutions based on dextran or polyvinylpyrrolidone); (IV) reveal the priority of ionic composition over the serum content in cryopreservation media; (V) determine a cooling rate of 1°C/min down to -40°C followed by immersion into liquid nitrogen as the optimal cryopreservation regime for this type of cells. This study demonstrates perspectives for creation of new defined cryopreservation methods towards GMP standards. PMID:26431528

  4. Transforming Growth Factor β1 (TGFβ1) and Progesterone Regulate Matrix Metalloproteinases (MMP) in Human Endometrial Stromal Cells

    PubMed Central

    Itoh, Hiroko; Kishore, Annavarapu Hari; Lindqvist, Annika; Rogers, David E.

    2012-01-01

    Context: Menstruation is preceded by progesterone withdrawal and endometrial matrix remodeling predominantly through induction of matrix metalloproteinases (MMP) and recruitment of invading neutrophils. Design: Using endometrial tissues from women during various phases of the menstrual cycle, we found that MMP2, MMP9, and MMP11 were up-regulated in the late secretory phase/premenstrual phase. Because TGFβ-responsive genes were also up-regulated in endometrium during this time, we tested the hypothesis that TGFβ1 and progesterone regulate expression of MMP in human endometrial stromal cells (HESC). Results: Treatment of HESC with TGFβ1 resulted in marked increases in MMP2 and MMP11 mRNA and pro- and active MMP2 activity. Progesterone inhibited TGFβ1-induced stimulation of MMP2 and MMP11 through its nuclear hormone receptors. Interestingly, TGFβ1 also decreased progesterone receptor (PR)-A and PR-B in HESC with a more pronounced effect on PR-A. Conclusions: These data support the hypothesis that TGFβ1 has endogenous anti-progestational effects in HESC and that the opposing effects of progesterone and TGFβ1 are important in regulation of matrix integrity in human endometrium. PMID:22466340

  5. Influence of vascular endothelial growth factor stimulation and serum deprivation on gene activation patterns of human adipose tissue-derived stromal cells.

    PubMed

    Tratwal, Josefine; Mathiasen, Anders Bruun; Juhl, Morten; Brorsen, Sonja Kim; Kastrup, Jens; Ekblond, Annette

    2015-04-13

    Stimulation of mesenchymal stromal cells and adipose tissue-derived stromal cells (ASCs) with vascular endothelial growth factor (VEGF) has been used in multiple animal studies and clinical trials for regenerative purposes. VEGF stimulation is believed to promote angiogenesis and VEGF stimulation is usually performed under serum deprivation. Potential regenerative molecular mechanisms are numerous and the role of contributing factors is uncertain. The aim of the current study was to investigate the effect of in vitro serum deprivation and VEGF stimulation on gene expression patterns of ASCs. Gene expressions of ASCs cultured in complete medium, ASCs cultured in serum-deprived medium and ASCs stimulated with VEGF in serum-deprived medium were compared. ASC characteristics according to criteria set by the International Society of Cellular Therapy were confirmed by flow cytometry. Microarray gene expressions were obtained using the Affymetrix HT HG-U133+ GeneChip®. Gene set enrichment analysis was performed using the Kyoto Encyclopedia of Genes and Genomes and gene ontology terms. Transcription of selected genes of interest was confirmed by quantitative PCR. Compared to ASCs in complete medium, 190 and 108 genes were significantly altered by serum deprivation and serum deprivation combined with VEGF, respectively. No significant differences in gene expression patterns between serum-deprived ASCs and serum-deprived ASCs combined with VEGF stimulation were found. Genes most prominently and significantly upregulated by both conditions were growth factors (IGF1, BMP6, PDGFD, FGF9), adhesion molecule CLSTN2, extracellular matrix-related proteins such as matricellular proteins SMOC2, SPON1 and ADAMTS12, and inhibitors of proliferation (JAG1). The most significantly downregulated genes included matrix metalloproteinases (MMP3, MMP1), and proliferation markers (CDKN3) and GREM2 (a BMP6 antagonist). The decisive factor for the observed change in ASC gene expression proves to

  6. The effect of growth factor supplementation on corneal stromal cell phenotype in vitro using a serum-free media.

    PubMed

    Lynch, Amy P; O'Sullivan, Finbarr; Ahearne, Mark

    2016-10-01

    In order to expand cells quickly and in high numbers for corneal tissue engineering applications corneal stromal cells, or keratocytes, are often cultured in the presence of serum. However, keratocytes become fibroblastic when exposed to serum leading to a downregulation of corneal stromal specific markers. The purpose of this current study was to determine if corneal stromal cells, made fibroblastic by serum, could display native quiescent keratocyte characteristics when cultured under serum-free conditions supplemented by different growth factors. Markers specific to a native keratocyte phenotype such as keratocan and aldehyde dehydrogenase 3A1 (ALDH3A1) and those specific to a fibrotic phenotype such as α-smooth muscle actin (αSMA) and collagen type III were examined. Cells were cultured in monolayer, self-assembled pellets or collagen hydrogels. Growth factors known to modulate keratocyte phenotype were chosen to supplement the serum free media, specifically insulin-like growth factor 1 (IGF-1) and transforming growth factor beta 1 and 3 (Tβ1 and Tβ3). The effects of serum-free media, growth factors and culture system on cell proliferation and morphology and extracellular matrix (ECM) synthesis were evaluated. The expression of keratocyte markers was evaluated by real-time PCR, immunofluorescent staining and western blotting. In addition, cell migration was tested using scratch assays. When serum was removed from the cells they displayed a reduction in proliferation and ECM synthesis (not significant), in addition to a significant decrease in migratory capacity (p < 0.05). Serum-free media promoted increased expression of keratocan (130.68 ± 47.44-fold increase; p < 0.05) and collagen type I (15.58 ± 9.49-fold increase; p < 0.05). However, there was no significant change in ALDH3A1 and αSMA expression, while collagen type III expression was significantly increased (44.66 ± 25.61-fold increase; p < 0.05). In addition, cells retained an

  7. Stromal Effects on Mammary Gland Development and Breast Cancer

    NASA Astrophysics Data System (ADS)

    Wiseman, Bryony S.; Werb, Zena

    2002-05-01

    Breast cancer manifests itself in the mammary epithelium, yet there is a growing recognition that mammary stromal cells also play an important role in tumorigenesis. During its developmental cycle, the mammary gland displays many of the properties associated with breast cancer, and many of the stromal factors necessary for mammary development also promote or protect against breast cancer. Here we review our present knowledge of the specific factors and cell types that contribute to epithelial-stromal crosstalk during mammary development. To find cures for diseases like breast cancer that rely on epithelial-stromal crosstalk, we must understand how these different cell types communicate with each other.

  8. Effects on proliferation and differentiation of multipotent bone marrow stromal cells engineered to express growth factors for combined cell and gene therapy.

    PubMed

    Fierro, Fernando A; Kalomoiris, Stefanos; Sondergaard, Claus S; Nolta, Jan A

    2011-11-01

    A key mechanism for mesenchymal stem cells/bone marrow stromal cells (MSCs) to promote tissue repair is by secretion of soluble growth factors (GFs). Therefore, clinical application could be optimized by a combination of cell and gene therapies, where MSCs are genetically modified to express higher levels of a specific factor. However, it remains unknown how this overexpression may alter the fate of the MSCs. Here, we show effects of overexpressing the growth factors, such as basic fibroblast growth factor (bFGF), platelet derived growth factor B (PDGF-BB), transforming growth factor β(1) (TGF-β(1) ), and vascular endothelial growth factor (VEGF), in human bone marrow-derived MSCs. Ectopic expression of bFGF or PDGF-B lead to highly proliferating MSCs and lead to a robust increase in osteogenesis. In contrast, adipogenesis was strongly inhibited in MSCs overexpressing PDGF-B and only mildly affected in MSCs overexpressing bFGF. Overexpression of TGF-β(1) blocked both osteogenic and adipogenic differentiation while inducing the formation of stress fibers and increasing the expression of the smooth muscle marker calponin-1 and the chondrogenic marker collagen type II. In contrast, MSCs overexpressing VEGF did not vary from control MSCs in any parameters, likely due to the lack of VEGF receptor expression on MSCs. MSCs engineered to overexpress VEGF strongly induced the migration of endothelial cells and enhanced blood flow restoration in a xenograft model of hind limb ischemia. These data support the rationale for genetically modifying MSCs to enhance their therapeutically relevant trophic signals, when safety and efficacy can be demonstrated, and when it can be shown that there are no unwanted effects on their proliferation and differentiation. Copyright © 2011 AlphaMed Press.

  9. Effects on Proliferation and Differentiation of Multipotent Bone Marrow Stromal Cells Engineered to Express Growth Factors for Combined Cell and Gene Therapy

    PubMed Central

    Fierro, Fernando A.; Kalomoiris, Stefanos; Sondergaard, Claus S.; Nolta, Jan A.

    2013-01-01

    A key mechanism for mesenchymal stem cells/bone marrow stromal cells (MSCs) to promote tissue repair is by secretion of soluble growth factors (GFs). Therefore, clinical application could be optimized by a combination of cell and gene therapies, where MSCs are genetically modified to express higher levels of a specific factor. However, it remains unknown how this overexpression may alter the fate of the MSCs. Here, we show effects of overexpressing the growth factors, such as basic fibroblast growth factor (bFGF), platelet derived growth factor B (PDGF-BB), transforming growth factor β1 (TGF-β1), and vascular endothelial growth factor (VEGF), in human bone marrow-derived MSCs. Ectopic expression of bFGF or PDGF-B lead to highly proliferating MSCs and lead to a robust increase in osteogenesis. In contrast, adipogenesis was strongly inhibited in MSCs overexpressing PDGF-B and only mildly affected in MSCs overexpressing bFGF. Overexpression of TGF-β1 blocked both osteogenic and adipogenic differentiation while inducing the formation of stress fibers and increasing the expression of the smooth muscle marker calponin-1 and the chondrogenic marker collagen type II. In contrast, MSCs overexpressing VEGF did not vary from control MSCs in any parameters, likely due to the lack of VEGF receptor expression on MSCs. MSCs engineered to overexpress VEGF strongly induced the migration of endothelial cells and enhanced blood flow restoration in a xenograft model of hind limb ischemia. These data support the rationale for genetically modifying MSCs to enhance their therapeutically relevant trophic signals, when safety and efficacy can be demonstrated, and when it can be shown that there are no unwanted effects on their proliferation and differentiation. PMID:21898687

  10. Intra-articular injection of human meniscus stem/progenitor cells promotes meniscus regeneration and ameliorates osteoarthritis through stromal cell-derived factor-1/CXCR4-mediated homing.

    PubMed

    Shen, Weiliang; Chen, Jialin; Zhu, Ting; Chen, Longkun; Zhang, Wei; Fang, Zhi; Heng, Boon Chin; Yin, Zi; Chen, Xiao; Ji, Junfeng; Chen, Weishan; Ouyang, Hong-Wei

    2014-03-01

    Meniscus injury is frequently encountered in clinical practice. Current surgical therapy involving partial or complete meniscectomy relieves pain in the short-term but often leads to osteoarthritis (OA) in the long-term. In this study, we report a new strategy of articular cartilage protection by intra-articular injection of novel human meniscus stem/progenitor cells (hMeSPCs). We found that hMeSPCs displayed both mesenchymal stem cell characteristics and high expression levels of collagen II. In the rat meniscus injury model, hMeSPC transplantation not only led to more neo-tissue formation and better-defined shape but also resulted in more rounded cells and matured extracellular matrix. Stromal cell-derived factor-1 (SDF-1) enhanced the migration of hMeSPCs, whereas AMD3100 abolished the chemotactic effects of SDF-1 on hMeSPCs, both in vitro and in vivo. In an experimental OA model, transplantation of hMeSPCs effectively protected articular cartilage, as evidenced by reduced expression of OA markers such as collagen I, collagen X, and hypoxia-inducible factor 2α but increased expression of collagen II. Our study demonstrated for the first time that intra-articular injection of hMeSPCs enhanced meniscus regeneration through the SDF-1/CXCR4 axis. Our study highlights a new strategy of intra-articular injection of hMeSPCs for meniscus regeneration.

  11. Stretching human mesenchymal stromal cells on stiffness-customized collagen type I generates a smooth muscle marker profile without growth factor addition

    NASA Astrophysics Data System (ADS)

    Rothdiener, Miriam; Hegemann, Miriam; Uynuk-Ool, Tatiana; Walters, Brandan; Papugy, Piruntha; Nguyen, Phong; Claus, Valentin; Seeger, Tanja; Stoeckle, Ulrich; Boehme, Karen A.; Aicher, Wilhelm K.; Stegemann, Jan P.; Hart, Melanie L.; Kurz, Bodo; Klein, Gerd; Rolauffs, Bernd

    2016-10-01

    Using matrix elasticity and cyclic stretch have been investigated for inducing mesenchymal stromal cell (MSC) differentiation towards the smooth muscle cell (SMC) lineage but not in combination. We hypothesized that combining lineage-specific stiffness with cyclic stretch would result in a significantly increased expression of SMC markers, compared to non-stretched controls. First, we generated dense collagen type I sheets by mechanically compressing collagen hydrogels. Atomic force microscopy revealed a nanoscale stiffness range known to support myogenic differentiation. Further characterization revealed viscoelasticity and stable biomechanical properties under cyclic stretch with >99% viable adherent human MSC. MSCs on collagen sheets demonstrated a significantly increased mRNA but not protein expression of SMC markers, compared to on culture flasks. However, cyclic stretch of MSCs on collagen sheets significantly increased both mRNA and protein expression of α-smooth muscle actin, transgelin, and calponin versus plastic and non-stretched sheets. Thus, lineage-specific stiffness and cyclic stretch can be applied together for inducing MSC differentiation towards SMCs without the addition of recombinant growth factors or other soluble factors. This represents a novel stimulation method for modulating the phenotype of MSCs towards SMCs that could easily be incorporated into currently available methodologies to obtain a more targeted control of MSC phenotype.

  12. Human Umbilical Cord Perivascular Cells Exhibited Enhanced Migration Capacity towards Hepatocellular Carcinoma in Comparison with Bone Marrow Mesenchymal Stromal Cells: A Role for Autocrine Motility Factor Receptor

    PubMed Central

    Aquino, Jorge B.; Malvicini, Mariana; Bolontrade, Marcela; Podhajcer, Osvaldo; Garcia, Mariana G.; Mazzolini, Guillermo

    2014-01-01

    Hepatocellular carcinoma (HCC) is the third cause of cancer-related death worldwide. Unfortunately, the incidence and mortality associated with HCC are increasing. Therefore, new therapeutic strategies are urgently needed and the use of mesenchymal stromal cells (MSCs) as carrier of therapeutic genes is emerging as a promising option. Different sources of MSCs are being studied for cell therapy and bone marrow-derived cells are the most extensively explored; however, birth associated-tissues represent a very promising source. The aim of this work was to compare the in vitro and in vivo migration capacity between bone marrow MSCs (BM-MSCs) and human umbilical cord perivascular cells (HUCPVCs) towards HCC. We observed that HUCPVCs presented higher in vitro and in vivo migration towards factors released by HCC. The expression of autocrine motility factor (AMF) receptor, genes related with the availability of the receptor on the cell surface (caveolin-1 and -2) and metalloproteinase 3, induced by the receptor activation and important for cell migration, was increased in HUCPVCs. The chemotactic response towards recombinant AMF was increased in HUCPVCs compared to BM-MSCs, and its inhibition in the conditioned medium from HCC induced higher decrease in HUCPVC migration than in BM-MSC. Our results indicate that HUCPVCs could be a useful cellular source to deliver therapeutic genes to HCC. PMID:25147818

  13. Human umbilical cord perivascular cells exhibited enhanced migration capacity towards hepatocellular carcinoma in comparison with bone marrow mesenchymal stromal cells: a role for autocrine motility factor receptor.

    PubMed

    Bayo, Juan; Fiore, Esteban; Aquino, Jorge B; Malvicini, Mariana; Rizzo, Manglio; Peixoto, Estanislao; Alaniz, Laura; Piccioni, Flavia; Bolontrade, Marcela; Podhajcer, Osvaldo; Garcia, Mariana G; Mazzolini, Guillermo

    2014-01-01

    Hepatocellular carcinoma (HCC) is the third cause of cancer-related death worldwide. Unfortunately, the incidence and mortality associated with HCC are increasing. Therefore, new therapeutic strategies are urgently needed and the use of mesenchymal stromal cells (MSCs) as carrier of therapeutic genes is emerging as a promising option. Different sources of MSCs are being studied for cell therapy and bone marrow-derived cells are the most extensively explored; however, birth associated-tissues represent a very promising source. The aim of this work was to compare the in vitro and in vivo migration capacity between bone marrow MSCs (BM-MSCs) and human umbilical cord perivascular cells (HUCPVCs) towards HCC. We observed that HUCPVCs presented higher in vitro and in vivo migration towards factors released by HCC. The expression of autocrine motility factor (AMF) receptor, genes related with the availability of the receptor on the cell surface (caveolin-1 and -2) and metalloproteinase 3, induced by the receptor activation and important for cell migration, was increased in HUCPVCs. The chemotactic response towards recombinant AMF was increased in HUCPVCs compared to BM-MSCs, and its inhibition in the conditioned medium from HCC induced higher decrease in HUCPVC migration than in BM-MSC. Our results indicate that HUCPVCs could be a useful cellular source to deliver therapeutic genes to HCC.

  14. Cyclooxygenase-2 or tumor necrosis factor-α inhibitors attenuate the mechanotransductive effects of pulsed focused ultrasound to suppress mesenchymal stromal cell homing to healthy and dystrophic muscle.

    PubMed

    Tebebi, Pamela A; Burks, Scott R; Kim, Saejeong J; Williams, Rashida A; Nguyen, Ben A; Venkatesh, Priyanka; Frenkel, Victor; Frank, Joseph A

    2015-04-01

    Maximal homing of infused stem cells to diseased tissue is critical for regenerative medicine. Pulsed focused ultrasound (pFUS) is a clinically relevant platform to direct stem cell migration. Through mechanotransduction, pFUS establishes local gradients of cytokines, chemokines, trophic factors (CCTF) and cell adhesion molecules (CAM) in treated skeletal muscle that subsequently infused mesenchymal stromal cells (MSC) can capitalize to migrate into the parenchyma. Characterizing molecular responses to mechanical pFUS effects revealed tumor necrosis factor-alpha (TNFα) drives cyclooxygenase-2 (COX2) signaling to locally increase CCTF/CAM that are necessary for MSC homing. pFUS failed to increase chemoattractants and induce MSC homing to treated muscle in mice pretreated with ibuprofen (nonspecific COX inhibitor) or etanercept (TNFα inhibitor). pFUS-induced MSC homing was also suppressed in COX2-knockout mice, demonstrating ibuprofen blocked the mechanically induced CCTF/CAM by acting on COX2. Anti-inflammatory drugs, including ibuprofen, are administered to muscular dystrophy (MD) patients, and ibuprofen also suppressed pFUS-induced homing to muscle in a mouse model of MD. Drug interactions with cell therapies remain unexplored and are not controlled for during clinical cell therapy trials. This study highlights potentially negative drug-host interactions that suppress stem cell homing and could undermine cell-based approaches for regenerative medicine.

  15. Identification of two distinct subsets of long-term nonprogressors with divergent viral activity by stromal-derived factor 1 chemokine gene polymorphism analysis.

    PubMed

    Balotta, C; Bagnarelli, P; Corvasce, S; Mazzucchelli, R; Colombo, M C; Papagno, L; Santambrogio, S; Ridolfo, A L; Violin, M; Berlusconi, A; Velleca, R; Facchi, G; Moroni, M; Clementi, M; Galli, M

    1999-08-01

    Stromal-derived factor (SDF)-1, the natural ligand for CXCR4, is present in a common polymorphic variant defined by a G-->A transition in the 3' untranslated region of the gene. In persons infected with human immunodeficiency virus type 1 (HIV-1), the homozygous genotype (SDF1-3'A/3'A) has been postulated to interfere with the appearance of T-tropic syncytium-inducing strains. The polymorphism of SDF1 was correlated with HIV-1 phenotype, plasma viremia, and unspliced and multiply spliced specific transcripts in 158 virologically characterized HIV-1-infected patients (39 recent seroconverters, 75 typical progressors, and 44 AIDS patients) and in 42 HIV-1-infected long-term nonprogressors (LTNPs). Analysis of SDF1 allele distribution revealed that SDF1-3'A/3'A status is associated with low CD4 cell count (P=.0449) but not with a specific HIV-1 phenotype. In LTNPs, SDF1-+/+ condition defined a subset of persons with lower HIV-1 replication than in heterozygous subjects. The low viral activity in SDF1-+/+ LTNPs suggests that other factors play a major role in vivo in determining the course of HIV-1 infection.

  16. Constitutive stabilization of hypoxia-inducible factor alpha selectively promotes the self-renewal of mesenchymal progenitors and maintains mesenchymal stromal cells in an undifferentiated state

    PubMed Central

    Park, In-Ho; Kim, Kwang-Ho; Choi, Hyun-Kyung; Shim, Jae-Seung; Whang, Soo-Young; June Hahn, Sang; Kwon, Oh-Joo; Oh, Il-Hoan

    2013-01-01

    With the increasing use of culture-expanded mesenchymal stromal cells (MSCs) for cell therapies, factors that regulate the cellular characteristics of MSCs have been of major interest. Oxygen concentration has been shown to influence the functions of MSCs, as well as other normal and malignant stem cells. However, the underlying mechanisms of hypoxic responses and the precise role of hypoxia-inducible factor-1α (Hif-1α), the master regulatory protein of hypoxia, in MSCs remain unclear, due to the limited span of Hif-1α stabilization and the complex network of hypoxic responses. In this study, to further define the significance of Hif-1α in MSC function during their self-renewal and terminal differentiation, we established adult bone marrow (BM)-derived MSCs that are able to sustain high level expression of ubiquitin-resistant Hif-1α during such long-term biological processes. Using this model, we show that the stabilization of Hif-1α proteins exerts a selective influence on colony-forming mesenchymal progenitors promoting their self-renewal and proliferation, without affecting the proliferation of the MSC mass population. Moreover, Hif-1α stabilization in MSCs led to the induction of pluripotent genes (oct-4 and klf-4) and the inhibition of their terminal differentiation into osteogenic and adipogenic lineages. These results provide insights into the previously unrecognized roles of Hif-1α proteins in maintaining the primitive state of primary MSCs and on the cellular heterogeneities in hypoxic responses among MSC populations. PMID:24071737

  17. Lim Mineralization Protein 3 Induces the Osteogenic Differentiation of Human Amniotic Fluid Stromal Cells through Kruppel-Like Factor-4 Downregulation and Further Bone-Specific Gene Expression

    PubMed Central

    Barba, Marta; Pirozzi, Filomena; Saulnier, Nathalie; Vitali, Tiziana; Natale, Maria Teresa; Logroscino, Giandomenico; Robbins, Paul D.; Gambotto, Andrea; Neri, Giovanni; Michetti, Fabrizio; Pola, Enrico; Lattanzi, Wanda

    2012-01-01

    Multipotent mesenchymal stem cells with extensive self-renewal properties can be easily isolated and rapidly expanded in culture from small volumes of amniotic fluid. These cells, namely, amniotic fluid-stromal cells (AFSCs), can be regarded as an attractive source for tissue engineering purposes, being phenotypically and genetically stable, plus overcoming all the safety and ethical issues related to the use of embryonic/fetal cells. LMP3 is a novel osteoinductive molecule acting upstream to the main osteogenic pathways. This study is aimed at delineating the basic molecular events underlying LMP3-induced osteogenesis, using AFSCs as a cellular model to focus on the molecular features underlying the multipotency/differentiation switch. For this purpose, AFSCs were isolated and characterized in vitro and transfected with a defective adenoviral vector expressing the human LMP3. LMP3 induced the successful osteogenic differentiation of AFSC by inducing the expression of osteogenic markers and osteospecific transcription factors. Moreover, LMP3 induced an early repression of the kruppel-like factor-4, implicated in MSC stemness maintenance. KLF4 repression was released upon LMP3 silencing, indicating that this event could be reasonably considered among the basic molecular events that govern the proliferation/differentiation switch during LMP3-induced osteogenic differentiation of AFSC. PMID:23097599

  18. Intra-Articular Injection of Human Meniscus Stem/Progenitor Cells Promotes Meniscus Regeneration and Ameliorates Osteoarthritis Through Stromal Cell-Derived Factor-1/CXCR4-Mediated Homing

    PubMed Central

    Shen, Weiliang; Chen, Jialin; Zhu, Ting; Chen, Longkun; Zhang, Wei; Fang, Zhi; Heng, Boon Chin; Yin, Zi; Chen, Xiao; Ji, Junfeng

    2014-01-01

    Meniscus injury is frequently encountered in clinical practice. Current surgical therapy involving partial or complete meniscectomy relieves pain in the short-term but often leads to osteoarthritis (OA) in the long-term. In this study, we report a new strategy of articular cartilage protection by intra-articular injection of novel human meniscus stem/progenitor cells (hMeSPCs). We found that hMeSPCs displayed both mesenchymal stem cell characteristics and high expression levels of collagen II. In the rat meniscus injury model, hMeSPC transplantation not only led to more neo-tissue formation and better-defined shape but also resulted in more rounded cells and matured extracellular matrix. Stromal cell-derived factor-1 (SDF-1) enhanced the migration of hMeSPCs, whereas AMD3100 abolished the chemotactic effects of SDF-1 on hMeSPCs, both in vitro and in vivo. In an experimental OA model, transplantation of hMeSPCs effectively protected articular cartilage, as evidenced by reduced expression of OA markers such as collagen I, collagen X, and hypoxia-inducible factor 2α but increased expression of collagen II. Our study demonstrated for the first time that intra-articular injection of hMeSPCs enhanced meniscus regeneration through the SDF-1/CXCR4 axis. Our study highlights a new strategy of intra-articular injection of hMeSPCs for meniscus regeneration. PMID:24448516

  19. Cyclooxygenase-2 or tumor necrosis factor-α inhibitors attenuate the mechanotransductive effects of pulsed focused ultrasound to suppress mesenchymal stromal cell homing to healthy and dystrophic muscle

    PubMed Central

    Tebebi, Pamela A.; Burks, Scott R.; Kim, Saejeong J.; Williams, Rashida A.; Nguyen, Ben A.; Venkatesh, Priyanka; Frenkel, Victor; Frank, Joseph A.

    2014-01-01

    Maximal homing of infused stem cells to diseased tissue is critical for regenerative medicine. Pulsed focused ultrasound (pFUS) is a clinically relevant platform to direct stem cell migration. Through mechanotransduction, pFUS establishes local gradients of cytokines, chemokines, trophic factors (CCTF) and cell adhesion molecules (CAM) in treated skeletal muscle that subsequently infused mesenchymal stromal cells (MSC) can capitalize to migrate into the parenchyma. Characterizing molecular responses to mechanical pFUS effects revealed tumor necrosis factor-alpha (TNFα) drives cyclooxygenase-2 (COX2) signaling to locally increase CCTF/CAM that are necessary for MSC homing. pFUS failed to increase chemoattractants and induce MSC homing to treated muscle in mice pretreated with ibuprofen (non-specific COX inhibitor) or etanercept (TNFα inhibitor). pFUS-induced MSC homing was also suppressed in COX2-knockout mice, demonstrating ibuprofen blocked the mechanically-induced CCTF/CAM by acting on COX2. Anti-inflammatory drugs, including ibuprofen, are administered to muscular dystrophy (MD) patients and ibuprofen also suppressed pFUS-induced homing to muscle in a mouse model of MD. Drug interactions with cell therapies remain unexplored and are not controlled for during clinical cell therapy trials. This study highlights potentially negative drug-host interactions that suppress stem cell homing and could undermine cell-based approaches for regenerative medicine. PMID:25534849

  20. Constitutive stabilization of hypoxia-inducible factor alpha selectively promotes the self-renewal of mesenchymal progenitors and maintains mesenchymal stromal cells in an undifferentiated state.

    PubMed

    Park, In-Ho; Kim, Kwang-Ho; Choi, Hyun-Kyung; Shim, Jae-Seung; Whang, Soo-Young; Hahn, Sang June; Kwon, Oh-Joo; Oh, Il-Hoan

    2013-09-27

    With the increasing use of culture-expanded mesenchymal stromal cells (MSCs) for cell therapies, factors that regulate the cellular characteristics of MSCs have been of major interest. Oxygen concentration has been shown to influence the functions of MSCs, as well as other normal and malignant stem cells. However, the underlying mechanisms of hypoxic responses and the precise role of hypoxia-inducible factor-1α (Hif-1α), the master regulatory protein of hypoxia, in MSCs remain unclear, due to the limited span of Hif-1α stabilization and the complex network of hypoxic responses. In this study, to further define the significance of Hif-1α in MSC function during their self-renewal and terminal differentiation, we established adult bone marrow (BM)-derived MSCs that are able to sustain high level expression of ubiquitin-resistant Hif-1α during such long-term biological processes. Using this model, we show that the stabilization of Hif-1α proteins exerts a selective influence on colony-forming mesenchymal progenitors promoting their self-renewal and proliferation, without affecting the proliferation of the MSC mass population. Moreover, Hif-1α stabilization in MSCs led to the induction of pluripotent genes (oct-4 and klf-4) and the inhibition of their terminal differentiation into osteogenic and adipogenic lineages. These results provide insights into the previously unrecognized roles of Hif-1α proteins in maintaining the primitive state of primary MSCs and on the cellular heterogeneities in hypoxic responses among MSC populations.

  1. Stretching human mesenchymal stromal cells on stiffness-customized collagen type I generates a smooth muscle marker profile without growth factor addition

    PubMed Central

    Rothdiener, Miriam; Hegemann, Miriam; Uynuk-Ool, Tatiana; Walters, Brandan; Papugy, Piruntha; Nguyen, Phong; Claus, Valentin; Seeger, Tanja; Stoeckle, Ulrich; Boehme, Karen A.; Aicher, Wilhelm K.; Stegemann, Jan P.; Hart, Melanie L.; Kurz, Bodo; Klein, Gerd; Rolauffs, Bernd

    2016-01-01

    Using matrix elasticity and cyclic stretch have been investigated for inducing mesenchymal stromal cell (MSC) differentiation towards the smooth muscle cell (SMC) lineage but not in combination. We hypothesized that combining lineage-specific stiffness with cyclic stretch would result in a significantly increased expression of SMC markers, compared to non-stretched controls. First, we generated dense collagen type I sheets by mechanically compressing collagen hydrogels. Atomic force microscopy revealed a nanoscale stiffness range known to support myogenic differentiation. Further characterization revealed viscoelasticity and stable biomechanical properties under cyclic stretch with >99% viable adherent human MSC. MSCs on collagen sheets demonstrated a significantly increased mRNA but not protein expression of SMC markers, compared to on culture flasks. However, cyclic stretch of MSCs on collagen sheets significantly increased both mRNA and protein expression of α-smooth muscle actin, transgelin, and calponin versus plastic and non-stretched sheets. Thus, lineage-specific stiffness and cyclic stretch can be applied together for inducing MSC differentiation towards SMCs without the addition of recombinant growth factors or other soluble factors. This represents a novel stimulation method for modulating the phenotype of MSCs towards SMCs that could easily be incorporated into currently available methodologies to obtain a more targeted control of MSC phenotype. PMID:27775041

  2. Effects of transforming growth factor beta1 released from biodegradable polymer microparticles on marrow stromal osteoblasts cultured on poly(propylene fumarate) substrates.

    PubMed

    Peter, S J; Lu, L; Kim, D J; Stamatas, G N; Miller, M J; Yaszemski, M J; Mikos, A G

    2000-06-05

    Recombinant human transforming growth factor beta1 (TGF-beta1) was incorporated into microparticles of blends of poly(DL-lactic-co-glycolic acid) (PLGA) and poly(ethylene glycol) (PEG) to create a delivery vehicle for the growth factor. The entrapment efficiency of TGF-beta1 in the microparticles containing 5% PEG was 40.3 +/- 1.2% for a TGF-beta1 loading density of 6.0 ng/1 mg of microparticles. For the same loading, 17.9 +/- 0.6 and 32.1 +/- 2.5% of the loaded TGF-beta1 was released after 1 and 8 days, respectively, followed by a plateau for the remaining 3 weeks. Rat marrow stromal cells showed a dose response to TGF-beta1 released from the microparticles similar to that of added TGF-beta1, indicating the activity of TGF-beta1 was retained during microparticle fabrication and after TGF-beta1 release. An optimal TGF-beta1 dosage of 1.0 ng/mL was determined through a 3-day dose response study for maximal alkaline phosphatase (ALP) activity. The TGF-beta1 released from the microparticles loaded with 6.0 ng TGF-beta1/1 mg of microparticles for the optimal dosage of TGF-beta1 enhanced the proliferation and osteoblastic differentiation of marrow stromal cells cultured on poly(propylene fumarate) substrates. The cells showed significantly increased total cell number, ALP activity, and osteocalcin production with values reaching 138,700 +/- 3300 cells/cm(2), 22.8 +/- 1.5 x 10(-7) micromol/min/cell, and 15.9 +/- 1.5 x 10(-6) ng/cell, respectively, after 21 days as compared to cells cultured under control conditions without TGF-beta1. These results suggest that controlled release of TGF-beta1 from the PLGA/PEG blend microparticles may find applications in modulating cellular response during bone healing at a skeletal defect site.

  3. Self-renewal and pluripotency is maintained in human embryonic stem cells by co-culture with human fetal liver stromal cells expressing hypoxia inducible factor 1alpha.

    PubMed

    Ji, Lei; Liu, Yu-xiao; Yang, Chao; Yue, Wen; Shi, Shuang-shuang; Bai, Ci-xian; Xi, Jia-fei; Nan, Xue; Pei, Xue-Tao

    2009-10-01

    Human embryonic stem (hES) cells are typically maintained on mouse embryonic fibroblast (MEF) feeders or with MEF-conditioned medium. However, these xenosupport systems greatly limit the therapeutic applications of hES cells because of the risk of cross-transfer of animal pathogens. The stem cell niche is a unique tissue microenvironment that regulates the self-renewal and differentiation of stem cells. Recent evidence suggests that stem cells are localized in the microenvironment of low oxygen. We hypothesized that hypoxia could maintain the undifferentiated phenotype of embryonic stem cells. We have co-cultured a human embryonic cell line with human fetal liver stromal cells (hFLSCs) feeder cells stably expressing hypoxia-inducible factor-1 alpha (HIF-1alpha), which is known as the key transcription factor in hypoxia. The results suggested HIF-1alpha was critical for preventing differentiation of hES cells in culture. Consistent with this observation, hypoxia upregulated the expression of Nanog and Oct-4, the key factors expressed in undifferentiated stem cells. We further demonstrated that HIF-1alpha could upregulate the expression of some soluble factors including bFGF and SDF-1alpha, which are released into the microenvironment to maintain the undifferentiated status of hES cells. This suggests that the targets of HIF-1alpha are secreted soluble factors rather than a cell-cell contact mechanism, and defines an important mechanism for the inhibition of hESCs differentiation by hypoxia. Our findings developed a transgene feeder co-culture system and will provide a more reliable alternative for future therapeutic applications of hES cells.

  4. Enhancement of osteoblastic differentiation of mesenchymal stromal cells cultured by selective combination of bone morphogenetic protein-2 (BMP-2) and fibroblast growth factor-2 (FGF-2).

    PubMed

    Maegawa, Naoki; Kawamura, Kenji; Hirose, Motohiro; Yajima, Hiroshi; Takakura, Yoshinori; Ohgushi, Hajime

    2007-01-01

    It is well known that bone marrow contains mesenchymal stromal cells (MSCs), which can show osteoblastic differentiation when cultured in osteogenic medium containing ascorbic acid, beta-glycerophosphate and dexamethasone. The differentiation results in the appearance of osteoblasts, together with the formation of bone matrix; thus, in vitro cultured bone (osteoblasts/bone matrix) could be fabricated by MSC culture. This type of cultured bone has already been used in clinical cases involving orthopaedic problems. To improve the therapeutic effect of the cultured bone, we investigated the culture conditions that contributed to extensive osteoblastic differentiation. Rat bone marrow was primarily cultured to expand the number of MSCs and further cultured in osteogenic medium for 12 days. The culture was also conducted in a medium supplemented with either bone morphogenetic protein-2 (BMP-2) or fibroblast growth factor (FGF-2), or with sequential combinations of both supplements. Among them, the sequential supplementation of FGF-2 followed by BMP-2 showed high alkaline phosphatase activity, sufficient bone-specific osteocalcein expression and abundant bone matrix formation of the MSC culture. These data implied that the number of responding cells or immature osteoblasts was increased by the supplementation of FGF-2 in the early phase of the culture and that these cells can show osteoblastic differentiation, of which capability was augmented by BMP-2 in the late phase. The sequential supplementation of these cytokines into MSC culture might be suitable for the fabrication of ideal cultured bone for use in bone tissue engineering.

  5. RhoA and RhoC are involved in stromal cell-derived factor-1-induced cell migration by regulating F-actin redistribution and assembly.

    PubMed

    Luo, Jixian; Li, Dingyun; Wei, Dan; Wang, Xiaoguang; Wang, Lan; Zeng, Xianlu

    2017-05-23

    Stromal cell-derived factor-1 (SDF-1) signaling is important to the maintenance and progression of T-cell acute lymphoblastic leukemia by inducing chemotaxis migration. To identify the mechanism of SDF-1 signaling in the migration of T-ALL, Jurkat acute lymphoblastic leukemia cells were used. Results showed that SDF-1 induces Jurkat cell migration by F-actin redistribution and assembly, which is dependent on Rho activity. SDF-1 induced RhoA and RhoC activation, as well as reactive oxygen species (ROS) production, which was inhibited by Rho inhibitor. The Rho-dependent ROS production led to subsequent cytoskeleton redistribution and assembly in the process of migration. Additionally, RhoA and RhoC were involved in SDF-1-induced Jurkat cell migration. Taken together, we found a SDF-1/CXCR4-RhoA and RhoC-ROS-cytoskeleton pathway that regulates Jurkat cell migration in response to SDF-1. This work will contribute to a clearer insight into the migration mechanism of acute lymphoblastic leukemia.

  6. Basic Fibroblast Growth Factor Inhibits Apoptosis and Promotes Proliferation of Adipose-Derived Mesenchymal Stromal Cells Isolated from Patients with Type 2 Diabetes by Reducing Cellular Oxidative Stress

    PubMed Central

    2017-01-01

    Type 2 diabetes (T2D) is a chronic metabolic disorder affecting increasing number of people in developed countries. Therefore new strategies for treatment of T2D and its complications are of special interest. Nowadays, cellular therapies involving mesenchymal stromal cells that reside in adipose tissue (ASCs) constitute a promising approach; however, there are still many obstacles concerning safety and effectiveness that need to be overcome before ASCs could be engaged for the treatment of diabetes mellitus. One of the challenges is preventing ASCs from deterioration caused by elevated oxidative stress present in diabetes milieu. In the current study we investigated the effect of basic fibroblast growth factor (bFGF) treatment on ASCs isolated from patients with diagnosed T2D. We demonstrate here that cell exposition to bFGF in 5 and 10 ng/mL dosages results in improved morphology, increased proliferative activity, reduced cellular senescence and apoptosis, and decreased oxidative stress, indicating recovery of ASCs' function impaired by T2D. Therefore our results provide a support for bFGF as a potential therapeutic agent for improving stem cell-based approaches for the treatment of diabetes mellitus and its complications. PMID:28168007

  7. In vitro formation of neuroclusters in microfluidic devices and cell migration as a function of stromal-derived growth factor 1 gradients.

    PubMed

    McCutcheon, Sean; Unachukwu, Uchenna; Thakur, Ankush; Majeska, Robert; Redenti, Stephen; Vazquez, Maribel

    2017-01-02

    Central nervous system (CNS) cells cultured in vitro as neuroclusters are useful models of tissue regeneration and disease progression. However, the role of cluster formation and collective migration of these neuroclusters to external stimuli has been largely unstudied in vitro. Here, 3 distinct CNS cell types, medulloblastoma (MB), medulloblastoma-derived glial progenitor cells (MGPC), and retinal progenitor cells (RPC), were examined with respect to cluster formation and migration in response to Stromal-Derived Growth Factor (SDF-1). A microfluidic platform was used to distinguish collective migration of neuroclusters from that of individual cells in response to controlled concentration profiles of SDF-1. Cell lines were also compared with respect to expression of CXCR4, the receptor for SDF-1, and the gap junction protein Connexin 43 (Cx43). All cell types spontaneously formed clusters and expressed both CXCR4 and Cx43. RPC clusters exhibited collective chemotactic migration (i.e. movement as clusters) along SDF-1 concentration gradients. MGPCs clusters did not exhibit adhesion-based migration, and migration of MB clusters was inconsistent. This study demonstrates how controlled microenvironments can be used to examine the formation and collective migration of CNS-derived neuroclusters in varied cell populations.

  8. Cilostazol Improves Proangiogenesis Functions in Human Early Endothelial Progenitor Cells through the Stromal Cell-Derived Factor System and Hybrid Therapy Provides a Synergistic Effect In Vivo.

    PubMed

    Tseng, Shih-Ya; Chao, Ting-Hsing; Li, Yi-Heng; Cho, Chung-Lung

    2016-01-01

    This study investigated the effect of cilostazol on proangiogenesis functions in human early endothelial progenitor cells (EPCs) in vitro and the therapeutic implication of hybrid therapy with cilostazol and human early EPCs in vivo. Cilostazol significantly increased colony-forming units and enhanced differentiation of EPCs toward endothelial lineage. Treatments resulted in antiapoptotic effects and stimulated proliferation and migration and in vitro vascular tube formation through activation of stromal cell-derived factor-1 (SDF-1)/C-X-C chemokine receptor type 4 (CXCR4)/phosphatidylinositol-3 kinase (PI3K)/Akt signaling pathway. Blood flow recovery and capillary density in murine ischemic hindlimbs were significantly improved in cilostazol-treated, human early EPCs-treated, and cotreatment groups. The effects were attenuated with SDF-1α inhibition. Plasma SDF-1α levels were significantly higher in 3 active treatment groups after surgery, with greatest effects observed in hybrid therapy. The angiogenic effects of transplanted EPCs pretreated with cilostazol ex vivo were superior to untreated EPCs using in vivo Matrigel assay. Implanted EPCs were incorporated into the capillary, with pretreatment or cotreatment with cilostazol resulting in enhanced effects. Taken together, cilostazol promotes a large number of proangiogenic functions in human early EPCs through activation of SDF-1/CXCR4/PI3K/Akt signaling, and hybrid therapy provides a synergistic effect in vivo. Cotreatment may be beneficial in ischemic disease.

  9. Cell-matrix Interactions of Factor IX (FIX)-engineered human mesenchymal stromal cells encapsulated in RGD-alginate vs. fibrinogen-alginate microcapsules.

    PubMed

    Sayyar, Bahareh; Dodd, Megan; Marquez-Curtis, Leah; Janowska-Wieczorek, Anna; Hortelano, Gonzalo

    2014-04-01

    The success of cell microencapsulation technology in tissue engineering and protein delivery applications depends on the viability and functionality of the encapsulated cells, which in turn are dependent upon cell/matrix interactions. In this work, we compared the viability of cord blood-derived mesenchymal stromal cells (CB MSCs), engineered to secrete factor IX (FIX) for hemophilia treatment, and encapsulated in arginine-glycine-aspartate (RGD)-alginate versus fibrinogen-alginate microcapsules. We evaluated the effect of the biomimetic matrix on cell attachment, proliferation, and secretion of FIX. Compared with nonsupplemented alginate matrix, RGD-alginate significantly enhanced the viability of the encapsulated MSCs. Further, cells in RGD-alginate displayed distinct attachment morphology, thus suggesting that RGD-alginate can potentially be used for the encapsulation of MSCs in tissue engineering applications that require enhanced cell attachment and viability. However, our data also showed that RGD-alginate microcapsules, in contrast to fibrinogen-alginate microcapsules, did not significantly improve cell proliferation of or FIX secretion by encapsulated MSCs. Our findings suggest that evidence of cell attachment alone may not accurately predict the functionality of cells in biomimetic microcapsules.

  10. Matrix metalloproteinase 10 contributes to hepatocarcinogenesis in a novel crosstalk with the stromal derived factor 1/C-X-C chemokine receptor 4 axis.

    PubMed

    García-Irigoyen, Oihane; Latasa, Maria U; Carotti, Simone; Uriarte, Iker; Elizalde, Maria; Urtasun, Raquel; Vespasiani-Gentilucci, Umberto; Morini, Sergio; Benito, Patricia; Ladero, Jose M; Rodriguez, Jose A; Prieto, Jesus; Orbe, Josune; Páramo, Jose A; Fernández-Barrena, Maite G; Berasain, Carmen; Avila, Matias A

    2015-07-01

    Matrix metalloproteinases (MMPs) participate in tissue repair after acute injury, but also participate in cancer by promoting a protumorigenic microenvironment. Previously, we reported on a key role for MMP10 in mouse liver regeneration. Herein, we investigated MMP10 expression and function in human hepatocellular carcinoma (HCC) and diethylnitrosamine (DEN)-induced mouse hepatocarcinogenesis. MMP10 was induced in human and murine HCC tissues and cells. MMP10-deficient mice showed less HCC incidence, smaller histological lesions, reduced tumor vascularization, and less lung metastases. Importantly, expression of the protumorigenic, C-X-C chemokine receptor-4 (CXCR4), was reduced in DEN-induced MMP10-deficient mice livers. Human HCC cells stably expressing MMP10 had increased CXCR4 expression and migratory capacity. Pharmacological inhibition of CXCR4 significantly reduced MMP10-stimulated HCC cell migration. Furthermore, MMP10 expression in HCC cells was induced by hypoxia and the CXCR4 ligand, stromal-derived factor-1 (SDF1), through the extracellular signal-regulated kinase 1/2 pathway, involving an activator protein 1 site in MMP10 gene promoter. MMP10 contributes to HCC development, participating in tumor angiogenesis, growth, and dissemination. We identified a new reciprocal crosstalk between MMP10 and the CXCR4/SDF1 axis contributing to HCC progression and metastasis. To our knowledge, this is the first report addressing the role of a MMP in hepatocarcinogenesis in the corresponding genetic mouse model. © 2015 by the American Association for the Study of Liver Diseases.

  11. Connective Tissue Growth Factor in Regulation of RhoA Mediated Cytoskeletal Tension Associated Osteogenesis of Mouse Adipose-Derived Stromal Cells

    PubMed Central

    Xu, Yue; Wagner, Diane R.; Bekerman, Elena; Chiou, Michael; James, Aaron W.; Carter, Dennis; Longaker, Michael T.

    2010-01-01

    Background Cytoskeletal tension is an intracellular mechanism through which cells convert a mechanical signal into a biochemical response, including production of cytokines and activation of various signaling pathways. Methods/Principal Findings Adipose-derived stromal cells (ASCs) were allowed to spread into large cells by seeding them at a low-density (1,250 cells/cm2), which was observed to induce osteogenesis. Conversely, ASCs seeded at a high-density (25,000 cells/cm2) featured small cells that promoted adipogenesis. RhoA and actin filaments were altered by changes in cell size. Blocking actin polymerization by Cytochalasin D influenced cytoskeletal tension and differentiation of ASCs. To understand the potential regulatory mechanisms leading to actin cytoskeletal tension, cDNA microarray was performed on large and small ASCs. Connective tissue growth factor (CTGF) was identified as a major regulator of osteogenesis associated with RhoA mediated cytoskeletal tension. Subsequently, knock-down of CTGF by siRNA in ASCs inhibited this osteogenesis. Conclusions/Significance We conclude that CTGF is important in the regulation of cytoskeletal tension mediated ASC osteogenic differentiation. PMID:20585662

  12. Stromal cell-derived factor-1 receptor CXCR4-overexpressing bone marrow mesenchymal stem cells accelerate wound healing by migrating into skin injury areas.

    PubMed

    Yang, Dazhi; Sun, Shijin; Wang, Zhengguo; Zhu, Peifang; Yang, Zailiang; Zhang, Bo

    2013-06-01

    Stromal cell-derived factor-1 (SDF-1) and its membrane receptor C-X-C chemokine receptor type 4 (CXCR4) are involved in the homing and migration of multiple stem cell types, neovascularization, and cell proliferation. This study investigated the hypothesis that bone marrow-derived mesenchymal stem cells (BMSCs) accelerate skin wound healing in the mouse model by overexpression of CXCR4 in BMSCs. We compared SDF-1 expression and skin wound healing times of BALB/c mice, severe combined immunodeficiency (SCID) mice, and immune system-deficient nude mice after (60)Co radiation-induced injury of their bone marrow. The occurrence of transplanted adenovirus-transfected CXCR4-overexpressing male BMSCs in the wound area was compared with the occurrence of untransfected male BALB/c BMSCs in (60)Co-irradiated female mice skin wound healing areas by Y chromosome marker analyses. The wound healing time of BALB/c mice was 14.00±1.41 days, whereas for the nude and SCID mice it was 17.16±1.17 days and 19.83±0.76 days, respectively. Male BMSCs could be detected in the surrounding areas of (60)Co-irradiated female BALB/c mice wounds, and CXCR4-overexpressing BMSCs accelerated the wound healing time. CXCR4-overexpressing BMSCs migrate in an enhanced manner to skin wounds in a SDF-1-expression-dependent manner, thereby reducing the skin wound healing time.

  13. Stromal cell-derived factor-1{alpha} (SDF-1{alpha}/CXCL12) stimulates ovarian cancer cell growth through the EGF receptor transactivation

    SciTech Connect

    Porcile, Carola; Bajetto, Adriana . E-mail: bajetto@cba.unige.it; Barbieri, Federica; Barbero, Simone; Bonavia, Rudy; Biglieri, Marianna; Pirani, Paolo; Florio, Tullio . E-mail: florio@cba.unige.it; Schettini, Gennaro

    2005-08-15

    Ovarian cancer (OC) is the leading cause of death in gynecologic diseases in which there is evidence for a complex chemokine network. Chemokines are a family of proteins that play an important role in tumor progression influencing cell proliferation, angiogenic/angiostatic processes, cell migration and metastasis, and, finally, regulating the immune cells recruitment into the tumor mass. We previously demonstrated that astrocytes and glioblastoma cells express both the chemokine receptor CXCR4 and its ligand stromal cell-derived factor-1 (SDF-1), and that SDF-1{alpha} treatment induced cell proliferation, supporting the hypothesis that chemokines may play an important role in tumor cells' growth in vitro. In the present study, we report that CXCR4 and SDF-1 are expressed in OC cell lines. We demonstrate that SDF-1{alpha} induces a dose-dependent proliferation in OC cells, by the specific interaction with CXCR4 and a biphasic activation of ERK1/2 and Akt kinases. Our results further indicate that CXCR4 activation induces EGF receptor (EGFR) phosphorylation that in turn was linked to the downstream intracellular kinases activation, ERK1/2 and Akt. In addition, we provide evidence for cytoplasmic tyrosine kinase (c-Src) involvement in the SDF-1/CXCR4-EGFR transactivation. These results suggest a possible important 'cross-talk' between SDF-1/CXCR4 and EGFR intracellular pathways that may link signals of cell proliferation in ovarian cancer.

  14. Stromal Cell-Derived Factor-1α Plays a Crucial Role Based on Neuroprotective Role in Neonatal Brain Injury in Rats

    PubMed Central

    Mori, Miki; Matsubara, Keiichi; Matsubara, Yuko; Uchikura, Yuka; Hashimoto, Hisashi; Fujioka, Toru; Matsumoto, Takashi

    2015-01-01

    Owing to progress in perinatal medicine, the survival of preterm newborns has markedly increased. However, the incidence of cerebral palsy has risen in association with increased preterm birth. Cerebral palsy is largely caused by cerebral hypoxic ischemia (HI), for which there are no effective medical treatments. We evaluated the effects of stromal cell-derived factor-1α (SDF-1α) on neonatal brain damage in rats. Left common carotid (LCC) arteries of seven-day-old Wistar rat pups were ligated, and animals were exposed to hypoxic gas to cause cerebral HI. Behavioral tests revealed that the memory and spatial perception abilities were disturbed in HI animals, and that SDF-1α treatment improved these cognitive functions. Motor coordination was also impaired after HI but was unimproved by SDF-1α treatment. SDF-1α reduced intracranial inflammation and induced cerebral remyelination, as indicated by the immunohistochemistry results. These data suggest that SDF-1α specifically influences spatial perception abilities in neonatal HI encephalopathy. PMID:26251894

  15. Recruitment of exogenous mesenchymal stem cells in mandibular distraction osteogenesis by the stromal cell-derived factor-1/chemokine receptor-4 pathway in rats.

    PubMed

    Cao, Jian; Wang, Lei; Du, Zhao-jie; Liu, Peng; Zhang, Ya-bo; Sui, Jian-fu; Liu, Yan-pu; Lei, De-lin

    2013-12-01

    Distraction osteogenesis is widely used in orthopaedic and craniofacial surgery. However, its exact mechanism is still poorly understood. The purpose of this study was to find out whether there is systemic recruitment of mesenchymal stem cells (MSC) to the neocallus in the distraction gap by the stromal cell-derived factor-1 (SDF-1)/CXC chemokine receptor 4 (CXCR4) axis during osteogenesis. We examined the migration of MSC towards a gradient of SDF-1 in vitro. We also transplanted MSC labelled with green fluorescent protein (GFP) intravenously, with or without treatment with CXCR4-blocking antibody, into rats that had had unilateral mandibular distraction osteogenesis, and investigated the distribution of cells labelled with GFP in the soft callus after 24 h. We found that SDF-1 facilitated the migration potency of MSC both in vitro and in vivo, and this migration could be inhibited by AMD3100, an antagonist of CXCR4, and promoted by local infusion of exogenous SDF-1 into the distraction gap. This study provides a new insight into the molecular basis of how new bone is regenerated during distraction osteogenesis. Copyright © 2013 The British Association of Oral and Maxillofacial Surgeons. Published by Elsevier Ltd. All rights reserved.

  16. Norepinephrine inhibits mesenchymal stem cell chemotaxis migration by increasing stromal cell-derived factor-1 secretion by vascular endothelial cells via NE/abrd3/JNK pathway.

    PubMed

    Wu, Baolei; Wang, Lei; Yang, Xi; Mao, Ming; Ye, Chen; Liu, Peng; Yang, Zihui; Yang, Xinjie; Lei, Delin; Zhang, Chenping

    2016-12-10

    Mesenchymal stem cells (MSCs), which are physiologically maintained in vascular endothelial cell (VEC)-based niches, play a critical role in tissue regeneration. Our previous studies demonstrated that sympathetic denervation could promote MSC mobilization, thereby enhancing bone formation in distraction osteogenesis (DO), a self-tissue engineering for craniofacial and orthopeadic surgeries. However, the mechanisms on how sympathetic neurotransmitter norepinephrine (NE) regulates MSC migration are not well understood. Here we showed that deprivation of NE by transection of cervical sympathetic trunk (TCST) inhibited stromal cell-derived factor-1 (SDF-1) expression in the perivascular regions in rat mandibular DO. In vitro studies showed that NE treatment markedly upregulated p-JNK and therefore stimulated higher SDF-1 expression in VECs than control groups, and siRNA knockdown of the abrd3 gene abolished the NE-induced p-JNK activation. On the other hand, osteoblasts differentiated from MSCs showed an increase in SDF-1 secretion with lack of NE. Importantly, NE-treated VECs inhibited the MSC chemotaxis migration along the SDF-1 concentration gradient as demonstrated in a novel 3-chamber Transwell assay. Collectively, our study suggested that NE may increase the SDF-1 secretion by VECs via NE/abrd3/JNK pathway, thereby inhibiting the MSC chemotaxis migration from perivascular regions toward bone trabecular frontlines along the SDF-1 concentration gradient in bone regeneration. Copyright © 2016. Published by Elsevier Inc.

  17. Human mast cells transmigrate through human umbilical vein endothelial monolayers and selectively produce IL-8 in response to stromal cell-derived factor-1 alpha.

    PubMed

    Lin, T J; Issekutz, T B; Marshall, J S

    2000-07-01

    Mature mast cells are generally considered to be less mobile cells residing within tissue sites. However, mast cell numbers are known to increase in the context of inflammation, and mast cells are recognized to be important in regulating local neutrophil infiltration. CXC chemokines may play a critical role in this process. In this study two human mast cell-like lines, HMC-1 and KU812, and human cord blood-derived primary cultured mast cells were employed to examine role of stromal cell-derived factor-1 (SDF-1) in regulating mast cell migration and mediator production. It was demonstrated that human mast cells constitutively express mRNA and protein for CXCR4. Stimulation of human mast cells with SDF-1, the only known ligand for CXCR4, induced a significant increase in intracellular calcium levels. In vitro, SDF-1 alpha mediated dose-dependent migration of human cord blood-derived mast cells and HMC-1 cells across HUVEC monolayers. Although SDF-1 alpha did not induce mast cell degranulation, it selectively stimulated production of the neutrophil chemoattractant IL-8 without affecting TNF-alpha, IL-1beta, IL-6, GM-CSF, IFN-gamma, or RANTES production, providing further evidence of the selective modulation of mast cell function by this chemokine. These findings provide a novel, SDF-1-dependent mechanism for mast cell transendothelial migration and functional regulation, which may have important implications for the local regulation of mast cells in disease.

  18. Stromal cell-derived factor 1 (SDF-1) and its receptor CXCR4 in the formation of postburn hypertrophic scar (HTS).

    PubMed

    Ding, Jie; Hori, Keijiro; Zhang, Rainny; Marcoux, Yvonne; Honardoust, Dariush; Shankowsky, Heather A; Tredget, Edward E

    2011-01-01

    Recent data support the involvement of stromal cell-derived factor 1 (SDF-1) in the homing of bone marrow-derived stem cells to wound sites during skeletal, myocardial, vascular, lung, and skin wound repair as well as some fibrotic disorders via its receptor CXCR4. In this study, the role of SDF-1/CXCR4 signaling in the formation of hypertrophic scar (HTS) following burn injury and after treatment with systemic interferon α2b (IFNα2b) is investigated. Studies show SDF-1/CXCR4 signaling was up-regulated in burn patients, including SDF-1 level in HTS tissue and serum as well as CD14+ CXCR4+ cells in the peripheral blood mononuclear cells. In vitro, dermal fibroblasts constitutively expressed SDF-1 and deep dermal fibroblasts expressed more SDF-1 than superficial fibroblasts. Lipopolysaccharide increased SDF-1 gene expression in fibroblasts. Also, recombinant SDF-1 and lipopolysaccharide stimulated fibroblast-conditioned medium up-regulated peripheral blood mononuclear cell mobility. In the burn patients with HTS who received subcutaneous IFNα2b treatment, increased SDF-1/CXCR4 signaling was found prior to treatment which was down-regulated after IFNα2b administration, coincident with enhanced remodeling of their HTS. Our results suggest that SDF-1/CXCR4 signaling is involved in the development of HTS by promoting migration of activated CD14+ CXCR4+ cells from the bloodstream to wound sites, where they may differentiate into fibrocyte and myofibroblasts and contribute to the development of HTS.

  19. Bryostatin-5 blocks stromal cell-derived factor-1 induced chemotaxis via desensitization and down-regulation of cell surface CXCR4 receptors.

    PubMed

    He, Xing; Fang, Liyan; Wang, Jue; Yi, Yanghua; Zhang, Shuyu; Xie, Xin

    2008-11-01

    The chemokine receptor CXCR4 and its ligand, stromal cell-derived factor-1 (SDF-1), play important roles in hematopoiesis regulation, lymphocyte activation, and trafficking, as well as in developmental processes, including organogenesis, vascularization, and embryogenesis. The receptor is also involved in HIV infection and tumor growth and metastasis. Antagonists of CXCR4 have been widely evaluated for drugs against HIV and tumors. In an effort to identify novel CXCR4 antagonists, we screened a small library of compounds derived from marine organisms and found bryostatin-5, which potently inhibits chemotaxis induced by SDF-1 in Jurkat cells. Bryostatin-5 is a member of the macrolactones, and its analogue bryostatin-1 is currently being evaluated in clinical trials for its chemotherapeutic potential. The involvement of bryostatins in the SDF-1/CXCR4 signaling process has never been reported. In this study, we found that bryostatin-5 potently inhibits SDF-1-induced chemotaxis but does not affect serum-induced chemotaxis. Further studies indicate that this inhibitory effect is not due to receptor antagonism but rather to bryostatin-5-induced receptor desensitization and down-regulation of cell surface CXCR4. We also show that these effects are mediated by the activation of conventional protein kinase C.

  20. PSA3, a Protein on the Stromal Face of the Thylakoid Membrane, Promotes Photosystem I Accumulation in Cooperation with the Assembly Factor PYG71[OPEN

    PubMed Central

    Williams-Carrier, Rosalind

    2017-01-01

    PSI is a large protein-pigment complex located in the thylakoid membrane in cyanobacteria, plants, and algae. Although the structure and components of PSI are well characterized, mechanisms that orchestrate its assembly are poorly understood. In this study, we discovered a novel nucleus-encoded protein, Photosystem I Assembly3 (PSA3), that is required for PSI accumulation. PSA3 is conserved among green photosynthetic eukaryotes but is lacking in cyanobacteria. Mutations in the psa3 gene cause the specific loss of PSI in Arabidopsis (Arabidopsis thaliana) and maize (Zea mays). Ribosome profiling and pulse-labeling analyses showed that chloroplast- encoded PSI subunits are synthesized at normal rates in psa3 mutants, indicating that PSA3 is involved in the biogenesis of PSI at a posttranslational step. PSA3 resides on the stromal face of the thylakoid membrane, where it is found in a complex that is slightly smaller than PSI. Structural predictions suggest that PSA3 binds a basic peptide in a manner that is sensitive to the oxidation state of Cys pairs flanking the predicted peptide binding groove. PSA3 and the previously described PSI biogenesis factor PYG7 interact in yeast two-hybrid and bimolecular fluorescence complementation assays, and they are found in thylakoid membrane complexes of similar size. These and other results indicate that PSA3 cooperates with PYG7 to promote the stable assembly of PSI, and that the PsaC subunit is likely to be the primary target of their action. PMID:28522455

  1. Combinatorial Treatment of Bone Marrow Stem Cells and Stromal Cell-Derived Factor 1 Improves Glycemia and Insulin Production in Diabetic Mice

    PubMed Central

    Cheng, H.; Zhang, Y.C.; Wolfe, S.; Valencia, V.; Qian, K.; Shen, L.; Tang, Y.L.; Hsu, W.H.; Atkinson, M.A.; Phillips, M.I.

    2011-01-01

    Transdifferentiation of stem cells into insulin-producing cells for the treatment of diabetes have shown promising but inconsistent results. We examined the potential for attracting bone marrow stem cells (BMSCs) to the pancreas using a chemokine, stromal cell derived factor-1 (SDF-1). SDF-1 treatment markedly increased the number of GFP labeled BMSCs in the pancreas, but surprisingly, the majority was observed in liver. The liver cells had typical pancreatic endocrine cell gene expression including insulin I, insulin II, PDX-1, somatostatin and glucagon. Combined treatment with SDF-1 and BMSC transplant reduced hyperglycemia and prolonged the long-term survival of diabetic mice, and a sub group had complete normoglycemia (<150 mg/dl), restored blood insulin levels, and normal glucose tolerance. Our results suggest that SDF-1 could potentially be used to improve the homing of stem cells and β-cell regeneration. The mechanism appears to involve an increase in insulin producing cells mainly in the liver. PMID:21801807

  2. Stromal cell-derived factor-1 is upregulated by dipeptidyl peptidase-4 inhibition and has protective roles in progressive diabetic nephropathy.

    PubMed

    Takashima, Satoru; Fujita, Hiroki; Fujishima, Hiromi; Shimizu, Tatsunori; Sato, Takehiro; Morii, Tsukasa; Tsukiyama, Katsushi; Narita, Takuma; Takahashi, Takamune; Drucker, Daniel J; Seino, Yutaka; Yamada, Yuichiro

    2016-10-01

    The role of stromal cell-derived factor-1 (SDF-1) in the pathogenesis of diabetic nephropathy and its modification by dipeptidyl peptidase-4 (DPP-4) inhibition are uncertain. Therefore, we studied this independent of glucagon-like peptide-1 receptor (GLP-1R) signaling using two Akita diabetic mouse models, the diabetic-resistant C57BL/6-Akita and diabetic-prone KK/Ta-Akita. Increased SDF-1 expression was found in glomerular podocytes and distal nephrons in the diabetic-prone mice, but not in kidneys from diabetic-resistant mice. The DPP-4 inhibitor linagliptin, but not the GLP-1R agonist liraglutide, further augmented renal SDF-1 expression in both Glp1r(+/+) and Glp1r(-/-) diabetic-prone mice. Along with upregulation of renal SDF-1 expression, the progression of albuminuria, glomerulosclerosis, periglomerular fibrosis, podocyte loss, and renal oxidative stress was suppressed in linagliptin-treated Glp1r(+/+) diabetic-prone mice. Linagliptin treatment increased urinary sodium excretion and attenuated the increase in glomerular filtration rate which reflects glomerular hypertension and hyperfiltration. In contrast, selective SDF-1 receptor blockade with AMD3100 reduced urinary sodium excretion and aggravated glomerular hypertension in the Glp1r(+/+) diabetic-prone mice. Thus, DPP-4 inhibition, independent of GLP-1R signaling, contributes to protection of the diabetic kidney through SDF-1-dependent antioxidative and antifibrotic effects and amelioration of adverse renal hemodynamics.

  3. Gastrointestinal Stromal Tumors Treatment (PDQ®)—Patient Version

    Cancer.gov

    Gastrointestinal stromal tumors (GIST) are usually found on the stomach or small intestine, but they can be found anywhere in or near the GI tract. Find out about risk factors, symptoms, tests to diagnose, prognosis, staging, and treatment for gastrointestinal stromal tumors.

  4. Plasma rich in growth factors (PRGF) eye drops stimulates scarless regeneration compared to autologous serum in the ocular surface stromal fibroblasts.

    PubMed

    Anitua, E; de la Fuente, M; Muruzabal, F; Riestra, A; Merayo-Lloves, J; Orive, G

    2015-06-01

    Autologous serum (AS) eye drops was the first blood-derived product used for the treatment of corneal pathologies but nowadays PRGF arises as a novel interesting alternative to this type of diseases. The purpose of this study was to evaluate and compare the biological outcomes of autologous serum eye drops or Plasma rich in growth factors (PRGF) eye drops on corneal stromal keratocytes (HK) and conjunctival fibroblasts (HConF). To address this, blood from healthy donors was collected and processed to obtain autologous serum (AS) eye drops and plasma rich in growth factors (PRGF) eye drops. Blood-derivates were aliquoted and stored at -80°C until use. PDGF-AB, VEGF, EGF, FGFb and TGF-β1 were quantified. The potential of PRGF and AS in promoting wound healing was evaluated by means of proliferation and migration assays in HK and HConF. Fibroblast cells were induced to myofibroblast differentiation after treatment with 2.5ng/mL of TGF-β1. The capability of PRGF and AS to prevent and inhibit TGF-β1-induced differentiation was evaluated. Results showed significant higher levels of all growth factors analyzed in PRGF eye drops compared to AS. Moreover, PRGF eye drops enhanced significantly the biological outcomes of both HK and HConF, and reduced TGF-β1-induced myofibroblast differentiation in contrast to autologous serum eye drops (AS). In summary, these results suggest that PRGF exerts enhanced biological outcomes than AS. PRGF may improve the treatment of ocular surface wound healing minimizing the scar formation compared to AS. Results obtained herein suggest that PRGF protects and reverses the myofibroblast phenotype while promotes cell proliferation and migration.

  5. Soluble toll-like receptor 4 reversed attenuating effect of Chinese herbal Xiao-Qing-Long-Tang on allergen induced nerve growth factor and thymic stromal lymphopoietin

    PubMed Central

    CHANG, REN-SHIU; WANG, YU-CHIN; KAO, SHUNG-TE

    2013-01-01

    Xiao-Qing-Long-Tang (XQLT) is known to regulate allergic immune reactions. The aim of this study was to investigate the effects of XQLT on allergen-induced cytokines and associated signaling pathways. An acute allergic mouse model was used to investigate the effects of XQLT on nerve growth factor (NGF) during an allergic reaction, while human pulmonary alveolar epithelial cells (HPAEpiCs) were used to investigate the effects of XQLT on Dermatophagoides pteronyssinus group 2 (Der p 2)-induced NGF, p75 neurotrophin receptor (p75NTR) and thymic stromal lymphopoietin (TSLP) expression. XQLT was demonstrated to inhibit NGF- and p75NTR-related allergic reactions in the mouse model. XQLT also reduced the expression of Toll-like receptor 4 (TLR4) in the lungs of the model mice. XQLT inhibited Der p 2-induced NGF, TSLP and p75NTR expression in the HPAEpiC cell line. The use of recombinant soluble TLR4 (sTLR4) in a competitive assay partially attenuated the inhibitory effect of XQLT on NGF, TSLP and p75NTR expression in HPAEpiC cells. The inhibitory effect of XQLT on the Ser536 phosphorylation of p65 (nuclear factor-κB; NF-κB), a TLR4-induced factor, was also attenuated by sTLR4. In conclusion, XQLT inhibited Der p allergen-induced NGF, p75NTR and TSLP expression. This inhibition was attenuated by sTLR4. The mechanism of action of XQLT may be correlated with TLR4, a primary receptor in the innate immune system. The findings of this study may focus the search for pharmacological targets of XQLT onto TLR4, which is important in the allergen presentation pathway. PMID:24223644

  6. 5th Ovarian Cancer Consensus Conference: Individualized Therapy and Patient Factors.

    PubMed

    McGee, J; Bookman, M; Harter, P; Marth, C; McNeish, I; Moore, K N; Poveda, A; Hilpert, F; Hasegawa, K; Bacon, M; Gatsonis, C; Brand, A; Kridelka, F; Berek, J; Ottevanger, N; Levy, T; Silverberg, S; Kim, B-G; Hirte, H; Okamoto, A; Stuart, G; Ochiai, K

    2017-01-24

    This manuscript reports the consensus statements regarding the design and conduct of clinical trials in patients with newly diagnosed and recurrent epithelial ovarian cancer (EOC), following deliberation at the 5th Ovarian Cancer Consensus Conference (OCCC), held in Tokyo in November 2015. Three important questions were identified for discussion prior to the meeting and achieved consensus during the meeting: 1) What are the most important factors to be evaluated prior to initial therapy? 2) What are the most important factors to be evaluated specifically in recurrent disease? 3) Are there specific considerations for special patient subpopulations? In addition, we report a list of important unmet needs compiled during the consensus process, which is intended to guide future research initiatives.

  7. Growth Factors: Production of Monocyte Chemotactic Protein-1 (MCP-1/JE) by Bone Marrow Stromal Cells: Effect on the Migration and Proliferation of Hematopoietic Progenitor Cells.

    PubMed

    Xu, Y. X.; Talati, B. R.; Janakiraman, N.; Chapman, R. A.; Gautam, S. C.

    1999-01-01

    Recombinant chemotactic cytokines (chemokines) have been shown to modulate in vitro proliferation of hematopoietic progenitor cells. Whether bone marrow stromal cells produce chemokines and the physiological role they may have in the regulation of hematopoiesis has largely remained unexamined. We have examined the expression of monocyte chemoattractant protein-1 (MCP-1/JE) in bone marrow stromal cells and its effect on the migration and proliferation of murine hematopoietic progenitor cells. Freshly derived murine bone marrow stromal cells were found to secrete abundant amounts of MCP-1/JE, which was further increased upon stimulation of stromal cells with pro-inflammatory agents LPS, IL1-alpha, IFN-gamma, or TNF-alpha. Although culture supernatant conditioned by stromal cells exhibited chemotactic activity toward hematopoietic progenitor cells, the chemotactic activity was not due to MCP-1/JE. Furthermore, rMCP-1/JE also failed to induce migration of progenitor cells. MCP-1/JE, however, caused 20 to 30% increase in the clonal expansion of progenitor cells. Thus, although MCP-1/JE does not chemoattract hematopoietic progenitor cells it may have a role in their proliferation and clonal expansion.

  8. A retrospective analysis of environmental risk factors for the diagnosis of deep stromal abscess in 390 horses in North Central Florida from 1991 to 2013.

    PubMed

    Proietto, Laura R; Plummer, Caryn E; Maxwell, Kathleen M; Lamb, Kenneth E; Brooks, Dennis E

    2016-07-01

    The purpose of this investigation was to identify potential environmental risk factors for the diagnosis of equine deep stromal abscesses (DSA) in the subtropical climate at the University of Florida Veterinary Medical Center (UFVMC). Cases included were selected from the UFVMC medical record and imaging database, and included all cases of equine DSA diagnosed during the period from December 1991 to December 2013 in patients residing in north central Florida. Patient date of diagnosis and atmospheric data was obtained for north central Florida for the corresponding time period. Univariate and multivariate general linear models were generated testing effects and interactions between environmental conditions. When year, sulfur dioxide (SO2 ) and wind were analyzed in the presence of each other, a one-mile per hour increase in wind (P = 0.005) significantly increased the number of DSA cases by 1.63 cases per year. When the influence of temperature was evaluated in conjunction with year and nitrogen dioxide (NO2 ), the number of cases decreased by 0.1534 per year for every degree increase in temperature (°C) (P = 0.039). Wind speed is the first significant atmospheric risk factor to be identified for DSA formation in the horse. The importance of environmental variance in the incidence of DSA indicates that the pathogenesis of DSA formation may be multifactorial, interdependent and provides support in some horses for the micropuncture hypothesis of DSA formation related to the involvement of environmental conditions causing precorneal tear film and epithelial damage. © 2015 American College of Veterinary Ophthalmologists.

  9. Stromal Cell-Derived Growth Factor-1 Alpha-Elastin Like Peptide Fusion Protein Promotes Cell Migration and Revascularization of Experimental Wounds in Diabetic Mice

    PubMed Central

    Yeboah, Agnes; Maguire, Tim; Schloss, Rene; Berthiaume, Francois; Yarmush, Martin L.

    2017-01-01

    Objective: In previous work, we demonstrated the development of a novel fusion protein containing stromal cell-derived growth factor-1 alpha juxtaposed to an elastin-like peptide (SDF1-ELP), which has similar bioactivity, but is more stable in elastase than SDF1. Herein, we compare the ability of a single topical application of SDF1-ELP to that of SDF1 in healing 1 × 1 cm excisional wounds in diabetic mice. Approach: Human Leukemia-60 cells were used to demonstrate the chemotactic potential of SDF1-ELP versus SDF1 in vitro. Human umbilical vascular endothelial cells were used to demonstrate the angiogenic potential of SDF1-ELP versus SDF1 in vitro. The bioactivity of SDF1-ELP versus SDF1 after incubation in ex-vivo diabetic wound fluid was compared. The in-vivo effectiveness of SDF1-ELP versus SDF1 was compared in diabetic mice wound model by monitoring for the number of CD31+ cells in harvested wound tissues. Results: SDF1-ELP promotes the migration of cells and induces vascularization similar to SDF1 in vitro. SDF1-ELP is more stable in wound fluids compared to SDF1. In vivo, SDF1-ELP induced a higher number of vascular endothelial cells (CD31+ cells) compared to SDF1 and other controls, suggesting increased vascularization. Innovation: While growth factors have been shown to improve wound healing, this strategy is largely ineffective in chronic wounds. In this work, we show that SDF1-ELP is a promising agent for the treatment of chronic skin wounds. Conclusion: The superior in vivo performance and stability of SDF1-ELP makes it a promising agent for the treatment of chronic skin wounds. PMID:28116224

  10. Effect of umbilical cord mesenchymal stromal cells on motor functions of identical twins with cerebral palsy: pilot study on the correlation of efficacy and hereditary factors.

    PubMed

    Wang, Xiaodong; Hu, Hezhen; Hua, Rongrong; Yang, Jing; Zheng, Pei; Niu, Xinxin; Cheng, Hongbin; Dai, Guanghui; Liu, Xuebin; Zhang, Zan; An, Yihua

    2015-02-01

    The objective of this study was to compare the impact of umbilical cord-derived mesenchymal stromal cell (UCMSC) transplantation on the motor functions of identical twins with cerebral palsy (CP) and to analyze the correlation between the efficacy and hereditary factors. Eight pairs (16 individuals) of identical twins with CP were recruited and received allogenic UCMSC transplantation by means of subarachnoid injection. The gross motor function measure (GMFM) and the fine motor function measure (FMFM) were performed before and 1 and 6 months after the treatment to analyze the results of individuals before and after the therapy, between two individuals of an identical twin and among twin pairs. Repeated-measured data variance was used to analyze the GMFM and FMFM scores of patients before and 1 and 6 months after the therapy. Eight pairs (16 individuals) of children with CP had significant improvement in the GMFM at the end of the 1st and 6th months after the therapy compared with that before the therapy, whereas the amelioration of the FMFM was not statistically significant. The improvements in motor functions between two individuals of an identical twin but not among twin pairs were correlated. UCMSC transplantation significantly improves GMFM in children with CP; motor function improvements in the GMFM between two individuals of an identical twin were closely correlated, but improvements among twin pairs were not correlated. We hypothesize that hereditary factors contribute to the mechanisms of UCMSC transplantation in motor function improvement in children with CP. Copyright © 2015 International Society for Cellular Therapy. Published by Elsevier Inc. All rights reserved.

  11. A transduced living hyaline cartilage graft releasing transgenic stromal cell-derived factor-1 inducing endogenous stem cell homing in vivo.

    PubMed

    Zhang, Feng; Leong, Wenyan; Su, Kai; Fang, Yu; Wang, Dong-An

    2013-05-01

    Stromal cell-derived factor-1 (SDF-1), also known as a homing factor, is a potent chemokine that activates and directs mobilization, migration, and retention of certain cell species via systemic circulation. The responding homing cells largely consist of activated stem cells, so that, in case of tissue lesions, such SDF-1-induced cell migration may execute recruitment of endogenous stem cells to perform autoreparation and compensatory regeneration in situ. In this study, a recombinant adenoviral vector carrying SDF-1 transgene was constructed and applied to transduce a novel scaffold-free living hyaline cartilage graft (SDF-t-LhCG). As an engineered transgenic living tissue, SDF-t-LhCG is capable of continuously producing and releasing SDF-1 in vitro and in vivo. The in vitro trials were examined with ELISA, while the in vivo trials were subsequently performed via a subcutaneous implantation of SDF-t-LhCG in a nude mouse model, followed by series of biochemical and biological analyses. The results indicate that transgenic SDF-1 enhanced the presence of this chemokine in mouse's circulation system; in consequence, SDF-1-induced activation and recruitment of endogenous stem cells were also augmented in both peripheral blood and SDF-t-LhCG implant per se. These results were obtained via flow cytometry analyses on mouse blood samples and implanted SDF-t-LhCG samples, indicating an upregulation of the CXCR4(+)(SDF-1 receptor) cell population, accompanied by upregulation of the CD34(+), CD44(+), and Sca-1(+) cell populations as well as a downregulation of the CD11b(+) cell population. With the supply of SDF-1-recruited endogenous stem cells, enhanced chondrogenesis was observed in SDF-t-LhCG implants in situ.

  12. Early in-situ cellularization of a supramolecular vascular graft is modified by synthetic stromal cell-derived factor-1α derived peptides.

    PubMed

    Muylaert, Dimitri E P; van Almen, Geert C; Talacua, Hanna; Fledderus, Joost O; Kluin, Jolanda; Hendrikse, Simone I S; van Dongen, Joost L J; Sijbesma, Eline; Bosman, Anton W; Mes, Tristan; Thakkar, Shraddha H; Smits, Anthal I P M; Bouten, Carlijn V C; Dankers, Patricia Y W; Verhaar, Marianne C

    2016-01-01

    In an in-situ approach towards tissue engineered cardiovascular replacement grafts, cell-free scaffolds are implanted that engage in endogenous tissue formation. Bioactive molecules can be incorporated into such grafts to facilitate cellular recruitment. Stromal cell derived factor 1α (SDF1α) is a powerful chemoattractant of lymphocytes, monocytes and progenitor cells and plays an important role in cellular signaling and tissue repair. Short SDF1α-peptides derived from its receptor-activating domain are capable of activating the SDF1α-specific receptor CXCR4. Here, we show that SDF1α-derived peptides can be chemically modified with a supramolecular four-fold hydrogen bonding ureido-pyrimidinone (UPy) moiety, that allows for the convenient incorporation of the UPy-SDF1α-derived peptides into a UPy-modified polymer scaffold. We hypothesized that a UPy-modified material bioactivated with these UPy-SDF1α-derived peptides can retain and stimulate circulating cells in an anti-inflammatory, pro-tissue formation signaling environment. First, the early recruitment of human peripheral blood mononuclear cells to the scaffolds was analyzed in vitro in a custom-made mesofluidic device applying physiological pulsatile fluid flow. Preferential adhesion of lymphocytes with reduced expression of inflammatory factors TNFα, MCP1 and lymphocyte activation marker CD25 was found in the bioactivated scaffolds, indicating a reduction in inflammatory signaling. As a proof of concept, in-vivo implantation of the bioactivated scaffolds as rat abdominal aorta interposition grafts showed increased cellularity by CD68+ cells after 7 days. These results indicate that a completely synthetic, cell-free biomaterial can attract and stimulate specific leukocyte populations through supramolecular incorporation of short bioactive SDF1α derived peptides.

  13. [Influence of transforming growth factor-beta1 inducing time on chondrogenesis of bone marrow stromal cells (BMSCs): in vitro experiment with porcine BMSCs].

    PubMed

    Liu, Tian-yi; Zhou, Guang-dong; Wei, Xian; Wu, Xiao-li; Chen, Fu-guo; Cui, Lei; Liu, Wei; Cao, Yi-lin

    2007-08-21

    To explore the influence of transforming growth factor (TGF)-beta1 inducing time on the chondrogenesis of bone marrow stromal cells (BMSC), and on the construction of tissue engineering cartilage. BMSCs were obtained from the greater trochanters of 3 pigs, cultured, seeded onto the cylindrical scaffolds made of polyglycolic acid at the density of 5.0 x 10(7) cells/ml, and then cultured with chondrogenesis media containing TGF-beta(1) (10 ng/ml), insulin-like growth factor-I (50 microg/L), and dexamethasone (40 microg/L) to be induced by TGF-beta(1) for 2 weeks (Group A), 4 weeks (Group B), 6 weeks (Group C), 8 weeks (Group D), or 10 weeks (Group E) respectively. 10 weeks later the cylindrical scaffolds underwent gross observation and histological examination. Alcin blue method was used to examine the content of proteoglycan (GAG). Immunochemistry and Western blotting were used to examine the type II collagen. The cylindrical scaffolds underwent biomechanical analysis. HE staining showed cartilage lacunae increasing in number from the periphery to center of the cylindrical scaffolds with the extended inducing time, and were arranged more uniformly progressively. Histochemical staining showed GAG accumulation. Immunohistochemistry and Western blotting showed that the content of type II collagen increased gradually, the amount of type II collagen stabilized after 6 weeks' culture, and there was no significant difference in the content of type II collagen between Group C and Group E. Biomechanical analysis showed that the modulus of elasticity and compression strength of the groups induced for more than 6 weeks (Groups C, D, and E) were all higher then those of Groups and B. TGF-beta(1) inducing time is correlated with the cartilage engineering characteristics with BMSC as seed cells. Induction for 6 weeks helps construct tissue engineered cartilage with good tissue structure, chemical composition and biomechanical properties.

  14. Nuclear Receptor Co-Regulator Krüppel-like Factor 9 in Human Endometrial Stromal Cell Differentiation

    USDA-ARS?s Scientific Manuscript database

    The biological actions of ligand-bound estrogen (E) and progesterone (P) receptors are dependent on coregulator partner proteins. We have identified Krüppel-like Factor 9 (KLF9) as important for E and P actions in endometrial cells. Ablation of KLF9 in mice resulted in subfertility due partly to alt...

  15. A GC-rich element confers epidermal growth factor responsiveness to transcription from the gastrin promoter.

    PubMed Central

    Merchant, J L; Demediuk, B; Brand, S J

    1991-01-01

    Epidermal growth factor (EGF) and transforming growth factor alpha are important determinants of mucosal integrity in the gastrointestinal tract, and they act both directly and indirectly to prevent ulceration in the stomach. Consistent with this physiological role, EGF stimulates transcription of gastrin, a peptide hormone which regulates gastric acid secretion and mucosal growth. EGF stimulation of gastrin transcription is mediated by a GC-rich gastrin EGF response element (gERE) (GGGGCGGGGTGGGGGG) which lies between -54 and -68 in the human gastrin promoter. The gERE sequence also confers weaker responsiveness to phorbol ester stimulation. The gERE sequence differs from previously described EGF response elements. The gERE DNA sequence specifically interacts with a GH4 DNA-binding protein distinct from previously described transcription factors (Egr-1 and AP2) which bind GC-rich sequences and mediate transcriptional activation by growth factors. Furthermore, the gERE element does not bind the Sp1 transcription factor even though the gERE sequence contains a high-affinity Sp1-binding site (GGCGGG). Images PMID:2017173

  16. Management of Gastrointestinal Stromal Tumors.

    PubMed

    von Mehren, Margaret

    2016-10-01

    Gastrointestinal stromal tumors had the reputation for poor outcomes because of their lack of response to nonsurgical interventions. The discovery of gain-of-function mutations involving receptor tyrosine kinase growth factor receptors altered the biological understanding and management. Beginning in 2000, management of these tumors has changed dramatically because of the availability of tyrosine kinase inhibitors. The role of surgery continues to be refined. This article reviews how surgery and systemic therapy are being used, incorporating definitions of risk. Decisions on how to treat a patient is based on the risk of progression, pathologic characteristics, and tumor location. Copyright © 2016 Elsevier Inc. All rights reserved.

  17. Overexpression of insulin-like growth factor 1 receptor and frequent mutational inactivation of SDHA in wild-type SDHB-negative gastrointestinal stromal tumors.

    PubMed

    Belinsky, Martin G; Rink, Lori; Flieder, Douglas B; Jahromi, Mona S; Schiffman, Joshua D; Godwin, Andrew K; Mehren, Margaret von

    2013-02-01

    Approximately 15% of gastrointestinal stromal tumors (GISTs) in adults and 85% in children lack mutations in KIT and PDGFRA and are known as wild-type GISTs. Wild-type GISTs from adults and children express high levels of insulin-like growth factor 1 receptor (IGF1R) and exhibit stable genomes compared to mutant GISTs. Pediatric wild-type GISTs, GISTs from the multitumor Carney-Stratakis syndrome, and the Carney triad share other clinicopathological properties (e.g., early-onset, multifocal GISTs with epitheliod cell morphology), suggesting a common etiology. Carney-Stratakis is an inherited association of GIST and paragangliomas caused by germline mutations in succinate dehydrogenase (SDH) genes. The connection between defective cellular respiration and GIST pathology has been strengthened by the utilization of SDHB immunohistochemistry to identify SDH deficiency in pediatric GISTs, syndromic GISTs, and some adult wild-type GISTs. SDHB and IGF1R expression was examined in 12 wild-type and 12 mutant GIST cases. Wild-type GISTs were screened for coding-region alterations in SDH genes and for chromosomal aberrations using genome-wide single-nucleotide polymorphism and MIP arrays. SDHB-deficiency, identified in 11/12 wild-type GIST cases, was tightly associated with overexpression of IGF1R protein and transcript. Biallelic inactivation of the SDHA gene was a surprisingly frequent event, identified in 5 of 11 SDHB-negative cases, generally due to germline point mutations accompanied by somatic SDHA allelic losses. As a novel finding, inactivation of the SDHC gene from a combination of a heterozygous coding-region mutation and hypermethylation of the wild-type allele was found in one SDHB-negative case. Copyright © 2012 Wiley Periodicals, Inc.

  18. Human adipose-derived mesenchymal stromal cell pigment epithelium–derived factor cytotherapy modifies genetic and epigenetic profiles of prostate cancer cells

    PubMed Central

    Zolochevska, Olga; Shearer, Joseph; Ellis, Jayne; Fokina, Valentina; Shah, Forum; Gimble, Jeffrey M.; Figueiredo, Marxa L.

    2014-01-01

    Background aims Adipose-derived mesenchymal stromal cells (ASCs) are promising tools for delivery of cytotherapy against cancer. However, ASCs can exert profound effects on biological behavior of tumor cells. Our study aimed to examine the influence of ASCs on gene expression and epigenetic methylation profiles of prostate cancer cells as well as the impact of expressing a therapeutic gene on modifying the interaction between ASCs and prostate cancer cells. Methods ASCs were modified by lentiviral transduction to express either green fluorescent protein as a control or pigment epithelium–derived factor (PEDF) as a therapeutic molecule. PC3 prostate cancer cells were cultured in the presence of ASC culture–conditioned media (CCM), and effects on PC3 or DU145. Ras cells were examined by means of real-time quantitative polymerase chain reaction, EpiTect methyl prostate cancer–focused real-time quantitative polymerase chain reaction arrays, and luciferase reporter assays. Results ASCs transduced with lentiviral vectors were able to mediate expression of several tumor-inhibitory genes, some of which correlated with epigenetic methylation changes on cocultured PC3 prostate cancer cells. When PC3 cells were cultured with ASC-PEDF CCM, we observed a shift in the balance of gene expression toward tumor inhibition, which suggests that PEDF reduces the potential tumor-promoting activity of unmodified ASCs. Conclusions These results suggest that ASC-PEDF CCM can promote reprogramming of tumor cells in a paracrine manner. An improved understanding of genetic and epigenetic events in prostate cancer growth in response to PEDF paracrine therapy would enable a more effective use of ASC-PEDF, with the goal of achieving safer yet more potent anti-tumor effects. PMID:24424267

  19. Platelet lysate and granulocyte-colony stimulating factor serve safe and accelerated expansion of human bone marrow stromal cells for stroke therapy.

    PubMed

    Yamauchi, Tomohiro; Saito, Hisayasu; Ito, Masaki; Shichinohe, Hideo; Houkin, Kiyohiro; Kuroda, Satoshi

    2014-12-01

    Autologous human bone marrow stromal cells (hBMSCs) should be expanded in the animal serum-free condition within clinically relevant periods in order to secure safe and effective cell therapy for ischemic stroke. This study was aimed to assess whether the hBMSCs enhance their proliferation capacity and provide beneficial effect in the infarct brain when cultured with platelet lysate (PL) and granulocyte-colony stimulating factor (G-CSF). The hBMSCs were cultured in the fetal calf serum (FCS)-, PL-, or PL/G-CSF-containing medium. Cell growth kinetics was analyzed. The hBMSCs-PL, hBMSC-PL/G-CSF, or vehicle was stereotactically transplanted into the ipsilateral striatum of the rats subjected to permanent middle cerebral artery occlusion 7 days after the insult. Motor function was assessed for 8 weeks, and the fate of transplanted hBMSCs was examined using immunohistochemistry. As the results, the hBMSCs-PL/G-CSF showed more enhanced proliferation than the hBMSCs-FCS and hBMSCs-PL. Transplantation of hBMSCs expanded with the PL- or PL/G-CSF-containing medium equally promoted functional recovery compared with the vehicle group. Histological analysis revealed that there were no significant differences in their migration, survival, and neural differentiation in the infarct brain between the hBMSCs-PL and hBMSCs-PL/G-CSF. These findings strongly suggest that the combination of PL and G-CSF may accelerate hBMSC expansion and serve safe cell therapy for patients with ischemic stroke at clinically relevant timing.

  20. Mesenchymal stem cells/multipotent stromal cells (MSCs) are glycolytic and thus glucose is a limiting factor of in vitro models of MSC starvation.

    PubMed

    Nuschke, Austin; Rodrigues, Melanie; Wells, Albin W; Sylakowski, Kyle; Wells, Alan

    2016-12-01

    Mesenchymal stem/multipotent stromal cells (MSCs) contribute to tissue repair but are challenged during wound healing when the blood supply is disrupted, thereby limiting nutrient delivery. Survival mechanisms against 'starvation' include autophagy, which we previously found to enhance differentiation efficiency. MSC response to models of in vitro nutrient deprivation are of great interest for improving MSC survival and therapeutic efficacy; however, the rate-limiting nutrients are unknown. MSC responses to culture nutrient and/or serum deprivations were assessed through light microscopy, cell survival, and measurements of metabolic levels. Glucose uptake was determined through conditioned media analyses over 3 days of culture. The Seahorse XF24 Flux analysis system was used to determine oxygen consumption and extracellular acidification for glycolytic metabolism. MSC autophagic response to these conditions was assessed via immunoblots for LC3-I and LC3-II, markers of autophagosome turnover. We more closely examined limiting nutritional factors to MSC survival in vitro, finding that glucose is rapidly utilized/depleted whereas amino acids and other required nutrients were used sparingly. This finding concurred with metabolic analyses that showed a primarily glycolytic character to the MSCs at steady state. MSC autophagy, previously linked to MSC function through a unique accumulated autophagosome phenotype, also responded quickly to changes in glucose concentration, with drastic LC3-II changes within 24 h of glucose concentration shifts. Our results demonstrated a rapid uptake of glucose in MSC cultures that was due to a highly glycolytic phenotype for the cells; MSC starvation with serum or other nutrients appears to have a less notable effect on the cells. These findings highlight the importance of glucose and glucose metabolism on MSC function. The conditions and cellular responses outlined here may be essential in modeling MSC nutrient deprivation.

  1. Self-assembling nanoparticles encapsulating zoledronic acid inhibit mesenchymal stromal cells differentiation, migration and secretion of proangiogenic factors and their interactions with prostate cancer cells

    PubMed Central

    Pivetta, Eliana; Colombatti, Alfonso; Boccellino, Mariarosaria; Amler, Evzen; Normanno, Nicola; Caraglia, Michele; De Rosa, Giuseppe; Aldinucci, Donatella

    2017-01-01

    Zoledronic Acid (ZA) rapidly concentrates into the bone and reduces skeletal-related events and pain in bone metastatic prostate cancer (PCa), but exerts only a limited or absent impact as anti-cancer activity. Recently, we developed self-assembling nanoparticles (NPS) encapsulating zoledronic acid (NZ) that allowed a higher intratumor delivery of the drug compared with free zoledronic acid (ZA) in in vivo cancer models of PCa. Increasing evidence suggests that Bone Marrow (BM) Mesenchymal stromal cells (BM-MSCs) are recruited into the stroma of developing tumors where they contribute to progression by enhancing tumor growth and metastasis. We demonstrated that treatment with NZ decreased migration and differentiation into adipocytes and osteoblasts of MSCs and inhibited osteoclastogenesis. Treatment with NZ reduced the capability of MSCs to promote the migration and the clonogenic growth of the prostate cancer cell lines PC3 and DU145. The levels of Interleukin-6 and of the pro-angiogenic factors VEGF and FGF-2 were significantly reduced in MSC-CM derived from MSCs treated with NZ, and CCL5 secretion was almost totally abolished. Moreover, treatment of MSCs with supernatants from PC3 cells, leading to tumor-educated MSCs (TE-MSCs), increased the secretion of IL-6, CCL5, VEGF and FGF-2 by MSCs and increased their capability to increase PC3 cells clonogenic growth. Treatment with NZ decreased cytokine secretion and the pro-tumorigenic effects also of TE-MSCS. In conclusion, demonstrating that NZ is capable to inhibit the cross talk between MSCs and PCa, this study provides a novel insight to explain the powerful anticancer activity of NZ on PCa. PMID:28477013

  2. Self-assembling nanoparticles encapsulating zoledronic acid inhibit mesenchymal stromal cells differentiation, migration and secretion of proangiogenic factors and their interactions with prostate cancer cells.

    PubMed

    Borghese, Cinzia; Casagrande, Naike; Pivetta, Eliana; Colombatti, Alfonso; Boccellino, Mariarosaria; Amler, Evzen; Normanno, Nicola; Caraglia, Michele; De Rosa, Giuseppe; Aldinucci, Donatella

    2017-06-27

    Zoledronic Acid (ZA) rapidly concentrates into the bone and reduces skeletal-related events and pain in bone metastatic prostate cancer (PCa), but exerts only a limited or absent impact as anti-cancer activity. Recently, we developed self-assembling nanoparticles (NPS) encapsulating zoledronic acid (NZ) that allowed a higher intratumor delivery of the drug compared with free zoledronic acid (ZA) in in vivo cancer models of PCa. Increasing evidence suggests that Bone Marrow (BM) Mesenchymal stromal cells (BM-MSCs) are recruited into the stroma of developing tumors where they contribute to progression by enhancing tumor growth and metastasis.We demonstrated that treatment with NZ decreased migration and differentiation into adipocytes and osteoblasts of MSCs and inhibited osteoclastogenesis. Treatment with NZ reduced the capability of MSCs to promote the migration and the clonogenic growth of the prostate cancer cell lines PC3 and DU145. The levels of Interleukin-6 and of the pro-angiogenic factors VEGF and FGF-2 were significantly reduced in MSC-CM derived from MSCs treated with NZ, and CCL5 secretion was almost totally abolished. Moreover, treatment of MSCs with supernatants from PC3 cells, leading to tumor-educated MSCs (TE-MSCs), increased the secretion of IL-6, CCL5, VEGF and FGF-2 by MSCs and increased their capability to increase PC3 cells clonogenic growth. Treatment with NZ decreased cytokine secretion and the pro-tumorigenic effects also of TE-MSCS. In conclusion, demonstrating that NZ is capable to inhibit the cross talk between MSCs and PCa, this study provides a novel insight to explain the powerful anticancer activity of NZ on PCa.

  3. Local delivery of chondroitinase ABC with or without stromal cell-derived factor 1α promotes functional repair in the injured rat spinal cord.

    PubMed

    Pakulska, Malgosia M; Tator, Charles H; Shoichet, Molly S

    2017-07-01

    Traumatic spinal cord injury (SCI) is a devastating event for which functional recovery remains elusive. Due to the complex nature of SCI pathology, a combination treatment strategy will likely be required for success. We hypothesized that tissue and functional repair would be achieved in a rat model of impact-compression SCI by combining degradation of the glial scar, using chondroitinase ABC (ChABC), with recruitment of endogenous neural precursor cells (NPCs), using stromal cell-derived factor 1α (SDF). To test this hypothesis, we designed a crosslinked methylcellulose hydrogel (XMC) for minimally invasive, localized, and sustained intrathecal drug delivery. ChABC was released from XMC using protein-peptide affinity interactions while SDF was delivered by electrostatic affinity interactions from polymeric nanoparticles embedded in XMC. Rats with SCI were treated acutely with a combination of SDF and ChABC, SDF alone, ChABC alone, or vehicle alone, and compared to injury only. Treatment with ChABC, both alone and in combination with SDF, resulted in faster and more sustained behavioural improvement over time than other groups. The significantly reduced chondroitin sulfate proteoglycan levels and greater distribution of NPCs throughout the spinal cord tissue with ChABC delivery, both alone and in combination with SDF, may explain the improved locomotor function. Treatment with SDF alone had no apparent effect on NPC number or distribution nor synergistic effect with ChABC delivery. Thus, in this model of SCI, tissue and functional repair is attributed to ChABC. Copyright © 2017 Elsevier Ltd. All rights reserved.

  4. Recruitment of mesenchymal stem cells and macrophages by dual release of stromal cell-derived factor-1 and a macrophage recruitment agent enhances wound closure.

    PubMed

    Kim, Yang-Hee; Tabata, Yasuhiko

    2016-04-01

    In this study, the wound closure of mouse skin defects was examined in terms of recruitment of mesenchymal stem cells (MSC) and macrophages. For the cells recruitment, stromal derived factor-1 (SDF-1) of a MSC recruitment agent and sphingosine-1 phosphate agonist (SEW2871) of a macrophages recruitment agent were incorporated into gelatin hydrogels, and then released in a controlled fashion. When applied to a skin wound defect of mice, gelatin hydrogels incorporating mixed 500 ng SDF-1 and 0.4, 0.8, or 1.6 mg SEW2871-micelles recruited a higher number of both MSC and macrophages than those incorporating SDF-1 or phosphate buffered saline. However, the number of M1 phenotype macrophages for the hydrogel incorporating mixed SDF-1 and SEW2871-micelles recruited was remarkably low to a significant extent compared with that for those hydrogel incorporating 0.4, 0.8, or 1.6 mg SEW2871-micelles. On the other hand, the number of M2 macrophages 3 days after the implantation of the hydrogels incorporating SDF-1 and 0.4 mg SEW2871-micelles significantly increased compared with that for other hydrogels. In vivo experiments revealed the hydrogels incorporating SDF-1 and 0.4 mg SEW2871-micelles promoted the wound closure of skin defect to a significant stronger extent than those incorporating SEW2871-micelles, SDF-1, and a mixture of SDF-1 and higher doses of SEW2871-micelles. It is concluded that the in vivo recruitment of MSC and macrophages to the defects may contribute to the tissue regeneration of skin wound.

  5. Systemic BMSC homing in the regeneration of pulp-like tissue and the enhancing effect of stromal cell-derived factor-1 on BMSC homing.

    PubMed

    Zhang, Li-Xia; Shen, Li-Li; Ge, Shao-Hua; Wang, Li-Mei; Yu, Xi-Jiao; Xu, Quan-Chen; Yang, Pi-Shan; Yang, Cheng-Zhe

    2015-01-01

    Pulp regeneration caused by endogenous cells homing has become the new research spot in endodontics. However, the source of functional cells that are involved in and contributed to the reconstituting process has not been identified. In this study, the possible role of systemical BMSC in pulp regeneration and the effect of stromal cell-derived factor-1 (SDF-1) on stem cell recruitment and angiogenesis were evaluated. 54 mice were divided into three groups: SDF-1 group (subcutaneous pockets containing roots with SDF-1 absorbed neutralized collagen gel and the green fluorescent protein (GFP) positive BMSCs transplantation via the tail vein), SDF-1-free group (pockets containing roots with gel alone and GFP + BMSCs transplantation) and Control group (pockets containing roots with gel alone). The animals were sacrificed after the roots were implanted into subcutaneous pockets for 3 weeks. Histomorphometric analysis was performed to evaluate the regenerated tissue in the canal by hematoxylin and eosin (HE) staining. The homing of the transplanted BMSCs was monitored with a fluorescence microscope and immunohistochemical analysis. The expression of ALP in new formed tissue was detected immunohistochemically. Dental-pulp-like tissue and new vessels were regenerated and GFP-positive BMSCs and expression of ALP could be observed in both SDF-1 group and SDF-1-free group. Furthermore, more GFP+ cells, stronger expression of ALP and stronger angiogenesis were found in the SDF-1 group than in the SDF-1-free group. To conclude, systemic BMSC can home to the root canal and participate in dental-pulp-like tissue regeneration. Intracanal application of SDF-1 may enhance BMSC homing efficiency and angiogenesis.

  6. Quantification of Intact and Truncated Stromal Cell-Derived Factor-1α in Circulation by Immunoaffinity Enrichment and Tandem Mass Spectrometry

    NASA Astrophysics Data System (ADS)

    Wang, Weixun; Choi, Bernard K.; Li, Wenyu; Lao, Zhege; Lee, Anita Y. H.; Souza, Sandra C.; Yates, Nathan A.; Kowalski, Timothy; Pocai, Alessandro; Cohen, Lucinda H.

    2014-04-01

    Stromal cell-derived factor 1α (SDF-1α) or CXCL12 is a small pro-inflammatory chemoattractant cytokine and a substrate of dipeptidyl peptidase IV (DPP-IV). Proteolytic cleavage by DPP-IV inactivates SDF-1α and attenuates its interaction with CXCR4, its cell surface receptor. To enable investigation of suppression of such inactivation with pharmacologic inhibition of DPP-IV, we developed quantitative mass spectrometric methods that differentiate intact SDF-1α from its inactive form. Using top-down strategy in quantification, we demonstrated the unique advantage of keeping SDF-1α's two disulfide bridges intact in the analysis. To achieve the optimal sensitivity required for quantification of intact and truncated SDF-1α at endogenous levels in blood, we coupled nano-flow tandem mass spectrometry with antibody-based affinity enrichment. The assay has a quantitative range of 20 pmol/L to 20 nmol/L in human plasma as well as in rhesus monkey plasma. With only slight modification, the same assay can be used to quantify SDF-1α in mice. Using two in vivo animal studies as examples, we demonstrated that it was critical to differentiate intact SDF-1α from its truncated form in the analysis of biomarkers for pharmacologic inhibition of DPP-IV activity. These novel methods enable translational research on suppression of SDF-1 inactivation with DPP-IV inhibition and can be applied to relevant clinical samples in the future to yield new insights on change of SDF-1α levels in disease settings and in response to therapeutic interventions.

  7. Modulation of stromal cell-derived factor 1 alpha (SDF-1α) and its receptor CXCR4 in Porphyromonas gingivalis-induced periodontal inflammation.

    PubMed

    Sun, Jiang; Nemoto, Eiji; Hong, Guang; Sasaki, Keiichi

    2016-07-22

    The production of chemokines by tissue resident cells during inflammation is considered one of the main mechanisms involved in the formation of inflammatory infiltrates. Fibroblasts are the main resident cell type in gingival and periodontal ligament tissues, and their ability to produce chemokine stromal cell-derived factor 1 alpha (SDF-1α) and its receptor CXCR4 under stimulation by gram negative bacteria, Porphyromonas gingivalis, commonly found in periodontal infections was investigated. Western blots were used to assess SDF-1α and CXCR4 protein expression levels in human gingival fibroblast cells (HGF-1) induced by Lipopolysaccharide (LPS) from P. gingivalis in the presence or absence of LY294002, a highly selective inhibitor of PI-3K/Akt. RT-PCR and quantitative Real-time PCR was performed using gingival mRNAs from periodontitis patients. Immunohistochemistry was performed to analyze the expression and subcellular localization of SDF-1α and CXCR4, together with NF-kβ phosphorylation, in specimens from patients with periodontitis and in an experimental rat periodontitis model. We found that P. gingivalis LPS up-regulated SDF-1α and CXCR4 protein levels and elevated phosphorylation of the SDF-1α-responsive NF-kβ and Akt at 24 h in HGF-1 cells. SDF-1α and CXCR4 mRNA and protein expression levels were high in all patients with periodontitis. In the P. gingivalis-induced rat experimental periodontitis model, SDF-1α and CXCR4 immunoreactivity was higher in gingival and periodontal ligament tissues compared to the control. Our data showed that PI-3K/Akt is an upstream participant in the P. gingivalis LPS-mediated induction of SDF-1α. Taken together, these results suggest that the chemokine SDF-1α and its receptor CXCR4 contribute to P. gingivalis-induced periodontal inflammation.

  8. Systemic BMSC homing in the regeneration of pulp-like tissue and the enhancing effect of stromal cell-derived factor-1 on BMSC homing

    PubMed Central

    Zhang, Li-Xia; Shen, Li-Li; Ge, Shao-Hua; Wang, Li-Mei; Yu, Xi-Jiao; Xu, Quan-Chen; Yang, Pi-Shan; Yang, Cheng-Zhe

    2015-01-01

    Pulp regeneration caused by endogenous cells homing has become the new research spot in endodontics. However, the source of functional cells that are involved in and contributed to the reconstituting process has not been identified. In this study, the possible role of systemical BMSC in pulp regeneration and the effect of stromal cell-derived factor-1 (SDF-1) on stem cell recruitment and angiogenesis were evaluated. 54 mice were divided into three groups: SDF-1 group (subcutaneous pockets containing roots with SDF-1 absorbed neutralized collagen gel and the green fluorescent protein (GFP) positive BMSCs transplantation via the tail vein), SDF-1-free group (pockets containing roots with gel alone and GFP + BMSCs transplantation) and Control group (pockets containing roots with gel alone). The animals were sacrificed after the roots were implanted into subcutaneous pockets for 3 weeks. Histomorphometric analysis was performed to evaluate the regenerated tissue in the canal by hematoxylin and eosin (HE) staining. The homing of the transplanted BMSCs was monitored with a fluorescence microscope and immunohistochemical analysis. The expression of ALP in new formed tissue was detected immunohistochemically. Dental-pulp-like tissue and new vessels were regenerated and GFP-positive BMSCs and expression of ALP could be observed in both SDF-1 group and SDF-1-free group. Furthermore, more GFP+ cells, stronger expression of ALP and stronger angiogenesis were found in the SDF-1 group than in the SDF-1-free group. To conclude, systemic BMSC can home to the root canal and participate in dental-pulp-like tissue regeneration. Intracanal application of SDF-1 may enhance BMSC homing efficiency and angiogenesis. PMID:26617734

  9. Sustained myocardial production of stromal cell-derived factor-1α was associated with left ventricular adverse remodeling in patients with myocardial infarction.

    PubMed

    Uematsu, Manabu; Yoshizaki, Toru; Shimizu, Takuya; Obata, Jun-ei; Nakamura, Takamitsu; Fujioka, Daisuke; Watanabe, Kazuhiro; Watanabe, Yosuke; Kugiyama, Kiyotaka

    2015-11-15

    The role of stromal cell-derived factor-1α (SDF-1α) expressed in infarcted myocardium is unknown in humans. We examined whether SDF-1α produced in an infarcted myocardial lesion may play a role in left ventricle (LV) remodeling and dysfunction in patients with acute myocardial infarction (AMI). We measured SDF-1α levels in plasma obtained from aortic root (AO) and anterior interventricular vein (AIV) in the early phase (2 wk after MI) and the chronic phase (6 mo after MI) in 80 patients with anterior MI. An increment in SDF-1α level from AO to AIV, reflecting SDF-1α release from infarcted myocardium, was more frequent in patients with MI in the early phase of MI [n = 52 (65%), P = 0.03] but not in the chronic phase of MI [n = 46 (58%), P = 0.11] compared with that in control patients [n = 6/17 (35%)]. On linear regression analysis, the transmyocardial gradient in SDF-1α level in the chronic phase of MI was correlated with percentage changes in LV end-diastolic volume index (r = 0.39, P < 0.001), LV end-systolic volume index (r = 0.38, P < 0.001), and LV ejection fraction (r = -0.26, P = 0.01) 6 mo after AMI. By contrast, the transmyocardial gradient of SDF-1α in the early phase of MI had no significant correlations. In conclusion, the production of SDF-1α in infarcted myocardium in the chronic phase of MI was associated with LV adverse remodeling and progressive dysfunction in AMI survivors.

  10. Selective JAK/STAT3 signalling regulates transcription of colony stimulating factor-2 and -3 in Concanavalin-A-activated mesenchymal stromal cells.

    PubMed

    Zgheib, Alain; Pelletier-Bonnier, Émilie; Levros, Louis-Charles; Annabi, Borhane

    2013-08-01

    Human bone marrow-derived mesenchymal stromal cells (MSCs) express Toll-like receptors (TLRs) and produce cytokines and chemokines, all of which contribute to these cells' immunomodulatory and proangiogenic properties. Among the secreted cytokines, colony-stimulating factors (CSFs) regulate angiogenesis through activation of endothelial cell proliferation and migration. Since MSC are recruited within hypoxic tumors where they signal paracrine-regulated angiogenesis, the aim of this study was to evaluate which CSF members are expressed and are inducible in activated MSC. Furthermore, we investigated the JAK/STAT signal transducing pathway that may impact on CSF transcription. MSC were activated with Concanavalin-A (ConA), a TLR-2/6 agonist as well as a membrane type-1 matrix metalloproteinase (MT1-MMP) inducer, and we found increased transcription of granulocyte macrophage-CSF (GM-CSF, CSF-2), granulocyte CSF (G-CSF, CSF-3), and MT1-MMP. Gene silencing of either STAT3 or MT1-MMP prevented ConA-induced phosphorylation of STAT3, and reversed ConA effects on CSF-2 and CSF-3. Treatment with the Janus Kinase (JAK)2 inhibitor AG490 antagonized the ConA induction of MT1-MMP and CSF-2, while the pan-JAK inhibitor Tofacitinib reversed ConA-induced CSF-2 and -3 gene expression. Silencing of JAK2 prevented the ConA-mediated increase of CSF-2, while silencing of JAK1, JAK3 and TYK2 prevented the increase in CSF-3. Given that combined TLR-activation and locally-produced CSF-2 and CSF-3 could regulate immunomodulation and neovascularization, pharmacological targeting of TLR-2/6-induced MT1-MMP/JAK/STAT3 signalling pathway may prevent MSC contribution to tumor development. Copyright © 2013 Elsevier Ltd. All rights reserved.

  11. Stromal cell-derived factor-1-directed bone marrow mesenchymal stem cell migration in response to inflammatory and/or hypoxic stimuli

    PubMed Central

    Yu, Yang; Wu, Rui-Xin; Gao, Li-Na; Xia, Yu; Tang, Hao-Ning; Chen, Fa-Ming

    2016-01-01

    ABSTRACT Directing cell trafficking toward a target site of interest is critical for advancing stem cell therapy in clinical theranostic applications. In this study, we investigated the effects of inflammatory and/or hypoxic stimuli on the migration of bone marrow mesenchymal stem cells (BMMSCs) during in vitro culture and after in vivo implantation. Using tablet scratch experiments and observations from a transwell system, we found that both inflammatory and hypoxic stimuli significantly enhanced cell migration. However, the combination of inflammatory and hypoxic stimuli did not result in a synergistic effect. The presence of stromal cell-derived factor-1 (SDF-1) significantly enhanced cell migration irrespective of the incubation conditions, and these positive effects could be blocked by treatment with AMD3100. Based on a time course experiment, we found that preconditioning cells with either inflammatory or hypoxic stimuli for 24 h or with both stimuli for 12 h led to high levels of chemokine receptor type 4 (CXCR4) expression. In vivo studies further demonstrated that pretreatment of BMMSCs with inflammatory and/or hypoxic stimuli resulted in an increased number of systemically injected cells migrating toward skin injuries, and local SDF-1 administration significantly increased cell migration. These findings suggest that in vitro control of either inflammatory or hypoxic stimuli has significant potential to enhance SDF-1-directed BMMSC migration via the upregulation of CXCR4 expression. Although combining the stimuli did not necessarily lead to a synergistic effect, the potential to reduce the dose and time required for cell preconditioning indicates that combinations of various strategies warrant further exploration. PMID:26745021

  12. Ultrasound-targeted stromal cell-derived factor-1-loaded microbubble destruction promotes mesenchymal stem cell homing to kidneys in diabetic nephropathy rats.

    PubMed

    Wu, Shengzheng; Li, Lu; Wang, Gong; Shen, Weiwei; Xu, Yali; Liu, Zheng; Zhuo, Zhongxiong; Xia, Hongmei; Gao, Yunhua; Tan, Kaibin

    2014-01-01

    Mesenchymal stem cell (MSC) therapy has been considered a promising strategy to cure diabetic nephropathy (DN). However, insufficient MSCs can settle in injured kidneys, which constitute one of the major barriers to the effective implementation of MSC therapy. Stromal cell-derived factor-1 (SDF-1) plays a vital role in MSC migration and involves activation, mobilization, homing, and retention, which are presumably related to the poor homing in DN therapy. Ultrasound-targeted microbubble destruction has become one of the most promising strategies for the targeted delivery of drugs and genes. To improve MSC homing to DN kidneys, we present a strategy to increase SDF-1 via ultrasound-targeted microbubble destruction. In this study, we developed SDF-1-loaded microbubbles (MB(SDF-1)) via covalent conjugation. The characterization and bioactivity of MB(SDF-1) were assessed in vitro. Target release in the targeted kidneys was triggered with diagnostic ultrasound in combination with MB(SDF-1). The related bioeffects were also elucidated. Early DN was induced in rats with streptozotocin. Green fluorescent protein-labeled MSCs were transplanted intravenously following the target release of SDF-1 in the kidneys of normal and DN rats. The homing efficacy was assessed by detecting the implanted exogenous MSCs at 24 hours. The in vitro results showed an impressive SDF-1 loading efficacy of 79% and a loading content of 15.8 μg/mL. MB(SDF-1) remained bioactive as a chemoattractant. In the in vivo study, SDF-1 was successfully released in the targeted kidneys. The homing efficacy of MSCs to DN kidneys after the target release of SDF-1 was remarkably ameliorated at 24 hours compared with control treatments in normal rats and DN rats. In conclusion, ultrasound-targeted MB(SDF-1) destruction could promote the homing of MSCs to early DN kidneys and provide a novel potential therapeutic approach for DN kidney repair.

  13. Spliced stromal cell-derived factor-1α analog stimulates endothelial progenitor cell migration and improves cardiac function in a dose-dependent manner after myocardial infarction

    PubMed Central

    Hiesinger, William; Frederick, John R.; Atluri, Pavan; McCormick, Ryan C.; Marotta, Nicole; Muenzer, Jeffrey R.; Woo, Y. Joseph

    2011-01-01

    Objectives Stromal cell-derived factor (SDF)-1α is a potent endogenous endothelial progenitor cell (EPC) chemokine and key angiogenic precursor. Recombinant SDF-1α has been demonstrated to improve neovasculogenesis and cardiac function after myocardial infarction (MI) but SDF-1α is a bulky protein with a short half-life. Small peptide analogs might provide translational advantages, including ease of synthesis, low manufacturing costs, and the potential to control delivery within tissues using engineered biomaterials. We hypothesized that a minimized peptide analog of SDF-1α, designed by splicing the N-terminus (activation and binding) and C-terminus (extracellular stabilization) with a truncated amino acid linker, would induce EPC migration and preserve ventricular function after MI. Methods EPC migration was first determined in vitro using a Boyden chamber assay. For in vivo analysis, male rats (n=48) underwent left anterior descending coronary artery ligation. At infarction, the rats were randomized into 4 groups and received peri-infarct intramyocardial injections of saline, 3 μg/kg of SDF-1α, 3 μg/kg of spliced SDF analog, or 6 μg/kg spliced SDF analog. After 4 weeks, the rats underwent closed chest pressure volume conductance catheter analysis. Results EPCs showed significantly increased migration when placed in both a recombinant SDF-1α and spliced SDF analog gradient. The rats treated with spliced SDF analog at MI demonstrated a significant dose-dependent improvement in end-diastolic pressure, stroke volume, ejection fraction, cardiac output, and stroke work compared with the control rats. Conclusions A spliced peptide analog of SDF-1α containing both the N- and C- termini of the native protein induced EPC migration, improved ventricular function after acute MI, and provided translational advantages compared with recombinant human SDF-1α. PMID:20951261

  14. Spliced stromal cell-derived factor-1α analog stimulates endothelial progenitor cell migration and improves cardiac function in a dose-dependent manner after myocardial infarction.

    PubMed

    Hiesinger, William; Frederick, John R; Atluri, Pavan; McCormick, Ryan C; Marotta, Nicole; Muenzer, Jeffrey R; Woo, Y Joseph

    2010-11-01

    Stromal cell-derived factor (SDF)-1α is a potent endogenous endothelial progenitor cell (EPC) chemokine and key angiogenic precursor. Recombinant SDF-1α has been demonstrated to improve neovasculogenesis and cardiac function after myocardial infarction (MI) but SDF-1α is a bulky protein with a short half-life. Small peptide analogs might provide translational advantages, including ease of synthesis, low manufacturing costs, and the potential to control delivery within tissues using engineered biomaterials. We hypothesized that a minimized peptide analog of SDF-1α, designed by splicing the N-terminus (activation and binding) and C-terminus (extracellular stabilization) with a truncated amino acid linker, would induce EPC migration and preserve ventricular function after MI. EPC migration was first determined in vitro using a Boyden chamber assay. For in vivo analysis, male rats (n = 48) underwent left anterior descending coronary artery ligation. At infarction, the rats were randomized into 4 groups and received peri-infarct intramyocardial injections of saline, 3 μg/kg of SDF-1α, 3 μg/kg of spliced SDF analog, or 6 μg/kg spliced SDF analog. After 4 weeks, the rats underwent closed chest pressure volume conductance catheter analysis. EPCs showed significantly increased migration when placed in both a recombinant SDF-1α and spliced SDF analog gradient. The rats treated with spliced SDF analog at MI demonstrated a significant dose-dependent improvement in end-diastolic pressure, stroke volume, ejection fraction, cardiac output, and stroke work compared with the control rats. A spliced peptide analog of SDF-1α containing both the N- and C- termini of the native protein induced EPC migration, improved ventricular function after acute MI, and provided translational advantages compared with recombinant human SDF-1α. Copyright © 2010 The American Association for Thoracic Surgery. Published by Mosby, Inc. All rights reserved.

  15. Resveratrol as a natural anti-tumor necrosis factor-α molecule: implications to dendritic cells and their crosstalk with mesenchymal stromal cells.

    PubMed

    Silva, Andreia M; Oliveira, Marta I; Sette, Laura; Almeida, Catarina R; Oliveira, Maria J; Barbosa, Mário A; Santos, Susana G

    2014-01-01

    Dendritic cells (DC) are promising targets for inducing tolerance in inflammatory conditions. Thus, this study aims to investigate the effects of the natural anti-inflammatory molecule resveratrol on human DC at phenotypic and functional levels, including their capacity to recruit mesenchymal stem/stromal cells (MSC). Primary human monocyte-derived DC and bone marrow MSC were used. DC immunophenotyping revealed that small doses of resveratrol (10 µM) reduce cell activation in response to tumor necrosis factor (TNF)-α, significantly decreasing surface expression of CD83 and CD86. Functionally, IL-12/IL-23 secretion induced by TNF-α was significantly reduced by resveratrol, while IL-10 levels increased. Resveratrol also inhibited T cell proliferation, in response to TNF-α-stimulated DC. The underlying mechanism was investigated by Western blot and imaging flow cytometry (ImageStreamX), and likely involves impairment of nuclear translocation of the p65 NF-κB subunit. Importantly, results obtained demonstrate that DC are able to recruit MSC through extracellular matrix components, and that TNF-α impairs DC-mediated recruitment. Matrix metalloproteinases (MMP) produced by both cell populations were visualized by gelatin zymography. Finally, time-lapse microscopy analysis revealed a significant decrease on DC and MSC motility in co-cultures, indicating cell interaction, and TNF-α further decreased MSC motility, while resveratrol recovered it. Thus, the current study points out the potential of resveratrol as a natural anti-TNF-α drug, capable of modulating DC phenotype and function, as well as DC-mediated MSC recruitment.

  16. The involvement of anterior gradient 2 in the stromal cell-derived factor 1-induced epithelial-mesenchymal transition of glioblastoma.

    PubMed

    Xu, Chunhua; Liu, Yue; Xiao, Limin; Guo, Changgui; Deng, Shengze; Zheng, Suyue; Zeng, Erming

    2016-05-01

    In recent years, it has been widely identified that the stromal cell-derived factor 1 (SDF-1) and anterior gradient 2 (AGR2) were implicated in the development of epithelial-mesenchymal transition (EMT) in a variety of cancers. However, the involvement of SDF-1-AGR2 pathway in the EMT of glioblastoma has not been investigated. In the present study, the in vitro assays were used to investigate the role of AGR2 in cell cycle, migration, and invasion. We found that the expressions of AGR2 and chemokine (C-X-C motif) receptor 4 (CXCR4) were obviously upregulated in glioblastoma cells T98G, A172, U87, and U251 than those in normal human astrocytes (NHA) (all p < 0.01), among which both U87 and U251 cells presented the highest expression (p > 0.05). Western blot revealed that SDF-1 induced the expression of p-AKT, AGR2, and EMT markers (N-cadherin, matrix metalloproteinase-2 (MMP2), and Slug) in a dose-dependent manner in U87 and U251 cells. However, the depletion of AGR2 reversed SDF-1-induced upregulation of EMT markers rather than p-AKT. Furthermore, functional analysis identified that knockdown of AGR2 induced cell cycle arrest in G0/G1 phase and suppressed the migration and invasion of U87 and U251 cells. Taken together, SDF-1-CXCR4 pathway induced the expression of AGR2 to control the progression of EMT likely via AKT pathway in the development of glioblastoma. Our findings lay a promising foundation for the SDF-1-AGR2 axis-targeting therapy in patients with glioblastoma.

  17. Sonic hedgehog signals to multiple prostate stromal stem cells that replenish distinct stromal subtypes during regeneration

    PubMed Central

    Peng, Yu-Ching; Levine, Charles M.; Zahid, Sarwar; Wilson, E. Lynette; Joyner, Alexandra L.

    2013-01-01

    The adult mouse prostate has a seemingly endless capacity for regeneration, and sonic hedgehog (SHH) signaling has been implicated in this stem cell-driven process. However, it is not clear whether SHH acts on the epithelium or stromal cells that secrete factors required for epithelial expansion. Because little is known about stromal stem cells compared with their epithelial counterparts, we used in vivo mouse genetics tools to characterize four prostate stromal subtypes and their stem cells. Using knockin reporter alleles, we uncovered that SHH signals from prostate basal epithelial cells to adjacent stromal cells. Furthermore, the SHH target gene Gli1 is preferentially expressed in subepithelial fibroblast-like cells, one of four prostate stromal subtypes and the subtype closest to the epithelial source of SHH. Using Genetic Inducible Fate Mapping to mark adult Gli1- or Smooth muscle actin-expressing cells and follow their fate during regeneration, we uncovered that Gli1-expressing cells exhibit long-term self-renewal capacity during multiple rounds of androgen-mediated regeneration after castration-induced involution, and depleted smooth muscle cells are mainly replenished by preexisting smooth muscle cells. Based on our Genetic Inducible Fate Mapping studies, we propose a model where SHH signals to multiple stromal stem cells, which are largely unipotent in vivo. PMID:24218555

  18. Schwann cells induce neuronal differentiation of bone marrow stromal cells.

    PubMed

    Zurita, Mercedes; Vaquero, Jesús; Oya, Santiago; Miguel, Miriam

    2005-04-04

    Bone marrow stromal cells are multipotent stem cells that have the potential to differentiate into bone, cartilage, fat and muscle. Recently, bone marrow stromal cells have been shown to have the capacity to differentiate into neurons under specific experimental conditions, using chemical factors. We now describe how bone marrow stromal cells can be induced to differentiate into neuron-like cells when they are co-cultured with Schwann cells. When compared with chemical differentiation, expression of neuronal differentiation markers begins later, but one week after beginning co-culture, most bone marrow stromal cells showed a typical neuronal morphology. Our present findings support the transdifferentiation of bone marrow stromal cells, and the potential utility of these cells for the treatment of degenerative and acquired disorders of the nervous system.

  19. Effects of lipid-related factors on adipocyte differentiation of bovine stromal-vascular cells in primary culture.

    PubMed

    Wu, P; Sato, K; Suzuta, F; Hikasa, Y; Kagota, K

    2000-09-01

    The effects of several factors related to lipids on bovine adipocyte differentiation were investigated in primary culture. Adipocyte differentiation was assessed by development of glycerol-3-phosphate dehydrogenase (GPDH) activity and morphological observation. Addition of triglyceride mixture (Intralipid), caprylic acid and very low-, low- and high-density lipoproteins (VLDL, LDL and HDL) stimulated bovine preadipocyte differentiation in serum-free condition. Especially, VLDL strongly increased both cell protein contents and GPDH activity, suggesting that it stimulated both proliferation and differentiation of bovine preadipocytes. Under Intralipid-induced condition, differentiation of preadipocytes from subcutaneous adipose tissues was more evident than those from omental adipose tissues. However, such depot difference was not observed in medium supplemented with indomethacin, which is a peroxisome proliferator-activated receptor (PPAR) gamma agonist. This suggests that the differentiation capacity of bovine preadipocytes was different between depots and such difference is dependent on the ability to utilize lipids as endogenous PPARgamma ligands. Therefore, lipid metabolites have the stimulatory effects on bovine adipocyte differentiation in vitro, and lipoproteins, especially VLDL, may play an important role in development of bovine adipose tissues in vivo.

  20. A DC-81-indole conjugate agent suppresses melanoma A375 cell migration partially via interrupting VEGF production and stromal cell-derived factor-1{alpha}-mediated signaling

    SciTech Connect

    Hsieh, Ming-Chu; Hu, Wan-Ping; Yu, Hsin-Su; Wu, Wen-Chuan; Chang, Long-Sen; Kao, Ying-Hsien; Wang, Jeh-Jeng

    2011-09-01

    Pyrrolo[2,1-c][1,4]benzodiazepine (PBD) chemicals are antitumor antibiotics inhibiting nucleic acid synthesis. An indole carboxylate-PBD hybrid with six-carbon spacer structure (IN6CPBD) has been previously demonstrated to induce melanoma cell apoptosis and reduce metastasis in mouse lungs. This study aimed at investigating the efficacy of the other hybrid compound with four-carbon spacer (IN4CPBD) and elucidating its anti-metastatic mechanism. Human melanoma A375 cells with IN4CPBD treatment underwent cytotoxicity and apoptosis-associated assays. Transwell migration assay, Western blotting, and ELISA were used for mechanistic study. IN4CPBD exhibited potent melanoma cytotoxicity through interrupting G1/S cell cycle progression, increasing DNA fragmentation and hypodipoidic DNA contents, and reducing mitochondrial membrane potential. Caspase activity elevation suggested that both intrinsic and extrinsic pathways were involved in IN4CPBD-induced melanoma apoptosis. IN4CPBD up-regulated p53 and p21, thereby concomitantly derailing the equilibrium between Bcl-2 and Bax levels. Transwell migration assay demonstrated that stromal cell-derived factor-1{alpha} (SDF-1{alpha}) stimulated A375 cell motility, while kinase inhibitors treatment confirmed that Rho/ROCK, Akt, ERK1/2, and p38 MAPK pathways were involved in SDF-1{alpha}-enhanced melanoma migration. IN4CPBD not only abolished the SDF-1{alpha}-enhanced chemotactic motility but also suppressed constitutive MMP-9 and VEGF expression. Mechanistically, IN4CPBD down-regulated Akt, ERK1/2, and p38 MAPK total proteins and MYPT1 phosphorylation. In conclusion, beyond the fact that IN4CPBD induces melanoma cell apoptosis at cytotoxic dose, the interruption in the VEGF expression and the SDF-1{alpha}-related signaling at cytostatic dose may partially constitute the rationale for its in vivo anti-metastatic potency. - Research Highlights: > A novel carboxylate-PBD hybrid as anti-melanoma drug. > IN4CPBD interrupts melanoma cell

  1. Noncontiguous domains of the alpha-factor receptor of yeasts confer ligand specificity.

    PubMed

    Sen, M; Marsh, L

    1994-01-14

    The Saccharomyces cerevisiae alpha-factor receptor has a 3400-fold higher affinity for the S. cerevisiae alpha-factor peptide (c-alpha-f) than for the Saccharomyces kluyveri alpha-factor peptide (k-alpha-f) as determined by competition for [3H] c-alpha-f binding. The S. kluyveri alpha-factor receptor has an approximately 2-fold higher affinity for k-alpha-f than for c-alpha-f. The S. kluyveri receptor gene (k-STE2) is incompletely regulated by S. cerevisiae mating type and poorly expressed on the surface of an S. cerevisiae mating type a strain. A chimeric receptor (c/k1) with amino acid residues 1-45 derived from S. cerevisiae and amino acid residues 46-427 from S. kluyveri exhibits the binding specificity of the S. kluyveri receptor. However, chimeric receptors containing residues 1-168 (c/k2) or 1-250 (c/k3) from S. cerevisiae and the remainder from the S. kluyveri receptor exhibit specificities similar to one another, but intermediate between the parent S. cerevisiae and S. kluyveri receptors. The relative ability of c-alpha-f and k-alpha-f to induce growth arrest in strains expressing chimeric receptors parallels relative affinity. Thus, two noncontiguous domains that include putative extracellular loops 1 and 3 and associated transmembrane segments, but exclude the extracellular NH2 terminus and loop 2, appear to contribute to alpha-factor receptor ligand specificity. COOH-terminal regions of the S. kluyveri receptor appear to confer a desensitization defect when expressed in S. cerevisiae. The S. cerevisiae receptor truncated at residue 296 retains ligand specificity for growth arrest.

  2. Regulation of growth by a nerve growth factor-like protein which modulates paracrine interactions between a neoplastic epithelial cell line and stromal cells of the human prostate.

    PubMed

    Djakiew, D; Delsite, R; Pflug, B; Wrathall, J; Lynch, J H; Onoda, M

    1991-06-15

    Nerve growth factor-like substance(s) were identified in both conditioned media of a human prostatic tumor epithelial cell line (TSU-pr1) and a human prostatic stromal cell line (HPS) by Western blot analysis and bioassay of neurite outgrowth of PC12 cells. Nerve growth factor-beta (NGF) immunofluorescence was also localized to secretory vesicles in the cytoplasm of both the TSU-pr1 and HPS cells. Western blot of the TSU-pr1 and HPS cell-secreted protein identified an Mr 65,000 major protein which immunoreacted with murine NGF antibody. NGF Western blot of HPS cell-secreted protein also identified an Mr 42,000 minor band under reduced and nonreduced conditions and an Mr 61,000 minor band under reduced conditions. The secreted protein from the TSU-pr1 cells (50 micrograms/ml) and HPS (50 micrograms/ml), as well as murine NGF (50 ng/ml) or human recombinant NGF (50 ng/ml), stimulated neurite outgrowth from PC12 cells. This neurite outgrowth activity was partially inhibited by treatment with NGF antibody. Neither the serum containing growth medium nor bovine serum albumin (50 micrograms/ml) stimulated neurite outgrowth. The NGF-like secretory protein appeared to play a role in the paracrine regulation of prostatic growth between TSU-pr1 cells and HPS cells. The relative growth of TSU-pr1 cells, as indicated by [3H]thymidine incorporation, in response to HPS secretory protein was stimulated 2.8-fold in a dose-dependent manner. In the converse interaction, the relative growth of HPS cells in response to TSU-pr1 secretory protein was stimulated 1.8-fold in a dose-dependent manner. Immunoneutralization of TSU-pr1 and HPS secretory protein was performed with antibody against NGF, acidic fibroblast growth factor, and basic fibroblast growth factor. Removal of the NGF-like protein from the maximal stimulatory dose of TSU-pr1 secretory protein (100 micrograms/ml) with NGF antibody reduced HPS proliferation to 52% of maximal levels, and immunoneutralization of the NGF

  3. Nerve regeneration by human corneal stromal keratocytes and stromal fibroblasts

    PubMed Central

    Yam, Gary Hin-Fai; Williams, Geraint P.; Setiawan, Melina; Yusoff, Nur Zahirah Binte M.; Lee, Xiao-wen; Htoon, Hla Myint; Zhou, Lei; Fuest, Matthias; Mehta, Jodhbir S.

    2017-01-01

    Laser refractive surgeries reshape corneal stroma to correct refractive errors, but unavoidably affect corneal nerves. Slow nerve regeneration and atypical neurite morphology cause desensitization and neuro-epitheliopathy. Following injury, surviving corneal stromal keratocytes (CSKs) are activated to stromal fibroblasts (SFs). How these two different cell types influence nerve regeneration is elusive. Our study evaluated the neuro-regulatory effects of human SFs versus CSKs derived from the same corneal stroma using an in vitro chick dorsal root ganglion model. The neurite growth was assessed by a validated concentric circle intersection count method. Serum-free conditioned media (CM) from SFs promoted neurite growth dose-dependently, compared to that from CSKs. We detected neurotrophic and pro-inflammatory factors (interleukin-8, interleukin-15, monocyte chemoattractant protein-1, eotaxin, RANTES) in SFCM by Bio-Plex Human Cytokine assay. More than 130 proteins in SFCM and 49 in CSKCM were identified by nanoLC-MS/MS. Proteins uniquely present in SFCM had reported neuro-regulatory activities and were predicted to regulate neurogenesis, focal adhesion and wound healing. Conclusively, this was the first study showing a physiological relationship between nerve growth and the metabolically active SFs versus quiescent CSKs from the same cornea source. The dose-dependent effect on neurite growth indicated that nerve regeneration could be influenced by SF density. PMID:28349952

  4. Neurogenic differentiation factor NeuroD confers protection against radiation-induced intestinal injury in mice

    PubMed Central

    Li, Ming; Du, Aonan; Xu, Jing; Ma, Yanchao; Cao, Han; Yang, Chao; Yang, Xiao-Dong; Xing, Chun-Gen; Chen, Ming; Zhu, Wei; Zhang, Shuyu; Cao, Jianping

    2016-01-01

    The gastrointestinal tract, especially the small intestine, is particularly sensitive to radiation, and is prone to radiation-induced injury as a result. Neurogenic differentiation factor (NeuroD) is an evolutionarily-conserved basic helix-loop-helix (bHLH) transcription factor. NeuroD contains a protein transduction domain (PTD), which allows it to be exogenously delivered across the membrane of mammalian cells, whereupon its transcription activity can be unleashed. Whether NeuroD has therapeutic effects for radiation-induced injury remains unclear. In the present study, we prepared a NeuroD-EGFP recombinant protein, and explored its protective effects on the survival and intestinal damage induced by ionizing radiation. Our results showed that NeuroD-EGFP could be transduced into small intestine epithelial cells and tissues. NeuroD-EGFP administration significantly increased overall survival of mice exposed to lethal total body irradiation (TBI). This recombinant NeuroD also reduced radiation-induced intestinal mucosal injury and apoptosis, and improved crypt survival. Expression profiling of NeuroD-EGFP-treated mice revealed upregulation of tissue inhibitor of metalloproteinase 1 (TIMP-1), a known inhibitor of apoptosis in mammalian cells. In conclusion, NeuroD confers protection against radiation-induced intestinal injury, and provides a novel therapeutic clinical option for the prevention of intestinal side effects of radiotherapy and the treatment of victims of incidental exposure. PMID:27436572

  5. Co-delivery and controlled release of stromal cell-derived factor-1α chemically conjugated on collagen scaffolds enhances bone morphogenetic protein-2-driven osteogenesis in rats

    PubMed Central

    SUN, HAIPENG; WANG, JINMING; DENG, FEILONG; LIU, YUN; ZHUANG, XIUMEI; XU, JIAYUN; LI, LONG

    2016-01-01

    There has been considerable focus in investigations on the delivery systems and clinical applications of bone morphogenetic protein-2 (BMP-2) for novel bone formation. However, current delivery systems require high levels of BMP-2 to exert a biological function. There are several concerns in using of high levels of BMP-2, including safety and the high cost of treatment. Therefore, the development of strategies to decrease the levels of BMP-2 required in these delivery systems is required. In our previous studies, a controlled-release system was developed, which used Traut's reagent and the cross-linker, 4-(N-maleimi-domethyl) cyclohexane-1-carboxylic acid 3-sulfo-N-hydroxysuccinimide ester sodium salt (Sulfo-SMCC), to chemically conjugate BMP-2 directly on collagen discs. In the current study, retention efficiency and release kinetics of stromal cell-derived factor-1α (SDF-1α) cross-linked on collagen scaffolds were detected. In addition, the osteogenic activity of SDF-1α and suboptimal doses of BMP-2 cross-linked on collagen discs following subcutaneous implantation in rats were evaluated. Independent two-tailed t-tests and one-way analysis of variance were used for analysis. In the present study, the controlled release of SDF-1α chemically conjugated on collagen scaffolds was demonstrated. By optimizing the concentrations of Traut's reagent and the Sulfo-SMCC cross-linker, a significantly higher level of SDF-1α was covalently retained on the collagen scaffold, compared with that retained using a physical adsorption method. Mesenchymal stem cell homing indicated that the biological function of the SDF-1α cross-linked on the collagen scaffolds remained intact. In rats, co-treatment with SDF-1α and a suboptimal dose of BMP-2 cross-linked on collagen scaffolds using this chemically conjugated method induced higher levels of ectopic bone formation, compared with the physical adsorption method. No ectopic bone formation was observed following treatment with a

  6. β-Arrestin1 and Distinct CXCR4 Structures Are Required for Stromal Derived Factor-1 to Downregulate CXCR4 Cell-Surface Levels in Neuroblastoma

    PubMed Central

    Clift, Ian C.; Bamidele, Adebowale O.; Rodriguez-Ramirez, Christie; Kremer, Kimberly N.

    2014-01-01

    CXC chemokine receptor 4 (CXCR4) is a G protein–coupled receptor (GPCR) located on the cell surface that signals upon binding the chemokine stromal derived factor-1 (SDF-1; also called CXCL 12). CXCR4 promotes neuroblastoma proliferation and chemotaxis. CXCR4 expression negatively correlates with prognosis and drives neuroblastoma growth and metastasis in mouse models. All functions of CXCR4 require its expression on the cell surface, yet the molecular mechanisms that regulate CXCR4 cell-surface levels in neuroblastoma are poorly understood. We characterized CXCR4 cell-surface regulation in the related SH-SY5Y and SK-N-SH human neuroblastoma cell lines. SDF-1 treatment caused rapid down-modulation of CXCR4 in SH-SY5Y cells. Pharmacologic activation of protein kinase C similarly reduced CXCR4, but via a distinct mechanism. Analysis of CXCR4 mutants delineated two CXCR4 regions required for SDF-1 treatment to decrease cell-surface CXCR4 in neuroblastoma cells: the isoleucine-leucine motif at residues 328 and 329 and residues 343–352. In contrast, and unlike CXCR4 regulation in other cell types, serines 324, 325, 338, and 339 were not required. Arrestin proteins can bind and regulate GPCR cell-surface expression, often functioning together with kinases such as G protein–coupled receptor kinase 2 (GRK2). Using SK-N-SH cells which are naturally deficient in β-arrestin1, we showed that β-arrestin1 is required for the CXCR4 343–352 region to modulate CXCR4 cell-surface expression following treatment with SDF-1. Moreover, GRK2 overexpression enhanced CXCR4 internalization, via a mechanism requiring both β-arrestin1 expression and the 343–352 region. Together, these results characterize CXCR4 structural domains and β-arrestin1 as critical regulators of CXCR4 cell-surface expression in neuroblastoma. β-Arrestin1 levels may therefore influence the CXCR4-driven metastasis of neuroblastoma as well as prognosis. PMID:24452472

  7. Modulation of Stromal Cell-Derived Factor-1/CXC Chemokine Receptor 4 Axis Enhances rhBMP-2-Induced Ectopic Bone Formation

    PubMed Central

    Wise, Joel K.; Sumner, Dale Rick

    2012-01-01

    Enhancement of in vivo mobilization and homing of endogenous mesenchymal stem cells (MSCs) to an injury site is an innovative strategy for improvement of bone tissue engineering and repair. The present study was designed to determine whether mobilization by AMD3100 and/or local homing by delivery of stromal cell-derived factor-1 (SDF-1) enhances recombinant human bone morphogenetic protein-2 (rhBMP-2) induced ectopic bone formation in an established rat model. Rats received an injection of either saline or AMD3100 treatment 1 h before harvesting of bone marrow for in vitro colony-forming unit-fibroblasts (CFU-F) culture or the in vivo subcutaneous implantation of absorbable collagen sponges (ACSs) loaded with saline, recombinant human bone morphogenetic protein-2 (rhBMP-2), SDF-1, or the combination of SDF-1 and rhBMP-2. AMD3100 treatment resulted in a significant decrease in CFU-F number, compared with saline, which confirmed that a single systemic AMD3100 treatment rapidly mobilized MSCs from the bone marrow. At 28 and 56 days, bone formation in the explanted ACS was assessed by microcomputed tomography (μCT) and histology. At 28 days, AMD3100 and/or SDF-1 had no statistically significant effect on bone volume (BV) or bone mineral content (BMC), but histology revealed more active bone formation with treatment of AMD3100, loading of SDF-1, or the combination of both AMD3100 and SDF-1, compared with saline-treated rhBMP-2 loaded ACS. At 56 days, the addition of AMD3100 treatment, loading of SDF-1, or the combination of both resulted in a statistically significant stimulatory effect on BV and BMC, compared with the saline-treated rhBMP-2 loaded ACS. Histology of the 56-day ACS were consistent with the μCT analysis, exhibiting more mature and mineralized bone formation with AMD3100 treatment, SDF-1 loading, or the combination of both, compared with the saline-treated rhBMP-2 loaded ACS. The present study is the first that provides evidence of the efficacy of AMD

  8. Expression of platelet-bound stromal-cell derived factor-1 (SDF-1) and number of CD34(+) progenitor cells in patients with congestive heart failure.

    PubMed

    Jorbenadze, Rezo; Schleicher, Erwin; Bigalke, Boris; Stellos, Konstantinos; Gawaz, Meinrad

    2014-01-01

    Platelet-bound stromal cell-derived factor-1 (SDF-1) plays a crucial role in attachment of circulating CD34(+) progenitor cells to the vascular wall, facilitating tissue healing after injury. However there is no evidence about expression of platelet-bound SDF-1 in patients with congestive heart failure (CHF). The aim of our study was to evaluate expression of platelet-bound SDF-1 and number of CD34(+) progenitor cells in patients with CHF. Forty-eight patients with idiopathic dilated cardiomyopathy (DCM) and 61 patients with ischaemic cardiomyopathy (ICM) were consecutively enrolled into the study. Blood taken from 109 consecutive patients was studied for surface expression of platelet-bound SDF-1 and number of CD34(+) progenitor cells by flow cytometry. The highest expression of platelet-bound SDF-1 was observed in patients with severe impairment of left ventricular systolic function compared with patients with mild or moderate impairment of left ventricular systolic function (mild vs. moderate vs. severe impairment of left ventricular systolic function: MFI ± SD: 35.6 ± 34 vs. 101.45 ± 73 vs. 124.86 ± 86.7, Kruskal-Wallis p < 0.001). Similar to platelet-bound SDF-1 number of CD34(+) progenitor cells was the highest in severe impairment of left ventricular systolic function (mild vs. moderate vs. severe impairment of left ventricular systolic function: mean ± SD: 260.4 ± 177.5 vs. 580.7 ± 340.5 vs. 640.82 ± 370.6, Kruskal-Wallis p < 0.001). Platelet-bound SDF-1 expression was associated with number of circulating CD34(+) progenitor cells (r = 0.454, p < 0.001) in patients with CHF. Expression of platelet-bound SDF-1 and number of CD34(+) cells were higher in patients with DCM compared with patients with ICM (p < 0.001 for both) and inversely correlated with age and aspirin therapy. Platelet-bound SDF-1 and CD34(+) progenitor cells are especially increased in patients with severe impairment of left

  9. Repair of mandibular defects by bone marrow stromal cells expressing the basic fibroblast growth factor transgene combined with multi-pore mineralized Bio-Oss.

    PubMed

    Yang, Chunyan; Liu, Yang; Li, Chunming; Zhang, Bin

    2013-01-01

    The aim of the present study was to evaluate the effect of combining Bio‑Oss with bone marrow stromal cells (BMSCs) transfected with the basic fibroblast growth factor (bFGF) gene on bone regeneration during mandibular distraction of rabbits. BMSCs obtained from rabbits were transfected with bFGF gene‑encoding plasmids and proliferation rate and the differentiation marker alkaline phosphatase activity were measured. Following seeding into Bio‑Oss collagen and 9‑day culture in vitro, the surface morphology of the Bio‑Oss was assessed using scanning electron microscopy analysis. Three mandibular defects were induced in the lower border of the bilateral mandibular ramus in each New Zealand white rabbit (total n=6). Three scaffolds, group A (seeded with BMSCs/bFGF), B (seeded with BMSCs/pVAX1) and C (cell‑free), which had been cultured in vitro under standard cell culture conditions for 18 days, were implanted into mandibular defects under sterile conditions. Animals were sacrificed by anesthesia overdose 12 weeks following surgery and the scaffolds were extracted for bone mineral density and histological analyses. Results indicate that bFGF was successfully transfected into BMSCs. Proliferation and osteoblast differentiation of BMSCs were stimulated by bFGF in vitro. No differences were identified in surface morphology for Bio‑Oss loaded with variable groups of cells. At week 12 following implantation of Bio‑Oss scaffolds, mineralization of BMSCs in Bio‑Oss scaffolds was observed to be increased by bFGF. New bone and cartilage formation was revealed in hematoxylin and eosin‑stained sections in Bio‑Oss scaffolds and was most abundant in group A (BMSCs transfected with bFGF). In the current study, the bFGF gene was transfected into BMSCs and expressed successfully. bFGF promoted proliferation and differentiation of BMSCs in vitro and implantation of bFGF‑expressing BMSCs combined with Bio‑Oss enhanced new bone regeneration more

  10. Effect of basic fibroblast growth factor on the growth and differentiation of adult stromal bone marrow cells: enhanced development of mineralized bone-like tissue in culture.

    PubMed

    Pitaru, S; Kotev-Emeth, S; Noff, D; Kaffuler, S; Savion, N

    1993-08-01

    Rat stromal bone marrow cells (SBMC) were shown to produce mineralized bone-like tissue in culture in the presence of dexamethasone, ascorbic acid, and beta-glycerophosphate. The addition of 3 ng/ml of basic fibroblast growth factor (bFGF) resulted in a significant increase in formation of mineralized tissue. The present study was aimed at assessing the effect of bFGF on the proliferation and differentiation of SBMC and on the sequential development of mineralized bone-like tissue in culture. Transmission electron microscopy of bFGF-treated cultures demonstrated the development of a multilayered structure resembling mineralized bone tissue consisting of cell layers embedded within a heavy extracellular matrix. The matrix was rich in bundles of collagen fibers associated with extensive mineral deposits consisting of hydroxyapatite as determined by infrared spectrophotometry. The addition of 3 ng/ml of bFGF resulted in significant enhancement of [3H]thymidine and [3H]proline incorporation and protein accumulation by 12-, 2.5-, and 2.5-fold, respectively. bFGF treatment increased cAMP responsiveness, alkaline phosphatase activity, osteocalcin level, 45Ca2+ deposition, and mineralized-like tissue formation and induced the earlier expression of these markers in the treated culture. A biphasic sequence of events was observed during the development of mineralized bone-like tissue in bFGF-treated and control cultures. The first phase is characterized by cell proliferation and matrix accumulation and is reflected by a progressive increase in [3H]thymidine and [3H]proline incorporation until day 11. The second phase, which follows, is characterized by a sharp decline in cell proliferation and matrix accumulation and a concomitant expression of osteoblast differentiation as reflected by the progressive increase in alkaline phosphatase activity, mineral deposition, and osteocalcin expression. Treatment of cultures with bFGF accentuated this biphasic sequence of events. These

  11. Proceedings of the international conference on nuclear power plant aging, availability factor and reliability analysis

    SciTech Connect

    Goel, V.S.

    1985-01-01

    This book presents the papers given at a conference on nuclear power plant life extension. Topics considered at the conference included availability, accelerated aging techniques, the qualification of electrical equipment, probabilistic risk assessment, reactor maintenance, outages, reliability, computer-aided design, seismic effects, mechanical vibrations, fatigue monitoring, steam generators, and materials degradation by aging and embrittlement.

  12. Fibroblast Growth Factor Receptor-2 Contributes to the Basic Fibroblast Growth Factor-Induced Neuronal Differentiation in Canine Bone Marrow Stromal Cells via Phosphoinositide 3-Kinase/Akt Signaling Pathway

    PubMed Central

    Nakano, Rei; Edamura, Kazuya; Nakayama, Tomohiro; Narita, Takanori; Okabayashi, Ken; Sugiya, Hiroshi

    2015-01-01

    Bone marrow stromal cells (BMSCs) are considered as candidates for regenerative therapy and a useful model for studying neuronal differentiation. The role of basic fibroblast growth factor (bFGF) in neuronal differentiation has been previously studied; however, the signaling pathway involved in this process remains poorly understood. In this study, we investigated the signaling pathway in the bFGF-induced neuronal differentiation of canine BMSCs. bFGF induced the mRNA expression of the neuron marker, microtubule associated protein-2 (MAP2) and the neuron-like morphological change in canine BMSCs. In the presence of inhibitors of fibroblast growth factor receptors (FGFR), phosphatidylinositol 3-kinase (PI3K) and Akt, i.e., SU5402, LY294002, and MK2206, respectively, bFGF failed to induce the MAP2 mRNA expression and the neuron-like morphological change. bFGF induced Akt phosphorylation, but it was attenuated by the FGFR inhibitor SU5402 and the PI3K inhibitor LY294002. In canine BMSCs, expression of FGFR-1 and FGFR-2 was confirmed, but only FGFR-2 activation was detected by cross-linking and immunoprecipitation analysis. Small interfering RNA-mediated knockdown of FGFR-2 in canine BMSCs resulted in the attenuation of bFGF-induced Akt phosphorylation. These results suggest that the FGFR-2/PI3K/Akt signaling pathway is involved in the bFGF-induced neuronal differentiation of canine BMSCs. PMID:26523832

  13. Hsp33 confers bleach resistance by protecting elongation factor Tu against oxidative degradation in Vibrio cholerae

    PubMed Central

    Wholey, Wei-Yun; Jakob, Ursula

    2012-01-01

    Summary The redox-regulated chaperone Hsp33 protects bacteria specifically against stress conditions that cause oxidative protein unfolding, such as treatment with bleach or exposure to peroxide at elevated temperatures. To gain insight into the mechanism by which expression of Hsp33 confers resistance to oxidative protein unfolding conditions, we made use of V. cholerae strain O395 lacking the Hsp33 gene hslO. We found that this strain, which is exquisitely bleach-sensitive, displays a temperature-sensitive (ts) phenotype during aerobic growth, implying that V. cholerae suffers from oxidative heat stress when cultivated at 43°C. We utilized this phenotype to select for E. coli genes that rescue the ts phenotype of V. cholerae ΔhslO when overexpressed. We discovered that expression of a single protein, the elongation factor EF-Tu, was sufficient to rescue both the ts and bleach-sensitive phenotypes of V. cholerae ΔhslO. In vivo studies revealed that V. cholerae EF-Tu is highly sensitive to oxidative protein degradation in the absence of Hsp33, indicating that EF-Tu is a vital chaperone substrate of Hsp33 in V. cholerae. These results suggest an “essential client protein” model for Hsp33’s chaperone action in Vibrio in which stabilization of a single oxidative stress-sensitive protein is sufficient to enhance the oxidative stress resistance of the whole organism. PMID:22296329

  14. Hsp33 confers bleach resistance by protecting elongation factor Tu against oxidative degradation in Vibrio cholerae.

    PubMed

    Wholey, Wei-Yun; Jakob, Ursula

    2012-03-01

    The redox-regulated chaperone Hsp33 protects bacteria specifically against stress conditions that cause oxidative protein unfolding, such as treatment with bleach or exposure to peroxide at elevated temperatures. To gain insight into the mechanism by which expression of Hsp33 confers resistance to oxidative protein unfolding conditions, we made use of Vibrio cholerae strain O395 lacking the Hsp33 gene hslO. We found that this strain, which is exquisitely bleach-sensitive, displays a temperature-sensitive (ts) phenotype during aerobic growth, implying that V. cholerae suffers from oxidative heat stress when cultivated at 43°C. We utilized this phenotype to select for Escherichia coli genes that rescue the ts phenotype of V. cholerae ΔhslO when overexpressed. We discovered that expression of a single protein, the elongation factor EF-Tu, was sufficient to rescue both the ts and bleach-sensitive phenotypes of V. cholerae ΔhslO. In vivo studies revealed that V. cholerae EF-Tu is highly sensitive to oxidative protein degradation in the absence of Hsp33, indicating that EF-Tu is a vital chaperone substrate of Hsp33 in V. cholerae. These results suggest an 'essential client protein' model for Hsp33's chaperone action in Vibrio in which stabilization of a single oxidative stress-sensitive protein is sufficient to enhance the oxidative stress resistance of the whole organism. © 2012 Blackwell Publishing Ltd.

  15. Parthenocarpic apple fruit production conferred by transposon insertion mutations in a MADS-box transcription factor

    PubMed Central

    Yao, Jia-Long; Dong, Yi-Hu; Morris, Bret A. M.

    2001-01-01

    Fruit development in higher plants normally requires pollination and fertilization to stimulate cell division of specific floral tissues. In some cases, parthenocarpic fruit development proceeds without either pollination or fertilization. Parthenocarpic fruit without seed has higher commercial value than seeded fruit. Several apple (Malus domestica) mutants (Rae Ime, Spencer Seedless and Wellington Bloomless) are known to produce only apetalous flowers that readily go on to develop into parthenocarpic fruit. Through genetics, a single recessive gene has been identified to control this trait in apple. Flower phenotypes of these apple mutants are strikingly similar to those of the Arabidopsis mutant pistillata (pi), which produces flowers where petals are transformed to sepals and stamens to carpels. In this study, we have cloned the apple PI homolog (MdPI) that shows 64% amino acid sequence identity and closely conserved intron positions and mRNA expression patterns to the Arabidopsis PI. We have identified that in the apetalous mutants MdPI has been mutated by a retrotransposon insertion in intron 4 in the case of Rae Ime and in intron 6 in the case of Spencer Seedless and Wellington Bloomless. The insertion apparently abolishes the normal expression of the MdPI gene. We conclude that the loss of function mutation in the MdPI MADS-box transcription factor confers parthenocarpic fruit development in these apple varieties and demonstrates another function for the MADS- box gene family. The knowledge generated here could be used to produce parthenocarpic fruit cultivars through genetic engineering. PMID:11158635

  16. Mutational activation of BRAF confers sensitivity to transforming growth factor beta inhibitors in human cancer cells

    PubMed Central

    Spender, Lindsay C.; Ferguson, G. John; Liu, Sijia; Cui, Chao; Girotti, Maria Romina; Sibbet, Gary; Higgs, Ellen B.; Shuttleworth, Morven K.; Hamilton, Tom; Lorigan, Paul; Weller, Michael; Vincent, David F.; Sansom, Owen J.; Frame, Margaret; Dijke, Peter ten; Marais, Richard; Inman, Gareth J.

    2016-01-01

    Recent data implicate elevated transforming growth factor-β (TGFβ) signalling in BRAF inhibitor drug-resistance mechanisms, but the potential for targeting TGFβ signalling in cases of advanced melanoma has not been investigated. We show that mutant BRAFV600E confers an intrinsic dependence on TGFβ/TGFβ receptor 1 (TGFBR1) signalling for clonogenicity of murine melanocytes. Pharmacological inhibition of the TGFBR1 blocked the clonogenicity of human mutant BRAF melanoma cells through SMAD4-independent inhibition of mitosis, and also inhibited metastasis in xenografted zebrafish. When investigating the therapeutic potential of combining inhibitors of mutant BRAF and TGFBR1, we noted that unexpectedly, low-dose PLX-4720 (a vemurafenib analogue) promoted proliferation of drug-naïve melanoma cells. Pharmacological or pharmacogenetic inhibition of TGFBR1 blocked growth promotion and phosphorylation of SRC, which is frequently associated with vemurafenib-resistance mechanisms. Importantly, vemurafenib-resistant patient derived cells retained sensitivity to TGFBR1 inhibition, suggesting that TGFBR1 could be targeted therapeutically to combat the development of vemurafenib drug-resistance. PMID:27835901

  17. Submergence Confers Immunity Mediated by the WRKY22 Transcription Factor in Arabidopsis[W

    PubMed Central

    Hsu, Fu-Chiun; Chou, Mei-Yi; Chou, Shu-Jen; Li, Ya-Ru; Peng, Hsiao-Ping; Shih, Ming-Che

    2013-01-01

    Transcriptional control plays an important role in regulating submergence responses in plants. Although numerous genes are highly induced during hypoxia, their individual roles in hypoxic responses are still poorly understood. Here, we found that expression of genes that encode members of the WRKY transcription factor family was rapidly and strongly induced upon submergence in Arabidopsis thaliana, and this induction correlated with induction of a large portion of innate immunity marker genes. Furthermore, prior submergence treatment conferred higher resistance to the bacterial pathogen Pseudomonas syringae in Arabidopsis. Among the WRKY genes tested, WRKY22 had the highest level of induction during the early stages of submergence. Compared with the wild type, WRKY22 T-DNA insertion mutants wrky22-1 and wrky22-2 had lower disease resistance and lower induction of innate immunity markers, such as FLG22-INDUCED RECEPTOR-LIKE KINASE1 (FRK1) and WRKY53, after submergence. Furthermore, transcriptomic analyses of wrky22-2 and chromatin immunoprecipitation identified several potential targets of WRKY22, which included genes encoding a TIR domain–containing protein, a plant peptide hormone, and many OLIGO PEPTIDE TRANSPORTER genes, all of which may lead to induction of innate immunity. In conclusion, we propose that submergence triggers innate immunity in Arabidopsis via WRKY22, a response that may protect against a higher probability of pathogen infection either during or after flooding. PMID:23897923

  18. The Stromal Contribution to the Development of Resistance to New-Generation Drugs by Castration-Resistant Prostate Cancer

    DTIC Science & Technology

    2015-10-01

    associated bone stromal cells to regular stromal cells. That laboratory has many samples of bone marrow stromal cells from patients and has offered to...So et al., Insulin -like growth factor binding protein-2 is a novel therapeutic target associated with breast cancer. Clin. Cancer Res. 14, 6944 (2008

  19. Stromal vascular fraction improves deep partial thickness burn wound healing.

    PubMed

    Atalay, Sibel; Coruh, Atilla; Deniz, Kemal

    2014-11-01

    The practice of early burn wound excision and wound closure by immediate autologous skin or skin substitutes is the preferred treatment in extensive deep partial and full-thickness burns. To date there is no proven definite medical treatment to decrease burn wound size and accelerate burn wound healing in modern clinical practice. Stromal vascular fraction is an autologous mixture that has multiple proven beneficial effects on different kinds of wounds. In our study, we investigated the effects of stromal vascular fraction on deep partial-thickness burn wound healing. In this study, 20 Wistar albino rats were used. Inguinal adipose tissue of the rats was surgically removed and stromal vascular fraction was isolated. Thereafter, deep second-degree burns were performed on the back of the rats by hot water. The rats were divided into two groups in a randomized fashion. The therapy group received stromal vascular fraction, whereas the control group received only physiologic serum by intradermal injection. Assessment of the burn wound healing between the groups was carried out by histopathologic and immuno-histochemical data. Stromal vascular fraction increased vascular endothelial growth factor, proliferating cell nuclear antigen index, and reduced inflammation of the burn wound. Furthermore, vascularization and fibroblastic activity were achieved earlier and observed to be at higher levels in the stromal vascular fraction group. Stromal vascular fraction improves burn wound healing by increasing cell proliferation and vascularization, reducing inflammation, and increasing fibroblastic activity. Copyright © 2014 Elsevier Ltd and ISBI. All rights reserved.

  20. TPL2 (Therapeutic Targeting Tumor Progression Locus-2)/ATF4 (Activating Transcription Factor-4)/SDF1α (Chemokine Stromal Cell-Derived Factor-α) Axis Suppresses Diabetic Retinopathy.

    PubMed

    Lai, De-Wei; Lin, Keng-Hung; Sheu, Wayne Huey-Herng; Lee, Maw-Rong; Chen, Chung-Yu; Lee, Wen-Jane; Hung, Yi-Wen; Shen, Chin-Chang; Chung, Tsung-Ju; Liu, Shing-Hwa; Sheu, Meei-Ling

    2017-09-01

    Diabetic retinopathy is characterized by vasopermeability, vascular leakage, inflammation, blood-retinal barrier breakdown, capillary degeneration, and neovascularization. However, the mechanisms underlying the association between diabetes mellitus and progression retinopathy remain unclear. TPL2 (tumor progression locus 2), a serine-threonine protein kinase, exerts a pathological effect on vascular angiogenesis. This study investigated the role of N(ε)-(carboxymethyl)lysine, a major advanced glycation end products, and the involved TPL2-related molecular signals in diabetic retinopathy using models of in vitro and in vivo and human samples. Serum N(ε)-(carboxymethyl)lysine levels and TPL2 kinase activity were significantly increased in clinical patients and experimental animals with diabetic retinopathy. Intravitreal administration of pharmacological blocker or neutralizing antibody inhibited TPL2 and effectively suppressed the pathological characteristics of retinopathy in streptozotocin-induced diabetic animal models. Intravitreal VEGF (vascular endothelial growth factor) neutralization also suppressed the diabetic retinopathy in diabetic animal models. Mechanistic studies in primary human umbilical vein endothelial cells and primary retinal microvascular endothelial cells from streptozotocin-diabetic rats, db/db mice, and samples from patients with diabetic retinopathy revealed a positive parallel correlation between N(ε)-(carboxymethyl)lysine and the TPL2/chemokine SDF1α (stromal cell-derived factor-α) axis that is dependent on endoplasmic reticulum stress-related molecules, especially ATF4 (activating transcription factor-4). This study demonstrates that inhibiting the N(ε)-(carboxymethyl)lysine-induced TPL2/ATF4/SDF1α axis can effectively prevent diabetes mellitus-mediated retinal microvascular dysfunction. This signaling axis may include the therapeutic potential for other diseases involving pathological neovascularization or macular edema. © 2017

  1. What's New in Gastrointestinal Stromal Tumor Research and Treatment?

    MedlinePlus

    ... Stromal Tumor (GIST) About Gastrointestinal Stromal Tumor What’s New in Gastrointestinal Stromal Tumor Research? Important research on ... Tumors? Key Statistics for Gastrointestinal Stromal Tumors What’s New in Gastrointestinal Stromal Tumor Research? More In Gastrointestinal ...

  2. Human breast adipose-derived stem cells transfected with the stromal cell-derived factor-1 receptor CXCR4 exhibit enhanced viability in human autologous free fat grafts.

    PubMed

    Xu, Fang-tian; Li, Hong-mian; Yin, Qing-Shui; Liu, Da-lie; Nan, Hua; Zhao, Pei-ran; Liang, Shuang-wu

    2014-01-01

    The main complication of autologous free fat tissue transplantation is fat resorption and calcification due to the ischemic necrosis of fat. The promotion of transplant neovascularization soon after autologous free fat grafts may reduce these outcomes. In adulthood, stromal cell-derived factor-1 (SDF-1) and its membrane receptor C-X-C chemokine receptor type 4 (CXCR4) are involved in the homing and migration of multiple stem cell types, neovascularization, and cell proliferation. We hypothesized that CXCR4 may improve the long-term survival of free fat tissue transplants by recruiting endothelial progenitor cells (EPCs) and may therefore improve graft revascularization. In this study, we aimed to determine the effect of human breast adipose-derived stem cells (HBASCs) transfected with the CXCR4 gene on the survival rate of human autologous free fat transplants in nude mice. Human breast adipose-derived stem cells (HBASCs) were expanded ex vivo for 3 passages, labeled with green fluorescent protein (GFP) and transfected with CXCR4 or left untransfected. Autologous fat tissues were mixed with the GFP-labeled, CXCR4-transfected HBASCs (group A), GFP-labeled HBASCs (group B), the known vascularization-promoting agent VEGF (group C), or medium (group D) and then injected subcutaneously into 32 nude mice at 4 spots in a random fashion. Six months later, the transplanted tissue volume and histology were evaluated, and neo-vascularization was quantified by counting the capillaries. CXCR4 and SDF-1α mRNA expression in the transplants was determined using real-time quantitative PCR analysis (qPCR). The data revealed that the control (group D) transplant volume survival was 28.3 ± 4.5%. Mixing CXCR4-transfected (group A) and untransfected (group B) HBASCs significantly increased transplant volume survival (79.5 ± 8.3% and 67.2 ± 5.9%, respectively), whereas VEGF-transfected HBASCs (group C) were less effective (41.2 ± 5.1%). Histological analysis revealed that both types

  3. Nuclear morphometric analysis in gastrointestinal stromal tumors: a preliminary study.

    PubMed

    Ozdamar, Sükrü Oğuz; Bektaş, Sibel; Erdem Ozdamar, Sevim; Gedikoğlu, Gökhan; Doğan Gün, Banu; Bahadir, Burak

    2007-06-01

    Gastrointestinal stromal tumors are considered a specialized group of mesenchymal neoplasms. In this study, the histomorphologic and immunohistochemical features of gastrointestinal stromal tumors are compared with nuclear morphometric results. Morphometric nuclear parameters such as mean area, mean roundness factor, mean form ellipse, mean length and mean perimeter were evaluated in hematoxylin and eosin stained slides of 22 gastrointestinal stromal tumors (9 benign and 13 malignant) by using a computer-assisted image analysis system. Morphometric results were compared with tumor behavior and tumor size, the presence of necrosis, mitotic index, and immunohistochemical expressions of p53 and proliferating cell nuclear antigen. We found that tumor necrosis was correlated with mean nuclear roundness factor, mean nuclear form ellipse, mean nuclear length and mean nuclear perimeter (p<0.05). Mitotic index was also correlated with mean nuclear roundness factor and mean nuclear form ellipse (p<0.05). However, no correlation was found between morphometric features and gastrointestinal stromal tumor behavior, tumor size, or index of proliferating cell nuclear antigen and p53 expressions (p>0.05). In this preliminary study, the relative concordance of the morphometric results and general histomorphologic data exhibited the importance of nuclear morphometric analysis in gastrointestinal stromal tumors. Studies including larger series of cases investigating detailed nuclear morphometric analysis of gastrointestinal stromal tumors are needed.

  4. Genetics Home Reference: gastrointestinal stromal tumor

    MedlinePlus

    ... Treatment of Gastrointestinal Stromal Tumours (Review of NICE Technology Appraisal Guidance 196) (National Institute for Health and ... Society: Treating Gastrointestinal Stromal Tumor (GIST) Cancer.Net: Gastrointestinal ...

  5. RhoA and Rac1 GTPases play major and differential roles in stromal cell–derived factor-1–induced cell adhesion and chemotaxis in multiple myeloma

    PubMed Central

    Azab, Abdel Kareem; Azab, Feda; Blotta, Simona; Pitsillides, Costas M.; Thompson, Brian; Runnels, Judith M.; Roccaro, Aldo M.; Ngo, Hai T.; Melhem, Molly R.; Sacco, Antonio; Jia, Xiaoying; Anderson, Kenneth C.; Lin, Charles P.; Rollins, Barrett J.

    2009-01-01

    The interaction of multiple myeloma (MM) cells with the bone marrow (BM) milieu plays a crucial role in MM pathogenesis. Stromal cell–derived factor-1 (SDF1) regulates homing of MM cells to the BM. In this study, we examined the role of RhoA and Rac1 GTPases in SDF1-induced adhesion and chemotaxis of MM. We found that both RhoA and Rac1 play key roles in SDF1-induced adhesion of MM cells to BM stromal cells, whereas RhoA was involved in chemotaxis and motility. Furthermore, both ROCK and Rac1 inhibitors reduced SDF1-induced polymerization of actin and activation of LIMK, SRC, FAK, and cofilin. Moreover, RhoA and Rac1 reduced homing of MM cells to BM niches. In conclusion, we characterized the role of RhoA and Rac1 GTPases in SDF1-induced adhesion, chemotaxis, and homing of MM cells to the BM, providing the framework for targeting RhoA and Rac1 GTPases as novel MM therapy. PMID:19443661

  6. Interactions between stromal cell--derived keratinocyte growth factor and epithelial transforming growth factor in immune-mediated crypt cell hyperplasia.

    PubMed Central

    Bajaj-Elliott, M; Poulsom, R; Pender, S L; Wathen, N C; MacDonald, T T

    1998-01-01

    Immune reactions in the gut are associated with increased epithelial cell proliferation. Here we have studied the role of keratinocyte growth factor (KGF; FGF7) and transforming growth factor-alpha (TGF-alpha) in the epithelial cell hyperplasia seen in explants of fetal human small intestine after activation of lamina propria T cells with the superantigen Staphylococcus aureus enterotoxin B (SEB). After the addition of SEB to the explants there is a 10-fold increase in KGF mRNA by 72 h of culture. KGF transcripts were abundant in the lamina propria using in situ hybridization and the culture supernatants contained elevated amounts of KGF protein. SEB had no direct effect on KGF mRNA and protein production by cultured lamina propria mesenchymal cells, but both were upregulated by TNF-alpha. Accompanying the increase in KGF there was also an increase in TGF-alpha precursor proteins in the culture supernatants and the phosphorylated form of the EGFR receptor was also detected in the tissue. Increased TGF-alpha precursor proteins were also detected in the supernatants of control explants stimulated with KGF alone. The direct addition of KGF and TGF-alpha enhanced epithelial cell proliferation and antibodies against KGF and TGF-alpha partially inhibited SEB-induced crypt hyperplasia. These results suggest molecular cross-talk between the KGF/KGFR and the TGF-alpha/EGFR in immune-mediated crypt cell hyperplasia. PMID:9788959

  7. Effect of hydrocortisone on multipotent human mesenchymal stromal cells.

    PubMed

    Shipunova, N N; Petinati, N A; Drize, N I

    2013-05-01

    We studied the effect of natural glucocorticosteroid hydrocortisone on total cell production, cloning efficiency, and expression of genes important for the function of mesenchymal stromal cells. Addition of hydrocortisone to the culture medium reduces the total cell yield by 2 times and significantly increased cloning efficiency by 2-3 times; this effect was more pronounced in multipotent mesenchymal stromal cells obtained from female donors. Hydrocortisone had no effect on the expression of immunomodulatory factors produced by multipotent mesenchymal stromal cells. Hydrocortisone inhibits the expression of bone differentiation markers, increases the expression of the early adipocyte differentiation marker at the beginning of culturing, and dramatically stimulates the expression of the late adipocyte differentiation marker throughout the culturing period. The findings suggest that hydrocortisone activates multipotent mesenchymal stromal cells.

  8. Epidermal growth factor (EGF) sustains in vitro primordial follicle viability by enhancing stromal cell proliferation via MAPK and PI3K pathways in the prepubertal, but not adult, cat ovary.

    PubMed

    Fujihara, Mayako; Comizzoli, Pierre; Keefer, Carol L; Wildt, David E; Songsasen, Nucharin

    2014-04-01

    This study examined the influences of epidermal growth factor (EGF) and growth differentiation factor 9 (GDF9) on in vitro viability and activation of primordial follicles in the ovarian tissue of prepubertal (age, <6 mo) versus adult (age, >8 mo) cats. Ovarian cortical slices were cultured in medium containing EGF and/or GDF9 for 14 days. EGF, but not GDF9, improved (P < 0.05) follicle viability in prepubertal donors in a dose-dependent fashion. Neither EGF nor GDF9 enhanced follicle viability in ovarian tissue from adults, and neither factor activated primordial follicles regardless of age group. We then explored how EGF influenced primordial follicles in the prepubertal donors by coincubation with an inhibitor of EGF receptor (AG1478), mitogen-activated protein kinase (MAPK; U0126), or phosphoinositide 3-kinase (PI3K; LY294002). EGF enhanced (P < 0.05) MAPK and AKT phosphorylation, follicle viability, and stromal cell proliferation. These effects were suppressed (P < 0.05) when the tissue was cultured with this growth factor combined with each inhibitor. To identify the underlying influence of age in response to EGF, we assessed cell proliferation and discovered a greater thriving stromal cell population in prepubertal compared to adult tissue. We conclude that EGF plays a significant role in maintaining intraovarian primordial follicle viability (but without promoting activation) in the prepubertal cat. The mechanism of action is via stimulation of MAPK and PI3K signaling pathways that, in turn, promote ovarian cell proliferation. Particularly intriguing is that the ability of cat ovarian cells to multiply in reaction to EGF is age-dependent and highly responsive in prepubertal females.

  9. Multipotent human stromal cells improve cardiac function after myocardial infarction in mice without long-term engraftment

    SciTech Connect

    Iso, Yoshitaka; Spees, Jeffrey L.; E-mail: Jeffrey.Spees@uvm.edu; Serrano, Claudia; Bakondi, Benjamin; Pochampally, Radhika; Song, Yao-Hua; Sobel, Burton E.; Delafontaine, Patrick; Prockop, Darwin J. . E-mail: dprocko@tulane.edu

    2007-03-16

    The aim of this study was to determine whether intravenously administered multipotent stromal cells from human bone marrow (hMSCs) can improve cardiac function after myocardial infarction (MI) without long-term engraftment and therefore whether transitory paracrine effects or secreted factors are responsible for the benefit conferred. hMSCs were injected systemically into immunodeficient mice with acute MI. Cardiac function and fibrosis after MI in the hMSC-treated group were significantly improved compared with controls. However, despite the cardiac improvement, there was no evident hMSC engraftment in the heart 3 weeks after MI. Microarray assays and ELISAs demonstrated that multiple protective factors were expressed and secreted from the hMSCs in culture. Factors secreted by hMSCs prevented cell death of cultured cardiomyocytes and endothelial cells under conditions that mimicked tissue ischemia. The favorable effects of hMSCs appear to reflect the impact of secreted factors rather than engraftment, differentiation, or cell fusion.

  10. Epidemiology, natural history, and risk factors: panel report from the Ninth International Research Conference on Otitis Media.

    PubMed

    Daly, Kathleen A; Hoffman, Howard J; Kvaerner, Kari Jorunn; Kvestad, Ellen; Casselbrant, Margaretha L; Homoe, Preben; Rovers, Maroeska M

    2010-03-01

    The 2007 Recent Advances in Otitis Media Research Conference Panel Report provides an update on otitis media (OM) research published from 2003 to 2007. This report summarizes important trends in disease incidence and prevalence, describes established and newly identified risk factors for acute and chronic OM and OM with effusion, and conveys information on newly discovered genetic factors. In this report, researchers have described declining rates of OM diagnosis, antibiotic prescriptions, offices visits for OM, and middle ear surgery since the licensure and routine use of pneumococcal conjugate vaccine in infants. The panel report also recommends short and long term goals for current and future OM research.

  11. Abstracts presented at the 7th World Alliance for Risk Factor Surveillance (WARFS) Global Conference. October 16-19, 2011. Toronto, Ontario, Canada.

    PubMed

    2012-01-01

    The 7th World Alliance for Risk Factor Surveillance (WARFS) Global Conference, hosted by the Public Health Agency of Canada, was held in Toronto, Ontario, Canada, from October 16 to 19, 2011. Previous WARFS conferences were held in USA (1999), Finland (2001), Australia (2003), Uruguay (2005) and Italy (2007, 2009). WARFS is a global working group on surveillance under the International Union for Health Promotion and Education (IUHPE) It supports the development of risk factor surveillance as a tool for evidence-based public health, acknowledging the importance of this source of information to inform, monitor and evaluate disease prevention and health promotion policies and programs. The theme of the 2011 Global Conference was the role of surveillance in the promotion of health. The Global Conference had 146 registered participants, making it the second most attended WARFS conference in its history. Over the three days, participants attended oral and poster presentations from 30 countries. The conference would not have been possible without the hard work of the International Scientific Committee and the Local Organizing Committee. To highlight the importance and the significance of this conference at an international level, Chronic Diseases and Injuries in Canada (CDIC) is pleased to publish this supplementary issue, which contains 70 abstracts presented at the 7th WARFS Global Conference. In the spirit the Global Conference, this collection of abstracts brings together surveillance material on risk factors, chronic diseases, infectious diseases and injuries from around the world. By making these abstracts widely available, CDIC hopes to further the conference objectives through a continued dialogue between those interested in linking risk factor surveillance to health promotion.

  12. Cellular and extracellular matrix modulation of corneal stromal opacity

    PubMed Central

    Torricelli, Andre A. M.; Wilson, Steven E.

    2014-01-01

    Stromal transparency is a critical factor contributing to normal function of the visual system. Corneal injury, surgery, disease and infection elicit complex wound healing responses that serve to protect against insults and maintain the integrity of the cornea, and subsequently to restore corneal structure and transparency. However, in some cases these processes result in prolonged loss of corneal transparency and resulting diminished vision. Corneal opacity is mediated by the complex actions of many cytokines, growth factors, and chemokines produced by the epithelial cells, stromal cells, bone marrow-derived cells, lacrimal tissues, and nerves. Myofibroblasts, and the disorganized extracellular matrix produced by these cells, are critical determinants of the level and persistence of stromal opacity after corneal injury. Decreases in corneal crystallins in myofibroblasts and corneal fibroblasts contribute to cellular opacity in the stroma. Regeneration of a fully functional epithelial basement membrane (BM) appears to have a critical role in the maintenance of corneal stromal transparency after mild injuries and recovery of transparency when opacity is generated after severe injuries. The epithelial BM likely has a regulatory function whereby it modulates epithelium-derived growth factors such as transforming growth factor (TGF) β and platelet-derived growth factor (PDGF) that drive the development and persistence of myofibroblasts from precursor cells. The purpose of this article is to review the factors involved in the maintenance of corneal transparency and to highlight the mechanisms involved in the appearance, persistency and regression of corneal opacity after stromal injury. PMID:25281830

  13. Traumatic ulcerative granuloma with stromal eosinophilia - Mystery of pathogenesis revisited.

    PubMed

    Sarangarajan, R; Vaishnavi Vedam, V K; Sivadas, G; Sarangarajan, Anuradha; Meera, S

    2015-08-01

    Oral ulcers are a common symptom in clinical practice. Among various causative factors, different types of ulcers in oral cavity exist. Among this, traumatic ulcerative granuloma with stromal eosinophilia (TUGSE) appears to be quite neglected by the clinicians due to the limited knowledge and awareness. On reviewing with a detailed approach to titles and abstracts of articles eliminating duplicates, 40 relevant articles were considered. Randomized studies, review articles, case reports and abstracts were included while conference papers and posters were excluded. Of importance, TUGSE cases been reported only to a minimal extent in the literature. Lack of its awareness tends to lead clinicians to a misconception of cancer. Thus, this particular lesion needs to be differentiated from other malignant lesions to provide a proper mode of treatment. The present article reviews various aspects of the TUGSE with emphasis on the clinical manifestation, pathogenesis, histological, and immunohistochemical study. This study provides the clinician contemporaries, a humble expansion to their knowledge of the disease, based on the searched literature, enabling a more comprehensive management of this rare occurrence.

  14. Dose-dependent insulin regulation of insulin-like growth factor binding protein-1 in human endometrial stromal cells is mediated by distinct signaling pathways.

    PubMed

    Lathi, R B; Hess, A P; Tulac, S; Nayak, N R; Conti, M; Giudice, L C

    2005-03-01

    IGF binding protein-1 (IGFBP-1) is a major product of decidualized human endometrial stromal cells and decidua, and as a modulator of IGF action and/or by independent mechanisms, it regulates cell growth and differentiation and embryonic implantation in these tissues. IGFBP-1 secretion is primarily stimulated by progesterone and cAMP and is inhibited by insulin and IGFs. The signaling pathways mediating the latter are not well defined, and the current study was conducted to determine which pathways mediate the effects of insulin on IGFBP-1 mRNA and protein expression by human endometrial stromal cells decidualized in vitro by progesterone. Cells were cultured and treated with different combinations of insulin; wortmannin, an inhibitor of the phosphatidylinositide-3-kinase (PI3-kinase) pathway; and PD98059, an inhibitor of the MAPK pathway. IGFBP-1 mRNA was determined by real-time PCR, and protein secretion in the conditioned medium was measured by ELISA. Activation of the PI3-kinase and the MAPK pathways was assessed by the detection of phosphorylated AKT and ERK in Western blots, respectively. Insulin inhibited IGFBP-1 mRNA and protein secretion in a dose-dependent fashion, with an ED(50) for the latter 0.127 ng/ml (21.6 pm). Inhibitor studies revealed that at low doses, insulin acts through the PI3-kinase pathway, whereas at higher levels it also activates the MAPK pathway in the inhibition of IGFBP-1. The data demonstrate that human endometrium is a target for insulin action in the regulation of IGFBP-1. At physiological levels insulin likely plays a homeostatic role for energy metabolism in the endometrium, and in hyperinsulinemic states, insulin action on the endometrium may activate cellular mitosis via the MAPK pathway and perhaps predispose this tissue to hyperplasia and/or cancer.

  15. In Vitro Characterization of Patches of Human Mesenchymal Stromal Cells

    PubMed Central

    Roux, Stephan; Bodivit, Gwellaouen; Bartis, Widy; Lebouvier, Angélique; Chevallier, Nathalie; Fialaire-Legendre, Anne; Bierling, Philippe

    2015-01-01

    Stem cells may represent an excellent strategy to improve the healing of skin ulcers. Today the administration mode of stem cells to skin defects remains unsatisfactory. Delivering stem cells with topical treatments represents a new strategy and answering the patients' need. Mesenchymal stromal cells (MSC) have been shown to improve wound healing of cutaneous lesions and amniotic membrane (AM) is known to represent a natural scaffold for cells. The aim of this study is to develop a tissue-engineered product combining MSC and AM for clinical use. In this work we investigated whether the stromal matrix of intact human AM could constitute a scaffold for human MSC derived from either bone marrow (BM) or adipose tissue (AT). For this purpose, clinical-grade AM, MSC, and culture medium were used. We performed experiments of short-term adherence and proliferation for 15 days after the seeding of the cells. Morphological aspects and secretion profiles of MSC onto AM were studied, respectively, by scanning electron microscopy and Luminex analysis. Results demonstrated that the stromal matrix allow the adherence in much greater amount of MSC from BM or AT compared to 2D material. Experiments of proliferation showed that both kinds of MSC could proliferate on the stromal matrix and remain viable 15 days after the seeding of the cells. The 3D analysis of MSC culture demonstrated that both types of MSC invaded the stromal matrix and grew in multiple layers while retaining their fibroblastic morphology. By studying the secretion profile of MSC onto the stromal matrix, we found that both kinds of MSC secrete important cytokines and growth factors for wound healing of cutaneous lesions, such as vascular endothelial growth factor, hepatocyte growth factor, and basic fibroblast growth factor. In conclusion, these results suggest that the stromal matrix of AM seeded with MSC represents a bioactive scaffold that should be evaluated in patients with a nonhealing cutaneous wound. PMID

  16. Stromal loss of TGFβ drives cancer growth in the epithelium via inflammation | Center for Cancer Research

    Cancer.gov

    Interactions between epithelial and stromal cells play an important role in cancer development and progression. Epithelial cancers develop when changes occur to tumor suppressor genes in stromal fibroblast cells. For example, loss of tumor suppressor, p53, in stromal fibroblasts leads to p53 inactivation in the epithelium in a prostate cancer model, and disruption of the transforming growth factor-b receptor II (TGF-βRII) in stromal fibroblasts results in intraepithelial dysplasia in prostate cancer and invasive squamous cell carcinoma (SCC) in mouse forestomach.

  17. Stromal reengineering to treat pancreas cancer

    PubMed Central

    Stromnes, Ingunn M.; DelGiorno, Kathleen E.; Greenberg, Philip D.; Hingorani, Sunil R.

    2014-01-01

    Pancreatic ductal adenocarcinoma co-opts multiple cellular and extracellular mechanisms to create a complex cancer organ with an unusual proclivity for metastasis and resistance to therapy. Cell-autonomous events are essential for the initiation and maintenance of pancreatic ductal adenocarcinoma, but recent studies have implicated critical non-cell autonomous processes within the robust desmoplastic stroma that promote disease pathogenesis and resistance. Thus, non-malignant cells and associated factors are culprits in tumor growth, immunosuppression and invasion. However, even this increasing awareness of non-cell autonomous contributions to disease progression is tempered by the conflicting roles stromal elements can play. A greater understanding of stromal complexity and complicity has been aided in part by studies in highly faithful genetically engineered mouse models of pancreatic ductal adenocarcinoma. Insights gleaned from such studies are spurring the development of therapies designed to reengineer the pancreas cancer stroma and render it permissive to agents targeting cell-autonomous events or to reinstate immunosurveillance. Integrating conventional and immunological treatments in the context of stromal targeting may provide the key to a durable clinical impact on this formidable disease. PMID:24908682

  18. Efficient generation of smooth muscle cells from adipose-derived stromal cells by 3D mechanical stimulation can substitute the use of growth factors in vascular tissue engineering.

    PubMed

    Parvizi, Mojtaba; Bolhuis-Versteeg, Lydia A M; Poot, André A; Harmsen, Martin C

    2016-07-01

    Occluding artery disease causes a high demand for bioartificial replacement vessels. We investigated the combined use of biodegradable and creep-free poly (1,3-trimethylene carbonate) (PTMC) with smooth muscle cells (SMC) derived by biochemical or mechanical stimulation of adipose tissue-derived stromal cells (ASC) to engineer bioartificial arteries. Biochemical induction of cultured ASC to SMC was done with TGF-β1 for 7d. Phenotype and function were assessed by qRT-PCR, immunodetection and collagen contraction assays. The influence of mechanical stimulation on non-differentiated and pre-differentiated ASC, loaded in porous tubular PTMC scaffolds, was assessed after culturing under pulsatile flow for 14d. Assays included qRT-PCR, production of extracellular matrix and scanning electron microscopy. ASC adhesion and TGF-β1-driven differentiation to contractile SMC on PTMC did not differ from tissue culture polystyrene controls. Mesenchymal and SMC markers were increased compared to controls. Interestingly, pre-differentiated ASC had only marginal higher contractility than controls. Moreover, in 3D PTMC scaffolds, mechanical stimulation yielded well-aligned ASC-derived SMC which deposited ECM. Under the same conditions, pre-differentiated ASC-derived SMC maintained their SMC phenotype. Our results show that mechanical stimulation can replace TGF-β1 pre-stimulation to generate SMC from ASC and that pre-differentiated ASC keep their SMC phenotype with increased expression of SMC markers.

  19. Targeting Stromal Androgen Receptor Suppresses Prolactin-Driven Benign Prostatic Hyperplasia (BPH)

    PubMed Central

    Lai, Kuo-Pao; Huang, Chiung-Kuei; Fang, Lei-Ya; Izumi, Kouji; Lo, Chi-Wen; Wood, Ronald; Kindblom, Jon; Yeh, Shuyuan

    2013-01-01

    Stromal-epithelial interaction plays a pivotal role to mediate the normal prostate growth, the pathogenesis of benign prostatic hyperplasia (BPH), and prostate cancer development. Until now, the stromal androgen receptor (AR) functions in the BPH development, and the underlying mechanisms remain largely unknown. Here we used a genetic knockout approach to ablate stromal fibromuscular (fibroblasts and smooth muscle cells) AR in a probasin promoter-driven prolactin transgenic mouse model (Pb-PRL tg mice) that could spontaneously develop prostate hyperplasia to partially mimic human BPH development. We found Pb-PRL tg mice lacking stromal fibromuscular AR developed smaller prostates, with more marked changes in the dorsolateral prostate lobes with less proliferation index. Mechanistically, prolactin mediated hyperplastic prostate growth involved epithelial-stromal interaction through epithelial prolactin/prolactin receptor signals to regulate granulocyte macrophage-colony stimulating factor expression to facilitate stromal cell growth via sustaining signal transducer and activator of transcription-3 activity. Importantly, the stromal fibromuscular AR could modulate such epithelial-stromal interacting signals. Targeting stromal fibromuscular AR with the AR degradation enhancer, ASC-J9®, led to the reduction of prostate size, which could be used in future therapy. PMID:23893956

  20. Dual regeneration of muscle and nerve by intravenous administration of human amniotic fluid-derived mesenchymal stem cells regulated by stromal cell-derived factor-1α in a sciatic nerve injury model.

    PubMed

    Yang, Dar-Yu; Sheu, Meei-Ling; Su, Hong-Lin; Cheng, Fu-Chou; Chen, Ying-Ju; Chen, Chun-Jung; Chiu, Wen-Ta; Yiin, Jia-Jean; Sheehan, Jason; Pan, Hung-Chuan

    2012-06-01

    Human amniotic fluid-derived mesenchymal stem cells (AFMSCs) have been shown to promote peripheral nerve regeneration. The expression of stromal cell-derived factor-1α (SDF-1α) in the injured nerve exerts a trophic effect by recruiting progenitor cells that promote nerve regeneration. In this study, the authors investigated the feasibility of intravenous administration of AFMSCs according to SDF-1α expression time profiles to facilitate neural regeneration in a sciatic nerve crush injury model. Peripheral nerve injury was induced in 63 Sprague-Dawley rats by crushing the left sciatic nerve using a vessel clamp. The animals were randomized into 1 of 3 groups: Group I, crush injury as the control; Group II, crush injury and intravenous administration of AFMSCs (5 × 10(6) cells for 3 days) immediately after injury (early administration); and Group III, crush injury and intravenous administration of AFMSCs (5 × 10(6) cells for 3 days) 7 days after injury (late administration). Evaluation of neurobehavior, electrophysiological study, and assessment of regeneration markers were conducted every week after injury. The expression of SDF-1α and neurotrophic factors and the distribution of AFMSCs in various time profiles were also assessed. Stromal cell-derived factor-1α increased the migration and wound healing of AFMSCs in vitro, and the migration ability was dose dependent. Crush injury induced the expression of SDF-1α at a peak of 10-14 days either in nerve or muscle, and this increased expression paralleled the expression of its receptor, chemokine receptor type-4 (CXCR-4). Most AFMSCs were distributed to the lung during early or late administration. Significant deposition of AFMSCs in nerve and muscle only occurred in the late administration group. Significantly enhanced neurobehavior, electrophysiological function, nerve myelination, and expression of neurotrophic factors and acetylcholine receptor were demonstrated in the late administration group. Amniotic

  1. Potential Effect of CD271 on Human Mesenchymal Stromal Cell Proliferation and Differentiation.

    PubMed

    Calabrese, Giovanna; Giuffrida, Raffaella; Lo Furno, Debora; Parrinello, Nunziatina Laura; Forte, Stefano; Gulino, Rosario; Colarossi, Cristina; Schinocca, Luciana Rita; Giuffrida, Rosario; Cardile, Venera; Memeo, Lorenzo

    2015-07-09

    The Low-Affinity Nerve Growth Factor Receptor (LNGFR), also known as CD271, is a member of the tumor necrosis factor receptor superfamily. The CD271 cell surface marker defines a subset of multipotential mesenchymal stromal cells and may be used to isolate and enrich cells derived from bone marrow aspirate. In this study, we compare the proliferative and differentiation potentials of CD271+ and CD271- mesenchymal stromal cells. Mesenchymal stromal cells were isolated from bone marrow aspirate and adipose tissue by plastic adherence and positive selection. The proliferation and differentiation potentials of CD271+ and CD271- mesenchymal stromal cells were assessed by inducing osteogenic, adipogenic and chondrogenic in vitro differentiation. Compared to CD271+, CD271- mesenchymal stromal cells showed a lower proliferation rate and a decreased ability to give rise to osteocytes, adipocytes and chondrocytes. Furthermore, we observed that CD271+ mesenchymal stromal cells isolated from adipose tissue displayed a higher efficiency of proliferation and trilineage differentiation compared to CD271+ mesenchymal stromal cells isolated from bone marrow samples, although the CD271 expression levels were comparable. In conclusion, these data show that both the presence of CD271 antigen and the source of mesenchymal stromal cells represent important factors in determining the ability of the cells to proliferate and differentiate.

  2. Potential Effect of CD271 on Human Mesenchymal Stromal Cell Proliferation and Differentiation

    PubMed Central

    Calabrese, Giovanna; Giuffrida, Raffaella; Lo Furno, Debora; Parrinello, Nunziatina Laura; Forte, Stefano; Gulino, Rosario; Colarossi, Cristina; Schinocca, Luciana Rita; Giuffrida, Rosario; Cardile, Venera; Memeo, Lorenzo

    2015-01-01

    The Low-Affinity Nerve Growth Factor Receptor (LNGFR), also known as CD271, is a member of the tumor necrosis factor receptor superfamily. The CD271 cell surface marker defines a subset of multipotential mesenchymal stromal cells and may be used to isolate and enrich cells derived from bone marrow aspirate. In this study, we compare the proliferative and differentiation potentials of CD271+ and CD271− mesenchymal stromal cells. Mesenchymal stromal cells were isolated from bone marrow aspirate and adipose tissue by plastic adherence and positive selection. The proliferation and differentiation potentials of CD271+ and CD271− mesenchymal stromal cells were assessed by inducing osteogenic, adipogenic and chondrogenic in vitro differentiation. Compared to CD271+, CD271− mesenchymal stromal cells showed a lower proliferation rate and a decreased ability to give rise to osteocytes, adipocytes and chondrocytes. Furthermore, we observed that CD271+ mesenchymal stromal cells isolated from adipose tissue displayed a higher efficiency of proliferation and trilineage differentiation compared to CD271+ mesenchymal stromal cells isolated from bone marrow samples, although the CD271 expression levels were comparable. In conclusion, these data show that both the presence of CD271 antigen and the source of mesenchymal stromal cells represent important factors in determining the ability of the cells to proliferate and differentiate. PMID:26184166

  3. Treatment for Gastrointestinal Stromal Tumors (GISTs) Based on Tumor Spread

    MedlinePlus

    ... Stromal Tumor Chemotherapy for Gastrointestinal Stromal Tumor Radiation Therapy for Gastrointestinal Stromal Tumor Treatment Choices for Gastrointestinal Stromal Tumor Based on Tumor ... Cancer Information Cancer Prevention & Detection Cancer Basics ...

  4. What Should You Ask Your Doctor about Gastrointestinal Stromal Tumors?

    MedlinePlus

    ... 227.2345 Phone Search Search Category Cancer A-Z Early Detection, Diagnosis, and Staging Can Gastrointestinal Stromal Tumors Be Found Early? Signs and Symptoms of Gastrointestinal Stromal Tumors Tests for Gastrointestinal Stromal Tumors Gastrointestinal Stromal Tumor Stages ...

  5. What Are the Key Statistics about Gastrointestinal Stromal Tumors?

    MedlinePlus

    ... Stromal Tumor (GIST) About Gastrointestinal Stromal Tumor Key Statistics for Gastrointestinal Stromal Tumors Gastrointestinal stromal tumors (GISTs) ... can occur in people at any age. Survival statistics for GIST are discussed in Survival Rates for ...

  6. Angiotensin II enhances epithelial-to-mesenchymal transition through the interaction between activated hepatic stellate cells and the stromal cell-derived factor-1/CXCR4 axis in intrahepatic cholangiocarcinoma.

    PubMed

    Okamoto, Koichi; Tajima, Hidehiro; Nakanuma, Shinichi; Sakai, Seisho; Makino, Isamu; Kinoshita, Jun; Hayashi, Hironori; Nakamura, Keishi; Oyama, Katsunobu; Nakagawara, Hisatoshi; Fujita, Hideto; Takamura, Hiroyuki; Ninomiya, Itasu; Kitagawa, Hirohisa; Fushida, Sachio; Fujimura, Takashi; Harada, Shinichi; Wakayama, Tomohiko; Iseki, Shoichi; Ohta, Tetsuo

    2012-08-01

    We previously reported that hepatic stellate cells (HSCs) activated by angiotensin II (AngII) facilitate stromal fibrosis and tumor progression in intrahepatic cholangiocarcinoma (ICC). AngII has been known as a growth factor which can promote epithelial-to-mesenchymal transition (EMT) in renal epithelial cells, alveolar epithelial cells and peritoneal mesothelial cells. However, in the past, the relationship between AngII and stromal cell-derived factor-1 (SDF-1) in the microenvironment around cancer and the role of AngII on EMT of cancer cells has not been reported in detail. SDF-1 and its specific receptor, CXCR4, are now receiving attention as a mechanism of cell progression and metastasis. In this study, we examined whether activated HSCs promote tumor fibrogenesis, tumor progression and distant metastasis by mediating EMT via the AngII/AngII type 1 receptor (AT-1) and the SDF-1/CXCR4 axis. Two human ICC cell lines and a human HSC line, LI-90, express CXCR4. Significantly higher concentration of SDF-1α was released into the supernatant of LI-90 cells to which AngII had been added. SDF-1α increased the proliferative activity of HSCs and enhanced the activation of HSCs as a growth factor. Furthermore, addition of SDF-1α and AngII enhanced the increase of the migratory capability and vimentin expression, reduced E-cadherin expression, and translocated the expression of β-catenin into the nucleus and cytoplasm in ICC cells. Co-culture with HSCs also enhanced the migratory capability of ICC cells. These findings suggest that SDF-1α, released from activated HSCs and AngII, play important roles in cancer progression, tumor fibrogenesis, and migration in autocrine and paracrine fashion by mediating EMT. Our mechanistic findings may provide pivotal insights into the molecular mechanism of the AngII and SDF-1α-initiated signaling pathway that regulates fibrogenesis in cancerous stroma, tumor progression and meta-stasis of tumor cells expressing AT-1 and CXCR4.

  7. The Brain-Derived Neurotrophic Factor Gene Confers Susceptibility to Bipolar Disorder: Evidence from a Family-Based Association Study

    PubMed Central

    Neves-Pereira, Maria; Mundo, Emanuela; Muglia, Pierandrea; King, Nicole; Macciardi, Fabio; Kennedy, James L.

    2002-01-01

    Bipolar disorder (BP) is a severe psychiatric disease, with a strong genetic component, that affects 1% of the population worldwide and is characterized by recurrent episodes of mania and depression. Brain-derived neurotrophic factor (BDNF) has been implicated in the pathogenesis of mood disorders, and the aim of the present study was to test for the presence of linkage disequilibrium between two polymorphisms in the BDNF gene and BP in 283 nuclear families. Family-based association test (FBAT) results for the dinucleotide repeat (GT)N polymorphism at position −1040 bp showed that allele A3 was preferentially transmitted to the affected individuals (Z=2.035 and P=.042). FBAT results for the val66met SNP showed a significant association for allele G (Z=3.415 and P=.00064). Transmission/disequilibrium test (TDT) haplotype analysis showed a significant result for the 3-G allele combination (P=.000394), suggesting that a DNA variant in the vicinity of the BDNF locus confers susceptibility to BP. Given that there is no direct evidence that either of the polymorphisms we examined alters function, it is unlikely that the actual risk-conferring allele is from these two sites. Rather, the causative site is likely nearby and in linkage disequilibrium with the 3-G haplotype that we have identified. PMID:12161822

  8. Ex Vivo Stromal Cell-Derived Factor 1-Mediated Differentiation of Mouse Bone Marrow Mesenchymal Stem Cells into Hepatocytes Is Enhanced by Chinese Medicine Yiguanjian Drug-Containing Serum

    PubMed Central

    Fu, Linlin; Pang, Bingyao; Zhu, Ying; Wang, Ling; Leng, Aijing; Chen, Hailong

    2016-01-01

    Yiguanjian is administered in traditional Chinese medicine for liver diseases and has been demonstrated to reduce liver fibrosis. This study investigated the effect of Yiguanjian drug-containing serum (YGJ) with Stromal Cell-Derived Factor 1 (SDF-1) and Hepatocyte Growth Factor (HGF) on the differentiation of murine bone-marrow-derived mesenchymal cells (BM-MSCs) into hepatocytes in vitro. Adherent MSCs were isolated from murine bone marrow. Differentiation was induced by 20 ng/mL HGF, 50 ng/mL SDF-1, and 20% Yiguanjian drug-containing serum for 7 to 28 days, and mature hepatocytes' marker albumin (ALB) and cholangiocytes' marker cytokeratin-18 (CK-18) were assessed by immunocytochemistry and western blot. BM-MSCs exhibited homogeneous spindle shape growth after subculture and stained positive for CD90 and negative for CD34. After induction with HGF + normal serum or YGJ for 14 days, HGF + SDF-1 + normal serum for 7 days, or HGF + SDF-1 + YGJ for 5 days, MSCs' morphology changed gradually and begun to resemble hepatocyte-like cells. Cultures supplemented with HGF + SDF-1 + YGJ contained significantly higher proportions of ALB and CK-18 positive cells than cultures supplemented with HGF + SDF-1 + normal serum at day 7. These observations corroborated the results of western blot. In conclusion, Yiguanjian drug-containing serum could facilitate the differentiation of murine BM-MSCs into hepatocytes in vitro and has a synergistic effect with SDF-1 and HGF. PMID:27190538

  9. Phosphatidylcholine-specific phospholipase C/heat shock protein 70 (Hsp70)/transcription factor B-cell translocation gene 2 signaling in rat bone marrow stromal cell differentiation to cholinergic neuron-like cells.

    PubMed

    Shao, Jing; Sun, Chunhui; Su, Le; Zhao, Jing; Zhang, Shangli; Miao, Junying

    2012-12-01

    Although bone marrow stromal cells (BMSCs) can differentiate into neuron-like cells, the mechanisms underlying neuronal differentiation are not well understood. We recently found that inhibition of phosphatidylcholine-specific phospholipase C (PC-PLC) by its inhibitor D609 promoted BMSCs' differentiation into cholinergic neuron-like cells. Using the effective small molecule D609 and gene microarray technology, we investigated the change of gene expression profile to identify key mediators involved in the neuronal differentiation. We selected heat shock protein 70 (Hsp70) and transcription factor B-cell translocation gene 2 (Btg2) that were maximally up-regulated for further study. We found that functional suppression of Hsp70 blocked D609-induced increase of Btg2 expression and cholinergic neuronal differentiation of BMSCs. These results demonstrated that Hsp70 was the pivotal factor in PC-PLC-medicated neuronal differentiation of BMSCs, and Btg2 might be its downstream target. Our findings provide new clues for controlling BMSCs' differentiation into cholinergic neuron-like cells and provide a putative strategy for neurodegenerative diseases therapies. Crown Copyright © 2012. Published by Elsevier Ltd. All rights reserved.

  10. Stromal-cell-derived factor (SDF) 1-alpha in combination with BMP-2 and TGF-β1 induces site-directed cell homing and osteogenic and chondrogenic differentiation for tissue engineering without the requirement for cell seeding.

    PubMed

    Chim, Harvey; Miller, Erin; Gliniak, Christy; Alsberg, Eben

    2012-10-01

    The clinical translation of tissue engineering approaches is limited by the requirement of a cell source. Cell guidance is a new concept that provides an alternative approach, obviating a requirement for an external cell source. This relies on site-specific homing and differentiation of the patient's own cells to an implanted scaffold through controlled delivery of cytokines. In this study, we used stromal-cell-derived factor 1-alpha (SDF-1α) in combination with bone morphogenic protein (BMP)-2 or transforming growth factor (TGF)-β1 to induce cell migration and osteogenic or chondrogenic differentiation, respectively, in implanted scaffolds in a rat model. A customized cytokine microdelivery apparatus was used to ensure the constant rate and concentration of cytokine delivery around the scaffold. The formation of osteoid or early cartilage was observed after 4 weeks in specimens treated with SDF-1α and either BMP-2 or TGF-β1. The density of cellular infiltrate and formation of differentiated tissue were lower in scaffolds treated only with BMP-2 or TGF-β1. Thus, controlled SDF-1α delivery induces cell migration into scaffolds and can result in enhanced osteogenesis and chondrogenesis when used in combination with differentiation cytokines for purposes of tissue engineering.

  11. Upregulation of long non-coding RNA HIF 1α-anti-sense 1 induced by transforming growth factor-β-mediated targeting of sirtuin 1 promotes osteoblastic differentiation of human bone marrow stromal cells

    PubMed Central

    XU, YAO; WANG, SHILONG; TANG, CHAOLIANG; CHEN, WENJUN

    2015-01-01

    The present study aimed to investigate the regulatory mechanism of long non-coding RNA hypoxia-inducible factor 1α-anti-sense 1 (lncRNA HIF1α-AS1) in osteoblast differentiation as well as its targeting by sirtuin 1 (SIRT1), which may be inhibited by transforming growth factor (TGF)-β in bone marrow stromal cells (BMSCs). Real-time polymerase chain reaction (PCR), western blot analysis, lncRNA PCR arrays and chromatin immunoprecipitation were performed in order to examine the interference of SIRT1 expression by TGF-β, the effects of SIRT1 overexpression on lncRNA HIF1α-AS1 and the regulation of the expression of homeobox (HOX)D10, which promotes BMSC differentiation, by lncRNA HIF1α-AS1. The results showed that TGF-β interfered with SIRT1 expression. Furthermore, lncRNA HIF1α-AS1 was significantly downregulated following overexpression of SIRT1. In addition, low expression of HIF1α-AS1 was sufficient to block the expression of HOXD10. The present study further demonstrated that downregulation of HOXD10 by HIF1α-AS1 interfered with acetylation, and subsequently resulted in the inhibition of osteoblast differentiation. These results suggested that HIF1α-AS1 is an essential mediator of osteoblast differentiation, and may thus represent a gene-therapeutic agent for the treatment of human bone diseases. PMID:26460121

  12. Conference on Bio-Social Factors in the Development and Learning of Disadvantaged Children. Conference Proceedings (Syracuse, New York, April 19-21, 1967).

    ERIC Educational Resources Information Center

    Yeshiva Univ., New York, NY. Ferkauf Graduate School of Humanities and Social Sciences.

    These conference proceedings contain two major papers. The paper by Susan S. Stodolsky and Gerald S. Lesser, "Learning Patterns in the Disadvantaged," reports a study of effects of social class and ethnic group influences on levels and patterns of mental ability. Scores for verbal ability, reasoning, number facility, and space conceptualization of…

  13. Zebrafish Stromal Cells have Endothelial Properties and Support Hematopoietic Cells

    PubMed Central

    Lund, Troy C.; Glass, Tiffany J.; Somani, Arif; Nair, Sethu; Tolar, Jakub; Nyquist, Mick; Patrinostro, Xiaobai; Blazar, Bruce R.

    2014-01-01

    Objective The goal of this study was to determine if we could establish a mesenchymal stromal line from zebrafish that would support hematopoietic cells. Such a co-culture system would be a great benefit to study the hematopoietic cell-stromal cell interaction in both the in vitro and in vivo environments. Methods Zebrafish stromal cells, ZStrC, were isolated from the “mesenchymal” tissue of the caudal tail and expanded in a specialized growth media. ZStrC were evaluated for phenotype, gene expression, and the ability to maintain zebrafish marrow cells in co-culture experiments. Results ZStrC showed mesenchymal and endothelial gene expression. Although ZStrC lacked the ability to differentiate into classic MSC lineages (osteocytes, adipocytes, chondrocytes), they did have the capacity for endotube formation on matrigel and LDL-uptake. ZStrC supported marrow cells for greater than 2 weeks in vitro. Importantly, the marrow cells were shown to retain homing ability in adoptive transfer experiments. ZStrC also were shown to improve hematopoietic recovery after sub-lethal irradiation after adoptive transfer. Conclusion As the zebrafish model grows in popularity and importance in the study of hematopoiesis, new tools to aid in our understanding of the hematopoietic cell-stromal cell interaction are required. ZStrC represent an additional tool in the study of hematopoiesis and will be useful to understand the factors that mediate the stromal cell-hematopoietic cell interaction that are important in hematopoietic maintenance. PMID:21920471

  14. Stromal cells in chronic inflammation and tertiary lymphoid organ formation.

    PubMed

    Buckley, Christopher D; Barone, Francesca; Nayar, Saba; Bénézech, Cecile; Caamaño, Jorge

    2015-01-01

    Inflammation is an unstable state. It either resolves or persists. Why inflammation persists and the factors that define tissue tropism remain obscure. Increasing evidence suggests that tissue-resident stromal cells not only provide positional memory but also actively regulate the differential accumulation of inflammatory cells within inflamed tissues. Furthermore, at many sites of chronic inflammation, structures that mimic secondary lymphoid tissues are observed, suggesting that chronic inflammation and lymphoid tissue formation share common activation programs. Similarly, blood and lymphatic endothelial cells contribute to tissue homeostasis and disease persistence in chronic inflammation. This review highlights our increasing understanding of the role of stromal cells in inflammation and summarizes the novel immunological role that stromal cells exert in the persistence of inflammatory diseases.

  15. GUY1 confers complete female lethality and is a strong candidate for a male-determining factor in Anopheles stephensi.

    PubMed

    Criscione, Frank; Qi, Yumin; Tu, Zhijian

    2016-09-20

    Despite their importance in sexual differentiation and reproduction, Y chromosome genes are rarely described because they reside in repeat-rich regions that are difficult to study. Here, we show that Guy1, a unique Y chromosome gene of a major urban malaria mosquito Anopheles stephensi, confers 100% female lethality when placed on the autosomes. We show that the small GUY1 protein (56 amino acids in length) causes female lethality and that males carrying the transgene are reproductively more competitive than their non-transgenic siblings under laboratory conditions. The GUY1 protein is a primary signal from the Y chromosome that affects embryonic development in a sex-specific manner. Our results have demonstrated, for the first time in mosquitoes, the feasibility of stable transgenic manipulation of sex ratios using an endogenous gene from the male-determining chromosome. These results provide insights into the elusive M factor and suggest exciting opportunities to reduce mosquito populations and disease transmission.

  16. Mesenchymal stromal cell cryopreservation.

    PubMed

    Renzi, Sabrina; Lombardo, Tina; Dotti, Silvia; Dessì, Sara S; De Blasio, Pasquale; Ferrari, Maura

    2012-06-01

    The advent of stem cells and stem cell-based therapies for specific diseases requires particular knowledge of laboratory procedures, which not only guarantee the continuous production of cells, but also provide them an identity and integrity as close as possible to their origin. Their cryopreservation at temperatures below -80°C and typically below -140°C is of paramount importance. This target can be achieved by incorporating high molar concentrations of cryoprotectant mixtures that preserve cells from deleterious ice crystal formation. Usually, dimethyl sulfoxide (DMSO) and animal proteins are used as protectant reagents, but unexpected changes in stem cell fate and downstream toxicity effects have been reported, limiting their wide use in clinical settings. In scientific reviews, there are not much data regarding viability of mesenchymal stromal cells (MSCs) after the freezing/thawing process. During our routine analysis, a poor resistance to cryopreservation of these cells was observed, as well as their weak ability to replicate. This is an important point in the study of MSCs; moreover, it represents a limit for preservation and long-term storage. For this reason, MSCs isolated from equine, ovine, and rodent bone marrow and equine adipose tissue were compared using different cryopreservation solutions for this study of vitality. Our findings showed the best results regarding cell viability using a solution of fetal bovine serum with addition of 10% DMSO. In particular, we noted an increase in survival of equine bone marrow MSCs. This parameter has been evaluated by Trypan blue staining at fixed times (0, 24, and 48 hours post-thaw). This result highlights the fact that equine bone marrow MSCs are the frailest we analyzed. Therefore, it could be useful to delve further into this topic in order to improve the storage possibility for these cells and their potential use in cell-based therapies.

  17. Ectopic Expression of JcWRKY Transcription Factor Confers Salinity Tolerance via Salicylic Acid Signaling

    PubMed Central

    Agarwal, Parinita; Dabi, Mitali; Sapara, Komal K.; Joshi, Priyanka S.; Agarwal, Pradeep K.

    2016-01-01

    Plants, being sessile, have developed intricate signaling network to specifically respond to the diverse environmental stress. The plant-specific WRKY TFs form one of the largest TF family and are involved in diverse plant processes, involving growth, development and stress signaling through auto and cross regulation with different genes and TFs. Here, we report the functional characterization of a salicylic acid -inducible JcWRKY TF. The JcWRKY overexpression confers salinity tolerance in transgenic tobacco, as was evident by increased chlorophyll content and seed germination potential. The transgenic plants showed increased soluble sugar, membrane stability, reduced electrolyte leakage and generation of reactive oxygen species (H2O2 and O2•-) as compared to the wild type. Furthermore, the low SA treatment along with salinity improved the tolerance potential of the transgenics by maintaining ROS homeostasis and high K+/Na+ ratio. The transcript expression of SA biosynthetic gene ICS1 and antioxidative enzymes (CAT and SOD) showed upregulation during stress. Thus, the present study reflects that JcWRKY is working in co-ordination with SA signaling to orchestrate the different biochemical and molecular pathways to maneuvre salt stress tolerance of the transgenic plants. PMID:27799936

  18. Signet ring stromal cell tumor revisited and related signet ring cell lesions of the ovary.

    PubMed

    Roth, Lawrence M; Ramzy, Ibrahim

    2014-03-01

    In this article, we revisit the first reported case of ovarian signet ring stromal cell tumor (SRSCT) using modern immunohistochemical techniques and compare it to a case of signet ring cell transformation of lutein cells in an ovarian stromal tumor having components of luteinized thecoma and sclerosing stromal tumor. We introduce a new classification of signet ring stromal cell lesions of the ovary that serves as a framework to distinguish pathogenetically distinct ovarian stromal lesions that may be confused with cases of true SRSCT. The SRSCT in our first case most likely arose directly from the ovarian stroma without an identifiable precursor neoplasm. In our second case, the association of the signet ring cells with lutein cells and the positive staining of the signet ring cells for inhibin and steroidogenic factor 1 confirm that in some instances signet ring cells are derived from lutein cells. © 2014 Elsevier Inc. All rights reserved.

  19. Chondrogenic induction of mesenchymal stromal/stem cells from Wharton's jelly embedded in alginate hydrogel and without added growth factor: an alternative stem cell source for cartilage tissue engineering.

    PubMed

    Reppel, Loïc; Schiavi, Jessica; Charif, Naceur; Leger, Léonore; Yu, Hao; Pinzano, Astrid; Henrionnet, Christel; Stoltz, Jean-François; Bensoussan, Danièle; Huselstein, Céline

    2015-12-30

    Due to their intrinsic properties, stem cells are promising tools for new developments in tissue engineering and particularly for cartilage tissue regeneration. Although mesenchymal stromal/stem cells from bone marrow (BM-MSC) have long been the most used stem cell source in cartilage tissue engineering, they have certain limits. Thanks to their properties such as low immunogenicity and particularly chondrogenic differentiation potential, mesenchymal stromal/stem cells from Wharton's jelly (WJ-MSC) promise to be an interesting source of MSC for cartilage tissue engineering. In this study, we propose to evaluate chondrogenic potential of WJ-MSC embedded in alginate/hyaluronic acid hydrogel over 28 days. Hydrogels were constructed by the original spraying method. Our main objective was to evaluate chondrogenic differentiation of WJ-MSC on three-dimensional scaffolds, without adding growth factors, at transcript and protein levels. We compared the results to those obtained from standard BM-MSC. After 3 days of culture, WJ-MSC seemed to be adapted to their new three-dimensional environment without any detectable damage. From day 14 and up to 28 days, the proportion of WJ-MSC CD73(+), CD90(+), CD105(+) and CD166(+) decreased significantly compared to monolayer marker expression. Moreover, WJ-MSC and BM-MSC showed different phenotype profiles. After 28 days of scaffold culture, our results showed strong upregulation of cartilage-specific transcript expression. WJ-MSC exhibited greater type II collagen synthesis than BM-MSC at both transcript and protein levels. Furthermore, our work highlighted a relevant result showing that WJ-MSC expressed Runx2 and type X collagen at lower levels than BM-MSC. Once seeded in the hydrogel scaffold, WJ-MSC and BM-MSC have different profiles of chondrogenic differentiation at both the phenotypic level and matrix synthesis. After 4 weeks, WJ-MSC, embedded in a three-dimensional environment, were able to adapt to their environment and

  20. Low Stromal Area and High Stromal Microvessel Density Predict Poor Prognosis in Pancreatic Cancer.

    PubMed

    Nishida, Takahiro; Yoshitomi, Hideyuki; Takano, Shigetsugu; Kagawa, Shingo; Shimizu, Hiroaki; Ohtsuka, Masayuki; Kato, Atsushi; Furukawa, Katsunori; Miyazaki, Masaru

    2016-04-01

    Excessive stroma is a unique property of cancer tissue of the pancreas. The aim of this study was to analyze the relationship of cancer stromal area (SA) and tumor microvessel density (MVD) with prognostic and clinicopathological findings. Pancreatic adenocarcinoma tissues obtained from 104 patients were subjected to cytokeratin 19 and CD31 double immunostaining to identify cancer cells and endothelial cells simultaneously. Stromal area and MVD were assessed in the same sections. Patients were divided into 2 groups for each analysis by the median value of the respective measure. Stromal area negatively correlated with MVD. The low SA group harbored more poorly differentiated carcinoma than the high SA group. Patients of the low SA group showed a higher incidence of hematogenous recurrence. As a consequence, patients in the low SA and the high MVD groups had poorer prognosis in terms of both disease-free survival and overall survival than their respective groups. Multivariate analysis showed that a low SA was an independent prognostic factor for disease-free and overall survival. Our data indicate that the stroma of pancreatic cancer may play an auxiliary role as a barrier to cancer cell invasion. The depletion of tumor stroma alone does not suppress pancreatic cancer progression.

  1. Changes in ventricular remodelling and clinical status during the year following a single administration of stromal cell-derived factor-1 non-viral gene therapy in chronic ischaemic heart failure patients: the STOP-HF randomized Phase II trial

    PubMed Central

    Chung, Eugene S.; Miller, Leslie; Patel, Amit N.; Anderson, Russell David; Mendelsohn, Farrell O.; Traverse, Jay; Silver, Kevin H.; Shin, Julia; Ewald, Gregory; Farr, Mary Jane; Anwaruddin, Saif; Plat, Francis; Fisher, Scott J.; AuWerter, Alexander T.; Pastore, Joseph M.; Aras, Rahul; Penn, Marc S.

    2015-01-01

    Background Stromal cell-derived factor-1 (SDF-1) promotes tissue repair through mechanisms of cell survival, endogenous stem cell recruitment, and vasculogenesis. Stromal Cell-Derived Factor-1 Plasmid Treatment for Patients with Heart Failure (STOP-HF) is a Phase II, double-blind, randomized, placebo-controlled trial to evaluate safety and efficacy of a single treatment of plasmid stromal cell-derived factor-1 (pSDF-1) delivered via endomyocardial injection to patients with ischaemic heart failure (IHF). Methods Ninety-three subjects with IHF on stable guideline-based medical therapy and left ventricular ejection fraction (LVEF) ≤40%, completed Minnesota Living with Heart Failure Questionnaire (MLWHFQ) and 6-min walk distance (6 MWD), were randomized 1 : 1 : 1 to receive a single treatment of either a 15 or 30 mg dose of pSDF-1 or placebo via endomyocardial injections. Safety and efficacy parameters were assessed at 4 and 12 months after injection. Left ventricular functional and structural measures were assessed by contrast echocardiography and quantified by a blinded independent core laboratory. Stromal Cell-Derived Factor-1 Plasmid Treatment for Patients with Heart Failure was powered based on change in 6 MWD and MLWHFQ at 4 months. Results Subject profiles at baseline were (mean ± SD): age 65 ± 9 years, LVEF 28 ± 7%, left ventricular end-systolic volume (LVESV) 167 ± 66 mL, N-terminal pro brain natriuretic peptide (BNP) (NTproBNP) 1120 ± 1084 pg/mL, MLWHFQ 50 ± 20 points, and 6 MWD 289 ± 99 m. Patients were 11 ± 9 years post most recent myocardial infarction. Study injections were delivered without serious adverse events in all subjects. Sixty-two patients received drug with no unanticipated serious product-related adverse events. The primary endpoint was a composite of change in 6 MWD and MLWHFQ from baseline to 4 months follow-up. The primary endpoint was not met (P = 0.89). For the patients treated with pSDF-1, there was a trend toward an

  2. A new case of Carney triad: gastrointestinal stromal tumours and leiomyoma of the oesophagus do not show activating mutations of KIT and platelet-derived growth factor receptor alpha.

    PubMed

    Knop, S; Schupp, M; Wardelmann, E; Stueker, D; Horger, M S; Kanz, L; Einsele, H; Kroeber, S M

    2006-10-01

    The Carney triad is a rare syndrome of unknown aetiology, with synchronous or metachronous appearance of rare neoplasms: gastrointestinal stromal tumours (GISTs), pulmonary chondromas and extra-adrenal paragangliomas. In most cases, the Carney triad is incomplete. The combination encountered typically, GISTs and pulmonary chondromas, was also seen in our patient, a 22-year-old woman. She was diagnosed with the triad after Billroth II gastrectomy for histologically proved gastric GISTs. The diagnosis of pulmonary chondromas was confirmed by transthoracic, computed tomography-guided needle biopsy. An oesophageal leiomyoma was resected 2 years after the initial diagnosis, on suspicion of paraganglioma. The clinical course of the patient has been uneventful since. The last follow-up was carried out 6 years after the initial diagnosis. On histological examination, the cells of gastric GIST were partly positive for CD34, whereas CD117 was expressed in all areas in variable intensity and S-100 protein was negative. The oesophageal tumour was classified as leiomyoma due to strong immunopositivity for smooth muscle actin and desmin, being negative for CD34 and CD117. Two different gastric GIST lesions as well as the oesophageal leiomyoma and normal tissue were analysed for activating mutations in common hot spots of KIT (exon 9 and 11) and platelet-derived growth factor receptor alpha (exon 18), but in all probes wild-type sequences were found. These results are in accordance with the first published analyses of GIST lesions from Carney patients.

  3. A new case of Carney triad: gastrointestinal stromal tumours and leiomyoma of the oesophagus do not show activating mutations of KIT and platelet‐derived growth factor receptor α

    PubMed Central

    Knop, S; Schupp, M; Wardelmann, E; Stueker, D; Horger, M S; Kanz, L; Einsele, H; Kroeber, S M

    2006-01-01

    The Carney triad is a rare syndrome of unknown aetiology, with synchronous or metachronous appearance of rare neoplasms: gastrointestinal stromal tumours (GISTs), pulmonary chondromas and extra‐adrenal paragangliomas. In most cases, the Carney triad is incomplete. The combination encountered typically, GISTs and pulmonary chondromas, was also seen in our patient, a 22‐year‐old woman. She was diagnosed with the triad after Billroth II gastrectomy for histologically proved gastric GISTs. The diagnosis of pulmonary chondromas was confirmed by transthoracic, computed tomography‐guided needle biopsy. An oesophageal leiomyoma was resected 2 years after the initial diagnosis, on suspicion of paraganglioma. The clinical course of the patient has been uneventful since. The last follow‐up was carried out 6 years after the initial diagnosis. On histological examination, the cells of gastric GIST were partly positive for CD34, whereas CD117 was expressed in all areas in variable intensity and S‐100 protein was negative. The oesophageal tumour was classified as leiomyoma due to strong immunopositivity for smooth muscle actin and desmin, being negative for CD34 and CD117. Two different gastric GIST lesions as well as the oesophageal leiomyoma and normal tissue were analysed for activating mutations in common hot spots of KIT (exon 9 and 11) and platelet‐derived growth factor receptor α (exon 18), but in all probes wild‐type sequences were found. These results are in accordance with the first published analyses of GIST lesions from Carney patients. PMID:17021135

  4. Analysis of genetic polymorphisms in CCR5, CCR2, stromal cell-derived factor-1, RANTES, and dendritic cell-specific intercellular adhesion molecule-3-grabbing nonintegrin in seronegative individuals repeatedly exposed to HIV-1.

    PubMed

    Liu, Huanliang; Hwangbo, Yon; Holte, Sarah; Lee, Jean; Wang, Chunhui; Kaupp, Nicole; Zhu, Haiying; Celum, Connie; Corey, Lawrence; McElrath, M Juliana; Zhu, Tuofu

    2004-09-15

    To determine the influence of host genetics on human immunodeficiency virus (HIV) type 1 infection, we examined 94 repeatedly exposed seronegative (ES) individuals for polymorphisms in multiple genes and compared the results with those for 316 HIV-1-seropositive and 425 HIV-1-seronegative individuals. The frequency of homozygous C-C chemokine receptor (CCR) 5- Delta 32 was higher in ES (3.2%) than in HIV-1-seropositive individuals (0.0%; P=.012). However, the CCR5-59029A, CCR2-64I, stromal cell-derived factor (SDF)-1-3'A, RANTES (regulated on activation, normally T cell-expressed and -secreted)-403A, and RANTES-28G polymorphisms were not associated with resistance to HIV-1 infection. Furthermore, we identified novel variants in the DC-SIGN (dendritic cell-specific intercellular adhesion molecule-3-grabbing nonintegrin) repeat region and observed that heterozygous DC-SIGN reduced the risk of HIV-1 infection (3.2% in ES individuals vs. 0.0% in HIV-1-seropositive individuals; P=.011).

  5. A role for the Rho-p160 Rho coiled-coil kinase axis in the chemokine stromal cell-derived factor-1alpha-induced lymphocyte actomyosin and microtubular organization and chemotaxis.

    PubMed

    Vicente-Manzanares, Miguel; Cabrero, José Román; Rey, Mercedes; Pérez-Martínez, Manuel; Ursa, Angeles; Itoh, Kazuyuki; Sánchez-Madrid, Francisco

    2002-01-01

    The possible involvement of the Rho-p160ROCK (Rho coiled-coil kinase) pathway in the signaling induced by the chemokine Stromal cell-derived factor (SDF)-1alpha has been studied in human PBL. SDF-1alpha induced activation of RhoA, but not that of Rac. RhoA activation was followed by p160ROCK activation mediated by RhoA, which led to myosin light chain (MLC) phosphorylation, which was dependent on RhoA and p160ROCK activities. The kinetics of MLC activation was similar to that of RhoA and p160ROCK. The role of this cascade in overall cell morphology and functional responses to the chemokine was examined employing different chemical inhibitors. Inhibition of either RhoA or p160ROCK did not block SDF-1alpha-induced short-term actin polymerization, but induced the formation of long spikes arising from the cell body, which were found to be microtubule based. This morphological change was associated with an increase in microtubule instability, which argues for an active microtubule polymerization in the formation of these spikes. Inhibition of the Rho-p160ROCK-MLC kinase signaling cascade at different steps blocked lymphocyte migration and the chemotaxis induced by SDF-1alpha. Our results indicate that the Rho-p160ROCK axis plays a pivotal role in the control of the cell shape as a step before lymphocyte migration toward a chemotactic gradient.

  6. Human skin melanocyte migration towards stromal cell-derived factor-1α demonstrated by optical real-time cell mobility assay: modulation of their chemotactic ability by α-melanocyte-stimulating hormone.

    PubMed

    Yamauchi, Akira; Hadjur, Christophe; Takahashi, Tadahito; Suzuki, Itaru; Hirose, Kunitaka; Mahe, Yann F

    2013-10-01

    To identify potential regulators of normal human melanocyte behaviour, we have developed an in vitro human melanocyte migration assay, using the optically accessible, real-time cell motility assay device TAXIScan. Coating of the glass surface with an extracellular matrix that served as scaffolding molecule was essential to demonstrate efficient melanocyte migration. Among several chemokines tested, stromal cell-derived factor (SDF)-1α/CXCL12 was the most effective driver of human normal skin melanocytes. Incubation of melanocytes with α-melanocyte-stimulating hormone (MSH) before the assay specifically enhanced CXCR4 expression and consequently chemotaxis towards SDF-1α/CXCL12. These results suggest that α-MSH acts on melanocytes to produce melanin as well as stimulates the cells to migrate to the site where they work through CXCR4 up-regulation, which is a new dynamic mode of action of α-MSH on melanocyte physiology. © 2013 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  7. The effect of a stromal cell-derived factor-1α /heparin coating of a biodegradable vascular graft on recruitment of both endothelial and smooth muscle progenitor cells and accelerated healing

    PubMed Central

    Yu, Jian; Wang, Aijun; Tang, Zhenyu; Henry, Jeffrey; Lee, Benjamin; Zhu, Yiqian; Yuan, Failei; Huang, Fengping; Li, Song

    2012-01-01

    Small-diameter synthetic vascular grafts have high failure rate and tissue-engineered blood vessels are limited by the scalability. Here we engineered bioactive materials for in situ vascular tissue engineering, which recruits two types of endogenous progenitor cells for the regeneration of blood vessels. Heparin was conjugated to microfibrous vascular grafts to suppress thrombogenic responses, and stromal cell-derived factor-1α (SDF-1α) was immobilized onto heparin to recruit endogenous progenitor cells. Heparin-bound SDF-1α was more stable than adsorbed SDF-1α under both static and flow conditions. Microfibrous grafts were implanted in rats by anastomosis to test the functional performance. Heparin coating improved the short-term patency, and immobilized SDF-1α further improved the long-term patency. SDF-1α effectively recruited endothelial progenitor cells (EPCs) to the luminal surface of the grafts, which differentiated into endothelial cells (ECs) and accelerated endothelialization. More interestingly, SDF-1α increased the recruitment of smooth muscle progenitor cells (SMPCs) to the grafts, and SMPCs differentiated into smooth muscle cells (SMCs) in vivo and in vitro. Consistently, SDF-1α-immobilized grafts had significantly higher elastic modulus. This work demonstrates the feasibility of simultaneously recruiting progenitor cells of ECs and SMCs for in situ blood vessel regeneration. This in situ tissue engineering approach will have broad applications in regenerative medicine. PMID:22884813

  8. Human mesenchymal stromal cells suppress T-cell proliferation independent of heme oxygenase-1.

    PubMed

    Patel, Seema R; Copland, Ian B; Garcia, Marco A; Metz, Richard; Galipeau, Jacques

    2015-04-01

    Mesenchymal stromal cells deploy immune suppressive properties amenable for use as cell therapy for inflammatory disorders. It is now recognized that mesenchymal stromal cells necessitate priming with an inflammatory milieu, in particular interferon-γ, to exert augmented immunosuppressive effects. It has been recently suggested that the heme-catabolizing enzyme heme oxygenase-1 is an essential component of the mesenchymal stromal cell-driven immune suppressive response. Because mesenchymal stromal cells upregulate indoleamine 2,3-dioxygenase expression on interferon-γ priming and indoleamine 2,3-dioxygenase requires heme as a cofactor for optimal catabolic function, we investigated the potential antagonism of heme oxygenase-1 activity on indoleamine 2, 3-dioxygenase and the impact on mesenchymal stromal cell immune plasticity. We herein sought to evaluate the molecular genetic effect of cytokine priming on human mesenchymal stromal cell heme oxygenase-1 expression and its functional role in differentially primed mesenchymal stromal cells. Contrary to previous reports, messenger RNA and protein analyses demonstrated that mesenchymal stromal cells derived from normal subjects (n = 6) do not express heme oxygenase-1 at steady state or after interferon-γ, tumor necrosis factor-α, and/or transforming growth factor-β priming. Pharmacological inhibition of heme oxygenase-1 with the use of tin protoporphyrin did not significantly abrogate the ability of mesenchymal stromal cells to suppress T-cell proliferation in vitro. Overall, these results unequivocally demonstrate that under steady state and after cytokine priming, human mesenchymal stromal cells immunoregulate T-cell proliferation independent of heme oxygenase-1.

  9. Identification of bone-derived factors conferring de novo therapeutic resistance in metastatic prostate cancer

    PubMed Central

    Lee, Yu-Chen; Lin, Song-Chang; Yu, Guoyu; Cheng, Chien-Jui; Liu, Bin; Liu, Hsuan-Chen; Hawke, David H.; Parikh, Nila U.; Varkaris, Andreas; Corn, Paul; Logothetis, Christopher; Satcher, Robert L.; Yu-Lee, Li-Yuan; Gallick, Gary E.; Lin, Sue-Hwa

    2015-01-01

    Resistance to currently available targeted therapies significantly hampers the survival of prostate cancer (PCa) patients with bone metastasis. Here we demonstrate an important resistance mechanism initiated from tumor-induced bone. Studies using an osteogenic patient-derived xenograft, MDA-PCa-118b, revealed that tumor cells resistant to cabozantinib, a Met and VEGFR-2 inhibitor, reside in a "resistance niche" adjacent to PCa-induced bone. We performed secretome analysis of the conditioned medium from tumor-induced bone to identify proteins (termed "osteocrines") found within this resistance niche. In accordance with previous reports demonstrating that activation of integrin signaling pathways confers therapeutic resistance, 27 of the 90 osteocrines identified were integrin ligands. We found that following cabozantinib treatment, only tumor cells positioned adjacent to newly-formed woven bone remained viable and expressed high levels of pFAK-Y397 and pTalin-S425, mediators of integrin signaling. Accordingly, treatment of C4-2B4 cells with integrin ligands resulted in increased pFAK-Y397 expression and cell survival, whereas targeting integrins with FAK inhibitors PF-562271 or defactinib inhibited FAK phosphorylation and reduced the survival of PC3-mm2 cells. Moreover, treatment of MDA-PCa-118b tumors with PF-562271 led to decreased tumor growth, irrespective of initial tumor size. Finally, we show that upon treatment cessation, the combination of PF-562271 and cabozantinib delayed tumor recurrence in contrast to cabozantinib treatment alone. Our studies suggest that identifying paracrine de novo resistance mechanisms may significantly contribute to the generation of a broader set of potent therapeutic tools that act combinatorially to inhibit metastatic PCa. PMID:26530902

  10. Identification of Bone-Derived Factors Conferring De Novo Therapeutic Resistance in Metastatic Prostate Cancer.

    PubMed

    Lee, Yu-Chen; Lin, Song-Chang; Yu, Guoyu; Cheng, Chien-Jui; Liu, Bin; Liu, Hsuan-Chen; Hawke, David H; Parikh, Nila U; Varkaris, Andreas; Corn, Paul; Logothetis, Christopher; Satcher, Robert L; Yu-Lee, Li-Yuan; Gallick, Gary E; Lin, Sue-Hwa

    2015-11-15

    Resistance to currently available targeted therapies significantly hampers the survival of patients with prostate cancer with bone metastasis. Here we demonstrate an important resistance mechanism initiated from tumor-induced bone. Studies using an osteogenic patient-derived xenograft, MDA-PCa-118b, revealed that tumor cells resistant to cabozantinib, a Met and VEGFR-2 inhibitor, reside in a "resistance niche" adjacent to prostate cancer-induced bone. We performed secretome analysis of the conditioned medium from tumor-induced bone to identify proteins (termed "osteocrines") found within this resistance niche. In accordance with previous reports demonstrating that activation of integrin signaling pathways confers therapeutic resistance, 27 of the 90 osteocrines identified were integrin ligands. We found that following cabozantinib treatment, only tumor cells positioned adjacent to the newly formed woven bone remained viable and expressed high levels of pFAK-Y397 and pTalin-S425, mediators of integrin signaling. Accordingly, treatment of C4-2B4 cells with integrin ligands resulted in increased pFAK-Y397 expression and cell survival, whereas targeting integrins with FAK inhibitors PF-562271 or defactinib inhibited FAK phosphorylation and reduced the survival of PC3-mm2 cells. Moreover, treatment of MDA-PCa-118b tumors with PF-562271 led to decreased tumor growth, irrespective of initial tumor size. Finally, we show that upon treatment cessation, the combination of PF-562271 and cabozantinib delayed tumor recurrence in contrast to cabozantinib treatment alone. Our studies suggest that identifying paracrine de novo resistance mechanisms may significantly contribute to the generation of a broader set of potent therapeutic tools that act combinatorially to inhibit metastatic prostate cancer. ©2015 American Association for Cancer Research.

  11. The corneal fibrosis response to epithelial-stromal injury.

    PubMed

    Torricelli, Andre A M; Santhanam, Abirami; Wu, Jiahui; Singh, Vivek; Wilson, Steven E

    2016-01-01

    The corneal wound healing response, including the development of stromal opacity in some eyes, is a process that often leads to scarring that occurs after injury, surgery or infection to the cornea. Immediately after epithelial and stromal injury, a complex sequence of processes contributes to wound repair and regeneration of normal corneal structure and function. In some corneas, however, often depending on the type and extent of injury, the response may also lead to the development of mature vimentin+ α-smooth muscle actin+ desmin+ myofibroblasts. Myofibroblasts are specialized fibroblastic cells generated in the cornea from keratocyte-derived or bone marrow-derived precursor cells. The disorganized extracellular matrix components secreted by myofibroblasts, in addition to decreased expression of corneal crystallins in these cells, are central biological processes that result in corneal stromal fibrosis associated with opacity or "haze". Several factors are associated with myofibroblast generation and haze development after PRK surgery in rabbits, a reproducible model of scarring, including the amount of tissue ablated, which may relate to the extent of keratocyte apoptosis in the early response to injury, irregularity of stromal surface after surgery, and changes in corneal stromal proteoglycans, but normal regeneration of the epithelial basement membrane (EBM) appears to be a critical factor determining whether a cornea heals with relative transparency or vision-limiting stromal opacity. Structural and functional abnormalities of the regenerated EBM facilitate prolonged entry of epithelium-derived growth factors such as transforming growth factor β (TGF-β) and platelet-derived growth factor (PDGF) into the stroma that both drive development of mature myofibroblasts from precursor cells and lead to persistence of the cells in the anterior stroma. A major discovery that has contributed to our understanding of haze development is that keratocytes and corneal

  12. The corneal fibrosis response to epithelial-stromal injury

    PubMed Central

    Torricelli, Andre A. M.; Santhanam, Abirami; Wu, Jiahui; Singh, Vivek; Wilson, Steven E.

    2014-01-01

    The corneal wound healing response, including the development of stromal opacity in some eyes, is a process that often leads to scarring that occurs after injury, surgery or infection to the cornea. Immediately after epithelial and stromal injury, a complex sequence of processes contributes to wound repair and regeneration of normal corneal structure and function. In some corneas, however, often depending on the type and extent of injury, the response may also lead to the development of mature vimentin+ α-smooth muscle actin+ desmin+ myofibroblasts. Myofibroblasts are specialized fibroblastic cells generated in the cornea from keratocyte-derived or bone marrow-derived precursor cells. The disorganized extracellular matrix components secreted by myofibroblasts, in addition to decreased expression of corneal crystallins in these cells, are central biological processes that result in corneal stromal fibrosis associated with opacity or “haze”. Several factors are associated with myofibroblast generation and haze development after PRK surgery in rabbits, a reproducible model of scarring, including the amount of tissue ablated, which may relate to the extent of keratocyte apoptosis in the early response to injury, irregularity of stromal surface after surgery, and changes on corneal stromal proteoglycans, but normal regeneration of the epithelial basement membrane (EBM) appears to be a critical factor determining whether a cornea heals with relative transparency or vision-limiting stromal opacity. Structural and functional abnormalities of the regenerated EBM facilitate prolonged entry of epithelium-derived growth factors such as transforming growth factor β (TGF-β) and platelet-derived growth factor (PDGF) into the stroma that both drive development of mature myofibroblasts from precursor cells and lead to persistence of the cells in the anterior stroma. A major discovery that has contributed to our understanding of haze development is that keratocytes and

  13. Social Factors in the Health of Families: A Public Health Social Work Responsibility. Proceedings of a Conference (Pittsburgh, Pennsylvania, March 23-26, 1986).

    ERIC Educational Resources Information Center

    St. Denis, Gerald C., Ed.

    This document contains a list of planning committee members, institute participants, an introduction by Gerald C. St. Denis a program agenda, and institute presentations from this conference. The following presentations are included: (1) "Social Factors in the Health of Families: A Public Health Social Work Responsibility" (Stanley F. Battle); (2)…

  14. Proceedings of the Conference on Ecological and Cultural Factors Related to Emotional Disturbances in Puerto Rican Children and Youth, Barranquitas, Puerto Rico, December 8-10, 1971.

    ERIC Educational Resources Information Center

    Moran, Roberto E., Ed.

    The Conference on Ecological and Cultural Factors Related to Emotional Disturbance in Puerto Rican Children and Youth was the primary attempt to bring together a group of behavioral scientists, medical doctors, and educators, so that the scientific findings of the former--behavioral and medical scientists--may be used by the latter--educators--in…

  15. Infant Mortality, Morbidity, and Childhood Handicapping Conditions: Psychosocial Factors. Based on Proceedings of a Bi-Regional Conference (Atlanta, Georgia, June 2-5, 1985).

    ERIC Educational Resources Information Center

    Watkins, Elizabeth L., Ed.; Melnick, Leslie R., Ed.

    In Part I, "Extent of Knowledge and Implications for Social Work Intervention," the following conference papers are presented: (1) "Unintended Pregnancy and Infant Mortality, Strategies and Interventions" (Alfred W. Brann, Jr.); (2) "Implications for Social Work Intervention in Biopsychosocial Factors Associated with Infant Mortality and…

  16. Analysis of stromal cell secretomes reveals a critical role for stromal cell-derived HGF and fibronectin in angiogenesis

    PubMed Central

    Newman, Andrew C.; Chou, Wayne; Welch-Reardon, Katrina M.; Fong, Ashley H.; Popson, Stephanie A.; Phan, Duc Thien; Sandoval, Daniel R.; Nguyen, Dananh P.; Gershon, Paul D.; Hughes, Christopher C. W.

    2013-01-01

    Objective Angiogenesis requires tightly coordinated cross-talk between endothelial cells and stromal cells such as fibroblasts and smooth muscle cells. The specific molecular mechanisms moderating this process are still poorly understood. Method and Results Stromal cell-derived factors are essential for endothelial cell sprouting and lumen formation. We therefore compared the abilities of two primary fibroblast isolates and a primary smooth muscle cell isolate to promote in vitro angiogenesis and analyzed their secretomes using a combination of nanoLC-MS/MS, qPCR and ELISA. Each isolate exhibited a different level of angiogenic ability. Using quantitative MS, we then compared the secretomes of a fibroblast isolate exhibiting low angiogenic activity, a fibroblast isolate exhibiting high angiogenic activity and human umbilical vein endothelial cells. High angiogenic fibroblast supernatants exhibited an over-abundance of proteins associated with extracellular matrix constituents compared to low angiogenic fibroblasts or endothelial cells. Finally, siRNA technology and purified protein were used to confirm a role for stromal cell-derived hepatocyte growth factor and fibronectin in inducing endothelial cell sprouting. Conclusion Differences in stromal cell ability to induce angiogenesis are due to differences in the secreted proteomes of both extracellular matrix proteins and pro-angiogenic growth factors. PMID:23288153

  17. Endometrial Stromal and Epithelial Cells Exhibit Unique Aberrant Molecular Defects in Patients With Endometriosis.

    PubMed

    Logan, Philip C; Yango, Pamela; Tran, Nam D

    2017-01-01

    Endometriosis is a chronic inflammatory disease that causes pain and infertility in women of reproductive age. To investigate the pathologic pathways in endometrial stromal and epithelial cells that contribute to the manifestation of endometriosis. In vitro cellular and molecular analyses of isolated eutopic endometrial stromal and epithelial cells. Eutopic stromal and epithelial cells from endometriotic and normal patients were isolated by fluorescence-activated cell sorting for paired sibling RNA sequencing and microRNA microarray. Aberrant pathways were identified using ingenuity pathway analysis networks and confirmed with in vitro modulation of the affected pathways in stromal and epithelial cell cultures. Both stromal versus epithelial cell types and paired endometriotic versus normal samples exhibited distinct hierarchical clustering. Compared to normal samples, there were 151 and 215 differentially expressed genes in the endometriotic stromal and epithelial populations, respectively, and concomitantly 9 and 16 differentially expressed microRNAs. Overall, endometriotic stromal and epithelial cells revealed distinct defects. In endometriotic stromal cells, key decidualization genes Zinc finger E-box Binding protein 1 (ZEB1), Heart And Neural crest Derivatives expressed 2 (HAND2), WNT4, and Interleukin 15 (IL-15) were found to be downregulated and Periostin (POSTN) and Matrix Metallopeptidase 7 (MMP7) were upregulated. Specifically, ZEB1 was downregulated in stromal cells by aberrant elevation in miR-200b. In contrast, ZEB1 was found to be upregulated in endometriotic epithelial cells through associated upregulation of transforming growth factor β1 (TGFβ1), inducer of the TGFβ1-Bone Morphogenetic Protein 2 (BMP2)-MMP2-Prostaglandin-endoperoxide Synthase 2 (COX2)-ZEB1 pathway, which activates epithelial-mesenchymal transition. Manifestation of endometriosis involves dysregulation of unique molecular pathways within the diseased endometrial stromal and

  18. Resistin-induced stromal cell-derived factor-1 expression through Toll-like receptor 4 and activation of p38 MAPK/ NFκB signaling pathway in gastric cancer cells.

    PubMed

    Hsieh, Yung-Yu; Shen, Chien-Heng; Huang, Wen-Shih; Chin, Chih-Chien; Kuo, Yi-Hung; Hsieh, Meng Chiao; Yu, Hong-Ren; Chang, Te-Sheng; Lin, Tseng-Hsi; Chiu, Yung-Wei; Chen, Cheng-Nan; Kuo, Hsing-Chun; Tung, Shui-Yi

    2014-06-14

    Stromal cell-derived factor-1 (SDF-1) (CXC chemokine ligand-12)/CXC chemokine receptor 4 (CXCR4) is involved in the carcinogenesis of human gastric cancer, where it stimulates angiogenesis and favors metastasis of tumor cells to distant organs. In addition, resistin is suggested to be an important link between obesity and the development of gastric cancer. Resistin has identified as an important player in inflammatory responses, and emerged as a mediator in inflammation-associated cancer. A limited number of studies have investigated the association of resistin and SDF-1 with gastric cancer. Herein, we investigated the molecular mechanisms by which resistin influences the expression of SDF-1 in gastric carcinoma cells. Human gastric cancer cell lines were exposed to doses of resistin; SDF-1 expression and secretion levels were then determined. Real-time polymerase chain reaction and western blotting analyses were performed to clarify molecular changes. Inhibition of Toll-like receptor 4 (TLR4) by a competitive antagonist inhibited resistin-induced SDF-1 expression. Pharmacological inhibitors and small interfering RNA (siRNA) demonstrated that activation of the p38 mitogen-activated protein kinase (MAPK) pathway is critical for resistin-induced SDF-1 expression mediated by TLR4. The promoter activity and transcription factor enzyme-linked immunosorbent assay revealed that resistin induced expression of SDF-1 mediated by NF-κB in gastric cancer cells. Inhibition of p38 MARK activation blocked the SDF-1-induced expression and the SDF-1 promoter activity in the cancer gastric cells. Chromatin immunoprecipitation assay revealed that inhibition of p38 MARK activation also blocked the resistin-increased NF-κB-DNA-binding activity. Resistin-induced SDF-1 upregulation by activation of TLR4, p38 MARK and NF-κB may explain a new role of resistin in the link of obesity and gastric cancer.

  19. Mesenchymal stromal cells expressing ErbB-2/neu elicit protective antibreast tumor immunity in vivo, which is paradoxically suppressed by IFN-gamma and tumor necrosis factor-alpha priming.

    PubMed

    Romieu-Mourez, Raphaëlle; François, Moïra; Abate, Amanda; Boivin, Marie-Noëlle; Birman, Elena; Bailey, Dana; Bramson, Jonathan L; Forner, Kathy; Young, Yoon-Kow; Medin, Jeffrey A; Galipeau, Jacques

    2010-10-15

    It is unknown whether mesenchymal stromal cells (MSC) can regulate immune responses targeting tumor autoantigens of low immunogenicity. We tested here whether immunization with MSC could break immune tolerance towards the ErbB-2/HER-2/neu tumor antigen and the effects of priming with IFN-γ and tumor necrosis factor-α (TNF-α) on this process. BALB/c- and C57BL/6-derived MSC were lentivirally transduced to express a kinase-inactive rat neu mutant (MSC/Neu). Immunization of BALB/c mice with nontreated or IFN-γ-primed allogeneic or syngeneic MSC/Neu induced similar levels of anti-neu antibody titers; however, only syngeneic MSC/Neu induced protective neu-specific CD8(+) T cell responses. Compared to immunization with nontreated or IFN-γ-primed syngeneic MSC/Neu, the number of circulating neu-specific CD8(+) T cells and titers of anti-neu antibodies were observed to be decreased after immunizations with IFN-γ- plus TNF-α-primed MSC/Neu. In addition, syngeneic MSC/Neu seemed more efficient than IFN-γ-primed MSC/Neu at inducing a protective therapeutic antitumor immune response resulting in the regression of transplanted neu-expressing mammary tumor cells. In vitro antigen-presenting cell assays performed with paraformaldehyde-fixed or live MSC showed that priming with IFN-γ plus TNF-α, compared to priming with IFN-γ alone, increased antigen presentation as well as the production of immunosuppressive factors. These data suggest that whereas MSC could effectively serve as antigen-presenting cells to induce immune responses aimed at tumor autoantigens, these functions are critically regulated by IFN-γ and TNF-α.

  20. Resistin-induced stromal cell-derived factor-1 expression through Toll-like receptor 4 and activation of p38 MAPK/ NFκB signaling pathway in gastric cancer cells

    PubMed Central

    2014-01-01

    Background Stromal cell-derived factor-1 (SDF-1) (CXC chemokine ligand-12)/CXC chemokine receptor 4 (CXCR4) is involved in the carcinogenesis of human gastric cancer, where it stimulates angiogenesis and favors metastasis of tumor cells to distant organs. In addition, resistin is suggested to be an important link between obesity and the development of gastric cancer. Resistin has identified as an important player in inflammatory responses, and emerged as a mediator in inflammation-associated cancer. A limited number of studies have investigated the association of resistin and SDF-1 with gastric cancer. Herein, we investigated the molecular mechanisms by which resistin influences the expression of SDF-1 in gastric carcinoma cells. Results Human gastric cancer cell lines were exposed to doses of resistin; SDF-1 expression and secretion levels were then determined. Real-time polymerase chain reaction and western blotting analyses were performed to clarify molecular changes. Inhibition of Toll-like receptor 4 (TLR4) by a competitive antagonist inhibited resistin-induced SDF-1 expression. Pharmacological inhibitors and small interfering RNA (siRNA) demonstrated that activation of the p38 mitogen-activated protein kinase (MAPK) pathway is critical for resistin-induced SDF-1 expression mediated by TLR4. The promoter activity and transcription factor enzyme-linked immunosorbent assay revealed that resistin induced expression of SDF-1 mediated by NF-κB in gastric cancer cells. Inhibition of p38 MARK activation blocked the SDF-1-induced expression and the SDF-1 promoter activity in the cancer gastric cells. Chromatin immunoprecipitation assay revealed that inhibition of p38 MARK activation also blocked the resistin-increased NF-κB-DNA-binding activity. Conclusions Resistin-induced SDF-1 upregulation by activation of TLR4, p38 MARK and NF-κB may explain a new role of resistin in the link of obesity and gastric cancer. PMID:24929539

  1. The NAC family transcription factor OsNAP confers abiotic stress response through the ABA pathway.

    PubMed

    Chen, Xu; Wang, Yaofeng; Lv, Bo; Li, Jie; Luo, Liqiong; Lu, Songchong; Zhang, Xuan; Ma, Hong; Ming, Feng

    2014-03-01

    Plants respond to environmental stresses by altering gene expression, and several genes have been found to mediate stress-induced expression, but many additional factors are yet to be identified. OsNAP is a member of the NAC transcription factor family; it is localized in the nucleus, and shows transcriptional activator activity in yeast. Analysis of the OsNAP transcript levels in rice showed that this gene was significantly induced by ABA and abiotic stresses, including high salinity, drought and low temperature. Rice plants overexpressing OsNAP did not show growth retardation, but showed a significantly reduced rate of water loss, enhanced tolerance to high salinity, drought and low temperature at the vegetative stage, and improved yield under drought stress at the flowering stage. Microarray analysis of transgenic plants overexpressing OsNAP revealed that many stress-related genes were up-regulated, including OsPP2C06/OsABI2, OsPP2C09, OsPP2C68 and OsSalT, and some genes coding for stress-related transcription factors (OsDREB1A, OsMYB2, OsAP37 and OsAP59). Our data suggest that OsNAP functions as a transcriptional activator that plays a role in mediating abiotic stress responses in rice.

  2. Frameshift Mutation Confers Function as Virulence Factor to Leucine-Rich Repeat Protein from Acidovorax avenae

    PubMed Central

    Kondo, Machiko; Hirai, Hiroyuki; Furukawa, Takehito; Yoshida, Yuki; Suzuki, Aika; Kawaguchi, Takemasa; Che, Fang-Sik

    2017-01-01

    Many plant pathogens inject type III (T3SS) effectors into host cells to suppress host immunity and promote successful infection. The bacterial pathogen Acidovorax avenae causes brown stripe symptom in many species of monocotyledonous plants; however, individual strains of each pathogen infect only one host species. T3SS-deleted mutants of A. avenae K1 (virulent to rice) or N1141 (virulent to finger millet) caused no symptom in each host plant, suggesting that T3SS effectors are involved in the symptom formation. To identify T3SS effectors as virulence factors, we performed whole-genome and predictive analyses. Although the nucleotide sequence of the novel leucine-rich repeat protein (Lrp) gene of N1141 had high sequence identity with K1 Lrp, the amino acid sequences of the encoded proteins were quite different due to a 1-bp insertion within the K1 Lrp gene. An Lrp-deleted K1 strain (KΔLrp) did not cause brown stripe symptom in rice (host plant for K1); by contrast, the analogous mutation in N1141 (NΔLrp) did not interfere with infection of finger millet. In addition, NΔLrp retained the ability to induce effector-triggered immunity (ETI), including hypersensitive response cell death and expression of ETI-related genes. These data indicated that K1 Lrp functions as a virulence factor in rice, whereas N1141 Lrp does not play a similar role in finger millet. Yeast two-hybrid screening revealed that K1 Lrp interacts with oryzain α, a pathogenesis-related protein of the cysteine protease family, whereas N1141 Lrp, which contains LRR domains, does not. This specific interaction between K1 Lrp and oryzain α was confirmed by Bimolecular fluorescence complementation assay in rice cells. Thus, K1 Lrp protein may have acquired its function as virulence factor in rice due to a frameshift mutation. PMID:28101092

  3. Functional Differentiation of Uterine Stromal Cells Involves Cross-regulation between Bone Morphogenetic Protein 2 and Kruppel-like Factor (KLF) Family Members KLF9 and KLF13

    USDA-ARS?s Scientific Manuscript database

    The inability of the uterine epithelium to enter a state of receptivity for the embryo to implant is a significant underlying cause of early pregnancy loss. We previously showed that mice null for the Progesterone Receptor (PGR)-interacting protein Kruppel-like Factor (KLF) 9 are subfertile and exhi...

  4. Multiple factors confer specific Cdc42 and Rac protein activation by dedicator of cytokinesis (DOCK) nucleotide exchange factors.

    PubMed

    Kulkarni, Kiran; Yang, Jing; Zhang, Ziguo; Barford, David

    2011-07-15

    DOCK (dedicator of cytokinesis) guanine nucleotide exchange factors (GEFs) activate the Rho-family GTPases Rac and Cdc42 to control cell migration, morphogenesis, and phagocytosis. The DOCK A and B subfamilies activate Rac, whereas the DOCK D subfamily activates Cdc42. Nucleotide exchange is catalyzed by a conserved DHR2 domain (DOCK(DHR2)). Although the molecular basis for DOCK(DHR2)-mediated GTPase activation has been elucidated through structures of a DOCK9(DHR2)-Cdc42 complex, the factors determining recognition of specific GTPases are unknown. To understand the molecular basis for DOCK-GTPase specificity, we have determined the crystal structure of DOCK2(DHR2) in complex with Rac1. DOCK2(DHR2) and DOCK9(DHR2) exhibit similar tertiary structures and homodimer interfaces and share a conserved GTPase-activating mechanism. Multiple structural differences between DOCK2(DHR2) and DOCK9(DHR2) account for their selectivity toward Rac1 and Cdc42. Key determinants of selectivity of Cdc42 and Rac for their cognate DOCK(DHR2) are a Phe or Trp residue within β3 (residue 56) and the ability of DOCK proteins to exploit differences in the GEF-induced conformational changes of switch 1 dependent on a divergent residue at position 27. DOCK proteins, therefore, differ from DH-PH GEFs that select their cognate GTPases through recognition of structural differences within the β2/β3 strands.

  5. Nonequilibrium Green's functions theory for the alpha factor of quantum cascade lasers (Conference Presentation)

    NASA Astrophysics Data System (ADS)

    Pereira, Mauro F.; Winge, David O.; Wacker, Andreas; Jumpertz, Louise; Michel, Florian; Pawlus, Robert; Elsaesser, Wolfgang E.; Schires, Kevin; Carras, Mathieu; Grillot, Frédéric

    2016-10-01

    The linewidth of a conventional laser is due to fluctuations in the laser field due to spontaneous emission and described by the Schalow-Townes formula. In addition to that, in a semiconductor laser there is a contribution arising from fluctuations in the refractive index induced by carrier density fluctuations. The later are quantitatively described by the linewidth enhancement or alpha factor [C. H. Henry, IEEE J. Quantum Electron. 18 (2), 259 (1982), W. W. Chow, S. W. Koch and M. Sargent III, Semiconductor-Laser Physics, Springer-Verlag (1994), M.F. Pereira Jr et al, J. Opt. Soc. Am. B10, 765 (1993). In this paper we investigate the alpha factor of quantum cascade lasers under actual operating conditions using the Nonequilibrium Greens Functions approach [A. Wacker et a, IEEE Journal of Sel. Top. in Quantum Electron.,19 1200611, (2013), T. Schmielau and M.F. Pereira, Appl. Phys. Lett. 95 231111, (2009)]. The simulations are compared with recent results obtained with different optical feedback techniques [L. Jumpertz et al, AIP ADVANCES 6, 015212 (2016)].

  6. Mitochondrial DNA Haplogroup R Confers a Genetic Risk Factor for Intrauterine Adhesion in Han Women Population.

    PubMed

    Zhang, Jing; Zhu, Libo; Huang, Xiufeng; Xu, Ping; Chen, Zhengyun; Huang, Qiongshi; Zhang, Xinmei

    2016-01-01

    To determine whether a specific mitochondrial DNA (mtDNA) haplogroup is implicated in the pathogenesis of intrauterine adhesion (IUA). Peripheral blood samples were collected from 486 women with (case group, n = 154) and without IUA (control group, n = 332) at the Women's Hospital, Zhejiang University School of Medicine. Genomic DNA was extracted from the blood, and the mtDNA haplogroups of Han women M, N and R were determined by sequencing hypervariable mtDNA segments and testing diagnostic polymorphisms in the mtDNA coding region. Women with mtDNA haplogroup R had an independently increased genetic risk factor for IUA with an OR 1.77 (95% CI 1.16-2.70, p = 0.009) compared with women without. Moreover, repeated intrauterine surgery within 1 month and number of intrauterine operations were both significantly associated with IUA (p < 0.001). These results suggest that mtDNA haplogroup R, one of the main mtDNA haplogroups in Han population, is a strong independent genetic risk factor for women with IUA. © 2015 S. Karger AG, Basel.

  7. The sunflower transcription factor HaWRKY76 confers drought and flood tolerance to Arabidopsis thaliana plants without yield penalty.

    PubMed

    Raineri, Jesica; Ribichich, Karina F; Chan, Raquel L

    2015-12-01

    Arabidopsis transgenic plants expressing the sunflower transcription factor HaWRKY76 exhibit increased yield and tolerance to drought and flood stresses. The genetic construct containing HaWRKY76 is proposed as a potential biotechnological tool to improve crops. Water deficit and water excess are abiotic stress factors that seriously affect crops worldwide. To increase the tolerance to such stresses without causing yield penalty constitutes a major goal for biotechnologists. In this survey, we report that HaWRKY76, a divergent sunflower WRKY transcription factor, is able to confer both dehydration and submergence tolerance to Arabidopsis transgenic plants without yield penalty. The expression pattern of HaWRKY76 was analyzed in plants grown in standard conditions and under different watering regimes indicating a regulation by water availability. The corresponding cDNA was isolated and cloned under the control of a constitutive promoter and Arabidopsis plants were transformed with this construct. These transgenic plants presented higher biomass, seed production and sucrose content than controls in standard growth conditions. Moreover, they exhibited tolerance to mild drought or flood (complete submergence/waterlogging) stresses as well as the same or increased yield, depending on the stress severity and plant developmental stage, compared with controls. Drought tolerance occurred via an ABA-independent mechanism and induction of stomatal closure. Submergence tolerance can be explained by the carbohydrate (sucrose and starch) preservation achieved through the repression of fermentation pathways. Higher cell membrane stability and chlorenchyma maintenance could be the nexus between tolerance responses in front of both stresses. Altogether, the obtained results indicated that HaWRKY76 can be a potential biotechnological tool to improve crops yield as well as drought and flood tolerances.

  8. Characterization of transcription factor gene SNAC2 conferring cold and salt tolerance in rice.

    PubMed

    Hu, Honghong; You, Jun; Fang, Yujie; Zhu, Xiaoyi; Qi, Zhuyun; Xiong, Lizhong

    2008-05-01

    Plants respond to adverse environment by initiating a series of signaling processes including activation of transcription factors that can regulate expression of arrays of genes for stress response and adaptation. NAC (NAM, ATAF, and CUC) is a plant specific transcription factor family with diverse roles in development and stress regulation. In this report, a stress-responsive NAC gene (SNAC2) isolated from upland rice IRA109 (Oryza sativa L. ssp japonica) was characterized for its role in stress tolerance. SNAC2 was proven to have transactivation and DNA-binding activities in yeast and the SNAC2-GFP fusion protein was localized in the rice nuclei. Northern blot and SNAC2 promoter activity analyses suggest that SNAC2 gene was induced by drought, salinity, cold, wounding, and abscisic acid (ABA) treatment. The SNAC2 gene was over-expressed in japonica rice Zhonghua 11 to test the effect on improving stress tolerance. More than 50% of the transgenic plants remained vigorous when all WT plants died after severe cold stress (4-8 degrees C for 5 days). The transgenic plants had higher cell membrane stability than wild type during the cold stress. The transgenic rice had significantly higher germination and growth rate than WT under high salinity conditions. Over-expression of SNAC2 can also improve the tolerance to PEG treatment. In addition, the SNAC2-overexpressing plants showed significantly increased sensitivity to ABA. DNA chip profiling analysis of transgenic plants revealed many up-regulated genes related to stress response and adaptation such as peroxidase, ornithine aminotransferase, heavy metal-associated protein, sodium/hydrogen exchanger, heat shock protein, GDSL-like lipase, and phenylalanine ammonia lyase. Interestingly, none of the up-regulated genes in the SNAC2-overexpressing plants matched the genes up-regulated in the transgenic plants over-expressing other stress responsive NAC genes reported previously. These data suggest SNAC2 is a novel stress

  9. Studies in fat grafting: Part IV. Adipose-derived stromal cell gene expression in cell-assisted lipotransfer.

    PubMed

    Garza, Rebecca M; Rennert, Robert C; Paik, Kevin J; Atashroo, David; Chung, Michael T; Duscher, Dominik; Januszyk, Michael; Gurtner, Geoffrey C; Longaker, Michael T; Wan, Derrick C

    2015-04-01

    Fat graft volume retention remains highly unpredictable, but addition of adipose-derived stromal cells to fat grafts has been shown to improve retention. The present study aimed to investigate the mechanisms involved in adipose-derived stromal cell enhancement of fat grafting. Adipose-derived stromal cells isolated from human lipoaspirate were labeled with green fluorescent protein and luciferase. Fat grafts enhanced with adipose-derived stromal cells were injected into the scalp and bioluminescent imaging was performed to follow retention of adipose-derived stromal cells within the fat graft. Fat grafts were also explanted at days 1, 5, and 10 after grafting for adipose-derived stromal cell extraction and single-cell gene analysis. Finally, CD31 immunohistochemical staining was performed on fat grafts enriched with adipose-derived stromal cells. Bioluminescent imaging demonstrated significant reduction in luciferase-positive adipose-derived stromal cells within fat grafts at 5 days after grafting. A similar reduction in viable green fluorescent protein-positive adipose-derived stromal cells retrieved from explanted grafts was also noted. Single-cell analysis revealed expression of multiple genes/markers related to cell survival and angiogenesis, including BMPR2, CD90, CD105, FGF2, CD248, TGFß1, and VEGFA. Genes involved in adipogenesis were not expressed by adipose-derived stromal cells. Finally, CD31 staining revealed significantly higher vascular density in fat grafts explanted at day 10 after grafting. Although adipose-derived stromal cell survival in the hypoxic graft environment decreases significantly over time, these cells provide multiple angiogenic growth factors. Therefore, improved fat graft volume retention with adipose-derived stromal cell enrichment may be attributable to improved graft vascularization.

  10. Induction of Hematopoietic Differentiation of Mouse Embryonic Stem Cells by an AGM-Derived Stromal Cell Line is Not Further Enhanced by Overexpression of HOXB4

    PubMed Central

    Gordon-Keylock, Sabrina A.M.; Jackson, Melany; Huang, Caoxin; Samuel, Kay; Axton, Richard A.; Oostendorp, Robert A.J.; Taylor, Helen; Wilson, Julie

    2010-01-01

    Hematopoietic differentiation of embryonic stem (ES) cells can be enhanced by co-culture with stromal cells derived from hematopoietic tissues and by overexpression of the transcription factor HOXB4. In this study, we compare the hematopoietic inductive effects of stromal cell lines derived from different subregions of the embryonic aorta-gonad-mesonephros tissue with the commonly used OP9 stromal cell line and with HOXB4 activation. We show that stromal cell lines derived from the aorta and surrounding mesenchyme (AM) act at an earlier stage of the differentiation process compared with the commonly used OP9 stromal cells. AM stromal cells were able to promote the further differentiation of isolated brachyury-GFP+ mesodermal cells into hematopoietic progenitors, whereas the OP9 stromal cells could not support the differentiation of these cells. Co-culture and analyses of individual embryoid bodies support the hypothesis that the AM stromal cell lines could enhance the de novo production of hematopoietic progenitors, lending support to the idea that AM stromal cells might act on prehematopoietic mesoderm. The induction level observed for AM stromal cells was comparable to HOXB4 activation, but no additive effect was observed when these 2 inductive strategies were combined. Addition of a γ-secretase inhibitor reduced the inductive effects of both the stromal cell line and HOXB4, providing clues to possible shared molecular mechanisms. PMID:20184433

  11. Induction of hematopoietic differentiation of mouse embryonic stem cells by an AGM-derived stromal cell line is not further enhanced by overexpression of HOXB4.

    PubMed

    Gordon-Keylock, Sabrina A M; Jackson, Melany; Huang, Caoxin; Samuel, Kay; Axton, Richard A; Oostendorp, Robert A J; Taylor, Helen; Wilson, Julie; Forrester, Lesley M

    2010-11-01

    Hematopoietic differentiation of embryonic stem (ES) cells can be enhanced by co-culture with stromal cells derived from hematopoietic tissues and by overexpression of the transcription factor HOXB4. In this study, we compare the hematopoietic inductive effects of stromal cell lines derived from different subregions of the embryonic aorta-gonad-mesonephros tissue with the commonly used OP9 stromal cell line and with HOXB4 activation. We show that stromal cell lines derived from the aorta and surrounding mesenchyme (AM) act at an earlier stage of the differentiation process compared with the commonly used OP9 stromal cells. AM stromal cells were able to promote the further differentiation of isolated brachyury-GFP(+) mesodermal cells into hematopoietic progenitors, whereas the OP9 stromal cells could not support the differentiation of these cells. Co-culture and analyses of individual embryoid bodies support the hypothesis that the AM stromal cell lines could enhance the de novo production of hematopoietic progenitors, lending support to the idea that AM stromal cells might act on prehematopoietic mesoderm. The induction level observed for AM stromal cells was comparable to HOXB4 activation, but no additive effect was observed when these 2 inductive strategies were combined. Addition of a γ-secretase inhibitor reduced the inductive effects of both the stromal cell line and HOXB4, providing clues to possible shared molecular mechanisms.

  12. Effects of Platelet-Rich Plasma & Platelet-Rich Fibrin with and without Stromal Cell-Derived Factor-1 on Repairing Full-Thickness Cartilage Defects in Knees of Rabbits

    PubMed Central

    Bahmanpour, Soghra; Ghasemi, Maryam; Sadeghi-Naini, Mohsen; Kashani, Iraj Ragerdi

    2016-01-01

    Background: The purpose of this study was to create biomaterial scaffolds like platelet-rich plasma (PRP) and platelet-rich fibrin (PRF) containing stromal cell-derived factor-1 (SDF1) as a chemokine to induce hyaline cartilage regeneration of rabbit knee in a full thickness defect. Methods: We created a full thickness defect in the trochlear groove of thirty-six bilateral knees of eighteen mature male rabbits. The knees were randomly divided into six groups (group I: untreated control, group II: PRP, group III: PRF, group IV: Gelatin+SDF1, group V: PRP+SDF1, and group VI: PRF+SDF1). After four weeks, the tissue specimens were evaluated by macroscopic examination and histological grading, immunofluorescent staining for collagen type II, and analyzed for cartilage marker genes by real-time PCR. The data were compared using statistical methods (SPSS 20, Kruskal-Wallis test, Bonferroni post hoc test and P<0.05). Results: Macroscopic evaluations revealed that international cartilage repair society (ICRS) scores of the PRF+SDF1 group were higher than other groups. Microscopic analysis showed that the ICRS score of the PRP group was significantly lower than other groups. Immunofluorescent staining for collagen II demonstrated a remarkable distribution of type II collagen in the Gel+SDF1, PRP+SDF1 and PRF+SDF1 groups compared with other groups. Real-time PCR analysis revealed that mRNA expression of SOX9 and aggrecan were significantly greater in the PRF+SDF1, PRP+SDF1, Gel+SDF1 and PRF groups than the control group (P<0.05). Conclusion: Our results indicate that implantation of PRF scaffold containing SDF1 led to the greatest evaluation scores of full-thickness lesions in rabbits. PMID:27853331

  13. Pigment epithelial-derived factor and melanoma differentiation associated gene-7 cytokine gene therapies delivered by adipose-derived stromal/mesenchymal stem cells are effective in reducing prostate cancer cell growth.

    PubMed

    Zolochevska, Olga; Yu, Gang; Gimble, Jeffrey M; Figueiredo, Marxa L

    2012-05-01

    Adipose-derived stromal/mesenchymal stem cells (ASC) have gained interest as promising tools for delivering cancer therapy. Adipose tissue can be obtained readily in amounts sufficient for ASC isolation, which can be expanded rapidly, allowing its use at low passage numbers, and can be transduced by viral and nonviral means. Our goal was to examine the potential of ASC to deliver cytokine gene therapies melanoma differentiation associated gene-7 (MDA-7) or pigment epithelial-derived factor (PEDF) to cancer cells. These novel cytokines are a potent proapoptotic and an antiangiogenesis mediator, respectively, with potential as antitumor agents. Expression of cytokine therapies did not adversely affect ASC biology, and these cells were still able to differentiate and retain normal viability. The ASC cytokine therapies were efficient in reducing tumor cell growth in coculture and also in suppressing in vitro angiogenesis phenotypes. We also observed that ASC retained their innate ability to migrate toward tumor cells in coculture, and this ability could be blocked by inhibition of CXCR4 signaling. The ASC were found to be nontumorigenic in vitro using a soft agar assay, as well as in vivo, utilizing 2 prostate cancer xenograft models. The ASC-MDA7 only reduced tumor growth in the TRAMP-C2-Ras (TC2Ras) prostate cancer model. The ASC-PEDF, however, reduced growth in both the TC2Ras and the PC3 highly aggressive prostate cancer models, and it was able to completely prevent prostate tumor establishment in vivo. In conclusion, ASC expressing PEDF and MDA7 could effectively reduce prostate tumor growth in vivo, suggesting ASC-cytokine therapies might have translational applications, especially the PEDF modality.

  14. Vascular endothelial growth factor gene-transferred bone marrow stromal cells engineered with a herpes simplex virus type 1 vector can improve neurological deficits and reduce infarction volume in rat brain ischemia.

    PubMed

    Miki, Yoshihito; Nonoguchi, Naosuke; Ikeda, Naokado; Coffin, Robert S; Kuroiwa, Toshihiko; Miyatake, Shin-ichi

    2007-09-01

    Several reports recently suggested that vascular endothelial growth factor (VEGF) may have a therapeutic benefit against experimental cerebral infarction animal models. In addition, bone marrow stromal cells (BMSCs) are known to have therapeutic potency in improving neurological deficits after occlusive cerebrovascular diseases. In the present study, we evaluated the hypothesis that intracerebral transplantation of VEGF gene-transferred BMSCs could provide a greater therapeutic effect than intracerebral transplantation of native (non-gene-transformed) BMSCs by using a transient middle cerebral artery occlusion (MCAO) rat model. Adult Wistar rats (Japan SLC, Inc., Hamamatsu, Japan) were anesthetized. VEGF gene-transferred BMSCs engineered with a replication-deficient herpes simplex virus type 1 1764/4-/pR19-hVEGF165 vector, native BMSCs, or phosphate-buffered saline were administered intracerebrally 24 hours after transient MCAO. All animals underwent behavioral testing for 28 days, and the infarction volume was determined 14 days after MCAO. The brain water contents in the ipsilateral and contralateral hemispheres of the MCAO were measured 2 and 7 days after the MCAO. Fourteen days after MCAO, immunohistochemical staining for VEGF was performed. The group receiving VEGF-modified BMSCs demonstrated significant functional recovery compared with those receiving native BMSCs. Fourteen days after the MCAO, there was a significantly lower infarct volume without aggravating cerebral edema in the group treated with VEGF gene-modified BMSCs compared with the control groups. The transplanted VEGF gene-modified BMSCs strongly expressed VEGF protein for at least 14 days. Our data suggest that the intracerebral transplantation of VEGF gene-transferred BMSCs may provide a more potent autologous cell transplantation therapy for stroke than the transplantation of native BMSCs alone.

  15. Stromal Cell-Derived Factor 1 Increases Tetrodotoxin-Resistant Sodium Currents Nav1.8 and Nav1.9 in Rat Dorsal Root Ganglion Neurons via Different Mechanisms.

    PubMed

    Qiu, Fang; Li, Yang; Fu, Qiang; Fan, Yong-Yan; Zhu, Chao; Liu, Yan-Hong; Mi, Wei-Dong

    2016-07-01

    Stromal cell-derived factor 1 (SDF-1)/chemokine CXC motif ligand 12 (CXCL12), a chemokine that is upregulated in dorsal root ganglion (DRG) during chronic pain models, has recently been found to play a central role in pain hypersensitivity. The purpose of present study is to investigate the functional impact of SDF-1 and its receptor, chemokine CXC motif receptor 4 (CXCR4), on two TTXR sodium channels in rat DRG using electrophysiological techniques. Preincubation with SDF-1 caused a concentration-dependent increase of Nav1.8 and Nav1.9 currents amplitudes in acutely isolated small diameter DRG neurons in short-term culture. As to Nav1.9, changes in current density and kinetic properties of Nav1.9 current evoked by SDF-1(50 ng/ml) was eliminated by CXCR4 antagonist AMD3100 and phosphatidylinositol 3-kinase (PI3K) inhibitor LY294002. The increase in Nav1.9 current was also blocked by pertussis toxin (PTX) but not cholera toxin (CTX), showing involvement of Gi/o but not Gs subunits. As to Nav1.8, inhibitors (AMD3100, PTX, CTX, LY294002) used in present study didn't inhibit the increased amplitude of Nav1.8 current and shifted activation curve of Nav1.8 in a hyperpolarizing direction in the presence of SDF-1 (50 ng/ml). In conclusion, our data demonstrated that SDF-1 may excite primary nociceptive sensory neurons by acting on the biophysical properties of Nav1.8 and Nav1.9 currents but via different mechanisms.

  16. Human factors in computing systems: focus on patient-centered health communication at the ACM SIGCHI conference.

    PubMed

    Wilcox, Lauren; Patel, Rupa; Chen, Yunan; Shachak, Aviv

    2013-12-01

    Health Information Technologies, such as electronic health records (EHR) and secure messaging, have already transformed interactions among patients and clinicians. In addition, technologies supporting asynchronous communication outside of clinical encounters, such as email, SMS, and patient portals, are being increasingly used for follow-up, education, and data reporting. Meanwhile, patients are increasingly adopting personal tools to track various aspects of health status and therapeutic progress, wishing to review these data with clinicians during consultations. These issues have drawn increasing interest from the human-computer interaction (HCI) community, with special focus on critical challenges in patient-centered interactions and design opportunities that can address these challenges. We saw this community presenting and interacting at the ACM SIGCHI 2013, Conference on Human Factors in Computing Systems, (also known as CHI), held April 27-May 2nd, 2013 at the Palais de Congrès de Paris in France. CHI 2013 featured many formal avenues to pursue patient-centered health communication: a well-attended workshop, tracks of original research, and a lively panel discussion. In this report, we highlight these events and the main themes we identified. We hope that it will help bring the health care communication and the HCI communities closer together.

  17. GUY1 confers complete female lethality and is a strong candidate for a male-determining factor in Anopheles stephensi

    PubMed Central

    Criscione, Frank; Qi, Yumin; Tu, Zhijian

    2016-01-01

    Despite their importance in sexual differentiation and reproduction, Y chromosome genes are rarely described because they reside in repeat-rich regions that are difficult to study. Here, we show that Guy1, a unique Y chromosome gene of a major urban malaria mosquito Anopheles stephensi, confers 100% female lethality when placed on the autosomes. We show that the small GUY1 protein (56 amino acids in length) causes female lethality and that males carrying the transgene are reproductively more competitive than their non-transgenic siblings under laboratory conditions. The GUY1 protein is a primary signal from the Y chromosome that affects embryonic development in a sex-specific manner. Our results have demonstrated, for the first time in mosquitoes, the feasibility of stable transgenic manipulation of sex ratios using an endogenous gene from the male-determining chromosome. These results provide insights into the elusive M factor and suggest exciting opportunities to reduce mosquito populations and disease transmission. DOI: http://dx.doi.org/10.7554/eLife.19281.001 PMID:27644420

  18. Guanine Nucleotide Exchange Factor OSG-1 Confers Functional Aging via Dysregulated Rho Signaling in Caenorhabditis elegans Neurons

    PubMed Central

    Duan, Zhibing; Sesti, Federico

    2015-01-01

    Rho signaling regulates a variety of biological processes, but whether it is implicated in aging remains an open question. Here we show that a guanine nucleotide exchange factor of the Dbl family, OSG-1, confers functional aging by dysregulating Rho GTPases activities in C. elegans. Thus, gene reporter analysis revealed widespread OSG-1 expression in muscle and neurons. Loss of OSG-1 gene function was not associated with developmental defects. In contrast, suppression of OSG-1 lessened loss of function (chemotaxis) in ASE sensory neurons subjected to conditions of oxidative stress generated during natural aging, by oxidative challenges, or by genetic mutations. RNAi analysis showed that OSG-1 was specific toward activation of RHO-1 GTPase signaling. RNAi further implicated actin-binding proteins ARX-3 and ARX-5, thus the actin cytoskeleton, as one of the targets of OSG-1/RHO-1 signaling. Taken together these data suggest that OSG-1 is recruited under conditions of oxidative stress, a hallmark of aging, and contributes to promote loss of neuronal function by affecting the actin cytoskeleton via altered RHO-1 activity. PMID:25527286

  19. An S-Locus Independent Pollen Factor Confers Self-Compatibility in ‘Katy’ Apricot

    PubMed Central

    Molina, Laura; Gisbert, Ana D.; Badenes, María L.; Romero, Carlos

    2013-01-01

    Loss of pollen-S function in Prunus self-compatible cultivars has been mostly associated with deletions or insertions in the S-haplotype-specific F-box (SFB) genes. However, self-compatible pollen-part mutants defective for non-S-locus factors have also been found, for instance, in the apricot (Prunus armeniaca) cv. ‘Canino’. In the present study, we report the genetic and molecular analysis of another self-compatible apricot cv. termed ‘Katy’. S-genotype of ‘Katy’ was determined as S1S2 and S-RNase PCR-typing of selfing and outcrossing populations from ‘Katy’ showed that pollen gametes bearing either the S1- or the S2-haplotype were able to overcome self-incompatibility (SI) barriers. Sequence analyses showed no SNP or indel affecting the SFB1 and SFB2 alleles from ‘Katy’ and, moreover, no evidence of pollen-S duplication was found. As a whole, the obtained results are compatible with the hypothesis that the loss-of-function of a S-locus unlinked factor gametophytically expressed in pollen (M’-locus) leads to SI breakdown in ‘Katy’. A mapping strategy based on segregation distortion loci mapped the M’-locus within an interval of 9.4 cM at the distal end of chr.3 corresponding to ∼1.29 Mb in the peach (Prunus persica) genome. Interestingly, pollen-part mutations (PPMs) causing self-compatibility (SC) in the apricot cvs. ‘Canino’ and ‘Katy’ are located within an overlapping region of ∼273 Kb in chr.3. No evidence is yet available to discern if they affect the same gene or not, but molecular markers seem to indicate that both cultivars are genetically unrelated suggesting that every PPM may have arisen independently. Further research will be necessary to reveal the precise nature of ‘Katy’ PPM, but fine-mapping already enables SC marker-assisted selection and paves the way for future positional cloning of the underlying gene. PMID:23342044

  20. An S-locus independent pollen factor confers self-compatibility in 'Katy' apricot.

    PubMed

    Zuriaga, Elena; Muñoz-Sanz, Juan V; Molina, Laura; Gisbert, Ana D; Badenes, María L; Romero, Carlos

    2013-01-01

    Loss of pollen-S function in Prunus self-compatible cultivars has been mostly associated with deletions or insertions in the S-haplotype-specific F-box (SFB) genes. However, self-compatible pollen-part mutants defective for non-S-locus factors have also been found, for instance, in the apricot (Prunus armeniaca) cv. 'Canino'. In the present study, we report the genetic and molecular analysis of another self-compatible apricot cv. termed 'Katy'. S-genotype of 'Katy' was determined as S(1)S(2) and S-RNase PCR-typing of selfing and outcrossing populations from 'Katy' showed that pollen gametes bearing either the S(1)- or the S(2)-haplotype were able to overcome self-incompatibility (SI) barriers. Sequence analyses showed no SNP or indel affecting the SFB(1) and SFB(2) alleles from 'Katy' and, moreover, no evidence of pollen-S duplication was found. As a whole, the obtained results are compatible with the hypothesis that the loss-of-function of a S-locus unlinked factor gametophytically expressed in pollen (M'-locus) leads to SI breakdown in 'Katy'. A mapping strategy based on segregation distortion loci mapped the M'-locus within an interval of 9.4 cM at the distal end of chr.3 corresponding to ∼1.29 Mb in the peach (Prunus persica) genome. Interestingly, pollen-part mutations (PPMs) causing self-compatibility (SC) in the apricot cvs. 'Canino' and 'Katy' are located within an overlapping region of ∼273 Kb in chr.3. No evidence is yet available to discern if they affect the same gene or not, but molecular markers seem to indicate that both cultivars are genetically unrelated suggesting that every PPM may have arisen independently. Further research will be necessary to reveal the precise nature of 'Katy' PPM, but fine-mapping already enables SC marker-assisted selection and paves the way for future positional cloning of the underlying gene.

  1. Physical and environmental factors affecting the persistence of explosives particles (Conference Presentation)

    NASA Astrophysics Data System (ADS)

    Papantonakis, Michael R.; Nguyen, Viet K.; Furstenberg, Robert; White, Caitlyn; Shuey, Melissa; Kendziora, Christopher A.; McGill, R. Andrew

    2017-05-01

    Knowledge of the persistence of trace explosives materials is critical to aid the security community in designing detection methods and equipment. The physical and environmental factors affecting the lifetimes of particles include temperature, airflow, interparticle distance, adlayers, humidity, particle field size and vapor pressure. We are working towards a complete particle persistence model that captures the relative importance of these effects to allow the user, with known environmental conditions, to predict particle lifetimes for explosives or other chemicals. In this work, particles of explosives are sieved onto smooth glass substrates using particle sizes and loadings relevant to those deposited by fingerprint deposition. The coupon is introduced into a custom flow cell and monitored under controlled airflow, humidity and temperature. Photomicroscopy images of the sample taken at fixed time intervals are analyzed to monitor particle sublimation and characterized as a size-independent radial sublimation velocity for each particle in the ensemble. In this paper we build on previous work by comparing the relationship between sublimation of different materials and their vapor pressures. We also describe the influence of a sebum adlayer on particle sublimation, allowing us to better model `real world' samples.

  2. A barley PHD finger transcription factor that confers male sterility by affecting tapetal development.

    PubMed

    Fernández Gómez, José; Wilson, Zoe A

    2014-08-01

    Controlling pollen development is of major commercial importance in generating hybrid crops and selective breeding, but characterized genes for male sterility in crops are rare, with no current examples in barley. However, translation of knowledge from model species is now providing opportunities to understand and manipulate such processes in economically important crops. We have used information from regulatory networks in Arabidopsis to identify and functionally characterize a barley PHD transcription factor MALE STERTILITY1 (MS1), which expresses in the anther tapetum and plays a critical role during pollen development. Comparative analysis of Arabidopsis, rice and Brachypodium genomes was used to identify conserved regions in MS1 for primer design to amplify the barley MS1 gene; RACE-PCR was subsequently used to generate the full-length sequence. This gene shows anther-specific tapetal expression, between late tetrad stage and early microspore release. HvMS1 silencing and overexpression in barley resulted in male sterility. Additionally, HvMS1 cDNA, controlled by the native Arabidopsis MS1 promoter, successfully complemented the homozygous ms1 Arabidopsis mutant. These results confirm the conservation of MS1 function in higher plants and in particular in temperate cereals. This has provided the first example of a characterized male sterility gene in barley, which presents a valuable tool for the future control of male fertility in barley for hybrid development.

  3. KIT mutations confer a distinct gene expression signature in core binding factor leukaemia.

    PubMed

    Lück, Sonja C; Russ, Annika C; Du, Juan; Gaidzik, Verena; Schlenk, Richard F; Pollack, Jonathan R; Döhner, Konstanze; Döhner, Hartmut; Bullinger, Lars

    2010-03-01

    Core binding factor (CBF) leukaemias, characterized by either inv(16)(p13.1q22) or t(8;21)(q22;q22), constitute acute myeloid leukaemia (AML) subgroups with favourable prognosis. However, 40-50% of patients relapse, emphasizing the need for risk-adapted treatment approaches. In this regard, studying secondary genetic aberrations, such as mutations of the KIT gene, is of great interest, particularly as they can be targeted by receptor tyrosine kinase inhibitors (TKI). However, so far little is known about the biology underlying KIT-mutated CBF leukaemias. We analysed gene expression profiles of 83 CBF AML cases with known KIT mutation status in order to gain novel insights in KIT-mutated CBF pathogenesis. KIT-mutated cases were characterized by deregulation of genes belonging to the NFkB signalling complex suggesting impaired control of apoptosis. Notably, a subgroup of KIT wildtype cases was also characterized by the KIT mutation signature due to yet unknown aberrations. Our data suggest that this CBF leukaemia subgroup might profit from TKI therapy, however, the relevance of the KIT mutation-associated signature remains to be validated prior to clinical implementation. Nevertheless, the existence of such a signature supports the notion of relevant biological differences in CBF leukaemia and might serve as diagnostic tool in the future.

  4. A novel insertion mutation on exon 20 of epidermal growth factor receptor, conferring resistance to erlotinib.

    PubMed

    Khan, Nawazish A; Mirshahidi, Saied; Mirshahidi, Hamid R

    2014-05-01

    The epidermal growth factor receptor (EGFR) is a transmembrane glycoprotein tyrosine kinase receptor. The small-molecule tyrosine kinase receptor inhibitors (TKIs) are in clinical use to treat non-small cell lung cancer with EGFR mutations. Variable tumor responses to erlotinib and gefitinib have been observed. The response to these TKIs varies by the type of EGFR mutations found in the tumor. The deletion on exon 19 and the L858R substitution on exon 21 constitute the most frequent mutations and are known to show good response to TKIs. However, mutations on exon 20 are less common and seem to respond poorly to TKIs. In clinical settings, the reported response of exon 20 mutations to reversible TKIs (both gefitinib and erlotinib) remains inconstant. The type of coexisting mutation seems to affect the response of these insertions to TKIs. We herein present a case of disease progression despite the use of erlotinib in a female patient who had a novel insertion mutation on exon 20. Our patient was a never-smoker and was identified to have a Pro772_His773insGlnCysPro mutation on exon 20. She had previously been treated with cisplatin and gemcitabine and then with carboplatin and pemetrexed. She was treated with erlotinib upon intolerance to second-line chemotherapy and did not respond. Our patient had a novel insertion mutation on exon 20, which was found to be resistant to erlotinib.

  5. Familial multiple gastrointestinal stromal tumours with associated abnormalities of the myenteric plexus layer and skeinoid fibres.

    PubMed

    Handra-Luca, A; Fléjou, J F; Molas, G; Sauvanet, A; Belghiti, J; Degott, C; Terris, B

    2001-10-01

    Multiple familial gastrointestinal stromal tumours are rare. We report the third family with two cases of multiple gastrointestinal stromal tumours showing skeinoid fibres. Associated abnormalities of the myenteric plexus layer are described and new hypotheses for the histogenesis of gastrointestinal stromal tumours are formulated. Multiple gastrointestinal stromal tumours developed in the duodenum and proximal jejunum were removed from mother and son. No history of a specific syndrome or of mastocytosis was known. Light microscopy revealed typical gastrointestinal stromal tumours with skeinoid fibres. An unusual abnormality of the myenteric plexus layer, showing a diffuse spindle cell hyperplasia, was noted in the macroscopically normal digestive wall. No abnormalities of the ganglion cells were associated. Tumours and the spindle cell hyperplasia showed similar morphological and immunohistochemical features with expression of CD34 and CD117 antigens. Follow-up revealed recurrences in the mother. The morphological characteristics of these two cases of familial gastrointestinal stromal tumours and of the associated abnormalities of the myenteric plexus layer, help to better explain the histogenesis of multiple familial gastrointestinal stromal tumours. The hyperplasia of the myenteric plexus could be considered a risk factor for recurrent tumours.

  6. Tumour-activated liver stromal cells regulate myeloid-derived suppressor cells accumulation in the liver.

    PubMed

    Zhang, H; He, G; Kong, Y; Chen, Y; Wang, B; Sun, X; Jia, B; Xie, X; Wang, X; Chen, D; Wei, L; Zhang, M; Zeng, H; Chen, H

    2017-04-01

    Regulating mechanisms underlying hepatic myeloid-derived suppressor cell (MDSC) accumulation remain to be described. Here, we provide evidence for the involvement of tumour-activated liver stromal cells in the process of hepatic MDSCs migration and accumulation. Our data showed an elevated frequency of MDSCs in the liver of tumour-bearing mice. Moreover, tumour-activated liver stromal cells promote MDSC migration into the liver site. Further investigation indicated higher levels of cytokine and chemokine expression in liver stromal cells after exposure to the tumour-conditioned supernatant. Notably, the expression levels of proinflammatory factors, mainly including macrophage colony stimulating factor (M-CSF), transforming growth factor-β (TGF-β), monocyte chemotactic protein-1 (MCP-1) and stromal-derived factor-1 (SDF-1), increased after treatment with tumour-conditioned supernatant, and blockade of MCP-1 or SDF-1 decreased the proportion of tumour infiltrated MDSCs in mice co-transplanted with liver stromal cells and tumour cells, but not in mice with only tumour cells injection. These findings demonstrate that tumour-activated liver stromal cells produce higher levels of chemokines and cytokines, which may contribute to MDSC accumulation into the liver site in patients with liver cancer. © 2017 British Society for Immunology.

  7. Transcription factor StWRKY1 regulates phenylpropanoid metabolites conferring late blight resistance in potato

    PubMed Central

    Yogendra, Kalenahalli N.; Kumar, Arun; Sarkar, Kobir; Li, Yunliang; Pushpa, Doddaraju; Mosa, Kareem A.; Duggavathi, Raj; Kushalappa, Ajjamada C.

    2015-01-01

    Quantitative resistance is polygenically controlled and durable, but the underlying molecular and biochemical mechanisms are poorly understood. Secondary cell wall thickening is a critical process in quantitative resistance, regulated by transcriptional networks. This paper provides compelling evidence on the functionality of StWRKY1 transcription factor, in a compatible interaction of potato–Phytophthora infestans, to extend our knowledge on the regulation of the metabolic pathway genes leading to strengthening the secondary cell wall. A metabolomics approach was used to identify resistance-related metabolites belonging to the phenylpropanoid pathway and their biosynthetic genes regulated by StWRKY1. The StWRKY1 gene in resistant potato was silenced to decipher its role in the regulation of phenylpropanoid pathway genes to strengthen the secondary cell wall. Sequencing of the promoter region of StWRKY1 in susceptible genotypes revealed the absence of heat shock elements (HSEs). Simultaneous induction of both the heat shock protein (sHSP17.8) and StWRKY1 following pathogen invasion enables functioning of the latter to interact with the HSE present in the resistant StWRKY1 promoter region. EMSA and luciferase transient expression assays further revealed direct binding of StWRKY1 to promoters of hydroxycinnamic acid amide (HCAA) biosynthetic genes encoding 4-coumarate:CoA ligase and tyramine hydroxycinnamoyl transferase. Silencing of the StWRKY1 gene was associated with signs of reduced late blight resistance by significantly increasing the pathogen biomass and decreasing the abundance of HCAAs. This study provides convincing evidence on the role of StWRKY1 in the regulation of downstream genes to biosynthesize HCAAs, which are deposited to reinforce secondary cell walls. PMID:26417019

  8. Co-Factor Binding Confers Substrate Specificity to Xylose Reductase from Debaryomyces hansenii

    PubMed Central

    Singh, Appu Kumar; Mondal, Alok K.; Kumaran, S.

    2012-01-01

    Binding of substrates into the active site, often through complementarity of shapes and charges, is central to the specificity of an enzyme. In many cases, substrate binding induces conformational changes in the active site, promoting specific interactions between them. In contrast, non-substrates either fail to bind or do not induce the requisite conformational changes upon binding and thus no catalysis occurs. In principle, both lock and key and induced-fit binding can provide specific interactions between the substrate and the enzyme. In this study, we present an interesting case where cofactor binding pre-tunes the active site geometry to recognize only the cognate substrates. We illustrate this principle by studying the substrate binding and kinetic properties of Xylose Reductase from Debaryomyces hansenii (DhXR), an AKR family enzyme which catalyzes the reduction of carbonyl substrates using NADPH as co-factor. DhXR reduces D-xylose with increased specificity and shows no activity towards “non-substrate” sugars like L-rhamnose. Interestingly, apo-DhXR binds to D-xylose and L-rhamnose with similar affinity (Kd∼5.0–10.0 mM). Crystal structure of apo-DhXR-rhamnose complex shows that L-rhamnose is bound to the active site cavity. L-rhamnose does not bind to holo-DhXR complex and thus, it cannot competitively inhibit D-xylose binding and catalysis even at 4–5 fold molar excess. Comparison of Kd values with Km values reveals that increased specificity for D-xylose is achieved at the cost of moderately reduced affinity. The present work reveals a latent regulatory role for cofactor binding which was previously unknown and suggests that cofactor induced conformational changes may increase the complimentarity between D-xylose and active site similar to specificity achieved through induced-fit mechanism. PMID:23049810

  9. Co-factor binding confers substrate specificity to xylose reductase from Debaryomyces hansenii.

    PubMed

    Biswas, Dipanwita; Pandya, Vaibhav; Singh, Appu Kumar; Mondal, Alok K; Kumaran, S

    2012-01-01

    Binding of substrates into the active site, often through complementarity of shapes and charges, is central to the specificity of an enzyme. In many cases, substrate binding induces conformational changes in the active site, promoting specific interactions between them. In contrast, non-substrates either fail to bind or do not induce the requisite conformational changes upon binding and thus no catalysis occurs. In principle, both lock and key and induced-fit binding can provide specific interactions between the substrate and the enzyme. In this study, we present an interesting case where cofactor binding pre-tunes the active site geometry to recognize only the cognate substrates. We illustrate this principle by studying the substrate binding and kinetic properties of Xylose Reductase from Debaryomyces hansenii (DhXR), an AKR family enzyme which catalyzes the reduction of carbonyl substrates using NADPH as co-factor. DhXR reduces D-xylose with increased specificity and shows no activity towards "non-substrate" sugars like L-rhamnose. Interestingly, apo-DhXR binds to D-xylose and L-rhamnose with similar affinity (K(d)∼5.0-10.0 mM). Crystal structure of apo-DhXR-rhamnose complex shows that L-rhamnose is bound to the active site cavity. L-rhamnose does not bind to holo-DhXR complex and thus, it cannot competitively inhibit D-xylose binding and catalysis even at 4-5 fold molar excess. Comparison of K(d) values with K(m) values reveals that increased specificity for D-xylose is achieved at the cost of moderately reduced affinity. The present work reveals a latent regulatory role for cofactor binding which was previously unknown and suggests that cofactor induced conformational changes may increase the complimentarity between D-xylose and active site similar to specificity achieved through induced-fit mechanism.

  10. Transcription factor StWRKY1 regulates phenylpropanoid metabolites conferring late blight resistance in potato.

    PubMed

    Yogendra, Kalenahalli N; Kumar, Arun; Sarkar, Kobir; Li, Yunliang; Pushpa, Doddaraju; Mosa, Kareem A; Duggavathi, Raj; Kushalappa, Ajjamada C

    2015-12-01

    Quantitative resistance is polygenically controlled and durable, but the underlying molecular and biochemical mechanisms are poorly understood. Secondary cell wall thickening is a critical process in quantitative resistance, regulated by transcriptional networks. This paper provides compelling evidence on the functionality of StWRKY1 transcription factor, in a compatible interaction of potato-Phytophthora infestans, to extend our knowledge on the regulation of the metabolic pathway genes leading to strengthening the secondary cell wall. A metabolomics approach was used to identify resistance-related metabolites belonging to the phenylpropanoid pathway and their biosynthetic genes regulated by StWRKY1. The StWRKY1 gene in resistant potato was silenced to decipher its role in the regulation of phenylpropanoid pathway genes to strengthen the secondary cell wall. Sequencing of the promoter region of StWRKY1 in susceptible genotypes revealed the absence of heat shock elements (HSEs). Simultaneous induction of both the heat shock protein (sHSP17.8) and StWRKY1 following pathogen invasion enables functioning of the latter to interact with the HSE present in the resistant StWRKY1 promoter region. EMSA and luciferase transient expression assays further revealed direct binding of StWRKY1 to promoters of hydroxycinnamic acid amide (HCAA) biosynthetic genes encoding 4-coumarate:CoA ligase and tyramine hydroxycinnamoyl transferase. Silencing of the StWRKY1 gene was associated with signs of reduced late blight resistance by significantly increasing the pathogen biomass and decreasing the abundance of HCAAs. This study provides convincing evidence on the role of StWRKY1 in the regulation of downstream genes to biosynthesize HCAAs, which are deposited to reinforce secondary cell walls.

  11. Somatic Hypermutations Confer Rheumatoid Factor Activity in Hepatitis C Virus–Associated Mixed Cryoglobulinemia

    PubMed Central

    Charles, Edgar D.; Orloff, Michael I. M.; Nishiuchi, Eiko; Marukian, Svetlana; Rice, Charles M.; Dustin, Lynn B.

    2014-01-01

    Objective Hepatitis C virus (HCV) is the most frequent cause of mixed cryoglobulinemia (MC), which is characterized by endothelial deposition of rheumatoid factor (RF)–containing immune complexes and end-organ vasculitis. MC is a lymphoproliferative disorder in which B cells express RF-like Ig, yet its precise antigenic stimulus is unknown. We have proposed that IgG–HCV immune complexes stimulate B cell expansion and somatic hypermutation (SHM)–induced affinity maturation in part via engagement of an RF-like B cell receptor. This study was undertaken to test the hypothesis that SHM augments RF activity. Methods RFs cloned from single B cells from 4 patients with HCV-associated MC (HCV-MC) were expressed as IgM, IgG, or IgG Fab. Selected Ig were reverted to germline. RF activity of somatically mutated Ig and germline-reverted Ig was determined by enzyme-linked immunosorbent assay. Results Ig with SHM had RF activity, with the preference for binding being highest for IgG1, followed by IgG2 and IgG4, and lowest for IgG3, where there was no detectable binding. In contrast, reverted germline IgG exhibited markedly diminished RF activity. Competition with 1 μg/ml of protein A abrogated RF activity, suggesting specificity for IgG Fc. Swapping of mutated heavy-chain pairs and light-chain pairs also abrogated RF activity, suggesting that context-specific pairing of appropriate IgH and Igκ, in addition to SHM, is necessary for RF activity. Conclusion SHM significantly contributes to RF activity in HCV-MC patients, suggesting that autoreactivity in these patients arises through antigen-dependent SHM, as opposed to nondeletion of autore-active germline Ig. PMID:23754128

  12. Factor XII full and partial null in rat confers robust antithrombotic efficacy with no bleeding.

    PubMed

    Cai, Tian-Quan; Wu, Weizhen; Shin, Myung K; Xu, Yiming; Jochnowitz, Nina; Zhou, Yuchen; Hoos, Lizbeth; Bentley, Ross; Strapps, Walter; Thankappan, Anil; Metzger, Joseph M; Ogletree, Martin L; Tadin-Strapps, Marija; Seiffert, Dietmar A; Chen, Zhu

    2015-12-01

    This report aims at exploring quantitatively the relationship between FXII inhibition and thromboprotection. FXII full and partial null in rats were established via zinc finger nuclease-mediated knockout and siRNA-mediated knockdown, respectively. The rats were subsequently characterized in thrombosis and hemostasis models. Knockout rats exhibited complete thromboprotection in both the arteriovenous shunt model (∼100% clot weight reduction) and the FeCl3-induced arterial thrombosis model (no reduction in blood flow), without any increase in cuticle bleeding time compared with wild-type control rats. Ex-vivo aPTT and the ellagic acid-triggered thrombin generation assay (TGA) exhibited anticoagulant changes. In contrast, ex-vivo PT or high tissue factor-triggered TGA was indistinguishable from control. Rats receiving single doses (0, 0.01, 0.03, 0.1, 0.3, 1 mg/kg) of FXII siRNA exhibited dose-dependent knockdown in liver FXII mRNA and plasma FXII protein (95 and 99%, respectively, at 1 mg/kg) at day 7 post dosing. FXII knockdown was associated with dose-dependent thromboprotection (maximal efficacy achieved with 1 mg/kg in both models) and negligible change in cuticle bleeding times. Ex-vivo TGA triggered with low-level (0.5 μmol/l) ellagic acid tracked best with the knockdown levels and efficacy. Our findings confirm and extend literature reports of an attractive benefit-to-risk profile of targeting FXII for antithrombotic therapies. Titrating of FXII is instructive for its pharmacological inhibition. The knockout rat is valuable for evaluating both mechanism-based safety concerns and off-target effects of FXII(a) inhibitors. Detailed TGA analyses will inform on optimal trigger conditions in studying pharmacodynamic effects of FXII(a) inhibition.

  13. Identification of a cis-acting regulatory element conferring inducibility of the atrial natriuretic factor gene in acute pressure overload.

    PubMed Central

    von Harsdorf, R; Edwards, J G; Shen, Y T; Kudej, R K; Dietz, R; Leinwand, L A; Nadal-Ginard, B; Vatner, S F

    1997-01-01

    To identify the cis-acting regulatory element(s) which control the induction of the atrial natriuretic factor (ANF) gene in acute pressure overload, DNA constructs consisting of promoter elements linked to a reporter gene were injected into the myocardium of dogs, which underwent aortic banding or were sham-operated. Expression of a reporter gene construct harboring the ANF promoter (-3400ANF) was induced 6-12-fold after 7 d of pressure overload. An internal deletion of 556 bp (nucleotide sequence -693 to -137) completely abrogated the inducibility of the ANF reporter gene construct. An activator protein-1 (AP1)-like site (-496 to -489) and a cAMP regulatory element (CRE) (-602 to -596) are located within the deleted sequence. Site-directed mutagenesis of the AP1-like site but not the CRE completely prevented the induction of this construct to acute pressure overload. Further, the AP1-like site was able to confer inducibility of a heterologous promoter (beta-myosin heavy chain) to higher values than controls. Gel mobility shift assay (GMSA) supershift analysis was performed using a radiolabeled probe of the ANF promoter (-506/-483) that included the AP1-like site (ATGAATCA) sequence, as well as a probe converted to contain an AP1 consensus sequence (ATGACTCA). GMSA analysis demonstrated that the ANF AP1-like element could bind both a constitutively expressed factor and the AP1 proteins, and conversion to a true AP1 site increased its affinity for AP1. However, 7 d after the onset of pressure overload, the AP1 proteins were present only at low levels, and the major complex formed by the ANF AP1-like probe was not supershifted by a jun antibody. Using a large animal model of pressure overload, we have demonstrated that a unique cis-acting element was primarily responsible for the overload induction of the ANF gene. PMID:9276748

  14. Gastrointestinal Stromal Tumor – An Evolving Concept

    PubMed Central

    Tornillo, Luigi

    2014-01-01

    Gastrointestinal stromal tumors (GISTs) are the most frequent mesenchymal tumors of the gastrointestinal tract. The discovery that these tumors, formerly thought of smooth muscle origin, are indeed better characterized by specific activating mutation in genes coding for the receptor tyrosine kinases (RTKs) CKIT and PDGFRA and that these mutations are strongly predictive for the response to targeted therapy with RTK inhibitors has made GISTs the typical example of the integration of basic molecular knowledge in the daily clinical activity. The information on the mutational status of these tumors is essential to predict (and subsequently to plan) the therapy. As resistant cases are frequently wild type, other possible oncogenic events, defining other “entities,” have been discovered (e.g., succinil dehydrogenase mutation/dysregulation, insuline growth factor expression, and mutations in the RAS-RAF-MAPK pathway). The classification of disease must nowadays rely on the integration of the clinico-morphological characteristics with the molecular data. PMID:25593916

  15. Estrogen-induced angiogenic factors derived from stromal and cancer cells are differently regulated by enterolactone and genistein in human breast cancer in vivo.

    PubMed

    Saarinen, Niina M; Abrahamsson, Annelie; Dabrosin, Charlotta

    2010-08-01

    Angiogenesis is a key in cancer progression and its regulators are released both by the tumor cells and the stroma. Dietary phytoestrogens, such as the lignan enterolactone (ENL) and the isoflavone genistein (GEN), may differently affect breast cancer growth. In this study, human breast cancer cells (MCF-7) were established in mice creating a tumor with species-specific cancer and stroma cells. Ovariectomized athymic mice supplemented with estradiol (E2) were fed basal AIN-93G diet (BD) or BD supplemented with 100 mg/kg ENL, 100 mg/kg GEN or their combination (ENL+GEN). We show that ENL and ENL+GEN inhibited E2-induced cancer growth and angiogenesis, whereas GEN alone did not. Microdialysis was used to sample extracellular proteins in tumors in vivo. ENL and ENL+GEN decreased both stroma- and cancer cell-derived VEGF, whereas cancer cell-derived PlGF increased. In subcutaneous Matrigel plugs in mice, ENL and ENL+GEN decreased E2-induced endothelial cell infiltration, whereas GEN alone did not. In endothelial cells, ENL inhibited E2-induced VEGFR-2 expression, whereas GEN did not. These results suggest that ENL has potent effects on breast cancer growth, even in combination with GEN, by downregulating E2-stimulated angiogenic factors derived both from the stroma and the cancer cells, whereas dietary GEN does not possess any antiestrogenic effects.

  16. Tumor cell and connective tissue cell interactions in human colorectal adenocarcinoma. Transfer of platelet-derived growth factor-AB/BB to stromal cells.

    PubMed Central

    Sundberg, C.; Branting, M.; Gerdin, B.; Rubin, K.

    1997-01-01

    Mechanisms underlying stimulation of platelet-derived growth factor (PDGF) beta-receptors expressed on connective tissue cells in human colorectal adenocarcinoma were investigated in this study. PDGF-AB/BB, but not PDGF receptors, was expressed by tumor cells in situ, as well as in tumor cell isolates of low passage from human colorectal adenocarcinoma. In an experimental co-culture system, conditioned medium from tumor cells only marginally activated PDGF beta-receptors expressed on fibroblasts. In contrast, co-culturing of the two cell types led to a marked PDGF beta-receptor activation. Functional PDGF-AB/BB was found to be associated with heparinase-I-sensitive components on the tumor cell surface. PDGF-AB/BB, isolated from heparinase-I-sensitive cell surface components, induced a marked activation of PDGF beta-receptors. Furthermore, co-culturing tumor cells together with fibroblasts led to a sustained activation of PDGF beta-receptors expressed on fibroblasts. Double immunofluorescence staining of tissue sections from human colorectal adenocarcinoma, combined with computer-aided image analysis, revealed that nonproliferating tumor cells were the predominant cellular source of PDGF-AB/BB in the tumor stroma. In addition, PDGF-AB/BB-expressing tumor cells were found juxtapositioned to microvascular cells expressing activated PDGF beta-receptors. Confocal microscopy revealed a cytoplasmic and cell-membrane-associated expression of PDGF-AB/BB in tumor cells situated in the stroma. In contrast, epithelial cells situated in normal or tumorous acinar structures revealed only a cell-membrane-associated PDGF-AB/BB expression. The is vitro and in situ results demonstrate that tumor cells not only facilitate but also have the ability to modulate connective tissue cell responsiveness to PDGF-AB/BB in a paracrine fashion, through direct cell-cell interactions in human colorectal adenocarcinoma. Images Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 Figure 6 PMID:9250160

  17. Progesterone reduces the migration of mast cells toward the chemokine stromal cell-derived factor-1/CXCL12 with an accompanying decrease in CXCR4 receptors.

    PubMed

    Belot, Marie-Pierre; Abdennebi-Najar, Latifa; Gaudin, Françoise; Lieberherr, Michèle; Godot, Véronique; Taïeb, Joelle; Emilie, Dominique; Machelon, Véronique

    2007-05-01

    Mast cell recruitment is implicated in many physiological functions and several diseases. It depends on microenvironmental factors, including hormones. We have investigated the effect of progesterone on the migration of HMC-1(560) mast cells toward CXCL12, a chemokine that controls the migration of mast cells into tissues. HMC-1(560) mast cells were incubated with 1 nM to 1 microM progesterone for 24 h. Controls were run without progesterone. Cell migration toward CXCL12 was monitored with an in vitro assay, and statistical analysis of repeated experiments revealed that progesterone significantly reduced cell migration without increasing the number of apoptotic cells (P = 0.0084, n = 7). Differences between progesterone-treated and untreated cells were significant at 1 microM (P < 0.01, n = 7). Cells incubated with 1 microM progesterone showed no rearrangment of actin filaments in response to CXCL12. Progesterone also reduced the calcium response to CXCL12 and Akt phosphorylation. Cells incubated with progesterone had one-half the control concentrations of CXCR4 (mRNA, total protein, and membrane-bound protein). Progesterone also inhibited the migration of HMC-1(560) cells transfected with hPR-B-pSG5 plasmid, which contained 2.5 times as much PR-B as the control. These transfected cells responded differently (P < 0.05, n = 5) from untreated cells to 1 nM progesterone. We conclude that progesterone reduces mast cell migration toward CXCL12 and that CXCR4 may be a progesterone target in mast cells.

  18. Computational Protein Design to Re-Engineer Stromal Cell-Derived Factor-1α (SDF) Generates an Effective and Translatable Angiogenic Polypeptide Analog

    PubMed Central

    Hiesinger, William; Perez-Aguilar, Jose Manuel; Atluri, Pavan; Marotta, Nicole A.; Frederick, John R.; Fitzpatrick, J. Raymond; McCormick, Ryan C.; Muenzer, Jeffrey R.; Yang, Elaine C.; Levit, Rebecca D.; Yuan, Li-Jun; MacArthur, John W.; Saven, Jeffery G.; Woo, Y. Joseph

    2014-01-01

    BACKGROUND After ischemic injury, cardiac secretion of the potent endothelial progenitor stem cell (EPC) chemokine SDF stimulates endogenous neovascularization and myocardial repair, a process insufficiently robust to repair major infarcts. Experimentally, exogenous administration of recombinant SDF enhances neovasculogenesis and cardiac function after MI. However, SDF has a short half-life, is bulky, and very expensive. Smaller analogs of SDF may provide translational advantages including enhanced stability and function, ease of synthesis, lower cost, and potential modulated delivery via engineered biomaterials. In this study, computational protein design was used to create a more efficient evolution of the native SDF protein. METHODS and RESULTS Protein structure model was used to engineer an SDF polypeptide analog (ESA) that splices the N-terminus (activation and binding) and C-terminus (extracellular stabilization) with a diproline segment designed to limit the conformational flexibility of the peptide backbone and retain the relative orientation of these segments observed in the native structure of SDF. EPCs in ESA gradient, assayed by Boyden chamber, showed significantly increased migration compared to both SDF and control gradients (ESA 567±74 cells/HPF vs SDF 438±46 p=0.037; vs Control 156±45 p=0.001). EPC receptor activation was evaluated by quantifying phosphorylated AKT. ESA had significantly greater pAKT levels than SDF and control (1.64±0.24 vs 1.26±0.187, p=0.01; vs. 0.95±0.08, p<0.001). Angiogenic growth factor assays revealed a distinct increase in Angiopoietin-1 expression in the ESA and SDF treated hearts. Also, CD-1 mice (n=30) underwent LAD ligation and peri-infarct intramyocardial injection of ESA, SDF-1α, or saline. At 2 weeks, echocardiography demonstrated a significant gain in EF, CO, SV, and Fractional Area Change (FAC) in mice treated with ESA when compared to controls and significant improvement in FAC when compared to SDF treated

  19. Heterologous expression of pneumococcal virulence factor PspC on the surface of Lactococcus lactis confers adhesive properties.

    PubMed

    Asmat, Tauseef M; Klingbeil, Katharina; Jensch, Inga; Burchhardt, Gerhard; Hammerschmidt, Sven

    2012-03-01

    Lactococcus lactis is a non-pathogenic bacterium that is used in the food industry but is also used as a heterologous host to reveal protein functions of pathogenic bacteria. The adhesin PspC from Streptococcus pneumoniae is a choline-binding protein that is non-covalently anchored to the bacterial cell wall. To assess the exclusive impact of pneumococcal surface protein C (PspC) on the interplay with its host we generated recombinant L. lactis producing a nisin-inducible and covalently anchored variant of PspC on the lactococcal cell surface. A translational fusion of the 5'-end of pspC3.4 with the 3'-end of hic (pspC11.4) was designed to decorate the surface of L. lactis with a chimeric PspC. The PspC3.4 part comprises the first 281 aa residues of PspC3.4, while the Hic sequence consists of the proline-rich and sortase-anchored domain. The results demonstrated that PspC is sufficient for adhesion and subsequent invasion of host epithelial cells expressing the human polymeric Ig receptor (hpIgR). Moreover, invasion via hpIgR was even more pronounced when the chimeric PspC was produced by lactococci compared with pneumococci. This study shows also for the first time that PspC plays no significant role during phagocytosis by macrophages. In contrast, recruitment of Factor H via the PspC chimer has a dramatic effect on phagocytosis of recombinant but not wild-type lactococci, as Factor H interacts specifically with the amino-terminal part of PspC and mediates the contact with phagocytes. Furthermore, L. lactis expressing PspC increased intracellular calcium levels in pIgR-expressing epithelial cells, thus resembling the effect of pneumococci, which induced release of Ca(2+) from intracellular stores via the PspC-pIgR mechanism. In conclusion, expression of the chimeric PspC confers adhesive properties to L. lactis and indicates the potential of L. lactis as a suitable host to study the impact of individual bacterial factors on their capacity to interfere with the

  20. Impact of local injection of brain-derived neurotrophic factor-expressing mesenchymal stromal cells (MSCs) combined with intravenous MSC delivery in a canine model of chronic spinal cord injury.

    PubMed

    Lee, Seung Hoon; Kim, Yongsun; Rhew, Daeun; Kim, Ahyoung; Jo, Kwang Rae; Yoon, Yongseok; Choi, Kyeung Uk; Jung, Taeseong; Kim, Wan Hee; Kweon, Oh-Kyeong

    2016-10-28

    The microenvironment of the chronically injured spinal cord does not allow for axonal regeneration due to glial scarring. To ameliorate this, several therapeutic strategies have been used. We investigated whether combined transplantation of chondroitinase ABC (chABC) and mesenchymal stromal cells (MSCs) genetically modified to secrete brain-derived neurotrophic factor (BDNF) with intravenous (IV) administration of MSCs can promote recovery of hindlimb function after chronic spinal cord injury (SCI). Canine BDNF-expressing MSCs were generated using a lentivirus packaging protocol. Twelve beagle dogs with experimentally induced chronic SCI were divided into chABC/MSC-green fluorescent protein (GFP), chABC/MSC-BDNF and chABC/MSC-BDNF/IV groups. The MSCs (1 × 10(7) cells) and chABC were transplanted 3 weeks after SCI in all groups, and IV injection of MSC-GFP (1 × 10(7) cells) was performed 1 and 2 weeks after MSC transplantation in the chABC/MSC-BDNF/IV group. Spinal cords were harvested 8 weeks after transplantation. The dogs in the chABC/MSC-BDNF included groups had significantly improved functional recovery 8 weeks after transplantation compared with those in the chABC/MSC-GFP group. The animals in the chABC/MSC-BDNF/IV group showed significant improvements in functional recovery at 6, 7 and 8 weeks compared with those in the chABC/MSC-BDNF group. Fibrotic changes were significantly decreased in the chABC/MSC-BDNF/IV group. We also observed significant decreases in the expression levels of tumor necrosis factor-α, interleukin-6, COX-2, glial fibrillary acidic protein and GalC and increased expression levels of BDNF, β3-tubulin neurofilament medium, and nestin in the chABC/MSC-BDNF/IV group. We suggest that transplantation of combined chABC and BDNF-expressing MSCs, along with IV injection of MSCs, is the optimal therapy for chronic SCI. Copyright © 2016 International Society for Cellular Therapy. Published by Elsevier Inc. All rights reserved.

  1. Thymic Stromal Lymphopoietin Is Up-Regulated in the Skin of Patients With Systemic Sclerosis and Induces Profibrotic Genes and Intracellular Signaling That Overlap With Those Induced by Interleukin-13 and Transforming Growth Factor β

    PubMed Central

    Christmann, Romy B.; Mathes, Allison; Affandi, Alsya J.; Padilla, Cristina; Nazari, Banafsheh; Bujor, Andreea M.; Stifano, Giuseppina; Lafyatis, Robert

    2015-01-01

    Objective To explore the expression of thymic stromal lymphopoietin (TSLP) in patients with diffuse cutaneous systemic sclerosis (dcSSc) and compare its effects in vivo and in vitro with those of interleukin-13 (IL-13) and transforming growth factor β (TGFβ). Methods Skin biopsy specimens from patients with dcSSc (n = 14) and healthy controls (n = 13) were analyzed by immunohistochemistry and immunofluorescence for TSLP, TSLP receptor, CD4, CD8, CD31, and CD163 markers. Wild-type, IL-4Rα1–, and TSLP-deficient mice were treated with TGFβ, IL-13, poly(I-C), or TSLP by osmotic pump. Human fibroblasts and peripheral blood mononuclear cells (PBMCs) were stimulated with TGFβ, IL-13, poly(I-C), or TSLP. Microarray analysis and quantitative polymerase chain reaction were performed to determine gene expression, and protein levels of phospho-Smad2 and macrophage marker CD163 were tested. Results TSLP was highly expressed in the skin of dcSSc patients, more strongly in perivascular areas and in immune cells, and was produced mainly by CD163+ cells. The skin of TSLP-treated mice showed up-regulated clusters of gene expression that overlapped strongly with those in IL-13– and TGFβ-treated mice. TSLP up-regulated specific genes, including CXCL9, proteasome, and interferon (IFN)–regulated genes. TSLP treatment in IL-4Rα1–deficient mice promoted similar cutaneous inflammation as in wild-type mice, though TSLP-induced arginase 1, CCL2, and matrix metalloproteinase 12 messenger RNA levels were blocked. In PBMCs, TSLP up-regulated tumor necrosis factor α, Mx-1, IFNγ, CXCL9, and mannose receptor 1 gene expression. TSLP-deficient mice treated with TGFβ showed less fibrosis and blocked expression of plasminogen activator inhibitor 1 and osteopontin 1. Poly(I-C)–treated mice showed high levels of cutaneous TSLP. Conclusion TSLP is highly expressed in the skin of dcSSc patients and interacts in a complex manner with 2 other profibrotic cytokines, TGFβ and IL-13

  2. Human corneal fibroblast migration and extracellular matrix synthesis during stromal repair: Role played by platelet-derived growth factor-BB, basic fibroblast growth factor, and transforming growth factor-β1.

    PubMed

    Gallego-Muñoz, Patricia; Ibares-Frías, Lucía; Garrote, José A; Valsero-Blanco, María Cruz; Cantalapiedra-Rodríguez, Roberto; Merayo-Lloves, Jesús; Carmen Martínez-García, M

    2016-11-15

    The development of treatments that modulate corneal wound healing to avoid fibrosis during tissue repair is important for the restoration of corneal transparency after an injury. To date, few studies have studied the influence of growth factors (GFs) on human corneal fibroblast (HCF) expression of extracellular matrix (ECM) proteins such as collagen types I and III, proteoglycans such as perlecan, or proteins implicated in cellular migration such as α5β1-integrin and syndecan-4. Using in vitro HCFs, a mechanical wound model was developed to study the influence of the GFs basic fibroblast GF (bFGF), platelet-derived GF (PDGF-BB) and transforming GF-β1 (TGFβ1) on ECM protein production and cellular migration. Our results show that mechanical wounding provokes the autocrine release of bFGF and TGFβ1 at different time points during the wound closure. The HCF response to PDGF-BB was a rapid closure due to fast cellular migration associated with a high focal adhesion replacement and a high expression of collagen and proteoglycans, producing nonfibrotic healing. bFGF stimulated nonfibrotic ECM production and limited the migration process. Finally, TGFβ1 induced expression of the fibrotic markers collagen type III and α5β1 integrin, and it inhibited cellular migration due to the formation of focal adhesions with a low turnover rate. The novel in vitro HCF mechanical wound model can be used to understand the role played by GFs in human corneal repair. The model can also be used to test the effects of different treatments aimed at improving the healing process. Copyright © 2016 John Wiley & Sons, Ltd. Copyright © 2016 John Wiley & Sons, Ltd.

  3. Angiogenic Potential of Multipotent Stromal Cells from the Umbilical Cord: an In Vitro Study.

    PubMed

    Arutyunyan, I V; Kananykhina, E Yu; Fatkhudinov, T Kh; El'chaninov, A V; Makarov, A V; Raimova, E Sh; Bol'shakova, G B; Sukhikh, G T

    2016-05-01

    The mechanisms of proangiogenic activity of multipotent stromal cells from human umbilical cord were analyzed in vitro. The absence of secreted forms of proangiogenic growth factor VEGF-A in the culture medium conditioned by umbilical cord-derived multipotent stromal cells was shown by ELISA. However, the possibility of paracrine stimulation of cell proliferation, mobility, and directed migration of endothelial EA.hy926 cells was demonstrated by using MTT test, Transwell system, and monolayer wound modeling. The capacity of multipotent stromal cells to acquire the phenotype of endothelium-like cells was analyzed using differentiation media of three types. It was found that VEGF-A is an essential but not sufficient inductor of differentiation of umbilical cord-derived multipotent stromal cells into CD31+ cells.

  4. High Stromal Carbonic Anhydrase IX Expression Is Associated With Decreased Survival in p16-Negative Head-and-Neck Tumors

    SciTech Connect

    Brockton, Nigel; Dort, Joseph; Lau, Harold; Hao, Desiree; Brar, Sony; Klimowicz, Alexander; Petrillo, Stephanie; Diaz, Roman; Doll, Corinne; Magliocco, Anthony

    2011-05-01

    Purpose: Head-and-neck squamous cell carcinoma (HNSCC) is the fifth most common malignancy worldwide. Alcohol use and tobacco use are the most established risk factors; however, human papilloma virus (HPV) infection is a major risk factor for a subset of HNSCCs. Although HPV-positive tumors typically present at a more advanced stage at diagnosis, they are associated with a better prognosis. Tumor hypoxia confers poor prognosis and treatment failure, but direct tumor oxygen measurement is challenging. Endogenous markers of hypoxia (EMHs) have been proposed but have not replicated the prognostic utility of direct oxygen measurement. The expression of endogenous markers of hypoxia may be influenced by oxygen-independent factors, such as the HPV status of the tumor. Methods and Materials: Consecutive cases of locally advanced HNSCC, treated with a uniform regimen of combined radiotherapy and chemotherapy, were identified. Tissue microarrays were assembled from triplicate 0.6-mm cores of archived tumor tissue. HPV status was inferred from semiquantitative p16 immunostaining and directly measured by use of HPV-specific chromogenic in situ hybridization and polymerase chain reaction. Automated quantitative fluorescent immunohistochemistry was conducted to measure epithelial and stromal expression of carbonic anhydrase IX (CAIX) and glucose transporter 1 (GLUT1). Results: High stromal CAIX expression was associated with significantly reduced overall survival (p = 0.03) in patients with p16-negative tumors. Conclusions: This is the first study to use quantitative immunohistochemistry to examine endogenous markers of hypoxia stratified by tumor p16/HPV status. Assessment of CAIX expression in p16-negative HNSCC could identify patients with the least favorable prognosis and inform therapeutic strategies.

  5. IFN type I and II induce BAFF secretion from human decidual stromal cells

    PubMed Central

    Lundell, Anna-Carin; Nordström, Inger; Andersson, Kerstin; Lundqvist, Christina; Telemo, Esbjörn; Nava, Silvia; Kaipe, Helen; Rudin, Anna

    2017-01-01

    B cell activating factor (BAFF) is a critical cytokine for maturation of immature B cells. In murine lymph nodes, BAFF is mainly produced by podoplanin-expressing stromal cells. We have previously shown that circulating BAFF levels are maximal at birth, and that farmers’ children exhibit higher BAFF levels in cord blood than non-farmers’ children. Here, we sought to investigate whether maternal-derived decidual stromal cells from placenta secrete BAFF and examine what factors could stimulate this production. We found that podoplanin is expressed in decidua basalis and in the underlying villous tissue as well as on isolated maternal-derived decidual stromal cells. Decidual stromal cells produced BAFF when stimulated with IFN-γ and IFN-α, and NK cells and NK-T-like cells competent of IFN-γ production were isolated from the decidua. Finally, B cells at different maturational stages are present in decidua and all expressed BAFF-R, while stromal cells did not. These findings suggest that decidual stromal cells are a cellular source of BAFF for B cells present in decidua during pregnancy. PMID:28057926

  6. IFN type I and II induce BAFF secretion from human decidual stromal cells.

    PubMed

    Lundell, Anna-Carin; Nordström, Inger; Andersson, Kerstin; Lundqvist, Christina; Telemo, Esbjörn; Nava, Silvia; Kaipe, Helen; Rudin, Anna

    2017-01-06

    B cell activating factor (BAFF) is a critical cytokine for maturation of immature B cells. In murine lymph nodes, BAFF is mainly produced by podoplanin-expressing stromal cells. We have previously shown that circulating BAFF levels are maximal at birth, and that farmers' children exhibit higher BAFF levels in cord blood than non-farmers' children. Here, we sought to investigate whether maternal-derived decidual stromal cells from placenta secrete BAFF and examine what factors could stimulate this production. We found that podoplanin is expressed in decidua basalis and in the underlying villous tissue as well as on isolated maternal-derived decidual stromal cells. Decidual stromal cells produced BAFF when stimulated with IFN-γ and IFN-α, and NK cells and NK-T-like cells competent of IFN-γ production were isolated from the decidua. Finally, B cells at different maturational stages are present in decidua and all expressed BAFF-R, while stromal cells did not. These findings suggest that decidual stromal cells are a cellular source of BAFF for B cells present in decidua during pregnancy.

  7. Differential expression of collagenase by human fibroblasts and bone marrow stromal cells.

    PubMed

    Takahashi, G W; Moran, D; Andrews, D F; Singer, J W

    1994-02-01

    The bone marrow stroma, represented in long-term marrow culture by cells of the adherent layer, is composed of a heterogenous mixture of macrophages and mesenchymal cells, including fibroblasts, endothelial cells and adipocytes, in association with a proteoglycan matrix. This matrix, which is synthesized by the stroma, is capable of binding hematopoietic growth factors, and likely plays a major role in hematopoietic regulation. Clonally-derived non-transformed bone marrow stromal cells, propagated in the presence of basic fibroblast growth factor, were studied for expression of collagenase, an enzyme whose substrate, collagen, is a major component of the extracellular matrix. Expression of steady-state collagenase mRNA was undetectable in both unstimulated dermal fibroblasts and non-transformed marrow stromal cells. However, stimulation with interleukin 1 alpha (10 U/ml) for 24 h resulted in marked accumulation of collagenase mRNA in dermal fibroblast cells, yet failed to elicit a similar response in bone marrow stromal cells. Both marrow stromal cells and dermal fibroblasts constitutively expressed transcripts of collagen I, and rhIL-1 alpha upregulated transcripts of interleukin 6 in both these cells as well. Although similar in morphology, these data indicate that bone marrow stromal cells differ from fibroblasts in their response to IL-1. In the marrow microenvironment, where IL-1 may be secreted by a variety of cell types, such suppression of collagenase expression may serve to prevent unwanted mobilization of collagen from the glycoprotein matrix by marrow stromal cells.

  8. Stromal-epithelial paracrine interactions in the neoplastic rat and human prostate.

    PubMed

    Djakiew, D; Pflug, B; Onoda, M

    1993-01-01

    Homotypic paracrine interactions in the rat and human prostate have been investigated using prostatic stromal cells and neoplastic epithelial cells (PA-III, rat; TSU-pr1, human). Secretory proteins prepared from each cell type were used to determine the dose dependent regulation of growth (DNA synthesis) of the corresponding homotypic responder cell, as determined by 3H-thymidine incorporation. PA-III secretory protein stimulated rat stromal cell proliferation by 1.8-fold. This stimulatory activity of PA-III protein on stromal cell proliferation was partially reduced (approximately 35%) by treatment with nerve growth factor (NGF) antibody, whereas neither acidic fibroblast growth factor (aFGF) antibody nor basic fibroblast growth factor (bFGF) antibody immunoneutralized the stimulatory activity of PA-III cell protein. In the corresponding opposite interaction, rat stromal cell protein modulated PA-III growth in a biphasic manner. At lower concentrations of stromal cell protein (1.25 micrograms/ml) PA-III cell growth was stimulated by 1.6-fold, whereas at higher concentrations of protein (100 micrograms/ml) PA-III cell growth was inhibited to 60%. Treatment of the stromal cell protein (1.25 micrograms/ml and 100 micrograms/ml) with NGF antibody reduced PA-III cell relative growth to approximately 30% and 5%, respectively. bFGF antibody treatment of stromal cell protein at 1.25 micrograms/ml did not influence relative growth, whereas bFGF antibody treatment of 100 micrograms/ml stromal cell protein reduced relative growth by an additional 40%. Treatment of the stromal cell protein (1.25 micrograms/ml and 100 micrograms/ml) with aFGF antibodies reduced relative growth from that observed at these two protein concentrations by approximately 50% in both cases. Human epithelial TSU-pr1 protein stimulated human stromal cell proliferation approximately 1.7-fold. Treatment of TSU-pr1 protein with NGF antibody resulted in stimulation of human stromal cell proliferation (4

  9. CXC chemokine receptor 4 expression and stromal cell-derived factor-1α-induced chemotaxis in CD4+ T lymphocytes are regulated by interleukin-4 and interleukin-10

    PubMed Central

    Jinquan, T; Quan, S; Jacobi, H H; Madsen, H O; Glue, C; Skov, P S; Malling, H-J; Poulsen, L K

    2000-01-01

    We report that interleukin (IL)-4 and IL-10 can significantly up- or down-regulate CXC chemokine receptor 4 (CXCR4) expression on CD4+ T lymphocytes, respectively. Stromal cell-derived factor-1α (SDF-1α)-induced CD4+ T-lymphocyte chemotaxis was also correspondingly regulated by IL-4 and IL-10. IL-4 and IL-10 up- or down-regulated CXCR4 mRNA expression in CD4+ T lymphocytes, respectively, as detected by real-time quantitative reverse transcription–polymerase chain reaction (RT–PCR). Scatchard analysis revealed a type of CXCR4 with affinity (Kd ≈ 6·3 nm), and ≈ 70 000 SDF-1α-binding sites per cell, among freshly isolated CD4+ T lymphocytes, and two types of CXCR4 with different affinities (Kd1 ≈ 4·4 nm and Kd2 ≈ 14·6 nm), and a total of ≈ 130 000 SDF-1α-binding sites per cell, among IL-4-stimulated CD4+ T lymphocytes. The regulation of CXCR4 expression in CD4+ T lymphocytes by IL-4 and IL-10 could be blocked by a selective inhibitor of protein kinase (staurosporine) or by a selective inhibitor of cAMP- and cGMP-dependent protein kinase (H-8), indicating that these cytokines regulate CXCR4 on CD4+ T lymphocytes via both cAMP and cGMP signalling pathways. The fact that cyclosporin A or ionomycin were able to independently change the CXCR4 expression and block the effects of IL-4 and IL-10 on CXCR4 expression implied that the capacity of IL-4 and IL-10 to regulate CXCR4 on CD4+ T lymphocytes is not linked to calcium-mobilization stimulation. These results indicate that the effects of IL-4 and IL-10 on the CXCR4–SDF-1 receptor–ligand pair may be of particular importance in the cytokine/chemokine environment concerning the inflammatory processes and in the progression of human immunodeficiency virus (HIV) infection. PMID:10712670

  10. Question of bone marrow stromal fibroblast traffic

    SciTech Connect

    Maloney, M.A.; Lamela, R.A.; Patt, H.M.

    1985-01-01

    Bone marrow stromal fibroblasts (CFU-F) normally do not exchange bone marrow sites in vivo. Restitution of the CFU-F after radiation damage is primarily recovery by the local fibroblasts from potentially lethal damage. Migration of stromal fibroblasts from shielded sites to an irradiated site makes a minimal contribution, if any, to CFU-F recovery. Determination of the relative contribution of donor stromal cells in bone marrow transplants by karyotyping the proliferating bone marrow stromal cells in vitro may not reflect the relative distribution of fibroblasts in the marrow. If there is residual damage to the host stromal fibroblasts from treatment before transplantation, these cells may not be able to proliferate in vitro. Therefore, an occasional transplanted fibroblast may contribute most of the metaphase figures scored for karyotype.

  11. Stromal Cell Subsets Directing Neonatal Spleen Regeneration

    PubMed Central

    Tan, Jonathan K. H.; Watanabe, Takeshi

    2017-01-01

    Development of lymphoid tissue is determined by interactions between stromal lymphoid tissue organiser (LTo) and hematopoietic lymphoid tissue inducer (LTi) cells. A failure for LTo to receive appropriate activating signals during embryogenesis through lymphotoxin engagement leads to a complete cessation of lymph node (LN) and Peyer’s patch development, identifying LTo as a key stromal population for lymphoid tissue organogenesis. However, little is known about the equivalent stromal cells that induce spleen development. Here, by dissociating neonatal murine spleen stromal tissue for re-aggregation and transplant into adult mouse recipients, we have identified a MAdCAM-1+CD31+CD201+ spleen stromal organizer cell-type critical for new tissue formation. This finding provides an insight into the regulation of post-natal spleen tissue organogenesis, and could be exploited in the development of spleen regenerative therapies. PMID:28067323

  12. Tumour cell-derived exosomes endow mesenchymal stromal cells with tumour-promotion capabilities.

    PubMed

    Lin, L Y; Du, L M; Cao, K; Huang, Y; Yu, P F; Zhang, L Y; Li, F Y; Wang, Y; Shi, Y F

    2016-11-17

    Mesenchymal stromal cells (MSCs) are a major component of the tumour microenvironment. A plethora of elegant studies focusing on tumour-derived MSCs have shown that they, unlike normal MSCs in other tissue, exhibit a strong ability to promote tumour progression. However, the mechanisms underlying the conversion of normal MSCs into tumour-associated MSCs are unknown. We report here a critical role of tumour cell-derived exosomes in endowing bone marrow-derived MSCs (BM-MSCs) with a tumour-favourable phenotype. Tumour cell-derived exosomes affected neither the growth factor production nor the immunosuppressive property of MSCs; rather, they endowed MSCs with a strong ability to promote macrophage infiltration into B16-F0 melanoma or EL-4 lymphoma. Ablation of macrophages by clodronate liposome administration reversed the tumour-promoting effect of MSCs educated by tumour cell-derived exosomes (TE-MSCs) on the tumour growth. By comparing the chemokine profile of BM-MSCs with that of TE-MSCs, we found that TE-MSCs produced a large amount of CCR2 ligands, CCL2 and CCL7, which are responsible for macrophage recruitment. CCR2-specific inhibitor was found to block the tumour-promoting effect of TE-MSCs. Thus, our investigations demonstrated that tumour cell-derived exosomes confer BM-MSCs the ability to enhance tumour growth. Therefore, we uncovered a novel mechanism underlying the conversion of normal MSCs to tumour-associated MSCs.

  13. Novel DREB A-5 subgroup transcription factors from desert moss (Syntrichia caninervis) confers multiple abiotic stress tolerance to yeast.

    PubMed

    Li, Haiyan; Zhang, Daoyuan; Li, Xiaoshuang; Guan, Kaiyun; Yang, Honglan

    2016-05-01

    Syntrichia caninervis Mitt. is a typical desiccation tolerant moss from a temperate desert which has been a good resource for stress tolerant gene isolation. Dehydration responsive element binding proteins (DREBs) was proven to play an important role in responding to abiotic stress, which has been identified in many plants, and were rarely reported in moss. In this study, we cloned ten DREB genes from S. caninervis, and investigated their abiotic stress response and stress tolerance. The results showed that ten ScDREB proteins belonged to the A-5 sub-group of the DREB sub-family. Six genes, ScDREB1, ScDREB2, ScDREB4, ScDREB6, ScDREB7, and ScDREB8 were involved in the ABA-dependent signal pathway and the desiccation, salt, and cold stress response. ScDREB3 also responded to desiccation, salt, and cold stresses, but was insensitive to ABA treatment. Another gene, ScDREB5, was involved in an ABA-independent cold stress-responsive signal pathway. Two genes, ScDREB9 and ScDREB10, responded slightly or had no response to neither stress factor or ABA treatment. We transformed four typical genes into yeast cells and the stress tolerance ability of transgenic yeast was evaluated. The results showed that ScDREB3 and ScDREB5 enhanced the yeast's cold and salt tolerance. ScDREB8 and ScDREB10 conferred the osmotic, salt, cold, and high temperature stresses tolerance, especially for osmotic and salt stresses. Our results indicated that A-5 sub-group DREB genes in S. caninervis played important roles in abiotic stresses response and enhanced stress tolerance to transgenic yeast. To our knowledge, this is the first report on DREB genes characterization from desiccation tolerant moss, and this study will not only provide insight into the molecular mechanisms of S. caninervis adaptation to environmental stresses, but also provides valuable gene candidates for plant molecular breeding.

  14. Stromal Modulators of TGF-β in Cancer

    PubMed Central

    Costanza, Brunella; Umelo, Ijeoma Adaku; Bellier, Justine; Castronovo, Vincent; Turtoi, Andrei

    2017-01-01

    Transforming growth factor-β (TGF-β) is an intriguing cytokine exhibiting dual activities in malignant disease. It is an important mediator of cancer invasion, metastasis and angiogenesis, on the one hand, while it exhibits anti-tumor functions on the other hand. Elucidating the precise role of TGF-β in malignant development and progression requires a better understanding of the molecular mechanisms involved in its tumor suppressor to tumor promoter switch. One important aspect of TGF-β function is its interaction with proteins within the tumor microenvironment. Several stromal proteins have the natural ability to interact and modulate TGF-β function. Understanding the complex interplay between the TGF-β signaling network and these stromal proteins may provide greater insight into the development of novel therapeutic strategies that target the TGF-β axis. The present review highlights our present understanding of how stroma modulates TGF-β activity in human cancers. PMID:28067804

  15. Stromal Modulators of TGF-β in Cancer.

    PubMed

    Costanza, Brunella; Umelo, Ijeoma Adaku; Bellier, Justine; Castronovo, Vincent; Turtoi, Andrei

    2017-01-06

    Transforming growth factor-β (TGF-β) is an intriguing cytokine exhibiting dual activities in malignant disease. It is an important mediator of cancer invasion, metastasis and angiogenesis, on the one hand, while it exhibits anti-tumor functions on the other hand. Elucidating the precise role of TGF-β in malignant development and progression requires a better understanding of the molecular mechanisms involved in its tumor suppressor to tumor promoter switch. One important aspect of TGF-β function is its interaction with proteins within the tumor microenvironment. Several stromal proteins have the natural ability to interact and modulate TGF-β function. Understanding the complex interplay between the TGF-β signaling network and these stromal proteins may provide greater insight into the development of novel therapeutic strategies that target the TGF-β axis. The present review highlights our present understanding of how stroma modulates TGF-β activity in human cancers.

  16. Dickkopf-related protein 3 promotes pathogenic stromal remodeling in benign prostatic hyperplasia and prostate cancer.

    PubMed

    Zenzmaier, Christoph; Sampson, Natalie; Plas, Eugen; Berger, Peter

    2013-09-01

    Compartment-specific epithelial and stromal expression of the secreted glycoprotein Dickkopf-related protein (Dkk)-3 is altered in age-related proliferative disorders of the human prostate. This study aimed to determine the effect of Dkk-3 on prostate stromal remodeling that is stromal proliferation, fibroblast-to-myofibroblast differentiation and expression of angiogenic factors in vitro. Lentiviral-delivered overexpression and shRNA-mediated knockdown of DKK3 were applied to primary human prostatic stromal cells (PrSCs). Cellular proliferation was analyzed by BrdU incorporation ELISA. Expression of Dkk-3, apoptosis-related genes, cyclin-dependent kinase inhibitors and angiogenic factors were analyzed by qPCR, Western blot analysis or ELISA. Fibroblast-to-myofibroblast differentiation was monitored by smooth muscle cell actin and insulin-like growth factor binding protein 3 mRNA and protein levels. The relevance of Wnt/β-catenin and PI3K/AKT signaling pathways was assessed by cytoplasmic/nuclear β-catenin levels and phosphorylation of AKT. Knockdown of DKK3 significantly attenuated PrSC proliferation as well as fibroblast-to-myofibroblast differentiation and increased the expression of the vessel stabilizing factor angiopoietin-1. DKK3 knockdown did not affect subcellular localization or levels of β-catenin but attenuated AKT phosphorylation in PrSCs. Consistently the PI3K/AKT inhibitor LY294002 mimicked the effects of DKK3 knockdown. Dkk-3 promotes fibroblast proliferation and myofibroblast differentiation and regulates expression of angiopoietin-1 in prostatic stroma potentially via enhancing PI3K/AKT signaling. Thus, elevated Dkk-3 in the stroma of the diseased prostate presumably regulates stromal remodeling by enhancing proliferation and differentiation of stromal cells and contributing to the angiogenic switch observed in BPH and PCa. Therefore, Dkk-3 represents a potential therapeutic target for stromal remodeling in BPH and PCa. © 2013 Wiley

  17. Inflammatory stromal keratopathies: medical management of stromal keratomalacia, stromal abscesses, eosinophilic keratitis, and band keratopathy in the horse.

    PubMed

    Brooks, Dennis E

    2004-08-01

    This article discusses the diagnosis and medical treatment of stromal keratomalacia or "melting ulcers," stromal abscesses, eosinophilic keratitis (EK), and calcific band keratopathy. These are common and important inflammatory keratopathies of the equine corneal stroma. Keratomalacia and stromal abscesses are associated with infection, leukocytic invasion of the stroma, and loss of tissue and tear film proteinase homeostasis. Eosinophils infiltrate the stroma in response to unknown stimuli in EK. Calcium is deposited in the stroma and epithelium secondary to chronic equine recurrent uveitis in calcific band keratopathy. They are all associated with varying degrees of iridocyclitis.

  18. WINGLESS (WNT) signaling is a progesterone target for rat uterine stromal cell proliferation.

    PubMed

    Rider, Virginia; Talbott, Alex; Bhusri, Anuradha; Krumsick, Zach; Foster, Sierra; Wormington, Joshua; Kimler, Bruce F

    2016-05-01

    Preparation of mammalian uterus for embryo implantation requires a precise sequence of cell proliferation. In rodent uterus, estradiol stimulates proliferation of epithelial cells. Progesterone operates as a molecular switch and redirects proliferation to the stroma by down-regulating glycogen synthase kinase-3β (GSK-3β) and stimulating β-catenin accumulation in the periluminal stromal cells. In this study, the WNT signal involved in the progesterone-dependent proliferative switch was investigated. Transcripts of four candidate Wnt genes were measured in the uteri from ovariectomized (OVX) rats, progesterone-pretreated (3 days of progesterone, 2mg/daily) rats, and progesterone-pretreated rats given a single dose (0.2µg) of estradiol. The spatial distribution of the WNT proteins was determined in the uteri after the same treatments. Wnt5a increased in response to progesterone and the protein emerged in the periluminal stromal cells of progesterone-pretreated rat uteri. To investigate whether WNT5A was required for proliferation, uterine stromal cell lines were stimulated with progesterone (1µM) and fibroblast growth factor (FGF, 50ng/mL). Proliferating stromal cells expressed a two-fold increase in WNT5A protein at 12h post stimulation. Stimulated stromal cells were cultured with actinomycin D (25µg/mL) to inhibit new RNA synthesis. Relative Wnt5a expression increased at 4 and 6 h of culture, suggesting that progesterone plus FGF preferentially increased Wnt5a mRNA stability. Knockdown of Wnt5a in uterine stromal cell lines inhibited stromal cell proliferation and decreased Wnt5a mRNA. The results indicate that progesterone initiates and synchronizes uterine stromal cell proliferation by increasing WNT5A expression and signaling. © 2016 The authors.

  19. Paracrine effects of uterine leucocytes on gene expression of human uterine stromal fibroblasts.

    PubMed

    Germeyer, Ariane; Sharkey, Andrew Mark; Prasadajudio, Mirari; Sherwin, Robert; Moffett, Ashley; Bieback, Karen; Clausmeyer, Susanne; Masters, Leanne; Popovici, Roxana Maria; Hess, Alexandra Petra; Strowitzki, Thomas; von Wolff, Michael

    2009-01-01

    The endometrium contains a distinct population of immune cells that undergo cyclic changes during the menstrual cycle and implantation. The majority of these leucocytes are uterine NK (uNK) cells, however how these cells interact with uterine stromal fibroblasts remains unclear. We therefore investigated the paracrine effect of medium conditioned by uterine decidual leucocytes (which are enriched for uNK cells) on the gene expression profile of endometrial stromal fibroblasts in vitro using a cDNA microarray. Our results, verified by real-time PCR, ELISA and FACS analysis, reveal that soluble factors from uterine leucocytes substantially alter endometrial stromal fibroblast gene expression. The largest group of up-regulated genes found was chemokines and cytokines. These include IL-8, CCL8 and CXCL1, which have also been shown to be stimulated by contact of stromal fibroblasts with trophoblast, suggesting that uNK cells work synergistically to support trophoblast migration during implantation. The decidual leucocytes also up-regulated IL-15 and IL-15Ralpha in stromal fibroblasts which could produce a niche for uNK cells allowing proliferation within and recruitment into the uterus, as seen in bone marrow. Overall this study demonstrates, for the first time, the paracrine communication between uterine leucocytes and uterine stromal fibroblasts, and adds to the understanding of how the uterine immune system contributes to the changes seen within the cycling endometrium.

  20. Giant gastrointestinal stromal tumor of the stomach.

    PubMed

    Ionescu, Sever; Barbu, Emil; Ionescu, Călin; Costache, Adrian; Bălăşoiu, Maria

    2015-01-01

    Gastrointestinal stromal tumors (GISTs) are the most common mesenchymal malignancies of the digestive tract. Gastric localization is the most frequent. The aim of this study is to evaluate the importance of immunohistochemical factors (CD117, CD34, α-SMA, vimentin, p53, Ki67) in diagnostic and size tumor and mitotic activity as prognostic factors for these tumors. We present the case of a 66-year-old male patient with a giant gastric GIST. Like in the vast majority, the symptomatology in this patient has long been faint, despite the large tumor size, and when it became manifest, it was nonspecific. Imagery wise, the computer tomography (CT) scan was the most efficient, showing the origin of the tumor from the greater curvature of the stomach, its dimensions, as well as the relations with the other abdominal viscera. Surgery in this patient was en-bloc, according to the principles of GIST. The histological aspect is characterized by a proliferation of spindle cells positive for CD117 and CD34. Despite complete microscopic resection, the size of the tumor (25×20×27 cm) and the mitotic activity (21÷5 mm2) remains important relapse factor.

  1. Decidualized Human Endometrial Stromal Cells Mediate Hemostasis, Angiogenesis, and Abnormal Uterine Bleeding

    PubMed Central

    Lockwood, Charles J.; Krikun, Graciela; Hickey, Martha; Huang, S. Joseph; Schatz, Frederick

    2011-01-01

    Factor VII binds trans-membrane tissue factor to initiate hemostasis by forming thrombin. Tissue factor expression is enhanced in decidualized human endometrial stromal cells during the luteal phase. Long-term progestin only contraceptives elicit: 1) abnormal uterine bleeding from fragile vessels at focal bleeding sites, 2) paradoxically high tissue factor expression at bleeding sites; 3) reduced endometrial blood flow promoting local hypoxia and enhancing reactive oxygen species levels; and 4) aberrant angiogenesis reflecting increased stromal cell-expressed vascular endothelial growth factor, decreased Angiopoietin-1 and increased endothelial cell-expressed Angiopoietin-2. Aberrantly high local vascular permeability enhances circulating factor VII to decidualized stromal cell-expressed tissue factor to generate excess thrombin. Hypoxia-thrombin interactions augment expression of vascular endothelial growth factor and interleukin-8 by stromal cells. Thrombin, vascular endothelial growth factor and interlerukin-8 synergis-tically augment angiogenesis in a milieu of reactive oxygen species-induced endothelial cell activation. The resulting enhanced vessel fragility promotes abnormal uterine bleeding. PMID:19208784

  2. Immunohistochemical expression of epithelial and stromal immunomodulatory signalling molecules is a prognostic indicator in breast cancer

    PubMed Central

    2012-01-01

    Background The immune system has paradoxical roles during cancer development and the prognostic significance of immune modulating factors is controversial. The aim of this study was to determine the expression of cyclooxygenase 2 (COX-2), transforming growth factor-beta (TGF- beta), interleukin-10 (IL-10) and their prognostic significance in breast cancers. Ki67 was included as a measure of growth fraction of tumor cells. Methods On immunohistochemical stained slides from 38 breast cancer patients, we performed digital video analysis of tumor cell areas and adjacent tumor stromal areas from the primary tumors and their corresponding lymph node metastases. COX-2 was recorded as graded staining intensity. Results The expression of TGF-beta, IL-10 and Ki67 were recorded in tumor cell areas and adjacent tumor stromal areas. In both primary tumors and metastases, the expression of COX-2 was higher in the tumor stromal areas than in the tumor cell areas (both P < 0.001). High stromal staining intensity in the primary tumors was associated with a 3.9 (95% CI 1.1-14.2) times higher risk of death compared to the low staining group (P = 0.036). The expression of TGF-beta was highest in the tumor cell areas of both primary tumors and metastases (both P < 0.001). High stromal expression of TGF-beta was associated with increased mortality. For IL-10, the stromal expression was highest in the primary tumors (P < 0.001), whereas in the metastases the expression was highest in tumor cell areas (P < 0.001). High IL-10 expression in tumor- and stromal cell areas of primary tumors predicted mortality. Ki67 was higher expressed in tumor stromal areas of the metastases, and in tumor cell areas of the primary tumors (P < 0.001). Ki67 expression in tumor cell areas and stromal areas of the metastases was independently associated with breast cancer mortality. Conclusions Stromal expression of COX-2, TGF-beta and Ki67 may facilitate tumor progression in breast cancer. PMID:22353218

  3. Heterogeneity of multipotent mesenchymal stromal cells: from stromal cells to stem cells and vice versa.

    PubMed

    Dominici, Massimo; Paolucci, Paolo; Conte, Pierfranco; Horwitz, Edwin M

    2009-05-15

    Discovered more than 40 years ago, the biological features of multipotent mesenchymal stromal cells (MSC) were progressively compared first with hematopoietic stem cells (HSC) and, more recently, with embryonic stem cells (ESC). Although these comparisons have been crucial in helping to clarify their nature, there is now a robust amount of data indicating that MSC in vitro represent an independent and heterogeneous group of progenitors with distinct self-renewal properties and established differentiation potentials. However, research developments both in humans and animals have progressively revealed the limits that MSC may face in vivo. To recognize these issues and challenge MSC stemness may seem to be a step backward. Nevertheless, it might also represent the beginning of a phase in which the introduction of novel preclinical approaches could provide better characterization and standardization of the in vivo factors influencing cell engraftment and survival, allowing a more successful impact of mesenchymal progenitors in several clinical settings.

  4. Effectiveness of Bone Marrow Stromal Cell Sheets in Maintaining Random-Pattern Skin Flaps in an Experimental Animal Model.

    PubMed

    Kira, Tsutomu; Omokawa, Shohei; Akahane, Manabu; Shimizu, Takamasa; Nakano, Kenichi; Nakanishi, Yasuaki; Onishi, Tadanobu; Kido, Akira; Inagaki, Yusuke; Tanaka, Yasuhito

    2015-11-01

    Bone marrow stromal cells can be applied therapeutically to enhance angiogenesis; however, the use of bone marrow stromal cell suspensions reduces efficiency because of low-level attachment. The authors hypothesized that bone marrow stromal cell sheets would facilitate cell fixation, thus enhancing angiogenesis. The authors investigated flap survival area and enhancement of angiogenic factors in a rat random-pattern skin flap model after application of bone marrow stromal cell sheets. Bone marrow stromal cell sheets (prepared from 7-week-old rat femurs) were cultured under four different hypoxic conditions. Sheets with the highest angiogenic potential, determined by an in vitro pilot study, were injected into subcutaneous layers of the rat dorsum (bone marrow stromal cell sheet group). A control group (phosphate-buffered saline only) was included. On day 2 after injection, caudally based random-pattern skin flaps (12 × 3 cm) were elevated. On day 7 after elevation, surviving skin flap areas were measured. Skin samples were harvested from each flap and gene expression levels of vascular endothelial growth factor and basic fibroblast growth factor were measured by quantitative real-time polymerase chain reaction. Skin flap survival area (71.6 ± 2.3 percent versus 51.5 ± 3.3 percent) and levels of vascular endothelial growth factor and basic fibroblast growth factor were significantly higher in the bone marrow stromal cell sheet group than in the control group (p < 0.05). Implantation of bone marrow stromal cell sheets increased the survival area of random-pattern skin flaps. Expression of angiogenic factors may have contributed to the increased flap survival.

  5. Treatment for Stromal Tumors of the Ovary

    MedlinePlus

    ... Epithelial Tumors of Low Malignant Potential Treatment for Germ Cell Tumors of the Ovary Treatment for Stromal Tumors ... used is what’s used in the treatment of germ cell tumors (PEB: cisplatin, etoposide, and bleomycin). The combination ...

  6. Conference Resolution

    NASA Astrophysics Data System (ADS)

    2009-04-01

    Since the first IUPAP International Conference on Women in Physics (Paris, March 2002) and the Second Conference (Rio de Janeiro, May 2005), progress has continued in most countries and world regions to attract girls to physics and advance women into leadership roles, and many working groups have formed. The Third Conference (Seoul, October 2008), with 283 attendees from 57 countries, was dedicated to celebrating the physics achievements of women throughout the world, networking toward new international collaborations, building each participant's capacity for career success, and aiding the formation of active regional working groups to advance women in physics. Despite the progress, women remain a small minority of the physics community in most countries.

  7. [Reorganization of actin cytoskeleton in the initial stage of transendothelial migration of bone marrow multipotent mesenchymal stromal cells].

    PubMed

    Aleksandrova, S A; Pinaev, G P

    2014-01-01

    The analysis of actin cytoskeleton reorganization in rat bone marrow multipotent mesenchymal stromal cells after one hour adhesion to a monolayer of endothelial cell line EA.hy 926 allowed us to identify three types of cells interacting with the endothelial cells. Approximately half of multipotent mesenchymal stromal cells retained a rounded shape, most of them contained large round actin aggregates, had irregular borders and contacted with the surface of the endothelial cells by microvilli or protrusions similar to small lamellae. Almost all other cells were surrounded by narrow lamellae along the entire perimeter. In addition, a small amount.of elongated flattened cells that contacting with endothelial cells by means of focal contacts was observed. Microenvironmental factors such as proinflammatory cytokine tumor necrosis factor α or plasma proteins affected the ratio of stromal cell types, with different types of organization of the actin cytoskeleton in multipotent mesenchymal stromal cells population.

  8. Mesenchymal stromal cell proliferation, gene expression and protein production in human platelet-rich plasma-supplemented media.

    PubMed

    Amable, Paola Romina; Teixeira, Marcus Vinicius Telles; Carias, Rosana Bizon Vieira; Granjeiro, José Mauro; Borojevic, Radovan

    2014-01-01

    Platelet-rich plasma (PRP) is increasingly used as a cell culture supplement, in order to reduce the contact of human cells with animal-derived products during in vitro expansion. The effect of supplementation changes on cell growth and protein production is not fully characterized. Human mesenchymal stromal cells from bone marrow, adipose tissue and Wharton's Jelly were isolated and cultured in PRP-supplemented media. Proliferation, in vitro differentiation, expression of cell surface markers, mRNA expression of key genes and protein secretion were quantified. 10% PRP sustained five to tenfold increased cell proliferation as compared to 10% fetal bovine serum. Regarding cell differentiation, PRP reduced adipogenic differentiation and increased calcium deposits in bone marrow and adipose tissue-mesenchymal stromal cells. Wharton's Jelly derived mesenchymal stromal cells secreted higher concentrations of chemokines and growth factors than other mesenchymal stromal cells when cultured in PRP-supplemented media. Bone marrow derived mesenchymal stromal cells secreted higher concentrations of pro-inflammatory and pro-angiogenic proteins. Mesenchymal stromal cells isolated from adipose tissue secreted higher amounts of extracellular matrix components. Mesenchymal stromal cells purified from different tissues have distinct properties regarding differentiation, angiogenic, inflammatory and matrix remodeling potential when cultured in PRP supplemented media. These abilities should be further characterized in order to choose the best protocols for their therapeutic use.

  9. Expression of neuroendocrine factor VGF in lung cancer cells confers resistance to EGFR kinase inhibitors and triggers epithelial-to-mesenchymal transition.

    PubMed

    Wen, Hwang; Chiu, Yu-Fan; Kuo, Ming-Han; Lee, Kuan-Lin; Lee, An-Chun; Yu, Chia-Cherng; Chang, Junn-Liang; Huang, Wen-Chien; Hsiao, Shih-Hsin; Lin, Sey-En; Chou, Yu-Ting

    2017-04-05

    Mutations in EGFR drive tumor growth but render tumor cells sensitive to treatment with EGFR tyrosine kinase inhibitors (TKIs). Phenotypic alteration in epithelial-to-mesenchymal transition (EMT) has been linked to the TKI resistance in lung adenocarcinoma. However, the mechanism underlying this resistance remains unclear. Here we report that high expression of a neuroendocrine factor termed VGF induces the transcription factor TWIST1 to facilitate TKI resistance, EMT, and cancer dissemination in a subset of lung adenocarcinoma cells. VGF silencing resensitized EGFR-mutated lung adenocarcinoma cells to TKI. Conversely, overexpression of VGF in sensitive cells conferred resistance to TKIs and induced EMT, increasing migratory and invasive behaviors. Correlation analysis revealed a significant association of VGF expression with advanced tumor grade and poor survival in patients with lung adenocarcinoma. In a mouse xenograft model of lung adenocarcinoma, suppressing VGF expression was sufficient to attenuate tumor growth. Overall, our findings show how VGF can confer TKI resistance and trigger EMT, suggesting its potential utility as a biomarker and therapeutic target in lung adenocarcinoma.

  10. Targeted therapy of gastrointestinal stromal tumours

    PubMed Central

    Jakhetiya, Ashish; Garg, Pankaj Kumar; Prakash, Gaurav; Sharma, Jyoti; Pandey, Rambha; Pandey, Durgatosh

    2016-01-01

    Gastrointestinal stromal tumours (GISTs) are mesenchymal neoplasms originating in the gastrointestinal tract, usually in the stomach or the small intestine, and rarely elsewhere in the abdomen. The malignant potential of GISTs is variable ranging from small lesions with a benign behaviour to fatal sarcomas. The majority of the tumours stain positively for the CD-117 (KIT) and discovered on GIST-1 (DOG-1 or anoctamin 1) expression, and they are characterized by the presence of a driver kinase-activating mutation in either KIT or platelet-derived growth factor receptor α. Although surgery is the primary modality of treatment, almost half of the patients have disease recurrence following surgery, which highlights the need for an effective adjuvant therapy. Traditionally, GISTs are considered chemotherapy and radiotherapy resistant. With the advent of targeted therapy (tyrosine kinase inhibitors), there has been a paradigm shift in the management of GISTs in the last decade. We present a comprehensive review of targeted therapy in the management of GISTs. PMID:27231512

  11. Stromal influences on breast cancer cell growth.

    PubMed Central

    van Roozendaal, C. E.; van Ooijen, B.; Klijn, J. G.; Claassen, C.; Eggermont, A. M.; Henzen-Logmans, S. C.; Foekens, J. A.

    1992-01-01

    Paracrine influences from fibroblasts derived from different sources of breast tissue on epithelial breast cancer cell growth in vitro were investigated. Medium conditioned (CM) by fibroblasts derived from tumours, adjacent normal breast tissue, and normal breast tissue obtained from reduction mammoplasty or from skin tissue significantly stimulated the growth of the steroid-receptor positive cell lines MCF-7 and ZR 75.1. The proliferation index (PI) on MCF-7 cells with CM from fibroblasts derived from breast tumour tissue was significantly higher than that obtained with fibroblasts derived from adjacent normal breast tissue (2p less than 0.05, n = 8). The PI obtained with CM from normal fibroblast cultures from reduction mammoplasty tissue, like normal tissue adjacent to the tumour, fell in the lower range of values. Skin fibroblast, like tumour tissue derived fibroblast, CM caused a high range PI. MDA-MB-231 and Evsa-T, two steroid-receptor negative cell lines, showed only a minor growth stimulatory responses with some of the fibroblast CM's. Evsa-T was occasionally inhibited by CM's. In conclusion, stromal factors play a role in the growth regulation of human breast cancer cells. The effects on cancer cell growth are, however, varying depending on the source of the stroma and the characteristics of the epithelial tumour cells. PMID:1733444

  12. Imatinib treatment for gastrointestinal stromal tumour (GIST)

    PubMed Central

    Lopes, Lisandro F; Bacchi, Carlos E

    2010-01-01

    Abstract Gastrointestinal stromal tumour (GIST) is the most common mesenchymal neoplasm of the gastrointestinal tract. GISTs are believed to originate from intersticial cells of Cajal (the pacemaker cells of the gastrointestinal tract) or related stem cells, and are characterized by KIT or platelet-derived growth factor receptor alpha (PDGFRA) activating mutations. The use of imatinib has revolutionized the management of GIST and altered its natural history, substantially improving survival time and delaying disease progression in many patients. The success of imatinib in controlling advanced GIST led to interest in the neoadjuvant and adjuvant use of the drug. The neoadjuvant (preoperative) use of imatinib is recommended to facilitate resection and avoid mutilating surgery by decreasing tumour size, and adjuvant therapy is indicated for patients at high risk of recurrence. The molecular characterization (genotyping) of GISTs has become an essential part of the routine management of the disease as KIT and PDGFRA mutation status predicts the likelihood of achieving response to imatinib. However, the vast majority of patients who initially responded to imatinib will develop tumour progression (secondary resistance). Secondary resistance is often related to secondary KIT or PDGFRA mutations that interfere with drug binding. Multiple novel tyrosine kinase inhibitors may be potentially useful for the treatment of imatinib-resistant GISTs as they interfere with KIT and PDGFRA receptors or with the downstream-signalling proteins. PMID:19968734

  13. Imatinib treatment for gastrointestinal stromal tumour (GIST).

    PubMed

    Lopes, Lisandro F; Bacchi, Carlos E

    2010-01-01

    Gastrointestinal stromal tumour (GIST) is the most common mesenchymal neoplasm of the gastrointestinal tract. GISTs are believed to originate from intersticial cells of Cajal (the pacemaker cells of the gastrointestinal tract) or related stem cells, and are characterized by KIT or platelet-derived growth factor receptor alpha (PDGFRA) activating mutations. The use of imatinib has revolutionized the management of GIST and altered its natural history, substantially improving survival time and delaying disease progression in many patients. The success of imatinib in controlling advanced GIST led to interest in the neoadjuvant and adjuvant use of the drug. The neoadjuvant (preoperative) use of imatinib is recommended to facilitate resection and avoid mutilating surgery by decreasing tumour size, and adjuvant therapy is indicated for patients at high risk of recurrence. The molecular characterization (genotyping) of GISTs has become an essential part of the routine management of the disease as KIT and PDGFRA mutation status predicts the likelihood of achieving response to imatinib. However, the vast majority of patients who initially responded to imatinib will develop tumour progression (secondary resistance). Secondary resistance is often related to secondary KIT or PDGFRA mutations that interfere with drug binding. Multiple novel tyrosine kinase inhibitors may be potentially useful for the treatment of imatinib-resistant GISTs as they interfere with KIT and PDGFRA receptors or with the downstream-signalling proteins.

  14. Biomedical Conferences

    NASA Technical Reports Server (NTRS)

    1976-01-01

    As a result of Biomedical Conferences, Vivo Metric Systems Co. has produced cardiac electrodes based on NASA technology. Frequently in science, one highly specialized discipline is unaware of relevant advances made in other areas. In an attempt to familiarize researchers in a variety of disciplines with medical problems and needs, NASA has sponsored conferences that bring together university scientists, practicing physicians and manufacturers of medical instruments.

  15. Overexpression of the pepper transcription factor CaPF1 in transgenic Virginia pine (Pinus Virginiana Mill.) confers multiple stress tolerance and enhances organ growth.

    PubMed

    Tang, Wei; Charles, Thomas M; Newton, Ronald J

    2005-11-01

    Transcription factors play an important role in regulating gene expression in response to stress and pathogen tolerance. We describe here that overexpression of an ERF/AP2 pepper transcription factor (CaPF1) in transgenic Virginia pine (Pinus virginiana Mill.) confers tolerance to heavy metals Cadmium, Copper, and Zinc, to heat, and to pathogens Bacillus thuringiensis and Staphylococcus epidermidis, as by the survival rate of transgenic plants and the number of decreasing pathogen cells in transgenic tissues. Measurement of antioxidant enzymes ascorbate peroxidase (APOX), glutathione reductase (GR), and superoxide dismutase (SOD) activities demonstrated that the level of the enzyme activities was higher in transgenic Virginia pine plants overexpressing the CaPF1 gene, which may protect cells from the oxidative damage caused by stresses, compared to the controls. Constitutive overexpression of CaPF1 gene enhanced organ growth by increasing organ size and cell numbers in transgenic Virginia pine plants over those in control plants.

  16. [Atypical stromal hyperplasia of the prostate (stromal proliferation of uncertain malignant potential)].

    PubMed

    Medina Pérez, M; Valero Puerta, J A; Pérez Martín, D

    2000-10-01

    To describe a case of atypical stromal hyperplasia of the prostate. A 62-year-old patient presented with prostatic syndrome. Physical examination disclosed an indurated prostate and PSA determination showed increased levels. A prostate biopsy was performed. The histological analysis showed atypical stromal proliferation with elongated nuclei and immunohistochemical expression for vimentine, smooth muscle actin and CD34 with glandular hyperplasia. The diagnosis was that of atypical stromal hyperplasia of the prostate (prostatic stromal proliferation of uncertain malignant potential). A careful histological study is necessary to make the correct diagnosis of prostatic stromal proliferation of uncertain malignant potential. CD34 expression is a characteristic finding. As its name indicates, its evolution is uncertain.

  17. Orthologs of the class A4 heat shock transcription factor HsfA4a confer cadmium tolerance in wheat and rice.

    PubMed

    Shim, Donghwan; Hwang, Jae-Ung; Lee, Joohyun; Lee, Sichul; Choi, Yunjung; An, Gynheung; Martinoia, Enrico; Lee, Youngsook

    2009-12-01

    Cadmium (Cd) is a widespread soil pollutant; thus, the underlying molecular controls of plant Cd tolerance are of substantial interest. A screen for wheat (Triticum aestivum) genes that confer Cd tolerance to a Cd hypersensitive yeast strain identified Heat shock transcription factor A4a (HsfA4a). Ta HsfA4a is most similar to the class A4 Hsfs from monocots. The most closely related rice (Oryza sativa) homolog, Os HsfA4a, conferred Cd tolerance in yeast, as did Ta HsfA4a, but the second most closely related rice homolog, Os HsfA4d, did not. Cd tolerance was enhanced in rice plants expressing Ta HsfA4a and decreased in rice plants with knocked-down expression of Os HsfA4a. An analysis of the functional domain using chimeric proteins constructed from Ta HsfA4a and Os HsfA4d revealed that the DNA binding domain (DBD) of HsfA4a is critical for Cd tolerance, and within the DBD, Ala-31 and Leu-42 are important for Cd tolerance. Moreover, Ta HsfA4a-mediated Cd resistance in yeast requires metallothionein (MT). In the roots of wheat and rice, Cd stress caused increases in HsfA4a expression, together the MT genes. Our findings thus suggest that HsfA4a of wheat and rice confers Cd tolerance by upregulating MT gene expression in planta.

  18. Endometrial Stromal Decidualization Responds Reversibly to Hormone Stimulation and Withdrawal

    PubMed Central

    Yu, Jie; Berga, Sarah L.; Johnston-MacAnanny, Erika B.; Sidell, Neil; Bagchi, Indrani C.; Bagchi, Milan K.

    2016-01-01

    Human endometrial stromal decidualization is required for embryo receptivity, angiogenesis, and placentation. Previous studies from our laboratories established that connexin (Cx)-43 critically regulates endometrial stromal cell (ESC) differentiation, whereas gap junction blockade prevents it. The current study evaluated the plasticity of ESC morphology and Cx43 expression, as well as other biochemical markers of cell differentiation, in response to decidualizing hormones. Primary human ESC cultures were exposed to 10 nM estradiol, 100 nM progesterone, and 0.5 mM cAMP for up to 14 days, followed by hormone withdrawal for 14 days, mimicking a biphasic ovulatory cycle. Reversible differentiation was documented by characteristic changes in cell shape. Cx43 was reversibly up- and down-regulated after the estradiol, progesterone, and cAMP treatment and withdrawal, respectively, paralleled by fluctuations in prolactin, vascular endothelial growth factor, IL-11, and glycodelin secretion. Markers of mesenchymal-epithelial transition (MET), and its counterpart epithelial-mesenchymal transition, followed reciprocal patterns corresponding to the morphological changes. Incubation in the presence of 18α-glycyrrhetinic acid, an inhibitor of gap junctions, partially reversed the expression of decidualization and MET markers. In the absence of hormones, Cx43 overexpression promoted increases in vascular endothelial growth factor and IL-11 secretion, up-regulated MET markers, and reduced N-cadherin, an epithelial-mesenchymal transition marker. The combined results support the hypothesis that Cx43-containing gap junctions and endocrine factors cooperate to regulate selected biomarkers of stromal decidualization and MET and suggest roles for both phenomena in endometrial preparation for embryonic receptivity. PMID:27035651

  19. Gastrointestinal Stromal Tumours: An Update

    PubMed Central

    Somerhausen, Nicolas De Saint Aubain

    1998-01-01

    Purpose. To study the evolution of concepts concerning gastrointestinal stromal tumours (GISTs) over 30 years. Discussion. GISTs have been, for more than 30 years, the subject of considerable controversy regarding their line of differentiation as well as the prediction of their behaviour. Furthermore, once they spread within the peritoneal cavity, they are extremely hard to control. The recent findings of c-Kit mutations and the immunohistochemical detection of the product of this gene, KIT or CD117, in the mainly non-myogenic subset of this family of tumours, has led to a reappraisal of this group of lesions, which, with some exceptions, is now thought to be derived from the interstitial cells of Cajal, and this has facilitated a clearer definition of their pathological spectrum. In this article, we review chronologically the evolution of the concept of GIST with the gradual application of electron microscopy, immunohistochemistry, DNA ploidy analysis. We discuss the impact of these techniques on the pathological assessment and clinical management of GISTs. PMID:18521245

  20. Ghrelin and gastrointestinal stromal tumors

    PubMed Central

    Zhu, Chang-Zhen; Liu, Dong; Kang, Wei-Ming; Yu, Jian-Chun; Ma, Zhi-Qiang; Ye, Xin; Li, Kang

    2017-01-01

    Ghrelin, as a kind of multifunctional protein polypeptide, is mainly produced in the fundus of the stomach and can promote occurrence and development of many tumors, including gastrointestinal tumors, which has been proved by the relevant researches. Most gastrointestinal stromal tumors (GISTs, about 80%), as the most common mesenchymal tumor, also develop in the fundus. Scientific research has confirmed that ghrelin, its receptors and mRNA respectively can be found in GISTs, which demonstrated the existence of a ghrelin autocrine/paracrine loop in GIST tissues. However, no reports to date have specified the mechanism whether ghrelin can promote the occurrence and development of GISTs. Studies of pulmonary artery endothelial cells in a low-oxygen environment and cardiac muscle cells in an ischemic environment have shown that ghrelin can activate the phosphatidylinositol 3-kinase/AKT/mammalian target of rapamycin (PI3K/AKT/mTOR) signaling pathway. Moreover, some studies of GISTs have confirmed that activation of the PI3K/AKT/mTOR pathway can indeed promote the growth and progression of GISTs. Whether ghrelin is involved in the development or progression of GISTs through certain pathways remains unknown. Can we find a new target for the treatment of GISTs? This review explores and summaries the relationship among ghrelin, the PI3K/AKT/mTOR pathway and the development of GISTs. PMID:28348480

  1. Ghrelin and gastrointestinal stromal tumors.

    PubMed

    Zhu, Chang-Zhen; Liu, Dong; Kang, Wei-Ming; Yu, Jian-Chun; Ma, Zhi-Qiang; Ye, Xin; Li, Kang

    2017-03-14

    Ghrelin, as a kind of multifunctional protein polypeptide, is mainly produced in the fundus of the stomach and can promote occurrence and development of many tumors, including gastrointestinal tumors, which has been proved by the relevant researches. Most gastrointestinal stromal tumors (GISTs, about 80%), as the most common mesenchymal tumor, also develop in the fundus. Scientific research has confirmed that ghrelin, its receptors and mRNA respectively can be found in GISTs, which demonstrated the existence of a ghrelin autocrine/paracrine loop in GIST tissues. However, no reports to date have specified the mechanism whether ghrelin can promote the occurrence and development of GISTs. Studies of pulmonary artery endothelial cells in a low-oxygen environment and cardiac muscle cells in an ischemic environment have shown that ghrelin can activate the phosphatidylinositol 3-kinase/AKT/mammalian target of rapamycin (PI3K/AKT/mTOR) signaling pathway. Moreover, some studies of GISTs have confirmed that activation of the PI3K/AKT/mTOR pathway can indeed promote the growth and progression of GISTs. Whether ghrelin is involved in the development or progression of GISTs through certain pathways remains unknown. Can we find a new target for the treatment of GISTs? This review explores and summaries the relationship among ghrelin, the PI3K/AKT/mTOR pathway and the development of GISTs.

  2. Proliferation and apoptosis in ovarian stromal hyperplasia and hyperthecosis.

    PubMed

    Sharabidze, N; Sabakhtarashvili, M; Burkadze, G

    2005-12-01

    The aim of our study was to investigate proliferation and apoptosis in ovarian stromal hyperplasia and hyperthecosis in reproductive women with and without polycystic ovarian disease. We have studied 104 patients with a histological diagnosis of ovarian stromal hyperthecosis and stromal hyperplasia. Paraffin sections were stained by hematoxylin-eosin, von Gieson and immunohistochemistry for Bcl-2 (anti-apoptotic protein) and Ki-67 antigen (proliferation marker). We assessed the number of Bcl-2-positive and Ki-67-positive cells. The patients were divided into 4 groups: group 1 -- 33 patients with polycystic ovarian disease and coexistent stromal hyperthecosis, group 2 -- 28 patients with polycystic ovarian disease and coexistent stromal hyperplasia, group 3 -- 24 patients with ovarian stromal hyperthecosis, group 4-19 patients with ovarian stromal hyperplasia. The results suggest that in ovarian stromal hyperthecosis coexistent with polycystic ovarian disease luteinized stromal cells show high proliferation activity and resistance to apoptosis, and internal and external theca cells show resistance to apoptosis. In ovarian stromal hyperplasia coexistent with polycystic ovarian disease, hyperplastic stromal cells show high proliferation activity and resistance to apoptosis. Proliferation activity is also increased in vascular endothelial and smooth muscle cells. Internal and external theca cells show resistance to apoptosis. In ovarian stromal hyperthecosis without polycystic ovarian disease, luteinized stromal cells show high proliferation activity and resistance to apoptosis. In ovarian stromal hyperplasia without polycystic ovarian disease, hyperplastic stromal cells show high proliferation activity and potential susceptibility to apoptosis. Proliferation activity is also increased in vascular endothelial and smooth muscle cells. These characteristics of proliferation and apoptosis have meaning in pathogenesis of ovarian stromal hyperthecosis and stromal

  3. Microarray analysis of colorectal cancer stromal tissue reveals upregulation of two oncogenic miRNA clusters.

    PubMed

    Nishida, Naohiro; Nagahara, Makoto; Sato, Tetsuya; Mimori, Koshi; Sudo, Tomoya; Tanaka, Fumiaki; Shibata, Kohei; Ishii, Hideshi; Sugihara, Kenichi; Doki, Yuichiro; Mori, Masaki

    2012-06-01

    Cancer stroma plays an important role in the progression of cancer. Although alterations in miRNA expression have been explored in various kinds of cancers, the expression of miRNAs in cancer stroma has not been explored in detail. Using a laser microdissection technique, we collected RNA samples specific for epithelium or stroma from 13 colorectal cancer tissues and four normal tissues, and miRNA microarray and gene expression microarray were carried out. The expression status of miRNAs was confirmed by reverse transcriptase PCR. Furthermore, we investigated whether miRNA expression status in stromal tissue could influence the clinicopathologic factors. Oncogenic miRNAs, including two miRNA clusters, miR-17-92a and miR-106b-25 cluster, were upregulated in cancer stromal tissues compared with normal stroma. Gene expression profiles from cDNA microarray analyses of the same stromal tissue samples revealed that putative targets of these miRNA clusters, predicted by Target Scan, such as TGFBR2, SMAD2, and BMP family genes, were significantly downregulated in cancer stromal tissue. Downregulated putative targets were also found to be involved in cytokine interaction and cellular adhesion. Importantly, expression of miR-25 and miR-92a in stromal tissues was associated with a variety of clinicopathologic factors. Oncogenic miRNAs were highly expressed in cancer stroma. Although further validation is required, the finding that stromal miRNA expression levels were associated with clinicopathologic factors suggests the possibility that miRNAs in cancer stroma are crucially involved in cancer progression.

  4. Inhibition of tumor-stromal interaction through HGF/Met signaling by valproic acid

    SciTech Connect

    Matsumoto, Yohsuke; Motoki, Takahiro; Kubota, Satoshi; Takigawa, Masaharu; Tsubouchi, Hirohito; Gohda, Eiichi

    2008-02-01

    Hepatocyte growth factor (HGF), which is produced by surrounding stromal cells, including fibroblasts and endothelial cells, has been shown to be a significant factor responsible for cancer cell invasion mediated by tumor-stromal interactions. We found in this study that the anti-tumor agent valproic acid (VPA), a histone deacetylase (HDAC) inhibitor, strongly inhibited tumor-stromal interaction. VPA inhibited HGF production in fibroblasts induced by epidermal growth factor (EGF), platelet-derived growth factor, basic fibroblast growth factor, phorbol 12-myristate 13-acetate (PMA) and prostaglandin E{sub 2} without any appreciable cytotoxic effect. Other HDAC inhibitors, including butyric acid and trichostatin A (TSA), showed similar inhibitory effects on HGF production stimulated by various inducers. Up-regulations of HGF gene expression induced by PMA and EGF were also suppressed by VPA and TSA. Furthermore, VPA significantly inhibited HGF-induced invasion of HepG2 hepatocellular carcinoma cells. VPA, however, did not affect the increases in phosphorylation of MAPK and Akt in HGF-treated HepG2 cells. These results demonstrated that VPA inhibited two critical processes of tumor-stromal interaction, induction of fibroblastic HGF production and HGF-induced invasion of HepG2 cells, and suggest that those activities serve for other anti-tumor mechanisms of VPA besides causing proliferation arrest, differentiation, and/or apoptosis of tumor cells.

  5. HB-EGF directs stromal cell polyploidy and decidualization via cyclin D3 during implantation.

    PubMed

    Tan, Yi; Li, Meiling; Cox, Sandra; Davis, Marilyn K; Tawfik, Ossama; Paria, Bibhash C; Das, Sanjoy K

    2004-01-01

    Stromal cell polyploidy is a unique phenomenon that occurs during uterine decidualization following embryo implantation, although the developmental mechanism still remains elusive. The general consensus is that the aberrant expression and altered functional activity of cell cycle regulatory molecules at two particular checkpoints G1 to S and G2 to M in the cell cycle play an important role in the development of cellular polyploidy. Despite the compelling evidence of intrinsic cell cycle alteration, it has been implicated that the development of cellular polyploidy may be controlled by specific actions of extracellular growth regulators. Here we show a novel role for heparin-binding EGF-like growth factor (HB-EGF) in the developmental process of stromal cell polyploidy in mice. HB-EGF, which is one of the earliest known molecular mediators of implantation in mice and humans, promotes stromal cell polyploidy via upregulation of cyclin D3. Adenoviral delivery of antisense cyclin D3 attenuates cyclin D3 expression and abrogates HB-EGF-induced stromal cell polyploidy in vitro and in vivo. Collectively, the results demonstrate that the regulation of stromal cell polyploidy and decidualization induced by HB-EGF depend on cyclin D3 induction.

  6. Expression of Pitx2 in stromal cells is required for normal hematopoiesis.

    PubMed

    Kieusseian, Aurélie; Chagraoui, Jalila; Kerdudo, Cécile; Mangeot, Philippe-Emmanuel; Gage, Philip J; Navarro, Nicole; Izac, Brigitte; Uzan, Georges; Forget, Bernard G; Dubart-Kupperschmitt, Anne

    2006-01-15

    Although the expression of Pitx2, a bicoid family homeodomain transcription factor, is highly regulated during hematopoiesis, its function during this process was not documented; we thus studied hematopoiesis in Pitx2-null mice. We found that Pitx2(-/-) embryos display hypoplastic livers with reduced numbers of hematopoietic cells, but these cells had normal hematopoietic potential, as evidenced by colony-forming assays, immature progenitor cell assays, and long-term repopulation assays. Because the microenvironment is also crucial to the development of normal hematopoiesis, we established Pitx2(-/-) and Pitx2(+/+) stromas from fetal liver and studied their hematopoietic supportive capacity. We showed that the frequency of cobblestone area-forming cells was 4-fold decreased when using Pitx2(-/-) stromal cells compared with Pitx2(+/+) stromal cells, whatever the Pitx2 genotype of hematopoietic cells tested in this assay. This defect was rescued by expression of Pitx2 into Pitx2(-/-) fetal liver stromal cells, demonstrating a major and direct role of Pitx2 in the hematopoietic supportive capacity of fetal liver stroma. Finally, we showed a reduced capacity of MS5 stromal cells expressing Pitx2 RNAi to support human hematopoiesis. Altogether these data showed that Pitx2 has major functions in the hematopoietic supportive capacity of fetal liver and adult bone marrow stromal cells.

  7. Expression of Pitx2 in stromal cells is required for normal hematopoiesis

    PubMed Central

    Kieusseian, Aurélie; Chagraoui, Jalila; Kerdudo, Cécile; Mangeot, Philippe-Emmanuel; Gage, Philip J.; Navarro, Nicole; Izac, Brigitte; Uzan, Georges; Forget, Bernard G.; Dubart-Kupperschmitt, Anne

    2006-01-01

    Although the expression of Pitx2, a bicoid family homeodomain transcription factor, is highly regulated during hematopoiesis, its function during this process was not documented; we thus studied hematopoiesis in Pitx2-null mice. We found that Pitx2–/– embryos display hypoplastic livers with reduced numbers of hematopoietic cells, but these cells had normal hematopoietic potential, as evidenced by colony-forming assays, immature progenitor cell assays, and long-term repopulation assays. Because the microenvironment is also crucial to the development of normal hematopoiesis, we established Pitx2–/– and Pitx2+/+ stromas from fetal liver and studied their hematopoietic supportive capacity. We showed that the frequency of cobblestone area-forming cells was 4-fold decreased when using Pitx2–/– stromal cells compared with Pitx2+/+ stromal cells, whatever the Pitx2 genotype of hematopoietic cells tested in this assay. This defect was rescued by expression of Pitx2 into Pitx2–/– fetal liver stromal cells, demonstrating a major and direct role of Pitx2 in the hematopoietic supportive capacity of fetal liver stroma. Finally, we showed a reduced capacity of MS5 stromal cells expressing Pitx2 RNAi to support human hematopoiesis. Altogether these data showed that Pitx2 has major functions in the hematopoietic supportive capacity of fetal liver and adult bone marrow stromal cells. PMID:16195330

  8. Pancreatic adenocarcinoma upregulated factor (PAUF) confers resistance to pancreatic cancer cells against oncolytic parvovirus H-1 infection through IFNA receptor-mediated signaling

    SciTech Connect

    Kaowinn, Sirichat; Cho, Il-Rae; Moon, Jeong; Jun, Seung Won; Kim, Chang Seok; Kang, Ho Young; Kim, Manbok; Koh, Sang Seok; Chung, Young-Hwa

    2015-04-03

    Pancreatic adenocarcinoma upregulated factor (PAUF), a novel oncogene, plays a crucial role in the development of pancreatic cancer, including its metastasis and proliferation. Therefore, PAUF-expressing pancreatic cancer cells could be important targets for oncolytic virus-mediated treatment. Panc-1 cells expressing PAUF (Panc-PAUF) showed relative resistance to parvovirus H-1 infection compared with Panc-1 cells expressing an empty vector (Panc-Vec). Of interest, expression of type I IFN-α receptor (IFNAR) was higher in Panc-PAUF cells than in Panc-Vec cells. Increased expression of IFNAR in turn increased the activation of Stat1 and Tyk2 in Panc-PAUF cells compared with that in Panc-Vec cells. Suppression of Tyk2 and Stat1, which are important downstream molecules for IFN-α signaling, sensitized pancreatic cancer cells to parvovirus H-1-mediated apoptosis. Further, constitutive suppression of PAUF sensitized Bxpc3 pancreatic cancer cells to parvovirus H-1 infection. Taken together, these results suggested that PAUF conferred resistance to pancreatic cancer cells against oncolytic parvovirus H-1 infection through IFNAR-mediated signaling. - Highlights: • PAUF confers resistance against oncolytic parvovirus H-1 infection. • PAUF enhances the expression of IFNAR in Panc-1 cells. • Increased activation of Tyk2 or Stat1 by PAUF provides resistance to parvovirus H-1-mediated apoptosis. • Constitutive inhibition of PAUF enhances parvovirus H-1-mediated oncolysis of Bxpc3 pancreatic cancer cells.

  9. The sunflower transcription factor HaHB11 confers tolerance to water deficit and salinity to transgenic Arabidopsis and alfalfa plants.

    PubMed

    Cabello, Julieta V; Giacomelli, Jorge I; Gómez, María C; Chan, Raquel L

    2017-09-10

    Homeodomain-leucine zipper (HD-Zip) transcription factors are unique to the plant kingdom; members of subfamily I are known to be involved in abiotic stress responses. HaHB11 belongs to this subfamily and it was previously shown that it is able to confer improved yield and tolerance to flooding via a quiescent strategy. Here we show that HaHB11 expression is induced by ABA, NaCl and water deficit in sunflower seedlings and leaves. Arabidopsis transgenic plants expressing HaHB11, controlled either by its own promoter or by the constitutive 35S CaMV, presented rolled leaves and longer roots than WT when grown under standard conditions. In addition, these plants showed wider stems and more vascular bundles. To deal with drought, HaHB11 transgenic plants closed their stomata faster and lost less water than controls, triggering an enhanced tolerance to such stress condition and also to salinity stress. Concomitantly, ABA-synthesis and sensing related genes were differentially regulated in HaHB11 transgenic plants. Either under long-term salinity stress or mild drought stress, HaHB11 transgenic plants did not exhibit yield penalties. Moreover, alfalfa transgenic plants were generated which also showed enhanced drought tolerance. Altogether, the results indicated that HaHB11 was able to confer drought and salinity tolerance via a complex mechanism which involves morphological, physiological and molecular changes. Copyright © 2016 Elsevier B.V. All rights reserved.

  10. Ectopic expression of Xylella fastidiosa rpfF conferring production of diffusible signal factor in transgenic tobacco and citrus alters pathogen behavior and reduces disease severity.

    PubMed

    Caserta, Raquel; Souza-Neto, Reinaldo Rodrigues; Takita, Marco Aurelio; Lindow, Steven; Souza, Alessandra De

    2017-08-04

    The pathogenicity of Xylella fastidiosa is associated with its ability to colonize the xylem of host plants. Expression of genes contributing to xylem colonization are suppressed, while those necessary for insect vector acquisition are increased, with increasing concentrations of diffusible signal factor (DSF) whose production is dependent on RpfF. We previously demonstrated that transgenic citrus plants ectopically expressing rpfF from a citrus strain of X. fastidiosa subsp. pauca exhibited less susceptibility to Xanthomonas citri subsp. citri, another pathogen whose virulence is modulated by DSF accumulation. Here we demonstrate that ectopic expression of rpfF in both transgenic tobacco and sweet orange also confers a reduction in disease severity incited by X. fastidiosa and reduces its colonization of those plants. Decreased disease severity in the transgenic plants was generally associated with increased expression of genes conferring adhesiveness to the pathogen, and decreased expression of genes necessary for active motility, accounting for the reduced population sizes achieved in the plants, apparently by limiting pathogen dispersal through the plant. Plant-derived DSF signal molecules in a host plant therefore can be exploited to interfere with more than one pathogen whose virulence is controlled by DSF signaling.

  11. Expression of vascular endothelial growth factor does not promote transformation but confers a growth advantage in vivo to Chinese hamster ovary cells.

    PubMed Central

    Ferrara, N; Winer, J; Burton, T; Rowland, A; Siegel, M; Phillips, H S; Terrell, T; Keller, G A; Levinson, A D

    1993-01-01

    Vascular endothelial growth factor (VEGF) is a mitogen with a specificity for endothelial cells in vitro and an angiogenic inducer in vivo. We tested the hypothesis that VEGF may confer on expressing cells a growth advantage in vivo. Dihydrofolatereductase--Chinese hamster ovary cells were transfected with expression vectors which direct the constitutive synthesis of VEGF. Neither the expression nor the exogenous administration of VEGF stimulated anchorage-dependent or anchorage-independent growth of Chinese hamster ovary cells in vitro. However, VEGF-expressing clones, unlike control cells, demonstrated an ability to proliferate in nude mice. Histologic examination revealed that the proliferative lesions were compact, well vascularized, and nonedematous. Ultrastructural analysis revealed that capillaries within the lesions were of the continuous type. These findings indicate that the expression of VEGF may confer on cells the ability to grow in vivo in the absence of transformation by purely paracrine mechanisms. Since VEGF is a widely distributed protein, this property may have relevance for a variety of physiological and pathological proliferative processes. Images PMID:8423215

  12. The stromal cell-derived factor-1alpha dependent migration of human cord blood CD34 haematopoietic stem and progenitor cells switches from protein kinase C (PKC)-alpha dependence to PKC-alpha independence upon prolonged culture in the presence of Flt3-ligand and interleukin-6.

    PubMed

    Kasenda, Benjamin; Kassmer, Susannah H; Niggemann, Bernd; Schiermeier, Sven; Hatzmann, Wolfgang; Zänker, Kurt S; Dittmar, Thomas

    2008-09-01

    Addition of the inflammatory cytokine interleukin (IL)-6 to the culture medium of human cord blood haematopoietic stem and progenitor cells (HSPCs) has been shown to lead to an altered stromal cell-derived factor-1alpha-dependent migratory phenotype. This study investigated whether this effect was attributed to a differential engagement of protein kinase C (PKC) isotypes. The migratory activity of both Flt3-ligand and Flt3-ligand/IL-6 cultured cord blood HSPCs was PKC-alpha dependent on day 1, but PKC-alpha independent after 5 d of cultivation. PKC-alpha expression was not down-regulated in cells cultured for 5 d indicating a switch of signalling molecules directing cell migration.

  13. Human-derived normal mesenchymal stem/stromal cells in anticancer therapies

    PubMed Central

    Zhang, Cheng; Yang, Shi-Jie; Wen, Qin; Zhong, Jiang F; Chen, Xue-Lian; Stucky, Andres; Press, Michael F; Zhang, Xi

    2017-01-01

    The tumor microenvironment (TME) not only plays a pivotal role during cancer progression and metastasis, but also has profound effects on therapeutic efficacy. Stromal cells of the TME are increasingly becoming a key consideration in the development of active anticancer therapeutics. However, dispute concerning the role of stromal cells to fight cancer continues because the use of mesenchymal stem/stromal cells (MSCs) as an anticancer agent is dependent on the specific MSCs subtype, in vitro or in vivo conditions, factors secreted by MSCs, types of cancer cell lines and interactions between MSCs, cancer cells and host immune cells. In this review, we mainly focus on the role of human-derived normal MSCs in anticancer therapies. We first discuss the use of different MSCs in the therapies for various cancers. We then focus on their anticancer mechanism and clinical application. PMID:28123601

  14. Cell adhesion and apoptosis in ovarian stromal hyperplasia and hyperthecosis.

    PubMed

    Sharabidze, N; Burkadze, G; Sabakhtarashvili, M

    2006-02-01

    The aim of our study was to investigate cell adhesion and apoptosis in ovarian stromal hyperplasia and hyperthecosis in reproductive women with and without polycystic ovarian disease. We have studied 104 patients with a histological diagnosis of ovarian stromal hyperthecosis and stromal hyperplasia. Paraffin sections were stained by hematoxylin-eosin, von Gieson and immunohistochemistry for Bcl-2 (anti-apoptotic protein) and E-cadherin (cell adhesion marker). We assessed the number of Bcl-2-positive and E-cadherin-positive cells. The patients were divided into 4 groups: group 1-33 patients with polycystic ovarian disease and coexistent stromal hyperthecosis, group 2-28 patients with polycystic ovarian disease and coexistent stromal hyperplasia, group 3-24 patients with ovarian stromal hyperthecosis, group 4-19 patients with ovarian stromal hyperplasia. Our results suggest that in ovarian stromal hyperthecosis and stromal hyperplasia coexistent with polycystic ovarian disease, E-cadherin-positivity in internal and external theca cells, and granulosa cells is associated with Bcl-2 expression. Therefore, ovarian cells expressing Bcl-2 and maintaining E-cadherin-positivity may be the viable cells that escape the apoptotic process. In ovarian stromal hyperthecosis without polycystic ovarian disease, luteinized stromal cells are potentially resistant to apoptosis as they are positive for Bcl-2. In ovarian stromal hyperplasia without polycystic ovarian disease, hyperplastic stromal cells are potentially susceptible to apoptosis as they are negative for Bcl-2. E-cadherin is negative both in stromal hyperthecosis and hyperplasia suggesting that E-cadherin expression in ovary is limited to granulosa and theca cells only. Described characteristics of cell adhesion and apoptosis may play a role in pathogenesis of ovarian stromal hyperthecosis and stromal hyperplasia with and without polycystic ovarian disease.

  15. Hepatic immune regulation by stromal cells.

    PubMed

    Schildberg, Frank A; Sharpe, Arlene H; Turley, Shannon J

    2015-02-01

    A metabolic organ, the liver also has a central role in tolerance induction. Stromal cells lining the hepatic sinusoids, such as liver sinusoidal endothelial cells (LSECs) and hepatic stellate cells (HSCs), are the first liver cells to encounter gut-derived and systemic antigens, thereby shaping local and systemic tolerance. Recent studies have demonstrated that stromal cells can modulate immune responses by antigen-dependent and independent mechanisms. Stromal cells interfere with the function of other antigen-presenting cells (APCs) and induce non-responsive T cells as well as regulatory T cells and myeloid-derived suppressor cells (MDSCs). The immunosuppressive microenvironment thus created provides a means to protect the liver from tissue damage. Such tolerized surroundings, however, can be exploited by certain pathogens, promoting persistent liver infections.

  16. Conference reports

    NASA Astrophysics Data System (ADS)

    Dongpei, Chen; Yulong, Ma

    1994-12-01

    The Ultrasonic Electronics Branch Society of the China Acoustics Society, and the Electronics Countermeasure Branch Society of the China Electronics Society held and All-China Applications Conference of Ultrasonic Electronics Devices in Electronic Countermeasures, Radar and Military Communication Technology. A total of 66 papers was received by the conference with contents relating to surface acoustic wave devices, high-frequency acoustic wave devices, acousto-optical devices, applications of devices in radar, applications of devices in electronic countermeasures, and applications of devices in military communication systems.

  17. Low-frequency vibration treatment of bone marrow stromal cells induces bone repair in vivo

    PubMed Central

    He, Shengwei; Zhao, Wenzhi; Zhang, Lu; Mi, Lidong; Du, Guangyu; Sun, Chuanxiu; Sun, Xuegang

    2017-01-01

    Objective(s): To study the effect of low-frequency vibration on bone marrow stromal cell differentiation and potential bone repair in vivo. Materials and Methods: Forty New Zealand rabbits were randomly divided into five groups with eight rabbits in each group. For each group, bone defects were generated in the left humerus of four rabbits, and in the right humerus of the other four rabbits. To test differentiation, bones were isolated and demineralized, supplemented with bone marrow stromal cells, and implanted into humerus bone defects. Varying frequencies of vibration (0, 12.5, 25, 50, and 100 Hz) were applied to each group for 30 min each day for four weeks. When the bone defects integrated, they were then removed for histological examination. mRNA transcript levels of runt-related transcription factor 2, osteoprotegerin, receptor activator of nuclear factor κ-B ligan, and pre-collagen type 1 α were measured. Results: Humeri implanted with bone marrow stromal cells displayed elevated callus levels and wider, more prevalent, and denser trabeculae following treatment at 25 and 50 Hz. The mRNA levels of runt-related transcription factor 2, osteoprotegerin, receptor activator of nuclear factor κ-B ligand, and pre-collagen type 1 α were also markedly higher following 25 and 50 Hz treatment. Conclusion: Low frequency (25–50 Hz) vibration in vivo can promote bone marrow stromal cell differentiation and repair bone injury. PMID:28133520

  18. Low-frequency vibration treatment of bone marrow stromal cells induces bone repair in vivo.

    PubMed

    He, Shengwei; Zhao, Wenzhi; Zhang, Lu; Mi, Lidong; Du, Guangyu; Sun, Chuanxiu; Sun, Xuegang

    2017-01-01

    To study the effect of low-frequency vibration on bone marrow stromal cell differentiation and potential bone repair in vivo. Forty New Zealand rabbits were randomly divided into five groups with eight rabbits in each group. For each group, bone defects were generated in the left humerus of four rabbits, and in the right humerus of the other four rabbits. To test differentiation, bones were isolated and demineralized, supplemented with bone marrow stromal cells, and implanted into humerus bone defects. Varying frequencies of vibration (0, 12.5, 25, 50, and 100 Hz) were applied to each group for 30 min each day for four weeks. When the bone defects integrated, they were then removed for histological examination. mRNA transcript levels of runt-related transcription factor 2, osteoprotegerin, receptor activator of nuclear factor κ-B ligan, and pre-collagen type 1 α were measured. Humeri implanted with bone marrow stromal cells displayed elevated callus levels and wider, more prevalent, and denser trabeculae following treatment at 25 and 50 Hz. The mRNA levels of runt-related transcription factor 2, osteoprotegerin, receptor activator of nuclear factor κ-B ligand, and pre-collagen type 1 α were also markedly higher following 25 and 50 Hz treatment. Low frequency (25-50 Hz) vibration in vivo can promote bone marrow stromal cell differentiation and repair bone injury.

  19. Interleukin-6 receptor in spindle-shaped stromal cells, a prognostic determinant of early breast cancer.

    PubMed

    Labovsky, Vivian; Martinez, Leandro Marcelo; Calcagno, María de Luján; Davies, Kevin Mauro; García-Rivello, Hernán; Wernicke, Alejandra; Feldman, Leonardo; Giorello, María Belén; Matas, Ayelén; Borzone, Francisco Raúl; Howard, Scott C; Chasseing, Norma Alejandra

    2016-10-01

    Spindle-shaped stromal cells, like carcinoma-associated fibroblasts and mesenchymal stem cells, influence tumor behavior and can serve as parameters in the clinical diagnosis, therapy, and prognosis of early breast cancer. Therefore, the aim of this study is to explore the clinicopathological significance of tumor necrosis factor-related apoptosis-induced ligand (TRAIL) receptors (Rs) 2 and 4 (TRAIL-R2 and R4), and interleukin-6 R (IL-6R) in spindle-shaped stromal cells, not associated with the vasculature, as prognostic determinants of early breast cancer patients. Receptors are able to trigger the migratory activity, among other functions, of these stromal cells. We conducted immunohistochemical analysis for the expression of these receptors in spindle-shaped stromal cells, not associated with the vasculature, of primary tumors from early invasive breast cancer patients, and analyzed their association with clinicopathological characteristics. Here, we demonstrate that the elevated levels of TRAIL-R2, TRAIL-R4, and IL-6R in these stromal cells were significantly associated with a higher risk of metastatic occurrence (p = 0.034, 0.026, and 0.006; respectively). Moreover, high expression of TRAIL-R4 was associated with shorter disease-free survival and metastasis-free survival (p = 0.013 and 0.019; respectively). Also, high expression of IL-6R was associated with shorter disease-free survival, metastasis-free survival, and overall survival (p = 0.003, 0.001, and 0.003; respectively). Multivariate analysis showed that IL-6R expression was an independent prognostic factor for disease-free survival and metastasis-free survival (p = 0.035). This study is the first to demonstrate that high levels of IL-6R expression in spindle-shaped stromal cells, not associated with the vasculature, could be used to identify early breast cancer patients with poor outcomes.

  20. Plant nuclear factor Y (NF-Y) B subunits confer drought tolerance and lead to improved corn yields on water-limited acres.

    PubMed

    Nelson, Donald E; Repetti, Peter P; Adams, Tom R; Creelman, Robert A; Wu, Jingrui; Warner, David C; Anstrom, Don C; Bensen, Robert J; Castiglioni, Paolo P; Donnarummo, Meghan G; Hinchey, Brendan S; Kumimoto, Roderick W; Maszle, Don R; Canales, Roger D; Krolikowski, Katherine A; Dotson, Stanton B; Gutterson, Neal; Ratcliffe, Oliver J; Heard, Jacqueline E

    2007-10-16

    Commercially improved crop performance under drought conditions has been challenging because of the complexity of the trait and the multitude of factors that influence yield. Here we report the results of a functional genomics approach that identified a transcription factor from the nuclear factor Y (NF-Y) family, AtNF-YB1, which acts through a previously undescribed mechanism to confer improved performance in Arabidopsis under drought conditions. An orthologous maize transcription factor, ZmNF-YB2, is shown to have an equivalent activity. Under water-limited conditions, transgenic maize plants with increased ZmNF-YB2 expression show tolerance to drought based on the responses of a number of stress-related parameters, including chlorophyll content, stomatal conductance, leaf temperature, reduced wilting, and maintenance of photosynthesis. These stress adaptations contribute to a grain yield advantage to maize under water-limited environments. The application of this technology has the potential to significantly impact maize production systems that experience drought.

  1. Bone marrow stromal cells from multiple myeloma patients uniquely induce bortezomib resistant NF-κB activity in myeloma cells

    PubMed Central

    2010-01-01

    Background Components of the microenvironment such as bone marrow stromal cells (BMSCs) are well known to support multiple myeloma (MM) disease progression and resistance to chemotherapy including the proteasome inhibitor bortezomib. However, functional distinctions between BMSCs in MM patients and those in disease-free marrow are not completely understood. We and other investigators have recently reported that NF-κB activity in primary MM cells is largely resistant to the proteasome inhibitor bortezomib, and that further enhancement of NF-κB by BMSCs is similarly resistant to bortezomib and may mediate resistance to this therapy. The mediating factor(s) of this bortezomib-resistant NF-κB activity is induced by BMSCs is not currently understood. Results Here we report that BMSCs specifically derived from MM patients are capable of further activating bortezomib-resistant NF-κB activity in MM cells. This induced activity is mediated by soluble proteinaceous factors secreted by MM BMSCs. Among the multiple factors evaluated, interleukin-8 was secreted by BMSCs from MM patients at significantly higher levels compared to those from non-MM sources, and we found that IL-8 contributes to BMSC-induced NF-κB activity. Conclusions BMSCs from MM patients uniquely enhance constitutive NF-κB activity in MM cells via a proteinaceous secreted factor in part in conjunction with IL-8. Since NF-κB is known to potentiate MM cell survival and confer resistance to drugs including bortezomib, further identification of the NF-κB activating factors produced specifically by MM-derived BMSCs may provide a novel biomarker and/or drug target for the treatment of this commonly fatal disease. PMID:20604947

  2. The Alfin-like homeodomain finger protein AL5 suppresses multiple negative factors to confer abiotic stress tolerance in Arabidopsis.

    PubMed

    Wei, Wei; Zhang, Yu-Qin; Tao, Jian-Jun; Chen, Hao-Wei; Li, Qing-Tian; Zhang, Wan-Ke; Ma, Biao; Lin, Qing; Zhang, Jin-Song; Chen, Shou-Yi

    2015-03-01

    Plant homeodomain (PHD) finger proteins affect processes of growth and development by changing transcription and reading epigenetic histone modifications, but their functions in abiotic stress responses remain largely unclear. Here we characterized seven Arabidopsis thaliana Alfin1-like PHD finger proteins (ALs) in terms of the responses to abiotic stresses. ALs localized to the nucleus and repressed transcription. Except AL6, all the ALs bound to G-rich elements. Mutations of the amino acids at positions 34 and 35 in AL6 caused loss of ability to bind to G-rich elements. Expression of the AL genes responded differentially to osmotic stress, salt, cold and abscisic acid treatments. AL5-over-expressing plants showed higher tolerance to salt, drought and freezing stress than Col-0. Consistently, al5 mutants showed reduced stress tolerance. We used ChIP-Seq assays to identify eight direct targets of AL5, and found that AL5 binds to the promoter regions of these genes. Knockout mutants of five of these target genes exhibited varying tolerances to stresses. These results indicate that AL5 inhibits multiple signaling pathways to confer stress tolerance. Our study sheds light on mechanisms of AL5-mediated signaling in abiotic stress responses, and provides tools for improvement of stress tolerance in crop plants.

  3. Stromal cell-derived CXCL12 and CCL8 cooperate to support increased development of regulatory dendritic cells following Leishmania infection.

    PubMed

    Nguyen Hoang, Anh Thu; Liu, Hao; Juaréz, Julius; Aziz, Naveed; Kaye, Paul M; Svensson, Mattias

    2010-08-15

    In the immune system, stromal cells provide specialized niches that control hematopoiesis by coordinating the production of chemokines, adhesion molecules, and growth factors. Stromal cells also have anti-inflammatory effects, including support for the differentiation of hematopoietic progenitors into dendritic cells (DCs) with immune regulatory properties. Together, these observations suggest that the alterations in hematopoiesis commonly seen in infectious disease models, such as experimental visceral leishmaniasis in mice, might result from altered stromal cell function. We report in this study that the stromal cell-derived chemokines CXCL12 and CCL8 cooperate to attract hematopoietic progenitors with the potential to differentiate into regulatory DCs. We also show that infection of murine bone marrow stromal cells by Leishmania donovani enhanced their capacity to support the development of regulatory DCs, as well as their capacity to produce CCL8. Likewise, in experimental visceral leishmaniasis, CCL8 production was induced in splenic stromal cells, leading to an enhanced capacity to attract hematopoietic progenitor cells. Thus, intracellular parasitism of stromal cells modifies their capacity to recruit and support hematopoietic progenitor differentiation into regulatory DCs, and aberrant expression of CCL8 by diseased stromal tissue may be involved in the switch from resolving to persistent infection.

  4. Conference Summary

    ERIC Educational Resources Information Center

    Doherty, Cait

    2009-01-01

    This article summarizes an original conference, organised by the Child Care Research Forum (http://www.qub.ac.uk/sites/ccrf/), which brought together experts from all over Northern Ireland to showcase some of the wealth of research with children and young people that is going on in the country today. Developed around the six high-level outcomes of…

  5. The conference

    Treesearch

    Gordon M. Heisler; Lee P. Herrington

    1977-01-01

    This is a report on the Conference on Metropolitan Physical Environment, held in August 1975 at Syracuse, N.Y., where some 160 scientists and planners met to discuss the use of vegetation, space, and structures to improve the amenities for people who live in metropolitan areas.

  6. Conference Space

    ERIC Educational Resources Information Center

    Tillett, Wade

    2016-01-01

    The following is an exploration of the spatial configurations (and their implications) within a typical panel session at an academic conference. The presenter initially takes up different roles and hyperbolically describes some possible messages that the spatial arrangement sends. Eventually, the presenter engages the audience members in atypical…

  7. Conference Space

    ERIC Educational Resources Information Center

    Tillett, Wade

    2016-01-01

    The following is an exploration of the spatial configurations (and their implications) within a typical panel session at an academic conference. The presenter initially takes up different roles and hyperbolically describes some possible messages that the spatial arrangement sends. Eventually, the presenter engages the audience members in atypical…

  8. Deletion of the znuA virulence factor attenuates Actinobacillus pleuropneumoniae and confers protection against homologous or heterologous strain challenge.

    PubMed

    Yuan, Fangyan; Liao, Yonghong; You, Wujin; Liu, Zewen; Tan, Yongqiang; Zheng, Chengkun; BinWang; Zhou, Danna; Tian, Yongxiang; Bei, Weicheng

    2014-12-05

    The znuA gene is known to be important for growth and survival in Escherichia coli, Haemophilus spp., Neisseria gonorrhoeae, and Pasteurella multocida under low Zn(2+) conditions. This gene is also present in Actinobacillus pleuropneumoniae serotype 1; therefore, the aim of this study was to investigate the existence of a similar role for the znuA gene in the growth and virulence of this organism. A precisely defined ΔznuA deletion mutant of A. pleuropneumoniae was constructed based on the sequence of the wild-type SLW01 using transconjugation and counterselection. This mutation was found to be lethal in low-Zn(2+) medium. Furthermore, the ΔznuA mutant strain exhibited attenuated virulence (≥22-fold) as well as reduced mortality and morbidity in a murine (Balb/C) model of infection. The majority of the bacteria were cleared from the lungs within 2 weeks. The ΔznuA mutant strain caused no adverse effects in pigs at doses of up to 1.0×10(9) CFU/mL. The ΔznuA mutant strain induced a significant immune response and conferred 80% and 100% protection on immunised pigs against challenge with A. pleuropneumoniae strains belonging to homologous or heterologous serovars, respectively, compared to the blank controls. The data obtained in this study indicate the potential of the mutant ΔznuA strain for development as a live vaccine capable of inducing reliable cross-serovar protection following intratracheal immunisation. Copyright © 2014 Elsevier B.V. All rights reserved.

  9. Stromal-dependent tumor promotion by MIF family members

    PubMed Central

    Mitchell, Robert A.; Yaddanapudi, Kavitha

    2014-01-01

    Solid tumors are composed of a heterogeneous population of cells that interact with each other and with soluble and insoluble factors that, when combined, strongly influence the relative proliferation, differentiation, motility, matrix remodeling, metabolism and microvessel density of malignant lesions. One family of soluble factors that is becoming increasingly associated with pro-tumoral phenotypes within tumor microenvironments is that of the migration inhibitory factor family which includes its namesake, MIF, and its only known family member, D-dopachrome tautomerase (D-DT). This review seeks to highlight our current understanding of the relative contributions of a variety of immune and non-immune tumor stromal cell populations and, within those contexts, will summarize the literature associated with MIF and/or D-DT. PMID:25277536

  10. Origin of hemopoietic stromal progenitor cells in chimeras

    SciTech Connect

    Chertkov, J.L.; Drize, N.J.; Gurevitch, O.A.; Samoylova, R.S.

    1985-12-01

    Intravenously injected bone marrow cells do not participate in the regeneration of hemopoietic stromal progenitors in irradiated mice, nor in the curetted parts of the recipient's marrow. The hemopoietic stromal progenitors in allogeneic chimeras are of recipient origin. The adherent cell layer (ACL) of long-term cultures of allogeneic chimera bone marrow contains only recipient hemopoietic stromal progenitors. However, in ectopic hemopoietic foci produced by marrow implantation under the renal capsule and repopulated by the recipient hemopoietic cells after irradiation and reconstitution by syngeneic hemopoietic cells, the stromal progenitors were of implant donor origin, as were stromal progenitors of the ACL in long-term cultures of hemopoietic cells from ectopic foci. Our results confirm that the stromal and hemopoietic progenitors differ in origin and that hemopoietic stromal progenitors are not transplantable by the intravenous route in mice.

  11. Skull metastasis from rectal gastrointestinal stromal tumours.

    PubMed

    Gil-Arnaiz, Irene; Martínez-Trufero, Javier; Pazo-Cid, Roberto Antonio; Felipo, Francesc; Lecumberri, María José; Calderero, Verónica

    2009-09-01

    Gastrointestinal stromal tumours (GIST) are the most common mesenchymal neoplasm of the gastrointestinal tract. Rectum localisation is infrequent for these neoplasms, accounting for about 5% of all cases. Distant metastases of GIST are also rare. We present a patient with special features: the tumour is localised in rectum and it has an uncommon metastatic site, the skull, implying a complex differential diagnosis approach.

  12. Bone-stromal cells up-regulate tumourigenic markers in a tumour-stromal 3D model of prostate cancer.

    PubMed

    Windus, Louisa C E; Glover, Tristan T; Avery, Vicky M

    2013-09-30

    The cellular and molecular mechanisms that mediate interactions between tumour cells and the surrounding bone stroma are to date largely undetermined in prostate cancer (PCa) progression. The purpose of this study was to evaluate the role of alpha 6 and beta 1 integrin subunits in mediating tumour-stromal interactions. Utilising 3D in vitro assays we evaluated and compared 1. Monocultures of prostate metastatic PC3, bone stromal derived HS5 and prostate epithelial RWPE-1 cells and 2. Tumour-stromal co-cultures (PC3 + HS5) to ascertain changes in cellular phenotype, function and expression of metastatic markers. In comparison to 3D monocultures of PC3 or HS5 cells, when cultured together, these cells displayed up-regulated invasive and proliferative qualities, along with altered expression of epithelial-to-mesenchymal and chemokine protein constituents implicated in metastatic dissemination. When co-cultured, HS5 cells were found to re-express N-Cadherin and chemokine receptor CXCR7. Alterations in N-Cadherin expression were found to be mediated by soluble factors secreted by PC3 tumour cells, while chemokine receptor re-expression was dependent on direct cell-cell interactions. We have also shown that integrins beta 1 and alpha 6 play an integral role in maintaining cell homeostasis and mediating expression of E-Cadherin, N-Cadherin and vimentin, in addition to chemokine receptor CXCR7. Collectively our results suggest that both PC3 and HS5 cells provide a "protective" and reciprocal milieu that promotes tumour growth. As such 3D co-cultures may serve as a more complex and valid biological model in the drug discovery pipeline.

  13. Experiences Sustaining a Conference and Building a Network

    ERIC Educational Resources Information Center

    Grady, Marilyn L.

    2016-01-01

    The Women in Educational Leadership Conference (WELC) was founded at the University of Nebraska-Lincoln in 1987. The 28th conference was held in October 2014. A database related to the conference has been maintained throughout these years. Using these data, the following report includes the factors that have sustained the conference, the network…

  14. What Good Are Conferences, Anyway?

    ERIC Educational Resources Information Center

    Pietro, David C.

    1996-01-01

    According to Frederick Herzberg's studies of employee motivation, humans are driven by motivating factors that allow them to grow psychologically and hygiene factors that help them meet physical needs. Good education conferences can enhance both factors by helping principals refocus their energies, exchange ideas with trusted colleagues, and view…

  15. What Good Are Conferences, Anyway?

    ERIC Educational Resources Information Center

    Pietro, David C.

    1996-01-01

    According to Frederick Herzberg's studies of employee motivation, humans are driven by motivating factors that allow them to grow psychologically and hygiene factors that help them meet physical needs. Good education conferences can enhance both factors by helping principals refocus their energies, exchange ideas with trusted colleagues, and view…

  16. Inhibition of Stromal PlGF Suppresses the Growth of Prostate Cancer Xenografts

    PubMed Central

    Zins, Karin; Thomas, Anita; Lucas, Trevor; Sioud, Mouldy; Aharinejad, Seyedhossein; Abraham, Dietmar

    2013-01-01

    The growth and vascularization of prostate cancer is dependent on interactions between cancer cells and supporting stromal cells. The primary stromal cell type found in prostate tumors is the carcinoma-associated fibroblast, which produces placental growth factor (PlGF). PlGF is a member of the vascular endothelial growth factor (VEGF) family of angiogenic molecules and PlGF mRNA levels increase after androgen deprivation therapy in prostate cancer. In this study, we show that PlGF has a direct dose-dependent proliferative effect on human PC-3 prostate cancer cells in vitro and fibroblast-derived PlGF increases PC-3 proliferation in co-culture. In xenograft tumor models, intratumoral administration of murine PlGF siRNA reduced stromal-derived PlGF expression, reduced tumor burden and decreased the number of Ki-67 positive proliferating cells associated with reduced vascular density. These data show that targeting stromal PlGF expression may represent a therapeutic target for the treatment of prostate cancer. PMID:24005860

  17. Ectopic Overexpression of SsCBF1, a CRT/DRE-Binding Factor from the Nightshade Plant Solanum lycopersicoides, Confers Freezing and Salt Tolerance in Transgenic Arabidopsis

    PubMed Central

    Zhang, Lili; Li, Zhenjun; Li, Jingfu; Wang, Aoxue

    2013-01-01

    The C-repeat (CRT)/dehydration-responsive element (DRE) binding factor (CBF/DREB1) transcription factors play a key role in cold response. However, the detailed roles of many plant CBFs are far from fully understood. A CBF gene (SsCBF1) was isolated from the cold-hardy plant Solanum lycopersicoides. A subcellular localization study using GFP fusion protein indicated that SsCBF1 is localized in the nucleus. We delimited the SsCBF1 transcriptional activation domain to the C-terminal segment comprising amino acid residues 193–228 (SsCBF1193–228). The expression of SsCBF1 could be dramatically induced by cold, drought and high salinity. Transactivation assays in tobacco leaves revealed that SsCBF1 could specifically bind to the CRT cis-elements in vivo to activate the expression of downstream reporter genes. The ectopic overexpression of SsCBF1 conferred increased freezing and high-salinity tolerance and late flowering phenotype to transgenic Arabidopsis. RNA-sequencing data exhibited that a set of cold and salt stress responsive genes were up-regulated in transgenic Arabidopsis. Our results suggest that SsCBF1 behaves as a typical CBF to contribute to plant freezing tolerance. Increased resistance to high-salinity and late flowering phenotype derived from SsCBF1 OE lines lend more credence to the hypothesis that plant CBFs participate in diverse physiological and biochemical processes related to adverse conditions. PMID:23755095

  18. Regenerative Potential of Mesenchymal Stromal Cells: Age-Related Changes

    PubMed Central

    Bruna, Flavia; Contador, David; Conget, Paulette; Erranz, Benjamín; Sossa, Claudia L.; Arango-Rodríguez, Martha L.

    2016-01-01

    Preclinical and clinical studies have shown that a therapeutic effect results from mesenchymal stromal cells (MSCs) transplant. No systematic information is currently available regarding whether donor age modifies MSC regenerative potential on cutaneous wound healing. Here, we evaluate whether donor age influences this potential. Two different doses of bone marrow MSCs (BM-MSCs) from young, adult, or old mouse donors or two doses of their acellular derivatives mesenchymal stromal cells (acd-MSCs) were intradermally injected around wounds in the midline of C57BL/6 mice. Every two days, wound healing was macroscopically assessed (wound closure) and microscopically assessed (reepithelialization, dermal-epidermal junction, skin appendage regeneration, granulation tissue, leukocyte infiltration, and density dermal collagen fibers) after 12 days from MSC transplant. Significant differences in the wound closure kinetic, quality, and healing of skin regenerated were observed in lesions which received BM-MSCs from different ages or their acd-MSCs compared to lesions which received vehicle. Nevertheless, our data shows that adult's BM-MSCs or their acd-MSCs were the most efficient for recovery of most parameters analyzed. Our data suggest that MSC efficacy was negatively affected by donor age, where the treatment with adult's BM-MSCs or their acd-MSCs in cutaneous wound promotes a better tissue repair/regeneration. This is due to their paracrine factors secretion. PMID:27247575

  19. Senescent stromal-derived osteopontin promotes preneoplastic cell growth.

    PubMed

    Pazolli, Ermira; Luo, Xianmin; Brehm, Sarah; Carbery, Kelly; Chung, Jun-Jae; Prior, Julie L; Doherty, Jason; Demehri, Shadmehr; Salavaggione, Lorena; Piwnica-Worms, David; Stewart, Sheila A

    2009-02-01

    Alterations in the tissue microenvironment collaborate with cell autonomous genetic changes to contribute to neoplastic progression. The importance of the microenvironment in neoplastic progression is underscored by studies showing that fibroblasts isolated from a tumor stimulate the growth of preneoplastic and neoplastic cells in xenograft models. Similarly, senescent fibroblasts promote preneoplastic cell growth in vitro and in vivo. Because senescent cells accumulate with age, their presence is hypothesized to facilitate preneoplastic cell growth and tumor formation in older individuals. To identify senescent stromal factors directly responsible for stimulating preneoplastic cell growth, we carried out whole-genome transcriptional profiling and compared senescent fibroblasts with their younger counterparts. We identified osteopontin (OPN) as one of the most highly elevated transcripts in senescent fibroblasts. Importantly, reduction of OPN protein levels by RNA interference did not affect senescence induction in fibroblasts; however, it dramatically reduced the growth-promoting activities of senescent fibroblasts in vitro and in vivo, showing that OPN is necessary for paracrine stimulation of preneoplastic cell growth. In addition, we found that recombinant OPN was sufficient to stimulate preneoplastic cell growth. Finally, we show that OPN is expressed in senescent stroma within preneoplastic lesions that arise following 7,12-dimethylbenz(a)anthracene/12-O-tetradecanoylphorbol-13-acetate treatment of mice, suggesting that stromal-derived OPN-mediated signaling events affect neoplastic progression.

  20. Senescent Stromal-Derived Osteopontin Promotes Preneoplastic Cell Growth

    PubMed Central

    Pazolli, Ermira; Luo, Xianmin; Brehm, Sarah; Carbery, Kelly; Chung, Jun-Jae; Prior, Julie L.; Doherty, Jason; Demehri, Shadmehr; Salavaggione, Lorena; Piwnica-Worms, David; Stewart, Sheila A.

    2008-01-01

    Alterations in the tissue microenvironment collaborate with cell autonomous genetic changes to contribute to neoplastic progression. The importance of the microenvironment in neoplastic progression is underscored by studies demonstrating that fibroblasts isolated from a tumor stimulate the growth of preneoplastic and neoplastic cells in xenograft models. Similarly, senescent fibroblasts promote preneoplastic cell growth in vitro and in vivo. Because senescent cells accumulate with age, their presence is hypothesized to facilitate preneoplastic cell growth and tumor formation in older individuals. To identify senescent stromal factors directly responsible for stimulating preneoplastic cell growth, we carried out whole genome transcriptional profiling and compared senescent fibroblasts to their younger counterparts. We identified osteopontin (OPN) as one of the most highly elevated transcripts in senescent fibroblasts. Importantly, reduction of OPN protein levels by RNAi did not impact senescence induction in fibroblasts; however, it dramatically reduced the growth-promoting activities of senescent fibroblasts in vitro and in vivo, demonstrating that OPN is necessary for paracrine stimulation of preneoplastic cell growth. In addition, we found that recombinant OPN was sufficient to stimulate preneoplastic cell growth. Finally, we demonstrate that OPN is expressed in senescent stroma within preneoplastic lesions that arise following DMBA/TPA treatment of mice, suggesting that stromal-derived OPN-mediated signaling events impact neoplastic progression. PMID:19155301

  1. Functional inhibition of mesenchymal stromal cells in acute myeloid leukemia.

    PubMed

    Geyh, S; Rodríguez-Paredes, M; Jäger, P; Khandanpour, C; Cadeddu, R-P; Gutekunst, J; Wilk, C M; Fenk, R; Zilkens, C; Hermsen, D; Germing, U; Kobbe, G; Lyko, F; Haas, R; Schroeder, T

    2016-03-01

    Hematopoietic insufficiency is the hallmark of acute myeloid leukemia (AML) and predisposes patients to life-threatening complications such as bleeding and infections. Addressing the contribution of mesenchymal stromal cells (MSC) to AML-induced hematopoietic failure we show that MSC from AML patients (n=64) exhibit significant growth deficiency and impaired osteogenic differentiation capacity. This was molecularly reflected by a specific methylation signature affecting pathways involved in cell differentiation, proliferation and skeletal development. In addition, we found distinct alterations of hematopoiesis-regulating factors such as Kit-ligand and Jagged1 accompanied by a significantly diminished ability to support CD34+ hematopoietic stem and progenitor cells in long-term culture-initiating cells (LTC-ICs) assays. This deficient osteogenic differentiation and insufficient stromal support was reversible and correlated with disease status as indicated by Osteocalcin serum levels and LTC-IC frequencies returning to normal values at remission. In line with this, cultivation of healthy MSC in conditioned medium from four AML cell lines resulted in decreased proliferation and osteogenic differentiation. Taken together, AML-derived MSC are molecularly and functionally altered and contribute to hematopoietic insufficiency. Inverse correlation with disease status and adoption of an AML-like phenotype after exposure to leukemic conditions suggests an instructive role of leukemic cells on bone marrow microenvironment.

  2. Anchored and soluble gangliosides contribute to myelosupportivity of stromal cells

    SciTech Connect

    Ziulkoski, Ana L.; Santos, Aline X.S. dos; Andrade, Claudia M.B.; Trindade, Vera M.T.; Daniotti, Jose Luis; Borojevic, Radovan; Guma, Fatima C.R.

    2009-10-09

    Stroma-mediated myelopoiesis depends upon growth factors and an appropriate intercellular microenvironment. Previous studies have demonstrated that gangliosides, produced by hepatic stromal cell types, are required for optimal myelosupportive function. Here, we compared the mielossuportive functions of a bone marrow stroma (S17) and skin fibroblasts (SF) regarding their ganglioside pattern of synthesis and shedding. The survival and proliferation of a myeloid precursor cell (FDC-P1) were used as reporter. Although the ganglioside synthesis of the two stromal cells was similar, their relative content and shedding were distinct. The ganglioside requirement for mielossuportive function was confirmed by the decreased proliferation of FDC-P1 cells in ganglioside synthesis-inhibited cultures and in presence of an antibody to GM3 ganglioside. The distinct mielossuportive activities of the S17 and SF stromata may be related to differences on plasma membrane ganglioside concentrations or to differences on the gangliosides shed and their subsequent uptake by myeloid cells, specially, GM3 ganglioside.

  3. Anchored and soluble gangliosides contribute to myelosupportivity of stromal cells.

    PubMed

    Ziulkoski, Ana L; dos Santos, Aline X S; Andrade, Cláudia M B; Trindade, Vera M T; Daniotti, José Luis; Borojevic, Radovan; Guma, Fátima C R

    2009-10-09

    Stroma-mediated myelopoiesis depends upon growth factors and an appropriate intercellular microenvironment. Previous studies have demonstrated that gangliosides, produced by hepatic stromal cell types, are required for optimal myelosupportive function. Here, we compared the mielossuportive functions of a bone marrow stroma (S17) and skin fibroblasts (SF) regarding their ganglioside pattern of synthesis and shedding. The survival and proliferation of a myeloid precursor cell (FDC-P1) were used as reporter. Although the ganglioside synthesis of the two stromal cells was similar, their relative content and shedding were distinct. The ganglioside requirement for mielossuportive function was confirmed by the decreased proliferation of FDC-P1 cells in ganglioside synthesis-inhibited cultures and in presence of an antibody to GM3 ganglioside. The distinct mielossuportive activities of the S17 and SF stromata may be related to differences on plasma membrane ganglioside concentrations or to differences on the gangliosides shed and their subsequent uptake by myeloid cells, specially, GM3 ganglioside.

  4. Microbicides 2006 conference

    PubMed Central

    Ramjee, Gita; Shattock, Robin; Delany, Sinead; McGowan, Ian; Morar, Neetha; Gottemoeller, Megan

    2006-01-01

    Current HIV/AIDS statistics show that women account for almost 60% of HIV infections in Sub-Saharan Africa. HIV prevention tools such as male and female condoms, abstinence and monogamy are not always feasible options for women due to various socio-economic and cultural factors. Microbicides are products designed to be inserted in the vagina or rectum prior to sex to prevent HIV acquisition. The biannual Microbicides conference took place in Cape Town, South Africa from 23–26 April 2006. The conference was held for the first time on the African continent, the region worst affected by the HIV/AIDS pandemic. The conference brought together a record number of 1,300 scientists, researchers, policy makers, healthcare workers, communities and advocates. The conference provided an opportunity for an update on microbicide research and development as well as discussions around key issues such as ethics, acceptability, access and community involvement. This report discusses the current status of microbicide research and development, encompassing basic and clinical science, social and behavioural science, and community mobilisation and advocacy activities. PMID:17038196

  5. Region 4 of Rhizobium etli Primary Sigma Factor (SigA) Confers Transcriptional Laxity in Escherichia coli

    PubMed Central

    Santillán, Orlando; Ramírez-Romero, Miguel A.; Lozano, Luis; Checa, Alberto; Encarnación, Sergio M.; Dávila, Guillermo

    2016-01-01

    Sigma factors are RNA polymerase subunits engaged in promoter recognition and DNA strand separation during transcription initiation in bacteria. Primary sigma factors are responsible for the expression of housekeeping genes and are essential for survival. RpoD, the primary sigma factor of Escherichia coli, a γ-proteobacteria, recognizes consensus promoter sequences highly similar to those of some α-proteobacteria species. Despite this resemblance, RpoD is unable to sustain transcription from most of the α-proteobacterial promoters tested so far. In contrast, we have found that SigA, the primary sigma factor of Rhizobium etli, an α-proteobacteria, is able to transcribe E. coli promoters, although it exhibits only 48% identity (98% coverage) to RpoD. We have called this the transcriptional laxity phenomenon. Here, we show that SigA partially complements the thermo-sensitive deficiency of RpoD285 from E. coli strain UQ285 and that the SigA region σ4 is responsible for this phenotype. Sixteen out of 74 residues (21.6%) within region σ4 are variable between RpoD and SigA. Mutating these residues significantly improves SigA ability to complement E. coli UQ285. Only six of these residues fall into positions already known to interact with promoter DNA and to comprise a helix-turn-helix motif. The remaining variable positions are located on previously unexplored sites inside region σ4, specifically into the first two α-helices of the region. Neither of the variable positions confined to these helices seem to interact directly with promoter sequence; instead, we adduce that these residues participate allosterically by contributing to correct region folding and/or positioning of the HTH motif. We propose that transcriptional laxity is a mechanism for ensuring transcription in spite of naturally occurring mutations from endogenous promoters and/or horizontally transferred DNA sequences, allowing survival and fast environmental adaptation of α-proteobacteria. PMID

  6. Region 4 of Rhizobium etli Primary Sigma Factor (SigA) Confers Transcriptional Laxity in Escherichia coli.

    PubMed

    Santillán, Orlando; Ramírez-Romero, Miguel A; Lozano, Luis; Checa, Alberto; Encarnación, Sergio M; Dávila, Guillermo

    2016-01-01

    Sigma factors are RNA polymerase subunits engaged in promoter recognition and DNA strand separation during transcription initiation in bacteria. Primary sigma factors are responsible for the expression of housekeeping genes and are essential for survival. RpoD, the primary sigma factor of Escherichia coli, a γ-proteobacteria, recognizes consensus promoter sequences highly similar to those of some α-proteobacteria species. Despite this resemblance, RpoD is unable to sustain transcription from most of the α-proteobacterial promoters tested so far. In contrast, we have found that SigA, the primary sigma factor of Rhizobium etli, an α-proteobacteria, is able to transcribe E. coli promoters, although it exhibits only 48% identity (98% coverage) to RpoD. We have called this the transcriptional laxity phenomenon. Here, we show that SigA partially complements the thermo-sensitive deficiency of RpoD285 from E. coli strain UQ285 and that the SigA region σ4 is responsible for this phenotype. Sixteen out of 74 residues (21.6%) within region σ4 are variable between RpoD and SigA. Mutating these residues significantly improves SigA ability to complement E. coli UQ285. Only six of these residues fall into positions already known to interact with promoter DNA and to comprise a helix-turn-helix motif. The remaining variable positions are located on previously unexplored sites inside region σ4, specifically into the first two α-helices of the region. Neither of the variable positions confined to these helices seem to interact directly with promoter sequence; instead, we adduce that these residues participate allosterically by contributing to correct region folding and/or positioning of the HTH motif. We propose that transcriptional laxity is a mechanism for ensuring transcription in spite of naturally occurring mutations from endogenous promoters and/or horizontally transferred DNA sequences, allowing survival and fast environmental adaptation of α-proteobacteria.

  7. Workshop Conference on Growth Factors in the Nervous System Held in Kent, United Kingdom on 24-16 March 1986,

    DTIC Science & Technology

    1986-06-10

    cells function and maintenance. In addition to give rise to type Ii astrocytes 14 days the role these factors play in neuronal postnatal, but never to...type I. Another development and function they appear to astrocyte precursor cell develops into be crucial to nerve regeneration and type I. The...cell tumors. About 10 percent of differentiation is a new idea. At this malignant astrocytomas are poorly dif- time, the function of astrocytes is un

  8. A wheat WRKY transcription factor TaWRKY10 confers tolerance to multiple abiotic stresses in transgenic tobacco.

    PubMed

    Wang, Chen; Deng, Pengyi; Chen, Liulin; Wang, Xiatian; Ma, Hui; Hu, Wei; Yao, Ningcong; Feng, Ying; Chai, Ruihong; Yang, Guangxiao; He, Guangyuan

    2013-01-01

    WRKY transcription factors are reported to be involved in defense regulation, stress response and plant growth and development. However, the precise role of WRKY transcription factors in abiotic stress tolerance is not completely understood, especially in crops. In this study, we identified and cloned 10 WRKY genes from genome of wheat (Triticum aestivum L.). TaWRKY10, a gene induced by multiple stresses, was selected for further investigation. TaWRKY10 was upregulated by treatment with polyethylene glycol, NaCl, cold and H2O2. Result of Southern blot indicates that the wheat genome contains three copies of TaWRKY10. The TaWRKY10 protein is localized in the nucleus and functions as a transcriptional activator. Overexpression of TaWRKY10 in tobacco (Nicotiana tabacum L.) resulted in enhanced drought and salt stress tolerance, mainly demonstrated by the transgenic plants exhibiting of increased germination rate, root length, survival rate, and relative water content under these stress conditions. Further investigation showed that transgenic plants also retained higher proline and soluble sugar contents, and lower reactive oxygen species and malonaldehyde contents. Moreover, overexpression of the TaWRKY10 regulated the expression of a series of stress related genes. Taken together, our results indicate that TaWRKY10 functions as a positive factor under drought and salt stresses by regulating the osmotic balance, ROS scavenging and transcription of stress related genes.

  9. A Wheat WRKY Transcription Factor TaWRKY10 Confers Tolerance to Multiple Abiotic Stresses in Transgenic Tobacco

    PubMed Central

    Chen, Liulin; Wang, Xiatian; Ma, Hui; Hu, Wei; Yao, Ningcong; Feng, Ying; Chai, Ruihong; Yang, Guangxiao; He, Guangyuan

    2013-01-01

    WRKY transcription factors are reported to be involved in defense regulation, stress response and plant growth and development. However, the precise role of WRKY transcription factors in abiotic stress tolerance is not completely understood, especially in crops. In this study, we identified and cloned 10 WRKY genes from genome of wheat (Triticum aestivum L.). TaWRKY10, a gene induced by multiple stresses, was selected for further investigation. TaWRKY10 was upregulated by treatment with polyethylene glycol, NaCl, cold and H2O2. Result of Southern blot indicates that the wheat genome contains three copies of TaWRKY10. The TaWRKY10 protein is localized in the nucleus and functions as a transcriptional activator. Overexpression of TaWRKY10 in tobacco (Nicotiana tabacum L.) resulted in enhanced drought and salt stress tolerance, mainly demonstrated by the transgenic plants exhibiting of increased germination rate, root length, survival rate, and relative water content under these stress conditions. Further investigation showed that transgenic plants also retained higher proline and soluble sugar contents, and lower reactive oxygen species and malonaldehyde contents. Moreover, overexpression of the TaWRKY10 regulated the expression of a series of stress related genes. Taken together, our results indicate that TaWRKY10 functions as a positive factor under drought and salt stresses by regulating the osmotic balance, ROS scavenging and transcription of stress related genes. PMID:23762295

  10. Novel NAC Transcription Factor TaNAC67 Confers Enhanced Multi-Abiotic Stress Tolerances in Arabidopsis

    PubMed Central

    Mao, Xinguo; Chen, Shuangshuang; Li, Ang; Zhai, Chaochao; Jing, Ruilian

    2014-01-01

    Abiotic stresses are major environmental factors that affect agricultural productivity worldwide. NAC transcription factors play pivotal roles in abiotic stress signaling in plants. As a staple crop, wheat production is severely constrained by abiotic stresses whereas only a few NAC transcription factors have been characterized functionally. To promote the application of NAC genes in wheat improvement by biotechnology, a novel NAC gene designated TaNAC67 was characterized in common wheat. To determine its role, transgenic Arabidopsis overexpressing TaNAC67-GFP controlled by the CaMV-35S promoter was generated and subjected to various abiotic stresses for morphological and physiological assays. Gene expression showed that TaNAC67 was involved in response to drought, salt, cold and ABA treatments. Localization assays revealed that TaNAC67 localized in the nucleus. Morphological analysis indicated the transgenics had enhanced tolerances to drought, salt and freezing stresses, simultaneously supported by enhanced expression of multiple abiotic stress responsive genes and improved physiological traits, including strengthened cell membrane stability, retention of higher chlorophyll contents and Na+ efflux rates, improved photosynthetic potential, and enhanced water retention capability. Overexpression of TaNAC67 resulted in pronounced enhanced tolerances to drought, salt and freezing stresses, therefore it has potential for utilization in transgenic breeding to improve abiotic stress tolerance in crops. PMID:24427285

  11. A maize stress-responsive NAC transcription factor, ZmSNAC1, confers enhanced tolerance to dehydration in transgenic Arabidopsis.

    PubMed

    Lu, Min; Ying, Sheng; Zhang, Deng-Feng; Shi, Yun-Su; Song, Yan-Chun; Wang, Tian-Yu; Li, Yu

    2012-09-01

    NAC proteins are plant-specific transcription factors that play essential roles in stress responses. However, only little information regarding stress-related NAC genes is available in maize. In this study, a maize NAC gene, ZmSNAC1, was cloned and functionally characterized. Expression analysis revealed that ZmSNAC1 was strongly induced by low temperature, high-salinity, drought stress, and abscisic acid (ABA) treatment, but downregulated by salicylic acid treatment. Subcellular localization experiments in Arabidopsis protoplast cells indicated that ZmSNAC1 was localized in the nucleus. Transactivation assays demonstrated that ZmSNAC1 functioned as a transcriptional activator. Overexpression of ZmSNAC1 in Arabidopsis led to hypersensitivity to ABA and osmotic stress at the germination stage, but enhanced tolerance to dehydration compared to wild-type seedlings. These results suggest that ZmSNAC1 functions as a stress-responsive transcription factor in positive modulation of abiotic stress tolerance, and may have applications in the engineering of drought-tolerant crops. ZmSNAC1 functioned as a stress-responsive transcription factor in response to abiotic stresses, and might be useful for crop tolerance improvement.

  12. Paraneoplastic Hypoglycaemia: A Rare Manifestation of Pelvic Gastrointestinal Stromal Tumour

    PubMed Central

    Hadi, Rahat; Mehrotra, Kiranpreet; Rastogi, Shivani; Masood, Shakeel

    2017-01-01

    Non-Islet Cell Tumour Induced Hypoglycaemia (NICTH), presenting with recurrent fasting hypoglycaemia is a very rare paraneoplastic syndrome. It usually presents with large metastatic mesenchymal tumours. NICTH secondary to Gastrointestinal Stromal Tumour (GIST) is even rarer. Diagnosis of NICTH is based on the low serum insulin level, low serum concentrations of Insulin Like Growth Factor (IGF-I) and IGF binding protein- III (IGFBP-III) in combination with elevated concentrations of pro-IGF-II. Various Immunohistochemical (IHC) markers are integral to diagnosis of GIST namely 2-deoxyglucose-6-phosphate phosphatase -1(DOG-1), Cluster Differentiation 34 (CD 34), Cluster Differentiation 117 (CD117). The management requires prompt intravenous hydration and glucose infusions followed by surgical resection. We hereby, report a rare case of a 65-year-old female with intractable fasting hypoglycaemia due to overproduction of "big" insulin-like growth factor II diagnosed to have pelvic GIST and managed by Steroids and Imatinib.

  13. Manipulation of human early T lymphopoiesis by coculture on human bone marrow stromal cells: potential utility for adoptive immunotherapy.

    PubMed

    Liu, Bing; Ohishi, Kohshi; Orito, Yuki; Nakamori, Yoshiki; Nishikawa, Hiroyoshi; Ino, Kazuko; Suzuki, Kei; Matsumoto, Takeshi; Masuya, Masahiro; Hamada, Hirofumi; Mineno, Junichi; Ono, Ryoichi; Nosaka, Tetsuya; Shiku, Hiroshi; Katayama, Naoyuki

    2013-04-01

    T cell precursors are an attractive target for adoptive immunotherapy. We examined the regulat