Journal articles on the topic 'Mice embryonic fibroblast'
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Singhal, Prabhat K., Slim Sassi, Lan Lan, Patrick Au, Stefan C. Halvorsen, Dai Fukumura, Rakesh K. Jain, and Brian Seed. "Mouse embryonic fibroblasts exhibit extensive developmental and phenotypic diversity." Proceedings of the National Academy of Sciences 113, no. 1 (December 22, 2015): 122–27. http://dx.doi.org/10.1073/pnas.1522401112.
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Analysis of embryonic fibroblasts from GFP reporter mice indicates that the fibroblast cell type harbors a large collection of developmentally and phenotypically heterogeneous subtypes. Some of these cells exhibit multipotency, whereas others do not. Multiparameter flow cytometry analysis shows that a large number of distinct populations of fibroblast-like cells can be found in cultures initiated from different embryonic organs, and cells sorted according to their surface phenotype typically retain their characteristics on continued propagation in culture. Similarly, surface phenotypes of individual cloned fibroblast-like cells exhibit significant variation. The fibroblast cell class appears to contain a very large number of denumerable subtypes.
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Niedermeyer, J., M. Kriz, F. Hilberg, P. Garin-Chesa, U. Bamberger, M. C. Lenter, J. Park, et al. "Targeted Disruption of Mouse Fibroblast Activation Protein." Molecular and Cellular Biology 20, no. 3 (February 1, 2000): 1089–94. http://dx.doi.org/10.1128/mcb.20.3.1089-1094.2000.
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ABSTRACT Human fibroblast activation protein (FAP), a member of the serine prolyl oligopeptidase family, is a type II cell surface glycoprotein selectively expressed by fibroblastic cells in areas of active tissue remodeling, such as the embryonic mesenchyme, areas of wound healing, the gravid uterus, and the reactive stroma of epithelial cancers. Homologues of FAP have been identified in the mouse and Xenopus laevis. FAP is a dual-specificity enzyme that acts as a dipeptidyl peptidase and collagenase in vitro. To explore the role of FAP in vivo, Fap −/− mice were generated by homologous recombination. RNase protection analysis and reverse transcription-PCR confirmed the absence of full-length Faptranscripts in mouse embryonic tissues. No FAP protein was detected inFap −/− animals by immunohistochemistry, and no FAP-specific dipeptidyl peptidase activity was found. We report thatFap −/− mice are fertile, show no overt developmental defects, and have no general change in cancer susceptibility.
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Patrinostro, Xiaobai, Allison R. O'Rourke, Christopher M. Chamberlain, Branden S. Moriarity, Benjamin J. Perrin, and James M. Ervasti. "Relative importance of βcyto- and γcyto-actin in primary mouse embryonic fibroblasts." Molecular Biology of the Cell 28, no. 6 (March 15, 2017): 771–82. http://dx.doi.org/10.1091/mbc.e16-07-0503.
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The highly homologous β (βcyto) and γ (γcyto) cytoplasmic actins are hypothesized to carry out both redundant and unique essential functions, but studies using targeted gene knockout and siRNA-mediated transcript knockdown to examine βcyto- and γcyto-isoform–specific functions in various cell types have yielded conflicting data. Here we quantitatively characterized actin transcript and protein levels, as well as cellular phenotypes, in both gene- and transcript-targeted primary mouse embryonic fibroblasts. We found that the smooth muscle αsm-actin isoform was the dominantly expressed actin isoform in WT primary fibroblasts and was also the most dramatically up-regulated in primary βcyto- or β/γcyto-actin double-knockout fibroblasts. Gene targeting of βcyto-actin, but not γcyto-actin, led to greatly decreased cell proliferation, decreased levels of cellular ATP, and increased serum response factor signaling in primary fibroblasts, whereas immortalization induced by SV40 large T antigen supported fibroblast proliferation in the absence of βcyto-actin. Consistent with in vivo gene-targeting studies in mice, both gene- and transcript-targeting approaches demonstrate that the loss of βcyto-actin protein is more disruptive to primary fibroblast function than is the loss of γcyto-actin.
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Jarboe, D. L., and T. F. Huff. "The mast cell-committed progenitor. II. W/Wv mice do not make mast cell-committed progenitors and S1/S1d fibroblasts do not support development of normal mast cell-committed progenitors." Journal of Immunology 142, no. 7 (April 1, 1989): 2418–23. http://dx.doi.org/10.4049/jimmunol.142.7.2418.
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Abstract We have previously reported that the population of mesenteric lymph node cells from normal BALB/c mice infected 14 days with the rodent nematode Nippostrongylus brasiliensis (Nb-MLN) contains a nongranulated mast cell-committed progenitor (MCCP) which does not require IL-3 for proliferation and differentiation if either a fibroblast monolayer or soluble factors produced by monolayers of 3T3 fibroblasts or embryonic skin are present in the culture. When Nb-MLN were cloned in a methylcellulose culture system using fibroblast conditioned medium as the only source of growth factors, numerous colonies of pure mast cells developed. We wished to determine whether the mast cell deficiency of W/Wv or S1/S1d mice could be explained by the failure of these mice to make either the MCCP or the factor to support proliferation and differentiation of the MCCP. We found that Nb-MLN from W/Wv mice were only able to produce mast cell colonies in response to a source of IL-3 such as conditioned medium from pokeweed mitogen-stimulated spleen cells (CM), and cultures given fibroblast conditioned medium as the only source of growth factors did not produce mast cell colonies. In contrast, Nb-MLN from mast cell deficient S1/S1d mice developed many mast cell colonies in methylcellulose cultures supplemented with either fibroblast conditioned medium or conditioned medium from PWM-stimulated spleen cells. These data suggest that S1/S1d mice but not W/Wv mice produce the mast cell progenitor that responds to fibroblast conditioned medium. To determine if mast cell deficient mice make the fibroblast derived factors that support development of the MCCP, monolayers were prepared from skin connective tissues of S1/S1d and W/Wv mice and Nb-MLN from normal BALB/c mice were cloned in the presence of conditioned medium from these monolayers. Fibroblast conditioned medium from monolayers prepared from W/Wv but not S1/S1d mice supported development of numerous mast cell colonies. Taken together, these data demonstrate that W/Wv mice are incapable of producing normal MCCP whereas S1/S1d fibroblasts fail to produce the appropriate factor to support the MCCP. In accordance with these data, a candidate for the gene product of each of these mutant alleles is discussed.
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DeClerck, Y., V. Draper, and R. Parkman. "Clonal analysis of murine graft-vs-host disease. II. Leukokines that stimulate fibroblast proliferation and collagen synthesis in graft-vs. host disease." Journal of Immunology 136, no. 10 (May 15, 1986): 3549–52. http://dx.doi.org/10.4049/jimmunol.136.10.3549.
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Abstract T lymphocyte clones were established from mice with acute and chronic graft-vs-host disease (GVHD) [C57B6/6 (B6) mice transplanted i.v. with LP/J spleen cells] and immune mice (LP/J mice immunized intraperitoneally with B6 spleen cells). To determine the role of leukokines in the increased collagen production that characterizes chronic GVHD, the supernatants of in vitro stimulated clones were assayed for their effect on 1) B6 embryonic fibroblast proliferation, 2) total fibroblast collagen secretion, and 3) collagen secretion per fibroblast. Four of nine chronic GVHD (CG) clones stimulated both total collagen secretion and collagen secretion per fibroblast, but none stimulated fibroblast proliferation. Six of 10 immune (I) clones stimulated fibroblast proliferation, and none stimulated collagen secretion per fibroblast. Acute GVHD (G) clones were heterogeneous; 2/10 G clones stimulated fibroblast proliferation, 5/10 stimulated total collagen secretion, and 4/10 stimulated collagen secretion per fibroblast. I and CG clones may act synergistically in vivo. The presence of CG-like clones in mice with acute GVHD suggest that the immunologic events that culminate in chronic GVHD are initiated early after transplantation.
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Kranc, Kamil R., Simon D. Bamforth, José Bragança, Chris Norbury, Maarten van Lohuizen, and Shoumo Bhattacharya. "Transcriptional Coactivator Cited2 Induces Bmi1 and Mel18 and Controls Fibroblast Proliferation via Ink4a/ARF." Molecular and Cellular Biology 23, no. 21 (November 1, 2003): 7658–66. http://dx.doi.org/10.1128/mcb.23.21.7658-7666.2003.
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ABSTRACT Cited2 (CBP/p300 interacting transactivator with ED-rich tail 2) is required for embryonic development, coactivation of transcription factor AP-2, and inhibition of hypoxia-inducible factor 1 transactivation. Cited2 is induced by multiple growth factors and cytokines and oncogenically transforms cells. Here, we show that the proliferation of Cited2 −/− mouse embryonic fibroblasts ceases prematurely. This is associated with a reduction in growth fraction, senescent cellular morphology, and increased expression of the cell proliferation inhibitors p16INK4a, p19ARF, and p15INK4b. Deletion of INK4a/ARF (encoding p16INK4a and p19ARF) completely rescued the defective proliferation of Cited2−/− fibroblasts. However, the deletion of INK4a/ARF did not rescue the embryonic malformations observed in Cited2 −/− mice, indicating that INK4a/ARF-independent pathways are likely to be involved here. We found that Cited2 −/− fibroblasts had reduced expression of the polycomb-group genes Bmi1 and Mel18, which function as INK4a/ARF and Hox repressors. Complementation with CITED2-expressing retrovirus enhanced proliferation, induced Bmi1/Mel18 expression, and decreased INK4a/ARF expression. Bmi1- and Mel18-expressing retroviruses enhanced the proliferation of Cited2 −/− fibroblasts, indicating that they function downstream of Cited2. Our results provide genetic evidence that Cited2 controls the expression of INK4a/ARF and fibroblast proliferation, at least in part via the polycomb-group genes Bmi1 and Mel18.
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Hollands, P. "Differentiation of embryonic haemopoietic stem cells from mouse blastocysts grown in vitro." Development 102, no. 1 (January 1, 1988): 135–41. http://dx.doi.org/10.1242/dev.102.1.135.
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Embryonic haemopoietic stem cells can differentiate from mouse blastocysts grown in vitro. Mouse blastocysts were cultured for 3 or 4 days and the resultant cells were injected intravenously into lethally X-irradiated or genetically anaemic recipient mice. Blastocysts grown in vitro did not maintain normal embryonic morphology. The presence of donor haemoglobin and donor lymphocytic glucose phosphate isomerase in grafted recipients, demonstrates the presence of embryonic haemopoietic stem cells. Recipients of embryonic haemopoietic stem cells, obtained from growth in vitro, were haematologically stable with no evidence of neoplasia. Pluripotent embryonic cells, maintained on fibroblast feeder layers, were unable to colonize X-irradiated or genetically anaemic mice. Recipients of pluripotent cells died at the same time as saline-injected controls.
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MACKINTOSH, Caroline A., and Anthony E. PEGG. "Effect of spermine synthase deficiency on polyamine biosynthesis and content in mice and embryonic fibroblasts, and the sensitivity of fibroblasts to 1,3-bis-(2-chloroethyl)-N-nitrosourea." Biochemical Journal 351, no. 2 (October 10, 2000): 439–47. http://dx.doi.org/10.1042/bj3510439.
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Mutant Gy male mice, which have previously been described as having disruption of the phosphate-regulating Phex gene and a spermine synthase gene [Meyer, Henley, Meyer, Morgan, McDonald, Mills and Price (1998) Genomics, 48, 289–295; Lorenz, Francis, Gempel, Böddrich, Josten, Schmahl and Schmidt (1998) Hum. Mol. Genet. 7, 541–547], as well as mutant Hyp male mice, which have disruption of the Phex gene only, were examined along with their respective normal male littermates. Biochemical analyses of extracts of brains, hearts and livers of 5-week-old mice showed that Gy males lacked any significant spermine synthase activity as well as spermine content. Organs of Gy males had a higher spermidine content. This was caused not only by the lack of conversion of spermidine into spermine, but also because of compensatory increases in the activities of other polyamine biosynthetic enzymes. Gy males were half the body weight of their normal male littermates at weaning age. Hyp males, however, were no different in size when compared with their controls. High mortality of Gy males occurs by weaning age and this mortality was shown to be largely post-natal. Embryonic fibroblasts were isolated from Gy males and their normal male littermates and were similarly shown to lack any significant spermine synthase activity as well as spermine content. The lack of spermine, however, had no significant effect on the growth of immortalized fibroblasts or of primary fibroblast cultures. Similarly, there was no difference in the time of senescence of primary fibroblast cultures from Gy males compared with cultures derived from normal male littermates. However, the lack of spermine did increase the sensitivity of immortalized fibroblasts to killing by the chloroethylating agent 1,3-bis(2-chloroethyl)-N-nitrosourea. Therefore both the Gy male mice and derived embryonic fibroblasts provide valuable models to study the importance of spermine and spermine synthase, without the use of inhibitors which may have additional side effects.
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Evans, S. M., L. J. Tai, V. P. Tan, C. B. Newton, and K. R. Chien. "Heterokaryons of cardiac myocytes and fibroblasts reveal the lack of dominance of the cardiac muscle phenotype." Molecular and Cellular Biology 14, no. 6 (June 1994): 4269–79. http://dx.doi.org/10.1128/mcb.14.6.4269-4279.1994.
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The molecular characterization of a cardiac determination gene has been an elusive goal for the past several years. Prior to cloning of the skeletal muscle determination factor MyoD, the presence of a dominantly acting skeletal muscle determination factor had been inferred from the observation that the skeletal muscle phenotype was dominant in skeletal muscle-fibroblast heterokaryons (H. M. Blau, G. K. Pavlath, E. C. Hardeman, C.-P. Chiu, L. Siberstein, S. G. Webster, S. C. Miller, and D. Webster, Science 230:758-766, 1985). In these experiments, we have examined cardiac-fibroblast heterokaryons to investigate the existence of a dominantly acting cardiac determination factor. We have employed a novel experimental approach using primary embryonic fibroblasts from transgenic mice as a means of assaying for the activation of a cardiac promoter-luciferase reporter transgene within fibroblast nuclei. This approach provides a potential means of genetic selection for a dominantly acting positive factor and can be generalized to other systems. We have examined the expression of three markers of the cardiac lineage: a myofibrillar protein promoter (MLC2), a secreted protein (ANF), and a transcription factor (MEF2). MEF2 is specific to both cardiac and skeletal muscle cells. Our results indicate that in a majority of heterokaryons with an equal ratio of cardiac to fibroblast nuclei, none of these cardiac markers are expressed, indicating that the cardiac phenotype is not dominant over the embryonic fibroblast phenotype. The distinction from previous results with skeletal muscle is emphasized by our results with MEF2, which is dominantly expressed in skeletal muscle-fibroblast but not cardiac-fibroblast heterokaryons, supporting its divergent regulation in the two cell types.
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Evans, S. M., L. J. Tai, V. P. Tan, C. B. Newton, and K. R. Chien. "Heterokaryons of cardiac myocytes and fibroblasts reveal the lack of dominance of the cardiac muscle phenotype." Molecular and Cellular Biology 14, no. 6 (June 1994): 4269–79. http://dx.doi.org/10.1128/mcb.14.6.4269.
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The molecular characterization of a cardiac determination gene has been an elusive goal for the past several years. Prior to cloning of the skeletal muscle determination factor MyoD, the presence of a dominantly acting skeletal muscle determination factor had been inferred from the observation that the skeletal muscle phenotype was dominant in skeletal muscle-fibroblast heterokaryons (H. M. Blau, G. K. Pavlath, E. C. Hardeman, C.-P. Chiu, L. Siberstein, S. G. Webster, S. C. Miller, and D. Webster, Science 230:758-766, 1985). In these experiments, we have examined cardiac-fibroblast heterokaryons to investigate the existence of a dominantly acting cardiac determination factor. We have employed a novel experimental approach using primary embryonic fibroblasts from transgenic mice as a means of assaying for the activation of a cardiac promoter-luciferase reporter transgene within fibroblast nuclei. This approach provides a potential means of genetic selection for a dominantly acting positive factor and can be generalized to other systems. We have examined the expression of three markers of the cardiac lineage: a myofibrillar protein promoter (MLC2), a secreted protein (ANF), and a transcription factor (MEF2). MEF2 is specific to both cardiac and skeletal muscle cells. Our results indicate that in a majority of heterokaryons with an equal ratio of cardiac to fibroblast nuclei, none of these cardiac markers are expressed, indicating that the cardiac phenotype is not dominant over the embryonic fibroblast phenotype. The distinction from previous results with skeletal muscle is emphasized by our results with MEF2, which is dominantly expressed in skeletal muscle-fibroblast but not cardiac-fibroblast heterokaryons, supporting its divergent regulation in the two cell types.
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García-Sastre, Adolfo, Russell K. Durbin, Hongyong Zheng, Peter Palese, Rachel Gertner, David E. Levy, and Joan E. Durbin. "The Role of Interferon in Influenza Virus Tissue Tropism." Journal of Virology 72, no. 11 (November 1, 1998): 8550–58. http://dx.doi.org/10.1128/jvi.72.11.8550-8558.1998.
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ABSTRACT We have studied the pathogenesis of influenza virus infection in mice that are unable to respond to type I or II interferons due to a targeted disruption of the STAT1 gene. STAT1−/− animals are 100-fold more sensitive to lethal infection with influenza A/WSN/33 virus than are their wild-type (WT) counterparts. Virus replicated only in the lungs of WT animals following intranasal (i.n.) virus inoculation, while STAT1−/− mice developed a fulminant systemic influenza virus infection following either i.n. or intraperitoneal inoculation. We investigated the mechanism underlying this altered virus tropism by comparing levels of virus replication in fibroblast cell lines and murine embryonic fibroblasts derived from WT mice, STAT−/− mice, and mice lacking gamma interferon (IFNγ−/− mice) or the IFN-α receptor (IFNαR−/− mice). Influenza A/WSN/33 virus replicates to high titers in STAT1−/− or IFNαR−/− fibroblasts, while cells derived from WT or IFNγ−/− animals are resistant to influenza virus infection. Immunofluorescence studies using WT fibroblast cell lines demonstrated that only a small subpopulation of WT cells can be infected and that in the few infected WT cells, virus replication is aborted at an early, nuclear phase. In all organs examined except the lung, influenza A WSN/33 virus infection is apparently prevented by an intact type I interferon response. Our results demonstrate that type I interferon plays an important role in determining the pathogenicity and tissue restriction of influenza A/WSN/33 virus in vivo and in vitro.
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Tucker, Rebecca M., and David T. Burke. "Transgenic mice for the establishment of histidinol‐resistant embryonic fibroblast feeder layers." FASEB Journal 10, no. 14 (December 1996): 1641–45. http://dx.doi.org/10.1096/fasebj.10.14.9002557.
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Popova, Svetlana N., Malgorzata Barczyk, Carl-Fredrik Tiger, Wouter Beertsen, Paola Zigrino, Attila Aszodi, Nicolai Miosge, Erik Forsberg, and Donald Gullberg. "α11β1 Integrin-Dependent Regulation of Periodontal Ligament Function in the Erupting Mouse Incisor." Molecular and Cellular Biology 27, no. 12 (June 15, 2007): 4306–16. http://dx.doi.org/10.1128/mcb.00041-07.
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ABSTRACT The fibroblast integrin α11β1 is a key receptor for fibrillar collagens. To study the potential function of α11 in vivo, we generated a null allele of the α11 gene. Integrin α11−/− mice are viable and fertile but display dwarfism with increased mortality, most probably due to severely defective incisors. Mutant incisors are characterized by disorganized periodontal ligaments, whereas molar ligaments appear normal. The primary defect in the incisor ligament leads to halted tooth eruption. α11β1-defective embryonic fibroblasts displayed severe defects in vitro, characterized by (i) greatly reduced cell adhesion and spreading on collagen I, (ii) reduced ability to retract collagen lattices, and (iii) reduced cell proliferation. Analysis of matrix metalloproteinase in vitro and in vivo revealed disturbed MMP13 and MMP14 synthesis in α11−/− cells. We show that α11β1 is the major receptor for collagen I on mouse embryonic fibroblasts and suggest that α11β1 integrin is specifically required on periodontal ligament fibroblasts for cell migration and collagen reorganization to help generate the forces needed for axial tooth movement. Our data show a unique role for α11β1 integrin during tooth eruption.
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Jarboe, D. L., J. S. Marshall, T. R. Randolph, A. Kukolja, and T. F. Huff. "The mast cell-committed progenitor. I. Description of a cell capable of IL-3-independent proliferation and differentiation without contact with fibroblasts." Journal of Immunology 142, no. 7 (April 1, 1989): 2405–17. http://dx.doi.org/10.4049/jimmunol.142.7.2405.
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Abstract We have identified a late, committed stage in the differentiation of the mast cell progenitor just before granulation. Mast cell committed progenitors (MCCP) are nongranulated cells with a density of 1.060 to 1.070 g/ml which can be harvested from the mesenteric lymph node of mice infected with Nippostrongylus brasiliensis. Mast cell-committed progenitors are able to proliferate and differentiate in the absence of IL-3 or IL-4 when cultured on a monolayer of embryonic skin or 3T3 fibroblasts and can form colonies in methylcellulose supplemented with fibroblast conditioned medium. Fibroblast conditioned medium appears to contain a soluble MCCP proliferation factor that maintains biologic activity when heated to 56 degrees C for 45 min but is destroyed by incubation with either trypsin or chymotrypsin. It can be selectively precipitated with 60 to 70% saturated ammonium sulfate. The factor is not absorbed by immobilized antibodies to nerve growth factor. The MCCP proliferation activity of the factor could not be mimicked by IL-1, IL-2, IL-4, granulocyte-macrophage-CSF, granulocyte-CSF, macrophage-CSF, IFN-alpha/beta, IFN-gamma, nerve growth factor, epidermal growth factor, serum fibronectin, heparin, or a number of glycosaminoglycans. At high salt concentrations, the factor passes through a 50-kDa membrane and can be concentrated above a 5-kDa membrane. MCCP acquire a connective tissue phenotype when cultured on a fibroblast monolayer and a mucosal phenotype when cloned in the presence of conditioned medium from PWM-stimulated spleen cells. When cultured in the absence of IL-3 on a monolayer of embryonic skin or 3T3 fibroblasts, mast cell-committed progenitors produce mast cells which stain with berberine sulfate suggesting a connective tissue phenotype; however, the mast cells that develop when mast cell-committed progenitors are cultured in the presence of IL-3 or conditioned media from PWM-stimulated spleen cells do not stain with berberine sulfate. MCCP intercalate into monolayers of embryonic skin or 3T3 fibroblasts, but T cells are not able to associate with the monolayer and can be completely washed away. Attempts to enrich mast cell-committed progenitors by intercalation and elution from embryonic skin monolayers proved unsuccessful, but some enrichment of mast cell-committed progenitors could be achieved by discontinuous Percoll gradients. Thus, we have identified a way to obtain late-stage, mast cell-committed progenitors in an environment that is virtually uncontaminated with other hematopoietic progenitors.
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Fortschegger, Klaus, Bettina Wagner, Regina Voglauer, Hermann Katinger, Maria Sibilia, and Johannes Grillari. "Early Embryonic Lethality of Mice Lacking the Essential Protein SNEV." Molecular and Cellular Biology 27, no. 8 (February 5, 2007): 3123–30. http://dx.doi.org/10.1128/mcb.01188-06.
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ABSTRACT SNEV (Prp19, Pso4, NMP200) is a nuclear matrix protein known to be involved in pre-mRNA splicing, ubiquitylation, and DNA repair. In human umbilical vein endothelial cells, SNEV overexpression delayed the onset of replicative senescence. Here we analyzed the function of the mouse SNEV gene in vivo by employing homologous recombination in mice and conclude that SNEV is indispensable for early mouse development. Mutant preimplantation embryos initiated blastocyst formation but died shortly thereafter. Outgrowth of SNEV-null blastocysts showed a lack of proliferation of cells of the inner cell mass, which subsequently underwent cell death. While SNEV-heterozygous mice showed no overt phenotype, heterozygous mouse embryonic fibroblast cell lines with reduced SNEV levels displayed a decreased proliferative potential in vitro. Our experiments demonstrate that the SNEV protein is essential, functionally nonredundant, and indispensable for mouse development.
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Johnson, Kevin A., Charles P. Lerner, Lena C. Di Lacio, Peter W. Laird, Arlene H. Sharpe, and Elizabeth M. Simpson. "Transgenic mice for the preparation of hygromycin-resistant primary embryonic fibroblast feeder layers for embryonic stem cell selections." Nucleic Acids Research 23, no. 7 (1995): 1273–75. http://dx.doi.org/10.1093/nar/23.7.1273.
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Linnell, Elizabeth R., Charles P. Lerner, Kevin A. Johnson, Craig A. Leach, Thomas R. Ulrich, William C. Rafferty, and Elizabeth M. Simpson. "Transgenic mice for the preparation of puromycin-resistant primary embryonic fibroblast feeder layers for embryonic stem cell selection." Mammalian Genome 12, no. 2 (February 1, 2001): 169–71. http://dx.doi.org/10.1007/s003350010238.
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Shen, Junhui, Zhong-Kai Cui, Fang Yao, Kai Li, Yue Zhang, Zhenguo Chen, Yuxia Zhou, et al. "TSC1 deletion in fibroblasts alleviates lipopolysaccharide-induced acute kidney injury." Clinical Science 132, no. 19 (October 2, 2018): 2087–101. http://dx.doi.org/10.1042/cs20180348.
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Mechanistic target of rapamycin complex 1 (mTORC1) signaling is active in inflammation, but its involvement in septic acute kidney injury (AKI) has not been shown. mTORC1 activation (p-S6) in renal fibroblasts was increased in a mouse AKI model induced by 1.5 mg/kg lipopolysaccharide (LPS). Deletion of tuberous sclerosis complex 1 (TSC1), an mTORC1 negative regulator, in fibroblasts (Fibro-TSC1−/−) inhibited the elevation of serum creatinine and blood urea nitrogen in AKI compared with that in TSC1fl/fl control mice. Endothelin-1 (EDN1) and phospho-Jun-amino-terminal kinase (p-JNK) were up-regulated in Fibro-TSC1−/− renal fibroblasts after LPS challenge. Rapamycin, an mTORC1 inhibitor, and bosentan, an EDN1 antagonist, eliminated the difference in renal function between TSC1fl/fl and Fibro-TSC1−/− mice after LPS injection. Rapamycin restored LPS-induced up-regulation of EDN1, endothelin converting enzyme-1 (ECE1), and p-JNK in TSC1-knockdown mouse embryonic fibroblasts (MEFs). SP600125, a Jun-amino-terminal kinase (JNK) inhibitor, attenuated LPS-induced enhancement of EDN1 and ECE1 in TSC1-knockdown MEFs without a change in phospho-S6 ribosomal protein (p-S6) level. The results indicate that mTORC1–JNK-dependent up-regulation of ECE1 elevated EDN1 in TSC1-knockout renal fibroblasts and contributed to improvement of renal function in Fibro-TSC1−/− mice with LPS-induced AKI. Renal fibroblast mTORC1 plays an important role in septic AKI.
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Kanakaraj, Palanisamy, Peter H. Schafer, Druie E. Cavender, Ying Wu, Karen Ngo, Patrick F. Grealish, Scott A. Wadsworth, et al. "Interleukin (IL)-1 Receptor–associated Kinase (IRAK) Requirement for Optimal Induction of Multiple IL-1 Signaling Pathways and IL-6 Production." Journal of Experimental Medicine 187, no. 12 (June 15, 1998): 2073–79. http://dx.doi.org/10.1084/jem.187.12.2073.
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Interleukin (IL)-1 is a proinflammatory cytokine with pleiotropic effects in inflammation. IL-1 binding to its receptor triggers a cascade of signaling events, including activation of the stress-activated mitogen-activated protein (MAP) kinases, c-Jun NH2-terminal kinase (JNK) and p38 MAP kinase, as well as transcription factor nuclear factor κB (NF-κB). IL-1 signaling results in cellular responses through induction of inflammatory gene products such as IL-6. One of the earliest events in IL-1 signaling is the rapid interaction of IL-1 receptor–associated kinases, IRAK and IRAK-2, with the receptor complex. The relative roles of IRAK and IRAK-2 in IL-1 signaling pathways and subsequent cellular responses have not been previously determined. To evaluate the importance of IRAK in IL-1 signaling, IRAK-deficient mouse fibroblast cells were prepared and studied. Here we report that IL-1–mediated activation of JNK, p38, and NF-κB were all reduced in embryonic fibroblasts deficient in IRAK expression. In addition, IL-6 production in response to IL-1 was also dramatically reduced in IRAK-deficient embryonic fibroblasts and in skin fibroblasts prepared from IRAK-deficient mice. Our results demonstrate that IRAK plays an essential proximal role in coordinating multiple IL-1 signaling pathways for optimal induction of cellular responses.
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Sakai, Shigeki, Noriko Aramaki-Hattori, and Kazuo Kishi. "Fetal Fibroblast Transplantation via Ablative Fractional Laser Irradiation Reduces Scarring." Biomedicines 11, no. 2 (January 26, 2023): 347. http://dx.doi.org/10.3390/biomedicines11020347.
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Scar treatments include fractional laser treatment, cell transplantation, surgery, skin needling, and dermal fillers. Fractional laser treatments are used to reduce scarring and blurring. Cell transplantation is promising, with mature fibroblasts and adipose-derived stem cells being used clinically, while embryonic fibroblasts are used experimentally. Herein, we developed a combination of ablative CO2 (carbon dioxide) fractional laser and cell transplantation for the treatment of scars. Eight-week-old male C57Bl/6 mice were used to create a full-layer skin defect in the back skin and create scars. The scar was then irradiated using a CO2 fractional laser. The cells were then transplanted onto the scar surface and sealed with a film agent. The transplanted cells were GFP-positive murine fetal fibroblasts (FB), fetal fibroblasts with a long-term sphere-forming culture (LS), and fetal skin with a short-term sphere-forming culture (SS). After transplantation, green fluorescent protein (GFP)-positive cells were scattered in the dermal papillary layer and subcutis in all the groups. LS significantly reduced the degree of scarring, which was closest to normal skin. In conclusion, the combination of ablative fractional laser irradiation and fetal fibroblast transplantation allowed us to develop new methods for scar treatment.
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Liu, Feng, Peng yu-huan, Li Qiang, and Liu Chanchan. "Expression of Imprinted Genes Kcnq1 and Cdkn1c During the Course of Differentiation from Mouse Embryonic Stem Cells into Islet-like Cells in vitro." Experimental and Clinical Endocrinology & Diabetes 126, no. 04 (September 19, 2017): 249–54. http://dx.doi.org/10.1055/s-0043-113254.
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AbstractTo study the effects of inducement on the expression of mouse embryonic stem cells SF1-G imprinted genes, Kcnq1 and Cdkn1c during the course of differentiation into islet-like cells in vitro. Mouse embryonic fibroblasts (MEFs) were isolated from pregnant mice embryos and fibroblast feeder cells were prepared by treating 3–5th generations MEFs with Mitomycin C. Moreover, mouse embryonic stem cells were induced to differentiate into islet-like cells directly. RT-PCR and Immunofluorescence staining were used to test the expression of islet cell-specific markers. Cells were collected at various stages throughout the differentiation process and the imprinted genes Kcnq1 and Cdkn1c were tested by reverse transcription-polymerase chain reaction fragment length polymorphism (RT-PCR/RFLP). In the present study, we found that cells appear islet cell-specific gene expression. Furthermore, immunofluorescence shows us that the islet cell-specific hormone protein can be measured at stage, which confirms that the embryonic stem cells can be successfully induced into islet-like cells in vitro. RT-PCR/RFLP analysis showsthat imprinted genes Kcnq1 and Cdkn1c are biallelic expression in the differentiated cells, suggestive of loss of imprinting (LOI), while these genes demonstrate maternal monoallelic expression in the undifferentiated cells’ continued subculture; this marks the maintenance of imprinting (MOI). Our data indicate that mouse embryonic stem cells are induced into islet-like cells in vitro. The gene imprinting status of Kcnq1 and Cdkn1c may be changed in differentiated cells during the induction in vitro.
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Labosky, P. A., D. P. Barlow, and B. L. Hogan. "Mouse embryonic germ (EG) cell lines: transmission through the germline and differences in the methylation imprint of insulin-like growth factor 2 receptor (Igf2r) gene compared with embryonic stem (ES) cell lines." Development 120, no. 11 (November 1, 1994): 3197–204. http://dx.doi.org/10.1242/dev.120.11.3197.
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Primordial germ cells of the mouse cultured on feeder layers with leukemia inhibitory factor, Steel factor and basic fibroblast growth factor give rise to cells that resemble undifferentiated blastocyst-derived embryonic stem cells. These primordial germ cell-derived embryonic germ cells can be induced to differentiate extensively in culture, form teratocarcinomas when injected into nude mice and contribute to chimeras when injected into host blastocysts. Here, we report the derivation of multiple embryonic germ cell lines from 8.5 days post coitum embryos of C57BL/6 inbred mice. Four independent embryonic germ cell lines with normal male karyotypes have formed chimeras when injected into BALB/c host blastocysts and two of these lines have transmitted coat color markers through the germline. We also show that pluripotent cell lines capable of forming teratocarcinomas and coat color chimeras can be established from primordial germ cells of 8.0 days p.c. embryos and 12.5 days p.c. genital ridges. We have examined the methylation status of the putative imprinting box of the insulin-like growth factor type 2 receptor gene (Igf2r) in these embryonic germ cell lines. No correlation was found between methylation pattern and germline competence. A significant difference was observed between embryonic stem cell and embryonic germ cell lines in their ability to maintain the methylation imprint of the Igf2r gene in culture. This may illustrate a fundamental difference between these two cell types.
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Trotter, Andreas, Markus Kipp, Roland Matthias Schrader, and Cordian Beyer. "Combined Application of 17-Estradiol and Progesterone Enhance Vascular Endothelial Growth Factor and Surfactant Protein Expression in Cultured Embryonic Lung Cells of Mice." International Journal of Pediatrics 2009 (2009): 1–8. http://dx.doi.org/10.1155/2009/170491.
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Preterm delivery is associated with disruption of the placental supply with 17-estradiol (E2) and progesterone (P). The aim is to evaluate the role of E2 and P on the regulation of key proteins in lung development in embryonic lung cells. Alveolar cell type II (AT-II) and central lung fibroblast cultures were established from mouse embryos. Cells were exposed for 24 hours to E2 and/or P, the estrogen receptor antagonist ICI 182.780 (ICI) and the progesterone receptor antagonist mifepristone (RU 486). The mRNA expression of vascular endothelial growth factor (VEGF) and surfactant protein B and C (SB-B, SB-C) was determined, and protein levels of VEGF were measured. Only the combined treatment with E2 and P increased mRNA expression and VEGF protein in AT-II cells and lung fibroblasts. Combined treatment also promoted SP-B and SP-C expression in AT-II cells. Pretreatment with ICI and RU 486 completely abolished the E2 and P induced effects. E2 and P enhanced expression of VEGF and surfactant proteins in primary embryonic lung cells and may be involved in regulating expression of key molecules for the prenatal lung development and postnatal lung function.
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Györfi, A. H., A. E. Matei, M. Fuchs, A. Rius Rigau, X. Hong, Z. Honglin, M. Luber, et al. "POS0328 ENGRAILED 1 COORDINATES CYTOSKELETAL ORGANIZATION TO PROMOTE MYOFIBROBLAST DIFFERENTIATION AND FIBROTIC TISSUE REMODELING." Annals of the Rheumatic Diseases 80, Suppl 1 (May 19, 2021): 391.2–391. http://dx.doi.org/10.1136/annrheumdis-2021-eular.1428.
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Background:Engrailed 1 (EN1) is a homeodomain-containing transcription factor with essential roles in embryonic development. In most cell types, the expression of EN1 is restricted to embryonic development. However, under pathological conditions, EN1 can be re-expressed to promote phenotypical adaptation. En1 is transiently expressed in the developing dermis of murine embryos in a distinct fibroblast lineage and silenced before birth (1). Former EN1-expressing cells give rise to a subpopulation of fibroblasts that has a high capacity for extracellular matrix production in adult murine skin. The role of EN1 in systemic sclerosis (SSc) was previously not explored.Objectives:To study the role of EN1 in the pathological activation of fibroblasts in tissue fibrosis.Methods:Bulk RNA-Seq and EN1 or SP1 ChIP-Seq were performed from cultured human dermal fibroblasts. The expression of EN1 was inhibited by siRNA. Cytoskeletal drugs paclitaxel, vinblastin and ROCK inhibitor (Y27632) were used to modulate the cytoskeleton in EN1 knockdown or overexpressing dermal fibroblasts. The role of EN1 in fibroblast activation was evaluated by functional experiments with EN1 knockdown or overexpression in standard 2D culture systems as well as in 3D skin equivalent models. The role of EN1 in skin fibrosis was further studied in En1fl/fl X Col6Cre mice, with fibroblast-specific knockout of En1 in three complementary mouse models: overexpression of a constitutively active TGFß-receptor I (TBRICA), bleomycin-induced skin fibrosis and TSK1 mice.Results:Pathologically activated dermal fibroblasts from SSc patients express higher levels of EN1 compared with age and sex matched healthy individuals in the skin and in vitro. TGFβ induces EN1 expression in fibroblasts in a SMAD3-dependent manner both in cultured fibroblasts and in murine skin. Knockdown of EN1 prevents TGFβ-induced fibroblast activation, whereas overexpression of EN1 fosters the pro-fibrotic effects of TGFβ with increased expression of αSMA, stress fibers and collagen. RNA sequencing demonstrates that EN1 induces a pro-fibrotic gene expression profile functionally related to cytoskeleton organization and ROCK activation. In silico analyses of the promoters of En1 target genes coupled with siRNA-mediated knockdown demonstrated that EN1 regulates these pro-fibrotic target genes by modulating the activity of regulatory modules that contain transcription factors of the specificity protein (SP) family. Functional experiments with selective modulators of ROCK and of microtubule polymerization confirm the coordinating role of EN1 on ROCK activity and the re-organization of cytoskeleton during myofibroblast differentiation in both conventional culture systems and 3D skin equivalents. Consistently, mice with fibroblast-specific knockout of En1 demonstrate impaired fibroblast-to-myofibroblast transition, reduced dermal thickening and impaired collagen deposition in the TBRICA, bleomycin-induced and TSK1 models.Conclusion:We characterize the homeodomain transcription factor EN1 as a molecular amplifier of TGFβ signaling in myofibroblast differentiation that coordinates cytoskeletal organization in a SP-dependent manner. EN1 might thus be a novel candidate for molecular targeted therapies to interfere with myofibroblast differentiation in fibrotic diseases.References:[1]Rinkevich Y, Walmsley GG, Hu MS, Maan ZN, Newman AM, Drukker M, et al. Skin fibrosis. Identification and isolation of a dermal lineage with intrinsic fibrogenic potential. Science. 2015;348(6232):aaa2151.Disclosure of Interests:Andrea-Hermina Györfi: None declared, Alexandru-Emil Matei: None declared, Maximilian Fuchs: None declared, Aleix Rius Rigau: None declared, Xuezhi Hong: None declared, ZHU Honglin: None declared, Markus Luber: None declared, Christina Bergmann: None declared, Clara Dees: None declared, Ingo Ludolph: None declared, Raymund Horch: None declared, Oliver Distler Consultant of: Actellion, AbbVie, Acceleron Pharma, Anamar, Amgen, Blade Therapeutics, CSL Behring, ChemomAb, Ergonex, Glenmark Pharma, GSK, Inventiva, Italfarmaco, iQvia, Medac, Medscape, Lilly, Sanofi, Target BioScience, UCB, Bayer, Boehringer Ingelheim, Catenion, iQone, Menarini, Mepha, Novartis, Mitsubishi, MSD, Roche, Pfizer, Georg Schett: None declared, Meik Kunz: None declared, Jörg H.W. Distler Consultant of: Actelion, Active Biotech, Anamar, ARXX, Bayer Pharma, Boehringer Ingelheim, Celgene, Galapagos, GSK, Inventiva, JB Therapeutics, Medac, Pfizer, RuiYi and UCB., Grant/research support from: Anamar, Active Biotech, Array Biopharma, aTyr, BMS, Bayer Pharma, Boehringer Ingelheim, Celgene, Galapagos, GSK, Inventiva, Novartis, Sanofi-Aventis, RedX, UCB
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Risau, W., H. Sariola, H. G. Zerwes, J. Sasse, P. Ekblom, R. Kemler, and T. Doetschman. "Vasculogenesis and angiogenesis in embryonic-stem-cell-derived embryoid bodies." Development 102, no. 3 (March 1, 1988): 471–78. http://dx.doi.org/10.1242/dev.102.3.471.
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Embryonic stem cells (ESC) have been established previously from the inner cell mass cells of mouse blastocysts. In suspension culture, they spontaneously differentiate to blood-island-containing cystic embryoid bodies (CEB). The development of blood vessels from in situ differentiating endothelial cells of blood islands, a process which we call vasculogenesis, was induced by injecting ESC into the peritoneal cavity of syngeneic mice. In the peritoneum, fusion of blood islands and formation of an in vivo-like primary capillary plexus occurred. Transplantation of ESC and ESC-derived complex and cystic embryoid bodies (ESC-CEB) onto the quail chorioallantoic membrane (CAM) induced an angiogenic response, which was directed by nonyolk sac endoderm structures. Neither yolk sac endoderm from ESC-CEB nor normal mouse yolk sac tissue induced angiogenesis on the quail CAM. Extracts from ESC-CEB stimulated the proliferation of capillary endothelial cells in vitro. Mitogenic activity increase during in vitro culture and differentiation of ESC. Almost all growth factor activity was associated with the cells. The ESC-CEB derived endothelial cell growth factor bound to heparin-sepharose. The identification of acidic fibroblast growth factor (FGF)in heparin-sepharose-purified material was accomplished by immunoblot experiments involving antibodies against acidic and basic FGF. We conclude that vasculogenesis, the development of blood vessels from in situ differentiating endothelial cells, and angiogenesis, the sprouting of capillaries from preexisting vessels are very early events during embryogenesis which can be studied using ESC differentiating in vitro. Our results suggest that vasculogenesis and angiogenesis are differently regulated.
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Cressman, Victoria L., Dana C. Backlund, Anna V. Avrutskaya, Steven A. Leadon, Virginia Godfrey, and Beverly H. Koller. "Growth Retardation, DNA Repair Defects, and Lack of Spermatogenesis in BRCA1-Deficient Mice." Molecular and Cellular Biology 19, no. 10 (October 1, 1999): 7061–75. http://dx.doi.org/10.1128/mcb.19.10.7061.
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ABSTRACT BRCA1 is a nuclear phosphoprotein expressed in a broad spectrum of tissues during cell division. The inheritance of a mutantBRCA1 allele dramatically increases a woman’s lifetime risk for developing both breast and ovarian cancers. A number of mouse lines carrying mutations in the Brca1 gene have been generated, and mice homozygous for these mutations generally die before day 10 of embryonic development. We report here the survival of a small number of mice homozygous for mutations in both the p53 andBrca1 genes. The survival of these mice is likely due to additional unknown mutations or epigenetic effects. Analysis of theBrca1−/− p53−/− animals indicates that BRCA1 is not required for the development of most organ systems. However, these mice are growth retarded, males are infertile due to meiotic failure, and the mammary gland of the female mouse is underdeveloped. Growth deficiency due to loss of BRCA1 was more thoroughly examined in an analysis of primary fibroblast lines obtained from these animals. Like p53−/− fibroblasts,Brca1−/− p53−/− cells proliferate more rapidly than wild-type cells; however, a high level of cellular death in these cultures results in reduced overall growth rates in comparison to p53−/− fibroblasts.Brca1−/− p53−/− fibroblasts are also defective in transcription-coupled repair and display increased sensitivity to DNA-damaging agents. We show, however, that after continued culture, and perhaps accelerated by the loss of BRCA1 repair functions, populations of Brca1−/−p53−/− fibroblasts with increased growth rates can be isolated. The increased survival of BRCA1-deficient fibroblasts in the absence of p53, and with the subsequent accumulation of additional growth-promoting changes, may mimic the events that occur during malignant transformation of BRCA1-deficient epithelia.
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Key, Jana, Patrick N. Harter, Nesli-Ece Sen, Elise Gradhand, Georg Auburger, and Suzana Gispert. "Mid-Gestation lethality of Atxn2l-Ablated Mice." International Journal of Molecular Sciences 21, no. 14 (July 20, 2020): 5124. http://dx.doi.org/10.3390/ijms21145124.
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Depletion of yeast/fly Ataxin-2 rescues TDP-43 overexpression toxicity. In mouse models of Amyotrophic Lateral Sclerosis via TDP-43 overexpression, depletion of its ortholog ATXN2 mitigated motor neuron degeneration and extended lifespan from 25 days to >300 days. There is another ortholog in mammals, named ATXN2L (Ataxin-2-like), which is almost uncharacterized but also functions in RNA surveillance at stress granules. We generated mice with Crispr/Cas9-mediated deletion of Atxn2l exons 5-8, studying homozygotes prenatally and heterozygotes during aging. Our novel findings indicate that ATXN2L absence triggers mid-gestational embryonic lethality, affecting female animals more strongly. Weight and development stages of homozygous mutants were reduced. Placenta phenotypes were not apparent, but brain histology showed lamination defects and apoptosis. Aged heterozygotes showed no locomotor deficits or weight loss over 12 months. Null mutants in vivo displayed compensatory efforts to maximize Atxn2l expression, which were prevented upon nutrient abundance in vitro. Mouse embryonal fibroblast cells revealed more multinucleated giant cells upon ATXN2L deficiency. In addition, in human neural cells, transcript levels of ATXN2L were induced upon starvation and glucose and amino acids exposure, but this induction was partially prevented by serum or low cholesterol administration. Neither ATXN2L depletion triggered dysregulation of ATXN2, nor a converse effect was observed. Overall, this essential role of ATXN2L for embryogenesis raises questions about its role in neurodegenerative diseases and neuroprotective therapies.
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Plotkin, Matthew D., and Michael S. Goligorsky. "Mesenchymal cells from adult kidney support angiogenesis and differentiate into multiple interstitial cell types including erythropoietin-producing fibroblasts." American Journal of Physiology-Renal Physiology 291, no. 4 (October 2006): F902—F912. http://dx.doi.org/10.1152/ajprenal.00396.2005.
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Mesenchymal cells have been isolated from embryos and multiple adult organs where they may differentiate into various connective tissue cell types and provide paracrine support for surrounding cells. With the use of a technique for culturing multipotent mesenchymal cells from adult tissues, a fibroblast-like cell clone (4E) was isolated from adult mouse kidney. 4E cells were able to differentiate along multiple mesodermal lineages including cell types located in the renal interstitium such as fibroblasts and pericytes. Coculture of 4E cells with ureteric bud and epithelial cell lines and analysis of resulting changes in gene expression revealed that these cells support angiogenesis and tubulogenesis and expressed genes characteristic of embryonic renal stromal cells. Following subcapsular injection after unilateral ischemia-reperfusion in adult mice, 4E cells migrated to a peritubular interstitial location and expressed interstitial cell markers, whereas cells injected in control kidneys remained stationary. Incubation in hypoxic or anoxic conditions resulted in erythropoietin expression in a small subset of ecto-5′-nucleotidase-positive cells and resulted in increased vascular endothelial growth factor expression in the same cell population. Our findings suggest that the adult kidney may contain interstitial mesenchymal cell progenitors with embryonic stromal cell characteristics that are able to provide paracrine support for surrounding vessels and tubular epithelial cells and differentiate into erythropoietin producing fibroblasts.
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Kim, Jin Woo, Tong-Shin Chang, Ji Eun Lee, Sung-Ho Huh, Seung Woo Yeon, Wan Seok Yang, Cheol O. Joe, et al. "Negative Regulation of the Sapk/Jnk Signaling Pathway by Presenilin 1." Journal of Cell Biology 153, no. 3 (April 24, 2001): 457–64. http://dx.doi.org/10.1083/jcb.153.3.457.
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Presenilin 1 (PS1) plays a pivotal role in Notch signaling and the intracellular metabolism of the amyloid β-protein. To understand intracellular signaling events downstream of PS1, we investigated in this study the action of PS1 on mitogen-activated protein kinase pathways. Overexpressed PS1 suppressed the stress-induced stimulation of stress-activated protein kinase (SAPK)/c-Jun NH2-terminal kinase (JNK) in human embryonic kidney 293 cells. Interestingly, two functionally inactive PS1 mutants, PS1(D257A) and PS1(D385A), failed to inhibit UV-stimulated SAPK/JNK. Furthermore, H2O2- or UV-stimulated SAPK activity was higher in mouse embryonic fibroblast (MEF) cells from PS1-null mice than in MEF cells from PS+/+ mice. MEFPS1(−/−) cells were more sensitive to the H2O2-induced apoptosis than MEFPS1(+/+) cells. Ectopic expression of PS1 in MEFPS1(−/−) cells suppressed H2O2-stimulated SAPK/JNK activity and apoptotic cell death. Together, our data suggest that PS1 inhibits the stress-activated signaling by suppressing the SAPK/JNK pathway.
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Zheng, Feng, Zhong Chen, Qiao-Li Tang, Xin-Ying Wang, Dan-Yang Chong, Tong-Yu Zhang, Ya-Yun Gu, Zhi-Bin Hu, and Chao-Jun Li. "Cholesterol metabolic enzyme Ggpps regulates epicardium development and ventricular wall architecture integrity in mice." Journal of Molecular Cell Biology 13, no. 6 (March 24, 2021): 445–54. http://dx.doi.org/10.1093/jmcb/mjab019.
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Abstract During embryonic heart development, the progenitor cells in the epicardium would migrate and differentiate into noncardiomyocytes in myocardium and affect the integrity of ventricular wall, but the underlying mechanism has not been well studied. We have found that myocardium geranylgeranyl diphosphate synthase (Ggpps), a metabolic enzyme for cholesterol biosynthesis, is critical for cardiac cytoarchitecture remodelling during heart development. Here, we further reveal that epicardial Ggpps could also regulate ventricular wall architecture integrity. Epicardium-specific deletion of Ggpps before embryonic day 10.5 (E10.5) is embryonic lethal, whereas after E13.5 is survival but with defects in the epicardium and ventricular wall structure. Ggpps deficiency in the epicardium enhances the proliferation of epicardial cells and disrupts cell‒cell contact, which makes epicardial cells easier to invade into ventricular wall. Thus, the fibroblast proliferation and coronary formation in myocardium were found enhanced that might disturb the coronary vasculature remodelling and ventricular wall integrity. These processes might be associated with the activation of YAP signalling, whose nuclear distribution is blocked by Ggpps deletion. In conclusion, our findings reveal a potential link between the cholesterol metabolism and heart epicardium and myocardium development in mammals, which might provide a new view of the cause for congenital heart diseases and potential therapeutic target in pathological cardiac conditions.
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Fuentes-Calvo, Isabel, Ana M. Blázquez-Medela, Nélida Eleno, Eugenio Santos, José M. López-Novoa, and Carlos Martínez-Salgado. "H-Ras isoform modulates extracellular matrix synthesis, proliferation, and migration in fibroblasts." American Journal of Physiology-Cell Physiology 302, no. 4 (February 15, 2012): C686—C697. http://dx.doi.org/10.1152/ajpcell.00103.2011.
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Ras GTPases are ubiquitous plasma membrane transducers of extracellular stimuli. In addition to their role as oncogenes, Ras GTPases are key regulators of cell function. Each of the Ras isoforms exhibits specific modulatory activity on different cellular pathways. This has prompted researchers to determine the pathophysiological roles of each isoform. There is a proven relationship between the signaling pathways of transforming growth factor-β1 (TGF-β1) and Ras GTPases. To assess the individual role of H-Ras oncogene in basal and TGF-β1-mediated extracellular matrix (ECM) synthesis, proliferation, and migration in fibroblasts, we analyzed these processes in embryonic fibroblasts obtained from H-Ras knockout mice ( H-ras−/−). We found that H- ras−/− fibroblasts exhibited a higher basal phosphatidylinositol-3-kinase (PI3K)/Akt activation than wild-type (WT) fibroblasts, whereas MEK/ERK 1/2 activation was similar in both types of cells. Fibronectin and collagen synthesis were higher in H -ras−/− fibroblasts and proliferation was lower in H -ras−/− than in WT fibroblasts. Moreover, H-Ras appeared indispensable to maintain normal fibroblast motility, which was highly restricted in H- ras−/− cells. These results suggest that H-Ras (through downregulation of PI3K/Akt activation) could modulate fibroblast activity by reducing ECM synthesis and upregulating both proliferation and migration. TGF-β1 strongly increased ERK and Akt activation in WT but not in H- ras−/− fibroblasts, suggesting that H-Ras is necessary to increase ERK 1/2 activation and to maintain PI3K downregulation in TGF-β1-stimulated fibroblasts. TGF-β1 stimulated ECM synthesis and proliferation, although ECM synthesis was higher and proliferation lower in H- ras−/− than in WT fibroblasts. Hence, H-Ras activation seems to play a key role in the regulation of these effects.
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Woo, Jiwon, Wonhee Suh, and Jonghyuk Sung. "Hair Growth Regulation by Fibroblast Growth Factor 12 (FGF12)." International Journal of Molecular Sciences 23, no. 16 (August 22, 2022): 9467. http://dx.doi.org/10.3390/ijms23169467.
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The fibroblast growth factor (FGF) family has various biological functions, including cell growth, tissue regeneration, embryonic development, metabolism, and angiogenesis. In the case of hair growth, several members of the FGF family, such as FGF1 and FGF2, are involved in hair growth, while FGF5 has the opposite effect. In this study, the regulation of the hair growth cycle by FGF12 was investigated. To observe its effect, the expression of FGF12 was downregulated in mice and outer root sheath (ORS) by siRNA transfection, while FGF12 overexpression was carried out using FGF12 adenovirus. For the results, FGF12 was primarily expressed in ORS cells with a high expression during the anagen phase of hair follicles. Knockdown of FGF12 delayed telogen-to-anagen transition in mice and decreased the hair length in vibrissae hair follicles. It also inhibited the proliferation and migration of ORS cells. On the contrary, FGF12 overexpression increased the migration of ORS cells. FGF12-overexpressed ORS cells induced the telogen-to-anagen transition in the animal model. In addition, FGF12 overexpression regulated the expression of PDGF-CC, MDK, and HB-EGF, and treatment of these factors exhibited hair growth promotion. Altogether, FGF12 promoted hair growth by inducing the anagen phase of hair follicles, suggesting the potential for hair loss therapy.
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Alipio, Zaida, Dan Xu, Jianchang Yang, Louis M. Fink, Wilson Xu, David C. Ward, and Yupo Ma. "Reprogrammed Murine Fibroblasts Differentiated into Hematopoietic Progenitors Are Able to Successfully Engraft and Repopulate the Bone Marrow." Blood 112, no. 11 (November 16, 2008): 389. http://dx.doi.org/10.1182/blood.v112.11.389.389.
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Abstract Cellular therapy using embryonic stem cells has always been an area of great interest due to the pluripotent characteristics of stem cells. In 2006, Takahashi and Yamanaka (Cell 126, 663–676) demonstrated that somatic cells can be reprogrammed into a stem cell-like state, termed induced pluripotent stem (iPS) cells, by ectopic expression of Oct4, Sox2, Klf4 and c Myc. A later report (Nakagawa et al. Nat. Biotechnol.26:101–106, 2008) showed that iPS cells can be produced in the absence of the c Myc oncogene. We have used this latter strategy to successfully reprogram somatic cells derived from C57BL/6 mouse tail fibroblast to iPS cells. Retrovirus infected fibroblasts exhibited stem cell-like morphology by 14 days post infection. These iPS cells were then infected with a retrovirus that expressed HOXB4. Recombinant leukemia inhibitor factor (LIF) supplement was removed from media at this time and the cells allowed to differentiate into embryoid bodies. These cells were screened for specific differentiation stem cell markers, such as Oct4, Nanog, Sall4 and SSEA-1. iPS cells were converted into embryonic bodies and then infected with retroviruses expressing HOXB4. Embryoid bodies stably expressing HOXB4 were induced to hematopoietic differentiation by treatment of thrombopoietin (TPO), stem cell factor (SCF), vascular endothelial growth factor (VEGF), interferon gamma (IFNg) and fms-like tyrosine kinase (FLT3 ligand). Evaluation of iPS-derived hematopoietic cells on smears show strikingly similarity in morphology to the W4 mouse embryonic stem (ES) cells differentiated into hematopoietic cells as a control. Flow cytometry analysis of iPS-derived hematopoietic cells after 1 week exposure to cytokines revealed 7% B220+ cells (B cells), 11% Ter119+ cells (erythroid), and 13% Gr-1+ cells (granulocytes) similar to W4 ES cells. The iPS-derived hematopoietic cells were transplanted into irradiated immunodeficient mice via lateral tail vein injection. Transplantation of these iPS-derived hematopoietic progenitors tagged with GFP into irradiated SCID mice revealed that the hematopoietic progenitors were able to home to the bone marrow after 1 week of transplantation. Importantly, after 1 month, GFP+ engrafted cells remained in the bone marrow suggesting a long-term engraftment. This long term engraftment of the iPS-derived hematopoietic cells to the bone marrow constitutes an important step toward potential therapy of numerous patient-specific blood based diseases.
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Lamanna, William C., Rebecca J. Baldwin, Michael Padva, Ina Kalus, Gerdy ten Dam, Toin H. van Kuppevelt, John T. Gallagher, Kurt von Figura, Thomas Dierks, and Catherine L. R. Merry. "Heparan sulfate 6-O-endosulfatases: discrete in vivo activities and functional co-operativity." Biochemical Journal 400, no. 1 (October 27, 2006): 63–73. http://dx.doi.org/10.1042/bj20060848.
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HS (heparan sulfate) is essential for normal embryonic development. This requirement is due to the obligatory role for HS in the signalling pathways of many growth factors and morphogens that bind to sulfated domains in the HS polymer chain. The sulfation patterning of HS is determined by a complex interplay of Golgi-located N- and O-sulfotransferases which sulfate the heparan precursor and cell surface endosulfatases that selectively remove 6-O-sulfates from mature HS chains. In the present study we generated single or double knock-out mice for the two murine endosulfatases mSulf1 and mSulf2. Detailed structural analysis of HS from mSulf1−/− fibroblasts showed a striking increase in 6-O-sulfation, which was not seen in mSulf2−/− HS. Intriguingly, the level of 6-O-sulfation in the double mSulf1−/−/2−/− HS was significantly higher than that observed in the mSulf1−/− counterpart. These data imply that mSulf1 and mSulf2 are functionally co-operative. Unlike their avian orthologues, mammalian Sulf activities are not restricted to the highly sulfated S-domains of HS. Mitogenesis assays with FGF2 (fibroblast growth factor 2) revealed that Sulf activity decreases the activating potential of newly-synthesized HS, suggesting an important role for these enzymes in cell growth regulation in embryonic and adult tissues.
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Vilarino, Marcela, Delia Alba Soto, Yanina Soledad Bogliotti, Leqian Yu, Yanli Zhang, Chunsheng Wang, Erika Paulson, et al. "Derivation of sheep embryonic stem cells under optimized conditions." Reproduction 160, no. 5 (November 2020): 761–72. http://dx.doi.org/10.1530/rep-19-0606.
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Until recently, it has been difficult to derive and maintain stable embryonic stem cells lines from livestock species. Sheep ESCs with characteristics similar to those described for rodents and primates have not been produced. We report the derivation of sheep ESCs under a chemically defined culture system containing fibroblast growth factor 2 (FGF2) and a tankyrase/Wnt inhibitor (IWR1). We also show that several culture conditions used for stabilizing naïve and intermediate pluripotency states in humans and mice were unsuitable to maintain ovine pluripotency in vitro. Sheep ESCs display a smooth dome-shaped colony morphology, and maintain an euploid karyotype and stable expression of pluripotency markers after more than 40 passages. We further demonstrate that IWR1 and FGF2 are essential for the maintenance of an undifferentiated state in de novo derived sheep ESCs. The derivation of stable pluripotent cell lines from sheep blastocysts represents a step forward toward understanding pluripotency regulation in livestock species and developing novel biomedical and agricultural applications.
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Piedrahita, Jorge A., Sehwon Koh, and Natasha Olby. "GENERATION AND CHARACTERISATION OF INDUCED PLURIPOTENT STEM CELLS (iPSCs) FROM ADULT CANINE FIBROBLASTS." Reproduction, Fertility and Development 24, no. 1 (2012): 285. http://dx.doi.org/10.1071/rdv24n1ab244.
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Pluripotent stem cells such as embryonic stem cells (ESCs) and induced pluripotent stem cells (iPSCs) can give rise to derivatives of all three germ layers and thus have great potential in regenerative medicine. In mice and humans, it has been shown that embryonic and adult fibroblasts can be reprogrammed into pluripotency by introducing four transcription factors, Oct3/4, Klf4, Sox2 and c-Myc (OKSM). In his presentation we will describe the derivation of iPS cells from adult canine fibroblast by retroviral OSKM transduction. The isolated canine iPS cells were expanded in three different iPS culture media (FGF2, LIF and FGF2 plus LIF) and only the cells cultured in FGF2 plus LIF showed strong AP activity expressed pluripotency markers, POU5F1 (OCT4), SOX2, NANOG and LIN28 as well as ES cells-specific genes (PODXL, DPPA5, FGF5, REX1 and LAMP1). In vitro differentiation by formation of embryoid bodies (EBs) and directed differentiation showed cell derivatives of all three germ layers as confirmed by expression for AFP, CXCR4 and SOX17 (endoderm), desmin (DES), vimentin (VIM), MSX1 and BMP2 (mesoderm) and glial fibrillary acidic protein (GFAP), TUJ1, NCAM and bIII-tubulin (TUBB, ectoderm). In vivo, the putative canine iPS cells formed simple teratomas that expressed markers for all three germ layers. In summary, we were able to derive induced pluripotent cells from adult somatic cells by using four transcription factors. The isolated canine iPSCs have similar characteristics to ESCs from other species, but the exact cellular mechanisms behind their unique co-dependency on both FGF and LIF is still unknown. This work was funded by a grant from the America Kennel Club to JAP.
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Sims-Lucas, Sunder, Luise Cullen-McEwen, Veraragavan P. Eswarakumar, David Hains, Kayle Kish, Brian Becknell, Jue Zhang, John F. Bertram, Fen Wang, and Carlton M. Bates. "Deletion of Frs2α from the ureteric epithelium causes renal hypoplasia." American Journal of Physiology-Renal Physiology 297, no. 5 (November 2009): F1208—F1219. http://dx.doi.org/10.1152/ajprenal.00262.2009.
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Fibroblast growth factor receptor 2 (Fgfr2) signaling is critical in maintaining ureteric branching architecture and mesenchymal stromal morphogenesis in the kidney. Fibroblast growth factor receptor substrate 2α (Frs2α) is a major docking protein for Fgfr2 with downstream targets including Ets variant (Etv) 4 and Etv5 in other systems. Furthermore, global deletion of Frs2α causes early embryonic lethality. The purpose of the study was to determine the role of Frs2α in mediating Fgfr2 signaling in the ureteric epithelium. To that end, we generated mice with conditional deletion of Frs2α in the ureteric epithelium ( Frs2α UB−/−) and mice with point mutations in the Frs2α binding site of Fgfr2 ( Fgfr2 LR/LR). Frs2α UB−/− mice developed mild renal hypoplasia characterized by decreased ureteric branching morphogenesis but maintained normal overall branching architecture and had normal mesenchymal stromal development. Reduced nephron endowment in postnatal mutant mice was observed, corresponding with the reduction in branching morphogenesis. Furthermore, there were no apparent renal abnormalities in Fgfr2 LR/LR mice. Interestingly, Etv4 and Etv5 expression was unaltered in Frs2α UB−/− mice, as was Sprouty1, an antagonist of Frs2α signaling. However, Ret and Wnt11 (molecules critical for ureteric branching morphogenesis) mRNA levels were lower in mutants vs. controls. Taken together, these findings suggest that Fgfr2 signals through adapter molecules other than Frs2α in the ureteric epithelium. Furthermore, Frs2α may transmit signals through other receptor kinases present in ureteric epithelium. Finally, the renal hypoplasia observed in Frs2α UB−/− mice is likely secondary to decreased Ret and Wnt11 expression.
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Kaufman, Dan S., Rachel L. Lewis, Eric T. Hanson, Robert Auerbach, Johanna Plendl, and James A. Thomson. "Functional endothelial cells derived from rhesus monkey embryonic stem cells." Blood 103, no. 4 (February 15, 2004): 1325–32. http://dx.doi.org/10.1182/blood-2003-03-0799.
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Abstract We have used rhesus monkey embryonic stem (ES) cells to study endothelial cell development. Rhesus ES cells (R366.4 cell line) exposed to medium containing vascular endothelial growth factor (VEGF), basic fibroblast growth factor (bFGF), insulin-like growth factor (IGF), and epidermal growth factor (EGF) assumed a relatively uniform endothelial cell morphology and could be propagated and expanded with a consistent phenotype and normal karyotype. When placed in Matrigel, these rhesus ES cell–derived endothelial cells (RESDECs) formed capillary-like structures characteristic of endothelial cells. Immunohistochemical and flow cytometric analysis of RESDECs showed that they take up acetylated low-density lipoprotein (LDL), express CD146, von Willebrand factor, and the integrin αvβ3, and bind the lectin ulex europaeus agglutinin-1. These cells also express the VEGF receptor Flk-1 and secrete VEGF. When introduced in a Matrigel plug implanted subcutaneously in mice, RESDECs formed intact vessels and recruited new endothelial cell growth. In vivo function was demonstrated by coinjection of RESDECs with murine tumor cells subcutaneously into immunocompromised adult mice. RESDECs injected alone did not form measurable tumors. Tumor cells grew more rapidly and had increased vascularization when coinjected with the RESDECs. Immunohistochemical staining demonstrated that the RESDECs participated in forming the tumor neovasculature. RESDECs provide a novel means to examine the mechanisms of endothelial cell development, and may open up new therapeutic strategies.
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Pozzi, Ambra, Kishore K. Wary, Filippo G. Giancotti, and Humphrey A. Gardner. "Integrin α1β1 Mediates a Unique Collagen-dependent Proliferation Pathway In Vivo." Journal of Cell Biology 142, no. 2 (July 27, 1998): 587–94. http://dx.doi.org/10.1083/jcb.142.2.587.
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Activation of integrins upon binding to extracellular matrix proteins is believed to be a crucial step for the regulation of cell survival and proliferation. We have used integrin α1-null mice to investigate the role of this collagen receptor in the regulation of cell growth and survival in vivo. α1-deficient animals, which are viable and fertile, have a hypocellular dermis and a deficiency in dermal fibroblast proliferation as embryos. In vitro analysis of α1-null embryonic fibroblasts has revealed that their proliferation rate is markedly reduced when plated on collagenous substrata, despite normal attachment and spreading. Moreover, on the same collagenous matrices, α1-null fibroblasts fail to recruit and activate the adaptor protein Shc. The failure to activate Shc is accompanied by a downstream deficiency in recruitment of Grb2 and subsequent mitogen-activated protein kinase activation. Taken together with the growth deficiency observed on collagens, this finding indicates that the α1β1 is the sole collagen receptor which can activate the Shc mediated growth pathway. Thus, integrin α1 has a unique role among the collagen receptors in regulating both in vivo and in vitro cell proliferation in collagenous matrices.
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Diatroptova, M. A., E. A. Ponomarenko, and M. E. Diatroptov. "Infradian Rhythm in Proliferative Activity of a Culture of Embryonic Fibroblast-Like Cells from C57BL/6 Mice." Bulletin of Experimental Biology and Medicine 169, no. 5 (September 2020): 714–17. http://dx.doi.org/10.1007/s10517-020-04962-y.
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Yagiz, Kader, Lion-Ying Wu, Charles P. Kuntz, D. James Morré, and Dorothy M. Morré. "Mouse embryonic fibroblast cells from transgenic mice overexpressing tNOX exhibit an altered growth and drug response phenotype." Journal of Cellular Biochemistry 101, no. 2 (2007): 295–306. http://dx.doi.org/10.1002/jcb.21184.
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Wu, Weiju, Chengfei Liu, Conrad A. Farrar, Liang Ma, Xia Dong, Steven H. Sacks, Ke Li, and Wuding Zhou. "Collectin-11 Promotes the Development of Renal Tubulointerstitial Fibrosis." Journal of the American Society of Nephrology 29, no. 1 (November 15, 2017): 168–81. http://dx.doi.org/10.1681/asn.2017050544.
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Collectin-11 is a recently described soluble C-type lectin, a pattern recognition molecule of the innate immune system that has distinct roles in host defense, embryonic development, and acute inflammation. However, little is known regarding the role of collectin-11 in tissue fibrosis. Here, we investigated collectin-11 in the context of renal ischemia-reperfusion injury. Compared with wild-type littermate controls, Collec11 deficient (CL-11−/−) mice had significantly reduced renal functional impairment, tubular injury, renal leukocyte infiltration, renal tissue inflammation/fibrogenesis, and collagen deposition in the kidneys after renal ischemia-reperfusion injury. In vitro, recombinant collectin-11 potently promoted leukocyte migration and renal fibroblast proliferation in a carbohydrate-dependent manner. Additionally, compared with wild-type kidney grafts, CL-11−/−mice kidney grafts displayed significantly reduced tubular injury and collagen deposition after syngeneic kidney transplant. Our findings demonstrate a pathogenic role for collectin-11 in the development of tubulointerstitial fibrosis and suggest that local collectin-11 promotes this fibrosis through effects on leukocyte chemotaxis and renal fibroblast proliferation. This insight into the pathogenesis of tubulointerstitial fibrosis may have implications for CKD mediated by other causes as well.
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Li, Zhuangwu, Marina Jerebtsova, Xue-Hui Liu, Pingtao Tang, and Patricio E. Ray. "Novel cystogenic role of basic fibroblast growth factor in developing rodent kidneys." American Journal of Physiology-Renal Physiology 291, no. 2 (August 2006): F289—F296. http://dx.doi.org/10.1152/ajprenal.00382.2005.
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Basic fibroblast growth factor (bFGF) is a heparin-binding growth factor that is accumulated in human dysplastic and cystic renal diseases. Previous studies have shown that bFGF can modulate the growth of developing renal tubules; however, its role in the pathogenesis of renal cyst formation is not clearly understood. Here, we tested the hypothesis that overexpression of bFGF in developing rodent kidneys induces cyst formation in vivo. We used two different adenoviral-mediated gene-transferring approaches to overexpress bFGF in developing rodent kidneys. Initially, metanephric kidney (MK) explants harvested from embryonic day 15 Sprague-Dawley rats were infected with adenoviral vectors (rAd) encoding human bFGF or LacZ genes and transplanted under the renal capsule of adult female rats. Subsequently, to determine whether bFGF could induce renal cysts in developing kidneys with an intact renal collecting system, we injected rAd-bFGF or LacZ vectors in the retroorbital plexus of newborn mice. Basic FGF induced a more efficient integration of the MK explants into the host kidneys and increased the vascularization and proliferation of developing tubules, leading to tubular dilatation and rapid formation of renal cysts. In addition, we successfully expressed human bFGF in the kidney of newborn mice in vivo and induced tubular dilatation and renal cysts. In contrast, mice injected with rAd- lacZ did not develop tubular dilatation or renal cysts. To the best of our knowledge, these experiments show for the first time that overexpression of bFGF in developing rodent kidneys can induce the formation of renal cysts in vivo.
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Holley, Rebecca J., Kate A. Meade, and Catherine L. R. Merry. "Using embryonic stem cells to understand how glycosaminoglycans regulate differentiation." Biochemical Society Transactions 42, no. 3 (May 22, 2014): 689–95. http://dx.doi.org/10.1042/bst20140064.
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Differentiation and subsequent specialization of every cell within an organism is an intricate interwoven process. A complex network of signalling pathways eventually leads to the specification of a multitude of different cell types able to function co-operatively. HS (heparan sulfate) is a highly sulfated linear polysaccharide that resides at the pericellular cell–matrix interface where it dictates the binding and activity of a large number of proteins, including growth factors and morphogens such as members of the FGF (fibroblast growth factor) and BMP (bone morphogenetic protein) families. Embryonic stem cells derived from mice with mutations in components of the HS biosynthetic pathway provide an opportunity to dissect the contribution of HS to signalling pathways critical for regulating stem cell maintenance and differentiation. In addition to improving our understanding of signalling mechanisms, this knowledge enables the selection of exogenous HS saccharides to improve the efficiency and selectivity of directed differentiation protocols, offering a cost-effective alternative to high concentrations of expensive growth factors to drive differentiation towards a particular therapeutically relevant cell type.
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Haase, Matthias, Matthias Schott, Stefan R. Bornstein, Ludwik K. Malendowicz, Werner A. Scherbaum, and Holger S. Willenberg. "CITED2 is expressed in human adrenocortical cells and regulated by basic fibroblast growth factor." Journal of Endocrinology 192, no. 2 (February 2007): 459–65. http://dx.doi.org/10.1677/joe-06-0083.
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CITED2 gene deletion in mice leads to adrenal agenesis. Therefore, we analyzed CITED2, a CBP/p300 interacting transactivator with transforming activity, in the human adrenal gland. In this study, we examined CITED2 expression in human embryonic and adult adrenal glands as well as adrenocortical carcinomas. As ACTH and basic fibroblast growth factor (bFGF) are connected to the physiology and growth of adrenocortical cells we studied the regulation of CITED2 by these factors in the NCI-H295R adrenocortical carcinoma cell line. We found CITED2 expression in the adult adrenal cortex as well in adrenocortical carcinomas. At an early stage of human adrenal organogenesis CITED2 could be located to the definitive zone of the developing adrenal gland using immunohistochemistry. In NCI-H295R cells, stimulation by bFGF led to a dose-dependent increase in CITED2 promotor activity, mRNA and protein expression while ACTH had no significant effect. The stimulatory effect of bFGF could be reduced by blocking mitogen-activated protein kinase activity using the MAPkinase kinase (MEK1)-inhibitor PD98059. CITED2 is expressed in embryonic and adult human adrenal glands as well as in adrenocortical cancer. It is connected to the signaling cascades of bFGF and its expression is modulated by mitogen-activated protein kinases. This suggests a novel role for CITED2 in human adrenal growth and possibly in adrenal tumorigenesis.
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Wu, Xiaoting, Hyeran Won, and David C. Rubinsztein. "Autophagy and mammalian development." Biochemical Society Transactions 41, no. 6 (November 20, 2013): 1489–94. http://dx.doi.org/10.1042/bst20130185.
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Autophagy is a highly conserved cytoplasmic degradation pathway that has an impact on many physiological and disease states, including immunity, tumorigenesis and neurodegeneration. Recent studies suggest that autophagy may also have important functions in embryogenesis and development. Many autophagy gene-knockout mice have embryonic lethality at different stages of development. Furthermore, interactions of autophagy with crucial developmental pathways such as Wnt, Shh (Sonic Hedgehog), TGFβ (transforming growth factor β) and FGF (fibroblast growth factor) have been reported. This suggests that autophagy may regulate cell fate decisions, such as differentiation and proliferation. In the present article, we discuss how mammalian autophagy may affect phenotypes associated with development.
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Li, Yong, Zhi-Yu Ni, Meng-Chu Zhu, Mei Dong, Si-Ming Wang, Qing-Wen Shi, Man-Li Zhang, et al. "Antitumour Activities of Sesquiterpene Lactones from Inula helenium and Inula japonica." Zeitschrift für Naturforschung C 67, no. 7-8 (August 1, 2012): 375–80. http://dx.doi.org/10.1515/znc-2012-7-804.
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3 Eight sesquiterpene lactones were isolated from the roots of Inula helenium and fl owers of I. japonica. Among them, isoalantolactone () and santamarine (6) exhibited significant growth inhibitory activities against gynecologic cancer cell lines, while others weakly inhibited the growth of the cell lines (IC50 ≤ 100 μM). In addition, 3 significantly inhibited the tumour growth of S180 tumour-bearing mice. Compounds 3 and 6 were not toxic to human embryonic lung fibroblast cells in vitro. These results demonstrated that the antitumour activities are closely related to the structures of the compounds, that is, an α-exomethylene- γ-lactone ring is necessary for these activities.
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Filmus, Jorge. "The function of glypicans in the mammalian embryo." American Journal of Physiology-Cell Physiology 322, no. 4 (April 1, 2022): C694—C698. http://dx.doi.org/10.1152/ajpcell.00045.2022.
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Glypicans are proteoglycans that are bound to the outer surface of the plasma membrane by a glycosylphosphatidylinositol anchor. The mammalian genome contains six members of the glypican family ( GPC1 to GPC6). Although the degree of sequence homology within the family is rather low, the three-dimensional structure of these proteoglycans is highly conserved. Glypicans are predominantly expressed during embryonic development. Genetic and biochemical studies have shown that glypicans can stimulate or inhibit the signaling pathways triggered by Wnts, hedgehogs, fibroblast growth factors, and bone morphogenetic proteins. The study of mutant mouse strains demonstrated that glypicans have important functions in the developmental morphogenesis of various organs. In addition, a role of glypicans in synapsis formation has been established. Notably, glypican loss-of-function mutations are the cause of three human inherited syndromes. Recent analysis of glypican compound mutant mice has demonstrated that members of this protein family display redundant functions during embryonic development.
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Symons, Rebecca A., Fabio Colella, Fraser L. Collins, Alexandra J. Rafipay, Karolina Kania, Jessica J. McClure, Nathan White, et al. "Targeting the IL-6–Yap–Snail signalling axis in synovial fibroblasts ameliorates inflammatory arthritis." Annals of the Rheumatic Diseases 81, no. 2 (November 29, 2021): 214–24. http://dx.doi.org/10.1136/annrheumdis-2021-220875.
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ObjectiveWe aimed to understand the role of the transcriptional co-factor Yes-associated protein (Yap) in the molecular pathway underpinning the pathogenic transformation of synovial fibroblasts (SF) in rheumatoid arthritis (RA) to become invasive and cause joint destruction.MethodsSynovium from patients with RA and mice with antigen-induced arthritis (AIA) was analysed by immunostaining and qRT-PCR. SF were targeted using Pdgfrα-CreER and Gdf5-Cre mice, crossed with fluorescent reporters for cell tracing and Yap-flox mice for conditional Yap ablation. Fibroblast phenotypes were analysed by flow cytometry, and arthritis severity was assessed by histology. Yap activation was detected using Yap–Tead reporter cells and Yap–Snail interaction by proximity ligation assay. SF invasiveness was analysed using matrigel-coated transwells.ResultsYap, its binding partner Snail and downstream target connective tissue growth factor were upregulated in hyperplastic human RA and in mouse AIA synovium, with Yap detected in SF but not macrophages. Lineage tracing showed polyclonal expansion of Pdgfrα-expressing SF during AIA, with predominant expansion of the Gdf5-lineage SF subpopulation descending from the embryonic joint interzone. Gdf5-lineage SF showed increased expression of Yap and adopted an erosive phenotype (podoplanin+Thy-1 cell surface antigen−), invading cartilage and bone. Conditional ablation of Yap in Gdf5-lineage cells or Pdgfrα-expressing fibroblasts ameliorated AIA. Interleukin (IL)-6, but not tumour necrosis factor alpha (TNF-α) or IL-1β, Jak-dependently activated Yap and induced Yap–Snail interaction. SF invasiveness induced by IL-6 stimulation or Snail overexpression was prevented by Yap knockdown, showing a critical role for Yap in SF transformation in RA.ConclusionsOur findings uncover the IL-6–Yap–Snail signalling axis in pathogenic SF in inflammatory arthritis.
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Son, H. Y., M. G. Kim, J. I. Yun, J. E. Kim, H. S. Kim, S. K. Kang, B. C. Lee, W. S. Hwang, and C. K. Lee. "179 ISOLATION AND CULTURE OF EMBRYONIC GERM-LIKE CELLS FROM PORCINE MESONEPHROS." Reproduction, Fertility and Development 17, no. 2 (2005): 240. http://dx.doi.org/10.1071/rdv17n2ab179.
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Many attempts to establish embryonic stem (ES) cells from pre-implantation stage embryos in pigs have failed. An alternate source of pluripotent stem cells is embryonic germ (EG) cells derived from primordial germ cells (PGCs) of the genital ridge, which is developed from the mesonephros. Mesonephros is a vestigial, transient renal organ that functions only during embryonic development. It is believed to be a source of multiple stem cells including somatic cells in the gonad, vascular endothelial cells, and hematopoietic stem cells. Therefore, we tried to obtain putative stem cells from cells isolated from porcine mesonephros under culture conditions used to establish EG cells. Porcine fetuses from crossbred gilts were collected by hysterectomy between Days 25 and 30 of pregnancy (estrus = Day 0). Mesonephros and genital ridges were separated from each other, and cells from the mesonephros and PGCs from genital ridges were isolated by a physical method. Isolated cells were cultured in PES medium [50:50 mixture of Dulbecco's modified Eagle's medium (DMEM) and Ham's F10 medium supplemented with 15% fetal bovine serum (FBS), l-glutamine (1.7 mM), β-mercaptoethanol (0.1 mM), 1% MEM non-essential amino acids and 1% antibiotic-antimycotic] containing the cytokines, soluble recombinant human basic fibroblast growth factor (bFGF; 20 ng/mL), and human leukemia inhibitory factor (hLIF; 10 ng/mL). Isolated cells were cultured on fresh primary murine embryonic fibroblast feeder cells (PMEF) in a humidified environment of 5% CO2 in air at 38°C. The colonies with EG-like morphology were dissociated with 0.25% trypsin/1 mM EDTA for 10 min, and passed to fresh feeders. After 5–8 days, colonies started to grow with typical EG-like morphology. More colonies were obtained from the culture of cells from mesonephros than from culture of PGCs. Porcine EG-like cells from mesonephros (pMN-EG-like cells) were passed to fresh feeder every 6–8 days and have been cultured up to 9 passages while maintaining typical EG-like morphology. pMN-EG-like cells were stained for alkaline phosphatase throughout the culture. Furthermore, these cells reacted with antibodies against Oct-4 and SSEA-1 by immunocytochemistry, indicating that these cells have characteristics of pluripotential stem cells. In order to characterize the pMN-EG-like cells with respect to their potential for differentiation, embryoid body (EB) formation was induced. EBs started to form in 4 days and cystic structures in 2 weeks. EBs were then attached to the dish and cultured without cytokines. Spontaneously, EBs from pMN-EG-like cells could give rise to differentiated cell types such as neuronal-like, epithelial-like, and fibroblast-like cells. Further studies to characterize differentiated cells from pMN-EG-like cells by immunocytochemistry and for teratoma formation by injection into SCID mice will be performed. In conclusion, EG-like cells could be obtained from culture of mesonephric cells from porcine fetus and further characterization of these cells is required.