scholarly journals Characterization of Mice with a Platelet-Specific Deletion of the Adapter Molecule ADAP

2019 ◽  
Vol 39 (9) ◽  
Author(s):  
Jochen Michael Rudolph ◽  
Karina Guttek ◽  
Gabriele Weitz ◽  
Clara Antonia Meinke ◽  
Stefanie Kliche ◽  
...  
Keyword(s):  
T Cells ◽  
Knockout Mice ◽  
Transforming Growth Factor ◽  
Severe Disease ◽  
Transforming Growth Factor Β1 ◽  
Regulatory Function ◽  
Platelet Factor ◽  
Tgf Β1

ABSTRACT The adhesion and degranulation-promoting adapter protein (ADAP) is expressed in T cells, NK cells, myeloid cells, and platelets. The involvement of ADAP in the regulation of receptor-mediated inside-out signaling leading to integrin activation is well characterized, especially in T cells and in platelets. Due to the fact that animal studies using conventional knockout mice are limited by the overlapping effects of the different ADAP-expressing cells, we generated conditional ADAP knockout mice (ADAPfl/fl PF4-Cretg) (PF4, platelet factor 4). We observed that loss of ADAP restricted to the megakaryocytic lineage has no impact on other hematopoietic cells even under stimulation conditions. ADAPfl/fl PF4-Cretg mice showed thrombocytopenia in combination with reduced plasma levels of PF4 and transforming growth factor β1 (TGF-β1). In vitro, platelets from these mice revealed reduced P-selectin expression, lower levels of TGF-β1 release, diminished integrin αIIbβ3 activation, and decreased fibrinogen binding after stimulation with podoplanin, the ligand of C-type lectin-like receptor 2 (CLEC-2). Furthermore, loss of ADAP was associated with impaired CLEC-2-mediated activation of phospholipase Cγ2 (PLCγ2) and extracellular signal-regulated kinase 1/2 (ERK1/2). Induction of experimental autoimmune encephalomyelitis (EAE) in mice lacking ADAP expression in platelets caused a more severe disease. In vivo administration of TGF-β1 early after T cell transfer reduced EAE severity in mice with loss of ADAP restricted to platelets. Our results reveal a regulatory function of ADAP in platelets in vitro and during autoimmune disease EAE in vivo.

Identification of Platelet Releasate Proteins that Bind to Mastoparan, a Peptide that Inhibits Shear-Induced TGF-β1 Activation,

Blood ◽  
2011 ◽  
Vol 118 (21) ◽  
pp. 3271-3271
Author(s):  
Teresa M Brophy ◽  
Jasimuddin Ahamed ◽  
Barry S. Coller
Keyword(s):  
Transforming Growth Factor ◽  
Coagulation Factor ◽  
Transforming Growth Factor Β1 ◽  
Affinity Column ◽  
Factor V ◽  
Disulfide Exchange ◽  
Control Peptide ◽  
Tgf Β1

Abstract Abstract 3271 Transforming growth factor β1 (TGF-β1) is a disulfide-bonded, 25 kD homodimeric protein produced by most cell types, including platelets, that functions as a cytokine in many physiologic and pathologic processes. Platelets contain 40–100 times more TGF-β1 than other cells and release it as an inactive large latent complex (LLC) comprised of TGF-β1 non-covalently associated with its latency-associated peptide (LAP), which is, in turn, disulfide-bonded to latent TGF-β binding protein 1 (LTBP-1). Thrombospondin-1 (TSP1), proteases, and reactive oxygen species have all been shown to activate TGF-β1 in vitro and a role for integrins in vivo has been inferred from studies of transgenic mice. Recently, we discovered that shear force can activate latent TGF-β1 released from platelets in vitro and that thiol-disulfide exchange contributes to shear-dependent TGF-β1 activation. A number of thiol isomerase enzymes that can catalyze thiol-disulfide exchange have been identified in platelets, including protein disulfide isomerase (PDI), ERp5, ERp57, ERp72, ERp44, ERp29, and TMX3. As shear-induced activation of TGF-β1 is partially thiol-dependent, we investigated if thiol isomerases can affect this process. Mastoparan is a non-thiol-containing wasp venom peptide known to inhibit the chaperone activity of PDI, ERp5, and perhaps other thiol isomerases. We recently showed that mastoparan, (INLKALAALAKKIL), inhibits stirring-induced TGF-β1 activation by more than 90% (100 μM; n=3, p=0.03), whereas no inhibition was observed with an inactive mastoparan-like control peptide (INLKAKAALAKKLL) at 100 μM (n=3, p=0.66). To identify the proteins that bind to mastoparan, either directly or indirectly, platelet releasates were chromatographed on a mastoparan affinity column prepared from N-hydroxysuccinimide Sepharose. Two control columns were employed: 1. unconjugated Sepharose, and 2. Sepharose conjugated with the mastoparan-like control peptide. Elution of bound proteins was achieved by increasing the NaCl concentration. Proteins identified by mass spectrometry as specifically binding to the mastoparan peptide column included LTBP-1, TGF-β1 precursor, clusterin, coagulation Factor V, multimerin-1, 14-3-3 protein zeta/delta, and α-actinin 4. These results were confirmed by immunoblotting. Furthermore, the thiol isomerases PDI, ERp5, ERp57, and ERp72 were all found to bind specifically to mastoparan as confirmed by immunoblotting. We conclude that mastoparan affinity chromatography identified a number of proteins in platelet releasates that may contribute to shear-induced TGF-β1 activation. Disclosures: Coller: Centocor/Accumetrics/Rockefeller University:.

CD28 disruption exacerbates inflammation in Tgf-β1-/- mice: in vivo suppression by CD4+CD25+ regulatory T cells independent of autocrine TGF-β1

Blood ◽  
2004 ◽  
Vol 103 (12) ◽  
pp. 4594-4601 ◽  
Author(s):  
Mizuko Mamura ◽  
WoonKyu Lee ◽  
Timothy J. Sullivan ◽  
Angelina Felici ◽  
Anastasia L. Sowers ◽  
...  
Keyword(s):  
T Cells ◽  
Regulatory T Cells ◽  
Transforming Growth Factor ◽  
Cytotoxic T Lymphocyte ◽  
Tumor Necrosis Factor Receptor ◽  
Transforming Growth Factor Β1 ◽  
Autoimmune Inflammatory Disease ◽  
Lymph Node Cells ◽  
Tgf Β1

Abstract Tgf-β1-/- mice develop a progressive, lethal, inflammatory syndrome, but mechanisms leading to the spontaneous activation of Tgf-β1-/- T cells remain unclear. Here we show the disruption of CD28 gene expression accelerates disease in Tgf-β1-/- mice, and we link this increase in severity to a reduction in the number of CD4+CD25+ regulatory T cells. CD4+CD25+ T cells develop normally in Tgf-β1-/- mice and display characteristic expression of cytotoxic T lymphocyte-associated antigen 4 (CTLA-4), glucocorticoid-induced tumor necrosis factor receptor (GITR), αEβ7 integrin, and Foxp3. Adoptive transfer of Tgf-β1-/- splenocytes to Tgf-β1+/+/Rag2-/- mice induced an autoimmune inflammatory disease with features similar to those of the Tgf-β1-/- phenotype, and disease transfer was accelerated by the depletion of Tgf-β1-/- CD4+CD25+ T cells from donor splenocytes. Cotransfer of Tgf- β1-/- CD4+CD25+ T cells clearly attenuated disease in Rag2-/- recipients of CD25+-depleted Tgf-β1-/- spleen and lymph node cells, but suppression was incomplete when compared with Tgf-β1+/+ CD4+CD25+ T cells. These data demonstrate that CD4+CD25+ regulatory T cells develop in complete absence of endogenous transforming growth factor-β1 (TGF-β1) expression and that autocrine TGF-β1 expression is not essential for these cells to suppress inflammation in vivo. (Blood. 2004;103:4594-4601)

Leflunomide induces immunosuppression in collagen-induced arthritis rats by upregulating CD4+CD25+ regulatory T cells

2010 ◽  
Vol 88 (1) ◽  
pp. 45-53 ◽  
Author(s):  
Ting-Yu Wang ◽  
Jun Li ◽  
Chang-Yu Li ◽  
Yong Jin ◽  
Xiong-Wen Lü ◽  
...  
Keyword(s):  
T Cells ◽  
T Lymphocytes ◽  
Regulatory T Cells ◽  
Transforming Growth Factor ◽  
Transforming Growth Factor Β1 ◽  
Con A ◽  
Collagen Induced Arthritis ◽  
Foxp3 Mrna ◽  
Tgf Β1

This study was to investigate the effect of leflunomide on the immunosuppressive CD4+CD25+ regulatory T cells (CD4+CD25+ Tregs) in collagen-induced arthritis (CIA) rats. CIA was induced by collagen type II in Wistar rats. Immunofluorescence flow cytometry and RT-PCR were used to determine the proportion of CD4+CD25+ Tregs and the expression of Foxp3 mRNA, respectively. Proliferation of T lymphocytes was assayed with MTT reagent, and the level of transforming growth factor β1 (TGF-β1) in the supernatant of concanavalin A (Con A)-induced T lymphocytes was determined by ELISA kit. Our investigations demonstrated that inhibition of arthritis by leflunomide was related to changes in CD4+CD25+ Tregs. In addition, A771726, which is the active metabolite of leflunomide, promoted the differentiation of spleen lymphocytes into CD4+CD25+ Tregs, increased antiinflammatory cytokine TGF-β1 secretion, and adjusted the activity of Con A-induced lymphocytes in vitro.

Comparison of Release Profiles of Various Growth Factors from Biodegradable Carriers

1998 ◽  
Vol 530 ◽  
Author(s):  
Y. Tabata ◽  
M. Yamamoto ◽  
Y. Ikada
Keyword(s):  
Growth Factors ◽  
Growth Factor ◽  
Transforming Growth Factor ◽  
Transforming Growth Factor Β1 ◽  
The Body ◽  
Bone Morphogenetic Protein 2 ◽  
Biodegradable Hydrogel ◽  
Tgf Β1

AbstractA biodegradable hydrogel was prepared by glutaraldehyde crosslinking of acidic gelatin with an isoelectric point (IEP) of 5.0 as a carrier to release basic growth factors on the basis of polyion complexation. Basic fibroblast growth factor (bFGF), transforming growth factor β1 (TGF-β1), and bone morphogenetic protein-2 (BMP-2) were sorbed from their aqueous solution into the dried gelatin hydrogels to prepare respective growth factor-incorporating hydrogels. Under an in vitro non-degradation condition, approximately 20 % of incorporated bFGF and TGF-β1 was released from the hydrogels within initial 40 min, followed by no further release, whereas a large initial release of BMP-2 was observed. After subcutaneous implantation of the gelatin hydrogels incorporating 125I-labeled growth factor in the mouse back, the remaining radioactivity was measured to estimate the in vivo release profile of growth factors. Incorporation into gelatin hydrogels enabled bFGF and TGF-β1 to retain in the body for about 15 days and the retention period well correlated with that of the gelatin hydrogel. Taken together, it is likely that the growth factors ionically complexed with acidic gelatin were released in vivo as a result of hydrogel biodegradation. On the contrary, basic BMP-2 did not ionically interact with acidic gelatin, resulting in no sustained released by the present biodegradable carrier system.

scholarly journals Targeting Follistatin like 1 ameliorates liver fibrosis induced by carbon tetrachloride through TGF-β1-miR29a in mice

2020 ◽  
Vol 18 (1) ◽  
Author(s):  
Xin-Yi Xu ◽  
Yan Du ◽  
Xue Liu ◽  
Yilin Ren ◽  
Yingying Dong ◽  
...  
Keyword(s):  
Liver Fibrosis ◽  
Cell Activation ◽  
Transforming Growth Factor ◽  
Stellate Cell ◽  
Neutralizing Antibody ◽  
Transforming Growth Factor Β1 ◽  
Chemokine Ligand ◽  
Tgf Β1

Abstract Background Hepatic fibrosis is a pathological response of the liver to a variety of chronic stimuli. Hepatic stellate cells (HSCs) are the major source of myofibroblasts in the liver. Follistatin like 1 (Fstl1) is a secreted glycoprotein induced by transforming growth factor-β1 (TGF-β1). However, the precise functions and regulation mechanisms of Fstl1 in liver fibrogenesis remains unclear. Methods Hepatic stellate cell (HSC) line LX-2 stimulated by TGF-β1, primary culture of mouse HSCs and a model of liver fibrosis induced by CCl4 in mice was used to assess the effect of Fstl1 in vitro and in vivo. Results Here, we found that Fstl1 was significantly up regulated in human and mouse fibrotic livers, as well as activated HSCs. Haplodeficiency of Fstl1 or blockage of Fstl1 with a neutralizing antibody 22B6 attenuated CCl4-induced liver fibrosis in vivo. Fstl1 modulates TGF-β1 classic Samd2 and non-classic JNK signaling pathways. Knockdown of Fstl1 in HSCs significantly ameliorated cell activation, cell migration, chemokines C-C Motif Chemokine Ligand 2 (CCL2) and C-X-C Motif Chemokine Ligand 8 (CXCL8) secretion and extracellular matrix (ECM) production, and also modulated microRNA-29a (miR29a) expression. Furthermore, we identified that Fstl1 was a target gene of miR29a. And TGF-β1 induction of Fstl1 expression was partially through down regulation of miR29a in HSCs. Conclusions Our data suggests TGF-β1-miR29a-Fstl1 regulatory circuit plays a key role in regulation the HSC activation and ECM production, and targeting Fstl1 may be a strategy for the treatment of liver fibrosis. Graphical abstract

scholarly journals Krüppel-like factor KLF10 regulates transforming growth factor receptor II expression and TGF-β signaling in CD8+ T lymphocytes

2015 ◽  
Vol 308 (5) ◽  
pp. C362-C371 ◽  
Author(s):  
Konstantinos A. Papadakis ◽  
James Krempski ◽  
Jesse Reiter ◽  
Phyllis Svingen ◽  
Yuning Xiong ◽  
...  
Keyword(s):  
T Cells ◽  
Growth Factor ◽  
Transforming Growth Factor ◽  
Critical Role ◽  
Effector Memory ◽  
Antiviral Immune Response ◽  
Ifn Γ ◽  
Tgf Β1

KLF10 has recently elicited significant attention as a transcriptional regulator of transforming growth factor-β1 (TGF-β1) signaling in CD4+ T cells. In the current study, we demonstrate a novel role for KLF10 in the regulation of TGF-β receptor II (TGF-βRII) expression with functional relevance in antiviral immune response. Specifically, we show that KLF10-deficient mice have an increased number of effector/memory CD8+ T cells, display higher levels of the T helper type 1 cell-associated transcription factor T-bet, and produce more IFN-γ following in vitro stimulation. In addition, KLF10−/− CD8+ T cells show enhanced proliferation in vitro and homeostatic proliferation in vivo. Freshly isolated CD8+ T cells from the spleen of adult mice express lower levels of surface TGF-βRII (TβRII). Congruently, in vitro activation of KLF10-deficient CD8+ T cells upregulate TGF-βRII to a lesser extent compared with wild-type (WT) CD8+ T cells, which results in attenuated Smad2 phosphorylation following TGF-β1 stimulation compared with WT CD8+ T cells. Moreover, we demonstrate that KLF10 directly binds to the TGF-βRII promoter in T cells, leading to enhanced gene expression. In vivo viral infection with Daniel's strain Theiler's murine encephalomyelitis virus (TMEV) also led to lower expression of TGF-βRII among viral-specific KLF10−/− CD8+ T cells and a higher percentage of IFN-γ-producing CD8+ T cells in the spleen. Collectively, our data reveal a critical role for KLF10 in the transcriptional activation of TGF-βRII in CD8+ T cells. Thus, KLF10 regulation of TGF-βRII in this cell subset may likely play a critical role in viral and tumor immune responses for which the integrity of the TGF-β1/TGF-βRII signaling pathway is crucial.

A major role of TGF-β1 in the homing capacities of murine hematopoietic stem cell/progenitors

Blood ◽  
2010 ◽  
Vol 116 (8) ◽  
pp. 1244-1253 ◽  
Author(s):  
Claude Capron ◽  
Catherine Lacout ◽  
Yann Lécluse ◽  
Valérie Jalbert ◽  
Hédia Chagraoui ◽  
...  
Keyword(s):  
Transforming Growth Factor ◽  
Bone Marrow Cells ◽  
Transforming Growth Factor Β1 ◽  
Inflammatory Disorder ◽  
Hematopoietic Stem ◽  
Murine Hematopoietic Stem Cell ◽  
In Vitro Effects ◽  
Tgf Β1

Abstract Transforming growth factor-β1 (TGF-β1) is a pleiotropic cytokine with major in vitro effects on hematopoietic stem cells (HSCs) and lymphocyte development. Little is known about hematopoiesis from mice with constitutive TGF-β1 inactivation largely because of important embryonic lethality and development of a lethal inflammatory disorder in TGF-β1−/− pups, making these studies difficult. Here, we show that no sign of the inflammatory disorder was detectable in 8- to 10-day-old TGF-β1−/− neonates as judged by both the number of T-activated and T-regulator cells in secondary lymphoid organs and the level of inflammatory cytokines in sera. After T-cell depletion, the inflammatory disease was not transplantable in recipient mice. Bone marrow cells from 8- to 10-day-old TGF-β1−/− neonates showed strikingly impaired short- and long-term reconstitutive activity associated with a parallel decreased in vivo homing capacity of lineage negative (Lin−) cells. In addition an in vitro–reduced survival of immature progenitors (Lin− Kit+ Sca+) was observed. Similar defects were found in liver cells from TGF-β1−/− embryos on day 14 after vaginal plug. These data indicate that TGF-β1 is a critical regulator for in vivo homeostasis of the HSCs, especially for their homing potential.

scholarly journals Pyrrole-Imidazole Polyamide Targeting Transforming Growth Factor β1 Ameliorates Encapsulating Peritoneal Sclerosis

2012 ◽  
Vol 32 (4) ◽  
pp. 462-472 ◽  
Author(s):  
Kazuo Serie ◽  
Noboru Fukuda ◽  
Shigeki Nakai ◽  
Hiroyuki Matsuda ◽  
Takashi Maruyama ◽  
...  
Keyword(s):  
Growth Factor ◽  
Mrna Expression ◽  
Messenger Rna ◽  
Transforming Growth Factor ◽  
Fluorescein Isothiocyanate ◽  
Transforming Growth Factor Β1 ◽  
Vegf Mrna ◽  
Tgf Β1

ObjectiveEncapsulating peritoneal sclerosis (EPS) is a devastating fibrotic complication in patients treated with peritoneal dialysis (PD). Transforming growth factor β1 (TGF-β1) is a pivotal factor in the induction of EPS.MethodsTo develop pyrrole-imidazole (PI) polyamide, a novel gene silencer, targeted to the TGF-β1 promoter (Polyamide) for EPS, we examined the effects of Polyamide on messenger RNA (mRNA) expression of TGF-β 1, vascular endothelial growth factor (VEGF), and extracellular matrix (ECM) in mesothelial cells in vitro, and on the thickness of injured peritoneum evaluated by histology and high- resolution regional elasticity mapping in rats in vivo.ResultsPolyamide significantly lowered mRNA expression of TGF-β 1 and ECM in vitro. Polyamide labeled with fluorescein isothiocyanate was taken up into the injured peritoneum and was strongly localized in the nuclei of most cells. Polyamide 1 mg was injected intraperitoneally 1 or 3 times in rats receiving a daily intraperitoneal injection of chlorhexidine gluconate and ethanol (CHX) for 14 days. Polyamide significantly suppressed peritoneal thickening and the abundance of TGF-β 1 and fibronectin mRNA, but did not affect expression of VEGF mRNA in the injured peritoneum. Elasticity distribution mapping showed that average elasticity was significantly lower in Polyamide-treated rats than in rats treated solely with CHX.ConclusionsPolyamide suppressed the stiffness, ECM formation, and thickening of the injured peritoneum that occurs during EPS pathogenesis. These data suggest that PI polyamide targeted to the TGF-β 1 promoter will be a specific and feasible therapeutic strategy for patients with EPS.

Transforming Growth Factor-β1 (TGF-β1) Induces Thrombopoietin From Bone Marrow Stromal Cells, Which Stimulates the Expression of TGF-β Receptor on Megakaryocytes and, in Turn, Renders Them Susceptible to Suppression by TGF-β Itself With High Specificity

Blood ◽  
1999 ◽  
Vol 94 (6) ◽  
pp. 1961-1970 ◽  
Author(s):  
Sumio Sakamaki ◽  
Yasuo Hirayama ◽  
Takuya Matsunaga ◽  
Hiroyuki Kuroda ◽  
Toshiro Kusakabe ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Growth Factor ◽  
Transforming Growth Factor ◽  
High Specificity ◽  
Transforming Growth Factor Β1 ◽  
Lineage Specificity ◽  
Β Receptor ◽  
Tgf Β1

Abstract The present study was designed to test the concept that platelets release a humoral factor that plays a regulatory role in megakaryopoiesis. The results showed that, among various hematoregulatory cytokines examined, transforming growth factor-β1 (TGF-β1) was by far the most potent enhancer of mRNA expression of bone marrow stromal thrombopoietin (TPO), a commitment of lineage specificity. The TPO, in turn, induced TGF-β receptors I and II on megakaryoblasts at the midmegakaryopoietic stage; at this stage, TGF-β1 was able to arrest the maturation of megakaryocyte colony-forming units (CFU-Meg). This effect was relatively specific when compared with its effect on burst-forming unit-erythroid (BFU-E) or colony-forming unit–granulocyte-macrophage (CFU-GM). In patients with idiopathic thrombocytopenic purpura (ITP), the levels of both TGF-β1 and stromal TPO mRNA were correlatively increased and an arrest of megakaryocyte maturation was observed. These in vivo findings are in accord with the aforementioned in vitro results. Thus, the results of the present investigation suggest that TGF-β1 is one of the pathophysiological feedback regulators of megakaryopoiesis.

scholarly journals Fedratinib Attenuates Bleomycin-Induced Pulmonary Fibrosis via the JAK2/STAT3 and TGF-β1 Signaling Pathway

Molecules ◽  
2021 ◽  
Vol 26 (15) ◽  
pp. 4491
Author(s):  
Hao Ruan ◽  
Jiaoyan Luan ◽  
Shaoyan Gao ◽  
Shuangling Li ◽  
Qiuyan Jiang ◽  
...  
Keyword(s):  
Pulmonary Fibrosis ◽  
Signaling Pathway ◽  
Kinase Inhibitors ◽  
Transforming Growth Factor ◽  
Transforming Growth Factor Β1 ◽  
In Vivo Studies ◽  
Matrix Deposition ◽  
Tgf Β1

Idiopathic pulmonary fibrosis (IPF) is a chronic, progressive interstitial lung disease with multiple causes, characterized by excessive myofibrocyte aggregation and extracellular matrix deposition. Related studies have shown that transforming growth factor-β1 (TGF-β1) is a key cytokine causing fibrosis, promoting abnormal epithelial–mesenchymal communication and fibroblast-to-myofibroblast transition. Fedratinib (Fed) is a marketed drug for the treatment of primary and secondary myelofibrosis, targeting selective JAK2 tyrosine kinase inhibitors. However, its role in pulmonary fibrosis remains unclear. In this study, we investigated the potential effects and mechanisms of Fed on pulmonary fibrosis in vitro and in vivo. In vitro studies have shown that Fed attenuates TGF-β1- and IL-6-induced myofibroblast activation and inflammatory response by regulating the JAK2/STAT3 signaling pathway. In vivo studies have shown that Fed can reduce bleomycin-induced inflammation and collagen deposition and improve lung function. In conclusion, Fed inhibited inflammation and fibrosis processes induced by TGF-β1 and IL-6 by targeting the JAK2 receptor.

Sign in / Sign up

Export Citation Format

Share Document