Growth Factor-beta Isoforms Research Articles

TGF-β Isoforms and GDF-15 in the Development and Progression of Atherosclerosis.

The effect of oxidised lipoproteins on the endothelium, monocytes, platelets, and macrophages is a key factor in the initiation and development of atherosclerosis. Antioxidant action, lipoprotein metabolism, and chronic inflammation are the fields of research interest for better understanding the development of the disease. All the fields are related to inflammation and hence to the secretion of cytokines, which are being investigated as potential diagnostic markers for the onset of atherosclerosis. Pathways of vascular damage are crucial for the development of new laboratory readouts. The very early detection of endothelial cell damage associated with the onset of atherosclerosis, allowing the initiation of therapy, remains a major research goal. This article summarises the latest results on the relationship of tumour growth factor beta (TGF-β) isoforms and growth differentiation factor 15 (GDF-15) to the pathogenesis of atherosclerosis: which cells involved in atherosclerosis produce them, which effectors stimulate their synthesis and secretion, how they influence atherosclerosis development, and the relationship between the levels of TGF-β and GDF-15 in the blood and the development and extent of atherosclerosis.

Response of TGF-β isoforms in epithelial-mesenchymal transition of enamel epithelial cells

ObjectiveDuring enamel formation, transforming growth factor-beta (TGF-β) isoforms exhibit different activities for gene expression, apoptosis, and endocytosis. This study aimed to investigate the differential response of TGF-β isoforms to epithelial-mesenchymal transition (EMT) in enamel epithelial cells. DesignUsing a mouse enamel epithelial cell line (mHAT9d) cultured in the presence of each TGF-β isoform, (1) the morphological changes in EMT were explored, (2) EMT-related genes were analyzed by next-generation sequencing (NGS), (3) TGF-β pathway for EMT was identified by inhibition experiments, and (4) the expression of the TGF-β receptor gene in response to the binding affinity of the TGF-β isoform were analyzed. ResultsEMT was observed in mHAT9d cultured in the presence of TGF-β1 and β3 but not TGF-β2. The expression of both epithelial and mesenchymal marker genes was observed in mHAT9d exhibiting EMT. NGS analysis suggested extracellular signal-regulated kinase (ERK) and Rho pathways as TGF-β signaling pathways associated with EMT. However, EMT in mHAT9d cultured in the presence of TGF-β1 or β3 occurred even in presence of an ERK1/2 inhibitor and was suppressed by Rho-kinase inhibitor. The expression of co-receptors for TGF-β signaling in mHAT9d cells reduced following stimulation with each TGF-β isoform. In contrast, endoglin levels increased following TGF-β1 or β3 stimulation, but no change was noted in response to TGF-β2. ConclusionsWe propose that in TGF-β-stimulated enamel epithelial cells, EMT mainly occurred via the Rho signaling pathway, and the differences in response across TGF-β isoforms were due to their endoglin-mediated binding affinity for the TGF-β receptor.

Transforming growth factor beta isoforms and TGF-βR1 and TGF-βR2 expression in systemic sclerosis patients.

Systemic sclerosis (SSc) is characterized by chronic inflammation and fibrosis, two processes associated with transforming growth factor β (TGF-β) functions. In the present study, we investigated the expression of TGF-β isoforms in serum and the skin distribution of TGF-β and two receptors (TGF-βR1 and TGF-βR2) and their relationship with some clinical, inflammatory, autoimmune (autoantibodies), and vascular (platelets) biomarkers in SSc patients. A total of 56 SSc patients and 120 control subjects (CS) were included. The serum levels of TGF-β isoforms were quantified by immunoassay with magnetic microspheres, and the skin biopsies were processed by immunohistochemistry. The soluble levels of the three active TGF-β isoforms were lower in SSc patients than in CS (p < 0.0001). However, sTGF-β1 and sTGF-β3 levels were positively correlated with C-reactive protein levels in SSc patients. Additionally, sTGF-β2 and sTGF-β3 levels were positively correlated with the number of platelets in SSc patients. In skin biopsies, TGF-β1, TGF-βR1, and TGF-βR2 expression levels were higher in SSc patients than CS. In conclusion, this is the first study showing a joint decrease of the 3 active TGF-β isoforms in SSc patients. However, TGF-β1, TGF-βR1, and TGF-βR2 are possibly increased in clinically involved skin. Therefore, it is likely that a distinct role is played by TGF-β at the local (skin lesions) and systemic levels in SSc patients.

TGF-β1 Production By Bone Marrow Mesenchymal Stem Cells of FANCD2 Differs from Other Fanconi Anemia Complementation Groups

TGF-β secretion from cells in the bone marrow (BM) niche affects hematopoietic stem cell (HSC) fate and has a cardinal role in HSC quiescence. As a component of the BM niche, BM- mesenchymal stem cells (BM-MSCs) may produce abnormal levels of TGF-β in Fanconi anemia (FA) as obtained for HSCs previously and may play a role in bone marrow failure.Herein, we present the molecular and cellular characterization of FA BM-MSCs by addressing their immunophenotypes, proliferation- and differentiation capacities, as well as expression and secretion levels of transforming growth factor beta (TGF-β) isoforms. Proliferative potency of FA cells was determined between P3 to P7 and compared with healthy donors by calculating cumulative population doubling (cPD) at each passage. Senescence of FA patient BM-MSCs was evaluated at P3 and P6 by SA-β-gal activity and compared with donor BM-MSCs. Diepoxybutane (DEB) effect on TGF-β1 expression of BM-MSCs from FA patients and donors was also assessed.In ten FA patients, mutations were classified as FANCA (n=7), FANCG (n=1) and FANCD2 (n=2). Of six were novel mutations, five in FANCA and one in FANCD2 . BM-MSCs immunophenotype and differentiation capacity of FA BM-MSCs were similar to healthy donors.The CD200 (OX-2) levels were slightly lower in FA MSCs compared to donors. The percentage of CD106 (VCAM-1) positive cells were slightly high especially in FANCD2 cells. FA BM-MSCs showed arrest in dip G2 following DEB treatment. BM-MSCs derived from FANCA- or FANCG -deficient patients displayed similar ex vivo proliferative capacity as healthy donor cells from P3 to P7. However, FANCD2-deficient cells presented lower proliferative capacity. FANCA BM-MSCs had notably increased β-gal positivity at P6, while cells displaying FANCD2 mutation had higher activity at P3 indicating premature aging of these cells. Relative gene expression of TGF-β1 was similar between donor and FA MSCs and did not change by DEB treatment. All cells did not secrete TGF-β2 and -β3. Since the variation of TGF-β1 secretion by FANCA and donor cells was high, a significant difference between the groups could not be obtained. On the other hand, FANCD2 -deficient cells were lack of TGF-β1 secretion. Our data suggest that absence of TGF-β1 secretion may lead to early onset of senescence of the FA-D2 BM-MSCs and may be of value to understand the Fanconi anaemia bone marrow dysfunction in humans. DisclosuresNo relevant conflicts of interest to declare.

Expression of Transforming Growth Factor Beta Isoforms in Canine Endometrium with Cystic Endometrial Hyperplasia-Pyometra Complex.

Simple SummaryPathomorphological changes and functional disorders of the uterus have long been a significant problem in the reproduction of dogs. The most commonly identified uterine disorders leading to permanent loss of fertility in dogs include cystic endometrial hyperplasia (CEH) and pyometra. These diseases may occur jointly as a CEH–pyometra complex. Despite numerous studies, the etiology of this disease remains unclear. TGF-β is considered to be one of the key factors in pathophysiological uterine disorders. The results indicate the significant expression of TGF-β1 in endometrial tissues in bitches affected by CEH–pyometra complex. Consequently, among all TGF-β isoforms, TGF-β1 is a potential biomarker involved in the regulation of a dog’s endometrium with proliferative and degenerative changes.Cystic endometrial hyperplasia (CEH) and pyometra are the most frequently diagnosed uterine diseases affecting bitches of different ages. Transforming growth factor beta (TGF-β) has been classified in females as a potential regulator of many endometrial changes during the estrous cycle or may be involved in pathological disorders. The aim of this study was to determine the expression of TGF-β1, -β2 and -β3 in the endometrium of bitches suffering from CEH or a CEH–pyometra complex compared to clinically healthy females (control group; CG). A significantly increased level of TGF-β1 mRNA expression was observed in the endometrium with CEH–pyometra compared to CEH and CG. Protein production of TGF-β1 was identified only in the endometrium of bitches with CEH–pyometra. An increase in TGF-β3 mRNA expression was observed in all the studied groups compared to CG. The expression of TGF-β2 mRNA was significantly higher in CEH and lower in CEH–pyometra uteri. The results indicate the presence of TGF-β cytokines in canine endometrial tissues affected by proliferative and degenerative changes. However, among all TGF-β isoforms, TGF-β1 could potentially be a key factor involved in the regulation of the endometrium in bitches with CEH–pyometra complex.

Human Cord Blood Derived Unrestricted Somatic Stem Cells Restore Aquaporin Channel Expression, Reduce Inflammation and Inhibit the Development of Hydrocephalus After Experimentally Induced Perinatal Intraventricular Hemorrhage.

Intraventricular hemorrhage (IVH) is a severe complication of preterm birth associated with cerebral palsy, intellectual disability, and commonly, accumulation of cerebrospinal fluid (CSF). Histologically, IVH leads to subependymal gliosis, fibrosis, and disruption of the ependymal wall. Importantly, expression of aquaporin channels 1 and 4 (AQP1 and AQP4) regulating respectively, secretion and absorption of cerebrospinal fluids is altered with IVH and are associated with development of post hemorrhagic hydrocephalus. Human cord blood derived unrestricted somatic stem cells (USSCs), which we previously demonstrated to reduce the magnitude of hydrocephalus, as having anti-inflammatory, and beneficial behavioral effects, were injected into the cerebral ventricles of rabbit pups 18 h after glycerol-induced IVH. USSC treated IVH pups showed a reduction in ventricular size when compared to control pups at 7 and 14 days (both, P < 0.05). Histologically, USSC treatment reduced cellular infiltration and ependymal wall disruption. In the region of the choroid plexus, immuno-reactivity for AQP1 and ependymal wall AQP4 expression were suppressed after IVH but were restored following USSC administration. Effects were confirmed by analysis of mRNA from dissected choroid plexus and ependymal tissue. Transforming growth factor beta (TGF-β) isoforms, connective tissue growth factor (CTGF) and matrix metalloprotease-9 (MMP-9) mRNA, as well as protein levels, were significantly increased following IVH and restored towards normal with USSC treatment (P < 0.05). The anti-inflammatory cytokine Interleukin-10 (IL-10) mRNA was reduced in IVH, but significantly recovered after USSC injection (P < 0.05). In conclusion, USSCs exerted anti-inflammatory effects by suppressing both TGF-β specific isoforms, CTGF and MMP-9, recovered IL-10, restored aquaporins expression towards baseline, and reduced hydrocephalus. These results support the possibility of the use of USSCs to reduce IVH consequences in prematurity.

IL11 is elevated in systemic sclerosis and IL11-dependent ERK signalling underlies TGFβ-mediated activation of dermal fibroblasts.

ObjectivesInterleukin 11 (IL11) is highly upregulated in skin and lung fibroblasts from patients with systemic sclerosis (SSc). Here we tested whether IL11 is mechanistically linked with activation of human dermal fibroblasts (HDFs) from patients with SSc or controls.MethodsWe measured serum IL11 levels in volunteers and patients with early diffuse SSc and manipulated IL11 signalling in HDFs using gain- and loss-of-function approaches that we combined with molecular and cellular phenotyping.ResultsIn patients with SSc, serum IL11 levels are elevated as compared with healthy controls. All transforming growth factor beta (TGFβ) isoforms induced IL11 secretion from HDFs, which highly express IL11 receptor α-subunit and the glycoprotein 130 (gp130) co-receptor, suggestive of an autocrine loop of IL11 activity in HDFs. IL11 stimulated ERK activation in HDFs and resulted in HDF-to-myofibroblast transformation and extracellular matrix secretion. The pro-fibrotic action of IL11 in HDFs appeared unrelated to STAT3 activity, independent of TGFβ upregulation and was not associated with phosphorylation of SMAD2/3. Inhibition of IL11 signalling using either a neutralizing antibody against IL11 or siRNA against IL11RA reduced TGFβ-induced HDF proliferation, matrix production and cell migration, which was phenocopied by pharmacological inhibition of ERK.ConclusionsThese data reveal that autocrine IL11-dependent ERK activity alone or downstream of TGFβ stimulation promotes fibrosis phenotypes in dermal fibroblasts and suggest IL11 as a potential therapeutic target in SSc.

Bone Morphogenetic Protein-9 Is a Potent Chondrogenic and Morphogenic Factor for Articular Cartilage Chondroprogenitors.

Articular cartilage contains a subpopulation of tissue-specific progenitors that are an ideal cell type for cell therapies and generating neocartilage for tissue engineering applications. However, it is unclear whether the standard chondrogenic medium using transforming growth factor beta (TGFβ) isoforms is optimal to differentiate these cells. We therefore used pellet culture to screen progenitors from immature bovine articular cartilage with a number of chondrogenic factors and discovered that bone morphogenetic protein-9 (BMP9) precociously induces their differentiation. This difference was apparent with toluidine blue staining and confirmed by biochemical and transcriptional analyses with BMP9-treated progenitors exhibiting 11-fold and 5-fold greater aggrecan and collagen type II (COL2A1) gene expression than TGFβ1-treated progenitors. Quantitative gene expression analysis over 14 days highlighted the rapid and phased nature of BMP9-induced chondrogenesis with sequential activation of aggrecan then collagen type II, and negligible collagen type X gene expression. The extracellular matrix of TGFβ1-treated progenitors analyzed using atomic force microscopy was fibrillar and stiff whist BMP9-induced matrix of cells more compliant and correspondingly less fibrillar. Polarized light microscopy revealed an annular pattern of collagen fibril deposition typified by TGFβ1-treated pellets, whereas BMP9-treated pellets displayed a birefringence pattern that was more anisotropic. Remarkably, differentiated immature chondrocytes incubated as high-density cultures in vitro with BMP9 generated a pronounced anisotropic organization of collagen fibrils indistinguishable from mature adult articular cartilage, with cells in deeper zones arranged in columnar manner. This contrasted with cells grown with TGFβ1, where a concentric pattern of collagen fibrils was visualized within tissue pellets. In summary, BMP9 is a potent chondrogenic factor for articular cartilage progenitors and is also capable of inducing morphogenesis of adult-like cartilage, a highly desirable attribute for in vitro tissue-engineered cartilage.

Orf Virus IL-10 and VEGF-E Act Synergistically to Enhance Healing of Cutaneous Wounds in Mice.

Orf virus (OV) is a zoonotic parapoxvirus that causes highly proliferative skin lesions which resolve with minimal inflammation and scarring. OV encodes two immunomodulators, vascular endothelial growth factor (VEGF)-E and interleukin-10 (ovIL-10), which individually modulate skin repair and inflammation. This study examined the effects of the VEGF-E and ovIL-10 combination on healing processes in a murine wound model. Treatments with viral proteins, individually and in combination, were compared to a mammalian VEGF-A and IL-10 combination. Wound biopsies were harvested to measure re-epithelialisation and scarring (histology), inflammation, fibrosis and angiogenesis (immunofluorescence), and gene expression (quantitative polymerase chain reaction). VEGF-E and ovIL-10 showed additive effects on wound closure and re-epithelialisation, and suppressed M1 macrophage and myofibroblast infiltration, while allowing M2 macrophage recruitment. The viral combination also increased endothelial cell density and pericyte coverage, and improved collagen deposition while reducing the scar area. The mammalian combination showed equivalent effects on wound closure, re-epithelialisation and fibrosis, but did not promote blood vessel stabilisation or collagen remodeling. The combination treatments also differentially altered the expression of transforming growth factor beta isoforms, Tgfβ1 and Tgfβ3. These findings show that the OV proteins synergistically enhance skin repair, and act in a complimentary fashion to improve scar quality.

Diverse Signaling by TGF\u03b2 Isoforms in Response to Focal Injury is Associated with Either Retinal Regeneration or Reactive Gliosis

Müller cells may have stem cell-like capability as they regenerate photoreceptor loss upon injury in some vertebrates, but not in mammals. Indeed, mammalian Müller cells undergo major cellular and molecular changes summarized as reactive gliosis. Transforming growth factor beta (TGFβ) isoforms are multifunctional cytokines that play a central role, both in wound healing and in tissue repair. Here, we studied the role of TGFβ isoforms and their signaling pathways in response to injury induction during tissue regeneration in zebrafish and scar formation in mouse. Our transcriptome analysis showed a different activation of canonical and non-canonical signaling pathways and how they shaped the injury response. In particular, TGFβ3 promotes retinal regeneration via Smad-dependent canonical pathway upon regulation of junb gene family and mycb in zebrafish Müller cells. However, in mice, TGFβ1 and TGFβ2 evoke the p38MAPK signaling pathway. The activation of this non-canonical pathway leads to retinal gliosis. Thus, the regenerative versus reparative effect of the TGFβ pathway observed may rely on the activation of different signaling cascades. This provides one explanation of the different injury response in zebrafish and mouse retina.

Role of Transforming Growth Factor-β in Skeletal Muscle Fibrosis: A Review.

Transforming growth factor-beta (TGF-β) isoforms are cytokines involved in a variety of cellular processes, including myofiber repair and regulation of connective tissue formation. Activation of the TGF-β pathway contributes to pathologic fibrosis in most organs. Here, we have focused on examining the evidence demonstrating the involvement of TGF-β in the fibrosis of skeletal muscle particularly. The TGF-β pathway plays a role in different skeletal muscle myopathies, and TGF-β signaling is highly induced in these diseases. In this review, we discuss different molecular mechanisms of TGF-β-mediated skeletal muscle fibrosis and highlight different TGF-β-targeted treatments that target these relevant pathways.

Potential function of TGF-β isoforms in maturation-stage ameloblasts

ObjectivesTo investigate potential functions of transforming growth factor-beta (TGF-β) isoforms in maturation-stage ameloblasts during amelogenesis. MethodsIn vivo activation of TGF-β was characterized by using matrix metalloproteinase 20 null (Mmp20-/-) and wild-type (Mmp20+/+) mice. Using mHAT9d cells cultured in the presence of each TGF-β isoform, (1) cell proliferation was determined by MTS assay, (2) immunostaining with anti-cleaved caspase-3 monoclonal antibody was performed and apoptotic indices were measured, (3) gene expression was analyzed by RT-qPCR, and (4) the uptake of amelogenin into mHAT9d cells was directly observed using a fluorescence microscope. ResultsTGF-β1 and TGF-β3 were present in the enamel matrix of developing teeth which were activated by MMP20 in vivo. A genetic study revealed that the three TGF-β isoforms upregulate kallikrein 4 (KLK4) mRNA levels but downregulate carbonic anhydrase II. Moreover, TGF-β1 and TGF-β2 significantly upregulated the mRNA level of amelotin, whereas TGF-β3 dramatically downregulated the mRNA levels of odontogenic ameloblast-associated protein (ODAM), family with sequence similarity 83 member H (FAM83H), and alkaline phosphatase (ALP). Immunostaining analysis showed that the apoptosis of mHAT9d cells is induced by three TGF-β isoforms, with TGF-β3 being most effective. Both TGF-β1 and TGF-β3 induced endocytosis of amelogenin. ConclusionsWe propose that TGF-β is regulated in an isoform-specific manner to perform multiple biological functions such as gene expression related to the structure of basal lamina/ameloblasts, mineral ion transport, apoptosis, and endocytosis in maturation-stage ameloblasts.

Inhibition of Human Corneal Myofibroblast Formation.

PurposeTransforming growth factor-beta (TGF-β) isoform 1 (T1) is involved in corneal fibrotic wound healing by stimulating myofibroblast transformation and altering fibrotic gene expression. In this study, two specific inhibitors were used to dissect the relationship between myofibroblast generation and the TGF-β/Smad- or TGF-β/p38-signaling pathway in human corneal fibroblasts (HCF).MethodsIn HCF, Trx-SARA (Smad-pathway inhibitor) was used to block the TGF-β/Smad-signaling pathway, and the p38 inhibitor (p38inh, SB202190) was used to inhibit p38MAPK, thus blocking the TGF-β/p38-signaling pathway. HCF ± Trx-SARA or Trx-GA (SARA control) were serum starved overnight in Eagle's minimum essential medium (EMEM) ± p38inh, grown in EMEM ± T1 ± p38inh for 24 hours, and then processed for indirect-immunofluorescence, Western blot, or quantitative real-time polymerase chain reaction to examine α-smooth muscle actin (αSMA) and other fibrotic genes, such as fibronectin, thrombospondin1, and type III collagen. In addition, the morphology and the effect of p38inh on myofibroblast phenotype after myofibroblast formation were examined.ResultsWe observed that Trx-SARA had little effect on αSMA expression, indicating that blocking the Smad pathway did not significantly inhibit myofibroblast formation. However, p38inh did significantly inhibit αSMA and other fibrotic genes, thus efficiently preventing the transition of HCFs to myofibroblasts. In addition, morphology changed and αSMA decreased in myofibroblasts exposed to p38inh medium, as compared with controls.ConclusionsHCF transition to myofibroblasts was mainly through the p38 pathway. Therefore, blocking the p38 pathway may be a potential therapeutic tool for human corneal fibrosis prevention/treatment, because it controls myofibroblast formation in human corneal cells, while leaving other functions of T1 unaffected.

Differential Expression and Clinical Significance of Transforming Growth Factor-Beta Isoforms in GBM Tumors.

Glioblastoma (GBM) represents the most common and aggressive malignant primary brain tumors in adults. Response to standard treatment is transitory and the survival of clinical trial cohorts are little more than 14 months. GBM are characterized by excessive proliferation, invasiveness, and radio-/chemoresistance features; which are strongly upregulated by transforming growth factor-beta (TGF-β). We hypothesized that TGF-β gene expression could correlate with overall survival (OS) and serve as a prognostic biomarker. TGF-β1 and -β2 expression were analyzed by qPCR in 159 GBM tumor specimens. Kaplan–Meier and multivariate analyses were used to correlate expression with OS and progression-free survival (PFS). In GBM, TGF-β1 and -β2 levels were 33- and 11-fold higher respectively than in non-tumoral samples. Kaplan–Meier and multivariate analyses revealed that high to moderate expressions of TGF-β1 significantly conferred a strikingly poorer OS and PFS in newly diagnosed patients. Interestingly, at relapse, neither isoforms had meaningful impact on clinical evolution. We demonstrate that TGF-β1 is the dominant isoform in newly diagnosed GBM rather than the previously acknowledged TGF-β2. We believe our study is the first to unveil a significant relationship between TGF-β1 expression and OS or PFS in newly diagnosed GBM. TGF-β1 could serve as a prognostic biomarker or target affecting treatment planning and patient follow-up.

Quantitative analysis of transforming growth factor beta isoforms mRNA TGF-β1-3 in the patients with psoriasis

Quantitative analysis of transforming growth factor beta isoforms mRNA TGF-β1-3 in the patients with psoriasis

Transforming Growth Factor Beta is regulated by a Glucocorticoid-Dependent Mechanism in Denervation Mouse Bone

Bone growth and remodeling is inhibited by denervation in adults and children, resulting in alterations of linear growth and bone mass and increased risk for osteoporosis and pathologic fractures. Transforming growth factor beta (TGF-β) isoforms are a key group of growth factors that enhance bone formation. To explore the relation between denervation-induced reduction of bone formation and TGF-β gene expression, we measured mRNA levels of TGF-β in denervation mouse bone and found decreased mRNA levels of TGF-β1, TGF-β2 and TGF-β3. These changes were accompanied by diminishing weight loss, bone mineral density (BMD), trabecular thickness, trabecular separation and trabecular number of femur and lumbar, serum osteocalcin, total calcium, intact parathyroid hormone, and increased serum C telopeptide. Recombinant human TGF-β1 (rhTGF-β1) prevented denervation-induced reduction of BMD further supporting our hypothesis that denervation-induced reduction of bone formation is a result of inhibition of TGF-β gene expression. In addition, antiprogestins RU 38486 blunted the denervation-induced decrease in mRNA levels of TGF-β group, while dexamethasone (DEX) decreased TGF-β group mRNA levels in normal mice. Furthermore, the denervated-mice exhibited a threefold increase in plasma corticosterone. These results suggest that denervation-induced reduction of bone formation may be regulated by glucocorticoids via inhibition of TGF-β gene expression at least in part.

Cardiosphere-Derived Cells Require Endoglin for Paracrine-Mediated Angiogenesis.

SummaryClinical trials of stem cell therapy to treat ischemic heart disease primarily use heterogeneous stem cell populations. Small benefits occur via paracrine mechanisms that include stimulating angiogenesis, and increased understanding of these mechanisms would help to improve patient outcomes. Cardiosphere-derived-cells (CDCs) are an example of these heterogeneous stem cell populations, cultured from cardiac tissue. CDCs express endoglin, a co-receptor that binds specific transforming growth factor β (TGFβ) family ligands, including bone morphogenetic protein 9 (BMP9). In endothelial cells endoglin regulates angiogenic responses, and we therefore hypothesized that endoglin is required to promote the paracrine pro-angiogenic properties of CDCs. Cre/LoxP technology was used to genetically manipulate endoglin expression in CDCs, and we found that the pro-angiogenic properties of the CDC secretome are endoglin dependent both in vitro and in vivo. Importantly, BMP9 pre-treatment of endoglin-depleted CDCs restores their pro-angiogenic paracrine properties. As BMP9 signaling is normally required to maintain endoglin expression, we propose that media containing BMP9 could be critical for therapeutic CDC preparation.

Uterine Wound Healing: A Complex Process Mediated by Proteins and Peptides.

Wound healing is the process by which a complex cascade of biochemical events is responsible of the repair the damage. In vivo, studies in humans and mice suggest that healing and post-healing heterogeneous behavior of the surgically wounded myometrium is both phenotype and genotype dependent. Uterine wound healing process involves many cells: endothelial cells, neutrophils, monocytes/macrophages, lymphocytes, fibroblasts, myometrial cells as well a stem cell population found in the myometrium, myoSP (side population of myometrial cells). Transforming growth factor beta (TGF-β) isoforms, connective tissue growth factor (CTGF), basic fibroblast growth factor (bFGF), platelet-derived growth factor (PDGF), vascular endothelial growth factor (VEGF), and tumor necrosis factor alpha (TNF-β) are involved in the wound healing mechanisms. The increased TGF- β1/β3 ratio reduces scarring and fibrosis. The CTGF altered expression may be a factor involved in the abnormal scars formation of low uterine segment after cesarean section and of the formation of uterine dehiscence. The lack of bFGF is involved in the reduction of collagen deposition in the wound site and thicker scabs. The altered expression of TNF-β, VEGF, and PDGF in human myometrial smooth muscle cells in case of uterine dehiscence, it is implicated in the uterine healing process. The over-and under-expressions of growth factors genes involved in uterine scarring process could represent patient's specific features, increasing the risk of cesarean scar complications.

Abstract 3741: Antimetastatic activity of ISTH0047, a potent and selective TGF-beta 2 antisense oligonucleotide, in syngeneic lung metastatic model of mouse 4T1 mammary carcinoma

Abstract Transforming growth factor beta (TGF-β) isoforms are the primary mediators for TGF-β signaling via TGF-β receptors and downstream phosphorylation/dephosphorylation cascade. TGF-β is associated with a wide range of biological processes in oncology, including tumor cell invasion, migration, angiogenesis, immunosuppression, as well as regulation of tumor stem cell properties. Mouse 4T1 mammary carcinoma cell line is a transplantable tumor cell line that is highly tumorigenic and invasive and, unlike most tumor models, can spontaneously metastasize from the primary tumor in the mammary gland to multiple distant sites including lymph nodes, blood, liver, lung, brain, and bone. Considering rather challenging preclinical evaluation of antitumor activity in tumor models, mouse 4T1 mammary carcinoma model has been widely used in the literature for evaluation of TGF-β antagonists. In this report, we describe the efficacy of ISTH0047 - a potent and selective TGF-β2 antisense oligonucleotide - in murine 4T1 primary tumors and lung metastasis following tumor cell injection into the mammary fat pad (orthotopic tumor model) of syngeneic Balb/c mice. Consistent with literature data generated with other classes of TGF-β antagonists (e.g., small-molecule kinase inhibitors, antibodies or TGF-β trap agents), although limited antitumor activity was demonstrated on primary tumor growth, marked and statistically significant decrease of lung metastasis number was observed upon subcutaneous administrations of ISTH0047. In addition, side by side comparison with murine surrogates of CTLA-4 or PD-1 antibodies indicated similar efficacy of all test items on lung metastasis in this model. Taken together, these encouraging results pave the way for in-depth preclinical evaluation of both ‘seed and soil’ theory and efficacy of combination regimen (immunomodulation) for better clinical outcome. Citation Format: Katja Wosikowski, Kathy Hasenbach, Jutta Petschenka, Diana Barea Roldan, Sebastian Kreiter, Ugur Sahin, Guillaume Serin, Julie-Orlane Redon, Marc Hillairet de Boisferon, Francis Bichat, Hanna Kohonen, Michel Janicot. Antimetastatic activity of ISTH0047, a potent and selective TGF-beta 2 antisense oligonucleotide, in syngeneic lung metastatic model of mouse 4T1 mammary carcinoma. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 3741.

Signalling by Transforming Growth Factor Beta Isoforms in Wound Healing and Tissue Regeneration

Transforming growth factor beta (TGFβ) signalling is essential for wound healing, including both non-specific scar formation and tissue-specific regeneration. Specific TGFβ isoforms and downstream mediators of canonical and non-canonical signalling play different roles in each of these processes. Here we review the role of TGFβ signalling during tissue repair, with a particular focus on the prototypic isoforms TGFβ1, TGFβ2, and TGFβ3. We begin by introducing TGFβ signalling and then discuss the role of these growth factors and their key downstream signalling mediators in determining the balance between scar formation and tissue regeneration. Next we discuss examples of the pleiotropic roles of TGFβ ligands during cutaneous wound healing and blastema-mediated regeneration, and how inhibition of the canonical signalling pathway (using small molecule inhibitors) blocks regeneration. Finally, we review various TGFβ-targeting therapeutic strategies that hold promise for enhancing tissue repair.