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Mesenchymal stromal cells are an important component of the bone marrow hematopoietic niche. Prior studies showed that signaling from members of the transforming growth factor (TGF) superfamily in mesenchymal stromal cells is required for normal niche development. Here, we assessed the impact of TGF family signaling on niche maintenance and stress responses by deleting Smad4 in mesenchymal stromal cells at birth, thereby abrogating canonical TGF signaling. No alteration in the number or spatial organization of CXCL12-abundant reticular (CAR) cells, osteoblasts, or adipocytes was observed in Osx-Cre, Smad4fl/fl mice, and expression of key niche factors was normal. Basal hematopoiesis and stress erythropoiesis responses to acute hemolytic anemia were normal. TGF-β potently inhibits stromal CXCL12 expression in vitro; however, G-CSF induced decreases in bone marrow CXCL12 expression and subsequent hematopoietic stem/progenitor cell mobilization were normal in Osx-Cre, Tgfbr2fl/fl mice, in which all TGF-β signaling in mesenchymal stromal is lost. Finally, although a prior study showed that TGF-β enhances recovery from myeloablative therapy, hematopoietic recovery following single or multiple doses of 5-flurauracil were normal in Osx-Cre, Tgfbr2fl/fl mice. Collectively, these data suggest that TGF family member signaling in mesenchymal stromal cells is dispensable for hematopoietic niche maintenance under basal and stress conditions., Competing Interests: The authors have declared that no competing interests exist.

Cancer associated fibroblasts (CAFs) as the dominant, long-lived and highly plastic cells within the tumor microenvironment (TME) with multi-faceted roles that are endowed with tumor aggressive features. They can instruct and shape the stroma of tumor into being a highly qualified bed for cellular recruitment, differentiation and plasticity in the host tissue or secondary organ/s. In this Review, we have a discussion over CAF reprogramming as a general concept, inducers and outcomes, pursued by suggesting potential strategies to combat this key promoter of tumor., (Copyright © 2019 Elsevier Inc. All rights reserved.)

Background: Chronic diabetic foot wounds are a leading cause of amputation, morbidity, and hospitalization for patients with diabetes. Negative-pressure wound therapy (NPWT) can putatively facilitate wound healing, but the underlying mechanisms remain unclear. Cellular fibronectin (cFN) and transforming growth factor-β1 (TGF-β1) play an important role in wound healing. This prospective randomized controlled trial evaluated the effects of NPWT on the production of cFN and the expression of TGF-β1 in diabetic foot wounds of patients., Methods: From January 2012 to January 2015, 40 patients with diabetic foot wounds were randomly and equally apportioned to receive either NPWT or advanced moist wound therapy (control) for 7 days. Granulation tissue was harvested before and after treatment. Immunohistochemistry and Western blot were performed to evaluate protein levels of cFN and TGF-β1, and real-time polymerase chain reaction (PCR) to measure corresponding mRNA expressions., Results: NPWT facilitated the expression of cFN and TGF-β1 in diabetic foot wounds. Immunohistochemical analysis revealed higher levels of cFN and TGF-β1 in the NPWT group than in the control group. Western blot and real-time PCR analysis further showed that protein and mRNA levels of cFN or TGF-β1 were higher in the NPWT group than that in the control group ( P < .01, both)., Conclusion: Our results showed that NPWT facilitated the production of cFN and the expression of TGF-β1 in granulation tissue in diabetic foot ulcers., Level of Evidence: Level I, randomized controlled study.

Rationale: Recent studies suggest that microRNAs (miRNAs) play important roles in regulation of pulmonary artery smooth muscle cell (PASMC) phenotype and are implicated in pulmonary arterial hypertension (PAH). However, the underlying molecular mechanisms remain elusive., Objectives: This study aims to understand the mechanisms regulating PASMC proliferation and differentiation by microRNA-17∼92 (miR-17∼92) and to elucidate its implication in PAH., Methods: We generated smooth muscle cell (SMC)-specific miR-17∼92 and PDZ and LIM domain 5 (PDLIM5) knockout mice and overexpressed miR-17∼92 and PDLIM5 by injection of miR-17∼92 mimics or PDLIM5-V5-His plasmids and measured their responses to hypoxia. We used miR-17∼92 mimics, inhibitors, overexpression vectors, small interfering RNAs against PDLIM5, Smad, and transforming growth factor (TGF)-β to determine the role of miR-17∼92 and its downstream targets in PASMC proliferation and differentiation., Measurements and Main Results: We found that human PASMC (HPASMC) from patients with PAH expressed decreased levels of the miR-17∼92 cluster, TGF-β, and SMC markers. Overexpression of miR-17∼92 increased and restored the expression of TGF-β3, Smad3, and SMC markers in HPASMC of normal subjects and patients with idiopathic PAH, respectively. Knockdown of Smad3 but not Smad2 prevented miR-17∼92-induced expression of SMC markers. SMC-specific knockout of miR-17∼92 attenuated hypoxia-induced pulmonary hypertension (PH) in mice, whereas reconstitution of miR-17∼92 restored hypoxia-induced PH in these mice. We also found that PDLIM5 is a direct target of miR-17/20a, and hypertensive HPASMC and mouse PASMC expressed elevated PDLIM5 levels. Suppression of PDLIM5 increased expression of SMC markers and enhanced TGF-β/Smad2/3 activity in vitro and enhanced hypoxia-induced PH in vivo, whereas overexpression of PDLIM5 attenuated hypoxia-induced PH., Conclusions: We provided the first evidence that miR-17∼92 inhibits PDLIM5 to induce the TGF-β3/SMAD3 pathway, contributing to the pathogenesis of PAH.

The present study examined whether 20-HETE production is reduced in the renal vasculature and whether this impairs myogenic or tubuloglomerular feedback (TGF) responses of the afferent arteriole (Af-Art). The production of 20-HETE was 73% lower in renal microvessels of Dahl salt-sensitive rats (SS) rats than in SS.5(BN) rats, in which chromosome 5 from the Brown Norway (BN) rat containing the CYP4A genes was transferred into the SS genetic background. The luminal diameter of the Af-Art decreased by 14.7 ± 1.5% in SS.5(BN) rats when the perfusion pressure was increased from 60 to 120 mmHg, but it remained unaltered in SS rats. Administration of an adenosine type 1 receptor agonist (CCPA, 1 μM) reduced the diameter of the Af-Art in the SS.5(BN) rats by 44 ± 2%, whereas the diameter of the Af-Art of SS rats was unaltered. Autoregulation of renal blood flow (RBF) and glomerular capillary pressure (PGC) was significantly impaired in SS rats but was intact in SS.5(BN) rats. Administration of a 20-HETE synthesis inhibitor, HET0016 (1 μM), completely blocked the myogenic and adenosine responses in the Af-Art and autoregulation of RBF and PGC in SS.5(BN) rats, but it had no effect in SS rats. These data indicate that a deficiency in the formation of 20-HETE in renal microvessels impairs the reactivity of the Af-Art of SS rats and likely contributes to the development of hypertension induced renal injury., (Copyright © 2014 the American Physiological Society.)

Uterine fibroids are a prevalent gynaecological condition in reproductive-aged women and are the commonest reason for hysterectomy. The cellular composition of clonal fibroids are heterogeneous, with phenotypically dissimilar cells that include smooth muscle cells (SMC), vascular SMC (VSMC) and fibroblasts. The aim of our study was to investigate genes that are commonly differentially expressed between fibroid and myometrial whole tissues in phenotypically different sub-populations of cells isolated from fibroid and myometrium. Genes to be investigated by fluorescence-activated cell sorting, quantitative real-time PCR and immunocytochemistry include transforming growth factor β (TGFB) and retinoic acid (RA) signalling families and steroid hormone receptors. We hypothesised that each cell population isolated from fibroid and myometrium would differ in the expression of fibroid-associated genes. We demonstrated that phenotypically different cellular constituents of uterine fibroids differentially express cellular RA-binding protein 2 (CRABP2), progesterone receptor B (PRB) and TGFB receptor 2 mRNA in fibroid-derived cells of VSMC and SMC phenotype. CRABP2 mRNA was also differentially expressed in fibroblasts and VSMC sub-populations from within clonal fibroid tumours. We conclude that differential regulation of RA, TGFB and PR pathway transcription occurs in fibroid-associated SMC and -fibroblasts and that investigation of paracrine interactions between different cell types within the fibroid microenvironment provides an important new paradigm for understanding the pathophysiology of this common disease.

New biological activities of bone morphogenetic proteins (BMPs) in the endocrine system have recently been revealed. The BMP system is composed of approximately 30 ligands and preferential combinations of type I and type II receptors. The BMP system not only induces bone formation but also plays unique tissue-specific roles in various organs. For instance, the ovarian BMP system is a physiological inhibitor of luteinization in growing ovarian follicles. In the ovary, the expression of oocyte-derived BMP-15 is critical for female reproduction. In the pituitary, BMP-4 is a key player for initial development of the anterior pituitary, while it is also functionally involved in some differentiated pituitary tumors, including prolactinoma and Cushing's disease. In the adrenal glands, BMP-6 and BMP-4 modulate aldosterone and catecholamine production, respectively, which contributes to a functional interaction between the cortex and medulla. In the present review, recent advances in BMP biology in the field of endocrinology are described and the possibility for clinical application of BMP activity is discussed.

Connexins in renal arterioles affect autoregulation of arteriolar tonus and renal blood flow and are believed to be involved in the transmission of the tubuloglomerular feedback (TGF) response across the cells of the juxtaglomerular apparatus. Connexin40 (Cx40) also plays a significant role in the regulation of renin secretion. We investigated the effect of deleting the Cx40 gene on autoregulation of afferent arteriolar diameter in response to acute changes in renal perfusion pressure. The experiments were performed using the isolated blood perfused juxtamedullary nephron preparation in kidneys obtained from wild-type or Cx40 knockout mice. Renal perfusion pressure was increased in steps from 75 to 155 mmHg, and the response in afferent arteriolar diameter was measured. Hereafter, a papillectomy was performed to inhibit TGF, and the pressure steps were repeated. Conduction of intercellular Ca(2+) changes in response to local electrical stimulation was examined in isolated interlobular arteries and afferent arterioles from wild-type or Cx40 knockout mice. Cx40 knockout mice had an impaired autoregulatory response to acute changes in renal perfusion pressure compared with wild-type mice. Inhibition of TGF by papillectomy significantly reduced autoregulation of afferent arteriolar diameter in wild-type mice. In Cx40 knockout mice, papillectomy did not affect the autoregulatory response, indicating that these mice have no functional TGF. Also, Cx40 knockout mice showed no conduction of intercellular Ca(2+) changes in response to local electrical stimulation of interlobular arteries, whereas the Ca(2+) response to norepinephrine was unaffected. These results suggest that Cx40 plays a significant role in the renal autoregulatory response of preglomerular resistance vessels.

HIV co-infection significantly impacts the natural history of hepatitis C virus (HCV) by increasing plasma HCV viral load, accelerating liver disease progression, and reducing rates of HCV clearance. Cytokines play an important role in regulating hepatic inflammation and fibrogenesis during chronic HCV infection, yet the impact of HIV on cytokine expression is unknown. In this study, an HCV continuous infection cell culture system was modified to permit co-infection with HIV to test the hypothesis that virus-induced disregulation of immune-response genes, particularly interferons and TGF-β, may create a permissive environment for the initial establishment of HIV/HCV co-infection in the host. CCR5-expressing Huh-7.5 hepatoma cells were transduced with human CD4 antigen to allow HIV infection in vitro. Co-infection of CD4⁺ Huh-7.5 cells with HIV and HCV or co-culture of HIV-infected CD4⁺ Huh-7.5 cells and HCV-infected Huh-7.5 cells increased the level of HCV RNA compared to HCV mono-infection. Quantitative gene expression analysis revealed HIV-induced up regulation of most tested IFN family genes when compared to HCV or co-infection. HCV infection induced up regulation of many TGF family genes that were subsequently down-regulated in the presence of HIV or HIV/HCV. Interestingly, co-infection resulted in down regulation of several IFN genes and significant up regulation of TGF-β genes leading to an overall enhancement of HCV replication. These data suggest that HIV infection may influence HCV replication in vitro by increasing levels of HCV RNA, possibly through the differential regulation of endogenous IFN and TGF family genes., (Copyright © 2012 Wiley Periodicals, Inc.)

Coronary vessel development depends on a subpopulation of epicardial cells that undergo epithelial to mesenchymal transformation (EMT) and invade the subepicardial space and myocardium. These cells form the smooth muscle of the vessels and fibroblasts, but the mechanisms that regulate these processes are poorly understood. Mice lacking the Type III Transforming Growth Factor β Receptor (TGFβR3) die by E14.5 due to failed coronary vessel development accompanied by reduced epicardial cell invasion. BMP2 signals via TGFβR3 emphasizing the importance of determining the relative contributions of the canonical BMP signaling pathway and TGFβR3-dependent signaling to BMP2 responsiveness. Here we examined the role of TGFβR3 in BMP2 signaling in epicardial cells. Whereas TGFβ induced loss of epithelial character and smooth muscle differentiation, BMP2 induced an ALK3-dependent loss of epithelial character and modestly inhibited TGFβ-stimulated differentiation. Tgfbr3(-/-) cells respond to BMP2 indicating that TGFβR3 is not required. However, Tgfbr3(-/-) cells show decreased invasion in response to BMP2 and overexpression of TGFβR3 in Tgfbr3(-/-) cells rescued invasion. Invasion was dependent on ALK5, ALK2, ALK3, and Smad4. Expression of TGFβR3 lacking the 3 C-terminal amino acids required to interact with the scaffolding protein GIPC (GAIP-interacting protein, C terminus) did not rescue. Knockdown of GIPC in Tgfbr3(+/+) or Tgfbr3(-/-) cells rescued with TGFβR3 decreased BMP2-stimulated invasion confirming a requirement for TGFβR3/GIPC interaction. Our results reveal the relative roles of TGFβR3-dependent and TGFβR3-independent signaling in the actions of BMP2 on epicardial cell behavior and demonstrate the critical role of TGFβR3 in mediating BMP2-stimulated invasion., (Copyright © 2011 Elsevier Inc. All rights reserved.)

Cutaneous scarring is often the epicenter of patient-related concerns, and the question "Will there be a scar?" is one that is all too familiar to the everyday clinician. In approaching this topic, we have reviewed the pathology, the embryology, and the molecular biology of cutaneous scarring., (Copyright © 2011 American Academy of Dermatology, Inc. Published by Mosby, Inc. All rights reserved.)

Intraoral bone grafting is routinely employed for implant site development prior or simultaneously to implant placement. Bone graft consolidation is a complex biological process depending on the formation of blood vessels into the augmented area. It is highly regulated by the angiogenesis and osteogenesis coupling phenomenon. The vascular system apart from supplying nutrients and oxygen to the developing and regenerating bone, also delivers critical signals which stimulate mesenchymal cell differentiation towards an osteogenic phenotype. Hypoxia inducible factors (HIFs) and mimicking agents (HMAs) (or alternatively HIF stabilizing agents) are considered to act as key stimulators of blood vessel formation. Under normoxia, HIFs are rapidly degraded. However, their degradation is prevented under hypoxia, which in turn, triggers angiogenesis. Hence, the major role of HMAs is to prevent degradation of HIFs under normoxic conditions. Recent studies suggest that HIFs and HMAs trigger the initiation and promotion of angiogenic-osteogenic cascade events. In vitro and animal studies involving genetic manipulation of individual components of the HIFs and HMAs have provided clues to how angiogenic-osteogenic coupling is achieved. Evidence from preclinical studies further suggests that topical application of HMAs enhance angiogenesis in intraoral augmented sites. In this article, we review the current understanding of the cellular and molecular mechanisms responsible for angiogenic-osteogenic coupling. We also discuss the therapeutic manipulation of HIFs and HMAs in intraoral bone repair and regeneration. Such discoveries suggest promising approaches for the development of novel therapies to improve intraoral bone repair and regeneration procedures., (Copyright © 2011 Elsevier Ltd. All rights reserved.)

The mechanisms that underlie the critical dendritic cell (DC) function in maintainance of peripheral immune tolerance are incompletely understood, although the β-catenin signaling pathway is critical for this role. The molecular details by which β-catenin signaling is regulated in DCs are unknown. Mechanical disruption of murine bone marrow-derived DC (BMDC) clusters activates DCs while maintaining their tolerogenic potential and this activation is associated with β-catenin signaling, providing a useful model with which to explore tolerance-associated β-catenin signaling in DCs. In this report, we demonstrate novel molecular features of the signaling events that control DC activation in response to mechanical stimulation. Non-canonical β-catenin signaling is an essential component of this tolerogenic activation and is modulated by adhesion molecules, including integrins. This unique β-catenin-dependent signaling pathway is constitutively active at low levels, suggesting that mechanical stimulation is not necessarily required for induction of this unique activation program. We additionally find that the immunomodulatory cytokine TGF-β antagonizes β-catenin in DCs, thereby selectively suppressing signaling associated with tolerogenic DC activation while having no impact on LPS-induced, β-catenin-independent immunogenic activation. These findings provide new molecular insight into the regulation of a critical signaling pathway for DC function in peripheral immune tolerance.

Endoglin (CD105) is an integral membrane glycoprotein that serves as a coreceptor for members of the transforming growth factor-β superfamily of proteins. A major role for endoglin in regulating transforming growth factor-β-dependent vascular remodeling and angiogenesis has been postulated based on the following: 1) endoglin is the gene mutated in hereditary hemorrhagic telangiectasia type 1, a disease characterized by vascular malformations; 2) endoglin knockout mice die at midgestation because of defective angiogenesis; 3) endoglin is overexpressed in neoangiogenic vessels, during inflammation, and in solid tumors; and 4) endoglin regulates the expression and activity of endothelial nitric oxide synthase, which is involved in angiogenesis and vascular tone. Besides the predominant form of the endoglin receptor (long endoglin isoform), two additional forms of endoglin have been recently reported to play a role in the vascular pathology and homeostasis: the alternatively spliced short endoglin isoform and a soluble endoglin form that is proteolytically cleaved from membrane-bound endoglin. The purpose of this review is to underline the role that the different forms of endoglin play in regulating angiogenesis, vascular remodeling, and vascular tone, as well as to analyze the molecular and cellular mechanisms supporting these effects.

The CCN (Cyr61 (cysteine-rich protein 61), CTGF (connective tissue growth factor), Nov (nephroblastoma overexpressed)) family consists of six members that belong to matricellular proteins of extracellular matrix (ECM). Like other matricellular proteins, CCN members do not primarily have a structural role; however, they modulate cell-ECM interactions. In general, CCN proteins are formed by four characteristic domain structures and thought to participate in various biological phenomena such as organ development, wound healing, angiogenesis, fibrosis, etc. In cancer, CCN proteins family expresses aberrantly; probably depending on the sites and types, expressions of different CCN proteins have been documented to be linked with either progression or inhibition of the pathological processes of cancer. Through various mechanisms like cell survival, apoptosis, inflammation, cell adhesion and migration and connection with several cytokines, CCN proteins perhaps influence the disease course including tumor metastasis. A majority of the above-mentioned effects are believed to be mediated by binding with integrins, a class of receptors that mediate cell-cell and cell-ECM interactions. Furthermore, the members of CCN family modulate the functions of several important growth factors and related pathways such as insulin-like growth factor (IGF), transforming growth factor-beta (TGF-beta) and Wnt signaling. Interestingly, a variety of factors/proteins linked with these signaling systems are reported to be associated with the carcinogenic process. Nevertheless, a precise knowledge about the pathophysiological activities including signaling pathways of CCN proteins would be helpful to identify molecular targets in order to design therapeutic strategies in the management of cancer.

There is increasing evidence that human development before implantation is regulated by embryonically and maternally derived growth factors. The "regulators" of embryonic origin such as soluble human leukocyte antigen G, platelet-activating factor, Th1/Th2 cytokines, insulin-like growth factor, epidermal growth factor, transforming growth factor alpha, colony-stimulating factor, platelet-derived growth factor may be used as indicators of embryo viability and implantation potential. The data prove the influence of growth factors on the development and growth of preimplantation embryos. Though there is a lot of research in the field of biomarkers during folliculogenesis and maternal-fetal interface, only few of them deal with regulators derived from embryonic cells to the cultivation medium. The aim of our study was to summarize the research dealing with immune markers produced by embryos in vitro and to estimate their impact on the cell growth, viability and implantation potential.

The bronchial epithelium is the barrier to the external environment and plays a vital role in protection of the internal milieu of the lung. It functions within the epithelial-mesenchymal trophic unit to control the local microenvironment and help maintain tissue homeostasis. However, in asthma, chronic perturbation of these homeostatic mechanisms leads to alterations in the structure of the airways, termed remodeling. Damage to the epithelium is now recognized to play a key role in driving airway remodeling. We have postulated that epithelial susceptibility to environmental stress and injury together with impaired repair responses results in generation of signals that act on the underlying mesenchyme to propagate and amplify inflammatory and remodeling responses in the submucosa. Many types of challenges to the epithelium, including pathogens, allergens, environmental pollutants, cigarette smoke, and even mechanical forces, can elicit production of mediators by the epithelium, which can be translated into remodeling responses by the mesenchyme. Several important mediators of remodeling have been identified, most notably transforming growth factor-beta, which is released from damaged/repairing epithelium or in response to inflammatory mediators, such as IL-13. The cross talk between the epithelium and the underlying mesenchyme to drive remodeling responses is considered in the context of subepithelial fibrosis and potential pathogenetic mechanisms linked to the asthma susceptibility gene, a disintegrin and metalloprotease (ADAM)33.

Hypertension and its sequelae are complex processes. Optimization of the care of the hypertensive patient requires not only attention to the regulation of arterial pressure but also attention to blunting the hypertension-related processes that lead to vascular disease. It is clear that the regulation of these processes is much more complex than previously understood. Here several new insights into the pathogenesis of hypertension-related vascular disease have been explored. While this review is not exhaustive, it does serve to point out the varied nature of the biologic processes that must be taken into account and it points to new avenues for the development of therapeutic agents.

Foxp3(+)CD25(+)CD4(+) regulatory T cells are produced in the thymus (natural T regs) but can also differentiate from peripheral Foxp3(-)CD4(+) precursors (induced or adaptive T regs). We assessed antigen presenting cell (APC) requirements for the latter differentiation. With added transforming growth factor (TGF)-beta, both immature and mature populations of dendritic cells (DCs) induced antigen-specific Foxp3(+) T regs from Foxp3(-) precursors. Using endogenous TGF-beta, DCs from gut-associated mesenteric lymph nodes were capable of differentiating Foxp3(+)T regs. Spleen DCs were 100-fold more potent than DC-depleted APCs for the induction of T regs and required 10-fold lower doses of peptide antigen. Interleukin-2 (IL-2) was essential, but could be provided endogenously by T cells stimulated by DCs, but not other APCs. The required IL-2 was induced by DCs that expressed CD80/CD86 costimulatory molecules. The DC-induced Foxp3(+)T regs divided up to 6 times in 6 days and were comprised of CD62L and CD103 positive and negative forms. The induced Foxp3(+)T regs exerted suppression in vitro and blocked tumor immunity in vivo. These results indicate that DCs are specialized to differentiate functional peripheral Foxp3(+)T regs and help set the stage to use DCs to actively suppress the immune response in an antigen-specific manner.

Problem: Cytokines are obviously very important in an established pregnancy, but what about human embryo implantation?, Methods: Literature review., Results: We first discuss the necessity and limits of animal models, and then review the few cytokines which have been demonstrated by knock-out methods to be absolutely necessary for embryo implantation using in animal models. We then review what is known or discussed about the role of other cytokines as deduced from quantitative and/or qualitative dysregulation in animals and in humans., Conclusions: Cytokines are indeed involved in implantation as they are in ongoing pregnancy and delivery. Relevance to infertility and recurrent pregnancy loss is discussed.

Adenosine triphosphate (ATP) is liberated from macula densa cells in response to increased tubular NaCl delivery. However, it is not known whether ATP from the macula densa is broken down to adenosine, or whether this adenosine mediates efferent arteriole (Ef-Art) tubuloglomerular feedback (TGF). We hypothesized that increased macula densa Ca(2+), release of ATP and degradation of ATP to adenosine are necessary for Ef-Art TGF. Rabbit Ef-Arts and adherent tubular segments (with the macula densa) were simultaneously microperfused in vitro while changing the NaCl concentration at the macula densa. The Ef-Art was perfused orthograde through the end of the afferent arteriole (Af-Art). In Ef-Arts preconstricted with norepinephrine (NE), increasing NaCl concentration from 10 to 80 mM at the macula densa dilated Ef-Arts from 7.5+/-0.7 to 11.1+/-0.3 microm. Buffering increases in macula densa Ca(2+) with the cell-permeant Ca(2+) chelator BAPTA-AM diminished Ef-Art TGF from 3.1+/-0.3 to 0.1+/-0.2 microm. Blocking adenosine formation by adding alpha-beta-methyleneadenosine 5'-diphosphate (MADP) blocked Ef-Art TGF from 2.9+/-0.5 to 0.1+/-0.2 microm. Increasing luminal NaCl at the macula densa from 10 to 45 mM caused a moderate Ef-Art TGF response, 1.3+/-0.1 microm. It was potentiated to 4.0+/-0.3 microm by adding hexokinase, which enhances conversion of ATP into adenosine. Our data show that in vitro changes in macula densa Ca(2+) and ATP release are necessary for Ef-Art TGF. ATP is broken down to form adenosine, which mediates signal transmission of Ef-Art TGF.

The vestibular schwannoma is a benign, slow-growing neoplasm that originates from the neurolemmal sheath of the vestibular branch of the VIIIth cranial nerve. This tumor entity accounts for 6 % of all intracranial tumors and the annual incidence of newly diagnosed vestibular schwannoma is reported as 13 per million. The molecular pathogenesis of both sporadic vestibular schwannoma and those occurring in neurofibromatosis type II appears to be associated with an aberration of a tumor suppressor gene on chromosome 22q12. The biological background for the various growth patterns of vestibular schwannoma is, however, largely unknown. This differing clinical and biological behaviour of vestibular schwannoma may be explained by the presence of neurotrophic factors. The results of recent immunohistochemical studies demonstrate the co-expression of transforming growth factor (TGF)-beta 1 and glial cell line-derived neurotrophic factor (GDNF) in vestibular schwannoma and suggest a trophic synergism of both neurotrophic factors in this tumor. Moreover, expression of numerous different neurotrophic factors has been shown in studies of nerve growth factor (NGF), vascular endothelial growth factor (VEGF), epidermal growth factor (EGF), fibroblast growth factor (FGF), neuregulin (NRG) and erythropoietin (EPO) indicating a biological role in development, maintainance or growth of vestibular schwannoma. In this article, we summarize the findings on neurotrophic factor expression and discuss their characteristics and biological role in vestibular schwannoma.

To achieve the goals of engineering large complex tissues, and possibly internal organs, vascularization of the regenerating tissue is essential. To maintain the initial volume after implantation of regenerated tissue, improved vascularization is considered to be important. Recent advances in understanding the process of blood vessel growth has offered significant tools for the neovascularization of bioengineered tissues and therapeutic angiogenesis. Several angiogenic growth factors including vascular endothelial growth factor (VEGF), basic fibroblast growth factor (bFGF) and hepatocyte growth factor (HGF) were used for vascularization of ischemic tissues. Other approaches such as prevascularization of the scaffold, prior to cell seeding, and incorporation of endothelial cells in the bioengineered tissue showed encouraging results. In this article, we will review recent advances in angiogenic growth factors, and discuss the role of these growth factors and endothelial cells in therapeutic angiogenesis and tissue engineering.

Embryonic stem cells (ESCs) are the pluripotent cell population derived from the inner cell mass of pre-implantation embryos and are characterised by prolonged self-renewal and the potential to differentiate into cells representing all three germ layers both in vitro and in vivo. Preservation of the undifferentiated status of the ESC population requires the maintenance of self-renewal whilst inhibiting differentiation and regulating senescence and apoptosis. In this review, we discuss the intrinsic and extrinsic factors associated with self-renewal process, together with possible signalling pathway interactions and mechanisms of regulation.

Endocrine tumours of the gut and pancreas originate from cells of the diffuse endocrine system and are characterised by the production of a wide variety of bioactive substances including growth factors. Two major tumour categories are distinguished-well-differentiated and poorly differentiated neoplasms-with distinct phenotypes and significantly diverse clinical behaviour. Here, genetic background data are summarised on an anatomical basis for tumours of foregut, midgut and hindgut derivatives. For well-differentiated tumours, independent techniques identified the abnormality of multiple chromosomal sites and genes, pointing to a complex genetic background. Differences in foregut tumours compared with midgut and hindgut tumours are, however, outlined. The multiple endocrine neoplasia syndrome type 1 (MEN1) gene is reported to be involved in about one-third of sporadic foregut endocrine tumours and exceptionally in midgut and hindgut tumours. Similarly, X chromosome markers are associated with malignant behaviour in foregut tumours only. For poorly differentiated carcinomas, a high degree of chromosomal instability is the common genetic trait independent of tumour site and frequently involving the p53 gene.

Genetic instability of tumor cells often leads to amplified expression of multiple growth factors that contribute to angiogenesis and tumor growth. Members of the platelet-derived growth factor (PDGF) family are frequently utilized growth factors by many tumors to support their growth. PDGFs have previously been found to induce tumor growth by directly stimulating cell growth of certain types of tumors. We have recently demonstrated that PDGFs are potent angiogenic factors. Particularly, the angiogenic activity of PDGFs can be potentiated in the presence of other angiogenic factors. In addition to stimulation of blood angiogenesis, we have recently found that PDGFs can directly stimulate lymphangiogenesis and lymphatic metastasis. In this review, multiple roles of PDGFs in control of tumor growth and metastasis are discussed.

Over the last decade, with the advent of new techniques and technologies in modern molecular biology, our understanding of the underlying mechanisms responsible for organ differentiation has developed rapidly. Despite this, our knowledge of these signaling pathways is still far from complete. Some of these advances, such as the creation of transgenic mouse models, have given us new tools to help us understand the interactions of the various transcription factors that are responsible for the creation of various cell types from a single cell type during embryogenesis. This knowledge then gives rise to the concept of creating new ways to manipulate stem cells in order to correct the deficiencies present in various disease processes. Here, we present work that focuses specifically on pancreatic development. The ultimate goal of our research in studying the mechanisms of the basic differentiation of pancreatic precursor cells is to gain the knowledge necessary to be able to engineer stem cells specifically into beta-cells in the treatment of diabetes.

The transforming growth factor beta (TGFbeta) superfamily has wide-ranging and profound effects on many aspects of cellular growth and development. Many TGFbeta-related ligands, receptors, and intracellular signaling proteins are expressed in the ovary and are critical for normal follicle development. Our laboratory and others have analyzed the in vivo function of the TGFbeta superfamily signal transduction pathways by using gene knockout and knockin approaches. Two TGFbeta superfamily ligands, growth differentiation factor 9 (GDF9) and bone morphogenetic protein 15 (BMP15), are expressed in developing oocytes. Based on in vivo data using knockout models, GDF9 is critical at both the primary and preovulatory stages of follicle development, and physiologically interacts with BMP15 during the latter stages of folliculogenesis. A knockin model of activin betaB expressed from the activin betaA locus, revealed that activin betaB can act as a hypomorphic protein and rescue some but not all of activin betaAs functions. Questions of functional redundancy of signaling components and multiple receptor utilization by different ligands still need to be addressed for these pathways. Answers will likely come from using existing single null mouse models to generate combinatorial ligand and receptor null mice. These new models may reveal the in vivo genetic interactions of TGFbeta superfamily ligands, receptors, binding proteins, and downstream signaling pathways.

Autoregulation of renal blood flow is an established physiological phenomenon, however the signalling mechanisms involved remain elusive. Autoregulatory adjustments in preglomerular resistance involve myogenic and tubuloglomerular feedback (TGF) influences. While there is general agreement on the participation of these two regulatory pathways, the signalling molecules and effector mechanisms have not been identified. Currently, there are two major hypotheses being considered to explain the mechanism by which TGF signals are transmitted from the macula densa to the afferent arteriole. The adenosine hypothesis proposes that the released adenosine triphosphate (ATP) is hydrolysed to adenosine and this product stimulates preglomerular vasoconstriction by activation of A(1) receptors on the afferent arteriole. Alternatively, the P2 receptor hypothesis postulates that ATP released from the macula densa directly stimulates afferent arteriolar vasoconstriction by activation of ATP-sensitive P2X(1) receptors. This hypothesis has emerged from the realization that P2X(1) receptors are heavily expressed along the preglomerular vasculature. Inactivation of P2X(1) receptors impairs autoregulatory responses while afferent arteriolar responses to A(1) adenosine receptor activation are retained. Autoregulatory behaviour is markedly attenuated in mice lacking P2X(1) receptors but responses to adenosine A(1) receptor activation remain intact. More recent experiments suggest that P2X(1) receptors play an essential role in TGF-dependent vasoconstriction of the afferent arteriole. Interruption of TGF-dependent influences on afferent arteriolar diameter, by papillectomy or furosemide treatment, significantly attenuated pressure-mediated afferent arteriolar vasoconstriction in wild-type mice but had no effect on the response in P2X(1) knockout mice. Collectively, these observations support an essential role for P2X(1) receptors in TGF-mediated afferent arteriolar vasoconstriction.

Aim: By crossing aquaporin 1 (AQP1)-/- and adenosine 1 receptor (A1AR)-/- mice, we generated an animal model that combines a proximal tubular absorption defect with absence of tubuloglomerular feedback (TGF) regulation of glomerular filtration rate (GFR). The aim of studies in these animals was to determine whether a TGF-induced reduction of GFR is a prerequisite for preventing potentially fatal fluid losses., Methods and Results: In contrast to AQP1 deficient mice, AQP1/A1AR-/- mice were found to have a normal GFR. TGF responses were abolished in these animals, in contrast to AQP1-/- mice in which TGF responses of single nephron glomerular filtration rate (SNGFR) were left-shifted. Proximal tubule fluid absorption in AQP1/A1AR-/- mice was reduced to levels previously reported for AQP1-/- mice. However, SNGFR was significantly higher in AQP1/A1AR-/- than AQP1-/- mice (10.6 +/- 0.8 nL min(-1) vs. 5.9 +/- 0.7 nL min(-1)). As a consequence of the normal GFR and the reduced proximal reabsorption distal fluid delivery was markedly higher in the double knockout compared with normal or AQP1-/- mice (5.5 +/- 0.5 nL min(-1) vs. 2.35 +/- 0.3 nL min(-1) in AQP1-/-). Despite the approximate doubling of distal fluid and Cl delivery, AQP1/A1AR-/- mice have a normal salt excretion, normal arterial blood pressure, and only a small increase in plasma renin concentration., Conclusion: The ability to compensate for proximal tubule malabsorption without a TGF-induced reduction of GFR attests to a remarkable adaptability of distal tubule transport mechanisms.

Inflammatory response and cytokine elaboration are particularly active after myocardial infarction and contribute to cardiac remodeling and eventual host outcome. The triggers of cytokine release in the acute postinfarction period include mechanical deformation, ischemic stimulus, reactive oxygen species (ROS), and cytokine self-amplification pathways. Acutely, the elaboration of tumor necrosis factor, IL-1 and IL-6, transforming growth factor families of cytokines, contribute to survival or deaths of myocytes, modulation of cardiac contractility, alterations of vascular endothelium, and recruitment of additional circulating cells of inflammation to the injured myocardium. This leads to further local oxidative stress and remodeling but also initiates the processes of wound healing. Chronically, sustained presence of cytokines leads to myocyte phenotype transition and activation of matrix metalloproteinases that modifies interstitial matrix, augmenting further the remodeling process. This in turn alters the local collagen composition and also the integrins that constitute the interface between myocytes and the matrix. These processes ultimately, when favorable, pave the way for angiogenesis and cellular regeneration. Thus, the insightful modulation of cytokines through current and future therapies could promote improved healing and cardiac remodeling postmyocardial infarction.

Brain tumors are a diverse group of malignancies that remain refractory to conventional treatment approaches, including radiotherapy and cytotoxic chemotherapy. Molecular neuro-oncology has now begun to clarify the transformed phenotype of brain tumors and identify oncogenic pathways that may be amenable to targeted therapy. Growth factor signaling pathways are often upregulated in brain tumors and may contribute to oncogenesis through autocrine and paracrine mechanisms. Excessive growth factor receptor stimulation can also lead to overactivity of the Ras signaling pathway, which is frequently aberrant in brain tumors. Receptor tyrosine kinase inhibitors, antireceptor monoclonal antibodies and antisense oligonucleotides are targeted approaches under investigation as methods to regulate aberrant growth factor signaling pathways in brain tumors. Several receptor tyrosine kinase inhibitors, including imatinib mesylate (Gleevec), gefitinib (Iressa) and erlotinib (Tarceva), have entered clinical trials for high-grade glioma patients. Farnesyl transferase inhibitors, such as tipifarnib (Zarnestra), which impair processing of proRas and inhibit the Ras signaling pathway, have also entered clinical trials for patients with malignant gliomas. Further development of targeted therapies and evaluation of these new agents in clinical trials will be needed to improve survival and quality of life of patients with brain tumors.

In Drosophila dorsoventral (DV) polarity arises during oogenesis when the oocyte nucleus moves from a central posterior to an asymmetrical anterior position. Nuclear movement is a symmetry-breaking step and establishes orthogonality between the anteroposterior and the DV axes. The asymmetrically anchored nucleus defines a cortical region within the oocyte which accumulates high levels of gurken messenger RNA (mRNA) and protein. Gurken is an ovarian-specific member of the transforming growth factor-alpha (TGF-alpha) family of secreted ligands. Secreted Gurken forms a concentration gradient that results in a dorsal-to-ventral gradient of EGF receptor activation in the follicle cells surrounding the oocyte. This leads to concentration-dependent activation or repression of target genes of the EGF pathway in the follicular epithelium. One outcome of this process is the restriction of pipe expression to a ventral domain that comprises 40% of the egg circumference. Pipe presumably modifies extracellular matrix components that are secreted by the follicle cells and are present at the ventral side of embryo after egg deposition. Here, they activate a proteolytic cascade that generates a gradient of the diffusible ligand, Spätzle. Spätzle activates the Toll receptor at the surface of the embryo that stimulates the nuclear uptake of the transcription factor Dorsal. This leads to a nuclear concentration gradient of Dorsal that specifies the cell types along the DV axis of the embryo.

Objective: To review research progress of the relation between growth factor and repair of intervertebral disc., Methods: The recent articles on growth factor and repair of intervertebral disc were extensively reviewed. The expression of growth factor in intervertebral disc and the effect of growth factor on disc cells were investigated., Results: Some growth factors play roles in the development and degeneration of intervertebral disc. Exogenous growth factor can increase proliferation of disc cells and production of proteoglycans and collagens. Gene of growth factor can be transferred to intervertebral disc cell by adenovirus., Conclusion: Growth factor plays an important role in the regulation of development and degeneration of interertebral disc. The above results show that the feasibility of usage of growth factor in the treatment of disc degeneration and in repair and reconstruction of disc.

The COP9 signalosome (CSN) is linked to signaling pathways and ubiquitin-dependent protein degradation in yeast, plant and mammalian cells, but its roles in Drosophila development are just beginning to be understood. We show that during oogenesis CSN5/JAB1, one subunit of the CSN, is required for meiotic progression and for establishment of both the AP and DV axes of the Drosophila oocyte. The EGFR ligand Gurken is essential for both axes, and our results show that CSN5 mutations block the accumulation of Gurken protein in the oocyte. CSN5 mutations also cause the modification of Vasa, which is known to be required for Gurken translation. This CSN5 phenotype - defective axis formation, reduced Gurken accumulation and modification of Vasa - is very similar to the phenotype of the spindle-class genes that are required for the repair of meiotic recombination-induced, DNA double-strand breaks. When these breaks are not repaired, a DNA damage checkpoint mediated by mei-41 is activated. Accordingly, the CSN5 phenotype is suppressed by mutations in mei-41 or by mutations in mei-W68, which is required for double strand break formation. These results suggest that, like the spindle-class genes, CSN5 regulates axis formation by checkpoint-dependent, translational control of Gurken. They also reveal a link between DNA repair, axis formation and the COP9 signalosome, a protein complex that acts in multiple signaling pathways by regulating protein stability.

Basic scientific research has demonstrated that epidermal growth factor (EGF), basic fibroblast growth factor (bFGF), platelet-derived growth factor-AA (PDGF-AA) and transforming growth factor-alpha (TGF-alpha) are induced by acute tympanic membrane (TM) perforation. The expression of these growth factors peaks during the inflammatory phase and then declines, suggesting their involvement in the healing process of the TM. Expression of EGF receptor, bFGF receptor, PDGF receptor and keratinocyte growth factor (KGF) receptor in the TM has also been reported. Identification of these receptors indicates that the target cells in the TM may be stimulated during the process of healing by the corresponding exogenous growth factor in vivo. Some reports from animal studies showed encouraging effects of EGF, bFGF and PDGF-AA in terms of accelerated or enhanced healing of acute and chronic TM perforations without significant adverse effects. Two reports of clinical trials of topical application of EGF or bFGF for TM perforations revealed mixed results. In this article special considerations for future directions of research into growth factors are discussed and related articles on healing of skin wounds and other lesions are reviewed. Further experimental and clinical studies on the mechanism of action of growth factors, timing of application, selection (either singly or in combination), delivery mode, dose and safety aspects, as well as more clinical trials, are warranted, and will pave the way for clinical application.

The establishment of axial polarity in the Drosophila egg and embryo depends on intercellular communication between two cell types in the ovary, the germline, and the soma. The genes gurken and egfr encode two essential players of this communication pathway. Gurken protein, a TGF-alpha-like molecule, is expressed in the germline, while the EGF-receptor homolog, Egfr, is expressed in the somatic cells of the ovary. Using the yeast two-hybrid system we show here, for the first time, that Gurken protein directly binds to the extracellular domain of Egfr. This direct physical association requires the presence of an intact EGF motif within Gurken protein. Furthermore, we provide evidence that this characteristic motif may be sufficient for interaction with the receptor, at list in vitro. Our results firmly establish Gurken as the germline ligand of Drosophila Egfr.

Astroglial-derived factors, as transforming growth factor (TGF)alpha and TGFbeta, act in the hypothalamus to activate luteinizing hormone-releasing hormone (LHRH) secretion. Hypothalamic hamartomas (HHs) contain normal nervous tissue in a heterotopic location. When symptomatic, they cause precocious puberty and/or characteristic gelastic seizures. Thus far, the pathogenesis of these alterations remains unknown. By examining two HHs associated with sexual precocity, we found that they contained astroglial cells expressing TGFalpha, but no LHRH neurons. In a third patient with HH, only epilepsy was present, but precocious puberty developed shortly after surgery, probably as a consequence of a surgery-induced lesion. These results imply that some HHs induce sexual precocity by activating endogenous LHRH secretion via astroglial-derived factors., (Copyright 2002 S. Karger AG, Basel)

We have analyzed the mechanism of activation of the Epidermal growth factor receptor (Egfr) by the transforming growth factor (TGF) alpha-like molecule, Gurken (Grk). Grk is expressed in the oocyte and activates the Egfr in the surrounding follicle cells during oogenesis. We show that expression of either a membrane bound form of Grk (mbGrk), or a secreted form of Grk (secGrk), in either the follicle cells or in the germline, activates the Egfr. In tissue culture cells, both forms can bind to the Egfr; however, only the soluble form can trigger Egfr signaling, which is consistent with the observed cleavage of Grk in vivo. We find that the two transmembrane proteins Star and Brho potentiate the activity of mbGrk. These two proteins collaborate to promote an activating proteolytic cleavage and release of Grk. After cleavage, the extracellular domain of Grk is secreted from the oocyte to activate the Egfr in the follicular epithelium.

In Drosophila, dorsoventral polarity is established by the asymmetric positioning of the oocyte nucleus. In egg chambers mutant for cap 'n' collar, the oocyte nucleus migrates correctly from a posterior to an anterior-dorsal position where it remains during stage 9 of oogenesis. However, at the end of stage 9, the nucleus leaves its anterior position and migrates towards the posterior pole. The mislocalisation of the nucleus is accompanied by changes in the microtubule network and a failure to maintain bicoid and oskar mRNAs at the anterior and posterior poles, respectively. gurken mRNA associates with the oocyte nucleus in cap 'n' collar mutants and initially the local secretion of Gurken protein activates the Drosophila EGF receptor in the overlying dorsal follicle cells. However, despite the presence of spatially correct Grk signalling during stage 9, eggs laid by cap 'n' collar females lack dorsoventral polarity. cap 'n' collar mutants, therefore, allow for the study of the influence of Grk signal duration on DV patterning in the follicular epithelium., (Copyright 2001 Academic Press.)

Background and Aims: Mice lacking epidermal growth factor (EGF), transforming growth factor alpha, and amphiregulin were used to identify roles for these EGF receptor (EGF-R) ligands in gastrointestinal development and mucosal integrity., Methods: Gastrointestinal tract development was examined in knockout mice and correlated with expression of EGF-R protein and EGF family members throughout the gut. Crossfostering experiments addressed roles of maternal- and neonatal-derived ligands in pup growth and intestinal development. Cysteamine-induced ulceration in EGF(-/-) mice was used to examine its role in mucosal cytoprotection., Results: Neonatal mice lacking all 3 ligands were growth retarded, even when reared by wild-type dams; conversely, lack of maternal ligands transiently impaired wild-type pup growth. Triple null neonates displayed spontaneous duodenal lesions, and ileal villi were truncated and fragile with reduced cellular proliferation in the crypts. However, maturation of digestive enzymes was unaffected. Adult EGF(-/-) mice displayed more severe lesions in response to cysteamine treatment compared with wild-type counterparts, although triple null mice were not more susceptible to dextran sulfate sodium-induced colitis, suggesting a differential role for these ligands in the injury response., Conclusions: EGF-R ligands are required for development and mucosal maintenance in mouse small intestine. Both maternal and neonatal sources of growth factors are required for optimal pup growth.

Pancreatic cancer is, indisputably, one of the most malignant gastrointestinal tumors. Although the etiology of this disease is unknown, it is clearly linked to alterations in the biologic activities of various signaling molecules. Aberrant signaling activities of growth factors and their receptors, transcription factors, and proteins that control the cell cycle have been increasingly implicated in the pathogenesis and dissemination of pancreatic tumors. It is indeed possible that several of these molecules are, in fact, part of a signaling network that has gone awry. This review summarizes some recent advances in an attempt to generate a working model for future investigations.

The body axes of Drosophila are established during oogenesis through reciprocal interactions between the germ line cells and the somatic follicle cells that surround them. The Notch pathway is required at two stages in this process: first, for the migration of the follicle cells around the germ line cyst and, later, for the polarization of the anterior-posterior (A-P) axis of the oocyte. Its function in these events, however, has remained controversial. Using clonal analysis, we show that Notch signaling controls cell proliferation and differentiation in the whole follicular epithelium. Notch mutant follicle cells remain in a precursor state and fail to switch from the mitotic cell cycle to the endocycle. Furthermore, removal of Delta from the germ line produces an identical phenotype, showing that Delta signals from the germ cells to control the timing of follicle cell differentiation. This explains the axis formation defects in Notch mutants, which arise because undifferentiated posterior follicle cells cannot signal to polarize the oocyte. Delta also signals from the germ line to Notch in the soma earlier in oogenesis to control the differentiation of the polar and stalk follicle cells. The germ line therefore regulates the development of the follicle cells through two complementary signaling pathways: Gurken signals twice to control spatial patterning, whereas Delta signals twice to exert temporal control.

-The predominant cause of restenosis after angioplasty is now thought to be inward remodeling, but the mechanisms responsible are unknown. Remodeling in normal vessels is regulated by the endothelium in response to altered shear stress. Although the endothelium is often damaged by angioplasty, restenosis rates after angioplasty have been correlated with impaired coronary flow. Thus, we examined how increases or decreases in blood flow through balloon catheter-injured rat carotid arteries affect vessel morphometry (4, 10, and 28 days), cell migration (4 days), and levels of promigratory mRNAs (2 and 10 days). After 28 days, the luminal area in vessels with low blood flow was significantly less than in those with normal and high blood flow (0.17+/-0.01 [low] versus 0.24+/-0.06 [normal] versus 0.30+/-0.02 [high] mm(2), P:<0.01), predominantly because of accentuated inward remodeling (or reduced area within the external elastic lamina; 0.42+/-0.02 [low] versus 0.54+/-0.07 [normal] versus 0.53+/-0.04 [high] mm(2), P:<0.05). Low flow also enhanced smooth muscle cell migration 4 days after injury by 90% above normal and high flows (P:<0.01). Two days after injury, low flow significantly increased levels of mRNAs encoding promigratory peptides (integrin alpha(v)ss(3), transforming growth factor-ss(1), CD44v6, MDC9, urokinase plasminogen activator receptor, and ss-inducible gene h3); these changes persisted 10 days after injury and were localized to the neointima. Low blood flow may promote restenosis after angioplasty because of its adverse effect on vessel remodeling, and it is associated with the augmented expression of multiple genes central to cell migration and restenosis.

Directed cell migration is important for many aspects of normal animal development, but little is known about how cell migrations are guided or the mechanisms by which guidance cues are translated into directed cell movement. Here we present evidence that signaling mediated by the epidermal growth factor receptor (EGFR) guides dorsal migration of border cells during Drosophila oogenesis. The transforming growth factor-alpha (TGF-alpha)-like ligand Gurken appears to serve as the guidance cue. To mediate this guidance function, EGFR signals via a pathway that is independent of Raf-MAP kinase and receptor-specific.