Hypoxia-mediated induction of acidic/basic fibroblast growth factor and platelet-derived growth factor in mononuclear phagocytes stimulates growth of hypoxic endothelial cells.

Wound repair and tumor vascularization depend upon blood vessel growth into hypoxic tissue. Although hypoxia slows endothelial cell (EC) proliferation and suppresses EC basic fibroblast growth factor (bFGF) expression, we report that macrophages (MPs) exposed to PO2 approximately 12-14 torr (1 torr = 133.3 Pa) synthesize and release in a time-dependent manner platelet-derived growth factor (PDGF) and acidic/basic FGFs (a/bFGFs), which stimulate the growth of hypoxic ECs. Chromatography of hypoxic MP-conditioned medium on immobilized heparin with an ascending NaCl gradient resolved three peaks of mitogenic activity: activity of the first peak was neutralized by antibody to PDGF; activity of the second peak was neutralized by antibody to aFGF; and activity of the third peak was neutralized by antibody to bFGF. Metabolically labeled lysates and supernatants from MPs exposed to hypoxia showed increased synthesis and release of immunoprecipitable PDGF and a/bFGF in the absence of changes in cell viability. Possible involvement of a heme-containing oxygen sensor in MP elaboration of growth factors was suggested by the induction of bFGF and PDGF by normoxic MPs exposed to nickel or cobalt, although metabolic inhibitors such as sodium azide were without effect. These results suggest a paracrine model in which hypoxia stimulates MP release of PDGF and a/bFGF, inducing EC proliferation and potentially promoting angiogenesis in hypoxic environments.

Induction of cell proliferation in mammalian inner-ear sensory epithelia by transforming growth factor alpha and epidermal growth factor.

Regenerative proliferation occurs in the inner-ear sensory epithelial of warm-blooded vertebrates after insult. To determine how this proliferation is controlled in the mature mammalian inner ear, several growth factors were tested for effects on progenitor-cell division in cultured mouse vestibular sensory epithelia. Cell proliferation was induced in the sensory epithelium by transforming growth factor alpha (TGF-alpha) in a dose-dependent manner. Proliferation was also induced by epidermal growth factor (EGF) when supplemented with insulin, but not EGF alone. These observations suggest that stimulation of the EGF receptors by TGF-alpha binding, or EGF (plus insulin) binding, stimulates cell proliferation in the mature mammalian vestibular sensory epithelium.

Fluency in interpreting: a native-like quality factor

Lo studio si occupa di fluency e analizza alcuni aspetti che la definiscono (pause vuote, pause piene, segnali discorsivi, riformulazioni). Si analizzano frequenza e durata di tali fenomeni, attraverso due corpora di produzioni orali di due gruppi di parlanti della lingua inglese: gli studenti italiani del corso di Mediazione Linguistica Interculturale della Scuola di Lingue, Letterature, Interpretazione e Traduzione di Forlì, Università di Bologna, e partecipanti britannici di un programma radiofonico. Si è ritenuto utile comparare le produzioni orali di studenti della lingua inglese a quelle di oratori pubblici madrelingua. Si è cercato di bilanciare i due corpora in termini di genere. Sono stati utilzzati i software Praat, per identificare la morfologia e la durata delle variabili, e Notetab Light, per l'annotazione dei corpora. I risultati della ricerca mostrano che le differenze maggiori tra i due gruppi risiedono nella durata delle pause vuote e nella frequenza, durata e e varietà di suoni delle pause piene, oltre a sillabe aggiuntive, sillabe allungate e riformulazioni. Le sillabe aggiuntive appaiono tipiche della produzione orale degli studenti italiani, in quanto, per la maggior parte, le parole della lingua italiana terminano con un suono vocalico. E' inoltre emersa una questione di genere. Le parlanti di sesso femminile, in entrambi i corpora, impiegano maggiormente le variabili della fluency prese in esame, rispetto ai parlanti di sesso maschile. Sulla base di questa ricerca e ricerche future si potranno ideare moduli di insegnamento dell'inglese basati sulla fluency come fattore primario di competenza linguistica. Il Capitolo 1 introduce lo studio. Il Capitolo 2 presenta lo stato dell'arte sul tema. Il Capitolo 3 presenta la metodologia dello studio. Il Capitolo 4 è dedicato a illustrare e discutere i risultati della ricerca. Il Capitolo 5 presenta considerazioni conclusive e future prospettive per l'insegnamento dell'inglese e per la ricerca.

Fibroblast growth factor 23 and markers of mineral metabolism in individuals with preserved renal function

Fibroblast growth factor 23 (FGF23) is a bone-derived hormone that regulates phosphate homeostasis. Circulating FGF23 is elevated in chronic kidney disease (CKD) and independently associated with poor renal and cardiovascular outcomes and mortality. Because the study of FGF23 in individuals with normal renal function has received little attention, we examined in a large, population based study of 1128 participants the associations of FGF23 with markers of mineral metabolism and renal function. The median estimated glomerular filtration rate (eGFR) of the cohort was 105 ml/min per 1.73 m2, and the median plasma FGF23 was 78.5 RU/ml. FGF23 increased and plasma 1,25-dihydroxyvitamin D3 decreased significantly below an eGFR threshold of 102 and 99 ml/min per 1.73 m2, respectively. In contrast, plasma parathyroid hormone increased continuously with decreasing eGFR and was first significantly elevated at an eGFR of 126 ml/min per 1.73 m2. On multivariable analysis adjusting for sex, age, body mass index, and GFR, FGF23 was negatively associated with 1,25-dihydroxyvitamin D3, and urinary absolute and fractional calcium excretion but not with serum calcium or parathyroid hormone. We found a positive association of FGF23 with plasma phosphate, but no association with urinary absolute or fractional phosphate excretion and, unexpectedly, a positive association with tubular maximum phosphate reabsorption/GFR. Thus, in the absence of CKD, parathyroid hormone increases earlier than FGF23 when the eGFR decreases. The increase in FGF23 occurs at a higher eGFR threshold than previously reported and is closely associated with a decrease in 1,25-dihydroxyvitamin D3. We speculate that the main demonstrable effect of FGF23 in the setting of preserved renal function is suppression of 1,25-dihydroxyvitamin D3 rather than stimulation of renal phosphate excretion.

Environmental Enteropathy and Fibroblast Growth Factor 21 are associated with early childhood growth in Bangladesh

Thesis (Ph.D.)--University of Washington, 2016-06

Vascular endothelial growth factor-B and retinal vascular development in the mouse

Purpose: Vascular endothelial growth factor-A (VEGF-A) is crucial to retinal vascular growth, both normal and pathological. VEGF-B, recently characterized, is reported to be expressed in retinal tissues, but the importance of VEGF-B to retinal vascular development remained unknown. The aim of this study was to analyse retinal vascular growth in the Vegfb (-/-) knockout mouse. Methods: Retinal vascular growth was measured in Vegfb (-/-) knockout mice raised under normal conditions, and Vegfb (-/-) knockout mice with an oxygen-induced proliferative retinopathy. Wild type Vegfb (+/+) mice served as controls. Vessels were perfused with ink and retinal flatmounts secondarily labelled with FITC-lectin (BS-1, Griffonia simplicifolia ). Area and diameter of retinal growth and retinal vascular growth were recorded over days 0-20, and capillary density and mean diameter recorded from day 17 pups. Results: A variety of techniques confirmed that Vegfb (+/+) mice expressed VEGF-B and that VEGF-B expression was absent in Vegfb (-/-) mice. Vegfb (-/-) mice raised in room air showed no significant differences from Vegfb (+/+) controls. No differences were found in oxygen-induced retinopathy between Vegfb (-/-) and Vegfb (+/+) pups in either the extent of the initial oxygen-induced ablation, or in the regrowth of retinal vessels or vitreal (neovascular) sprouts; vitreal sprouts are important markers of the abnormal proliferative response, and are maximally expressed on day 17 in this model of oxygen-induced retinopathy. Conclusions: These results indicate that a lack of VEGF-B does not significantly affect development of the retinal vasculature under normal conditions, nor does it appear to affect the proliferative retinal responses seen in oxygen-induced retinopathy.

CSF-1 as a regulator of macrophage activation and immune responses

Macrophage activation is a key determinant of susceptibility and pathology in a variety of inflammatory diseases. The extent of macrophage activation is tightly regulated by a number of pro-inflammatory cytokines (e.g. IFN-gamma, IL-2, GM-CSF, IL-3) and anti-inflammatory cytokines (e.g. IL-4, IL-10, TGF-beta). Macrophage colony-stimulating factor (CSF-1/M-CSF) is a key differentiation, growth and survival factor for monocytes/macrophages and osteoclasts. The role of this factor in regulating macrophage activation is often overlooked. This review will summarize our current understanding of the effects of CSF-1 on the activation state of mature macrophages and its role in regulating immune responses.

The colony-stimulating factor 1 receptor is expressed on dendritic cells during differentiation and regulates their expansion

The lineage of dendritic cells (DC), and in particular their relationship to monocytes and macrophages, remains obscure. Furthermore, the requirement for the macrophage growth factor CSF-1 during DC homeostasis is unclear. Using a transgenic mouse in which the promoter for the CSF-1R (c-fms) directs the expression of enhanced GFP in cells of the myeloid lineage, we determined that although the c-fms promoter is inactive in DC precursors, it is up-regulated in all DC subsets during differentiation. Furthermore, plasmacytoid DC and all CD11c(high) DC subsets are reduced by 50-70% in CSF-1-deficient osteopetrotic mice, confirming that CSF-1 signaling is required for the optimal differentiation of DC in vivo. These data provide additional evidence that the majority of tissue DC is of myeloid origin during steady state and supports a close relationship between DC and macrophage biology in vivo.

Overexpression of transforming growth factor-beta is associated with increased hyaluronan content and impairment of repair in Marfan syndrome aortic aneurysm

Background-Marfan syndrome (MFS), a condition caused by fibrillin-1 gene mutation is associated with aortic aneurysm that shows elastic lamellae disruption, accumulation of glycosaminoglycans, and vascular smooth muscle cell (VSMC) apoptosis with minimal inflammatory response. We examined aneurysm tissue and cultured cells for expression of transforming growth factor-beta1 to -beta3 (TGF beta 1 to 3), hyaluronan content, apoptosis, markers of cell migration, and infiltration of vascular progenitor cells (CD34). Methods and Results-MFS aortic aneurysm (6 males, 5 females; age 8 to 78 years) and normal aorta (5 males, 3 females; age 22 to 56 years) were used. Immunohistochemistry showed increased expression of TGF beta 1 to 3, hyaluronan, and CD34-positive microcapillaries in MFS aneurysm compared with control. There was increased expression of TGF beta 1 to 3 and hyaluronan in MFS cultured VSMCs, adventitial fibroblasts (AF), and skin fibroblasts (SF). Apoptosis was increased in MFS (VSMC: mean cell loss in MFS 29%, n of subjects = 5, versus control 8%, n = 3, P < 0.05; AF: 28%, n = 5 versus 7%, n = 5, P < 0.05; SF: 29%, n = 3 versus 4%, n = 3, not significant). In MFS, there was a 2-fold increase in adventitial microcapillaries containing CD34-positive cells compared with control tissue. Scratch wound assay showed absence of CD44, MT1-MMP, and beta-3 integrin at the leading edge of migration in MFS indicating altered directional migration. Western blot showed increased expression of TGF beta 1 to 3 in MFS but no change in expression of CD44, MT1-MMP, or beta-3 integrin compared with controls. Conclusions-There was overexpression of TGF-beta in MFS associated with altered hyaluronan synthesis, increased apoptosis, impaired progenitor cell recruitment, and abnormal directional migration. These factors limit tissue repair and are likely to contribute to aneurysm development.

Solution structure and characterization of the LGR8 receptor binding surface of insulin-like peptide 3

Insulin-like peptide 3 (INSL3), a member of the relaxin peptide family, is produced in testicular Leydig cells and ovarian thecal cells. Gene knock-out experiments have identified a key biological role in initiating testes descent during fetal development. Additionally, INSL3 has an important function in mediating male and female germ cell function. These actions are elicited via its recently identified receptor, LGR8, a member of the leucine-rich repeat-containing G-protein- coupled receptor family. To identify the structural features that are responsible for the interaction of INSL3 with its receptor, its solution structure was determined by NMR spectroscopy together with in vitro assays of a series of B-chain alanine-substituted analogs. Synthetic human INSL3 was found to adopt a characteristic relaxin/ insulin-like fold in solution but is a highly dynamic molecule. The four termini of this two-chain peptide are disordered, and additional conformational exchange is evident in the molecular core. Alanine-substituted analogs were used to identify the key residues of INSL3 that are responsible for the interaction with the ectodomain of LGR8. These include Arg(B16) and Val(B19), with His(B12) and Arg(B20) playing a secondary role, as evident from the synergistic effect on the activity in double and triple mutants involving these residues. Together, these amino acids combine with the previously identified critical residue, Trp(B27), to form the receptor binding surface. The current results provide clear direction for the design of novel specific agonists and antagonists of this receptor.

Role of fibroblast growth factor receptor in regulation of membrane traffic

Several studies show that membrane transport mechanisms are regulated by signalling molecules. Recently, genome-wide screen analyses in C.elegans have enabled scientists to identify novel regulators in membrane trafficking and also signalling molecules which are found to couple with this machinery. Fibroblast growth factor (FGF) via binding to fibroblast growth factor receptor (FGFR) mediate signals which are essential in the development of an organism, patterning, cell migration and tissue homeostasis. Impaired FGFR-mediated signalling has been associated with various developmental, neoplastic, metabolic and neurological diseases and cancer. In this study, the potential role of FGFR-mediated signalling pathway as a regulator of membrane trafficking was investigated. The GFP-tagged yolk protein YP170-GFP trafficking was analysed in worms where 1) FGFR signalling cascade components were depleted by RNAi and 2) in mutant animals. From these results, it was found that the disruption of the genes egl-15 (FGFR), egl-17(FGF), let-756(FGF), sem-5, let-60, lin-45, mek-2, mpk-1 and plc-3 lead to abnormal localization of YP170-GFP, suggesting that signalling downstream of FGFR via activation of MAPK and PLC-γ pathway is regulating membrane transport. The route of trafficking was further investigated, to pinpoint which membrane step is regulated by worm FGFR, by analysing a number of GFP-tagged intracellular membrane markers in the intestine of Wild Type (WT) and FGFR mutant worms. FGFR mutant worms showed a significant difference in the localisation of several endosomal membrane markers, suggesting its regulatory role in early and recycling steps of endocytosis. Finally, the trafficking of transferrin in a mammalian NIH/3T3 cell line was investigated to identify the conservation of these membrane trafficking regulatory mechanisms between organisms. Results showed no significant changes in transferrin trafficking upon FGFR stimulation or inhibition.

Experiences of patients with age-related macular degeneration receiving anti-vascular endothelial growth factor therapy:a qualitative study

The societal cost for the average health authority in the United Kingdom for the care of wet age-related macular degeneration (AMD) has been suggested to be around £7.4 million. It is vital that the best possible care based on the best available evidence is provided to reduce the impact of AMD on patients' lives and the financial cost to the health-care system. This study explored the experiences of AMD patients treated with intravitreal ranibizumab injections. Three semistructured interviews were conducted with seven participants over the course of 18 months. Transcripts were analysed using interpretative phenomenological analysis. Analysis identified four themes: preparing for treatment, the treatment process, patient-provider communication, and results of treatment. Patient experiences highlighted the need to move away from the reliance on letters for information provision, and the need for clearer guidelines about when to cease AMD treatment. Interviews highlighted the need for the inclusion of rigorous qualitative evidence with experiential data in future good clinical practice guideline development for AMD. © The Author(s) 2013.

Negative regulation of soluble Flt-1 and soluble endoglin release by heme oxygenase-1

Preeclampsia is characterized clinically by hypertension and proteinuria. Soluble Flt-1 (sFlt-1; also known as soluble vascular endothelial growth factor receptor-1 [VEGFR-1]) and soluble endoglin (sEng) are elevated in preeclampsia, and their administration to pregnant rats elicits preeclampsia-like symptoms. Heme oxygenase-1 (HO-1) and its metabolite carbon monoxide (CO) exert protective effects against oxidative stimuli. Thus, we hypothesized that HO-1 upregulation may offer protection against preeclampsia by inhibiting sFlt-1 and sEng release.

Vascular endothelial growth factor promotes physical wound repair and is anti-apoptotic in primary distal lung epithelial and A549 cells

Objective: There is evidence to suggest a beneficial role for growth factors, including vascular endothelial growth factor (VEGF), in tissue repair and proliferation after injury within the lung. Whether this effect is mediated predominantly by actions on endothelial cells or epithelial cells is unknown. This study tested the hypothesis that VEGF acts as an autocrine trophic factor for human adult alveolar epithelial cells and that under situations of pro-apoptotic stress, VEGF reduces cell death. Design: In vitro cell culture study looking at the effects of 0.03% H2O2 on both A549 and primary distal lung epithelial cells.Measurement and Main Results: Primary adult human distal lung epithelial cells express both the soluble and membrane-associated VEGF isoforms and VEGF receptors 1 and 2. At physiologically relevant doses, soluble VEGF isoforms stimulate wound repair and have a proliferative action. Specific receptor ligands confirmed that this effect was mediated by VEGF receptor 1. In addition to proliferation, we demonstrate that VEGF reduces A549 and distal lung epithelial cell apoptosis when administered after 0.03% H2O2 injury. This effect occurs due to reduced caspase-3 activation and is phosphatidylinositol 3′–kinase dependent. Conclusion: In addition to its known effects on endothelial cells, VEGF acts as a growth and anti-apoptotic factor on alveolar epithelial cells. VEGF treatment may have potential as a rescue therapy for diseases associated with alveolar epithelial damage such as acute respiratory distress syndrome.

Vascular endothelial growth factor-induced endothelial cell proliferation is regulated by interaction between VEGFR-2, SH-PTP1 and eNOS

VEGF receptor-2 plays a critical role in endothelial cell proliferation during angiogenesis. However, regulation of receptor activity remains incompletely explained. Here, we demonstrate that VEGF stimulates microvascular endothelial cell proliferation in a dose-dependent manner with VEGF-induced proliferation being greatest at 5 and 100 ng/ml and significantly reduced at intermediate concentrations (>50% at 20 ng/ml). Neutralization studies confirmed that signaling occurs via VEGFR-2. In a similar fashion, ERK/MAPK is strongly activated in response to VEGF stimulation as demonstrated by its phosphorylation, but with a decrease in phosphoryation at 20 ng/ml VEGF. Immunoblotting analysis revealed that VEGF did not cause a dose-dependent change in expression of VEGFR-2 but instead resulted in reduced phosphorylation of VEGFR-2 when cells were exposed to 10 and 20 ng/ml of VEGF. VEGFR-2 dephosphorylation was associated with an increase in the protein tyrosine phosphatase, SH-PTP1, and endothelial nitric oxide synthase (eNOS). Immunoprecipitation and selective immunoblotting confirmed the association between VEGFR-2 dephosphorylation and the upregulation of SH-PTP1 and eNOS. Transfection of endothelial cells with antisense oligonucleotide against VEGFR-2 completely abolished VEGF-induced proliferation, whereas anti SH-PTP1 dramatically increased VEGF-induced proliferation by 1 and 5-fold at 10 and 200 ng/ml VEGF, respectively. Suppression of eNOS expression only abolished endothelial cell proliferation at VEGF concentrations above 20 ng/ml. Taken together, these results indicate that activation of VEGFR-2 by VEGF enhances SH-PTP1 activity and eNOS expression, which in turn lead to two diverse events: one is that SH-PTP1 dephosphorylates VEGFR-2 and ERK/MAPK, which weaken VEGF mitogenic activity, and the other is that eNOS increases nitric oxide production which in turn lowers SH-PTP1 activity via S-nitrosylation.