We analyzed the protein composition of human aqueous humor. Samples were obtained by paracentesis from 25 human eyes (age range 64-92 years) at elective cataract surgery, and from 20 age-matched post-mortem eyes within 1.5 to 18 hr after death. Individual samples were assayed for total protein, and the polypeptides were separated by qualitative SDS-PAGE into high-, medium- and low-molecular-weight ranges and then silver-stained. The clinical samples showed a remarkable consistency in the total protein values (mean +/- SEM: 12.4 +/- 2.0 mg per 100 ml) and no detectable variations in the profiles of the silver-stained proteins. Twelve major protein fractions, with apparent molecular weights of 140, 80 (doublet), 67, 60 (doublet), 35, 27, 25, <em>17</em>, 14.6 and 9 kDa, were present. A preliminary analysis showed that the <em>17</em> kDa band contained a molecule resembling basic <em>fibroblast</em> <em>growth</em> <em>factor</em>. Two additional samples of aqueous humor from patients whose blood/aqueous barrier was compromised during paracentesis showed a quantitative and qualitative increase in the polypeptides that were present. Compared with the samples of aqueous humor obtained at surgery, the post-mortem samples exhibited a greater variability in total protein content (56.1 +/- 11.6 mg per 100 ml) and an increased number of high- and low-molecular-weight protein fractions. In view of wide differences in the clinical parameters, including ocular and systemic medications, systemic illness, surgical premedications, anesthesia and total serum protein values, the similarity in the protein profiles of the carefully drawn surgical samples is most remarkable. Our results indicate that, in patients who underwent elective cataract surgery, the levels of major proteins in human aqueous humor are not affected by wide individual variations in the clinical parameters. We attribute this finding to the care taken in the collection of aqueous humor samples.

Evidence suggests epithelial-mesenchymal transition (EMT) as one potential source of <em>fibroblasts</em> in idiopathic pulmonary fibrosis. To assess the contribution of alveolar epithelial cell (AEC) EMT to <em>fibroblast</em> accumulation in vivo following lung injury and the influence of extracellular matrix on AEC phenotype in vitro, Nkx2.1-Cre;mT/mG mice were generated in which AECs permanently express green fluorescent protein (GFP). On days <em>17</em>-21 following intratracheal bleomycin administration, ~4% of GFP-positive epithelial-derived cells expressed vimentin or α-smooth muscle actin (α-SMA). Primary AECs from Nkx2.1-Cre;mT/mG mice cultured on laminin-5 or fibronectin maintained an epithelial phenotype. In contrast, on type I collagen, cells of epithelial origin displayed nuclear localization of Smad3, acquired spindle-shaped morphology, expressed α-SMA and phospho-Smad3, consistent with activation of the transforming <em>growth</em> <em>factor</em>-β (TGFβ) signalling pathway and EMT. α-SMA induction and Smad3 nuclear localization were blocked by the TGFβ type I receptor (TβRI, otherwise known as Alk5) inhibitor SB431542, while AEC derived from Nkx2.1-Cre;Alk5(flox/KO) mice did not undergo EMT on collagen, consistent with a requirement for signalling via Alk5 in collagen-induced EMT. Inability of a pan-specific TGFβ neutralizing antibody to inhibit effects of collagen together with absence of active TGFβ in culture supernatants is consistent with TGFβ ligand-independent activation of Smad signalling. These results support the notion that AECs can acquire a mesenchymal phenotype following injury in vivo and implicate type I collagen as a key regulator of EMT in AECs through signalling via Alk5, likely in a TGFβ ligand-independent manner.

We investigated the effects of various culture conditions on the <em>growth</em> of normal human corneal endothelial cells in culture. Falcon Primaria tissue culture plastic was found to provide a more suitable surface for endothelial cell <em>growth</em> than the conventional Corning tissue culture plastic. Also, media containing 10% fetal bovine serum and 5% calf serum (complete media) facilitated the <em>growth</em> of human cells better than those containing Nu-serum. Supplementation with epidermal or <em>fibroblast</em> <em>growth</em> <em>factor</em> (10 and 100 ng/ml) to the complete media had no effect on human endothelial cell <em>growth</em>. Chondroitin sulfate at low concentrations (100 micrograms/ml to 1 mg/ml) also showed little effect. At high concentrations (13.5 and 25 mg/ml), however, chondroitin sulfate significantly promoted human corneal endothelial cell <em>growth</em> during a 1- to 2-week incubation period. From the 37 cultures initiated, out<em>growth</em> from explants appeared within 3 to 7 days. Cells were polygonal in shape and, at confluency, formed a continuous monolayer. We attained a success rate of 87% (7/8) <em>growing</em> primary cultures from donors under 20 years of age and a 59% (<em>17</em>/29) success rate from older donors.

BACKGROUND

Elevated fibroblast growth factor 23 (FGF-23) concentrations associate with left ventricular hypertrophy (LVH) and adverse outcomes in adult patients with chronic kidney disease. We hypothesized that similar associations are present in pediatric patients on maintenance hemodialysis.

METHODS

In this retrospective study of 26 young patients on chronic hemodialysis, aged 6-21 years, cardiac structure and geometry were measured by echocardiography, and circulating levels of FGF-23 and calciotropic hormones were obtained.

RESULTS

FGF-23 levels were uniformly elevated in all patients from three- to 835-fold above the upper limit of normal. The average LV mass index (LVMI) was 43 ± 13 g/m(2.7) and reflected LVH in 55 % of patients. Log-transformed FGF-23 concentrations correlated with LVMI (p = 0.03) and were independently associated with the interventricular septal thickness Z-score (p < 0.001). Concentric LVH was associated with the highest FGF-23 concentrations and the highest LVMI measurements (p < 0.001). Each 1 standard deviation increase in log-transformed FGF-23 levels was associated with a 17 % increase in LVMI.

CONCLUSIONS

FGF-23 levels are strongly associated with increased LVMI and with prevalent LVH in pediatric hemodialysis patients. Our cross-sectional findings provide observational evidence supporting the hypothesis linking FGF-23 to cardiac hypertrophy in patients with chronic kidney disease.

Neonatal skin <em>fibroblasts</em> were cultured in supernatants of peripheral blood monocytes that had been cultured with and without lactoferrin. Granulocyte-monocyte colony-stimulating activity (CSA) was measured in supernatants of the <em>fibroblast</em> cultures with normal T lymphocyte-depleted, phagocyte-depleted, low density bone marrow target cells in colony <em>growth</em> (colony-forming unit granulocyte/macrophage) assays. Monocyte-conditioned medium contained a nondialyzable <em>factor</em> that enhanced by <em>17</em>-50-fold the production of CSA by <em>fibroblasts</em>. The addition of lactoferrin to monocyte cultures reduced the activity of this monokine by 75-100%. Lactoferrin did not inhibit CSA production by monokine-stimulated <em>fibroblasts</em>. We conclude that under appropriate conditions human <em>fibroblasts</em> are potent sources of CSA, that the production of CSA by these cells is regulated by a stimulatory monokine, and that the production and or release of the monokine is inhibited by lactoferrin, a neutrophil-derived putative feedback inhibitor of granulopoiesis. We propose that the major role of mononuclear phagocytes in granulopoiesis is played not by producing CSA, but by recruiting other cells to do so, and that in the steady state, feedback regulation of neutrophil production may occur as a result of a mechanism that inhibits the recruitment phenomenon.

Ets-1 proto-oncogene is a transcription <em>factor</em> involved in several cellular functions, including the activation of several proteases participating in tumor invasion and metastasis. The objective of this study was to analyze the possible correlation between Ets-1 mRNA expression and survival in advanced-stage ovarian carcinomas, studying two patient groups with extremely different disease outcome. Sections from 66 primary ovarian carcinomas and metastatic lesions from 41 patients diagnosed with advanced-stage ovarian carcinoma (International Federation of Gynecologists and Obstetricians stages III and IV) were evaluated for expression of Ets-1 using mRNA in situ hybridization. Patients were divided into long-term (n = <em>17</em>) and short-term (n = 24) survivors. The mean values for disease-free survival and overall survival were 116 and 133 months for long-term survivors, as compared to 3 and 21 months for short-term survivors, respectively. Expression of Ets-1 mRNA was detected in carcinoma cells and stromal cells in 28 of 66 (42%) and 22 of 66 (33%) lesions, respectively. Ets-1 expression showed an association with mRNA expression of vascular endothelial <em>growth</em> <em>factor</em> (P = 0.001 for carcinoma cells; P = 0.004 for stromal cells), basic <em>fibroblast</em> <em>growth</em> <em>factor</em> (P = 0.049 for carcinoma cells), and membrane type-1 matrix metalloproteinase (P = 0.045), which were previously studied in this patient cohort. Ets-1 mRNA was detected more often in both carcinoma and stromal cells in tumors of short-term survivors (P = 0.038 for carcinoma cells). In univariate survival analysis for all cases, Ets-1 expression in both tumor (P = 0.018) and stroma (P = 0.026) correlated with poor survival. These findings were reproduced in an analysis of primary tumors alone (P = 0.039 for tumor cells; P < 0.001 for stromal cells). Ets-1 mRNA expression in stromal cells retained its predictive power in a multivariate survival analysis in which all molecules studied previously in this patient cohort were included (P = 0.007). To our knowledge, this is the first evidence associating Ets-1 mRNA expression and poor survival in human epithelial malignancy. Ets-1 is thus a novel prognostic marker in advanced-stage ovarian carcinoma. The association between Ets-1 mRNA expression and the expression of membrane type-1 matrix metalloproteinase and angiogenic genes, first documented here in a study of patient material, points to the central role of this transcription <em>factor</em> in tumor progression in ovarian carcinoma.

<em>Fibroblast</em> <em>growth</em> <em>factor</em>-2 (FGF-2) and epidermal <em>growth</em> <em>factor</em> (EGF) have both been reported to stimulate precursors in the developing CNS. To clarify these effects, we used clonal analysis to determine the lineage potential of precursors stimulated with each <em>factor</em>, at two stages of development. It was found that in cells isolated from the cerebrum of embryonic day <em>17</em> (E<em>17</em>) mice, FGF-2 stimulated both a multipotential precursor, which gave rise to neurons and astrocytes, and a committed glial precursor. In contrast, EGF only stimulated the glial restricted precursor. Thus, it appears that the multipotential cell, previously identified to be present at E10, remains selectively responsive to FGF-2, and that the EGF responsiveness observed at E<em>17</em> reflects the presence of a new restricted class of precursors, rather than a switch in <em>factor</em> specificity of the multipotential cell.

There is <em>growing</em> evidence for the intracellular role of cytokines and <em>growth</em> <em>factors</em>, but the pathways by which these activities occur remain largely obscure. Previous work from our laboratory identified the constitutive, aberrant expression of the 31-kDa IL-1 alpha precursor (pre-IL-1 alpha) in the nuclei of <em>fibroblasts</em> from the lesional skin of patients with systemic sclerosis (SSc). We established that pre-IL-1 alpha expression was associated with increased <em>fibroblast</em> proliferation and collagen production. Further investigation has led to the identification of a mechanism by which nuclear expression of pre-IL-1 alpha affects <em>fibroblast</em> <em>growth</em> and matrix production. By using a yeast two-hybrid method, we found that pre-IL-1 alpha binds necdin, a nuclear protein with <em>growth</em> suppressor activity. We mapped the region of pre-IL-1 alpha responsible for necdin binding and found it to be localized near the N terminus, a region that is present on pre-IL-1 alpha, but not the mature <em>17</em>-kDa cytokine. Expression studies demonstrated that pre-IL-1 alpha associates with necdin in the nuclei of mammalian cell lines and regulates cell <em>growth</em> and collagen expression. Our results provide the first evidence, to our knowledge, of a nuclear target for pre-IL-1 alpha. Based on these findings, we propose that the constitutively up-regulated expression of pre-IL-1 alpha in the nuclei of SSc <em>fibroblasts</em> up-regulates proliferation and matrix production of SSc <em>fibroblasts</em> through binding necdin, and by counteracting its effects on cell <em>growth</em> and collagen production.

Our previous work demonstrated that both polymorphonuclear leukocytes (PMNs) and protein fractions released from PMNs induced de novo synthesis of <em>fibroblast</em> <em>growth</em> <em>factor</em> 2 (FGF-2), which in turn becomes the direct mediator of endothelial mesenchymal transformation observed in corneal endothelial cells (CECs). To identify the protein <em>factor</em>, we used ProteinChip Array technology. Protein fractions obtained from the conditioned medium released by PMNs were resolved by two-dimensional electrophoresis with immobilized pH gradient strips. Most of the protein spots, with molecular masses of <em>17</em> kDa, were sequentially subjected to in-gel trypsin digestion and mass spectrometry. The <em>17</em>-kDa peptide band was identified as interleukin-1 beta (IL-1 beta). Biological activities of IL-1 beta were further determined; IL-1 beta altered the shape of CECs from polygonal to <em>fibroblast</em>ic morphologies in a time- and dose-dependent manner, whereas neutralizing IL-1 beta antibody, neutralizing antibody to FGF-2, and LY294002 blocked the action of IL-1 beta. IL-1 beta greatly increased the levels of FGF-2 mRNA in a time- and dose-dependent manner; IL-1 beta stimulated expression of all isoforms of FGF-2. IL-1 beta initially induced nuclear accumulation of FGF-2 and facilitated translocation of FGF-2 to plasma membrane and extracellular matrix. IL-1 beta activated phosphatidylinositol (PI) 3-kinase, the enzyme activity of which was greatly stimulated after a 5-min exposure to IL-1 beta. This early and rapid activation of PI 3-kinase greatly enhanced FGF-2 production in CECs; pretreatment with LY294002 hampered the induction activity of IL-1 beta. These observations suggest that IL-1 beta takes part in endothelial to mesenchymal transformation of CECs through its inductive potential on FGF-2 via the action of PI 3-kinase.

We isolated the cDNA encoding a novel member (207 amino acids) of the <em>fibroblast</em> <em>growth</em> <em>factor</em> (FGF) family from rat embryos. Because this protein is the 18th documented member of the FGF family, we tentatively termed it FGF-18. We have also determined mouse and human FGF-18 with high amino acid identity (99.5 and 99.0%) to rat FGF-18, respectively. Among FGF family members, FGF-18 is most similar (52.7% amino acid identity) to FGF-8 and FGF-<em>17</em>. FGF-18 has a typical signal sequence at its amino terminus. Recombinant rat FGF-18, which was efficiently secreted by High Five insect cells infected with recombinant baculovirus containing the cDNA, induced neurite out<em>growth</em> in PC12 cells. The expression of FGF-18 mRNA was examined in adult rat tissues and embryos by Northern blotting analysis and in situ hybridization. FGF-18 mRNA of approximately 2. 7 kilobases was preferentially detected in the lung among adult rat tissues examined. In rat embryos, FGF-18 mRNA was detected in several discrete regions at embryonic days 14.5 and 19.5 but not at E10.5. The temporal and spatial patterns of FGF-18 mRNA expression in embryos are quite different from those of FGF-8 and FGF-<em>17</em> mRNAs reported. The present results indicate that FGF-18 is a unique secreted signaling molecule in the adult lung and developing tissues.

The bovine papillomavirus E5 protein is a 44-amino-acid membrane-associated protein that forms a stable complex with the endogenous platelet-derived <em>growth</em> <em>factor</em> (PDGF) beta receptor in rodent and bovine <em>fibroblasts</em>, resulting in sustained receptor activation and cell transformation. We report here that high-level expression of the E5 protein caused a reduction in the level of the mature form of the PDGF beta receptor in acutely and stably transformed mouse C127 cells. To explore in more detail the interaction of the E5 protein and the PDGF beta receptor, we tested the abilities of various E5 point mutants to bind the PDGF receptor, to induce PDGF receptor down-regulation and tyrosine phosphorylation, and to transform cells. A transformation-competent mutant, like the wild-type E5 protein, bound the receptor and induced receptor tyrosine phosphorylation and down-regulation. Transformation-defective E5 proteins either failed to interact with the endogenous PDGF beta receptor in mouse <em>fibroblasts</em> or underwent an aberrant interaction with the receptor. Mutation of glutamine at position <em>17</em>, aspartic acid at position 33, or both carboxyl-terminal cysteine residues required for E5 homodimerization interfered with stable complex formation with the PDGF receptor, tyrosine phosphorylation and down-regulation of the receptor, and cell transformation. Point mutations at several other carboxyl-terminal positions generated transformation-defective E5 proteins that formed a complex with the PDGF receptor and induced receptor tyrosine phosphorylation but did not induce PDGF receptor down-regulation. Either PDGF receptor activation is not sufficient for transformation of C127 cells or the receptors that are tyrosine phosphorylated in response to these mutant E5 proteins are not fully activated and therefore are not able to deliver a mitogenic signal.

The effects of fluoride (20 mumol/L) and bovine bone extract (<em>17</em> micrograms/ml) were determined on cultures of human bone cells, embryonic chick bone cells, and human skin <em>fibroblasts</em>. The incorporation of [3H]thymidine into DNA was measured 16 hours after the addition of <em>factors</em>. After three to five days treatment, Triton X-100 extracts of the cells were assayed for acid phosphatase activity, in the presence and absence of tartrate, and for alkaline phosphatase activity. Fluoride stimulated [3H]thymidine incorporation and specific activity of alkaline phosphatase in human bone cells and chick bone cells but not in human skin cells. Fluoride also stimulated the cell population doubling rate of the human bone cells with an optimum of approximately 20 mumol/L. Bovine bone extract stimulated thymidine uptake into DNA several-fold and decreased alkaline phosphatase activity in all three types of cultured cells. The specific activity of tartrate-resistant acid phosphatase was increased in bone cells but not in skin <em>fibroblasts</em>. These results suggest that fluoride specifically stimulates the proliferation and differentiation of osteoblasts, while the <em>growth</em> <em>factors</em> in bovine bone extract primarily stimulate proliferation of bone cells. Cultures of human bone cells respond similarly to chick calvarial cells when treated with fluoride or bovine bone extract.

Melanomas depend on autocrine signals for proliferation and survival; however, no systematic screen of known receptor tyrosine kinases (RTKs) has been performed to identify which autocrine signaling pathways are activated in melanoma. Here, we performed a comprehensive analysis of 42 RTKs in six individual human melanoma tumor specimens as well as <em>17</em> melanoma cell lines, some of which were derived from the tumor specimens. We identified five RTKs that were active in almost every one of the melanoma tissue specimens and cell lines, including two previously unreported receptors, insulin-like <em>growth</em> <em>factor</em> receptor 1 (IGF-1R) and macrophage-stimulating protein receptor (MSPR), in addition to three receptors (vascular endothelial <em>growth</em> <em>factor</em> receptor, <em>fibroblast</em> <em>growth</em> <em>factor</em> receptor, and hepatocyte <em>growth</em> <em>factor</em> receptor) known to be autocrine activated in melanoma. We show, by quantitative real time PCR, that all melanoma cell lines expressed genes for the RTK ligands such as HGF, IGF-1, and MSP. Addition of antibodies to either IGF-1 or HGF, but not to MSP, to the culture medium blocked melanoma cell proliferation, and even caused net loss of melanoma cells. Antibody addition deactivated IGF-1R and hepatocyte <em>growth</em> <em>factor</em> receptors, as well as mitogen-activated protein kinase signaling. Thus, IGF-1 is a new <em>growth</em> <em>factor</em> for autocrine driven proliferation of human melanoma in vitro. Our results suggest that IGF-1-IGF-1R autocrine pathway in melanoma is a possible target for therapy in human melanomas.

Proteolytic processing and ectodomain shedding have been described for a broad spectrum of transmembrane proteins under both normal and pathophysiological conditions and has been suggested as one mechanism to regulate a protein's function. It has also been documented for the receptor-like protein tyrosine phosphatase PTP-LAR, induced by treating cells with the tumor promoter TPA or the calcium ionophor A23187. Here we identified the epidermal <em>growth</em> <em>factor</em> receptor (EGFR) as both an association partner of PTP-LAR, that mediates phosphorylation of the latter, as well as an inducer of LAR-cleavage. Both overexpression of this kinase and stimulation of endogenous EGFR in various tumor cell lines were shown to induce proteolytic processing of the catalytic LAR-P-subunit. In contrast to TPA-induced shedding of PTP-LAR, EGFR-mediated cleavage did not require PKC-activity. For both stimuli, however, processing of the P-subunit turned out to be dependent on the activation of the MAP kinases ERK1 and ERK2, and was completely abrogated upon pre-treating cells with Batimastat, indicating the involvement of a metalloproteinase in this pathway. Being strongly impaired in <em>fibroblasts</em> derived from ADAM-<em>17</em>/TACE-knockout-mice or tumor cells that express a dominant negative mutant of ADAM-<em>17</em>/TACE, cleavage of PTP-LAR is suggested to be mediated by this metalloproteinase. Paralleled by rapid reduction of cell surface-localized LAR-E-subunit, EGFR-induced cleavage could be shown to lead to degradation of the catalytic LAR-P-subunit, thereby resulting in a significantly reduced overall cellular phosphatase activity of PTP-LAR. These results for the first time identify a protein tyrosine phosphatase as a potential substrate of TACE and describe proteolytic processing of PTP-LAR as a means of regulating phosphatase activity downstream and thus under the control of EGFR-mediated signaling pathways.

Antley-Bixler syndrome (ABS, MIM 207410) is a skeletal abnormality syndrome primarily affecting head and limbs. Little is known of the origin of the condition but inactivating mutations in the <em>fibroblast</em> <em>growth</em> <em>factor</em> receptor (FGFR2) has been found in some patients. Genital ambiguity is seen occasionally in this condition, suggesting possible disordered steroidogenesis in early pregnancy. We report the steroid excretion of eight patients diagnosed with the syndrome and one with a related condition, a mild phenotype of the disorder since skeletal and genital abnormalities were not evident. The steroid excretion pattern was consistent and very distinctive in all nine patients. Metabolites of the two primary precursors of steroid hormones, pregnenolone and progesterone, were elevated as were the classical diagnostic metabolites for <em>17</em>- and 21-hydroxylase deficiencies. Cortisol production was typically within the normal range but generally had blunted response to ACTH. Androgen metabolite excretion tends to be low in patients over 2 months of age, but may be elevated in the newborn period. The metabolome suggested attenuated steroid hydroxylation (including <em>17</em>,20-lyase activity) although underlying cause is yet to be established. Mutations in CYP<em>17</em> and CYP21 have not been found and currently the prime suspect is an abnormality in an essential redox partner (P450 oxidoreductase). This paper proposes use of the distinctive steroid metabolome as the primary biochemical parameter for diagnosis of ABS, at least the form not associated with FGFR2 mutations.

<em>Fibroblast</em> <em>growth</em> <em>factor</em> (FGF) receptors constitute a family of four membrane-spanning tyrosine kinases (FGFR1-4) which serve as high-affinity receptors for <em>17</em> <em>growth</em> <em>factors</em> (FGF1-<em>17</em>). To study functions of FGF/ FGFR signals in development, mice that carry mutations in each receptor have been created by gene targeting. Analysis of these mutant mice revealed essential functions of FGF receptors in multiple biological processes, including mesoderm induction and patterning, cell <em>growth</em> and migration, organ formation and bone <em>growth</em>. In this review we discuss recent work with FGF receptors to illustrate mechanisms, through which the FGF/FGFR signals specify vertebrate limb initiation, out<em>growth</em> and patterning.

<em>Growth</em> <em>factors</em> and morphogens need to be secreted to act on distant cells during development and in response to injury. Here, we report evidence that efficient export of a <em>fibroblast</em> <em>growth</em> <em>factor</em> (FGF), EGL-<em>17</em>, from the Caenorhabditis elegans developing vulva requires the lipoprotein receptor-related proteins Ce-LRP-1 and Ce-LRP-2 and a cytoplasmic adaptor protein, Ce-DAB-1 (Disabled). Lipoprotein receptors are transmembrane proteins best known for their roles in endocytosis. Ce-LRP-1 and Ce-LRP-2 possess a conserved intraluminal domain that can bind to EGL-<em>17</em>, as well as a cytosolic FXNPXY motif that can bind to Ce-DAB-1. Ce-DAB-1 contains signals that confer subcellular localization to Golgi-proximal vesicles. These results suggest a model in which Ce-DAB-1 coordinates selection of receptors and cargo, including EGL-<em>17</em>, for transport through the secretory pathway.

In rat osteosarcoma (ROS <em>17</em>/2.8) cells, which express osteoblastic features in culture, basic <em>fibroblast</em> <em>growth</em> <em>factor</em> (bFGF) reduces the level of alkaline phosphatase, type I collagen, and osteocalcin mRNA and increases osteopontin mRNA, independent of <em>growth</em> stimulation. The <em>fibroblast</em> <em>growth</em> <em>factor</em> (FGF) effects are dose dependent (EC50 about 6 pM) and are detected 24 h after addition of the <em>growth</em> <em>factor</em>. bFGF also reduces parathyroid hormone-stimulatable adenylate cyclase and alkaline phosphatase activity in these cells. Concomitant treatment with pertussis toxin (20 ng/ml) opposes the FGF effects. Although cyclic AMP elevating agents mimic pertussis toxin action on some parameters, they produce opposite effects on others, indicating that antagonism between pertussis toxin and bFGF is not mediated by cyclic AMP. bFGF caused a small reduction in steady state NAD-dependent ADP-ribosylation and had no detectable effects on the steady-state levels of the Gi alpha (alpha subunit of the inhibitory G protein) 1, 2, and 3, visualized with specific antibodies in these cells. Although the site of interaction of pertussis toxin and FGF remains to be determined, the findings presented here suggest separate control of <em>growth</em> and differentiation by bFGF and show that pertussis toxin treatment can modulate differentiation in these cells, presumably via Gi proteins.

OBJECTIVE

Moyamoya disease (MMD) is a rare cerebrovascular disorder involving stenosis of the major vessels of the circle of Willis and proximal portions of its principal branches. Despite concerted investigation, the pathophysiology of the disorder has not been fully elucidated. Currently, the major proteins believed to play an active role in the pathogenesis include vascular endothelial <em>growth</em> <em>factor</em> (VEGF), basic <em>fibroblast</em> <em>growth</em> <em>factor</em> (bFGF), hepatocyte <em>growth</em> <em>factor</em> (HGF), transforming <em>growth</em> <em>factor</em>-β₁ (TGFβ₁), and granulocyte colony-stimulating <em>factor</em> (G-CSF). In terms of the genetics, recent literature suggests a low penetrance autosomal dominant or polygenic mode of transmission involving chromosomes 3, 6, 8, 12, and <em>17</em> for familial MMD. This review summarizes the current knowledge on the histopathology, pathophysiology and genetics of MMD.

METHODS

A PubMed/Medline systematic study of the literature was performed, from which 45 articles regarding MMD pathophysiology were identified and analyzed.

CONCLUSIONS

Moyamoya disease is characterized by the intimal thickening and media attenuation of the proximal vessels of the circle of Willis as well as the development of an aberrant distal vascular network. The primary proteins that are currently implicated in the pathophysiology of MMD include VEGF, bFGF, HGF, TGFβ₁, and G-CSF. Furthermore, the current literature on familial MMD has pointed to a low penetrance autosomal dominant or polygenic mode of transmittance at loci on chromosomes 3, 6, 8, 12, and <em>17</em>.

Rheumatoid arthritis (RA) is a chronic inflammatory disease characterized by synovial <em>fibroblast</em> hyperplasia and bone and cartilage erosion. Synovial <em>fibroblast</em>- and T cell-mediated inflammation plays crucial roles in the pathogenesis of RA. However how this inflammation is initiated, propagated, and maintained remains controversial. Here, we systemically examined the contribution of toll-like receptors (TLRs) to the inflammatory mediator production as well as Th1 and Th<em>17</em> cell hyperactivity in RA. Our results show that rheumatoid arthritis synovial <em>fibroblasts</em> (RASF) express a series of TLRs, including TLR2, TLR3, TLR4, and TLR9, with the predominant expression of TLR3. Moreover, the expression levels of these TLRs were higher than those in osteoarthritis synovial <em>fibroblasts</em> (OASF). Ligation of TLR3, as well as TLR2 and TLR4, resulted in vigorous production of inflammatory cytokines, matrix metalloproteinases (MMPs), and vascular endothelial <em>growth</em> <em>factor</em> (VEGF) in RASF, with activation of the NF-κB, MAPK, and IRF3 pathways. More important, activation of these TLRs expressed by RASF exacerbated inflammatory Th1 and Th<em>17</em> cell expansion both in cell-cell contact-dependent and inflammatory cytokine-dependent manners, which induced more IFN-γ and IL-<em>17</em> accumulation. Targeting TLRs may modulate the inflammation in RA and provide new therapeutic strategies for overcoming this persistent disease.

OBJECTIVE

To determine whether primary drug resistance or rapid relapse explains the poor prognosis seen in t(4;14)-positive multiple myeloma (MM).

METHODS

A total of 131 patients treated with high-dose therapy (HDT) were assessed, of whom 19 were t(4;14) positive. We examined the presentation features, chemotherapy responsiveness at presentation and to salvage therapies at relapse, and overall survival outcomes.

RESULTS

t(4;14)-positive patients had a predominance of the immunoglobulin A isotype (52.6%) but otherwise baseline characteristics were indistinguishable. After treatment with vincristine, doxorubicin, and dexamethasone or dexamethasone alone, <em>17</em> (89.7%) of the 19 patients achieved a partial response and 11 patients (57.9%) demonstrated an additional 50% reduction in paraprotein after HDT. Thus, t(4;14)-positive patients are chemotherapy sensitive; however, early progression was common, with 26% of patients progressing before HDT and a median progression-free survival after HDT of only 14.1 months. At relapse, a resistance to alkylating agents was evident, with no responses (zero of 11 patients) seen with conventional-dose alkylating agents. Salvage regimens using thalidomide and/or dexamethasone achieved at least minimal response in 59% of patients. The duration of response was short, however, with a median of only 4.7 months. The median overall survival after HDT was 24.2 months.

CONCLUSIONS

We conclude that t(4;14)-positive MM is chemotherapy sensitive but rapid relapse occurs. Resistance to alkylating agents is evident at relapse. The development of novel therapeutic agents is required, including the early clinical study of targeted fibroblast growth factor receptor 3 tyrosine kinase inhibitors, which have shown promise in preclinical studies.

Background <em>Fibroblast</em> <em>growth</em> <em>factor</em>-23 (FGF-23) has been hypothesized to play a role in the increased risk of cardiovascular disease in patients with CKD.Methods We identified prospective studies reporting associations between FGF-23 concentration and risk of cardiovascular events. Maximally adjusted risk ratios (RRs) were extracted for each outcome and scaled to a comparison of the top versus bottom third of the baseline FGF-23 concentration, and the results aggregated.Results Depending on the assay used, median FGF-23 concentrations were 43-74 RU/ml and 38-47 pg/ml in <em>17</em> general population cohorts; 102-392 RU/ml in nine cohorts of patients with CKD not requiring dialysis; and 79-4212 RU/ml and 2526-5555 pg/ml in eight cohorts of patients on dialysis. Overall, comparing participants in the top and bottom FGF-23 concentration thirds, the summary RRs (95% confidence intervals [95% CIs]) were 1.33 (1.12 to 1.58) for myocardial infarction, 1.26 (1.13 to 1.41) for stroke, 1.48 (1.29 to 1.69) for heart failure, 1.42 (1.27 to 1.60) for cardiovascular mortality, and 1.70 (1.52 to 1.91) for all-cause mortality. The summary RR for noncardiovascular mortality, calculated indirectly, was 1.52 (95% CI, 1.28 to 1.79). When studies were ordered by average differences in FGF-23 concentration between the top and bottom thirds, there was no trend in RRs across the studies.Conclusions The similarly-sized associations between increased FGF-23 concentration and cardiovascular (atherosclerotic and nonatherosclerotic) and noncardiovascular outcomes, together with the absence of any exposure-response relationship, suggest that the relationship between FGF-23 and cardiovascular disease risk may be noncausal.

OBJECTIVE

Fibroblast growth factor 19 (FGF19) is a hormone released from the small intestine; recently, it has emerged as an endocrine regulator of glucose and lipid metabolism. The aim of this study was to investigate the role of FGF19 in the development of nonalcoholic fatty liver disease (NAFLD).

METHODS

This study included 23 (17 boys) obese adolescents (mean age of 14.1 years) with NAFLD. The control group consisted of 34 (13 boys) obese peers with normal ultrasonographic imaging and normal liver function tests.

METHODS

The definition of NAFLD was based on clinical criteria: elevated alanine aminotransferase (>35 U/L) and liver steatosis features on ultrasound imaging. Serum FGF19 levels were measured in a fasting blood sample. The definition of insulin resistance was based on the homeostasis model assessment (HOMA) threshold: >2.5.

RESULTS

There was a significant difference between mean FGF19 levels in patients with NAFLD and controls (142.2 vs. 206 pg/mL, p=0.04). Mean fasting FGF19 levels were decreased in insulin-resistant patients in comparison with the non-insulin-resistant group (155.0 vs. 221.0 pg/mL, p=0.05). There was an inverse correlation between FGF19 and alanine aminotransferase levels (R=-0.3, p<0.05) and triglycerides (R=-0.27, p<0.05).

CONCLUSIONS

A decrease in fasting FGF19 is associated with the development of NAFLD in obese adolescents. A decrease in fasting FGF19 levels may be a new important risk factor for NAFLD and the metabolic syndrome in adolescents. Further studies are needed to explain whether exogenous delivery of FGF19 might be therapeutically beneficial.

Many implanted devices fail due to the formation of an avascular capsule surrounding the device. Additionally, fat has long been known to promote healing and vascularization. The goals of this study were to identify potential mechanisms of the provascular actions of adipose-derived stromal cells (ASCs) and to improve implant biocompatibility. First, adult ASCs and <em>fibroblasts</em> from rats were attached to polyurethane and polystyrene in vitro and their cytokine secretion profile was analyzed. Secretion of vascular endothelial <em>growth</em> <em>factor</em> (VEGF) from ASCs was 10-70 times higher than <em>fibroblasts</em> after 3 and 6 days. Next, polyurethane, bare and with cellular coatings, was implanted subcutaneously in rats. The fibrous capsule surrounding bare polyurethane implants was <em>17</em>%-32% thicker and the amount of collagen was 27% greater than the capsule surrounding ASC-coated implants. Finally, the microvessel density adjacent to ASC-coated polyurethane was approximately 50%-80% higher than bare polyurethane. In summary, ASCs attached to polyurethane have a dramatically increased VEGF production compared with <em>fibroblasts</em> in vitro, and these cells also produce an increased microvessel density in the surrounding tissue when implanted subcutaneously in rats. Disclosure of potential conflicts of interest is found at the end of this article.