Fibroblast Growth Factor Immunoreactivity Research Articles

Distribution of fibroblast growth factor in cultured dorsal root ganglion neurons and Schwann cells. I. Localization during maturation in vitro.

The distribution of acidic and basic fibroblast growth factor in co-cultures of dorsal root ganglion neurons and Schwann cells was examined as a function of time in culture. After two days in vitro, the cytoplasm of the neuronal cell bodies demonstrated both acidic and basic FGF immunoreactivity, whereas the cytoplasm of the neurites was not immunoreactive. Schwann cells, in contrast, exhibited both acidic and basic fibroblast growth factor cytoplasmic immunoreactivity. After two days in culture, immunoreactivity was not detected on the plasma membrane surface of either the neurons or the Schwann cells. By 10 days in vitro, fibroblast growth factor immunoreactivity was observed in the cytoplasm of the most proximal portion of some, but not all, neurites but was unchanged in Schwann cells. At 20 days in vitro, immunoreactivity was still restricted to the intracellular compartment of both Schwann cells and neurons. Acidic fibroblast growth factor was primarily localized to the cytoplasm of Schwann cells, neuron cell bodies and along the entire length of the neurites. In contrast, basic fibroblast growth factor was predominantly localized to the nuclei of Schwann cells and small to medium size neurons. In many cases, the nucleolar region demonstrated the most intense basic fibroblast growth factor. The cytoplasm of the neurites was also immunoreactive for basic fibroblast growth factor. At 30 days in vitro the intracellular distribution of fibroblast growth factor immunoreactivity was similar to that observed at 20 days. However, both acidic and basic fibroblast growth factor were detected on the surface of the neurites. In contrast, no fibroblast growth factor immunoreactivity was detected at the Schwann cell surface at any time point examined. The distribution of fibroblast growth factor in Schwann cells cultured by themselves was similar to that of Schwann cells co-cultured with neurons after 20 days in vitro. Both Schwann cells and dorsal root ganglia exhibited increased fibroblast growth factor immunoreactivity with increased time in culture and an increased expression of basic fibroblast growth factor in the nucleus. Of particular interest was the appearance of fibroblast growth factor on the surface of neurites after 30 days in vitro where it could function to modulate neuron-glial cell interactions.

Corticosterone increases FGF-2 (bFGF) immunoreactivity in the substantia nigra of the rat.

The effects of acute and subchronic (7 days) administrations of the adrenocortical hormone corticosterone on basic fibroblast growth factor (bFGF, FGF-2) immunoreactivity were studied in the substantia nigra of the rat by semiquantitative immunocytochemistry coupled with image analysis. Corticosterone was able to increase FGF-2 immunoreactivity in different nigral subregions and cell types (astrocytes and neurones) depending on the duration of the treatment. These results open up the possibility that stress hormones can modulate the trophic state of the substantia nigra through an action on FGF-2.

Immunohistochemical localization of basic fibroblast growth factor in cultured rat astrocytes and oligodendrocytes

The distribution of basic fibroblast growth factor in cultured astrocytes and oligodendrocytes was examined using immunocytochemistry. The results demonstrate a localization of basic fibroblast growth factor immunoreactivity predominantly in astrocyte nuclei at all stages of differentiation. Cytoplasmic and process staining was best detected during early stages of differentiation, under normal growth conditions or as a result of treatment with dibutyryl cyclic adenosine monophosphate. Astrocytes at all stages of differentiation bound antibody-complexed bFGF, suggesting the presence of cell-associated low affinity binding sites for the growth factor. Our studies also show the presence of immunoreactivity for basic fibroblast growth factor in process-bearing oligodendrocytes. These results suggest a role for endogenous basic fibroblast growth factor in astrocyte and oligodendrocyte growth and function.

Altered Acidic and Basic Fibroblast Growth Factor Expression Following Spinal Cord Injury

In normal spinal cord, acidic fibroblast growth factor (aFGF) immunoreactivity was localized in the cytoplasm of ventral motor neurons and sensory fibers in the dorsal columns. Basic FGF (bFGF) immunoreactivity was restricted to astrocyte nuclei and the cytoplasm of a few neurons in the intermediate gray matter. Spinal cord lesions resulted in complete destruction of the dorsal columns at T8. Two days postlesion, aFGF immunoreactivity was increased in ventral motor neurons and was now seen in intermediate gray matter neurons. Acidic FGP was not detected in the lesioned fasciculus gracilis at T4-5, but markedly increased in the fasciculus cuneatus. At L1-2, aFGF-immunoreactive fibers in the fasciculus gracilis also increased. This aFGF immunostaining was maintained 5 and 12 days postlesion. A lesion-induced loss of aFGF immunoreactivity in the nucleus gracilis suggests that aFGF is anterogradely transported in ascending sensory fibers. Two days postlesion, glial fibrillary acidic protein immunoreactivity increased at the lesion site, as well as at T4-5 and L1-2, with no change in bFGF staining. Five days postlesion, increased bFGF immunoreactivity appeared at the edge of the cystic cavity and the dorsal columns at T4-5 in both the nucleus and the cytoplasm of reactive astrocytes, and was increased at 12 days postlesion. The differential cellular, temporal, and spatial expression of aFGF and bFGF following spinal cord lesion suggest they subserve distinct roles in the response to CNS injury.

Basic fibroblast growth factor: an autocrine mitogen of rat thyroid follicular cells?

Basic fibroblast growth factor (FGF) is a mitogen for the rat thyroid cell line FRTL-5. A possible autocrine role for this growth factor has been investigated in rat thyroid follicular cells both in vitro and in vivo. We report here the synthesis and localisation of basic FGF and one of its high affinity receptors (flg) in FRTL-5 cells, shown by Northern hybridization analysis, Western blotting, and immunohistochemistry. Two major species of basic FGF mRNA of approximately 2.2 and 7.0 kilobases and one major species of flg mRNA of approximately 4.2 kilobases were identified in FRTL-5 cells. The basic FGF immunoreactivity observed histologically was attributed to a heparin-binding protein of approximately 20 kilodaltons mol wt. The physiological relevance of basic FGF to the thyroid is underlined by the demonstration of significant stores of immunoreactive protein, predominantly in the basement membrane of thyroid follicular cells, in paraffin sections of the normal rat thyroid, although basic FGF mRNA was not detected by in situ or Northern hybridization analysis. The mitogenic response of FRTL-5 cells to human recombinant basic FGF has been further characterized, and the factor shown to stimulate with an ED50 of 4 ng/ml. The mitogenic effects of exogenously supplied and endogenously produced basic FGF were shown to be potentiated by heparin. Examination of the mitogenic activity of both exogenous and endogenous basic FGF and its immunoneutralization in vitro suggests that locally produced basic FGF may be an important autocrine regulator of thyroid follicular cell growth.

Basic fibroblast growth factor: an autocrine mitogen of rat thyroid follicular cells?

Basic fibroblast growth factor (FGF) is a mitogen for the rat thyroid cell line FRTL-5. A possible autocrine role for this growth factor has been investigated in rat thyroid follicular cells both in vitro and in vivo. We report here the synthesis and localisation of basic FGF and one of its high affinity receptors (flg) in FRTL-5 cells, shown by Northern hybridization analysis, Western blotting, and immunohistochemistry. Two major species of basic FGF mRNA of approximately 2.2 and 7.0 kilobases and one major species of flg mRNA of approximately 4.2 kilobases were identified in FRTL-5 cells. The basic FGF immunoreactivity observed histologically was attributed to a heparin-binding protein of approximately 20 kilodaltons mol wt. The physiological relevance of basic FGF to the thyroid is underlined by the demonstration of significant stores of immunoreactive protein, predominantly in the basement membrane of thyroid follicular cells, in paraffin sections of the normal rat thyroid, although basic FGF mRNA was not detected by in situ or Northern hybridization analysis. The mitogenic response of FRTL-5 cells to human recombinant basic FGF has been further characterized, and the factor shown to stimulate with an ED50 of 4 ng/ml. The mitogenic effects of exogenously supplied and endogenously produced basic FGF were shown to be potentiated by heparin. Examination of the mitogenic activity of both exogenous and endogenous basic FGF and its immunoneutralization in vitro suggests that locally produced basic FGF may be an important autocrine regulator of thyroid follicular cell growth.

Increase of basic fibroblast growth factor immunoreactivity and its mRNA level in rat brain following transient forebrain ischemia.

We examined the time course of basic fibroblast growth factor (bFGF) immunoreactivity and its mRNA level mainly in the hippocampus after transient forebrain ischemia using immunohistochemistry, enzyme immunoassay (EIA), Western blot analysis and in situ hybridization. Neuronal death in the hippocampal CA1 subfield was observed 72 h after 20 min of ischemia. The number of bFGF-immunoreactive(IR) cells increased 48 h-5 days after ischemia in all hippocampal regions. At 10 and 30 days, the bFGF-IR cells in the CA1 subfield had further increased in numbers and altered their morphology, enlarging and turning into typical reactive astrocytes with the advancing neuronal death in that area. In contrast, the number of bFGF-IR cells in other hippocampal regions had decreased 30 days after ischemia. The EIA study showed a drastic increase in bFGF levels in the hippocampus 48 h after ischemia (150% of that in normal rat) which was followed by further increases. In Western blot analysis, three immunoreactive bands whose molecular weights correspond to 18, 22 and 24 kDa were observed in normal rat and ischemia increased all their immunoreactivities. In the in situ hybridization study of the hippocampus, bFGF mRNA positive cells were observed in the CA1 subfield in which many bFGF-IR cells existed after ischemia. These data demonstrate that transient forebrain ischemia leads to an early and strong induction of bFGF synthesis in astrocytes, suggesting that the role of bFGF is related to the function of the reactive astrocytes which appear following brain injury.

Never ending growth and a growth factor. I. Immunocytochemical evidence for the presence of basic fibroblast growth factor in a tapeworm.

Basic fibroblast growth factor immunoreactivity (bFGF-IR) was detected in a specific set of nerve cells in the central nervous system of the gull-tapeworm Diphyllobothrium dendriticum (Cestoda)--an obligatory endoparasite. The long varicose processes of the bFGF-IR neurons extend along the two main nerve cords, which contain an actively growing population of nerve cells. A neurotrophic function for the bFGF-like material is proposed. The adult tapeworm lives in the intestine of a homeothermic host and has a pattern of very active and never ending growth. The larval stages live in poikilothermic hosts and grow very slowly. The bFGF-IR nerve cells occur both in II stage larvae and in adult worms. Thus, no correlation between the presence of bFGF-IR and the general growth rate of the worm was found. Western blot analysis revealed the presence of an anti-bFGF immunoreactive peptide with a molecular mass of 47 kDa in both larval and adult worms.

Appearance of Basic Fibroblast Growth Factor Immunoreactivity in Astrocytes at the Site of Cerebral Lesions and Edematous Areas in SHRSP

Appearance of Basic Fibroblast Growth Factor Immunoreactivity in Astrocytes at the Site of Cerebral Lesions and Edematous Areas in SHRSP

Distribution of Fibroblast Growth Factor Immunoreactivity in Skin during Wool Follicle Development

Distribution of Fibroblast Growth Factor Immunoreactivity in Skin during Wool Follicle Development

Basic FGF is present in dopaminergic neurons of the ventral midbrain of the rat

Immunocytochemistry procedures and 6-hydroxy-dopamine-induced degeneration of dopamine nerve cells provided evidence that practically all tyrosine hydroxylase immunoreactive (IR) neurons of the substantia nigra and ventral tegmental area contain cytoplasmic basic fibroblast growth factor immunoreactivity (bFGF IR), while many astroglial cells in the neostriatum and substantia nigra contain nuclear bFGF IR. RNA blot analysis demonstrated strong expression of a major 3.7 kb bFGF mRNA in the substantia nigra and ventral tegmental area. These results indicate that bFGF may be a significant growth factor in the DA neurons.

Possible roles of basic fibroblast growth factor in the pathogenesis of moyamoya disease: an immunohistochemical study

Prominent features of moyamoya disease are fibrocellular thickening of the intima and enhanced angiogenesis. The pathogenesis of moyamoya disease is, however, unknown. Basic fibroblast growth factor (FGF) is an angiogenic factor as well as a potent mitogen for a number of cell types including vascular endothelial and smooth-muscle cells. In order to test the possibility that basic FGF takes part in the pathogenesis of moyamoya disease, the authors tested for the presence of this factor using a mouse monoclonal antibody against human recombinant basic FGF. The surgical specimens studied included two sections of the superficial temporal artery (STA) and four samples of dura mater from four patients with moyamoya disease. Surgical specimens were obtained from three patients with other diseases as control tissue. Sections of the STA obtained from the patients with moyamoya disease showed strong basic FGF immunoreactivity in endothelial and smooth-muscle cells, while control sections had only faint and scattered immunoreactivity. All sections of the dura mater obtained from the patients with moyamoya disease also revealed more intense immunohistochemical staining of basic FGF in meningeal and vascular cells than did control sections. These observations indicate that the amount of basic FGF is increased in the tissues of patients with moyamoya disease; thus, basic FGF may play an important role in the pathogenesis of moyamoya disease.

Internalization and processing of basic fibroblast growth factor by neurons and astrocytes

The fate of iodinated basic fibroblast growth factor (FGF) after its binding to cultured astrocytes and hippocampal neurons was studied. Autoradiography after light and electron microscopy establishes that, if cells are returned to 37 degrees C, the 125I-basic FGF bound internalizes into vesicles in the cytoplasm, localizes to the perinuclear cytoplasm, and is translocated to chromatin structures of the nucleus. The radiolabeled protein is long-lived, a finding confirmed by biochemical analyses. Polyacrylamide gel electrophoresis and autoradiography of both hippocampal neurons and astrocyte extracts reveal that these cells internalize 125I-basic FGF and then metabolize it to three major heparin-binding peptides with molecular weights of 15.5, 9, and 4 kDa. These peptides are initially detected 16 hr after binding to neurons and 4 hr after binding to astrocytes but are still detectable 48 and 16 hr, respectively, after initial binding (though present at lower levels). Immunoprecipitation with sequence-specific antisera to basic FGF reveals that the 15.5-kDa fragment is generated by cleavage at the carboxyl terminus, that the 9-kDa peptide contains the sequences between residues 30 and 87, and the 4-kDa peptide is a C-terminus fragment containing the sequence of basic FGF(106-120) but without basic FGF(139-146) immunoreactivity. The internalization of basic FGF is required for this processing; the treatment of cells with trypsin and 2 M NaCl at different times after binding can only prevent the metabolism of basic FGF if it is performed immediately after binding. Similarly, WGA, which inhibits basic FGF binding to its high-affinity receptor, prevents the metabolism of basic FGF. The possible significance of a metabolic pathway that is responsible for the processing of basic FGF after its internalization by cells in the CNS is discussed in light of its potential function as a neurotrophic factor.

REPLY FROM R. WESTERMANN ET AL.

Journal of NeurochemistryVolume 56, Issue 3 p. 1088-1088 Free Access REPLY FROM R. WESTERMANN ET AL. Reiner Westermann, Reiner Westermann Department of Anatomy and Cell Biology University of Marburg Marburg, F.R.G.Search for more papers by this authorKlaus Unsicker, Klaus Unsicker Department of Anatomy and Cell Biology University of Marburg Marburg, F.R.G.Search for more papers by this authorClaudia Grothe, Claudia Grothe Department of Anatomy and Cell Biology University of Marburg Marburg, F.R.G.Search for more papers by this author Reiner Westermann, Reiner Westermann Department of Anatomy and Cell Biology University of Marburg Marburg, F.R.G.Search for more papers by this authorKlaus Unsicker, Klaus Unsicker Department of Anatomy and Cell Biology University of Marburg Marburg, F.R.G.Search for more papers by this authorClaudia Grothe, Claudia Grothe Department of Anatomy and Cell Biology University of Marburg Marburg, F.R.G.Search for more papers by this author First published: March 1991 https://doi.org/10.1111/j.1471-4159.1991.tb02035.xAboutPDF ToolsRequest permissionExport citationAdd to favoritesTrack citation ShareShare Give accessShare full text accessShare full-text accessPlease review our Terms and Conditions of Use and check box below to share full-text version of article.I have read and accept the Wiley Online Library Terms and Conditions of UseShareable LinkUse the link below to share a full-text version of this article with your friends and colleagues. Learn more.Copy URL No abstract is available for this article. REFERENCES Westermann R., Stögbauer F., Unsicker K., and Lietze R. (1988) Calcium-dependence of chromogranin A-catecholamine interaction. FEBS Lett. 239, 203– 206. Westermann R., Johannsen M., Unsicker K., and Grothe C. (1990) Basic fibroblast growth factor (bFGF) immunoreactivity is present in chromaffin granules. J. Neurochem. 55, 285– 292. Winkler H. and Smith A. D. (1975) The chromaffin granule and the storage of catecholamines. Handbook of Physiol, Sec. 7: Endocrinology, Vol. VI ( R. O. Creep and E. B. Astwood eds), pp. 321– 339. Williams & Wilkins, Baltimore . Volume56, Issue3March 1991Pages 1088-1088 ReferencesRelatedInformation

Identification and Characterization of Basic Fibroblast Growth Factor in Porcine Thyroids*

Fibroblast growth factor (FGF) is one of the most potent endothelial cell growth factors and has been found in almost all tissues in the body. However, there is no definitive report showing that FGF exists in the thyroid. In this report, we describe heparin binding endothelial cell growth factor activity in porcine thyroids. The extract from normal adult porcine thyroids was loaded onto a heparin-sepharose affinity column. The mitogenic activity on endothelial cells was found to elute from the column with 0.9 M-2.0 M NaCl. Polyacrylamide gel electrophoresis and an immunoblotting of bioactive fractions showed the basic FGF immunoreactivity in a double band with a molecular weight of 18K and 20K. These results strongly imply that the adult porcine thyroid-derived heparin binding endothelial cell growth factor may be authentic basic FGF and one of its high molecular weight forms. In support of this conclusion, basic FGF mRNA was detected in poly A+ RNA isolated from cultured porcine thyroid cells. On the other hand, recombinant human basic FGF at the concentration of 0.1-10 ng/ml increased the incorporation of 3H-thymidine into FRTL-5 cells and porcine thyroid follicular cells in culture. The stimulatory effects were also observed when the biologically active fractions of heparin-sepharose column of the thyroid extract were added to the culture. In addition, both recombinant human basic FGF at 10 ng/ml and heparin binding fractions from porcine thyroids inhibited TSH-induced iodide uptake by porcine thyroid cells in culture. These results suggest that basic FGF may be one of the important local modulators of thyroid function, cell growth, and angiogenesis.

Immunocytochemical mapping of basic fibroblast growth factor in the developing and adult rat adrenal gland

We studied the spatial and temporal pattern of basic fibroblast growth factor (bFGF) immunoreactivity in the rat adrenal gland during postnatal development. In the cortex the glomerulosa zone reveals a strong anti-bFGF immunoreactivity at all developmental ages studied. In the fasciculata zone the high number of anti-bFGF immunoreactive cells in the first week decreases during the second and third week. The late developing reticularis zone shows only few anti-bFGF labeled cells at all postnatal ages. This distributional pattern of bFGF immunoreactivity matches that of mitotic activity in the rat adrenal cortex strengthening the role of bFGF as an autocrine growth factor for adrenocortical cells. In the medulla anti-bFGF positive chromaffin cells become detectable at postnatal day (P) 8 and increase in number during the second and third week. In the adult rat the staining intensity of the chromaffin cells was higher than at P18. In the adult medulla bFGF colocalizes with noradrenaline suggesting its presence in a chromaffin cell subpopulation. In accordance with previous results the role of the chromaffin cell bFGF as a neurotrophic factor for preganglionic sympathetic neurons is discussed.

Immunocytochemical localization of basic fibroblast growth factor in bovine adrenal gland, ovary, and pituitary.

We studied the distribution of basic fibroblast growth factor (bFGF) immunoreactivity in bovine adrenal gland, ovary, and pituitary, using a polyclonal anti-bFGF antibody. In the adrenal gland, the inner layers of the capsule, the zona glomerulosa of the cortex, and the chromaffin cells of the adrenal medulla were intensely stained. In the ovary, follicular epithelial cells of growing follicles and granulosa cells of mature follicles showed strong bFGF-like immunoreactivity. Endocrine cells of the pituitary anterior and intermediate lobes displayed a positive immunoreaction. Blood vessels, including endothelial and smooth muscle cells, as well as stromal cells in all three organs studied, were not stained. This distribution pattern of bFGF immunoreactivity is only partially compatible with the established mitogenic role of this protein, and suggests a wider spectrum of bFGF functions.

Increased basic fibroblast growth factor (bFGF) immunoreactivity at the site of focal brain wounds

Basic fibroblast growth factor (bFGF) is a polypeptide found within the CNS with potent effects on the survival and proliferation of CNS glia and endothelial cells, and on the survival and outgrowth of CNS neurons. Immunohistochemical methods were used to examine relative changes in the levels and distribution of bFGF following focal brain injury. Two monospecific antisera to bFGF were used to immunostain intact mature rat brain, and brain in which a focal mechanical lesion had been made in the dorsolateral cerebral cortex one week previously. In the intact brain, staining was localized primarily in neuronal cell bodies, especially in limbic structures. In injured brain, a marked increase of bFGF immunoreactivity was found at the borders of lesions, localized to the dense accumulation of cells, many of which resembled ‘reactive’ astroglia. Such increases in local bFGF concentrations may contribute to the cascade of cellular changes — including glial and capillary proliferation, and neural sprouting — that follows focal brain injury.

Immunocytochemical demonstration of fibroblast growth factor in cultured chick and rat neurons.

The presence of fibroblast growth factor (FGF) was investigated by immunocytochemistry in cultured neuronal cells derived from the peripheral (PNS) and the central nervous system (CNS) of chick and rat embryos. Polyclonal antimouse FGF antibodies, which cross-react with basic and acidic FGF, were used in the peroxidase immunocytochemical staining method. FGF immunoreactivity was found in neurons. Staining intensity in chick and rat brain neuronal cells increased during the culture period, reached a maximum after 6-8 days, and subsequently declined. Embryonic chick ciliary and dorsal root ganglionic (DRG) neurons as well as DRG neurons from newborn rat displayed intense FGF immunoreactivity at 12 and 48 hr in culture. Nonneuronal cells were not stained. The data demonstrate for the first time that cultured neurons from the CNS and PNS contain FGF.

Immunoreactive fibroblast growth factor (FGF) in a transplantable chondrosarcoma: inhibition of tumor growth by antibodies to FGF.

Using a radioimmunoassay for bovine pituitary fibroblast growth factor (FGF), we have established the presence of the immunoreactive mitogen in extracts of a transplantable mouse chondrosarcoma. Both neutral and acidic extracts of the tumor contain an immunoreactive FGF (ir-FGF) that cross-reacts in a parallel and dose-dependent fashion in the radioimmunoassay. The ir-FGF is retained on heparin-Sepharose affinity columns and can be detected in the same molecular weight forms as rat pituitary FGF. Mice (C57/Bl) inoculated with the tumor (10 mg, im) show a decreased rate of tumor growth when passively immunized with the antiserum to FGF. The results establish the presence of FGF in this tumor and implicate its role in the etiology of its development.