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Biomedical subjects

W F Herblin

Publications and source records attributed to W F Herblin.

At least 19 recordsLinked to original sources
PubMed1998

A novel angiotensin II binding site in murine tissues.

A novel binding site for angiotensin II has been identified in certain murine tissues. This site, denoted ATm, is distinct from both the AT1 and AT2 sites, as well as the various atypical sites that have been described. The site has a low affinity for angiotensin II (100 nM) and is not blocked by losartan, PD123177 or saralasin. The site shows structural specificity for angiotensin II since both angiotensin III and angiotensin II (1-7) failed to inhibit the binding. Numerous standard drugs, including various receptor blockers, enzyme inhibitors and therapeutic agents, showed no affinity for the site. In murine tissues the site is associated with active tissue generation, as found in spleen, placenta and growing tumors, but its occurrence appears to be rare outside of the mouse.

Amino Acid Sequence↗
PubMed1996

Nonpeptide angiotensin II receptor antagonists: in vivo inhibition of [125I-Sar1,Ile8]angiotensin II binding by losartan, EXP597 and L-159,282 in rats.

Effects of losartan, L-159,282 and EXP597 on the in vivo binding of [125I-Sar1,Ile8]angiotensin II to kidney cortex and adrenal were examined in rats. Losartan, an AT1 receptor antagonist, completely blocked [125I-Sar1,Ile8]angiotensin II binding to the kidney cortex which contains only AT1 binding sites with an ID50 of 0.06 mg/kg. Losartan partially inhibited [125I-Sar1,Ile8]angiotensin II binding to the adrenal which contains equal amounts of AT1 and AT2 binding sites. Blockade by the AT1 receptor antagonist L-159,282 sufficiently increased the plasma levels of angiotensin II to block the AT2 receptor. EXP597 inhibited [125I-Sar1,Ile8]angiotensin II binding to the kidney cortex and adrenal almost totally with ID50s of 0.05 and 0.06 mg/kg, respectively. This result suggests that EXP597 exhibits almost equal binding affinity for AT1 and AT2 binding sites in vivo in rats.

Angiotensin II↗
PubMed1995

A rapid, high capacity assay for basic fibroblast growth factor binding.

A rapid, high capacity assay for the binding of basic fibroblast growth factor has been developed. Rat lung tissue was selected as the optimum source of membranes and 2M sodium chloride used to remove endogenous growth factor. The assay has been adapted to the Millipore Multi-Screen system so that it can be run in 96-well format with a volume of 300 microliters. The assay has been validated through the demonstration of inhibition by standard inhibitors such as suramin and protamine sulfate. The assay has proven useful for the screening of random compounds as well as the more detailed examination of suspected inhibitors. By running compounds in the presence and absence of a mid-range concentration of unlabeled bFGF, an estimate of the proportion of the inhibition due to high affinity binding can be obtained. Suramin and protamine sulfate show no selectivity and inhibit high affinity binding and overall binding with similar potencies. Another inhibitor, dimercaptothiadiazole, is more potent against high affinity binding.

Animals↗
PubMed1994

Identification and concerted function of two receptor binding surfaces on basic fibroblast growth factor required for mitogenesis.

Members of the fibroblast growth factor (FGF) family promote angiogenesis and wound repair, modulate early developmental events and survival of neurons, and have been associated with the pathogenesis of various diseases. FGFs interact with specific FGF receptors (FGFRs) and heparan sulfate proteoglycans on cell surfaces to mediate mitogenesis. Using protein structure-based site-directed mutagenesis of basic FGF (bFGF), we have identified two FGFR binding sites on bFGF which act in concert to initiate signal transduction. Both FGFR binding surfaces are distinct from the heparan sulfate proteoglycan binding domain. The primary, higher affinity, binding interaction comprises a cluster of solvent exposed hydrophobic amino acids (Tyr-24, Tyr-103, Leu-140, and Met-142), and two polar residues (Arg-44 and Asn-101). The hydrophobic contacts dominate the primary binding interaction and provide approximately 75% of the binding affinity. The secondary FGFR binding site on bFGF has an approximately 250-fold lower affinity and is composed of amino acids Lys-110, Tyr-111, and Trp-114 in a surface-exposed type I beta-turn (formerly known as the putative receptor binding loop). Binding of FGFR to both bFGF surfaces in a stoichiometry of 2FGFR:1bFGF is required for growth factor mediated cell proliferation. This represents a mechanism for the fibroblast growth factor/receptor family in which FGF facilitates FGFR dimerization and subsequent signal transduction events as a monomeric ligand.

Animals↗
PubMed1994

Suramin, an anticancer and angiosuppressive agent, inhibits endothelial cell binding of basic fibroblast growth factor, migration, proliferation, and induction of urokinase-type plasminogen activator.

Suramin, an anticancer agent in current clinical trials, is a prototype of a pharmacological antagonist of growth factors, including basic fibroblast growth factor (bFGF). Suramin inhibited angiogenesis in the chick chorioallantoic membrane assay in a dose-dependent fashion. Suramin, 200 mg/kg i.v., inhibited rat corneal angiogenesis induced by bFGF-impregnated polymers; addition of heparin stimulated angiogenesis and counteracted the inhibition of suramin. The half-maximal inhibitory concentration (IC50) of suramin was determined for key cellular mechanisms that regulate angiogenesis: (a) low and high affinity cellular binding of bFGF to bovine capillary endothelial (BCE) cells with IC50s, respectively, of 24.3 and 71.5 micrograms/ml; (b) spontaneous migration of bovine pulmonary artery endothelial and normal AG 7680 fetal bovine aortic endothelial cells; bFGF-stimulated migration of BCE and transformed GM 7373 fetal bovine aortic endothelial cells with IC50s of 200-320 micrograms/ml; (c) proliferation of bovine pulmonary artery endothelial cells at > 100 micrograms/ml and of BCE cells at > 250 micrograms/ml; and (d) urokinase-type plasminogen activator activity of GM 7373 endothelial cells stimulated by bFGF with an IC50 of 211 micrograms/ml and of BCE cells stimulated by bFGF at > 100 micrograms/ml, but not plasminogen activator activity induced by phorbol 12-myristate 13-acetate. Suramin inhibited multiple control points of angiogenesis, including those stimulated by bFGF. Because tumor growth is angiogenesis dependent, the clinical efficacy of suramin may relate, in part, to angiosuppression.

Animals↗
PubMed1994

Basic fibroblast growth factor and receptor expression in human ovarian cancer.

Basic fibroblast growth factor (bFGF) and other members of the FGF family share several biological properties that have the potential to mediate neoplastic cell growth. To test the hypothesis that bFGF may play a role in human ovarian cancer cell growth, three ovarian cancer cell lines, A90, A121(P), and A121(A), were investigated for their ability to respond to bFGF as a mitogen, to express endogenous bFGF protein or message for FGF proteins, and to exhibit FGF receptor or its message. Addition of bFGF to cultures of all three cell lines maintained in chemically defined media resulted in a statistically significant increase in cell number. Cell extracts from A90, A121(P), and A121(A) contained an immunoreactive protein that comigrated with hr-bFGF by Western blot analysis. Several bands of higher molecular weight were also noted. Immunohistochemical staining for bFGF demonstrated a cytoplasmic distribution of bFGF in the three cell lines. Both high- and low-affinity binding sites for human recombinant bFGF (hr-bFGF) were expressed by all three lines. High-affinity sites varied from 2700 sites per cell (Kd = 29 pM) to 13,500 sites per cell (Kd = 71 pM). All three cell lines were screened for mRNA expression for seven FGF proteins and four FGF receptors. In all three lines, mRNA for FGF2 (bFGF) was detected by PCR analysis, and in two lines, mRNA for FGF1 (aFGF) and FGF5 were also found. The FGFR1 receptor subtype (flg) was common to all of the cell lines. Finally, suramin inhibited proliferation of A90 and A121 (P and A) with IC50's of 60 and 210 micrograms/ml, respectively. This is consistent with the A90 cell line having higher levels of endogenous bFGF and flg and therefore being more responsive to suramin inhibition than the A121 cell line. The results indicate that these ovarian cancer cell lines can produce bFGF as well as other members of the FGF family of genes and have the ability to respond to bFGF.

Base Sequence↗
PubMed1993

Effects of modulation of basic fibroblast growth factor on tumor growth in vivo.

BACKGROUND: Recombinant human basic fibroblast growth factor (rHu-bFGF) is known to stimulate proliferation in some tumor cells and to modulate tumor vascularization. PURPOSE: The purpose of this study was to examine the possible role of this agent in the development of tumors. The study was designed to determine the effects of modulating bFGF activity in vivo in tumor models from cell lines with different responses to bFGF and with different content and receptor levels of bFGF. METHODS: Two tumor cell lines (human DLD-2 colon carcinoma and rat C6 glioma) were characterized for bFGF content and bFGF receptor levels by Western blot analysis in cultured cells and by studies of [125I]rHu-bFGF binding to sections from xenografts grown in nude mice. Tumor cell proliferation was monitored after treatment with rHu-bFGF or the DG2 or DE6 IgG monoclonal antibody to rHu-bFGF in culture and in vivo. RESULTS: C6 cells exhibited 7800 high-affinity receptors for rHu-bFGF per cell (dissociation constant [Kd] = 46 pM), while DLD-2 cells lacked high-affinity receptors. rHu-bFGF stimulated [3H]thymidine uptake by C6 cells, but the addition of DG2 IgG prevented this stimulation; rHu-bFGF had no effect on [3H]thymidine incorporation by DLD-2 cells. C6 cells had higher levels of immunoreactive bFGF than did DLD-2 cells. The xenografts from both cell lines exhibited high-affinity [125I]rHu-bFGF binding that was concentrated on vascular-like structures. rHu-bFGF at a dosage of 0.25 mg/kg given intraperitoneally daily for 18 days caused a twofold increase in DLD-2 tumor weight but had little effect on the growth of C6 xenografts. In contrast, daily intravenous injections of DG2 IgG given to mice had no effect on DLD-2 tumor growth but reduced growth of C6 tumors by approximately 30%--a statistically significant difference. CONCLUSIONS: The addition of exogenous rHu-bFGF or of a neutralizing antibody resulted in significant alterations in tumor growth in vivo, which were specific for tumor type and bFGF characteristics. While some of these effects may be mediated by the bFGF-responsive endothelial cells of the tumor vasculature (DLD-2 colon carcinoma), others may result from inhibition of bFGF-dependent tumor cell proliferation (C6 glioma). IMPLICATIONS: Studies that measure tumor blood flow are necessary to confirm that these effects are mediated by changes in tumor vasculature.

Animals↗
PubMed1993

Angiotensin II receptors in rabbit vascular grafts.

New Zealand White rabbits were used as a model of autologous venous bypass grafting. A section of jugular vein was sutured into the divided common carotid artery. The animals were euthanized one month after surgery. At that time histopathology showed myointimal thickening of the graft segments. Angiotensin II receptor assays showed that the AT1 receptor was present in the grafts, concentrated in the intima. These localized receptors were shown to be similar to the prototypic vascular AT1 receptor through the use of the specific antagonist DuP 753.

Angiotensin II↗
PubMed1993

New perspectives in angiotensin system control.

The 'angiotensin system' is expressed at the whole body, organ/tissue and cellular levels through the action of angiotensin II at specific receptors. An appreciation of the full scope of the actions of angiotensin II (endocrine, paracrine and autocrine) has been made possible by the discovery of the non-peptide angiotensin II receptor antagonists, losartan (DuP 753/MK954)(AT1-selective) and PD123177 (AT2-selective). Virtually all of the known effects of angiotensin II are blocked by losartan and designated AT1. Selective AT1 receptor blockade with losartan lowers BP in angiotensin II-dependent models of hypertension, reduces cardiac hypertrophy, improves haemodynamics in models of cardiac failure and reduces the intimal response to vascular injury. AT2 sites have been localised in distinct parts of the brain and in foetal tissue. The functional role of the AT2 sites remains controversial, but possible roles in neuronal ion channel function and collagen metabolism in fibroblasts have been reported. AT1 (losartan-sensitive) receptor subtypes have now been cloned from several rat tissues, suggesting that selective agents of the future may be even more specifically targeted. New perspectives in the control of the angiotensin system continue to evolve rapidly as the new receptor antagonists and molecular biology techniques expand our understanding of angiotensin II.

Amino Acid Sequence↗
PubMed1992

Immunolocalization of basic fibroblast growth factor to the microvasculature of human brain tumors.

BACKGROUND: Microvascular proliferation, a prominent feature of tumors of the central nervous system, is a prime target for anti-cancer therapy. METHODS: Because basic fibroblast growth factor (bFGF) plays a key role in the regulation of angiogenesis, surgical specimens from 52 human brain tumors were examined by immunocytochemical studies with a murine monoclonal antibody to bFGF. Sections from these tumors also were incubated with Ki-67 monoclonal antibody to measure the growth fraction. RESULTS: Immunostaining for bFGF was observed in 45 of 52 (87%) neoplasms, reacting with 97% of the malignant brain tumors and 67% of benign tumors (P < 0.01). The nonreactive tumors were a medulloblastoma and 7 of 21 (33%) benign, noninvasive, slow-growing neoplasms (1 acoustic schwannoma, 3 meningiomas, 2 pituitary adenomas, and 1 cholesteatoma). The indices of proliferation (Ki-67 labeling) were lower for the 21 benign tumors (1.2 +/- 1.1%) than the 31 malignant tumors (10.3 +/- 10.5%; P < 0.001). The bFGF was immunolocalized in the tumor cell nuclei in 23 of 52 tumors (44%) and in the cytoplasm of 8 of 52 (15%) tumors. Immunostaining to bFGF was prominent in the microvascular endothelial compartment in 84% of the malignant tumors and only 52% of benign tumors (P < 0.01). Immunostaining was not present after preabsorption of the antibody with pure human recombinant bFGF. CONCLUSIONS: The presence of bFGF predominantly within the tumor microvasculature indicates a cellular depot for this potent growth factor that mediates angiogenesis and tumorigenesis. These data support a role for bFGF in the transition from the benign to the malignant phenotype.

Blotting, Western↗
PubMed1992

Angiotensin II receptors and functional correlates.

Angiotensin II (ANG II) is the primary mediator of the renin-angiotensin system, which has an important functional role in cardiovascular homeostasis. The angiotensin receptor and its functional correlates have been redefined by the cloning of angiotensin receptors and the discovery and widespread study of specific nonpeptide ANG II-receptor antagonists losartan (AT1 selective) and PD123177 (AT2 selective). With these antagonists, it has been possible to extend the concept of ANG II-receptor heterogeneity to virtually every tissue and species. The losartan-sensitive sites have been shown to mediate all of the major ANG II-induced biologic effects, including vasoconstriction, aldosterone and catecholamine release, and central, ANG II-induced drinking behavior. The function of the AT2 site is not fully understood, but it may be involved in neuronal ion channel modulation and in fibroblast collagen metabolism. The presence of AT2 sites in fetal tissues and in discrete locations in the brain has encouraged continued research. Losartan, which represents the first of a new class of therapeutic agents, is currently undergoing clinical trials. A growing number of other AT1-selective ANG II-receptor antagonists are under development, including L-158,809, SKF 108566, and GR117285. Rat AT1-receptor subtypes have been cloned and sequenced (AT1A and AT1B). Human ANG II receptors have also been cloned and shown to have high affinity for losartan. A number of atypical angiotensin-binding sites have been identified from mycoplasma, amphibians, and mouse neuroblastoma, which are not sensitive to either losartan or PD123177.(ABSTRACT TRUNCATED AT 250 WORDS)

Acrylates↗
PubMed1992

Angiotensin II receptor subtypes.

The octapeptide, angiotensin II (Ang II), the biologically active component of the renin-angiotensin system, elicits its multiple actions through the stimulation of specific surface receptors on various target organs. Although the existence of Ang II receptor subtypes has been suspected for some time, definitive evidence for Ang II receptor heterogeneity has been obtained only with the recently introduced nonpeptide Ang II receptor antagonists, exemplified by the prototypic compounds DuP 753 and PD 123177. The sites having high affinity for DuP 753 are designated as site 1 (AT1 receptors) and those having a high affinity for PD 123177 as site 2 (AT2 receptors). Unlike Ang I, Ang II, Ang III, and peptide antagonists, such as saralasin, which all are relatively nonselective ligands for both Ang II receptors, the peptides CGP42112A and p-aminophenylalanine6-Ang II show a marked preference for the AT2 site. The occurrence of the AT1 and AT2 receptor subtypes/binding sites identified so far appears widespread. The presence and proportion of these receptors vary significantly among different tissues/organs of the same species and within the same tissue/organ of different species. Despite the abundance of the AT2 site, its functional correlates remain to be determined. The DuP 753-sensitive site (AT1 receptor) mediates all the major Ang II-induced biological effects, including adrenal aldosterone and catecholamine secretion, release of catecholamines from sympathetic ganglia, central nervous system responses, and vasoconstriction.

Angiotensin Receptor Antagonists↗