PubMed Health⌕ Search

Biomedical subjects

L I Gold

Publications and source records attributed to L I Gold.

At least 73 records · Page 4Linked to original sources
PubMed1992

Immunohistochemical staining for transforming growth factor beta 1 associates with disease progression in human breast cancer.

The transforming growth factor beta s (TGF-beta) comprise a family of M(r) 25,000 pluripotent growth factors which have been implicated in the development and progression of human breast cancer. Conflicting data suggest that TGF-beta has the potential to either inhibit or promote the progression of mammary neoplasia. We therefore examined a pathological library of malignant breast biopsy specimens to determine the prevalence and distribution of immunoreactivity with antibodies specific for the three mammalian isoforms of TGF-beta (beta 1, beta 2, and beta 3). We found that intense staining for TGF-beta 1 was positively associated with rate of disease progression, and that this was independent of age, stage, nodal status, or estrogen receptor status (P = 0.009).

Adult↗
PubMed1992

Localization of the cleavage sites on fibronectin following digestion by urokinase.

Urokinase (u-PA) proteolytically cleaves both human plasma (pFn) and cellular (cFn) dimeric fibronectin (M(r) 440,000) into four major polypeptides of approximately M(r) 210,000, 200,000, 25,000, and 6,000. Amino acid sequence analysis of the polypeptide fragments indicated that the enzymatic cleavage of Fn occurs at two sites: 1) between an arginine/alanine peptide bond located C-terminal to residue 259; this cleavage liberates the N-terminal M(r) 25,000 fragment and the M(r) 210,000 and M(r) 200,000 polypeptides derived from the A and B chains of Fn, respectively; and 2) between an arginine/threonine peptide bond located C-terminal to residue 2,299, thereby yielding an M(r) 6,000 dimeric fragment containing the C-terminal interchain disulfide bonds. Predigestion of Fn with u-PA increased the molecule's vulnerability to further attack by the enzymes plasmin and cathepsin D. These data provide further biochemical evidence for the proteolytic cleavage of fibronectin by plasminogen activators and substantiate that u-PA digestion of Fn may be an initial event in the local degradation of the extracellular matrix by malignant cells, possessing elevated levels of these enzymes.

Amino Acid Sequence↗
PubMed1992

Effect of topical retinoic acids on the levels of collagen mRNA during the repair of UVB-induced dermal damage in the hairless mouse and the possible role of TGF-beta as a mediator.

Topically applied retinoic acids have been found to enhance the gene expression for collagen types I and III in the skin of UVB-irradiated hairless mice. Prior damage is required because the effect is not observed in the skin of age-matched, non-irradiated control animals. Immunochemical methods have shown an increase in TGF-beta 1 and, to a lesser extent, of TGF-beta 2 in the epidermis following retinoic acid treatment. There were no changes in mRNA levels for any of the isotypes of TGF-beta induced by retinoic acid treatment. This study suggests that TGF-beta may mediate the effect of retinoic acids on dermal repair through the stimulation of collagen gene expression.

Administration, Topical↗
PubMed1992

Transforming growth factor-beta and transforming growth factor beta-receptor expression in human meningioma cells.

The transforming growth factor-beta (TGF beta) family in mammals includes three closely related peptides that influence proliferation and numerous physiologic processes in most mesenchymal cells. In this study, Northern blots, immunohistochemistry, TGF beta radioreceptor assays, TGF beta receptor affinity labeling and [3H] thymidine incorporation were used to evaluate whether primary cell cultures of human meningiomas synthesize the three TGF beta isoforms, bear TGF beta receptors, and respond to TGF beta. Transcripts for TGF beta 1 and 2 were detected in the three cases analyzed. Transforming growth factor-beta 1 immunoreactivity was detected in three of six cases, and TGF beta 2 and 3 immunoreactivity were detected in each case analyzed. Media conditioned by cells cultured from six meningiomas also contained latent TGF beta-like activity. Transforming growth factor-beta receptor cross-linking studies identified TGF beta binding sites corresponding to the type 1, type 2, and type 3 receptors on meningioma cells. Treatment with active TGF beta 1 produced a statistically significant reduction in [3H] thymidine incorporation after stimulation with 10% fetal calf serum and epidermal growth factor in all six cases studied.

Blotting, Northern↗
PubMed1992

Evidence for transforming growth factor-beta expression in human leptomeningeal cells and transforming growth factor-beta-like activity in human cerebrospinal fluid.

BACKGROUND: Little is known about the factors regulating growth and maintenance of human leptomeningeal cells. The influence of cerebrospinal fluid on these functions is also unknown. Possible mediators include the transforming growth factor-beta (TGF beta) family, three closely related peptides that regulate proliferation and numerous other physiologic processes in most mesenchymal cells. EXPERIMENTAL DESIGN: Expression of both mRNA and protein for TGF beta isoforms TGF beta 1, TGF beta 2, and TGF beta 3 as well as TGF beta-competing activity were evaluated in primary human leptomeningeal cultures by Northern blot analysis, immunohistochemistry, and a radioreceptor assay, respectively. TGF beta 1, TGF beta 2, and TGF beta 3 immunoreactivity was also evaluated in brain sections containing leptomeninges from which these cell cultures were established. An additional study analyzed human cerebrospinal fluid for TGF beta-like activity. RESULTS: Transcripts for TGF beta 1, TGF beta 2 and TGF beta 3 were detected in RNA from each of the eight leptomeningeal cultures. Significant TGF beta 1 immunoreactivity was detected in leptomeningeal tissue from five of eight cases. TGF beta 2 and TGF beta 3 immunostaining was seen in eight and seven of the cases, respectively. Similarly, cells cultured from these meninges exhibited variable TGF beta 1 and extensive TGF beta 2 and TGF beta 3 immunoreactivity. Radioreceptor assays of conditioned media from four cultures demonstrated significant latent TGF beta-like activity. TGF beta radioreceptor competing activity was also detected by radioreceptor assay in normal blood-free cerebrospinal fluid from 32 patients without neurological disease. In addition, pooled cerebrospinal fluid (from six additional patients) exhibited dose dependent TGF beta-like activity in the radioreceptor assay, stimulation of AKR-2B cell growth in soft agar and inhibition of growth in CCL-64 cell assays suggesting that cerebrospinal fluid contains TG beta-like activity. CONCLUSIONS: These findings suggest that the human leptomeninges synthesize TGF beta 1, TGF beta 2 and TGF beta 3 and secrete latent TGF beta s at least in vitro. Human cerebrospinal fluid may also contain TGF beta isoforms. Collectively, these observations raise the possibility that members of the TGF beta family contribute to biologic processes of the leptomeninges.

Agar↗
PubMed1991

Transforming growth factor beta 1, a potent chemoattractant for human neutrophils, bypasses classic signal-transduction pathways.

Transforming growth factor beta 1 (TGF-beta 1), a homodimeric polypeptide (Mr 25,000), derives from inflammatory cells and acts as a chemoattractant for monocytes and fibroblasts. We report here that TGF-beta 1 is also the most potent chemoattractant yet described for human peripheral blood neutrophils. Recombinant TGF-beta 1 elicited dose-dependent directed migration of neutrophils under agarose that was inhibited in the presence of a neutralizing antibody to TGF-beta 1. Maximal chemotaxis was evoked by TGF-beta 1 at femtomolar concentrations, whereas conventional chemoattractants act at nanomolar concentrations: on a molar basis, TGF-beta 1 was 150,000 times more potent than fMet-Leu-Phe. In contrast, TGF-beta 1 provoked neither exocytosis nor the production of superoxide by neutrophils. We further analyzed the mechanism by which TGF-beta 1 elicits chemotaxis (GTPase activity, [Ca2+], and actin polymerization). In contrast to the conventional chemoattractant fMet-Leu-Phe, TGF-beta neither activated classic heterotrimeric guanine nucleotide-binding proteins nor provoked global mobilization of intracellular Ca2+. Chemoattraction by both fMet-Leu-Phe and TGF-beta 1 was inhibited by cycloheximide and actinomycin D. Moreover, chemotaxis in response to TGF-beta 1 was associated with the polymerization of actin. The selectivity and potency of TGF-beta 1 as a chemoattractant suggest that it elicits directed cell migration by means of a pathway that depends not on classic intracellular signals but on protein synthesis.

Calcium↗
PubMed1991

Immunohistochemical localization of TGF beta 1, TGF beta 2, and TGF beta 3 in the mouse embryo: expression patterns suggest multiple roles during embryonic development.

Isoform-specific antibodies to TGF beta 1, TGF beta 2, and TGF beta 3 proteins were generated and have been used to examine the expression of these factors in the developing mouse embryo from 12.5-18.5 d post coitum (d.p.c.). These studies demonstrate the initial characterization of both TGF beta 2 and beta 3 in mammalian embryogenesis and are compared with TGF beta 1. Expression of one or all three TGF beta proteins was observed in many tissues, e.g., cartilage, bone, teeth, muscle, heart, blood vessels, lung, kidney, gut, liver, eye, ear, skin, and nervous tissue. Furthermore, all three TGF beta proteins demonstrated discrete cell-specific patterns of expression at various stages of development and the wide variety of tissues expressing TGF beta proteins represent all three primary embryonic germ layers. For example, specific localization of TGF beta 1 was observed in the lens fibers of the eye (ectoderm), TGF beta 2 in the cortex of the adrenal gland (mesoderm), and TGF beta 3 in the cochlear epithelium of the inner ear (endoderm). Compared to the expression of TGF beta mRNA transcripts in a given embryonic tissue, TGF beta proteins were frequently colocalized within the same cell type as the mRNA, but in some cases were observed to localize to different cells than the mRNA, thereby indicating that a complex pattern of transcription, translation, and secretion for TGF beta s 1-3 exists in the mouse embryo. This also indicates that TGF beta 1, beta 2, and beta 3 act through both paracrine and autocrine mechanisms during mammalian embryogenesis.

Animals↗
PubMed1991

Expression of transforming growth factor-beta 1, -beta 2, and -beta 3 mRNA and protein in the murine lung.

Evidence has accumulated suggesting that the various isoforms of beta-type transforming growth factors (TGF-beta s) regulate important functions in the lung; however, the cellular source of these proteins is not well defined. Northern blot analysis of murine lung tissue demonstrates that mRNA transcripts for all three TGF-beta isoforms are found from birth through adulthood. Although the level of expression for each TGF-beta is variable during the first 2 wk post partum, all three isoforms are equal in the adult lung. Using in situ hybridization and immunohistochemical analysis, we have localized both mRNA and protein expression for all three isoforms of TGF-beta in the adult murine lung. At low magnification, immunohistochemical localization of TGF-beta proteins appears coincident in their pattern of expression with TGF-beta mRNAs in the large proximal conducting airways of the lung. However, on closer analysis, protein expression of all three TGF-beta isoforms is confined to the bronchiolar epithelium, while TGF-beta mRNA transcripts for each of the TGF-beta genes are found in smooth muscle cells and connective tissue fibroblasts lying subjacent to the epithelium. Although the levels of both TGF-beta mRNA and protein expression are high in the proximal bronchiolar tree, their signal intensities completely disappear as the terminal bronchioles progress to respiratory bronchioles. Additionally, in the lung vasculature, there is very high expression of all three TGF-beta mRNA transcripts in the smooth muscle cells of the large vessels. TGF-beta2 and TGF-beta but not TGF-beta1 proteins are expressed in these same smooth muscle cells.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1989

Biochemical characterization of the fibronectin binding sites for IgG.

Plasma fibronectin (Fn) is a constituent of cryoglobulins and has been shown to interact with immune complexes. In a previous report we demonstrated that Fn specifically bound to IgG immobilized on a solid matrix. To localize and biochemically characterize the sites on the Fn molecule involved in this interaction, Fn was enzymatically cleaved with subtilisin and subjected to IgG affinity chromatography. Three major polypeptide fragments of 16 kDa, 22 kDa, and a triplet of 26- to 29-kDa bound IgG. They were localized to three separate regions of the molecule by Western blot analysis using antisera to specific regions of the Fn molecule, by amino acid sequencing, and by their previously described heparin binding affinities. The 22-kDa fragment interacted with IgG under physiologic conditions and it is localized at the N-terminal of the Fn molecule. The 16-kDa and 26- to 29-kDa fragments bound to IgG under conditions of lower ionic strength; the former commences at residue 588, carboxyl-terminal to the collagen binding region and the latter begins at residue 1597, carboxyl-terminal to the cell binding domain. The interaction of Fn with Ig has significant implications in host defense and also in immune complex disease where basement membrane Fn may sequester immune complexes from the circulation.

Amino Acid Sequence↗
PubMed1989

Human plasma fibronectin as a substrate for human urokinase.

An early event in malignant transformation is the increased expression of proteases, such as plasminogen activator, which can degrade surrounding extracellular matrices, thereby conferring an advantage for tumour cell invasion and metastasis. The present studies provide evidence that plasma fibronectin (Fn), which is a component of the extracellular matrix, is a direct substrate for the plasminogen activator urokinase (UK). Human plasma Fn was incubated with human UK under plasminogen-free conditions. Fn cleavage was both time- and dose-dependent and was evident within 30 min. The proteolytic digestion was limited and complete within 12 h at an enzyme/substrate ratio of 1:20. Analysis of the final proteolytic digestion products demonstrated the disappearance of the native dimeric 440 kDa structure of Fn with the concomitant appearance of three proteolytic fragments of 210, 200 and 25 kDa. Since two large fragments of similar size to the 220 kDa monomeric chains of Fn were obtained following proteolysis, it is proposed that UK cleaves Fn at two sites, one towards the N-terminal and one close to the C-terminal, but N-terminal to its interchain disulphide bonds. These studies suggest that the local proteolytic digestion and release of Fn from the extracellular matrix by tumour cells possessing high levels of UK may involve the direct proteolytic breakdown of Fn by UK.

Fibrinolysin↗
PubMed1987

Limited cleavage of cellular fibronectin by plasminogen activator purified from transformed cells.

The substrate specificity and direct catalytic activity of plasminogen activator (PA) was examined under conditions where its natural substrate, plasminogen, was missing or inhibited. PA, purified from cultures of transformed chicken fibroblasts, was incubated with purified preparations of potential substrates. The adhesive glycoprotein fibronectin, isolated from normal chicken fibroblast extracellular matrix, underwent limited but specific cleavage by PA in the absence of plasminogen. Analysis of the cleavage products by polyacrylamide gels under both reducing and nonreducing conditions indicated that PA-mediated cleavage occurred near the carboxyl terminus of fibronectin but on the amino-terminal side of the interchain disulfide bridge, thus disrupting the native dimeric fibronectin molecule. Under the identical conditions, chicken ovalbumin was not cleaved while the established substrate, chicken plasminogen, was extensively converted to plasmin. A monoclonal antibody, directed against avian PA and shown to inhibit plasminogen-free, cell-mediated matrix degradation, specifically inhibited the fibronectin cleavage. A human PA, urokinase, also cleaved fibronectin under plasminogen-free conditions yielding a limited number of high molecular weight cleavage products.

Animals↗
PubMed1980

Fibronectin-collagen binding and requirement during cellular adhesion.

Fibronectin isolated from human plasma and from the extracellular matrices of cell monolayers mediates the attachment in vitro and spreading of trypsin-treated cells on a collagen substratum. Fibronectin-dependent kinetics of cellular attachment to collagen were studied for several adherent cell types. It was shown that trypsin-treated human umbilical-cord cells, mouse sarcoma CMT81 cells, endothelial cells, and human fibroblasts from a patient with Glanzmann's disease were completely dependent on fibronectin for their attachment to collagen, whereas guinea-pig and monkey smooth-muscle cells and chick-embryo secondary fibroblasts displayed varying degrees of dependence on fibronectin for their attachment. Radiolabelled human plasma fibronectin possessed similar affinity for collagen types I, II and III from a variety of sources. The fibronectin bound equally well to the collagens with or without prior urea treatment. However, in the fibronectin-mediated adhesion assay using PyBHK fibroblasts, a greater number of cells adhered and more spreading was observed on urea-treated collagen. Fibronectin extracted from the extracellular matrix of chick-embryo fibroblasts and that purified from human plasma demonstrated very similar kinetics of complexing to collagencoated tissue-culture dishes. Fibronectin from both sources bound to collagen in the presence of 0.05-4.0m-NaCl and over the pH range 2.6-10.6. The binding was inhibited when fibronectin was incubated with 40-80% ethylene glycol, the ionic detergents sodium dodecyl sulphate and deoxycholate, and the non-ionic detergents Nonidet P-40, Tween 80 and Triton X-100, all at a concentration of 0.1%. From these results we proposed that fibronectin-collagen complexing is mainly attributable to hydrophobic interactions.

Animals↗
PubMed1979

Stability of fibronectin biological activity following chemical modification.

Fibronectin isolated from human plasma functions in vitro as a mediator of adhesion and spreading of trypsinized fibroblasts on native or denatured collagen. As a means of elucidating structural characteristics which might contribute to fibronectin's biological activity, we have modified and digested the protein with several chemicals. Following various treatments, the protein was utilized to mediate cell adhesion and spreading on collagen to determine which alteration disrupted its activity. Fibronectin remained functionally intact after partial or complete reduction and alkylation, oxidation of 59% of the carbohydrates with sodium periodate, citraconylation, carbodiimide-catalyzed amide formation, and oxidation of 35.2 residues of tryptophan/molecule with N-bromosuccinimide. Dinitrofluorobenzene treatment, which phenylated ten residues/molecule of fibronectin, successfully inactivated fibronectin's in vitro biological function. Effective modification of the protein was determined by appropriate analytical procedures. Since fibronectin retained its biological function after several treatments that presumably affected its molecular conformation, we concluded that its secondary or tertiary structure appears not to be essential for its in vitro activity, or alternatively that the protein possesses a biologically active domain relatively resistant to chemical modification.

Bromosuccinimide↗
PubMed1979

Subtilisin and cyanogen bromide cleavage products of fibronectin that retain gelatin-binding activity.

The gelatin-binding region of fibronectin has been obtained by subtilisin digestion and cyanogen bromide cleavage of the molecule. Enzymatic digestion yielded two fragments of molecular weights 50,000 (S50K) and 30,000 (S30K) which were isolated by elution from gelatin-Sepharose affinity columns. Because the S50K fragment also mediated the adhesion of fibroblasts to collagen, it contains both the collagen and cell binding sites on the fibronectin molecule. Both fragments had valine as the NH2-terminal residue, were enriched in half-cystine and methionine residues compared to the whole molecule, and were identical by immunodiffusion. The S50K fragment begins with the sequence Val-Tyr-Gln-Pro-Gln-Pro-His-Pro-Gln-Pro-(Pro)-(Gly)-Tyr-Gly-His-( )-Val, a region with an extended conformation which is susceptible to proteolysis and connects this domain to the remainder of the fibronectin molecule. The S50K fragment appears to be located in the COOH-terminal one-third of the fibronectin molecule but does not contain the interchain disulfide bridge(s); the S30K fragment is probably derived from the NH2-terminal region of S50K.

Amino Acids↗