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Biomedical subjects

A B Roberts

Publications and source records attributed to A B Roberts.

At least 199 records · Page 11Linked to original sources
PubMed1990

Fructosamine compared with a glucose load as a screening test for gestational diabetes.

Five hundred seven women were screened for gestational diabetes between 20-36 weeks' gestation. All received a 100-g glucose (polycose) load at 28 weeks with measurement of plasma glucose 1 hour later. Fructosamine levels were measured at 4-week intervals from 20-36 weeks. At 36 weeks, a full 100-g 3-hour glucose tolerance test was performed on all subjects. Eighteen women were diagnosed as having gestational diabetes. The glucose load had a sensitivity of 81% in detection of gestational diabetes, compared with 50% for fructosamine at 36 weeks. Fructosamine is not useful as a screening test for gestational diabetes as currently defined.

Diabetes Mellitus↗
PubMed1990

Autoinduction of transforming growth factor beta 1 is mediated by the AP-1 complex.

The multifunctional actions of transforming growth factor beta 1 (TGF-beta 1) indicate that it has a pivotal control function in many physiological and pathological processes. An important property of TGF-beta 1 is its ability to activate its own mRNA expression and thereby increase its own secretion. Two distinct regions of the promoter of the TGF-beta 1 gene are responsive to autoregulation: one 5' to the upstream transcriptional start site and another located between the two major start sites. In both promoter regions, autoinduction is mediated by binding of the AP-1 (Jun-Fos) complex. An important contribution to this positive regulation is the autoactivation of c-jun transcription by AP-1. Cotransfection of antisense c-jun or antisense c-fos expression vectors prevents TGF-beta 1 autoinduction. These results demonstrate that both components of the AP-1 complex are required for TGF-beta 1 autoinduction. Induction of jun expression by TGF-beta 1, as well as jun autoinduction, may amplify the action of TGF-beta 1 during normal development and oncogenesis.

Adenocarcinoma↗
PubMed1990

Transcriptional regulation of the transforming growth factor beta 1 promoter by v-src gene products is mediated through the AP-1 complex.

Growth factor-independent 32D-src and 32D-abl cell lines, established by infecting the interleukin-3-dependent myeloid precursor cell line (32D-123) with retroviruses containing the src or abl oncogene, were used to study transcriptional regulation of transforming growth factor beta 1 (TGF-beta 1) mRNA. Analysis of different TGF-beta 1 promoter constructs regulated by pp60v-src indicated that sequences responsive to high levels of src induction contain binding sites for AP-1. Both src and serum induced expression of the c-fos and c-jun genes in myeloid cells, resulting in transcriptional activation of the TGF-beta 1 gene. We found that serum treatment increased TGF-beta 1 mRNA levels in 32D-123 cells and that the v-Src protein could replace the serum requirement by stimulating binding to the AP-1 complex of the TGF-beta 1 promoter, thereby mediating the induction of TGF-beta 1 transcription.

Animals↗
PubMed1990

Interleukin-1 stimulates and all-trans-retinoic acid inhibits collagenase gene expression through its 5' activator protein-1-binding site.

Collagenase production by synovial fibroblast-like cells (synoviocytes) plays a major role in cartilage and bone destruction in rheumatoid arthritis. Interleukin-1 (IL-1) increases collagenase secretion by elevating the steady state levels of collagenase mRNA in cultured rheumatoid synoviocytes, while all-trans-retinoic acid (RA) has the opposite effect. We have studied the regulation of collagenase gene transcription by IL-1 and RA in synoviocytes by transient transfection of plasmid constructs containing deletion mutants of the 5'-flanking region of the collagenase gene or the isolated phorbol ester-responsive element ligated to a chloramphenicol acetyltransferase reporter gene. We show that the phorbol ester-responsive element of the collagenase gene mediates both positive and negative regulatory effects, respectively, of IL-1 and RA on transcription. In addition, we show that IL-1 and 12-O-tetradecanoyl-phorbol-13-acetate transiently induce c-jun and c-fos expression and that retinoic acid inhibits IL-1 and 12-O-tetradecanoyl-phorbol-13-acetate induction of c-fos, but not c-jun. These results suggest that RA inhibits collagenase transcription at least in part through inhibition of c-fos.

Binding Sites↗
PubMed1990

Colocalization of TGF-beta 1 and collagen I and III, fibronectin and glycosaminoglycans during lung branching morphogenesis.

The possible in vivo role of TGF-beta 1 in regulating various proteins of the extracellular matrix, including fibronectin, collagen I and III, and glycosaminoglycans, was examined by immunohistochemical methods during critical stages of lung morphogenesis in the 11- to 18-day-old mouse embryo. Sections of Bouin-fixed, paraffin-embedded whole embryos were exposed to polyclonal antibodies specific to synthetic peptides present in the precursor part of TGF-beta 1 (pro-TGF-beta 1), in the processed TGF-beta 1 (antibody CC), collagen I and III, fibronectin, followed by the PAP or ABC technique to visualize the location of the antibody. GAG were stained with Alcian Blue 8GX. Our results indicate colocalization of TGF-beta 1 expression and that of matrix proteins in the developing lung when branching morphogenesis (cleft formation) and tissue stabilization occur. The presence of TGF-beta 1 at the epithelial-mesenchymal interfaces of stalks and clefts at a time when matrix proteins can first be visualized in these areas, suggests a direct participation of the growth factor in the development of the basic architecture of the lung.

Animals↗
PubMed1990

Transforming growth factor-beta in human aqueous humor.

The transforming growth factor-beta s are peptide growth factors known to play a central role in wound healing. Using a specific, in vitro assay of cell growth inhibition, we have detected transforming growth factor-beta (TGF-beta) in 24/24 aqueous humor specimens from eyes undergoing cataract extraction with intraocular lens implantation. The amount of TGF-beta ranged from 2.3 to 8.1 ng/ml (mean +/- SD = 4.5 +/- 1.7 ng/ml), with 61% present in the active form. Subtyping of TGF-beta was performed by addition of antibodies specific for the beta 1 and beta 2 isoforms to the growth inhibition assay, and confirmed with a sandwich enzyme-linked immunosorbent assay. None of the TGF-beta detected was of the beta 1 isoform; in contrast, the beta 2 isoform was present in every sample, implying that it might have originated from ocular tissues. The presence of this potent modulator of tissue repair in aqueous humor suggests a role in the healing processes following intraocular surgery, including glaucoma filtration surgery.

Aged↗
PubMed1990

Mesoderm induction in Xenopus laevis distinguishes between the various TGF-beta isoforms.

Induction of mesoderm in ectodermal explants of Xenopus laevis blastula embryos had previously been shown to respond selectively to TGF-beta 2, with TGF-beta s 1 and 5 having no activity in this assay. As TGF-beta s 1, 2, and 3 are frequently coexpressed in tissues, we wished to examine the activity of TGF-beta 3 relative to that of TGF-beta s 1 and 2 in this assay as well as in other in vitro assays. We report here that when the activity of recombinant TGF-beta 3 is normalized to that of TGF-beta 1 in the assay for growth inhibition in CCL-64 cells, it is also equal to that of TGF-beta 1 in assays for stimulation of both anchorage-independent growth of rat NRK cells and chemotaxis of human monocytes. In contrast, in the assay for mesoderm induction, recombinant TGF-beta 3 is 10-fold more active than TGF-beta 2, inducing expression of muscle specific alpha-actin at concentrations as low as 1 ng/ml. These results suggest that more complex systems, in contrast to individual cell types, may respond selectively to the various TGF-beta isoforms and that there might be biological consequences of TGF-beta isoform switching in vivo.

Animals↗
PubMed1990

Antibodies to transforming growth factor-beta 2 peptides: specific detection of TGF-beta 2 in immunoassays.

Polyclonal antibodies have been raised to synthetic peptides corresponding to several regions of transforming growth factor-beta 2 (TGF-beta 2). All antisera were tested for their ability to react with either the native or reduced forms of both TGF-beta 1 and TGF-beta 2 in enzyme-linked immunosorbent assays, Western blots, and immunoprecipitation assays. On Western blots, antisera raised to a peptide corresponding to residues 50-75 of TGF-beta 2 specifically detected 5 ng TGF-beta 2, while antisera raised to regions 1-30 and 79-108 cross-reacted with TGF-beta 1. Anti-P 50-75(2) also localized TGF-beta 2 in murine placenta in immunohistochemical studies. In immunoprecipitation assays with either iodinated TGF-beta s or with media conditioned by cells labeled with [35S]cysteine, both anti-P 50-75(2) and anti-P 79-108(2) specifically immunoprecipitated TGF-beta 2 under reducing conditions only, while anti-P 79-108(2) also reacted with TGF-beta 1. None of the TGF-beta 2 peptide antibodies was able to block receptor binding of either TGF-beta 1 or 2. Analysis of the cross-reactivity patterns of these peptide antibodies suggests conformational differences between TGF-beta 1 and TGF-beta 2.

Amino Acid Sequence↗
PubMed1990

Complementary deoxyribonucleic acid cloning of an mRNA encoding transforming growth factor-beta 2 from chicken embryo chondrocytes.

Using a simian transforming growth factor-beta 2 (TGF-beta 2) cDNA probe, we have isolated chicken cDNA clones for TGF-beta 2 from a chicken embryo chondrocyte cDNA library. The predicted precursor protein of chicken TGF-beta 2 is 412 amino acids long, two amino acids shorter than the human form to which it shows 90% identity. Cleavage of the chicken TGF-beta 2 precursor at a pentabasic Arg-Arg-Lys-Lys-Arg site would produce a 112-amino acid processed peptide differing by only one amino acid from the human TGF-beta 2 peptide. In contrast to TGF-beta 3 and 4 mRNAs, TGF-beta 2 mRNA is expressed in both cultured and non-cultured chicken embryo chondrocytes and at higher levels in chicken embryo fibroblasts. In chondrocytes and fibroblasts, the 3.9- and 4.3-kb TGF-beta 2 mRNAs are both expressed at higher levels than the 8-kb TGF-beta 2 mRNA; however, in developing chicken embryos, the level of expression of the 8-kb mRNA is higher than that of the 3.9- and 4.3-kb mRNAs.

Amino Acid Sequence↗
PubMed1990

Isolation and characterization of TGF-beta 2 and TGF-beta 5 from medium conditioned by Xenopus XTC cells.

TGF-beta 2 and -beta 5 have been purified from medium conditioned by Xenopus cultured cells (XTC) and identified based on their N-terminal amino acid sequence analysis and biological activity. When applied in high concentrations, Xenopus TGF-beta 2, like porcine TGF-beta 2, induces expression of mesodermal markers from cultured Xenopus ectodermal explants, whereas TGF-beta 5 is inactive in this assay. However, the TGF-beta 's could be separated from the major mesoderm-inducing activity present in XTC medium. Xenopus TGF-beta 2 and -beta 5 are approximately equivalent to TGF-beta 1 in their abilities to inhibit the growth of mink lung CCL-64 cells, induce anchorage-independent growth of rat NRK cells, inhibit the proliferation and antibody secretion of human B-lymphocytes, and stimulate chemotaxis of human monocytes. These data establish the functional activity of TGF-beta 5 and suggest that more complex multicellular systems, in contrast to most isolated cells, discriminate between the different TGF-beta s.

Amino Acid Sequence↗
PubMed1990

Cloning by polymerase chain reaction of a new mouse TGF-beta, mTGF-beta 3.

Two TGF-beta s, TGF-beta 1 and 2, have previously been isolated from the mouse. Here we report the isolation of a murine TGF-beta 3 cDNA. RNAs extracted from 15-day-old mouse embryos and several mouse cell lines were reverse-transcribed. These cDNA mixtures were used as substrates for polymerase chain-reaction amplifications, using oligonucleotides designed on the basis of known human and chicken TGF-beta 3 sequences, including the initiation and stop codons. Several overlapping cDNAs containing either the amino-terminal domain or the carboxy-terminal domain, as well as the complete 1.2-kb coding region of the mouse TGF-beta 3 cDNA were obtained. The mouse TGF-beta 3 coding region is 1230 nucleotides long and codes for a 410 amino acid polypeptide very similar to its human counterpart. This cDNA hybridizes to a unique 3.5-kb RNA and is differentially expressed in various mouse tissues and at different embryonic stages.

Amino Acid Sequence↗
PubMed1990

Transforming growth factor-beta 1 specifically localizes in elastin during synovial inflammation: an immunoelectron microscopic study.

We report here that extracellular TGF-beta 1 is associated exclusively with microfibrils of elastin which are present in the extracellular matrix of the inflamed articular joint of the rat. Inflammation was initiated by bacterial cell walls localized in the synovium following intraperitoneal injection of the bacterial components. This synovitis is associated with both destruction of connective tissue components and matrix deposition. The growth factor was localized by using a polyclonal antibody raised to a synthetic peptide corresponding to amino terminal 30 amino acids of TGF-beta 1 in conjunction with a gold-labeled secondary antibody. The results suggest a close association of TGF-beta 1 with proteoglycans which are known to be a major component of the microfibrils in elastin. Proteoglycan-mediated binding and concentration of TGF-beta 1 in specific areas of the extracellular matrix may constitute a mechanism whereby the growth factor could be targeted to specific sites of action.

Animals↗
PubMed1990

Cytokines and growth regulation of synoviocytes from patients with rheumatoid arthritis and rats with streptococcal cell wall arthritis.

Paracrine growth factors probably stimulate the pathologic proliferation of synovial fibroblast-like cells (synoviocytes) in rheumatoid arthritis (RA), but the relative importance of various factors is highly controversial. To address this problem, we compared the effects of highly purified or recombinant cytokines, in serum-free medium, on the in vitro long-term growth of synoviocytes from patients with RA and rats with streptococcal cell wall (SCW) arthritis. Of the factors tested (PDGF, aFGF, bFGF, EGF, TGF-beta, IL-1-alpha, TNF-alpha and IFN-gamma), PDGF, was clearly the most potent stimulant of long-term growth of both rat and human synoviocytes. The strong mitogenic activity of rheumatoid synovial fluids was significantly inhibited by neutralizing anti-PDGF antibody, thus confirming the importance of PDGF. EGF, TGF-beta, IL-1-alpha, TNF-alpha, and IFN-gamma had minimal effects. Similar to the effects on anchorage-independent growth, TGF-beta 1 and 2, inhibited serum- or PDGF-stimulated anchorage-dependent growth. Considered in the context of other reports, these data support the view that cytokines such as PDGF, and possibly aFGF and bFGF, play major roles in stimulating synoviocyte hyperplasia in RA and SCW arthritis, whereas TGF-beta may inhibit synoviocyte growth.

Animals↗
PubMed1989

Activation of the second promoter of the transforming growth factor-beta 1 gene by transforming growth factor-beta 1 and phorbol ester occurs through the same target sequences.

Two distinct regions of the transforming growth factor-beta 1 (TGF-beta 1) promoter are responsive to autoregulation and activation by phorbol ester (12-O-tetradecanoylphorbol-13-acetate): sequences located between nucleotides -454 to -323 (first promoter) and between the two transcriptional start sites. We have now characterized in detail the induction of the second promoter (sequences between nucleotides + 1 to +271) of the TGF-beta 1 gene by both TGF-beta 1 and phorbol ester. By assaying progressively deleted mutations in the second promoter, we have found two regions responsible for the induction; each contains a phorbol ester-responsive element. In vitro transcription of the second promoter-chloramphenicol acetyltransferase chimeric genes using nuclear extracts of A-549 cells showed that deletion of the putative phorbol ester-responsive elements results in a 70-80% decrease in activity. DNase I footprinting and gel mobility shift assays showed that binding to an Sp1 site and the putative TRE elements are required for maximal expression of the second promoter region of the TGF-beta 1 gene. These results suggest that AP-1, which is capable of conferring phorbol ester or TGF-beta 1 responsiveness, is the major transcription factor involved in the second promoter-derived transcription of the TGF-beta 1 gene.

Animals↗