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Biomedical subjects

H L Moses

Publications and source records attributed to H L Moses.

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

Transforming growth factor beta 1 regulation of c-myc expression, pRB phosphorylation, and cell cycle progression in keratinocytes.

Transforming growth factor beta 1 (TGF-beta 1) is a potent inhibitor of cellular proliferation in a variety of cell types, including skin keratinocytes. TGF-beta 1 suppression of c-myc transcription has been implicated in the mechanism of TGF-beta 1 inhibition of keratinocytes, and evidence suggests that the protein product of the retinoblastoma gene (pRB) is a necessary component in this pathway. Following growth factor stimulation of quiescent keratinocytes, TGF-beta 1 can inhibit cell cycle progression into S phase at any point prior to the G1-S transition but does not inhibit progression through the S phase of the cell cycle. Since pRB is hypophosphorylated during G1 and hyperphosphorylated during S and G2, the G1-S-specific phosphorylation of pRB becomes an attractive target for the growth-inhibitory activities of TGF-beta 1. However, in TGF-beta 1-treated primary human keratinocytes and in a series of human papilloma virus and SV40 immortalized human keratinocyte cell lines, the phosphorylation status of pRB strictly correlated with cell growth. No evidence was found for a direct effect of TGF-beta 1 on the phosphorylation state of pRB in these cells. It was further demonstrated that synthesis of c-myc protein can be rapidly inhibited by TGF-beta 1 addition throughout G1 and S phases, indicating that the phosphorylation state of pRB, at least as it varies during the cell cycle, does not alter the ability of TGF-beta 1 to suppress c-myc expression.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1991

Factor-binding element in the human c-myc promoter involved in transcriptional regulation by transforming growth factor beta 1 and by the retinoblastoma gene product.

Previous studies have shown that transforming growth factor beta 1 (TGF-beta 1) inhibition of keratinocyte proliferation involves suppression of c-myc transcription, and indirect evidence has suggested that the retinoblastoma gene product (pRB) may be involved in this process. In this study, transient expression of pRB in skin keratinocytes was shown to repress transcription of the human c-myc promoter as effectively as TGF-beta 1. The same c-myc promoter region was required for regulation by both TGF-beta 1 and pRB. These sequences, termed the TGF-beta control element (TCE), lie between positions -86 and -63 relative to the P1 transcription start site. Oligonucleotides containing the TCE bound to several nuclear factors in mobility-shift assays using extracts from cells with or without normal pRB. Binding of some factors was inhibited by TGF-beta 1 treatment of TGF-beta-sensitive but not TGF-beta-insensitive cells. These data indicate that pRB can suppress c-myc transcription and suggest the involvement of cellular factors in addition to pRB in the TGF-beta 1 pathway for the inhibition of c-myc transcription and growth inhibition.

Animals↗
PubMed1991

Telomeric associations and consistent growth factor overexpression detected in giant cell tumor of bone.

Tumor specimens from 15 patients with giant cell tumor (GCT) of bone were cytogenetically analyzed. A subset of five individuals had tumor cells harvested and polyadenylated RNA isolated. Multiple Northern blots were performed utilizing radiolabeled probes for the growth factors TGF beta 1, TGF beta 2, TGF beta 3, and TGF alpha (TGF, transforming growth factor). RNAs from other types of neoplasms and nonneoplastic cells were examined as controls. The most consistent cytogenetic abnormality detected involved multiple telomeric associations (TAs), most frequently involving the terminus of the long arm of chromosome 19 (19q). Northern blot analysis revealed a consistent expression of TGF beta 1 and TGF beta 2 with an inconsistent mRNA expression for the other TGFs. There was a relative overexpression of mRNA for TGF beta 2. The gene location for TGF beta 1 is near the 19q terminus and thus it is speculated that TGF beta may play a role in the neoplastic transformation of GCT.

Adult↗
PubMed1991

Human carcinoid cell production of paracrine growth factors that can stimulate fibroblast and endothelial cell growth.

A serotonin-secreting human pancreatic carcinoid cell line (BON) is demonstrated to express transcripts for all three mammalian types of transforming growth factor beta (TGF beta 1, 2, and 3). Similarly, freshly excised carcinoid tumors from six patients were also found to express mRNA for all three of the type-beta TGFs. Medium conditioned by BON cells had detectable TGF beta activity, although most of the activity was latent as determined by radioreceptor assay with and without prior acid treatment. However, nonactivated BON-conditioned medium stimulated DNA synthesis, soft agar growth, and an increase in TGF beta 1 and fibronectin mRNA expression in AKR-2B fibroblasts. In addition, BON-conditioned medium had a potent endothelial cell growth-stimulatory activity. Since the TGF beta s inhibit growth of endothelial cells, the presence of other growth factors was suspected. TGF alpha, c-sis, and basic fibroblast growth factor transcripts were also found to be expressed by the BON carcinoid cells. These data indicate that multiple peptide growth factors may have a paracrine role in the desmoplastic reaction accompanying carcinoid tumors.

Amino Acid Isomerases↗
PubMed1991

TGF beta 1 and TGF beta 2 are potential growth regulators for low-grade and malignant gliomas in vitro: evidence in support of an autocrine hypothesis.

Low-grade astrocytomas, anaplastic astrocytomas and glioblastomas in vitro were found to ubiquitously produce the mRNA of transforming growth factor-beta (TGF beta). TGF beta 1 and TGF beta 2 mRNA were expressed to a lesser degree among the hyperdiploid malignant gliomas. By radioreceptor assay of conditioned medium, TGF beta was secreted predominantly in latent form, in both latent and active form, or only in active form within a panel of low-grade and malignant gliomas. The TGF beta receptor (types I, II, and III) was evident among the glioma lines. Many near-diploid gliomas were growth-inhibited by TGF beta 1 and TGF beta 2 in vitro. Most hyperdiploid glioblastomas showed a positive mitogenic response to exogenous TGF beta 1 and TGF beta 2. A synergistic or additive mitogenic interaction with epidermal growth factor and insulin was observed among some. Under serum-free conditions, anti-TGF beta antibody neutralized the expected growth-regulatory effect of endogenous TGF beta, thus establishing the specificity of the response in vitro. TGF beta 1 also enhanced the clonogenicity of certain gliomas which had been growth-stimulated in monolayer. Thus, basic elements in support of an autocrine hypothesis have been demonstrated: TGF beta mRNA was expressed among low-grade and malignant gliomas, TGF beta was secreted in latent and/or active form into conditioned media and appeared to serve as an endogenous regulator of glioma proliferation in vitro. The mitogenic response, either positive or negative, correlated with the degree of anaplasia and karyotypic divergence.

Blotting, Northern↗
PubMed1991

Regulation of epithelial proliferation by TGF-beta.

The closely related mammalian TGF-betas (TGF-beta 1, TGF-beta 2 and TGF-beta 3) are potent inhibitors of proliferation of many cell types in vitro. TGF-beta 1 has been demonstrated to be growth inhibitory in vivo for epithelial, endothelial, myeloid and lymphoid cells. Utilizing skin keratinocytes as a model system for studying the mechanism of TGF-beta 1-induced growth inhibition, it has been demonstrated that TGF-beta 1 rapidly inhibits transcription of the c-myc gene. Antisense c-myc oligonucleotides inhibit proliferation of keratinocytes as effectively as does TGF-beta 1, indicating that TGF-beta 1 suppression of c-myc expression is an important component of this growth inhibition. Studies utilizing DNA tumour virus transforming gene constructs have shown that the retinoblastoma gene product, pRb, or a related protein, is needed for TGF-beta 1 suppression of c-myc transcription. Thus, TGF-beta 1 may act through a tumour suppressor gene product, pRb, to suppress transcription of a proto-oncogene, c-myc, and subsequently inhibit cell proliferation.

Animals↗
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

c-myc and pRB: role in TGF-beta 1 inhibition of keratinocyte proliferation.

The TGF-beta s are potent inhibitors of proliferation of most cell types in culture and in vivo. Previous studies have demonstrated that TGF-beta inhibition of skin keratinocyte proliferation involves suppression of c-myc transcription. Evidence derived from use of expression plasmids for certain DNA viral oncoproteins has suggested that the retinoblastoma gene (RB) may be involved in this process. Transient expression of pRB, like TGF-beta 1, in skin keratinocytes represses expression of a human c-myc reporter plasmid, and the same c-myc promoter region (TCE) is required for repression by either TGF-beta 1 or pRB. We showed here that proliferation and c-myc expression in a cell line lacking normal pRB (DU145 human prostate adenocarcinoma cells) are not inhibited by TGF-beta 1. Oligonucleotides containing the TCE were found to bind to a cellular protein of approximately 106 kD (termed p106) in Southwestern assays, utilizing extracts from both the skin keratinocytes and DU145 cells. TCE binding to p106 was diminished by TGF-beta in TGF-beta-sensitive skin keratinocytes but not in TGF-beta-insensitive SV40-transformed keratinocytes. These data support the hypothesis that pRB is required for TGF-beta 1 suppression of c-myc transcription and suggest the involvement of a cellular factor(s) in addition to pRB in the TGF-beta 1 pathway for inhibition of c-myc transcription and growth inhibition.

Animals↗
PubMed1991

Biochemical and biological differences between E7 oncoproteins of the high- and low-risk human papillomavirus types are determined by amino-terminal sequences.

Differences in the biological characteristics of the high-risk human papillomavirus type 16 (HPV-16) and the low-risk HPV-6 E7 proteins were analyzed and shown to correlate with certain biochemical properties. To ascertain which region of E7 conferred these properties, chimeric E7 genes were constructed by the exchange of the amino and carboxyl coding halves of the HPV-6 and HPV-16 E7 genes. The amino-terminal half of E7 determined the affinity for binding to the retinoblastoma protein pRB, the transformation properties, and the ability to abrogate transforming growth factor beta-mediated repression of the c-myc promoter. This region of E7 is therefore responsible for the biological and biochemical differences between the E7 proteins of the low-risk and the high-risk HPVs and consequently is one of the critical determinants distinguishing these two groups of viruses. Transcriptional transactivation of the adenovirus E2 promoter, in contrast, was a property shared by E7 proteins of both low-risk and high-risk HPVs.

Adenovirus Early Proteins↗
PubMed1991

Production of transforming growth factor-alpha by normal rat small intestine.

Transforming growth factor-alpha (TGF-alpha) and epidermal growth factor (EGF) are similar in structure and biological activity. In the present study, the distributions of TGF-alpha mRNA, TGF-alpha immunoreactivity, and TGF-alpha-EGF receptor mRNA were examined in epithelial and nonepithelial compartments of the jejunum, and the effect of TGF-alpha on growth of a jejunal crypt cell line (IEC-6) was determined. Epithelial cells eluted from the rat jejunal cryptvillus axis expressed TGF-alpha mRNA at twofold higher levels in the villus tip than in the crypt and EGF receptor mRNA at sevenfold higher levels in the villus tip. Expression of these two mRNA transcripts in the subepithelium was low. Immunohistochemical staining showed TGF-alpha immunoreactivity predominantly in the epithelium and muscularis. Immunostaining of villus cells was uniform, whereas crypt cells did not stain. IEC-6 cells bound 125I-EGF to a single class of high-affinity (dissociation constant = 833 pM) receptors. EGF and TGF-alpha (10 ng/ml) only modestly stimulated IEC-6 cell growth in the presence of 5% serum but increased expression of the protooncogenes c-jun and c-myc threefold over control cells. These findings suggest that, among the potential physiological roles for TGF-alpha produced by the jejunal epithelium, promotion of cell migration and modulation of fluid and electrolyte transport may be as relatively important as stimulation of cell proliferation.

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↗
PubMed1991

Differential binding of transforming growth factor-beta 1, -beta 2, and -beta 3 by fibroblasts and epithelial cells measured by affinity cross-linking of cell surface receptors.

A murine fibroblast cell line (AKR-2B clone 84A) and an epithelial cell line (BALB/MK) were compared for their ability to bind different transforming growth factor-beta (TGF beta) species. The results of competitive binding assays indicated that the epithelial cells had a higher affinity for TGF beta than the fibroblasts. This difference may be the basis for the sensitivity of epithelial cells to much lower concentrations of TGF beta than fibroblasts. Affinity cross-linking studies showed that both cell types express the three cell surface TGF beta-binding molecules that have been previously described for a variety of cell types. The complexity of these cell surface binding proteins was further evaluated using all possible combinations of radiolabeled ligands in competition with each of the three unlabeled TGF beta species. Differences in the ability of specific TGF beta types to compete with radiolabeled TGF beta 2 for binding to the type I and II receptors were observed, with TGF beta 1 being more potent for epithelial cells, and TGF beta 2 being more potent for fibroblasts. In addition, a difference in the ability of different TGF beta species to compete the [125I]TGF beta 3 from epithelial cell surface receptors was apparent. TGF beta 2 was not able to compete with [125I]TGF beta 3 for binding to the type II receptor at any concentration tested, while TGF beta 1 and TGF beta 3 were about equally potent in competition for this receptor type. These differences in cell surface receptor binding of structurally and biologically similar molecules may reflect different functions for these molecules.

Animals↗
PubMed1991

TGF beta regulation of epithelial cell proliferation: role of tumor suppressor genes.

Transforming growth factor beta 1 (TGF beta 1) is the prototype of a large family of polypeptides involved in growth control, extracellular matrix production, and development. The TGF beta s have marked stimulatory effects on connective tissue formation. They are chemotactic for fibroblasts, indirect mitogens for certain mesenchymal cells and stimulators of extracellular matrix deposition. The TGF beta s are also potent inhibitors of proliferation of most cell types in culture, and in vivo studies have indicated that the predominant effect of TGF beta 1 on cell proliferation is inhibition. We have investigated the mechanism of TGF beta 1 inhibition of skin keratinocyte growth. Earlier studies demonstrated that TGF beta 1 inhibition of keratinocyte proliferation involves suppression of c-myc transcription, and indirect evidence suggested that the product of the retinoblastoma tumor susceptibility gene, pRB, may be involved in this process. More recently, we have shown that transient expression of pRB in skin keratinocytes can repress human c-myc promoter/CAT transcription as effectively as TGF beta 1. The same c-myc promoter region, termed the TGF beta control element (TCE), was required for regulation by both TGF beta 1 and pRB. Oligonucleotides containing the TCE bound to several nuclear factors in mobility shift assays and a cellular protein of approximately 106 kD in Southwestern assays. Binding of these factors could be demonstrated in cells with or without normal pRB, and the binding of some factors was rapidly inhibited by TGF beta 1 treatment of TGF beta-sensitive but not TGF beta-insensitive cells. These data indicate that pRB can function to inhibit c-myc transcription and suggest the involvement of cellular factor(s) in addition to pRB in the TGF beta 1 pathway for suppression of c-myc transcription. Studies with other cell types have shown that another tumor suppressor gene, p53, can also regulate transcription of c-myc in transient assays. Whereas wild type p53 markedly suppressed transcription, four different mutant p53 clones caused transactivation. The data support the hypothesis that pRB and p53 can both cause growth inhibition by blocking the expression of the protooncogene, c-myc, and indicate that tumor suppressor genes may function in the response pathway for diffusible negative growth regulators such as TGF beta.

Animals↗
PubMed1990

Synthesis of transforming growth factor-beta 1 by megakaryocytes and its localization to megakaryocyte and platelet alpha-granules.

We have directly demonstrated that megakaryocytes are a major site of synthesis and storage of transforming growth factor-beta 1 (TGF/beta 1) by combined immunohistochemical, immunocytochemical, and in situ hybridization methods. The presence of TGF/beta 1 messenger RNA (mRNA) in mature megakaryocytes in adult rat spleen and bone marrow (BM) was established by in situ hybridization. Localization of TGF/beta 1 protein to intact alpha-granules of megakaryocytes, its putative storage site, was accomplished in glycol-methacrylate embedded porcine BM with an immunoperoxidase technique and light microscopy. The TGF/beta 1 was sequestered in intracytoplasmic granules in a pattern virtually identical to that of another alpha-granule marker protein, fibrinogen. This observation strongly suggests packaging of TGF/beta 1 into this organelle within megakaryocytes. That TGF/beta 1 mRNA was localized to megakaryocytes suggests that the TGF/beta 1 found in the alpha-granules in platelets originates with megakaryocyte synthesis. The alpha-granule localization of TGF/beta 1, as well as fibrinogen, was also demonstrated in isolated platelets at the ultrastructural level by electronmicroscopy (EM) and postembedding colloidal-gold immunocytochemistry, thus directly demonstrating that alpha-granules are the final storage site for TGF/beta 1 in mature platelets.

Animals↗
PubMed1990

TGF-beta 1 inhibition of c-myc transcription and growth in keratinocytes is abrogated by viral transforming proteins with pRB binding domains.

TGF-beta 1 is demonstrated to inhibit skin keratinocyte proliferation when added during the G1 phase of the cell cycle. Human foreskin keratinocytes transformed with either HPV-16 or -18 or SV40, however, were resistant to the growth inhibitory effects of TGF-beta 1. Since TGF-beta 1 appears to inhibit keratinocyte growth through down-regulation of c-myc, it was hypothesized that these DNA tumor viruses might be modulating the response to TGF-beta 1 via this pathway. Transient expression of proteins HPV-16 E7, adenovirus type 5 E1A, and SV40 large T antigen is demonstrated to block TGF-beta 1 suppression of c-myc transcription. This effect was not observed with DNA tumor virus transforming proteins mutated in their pRB binding domain. These observations indicate that pRB or another protein that interacts with this binding domain mediates TGF-beta 1 regulation of c-myc gene expression and growth inhibition.

Adenovirus Early Proteins↗