Modulation of skin cell functions by transforming growth factor-beta1 and ACTH after ultraviolet irradiation

A new method was developed for binding poly-(ethylene oxide) (PEO) to polymer surfaces that involves the use of electron beam irradiation in two steps. In the first, methacrylic acid was grafted and polymerized to a polymer surface, changing it from hydrophobic to hydrophilic. Exposure of this surface to aqueous PEO solutions resulted in strong hydrogen bonding of the PEO, which was covalently grafted in a second radiation step. The PEO grafts were stable; they could not be removed with extensive washing with water, soaking in basic solution, or gentle mechanical scraping. Both monolayers and multilayers of PEO were formed. The density of the monolayers were found to have little dependence on the molecular weight or concentration of the PEO solution; multilayers could be controlled by varying the viscosity of the PEO solution and the method of application. The PEO-grafted monolayers were tested for their ability to prevent protein adsorption of cytochrome-c, albumin, and fibronectin. Monolayers of star PEO were the most effective, at best showing a 60% decrease in adsorption from untreated controls. One million molecular wight linear PEO monolayers were almost as effective as star monolayers, and 35,000 g/mol linear PEO was bound too closely to the surface, owing to its small size, to have much impact in preventing protein adsorption. The reason for the continued protein adsorption was believed to be due to a close grafting of the PEO to the surface, as well as the grafted methacrylic acid chains being long enough to extend through the PEO monolayer, thus being accessible on the surface.     

Mandibular reconstruction with transforming growth factor-beta1

This is a case report of a 43-year-old woman who received a transplant for end-stage liver disease due to hereditary hemorrhagic telangiectasia and fibropolycystic liver disease. This is an uncommon association of two autosomal-dominant conditions with defined genetic and molecular defects. The liver showed extensive vascular malformations of arteries and veins as well as telangiectasia and fibrosis. In addition, there were cystically dilated ducts containing inspissated bile and extensive von Meyenburg complexes. This case raises interesting questions about the possible relationship of these genes and their gene products, both of which are related to cell-matrix interactions and are strongly associated with blood vessels, one of them being expressed on endothelial cells and the other being developmentally important in blood vessels.     

Growth-inhibitory effects of transforming growth factor-beta 1 on myeloid leukemia cell lines

Transforming growth factor-beta1 is a pleiotropic cytokine involved in a variety of biological processes in both transformed and normal cells, including regulation of cellular proliferation and differentiation; its predominant action on hematopoietic cells is to inhibit cell growth. We used growth factor-dependent cell lines to assess TGF-beta1 effects on human myeloid leukemia cell growth. While four lines were completely or predominantly resistant, TGF-beta1 inhibited effectively, albeit to various extents, the growth of 12 other cell lines. This effect was dose dependent and specific, because a neutralizing anti-TGF-beta1 antibody prevented TGF-beta1-induced growth suppression. In the present system, basic fibroblast growth factor, known as an antagonist of TGF-beta1 counteracting its inhibitory effects, did not abrogate the suppressive effects of TGF-beta1. Other growth-stimulatory cytokines negated the TGF-beta1-induced inhibition in several cell lines, again to various extents. When proliferation was enhanced by growth-promoting cytokines (e.g. granulocyte-macrophage colony-stimulating factor, GM-CSF, stem cell factor, SCF, or PIXY-321), some previously TGF-beta1-sensitive cell lines acquired cellular resistance toward TGF-beta1-mediated growth suppression, whereas four other cell lines remained susceptible to TGF-beta1 growth inhibition despite possible counteraction by other cytokines. Thus, three growth response patterns to TGF-beta1 were seen: (1) constitutive resistance; (2) factor-dependent relative resistance; and (3) sensitivity to growth inhibition indifferent to counteracting cytokines. In the latter case, TGF-beta1 did not downregulate expression of one specific growth factor receptor. These studies indicate that human myeloid leukemia cells, represented here by leukemia cell lines as model systems, exhibit heterogeneous growth responses to TGF-beta1; its inhibitory effects can be modulated or completely alleviated by positive antagonistic cytokines. The availability of TGF-beta1-susceptible and -refractory cell lines allows for detailed investigations on the mechanisms of these regulatory pathways, the nature of TGF-beta1-resistance, and the possible contribution of acquired TGF-beta1-resistance to disease progression.     

Expression of transforming growth factor-beta 1 messenger ribonucleic acid and the modulation of deoxyribonucleic acid synthesis by transforming growth factor-beta 1 in human endometrial cells

OBJECTIVE: The purpose of this study was (1) to evaluate the potential sites of transforming growth factor-beta 1 synthesis in human endometrium by analyzing separated endometrial glands and stromal cells for transforming growth factor-beta 1 messenger ribonucleic acid by Northern analysis of total ribonucleic acid and (2) to investigate the effects of transforming growth factor-beta 1 on deoxyribonucleic acid synthesis in endometrial epithelial and stromal cells in culture. STUDY DESIGN: Endometrial glands and stroma from proliferative and secretory endometrium were isolated after collagenase treatment of endometrial tissue minces and were analyzed for transforming growth factor-beta 1 messenger ribonucleic acid by Northern analysis. We studied the effects of estradiol-17 beta and transforming growth factor-beta 1 on deoxyribonucleic acid synthesis in endometrial epithelium and transforming growth factor-beta 1 on stromal cells in culture by evaluating tritiated thymidine incorporation into trichloroacetic acid-precipitable material. RESULTS: Transforming growth factor-beta 1 messenger ribonucleic acid was detected for Northern analysis in separated endometrial stromal cells in levels that were greatest during the secretory phase and in greater levels than in epithelial cells from that same tissue. Transforming growth factor-beta 1 messenger ribonucleic acid in glandular epithelium in culture was not increased to detectable levels by treatment with transforming growth factor-beta 1. Deoxyribonucleic acid synthesis in endometrial glandular epithelium was inhibited by transforming growth factor-beta 1, but transforming growth factor-beta 1 stimulated deoxyribonucleic acid synthesis in endometrial stromal cells in culture. After treatment for 5 days with estradiol-17 beta (10(-8) mol/L), deoxyribonucleic acid synthesis in endometrial glands in culture was decreased by 40%. Transforming growth factor-beta 1 (1 ng/ml) did not alter this effect of estradiol-17 beta on deoxyribonucleic acid synthesis. CONCLUSIONS: Transforming growth factor-beta 1 acts to decrease deoxyribonucleic acid synthesis in epithelial cells and to increase it in stromal cells isolated from human endometrium and maintained in monolayer culture. Transforming growth factor-beta 1, potentially of stromal cell origin, could participate in the regulation of endometrial cell proliferation and differentiation in vivo.     

Probenecid inhibits transforming growth factor-beta 1 induced pyrophosphate elaboration by chondrocytes

OBJECTIVE: The elaboration of excess extracellular inorganic pyrophosphate (ePPi) by cartilage contributes to calcium pyrophosphate dihydrate (CPPD) crystal deposition disease. Transforming growth factor-beta 1 (TGF beta 1) is the only defined physiologic stimulant of cartilage ePPi elaboration. The mechanism of ePPi generation by chondrocytes is unknown, but current evidence suggests that TGF beta 1 induced ePPi is made intracellularly. An active transport mechanism such as an anion transporter would then be necessary to export ePPi to the matrix where crystals form. We determined the effect of probenecid (PB), an anion transport inhibitor, on TGF beta 1 induced ePPi elaboration. METHODS: Porcine hyaline articular chondrocytes in high density monolayer cultures were exposed to serum-free media with and without TGF beta 1 and/or PB. ePPi was measured in the media after 48-96 h of exposure. Cell injury was measured by examining the release of 3H-deoxyglucose from chondrocytes. The activity of the ePPi generating ectoenzyme nucleoside triphosphate pyrophosphohydrolase (NTPPPH) and media lactate concentrations were measured with standard colorimetric assays. As PB may inhibit phosphodiesterase (PDE), its effects on ePPi generation were compared with isobutylmethylxanthine (IBMX), a specific PDE inhibitor. RESULTS: PB inhibited TGF beta 1 induced ePPi elaboration by chondrocytes. PB did not cause membrane injury or decrease NTPPPH activity. Lactate production was decreased by PB but did not correlate with the effects of PB on ePPi elaboration. IBMX did not inhibit TGF beta 1 effect on ePPi elaboration. CONCLUSION: PB blocks TGF beta 1 induced ePPi elaboration. This effect is independent of cell membrane injury, decreased NTPPPH activity, or PDE inhibition. Our data implicate a role for anion transport in TGF beta 1 induced ePPi elaboration, and suggest a potential therapy for CPPD disease.     

Immunolocalization of transforming growth factor-beta 1 and transforming growth factor-beta 2 in the mouse ovary during gonadotrophin-induced follicular maturation

An immunohistochemical approach was utilized to evaluate the cellular distribution of transforming growth factor-beta 1 (TGF beta 1) and transforming growth factor beta 2 (TGF beta 2) at different stages of follicle development in the prepubertal mouse ovary under the following conditions: (i) after pregnant mare's serum gonadotrophin (PMSG) treatment; (ii) after PMSG and human chorionic gonadotrophin (HCG) treatment; (iii) after PMSG and HCG treatment plus mating. In the immature ovary, TGF beta 1 and TGF beta 2 immunoreactivities are localized in theca and granulosa cells and in oocytes. After PMSG treatment, TGF beta 1 and TGF beta 2 immunoreactivities are localized in granulosa cells; in addition, TGF beta 2 staining is noted in the matrix surrounding antral cells. Staining for both TGR beta 1 and TGF beta 2 drops in the theca but persists in the oocyte. PMSG plus HCG treatment results in a significant increase in TGF beta 1 and TGF beta 2 immunoreactivity in the theca and in the maintenance of TGF beta 1 staining in both basal granulosa cells and cumulus cells whereas TGF beta 2 immunoreactivity is essentially localized in the matrix surrounding cumulus cells. Staining for TGF beta 1 and TGF beta 2 persists in the oocyte. Following PMSG plus HCG treatment and mating, TGF beta 1 immunoreactivity is localized in the luteal cells of corpora lutea and TGF beta 2 shows a similar localization pattern. This study provides evidence that TGF beta 1 and TGF beta 2 peptides are expressed in specific cell types during induced follicular maturation in the mouse ovary.     

Modulation of collagen gene expression by cytokines: stimulatory effect of transforming growth factor-beta1, with divergent effects of epidermal growth factor and tumor necrosis factor-alpha on collagen type I and collagen type IV

Transforming growth factor-beta1 (TGF-beta1) is well recognized as a potent mediator of both fibrillar (collagen type I) and basement membrane (collagen type IV) production. However, tissue injury is characterized by the concomitant expression of many cytokines and/or growth factors in addition to TGF-beta1, and the ultimate extent of extracellular-matrix (ECM) deposition may reflect the interacting effects of TGF-beta1 and these other cytokines and/or growth factors. We, therefore, sought to determine whether other cytokines and/or growth factors, known to be produced after tissue injury, are capable either alone or in combination with TGF-beta1 of modulating collagen gene expression. Collagen type I and collagen type IV gene expression was assessed in NIH-3T3 cells, a murine fibroblast-like cell line that responds to TGF-beta1, with increases in both collagen type I and collagen type IV production. TGF-beta1 coordinately induced production of collagen type IV messenger ribonucleic acid (mRNA) to a level 3.8-fold above its baseline value (p < 0.001) and collagen type I mRNA to a level 2.6-fold above its baseline value (p < 0.001). Of the other cytokines and/or growth factors tested, only epidermal growth factor (EGF) had significant effects on collagen mRNA expression. We report the novel observation that EGF significantly induced collagen type IV mRNA (3.0-fold; p < 0.001) but did not alter collagen type I mRNA expression. Platelet-derived growth factor (PDGF), basic fibroblast growth factor (bFGF), tumor necrosis factor-alpha (TNF-alpha), interleukin-1 (IL-1), and insulin-like growth factor-1 (IGF-1) did not alter the expression of mRNA for collagen type IV or collagen type I. Addition of TGF-beta1 to cytokine- and/or growth factor-treated cells increased both collagen type IV and collagen type I mRNA levels. However, collagen type IV mRNA levels were similar in cultures given TGF-beta1 alone and cultures given TGF-beta1 with other cytokines and/or growth factors; there were no additive, synergistic, or antagonistic effects after coadministration of TGF-beta1 and other cytokines and/or growth factors. With regard to collagen type I mRNA expression, all cytokines and/or growth factors tested, with the exception of TNF-alpha, had no effect on collagen type I mRNA levels in TGF-beta1-treated cultures. Importantly, TNF-alpha antagonized the stimulatory effect of TGF-beta1 on collagen type I mRNA levels. These observations support a dominant role for TGF-beta1 in stimulating coordinate expression of collagen type I and collagen type IV mRNAs by NIH-3T3 cells; EGF and TNF-alpha are capable of inducing divergent expression of the genes for these two types of collagen.     

Down-regulation of murine fibrosarcoma transforming growth factor-beta 1 expression by interleukin 7

BACKGROUND: Cytokine genes encode proteins that modulate immune system responses. Modification of tumor cells by the introduction of cytokine genes has been used as a strategy to augment host immunity. Interleukin 7 (IL-7) gene transfer enhances the immune response to tumor cells and can result in tumor regression. Transforming growth factor-beta 1 (TGF-beta 1) is a potent immunosuppressive cytokine produced by many tumors. We have previously reported that recombinant IL-7 decreases the expression of TGF-beta 1 by murine macrophages. PURPOSE: This study investigates the inhibition of tumor-derived TGF-beta 1 production as a possible mechanism for the enhanced antitumor immunity that accompanies IL-7 gene transfer. METHODS: A fibrosarcoma cell line (FSA-JmIL-7) genetically modified to produce IL-7 was used to evaluate the effects of IL-7 on tumor production of TGF-beta 1. The control cell line (FSA-Jneo) originated from the same parental fibrosarcoma cell line (FSA) and was produced by transduction with the same retroviral vector without the IL-7 gene. FSA-Jneo and FSA-JmIL-7 tumor cells were evaluated for the expression of TGF-beta 1 messenger RNA (mRNA). To determine if the observed change in TGF-beta 1 mRNA was associated with an alteration in protein secretion, we compared supernatants from tumor cell cultures for TGF-beta 1 production. Specific anti-TGF-beta 1 monoclonal antibody (MAb) was used to confirm the role of TGF-beta 1 in these assays. RESULTS: Compared with FSA parental and FSA-Jneo cells, FSA-JmIL-7 cells expressed TGF-beta 1 mRNA at a lower level. Compared with supernatants from FSA-Jneo cells, FSA-JmIL-7 supernatants contained consistently lower levels of TGF-beta 1 activity (P < .05). In addition, FSA-Jneo supernatants suppressed lymphocyte proliferation to a significantly greater degree than supernatants from FSA-JmIL-7 cells (P < .05). Studies with anti-TGF-beta 1 MAb added to the supernatants confirmed the role of TGF-beta 1 in inhibition of lymphocyte proliferation. CONCLUSION: These findings suggest that IL-7 gene transfer inhibits the production of TGF-beta 1 by tumor cells and thus may enhance the efficacy of the host's antitumor immune response. IMPLICATION: The regulation of endogenous tumor-derived cytokines in response to cytokine gene transfer may contribute to altered immune responses in the tumor microenvironment and thus may be an important additional parameter to assess in gene therapy.     

Autoregulation of periodontal ligament cell phenotype and functions by transforming growth factor-beta1

During orthodontic tooth movement, mechanical forces acting on periodontal ligament (PDL) cells induce the synthesis of mediators which alter the growth, differentiation, and secretory functions of cells of the PDL. Since the cells of the PDL represent a heterogeneous population, we examined mechanically stress-induced cytokine profiles in three separate clones of human osteoblast-like PDL cells. Of the four pro-inflammatory cytokines investigated, only IL-6 and TGF-beta1 were up-regulated in response to mechanical stress. However, the expression of other pro-inflammatory cytokines such as IL-1 beta, TNF-alpha, or IL-8 was not observed. To understand the consequences of the increase in TGF-beta1 expression following mechanical stress, we examined the effect of TGF-beta1 on PDL cell phenotype and functions. TGF-beta1 was mitogenic to PDL cells at concentrations between 0.4 and 10 ng/mL. Furthermore, TGF-beta1 down-regulated the osteoblast-like phenotype of PDL cells, i.e., alkaline phosphatase activity, calcium phosphate nodule formation, expression of osteocalcin, and TGF-beta1, in a dose-dependent manner. Although initially TGF-beta1 induced expression of type I collagen mRNA, prolonged exposure to TGF-beta1 down-regulated the ability of PDL cells to express type I collagen mRNA. Our results further show that, within 4 hrs, exogenously applied TGF-beta1 down-regulated IL-6 expression in a dose-dependent manner, and this inhibition was sustained over a six-day period. In summary, the data suggest that mechanically stress-induced TGF-beta1 expression may be a physiological mechanism to induce mitogenesis in PDL cells while down-regulating its osteoblast-like features and simultaneously reducing the IL-6-induced bone resorption.     

Sequential effects of high glucose on mesangial cell transforming growth factor-beta 1 and fibronectin synthesis

BACKGROUND: Transforming growth factor (TGF)-beta is recognized as the final common mediator of the principal lesions of diabetic nephropathy such as renal hypertrophy and mesangial expansion. To gain better understanding of the temporal relationships between high glucose (HG) and mesangial cell (MC) TGF-beta 1 synthesis and between TGF-beta 1 and extracellular matrix (ECM) synthesis, the present study examined early and sequential effects of HG on TGF-beta 1 and fibronectin (FN) mRNA expression and protein synthesis. METHODS: Confluent primary rat MC was stimulated with 5.6 (control) or 30 (high) mM glucose after synchronizing the growth by incubation with serum-free media for 48 hours. RESULTS: Mesangial cell TGF-beta 1 mRNA expression increased significantly in six hours and continued to increase until 48 hours in response to HG. The level of TGF-beta 1 mRNA was 1.5-fold higher than that of control glucose at six hours and 1.8-fold at 48 hours. TGF-beta activity in heat-activated conditioned media under HG increased 1.5- and 1.6-fold at 24 and 48 hours, respectively, compared to control glucose. FN mRNA increased significantly at 24 and 48 hours and 1.4-fold that of control glucose at both time points. FN protein also increased 1.5-fold that of control glucose at 48 hours. Anti-TGF-beta antibody completely abolished HG-induced FN synthesis. CONCLUSIONS: The present finding demonstrate that HG stimulates TGF-beta 1 very early and prior to FN production and that HG-induced FN production is mediated by TGF-beta. This finding is consistent with the view that TGF-beta mediates increased ECM accumulation by MC under high glucose conditions.     

Hepatocyte growth factor releases epithelial and endothelial cells from growth arrest induced by transforming growth factor-beta1

Human lung fibroblasts and Mv1Lu mink lung epithelial cells were used as a model to study the role of extracellular matrix in epithelial-mesenchymal interactions. Extracellular matrices of fibroblasts were found to contain growth promoting activity that reduced the sensitivity of Mv1Lu cells to the growth inhibitory effects of transforming growth factor-beta (TGF-beta). The majority of the activity was identified as hepatocyte growth factor/scatter factor (HGF) by inhibition with specific antibodies and by reconstitution of the effect by recombinant HGF. HGF induced cell proliferation when contact-inhibited Mv1Lu cells were trypsinized and plated in the presence of TGF-beta1. The effect was valid also in assays where Madin-Darby canine kidney epithelial cells or bovine capillary endothelial cells were used. The multiplication of chronically TGF-beta1 inhibited Mv1Lu cells was also induced by HGF. In addition, HGF induced anchorage independent growth of Mv1Lu cells that was refractory to TGF-beta1 growth inhibition. Immunoprecipitation analysis indicated that HGF prevented the suppression of Cdk4 and Cdk2, but not the induction of p21, by TGF-beta1. Since both TGF-beta1 and HGF require proteolysis for activation, the results imply that proteolytic activity of epithelial and endothelial cells directs their responses to signals from mesenchymal-type extracellular matrices, and that during development, matrix-bound growth and invasion promoting and suppressing factors are activated in a coordinated manner.     

IFN-gamma inhibits the production of latent transforming growth factor-beta1 by mouse inflammatory macrophages

Transforming growth factor (TGF)-beta is a multifunctional cytokine, which in mammals exists in three isoforms (TGF-beta1, 2 and 3). It is synthesized by a variety of cells including macrophages, and exerts potent immunoregulatory effects such as the inhibition of Th1 development and the suppression or reversal of IFN-gamma-induced macrophage activation. In this study we analyzed the effect of IFN-gamma on the production of TGF-beta1 by thioglycolate-elicited mouse peritoneal macrophages under serum-free conditions. Untreated macrophages released TGF-beta1 in its latent form, which became detectable in a capture ELISA specific for active TGF-beta1 after acid activation of the culture supernatants. Treatment with IFN-gamma reduced the amount of latent TGF-beta1 in the culture supernatants in a dose-dependent fashion. The effect of IFN-gamma was confirmed by a newly developed Western blot system for the detection of mouse TGF-beta1 protein. IFN-gamma only weakly (16-24 %) reduced the levels TGF-beta1 mRNA at early and late time points of stimulation, and no evidence was obtained that IFN-gamma suppresses the secretion of latent TGF-beta1. Thus, inhibition of TGF-beta1 production by IFN-gamma is most likely due to decreased synthesis and/or stability of the TGF-beta1 protein, and might be important for the generation of fully activated macrophages and a Th1 response.     

Molecular mechanisms of transforming growth factor-beta signaling

The present experiments examined the effects of posttraining intrahippocampal injections of the degradative enzyme-resistant methylcarbamyl analog of the bioactive phospholipid platelet-activating factor (mc-PAF) and the platelet-activating factor (PAF) receptor antagonists BN52021 and BN 50730 on memory in male Long-Evans rats trained in a hidden platform version of the Morris water maze. Following an eight-trial training session, rats received a unilateral intrahippocampal injection of mc-PAF (0.5, 1.0, or 2.0 microgram/0.5 microliter), lyso-PAF (1.0 microgram/0.5 microliter), the cell surface PAF receptor antagonist BN 52021 (0.25, 0.5, or 1.0 micrigram/0.5 microliter/, the intracellular PAF receptor antagonist BN 50730 (2.0, 5.0, or 10.0 microgram/0.5 microliter), or vehicle (50% DMSO in 0.9% saline; 0.5 microliter). On a retention test conducted 24 h after training, the escape latencies of rats administered mc-PAF (1.0 or 2.0 microgram) were significantly lower than those of the vehicle-injected controls, demonstrating a memory-enhancing effect of mc-PAF. Injections of lyso-PAF, a structurally similar metabolite of PAF, had no influence on memory, indicating that the memory-enhancing effect of mc-PAF is not caused by membrane perturbation by the phospholipid. The retention test escape latencies of rats administered BN 52021 (0.5 microgram) and BN 50730 (5.0 or 10 microgram) were significantly higher than those of the controls, indicating a memory impairing effect of both PAF antagonists. When mc-PAF, BN 52021, or BN 50730 was administered 2 h posttraining, no effect on retention was observed, indicating a time-dependent effect of the neuroactive substances on memory storage. The findings suggest a role for endogenous PAF in hippocampal-dependent memory processes.     

Latent transforming growth factor-beta binding protein domains involved in activation and transglutaminase-dependent cross-linking of latent transforming growth factor-beta

Transforming growth factor-beta (TGF-beta) is secreted by many cell types as part of a large latent complex composed of three subunits: TGF-beta, the TGF-beta propeptide, and the latent TGF-beta binding protein (LTBP). To interact with its cell surface receptors, TGF-beta must be released from the latent complex by disrupting noncovalent interactions between mature TGF-beta and its propeptide. Previously, we identified LTBP-1 and transglutaminase, a cross-linking enzyme, as reactants involved in the formation of TGF-beta. In this study, we demonstrate that LTBP-1 and large latent complex are substrates for transglutaminase. Furthermore, we show that the covalent association between LTBP-1 and the extracellular matrix is transglutaminase dependent, as little LTBP-1 is recovered from matrix digests prepared from cultures treated with transglutaminase inhibitors. Three polyclonal antisera to glutathione S-transferase fusion proteins containing amino, middle, or carboxyl regions of LTBP-1S were used to identify domains of LTBP-1 involved in cross-linking and formation of TGF-beta by transglutaminase. Antibodies to the amino and carboxyl regions of LTBP-1S abrogate TGF-beta generation by vascular cell cocultures or macrophages. However, only antibodies to the amino-terminal region of LTBP-1 block transglutaminase-dependent cross-linking of large latent complex or LTBP-1. To further identify transglutaminase-reactive domains within the amino-terminal region of LTBP-1S, mutants of LTBP-1S with deletions of either the amino-terminal 293 (deltaN293) or 441 (deltaN441) amino acids were expressed transiently in CHO cells. Analysis of the LTBP-1S content in matrices of transfected CHO cultures revealed that deltaN293 LTBP-1S was matrix associated via a transglutaminase-dependent reaction, whereas deltaN441 LTBP-1S was not. This suggests that residues 294-441 are critical to the transglutaminase reactivity of LTBP-1S.     

Smad1 recognition and activation by the ALK1 group of transforming growth factor-beta family receptors

Two structural elements, the L45 loop on the kinase domain of the transforming growth factor-beta (TGF-beta) family type I receptors and the L3 loop on the MH2 domain of Smad proteins, determine the specificity of the interactions between these receptors and Smad proteins. The L45 sequence of the TGF-beta type I receptor (TbetaR-I) specifies Smad2 interaction, whereas the related L45 sequence of the bone morphogenetic protein (BMP) type I receptor (BMPR-I) specifies Smad1 interactions. Here we report that members of a third receptor group, which includes ALK1 and ALK2 from vertebrates and Saxophone from Drosophila, specifically phosphorylate and activate Smad1 even though the L45 sequence of this group is very divergent from that of BMPR-I. We investigated the structural elements that determine the specific recognition of Smad1 by ALK1 and ALK2. In addition to the receptor L45 loop and the Smad1 L3 loop, the specificity of this recognition requires the alpha-helix 1 of Smad1. The alpha-helix 1 is a conserved structural element located in the vicinity of the L3 loop on the surface of the Smad MH2 domain. Thus, Smad1 recognizes two distinct groups of receptors, the BMPR-I group and the ALK1 group, through different L45 sequences on the receptor kinase domain and a differential use of two surface structures on the Smad1 MH2 domain.     

Dehydroepiandrosterone induces the transforming growth factor-beta production by murine macrophages

Dehydroepiandrosterone(DHEA), a predominant androgen secreted by the adrenal cortex, and dehydroepiandrosterone sulfate (DHEAS). Its predominant form in serum, were investigated for their role in the regulation of transforming growth factor-beta (TGF beta) production by murine macrophages. Using a bioassay based on the growing inhibition to Mv-1-Lu cells and RT-PCR analysis, the effect of DHEA and DHEAS on the TGF-beta production and gene expression was studied. Results suggested that DHEA at relatively high concentration (10 microM) significantly induced TGF-beta secretion by both peritoneal cells and P388D1 macrophage-like cells. For the cells treated with DHEAS, no significant increase in TGF-beta secretion was found statistically. Result of RT-PCR confirmed the observation that cDNA from the cells pretreated with DHEA generated a significant amount of amplicons but cDNA samples obtained from both control cells and DHEAS-treated cells showed relatively weak signals. In a quantitative RT-PCR analysis, both DHEAS-treated cells and control cells failed to compete with internal standards and failed to produce any detectable amplicons. Dexamethasone, one of the commonly used glucocorticoids, induced an increase in TGF-beta secretion and in mRNA level. Dexamethasone and DHEA failed to show a synergistic effect on the DHEA-induced increase in TGF-beta secretion and gene expression. The biological significance for DHEA to act as a positive stimulator for TGF-beta production and its role in glucocorticoid-mediated immunoregulation needs to be further delineated.     

Retinoids potentiate transforming growth factor-beta activity in bovine endothelial cells through up-regulating the expression of transforming growth factor-beta receptors

The present paper reports data on secular trends in the stature of Brazilian Navy recruits born from 1940 to 1965. The final sample included 3269 individuals aged 18.00-18.99. Statistics performed were: ANOVA (one-way and two-way), Sheffe test, simple linear regression between stature and year of birth, and multiple linear regression adjusting for level of schooling (beta coefficient) and chi-square. Results indicated a progressive growth trend in stature of 0.1 cm/yr. for the country as a whole. The trend was also observed for nearly all regions and two out of three levels of schooling and can be explained by improvement in some of the country's health indicators. One important characteristic was a higher level of schooling observed among Navy recruits, suggesting that these individuals represent a highly select group, and that therefore data on the Navy cannot be applied directly to the Brazilian population as a whole.     

Role of transforming growth factor-beta1 in the pathogenesis of moyamoya disease

N-Nitroso propoxur (NP) can be synthesized from a widely used N-methylcarbamate insecticide, propoxur, in vitro in the laboratory. Because of the extensive use of aerosol propoxur, the adverse effect on cells of respiratory origin is worth elucidating. In this report, two mammalian cell cultures from respiratory tissues [a hamster lung fibroblast, V79, and a primary rat tracheal epithelial cell (RTE)], were used to investigate the genotoxicity of propoxur and NP. NP was more cytotoxic than propoxur, with LC50s (20 and six times smaller, respectively in V79 and RTE cells. NP significantly induced sister chromatid exchange (> or = 0.01 microg/ml), chromosome aberration (> or = 2.5 microg/ml) and hprt gene mutation (> or = 0.5 microg/ml) in V79 cells, and cell transformation (> or = 0.2 microg/ml) in RTE cells. Results of chromosome aberration and hprt gene mutation indicated that the major pre-mutagenic lesion induced by NP must be the O6-methylguanine adduct, which frequently mispairs with thymine and thus gives rise to a GC-->AT transition. Propoxur was not mutagenic to either type of cells. However, it inhibited gap-junctional intercellular communication in V79 cells, which indicates that propoxur could act through some epigenetic mechanisms, such as tumor promotion or cell proliferation, in the multiple process of chemical carcinogenesis.