Altered metabolic and adhesive properties and increased tumorigenesis associated with increased expression of transforming growth factor beta 1

Transforming growth factor-beta (TGF-beta) is a potent mediator of cell proliferation and extracellular matrix formation, depending on the cell type and the physiological conditions. TGF-beta is usually secreted in a "latent" complex that needs activation before it can exert its effects. Several observations correlate increased expression of TGF-beta 1 with tumorigenesis. To evaluate the physiological relevance of increased TGF-beta 1 synthesis in tumor cells we established cell clones overexpressing TGF-beta 1 and observed the resulting physiological changes in TGF-beta overproducing cells in vitro and in vivo. As a model system we used the human E1A-transformed 293 tumor cells, which are insensitive to the direct growth modulatory effects of TGF-beta. The selection of this cell line allows an assessment of physiological alterations independent of TGF-beta induced proliferative changes. The use of two TGF-beta 1 expression vectors containing either the natural or a modified TGF-beta 1 precursor cDNA permitted the establishment of separate 293 cell lines overexpressing latent or active TGF-beta. Comparison of the resulting changes in glycolytic rate, adhesiveness and integrin and plasminogen activator expression established that, in vitro, both types of clones behaved similarly, indicating that expression of latent TGF-beta induces autocrine changes in the tumor cells and thus suggesting that some level of cell-associated activation occurs. TGF-beta overexpression resulted in an increased metabolic rate due to enhanced glycolysis, a property long associated with tumor cells. This increased glycolysis was not associated with altered proliferation. Cells overexpressing TGF-beta also displayed enhanced fibronectin mRNA and plasminogen activator synthesis and increased adhesiveness in vitro. They showed enhanced survival when plated sparsely on plastic in the absence of serum, and attached more readily to laminin. In addition, synthesis of several beta 1 integrins, in particular the alpha 1/beta 1, alpha 2/beta 1, and alpha 3/beta 1, all of which recognize laminin, were enhanced. Finally, cells overexpressing active TGF-beta, but not latent TGF-beta, also showed increased tumorigenicity in nude mice. Thus, an increase in endogenous TGF-beta synthesis confers several proliferation-independent phenotypic changes which may be of significance for the survival of the tumor cell inoculum or its subsequent growth, and for tumor formation and development. In the case of cells expressing active TGF-beta, the release of active TGF-beta into the vicinity of the tumor cells may also result in a more hospitable environment for tumor growth.     

Demonstration of the capacity of nacre to induce bone formation by human osteoblasts maintained in vitro.

Nacre implanted in vivo in bone is osteogenic suggesting that it may possess factor(s) which stimulate bone formation. The present study was undertaken to test the hypothesis that nacre can induce mineralization by human osteoblasts in vitro. Nacre chips were placed on a layer of first passage human osteoblasts. None of the chemical inducers generally required to obtain bone formation in vitro was added to the cultures. Osteoblasts proliferated and were clearly attracted by nacre chips to which they attached. Induction of mineralization appeared preferentially in bundles of osteoblasts surrounding the nacre chips. Three-dimensional nodules were formed by a dense osteoid matrix with cuboidal osteoblasts at the periphery and osteocytic-like cells in the center. These nodules contained foci with features of mineralized structures and bone-like structures, both radiodense to X-ray. Active osteoblasts (e.m.) with abundant rough endoplasmic reticulum, extrusion of collagen fibrils and budding of vesicles were observed. Matrix vesicles induced mineral deposition. Extracellular collagen fibrils appeared cross-banded and electrodense indicating mineralization. These results demonstrate that a complete sequence of bone formation is reproduced when human osteoblasts are cultured in the presence of nacre. This model provides a new approach to study the steps of osteoblastic differentiation and the mechanisms of induction of mineralization.     

Inhibition by L-arginine of the endothelin-mediated increase in cytosolic calcium in human neutrophils

The effect of endothelin (ET) on the cytosolic-free calcium [(Ca2+]i) changes in polymorphonuclear leukocytes (PMN) from normal humans and Wistar rats was investigated. ET induced a dose-related [Ca2+]i peak. This [Ca2+]i transient was blunted by TMB-8 (10(-5)M) and by Ca(2+)-free EGTA medium, therefore suggesting a role of both intracellular Ca2+ release and Ca2+ influx in the generation of the [Ca2+]i peak. Preincubation of PMN with the nitric oxide (NO)-donor L-arginine (L-Arg) markedly blocked the ET-induced [Ca2+]i transient in an enantiomerically-specific manner. A similar blunting effect of L-Arg on the fMLP (10(-7)M)-induced [Ca2+]i transient was detected. The L-Arg antagonist, NG-monomethyl-L-arginine (L-NMMA), reverted the L-Arg blocking effect on both ET- and fMLP-induced [Ca2+]i transients. These data suggest that ET has a potential role in activating Ca2+ mobilization in PMN, an effect that can be inhibited by L-Arg.     

Sequence of the Streptococcus pneumoniae bacteriophage HB-3 amidase reveals high homology with the major host autolysin.

We have sequenced a DNA fragment containing the pneumococcal bacteriophage HB-3 hbl gene, which codes for the phage lytic amidase. A remarkable nucleotide similarity (87.1%) between the lytA gene, coding for the pneumococcal amidase, the major autolysin of Streptococcus pneumoniae, and the hbl gene was found. This similarity completely disappeared outside the open reading frames coding for both amidases. The hbl gene transformed amidase-deficient strains of S. pneumoniae to the wild-type phenotype, and Southern blotting experiments provided evidence for recombination between donor and recipient genes. A comprehensive evaluation of these and previous results on the peptidoglycan hydrolases of S. pneumoniae and its bacteriophages suggested that recombination mechanisms participate in the evolution of the genes coding for these enzymes.     

Growth of rural and urban children in the Valley of Oaxaca,Mexico

Weight, stature, arm circumference, and the triceps skinfold were measured in 1,410 school children, 6 through 14 years of age, from two urban colonias in the city of Oaxaca de Juarez (n = 479), and from two rural Ladino (n = 467) and two rural Zapotec (n = 464) communities in the Valley of Oaxaca in southern Mexico. Children from rural Ladino communities and urban colonias are significantly taller, heavier, and more muscular than children in rural Zapotec communities. The differences between rural Ladino and urban colonia children favor the former, particularly for weight and stature. These observations thus suggest (1) that children in the rural, indigenous communities in the Valley of Oaxaca are relatively undernourished compared to children in Ladinoized and urban communities, and (2) that rural-to-urban migration does not necessarily result in improved growth status.     

Growth of rural and urban children in the valley of Oaxaca,Mexico

Weight, stature, arm circumference, and the triceps skinfold were measured in 1,410 school children, 6 through 14 years of age, from two urban colonias in the city of Oaxaca de Juarez (n = 479), and from two rural Ladino (n = 467) and two rural Zapotec (n = 464) communities in the Valley of Oaxaca in southern Mexico. Children from rural Ladino communities and urban colonias are significantly taller, heavier, and more muscular than children in rural Zapotec communities. The differences between rural Ladino and urban colonia children favor the former, particularly for weight and stature. These observations thus suggest (1) that children in the rural, indigenous communities in the Valley of Oaxaca are relatively undernourished compared to children in Ladinoized and urban communities, and (2) that rural-to-urban migration does not necessarily result in improved growth status.     

In vitro metabolism of C19 steroids in human endometrium

In order to quantitate the extent of intracellular metabolic conversions of C19 steroids in human endometrium, specimens of proliferative and secretory tissue were superfused at a constant rate with several pairs of labeled compounds at low concentrations. About 16% of dehydroepiandrosterone sulfate interacting with endometrial cells was converted to dehydroepiandrosterone and about 3% of this compound was converted to androstenedione. Androstenedione was reversibly reduced to testosterone and the extent of this conversion was shown to be several-fold higher in secretory than in proliferative tissue. About 1% of testosterone entering the cells was reduced to 5 alpha-dihydrotestosterone. These results demonstrate that the conversion of the main circulating C19 steroids in women, i.e. dehydroepiandrosterone sulfate and androstenedione, to 5 alpha-dihydrotestosterone, the compound considered to be the true intracellular androgen, is very small. In contrast, formation of testosterone from androstenedione is extensive and increases during the luteal phase under the influence of progesterone, a hormone known to stimulate the activity of 17 beta-hydroxysteroid dehydrogenase in human endometrium.     

Nitrofurantoin prompts the stringent response in Bacillus subtilis

Nitrofurantoin causes the stringent response in Bacillus subtilis. After exposure of a stringent strain to this drug, the intracellular concentrations of guanosine 3'-diphosphate 5'-diphosphate (ppGpp), guanosine 3'-diphosphate 5'-triphosphate (pppGpp) and ATP increased, while that of GTP decreased. In a relaxed strain no accumulation of ppGpp or pppGpp was observed, but both GTP and ATP declined after the addition of nitrofurantoin. Protein synthesis was equally sensitive to nitrofurantoin in both the stringent and relaxed strains, but the drug inhibited RNA accumulation only in the stringent strain, not in the relaxed strain. Nitrofurantoin also caused the accumulation of ppGpp in Escherichia coli and Serratia marcescens.