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PubMed2004

The response regulator PhoP of Yersinia pseudotuberculosis is important for replication in macrophages and for virulence.

Yersinia pestis and Yersinia pseudotuberculosis are closely related facultative intracellular pathogens. The response regulator PhoP was previously shown to be important for Y. pestis survival in macrophages and for virulence in a murine bubonic plague infection assay. Here the importance of PhoP for Y. pseudotuberculosis pathogenesis was investigated. Y. pseudotuberculosis phoP mutants were unable to replicate in low-Mg(2+) medium or in macrophages. phoP(+) Y. pseudotuberculosis strains initiated replication in macrophages after a lag period of approximately 5 h, as shown by fluorescence microscopy and viable count assays. Y. pseudotuberculosis phoP mutants died at a low rate in macrophages; there was no decrease in viability over the first 5 h of infection, and there was a 10-fold decrease in viability between 5 and 24 h of infection. Trafficking of phagosomes containing phoP(+) or phoP mutant Y. pseudotuberculosis was studied by using immunofluorescence microscopy and cathepsin D as a marker for lysosomes. Phagosomes containing phoP mutant Y. pseudotuberculosis acquired cathepsin D at a higher rate than phagosomes containing phoP(+) bacteria. However, the increased rate of marker acquisition for phagosomes containing mutant bacteria was only evident approximately 5 h after infection, suggesting that phoP mutants are able to retard phagosome maturation during the lag phase of intracellular growth. The results obtained with a Y. pestis phoP mutant were similar to those described above, except that the rates of intracellular killing and trafficking to cathepsin D-positive vacuoles were significantly higher. A Y. pseudotuberculosis phoP mutant was 100-fold less virulent than the wild-type strain in a murine intestinal infection model, suggesting that survival and replication in macrophages are important for Y. pseudotuberculosis pathogenesis.

Amino Acid Sequence↗

PubMed1998

Comparison of the ABI 7700 system (TaqMan) and competitive PCR for quantification of IS6110 DNA in sputum during treatment of tuberculosis.

Mycobacterium tuberculosis can persist in sputum for long periods of time after the initiation of antituberculosis chemotherapy. The purpose of this study was to determine whether quantitative estimates of M. tuberculosis DNA in sputum correlate with the numbers of viable bacilli and thus measure the therapeutic response of patients during treatment. Two methods of M. tuberculosis DNA quantification were examined by using DNA isolated from sputum specimens serially collected during the course of chemotherapy. A competitive PCR assay was compared to an automated system of real-time quantification with the ABI Prism 7700 Sequence Detection System (TaqMan). The ABI 7700 system uses standard PCR in conjunction with a fluorogenic probe in which the intensity of fluorescence is proportional to the amount of target DNA present. The results showed that both PCR systems are reproducible and accurate. The amounts of M. tuberculosis DNA quantified in sputum corresponded well with the numbers of acid-fast bacilli (AFB) counted by microscopy. Before initiation of antituberculosis therapy, measures of AFB, M. tuberculosis DNA, and cultivable bacilli were similar, suggesting that quantification of DNA is a good method for measuring the initial bacillary load. However, the rate of disappearance of both AFB and M. tuberculosis DNA did not correlate with the decline in cultivable bacilli in the specimen; therefore, these tests are not appropriate for monitoring treatment efficacy.

Antitubercular Agents↗

PubMed1991

Analysis of premature termination in c-myc during transcription by RNA polymerase II in a HeLa nuclear extract.

Transcriptional regulation of the human c-myc gene, an important aspect of cellular differentiation, occurs in part at the level of transcript elongation. In vivo, transcriptional arrest, due to either pausing or termination, occurs near the junction between the first exon and first intron and varies with the growth state of the cell. We have tested the transcription of c-myc templates in HeLa nuclear extracts. We did not observe significant arrest under standard conditions, but we found that a considerable fraction of transcription complexes stopped at the c-myc TII site (just past the first exon-intron junction) when the KCl concentration was raised to 400 mM during elongation. Transcriptional arrest at TII also was observed at KCl concentrations as low as 130 mM and when potassium acetate or potassium glutamate was substituted for KCl. Under these conditions, arrest occurred at the TII site when transcription was initiated at either the c-myc P2 promoter or the adenovirus 2 major late promoter. Further, the TII sequence itself, in forward but not reverse orientation, was sufficient to stop transcription in a HeLa nuclear extract. By separating the TII RNA from active transcription complexes by using gel filtration, we found that arrest at TII at 400 mM KCl resulted in transcript release and thus true transcriptional termination. The efficiency of termination at TII depended on the growth state of the cells from which the extracts were made, suggesting that some factor or factors control premature termination in c-myc.

Acetates↗

PubMed1992

Monocytes may amplify their recruitment into inflammatory lesions by inducing monocyte chemotactic protein.

By Northern analysis, freshly isolated monocytes contained no detectable mRNA for monocyte chemotactic protein-1 (MCP-1). However, after 4 hours of incubation at 37 degrees C, MCP-1 mRNA was clearly induced in the monocytes and was found to be highly dependent and directly proportional to the monocyte density. The level of MCP-1 mRNA continued to increase, reaching a peak after 22 hours of incubation. After 3 days in culture, MCP-1 mRNA levels had declined substantially and after 8 days were undetectable in the monocytes/macrophages. The amount of MCP-1 protein secreted correlated with the density-dependent increase in MCP-1 message. We hypothesize that the migration of monocytes into inflammatory lesions may be amplified by the density and time-dependent induction of MCP-1.

Base Sequence↗

PubMed1996

Cellular hyperplasia in rats following continuous intravenous infusion of recombinant human epidermal growth factor.

In this study, we determined in vivo morphologic effects of continuous intravenous infusion of recombinant human epidermal growth factor (EGF) in adult Wistar rats. The EGF used consisted of the amino acid residues 1-48 of the human 53-amino-acid EGF molecule, purified from transfected Escherichia coli. Doses of 25, 100, or 250 micrograms/kg body weight were administered using Harvard digital syringe infusion pumps for 4 weeks. At necropsy, the submandibular salivary glands, Harderian glands, liver, kidneys (females only), and ovaries were enlarged and urinary bladders were thickened in 100- and 250-micrograms/kg rats. Numerous tissues of the 100- and 250-micrograms/kg rats contained hyperplastic epithelial cells, and selected organs also had mesenchymal cell proliferation. Epithelial proliferation was most pronounced in the trachea, nasal cavity, nasolacrimal duct, tongue, stomach, small intestine, large intestine, urinary tract, salivary gland ducts, and Harderian gland. Periportal hepatocytes were hypertrophic, correlating with increased liver weight. In addition, mesenchymal cell proliferation was evident in the gastric mucosa lamina propria and in heart valves in 100- and 250-micrograms/kg rats. Increased ovarian weight correlated with increased number and size of corpora lutea and an increased incidence of luteal cysts. Continuous systemic exposure of adult Wistar rats to high doses of EGF resulted in generalized epithelial hyperplasia and tissue-selective mesenchymal proliferation.

Amino Acid Sequence↗

PubMed2006

Prolactin-like protein-f subfamily of placental hormones/cytokines: responsiveness to maternal hypoxia.

The prolactin (PRL) family of hormones/cytokines is involved in the maintenance of pregnancy and adaptations to physiological stressors. In this report, we identify and characterize a new member of the rat PRL family, examine the impact of maternal hypoxia on placental PRL family gene expression, and investigate maternal adaptive responses to hypoxia. Perusal of the PRL gene family locus in the rat genome resulted in the identification of a putative new member of the rat PRL family. The new member is closely related to the previously reported PRL-like protein-F (PLP-F) and has been named PLP-Fbeta and the originally characterized PLP-F, now termed PLP-Falpha. The two proteins exhibit structural similarities but possess distinct cell- and temporal-specific expression profiles. In vivo hypoxia stimulates placental PLP-Falpha and PLP-E mRNA expression in the rat and mouse, respectively. Rcho-1 trophoblast cells can differentiate into trophoblast giant cells, express PLP-Falpha, and exhibit enhanced PLP-Falpha mRNA levels when cultured under low oxygen tension (2%). Exposure to hypobaric hypoxia during latter part of pregnancy did not significantly impact the expression of PLP-Fbeta mRNA. Finally, exposure to hypobaric hypoxia during midpregnancy led to increased maternal red blood cells, hemoglobin concentrations, hematocrit, and increased concentrations of maternal splenic mRNAs for key proteins involved in hemoglobin synthesis, erythroid Krüppel-like factor, erythroid 5-aminolevulinate synthase-2, and beta-major globin. In summary, adaptive responses to maternal hypoxia include activation of placental PLP-Falpha/E gene expression, which may then participate in maternal hematological adjustments required for maintaining maternal and fetal oxygen delivery.

Adaptation, Physiological↗

PubMed1992

Tripeptide RGD-dependent adhesion of articular chondrocytes to synovial fibroblasts.

Cell-cell interactions play an important role in the development of cartilage. Heterologous and homologous cell-cell interactions are critical for chondrogenic differentiation during development. Cell-cell interactions in the formation of fracture callus and cartilage neoplasia also invoke the process of cartilage differentiation. We have investigated cell-cell interactions between articular chondrocytes and synovial fibroblasts and show that there was enhanced binding between these two cell types compared to background binding of the labelled cells to the tissue culture plastic surface. The binding of chondrocytes to fibroblasts was temperature- and calcium-dependent, suggesting ligand-integrin involvement. The peptide, GRGDSP, which competes with the ligand-integrin through the tripeptide RGD (arginine-glycine-aspartic acid), almost completely inhibited chondrocyte attachment to synovial fibroblasts. The control peptide, GRGESP, had no inhibitory effect on binding. Antibodies to fibronectin (Fn) inhibited chondrocyte attachment by about 50%. Monoclonal antibodies to the alpha and beta chains of the fibronectin receptor (FnR) interfered with the attachment of chondrocytes to synovial fibroblasts. A combination of antibodies to Fn and to FnR did not completely abrogate chondrocyte binding, suggesting that other ligand-receptors were involved in the adhesion process. Chondrocytes and fibroblasts were shown to express membrane-associated Fn and FnR, by immunofluorescence. The alpha and beta chains of FnR, migrating at 110 and 140 kDa, respectively, could be immunoprecipitated from [35S]methionine-labelled synovial fibroblasts and chondrocytes. Northern blots showed the presence of mRNA for the alpha and beta chains of fibronectin receptors in fibroblasts and chondrocytes. Changes in cell shape were observed in chondrocytes on attachment to fibroblasts, i.e. the chondrocytes appeared fibroblast-like, suggesting that the chondrocytes had dedifferentiated. These studies suggest that chondrocytes specifically bind to synovial fibroblasts through RGD-dependent receptors. beta 1 Integrins are involved in this adhesion process and these heterlogous cell interactions appear to have a negative influence on chondrogenic differentiation.

Amino Acid Sequence↗

PubMed2003

Activation of antigen-specific CD8 T cells results in minimal killing of bystander bacteria.

Memory CD8 T cells play a critical role in protective immunity against intracellular pathogens. In addition to their ability to specifically recognize and lyse infected targets, activated CD8 T cells secrete cytokines that induce phagocytic cells to engulf and kill bacterial pathogens. In this study, we asked whether activation of Ag-specific CD8 T cells results in nonspecific killing of bystander bacteria during a mixed infection. Mice with epitope-specific memory CD8 T cells were coinfected with two isogenic strains of recombinant Listeria monocytogenes that differ in the cognate epitope. Recall responses by epitope-specific CD8 T cells rapidly inhibited the growth of epitope-bearing bacteria, impeding the course of infection within 6 h after challenge. This rapid inhibition was highly specific and did not affect the growth of coinfecting bacteria without the epitope. CTL recall did not enhance activation of innate immune cells, as evidenced by the absence of inducible NO synthase production in infectious foci. Our observations demonstrate the remarkable specificity of the bactericidal mechanisms of CTL and reveal the possibility for escape mutants to prevail in the hostile environment of a specific immune response. This implication has a bearing on subunit vaccine design strategies and understanding failure of immunization against bacterial infection.

Amino Acid Sequence↗

PubMed1975

Improvement of scintigram reliability by isocount scanning and multilevel analysis.

Given the target source conditions and the detector parameters, the isocount scanning uses optimally the available observation time by maintaining the statistical reliability of each measurement in the composition of a scintigram and by devoting all the observation time to the region of interest through logic circuits which allows quick skipping of the background region. The multilevel analysis is an effective enlargement of the dynamic range of the displayer screen density characteristics that emphasizes small uptake ratios in any density level. It also makes the features extraction very easy by producing an animated sequence of pictures of the observed organ on the screen. The detectability of small uptake ratios significantly improved, specially in regions of low counting rate where the fluctuation of data severely degrades the picture quality in the conventional constant speed scanning method.

Brain Neoplasms↗

PubMed1996

NK 1+ CD4- CD8- alphabeta T cells in the peritoneal cavity: specific T cell receptor-mediated cytotoxicity and selective IFN-gamma production against B cell leukemia and myeloma cells.

NK1+ double negative (DN) alphabeta T cells were present in the peritoneal exudate cells (PEC) of both normal and athymic B6 mice, accounting for as much as 25% of the total T cells, while their numbers were far less in the PEC of BALB/c and (BALB/c x B6)F1 mice. IL-2-dependent clones established from the DN alphabeta T cell population in the PEC of IL-2 receptor alpha-chain transgenic B6 mice exhibited potent cytotoxicity against a series of B cell lineage leukemias and myelomas, such as CD5+BCL1 and MOPC, without affecting NK-susceptible targets. The cytotoxicity of the clones against BCL1 and MOPC was specifically inhibited by anti-CD3, anti-alphabetaTCR, or anti-relevant Vbeta (Vbeta8) Ab, but not by control Abs, indicating that it was mediated by the specific alphabetaTCR/CD3. Other BALB/c-derived target cells expressing both MHC class I and class II were not affected, and neither Ab against them affected the cytotoxicity, strongly suggesting that the cytotoxicity of NK1+ DN alphabeta T cell clones was independent of the particular MHC Ags. The clones produced IFN-gamma, but little IL-2 or IL-4, in response to anti-CD3 stimulation, to the susceptible, but not resistant, targets, and to IL-12. The clones exhibited TCRalpha (Valpha8) distinct from an invariant TCRalpha (Valpha14) reported to dominate in thymic NK1+ alphabeta T cells. The presence of DN alphabeta T cells with similar functional features in the normal PEC was confirmed by the short term stimulation in vitro. The present results along with other recent reports strongly suggested that, like the mainstream alphabeta T cells, the NK1+ DN alphabeta T cell population consisted of functionally heterogeneous subsets.

Amino Acid Sequence↗

PubMed1999

Mycobacterium bovis bacille Calmette-Guérin enhances pathogenicity of simian immunodeficiency virus infection and accelerates progression to AIDS in macaques: a role of persistent T cell activation in AIDS pathogenesis.

It has recently been proposed that Mycobacterium tuberculosis may enhance the pathogenicity of HIV infections and accelerate the course of HIV disease. This hypothesis has been tested in the present study using a simian immunodeficiency virus of macaques (SIVmac)/Mycobacterium bovis bacille Calmette-Guérin (BCG)-coinfected macaque model. Naive and chronically SIVmac-infected monkeys were evaluated. Following BCG inoculation, the SIVmac-infected monkeys exhibited the dominant responses of TCR-beta complementarity-determining region 3-restricted T cell subpopulations. This BCG-driven T cell activation correlated with a marked increase in viral loads in SIVmac-infected monkeys. Moreover, the prolonged T cell activation coincided with the enhanced decline of CD4+ PBL counts and the accelerated progression to clinical AIDS in the coinfected monkeys, suggesting that Mycobacterium-driven T cell activation may be the mechanism underlying the enhanced pathogenicity of AIDS virus infection in the coinfected individuals. Within 2 to 7 mo after BCG coinfection, all chronically SIVmac-infected monkeys died from SIV-induced AIDS including tuberculosis-like disease. Surprisingly, the naive monkeys manifested a T cell activation-related toxic shock syndrome and a profound depletion of CD4+ lymphocytes 2 wk after simultaneous SIVmac/BCG inoculation. These naive animals died 2 mo after SIVmac/BCG inoculation, with the evidence of the persistent SIV p27 antigenemia and SIVmac-induced disease. In contrast, the normal monkeys not infected with SIVmac survived BCG infection; the control SIVmac-infected animals showed a natural course of chronic SIV infection. Thus, results from this SIV/BCG coinfection model strongly support the hypothesis that active coinfection with HIV and Mycobacterium can impact remarkably on the AIDS virus-induced disease.

Acquired Immunodeficiency Syndrome↗

PubMed2006

Non-Helicobacter bacterial flora rarely develops in the gastric mucosal layer of children.

Non-Helicobacter bacteria can be cultured from the gastric mucosa in adults but in children, there are no studies about such microflora. The purpose of this study, therefore, was to clarify whether gastric biota develops in children. In 10 children and 10 adults or elderly (5 H. pylori-infected and 5 uninfected in each group), biopsy specimens of the gastric antrum and corpus and gastric juice were studied for bacterial examinations and the data were compared between both age groups in relation to H. pylori status and luminal pH. Bacterial genera and species were analyzed using both culture and real-time polymerase chain reaction (PCR) with the 52 genus- and species-specific primer sets. Non-Helicobacter bacteria in the mucosa were cultured from all adult patients, whereas microorganisms were cultured in only one child (p < .001). Gastric pH was lower in children (median, 1.4) than in adults (median, 2.6) (p < .005). The grade of endoscopic gastric atrophy was moderate or severe in 8 adults, but absent or mild in all 10 children. Among adults, there was a significant positive correlation between gastric pH and total bacterial counts of both the mucosa and juice. These data indicate that impaired gastric acid secretion associated with long-term H. pylori infection enables non-Helicobacter bacteria to colonize in the human stomach. Such microorganisms rarely colonize in the gastric mucosa in children regardless of H. pylori status.

Adolescent↗

PubMed2005

Silencing the epidermal growth factor receptor gene with RNAi may be developed as a potential therapy for non small cell lung cancer.

Lung cancer has emerged as a leading cause of cancer death in the world. Non-small cell lung cancer (NSCLC) accounts for 75-80% of all lung cancers. Current therapies are ineffective, thus new approaches are needed to improve the therapeutic ratio. Double stranded RNA (dsRNA)-mediated RNA interference (RNAi) has shown promise in gene silencing, the potential of which in developing new methods for the therapy of NSCLC needs to be tested. We report here RNAi induced effective silencing of the epidermal growth factor receptor (EGFR) gene, which is over expressed in NSCLC. NSCLC cell lines A549 and SPC-A1 were transfected with sequence- specific dsRNA as well as various controls. Immune fluorescent labeling and flow cytometry were used to monitor the reduction in the production of EGFR protein. Quantitative reverse-transcriptase PCR was used to detect the level of EGFR mRNA. Cell count, colony assay, scratch assay, MTT assay in vitro and tumor growth assay in athymic nude mice in vivo were used to assess the functional effects of EGFR silencing on tumor cell growth and proliferation. Our data showed transfection of NSCLC cells with dsRNA resulted in sequence specific silencing of EGFR with 71.31% and 71.78 % decreases in EGFR protein production and 37.04% and 54.92% in mRNA transcription in A549 and SPC-A1 cells respectively. The decrease in EGFR protein production caused significant growth inhibition, i.e.: reducing the total cell numbers by 85.0% and 78.3%, and colony forming numbers by 63.3% and 66.8%. These effects greatly retarded the migration of NSCLC cells by more than 80% both at 24 h and at 48 h, and enhanced chemo-sensitivity to cisplatin by four-fold in A549 cells and seven-fold in SPC-A1. Furthermore, dsRNA specific for EGFR inhibited tumor growth in vivo both in size by 75.06% and in weight by 73.08%. Our data demonstrate a new therapeutic effect of sequence specific suppression of EGFR gene expression by RNAi, enabling inhibition of tumor proliferation and growth. However, in vivo use of dsRNA for gene transfer to tumor cells would be limited because dsRNA would be quickly degraded once delivered in vivo. We thus tested a new bovine lentiviral vector and showed lentivector-mediated RNAi effects were efficient and specific. Combining RNAi with this gene delivery system may enable us to develop RNAi for silencing EGFR into an effective therapy for NSCLC.

Journal Article↗

PubMed2003

Aeromicrobium marinum sp. nov., an abundant pelagic bacterium isolated from the German Wadden Sea.

An obligately salt-dependent Gram-positive bacterium, designated strain T2(T), was isolated from surface waters of the German Wadden Sea. The organism exhibited optimum growth at salt concentrations similar to that of sea water. On the basis of phenotypic, chemotaxonomic and phylogenetic differences, it is concluded that strain T2(T) (=DSM 15272(T)=LMG 21768(T)) is the first marine species of the genus Aeromicrobium to be identified, for which the name Aeromicrobium marinum is proposed. It is also the first described marine bacterium within the family Nocardioidaceae. Strain T2(T) is a rod-shaped, aerobic, heterotrophic bacterium containing LL-diaminopimelic acid in the peptidoglycan and MK-9(H(4)) as the major menaquinone. The bacterium is characterized by high proportions of the fatty acids palmitic acid, oleic acid, tuberculostearic acid and hydroxypalmitic acid. DNA-DNA hybridization analysis showed the marine bacterium to display 29.1 % relatedness with Aeromicrobium fastidiosum DSM 10552(T) and 44.4 % relatedness with Aeromicrobium erythreum DSM 8599(T). A. marinum was demonstrated to be an abundant member of the pelagic bacterial community in the German Wadden Sea since it represented about 1 % of the total bacterial population as revealed by dot-blot hybridization and most-probable-number counts.

DNA, Bacterial↗

PubMed1996

Pentoxifylline inhibits acute HIV-1 replication in human T cells by a mechanism not involving inhibition of tumour necrosis factor synthesis or nuclear factor-kappa B activation.

OBJECTIVE AND DESIGN: To study the in vitro activity of pentoxifylline (PTX), which may be of benefit in AIDS, on cell proliferation, tumour necrosis factor (TNF)-alpha, interferon (IFN)-gamma (a type 1 cytokine) and interleukin (IL)-10 (a type 2 cytokine) production, viral replication and CD4+ depletion in acutely HIV-1-infected human T cells. METHODS: T cells were stimulated with anti-CD3 antibody or phytohaemagglutinin (PHA) and infected with HIV-1 in presence or absence of PTX. Cell proliferation, CD4+ cell number, nuclear factor (NF)-kappa B activation, p24 antigen release or lymphokine content of the supernatants were evaluated by [3H]-thymidine incorporation, cytofluorimetry, electrophoretic mobility shift assays and specific enzyme-linked immunosorbent assay, respectively. RESULTS: In HIV-1-infected T cells, PTX inhibited cell proliferation and p24 release and prevented CD4+ depletion associated with HIV replication. Moreover, PTX reduced TNF-alpha, IFN-gamma and IL-10 production and NF-kappa B activation. PTX inhibited with similar potency IFN-gamma, TNF-alpha and cell proliferation. However, the inhibition of p24 release and specially of IL-10 production required significantly lower doses of PTX. Exogenous addition of IL-2 or TNF-alpha in presence of PTX restore T-cell proliferation and NF-kappa B activation respectively, but did not affect p24 inhibition. CONCLUSIONS: Our data suggest that the inhibitory effect of PTX on HIV replication cannot be satisfactorily explained by the inhibition of NF-kappa B or TNF-alpha. Moreover, PTX cannot be primarily considered as a TNF-alpha inhibitor and has several immunomodulatory and antiviral properties which could be of benefit against HIV-1 at various levels.

Antiviral Agents↗

PubMed1995

Nucleated erythrocytes in maternal blood: quantity and quality of fetal cells in enriched populations.

The discovery of nucleated erythrocytes in maternal circulation provides a potential source for non-invasive prenatal diagnosis. We have evaluated the use of a three-stage procedure to determine the number of cells that are of fetal rather than maternal origin. First, monoclonal antibodies specific for CD45 and CD14 were used in conjunction with a magnetic (MACS) column to deplete unwanted leukocytes from maternal blood. This was followed by a positive MACS enrichment for nucleated erythrocytes, using an anti-CD71 (transferrin receptor) monoclonal antibody. To discriminate between fetal nucleated erythrocytes and those of maternal origin, enriched fractions were simultaneously stained with an anti-fetal haemoglobin (HbF) antibody and hybridized with probes specific for X and Y chromosomes. Samples were then subjected to blind analysis along with negative control samples from non-pregnant volunteers. Using this dual analysis, we were able to determine that less than one nucleated erythrocyte per ml of maternal blood was of fetal origin. Small numbers of these fetal cells were found in 87.5% of pregnancies, ranging from 6 to 35 weeks gestational age. Comparison of HbF and X/Y probe data also suggests that the fetal cells are less suitable for fluorescence in-situ hybridization (FISH) analysis than similar preparations from other sources.

Antibodies, Monoclonal↗

PubMed1993

Serum deprivation response gene is induced by serum starvation but not by contact inhibition.

The relationship between quiescence induced by serum deprivation and that induced by high cell density (contact inhibition) is still unclear. Here, we describe a gene, named sdr (serum deprivation response), whose mRNA level is highly induced in serum starved but not in density dependent growth arrested NIH3T3 cells. sdr induction seems to be coupled to growth arrest since it does not become apparent when transformed NIH3T3 cells are cultured in low serum. The expression of sdr is down-regulated within 6 h after the addition of serum or epidermal growth factor to serum starved cells. Although a transient reinduction is noticed at later times after fetal calf serum stimulation, this is not the case for epidermal growth factor, for which the sdr mRNA level remains down-regulated. The data presented here point to the possibility that the density dependent growth arrest state cannot be simply explained by local growth factor depletion, as occurs in overcrowded cultures, but it could be the consequence of a more complex pathway mediated by cellular interactions.

3T3 Cells↗

PubMed2003

Enumeration and characterization of acidophilic microorganisms isolated from a pilot plant stirred-tank bioleaching operation.

Microorganisms were enumerated and isolated on selective solid media from a pilot-scale stirred-tank bioleaching operation in which a polymetallic sulfide concentrate was subjected to biologically accelerated oxidation at 45 degrees C. Four distinct prokaryotes were isolated: three bacteria (an Acidithiobacillus caldus-like organism, a thermophilic Leptospirillum sp., and a Sulfobacillus sp.) and one archaeon (a Ferroplasma-like isolate). The relative numbers of these prokaryotes changed in the three reactors sampled, and the Ferroplasma isolate became increasingly dominant as mineral oxidation progressed, eventually accounting for >99% of plate isolates in the third of three in-line reactors. The identities of the isolates were confirmed by analyses of their 16S rRNA genes, and some key physiological traits (e.g., oxidation of iron and/or sulfur and autotrophy or heterotrophy) were examined. More detailed studies were carried out with the Leptospirillum and Ferroplasma isolates. The data presented here represent the first quantitative study of the microorganisms in a metal leaching situation and confirm that mixed cultures of iron- and sulfur-oxidizing prokaryotic acidophiles catalyze the accelerated dissolution of sulfidic minerals in industrial tank bioleaching operations. The results show that indigenous acidophilic microbial populations change as mineral dissolution becomes more extensive.

Bacteria↗