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Biomedical subjects

A Camilli

Publications and source records attributed to A Camilli.

At least 37 records · Page 2Linked to original sources
PubMed1997

[Hormone replacement therapy (HRT). The effects at 12 months on the risk factors for menopausal osteoporosis].

Cessation of ovarian activity is accompanied by a more or less marked and more or less accelerated reduction in bone mass; the degree and speed of the process--which occurs in all women--depends on individual (genetic factors influencing peak bone mass, duration of child-bearing period), iatrogenic (treatment with corticosteroids or thyroid hormones) or accidental factors (post-traumatic immobilization). Whatever the factor that triggers off the process, the end result is the destruction of bone tissue. This process may be documented by hematochemical (Nordin's test) and instrumental parameters (MOC and similar techniques). Oestroprogestin (and to a lesser extent calcitonin) hormone replacement therapy has been demonstrated to be highly efficacious in countering this involutive process. The authors report data obtained following the evaluation of 35 women in menopause undergoing. Nordin's test and measurement of the plasma level of estradiol using RIA, before and after twelve months after the start of osteoprotective treatment. Of the 35 patients 9 received only progestin, 22 an oestroprogestin combination (of these 18 patients received estrogen transdermally and 4 orally), and 4 calcitonin administered parenterally. A statistically significant positive correlation with Nordin's test was only found in the group receiving oestrogen therapy. In conclusion, it may be affirmed that in the absence of contraindications oestrogens represent the elective form of treatment for menopausal osteoporosis. Acceptable results have been reported in the literature also using calcitonin, but this treatment could not be evaluated in this study owing to the reduced number of the sample treated.

Calcitonin↗
PubMed1997

[The effects of hormone replacement therapy (HRT) on lipid metabolism in the menopause].

INTRODUCTION AND AIMS: During menopause the number of cardiovascular attacks increases parallel to the elimination of estradiol production. The administration of the latter reverses this tendency owing to a compound mechanism (improved HDL) cholesterol/total cholesterol ratio, diminished vasal resistance). MATERIALS AND METHODS: The authors present a study performed in 34 patients in menopause receiving oestroprogestinic replacement therapy using an oral or transdermal route. Metabolic status (serum concentrations of total cholesterol (TC) and HDL cholesterol (C.HDL) was evaluated in all patients before treatment and after 12 months. RESULTS: A statistically significant positive correlation (p < 0.0001) was found between the use of oestrogen and serum levels of HDL. This correlation appeared to be more evident in patients using transdermal treatment compared to the oral form. CONCLUSIONS: The authors conclude that in the absence of contraindications, hormone replacement therapy in menopause exercises a beneficial effect on the lipid status, contributing to diminishing the risk of cardiovascular attacks. The possibility of an increased incidence of breast cancer is now being evaluated, whereas effective protection of the endometrium against the risk of hyperplasia and cancer was shown using the doses of progestin used in this study, which coincide with those currently prescribed in the literature.

Estradiol↗
PubMed1995

Use of recombinase gene fusions to identify Vibrio cholerae genes induced during infection.

A complete understanding of host-parasite interactions must necessarily include the identification and characterization of gene products expressed by both parties during the infectious process. We have developed a new screen to identify bacterial genes that are transcriptionally induced during infection of a host animal. The method is based on pre-selection of strains carrying tnpR operon fusions (encoding resolvase, a site-specific DNA recombinase) which are not expressed in vitro, followed by screening for a subset of these strains that subsequently express resolvase within the host environment. The latter subset was recognized as recombinants that had deleted a resolvase-specific reporter construct. Thirteen transcription units of Vibrio cholerae were identified that were induced during infection in an infant mouse model of cholera. Five of these were predicted to encode polypeptides with diverse functions in metabolism, biosynthesis and motility; one encoded a secreted lipase; two appear to be antisense to genes involved in motility; and five are predicted to encode polypeptides of unknown function. Three of the transcripts were shown to be required for full virulence in infant mice, as assessed by competition experiments.

Amino Acid Sequence↗
PubMed1994

Use of genetic recombination as a reporter of gene expression.

An understanding of the patterns of gene expression in response to specific environmental signals can yield insight into a variety of complex biological systems such as microbial-host interactions, developmental cycles, cellular differentiation, ontogeny, etc. To extend the utility of the reporter gene fusion approach to such studies, we have constructed a gene expression reporter cassette that permits the generation of transcriptional fusions to tnpR encoding resolvase, a site-specific recombinase of the transposable element gamma delta. Induction of the transcriptional fusions results in production of resolvase, which in turn, catalyzes excision of a linked tetracycline-resistance reporter gene flanked by direct repeats of res, the DNA sequences at which resolvase functions. The loss of tetracycline resistance in descendant bacteria serves as a permanent and heritable marker of prior gene expression. This gene fusion approach will allow us to assay the induction of gene expression in as few as one cell. Additionally, gene expression can be assayed at a later time and/or different place from the inducing environment facilitating the study of gene expression in complex environments such as animal tissues.

Alkaline Phosphatase↗
PubMed1994

Overproduction of a dextranase inhibitor by Streptococcus sobrinus mutants.

An inhibitor of Streptococcus sobrinus endodextranase was detected in the extracellular fractions of UAB66 mutants identified following ethyl methanesulfonate mutagenesis as either devoid of dextranase activity (Dex-) or overproducing water-soluble glucan. The two groups of mutants had the same phenotype and displayed no dextranase activity in assays of extracellular fractions (H. Murchison, S. Larrimore, and R. Curtiss III, Infect. Immun. 34:1044-1055, 1981) and had been shown to be defective in adherence (Adh-) and capable of inhibiting adherence of wild-type strains during cocultivation in vitro (H. Murchison, S. Larrimore, and R. Curtiss III, Infect. Immun. 50:826-832, 1985) and in vivo in gnotobiotic rats (K. Takada, T. Shiota, R. Curtiss III, and S. M. Michalek, Infect. Immun. 50:833-843, 1985). By analysis of proteins in Western blots (immunoblots) and following blue dextran-sodium dodecyl sulfate-polyacrylamide gel electrophoresis (BD-SDS-PAGE), it was demonstrated that these Dex- mutants did synthesize enzymatically active dextranase. From the results of mixing experiments, it was determined that these Dex- Adh- mutants produced enhanced amounts of a cell surface-localized or a cell-associated dextranase inhibitor (Dei). Dei was heat stable but trypsin sensitive. By adding excess dextranase following BD-SDS-PAGE, Dei was detected as blue bands with apparent molecular masses of 43, 40, 37, 27, and 23 kDa. Dei competitively inhibits dextranase activity and is synthesized by wild-type S. sobrinus strains, with the amount varying depending upon growth medium and stage in the growth cycle. R. M. Hamelik and M. M. McCabe (Biochem. Biophys. Res. Commun. 106:875-880, 1982) previously described a Dei in a wild-type S. sobrinus strain.

Bacterial Adhesion↗
PubMed1994

Cloning and DNA sequencing of the dextranase inhibitor gene (dei) from Streptococcus sobrinus.

Some dextranase-deficient (Dex-) mutants of Streptococcus sobrinus UAB66 (serotype g) synthesize a substance which inhibits dextranase activity (S.-Y. Wanda, A. Camilli, H. M. Murchison, and R. Curtiss III, J. Bacteriol. 176:7206-7212, 1994). This substance produced by the Dex- mutant UAB108 was designated dextranase inhibitor (Dei) and identified as a protein. The Dei gene (dei) from UAB108 has been cloned into pACYC184 to yield pYA2651, which was then used to generate several subclones (pYA2653 to pYA2657). The DNA sequence of dei was determined by using Tn5seq1 transposon mutagenesis of pYA2653. The open reading frame of dei is 990 bp long. It encodes a signal peptide of 38 amino acids and a mature Dei protein of 292 amino acids with a molecular weight of 31,372. The deduced amino acid sequence of Dei shows various degrees of similarity with glucosyltransferases and glucan-binding protein and contains A and C repeating units probably involved in glucan binding. Southern hybridization results showed that the dei probe from UAB108 hybridized to the same-size fragment in S. sobrinus (serotype d and g) DNA, to a different-size fragment in S. downei (serotype h) and S. cricetus (serotype a), and not at all to DNAs from other mutans group of streptococci.

Amino Acid Sequence↗
PubMed1993

Expression and phosphorylation of the Listeria monocytogenes ActA protein in mammalian cells.

Movement of Listeria monocytogenes within infected eukaryotic cells provides a simple model system to study the mechanism of actin-based motility in nonmuscle cells. The actA gene of L. monocytogenes is required to induce the polymerization of host actin filaments [Kocks, C., Gouin, E., Tabouret, M., Berche, P., Ohayon, H. & Cossart, P. (1990) Cell 68, 521-531; Domann, E., Wehland, J., Rohde, M., Pistor, S., Hartl, M., Goebel, W., Leimeister-Wachter, M., Wuenscher, M. & Chakraborty, T. (1992) EMBO J. 11, 1981-1990]. In this study, an in-frame deletion mutation within the actA gene was constructed and introduced into the L. monocytogenes chromosome by allelic exchange. This mutation resulted in a decrease (3 orders of magnitude) in virulence for mice. In tissue culture cells, the actA mutant was absolutely defective for the nucleation of actin filaments and consequently was impaired in cell-to-cell spread. Antiserum raised to a synthetic peptide encompassing the proline-rich repeat (DFPPPPTDEEL) of ActA was used to characterize the expression of the ActA protein. The ActA protein derived from extracellular bacteria migrated as a 97-kDa polypeptide upon SDS/PAGE, whereas the protein from infected cells migrated as three distinct polypeptides, one that comigrated with the 97-kDa extracellular form and two slightly larger species. Treatment of infected cells with okadaic acid resulted in decreased amounts of all forms of ActA and the appearance of a larger species of ActA. Phosphatase treatment of ActA immunoprecipitated from intracellular bacteria resulted in conversion of the larger two species to the 97-kDa form. Labeling of infected cells with 32Pi followed by immunoprecipitation showed that the largest molecular form of ActA was phosphorylated. Taken together, these data indicate that ActA is phosphorylated during intracellular growth. The significance of the intracellular modification of ActA is not known, but we speculate that it may modulate the intracellular activity of ActA.

Amino Acid Sequence↗
PubMed1993

Dual roles of plcA in Listeria monocytogenes pathogenesis.

The plcA gene of Listeria monocytogenes encodes a secreted phosphatidylinositol-specific phospholipase C (Pl-PLC). Recent studies have established that transposon mutations within plcA result in avirulence for mice and pleiotropic effects when examined in tissue-culture models of infection. Genetic analysis reveals that many of the effects of the transposon insertions are due to loss of readthrough transcription from plcA into the downstream gene prfA, which encodes an essential transcription factor of numerous L. monocytogenes virulence genes. Construction of an in-frame deletion within plcA had no effect on expression of prfA thus allowing direct assignment of a role of the Pl-PLC in pathogenesis. Pl-PLC was shown to play a significant role in mediating escape of L. monocytogenes from phagosomes of primary murine macrophages. Interestingly, this defect manifested itself in vivo in the liver but not in the spleen of infected mice.

Animals↗
PubMed1991

Listeria monocytogenes mutants lacking phosphatidylinositol-specific phospholipase C are avirulent.

A number of bacterial species secrete phosphatidylinositol-specific phospholipase C (PI-PLC). In this report, we show that the facultative intracellular bacterial pathogen, Listeria monocytogenes, contains a gene, plcA, predicting a polypeptide with 31% amino acid identity to a Bacillus thuringiensis PI-PLC. Accordingly, L. monocytogenes secretes PI-PLC activity, while a mutant with a transposon insertion in plcA lacks detectable PI-PLC activity. In addition, expression of plcA in B. subtilis resulted in secretion of PI-PLC activity. The L. monocytogenes PI-PLC-defective mutant was three logs less virulent for mice and failed to grow in host tissues. The mutant was also defective for in vitro growth in mouse peritoneal macrophages. These results strongly suggest that PI-PLC is an essential determinant of L. monocytogenes pathogenesis. Whether the PI-PLC acts on a bacterial or host substrate remains to be determined.

Animals↗
PubMed1990

Isolation of Listeria monocytogenes small-plaque mutants defective for intracellular growth and cell-to-cell spread.

To dissect the regulatory and structural requirements for Listeria monocytogenes intracellular growth and cell-to-cell spread, we designed a protocol based on transposon mutagenesis and the isolation of mutants which form small plaques in monolayers of mouse L2 cell fibroblasts. Two different transposable elements were used to generate libraries of insertion mutants: Tn916 and a derivative of Tn917-lac, Tn917-LTV3. Ten classes of mutants were isolated and evaluated for growth and cell-to-cell spread in J774 mouse macrophagelike cells, Henle 407 human epithelial cells, and mouse bone marrow-derived macrophages. Mutants were also evaluated for secretion of hemolysin and phospholipase (assayed by egg yolk opacity) and association with F-actin in the cytoplasm of cells, using NBD-phallacidin staining. The ten classes of mutants included (i) mutants showing abortive intracellular and extracellular growth; (ii) mutants showing abortive intracellular growth; (iii) rough mutants; (iv) mutants showing greatly reduced hemolysin and phospholipase secretion but showing normal growth in cells and little or no association with F-actin; (v) mutants with mutations mapping to an open reading frame (ORF) adjacent to hlyA and referred to as ORF U, lacking phospholipase activity, and with 50% normal hemolysin activity; (vi) mutants with reduced secretion of both hemolysin and phospholipase; (vii) nonhemolytic mutants with mutations mapping to the structural gene, hlyA; (viii) mutants with 25% normal hemolysin secretion and absolutely no association with F-actin; (ix) mutants with mutations mapping to ORF U, lacking phospholipase activity, and with normal hemolysin activity; and (x) mutants showing a mixed-plaque morphology but normal for all other parameters.

Actins↗
PubMed1990

Insertional mutagenesis of Listeria monocytogenes with a novel Tn917 derivative that allows direct cloning of DNA flanking transposon insertions.

To carry out efficient insertional mutagenesis in Listeria monocytogenes and to facilitate the characterization of disrupted genes, two novel derivatives of Tn917 were constructed, Tn917-LTV1 and Tn917-LTV3. The derivatives (i) transpose at a significantly elevated frequency, (ii) generate transcriptional lacZ fusions when inserted into a chromosomal gene in the appropriate orientation, and (iii) allow the rapid cloning in Escherichia coli of chromosomal DNA flanking transposon insertions. The rapid cloning of DNA flanking insertions is possible because the transposon derivatives carry ColE1 replication functions, a cluster of polylinker cloning sites, and antibiotic resistance genes selectable in E. coli (bla in the case of Tn917-LTV1; neo and ble in the case of Tn917-LTV3). The enhanced transposition frequency of Tn917-LTV1 and Tn917-LTV3 (about 100-fold in Bacillus subtilis) is believed to be due to the fortuitous placement of vector-derived promoters upstream from the Tn917 transposase gene. In L. monocytogenes, Tn917-LTV3 transposed at a frequency of 8 x 10(-4) when introduced on a pE194Ts-derived vector and generated at least eight different auxotrophic mutations. Two nonhemolytic insertion mutants of L. monocytogenes were isolated, and DNA flanking the transposon insertions was cloned directly into E. coli, making use of the ColE1 rep functions and neo gene carried by Tn917-LTV3. Both insertions were shown to be within hlyA, the L. monocytogenes hemolysin structural gene. Although Tn917-LTV1 and Tn917-LTV3 were constructed specifically for genetic analysis of L. monocytogenes, their enhanced transposition frequency and convenience for cloning of DNA adjacent to sites of insertions make them the transposon derivatives of choice for insertional mutagenesis in any gram-positive bacteria that support replication of pE194Ts.

Bacillus subtilis↗
PubMed1989

Intracellular methicillin selection of Listeria monocytogenes mutants unable to replicate in a macrophage cell line.

To dissect the determinants of Listeria monocytogenes that are required for pathogenicity, we designed an intracellular selection protocol based on penicillin selection to isolate mutants defective for intracellular growth. Eight independent mutants obtained by insertion of Tn916 were isolated that were resistant to methicillin treatment following internalization by the J774 macrophage-like cell line. Seven mutants were absolutely defective for intracellular growth, whereas one showed abortive intracellular growth. The majority of the mutants were nonhemolytic and lacked a secreted 58-kDa polypeptide thought to be the L. monocytogenes hemolysin, listeriolysin O. Southern blot analysis indicated that one mutant contained a Tn916 insertion in hlyA, the listeriolysin O structural gene, which resulted in a truncated listeriolysin O polypeptide, whereas another mutant contained an insertion immediately upstream of hlyA, which resulted in reduced expression of listeriolysin O. The other mutants contained Tn916 insertions in genes other than hlyA, although all but one were nonhemolytic. Revertants isolated by their ability to grow within tissue culture cells regained hemolytic activity. These data show that intracellular methicillin selection facilitates isolation of mutations in genes required for intracellular growth and strengthens the premise that listeriolysin O is essential for intracellular growth.

Animals↗
PubMed1989

On the temporal relationships between lung function and somatic growth.

A sample population of healthy nonsmoking male (n = 416) and female (n = 608) subjects derived from all non-Hispanic white subjects 5 to 60 yr of age enrolled in the Tucson Epidemiological Study of Airway Obstructive Disease was studied to evaluate the temporal relationships between various pulmonary function measures and somatic growth. Pulmonary function measures derived from the maximal expiratory flow volume (MEFV) curve included FVC, FEV1, maximal expiratory flow after 50% of vital capacity had been expired (Vmax50), and maximal midexpiratory flow (FEF25-75). The lung function and somatic growth longitudinal data were characterized by a nonparametric polynomial smoothing spline model. This procedure yields an optimal fitted curve through the data, an estimate of the growth velocity curve, and 95% confidence bands. Temporal relationships between the fitted growth curves were determined examining the ages at which the growth velocity peaks (GVP) occur. In addition, the age of growth cessation was estimated using lower limits of the 95% confidence bands of each growth velocity curve. The results indicate that the point estimates of the somatic GVPs precede all peak lung function measurements derived from the MEFV curve in both males and females. The only statistically significant difference between when the GVPs for somatic growth and functional lung growth occurred was for FEV1 and FEF25-75 in males, though similar trends were apparent in all other variables, suggesting that the timing of maximal body growth velocity precedes that of maximal lung functional growth and that the early growth spurt in females includes the lungs.(ABSTRACT TRUNCATED AT 250 WORDS)

Adolescent↗
PubMed1987

Respiratory effects of non-tobacco cigarettes.

Data from the Tucson epidemiological study of airways obstructive disease on smoking of non-tobacco cigarettes such as marijuana were analysed to determine the effect of such smoking on respiratory symptoms and pulmonary function. Among adults aged under 40, 14% had smoked non-tobacco cigarettes at some time and 9% were current users. The prevalence of respiratory symptoms was increased in smokers of non-tobacco cigarettes. After tobacco smoking had been controlled for men who smoked non-tobacco cigarettes showed significant decreases in expiratory flow rates at low lung volumes and in the ratio of the forced expiratory volume in one second to the vital capacity. This effect on pulmonary function in male non-tobacco cigarette smokers was greater than the effect of tobacco cigarette smoking. These data suggest that non-tobacco cigarette smoking may be an important risk factor in young adults with respiratory symptoms or evidence of airways obstruction.

Adolescent↗