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Biomedical subjects

M Fons

Publications and source records attributed to M Fons.

At least 19 recordsLinked to original sources
PubMed1994

Study of gene transfer in vitro and in the digestive tract of gnotobiotic mice from Lactococcus lactis strains to various strains belonging to human intestinal flora.

The use of genetically modified organisms (GMO) in dairy products requires evaluation of the DNA transfer capacity from such organisms among the human intestinal microflora. Thus, both in vitro and in vivo [in the digestive tract (DT) of mice] transfer from Lactococcus lactis donor strains of the conjugative plasmid pIL205 (CmR) and the non-conjugative plasmid pIL253 (EmR) to: (1) recipient strains isolated from human faecal flora Bacteroides sp., Bifidobacterium sp., Peptostreptococcus sp. (strictly anaerobic bacterial strains) and Enterococcus faecalis, (2) a whole human faecal flora, was studied. In both cases, no gene transfer was observed to strictly anaerobic bacterial strains. DNA transfer was only observed to the E. faecalis strain: in vivo CmR E. faecalis transconjugants were isolated from sequentially multi-associated mice and when the recipient strains associated with the mice, they were a defined mixture of Bacteroides sp., Bifidobacterium sp., Peptostreptococcus sp. and E. faecalis strains. When mice were associated with the whole human faecal flora, the plasmid pIL205 was transferred into some facultative anaerobic streptococci. It was also shown that DNA transfer occurred even when the lactococcal donor strain was transient in the DT of the gnotobiotic host animals.

Animals↗
PubMed1991

Possible involvement of a L-delta 1-pyrroline-5-carboxylate (P5C) reductase in the synthesis of proline in Desulfovibrio desulfuricans Norway.

A L-delta 1-pyrroline-5-carboxylate reductase activity has been detected in crude extracts of Desulfovibrio desulfuricans Norway. This P5C reductase activity is also found when a 2.5 kb D. desulfuricans DNA fragment is introduced into an Escherichia coli proC mutant. Although it restores growth of the proC mutant, the ProDd enzyme might be detrimental to the E. coli host since the plasmid carrying the cognate proDd gene is segregated at high rate by the cells but is stabilized by small deletions which lead to a loss of the P5C reductase activity.

DNA, Bacterial↗
PubMed1991

Amiloride inhibition of human cytomegalovirus replication.

Amiloride, an inhibitor of Na+/H+ exchange, interfered with cytomegalovirus (CMV) DNA synthesis, blocked the formation of nuclear inclusions, and reduced CMV infectious yields. The reduction of CMV infectious yields was concentration dependent with an ED90 of 46 microM. Amiloride at a concentration of 150 microM reduced CMV yields by about 100-fold. Reduction of infectious yields appeared to be related to interference with the formation of nuclear inclusions and to inhibition of CMV DNA synthesis. Nuclear inclusions were much reduced in size and demonstrated poorly defined cellulae in the amiloride-treated cells. CMV DNA synthesis was inhibited by approximately 70% when cells were treated with 150 microM amiloride. The reduction in CMV yields could not be related to the reported inhibitory effect of amiloride on protein synthesis. In amiloride (150 microM)-treated, CMV-infected cells, late, yet not immediate-early or early, protein synthesis was markedly decreased relative to untreated, CMV-infected cells. Accordingly, CMV DNA synthesis and the replication of CMV may be related to Na+ entry through an amiloride-sensitive pathway.

Amiloride↗
PubMed1990

Characterization of transposon insertion out- mutants of Erwinia carotovora subsp. carotovora defective in enzyme export and of a DNA segment that complements out mutations in E. carotovora subsp. carotovora, E. carotovora subsp. atroseptica, and Erwinia chrysanthemi.

Soft-rotting Erwinia spp. export degradative enzymes to the cell exterior (Out+), a process contributing to their ability to macerate plant tissues. Transposon (Tn5, Tn10, Tn10-lacZ) insertion Out- mutants were obtained in Erwinia carotovora subsp. carotovora 71 by using plasmid and bacteriophage lambda delivery systems. In these mutants, pectate lyases, polygalacturonase, and cellulase, which are normally excreted into the growth medium, accumulated in the periplasm. However, localization of the extracellular protease was not affected. The Out- mutants were impaired in their ability to macerate potato tuber tissue. Out+ clones were identified in a cosmid library of E. carotovora subsp. carotovora 71 by their ability to complement mutants. Localization of cyclic phosphodiesterase in the periplasm indicated that the Out+ plasmids did not cause lysis or a nonspecific protein release. The Out+ derivatives of the E. carotovora subsp. carotovora 71 mutants regained the ability to macerate potato tuber tissue. Our data indicate that a cluster of several genes is required for the Out+ phenotype. While one plasmid, pAKC260, restored the Out+ phenotype in each of the 31 mutants of E. carotovora subsp. carotovora, E. carotovora subsp. atroseptica, and Erwinia chrysanthemi, it failed to render Escherichia coli export proficient. Homologs of E. carotovora subsp. carotovora 71 out DNA were detected by Southern hybridizations in subspecies of E. carotovora under high-stringency conditions. In contrast, E. chrysanthemi sequences bearing homology to the E. carotovora subsp. carotovora 71 out DNA were detectable only under low-stringency hybridization. Thus, although the out genes are functional in these two soft-rotting bacterial groups, the genes appear to have diverged.

Biological Transport↗
PubMed1990

Cell activation signals and the pathogenesis of human cytomegalovirus.

Cytomegalovirus (CMV) infection induces a series of cellular responses that resemble those observed in cells activated by growth factors or hormones including: hydrolysis of phosphatidylinositol-4,5-bisphosphate; Ca2+ influx and an increase in the cytosolic free [Ca2+]; an increase in Na+ entry; and, increases in cellular levels of cyclic AMP and cyclic GMP. The time courses for some of these responses appear to be markedly protracted relative to those observed for growth factors. The prolonged physiologic responses in CMV-infected cells appear to be related to modifications in the intracellular environment that are associated with the development of cytomegaly and with the phasing of CMV-directed macromolecular synthesis. For example, as the infected cell enlarges, the rate of CMV DNA synthesis increases by about 4-fold, late nuclear inclusions develop and progeny viruses are formed. When the CMV-induced activation signals are inhibited or their physiologic responses are blocked, then the yields of infectious CMV are substantially reduced. Furthermore, perturbation of the cell cycle resulting from induction of the cell activation process by CMV may be causally related to the induction of cellular damage by CMV, even in the absence of productive infection. Accordingly, the CMV-induced pathophysiologic cell activation responses represent potential targets for novel antiviral therapy.

Amiloride↗
PubMed1989

Interleukin 2 induced non MHC-restricted killing of herpes simplex type-1 (HSV-1) infected allogeneic and autologous lymphoblasts.

Peripheral blood lymphocytes (PBL) from herpes simplex type-1 (HSV-1) seropositive (sero+) and seronegative (sero-) individuals were assessed before and after in vitro culture with IL-2 for their ability to kill different tumor cells and to kill HSV-1 infected PHA lymphoblasts using a 4 hr 51Cr release assay. Fresh PBL from either sero+ or sero- individuals had little cytotoxic activity against HSV-1 infected autologous PHA lymphoblasts or against different NK-insensitive tumor targets. When PBL from either sero+ or sero- individuals were incubated 3-4 days in vitro with IL-2, significant levels of cytotoxic activity was observed against both allogeneic and autologous HSV-1 infected targets and against different NK-insensitive tumor cells. When comparing PBL from sero+ versus sero- individuals for IL-2 induced cytotoxic activity against HSV-1 infected autologous targets, PBL from sero+ individuals always had significantly higher levels of cytotoxic activity. Allogeneic HSV-1 infected targets were also killed by IL-2 activated PBL obtained from either sero+ or sero- individuals, but IL-2 activated PBL from sero+ individuals showed higher cytotoxic activity on HSV-1 infected autologous targets when compared to HSV-1 infected allogeneic targets. When IL-2 induced cytotoxic effector cells were treated with HSV-1 for 1 hr and recultured with IL-2, significant inhibition of cytotoxic activity against all target cells was observed. Greater than 50% inhibition of cytotoxic activity was observed when IL-2 induced cytotoxic effector cells were treated with HSV-1 and cultured for 6 hr.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1988

Aerobic expression of the nar operon of Escherichia coli in a fnr mutant.

Mutations allowing aerobic expression of the anaerobically controlled nar operon have been located in the autoregulated fnr gene. Cloning and sequencing of the mutant fnrd20 allele, and fnr mRNA quantitation by dot blot assay, revealed that the mutation was the result of an IS5 insertion into the control region of fnr that enhanced transcription of the fnr gene at least ten-fold. Examination of the regulatory region of the negatively autoregulated fnr gene indicated that it shared homologous sequences with the positively Fnr-controlled frd and nar operons. The increase in fnr transcription in the fnrd20 mutated allele could be partly the result of loss of autoregulation, since the IS5 separated the Fnr target site from the '-35' region of the promoter.

Alleles↗
PubMed1987

Cloning in Escherichia coli of genes involved in the synthesis of proline and leucine in Desulfovibrio desulfuricans Norway.

A library of Desulfovibrio desulfuricans Norway genomic DNA was constructed in Escherichia coli with pBR322 as vector and plasmids able to complement the proA and leuB mutations of the host were screened. It was observed that all the plasmids studied were highly unstable, the insert DNA being rapidly lost under non-selective growth conditions. A 2.75 kb DNA fragment of D. desulfuricans Norway was found to complement E. coli ProA, ProB and ProC deficiencies. From the results of restriction analysis and Southern hybridizations, it is proposed that the genes involved in proline and leucine biosynthesis are clustered on the chromosome of D. desulfuricans Norway.

Cloning, Molecular↗
PubMed1986

Defective NK cell activity following thermal injury.

Peripheral blood mononuclear lymphocytes (PBL) from thermal injury patients were examined for their ability to mediate natural killer (NK) cell activity against K562 tumour cells and against herpes simplex virus type 1 (HSV-1) infected Raji tumour cells. Using fluorescein isothiocyanate-conjugated monoclonal antibodies, the number of T3, T4, T8, Leu11, and Leu7 positive cells in PBL obtained from patients and normal controls was determined. Thermal injury patients had decreased levels of T3+ cells and a T4:T8 ratio which was significantly lower than that found in normal control individuals. Although patients had normal percentages of Leu7+ and Leu11+ cells, they had depressed NK cell activity against both K562 tumour cells and HSV-1 infected Raji cells. NK cell activity against K562 tumour cells was severely depressed during the first 20 days after injury. This defective NK cell activity did not appear to be due to a defect in PBL binding to the K562 tumour cells. In patients, the level of NK cell activity against HSV-1 infected cells did not correlate with the level of NK cell activity against K562 tumour cells. This finding further supports previous reports showing that NK cells which kill K562 tumour cells are different from the NK cell population which kills HSV-1 infected cells. Pretreatment of PBL obtained from patients with IL-2 or IFN-alpha, in some cases greatly enhanced NK cell killing of K562 tumour cells. However, IL-2 or IFN-alpha did not enhance NK cell activity in patients who had severely depressed levels of NK cell activity. Interestingly, in some patients, differential responsiveness to IL-2 and IFN-alpha was observed. In some patients, NK cell activity was enhanced by IL-2 but not by IFN-alpha. These results, while only suggestive, may indicate that different populations of NK cells respond preferentially to IL-2 and that IFN-alpha and/or IL-2 enhance NK cell activity in PBL obtained from some, but not all, thermal injury patients. Finally, this study clearly shows that thermal injury patients have defective NK cell activity not only against K562 tumour cells but also against virus-infected cells.

Adult↗
PubMed1981

A (carbonmonoxy)heme complex with a weak proximal bond. Molecular stereochemistry of carbonyl(deuteroporphinato)(tetrahydrofuran)iron(II).

The synthesis and structural characterization of a six-coordinate (carbonmonoxy)(porphyrin)iron(II), carbonyl(deuteroporphinato)(tetrahydrofuran)iron(II), is described. The choice of tetrahydrofuran as the sixth ligand represents one of the weakest field and weakest binding ligands possible and leads to stronger than usual bonding of CO to the heme. The Fe--C(CO) bond distance is 1.706 A, unusually short. The Fe--O(THF) bond distance in the low-spin complex is 2.127 A. The Fe--C--O and O--Fe--C groups are essentially linear with angles of 178.3 degrees and 177.4 degrees, respectively. The iron atom is displaced out of plane 0.10 A toward the tightly bound CO. The results suggest that the different CO stretching frequencies observed in myoglobin might result from modulating the bonding of the histidine trans to CO. Crystal data are as follows: a = 11.524 (3) A, b = 14.915 (5) A, c = 10.583 (2) A; alpha = 91.51 (2) degrees, beta = 108.99 (2) degrees, gamma = 102.86 (3) degrees; triclinic, space group P1, Z = 2; rho calcd = 1.363 g/cm3, rho obsd = 1.370 g/cm3.

Computers↗