Biomedical subjects

J Frey

Publications and source records attributed to J Frey.

At least 217 records · Page 12Linked to original sources

PubMed1989

Study of basement membrane formation in dermal-epidermal recombinants in vitro.

Two different dermal-epidermal recombinants were prepared in vitro and used to study the synthesis and formation of basement membrane. The first was obtained by culturing keratinocytes on the surface of a collagen lattice populated by fibroblasts. The second was prepared by coculture of both keratinocytes and fibroblasts in a collagen lattice. After 6 weeks of culture, the basal lamina was observed with electron microscopy only if keratinocytes were cultivated on top of the collagen lattice populated by fibroblasts. In the second model, however, type IV collagen, laminin, and pemphigoid bullosa antigen could be detected by immunofluorescence as well as synthesis of type IV collagen in the culture, but no basement membrane was observed by electron microscopy. These data demonstrate that, in vitro, basement membrane formation depends not only on the presence of the macromolecular components but also on the culture conditions.

Basement Membrane↗

PubMed1989

Omegon-Km: a transposable element designed for in vivo insertional mutagenesis and cloning of genes in gram-negative bacteria.

To combine the features of the omega interposons with the advantages of in vivo transposition mutagenesis, we have constructed an artificial transposon, called Omegon-Km. The Omegon-Km transposon is carried on the plasmid pJFF350 which can be conjugally mobilized into a broad range of Gram-negative bacteria. Omegon-Km is flanked, in inverted orientation, by synthetic 28-bp repeats derived from the ends of IS1. In addition, each end of Omegon-Km has the very efficient transcription and translation terminators of the omega interposon. Internally, Omegon-Km carries the selectable kanamycin (Km)-neomycin resistance gene (alph A) which is expressed well in many Gram-negative bacteria. The IS1 transposition functions are located on the donor plasmid but external to Omegon-Km. Thus, insertions of Omegon-Km are very stable because they lack the capacity for further transposition. Omegon-Km mutagenesis is performed by conjugal transfer of pJFF350 from Escherichia coli into any Gram-negative recipient strain in which this plasmid is unable to replicate. Those cells which have had a transposition event are selected by their resistance to Km. Very high frequencies of Omegon-Km transposition were observed in Pseudomonas putida. Preliminary experiments with other Gram-negative soil and water bacteria (Rhizobium leguminosarum, Paracoccus denitrificans) yielded mutants at reasonable levels. The presence of an E. coli-specific origin of replication (ori) within Omegon-Km allows the rapid and easy cloning, in E. coli, of the nucleotide sequences flanking the site of the transposition event.

Blotting, Southern↗

PubMed1989

Collagen and lipid biosynthesis in a case of epitheliogenesis imperfecta in cattle.

An analysis of the dermis and a study of collagen and lipid biosynthesis by fibroblast cultures were carried out in one case of epitheliogenesis imperfecta in a new born calf. This was found to be not only an epidermal disease with fissures and blisters within basal cells on the basement membrane but also a metabolic disease affecting the dermal fibroblasts. These fibroblasts showed a significant decrease in collagen biosynthesis with an increase in the percentage of type III collagen and also a decrease in the biosynthesis of lipids, especially glycerides and cholesterol.

Animals↗

PubMed1989

Cloning and expression of a cohemolysin, the CAMP factor of Actinobacillus pleuropneumoniae.

The genetic determinant of the cohemolysin which is responsible for the CAMP phenomenon, a cohemolysis, of Actinobacillus pleuropneumoniae was cloned in Escherichia coli. Total DNA from the A. pleuropneumoniae serotype 1 type strain 4074 was used to construct a gene library in plasmid pUC18 in E. coli JM83. A total of 10,500 clones containing recombinant plasmids have been screened for hemolysis on blood plates. Fifty-five clones which showed a weak hemolytic response after 24 to 48 h of incubation were screened for the CAMP reaction with Staphylococcus aureus. This led to the identification of one clone which showed a positive CAMP reaction. Immunoblot analysis revealed that the recombinant strain expressed a protein with a molecular mass of 27,000 daltons, similar in size to the CAMP protein of the group B streptococci. Rabbit antibodies against the CAMP+ clone neutralized the CAMP reaction mediated by the E. coli strain containing the cloned CAMP gene as well as that of A. pleuropneumoniae. Antibodies raised against the cloned CAMP cohemolysin cross-reacted with Streptococcus agalactiae protein B. We designate the 27,000-dalton molecule CAMP factor protein and name its corresponding gene cfp.

Actinobacillus↗

PubMed1989

Informed consent as a framework for treatment: ethical and therapeutic considerations.

Significant changes in the nature of doctor-patient relationships have required more patient involvement in decision making and the therapeutic use of informed consent. Particularly in family therapy, open acknowledgment and discussion of potential hazards of psychotherapy not only establish trust, but also can deflect patients' resistance to therapy and strengthen their commitment to the therapeutic work. Three case reports of the successful use of "informed consent as a treatment strategy" are described.

Child↗

PubMed1989

Collagen biosynthesis anomalies in prolidase deficiency: effect of glycyl-L-proline on the degradation of newly synthesized collagen.

Prolidase deficiency is a rare hereditary disease characterized by an iminodipeptiduria especially composed by glycyl-L-proline which is not further degraded. The study of collagen metabolism in fibroblast cultures from three prolidase-deficient patients showed an increase in the rapidly degraded collagen and a decrease in the proline pool. In order to elucidate the mechanism of this metabolic disturbance, glycyl-L-proline was added to the cell cultures. In the control cultures, the addition of this dipeptide caused an increase in the rapidly degraded collagen and a decrease in the proline pool. The effects on the patient fibroblasts depended on the severity of the deficiency. The metabolic function of the dipeptide glycyl-L-proline was discussed in the light of these results.

Adult↗

PubMed1988

Genetic evidence for the existence of a repressor that modulates colicin D expression on plasmid ColD-CA23.

The plasmid ColD-CA23, a high copy number plasmid of 5.12 kb, contains genes for colicin D (cda), for immunity colicin D (cdi), and for a lysis function (cdl). These genes are arranged on a contiguous 2.4 kb fragment in the following sequence: cda, cdi, cdl. They are transcribed in two operons, one transcribing cda and cdl from a SOS inducible promoter, the other transcribing cdi in the opposite direction. The expression of cda and cdl is modulated by a repressor, cdr, which is encoded on the same transcript as cda and cdl. In the absence of this repressor, transcription from the SOS inducible colicin D promoter is exceptionally strong and leads to protein contents up to 50% of total cellular proteins. This autoregulative repressor is a new finding in the control mechanisms of expression of colicins. We have also identified the gene product of cdl to be a 10,000 dalton protein.

Binding Sites↗

PubMed1988

pUB307 mobilizes resistance plasmids from Escherichia coli into Neisseria gonorrhoeae.

The plasmid pUB307, a derivative of RP1, is a conjugative, broad-host-range plasmid. We have shown that this element mobilizes gonococcal resistance plasmids from Escherichia coli to Neisseria gonorrhoeae, thus providing evidence that extrachromosomal elements can efficiently enter gonococci by conjugation. Furthermore, pUB307 can also be used as a helper element to mobilize the cloning vector pLES2 into N. gonorrhoeae. This finding significantly increases the usefulness of pLES2 as a shuttle vector between E. coli and gonococcus.

Cloning, Molecular↗

PubMed1988

A bacteriophage T4 expression cassette that functions efficiently in a wide range of gram-negative bacteria.

We have constructed a derivative of the broad-host-range vector RSF1010. This plasmid, p alpha omega, contains an expression cassette derived from bacteriophage T4 gene 32, into which we have inserted the coding sequence for the xylE enzyme (C2,3O) of the TOL plasmid pWWO. The composite plasmid, p alpha xylE omega, was transferred by conjugal mobilisation into a variety of Gram-negative bacteria (Agrobacter, Paracoccus, Erwinia, Pseudomonas, Rhizobium and Xanthomonas). High levels of C2,3O activity were found in almost all of the extracts. Polyacrylamide gel electrophoresis of these extracts revealed a prominent protein band at 35 kDa whose identity as the C2,3O gene product was confirmed by immunoblotting. We have mapped the 5' ends of the gene 32/xylE hybrid transcripts. In all of the Gram-negative bacteria, the proximal P2 promoter is the most efficient promoter in the cassette. In most of the strains a weaker and more distal promoter activity (Pl) was also detected. In both uninfected and phage-infected Escherichia coli cells, the transcript produced from this promoter is processed at a specific site upstream from the gene 32 start codon. The same processing occurred in all the bacterial species examined. The decay of the hybrid xylE transcript has been analyzed in E. coli and Erwinia, and in both strains this mRNA was among the most stable.

Cloning, Molecular↗

PubMed1988

Regulation of hemolysin expression in Actinobacillus pleuropneumoniae serotype 1 by Ca2+.

Actinobacillus pleuropneumoniae, the causative agent of swine pleuropneumonia, secretes a hemolytic activity which is thought to be a factor involved in the pathogenesis of the disease. The biosynthesis of hemolysin by serotype 1 strain 4074 was strongly dependent on the activity of free Ca2+ in the growth medium. At activities of free Ca2+ below 50 microM, very low hemolytic activities could be detected in the growth medium and in cell extracts. Maximal hemolytic activities of up to 400 hemolytic units per ml could be measured in growth medium containing free Ca2+ activities above 3 mM. Other bivalent cations did not stimulate the production of hemolysin. Neither the growth rate nor the secretion of hemolysin was affected by increasing Ca2+ concentrations in the medium. The hemolysin of serotype 1 did not require Ca2+ as a cofactor for the lysis of erythrocytes. Ca2+ induced the expression of a 105-kilodalton protein, which was secreted. This protein comigrated with purified hemolysin on sodium dodecyl sulfate-polyacrylamide gel electrophoresis and exhibited hemolysin activity upon purification. Inhibition experiments with rifampin suggest that the hemolysin of A. pleuropneumoniae is regulated by Ca2+ at the transcriptional level. The threshold of hemolysin induction was around 700 microM free Ca2+, a concentration which is similar to that found in blood serum. The Ca2+-inducible hemolysin represents a novel type of positively regulated bacterial gene expression.

Actinobacillus↗

PubMed1988

[Association between immunity to collagen and the pathology of extracellular matrices].

In a first part, the data of immunogenicity of collagen type II and type IV in physiopathology of extracellular matrices as cartilage and basement membrane, are reviewed. However, in the second part, this conclusion will be examined by basing on the knowledge of the nature of specific sites of these collagens and the variable or non specific humoral immune response towards these collagens in human diseases.

Animals↗

PubMed1988

Multiple clustered dermatofibromas. Evolution over 20 years.

A 37 year-old man developed, over 20 years, multiple clustered dermatofibromas on the left thigh. Three similar cases have been described previously and the benign course of this lesion has been referred to. Analysis of collagen amino acids revealed a sharp increase of hydroxylysine on the fibrous central zone, suggesting an abnormal accumulation of type IV collagen.

Adult↗

PubMed1988

[DS-1000--a new digital image intensifier film-screen system in thoracic diagnosis].

The characteristics of the DS-1000, a new digital imaging equipment, using an image-intensifier-video-system are described. The digital images are displayed on a monitor with 1024 x 1024 matrix, are stored on a hard disc and can be postprocessed with regard to gray-scale-level and recognition of details. The spatial resolution ist 1,1 Lp/mm, using the 47-cm-image-intensifier. The DS 1000 was employed for chest-examinations. The diagnostic accuracy of 70 selected cases with a great variety of pathologic conditions was verified in comparison to conventional chest films. There was no significant difference in the recognition of normal anatomic structures between the two imaging systems. Pathologic states with low contrast to the surrounding structures (mediastinal and hilar masses, airspace disease, pulmonary nodules) were displayed at least equivalent with the digital technique. The accuracy in recognizing pathologic states with the digital equipment as compared to the conventional technique was inferior for conditions requiring high spatial resolution (subtle interstitial infiltrations, septal lines, scars, subtle calcifications).

Diagnosis, Differential↗

PubMed1988

Accumulation of glycolic acid and glyoxylic acid in serum in cases of transient hyperglycinemia after transurethral surgery.

Experimental data are presented here proving the accumulation of glycine in serum after transurethral prostatectomy and increased production of glycine metabolites: serine, alanine, glyoxylic acid, and glycolic acid. The presence of the metabolites glyoxylic acid and glycolic acid was demonstrated by gas-liquid chromatography and mass spectrometry. Glycine, glyoxylic acid, and glycolic acid possess neurological activity, so we examined the pathophysiology of the transurethral prostatectomy syndrome in view of the transient accumulation of these compounds in serum.

Aged↗

PubMed1988

Collagen biosynthesis in a case of infantile myofibromatosis.

Cells from a skin nodule from a patient with a recurrent form of familial myofibromatosis were cultivated in vitro. A metabolic study showed that these cells behaved like fibroblasts with collagen synthesis, a normal percentage of type III collagen, hydroxylation rate and the ability to contract a collagen gel. The main disturbances were the decreased synthesis and increased cell multiplication rate after a lag phase. This behavior was compared with that of a fibroblast culture from normal skin.

Collagen↗

PubMed1987

Characterization and structural analysis of Fc gamma receptors of human monocytes, a monoblast cell line (U937) and a myeloblast cell line (HL-60) by a monoclonal antibody.

A monoclonal antibody, FR51, raised against the IgG Fc receptor (Fc gamma R) of the human monoblast cell line U937 was used to analyze the distribution of this antigen on various human cells. This antibody inhibited the binding of human IgG to the Fc gamma R on U937 cells, HL-60 cells and human peripheral blood monocytes. In contrast, the Fc gamma R on human granulocytes (neutrophil cells) and on an Epstein-Barr virus-transformed human lymphoblastoid cell line (Raji) were not recognized, indicated by the failure of blocking the binding of human IgG ligand to the Fc gamma R on these cells. By affinity chromatography of detergent-containing cell free lysates of surface-iodinated U937 cells, HL-60 cells and monocytes, a protein of 70-kDa was isolated. This protein was identified as the Fc gamma R by rebinding the isolated protein to immobilized human IgG. Removal of the carbohydrate moiety with endo-beta-N-acetylglucosaminidase F demonstrated that the receptors consist of a 40-kDa polypeptide. Analysis of the polypeptide patterns obtained by proteolytic digestion of either mature (70-kDa) or deglycosylated (40-kDa) receptors isolated from monocytes, U937 cells and HL-60 cells strongly suggests that the Fc gamma R are identical. The monoclonal antibody FR51 specifically reacts with Fc gamma R on human monocytes, a myeloblast and a monoblast cell line but not with the receptors on a B cell line and neutrophil cells.

Antibodies, Monoclonal↗

PubMed1987

Collagen biosynthesis and isomorphism in a case of Ehlers-Danlos syndrome type VI.

Collagen metabolism was studied in fibroblasts grown from a skin biopsy specimen of a patient who presented the striking clinical features of Ehlers-Danlos syndrome and, in particular, hyperextensibility of the skin, hypermobility of the joints, and kyphoscoliosis. A reduction in lysine hydroxylation, characteristic of Ehlers-Danlos Type VI, was observed after labelling of the collagen with 14C-proline and 3H-lysine. Other modifications in the collagen metabolism of fibroblast cultures were noted, including an increase in collagen and total protein synthesis, and an increase in both the Type I and Type III collagen. The percentage of Type III collagen was, however, lower than in the control fibroblasts. The results point out the complexity of collagen disturbances in Ehlers-Danlos Type VI.

Adult↗

PubMed1987

Interposon mutagenesis of soil and water bacteria: a family of DNA fragments designed for in vitro insertional mutagenesis of gram-negative bacteria.

We have constructed a series of derivatives of the omega interposon [Prentki and Krisch, Gene 29 (1984) 303-313] that can be used for in vitro insertional mutagenesis. Each of these DNA fragments carries a different antibiotic or Hg2+ resistance gene (ApR, CmR, TcR, KmR or HgR) which is flanked, in inverted orientation, by transcription and translation termination signals and by synthetic polylinkers. The DNA of these interposons can be easily purified and then inserted, by in vitro ligation, into a plasmid linearized either at random by DNase I or at specific sites by restriction enzymes. Plasmid molecules which contain an interposon insertion can be identified by expression of its drug resistance. The position of the interposon can be precisely mapped by the restriction sites in the flanking polylinker. To verify their properties we have used these omega derivatives to mutagenize a broad host range plasmid which contains the entire meta-cleavage pathway of the toluene degradation plasmid pWW0 of Pseudomonas putida. Insertion of these interposons in the plasmid between the promoter and the catechol 2,3-dioxygenase (C23O) gene dramatically reduced the expression of this enzyme in Escherichia coli. We also show that when a plasmid containing an omega interposon is transferred by conjugal mobilization from E. coli to P. putida, Agrobacterium tumefaciens, Erwinia chrysanthemi, Paracoccus denitrificans or Rhizobium leguminosarum, the appropriate interposon drug resistance is usually expressed and, compared to the non-mutated plasmid, much reduced levels of C23O activity are detected. Thus, the selection and/or characterization of omega insertional mutations can be carried out in these bacterial species.

DNA Restriction Enzymes↗