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PubMed2002

[Echocardiographic evaluation of left atrial function in dilated cardiomyopathy with restricted and non-restricted Doppler transmitral flow].

UNLABELLED: Left atrial (LA) function is of great importance in left ventricular (LV) filling. There is evidence that echocardiographic Doppler evaluation of transmitral flow, routinely used for LV filling estimation, is dependent on LA function. Information regarding the relation of LA size and function to transmitral flow in heart failure is limited. We used 2D echocardiographic acoustic quantification methods to assess LA function in patients with dilated, (non-ischemic) cardiomyopathy (DCM) and a control group. The DCM group was divided into 2 subgroups: Group 1-with restrictive LV Doppler filling pattern-18 patients (DCM-R) and Group 2-with non-restrictive LV Doppler filling pattern-11 patients (DCM-NR) with similar heart rate, age and degree of mitral regurgitation. LA maximal area, total emptying fraction and absolute and fractional area change during rapid emptying and atrial contraction were calculated. The LA was enlarged only in DCM-R. Both DCM groups had decreased total emptying fractions and rapid emptying area changes compared to controls. An enlarged LA area and more decreased total emptying were found in DCM-R with high LV filling pressures compared to DCM-NR. The restrictive group had a significantly smaller LA rapid emptying area change, as well as a smaller LA area change and emptying fraction during atrial contraction compared to DCM-NR. Within < or = 2 hrs after the echocardiography study, cardiac catheterization was performed in the DCM group. We found significantly higher LV filling pressures and lower LV ejection fractions in DCM-R compared to DCM-NR. Significant correlations were found between LA function and invasive parameters like capillary wedge and LV enddiastolic pressures and LV EF. CONCLUSION: Patients with DCM-R had significantly enlarged LA areas with more depressed total emptying fractions and smaller LA area changes during contraction compared to DCM-NR. Thus, left atrial function plays an important role in LV filling and its dysfunction can be a marker of poor prognosis.

Adult↗

PubMed1990

The predominance of CD8+ T cells after infection with measles virus suggests a role for CD8+ class I MHC-restricted cytotoxic T lymphocytes (CTL) in recovery from measles. Clonal analyses of human CD8+ class I MHC-restricted CTL.

Stimulation of PBMC, in children recovering from acute measles, with autologous EBV-transformed and measles virus (MV)-infected lymphoblastoid cell lines (B-LCL) expanded primarily MV-specific CD8+ T cells. A large number of CD8+ T cell clones were obtained either by passaging of bulk cultures at limiting dilutions or by direct cloning of PBMC without previous stimulation in bulk culture. The MV-specific CD8+ T cell clones responding in a proliferative and a CTL assay were found to be class I MHC restricted. In contrast, CD4+ MV-specific T cell clones, which were generated by the same protocol, recognized MV in association with class II MHC molecules. Analysis of processing requirements for Ag presentation to CD8+ and CD4+ T cell clones, measured by the effect of chloroquine in a proliferative T cell response, revealed that both types of T cells recognized MV Ag processed via the endogenous/cytoplasmic pathway. Thus, these studies indicate that, as in most other viral infections and in contrast to previous suggestions, the class I MHC-restricted CTL response by CD8+ T cells may be an important factor in the control and elimination of MV infection. Therefore, the role proposed for CD4+ class II-restricted T cells in recovery from measles needs to be reevaluated.

Antigen-Presenting Cells↗

PubMed1977

[Long-term study on the influence of running exercise, food restriction, running exercise + food restriction and of parenterally applicated testis cells on age-parameters of the rat (author's transl)].

Preliminary results of a long-term study on 1100 male Sprague-Dawley rats are presented, showing the influence of running exercise, of restricted diet, of both of these together and of the s.c. application of lypholized testis cells. Up to now animals aged 9, 15 and 24 mths were investigated. The test-animals were exposed to the experimental conditions from their 6th month of life. The running exercise was carried out on a treadmill (30 m at a speed of 25 m/min, horizontal) 5 days a week. A food restriction of about 20% was achieved by 2 fasting-days a week. The lyophilized testis cells were injected for the first time at an age of 9 mths and afterwards in 6 mths intervals. Between the injection and the assessment of the age-parameters there was an interval of 6 mths. S o fare the following parameters of the comprehensive test-program have been evaluated: 1) running performance in m (treadmill, 20 m/min, ascent 15 degrees), 2) motor activity (Animex Activity Meter), 3) chemical contraction-relaxation of tail-tendon-fibers, 4) total lipids and total cholesterol in the plasma. The results obtained so far show that a mild regular training, moderate food restriction and the s.c. application of testis-cells are able to cause a significant shift in the dirction of a younger biological age in at least some of the age parameters. The action of the testis-cells seems to affect most of the age parameters but shows the tendency to be more distinct at a higher age. More concrete statements about the influence on the biological age or the vitality will only be possible after the multivariate analysis of the results.

Age Factors↗

PubMedDate not supplied

Reproduction and organ weight responses in female rats fed a restricted balanced diet or a restricted nonprotein calorie diet with or without supplemental glutamine or cysteine.

Primiparous Sprague-Dawley rats were used in two experiments to test the hypothesis that restriction of carbohydrate and fat calories alone versus comparable restriction of a balanced diet through gestation produces differential effects on gastrointestinal tract, reproductive tract and conceptus development. Rats were assigned on d 1 of pregnancy (13 rats/diet) to the following diets in Expt. 1: (1) balanced control diet fed ad libitum, C; (2) control diet fed at 50% of ad libitum intake, R; (3) restricted nonprotein energy diet containing twice the concentrations of protein, vitamins and mineral elements as the balanced control diet but fed at 50% of ad libitum intake of control diet, RCal; (4) R diet modified plus 0.21% L-cysteine, R + Cys; (5) R diet modified plus 1.54% L-glutamine, R + Glu. In Expt. 2, the treatment groups were: (1) Control diet, C; (2) C diet fed at 50% of ad libitum, R; (3) RCal diet fed at 50% of ad libitum intake of C; (4) RCal diet plus 2% L-glutamine fed at 50% of ad libitum intake of C. All pregnant rats were euthanized on d 20 postcoitum. Body weight of control dams was greater (P less than .01) than that of other dams in both experiments. Uterus weight and litter weight were less in RCal than in other dams in both experiments.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

PubMed1997

Amplified restriction fragment length polymorphism-based mRNA fingerprinting using a single restriction enzyme that recognizes a 4-bp sequence.

Using amplified restriction fragment length polymorphism (AFLP) technology, we have developed a new protocol for the fingerprinting of mRNA that allows systematic comparison of the differential expression of genes between mRNA samples. The major advantage of our protocol is the use of only a single restriction enzyme that recognizes a 4-bp sequence but allows screening of large numbers of different cDNAs. Using this new protocol, we compared mRNA samples obtained from the flower buds of two lines of the common morning glory (Ipomoea purpurea) with red and white flowers, respectively. Approximately 50 bands were observed in each lane of a denaturing polyacrylamide gel and the results were highly reproducible, as indicated by the results of analysis of two sets of independent mRNA samples. Two cDNA fragments, which were differentially amplified in the samples from the two lines, were shown to have been derived from a single gene that was actively expressed in the buds of red flowers but not in those of white flowers. A full-length cDNA of this gene was cloned from a bud cDNA library. Sequence analysis showed that this cDNA carries a sequence highly homologous to the chalcone synthase (CHS) genes, the key enzyme in the flavonoid biosynthetic pathway.

Amino Acid Sequence↗

PubMed1989

The EcoDXX1 restriction and modification system: cloning the genes and homology to type I restriction and modification systems.

The Escherichia coli plasmid pDXX1 codes for a type I restriction and modification system, EcoDXX1. A 15.5-kb BamHI fragment from pDXX1 has been cloned and contains the hsdR, hsdM, and hsdS genes that encode the EcoDXX1 system. The EcoDXX1 hsd genes can complement the gene products of the EcoR124 and EcoR124/3 hsd systems, but not those of EcoK and EcoB. Hybridization experiments using EcoDXX1 hsd genes as a probe demonstrate homology between EcoDXX1 and EcoR124 and EcoR124/3 restriction-modification systems, but weak or no homology between EcoDXX1 and EcoK or EcoB systems.

Blotting, Southern↗

PubMed1985

The EcoDXX1 restriction and modification system of Escherichia coli ET7. Purification, subunit structure and properties of the restriction endonuclease.

The Escherichia coli plasmid pDXX1 codes for a new restriction-modification system. The specific restriction endonuclease coded by this system has been purified by a procedure that includes phosphocellulose and heparin-agarose chromatography. Sedimentation on glycerol gradients showed one peak of activity with a value of about 12 S. The highly purified enzyme require ATP and Mg2+ for activity as well as S-adenosylmethionine, although some S-adenosylmethionine molecules are probably bound to the enzyme. The enzyme does not cleave lambda DNA at well-defined sites and has a strong non-modified DNA-dependent ATPase activity. The enzyme has also methylase activity acting against non-modified DNA.

Adenosine Triphosphatases↗

PubMed2005

Improved assessment of denitrifying, N2-fixing, and total-community bacteria by terminal restriction fragment length polymorphism analysis using multiple restriction enzymes.

A database of terminal restriction fragments (tRFs) of the 16S rRNA gene was set up utilizing 13 restriction enzymes and 17,327 GenBank sequences. A computer program, termed TReFID, was developed to allow identification of any of these 17,327 sequences by means of polygons generated from the specific tRFs of each bacterium. The TReFID program complements and exceeds in its data content the Web-based phylogenetic assignment tool recently described by A. D. Kent, D. J. Smith, B. J. Benson, and E. W. Triplett (Appl. Environ. Microb. 69:6768-6766, 2003). The method to identify bacteria is different, as is the region of the 16S rRNA gene employed in the present program. For the present communication the software of the tRF profiles has also been extended to allow screening for genes coding for N2 fixation (nifH) and denitrification (nosZ) in any bacterium or environmental sample. A number of controls were performed to test the reliability of the TReFID program. Furthermore, the TReFID program has been shown to permit the analysis of the bacterial population structure of bacteria by means of their 16S rRNA, nifH, and nosZ gene content in an environmental habitat, as exemplified for a sample from a forest soil. The use of the TReFID program reveals that noncultured denitrifying and dinitrogen-fixing bacteria might play a more dominant role in soils than believed hitherto.

Bacteria↗

PubMed1993

Epidemiological fingerprinting of Enterobacter cloacae by small-fragment restriction endonuclease analysis and pulsed-field gel electrophoresis of genomic restriction fragments.

A cluster of infections caused by Enterobacter cloacae was observed among preterm neonates in a neonatal intensive care unit (NICU) of a pediatric hospital in Osnabrück, Germany. The presence of similar antimicrobial susceptibility patterns among the bacterial isolates prompted an investigation to determine whether a limited spread of a single strain existed. All 12 E. cloacae isolates from the NICU and 50 nonrelated strains were fingerprinted by small-fragment restriction endonuclease analysis (SF-REA) of EcoRI DNA digests. Selected isolates were further characterized by pulsed-field gel electrophoresis (PFGE) of NotI- or XbaI-generated genomic restriction fragments. Epidemiologically unrelated strains were clearly discriminated by both methods. Results achieved by SF-REA and PFGE revealed that of the 12 isolates from the NICU, 11 belonged to the same genotypic cluster. Since all reagents and equipment for both techniques are commercially available, DNA fingerprinting by SF-REA or PFGE is proposed as a useful tool in the microbiology laboratory for investigating the epidemiological relatedness of E. cloacae strains of clinical and environmental origin.

Cross Infection↗

PubMed1996

Restriction fragment analysis of fowlpox virus using Bgl I, Bam HI, Hha I and Sma I restriction endonucleases.

Fowlpox virus vaccine strain and two field isolates collected from out breaks of disease were purified from cell culture using sucrose gradient ultracentrifugation. The viral DNAs were digested with Bgl I, Bam HI, Hha I and Sma I restriction endonucleases and the fragment pattern was analysed on 0.7% agarose gel. Bgl I digestion produced 54 fragments of size ranging from 31.50 to 0.60 kb, having similar electrophoretic mobilities in both the vaccine strain and two field isolates. Only 9 well resolved and one unresolved or partially digested fragment were obtained after Bam HI digestion. A total number of 29 and 41 fragments were obtained with Hha I and Sma I respectively. Almost similar restriction fragment pattern was observed in vaccine strain and the field isolates. The total genomic size was calculated to be between 265.00 and 302.91 kb. The three viruses were found to be genetically similar.

Animals↗

PubMed1975

Restriction and modification in B. subtilis. Nucleotide sequence recognised by restriction endonuclease R. Bsu R from strain R.

Restriction endonuclease R from Bacillus subtilis strain R cleaves nonmodified SPP 1 DNA in approximately 80, and lambda DNA in about 200 different sites. DNA digests with this endonuclease and with endonuclease Hae III from Haemophilus aegyptius show identical fragmentation patterns on gel electrophoresis, indicating that the two enzymes recognise the same nucleotide sequence. The polynucleotide kinase reaction was used in conjunction with two-dimensional ionophoretic nucleotide mapping methods to identify the 5'-nucleotide sequences at the sites of cleavage by the B. subtilis restriction endonuclease. The results show that the recognition sequence is (see article) where arrows indicate the points of strand scission. Each of the four possible nucleotides can occur in the positions flanking the recognition site.

Bacillus subtilis↗

PubMed1989

Alcelaphine herpesviruses 1 and 2 SDS-PAGE analysis of virion polypeptides, restriction endonuclease analysis of genomic DNA and virus replication restriction in different cell types.

Herpesviruses have been isolated from white-tailed, white-bearded and blue wildebeest, as well as from Jimela topi and Cape hartebeest. These animals are members of the sub-family Alcelaphinae of the family Bovidae. Viruses isolated from wildebeest cause malignant catarrhal fever (MCF) in susceptible ruminant species. Alcelaphine herpesviruses (AHV) isolated from wildebeest replicate in both fetal aoudad sheep kidney (FAK) cells and bovine embryonic lung (BEL) cells. However, virus isolates from topi and hartebeest, which have not been linked to clinical MCF, replicate only in FAK cells. Buoyant density analysis by analytical ultracentrifugation, restriction endonuclease analysis and blot hybridization of virus genomic DNA from both alcelaphine herpesviruses as well as from bovine herpesviruses 1, 2, and 4 demonstrate that there are two types of alcelaphine herpesviruses, each distinct and different from the other bovine herpesviruses. Genomic size of both alcelaphine herpesviruses, estimated from DNA restriction fragments, is approximately 110 kilobase pairs. Alcelaphine herpesvirus DNA resembles Herpesvirus saimiri DNA during equilibrium sedimentation in that the majority of the DNA bands as a light (L) fraction with a minor heavy (H) component. Polyacrylamide gel analysis of virion proteins indicates that both viruses have distinct patterns, each consisting of 36 polypeptides ranging in molecular weight from 12,000 to 275,000. Virus isolates from wildebeest have been designated AHV-1, while viruses isolated from topi and hartebeest have been designated AHV-2.

Animals↗

PubMed1995

Cloning of a pair of genes encoding isoschizomeric restriction endonucleases from Bacillus species: the BspEI and BspMII restriction and modification systems.

The respective genes (R-M) encoding restriction and modification systems from two Bacillus species which recognise the same nucleotide sequence, 5'-TCCGGA, have been cloned and expressed in Escherichia coli. The BspEI R-M genes were cloned on a 3.6-kb HindIII fragment, whereas the BspMII R-M genes were cloned on three contiguous HindIII fragments totalling 9.8 kb. Upon thermal induction, E. coli carrying the bspEIR clones under the control of the phage lambda PL promoter, express high levels of R.BspEI (10(6) units/g wet cell paste). The bspMIIR gene, on the other hand, is only poorly expressed (about 4 x 10(3) units/g wet cell paste) following induction. Although the enzymes of both R-M systems recognize the same sequence and the restriction endonucleases (ENases) cleave DNA at the same position, the modification specified by the methyltransferases (MTases) differ. The internal cytosine is the site of M.BspMII modification (TCmeCGGA), whereas the external cytosine is modified by M.BspEI (TmeCCGGA). The two R-M systems probably evolved independently.

Bacillus↗

PubMed1991

The distribution of restriction fragment lengths for non-overlapping restriction sites in a random DNA sequence model.

Overlapping subsequences in a DNA sequence are not independent even if independence is supposed for the single nucleotides. Therefore the often used geometric distribution for the length of restriction fragments is not exact. The exact distribution of this random variable is derived for non-overlapping restriction sites in a DNA sequence with an infinite (or very large) number of nucleotides. Correction to the finite case is easy. It is shown that the simple geometric distribution is a good approximation as long as the basic probability for the occurrence of the recognition sequence at a given site is small.

Base Sequence↗

PubMed2006

First restriction and genetic mapping of the genomic DNA of urease-positive thermophilic campylobacters (UPTC), and small restriction fragment sequencing.

A restriction and genetic map of urease-positive thermophilic campylobacter (UPTC) CF89-12 genome DNA is constructed using a pulsed-field gel electrophoresis procedure after digestion with SalI and SmaI and Southern blot hybridisation. Each of the six gene fragments (flaA, glyA, lysS, recA, sodB and ureAB) selected are mapped in only a fragment on the restriction map. Three DNA fragments for rrn operon probes are mapped in multiple regions on the map. When two SmaI-digested neighbouring small fragments hybridised with rrn probes are cloned and sequenced, a total sequence length of 7487 bp is determined. In the sequence, part of the pnp gene (734 bp) bearing a p-independent transcriptional termination region, a cluster of five tRNA genes including the putative promoter region, a hypothetical Cj0171-like 507-bp sequence containing an internal termination codon, and a part of the rrn operon including the putative promoter region (4700 bp) are identified. The 507 bp sequence carried both putative transcriptional promoter sequences, including a ribosome binding site upstream of the ATG start codon and a characteristic G9 structure, and a possible p-independent transcriptional termination region. A hypothetical Cj0170-like 204-bp sequence containing an internal termination codon also occurred, overlapping partly with the Cj0171-like sequence. Based on nucleotide sequence alignment analysis between the UPTC rrn operon examined here and the previously reported one, two different 16S-23S ribosomal DNA (rDNA) internal spacer regions are shown to exist.

Base Sequence↗

PubMed2001

Families of restriction enzymes: an analysis prompted by molecular and genetic data for type ID restriction and modification systems.

Current genetic and molecular evidence places all the known type I restriction and modification systems of Escherichia coli and Salmonella enterica into one of four discrete families: type IA, IB, IC or ID. StySBLI is the founder member of the ID family. Similarities of coding sequences have identified restriction systems in E.coli and Klebsiella pneumoniae as probable members of the type ID family. We present complementation tests that confirm the allocation of EcoR9I and KpnAI to the ID family. An alignment of the amino acid sequences of the HsdS subunits of StySBLI and EcoR9I identify two variable regions, each predicted to be a target recognition domain (TRD). Consistent with two TRDs, StySBLI was shown to recognise a bipartite target sequence, but one in which the adenine residues that are the substrates for methylation are separated by only 6 bp. Implications of family relationships are discussed and evidence is presented that extends the family affiliations identified in enteric bacteria to a wide range of other genera.

Amino Acid Sequence↗

PubMed1993

Delayed expression of in vivo restriction activity following conjugal transfer of Escherichia coli hsdK (restriction-modification) genes.

Following conjugal transfer of the hsdK genes (hsdRK, hsdMK, and hsdSK) of Escherichia coli K-12, restriction activity was first detected only after approximately 15 generations, whereas modification activity was observed immediately. This sequential expression explains the establishment of hsdK genes in a nonmodified host and suggests regulation of restriction activity after conjugal transfer.

Conjugation, Genetic↗

PubMed2003

Identification of Mycobacterium avium genotypes with distinctive traits by combination of IS1245-based restriction fragment length polymorphism and restriction analysis of hsp65.

One-hundred eight Mycobacterium avium isolates from pigs, humans, birds, and bovines were typed by the IS1245-based restriction fragment length polymorphism (RFLP) method and PCR-restriction enzyme analysis (PRA) of hsp65. Nine clusters of isolates showing more than 80% similarity in their RFLP profiles were detected. The largest cluster (cluster B) included 32 of 79 pig isolates (40.5%), 3 of 25 human isolates (12%), and 1 of 2 bovine isolates, comprising 33% of all isolates. The second largest cluster (cluster A) included 18 pig isolates (22.8%) and 6 human isolates (24%). Six smaller clusters included six pig isolates (clusters C and D), four and two human isolates (clusters E and F, respectively), two pig isolates (cluster I), and two pig isolates plus one bovine isolate and the avian purified protein derivative strain (cluster H). Cluster G represented the "bird-type" profile and included the bird isolate in this series, one pig isolate, plus reference strain R13. PRA revealed four allelic variants. Seventy-seven isolates were identified as M. avium PRA variant I, 24 were identified as M. avium PRA variant II, 6 were identified as M. avium PRA variant III, and 1 was identified as M. avium PRA variant IV. Except for three isolates from cluster B, each of the RFLP clusters was associated with a single PRA pattern. Isolates with unique (nonclustered) RFLP profiles were distributed between PRA variants I and II, and there was one unique isolate of PRA variant IV. These observations are consistent with divergent evolution within M. avium, resulting in the emergence of distinct lineages with particular competence to infect animals and humans.

Animals↗