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Biomedical subjects

H Schmidt

Publications and source records attributed to H Schmidt.

At least 541 records · Page 30Linked to original sources
PubMed1993

Direct desaturation of intact galactolipids by a desaturase solubilized from spinach (Spinacia oleracea) chloroplast envelopes.

In plants, polyenoic fatty acids are synthesized by desaturase enzymes which use acyl groups of membrane lipids as substrates. To provide direct 'in vitro' evidence for this reaction, we solubilized envelope membranes from spinach (Spinacia oleracea) chloroplasts with Triton X-100 to release a membrane-bound n-6 desaturase. In the presence of oxygen and reduced ferredoxin, the solubilized enzyme desaturated a variety of substrates, such as free oleic acid, free erucic acid, 1-oleoyl-sn-glycerol 3-phosphate and the three galactolipids 1-oleoyl-2-(7'-cis-hexadecenoyl)-3-beta-D-galactopyranosyl-sn-glycerol, 1,2-dioleoyl-3-beta-D-galactopyranosyl-sn-glycerol and the ether analogue 1,2-di-(9'-cis-octadecenyl)-3-beta-D-galactopyranosyl-sn- glycerol. The in vitro desaturation of these exogenously added complex lipids with ester- and ether-linked substrate chains is unambiguous evidence for lipid-linked desaturation. The enzyme measures the insertion of the new double bond from the methyl end and the existing (n-9)-cis-double bond of an appropriate acyl or alkyl chain. The distal part of the substrate group, normally the carboxy end of a fatty acyl residue, is of less importance and, in particular, its activation in thioester form is not required.

Chloroplasts↗
PubMed1993

Characterization of acetyl-CoA: L-lysine N6-acetyltransferase, which catalyses the first step of carbon catabolism from lysine in Saccharomyces cerevisiae.

The carbon catabolism of L-lysine starts in Saccharomyces cerevisiae with acetylation by an acetyl-CoA:L-lysine N6-acetyltransferase. The enzyme is strongly induced in cells grown on L-lysine as sole carbon source and has been purified about 530-fold. Its activity was specific for acetyl-CoA and, in addition to L-lysine, 5-hydroxylysine and thialysine act as acetyl acceptor. The following apparent Michaelis constants were determined: acetyl-CoA 0.8 mM, L-lysine 5.8 mM, DL-5-hydroxylysine, 2.8 mM, L-thialysine 100 mM. The enzyme had a maximum activity at pH 8.5 and 37 degrees C. Its molecular mass, estimated by sodium dodecyl sulphate-polyacrylamide gel electrophoresis, was 52 kDa. Since the native molecular mass, determined by gel filtration, was 48 kDa, the enzyme is a monomer.

Acetylation↗
PubMed1993

Recovery after anaerobic metabolism in the leech (Hirudo medicinalis L.).

Medicinal leeches (Hirudo medicinalis L.) responded to self-induced hypoxia (72 h) with typical anaerobic metabolism characterized by a decrease in adenylate energy charge, utilization of the substrates glycogen and malate, and accumulation of the main anaerobic end-products succinate and propionate. Propionate was also excreted into the medium. Ammonia excretion was suppressed. Aerobic recovery resulted in a profound O2 debt. Resynthesis of ATP was completed within 30 min. Disposal of succinate and restoring of malate required 2-3 h, and clearance of propionate and recharging of glycogen 6-12 h. Ammonia excretion did not exceed normoxic rates and excretion of propionate during recovery accounted for only 10% of total propionate accumulated during hypoxia. It is postulated that the clearance of succinate and propionate involves oxidation but also resynthesis of malate and glycogen. During hypoxia and recovery blood osmolality remained constant. The Na+ and Cl- ion concentrations in blood, the decrease of which was nearly equimolar during hypoxia, were re-established following different time-courses. Na+ concentration returned to normoxic levels after 2-3 h. The delayed increase in Cl- concentration, however, correlating with 6-12 h necessary to clear blood propionate, is interpreted as an anion regulating effect.

Acetates↗
PubMed1993

Effective long range mapping in Schizosaccharomyces pombe with the help of swi5.

The switching gene swi5 has a function in mating-type switching. In addition, the swi5 mutation causes an increased radiation sensitivity and reduces meiotic recombination about ten-fold. Based on the latter property, an experimental protocol was developed for using swi5 in long-range mapping in S. pombe. It is suitable for a speedy mapping of any new gene which has not yet been cloned. The procedure was used to clarify the map positions of some genes.

Chromosome Mapping↗
PubMed1993

Detection of Borrelia burgdorferi by nested polymerase chain reaction in cerebrospinal fluid and urine of children with neuroborreliosis.

Diagnosis of neuroborreliosis is often difficult since history and clinical presentation may be non-specific and serological tests may initially be negative. We therefore tested the polymerase chain reaction (PCR) for the detection of borrelial sequences in CSF and urine samples of consecutive children with neuroborreliosis seen in a single summer season. Four of eight children were negative in serum for antibodies to Borrelia burgdorferi. Two of eight children were PCR-positive in CSF and one other child was positive in urine. In two out of four children PCR was the only laboratory test confirming the clinical diagnosis. All children recovered after treatment with third generation cephalosporins. When seven of eight children were re-examined 6 months later all were healthy and antibodies to B. burgdorferi were detected in their serum. PCR may assist the paediatrician in establishing a diagnosis of neuroborreliosis; however, a negative result does not rule out neuroborreliosis. PCR is an adjunct, but no substitute for clinical judgement and serology.

Adolescent↗
PubMed1993

Cytogenetic analysis of a benzpyrene induced osteosarcoma in the rat (Rattus norvegicus).

A cytogenetic comparison of primary and transplant tumor cell-lines, both originating from a benzpyrene induced osteosarcoma, with normal rat cell-lines (Rattus norvegicus) is presented here. In all tumor cell-lines tested, the number of chromosomes was increased by one or two. Using Giemsa-banding, structural chromosomal changes, i.e. a Robertsonian translocation t(4;4)(q10;q10) and an interstitial deletion del(6)(q11q16) could be recorded. Furthermore, staining of nucleolus organizer regions (NORs) revealed a shift in NOR activity from chromosome number 11 to 12 and a decrease in NOR activity at chromosome number 3.

Aneuploidy↗
PubMed1993

Analysis of DNA repair pathways of Schizosaccharomyces pombe by means of swi-rad double mutants.

In Schizosaccharomyces pombe 11 different switching genes (swi1 to swi10 and rad22) are known which are involved in mating-type (MT) switching. Mutations in swi5, swi9, swi10 and rad22 also cause an increased radiation sensitivity. We tested whether the survival of these mutants after UV irradiation is influenced by caffeine. We included rad1 and rad13 mutants in our experiments which do not affect MT switching. Several double and triple mutants were constructed. We were able to assign the switching genes to different repair pathways: swi9 and swi10 are involved in excision repair, rad22 has a function in recombination repair, while swi5 appears to be involved in a hitherto unknown pathway. This 'swi5 pathway' is stimulated (!) by caffeine. Previously it was found that the swi5 mutation also reduces meiotic recombination. As to rad genes, we found a few inconsistencies with previous reports in the literature.

Alleles↗
PubMed1993

Differential regulation of human leukocyte antigen class I genes by interferon in vivo and in vitro.

Modulation of human leukocyte antigen (HLA) class I antigens on peripheral blood lymphocytes, monocytes, and hematopoietic precursors during interferon-alpha (IFN-alpha) therapy was investigated in 18 patients with myeloproliferative syndrome. After 1 month of IFN-alpha therapy, an increased number of monocytes and hematopoietic precursor cells but not of lymphocytes expressed HLA-DQ antigens. In addition, a strong induction of HLA class I antigens was found on both hematopoietic progenitors and normal peripheral blood mononuclear cells. By daily injections of IFN in the first month of therapy, stimulation continuously increased, suggesting a major effect of IFN alpha on hematopoietic progenitors with sustained enhanced expression of HLA class I antigens during differentiation of myelomonocytic cells. HLA class I antigen expression was consistently augmented by IFN alpha in all patients irrespective of their hematologic response. Differential in vivo regulation of HLA class I antigens by IFN was confirmed by comparison of HLA-A2 with HLA-B antigen expression. In vitro expression of the HLA-B7 and -Bw64 genes had been shown earlier to be significantly more inducible by IFN than the genes coding for several other HLA class I antigens after transfection into mouse L cells. Modification of the 5' ends of the HLA-B7 and HLA-B27 genes before transfection in mouse L cells showed the presence of enhancer sequences responding to IFN treatment in the 5' untranslated region of the HLA-B7 but not of the HLA-B27 gene and suggested the presence of independently acting regulatory mechanisms independent of these enhancers.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1993

Dependence of induction of enterobacterial AmpC beta-lactamase on cell-wall peptidoglycan, as demonstrated in Proteus mirabilis and its wall-less protoplast L-form.

The mobilizable plasmid pMD101 (ampR, ampC) was constructed by inserting cloned ampC, the structural gene for the chromosomal AmpC beta-lactamase of Citrobacter freundii, and the closely linked ampR encoding the transcriptional regulator essential for enzyme induction, into the broad host-range plasmid pKT231. Plasmid pMD101 was transconjugated into Proteus mirabilis VI and its isogenic, cell-wall-less protoplast L-form LVI. AmpC beta-lactamase was expressed constitutively from cloned ampR and ampC in bacteria and in some L-form protoplasts. However, induction of the enzyme by beta-lactam antibiotics occurred only in bacterial cells and not in the cell-wall- and peptidoglycan-deficient L-form. In agreement with current models, induction of AmpC beta-lactamase is thought to be initiated by an induction signal arising from the metabolic disturbance of cell-wall peptidoglycan.

Bacterial Proteins↗
PubMed1993

AmpG, a signal transducer in chromosomal beta-lactamase induction.

The chromosomal ampC beta-lactamase in Citrobacter freundii and Enterobacter cloacae is inducible by beta-lactam antibiotics. When an inducible ampC gene is introduced on a plasmid into Escherichia coli together with its transcriptional regulator ampR, the plasmid-borne beta-lactamase is still inducible. We have isolated mutants, containing alterations in a novel E. coli gene, ampG, in which a cloned C. freundii ampC gene is unable to respond to beta-lactam inducers. The ampG gene was cloned, sequenced and mapped to minute 9.6 on the E. coli chromosome. The deduced amino acid sequence predicted AmpG to be a 53 kDa, transmembrane protein, which we propose acts as a signal transducer or permease in the beta-lactamase induction system. Immediately upstream of ampG there is another 579-base-pair-long open reading frame (ORF) encoding a putative lipoprotein shown to be non-essential for beta-lactamase induction. We have found that ampG and this ORF form an operon, whose promoter is located in front of the ORF. Located closely upstream of the putative promoter is the morphogene bolA, which is transcribed in the opposite orientation. However, using transcription fusions, we have found that the ampG transcription is not regulated by bolA. In addition, we show that transcription is probably not regulated by either the starvation specific sigma factor RpoS, which controls bolA, or by AmpD the negative regulator for ampC transcription.

Amino Acid Sequence↗
PubMed1993

A contrast enhanced lesion of the III nerve on MR of a patient with ophthalmoplegic migraine as evidence for a Tolosa-Hunt syndrome.

A 23-year-old female with a six year history of migraine without aura twice developed a nearly complete internal and external III nerve paresis ipsilateral to her headache, two days after the onset of migraineous headache. An MR scan performed one week after the second episode showed a contrast enhanced lesion of the prepontine III nerve, where it enters the cavernous sinus on the left side. The headache, as well as the paresis ameliorated spontaneously. We suggest this is a further well documented case of "ophthalmoplegic migraine" which might reflect Tolosa-Hunt syndrome.

Adult↗
PubMed1993

IFN-gamma and TNF-alpha secretion by CD4+ and CD8+ TCR alpha beta + T-cell clones derived early after allogeneic bone marrow transplantation.

Secretion of the potentially antileukaemic cytokines IFN-gamma and TNF-alpha was investigated for CD4+ and CD8+ TCR alpha beta + T-cell clones derived from 4 leukaemia patients 3-6 weeks after allogeneic BMT. We investigated cytokine secretion in response to the activation signal accessory cells+phytohaemagglutinin+Interleukin 2. All clones derived after BMT were capable of IFN-gamma and TNF-alpha secretion, and both for CD4+ (n = 96) and CD8+ (n = 8) T cells quantities of IFN-gamma and TNF-alpha were significantly correlated with one another. When comparing the overall results for posttransplant and normal T-cell clones derived from 2 bone marrow donors (n = 65), both CD4+ and CD8+ TCR alpha beta + T-cell clones showed increased IFN-gamma production, and CD4+ but not CD8+ clones showed a decreased TNF-alpha secretion. The results suggest that noncytotoxic T cells derived after allogeneic BMT can produce IFN-gamma and TNF-alpha and may thus be capable of mediating antileukaemic effects.

Adult↗
PubMed1993

Inhibition of hepatic microsomal drug metabolism by atracurium administration in the rat.

The muscle relaxant atracurium is known to undergo extrahepatic degradation via Hofmann elimination and ester hydrolysis. The purpose of the present study was to evaluate the effects of atracurium on hepatic P450-dependent enzyme activities. Thirty-two male Sprague-Dawley rats were anaesthetized, mechanically ventilated, and randomly allocated to one of four study groups: group 1 received saline, group 2 atracurium, group 3 vecuronium, and group 4 pancuronium intravenously for a period of 3 hr. Equipotent doses of the muscle relaxants were applied; the doses had been obtained in a pilot study using evoked electromyography. At the end of the study period, the livers were removed and analyzed. All three muscle relaxants may lead to inhibition of hepatic drug metabolism. Atracurium influences hepatic P450, although it is predominantly degraded in extrahepatic tissues. Further studies are needed to evaluate the contribution of the major metabolite laudanosine to this inhibitory action.

Aminopyrine N-Demethylase↗
PubMed1993

Virulence determinants in nontoxinogenic Escherichia coli O157 strains that cause infantile diarrhea.

Ten sorbitol-fermenting Escherichia coli O157 strains that cause infantile diarrhea and are positive in the fluorescence actin staining test were determined to be negative for Shiga-like toxin (SLT) genes. We amplified their complete eae genes, contrasting them with those of SLT-producing E. coli O157 by restriction fragment length polymorphism analysis and nucleotide sequence analysis of a 400-bp stretch of the 3' end of eae. The data substantiated the presence of two eae genotypes within serogroup O157, one resembling eae of enteropathogenic E. coli (EPEC) strain E2348/69, found in nontoxinogenic E. coli O157 strains, and the other resembling eae of EHEC strain EDL 933, found in toxinogenic E. coli O157 strains. Another EPEC-specific virulence determinant was also shown to be large plasmids harboring EPEC adherence factor sequences. The SLT-negative E. coli O157 strains described here fall under the heading of EPEC, which serves as an explanation for their virulence in infants, and represent a third pathogroup within serogroup O157.

Adhesins, Bacterial↗
PubMed1993

Shiga-like toxin II-related cytotoxins in Citrobacter freundii strains from humans and beef samples.

By hybridizing colonies grown from 928 individual stool samples of patients suffering from diarrhea with oligonucleotide probes 772 and 849 complementary to Shiga-like toxin I (SLT-I) and SLT-II gene sequences, respectively, we identified two strains that hybridized with probe 849, which biochemical identification revealed as Citrobacter freundii. An additional five slt-II probe-positive isolates were screened from 81 beef samples. Polymerase chain reaction analysis and restriction of amplified products provided evidence for slt-II-related genes in all seven strains. From C. freundii LM 76, the genes encoding the A and B subunits were cloned in pUC 18 vectors and sequenced. The gene encoding the A subunit differed from that of Escherichia coli slt-IIvhc in 4 bases, resulting in two amino acid residue differences. In 11, 13, and 11 nucleotides, differentiation of slt-IIA, slt-IIcA, and vtx2haA, respectively, was found. These differences affected the predicted amino acid sequence as follows: there were six amino acid differences with SLT-IIA, five with SLT-IIcA, and four with VTx2haA. The nucleotide sequence of the gene encoding the B subunit is identical to slt-IIvhcB and differed from slt-IIcB and vtx2haB by only a single nucleotide base, but this resulted in a predicted amino acid sequence identical to that reported for these toxins. We therefore termed the toxin genes C. freundii slt-IIcA and slt-IIcB. Culture filtrates inoculated with material from the colonies from primary cultures were cytotoxic to Vero cells. Neutralization assays with antisera to E. coli SLT-I, SLT-II, and SLT-IIvhc revealed that antibodies against SLT-IIvhc reduced the C. freundii cytotoxic activity specifically and to the same degree as with the E. coli SLT-IIvhc control strain. In five of the seven strains tested, subcultivation on both a liquid or solid medium resulted in loss of cytotoxic activity. With polymerase chain reaction, we demonstrated that loss of cytotoxic activity ran parallel with the loss of slt genes. These data demonstrate the intergeneric occurrence of SLT-II-related toxins, which may well be a new marker of enteropathogenicity in C. freundii. Our findings that the toxin genes belong to the slt-II family plus their evident instability in the majority of strains should help pave the way to a better understanding of their role in diarrhea or food poisoning.

Animals↗