Biomedical subjects
Publications and source records attributed to C Koch.
Life-threatening polymicrobial bacteremia in a 7 1/2-year-old boy, was found to be caused by willful contamination of i.v. drips by the mother. The boy, as well as his diseased twin sister, had histories of long-lasting chronic otitis of unknown etiology. The importance of obvious pathologic psycho-social factors was overlooked, and diagnosis was only reached by close collaboration with clinical microbiologists.
N-acetylcysteine (NAC) is known to be a scavenger of free oxygen radicals, and recent in vitro studies have demonstrated that it is also able to inhibit leukocyte function. The clinical significance of these effects is, however, not known. In this study we have measured the effect on human blood neutrophil and monocyte function of a single 400 mg dose of NAC administered orally. Administration of NAC to ten healthy volunteers resulted in significant reduction of neutrophil chemiluminescence response following activation by opsonized zymosan as compared to four non-treated persons acting as controls. No effect was observed on the chemotaxis of either cell type or on monocyte chemiluminescence response. These findings suggest that NAC may be beneficial in clinical conditions like cystic fibrosis, where tissue damage may be a consequence of the effects of increased release of toxic oxygen radicals and proteolytic enzymes.
A hemolytic assay was developed with the primary aim of being able to identify human lymphocytes producing anti-dsDNA antibodies found in patients with systemic lupus erythematosus (SLE). The coating of sheep red-blood cells with DNA was performed after precoating the cells with poly-L-lysine. The DNA-SRBC were lysed by anti-DNA antibodies from SLE sera, and the percent hemolysis was found to correlate with the anti-DNA activity demonstrated by the Farr assay (r = 0.87). Single-stranded DNA at the surface of the coated cells could be removed after digestion with nuclease S1. The effect of the digestion was verified by SLE serum specific for single-stranded DNA. With slight modifications, the target cells may be used to determine not only the titer of anti-DNA antibodies but also the complement-consumption and immunoglobulin classes of the anti-DNA antibodies.
Using a mice in a tetanus vaccination system, we have examined the mode of action of the immunological adjuvant, aluminium hydroxide (Al(OH)3), on antibody titers as measured by an ELISA-method. A profound effect of the adjuvant was observed after primary immunizations, increasing titers about 100-fold compared to plain (not adjuvanted) controls. However, no effect on titers could be attributed to a content of Al(OH)3 in the vaccines when used for booster immunizations, as the increase in titers after boosting was either the same for groups given plain or adsorbed toxoids, or even higher in the groups receiving the plain vaccine. Thus, the effect of Al(OH)3 as an adjuvant seems to be exerted mainly on the primary antibody response. Investigating the mode of action by the adjuvant, we adsorbed a certain amount of toxoid onto varying doses of Al(OH)3. An antigen ELISA revealed that all toxoid had been adsorbed to the Al-gel in the preparations. The antibody titers after immunization showed a significantly higher response to toxoid adsorbed to the higher amount of adjuvant, indicating that this effect was most probably due to free, not antigen-bound, Al-particles. Moreover, this effect could be inhibited when BSA was added in excess to the preparation, thereby blocking the free residues on the Al-gel.
A combined method of the Limulus amebocyte lysate (LAL) test and the enzyme-linked immunosorbent assay for quantitation of endotoxin was developed based on our observation that the antigenicity of coagulogen, a major protein in LAL, was lost when LAL reacted with endotoxin as shown by immunoblotting. Determination of the residual coagulogen by an enzyme-linked immunosorbent assay system with monoclonal antibody against coagulogen revealed that the loss of the antigenicity of coagulogen was proportional to the concentration of endotoxin. An inverse linear curve was established between the endotoxin concentration and absorbance. Standard curves of the LAL enzyme-linked immunosorbent assay with different detection limits (from 0.1 to 100 pg of the control standard endotoxin per ml) were obtained from one batch of commercial LAL by adjusting incubation time and dilution of LAL. The reaction curves of various endotoxins were parallel to one another, whereas the kinetics differed from that of (1-3)-beta-D-glucan. The LAL enzyme-linked immunosorbent assay is a highly reproducible microassay, using only 10 microliter of test sample and LAL reagent; because the color and turbidity of plasma samples do not interfere with the assay, it is well suited for quantitation of endotoxins in clinical specimens.
A 5.5-year survey revealed 9 patients with allergic bronchopulmonary aspergillosis (ABPA) and 1 patient with a highly probable diagnosis among 200 Danish patients with cystic fibrosis (CF). The incidence of ABPA was 0.9 (0.4-1.7, 95% confidence limits) per 100 patients per year. All the patients with definite or probable ABPA had serum antibodies to the Aspergillus fumigatus catalase antigen, and in 5 patients the appearance of catalase antibodies during the incipient phase of ABPA was documented. 29 patients without ABPA also had catalase antibodies, but in lower levels than ABPA patients (p = 0.006). The 6-month prevalence rate of A. fumigatus in sputum was 80 and 72% in the 2 groups, respectively. The finding of catalase antibodies in some CF patients without ABPA may indicate the occurrence of a symptom-poor form of pulmonary aspergillosis.
71 children and adults (median age 7 years) with chronic obstructive pulmonary disease in which ampicillin-sensitive Haemophilus influenzae were isolated from lower airway secretions were included in a single-blind study comparing amoxycillin/clavulanate and amoxycillin alone. The dosage of amoxycillin was 50 mg/kg/day given together with probenecid and divided in 3 doses. Duration of treatment was 14 days. Clinical and bacteriological examinations were performed at study entry and again immediately after the treatment period. A late bacteriological follow-up 1.5 months after entry was performed. 65 patients were eligible for analysis of clinical outcome, and no difference between the groups was found. Side-effects were mild at a frequency of 3% for either preparation. In terms of eradication of the initially isolated H. influenzae amoxycillin/clavulanate tended to be better than amoxycillin, although the difference was not significant (70% and 57%, respectively). In a subset of 33 patients with polymicrobial flora amoxycillin/clavulanate was significantly more effective than amoxycillin. However, amoxycillin/clavulanate did not significantly reduce the emergency of beta-lactamase producing H. influenzae during treatment, and thus offers no advantage over amoxycillin in patients with amoxycillin-sensitive H. influenzae. The combination should be reserved to patients with either polymicrobial flora or ampicillin-resistant H. influenzae.
The beta-cell response to an oral glucose load was studied in 22 patients with cystic fibrosis (CF) by means of insulin, C-peptide and proinsulin, and the results compared with those from 20 healthy sex and age matched controls. According to WHO criteria two had diabetes mellitus, eight had impaired glucose tolerance and twelve had normal glucose tolerance. All patients showed lower insulin and C-peptide levels than the controls at 30 minutes. However the insulin and C-peptide responses were sustained so that the areas under the curves were comparable between controls, CF patients with normal glucose tolerance, and CF patients with impaired glucose tolerance. By contrast, the area under the proinsulin curve was significantly higher in the CF patients with impaired glucose tolerance compared with both controls (p less than 0.05) and with the CF normal glucose tolerance group (p less than 0.02). In the CF patients with impaired glucose tolerance the proinsulin level was significantly elevated at 120 minutes (median 50 pmol/l) and at 180 minutes (29 pmol/l) as compared with the controls (24 and 19 pmol/l p less than 0.01) and with the CF patients with normal glucose tolerance (21 and 16 pmol/l p less than 0.01). These data confirm that impaired glucose tolerance and diabetes is frequent in cystic fibrosis. The elevated proinsulin immunoreactive material in CF patients with impaired glucose tolerance may partly compensate for the relative insufficient insulin response found in these patients.
A murine hybridoma cell line (aDNA35I9) secreting anti-DNA antibodies was used as a model for haemolytic plaque forming cells in an assay where DNA-conjugated sheep red blood cells (DNA-SRBC) were used as target cells. DEAE-dextran, employed to abolish the anti-complementary activity of agar gel, completely inhibited anti-DNA plaques. This problem was solved by using agarose instead of agar. Since the occurrence of small plaques may make reading of the test difficult, it was established that plaque size could be increased by decreasing the antigen density on the target cells. Free DNA in the gel inhibited plaque formation, indicating the specificity of the assay. Spleen cells from mice of the strains MRL/MP and NZB/W which are known to develop a stage of autoimmunity, produced plaques in numbers which were correlated to the age of the mice and to the anti-DNA antibody level in serum.
The effect of peroral N-acetylcysteine (NAC) in patients with cystic fibrosis (CF) and primary ciliary dyskinesia (PCD) was investigated. 41 CF patients and 13 PCD patients completed the study which was a double-blind, placebo-controlled, cross-over trial. The patients received either NAC or placebo for two periods of three months followed by a three month follow-up period. Active treatment consisted of NAC, either 200 mg x 3 daily (patients weighing less than 30 kg) or 400 mg x 2 daily (greater than 30 kg). The effect was evaluated in terms of a subjective clinical score, weight, sputum bacteriology, blood leucocyte count, sedimentation rate, titres of specific antimicrobial antibodies, lung function parameters and measurement of the ciliary function. No effect was seen in PCD patients, but in CF patients an improved lung function was seen in the period when the patients suffer most from lower airway infections.
Here we characterize FIS (factor for inversion stimulation), a new cellular component of the lambda site-specific recombination pathway. This host protein binds to a specific region in the lambda attP overlapping the Xis binding sites and can bind cooperatively with Xis to these sites. FIS stimulates lambda excision up to 20-fold in vitro in the presence of suboptimal Xis concentrations, but has no effect in the presence of saturating Xis; FIS has no effect on integrative recombination. FIS can replace one Xis molecule in a series of cooperative and competitive interactions but cannot carry out excision in the absence of Xis. FIS's role in the regulation of recombination has been inferred from in vivo modification of DNA. In exponentially growing cells the lambda FIS site is fully occupied, whereas in stationary-phase cells this binding site is vacant.
DNA was conjugated to sheep red blood cells (SRBC) by chemical methods using CrCl3, poly-L-lysine or methylated bovine serum albumin as conjugation agents and by a physical method where conjugation was accomplished by incubation at 45 degrees C. The degree of conjugation was estimated using 32P-DNA (mean size 1 kbase pairs). Employing the CrCl3 method 5.8 +/- 3.6 micrograms DNA were conjugated per 10(8) SRBC at a concentration of 70 micrograms DNA/10(8) cells. At the same DNA concentration in the incubation medium 3.0 +/- 0.6 microgram DNA/10(8) cells were conjugated by poly-L-lysine, 4.1 +/- 0.8 microgram DNA/10(8) cells by methylated bovine serum albumin and approximately 4 micrograms DNA/10(8) cells when the cells were incubated at 45 degrees C. Cells conjugated with DNA by CrCl3 showed linearly increasing conjugation with increasing concentration of DNA. Cells conjugated by poly-L-lysine (pLL) or methylated bovine serum albumin seemed to be saturated by DNA at 30 micrograms DNA per 10(8) cells. At 45 degrees C the spontaneous adhesion of DNA to SRBC increased in the concentration range investigated. The degree of conjugation of DNA to SRBC was influenced by pH, and Ca2+.pLL-conjugated DNA-SRBC, but none of the other preparations were lysed in a hemolytic assay using anti-DNA antiserum from a patient with systemic lupus erythematosus.
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Increased levels of intracellular calcium at either pre- or postsynaptic sites are thought to precede changes in synaptic strength. Thus, to induce long-term potentiation in the hippocampus, periods of intense synaptic stimulation would have to transiently raise the levels of cytosolic calcium at postsynaptic sites--dendritic spines in the majority of cases. Since direct experimental verification of this hypothesis is not possible at present, calcium levels have been studied by numerically solving the appropriate electro-diffusion equations for two different postsynaptic structures. Under the assumption that voltage-dependent calcium channels are present on dendritic spines, free intracellular calcium in spines can reach micromolar levels after as few as seven spikes in 20 milliseconds. Moreover, a short, but high-frequency, burst of presynaptic activity is more effective in raising levels of calcium and especially of the calcium-calmodulin complex than sustained low-frequency activity. This behavior is different from that seen at the soma of a typical vertebrate neuron.
Poggio and Reichardt (Kybernetik, 13:223-227, 1973) showed that if the average response of a visual system to a moving stimulus is directionally selective, then this sensitivity must be mediated by a nonlinear operation. In particular, it has been proposed that at the behavioral level, motion-sensitive biological systems are implemented by quadratic nonlinearities (Hassenstein and Reichardt: Z. Naturforsch., 11b:513-524, 1956; van Santen and Sperling: J. Opt. Soc. Am. [A] 1:451-473, 1984; Adelson and Bergen: J. Opt. Soc. Am. [A], 2:284-299, 1985). This paper analyzes theoretically two nonlinear neural mechanisms that possibly underlie retinal direction selectivity and explores the conditions under which they behave as a quadratic nonlinearity. The first mechanism is shunting inhibition (Torre and Poggio: Proc. R. Soc. Lond. [Biol.], 202:409-416, 1978), and the second consists of the linear combination of the outputs of a depolarizing and a hyperpolarizing synapse, followed by a threshold operation. It was found that although sometimes possible, it is in practice hard to approximate the Shunting Inhibition and the Threshold models for direction selectivity by quadratic systems. For instance, the level of the threshold on the Threshold model must be close to the steady-state level of the cell's combined synaptic input. Furthermore, for both the Shunting and the Threshold models, the approximation by a quadratic system is only possible for a small range of low contrast stimuli and for situations where the rectifications due to the ON-OFF mechanisms, and to the ganglion cells' action potentials, can be linearized. The main question that this paper leaves open is, how do we account for the apparent quadratic properties of motion perception given that the same properties seem so fragile at the single cell level? Finally, as a result of this study, some system analysis experiments were proposed that can distinguish between different instances of the models.
The clinical efficacy of the conventional aminoglycoside plus beta-lactam treatment was compared to that of monotherapy with oral quinolones in 26 adult cystic fibrosis patients in an open prospective clinical trial in which six two-week courses of antipseudomonas treatment were administered with an interval of approximately three months between treatments. In each patient two courses of conventional treatment were followed by two courses of quinolone treatment and then by another two courses of conventional treatment. The observed improvements in pulmonary function were somewhat higher when the patients received conventional treatments, and in the most seriously affected patients conventional treatment was significantly better than quinolone treatment. On the basis of these findings it is suggested that quinolone monotherapy cannot replace conventional antipseudomonal chemotherapy in patients with severe pulmonary involvement.
We report a family in which three males in two generations had meningococcal infections; one of them died. Hemolytic activity of the alternative complement pathway in the two survivors and in one healthy boy belonging to the family was reduced, as measured in a kinetic system. These three individuals had 10-11% of normal properdin concentration in plasma, as measured by a catching ELISA method, while the other complement components were normal. Hemolytic complement activity was normalized when purified properdin was added. The data are compatible with an X-linked mode of inheritance of properdin deficiency.
A polymorphic epitope in the third component of human complement was studied. This allotypic system is distinct from the electrophoretically determined C3 S/F polymorphism and is defined by the recognition of one allotype by a monoclonal antibody. Allotypic protein variants, C3F+ (reactive with this antibody) and C3S- (non-reactive with the antibody), were purified. Deglycosylation studies and N-terminal sequencing of CNBr fragments, reactive with the antibody, revealed that the polymorphic epitope was present in a beta chain fragment of mol. wt 20,000. In the intact C3 molecule, this fragment is situated with N-terminus at residue No. 202, using the numbering of the cDNA derived amino acid sequence of human prepro C3. Addition of Fab fragments from the alloselective antibody preferentially inhibited the activity of C3F+ in a haemolytic assay which is selective for the C3 activity in the alternative complement pathway.