Familial scleroderma-like deformity of the fingers.
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Adolescent↗
Biomedical subjects
J C Olson
Publications and source records attributed to J C Olson.
Systemic lupus erythematosus presenting with a painful goiter.
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Acute Disease↗
Childhood stroke and lupus anticoagulant.
Despite extensive evaluation, the cause of many childhood ischemic strokes remains unexplained. Two children are reported with stroke and lupus anticoagulant. Both had family members with features of the antiphospholipid antibody syndrome. Children with unexplained ischemic stroke should be evaluated for lupus anticoagulant and anticardiolipin antibodies.
Antiphospholipid Syndrome↗
Cutaneous polyarteritis nodosa of childhood.
BACKGROUND: The clinical presentation of childhood polyarteritis nodosa (PAN) can range from isolated cutaneous findings to widespread multisystem involvement. Both the systemic and cutaneous forms are known to occur after streptococcal infection. OBJECTIVE: Our purpose was to emphasize the frequent association of childhood cutaneous PAN with antecedent streptococcal infection. METHODS: We discuss four cases of cutaneous PAN that were associated with streptococcal infection and briefly review the pertinent literature. RESULTS: All patients had evidence of preceding streptococcal infection at the onset of their illness. Although mild systemic symptoms were present in all, their course was benign. The use of nonsteroidal antiinflammatory agents or corticosteroids resulted in clinical improvement. Antibiotic therapy was also used in the treatment of these patients. CONCLUSION: In children with PAN, evaluation should include laboratory studies to detect streptococcal infection.
Adolescent↗
Flow cytometric immunodetection of human immunodeficiency virus type 1 proviral DNA by heminested PCR and digoxigenin-labeled probes.
PCR is the most sensitive and direct method for detecting blood-borne viruses, as well as an efficient means for producing vector-free probes. However, the application of PCR, especially in the laboratory diagnosis of human immunodeficiency virus (HIV) infection, is impeded by the current use of radiolabeled oligonucleotide probes. Therefore, we have developed a nonisotopic PCR immunoreactive bead (PCR-IRB) assay to detect HIV type 1 proviral DNA from peripheral blood mononuclear cells (PBMC). We used a biotinylated primer in a set of three oligonucleotides selected from the HIV long terminal repeat region for heminested PCR amplification. An internal probe was synthesized by PCR with incorporation of digoxigenin-labeled dUTP. After solution hybridization of the probe with PCR-amplified products (amplicons), the hybridized DNA was captured with streptavidin-coated magnetic beads. For the detection of hybrids, flow cytometric analyses were carried out by two procedures: (i) direct detection with fluorescein isothiocyanate (FITC)-labeled antidigoxigenin immunoglobulin G (IgG) antibody and (ii) indirect detection with antidigoxigenin sheep IgG antibody followed by FITC-labeled anti-sheep IgG antibody. Both procedures in the PCR-IRB assay detected two to three copies of HIV proviral DNA sequences, a sensitivity that is comparable with that of the conventional radioactive detection of amplicons following probe hybridization and electrophoresis. To compare the PCR-IRB assay with the conventional method, we tested 53 pedigreed PBMC specimens from blood donors and newborns; the results obtained were identical. This nonisotopic PCR-IRB assay can also be automated for potential application in laboratory diagnosis of HIV infection, blood bank screening, and therapeutic monitoring of viremia and perinatal transmission.
Base Sequence↗
Secreted LasA of Pseudomonas aeruginosa is a staphylolytic protease.
Full expression of the elastolytic phenotype of Pseudomonas aeruginosa depends on LasA, an extracellular protease with restricted specificity whose mode of action on elastin and biological role is not understood. LasA exhibits amino acid sequence homology to some bacteriolytic proteases and shares several physicochemical properties with the staphylolytic protease of P. aeruginosa. This led us to examine whether the two proteases are the same. Production of LasA and staphylolytic protease by prototrophic and lasA mutant strains of P. aeruginosa was investigated. The two prototrophic strains examined, PAO1 and FRD2, exhibited extracellular staphylolytic activity and secreted LasA. LasA mutants, PAO-E64 lasA1 (Ts), FRD2128 delta lasA, and FRD244 lasA::mTn10, did not exhibit staphylolytic activity. A low level of the LasA protein was detected in the culture filtrate of the temperature-sensitive lasA mutant PAO-E64, but none was detectable in those of the deletion and insertion mutants, FRD2128, and FRD244, respectively. The staphylolytic protease was purified from the culture filtrate of P. aeruginosa strain FRD2 by DEAE-cellulose chromatography. The purified enzyme hydrolyzed pentaglycine into the respective di- and tripeptides and reacted specifically with antibodies against a synthetic peptide identical in sequence to positions 77-98 in LasA. The amino-terminal sequence of the first 15 amino acid residues of the staphylolytic protease was found to be identical with that of the secreted LasA. These results clearly indicate that LasA is a staphylolytic protease. In addition to lysing staphylococci, it may enhance elastolysis by cleaving Gly-Gly bonds, which are abundant in elastin.
Amino Acid Sequence↗
Cholinergic stimulation of the Na+/K+ adenosine triphosphatase as revealed by microphysiometry.
The activation of a wide range of cellular receptors has been detected previously using a novel instrument, the microphysiometer. In this study microphysiometry was used to monitor the basal and cholinergic-stimulated activity of the Na+/K+ adenosine triphosphatase (ATPase) (the Na+/K+ pump) in the human rhabdomyosarcoma cell line TE671. Manipulations of Na+/K+ ATPase activity with ouabain or removal of extracellular K+ revealed that this ion pump was responsible for 8.8 +/- 0.7% of the total cellular energy utilization by those cells as monitored by the production of acid metabolites. Activation of the pump after a period of inhibition transiently increased the acidification rate above baseline, corresponding to increases in intracellular [Na+] ([Na+]i) occurring while the pump was off. The amplitude of this transient was a function of the total [Na+]i excursion in the absence of pump activity, which in turn depended on the duration of pump inhibition and the Na+ influx rate. Manipulations of the mode of energy metabolism in these cells by changes of the carbon substrate and use of metabolic inhibitors revealed that, unlike some other cells studied, the Na+/K+ ATPase in TE671 cells does not depend on any one mode of metabolism for its adenosine triphosphate source. Stimulation of cholinergic receptors in these cells with carbachol activated the Na+/K+ ATPase via an increase in [Na+]i rather than a direct activation of the ATPase.
Biophysical Phenomena↗
Pseudomonas aeruginosa lasB1 mutants produce an elastase, substituted at active-site His-223, that is defective in activity, processing, and secretion.
Pseudomonas aeruginosa secretes elastase in a multistep process which begins with the synthesis of a preproelastase (53.6 kDa) encoded by lasB, is followed by processing to proelastase (51 kDa), and concludes with the rapid accumulation of mature elastase (33 kDa) in the extracellular environment. In this study, mutants of P. aeruginosa were constructed by gene replacement which expressed lasB1, an allele altered in vitro at an active-site His-223-encoding codon. The lasB1 allele was exchanged for chromosomal lasB sequences in two strain backgrounds, FRD2 and PAO1, through a selectable-cassette strategy which placed a downstream Tn501 marker next to lasB1 and provided the selection for homologous recombination with the chromosome. Two lasB1 mutants, FRD720 and PDO220, were characterized, and their culture supernatants contained greatly reduced proteolytic (9-fold) and elastolytic (14- to 20-fold) activities compared with their respective parental lasB+ strains. This was primarily due to the effect of His-223 substitution on substrate binding by elastase and thus its proteolytic activity. However, the concentration of supernatant elastase antigen was also reduced (five- to sevenfold) in the mutant strains compared with the parental strains. An immunoblot analysis of cell extracts showed a large accumulation of 51-kDa proelastase within lasB1 mutant cells which was not seen in wild-type cell extracts. A time course study showed that production of extracellular elastase was inefficient in the lasB1 mutants compared with that of parental strains. This showed that expression of an enzymatically defective elastase inhibits proper processing of proelastase and provides further evidence for autoproteolytic processing of proelastase in P. aeruginosa. Unlike the parental strains, culture supernatants of the lasB1 mutants contained two prominent elastase species that were 33 and 36 kDa in size. Extracellular 51-kDa proelastase was barely detectable, even though it accumulated to high concentrations within the lasB1 mutant cells. These data suggest that production of an enzymatically defective elastase affects proper secretion because autoproteolytic processing of proelastase is necessary for efficient localization to the extracellular milieu. The appearance of reduced amounts of extracellular elastase and their sizes of 33 and 36 kDa suggest that lasB1-encoded elastase was processed by alternate, less-efficient processing mechanisms. Thus, proelastase must be processed by removal of nearly all of the 18-kDa propeptide before elastase is a protein competent for extracellular secretion.
Alleles↗
Use of synthetic peptides and site-specific antibodies to localize a diphtheria toxin sequence associated with ADP-ribosyltransferase activity.
Diphtheria toxin (DT) and Pseudomonas aeruginosa exotoxin A have the same molecular mechanism of toxicity; both toxins ADP-ribosylate a modified histidine residue in elongation factor 2. To help identify amino acids involved in this reaction, sequences in DT that share homology with P. aeruginosa exotoxin A were synthesized and examined for a role in the ADP-ribosyltransferase reaction. By using this approach, residues 32 to 54 of DT were found to define an epitope associated with antibody-mediated inhibition of DT enzyme activity. This lends further support to the notion that residues in this region of DT are involved in the enzymatic reaction.
ADP Ribose Transferases↗
Biochemical characterization of the cystic fibrosis transmembrane conductance regulator in normal and cystic fibrosis epithelial cells.
Affinity-purified polyclonal antibodies, raised against two synthetic peptides corresponding to the R domain and the C terminus of the human cystic fibrosis transmembrane conductance regulator (CFTR), were used to characterize and localize the protein in human epithelial cells. Employing an immunoblotting technique that ensures efficient detection of large hydrophobic proteins, both antibodies recognized and approximately 180-kDa protein in cell lysates and isolated membranes of airway epithelial cells from normal and cystic fibrosis (CF) patients and of T84 colon carcinoma cells. Reactivity with the anti-C terminus antibody, but not with the anti-R domain antibody, was eliminated by limited carboxypeptidase Y digestion. When normal CFTR cDNA was overexpressed via a retroviral vector in CF or normal airway epithelial cells or in mouse fibroblasts, the protein produced had an apparent molecular mass of about 180 kDa. The CFTR expressed in insect (Sf9) cells by a baculovirus vector had a molecular mass of about 140 kDa, probably representing a nonglycosylated form. The CFTR in epithelial cells appears to exist in several forms. N-glycosidase treatment of T84 cell membranes reduces the apparent molecular mass of the major CFTR band from 180 kDa to 140 kDa, but a fraction of the T84 cell CFTR could not be deglycosylated, and the CFTR in airway epithelial cell membranes could not be deglycosylated either. Moreover, wheat germ agglutinin absorbs the majority of the CFTR from detergent-solubilized T84 cell membranes but not from airway cell membranes. The CFTR in all epithelial cell types was found to be an integral membrane protein not solubilized by high salt or lithium diiodosalicylate treatment. Sucrose density gradient fractionation of crude membranes prepared from the airway epithelial cells, previously surface-labeled by enzymatic galactosidation, showed a plasma membrane localization for both the normal CFTR and the CFTR carrying the Phe508 deletion (delta F 508). The CFTR in all cases co-localized with the Na+, K(+)-ATPase and the plasma membrane calcium ATPase, while the endoplasmic reticulum calcium ATPase and mitochondrial membrane markers were enriched at higher sucrose densities. Thus, the CFTR appears to be localized in the plasma membrane both in normal and delta F 508 CF epithelial cells.
3T3 Cells↗
Painful digital vesicles and acrocyanosis in a toddler.
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Biopsy↗
Efficient production and processing of elastase and LasA by Pseudomonas aeruginosa require zinc and calcium ions.
The ability of Pseudomonas aeruginosa to degrade elastin, a major component of connective tissue, likely contributes to its pathogenicity and multiplication in human tissues. Two extracellular enzymes are required for P. aeruginosa elastolytic activity: elastase and LasA. Elastase is a zinc metalloprotease, but little is known about the structure of LasA. When grown under metal ion-deficient conditions, P. aeruginosa culture supernatants were found to exhibit a low level of elastolytic activity, which coincided with production of low levels of the 51-kDa proelastase and no detectable LasA. By using this fact to identify factors that promote elastolytic activity, P. aeruginosa PAO1, FRD2, and DG1 were grown in metal ion-deficient medium supplemented with zinc (10(-4) M ZnCl2), calcium (2.5 x 10(-3) M CaCl2), or iron (10(-4) M FeCl3). High levels of proteolytic and elastolytic activity were exhibited by all strains when cultured in the presence of both zinc and calcium, and this was associated with the production of mature 33-kDa elastase and 21-kDa LasA. Supplementing DG1 and PAO1 cultures with zinc alone stimulated the production of 33-kDa elastase, which, because of the calcium-deficient conditions, exhibited low proteolytic and elastolytic activities. Zinc also stimulated the production of a 41-kDa form of LasA in DG1 and PAO1 culture supernatants. Elastase production by FRD2 cultured in the presence of zinc alone differed from that by the other two strains in that supernatants contained 33-kDa elastase, a 21-kDa form of LasA, and exhibited high proteolytic and elastolytic activities. Such strain-associated differences in LasA processing and elastase activity can be explained by differences in metal ion-scavenging mechanisms adapted by the strains. Supplementing cultures with calcium stimulated the production of elastase but had no effect on LasA production. The elastase produced exhibited variable sizes, possibly resulting from aberrant processing reactions, and showed little proteolytic activity. Proteolytic activity could be recovered from 33-kDa elastase produced in the presence of calcium by inclusion of zinc in the enzymatic assay. Although iron was previously found to exert a repressive effect on P. aeruginosa elastolytic activity, iron exerted little effect on elastolytic activity when added to cultures containing both zinc and calcium. These studies support the conclusion that elastase production and processing are promoted by both zinc and calcium. LasA production, in comparison, is stimulated by zinc, with both zinc and calcium facilitating its processing. The association of 41-kDa LasA with a low level of elastolytic activity and of 21-kDa LasA with a high level of activity supports the conclusion that lasA encodes a larger, precursor protein which is processed to an active 21-kDa form during secretion.
Amino Acid Sequence↗
Juvenile dermatomyositis.
Juvenile dermatomyositis is a chronic inflammatory disease of skeletal muscle accompanied by skin involvement. It is a presumed autoimmune disease. Other organs may be involved. Corticosteroids are standard therapy and are associated with decreased mortality. Significant morbidity may occur.
Child↗
Pulmonary disease with hemorrhage in Henoch-Schöenlein purpura.
Henoch-Schönlein purpura is a common vasculitic syndrome of childhood. Pulmonary hemorrhage is a rare complication, which resulted in death in three of four previously reported cases. We report concerning four additional patients with Henoch-Schönlein purpura and pulmonary hemorrhage. Three survived the episode. Aggressive and supportive care may improve survival.
Adolescent↗
Identification of functional epitopes of Pseudomonas aeruginosa exotoxin A using synthetic peptides and subclone products.
The structure-function relationship of P. aeruginosa exotoxin A (ETA) was examined using synthetic peptides and genetically engineered ETA deletion mutants. Antibodies directed against synthetic peptides have allowed the identification of three ETA epitopes, two within domain I and one within the last 33 amino acids of domain III. In addition two distinct neutralizing determinants have been identified by antibodies directed against subclone products. One was associated with the amino-terminal half of ETA, the proposed receptor binding region. The second was associated with the carboxy-terminal half of ETA, a region previously not associated with receptor-binding. The amino-terminal subclone also offers potential as an ETA vaccine, since it produces a stable, non-enzymatically active product, effective in inducing ETA neutralizing antibodies. Data derived from these studies were used in a re-evaluation of structure-function relationships between ETA and diphtheria toxin.
ADP Ribose Transferases↗
Urticarial vasculitis and Lyme disease.
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Child, Preschool↗
Arthropathy of Down syndrome.
A juvenile rheumatoid arthritis-like arthropathy has previously been documented in 12 patients with Down syndrome. An additional 9 patients are described and the literature is reviewed. It is unknown whether these patients have juvenile rheumatoid arthritis or a unique arthropathy in light of the genetic and immunologic abnormalities associated with Down syndrome. Most of the patients had a progressive course with polyarticular disease complicated by subluxations and a long lag time to diagnosis. The purpose of reporting these children is to increase awareness of this association and facilitate more appropriate and timely diagnosis of arthritis in Down syndrome patients.
Arthritis, Rheumatoid↗
Two sisters with telangiectases [corrected], spondyloepiphyseal dysplasia, hypothyroidism, neovascularization and tractional retinal detachments: a new entity?
Two sisters had an unusual constellation of findings, including telangiectases [corrected] involving the face and limbs, unclassifiable spondyloepiphyseal dysplasia, hypothyroidism, and tractional retinal detachment preceded by retinal and iris neovascularization. To our knowledge, this combination of defects has not been previously documented in the literature.
Abnormalities, Multiple↗