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Biomedical subjects

K D Wuepper

Publications and source records attributed to K D Wuepper.

At least 37 records · Page 2Linked to original sources
PubMed1981

Purification and characterization of streptococcal proliferative factor.

Group A streptococcal infections are often associated with scarlet fever and flares of guttate psoriasis. Previous investigation has demonstrated the presence of a factor in streptococcal culture filtrates capable of stimulating proliferation of rabbit keratinocytes in vivo and human lymphocytes in vitro. This report outlines an in vivo method for the production of streptococcal proliferation factor, its purification, and characterization of its physical properties. We cultured Group A streptococci (Type 12, Strain NY5) in synthetic media by in vivo incubation within dialysis casing surgically implanted in rabbit peritoneum. Streptococcal exoproteins were isolated by centrifugation of the bacteria and millipore filtration. Purification of streptococcal proliferative factor was accomplished by differential solubility and molecular sieve was discovered in the resulting product. The relative by SDS gels and molecular sieve chromatography. The sedimentation coefficient determined by sucrose gradient ultracentrifugation is 2.7S. Isoelectric focusing showed minimal microheterogeneity with the pI of the major band being 5.0. Thus, streptococcal proliferative factor can be produced by in vivo incubation of streptococci in synthetic media. Purification entails a rapid 2-step process. The relative molecular weight, sedimentation coefficient and isoelectric points have been established.

Animals↗
PubMed1980

Epidermal transglutaminase. Identification and purification of a soluble substrate with studies of in vitro cross-linking.

A Mr = 36,000 substrate for epidermal transglutaminase was identified immunochemically in buffer extracts of bovine snout epidermis, using antiserum to isolated high molecular weight substrate proteins recovered after cross-linking by transglutaminase. The high molecular weight proteins were not present in soluble epidermal extracts in EDTA prior to cross-linking. The substrate was purified by DEAE-Sepharose CL-6B and ACA-34 gel filtration, where it demonstrated an apparent molecular weight of 36,000. In the presence of Ca2+ and transglutaminase, the purified protein was converted to high molecular weight polymers which, under conditions of high protein concentration, included a protein aggregate insoluble in urea, sodium dodecyl sulfate, or beta-mercaptoethanol. Cross-linking did not occur with Ca2+ alone or in the presence of EDTA or putrescine, a competitive inhibitor. The epsilon-(gamma-glutamyl) lysine isodipeptide was identified in high molecular weight products of cross-linking but not in the Mr = 36,000 precursor. We postulate the intracellular assembly of Mr = 36,000 substrate, with subsequent cross-linking by transglutaminase and insolubilization into the keratinocyte membrane.

Animals↗
PubMed1980

Immune complexes in erythema multiforme and the Stevens-Johnson syndrome.

Early cutaneous lesions of erythema multiforme or mucosal lesions of the Stevens-Johnson syndrome contain delicate granular deposits of immune reactants and/or complement components lodged in the walls of vessels of the papillary dermis. Such deposits are not present in normal, unaffected skin although they can be caused to occur there by injection of substances which increase vascular permeability. Factors which cause accumulation of mononuclear cells in the papillary dermis have not been elucidated but we suggest that receptors for the C3d fragment of the third component of complement could be responsible. Factors which cause intercellular edema in the epidermis, epidermal-dermal separation, or necrotic keratinocytes are unknown. We, and others, may now measured immune complexes in the serum of patients experiencing these reaction patterns by means of Raji cell radioimmunoassay, C1q precipitation, 125I-C1q binding, cryoprecipitation and with monoclonal rheumatoid factor. Cryoimmunoglobulins have also been described and antigens of Herpesvirus hominus have been measured in the cryoprecipitate of 2 patients who experienced erythema multiforme following recurrent herpesvirus infection. Erythema multiforme commonly accompanied serum sickness reactions which occurred following injection of immune horse serum in the 1980s (von Pirquet and Schick). Such reactions are now well established as due to circulating immune complexes in antigen excess.

Antigen-Antibody Complex↗
PubMed1979

Purification of a mannan from Candida albicans which activates serum complement.

Candida species activate complement by the alternative pathway, induce leukocyte migration and, when applied epicutaneously, cause epidermal microabscesses of neutrophils in man and experimental animals. Complement activation by C. albicans appears to be a property of the cell wall. To biochemically identify the complement-activating constituent(s) of C. albicans, an ethyleneglycol extract of growth phase blastospores was prepared. Acid hydrolysis and neutral sugar analysis revealed mannose (82%), fucose (7%), and glucose (11%). The soluble, mannose-rich cell wall polysaccharide of C. albicans activates serum complement via the alternative pathway, induces neutrophil chemotaxis and is antigenically reactive with antisera to C. albicans. This constituent exhibits in vitro endotoxin-like activity as measured by Limulus lysate gelation, but is nonpyrogenic in rabbits. The extracts produced precipitin lines in double immunodiffusion studies against serum from patients with invasive candidiasis and rabbit antisera to mycelial and blastospore preparations of C. albicans, but not against normal serum. Thus, pathogenic properties and reactive phenomenon of C. albicans are in part attributable to a cell wall polysaccharide, mannan.

Animals↗
PubMed1978

Experimental cutaneous candidiasis in rodents; II. Role of the stratum corneum barrier and serum complement as a mediator of a protective infalmmatory response.

Of six species of Candida applied epicutaneously to rodents, only C albicans and C stellatoidea penetrate the stratum corneum and produce inflammation. The role of the stratum corneum and produce inflammation. The role of the stratum corneum in experimental murine Candida infections was studied, therefore, by intraepidermal injection of blastospores in mice that received a prior injection of staphylococcal epidermolysin. All six species caused accumulation of neutrophils if placed within an intraepidermal cleft in contact with viable epidermis. The role of serum complement in cutaneous candidiasis was also studied in vivo in rodents depleted of complement with cobra venom factor or deficient in the fifth component of complement. These animals failed to develop neutrophilic inflammatory responses to the six Candida species tested in contrast to control animals. Candida albicans, C stellatoidea, and C tropicalis developed extensive hyphal proliferation with invasion of the entire cutis and subcutis in these animals. The stratum corneum provides an effective barrier to some, but not all Candida species. When this barrier is penetrated, complement mediates an acute neutrophilic pustular response that resticts Candida proliferation and prevents deep invasion of tissue.

Animals↗
PubMed1978

The epidermolytic toxin of Staphylococcus aureus: its failure to bind to cells and its detection in blister fluids of patients with bullous impetigo.

Radioiodinated staphylococcal epidermolytic toxin was found not to bind to erythrocytes, blood leukocytes, trypsin-dispersed keratinocytes, epidermis or whole skin. Moreover the toxin could not be found to bind to murine epithelia by indirect immunofluorescence methods. However, the toxin, measured by radioimmunoassay, could be eluted from the skin of mice undergoing epidermolysis following intraperitoneal injection of toxinogenic Staphylococcus aureus. Furthermore, epidemolysin was measured in the blister fluid of 3 of 5 children with bullous impetigo but not in blister fluid from control patients with other blistering eruptions. Thus epidermolysin has been demonstrated to be present in lesions of the staphylococcal epidermolytic toxin syndrome but its mechanism of action does not involve binding to cells.

Animals↗
PubMed1977

[Staphylogenous Lyell's syndrome].

The scalded skin syndrome or Lyell syndrome can now be divided into two distinct forms. One form is associated with staphylococci that are usually phage group II Staphylococcus aureus, are penicillin resistant, and produce a protein exotoxin which produces epidermolysis by causing a split to develop in the epidermis between the stratum granulosum and the stratum spinosum. The other form is associated usually with drug reactions and is characterized by necrosis of the epidermis with a split between epidermis and dermis. The staphylococcal Lyell syndrome should be treated with penicillinase resistant antibiotics and not with corticosteroids; the non-staphylococcal form of the Lyell syndrome often requires systemic corticosteroids. Therefore the correct diagnosis must be established early. This is possible since the histopathology of the two forms is different.

Adolescent↗
PubMed1976

Isolation, purification and characterization of bovine epidermal transglutaminase.

A crosslinking enzyme, epidermal transglutaminase, was isolated from soluble proteins of glabrous cow snout epidermis. This enzyme stabilized fibrin clots rendering them insoluble in 2% acetic acid. It also catalyzed the incorporation of the fluorescent amine, dansyl cadaverine, into casein. Epidermal transglutaminase was purified by chromatography upon DEAE-Sephadex A-50, zone electrophoresis in Pevikon, and Sephadex G-200 gel permeation chromatography. The highly purified substance, which had a specific activity of 3267 amine-incorporating units/mg per h and a molecular weight of 55000, behaved as a single molecular species in the analytical ultracentrifuge. It had a sedimentation coefficient of 4.4 S and migrated as a gamma-globulin at pH 8.6; it displayed anomalous migration in polyacrylamide gels containing sodium dodecyl sulfate. The enzyme was dependent upon free calcium ions and a reduced sulfhydryl group for activity. The apparent Km for dansyl cadaverine was 1.2 - 10(-4) at pH 7.5. Monospecific antiserum to bovine epidermal transglutaminase precipitated with the enzyme in agar. The antiserum prevented fibrin crosslinking but enhanced incorporation of dansyl cadaverine into casein by the enzyme. The epidermal enzyme differed biochemically and immunochemically from bovine plasma transglutaminase (Factor XIII).

Animals↗
PubMed1976

Clinical and physiologic studies of two siblings with prekallikrein (Fletcher factor) deficiency.

Two siblings with hereditary Fletcher factor (prekallikrein) deficiency were studied for alterations of fibrinolysis, platelet function, skin inflammatory responses, permeability factor (PF/dil) formation and leukocyte chemotaxis. In vivo stimulation of fibrinolytic activity was normal; the bleeding time and platelet functions (adhesivity, aggregation, release reaction) were also normal. Both immediate (wheal-flare reaction to histamine, bradykinin, prostaglandin E1, physical agents) and delayed sensitivity skin test reactions were within normal limits. Migration of subjects' leukocytes to attractants in skin windows and in Boyden-type chambers was the same as that of control leukocytes. Serum complement components were essentially normal. One subject's leukocytes showed normal tissue factor production on stimulation by endotoxin, although prekallikrein deficiency did impair the endotoxin-stimulated generation of serum procoagulant activity. PF/dil caused increased vessel permeability in human skin; in vitro generation of PF/dil required both the Hageman factor and prekallikrein. The Fletcher factor-deficient subjects responded in a normal manner to PF/dil. Based on the Fletcher factor-coagulation assay, the biologic half-disappearance time of prekallikrein (after the transfusion of normal plasma in one of the subjects) was estimated at 35 hours. Therefore, these studies suggest that severe prekallikrein (Fletcher factor) deficiency in man is not associated with any clinically significant impairment in hemostasis, fibrinolysis, inflammatory responses or leukocyte function.

Adolescent↗
PubMed1976

Experimental cutaneous candidiasis in rodents.

Epicutaneous application of viable Candida albicans or Candida stellatoidea under an occlusive dressing resulted in subcorneal microabscesses in newborn rats and mice. Neither heat-killed C. albicans nor occlusion alone resulted in epidermal micropustules. These results were dependent upon the number of organisms applied and the duration of application. Four additional species of Candida tested did not produce epidermal micropustules. The microscopic pathology of the abscesses revealed penetration of the epidermis by Candida pseudohyphae; the tips of the hyphal processes were seen within the pustules when sections were stained by the periodic acid-Schiff method. The resident flora of the newborn mouse or rat was not substantially altered by occlusion with or without C. albicans during the interval tested. The animal model of cutaneous candidiasis with permit greater manipulation and control in investigations of cutaneous candidiasis and the factors influencing neutrophil accumulation in the epidermis.

Animals↗
PubMed1976

Activation of the alternative (properdin) pathway of complement by Candida albicans and related species.

Accumulations of neutrophilic granulocytes within the epidermis and beneath the stratum corneum of the skin are a prominent histologic feature of experimental and clinical candidiasis. The mechanism of cell accumulation was studied by standard chemotactic methods. Suspensions of viable or heat-killed Candida sp caused marked chemotaxis of human neutrophils in fresh serum. Culture supernatants of Candida sp were not chemotactic. Chemotaxis was dependent upon fresh serum, and could be abolished by heating the serum to 56 degress C for 30 min, suggesting that interaction of these organisms with a heat-labile serum factor generated a chemoattractant. Incubation of Candida sp with fresh human serum resulted in the conversion of the third component of complement and properdin factor B, as measured by immunoelectrophoresis. Conversion did not occur in serum chelated with EDTA, or heated to 50 degress C for 30 min (to destroy factor B). Conversion was present in serum chelated with EGTA (to deplete calcium), or genetically deficient in the fourth component of complement. By contrast, the three components of the kinin-forming system (Hageman factor, prekallikrein, high-molecular-weight kininogen) were not activated by Candida sp. We suggest that Candida sp do not release a chemotactic substance but, in the presence of serum, activate the alternative pathway of complement, generating chemotactic factors.

Candida↗