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Biomedical subjects

E A Bauer

Publications and source records attributed to E A Bauer.

At least 91 records · Page 5Linked to original sources
PubMed1982

Colchicine-induced modulation of collagenase in human skin fibroblast cultures. II. A probe for defective regulation in epidermolysis bullosa.

The addition of colchicine to cultures of normal human skin fibroblasts produces a significant stimulation of collagenase. Because this finding implies a role for the microtubule system in the regulation of normal collagenase synthesis, we have used colchicine as a probe for aberrations in this enzyme in epidermolysis bullosa. In fibroblast cultures from the dominant simplex, dominant dystrophic, and recessive letalis forms of epidermolysis bullosa, 10(-6) M colchicine produced approximately a 2-fold increase in collagenase in the culture medium, a finding shown by biosynthetic studies to be attributable to enhanced synthesis of enzyme protein. In the case of typical recessive dystrophic epidermolysis bullosa, a disease characterized by excessive collagenase synthesis, the fibroblasts could also be stimulated to produce additional collagenase, despite having elevated baseline synthetic rates. In contrast, fibroblasts isolated from one recessive epidermolysis bullosa patient were resistant to the stimulatory effects of colchicine in concentrations up to 5 x 10(-6) M. In the absence of colchicine, collagenase synthesis in this patient's cells (termed REBc-) was 3-4 times that of normal controls, suggesting that the as yet undefined cellular function that is abrogated (or stimulated) by colchicine in normal cells may have been genetically impaired in these REBc- cells. Despite the resistance to colchicine, as manifested by the failure to stimulate collagenase, gross parameters of microtubular function, such as cell replication, were intact. Phenotypically, this patient had a form of epidermolysis bullosa intermediate between typical recessive dystrophic and recessive letalis forms of the disease. Although an experimentally induced blister was located in the lamina lucida, hypoplastic anchoring fibrils were also observed. These findings, in addition to the marked increase in collagenase synthesis, suggest the possibility that this patient may represent a compound heterozygote of two forms of epidermolysis bullosa and that colchicine may be useful in defining other such patients.

Cells, Cultured↗
PubMed1982

Abnormalities in collagenase expression as in vitro markers for recessive dystrophic epidermolysis bullosa.

In order to correlate a characteristic clinical phenotype with biochemical abnormalities in recessive dystrophic epidermolysis bullosa, fibroblast cultures were established from 4 typical patients with the severe form of the disease. Collagenase, the enzyme implicated in the pathogenesis of blistering, was present in vitro in 2- to 4-fold greater concentrations than in control fibroblast cultures. Partially purified preparations of this enzyme displayed marked thermal lability and diminished affinity for Ca2+, a metal cofactor, suggesting the existence of a mutant enzyme. The data suggest that these 3 biochemical abnormalities, increased synthesis, decreased thermal stability and diminished affinity for Ca2+, should serve as reliable in vitro markers for genetic discrimination of recessive dystrophic epidermolysis bullosa.

Cells, Cultured↗
PubMed1982

Upper airway and external genital involvement in epidermolysis bullosa dystrophica.

Radiographic findings in two patients with uncommon manifestations of epidermolysis bullosa dystrophica are described. A girl with recurrent urinary tract infections had scarring of the external genitalia producing chronic vaginal and uterine reflux and retention of urine. A second patient with long-standing stridor was found to have subglottic narrowing due to localized subglottic edema associated with an inflammatory membrane. The second case illustrates the occasional involvement of columnar epithelial surfaces which can occur in both epidermolysis bullosa dystrophica and epidermolysis bullosa hereditaria letalis.

Airway Obstruction↗
PubMed1982

Human skin collagenase in recessive dystrophic epidermolysis bullosa. Purification of a mutant enzyme from fibroblast cultures.

Recessive dystrophic epidermolysis bullosa, a genodermatosis characterized by dermolytic blister formation in response to minor trauma, is characterized by an incresaed collagenase synthesis by skin fibroblasts in culture. Since preliminary studies of partially purified recessive dystrophic epidermolysis bullosa collagenase suggested that the protein itself was aberrant, efforts were made to purify this enzyme to homogeneity, so that detailed biochemical and immunologic comparisons could be made with normal human skin fibroblast collagenase. Recessive dystrophic epidermolysis bullosa skin fibroblasts obtained from a patient documented to have increased synthesis of the enzyme were grown in large scale tissue culture and both serum-free and serum-containing medium collected as a source of collagenase. The recessive dystrophic epidermolysis bullosa collagenase was purified to electrophoretic homogeneity using a combination of salt precipitation, ion-exchange, and gel-filtration chromatography. In contrast to the normal enzyme, the recessive dystrophic epidermolysis bullosa collagenase bound to carboxymethyl-cellulose at Ca(2+) concentrations at least 10 times higher than those used with the normal enzyme. Additionally, this enzyme was significantly more labile to chromatographic manipulations, particularly when serum-free medium was used. However, rapid purification from serum-containing medium yielded a preparation enzymatically equivalent to normal human skin collagenase. Like the normal enzyme, the recessive dystrophic epidermolysis bullosa collagenase was secreted as a set of two closely related zymogens of approximately 60,000 and approximately 55,000 daltons that could be activated by trypsin to form enzymically active species of approximately 50,000 and approximately 45,000 daltons, respectively. Amino acid analysis suggested slight variations between the normal and recessive dystrophic epidermolysis bullosa collagenases. Cyanogen bromide digests demonstrated peptides unique to the enzyme from each source. The recessive dystrophic epidermolysis bullosa proenzyme was significantly more thermolabile at 60 degrees C than the normal, a finding that correlated with an approximate fourfold decrease in the affinity of the mutant enzyme for Ca(2+), a known activator and stabilizer of human skin collagenase. Aside from the altered affinity for this metal cofactor, kinetic analysis of the structurally altered recessive dystrophic epidermolysis bullosa collagenase revealed that its reaction rates and substrate specificity for human collagen types I-V were identical to those for the normal enzyme. Likewise, enzymes from both sources displayed identical energies of activation and deuterium isotope effects. Antisera were raised to the normal and putatively mutant procollagenases respectively, and, although they displayed a reaction of identity in double diffusion analysis, immunologic differences were present in enzyme inhibition and quantitative precipitation studies. These studies indicate that recessive dystrophic epidermolysis bullosa is characterized by the increased synthesis of an enzymically normal, but structurally aberrant, collagenase.

Amino Acids↗
PubMed1981

Human skin fibroblasts in culture: procollagen synthesis in the presence of sera from normal human subjects and from patients with dermal fibroses.

Various dermal fibrotic conditions, such as progressive systemic sclerosis, localized morphea and familial cutaneous collagenoma, are characterized by excessive deposition of collagen in the skin. In the present study, we examined the possibility that a circulating serum factor(s) is responsible for increased collagen production in these diseases. The effects of human serum on the synthesis of procollagen were examined by incubating normal human dermal fibroblasts with [3H]proline and varying concentrations of dialyzed heat-inactivated serum. The synthesis of procollagen was measured as formation of nondialyzable [3H]hydroxyproline and collagenase-digestible 3H]polypeptides. In the absence of serum little procollagen was formed but the synthesis was markedly stimulated by the addition of normal serum in a concentration-dependent manner. THe ratio of genetically distinct 3H-procollagens of type I and type III, assayed by DEAE-cellulose chromatography and SDS-polyacrylamide gel electrophoresis after limited pepsin proteolysis, was unaffected by the addition of serum. Thus, normal human serum contains a nondialyzable factor(s) which stimulates the synthesis of procollagens type I and type III equally. Sera from 5 patients with progressive systemic sclerosis, 3 with localized scleroderma, and 2 with familial cutaneous collagenoma were also tested. Sera from these patients failed to stimulate 3H-procollagen production more than sera from healthy age-matched controls. Therefore, no increased quantities or qualitatively aberrant factors were shown to be present in the sera of these patients.

Blood Physiological Phenomena↗
PubMed1981

Lipoid proteinosis: in vivo and in vitro evidence for a lysosomal storage disease.

Tissue and cultured fibroblasts derived from one patient with the classical findings of lipoid proteinosis have been used to examine pathologic mechanisms in the disease. Ultrastructural examination of the skin revealed not only extracellular deposits of finely granular, moderately electron dense material, but in addition the dermal fibroblasts characteristically demonstrated marked cytoplasmic vacuolization. Phase contrast microscopy of the cultured skin fibroblasts also showed strikingly abnormal cells with many inclusions, which by electron microscopy were delimited by a single membrane. Membranous lamellar material was also increased in these cells. Biochemical analysis of the fibroblasts revealed a 3- to 4-fold elevation in intracellular hexuronic acid. These morphologic and biochemical findings suggest certain similarities with known storage diseases and support the postulate that lipoid proteinosis may represent a lysosomal storage disease.

Cells, Cultured↗
PubMed1981

Epidermolysis bullosa.

The disorders known as epidermolysis bullosa constitute a group of genetic diseases of the skin which in the mildest form are debilitating throughout life and in the most severe forms result in death at an early age. The profound morbidity associated with all forms of the disease results not only in the loss of an economically productive life but also, in many cases, in the need for continuing medical care throughout life. Inroads are being made into potential pathogenic mechanisms in certain forms of EB. The current state of knowledge can best be characterized as a number of potentially significant disparate observations from various disciplines. The ultimate goals of research in EB are not only to understand the disease but also to provide the biochemical tools for reliable genetic classification, for antenatal diagnosis, and for rational modes of therapy.

Epidermolysis Bullosa↗
PubMed1980

Phenytoin therapy of recessive dystrophic epidermolysis bullosa. Clinical trial and proposed mechanism of action on collagenase.

We administered phenytoin (diphenylhydantoin) by mouth to 17 unselected patients to assess its ability to reduce blistering in recessive dystrophic epidermolysis bullosa (RDEB). Therapeutic response was correlated with blood levels of the drug. Although there was a decrease in blistering of 53 +/- 6 per cent (mean +/- S.E.) among all patients at levels of more than 8 microgram of phenytoin per milliliter, the response was variable, with 12 of 17 patients having a decrease in blistering of more than 40 per cent. Since increased collagenase in human skin has been implicated in the pathogenesis of blistering in RDEB, we examined the effect of phenytoin on this enzyme. Although the drug did not inhibit collagenase activity directly, its addition to human-skin explant and fibroblast cultures produced a 50 to 60 per cent decrease in collagenase activity and immunoreactive protein concentrations. These in vitro studies suggest that phenytoin inhibits synthesis or secretion of collagenase of both, and that the favorable clinical response can be explained by this inhibition.

Administration, Oral↗
PubMed1980

Hormonal interactions in mammalian collagenase regulation. Comparative studies in human skin and rat uterus.

The production of collagenase by human skin explants in culture is prevented by 10(-8) M dexamethasone, 5 . 10(-4) M dibutyryl cyclic AMP, or 2.5 . 10(-3) M theophylline. Decreases in collagenase activity are paralleled by reductions in the degradation of explant collagen during the culture period. Progesterone, which effectively inhibits collagenase production in rat uterine explant cultures, has no effect on human skin explants. The inhibition by cyclic AMP is nucleotide specific. When partially inhibitory concentrations of dexamethasone and dibutyryl cyclic AMP, or dexamethasone and theophylline, are added to culture medium together, the resultant inhibition is that predicted by additivity. Synergistic inhibition, as observed in rat uterus between progesterone and dibutyryl cyclic AMP, fails to occur. Dexamethasone inhibits the production of collagenase by cultured explants of rat uterus, with complete inhibition occurring at 10(-7) M steroid. Synergism between glucocorticoids and dibutyryl cyclic AMP or between dexamethasone and progesterone could not be demonstrated in the uterine culture system. These results suggest the existence of three regulatory systems for the control of collagenase production in mammalian tissues, and that cooperativity between systems may occur on a tissue-specific basis.

Animals↗
PubMed1980

Collagenase in human cornea: immunologic localization.

Collagenase has been localized in human corneas using immunocytochemical techniques employing a specific antiserum to human skin collagenase. The enzyme was found in vivo in ulcerating corneas, whereas in corneas obtained from a variety of nonulcerative conditions, no immunologic evidence of the enzyme was seen. Corneal collagenase was present only in the stromal portion of the cornea, suggesting that cells from this tissue are the source of the enzyme. The localization of collagenase to the corneal stroma was reproduced by placing either ulcerating or nonulcerating corneas in tissue culture, indicating that the organ culture system may represent a model that duplicates some conditions present in corneal disease.

Adult↗
PubMed1980

Morphea and lichen sclerosus et atrophicus. Clinical and histopathologic studies in patients with combined features.

Ten patients with skin lesions clinically consistent with morphea and lichen sclerosus et atrophicus were studied. In all cases, histopathologic changes of both morphea and lichen sclerosus et atrophicus were also present. The coexistence of morphea and lichen sclerosus et atrophicus in the same patient suggests that these lesions represent a spectrum which may reflect similar etiologic events or closely related pathologic processes in these two diseases.

Adolescent↗
PubMed1980

Enhanced biosynthesis of human skin collagenase in fibroblast cultures from recessive dystrophic epidermolysis bullosa.

Using a sensitive, specific immunoprecipitation method, the biosynthesis of human skin collagenase was studied in fibroblast cultures from patients with recessive dystrophic epidermolysis bullosa. Sodium dodecyl sulfate polyacrylamide gel electrophoresis of solubilized immunoprecipitates showed two 3H-labeled procollagenase species that comigrated with those harvested from control cultures. Recessive dystrophic epidermolysis bullosa cultures accumulated increased amounts of collagenase. Both the initial rate of accumulation of intracellular enzyme and the rate of secretion were enhanced, suggesting that excessive accumulation is related to increased synthesis. Because the turnover of labeled collagenase was unaltered, the accumulation could not be attributed to diminishing enzyme degradation. No preferential incorporation of [3H]leucine into recessive dystrophic epidermolysis bullosa collagenase occurred. Furthermore, the mutant cultures displayed no alteration in total protein synthesis, the intracellular leucine pool, or the growth kinetics of the cells. Cells from a patient with dominant epidermolysis bullosa did not show enhanced accumulation of collagenase. The levels of collagenase synthesized in vitro correlated with those observed previously in vivo in recessive dystrophic epidermolysis bullosa patients, suggesting that this biochemical trait is pathogenetically significant in the disorder.

Cells, Cultured↗
PubMed1979

Environmental pH modulation of collagenase in normal human fibroblast cultures.

Normal human skin fibroblast cultures have been used to assess the effects of relatively minor changes in environmental pH on collagenase, a major extracellular gene product. Collagenase accumulation in the culture medium, measured both as enzyme activity and immunoreactive material, was 2- to 10-fold greater at pH 7.6--8.2 than at pH 6.8--7.2. The pH-associated increase in collagenase was paralleled by an increase in general protein synthesis. Nevertheless, prototypic lysosomal and cytoplasmic enzymes changed very little under identical culturing conditions. Although substantial intracellular protein degradation occurred at all pH values, the small differences either in general protein degradation or in specific collagenase degradation in the medium were of insufficient magnitude to account for the increased accumulation of collagenase.

Cells, Cultured↗
PubMed1979

Carcinoembryonic antigen: increased plasma levels in recessive epidermolysis bullosa.

Circulating plasma levels of the oncofetal antigen, carcinoembryonic antigen, were examined in 18 patients with various forms of epidermolysis bullosa. Carcinoembryonic antigen was markedly elevated in the plasma of 4 of 6 patients with recessive dystrophic epidermolysis bullosa and in 1 of 2 patients with recessive epidermolysis bullosa letalis. In contrast, patients with dominantly inherited forms of the disease, dominant epidermolysis bullosa simplex and dominant dystrophic epidermolysis bullosa, had normal levels of antigen. In the recessive patients, the plasma levels of carcinoembryonic antigen appeared to correlate with the severity of cutaneous involvement. Alternatively, it is possible that expression of carcinoembryonic antigen is genetically linked to certain forms of recessive epidermolysis bullosa or is part of a pleiotropic effect of the gene coding for the disease.

Adolescent↗