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Biomedical subjects

J Uitto

Publications and source records attributed to J Uitto.

At least 469 records · Page 26Linked to original sources
PubMed1984

Association of hemolytic anemia and early-onset pulmonary emphysema in three siblings.

Three of four siblings born to nonconsanguineous parents of Italian origin were affected with severe congenital hemolytic anemia of unknown cause, and early-onset pulmonary emphysema. Two of the three affected siblings died of septic shock after splenectomy, at the ages of 7 and 3 1/2 years, respectively. The remaining affected sibling was shown to have cutis laxa and severe pulmonary emphysema at 15 years of age. Assay of serum components indicated that alpha 1-antitrypsin and alpha 2-macroglobulin levels were normal or slightly elevated. However, there was markedly elevated activity of an elastase-like serum enzyme. The relation of the hemolytic anemia to the pulmonary findings in this family is not clear; pedigree analysis suggests a recessively inherited defect.

Adolescent↗
PubMed1984

Nonenzymatic glucosylation of lysyl and hydroxylysyl residues in type I and type II collagens.

Nonenzymatic glucosylation of type I and type II collagens was examined by incubating collagen substrates with D-glucose in vitro. In one set of experiments, unlabeled collagen was incubated with [14C]-glucose and the incorporation of [14C]-radioactivity into protein was determined by TCA precipitation. The incorporation was dependent on the concentration of glucose and the time of incubation. The glucosylated product was also examined by SDS-polyacrylamide slab gel electrophoresis. The results indicated that both alpha 1(I)- and alpha 2(I)-chains of type I collagen were glucosylated and the glucosylation occurred both with native and denatured collagen as substrate. In further studies [3H]-lysine-labeled collagens were glucosylated, the products reduced by NaBH4, and the [3H]-lysine-derived residues were separated by amino acid analyzer. After a 144 h incubation in vitro, 18.9% of [3H]-lysyl residues and 36.5% of [3H]-hydroxylysyl residues in type I collagen were substituted with glucose. In contrast, 47.9% of [3H]-lysyl residues and 68.1% of [3H]-hydroxylysyl residues in type II collagen were glucosylated after 144 h incubation. Based on quantitative amino acid analyses of the substrates, these values represent 27.6 lysine plus hydroxylysine residues substituted per triple-helical type I collagen molecule and 65.3 residues per triple-helical type II collagen molecule. Thus, type I and type II collagens display differential susceptibilities to nonenzymatic glucosylation. Finally, [3H]-proline-labeled type I collagen was glucosylated to varying extents, and the glucosylated products were used as substrates for human polymorphonuclear leukocyte collagenase. No difference in susceptibility to this collagenase was noted, irrespective of the extent of glucosylation.

Amino Acids↗
PubMed1984

Anetoderma: biochemical and ultrastructural demonstration of an elastin defect in the skin of three patients.

Three patients with localized cutaneous lesions characteristic of anetoderma were studied. Clinically, the onset of the disease was between the ages of 17 and 25, and numerous flaccid, saclike skin lesions developed over several subsequent years. Histologically, the lesions were characterized by paucity and fragmentation of the elastic fibers. Electron microscopy demonstrated that the elastic fibers, both in papillary and deep reticular dermis in the lesional skin, were fragmented and irregular in appearance. The concentration of elastin, determined by a radioimmunoassay of desmosine, an elastin-specific cross-link compound, was markedly reduced in the lesions, as compared with unaffected skin from the same patients or with normal skin from unrelated control subjects. In contrast, the concentrations of hydroxyproline, an index of collagen, or deoxyribonucleic acid (DNA), a measure of cellularity, were not changed in the lesions. Thus, the results indicate that in the three patients studied, the elastic fibers are defective and reduced in quantity. These observations suggest that the deficiency of elastin in the dermis may lead to development of the cutaneous lesions of anetoderma.

Adult↗
PubMed1984

Control of connective tissue metabolism by lasers: recent developments and future prospects.

Various laser modalities are currently in extensive use in dermatology and plastic surgery, particularly for treatment of vascular and pigmented lesions. A relatively new area of laser utilization involves the possible biologic effects of the lasers. In this overview, we are summarizing our recent studies, which indicate that lasers at specific wavelengths and energy densities modulate the connective tissue metabolism by skin fibroblasts both in vitro and in vivo. Specifically, the neodymium-yttrium-aluminum-garnet (Nd: YAG) laser was shown to selectively suppress collagen production both in fibroblast cultures and in normal skin in vivo, thus suggesting that this laser modality may be useful for the treatment of fibrotic conditions such as keloids and hypertrophic scars. Furthermore, two low-energy lasers, helium-neon (He-Ne) and gallium-arsenide (Ga-As), were shown to stimulate collagen production in human skin fibroblast cultures, suggesting that these lasers could be used for enhancement of wound healing processes. These experimental approaches illustrate the future possibilities for applying lasers for the modulation of various biologic functions of cells in tissues and attest to the potential role of lasers in the treatment of cutaneous disorders.

Animals↗
PubMed1984

Ultraviolet radiation-induced connective tissue changes in the skin of hairless mice.

Hairless mice (Skh/ hr1 ) were exposed to ultraviolet A (UVA; peak irradiance at 365 nm), or to ultraviolet B (UVB; peak irradiance at 313 nm) radiation. The animals received 12 treatments on alternate days. Connective tissue changes in the skin were monitored by assaying hydroxyproline and desmosine as an indication of collagen and elastin concentrations, respectively. The activities of prolyl hydroxylase and collagen glucosyl-transferase, enzymes participating in the biosynthesis of collagen, were also assayed. The concentration of elastin was significantly increased in mice treated with UVA or UVB. The concentration of collagen was unaffected by the treatments, but the activity of prolyl hydroxylase, reflecting collagen synthetic capacity, was decreased in UVA-treated mice. The collagen glucosyl-transferase activity was unchanged. Irradiation of purified human prolyl hydroxylase with UVA in vitro decreased the enzyme activity at higher doses, but UVB had no effect. The results indicate that definitive changes in the biochemistry of dermal connective tissues can be induced by exposure of mice to UV irradiation.

Animals↗
PubMed1984

Pharmacological inhibition of excessive collagen deposition in fibrotic diseases.

Excessive accumulation of collagen is a major pathological feature in diseases characterized by tissue fibrosis. Although several therapeutic approaches have been attempted in such patients, currently no treatment modality would specifically reduce collagen deposition in tissues. In this paper we discuss the mode of action of compounds that interfere with collagen production on the posttranslational level. First, structural analogs of proline, cis-4-hydroxy-L-proline and L-azetidine-2-carboxylic acid, which are incorporated into the newly synthesized polypeptides of procollagen during translation, prevent the polypeptides from folding into a stable triple-helical conformation. As a consequence, the nonhelical polypeptides are subject to degradation by proteases, thus leading to reduced deposition of collagen fibers. Second, several naturally occurring amino acids, polyamines, and their structural analogs prevent the removal of the carboxy-terminal extensions during the extracellular conversion of procollagen to collagen. Because the precursor molecules that contain the carboxy-terminal extensions are unable to assemble into functional fibers, collagen deposition is again reduced. Further development of these and related compounds, with appropriate tissue targeting, could potentially provide us with novel means to reduce the excessive deposition of collagen in fibrotic processes.

Animals↗
PubMed1983

Proteinases in human polymorphonuclear leukocytes. Purification and characterization of an enzyme which cleaves denatured collagen and a synthetic peptide with a Gly-Ile sequence.

Polymorphonuclear leukocytes have been shown to contain proteolytic enzymes which are capable of degrading connective tissue proteins such as native collagen. In this study, proteolytic enzymes were extracted from human polymorphonuclear leukocytes and a neutral proteinase was extensively purified and characterized. The activity of this enzyme was monitored by degradation of denatured [ 3H ]proline-labeled type I collagen or by cleavage of a synthetic dinitrophenylated peptide with a Gly-Ile sequence. The enzyme was readily separated from leukocyte collagenase by concanavalin-A--Sepharose affinity chromatography and further purified by QAE-Sephadex ion-exchange chromatography and gel filtration on Sephacryl S-200. The purified enzyme had a molecular weight of approximately 105000, its pH optimum was about 7.8, and it was inhibited by Na2EDTA and dithiothreitol, but not by fetal calf serum. The enzyme degraded genetically distinct type I, II, III, IV and V collagens, when in a non-helical form, but not when in native triple-helical conformation. Dansyl-monitored end-group analyses, combined with digestion by carboxypeptidase A, indicated that the enzyme cleaved denaturated type I collagen at Gly-Xaa sequences, in which Xaa can be leucine, isoleucine, valine, phenylalanine, lysine, or methionine. Thus, the purified enzyme referred to here as Gly-Xaa proteinase, is a neutral proteinase, which may be of importance in inflammatory disease processes by degrading further collagen peptides which have been rendered non-helical as a result of collagenase cleavage.

Amino Acid Sequence↗
PubMed1983

Conformational stability of type I collagen triple helix: evidence for temporary and local relaxation of the protein conformation using a proteolytic probe.

Native collagen polypeptides exist in a unique triple helical conformation resistant to most proteinases. In this study, the stability of type I collagen triple helix, employing a mixture of trypsin and alpha-chymotrypsin as a proteolytic probe, was examined. The degradation of type I [3H]collagen was monitored as 3H-labeled peptides soluble in trichloroacetic acid (TCA) or by sodium dodecyl sulfate (SDS)-polyacrylamide slab gel electrophoresis. In one set of experiments, collagen substrates were preincubated at various temperatures for up to 8 h, followed by a 15-min proteolytic treatment at the same temperature. At 43 degrees C, most of the collagen was degraded, while the fraction of the substrate degraded at 40, 38, and 35 degrees C was 53, 41 and 19%, respectively. This fraction was independent of the preincubation time which varied from 10 to 480 min. Thus, at any given temperature, a constant fraction of the collagen substrate was susceptible to proteolysis. Measurement of the midpoint temperature (Tm) of the helix to coil transformation for type I collagen, at neutral pH employing an increasing temperature gradient and brief proteolysis at the individual temperatures, indicated a value of 38.8 degrees C. However, determination of the Tm by employing proteolytic digestions at a constant temperature (30 degrees C) using conditions under which the nonhelical peptides are readily digested to TCA-soluble peptides while native collagen resists such proteolysis, indicated a value of 42.7 degrees C. In further studies, collagen was subjected to continuous proteolysis for up to 24 h. A large fraction of collagen was digested at 30 or 34 degrees C, temperatures well below the Tm of the helix to coil transformation. SDS-polyacrylamide gel electrophoresis of the degradation products obtained at these temperatures revealed multiple cleavage fragments. Finally, temperature double-jump experiments indicated that the destabilization of the triple helix is reversible provided that the Tm of the substrate is not exceeded. The results provide evidence for reversible and local relaxation of the collagen triple helix.

Collagen↗
PubMed1983

Verrucous lupus erythematosus: ultrastructural studies on a distinct variant of chronic discoid lupus erythematosus.

Ten patients with classic discoid lupus erythematosus of the face associated with verrucous, papulonodular lesions on the arms and hands were studied by electron microscopy. The ultrastructural findings on the verrucous lesions included apoptotic keratinocytes, intraepidermal lymphocytes, and gapping, detachment, and reduplication of the basal lamina. Also, tubuloreticular inclusion bodies were present in the endothelial cells. These observations, together with the clinical, histopathologic, and immunofluorescence findings, suggest that the verrucous lesions represent a rare, but distinct, variant of chronic discoid lupus erythematosus.

Chronic Disease↗
PubMed1983

Wound healing: biological effects of Nd:YAG laser on collagen metabolism in pig skin in comparison to thermal burn.

Pig skin was treated with the Nd:YAG laser at 1,060 nm or electrocautery, at energy densities of 649 +/- 20 J/cm2 and 612 J/cm2, respectively. Biopsies of treated areas and of normal skin were performed at 7, 14, and 60 days after treatment and processed for histology, electron microscopy and biochemical assays. Wound healing, as shown histologically, was similar in both treated groups. Depth of injury appeared to reach reticular dermis at day 7 in each treated group. However, thermal burn was more destructive of regular collagen, whereas the laser appeared to damage deep dermal blood vessels without destroying surrounding connective tissue. Biochemical assays revealed increased collagen production and increased collagenolytic activity 7 days after laser injury. However, by day 60, there was a reduction in total collagen content in laser treated skin below that of normal skin, which correlated with decreased collagen synthesis and unchanged collagenolytic activity. In burn specimens there was an initial decrease in total collagen content which reverted to normal by day 60. Active collagen degradation occurred at all 3 time points, but a marked increase in synthetic activity occurred as the burn scar was laid down. Laser treatment resulted in reduction of the amount of collagen below that in burn scarred or normal skin, suggesting that classical scar formation may be inhibited. These results indicate that the Nd:YAG laser may be useful for the treatment of keloids and hypertrophic scars.

Animals↗
PubMed1983

Demonstration of collagenase and elastase activities in the blister fluids from bullous skin diseases. Comparison between dermatitis herpetiformis and bullous pemphigoid.

We have investigated potential mechanisms for blister formation by assaying proteolytic enzymes in the blister fluids of patients with various bullous diseases. Blister fluids were obtained from patients with dermatitis herpetiformis (DH), bullous pemphigoid (BP), chronic bullous disease of childhood (CBDC), and pemphigus vulgaris (PV). The cells were recovered by centrifugation, and the supernatants as well as the cell pellets were assayed first for collagenase activity using [3H]proline-labeled type I collagen as substrate. Collagenase activity could be detected in most cases with DH, BP, and CBDC, while no activity was found in 2 cases of PV or in 5 control blister fluids obtained from suction blisters induced in healthy control subjects. Elastase activity was assayed in the same blister fluids by using a synthetic substrate succinyl-(L-alanyl)3-paranitroanilide or soluble [14C]valine-labeled tropoelastin. High levels of elastase activity were present in all DH patients, while lower, but clearly detectable, levels were found in BP, CBDC, and PV. The enzyme activity in BP was inhibited by Na2EDTA, but not by phenylmethylsulfonyl fluoride (PMSF), and Ca2+ stimulated the activity, suggesting that the enzyme in BP was a metalloproteinase. In cell-free supernatants of the DH blister fluids, the elastase activity was markedly decreased by PMSF, indicating that most of the enzyme activity was due to a serine protease. The cells recovered from DH blister fluids also contained high levels of elastase activity which could be inhibited by PMSF but not by Na2EDTA. Thus, in DH, the elastase activity is probably derived from polymorphonuclear leukocytes abundantly present in the lesions. The results indicate that active proteases are present in the blister fluids of skin diseases, and they may play a mechanistic role in the blister formation by degrading connective tissue components of the dermis and the dermal-epidermal junction.

Adult↗
PubMed1983

Proline analogues inhibit human skin fibroblast growth and collagen production in culture.

Several structural analogues of proline have been shown to be incorporated into proteins in place of proline. As a consequence, the proliferation of cells in culture and the extracellular deposition of collagen in animal systems are reduced. In this study, the effects of two proline analogues, cis-4-hydroxy-L-proline and L-azetidine-2-carboxylic acid, on the growth parameters and procollagen production by cultured normal human skin fibroblasts were examined. The results indicated that incubation of the cells with the analogues reduced the rate of fibroblast proliferation and lowered the plating efficiency. Further experiments demonstrated that fibroblasts in the presence of L-azetidine-2-carboxylic acid synthesized procollagen polypeptides which were not in a triple-helical conformation, as judged by limited pepsin proteolysis. Also, a significantly increased fraction of the newly synthesized collagenous peptides was in a dialyzable form, suggesting increased degradation of the nonhelical chains. The rate of translation of collagenous polypeptides and the preprocollagen messenger RNA activity in the cells were not affected by the analogues. The proline analogues thus appear to inhibit the production of procollagen on the posttranslational level by preventing the polypeptides from folding into a stable triple-helical conformation. The nonhelical polypeptides are then readily susceptible to proteolysis leading to reduced deposition of extracellular collagen fibers. Similar experiments were also performed with fibroblasts cultured from patients with active progressive systemic sclerosis. Quantitatively and qualitatively comparable inhibition of procollagen production by L-azetidine-2-carboxylic acid was noted with scleroderma cells as with control fibroblast cultures. The results suggest, therefore, that proline analogues may, in the future, prove useful in limiting excessive collagen deposition in scleroderma and other forms of dermal fibrosis.

Azetidinecarboxylic Acid↗
PubMed1983

Elastic fibers in human skin: quantitation of elastic fibers by computerized digital image analyses and determination of elastin by radioimmunoassay of desmosine.

The elastic fibers in the skin and other organs can be affected in several disease processes. In this study, we have developed morphometric techniques that allow accurate quantitation of the elastic fibers in punch biopsy specimens of skin. In this procedure, the elastic fibers, visualized by elastin-specific stains, are examined through a camera unit attached to the microscope. The black and white images sensing various gray levels are then converted to binary images after selecting a threshold with an analog threshold selection device. The binary images are digitized and the data analyzed by a computer program designed to express the properties of the image, thus allowing determination of the volume fraction occupied by the elastic fibers. As an independent measure of the elastic fibers, alternate tissue sections were used for assay of desmosine, an elastin-specific cross-link compound, by a radioimmunoassay. The clinical applicability of the computerized morphometric analyses was tested by examining the elastic fibers in the skin of five patients with pseudoxanthoma elasticum or Buschke-Ollendorff syndrome. In the skin of 10 healthy control subjects, the elastic fibers occupied 2.1 +/- 1.1% (mean +/- SD) of the dermis. The volume fractions occupied by the elastic fibers in the lesions of pseudoxanthoma elasticum or Buschke-Ollendorff syndrome were increased as much as 6-fold, whereas the values in the unaffected areas of the skin in the same patients were within normal limits. A significant correlation between the volume fraction of elastic fibers, determined by computerized morphometric analyses, and the concentration of desmosine, quantitated by radioimmunoassay, was noted in the total material. These results demonstrate that computerized morphometric techniques are helpful in characterizing disease processes affecting skin. This methodology should also be applicable to other tissues that contain elastic fibers and that are affected in various heritable and acquired diseases.

Amino Acids↗
PubMed1983

Effects of the Nd:YAG laser on DNA synthesis and collagen production in human skin fibroblast cultures.

Human skin fibroblasts were subjected to treatment with a Neodymium:YAG laser at 1060 nm with varying levels of energy determined by a reproducible method of dosimetry. DNA synthesis in the cells was measured by the incorporation of [3H]thymidine, and collagen production was monitored by the synthesis of nondialyzable [3H]hydroxyproline after incubation of cells with [3H]proline. Using energy levels equal to 1.7 X 10(3) J/cm2, a significant reduction in DNA synthesis was noted, while the cells remained viable as tested by the trypan blue exclusion test. With energy levels higher or equal to 2.3 X 10(3) J/cm2, the suppression of DNA synthesis was accompanied by cell nonviability. The collagen production, when measured immediately following the treatment with 1.7 X 10(3) J/cm2, was markedly reduced, and similar effects were observed with higher energy levels. However, when the cells were tested for collagen production at 20 hours following laser treatment, there was a significant decrease in collagen production at energy levels as low as 1.1 X 10(3) J/cm2, a dose that did not affect DNA synthesis or cell viability. Thus, the results indicate that the Nd:YAG laser can selectively suppress collagen production without affecting cell proliferation. These observations suggest that laser treatment could potentially be used to reduce collagen deposition in conditions such as keloids and hypertrophic scars.

Cells, Cultured↗
PubMed1982

Assay of collagenase activity by a rapid, sensitive, and specific method.

A new method for the assay of collagenase activity has been developed, whereby the collagen cleavage products, after initial collagenase digestion, are degraded further by a mixture of trypsin and alpha-chymotrypsin. The degradation products are soluble in TCA and can be conveniently separated from the remaining uncleaved collagen substrate by rapid filtration. The enzyme assay is shown to be reproducible and sensitive, and it lends itself to a convenient and rapid determination of collagenase activity in relatively large numbers of samples. The applicability of this method is demonstrated by the detection of increased collagenase activity in skin fibroblast cultures derived from a patient with recessive dystrophic epidermolysis bullosa.

Bacteria↗