Gene/Protein Disease Symptom Drug Enzyme Compound
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Adult Swiss webster mice were injected with 3 x 10(6) colony-forming units (cfu) of group G or 2.5 x 10(6) cfu of group A streptococci at intradermal injection sites on the right and left paralumbar areas of the back. The mice were sacrificed at intervals between 4 hours and 14 days post-injection (p.i.) and full thickness biopsies of skin 10 mm in diameter encompassing the sites of injection were taken. One tissue specimen was homogenized in PBS and plated to determine the number of cfu, while another was used for histopathological studies. The number of viable group A and group G streptococci in the tissue increased to 3 x 10(9) cfu by 96 hours p.i.: after 192 hours p.i. the group A cells had declined to 2.7 x 10(6) cfu compared to 1.1 x 10(8) cfu for group G cells. No streptococci of either group were detected at 336 hours (14 days p.i.). Gross edematous lesions induced by either streptococcus group were evident on all animals at 24 hours (p.i.). Group G streptococci lesions were larger and persisted longer than lesions induced by group A. Histological examination consistently revealed more inflammation and necrosis in tissue sections from mice injected with group G streptococci.
Exp Dermatol 1992 Dec
PMID:Comparison of skin changes induced on mice by either group A type 12 or group G streptococci. 136 27

Most techniques used to visualize cells in tissues are accompanied by some distortion of the tissue due to fixation and sectioning. We here present a technique involving "optical sectioning" and three-dimensional reconstruction of fluorescence-labelled Langerhans' cells in human skin that avoids such problems. The instrument used to study the specimens was a PHOIBOS Confocal Scanning Laser Microscope (CSLM) built around a Zeiss Universal microscope. A software package for three-dimensional reconstructions of confocal sections was employed for calculations in a Hewlett-Packard HP9000-350 minicomputer running UNIX. Normal human skin obtained at plastic surgery or skin biopsy specimens from allergic, irritant, or control patch test reactions was studied. Epidermis separated from dermis by incubation in thermolysin or thick (25 microns) cryostat sections were incubated with fluorescein-isothiocyanate labelled mouse monoclonal antibodies directed against HLA-DR. To reduce the fading of immunofluorescence during microscopy, the specimens were mounted with glycerol in PBS containing p-phenylenediamine. The results indicate that CSLM is a powerful tool for investigations on Langerhans' cells in different conditions of the skin allowing a three-dimensional view of cells in unfixed or acetone-fixed preparations at the light microscope level.
Arch Dermatol Res 1990
PMID:Three-dimensional visualization of human Langerhans' cells using confocal scanning laser microscopy. 218 43

In order to characterize immunoglobulins found on amyloid deposits of lichen amyloidosus and macular amyloidosis, and elution from cryostat sections was performed with citrate buffer, glycine buffer, NaCl, and PBS. Resulting eluates (mainly IgG) were examined with dot immunoblotting and SDS-PAGE immunoblotting and were found to react with the human epidermal keratin of 50 and 67 kD. Antikeratin autoantibody activities in normal murine and human sera were examined using a dot immunoblotting assay. In murine sera, titers of IgG and IgM autoantibodies were higher in older mice. The human cord blood showed significantly lower IgM autoantibody titers, whereas IgG antibody titers showed no significant differences from adults' sera, probably due to the permeability of IgG through the placental barrier. A stronger antibody activity in older individuals was thought to be due to the repeated exposures to keratin proteins derived from apoptotic keratinocytes. Sera from lichen amyloidosus and macular amyloidosis patients did not show any difference from normal controls in their antikeratin titers. It was concluded that the patients with lichenoid or macular amyloidosis are capable of producing a normal level of antikeratin autoantibodies. However, the removal of opsonized keratin-type amyloid from the skin is slow or deficient due to as yet unknown factors.
Arch Dermatol Res 1989
PMID:Antikeratin autoantibodies in the amyloid deposits of lichen amyloidosus and macular amyloidosis. 248 Jul 51

In newborn rat skin, disulfide bonds exist in the cell membrane of the stratum corneum and in the cytoplasm of the stratum granulosum. However, in the latter region, they can be detected after ethanol fixed epidermis remains for 2 weeks at 4 degrees C. Thus, the idea arose that the visualization of the disulfide bonds in the stratum granulosum may be caused by proteases. Therefore, the in situ effects by both activation and inhibition of epidermal proteases were examined in tissue sections. Cryostat sections which were fixed in cold ethanol (-20 degrees C, 98%) were incubated either in PBS, pH 7.4, containing 10mM 2-ME as a control or in an epidermal homogenate (15,000 x g supernatant fraction containing 2-ME) at 37 degrees C for 30 minutes and 4 hours, respectively, in order to activate the epidermal proteases and then reacted with NEM, 2-ME, and DACM in steps. Many minute fluorescent particles were seen in the stratum granulosum under both of the above conditions. However, with incubation in PBS without 2-ME, no fluorescent particles were seen. In addition, when the tissue was treated with NEM and zinc chloride before incubation, no fluorescent particles were seen. O-phenanthrolin remarkably inhibited the appearance of these fluorescent particles. In contrast, leupeptin and antipain only slightly inhibited the appearance of these fluorescent particles, and pepstatin and phenylmethylsulfonyl fluoride didn't inhibit at all. In general, as NEM and zinc chloride strongly inhibit thiol proteases, these findings suggest that presence of the minute fluorescent particles in the stratum granulosum could be due to the effects of proteases, especially thiol proteases.
J Dermatol 1989 Feb
PMID:The substrate of epidermal proteases--effects of in situ activation of epidermal thiol proteases on stratum granulosum cells. 265 3

To better understand the role of autoimmunity in the pathogenesis of dermatitis herpetiformis, linear IgA bullous dermatosis or other skin disorders, the antigenic specificity of the immune reactants bound in vivo in the skin must be identified. In order to do so, one must first be able to elute these immune reactants from the skin. We describe here a simple method of eluting not only specifically bound IgG, but also IgA and other immunoglobulins and complement components from skin biopsy material. The method involves cutaneous washing of the entrapped serum proteins in PBS pH 7 and pH 5 buffers followed by specific immunoglobulin elutions at pH 3 and 2. The IgA deposits which could not be removed by this treatment were eluted by a combination of low pH (0.5 M citrate pH 2) and a chaotropic agent (2 M NaCl). The relative concentration of IgA in eluates when quantitated by fluoroimmunoassay were three- to five-fold higher in dermatitis herpetiformis skin biopsy specimens, than in eluates of bullous pemphigoid or normal skin biopsy specimens.
Arch Dermatol Res 1989
PMID:A simple method for elution of IgA deposits from the skin of patients with dermatitis herpetiformis. 268 62

A Japanese patient with epidermolysis bullosa acquisita (EBA) was autopsied, and direct immunofluorescence (DIF) testing was performed. Using this patient's serum (EBA serum) and three bullous pemphigoid (BP) sera, the anatomical distribution and immunological characteristics of EBA antigen and BP antigen were investigated by indirect immunofluorescence (IIF). EBA antigen showed the same anatomical distribution as BP antigen in DIF and IIF studies; both antigens were limited to the skin, tongue, oesophagus, trachea, cornea and bladder. EBA antigen was located on the dermal side of both NaCl and PBS-separated skin, whereas BP antigen was limited to the epidermal side. Ethanol fixation abrogated the antigenic stability of BP antigen, but not that of EBA antigen. No difference was found when acetone or formalin fixation was used. The separation methods and prefixation in ethanol could be useful techniques applicable to the classification of the bullous disorders which manifest circulating anti-BMZ antibodies.
Br J Dermatol 1986 Jun
PMID:Anatomical distribution and immunological characteristics of epidermolysis bullosa acquisita antigen and bullous pemphigoid antigen. 352 10

Bullous pemphigoid antigen (BP Ag) is a cell surface marker of epidermal basal cells. The functional role of this molecule is unknown. Epidermal cell suspensions obtained by trypsinization of skin show a population of epidermal basal cells with a polar rim of antigen as demonstrated by indirect immunofluorescence technique. This study shows that treatment of these cells suspensions with a variety of proteolytic and glycosidic enzymes failed to remove the antigen from these basal cells. BP Ag was also stable upon incubation with distilled water, Triton X-100, PBS, and 1 M NaCl. Treatment of epidermal basal cells with 2 N NaSCN, 1% periodic acid, and 4 M urea, as well as acidic pH or 56 degrees C temperature, abolished the reactivity of these cells with BP antibodies.
J Invest Dermatol 1981 Mar
PMID:Some biochemical properties of pemphigoid antigen bound to the surface of dissociated epidermal basal cells. 701 11

Water-soluble extracts from psoriatic scales and normal human skin were prepared using either phosphate-buffered saline, pH 7.2, or 0.1 M carbonate buffer, pH 10.8. Anaphylatoxin C5a des Arg was quantified using a novel sandwich enzyme-linked immunosorbent assay (ELISA) using neoepitope-specific monoclonal antibodies. Alkali was about five to eight times more efficient than PBS in extracting C5a des Arg from scales, probably via dissociation of bound C5a des Arg. C5a des Arg concentration in scales from three patients suffering from psoriasis vulgaris varied between 2.5 and 4.6 ng/mg scale. No C5a des Arg was detectable in normal skin extracts. The biological activity of alkali-extractable C5a des Arg, i.e. chemotaxis, was preserved. The concentration of C5a des Arg relative to the concentration of albumin was taken as a parameter of the degree of complement activation in the psoriatic lesions, and was found to be more than six times higher than values attained in serum after maximum complement activation by zymosan. We conclude that complement activation may play a quantitatively important role in the inflammatory process in psoriasis.
Arch Dermatol Res 1993
PMID:Surprisingly high levels of anaphylatoxin C5a des Arg are extractable from psoriatic scales. 850 93

Because interleukin (IL)-10 is an immunoregulatory cytokine that is produced by keratinocytes exposed to UVB radiation (UVR), we determined whether IL-10 participates in either failed contact hypersensitivity (CH) induction or tolerance after acute, low-dose UVR. Murine recombinant IL-10 (200 ng) was injected intradermally on shaved abdominal skin. To assess the effects of IL-10 on CH induction, dinitrofluorobenzene (DNFB, 185 microg) was painted on the skin within 30 min after IL-10 was injected, and the mice were assayed 5 d later by ear challenge with dilute DNFB. To assess tolerance, DNFB (185 microg) was painted a second time on normal body-wall skin 14 d after DNFB was first painted on IL-10-injected skin; CH was then assayed on day 19. We found that mice that received DNFB on IL-10-injected skin developed CH comparable in intensity to that observed in PBS-injected controls. Thus, this dose of IL-10 did not prove to be deleterious to CH induction if hapten was painted on the injected site within 30 min. By contrast, mice that first experienced DNFB within 30 min, 1 d, or 3 d after IL-10 had been injected intracutaneously displayed hapten-specific tolerance. Moreover, intraperitoneally injected anti-IL-10 antibody prevented UVR- and cis-urocanic acid-dependent tolerance; anti-IL-10 antibody had no effect on TNF-alpha-induced tolerance and failed to restore CH induction after UVR exposures. These data indicate that IL-10 is an important mediator of the tolerance induced when hapten is painted on the skin of animals exposed to acute, low-dose UVR.
J Invest Dermatol 1997 Jul
PMID:Hapten-specific tolerance induced by acute, low-dose ultraviolet B radiation of skin is mediated via interleukin-10. 920 50

Dexamethasone palmitate (D-PAL) incorporated into lipid microspheres (D-PAL emulsion) is taken up by the reticuloendothelial system and by some inflammatory cells. Therefore, it has a stronger anti-inflammatory activity than free corticosteroids in vivo. To study the effect of D-PAL emulsion on systemic lupus erythematosus (SLE), we administered D-PAL emulsion to MRLlpr/lpr mice, an animal model for human SLE. The effect of D-PAL emulsion was compared with that of methylprednisolone (m-PSL), a water-soluble steroid. Percent survival was higher in the group treated with 0.25 mg of D-PAL emulsion intravenously once every 4 weeks than in those groups treated similarly with m-PSL or PBS control. Swelling of lymph nodes was frequent in the group treated with m-PSL or with PBS, while rarely observed in the group treated with D-PAL emulsion. Proteinuria was more frequent in the groups treated with m-PSL or PBS than in the group treated with D-PAL emulsion. Although the frequency of skin lesions was not different between these three groups, the control and m-PSL treated mice had severe skin lesions, such as hair loss of erythematous skin with scales and crusts at the nape, while D-PAL emulsion treated animals showed only facial alopecia without inflammatory skin changes. These data demonstrate that D-PAL emulsion was more effective than a corresponding dose of m-PSL on autoimmune prone mice. This suggests that intermittent administration of D-PAL emulsion may be effective in the treatment of human SLE.
J Dermatol Sci 1997 Nov
PMID:Effects of liposteroid on skin lesions in autoimmune MRLlpr/lpr mice. 943 7


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