Fungi ; isolation & purification ; Histocytochemistry ; methods ; Humans ; Immunohistochemistry ; methods ; In Situ Hybridization ; Periodic Acid-Schiff Reaction ; Polymerase Chain Reaction ; methods ; Polymorphism, Restriction Fragment Length ; Polymorphism, Single-Stranded Conformational ; Random Amplified Polymorphic DNA Technique ; Rhinitis ; microbiology ; Sinusitis ; microbiology

BACKGROUND: The cornified cell envelope(CE) which is formed during the terminal differentiation of keratinocytes, is a specialized structure which forms a structurally and functionally complete permeability barrier. OBJECTIVE: The purpose of our study is to investigate the effects of changes in the calcium ions on keratinocyte differentiation, especially in the expression of CE protein. METHODS:The permeability barrier of hairless mice was disrupted by tape-stripping and then exposed to the air or occluded with a water-vapor impermeable membrane, and iontophoresis was done without permeability barrier perturbation. Skin specimens were prepared for ion capture cytochemistry and immunohistochemistry with anti-K5, anti-K10, anti-K6, anti-involucrin and anti-loricrin. RESULTS: The calcium gradient which disappeared after tape-stripping was restored at 36 h after tape-stripping with air exposure and at 60 h after tape-stripping with occlusion. The change in calcium ions produced by both positive and negative iontophoresis showed recovery at 6 h. Expression of basal K5 showed a slight decrease and expression of suprabasal K10 showed an increase at 12 h with air exposure after tape-stripping, tape stripping with occlusion, and iontophoresis. Expression of K6 appeared at 12 h after tape-stripping and then in the whole epidermis at 36 h with air exposure after tape-stripping and tape stripping with occlusion and focally appeared in the stratum granulosum and stratum spinosum after iontophoresis. Expression of involucrin was increased at 12 h with air exposure after tape-stripping and iontophoresis and was extended to the lower spinous layers in tape-stripping with occlusion. Expression of loricrin in air exposure after tape-stripping, tape-stripping with occlusion and iontophoresis was similar to that of normal skin. CONCLUSION: The changes in calcium ions without permeability barrier perturbation are related to the expression of CE protein. It is thought that calcium ions in the epidermis have an important role in the terminal differentiation of keratinocytes.
Animals ; Calcium* ; Epidermis ; Histocytochemistry ; Immunohistochemistry ; Ions ; Iontophoresis ; Keratinocytes* ; Membranes ; Mice ; Mice, Hairless ; Permeability ; Skin

BACKGROUND: The cornified cell envelope(CE) which is formed during the terminal differentiation of keratinocytes, is a specialized structure which forms a structurally and functionally complete permeability barrier. OBJECTIVE: The purpose of our study is to investigate the effects of changes in the calcium ions on keratinocyte differentiation, especially in the expression of CE protein. METHODS:The permeability barrier of hairless mice was disrupted by tape-stripping and then exposed to the air or occluded with a water-vapor impermeable membrane, and iontophoresis was done without permeability barrier perturbation. Skin specimens were prepared for ion capture cytochemistry and immunohistochemistry with anti-K5, anti-K10, anti-K6, anti-involucrin and anti-loricrin. RESULTS: The calcium gradient which disappeared after tape-stripping was restored at 36 h after tape-stripping with air exposure and at 60 h after tape-stripping with occlusion. The change in calcium ions produced by both positive and negative iontophoresis showed recovery at 6 h. Expression of basal K5 showed a slight decrease and expression of suprabasal K10 showed an increase at 12 h with air exposure after tape-stripping, tape stripping with occlusion, and iontophoresis. Expression of K6 appeared at 12 h after tape-stripping and then in the whole epidermis at 36 h with air exposure after tape-stripping and tape stripping with occlusion and focally appeared in the stratum granulosum and stratum spinosum after iontophoresis. Expression of involucrin was increased at 12 h with air exposure after tape-stripping and iontophoresis and was extended to the lower spinous layers in tape-stripping with occlusion. Expression of loricrin in air exposure after tape-stripping, tape-stripping with occlusion and iontophoresis was similar to that of normal skin. CONCLUSION: The changes in calcium ions without permeability barrier perturbation are related to the expression of CE protein. It is thought that calcium ions in the epidermis have an important role in the terminal differentiation of keratinocytes.
Animals ; Calcium* ; Epidermis ; Histocytochemistry ; Immunohistochemistry ; Ions ; Iontophoresis ; Keratinocytes* ; Membranes ; Mice ; Mice, Hairless ; Permeability ; Skin

The cytologic distinction of carcinoma cells from reactive mesothelial cells can be difficult, especially in specimens containing abundant reactive mesothelial cells and inflammatory cells with scant carcinoma cells. This study evaluates the usefulness of mucin and immunocytochemistry for discrimination between reactive mesothelial cells and carcinoma cells, and sensitivity and specificity of these stains for the detection of metastatic carcinoma in serous effusions. Immunocytochemical panel including mucin cytochemistry with the periodic acid-Schiff(PAS) reaction after or without diastase digestion was undertaken on 127 serous effusion specimens with histologically confirmed diagnoses. The specimens including cell smears and cell blocks were stained with PAS and antibodies to carcinoembryonic antigen(CEA), epithelial membrane antigen(EMA), cytokeratin(CK), and vimentin. The sensitivities of these stains for metastatic carcinoma(127 cases) were 49%(46/94) in PAS, 48%(60/124) in CEA, 89%(97/109) in EMA, 88%(93/106) in CK, and 25%(20/81) in vimentin. The sensitivities of stains for reactive mesothelial cells(36 cases) were 19%(7/36) in EMA, 78%(28/36) in CK, and 75%(27/36) in vimentin. The PAS and CEA stains were not reacted with all cases of benign reactive serous effusions containing abundant reactive mesothelial cells. The specificities of stains for metastatic carcinoma(127 cases) were 100% in PAS, 100% in CEA, 81% in EMA, 22% in CK, and 25% in vimentin. The optimal combination of stains for use in a panel was PAS and CEA. Combined results from these two stains yielded an advanced sensitivity of 8% in PAS and 4% in CEA for metastatic carcinoma. EMA wasalso cosiderably useful for identification of carcinoma cells. CK and vimentin were not suitable for distinguishing between reactive mesothelial cells and carcinoma cells.
Amylases ; Antibodies ; Coloring Agents ; Diagnosis ; Digestion ; Discrimination (Psychology) ; Histocytochemistry ; Immunohistochemistry ; Membranes ; Mucins ; Sensitivity and Specificity ; Vimentin

We investigated the response to challenge infection with Cryptosporidium parvum oocysts in immunosuppressed C57BL/6N mice. In the primary infection, fecal oocyst shedding and parasite colonization were greater in immunosuppressed mice than in nonimmunosuppressed mice. Compared with primary infection, challenge infection with C. parvum didn't show any oocyst shedding and parasite colonization. Especially, oocyst shedding and parasite colonization from the mice infected with heatkilled oocysts were not detected. After challenge infection with C. parvum oocysts, however, these mice were shedding small numbers of oocysts and parasite colonization. Except normal control and uninfected groups, the antibody titers of other groups appear similar. Based on the fecal oocyst shedding, parasite colonization of ilea, and antibody titers in the mice, these results suggest that the resistance to challenge infection with C. parvum in immunosuppressed C57BL/6N mice has increased.
Animals ; Antibodies, Protozoan/blood ; Cryptosporidiosis/*immunology/*parasitology ; Cryptosporidium parvum/*immunology ; Dexamethasone/immunology ; Enzyme-Linked Immunosorbent Assay ; Feces/parasitology ; Female ; Fluorescent Antibody Technique, Indirect ; Histocytochemistry ; Ileum/parasitology ; Immunocompromised Host ; Mice ; Mice, Inbred C57BL ; Oocysts/immunology ; Random Allocation

Abrupt reoxygenation (or reperfusion) after anoxia (or ischemia)-induced injury resulted in the loss of contractile property, destruction of cell organelles, and ultimately, cell death in cardiac myocytes. This phenomenon has been called 'oxygen paradox' or 'reperfusion injury'. The purpose of this study was to investigate the changes of fine structures and enzyme activities associated with oxygen paradox during 60 min. of anoxia, followed by a 30 min. of reoxygenation. Cardiac myocytes were dissociated from neonatal rat ventricles and cultured for three days. While they were exposed to anoxia and reoxygenation, the cardiac myocytes were investigated through beating counts, enzyme cytochemistry, immunofluorescence, electron microscopy for morphological study. Activity staining and Western blot for Cu, Zn-SOD, NADPH-diaphorase stain and nitrite concentration mesurement for nitric oxide synthase, and catalase activity measurement were performed. After 60 min. of anoxia, the beating rate increased remarkably. Swollen mitochondria with amorphous dense clumps, mild contracture of myofibrils and retraction of cytoplasmic processes were observed in cardiac myocytes. Under confocal microscope, weak reaction of Mn-SOD and myosin were observed, whereas reaction of Cu, Zn-SOD was enhanced in perinuclear region. Cu, Zn-SOD and catalase activity in cardiac myocytes increased markedly. Nitric oxide synthase activity increased gradually with time. After 30 min. of reoxygenation following 60 min anoxia, structural changes of myocardial cells was more pronounced than in the cells of anoxic group. Beating rate was variable but decreased gradually. Myocardial cells showed evidence of severe structural alterations, including marginal clumping of chromatids, varying-sized bleb formation, many vacuoles, mitochondrial matrix exposed to cytoplasm and fragmen-tation of cristae, myofibrillar hypercontracture. Decline of immunocytochemical reaction of Mn-SOD, myosin and Cu, Zn-SOD were observed under confocal microscope. The declines of activity and quantity of Cu, Zn-SOD were severe compared to control. In contrast, nitric oxide synthase activity significantly increased. Catalase activity was lower than in anoxic group, but still higher than in control activity. These results suggested that there were two possible mechanisms for the drastic morphological changes induced by anoxia-reoxygenation; 1) direct effect of oxygen free radicals, and 2) reaction of nitric oxide with superoxide radicals, which resulted in generation of toxic metabolites of nitric oxide, exacerbated myocardial cellular damages.
Animals ; Anoxia ; Blister ; Blotting, Western ; Catalase ; Cell Death ; Chromatids ; Contracture ; Cytoplasm ; Fluorescent Antibody Technique ; Free Radicals ; Histocytochemistry ; Microscopy, Electron ; Mitochondria ; Myocytes, Cardiac* ; Myofibrils ; Myosins ; Nitric Oxide ; Nitric Oxide Synthase ; Organelles ; Oxygen ; Rats ; Superoxide Dismutase ; Superoxides ; Vacuoles

BACKGROUND: Immunophenotyping is an important technique for the diagnosis and classification of acute leukemia, as well as French-American-British (FAB) classification on the basis of morphologic characteristics and cytochemistry. We evaluated the expression patterns of immunologic surface markers in acute leukemia. METHODS: Peripheral or bone marrow leukemic cells from 75 leukemic patients (acute lymphoblastic leukemia, ALL 40 cases; children (26 cases), adults (14 cases) and acute myeloid leukemia, AML 35 cases; children (9 cases), adults (26 cases)) were studied. Monoclonal antibodies which were designed for two color direct immunofluorescence (IF) analysis with combination of fluoresceinisothiocynate (FITC) and phycoerythrin (PE) conjugated, CD10/CD19, CD20/CD5, CD3/CD22, CD7/CD33, HLA-DR/CD13 (Acute Leukemia Phenotyping Kit, Becton Dickinson; BD, USA) were analyzed by flow cytometry. RESULTS: Blasts from these patients could be classified as CALLA (+)B-ALL (26 cases, 65.0%), CALLA (-)B-ALL (6 cases, 15.0%), T-ALL (6 cases, 15.0%), biphenotypic ALL (2 cases, 5.0%). The positive expression rates were CD19 (100%), CD10 (78.1%), CD22 (75.0%) and CD20 (50.0%) in B-ALL, CD7 (100%), CD3 (50.0%) and CD5 (50.0%) in T-ALL and CD33 (85.7%), CD13 (74.3%) in AML, respectively. The incidence of acute mixed lineage leukemia (AMLL) was 26.7% and leukocytosis, anemia and thrombocytopenia were frequently seen in AMLL. CONCLUSION: By the study of immunophenotyping we could more exactly diagnosed ALL and AML, as well as AMLL which was not exactly diagnosed by characteristics of morphology and cytochemistry only. Therefore the best method for the diagnosis of acute leukemia will be achieved by using of immunophenotyping and FAB classification on the basis of morphology and cytochemistry.
Adult ; Anemia ; Antibodies, Monoclonal ; Antigens, Surface* ; Bone Marrow ; Child ; Classification ; Diagnosis ; Flow Cytometry ; Fluorescent Antibody Technique, Direct ; Histocytochemistry ; Humans ; Immunophenotyping ; Incidence ; Leukemia* ; Leukemia, Myeloid, Acute ; Leukocytosis ; Phycoerythrin ; Precursor Cell Lymphoblastic Leukemia-Lymphoma ; Precursor T-Cell Lymphoblastic Leukemia-Lymphoma ; Thrombocytopenia

In view of the sparsity of report on normal and abnormal pattern of the major arteries during fetal life, the authors undertook investigation of the aortas, coronary arteries, renal arteries, and the umbilical arteries of 76 korean fetuses, ranging from 21/2 months to full-term, A normal pattern and its evolution of the intima, media, and the adventitia was described. The P. A. S. positive substance was most abundantly found in the media of the umbilical arteries, medium amount in the media and intima of the renal arteries and the aortas, and lesser amount in the media of the coronary arteries. A surprisingly high incidence of the alteration of the internal elastic membrane of the aorta simulating to the early lesions of atherosclerosis in adult and neonatal life was observed. In e1even instances, microcystic degeneration of the inner media of the aorta was observed, and its relationship to the idiopathic cystic medial necrosis and dissecting aneurysm was discussed.
Arteries/*embryology ; Arteriosclerosis/*etiology ; Female ; Histocytochemistry ; Human ; Korea ; Pregnancy

OBJECTIVETo study the histochemical staining in the diagnosis of osteosarcoma.

METHODSTo compare the effectiveness of picrosirius red, improved Ponceau trichrome and Masson trichrome staining methods on bone formation tissues in conventional osteosarcoma, paraosteal osteosarcoma, periosteal osteosarcoma, extraskeletal osteosarcoma, inflammatory myofibroblastic tumour, malignant fibrohistiocytoma, chondrosarcoma, fibrosis with ossification and calcification.

RESULTSWith modified Ponceau trichrome staining, bone formation tissues showed a homogenous, orange-red interblended with blue in color. From osteoid to mature bone the color changed from orange-red, light blue to dark blue. Fibrotic tissue was stained blue in color with striated appearance. Cartilage was not stained. Picrosirius red method gave bone formation tissues homogenous staining. Along with bone maturation, from osteoid tissue to mineralized bones, the color showed changes from light red, yellow, orange-red, red to dark purple. The cartilage demonstrated homogenous light red in color. Fibrous tissue stained red interblended with yellow in color, striated in shape. With Masson trichrome staining osteoid displayed pale blue and mineralized bone showed dark blue in color. Fibrotic tissue showed a striated blue staining.

CONCLUSIONThe modified Ponceau trichrome and Picrosirius red staining methods are better than Masson trichrome to demonstrate bone formation tissue in osteosarcoma. The former two methods could be also used in study on bone formation.

BACKGROUND: Cold is considered an important environmental factor on the skin's barrier function. It is also known that cold can disrupt the barrier's permeability function. Historically irritant contact dermatitis develops frequently amongst fish processing workers. This means that cold may disrupt the skin barrier function. However, there are only a few studies of the effect of cold on the skin barrier. OBJECTIVE: The aim of our study was to investigate the effects of exposure to the cold on the skin's barrier permeability, calcium gradient and cornified enveloped proteins. METHODS: Hairless mice were classified into four groups as follows; tape-stripping and exposure to cold (4degrees C) for 5 hours group (Group 1), exposure to cold (4degrees C) for 5 hours after tape-stripping and occlusion with Gore-Tex group (Group 2), exposure to cold (4degrees C) for 5 hours after tape-stripping and applying with petrolatum group (Group 3), and tape-stripping and exposure at room temperature (25degrees C) for 5 hours group (Group 4). Next, rate of barrier recovery was measured, and skin of cold exposure site was taken for immunohistochemistry of cornified enveloped protein, calcium inon capture chemistry and in situ hybridization. RESULTS: Rate of barrier recovery of Group 1 was measured at -56.33%, the most delayed rate of all groups. Rate of barrier recovery of other groups were measured as follows; Group 2 (20%), Group 3 (55%) and Group 4 (41.78%). In calcium-ion capture cytochemistry, Group 1 appeared to absent calcium deposit in the entire epidermis. Group 3 appeared to the highest density of calcium in entire epidermis. In immunohistochemical staining of cornified enveloped proteins including involucrin, loricrin and filaggrin, Group 1 resulted in the lowest expression of cornified enveloped proteins. Of all groups, Group 3 appeared to have the highest expressionof cornified enveloped proteins. In situ hybridization of loricrin and filaggrin, expression were similar with results of immunohistochemical staining. CONCLUSION: Our results provide that cold impairs the skin's barrier permeability by having a negative effect on recovery of calcium gradient and differentiation of cornifed enveloped proteins. But occlusion with vapor permeable membranes such as Gore-Tex or petrolatum could improve conditions of skin barrier impaired by cold by restoring the recovery rate of calcium gradient and differentiation of cornified enveloped protein.
Animals ; Calcium ; Cold Temperature ; Dermatitis, Contact ; Epidermis ; Histocytochemistry ; Immunohistochemistry ; In Situ Hybridization ; Intermediate Filament Proteins ; Membrane Proteins ; Membranes ; Mice ; Mice, Hairless ; Permeability ; Petrolatum ; Polytetrafluoroethylene ; Protein Precursors ; Proteins ; Skin