Glycosaminoglycans (GAGs)are essential components of the extracellular matrix of various tissues.They have been known to regulate cellular and extracellular environments for cell survival and differentiation in various physiologic and pathologic conditions.The present study was conducted to identify the distribution of GAGs in subepithelial cortical fibers of normal and traumatic cataractous lens of rabbits. Traumatic cataract was made by piercing the anterior lens capsule with 25-gauge sharp needle and the rabbits were killed at different time intervals (1, 2, 3 and 4 weeks).From the enucleated eyes subepithelial lens cortical fibers were obtained.A normal rabbit lens was used for the control. The specimens were stained with 0.05%ruthenium red (RR)and processed for histochemical electron microscopy. RR reactive materials were identified as fine granular or filamentous structures.In normal rabbit lens they were present mainly along the surface of lens epithelial cells, the surface of the subepithelial cortical fibers. In cataractous rabbits lens, strong RR positive reactions were observed along the surface of the lens epithelial cells and subepthelial cortical fibers as well as at the intercortical fiber spaces and even within its micro-organelles. This investigation resulted in an illustration of the ultrastructural distribution of GAGs in normal and traumatic cataractous lens of rabbit.this may suggest altered GAG distribution may closely related to the formation of lens opacity.
Cataract* ; Cell Survival ; Epithelial Cells ; Extracellular Matrix ; Glycosaminoglycans ; Microscopy, Electron ; Needles ; Rabbits* ; Ruthenium Red* ; Ruthenium*

N-acetyl-L-cysteine (NAC) and cysteine have been implicated in a number of human neutrophils' functional responses. However, though Ca²⁺ signaling is one of the key signalings contributing to the functional responses of human neutrophils, effects of NAC and cysteine on intracellular calcium concentration ([Ca²⁺]ᵢ) in human neutrophils have not been investigated yet. Thus, this study was carried out with an objective to investigate the effects of NAC and cysteine on [Ca²⁺]ᵢ in human neutrophils. We observed that NAC (1 µM ~ 1 mM) and cysteine (10 µM ~ 1 mM) increased [Ca²⁺]ᵢ in human neutrophils in a concentration-dependent manner. In NAC pre-supplmented buffer, an additive effect on N-formyl-methionine-leucine-phenylalanine (fMLP)-induced increase in [Ca²⁺]ᵢ in human neutrophils was observed. In Ca²⁺-free buffer, NAC- and cysteine-induced [Ca²⁺]ᵢ increase in human neutrophils completely disappeared, suggesting that NAC- and cysteine-mediated increase in [Ca²⁺]ᵢ in human neutrophils occur through Ca²⁺ influx. NAC- and cysteine-induced [Ca²⁺]ᵢ increase was effectively inhibited by calcium channel inhibitors SKF96365 (10 µM) and ruthenium red (20 µM). In Na⁺-free HEPES, both NAC and cysteine induced a marked increase in [Ca²⁺]ᵢ in human neutrophils, arguing against the possibility that Na⁺-dependent intracellular uptake of NAC and cysteine is necessary for their [Ca²⁺]ᵢ increasing activity. Our results show that NAC and cysteine induce [Ca²⁺]ᵢ increase through Ca²⁺ influx in human neutrophils via SKF96365- and ruthenium red-dependent way.
Acetylcysteine* ; Calcium Channels ; Calcium* ; Cysteine* ; HEPES ; Humans* ; Neutrophils* ; Ruthenium ; Ruthenium Red

The distribution of glycosaminoglycans in Bruch`s membrane and sensory retina of human eye was evaluated with histochemical electron microscope using 0.05%ruthenium red (RR). Seven donated or enucleated eyeballs without chorioretinal disease were used.Electron-dense RR reactive materials were identified as fine discrete granules along the basement membrane of the retinal pigment epithelium, in the inner and outer collagen layer and central elastic layer of Bruch 's membrane, and in the interphotorecep- tor matrix. In addition, they were observed at the cytoplasmic wall of photoreceptor cells, their intercellular spaces, and the internal limiting membrane. These results demonstrated that GAGs are widely distributed within the interphotoreceptor matrix, Bruch`s membrane, and intercellular spaces of retinal neurons. They are considered to function as a mechanical supporter as well as a selective filtration barrier through which the metabolitespass.
Basement Membrane ; Collagen ; Cytoplasm ; Extracellular Space ; Filtration ; Glycosaminoglycans* ; Humans ; Membranes* ; Photoreceptor Cells ; Retina* ; Retinal Neurons ; Retinal Pigment Epithelium ; Ruthenium Red* ; Ruthenium*

BACKGROUND: Moisturizers induce skin hydration and then increase flexibility and elasticity, making the skin soft and smooth, and protecting it against environmental stimuli. OBJECTIVE: The aim of this work was to study the role of intercorneocyte lipid layers and desmosomes in the mechanism of moisturization. METHODS: Transepidermal water loss (TEWL) and capacitance were measured and the morphologic changes of the intercorneocyte lipid layers and desmosomes with electron microscopy, using ruthenium tetroxide (RuO4) postfixation, following the application of glycerin, propylene glycol, and a mixture of glycerin and propylene glycol for a 2 hour period to the epidermis of hairless mice were measured. RESULTS: 1. The TEWL was significantly increased in all three groups; glycerin, propylene glycol, and mixture of glycerin and propylene glycol. The increase of TEWL after the application of glycerin was maintained from the second to the forth hour after application which was statistically significant, after the application of propylene glycol it was maintained for 5 hours, and after the application of a mixture of glycerin and propylene glycol, for 6 hours. 2. The capacitance also was increased in all three experimental groups, compared to the control group. However there was no statistical significance. 3. Light microscopic findings showed no specific changes in all three groups, compared to the control group. 4. Ultrastructural observation by electron microscope, using RuO4 postfixation, showed widening of the intercorneocyte lipid layers in all three groups. In contrast to glycerin in which the results showed detachment of the desmosomes without changes in the intercorneocyte lipid layers, propylene glycol showed interruption and undulation of the intercorneocyte lipid layers and expansion of the lacunae spaces. A mixture of glycerin and propylene glycol showed interruption and undulation of the intercorneocyte lipid layers, detachment of the desmosomes, and, partial, formation of lacunae. CONCLUSION: These results suggest that the moisturizing effects of glycerin result from an increased detachment of the desmosomes and widening of the intercorneocyte lipid layers and then an increase in the water holding capacity of the stratum corneum. Propylene glycol, a chemical penetration enhancer, induce widening, interruption, and undulation of the lipid layers and expansion of the lacunae space. In the mixture of glycerin and propylene glycol, propylene glycol potentiate and continue the moisturizing effects of the glycerin.
Animals ; Desmosomes* ; Elasticity ; Epidermis ; Glycerol* ; Mice ; Mice, Hairless ; Microscopy, Electron ; Pliability ; Propylene Glycol* ; Ruthenium ; Skin

BACKGROUND: Bentonite clay, which is a major component of mud pack, has been used for various purposes in cosmetics. Glycolic acid is known to be effective in the treatment of acne. Al-though those products are used widely, information on the mode of action and effects on the skin are little and controversial till now. OBJECTIVE: To investigate whether bentonite alone, or bentonite with glycolic acid in mixed formulation affect the stratum corneum leading to alteration on cutaneous barrier function and whether those products alter the lipid lamellae and desmosomes of corneocytes. MATERIALS AND METHODS: Mud pack-type ointment of bentonite, bentonite and 5% glycolic acid formulation, bentonite and 10% glycolic acid formulation were applied on the volar fore-arm of the five healthy men and flank skin of five 6-8 week old hairless mice. Transepidermal water loss and capacitance were measured. Electron microscopic examination after ruthenium tetroxide postfixation was performed on the flank skin of the mice. RESULTS: Transepidermal water loss(TEWL) increased immediately and normalized 4 to 6 hours later after removal of vapor permeable membrane in both mouse and human. Capacitance did not show any evidence of change in the water content of the stratum corneum. Electron microscopic examination revealed that lipid lamellae and desmosome of corneocytes were not de-graded, but lamellar body secretion and partially electron-lucent material was-increased in 10% glycolic acid and bentonite mixture-treated area. CONCLUSION: Barrier function of stratum corneum is not disturbed by bentonite and glycolic acid formulations at the concentration used. Barrier structures are not disrupted, but lamellar body secretion and partially electron-lucent material was increased by bentonite and glycolic acid formulations at higher concentration.
Acne Vulgaris ; Animals ; Bentonite* ; Desmosomes ; Humans ; Male ; Membranes ; Mice ; Mice, Hairless ; Mud Therapy ; Ruthenium ; Skin ; Water

BACKGROUND: Acetone disrupts the cutaneous permeability barrier through the removal of stratum corneum lipids. This pertubation of barrier integrity stimulates a variety of homeostatic repair that ultimately results in the normalization of barrier function. OBJECT: To measure the effect of steroid on the barrier recovery of acetone applied skin. MATERIAL AND METHODS: The flank skin of 8~10 week old hairless mice was treated with acetone and then topical and systemic steroids were applied. Transepidermal water loss(TEWL) was checked after 0, 3, 6, 12 and 24 hours. Electron and light microscopic examination and ion capture cytochemistry were performed after 3, 6, 12 and 24 hours after systemic and topical steroids had been applied. RESULTS: The results were as follows ; 1) During 3~6 hours after experiment, the recovery rate of TEWL was most prominent in the group of acetone applied animal than other groups. 2) After 12 hours after acetone applied, formation of new stratum corneum was found in the groups of acetone applied or acetone applied skin with topical steroid application. But loss of stratum corneum was observed in the groups of high or low dose steroid injection. 3) Ruthenium tetroxide staining of acetone alone or topical steroid treated specimens after 12 hours experiment revealed that the lipid bilayer was partly impaired and fragmented. Intercellular spaces were widening and the lipid bilayer disappeared or was damaged in the groups of high or low dose steroid injection. 4) Six hours after acetone application, pattern of calcium distribution had been partially reestabilished in the group of acetone alone or topical steroid treated animals. But calcium content was still sparse and decreased from the stratum granulosum to basale in the groups of high or low dose steroid injection. CONCLUSION: In summary the present study demonstrates that steroid treatment acutely delays recovery rate of TEWL, inhibits normalization of calcium gradient or epidermal lipid synthesis that leads to abnormalities in permeability barrier homeostasis.
Acetone ; Animals ; Calcium ; Extracellular Space ; Histocytochemistry ; Homeostasis ; Lipid Bilayers ; Mice ; Mice, Hairless* ; Permeability ; Ruthenium ; Skin ; Steroids

BACKGROUND: Elecsys 2010 immunoassay system is based on the electrochemiluminescence immunoassay using a ruthenium (II) tris (bipyridyl) label. Since it was the first time to use the system in our laboratory, we would like to evaluate the analytical performances (precision, linearity and recovery rate) and correlation with radioimmunoassay (RIA) and microparticle enzyme immunoassay (MEIA) methods. METHODS: We used precicontrol tumor marker (TM1, TM2) for alpha-fetoprotein (AFP), prostatic specific antigen (PSA) and carcinoembryonic antigen (CEA), Precicontrol universal (Ul, U2) for triiodothyronine (T3) and thyroxine (T4), Precicontrol-TSH for thyrotropin (TSH) and pooled serum for the evaluation of precision and recovery rate. Patients' sera were used for the linearity and comparison study. RESULTS: The coefficients of variatron of Imprecision study were below; 4.0%, 8.7% and 10.2%, respectively in the within-run, within-day and between-day analysis. The recovery rates were 100.5%, 96.1% and 102.5%, respectively in T4, TSH, and AFP. The linearity were y=1.02x-0.182(r=0.99) for T4, y=1.01x+0.12 (r=0.99) for TSH and y=1.01x+0.54(r=1.00) for AFP. T3, T4, TSH, CEA and PSA results showed good correlation with RIA (r>0.90), but AFP showed r=0.88. Also, AFP, CEA and PSA results showed excellent correlation with AxSYM (r>0.99). CONCLUSION: Elecsys 2010 immunoassay system showed excellent precision, recovery rate, clinically acceptable linearity and good correlation with the results obtained by RIA and MEIA methods.
alpha-Fetoproteins ; Carcinoembryonic Antigen ; Immunoassay* ; Immunoenzyme Techniques ; Radioimmunoassay ; Ruthenium ; Thyrotropin ; Thyroxine ; Triiodothyronine

This study was performed to evaluate the histochemical changes of the conjunctival epithelial cell in eyes with dry eye syndrome. The authors selected 10 eyes clinically diagnosed as dry eye syndrome and 2 eyes which were enucleated due to choroidal melanoma as control group. The conjunctiva was stained with ruthenium red to enhance the detection of glycoprotein in conjunctival epithelium and observed by electron microscopy. The conjunctiva with dry eye syndrome showed the epithelial stratification, elon gation of the epithelial cells, reduction of microplicae in number, and irregular distribution of mucin. These results suggest that the mucin in the conjunctiva with dry eye syndrome may be abnormal in function and may lead to clumping.
Choroid ; Conjunctiva ; Dry Eye Syndromes* ; Epithelial Cells ; Epithelium ; Glycoproteins ; Melanoma ; Microscopy, Electron* ; Mucins ; Ruthenium Red

BACKGROUND: Topical irritants disrupt the cutaneous permeability barrier through the removal of stratum comeum lipids. This perturbation of barrier integrity stimulates a variety of homeostatic repair responses that ultimately result in the normalization of bamer function. Object To measure the effect of desoxymethasone ointment, vaseline and hydrobase on the barrier recovery of benzalkonium chloride (BKC) imtated skin. MATERIALS AND METHODS: The left flank skin of 2-3 monthold hairless mice was treated with BKC and then desoxymethasone ointment, vaseline and hydrobase were applied. Transepidermal water loss (TEWL) was checked after 0, 3, 6, 9, 12, 15, 18, 21 and 24 hours. Electron microscopic examination was performed after 3 and 24 hours after desoxymethasone, vaseline and hydrobase had been applied. RESULTS: The recovery of TEWL was most prominantly observed in the desoxymethasone ointment treated group followed by vaseline and hydrobase. Electron microscopic examination using ruthenium tetroxide fixation revealed that secretion and numbers of lamellar bodies and complete formatice of lipid bilayers were most prominent at desoxymethasone ointment and vaseline treated group. CONCLUSION: Desoxymethasone ointment, vaseline and hydrobase can be good agents in improving bamer recovery after exposure to irritant material.
Animals ; Benzalkonium Compounds* ; Desoximetasone* ; Irritants ; Lipid Bilayers ; Mice ; Mice, Hairless* ; Permeability ; Petrolatum* ; Ruthenium ; Skin*

BACKGROUND: The epidermal permeability barrier necessary for terrestrial life resides in the intercellular spaces of the stratum corneum and is composed of lipids. OBJECTIVE: Since strrtum corneum lipid may be important for the permeability barrier, we studied the differences and effects of experimentally altered barrier function using acetone and tape-stripping technique. METHODS: The permeability barrier of hairless mouse was disrupted by tape-stripping and acetone treatment and the recovery rate was assessed by histochemical staining, electron microscopic examination and lipid analysis. RESULTS: Although the transepidermal water loss recovered completely by 48 hours in both of the acute models, acetone treated samples seem to have on over-all better recovery rate than tape-stripped samples. The return of barrier function to normal in both tape-stripped and acetone-treated skin was accompanied by a comparable return of normal nile red and ruthenium tetroxide staining. The amount of lipid in stratum corneum paralleled both the return of barrier function towards normal and the extent of prior damage to the barrier in acetone treated skin, yet, the lipid synthesis in tape-stripped skin showed a slower return of lipid content. CONCLUSION: The difference in the recovery rate of the two acute models may be due to the fact that acetone mainly extracts intercellular lipids, whereas, tape-stripping has a prolonged effect by removal of comeocyte in addition to the intercellular lipids. This shows the importance of comeocytes as well as the intercellular lipid bilayer in the recovery of normal barrier function.
Acetone* ; Animals ; Extracellular Space ; Lipid Bilayers ; Mice ; Mice, Hairless ; Permeability ; Ruthenium ; Skin ; Water