PURPOSE: To investigate the effects of dipyridamole (DPD) on the production of reactive oxygen species (ROS) and oxidative stress in cultured human trabecular meshwork cells (HTMC). METHODS: Antioxidant activity of DPD was determined by DPPH assay. Primarily cultured HTMC were exposed to 0, 20, and 50 microm DPD using serum-deprived media. The effect of DPD on the production of ROS was assessed with the DCHFDA assay. The effect of DPD on the t-butyl hydroperoxide (tBHP)-induced oxidative stress was assessed with resazurin assay. RESULTS: DPD showed significant antioxidant activity. DPD significantly decreased the production of ROS (p < 0.05) and improved cellular activity significantly after treatment with t-BHP (p < 0.05). DPD did not affect the generation of nitric oxides. CONCLUSIONS: DPD suppressed the formation of ROS and possessed cytoprotective activity against the oxidative stress in HTMC.
Dipyridamole ; Humans ; Oxazines ; Oxidative Stress ; Reactive Oxygen Species ; tert-Butylhydroperoxide ; Trabecular Meshwork ; Xanthenes

PURPOSE: To evaluate the protective effect of creatine on the survival of retinal ganglion cells after serum deprivation. METHODS: RGC-5 cells were exposed to 5 mM creatine with serum-free media for 4 days. Cellular survival and mitochondrial respiratory activity were measured with MTT assay and resazurin assay, respectively. Degree of apoptosis was evaluated with vital staining using acridine orange/Hoechest 33342 and flow cytometric analysis using annexin/PI, respectively. RESULTS: Creatine increased cellular survival of RGC-5 cells significantly after serum deprivation. Additionally, creatine increased mitochondrial respiratory activity and inhibited apoptosis of RGC-5 cells. CONCLUSIONS: The energy precursor creatine increased survival of retinal ganglion cells after serum deprivation. Creatine could be relevant for the cytoprotection of retinal ganglion cells.
Apoptosis ; Creatine ; Culture Media, Serum-Free ; Cytoprotection ; Oxazines ; Retinal Ganglion Cells ; Xanthenes

BACKGROUND: Cellulite is a 'cottage cheese-like' cutaneous change caused by subcutaneous fat bulging into the dermis that usually leads to cosmetic problems. Slimming cream containing 3.5% water-soluble caffeine and xanthenes exhibits a lipolytic effect with penetration into the dermis. OBJECTIVE: To evaluate the efficacy and safety of slimming cream for the treatment of cellulite. METHODS: Fifteen subjects with cellulite applied slimming cream to the thighs and inner side of the upper arms twice daily for 6 weeks. Efficacy was assessed using a standard visual scale, changes in the circumferences of the thighs and upper arms, and patient satisfaction by a questionnaire at baseline, week 3, and week 6. Safety was assessed by inquiring about adverse events through questionnaires. RESULTS: The standard visual scale score improved significantly by 0.49 points (19.8%) at week 6. Thigh and upper-arm circumferences decreased by 0.7 cm (1.7%) and 0.8 cm (2.3%), respectively, at week 6. Slight itching and transient flushing were commonly reported, but no serious adverse event occurred. CONCLUSION: The slimming cream tested appears to be effective for the treatment of cellulitis without serious adverse effects. However, additional large clinical trials are required to confirm the efficacy and safety of slimming cream for the treatment of cellulitis.
Arm ; Caffeine* ; Cellulitis ; Dermis ; Flushing ; Patient Satisfaction ; Pruritus ; Subcutaneous Fat ; Thigh ; Xanthenes* ; Surveys and Questionnaires

OBJECTIVETo study the spatial distribution of free Ca2+ in osteoclast-like cells cultured on glass.

METHODSTo detect the free Ca2+ in osteoclast-like cells, the images were analyzed with image software, using the laser scanning confocal microscope and fluorescent probe.

RESULTSAt 37 degrees C the free Ca2+ in osteoclast-like cells could be labelled effectively with 10 micromol/L Fluo-3/AM, the intensity of Ca2+ fluorescent signal in the central part was greater than that in the peripheral part and in the same section the signal was not distributed evenly.

CONCLUSIONThe intensity of Ca2+ fluorescent signal is different among various organellae in osteoclast-like cell, which suggests the osteoclast-like cell modulate its own function through the spatial difference of free Ca2+ concentration.

Aniline Compounds ; Calcium ; Cell Line ; Cells, Cultured ; Microscopy, Confocal ; Osteoclasts ; Xanthenes

OBJECTIVETo observe the effects of 17beta-estradiol on the intracellular calcium of masticatory muscles myoblast.

METHODSMyoblasts from maxillofacial skeletal muscle of one week old female Sprague-Dawley rats were cultured. Fluo-4-AM as the Ca2+ indicator and the laser confocal microscope system were used to observe the effects of estrogen on the cytoplasmic Ca2+ concentration in the normal pH condition and the acid condition (pH = 6.7).

RESULTSIn the normal pH condition, when 17beta-estradiol (10(-9), 10(-8), 10(-7) mol/L) were added to cells cytoplasmic Ca2+ immediately increased then decreased right away, and in the end came into a new Ca2+ homeostasis in the base line. In the acid condition, 17beta-estradiol (10(-9), 10(-8), 10(-7) mol/L) made the cytoplasmic Ca2+ decreased immediately then came into a new Ca2+ homeostasis under the base line.

CONCLUSIONThe results suggest that estrogen may maintain the skeletal cytoplasmic Ca2+ concentration in a lower level and reduce the cytoplasmic Ca2+ accumulation to keep the normal functions of masticatory muscles myoblast.

Aniline Compounds ; Animals ; Calcium ; Estradiol ; Female ; In Vitro Techniques ; Masticatory Muscles ; Myoblasts ; Rats ; Rats, Sprague-Dawley ; Xanthenes

OBJECTIVETo study the change of Ca2+ density in cultured osteoclast-like cells in response to fluid shear stress.

METHODSLaser scanning confocal microscope and fluorescent probe were used to detect the free Ca2+ in osteoclast-like cells before and after undergoing fluid shear stress. The images were analyzed and compared with image software.

RESULTSAt 37 degrees C the free Ca2+ in osteoclast-like cells could be labelled effectively with 10 micromol/L Fluo-3/AM. Compared with contol group, the average intensity of Ca2+ fluorescent signal in osteoclast-like cells undergoing fluid shear stress increased significantly.

CONCLUSIONThe Cal2+ concentration in bone-marrow derived osteoclast-like cells is sensitive to fluid shear stress, which suggests osteoclast-like cells modulate their function in response to fluid shear stress through the change of free Ca2+ concentration.

Aniline Compounds ; Bone Marrow ; Bone Marrow Cells ; Calcium ; Cells, Cultured ; Osteoclasts ; Stress, Mechanical ; Xanthenes

BACKGROUND: Clostridium difficile is the most common pathogen of antibiotic-associated diarrhea. Toxigenic strains produce toxin A and toxin B. The pathogenicity of C. difficile is due to the production of these two exotoxins. This study aimed to evaluate diagnostic value of two enzyme immunoassay by comparison of concordance rate to diagnose C. difficile-associated infection. METHODS: C. DIFF QUIK CHEK (TECHLAB, USA) that detect glutamate dehydrogenase antigen and VIDAS C. difficile Toxin A&B (BioMerieux, France) that detect toxin A and toxin B were done in 122 fecal specimens to detect C. difficile. RESULTS: In the total 122 stool specimens, 17 cases showed positive results in both tests. One specimen showed discrepancy that positive result in VIDAS C. difficile Toxin A&B (relative fluorescence value, RFV=2.93) but negative result in C. DIFF QUIK CHEK. Therefore, the concordance rate between two tests was 95.1% (116/122). Both anaerobic culture and in-house PCR for toxin B were negative in the discrepant fecal specimen and there was no clinical evidence that support C. difficile-associated diarrhea, so we concluded result in VIDAS C. difficile Toxin A&B as false positive. CONCLUSIONS: Although these two enzyme immunoassays targeted different antigen, they showed high concordance rate. The discrepant case was concluded to false positive in VIDAS C. difficile Toxin A&B test because it showed negative results in culture and PCR for toxin B and there were no clinical evidences of C. difficile-associated infection. It could be needed for analysis about conditions that cause false positive result in enzyme immunoassays to detect C. difficile toxin.
Azure Stains ; Clostridium difficile ; Diarrhea ; Exotoxins ; Fluorescence ; Glutamate Dehydrogenase ; Immunoenzyme Techniques ; Methylene Blue ; Polymerase Chain Reaction ; Xanthenes

AIMTo assess that metabolic status of spermatozoa could provide a useful tool for evaluation of semen quality.

METHODSThe accuracy of the spectrophotometric application of the resazurin reduction assay was assessed using receiver operating characteristic (ROC) analysis.

RESULTSAreas under ROC curves (AUC) for motile sperm concentration and sperm index (SI) (sperm concentration multiplied by the square root of percentage sperm motility multiplied by the percentage normal sperm morphology) were 0.922. The best discrimination between poor and good semen samples according to the SI was achieved at a cut-off point of A610 = 0.209, where high sensitivity (94.1%) and specificity (91.7%) were calculated. The assay was less accurate when motile sperm concentration was used as the criterion value, yielding sensitivity of 88.2% and specificity of 87.5%, respectively. Likelihood ratios (LR) indicate that absorbances lower than 0.209 were at least 11.3 times as likely to be found in good semen samples than those in poor according to the SI, whereas in the case of motile sperm concentration, the LR was calculated to be 7.06.

CONCLUSIONThese results show that the resazurin reduction assay combined with spectrophotometry is an accurate method of assessing the quality of boar semen.

Animal Husbandry ; Animals ; Area Under Curve ; Male ; Oxazines ; pharmacokinetics ; pharmacology ; Semen ; physiology ; Sperm Motility ; Swine ; Xanthenes ; pharmacokinetics ; pharmacology

PURPOSE: The aim of this in vitro study was to investigate the adhesion of initial colonizer, Streptococcus sanguis, on resin, titanium and zirconia under the same surface polishing condition. MATERIALS AND METHODS: Specimens were prepared from Z-250, cp-Ti and 3Y-TZP and polished with 1 microm diamond paste. After coating with saliva, each specimen was incubated with Streptococcus sanguis. Scanning electron microscope, crystal violet staining and measurement of fluorescence intensity resulting from resazurin reduction were performed for quantifying the bacterial adhesion. RESULTS: Surface of resin composite was significantly rougher than that of titanium and zirconia, although all tested specimens are classified as smooth. The resin specimens showed lower value of contact angle compared with titanium and zirconia specimens, and had hydrophilic surfaces. The result of scanning electron microscopy demonstrated that bound bacteria were more abundant on resin in comparison with titanium and zirconia. When total biofilm mass determined by crystal violet, absorbance value of resin was significantly higher than that of titanium or zirconia. The result of relative fluorescence intensities also demonstrated that the highest fluorescence intensity was found on the surface of resin. Absorbance value and fluorescence intensity on titanium was not significantly different from those on zirconia. CONCLUSION: Resin specimens showed the roughest surface and have a significantly higher susceptibility to adhere Streptococcus sanguis than titanium and zirconia when surfaces of each specimen were polished under same condition. There was no significant difference in bacteria adhesion between titanium and zirconia in vitro.
Bacteria ; Bacterial Adhesion ; Biofilms ; Colon ; Diamond ; Electrons ; Fluorescence ; Gentian Violet ; Microscopy, Electron, Scanning ; Oxazines ; Saliva ; Streptococcus sanguis ; Titanium ; Xanthenes ; Zirconium

We prepared protoplasts from Salvia miltiorrhiza Bunge suspension culture cells. Then, the protoplasts' vitality and functions were tested by fluorescein diacetate staining method and Fluo-3/AM flourescent probe. The optimal condition of protoplast isolation was Cellulase R-10 1.5%, Pectinase Y-23 0.3%, Macerozyme R-10 0.5%, 40 r/min 12 h, 600 r/min 5 min, and the protoplasts yield was 1.1x10(6) cells/g FW, the vitality was more than 95% by using fluorescein diacetate staining method. It has been confirmed that calcium fluorescent probe Fluo-3/AM can be successfully loaded into protoplasts.
Aniline Compounds ; chemistry ; Cell Culture Techniques ; Cellulase ; chemistry ; Fluorescent Dyes ; chemistry ; Protoplasts ; chemistry ; Salvia miltiorrhiza ; growth & development ; Xanthenes ; chemistry