Evaluation of tear production in juvenile opossum using three different methods / Avaliação da produção lacrimal em gambás filhotes por três diferentes métodos

The establishment of parameters for tear production in different species is important for better understanding eye´s health and is one of the components of the ophthalmic semiological technique. Particularities derived from the anatomophysiology of non-domestic species induce the search for more reliable methodologies. The aim was to evaluate and compare tear production of white-eared opossum (Didelphis albiventris) and Brazilian common opossum (Didelphis aurita) by three different methods. Fifteen individuals of each species, juveniles, healthy, of both sexes, with 60 to 90 days of life, were physically restrained. Phenol red thread test (PRTT), endodontic absorbent paper point tear test (EAPPTT) and modified -Schirmer tear test (mSTT) were performed. PRTT was the most difficult to perform because of the wire malleability, while EAPPTT was more feasible for both species. The median ± semi-quartile range for PRTT were 19.79±2.61mm/15 "and 5.22±2.92mm/15", for EAPPTT were 16.25±1.82mm/min and 10.9±3.04mm/min, and for STTm were 0±1.63mm/min and 0±1.63mm/min for white-eared opossum and Brazilian common opossum respectively. There was no difference between the right and left eye neither sex. A significant difference was obtained for the same test to different species. No significant correlation was found between the tests for both species. The description of tear production parameters for juvenile white-eared opossum and Brazilian common opossum may be used as a tool, which will allow the early diagnosis of ocular diseases.(AU)

O estabelecimento do parâmetro de produção lacrimal nas diferentes espécies é importante para o entendimento da saúde do olho e é um dos componentes da semiotécnica oftálmica. Particularidades derivadas da anatomofisiologia das espécies não domésticas induzem a busca de metodologias que sejam mais fidedignas aos parâmetros. Objetivou-se com este estudo avaliar e comparar a produção lacrimal de gambás-de-orelha-branca (Didelphis albiventris) e gambás-de-orelha-preta (Didelphis aurita) por três diferentes métodos. Quinze indivíduos de cada espécie, juvenis, hígidos, de ambos os sexos, com 60 a 90 dias de vida, foram contidos fisicamente para realização do teste lacrimal do vermelho de fenol (TLVF), da ponta de papel absorvente estéril e do teste lacrimal de Schirmer modificado (TLSm). O TLVF foi o mais difícil de ser executado devido à maleabilidade do fio, enquanto a TEPA se mostrou mais exequível para ambas as espécies. A mediana ± intervalo semi-interquartil para o TLVF foi de 19,79±2,61mm/15" e 5,22±2,92mm/15", para a TEPA foram de 16,25±1,82mm/min e 10,93±3,04mm/min, e para o TLSm foram de 0±1,63mm/min e 0±1,63mm/min, para gambás-de-orelha-branca e gambás-de-orelha-preta, respectivamente. Não houve diferença entre o olho direito e esquerdo e nem quanto ao sexo. Obteve-se diferença significativa para um mesmo teste entre as espécies. Não foi encontrada correlação significativa entre os testes para ambas as espécies. A quantificação da porção aquosa da lágrima poderá auxiliar no diagnóstico precoce de doenças oculares nas espécies estudadas.(AU)

Método rápido para la detección de la sensibilidad a cefotaxima en enterobacterias / Rapid test for detection of susceptibility to cefotaxime in Enterobacteriaceae

En este trabajo se evalúa una prueba rápida in house para la detección de enterobacterias sensibles a cefotaxima, basada en el cambio de pH del rojo fenol debido a la hidrólisis de este antibiótico. Las cepas de enterobacterias procedentes de 1.947 urocultivos se evaluaron mediante los paneles MicroScan y esta prueba in house. Mediante los paneles de MicroScan se estudiaron 499 aislados de enterobacterias, entre los cuales había 27 aislados de Escherichia coli productora de β-lactamasa de espectro extendido (BLEE), 16 de Klebsiella pneumoniae BLEE y una de Klebsiella oxytoca BLEE. La prueba in house mostró una sensibilidad del 98% y una especificidad del 97%, con un valor predictivo negativo del 100% y un valor predictivo positivo del 78%. La prueba in house basada en el cambio de pH es útil en nuestro medio para detectar presuntivamente de forma rápida cepas de enterobacterias con cierta resistencia a cefotaxima.

In this work an "in house" rapid test based on the change in pH that is due to hydrolysis for detecting Enterobacteriaceae susceptible to cefotaxime is evaluated. The strains of Enterobacteriaceae from 1947 urine cultures were assessed using MicroScan panels and the "in house" test. This rapid test includes red phenol solution and cefotaxime. Using MicroScan panels, 499 Enterobacteriaceae isolates were evaluated, which included 27 isolates of Escherichia coli producing extended-spectrum beta-lactamases (ESBL), 16 isolates of Klebsiella pneumoniae ESBL and 1 isolate of Klebsiella oxytoca ESBL. The "in house" test offers the following values: sensitivity 98% and specificity 97%, with negative predictive value 100% and positive predictive value 78%. The "in house" test based on the change of pH is useful in our area for detecting presumptively cefotaxime-resistant Enterobacteriaceae strains.

Rat prostate glandular epithelial cells cultured in vitro and their barrier function / 中华男科学杂志

<p><b>OBJECTIVE</b>To culture rat prostate glandular epithelial cells and study their barrier functions in vitro.</p><p><b>METHODS</b>Rat prostate glandular epithelial cells were cultured in vitro. The expression of the tight junction protein claudin-1 was determined by immunohistochemistry, the structure and composition of the epithelial cells observed under the inverted microscope and transmission electron microscope. The transepithelial electrical resistances (TEERs) were monitored with the Millicell system. The permeability of the prostate glandular epithelial cells was assessed by the phenol red leakage test.</p><p><b>RESULTS</b>Compact monolayer cell structures were formed in the prostate glandular epithelial cells cultured in vitro. Immunohistochemistry showed the expression of the tight junction protein claudin-1 and transmission electron microscopy confirmed the formation of tight junctions between the adjacent glandular epithelial cells. The TEERs in the cultured prostate glandular epithelial cells reached the peak of about (201.3 ± 3.5) Ω/cm2 on the 8th day. The phenol red leakage test manifested a decreased permeability of the cell layers with the increase of TEERs.</p><p><b>CONCLUSION</b>The structure and function of rat prostate glandular epithelial cells are similar to those of brain capillary endothelial cells, retinal capillary endothelial cells, and intestinal epithelial cells. In vitro cultured prostate glandular epithelial cells have the barrier function and can be used as a model for the study of blood prostate barrier in vitro.</p>

The effect of progesterone and 17-β estradiol on membrane-bound HLA-G in adipose derived stem cells

Membrane-bound HLA-G (mHLA-G) discovery on adipose derived stem cells (ADSCs) as a tolerogenic and immunosuppressive molecule was very important. Many documents have shown that HLA-G expression can be controlled via some hormones such as progesterone (P4) and estradiol (E2). Therefore, this study was designed to evaluate progesterone and estradiol effects on mHLA-G in ADSCs at restricted and combination concentrations. Three independent cell lines were cultured in complete free phenol red DMEM and subcultured to achieve suffi cient cells. These cells were treated with P4, E2 and P4 plus E2 at physiologic and pregnancy concentrations for 3 days in cell culture conditions. The HLA-G positive ADSCs was measured via monoclonal anti HLA-G-FITC/MEMG-09 by means of flow cytometry in nine groups. Data were analyzed by one way ANOVA and Tukey's post hoc tests. There were no signifi cant values of the mean percentage of HLA-G positive cells in E2-treated and the combination of P4 plus E2-treated ADSCs compared to control cells (p value>0.05) but P4 had a signifi cant increase on mHLA-G in ADSCs (p value<0.05). High P4 concentration increased mHLA-G but E2 and the combination of P4 plus E2 could not change mHLA-G on ADSCs.

Evidence for the involvement of peripheral -adrenoceptors in delayed liquid gastric emptying induced by dipyrone, 4-aminoantipyrine, and antipyrine in rats

Dipyrone (Dp), 4-aminoantipyrine (AA), and antipyrine (At) delay liquid gastric emptying (GE) in rats. We evaluated adrenergic participation in this phenomenon in a study in male Wistar rats (250-300 g) pretreated subcutaneously with guanethidine (GUA), 100 mg·kg−1·day−1, or vehicle (V) for 2 days before experimental treatments. Other groups of animals were pretreated intravenously (iv) 15 min before treatment with V, prazosin (PRA; 1 mg/kg), yohimbine (YOH; 3 mg/kg), or propranolol (PRO; 4 mg/kg), or with intracerebroventricular (icv) administration of 25 µg PRO or V. The groups were treated iv with saline or with 240 µmol/kg Dp, AA, or At. GE was determined 10 min later by measuring the percentage of gastric retention (%GR) of saline labeled with phenol red 10 min after gavage. %GR (mean±SE, n=8) indicated that GUA abolished the effect of Dp (GUA vs V=31.7±1.6 vs 47.1±2.3%) and of At (33.2±2.3 vs 54.7±3.6%) on GE and significantly reduced the effect of AA (48.1±3.2 vs 67.2±3.1%). PRA and YOH did not modify the effect of the drugs. %GR (mean±SE, n=8) indicated that iv, but not icv, PRO abolished the effect of Dp (PRO vs V=29.1±1.7 vs 46.9±2.7%) and At (30.5±1.7 vs 49±3.2%) and significantly reduced the effect of AA (48.4±2.6 vs 59.5±3.1%). These data suggest activation of peripheral β-adrenoceptors in the delayed GE induced by phenylpyrazolone derivatives.

Valuation on analgesic, expectorant and antitussive effects of compatible use of Aconiti radix cocta and Fritillaria cirrhosa or Fritillaria thunbergii / 中国中药杂志

<p><b>OBJECTIVE</b>To study the analgesic, expectorant and antitussive effects of the compatible use of Aconiti Radix Cocta and Fritillaria cirrhosa or F. thunbergii with different matching ratio or dose in mice.</p><p><b>METHOD</b>The two-factor, seven-level uniform design method was adopted to observe the analgesic, expectorant and antitussive effects of the oral administration with the two combined decoctions in rats, with frequency of body torsions induced by acetum, secretion of phenol red in tracheas and frequency of coughs as indexes. Significant matching proportions and doses were collected for verification.</p><p><b>RESULT</b>The effect on the frequency of body torsions: The combined decoctions could effectively reduce the frequency of body torsions. According to a regression analysis, Aconiti Radix Cocta and F. cirrhosa had the synergistic effect, which was maximized with a ratio of 1: 1. The 1: 1 combined decoction played the least role in reducing the frequency of body torsions with a total dose of more than 5 g x kg(-1). The effect on the secretion of phenol red in tracheas. The combined decoctions could effectively increase the secretion of phenol red in tracheas. According to a regression analysis, Aconiti Radix Cocta and F. thunbergii had the antagonism, which was maximized at the ratio of 1: 1, and minimized with a total dose of less than 10 g x kg(-1) and a ratio of 5: 1 between F. thunbergii and Aconiti Radix Cocta. The effect on the frequency of coughs. The combined decoctions could effectively reduce the frequency of coughs. According to a regression analysis, Aconiti Radix Cocta and F. cirrhosa had the antagonism, which was maximized at the ratio of more than 1: 5 and less than 10: 1. There was no interaction between Aconiti Radix Cocta and F. thunbergii. F. thunbergii could reduce the frequency of coughs, whereas Aconiti Radix Cocta showed no effect.</p><p><b>CONCLUSION</b>The compatible application of Aconiti Radix Cocta and F. cirrhosa could enhance the analgesic effect of Aconiti Radix Cocta and reduce the expectorant and antitussive effects of F. cirrhosa, which vary according to different matching ratio and dose. The compatible application of Aconiti Radix Cocta and F. thunbergii shows no effect on the antitussive effect of F. thunbergii. This study provides experimental basis for in-depth studies on the combined effect of Aconiti Radix Cocta and Fritillaria--two of eighteen incompatible pairs.</p>

Clinical Usefulness of the Phenol Red Thread Test as Diagnostic Tool in Dry Eye Patient

PURPOSE: To evaluate the clinical usefulness of the phenol red thread test as a diagnostic tool of dry eye by comparing the phenol red thread test, Schirmer's test and tear break-up time. METHODS: The present study included 30 dry eye patients belonging to dry eye workshop grade 1 or 2 and 25 normal subjects. Phenol red thread test, Schirmer's test, and tear break-up time were performed on each subject's right eye. The sensitivity, specificity and repeatability of each test were compared, and the correlations between the 3 tests were also analyzed. RESULTS: Tear break-up time was superior to the other tests in terms of sensitivity and repeatability. The phenol red thread test was better than Schirmer's test in terms of specificity and repeatability. In all 55 patients including dry eye patients and normal subjects, the phenol red thread test showed a greater correlation with tear break-up time than did Schirmer's test. In addition, in 25 dry eye patients, the correlation between the phenol red thread test and Schirmer's test increased significantly. CONCLUSIONS: The phenol red thread test is less irritating and requires a shorter testing time than Schirmer's test. Additionally, the phenol red thread test is superior to Schirmer's test in terms of specificity, repeatability, and relation to tear break-up time. In addition, the correlation between the phenol red thread test and Schirmer's test significantly increases in dry eye patients. Therefore, the phenol red thread test is a good substitute option for Schirmer's test in diagnosing dry eye.

A method for simultaneous assay of propulsion and absorption in small intestine / 浙江大学学报·医学版

<p><b>OBJECTIVE</b>To develop a method for simultaneous assay of propulsion and absorption in small intestine.</p><p><b>METHODS</b>The mice were administrated through gastric tube with mixed reagents containing 0.12% phenol red, D-xylose (1.25%, 2.5% and 5%) and 15% gelatin. The influence of phenol red on D-xylose absorption and the influence of D-xylose on small intestine propulsion rate were investigated by measuring serum concentration of D-xylose with phloroglucinol method.</p><p><b>RESULTS</b>At 10 min, no significant difference was found between 5% D-xylose mixed reagent group and 5% D-xylose control. At 15 min, small intestine propulsion rate in 5% D-xylose mixed reagent group, but not in 2.5% and 1.25% D-xylose mixed reagent groups, was significantly higher than in phenol red control (P<0.05).</p><p><b>CONCLUSION</b>Gastric administration of mixed reagent containing 0.12% phenol red, 5% D-xylose and 15% gelatin can simultaneously assay propulsion and absorption of small intestine in mice.</p>

Optimal Media Conditions for the Detection of Extracellular Cellulase Activity in Ganoderma neo-japonicum

To determine the optimal media conditions for the detection of the extracellular cellulase activity in Ganoderma neo-japonicum, we varied three media conditions: dye reagent, pH, and temperature. We evaluated the use of four dyes, Congo red, phenol red, remazol brilliant blue, and trypan blue. To observe the effect of pH on the chromogenic reaction, we tested media ranging from 4.5 to 8.0. To research the effect of temperature on the clear zone and the fungus growing zone, we tested temperatures ranging from 15 to 35degrees C. On the whole, the best protocol called for Ganoderma neo-japonicum transfer onto media containing Congo red with a pH of 7.0, followed by incubation at 25degrees C for 5 days. Our results will be useful to researchers who study extracellular enzyme activity in Ganoderma neo-japonicum.

Development of Detection Methods for Cellulolytic Activity of Auricularia auricula-judae

To obtain basic information on the detection of cellulolytic activity in Auricularia auricula-judae, the influences of dye reagent, pH, and temperature were assessed. Chromogenic dye (congo red, phenol red, remazol brilliant blue, and trypan blue) was individually incorporated into a medium containing either carboxymethyl-cellulose, Avicel, or D-cellobiose as a polysaccharide carbon substrate. The other assessments utilized pHs ranging from 4.5 to 8.0 and temperatures from 15~35degrees C. Overall, when A. auricula-judae species were transferred onto media contained Congo red and adjusted pH 7.0 and then incubated at 25degrees C for 5 days, the clear zone indicative of cellulolytic activity was more pronounced.

Optimal Medium Conditions for the Detection of Cellulolytic Activity in Ganoderma lucidum

To determine the optimal medium conditions for the detection of the cellulolytic activity in Ganoderma lucidum, we varied three media conditions: dye reagent, pH, and temperature. First, we evaluated the use of four dyes, Congo Red, Phenol Red, Remazol Brilliant Blue, and Trypan Blue. To observe the effect of pH on the chromogenic reaction, we also made and tested various media spanning acidic and alkaline pHs, ranging from 4.5 to 8.0. Furthermore, in order to research the effect of temperature on the clear zone and the fungus growing zone, we tested temperatures ranging from 15 to 35degrees C. On the whole, the best protocol called for Ganoderma lucidum transfer onto media containing Congo red with pH adjusted to 7.0, followed by incubation at 25degrees C for 5 days. Our results will be useful to researchers who aim to study extracellular enzyme activity in Ganoderma lucidum.

Sjögren-like syndrome after bone marrow transplantation.

OBJECTIVE: To study the incidence of dry eye in Sjögren-like syndrome, graft-versus-host disease (GVHD) in hematological patients undergoing bone marrow transplantation (BMT). MATERIAL AND METHOD: Prospective, cross-sectional study in twenty-six patients that were planned for BMT (group I). Twenty-nine patients undergoing BMT before study were classified as group II no GVHD (9), and group III with GVHD (20). Thirty-two normal subjects were controls. All subjects were examined by slit lamp biomicroscopy and had their tear samples analyzed about tear osmolarity. They were also evaluated for aqueous tear production by phenol red thread test, Schirmer test without anesthesia, tear film stability by tear break-up time (TBUT), and rose bengal staining 2 weeks before BMT (for group I) as well as 6 weeks, 3 months, and 6 months after BMT. The patients with GVHD were followed up 1 month later. Main outcome measures were amount of tear production, tear film stability, and dry eye symptoms. RESULTS: Average aqueous tear production in group III was less than control and group II (p < 0.001). Mean TBUT in group III was faster than control (p < 0.001) and group I before BMT (p = 0.001). Mean score of rose bengal staining in group III was more than control and group I before BMT (p < 0.001). Keratoconjunctivitis sicca and red eye developed in 27.5%, and 20% of group III, with incidence of dry eye by Schirmer test without anesthesia (67.5%). This compares with group II having incidence of dry eye of 16.7%. However, 42.3% of group I before BMT had dry eye compared with 9.4% in the controls (p < 0.001). CONCLUSION: Trend of dry eye in patients with BMT and GVHD were higher than no-GVHD group. Doctors should be aware of ocular symptoms and signs of dry eye in patients with BMT and follow-up for proper management.

Comparison of the 13C-urea breath test and the endoscopic phenol red mucosal pH test in the quantification of Helicobacter pylori infection loading

BACKGROUND/AIMS: The 13C-urea breath test (UBT) is a semiquantitative test for measuring Helicobacter pylori infection loading. H. pylori produces ammonia, which elevates the pH of the gastric mucosa and is detectable via endoscopy using a phenol red indicator. We evaluated whether this test could be used to diagnose H. pylori infection and whether phenol red staining was correlated with 13C-UBT results. METHODS: One hundred and twenty-three patients participated. The UBT was performed after ingestion of a capsule containing urea. A change in 13C-UBT >2 ppt was selected as the cutoff value for diagnosing infection. After spraying evenly with a 0.1% phenol red solution, the pH of the gastric mucosal surface was measured using an antimony electrode through the biopsy channel. RESULTS: The pH of stained mucosa (6.9+/-0.4) was significantly higher than that of unstained mucosa (1.9+/-0.8; p<0.001), and the H. pylori detection rate confirmed via histology was higher in stained versus unstained mucosa (p<0.01). Extensive mucosal staining resulted in a higher detection rate (p<0.001). The UBT produced results were very similar to those obtained via histological detection in stained mucosa (p<0.001). The extent of staining, expressed as a staining score, was positively correlated with the change in 13C-UBT (r=0.426, p<0.001). A significant correlation was also observed between the histologically determined H. pylori density and 13C-UBT results (r=0.674, p<0.001). CONCLUSIONS: H. pylori infection elevates gastric mucosal surface pH, and endoscopic phenol red staining may be an alternative method for the diagnosis of H. pylori infection.

Comparison of Dyes for Easy Detection of Extracellular Cellulases in Fungi

To evaluate which dye is effective in a plate assay for detecting extracellular cellulase activity produced by fungi, four chromogenic dyes including remazol brilliant blue, phenol red, congo red, and tryphan blue, were compared using chromagenic media. For the comparison, 19 fungal species belonging to three phyla, ascomycota, basidiomycota, and zygomycota were inoculated onto yeast nitrogen-based media containing different carbon substrates such as cellulose (carboxylmethyl and avicel types) and cellobiose labeled with each of the four dyes. Overall, the formation of clear zone on agar media resulting from the degradation of the substrates by the enzymes secreted from the test fungi was most apparent with media containing congo red. The detection frequency of cellulase activity was also most high on congo red-supplemented media. The results of this study showed that congo red is better dye than other three dyes in a plate assay for fungal enzyme detection.

Alternative methods for assessing experimental colitis in vivo and ex vivo

To develop and validate methods for the assessment of colonic damage in the rat that are rapid, simple to perform and quantitative. Rats [Sprague Dawley, male, N = 52] were administered 0.5 mL of 2,4,6-trinitrobenzenesulfonic acid [TNBS, 60 mg mL -1 in 50% ethanol] intracolonically to induce reproducible experimental colitis. The rats were administered either orally or intracolonically 1 g of the non-absorbable permeability marker phenol red in 0.5 mL of water 24-72 h post-colitis induction. Urine was collected for the next 24 h and analyzed for phenol red at 559 nm using a spectrophotometer. In separate studies, colonic tissue was excised 24-72 h post-colitis induction for mitochondrial DNA damage determination. Baseline permeability indicated that < 4% of phenol red dose was excreted in urine after oral or intracolonic administration to vehicle control rats. A 3-4 fold increase in colonic permeability was apparent in colitic rats 48 h post TNBS, which correlated with gross macroscopic ulceration in the large bowel and colonic mitochondrial damage. Administration of dexamethasone 2 mg kg -1, or 5-aminosalicylic acid [5-ASA] 100 mg kg -1 resulted in significant gross macroscopic protection and reduction of colonic permeability and mitochondrial damage. In a group of rats TNBS was administered and then allowed to recover for 6 weeks. Subsequent tail vein administration of TNBS 5 mg kg -1 for 3 days reactivated the disease, which was also detected by an increase in colonic permeability of phenol red and oxidative mitochondrial damage. Several non-steroidal anti-inflammatory drugs administered in this model also appeared to exacerbate this disease and increase colonic permeability and mitochondrial damage. Oral or rectal administration of phenol red in experimental colitis can reproducibly detect colonic damage. This marker can also be used to non-invasively evaluate potential treatments for experimental colitis as well as its reactivation. Mitochondrial damage to colonic tissue parallels the increase in colonic permeability and appears to be a sensitive marker of disease activity in experimental colitis

Detection of Cellulolytic Activity in Ophiostoma and Leptographium species by Chromogenic Reaction

To understand the ability of producing cellulolytic enzyme activity in the sapstaining fungi, four species of Ophiostoma and two species of Leptographium were investigated in the culture media containing each of cellulose substrates such as CM-cellulose, Avicel and D-cellobiose and each of chromogenic dyes such as Congo-Red, Phenol Red, Remazol Brilliant Blue and Tryphan Blue. When the fungi were grown for 5~7 days at 25degrees C, the formation of clear zone by chromogenic reaction around the margin of the fungal colony was demonstrated in all the culture media Congo-Red containing CM-cellulose. There was difference in the formation of clear zone among the dyes. Only Ophiostoma setosum and Leptographium spp. showed cellulolytic activity to the three substrates. Overall, the results of this study show that ophiostomatoid sapstaining fungi can produce cellulolytic enzymes.

Development of Luciferase Reporter Gene-based Cell Bioassay for the Aromatic Hydrocarbon Receptor Agonists

The aromatic hydrocarbon receptor (AhR) is a ligand-activated transcription factor that mediates many of the biological and toxicological effects of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD, dioxin) and related chemicals. The application of recombinant reporter plasmid such as the firefly luciferase gene has proven to be a very effective method to detect these chemicals. The bioassay system, CALUX, is sensitive in directly detecting AhR-agonists from a variety of environmental and biologic materials. However, responses of the AhR-dependent bioassays are dependent on the cell types used. Thus, we developed a sensitive bioassay using the recombinant mouse hepatoma cell (Hepa1c1c7) for the determination of dioxins. The recombinant cell line was stably transfected with firefly luciferase reporter gene (pGudLuc1.1). The transfected cells showed the highest induction of luciferase activity at 4.5 hr and a decrease beyond this time point. The system showed the highest sensitivity of detection ever reported. Upon TCDD exposure cells showed 2 fold increase at 10 pM and 7 fold increase at 100 pM, respectively. The passage number after the transfection played an important role in the sensitivity. The increase of passage number tended to increase the sensitivity of the cells up to 15. The media without phenol red showed a higher induction rate than with phenol red, suggesting the preferable use of phenol red-free media for the bioassay. Since each of the assays has unique characteristics that make them suitable for some screening applications and not others, development of sensitive bioanalytical methods based on a variety of cellular systems in a key to the successful determination of dioxins. The bioassay system developed in this study will contribute to further development of successful screening the AhR agonists among the environmental mixture. In addition, the rapid and sensitive nature of this cellular system can be applied as a valuable tool to screen the dioxin-like moieties among the prodrugs at the initial stage, thereby expediting the new drug discovery.

Estudo quantitativo da lágrima pelo teste de fenol vermelho na população brasileira / Quantitative tear study using the red phenol test in the Brazilian population

OBJETIVO: Determinar parâmetros normais do teste de fenol vermelho na população brasileira, e comparar entre diferentes raças, idade e sexo. MÉTODOS: Foram avaliados, com o teste de fenol vermelho, 280 indivíduos (560 olhos) da raça branca e 280 indivíduos (560 olhos) da raça não branca, que foram divididos de acordo com a idade e o sexo. Foram excluídos indivíduos com qualquer doença ocular, usuários de lente de contato e que usavam medicação ocular. RESULTADOS: Dos 1.120 olhos avaliados, o resultado médio foi de 19,77±7,90 mm. CONCLUSAO: O resultado médio encontrado foi um valor intermediário entre as duas populações previamente estudadas (japonesa e norte-americana).

New Selective Medium for Rapid Identification of Vibrio vulnificus from Patients with V. vulnificus Sepsis / 대한피부과학회지

BACKGROUND: Vibrio(V.) vulnificus is a halophilic, gram-negative bacillus that causes a fatal sepsis in patients with underlying chronic disease such as liver cirrhosis and alcoholic abuse. Because V. vulnificus infection has a fulminant course and high mortality rate, early recognition and rapid diagnosis with prompt therapy are necessary to improve survival rate. OBJECTIVE: The purpose of this study was to develop a new selective medium for rapid identification of V. vulnificus through color change of medium according to pH from patients suspected of having V. vulnificus sepsis. METHODS: Rapid isolation and identification of V. vulnificus can be possible by modifying the component of PNC(5% peptone, 1% NaCl, and 0.08% cellobiose [pH 8.0]) broth medium. From this PNC broth, a basal broth(5% peptone+1% NaCl+cellobiose) was prepared and used to evaluate additional medium supplements(cellobiose concentration [0.08, 0.2, 0.1%], pH [6.8, 7.5, 8.0] and pH indicator dye [bromthymol blue, thymol blue, phenol red, bromcresol purple, crystal violet, cresol red, and neutral red]). To examine the rapid identification and selectivity of this basal medium according to various conditions, V. vulnificus was tested by using saline and normal human blood containing these bacteria(1, 000 bacteria/ml), respectively at 37degrees C. A positive reaction(V. vulnificus growth) appeared as color change. The selectivity and identification capacity of this new broth was tested by using other 6 Vibrio species and 14 strains of other bacteria. RESULTS: Color change appeared only in the medium including bromthymol blue and thymol blue as a pH indicator dye. It was called the basal medium containing blue dyes as PNCB(peptone, NaCl, cellobiose and blue dye) medium. It took an average time of 4.8hr for becoming aware of yellow color change in PNCB broth after cultivating with saline mixed with V. vulnificus and 6hr in PNCB broth after cultivating with blood mixed with V. vulnificus. One Vibrio species and another 3 bacteria produced color change. So we confirmed that the final composition and pH of PNCB broth medium was 5% peptone, 1% NaCl, 0.2% cellobiose, 0.0004% bromthymol blue and 0.0004% thymol blue [pH 7.5] CONCLUSIONS: PNCB broth could be used as a selective and differential medium for rapid isolation and identification of V. vulnificus in patients with V. vulnificus sepsis.

The effect of Ni2+ on the intracellular Ca2+ increase of the mouse early 2-cell embryos / 대한불임학회지

OBJECTIVE: We reported the overcoming effect of Ni2+ on the in vitro 2-cell block of mouse embryos. In this study, we aim to investigate whether Ni2+ should induce intracellular Ca2+ transient in the mouse embryos. MATERIALS AND METHODS: Embryos were collected at post hCG 32hr from the oviduct of the ICR mouse and cultured in M2 medium omitted phenol red. Intracellular Ca2+ was checked by using a confocal laser scanning microscope and fluo-3AM by using various intracellular Ca2+ antagonists. RESULTS: In 1mM Ni2+ treated medium which contained Ca2+(1.71mM), 75.7% of the embryos showed [Ca2+]i transient about 200 sec later. In the Ca2+-free medium, 69.8% of the embryos showed [Ca2+]i transient. In U73122, phospholipaseC(PLC) inhibitor (5uM, 10min) pretreated group, 33.3% of the embryos showed [Ca2+]i transient. Heparine, inositol 1,4,5-triphosphate receptor(IP3R) antagonist preinjected embryos showed no response with 1mM Ni2+. In danthrolene treatment, ryanodine receptor(RyR)-antagonist, 43% embryos showed [Ca2+]i transient but they showed delayed response about 340sec in the presence of Ca2+. CONCLUSIONS: Summing up the above results, Ni2+ seems to induce Ca2+-release from the Ca2+-store even in the Ca2+-free medium. IP3 receptors of the mouse 2-cell embryos might have an essential role for the intracellular Ca2+ increase by Ni2+.