Effect of different obturation techniques on sealer penetration into dentinal tubules after final irrigation with XP-Endo finisher file

The first objective of this research was to evaluate the effectiveness of XP-Endo Finisher on dentinal tubule penetration of irrigation solution using confocal laser scanning microscopy. The main purpose of this research was to compare the effect of cold lateral condensation, continuous wave obturation and core-carrier based techniques on sealer penetration. Sixty mandibular premolars were prepared and allocated into two experimental groups (n=30) as the final irrigation technique and obturation technique experiment. In the final irrigation technique experiment, final irrigation was performed with XP-Endo Finisher, passive ultrasonic irrigation (PUI) and conventional needle irrigation (CNI) (n=10). The roots in the obturation technique experiment were also assigned into 3 groups and obturated with cold lateral condensation, continuous-wave obturation and core-carrier techniques (n=10). The most effective activation method, which emerged as a result of the first part of this study, was used as the final irrigation method in the obturation technique experiment. Then, all roots were sectioned in 1-mm-thick slices at 3mm from the apex for scanning. In terms of depth and percentage of material penetration, CNI exhibited significantly the lowest values and no significant difference was found between others. Also, there was no significant difference among obturation methods. In conclusion, XP-Endo Finisher and PUI are more effective than CNI on irrigant penetration. Sealer penetration into dentinal tubules is independent of obturation techniques.

El objetivo principal de esta investigación fue evaluar la eficacia de XP- Endo Finisher en la penetración de la solución de irrigación en los túbulos dentinarios mediante microscopía de láser confocal. Se prepararon sesenta premolares mandibulares y se distribuyeron en dos grupos experimentales (n=30) según el tipo de método de evaluación utilizado. En el experimento de la técnica de irrigación final, la irrigación final se realizó con XP-Endo Finisher, irrigación ultrasónica pasiva (PUI) e irrigación con aguja convencional (CNI) (n=10). Las raíces en el experimento de la técnica de obturación también se asignaron en 3 grupos y se obturaron con técnicas de condensación lateral fría, obturación de onda continua y portador de núcleo (n=10). El método de activación más eficaz, que surgió como resultado de la primera parte de este estudio, se utilizó como método de irrigación final en el experimento de la técnica de obturación. Luego, todas las raíces se seccionaron en muestras de 1mm de espesor. En términos de profundidad y porcentaje de penetración del material, CNI exhibió significativamente los valores más bajos y no se encontraron diferencias significativas entre los demás. Además, no hubo diferencias significativas entre los métodos de obturación. En conclusión, XP-Endo Finisher y PUI son más efectivos que CNI en la penetración del irrigante. La penetración del sellador en los túbulos dentinarios es independiente de las técnicas de obturación.

Characteristics of corneal stromal demarcation line after different cross-linking methods in keratoconus and its influence on the cross-linking effect / 中华实验眼科杂志

Objective:To compare the characteristics of corneal stromal demarcation line after different surgical methods of riboflavin/ultraviolet A corneal collagen cross-linking (CXL) in early keratoconus, and analyze the influence of the demarcation line on the cross-linking effect.Methods:A non-randomized controlled clinical study was conducted.Sixty-nine eyes of 69 patients treated with riboflavin/ultraviolet A CXL in the Eye Hospital of Shandong First Medical University from May 2019 to February 2021 were included.According to the cross-linking methods, the patients were divided into epithelium-on treatment group (21 eyes) and epithelium-off treatment group (48 eyes). There were 25 eyes in 5.4 J energy group and 44 eyes in 7.2 J energy group.The morphology and changes of corneal stromal cross-linking reaction (corneal stromal demarcation line) were observed at 2 weeks, 1, 3 and 4 months after operation.Changes in the thinnest corneal thickness (TCT), uncorrected visual acuity (UCVA, LogMAR), best corrected visual acuity (BCVA, LogMAR) and corneal maximum curvature (Kmax) were recorded.This study adhered to the Declaration of Helsinki.The study protocol was approved by the Ethics Committee of Eye Hospital of Shandong First Medical University (No.2019.05). Written informed consent was obtained from each subject.Results:Of the 69 eyes after operation, 44 eyes (63.77%) had demarcation lines, and 25 eyes (36.23%) had no demarcation lines.The occurrence rate of demarcation lines in the epithelium-on treatment group was 79.17%(38/48), which was significantly higher than 28.57%(6/21) in the epithelium-off treatment group ( χ2=16.186, P<0.01). The occurrence rate of demarcation line in 5.4 J energy group was 72.00%(18/25), and the 7.2 J energy group was 56.80%(25/44), with no significant difference ( χ2=1.565, P=0.302). Slit lamp microscopy and anterior segment-optical coherence tomography showed that the demarcation line appeared at 1-2 weeks after operation, gradually converged and strengthened after 1 month, turned diffuse, blurred and faded by degrees after 2-3 months, and basically disappeared after 4 months.The depth of the demarcation line reached 141-423 μm, with an average depth of (263.44±84.22)μm.Scanning laser confocal microscopy showed that corneal stromal cells were activated and light reflection was enhanced after CXL.Collagen fibers extended vertically and horizontally, crisscrossed, and were in a reticular arrangement.The TCT decreased from preoperative (458.69±38.28)μm to (443.86±36.54)μm at 4 months after operation, showing a statistically significant difference ( t=6.705, P<0.001). There was no significant difference in the TCT reduction between groups with and without demarcation lines ( t=1.684, P=0.100). At 4 months postoperatively, the UCVA of all eyes increased from preoperative 0.74±0.37 to 0.69±0.38, and the difference was statistically significant ( t=2.109, P=0.039). There was no significant difference in BCVA between before and after operation ( t=1.006, P=0.319). There was no significant difference in change of UCVA and BCVA between groups with and without demarcation lines ( t=0.065, P=0.949; t=0.346, P=0.730). There was no significant difference in Kmax in all patients between before and after operation ( t=0.050, P=0.950). There was no significant difference in the Kmax change between groups with and without demarcation lines ( t=-0.739, P=0.464). The change in TCT in the epithelium-off treatment group was significantly greater than that in the epithelium-on treatment group ( t=2.815, P=0.008). There was no significant difference in UCVA, BCVA and Kmax changes between epithelium-on and epithelium-off treatment groups (all at P>0.05). There was no obvious corneal scarring, infectious keratitis, corneal endothelial decompensation or other complications. Conclusions:The demarcation line after CXL may be a sign of the depth of cross-linking reaction, which is more prone to occur after the epithelium-off operation method.Both the epithelium-on and epithelium-off operation methods have similar therapeutic effects.Demarcation line after different cross-linking methods has no significant influence on the cross-linking effect in keratoconus.

Treatment of rheumatoid arthritis by injection of sinomenine solid lipid nanoparticles under a fluorescence endoscopic laser confocal microscope / 中国中药杂志

A fluorescence endoscopic laser confocal microscope(FELCM) was used to direct the injection of sinomenine solid lipid nanoparticles(Sin-SLN) into the joint, and the in vitro effectiveness of Sin-SLN in the treatment of rheumatoid arthritis(RA) was evaluated. Sin-SLN was prepared with the emulsion evaporation-low temperature curing method. The Sin-SLN prepared under the optimal conditions showed the encapsulation efficiency of 64.79%±3.12%, the drug loading of 3.84%±0.28%, the average particle size of(215.27±4.21) nm, and the Zeta potential of(-32.67±0.84) mV. Moreover, the Sin-SLN demonstrated good stability after sto-rage for 30 days. The rabbit model of RA was established by the subcutaneous injection of ovalbumin and complete Freund's adjuvant. Five groups were designed, including a control group, a model group, a Sin(1.5 mg·kg~(-1)) group, a Sin-SLN(1.5 mg·kg~(-1)) group, and a dexamethasone(positive drug, 1.0 mg·kg~(-1), ig) group. The control group and the model group only received puncture treatment without drug injection. After drug administration, the local skin temperature and knee joint diameter were monitored every day. The knee joint diameter and the local skin temperature were lower in the drug administration groups than in the model group(P<0.05, P<0.01). FELCM recorded the morphological alterations of the cartilage of knee joint. The Sin-SLN group showed compact tissue structure and smooth surface of the cartilage. Enzyme-linked immunosorbent assay(ELISA) was employed to determine the serum le-vels of interleukin-1(IL-1) and tumor necrosis factor-α(TNF-α). The findings revealed that the Sin-SLN group had lower IL-1 and TNF-α levels than the model group(P<0.05, P<0.01). Hematoxylin-eosin(HE) staining was employed to reveal the pathological changes of the synovial tissue, which were significantly mitigated in the Sin-SLN group. The prepared Sin-SLN had uniform particle size and high stability. Through joint injection administration, a drug reservoir was formed. Sin-SLN effectively alleviate joint swelling and cartilage damage of rabbit, down-regulated the expression of inflammatory cytokines, and inhibited the epithelial proliferation and inflammatory cell infiltration of the synovial tissue, demonstrating the efficacy in treating RA.

Effect of cavity disinfection protocols on microtensile bond strength of universal adhesive to dentin / Efecto de los protocolos de desinfección de cavidades en la fuerza de adhesión microtensil del adhesivo universal a la dentina

Abstract The purpose of this study was to compare the effect of different disinfection protocols of dentin on bond strength of an MDP-containing universal adhesive. Twelve extracted mandibular third molars were separated horizontally at the mid-coronal of crown to get smooth and sound dentin surfaces using low-speed diamond saw. The teeth were randomly fallen into four groups: chlorhexidine (CHX), ozone, Er,Cr:YSGG laser irradiation (LASER) and no treatment (control). After cavity disinfection application, a universal adhesive (G-Premio Bond) was applied to the surface of dentin according to self-etch mode as instructed by the manufacturer. After incremental built-up of composite resin (Charisma Smart), the specimens were immersed in distilled water at 37°C for 24h. Dentin/composite beams with 1 mm² cross sectional area were produced and micro-tensile bond strength (µTBS) was applied on these beams (n=20). Failure mods were determined under a stereomicroscope at ×40. The resin penetration of samples stained with Rhodamine B fluorochrome dye was examined with a confocal laser scanning microscope. Statistical analysis was performed with SPSS-22. Test results were analyzed using One-way Anova and Tukey HSD Post-Hoc tests (p0.5). All applications of cavity disinfection procedures decreased the µTBS of the resin-dentin interface.

Resumen El propósito de este estudio fue comparar el efecto de diferentes protocolos de desinfección de la dentina sobre la fuerza de unión de un adhesivo universal que contiene MDP. Doce terceros molares mandibulares extraídos se quebraron horizontalmente en la mitad de la corona para obtener superficies de dentina lisas y sólidas utilizando una sierra de diamante de baja velocidad. Los dientes se dividieron aleatoriamente en cuatro grupos: clorhexidina (CHX), ozono, irradiación con láser Er,Cr:YSGG (LASER) y ningún tratamiento (control). Después de la aplicación de la desinfección de la cavidad, se aplicó un adhesivo universal (G-Premio Bond) a la superficie de la dentina según el modo de autograbado indicado por el fabricante. Después de la obturación con resina compuesta (Charisma Smart), las muestras se sumergieron en agua destilada a 37°C durante 24h. Se produjeron porciones de dentina/resina con un área de sección transversal de 1 mm² y se aplicó una fuerza de adhesión microtensile (µTBS) (n=20). Los modos de falla se determinaron bajo un microscopio estereoscópico a ×40. La penetración de la resina de las muestras teñidas con colorante fluorocromo rodamina B se examinó con un microscopio de barrido láser confocal. El análisis estadístico se realizó con SPSS-22. Los resultados de las pruebas se analizaron utilizando las pruebas post-hoc Anova unidireccional y Tukey HSD (p0.5). Todas las aplicaciones de procedimientos de desinfección de cavidades redujeron el µTBS de la interfaz resina-dentina.

Safety Evaluation of Jiaotaiwan Based on Zebrafish Model / 中国实验方剂学杂志

Objective:To take zebrafish embryo as the research object, in order to investigate the development toxicity, cardiotoxicity, liver toxicity and kidney toxicity of water extract of Jiaotaiwan (JTW) on zebrafish embryo. Method:Zebrafish embryos with normal development at 12 h (hpf) after fertilization were selected as model animals for the growth and cardiotoxicity experiments. The embryos were treated with 125, 250, 500 mg·L-1 of JTW water extracts, and the effects of the drugs on the heart rate and morphology of the embryos and LD50 were observed at 72 h (hpf) after fertilization. Zebrafish embryos with normal development at 72 h (hpf) after fertilization were used as model animals for the liver and kidney toxicity experiments. The embryos were treated with 125，250，500 mg·L-1 of JTW water extracts, and the effect of the drugs on morphological changes, Alanine aminotransferase(ALT), Aspartate aminotransferase (AST) activity, and creatinine content of the larvae and LD50 were observed at 72 d (dpf) after fertilization. Result:The zebrafish embryos in control group developed normally, the heart was well developed, and the heartbeat was even and powerful. The LD50 of JTW water extract on zebrafish embryos for 72 h was 1 023 mg·L-1. Compared with the embryos in the control group, 250，500 mg·L-1 treatment groups in the development toxicity had a smaller head, shorter body lengths (P<0.05), and decreased eye size (P<0.05). Compared with the control group embryos, the pericardial edema was observed in the 500 mg·L-1 group, the heart rate was significantly decreased in the 250，500 mg·L-1 JTW water extract groups (P<0.01), the atrial and ventricular areas were significantly reduced (P<0.05), the distance of SV-BA became significantly larger (P<0.05), the distance of AV channel became significantly larger (P<0.01), and the in-flow distance was significantly shorter (P<0.01). In the acute toxicity experiment, the LD50 of JTW water extract for zebrafish larvae for 72 h was 1 067 mg·L-1. Compared with control group, JTW water extract significantly reduced ALT activity in zebrafish larvae (P<0.05). Conclusion:This experiment found that JTW has an obvious toxicity in embryonic development, which is mainly manifested as delayed growth and severe cardiotoxicity. Great attention shall be paid to clinical administration to pregnant women, lactating women and patients with heart disease.

In vivo imaging in tumor-bearing animals and tissue distribution of nanostructured lipid carrier modified by hyaluronic acid loaded with three components in Panax ginseng / 中草药

Objective: Human hepatoma SMMC-7721 cells were transplanted into nude mice to study the tissue distribution of nanostructured lipid carrier modified by hyaluronic acid (HA-OUR-NLC) loaded with three components in Panax ginseng (oleanolic acid, ursolic acid, and ginsenosider Rg3, OUR). Methods: FITC and DiR were used as fluorescent probes to dynamically monitor the HA-OUR-NLC targeted behavior of various tissues and organs through fluorescence endoscopic confocal imaging and in vivo imaging studies. Results: RUE values of oleanolic acid, ursolic acid, and ginsenosider Rg3 in tumors were significantly increased in HA-OUR-NLC group, reaching 2.51 ± 1.23, 2.27 ± 1.43, and 2.77 ± 0.25, respectively, which indicated that nanoparticles modified by hyaluronic acid could enhance drug uptake in tumors. The DiR accumulation in tumors of DiR-HA-OUR-NLC was higher than that of DiR-OUR-NLC by the visualized fluorescence of in vivo imaging. Conclusion: It indicated that nanoparticles modified by hyaluronic acid loaded with three components in P. ginseng can be enriched in the tumor site of liver cancer, which is in line with the expectation and can significantly improve the tumor targeting of the drug delivery system.

Influencia de la medicación intracanal con pasta de hidróxido de calcio en la penetración del cemento obturador / obturadorInfluence of intracanal medication with calcium hydroxide paste in the penetration of the shutter cement / Influência da medicação intracanal com pasta de hidróxido de cálcio na infiltração do cimento obturador

La eliminación del hidróxido de calcio en el conducto radicular es determinante para el éxito del tratamiento endodóntico, los remanentes pueden interaccionar negativamente con los selladores endodónticos aumen-tando las filtraciones y disminuyendo la calidad de la obturación. Objetivo: Evaluar el efecto de la medicación intraconducto con pasta de hidróxido de calcio en la penetración del cemento obturador en el interior de los túbulos dentinarios. Materiales y métodos: 20 raíces distales de molares superiores se instrumentaron uti-lizando el Sistema Wave One Large 40/.08. Se dividieron aleatoriamente en dos grupos: uno obturado con técnica de cono único y cemento Ah plus con rodamina-B y otro obturado con la misma técnica y cemento Ah plus con rodamina B, previa colocación por 15 días y remoción mediante recapitulación de la pasta de hidróxido de calcio. Posteriormente los dientes fueron cortados transversalmente y se realizaron fotomicro-grafías del tercio cervical, medio y apical a través de la técnica de microscopia confocal de barrido por láser. La profundidad máxima de penetración fue determinada a través del programa Image J. Resultados: El ce-mento sellador Ah plus presentó menores valores de penetración cuando fue empleada previamente la pasta de hidróxido de calcio como medicación intracanal (p<0,01). El tercio del conducto con mayor penetración fue el tercio cervical seguido por el tercio medio y por último el apical (p<0,01). Conclusión: Los remanentes de hidróxido de calcio disminuyen la penetración del cemento sellador Ah plus en los túbulos dentinarios en todos los tercios del conducto radicular.

The elimination of calcium hydroxide in the root canal is decisive for the success of endodontic treatment, the remnants can interact negatively with endodontic sealants increasing filtrations and decreasing the quality of the seal. Objective: To evaluate the effect of intra-duct medication with calcium hydroxide paste on the pen-etration of the sealing cement inside the dentinal tubules. Materials and methods: 20 distal roots of upper molars were instrumented using the Wave One Large 40 / .08 System. They were randomly divided into two groups: one sealed with a single cone technique and Ah plus cement with rhodamine-B and another sealed with the same technique and Ah plus cement with rhodamine B, previous placement for 15 days and removal by recapping the paste calcium hydroxide. Subsequently, the teeth were cut transversely and photomicro-graphs of the cervical, middle and apical third were performed using the laser scanning confocal microscopy technique. The maximum depth of penetration was determined through the Image J program. Results: The Ah plus sealing cement had lower penetration values when the calcium hydroxide paste was previously used as an intra-channel medication (p <0.01). The third of the duct with the highest penetration was the cervical third followed by the middle third and finally the apical (p <0.01). Conclusion: Remaining calcium hydroxide decreases the penetration of the sealing cement Ah plus in the dentinal tubules in all thirds of the root canal.

A eliminação do hidróxido de cálcio do canal radicular é determinante para o sucesso do tratamento endo-dôntico; os remanescentes podem interagir negativamente com os cimentos endodônticos, aumentando as filtrações e diminuindo a qualidade do selamento. Objetivo: Avaliar o efeito da medicação intracanal com pasta de hidróxido de cálcio na penetração do cimento de selamento no interior dos túbulos dentinários. Materiais e métodos: 20 raízes distais de molares superiores foram instrumentadas usando o sistema Wave One Large 40 /.08. Eles foram divididos aleatoriamente em dois grupos: um selado com técnica de cone único mais cimento Ah plus com rodamina-B e outro selado com a mesma técnica e cimento Ah plus com rodami-na-B, prévia colocação por 15 dias e remoção recolocação da pasta hidróxido de cálcio. Posteriormente, os dentes foram cortados transversalmente e fotomicrografias do terço cervical, médio e apical foram realizadas pela técnica de microscopia confocal de varredura a laser. A profundidade máxima de penetração foi de-terminada pelo programa Image J. Resultados: O cimento selante Ah plus apresentou menores valores de infiltração quando a pasta de hidróxido de cálcio foi utilizada anteriormente como medicamento intracanal (p <0,01). O terço do conduto com maior penetração foi o terço cervical, seguido pelo terço médio e finalmente o apical (p <0,01). Conclusão: O hidróxido de cálcio restante diminui a penetração do cimento selante Ah plus nos túbulos dentinários em todos os terços do canal radicular.

Application of corneal laser confocal microscopy in diabetic retinopathy patients / 眼科新进展

Objective To investigate the value of corneal laser confocal microscope in the diagnosis of corneal subepithelial plexus,corneal cell density and morphological changes of patients with diabetic retinopathy (DR).Methods Together 94 cases of confirmed DR (114 eyes),including 41 cases of non-proliferative diabetic retinopathy (NPDR group,52 eyes) and 53 cases of proliferative diabetic retinopathy (PDR group,62 eyes) were selected from January 2016 to April 2017,and meanwhile,40 diabetic patients (40 eyes) with no fundus abnormality were grouped as control group.Corneal laser confocal microscopy was used to compare the corneal subepithelial plexus,corneal cell density and morphological changes in the three groups,Results The cell densities of the basal layer,the superficial stromal layer,the medium stromal layer and the deep stromal layer of the cornea in NPDR and PDR were significantly lower than those of the control group (all P ＜ 0.05),and the PDR group was significantly lower than the NPDR group (all P ＜ 0.05).The corneal endothelial cell density,hexagonal cell ratio,nerve fiber density,and nerve fiber length in the NPDR and PDR group were significantly lower than those in the control group (all P ＜ 0.05);and the variability of endothelial cell and nerve branch density in NPDR and PDR patients were significantly higher than those in the control group (all P ＜ 0.05).The corneal endothelial cell density,hexagonal cell ratio,nerve fiber density,and nerve fiber length in the PDR group was (1962.0-± 117.3) · mm-2,46.1％ ± 5.5％,(15.4 ± 3.3) · mm-2,(6.2 ± 2.7) mm · mm-2,respectively,which were significantly lower than those in the NPDR group [(2381.4 ± 144.0) · mm-2,58.2％ ±7.0％,(20.6 ±3.8) ·mm-2,(8.6 ± 2.4)mm · mm-2,respectively] (all P ＜ 0.05),but the variability of endothelial cell and nerve branch density in PDR group were significantly higher than those in the NPDR group (all P ＜ 0.05).Conclusion Corneal confocal microscopy can effectively observe the density and morphological changes in corneal subepithelial plexus and corneal cell in DR patients so as to provide guidance for clinical diagnosis and treatment.

Analysis of the polymorphs of carbamazepin tablets by laser confocal micro-Raman spectrometer and establishment of the identification test model / 中国药学杂志

OBJECTIVE: To analyze the polymorphs of carbamazepin tablets by laser confocal micro-Raman spectrometer and establish the identification test model. METHODS: Reference Raman spectra of the four crystal forms of carbamazepin were established; then the polymorphs of the carbamazepin tablets from different manufacturers were analyzed, and an identification test model was established to distinguish the unknown samples. RESULTS: According to the results of the cluster analysis, the four batches of carbamazepin tablets from domestic manufacturers belonged to crystal form III. A qualitative model was created by choosing the first derivative and vector normalization method to process the spectra, using the factorization method to calculate, and selecting the 200-500, 650-850, 1 000-1 100, 1 500-1 700 cm-1 regions to compare. This model was verified to have high sensitivity, specificity, precisicion, negative prediction ability, and detection efficiency. CONCLUSION: The laser confocal micro-Raman spectrometer canbe used to analyze the polymorphs of carbamazepin tablets and the qualitative identification is rapid and accurate.

Effects of chronic alcohol exposure on hippocampus and cerebral cortex neurons in mice / 药学学报

This study was performed to investigate the changes of the number, morphology and ultrastructure of the central nervous system of mice during the long-term alcohol exposure. Mice at 60 days in age were used to establish the long-term alcohol exposure model. The structure of the central nervous system, such as nuclear antigen, dendritic spines and synapses, were labeled by the methods of immunocytochemistry and DiI (1, 1'-dioctadecyl-3, 3, 3', 3'-tetramethy lindocarbocyanine perchlorate) scattering. The results showed that prolonged alcohol exposure could promote apoptosis of nerve cells in the central nervous system, and inhibit the proliferation of neural stem cells, which reduced the number of nerve cells in the central nervous system. Long-term ethanol exposure can also lead to a decrease in the density of dendritic spines of neuron, a smaller number of synapses (connections between nerve cells), and some changes in synaptic ultrastructure. The density of nerve cells and their dendritic spines, as well as the changes of synaptic ultrastructure, suggest that the function of nerve cells may be low.

Effect of hsa-microRNA-218 on granulysin expression / 中国免疫学杂志

Objective:To elucidate the effect of hsa-microRNA-218(hsa-mir-218)on exogenous granulysin (GLS) expression in 293T cells.Methods:Total RNA was extracted from THP-1 cells induced by phorbol 12-myristate 13-acetatefor (PMA), and GLS gene was amplified by RT-PCR, and then cloned into pDsRed-Express-C1 to construct the GLS expression vector pDsRed-GLS.Then 293T cells were co-transfected with pDsRed-GLS and pGenesil-mir-218 (pGenesil-mir-control) and laser confocal microscopy was per-formed 36 h later to detect their co-expression .Total RNA and protein were extracted 48 h post transfection , and RT-PCR and Western blot were performed to detect the effect of hsa-mir-218 on exogenous GLS expression .Results:The GLS expression vector pDsRed-GLS was constructed successfully and laser confocal microscopy indicated that it was co -expressed with the interference vector .Compared with that of cells transfected with pGenesil-mir-control, Western blot showed a markedly decrease of GLS protein expression (50%) in the cells transfected with pGenesil-mir-218.However, GLS mRNA expression remained unchanged .Conclusion: hsa-mir-218 nega-tively regulates GLS expression at a post-transcriptional level , and this provides an experimental basis for future study of mechanism of GLS expression regulated by mir-218 .

Phenotypes,distribution, and morphological features of antigen-presenting cells in the normal murine iris / 中华实验眼科杂志

Background The conventional study of antigen-presenting cells(APCs)in eye relies on in vitro histoimmunochemistry,but its outcome is influenced by many factors.The anterior chamber injection of fluoresceinmarked antibody was used as a new approach before,however,it is liable to lead to injury of cornea.The intravitreal injection of fluorescein-labeled antibody may be important for the in vivo study of the phenotype features of APCs in iris,which is significant for evaluating the function of APCs in immune homeostasis.Objective This study was to investigate the phenotype characters,distribution and morphology of different types of APCs in the normal murine iris.Methods Fifty-one SPF female BALB/c mice(from 6-to 8-week old)were randomized into 17 groups according to the injection of different antibodies.Alexa Fluor 594 or Alexa Fluor 488-tagged ovalbumin (OVA),CD11 c,major histocompatibility complex Ⅱ (MHC-Ⅱ),F4/80,B7-1 and B7-2 monoclonal antibodies or mixtures of two antibodies (2.0 μl)were intravitreally injected at 0.5 mm far from corneal limbus with microneedle under the biomicroscope.The iris tissues were isolated 24 hours after injection.The phenotype characters,precise distribution and morphology of different types of APCs were identified by epifluorescence microscope and laser confocal microscope.In vitro staining was also performed to validate the in vivo staining results.Results After in vivo staining via intravitreal injection,the cell positive for OVA as well as MHC-Ⅱ,F4/80,CD11 c,B7-1 and B7-2 were exhibited with the regular networkline appearance throughout the normal murine iris.Positive cells tagged with Alexa Fluor 594 or Alexa Fluor 488 presented the red or green fluorescence.Double-fluorescein staining showed that about 90％ of F4/80+ cells were OVA+,and MHC-Ⅱ was expressed in about 60％ of F4/80+ cells and CD11c+cells,and about 35％ of F4/80+ cells and CD1 1 c+ cells expressed B7-1 and B7-2 simultaneously,and over 70％ of OVA+ cells were positive to MHC-Ⅱ.These labeled cells were identified as two populations based on their shape.One type was dendritiform cell (DC) with a small cell body and many long dendrites,including OVA+,CD1 1 c+,F4/80+ cells and MHC-Ⅱ + cells ; and the other types were polymorphic population being round,pleomorphic or irregular shape with a large cell body and a few short dendrities,including B7-1 + and B7-2+ cells.Conclusions In vivo intravitreal injection of labeled antibodies can be adapted to visualize the labeled cells in the murine iris.APCs with distinct morphologies,phenotypes and distribution may contribute to the immunologically privileged feature and inflammation of the eye.

A cell co-culture model for studying bystander effect and its application on bystander DNA double-strand breaks induced by alpha-particles irradiation / 中华放射医学与防护杂志

Objective To establish an experimental model for the study of α-particle-induced bystander effect of DNA damage and investigate the characteristics of bystander DNA double-strand break (DSB).Methods The red fluorescence fusion protein of HsBrkl-RFP was used to mark the cytoplasm of one cell line to distinguish the irradiated target cells (HFS-RFP) and the non-irradiated bystander cells (HFS) in the co-culture cellular model.After α-particle irradiation,cellular DSB and its repair kinetics were analyzed by the immunofluorescence staining of γH2AX and laser confocal microscope observation.Results A bystander studying model was established by co-culturing human HFS-RFP cells with its partner HSF cells.After 0.1 Gy or 0.2 Gy α-particle irradiation,the similar kinetics of γH2AX foci production and abatement were observed in both irradiated HFS-RFP cells and non-irradiated bystander HFS cells,in which the highest level of γH2AX foci was detected at 1 h post-irradiation.The second peak of γH2AX foci formation appeared at 8 h post-irradiation,which possibly indicates the occurrence of secondary DSB.However,the production of secondary DSB in the bystander cells was weaker than that in the irradiated cells.Conclusions The cell co-culture model can be used for bystander effect investigation.Bystander DSB can be effectively induce by irradiation and the secondary breakage of DNA DSB in the bystander cells may relative to the consequential biochemical processing of clustered DNA damage.

Analysis of microvessel density in pterygium tissue with corneal laser confocal microscopyc in vivo and immunohistochemistry in vitro / 中华实验眼科杂志

BackgroundPterygium is an ocular surface disease of abnormal cell proliferative kind and angiogenesis plays an important role in its development and recurrence.Several anti-angiogenic therapies have been used to treat pterygium,but there very few studtes for the in vivo observation of the microvessles in pterygium.ObjectiveThis study was to observe angiogenesis in pterygium with a high-resolution confocal microscope in vivo and to perform immunohistochemical study in vitro.MethodsA prospective case-controlled study was designed.Twenty eyes of 20 consecutive patients with primary pterygia and 20 age- and sex-matched patients with inner eye diseases and strabismus with normal conjunctiva were enrolled in this study.An in vivo confocal microscopy imaging system (Heidelberg Retina Tomograph Ⅱ Rostock Cornea Module) was used to collect microvascular pictures from the anterior part of pterygia and normal nasal conjunctiva of controls,and then immunochemistry was performed to examine the expression of CD31 in microvessel in vitro.The vascular density values were compared between these two groups.The correlation of vascular density values between in vivo Heidelberg Retina Tomograph and in vitro immunohistochemistry was calculated.Written informed consent was obtained from pationts before any examination and surgery.ResultsUnder the in vivo confocal microscope,the microvessel density was (8929±2993) μm/mm2 and (4202 ±692)μm/mm2,respectively,in pterygium and the normal conjunctiva group with a statistically significant difference between them (t =6.881,P＜0.01 ).Immunochemistry revealed that the expression of CD31 to measure vascular density was ( 21.00 ± 4.06/400 × field ) and ( 6.07 ± 1.75/400 × field ) in pterygium and the normal conjunctiva group,showing significant difference (t =12.312,P＜0.01 ).Positive correlations were found in the vascular density values between in vivo corneal laser confocal microscopy examination and in vitro immunochemistry examination in both the pterygium group and normal conjunctiva group (pterygium group:r=0.649,P＜0.01 ;normal conjunctiva group: r=0.572,P＜0.01 ) ConclusionsIn vivo confocal microscopy imaging is superior to in vitro immunochemistry in evaluating the microvessel of pterygium.The results of this study offer a new way index for further investigation of the biological behavior of pterygium and its mechanism.

Microscopia confocal de barrido láser y su relación con la morfología de la bula de filtración / Confocal laser scanning microscopy and its association to the morphology of the filtering bleb

Objetivo: describir los hallazgos de la microscopia confocal de barrido láser y su relación con la morfología de la bula de filtración.Métodos: estudio observacional descriptivo de corte transversal en 100 ojos. Se les realizó microscopia confocal de barrido láser con el HRT II y el módulo corneal de Rostock. Se aplicó la prueba de significación estadística de Mc. Nemar para una p=0,05.Resultados: predominó el grupo entre 61 y 80 años de edad (55,8 porciento) y el color de la piel blanca (46,8 porciento). En el grupo de descenso de la presión intraocular en más del 30 porciento se ubicó la mayor cantidad de bulas de todos los tamaños. Las bulas de mediano tamaño fueron las que más disminuyeron las cifras de presión intraocular, con 24 ojos (55,8 porciento, p=0,00), seguidas por las de pequeño tamaño (p=0,14). Las bulas de filtración aplanadas fueron las más frecuentes en 55 porciento de los casos (p=0,00), y 67,4 porciento de estas se ubicaron en el grupo de descenso de presión intraocular de más de 30 porciento (p=0,01). Las bulas medianas presentaron la mayor cantidad de estroma en malla porosa (60 porciento) y de microquistes epiteliales (56 porciento) p=0,00.Conclusiones: la configuración aplanada y el tamaño mediano de la bula de filtración se relacionaron con la presencia de variables histológicas que infieren buen funcionamiento de la bula (microquistes epiteliales y estroma en malla porosa). También se relacionaron con un mayor descenso de la presión intraocular

Objective: to describe the findings in the confocal laser scanning microscopy and its relation with the morphology of the filtering bleb. Methods: cross-sectional, observational and descriptive study of 100 eyes that underwent confocal laser scanning microscopy with the Retinal Heidelberg Tomography and the Rostock corneal module. Mc NemarÆs statistical significance test was made to obtain p=0,05. Results: the 61-80 years old age group (55,8 per cent) and the caucasians predominated. The group of eyes with over 30 per cent decrease of the intraocular pressure showed the highest amount of filtering blebs of all sizes. The medium-size blebs were the ones that decreases the IOP in a higher percentage (24 eyes; 55,8 per cent) for p=0,00, followed by the small blebs (p=0,14). The flat blebs were predominant in 55 per cent of cases (p=0,00) and the 67,4 per cent of them were consistent with the major range of decrease of the intraocular pressure (p=0,01). The medium size blebs presented the major number of stromas in porous mesh (60 per cent), and of epithelial mycrocists (56 per cent) for p=0,00. Conclusions: the flat configuration and the medium size of the blebs were related with the presence of the histological variables that infer adequate function of the filtrating blebs (stroma in porous mesh and epithelial mycrocists). They were also associated with higher decrease of intraocular pressure

Schistosoma mansoni specimens first described by Pirajá da Silva in Brazil (1908) re-examined by confocal laser scanning microscopy / Espécimes de Schistosoma mansoni descritos por Pirajá da Silva no Brasil (1908) reexaminados pela técnica de microscopia de varredura a laser confocal

INTRODUCTION: Pirajá da Silva made a seminal contribution to helminthology by demonstrating both schistosome eggs in feces of patients from the State of Bahia and the morphology of Schistosoma mansoni adult worms. METHODS: In this study, a microscopic investigation of the whole-mounts deposited at the Helminthological Collection of the Oswaldo Cruz Institute is presented. Confocal laser scanning microscopy was used as the main investigation technique. RESULTS: At the anterior end of the adult male, suckers with well-developed musculature and germinative cells inside the testicular lobes were observed, as well as spines located in the mid region of the male gynecophoric canal. Tegumental tubercles and transversal and longitudinal muscular bundles were observed at the dorsal surface. The female reproductive system presented a uterus lacking eggs and an elongated ovary with germinative cells. The vitellaria were restricted to the extreme posterior end of each female connected to a short vitelline duct. CONCLUSIONS: The results reported in this study demonstrate that the characteristic studied here are similar to those previously reported, using fresh worms. Moreover, this study also highlights the importance of deposits of specimens in helminthological collections, which further permit revisiting whole-mounts in such institutions.

INTRODUÇÃO: Pirajá da Silva fez contribuição magnífica à helmintologia ao descrever ovos de Schistosoma mansoni nas fezes de um paciente, no Estado da Bahia e a morfologia de vermes adultos. MÉTODOS: Neste estudo, apresentamos uma avaliação microscópica das lâminas montadas e depositadas na Coleção Helmintológica do Instituto Oswaldo Cruz. A técnica empregada nesta nova análise foi a microscopia de varredura a laser confocal. RESULTADOS: Na parte anterior dos vermes adultos machos, observamos ventosas com musculatura bem desenvolvida e células germinativas dentro dos lobos testiculares. Visualizamos, também, espinhos localizados na região mediana do canal ginecóforo. Na superfície dorsal, encontramos tubérculos e feixes musculares transversais e longitudinais. Em relação ao aparelho reprodutivo feminino, pudemos distinguir um ovo no interior do útero e o ovário alongado com células germinativas. As glândulas vitelínicas estavam restritas à parte posterior das fêmeas conectadas por um ducto vitelínico curto. CONCLUSÕES: As características morfológicas são similares as estudadas anteriormente por Pirajá da Silva com vermes frescos. Além disso, este estudo demonstra a importância de se depositar espécimes nas coleções helmintológicas abrindo possibilidade de novos estudos com estas lâminas.

Establishment of co-culture model in vitro to induce bone marrow mesenchymal stem cells differentiate into lung epithelial cells / 西安交通大学学报(医学版)

Objective To establish the co-culture model in vitro and induce bone marrow mesenchymal stem cells (MSCs) to differentiate into lung alveolar epithelial cells. Methods Each group had 6 samples, control group was MSCs alone; Group A was the MSCs cultured with the cells from normal lung; and Group B was the MSCs with the cells from injuried lung. Each group was cultured for 8 days and the two markers of lung alveolar epithelial cells including AQP5 and SP-C were tested by laser confocal microscopy and RT-PCR. Results Only AQP5 was detected in the control group and Group A, both AQP5 and SP-C were detected in Group B, the AQP5 mRNA expression in Group B was significantly increased compared with that in the control group(P<0.01). The AQP5 mRNA expression in Group B was also significantly increased compared with that in Group A (P<0.01). But there was no significant difference in AQP5 mRNA expression between Group A and control group. Conclusion We have successfully established the co-culture model in vitro to induce bone marrow mesenchymal stem cells to differentiate into lung epithelial cells.

Effect of Gross Saponin Tribulus Terrestris on protein kinase Cexpression in neonate rat cardiocytes injured by hypoxia / 吉林大学学报(医学版)

Objective To study the protective mechanism of Gross Saponin Tribulus Terrestris(GSTT) on the neonatal rat ventricular cardiocytes injured by hypoxia and the effect on protein kinase C(PKC).Methods The hypoxia-ischemia model was performed by treating the cultured neonatal rat ventricular cardiocytes with NaCN.The effect of GSTT(100 and 30 mg?L~(-1)) on the contents of ?PKC and ?PKC were detected by flow cytometry and laser confocal microscopy system.Results GSTT up-regulated ?PKC and ?PKC expressions.The contents of ?PKC and ?PKC in GSTT 100 mg?L~(-1) group(1 325.00?53.25,810.55?36.89;66.22?6.23,40.12?2.21) were increased significantly than those in model group(792.00?32.36,492.40?30.15;32.70?2.78,29.28?4.82)(P

Comparative study on different methods for quantitative analysis of oral common microorganisms / 上海第二医科大学学报

flow cytometry.A strong linear relationship was observed in the standard curve of real-time PCR of each bacteria. Conclusion These three non-culture methods can be used in the quantitative analysis of oral microorganisms.Real-time PCR and laser scanning confocal microscopy are better than the traditional culture-based CFU count,and real-time PCR is the most sensitive method.

Protective Effect of Scutellarin on Ischemic Injury in PC12 Cells / 中国药房

OBJECTIVE:To investigate the protective effect of scutellarin on ischemic injury in PC12 cells and explore its action mechanisms.METHODS:PC12 cells were treated with Na2S2O4 in combination with sugar deficiency to induce ischemic injury model.The survival rate of the PC12 cells were detected using MTT assay;and the activity of lactate dehydrogenase(LDH)were detected.The apoptosis was confirmed by Hoechst 33342 staining and flow cytometric assay was performed to determine the apoptotic rate and the mitochondrial membrane potential(MMP)in PC12 cells.The concentration of cellular calcium ion(Ca2+)was detected by laser scanning confocal microscopy.RESULTS:Within the range of 10-7～10-5 mol?L-1,Scutellarin significantly increased the survival rate of the injured cells,decreased ischemic injury-induced LDH release within the cultured medium in a concentration dependent manner,decreased the apoptotic rate,stabilized mitochondrial transmembrane potential and decreased the intracellular concentration of Ca2+ in concentration-dependent manner.CONCLUSION:Scutellarin has remarkable protective effect on PC12 cells against the ischemic injury.The mechanism of the protection might be attributed to its action of stabilizing mitochondrial transmembrane potential and inhibiting on the inflow of Ca2+.