The effect of sonic activated device used for different time on intracanal smear layer removal: An SEM analysis / 口腔医学

Objective @#To observe the clearance of smear layer on the root canal wall in different action time by scanning electron microscope (SEM), and to determine the optimal amount of time using sonically activated irrigation to wash root canal in clinic. @*Methods@# Fifty-six ex vivo human anterior teeth with single straight root canal were selected. After routine mechanical preparation, they were divided into two experimental groups according to different irrigating agents: saline group and EDTA group. Each group was assisted by VDW sonic activation EDDY. The saline group was divided into three subgroups according to the irrigating time: 5 s, 30 s and 50 s; EDTA group was divided into six subgroups according to the irrigating time: 5 s, 10 s, 20 s, 30 s, 40 s and 50 s. The control group did not undergo root canal irrigation. After irrigation, the root was cut longitudinally. The smear layer of crown, middle and apical of root canal wall was observed by SEM.@* Results@# After irrigating for 30 seconds, there was a significant difference between the normal saline group and the control group and the 5 second group (P<0.05), and there was no difference in the middle and apical part (P>0.05). After 50 seconds, there was a significant difference in the score of the smear layer between the apical area and the other groups (P<0.05). After irrigating for 5 seconds or 10 seconds in EDTA group, there was a significant difference between the scores of the crown and middle area of the root canal and the control group (P<0.05), and there was no significant difference in the apical area (P>0.05). There was a significant difference between the 20-40 second group and the first two groups (P<0.05). There was a significant difference between the 50 second group and the other groups (P<0.05). Comparing the cleaning effect on the smear layer after 50 seconds of irrigating between the two experimental groups, the whole root canal showed significant statistical difference (P<0.05). @*Conclusion @#The EDTA-assisted sonic activated device used for 50 seconds has the best cleaning effect.

Effect study of Sonic hedgehog overexpressed hair follicle stem cells in hair follicle regeneration / 中国修复重建外科杂志

OBJECTIVE@#To determine the expression level of Sonic hedgehog (Shh) in the passage of hair follicle stem cells (HFSCs), analyze the effect of Shh overexpression on the proliferation activity of HFSCs, and explore the survival of HFSCs after Shh overexpression and its effect on hair follicle regeneration.@*METHODS@#Hair follicles from the normal area (H1 group) and alopecia area (H2 group) of the scalp donated by 20 female alopecia patients aged 40-50 years old were taken, and the middle part of the hair follicle was cut under the microscope to culture, and the primary HFSCs were obtained and passaged; the positive markers (CD29, CD71) and negative marker (CD34) on the surface of the fourth generation HFSCs were identified by flow cytometry. The two groups of HFSCs were transfected with Shh-overexpressed lentivirus. Flow cytometry and cell counting kit 8 assay were used to detect the cell cycle changes and cell proliferation of HFSCs before and after transfection, respectively. Then the HFSCs transfected with Shh lentivirus were transplanted subcutaneously into the back of nude mice as the experimental group, and the same amount of saline was injected as the control group. At 5 weeks after cell transplantation, the expression of Shh protein in the back skin tissue of nude mice was detected by Western blot. HE staining and immunofluorescence staining were used to compare the number of hair follicles and the survival of HFSCs between groups.@*RESULTS@#The isolated and cultured cells were fusiform and firmly attached to the wall; flow cytometry showed that CD29 and CD71 were highly expressed on the surface of the cells, while CD34 was lowly expressed, suggesting that the cultured cells were HFSCs. The results of real-time fluorescence quantitative PCR and Western blot showed that the expression levels of Shh protein and gene in the 4th, 7th, and 10th passages of cells in H1 and H2 groups decreased gradually with the prolongation of culture time in vitro. After overexpression of Shh, the proliferation activity of HFSCs in the two groups was significantly higher than that in the blank group (not transfected with lentivirus) and the negative control group (transfected with negative control lentivirus), and the proliferation activity of HFSCs in H1 group was significantly higher than that in H2 group before and after transfection, showing significant differences ( P<0.05). At 5 weeks after cell transplantation, Shh protein was stably expressed in the dorsal skin of each experimental group; the number of hair follicles and the expression levels of HFSCs markers (CD71, cytokeratin 15) in each experimental group were significantly higher than those in the control group, and the number of hair follicles and the expression levels of HFSCs markers in H1 group were significantly higher than those in H2 group, and the differences were significant ( P<0.05).@*CONCLUSION@#Lentivirus-mediated Shh can be successfully transfected into HFSCs, the proliferation activity of HFSCs significantly increase after overexpression of Shh, which can secrete and express Shh continuously and stably, and promote hair follicle regeneration by combining the advantages of stem cells and Shh.

Comparison of four methods that remove calcium hydroxide from root canals / 口腔疾病防治

Objective@# To compare the efficiency of four methods that remove calcium hydroxide in root canals and to guide clinical practice. @* Methods @# Sixty-five isolated mandibular single root canal premolars were collected. After crown cutting and root canal preparation, a tooth was randomly selected as the blank control group, and the remaining 64 teeth were equally divided into Groups A and B (n = 32). Group A was injected with water-soluble calcium hydroxide, and Group B was injected with oil-soluble calcium hydroxide. After 2 weeks of drug sealing, Groups A and B were randomly divided into 4 groups (n = 8), including the lateral opening syringe group, sonic vibration group, ultrasonic group, and Er: YAG laser group. Before and after calcium hydroxide removal, the samples were scanned by cone-beam CT, and the data were imported into Mimics for 3D reconstruction. The root canal was divided into the following segments: superior root segment, middle and apical, and the calcium hydroxide volume of each segment of the root canal was calculated. The volumes of calcium hydroxide before and after removal were V1 and V2, respectively, with a clearance rate = (V1-V2)/V1×100%. Three-factor ANOVA was used for statistical analysis. After Groups A and B were reconstructed, the apical region with residual calcium hydroxide was selected, and the blank control was observed by scanning electron microscopy (SEM). @*Results @# Two types of calcium hydroxide could not be completely removed by the four flushing methods. The clearance rate of water-soluble calcium hydroxide was higher than that of oil-soluble calcium hydroxide (P<0.001). Among the three segments of the root canal, the clearance rate of the apical segment was lower (P<0.05). The Er: YAG laser treatment group showed the highest removal efficiency of two kinds of calcium hydroxide, which was higher than that of the other groups, especially in apical of the root. Compared with the sonic wave washing group and the syringe washing group, the ultrasonic wave washing group exhibited significant advantages (P<0.05). The clearance rate of the sonic wave washing group was higher in the oily calcium hydroxide root middle group than in the syringe washing group (P<0.05). SEM showed that the two kinds of calcium hydroxide could not be completely removed, but the residual rate of oil-soluble calcium hydroxide was large.@*Conclusion @# Both types of calcium hydroxide could not be completely removed, and compared to water-soluble calcium hydroxide, oil-soluble calcium hydroxide was more difficult to remove. Among the four cleaning methods, Er:YAG laser swing washing showed the higher cleaning efficiency.

O papel de proteínas das vias WNT/ß-Catenina, TGF-ß/BMP4 e SHH na odontogênese e odontomas / The role of proteins from the WNT/ß-Catenin, TGF-ß/BMP4 and SHH pathways in odontogenesis and odontomas

O desenvolvimento do dente depende de uma série de interações sinalizadoras recíprocas entre o epitélio oral (EO) e o ectomesênquima derivado da crista neural, a via WNT com o TGF-ß e BMP4 tem sido implicada na tumorigênese. A via de sinalização tipo Wingless (Wnt) / ß-catenina é essencial para a ativação precoce da odontogênese e no desenvolvimento de tumores odontogênicos. O TGF-ß e as BMPs tem sido associadas aos processos de dentinogênese reacionária e reparadora. A sinalização de Shh pode regular a proliferação celular no ectomesênquima dentário, controlando assim a morfogênese dentária. O objetivo da pesquisa foi investigar a atuação de algumas proteínas das vias na odontogênese e na formação de odontomas e tumores odontogênicos mistos benignos, para isto, foi desenvolvido um estudo seccional restrospectivo e imuno-histoquímico contendo 23 odontomas compostos, 21 odontomas complexos, 17 germes dentários, 05 fibro-odontomas ameloblásticos e 01 fibroma ameloblástico. Os resultados encontrados demonstraram maiores imunoexpressões da via WNT/ß-catenina no epitélio dos germes dentários (p<0,001) e no fibroma ameloblástico, enquanto que, esteve no ectomesênquima dos odontomas (p<0,001) e fibro-odontomas ameloblásticos. A via WNT/ßcatenina correlacionou-se moderadamente e significativamente com a CK14 no epitélio (p = 0,007) dos odontomas. A BMP4 foi imunoexpressa, especialmente, no ectomesênquima dos odontomas complexos (mediana = 33,7; p<0,001). A via Shh foi mais imunoexpressa no epitélio dos germes dentários (p<0,001) e no ectomesênquima dos odontomas complexos (p=0,029). De forma similar, o TGFß apresentou maior imunoexpressão no epitélio dos germes dentários (p<0,001) e no ectomesênquima dos odontomas complexos (p = 0,002). O dente em desenvolvimento exibiu maiores concentrações para estas proteínas no epitélio odontogênico nas fases de botão e capuz e a expressão diferencial ocorreu, principalmente, no ectomesênquima dos tumores, o que indica que esse componente é de fato mais proliferativo (AU).

Tooth development depends on a series of reciprocal signaling interactions between oral epithelium (EO) and neural crest-derived ectomesenchyme, the WNT pathway with TGF-ß and BMP4 has been implicated in tumorigenesis. The Wingless (Wnt)/ß-catenin signaling pathway is essential for the early activation of odontogenesis and the development of odontogenic tumors. TGF-ß and BMPs have been associated with reactionary and reparative dentinogenesis processes. Shh signaling can regulate cell proliferation in dental ectomesenchyme, thus controlling dental morphogenesis. The objective of the research was to investigate the role of some proteins in the pathways in odontogenesis and in the formation of odontomas and benign mixed odontogenic tumors. tooth germs, 05 ameloblastic fibro-odontomas and 01 ameloblastic fibroma. The results found showed higher immunoexpressions of the WNT/ß-catenin pathway in the epithelium of tooth germs (p<0.001) and in ameloblastic fibroma, while it was in the ectomesenchyme of odontomas (p<0.001) and ameloblastic fibroodontomas. The WNT/ß-catenin pathway correlated moderately and significantly with CK14 in the epithelium (p = 0.007) of odontomas. BMP4 was immunoexpressed, especially in the ectomesenchyme of complex odontomas (median = 33.7; p<0.001). The Shh pathway was more immunoexpressed in the epithelium of tooth germs (p<0.001) and in the ectomesenchyme of complex odontomas (p=0.029). Similarly, TGF-ß showed higher immunoexpression in the epithelium of tooth germs (p<0.001) and in the ectomesenchyme of complex odontomas (p = 0.002). The developing tooth exhibited higher concentrations of these proteins in the odontogenic epithelium in the bud and cap phases and the differential expression occurred mainly in the ectomesenchyme of the tumors, which indicates that this component is in fact more proliferative (AU).

Potential mechanism of NVP-LDE225 combined with radiotherapy for melanoma / 中华放射肿瘤学杂志

Objective:To explore the potential mechanism and feasibility of sonic hedgehog (SHH) signal pathway inhibitor NVP-LDE225 combined with radiotherapy in the treatment of melanoma.Methods:The Gli1 mRNA expression of the melanoma cells (Melan-A and SK-MEL-2) was detected by qPCR. After treatment with NVP-LDE225, GDC-0449 or combined with radiation for 24 hours, the number and activity of Melan-A and SK-MEL-2 were detected by cell counting and CCK-8 kit. The survival and proliferation ratio of Melan-A and SK-MEL-2 were detected by cell cloning. The changes of cell cycle of melanoma Melan-A cells were determined by flow cytometry. The levels of the Bax and Caspase3 of Melan-A apoptosis protein in melanoma cells were detected by Western blot.Results:NVP-LDE225, an inhibitor of Smo, decreased the mRNA expression of Gli1 in the melanoma cells in a dose-dependent manner, inhibited the proliferation ratio of the melanoma cells, induced apoptosis, and arrested melanoma cells in G 0 / G 1 phase. Gamma ray irradiation after NVP-LDE225 treatment further inhibited the SHH signal pathway, arrested the melanoma cells in G 2 / M phase, and further increased the inhibitory effect on melanoma cell proliferation. Conclusions:NVP-LDE225, an inhibitor of Smo, can inhibit SHH signal pathway in melanocytes, suppress cell proliferation, and further increases the effect of inhibiting cell proliferation after combining with irradiation. It can be used as a potential drug in combination with radiotherapy in the treatment of melanoma, which is worthy of further study.

The role of nuclear transcription factor Gli in the regulation of hepatic epithelial mesenchymal transition in biliary atresia mice caused by Rhesus monkey rotavirus infection / 中华实用儿科临床杂志

Objective:To investigate the role of nuclear transcription factor Gli1/Gli2 of the sonic hedgehog (Shh) signaling pathway in the hepatic epithelial mesenchymal transition (EMT) of biliary atresia mice caused by Rhesus rotavirus (RRV) infection.Methods:The biliary atresia model in mice was generated by RRV infection.Mice were divided into normal group, model group, Gli1 overexpression group, Gli1 shRNA group, Gli2 overexpression group and Gli2 shRNA group.Real-time fluorescence quantitative polymerase chain reaction and Western blot were used to detect the mRNA and protein expressions of regulatory factors for EMT (Snail/Slug) and characteristic cytokines of EMT [Vimentin, α-smooth muscle actin(α-SMA), E-cadherin] in mouse liver tissues.Additionally, hematoxylin-eosin staining and Masson staining were performed to calculate the percentage of liver fibrous tissue expression area.The data were analyzed by One- Way ANOVA and LSD- t test. Results:The relative mRNA expression of Snail, Slug, Vimentin, α-SMA and E-cadherin in Gli2 overexpression group, Gli2 shRNA group and model group were 15.13±3.40, 5.48±0.46, 8.78±1.06, 12.40±2.18 and 3.06±0.53; 3.73±1.16, 5.62±1.75, 3.56±1.06, 3.88±1.16 and 10.51±1.83; 8.13±1.27, 5.32±0.98, 5.05±0.98, 4.02±0.77 and 5.12±1.60.Compared with those of the model group, mRNA levels of Snail, Vimentin and α-SMA were significantly higher in Gli2 overexpression group, while that of E-cadherin was significantly lower( t=4.53, 5.29, 8.12, -2.13; all P<0.05); compared with those of the model group, mRNA levels of Snail and Vimentin in Gli2 shRNA group significantly decreased, while that of E-cadherin significantly increased( t=-2.86, -2.12, 5.62; all P<0.05). In Gli2 overexpression group, Gli2 shRNA group and model group, the protein levels of Snail, Slug, Vimentin, α-SMA and E-cadherin were 2.02±0.39, 0.31±0.08, 0.95±0.17, 1.07±0.17 and 0.42±0.06; 0.53±0.13, 0.40±0.18, 0.20±0.04, 0.28±0.07 and 1.09±0.31; 0.70±0.15, 0.42±0.22, 0.64±0.13, 0.81±0.11 and 0.42±0.09.Compared with those of the model group, protein levels of Snail, Vimentin and α-SMA were significantly higher in Gli2 overexpression group( t=12.71, 4.28, 3.70; all P<0.05); compared with those of the model group, protein levels of Vimentin and α-SMA in Gli2 shRNA group significantly decreased, while that of E-cadherin significantly increased( t=-6.14, -7.57, 5.96; all P<0.05). However, no significant change trend were detected in expression levels of characteristic cytokines of EMT between Gli1 overexpression group and Gli1 shRNA group.The area percentage of liver fiber expression in normal group, model group, Gli1 overexpression group, Gli1 shRNA group, Gli2 overexpression group and Gli2 shRNA group were (1.03±0.58)%, (33.02±11.39)%, (39.81±5.67)%, (26.06±1.29)%, (49.81±8.57)% and (17.55±0.66)%, respectively.Besides, in terms of percentage of area expressed in liver fiber tissue, the Gli2 overexpression group and Gli2 shRNA group were statistically significant compared with the model group( t=3.21, -2.96; all P<0.05), while the Gli1 overexpression group and Gli1 shRNA group were not statistically significant compared with the model group (all P>0.05). Conclusions:The Shh signaling pathway plays an important role in liver fibrosis in mice with biliary atresia.Gli2, a key transcription factor of Shh signaling pathway, can significantly regulate liver EMT process in mice with biliary atresi.

Role of Shh/Gli1 signaling pathway in sleep deprivation-induced cognitive impairment in young mice / 中华麻醉学杂志

Objective:To evaluate the role of sonic hedgehog (Shh)/glioma-associated oncogene homolog 1 (Gli1) signaling pathway in sleep deprivation-induced cognitive impairment in young mice.Methods:Forty-eight SPF healthy male C57BL/6 mice, aged 4 weeks, weighing 14-16 g, were divided into 3 groups ( n=16 each) by the random number table method: control group (C group), sleep deprivation group (SD group) and Shh agonist SAG group (SD+ SAG group). Multi-platform water environment method was used to prepare the sleep deprivation model in mice, and the sleep deprivation was 20 h a day for 10 consecutive days.In SD+ SAG group, SAG 10 mg/kg was intraperitoneally injected at 5 min before each sleep deprivation, while the equal volume of normal saline was intraperitoneally injected in group C and group SD.The mice underwent novel object recognition and Y-maze tests at 24 h after development of the model.Mice were sacrificed after the behavioral testing, and the hippocampi were isolated for determination of the density of dendritic spines in hippocampal CA1 region (by Golgi staining), expression of Gli1 and brain-derived neurotrophic factor (BDNF) in hippocampal tissues (by Western blot), and expression of Gli1 and BDNF mRNA in hippocampal tissues (by quantitative real-time polymerase chain reaction). Results:Compared with group C, the preference index in novel object recognition and Y-maze tests and density of dendritic spines in CA1 region were significantly decreased, and the expression of Gli1 and BDNF protein and mRNA in hippocampus was down-regulated in group SD ( P<0.05). Compared with group SD, the preference index in novel object recognition and Y-maze tests and density of dendritic spines in CA1 region were significantly increased, and the expression of Gli1 and BDNF protein and mRNA in hippocampus was up-regulated in group SD+ SAG ( P<0.05). Conclusions:Inhibition of Shh/Gli1 signaling pathway and reduction of plasticity of dendritic spines of hippocampal neurons are involved in sleep deprivation-induced cognitive impairment in young mice.

Effects of SET7 on angiotensin II-mediated proliferation and collagen synthesis of myocardial fibroblasts and its mechanisms / 中南大学学报(医学版)

OBJECTIVES@#Silence of SET domain containing lysine methyltransferase 7 (SET7) alleviates myocardial tissue injury caused by ischemia-reperfusion. But the effects of SET7 on angiotensin II (Ang II)-induced myocardial fibroblast proliferation and the collagen synthesis are not clear. The purpose of this study was to explore the effect of SET7 on the proliferation and collagen synthesis of myocardial fibroblasts and its mechanisms.@*METHODS@#Myocardial fibroblasts were isolated and identified by immunofluorescence. Myocardial fibroblasts were randomly divided into 4 groups: a control group (cells were normally cultured), an Ang II group (cells were treated with 100 nmol/L Ang II for 24 h), a siCtrl group (cells were transfected with siRNA control and were then treated with 100 nmol/L Ang II for 24 h), and a siSET7 group (cells were transfected with siRNA SET7 and were then treated with 100 nmol/L Ang II for 24 h). Cell counting kit-8 (CCK-8) and 5-ethynyl-2'-deoxyuridine (EdU) assay were used to evaluate cell proliferation. Real-time PCR was used to detect the mRNA levels of SET7, collagen I, collagen III, and α-smooth muscle actin (α-SMA). Western blotting was used to detect the protein expression of SET7, collagen I, collagen III, α-SMA, sonic hedgehog (Shh), ptched1 (Ptch1), and glioma-associated oncogene homolog 1 (Gli1).@*RESULTS@#Fluorescence microscopy showed positive vimentin staining, and myocardial fibroblasts were in good condition. As compared to the control group, the mRNA and protein levels of SET7 in the Ang II group were significantly upregulated; cell proliferation rate and EdU fluorescence intensity in the Ang II group were significantly increased; the mRNA and protein levels of collagen I, collagen III, and α-SMA were significantly upregulated (all @*CONCLUSIONS@#Silence of SET7 gene inhibits Ang II-induced proliferation and collagen synthesis of myocardial fibroblasts. Shh signaling pathway may be involved in this process.

Canal Cleanliness and Smear Layer Removal Using Different Irrigant Activation Methods - A Scanning Electron Microscopy Study

rrigation is one the most important aspects during root canal treatment of the teeth which can be achieved by mechanical cleaning and shaping with the aid of irrigants. However, the irrigant does not travel to all the places of the root canal especially the apical third. Hence, to make this irrigant reach the apical third, we need irrigant activation methods which will agitate the irrigant and help in accessing the places which are difficult to reach in the root canal system. The objective of this research was to check the canal cleanliness and the removal of the debris after irrigant activation using manual dynamic agitation (MDA), plastic F file, sonic irrigation (SI) and conventional syringe irrigation (CSI). METHODSSixty single rooted teeth were chosen for this study which were biomechanically prepared using ProTaper system (Dentsply Maillefer, USA) up to a preparation of F2 and 3 % NaOCl and 17 % EDTA were used as irrigants. All the samples were equally divided into groups of fifteen each depending on the mode of irrigant activation method used - Group 1, Manual Dynamic Agitation (MDA); Group 2, plastic F file; Group 3, EndoActivator (SI); and Group 4, control group (C). These teeth were then split along the long axis and were observed under the SEM for any debris and to determine the degree of canal cleanliness. RESULTSGroup 4 (control group) showed the maximum debris under SEM with a statistically significant difference with a P value less than 0.05; next was the manual dynamic agitation group. Plastic F file group and sonic irrigation groups showed almost similar results in terms of debris. CONCLUSIONSIrrigant activated using sonic mode and plastic F file efficiently removed the debris in comparison to the other two groups of syringe irrigation and manual dynamic agitation.

Removal Techniques for Intracanal Medicament- A Review

This article reviews irrigation techniques for removal of intracanal medicament in endodontic practice. Microorganisms are the primary etiological factors for pulpal and periradicular diseases. So primary purpose is to completely eradicate microorganism from the root canal. It is done through chemo-mechanical preparation of the canal. Complete disinfection of the pulp space cannot be achieved with most sophisticated instrumentation techniques. Therefore use of inter appointment intracanal medicaments is mandatory. Removal of the medicament is mandatory, as its remnants may mechanically block the apical area of the root canal system. Also affects viscosity, working time, tubule penetration and adhesion of root canal sealers. Remnants of Ca(OH)2 in the canal react with unreacted eugenol present in ZOE based sealer to form calcium eugenolate. Today’s irrigation armamentarium presents a diverse variety of tools and techniques , that can assist the practitioner in reducing bacteria, debris, intracanal medicament within the canal system. Conventional syringe irrigation is a routinely practiced method for removal of medicament. It consists of delivering the irrigant in the canal passively or by agitation. Rotary brush does not actually render irrigating solution for removal of medicament. This acts like auxiliaries during removal of medicament from canal or for increased movement of irrigating solution. Ultrasonic irrigation is done with or without simultaneous ultrasonic instrumentation. EndoVac is negative pressure irrigation, which can be used as an alternative method that helps in safe removal of medicament in apical thirds. RinsEndo is also based on pressure alteration technology like EndoVac. Sonically driven system safely acti­vates various intracanal reagents and vigorously produces the hydrodynamic phenomenon as it includes EndoActivator and Vibringe. Laser activated irrigation is more effective for cleaning of root canal. Er:YAG is most commonly used laser in endodontics. Therefore, the aim of this article is to highlight the irrigation techniques used for removal of the intracanal medicament in endodontic practice.

Efficacy of diode laser and sonic agitation of Chlorhexidine and Silver-nanoparticles in infected root canals / Eficácia do laser de diodo e agitação sônica de clorexidina e nanopartículas de prata em canais radiculares infectados

Objective: To assess the efficacy of agitation of chlorohexidine (CHX) and Silver nanoparticles "AgNps" with 810nm diode laser or sonic endoactivator compared to side ­vented needle on infected root canals with Enterococcus "E" Faecalis biofilms. Material and Methods: Sixty-five extracted human premolars with single oval canals were instrumented by protaper system up to F3. Biofilms of E. faecalis were generated based on a previously established protocol. Two teeth were used to check the biofilm formation, then the remaining Teeth were randomly divided into three equal experimental groups according to agitation techniques used: group 1 (810 nm diode laser with 1 watt) , group 2 (sonic endoactivator) and group 3 (Side vented needle). Each group was further divided into three equal subgroups according to the irrigant solution into; subgroup A: chlorohexidine, subgroup B: silver nanoparticles and subgroup C: distilled water: Confocal laser scanning microscopy "CLSM" was used to assess bacterial viability. Data were analyzed by appropriate statistical analyses with P = 0.05. Results: Regarding the activation method, all groups had a significantly high percentage of dead bacteria (P < 0.05). However, Laser was significantly the highest and Endoactivator the least (P < = 0.001). Diode laser agitation of AgNps irrigant showed the highest reduction percentage of bacteria (78.1%) with a significant difference with both CHX and water irrigation, Conclusion: Under the condition of the present study; results reinforced that laser activation is a useful adjunct, 810 nm diode laser agitation of AgNps or chlorhexidine was more effective in disinfection of oval root canals than endoactivator and side vented needle techniques. (AU)

Objetivo: Avaliar a eficácia da agitação de clorohexidina (CHX) e nanopartículas de prata (AgNps) , com laser de diodo de 810 nm ou endoativador sônico, em comparação à agulha de ventilação lateral, em canais radiculares infectados com biofilmes de Enterococcus "E"; Faecalis. Material e Métodos: Sessenta e cinco pré-molares humanos com um único canal oval, extraídos, foram instrumentados pelo sistema protaper até F3. Os biofilmes de E. faecalis foram gerados com base em um protocolo previamente estabelecido. Foram utilizados dois dentes para verificar a formação do biofilme, e os dentes restantes foram divididos aleatoriamente em três grupos experimentais iguais, de acordo com as técnicas de agitação utilizadas: grupo 1 (laser de diodo 810 nm com 1 watt), grupo 2 (endoativador sônico) e grupo 3 (Agulha com ventilação lateral). Cada grupo foi dividido em três subgrupos iguais, de acordo com a solução irrigante; subgrupo A: clorohexidina, subgrupo B: nanopartículas de prata e subgrupo C: água destilada: A microscopia confocal de varredura a laser foi usada para avaliar a viabilidade bacteriana. Os dados foram analisados por análises estatísticas apropriadas com P = 0,05. Resultados: Em relação ao método de ativação, todos osgrupos apresentaram percentual significativamente alto de bactérias mortas (P < 0.05). No entanto, para o laser foi significativamente o mais alto e, para oendoativador, o menos alto (P < = 0.001). A agitação com laser de diodo doirrigante AgNps apresentou a maior porcentagem de redução de bactérias (78,1%), com diferença significativa tanto para irrigação com CHX quanto comágua. Conclusão: Sob as condições do presente estudo; os resultadosreforçaram que a ativação a laser é um complemento útil, a agitação por laserde diodo de 810 nm de AgNps ou clorexidina foi mais eficaz na desinfecção dos canais radiculares ovais do que as técnicas de endoativador e agulha com ventilação lateral. (AU)

Sonic hedgehog-polydopamine-fibrin scaffold promotes recovery of spinal cord injury in rats / 中国组织工程研究

BACKGROUND: Recently, most studies have combined tissue engineering materials with stem cells or factors to improve the microenvironment of animal models of spinal cord injury to increase the duration of action, improve the recovery effect and prognosis. OBJECTIVE: To investigate the effect of sonic hedgehog-polydopamine-fibrin scaffold on the repair of spinal cord injury in rats. METHODS: Fibrin glue was made using a vacuum freeze-dryer. The prepared fibrin glue was immersed in a dopamine hydrochloride solution for 24 hours for cross-linking. Then the cross-linked scaffold was placed in a factor solution for adsorption and cross-linking for 24 hours. Sonic hedgehog-polydopamine-fibrin scaffolds were prepared. Sixty female SD rat models of spinal cord injury were established and then divided into four groups: In the group A, no material was implanted. In the groups B, C and D, fibrin scaffolds, polydopamine-fibrin scaffolds, and sonic hedgehog-polydopamine-fibrin scaffolds were implanted respectively. The Basso, Beattie and Bresnahan (BBB) locomotor scale score of lower limb locomotor function was evaluated within 12 weeks after surgery. At 12 weeks post-surgery, the tissue at the site of spinal cord injury was collected for histological observation (hematoxylin-eosin and immunohistochemical staining) and western blot assay. This study was approved by the Animal Ethics Committee of Jiangsu University, China. RESULTS AND CONCLUSION: (1) From 2 weeks after surgery, the lower limb locomotor function of rats in each group began to recovery. At 5-12 weeks after surgery, the BBB score of group D was significantly higher than that of the other three groups (P < 0.05). Rats in group D had the best recovery of locomotor function of the lower limb. (2) Hematoxylin-eosin staining revealed newly generated nerve fibers in the groups C and D, and that the number of density of new nerve fibers in group C was lower than that in group D. (3) Immunohistochemical staining showed that a large amount of linearly arranged new nerve fibers were observed in the completely transected site of rat spinal cord. In group D, myelin basic protein-, growth related protein- and neurofilament protein-positive rates were significantly higher (P < 0.05), and glial fibrillary acidic protein-positive rate was significantly lower, compared with the other three groups. (4) Western blot assay revealed that in group D, the protein expression of myelin basic protein, growth related protein and neurofilament protein was significantly higher (P < 0.05), and the protein expression of glial fibrillary acidic protein was significantly lower (P < 0.05), compared with the other three groups. (5) These results suggest that sonic hedgehog-polydopamine-fibrin has a good sustained-release performance, which can greatly promote the repair of spinal cord injury in rats.

Lycium barbarum polysaccharide inhibits the proliferation and promots the apoptosis of osteosarcoma HOS cells by blocking Shh signaling pathway / 中国临床药理学与治疗学

AIM: To study the effects and mechanism of Lycium barbarum polysaccharide on the proliferation and apoptosis of osteosarcoma HOS cells. METHODS: Osteosarcoma HOS cells were divided into four groups: control group, LBP-I group, LBP-II group and LBP-III group. MTT was used to detect cell proliferation; PI single staining was used to detect cell cycle; Annexin V-FITC/PI double staining was used to detect apoptosis; Western blot was used to detect the expression of Cleaved Caspase-3, Cleaved PARP, cyclin-dependent kinase 4 (CDK4), cyclin D1, Shh and Gli1. Shh signal activator and Lycium barbarum polysaccharide treated osteosarcoma HOS cells together; the changes of cell proliferation, cell cycle and apoptosis were observed.RESULTS: Compared with the control group, the proliferation ability of LBP-I group, LBP-II group and LBP-III group decreased, the proportion of cells in G0/G1 phase increased[(51.2±4.1)% vs. (59.1±3.2)%, (66.8±2.0)%, (72.3±3.2)%, F=72.76, P<0.001], the level of apoptosis increased[(3.9±0.3)% vs. (13.2±1.2)%, (17.6±1.3)%, (24.8±2.1)%, F=364.50, P<0.001], the expression levels of Cleaved Caspase-3, Cleaved PARP protein increased, the expression levels of CDK4, cyclin D1, Shh and Gli1 decreased (P<0.05). Compared with cells not treated with Shh signal activator, the cells treated with Shh signal activator could reverse the effect of Lycium barbarum polysaccharide on the proliferation, cycle arrest and apoptosis of osteosarcoma HOS cells. CONCLUSION: Lycium barbarum polysaccharides blocked the cell cycle of osteosarcoma HOS cells, inhibited cell proliferation and promoted cell apoptosis by inhibiting Shh signaling pathway.

Desarrollo embriofetal del iris: inmunotinción del morfógeno Shh / Embryo-fetal development of iris: immunohistochemestry of morphogen Shh

RESUMEN: Para que se desarrolle el iris, se requiere una especificación de la capa periférica de la copa óptica a un destino no neuronal y además la migración de células mesenquimales perioculares. Nuestro objetivo fue reconocer los cambios histológicos de los derivados periféricos de la copa óptica y mesénquima periocular, como también reconocer la presencia del morfógeno Sonic hedgehog (Shh) en las capas que constituyen el esbozo de iris. Se utilizaron 15 ratones hembras (Mus musculus) adultas jóvenes gestantes. Se realizó eutanasia con tiopental sódico. Los embriones y fetos de 12, 14,5 y 17 días post-coital (dpc) fueron procesados con técnica histológica e inmunohistoquímica con anticuerpo anti-Shh (scbt, H-160, conejo) con dilución 1:100 en PBS. A los 12 dpc, se observa una cópa óptica que presenta capas retinianas interna y externa, y el iris no se observa. Entre el cristalino y el ectodermo superficial se identifican 4 capas de células mesenquimales. A los 14,5 dpc, el iris contiene dos capas epiteliales (interna y externa) que se continúan con las capas neural y pigmentaria de la retina. Se observan 8 capas de células mesenquimales. A los 17 dpc, la capa epitelial interna del iris presenta un segmento más elongado con inmunotinción positiva a Shh y otra parte que constituye un epitelio de células cilíndricas simples negativas a este anticuerpo. La capa epitelial externa presenta el mismo epitelio inmunonegativo. Las capas de la retina también son positivas, como también la periferia del cristalino. No esta formado el iris ni tampoco el cuerpo ciliar. La inmunopositividad en el cristalino, en el primer segmento de la capa interna del esbozo del iris y en la capa ganglionar retinal a los 17 dpc, se relaciona con la diferenciación tardía del iris y con los ojos cerrados de las crías al nacimiento.

SUMMARY: In order for the iris to develop, a specification of the peripheral layer of the optic cup to a non-neuronal target is required, as well as the migration of periocular mesenchymal cells. Our aim was to recognize the histological changes of peripheral derivatives of the optic cup and periocular mesenchyme, as well as recognize the presence of the morphogen Sonic hedgehog (Shh) in the layers constituting the outline of the iris. 15 female mice (Mus musculus) pregnant young adults were used. Euthanasia was performed with sodium thiopental. Embryos and fetuses of 12, 14.5 and 17 days post-coital (dpc) were processed with histological and immunohistochemical technique with anti-Shh antibody (scbt, H 160, rabbit) with dilution 1:100 in PBS. At 12 dpc, an optic cup showing internal and external retinal layers is observed, and the iris is not observed. Between the lens and the superficial ectoderm, 4 layers of mesenchymal cells are identified. At 14.5 dpc, the iris contains two epithelial layers (internal and external) that are continued with the neural and pigmentary layers of the retina. 8 layers of mesenchymal cells are observed. At 17 dpc, the inner epithelial layer of the iris presents a more elongated segment with positive immunostaining to Shh and another part that constitutes an epithelium of simple cylindrical cells negative to this antibody. The outer epithelial layer presents the same immunonegative epithelium. The layers of the retina are also positive, as well as the periphery of the lens. The iris is not formed nor is the ciliary body.The immunopositivity in the lens, in the first segment of the inner layer of the iris outline and in the retinal ganglion layer at 17 dpc, is related to the late differentiation of the iris and the closed eyes of the offspring at birth.

Comparación de tres técnicas de irrigación en la remoción de hidróxido de calcio / Comparison of three irrigation techniques in the removal of calcium hydroxide

Objetivos: Comparar la eficacia de tres de protocolos de irrigación en la remoción de hidróxido de calcio (Ca(OH)2) Material y Métodos: Mediante un diseño de estudio comparativo in vitro, se instrumentaron 106 conductos radiculares de incisivos bovinos hasta una lima de diámetro 60, la raíces se fraccionaron en dos mitades siguiendo el eje mayor del diente y se creó un surco estandarizado a 2 mm del agujero apical que fue rellenado con una pasta de hidróxido de calcio, luego se reensamblaron las mitades, se incubaron por 7 días y se realizaron los protocolos de irrigación: ultrasónica pasiva (PUI), sónica con Endo Activator (EA) y activación dinámica manual (MDA), donde se usó como irrigante hipoclorito de sodio (NaOCl) al 5%. Se utilizó un control negativo y positivo. Con un estereomicroscópio se observó la cantidad de residuo, las imágenes se examinaron y se les asignó una puntación de acuerdo a la escala de van der Sluis, finalmente los datos se analizaron con la prueba estadística del Chi Cuadrado. Resultados: Los porcentajes de eficacia para los protocolos de irrigación PUI, EA y MDA fueron del 87,5%; 46,9% y 28,1% respectivamente, la técnica PUI fue superior a EA y MDA y obtuvo diferencias estadísticamente significativas (p<0,001). Conclusiones: PUI fue el método más efectivo en la remoción de Ca(OH)2 de los surcos simulados en los conductos radiculares.

Objectives: To compare efficacy of three irrigation protocols in the removal of calcium hydroxide (Ca(OH)2). Materials and Methods: Through an in vitro comparative study design, 106 root canals of bovine incisors were instrumented up to a file of diameter 60, the roots were fractioned into two halves following the major axis of the tooth and a standardized groove was created 2mm from the apical foramen that was filled with a paste of calcium hydroxide, then the halves were reassembled, incubated for 7 days and the irrigation protocols were carried out: passive ultrasonic (PUI), sonic with Endo Activator (EA) and manual dynamic activation (MDA), where 5% sodium hypochlorite (NaOCl) was used as irrigant. A negative and positive control was used. With a stereomicroscope the amount of residue was observed, the images were examined and assigned a score according to the van der Sluis scale, finally the data were analyzed with the statistical test of Chi-square. Results: The percentages of efficacy for PUI, EA and MDA irrigation protocols were 87.5%; 46.9% and 28.1% respectively, the PUI technique was superior to EA and MDA and obtained statistically significant differences (p <0.001). Conclusions: PUI was the most effective method in the removal of Ca(OH)2 from the simulated grooves in the root canals.

Inmunolocalización de Sonic hedgehog en el desarrollo embrio-fetal de ratones ( Mus musculus ) / Immunolocalization of Sonic hedgehog in embryo-fetal mouse ( Mus musculus ) development

Sonic hedgehog (Shh) es un morfógeno esencial para el desarrollo de diversas estructuras, tales como notocorda, placa del piso del tubo neural, miembros, entre otros. Se buscó determinar la inmunolocalización de Shh en embriones y fetos de ratón. Para ello, se eutanasiaron 10 ratones gestantes (Mus musculus) BALB/c, un grupo de 5 animales a los 12,5 días post-coito (dpc), y otro grupo a los 17,5 dpc. Los embriones y fetos obtenidos fueron fijados en formalina al 10 % tamponada en PBS e incluidos en paraplast. Se realizaron cortes transversales seriados. Se utilizó anticuerpo policlonal Shh (Santa Cruz Biotechnology, H-160, conejo), dilución 1:100. Se identificó y describió la inmunolocalización de las muestras marcadas positivamente. La expresión de Shh en los embriones de 12,5 dpc fue inmunopositiva en notocorda, placa del piso del tubo neural, precartílago de radio y ulna, y prácticamente todos los epitelios: bronquial, intestinal, vejiga y uretra. En la etapa fetal, a los 17,5 dpc la inmunopositividad desaparece en el cartílago a excepción de zonas de osificación, disminuye en la epidermis pero aparece en folículos pilosos. La mucosa intestinal se ha diferenciado en segmentos, mostrando una inmunotinción mayor a nivel de las vellosidades intestinales. Shh actúa en distintos estadios del periodo gestacional, siendo clave en la diferenciación de distintas estructuras. En etapas embrionaria, es vital en la formación del sistema nervioso, organogénesis y formación de miembros, por lo que su expresión se encuentra en estas zonas. Sin embargo, en la etapa fetal la expresión cambia a estructuras de mayor especialización como folículo piloso y vellosidades intestinales.

Sonic hedgehog (Shh) is an essential morphogen for the development of various structures, such as notochord, neural tube floor plate, limbs, among others. We sought to determine the immunolocalization of Shh in embryos and mouse fetuses. To do this, 10 pregnant mice (Mus musculus) BALB /c were euthanized, a group of 5 animals at 12.5 days postcoitus (dpc), and another group at 17.5 dpc. Embryos and fetuses obtained were fixed in 10 % formalin buffered in PBS and embedded in paraplast. Serial cross sections were made. Polyclonal antibody Shh (Santa Cruz Biotechnology, H-160, rabbit), dilution 1:100 was used. The immunolocalization of the positively labeled samples was identified and described. Shh expression in 12.5 dpc embryos was immunopositive in notochord, neural tube floor plate, radius precartilage and ulna, and practically all epithelia: bronchial, intestinal, bladder and urethra. In the fetal stage, at 17.5 dpc the immunopositivity disappears in the cartilage except for areas of ossification, decreases in the epidermis but appears in hair follicles. The intestinal mucosa has differentiated into segments, showing greater immunostaining at the level of the intestinal villi. Shh acts in different stages of the gestational period, being key in the differentiation of different structures. In embryonic stages, it is vital in the formation of the nervous system, organogenesis and formation of limbs, so its expression is found in these areas. However, in the fetal stage the expression changes to more specialized structures such as hair follicles and intestinal villi.

Effect of sonic versus ultrasonic activation on aqueous solution penetration in root canal dentin

The aim of this study was to observe the penetration of an aqueous solution into the root canal dentin under sonic activation and ultrasonic activation. Materials and Method: this study consisted of experimental in vitro research. In order to achieve a closed system, the apex of 45 single-rooted teeth was sealed with wax. The step-back technique was manually performed using a K50 apical master file and 3 groups were organized according to the protocol of the final irrigant activation: group I: non-activated chinese ink for 30 seconds, group II: chinese ink sonically activated with EndoActivator for 30 seconds, and group III: chinese ink ultrasonically activated with Varios 350 equipment for 30 seconds. Teeth were sectioned longitudinally, and the samples obtained were observed under a stereomicroscope at 1X magnification in order to be photographed and scanned to calculate the penetration area using the Image J software. The tinted radicular area was evaluated in relation to the total area of the root dentin. The tukey's post-hoc test and ANOVA were used for the statistical analysis (p<0.05). Results: group I and II obtained 9.13 percent and 9.42 percent penetration respectively, while in group III the highest degree of dye infiltration was achieved (13.9 percent), being statistically significant (p<0.001). Conclusions: ultrasonic activation produced a significantly higher penetration of the dye when compared to conventional activation and sonic activation.

Progress of targeting inhibition of Sonic Hedgehog signaling pathway in leukemia / 白血病·淋巴瘤

Sonic Hedgehog (SHH) signaling pathway participates in the regulation of various organs and growth of tissue cells, maintains normal function and structure in the development of embryogenesis, however, SHH signaling pathway is in a state of inhibition. Meanwhile, the abnormal regulation of SHH signaling pathway plays an important role in the occurrence, development and drug resistance of leukemia. The mechanism of SHH signaling pathway in leukemia is similar to the solid tumor. With the further understanding of SHH signaling pathway, there are many antitumor drugs targeting SHH signaling pathway. The targeting inhibitors targeted SHH signaling pathway will become a new method for the treatment of leukemia. This paper reviews the application of the inhibitors targeting SHH signaling pathway in leukemia.

Activation of Sonic hedgehog signaling in ventricular cardiomyocytes exerts DNA protection against hypoxia / 中国药理学通报

Aim To investigate the protective roles of sonic hedgehog( Shh) signaling pathway in hypoxia-in-duced DNA damage with the neonatal rat cardiomyo-cytes. Methods The hypoxia model on neonatal car-diomyocytes was established with one to two days old Sprague Dawley rats by deprivation of oxygen and glu-cose ( OGD) . After pretreated with Shh pathway ago-nist SAG1.3 or antagonist GANT61, the survival rates of cardiomyocytes were assayed by MTT after OGD 6 hours or 12 hours. The protein levels of Shh pathway, phosphorylated histone H2AX at serine 139 (γH2AX), phosphorylated ATM (p-ATM), phospho-rylated p53 ( p-p53 ) , cleaved-caspase-3, Bcl-2 and Bax were detected by Western blot. The γH2AX foci was detected by immunofluorescence. Results Com-pared to control group, the protein expression of γH2AX, p-ATM, cleaved-caspase-3, p-p53 in OGD cardiomyocytes significantly increased, and Bcl-2/Bax ratio proportionally decreased. Particularly, the ex-pression of γH2AX, p-ATM was highest at OGD 6 h, and then gradually declined after OGD 12 h. After SAG1.3 pretreatment, the expression of γH2AX, p-ATM, cleaved-caspase-3 and p-p53 dramatically de-creased and the Bcl2/Bax ratio increased in OGD 6 h or OGD 12 h cardiomyocytes. On the contrary, in GANT61 pretreatment group, the expression of γH2AX, p-ATM, cleaved-caspase-3 and p-p53 signifi-cantly increased and the Bcl-2/Bax ratio decreased compared to the OGD 6 h or OGD 12 h cardiomyo-cytes. Conclusion The activation of Shh pathway protects cardiomyocytes against hypoxia-induced apop-tosis through inhibition of DNA damage.

The study on the relationship between the levels of sonic hedgehog and vascular endothelial growth factor in serum and cerebral collateral circulation in patients with symptomatic cerebral artery stenosis / 中国医师进修杂志

Objective To investigate the changes of the levels of sonic hedgehog (SHH) and vascular endothelial growth factor (VEGF) in serum and its relationship with collateral circulation in patients with symptomatic middle cerebral artery stenosis. Methods From January 2015 to January 2018, a total of 268 patients with acute ischemic stroke confirmed as unilateral middle cerebral artery M1 segment (MCA-M1) severe stenosis or occlusion by digital subtract angiography (DSA) were enrolled. The baseline clinical data were collected. According to the establishment of collateral circulation shown by DSA, they were divided into good collateral circulation group (152 patients) and poor collateral circulation group (116 patients). The levels of SHH and VEGF in serum were detected by enzyme linked immunosorbent assay (ELISA), the expression characteristics of SHH and VEGF in serum and the relative factors influencing the establishment of collateral circulation were analyzed. Results The levels of serum SHH and VEGF in good collateral circulation group were significantly higher than those in poor collateral circulation group (P < 0.01). Pearson correlation analysis showed that there was a positive correlation between SHH and VEGF (r=0.758, P < 0.01). Multivariate Logistic regression analysis showed that the levels of serum SHH ( OR=0.310, 95% CI 0.117-0.819, P=0.018) and VEGF ( OR=0.361, 95% CI 0.147-0.887, P=0.026) were independent protective factors for the establishment of collateral circulation. Diabetes ( OR=3.094, 95% CI 1.321-7.245, P=0.009) was independent risk factor for the establishment of collateral circulation. Conclusions The levels of serum SHH and VEGF are closely related to the formation of collateral circulation and they are independent protective factors. SHH may be involved in the establishment of cerebral collateral circulation by regulating the expression of VEGF and diabetes is not conducive to the formation of collateral circulation.