Grape seed extract inhibits apoptosis of rat growth plate chondrocytes and promotes tibial bone growth / 中国组织工程研究

BACKGROUND:Grape seed extract can inhibit chondrocyte apoptosis and aging,and improve osteoarthritis.However,the effects of grape seed extract on the apoptosis of chondrocytes in the growth plate and tibial growth are still unclear. OBJECTIVE:To investigate the effect of grape seed extract on interleukin-1β-induced apoptosis in rat growth plate cells and on tibial bone growth. METHODS:(1)Cell experiment:Growth plate chondrocytes from Sprague-Dawley rats were isolated,cultured,and identified.The cells were then randomly divided into control group,model group,grape seed extract group,miR-138-5p NC group and miR-138-5p inhibitor group.In the model group,20 ng/mL interleukin-1β was used to induce apoptosis in rat growth plate chondrocytes.In the grape seed extract group,20 ng/mL interleukin 1β was added along with 10 μmol/L grape seed extract solution for 48 hours.Cells in the miR-138-5p NC and inhibitor groups were transfected with 5 nmol/L miR-138-5p NC and 5 nmol/L miR-138-5p inhibitor for 12 hours,respectively,followed by addition of 20 ng/mL interleukin-1β.qRT-PCR was used to detect miR-138-5p and caspase-3 expression.Luciferase reporter assay was used to analyze the relationship between miR-138-5p and caspase-3 targeting.Cell counting kit-8 was used to detect cell proliferation activity.Flow cytometry was used to detect cell apoptosis.Caspase-3 and Bcl-2 protein expressions were detected by western blot.(2)Animal experiment:The animals were divided into normal control group,grape seed extract group and miR-138-5p inhibitor group.The effects of grape seed extract on epiphyseal closure and tibial growth of the tibial plateau in rats were observed. RESULTS AND CONCLUSION:miR-138-5p had a targeting relationship with caspase-3.Compared with the control group,cell proliferation was significantly reduced,apoptosis was significantly increased(P＜0.01),miR-138-5p,Bcl-2 expression was reduced(P＜0.01),and caspase-3 expression was increased(P＜0.01)in the model group.Compared with the mod group,the grape seed extract group showed a significant increase in cell proliferation,a significant decrease in apoptosis(P＜0.01),an increase in miR-138-5p and Bcl-2 expression(P＜0.01)and a decrease in caspase-3 expression(P＜0.01).Compared with the grape seed extract group,the miR-138-5p inhibitor group showed a significant decrease in cell proliferation,a significant increase in apoptosis(P＜0.01),a decrease in miR-138-5p and Bcl-2 expression(P＜0.05 or P＜0.01),and an increase in caspase-3 expression(P＜0.05).Intragastric administration of grape seed extract could delay epiphyseal closure of the tibial plateau and promote tibial bone growth in rats,whereas miR-138-5p inhibitor intervention inhibited the effect of grape seed extract on tibial bone growth in rats.To conclude,grape seed extract can inhibit apoptosis of rat growth plate chondrocytes through regulating the miR-138-5p/caspase-3 pathway,improve epiphyseal closure of rat tibial plateau and promote tibial bone growth.

Grape seed extract inhibits apoptosis in growth plate chondrocytes and promotes tibial growth in rats / 中国组织工程研究

BACKGROUND:Grape seed extract has been shown to be effective in inhibiting the growth of androgen-dependent tumors(e.g.,breast cancer),and thus grape seed extract could theoretically inhibit epiphyseal closure induced by estrogen in late adolescence. OBJECTIVE:To screen the effects of grape seed extract on apoptosis of growth plate chondrocytes and epiphyseal closure in rats. METHODS:(1)In vitro experiment:Growth plate chondrocytes from rat large tibia and femur at logarithmic growth stage were obtained and cultured in groups:normal control group,model control group(adding 17β-estradiol to induce apoptosis),positive control group(adding letrozole and 17β-estradiol),grape seed extract group(adding 17β-estradiol and 10 μg/mL grape seed extract),Caspase-9 inhibitor group(adding 17β-estradiol and Caspase-9 inhibitor),Caspase-9 agonist group(adding 17β-estradiol and Caspase-9 agonist).Cell apoptosis was detected by flow cytometry after 48 hours of culture.(2)In vivo experiment:Thirty 3-month-old Sprague-Dawley rats were randomly divided into model control group,positive control group and low-,medium-and high-dose groups,with five rats in each group.All rats were injected subcutaneously with 17β-estradiol(3 times per week)to establish epiphyseal closure models,followed by intragastric administration of letrozole in positive control group and 0.05,0.2 and 0.8 g/kg grape seed extract in low-,medium-and high-dose groups,respectively,once a day until over 2/3 of the epiphyseal plate in the model control group was closed.The length of the tibia was then observed.Another 18 Sprague-Dawley rats were randomly divided into model control group,positive control group,and medium-dose group,with 6 rats in each group,treated as above for 1.5 continuous months.The expression of Caspase-9 protein in rat growth plate cartilage was detected by western blot. RESULTS AND CONCLUSION:(1)In vitro experiment:17β-estradiol could induce apoptosis in growth plate chondrocytes,and letrozole,grape seed extract,and caspase-9 inhibitors could all inhibit apoptosis in growth plate chondrocytes.(2)In vivo experiment:When more than 2/3 of the epiphyseal plate in the model control group was closed,the number of rats with epiphysis closure in the positive control and medium-dose groups was less than that in the model control group(P＜0.05),and the tibial length was longer than that in the model control group(P＜0.05),and the Caspase-9 protein expression in the tibial growth plate was lower than that in the model control group(P＜0.05).To conclude,the appropriate dose of grape seed extract can effectively inhibit the apoptosis of growth plate chondrocytes and delay epiphyseal closure,which has the potential to promote bone growth.

Low concentrations of grape seed extract maintain osteoblast morphology, cell adhesion, and mineralization

Abstract The increase in life expectancy has led to a higher incidence of osteoporosis, characterized by an imbalance in bone remodeling. Several drugs are used for its treatment, but most promote undesirable side effects. The present investigation evaluated the effects of two low concentrations of grape seed extract (GSE) rich in proanthocyanidins on MC3T3-E1 osteoblastic cells. The cells were cultured in an osteogenic medium and divided into control (C), 0.1 µg/mL GSE (GSE0.1), and 1.0 µg/mL GSE (GSE1.0) groups to evaluate cell morphology, adhesion, and proliferation, in situ alkaline phosphatase (ALP) detection, mineralization and immunolocalization of osteopontin (OPN). The data obtained were analyzed by statistical tests for a significance of 5%. Cell morphology was maintained with both GSE concentrations, whereas cell adhesion significantly increased within three days in all groups. Cell proliferation increased significantly at seven days of culture, followed by a significant decrease in all experimental periods, with no statistical difference among them. In situ detection of ALP and mineralization increased with time, but within each period, no statistical differences among groups were observed. The expression of osteopontin was distributed regularly with more intensity after 24 hours in the GSE0.1 group. After three days, OPN expression was more intense in the control group, followed by GSE0.1 and GSE1.0 groups. Data obtained suggest that low concentrations of GSE do not affect the morphology and may stimulate the functional activity of osteoblastic cells.

Resumo O aumento da expectativa de vida tem levado a uma maior incidência de osteoporose, caracterizada por um desequilíbrio na remodelação óssea. Vários medicamentos são utilizados para o seu tratamento, contudo, a maioria promove efeitos colaterais indesejáveis. A presente investigação avaliou os efeitos de duas baixas concentrações de extrato de semente de uva (GSE) rico em proantocianidinas em células osteoblásticas MC3T3-E1. As células foram cultivadas em meio osteogênico e divididas em grupos controle (C), 0,1 µg/mL de GSE (GSE0.1) e 1,0 µg/mL de GSE (GSE1.0) para avaliar morfologia, adesão e proliferação celular, detecção in situ de fosfatase alcalina (ALP), mineralização e imunolocalização da proteína osteopontina (OPN). Os dados obtidos foram analisados por testes estatísticos para um nível de significância de 5%. A proliferação celular aumentou significativamente aos sete dias de cultura, seguido de uma diminuição significativa em todos os períodos experimentais, sem diferença estatística entre eles. A detecção in situ de ALP e mineralização aumentou com o tempo, mas dentro de cada período não foram observadas diferenças estatísticas entre os grupos. A morfologia celular foi mantida com ambas as concentrações de GSE, enquanto a adesão celular aumentou significativamente aos três dias em todos os grupos. A expressão de osteopontina distribuiu-se regularmente com maior intensidade após 24 horas no grupo GSE0.1. Após três dias, a expressão de OPN foi mais intensa no grupo controle, seguida pelos grupos GSE0.1 e GSE1.0. Os dados obtidos sugerem que baixas concentrações de GSE não afetam a morfologia e podem estimular a atividade funcional das células osteoblásticas.

Grape Seed Extract Attenuates Demyelination in Experimental Autoimmune Encephalomyelitis Mice by Inhibiting Inflammatory Response of Immune Cells / 中国结合医学杂志

OBJECTIVE@#To examine the anti-inflammatory effect of grape seed extract (GSE) in animal and cellular models and explore its mechanism of action.@*METHODS@#This study determined the inhibitory effect of GSE on macrophage inflammation and Th1 and Th17 polarization in vitro. Based on the in vitro results, the effects and mechanisms of GSE on multiple sclerosis (MS)-experimental autoimmune encephalomyelitis (EAE) mice model were further explored. The C57BL/6 mice were intragastrically administered with 50 mg/kg of GSE once a day from the 3rd day to the 27th day after immunization. The activation of microglia, the polarization of Th1 and Th17 and the inflammatory factors such as tumor necrosis factor- α (TNF- α), interleukin-1 β (IL-1 β), IL-6, IL-12, IL-17 and interferon-γ (IFN-γ) secreted by them were detected in vitro and in vivo by flow cytometry, enzyme linked immunosorbent assay (ELISA), immunofluorescence staining and Western blot, respectively.@*RESULTS@#GSE reduced the secretion of TNF-α, IL-1 β and IL-6 in bone marrow-derived macrophages stimulated by lipopolysaccharide (P<0.01), inhibited the secretion of TNF-α, IL-1 β, IL-6, IL-12, IL-17 and IFN-γ in spleen cells of EAE mice immunized for 9 days (P<0.05 or P<0.01), and reduced the differentiation of Th1 and Th17 mediated by CD3 and CD28 factors (P<0.01). GSE significantly improved the clinical symptoms of EAE mice, and inhibited spinal cord demyelination and inflammatory cell infiltration. Peripherally, GSE downregulated the expression of toll-like-receptor 4 (TLR4) and Rho-associated kinase (ROCKII, P<0.05 or P<0.01), and inhibited the secretion of inflammatory factors (P<0.01 or P<0.05). In the central nervous system, GSE inhibited the infiltration of CD45+CD11b+ and CD45+CD4+ cells, and weakened the differentiation of Th1 and Th17 (P<0.05). Moreover, it reduced the secretion of inflammatory factors (P<0.01), and prevented the activation of microglia (P<0.05).@*CONCLUSION@#GSE had a beneficial effect on the pathogenesis and progression of EAE by inhibiting inflammatory response as a potential drug and strategy for the treatment of MS.

Efecto de distintas Concentraciones de un Gel Experimental a Base de Extracto Etanólico de Semillas de Uva (Vitis vinifera L) sobre la Resistencia de Adhesión de Resina Nanohíbrida

El objetivo del estudio es determinar el efecto de distintas concentraciones de un gel experimental a base de extracto etanólico de semillas de uva (Vitis vinifera L) sobre la resistencia a la adhesión de resina nanohíbrida. Se obtuvo el extracto etanólico a base de Semillas de Uva (EESU) por maceración en alcohol al 70 % durante 45 días. Se reconstituyó el principio activo de las semillas de uva en forma de polvo mezclando 200 ml de agua ultrapura, 2 g. de Carbopol 940, 20 g. de glicerol, 20g. de propilenglicol, 1g. de benzoato de sodio, 1.5 de trietanolamina, obteniendo un gel con un pH de 14, en concentraciones de 5 %, 10 % y 15 %. La sustancia madre fue sometida a cromatografía en capa fina HPLC para identificar sus componentes químicos hallando compuestos fenólicos (ácido Gálico), taninos y en menor cantidad flavonoides. No existieron diferencias significativas entre los grupos control y experimentales cuya tracción se realizó inmediatamente al tratamiento de aclaramiento dental, a los 7 días y 14 días. El extracto etanólico de semillas de uva (Vitis Vinífera L) en concentraciones de 5 %, 10 % y 15 % tuvieron el mismo efecto sobre la resistencia a la adhesión de resina nanohíbrida.

The objective of the study is to determine the effect of different concentrations of an experimental gel based on ethanolic extract of grape seeds (Vitis vinifera L) on the resistance to adhesion of nanohybrid resin The ethanolic extract based on Grape Seeds (EESU) by maceration in 70 % alcohol for 45 days. The active principle of grape seeds was reconstituted in powder form by mixing 200 ml of ultrapure water, 2 g. of Carbopol 940, 20 g. glycerol, 20g. of propylene glycol, 1g. of sodium benzoate, 1.5 of triethanolamine, obtaining a gel with a pH of 14, in concentrations of 5 %, 10 % and 15 %. The base substance was subjected to HPLC thin-layer chromatography to identify its chemical components, finding phenolic compounds (Gallic acid), tannins and, to a lesser extent, flavonoids. There were no significant differences between the control and experimental groups whose traction was performed immediately after dental whitening treatment, at 7 days and 14 days. The ethanolic extract of grape seeds (Vitis vinifera L) in concentrations of 5 %, 10 % and 15 % had the same effect on the resistance to the adhesion of nanohybrid resin.

Functional and chemical qualities of Vitis labrusca grape seed oil extracted by supercritical CO 2 / Calidad funcional y química del aceite de semilla de Vitis labrusca extraído mediante CO 2 supercrítico / Qualidade funcional e química do óleo de semente de Vitis labrusca extraído por CO 2 supercrítico

Abstract Grape seed oil, which is usually extracted with highly toxic organic solvents that are harmful to human health, is produced from tons of grape pomace waste, generated during winemaking. Sometimes, this waste is used to make compost or is burnt, which causes environmental contamination. The functional qualities, antioxidant capacity (AC), α-tocopherol and total phenolic compounds content (TPC) of Black Borgoña (Vitis labrusca) grape seed oil, extracted by supercritical CO2, were evaluated. The high content of linoleic acid (ω-6) and monounsaturated fatty acids contributed to the beneficial effect on the functional quality indices, which were 0.20, 0.23, 11.80 for IA, IT and H:H, respectively. In addition, a POV of 6.23 ± 0.08 milliequivalents of peroxide/kg oil and an anisidine index of 2.70 ± 0.05 indicated a good quality oil. Also, a high concentration of a-tocopherol (9.82 ± 0.02 mg/100 g oil) and a high TPC ("4.14 ± 3.24 mg GAE/kg oil) were obtained. This study demonstrated that supercritical CO2 extraction is a suitable method for the delivery of a high-quality grape seed oil.

Resumen El aceite de semilla de uva que generalmente se extrae con disolventes orgánicos altamente tóxicos y perjudiciales para la salud humana, se produce a partir de toneladas de residuos de orujo de uva, generados durante la elaboración del vino. A veces, estos residuos se utilizan para hacer compost o se queman, lo que provoca la contaminación del medio ambiente. Se evaluaron las cualidades funcionales, la capacidad antioxidante (AC), el contenido de a-tocoferol y los compuestos fenólicos totales (TPC) del aceite de semilla de uva Borgoña Negra (Vitis labrusca), extraído mediante CO2 supercrítico. El alto contenido de ácido linoleico (ω-6) y de ácidos grasos monoinsaturados contribuyó al efecto beneficioso sobre los índices de calidad funcional que fueron de 0.20, 0.23, ''.80 para IA, IT y H:H, respectivamente. Además, un POV de 6.23 ± 0.08 miliequivalentes de peróxido/ kg de aceite y un índice de anisidina de 2.70 ± 0.05 indicaban una buena calidad del aceite. También se obtuvo una alta concentración de α-tocoferol (9.82 ± 0.02 mg/100 g de aceite) y un alto TPC ("4.14 ± 3.24 mg de GAE/ kg de aceite). Este estudio demostró que la extracción con CO2 supercrítico es un método adecuado para obtener un aceite de semilla de uva de alta calidad.

Resumo O óleo de semente de uva é geralmente extraído com solventes orgânicos altamente tóxicos que são prejudiciais à saúde humana, é produzido a partir de toneladas de resíduos de bagaço de uva, gerados durante a vinificação. Às vezes, esses resíduos são usados para fazer adubo ou são queimados, o que causa contaminação ambiental. Foram avaliadas as qualidades funcionais, capacidade antioxidante (AC), a-tocoferol e o teor total de compostos fenólicos (TPC) do óleo de semente de uva Borgoña Negra (Vitis labrusca), extraído por CO2 supercrítico. O alto teor de ácido linoleico (ω-6) e ácidos graxos monoinsaturados contribuiu para o efeito benéfico sobre os índices de qualidade funcional que foram 0.20, 0.23, 11.80 para IA, IT e H:H, respectivamente. Além disso, um POV de 6.23 ± 0.08 miliequivalentes de peróxido/ kg de óleo e um índice de anisidina de 2.70 ± 0.05 indicava uma boa qualidade de óleo. Também foi obtida uma alta concentração de α-tocoferol (9.82 ± 0.02 mg/100 g de óleo) e um alto TPC ("4.14 ± 3.24 mg de óleo GAE/ kg). Este estudo mostrou que a extração de CO2 supercrítico é um método adequado para a entrega de um óleo de semente de uva de alta qualidade.

Use of two kinds of antioxidants to restore the bond strength of bleached enamel / 华西口腔医学杂志

OBJECTIVES@#This work aimed to evaluate the ability of two kinds of antioxidants, namely, grape-seed extract and sodium ascorbate, in restoring bond strength at the resin-enamel interface after bleaching.@*METHODS@#Ten groups of samples with 15 teeth per group were prepared for shear-bond-strength test at the resin-enamel interface after bleaching. The groups were as follows: control; no antioxidant; 2.5%, 5%, 10%, or 15% grape-seed extract; and 2.5%, 5%, 10%, or 15% sodium ascorbate. The peak values of shear bond strength when resin was debonded from teeth and the failure modes under a microscope were recorded. Ten other groups of teeth with two teeth per group were prepared and treated in a similar approach before resin bonding. The samples were cut vertically to the bonding interface. The structures of the bonding interface were compared by scanning electron microscopy.@*RESULTS@#No statistically significant difference in shear bond strength was found among the no-antioxidant, 2.5% grape-seed extract, and 2.5%, 5%, or 10% sodium ascorbate groups (@*CONCLUSIONS@#Immediately after bleaching, the bond strength of dental enamel significantly decreased. Bond strength can be restored by 5% grape-seed extract or 15% sodium ascorbate in 5 min.

Effectiveness of grape seed extract-based intracanal dressings against Enterococcus Faecalis and its influence on dentin microhardness and bond strength of filling material / Eficácia de medicamentos intracanais à base de extrato de semente de uva contra o Enterococcus Faecalis e sua influência na micro-dureza da dentina e na resistência de união do material obturador

The aim of this study was to evaluate the effectiveness of grape seed extract (GSE)-based intracanal dressings against Enterococcus faecalis (E. faecalis) and its influence on dentin microhardness and bond strength of the filling material. The root canals of 126 human teeth were distributed into three test groups: antimicrobial activity (60 teeth), dentin microhardness (30 teeth) and bond strength (36 teeth). In all three groups, specimens were subdivided into six groups, according to intracanal dressing protocols: G1 ­ distilled water (DW); G2 ­ 2% chlorhexidine gel (CHX); G3 ­ calcium hydroxide (Ca[OH]2)+DW; G4 ­ GSE+DW; G5 ­ Ca(OH)2+CHX; G6 ­ GSE+CHX. The counting of colony-forming units (CFUs), the Vickers microhardness tester and the push-out test were performed to evaluate the antimicrobial activity, dentin microhardness and bond strength, respectively. Specific statistical analysis was performed for each evaluation (α=5%). The greatest bacterial reduction was observed in G5 (Ca[OH]2+CHX) and G6 (GSE+CHX) (p<0.05). There was no statistically significant difference among groups in the dentin microhardness evaluation (p<0.05). The highest bond strength in the immediate evaluation was observed in G4 (GSE+DW) and G6 (GSE+CHX), whereas the highest bond strength after 12 months of storage was observed in G2 (CHX), G3 (Ca[OH]2+DW), G4 (GSE+DW), and G6 (GSE+CHX) (p<0.05). After the storage period, bond strength was increased in G2 (CHX) and G3 (Ca[OH]2+DW), and remained unchanged in G4 (GSE+DW) and G6 (GSE+CHX) (p<0.05). GSE-based intracanal dressings have antimicrobial potential against E. faecalis, have no influence in dentin microhardness and preserve the high bond strength of filling materials for root dentin over time.

O objetivo deste estudo foi avaliar a eficácia de medicamentos intracanal à base de extrato de semente de uva (GSE) contra Enterococcus faecalis (E. faecalis) e sua influência na microdureza da dentina e na resistência de união do material de obturação. Os canais radiculares de 126 dentes humanos foram distribuídos em três grupos de teste: atividade antimicrobiana (60 dentes), microdureza da dentina (30 dentes) e resistência adesiva (36 dentes). Nos três grupos, as amostras foram subdivididas em seis grupos, de acordo com os protocolos de curativos intracanal: G1 ­ água destilada (DW); G2 ­ gel de clorexidina a 2% (CHX); G3 ­ hidróxido de cálcio (Ca[OH]2) +DW; G4 ­ GSE+DW; G5 ­ Ca(OH)2+CHX; G6 ­ GSE+CHX. A contagem de unidades formadoras de colônias (UFCs), o testador de microdureza Vickers e o teste push-out foram realizados para avaliar a atividade antimicrobiana, a microdureza da dentina e a resistência adesiva, respectivamente. Análise estatística específica foi realizada para cada avaliação (α=5%). A maior redução bacteriana foi observada no G5 (Ca[OH]2+CHX) e G6 (GSE+CHX) (p<0,05). Não houve diferença estatisticamente significativa entre os grupos na avaliação da microdureza da dentina (p<0,05). A maior resistência adesiva na avaliação imediata foi observada no G4 (GSE+DW) e G6 (GSE+CHX), enquanto a maior resistência adesiva após 12 meses de armazenamento foi observada no G2 (CHX), G3 (Ca[OH]2+DW), G4 (GSE+DW) e G6 (GSE+CHX) (p<0,05). Após o período de armazenamento, a resistência de união aumentou no G2 (CHX) e G3 (Ca[OH]2+DW), permanecendo inalterada no G4 (GSE+DW) e G6 (GSE+CHX) (p<0,05). Os medicamentos intracanal à base de GSE têm potencial antimicrobiano contra E. faecalis, não influenciam na microdureza da dentina e preservam a alta resistência adesiva dos materiais de obturação da dentina radicular ao longo do tempo.

Antimicrobial activity of intracanal dressings composed by natural products associated to chlorhexidine and its influence on dentinal colour change / Avaliação da ação antimicrobiana de medicações intracanais à base de produtos fitoterápicos associados à clorexidina e sua influência na alteração de cor da estrutura dentinária

Purpose: evaluate the antimicrobial activity of intracanal dressings and their influence on dentinal colour changes. Material and methods: eighty single-rooted human extracted teeth were decoronated and divided into eight groups (n=10) according to intracanal dressing protocols inserted into the root canals: G1­distilled water (DW); G2­2% chlorhexidine gel (CHX); G3­calcium hydroxide (Ca[OH]2)+DW; G4­grape seed extract (GSE)+DW; G5­ginger extract (GE)+DW; G6­Ca(OH)2+CHX; G7­GSE+CHX; and G8­GE+CHX. The antimicrobial activity was evaluated by colony-forming units (CFUs) counting and dentinal colour changes was evaluated by digital spectrophotometry. Data were statistically analysed by One-way ANOVA followed by Tukey´s post hoc test (antimicrobial evaluation) and non-parametric Wilcoxon followed by the Mann- Whitney-U test (colour change evaluation) (α=0.05). Results: the highest bacterial reduction was observed in groups 4, 6, 7 and 8, with no significant difference between them (p<0.05). Groups 4 and 7 showed the highest medians of dentinal colour change (p<0.05). Conclusion: the addition of CHX improved the antimicrobial activity of GE-based intracanal dressing, with no effect in GSE-based intracanal dressing; moreover, these protocols induced significant dentinal colour changes. (AU)

Objetivo: avaliar a atividade antimicrobiana de medicações intracanais e sua influência na alteração da cor dentinária. Materiais e métodos: oitenta dentes humanos extraídos unirradiculares foram seccionados e divididos em oito grupos (n = 10), de acordo com os protocolos de medicação intracanal inseridos nos canais radiculares: água destilada G1 (DW); G2-2% de gel de clorexidina (CHX); hidróxido de cálcio G3 ­ (Ca [OH] 2) + DW; extrato de semente de uva G4 (GSE) + DW; extrato de gengibre G5 (GE) + DW; G6- Ca (OH) 2 + CHX; G7 ­ GSE + CHX; e G8-GE + CHX. A atividade antimicrobiana foi avaliada por contagem de unidades formadoras de colônias (UFCs) e as alterações de cor dentinária foram avaliadas por espectrofotometria digital. Os dados foram analisados estatisticamente por ANOVA one-way, seguida pelo teste post hoc de Tukey (avaliação antimicrobiana) e Wilcoxon não paramétrico, seguido pelo teste de Mann- Whitney-U (avaliação da mudança de cor) (α = 0,05). Resultados: a maior redução bacteriana foi observada nos grupos 4, 6, 7 e 8, sem diferença significativa entre eles (p < 0,05). Os grupos 4 e 7 apresentaram as maiores medianas da alteração da cor dentinária (p < 0,05). Conclusão: a adição de CHX melhorou a atividade antimicrobiana da medicação intracanal baseado em GE, sem efeito na medicação intracanal baseado em GSE; além disso, esses protocolos induziram alterações significativas na cor dentinária.(AU)

Effect of oligomeric grape seed proanthocyanidins on DSS-induced ulcerative colitis in mice and its mechanism / 中草药

Objective: To investigate the effect of oligosaccharide grape seed proanthocyanins (GSPE) on dextran sulphate sodium salt (DSS)-induced ulcerative colitis (UC) in mice and its mechanisms. Methods: SPF-class C57 mice were randomly divided into normal group, model group, positive group (sulfasalazine group), and GSPE groups (125, 250, 500 mg/kg). The normal group was given pure water, and the other groups were free to drink 3% DSS aqueous solution for 7 d to induce the model of UC in mice. The changes of body weight, hematochezia and stool type were recorded every day. After seven days of treatment, blood, colons and spleens were collected, and the length of the colon and the weight of the spleen were recorded. HE staining was used to evaluate the pathological changes of colonic mucosa in mice. The expression of interleukin-6 (IL-6), interleukin-1β (IL-1β), and tumor necrosis factor TNF-α in serum and colon tissues and the levels of NO, MDA, and SOD were detected by ELISA. The changes of HO-1, NF-κB, and Nrf2 in colonic epithelial cells were analyzed by immunohistochemistry. Results: Compared with the model group, the body weight of mice in GSPE groups (250, 500 mg/kg) decreased relatively slowly, and the symptoms of diarrhea and hematochezia were improved significantly. The content of IL-1β, IL-6, TNF-α, NO, and MDA in serum and colon tissues was much lower in the administration groups than those in the model group, while the content of SOD was significantly higher (P < 0.01). The pathological tissue analysis showed that the pathological damage of colonic mucosa in the high dose group of GSPE was obviously decreased. Immunohistochemical analysis showed that GSPE groups significantly decreased the expression of NF-κB and increased the expression of Nrf2 and HO-1 (P < 0.01). Conclusion: GSPE could effectively improve the symptoms of UC induced by DSS, and regulate the expression of oxidative stress-related proteins Nrf2, HO-1 and inflammatory pathway protein NF-κB, and then affect the changes of oxidative stress indicators SOD, MDA and inflammatory factors. Therefore, GSPE plays an important role in the treatment and prevention of UC.

Grape seed proanthocyanidins antagonize osteocyte apoptosis due to steroid-induced osteonecrosis of the femoral head / 中国组织工程研究

BACKGROUND: Steroid-induced avascular necrosis of the femoral head has a complex biological process, and its pathogenesis is unknown. To date, there is no effective treatment in clinical practice. Therefore, exploring the etiology of steroid-induced avascular necrosis of the femoral head is still an important content of research in this field. OBJECTIVE: To explore the effect of grape seed proanthocyanidins on osteocyte apoptosis due to steroid-induced osteonecrosis of the femoral head. METHODS: Twenty-seven Japanese white rabbits were randomly divided into three groups. The model group was intravenously injected with E. coli endotoxin, 100 μg/kg, twice at an interval of 24 hours. Two injections of E. coli endotoxin were followed by intramuscular injection of methylprednisolone 20 mg/kg, for 3 times. The interval was 24 hours. The treatment group was intravenously injected with E. coli endotoxin, 100 μg/kg, twice at an interval of 24 hours. After the second injection of E. coli endotoxin, methylprednisolone 20 mg/kg and grape seed proanthocyanidin extract 200 μg/kg were injected intramuscularly three times at an interval of 24 hours. The control group was treated with the same dose of saline intravenously. The animals were killed by air embolization at the 4th week after the last injection. Under the relatively aseptic condition, the bilateral femoral heads were routinely fixed, decalcified, embedded and sliced. Histomorphological observation and hematoxylin-eosin staining were performed to count empty bone lacunae under light microscope. Hoechst staining was used to detect cell apoptosis. The expressions of Caspase-9 and Bcl-2 in the femoral head were detected by immunohistochemical staining.

Effects of grape seed proanthocyanidin B2 pretreatment on oxidative stress and renal tubular epithelial cell apoptosis after renal ischemia reperfusion in mice

Abstract Purpose To investigate the effects of grape seed proanthocyanidin B2 (GSPB2) preconditioning on oxidative stress and apoptosis of renal tubular epithelial cells in mice after renal ischemia-reperfusion (RIR). Methods Forty male ICR mice were randomly divided into 4 groups: Group A: mice were treated with right nephrectomy. Group B: right kidney was resected and the left renal vessel was clamped for 45 minutes. Group C: mice were intraperitoneally injected with GSPB2 before RIR established. Group D: mice were intraperitoneally injected with GSPB2 plus brusatol before RIR established. Creatinine and urea nitrogen of mice were determined. Pathological and morphological changes of kidney were checked. Expressions of Nrf-2, HO-1, cleaved-caspase3 were detected by Western-blot. Results Compared to Group B, morphology and pathological damages of renal tissue were less serious in Group C. Western-blot showed that expressions of Nrf-2 and HO-1 in Group C were obviously higher than those in Group B. The expression of cleaved-caspase3 in Group C was significantly lower than that in Group B. Conclusion GSPB2 preconditioning could attenuate renal oxidative stress injury and renal tubular epithelial cell apoptosis by up-regulating expressions of Nrf-2 and HO-1 and down-regulating the expression of cleaved-caspase-3, but the protective effect could be reversed by brusatol.

Effect of feeding grape seed Proanthocyanidin extract on production performance, metabolic and anti-oxidative status of dairy cattle / Efeito da alimentação do extrato de semente de uva Proanthocyanidin sobre o desempenho da produção e o status metabólico e antioxidante de gado leiteiro

Aims to investigate the effects of grape seed proanthocyanidin extract (GSPE) on production performance, metabolism, and anti-oxidative status of Holstein dairy cattle in early lactation. Forty-eight multiparous Holstein dairy cattle were assigned to four groups (CON, G20, G40 and G80) and supplied with 0, 20, 40, and 80mg GSPE/kg of body weight/day. G20 significantly increased milk yield compared with other groups. Milk protein and non-fat-solids were increased in G20, G40 and G80 groups compared with the control group only at the 7th day during the experiment. No significant difference was observed in milk fat and somatic cell count, nor on parameters of energy metabolism in blood, liver function and kidney function between the four groups. There was no significant difference in glutathione peroxidase, superoxide dismutase, total antioxidant capacity, and hydrogen peroxide between the groups; but the malondialdehyde content of G20 significantly increased at day 14 in comparison with CON, and tended to increase at the 28th day. In conclusion, feeding 20mg GSPE/kg of body weight/day was associated with a significant increase in milk yield without detrimental effects on liver or kidney function and with substantial energy metabolism and antioxidant parameters improvement in early lactation dairy cattle.(AU)

O presente trabalho visa investigar os efeitos do extrato de semente de uva Proanthocyanidin (GSPE) sobre o desempenho da produção, o metabolismo e o status antioxidante de gado leiteiro Holstein em lactação precoce. Quarenta e oito vacas leiteiras multíparas Holstein foram divididas em quatro grupos (CON, G20, G40 e G80) e receberam 0, 20, 40 e 80mg de GSPE/kg de peso corporal/dia, respectivamente. O G20 aumentou significativamente o rendimento do leite em comparação com os outros grupos. A proteína e os sólidos não gordurosos do leite foram aumentados nos grupos G20, G40 e G80 somente no sétimo dia durante a experiência. Não foi observada diferença significativa na gordura do leite e na contagem de células somáticas, bem como nos parâmetros de metabolismo energético no sangue, na função hepática e na função renal entre os grupos em relação ao grupo controle. Não houve diferença significativa na glutationa peroxidase, na dimutase de superóxido, na capacidade antioxidante total e no peróxido de hidrogênio entre os grupos, mas o conteúdo malondialdeído do G20 aumentou significativamente no dia 14 em comparação com o CON, e tendia a aumentar no dia 28. Em conclusão, a alimentação de 20mg de GSPE/kg de peso corporal/dia foi associada a um aumento significativo no rendimento do leite, sem efeitos nocivos sobre a função hepática ou a renal, com o metabolismo de energia substancial e a melhoria dos parâmetros antioxidantes de gado leiteiro no início da lactação.(AU)

Radioprotective effect of grape seed extract against gamma irradiation in mouse bone marrow cells

Introduction: Ionizing radiations produce free radicals which are often responsible for DNA damage or cell death. Grape seed extract (GSE) is a natural compound having an antioxidant that protects DNA, lipids, and proteins from free radical damages. In this study, radioprotective effect of the GSE has been investigated in mouse bone marrow cells using micronucleus test. Materials and Methods: Four groups of mice were investigated in this study: Mice in Group 1 were subjected to injection of distilled water with no irradiation. Mice in Group 2 were exposed to 3 Gy gamma radiation after the injection of distillated water. Mice in Group 3 were injected with 200 mg/kg of the GSE without any irradiation. In another group, mice were exposed to three gray gamma irradiation after the injection of GSE. Animals were killed, and slides were prepared from the bone marrow cells 24 h after irradiation. The slides were stained with May Grunwald–Giemsa method and analyzed microscopically. The frequency of the micronucleated polychromatic erythrocytes (MnPCEs), micronucleated normochromatic erythrocyte (MnNCEs), and polychromatic erythrocyte/polychromatic erythrocyte + normochromatic erythrocyte (PCE/PCE + NCE) ratios was calculated. Results: Injection of GSE significantly decreased the frequency of MnPCEs (P < 0.0001) and MnNCEs (P < 0.05) and increased the ratio of PCE/PCE + NCE (P < 0.0001) compared to the irradiated control group. Discussion and Conclusions: GSE could reduce clastogenic and cytotoxic effects of gamma irradiation in mice bone marrow cells; therefore, it can be concluded that the GSE is a herbal compound with radioprotective effects against gamma irradiation. Free radical scavenging and the antioxidant effects of the GSE probably are responsible mechanisms for the GSE radioprotective effects

Grape seed proanthocyanidin extract inhibits platelet-derived growth factor induced proliferation of airway smooth muscle cells / 中国小儿急救医学

Objective To explore the effect of grape seed proanthocyanidin extract(GSPE) on the proliferation of airway smooth muscle cells( ASMCs) induced by platelet-derived growth factor( PDGF) and the underlying molecular mechanism. Methods ASMCs of primary rat were cultured. MTT and flow cytom-etry were used to detect the cell proliferation activity and cell cycle distribution of ASMCs which were treated with PDGF and GSPE respectively. The expression levels of cyclin D1,extracellular regulated protein kinases ( ERK)1/2,p-ERK1/2 and β-actin protein in each group ASMCs were analyzed using western blotting assay after ERK1/2 inhibitor PD98059 intervention. Results Compared with control group,cell proliferative activ-ity,S phase fraction and the expression of cyclin D1 and p-ERK1/2 protein increased in PDGF induced group (P<0. 05). These effects induced by PDGF could be reversed by GSPE. PD98059 also could block PDGF induced higher expression of p-ERK1/2 and cyclin D1 proteins in rat ASMCs. Conclusion GSPE can inhib-it PDGF induced cell proliferation and via ERK1/2 signaling pathway in rat ASMCs,which provide a new way for treatment of bronchial asthma.

Effect of grape seed extract (GSE) on functional activity and mineralization of OD-21 and MDPC-23 cell lines

Abstract Recent studies on functional tissue regeneration have focused on substances that favor cell proliferation and differentiation, including the bioactive phenolic compounds present in grape seed extract (GSE). The aim of this investigation was to evaluate the stimulatory potential of GSE in the functional activity of undifferentiated pulp cells and odontoblast-like cells. OD-21 and MDPC-23 cell lines were cultivated in odontogenic medium until subconfluence, seeded in 24-well culture plates in a concentration of 2x104/well and divided into: 1) OD-21 without GSE; 2) OD-21+10 µg/mL of GSE; 3) MDPC-23 without GSE; 4) MDPC-23+10 µg/mL of GSE. Cell proliferation, in situ detection of alkaline phosphatase (ALP) and total protein content were assessed after 3, 7 and 10 days, and mineralization was evaluated after 14 days. The data were analyzed by ANOVA statistical tests set at a 5% level of significance. Results revealed that cell proliferation increased after 10 days, and protein content, after 7 days of culture in MDPC-23 cells. In situ ALP staining intensity was higher in undifferentiated pulp cells and odontoblast-like cells after 7 and 10 days, respectively. A discrete increase in MDPC-23 mineralization after GSE treatment was observed despite OD-21 cells presenting a decrease in mineralized nodule deposits. Data suggest that GSE favors functional activity of differentiated cells more broadly than undifferentiated cells (OD-21). More studies with different concentrations of GSE must be conducted to confirm its benefits to cells regarding dentin regeneration.

Antibacterial efficacy of synthetic and natural-derived novel endodontic irrigant solutions

Abstract The aim of this study was to compare the efficacy of grape seed extract (GSE), calcium hypochlorite [Ca(ClO)2], and sodium hypochlorite (NaOCl) irrigant solutions with rotary or reciprocating instrumentation for disinfection of root canals inoculated with Enterococcus faecalis. The mesiobuccal root canals of mandibular molars were prepared and inoculated with Enterococcus faecalis for 21 days. The roots were then randomly divided into the following eight experimental groups (n=11) according to the instrumentation technique and disinfection protocol: ProTaper Next or Reciproc R25 with sodium chloride (control group), 6% NaOCl, 6% Ca(ClO)2, or 50% GSE used for irrigation during instrumentation. The antimicrobial activity was determined on the basis of a reduction in colony-forming units (CFUs) counted on bacterial samples collected before and after root canal instrumentation and expressed as a percentage of reduction. Data were evaluated by two-way ANOVA followed by Tukey HSD post-hoc tests (p<0.05). No significant differences were observed in bacterial reduction between the ProTaper Next and Reciproc R25 systems (p>0.05), regardless of the irrigant solution used. Furthermore, all active solutions (6% NaOCl, 50% GSE, and 6% Ca(ClO)2) showed similar potential to reduce bacterial counts (p>0.05) and were significantly more effective than sodium chloride (control) (p<0.05). The results suggest that the GSE and Ca(ClO)2 have potential clinical application as irrigant solutions in endodontic therapy since they present bactericidal efficacy against Enterococcus faecalis.

Resumo O objetivo deste estudo foi comparar a eficácia do extrato de semente de uva (ESU), hipoclorito de cálcio [Ca(ClO)2] e hipoclorito de sódio (NaOCl) como soluções irrigadores quando utilizadas com instrumentos reciprocantes e rotatórios para desinfecção de canais radiculares infectados com Enterococcus faecalis. Raízes mesio-vestibulares de molares inferiores foram preparados e inoculados com E. faecalis por 21 dias. As raízes foram aleatoriamente divididas em 8 grupos (n=11) de acordo com a técnica de instrumentação e protocolo de irrigação: ProTaper Next ou Reciproc R25 associados com soro fisiológico (grupo controle), Ca(ClO)2 6%, NaOCl 6% ou ESU 50%. A atividade antimicrobiana foi determinada pela redução do número de Unidades Formadoras de Colonias (UFCs) coletadas antes e após a instrumentação e expressas em porcentagens de redução. Os dados foram analisados estatisticamente pelos testes ANOVA seguido pelo teste complementar de Tukey HSD (p<0,05). Não foi encontrado diferença estatisticamente significante na redução bacteriana entre os sistemas ProTaper Next e Reciproc R25 (p>0.05), independente da solução irrigadora usada. Além disso, todas as soluções ativas (NaOCl, ESU e Ca(ClO)2) mostraram similar potencial em reduzir a quantidade de bactérias (p>0.05) e foram significativamente mais efetivas que o soro fisiológico (p<0.05). Pode-se concluir que o ESU e o Ca(ClO)2 apresentam potencial para aplicação clínica como irrigantes endodônticos uma vez que apresentaram efetividade antimicrobiana contra o E. faecalis.

Comparative Evaluation Of The Effect Of Various Remineralizing Agents On Root Dentin: An In Vitro Study

50 root dentin sections were obtained from 25 extracted premolars and divided randomly into 5 groups of 10 samples each. Except group 1, all the other group specimens were demineralised by immersing in a demineralising solution for 96 hours at 370C. The specimens were then subjected to surface treatment with grape seed extract, silverdiamine fluoride and calcium sucrose orthophosphate complex according to the groups. The samples were stored in artificial saliva during the intervals and subjected to pH cycling and evaluated using SEM-EDAX and the results statistically compared. The tested remineralizing agents showed statistically significant increase in remineralization when compared to the demineralized group. Calcium sucrose orthophosphate showed the highest Ca/Pvalues followed by Silver diamine flouride and grape seed extract

Grape seed proanthocyanidin extract ameliorates murine autoimmune arthritis through regulation of TLR4/MyD88/NF-κB signaling pathway

BACKGROUND/AIMS: Grape seed proanthocyanidin extract (GSPE) has been reported to have a beneficial effect on regulating inf lammation. However, the anti-inflammatory mechanism of GSPE remains unclear. The aim of this study was to verify the influence of GSPE on the Toll-like receptor 4 (TLR4)-mediated signaling pathway in the regulation of murine autoimmune arthritis. METHODS: Collagen-induced arthritis (CIA) was induced in dilute brown non-agouti (DBA)/1J mice. The mice were treated with GSPE (0 or 100 mg/kg) intraperitoneally. The severity of arthritis was assessed clinically, biochemically, and histologically. Immunostaining for TLR4 was performed. The expressions of TLR4 and downstream signaling molecules were analyzed by Western blot. The effect of GSPE on lipopolysaccharide (LPS)-induced TLR4 activation was also evaluated using RAW264.7 cells and fibroblast-like synoviocytes (FLSs) from patients with rheumatoid arthritis and from those with osteoarthritis. RESULTS: GSPE attenuated the clinical severity of arthritis and decreased histological damage. GSPE treatment reduced the number of TLR4-stained cells in the synovium of mice with CIA. GSPE also downregulated the expression of TLR4, myeloid differentiation factor 88 (MyD88) and phosphorylated IκBα synovial protein in CIA mice. Concurrently, GSPE inhibited the nuclear translocation of nuclear factor-κB (NF-κB) subunits (p65 and p50). LPS-induced TLR4 activation was suppressed by GSPE in human FLS as well as in murine macrophages in vitro. CONCLUSIONS: Our results demonstrated that GSPE ameliorated CIA by regulating the TLR4-MyD88-NF-κB signaling pathway.

Avaliação da expressão de citocinas pró e anti-inflamatórias após estímulo celular com diferentes agentes irrigantes / Evaluation of the expression of pro and anti-inflammatory cytokines after cellular stimulation with different irrigating agents

O objetivo do trabalho foi investigar o efeito de diferentes soluções químicas auxiliares, em concentrações subcitotóxicas, na expressão de citocinas pró e anti-inflamatórias, quando em contato com células da linhagem de linfoma humano, diferenciadas em macrófagos, human macrophage-like (U937), através da adição de 125ng/mL de PMA. As concentrações, citotóxica e subcitotóxica, de cada solução, foram determinadas de acordo com padrão ISO, utilizando-se fibroblastos de camundongos (L929). As células L929 foram cultivadas em meio MEM completo e mantidas em placas de 96 poços. As células foram então colocadas em contato com as diluições das soluções químicas auxiliares (a partir das concentrações recomendadas para uso, NaOCL 5,25%; CHX 2%; Quitosana 0,2%; HEBP 18%; GSE de Vitis vinífera, 6,5%), por 24 horas a 37ºC em estufa de CO2. Em seguida, o meio de cultivo contendo 1mg/mL de MTT foi adicionado aos poços em triplicata, para avaliação da viabilidade celular e determinação das concentrações citotóxicas (30% de morte das células L929) e subcitotóxicas (15% de morte) por regressão linear por meio do programa GraphPad Prism versão 6.0. As concentrações subcitotóxicas foram colocadas em contato com as células human macrophage-like (U937) por 60 min. Os sobrenadantes da pré-incubação (controles sem substâncias químicas auxiliares), e incubação com as soluções químicas auxiliares em meio DMEM completo foram congelados em freezer ultrafrio (­ 80ºC) e avaliados para as concentrações de dezessete citocinas através do kit Bio-Plex Pro Human Th17 Cytokine Panel® pelo método Luminex®. Foi possível obter valores de expressão de 7 citocinas pró-inflamatórias: IL-1ß, TNFα, IL-6, IL-17A, IL-17F, IL-22 e IL-23. As substâncias com maior atividade citotóxica (padrão ISO para morte celular) foram em ordem de grandeza, a quitosana, a CHX, o NaOCl, o HEBP e o GSE de Vitis vinifera. As concentrações subcitotóxicas das soluções químicas auxiliares, CHX e NaOCl, induziram ao aumento da maioria das citocinas pró-inflamatórias, exceto para a IL-6. Quanto a IL-1ß, a quitosana e a CHX, estas foram as que mais induziram a expressão pelas células U937 (P<0,05),enquanto o NaOCl induziu maior expressão de TNFα (P>0,05). Dentre as citocinas relacionadas ao fenótipo Th17, a CHX foi a que mais induziu a expressão de IL-17A (P<0,05), IL-17F e IL-23, seguido do NaOCl(P<0,05). O NaOCl induziu maior expressão de IL22 seguido da CHX (p<0,05). Quanto a IL-6, todas as substâncias modularam a sua expressão pelas células U937(p>0.05). O HEBP atuou como um excelente modulador da expressão de citocinas pró-inflamatórias, induzindo a redução da expressão de todas as citocinas testadas (P<0,05). O GSE de Vitis vinifera, apesar de ser o menos citotóxico não apresentou diferença estatística para o controle em nenhuma das situações avaliadas, exceto para a redução da IL-6 (assim como o HEBP e a quitosana) (p<0,05). A quitosana comportou-se de forma similar ao GSE de Vitis vinífera (p>0,05) exceto para a IL-1ß. Conforme observado, as substâncias químicas auxiliares são capazes de induzir a expressão de citocinas pró-inflamatórias diversas. O HEBP foi o agente químico que melhor modulou a expressão de citocinas pró-inflamatórias.

The aim of this investigation was to analyze the effect of different chemical endodontic solutions, in subcytotoxic concentrations, on the expression of pro and anti-inflammatory cytokines when in contact with human lymphoma lineage differentiated into human macrophages, human macrophage-like (U937), by the addition of 125ng/ml PMA. The cytotoxic and subcytotoxic concentrations of each solution were determined according to ISO standard using mouse fibroblasts (L929). L929 cells were cultured in complete MEM medium and maintained in 96-well plates. Dilutions of the auxiliary chemical solutions, from the recommended concentrations for use: NaOCL 5,25%; CHX 2%; Chitosan 0.2%; Etidronic acid 18% and Grape seed extract, Vitis vinifera, 6.5%, were carried out in complete MEM medium, and placed in contact with the cells for 24 hours at 37ºC in CO2 cell incubator, in triplicate. Thereafter, the culture medium containing 1 mg/mL MTT was added to the wells. Cytotoxic concentrations (30% death of L929 cells) and subcytotoxic (15% death) were determined by linear regression using GraphPad Prism version 6.0. Subcytotoxic concentrations were obtained in complete DMEM medium and maintained in contact for 60 min. with U937 cells. Controls without chemical substances were also performed. Supernatants from the pre-incubation, incubation with the chemical solutions and complete DMEM were removed, frozen in ultra-cold freezer (-80ºC) and evaluated for the concentrations of 17 cytokines through the Bio-Plex Pro Human Th17 Cytokine Panel® kit by Luminex®. The substances with higher cytotoxic activity were in order of magnitude: chitosan, CHX, NaOCl, etidronic acid and Vitis vinifera extract. The values of 7 pro-inflammatory cytokines: IL-1ß, TNFα, IL-6, IL-17A, IL-17F, IL-22 and IL-23 were obtained. Subcytotoxic concentrations of the irrigant agents (chlorhexidine and NaOCl) generally induced the increase of most pro-inflammatory cytokines, except for IL-6. Chitosan and CHX were the agents that most induced the expression of IL-1ß by U937 cells, whereas NaOCl induced higher TNFα expression (P> 0,05). Among the cytokines related to the Th17 phenotype, CHX was the agent that induced the highest expression of IL-17A (P <0,05), IL-17F and IL-23, followed by NaOCl (P<0,05). NaOCl induced the highest expression of IL22 followed by CHX. All of the substances modulated the expression of IL-6 by U937 cells (P >0,05). HEBP was an excellent modulator of the expression of pro-inflammatory cytokines, inducing the reduction of the expression of all cytokines tested. Vitis vinifera extract, despite being the least cytotoxic agent, did not present statistical differences compared to the control for any of the evaluated cytokines, except for the reduction of IL-6 (as well as etidronic acid and chitosan) (P<0,05). Chitosan, except for IL-1 ß, was very similar to the Vitis vinifera (P>0,05) extract. As observed, the auxiliary substances (especially the irrigants) are capable of inducing the expression of various pro-inflammatory cytokines. Etidronic acid was the chemical that modulated the expression of pro-inflammatory cytokines.