Efeito tempo-dose-resposta do montelucaste no processo de reparo alveolar em camundongos 129Sv/Ev / Time-dose-response effect of Montelukast in the process of alveolar repair in 129Sv / Ev mice

Metabólitos do ácido araquidônico são conhecidos por exercerem importante papel nos processos inflamatórios e no metabolismo do tecido ósseo. No entanto, as ações pontuais, especialmente dos leucotrienos derivados da 5-lipoxigenase (5-LO) sobre o processo de reparo ósseo intramembranoso são pouco exploradas. O presente estudo tem como objetivo analisar os efeitos tempo-dose-resposta da droga montelucaste (MTK), potente antagonista dos receptores de cisteinil leucotrienos tipo 1 (CisLT1Rs), no curso do reparo alveolar pós-exodontia em camundongos 129Sv/Ev, bem como nos níveis plasmáticos de marcadores ósseos bioquímicos. Para tanto, foram utilizados 70 camundongos machos jovens divididos em quatro grupos, de acordo com o tratamento: C - Grupo Controle (não tratados); CV - Grupo Controle Veículo, 20 µL de solução fisiológica (SF) 0,9%; MTK2 ­2 mg/kg de MTK e MTK4 ­ 4 mg/kg de MTK. Os animais dos grupos CV, MTK2 e MTK4 foram tratados diariamente por via oral, iniciando 24 horas antes do procedimento cirúrgico, continuando até o final dos períodos experimentais de 7, 14 e 21 dias pós-operatórios. Ao final dos períodos determinados, os animais foram submetidos à eutanásia para coleta de sangue para análise bioquímica dos níveis de cálcio, fosfato, fosfatase ácida resistente ao tartarato (TRAP) total e fosfatase alcalina (FAL), coleta da maxila direita contendo os alvéolos dentários para serem analisados por meio de microtomografia computadorizada (microCT), e análise histopatológica. Os resultados obtidos foram submetidos à testes estatísticos considerando-se nível de confiança de 5%. Observou-se aumento do BV/TV para os animais tratados com MTK em relação aos grupos C e CV, tanto aos 14 dias quanto aos 21 dias, sendo maior no grupo MTK4 aos 14 dias em relação ao grupo MTK2. Do mesmo modo, os animais tratados com MTK em ambas doses apresentaram aumento significativo de Tb.Th em comparação aos grupos C e CV aos 21 dias. Chamou a atenção valores de BV/TV e Tb.Th significativamente reduzidos no grupo CV em comparação ao C, indicando um efeito negativo da manipulação do animal. Na análise histopatológica observou-se reparo ósseo precoce nos animais MTK2 e MTK4 em todos os períodos avaliados, em comparação aos do grupo C, bem como atraso no processo de reparo no grupo CV aos 21 dias. Quanto aos marcadores plasmáticos, observou-se aumento do cálcio no grupo MTK4 em relação ao grupo C aos 7 dias, e aos 21 dias também em relação ao grupo MTK2. Já o fosfato mostrouse significantemente elevado nos períodos de 7 e 21 dias no grupo MTK2 em relação aos demais grupos. FAL e TRAP total não apresentaram níveis plasmáticos significativamente diferentes comparando-se os grupos e períodos. Considerando os resultados obtidos, concluiu-se que o MTK exerceu efeito tempo-dose-dependente, acelerando o processo de reparo ósseo intramembranoso alveolar e interferindo nos níveis plasmáticos de cálcio e fosfato no presente modelo animal(AU)

Arachidonic acid metabolites are known to play an important role in inflammatory processes and in bone metabolism. However, the role of these products on alveolar bone repair post tooth extraction remains to be explored, especially leukotrienes, derived from 5-lipoxygenase (5-LO). The present study aims to analyze the time-doseresponse effects of the drug montelukast (MTK), a potent type 1 leukotriene cystenyl antagonist (CisLT1Rs), in the alveolar repair process after extraction in male 129Sv/Ev mice. For this purpose, 70 young male mice were used, divided into four groups: C - Control Group (no treatment); VC - Vehicle Control Group, treated with 20 µL of 0.9% SF; MTK2 - treated with 2mg / Kg of MTK and MTK4 - treated with 4mg / Kg of MTK. The animals of the CV, MTK2 and MTK4 groups were treated daily orally (V.O.), starting 24 hours before the surgical procedure, continuing until the end of the experimental periods of 7, 14 and 21 days postoperatively. At the end of the experimental periods, the animals were euthanized for blood collection for serum markers as calcium, phosphate, tartrate-resistant acid phosphate (TRAP) and alkaline phosphatasis (FAL), and to removal of the right maxilla containing the dental socket to be analyzed under computed microtomography (microCT) and histopathology. The results obtained were subjected to statistical tests considering a confidence level of 5%. Results revealed an increase in BV/TV for MTK vs. C and CV groups, in both 14 and 21 days time points. Of note, this increase was higher in MTK4 than in the MTK2 at 14 days. Considering Tb.Th, both MTK2 e MTK4 groups presented positive effects in the BV/TV and Tb.Th increase when compared to controls groups (C and CV) at 21 days. A decrease in BV/TV and Tb.Th was observed in CV compared to C, as a negative effect of animal manipulation. As observed in H&E sections, both MTK2 and MTK4 experimental groups presented an early bone repair in comparison with C group from 7 to 21 days. CV group presented a slight delayed bone healing compared to C. Levels of calcium was increased in MTK4 in comparison to C and MTK2 at 7 and 21 days. Phosphate was significantly elevated at 7 and 21 days in MTK2 in comparison to the other groups. Despite of beneficial effects on observed on morphological levels on sites of healing (microCT and HE), no significant changes were found for bone markers of remodeling in blood plasma (FAL and TRAP). Taken together, these results indicate that MTK induced early bone healing post tooth extraction in 129Sv/Ev mice. Thus, the inhibition of CysLT is suggested to exert a positive influence on intramembranous bone repair post tooth extraction(AU)

Beneficial effects of methanolic extract of Indigofera linnaei Ali. on the inflammatory and nociceptive responses in rodent models

ABSTRACT Indigofera linnaei Ali. (Tamil Name: Cheppu Nerinjil) belongs to the family Fabaceae, used for the treatment of various ailments in the traditional system of medicine. In the present study, the beneficial effects of methanol extract of whole plant of I. linnaei (MEIL) were evaluated on inflammation and nociception responses in rodent models. In vitro nitric oxide (NO), lipoxygenase (LOX) and cyclooxygense (COX) inhibitory activities were also performed to understand the mode of action. MEIL at the dose of 200 & 400 mg/kg, p.o. significantly inhibited carrageenan induced rat paw volume and reduced the weight of granuloma in cotton pellet granuloma model. The results obtained were comparable with the standard drug aceclofenac. The anti-nociceptive effect of MEIL in mice was evaluated in hot plate and acetic acid induced writhing model. The plant extract significantly reduced the number of writhes and the analgesic effect was higher than that of the standard drug aspirin. However, the extract fails to increase the latency period in hot plate method suggesting that the extract produce nociception by peripheral activity. The extract produced inhibitory effect on NO, LOX and COX in concentration dependent manner. The extract exhibited pronounced and selective COX-2 inhibition. Altogether, these results suggested that the methanol extract of Indigofera linnaei could be considered as a potential anti-inflammatory and analgesic agent.

RESUMO Indigofera linnaei Ali pertence à família Leguminosae e é utilizada para o tratamento de várias doenças na medicina tradicional. No presente estudo, os efeitos benéficos do extrato metanólico da planta inteira de I. linnaei (MEIL) foram avaliados em respostas inflamatórias e nocicepção em modelos de roedores. Testes in vitro de atividade inibitória do óxido nítrico (NO), lipoxigenase (LOX) e ciclooxigenase (COX) também foram realizados para compreender o modo de ação. MEIL nas doses de 200 e 400 mg/kg, p.o. inibiu significativamente o volume da pata de rato induzido por carragenana e reduziu o peso do granuloma no modelo de pélete de algodão. Os resultados obtidos foram comparáveis ao do fármaco padrão, aceclofenaco. O efeito anti-nociceptivo de MEIL foi avaliado em camundongos no modelo de placa quente e de contorção induzida por ácido acético. O extrato da planta reduziu significativamente o número de contorções e o efeito analgésico foi maior do que o do fármaco padrão, ácido acetilsalicílico. Porém, o extrato não conseguiu aumentar o período de latência no método da placa quente, sugerindo que este produz nocicepção por atividade periférica. O extrato produziu efeito inibitório sobre o NO, LOX e COX dependente da concentração. O extrato exibiu inibição acentuada e seletiva da COX-2. No seu conjunto, estes resultados sugerem que o extrato metanólico de Indigofera linnaei poderia ser considerado como agente anti-inflamatório e analgésico potencial.

Antioxidant lipoxygenase inhibitors from the leaf extracts of Simmondsia chinensis

OBJECTIVE@#To isolate and identify chemical constituents with antioxidant and lipoxygenase inhibitory effects of the ethanolic extract of Simmondsia chinensis (Jojoba) leaves.@*METHODS@#The alcoholic extract was subjected to successive solvent fractionation. The antioxidant active fractions (chloroform, ethyl acetate and aqueous fractions) were subjected to a combination of different chromatographic techniques guided by the antioxidant assay with DPPH. The structures of the isolated compounds were elucidated on the basis of spectroscopic evidences and correlated with known compounds. The antioxidant activity was assessed quantitively using DPPH and β-carotene methods. The inhibitory potential against enzyme lipoxygenase was assessed on soybean lipoxygenase enzyme.@*RESULTS@#Ten flavonoids and four lignans were isolated. Flavonoid aglycones showed stronger antioxidant and lipoxygenase inhibitory effects than their glycosides. Lignoid glycosides showed moderate to weak antioxidant and lipoxygenase inhibitory effects.@*CONCLUSIONS@#A total of 14 compounds were isolated and identified from Simmondsia chinensis; 12 of them were isolated for the first time. This is the first report that highlights deeply on the phenolic content of jojoba and their potential biological activities and shows the importance of this plant as a good source of phenolics in particular the flavonoid content.

Antioxidant lipoxygenase inhibitors from the leaf extracts of Simmondsia chinensis

Objective: To isolate and identify chemical constituents with antioxidant and lipoxygenase inhibitory effects of the ethanolic extract of Simmondsia chinensis (Jojoba) leaves. Methods: The alcoholic extract was subjected to successive solvent fractionation. The antioxidant active fractions (chloroform, ethyl acetate and aqueous fractions) were subjected to a combination of different chromatographic techniques guided by the antioxidant assay with DPPH. The structures of the isolated compounds were elucidated on the basis of spectroscopic evidences and correlated with known compounds. The antioxidant activity was assessed quantitively using DPPH and β-carotene methods. The inhibitory potential against enzyme lipoxygenase was assessed on soybean lipoxygenase enzyme. Results: Ten flavonoids and four lignans were isolated. Flavonoid aglycones showed stronger antioxidant and lipoxygenase inhibitory effects than their glycosides. Lignoid glycosides showed moderate to weak antioxidant and lipoxygenase inhibitory effects. Conclusions: A total of 14 compounds were isolated and identified from Simmondsia chinensis; 12 of them were isolated for the first time. This is the first report that highlights deeply on the phenolic content of jojoba and their potential biological activities and shows the importance of this plant as a good source of phenolics in particular the flavonoid content.

Benzoxazole Derivative B-98 Ameliorates Dextran Sulfate Sodium-induced Acute Murine Colitis and the Change of T Cell Profiles in Acute Murine Colitis Model / 대한소화기학회지

BACKGROUND/AIMS: The unique role of enzyme 5-lipoxygenase (5-LO) in the production of leukotrienes makes it a therapeutic target for inflammatory bowel disease (IBD). The aim of this study was to evaluate the effects of B-98, a newly synthesized benzoxazole derivatives and a novel 5-LO inhibitor, in a mouse model of IBD induced by dextran sulfate sodium (DSS). METHODS: C57BL/6 mice were randomly assigned to four groups: normal control, DSS colitis (DSS+saline), low dose B-98 (DSS+B-98 20 mg/kg) and high dose B-98 (DSS+B-98 100 mg/kg). B-98 was administered with 3% DSS intraperitoneally. The severity of the colitis was assessed via the disease activity index (DAI), colon length, and histopathologic grading. The production of inflammatory cytokines interleukin (IL)-6 was determined by RT-PCR. Th cells were examined for the proportion of Th1 cell, Th2 cell, Th9 cell, Th17 cell and Treg cell using intracellular cytometry. RESULTS: The B-98 group showed lower DAI, less shortening of the colon length and lower histopathologic grading compared with the DSS colitis group (p<0.01). The expression of IL-6 in colonic tissue was significantly lower in the B-98 groups than the DSS colitis group (p<0.05). The cellular profiles revealed that the Th1, Th9 and Th17 cells were increased in the DSS colitis group compared to the B-98 group (p<0.05). CONCLUSIONS: Our results suggest that acute intestinal inflammation is reduced in the group treated with B-98 by Th1, Th9 and Th17 involved cellular immunity.

Effects of lipoxygenase inhibitor NDGA on expression of 5-LOX and its apoptosis related genes in HepG2 cell line / 国际外科学杂志

Objective To investigate the effects of lipoxygenase inhibitor NDGA on expression of 5-LOX and its apoptosis related genes in HepG2 cell line.Methods The expression of 5-LOX and apoptosis related genes hTERT,bcl-2 and bax mRNA was determined by reverse transcriptasepolymerase chain reaction (RT-PCR).Results After 25,50,100,200 μmol/L NDGA treatment for24,48 h,the expression of 5-LOX of HepG2 cell decreased,but the expression of bax was up-regulated and the expressions of bcl-2 and hTERT mRNA were down-regulated,(P＜0.05 compared with the control group).The decrease in the expression of 5-LOX,hTERT and bcl-2 in HepG2 cell was negtively correlated with the dose duration of action of NDGA.Conclusion In vitro,5-LOX is expressed highly in HepG2 cell.Overexpression of 5-LOX may be related to the progression of hepatocellular carcinoma,NDGA can significantly decrease the expression of 5-LOX,up-regulate of bax and downregulation of bcl-2 and telomerse.Lipoxygenase might be a novel therapeutic target for the hepatocellular carinoma.

The effect of lipoxygenase inhibitor (NDGA) on the expression of VEGF and CD44V6 in breast cancer MCF-7 cells / 中国现代普通外科进展

Objective:To explore the effects of VEGF and CD44V6 on breast cancer MCF-7 cells by NDGA.Methods:MCF-7 cells in the logarithmic growth state were selected and measured.There were four cell groups(concentration:0,1,10,100?mol/L)in the experiment.After 48h treatment of NDGA,the expression of VEGFmRNA was measured by RT-PCR and the expression of CD44V6 was examined by flow cytometry.Results:After treatment of NDGA,the expressions of VEGFmRNA and CD44V6 in NDGA groups were significantly lower than that of the control group(F=2996.13,P

Effects of lipoxygenase inhibitor NDGA on telomerase and bcl-2 in breast cancer MCF-7 cells / 中国现代普通外科进展

Objective:To explore the effects of lipoxygenase inhibitor NDGA on telomerase and bcl-2 of breast cancer MCF-7 cells.Methods:MCF-7 cells in the logarithmic growth state were selected and measured.Four cell groups (concentration:0,1,10,100?mol/L)were divided in the experiment.After 48h treatment of NDGA,the mRNA expression of telomerase was examined by RT-PCR and the expression of bcl-2 was measured by flow cytometry.Results:After 48h treatment of different concentration of NDGA,the expressions of hTERTmRNA and bcl-2 in NDGA groups were both significantly lower than the control group (P0.001).Conclusion:NDGA can prevent proliferation and induce apoptosis in MCF-7 cells by means of the down-regulated expressions of hTERTmRNA and bcl-2.

Advances in studies on lipoxygenase and its inhibitors / 中国药理学通报

Lipoxygenase(LO) pathway had been implicated in the pathogenesis of such cardiovascular diseases as hypertension, atherosclerosis, restenosis, and palys and important role in the development of these disease. LO inhibitiors could suppress vascular contractile responses significantly, reduce blood pressure, inhibit migration of vascular smooth muscle cells(VSMC), attenuate neointimal thickening in the injuried arteries and generation reactive oxygen species(ROS), block monocyte binding to VSMC, etc. The effects of LO inhibitors were associated with marked inhibition of MAPK pathway. Therefore, inhibition of LO pathway may provide a new strategy for preventing and treating above diseases, suggesting that LO mat be a novel taget for such purposes.

Research on apoptosis mechanism of HT-29 colon cancer cell induced by NDGA / 中国现代普通外科进展

Objective:To find out the apoptosis mechanism of HT-29 colon cancer cell induced by NDGA,the lipoxygenase inhibitor in vitro.Methods:We applied respectively:RT-PCR to detect colon cancer cell’s expression of 5-LOX messenger ribonucleic acid(5-LOXmRNA) and that of messenger ribonucleic acid about human telomerase reverse transcriptase(hTERTmRNA)respectively,confocal laser scanning electron microscope to examine intracellular free calcium.Results:Colon cancer cell lines HT-29 showed positive expression of 5-LOXmRNA.This expression became weaker following the rise of cell’s apoptosis and so were hTERTmRNA.Intracellular Free calcium increased following the rise of cell’s apoptosis.Conclusion:The apoptosis of tumor cell is caused by a combination of factors,with 5-LOX,telomerase and free calcium all active in the course.

Study on the action by PAF on IL-1 modulation in alveolar macrophages: Involvement of endogenous arachidonate metabolites and intracellular Ca++ mobilization

Platelet-activating factor (PAF) enhanced interleukin-1 (IL-1) activity by the interaction with a specific receptor in rat alveolar macrophages. In this study, we investigated the role of endogenous arachidonate metabolites and intracellular calcium mobilization in the PAF-induced IL-1 activity. Alveolar macrophages were preincubated with 5-lipoxygenase and cyclooxygenase inhibitors 30 min before the addition of PAF and lipopolysaccharide (LPS). After 24 h culture, IL-1 activity was measured in the supernate of sample using the thymocyte proliferation assay. Inhibition of 5-lipoxygenase by nordihydroguaiaretic acid and AA-861 completely blocked the PAF-induced enhancement of IL-1 activity with IC50 of 2 micrometer and 5 micrometer, respectively. In contrast, the inhibition of cyclooxygenase pathway by indomethacin and ibuprofen resulted in the potentiation in PAF-induced IL-1 activity with maximal effect at 1 micrometer and 5 micrometer, respectively. In addition, leukotriene B4 and prostaglandin E2 production were observed in PAF-stimulated alveolar macrophage culture. As could be expected, 5-lipoxygenase and cyclooxygenase inhibitors abolished PAFstimulated leukotriene B4 and prostaglandin E2 production, respectively. The effects of PAF on intracellular calcium mobilization in alveolar macrophages were evaluated using the calcium-sensitive dye fura-2 at the single cell level. PAF at any dose between 10-16 and 10-8M did not increase intracellular calcium. Furthermore, there was no effective change of intracellular calcium level when PAF was added to alveolar macrophages in the presence of LPS or LPS + LTB4, and 4, 24 and 48h after treatment of these stimulants. Together, the results indicate that IL-1 activity induced by PAF is differently regulated through subsequent induction of endogenous 5-lpoxygenase and cyclooxygenase pathways, but not dependent on calcium signalling pathway.

Effects of nimesulide on proliferation and apoptosis of human cholangiocarcinoma cell line QBC939 / 中国普通外科杂志

Objective To investigate the effect of a selective inhibitor of COX-2 nimesulide on growth and apoptosis of human cholangiocarcinoma cell line QBC939 in vitro. Methods MTT assay was used to determine the influence of nimesulide on the proliferation of QBC939 cells, apoptosis of QBC939 was measured by transmission electron microscopy and flow cytometry.Expression of apoptosis related genes mRNA and bcl-2 ,bax, survivin were detected by RT-PCR and immunocytochemistry. Results Nimesulide effects a dose-dependent and time-dependent growth inhibition on QBC939 cells. High concentration of nimesulide (200 ?mol/L) not only inhibits the growth of QBC939 cells but also induces apoptosis cell nuclear condensation and apoptotic bodies were seen by transmission electron microscopy. Immunocytochemistry and RT-PCR shows upregulation of bax and down regulation of bcl-2 and survivin. Conclusion Nimesulide significantly inhibits the proliferation of QBC939 in vitro by induction of apoptosis in a dose- and time- dependent manner.