砂仁水提物对5-FU致大鼠肠黏膜屏障损伤的保护作用
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摘要
目的研究砂仁水提物对5-氟尿嘧啶(5-FU)诱导的大鼠肠黏膜损伤的保护作用。方法清洁级SD大鼠随机分成正常对照组、模型组、阳性药组、砂仁水提物48,96,192mg·kg~(-1)组,每组8只。正常对照组腹腔注射生理盐水5d,其余各组腹腔注射35 mg·kg~(-1) 5-FU 5 d,且从实验开始正常对照组空腹灌胃生理盐水、其余各组空腹灌胃相应治疗药物,给药12 d。实验期间记录大鼠腹泻情况、体质量、食量;实验结束后取小肠做病理切片;ELISA法检测大鼠IL-6、ROS、NF-κB、MPO、TNF-α、LPS的含量,流式细胞术检测肠道紧密连接蛋白ZO-1、occludin及caspase-3的含量。结果砂仁水提物可显著改善5-FU导致的大鼠体质量、食量下降;显著改善5-FU诱导的大鼠肠屏障损伤及腹泻;显著降低大鼠血液中炎症因子IL-6、ROS的含量;显著抑制5-FU导致的大鼠小肠中NF-κB、TNF-α、MPO的升高与阳性药效果相当。同时砂仁水提物可增加occludin、抑制caspase-3的表达,在一定程度上抑制肠源性LPS易位,保护肠屏障功能。结论砂仁水提物对5-FU引起的肠黏膜屏障损伤具有一定保护作用,其中96 mg·kg~(-1)的效果较好,推测其作用机制可能与砂仁水提物抑制肠黏膜炎前体物质ROS的产生,从而抑制核因子NF-κB通路,降低炎症反应;抑制MPO的激活及caspase-3表达,维持细胞间紧密完整性进而抑制肠源性LPS易位有关。
OBJECTIVE To study the effect of Amomi Fructus water extraction on intestinal mucosal barrier function in rats induced by 5-FU. METHODS SPF SD rats were divided into normal control group, model group, positive drug group and water extract of Amomi Fructus(48, 96, 192 mg·kg~(-1)) groups, 8 rats in each group. The normal control group was intraperitoneally injected with saline for 5 d, and the other groups were intraperitoneally injected with 35 mg·kg~(-1) 5-FU for 5 d. From the beginning of the experiment, the normal control group were given saline, and the other groups were given the corresponding therapeutic drugs for fasting and gavage for 12 d. Diarrhea occurrence, weight and food intakes were recorded daily throughout the study in rats. Blood samples and the small intestinal were collected after killing of rats for evaluation of histopathology, the content of cytokines(IL-6, ROS, NF-kB, MPO and TNF-α) and LPS were detected by ELISA. The intestinal tight junction protein ZO-1, occludin and caspase-3 were measured by flow cytometry. RESULTS Water extract of Amomi Fructus significantly improved rats body weight, food intakes diarrhea and significantly reversed the hisopathological changes in intestinal mucositis induced by 5-FU and reduced the inflammatory parameters(IL-6, ROS) in blood sample. It could also significantly effectively reduce the content of NF-κB, TNF-α, MPO in small intestinal tissue and the effect was similar to the positive drug. Meanwhile, it showed that water extract of Amomi Fructus could increase the expression of occludin and decrease the expression of caspase-3, inhibit the translocation of LPS, protect the integrity of intestinal barrier function. CONCLUSION It is suggested that the water extract of Amomi Fructus may play an important protective role in intestinal mucositis and gut barrier injury in rats challenged by 5-FU, 96 mg·kg~(-1) shows best effect, which may contribute to inhibit ROS generated, decrease pro-inflammatory cytokines secrete, inhibit NF-κB signaling, caspase-3 signaling and suppress the MPO activity, further decrease apoptosis, inhibit the translocation of LPS and protect the integrity of mucosal.
OBJECTIVE To study the effect of Amomi Fructus water extraction on intestinal mucosal barrier function in rats induced by 5-FU. METHODS SPF SD rats were divided into normal control group, model group, positive drug group and water extract of Amomi Fructus(48, 96, 192 mg·kg~(-1)) groups, 8 rats in each group. The normal control group was intraperitoneally injected with saline for 5 d, and the other groups were intraperitoneally injected with 35 mg·kg~(-1) 5-FU for 5 d. From the beginning of the experiment, the normal control group were given saline, and the other groups were given the corresponding therapeutic drugs for fasting and gavage for 12 d. Diarrhea occurrence, weight and food intakes were recorded daily throughout the study in rats. Blood samples and the small intestinal were collected after killing of rats for evaluation of histopathology, the content of cytokines(IL-6, ROS, NF-kB, MPO and TNF-α) and LPS were detected by ELISA. The intestinal tight junction protein ZO-1, occludin and caspase-3 were measured by flow cytometry. RESULTS Water extract of Amomi Fructus significantly improved rats body weight, food intakes diarrhea and significantly reversed the hisopathological changes in intestinal mucositis induced by 5-FU and reduced the inflammatory parameters(IL-6, ROS) in blood sample. It could also significantly effectively reduce the content of NF-κB, TNF-α, MPO in small intestinal tissue and the effect was similar to the positive drug. Meanwhile, it showed that water extract of Amomi Fructus could increase the expression of occludin and decrease the expression of caspase-3, inhibit the translocation of LPS, protect the integrity of intestinal barrier function. CONCLUSION It is suggested that the water extract of Amomi Fructus may play an important protective role in intestinal mucositis and gut barrier injury in rats challenged by 5-FU, 96 mg·kg~(-1) shows best effect, which may contribute to inhibit ROS generated, decrease pro-inflammatory cytokines secrete, inhibit NF-κB signaling, caspase-3 signaling and suppress the MPO activity, further decrease apoptosis, inhibit the translocation of LPS and protect the integrity of mucosal.
引文
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[20]BEUTHEU Y S,BELMONTE L,GALAS L,et al.Methotrexate modulates tight junctions through NF-?B,MEK and JNKpathways[J].J Pediatr Gastroenterol Nutr,2012,54(4):463-470.
[21]NAKAO T,KURITA N,KOMATSU M,et al.Irinotecan injures tight junction and causes bacterial translocation in rat[J].J Surg Res,2012,173(2):341-347.
[2]FILICKO J,LAGARUS H M,FLOMENBERG N.Mucosal injury in patients undergoing hematopoietic progenitor cell transplantation:new approaches to prophylaxis and treatment[J].Bone Marrow Transplant,2003,31(1):1-10.
[3]WEI W,LI B L,WANG C.Synthesis of 5-fluorouracil doped silica nanoparticles and their cytotoxicity[J].Chin J Mod Appl Pharm(中国现代应用药学),2016,33(1):59-62.
[4]WISCHMEYER P E.Glutamine:role in gut protection in critical illness[J].Curr Opin Clin Nutr Metab Care,2006,9(5):607-612.
[5]SOH H,WASA M,WANG H S,et a1.Glutamine regulates amino acid transport And glutamine levels in a human neuroblastoma cell line[J].Pediatr Surg Int,2005,21(1):29-33.
[6]中国药典.一部[S].2015:236.
[7]胡玉兰,张忠义,王文静,等.砂仁挥发油对大鼠乙酸性胃溃疡的影响及其机制探讨[J].中药材,2005,28(11):1022-1024.
[8]LI X G,YE F Q,XU H H.Study on pharmacological effects of bornyl acetate in volatile oil of Amomum villosum[J].West China J Pharm Sci(华西药学杂志),2001,16(5):356-358.
[9]郑虎占,董泽宏,余靖.中药现代研究与应用[M].第4卷.北京:学苑出版社,1998:3214-3229.
[10]邱赛红,陈立峰,柳克玲,等.芳香化湿药开胃作用机制的实验研究[J].中药药理与临床,1995,11(4):24-27.
[11]SAEGUSA Y,ICHIKAWA T,IWAI T,et al.Changes in the mucus barrier of the rat during 5-fluorouracil-induced gastrointestinal mucositis[J].Scand J Gastroenterol,2008,43(1):59-65.
[12]SONIS S T.The pathobiology of mucositis[J].Nat Rev Cancer,2005,4(4):277-284.
[13]ARIFA R D,MADEIRA M F,DE PAULA T P,et al.Inflammasome activation is reactive oxygen species dependent and mediates irinotecan-induced mucositis through IL-1βand IL-18 in mice[J].Am J Pathol,2014,184(7):2024-2034.
[14]SONIS S T.The biologic role of ueclear factor-κB in disease and its potential involvement in mucosal injury associated with antinepplastic therapy[J].Crit Rev Oral Biol Med,2002,13(5):380-389.
[15]RINKENBAUGH A L,BALDWIN A S.The NF-κB pathway and cancer stem cells[J].Cells,2016,5(2):2-19.
[16]赵丽云,叶铁龙.核因子-κB对细胞因子的基因调控与炎症反应[J].麻醉与监护论坛,2002,9(3):37-39.
[17]CHANG C T,HO T Y,LIN H,et al.5-Fluorouracil induced intestinal mucositis via nuclear factor-κb activation by transcriptomic analysis and in vivo bioluminescence imaging[J].PLoS One,2012,7(3):e31808.
[18]HAN F F,LU Z Q,LIU Y F,et al.Cathelicidin-BF ameliorates lipopolysaccharide-induced intestinal epithelial barrier disruption in rat[J].Life Sci,2016(152):199-209.
[19]SHETH P,DELOS SANTOS N,SETH A,et al.Lipopiysaccharide dispruts tight junctions in cholangicocyte monolayers by a c-Src-,TLR4-,and LBP-dependent mechanism[J].Am J Physiol Gastrointest Liver Physiol,2007,293(1):308-318.
[20]BEUTHEU Y S,BELMONTE L,GALAS L,et al.Methotrexate modulates tight junctions through NF-?B,MEK and JNKpathways[J].J Pediatr Gastroenterol Nutr,2012,54(4):463-470.
[21]NAKAO T,KURITA N,KOMATSU M,et al.Irinotecan injures tight junction and causes bacterial translocation in rat[J].J Surg Res,2012,173(2):341-347.