DAI Lu-ming, ZHU Lei, SHEN Hong. Effect of Qingchang Huashi Granule on AR/-arrestin2/NF-B Signaling Pathway in Ulcerative Colitis[J]. Chinese journal of experimental traditional medical formulae, 2018, 24(9): 86-94.
DAI Lu-ming, ZHU Lei, SHEN Hong. Effect of Qingchang Huashi Granule on AR/-arrestin2/NF-B Signaling Pathway in Ulcerative Colitis[J]. Chinese journal of experimental traditional medical formulae, 2018, 24(9): 86-94. DOI: 10.13422/j.cnki.syfjx.20180837.
目的:观察清肠化湿颗粒对溃疡性结肠炎(ulcerative colitis,UC)模型大鼠及人结肠癌上皮细胞株HT-29炎症模型β2肾上腺素受体(β2AR)/β-抑制蛋白2(β-arrestin2)/核转录因子-κB(NF-κB)信号通路的干预作用。方法:取8只大鼠作为空白组,其余大鼠采用三硝基苯磺酸(trinitro-benzene-sulfonic acid,TNBS)灌肠法复制UC大鼠模型;待模型建立成功后,随机分为模型组,柳氮磺胺吡啶(SASP,1.0 g ·kg-1)组,清肠化湿颗粒低、中、高剂量(2.8,5.5,11.0 g ·kg-1)组,每组8只,灌胃给药10 d,每日1次;酶联免疫吸附法(enzyme linked immunosorbent assay,ELISA)测定UC大鼠结肠组织中巨噬细胞移动抑制因子(MIF),基质金属蛋白酶-1(MMP-1),MMP-2,MMP-9,胰岛素样生长因子-1(IGF-1),超氧化物歧化酶(SOD),丙二醛(MDA),髓过氧化物酶(MPO),一氧化氮(NO),一氧化氮合成酶(iNOS),谷胱甘肽-过氧化物酶(GSH-Px);免疫组化法检测模型大鼠结肠黏膜β2AR,β-arrestin2,NF-κB定位表达;采用肿瘤坏死因子与脂多糖诱导HT-29细胞炎症模型;噻唑蓝(MTT)比色法检测清肠化湿颗粒对细胞增殖的影响,蛋白免疫印迹法(Western blot)检测细胞中β2AR,β-arrestin2,NF-κB蛋白表达量。结果:造模后,模型组大鼠较空白组大鼠结肠黏膜的结肠损伤分数明显升高;与模型组比较,3个剂量的清肠化湿颗粒给药组结肠黏膜损伤明显下降(P<0.05,P<0.01),大鼠结肠组织IGF-1,SOD,GSH-Px含量与β2AR,β-arrestin2表达明显上升,MIF,MMP-2,MMP-9,MDA,MPO,NO,iNOS含量及NF-κB表达明显下降(P<0.05,P<0.01)。结论:清肠化湿颗粒可缓解UC氧化应激反应及肠道炎症,其作用与激活β2AR/β-arrestin2/NF-κB信号通路有关。
Abstract
Objective: To study the effect of Qingchang Huashi granule on β2 adrenergic receptor(β2AR)/β-arrestin2/nuclear transcription factor-κB(NF-κB) signaling pathway in rat and human ulcerative colitis. Method: Eight rats were randomly selected as control group
while the other rats were used to duplicate the ulcerative colitis model induced by trinitrobenzene sulfonic acid; after successful modeling
the rats were randomly divided into model group
salazosulfapyridine(SASP) group (1.0 g · kg-1)
and low
medium and high-dose Qingchang Huashi granule groups (2.8
5.5
11.0 g · kg-1)
with 8 rats in each group. The drugs were given once a day by gavage for 15 days. The levels of macrophage mobile inhibitory factor(MIF)
matrix metalloproteinase-1(MMP-1)
macrophage migration inhibitory factor (MIF)
matrix metalloproteinase-1 (MMP-1)
MMP-2
MMP-9
insulin-like growth factor-1 (IGF-1)
superoxide dismutase (SOD)
malondialdehyde (MDA)
myeloperoxidase (MPO)
nitric oxide (NO)
nitric oxide synthase (iNOS)
glutathione peroxidase (GSH-Px) in experimental UC rats were measured by enzyme linked immunosorbent assay method(ELISA); immunohistochemical method was used to detect the location expressions of β2AR
β-arrestin2
NF-κB in rat models; tumor necrosis factor alpha and lipopolysaccharide were used to induce the HT-29 cell inflammation model; the effect of Qingchang Huashi granule on cell growth was detected by MTT method
the expressions of β2AR
β-arrestin2
NF-κB were measured with Western blot. Result: The injury score of colonic mucosa in model group rats was significantly higher than that of the normal group
expressions of IGF-1
SOD
GSH-Px
β2AR and β-arrestin2 decreased significantly
while expressions of MIF
MMP-2
MMP-9
MDA
MPO
NO
iNOS and NF-κB increased in UC model group (P<0.05
P<0.01). After intervened with Qingchang Huashi granule
these indicators all recovered to normal. Conclusion: Qingchang Huashi granule can reduce the pathological process of ulcerative colitis through β2AR/β-arrestin2/NF-κB signaling pathway.