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1.安徽中医药大学,合肥 230012
2.安徽中医药大学 新安医学教育部重点实验室,合肥 230038
李姿慧,博士,副教授,从事经方防治脾胃病研究,E-mail:lizihui001@126.com
蔡荣林,博士,教授,从事中医药防治脾胃病研究,E-mail:ronglincai@163.com
收稿日期:2020-02-18,
网络出版日期:2020-07-29,
纸质出版日期:2020-10-05
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李姿慧,蔡荣林,孙娟等.参苓白术散对脾虚湿困型溃疡性结肠炎大鼠结肠组织NF-κB p65,IκBα,IκKβ蛋白及mRNA表达的影响[J].中国实验方剂学杂志,2020,26(19):108-113.
LI Zi-hui,CAI Rong-lin,SUN Juan,et al.Effect of Shenling Baizhusan on Protein and mRNA Expression of NF-κB p65,IκBα,IκKβ in Ulcerative Colitis Rats with Syndrome of Dampness Stagnancy Due to Spleen Deficiency[J].Chinese Journal of Experimental Traditional Medical Formulae,2020,26(19):108-113.
李姿慧,蔡荣林,孙娟等.参苓白术散对脾虚湿困型溃疡性结肠炎大鼠结肠组织NF-κB p65,IκBα,IκKβ蛋白及mRNA表达的影响[J].中国实验方剂学杂志,2020,26(19):108-113. DOI: 10.13422/j.cnki.syfjx.20201936.
LI Zi-hui,CAI Rong-lin,SUN Juan,et al.Effect of Shenling Baizhusan on Protein and mRNA Expression of NF-κB p65,IκBα,IκKβ in Ulcerative Colitis Rats with Syndrome of Dampness Stagnancy Due to Spleen Deficiency[J].Chinese Journal of Experimental Traditional Medical Formulae,2020,26(19):108-113. DOI: 10.13422/j.cnki.syfjx.20201936.
目的
2
观察参苓白术散对脾虚湿困型溃疡性结肠炎(UC)大鼠结肠组织核转录因子-
κ
B(NF-
κ
B)抑制蛋白激酶(I
κ
K)/NF-
κ
B抑制蛋白(I
κ
B)/NF-
κ
B信号通路的影响,探讨参苓白术散改善UC的作用机制。
方法
2
将48只Wistar大鼠按照随机数字表法分为正常组、模型组、参苓白术散(15.6 g·kg
-1
)组、奥沙拉嗪钠(0.68 g·kg
-1
)组,每组12只,脾虚湿困型UC大鼠模型采用2,4,6-三硝基苯磺酸(TNBS)/乙醇灌肠结合环境与饮食干预法复制,给药组连续灌胃14 d。采用酶联免疫吸附测定(ELISA)检测各组大鼠血清肿瘤坏死因子-
α
(TNF-
α
),白细胞介素-6(IL-6)及白细胞介素-1
β
(IL-1
β
)含量,免疫组化和蛋白免疫印迹法(Western blot)检测各组大鼠结肠组织中NF-
κ
B p65,I
κ
B
α
,I
κ
K
β
蛋白表达,实时荧光定量聚合酶链式反应(Real-time PCR)检测各组大鼠结肠组织NF-
κ
B p65,I
κ
B
α
,I
κ
K
β
mRNA表达。
结果
2
与正常组比较,模型组大鼠血清TNF-
α
,IL-6及IL-1
β
含量均显著升高(
P
<
0.01),结肠组织NF-
κ
B p65,I
κ
K
β
蛋白和mRNA表达显著升高(
P
<
0.01),I
κ
B
α
蛋白和mRNA表达显著降低(
P
<
0.01);与模型组比较,参苓白术散组和奥沙拉嗪钠组大鼠血清TNF-
α
,IL-6及IL-1
β
水平均显著下降(
P
<
0.01),结肠组织NF-
κ
B p65,I
κ
K
β
蛋白和mRNA均显著下降(
P
<
0.01),I
κ
B
α
蛋白和mRNA表达显著升高(
P
<
0.01)。
结论
2
参苓白术散能明显下调脾虚湿困型UC大鼠结肠组织NF-
κ
B p65,I
κ
K
β
,蛋白和mRNA表达,上调I
κ
B
α
蛋白和mRNA表达;参苓白术散抑制UC大鼠I
κ
K/I
κ
B/NF-
κ
B信号通路活化是其发挥肠黏膜保护作用的重要机制。
Objective
2
To observe the effect of Shenling Baizhusan on the protein and mRNA expression of inhibitor of nuclear factor kappa B kinase (I
κ
K)/inhibitor of nuclear factor kappa B(I
κ
B)/nuclear factor kappa B(NF-
κ
B) signaling pathway in the colon of rats with ulcerative colitis (UC) of spleen deficiency and dampness stagnation type
and to explore the mechanism of Shenling Baizhusan in the treatment of UC.
Method
2
The 48 Wistar rats were randomly divided into normal group
model group
Shenling Baizhusan group (15.6 g·kg
-1
) and osalazine sodium group (0.68 g·kg
-1
)
12 rats in each group. The model of UC with spleen deficiency and dampness stagnation was reproduced by trinitrobenzene sulfonic acid (TNBS)/ethanol enema combined with environment and diet intervention.Serum levels of tumour necrosis factor-
α
(TNF-
α
)
interleukin-6 (IL-6) and interleukin-1
β
(IL-1
β
) were determined by enzyme-linked immunosorbent assay(ELISA). The expression of NF-
κ
B p65
I
κ
B
α
I
κ
K
β
protein in colon tissue was measured by Western blot and immunohistochemical method
and the mRNA expression of NF-
κ
B p65
I
κ
B
α
and I
κ
K
β
in colon tissue of rats in each group was detected and compared by real time polymerase chain reaction (Real-time PCR).
Result
2
Compared with normal group
the levels of TNF-
α
IL-6 and IL-1
β
in the serum
the protein and mRNA expression of NF-
κ
B p65
I
κ
K
β
in colon tissue of the model group was significantly higher than that of normal group (
P
<
0.01)
and the protein and mRNA expression of I
κ
B
α
was significantly lower than that of normal group (
P
<
0.01). Compared with model group
the protein and mRNA expression of NF-
κ
B p65
I
κ
K
β
in colon tissue of the Shenling Baizhusan group and osalazine sodium group were significantly decreased (
P
<
0.01)
and the protein and mRNA expression of I
κ
B
α
was significantly increased (
P
<
0.01).
Conclusion
2
Shenling Baizhusan can obviously down regulate the protein and mRNA expression of NF-
κ
B p65
I
κ
K
β
up regulate the expression of I
κ
B
α
in colon tissue of UC rats with spleen deficiency and dampness stagnation. The inhibition of I
κ
K/I
κ
B/NF-
κ
B signal pathway activation by Shenling Baizhusan is an important mechanism of its role in protecting intestinal mucosa.
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