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1. 安徽中医药大学第二临床医学院
2. 安徽中医药大学第二附属医院/安徽省针灸医院名医堂
3. 安徽省中医药科学院针灸临床研究所
4. 安徽省针灸临床医学研究中心
纸质出版:2023
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李奎武, 储浩然, 阮静茹, 等. 基于SCF/c-kit信号通路探讨艾灸干预对腹泻型肠易激综合征大鼠免疫功能稳态的影响[J]. 中国针灸, 2023,43(2):177-185.
Effect of moxibustion on immune function homeostasis in rats with diarrhea irritable bowel syndrome based on SCF/c-kit signaling pathway[J]. Chinese Acupuncture and Moxibustion, 2023, 43(2): 177-185.
李奎武, 储浩然, 阮静茹, 等. 基于SCF/c-kit信号通路探讨艾灸干预对腹泻型肠易激综合征大鼠免疫功能稳态的影响[J]. 中国针灸, 2023,43(2):177-185. DOI: 10.13703/j.0255-2930.20220108-k0007.
Effect of moxibustion on immune function homeostasis in rats with diarrhea irritable bowel syndrome based on SCF/c-kit signaling pathway[J]. Chinese Acupuncture and Moxibustion, 2023, 43(2): 177-185. DOI: 10.13703/j.0255-2930.20220108-k0007.
目的:观察艾灸对腹泻型肠易激综合征(IBS-D)模型大鼠干细胞因子(SCF)/酪氨酸激酶受体(c-kit)信号通路及免疫功能的影响,探讨艾灸干预IBS-D的机制。方法:从6只健康SPF级孕鼠产下的52只幼鼠中随机选取12只作为正常组,剩余40只采用母子分离、醋酸灌肠、慢性束缚应激三因素联合复制IBS-D大鼠模型,将造模成功的36只大鼠随机分为模型组、艾灸组、西药组,每组12只。艾灸组予悬灸“天枢”“上巨虚”,西药组予利福昔明悬浊液(150 mg/kg)灌胃,均每天1次,连续干预7 d。分别于醋酸灌肠前(35日龄)、造模后(45日龄)、干预后(53日龄)检测各组大鼠体质量、稀便率(LSR)、腹壁回缩反射(AWR)评分达3分时最小容量阈值;干预后(53日龄),HE染色观察大鼠结肠组织形态,测定脾脏及胸腺脏器系数,ELISA法检测血清炎性因子[肿瘤坏死因子α(TNF-α)、白介素(IL)-10、IL-8]、T淋巴细胞亚群(CD_4
+
、CD_8+、CD_8
+
、CD_(45)+、CD_(45)
+
)、CD_4+)、CD_4
+
/CD_8+/CD_8
+
值及免疫球蛋白(Ig A、Ig G、Ig M)含量,实时荧光定量PCR法、Western blot法检测大鼠结肠组织SCF、c-kit m RNA及蛋白表达,免疫荧光法检测大鼠结肠组织SCF、c-kit阳性表达。结果:干预后,与正常组比较,模型组体质量、AWR评分达3分时最小容量阈值降低(P
<
0.01)
LSR、脾脏及胸腺系数与血清TNF-α、IL-8、CD_4+值及免疫球蛋白(Ig A、Ig G、Ig M)含量,实时荧光定量PCR法、Western blot法检测大鼠结肠组织SCF、c-kit m RNA及蛋白表达,免疫荧光法检测大鼠结肠组织SCF、c-kit阳性表达。结果:干预后,与正常组比较,模型组体质量、AWR评分达3分时最小容量阈值降低(P
<
0.01)
LSR、脾脏及胸腺系数与血清TNF-α、IL-8、CD_4
+
、CD_8+、CD_8
+
、CD_(45)+、CD_(45)
+
、CD_4+、CD_4
+
/CD_8+/CD_8
+
值、Ig A、Ig G、Ig M含量升高(P
<
0.01),血清IL-10与结肠组织SCF、c-kit蛋白及m RNA表达降低(P
<
0.01),结肠组织SCF、c-kit阳性表达降低(P
<
0.01)。与模型组比较,艾灸组和西药组体质量、AWR评分达3分时最小容量阈值升高(P
<
0.01
P
<
0.05)
LSR、脾脏及胸腺系数与血清TNF-α、IL-8、CD_4+值、Ig A、Ig G、Ig M含量升高(P
<
0.01),血清IL-10与结肠组织SCF、c-kit蛋白及m RNA表达降低(P
<
0.01),结肠组织SCF、c-kit阳性表达降低(P
<
0.01)。与模型组比较,艾灸组和西药组体质量、AWR评分达3分时最小容量阈值升高(P
<
0.01
P
<
0.05)
LSR、脾脏及胸腺系数与血清TNF-α、IL-8、CD_4
+
、CD_8+、CD_8
+
、CD_(45)+、CD_(45)
+
、CD_4+、CD_4
+
/CD_8+/CD_8
+
值、Ig A、Ig G、Ig M含量降低(P
<
0.01
P
<
0.05),血清IL-10与结肠组织SCF、c-kit蛋白及m RNA表达升高(P
<
0.01),结肠组织SCF及c-kit阳性表达升高(P
<
0.01)。与西药组比较,艾灸组血清CD_4+值、Ig A、Ig G、Ig M含量降低(P
<
0.01
P
<
0.05),血清IL-10与结肠组织SCF、c-kit蛋白及m RNA表达升高(P
<
0.01),结肠组织SCF及c-kit阳性表达升高(P
<
0.01)。与西药组比较,艾灸组血清CD_4
+
含量降低(P
<
0.05)
CD_4+含量降低(P
<
0.05)
CD_4
+
/CD_8+/CD_8
+
值升高(P
<
0.01),其余指标比较差异无统计学意义(P
>
0.05)。SCF、c-kit m RNA表达与AWR评分达3分时最小容量阈值、IL-10呈正相关(P
<
0.01),与其余指标均呈负相关(P
<
0.01
P
<
0.05)。结论:艾灸干预可降低IBS-D模型大鼠内脏高敏感性,改善其腹痛、腹泻等症状,其作用机制可能与上调SCF/c-kit信号通路的表达、改善IBS-D免疫功能状态有关。
Objective To observe the effects of moxibustion on the stem cell factor(SCF)/tyrosine kinase receptor(c-kit) signaling pathway and immune function in rats with diarrhea irritable bowel syndrome(IBS-D)
and to explore the mechanism of moxibustion for IBS-D. Methods Among 52 young rats born from 6 healthy pregnant SPF rats
12 rats were randomly selected into the normal group
and the remaining 40 rats were treated with the three-factor combination method of maternal separation
acetic acid enema and chronic restraint stress to establish the IBS-D rat model. Thirty-six rats with successful IBS-D model were randomly divided into a model group
a moxibustion group
and a medication group
12 rats in each group. The rats in the moxibustion group were treated with suspension moxibustion at "Tianshu"(ST 25) and "Shangjuxu"(ST 37); the rats in the medication group were treated with intragastric administration of rifaximin suspension(150 mg/kg). All the treatments were given once a day for 7 consecutive days. The body mass
loose stool rate(LSR)
the minimum volume threshold when abdominal withdrawal reflex(AWR) scored 3 were measured before acetic acid enema(35 days old)
after modeling(45 days old)
and after intervention(53 days old). After intervention(53 days old)
HE staining was used to observe the morphology of colon tissue
and spleen and thymus coefficients were measured; ELISA method was used to detect serum inflammatory factors(tumor necrosis factor α[TNF-α]
interleukin [IL]
-10
IL-8)
T-lymphocyte subsets(CD
4
+
CD
8
+
CD
45
+
)
value of CD
4
+
/CD
8
+
and immune globulin(Ig A
Ig G
Ig M); real-time PCR method and Western blot method was used to detect the expression of SCF
c-kit m RNA and protein in colon tissue; immunofluorescence staining method were used to detect positive expression of SCF and c-kit.Results After intervention
compared with the normal group
in the model group
the body mass and the minimum volume threshold when AWR scored 3 were decreased(P
<
0.01)
LSR
spleen and thymus coefficients
serum levels of TNF-α
IL-8
CD
4
+
CD
45
+
CD
4
+
/CD
8
+
Ig A
Ig G
Ig M were increased(P
<
0.01)
serum IL-10 level and protein and m RNA expression of SCF and c-kit in colon tissue were decreased(P
<
0.01)
and the positive expression of SCF and c-kit was decreased(P
<
0.01). Compared with the model group
in the moxibustion group and the medication group
the body mass and the minimum volume threshold when AWR scored 3 were increased(P
<
0.01
P
<
0.05)
LSR
spleen and thymus coefficients
serum levels of TNF-α
IL-8
CD
4
+
CD
8
+
CD
45
+
CD
4
+
/CD
8
+
Ig A
Ig G
Ig M were decreased(P
<
0.01
P
<
0.05)
serum IL-10 level and protein and m RNA expression of SCF and c-kit in colon tissue were increased(P
<
0.01)
and the positive expression of SCF and c-kit was increased(P
<
0.01). Compared with the medication group
in the moxibustion group
the level of serum CD
4
+
was decreased(P
<
0.05)
the value of CD
4
+
/CD
8
+
was increased(P
<
0.01)
and there was no significant difference in other indexes(P
>
0.05). The expression of SCF and c-kit m RNA was positively correlated with the minimum volume threshold when AWR scored 3 and IL-10(P
<
0.01)
and negatively correlated with remaining indexes(P
<
0.01
P
<
0.05). Conclusion Moxibustion could reduce visceral hypersensitivity
improve symptoms of abdominal pain and diarrhea in IBS-D rats
and its mechanism may be related to up-regulation of the expression of SCF/c-kit signaling pathway and improvement of IBS-D immune function.
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