白术破壁饮片对慢传输型便秘小鼠肠神经递质及ICC的影响

Effect of Atractylodes macrocephala decoction pieces on enteric neurotransmitter and interstitial cells of Cajal in mice with slow transit constipation

摘要:

目的：探究白术破壁饮片对慢传输型便秘（slow transit constipation，STC）小鼠肠神经递质、Cajal间质细胞（interstitial cells of Cajal，ICC）的干细胞因子（stem cell factor，SCF）/干细胞生长因子受体（c-kit）信号通路的影响。方法：采用皮下注射2.5 mg/（kg·d）盐酸吗啡注射液法复制STC小鼠模型，选取48只雌雄各半昆明小鼠随机分为正常组，模型组，白术破壁饮片低、中、高剂量组（25、50、100 mg/kg）和莫沙必利组（1.5 mg/kg），每组8只。各组予以不同药物干预1周后，检测造模后和给药后各组小鼠粪便性状、肠道推进率；采用酶联免疫反应（enzyme-linked immunosorbent assay，ELISA）测定各组小鼠结肠神经递质P物质（substance P，SP）、乙酰胆碱（acetylcholine，Ach）、血管活性肠肽（vasoactive intestinal peptide，VIP）、一氧化氮（nitric oxide，NO）和5-羟色胺（5-hydroxytrytamine，5-HT）含量；采用Western blot检测小鼠结肠组织SCF和c-kit蛋白的表达。结果：ELISA结果显示，与正常组比较，模型组SP和Ach含量均明显降低（SP：230.41±16.41 vs. 146.69±15.59；Ach：577.68±39.35 vs. 281.50±39.40）。与模型组比较，白术破壁饮片低剂量组、白术破壁饮片中剂量组和白术破壁饮片高剂量组SP含量均明显升高（146.69±15.59 vs. 188.25±4.60；146.69±15.59 vs. 201.78±4.44；146.69±15.59 vs. 307.51±16.94）；白术破壁饮片高剂量组和莫沙必利组Ach含量均明显升高（281.50±39.40 vs. 400.93±12.21；281.50±39.40 vs. 422.22±22.70）。与白术破壁饮片高剂量组比较，白术破壁饮片低剂量组和白术破壁饮片中剂量组SP和Ach含量均明显降低（SP：307.51±16.94 vs.188.25±4.60；307.51±16.94 vs. 201.78±4.44；Ach：400.93±12.21 vs. 283.79±11.53；400.93±12.21 vs. 327.81±14.77）；莫沙必利组SP含量均明显降低（307.51±16.94 vs. 164.08±12.54）。与正常组比较，模型组VIP、NO和5-HT含量均明显升高（VIP： 64.47±2.69 vs. 87.74±2.93；NO：38.21±1.76 vs. 42.78±1.69；5-HT：219.58±11.60 vs. 276.08±7.97）。与模型组比较，白术破壁饮片低剂量组、白术破壁饮片中剂量组、白术破壁饮片高剂量组和莫沙必利组VIP含量均明显降低（87.74±2.93 vs. 75.58±2.08；87.74±2.93 vs. 69.34±2.23；87.74±2.93 vs. 66.37±1.93；87.74±2.93 vs. 65.31±3.32 ）；白术破壁饮片中剂量组、白术破壁饮片高剂量组和莫沙必利组NO和5-HT含量均明显降低（NO：42.78±1.69 vs. 39.27±1.90；42.78±1.69 vs. 37.43±1.30；42.78±1.69 vs. 35.65±2.01；5-HT：276.08±7.97 vs. 257.89±6.16；276.08±7.97 vs. 226.79±10.49；276.08±7.97 vs. 242.05±12.15）。与白术破壁饮片高剂量组比较，白术破壁饮片低剂量组VIP、NO和5-HT含量均明显升高（VIP：66.37±1.93 vs. 75.58±2.08；NO：37.43±1.30 vs. 42.73±2.19；5-HT：226.79±10.49 vs. 269.87±10.91）；白术破壁饮片中剂量组VIP、5-HT，莫沙必利组5-HT含量均明显升高。Western blot结果显示，与模型组比较，白术破壁饮片低剂量组c-kit蛋白含量明显升高（0.22±0.10 vs. 0.37±0.08）；白术破壁饮片中剂量组、白术破壁饮片高剂量组和莫沙必利组SCF和c-kit蛋白含量均明显升高（SCF：0.60±0.19 vs. 0.99±0.28；0.60±0.19 vs. 1.17±0.34；0.60±0.19 vs. 1.02±0.30；c-kit：0.22±0.10 vs. 0.47±0.10；0.22±0.10 vs. 0.58±0.13；0.22±0.10 vs. 0.49±0.13）。结论：白术破壁饮片可通过升高STC模型小鼠结肠组织 SP、Ach和SCF/c-kit含量，以及降低VIP、NO和5-HT含量，从而调节小鼠胃肠道功能，促进肠道蠕动，改善便秘症状，进而达到治疗STC的效果。

Abstract:

Objective：To investigate the effects of Atractylodes macrocephala decoction pieces on enteric neurotransmitter and stem cell factor（SCF）/stem cell growth factor receptor（c-kit） signaling pathway of interstitial cells of Cajal（ICC） in mice with slow transit constipation（STC）. Methods：The STC mouse model was replicated by subcutaneous injection of 2.5 mg/（kg·d） morphine hydrochloride injection. A total of 48 Kunming mice were randomly divided into normal group，model group，Atractylodes macrocephala decoction pieces low，medium，high-dose groups（25，50，100 mg/kg） and mosapride group（1.5 mg/kg），each with 8 mice. After each group was treated with different drugs for 1 week，the stool characteristics and intestinal propulsion rate of mice in each group were tested after modeling and administration. Enzyme-linked immunosorbent assay（ELISA） was conducted to determine neurotransmitter substance P（SP），acetylcholine（Ach），vasoactive intestinal peptide（VIP），nitric oxide（NO） and 5-hydroxytrytamine（5-HT） contents in the colon of the mice in each group. And Western blot was used to detect the expression of SCF and c-kit protein in mouse colon tissue. Results：ELISA results showed that compared with the normal group，the SP and Ach contents of the model group were significantly reduced（SP：230.41±16.41 vs. 146.69±15.59；Ach：577.68±39.35 vs. 281.50±39.40）. Compared with the model group，the SP content in the low-dose group of Atractylodes macrocephala，the middle-dose group and the high-dose group of Atractylodes macro－cephala fragments were significantly increased（146.69±15.59 vs. 188.25±4.60；146.69±15.59 vs. 201.78±4.44；146.69±15.59 vs. 307.51±16.94）；Ach content in high-dose Atractylodes macrocephala and mosapride group were significantly increased（281.50±39.40 vs. 400.93±12.21；281.50±39.40 vs. 422.22±22.70 ）. Compared with the high-dose group of Atractylodes macrocephala decoction pieces，the SP and Ach contents of the low-dose group of Atractylodes macrocephalus decoction pieces and the middle-dose group of Atractylodes macrocephalus decoction pieces were significantly reduced（SP：307.51±16.94 vs. 188.25±4.60；307.51±16.94 vs. 201.78±4.44；Ach：400.93±12.21 vs. 283.79±11.53；400.93±12.21 vs. 327.81±14.77）；SP content in the mosapride group was significantly reduced（307.51±16.94 vs. 164.08±12.54）. Compared with the normal group，the contents of VIP，NO and 5-HT in the model group were significantly increased（VIP：64.47±2.69 vs. 87.74±2.93；NO：38.21±1.76 vs. 42.78±1.69；5-HT：219.58±11.60 vs. 276.08±7.97）. Compared with the model group，the VIP content of the low-dose Atractylodes macrocephala decoction pieces，the medium-dose Atractylodes macrocephalus decoction pieces，the high-dose Atractylodes macrocephalus decoction pieces，and the mosapride group were significantly reduced（87.74±2.93 vs. 75.58±2.08；87.74±2.93 vs. 69.34±2.23；87.74±2.93 vs. 66.37±1.93；87.74±2.93 vs. 65.31±3.32）；Atractylodes macrocephala medium dose group，Atractylodes macrocephala fragment high dose group and mosapride group NO and The content of 5-HT was significantly reduced（NO：42.78±1.69 vs. 39.27±1.90；42.78±1.69 vs. 37.43±1.30；42.78±1.69 vs. 35.65±2.01；5-HT：276.08±7.97 vs. 257.89±6.16；276.08±7.97 vs. 226.79±10.49；276.08±7.97 vs. 242.05±12.15）. Compared with the high-dose Atractylodes macrocephala decoction pieces group，the contents of VIP，NO and 5-HT in the low-dose Atractylodes macrocephalus decoction pieces group were significantly increased（VIP：66.37±1.93 vs. 75.58±2.08；NO：37.43±1.30 vs. 42.73±2.19；5-HT：226.79±10.49 vs. 269.87±10.91）；VIP，5-HT in the middle-dose group of Atractylodes macrocephala and 5-HT in the mosapride group were significantly increased. Western blot results showed that compared with the model group，the c-kit protein content in the low-dose group of Atractylodes macrocephala was significantly increased（0.22±0.10 vs. 0.37±0.08）；the middle-dose group of Atractylodes macrocephala and the high-dose group of Atractylodes macrocephala both the SCF and c-kit protein content in the mosapride group and the mosapride group were significantly increased（SCF：0.60±0.19 vs. 0.99±0.28；0.60±0.19 vs. 1.17±0.34；0.60±0.19 vs. 1.02±0.30；c-kit：0.22±0.10 vs. 0.47±0.10；0.22±0.10 vs. 0.58±0.13；0.22±0.10 vs. 0.49±0.13）. Conclusion：Atractylodes macrocephala decoction pieces can increase the contents of SP，Ach and SCF/c-kit in the colon tissue of STC model mice，and reduce the contents of VIP，NO and 5-HT，thereby regulating the gastrointestinal function of mice，promoting the intestinal tract peristalsis，improving constipation symptoms，and then achieving the effect of treating STC.