Mechanism of Buyang Huanwutang Combined with Bone Marrow Mesenchymal Stem Cell Transplantation in Treatment of Spinal Cord Injury Based on PI3K/Akt Signaling Pathway

XU Yang; WU Chenglin; GUO Dehua; YU Yinzhen; ZHANG Guofu

doi:10.13422/j.cnki.syfjx.20221805

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Mechanism of Buyang Huanwutang Combined with Bone Marrow Mesenchymal Stem Cell Transplantation in Treatment of Spinal Cord Injury Based on PI3K/Akt Signaling Pathway

更新时间：2023-01-17

- Mechanism of Buyang Huanwutang Combined with Bone Marrow Mesenchymal Stem Cell Transplantation in Treatment of Spinal Cord Injury Based on PI3K/Akt Signaling Pathway
- Chinese Journal of Experimental Traditional Medical Formulae Vol. 29, Issue 4, Pages: 9-17(2023)
- 作者机构：
  
  1.江西中医药大学，南昌 330004
  2.江西中医药大学附属医院，南昌 330006
- 作者简介：
- 基金信息：
- DOI：10.13422/j.cnki.syfjx.20221805
  CLC： R2-0;R22;R285.5;R289;R33
- Published：20 February 2023，
  
  Published Online：20 September 2022，
  
  Received：11 July 2022，
- 稿件说明：
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许洋,吴成林,郭德华等.基于PI3K/Akt信号通路探究补阳还五汤联合骨髓间质干细胞移植治疗脊髓损伤的作用机制[J].中国实验方剂学杂志,2023,29(04):9-17.

XU Yang,WU Chenglin,GUO Dehua,et al.Mechanism of Buyang Huanwutang Combined with Bone Marrow Mesenchymal Stem Cell Transplantation in Treatment of Spinal Cord Injury Based on PI3K/Akt Signaling Pathway[J].Chinese Journal of Experimental Traditional Medical Formulae,2023,29(04):9-17.
许洋,吴成林,郭德华等.基于PI3K/Akt信号通路探究补阳还五汤联合骨髓间质干细胞移植治疗脊髓损伤的作用机制[J].中国实验方剂学杂志,2023,29(04):9-17. DOI： 10.13422/j.cnki.syfjx.20221805.

XU Yang,WU Chenglin,GUO Dehua,et al.Mechanism of Buyang Huanwutang Combined with Bone Marrow Mesenchymal Stem Cell Transplantation in Treatment of Spinal Cord Injury Based on PI3K/Akt Signaling Pathway[J].Chinese Journal of Experimental Traditional Medical Formulae,2023,29(04):9-17. DOI： 10.13422/j.cnki.syfjx.20221805.

摘要

目的

探讨补阳还五汤联合骨髓间质干细胞（BMSCs）移植治疗脊髓损伤（SCI）的作用机制。

方法

用不同浓度（12.5、25、50 g·kg

-1

）的补阳还五汤灌胃，通过改良Tarlov评分测定大鼠脊髓功能，筛选出补阳还五汤最合适的浓度。再将SD大鼠分为6组，即假手术组（灌胃等体质生理盐水）、模型组（灌胃等体质生理盐水）、补阳还五汤组（灌胃25 g·kg

-1

补阳还五汤）、BMSCs移植组（尾静脉注射BMSCs 1 mL）、补阳还五汤+BMSCs组（灌胃25 g·kg

-1

补阳还五汤同时尾静脉注射BMSCs 1 mL）、补阳还五汤+BMSCs+LY294002组（灌胃25 g·kg

-1

补阳还五汤，同时尾静脉注射BMSCs 1 mL和LY294002 40 mg·kg

-1

），各10只。改良Tarlov评分、斜板试验和皮层体感诱发电位波潜伏期测定大鼠脊髓功能；免疫组化法检测脊髓组织中5-溴-2-脱氧尿嘧啶核苷（Brdu）标记的阳性细胞数；蛋白免疫印迹法（Western blot）检测脊髓组织中磷酸化蛋白激酶B（p-Akt）、糖蛋白130（gp130）、白细胞介素-6（IL-6）蛋白表达。

结果

与假手术组比较，模型组的Tarlov评分和倾斜平面临界角度明显降低（

0.05），皮层体感诱发电位波潜伏期和p-Akt、gp130、IL-6蛋白表达均明显升高（

0.05）。与模型组比较，假手术组和各治疗组的Tarlov评分和倾斜平面临界角度均明显升高（

0.05），皮层体感诱发电位波潜伏期和p-Akt、gp130、IL-6蛋白表达均明显降低（

0.05）。和BMSCs组比较，补阳还五汤+BMSCs组Tarlov评分和倾斜平面临界角度明显升高（

0.05），皮层体感诱发电位波潜伏期和p-Akt、gp130、IL-6蛋白表达明显降低（

0.05），移植5周后，补阳还五汤+BMSCs组Brdu标记阳性细胞数明显增多（

0.05）。和补阳还五汤+BMSCs组比较，补阳还五汤+BMSCs+LY294002组Tarlov评分和倾斜平面临界角度明显升高（

0.05），皮层体感诱发电位波潜伏期和p-Akt、gp130、IL-6蛋白表达明显降低（

0.05），移植5周后，补阳还五汤+BMSCs+LY294002组Brdu标记阳性细胞数明显增多（

0.05）。

结论

补阳还五汤通过抑制磷脂酰肌醇3-激酶（PI3K）/Akt信号促进BMSCs迁移进而恢复脊髓功能。

Abstract

Objective

To explore the mechanism of Buyang Huanwutang combined with bone marrow mesenchymal stem cell （BMSC） transplantation in the treatment of spinal cord injury （SCI）.

Method

Different concentrations （12.5， 25， 50 g·kg

-1

） of Buyang Huanwutang were administrated to rats by gavage. The spinal cord function of rats was measured by modified Tarlov score， and the most suitable concentration of Buyang Huanwutang was screened out. SD rats were then divided into 6 groups， namely， the sham operation group （gavage of equal amount of normal saline）， the model group （gavage of equal amount of normal saline）， the Buyang Huanwutang group （gavage of 25 g·kg

-1

Buyang Huanwutang）， the BMSC transplantation group （tail vein injection of BMSCs 1 mL）， the Buyang Huanwutang+BMSC group （gavage of 25 g·kg

-1

Buyang Huanwutang and tail vein injection of BMSCs 1 mL）， the Buyang Huanwutang+BMSC+LY294002 group （gavage of 25 g·kg

-1

Buyang Huanwutang and tail vein injection of BMSCs 1 mL and 40 mg·kg

-1

LY294002）， with 10 rats in each group. The spinal cord function was measured by the modified Tarlov score， inclined plate test， and latency of cortical somatosensory evoked potential. Immunohistochemistry was used to detect the number of 5-bromo-2-deoxyuracil nucleoside （Brdu）-labeled positive cells in the spinal cord tissue. The protein expression levels of phosphorylated protein kinase B （p-Akt）， glycoprotein 130 （gp130）， and interleukin-6 （IL-6） in spinal cord were detected by Western blot.

Result

As compared with the sham operation group， the Tarlov score and the critical angle of tilt plane in the model group were significantly decreased （

0.05）， and the latency of cortical somatosensory evoked potential wave and the protein expression levels of p-Akt， gp130， and IL-6 were significantly increased （

0.05）. As compared with the model group， the Tarlov score and the critical angle of tilt plane in the sham operation group and each treatment group were significantly increased （

0.05）， and the latency of cortical somatosensory evoked potential wave and the protein expression levels of p-Akt， gp130， and IL-6 were significantly decreased （

0.05）. As compared with the BMSC group， the Tarlov score and the critical angle of inclined plane in the Buyang Huanwutang+BMSC group increased （

0.05）， the latency of cortical somatosensory evoked potential wave and the protein expression levels of p-Akt， gp130， and IL-6 decreased （

0.05）， and the number of Brdu-labeled positive cells increased 5 weeks after transplantation （

0.05）. As compared with the Buyang Huanwutang+BMSC group， the Tarlov score and the critical angle of the inclined plane in the Buyang Huanwutang+BMSC+LY294002 group increased （

0.05）， and the latency of cortical somatosensory evoked potential wave and the protein expression levels of p-Akt， gp130， and IL-6 decreased significantly （

0.05）. Five weeks after transplantation， the number of Brdu-labeled positive cells increased significantly in the Buyang Huanwutang+BMSC+LY294002 group （

0.05）.

Conclusion

Buyang Huanwutang can promote BMSCs migration and restore spinal cord function by inhibiting phosphatidylinositol 3-kinase （PI3K）/protein kinase B （Akt） signal.

关键词

脊髓损伤补阳还五汤骨髓间充质干细胞磷脂酰肌醇3-激酶（PI3K）/蛋白激酶B（Akt）信号通路

Keywords

spinal cord injuryBuyang Huanwutangbone marrow mesenchymal stem cellsphosphoinositide 3-kinase/protein kinase B （PI3K/Akt） signaling pathway

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