The Effect of <italic style="font-style: italic">Zhiqiao Gancao Decoction</italic> （枳壳甘草汤） on Intervertebral Disc Nucleus Pulposus Cell Apoptosis and the Hippo-YAP/TAZ Signaling Pathway in Tail Intervertebral Disc Degeneration Model Rats

ZHU Zaishi; HUANG Zeling; CHEN Junming; XU Bo; LU Binjie; CHEN Hua; DUAN Xingxing; LI Yuwei; SHEN Xiaofeng

doi:10.13288/j.11-2166/r.2025.05.012

您当前的位置：

首页 >

文章列表页 >

The Effect of Zhiqiao Gancao Decoction （枳壳甘草汤） on Intervertebral Disc Nucleus Pulposus Cell Apoptosis and the Hippo-YAP/TAZ Signaling Pathway in Tail Intervertebral Disc Degeneration Model Rats

更新时间：2025-03-06

- The Effect of Zhiqiao Gancao Decoction （枳壳甘草汤） on Intervertebral Disc Nucleus Pulposus Cell Apoptosis and the Hippo-YAP/TAZ Signaling Pathway in Tail Intervertebral Disc Degeneration Model Rats
- Journal of Traditional Chinese Medicine Vol. 66, Issue 5, Pages: 509-517(2025)
- 作者机构：
  
  1.南京中医药大学附属苏州市中医医院，江苏省苏州市杨素路18号，215009
  2.苏州市吴门医派研究院骨伤研究所
- 作者简介：
- 基金信息：
- DOI：10.13288/j.11-2166/r.2025.05.012
  CLC：
- Received：12 November 2024，
  
  Revised：2024-12-23，
  
  Published：02 March 2025
- 稿件说明：
移动端阅览
朱在师,黄泽灵,陈俊名等.枳壳甘草汤对椎间盘退变模型大鼠尾椎间盘髓核组织细胞凋亡及Hippo-YAP/TAZ信号通路的影响[J].中医杂志,2025,66(05):509-517.

ZHU Zaishi,HUANG Zeling,CHEN Junming,et al.The Effect of Zhiqiao Gancao Decoction （枳壳甘草汤） on Intervertebral Disc Nucleus Pulposus Cell Apoptosis and the Hippo-YAP/TAZ Signaling Pathway in Tail Intervertebral Disc Degeneration Model Rats[J].Journal of Traditional Chinese Medicine,2025,66(05):509-517.
朱在师,黄泽灵,陈俊名等.枳壳甘草汤对椎间盘退变模型大鼠尾椎间盘髓核组织细胞凋亡及Hippo-YAP/TAZ信号通路的影响[J].中医杂志,2025,66(05):509-517. DOI： 10.13288/j.11-2166/r.2025.05.012.

ZHU Zaishi,HUANG Zeling,CHEN Junming,et al.The Effect of Zhiqiao Gancao Decoction （枳壳甘草汤） on Intervertebral Disc Nucleus Pulposus Cell Apoptosis and the Hippo-YAP/TAZ Signaling Pathway in Tail Intervertebral Disc Degeneration Model Rats[J].Journal of Traditional Chinese Medicine,2025,66(05):509-517. DOI： 10.13288/j.11-2166/r.2025.05.012.

摘要

目的

基于河马（Hippo）-Yes相关蛋白（YAP）/带有PDZ结合基序的转录共激活因子（TAZ）信号通路探讨吴门枳壳甘草汤延缓椎间盘退变的可能作用机制。

方法

50只SD大鼠随机分为假手术组、模型组、枳壳甘草汤低剂量组、枳壳甘草汤高剂量组、枳壳甘草汤高剂量+抑制剂组，每组10只。假手术组大鼠采用21G针头在尾部Co6/7/8节段穿刺皮肤和肌肉（深度约为2 mm），不损伤椎间盘，其余各组大鼠在尾部Co6/7/8节段用21G针头穿刺尾椎间盘5 mm建立椎间盘退变大鼠模型。造模1周后，枳壳甘草汤低、高剂量组分别予6.24、12.24 g/（kg·d）枳壳甘草汤灌胃，枳壳甘草汤高剂量+抑制剂组大鼠给予12.24 g/（kg·d）枳壳甘草汤灌胃后，按10 mg/kg腹腔注射YAP/TAZ抑制剂Verteporfin，假手术组与模型组大鼠予5 ml/ （kg·d）生理盐水灌胃。灌胃每日1次，腹腔注射隔日1次。各组均连续干预4周后，采用HE染色、番红O-固绿染色及甲苯胺蓝染色观察大鼠尾椎间盘病理改变；免疫组化法检测椎间盘髓核组织聚集蛋白聚糖（Aggrecan）、基质金属蛋白酶3（MMP3）表达；Tunel荧光染色检测髓核组织细胞凋亡，计算凋亡率；Western blot法检测髓核组织中Hippo-YAP/TAZ信号通路［包括YAP、磷酸化Yes相关蛋白（p-YAP）、磷酸化20样激酶1/2（p-MST1/2）、带有PDZ结合基序的磷酸化转录共激活因子（p-TAZ）］及凋亡相关蛋白［包括剪切的含半胱氨酸的天冬氨酸蛋白水解酶（Cleaved Caspase 3）、肿瘤基因P53（P53）、B淋巴细胞瘤2（Bcl-2）、Bcl-2相关X蛋白（Bax）］水平。

结果

与假手术组比较，模型组大鼠椎间盘发生明显退行性改变，椎间盘髓核组织Aggrecan、Bcl-2、YAP蛋白水平降低，p-MST1/2、p-YAP、p-TAZ、P53、Bax、Cleaved Caspase 3、MMP3蛋白水平及细胞凋亡率均上升（

＜0.01）。与模型组比较，各药物干预组椎间盘退变均得到一定恢复，Aggrecan、Bcl-2、YAP蛋白水平均升高，p-MST1/2、p-YAP、p-TAZ、P53、Bax、Cleaved Caspase 3、MMP3蛋白水平及细胞凋亡率均降低（

＜0.05或

＜0.01）。与枳壳甘草汤低剂量组比较，枳壳甘草汤高剂量组大鼠椎间盘退变恢复更显著，p-MST1

/2、p-YAP、p-TAZ、P53、Bax、Cleaved Caspase 3、MMP3蛋白水平及细胞凋亡率降低，Aggrecan、Bcl-2、YAP蛋白水平升高（

＜0.05或

＜0.01）。与枳壳甘草汤高剂量组比较，枳壳甘草汤高剂量+抑制剂组大鼠椎间盘退变恢复程度降低，p-MST1/2、p-YAP、p-TAZ、P53、Bax、Cleaved Caspase 3、MMP3蛋白水平及细胞凋亡率升高，Aggrecan、Bcl-2、YAP蛋白水平降低（

＜0.05或

＜0.01）。

结论

枳壳甘草汤可能通过抑制椎间盘髓核组织YAP磷酸化，维持Hippo-YAP/TAZ信号通路功能，降低髓核细胞凋亡，从而延缓椎间盘退变。

Abstract

Objective

To investigate the possible mechanism by which

Zhiqiao Gancao Decoction

（枳壳甘草汤， ZGD） delays intervertebral disc degeneration （IDD） based on the Hippo-yes-associated protein （YAP）/transcriptional co-activator with PDZ-binding motif （TAZ） signaling pathway.

Methods

A total of 50 SD rats were randomly divided into sham surgery group， model group， low-dose ZGD group， high-dose ZGD group， and high-dose ZGD + inhibitor group， with 10 rats in each group. In the sham surgery group， the rats were pierced in the skin and muscle at the Co6/7/8 segments of the tail with a 21G needle （depth approximately 2 mm） without damaging the intervertebral disc. In the other groups， rats were injected with a 21G needle at the Co6/7/8 segments of the tail to establish an IDD model by piercing the tail intervertebral disc 5 mm. One week after modeling， rats in the low-dose and high-dose ZGD groups were given 6.24 and 12.24 g/（kg·d） of the decoction via gastric gavage， respectively. The high-dose ZGD + inhibitor group was given 12.24 g/（kg·d） of the decoction and an intraperitoneal injection of YAP/TAZ inhibitor Verteporfin 10 mg/kg. The sham surgery and model groups were given 5 ml/（kg·d） of normal saline via gavage. The gavage was given once a day， and the intraperitoneal injection was given every other day. After 4 weeks of continuous intervention， the pathological changes of the tail intervertebral discs were observed using HE staining， Oil Red O-Green staining， and Toluidine Blue staining. Immunohistochemistry was used to detect the expression of aggrecan and MMP3 in the nucleus pulposus. TUNEL fluorescence staining was performed to detect apoptosis in the nucleus pulposus， and the apoptosis rate was calculated. Western blot was used to detect the Hippo-YAP/TAZ signaling pathway， including YAP， phosphorylated YAP （p-YAP）， phosphorylated MST1/2 （p-MST1/2）， phosphorylated TAZ （p-TAZ） and apoptosis-related proteins， such as Cleaved Caspase 3， P53， Bcl-2 and Bax.

Results

Compared with sham surgery group， the rats in the model group showed significant degenerative changes in the intervertebral disc. The levels of aggrecan， Bcl-2， and YAP proteins in the nucleus pulposus decreased， while the levels of p-MST1/2， p-YAP， p-TAZ， P53， Bax， Cleaved Caspase 3， MMP3 proteins， and the apoptosis rate increased （

0.01）. Compared with the model group， the drug intervention groups showed partial recovery in intervertebral disc degeneration. The levels of aggrecan， Bcl-2， and YAP proteins increased， while the levels of p-MST1/2， p-YAP， p-TAZ， P53， Bax， Cleaved Caspase 3， MMP3 proteins， and the apoptosis rate decreased （

＜0.05 or

＜0.01）. The high-dose ZGD group showed more significant recovery in intervertebral disc degeneration compared to the low-dose ZGD group， with a decrease in the levels of p-MST1/2， p-YAP， p-TAZ， P53， Bax， Cleaved Caspase 3， MMP3 proteins， and apoptosis rate， and an increase in the levels of aggrecan， Bcl-2， and YAP proteins （

＜0.05 or

＜0.01）. Compared with the high-dose ZGD group， the high-dose ZGD + inhibitor group showed a reduced recovery in intervertebral disc degeneration， with an increase in the levels of p-MST1/2， p-YAP， p-TAZ， P53， Bax， Cleaved Caspase 3， MMP3 proteins， and apoptosis rate， and a decrease in the levels of aggrecan， Bcl-2， and YAP proteins （

＜0.05 or

＜0.01）.

Conclusion

ZGD may delay intervertebral disc degeneration by inhibiting the phosphorylation of YAP in the nucleus pulposus， maintaining the function of the Hippo-YAP/TAZ signaling pathway， and reducing apoptosis of nucleus pulposus cells.

关键词

Keywords

references

MOLINOS M ， FIORDALISI MF ， CALDEIRA J ， et al . Alterations of bovine nucleus pulposus cells with aging ［J］. Aging Cell ， 2023 ， 22 （ 8 ）： e13873 .

吕圭源，陈素红，王婷 . 面向应用是中药研究的核心［J］. 世界科学技术-中医药现代化， 2018 ， 20 （ 4 ）： 479 - 487 .

龚正丰 . 枳壳甘草汤［J］. 江苏中医药， 2011 ， 43 （ 6 ）： 14 .

陈紫轩，沈晓峰，徐波，等 . 枳壳甘草汤结合腰椎斜扳手法治疗慢性非特异性腰痛的临床观察［J］. 中医药临床杂志， 2024 ， 36 （ 2 ）： 329 - 333 .

刘昱江，汪松林，蔡学峰，等 . 基于Pfirrmann分级观察枳壳甘草汤治疗腰椎间盘突出症的临床疗效［J］. 浙江中医药大学学报， 2023 ， 47 （ 6 ）： 651 - 658 .

HUANG Z ， CAI X ， SHEN X ， et al . Identification and experimental verification of immune-related hub genes in intervertebral disc degeneration ［J］. Heliyon ， 2024 ， 10 （ 14 ）： e34530 .

徐波，刘昱江，汪松林，等 . 枳壳甘草汤抑制巨噬细胞M1极化延缓椎间盘退变的实验研究［J］. 环球中医药， 2023 ， 16 （ 12 ）： 2421 - 2426 .

VANYAI HK ， PRIN F ， GUILLERMIN O ， et al . Control of skeletal morphogenesis by the Hippo-YAP/TAZ pathway ［J］. Development ， 2020 ， 147 （ 21 ）： dev187187 .

HAN J ， ZHANG J ， ZHANG X ， et al . Emerging role and function of Hippo-YAP/TAZ signaling pathway in musculoskeletal disorders ［J］. Stem Cell Res Ther ， 2024 ， 15 （ 1 ）： 386 .

YU C ， LI J ， KUANG W ， et al . PRDM1 promotes nucleus pulposus cell pyroptosis leading to intervertebral disc degeneration via activating CASP1 transcription ［J］. Cell Biol Toxicol ， 2024 ， 40 （ 1 ）： 89 .

颜昊，贾永伟，翟伟峰 . 椎间盘退变动物模型的研究进展［J］. 中国实验动物学报， 2021 ， 29 （ 3 ）： 387 - 392 .

LOTZ JC . Animal models of intervertebral disc degeneration： lessons learned ［J］. Spine （Phila Pa 1976）， 2004 ， 29 （ 23 ）： 2742 - 2750 .

魏伟，吴希美，李元建 . 药理实验方法学［M］. 北京：人民卫生出版社， 2010 ： 69 - 72 .

霍雨佳，阳远舟，邹成丽，等 . 汉黄芩素调节Hippo/YAP信号通路对慢性心力衰竭大鼠心肌纤维化的影响［J］. 中西医结合心脑血管病杂志， 2024 ， 22 （ 15 ）： 2740 - 2745 .

高兴强，窦晓静，郭燕春，等 . YAP抑制药Verteporfin增强肝细胞癌导管动脉化疗栓塞术治疗大鼠的疗效［J］. 中国临床药理学杂志， 2023 ， 39 （ 20 ）： 2970 - 2974 .

XIA Q ， ZHAO Y ， DONG H ， et al . Progress in the study of molecular mechanisms of intervertebral disc degeneration ［J］. Biomed Pharmacother ， 2024 ， 174 ： 116593 . doi： 10.1016/j.biopha.2024.116593 http://dx.doi.org/10.1016/j.biopha.2024.116593 .

XIN J ， WANG Y ， ZHENG Z ， et al . Treatment of intervertebral disc degeneration ［J］. Orthop Surg ， 2022 ， 14 （ 7 ）： 1271 - 1280 .

ZOU X ， ZHANG X ， HAN S ， et al . Pathogenesis and therapeutic implications of matrix metalloproteinases in intervertebral disc degeneration： a comprehensive review ［J］. Biochimie ， 2023 ， 214 （ Pt B ）： 27 - 48 .

YANG YH ， GU XP ， HU H ， et al . Ginsenoside Rg1 inhibits nucleus pulposus cell apoptosis， inflammation and extracellular matrix degradation via the YAP1/TAZ pathway in rats with intervertebral disc degeneration ［J］. J Orthop Surg Res ， 2022 ， 17 （ 1 ）： 555 .

EDGAR BA . From cell structure to transcription： hippo forges a new path ［J］. Cell ， 2006 ， 124 （ 2 ）： 267 - 273 .

LIANG T ， QIU J ， LI S ， et al . Inverse agonist of retinoid-related orphan receptor-alpha prevents apoptosis and degeneration in nucleus pulposus cells via upregulation of YAP ［J］. Mediators Inflamm ， 2021 ， 2021 ： 9954909 . doi： 10.1155/2021/9954909 http://dx.doi.org/10.1155/2021/9954909 .

WANG Z ， CUI M ， QU Y ， et al . Hypoxia protects rat bone marrow mesenchymal stem cells against compression-induced apoptosis in the degenerative disc microenvironment through activation of the HIF-1alpha/YAP signaling pathway ［J］. Stem Cells Dev ， 2020 ， 29 （ 20 ）： 1309 - 1319 .

SUDO H ， MINAMI A . Regulation of apoptosis in nucleus pulposus cells by optimized exogenous Bcl-2 overexpression ［J］. J Orthop Res ， 2010 ， 28 （ 12 ）： 1608 - 1613 .

DING F ， SHAO ZW ， XIONG LM . Cell death in intervertebral disc degeneration ［J］. Apoptosis ， 2013 ， 18 （ 7 ）： 777 - 785 .

张文捷，张勇，史明，等 . 淫羊藿苷调控髓核来源间充质干细胞凋亡修复椎间盘退变［J］. 中国组织工程研究， 2023 ， 27 （ 24 ）： 3803 - 3809 .

HUANG Z ， SHEN X ， CHEN H ， et al . Zhiqiao Gancao decoction regulated JAK2/STAT3/ macrophage M1 polarization to ameliorate intervertebral disc degeneration ［J］. Heliyon ， 2024 ， 10 （ 15 ）： e34715 .

HUANG Z ， LU B ， ZHANG X ， et al . Zhiqiao Gancao decoction ameliorates hyperalgesia in lumbar disc herniation via the CCL2/CCR2 signaling pathway ［J］. Drug Des Devel Ther ， 2023 ， 17 ： 2239 - 2257 . doi： 10.2147/DDDT.S415127 http://dx.doi.org/10.2147/DDDT.S415127 .

查看原文

Views

下载量

CSCD

Alert me when the article has been cited

提交

Tools

Publicity Resources

Effects of Huatan Sanjie Formula （化痰散结方） on Tumor Tissue Stiffness and the Integrin β1/FAK/YAP Mechanotransduction Signaling Pathway in Triple Negative Breast Cancera Model Mice

基于NLRP3/Caspase-1/IL-1β信号通路探讨通窍明目Ⅳ号方含药血清对L-谷氨酸诱导R28细胞损伤的影响

Effects of Bushen Tiaojing Formula （补肾调经方） on Blastocyst Cell Apoptosis and Endoplasmic Reticulum Stress in Repeated Controlled Ovarian Stimulation Model Mice

Effects of Jianpi Yishen Huatan Formula （健脾益肾化痰方）-Medicated Serum on Apoptosis，Migration and the LncRNA ALAL-1/USP4/HDAC2 Pathway in Human Lung Squamous Carcinoma Cells

The Effect of Ziyin Mingmu Formula （滋阴明目方） on Retinal Tissue Apoptosis and Endoplasmic Reticulum Stress in Retinitis Pigmentosa Model Mice

Related Author

ZHAO Xiangyu

LIU Jingyang

ZHANG Minpu

WANG Xue

SUN Changgang

GAO Qing

SUN He

WANG Liyuan

Related Institution

School of Traditional Chinese Medicine，Shandong Second Medical University

School of Traditional Chinese Medicine，Macau University of Science and Technology

Cancer Diagnosis and Treatment Center，Weifang Hospital of Traditional Chinese Medicine

School of Life Sciences，Zhejiang Chinese Medical University

Heilongjiang University of Chinese Medicine

Address：No.16, Nanxiao street, Dongzhimen, Dongcheng District, Beijing Postal code：100700
Tel：010-64035632 Email：jtcmcn@188.com
Technical support is provided by Beijing Founder electronics co., LTD 京ICP备09064830号-19 京公网安备11010102007558号
It is recommended to read the content of this site in Chrome&IE9+. Please switch to extreme mode in browser 360.
Cookies We use cookies to help provide and enhance our service and tailor content. By continuing, you agree to the use of cookies.

⁰

The Effect of Zhiqiao Gancao Decoction （枳壳甘草汤） on Intervertebral Disc Nucleus Pulposus Cell Apoptosis and the Hippo-YAP/TAZ Signaling Pathway in Tail Intervertebral Disc Degeneration Model Rats

DOI：10.13288/j.11-2166/r.2025.05.012

摘要

Abstract

关键词

Keywords

references