中国临床药理学杂志

苦参酮通过Ctsk调控TLR9信号通路拮抗破骨细胞分化的作用机制研究

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何欢 ^1a^,² , 方翰林 ^1a , 陈炜 ^1a^,² , 徐荣 ^1a^,² , 吴双双 ^1a^,² , 刘文锴 ^1b , 平静 ^1a^,²

中国临床药理学杂志 | 临床与基础桥接研究 2026,42(4): 516-522

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中国临床药理学杂志 |临床与基础桥接研究 2026 , 42 (4) : 516 -522

苦参酮通过Ctsk调控TLR9信号通路拮抗破骨细胞分化的作用机制研究

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何欢^1a^,², 方翰林^1a, 陈炜^1a^,², 徐荣^1a^,², 吴双双^1a^,², 刘文锴^1b, 平静^1a^,²

作者信息

^1a.抚州医药学院药学院，江西抚州 344000

^1b.抚州医药学院临床医学专业，江西抚州 344000

^2.江西省教育厅AI创新药物重点实验室，江西抚州 344000

通讯作者:

平静，副教授 MP: 13507049200 E-mail: hehuan_0118@163.com

作者简介:

何欢（1986-），女，副教授，主要从事内分泌药理学的相关工作和研究

Study of mechanism of kurarinone in antagonizing RANKL-induced osteoclast differentiation through inhibiting Ctsk-regulated TLR9 signaling pathway

Huan HE^1a^,², Han-lin FANG^1a, Wei CHEN^1a^,², Rong XU^1a^,², Shuang-shuang WU^1a^,², Wen-kai LIU^1b, Jing PING^1a^,²

Affiliations

^1.Fuzhou Medical University, School of Pharmacy, Clinical Medicine Program, Fuzhou 344000, Jiangxi Province, China

^2.Jiangxi Provincial Department of Education AI Innovation Drug Key Laboratory, Fuzhou 344000, Jiangxi Province, China

出版时间: 2026-02-28 doi: 10.13699/j.cnki.1001-6821.2026.04.010

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摘要

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目的

探讨苦参酮通过组织蛋白酶K（Ctsk）调控Toll样受体9（TLR9）信号通路对骨髓巨噬细胞（BMDM）细胞的作用及其机制。

方法

BMDM细胞分为对照组、模型组[核因子κB受体活化因子配体（RANKL）50 ng·mL^-1]、苦参酮组（RANKL 50 ng·mL^-1，苦参酮10 μmol·L^-1）、苦参酮+oe-NC组（感染oe-NC，RANKL 50 ng·mL^-1，苦参酮10 μmol·L^-1）、苦参酮+oe-Ctsk组（感染oe-Ctsk，RANKL 50 ng·mL^-1，苦参酮10 μmol·L^-1）。用实时荧光定量聚合酶链反应（qRT-PCR）法检测Ctsk mRNA相对表达水平；用蛋白免疫印迹法检测抗酒石酸酸性磷酸酶（TRAP）、TLR9等信号通路相关蛋白表达水平；用免疫荧光法检测破骨细胞分化相关蛋白表达水平；用酶联免疫吸附试验（ELISA）法检测炎症因子水平；用体外骨吸收试验检测骨吸收情况。

结果

对照组、模型组、苦参酮组、苦参酮+oe-NC组和苦参酮+oe-Ctsk组Ctsk mRNA相对表达水平分别为1.00±0.15、2.07±0.28、1.31±0.18、1.34±0.14和1.79±0.27，TRAP蛋白相对表达水平分别为1.00±0.16、2.31±0.44、1.43±0.17、1.38±0.22和1.91±0.30，基质金属蛋白酶9（MMP-9）蛋白相对荧光水平分别为1.00±0.15、2.18±0.37、1.33±0.19、1.28±0.17和1.62±0.31，活化T细胞核因子1（NFATc1）蛋白相对荧光水平分别为1.00±0.13、1.87±0.39、1.13±0.21、1.18±0.19和1.38±0.22，整合素β1（Itgb1）蛋白相对荧光水平分别为1.00±0.19、1.49±0.25、1.19±0.22、1.14±0.17和1.32±0.18，白细胞介素-1β（IL-1β）水平分别为（15.28±2.05）、（44.68±7.93）、（31.40±6.08）、（33.75±5.76）和（39.62±5.52）pg·mL^-1，白细胞介素-18（IL-18）水平分别为（122.45±22.39）、（317.56±51.51）、（191.72±33.34）、（185.93±32.19）和（217.89±33.58）pg·mL^-1，肿瘤坏死因子-α（TNF-α）水平分别为（81.36±12.27）、（216.93±34.39）、（130.89±16.18）、（125.76±15.15）和（162.94±26.59）pg·mL^-1，骨吸收陷窝数分别为每片（35.26±6.35）、（84.72±16.10）、（50.38±9.07）、（47.85±9.09）和（61.63±10.89），骨吸收陷窝相对面积分别为（0.26±0.05）%、（1.38±0.24）%、（1.04±0.18）%、（1.06±0.16）%和（1.27±0.19）%，TLR9蛋白相对表达水平分别为1.00±0.15、2.05±0.31、1.32±0.16、1.27±0.15和1.53±0.24，髓分化因子88（MyD88）蛋白相对表达水平分别为1.00±0.13、2.18±0.33、1.42±0.18、1.36±0.17和1.62±0.26，p-p65/t-p65分别是1.00±0.11、2.22±0.34、1.40±0.19、1.39±0.19、1.76±0.27。模型组的上述指标与对照组比较，苦参酮组的上述指标与模型组比较，苦参酮+oe-Ctsk组的上述指标与苦参酮组比较，在统计学上差异均有统计学意义（P<0.05，P<0.01，P<0.001）。

结论

苦参酮能通过抑制Ctsk，抑制RANKL诱导的破骨细胞分化、炎症因子分泌及骨吸收功能，可能是通过抑制TLR9通路实现的。

关键词

苦参酮 / 组织蛋白酶K / Toll样受体9 / 破骨细胞分化 / 骨质疏松

Abstract

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Objective

To investigate the effect and mechanism of kurarinone on bone marrow-derived macrophages (BMDM) cells through regulating toll-like receptor 9 (TLR9) signaling pathway via cathepsin K (Ctsk).

Methods

BMDM cells were divided into control group, model group [receptor activator of nuclear factor-κB ligand (RANKL) 50 ng·mL^-1], kurarinone group (RANKL 50 ng·mL^-1, kurarinone 10 μmol·L^-1), kurarinone+oe-NC group (infected oe-NC, RANKL 50 ng·mL^-1, kurarinone 10 μmol·L^-1), and kurarinone+oe-Ctsk group (infected oe-Ctsk, RANKL 50 ng·mL^-1, kurarinone 10 μmol·L^-1). Quantitative real time polymerase chain reaction (qRT-PCR) was used to detect the relative expression level of Ctsk mRNA; Western blot was used to detect the expression levels of tartrate-resistant acid phosphatase (TRAP) and TLR9 signaling pathway-related proteins; immunofluorescence was used to detect the expression levels of osteoclast differentiation-related proteins; enzyme-linked immunosorbent assay (ELISA) was used to detect the levels of inflammatory factors; in vitro bone resorption assay was used to detect bone resorption.

Results

The relative expression levels of Ctsk mRNA in the control group, model group, kurarinone group, kurarinone+oe-NC group, and kurarinone+oe-Ctsk group were 1.00±0.15, 2.07±0.28, 1.31±0.18, 1.34±0.14 and 1.79±0.27, respectively; the relative expression levels of TRAP protein were 1.00±0.16, 2.31±0.44, 1.43±0.17, 1.38±0.22 and 1.91±0.30, respectively; the relative fluorescence levels of matrix metalloproteinase-9 (MMP-9) protein were 1.00±0.15, 2.18±0.37, 1.33±0.19, 1.28±0.17 and 1.62±0.31, respectively; the relative fluorescence levels of nuclear factor of activated T-cells 1 (NFATc1) protein were 1.00±0.13, 1.87±0.39, 1.13±0.21, 1.18±0.19 and 1.38±0.22, respectively; the relative fluorescence levels of integrin beta-1 (Itgb1) protein were 1.00±0.19, 1.49±0.25, 1.19±0.22, 1.14±0.17 and 1.32±0.18, respectively; the levels of interleukin-1β (IL-1β) were (15.28±2.05), (44.68±7.93), (31.40±6.08), (33.75±5.76) and (39.62±5.52) pg·mL^-1, respectively; the levels of interleukin-18 (IL-18) were (122.45±22.39), (317.56±51.51), (191.72±33.34), (185.93±32.19) and (217.89±33.58) pg·mL^-1, respectively; the levels of tumor necrosis factor-α (TNF-α) were (81.36±12.27), (216.93±34.39), (130.89±16.18), (125.76±15.15) and (162.94±26.59) pg·mL^-1, respectively; the number of bone resorption pits were (35.26±6.35), (84.72±16.10), (50.38±9.07), (47.85±9.09) and (61.63±10.89) per piece, respectively; the relative area of bone resorption pits were (0.26±0.05)%, (1.38±0.24)%, (1.04±0.18)%, (1.06±0.16)% and (1.27±0.19)%, respectively; the relative expression levels of TLR9 protein were 1.00±0.15, 2.05±0.31, 1.32±0.16, 1.27±0.15 and 1.53±0.24, respectively; the relative expression levels of myeloid differentiation factor 88 (MyD88) protein were 1.00±0.13, 2.18±0.33, 1.42±0.18, 1.36±0.17 and 1.62±0.26, respectively; p-p65/t-p65 were 1.00±0.11, 2.22±0.34, 1.40±0.19, 1.39±0.19, 1.76±0.27, respectively. Compared model group with control group, compared kurarinone group with the model group, compared the kurarinone+oe-Ctsk group with the kurarinone group, the differences of above indicators were all statistically significant (P<0.05, P<0.01, P<0.001).

Conclusion

Kurarinone can inhibit RANKL-induced osteoclast differentiation, inflammatory factor secretion, and bone resorption function by suppressing Ctsk, which may be achieved through inhibiting the TLR9 pathway.

Key words

kurarinone / cathepsin K / toll-like receptor 9 / osteoclast differentiation / osteoporosis

引用本文

何欢, 方翰林, 陈炜, 徐荣, 吴双双, 刘文锴, 平静. 苦参酮通过Ctsk调控TLR9信号通路拮抗破骨细胞分化的作用机制研究. 中国临床药理学杂志, 2026 , 42 (4) : 516 -522 . DOI: 10.13699/j.cnki.1001-6821.2026.04.010

Huan HE, Han-lin FANG, Wei CHEN, Rong XU, Shuang-shuang WU, Wen-kai LIU, Jing PING. Study of mechanism of kurarinone in antagonizing RANKL-induced osteoclast differentiation through inhibiting Ctsk-regulated TLR9 signaling pathway[J]. Chinese Journal of Clinical Pharmacology, 2026 , 42 (4) : 516 -522 . DOI: 10.13699/j.cnki.1001-6821.2026.04.010

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2026年第42卷第4期

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doi: 10.13699/j.cnki.1001-6821.2026.04.010

接收时间：2026-01-30
首发时间：2026-08-06
出版时间：2026-02-28

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收稿日期：2026-01-30

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^1a.抚州医药学院药学院，江西抚州 344000

^1b.抚州医药学院临床医学专业，江西抚州 344000

^2.江西省教育厅AI创新药物重点实验室，江西抚州 344000

通讯作者:

平静，副教授 MP: 13507049200 E-mail: hehuan_0118@163.com

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2种不同金属材料的力学参数

科 Family	属数 Number of genus	种数 Number of species	占总种数比例 Percentage of total species (%)	属 Genus	种数 Number of species	占总种数比例 Percentage of total species (%)
鹅膏菌科Amanitaceae	2	11	5.26	鹅膏菌属 Amanita	10	4.78
小菇科 Mycenaceae	2	12	5.74	丝盖伞属 Inocybe	5	2.39
多孔菌科 Polyporaceae	8	14	6.70	蜡蘑属 Laccaria	5	2.39
红菇科 Russulaceae	3	23	11.00	小皮伞属 Marasmius	6	2.87
				小菇属 Mycena	11	5.26
				光柄菇属 Pluteus	5	2.39
				红菇属 Russula	17	8.13
				栓菌属 Trametes	5	2.39

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