Miao medicine Sidaxue regulates macrophage polarization and alleviates synovial inflammation in collagen-induced arthritis rats by inhibiting HIF-1α/PDK1-mediated glycolytic reprogramming

doi:10.12122/j.issn.1673-4254.2026.09.08

Abstract

Abstract:

Objective To investigate the mechanism of Miao medicine prescription Sidaxue (SX) for inhibiting glycolysis in the synovial tissue and promoting macrophage polarization in rats with collagen-induced arthritis (CIA). Methods Thirty-two SD rats were randomized into normal control group, CIA model group, tripterygium glycosides (40 mg/kg; positive control) treatment group, and SX (40 g/kg) treatment group (n=8). Except for those in the normal control group, all the rats were subjected to CIA modeling and treated with saline, tripterygium glycosides or SX by gavage for 21 consecutive days. Paw swelling, arthritis index, synovial pathology, inflammatory factors, glycolysis-related indicators and gene expressions were detected. UHPLC-MS was used to screen the components of SX in the blood of SX-treated rats. The target genes of the active components of SX and rheumatoid arthritis (RA) were predicted using bioinformatics analysis, and the hub genes were identified using machine learning algorithms. The binding sites between HIF-1α and PDK1 were predicted using AlphaFold3, and the interactions between SX components and their targets were simulated using molecular docking. Results SX treatment significantly alleviated joint swelling, lowered arthritis scores, improved synovial lesions, and reduced serum levels of TNF-α, IL-1β and IL-6 in CIA rats, resulting in also inhibition of macrophage polarization and down-regulation of HIF-1α, PDK1 and HK2 expressions in the synovial tissues. Twenty-nine components of SX were detected in the blood of SX-treated rats, and 9 key target genes were identified. AlphaFold3 predicted a strong interaction between HIF-1α and PDK1, and their virtual knockout was shown to disturb the inflammatory pathways in the macrophages. Molecular docking studies suggested stable docking between the active components of SX and their key targets. Conclusion SX alleviates joint inflammation in CIA rats by inhibiting the HIF-1α/PDK1 signaling pathway, correcting glycolytic reprogramming, and promoting M2 polarization of macrophages.

Key words: rheumatoid arthritis, Sidaxue, HIF-1α/PDK1 pathway, glycolytic reprogramming, macrophage polarization

Wanting SHEN, Yizhe CHEN, Xinyu FU, Jiatao YUAN, Jun ZHOU, Li HUANG, Ning WU. Miao medicine Sidaxue regulates macrophage polarization and alleviates synovial inflammation in collagen-induced arthritis rats by inhibiting HIF-1α/PDK1-mediated glycolytic reprogramming[J]. Journal of Southern Medical University, 2026, 46(9): 2078-2090.

Figures/Tables 12

References 40

[1]	Di Matteo A, Bathon JM, Emery P. Rheumatoid arthritis[J]. Lancet, 2023, 402(10416): 2019-33. doi：10.1016/s0140-6736(23)01525-8
[2]	Parab A, Bhatt LK. T-cell metabolism in rheumatoid arthritis: focus on mitochondrial and lysosomal dysfunction[J]. Immunopharmacol Immunotoxicol, 2024, 46(3): 378-84. doi：10.1080/08923973.2024.2330645
[3]	Xu J, Feng ZT, Chen SX, et al. Taxol alleviates collagen-induced arthritis in mice by inhibiting the formation of microvessels[J]. Clin Rheumatol, 2019, 38(1): 19-27. doi：10.1007/s10067-017-3646-1
[4]	Touhidul Islam SM, Won J, Khan M, et al. Hypoxia-inducible factor-1 drives divergent immunomodulatory functions in the pathogenesis of autoimmune diseases[J]. Immunology, 2021, 164(1): 31-42. doi：10.1111/imm.13335
[5]	Damerau A, Kirchner M, Pfeiffenberger M, et al. Metabolic reprogramming of synovial fibroblasts in osteoarthritis by inhibition of pathologically overexpressed pyruvate dehydrogenase kinases[J]. Metab Eng, 2022, 72: 116-32. doi：10.1016/j.ymben.2022.03.006
[6]	Škorja Milić N, Dolinar K, Miš K, et al. Suppression of pyruvate dehydrogenase kinase by dichloroacetate in cancer and skeletal muscle cells is isoform specific and partially independent of HIF-1α[J]. Int J Mol Sci, 2021, 22(16): 8610. doi：10.3390/ijms22168610
[7]	雷海桃, 王金海, 高韵奇, 等. 己糖激酶2在类风湿关节炎发病机制中的研究进展[J]. 中国骨质疏松杂志, 2025, 31(1): 90-5.
[8]	Chen JX, Li GR, Sun DJ, et al. Research progress of hexokinase 2 in inflammatory-related diseases and its inhibitors[J]. Eur J Med Chem, 2024, 264: 115986. doi：10.1016/j.ejmech.2023.115986
[9]	Su PF, Li O, Ke K, et al. Targeting tumor-associated macrophages: Critical players in tumor progression and therapeutic strategies (Review)[J]. Int J Oncol, 2024, 64(6). doi：10.3892/ijo.2024.5648
[10]	Cersosimo F, Lonardi S, Bernardini G, et al. Tumor-associated macrophages in osteosarcoma: from mechanisms to therapy[J]. Int J Mol Sci, 2020, 21(15): 5207. doi：10.3390/ijms21155207
[11]	Peng Y, Zhou MX, Yang H, et al. Regulatory mechanism of M1/M2 macrophage polarization in the development of autoimmune diseases[J]. Mediat Inflamm, 2023, 2023: 8821610. doi：10.1155/2023/8821610
[12]	Kzhyshkowska J, Shen JX, Larionova I. Targeting of TAMs: can we be more clever than cancer cells?[J]. Cell Mol Immunol, 2024, 21(12): 1376-409. doi：10.1038/s41423-024-01232-z
[13]	Siouti E, Andreakos E. The many facets of macrophages in rheumatoid arthritis[J]. Biochem Pharmacol, 2019, 165: 152-69. doi：10.1016/j.bcp.2019.03.029
[14]	余芸, 蔡伟伟, 周静, 等. 代谢重编程调控巨噬细胞极化及其在类风湿关节炎中的作用[J]. 药学学报, 2020, 55(9): 2027-34. doi：10.16438/j.0513-4870.2020-0386
[15]	Kumar V. Inflammation research sails through the sea of im-munology to reach immunometabolism[J]. Int Immunopharmacol, 2019, 73: 128-45. doi：10.1016/j.intimp.2019.05.002
[16]	欧慧萍, 吴趋荟, 沈枭, 等. 中医药防治类风湿关节炎的作用机制研究进展[J]. 吉林中医药, 2021, 41(12): 1685-8.
[17]	袁桃花, 黄宇键, 李佳豪, 等. 贵州苗药方“四大血”对CIA大鼠滑膜血管增生的作用机制[J]. 西安交通大学学报: 医学版, 2019, 40(5): 824-9, 843. doi：10.7652/jdyxb201905031
[18]	吴宁, 曾佳, 王时敏, 等. 苗药四大血对大鼠佐剂性关节炎作用的研究[J]. 中国实验方剂学杂志, 2014, 20(2): 120-3. doi：10.11653/syfj2014020120
[19]	梁玺, 覃淇, 张艺馨, 等. 苗药验方“四大血”对类风湿性关节炎大鼠关节炎症的作用及机制[J]. 贵州医科大学学报, 2023, 48(7): 759-66.
[20]	吴宁, 袁桃花, 姬进忠, 等. 苗药验方四大血对人类风湿性关节炎成纤维样滑膜细胞凋亡和焦亡的影响[J]. 南方医科大学学报, 2021, 41(10): 1473-83. doi：10.12122/j.issn.1673-4254.2021.10.05
[21]	Wu N, Yuan TH, Yin ZX, et al. Network pharmacology and molecular docking study of the Chinese Miao medicine sidaxue in the treatment of rheumatoid arthritis[J]. Drug Des Devel Ther, 2022, 16: 435-66. doi：10.2147/dddt.s330947
[22]	马麦丽娜, 李喜媛, 马晓飞, 等. 中医药调控M1/M2巨噬细胞极化治疗类风湿关节炎研究进展[J]. 中国医药导刊, 2025, 27(8): 780-5.
[23]	胡白飞, 高倩茹, 胡海明, 等. 中药物质基础研究进展：从化学物质组学到肠道转化物质组学[J]. 中国生物工程杂志, 2026, 46(4): 142-54.
[24]	陈旺宇, 节笑笑, 陈甜甜, 等. 中药血清药物化学研究现状[J]. 中国医药科学, 2023, 13(4): 26-9, 86.
[25]	彭鹏, 李佳欣, 杨新月, 等. 基于UPLC-Q-Exactive Orbitrap MS分析泻白散在正常和慢性支气管炎小鼠体内的入血成分及其代谢差异[J]. 中国实验方剂学杂志, 2026, 32(1): 219-27.
[26]	郭帅, 褚小磊, 沈桐, 等. 两种佐剂制备CIA大鼠模型的效果比较[J]. 蚌埠医学院学报, 2022, 47(3): 281-4.
[27]	王琎, 尹葳, 蔡雪阳, 等. 基于SOCS1/STAT3信号通路探究红禾麻乙酸乙酯部位对Ⅱ型胶原诱导关节炎大鼠的影响[J]. 中成药, 2023, 45(7): 2364-7.
[28]	王远迎, 戴兴月, 陈春彦, 等. 苗药验方四大血可减少关节炎大鼠滑膜组织的坏死性凋亡和血管新生: 基于抑制RIPK1/RIPK3/MLKL通路[J]. 南方医科大学学报, 2022, 42(12): 1774-82. doi：10.12122/j.issn.1673-4254.2022.12.04
[29]	Li XJ, Li FX, Jing WY, et al. Effect of heat-reinforcing needling on hypoxia-inducible factor 1α and glycolysis activity in rabbits with cold syndrome of rheumatoid arthritis[J]. Acupunct Res, 2024, 49(6): 585-93. doi：10.13702/j.1000-0607.20230487
[30]	Wu TZ, Hu EQ, Xu SB, et al. clusterProfiler 4.0: a universal enrichment tool for interpreting omics data[J]. Innovation, 2021, 2(3): 100141. doi：10.1016/j.xinn.2021.100141
[31]	Gu ZG, Eils R, Schlesner M. Complex heatmaps reveal patterns and correlations in multidimensional genomic data[J]. Bioinformatics, 2016, 32(18): 2847-9. doi：10.1093/bioinformatics/btw313
[32]	Osorio D, Zhong Y, Li GX, et al. scTenifoldKnk: an efficient virtual knockout tool for gene function predictions via single-cell gene regulatory network perturbation[J]. Patterns, 2022, 3(3): 100434. doi：10.1016/j.patter.2022.100434
[33]	Wang Q, Ren JH, Lin X, et al. Inflammatory stimulus-responsive polymersomes reprogramming glucose metabolism mitigates rheumatoid arthritis[J]. Biomaterials, 2025, 312: 122760. doi：10.1016/j.biomaterials.2024.122760
[34]	尹煜辉, 李琰, 齐保玉, 等. 基于有氧糖酵解探究补骨生髓方含药血清通过介导HIF-1α促进MC3T3-E1细胞成骨分化的作用[J].湖南中医药大学学报, 2025, 45(5): 818-24.
[35]	Jin ZX, Kho J, Dawson B, et al. Nitric oxide modulates bone anabolism through regulation of osteoblast glycolysis and differentiation[J]. J Clin Invest, 2021, 131(5): e138935. doi：10.1172/jci138935
[36]	Pei XY, Lu LJ, Huang ZQ, et al. Activation of M1 macrophages promotes diabetic kidney disease by modulating glycolysis via HIF-1α‑HK2 signaling pathway[J]. Diabetol Metab Syndr, 2025, 17(1): 362. doi：10.1186/s13098-025-01894-3
[37]	Tardito S, Martinelli G, Soldano S, et al. Macrophage M1/M2 polarization and rheumatoid arthritis: a systematic review[J]. Autoimmun Rev, 2019, 18(11): 102397. doi：10.1016/j.autrev.2019.102397
[38]	Ye L, Wen ZY, Li YQ, et al. Interleukin-10 attenuation of collagen-induced arthritis is associated with suppression of interleukin-17 and retinoid-related orphan receptor γt production in macrophages and repression of classically activated macrophages[J]. Arthritis Res Ther, 2014, 16(2): R96. doi：10.1186/ar4544
[39]	Muraille E, Leo O, Moser M. TH1/TH2 paradigm extended: macrophage polarization as an unappreciated pathogen-driven escape mechanism[J]? Front Immunol, 2014, 5: 603. doi：10.3389/fimmu.2014.00603
[40]	李淼, 李瑞, 杨晓芳, 等. 橙皮苷调控mTOR/HIF-1α通路对高糖诱导的肾小球系膜细胞自噬的影响[J]. 天津中医药, 2025, 42(1): 78-83.

Primer	Sequences (5' to 3')
HK2	Forward Reverse	GCTGCTGCTGAAGAAGATGG
HK2	Forward Reverse	CCAGAGGGAGTCGATGTTGA
HIF-1α	Forward Reverse	GCTGATTTGTGAGCCTTCTATG
HIF-1α	Forward Reverse	TCATCTGTAAGTGGGTCTCGT
PDK1	Forward Reverse	ATGGCGTCCGTGGAGAAGTA
PDK1	Forward Reverse	TCAGTCTTGGCGTAGTCGTT
GAPDH	Forward Reverse	GAAGCTGGTCATCAACGGGA
GAPDH	Forward Reverse	GGCGGAGATGATGACCCTTT

Primer	Sequences (5' to 3')
HK2	Forward Reverse	GCTGCTGCTGAAGAAGATGG
HK2	Forward Reverse	CCAGAGGGAGTCGATGTTGA
HIF-1α	Forward Reverse	GCTGATTTGTGAGCCTTCTATG
HIF-1α	Forward Reverse	TCATCTGTAAGTGGGTCTCGT
PDK1	Forward Reverse	ATGGCGTCCGTGGAGAAGTA
PDK1	Forward Reverse	TCAGTCTTGGCGTAGTCGTT
GAPDH	Forward Reverse	GAAGCTGGTCATCAACGGGA
GAPDH	Forward Reverse	GGCGGAGATGATGACCCTTT

Group	TNF-α	IL-1β	IL-6
Normal	30.81±4.56	12.14±1.85	32.79±4.26
Model	62.86±5.68**	54.52±4.51**	79.78±5.35**
GTW	39.82±5.88^#	32.36±3.97^#	42.65±6.31^#
SX	32.62±6.43^##	30.89±2.36^##	33.52±4.54^##

Group	TNF-α	IL-1β	IL-6
Normal	30.81±4.56	12.14±1.85	32.79±4.26
Model	62.86±5.68**	54.52±4.51**	79.78±5.35**
GTW	39.82±5.88^#	32.36±3.97^#	42.65±6.31^#
SX	32.62±6.43^##	30.89±2.36^##	33.52±4.54^##

Serial number	t (min)	Component	Lon species	Measured value (m/z)	Score
1	3.32	Fumaritine	[M+H]⁺	356.15	0.99
2	3.46	Heriguard	[M+H C₇H₁₂O₆]⁺	163.04	0.99
3	3.55	Procyanidin B2	[M+H]⁺	579.15	0.99
4	3.63	Magnoflorine	[M]⁺	342.17	0.93
5	4.26	3-O-Feruloylquinic acid	[M+H]⁺	369.12	0.99
6	4.31	Glu-Thr	[M+H]⁺	249.11	0.93
7	5.16	N-Malonyltryptophan	[M+H]⁺	291.10	0.83
8	5.43	7-hydroxy-4-(methoxymethyl)coumarin	[M+H]⁺	207.06	0.89
9	5.51	Brutieridin	[M+H C₁₈H₂₈O₁₃]⁺	303.09	0.92
10	5.90	4-Hydroxy-7-methyl-2-[2-[3,4,5-trihydroxy-6-(hydroxymethyl)oxan-2-yl] oxypropan-2-yl]-2,3-dihydrofuro[3,2-g]chromen-5-one	[M+H]⁺	439.16	0.97
11	7.15	Toddalolactone	[M+H]⁺	309.13	1.00
12	7.89	Visamminol	[M+H]⁺	277.11	0.96
13	8.48	Skimmianine	[M+H]⁺	260.09	0.95
14	9.34	Isopimpinellin	[M+H]⁺	247.06	1.00
15	10.01	Pimpinellin	[M+H]+	247.06	1.00
16	12.12	Phellopterin	[M+H]⁺	301.11	0.98
17	2.84	Neochlorogenic acid	[M-H]^-	353.09	0.97
18	3.36	Wikstrol A	[M-H C₆H₆O₃]-	417.10	0.73
19	3.95	Catechin	[M-H]^-	289.07	1.00
20	4.09	2-beta-D-Glucopyranosyloxy-4-methoxy-cis-cinnamic acid	[M-H-H₂O]^-	337.09	0.98
21	4.37	(-)-Lyoniresinol 9'-O-glucoside	[M-H]^-	581.22	0.95
22	4.79	N-Acetyl-L-phenylalanine	[M-H]^-	206.08	0.98
23	5.15	Asn-Asn	[M-H]^-	245.09	0.73
24	5.51	Hesperidin	[M-H]^-	609.18	1.00
25	6.32	Azelaic acid	[M-H]^-	187.10	1.00
26	6.60	Salicylic acid	[M-H]^-	137.02	1.00
27	8.35	1-O-[(2beta,3beta,5xi,9xi,18xi)-2,23-Dihydroxy-28,29-dioxo-3- (beta-D-xylopyranosyloxy)olean-12-en-28-yl]-beta-D-glucopyranose	[M-H]^-	825.43	0.99
28	10.76	Mocimycin	[M-H]^-	795.42	0.97
29	11.22	Medicagenic acid 3-O-beta-D-glucoside	[M-H]^-	663.37	0.97