Multifunctional Prussian blue nanoparticles loading with Xuetongsu for efficient rheumatoid arthritis therapy through targeting inflammatory macrophages and osteoclasts

Abstract

Abnormal activation of macrophages and osteoclasts (OCs) contributes significantly to rheumatoid arthritis (RA) development by secretion of numerous inflammatory factors. Notably, these cells exhibit significant upregulation of folate receptor proteins on their surfaces. Unfortunately, there is a current lack of safe and effective therapeutic drugs for RA. Xuetongsu (XTS), a triterpenoid compound extracted from Kadsura heteroclita Roxb Craib, has demonstrated the ability to significantly inhibit the proliferation of RA fibroblast-like synoviocytes (RAFLS). However, its clinical application is hampered by poor targeting and short half-life. To address these drawbacks, we previously developed a nano-drug system named HRPS nanoparticles (NPs), which effectively targets RAFLS and inhibits synovial hyperplasia. However, this system overlooked the essential role of OCs in RA-related bone destruction. Therefore, we designed a novel folate-modified biomimetic Prussian blue (PB)-XTS NP (FMPX NP) for the selective delivery of XTS into inflammatory macrophages and OCs. The NP exhibits an excellent photothermal effect when assisted by laser irradiation, facilitating targeted release of XTS within inflammatory macrophages and OCs. The synergistic anti-inflammatory and reactive oxygen species scavenging effects of PB NPs and XTS are mediated by the inhibition of the NF-κB signaling pathway in inflammatory macrophages and RANK/RANKL/NFATc1 signaling pathway in OCs. In vivo experiments showed that FMPX NPs extended the half-life of XTS by 2.32 times, decreased hind foot swelling from 12.10 ± 0.49 mm to 8.24 ± 0.09 mm in the model group, and prevented bone damage. In conclusion, this study introduces a novel dual-targeted nano-based therapy for RA joints and highlights its potential for biochemical photothermal triple therapy for RA. FMPX NPs inhibit arthritis-related inflammation and bone destruction through a dual-target strategy, providing new insights for targeted drug therapies in clinical RA treatment.

Keywords

Rheumatoid arthritis / Xuetongsu / Nano drug delivery system / Macrophage / Osteoclast

Conflicts of interest

The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper. The authors report no conflicts of interest. The authors alone are responsible for the content and writing of this article.

Acknowledgments

This work was financially supported by National Natural Science Foundation of China (82204766, 82074122, 82174078); Natural Science Foundation of Hunan province (2023JJ40490); Changjiang Scholars Program in Ministry Education, People's Republic of China (T2019133); Xiaohe Sci-Tech Talents Special Funding under Hunan Provincial Sci-Tech Talents Sponsorship Program (2023TJ-X71); Science and Technology Innovation Program of Hunan Province (2024RC3201); Scientific research project of Hunan Provincial Education Department (21B0394, 21A0239); Research Project of Hunan Administration of Traditional Chinese Medicine (B2023055); Changsha Outstanding Innovative Youth Training Program (kq2306021); Outstanding Youth Program of Hunan University of Chinese Medicine (202202); Postgraduate Innovation Project of Hunan University of Chinese Medicine (2024CX090); Open Foundation Project of Hunan International Joint Laboratory of Traditional Chinese Medicine (2022GJSYS02); Undergraduate Research and Innovation Foundation of Hunan University of Chinese Medicine (2023BKS097).

Supplementary materials

Supplementary material associated with this article can be found, in the online version, at doi:10.1016/j.ajps.2025.101037. The figures and tables with "S" before the serial number are included in the Supplementary material.

References

[1] Matteo AD, Bathon JM, Emery P. Rheumatoid arthritis. Nat Rev Dis Primer 2018; 4:18002.

[2] Finckh A, Gilbert B, Hodkinson B, Bae S-C, Thomas R, Deane KD, et al. Global epidemiology of rheumatoid arthritis. Nat Rev Rheumatol 2022; 18:591-602.

[3] Ray LB. RANKL in bone homeostasis. Science 2018; 362(42):1-43.

[4] Buckley CD, Ospelt C, Gay S, Midwood KS. Location, location, location: how the tissue microenvironment affects inflammation in RA. Nat Rev Rheumatol 2021; 17:195212.

[5] Huang R, Wang X, Zhou Y, Xiao Y. RANKL-induced M1 macrophages are involved in bone formation. Bone Res 2017; 5:17019.

[6] Bae S, Kim K, Kang K, Kim H, Lee M, Oh B, et al. RANKL-responsive epigenetic mechanism reprograms macrophages into bone-resorbing osteoclasts. Cell Mol Immunol 2022; 20:94-109.

[7] Jia N, Gao Y, Li M, Liang Y, Li Y, Lin Y, et al. Metabolic reprogramming of proinflammatory macrophages by target delivered roburic acid effectively ameliorates rheumatoid arthritis symptoms. Signal Transduct Target Ther 2023; 8(1):280.

[8] Yang Y, Guo L, Wang Z, Liu P, Liu X, Ding J, et al. Targeted silver nanoparticles for rheumatoid arthritis therapy via macrophage apoptosis and re-polarization. Biomaterials 2021; 264:120390.

[9] Burmester GR, Pope JE. Novel treatment strategies in rheumatoid arthritis. Lancet 2017; 389:2338-48.

[10] Wu H, He Y, Wu H, Zhou M, Xu Z, Xiong R, et al. Near-infrared fluorescence imaging-guided focused ultrasound-mediated therapy against rheumatoid arthritis by MTX-ICG-loaded iRGD-modified echogenic liposomes. Theranostics 2020; 10:10092-105.

[11] Heo R, You DG, Um W, Choi KY, Jeon S, Park J-S, et al. Dextran sulfate nanoparticles as a theranostic nanomedicine for rheumatoid arthritis. Biomaterials 2017; 131:15-26.

[12] Jeong M, Jung Y, Yoon J, Kang J, Lee SH, Back W, et al. Porous silicon-based nanomedicine for simultaneous management of joint inflammation and bone erosion in rheumatoid arthritis. ACS Nano 2022; 16:16118-32.

[13] Moltó A, Dougados M. Novel DMARD monotherapy in rheumatoid arthritis. The Lancet 2019; 393:2277-8.

[14] Harigai M, Honda S. Selectivity of janus kinase inhibitors in rheumatoid arthritis and other immune-mediated inflammatory diseases: is expectation the root of all headache? Drugs 2020; 80:1183-201.

[15] Schwartz DM, Kanno Y, Villarino A, Ward M, Gadina M, O'Shea JJ. JAK inhibition as a therapeutic strategy for immune and inflammatory diseases. Nat Rev Drug Discov 2017; 16:843-62.

[16] Aletaha D, Smolen JS. Diagnosis and management of rheumatoid arthritis: a review. JAMA 2018; 320:1360.

[17] Yu H, Lin Y, Zeng R, Li X, Zhang T, Tasneem S, et al. Analgesic and anti-inflammatory effects and molecular mechanisms of Kadsura heteroclita stems, an anti-arthritic Chinese Tujia ethnomedicinal herb. J Ethnopharmacol 2019;238: 111902.

[18] Shehla N, Li B, Cao L, Zhao J, Jian Y, Daniyal M, et al. Xuetonglactones A-F: highly oxidized lanostane and cycloartane triterpenoids from Kadsura heteroclita Roxb. Craib. Front Chem 2020; 7:935.

[19] Yu H, Zeng R, Lin Y, Li X, Tasneem S, Yang Z, et al. Kadsura heteroclita stem suppresses the onset and progression of adjuvant-induced arthritis in rats. Phytomedicine 2019; 58:152876.

[20] Liu R, Liu Q, Li B, Liu L, Cheng D, Cai X, et al. Pharmacokinetics, bioavailability, excretion, and metabolic analysis of Schisanlactone E, a bioactive ingredient from Kadsura heteroclita (Roxb) Craib, in rats by UHPLC-MS/MS and UHPLC-Q-Orbitrap HRMS. J Pharm Biomed Anal 2020; 177:112875.

[21] Zheng H, Li Y, Deng Y, Li H, Shen X, Lin H, et al. Xuetongsu attenuates bone destruction in collagen-induced arthritis mice by inhibiting osteoclast differentiation and promoting osteoclast apoptosis. Int J Biochem Cell Biol 2024; 169:106550.

[22] Wang Z, Long Y, Fan J, Xiao C, Tong C, Guo C, et al. Biosafety and biocompatibility assessment of prussian blue nanoparticles in vitro and in vivo. Nanomed 2020; 15:2655-70.

[23] Long Y, Fan J, Zhou N, Liang J, Xiao C, Tong C, et al. Biomimetic Prussian blue nanocomplexes for chemo-photothermal treatment of triple-negative breast cancer by enhancing ICD. Biomaterials 2023; 303:122369.

[24] Tong C, Zhong X, Yang Y, Liu X, Zhong G, Xiao C, et al. PB@PDA@Ag nanosystem for synergistically eradicating MRSA and accelerating diabetic wound healing assisted with laser irradiation. Biomaterials 2020; 243:119936.

[25] Gan J, Huang D, Che J, Zhao Y, Sun L. Biomimetic nanoparticles with cell-membrane camouflage for rheumatoid arthritis. Matter 2024; 7(3):794-825.

[26] Lyu J, Wang L, Bai X, Du X, Wei J, Wang J, et al. Treatment of rheumatoid arthritis by serum albumin nanoparticles coated with mannose to target neutrophils. ACS Appl Mater Interfaces 2021; 13:266-76.

[27] Yu H, Fan J, Shehla N, Qiu Y, Lin Y, Wang Z, et al. Biomimetic hybrid membrane-coated xuetongsu assisted with laser irradiation for efficient rheumatoid arthritis therapy. ACS Nano 2022; 16:502-21.

[28] Kumar P, Huo P, Liu B. Formulation strategies for folate-targeted liposomes and their biomedical applications. Pharmaceutics 2019; 11:381.

[29] Ma Y, Lu Z, Jia B, Shi Y, Dong J, Jiang S, et al. DNA origami as a nanomedicine for targeted rheumatoid arthritis therapy through reactive oxygen species and nitric oxide scavenging. ACS Nano 2022; 16:12520-31.

[30] Ni R, Song G, Fu X, Song R, Li L, Pu W, et al. Reactive oxygen species-responsive dexamethasone-loaded nanoparticles for targeted treatment of rheumatoid arthritis via suppressing the iRhom2/TNF- a /BAFF signaling pathway. Biomaterials 2020; 232:119730.

[31] Liu B, Wang W, Fan J, Long Y, Xiao F, Daniyal M, et al. RBC membrane camouflaged prussian blue nanoparticles for gamabutolin loading and combined chemo/photothermal therapy of breast cancer. Biomaterials 2019; 217:119301.

[32] Fan J, Qin Y, Xiao C, Yuan L, Long Y, Zhao Y, et al. Biomimetic PLGA-based nanocomplexes for improved tumor penetration to enhance chemo-photodynamic therapy against metastasis of TNBC. Mater Today Adv 2022; 16:100289.

[33] Xiao C, Sun Y, Fan J, Nguyen W, Chen S, Long Y, et al. Engineering cannabidiol synergistic carbon monoxide nanocomplexes to enhance cancer therapy via excessive autophagy. Acta Pharm Sin B 2023; 13:4591-606.

[34] Wang M, Jiang S, Hussain N, Zafar S, Xie Q, Huang F, et al. Anti-RAFLS triterpenoids and hepatoprotective lignans from the leaves of Tujia ethnomedicine Kadsura heteroclita (Xuetong). Front Chem 2022; 10:878811.

[35] Deng Y, Zheng H, Liu S, Deng Y, Chen Y, Liang L, et al. Effective extraction of Xuetongsu and its role in preventing RA synovial hyperplasia by targeting synovial cell migration and apoptosis. Sci Rep 2024; 14(1):23345.

[36] Li T, Guo R, Zong Q, Ling G. Application of molecular docking in elaborating molecular mechanisms and interactions of supramolecular cyclodextrin. Carbohydr Polym 2022; 276:118644.

[37] Lin Y, Yi O, Hu M, Hu S, Su Z, Liao J, et al. Multifunctional nanoparticles of sinomenine hydrochloride for treat-to-target therapy of rheumatoid arthritis via modulation of proinflammatory cytokines. J Controlled Release 2022; 348:42-56.

[38] Fan X, Xu M, Leung EL-H, Jun C, Yuan Z, Liu L. ROS - responsive berberine polymeric micelles effectively suppressed the inflammation of rheumatoid arthritis by targeting mitochondria. Nano-Micro Lett 2020; 12:76.

[39] Deng Y, Chen Y, Zheng H, Li B, Liang L, Su W, et al. Xuetongsu ameliorates synovial inflammatory hyperplasia in rheumatoid arthritis by inhibiting JAK2/STAT3 and NF- $\kappa$ b signaling pathways. J Ethnopharmacol 2024; 337(1):118786.

[40] Liang S, Hu D, Li G, Gao D, Li F, Zheng H, et al. NIR-II fluorescence visualization of ultrasound-induced blood-brain barrier opening for enhanced photothermal therapy against glioblastoma using indocyanine green microbubbles. Sci Bull 2022; 67(22):2316-26.

[41] Wang M, Lv CY, Li SA, Wang JK, Luo WZ, Zhao P-C, et al. Near infrared light fluorescence imaging-guided biomimetic nanoparticles of extracellular vesicles deliver indocyanine green and paclitaxel for hyperthermia combined with chemotherapy against glioma. J Nanobiotechnology 2021; 19(1):210.

[42] Xiong H, Zhao Y, Xu Q, Xie X, Wu J, Hu B, et al. Biodegradable hollow - structured nanozymes modulate phenotypic polarization of macrophages and relieve hypoxia for treatment of osteoarthritis. Small 2022; 18(32):2203240.

[43] Chen X, Dai C, Hu R, Yu L, Chen Y, Zhang B. Engineering ROS-scavenging prussian blue nanozymes for efficient atherosclerosis nanotherapy. J Mater Chem B 2023; 11(9):1881-90.

[44] Ma X, Hao J, Wu J, Li Y, Cai X, Zheng Y. Prussian blue nanozyme as a pyroptosis inhibitor alleviates neurodegeneration. Adv Mater 2022; 34(15):2106723.

[45] Lan R, Lv J, Gao D, Hu D, Liu C, Jia J, et al. Folate receptor-targeted NIR-II dual-model nanoprobes for multiscale visualization of macrophages in rheumatoid arthritis. Adv Funct Mater 2023; 33(29):2300673.

[46] Deng C, Zhang Q, He P, Zhou B, He K, Sun X, et al. Targeted apoptosis of macrophages and osteoclasts in arthritic joints is effective against advanced inflammatory arthritis. Nat Commun 2021; 12:2174.

[47] Guo Q, Jin Y, Chen X, Ye X, Shen X, Lin M, et al. NF- $\kappa$ b in biology and targeted therapy: new insights and translational implications. Signal Transduct Target Ther 2024; 9(1):53.

[48] Ilchovska DD, Barrow DM. An overview of the NF-kB mechanism of pathophysiology in rheumatoid arthritis, investigation of the NF-kB ligand RANKL and related nutritional interventions. Autoimmun Rev 2020; 20(2):102741.

[49] Zheng H, Liu Y, Deng Y, Li Y, Liu S, Yang Y, et al. Recent advances of NFATc1 in rheumatoid arthritis-related bone destruction: mechanisms and potential therapeutic targets. Mol Med 2024; 30(1):20.

[50] Bae S, Kim K, Kang K, Kim H, Lee M, Oh B, et al. RANKL-responsive epigenetic mechanism reprograms macrophages into bone-resorbing osteoclasts. Cell Mol Immunol 2022; 20(1):94-109.

[51] Park JH, Jeong E, Lin J, Ko R, Kim JH, Yi S, et al. RACK1 interaction with c-src is essential for osteoclast function. Exp Mol Med 2019; 51(7):1-9.