2025 Volume No 51 – pages 103-119
Title: Alendronate nanoparticles promote fracture healing by inhibiting LRP4 to activate the Wnt/β-catenin pathway |
Authors: ZW Sun, QL Zhu, JP Gong, YH Sui, XY Liu |
Address: Pharmaceutical Department, Yantai Yuhuangding Hospital, 264000 Yantai, Shandong, China |
E-mail: 13583544489 at 163.com; 13553106682 at 163.com |
Abstract: Background: Fracture is a common skeletal disorder, and fracture healing is a complex biological process. The Wingless-Int-Integrated/beta-catenin (Wnt/β-catenin) signaling pathway plays an important role in bone formation and fracture healing. Low-density lipoprotein receptor-related protein 4 (LRP4) has been reported to negatively regulate the Wnt/β-catenin pathway. This study investigated whether alendronate nanoparticles (alendronate-NPs) can activate the Wnt/β-catenin pathway by inhibiting LRP4, thereby promoting fracture healing. Methods: We initially synthesized alendronate-NPs. Then, the impact of alendronate, alendronate-NPs, and LRP4 on the osteogenic differentiation of rat osteoblasts was investigated. The proliferation capacity of osteoblasts was evaluated using cell-counting kit-8. The effects on the messenger ribonucleic acid (mRNA) and protein levels of proliferating cell nuclear antigen (PCNA), runt-related transcription factor 2 (Runx2), osteocalcin (OCN), osteopontin (OPN), LRP4, Wingless-type MMTV integration site family, member 4 (Wnt4), glycogen synthase kinase 3 beta (GSK-3β), p-GSK-3β, and β-catenin were analyzed by quantitative reverse-transcription polymerase chain reaction and Western blot. The impact on osteoblast apoptosis and calcium deposition was assessed using terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) and alizarin red staining assays. A rat fracture model was then established, followed by treatments with either alendronate or alendronate-NPs. Bone formation during fracture healing was evaluated using alkaline phosphatase. The effects of alendronate and alendronate-NPs on fracture healing were assessed through hematoxylin-eosin (HE) staining and immunohistochemistry staining of collagen I. Results: Alendronate and alendronate-NPs significantly downregulated the expression of LRP4 during osteoblast differentiation and upregulated the expression of downstream targets of the Wnt/β-catenin pathway (p < 0.01). The co-treatment of osteoblasts with alendronate-NPs and LRP4 overexpression partially offset the bone-formation-promoting effects of alendronate-NPs. LRP4 overexpression reversed the proliferative and osteogenic effects of alendronate-NPs (p < 0.01). Compared with alendronate-NP treatment alone, the combined treatment of alendronate-NPs and LRP4 overexpression downregulated the expression of downstream targets of the Wnt/β-catenin pathway in osteoblasts (p < 0.01). Alendronate and alendronate-NPs significantly accelerated fracture healing in rats. They also promoted the proliferation and bone-formation capacity of osteoblasts, with alendronate-NPs showing a more pronounced therapeutic effect on fracture healing than alendronate. Conclusions: Alendronate-NPs may promote fracture healing by inhibiting LRP4 to activate the Wnt/β-catenin pathway. This study provides experimental evidence for a new mechanism of alendronate-NPs in promoting fracture healing. |
Keywords: Alendronate nanoparticles, LRP4, Wnt/β-catenin pathway, bone-fracture healing. |
Publication date: 26th June 2025 |
Copyright policy: © 2025 The Author(s). Published by Forum Multimedia Publishing, LLC. This article is distributed in accordance with Creative Commons Attribution Licence (http://creativecommons.org/licenses/by/4.0/). |
Article download: Pages 103-119 (PDF file) |
