Lumbar degenerative disc disease is a common disabling condition that often leads to back pain, leg pain, and spinal deformities (1). When conservative treatment fails, radiculopathy caused by disc herniation is often treated surgically through lumbar interbody fusion (2). Spinal fusion is a definitive method for treating progressive spinal deformities, instability, and certain spinal infections or tumors (3). At present, there are several surgical methods for spinal fusion, the main difference lying in the surgical approach to the intervertebral disc space. Lateral lumbar interbody fusion (LLIF) directly enters the intervertebral disc laterally through the retroperitoneal space, which can reduce the dissection of the spine and paravertebral muscles. Compared to posterior lumbar interbody fusion (PLIF), LLIF provides more graft space, reduces surgical trauma and intraoperative bleeding, which is more beneficial to postoperative recovery (4–6). Despite these advantages of LLIF, it is still challenging to achieve the interbody fusion. For example, nonunion or postoperative pseudarthrosis is still a major concern (7).

Exploring efficient and convenient fusion strategies is crucial to improving the effectiveness of LLIF. Autologous bone, which does not trigger an immune response, can be used as a scaffold for the growth of blood vessels and bone tissue and provides some biological factors beneficial to osteogenesis. This is considered as the gold standard for interbody fusion (8, 9). However, harvesting autologous bone is accompanied by the risk of surgical complications and surgical pain, which limits the application in LLIF. Previous studies have shown that combining some bone biomaterials with bone morphogenetic protein (BMP) can significantly improve lumbar fusion rate (7, 10). However, excessive use of BMP would lead to adverse events, such as soft tissue swelling, local inflammation, aseptic cyst formation, osteolysis, implant displacement, ectopic bone formation, retrograde ejaculation and radiculitis (11–13). The failure rate of LLIF is between 7%–20% (14–16). In addition to the strategy of using autogenous bone or bone biomaterials during operation, postoperative medication can also improve the fusion effect. Recently, it was reported that in the posterolateral spinal fusion model of rabbits, intramuscular injection of abaloparatide combined with spinal fusion surgery significantly increased the rate and effect of fusion (17, 18). Postoperative drug therapy can achieve significant therapeutic effects and offers higher safety, providing a promising strategy to improve lumbar fusion rate.

In recent years, with the development of bone tissue engineering (BTE), an increasing number of researchers have begun to analyze various bioactive components which are derived from traditional Chinese medicine (TCM) and use them to treat orthopedic diseases. For example, Eupolyphaga sinensis (ES) is a beneficial medicinal insect, commonly used in TCM, having the function of treating fractures. ES belongs to Eupolyphaga, which is widely distributed in Asia such as Thailand, India, Malaysia, and China (19). In TCM, ES is described as having properties of invigorating blood circulation, removing blood stasis, and healing bones and tendons. Due to its accessibility and efficacy, it is widely used in folk medicine for treating orthopedic conditions in China. Previous studies have shown that ES significantly enhances vascular regeneration and promotes bone regeneration (20, 21). At the cellular level, ES can promote the differentiation of rat bone marrow mesenchymal stem cells (BMSCs) towards osteoblasts by regulating BMP (22). From both folk application and scientific research perspectives, it is evident that ES has the ability to promote bone regeneration. This suggests its potential use for enhancing lumbar fusion, although its research and application in spinal surgery are relatively limited at present.

In LLIF, a cage and bone grafts are implanted into the intervertebral space to maintain intervertebral height and provide a scaffold for bone growth. During the fusion process, blood vessels and osteoblasts from cartilage endplate grow into intervertebral space with the support of the graft and cage leading to bone bridging and interbody fusion. Therefore, it is an ideal method to enhance the biological activity of endplate through adjunctive drug therapy based on surgical strategy to promote interbody fusion. The purpose of this study is to enhance the activity of blood vessels and bone cells in the endplate by using ES, thereby improving the lumbar fusion rate.

In the historical development of traditional Chinese medicine (TCM), the application of ES and its derivatives in the treatment of orthopedic diseases can be traced back to the Eastern Han Dynasty nearly 2,000 years ago. Classic Chinese medicine, including Synopsis of the Golden Chamber and Compendium of Materia Medica, clearly recorded the safety and effectiveness of oral ES in treating tendon and bone injuries (23). In addition, some studies related to TCM also show that ES has high safety. Previous reports show that ES has a high safety profile in animal research, and an LD50 exceeding 10 g/kg in rats (24). Although ES has a high safety threshold, its effective therapeutic concentration has not been fully studied. Li et al. found that administering ES at 1,000 mg/kg/day significantly promoted the healing of mandibular fracture in rabbits (25). In this study, the raw material dosage of ES was set at 1,000 mg/kg/day, and the actual dosage was 162 mg/kg/day based on the extraction rate of 16.2%. In the previous clinical application and research, ES and its preparations were made into decoction by boiling and eaten immediately. However, this method is affected by variables such as boiling conditions, making dosage control difficult.

In our study, ES extract powder was prepared by freeze-drying, which can form uniform and adjustable drug concentration after dissolution. It is verified that ES powder can be stored at room temperature for at least 3 months, ensuring consistency of composition and efficacy during long-term use. During the experimental period, the freeze-dried ES powder was reconstituted into a 5 mg/ml solution and administered based on each rabbit's weight so that the dose of the extract can be accurately controlled and the influence of dose error on the experimental results can be reduced. Clinically, the drug concentration can be adjusted according to the individual weight difference between rabbits and humans to reduce the total volume.

At present, the extraction methods of ES primarily include water extraction and alcohol extraction. Studies have shown that ES water extract can promote the proliferation of human endothelial cells, whereas alcohol extract inhibit the proliferation of endothelial cells (20, 26). The major components of ES include amino acids, proteins, bioactive peptides, polysaccharides, lipids, alkaloids, nitrogen-containing compounds, nucleotides, fat-soluble vitamins, and inorganic substances. The differences between water and alcohol extraction are largely determined by the solubility characteristics of these components. Components with inhibitory effects on tumor cell proliferation, such as polysaccharides, oleic acid, and unsaturated fatty acids, are more efficiently extracted by alcohol. Components promoting angiogenesis and bone regeneration, such as bioactive peptides, alkaloids, and inorganic substances, are more efficiently extracted using water. ES has diverse biological functions, and different extraction methods can be used to obtain different active components according to practical therapeutic needs (27). However, comprehensive research on the complete chemical composition of ES remains limited and warrants further investigation.

This study is the first to investigate the relationship between endplate changes and interbody fusion using a rabbit interbody fusion model. Previous studies have shown that cages and bone graft serve as scaffolds and growth factors for intervertebral bone growth, thus improving the fusion rate (7). To further clarify the factors that influence interbody fusion, this study compared and analyzed the respective effects of drugs and cage between groups. Compared with the CON group, the ES group exhibited improved bone tissue parameters, likely due to the ability of ES extract which enhances endplate angiogenesis and increases intervertebral blood supply. Wei et al. reported that the culture medium containing ES extract promoted the proliferation of endothelial cells and bone marrow mesenchymal stem cells—two key cell types involved in bone tissue repair (20–22). Abundant local blood supply provides sufficient nutrients and growth factors for bone growth while removing metabolic wastes. The blood supply of intervertebral region decreases with the aging of the body, which makes the promotion of vascular ingrowth become increasingly important for interbody fusion (28). In addition, the ability of ES to promote the proliferation and osteogenic differentiation of bone marrow mesenchymal stem cells is more beneficial to enhance interbody fusion. However, due to the lack of intervertebral support in ES group, the growth of blood vessels and bone tissue into intervertebral space was hindered, resulting in the formation of bony spurs. During the fusion process, the cage and internal fixation device provide stable space support and scaffolding between the upper and lower endplates, thereby facilitating the sustained ingrowth of blood vessels and bone cells into the intervertebral space. This structural support explains why stable bone connection is finally achieved only in IBF/ES group.

Interbody fusion is characterized by “the formation of bone connections between the upper and lower endplates, including bone connections within the endplate and bone bridge connections at the endplate edge” (29, 30). Previous studies reported that the average time required for spinal fusion for small animals such as rabbits and rats was 6–8 weeks (31, 32). However, these studies employed transverse process fusion models rather than interbody fusion. Because the intervertebral region is subjected to higher mechanical stress and has relatively poor blood supply, the fusion process is typically slower and less efficient. Therefore, a more clinically relevant interbody fusion model was adopted in our study, and this model was established according to the fusion cage and graft materials currently used in clinic. Our previous research demonstrated that the lateral lumbar fusion in rabbits achieved a fusion rate of 91.7% at approximately 12 weeks postoperatively (33). However, in the current study, the IBF/ES group achieved bone fusion by the 10th week after operation, and the fusion rate was 100%. These findings suggest that ES extract significantly accelerates and enhances process of interbody fusion.

Cage subsidence and endplate collapse are common complications following lumbar fusion. When the cage sinks, it directly reduces intervertebral height, compromises anterior spinal support, and alters adjacent soft tissue structure, which may ultimately impair the result of surgical decompression. Liu et al. reported a case in which the cage subsidence led to a reduction in intervertebral height and ligament laxity, further compressing nerve roots and led to recurrence (34). Therefore, preventing cage subsidence is crucial for maintaining sagittal balance and optimizing surgical outcomes. Our study not only addressed the issue of improving fusion rates but also the prevention of cage subsidence. Oral ES extract after operation promoted angiogenesis and enhanced bone density in adjacent endplate regions, which may help prevent cage subsidence. The results of micro-CT showed significant increases in BV/TV, Tb.Th, and Tb.N in the adjacent vertebrae of the ES and IBF/ES groups, indicating a beneficial effects of ES extract on surrounding vertebral structures. Finally, the improvement of trabecular structure and the increase of adjacent vertebrae density are beneficial to prevent the loss of intervertebral height caused by cage subsidence.

One possible pathway using oral ES extract is that it can be used as an adjunctive therapy in human lumbar fusion. From a clinical standpoint, lumbar fusion typically involves the use of autologous bone grafts or allogeneic materials to promote bone regeneration. This study demonstrated that oral administration of ES extract significantly promotes angiogenesis and bone regeneration within the intervertebral region. Thus, oral ES extract has the potential to be routinely employed postoperatively for human lumbar fusion, even replacing the need for bone grafts. From the aspect of BTE, although the pharmacological composition, functions and mechanisms of ES are complex, its pronounced therapeutic effects warrant further investigation. Therefore, the future research will focus on the basic research of ES and the development of local application methods of ES drugs from combined BTE.

In conclusion, this study suggested that oral ES extract after spine surgery can significantly enhance the effectiveness and success rate of lumbar lateral fusion surgery by promoting cartilage endplate vascularization and bone formation. This study provides preclinical evidence supporting the use of ES extract to promote interbody fusion and highlights its potential application in treatment of bone-related diseases.