Bone remodeling is a continuous process executed by the coordinated actions of osteoclasts, which resorb bone, and osteoblasts, which synthesize new bone matrix. This osteoblast-osteoclast coupling is essential for skeletal homeostasis and is profoundly influenced by mechanical loading (18). The process of fracture healing recapitulates embryonic skeletal development, involving an inflammatory phase, soft and hard callus formation, and remodeling (19). During this cascade, various signaling pathways, including Wnt/β-catenin and BMP/Smad, are sequentially activated to ensure proper repair (20). Recent evidence has highlighted the critical role of skeletal stem cells and their niche interactions in coordinating regenerative responses (21). Pathological conditions, such as stress fractures common in athletes, arise when this delicate balance is disrupted, leading to an imbalance between bone resorption and formation (22). Moreover, immune cells and vascular endothelial cells contribute significantly to the regenerative microenvironment, influencing the recruitment and differentiation of skeletal progenitor cells (23).

Exosomes are nano-sized extracellular vesicles (30-150 nm in diameter) that mediate paracrine and endocrine signaling by transferring bioactive molecules, including proteins, lipids, and ncRNAs, between cells (24). Their biogenesis involves the endosomal pathway, beginning with the formation of intraluminal vesicles within multivesicular bodies, which subsequently fuse with the plasma membrane to release exosomes into the extracellular space (25). In the musculoskeletal system, exosomes facilitate critical crosstalk between various cell types, such as between skeletal muscle and bone, a concept central to the muscle-bone axis (26). For instance, exosomes derived from myoblasts can influence the activity of osteoclasts, highlighting their role in systemic skeletal regulation (27). The specific cargo of these vesicles, particularly ncRNAs, dictates their functional impact on recipient cells. Notably, the molecular composition of exosomes reflects the physiological or pathological state of the parent cell, making them attractive targets for biomarker discovery (28).

ncRNAs encapsulated within exosomes are pivotal regulators of gene expression in bone cells. Among these, miRNAs are the most extensively studied, functioning by binding to complementary sequences in target mRNAs to inhibit translation or promote degradation (24). In the context of bone, exosomal miRNAs have been shown to both promote and inhibit osteogenesis. For example, exosomal miR-26a derived from stem cells enhanced bone regeneration by targeting specific inhibitory factors (29), whereas other miRNAs suppressed osteoblast differentiation by modulating key transcription factors (30). LncRNAs and circRNAs add another layer of complexity, acting as competing endogenous RNAs that sponge miRNAs to modulate their activity. For instance, lncRNAs can sequester osteogenic suppressor miRNAs, thereby derepressing osteogenic gene expression (31). This complex regulatory network is now recognized as a fundamental mechanism governing the fate of bone marrow mesenchymal stem cells (BMSCs) and other skeletal progenitors (32). Furthermore, exosomal ncRNAs can simultaneously target multiple signaling nodes, providing a coordinated regulatory mechanism that single soluble factors cannot achieve (8).

Physical activity acts as a potent systemic cue that profoundly influences the exosomal landscape. Exercise, particularly resistance and high-intensity interval training, alters the number of circulating extracellular vesicles and modifies their molecular cargo (33). Garner et al (34) demonstrated that acute exercise rapidly increases the expression of genes involved in the multivesicular body and exosome pathways in skeletal muscle. Mechanical strain on skeletal muscle is a potent driver of exosome secretion, with these muscle-derived vesicles carrying specific myomiRs and other factors that can target distant organs, including bone (35). Recent research has shown that mechanically stimulated osteocytes also release exosomes enriched with osteogenic miRNAs that promote osteoblast differentiation (36). Furthermore, the metabolic and mechanical stress associated with exercise can be sensed by various tissues, such as adipose tissue and the vascular endothelium, leading to the secretion of tissue-specific exosomes (37). This exercise-induced 'exosomal signature' is dynamic, with distinct differences observed between acute bouts of exercise and long-term training, as well as being influenced by factors such as age and sex (38). Notably, the dose-effect relationship between exercise parameters, including intensity, duration, and frequency, and the resulting exosomal ncRNA profile remains incompletely characterized, representing a critical knowledge gap (39).

Different exercise modalities elicit distinct exosomal ncRNA expression profiles, reflecting the unique physiological demands of each training type. As summarized in Table I, resistance exercise has been shown to alter the surface profile and miRNA cargo of circulating extracellular vesicles, with Just et al (33) demonstrating that blood flow-restricted resistance exercise modifies the miRNA landscape in a manner distinct from traditional resistance training. Similarly, Garner et al (34) reported that a single bout of acute exercise comprising 45 min of cycling followed by resistance exercise rapidly upregulates genes involved in multivesicular body and exosome pathways in skeletal muscle, establishing a mechanistic link between mechanical stress and exosome biogenesis. By contrast, aerobic exercise training regulates serum extracellular vesicle miRNAs linked to metabolic health, as shown by de Mendonça et al (40), who observed that aerobic training modulates miRNA profiles associated with obesity-related pathways. High-intensity interval training (HIIT) was shown to induce muscle-derived exosomal miRNAs that improve insulin sensitivity through hepatic forkhead box protein O1 (FoxO1) downregulation, as demonstrated by Castaño et al (41). Maggio et al (42) further characterized that different exercise regimens, including moderate continuous training and HIIT, produce distinct modulatory effects on circulating extracellular vesicle cargo and their inflammatory properties. Collectively, these findings indicate that exercise modality serves as a critical determinant of exosomal ncRNA composition, with implications for targeted therapeutic applications.

Skeletal muscle represents the predominant tissue source of exercise-induced exosomal ncRNAs, functioning as a central mediator of the muscle-bone axis. Vechetti et al (43) demonstrated that mechanical overload-induced muscle-derived extracellular vesicles carry specific miRNAs that promote adipose tissue lipolysis, highlighting the endocrine capacity of myogenic exosomes. The release of muscle-derived exosomes into the circulation following exercise has been well characterized, with Guescini et al (44) providing foundational evidence that skeletal muscle releases α-sarcoglycan-positive extracellular vesicles carrying miRNAs into the bloodstream. Beyond skeletal muscle, adipose tissue contributes significantly to the exercise-responsive exosomal pool. Burke et al (37) showed that extracellular vesicle transfer of miR-1 to adipose tissue modifies lipolytic pathways following resistance exercise, establishing a muscle-adipose crosstalk mechanism. Additionally, Zhao et al (45) identified that small extracellular vesicles from brown adipose tissue mediate exercise-induced cardioprotection, expanding the understanding of adipose-derived exosomal signaling. Vascular endothelial cells and bone tissue also contribute to the exosomal response, with Lou et al (46) demonstrating that liver-derived extracellular vesicles carrying miR-122-5p promote exercise-induced angiogenesis. The multiplicity of tissue sources underscores the complexity of exercise-induced exosomal signaling networks.

The relationship between exercise parameters, including intensity, duration, frequency, and training cycle, and exosomal ncRNA expression follows dose-dependent patterns, although systematic characterization remains incomplete. Sapp et al (47) reported that moderate and high-intensity exercise differentially affect circulating markers of endothelial integrity, with intensity-dependent effects on vesicle release. The duration of exercise bouts also influences exosomal cargo, as demonstrated by Lovett et al (39), who analyzed plasma-derived small extracellular vesicle characteristics following exercise-induced skeletal muscle damage and observed time-dependent alterations in miRNA cargo. Regarding training frequency and cycle length, Warnier et al (48) examined the effects of a 6-week sprint interval training protocol at different altitudes on circulating extracellular vesicles, revealing that sustained training regimens produce cumulative effects on exosomal profiles. Doncheva et al (49) further demonstrated that extracellular vesicles and miRNAs are altered in response to exercise in conjunction with insulin sensitivity and overweight status, suggesting that the dose-response relationship is modulated by underlying metabolic conditions. The emerging evidence supports a non-linear, context-dependent relationship between exercise dose and exosomal ncRNA expression.

As detailed in Table I, individual characteristics substantially modify the exosomal ncRNA response to exercise. Age-related differences have been systematically characterized, with Xhuti et al (38) demonstrating that circulating exosome-like vesicle and skeletal muscle miRNAs are altered with age and resistance training, indicating that older adults exhibit distinct exosomal responses compared with younger individuals. Specifically, aging is associated with changes in exosome concentration and altered miRNA cargo, including increased levels of senescence-associated miRNAs such as miR-34a and miR-126-3p, which may contribute to age-related declines in muscle-bone crosstalk. Similarly, Estébanez et al (50) reported that resistance training diminishes the expression of exosome CD63 protein in the elderly without modifying plasma miR-146a-5p levels, highlighting age-dependent differences in exosome biogenesis and cargo. Sex differences represent another critical modifying factor, as Kargl et al (51) recently demonstrated that circulating extracellular vesicle characteristics differ between men and women following 12 weeks of concurrent exercise training. This includes baseline differences in EV concentration, size distribution, and protein cargo between sexes, as well as sex-specific responses to exercise training. Moreover, Kargl et al (52) further elucidated the varying influence of menstrual cycle phase and hormonal contraceptive use on resistance exercise-induced circulating extracellular vesicle signaling, emphasizing the complexity of sex hormone interactions with exosomal responses. Physical fitness level also modulates exosomal profiles, with Fernandez-Sanjurjo et al (53) showing that next-generation sequencing reveals distinct plasma extracellular vesicle miRNA cargo differentiating sedentary young males from athletes. Athletes exhibited significantly different levels of miR-16-5p, miR-19a-3p, miR-451a, and miR-25-3p compared with sedentary controls, suggesting that long-term training status induces lasting changes in exosomal miRNA signatures. Injury status further influences exosomal responses, as demonstrated by Lovett et al (39), who analyzed exosomal characteristics following exercise-induced skeletal muscle damage.

The temporal dynamics of exosomal ncRNA expression distinguish acute exercise responses from chronic training adaptations, with distinct molecular signatures characterizing each physiological state. Acute exercise induces rapid but transient changes in exosomal cargo, as demonstrated by Garner et al (34), who observed that acute exercise rapidly increases the expression of genes involved in multivesicular body and exosome pathways in skeletal muscle. Similarly, D'Souza et al (54) reported that circulatory exosomal miRNA following intense exercise is unrelated to muscle and plasma miRNA abundances, suggesting that acute exercise mobilizes pre-existing exosomal pools rather than reflecting ongoing transcriptional activity. Silver et al (55) corroborated these findings, demonstrating that extracellular vesicular miRNA expression is not a proxy for skeletal muscle miRNA expression following acute moderate-intensity exercise, indicating compartmentalized regulation. By contrast, chronic training has been shown to induce sustained alterations in exosomal ncRNA profiles. Hou et al (56) demonstrated that long-term exercise-derived exosomal miR-342-5p functions as a novel exerkine for cardioprotection, establishing that sustained training produces durable exosomal signatures. Di et al (57) further reported that long-term exercise-secreted extracellular vesicles promote browning of white adipocytes by suppressing miR-191a-5p, confirming that chronic exercise induces persistent changes in exosomal cargo. Beyond these specific miRNAs, profiling studies have identified numerous other exosomal miRNAs altered by chronic exercise training, including miR-16-5p, miR-19a-3p, miR-451a, and miR-25-3p, which differentiate athletes from sedentary individuals (53). The temporal resolution of exosomal responses, ranging from minutes following acute exercise to weeks of training adaptation, reveals a dynamic continuum of molecular regulation.

Exercise-induced exosomal ncRNAs promote osteoblast differentiation and bone formation primarily through activation of the Wnt/β-catenin, BMP/Smad, and MAPK/ERK signaling cascades (58). Mechanical strain-stimulated osteocyte-derived exosomes have been shown to enhance osteogenic differentiation via miRNA-mediated pathway activation. Lv et al (59) demonstrated that mechanically induced osteocyte-derived exosomes promote periodontal ligament stem cell proliferation and osteogenic differentiation through the miR-181b-5p/phosphatase and tensin homolog (PTEN)/protein kinase B (AKT) signaling axis. This finding establishes a direct mechanotransduction link where exosomal miRNAs modulate PTEN expression to relieve AKT pathway inhibition. Similarly, Zhu et al (36) reported that mechanically strained osteocyte-derived exosomes promote osteoblastic differentiation, further supporting the concept that mechanical loading generates osteogenic exosomal signals.

The Wnt/β-catenin pathway represents a central convergence point for exosomal ncRNA-mediated osteogenesis. Ibrahim et al (60) demonstrated that augmenting canonical Wnt signaling in therapeutically inert cells converts them into potent exosome factories, highlighting the capacity of the pathway to enhance exosomal therapeutic efficacy. In the context of muscle-bone crosstalk, skeletal muscle-derived exosomes have been identified as key mediators of osteogenesis. Chen et al (27) showed that histone deacetylase inhibition enhances extracellular vesicles from muscle to promote osteogenesis via miR-873-3p, indicating that muscle-derived exosomes can directly activate osteogenic programs in bone cells. Xu et al (61) provided complementary evidence that exosomes from C2C12 myoblasts enhance osteogenic differentiation of MC3T3-E1 pre-osteoblasts by delivering miR-27a-3p. Across these studies, a common regulatory pattern emerges: Exosomal ncRNAs from both osteocytes and myocytes converge on the Wnt/β-catenin pathway as a central effector. Specifically, these ncRNAs function by relieving endogenous inhibitors of the pathway (such as PTEN and DKK2) or by directly targeting transcriptional repressors of osteogenic genes. This pattern suggests that diverse exercise stimuli may converge on a shared exosomal ncRNA-mediated mechanism to activate Wnt/β-catenin signaling, representing a potential therapeutic node for bone regeneration.

Collectively, these studies demonstrate that exosomal miRNAs from both osteocytes and myocytes converge on osteogenic signaling pathways, with the Wnt/β-catenin cascade serving as a primary effector mechanism. It should be noted, however, that the majority of these studies are correlative in nature, reporting associations between exosomal ncRNA levels and osteogenic outcomes. Direct causal evidence through loss-of-function (such as exosomal ncRNA knockdown) or gain-of-function (such as ncRNA overexpression or mimic delivery) approaches remains limited, leaving the necessity and sufficiency of specific exosomal ncRNAs for exercise-induced bone formation incompletely validated.

The inhibitory effects of exercise-induced exosomal ncRNAs on osteoclastogenesis involve targeting the RANKL/RANK/OPG axis and suppressing NF-κB and nuclear factor of activated T-cells 1 (NFATc1) signaling. Nie et al (62) demonstrated that skeletal muscle-derived exosomes regulate endothelial cell functions via reactive oxygen species-activated NF-κB signaling, establishing a mechanistic link between exercise-induced oxidative stress and downstream transcriptional regulation. This pathway also modulates osteoclast precursor sensitivity to RANKL stimulation. Wang et al (63) reported that circ_0008542 in osteoblast exosomes promotes osteoclast-induced bone resorption through m6A methylation, revealing that exosomal circRNAs can influence the osteoblast-osteoclast coupling balance. The opposing effects of different exosomal ncRNA species on osteoclastogenesis underscore the complexity of exercise-mediated skeletal regulation.

Muscle-derived exosomes also contribute to anti-resorptive effects through delivery of miRNAs that target osteoclast differentiation factors. Fulzele et al (64) demonstrated that muscle-derived miR-34a increases with age in circulating extracellular vesicles and induces senescence of bone marrow stem cells, suggesting that age-related changes in exosomal cargo may compromise bone homeostasis. Conversely, Qu et al (65) showed that mechanically stimulated periodontal ligament cell-derived exosomes promote osteoblast differentiation via the miR-181d-5p/tumor necrosis factor (TNF) signaling pathway, indicating that mechanical loading generates exosomal signals that suppress inflammatory osteoclastogenesis.

Synthesizing these findings, a recurrent mechanistic theme is that exosomal ncRNAs modulate osteoclastogenesis by targeting three distinct nodes: i) The RANKL/RANK/OPG ratio, ii) NF-κB transcriptional activity, and iii) NFATc1-mediated downstream gene expression. Notably, different exosomal ncRNA species can exert opposing effects on the same pathway depending on cellular context, suggesting that the net anti-resorptive outcome of exercise is determined by the balance of pro-osteogenic and anti-osteoclastogenic exosomal signals. This balance, rather than the absolute level of any single ncRNA, likely dictates the overall effect on bone mass.

Exercise-induced exosomal ncRNAs exert pleiotropic effects on the bone microenvironment by coordinating mesenchymal stem cell lineage commitment, angiogenesis, and immune homeostasis. Ma et al (26) demonstrated that skeletal muscle-derived extracellular vesicles transport glycolytic enzymes to mediate muscle-to-bone crosstalk, revealing a metabolic dimension to exosomal communication beyond ncRNA cargo. This finding expands the understanding of how exercise generates integrated signals that support bone formation. For angiogenesis coupling, Hayashi et al (66) reported that exosomal miR-206 secreted from growing muscle promotes angiogenic response in endothelial cells, establishing a direct mechanism by which muscle-derived exosomes support the vascular niche required for bone remodeling. Similarly, Lou et al (46) demonstrated that exercise promotes angiogenesis by enhancing endothelial cell fatty acid utilization via liver-derived extracellular vesicle miR-122-5p, highlighting the multi-organ contribution to exercise-induced vascular adaptation.

Mesenchymal stem cell lineage commitment is critically regulated by exosomal ncRNAs from various sources. Xing et al (67) showed that skeletal muscle-derived exosomes prevent osteoporosis by promoting osteogenesis, providing direct evidence for the therapeutic potential of muscle-derived exosomes in bone disorders. By contrast, Peruzzi et al (68) demonstrated that circulating extracellular vesicles from adolescents with obesity impair mesenchymal stromal cell differentiation, favoring adipogenic rather than osteogenic differentiation, indicating that pathological states alter exosomal cargo to disrupt bone homeostasis. Immune modulation represents another key mechanism, as Qiu et al (69) reported that exosomes derived from BMSCs of exercise-trained mice improve wound healing by inhibiting macrophage M1 polarization, suggesting that exercise-conditioned exosomes promote a regenerative immune environment.

The transduction of mechanical forces into biochemical signals via exosomal ncRNAs constitutes a fundamental mechanobiological coupling mechanism in bone adaptation. Osteocytes serve as primary mechanosensors in bone, and their exosomal response to mechanical loading mediates downstream effects on osteoblasts and osteoclasts. Shen et al (70) demonstrated that mechanically activated mesenchymal-derived bone cells drive vessel formation via an extracellular vesicle-mediated mechanism, establishing that mechanical stimulation generates pro-angiogenic exosomal signals that coordinate bone formation with vascular support. Shang et al (71) further elucidated that extracellular vesicles allow epigenetic mechanotransduction between chondrocytes and osteoblasts, revealing that mechanical signals can be propagated across cell types through exosomal delivery of regulatory ncRNAs.

The molecular machinery underlying exosomal mechanotransduction involves specific miRNA species that respond to mechanical strain. Zheng et al (72) reported the biological characteristics of miRNAs secreted by exosomes of periodontal ligament stem cells due to mechanical force, demonstrating that mechanical loading induces distinct exosomal miRNA profiles. Qu et al (65) confirmed that cyclic stretch-induced exosomes from periodontal ligament cells promote osteoblast differentiation via the miR-181d-5p/TNF signaling pathway, providing mechanistic insight into how cyclical mechanical forces are converted into anabolic signals. Collectively, these studies establish that exosomal ncRNAs function as mechanotransducers, converting physical stimuli into molecular signals that orchestrate adaptive bone remodeling.

The muscle-bone axis represents a paradigm of inter-organ communication mediated by exercise-induced exosomal ncRNAs, where synergistic and antagonistic signaling interactions coordinate skeletal adaptation. Multiple recent reviews have synthesized this emerging field. Zhang et al (8) comprehensively summarized exosomal miRNAs in muscle-bone crosstalk, detailing mechanistic links and exercise modulation relevant to sarcopenia and osteoporosis. Li et al (73) provided a systematic overview of molecular communication from bone to skeletal muscle, establishing that exosomal signaling is bidirectional between these tissues. Kirk et al (74) further contextualized bone and muscle crosstalk in ageing and disease, highlighting the clinical relevance of exosome-mediated inter-organ communication.

Specific molecular mechanisms underlying muscle-bone crosstalk have been elucidated. Qin et al (75) demonstrated that myostatin inhibits osteoblastic differentiation by suppressing osteocyte-derived exosomal miR-218, revealing a novel mechanism by which muscle-derived factors regulate bone formation through exosomal pathways. Li et al (76) showed that myoblast-derived exosomal paired related homeobox 2 attenuates osteoporosis via transcriptional regulation of lncRNA-MIR22HG to activate the Hippo pathway, establishing a non-canonical signaling axis in muscle-bone communication. Conversely, Wang et al (77) reported that slowpoke homolog 1 (Slo1)-deficient myoblast exosome-derived miR-222-3p inhibits osteogenic differentiation via targeting signal transducer and activator of transcription 3 (STAT3), demonstrating that pathological alterations in muscle exosomal cargo can negatively impact bone. The emerging evidence supports that exercise optimizes muscle-bone crosstalk by enriching exosomal ncRNA cargo with pro-osteogenic and anti-osteoclastogenic signals, while antagonistic signals from pathological muscle states can disrupt skeletal homeostasis.

The regulatory roles of exercise-induced exosomal ncRNAs in traumatic fracture healing span the sequential phases of inflammation, soft callus formation, hard callus mineralization, and remodeling. During the inflammatory phase, exosomal miRNAs modulate macrophage polarization and cytokine secretion, creating a pro-regenerative environment. Recent evidence from Lu et al (78) demonstrated that an osteoblast/osteoclast and immune cocktail therapy using exosome/drug delivery multi-functional hydrogel accelerates fracture repair, highlighting the potential of exosome-based strategies to coordinate multicellular responses during bone healing. The transition to soft callus formation involves exosomal transfer of osteogenic signals between mesenchymal stem cells and osteoprogenitors. Liu et al (79) showed that hypoxic mesenchymal stem cell-derived exosomes promote bone fracture healing by transferring miR-126, which enhances angiogenesis and osteogenesis simultaneously.

During hard callus mineralization, exosomal ncRNAs regulate osteoblast differentiation and matrix deposition. Jiang et al (80) reported that BMSC-derived exosomal miR-25 regulates the ubiquitination and degradation of Runx2 by SMAD specific E3 ubiquitin protein ligase 1 (SMURF1) to promote fracture healing in mice. More recent work by Yang et al (81) demonstrated that mesenchymal stem cell-derived extracellular vesicles ameliorate temporomandibular joint osteoarthritis by suppressing osteoclast activity via the let-7a-5p/integrin β3 axis, establishing that exosomal mechanisms are relevant across different joint contexts. In the remodeling phase, exosomal signals coordinate osteoclast-mediated resorption with new bone formation. The stage-dependent effects of exosomal ncRNAs suggest that therapeutic strategies targeting specific healing phases may optimize fracture repair outcomes.

Stress fractures and overuse bone injuries in athletes represent a distinct pathological entity characterized by repetitive mechanical loading-induced bone fatigue damage, where exercise-induced exosomal ncRNAs may serve as early warning signals and repair mediators. The identification of circulating exosomal miRNA signatures associated with bone fatigue damage offers potential for early detection before radiographic changes appear. Xu et al (82) demonstrated that exosomal miR-128-3p from mesenchymal stem cells of aged rats regulates osteogenesis and bone fracture healing by targeting Smad5. The mechanosensitive nature of exosomal release positions these vesicles as ideal candidates for monitoring cumulative bone fatigue. Shang (83) comprehensively reviewed mechanosensitive miRNAs in cartilage and subchondral bone remodeling, identifying them as emerging targets for osteoarthritis therapy, with direct relevance to overuse injuries.

Furthermore, Yang et al (84) confirmed that cyclic stretch-induced exosomes from periodontal ligament cells promote osteoblast differentiation via the miR-181d-5p/TNF signaling pathway, indicating that mechanically stimulated exosomes actively participate in the repair of fatigue-damaged bone. Recent work by Tian et al (85) demonstrated that miR-877, an exosomal miRNA from mechanical stretch-induced adipose derived stromal cells, enhances fracture healing in nonunion rats with type 2 diabetes mellitus, providing evidence that mechanically conditioned exosomes can overcome healing impairments. The dual role of exercise-induced exosomes, both as sensors of mechanical overload and as mediators of adaptive repair, positions them as critical regulators of the bone fatigue response.

Osteochondral injuries and post-traumatic osteoarthritis represent common sequelae of sports-related joint trauma, with exercise-induced exosomal ncRNAs modulating cartilage-subchondral bone crosstalk and disease progression. The bidirectional communication between cartilage and bone via exosomal ncRNAs influences both acute injury responses and chronic degenerative changes. Shang et al (71) elucidated that extracellular vesicles allow epigenetic mechanotransduction between chondrocytes and osteoblasts, revealing that mechanical signals can be propagated across joint tissues through exosomal delivery of regulatory ncRNAs. This mechanism is particularly relevant to sports injuries where aberrant joint loading disrupts normal cartilage-bone homeostasis. Wu et al (86) reported that osteoarthritic subchondral bone releases exosomes that promote cartilage degeneration, establishing a pathological feedback loop that accelerates osteoarthritis progression. Recent advances by Lin et al (87) demonstrated that osteocyte-derived extracellular vesicles mediate the bone-to-cartilage crosstalk and promote osteoarthritis progression, providing mechanistic insight into how bone pathology drives cartilage degeneration.

Conversely, protective exosomal signals from healthy tissues can mitigate injury-induced degeneration. Wang et al (88) demonstrated that mesenchymal stem cell-derived extracellular vesicles inhibit osteoporosis via miR-27a-induced inhibition of the DKK2-mediated Wnt/β-catenin pathway. More recent work by Gu et al (16) showed that bone remodeling stimulated by Wnt-mediated mitophagy-regulated extracellular vesicles in subchondral bone contributes to osteoarthritis development, highlighting the interconnected nature of bone and cartilage pathology. The identification of osteoarthritis-specific exosomal miRNA signatures provides potential biomarkers for early detection of post-traumatic osteoarthritis following sports injuries. Qiu et al (89) comprehensively reviewed the therapeutic potential of exosomal ncRNAs in bone metabolic diseases, focusing on osteoarthritis and rheumatoid arthritis.

Immobilization and disuse following sports injury create a catabolic environment characterized by rapid bone loss, where exercise-induced exosomal ncRNAs may exert protective effects against disuse osteopenia. The absence of mechanical loading during immobilization reduces endogenous exosomal signals that normally maintain bone mass, creating an opportunity for exogenous or exercise-mimetic exosome therapies. Van Pelt et al (90) demonstrated that serum extracellular vesicle miR-203a-3p content is associated with skeletal muscle mass and protein turnover during disuse atrophy and regrowth, establishing a link between exosomal cargo and disuse-induced tissue loss. This finding suggests that monitoring exosomal miRNAs could guide rehabilitation strategies following immobilization. Yang et al (91) reported that human umbilical cord mesenchymal stem cell-derived exosomes act via the miR-1263/Mps one binder 1 (Mob1)/Hippo signaling pathway to prevent apoptosis in disuse osteoporosis. Recent work by Shao et al (92) demonstrated that atrophic skeletal muscle-derived extracellular vesicles transfer miR-125a-5p to inhibit bone formation in osteoporosis during aging, revealing that pathological changes in muscle during disuse can actively promote bone loss. This finding underscores the importance of maintaining muscle activity during immobilization. Bao and He (93) showed that skeletal muscle-derived exosomes prevent osteoporosis by promoting osteogenesis, indicating that delivering muscle-derived exosomes could counteract disuse osteopenia. More recently, Wang et al (77) reported that Slo1-deficient myoblast exosomes-derived miR-222-3p inhibits osteogenic differentiation via targeting STAT3, demonstrating that the quality of muscle-derived exosomes critically determines their effects on bone. The concept of using exercise-conditioned exosomes as a replacement therapy during immobilization represents a promising translational strategy for athletes recovering from injury.

Circulating exosomal ncRNAs have emerged as promising non-invasive biomarkers for early diagnosis, risk stratification, and prognosis evaluation of sports-related bone injuries. Their stability in body fluids and tissue-specific origin make them attractive candidates for liquid biopsy approaches. Shi et al (94) identified miRNAs in serum exosomes as circulating biomarkers for postmenopausal osteoporosis, demonstrating the feasibility of exosomal miRNA-based diagnostics for bone disorders. Similarly, Shao et al (95) established serum exosomal miRNA expression profiling in menopausal females with osteoporosis using high-throughput sequencing, providing a reference for biomarker discovery. The prognostic value of exosomal ncRNAs has been demonstrated in fracture healing complications. Hou et al (96) reported a prospective analysis showing that elevated exosomal miR-21 levels are associated with nonunion in clavicular fractures post-open reduction and internal fixation (ORIF), providing direct clinical evidence linking exosomal cargo to fracture healing outcomes. For osteoarthritis, Zhao and Xu (97) identified synovial fluid-derived exosomal lncRNA PCGEM1 as a biomarker for different stages of osteoarthritis, highlighting the potential for staging sports-related joint injuries. More recently, Sun et al (98) identified and evaluated circulating exosomal miRNAs for the diagnosis of postmenopausal osteoporosis, further validating the biomarker potential of exosomal ncRNAs. The specificity of exosomal signatures for distinct injury types and healing stages suggests that multi-panel approaches may achieve higher diagnostic accuracy than single biomarkers.

Native exosomes derived from exercise-conditioned tissues offer naturally optimized therapeutic vehicles for bone injury repair. These vesicles inherently carry the molecular cargo that mediates the beneficial effects of exercise, potentially circumventing safety concerns associated with synthetic delivery systems. Deininger et al (99) demonstrated that anatomical implant region critically determines the osteogenic potency of small extracellular vesicles, highlighting the importance of delivery site optimization. The extraction and standardization of exercise-modulated exosomes face challenges related to donor variability, exercise protocol standardization, and scalable production. Deluca et al (100) showed a synergistic effect of umbilical cord extracellular vesicles and recombinant human bone morphogenetic protein-2 (rhBMP-2) to enhance regeneration of metaphyseal femoral defects in osteoporotic rats, suggesting that exosomes can be combined with established growth factors for enhanced efficacy. Wu et al (101) demonstrated that regulatory T cell-derived exosomes mediated macrophage polarization for osteogenic differentiation in fracture repair, revealing that immune cell-derived exosomes may complement exercise-derived vesicles in promoting bone healing. The clinical application potential of exercise-derived exosomes requires rigorous quality control and standardization comparable to pharmaceutical products.

Engineered exosomes designed to deliver specific ncRNA cargo represent a next-generation therapeutic strategy for refractory bone injuries. These approaches leverage the natural targeting capabilities of exosomes while incorporating synthetic modifications to enhance therapeutic efficacy. Strategies include direct engineering of parent cells to overexpress therapeutic ncRNAs, which are then naturally packaged into secreted exosomes, as well as post-isolation modification of exosomes with targeting ligands or therapeutic cargo. Li et al (102) demonstrated the use of engineered stem cells to produce exosomes with enhanced bone regeneration effects, presenting an alternative strategy for gene therapy that avoids direct stem cell transplantation. The delivery of ncRNA therapeutics through engineered exosomes offers advantages including protection from degradation, targeted delivery, and sustained release. For example, exosomes can be surface-modified with targeting peptides to achieve bone-specific delivery, or loaded with therapeutic ncRNAs via electroporation or chemical transfection to enhance their osteogenic or anti-resorptive potency. Huang et al (103) developed engineered exosomes as targeted lncRNA MEG3 delivery vehicles for osteosarcoma therapy, demonstrating the versatility of exosome engineering for musculoskeletal applications. Xu et al (104) developed osteoclast-targeted delivery of anti-miRNA oligonucleotides by red blood cell extracellular vesicles, showing that cell-specific targeting can be achieved through engineering approaches. For osteoarthritis, Wang et al (105) described extracellular vesicle-mediated miR-150-3p delivery in joint homeostasis as a potential treatment, illustrating the therapeutic potential of miRNA-based exosome therapies. More recently, Yan et al (106) engineered delta-like ligand 4 (Dll4)-overexpressing osteocyte-derived exosomes to enhance bone regeneration by regulating osteogenesis and angiogenesis, providing a blueprint for tissue-specific exosome engineering.

Beyond these established approaches, recent advances have focused on optimizing the stoichiometry and spatio-temporal control of ncRNA loading to enhance therapeutic efficacy. Comparative studies have shown that electroporation achieves the highest loading efficiency for small ncRNAs (miRNAs and siRNAs) into exosomes, whereas chemical transfection methods (such as Lipofectamine) yield lower encapsulation rates and may introduce residual carrier toxicity that confounds functional outcomes (15). For bone-targeted delivery, surface modification of exosomes with bone-homing peptides, such as the (DSS)₆ bone-homing peptide (a six-repeat aspartate-serine-serine sequence) that binds hydroxyapatite, has been successfully employed to concentrate therapeutic ncRNAs at fracture sites while minimizing systemic off-target effects (17,21). Furthermore, emerging evidence supports the use of sequential loading strategies, where pro-angiogenic ncRNAs (such as miR-126) and osteogenic ncRNAs (such as miR-26a) are packaged into distinct exosome batches and administered according to the temporal phases of bone healing, first promoting vascularization during soft callus formation, followed by enhancing mineralization during hard callus remodeling (23,79). This phased approach mimics the natural time-course of fracture repair and has shown superior outcomes compared with single-cargo or bolus administration in preclinical models (23). However, the clinical translation of such multi-cargo, temporally programmed exosome therapeutics requires rigorous validation of batch-to-batch consistency, long-term storage stability, and scalable manufacturing under Good Manufacturing Practice conditions, barriers that remain incompletely addressed (15,21).

The integration of exercise-induced exosomal ncRNAs with existing treatment modalities offers synergistic opportunities for enhanced therapeutic outcomes. Combination strategies encompass stem cell therapy, biomaterials, physical therapy, and pharmacotherapy. Yang et al (107) demonstrated combination therapy with BMSC-derived exosomes and porous tantalum for the repair of femur supracondylar defects, showing that exosomes can enhance the osteointegration of metallic implants. Similarly, Zheng et al (108) incorporated small extracellular vesicles in polyethylene glycol (PEG)/hyaluronic acid (HA)-Bio-Oss hydrogel composite scaffold for bone regeneration, illustrating the synergy between exosomes and biomaterial scaffolds. Yu et al (109) demonstrated synergistic effects of Shed-derived exosomes, Cu²⁺, and an injectable hyaluronic acid hydrogel on antibacterial, anti-inflammatory, and osteogenic activity for periodontal bone regeneration, showcasing multi-component combination approaches. Zhang et al (110) developed exosome-loaded hyaluronic acid hydrogel composite with oxygen-producing 3D printed polylactic acid scaffolds, combining advanced manufacturing with exosome delivery for bone tissue repair. For fracture repair, Wu et al (101) showed that regulatory T cell-derived exosomes mediated macrophage polarization for osteogenic differentiation, suggesting that immune modulation combined with osteogenic stimulation may optimize healing. The comparative analysis of these combination strategies reveals that scaffold-based delivery provides sustained local release, while systemic administration may benefit multifocal injuries. Future studies should systematically evaluate the relative efficacy of different combination approaches to establish evidence-based guidelines for clinical translation.

Clinical evidence linking exercise-induced exosomal ncRNAs to the occurrence, severity, and prognosis of sports-related bone injuries is emerging through human cohort studies. Circulating exosomal ncRNAs have been investigated as non-invasive biomarkers for fracture healing outcomes and complication risk. Shi et al (94) identified miRNAs in serum exosomes as circulating biomarkers for postmenopausal osteoporosis, establishing a foundation for similar approaches in sports-related bone injuries. The association between exosomal miRNA profiles and fracture healing complications has been demonstrated in clinical populations. Hou et al (96) reported a prospective analysis showing that elevated exosomal miR-21 levels are associated with nonunion in clavicular fractures post-ORIF, providing direct clinical evidence linking exosomal cargo to fracture healing outcomes. This finding supports the potential of exosomal miRNAs to predict delayed healing or nonunion following sports-related fractures. Recent advances by Frank et al (111) demonstrated that circulating miRNAs are associated with successful bone regeneration, providing a framework for using exosomal ncRNAs as prognostic markers. The prognostic value of exosomal signatures extends to bone metabolic status following injury. Mihanfar et al (112) demonstrated that extracellular vesicles and miRNAs are altered in response to exercise, insulin sensitivity and overweight status, indicating that metabolic factors influence exosomal profiles and may confound biomarker interpretation. Gao et al (113) recently established a plasma-derived exosomal miRNA signature by small RNA sequencing for early detection of postmenopausal osteoporosis, demonstrating the feasibility of exosomal miRNA-based diagnostics. Zhang et al (8) comprehensively reviewed exosomal miRNAs in muscle-bone crosstalk, detailing mechanistic links and exercise modulation with direct implications for sarcopenia, osteoporosis, and osteosarcopenia. Longitudinal studies tracking exosomal ncRNA changes during rehabilitation are needed to establish causality and clinical utility. The integration of exosomal biomarkers into clinical practice for sports-related bone injuries requires standardization of isolation protocols and validation across diverse athlete populations.

Current clinical evidence on exercise-induced exosomal ncRNAs in bone metabolism is predominantly derived from studies with inherent methodological constraints that limit generalizability. The majority of investigations feature small sample sizes and lack multi-center validation, compromising statistical power and external validity. For instance, studies examining circulating exosomal miRNA signatures in fracture healing typically enroll fewer than 100 participants, precluding robust subgroup analyses (114,115). Population heterogeneity represents another critical concern, as individual characteristics such as age, sex, fitness level, and hormonal status profoundly influence exosomal responses, yet stratification is frequently inadequate (116,117). Furthermore, exercise intervention protocols vary considerably across studies, with inconsistent parameters regarding intensity, duration, and training modality, impeding direct comparisons and meta-analytic synthesis (118,119). The absence of standardized pre-analytical procedures, including sample collection timing relative to exercise bouts and anticoagulant selection, introduces additional variability (120). Collectively, these limitations underscore the urgent need for prospective, multi-center large-cohort studies employing harmonized protocols to establish clinically actionable exosomal biomarkers.

Fundamental mechanistic questions persist regarding whether exercise-induced exosomal ncRNAs exert direct causal effects on bone remodeling or merely represent correlative biomarkers of systemic adaptation. A substantial proportion of published studies relies on descriptive profiling without functional validation, leaving causal inference inadequately supported. Specifically, numerous studies cited in this review (such as those reporting acute exercise-induced changes in circulating exosomal miRNA levels) demonstrate correlation rather than causation. While these findings are valuable for hypothesis generation, they do not constitute molecular mechanism demonstration. Although in vitro and animal models have demonstrated that exosomal miRNAs such as miR-181b-5p and miR-27a-3p directly modulate osteogenic pathways (121,122), the translational relevance of these findings to human physiology remains uncertain due to species differences in ncRNA sequences, target repertoires, and exosome biogenesis machinery (123).

A more fundamental issue is that even in animal studies, few have employed rigorous causal validation frameworks. The minimum requirements for establishing causality in this field should include: i) Selective knockout or knockdown of specific exosomal ncRNAs in relevant parent cells (such as skeletal muscle or osteocytes) followed by exercise intervention; ii) rescue experiments using ncRNA mimics or engineered exosomes; and iii) tissue-specific ablation of exosome biogenesis machinery (such as Rab27a or nSMase2 knockout) to determine whether exosomal ncRNAs are necessary for exercise-mediated bone protection. To date, no study has comprehensively met these criteria, representing a critical gap in the literature.

Cell and tissue specificity of exosomal ncRNA actions constitutes another unresolved controversy. Current evidence suggests that exosomes from distinct sources, including skeletal muscle, osteocytes, and adipose tissue, may exert overlapping yet distinct effects on bone cells (124,125); however, the molecular determinants governing recipient cell specificity remain poorly characterized. Additionally, the potential off-target effects of exosomal ncRNAs on non-skeletal tissues raise safety concerns that have not been systematically evaluated. Moreover, most current studies have examined the effect of a given exosomal ncRNA on only one cell type in isolation, without systematically comparing its functional consequences across osteoblasts, osteoclasts, and BMSCs; direct extrapolation of these reductionistic in vitro findings to in vivo bone adaptation lacks scientific validity. The field therefore requires a paradigm shift from descriptive correlative studies to hypothesis-driven mechanistic investigations using loss-of-function and gain-of-function approaches with cell-type-specific targeting.

The absence of standardized protocols for exosome isolation and characterization represents a fundamental barrier to data reproducibility and cross-study comparability. Ultracentrifugation, the most widely employed isolation method, exhibits variable efficiency and co-pellets non-vesicular contaminants, including protein aggregates and lipoproteins (126,127). Size-exclusion chromatography offers improved purity but suffers from lower yield and longer processing times, while polymer-based precipitation methods frequently co-precipitate non-exosomal materials that confound downstream analyses (120,128). The distinction between exosomes and other extracellular vesicle subtypes, as well as lipoprotein particles of similar size, remains technically challenging, with conventional characterization methods lacking sufficient resolution (129). For ncRNA detection, the choice of isolation method profoundly influences miRNA profiles, yet consensus guidelines for method selection are lacking. Functional validation methodologies also exhibit heterogeneity, with limited standardization in exosome labeling, tracking, and uptake assays (128). The development of reference materials and validation protocols, as recently advocated (130), represents a critical priority for advancing the field toward clinical implementation.

Translating exercise-induced exosomal ncRNAs into clinical applications for sports-related bone injuries confronts formidable obstacles spanning manufacturing, safety, and regulatory domains. Scalable production of therapeutic exosomes with consistent quality attributes remains technically demanding, as current culture conditions for exercise-mimetic exosome generation lack standardization (131). Good Manufacturing Practice compliance necessitates rigorous quality control measures, including characterization of size distribution, cargo composition, and batch-to-batch consistency, which are not routinely implemented in research settings (132). The optimal route of administration for exosome-based therapies in bone injuries remains undefined, with limited comparative data on local vs. systemic delivery, dosing frequency, and tissue biodistribution (133). Long-term safety evaluations, including immunogenicity, potential for off-target effects, and carcinogenicity, have not been systematically conducted (134). Furthermore, regulatory frameworks for exosome-based therapeutics are still evolving, creating uncertainty regarding approval pathways and quality requirements (132,135). These translational barriers necessitate coordinated efforts between academic researchers, regulatory agencies, and industry partners to establish clear guidelines and accelerate clinical development.