Introduction

Stem cells are a population of undifferentiated cells with unique abilities to self-renew and recreate functional tissues. They are primarily classified by their differentiation potential, origin and lineage progression. According to their potency, stem cells can be totipotent, pluripotent, multipotent, oligopotent and unipotent.1 Stem cells exist both in embryos and adult cells. Embryonic stem cells (ESCs) and induced pluripotent stem cells (iPSCs) are best examples of pluripotent stem cells,2 whereas adult multipotent stem cells are exemplified by hematopoietic stem cells (HSCs),3 mesenchymal stem cells (MSCs),4 neural stem cells (NSCs),5 and endothelial stem/progenitor cells (EPCs)6 (Fig. 1a). All these subtypes of stem cells have been extensively trialed for the treatment of human diseases.

Fig. 1
figure 1

Illustration of the upstream measures of exosome therapy (figure generated using Autodesk 3ds Max 2023). a production and purification of exosomes (MSCs and NSCs are used as examples for multipotent stem cells). b content of natural exosomes. c modification of exosomes. (BM bone marrow, DC dendritic cell, IAC immunoaffinity chromatography, iPSC induced pluripotent stem cell, MHC major histocompatibility complex, miRNA microRNA, MSC mesenchymal stem cell, MVB multivesicular body, NSC neural stem cell, SEC size-exclusion chromatography, UC umbilical cord)

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Stem cell-based therapy, as a modality of regenerative medicine, has generated tremendous attention, as it offers new options for patients suffering from previously incurable diseases. Subsequently, thousands of related clinical trials have been registered, covering a wide spectrum of medical problems, such as musculoskeletal and neurological disorders, immune diseases, hematological dysfunctions, and degenerative conditions.7 However, some trials have failed to show any benefit in the clinic. This is likely due to the inevitable limitations of stem cell therapy, such as infusion toxicity, immunogenicity, tumorigenic potentials and ethical issues.8 Exosome, secreted by almost all cell types including stem cells (Fig. 1a), has been posited as a safer and more versatile alternative to stem cell therapy.9

Exosomes are nanoscale, spherical, and lipid bi-layered single membrane extracellular vesicles, which act as intercellular messengers.10 Exosomes have been regarded as miniature versions of their parental cells, partially because exosomes from a certain cell type provide cell-specific or unique sets of biomolecules. In addition, the stem cells have been found to function in a paracrine fashion through their soluble secretome including exosomes.11 In other words, stem cell-derived exosomes (SC-Exo) inherit similar therapeutic effects from their parental cell of origin, e.g., anti-inflammation, immunomodulation and tissue regeneration.12 Collectively, stem cell-derived exosomes are a potent surrogate for stem cell therapy without exhibiting the disadvantages their cellular counterparts present13 (Table 1).

Table 1 The comparison between stem cell therapy and stem cell-derived exosome therapy
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Prior to clinical applications, exosomes must be prepared and optimized in terms of production, purification, and modification (Sections 2.3 and 2.4). A wide range of medical reviews analyzing these upstream measures of exosome therapy have been published in recent years. Nevertheless, some research avenues remain under-investigated: in particular, systematic investigation dedicated to downstream clinical applications is lacking, especially from a surgical perspective. Tissues that have been damaged, whether by disease or a surgeon’s scalpel, respond by inflammatory and regenerative dynamics,14 making surgery a perfect arena for stem cell-derived exosome therapy.15 Stem cell-derived exosomes inherit similar therapeutic effects from their parental cell of origin, e.g., tissue regeneration, anti-inflammation and immunomodulation.12,16,17,18

In this work, we will dissect relevant publications from the last five years in order to present a comprehensive, up-to-date, specialty-specific and disease-oriented review (Fig. 2). Our aim is to bridge the gap that currently exists between surgeons, nanomedicine practitioners, and stem cell researchers.

Fig. 2
figure 2

Illustration of the downstream surgical applications of exosome therapy (figure generated using Adobe Photoshop 2023 and Adobe Illustrator 2023). The therapeutic effects of exosomes are a hierarchical translation through disease-specific tissue responses, tissue-specific cellular alterations, and cell-specific molecular signaling pathways

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General background of exosomes and exosome therapy

Biogenesis, composition, and uptake of exosomes

Exosomes differ from other types of primary extracellular vesicles (e.g., apoptotic bodies and microvesicles) in terms of size, content, and production mechanism.19 The most popularly accepted mechanism of exosome formation, i.e., an endosomal route, is as follows (Fig. 1a). The initial endosomes are produced by cell membrane invagination during which the bioactive substances begin to accumulate within the early sorting endosomes. The late sorting endosomes then form multivesicular bodies (MVBs) after a second indentation. Finally, the MVBs fuse with the cell membrane, releasing the carried exosomes to the outside. Non-endosomal route of exosome biogenesis, such as plasma membrane budding, has also been reported.20

As the three major exosome databases (i.e., ExoCarta, Vesiclepedia, and EVpedia) summarize, exosomes contain numerous molecules, including proteins, glycoconjugates, lipids, nucleic acids, metabolites, and other bioactive substances (Fig. 1b). The examples of each category and the corresponding functions have been thoroughly reviewed elsewhere.21,22 On the one hand, exosomes comprise a complex protein network including external proteins (e.g., tetraspanins, antigen-presenting complexes, and adhesion molecules) and internal proteins (e.g., heat shock proteins, ESCRT machinery, cytokines and chemokines, and membrane transporters).23 On the other hand, as the most abundant in human exosomal nucleic acids, microRNA (mRNA) could participate in hematopoiesis, exocytosis, and nerve and vascular regeneration through exosome-mediated cellular communication.24

There are various uptake mechanisms once exosomes reach the recipient cell, all of which can be categorized into membrane fusion, receptor interaction, and internalization21 (Fig. 1b). Finally, the exosomal cargos are released into the cytoplasm, the process of which depends on the source of the exosome, nature of the cargo, and the metabolic state of the recipient cell.25 The entire lifecycle from exosome biogenesis to uptake and intracellular signaling can be tracked using fluorescent, luminescent, and radioactive techniques.26,27

Source and classification of exosomes

Depending on whether exosomes have been artificially modified, they are broadly classified into natural exosomes and engineered exosomes (Section 2.4). Depending on the species of origin, exosomes are divided into animal-derived and plant-derived exosomes. Currently, exosomes are mainly classified according to the type of their parental cells. Almost all types of human cells can produce exosomes. These include, but are not limited to, macrophages, dendritic cells (DCs), platelets, stem cells, and even tumor cells28 (Fig. 1a).

For example, macrophage-derived exosomes contribute to disease progression (e.g., diabetes, atherosclerosis and heart failure)29 and disease treatment (e.g., cutaneous wound, inflammatory bowel disease, and fungal and viral infection).30 However, they seem to play paradoxical roles in suppressing and promoting tumors.31 Like DCs, DC-derived exosomes (Dex) could also interact with immune cells (e.g., T cells, B cells, and NK cells) through their surface proteins such as major histocompatibility complexes (MHCs).32 Some preclinical and clinical trials have demonstrated the effectiveness and safety of Dex-based immunotherapy for cancers.33 Furthermore, tumor-derived exosomes (Tex) not only are involved during tumor proliferation, invasion, metastasis, and immunity but also can be used as biomarkers for cancer diagnosis and treatment.34 Lately, Tex has been used as an anti-tumor drug and an antigen presenter for DC vaccination, serving as a promising cell-free cancer immunotherapy.35 Finally, the clinical applications of stem cell-derived exosomes will be discussed in detail in the following sections.

Exosomes can be found in all body fluids such as blood, saliva, urine, plasma, tears, semen, amniotic fluid, and even breast milk.36 Body fluid-derived exosomes are a highly stable reservoir of disease biomarkers, assisting liquid biopsy in various clinical settings such as cancers, cardiovascular diseases, and perinatal disorders.37,38 However, the coexisting contents and availability of each type of body fluid might create challenges to exosome isolation.

Production, isolation and purification of exosomes

One of the major obstacles preventing exosome-based therapeutics from entering clinical practice is the low yield and efficiency of exosomes. For example, only less than 1 μg exosomal protein could be harvested from 1 ml culture medium in a laboratory setting.39 There are various methods of upscaling exosome production, which are categorized into biochemical strategies (e.g., LPS, BMP-2, HIF-1α, and IFN-γ and TNF-α), physical strategies (hypoxia, thermal stress, and starvation), mechanical strategies (shear stress and 3D culturing) and instrumental strategies (hollow-fiber bioreactors and stirred tank bioreactors).40

Exosomes are heterogeneous in terms of size, content, surface markers, and source, which makes their isolation difficult. The currently available techniques for exosome isolation and purification are based on their size, surface charge, or immunoaffinity26 (Fig. 1a). However, there is no ‘one-fits-all’ approach as these techniques all have advantages and disadvantages.

For example, ultracentrifugation is deemed the gold standard for exosome extraction. Although it requires minimal reagents and expertize, the time consumption, high cost, low efficiency, and lipoprotein co-separation have limited its large-scale use.41 Immunoaffinity chromatography is a separation technology based on the specific binding of antibodies and ligands. It is rapid and provides high purity, specificity, and yield. However, the antigen/protein coupling used needs to be expressed on the surface of exosomes.19 Size-based isolation techniques mainly refer to ultrafiltration and size-exclusion chromatography, both of which are quick and suitable for large-scale applications. But pore clogging, exosome loss, and low purity are making this method difficult to popularize.42 Although no single technique is perfect, combining the above techniques with others (e.g., precipitation-based and microfluidics-based) might be a solution to simultaneously meet multiple requirements for exosome isolation and purification.

Modification of exosomes

Exosomes can be biochemically modified to broaden, change, or improve their therapeutic effects. The modification of exosomes is classified into internal strategies (e.g., drug loading) and external strategies (e.g., surface modification). On the one hand, exosomes may be an ideal therapeutic carrier to deliver drugs, nucleic acids, and vaccines due to their advantages in stability, non-immunogenicity, and targeting recipient cells.43 There are various cargo loading techniques including pre-production loading methods (e.g., transfection, co-incubation, and electroporation) and post-production loading methods (e.g., freeze-thaw cycles, incubation, sonication, extrusion, and hypotonic dialysis) depending on whether they are applied before or after exosome biogenesis10,26,44,45,46 (Fig. 1c). For example, Tian et al. loaded doxorubicin in Dex using electroporation for the treatment of breast cancer.47 Kim et al. loaded paclitaxel in RAW 264.7-derived exosomes using incubation and sonication to overcome multidrug resistance in cancer cells.48 Ohno et al. loaded antitumor let-7a miRNA in HEK293-derived exosomes using transfection to manage breast cancer.49

On the other hand, surface modification of exosomes is exemplified by genetic engineering of exosomal membrane or parental cells, chemical connection of targeting ligands, electrostatic interaction, and magnetic nanoparticle technology.10 The main purpose of surface modification is to selectively deliver exosomes to target cells for precise treatment. For example, Alvarez-Erviti et al. modified DCs using genetic engineering to express Lamp2b and RVG peptides, thereby targeting the central nervous system (CNS).50 Zhu et al. inserted tumor-targeting peptides, c(RGDyK), into the exosome surface using a chemical reaction to target glioblastoma.51 Nakase et al. bound exosomes with a complex formed by pH-sensitive fusion peptide and cationic lipid using electrostatic interaction, thereby achieving enhanced cytosolic delivery.52

Characterization and verification of exosomes

Exosomes need to undergo characterization and verification before therapeutic applications. Current methods used for exosome characterization mainly focus on the size, morphology, and cargo profile of exosomes.43 Size-oriented verification includes nanoparticle tracking analysis (NTA), dynamic light scattering (DLS), and tunable resistive pulse sensing (TRPS), whereas morphology-oriented analysis includes scanning electron microscopy (SEM) and transmission electron microscopy (TEM).19 In addition, cargo profiling is further subdivided into proteomic, lipidomic, and genomic analyses including western blotting, ELISA, flow cytometry, mass spectroscopy, and PCR.36 Since each of the above characterization methods has advantages and disadvantages, it is a universal practice to combine analyses from three different aspects, e.g., a package of TEM, NTA, and western blotting, to identify isolated exosomes.

For example, microscopy-based methods, such as SEM and TEM, can directly visualize the surface topography and internal structure, respectively. However, TEM is not suitable for quick measurement of a large number of samples due to complicated operation and tedious sample preparation.53 NTA facilitates fast detection and real-time exosome observation while having a higher resolution than flow cytometry. The main disadvantage of NTA is its difficulty in distinguishing exosomes from contaminated proteins.54 As a mature technique, western blotting can qualitatively and quantitatively detect the expression of exosomal protein biomarkers, especially exosomes from cell culture media. However, it is time-consuming and not suitable for the detection of exosomes from biological fluids.55,56

Storage of exosomes

The currently used preservation methods for long-term storage of exosomes mainly include cryopreservation, lyophilization, and spray-drying.10 Temperature and antifreeze are the two most important ingredients for cryopreservation. Storage at 4 °C might weaken the biological activity and reduce the protein cargo of exosomes, whereas −80 °C is considered the optimal temperature causing the least impact on exosome morphology and content.57,58 Non-permeable disaccharide antifreeze, especially trehalose, represents the best choice as it prevents exosome aggregation and cryodamage.59 Heat-sensitive materials, e.g., exosomes and vaccines, treated by lyophilization of freeze-drying can be easily stored and reconstituted by simply adding water. A recent study showed that lyophilization with cryoprotectant could retain the activity of exosomal proteins and RNA for approximately 4 weeks even when stored at room temperature.60 Finally, in contrast to freeze-drying, spray-drying is a single-step process, thereby reducing the need for expensive equipment and lengthy multi-step milling. However, core parameters of spray-drying such as exosome feeding rate, atomization pressure, and outlet temperature, can all affect exosome stability and cargo integrity.61

Orthopedic and trauma surgery and SC-Exo therapy

Fracture

Fractures are the most common traumatic large-organ injuries, and approximately 10% heal improperly.62 Fracture healing involves an anabolic tissue-bulking phase and a catabolic tissue-remodeling phase, which are controlled by various factors such as stem cells, innate and adaptive immune functions, and stability.63 Biopharmacological treatment for fractures can be given locally (e.g., bone morphogenetic protein, BMP) or systemically (e.g., parathyroid hormone, PTH). As a promising alternative, exosome therapy for fracture healing mostly utilizes bone marrow-derived MSCs as a cellular supplier (Table 2).

Table 2 Stem cell-derived exosomes for the treatment of diseases in orthopedic surgery and related specialties
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The presumed mechanism of how MSC-derived exosomes promote fracture healing is as follows. Firstly, the progression of bone repair needs a variety of cells, e.g., inflammatory cells in the inflammation stage, endothelial and mesenchymal progenitor cells in the fibrovascular stage, osteoblasts and chondrocytes during bone formation, and osteoclasts during callus remodeling.62 Secondly, most of these cells can uptake exosomes, especially osteoblasts and vascular endothelial cells,64 which are most related to fracture healing. Lastly, upon exosome absorption, the gene expression of the recipient cells is modified, thereby activating various signaling pathways (Fig. 3a), causing various cellular and tissue responses (Fig. 3b) and ultimately leading to improved fracture healing.

Fig. 3
figure 3

Mechanisms of stem cell-derived exosome therapy (figure generated using Adobe Photoshop 2023 and Adobe Illustrator 2023). a activation and regulation of various signaling pathways. b disease-specific cellular and tissue responses

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Early research has employed various animal models of fracture healing. In a transverse femoral shaft fracture model, exosomes were found to not only promote osteogenesis in wild-type mice, but also rescue retardation of fracture healing in CD9−/− mice, a strain known to have a lower bone union rate.65 In a femoral nonunion model, exosomes enhanced fracture healing by promoting osteogenesis and angiogenesis possibly via the BMP-2/Smad1/RUNX2 pathway.66 In a tibial distraction osteogenesis model, exosomes secreted by young MSCs promoted osteogenic capacity of older MSCs and enhanced new bone formation in older rats.67 In addition, EPC-derived exosomes accelerated bone regeneration during distraction osteogenesis by stimulating angiogenesis.68

As a major cargo of exosomes (Section 2.1), RNA can alter recipient cell gene expression and phenotypic function, with microRNA (miRNA) and long non-coding RNA (lncRNA) being the most widely studied.69 From the perspective of an exosomal miRNA, one group discovered that miR-136-5p from bone marrow MSC-derived exosomes promoted osteoblast proliferation and differentiation in vitro, thereby promoting fracture healing in vivo.70 This was achieved by inhibiting the downstream target gene of miR-163-5p, low-density lipoprotein receptor-related protein 4 (LRP4), through the Wnt/β-catenin pathway. The other group found that MSC-derived exosomal miR-25 could regulate the ubiquitination and degradation of Runt-related transcription factor 2 (Runx2) by Smad ubiquitination regulatory factor 1 (SMURF1) to promote fracture healing in mice.71 From a lncRNA perspective, especially the bone-specific lncRNA H19, a Chinese group revealed that although a high-fat diet reduced osteogenic differentiation and weakened fracture healing, this could be reversed by MSC-derived exosomal lncRNA H19 via miR-467/HoxA10 axis in an obesity-induced fracture model.72 In addition, an American group demonstrated that exosomal lncRNA H19 not only improved osteogenesis but also angiogenesis through the angiopoietin 1/Tie2-NO signaling pathway in an immunocompromised nude mouse model.73

Instead of using naturally derived exosomes from MSCs, some researchers have conducted pre-isolation modification of exosomes to achieve better results. Liang et al. preconditioned MSCs with low doses of dimethyloxaloylglycine (DMOG), a small angiogenic molecule, to prepare the exosomes for an enhanced angiogenesis and bone regeneration in a critical-sized calvarial defect model by targeting the protein kinase B/mechanistic target of rapamycin (Akt/mTOR) pathway.74 Alternatively, Lu et al. loaded MSC-derived exosomes with miR-29a, which showed a robust ability in promoting angiogenesis and osteogenesis by targeting vasohibin 1.75 Furthermore, umbilical cord MSC-derived exosomes demonstrated comparable results to their bone marrow counterparts during fracture healing.76 In addition, exosomes derived from MSCs under hypoxia exhibited better effects on bone fracture healing than those under normoxia. Mechanistically, hypoxia preconditioning enhanced the production of exosomal miR-126 through the activation of hypoxia-inducible factor 1 (HIF-1α). Various studies have shown that hypoxia preconditioning represents an effective and promising optimization of the therapeutic effects of MSC-derived exosomes for bone fracture healing.

Osteoarthritis

Osteoarthritis (OA) is the most common joint disease and most frequent reason for activity limitation in adults, affecting approximately 240 million patients globally.77 The pathology of OA has evolved from being viewed as cartilage-only to a multi-tissue disease that affects all components of the whole joint, including bone, synovium, muscle, ligament, and periarticular fat.78 Clinical trials have successfully revealed systemic compounds that arrest structural progression (e.g., cathepsin K and Wnt inhibitors) or reduce OA pain (e.g., nerve growth factor inhibitors). As a potential treatment option for OA, most MSC-derived exosome therapy used chondrocytes as a target in in vitro models. These MSCs could originate from various tissues, such as bone marrow, synovium, gingiva, and infrapatellar fat pads (IPFPs).

Some studies focusing on chondrogenesis demonstrated a particular interest in the role of miRNA. Wu et al. found that IPFP MSC-derived exosomes protect articular cartilage from damage and ameliorate gait abnormality in OA mice by miR100-5p-regulated inhibition of mTOR-autophagy pathway.79 Since it is easy to retrieve human IPFP from OA patients by arthroscopic operation within a clinic, this type of exosome therapy might simplify and accelerate the process from bench to bedside. Liu et al. discovered that MSC-derived exosomes could promote proliferation and inhibit apoptosis of chondrocytes via lncRNA-KLF3-AS1/miR-206/GIT1 axis in OA.80 The cellular work conducted by Kong et al. showed that synovial MSC-derived exosomal miR-320c could enhance chondrogenesis by targeting ADAM19.81 In addition, Mao et al. suggested that exosomal miR-92a-3p from chondrogenic MSCs could enhance chondrogenesis and suppress cartilage degradation via targeting Wnt5a.82 In contrast to these studies using original exosomes, few groups modified exosomes prior to their systemic administration. Tao et al. modified exosomes by transfecting synovial MSCs with miR-140-5p and found that exosomal miR-140-5p-overexpression could enhance cartilage tissue regeneration and prevent OA of the knee in a rat model.83 Meanwhile, Wang et al. used TGF-β1 to stimulate MSCs, and the resultant exosomal miR-135b increased chondrocyte proliferation by regulating specificity protein-1.84 In a comparative study, Zhu et al. demonstrated that exosomes from iPSC-derived MSCs could provide a stronger therapeutic effect on OA than synovial membrane MSC-derived exosomes.85

Other studies have focused on not only chondrogenesis but also anti-inflammation and immune modulation during OA treatment. For example, MSC-derived exosomes inhibited inflammatory factors, glutamine metabolic activity-related proteins, glutamine, and GSH/GSSG ratio in vitro, while improving mice’s chondrocyte function, tissue inflammation, and exercise capacity in vivo, thereby alleviating OA progression.86 Using a holistic approach, recent studies have shifted the attention away from cartilage towards other tissues (e.g., bone) in a diarthrodial joint. Firstly, bone marrow MSC-derived exosomal miR-206 promoted proliferation and osteogenic differentiation of osteoblasts in OA by reducing E74-like factor 3 (Elf 3), and ameliorated inflammation and increased expression of osteocalcin and BMP2 in mouse femoral tissues.87 Secondly, MSC exosome-treated osteochondral defects demonstrated a regenerative immune phenotype, characterized by a higher infiltration of CD163+ M2 macrophages over CD86+ M1 macrophages, with a concomitant reduction in pro-inflammatory synovial cytokines IL-1β and TNF-α.88 Lastly, gingival MSC-derived exosomes proved to be immunosuppressive in preventing collagen-induced arthritis.89 Compared with parental cells, these exosomes had the same or stronger effects in inhibiting IL-17A and promoting IL-10, reducing incidences and bone erosion by arthritis, via inhibiting the IL-17RA-Act1-TRAF6-NF-κB signaling pathway.

Currently, there is no single ‘one size fits all’ drug that may be suitable for all OA patients. Disease-modifying OA drugs (DMOADs) might become the next-generation OA treatment.90 It is very valuable and relevant that MSC-derived exosome therapy for OA coincides with DMOADs: both are capable of targeting inflammatory cytokines, matrix-degrading enzymes, and the Wnt pathway. Thus, emerging approaches for DMOAD development, such as miRNA-based modality and targeting cellular senescence, might also be used to refine MSC-based exosome therapy for OA.

Spinal cord injury

Traumatic spinal cord injury (SCI) is a devastating global health issue that poses a significant functional and economic burden both on the patient and society.91 The pathophysiology of SCI includes primary injuries caused by mechanical trauma and secondary injury cascade characterized by apoptosis, edema, ischemia, inflammatory cell infiltration, and excitotoxicity.92 Despite surgical intervention, clinical studies involving pharmacotherapy can be broadly classified as either neuroprotective or neuroregenerative.93 Targeting each event of the above mechanistic chain, both MSC- and NSC-derived exosome therapy could exert a beneficial influence on spinal cord protection and regeneration.

Some groups have targeted neuronal cell death. Ma et al. revealed that insulin-like growth factor 1 (IGF-1)-stimulated NSC-derived exosomes could inhibit neuronal apoptosis while promoting functional recovery after SCI through a miR-219a-2-3p/YY1 pathway.94 Alternatively, Zhang et al. discovered that subarachnoid injection of NSC-derived exosomes could suppress neuronal cell apoptosis by activating autophagy via miR-374-5p/STK-4 axis for enhanced functional recovery in SCI.95 Shao et al. explored other forms of cell death (e.g., ferroptosis) using MSC-derived exosomes, and found that exosomal lncGm36569 could inhibit neuronal cell ferroptosis via miR-5627-5p/FSP1 axis, thereby decreasing neuronal dysfunction.96

Some groups have targeted anti-inflammation and immunomodulation. Nakazaki et al. discovered that fractionated intravenous infusion of MSC-derived exosomes could target M2 macrophages and upregulate TGF-β, thereby stabilizing microvessels and improving functional recovery.97 Similarly, Liu et al. demonstrated that in addition to hypoxia increasing exosome production from bone marrow MSCs, preconditioned exosomal miR-216a-5p could also repair traumatic SCI by shifting microglial M1/M2 polarization via the TLR4/NF-κB/PI3K/Akt pathway.98 Huang et al. valuably proved that epidural fat MSC-derived exosomes could attenuate NLRP3 inflammasome and improve functional recovery in SCI.99 Compared to MSC-derived exosomes, exosomes derived from EPCs could provide comparable anti-inflammatory effect. Yuan et al. showed that the exosomal miR-222-3p from EPCs could promote anti-inflammatory macrophages via the SOC3/JAK2/STAT3 pathway and improve mouse functional repair after SCI.100

Some groups have targeted angiogenesis and blood-spinal cord barrier (BSCB) integrity. For example, Zhong et al. used unmodified NSC-derived exosomes, found that they were highly enriched in VEGF-A, and could therefore enhance the angiogenic activity of spinal cord microvascular endothelial cells (SCMECs).101 In comparison, Chen and co-workers modified NSC-derived exosomes with FTY720, an immune modulator and microvascular regulator, to protect the barrier function of SCMECs via the PTEN/Akt pathway, thereby ameliorating hindlimb function.102 It is well-known that the connection between the microvascular endothelium of the spinal cord and the pericyte is crucial in maintaining the structural integrity of BSCB.103 Thus, Zhou’s team attempted to verify the role of exosome therapy in pericyte homeostasis.104 They proved that bone marrow MSC-derived exosomes could reduce pericyte pyroptosis and increase pericyte survival rate in vitro, while improving BSCB integrity and locomotor recovery in vivo.

Finally, some groups have targeted other aspects during neuroprotection and neuroregeneration, such as neurotoxic astrocytes and endogenous NSC sustainability. Lai et al. proved that human umbilical cord MSC-derived exosomes could facilitate recovery of spinal cord function by targeting neurotoxic astrocytes.105 In addition, miR-146a-5p-modified exosomes exerted a more powerful effect than unmodified exosomes. Li et al. discovered that exosomes derived from nerve growth factor (NGF)-overexpressing bone marrow MSCs could enhance neuronal differentiation of NSCs and axonal regeneration.106 Zhou et al. demonstrated that placental MSC-derived exosomes could promote the activation of proliferating endogenous NSCs, thereby improving both locomotor activity and bladder dysfunction,107 which is a frequent sequelae that could further worsen the quality of life of SCI patients.108

Muscle and tendon tear

Muscle and tendon tears can result from either acute trauma (e.g., fractures, Section 3.1) or chronic overuse (e.g., sports injury).109 Healing of muscle strain and tendon tear follows the typical wound healing course, involving the inflammatory, proliferative, and remodeling phases. Multiple non-surgical strategies have been trialed to improve healing, including cell-based and growth factor-based therapies.110 The following proof-of-concept studies indicate that MSC-derived exosomes could become the next-generation musculoskeletal treatment.

On the one hand, some groups have focused on individual components of the muscle-tendon-bone unit. Nakamura et al. claimed that MSC-derived exosomes could improve in vitro myogenesis in C2C12 myoblasts and angiogenesis in HUVECs, while accelerating in vivo skeletal muscle regeneration in a cardiotoxin-induced muscle injury model.111 These benefits were at least in part mediated by miRNAs such as miR-494. Chen et al. discovered that exosomes from adipose MSCs could enhance the proliferation and the migration of primary tenocytes, while also improving mechanical strength of repaired tendons by upregulating decorin and biglycan in a rabbit Achilles tendon rupture model.112

On the other hand, some groups have regarded the muscle-tendon-bone unit as a single functional system and used rotator cuff tear as the disease model, which is the most common shoulder condition for which patients seek treatment.113 One group of researchers published two consecutive studies using adipose MSC-derived exosomes. In a rat model of massive rotator cuff tear, exosome therapy could prevent the atrophy, inflammation, and vascularization of muscles.114 In a rabbit model of chronic rotator cuff tear, exosome therapy could prevent fatty infiltration and improve biomechanical properties.115 Another group reported that bone marrow MSC-derived exosomes could increase the breaking load and stiffness of the rotator cuff after reconstruction, induce angiogenesis around the rotator cuff endpoint, and promote growth of the tendon-bone interface.116

Other orthopedic diseases

Osteoporosis is a metabolic bone disease characterized by low bone density and weakening of bone architecture, which increase the risk of fractures. It results from osteoclastic bone resorption undercompensated by osteoblastic bone formation.117 In a cellular study, adipose MSC-derived exosomes could antagonize hypoxia/serum deprivation-induced osteocyte apoptosis and osteocyte-mediated osteoclastogenesis.118 Further animal studies revealed that umbilical cord MSC-derived exosomes could inhibit bone marrow MSC apoptosis and prevent disuse osteoporosis via miR-1263/Mob1/Hippo pathway,119 and improve tibial density and reverse estrogen-deficient osteoporosis via miR-2110 and miR-328-3p.120

Compared to SCI, damage to peripheral nerve (e.g., sciatic nerve injury) is considerably more common. The subsequent nerve regeneration is controlled by the interplay between neurons and Schwann cells, and further complicated by inflammatory cell infiltration.121 It was shown that adipose MSC-derived exosomes could target neurons by increasing neurite outgrowth in vitro and axonal regeneration and walking behavior in vivo.122 Adipose MSC-derived exosomes could target Schwann cells by promoting the proliferation, migration and secretion of neurotrophic factors in vitro and restore denervation muscle atrophy in vivo.123 LPS-preconditioned MSC-derived exosomes could target inflammatory cells by enhancing M2 macrophage polarization in vitro and accelerate peripheral nerve regeneration in vivo.124

Intervertebral disc (IVD) degeneration is a major cause of lower back pain which is the leading injury in total global years lived with disability. Its molecular processes include extracellular matrix (ECM) degeneration, inflammation, oxidative stress, apoptosis, senescence and reduced autophagy.125 The emerging avenues of exosome therapy attempt to solve some of these issues. Cheng et al. demonstrated that intradiscal injection of bone marrow MSC-derived exosomes could inhibit nucleus pulposus cell (NPC) apoptosis and alleviate IVD degeneration via exosomal miR-21.126 On the other hand, Chen et al. discovered that human ESC-derived exosomes could inhibit NLRP3 inflammasome to alleviate pyroptosis in nucleus pulposus cells by delivering miR-302c.127 In addition to cell death and mitochondrial damage, oxidative stress in NPCs was also found to be inhibited by MSC-derived exosomes.128 Since IVD degeneration and OA share a common molecular disease spectrum125 the positive results of OA treatment using MSC-derived exosomes (Section 3.2) could be used as a reference for IVD degeneration research.

Osteonecrosis, aka., avascular necrosis, of the femoral head (ONFH) is a disabling condition affecting a younger population, which often results in total hip arthroplasty.129 Glucocorticoid (GC)-induced osteonecrosis is one of the most common causes of ONFH, whose pathogenesis is manifested in two aspects: compromised blood supply to the femoral head and dampened osteogenic activity. Liu et al. showed that exosomes from iPSC-derived MSCs could prevent GC-induced ONFH by promoting angiogenesis and osteogenesis via the PI3K/Akt pathway.130 Zuo et al. demonstrated that miR-26a-overexpressing exosomes derived from HSCs could provide similar therapeutic effects.131

Neurosurgery and SC-Exo therapy

Ischemic stroke

Strokes are the second highest cause of death and the third leading cause of disability globally, with ischemic stroke being the most common subtype.132 The key events during the ischemic cascade include neuronal dysfunction, excitotoxicity, neurochemical injury, and neuroinflammation.133 In terms of treatment, a new generation of clinical trials is now underway, which uses cytoprotective drugs, such as immunomodulators, IL-6 receptor antagonists, Rho kinase inhibitors, and free radical scavengers.134 Targeting each event of the above pathophysiology, nearly all subtypes of SC-exo demonstrated potent therapeutic effects on stroke recovery (Table 3).

Table 3 Stem cell-derived exosomes for the treatment of diseases in neurosurgery and related specialties
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Some groups have targeted neuroprotection and neurogenesis. Firstly, SC-exo therapy could inhibit neuronal cell death. Luo et al. found that NSC-derived exosomes could inhibit apoptosis while promoting the proliferation of SH-SY5Y cells both under normal and oxygen-glucose deprivation (OGD) conditions.135 This was also tested in a middle cerebral artery occlusion (MCAO) model as a reduced infarction area and neuronal apoptosis via exosomal miR-150-3p. Other in vitro and in vivo studies showed similar anti-apoptotic effects using EPC-derived exosomes.136,137 Zhang et al. discovered that the exosomal anti-apoptotic effect could be improved by preconditioning the parental NSCs with interferon gamma (IFN-γ).138 Secondly, SC-exo therapy could protect cells of the CNS. Kang et al. revealed that exosomes derived from bone marrow MSCs could rescue OGD-induced injury in neural cells by suppressing NLRP3 inflammasome-mediated pyroptosis.139 Exosomes sourced from hypoxic cultures had a more pronounced neuroprotective effect than their counterparts from normal cultures. Similarly, Li et al. discovered that exosomes derived from human iPSC-derived neural progenitor cells exhibited a neuroprotective effect on OGD neurons and neurite outgrowth.140 This protection of neuronal function under ischemic conditions was regulated through the PTEN/Akt pathway. In addition, Sun et al. proved that NSC-derived exosomes could also protect astrocytes, which become supporting reactive astrocytes (RAs) after strokes.141 Thirdly, SC-exo therapy could improve post-stroke neurogenesis. Wei et al. suggested that Zeb2/Axin2 from bone marrow MSC-derived exosomes could improve post-stroke neurogenesis, neural plasticity, and spatial memory and nerve function, likely via the SOX10, Wnt/β-catenin, and endothelin-3/EDNRB pathways.142 Wang et al. illustrated that miR-126-modified EPC-derived exosomes could alleviate acute brain injury and promote functional recovery after stroke by enhancing neurogenesis.143

Some groups have targeted the inhibition of the neuroinflammation. Firstly, unmodified SC-exo therapy exhibited an anti-inflammatory effect through exosomal miRNAs. Dong et al. showed that bone marrow MSC-derived exosomes could induce BV2 microglia deactivation and M2 polarization in vitro, while reducing infarct size and improving neuronal function in vivo via transferring miR-23a-3p.144 Similarly, Zhang et al. unveiled that umbilical cord MSC-derived exosomal miR-146a-5p could attenuate microglia-mediated neuroinflammation after OGD in vitro, while improving behavioral deficits and microglia activation in vivo via the IRAK1/TRAF6 signaling pathway.145 Secondly, the anti-inflammatory effect of SC-exo therapy could be enhanced by modifying the exosomes. Yoon and co-workers established tumor susceptibility gene (TSG)101-overexpressing human NSCs, thereby increasing exosome secretion.146 The engineered exosomes not only attenuated LDH release and proinflammatory factors in vitro, but also reduced infarction volume, inhibited DNA-damage pathway, and upregulated neurotrophic factors in vivo. Furthermore, Tian’s team broke new ground by ingeniously attaching RGD peptide onto an NSC-derived exosome membrane, which targeted the lesion region of the ischemic brain after intravenous administration, thereby suppressing the inflammatory response after cerebral ischemia by inhibiting the MAPK pathway.147 Interestingly, Gao et al. used induced NSCs (iNSCs) reprogrammed from mouse fibroblasts for stroke treatment. They showed that iNSC-derived exosomes, bearing similar therapeutic effects with NSC-derived ones, could not only promote neurogenesis but also inhibit neuroinflammation.148

Finally, some groups have targeted other aspects during stroke recovery, such as neurochemical injury and oxidative stress. Zhu et al. loaded brain-derived neurotrophic factor (BDNF) into exosomes derived from NSCs to construct engineered exosomes.149 In a model of H2O2-induced oxidative stress, exosome therapy significantly enhanced NSC survival. In a rat MCAO model, exosome therapy not only inhibited microglial activation, but also boosted the differentiation of endogenous NSCs into neurons. Collectively, BDNF-based modification of NSC-derived exosomes has improved effects in the treatment of ischemic stroke. On the other hand, miR-210-modified EPC-derived exosomes could protect neurons from hypoxia and reoxygenation (H/R)-induced apoptosis, oxidative stress, and decreased viability, thereby supporting the treatment of ischemic stroke.150,151 The exosomal miR-17-5p from ACE2-enriched EPC-derived exosomes could ameliorate cerebral ischemic injury in aged mice.152 In an intriguing study conducted by Xu and co-workers, combination of NSC-exo and EPC-exo with miR-210 and miR-123 overexpression exerted better therapeutic effects on ischemic stroke by protecting H/R injured neurons through the BDNF-TrkB and Nox2/ROS pathways.153

In contrast to ischemic stroke, hemorrhagic stroke poses a deadlier threat and worse disability in most survivors.154 miR-137 overexpression was found to boost the neuroprotective effects of EPC-derived exosomes against apoptosis, ferroptosis, and mitochondrial dysfunction in oxyhemoglobin-treated SH-SY5Y cells, an in vitro hemorrhagic stroke model, partially through the COX2/PGE2 pathway.155

Traumatic brain injury

Approximately 70 million patients suffer from traumatic brain injury (TBI) globally each year, which poses serious physical, psychosocial and economic threats.156 TBI can be categorized as primary injuries (e.g., axonal death, neuroinflammation, neurochemical change, and metabolic dysfunction) and secondary injuries (e.g., ischemic and hypoxic damage, cerebral edema, raised intracranial pressure, hydrocephalus, and infection).157 Each patient with a TBI has a unique set of circumstances depending on variables such as the location and severity of the injury, making medical and surgical treatment quite challenging.158 Therefore, systemic therapy using SC-exo may become a ‘one-size-fits-all’ option for managing TBI.

A series of animal studies published initially focused on the functional recovery and macroscopic aspects of MSC-derived exosome therapy. In a rat TBI model, exosome-treated animals showed significant improvement in spatial learning and sensorimotor function.159 In addition, exosome treatment significantly increased the number of newborn endothelial cells in the lesion boundary zone, and newborn immature and mature neurons in the dentate gyrus. In another rat TBI model with similar findings, exosomes derived from MSCs cultured in a 3D system provided better outcomes than those in a conventional 2D condition.160 In a monkey model of TBI to the primary motor cortex, exosome-treated animals returned to pre-operative grasp patterns and latency to retrieve a food reward in the first 3–5 weeks of recovery.161 In an even more complicated and clinically realistic large animal model, in which both TBI and hemorrhagic shock were investigated, exosome therapy attenuated the severity of neurologic injury and enabled faster neurologic recovery.162

In comparison, studies completed in recent years shed new light on the molecular mechanisms underlying SC-exo therapy for TBI. Chen et al. reported that adipose MSC-derived exosomes could promote functional recovery, suppress neuroinflammation, reduce neuronal apoptosis, and increase neurogenesis. This was achieved through the uptake of exosomes specifically by microglia and suppression of their activation by inhibiting the NF-κB & MAPK pathways.163 Wen et al. showed that bone marrow MSC-derived exosomes could reduce cell apoptosis in cortical tissue of mouse models of TBI, inhibit neuroinflammation, and promote the transformation of microglia to the anti-inflammatory phenotype. This was realized by the action of miR-181b on the IL-10/STAT3 pathway.164 Abedi et al. proved that NSC-derived exosomes could improve neurobehavioral performance, inhibit astrocyte neuroinflammation, enhance neurogenesis, while maintaining NSC stemness.165 A valuable additional finding was that exosomes seemed to be superior to the parent NSCs in terms of sensorimotor functional recovery. Finally, a dose-response and therapeutic window demonstrated that MSC-derived exosomes could improve angiogenesis and neurogenesis, and sensorimotor and cognitive function, while reducing neuroinflammation and hippocampal neuronal cell loss.166 Although 100 µg and 1 day might be the optimal dose and therapeutic window respectively, exosomes exhibited a wide range of effective doses for treatment of TBI within a therapeutic window of at least 7 days post-injury.

TBI and SCI are two of the most severe CNS traumas, which are increasingly recognized as global health priorities. The emerging evidence presented in Sections 3.3 and 4.2 are mutually beneficial for these two closely related research subspecialties. Henceforth, future research on SC-exo therapy for TBI and SCI could be either mechanism-based (e.g., the role of brain-gut axis167 transcriptional factors168 inflammasome169 and the complement system170) or modification-based (loading exosomes with drugs, e.g., immunomodulators171 antioxidants172 circular RNAs173 and microRNAs174).

Alzheimer’s disease

Unlike TBI and SCI, which are traumatic in nature, Alzheimer’s disease (AD) and Parkinson’s disease (PD) are the most common neurodegenerative diseases (NDD). The hallmarks of NDD include, but are not limited to, pathological protein aggregation, synaptic and neuronal network dysfunction, aberrant proteostasis, cytoskeletal abnormalities, altered energy homeostasis, DNA and RNA defects, inflammation, and neuronal cell death.175 AD is the most common form of dementia globally and accounts for 25 million cases.176 Currently, only two classes of drugs are approved for symptomatic AD treatment, including cholinesterase inhibitors and NMDA antagonists. Although several therapeutics are actively undergoing clinical trials, none of them are near curative for AD.177 The challenges of brain-drug delivery, e.g., the blood-brain barrier (BBB) and pharmacokinetic drawbacks, are very likely to be solved by nanosized exosomes, which are additionally packaged with potent biomolecules. Most SC-exo therapy involves amyloid-β (Aβ), which is positioned at the center of AD pathophysiology.178

The initial work focused on the clearance of aggregation of the pathological protein, Aβ peptide. The intracerebral injection of MSC-derived exosomes by Elia and co-workers reduced Aβ plaque burden and dystrophic neurites in both the cortex and hippocampus in the early stages of a preclinical model of AD.179 In addition, using immunoblotting, the authors confirmed the presence of Neprilysin, a neutral endopeptidase capable of Aβ degradation, in the exosome’s lysates and its mRNA.

Some teams have focused on relieving synaptic dysfunction and oxidative stress. Wang et al. found that MSC-derived exosomes could rescue synaptic impairment and improve cognitive behavior in APP/PS1 mice, while alleviating exogenous Aβ-induced inducible nitric oxide synthase (iNOS) expression.180 Instead of using MSC-derived exosomes, Li et al. administered NSC-derived exosomes and enhanced mitochondrial function, sirtuin 1 activation, synaptic activity, and rescued cognitive deficits.181 Using alternative methods, Huber et al. noticed that heat shock-induced exosomes derived from NSCs exhibited greater neuroprotection against oxidative stress as well as Aβ-induced neurotoxicity.182

Some teams have centered their research around energy homeostasis. Chen et al. found that MSC-derived exosomes could improve brain glucose metabolism and cognitive function in AD transgenic mice using 18F-FDG PET/CT imaging and NOR testing, respectively.183

Some teams have focused on microglial neuroinflammation. In Zavatti’s cellular study, it was found that amniotic fluid MSC-derived exosomes could mitigate neuroinflammatory microglial phenotype and recover neurotoxicity from Aβ using LPS-stimulated BV2 microglia and SH-SY5Y neuroblastoma cells as models, respectively.184 Ding et al. showed that umbilical cord MSC-derived exosomes could alleviate neuroinflammation and reduce Aβ deposition by modulating microglial activation, thereby increasing spatial learning and memory function in AD mice.185

Some teams have focused on neuronal cell death and neurogenesis hoping to counteract AD progression. Reza-Zaldivar and co-workers gave MSC-derived exosomes to AD mice and the SC-exo therapy stimulated neurogenesis in the subventricular zone and alleviated Aβ-induced cognitive impairment.186 These effects are comparable to those shown in the MSCs.

Some teams have focused on the BBB, the dysfunction of which leads to increased permeability, microbleeds, impaired glucose transport, and degeneration of pericytes and endothelial cells.187 Liu et al. indicated that BBB breakdown in 5xFAD (familial Alzheimer’s disease) mice occurred at 4 months of age, and more importantly, treatment with NSCs-derived exosomes reversed AD-caused BBB deficiency.188

Finally, some groups have focused on improving the technicality of SC-exo therapy for AD. When exosomes are injected intravenously, they could be tracked in other organs instead of the targeted regions in the brain. Cui et al. conjugated MSC-derived exosomes with CNS-specific rabies viral glycoprotein (RVG) to target them to the brain of transgenic AD mice.189 The modified exosomes not only exhibited increased delivery to the cortex and hippocampus, but also significantly improved learning and memory capabilities with reduced Aβ deposition. On the other hand, Gao et al. obtained iNSCs through somatic cell reprogramming, which opened a new window for sourcing therapeutic exosomes. They demonstrated that iNSC-derived exosomes, bearing comparable therapeutic effects with NSC-derived ones, could mitigate various AD phenotypes, e.g., cognitive function, Aβ deposition, neuroinflammation, and neuroregeneration, in a preclinical mouse model.190

Parkinson’s disease

Parkinson’s disease is the second most common neurodegenerative disease among the elderly, affecting more than 6 million patients worldwide.191 PD is caused by the necrosis of dopaminergic neurons in the substantia nigra and the presence of protein inclusions named Lewy bodies. The molecular pathophysiology includes α-synuclein proteostasis, mitochondrial dysfunction, oxidative stress, calcium imbalance, and neuroinflammation.192

In a study using bone marrow MSC-derived exosomes, Huang et al. discovered that exosome-derived TNF-stimulated gene-6 (TSG-6) could attenuate 1-methyl-4-phenylpyridinium ion (MPP + , metabolite of a neurotoxin MPTP)-induced neurotoxicity. In this in vitro PD model using SH-SY5Y and SK-N-SH cells, the exosomal anti-PD progression effect was found to be mediated through the STAT3/miR-7/NEDD4 axis.193

In a study using NSC-derived exosomes, Lee et al. revealed that SC-exo therapy could help to prevent the neuropathology and progression of PD.194 Working in vitro on SH-SY5Y and BV2 cells, NSC-derived exosomes could reduce the intracellular reactive oxygen species (ROS) and associated apoptotic pathways. Working in vivo on 6-hydroxydopamine-induced PD mice, NSC-derived exosomes could downregulate pro-inflammatory factors and significantly reduce dopaminergic neuronal loss. The presence of NSC-specific microRNAs, such as miR-182-5p, miR-183-5p, miR-9 and let-7, was confirmed and found to be involved in cell differentiation, neurotrophic function, and immune modulation.

Multiple sclerosis

Multiple sclerosis (MS) is the most common non-traumatic, neurodegenerative, and disabling CNS disease affecting young adults. The pathological hallmark of MS is the formation of demyelinating lesions in the brain and spinal cord, with an inflammatory and autoimmune involvement.195 Currently licensed disease-modifying therapies include interferon-based, immunomodulatory, immunosuppressive, and immune reconstitution drugs.196 A few preliminary studies have highlighted the potential of MSC-derived exosomes for MS treatment.

In an animal experiment using experimental autoimmune encephalomyelitis (EAE) rats, Li et al. showed that SC-exo therapy significantly decreased neural behavioral scores, reduced the infiltration of inflammatory cells into the CNS, and decreased demyelination.197 In addition, exosome treatment upregulated M2-related cytokines while downregulating M1-related ones by regulating the polarization of microglia.

In an animal study using two mouse models of demyelination (the EAE model and the cuprizone diet model), Zhang et al. found that SC-exo therapy could promote remyelination by acting both directly on oligodendrocyte (OL) progenitors and indirectly on microglia.198 MSC-derived exosomes could improve neurological outcomes, increase the numbers of newly generated and mature OLs, decrease Aβ precursor protein density, decrease neuroinflammation by shifting from M1 to M2 phenotype, and inhibit the TLR2/IRAK1/NF-κB pathway.

Other neurosurgical and related diseases

The surgical potential of MSC- and NSC-derived exosome therapy in four major types of neurosurgical or neurological diseases has been thoroughly discussed above. In addition to vascular disruption-, trauma-, neurodegeneration-, and autoimmune-related disorders, other diseases have been proven suitable targets for SC-exo therapy recently (Table 3). These include, but are not limited to: 1, dementia, such as vascular dementia.199 HIV-associated neurocognitive disorders200 and radiation-induced cognitive dysfunction201,202; 2, functional disorders, such as epilepsy203 and mechanical allodynia204; 3, congenital abnormalities, such as spina bifida aperta205; 4, neuropsychological conditions, such as depression206 and stress207; 5, brain aging208,209,210; 6, iatrogenic brain problems, such as deep hypothermic circulatory arrest.211

Plastic surgery and SC-Exo therapy

Wound healing occurs in all parts of the human body, with cutaneous wounds being the most common. The highest wound-related expenses were attributed to surgical wounds followed by diabetic ulcers.212 The overall but overlapping phases of wound healing include hemostasis, inflammation, angiogenesis, proliferation and remodeling, each of which is governed by distinct cell types and modulated by various signaling pathways.213 More than half of relevant work using SC-exo therapy to boost cutaneous wound healing is MSC-based (Table 4).

Table 4 Stem cell-derived exosomes for the treatment of diseases in plastic surgery and related specialties
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In the inflammatory stage, exosomes could inhibit the proliferation of peripheral blood mononuclear cells and promote the transformation of regulatory T cells in vitro, and reduce the number of lymphocytic infiltrations in the skin.214 In addition, exosomes could reduce IgE, eosinophil and mast cell count, and downregulate inflammatory cytokines.215 In the angiogenic stage, educated exosomes (e.g., atorvastatin and deferoxamine) could promote angiogenesis in diabetic wounds via the Akt/eNOS and PTEN/PI3K/Akt pathways.216,217,218 EPC-derived exosomes could accelerate cutaneous wound healing by promoting angiogenesis219 through the Erk1/2 pathway220 and p53 pathway.221 In the proliferative stage, stem cell-derived exosomes could promote the proliferation and migration of fibroblasts and keratinocytes. Some were achieved through the PI3K/Akt222 Akt/HIF-1α223 ERK1/2224 and Wnt/β-catenin225 pathways, while others through inhibition of LATS2226 PPARγ227 and AIF nucleus translocation228 In the final remodeling stage of wound healing, granulation tissue is replaced by permanent scar, during which abnormal wound healing might occur (e.g., keloids and hypertrophic scars). MSC-derived exosomes could suppress fibroblast-myofibroblast transition via the TGF-β/Smad2 pathway229,230 and increase collagen synthesis in early stage and reduce in late stage231 thereby reducing scar formation.

Furthermore, ESC-derived exosomes were found to exert similar therapeutic effect for wound healing to MSC-derived ones. Chen et al. used human ESC-derived exosomes to help healing of pressure ulcer.232 They noticed that exosomes could ameliorate endothelial senescence by activating Nrf2 and recover aging-related angiogenic dysfunction, thereby accelerating wound healing.232 In addition, Bae et al. revealed that the exosomal mmu-miR-291a-3p from ESCs could inhibit cellular senescence in human dermal fibroblasts through the TGF-β receptor 2 pathway, thereby accelerating the excisional skin wound healing process.233

In addition to wound healing, other plastic surgery-related diseases have been proven to be suitable targets for SC-exo therapy (Table 4). These include, but are not limited to: 1, skin grafting, such as skin flaps234; 2, tissue loss, such as craniofacial defect235; 3, autoimmune skin diseases, such as scleroderma236; 4, skin infections, such as leishmaniasis237; 5, hair transplantation, such as for alopecia238; 6, skin aging239

General surgery and SC-Exo therapy

As a major subspecialty of general surgery, hepatobiliary surgery has attracted tremendous attention to SC-exo therapy. Firstly, acute liver injury (ALI)/acute liver failure (ALF) is a rare but challenging syndrome manifested by hepatic dysfunction, coagulopathy, encephalopathy, and multiorgan failure. About 60% of cases with ALF require and undergo orthotopic liver transplantation or result in death.240 In one study, Lin’s team focused on the cell death aspect of ALI, and found that MSC-derived exosomes could protect against ferroptosis via stabilization of SLC7A11 in carbon tetrachloride-induced ALI.241 Alternatively, Shao’s team focused on the pre-isolation modification of the exosomes, and revealed that exosomes derived from umbilical cord MSCs could ameliorate IL-6-induced ALI through exosomal miR-455-3p.242 Secondly, in contrast to ALI, liver fibrosis occurs when the liver sustains a chronic injury, which may progress into cirrhosis, liver failure, hepatocellular carcinoma, and even death.243 Ma et al. discovered that MSC-originated exosomal circDIDO1 could suppress hepatic stellate cell activation by miR-141-3p/PTEN/Akt pathway in human liver fibrosis.244 In addition, Wang et al. found that exosomes derived from 3D human ESC spheroids could attenuate hepatic stellate cell activation and inhibit liver fibrosis through inactivation of the Smad pathway by exosomal miR-6766-3p.245 For those with end-stage liver fibrosis needing a liver transplant, liver ischemia reperfusion injury (IRI) is a serious complication for graft dysfunction and organ rejection.246 Yang et al. demonstrated that bone marrow MSC-derived exosomes could relieve hepatic IRI, reduce hepatocyte apoptosis, and decrease liver enzyme levels by enhancing autophagy.247 Du et al. showed that exosomes from iPSC-derived MSCs could protect liver against hepatic IRI via activating sphingosine kinase and the sphingosine-1-phosphate pathway.248 Thirdly, non-alcoholic fatty liver disease (NAFLD) is known to adversely affect stroke recovery. Using a type 2 diabetes mellitus mouse model, Venkat et al. demonstrated that HSC-derived exosomes could simultaneously reduce liver dysfunction and improve neurological and cognitive function.249 Lastly, acute pancreatitis is an unpredictable and potentially lethal disease, the prognosis of which mainly depends on whether it develops into multiple organ dysfunction syndrome.250 Chen et al. revealed that exosomes from iPSC-derived MSCs could improve myocardial injury caused by severe acute pancreatitis through the Akt/Nrf2/HO-1 pathway.251

Peripheral artery disease affects 200 million patients worldwide and, in its most severe stage, can cause critical limb ischemia, subjecting patients to increased risk of cardiovascular events, amputation and death.252 As a cell-free therapy, placenta MSC-derived exosome infusion could enhance angiogenesis in a murine auricle ischemic injury model using laser Doppler blood flow analysis.253 Mechanistically, MSC-derived exosomes not only promote tube-like structure formation in vitro, but also mobilize endothelial cells into subcutaneous Matrigel plug in vivo, mainly through exosomal pro-angiogenic microRNAs, such as miR-30b.254 In addition, HSC-derived exosomes could repair ischemic hindlimb in mice by improving limb perfusion, capillary density, motor function and their amputation.255 This was most likely caused by internalization of exosomal miR-126-3p by endothelial cells relative to smooth muscle cells and fibroblasts. Human iPSC-derived exosomes demonstrated similar neoangiogenic effect through the exosomal miR-199b-5p.256 On the other hand, endovascular re-canalization is increasingly being used to reestablish blood flow to ischemic areas and restore tissue loss or gangrene for patients with peripheral artery disease.257 Three independent teams all proved that EPC-derived exosomes could promote vascular repair and accelerate reendothelialization in rat models of balloon-induced vascular injury by enhancing endothelial cell function.258,259,260 In addition, Kong et al. demonstrated similar protective effect of EPC-derived exosomes against balloon injury by inhibiting neo-intimal hyperplasia.261 This was achieved through promotion of reendothelialization and suppression of restenosis rather than through the direct inhibition of proliferation and migration of smooth muscle cells.

McCulloh and his co-workers published an intriguing study on the SC-exo therapy for necrotizing enterocolitis (NEC) which has an overall mortality of over 30% for premature infants requiring surgery.262 The authors compared the therapeutic effect of exosomes derived from four different types of stem cells, i.e., amniotic fluid MSCs, bone marrow MSCs, amniotic fluid NSCs and neonatal enteric NSCs.263 When injected at a concentration of at least 4 × 108, all types of SC-exo were shown to reduce the incidence and severity of experimental NEC as effectively as their parental stem cells.

Sepsis is a deadly and potentially preventable complication in general surgery, in which microvascular dysfunction leads to multi-organ failure and mortality.264 Using a murine sepsis model by cecal ligation and puncture (CLP), Zhou and co-workers demonstrated that EPC-derived exosomes could improve sepsis outcome.265 This was manifested by reduced lung and renal vascular leakage, improved organ function, and increased survival through the exosomal miR-126-5p and miR-126-3p. Similarly, Liu et al. exhibited protective effect of EPC-derived exosomes on sepsis-induced organ damage and immune suppression by the exosomal miR-382-3p through the IκBα/NF-κB pathway.266

Cardiothoracic surgery and SC-Exo therapy

The world’s leading mortality is ischemic heart disease (IHD) which is primarily caused by obstructive coronary atherosclerosis. The rupture of an atherosclerotic plaque is the most common trigger of acute arterial thrombosis causing myocardial infarction (MI). Prolonged oxygen deprivation to the myocardium can lead to cardiomyocyte death. Although timely reperfusion is essential, myocardial IRI might occur, thus mitigating the beneficial effects of reperfusion. Despite modern coronary reperfusion, the mortality and morbidity associated with the development of heart failure as a consequence of acute MI remain substantial, highlighting the importance of next-generation cardioprotective therapies, such as SC-exo.267

The work conducted by Xing et al. focused on atherosclerosis. The adipose MSC-derived exosomal miR-342-5p was shown to protect endothelial cells against atherosclerosis by targeting PPP1R12B in a H2O2-challenged HUVEC model.268 The work conducted by Peng et al. and Gao et al. focused on MI. The exosomal miR-25-3p from MSCs could alleviate MI by targeting pro-apoptotic proteins and EZH2 in an OGD cardiomyocytes model and left anterior descending artery ligation animal model.269 Similarly, human iPSC-derived exosomes could improve recovery from MI without increasing the frequency of arrhythmogenic complications in a swine model.270 The work conducted by Wen et al. and Santoso et al. focused on the death of cardiomyocytes. MSC-derived exosomes could ameliorate cardiomyocyte apoptosis in hypoxic conditions through miR-144 by targeting the PTEN/Akt pathway271 whereas iPSC-derived exosomes could regulate autophagy in hypoxic cardiomyocytes.272 Both Katsur et al. and Chen et al. focused on myocardial IRI. The former team found that exosomes derived from non-cardiomyocyte-related cells, i.e., CTX0E03 NSCs, could reduce infarct size while delaying cardiomyocyte mitochondrial permeability transition pore opening through the JAK/STAT pathway.273 The latter team discovered that MSC-derived exosomal miR-143-3p could suppress myocardial IRI by regulating autophagy via the CHK2-Beclin2 pathway.274 Finally, Chen and co-workers proved that bone marrow MSC-derived exosomes could attenuate cardiac hypertrophy and fibrosis in pressure overload-induced remodeling, thereby providing a promising potential treatment for heart failure.275

In comparison to MSC-derived exosomes, exosomes derived from ESCs exhibited comparable therapeutic effect for cardiac conditions. In terms of protection of cardiomyocytes, Khan et al. discovered that the ESC-derived exosomal miR-294 could improve cardiomyocyte survival, promote neovascularization and inhibit fibrosis after MI, thereby augmenting post-MI cardiac function.276 In addition, Tavakoli Dargagni et al. demonstrated that ESC-derived exosomes could alleviate doxorubicin-induced cardiotoxicity by inhibiting TLR4-NLRP3-mediated pyroptotic cell death in cardiomyocytes.277 Similarly, Singla’s team showed that ESC-derived exosomes could improve cardiac remodeling by enhancing anti-inflammatory M2 macrophages and reducing inflammation-induced pyroptosis.278 In terms of management of heart failure, Pang et al. exhibited that ESC-derived exosomes could attenuate heart failure, improve cardiac function and promote myocardial angiogenesis through the FGF2 signaling in a transverse aortic constriction-induced heart failure model.279 Using a coronary artery occlusion-induced heart failure model, Kervadec et al. showed that exosomes secreted by ESC-derived cardiovascular progenitors could recover cardiac functions such as reduced left ventricular end-systolic and end-diastolic volumes.280 Finally, the same research group later demonstrated that exosomes derived from more readily available cell sources, e.g., iPSCs, were capable of cardioprotective effects similar to those offered by ESC-derived ones.281

Other important subtypes of SC-Exo, such as iPSC-exo, HSC-exo and EPC-exo, have also exhibited cardio-protective effects. For example, exosomes secreted by iPSCs could exert cytoprotective effects on maintaining intracellular Ca2+ homeostasis and promoting cardiomyocyte survival, thereby improving recovery from MI.282 HSC-exo could reduce the cardiac injury-related indices and the degree of cardiac fibrosis while elevating the ejection fraction in an animal model of heart failure.283 In addition, systemic infusion of HSC-derived exosomes could improve ischemic cardiomyopathy in a rat model of acute MI, with additional benefits in treating the side effects such as kidney damage.284 Modification of HSCs using sonic hedgehog (Shh), an angiogenic factor, could preserve cardiac function after acute MI by delivery of exosomal Shh to ischemic myocardium.285 Ke et al. proved that EPC-derived exosomes could enhance the proliferation and angiogenesis of cardiac fibroblasts by activating mesenchymal-endothelial transition and decreasing the expression of HMGB1286 and later revealed that the exosomal miR-218-5p and miR-363-3p from EPC-derived exosomes could ameliorate MI by targeting the p53/JMY pathway.287 In an interesting study by Yue et al., IL-10 deficiency-induced systemic inflammation was found to compromise the reparative properties of EPC-derived exosomes on myocardial repair by upregulating integrin-linked kinase (ILK) enrichment in exosomes, and ILK-mediated activation of NF-κB pathway in recipient cells.288

In terms of treatment of thoracic disorders, Liu et al. proved that human ESCs-derived exosomes could alleviate inflammation, prevent excessive collagen deposition and preserve alveolar architecture in the lungs of mice with bleomycin-induced pulmonary fibrosis.289 This was achieved by the exosomal mi-17-5p targeting thrombospondin-2. Similarly, Zhou et al. demonstrated that the exosomal miR-302a-3p from iPSC-derived exosomes could suppress M2 macrophages via TET1, thereby mitigating pulmonary fibrosis.290 Liu et al. showed that EPC-derived exosomes could inhibit pulmonary artery smooth muscle cells proliferation and their resistance to apoptosis by regulating the Mitofusin-2 and Ras-Raf-ERK1/2 pathways, thereby acting as a potential therapeutic candidate for the treatment of pulmonary arterial hypertension.291 Two independent teams both revealed that human EPC-derived exosomes could improve outcomes of the LPS-induced acute lung injury partially through the delivery of miR-126 into the injured alveolus.292,293 Zhang et al. found that EPC-derived exosomes could improve the bioactivity of pulmonary microvascular endothelial cells and protect them from hyperoxic injury in the developing lung vasculature, thereby contributing to the treatment of bronchopulmonary dysplasia.294 Moreover, Montay-Gruel et al. demonstrated that human ESC-derived exosomes could improve the adverse late normal tissue complications associated with exposure of the lungs to ionizing radiation, such as those encountered during postoperative treatment of lung cancer.295

Urology and SC-Exo therapy

Chronic kidney disease (CKD) is a syndrome characterized by persistent changes in kidney structure, function, or both, affecting 10−14% of the global population.296 The most common pathological feature and final manifestation of CKD is some form of renal fibrosis. Kidney fibrosis occurs when wound healing is deregulated, which leads to excessive accumulation of ECM proteins, such as collagen and fibronectin. In their study, Liu et al. discovered that bone marrow MSC-derived exosomes could alleviate vascular calcification, a detrimental indicator of morbidity and mortality in CKD.297 This was achieved through exosomal miR-381-3p by targeting NFAT5, which was further verified in severe arterial calcification in dialysis patients. In alternative studies, several groups demonstrated the capacity of bone marrow MSC-derived exosomes in treating renal fibrosis, each with a distinct mechanistic interpretation. In a cellular study, Yin et al. found that exosomes could prevent TGF-β1-induced epithelial-mesenchymal transition of renal tubular epithelial cells by transporting Nedd4L, which activates autophagy. In a 5/6 subtotal nephrotomy rat model, Liu et al. revealed that exosomes could improve renal function and reduce fibrotic size by regulating the Smurf2/Smad7 axis.298 In a unilateral ureteral occlusion-induced interstitial fibrosis mouse model, Lu et al. demonstrated that exosomes could improve renal fibrosis by reducing the polarization of M1 and M2 macrophages by activating EP2 receptors.299

Acute kidney injury (AKI) and CKD are closely connected, with each a risk factor for developing the other. Renal IRI is a leading cause of AKI and acute kidney failure.300 Lim et al. proved that exosomes from iPSC-derived MSCs could correct serum creatinine level, tubular necrosis, apoptosis, inflammatory cytokine production, and oxidative stress in AKI mice by activating the ERK1/2 signaling pathway.301 In a similar work by Zhang et al., the exosomal miR-21-5p from EPCs were found to alleviate sepsis-induced AKI by inhibiting RUNX1 expression in CLP rats.302

Otorhinolaryngology and head & neck surgery and SC-Exo therapy

Hearing loss is the most common sensory deficit worldwide, affecting nearly 20% of the global population.303 The causes of sensorineural hearing loss (SNHL) can be very diverse, such as presbycusis, ototoxic medication-induced, noise-induced, and idiopathic sudden SNHL. Tsai’s team demonstrated that umbilical cord MSC-derived exosomes could rescue the loss of outer hair cells and repair cochlear damage in cisplatin-induced hearing loss.304 The underlying mechanism for the cochlea-protective effects is mediated by the miRNAs (e.g., miR-125a-5p and miR-125b-5p) and remodeling factors (e.g., fibronectin and galectin-3).

Cochlear IRI is one of the main reasons for idiopathic sudden and noise-induced SNHL, which can lead to irreversible damage of sensory hair cells and bipolar cochlear spiral ganglion.305 Its pathophysiology includes oxidative stress, excess cell death and dysregulated inflammation. Hao et al. discovered that exosomes derived from miR-21-overexpressing NSCs could prevent hearing loss from IRI by inhibiting the inflammatory process in the mouse cochlea.306 This was evidenced by a reduced auditory brainstem response threshold, upregulated IL-10 and downregulated TNF-α and IL-1β.

Hypothyroidism is a very common disease which could result from thyroidectomy or radioactive ablation to treat hyperthyroidism or thyroid cancer.307 Stem cell therapy and stem cell-derived exosome therapy have emerged as a promising management for hypothyroidism through thyroid regeneration. Using an in vitro culture system of thyroid lobes, Degosserie et al. suggested that EPC-derived exosomes could facilitate thyrocyte organization into thyroid follicles and lumen expansion (i.e., folliculogenesis), which was promoted by laminin-α1.308

Temporomandibular joint (TMJ) disorders are the second most common musculoskeletal condition affecting 31% of adults and 11% of children.309 Like other synovial joints in the body, TMJ is also prone to OA, sharing common pathophysiological processes (Section 3.2). Zhang and co-workers proved that MSC-derived exosomes could alleviate TMJ OA in an immunocompetent rabbit model by attenuating inflammation and restoring matrix homeostasis.310 The exosome-mediated joint repair was attributed to adenosine activation of Akt, ERK and AMPK signaling, as well as enhanced s-GAG synthesis.

Ophthalmology and SC-Exo therapy

Acquired optic neuropathy is a major cause of blindness in adults, and has various etiologies, such as vascular, inflammatory, traumatic, toxic, compressive, and nutritional etiologies. Retinal ganglion cell (RGC) loss is the hallmark of optic neuropathies, via multiple cell death pathways.311 Mead and Tomarev found that bone marrow MSC-derived exosomes could promote survival of RGCs, and regeneration of their axons, in a rat optic nerve crush model.312 The exosomal neuroprotective and neuritogenic effects were accomplished through exosomal miRNAs, demonstrating a cell-free potential for traumatic and degenerative ocular diseases.

Retinal degenerative diseases (e.g., age-related macular degeneration and retinitis pigmentosa) are the leading cause of bilateral irreversible vision loss worldwide.313 It is characterized by progressive degeneration of photoreceptors, RGCs or retinal pigment epithelium (RPE) cells. Bian’s team discovered that exosomes derived from NSCs could preserve photoreceptors, visual function and prevent thinning of the outer nuclear layer in an RCS retinal degeneration rat model.314 This was achieved by marked inactivation of microglial inflammation via exosomal targeting of TNF-α, IL-1β and COX-2. In two consecutive studies, Gao’s team showed that ESC-derived exosomes could alleviate retinal degeneration by enhancing the proliferation and retrodifferentiation of retinal Müller cells as replacement retinal neuronal precursors. On one hand, this was accomplished by regulating the expression of Oct4 in Müller cells through exosomal HSP90.315 On the other hand, activation of the Wnt signaling pathway by delivering BDNF protein to Müller cells also played an important role.316 Furthermore, using a rat model of inherited retinal degeneration, Park et al. demonstrated that both subretinal and intravitreal injection of human HSC-derived exosomes could provide functional rescue of a degenerating retina.317

Failed healing of corneal defect often leads to corneal blindness which has been reported as second only to cataract in the leading causes of blindness.318 The most severe and recalcitrant cases would need corneal transplantation. Wang et al. compared exosomes derived from iPSCs and MSCs as therapeutic providers for the treatment of corneal epithelial defects.319 It was found that both types of exosomes could promote proliferation, cell cycle progression and migration while inhibiting apoptosis in vitro, and accelerate corneal epithelium defect healing in vivo. More importantly, the iPSC-derived exosomes had a stronger therapeutic effect than the MSC-derived exosomes.

Corneal transplantation is one of the most successful forms of solid organ transplantation. However, graft rejection can occur in up to 90% of high-risk recipients.320 Both innate and adaptive immunity are the predominant reason for graft failure. Immunosuppressive drugs have shown only partial effectiveness. Jia et al. showed that MSC-derived exosomes could cross biological barriers and prolong graft survival time in a rat model of corneal allograft rejection.321 This was likely caused by inhibition of the infiltration of CD4+ and CD25+ T cells and the reduction of IFN-γ and CXCL11 via the Th1 signaling pathway.

Obstetrics and gynecology and SC-Exo therapy

Primary ovarian insufficiency (POI), or premature ovarian failure (POF), is defined as loss of ovarian function before the age of 40. Non-genetic causes of POI include autoimmune disorders, metabolic conditions, infections, and iatrogenic procedures (e.g., chemotherapy, radiotherapy, and surgery). Women with POI suffer from various complications, such as osteoporosis, infertility, cardiovascular disorders and depression.322 Although promptly initiating hormone replacement therapy is critical to control these symptoms and complications, it fails to restore ovarian function. Currently-tested experimental therapies include mitochondrial activation, in vitro activation, stem cell therapy, and exosome therapy.323

The work conducted by Li et al. showed that umbilical cord MSC-derived exosomes could improve ovarian function in a cyclophosphamide (CTX)-induced POI mouse model.324 The SC-exo therapy not only restored ovarian function-related hormone levels and the number of ovarian follicles, but also improved the reproductive ability of POI mice. In addition, the exosomes promoted the proliferation of ovarian granulosa cells (GCs) by regulating the Hippo pathway, and the effect was neutralized by a YAP inhibitor. Similar results were obtained using exosomes from iPSC-derived MSCs.325 The work performed by Ding et al. illustrated that umbilical cord MSC-derived exosomes could restore ovarian phenotype and function in a POI mouse model, promote proliferation of CTX-damaged human GCs and oocytes, and alleviate ROS accumulation by delivering exosomal miR-17-5p and targeting its downstream mRNA SIRT7.326 It was further elucidated that miR-17-5p down-regulated PARP1, γH2AX, and XRCC6 expression by inhibiting SIRT7. Lastly, the work completed by Yang et al. revealed that bone marrow MSC-derived exosomes could recover the estrus cycle, increase the number of basal and sinus follicles, increase estradiol E2 and anti-Mullerian hormone levels, and reduce follicle stimulating hormone and luteinizing hormone levels in a chemotherapy-induced POF rat model.327 Mechanistically, this was achieved by exosomal miR-114-5p that targets PTEN.

From preclinical studies to clinical trials of exosome therapy

Many preclinical studies, as discussed in Sections 3 to 11, have confirmed the advantages of MSC-derived and NSC-derived exosomes to treat many diseases spanning the subspecialties of surgical practice. Without restricting the scope to stem cell-derived exosomes only, many clinical trials have demonstrated the role of exosomes to be twofold: biological markers and therapeutic agents. A search on ClinicalTrials.gov using ‘exosome therapy’, ‘exosome treatment’, and ‘exosome’ as keywords generated 188 records. However, only 60 (32%) of these directly relate to interventional studies using exosomes as therapeutic agents (Table 5). The rest, especially oncology-related trials, mostly used exosomes as biomarkers, such as key players during disease pathogenesis (e.g., NCT04288141, NCT04154332), diagnostic markers and guidance before treatment (e.g., NCT04629079, NCT03791073, NCT05451342, NCT03432806), monitoring indices and predictive tools for treatment efficiency (e.g., NCT05427227, NCT04499794, NCT04852653, NCT05370105, NCT03800121, NCT05370105, NCT05328089), and prognostic indicators after treatment (e.g., NCT06026735, NCT05705583, NCT05411445, NCT04167722, NCT05575622). The clinical applications of exosomes as biomarkers have been extensively explored in other reviews328,329,330,331,332 and are therefore beyond discussion in this review. In terms of the clinical characteristics of the 60 clinical trials on exosome therapy (Table 5), there are several highlights worth mentioning.

Table 5 Clinical trials of exosome therapy
Full size table

Firstly, the spectrum of diseases covered is very broad, as both surgical and medical conditions are included. These include many surgical disorders discussed in Sections 3 to 11, such as orthopedic diseases (osteoarthritis of the knee in NCT05060107, bone loss in NCT04998058, and intervertebral disc degeneration in NCT04849429), neurosurgical diseases (ischemic stroke in NCT03384433 and Alzheimer’s disease in NCT04388982), plastic surgical diseases (cutaneous wound healing in NCT02565264 and NCT05475418), general surgical diseases (liver cirrhosis in NCT05871463), cardiothoracic diseases (myocardial infarction in NCT05669144), and ophthalmology diseases (retinitis pigmentosa in NCT05413148). In other words, some preclinical studies have not yet developed into clinical trials. These include, but are not limited to, exosome therapy for fracture333 spinal cord injury334 traumatic brain injury335 acute liver injury336 and hearing loss304,306 which might serve as future directions for exosome therapy-related clinical trials.

Secondly, an increasing number of clinical trials focused on two medical conditions, i.e., COVID-19 (16 trials, 27%) and cancer (5 trials, 8%). MSC-derived exosomes can manage viral infection and lung damage in COVID-19 through both reparative actions and regenerative effects.337 The former manifests as blockage of viral entry and replication, and suppression of the cytokine storm, whereas the latter as prevention of inflammation, and fluid-clearance and restoration of lung permeability. In addition, exosomes can be engineered into a drug (e.g., CD24, a potent immune regulator) delivery platform338 and even a vaccine339 to combat COVID-19. In contrast to the MSC-derived exosomes for COVID-19 management, exosomes used for cancer treatment mainly rely on non-stem cells and cargo engineering. This is partially because MSC-derived exosomes demonstrate controversial effects on tumorigenesis and metastasis.340,341 Although the exact interaction between MSC-derived exosomes and tumor cells remains open to debate, scientists have overcome several obstacles by modifying exosomal cargos to deliver antioncogenic nucleic acids and anticancer medications, and exosomal membranes for specific tumor targeting.342,343,344,345,346

Finally, among the 40 clinical trials using stem cell-derived exosomes for disease treatment, 38 (95%) used MSC and 2 (5%) used iPSC as the cellular source for exosomes. However, this differs significantly from the preclinical studies discussed in Sections 3 to 11. The lack of use of NSC-derived exosomes in clinical trials might be partially because of the supply constraints of their parental cells.347 Currently, NSCs can be obtained from three sources348 (Fig. 1a): 1, isolation from primary CNS tissues (e.g., adult and fetal brain); 2, differentiation from pluripotent stem cells (e.g., iPSCs and ESCs); 3, reprogramming of somatic cells (e.g., fibroblasts and blood cells) to iNSCs.349 Recent studies have developed fibroblast-derived iNSCs, opening a new window for obtaining exosomes from NSC-like cells. These iNSC-exo could not only promote cell survival and proliferation no less than NSC-exo in vitro350,351 but also enhance recovery after ischemic stroke148 and mitigate AD-like phenotypes in preclinical models.190 Therefore, iNSCs might be an excellent cellular source to produce clinical-grade exosomes in clinical trials.

In summary, the progression from preclinical studies to clinical trials of exosome therapy has been expeditious. However, issues like insufficient clinical indications for exosome treatment and limited sources for parental stem cells remain to be addressed. In addition, once the limitations (Table 1) in upscaling of manufacturing, compliance with good manufacturing practice, and regulatory framework are overcome329 stem cell-derived exosome therapy will soon be incorporated into clinical practice and serve at the patient’s bedside.

Conclusion and future perspectives

Exosomes have been pursued recently as a cell-free alternative to stem cell-based therapy. ESC-, iPSC-, HSC-, MSC-, NSC- and EPC-derived exosomes are of particular interest, partially due to the pluripotency or multipotency of their parental cells. After going through production and purification with or without modification, stem cell-derived exosomes have demonstrated tremendous potential in treating numerous diseases encountered during surgical practice. These are exemplified by disorders in orthopedic surgery (e.g., fracture, osteoarthritis, and spinal cord injury); neurosurgery (e.g., ischemic stroke, traumatic brain injury, and Alzheimer’s disease); plastic surgery (e.g., wound healing); general surgery (e.g., acute liver injury); cardiothoracic surgery (e.g., myocardial infarction); urology (e.g., chronic kidney disease); head and neck surgery (e.g., sensorineural hearing loss); ophthalmology (e.g., acquired optic neuropathies), and gynecology (e.g., primary ovarian insufficiency). Mechanistically, the diverse therapeutic effects of stem cell-derived exosomes are achieved through disease-specific cellular and tissue responses (e.g., tissue regeneration, anti-inflammation, anti-cell death, immunomodulation, and anti-oxidative stress) and tissue-specific molecular signaling pathways (e.g., Wnt/β-catenin, PTEN/PI3K/Akt/HIF-1α, MAPK, and JAK/STAT pathways). Collectively, stem cell-derived exosome therapy has been proven to be a potent and versatile surrogate to stem cell therapy in the surgical arena.

Future emphasis of clinical applications of stem cell-derived exosomes should be placed on various nodes of this therapeutic pipeline. Firstly, targeting the pretherapeutic large-scale production of exosomes, a high-throughput cellular source as well as a reproducible and scalable production and isolation protocol are required. Compared to the static system growing monolayer cells, dynamic system in the form of bioreactor, e.g., hollow-fiber bioreactor and stirred tank bioreactor, can increase the efficiency by producing copious cells and exosomes in a short period of time. However, the phenotype of parental cells and derived exosomes might change due to physical and shear stress encountered in a reactor. Thus, the working parameters of the bioreactor must be optimized to facilitate large-scale production of stem cell-derived exosomes. Secondly, targeting the therapeutic modality of exosomes, delivery methods other than systemic administration need to be explored. When delivered through the venous system, exosomes are rapidly cleared from blood circulation and accumulate in the liver, spleen and lungs, which can be overcome by local delivery. Various biomaterials have been recently used to protect, assist and augment locally delivered exosomes to maximize their therapeutic effects. These biomaterials could be designed according to their sources (e.g., natural, synthetic, and hybrid polymers), format (e.g., scaffold, patch, spray, and microneedle), and responsiveness (e.g., temperature, pH, and protein), thereby allowing disease-specific customization. Lastly, targeting the therapeutic indications of exosome therapy, more diseases than the ones discussed in this review should be included into future preclinical studies and clinical trials. For example, airway inflammatory conditions (e.g., allergic rhinitis and asthma) could be suitable candidates for exosome therapy, considering the immunomodulatory effect of MSC-derived exosomes. Disorders that are best treated using surgical implants (e.g., cochlear implant, intraocular lenses, and contraceptive intrauterine devices) could be managed in the form of implant-based local release of exosomes. In addition to primary diseases, secondary conditions including surgical operation- and general anesthesia-related complications (e.g., cognitive impairment, wound paresthesia, and malignant hyperthermia) might become therapeutic targets of exosome therapy. Collectively, efforts to upscale exosome production in conjunction with multimodal exosome delivery will accelerate the clinical applications of stem cell-derived exosomes in a rapidly expanding disease spectrum.