Open access peer-reviewed chapter
Abstract
This chapter focuses on the emerging field of exosomes in the context of reperfusion injuries. Exosomes, nano extracellular vesicles with diverse cargo, play a crucial role in cell-to-cell communication. Exosome quantity and content changes have been implicated in various pathologies, including kidney, brain, heart, and liver ischemia-reperfusion injury. Particularly, exosomes derived from mesenchymal stem cells (MSCs) have shown promising potential as a treatment approach. This chapter aims to provide a comprehensive understanding of exosome biogenesis, the role of organ-specific exosomes in reperfusion injury pathophysiology, and the advantages and prospects of exosome-based treatments. By exploring the multifaceted aspects of exosomes in reperfusion injuries, this chapter will contribute to the advancement of knowledge in this field.
Keywords
- exosomes
- reperfusion
- kidney
- heart
- brain cells
- liver
1. Introduction
Exosomes are small extracellular vesicles released by various cell types in different cell conditions, including those involved in tissue repair and regeneration. These particles have been linked to a variety of biological phenomena, including immune responses, viral pathogenicity, pregnancy, diseases related to pregnancy, cardiovascular diseases, diseases related to the central nervous system, and the progression of cancer [1]. They play a crucial role in cell-to-cell communication by transferring bioactive macromolecules, such as proteins, nucleic acids, and lipids, between cells [2]. Exosomes have gained significant attention in physiology due to their unique formation process within cells [3], which is thought to play a crucial role in determining composition and potential functions after they are released into the extracellular space. In the context of reperfusion injuries, the potential protective effects of exosomes have garnered considerable attention.
Reperfusion injury refers to tissue damage that occurs when blood supply returns to previously ischemic tissue. While restoring blood flow, it is necessary to prevent further damage; re-establishing oxygen and nutrients can also lead to the generation of reactive oxygen species (ROS) and inflammatory responses, which can exacerbate tissue injury. Acute damage caused by reperfusion is characterized by cellular injury, upregulation of cytokines, infiltration of leukocytes, and the formation of neutrophil extracellular traps (NETs) among other cellular and tissue changes.
The study of exosomes has gained high importance given their potential use in different areas of biology and health, which will be described in this book chapter. Some of the topics most currently addressed are:
Exosomes as rich source of biomarkers: Exosomes can carry specific proteins, nucleic acids, and miRNAs that are indicative of cellular stress, injury, or regenerative processes. Studying the composition and content of exosomes released during reperfusion injuries can help identify potential biomarkers for early detection, prognosis, and monitoring of the extent of tissue damage.
Exosomes in cellular communication and cytoprotection: Exosomes released by certain cell types, such as endothelial cells, cardiomyocytes, and stem cells, have been shown to promote cell survival, reduce inflammation, and protect against reperfusion injury. Understanding the molecular cargo and mechanisms of exosomal transfer can provide insights into intercellular signaling and potential therapeutic targets.
Regenerative potential of exosomes content: Exosomes derived from stem cells, including mesenchymal stem cells (MSCs), have been reported to exhibit regenerative properties and promote tissue repair. These exosomes can transfer regenerative factors and modulate inflammatory responses, thereby enhancing the regenerative potential of damaged tissues after reperfusion injury.
Therapeutic Applications: Harnessing the therapeutic potential of exosomes is an active area of research. Researchers are exploring strategies to engineer exosomes with specific cargo or modify their composition to enhance their protective effects. Exosomes could be utilized as targeted delivery vehicles for therapeutic agents, such as anti-inflammatory drugs or regenerative factors, to mitigate reperfusion injury.
In this book chapter, we will review from the biological basis of exosome formation and structure to the physiology and involvement of exosomes in reperfusion damage. We will also analyze the most important studies that to date have given insights into the best techniques for isolating these particles. Finally, we will explore a series of future perspectives that involve the use of exosomes as possible tools for treatments in complicated pathologies.
2. Biogenesis of exosomes
2.1 Definition and characteristics of exosomes
Exosomes are a group of biomolecules between 50 to 100 nanometers in diameter. They have a density of 1.13–1.19 g/mL and exhibit characteristic morphology when viewed under a transmission electron microscope. Among their main protein markers, we find tetraspanins, TSG101, and Hsp70 primarily [4].
Exosomes are the smallest characterized molecules after exomeres [5]. With a size ranging from 30 to 200 nm, they are formed by a membrane system that contains the same biological structures as a cell, including nucleic acids, proteins, lipids, amino acids, and metabolites, see Figure 1. These components can reflect their cellular origin. This particular architecture exhibits an important array of membrane-associated, oligomeric proteins, resulting in protein heterogeneity [6].
Figure 1.
Biogenesis of the exosomes.
An important role of exosomes is to communicate information between distant and different cells while also participating in the microenvironmental remodeling of the extracellular matrix. The size of exosomes has been determined using various laboratory techniques; however, each of these techniques provides different size variations. The techniques used to determine the size of exosomes include single-particle interferometric reflectance (SPIR) imaging, fluorescence microscopy, nanotracking particle analysis, and resistive pulse sensing [6, 7]. Transmission electron microscopy is a technique that only determines the diameter of the exosome membrane. For this reason, when determining the diameter of these particles, we consider the method used for measurement. This measurement can be affected by different variables, including shrinking, swelling, or flattening. These processes are susceptible to affecting our samples.
Exosomes can originate from various sources, such as urine, blood, saliva, or even tears. The main role of great interest for which exosomes have been studied in recent years is their mediation of cell communication in both health and disease processes, directly affecting cell physiology [8].
2.2 Mechanisms of exosome formation and cargo loading
The process for the formation of exosomes is called biogenesis. This process describes how exosomes are formed inside cells and involves a crucial role of the plasma membrane in creating multivesicular bodies (MVBs). Exosomes are released from these MVBs out of the cell [4]. The process begins within an early endosome, containing endocytosed components. This early endosome can undergo three fates: it can either proceed toward the Golgi network, continue maturing as a late endosome, or recycle through endosome-lysosome fusion (Figure 2) [9].
Figure 2.
Exosomes cargo.
MVBs are characterized by being formed as biological structures through the creation of cell membranes. Vesicles captured by these structures are deposited inside as intraluminal vesicles (ILVs). These vesicles, present within this newly formed component, are released outside the cell through the fusion of the intraluminal vesicles with the plasma membrane. This is how exosomes are released outside the cell [8]. The formed body has three potential destinations: one leads to the lysosomal pathway, where proteins are recycled through lysosomes [10], while the other involves the exocytic pathway, allowing newly formed exosomes to be released outside the cell (see Figure 3) [8].
Figure 3.
Exosomes internalization.
Exosome formation involves a crucial role played by various proteins, including Rab GTPases and ESCRT proteins. Additionally, specific molecules can serve as markers for exosomes, including CD9, CD81, CD63, flotillin, TSG101, ceramide, and Alix [8]. Regarding the components present on the surfaces of these vesicles, tetraspanins, integrins, immunomodulatory proteins, and others can be mentioned [11]. In the process of exosome formation, various molecules are associated with the endosomal sorting complex required for transport (ESCRT). Generally, the proteins involved in this process can be divided into two categories: ESCRT-dependent and ESCRT-independent. ESCRT is a group of proteins responsible for coordinating and deforming membranes through its multi-protein machinery. This machinery facilitates the generation of new exosomes within intraluminal vesicles. ESCRT has the capability to form four distinct complexes: ESCRT 0, I, II, and III [12].
An important role of ESCRT 0 to II is direct participation in sorting cargo. This process occurs through the formation of microdomains on the vesicle’s surface. These domains act as organizing centers, facilitating signal transduction within the vesicle’s interior. Following this, ESCRT manages the budding and cleavage of these domains to generate intraluminal vesicles (ILVs). Vps4 (Vacuolar protein sorting-associated protein 4) assists in dissociating ESCRT III after the membrane scission process is completed. The SNF7/CHMP4 protein complex plays a role in contributing to membrane formation by inducing inward budding and generating ILVs. This is achieved through the polymerization of the mentioned complex, resulting in the formation of spirals that accumulate potential energy. This accumulated energy enables the creation of negative curvature, leading to the generation of exosomes [13].
The second group of molecules involved in the process of exosome formation consists of those that are independent of ESCRT. These molecules can be either lipids or proteins. Among the most crucial for this process are tetraspanins, which play various roles in the pathway of exosome generation. Tetraspanins play a direct role in loading compounds into multivesicular bodies. They enhance the secretion of specific compounds into exosomes and enable the compartmentalization of the exosomal membrane into functional domains enriched with these proteins [14]. There are other lipid compounds strongly associated with promoting exosome formation, including ceramides, membrane sphingolipids, cholesterol, and lipid raft microdomains. These compounds induce spontaneous negative curvature of the membrane, facilitating the generation of intraluminal vesicles independently of ESCRT [15].
The process by which the contents of exosomes are introduced is referred to as cargo sorting. This process ensures that the relevant content is loaded inside the vesicles. The cargo can encompass various components, including amino acids, proteins, metabolites, lipids, as well as different types of RNA such as messenger RNA (mRNA), microRNA (miRNA), long noncoding RNA (lncRNA), circular RNA (circRNA), PIWI-interacting RNA (piRNA), and DNA [16]. The mechanisms to load these components have a different nature as shown in Figure 2. The process initiates with a signal that enables proteins to enter the endosomal pathway. This signal involves monoubiquitination, which allows the cargo to be recognized and bound by the ubiquitin-interacting motif (UIM) domain of the ESCRT 0 Vps27/Hrs protein [17].
Ubiquitination is recognized for facilitating the direct entry of proteins into exosomes. However, the precise mechanism for nonprotein compounds remains unclear. For instance, in the case of microRNA, the exact mechanism is not fully understood. It is known that microRNA can enter exosomes through selective binding to heterogeneous nuclear ribonucleoprotein A2B1 (hnRNA2B1), which is expressed on the exosome membrane [18]. The cellular state in which exosomes are generated plays a crucial role in determining the content of the generated exosomes. Furthermore, the protein content can be influenced by both ESCRT-dependent and ESCRT-independent factors, including but not limited to ESCRT-III, Alix, Syntenin-1, and ceramides. These factors contribute to shaping the composition of exosomes [19]. Finally, prior to sealing the vesicle that gives rise to the exosome, a deubiquitination process takes place to remove ubiquitin from the proteins associated with the cargo [20].
2.3 Release and uptake of exosomes by recipient cells
Released exosomes can engage with the plasma membranes of neighboring or remote cells, facilitating the internalization and subsequent release of their contents. This intricate process involves SNARE proteins, which play a pivotal role in mediating membrane fusion to enable exosome release [21].
The transportation of multivesicular bodies (MVBs) to the plasma membrane, facilitating the release of exosomes, is orchestrated by the cytoskeleton. Specifically, actin filaments and microtubules play a pivotal role in transporting MVBs, ultimately enabling the release of exosomes [19]. An essential aspect to consider regarding exosome distribution is that while exosomes can be present within the organism, their biodistribution is influenced by various factors. These factors include considerations such as their retention in lymph nodes and bone tissue, as well as potential competition with exosomes secreted by other cells. After exosomes have reached their recipient cell, they can engage in three distinct modes of interaction with the target cell. These interaction modes encompass internalization, membrane fusion, and binding to receptor ligands such as MHC and TNF ligands. These binding interactions can subsequently trigger the initiation of signaling cascades [22].
3. Isolation, characterization, and cargo analysis of exosomes
To comprehend the content of exosomes, various laboratory tests are essential to ascertain the internal composition of these vesicles. The initial step involves isolation, which determines their origin. As previously mentioned, the source can vary, encompassing cell cultures, plasma, sweat, tears, urine, and other bodily fluids. Characterization of exosomes involves assessing their physical attributes such as size, protein-lipid ratio, density, and other physicochemical parameters. Additionally, this characterization extends to quantitatively determining the contents of proteins, lipids, carbohydrates, and nucleic acids within the exosomes. Given the diverse range of tests that can be conducted on exosomes, it is crucial to discern which characteristics are the most pivotal parameters for addressing the specific questions we seek to answer.
3.1 Methods for isolating exosomes from different biological sources
The methods and techniques used to isolate exosomes can be categorized into conventional and novel approaches. Within conventional techniques, we encounter ultracentrifugation-based separation, size-based separation, and precipitation.
Ultracentrifugation stands as the primary technique employed for exosome isolation. This method exploits centrifugal force to create gradients within the sample. Notably, ultracentrifugation boasts simplicity, cost-effectiveness, high purity, and the potential for protocol standardization [23]. The second technique used is the separation based on size; this technique combines ultracentrifugation with size-exclusion chromatography; the advantages that this technique offers allow obtaining large amounts of exosomes with high purity; with this technique, urinary exosomes and those from platelets can be obtained [24]. The third technique involves precipitation using agents such as polyethylene glycol. While this method offers relative simplicity and speed in isolation, it comes with higher costs and inadequate specificity. This technique is applicable for extracting exosomes from serum, plasma, or cell supernatants [25].
On the other hand, novel isolation techniques offer technical advantages for exosome extraction; however, ensuring specificity and purity demands careful attention. Several of these techniques include:
Immunoaffinity Separation: This method relies on antigen-antibody interactions within a conjugated platform featuring specific antibodies on the exosome surface. It is a straightforward and specific technique, although it tends to be expensive [26]. Magnetic separation method involves employing magnetic beads conjugated with antibodies. Like immunoaffinity separation, magnetic separation offers the benefits of specificity and ease of execution. Nonetheless, it comes with a higher cost [27].
The subsequent category of techniques for exosome isolation involves separation based on their physical properties, particularly their size. These methods often utilize specialized membranes that serve as filters, akin to molecular exclusion chromatography. This type of technique offers a wide range of applications for exosome extraction [28].
Lipid-mediated separation: The most used technique involves the lipid nanoprobe coupled with high-affinity silica-based TiO2. This approach offers the advantage of causing minimal damage to extracellular structures, thereby facilitating effective and efficient isolation. However, potential interference due to contamination remains a consideration with this method [29].
Thermophoretic enrichment: This method is both simple and quick to apply, utilizing thermophoresis to achieve efficient isolation of extracellular vesicles [30].
Microfluidic-based acoustics: It employs acoustic radiation as an active means to separate extracellular vesicles, effectively facilitating the efficient separation of exosomes [31].
3.2 Techniques for exosome characterization and cargo analysis
More contemporary techniques for exosome characterization encompass diverse approaches for their detection. The first approach employs colorimetric detection utilizing a reaction between hydrogen peroxide and TMB (3,3′,5,5′-Tetramethylbenzidine). Sample volumes required typically span from 100 to 500 μL. This method has been employed to evaluate exosomes sourced primarily from breast cancer cells, prostate cells, and urine [16]. The second characterization technique involves fluorescence-based detection. It is applicable to a variety of samples, including plasma, cell supernatants, and blood samples. Fluorophores are utilized, detectable by electronic equipment, with a detection limit ranging from 21 exosomes to 4 million particles per microliter of sample [16]. Electrochemical detection constitutes another technique, often using sample volumes ranging from 10 to 30 μL. Detection is achieved through specific biosensors and antibody microarray sensors [16].
Surface plasmon resonance detection methods (SPR-detection) involve utilizing specific antibodies to target antigens within exosomes. This method can be applied to exosomes from sources such as breast cancer cells (HER2 positive), lung cell lines, plasma, and urine samples from lung cancer patients. Sample volumes may range from 250 to 1500 μL [16].
Surface-Enhanced Raman Scattering (SERS) is a characterization technique utilizing sample volumes ranging from less than 1 to 400 μL. This technique enhances the Raman signal of small molecules attached to a rough metal surface through electromagnetic and chemical mechanisms. Exosomes for this method can originate from various sources including plasma, cell cultures, serum samples, and lung cells [16].
3.3 Relevance of cargo analysis in understanding exosome-mediated effects
It is very important to know the content of the exosomes to determine biomolecule profiles that it contains inside; it is important because it has been verified through these analyses that their content is different between the cells in the organism, even according to the physiological state of each of them specially in cancer [28]. Furthermore, research has shown that dysregulation of exosome-related proteins can contribute to the development and progression of disease states, highlighting their potential as diagnostic and therapeutic targets [32].
Various research teams have recognized exosomes as valuable sources of potential markers or biomarkers. These include applications for early diagnosis of various pathologies across different organs, including the kidneys. Consequently, thorough characterization and analysis of the content within these vesicles provide a means to comprehensively understand the intricate molecular and cellular mechanisms underpinning the pathophysiology of diverse diseases. This knowledge has the potential to revolutionize our understanding of disease development and progression and potentially aid in the development of novel diagnostic and therapeutic approaches.
4. Exosomes in reperfusion injury pathophysiology
The role of exosomes has been extensively investigated in various diseases such as cancer, neurodegenerative diseases, and heart failure. Additionally, pathogens exploit exosomes to infect host cells [33]. Numerous studies have highlighted augmented exosome release and alterations in their contents in hypoxic conditions [34]. Keeping up with the advancements in the study of reperfusion injury pathophysiology includes a comprehensive understanding of exosomes and their potential involvement in this phenomenon.
4.1 Kidney-derived exosomes and their role in reperfusion injury
The proximal tubule is considered the most sensitive cellular entity to ischemia, hypoxia, or nephrotoxic damage. This sensitivity is largely related to the high metabolic rate and strong reliance on oxidative phosphorylation of these cells [35]. Exosomes released by proximal tubular epithelial cells are believed to play a significant role in the context of ischemia-reperfusion injury. Many studies have demonstrated that these cells increased the production of exosomes under hypoxic conditions [36].
In a model of ischemia-reperfusion injury, exosomes demonstrated a significant role in mediating epithelial–mesenchymal communication (EMC) during fibrinogenesis. Another interesting observation was that the renal proximal tubular epithelium shows the major release of exosomes. The upregulation in exosome production was not limited to the model of ischemia-reperfusion injury; it was also observed in cases of unilateral obstruction and partial nephrectomy, showing its importance in different models of kidney damage [37].
A study of tubular epithelial cells (TECs) under hypoxic conditions revealed the capacity of exosomes to facilitate the activation of fibroblasts. Exosomes play a role in cell-to-cell communication with protein transport and mRNA transfer. The delivery of TGF-β1 mRNA
The capacity of exosomes to transport miRNA is also important. A comparison between renal tissue samples with and without ischemia-reperfusion identified upregulation of miR-150p that promotes fibroblast activation [40]. MiR-374b-5p found in exosomes of TECs is related to promoted macrophage activation in IR and increased kidney damage [40]. Both miRNAs are related to the inhibition of the suppressor of cytokine signaling (SOCS-1), which is a negative regulator of NF-κB signaling pathways [41].
Several well-known proteins that play a significant role in ischemia-reperfusion can be encapsulated within exosomes [42]. One way to study proteins around pathophysiology or as biomarkers of kidney disease is the study of urinary exosomes. This gives us a view of what is happening in the kidney, and it became an important part of kidney study. Increased fetuin-A and decrease of Na+/H+ exchanger isoform 3 (NHE3) and aquaporin 1 are examples of changes observed in urinary exosomes after ischemia-reperfusion [43].
4.2 Brain-derived exosomes: implications for ischemic stroke and reperfusion injury
Brain cells such as neurons, astrocytes, oligodendrocytes, endothelial cells, and microglia are capable of releasing exosomes [44]. Under pathologic conditions, neuron exosomes mediate nutritional metabolic support, nerve regeneration, inflammation, and propagation of toxic components, while exosomes released by astrocytes and microglia exert neuroprotective effects [45].
In a model of oxygen and glucose deprivation, astrocyte-derived exosomes inhibited neuron apoptosis and autophagy. The same results were found
Microglia exosomal miRNA-317 has a protective effect after ischemic injury with Notch-1 as a target [49]. Additionally, microglia exosomes release miRNA-137 and -34a, both of which are capable of inhibiting Notch-1 [50].
The study of exosomal miRNA in the plasma of patients after ischemic stroke has associated miRNAs-9, −124, −134, −152-3p, and −223 with stroke severity and miRNAs-134 and -223 with poor prognosis [51].
Use of exosomes derived from plasma of a murine model of ischemic preconditioning decreases the damage in a cellular model of oxygen–glucose deprivation and restoration of N2a. The study identified miR-451a as a protective factor through Rac1 inactivation [52]. Rac1 suppression reduced infarction volume and promoted neural stem cell activity in brain ischemia-reperfusion [53].
Exosomes have different roles in angiogenesis, neurogenesis, autophagy, and blood–brain barrier during ischemic stroke, which made them a good source of biomarkers for diagnosis and prognostic of this neurological disease [54].
4.3 Cardiac exosomes: involvement in myocardial ischemia and reperfusion injury
The heart is another organ affected by ischemia-reperfusion. Myocardium ischemia-reperfusion injury involves mechanisms such as oxidative stress, intracellular calcium overload, energy metabolism disorder, apoptosis, endoplasmic reticulum stress autophagy, pyroptosis, ferroptosis, and necroptosis [55]. Cardiomyocytes can uptake exosomes from various cell types. Cardiac fibroblast exosomes can be internalized by cardiomyocytes. During ischemia-reperfusion injury, exosomes derived from cardiac fibroblasts elevate the levels of miR-133a, which has the capability to mitigate cardiomyocyte pyroptosis [56]. Additionally, cardiomyocytes are capable of uptaking exosomes released by polymorphonuclear neutrophils (PMNs). Exosomes originating from PMNs that have been stimulated with the calcium-sensing receptor (CaSR) demonstrate the capacity to mitigate ischemia-reperfusion damage in cardiomyocytes [57]. Cardiomyocytes can also take exosomes released by endothelial cells [58].
Angiogenesis is one of the most vital pathways involved in ischemia-reperfusion, playing a crucial role in the recovery following the insult. Therapeutic angiogenesis has emerged as a significant alternative for treating ischemic heart disease [59]. Pro-angiogenic miRNAs have been detected in exosomes such as miR-126-3, −125, −30c, −126, −17, and 19a/b [60].
Use of exosomes as a therapy of ischemic preconditioning is also considered in the heart. We have found cardioprotective proteins in cardiomyocyte exosomes such as IL-6 and heat shock proteins [61]. Serum exosomes of ischemic preconditioning rats have a protective effect in ischemia-reperfusion injury activating PI3K/AKT signaling pathway [62]. The cardioprotective effects of ischemic preconditioning (IPC) exosomes have also been attributed to the presence of miR-126a-3p, which influences Akt and Erk1/2 signaling pathways [63].
4.4 Liver-derived exosomes and their contribution to hepatic reperfusion injury
Hepatic ischemia-reperfusion injury (HIRI) arises during liver surgery and transplantation. Oxidative stress and inflammation are among the cellular and biochemical factors contributing to liver damage during IR. However, the complete process is still under study [64]. Liver cells are capable of releasing and taking exosomes. This included hepatocytes, Kupffer cells, and hepatic stellate cells, as well as biliary epithelial cells and endothelial cells [65]. One of the mechanisms involving hepatocyte exosomes promoting cell proliferation has been identified
The liver can sustain damage during intestinal ischemic reperfusion due to the ability of intestinal cell exosomes to migrate and impact the liver. Zhao et al. conducted a study to investigate the potential influence of these exosomes on the liver. The study’s findings indicated that exosomes generated after intestinal ischemia-reperfusion (IR) contributed to liver damage by polarizing M1 macrophages. Suppressing the release of exosomes resulted in a reduction of hepatic injury [67]. Indeed, this is an interesting perspective of how exosomes’ effects extend beyond the cells of their originating organ.
5. Mesenchymal stem cell-derived exosomes as a potential treatment
The use of mesenchymal stem cells (MSCs) in immunomodulation and regenerative medicine is not new. Like all the cells that we have mentioned, MSCs are capable of producing exosomes. These exosomes have intrinsic characteristics of protein and miRNA that could have an effect in targets cells and also could be interesting vehicles of genes, drugs, enzymes, or RNA [68]. Another important characteristic is that MSCs are easy to be cultured and manipulated [69].
Exosomes have emerged as an important therapeutic option for several reasons. Their biological origin ensures that their components are easily metabolized by cells, and they possess low immunogenicity due to this inherent compatibility [70]. They also possess great stability and low toxicity [71]. In the context of brain, exosomes represent an excellent option of therapy and drug delivery for their capacity to cross BBB [72]. The use of MSC-derived exosomes shows promise as a treatment for ischemia-reperfusion injuries. Angiogenic factors like VEGF, FGF, Ang-1, and Ang-2, among others can be found in MSC exosomes. Also, important angiogenic miRNA as −107, −31, −126, 26a, and -20b-5p [73]. Their anti-inflammatory and immunomodulatory properties made them capable to regulate macrophage polarization [74, 75].
The origin of MSCs determines their cargo and effects on target cells. Even the exosomes from bone marrow MSCs (BMSCs) contain mRNA that is relevant to cell transcription and the regulation of proliferation; exosomes from human liver stem cells contain specific mRNA related to liver metabolism and its proliferation [76].
The
MSC-derived exosomes have been tested in ischemia reperfusion with interesting results. MSCs for bone, adipose tissue, placental, kidney, and umbilical cord have been tried in clinical models of kidney ischemia reperfusion. In preclinical studies, these exosomes have demonstrated being anti-inflammatory and anti-fibrotic, as well as decreased tubular damage, creatinine, BUN, and proteinuria [78]. MSC exosomes in ischemic stroke have shown promoted angiogenesis, neurogenesis, oligodendrogenesis, and astrogenesis in experimental therapy [79].
6. Challenges and future perspectives
One of the primary challenges in exosome research is the isolation and purification of exosomes from complex biological samples. Existing isolation methods, such as ultracentrifugation, immunomagnetic bead isolation kits, and density gradient separation, have limitations in terms of yield, purity, and scalability [80]. Standardization of isolation protocols and characterization methods is essential to ensure consistency and comparability among studies; however, at present, there is still no consensus on the most efficient and effective method of extraction [81].
The therapeutic potential of exosomes heavily depends on their ability to carry and deliver specific cargo molecules, such as miRNAs, siRNAs, metabolites, and proteins, to target organs [82]. Nevertheless, efficiently loading therapeutic cargo into exosomes remains a challenge. Moreover, targeted delivery to affected tissues and cells needs improvement, as the passive targeting of exosomes may not always suffice for optimal therapeutic outcomes. These exosome-based therapies demand large-scale production to meet clinical demand. Cost-effective and scalable manufacturing processes that maintain the integrity and biological activity of exosomes are still under development [83]. In the same order of ideas, establishing Good Manufacturing Practice guidelines for exosome production is crucial for ensuring product quality and safety.
Understanding the
Currently, bioengineering approaches offer exciting possibilities for exosome modification and optimization. Engineering exosomes with specific surface ligands or coatings can enhance their targeting ability and cargo-loading efficiency, enabling precise therapeutic interventions [86]. It is very interesting to think about combined therapies that include exosomes with other treatment modalities. Combining exosomes with drugs, gene therapy, or nanoparticles holds promise for synergistic effects and improved treatment outcomes. The synergistic potential of these combination therapies must be thoroughly investigated in preclinical and clinical studies.
Due to the variability in individuals, the goal of medicine in the present and into the future is to implement personalized treatments. The ability to derive patient-specific exosomes and make therapeutic approaches accordingly is an area of significant interest too. Personalized exosome-based therapies may enhance treatment efficacy and reduce the risk of adverse reactions. Preconditioning of donor cells, through either genetic modifications or exposure to specific stimuli, can influence the cargo content and therapeutic efficacy of exosomes [87]. These strategies could enhance the therapeutic potential of exosomes for reperfusion injuries.
To establish a future where exosomes are properly used in health, it is important to establish regulatory considerations for their correct commercialization. The regulatory landscape for exosome-based therapies is complex and dynamically evolving [88, 89]. Close collaboration between researchers, clinicians, and regulatory agencies is essential to establish appropriate guidelines for preclinical testing, clinical trials, and eventual approvals. In the same way, it is critical to protect intellectual property rights to incentivize investment in exosome research and development. Clear guidelines and policies related to patenting and licensing of exosome-based therapies are necessary to advance innovation.
7. Conclusions
Exosomes represent a promising avenue for addressing the challenges posed by reperfusion injuries, providing novel and targeted therapeutic options. However, several hurdles, including isolation standardization, cargo loading, scalability, safety, and regulatory approvals, must be overcome to realize their full potential. Emerging trends in engineering, combination therapies, personalized medicine, and
Acknowledgments
We would like to express our gratitude to all those who have contributed to the creation of this book chapter. We extend our appreciation to all scientists of CIBO and CIATEJ for their valuable collaboration. And our families to be always with us, thanks.
Nomenclature
reactive Oxygen Species | |
neutrophil extracellular traps | |
mesenchymal Stem Cells | |
heat shock protein | |
tumor susceptibility gene | |
multivesicular bodies | |
intraluminal vesicles | |
endosomal sorting complex required for transport | |
messenger RNA | |
microRNA | |
long noncoding RNA | |
circular RNA | |
PIWI-interacting RNA | |
epithelial–mesenchymal communication | |
tubular epithelial cells | |
calcium-sensing receptor | |
ischemic preconditioning | |
sphingosine 1-phosphate | |
blood-brain barrier | |
blood urea nitrogen |
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