Therapeutic potential of exosomal lncRNAs derived from stem cells in wound healing: focusing on mesenchymal stem cells

Abstract

The self-renewal ability and multipotency of stem cells give them great potential for use in wound healing. Stem cell-derived exosomes, owing to their close biological resemblance to their parent cells, offer a more efficient, safer, and economical approach for facilitating cellular communication and interactions within different environments. This potential makes them particularly valuable in the treatment of both acute and chronic wounds, such as lacerations, burns, and diabetic ulcers. Long non-coding RNAs (lncRNAs) enclosed in exosomes, as one of the leading actors of these extracellular microvesicles, through interaction with miRNAs and regulation of various signaling pathways involved in inflammation, angiogenesis, cell proliferation, and migration, could heal the wounds. Exosome-derived lncRNAs from stem cells facilitate extracellular matrix remodeling through interaction between macrophages and fibroblasts. Moreover, alongside regulating the expression of inflammatory cytokines, controlling reactive oxygen species levels, and enhancing autophagic activity, they also modulate immune responses to support wound healing. Regulating the expression of genes and signaling pathways related to angiogenesis, by increasing blood supply and accelerating the delivery of essential substances to the wound environment, is another effect exosomal lncRNAs derived from stem cells for wound healing. These lncRNAs can also enhance skin wound healing by regulating homeostasis, increasing the proliferation and differentiation of cells involved in the wound-healing process, and enhancing fibroblast viability and migration to the injury site. Ultimately, exosome-derived lncRNAs from stem cells offer valuable and novel insights into the molecular mechanisms underlying improved wound healing. They can pave the way for potential therapeutic strategies, fostering further research for a better future. Meanwhile, exosomes derived from mesenchymal stem cells, due to their exceptional regenerative properties, as well as the lncRNAs derived from these exosomes, have emerged as one of the innovative tools in wound healing. This review article aims to narrate the cellular and molecular roles of exosome-derived lncRNAs from stem cells in enhancing wound healing with a focus on mesenchymal stem cells.

Graphical Abstract

Introduction

Wound healing is one of the most intricate biological phenomena that occurs throughout the human lifespan. Skin wounds are divided into acute and chronic wounds. Acute wounds include burns, cuts, trauma, needle punctures, scrapes, and surgical incisions. Chronic wounds encompass conditions such as diabetic foot ulcers, pressure ulcers, venous ulcers, arterial ulcers, infectious wounds, and ischemic wounds [1, 2]. Wound healing is a very sequential process of repair and includes steps that not only overlap in time but can also depend on each other [3, 4]. Wound healing involves inflammation, proliferation, hemostasis, contraction, and regeneration [5]. Wound healing relies on the coordination of matrix metalloproteinases (MMPs), growth factors, cytokines, inflammatory cells, fibroblasts, keratinocytes, and endothelial cells [6].

Conventional wound care methods, such as wound dressings, skin substitutes, and growth factors, have drawbacks including extended recovery times, immune system rejection, high costs, and vulnerability to infection, which constrain their use [7,8,9]. In recent years, the regenerative and restorative properties of stem cells have led to their use in wound healing [10]. From undifferentiated pluripotent stem cells to more specialized multipotent progenitor cells, they have been used to promote wound healing in numerous animal models and clinical trials [11]. However, due to the presence of reactive oxygen species (ROS) in the wound environment, stem cells often do not survive easily in the wound microenvironment, and their use in wound healing may be complex [12, 13]. Compared to stem cells, exosomes derived from stem cells offer several advantages, including non-immunogenicity, lack of infusion toxicity, ease of access, and the absence of tumorigenic risk and ethical concerns [14].

Exosomes are intracellular vesicles with size of 30–150 nm that could be released by nearly all types of cells [15]. Exosomes are commonly distributed in body fluids including serum, urine, milk, semen, cerebrospinal fluid, and saliva [16]. Stem cell exosomes have unique biological functions akin to stem cells because the stem cells are considered as their parent [17]. Stem cell-derived exosomes have tremendous therapeutic potential in all stages of the wound healing process, including reducing inflammation, and angiogenesis, and promoting the proliferation and migration of fibroblasts and epithelial cells [18,19,20]. Exosomes can encompass cytokines including interleukin-6 (IL-6), interleukin-10 (IL-10), transforming growth factor-β1 (TGF-β1), vascular endothelial growth factor (VEGF), and hepatocyte growth factor (HGF) and many other cases [21, 22]. Also, exosomes facilitate communication and material transfer between cells and comprise a noteworthy quantity of RNA types including mRNA, rRNA, tRNA, piRNA, snoRNA, snRNA, miRNA, and lncRNA involved in various biological procedures [23, 24]. Non-coding RNAs (ncRNAs) including circRNA, miRNA, and lncRNA are recognized as active components within exosomes [25].

Long non-coding RNAs (lncRNAs) are a diverse class of RNA molecules exceeding 200 base pairs in length that do not encode proteins. They primarily function in the modulation of gene methylation, activation of transcription, and influencing the progression of translation [26]. lncRNAs undergo extensive synthesis and processing within the nucleus and play their regulatory activities in the cytoplasm [27]. The secondary structure and domain of lncRNAs are their essential features. However, they lack significant conservation across different species, mediate lncRNA:DNA or lncRNA:protein interactions, and affect most of their functions (Fig. 1) [28, 29].

Fig. 1

Cellular and molecular functions of LncRNAs. These molecules as non-coding RNAs, although they do not code proteins, play key regulatory roles in the expression of genes and other cellular processes. LncRNAs activate or suppress genes by interacting with chromosomes or attracting and absorbing regulatory proteins and somehow causing transcriptional regulation. Also, they can increase their expression and strengthen the transcription process through interaction with the enhancer of genes. LncRNAs can participate in mRNA splicing and play an essential role in the formation of mature mRNA. These non-coding RNAs also prevent miRNAs from binding to the target mRNA by sponging them and preventing its degradation by miRNAs. In addition, by interacting with mRNA and preventing its degradation, they can also help maintain the stability of mRNA molecules. Another ability of LncRNAs is its direct interaction with mRNA and ribosomes to help the protein translation process

In recent years, exosomes derived from mesenchymal stem cells (MSC-Exos) have been proposed as one of the new tools in skin wound healing and scar reduction due to their unique regenerative properties [30]. These nanovesicles accelerate wound healing by carrying proteins, lipids, and non-coding RNAs [31]. lncRNAs in MSC-Exos play a pivotal role in regulating regenerative signaling pathways. MSC-Exos carrying lncRNAs can increase the proliferation of fibroblasts and facilitate wound healing by epigenetically regulating the expression of collagen and extracellular matrix genes [32, 33]. In addition, these lncRNAs can reduce inflammation by affecting inflammatory pathways and cell signaling [34]. On the other hand, by increasing the expression level of pro-angiogenic factors such as VEGF, MSC-Exos can enhance angiogenesis, contributing to wound healing and tissue regeneration [35,36,37,38]. Also, lncRNAs present in MSC-Exos can play a role in improving diabetic wound healing by enhancing mitochondrial function and reducing oxidative stress [39, 40]. Finally, the unique properties of MSC-Exos, due to the lncRNAs they carry, in regulating cellular pathways and reparative processes, can make them a highly potent and non-invasive candidate for skin wound healing. So, the purpose of this review article is to narrate the cellular and molecular roles of exosomal lncRNAs derived from stem cells for wound healing, focusing on the mesenchymal stem cells.

Methodology

To identify relevant articles, electronic databases such as PubMed, Scopus, Web of Science, and Google Scholar were searched using keywords including "stem cell", "mesenchymal stem cell", "long non-coding RNA", "lncRNA", "exosome", "exosomal lncRNAs", "molecular mechanisms and pathways", and "wound healing". English-language articles up to August 2024 were assessed. This search aimed to find articles that studied the cellular and molecular roles of stem cell-derived exosomal lncRNAs in the wound healing process. The reference list of all selected articles was carefully evaluated to avoid missing relevant papers. To ensure the comprehensiveness of the search, the titles and abstracts of each article were carefully assessed, and the authors independently determined the relevance and importance of the articles. Finally, based on the selected articles, the main titles and structure of our article were designed and adjusted. This approach helped us to provide a comprehensive and detailed review of the role of exosomal lncRNAs in wound healing.

The role of stem cell-derived exosomes in wound healing: effect of long non-coding RNAs

Mesenchymal stem cells (MSCs) from sources such as adipose tissue (ADSCs), and bone marrow (BMSCs) have shown significant potential in wound healing due to their ability to produce exosomes rich in bioactive components, including lncRNAs [30]. These MSC-derived exosomes not only facilitate cell communication but also deliver functional lncRNAs, DNA, proteins, and metabolites to wound sites, promoting tissue repair through improved blood circulation, reduced inflammation, and stimulation of resident cells like fibroblasts and endothelial cells [41].

ADSC-derived exosomes (ADSC-Exos) have demonstrated promising therapeutic potential by modulating immune responses and oxidative stress and enhancing ECM composition [42]. Notably, certain lncRNAs within ADSC-Exos, such as double homeobox A pseudogene 10 (DUXAP10) and the SAN-miR-143-3p-ADD3 axis, play essential roles in rejuvenating aged ADSCs and enhancing wound healing efficacy by regulating associated signaling pathways [38, 43].

BMSC-derived exosomes have been shown to accelerate the healing of wounds, including diabetic cutaneous wounds, by promoting angiogenesis. For example, the exosomal lncRNA KLF3-AS1 from BMSCs has been shown to increase VEGFA expression by interacting with a miRNA, which stimulates angiogenesis and supports wound repair [44].

The diverse roles of MSC-derived exosomes in wound healing highlight their ability to deliver targeted lncRNAs that regulate inflammation, extracellular matrix (ECM) remodeling, and cellular proliferation. Consequently, MSC exosomes, particularly those rich in specific lncRNAs, represent a promising and targeted approach in regenerative medicine for wound healing [45, 46].

Effect of exosomal lncRNAs on immune modulation in wound healing

Wound healing is a complex process in response to tissue damage, involving the release of chemokines, cytokines, and growth factors from injured tissue [47]. Exosomes containing lncRNAs can play a role in modulating immune responses and inflammation, thus supporting wound healing [48].

M2 macrophages, active in the later stages of wound healing and hypertrophic scar microenvironment, reduce inflammation and promote collagen deposition, which is essential for tissue repair [49]. Exosomes containing lncRNA-ASLNCS5088, derived from M2 macrophages in a TGF-β1-rich environment, promote fibroblast activation and extracellular matrix (ECM) production, likely through suppressing miR-200c-3p, which regulates glutaminase. This process identifies lncRNA-ASLNCS5088 as a potential therapeutic target for hypertrophic scars [50, 51]. Similarly, hBMSC-Exos containing lncRNA maternally expressed gene 3 (MEG3) have been shown to prevent keloid formation by reducing fibrosis-related protein and collagen expression, further aiding wound healing [33].

M1 macrophages increase ROS production, which is essential early in wound healing but can impair healing if prolonged. lncRNA GAS5 upregulates M1 markers like IL-1 and TNF-a via the STAT1 pathway, while lncRNA Lethe inhibits NFκB target genes like NOX2, exerting anti-inflammatory and antioxidative effects [48, 52].

Long non-coding RNA heart and neural crest derivatives expressed 2-antisense RNA 1 (HAND2-AS1) can be packaged in hMSC-derived exosomes and absorbed by rheumatoid arthritis (RA) fibroblast-like synoviocytes (RA-FLSs) and through inactivation of the NF-κB pathway induces apoptosis in these cells. HAND2-AS1 directly sponges miR-143-3p and positively regulates the tumor necrosis factor alpha-inducible protein 3 (TNFAIP3) expression. Exosomes derived from MSC participate in the intercellular transfer of HAND2-AS1 and suppress the RA-FLSs activation through the miR-143-3p/TNFAIP3/NF-kB axis and exert their anti-inflammatory effect [53]. ADSCs-exo-carrying lncRNA Neat1 induces the expression of Ulk1 through miR-17-5p sponging in mouse keratinocytes, which leads to the up-regulation of autophagic activity to promote wound healing. Therefore, the improved autophagy pathway through Neat1/miR-17-5p/Ulk1 axis could enhance the wound healing process [54]. In a study, GAS5 lncRNA-enriched ADSC-Exos in human dermal fibroblast cells treated with lipopolysaccharide in a low-grade chronic inflammatory environment were shown to regulate toll-like receptor 7 (TLR7) levels as well as by affecting the genes of the inflammatory pathway, they accelerate the healing of resistant chronic wounds [34] (Fig. 2).

Fig. 2

Wounds healing by exosomes derived from adipose stem cells containing lncRNA. Adipose stem cells, as pluripotent cells with high self-renewal capability, can play a prominent role in wound healing by releasing exosomes containing specific cellular contents, including LncRNAs. Among the most known and effective LncRNAs present in these exosomes are NEAT1, XIST, GAS5, H19, MALAT1, SENCR, and Linc00511, each of which contributes to the improvement of wounds through their unique molecular pathways. NEAT1 promotes the expression of UIL1 by inhibiting miR-17-5p and thus increases autophagic activity. MALAT1 increases cell migration and angiogenesis by inhibiting miR-124 and interacting with the Wnt/β-catenin pathway. Also, by inhibiting miR-374a-5p, MALAT1 improves mitochondrial function and inhibits apoptosis in fibroblasts. Furthermore, MALAT1 promotes the proliferation and migration of human skin fibroblasts (HSFs) by inhibiting miR-378a and increasing the expression of FGF2, and H19 enhances the proliferation and migration of HSFs by inhibiting miR-29b and increasing the expression of FBN1, and inhibiting miR-19b and increasing the expression of SOX9, then interacting with the Wnt/β-catenin pathway. H19 also supports fibroblast proliferation, migration, and angiogenesis by activating STAT3. SENCR promotes angiogenesis in damaged tissue by increasing the expression of VEGF-A and Linc00511 by inhibiting PAQR3, which increases TWIST1. XIST increases DDR2 expression and proliferation and migration in mouse dermal fibroblasts (MDFs) cells by inhibiting miR-96-5p. GAS5 accelerates wound healing by reducing inflammation through interaction with TLR7 and regulation of inflammatory pathway genes

Regulation of angiogenesis in wound healing by stem cell-derived exosomal lncRNAs

Angiogenesis is crucial for the regeneration of skin tissue during wound healing, facilitating the delivery of growth factors, oxygen, and nutrients to the injured area [55]. The angiogenesis can be regulated by various lncRNAs and miRNAs [56]. Sun et al. reported that hADSC-Exos could neutralize the inhibitory effect of short-hairpin RNA SENCR (shSENCR) on angiogenesis and accelerate wound healing. The hADSC-Exos enhance the expression of early growth response factor-1 (EGR-1), which binds to the lncRNA-SENCR promoter, causing its overexpression and thus increasing the expression of VEGF-A and suppressing the inhibitory impact of shSENCR on human umbilical vein endothelial cells (HUVECs). Therefore, it can be concluded that hADSC-Exo EGR-1 positively regulates the expression of Dyskerin pseudouridine synthase 1 (DKC1) or lncRNA-SENCR that interact with each other to promote angiogenesis in the wound healing process by activating VEGF-A [57] (Fig. 2).

Recently, it has been reported that HOX transcript antisense intergenic RNA (HOTAIR), an exosomal lncRNA from endometrial stromal cells (ESCs), can competitively downregulate miR-761 both in vitro and in vivo, upregulate histone deacetylase 1 (HDAC1), and promote angiogenesis by activating STAT3-associated pro-inflammatory cytokines [58]. A study on human normal wound and burn tissues and HUVECs showed that lncRNA HOTAIR sponged miR-126, which is highly expressed under heat stress, and indirectly downregulates Sciellin (SCEL) as a target of miR-126. Therefore, it can be stated that HOTAIR by regulating the miR-126/SCEL axis can increase angiogenesis in the healing process of wounds and burns [59, 60]. Supporting these results, Born et al. reported that LncRNA HOTAIR overexpressed in MSCs through upregulation of the VEGF gene significantly promotes angiogenesis in endothelial cells and accelerates diabetic wound healing in db/db mice [35].

Exosomes derived from HUVECs exposed to H₂O₂ oxidative stress significantly enhanced the pro-angiogenic capacity of endothelial progenitor cells (EPCs) via the Wnt/β-catenin pathway regulated by Lnc NEAT1, thereby enhancing random flap survival in vivo. Thus, the H₂O₂-HUVEC-Exos might serve as an alternative therapeutic procedure to improve the survival of the randomized skin flap [61]. In ADSC cells under hypoxia, inhibition of LINC02913 through direct binding of hypoxia-inducible factor-1alpha (HIF1A) to its promoter region and interaction with insulin-like growth factor 1 receptor (IGF1R), activated PI3K/AKT pathway and increased proliferation and angiogenic ability in the wound area [62].

Human amniotic mesenchymal stem cells (hAMSC)-Exos lncRNAs related to angiogenesis include PANTR1, H19, OIP5-AS1, and NR2F1-AS1 [63]. ADSCs-Exos carrying lncRNA H19 promote M2 macrophage polarization, boosting fibroblast proliferation, migration, and angiogenesis of endothelial cells (ECs). miR-130b-3p, which directly targets STAT3 or PPARγ, participates as a downstream effector in H19-mediated biological impacts. Therefore, the H19 in exosome derived from ADSCs accelerates skin wound healing through the miR-130b-3p/PPARγ/STAT3 pathway by increasing fibroblast proliferation, migration, and angiogenesis of endothelial cells [64] (Fig. 2). Also, confirming previous findings in a study that used extracellular vesicle-mimetic nanovesicles (EMNVs), it was demonstrated that H19EMNVs possess a potent capability to counteract the regeneration-inhibiting impacts of hyperglycemia, significantly speeding up chronic wound healing [65]. Shi et al. reported that lncRNA OIP5-AS1 induces angiogenesis by regulating miR-3163/VEGFA [66]. Exosomes highly enriched in the lncRNA OIP5-AS1 regulate angiogenesis through the OIP5-AS1/miR-153/ATG5 axis [67]. lncRNA NR2F1-AS1 increased the expression of insulin-like growth factor-1 (IGF-1) by sponging miR-338-3p, and then by regulating the IGF-1R receptor and extracellular signal, it activated the ERK signaling pathway in HUVECs and caused angiogenesis occurs. Therefore, NR2F1-AS1 may promote angiogenesis via the IGF-1/IGF-1R/ERK axis [68].

The function of exosomal lncRNAs in the regulation of cell proliferation and migration throughout wound healing

Cell migration is essential to maintain suitable organization in multicellular organisms. Proper morphogenesis could be considered, in part, as a result of cell migration. In an adult organism, cell migration is crucial for an effective immune response, wound healing, and maintaining tissue homeostasis [69]. Fibroblasts have a crucial role in wound healing, and by migrating into wounds, they repair skin damage [70]. ADSC-derived exosomes boost wound healing by stimulating the migration of skin fibroblasts [71]. Cooper et al. reported that ADSC-derived exosomes containing MALAT1 can stimulate the migration of human dermal fibroblasts (HDFs) and angiogenesis in mouse ischemic wounds. In vitro, nuclear MALAT1 can increase cell migration in scratch assays. In fact, hADSCs and exosomes containing MALAT1 increase cell migration to heal ischemic wounds and make it possible to use the paracrine signaling abilities of stem cells without using cells [72]. The lncRNA MALAT1 as a sponge miRNA is also involved in cell migration and wound healing [73]. MALAT1 present in ADSC-Exos activates the Wnt/β-catenin signaling pathway by binding to miR-124 to accelerate skin wound healing [74]. Pi et al. reported that overexpression of miR-378a decreases the expression of FGF2 and prevents the migration and proliferation of human skin fibroblasts (HSFs). However, ADSC-derived exosomal lncRNA MALAT1 upregulates FGF2 through sponging miR-378a [36]. Therefore, it can be deduced that MALAT1 promotes the proliferation and migration of HSFs by manipulating the miR-378a/FGF2 axis and leads to wound healing [75] (Fig. 2).

LncRNA H19 in ADSC-Exos could control the SOX9 expression via miR-19b to accelerate the proliferation, migration, and invasion of HSFs and accelerate the healing of skin wounds by activating the Wnt/β-catenin pathway [76]. Upregulation of lncRNA-H19 could also enhance the FBN1 expression by competitively binding to miR-29b, thereby increasing the proliferation and migration of fibroblasts and inhibiting their apoptosis, ultimately accelerating wound healing [77] (Fig. 2).

Exosomes derived from bone marrow mesenchymal stem cells (BMMSCs) can prevent tissue damage by regulating the progression of sepsis. Exosomes derived from BMMSCs, in which KCNQ10T1 lncRNA is overexpressed, sponge miR-154-3p and, by affecting RNF19A, which is the downstream target gene of miR-154-3p, inhibit cell proliferation, metastasis, and inflammatory response caused by LPS and prevents the development of sepsis [78]. Over-expressed lncRNA FOXD2-AS1 in ADMSC exosomes enhances HaCaT cell proliferation and migration by down-regulating miR-185-5p and up-regulating rho-associated coiled-coil-containing protein kinase 2 (ROCK2). In summary, the overexpression of LncRNA FOXD2-AS1 in exosomes derived from ADMSCs may significantly promote HaCaT cell migration and proliferation through modulation of the miR-185-5p/ROCK2 signaling pathway [79]. Synthesis of ceramide, through regulating MALAT1 exosomal biogenesis and packaging derived from ADSC, can promote skin fibroblast migration and enhance mitochondrial function, potentially leading to improved wound healing [80]. Xiong et al. showed that a lncRNA known as NORAD, enriched in exosomes derived from ADSC, can modulate the wound microenvironment under hypoxic conditions. in fact, NORAD enhances fibroblast proliferation and migration by interacting with miR-524-5p and influencing the activity of key proteins like Pumilio, thereby promoting wound repair [81].

ADSC-exosome-loaded X-inactive-specific transcript (XIST) binds to miR-96-5p and, by inhibiting it, induces discoidin domain receptor 2 (DDR2) mRNA expression. The silencing of miR-96-5p or overexpression of DDR2 significantly affects the proliferation and migration of mouse dermal fibroblasts (MDFs) and accelerates wound healing [82] (Fig. 2).

lncRNAs of exosomes derived from stem cells and diabetic wounds healing

Diabetes, a prevalent metabolic disease worldwide, in addition to increasing blood glucose, can cause severe mental, physical, and financial burdens for diabetic people by bringing other severe complications such as severe foot ulcers, gangrenous wounds, and even amputation [83]. The healing of skin wounds in diabetes is impaired and can turn into non-healing wounds [84]. However, treatments utilizing exosomal lncRNAs offer promising prospects for the future. Exosomes derived from adipose progenitor cells can effectively restore damaged diabetic wound healing by delivering lncRNA-H19 to the damaged tissue. Cutaneous platelet-derived growth factors (PDGFs) fibroblast-expressing lncH19 accelerates wound healing by promoting the proliferation of dermal fibroblasts and infiltration of macrophages into damaged skin. There is a lower level of PDGFRα + cell-derived lncH19 in the skin tissue of patients and mice with type 2 diabetes. lncH19 reduces the cell cycle arrest of fibroblasts and by inhibiting p53 activity and releasing GDF15, increases the infiltration of macrophages in damaged tissues [85]. Notably, since lncRNA H19 is abundantly found in exosomes derived from MSCs, it is suggested that MSC-derived exosomal lncRNA may similarly be able to play a therapeutic role by regulating macrophage infiltration and fibroblast proliferation in diabetic wounds.

In the fibroblasts of patients with diabetic foot ulcers (DFU), the expression of miRNA-152-3p is increased, and the expression of PTEN is decreased, which makes it susceptible to diabetic nephropathy by disrupting glomerular function. MSC-released exosomal lncRNA H19 by binding and inhibiting miR-152-3p and increasing the expression of PTEN increases the proliferation and migration of fibroblasts. It inhibits apoptosis in them, thus reducing the damage caused by DFU and accelerating the wound healing process [40, 86]. Overexpression of connective tissue growth factor (CTGF) activates the MAPK pathway to enhance cell proliferation, angiogenesis, ECM remodeling, and wound healing while preventing cell apoptosis. By recruiting serum response factor (SRF) to the CTGF promoter region, lncRNA H19 enhances its expression. It activates the MAPK pathway, thereby accelerating cell proliferation, ECM regeneration, wound healing, and suppression of cell apoptosis. Therefore, lncRNA H19 not only accelerates wound healing in DFU via the upregulation of CTGF and activation of the MAPK pathway but also reflects a broader potential mechanism by which stem cell-derived lncRNAs regulate similar pro-regenerative pathways in diabetic wounds [87]. Hair follicle mesenchymal stem cell exosomal lncRNA H19, by increasing the proliferation and migration of HaCaT as well as reversing the stimulation of NLRP3 inflammasome, which inhibited pyroptosis, thereby improved the skin wound of diabetic mice [88] (Fig. 3). This shows that lncRNA H19 has the potential to play a key role in wound healing, but more studies are needed to understand its exact involvement in stem cell-derived exosomes during DFU.

Fig. 3

LncRNAs involved in diabetic foot ulcer healing. Adequate blood supply to the damaged tissue in diabetic foot ulcers is one of the most essential wound-healing processes, which is promoted by lncRNA H19, LINC01435, MALAT1, and linc00511. LncRNA H19 supports angiogenesis through histone methylation mediated by the Enhancer of Zeste Homolog 2 (EZH2) and modulation of the HIF-1α signaling pathway. MALAT1 enhances VEGF expression, Linc00511 reduces PAQR3 and increases Twist1, and the reduction of LINC01435 leads to decreased nuclear translocation of YY1, causing knockdown of HDAC8, thereby supporting angiogenesis. To regenerate the damaged tissue and return to its normal morphology, some cells involved in wound healing need to migrate. lncRNAs H19, MALAT1, and CASC2 promote the migration of these cells. H19 does this by inhibiting miR-152-3p and increasing PTEN expression. MALAT1 supports migration through the inhibition of miR-374a-5p, and CASC2 by inhibiting miRNA 155, both enhancing HIF-1α expression and supporting cell migration and proliferation. The regeneration of ECM and extracellular space is another primary step in the wound healing, which can be related to lncRNAs URIDS, MALAT1, and H19. Reducing URIDS promotes collagen production by increasing Plod1 protein stability. MALAT1 accelerates the expression of MFGE8 through inhibition of miR-1914-3p. By recruiting SRF to the CTGF promoter region, H19 increases its expression and accelerates cell proliferation, ECM regeneration, and diabetic foot ulcer healing. To properly heal wounds, inflammation must be regulated alongside other wound healing-promoting factors. LncHAR1B supports diabetic foot ulcer healing by increasing BHLHE23, which regulates KLF4, a gene that inhibits inflammation

BMSCs-derived exosomal lncRNA KLF3-AS1 is a competitor for miR-383 and, by inhibiting it, enhances VEGF-A signaling to induce angiogenesis and diabetic wound healing process. In diabetic conditions, exosomal KLF3-AS1 significantly promotes the HUVECs proliferation and up-regulates pro-apoptotic proteins, while anti-apoptotic proteins are reduced due to the increased expression level of VEGF-A. Interestingly, the presence of KLF3-AS1-Exos also suppresses inflammation, which causes more and faster wound healing [44]. ADSC exosomes by overexpressing linc00511 restore the biological capability of EPCs and accelerate angiogenesis by preventing PAQR3 degradation caused through Twist1 ubiquitination, thus improving DFU [89].

HaCaT-derived exosomal LINC01435 might control vascular endothelial cell function and affect diabetic wound healing. Exososom from high glucose-pretreated immortalized human epidermal cells (HG-Exos), which are rich in LINC01435, can increase the expression of histone deacetylase 8 (HDAC8) in endothelial cells by inducing the translocation of transcription factor Yin Yang 1 (YY1) to the nucleus. As a result, this inhibits tube formation and the migratory ability of endothelial cells, ultimately impairing angiogenesis. To positively regulate wound healing, HDAC8 knockdown can enhance tube formation and HUVEC migration. However, simultaneous LINC01435 overexpression can prevent these procedures. These outcomes suggest that LINC01435/YY1/HDAC8 axis might be an essential signaling pathway that affects diabetic wound healing [90]. Guo et al. showed that lncRNA H19 accelerates fibroblast activation by enhancing histone methylation mediated by enhancer of zeste homolog2 (EZH2) and regulating the HIF-1α pathway. This, in turn, enhances the process of modified preservative fluid-preserved autologous blood and improves postoperative wound healing in diabetic mice [91]. Exosomal human keratinocyte-derived MALAT1 may induce the deposition of collagen, remodeling of ECM, and expression of CD31, VEGF, and MFGE8, but downregulate TGFB and SMAD3 expression. MALAT1 accelerates the MFGE8 expression via binding competitively to miR-1914-3p, influencing the macrophage function and the TGFB1/SMAD3 axis, and ultimately causing the diabetic wounds to heal [92]. By overexpressing lncRNA cancer susceptibility candidate 2 (CASC2) in fibroblasts, it can quickly heal DFU wounds. Through interaction with miRNA 155, lncRNA CASC2 upregulates the HIF-1α gene, prevents fibroblast apoptosis, promotes cell proliferation and migration, and facilitates wound repair. Therefore, increasing the level of HIF-1α by lncRNAs can be regarded as an influential approach to the wound healing process [93]. The MALAT1 overexpression or miR-374a-5p knockdown can enhance fibroblasts viability and suppress pyroptosis and apoptosis in DFU. Overexpression of miR-374a-5p counteracts the effect of overexpression of platelet-rich plasma (PRP-Exos) or MALAT1 on cell viability, apoptosis, and pyroptosis. In general, it can be deduced that the MALAT1-mediated signaling axis plays a role in the function of PRP-exosomes in promoting DFU wound healing [39] (Fig. 3). The lncRNA Upregulated in Diabetic Skin (lnc-URIDS) interacts with procollagen-lysine, 2-oxoglutarate 5-dioxygenase 1 (Plod1), a key enzyme in collagen cross-linking. This interaction reduces the stability of the Plod1 protein, resulting in impaired collagen production and deposition; consequently, wound healing is delayed [94]. Overexpression of lncHAR1B leads to upregulation of basic helix-loop-helix family member E23 (BHLHE23), and its knock-out causes downregulation of BHLHE23. Also, the expression level of Kruppel-like factor 4 (KLF4) as a target gene for the transcription factor BHLHE23 is regulated after its binding with lncHAR1B. KLF4 plays a central role in diabetic wound healing by inhibiting inflammation and modulating their polarization towards M2 macrophages [95,96,97,98] (Fig. 3). While some of these lncRNAs have not yet been studied in the context of stem cell-derived exosomes in wound healing, their involvement in similar pathways suggests a potential role that warrants further exploration in stem cell-based therapeutic strategies. A summary of the role of exosomal lncRNAs derived from stem cells in wound healing is presented in Table 1.

Engineered exosomes derived from stem cells and their application in wound healing

By using advanced bioengineering technologies, it is possible to modify exosomes so that new capabilities of expressing specific markers or carrying therapeutic cargo are provided. By turning exosomes into new platforms for treating complex diseases and challenging regenerative medicine, these changes have created a promising prospect for healing damaged tissues and organs [37, 99]. Engineered exosomes modified with surface decoration or internal therapeutic molecules can treat many diseases [100]. Although these exosomes do not differ significantly from natural exosomes in size or shape, the cargo or content placed in them may differ based on the purpose of their synthesis [99]. Modifying the carrying capacity of exosomes or loading therapeutic cargo with different and new methods will ultimately improve their targeting ability and therapeutic efficiency [101]. Stem cell-derived exosomes can be engineered through direct, indirect, and synthetic methods including parent cell surface modification, co-incubation, genetic engineering, and artificial synthesis [37]. These engineered exosomes have improved therapeutic properties for precise targeting, therapeutic cargo loading, bioavailability, and high efficiency, and they show multiple and effective functions in tissue regeneration and wound healing. These exosomes improve collagen synthesis and have anti-inflammatory, pro-angiogenic, and epithelialization properties. They also facilitate specific pathological pathways that underlie resistant DFUs, including peripheral neuropathy, macro- and micro-angiopathy, persistent inflammation, infection, and re-epithelialization [102].

Engineered stem cell exosomes, by targeting specific anti-inflammatory signaling pathways, affect multiple biological activities involved in wound healing, including diabetic wounds. These exosomes can accelerate wound healing by reducing the inflammatory response and inhibiting the secretion of pro-inflammatory factors [37, 103]. In a study, it has been shown that using MSC exosomes modified with melatonin can reduce inflammation in diabetic wounds by increasing PTEN expression and inhibiting AKT phosphorylation by affecting the PTEN/AKT signaling pathway [18]. Also, treatmenting these exosomes with lipopolysaccharide can influence the TLR4/NF-κB/STAT3/AKT signaling pathway through miRNA let-7b and improve diabetic wounds by reducing chronic inflammation [104]. Exosomes derived from MSCs have been shown to reduce the release of pro-inflammatory cytokines and eventually inflammation when stimulated by inflammatory factors such as TNFα and IFNγ [105]. Also, overexpression of transcription factors including Nrf2 in engineered exosomes derived from ADSCs inhibits the production of ROS and inflammatory cytokines in the direction of diabetic wound healing [106].

Using engineered exosomes derived from stem cells can promote the healing of skin wounds by increasing the proliferation of fibroblasts [107, 108]. MSC-derived exosomal lncRNA H19 inhibited fibroblast apoptosis and inflammation by disrupting miR-152-3p's inhibition of PTEN, which enhanced the wound-healing process in diabetic foot ulcers [40]. Also, artificially synthesized exosomes can be absorbed by recipient cells and cause wound healing. It has been observed that nanovesicles extracted from embryonic stem cells promote the proliferation of fibroblasts and wound healing by activating the MAPK signaling pathway [109].

Engineered exosomes of stem cells can increase the proangiogenic ability of vascular endothelial cells by absorbing miRNA and cytokines [37]. Blue light treatment has been shown to ultimately increase proliferation, migration, and angiogenesis in HUVECs by regulating the levels of miR-135b-5p and miR-499a-3p in hUC-MSC-derived exosomes [110]. Exosomes treated with superparamagnetic ferric oxide accumulate in the damaged area with precise targeting and significantly increase angiogenesis [111]. Wu et al. reported that BMSCs-derived exosomes stimulated by iron oxide (Fe₃O₄) and static magnetic field targeted SPRY2 by upregulating miR-21-5p and activating PI3K/AKT and ERK1/2 involved in wound healing, enhanced angiogenesis [112]. In addition, artificially synthesized exosomes can significantly enhance angiogenesis with specific protein composition and RNA load [65, 113].

Engineered exosomes can also effectively increase the endogenous secretion of growth factors [102]. Exosomes derived from MSCs loaded with an inhibitor of miR-155 can restore FGF-7 levels and increase the migration of keratinocytes, ultimately accelerating wound healing through the negative regulation of miR-155 [114]. Engineered exosomes loaded with RL-QN15 peptide can express vascular endothelial growth factor B. They can also promote migration and proliferation of HaCaT cells in a high-glycemic environment and promote angiogenesis and re-epithelialization of DFUs [115, 116].

Challenges and limitations of exosomal lncRNAs derived from stem cells in wound healing

Exosomes derived from stem cells, especially MSCs, due to their unique characteristics in easy transfer of cell contents from one cell to another, low immunogenicity, and high biocompatibility, have been proposed as a new and promising tool in the treatment of complex diseases in regenerative medicine. However, using these exosomes faces various challenges, such as different exosome isolation methods affecting their density, shape, and surface markers, absorption into other body tissues, and relatively sensitive storage conditions [117,118,119,120,121,122]. lncRNAs enriched in MSC-derived exosomes have also been proposed as one of the useful therapeutic tools due to their important role in tissue healing and regeneration processes, but their use is limited [45]. Among these limitations is the tissue specificity of lncRNAs. In fact, lncRNAs are expressed differently in various types of diabetic and burn wounds, so their use in one type of wound may not be efficient in other types of wounds [123, 124]. Another limitation of using lncRNAs is the incomplete understanding of their exact mechanisms in wound healing processes. These molecules implement various mechanisms, many of which are still unknown. Therefore, understanding their exact functioning at each stage of wound healing is essential [125].

Since exosomes are considered as a whole, containing a mixture of different proteins, nucleic acids, and other components, it is challenging to investigate the precise and exclusive role of their lncRNAs. However, their detailed investigation in studies, especially the use of lncRNAs exclusive assay methods in related studies, can determine their role in molecular and cellular mechanisms. For example, the studies discussed in this review, most of them used techniques such as quantitative real-time polymerase chain reaction [53] and the use of specific siRNAs for the downregulation of lncRNAs [34], which indicates the exclusive role of lncRNAs in wound studies. Also, bioengineered exosomes that contain specific lncRNAs would determine the more precise role of lncRNAs in wound healing. However, due to the complexity of these molecules and their interactions with other genetic and protein molecules, identifying the exact function of each lncRNA is challenging and requires further research. For example, MSC-derived exosomal lncRNAs can have opposite effects in various biological processes [126,127,128,129].

On the other hand, the presence of destructive enzymes such as RNases can reduce the stability of lncRNAs and limit their efficiency. Therefore, accurate and efficient delivery of lncRNAs to the wound site is one of the main challenges in the clinical use of these molecules [130]. The existing delivery systems may not be able to deliver lncRNAs accurately and targeted to the damaged tissue. Hence, studying and improving lncRNA delivery methods can be very helpful [127]. In addition, the lack of advanced technologies for precisely regulating the timing and location of lncRNA expression leads to their premature or delayed expression, which can result in undesirable and unintended outcomes [127, 131]. For example, the source of MSCs can have diverse effects on the function and expression of lncRNAs, and this expression may vary according to the type of tissue and timing. Unbalanced expression of these lncRNAs may lead to various diseases and even cancer. Also, the cultural conditions and different passages of MSCs in the laboratory environment affect their fundamental characteristics and performance. These conditions can change the behavior of cells and cause inconsistency in the function of lncRNAs [132]. Finally, to fully utilization of the therapeutic potential of lncRNAs enriched in MSC-derived exosomes, more extensive research and more advanced molecular technologies are needed to identify their specific targets and pathways as well as protein interactions. Despite these challenges, these lncRNAs can still be considered as high-value diagnostic and medicinal targets for the treatment and diagnosis of diseases [133].

Future perspectives

Wound healing, as an increasing challenge, is a clinically demanding problem, making the proper and efficient management of wound healing essential [134]. Biological behaviors of critical cells in different stages of wound healing can change this process under the influence of molecular inhibitors and genetic changes [135]. Stem cell-derived exosomes, carrying lncRNAs, are one of the main factors altering the biological behaviors of cells involved in wound healing [2]. Although the use of lncRNA-containing exosomes has shown promising results, it should be noted that experimental treatments for wound healing have primarily been performed in rodent models. Due to the noteworthy morphological and structural differences between human skin and rodent skin, these results cannot be fully extrapolated to human wounds [136]. Therefore, to achieve reliable results, it is strongly recommended to conduct human clinical studies with large sample sizes to confirm previous findings. Achieving fruitful and promising results of in vivo and in vitro research on using exosomes derived from stem cells in wound healing can make this type of treatment a suitable candidate for use in clinics. In the future, by targeting distinct pathways involved in the wound healing process, each patient can be treated according to their specific molecular and genetic profile [137]. Exosomes become valuable due to the lncRNAs they carry and can exert their effectiveness on the healing of wounds. In pursuit of a better future, further research in wound healing may lead to more precise and effective treatments by fostering a deeper and more comprehensive understanding of the underlying molecular mechanisms, thereby minimizing risks, hazards, and potential errors [138]. In the future, exosomes derived from stem cells as a low-cost but long-term effective approach can be available to all patients as a novel and advanced treatment. It is the lncRNAs within exosomes that confer these properties by influencing wound healing. On the other hand, advancements in innovative technologies today, facilitated by the emergence of gene editing technology and the development and enhancement of methods for delivering exosomal lncRNAs to wound sites, can create a promising and hopeful outlook for regenerative medicine and wound healing in the future, increasing the precision and efficiency of treatments [139]. Ultimately, exosomes derived from stem cells, due to their unique contents, particularly various lncRNAs, represent an innovative, pioneering, and promising therapeutic approach to wound healing. They hold promise for significant advancements in this field and have the potential to impact the lives of these patients greatly.

Table 1 The role and effect of lncRNAs enclosed in exosomes derived from stem cells in the wound healing

Conclusion

The emergence of stem cells, particularly MSCs, as effective tools in regenerative medicine has enabled innovative strategies, which have shown remarkable efficiency in wound healing. In addition to their structural and transportive roles, the exosomes of these cells contain a set of bioactive molecules, especially lncRNAs, which play an essential role in the wound regeneration process. Key signaling related to inflammation, angiogenesis, cell proliferation and migration play a significant role in wound healing. Through complex interactions with miRNAs, cytokines, and other molecular regulators, lncRNAs induce changes in gene expression and cellular responses that optimize the immune system and regulate oxidative stress and tissue regeneration. These effects make exosomal lncRNAs of stem cells as a strong factor in wound healing, supporting fibroblast survival, cell differentiation, and ECM regeneration, thus enabling effective and sustainable tissue repair. To advance these studies, it is necessary to fully elucidate the molecular mechanisms and pathways through which exosomal lncRNAs act. Understanding these interactions can aid in developing targeted therapeutic strategies, especially for chronic and non-healing wounds, and lead to more personalized and precise treatments. With the expansion of knowledge about the roles of exosomal lncRNAs derived from stem cells, particularly MSCs, the potential of these extracellular vesicls to revolutionize wound healing and broader applications in regenerative medicine continues to grow. Continuous and innovative research in this field promises a bright future.