Orchestration of Mesenchymal Stem/Stromal Cells and Inflammation During Wound Healing

Abstract

Wound healing is a complex process and encompasses a number of overlapping phases, during which coordinated inflammatory responses following tissue injury play dominant roles in triggering evolutionarily highly conserved principals governing tissue repair and regeneration. Among all nonimmune cells involved in the process, mesenchymal stem/stromal cells (MSCs) are most intensely investigated and have been shown to play fundamental roles in orchestrating wound healing and regeneration through interaction with the ordered inflammatory processes. Despite recent progress and encouraging results, an informed view of the scope of this evolutionarily conserved biological process requires a clear understanding of the dynamic interplay between MSCs and the immune systems in the process of wound healing. In this review, we outline current insights into the ways in which MSCs sense and modulate inflammation undergoing the process of wound healing, highlighting the central role of neutrophils, macrophages, and T cells during the interaction. We also draw attention to the specific effects of MSC-based therapy on different pathological wound healing. Finally, we discuss how ongoing scientific advances in MSCs could be efficiently translated into clinical strategies, focusing on the current limitations and gaps that remain to be overcome for achieving preferred functional tissue regeneration.

Graphical Abstract

Graphical Abstract.

Significance Statement.

In this review, we outline current insights into the ways in which MSCs sense and modulate inflammation in the process of wound healing, highlighting the central role of neutrophils, macrophages, and T cells during the interaction. We also draw attention to the specific effects of MSC-based therapy on different pathological wound healing. Finally, we discuss how ongoing scientific advances in MSCs could be efficiently translated into clinical strategies, focusing on the current limitations and gaps that remain to be overcome for achieving desired tissue regeneration.

Introduction

Wound healing is dynamic but a fine-tuned biological process in which damaged tissue acquires the ability to regenerate itself following impairment. Under a desirable condition of skin wound, this process undergoes distinct but overlapping well-staged phases of inflammation, cell proliferation, and remodeling,¹ which involves a complex interplay between many cellular players of the skin, comprising primarily keratinocytes, fibroblasts, endothelial cells of vessels, and recruited immune cells, and their associated extracellular matrix (ECM).^2,3 During the early phase of wound healing, various types of leukocytes, including neutrophils, macrophages, and T cells, are attracted to the site of injury by chemotactic signals. Neutrophils play a crucial role in eliminating bacteria and debris from the wound, while macrophages are responsible for removing dead cells and other debris and producing growth factors that promote healing. T cells become activated in response to cytokines and chemokines released by other cells such as macrophages and dendritic cells. Once activated, T cells can produce cytokines and growth factors that promote the proliferation of other cells involved in wound healing, such as fibroblasts and keratinocytes. During the proliferative phase of wound healing, fibroblasts are activated and differentiate into myofibroblasts to reconstruct the wound. ECM deposition, especially collagen triggers the re-epithelialization process.⁴ Keratinocytes are the primary cells involved in re-epithelialization and are stimulated to migrate and proliferate during this phase. Once they arrive at the wound site, keratinocytes divide and migrate over the wound bed to close the wound. Endothelial cells are also involved in the proliferative phase, as they aid in the formation of new blood vessels through a process called angiogenesis which is critical for providing the new tissue with essential nutrients and oxygen. Overall, these cell types work together during the proliferative phase to repair the damaged tissue and restore normal tissue function. Finally, the collagen fibers reorganize from collagen types III to I, and the tissue remodels, slowly gaining strength and flexibility by promoting epithelialization and neovascularization.^5-7 However, the complex healing process of tissue repair and regeneration after the damage is still far from fully understood. Many risk factors, limitations of effective treatment, and associated pathological processes still cause deficient wound closure, and even the failure of the conserved wound healing cascade often leads to pathological fibrosis, which could lead to malignant transformation.

The application of MSC-based cell therapy in the treatment of wounds is currently an active area of investigation, providing a promising opportunity for those patients with refractory wounds. MSCs, identified as mesenchymal stem/stromal cells, have multi-lineage differentiation potential and immune-modulatory properties. They exist in almost all tissues and play an essential role in tissue regeneration and closely interact with cells of the immune system in the tissue microenvironment during the repair of damaged tissues. Technically, MSCs can be derived and amplified from several sources, including the umbilical cord, bone marrow, fat pad, and other tissues. These impressive characteristics make them a promising cell type for cell therapy.

A series of studies have shown that MSCs are able to migrate into injured tissue sites and contribute to wound repair by promoting the generation and differentiation of multiple skin cell types.^1,8,9 Cell tracking and fate tracing of transplanted exogenously expanded MSCs have demonstrated that those cells can differentiate into fibroblasts, endothelial cells, keratinocytes, and skin appendages, and even form glandular structures and hair follicles, subsequently participating in cutaneous wound healing and hair regeneration.^8,10 However, the proportion of transplanted MSCs differentiated is very small and the functionality of the differentiated cells remains unknown, especially when allogeneic MSCs are used. The biggest question is what is the role of the short-lived infused MSCs in promoting tissue regeneration during wound healing.

Of note, inflammation is the first step in wound healing, and a moderate inflammatory response is beneficial for wound repair. Some pro-inflammatory factors, such as interleukin-1, 6, and 8, can promote vascularization and accelerate wound healing by inhibiting endothelial cell apoptosis and activating the endothelial cells to form blood vessels.^11,12 However, in the experimental injury models, hyper-inflammation has been shown to delay wound healing and result in increased scarring. Furthermore, chronic inflammation, a hallmark of the non-healing wound, predisposes to a protracted course of wound healing, unhealed wound, or even pathological transformation. Therefore, inflammation at the damaged tissue sites should be well-staged and precisely orchestrated along with the wound healing cascade. It has been shown that inflammation is critical for attracting MSCs to wound sites and increasing evidence strongly indicates that MSCs can accelerate wound healing by modulating inflammation, improving the motility of various cell types such as fibroblasts and endothelial cells, and promoting angiogenesis and extracellular matrix deposition and remodeling.^2,13-15 Therefore, a meticulous understanding of the orchestration of MSCs and inflammation during the entire process of wound healing should provide key information for better treatment of wounds, as well as the acquired knowledge about tissue microenvironment during pathological processes such as fibrosis and tumor development.

MSCs Modulate the Inflammatory Responses During Wound Healing

Once an acute wound occurs, the exposure of collagen initiates the intrinsic and extrinsic clotting cascades. Acute inflammation follows immediately, causing an increase in blood vessel leaking and local swelling. It is an inevitable consequence of injury, aiming for controlling bleeding, preventing infection, and triggering the repair process. But excessive inflammation will cause chronic wounds even the failure of wound healing. Indeed, altered inflammation in the early stage has been deemed essential to affect subsequent steps of the repair process that could influence cell proliferation, proper wound healing, and remodeling. In a bone marrow transplantation mouse model, green fluorescent protein (GFP) labeled endogenous MSCs can be found preferentially in areas of cutaneous injury, which are probably activated by early signals of pathological and inflammatory changes to initiate a cascade of wound healing.⁹ Available evidence shows that MSCs-based therapy is likely to reduce inflammation but not replace damaged cells. For instance, in treating acute and chronic liver injury, the administration of MSCs is a strategy widely used in the early stage to regulate organ inflammation in different tissue sites. Even for SARS-CoV-2 infection, MSCs intravenous infusion was suggested as a treatment for COVID-19 pneumonia to suppress pulmonary inflammatory storm.¹⁶

Major Effector Molecules Released by MSCs

Numerous studies have shed light on the immunomodulatory properties of MSCs in treating inflammatory diseases and found several factors and molecules derived from MSCs linked to this function. These include vascular endothelial growth factor C (VEGFC), transforming growth factor-β1 (TGF-β1), IL-6, nitric oxide (NO), indoleamine 2, 3-dioxygenase (IDO), tumor necrosis factor stimulated gene-6 (TSG6), prostaglandin E2, IL-1 receptor antagonist, IL-10, hepatocyte growth factor (HGF), platelet-derived growth factor (PDGF), insulin growth factor (IGF), stromal cell-derived factor-1 (SDF-1), and an antagonistic variant of the chemokine CCL2.^13,17-19 In addition, MSCs can also secrete a large amount of MMPs, which could regulate the inflammation by regulating the activity of chemokines and mediate the phagocytic functions and immune responses of inflammatory cells through matricryptins.^20,21 At least in principle, some of these regulations could be exerted by p63, an epithelial regulator able to impinge on several interleukins.^22-24 The ability of MSCs to modulate the inflammatory response in wounds supports their favorable effect on the healing response. A severe burning rat model, which adopted the human umbilical cord-derived MSCs treatment, demonstrated a significantly decreased number of inflammatory cells and pro-inflammatory cytokines such as IL-1, IL-6, and TNF-α. Importantly, these wounds also exhibited a shorter recovery time and faster healing rate.²⁵ MSCs can also lower the level of the pro-inflammatory cytokine and accelerate wound healing in the cornea undergoing chemical injury.²⁶ TSG-6 is a crucial immunosuppressive factor conferring MSCs with prominent anti-inflammatory properties for treating myocardial infarction,²⁷ peritonitis_,²⁸ acute lung injury,²⁹ and corneal injury_.³⁰ The TSG-6 released from MSCs infused into wound margins contributed to the suppression of the release of TNF-α from activated macrophages, accelerating wound healing, and reducing tissue fibrosis in murine full-thickness skin wounds.³¹ Interestingly, TSG-6-modified MSCs effectively ameliorated pathological scarring, decreased inflammatory molecular secretion such as monocyte chemoattractant protein-1 (MCP-1), TNF-α, IL1β, and IL6, and attenuated collagen deposition in a mouse skin wound model. Notably, the pro-inflammatory cytokines were decreased.³²

Plasticity of MSCs in Immunomodulation

It is important to note that MSCs can be “licensed” to exert their immunomodulatory effects after stimulation with interferon-γ (IFN-γ) in the presence of one (or more) other cytokines (s), including TNF, IL-1α, or IL-1β. But this exclusive feature also depends upon the kinds and concentrations of inflammatory mediators present in the tissue microenvironment. Low concentrations of cytokines were sufficient to upregulate chemokine secretion but were not enough to induce substantial expression of iNOS or IDO and ultimately led to greater inflammation.³³ In addition, the types of cytokines present in the environment can further enhance or weaken the immunosuppressive effects of MSCs induced by TNF and IFN. For example, IL-17 present in the environment further strengthens the immunosuppressive ability of MSCs.³⁴ Reversely, TGF-β and IL-10 result in the MSCs less immunosuppressive.^35,36 Another factor that affects the immunoregulatory ability of MSCs is the recognition of microbial molecular patterns through Toll-like receptors (TLRs). For instance, TLR-4 activation induces a pro-inflammatory response, while TLR-3 activation results in an anti-inflammatory signal. Interestingly, TLRs have also been reported to play an important role in skin injury repair.^37,38 Therefore, specific stimuli are crucial in provoking the immunomodulatory capacities of MSCs during MSC-based therapy for wound healing.

A growing number of studies on the crosstalk between inflammation and MSC-mediated immunosuppression strongly support the notion that the immunoregulatory function of MSCs is highly plastic. An impressive study in the mouse model of liver fibrosis has demonstrated the therapeutic roles of MSC treatment for the initial and progressive stages of liver fibrotic injury, indicating distinct effects of variable inflammatory stages of diseases with various kinds of cytokines.³⁹ Also, concurrent administration of steroids, a widely used clinical immunosuppressor, abolished the therapeutic effects of MSCs on liver cirrhosis by reversing the immunosuppressive property of MSCs.⁴⁰ Likewise, a similar outcome found in a GvHD mouse model, which was attributed to the effects of VEGFC, a novel immunomodulatory factor derived from MSCs.⁴¹ Those findings suggest that the appropriate inflammatory state of the disease for intervention is one of the most critical conditions for MSC therapy to be effective in wound healing.

The Interaction Between MSCs and Various Immune Cells

Upon injury, several inflammatory cell types,⁴² which bridge innate and adaptive immunity, have been found to play a critical role in wound healing.^43-45 Especially during the inflammation phase, the infiltrated immune cells and inflammatory cytokines are in a dynamic change, which significantly affects the immunoregulation of MSCs. Correspondingly, several studies have demonstrated that MSCs also can exert immunomodulatory function by regulating the activation of various immune cells, including macrophages, neutrophils, natural killer cells, dendritic cells, T lymphocytes, and B lymphocytes^46,47 (Fig. 1).

Figure 1.

The immunoregulation of MSCs during wound healing. The inflammatory phase is one of the main steps for deciding on an ordinary or impaired wound healing course; this phase is necessary to clean bacteria, tissue debris, apoptotic cells, and clots from the wound. MSCs can exert immunomodulatory function by regulating various cells’ activation and operation of the innate and adaptive immune systems, including mast cells, neutrophils, macrophages, and T cells.

Neutrophils

Neutrophils, which are usually not observed in the normal skin, are produced in the bone marrow from promyelocytes and are recruited as “first responders” from blood in response to “find me” signals, including DAMPs, hydrogen peroxide, lipid mediators, and chemokines released from regions of injury or infection.⁴⁸ Neutrophils reach the injury site immediately after injury and peak after 24 h. These sites combat infectious threats by releasing toxic granules, producing an oxidative burst, initiating phagocytosis, and generating neutrophil extracellular traps (NETs).⁴² Increasing evidence suggests that the injected MSCs promote wound healing by regulating neutrophil activation and NETs formation, which is associated with protection from infection.^49,50

However, neutrophils are not just occurred in response to infection. It has been reported that they also penetrate the injury site and perform essential repair functions in a fully repairing sterile thermal hepatic injury.⁵¹ We also reported that the lung-resident MSCs promote neutrophil recruitment and the formation of NETs, which facilitate cancer cell metastasis to the lungs.⁵² The above studies indicate that neutrophils have alternative functions in addition to their classic bactericidal functions. Interestingly, apoptotic neutrophils significantly induce a repair macrophage phenotype that involves the induced production of TGF-β and IL-10.⁵³ Moreover, it is reported that MSCs-conditioned medium is responsible for tissue repair by inducing neutrophil apoptosis in the lipopolysaccharide (LPS)-induced acute lung injury (ALI) mouse model.⁵⁴ However, it does not always occur the same way. A growing number of studies have demonstrated excessive neutrophil infiltration during this phase may increase inflammation and impede wound healing. NETs could destroy wound structures, impair angiogenesis in the wound area, and also affect the number or functions of cells participating in wound repair, eventually leading to delayed wound healing.⁵⁵ In the ALI mouse model, MSCs or MSCs conditioned medium treatment significantly reduced the number of infiltrated neutrophils and alleviated the progression.^56,57 Moreover, we found that human adipose-derived MSCs (ADSCs) could effectively promote the clearance of neutrophils, reduce the NETs formation during the granulation stage, inhibit neovascularization, and reduce the opacification of the ethanol-injured cornea.⁵⁸ The importance of neutrophils in wound healing was clearly demonstrated in mice genetically deficient for CXCR2- a chemokine receptor crucial for neutrophils recruitment to wound sites. These mice exhibited delayed re-epithelialization of skin wounds compared to the WT mice.⁵⁹ However, in another study, the author found that the re-epithelialization was accelerated when the neutrophils were depleted after the mice were injected with anti-Gr1 antibody.⁶⁰

Macrophages

Macrophages, which arrive at the injured site followed by neutrophils, are essential to routine wound healing and tissue regeneration. The wound-accumulated macrophages originated from monocytes-derived macrophages or expanded tissue-resident macrophages.⁴² Significantly, in a murine wound model, depletion of macrophages shows delayed wound closure.^61,62 Conversely, increasing the number of monocytes or macrophages in the wound can significantly accelerate both normal and diabetic murine wound healing.⁶³ In the early stages of wound healing, macrophages are microbicidal and pro-inflammatory, expressing TNF-α and IL-6 and IL-1, referred to as M1. Macrophage depletion in this phase has demonstrated roles in the formation of vascularized granulation tissue, epithelialization, and diminishment of scar formation.^64,65 With the resolution of inflammation, the transition of macrophages from M1 to anti-inflammatory phenotype (M2) is critical for the subsequent proliferation stage and, eventually, tissue regeneration. Depletion of macrophage during the proliferation stage led to hemorrhage and a failure of wound closure.⁶⁴

The functionally converted macrophages further enhance the migration and proliferation abilities of fibroblasts to facilitate the wound-healing process.⁶⁶ Several early studies have demonstrated that apoptotic bodies or exosomes derived from MSCs could also promote cutaneous wound healing by triggering macrophages to polarize towards the M2 phenotype. Exosomes derived from MSCs are also reported to alter the M2 polarization and enhance wound healing.⁶⁷ The same phenomenon was observed in a full-thickness wound experiment. It was found that miRNAs Let-7b in exosomes from LPS-pretreated MSCs regulate macrophage M2 polarization via the TLR4/NF-κB/STAT3/AKT signaling cascade.⁶⁸ In addition, exosomes derived from melatonin-stimulated MSCs have been applied to promote diabetic wound healing by regulating macrophage M2 polarization and increasing the expression of IL-10 and Arg-1 via targeting the PTEN/AKT pathway.⁶⁹ Moreover, IL-6 and GM-CSF produced by MSCs are reported to significantly elicit the M2 polarization process of macrophages and contribute to a marked acceleration of wound healing.⁷⁰ To develop technologies that can be used for clinical treatment, CCR2-engineered MSCs were created to induce a regenerative immune microenvironment for tissue repair by modulating the inflammatory properties of macrophages.⁶⁰ Our studies have shown that hypoxia-conditioned MSCs allow macrophages to acquire an anti-inflammatory phenotype by producing IGF-2, thereby alleviating experimental autoimmune encephalomyelitis (EAE).⁷¹ Mechanistically, nuclear translocation of ligated IGF2R complexed with GSK3 can induce proton rechanneling and promote oxidative phosphorylation (OXPHOS) in maturing macrophages.⁷² These studies provide novel strategies for the clinical use of MSCs in wound healing by regulating macrophage polarization.

Mast Cells

The mast cell is a type of granulocyte derived from the myeloid stem cell, which is widely distributed around the microvessels under the skin.⁷³ These cells represent up to 8% of the total nucleated cells within the dermis and their cutaneous versions are localized adjacent to the epidermis and the subdermal vasculature and nerves. At the time of injury, mast cells become activated and secret a large number of pre-existed mediators in the granules, including vasoactive amines (eg, histamine, serotonin), lipid mediators (eg, LTB4, PGE2), cytokines (eg, IL-1β, IL4, IL6, TNF-α), and growth factors (eg, FGF-2, PDGF, VEGF). These mediators have been investigated during various stages of wound healing, including inflammation, proliferation, and remodeling, with a number of contradictory findings in the wound healing outcomes in mast cell-deficient mice using different skin injury mouse models (WBB6F1-Kit^W/Kit^W-v VS Mcpt5Cre/DTR mice).^74-76 It has been proved that mast cells are mechanoresponsive and able to modulate the recruitment of neutrophils into injury sites using mast cell-deficient mice.⁷⁶ In addition to mediating the early inflammatory response, the mast cells also directly contribute to skin fibrosis and scar formation-based wound contraction by activating fibroblast proliferation and collagen synthesis.^77,78 On the other hand, it was observed that these cells could stimulate keratinocyte proliferation and re-epithelialization to functionally facilitate wound contraction and skin regeneration by generating histamine,⁷⁹ and even inhibit fibrosis by suppressing the differentiation of endogenous mesenchymal stem cells to myofibroblasts.⁸⁰ A balance between 2 distinct patterns of repair engaged by mast cells is likely to determine the fate of wound repair. Accordingly, increasing studies investigated the potential of using MSCs to regulate mast cells for wound repair based on the biological properties of these cells. In a murine atopic dermatitis model, MSC administration significantly inhibited mast cell degranulation and alleviated inflammation by producing PGE2 and TGF-β1.⁸¹ Of note, numerous studies are focused on the effects of MSC exosome components on mast cells. MSC exosomes have been shown to inhibit the increase in the number of dermal mast cells in TLR7 agonist-treated mice.⁸² Likewise, the culture medium from TNF-α-stimulated MSCs could reduce mast cell activation and histamine release through a COX2-dependent mechanism and attenuate experimental allergic conjunctivitis.⁸³ Although regulatory effects have been identified in animal studies, further research is needed to focus on the key molecular mechanisms by which human MSCs regulate mast cell function for clinical precision therapy.

T cells

T cells are one of the important types of the immune system and play a central role in the adaptive immune response, drawn by chemokines produced in the wound, such as CCL9 and CCL10, are the final immune cells to infiltrate into the wound.⁸⁴ Since the immune system is intimately involved in wound healing, extensive work with innovative tools has examined the impact of T cells and their cytokines on the final wound outcome. T cell-mediated immune responses are dynamic in wound healing. They are recruited in smaller numbers at the late stages with a Th1 response in the acute inflammatory phase, which is primarily driven by interferon-gamma (IFN-γ). As a proinflammatory cytokine, IFN-γ is able to aid in the polarization of macrophages to the M1 phenotype. When the inflammatory phase progresses, it is shifted into a Th2 response as T cells are progressively polarized. These cells produce IL-4, IL-5, IL-10, and IL-13, which, although anti-inflammatory, also have significant profibrotic effects. Murine studies support this observation and demonstrate that the interleukins, IL-4, IL-5, and IL-13, secreted from the Th2 cells resulted in scar formation.^85,86 Consistently, impaired wound healing, characterized by excessive inflammation, is associated with increases in Th2 cell-derived TNF-α level. Overlapping with the Th1 and Th2 immune responses, the immune responses driven by Th17 cells result in re-epithelialization of the wound as well as hair follicle neogenesis through neutrophil recruitment. In addition, regulatory T cells (Treg cells) were found to promote tissue homeostasis and regeneration following tissue damage via restraining excessive inflammation. Strikingly, MSCs have been acknowledged for their ability to regulate T-cell immune responses and are therefore widely used for the treatment of inflammatory diseases, including refractory wounds. MSCs broadly suppress T-cell activation and proliferation in vitro via a plethora of soluble and cell contact-dependent mediators. These mediators may act directly upon CD4⁺T cells and CD8+T cells or indirectly via modulation of antigen-presenting cells and other accessory cells. The multiple effects of MSCs on cellular immunity may reflect their diverse influences on the different T-cell effector subpopulations and their capacity to specifically protect or induce Treg populations. For instance, the CCR2 over-expressed MSCs were reported could promote the accumulation of Treg cells in diabetic wounds and accelerate wound closure.⁸⁷ MSCs also reduce collagen deposition¹³ and attenuate scar formation.^32,88 It’s demonstrated that MSCs usually perform their therapeutic function by balancing pro-inflammatory and anti-inflammatory responses and directly or indirectly regulating disease-associated T-cell subsets (Th1, Th2, Th17, Treg).⁸⁹ Although the inflammatory pathological process of wounds and the immunomodulatory properties of MSCs are clearly characterized, the specific mechanisms of interaction and regulatory targets between MSCs and T-cell subsets remain to be investigated for the clinical use of MSCs in the treatment of wounds in order to optimize the period and route of cell delivery.

In skin, one specific type of T cell that is important in skin wound healing is γδ T cells. They are known to recruit immune cells, stimulate angiogenesis, and activate keratinocytes.⁹⁰ In addition, skin TRMs (tissue-resident memory T cells) are another unique subset of T cells that reside in the skin and provide long-lasting immune protection against re-infection by a pathogen. They are critical for local immunity in the skin and have been shown to play a key role in wound healing by modulating the immune response and promoting tissue repair. However, few articles focus on the impact of MSCs-based therapy on them in wound healing treatment.

Cancer and Wound Healing

Cancer, regarded as “non-healing wounds,”⁹¹ is believed to take advantage of the pro-regenerative ability of host cells to facilitate local cancer growth, resistance to therapy, and metastases to remote organs.^92-94 It shares the same process with routine wound healing, including inflammation, tissue proliferation, and remodeling. However, the wound healing response of tumorigenesis possesses one or multiple prolonged, uncompleted phases, and tumors can therefore be considered as an “over-healing wound.”⁹⁵ MSCs have recently been studied for their role in the tumor microenvironment. First, the tumor-associated MSCs (TA-MSCs) are involved in the pathological inflammatory process of tumors. It has been found that TA-MSCs are derived from normal resident MSCs altered by the tumor microenvironment. These cells are able to recruit monocytes and neutrophils to the tumor by secreting chemokines and inhibit the function of T cells and NK cells, thus maintaining tumor growth.^96-99 Secondly, increasing evidence demonstrated that MSCs could promote an immunosuppressive environment that ultimately favors tumor progression and metastasis by inducing differentiation of leukocytes into immunosuppressive myeloid-derived suppressor cells (MDSCs)¹⁰⁰ and increasing the proportion of Treg cells.¹⁰¹ Of note, we recently reported that the C3 secreted by lung resident MSCs could lead to pulmonarymetastasis of tumors by inducing NETs formation,⁵² which has been proved to act as a “trap” for circulatingtumorcells in previous studies.¹⁰² These results shed light on the mechanisms involved in the promotion of tumor progression by MSCs from an immunological perspective.

The Application of MSC Therapy on Different Pathological Wounds

Skin covers the surface of the human body, serving as a protective barrier against pathogens, this largest organ of the human body plays a vital role in maintaining the homeostasis of the body. In addition to trauma and burns, vascular insufficiency, diabetes mellitus, local pressure effects and systemic factors such as poor nutritional condition, infection as well as altered immunological status can lead to refractory wounds. Many studies have been performed on the therapeutic potential of MSCs for different chronic refractory wounds, including burn wounds, diabetic ulcers, and pressure sores (Table 1).

Table 1.

Outcome assessment parameters from clinical trials.

Study	Wound healing	Patients	Cell type	Cell number	Cell source	Cell administration	Outcome
Rasulov103	Thermal burn	Female, 45 years, 1	BM-MSC	20-30 × 103cells/cm2	Allogenic	BM-MSC suspension was applied to burn wounds and the first skin graft transplantation was carried out in 4 days. Additional transplantations of MSCs onto wounds and immediately made.	Proved safety, improved new vessel, and granulation formation and reduced coarse cicatrices.
Eduardo Mansilla104	Thermal burn	Male, 26 years, 1	BM-MSC	106/cm2	Allogenic	Spraying the MSCs onto the burn wound surface after deep escharotomy using 2 sterile conical tubes. Then the surface was dressed with a sterile polymeric transparent film and keep changing the film.	Good blood supply without infection, better granulation tissue formation, and better re-epithelialization.
Xu105	Thermal burn	Male, 19 years, 1	BM-MSC	1 × 104 cells/cm2	Autologous	The autologous split-skin graft was spread over the dermal matrix sheet to cover the wound.	No pain, infection or other side effects during wound healing and reduced skin contraction after 2 years.
Nikolaos Arkoulis106	Thermal burn	Female, 42, 55 years, 2	Adipose stem MSCs	46 400 cells/cm2	Autologous	Surface of burn + skin-graft	Positive
Wael Abo-Elkheir107	Thermal burn	All, 15 to 50 years, 60	BM-MSC/UC-MSC	Not mentioned	Allogenic	Two days after the surgical excision of deeply burned tissue, stem cells were injected into the burned area and the application was repeated in 10 days.	Improved rate of healing and reduced hospitalization period in both BM-MSC and UC-MSC groups.
Vincent Falanga108	Lower extremity wounds	7	BM-MSC	2 × 106/cm2	Autologous	Delivered topically in a fibrin spray, using the same methodology with respect to fibrin (no more than 2 mL).	Positive
Nihar Ranjan Dash109	Lower extremity wounds	6	BM-MSC	>1 × 106/cm2	Allogenic	Along the edges of nonhealing ulcers	Improved recovery of the ulcer area after transplantation.
Lafosse Aurore110	Lower extremity wounds	Two male, 46 and 21 years, 1 female, 41 years	Adipose stem MSCs	5.8 × 105 cells/cm2	Allogenic	Covered with human acellular collagen matrix surface	Positive
Qin111	Lower extremity wounds	Not mentioned	UC-MSC	Between 5 and 10 × 106 cells/cm2	Allogenic	Multiple points	Positive
Xiangxia Zeng112	Lower extremity wounds	Female, 57 years, 1	Placenta-derived Mesenchymal Stem Cells	1  ×  106 cells/cm2	Allogenic	On topical MSCs hydrogel	After 3 weeks, the wound had completely healed.
JJ Lataillade113	Radiation burn	Male, 27 years, 3	BM-MSC	200 × 103 cells/cm2	Autologous	Excision plus skin autograft and a local MSC therapy	the healing was complete without any functional impairment.
Marc Benderitter114	Radiation burn	3	BM-MSC	Totally 76 × 106 cells	Autologous	Surface of burn	Perfect healing persist 3 years after the implantation.
Portas115	Radiation Burn	Male, 66 years, 1	BM-MSC	Not mentioned	Allogenic	Surface of burn	Ulcer dimensions were reduced with local recovery and remission of signs and symptoms.
Damián García-Olmo116	Perianafistula	5	Adipose Mesenchymal Stem Cells	10-15 × 103 cells/cm2	Allogenic	Injected into the tract walls	Positive
Garcia-Olmo117	Perianal fistula	35	Adipose Mesenchymal Stem Cells	103 cells/cm2	Allogenic	Injected into the tract walls	Quality of life scores were higher in patients who received ASCs than in those who received fibrin glue alone.
Takafumi Yoshikawa118	Burn, lower extremity wounds, pressure ulcers	All, average 64.8 years, 2	BM-MSC	Not mentioned	Allogenic	Artificial dermis was cut to match the shape of skin wounds and the cultured marrow mesenchymal cells were combined in the artificial dermis.	Confirmed skin regeneration and epithelialization in 2 to 4 weeks. One year after graft, the scarring degree was milder.
González Sarasúa119	Pressure ulcers	19 male, 3 female	BM-MSC	60 million cells	Autologous	Injected into the ulcers	Positive

Radiation Burns

Severe radiation burns continue to be a significant challenge for wound care therapies, as they are often insensitive or resistant to conventional treatments, which are very lengthy and difficult.¹²⁰ Several studies have explored the therapeutic potential of MSCs for radiation burns. Alain Chapel et al investigated the potential of a combined infusion of autologous ex vivo expanded hematopoietic cells with MSCs for the treatment of multi-organ failure syndrome following irradiation in a non-human primate model in 2003. And then they reported remarkable outcome of a clinical trial in which bone marrow-derived MSCs were first used to treat radiation-induced proctitis in patients over-irradiated at Epinal Hospital. It was demonstrated that MSCs reduced pain and the number of episodes of diarrhea in three patients who were resistant to symptomatic treatments.^121,122 In another trial, a decline of CD4⁺CD127⁺CD25⁺ T lymphocytes and an increase of CD4⁺CD25⁺ (potentially regulatory) T lymphocytes were observed in patients after receiving MSC treatments.¹²² Besides autologous BM-MSCs, allogenic BM-MSCs have also been used to treat a radiation burn, showing significant improvement in vasculature and skin quality with inflammatory remission. The activation of T and B cells was suppressed along with the decreased levels of C-reactive protein (CRP)¹¹⁵ after MSCs administration. These promising clinical findings also provide a wealth of prospective information for the use of MSCs in the treatment of severe radiation burns.

Lower Extremity Wounds

Lower extremity wounds are a prevalent and reoccurring type of complicated wound, becoming a big public healthcare issue with critical social and economic concern. They can be caused by one or a combination of problems including poor circulation, critical limb ischemia, diabetes, and other medical problems. Currently, nearly 1% of adults and 3.6% of older patients are reported to suffer from this chronic condition.¹²³ Several investigators have reported their studies regarding the wound-healing potential therapeutic effects of MSCs on these refractory wounds.

Lafosse et al reported that implantation of adipose-derived MSCs seeded onto human acellular collagen matrix (biological dressing) represents a promising therapy for nonhealing wounds, offering improvement in dermal angiogenesis and remodeling. Significant macrophagic recruitment was also found in injured tissues after treatment without modification of lymphocyte infiltration.¹¹⁰

Another clinical study also showed a tremendous therapeutic effect of hUC-MSC on diabetic foot, and it demonstrated that the ratios of Treg/Th17 and Treg/Th1 cells were significantly increased after hUC-MSC transplantation. In contrast, the ratio of Th17/Th1 cells remained unchanged. CRP and TNF-α levels were also considerably reduced after hUC-MSC treatment.¹²⁴ It indicated that hUC-MSC exerted an anti-inflammatory effect on diabetic foot ulcers, offering great potential for clinical application as an alternative to amputation.

Pressure Sores

Pressure sores are a significant cause of morbidity resulting from unrelieved pressure against the skin. Although many preventative and treatment modalities have been developed in recent years, the treatment of pressure ulcers remains frustrating and time-consuming. Considering that MSCs can promote skin wound healing, several clinical studies have been performed to determine the therapeutic potential of MSCs for pressure sores.

In 2008, Yoshikawa et al treated 11 patients suffering from pressure sores that had persisted for more than 3 months with autologous BM-MSCs impregnated on a collagen sponge. In 9 of the 11 patients, the ulcers almost healed; in the remaining 2 patients, the ulcers became smaller after treatment. Long-term follow-up of those surviving patients was favorable. Except for 2 patients with worsened nutritional states, the decubitus ulcers in the other 7 patients did not recur for at least 1-year post-treatment.¹¹⁸ We hypothesize that transplantation of BM-MSCs at a wounded site could reconstitute and modulate the dermal fibroblast population to facilitate the local cutaneous angiogenic formation, accommodate collagen deposition, and manipulate inflammation through anti-inflammatory effects or by suppressing the immune response.

Conclusion and Prospects

Though increasing investigations have shown the therapeutic effects of MSCs on wound healing, the underlying mechanisms remain largely undefined. The ability of MSCs to modulate the inflammatory response in wounds supports their favorable impact on the healing response. However, the inflammation also alters the MSC’s secretome, including a source of growth factors that promote the proliferation and differentiation of tissue cells. Therefore, a more in-depth understanding of the complicated link between MSCs and inflammatory conditions should be investigated in the future, which is essential for translation into clinical application.

In addition, many challenges need to be addressed before MSCs fulfill their therapeutic potential for clinical treatment. Allogeneic MSCs have the advantage of enabling complete cell characterization prior to treatment and increasing the possibility of rapidly applying cells to patients with low immunogenicity. However, some studies have described the generation of antibodies against and immune rejection of allogeneic donor MSCs.¹²⁵ In addition, the MSCs immunoregulation capability is affected by the status of the disease and the age of donors.^126,127 Moreover, the dosage of MSCs application is another issue that needs to be concerned, as chronic inflammation delay wound healing, however, a moderate inflammatory response is beneficial for wound repair. Therefore, the application of MSCs should control inflammation at an appropriate level to aid the wound healing transition from the prolonged inflammation phase to the proliferative phase. In order to maximize the effect of MSCs application in clinical treatment, all of these factors should be considered. In addition, the delivery route, and administration period should also be taken into consideration.

Moreover, one of the most significant strategies for optimizing MSCs-based therapy is to license MSCs and make them produce specific factors for treating various diseases. It has been reported that human umbilical cord-derived MSCs treated with TNF-α and IFN-γ could secret a large amount of VEGFC, which promotes angiogenesis, thus accelerating skin wound healing. Therefore, empowerment with inflammatory cytokines may have tremendous therapeutic potential in terms of enhancing the ability of MSCs to achieve an appropriate therapeutic effect in wound healing.

Funding

This study was supported by grants from the National Key R&D Program of China (2022YFA0807300), National Natural Science Foundation of China (81930085), Jiangsu Province International Joint Laboratory for Regenerative Medicine Fund, Suzhou Foreign Academician Workstation Fund (SWY202202). Wuxi Taihu Tanlent Top Talent Team, Jiangsu Province, China (2021).

Conflict of Interest

The authors declared no potential conflicts of interest.

Author Contributions

M.Z.: conception and design, writing—original draft preparation. L.C.: conception and design, writing—original draft preparation. S.M., E.C., Y.W., C.S.: conception and design. G.M.: conception and design, writing—review and editing. Y.S., X.C.: conception, design, supervision of study conduct and operations, writing—review and editing, final approval of manuscript.

Data Availability

The data that support the findings of this study are available from the corresponding author upon reasonable request.