The global number of individuals suffering from organ dysfunction as a result of acute injuries, chronic disorders, or aging has been on the rise and thus inadvertently places a high demand for organ transplantation. However, organ and tissue transplantation is obviously limited by the shortage of donors and side effects associated with the use of immunosuppressants (1), placing stress upon current methodology and creating a need for an alternative therapeutic avenue. By virtue of its self-renewal properties and capability in differentiating into multiple cell types, recent advances in human pluripotent stem cell research has offered a literally unlimited amount and varieties of therapeutic cells for transplantation (2, 3). Nevertheless, there is a lack of clinical evidence showing their long-term engraftment following transplantation possibly due to poor cell survival and chronic immune rejection (4, 5). Moreover, regenerative therapies stimulating endogenous regeneration such as growth factor-based strategies have shown mixed results in the clinic due to safety concerns and cost-effectiveness (6, 7). Therefore, it is necessary to find new ways to improve regenerative strategies and one of them is to control and utilize the host immune system. Nevertheless, in order to design immune-centric regenerative therapies, it is imperative to understand how the various immune components modulate tissue repair and regeneration.

Since decades, the immune system is well known to be implied in tissue repair and regeneration. For instance, inflammation following injury greatly contributes to tissue repair and scar formation, while excessive inflammation led by immune cells causes pathological fibrosis that debilitates tissue function and may lead to organ failure. Immune-mediated tissue healing processes are complex, yet, highly orchestrated. After injury, invading pathogens, necrotic debris, the clotting reaction, and tissue-resident immune cells trigger an inflammatory response, which result in the recruitment of various immune cells. The activity of immune cells during wound healing can be separated in three phases (8): First, pro-inflammatory cells are recruited to the site of injury for host defense and phagocytosis of necrotic tissues. Second, the pro-inflammatory response is dampened via immune cells such as anti-inflammatory macrophages, while immune cells also directly participate in stimulating angiogenesis, myofibroblast activation, and tissue progenitor cell proliferation. Last, most immune cells exit the site of injury or are eliminated by apoptosis and the tissue homeostasis is restored. Nonetheless, the role of the various immune cells and their subsets as well as the mechanisms by which they regulate tissue healing remain largely elusive. It is, therefore, imperative to understand how tissue healing is controlled by the immune system and harnessing the endogenous regenerative capacity has recently become an active area of research.

An interesting observation supporting the critical role of immunity in regeneration (as opposed to tissue repair and scarring) comes from the evolution of the immune system among species and during development. Compared to lower vertebrates such as amphibians and teleost fishes that are capable of completely regenerating many body parts, mammals have a limited regenerative potential. To explain this difference, it has been postulated that the loss of regenerative capacity in mammals is in part associated with maturation of their immune system compared to lower vertebrates (9, 10). The immune system also changes during development and throughout life. For example, some organs such as the mammalian heart is notorious for not being able to regenerate and the necrotic cardiac muscles are replaced by dysfunctional scar tissues after injury. However, accumulating evidence shows that the neonatal hearts of mammals including humans have a transient regenerative capacity compared to adults (11–13). Indeed, the mammalian adaptive immune system is relatively immature after birth, which coincides with the period of neonatal regeneration. In contrast to adults, neonates do not mount a robust fibrotic but a more angiogenic response that facilitates tissue regeneration after injury (10). Therefore, since immune cells regulate both fibrosis and angiogenesis during tissue healing, targeting the immune system to promote neoangiogenesis with minimal fibrosis would be an interesting approach to stimulate regeneration. Therefore, understanding how immunity regulates tissue fibrosis and neoangiogenesis would shed light on the development of potential therapeutics targeting endogenous tissue regeneration. During the last decade, innate immunity, in particular, macrophages and their various polarization states, has been considered as a central regulator of the tissue healing process. However, recent evidences suggest that the adaptive immune system is also a critical actor. In this review, we focus on the role of regulatory T-cells (Treg).

Treg are required for maintenance of self-tolerance, preventing excessive inflammation and autoimmune diseases. The most reliable cell-specific marker of Treg is Forkhead box P3 (FOXP3), which is essential for the maturation and function of Treg. Congenital deficiency in Treg, due to mutation of the Foxp3 gene, causes fatal autoimmunity in mice, the scurfy phenotype, and enteropathy, X-linked (IPEX) syndrome in human (14, 15). Treg are normally present in lymphoid organs but have been shown to accumulate in damaged tissues to some extent. Long recognized as potent suppressors of the immune system, Treg have been recently rediscovered as indirect and direct regulators of tissue healing, while the mechanisms are still largely unknown (16–18).

Uncontrolled inflammation after tissue injury can lead to impaired healing and tissue remodeling. In many tissues, Treg are recruited to the damaged site to facilitate inflammation resolution and to regulate immunity after injury (19). For instance, Treg can indirectly modulate regeneration by controlling neutrophils (20–22), inducing macrophage polarization (23, 24), and regulating helper T-cells (22, 25) (Figure 1) Moreover, Treg have been shown to directly facilitate regeneration via activating progenitor cells locally (16, 17).

Treg are able to control the functions of neutrophils and macrophages, which have been widely shown to be involved in the tissue healing process. Neutrophils are among the first leukocytes recruited to the injury site, and they directly modulate tissue healing either positively or negatively. For instance, after skeletal muscle injury, it has been demonstrated that neutrophils impair restoration of muscle structures and function through the release of hypochlorous acid, NAPDH oxidase, and other cytokines (26, 27). A negative role of neutrophils has also been demonstrated in a lung ischemia-reperfusion model, where neutrophils enhance the injury (28). However, in an inflammatory lung disease model, mice treated with intratracheal LPS, which induces neutrophil transmigration show activated β-catenin signaling in lung epithelial cells, triggering repair of the lung epithelium (29). Therefore, it is likely that neutrophils modulate tissue healing in a context-specific manner.

Treg have shown ability to modulate tissue healing via controlling neutrophil behavior. For instance, an in vitro study has demonstrated that activated Treg promote neutrophils to secrete anti-inflammatory molecules including IL-10 and TGF-β, heme oxygenase-1, and indoleamine 2,3-dioxygenase (IDO). This is also preceded by inhibition of neutrophil’s IL-6 production, suggesting that Treg can modulate inflammation through inhibition of neutrophil activity (30). Concomitantly, Treg have been shown to induce neutrophil apoptosis and death both in vitro and in vivo (21, 31). For example, in an acute lung injury model, Treg mediate resolution of lung injury via TGF-β-induced neutrophil apoptosis (21). In addition, Treg can modulate neutrophil infiltration to the site of injury. For instance, deletion of Treg leads to increased infiltration of neutrophils after cardiac injury and subsequently results in impaired healing (20, 22). In a model of kidney ischemia reperfusion injury, Treg suppress infiltration of neutrophils and attenuate kidney injury via IL-10 secretion (32). These studies indicate that Treg-mediated modulation of neutrophil behavior and activity is an important step toward regulating tissue healing.

Asides from neutrophils, Treg interact with other key innate immune cells involved in the inflammatory response such as macrophages. In addition to being scavengers that phagocytose cellular debris including apoptotic neutrophils and other cells, macrophages have been shown to play a pivotal role in tissue repair and regeneration. As a remarkable example, salamanders are well-known to be able to regenerate limbs, but depletion of macrophages leads to failure of limb blastemal formation and regeneration (33). Similarly, genetic ablation of macrophages during blastemal proliferation leads to failure of tail fin regeneration in adult zebrafish (34). In mice, depletion of macrophages leads to excessive fibrosis and lack of neoangiogenesis, resulting in failure in neonatal heart regeneration after myocardial infarction (MI) or apex resection (11, 24). Likewise, macrophages are important for cardio protection driven by cardiosphere-derived cells (CDCs), a stem-like population derived from cardiac biopsies ex vivo. Indeed, systemic depletion of macrophages with clodronate abolishes CDC-mediated cardioprotection and inhibits their regenerative capability in adult hearts after MI (35).

Importantly, during tissue healing, there are at least two different subsets of monocyte-derived macrophages, namely M1 and M2 macrophages. M1 are pro-inflammatory macrophages usually induced by IFN-γ or TNF-α, while M2 are anti-inflammatory usually induced by IL-4/IL-13 or IL-10. In this context, Treg are important regulator of macrophage phenotypes and functions (36–38). For example, monocytes cocultured with Treg produce decreased level of TNF-α and IL-6 in response to LPS; and the inhibition is associated with secretion of IL-10, IL-4, and IL-13 by Treg (39). Additionally, coculture of monocytes with Treg induces macrophages to polarize toward a M2 phenotype characterized by the upregulation of CD206, CD163, and decreased expression of HLA-DR (40). Treg can attenuate tissue injury and help tissue repair also by modulating macrophage activity and survival. For example, in a chronic kidney disease model, Treg protect kidney injury through inhibition of macrophage activity, which is dependent on Treg-derived TGF-β (41). In this context, Treg also inhibit monocyte survival through the Fas/FasL pathway (41).

Mounting evidence suggest that conventional T-cells are most likely detrimental for tissue healing (42). For example, CD4- and CD8-deficient mice have improved renal function in renal ischemia reperfusion model. SCID mice, which lack lymphocytes have significantly decreased intestinal leakage of albumin compared to wild-type mice after mesenteric artery ischemia and reperfusion (43, 44). In a MI model, CD8⁺ cytotoxic T-cells can respond to cardiomyocytes after being exposed to autoantigen in vivo and kill healthy cardiomyocytes in vitro (45, 46). Moreover, Rag^−/− mice, which lack T-cells have significantly smaller infarct size compared to control mice (47). In the context of bone, conventional T-cells may inhibit regeneration by promoting osteoclast differentiation (48). In addition, recruitment of CD8⁺ effector T-cells is correlated with delayed fracture healing and osteogenesis, due to secretion of IFN-γ and TNF-α (49). Deletion of CD8⁺ T-cells in mouse osteotomy model enhances fracture healing while adoptive transfer of CD8⁺ T-cells results in impaired healing (49).

The negative role of conventional T-cells in tissue injury is most likely mediated by the expression of inflammatory cytokines such as TNF-α and IFN-γ (50–52), but Treg can suppress conventional T-cells through various mechanisms including secretion of anti-inflammatory cytokines such as IL-10, TGF-β, and IL-35 (53–56). For example, it has been shown that deletion of Treg increase CD4⁺ and CD8⁺ cell number in the heart injury zone. In this context, both CD8⁺ and CD4⁺ T-cells show an increased secretion of IFN-γ and TNF-α, suggesting that Treg not only decrease the infiltration of conventional T-cells, but also attenuate their activity (22). Yet, the mechanisms by which Treg modulate tissue repair and regeneration are most likely tissue-dependent. In the next sections, we will discuss the role of Treg in the repair and regeneration of various tissues including skeletal and heart muscle, skin, lung, bone, and the central nervous system (CNS) (Figure 2).

Recently, exploring the function of the immune system during tissue repair and regeneration has gained a lot of interest in regenerative medicine. Nevertheless, we still have sparse knowledge on how immunity—in particular the adaptive immune system—controls the tissue healing process. For instances, what are the neo antigens, if any, released to initiate adaptive immunity-mediated tissue healing? Similarly, is adaptive immunity-mediated tissue healing an antigen-specific process? If so, how are T-cells recruited, activated and function in response to self-antigens during injury that are different from responding to non-self antigens? Currently, the development of treatments targeting the immune system is hindered by the lack of markers that specifically define distinct subsets of immune cells. Recent advances in single-cell genomics could offer unprecedented delineation of lineage-specific markers and function of various subsets of immune cells operating during tissue repair and regeneration.

Furthermore, it is still unclear why scars are absent in some tissues such as in bone, but are forming in others such as in heart. Understanding how both innate and adaptive immune cells interact with tissue-resident progenitor cells and myofibroblasts would shed light on developing therapeutic strategies for improving healing and regeneration in the clinic. Moreover, accumulating evidence has shown that the function of the immune system declines with age (101, 102). Given that the immune system plays a crucial role in tissue repair and regeneration, whether the reduced tissue repair capacity is related to a degenerated immune system during aging awaits further investigations.

Overall, studies investigating the role of Treg during tissue regeneration have been largely based on the use of Foxp3^DTR mouse model (17, 18, 22, 25, 57, 77, 90). However, one caveat of using such model is that the mice develop spontaneous systemic autoimmunity when Treg are depleted for long term (103). Careful data analysis should also include gain-of-function experiments such as adoptive transfer of purified Treg into Rag1^−/− mice to determine the role of Treg in tissue repair and regeneration.

From a regenerative point of view, one could control tissue Treg to promote regeneration. Treg of adipose tissues, skeletal muscle, and colonic lamina propria are the best characterized tissue Treg that maintain organismal homeostasis (104). Similar to regenerative Treg as aforementioned, IL-33 has been reported to expand tissue Treg in colonic lamina propria (105) that are well equipped to participate in local repair responses with expression of the tissue repair factor, amphiregulin. The exact role of these tissue Treg in intestinal regeneration awaits further investigations. Nevertheless, manipulating Treg for alleviating inflammatory diseases has been tested in several clinical trials. For instances, the use of low dosage of IL-2 in selective expansion of Treg in human patients (106) as well as ex vivo expansion and adoptive transfer of Treg to treat Type 1 diabetes have been reported and tested in the clinic (107–109). Further studies are needed to investigate if these Treg-mediated strategies can also be utilized for inducing tissue regeneration.

Although it has been reported that superagonistic anti-CD28 mAb increases Treg infiltration or activities in mice, the use of humanized superagonisticanti-CD28 antibody TGF1412 caused cytokine storm, leading to organ failure in a previous trial (110). Furthermore, even though IL-33 plays an important role in recruitment and function of Treg in mice (60), IL-33 is dispensable in humans as individuals lacking IL-33 have no obvious health problems such as autoimmunity, indicating that the pro-regenerative function of IL-33 on Treg could also be different between mice and humans (111). Therefore, novel strategies in empowering Treg-mediated tissue regeneration for potential clinical uses would be needed in the future.

JL and KL wrote the manuscript; JT and MM made the figures; MM and KL revised the manuscript.

The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest. The reviewer AA and handling Editor declared their shared affiliation.