Aging is accompanied by a decline in physiological integrity and a loss of regenerative capacity in many tissues. The development of interventions that prevent or reverse age‐related disease requires a better understanding of the interplay of cell intrinsic, inter‐cellular communication and systemic deregulations that underlie the aging process. Immune dysfunction and changes in inflammatory pathways are transversal contributors to the aging process and are essential propagators of tissue deterioration. Here, we propose and discuss the rejuvenation potential of interventions that target chronic inflammation and how modulation of tissue repair capacity could be an important mediator of such anti‐aging strategies. We highlight how current knowledge on the systemic nature of inflammatory dysregulation in old organisms, together with the development of new animal models that allow for the isolation of the inflammatory component of aging, could provide new targets for interventions in aging based on the modulation of inflammatory pathways.
- chemokine (C‐X3‐C motif) receptor, also known as Fractalkine receptor
- gonadotropin‐releasing hormone
- Interleukin 10
- Janus kinases/signal transducer and activator of transcription proteins
- loss of function
- MANF heterozygous
- mesencephalic astrocyte‐derived neurotrophic factor
- nuclear factor kappa‐light‐chain‐enhancer of activated B cells
- Senescence Associated Secretory Phenotype
Aging is characterized by a decline in the organism’s physiological integrity, leading to the loss of tissue function and vulnerability to disease. Although the consequences of aging on human health are broadly apparent, the causes and drivers of the aging process are just beginning to be understood . The age‐related decline in tissue homeostasis results from the interplay of cell intrinsic, inter‐cellular communication and systemic deregulations that have to be considered in the development of an integrated view of the basic mechanisms of aging . Among these changes, alterations in the nature of the inflammatory response in old organisms have been postulated to be an important contributor to tissue dysfunction (Fig. Fig. 1 ), but experimental evidence providing mechanistic support to this idea is still limited.
Inflammation is a concept used to define a broad group of biological processes with a wide range in duration, and cellular and molecular composition. In the context of this discussion, we refer to inflammation as a state of response to tissue insult (either external, such as infection, or internal, such as tissue injury) that involves persistent elevated levels of cytokines capable of activating both innate and adaptive immune system. Thus, inflammation is an integral part of many physiological functions and, as a consequence, the inability to regulate an inflammatory response has multiple detrimental consequences for organismal homeostasis. Interventions that target inflammatory pathways and restore a regulated inflammatory response are promising strategies to prevent disease progression (Fig. Fig. 1).
The aging process is accompanied by a state of low grade nonresolving activation of inflammatory pathways, a phenomenon known as ‘inflammaging’ . The precise sources of age‐related chronic inflammation and the extent of the causal relation between inflammaging and tissue dysfunction in aging is still largely unknown. However, it is clear that inflammation is a common denominator between several age‐related pathologies and, independently of its role as the initial trigger driving the aging process, contributes to the progression of tissue dysfunction. A detailed discussion of the potential sources of age‐related chronic inflammation, as well as a comprehensive description of the studies that advance possible mechanistic links between age‐related inflammation and disease has been extensively reviewed elsewhere [3–5] and will not be discussed in detail. Here, we provide a view point on how inflammatory pathways can be used as targets for immune modulatory rejuvenating interventions to ameliorate age‐related tissue dysfunction and, thus, to prevent or reverse age‐related disease (Fig. Fig. 1). Further, we highlight how optimization of the tissue repair capacity is an expected outcome of such immune modulatory interventions and a potential effector of the anti‐aging effects (Fig. Fig. 1). We will also propose new approaches and animal models that allow testing the application of rejuvenating interventions based on immune modulation, while contributing to a better understanding of the links between chronic inflammation and age‐related tissue dysfunction.
Chronic inflammation and anti‐aging interventions
Although yet poorly defined, the chronic inflammation that accompanies the aging process differs in intensity, duration and possibly in the composition of the cytokines involved from the acute inflammatory response originating from infection or tissue injury. While effective inflammation is fundamental for tissue repair and clearance of pathogens, the nonresolved persistent inflammatory state in aging can be understood as a generally maladaptive response, occurring at a stage of life not subjected to selection during evolution.
A major stimulus continuously activating inflammatory signaling in aging is the presence of damaged organelles and macromolecules that accumulate in old organisms . The self‐debris and self‐molecules that result from unhealthy or dead cells are produced at a higher rate in aged tissues, while the mechanisms responsible for the disposal of harmful products of cellular damage progressively decline. Many well described strategies to delay aging and improve health and lifespan, including caloric restriction, are interventions that can potentially reduce inflammation by activating mechanisms of cellular damage disposal [5, 6]. Another important source of inflammation in aging is the accumulation of senescent cells and their associated pro‐inflammatory secretome [7, 8]. Also in this case, the removal of senescent cells, achieved through the use of senolytics, or in mouse models where the selective elimination of senescent cells can be chemically induced, displays a reversal or improvement of key aspects of aging phenotypes [9–11]. Finally, recent studies point to changes in gut microbiota composition with aging as a source of inflammatory activation [12, 13]. Mice maintained under germ‐free conditions live longer than their conventional counterparts with reduced levels of circulatory pro‐inflammatory cytokines. Co‐housing of these germ‐free mice with old, but not young, conventionally raised mice resulted in an increase of inflammatory markers in circulation . Recent work in short‐lived African turquoise killifish (Nothobranchius furzeri) indicate that microbiome transplantation from young to middle‐aged killifish can increase lifespan, pointing to microbiome composition as an aspect of health that can be modulated to impact the aging process .
Aging research has also relied in the analysis animal models of progeria, where accelerated aging is observed due to specific genetic alterations that phenocopy accelerated aging, activating several of the mechanisms described above [16, 17]. Although, in these models, the initial trigger that drives tissue dysfunction is not related to inflammatory pathways, it has been extensively documented that chronic activation of inflammatory signaling accompanies the premature development of age‐related pathologies. Importantly, interventions that reduce inflammation delay the onset of age‐associated features in progeroid mouse models, extending their longevity [18–20]. From these observations, it becomes apparent that targeting inflammation could be envisioned as an intervention to allay age‐related tissue dysfunction also during physiological aging, an idea that is already beginning to be tested [21–24].
The above described interventions are examples of strategies that can alter the rate of aging and can potentially act through pathways that are linked with the sources of increased pro‐inflammatory signaling in old organisms. However, age‐related immune alterations can be better understood as manifestations of an unbalance between pro‐inflammatory and anti‐inflammatory responses. This concept is critical to explain what is commonly referred to as the ‘paradox’ of centenarians: individuals with remarkable longevity present increased levels of circulating pro‐inflammatory cytokines. This apparent contradiction can be resolved if we consider that the same group of individuals display higher levels of anti‐inflammatory factors in circulation . These findings point to the importance of mechanisms involved in anti‐inflammatory processes to our understanding of immune dysfunction as a driver of aging and in the design of new rejuvenating interventions.
Experimental models of chronic inflammation
In order to refine our knowledge on the mechanisms linking inflammation and age‐related tissue dysfunction, it is critical to develop reliable experimental models that allow for the isolation of the inflammatory component of aging and the evaluation of its independent contribution to other age‐related alterations. These models would also be valuable tools to test the idea that immune modulatory interventions can indeed be applied to delay age‐related tissue alterations and improve health span.
While the etiology of age‐related inflammation remains to be fully elucidated, some recent tools are starting to be used to understand and mimic the state of ‘inflammaging’ and evaluate its consequences. Although accumulation of senescent cells has been postulated to be an important source of age‐related chronic inflammation , studies that directly test the contribution of the pro‐inflammatory component of the Senescence Associated Secretory Phenotype (SASP) on tissue function and physiology in the context of aging are yet to be performed. Experiments in which young animals are exposed to senescent cells support the notion that signals derived from senescent cell contribute to the propagation of aging phenotypes . The next step would be to directly test the effects of long‐term exposure of young mice to combinations of exogenous SASP factors, in an attempt to establish a new model of accelerated aging driven by senescence‐associated inflammatory factors. Such manipulation could be envisioned as a new experimental paradigm to model age‐related inflammation and its consequences. As an alternative approach, researchers have used a mouse model of increased NFκB signaling [26, 27], a key signaling pathway driving SASP‐related pro‐inflammatory cytokines , and a pro‐inflammatory mediator over‐activated during normal aging, and in premature aging models . Supporting a causal relation between chronic inflammation and age‐related tissue dysfunction, the low‐level elevated inflammatory phenotype of NFκB gain of function mice (Nfkb1−/−) is associated with a shorter lifespan and signs of accelerated aging [26, 27]. In this mouse model, abnormalities emerge at middle age and include alopecia, kyphosis, osteoporosis, central nervous system alterations, enhanced cellular senescence, and reduced regenerative capacity [26, 27]. Importantly, anti‐inflammatory treatments in these mice reverse at least part of these phenotypes , demonstrating that, in this model, inflammation has a causal relation to the development of other age‐related phenotypes.
An alternative model of chronic inflammation relies on the loss of function (LOF) of the anti‐inflammatory cytokine, interleukin 10 (IL‐10). Here, the loss of anti‐inflammatory signaling results in a low grade pro‐inflammatory state that is accompanied by signs of accelerated frailty at middle age. IL‐10 LOF mice are characterized by multiple alterations in skeletal muscle [30–32], along with changes in the cardiovascular system  and metabolic dysfunction , commonly observed in human frailty.
Recently, we described a new mouse model of chronic inflammation  that relies on the LOF of mesencephalic astrocyte‐derived neurotrophic factor (MANF), an immune modulatory molecule with pro‐repair activity . MANF is a secreted factor and its levels decline with age in circulation and several tissues of Drosophila, mice and humans . MANF null mice are embryonic lethal  or die before adulthood due to acute beta cell loss , depending on the genetic background. Interestingly, we found that heterozygous carriers of the MANF null allele develop signs of chronic inflammation, characterized by increased levels of pro‐inflammatory cytokines and accumulation of activated macrophages in different organs. This inflammatory state is accompanied by liver disease at middle age, resembling phenotypes observed at old age .
One interesting observation arising from these multiple models is that creating a pro‐inflammatory phenotype does not have immediate consequences. In fact, in all three models, aging‐like phenotypes emerge at middle age (around 1 year in mice), suggesting that it is the long‐term exposure to inflammatory signaling that contributes to alterations in other cell types and organismal physiology. How this happens, however, remains unknown: one hypothesis is that the pro‐inflammatory signals act on specific receptors on target cells and that their continuous activation results in cell intrinsic alterations that contribute to aging; one other possibility is that inflammation acts as an amplifier of additional aging ‘triggers’ and contributes to the establishment of a ‘vicious circle’ that accelerates the progression of aging phenotypes. The models of chronic inflammation described above could be used to test these and other hypothesis. It would be particularly interesting to use them in combination with other stressors that can provide a second trigger and evaluate the consequences in animal physiology.
There are also evident limitations to the use of the models described above: on one hand, the pathways manipulated in such genetic models are also involved in the regulation of other aspects of cellular physiology, not necessarily related to inflammation, which may introduce confounding effects; on the other hand, each of the pathways alone is likely responsible for only part of the inflammatory milieu that exists during physiological aging. We envision several approaches that could be taken to mitigate these limitations. One such approach is to target these pathways specifically in the immune system or in cell types that are distributed systemically and can influence immune cell phenotypes (see next chapter). As an example of this approach applied to a mechanistically poorly defined model of premature aging, our recent work demonstrated that ablation of MANF specifically in Cx3Cr1+ monocytes/macrophages is sufficient to recapitulate the phenotypes of inflammation and tissue damage observed in MANFHet mice . As a complementary approach, it is also possible to consider the overlapping analysis of multiple models, including the genetic models described above. There is evidence that other nonaging related physiological conditions, such as obesity or infection (e.g., sepsis), can result in phenotypes of tissue damage and metabolic dysfunction that resemble age‐related alterations and are also mechanistically linked to inflammation and/or their age‐related sources [37–43]. Thus, a possible approach to understand the mechanistic link between inflammation and age‐related tissue dysfunction would take advantage of the combined analysis of these models, allowing for the identification of common altered pathways that are also affected during physiological aging. The knowledge of these common mechanisms would be extremely relevant to develop strategies that target inflammation to allay age‐related disease.
Systemic regulation of age‐related chronic inflammation and rejuvenation
One important feature of chronic inflammation is its systemic nature. While this means that age‐related chronic inflammation has the potential to propagate the effects of localized age‐related alterations, such as an altered microbiota, a local accumulation of senescent cells or an unresolved site of tissue damage, it also anticipates that intervening at the level of inflammation has the potential to delay the aging process by slowing the propagation of the detrimental consequences of localized events.
Heterochronic parabiosis studies represent one important landmark in aging biology demonstrating the systemic component of the aging process. The observation that it is possible to rejuvenate multiple organs by joining the circulatory system of an old mouse with that of a young mouse [44–46], or simply by transfusing the old mouse with young plasma , supports the notion that systemic mediators of the aging process exist and that the blood can function as a vehicle of propagation. Consistently, old blood also has pro‐aging effects in young organisms. Multiple circulatory factors have been identified as mediators of the pro‐rejuvenation and pro‐aging systemic effects of heterochronic parabiosis, including molecules involved in the regulation of inflammation [23, 45], but also growth factors or hormones not directly involved in the regulation of inflammatory pathways [46, 48–50], highlighting the existence of systemic regulators of aging beyond inflammation.
The mechanisms and cellular targets of systemic mediators of inflammation emerging from heterochronic parabiosis studies are not fully elucidated. While it is clear that circulatory proteins are, at least in part, responsible for these effects, it is still unclear whether they act directly on somatic cells from multiple target tissues, or whether other cellular mediators are involved in the propagation of systemic effects. Potential candidates to mediate the effect are cell types that exist in circulation or that are an integral part of all tissues of an organism, such as immune cells, endothelial cells that form the vasculature, or innervating neural fibers and associated glial cells. In fact, evidence exist that endothelial cells  and macrophages  are targets and mediators of rejuvenating effects elicited by young plasma, at least in the brain. Thus, it is tempting to speculate that in the context of age‐related chronic inflammation, immune cells and the vasculature cooperate as systemic propagators of the inflammatory signal and are potential targets for intervention. Indeed, immune cells and endothelial cells are important nodes of integration of innate immune pathways, and could be, not only targets, but also sources of systemic factors that affect aging and longevity. These cell types constitute ideal candidates for the development of cell type specific genetic models of chronic inflammation, as described above. The role of the nervous system in controlling systemic aging is also supported by experimental evidence, and involves the propagation of local inflammatory signaling originating in the hypothalamus through modulation of the systemic levels of gonadotropin‐releasing hormone (GnRH): age‐related increase in NF‐κB signaling in the hypothalamus results in decreased levels of GnRH that contribute to acceleration of aging phenotypes. Reducing hypothalamic inflammation of restoring GnRH levels are effective anti‐aging interventions .
While the organismal‐level effects of pro‐longevity interventions, such as rapamycin treatment or caloric restriction, are indicators of a mechanism involving systemic signals, it is still unclear whether circulatory factors responsible for the rejuvenating effects of young blood are also mediators of the effects of other rejuvenating interventions. Nevertheless, experiments in simpler organisms already point to the existence of a systemic component involving innate immunity in mediating the pro‐longevity effects of caloric restriction. In Caenorhabditis elegans metabolic regulation of immunity‐associated genes was shown to be an integral part of the pro‐longevity effects of regulated dietary input, demonstrating that inflammatory pathways, usually associated with the control of infections, are an integral part of the systemic network controlling aging and organismal homeostasis .
Immune modulation: a strategy to limit inflammaging and promote repair in older organisms
Aging is associated with a loss of regenerative capacity in multiple tissues caused by changes in the systemic environment and the niche, as well as intrinsic limitations of stem cells . A key change in the aged environment preventing effective regeneration is the altered composition of inflammatory signals that arises from immune dysfunction (Fig. Fig. 2). The inflammatory component of the repair response consists of a biphasic activation of the immune system (Fig. Fig. 2, top), which is dynamically coordinated with the function of stem cells to optimize tissue reconstruction . The initial phase of the repair response is dominated by the activation of immune signals involved in pathogen and debris clearance and which synergistically contribute of the activation of stem cells, promoting their proliferative activity. This first wave of activation is defined as pro‐inflammatory. The second wave of immune activation, usually referred to as anti‐inflammatory or pro‐repair, represents a shift in cellular phenotypes and molecular effectors and involves signals promoting stem cell differentiation and tissue remodeling . An effective repair response requires the proper activation of both phases on the inflammatory response, a timely transition between the two phases, and a resolution process that ensure the de‐activation of the immune system once injury is eliminated, and the resulting cessation of inflammation . Thus, although inflammatory pathways are essential regulators of the regenerative response, their continuous activation in aging is considered a limiting factor for optimal stem cell function (Fig. Fig. 2, bottom).
Several studies with fly and mouse models support the notion that age‐related inflammation contributes to the loss of regenerative capacity. The NF‐κB/Rel and JAK/STAT signaling pathways, in particular, have been repeatedly linked with regenerative homeostasis and seem to play a central role in the age‐associated decline in regenerative potential. The importance of altered NF‐κB signaling in limiting regeneration in aged tissues is suported by the analysis of genetic mouse models of chronic low‐grade inflammation induced by NF‐κB over activation (see above), which present an impaired regenerative response in the liver and gut and can be rescued by pharmacological anti‐inflammatory interventions . Work done in skeletal muscle, point to local effects of increased NF‐κB activity in aged muscle fibers, as well as an hyperactivation of the JAK/STAT signaling pathway, as important factors limiting the myogenic potential of old muscle stem cells. Muscle stem cell‐specific inhibition of JAK/STAT signaling or the systemic administration of an NF‐κB inhibitor proved to be effective in improving regenerative capacity of aged skeletal muscle [24, 57]. The inhibition of JAK/STAT signaling is similarly effective in enhancing stem cell activity in the aging skin, where alterations in cytokine composition directly impact and inhibit epidermal stem cell function . Also in flies, chronic activation of JAK/STAT signaling in the aging intestine has a detrimental impact in stem cell function, inducing metaplasia of the gastric epithelium . Consistently, strategies that prevent chronic activation of the pro‐inflammatory NF‐κB/Rel and JAK/STAT signaling pathways in the fly intestine are effective in promoting regenerative homeostasis and extending lifespan [59, 60].
The accumulation of senescent cells is a prominent feature of aging tissues that can directly impact regeneration. Although cellular senescence can serve important roles in tissue repair [61–64], the SASP is a central contributor to the general state of chronic inflammation and dysregulated immune microenvironments found in older organisms. Thus, interventions that can selectively eliminate senescent cells have the potential improve regenerating tissue microenvironments. This idea is well supported by results from a study using the senolytic agent ABT263, which can effectively deplete senescent cells and was shown to improve the function of hematopoietic stem cells and muscle stem cells in aging mice . Other genetic and pharmacological interventions capable of clearing senescent cells from aging animals were shown to be effective in attenuating age‐related deterioration of several organs [9–11], suggesting that senolytics can have a broad application in regenerative medicine.
Given the central role of immune cells as sources and integrators of inflammatory signals, and the systemic potential of their action, modulation of immune cell phenotypes could be a target for intervention to limit ‘inflammaging’ and restore repair capacity in older organisms. Here, we define the concept of immune modulation (Fig. Fig. 2) as a set of manipulations that target immune cell phenotypes to overcome their state of persistent activation and re‐establish their ability to autoregulate an acute response to tissue insults. Immune modulatory interventions should thus include signals that regulate the timely transition between the different phases of the inflammatory response, as well as molecules that promote the resolution of inflammation when the repair process has terminated. The resolution phase of the inflammatory response is a subject of active investigation with several classes of pro‐resolving molecules identified so far . Moreover, there is evidence of delayed resolution of inflammation in old mice under infection conditions  and modulation of macrophage phenotypes with pro‐resolving compounds is sufficient to re‐establish the resolution phase , suggesting that such molecules may also be useful when combined with regenerative interventions. In contrast, factors responsible for the timely transition between the two phases of the inflammatory response are still vastly unknown, although there is evidence that phagocytosis and metabolic pathways intrinsically regulate this transition in macrophages [68, 69]. Phagocytic activity is reduced in macrophages from old individuals  and metabolic pathways are major targets of the aging process that impact innate immune signaling . Thus, the discovery of molecular regulators of these biological functions of macrophages may lead to identification of new targets for immune modulatory interventions in aging.
Indeed, proof of principle studies have emerged in recent years that support the application of immune modulatory interventions to improve regenerative success in aging: In the retina, we showed that MANF, an immune modulator with the ability to promote pro‐repair activity of macrophages, can be used as a co‐adjuvant in retinal transplants and promote functional integration of replacement cells, accelerating and enhancing vision recovery . Interestingly, systemic administration of MANF can improve immune and metabolic homeostasis in older organisms . Thus, it is likely that such systemic intervention applied in older organisms could also be used to improve regenerative success [71, 72]. In traumatic muscle wounds, modulation of the immune microenvironment by extracellular matrix‐derived biomaterial scaffolds can be used to promote regenerative success. Mechanistically, biomaterials promote T helper 2 cells mediated anti‐inflammatory polarization of local macrophages, potentiating functional tissue regeneration . One important observation arising from this study is that the local administration of the scaffolds stimulates T cell responses systemically, highlighting once again the potential of immune modulatory interventions to affect tissue function at the organism level. This may be beneficial in situations where chronic inflammation is detrimental for regenerative capacity in several tissues simultaneously, as observed in old organisms. Although these interventions have not been directly tested in aging, a recent study supports the notion that rejuvenation of the immune system is indeed a promising strategy to improve regenerative capacity and stem cell function in older organisms .
Future directions and outstanding questions
It is now widely recognized that the loss of a regulated inflammatory response has important contributions to the aging process. Nevertheless, the evidence linking ‘inflammaging’ with the overall decline in physiological function is still mostly correlational. Good models of chronic inflammation that can isolate and mimic the inflammatory component of aging are just beginning to be developed, mostly due to emerging knowledge on the determinants underlying chronic activation of inflammatory pathways in old organisms. Still, considering that the molecular signature of ‘inflammaging’ is likely driven by a multitude of alterations in inflammatory pathways , a possible approach to the problem could involve a transversal and overlapping analysis of multiple existing models, to identify common nodes of integration that can be defined as pillars of ‘inflammaging’ as we suggest above.
Since it is now clear that an intact inflammatory response is essential for many physiological functions, the overall approach in targeting inflammaging to prevent age‐related disease is shifting from strategies that involve the inhibition of inflammation to a concept of immune modulation defined by interventions that re‐establish a regulated inflammatory response. One expected outcome of such interventions is the optimization of tissue repair responses that are naturally decreased during aging, and postulated as an important contributor to age‐related tissue dysfuntion . Thus, improved regenerative capacity could be one of the mediators of rejuvenating interventions based on immune modulation.
While promising steps have been given in this direction, there are still a number of outstanding questions in the field that need to be addressed so that we can maximize the success of such interventions: First, we will need to better understand the complex network of cell types that is involved in the regulation of the inflammatory response – including immune cell subsets, but also other potential regulators such as cells integrating the vasculature, or cells integrating neuronal networks – and how their function is affected by aging. Secondly, we will need to elucidate the mechanisms balancing pro‐inflammatory and anti‐inflammatory responses and their de‐regulation during aging. This will include characterizing pathways promoting the transition between these two phases of the inflammatory response, and the factors responsible for the resolution of the inflammatory response at the end of the repair process. Finally, we will need to understand whether long‐term exposure to inflammatory signals also has secondary consequences in other cell types, including stem cells. Stem cells are, like immune cells, equipped with an inflammatory memory  and may thus be affected by long‐term exposure to inflammatory pathways. If this is the case, interventions in aging that aim to restore a regulated inflammatory response should also consider the intrinsic transformation that has already occurred in other cell types for a targeted combinatorial approach.