Aging is a progressive process that entails changes in cellular function due to genetic, metabolic, or physical alterations. It involves reduced functionality and a higher probability of death [1]. CD38 is a transmembrane glycoprotein widely expressed in the body, with enzymatic functions related to reducing NAD⁺ levels, and receptor functions promoting cytokine production, migration, and activation [2]. CD38 is associated with differentiation, regulatory functions, and activation of inflammatory cell subsets [3]. The involvement of CD38 in inflammation is complex, as it can both promote and inhibit inflammatory responses depending on the conditions [4]. It is evaluated as a contributor to inflammation and as an activation marker in various diseases, including obesity, chronic inflammatory conditions, cardiovascular diseases, and Human Immunodeficiency Virus infection [5]. In aging, CD38 is overexpressed, contributing to NAD⁺ depletion and mitochondrial alterations [6]. Studies suggest that CD38 promotes inflammaging and immunosenescence, although further research is needed to understand its specific effects on T lymphocyte populations associated with aging. T cell senescence refers to the progressive deterioration and dysfunction of T cells due to aging or chronic immune stimulation [7]. Senescent T cells display various hallmarks, including decreased proliferative capacity, shortened telomeres, CD27 and CD28 downregulation, and Senescent Associated β-galactosidase (SA-β-gal) expression [8]. Inflammaging refers to the chronic low-grade inflammation that occurs with aging [9]. The immune system changes with age, leading to increased production of pro-inflammatory cytokines and activation of immune cells. Senescent T cells and memory T cells with an exhausted phenotype accumulate during aging and contribute to the pro-inflammatory environment. These cells display altered cytokine production patterns and express receptors associated with immune exhaustion, such as programmed death 1 (PD-1) and T cell immunoglobulin and mucin domain 3 (TIM-3) [10-12]. Inflammaging can lead to dysregulated immune responses, impaired immune surveillance, and increased susceptibility to infections and chronic diseases; it also contributes to tissue damage and dysfunction in various organs [13]. Chronic inflammation affects the overall functionality of the immune system, including T cell responses affected by the expression of inhibitory receptors conferring limited functionality.

CD38 activity impacts NAD⁺ metabolism, cADPR, and intracellular calcium levels [14,15]. We hypothesize that dysregulation of these processes in inflammaging, triggered by increased CD38 expression, could affect T cell function and contribute to immunosenescence and exhaustion. Here we review the involvement of CD38 in proinflammatory responses and its possible effects on T cell phenotypic and functional changes throughout life. We suggest that regulatory, effector, and memory T cell subpopulations are affected by the contribution of CD38 to the mechanisms connecting inflammaging, immunosenescence, and exhaustion.

From a biological perspective, aging is a progressive process that increases with the accumulation of years of life. It highlights changes in cellular function due to genetic, metabolic, or physical alterations. Additionally, it involves reduced functionality and a higher probability of death [1]. Aging is accompanied by the accumulation of senescent cells, low-grade chronic inflammation (inflammaging), and immunosenescence. Despite their different causes, these processes might be interconnected in aging. Cellular senescence is a biological state of permanent growth arrest, occurring after numerous cell divisions or in response to prolonged stress signals such as DNA damage and oncogene activation, preventing damage propagation in cells [16,17]. Senescent cells present the SASP, driven by Nuclear Factor kappa B (NF-κB) via DNA damage response mechanisms and cyclic GMP–AMP synthase (cGAS)–Stimulator of Interferon Genes (STING) pathway. SASP is represented by markers such as interleukins (IL-1, IL-6, IL-8), chemokines, and matrix metalloproteinases [18,19]. These factors induce inflammation, tissue remodeling, and immune cell recruitment, impacting aging and tumorigenesis [20]. Also, during senescence, the downregulation of sirtuin 1 (SIRT1) enhances IL-6 and IL-8 expression, reinforcing the senescent state [21]. Although SASP significantly reinforces the senescent cell state, its expression is tissue-dependent.

Aging is not synonymous with senescence, but there is an accumulation of senescent cells during aging. When a cell becomes senescent, the organism initiates mechanisms for its remotion by CD8⁺ T cells, CD4⁺ cytotoxic T cells, M1 macrophages, natural killer cells, and neutrophils [22,23]. Nevertheless, not all senescent cells are eliminated; therefore, they accumulate with age [24]. The inefficient removal of senescent cells is explained mainly by the fact that immune system cells are also affected by this cellular state [25,26].

In addition to senescent cell accumulation in aging, in aged adults or murine models, the production of inflammatory mediators is increased and sustained without antigen stimulation. This scenario, called inflammaging, describes the sterile inflammation present in aged individuals with elevated concentrations of proinflammatory mediators such as C-reactive protein, IL-12, IL-1β, tumor necrosis factor-alpha (TNF-α), IL-6, interferon-gamma (IFN-γ), IL-17, and IL-23, and increased concentrations of some anti-inflammatory mediators like IL-10 and Transforming Growth Factor-beta (TGF-β), compared to younger counterparts [27]. Since senescent cells accumulate within age and present SASP, these environmental alterations would contribute to inflammaging.

Another scenario interrelated with senescence, inflammaging, and aging is immunosenescence. Immunosenescence denotes the changes or loss of functionality of immune cells related to aging. Three contrasting processes characterize immunosenescence: chronic activation, moderate inflammation (inflammaging), and reduced capacity to mount efficient immune responses against infection or vaccination [28].

CD38 is a transmembrane glycoprotein broadly expressed in the organism. This glycoprotein was first described in human thymocytes [29]. It is known that CD38 possesses enzymatic functions as a glycohydrolase of NAD⁺ and as an ADPR-ribosyl cyclase, contributing to NAD⁺ depletion in aging [30,31]. Also, CD38 participates in nicotinic acid adenine dinucleotide phosphate (NAADP) generation by catalyzing the exchange of the nicotinamide group of nicotinamide adenine dinucleotide phosphate with nicotinic acid [32,33]. In addition, the role of CD38 in the activity of immune system cells has also been described as a receptor. In this sense, CD38 is linked to differentiation and regulatory functions in immune cells [34-37]. However, some of its principal roles are as a promoter of inflammatory cell subsets, as described below.

The role of CD38 in inflammation is studied mainly in innate immune system cells. The in vitro activation of macrophages with lipopolysaccharide (LPS) and IFN-γ has proposed CD38 as a marker of M1 macrophages. Also, in murine models, its enhanced expression promotes the expansion of TNF-α+ macrophages (Figure 1) [38]. Later, this was also observed in human macrophages, where CD38 expression was associated with higher secretion of IL-1β, IL-6, and IL-12p40 in response to LPS stimulation [39]. In addition, CD38 activation promotes the expression of mRNA of Il-10, IL-1β, and IL-6 in lymphocytes [40,41]. In this regard, the expression of CD38 related to Liver X receptor (LXR) activation promotes the protection of macrophages in the face of Salmonella typhimurium infection [42]. In murine models of infection by Listeria monocytogenes, CD38 is necessary for the resolution of infection since there is greater death among infected CD38 knockout (KO) mice compared with wild-type (WT) counterparts [43]. Moreover, CD38 could limit the growth of bacteria like H. influenzae, H. aegyptius, H. haemolyticus, H. parainfluenzae, and H. parahaemolyticus by NAD⁺ consumption [44]. According to this, the contribution of CD38 to innate immune responses is related not only to the increase in the number of proinflammatory macrophages but also to the production of its effector molecules, thus favoring inflammation and antibacterial clearance.

However, the proinflammatory effects of CD38 are not restricted to immune cells against potential pathogens since, in rats with cecal ligation puncture, the presence of CD38 promotes apoptosis and oxidative stress and favors structural damage in the hippocampus [45]. In the same model, inhibiting the enzymatic pathway elicited by CD38 reduces sepsis-induced intracellular calcium mobilization and mitigates injury in heart, liver, and kidney cells [46]. It is possible to think about CD38 as a promoter of infection-induced tissue damage. In this regard, in a colitis model induced by Dextran Sodium Sulfate, there is infiltration of CD38⁺ cells related to the development of severe colitis seen as an enrichment of CD38⁺ CD4⁺ T cells in the large intestine, and CD4⁺ and CD8⁺ CD38⁺ in the spleen [47]. Additionally, there is evidence that CD38 favors the recruitment of neutrophils in a mouse model of Streptococcus pneumoniae infection, proposing that the abrogation of its enzymatic activity could avoid or mitigate tissue damage [48]. According to this, knocking down or enzymatic inhibition of CD38 reduces M1 macrophage polarization in an NF-κB-dependent manner and mitigates acute kidney injury due to lower secretion of IL-1β, IL-6, IL-12β, and inducible nitric oxide synthase (iNOS) [49]. In this regard, experiments are necessary to elucidate the contribution of its enzymatic activity and the possible effect on NF-κB proinflammatory effects. Also, the reports described here demonstrate the role of CD38 in the acute inflammatory response in innate immune cells of young models. However, the possible effects of aging and the contribution of CD38 activity in adaptative immune cells are yet to be fully understood since acute vs. chronic inflammation suppose contrasting scenarios in immune function (Figure 1).

The glycoprotein CD38 also contributes to chronic inflammatory diseases related to aging, such as autoimmunity. In a CD38-deficient murine model with induced arthritis, there is less bone irregularity, less enrichment of T-helper (Th) 1 and Th17 cell phenotype, and, in contrast, lower levels of IL-1β, TNF-α, IL-6, IFN-γ, IL-17, IL-10, and TGF-β1. Proposing CD38 as a promoter of Th1 polarization and inflammation in autoimmune arthritis [50] (Figure 2). Furthermore, in CD38 KO mice, there is less severe glomerular damage, fewer proinflammatory Ly6C^hi monocytes, and TNF-α production by Ly6C^hi monocytes and granulocytes in a lupus erythematosus-induced model [51]. Moreover, in patients with systemic lupus erythematosus (SLE), there was an overrepresentation of non-classical monocytes and an increase in CD38 expression in these cells [39]. It seems that the presence of CD38 promotes the inflammatory scenario observed in SLE and contributes to the characteristic tissue damage in this pathology. However, it is imperative to focus on the cell types, model, or disease where CD38 is evaluated as we observed that some reports might be contradictory to the pro-inflammatory role of CD38, e.g., in healthy volunteers there is an enriched CD8⁺ T lymphocytes with a regulatory phenotype expressing CD38⁺ and T cell immunoglobulin and ITIM domain (TIGIT) showing higher production of IL-10 and fewer expression of IFN-γ compared to patients with inflammatory bowel disease (IBD) where this population is reduced [52]. This elucidates the role of CD38 in autoimmune diseases and contributes to understanding its function beyond innate immune cells. It also addresses its possible function in adaptative immune cells since, in aging, CD38 overexpression is increased alongside the enrichment of CD38-expressing cells. Therefore, it would be valuable to evaluate the role of CD38 in aging since its effects could also influence chronic pathologies such as autoimmunity.

Data from our group demonstrates that CD38 is also involved in regulatory B cell function in autoimmunity. We observed that CD38 KO mice show decreased survival after 12 months of age compared to WT animals; then, the absence of CD38 was accompanied by higher titers of IgG anti-double-stranded DNA, anti-nuclear antibodies, and severe kidney damage [36]. However, the functionality of CD38 on Bregs was evaluated on in vitro assays with cells from younger animals (8-12 weeks). Therefore, it is necessary to elucidate the effect of aging in regulatory B cells (Bregs) activation in response to proinflammatory stimulus regarding CD38 activation and enzymatic activity on in vivo models.

The regulatory role of CD38 is not restricted to Bregs; data from our group also addressed its potential role in Tregs. In these cells, CD38 positively correlates with Forkhead Box P3 (FoxP3) transcription factor expression and immunosuppressive molecules such as cytotoxic T-lymphocyte associated protein 4 (CTLA-4), PD-1, and CD69 in CD4⁺CD25- and CD4⁺CD25⁺ T cells. Also, the data show higher mean fluorescence intensity (MFI) of IL-10 in CD38⁺ Tregs compared to CD38- Tregs. Finally, this data also evidenced a higher IFN-γ/IL-10 ratio after T cell receptor (TCR) engagement in CD38 KO mice, suggesting a promoter role of CD38 in regulatory T cell function [37]. Others have observed fewer Tregs cells in CD38 KO mice and proposed that higher NAD⁺ availability due to the absence of CD38 promotes apoptosis in Tregs via ribosylation of P2X7 cytosolic receptor mediated by mono–ADP-ribosyl transferase (ART) ART2.2 [53]. While the role of CD38 on adaptive immune cells is being studied, more studies are necessary addressing its role on these cells in aging. In this sense, we also proposed that CD38 function is not limited to the regulatory function of adaptative immune cells since we have observed an altered phenotype composition of effector and memory T cells in aged CD38 KO mice (Figure 2) (unpublished). We hypothesize that CD38’s function is not limited to innate-proinflammatory effects but could play different roles in regulatory and, importantly, in effector T cells.

Beyond autoimmune diseases and chronic infections, metabolic-related pathologies also have a chronic aspect that alters homeostasis. In this regard, obesity is another disease with a chronic inflammatory profile. A study demonstrated that CD38 is necessary for the induction of obesity since CD38 depletes NAD⁺ and, therefore, less SIRT1/PGC1α (peroxisome proliferator-activated receptor-gamma coactivator 1 alpha) axis activation was observed in muscle and liver, regulating obesity development [54]. Later, it was described that CD38 promotes adipogenesis and lipogenesis in a SIRT1/PPARγ (peroxisome proliferator-activated receptor-γ dependent pathway) [55]. Obesity and aging are both associated with chronic low-grade inflammation. First, fat cells can produce proinflammatory cytokines such as IL-6 and TNF-α, among other products [56]. Then, an increase in the burden of senescent cells and the presence of SASP mediators in white adipose tissue (WAT) was observed in aged models [57,58]. In aging, there is a gradual accumulation of fat mass along with compromised stemness and adipogenesis of aged adipose progenitors and stem cells, constituting a setting for more negative effects of inflammaging-senescence [59]. Considering that the higher enzymatic activity of CD38 due to its overexpression in aging promotes inflammation, it is possible to think of a specific role of CD38 in senescence related to its metabolic processes. Metabolic stress, seen as an intracellular cholesterol accumulation, promotes the activation of CD38 via LXR-α. Authors suggest that the lysosomal processing of cholesterol is related to the effect of LXR-α promoting CD38 activation, which in turn promotes lysosomal cholesterol hydrolysis via NAADP/Ca²⁺ dependent manner and cholesterol efflux via ATP binding cassette subfamily A member 1 and G1 pathway [60,61]. Additionally, from an immunologic perspective, there is evidence that obesity favors the presence of characteristics also seen in aging, like reduced thymic cellularity, promotion of T cell differentiation towards effector memory T cell (Tem), and higher apoptosis [62,63]. Also, naïve, Tem, and Tregs from obese mice showed exhaustion markers like PD-1, TIM-3, and lymphocyte activation gene 3 (LAG-3) [62,63]. In aged individuals with obesity or metabolic alterations, CD38 could be involved in the mechanism of immune exhaustion and probably immunosenescence promoted by inflammaging. However, its effect on metabolic alterations in aging and immune cells is mainly described in macrophages. Therefore, it is necessary to evaluate the effects or contribution of CD38 to immunosenescence in adaptative immune cells. This assessment should not only consider the expression of inhibitor markers but also measure their effector and metabolic functionality in the face of acute infections in aged models (Figure 3). Also, since NAADP is a potent Ca²⁺ mobilizing second messenger that also impacts TCR signaling in T cells [64], it would be necessary to address the enzymatic activity of CD38 in calcium mobilization for T cell activation to evaluate possible alterations in T cell activation during aging.

The COVID-19 pandemic highlighted the vulnerability of elderly people to acute infections [65]. In aged individuals with SARS-CoV-2 infection, effects of inflammation and calcium signaling in a CD38-dependent manner have been hypothesized [66]. In this regard, T cell depletion related to CD38 overexpression in severe SARS-CoV-2 infection has also been reported (Figure 2) [67]. Therefore, since CD38 is proposed as a contributor to severe SARS-CoV-2 infection and aging, its study will help to understand the functionality of immunity against acute infections in aging and would even be a possible target for optimizing the response to vaccines at advanced ages. It would be necessary to evaluate the specific role of CD38 in non-immune cells in the lung and innate and adaptative immune cells with cellular metabolic and receptor perspectives.

Reports studying CD38 in aging are increasing, helping to clarify the implication of its enzymatic activity in health and lifespan. First, it is demonstrated that CD38 rises in the liver but also in adipose tissue, skeletal muscle, and spleen in aging. This CD38 elevation was related to NAD⁺ depletion, provoking mitochondrial alterations in aged mice [6]. Additionally, it would be necessary to evaluate enriched CD38⁺ subpopulations and their functionality. Also, to measure the expression of CD38 seen as MFI and its cellular effects, and finally, to distinguish CD4⁺ and CD8⁺ depending on their effector, memory, or regulatory phenotype and functionality regarding CD38 activity in aged models (Figure 3).

Moreover, since NAD⁺ depletion is related to the development of complications in aging, restoring the levels of this metabolite has gained interest [68]. 78c, a non-competitive inhibitor of CD38, reestablishes NAD⁺ levels in aged mice in hepatocytes and muscle cells, improving muscle function and reducing fibrosis and the accumulation of DNA damage [69]. Some studies have focused on the mechanisms that promote CD38 expression and the cells that overexpress CD38 in aging. To this end, in vitro stimulation with LPS of human cells and bone marrow-derived macrophages from aged mice demonstrates that the presence of SASP products promotes the expression of CD38 mRNA, favoring M1 macrophages polarization and the production of IL-6, IL-8, and iNOS [70]. Additionally, another work demonstrated the overexpression of CD38 in monocytes, macrophages, and CD8⁺ T lymphocytes from aged mice. In this work, the induction of CD38 expression by SASP contributed to NAD⁺ depletion in WAT, promoting senescent-associated release products (Figure 1) [71]. Although SASP and senescent products are related to CD38 overexpression, further investigations are needed to elucidate if other products, independently of senescence, can increase CD38 expression. In this sense, LPS, acting per se as a proinflammatory inducer, favors CD38 expression and contributes to the production of proinflammatory cytokines as described in vitro and in vivo models [39,72]. In this regard, alterations in gut microbiota with contributions to LPS production have been reported in aged mice. In this study, researchers found higher fecal and systemic levels of LPS in aged mice compared to younger mice. Also, they described increased p16 expression, an inhibitor of cyclin-dependent kinases (CDK) CDK4 and CDK6, and a senescent marker, in gut and peritoneal macrophages [39,72,73].

The elevation of senescent markers such as CDK inhibitor p16 and p21 was also observed in M1 macrophages from aged mice with higher mRNA expression of IL-6, IL-1β, and CD38 [72]. Here, researchers described that senescent products promote the expression of CD38, contributing to M1 proinflammatory macrophage phenotype and NAD⁺ depletion in aging [72]. As reviewed elsewhere, SASP mediators and pathogen-associated molecular patterns could induce CD38-dependent NAD⁺ depletion and mitochondrial defects in aging [74]. The role of CD38 in NAD⁺-dependent mitochondrial alterations in T cells has also been addressed. CD38^hi CD8⁺ T cells showed reduced NAD⁺ levels, affecting their cytotoxic function, which leads to infection susceptibility in SLE patients [75]. Another study addressed the coexpression of CD38 and PD-1 on T cells as an exhausted phenotype. In the melanoma murine model, they demonstrated that genetic ablation or antibody-mediated targeting of CD38 increases NAD⁺ levels, promoting memory-associated genes and antitumor response of T helper cells in a SIRT1-Foxo1 dependent manner [76]. Also, in SLE patients, type I IFN promotes CD38 expression, leading to NAD⁺ depletion and mitochondrial damage, impeding the cytotoxic function of CD8⁺ T cells. This effect was also observed in a lupus-prone mouse with chronic lymphocytic choriomeningitis virus infection, where CD38^hi CD8⁺ T cells had defective virus clearance (Figure 2) [77]. In addition, in a mouse model with ovalbumin peptide chronic stimulation, the CD38 deficiency protects CD8⁺ T cells from acquiring the TIM-3+ PD-1+ exhausted phenotype and the expression of Thymocyte-selection associated high mobility group-box protein expression (TOX), alongside a higher proportion of TNF-α+ IFN-γ+ T cells compared to CD38-expressing T cells. Nevertheless, neither antitumoral capacity was improved nor the exhaustion phenotype acquisition (Figure 2) [78]. Although some reports address the effects of CD38 in chronic conditions and T cell functionality, more research is still needed to assess specific populations of T lymphocytes under pathological conditions observed in aging, such as autoimmunity, cancer, and acute infections. Data from our research group suggests that LPS promotes the enrichment of CD4⁺ CD38⁺ effector memory (Tem) and CD8⁺CD38⁺ central memory T cells (Tcm) in aged mice (unpublished). We also observed that LPS promotes the expansion of CD4⁺ and CD8⁺ Tcm cells expressing CD69 or Ki67 in a CD38-dependent manner, suggesting a T cell bystander activation in aging promoted by CD38 (unpublished).

Here, we describe the main age-related changes in T lymphocytes. Several factors affect the maintenance of T cells, including thymic involution, genetic load, stressors, and lifestyles [79]. The thymus is a primary lymphoid organ where T cell precursors find a niche to develop, mature, and differentiate, and its involution is one of the main changes in the immune system during aging, affecting T cell numbers, functionality, and TCR repertoire [80]. In mice, this lymphoid organ reaches its maximal development 4 weeks after birth, and thymic involution begins [81]. In humans, thymic involution is evident as early as one year of age, and therefore, T cell homeostasis depends on the peripheral proliferation of naïve and memory T cells [82]. Considering that thymic involution starts early in both mice and humans, this process occurs in two distinct phases: one related to development and the other related to aging, as reviewed elsewhere [83]. The first tries to explain that early in life, it is imperative to achieve sufficient and diverse T cell repertoire, which implies a highly energetic demand on thymic activity to produce auto-tolerable, self-restricted, and functional T cells. Considering that there is high thymocyte apoptosis in this process, it has been proposed that thymic activity diminishes once T cell repertoire is achieved. The second phase of thymic involution regards age-dependent changes throughout life, such as senescence, inflammation, and loss of cellular functionality within the thymus microenvironment. Thymic epithelial cells are essential for T lymphocyte development. Some studies report a decrease in these cells over time, and an increase of adipose tissue, senescent cells, and fibroblasts in the thymus is observed in older individuals [81,84,85]. CD38 is differentially expressed in thymocytes across its maturation process, and it is also used together with CD4, CD8, CD25, and CD44 for monitoring T cell development in the thymus [86,87]. However, in CD38 KO mice, no differences in the frequencies of thymocytes (double negative, double positive, CD4⁺ or CD8⁺ single positive) or splenic naïve (CD62L^hiCD44^low), Tcm (CD62L^hiCD44^hi), Tem (CD62L^lowCD44^hi), and double negative (CD62L^negCD44^neg) T lymphocytes have been found in comparison with WT mice [78], suggesting that thymic and post-thymic differentiation does not require CD38. Nevertheless, since the models used in this study were 8-12 weeks old, it is necessary to evaluate these subpopulations and their functions at an advanced age to assess the role of CD38 in post-thymic differentiation across aging, which requires very different events than those involved in thymopoiesis as we have reviewed here. Indeed, we found that aged CD38 KO mice show an altered memory T cell composition with an overrepresentation of naïve and central memory CD8⁺ T cells and lower proportions of effector memory (Tem) compared to aged WT mice (Figure 2) (unpublished). This could support our hypothesis that CD38 is dispensable for post-thymic differentiation at an early age but is a potential contributor factor for peripheral T lymphocyte differentiation and activation through aging.

T lymphocyte composition undergoes reorganization throughout life. In elderly adults or aged mice, a decrease in naïve T cells and enrichment of memory T cell phenotype is observed, particularly in CD4⁺ T lymphocytes, seen as memory CD4⁺ T cells with a phenotype characterized by CD44, PD-1, and CD153 expression [88]. In addition, researchers described some phenotypes accumulated in aged mice like Tregs, Tem, cytotoxic and exhausted T cells. The transcriptome analysis identified CD81, CCL5, Eomesodermin transcription factor (Eomes), and PD-1 expression as markers for these highly differentiated cytotoxic, activated Tregs and exhausted CD4⁺ T cells present in aged mice [10].

In addition, young adults who were thymectomized in early childhood present an aged-like T cell phenotype measured by limited proliferation and diminished frequency of CD4⁺ and CD8⁺ T lymphocytes. This work describes peripheral homeostatic proliferation in naïve T cell count from thymectomized adults; nevertheless, this does not reach optimal levels to compensate for the absence of the thymus [89]. Also, the accumulation of Tregs in mice and humans is evident with aging and is related to limitations in the responsiveness of effector T cells (Teffec) since Tregs accumulation in aged individuals promotes chronic infections [90]. Others have reported less apoptosis of Treg cells in aging and increased immunosuppression markers such as CTLA-4, PD-1, IL-10, and TFG-β (Figure 3) [91]. As we previously reported, CD38 promotes the regulatory functions of Bregs and Tregs. CD38 stimulation also promotes IL-10 production by Tregs [36,37]. According to its function in regulatory T cells and its contrasting contributions on inflammatory cells (promotion of inflammation vs. regulation), some experiments are necessary to elucidate the potential role of CD38 in the Treg-suppressive role over effector functions of Teffec cells in chronic infections and cancer. First, evaluate the changes in aging in Tregs proportions and function in CD38 KO mice compared to WT mice. Furthermore, whether Tregs function is affected by CD38 activation or by its enzymatic function in aged models remains to be determined. We wonder if the age-associated proinflammatory role of CD38 could intrinsically affect the function of Teff cells or if this altered function of Tffec cells depends on regulatory mechanisms by other cells. Potentially, this could have implications for the effector/regulatory balance between T lymphocytes in aging. In this regard, does the pro-inflammatory role of CD38 overshadow its potential regulatory effect on regulatory T cells? It is still necessary to perform some experiments to evaluate if the presence of CD38 promotes the expression of inhibitory markers in Tregs, enhancing their regulatory function in aging (Figure 3).

Transcriptome analysis in aged mice suggests the aberrant function of CD4⁺ and CD8⁺ T cells according to higher activation and improved immune activity but exhausted phenotype, demonstrated by the expression of the inhibitory checkpoint receptors LAG-3 on Tcm and TIGIT, PD-L1, and CTLA-4 on Tem cells [92]. Since IL-2, IL-7, and IL-15 are homeostatic cytokines for T lymphocyte activation, maintenance, and survival, these changes could also be related to alterations in these cytokine levels. In old mice, virtual memory T cell (T_VM) accumulation has been observed; these cells expressed high amounts of CD122 (IL-2R/IL-15Rβ) and showed better proliferative potential compared to naïve T cells in response to IL-7 and IL-15 stimulation, but, when challenged with ovalbumin peptide this advantage was not observed, suggesting that aged T_VM cells show better proliferative response to homeostatic cytokine instead of TCR engagement [93]. Besides, it has been described that T cell exhaustion, seen as high expression of inhibitory receptors and increased Treg numbers, affects sepsis recovery in the elderly; researchers proposed that in aged mice, IL-15 administration rescues CD4⁺ and CD8⁺ persistent T cell exhaustion by increasing Naïve T cells and depleting CD4⁺ and CD8⁺ PD-1⁺ T cells and Treg numbers [94]. Also, severe infections are related to immunosuppression due to induction of thymic involution. There is evidence that IL-15 promotes the proliferation of memory-like CD8⁺ CD38^hi T cells with a regulatory phenotype [95]. However, since IL-15 has not been measured in CD38 KO mice, it would be necessary to determine possible alterations depending on CD38 expression and activation in aged mice.

Inflammaging negatively affects the responses elicited by the immune system and, in some cases, favors the development of inflammatory T cell subsets. Here, we recapitulate that senescent-associated cell accumulation in aging could promote proinflammatory mediators elevation, and metabolic and genotoxic alterations could also contribute to this scenario. Likewise, increased microbial translocation in aging could activate inflammation. As a low-grade chronic inflammatory state persists in aging, we suggest that inflammaging could also promote the terminal differentiation of T cells and potentially lead to cell exhaustion.

The increased Th17/Treg ratio is related to inflammatory or autoimmune diseases [96]. However, individuals over the age of 100, called centenarians, have lower inflammatory markers and are considered exemplary models of successful aging. Old adults (69.1 ± 0.6 years) present elevated Th17/Tregs ratio compared to younger adults (20-45 years old). In contrast, centenarians (102.2 ± 0.1 years) downregulate the activity of Th17 lymphocytes by limiting the release of proinflammatory cytokines from Th17 cells and increasing the production of IL-10 by Tregs, showing lower levels of proinflammatory cytokines (IL-17A, IL-12, TNF-α, and IFN-γ) in Th17 polarizing conditions compared to old adults. Interestingly, they presented lower levels of IL-6, IL-17A, IL-1β, IL-23, IL-12, and TNF-α compared to younger individuals (28.6 ± 1.6) [97]. This data could explain alleviated inflammaging in centenarians compared to old adults; however, the mechanism is still to be elucidated [97].

In contrast, Tregs with an inflammatory phenotype, called proinflammatory Tregs, are associated with less tissue damage repair in a pneumonia model because of elevated expression of T-box transcription factor T-bet, IFN-γ, the retinoic acid-related orphan receptor RORγt, and IL-17 in the aged murine model [98]. Analyzing the data here exposed, Tregs involves different phenotypes in aging. We wonder if the accumulation of Tregs in aging is still strictly related to its classical regulatory functions or if there is a broader uncharacterized phenotype of non-classical Tregs associated with malfunctioning of effector T cells in aging. In this regard, it would be interesting to evaluate Treg pool differentiation and functionality across aging and to evaluate the possible contribution of CD38 in this process since CD38 KO mice show reduced Tregs due to higher NAD⁺ availability [37,53].

Some studies have focused on changes in the metabolism of T cells as a possible cause of inflammaging. As memory T cells accumulate in aging, researchers have described increased respiratory capacity in these cells compared to naïve T cells. However, memory CD4⁺ T cells from aged adults contain higher mitochondrial content and produce higher levels of reactive oxygen species and proinflammatory cytokines after TCR activation compared to memory CD4⁺ T cells from young adults [99]. Since NAD⁺ contributes to mitochondrial homeostasis, evaluating CD38-dependent NAD⁺ levels in T cell subsets in aging would be necessary. It has been reported that naïve T cells possess a lower concentration of NAD⁺ compared to effector T cells. However, this determination did not consider stratification by age [100]. Considering the role of CD38 as a consumer of NAD⁺, it will also be necessary to elucidate its contribution to metabolic alterations in naïve and memory subpopulations due to aging. Also, it is important to mention that in nonpathological conditions, there is a higher percentage of CD4⁺ and CD8⁺ CD38⁺ naïve T cells than CD38⁺ Teffect lymphocytes. In addition, higher expression of CD38 was observed in naïve T cells and lower concentration of NAD⁺, demonstrating the NADase activity of CD38 [100]. In aged murine models, mitochondrial and NAD⁺ level alterations in T cells are related to a senescent phenotype characterized by less proliferation, activation, and higher apoptosis, contributing to chronic inflammation and promoting premature death [101,102].

Phenotypic markers are used to identify cells under senescence. The expression of p53, p21, and p16 are linked to the start of the senescence program [17,103]. Also, in senescent cells, the accumulation of lysosomal content and reduction of lysosomal pH causes the expression of another marker: β-galactosidase (lysosomal hydrolase that cleaves β-linked terminal galactosyl residues), which has been called Senescent-associated-β-Galactosidase (SA-β-Gal) [104]. Here, we have commented on accumulating more differentiated lymphocytes in the elderly. In this regard, in humans older than 60, effector memory re-expressing CD45RA, T_EMRA (CD45RA+CCR7−) lymphocytes with elevated expression of SA- β-Gal are enriched in CD8⁺ T cells. In this way, SA- β-Gal is helpful for the identification of senescent CD8⁺ T lymphocytes in aging [105]. Moreover, in CD8⁺ T cells from aged individuals, the loss of CD28 expression has been observed. Then in this population, there is an increase in CD8⁺CD28- T cells [106,107].

Among other markers associated with immunosenescence, the expression of inhibitory receptors such as PD-1 and CTLA-4 is also related to the reduced proliferative capacity of murine CD4⁺ T lymphocytes [108]. Helios and the immune receptor TIGIT are other receptors described for senescent CD4⁺ T lymphocytes. In aged individuals, Tregs cells with an activated phenotype (CD4⁺Foxp3+CD25^hiHelios^hiTIGIT+) are increased, and therefore, there is an inefficient response elicited by Teffect cells. The above is related to increased susceptibility to Streptococcus pneumoniae colonization and developing more severe symptoms in aging [109]. Besides, these markers identify not only CD4⁺ senescent cells but also CD8⁺ T cells. However, the co-expression of TIGIT and Helios in CD8⁺ T cells identify a subpopulation of CD8⁺ T lymphocytes with lower expression of the activation marker CD69 and antigen non-specificity. This phenotype is enriched in CD27+CD28- and CD27-CD28- subsets, corresponding to intermediated and late senescent phenotype, respectively, and are associated with a longer duration of coughing [110]. In this regard, the coexpression of these inhibitory markers, associated with senescence and CD38, need to be studied and evaluated, as their possible correlation to inflammaging and exhaustion. For example, as we mentioned above, CD38 and TIGIT denote CD8⁺ T cells with regulatory phenotype [52]; however, its regulatory function is mediated by IFN-γ secretion, and as a proinflammatory cytokine, it is necessary to evaluate its contribution to inflammaging since IFN-γ promotes CD38 expression and, therefore, could contribute to more low-grade inflammation. Also, does the correlation between CD38 and CTLA-4 and PD-1 [37] expression denote a better suppressive capacity of Treg cells in old models? If this coexpression is also present in Teffec, could it suggest an exhausted phenotype? In this regard, characterization of not only Tregs but also memory and effector T cell composition could be performed by analyzing pool composition, absolute numbers, and functionality in the face of acute infection in CD38 KO compared to WT-aged mice. Then, CD38⁺ cell numbers and CD38 MFI need to be evaluated in these subpopulations, along with the coexpression of inhibitory receptors and CD38 about effector and regulatory functions in aged WT mice. An additional question to address is whether the expression of inhibitory receptors, seen as exhaustion, is an effect of senescence, a senescent phenotype itself, or an effect of inflammaging mediated by CD38. Therefore, phenotypic senescent markers such as SA-β-Gal, p16, and p21 need to be evaluated in T cells, along with proliferation capacity and SASP production by T cell subpopulation. We suggest using senolytic agents, senomorphic compounds, and CD38 inhibitors for the specific identification of senescent cells among the exhausted T cells present in aging.

Finally, a cellular phenotype under debate in chronic immune responses or aging is Tcm cells since these cells are enriched in aging; however, there is evidence that 90% of these cells are T_VM. These cells have been phenotypically classified as Tcm, as they possess memory phenotype (CD44^hiD62L^hi). However, T_VM cells express CD122⁺CD62L⁺CD49d^low [111]. Indeed, T_VM are antigen-inexperienced cells [112]. These cells also have less proliferative capacity and poor IFN-γ production under TCR stimulation and interestingly accumulate in aging [113]. In this scenario, it is necessary to focus on and not misinterpret accurate Tcm with T_VM cells in aging-related studies [114]. These cells depend on IL-15 disposition [93]. Additionally, IL-12 and IL-18 can activate T_VM CD8⁺ cells, promoting rapid production of IFN-γ and enhancing antitumor response [115]. As mentioned, we observed an overrepresentation of CD8⁺ Tcm cells in CD38 KO (manuscript in preparation). It would be necessary to delineate T_VM phenotype and function in a CD38-dependent manner.

The low-grade inflammation associated with aging has taken on the name inflammaging, just as the senescence of the immune system is now called immunosenescence. Although senescence is an event that is not strictly age-related, aging promotes cellular and metabolic changes that promote the accumulation of senescent cells and their pro-inflammatory products. It is difficult to consider inflammaging as an event isolated from senescence. Thus, it is possible to think that inflammaging promotes immunosenescence and vice versa. In this sense, if we talk about a sustained inflammaging state in aging and malfunctioning of immune cells, we cannot leave aside immune exhaustion. Here, we have set out the background linking CD38 to the acute inflammatory response and inflammation associated with aging and its possible relationship to the accumulation of exhausted T cell phenotypes and their functionality in aging. The inflammatory responses elicited by CD38 in innate and adaptative immune cells should be addressed in aged models since they are mainly studied in young mice and humans. Studies regarding sepsis, cancer, and autoimmunity must consider the role of aging, inflammaging, and CD38 function as contributors to T cell exhaustion and immunosenescence in the elderly population as chronic diseases are increasing. Focusing on CD38 function in Tregs, Teffec, and memory T cell activation, differentiation, and maintenance throughout life will be helpful to understand the function of CD38 in different pathological and non-pathological conditions, allowing us to determine in which conditions it is favorable to inhibit or modulate CD38 activity. Lastly, modulating CD38 activity and its downstream signaling pathways in T cells may offer potential therapeutic avenues to mitigate inflammaging and attenuate immunosenescence in aging individuals (Figure 4).

The authors declare that they have no conflict of interest.

This work was supported by the scholarship provided by Consejo Nacional de Ciencia y Tecnología (CONACyT) No. 779719.

We thank Dr. Enrique Espinosa from Research in Integrative Immunology, INER for reviewing the manuscript.

Wendolaine Santiago-Cruz performed conceptualization, methodology, supervision, investigation, writing of original draft, review editing, and visualization. F. García-García performed writing, review, editing, and funding acquisition.