Nuclear Sirtuins And The Aging Of The Immune System Part 2

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4. Monocytes and Macrophages

Macrophages reside in all tissues of the body [59]. They are critical for tissue homeostasis during the development of context-specific functions, as is the case of alveolar macrophages in the lung or microglial cells in the central nervous system.

In addition to their tissue-specific roles, macrophages are also well known for their ability to phagocyte pathogens, which leads to antigen presentation and inflammation. Macrophages originate from erythro-myeloid precursors during early embryo development, or from infiltrating monocytes in adulthood. Embryo-derived and monocyte-derived macrophages are both capable of maintaining their abundance through self-renewal when required. Macrophages respond to the environment, resulting in the acquisition of a spectrum of functional states. Upon antigen stimulation, macrophages activate and po-larize into a pro- or anti-inflammatory phenotype, the so-called classically activated Ml and alternative M2 macrophages, respectively. Ml macrophages perform cytotoxic and tissue-damage proinflammatory functions, while M2 macrophages are important for re-solving inflammation and tissue repair. Macrophage function is altered in aged hosts, resulting in poor outcomes after infections and tissue degeneration [60]. The phenotypic features of aged macrophages may differ depending on the macrophage population, but many studies indicate that macrophages from old hosts have impaired phagocytic capacity and are skewed towards a more pro-inflammatory phenotype. Macrophage depletion in aged mice under immunotherapy administration is associated with reduced proinflammatory cytokine production and survival [61]. Similarly, macrophage targeting in aged mice improves peripheral nerve structure and muscular performance [62]. Together this data indicates that macrophage deregulation with age is a major contributor to overall organismal aging.

Various lines of evidence indicate that nuclear sirtuins support immunosuppressive functions and M2-associated responses (Figures 2 and 4). For instance, SIRT6 expression increases in mouse BM macrophages under M2-polarizing conditions [55]. Similarly, SIRT2 expression decreases in mouse microglia upon LPS stimulation, which induces Ml polarization [56]. cistanche cholesterol Sirtuins support macrophage biology at many levels, including its cellular differentiation, self-renewal, polarization, and activation. SIRT1 and SIRT2 protein levels increase during the differentiation of human monocytes to macrophages, and their inhibition with cannabinol (Table 1) or deficiency prompts the development of a proinflammatory phenotype[46]. SIRT1 and SIRT2 prevent the premature expression of proinflammatory genes through the control of their chromatin structure. Mechanistically, SIRTl and SIRT2 interact with DNA methyltransferase 3B(DNMT3B) enzyme to promote DNA methylation in addition to limiting H3K4me3 and H3K27ac deposition [47]. BM-derived macrophages from Sirt6W LysM-Cre mice, in which the SIRT6 gene is specifically deleted in myeloid cells,have increased levels of expression of proinflammatory cytokines, including interleukin (IL)-6, tumor necrosis factor α(TNF-α), and interferon β (IFN-β), and increased migration capabilities compared to WT controls, but it is not known whether this is due to impaired cellular differentiation[55].

SIRT1,SIRT2, SIRT6 and SIRT7 activities are also important for the balance of macrophage polarization (Figure 4), which depends on specific stimuli and downstream signaling events [66]. M1 polarization happens in response to triggers sluch as LPS or IFN-y and strongly depends on the nuclear factor-kB(NF-KB) transcription factor, a master regulator of inflammation and age-related pathways [13,66]. M2 polarization is induced byy stimuli such as IL-4 or ⅡL-10 and uses different cascade signals, including signal transducer and activator of transcription 6(STAT6)and peroxisome proliferator-activated receptor y (PPARy) activation. Many studies have shown that SIRT1, SIRT2 and SIRT6 limit macrophage inflammation through NF-kB regulation [55,57,58]. SIRT1, SIRT2, and SIRT6 knockout BM macrophages display hyperacetylation of the NF-kB p65 subunit, which raises its transcriptional activity, and increased expresssion of NF-kB target genes, including IL-6, TNF-a, and IL-1β. The biochemical and functional interplay of SIRT1, SIRT2, and SIRT6 with NF-kB is well documented in many cell types [13,65,67], suggesting that similar mechanisms of NF-kB regulation may exist in macrophages. In HeLa cells, SIRT6:silences the expression of NF-kB targets genes by deacetylating H3K9ac [13]. cistanche deserticola side effects In addition, in 293Fcells, SIRT6 promotes the expression of the NF-KB repressor,IkBa (nuclear factor of kappa light polypeptide gene enhancer in B-cells inhibitor, α), via a mechanism that involves cysteine monoubiquitination of the histone methyltransferase SUV39H1, which results in its dissociation from the IkBa gene promoter and, consequently, gene activation [68]. In mouse embryonic fibroblasts, SIRT2 directly deacetylates the p65 subunit of NF-kB at lysine 310, repressing its transcriptional activity [67]. SIRT2 deacetylates H4K16ac during the G2/M transition, but whether SIRT2 epigenetically regulates NF kB targets genes during inflammation or under similar conditions has not been explored. Finally, SIRT7 expression has been reported to decrease in an age-dependent manner in leukocytes from healthy patients. In the monocytic THP-1 cell line, PMA-mediated monocyte-to-macrophage differentiation increases SIRT7 expression, while SIRT7 overexpression increases differentiation markers in non-stimulated THP-1 cells [64].

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SIRTI and SIRT2 also play important roles in microglia activation and brain inflammation, which have significant implications for age-dependent neurodegenerative dis-eases [56,69]. SIRT1 overexpression in microglial cells also protects neural cells from amyloid-β peptide-induced death, a neurotoxic pathway related to the pathogenesis of Alzheimer disease[69]. Sirt2/mice and SIRT2 KD microglial cells challenged with LPS have a stronger microglial proinflammatory response, including higher levels of cytokine secretion and free-radical production, and celll death [56]. At the molecular level, SIRT1 and SIRT2 exert their anti-inflammatory properties by downregulating NF-kB activity. SIRT2 enzymatic capabilities are modulated by phosphorylation, and the absence of this post-translational modification on serine S331 of SIRT2 prevents NF-kB acetylation in microglial cells. Indeed, overexpression of the phospho-resistant SIRT2 S331A mutant, but not of the phospho-mimetic SIRT2 S331D mutant, in microglia results in diminished p65 subunit acetylation at lysine 310,possibly bringing about NF-kB target-gene silencing.

Although macrophages are terminally differentiated cells, they have the capacity to self-maintain through local proliferation independently of hematopoietic precursor differentiation, a feature normally associated with stem cells [70]. SIRTl participates in macrophage self-renewal by controlling cell-cycle progression and proliferation [42]. cistanche dosage reddit SIRT1 KD macrophages are less efficient in colony formation assays and display a Gl cell cycle arrest that is associated with Myc downregulation, impaired phosphorylation of E2 factor (E2F), and increased nuclear translocation of FOXOl transcription factor. Accordingly, SIRT1 deficiency results in gene silencing of the Myc and E2F pathways, which play important roles in self-renewal, and upregulation of those pathways involving FOXO factors, which are known to induce cell-cycle arrest. A similar phenotype is observed in macrophages treated with NAM (Table 1), which raises the possibility that other sirtuins are also involved in the self-renewal process.

Cistanche can anti-aging

Despite the well-established anti-inflammatory roles of sirtuins, only one study has addressed their contribution to macrophage aging and the development of age-related dis-eases. The presence of senescent cells in aged tissues promotes M1 macrophage polarization and activation, resulting in tissue inflammation and compromised insulin signaling [71]Indeed, chronic low-grade inflammation in the elderly is associated with insulin resistance and diabetes [72]. In this regard, myeloid SIRT2 has been shown to protect against glucose intolerance by controlling aged-related inflammation [63]. How SIRT2 regulates this process is explained by its functional interplay with the NLRP3 inflammasome (Figure 4), as also reported in HSCs. In macrophages, SIRT2 interacts and deacetylates the NLRP3 scaffold protein to suppress NLRP3 inflammasome assembly and activity. Importantly, SIRT2 levels decrease with age in macrophages in conjunction with increased NLPR3 acety-lation and activation. In addition, white adipose tissue previously co-cultured with aged macrophages exhibits impaired insulin signaling compared with young controls, which can be rescued with old macrophages transduced with SIRT2 or with a constitutive deacetylated NLPR3 form. This study highlights the SIRT2-NLPR3 axis in macrophages as an interesting target for reversing age-associated inflammation and improving glucose homeostasis.

5. Eosinophils

Eosinophils play important roles in defending against helminth parasite infections and allergic inflammation, such as allergic rhinnitis and asthma. Other roles include cellular metabolism, thermogenesis, and antitumoral responses. Eosinophils are produced in the bone marrow in the presence of IL-5, a process that critically depends on the GATA-1 transcription factor [73]. In humans and mice, recent evidence indicates that eosinophil frequencies diminish in the white adipose tissue of aged hosts [74]. This age-dependent drop in eosinophil abundance is correlated with the occurrence of inflammation and the development of different age-related conditions, including frailty and impaired immune response to immunization. Importantly, the transfer of young eosinophils into aged mouse recipients reduces systemic low-grade inflammation, improves physical performance, and immune differentiation and activity, highlighting the role of young eosinophils as rejuvenating agents.

Our knowledge of the role of sirtuins and eosinophils is very limited (Figures 2and 5). There is only one report describing the functional interplay between them (Figure 5A)[75]. However, some evidence suggests that sirtuins play imoortant roles in eosinophil biology. For instance, the DNA damage response is a process that is strongly connected to nuclear sirtuin activity [76], and it is known to be more robust in eosinophils than in other innate immune cells[77]. SIRT6 is important for eosinophil differentiation and function [75]. In vitro differentiation of BM cells into eosinophils is altered in the absence of SIRT6. In addition, eosinophil-mediated M2 macrophage polarization, a process that depends on eosinophiil IL-4 secretion, is also impaired in the presence of Sirt67 eosinophils. SIRT6 regulates the abundance and activity of GATA-1, a transcription factor required for eosinophil lineage commitment anddifferentiation [73]. Intriguingly, SIRT6 promotes GATA-1 transcriptional activity independent of its enzymatic activity. SIRT6 forms a ternary complex with GATA-1 and p300 to positively regulate GATA-1 activity, so it is possible that SIRT6 acts as a scaffold protein to recruit p300 acetyltransferase to this complex. Similar to what happens in aged hosts,after exposure to the cold, myeloid.Sirt6-mice have lower frequencies of eosinophils in the white adipose tissue than do WT mice. Adaptive thermogenesis requires production of cytokines, including IL-4 by eosinophils, which leads to M2 macrophage polarization and in turn facilitates the browning of white adipocytes and heat generation. Although this study addressed the role of eosinophil SIRT6 in brown adipocyte activity, it is of note that brown adipose tissue depots and function decline in the elderly [78], suggesting novel avenues for understanding the mechanisms of organismal aging and their potential relationship with eosinophil SIRT6.

differentiation, possibly through GATA-1 stabilization and activation [75]. (B)SIRT2 enhances NK cell-mediated cytotoxicity versus hepatocellular carcinoma cells, but the underlying rnolecular mechanism remains mostly unknown [79]. (C)In DCs, SIRT1 regulates cytokine expression with important consequences for subsequent Th differentiation [80-82]. SIRT1 promotes Treg differentiation through TGF-β1 productior in a HIF1-α dependent manner. SIRT1 also promotes Th17 differentiation through IRF1 deacetylation, thereby limiting its binding to the il-27p28 promoter and silencing its expression. Moreover, in response to zymosan, SIRT1 gets recruited to the il-12a gene promoter to repress its expression and limit Th1 differentiation. (D) SIRT6 is required for both DC differentiation and maturation, but the molecular mechanisms involved have not been explored [48].

6.NK Cells

NK cells are cytotoxic lymphocytes with important roles in innate immunity against viral-infected cells and tumors. NK cells secrete perforins and granzymes and express cell-death ligands on their surface to induce target cell apoptosis. In addition, NK cells secrete a variety of pro-inflammatory cytokines, including TNF-x and IFN-β, that have important roles in sustaining and amplifying immune responses through macrophage and dendritic cell activation. In aged humans, the NK cell compartment is associated with an increase in mature long-lived circulating NK cells[83]. Despite this increase, NK cell-mediated cytotoxicity, including granulesecretion and death receptor-mediated killing, is impaired, resulting in poor responses to viruses and an increase in cancer development The role of sirtuins in NK cell function has been little investigated (Figures 2 and 5). Levels of human SIRT1 expression are high in aged NK cells [84].In particular, the level of SIRT1 expression is significantly higher in human NK cells of subjects older than 85 years than in senior and young people with average ages of75 and 21 years, respectively. Similarly, the levels of heat shock protein 70 (HSP70), a protein with important roles in protein folding and a downstream effector of SIRT1 activity in the control of protein quality [85], arealso high in senior people aged over 85 years. In the same group, superoxide dismutase 2(SOD2), a major antioxidant enzyme regulated by SIRT1 in many cells [86], is also strongly expressed ir activated NK cells. cistanche extract benefits Further investigation may help us understand the role of SIRT1 in aged NFK cells and show whether SIRT1 has a functional relationship with HSP70 and SOD2.

SIRT2 promotes the activity of liver NK cells in response to hepatocellular carcinoma (Figure 5B) (HCC)[79].SIRT2 expression specifically increases in liver NK cells from HCC. induced mice, where it promotes NK cell activity. SIRT2-overexpressing NK cells secrete higher proinflammatory cytokines, cytotoxic granules, and have enhanced tumoricidal activity, whereas SIRT2 KD impairs NK cellcytotoxic activity. SIRT2 activity is correlated with increased phosphorylation of extracellular regulated kinase 1/2 (Erk1/2) and p38, two signaling pathways important for NK cell activity. Despite the importance of SIRT2 to the liver NK cell-mediated anti-tumoral response, the role of this sirtuin in NK cell aging is yet to be explored. 7.Dendritic Cells

Dendritic cells (DCs) are antigen-presenting cells that have important roles in adaptive immunity and the maintenance of self-toleralnce. At steady state, dendritic cells are highly phagocytic and continuously present self-antigens to limit T cell reactivity. Upon infection, DCs mature, resulting in the increased expression of costimulatory receptors, including CD80, CD86, and MHC-II molecules, secretion of proinflammatory cytokines, and in T cell priming. Major dendritic subtypes include conventional dendritic cells (cDCs), of myeloid origin, and plasmacytoid dendritic cells (pDCs), which originate from a lymphoid precursor. Aging causes major changes in DC activity. In general terms, while DCresponse to pathogens is diminished, there is an increased reactivity to self-antigens and increased expression of proinflammatory cytokines, which contribute to tolerance breakdown and inflammaging[87].

While SIRTl is dispensable for DC differentiation and maturation, DC SIRTl is of great importance in maintaining the balance of Th-mediated immune responses (Figures 2 and 5). Indeed, SIRT1 expression increases in DCs upon Toll-like receptor (TLR)stimulation in humans and mice, and its deletion in mice results in altered T cell polarization [80,81]However, several research groups have reported contrasting roles for SIRTl in this cell type (Figure 5C).Yang and colleagues found that DC SIRT1 directs the production of Th17 cells, a T cell subset with inflammatory properties, by restraining the production of IL-27, an anti-inflammatory cytokine that suppresses Th17 differentiation. Indeed, Sirt10 CD110-Cre mice, in which SIRTl is specifically deleted in DCs, have lower percentages of Th17 cells. At the molecular level, SIRTl interacts with and deacetylates interferon regulatory factor 1 (IRF1), a transcription factor related to IL-27 expression. IL-27 is a protein heterodimer composed of subunits p28 and Epstein-Barr-induced gene 3(EBI3)SIRT1-dependent IRF1 deacetylation reduces IRF1 binding to the il-27p28 gene promoter, resulting in its silencing, hence leading to reduced IL-27 production and the promotion of Th17 differentiation. Using a similar mouse model of Sirt1 deletion in DCs,Liu and colleagues reported that DC SIRT1 dictates the balance of Thl and regulatory T (Treg) cell production upon DC stimulation, with no alteration in the Th17 lineage. Mice with Sirt1/DCs have higher percentages of IFNyt T cells and IFNy secretion and lower percentages of FOXP3+ T cells and levels of FOXP3 mRNA.In this study,SIRT1 regulates the production of IL-12 and TGF-β1, two hallmark cytokines for Thl and Treg differentiation, respectively in a hypoxia-inducible factor a (HIFla)-dependent manner. In human DCs, SIRTl also limits the production of IL-12p70 in response to zymosan, a TLR2 stimulus involved in immunotolerance and Thl cytokine downregulation [82]. IL-12p70 is a heterodimer of p35 and p40 subunits, which are encoded by the IL-12a and IL-12b genes, respectively. Mechanistically,zymosan prompts SIRTl recruitment to the IL-12a gene promoter, resulting in chromatin compaction at nucleosome 1 and histone deacetylation, which limits IL-12p35 expression. Overall, these studies indicate that SIRTl is a major regulator of cytokine production in DCs and has important implications for subsequent T cell subset generation.

In humans and mice, SIRT6 participates in dendritic cell differentiation and maturation (Figure 5D)[48]. Compared with WT controls, Sirt6/mice have fewer cDC precursors in their bone marrow. In addition, in vitro differentiation and maturation of mouse DCs from BM cells are impaired in the absence of SIRT6. More striking results have been obtained in a human modelof cDC generation, in which SIRT6 inhibition with S6 inhibitor(Table 1)severely impairs monocyte differentiation into DCs. From a phenotypic perspective, mouse Sirt6// BM derived DCs are less mature,as measured by reduced expression of CD86,CD80, and MHCII, increased endocytic capacity, and a reduced ability to stimulate lymphocyte proliferation. Importantly,TLR engagement with LPS in Sirt6/BM derived DCs results in increased percentages of TNF-o-and IL-6-producing cells, implying that SIRT6 fine-tunes cytokine production in these cells. Overall, this study highlights the important role of SIRT6 in DCs and suggests that lack of SIRT6 in aged DC could be partially responsible for the poor immune responses and inflammaging. 8.Adaptive Immunity

While the innate immune system recognizes low-specificity repetitive motifs present in a wide range of pathogens and damaged host cells, the adaptive immune system is remarkable for its high degree of antigen specificity. B and T lymphocytes, the two cellular members of adaptive immunity, are generated in the bone marrow (Figure 2),although T cell progenitors subsequently migrate into the thymus to complete their maturation. Mature B and T cells circulate in the bloodstream and lymph system, and both express B cell receptors (BCRs) or T cell receptors (TCRs) in their membranes that are raised to recognize almost any exogenous or malignant antigens while tolerating self-antigens. Clonal diversity of antigen specificity is thus the cornerstone of the adaptive immune system. Upon recognition of infectious agents, B and T cells are activated and differentiate into effector cells or long-living memory cells. Effector lymphocytes either amplify the innate immune response by specifically targeting pathogens or through cytokine secretion, or induce the death of infected and malignant host cells, or end the immune response once the challenge has been eliminated. In the context of immunosenescence, B and T cell clonal diversity is compromised, and a substantial reduction in the ability to respond to vaccines and new pathogenic agents is observed. 9.T Cells

T lymphocytes are subdivided into helper CD4t T and cytotoxic CD8t T cells and are activated by TCR-specific antigens in a process involving cell-to-cell contacts. Cytotoxic CD8t T cells recognize malignant cells or cells infected and target them for cell death by various mechanisms, including the production of granzymes and perforins, two major pro-apoptotic factors. Helper CD4+ T cells have immunomodulatory functions and are further divided into myriad subsets, including Th1, Th2, Th9, Th17, and Treg, each of which has a distinctive group of immune cell targets and cytokine expression pattern (Figure 2)[88,89].

Although a certain degree of naive T cell production is thought to be maintained until old age, it is accepted that the main Tr cell pool is established early in life. During age-related thymus atrophy, a progressive reduction in thymic cellularity and a marked loss of tissue architecture take place. This is accompanied by an exponential decrease in thymopoiesis, with a half-life of 16 years in humans [9]. The decline in the production of new T cells with age has consequences for the pre-existing naive T cell pool and for the rest of the immune system.

On one hand, aged naive T cells become responsible for maintaining the compartment in the absence of a substantial T cell production, which makes them enter a stem-like state [91]. On the other hand, continuous exposure to new pathogens and the appearance of autoimmune disorders and chronic infections result in an enlarged pool of clonally expanded memory T cells at the expense of the peripheral naive T cell pool. This ultimately limits TCR diversity, dampens the ability of the adaptative immune system to confront new and preexisting challenges, and is a hallmark of immunosenescence [3,92]. T cell aging is accompanied by a series of T cell-intrinsic defects that include T cell exhaustion, extensive genetic and epigenetic alterations, impaired TCR signaling and loss of proteostasis and mitochondrial homeostasis [92]. cistanche genghis khan Sirtuins contribute to T cell biology (Figure 6) and to the preservation of the health-span in the T cell compartment at multiple levels: SIRT1 has a complex role regulating TCR-mediated T cell responses, senescence, and helper T cell polarization; Sirt6 knockout in T cells produces systemic inflammation in mice, and high levels of SIRT7 expression in breast cancer are linked to T cell exhaustion. Proportional activation of T cells during the immune response depends strictly on a threshold in TCR signaling that determines whether stimulated T cells mount an effective immune response or whether they become anergic. This threshold is crucial to preclude autoimmunity and can become dysregulated during aging [93,94]. TCR signaling is care-fully controlled by co-stimulatory or co-repressor receptors at the plasma membrane,and by intracellular modules that fine-tune the intensity of TCR signaling. SIRTl has emerged as an important factor for adjusting T cell responses by regulating the termination of TCR signaling (Figure 6A and Table 1). In the absence of SIRT1,T cell activation with anti-CD3 antibodies is permitted regardless of CD28 co-stimulation. In mice,TCR-stimulated Sirtl'T cells disproportionally proliferate, produce increased levels of IL-2, and are unable to enter anergy, implying that SIRTl negatively regulates TCR signaling in vivo [40,95]. In turn, this results in a loss of tolerance in Sirt1-/T cells. Indeed, while naive and activated T cells display similar SIRT1 expression levels, that of anergic T is significantly higher. Mechanistically, SIRT1-mediated termination of TCR signaling involves the transcription factor AP-1, which must be transcriptionally active if T cell effector responses are to be effective [95]. Acetylation of the c-Jun member of the AP-1 heterodimer is needed for it to be active,and SIRT1 dynamically regulates c-Jun acetylation during TCR activation [95-97]Therefore, upon TCR stimulation, when c-Jun acetylation peaks, SIRTl interacts with c-Jun to reduce its acetylation levels and thereby extinguish the AP-1-mediated TCR response Providing further evidence that SIRT1 acts as a feedback modulator of T cell activation and anergy, one study demonstrated that IL-2, which can reverse anergy in T cells, sup-presses SIRTI expression by preventing FOXO3a from binding to the Sirtl promoter. This represents a plausible mechanism for recovering TCR sensitivity [98].

(B)In B cells SIRTI deficiency results in reduced levels of MHC-II, which results in impaired cross-presentation to CD4*T cells [104]. SIRTl is also important for class switching recombination, as it represses AID expression by deacetylation of H3K9Ac and H3K14Ac at the AID promoter [46]. Contrarily, Sirt7-/splenic B cells display defective class switching recombination [18]. Faded lines indicate age-related loss offunction, and comments in red indicate age-related changes. Figure created with BioRender.com. SIRT1 deficiency results in reduced levels of MHC-Ⅱ, which results in impaired cross-presentation to CD4+ T cells [104]. SIRT1 is also important for class switching recombination, as it represses AID expression by deacetylation of H3K9Ac and H3K14Ac at theAID promoter[46]. Contrarily, Sirt7-/splenic B cells display defective class switching recombination [18]. Faded lines indicate age-related loss of function, and comments in red indicate age-related changes. Figure created with BioRender.com.

In aged T cells, different conflicting studies have reported both increased and decreased SIRTl protein and mRNA levels, indicating the existence of a complex signaling network governing SIRT1 activity in this context (Figure 6A). During CD8t T cell differentiation, IL-12 stimulation increases histone acetylation and the transcription factor basic leucine zipper ATF-like transcription factor (BATF). BATF cooperates with c-Jun to repress transcription of the SIRTI gene, ensuring a high level of histone acetylation at the T-bet promoter to drive increased ATP production and T cell differentiation into effector cells [99]. The level of BATF expression is higher in old CD4+ T cells, and the accessibility of BATF binding motifs increasesas CD8+T cells age [105]. These observations indicate that SIRT1 downregulation may couple T cell activation with T cell aging[106]. In agreement with this model, Sirtl/mice develop spontaneous autoimmunity indicating that SIRT1's role in the maintenance of peripheral tolerance may be important to prevent this pattern of T immunosenescence [107]. In human elderly individuals, SIRT1 expression is significantly reduced in peripheral blood mononuclear cells but whether this is due to aberrant epigenetic regulation of the SIRT1 locus and whether it has a significant impact on T cell responsiveness is not known [108].

During T cell aging, downregulation of the microRNA miR-18la in aged naive and memory T cells has also been described to impact SIRT1 levels. miR-18la fine-tunes T cell activation by regulating the expression of proteins that affect the intensity and outcome of TCR signaling. In aged human T cells, miR-18la downregulation enhances the expression of several negative feedback regulators of TCR signaling, including SIRT1 thus raising the threshold of T cell activationand reducing T cell sensitivity [100]. Notably, SIRTl inhibition or silencing in cycling aged human T cells not only restores cell cycle progression but also reduces their replication stress [109]. This is in contrast to observations in primary mouse fibroblasts, in which the absence of SIRTl is associated with abnormal DNA replication [110].

Overall, these studies indicate that SIRTI expression is strictly calibrated to ensure proper T cell responses and that SIRT1 deregulation in aged T cells may either predispose to enhanced T responsiveness in the case of SIRT1 downregulation and to poor T cell responses in the case of SIRT1 upregulation.

The accumulation of terminally differentiated CD8*CD28 T cells is another hallmark of immunosenescence, and SIRT1 has been linked to the aging of these cells [1,112]. In the absence of CD28 co-stimulation, CD8*CD28 T cells are highly cytotoxic, express proinflammatory cytokines and acquire characteristics of replicative senescence [113]. During aging, SIRT1 undergoes autophagy-mediated degradation in multiple murine organs, including the spleen and thymus. In aged CD8+CD28 memory T cells, SIRTl is downregulated at the protein level, with SIRT1 transcription being unaltered, and inhibition of autophagic degradation restores SIRTl abundancy [49]. Similar findings were obtained by Jeng and coworkers, who observed that human CD8+ memory T cells and, more prominently, termi-nally differentiated CD8*CD28 T cells exhibit dramatically decreased SIRT1 (but not SIRT6 or SIRT7) protein levels, without any alteration in its gene expression. Mechanistically, loss of SIRT1 enhances the proteasomal degradation of its target FOXOl and thereby in-creases the glycolytic capacity and the cytotoxic effector functions of these memory T cells (Figure 6A)[102]. Indeed, FOXOl has recently been reported to prevent senescence and negatively regulate activation and terminal differentiation in CD8+T cells [114]. Therefore, SIRT1 loss during the latest stages of CD8+Tcell differentiation could contribute to inflam-maging by promoting the accumulation of active and highly cytotoxic CD8t T cells. In contrast with the anti-inflammatory roles generally attributed to sirtuins, SIRT1 has been shown to contribute to a general pro-inflammatory phenotype by suppressing Treg activity. This is relevant because activated Tregs accumulate at the periphery in aged individuals, probably due to the pro-inflammatory context imposed by age [45,15,16]. FOXP3 deacetylation by SIRT1 makes it more prone to proteasomal degradation, thereby reducing the suppressive Treg function in in vitro suppression assays. Conversely, sirtuin inhibition with NAM signifi-cantly increases Treg cell frequency and function in vitro(Table 1)[43,44]. Furthermore, specific deletion of Sirtl in FOXP3+ cells increases FCXP3 levels and Treg function in vivo, thereby improving survival in allograft transplants [118]. As further evidence that SIRTl promotes pro-inflammatory T cell phenotypes, SIRT1 hasbeen found to be involved in the differentiation of Th17 cells. These are CD4+ T cells that havean important inflammatory function in bacterial and fungal infections and that have been linkeed to several inflammation-associated diseases SIRT1 is upregulated during Th17 differentiatior and deacetylates the central Th17 transcription factor RORyt to optimize its transcriptional acttivity, so SIRT1 inhibition suppresses both Th17 differentiation and function [101]. Conversely, 5SIRT1 regulates STAT3 acetylation to determine its cellular distribution. SIRT1 activation with different agonists(Table 1)reduces STAT3 translo-cation to the nucleus and in turn impairs transcription of the STAT3 target RORC(which codes for RORy), thereby blocking Th17 differentiation [41]. SIRT1 then negatively regulates RORy levels while increasing its transcriptional activity. Whether these two opposing functions need to be balanced to control Th17 generation and whether this is context-dependent remain to be unexplored. Finally, SIRT1 has also been described as negatively regulating CD4t T cell differentiation into Th9 cells via a mechanistic target of rapamycin (mTOR)-HIF1α-dependent mechanism [103]. Although the role of SIRT1in effector helper T cells during aging has not yet been investigated, the established importance of SIRT1 in fate decisions during helper T cell differentiation suggests that SIRT1-deregulated expression and activity probably affect the imbalance in CD4+ T cell subpopulations thatis observed at the onset of aging and disease.

In a paper studying the gene expression changes occurring during immunosenesce in rats, SIRT2 protein levels were found to be significantly decreased in the spleen and, more prominently, in the thymus of aged rats. This was in contrast with the fact that old thymus also displayed reduced levels of the SIRT2 target H4K16Ac [18]. More generally, H4K16Ac hypoacetylation has previously been linked to replicative senesce and found to be relatively weak in several aged murine tissues. The authors proposed a plausible explanation for the lower levels of both SIRT2 and H4K16Ac, wherein the weaker association of MOF, the main H4K16-acetyltransferase, with the nuclear lamina was responsible for the hypoacetylation of H4K16 [119].

Changes in DNA methylation and loss of silent heterochromatin regions during aging, and, in particular, in immunosenescence are the epigenetic disorders most commonly recognized as appearing with advancing age. In this context, the heterochromatin mark H3K9me3 is known to be weaker in aged humans. Although its age-dependent changes seem to be context- and species-dependent,lower levels of H3K9me3 are also observed in the spleens of aged rats, which exhibit a concomitant decrease in the levels of the H3K9 methyltransferase SUV39H1 [118]. Indeed, double knockout of Sua39hl and Suv39h2 recapitulates many defects of immunosenescence in mice, including thymic involution, decreased lymphocyte production, higher memory/naive cell ratio and more HSC priming toward the myeloid lineage. Moreover, regulation of H3K9me3 by SUV39H1 has been shown to determine fate decisions in naive CD8+ T cells. In the absence of SUV39H1 CD8+T cells are unable to repress memory transcriptional programs and therefore impair the capacity to acquire effector functions. Instead, a higher percentage of CD8+ T cells develop into memory T cells, which results in sustained survival and increased long-term memory. Therefore, SUV39H1-mediated H3K9me3 seems to be important for silencing memory programs upon activation in T cells, potentially influencing the decrease in the naive repertoire observed during aging [120].

Of the many roles that sirtuins play in maintaining heterochromatin in non-immune cells, SUV39Hl is one key target of SIRT6 and SIRT1, suggesting that they may also be im-portant for regulating H3K9me3 in immune cells. SIRT6 mediates the monoubiquitination of SUV39H1, which prevents its binding to chromatin and thereby its H3K9methylation activity [68]. Conversely, SIRT1 directly regulates SUV39H1 function by deacetylation. In the absence of SIRT1, SUV39H1 activity is dramatically impaired, resulting in los of H3K9Ac and heterochomatin protein 1 α (HPlo) foci and, in turn, heterochromatin destabilization [11].

Finally,SIRT6 is also involved in immunosenescence and inflammaging since it regu-lates T cell inflammatory responses. Seminal studies of the role of SIRT6 in aging showed that Sirt6/mice displayed a severe progeroid phenotype involving profound lymphopenia and died within the first month of life. However, Sirt6/ lymphocytes normally developed in competitive transplantation assays, indicating a cell-extrinsic phenotype [25]. A subse-quent study reported massive multiorgan inflammation in Sirt6/mice, most prominently in their livers. Histological analysis indicatted strong infiltration of CD3+ T cells and, to a lesser extent, of F4/80+macrophages [121]. In this study, targeted deletion of Sirt6 in T cells or in the myeloid lineage, but not in hepatocytes, recapitulated the inflammatory and fibrotic phenotype in the liver, indicating that SIRT6 regulates inflammation in an immune-cell-autonomous manner. While SIRT7 has not been explicitly studied in the context of immunosenescence, Huo and coworkers reported that high levels of SIRT7 expression in breast cancer cells are correlated with poor prognosis, T cell exhaustion, and infiltration of pro-inflammatory M1-type macrophages[122], suggesting that SIRT7 activity may contribute to inflammaging and be detrimental to T cell homeostasis during aging.

This article is extracted from Genes 2021, 12, 1856. https://doi.org/10.3390/genes12121856 https://www.mdpi.com/journal/genes