Caloric restriction promotes longevity in multiple animal models. Compounds modulating nutrient-sensing pathways have been suggested to reproduce part of the beneficial effect of caloric restriction on aging. However, none of the commonly studied caloric restriction mimetics actually produce a decrease in calories. Sodium-glucose cotransporter 2 inhibitors (SGLT2-i) are a class of drugs which lower glucose by promoting its elimination through urine, thus inducing a net loss of calories. This effect promotes a metabolic shift at the systemic level, fostering ketones and fatty acids utilization as glucose-alternative substrates, and is accompanied by a modulation of major nutrient-sensing pathways held to drive aging, e.g., mTOR and the inflammasome, overall resembling major features of caloric restriction. In addition, preliminary experimental data suggest that SGLT-2i might also have intrinsic activities independent of their systemic effects, such as the inhibition of cellular senescence. Consistently, evidence from both preclinical and clinical studies have also suggested a marked ability of SGLT-2i to ameliorate low-grade inflammation in humans, a relevant driver of aging commonly referred to as inflammaging. Considering also the amount of data from clinical trials, observational studies, and meta-analyses suggesting a tangible effect on age-related outcomes, such as cardiovascular diseases, heart failure, kidney disease, and all-cause mortality also in patients without diabetes, here we propose a framework where at least part of the benefit provided by SGLT-2i is mediated by their ability to blunt the drivers of aging. To support this postulate, we synthesize available data relative to the effect of this class on: 1- animal models of healthspan and lifespan; 2- selected molecular pillars of aging in preclinical models; 3- biomarkers of aging and especially inflammaging in humans; and 4- COVID-19-related outcomes. The burden of evidence might prompt the design of studies testing the potential employment of this class as anti-aging drugs.

Obesity is a major risk factor for type 2 diabetes and a trigger of chronic and systemic inflammation. Recent evidence suggests that an increased burden of senescent cells (SCs) in the adipose tissue of obese/diabetic animal models might underlie such pro-inflammatory phenotype. However, the role of macrophages as candidate SCs, their phenotype, the distribution of SCs among fat depots, and clinical relevance are debated. The senescence marker β-galactosidase and the macrophage marker CD68 were scored in visceral (vWAT) and subcutaneous (scWAT) adipose tissue from obese patients (n=17) undergoing bariatric surgery and control patients (n=4) subjected to cholecystectomy. A correlation was made between the number of SCs and BMI, serum insulin, and the insulin resistance (IR) index HOMA. The monocyte cell line (THP-1) was cultured in vitro in high glucose milieu (60 mM D-glucose) and subsequently co-cultured with human adipocytes (hMADS) to investigate the reciprocal inflammatory activation. In obese patients, a significantly higher number of SCs was observed in vWAT compared to scWAT; about 70% of these cells expressed the macrophage marker CD68; and the number of SCs in vWAT, but not in scWAT, positively correlated with BMI, HOMA-IR, and insulin. THP-1 cultured in vitro in high glucose milieu acquired a senescent-like phenotype (HgSMs), characterized by a polarization toward a mixed M1/M2-like secretory phenotype. Co-culturing HgSMs with hMADS elicited pro-inflammatory cytokine expression in both cell types, and defective insulin signaling in hMADS. In morbid obesity, expansion of visceral adipose depots involves an increased burden of macrophages with senescent-like phenotype that may promote a pro-inflammatory profile and impair insulin signaling in adipocytes, supporting a framework where senescent macrophages fuel obesity-induced systemic inflammation and possibly contribute to the development of IR.

Olive tree by-products have been deeply studied as an invaluable source of bioactive compounds. Several in vitro and in vivo studies showed that olive leaf extract (OLE) has anti-inflammatory and antioxidant properties. Here, we wanted to assess the valuable benefits of two less-studied OLE components—3,4-DHPEA-EDA (Oleacin, OC) and 3,4-DHPEA-EA (Oleuropein-Aglycone, OA)—directly purified from OLE using a cost-effective and environmentally sustainable method, in line with the principles of circular economy. OLE, OC and OA were then tested in human cellular models involved in acute and chronic inflammation and in the pathogenesis of viral infections, i.e., lipopolysaccharide (LPS)-treated monocyte/macrophages (THP-1) and endothelial cells (HUVECs), senescent HUVECs and Poly(I:C)-treated small airway epithelial cells (hSAECs). Results showed that OC and OA are efficient in ameliorating almost all of the pro-inflammatory readouts (IL-1β, TNF-α, IL-8, ICAM, VCAM) and reducing the release of IL-6 in all the cellular models. In hSAECs, they also modulate the expression of SOD2, NF-kB and also ACE2 and TMPRSS2, whose expression is required for SARS-CoV-2 virus entry. Overall, these data suggest the usefulness of OLE, OC and OA in controlling or preventing inflammatory responses, in particular those associated with viral respiratory infections and aging.

Chronic hyperglycemia, the diagnostic biomarker of Type 2 Diabetes Mellitus (T2DM), is a condition that fosters oxidative stress and proinflammatory signals, both involved in the promotion of cellular senescence. Senescent cells acquire a proinflammatory secretory phenotype, called SASP, exacerbating and perpetuating the detrimental effects of hyperglycemia. Bioactive compounds can exert antioxidant and anti-inflammatory properties. However, the synergistic anti-inflammatory and antioxidant effects of the most extensively investigated natural compounds have not been confirmed yet in senescent cells and in hyperglycemic conditions. Here, we exposed young and replicative senescent HUVEC (yHUVEC and sHUVEC) to a high-glucose (HG) condition (45 mM) and treated them with Polydatin (POL), Curcumin (CUR) and Quercetin (QRC), alone or in combination (MIX), to mirror the anti-inflammatory component OxiDef^TM contained in the novel nutraceutical Glicefen^TM (Mivell, Italy). In both yHUVEC and sHUVEC, the MIX significantly decreased the expression levels of inflammatory markers, such as MCP-1, IL-1β and IL-8, and ROS production. Importantly, in sHUVEC, a synergistic effect of the MIX was observed, suggesting its senomorphic activity. Moreover, the MIX was able to reduce the expression level of RAGE, a receptor involved in the activation of proinflammatory signaling. Overall, our data suggest that the consumption of nutraceuticals containing different natural compounds could be an adjuvant supplement to counteract proinflammatory and pro-oxidative signals induced by both hyperglycemic and senescence conditions.

One of the main challenges of current research on aging is to identify the complex epigenetic mechanisms involved in the acquisition of the cellular senescent phenotype. Despite some evidence suggested that epigenetic changes of DNA repetitive elements, including transposable elements (TE) sequences, are associated with replicative senescence of fibroblasts, data on different types of cells are scarce. We previously analysed genome-wide DNA methylation of young and replicative senescent human endothelial cells (HUVECs), highlighting increased levels of demethylated sequences in senescent cells. Here, we aligned the most significantly demethylated single CpG sites to the reference genome and annotated their localization inside TE sequences and found a significant hypomethylation of sequences belonging to the Long-Interspersed Element-1 (LINE-1 or L1) subfamilies L1M, L1P, and L1HS. To verify the hypothesis that L1 demethylation could be associated with increased transcription/activation of L1s and/or Alu elements (non-autonomous retroelements that usually depend on L1 sequences for reverse transcription and retrotransposition), we quantified the RNA expression levels of both L1 (generic L1 elements or site-specific L1PA2 on chromosome 14) and Alu elements in young and senescent HUVECs and human dermal fibroblasts (NHDFs). The RNA expression of Alu and L1 sequences was significantly increased in both senescent HUVECs and NHDFs, whereas the RNA transcript of L1PA2 on chromosome 14 was not significantly modulated in senescent cells. Moreover, we found an increased amount of TE DNA copies in the cytoplasm of senescent HUVECs and NHDFs. Our results support the hypothesis that TE, which are significantly increased in senescent cells, could be retrotranscribed to DNA sequences.

During aging, bone marrow mesenchymal stromal cells (MSCs)—the precursors of osteoblasts—undergo cellular senescence, losing their osteogenic potential and acquiring a pro-inflammatory secretory phenotype. These dysfunctions cause bone loss and lead to osteoporosis. Prevention and intervention at an early stage of bone loss are important, and naturally active compounds could represent a valid help in addition to diet. Here, we tested the hypothesis that the combination of two pro-osteogenic factors, namely orthosilicic acid (OA) and vitamin K2 (VK2), and three other anti-inflammatory compounds, namely curcumin (CUR), polydatin (PD) and quercetin (QCT)—that mirror the nutraceutical BlastiMin Complex^® (Mivell, Italy)—would be effective in promoting MSC osteogenesis, even of replicative senescent cells (sMSCs), and inhibiting their pro-inflammatory phenotype in vitro. Results showed that when used at non-cytotoxic doses, (i) the association of OA and VK2 promoted MSC differentiation into osteoblasts, even when cultured without other pro-differentiating factors; and (ii) CUR, PD and QCT exerted an anti-inflammatory effect on sMSCs, and also synergized with OA and VK2 in promoting the expression of the pivotal osteogenic marker ALP in these cells. Overall, these data suggest a potential role of using a combination of all of these natural compounds as a supplement to prevent or control the progression of age-related osteoporosis.