Introduction

Healthy aging is a critical issue in the current literature due to the increase in overall survival and age-related pathological conditions affecting older people [1,2,3]. Moreover, age-related changes in cardiorespiratory, nervous and musculoskeletal systems might significantly affect the risk of developing physical function impairment and limitations in independence in the activities of daily living, with crucial implications on global health care burden and social costs [4, 5].

In this scenario, understanding the biological mechanisms underpinning the aging process might be a cornerstone of a comprehensive and tailored management of age-related pathological conditions. Interestingly, mitochondrial modifications have been proposed as biological targets in cellular aging due to their key role in molecular processes involved in energy production, oxidative stress regulation, mitophagy, and regulation of programmed cell death [6]. In addition, muscle mitochondrial modifications in older people are characterized by alterations in terms of bioenergetic capacity, mitochondrial oxidative and antioxidant capacity, and transcriptome, leading to inflammatory response dysregulation and increased oxidative stress [7,8,9,10]. In this context, recent research underlined that mitochondrial modifications might be associated with the impairment of cardiorespiratory, nervous and musculoskeletal systems with detrimental consequences on body function and physical performance [11,12,13,14].

Although the mechanisms underpinning age-related mitochondrial impairment are far from being fully understood, in the recent years, a growing amount of literature underlined the potential role of different interventions targeting age-related mitochondrial modifications to slow down the complex framework of the aging process [11, 15,16,17,18,19]. Recently, our research group focused on the role of physical activity in improving mitochondrial function to pave the way to a future concept integrating a translational approach in lifestyle medicine [12]. Concurrently, growing evidence suggests a positive role of nutraceuticals and nutritional interventions to promote healthy aging with several reports underlining positive effects of specific integration in age-related diseases [13, 14]. In particular, lifestyle interventions, physical exercise and supplementation are widely used interventions to treat osteoporosis (i.e., dietary supplements, vitamin D, calcium, probiotics) in older subjects [20,21,22]. Accordingly, protein and amino acid supplementations might counteract sarcopenia boosting the positive effect of physical exercise in frail patients [23,24,25].

Recent research is now focusing on nutraceuticals with antioxidant activity to reduce autoxidation by reacting with reactive oxygen species (ROS) and consequently reduce oxidative stress, one of the major contributors to the aging process [26, 27]. ROS increase is a major hallmark of aging and is strictly related to the progressive impairment of mitochondrial oxidative phosphorylation due to constitutive age-related changes in mitochondrial structure [28, 29].

On the other hand, to the best of our knowledge, no previous systematic reviews have investigated the effects of nutraceuticals on mitochondria modifications of healthy older adults. Moreover, strong evidence supporting the role of integrating biological advances in the tailored nutritional management of older people is still lacking.

Therefore, this systematic review of randomized controlled trials (RCTs) aimed at summarizing the role of nutraceuticals and dietary supplements in modifying age-related mitochondria modifications, to guide clinicians to integrate these interventions in the complex treatment framework of mitochondria modifications in older subjects.

Methods

Registration and search strategy

This systematic review of randomized clinical trials (RCTs) has been performed in accordance with the Preferred Reporting Items for Systematic Reviews and Meta-analyses (PRISMA) statement [30]. The study protocol has been realized before study initiation and has been submitted to the International prospective register of systematic reviews (PROSPERO, https://www.crd.york.ac.uk/prospero; registration number: CRD42022313118).

Four databases (PubMed, Scopus, Web of Science, and Cochrane Central Register of Controlled Trials (CENTRAL) were systematically searched for studies published until February 22nd, 2022, adopting the strategy depicted by Supplementary Table 1. The keywords considered were: "aged”, "elderly", "aging", "diet", "supplement", "nutrition", "macronutrient", "micronutrients", and "mitochondria". Additionally, the reference lists of all the primary studies have been screened for all the potentially relevant papers and these articles were reviewed for additional references. The analysis was performed independently and synchronously by two of the authors.

Selection criteria

After duplicates removal, two authors independently screened the title and abstract of all papers identified for eligibility. If agreement was not achieved by the two reviewers, a third reviewer was consulted. The selected articles were subsequently screened in full text by the two authors. Likewise, if a consensus was not achieved, the discrepancy was resolved by consulting one of the other authors.

The inclusion criteria for the screened articles were set in accordance with the PICO model:

  1. 1.

    (P) Participants: healthy older people (aged more than 60 years), according to the definition by World Health Organization [31];

  2. 2.

    (I) Intervention: any nutraceutical treatment administered as exclusive intervention;

  3. 3.

    (C) Comparator: any comparator, including placebo, other pharmacological treatment, non-pharmacological treatment, or no treatment;

  4. 4.

    (O) Outcomes: mitochondrial modifications assessed by plasmatic changes or tissue biopsies. Mitochondrial modifications assessed included: (a) mitochondrial oxidative capacity; (b) mitochondrial antioxidant capacity; (c) mitochondrial volume; (d) mitochondrial bioenergetic capacity; (e) mitochondrial transcriptome.

We included only RCTs, published in peer-reviewed international journals. Exclusion criteria were: (a) language other than English; (b) study design different from RCTs; (c) papers involving animal models; (d) participants with metabolic, cardiovascular, pulmonary, or other pathological conditions related to potential mitochondrial alterations; (e) other treatments as main intervention.

Data extraction and synthesis

Full-text assessments and data extraction were performed by two authors, independently. Any divergences between the two reviewers were solved by collegial discussion among the authors. In case of further disagreement, a third author was asked.

The following data were extracted: (1) title; (2) authors; (3) publication year; (4) nationality; (5) participants (number, mean age and age range, gender, body mass index — BMI); (6) interventions’ characteristics; (7) comparator; (8) study results. For all studies, both study characteristics and data extracted were summarized.

Quality assessment and risk of bias

The quality of the RCTs included was assessed by two of the authors independently, according to the Jadad scale [32]. Discordance between the two authors was resolved collegially among the research team. Jadad scale is composed by five items, with a total score ranging from zero to five based on adherence and the items evaluated were the following: (a) random sequence generation; (b) appropriate randomization; (c) blinding of participants or personnel; (d) blinding of outcome assessors; (e) withdrawals and dropouts. A Jadad score between three to five points was considered high quality.

The risk of bias was assessed by the Cochrane risk-of-bias tool for randomized trials (RoBv.2) [33]. Bias was classified as low, high, or with some concerns based on the item of the RoBv.2. In particular, the following domains were assessed by RoBv.2: (a) randomization process; (b) deviation from the intended interventions; (c) missing outcome data; (d) measurement of the outcome; (e) selection of the reported result.

Results

Study characteristics

Altogether, 8489 records were identified from the 4 databases. After duplication removal, 4534 studies were assessed for eligibility and screened for title and abstract. Therefore, 4495 records were excluded, and 39 full-text studies were screened. Subsequently, 33 articles were excluded for inconsistency with the eligibility criteria. Supplementary Table 2 reported the list of excluded studies along with the reasons for exclusion. Finally, 6 studies [34,35,36,37,38,39] were included in the qualitative synthesis. The search process is shown in detail in the PRISMA flow diagram depicted in Fig. 1.

Fig. 1
figure 1

PRISMA 2020 Flow Diagram

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The studies included in this systematic review were published between 2016 [36] and 2022 [38]. Three studies (50%) were conducted in the USA [35, 36, 38], while the remaining 3 studies were carried out in Serbia (n = 1, 16.7% [37]), the UK (n = 1, 16.7% [34]), and the Netherlands (n = 1, 16.7% [39]). The sample size of the RCTs included ranged from 12 [34] to 66 [38], for a total of 201 healthy older adults included in the systematic review.

The mean age of the subjects included ranged from 67.0 ± 1.0 years [35] to 76.0 ± 5.6 years [37]. It should be noticed that Yoshino et al. [36] did not characterize the study sample for gender differences. As a result, based on the remaining 181 participants, the sample of the present study was composed of 83 males and 98 females, with a body composition assessed by BMI ranging from 25.3 ± 1.3 kg/m2 [36] to 28.6 ± 3.9 kg/m2 [37]. Interestingly, 4 studies did not report any standardization in terms of diet and physical activity during the study protocol, while both Zanini et al. [37] and Yoshino et al. [36] instructed study participants to maintain usual diet and physical activity.

Control groups were characterized by placebo administration in all the studies included in the present review (n = 6, 100% [34,35,36,37,38,39]), indistinguishable from the nutraceutical treatment administered in the intervention group. The characteristics of the studies included are presented in detail in Table 1.

Table 1 Main characteristics of the articles included in the present systematic review
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Nutraceutical intervention

The intervention was characterized by the administration of several nutraceuticals based on the study included. More in detail, the experimental treatment included: sodium nitrite [35], fish oil-derived n-3 polyunsaturated fatty acids (PUFA) [36], hydrogen-rich water [37], nicotinamide riboside (NR) [34], urolithin A [38], and whey protein powder (L-Tryptophan [L-Trp], nicotinic acid [NA], and nicotinamide [NAM]) [39].

  • Sodium Nitrite. Sodium nitrite was considered as a key intermediary of nitrate–nitrite–NO pathway, increasing NO bioavailability and affecting nitrite-mediated oxidative stress [40]. It was administrated in the study of Rossman et al. [35] for a total of 80 mg per day (40 mg, 2 times per day). The duration of sodium nitrite supplementation was 12 weeks [35].

  • Fish oil-derived n-3 PUFA. N-3 PUFAs supplementation was performed in older subjects according to the preclinical evidence about the increased gene expression of master regulators of mitochondrial function (PPARGC1A and UCP3) and the effects on metabolic and regenerative pathways potentially related to the inhibition of autophagy and atrophy pathways [41, 42]. Fish oil-derived n-3 PUFA was administrated in the study by Yoshino et al. [36]. The study participants in the n-3 PUFA group received four 1-g (LOVAZA®) pills per day, composed of 1.86 g eicosapentaenoic acid (20:5 n-3 PUFA) and 1.50 g docosahexaenoic acid (22:6 n-3 PUFA. The protocol duration was 6 months [36].

  • Hydrogen-rich water. Hydrogen-rich water supplementation has been proposed due to the evidence supporting hydrogen selective interaction with hydroxyl radical and peroxynitrite, promoting the reduction of oxidative stress and positive effects in inflammatory and apoptotic pathways [43]. Hydrogen-rich water was administrated in the study by Zanini et al. [37]. Hydrogen-rich water was administered 2 times per day. The protocol duration of hydrogen-rich water supplementation was 6 months [37].

  • Nicotinamide Riboside. Nicotinamide riboside is a NAD precursor, increasing NAD+ availability with potential implications in preventing age-related mitochondria functional decline and mitochondria bioenergetic capacity in animal models [44]. NR was administrated in the study by Elhassan et al. [34]. The study participants in the intervention group were orally supplemented with 2 capsules of 250 mg of NR (Niagen, ChromaDex, Irvine, CA), administered twice a day for a total of 1000 mg per day. The protocol duration was 21 days [34].

  • Urolithin A. Urolithin A has been proposed to induce mitochondrial gene expression, stimulating mitophagy and improving muscle function in preclinical studies [45, 46]. Urolithin A supplementation was assessed in the study by Liu et al. [38]. The study participants in the intervention group were orally supplemented with 250 mg of softgel containing urolithin A, 4 times per day, for a total of 1000 mg daily. The protocol duration was 4 months [38].

  • Whey protein powder (L-Trp, NA, and NAM). L-Trp, NA, and NAM are involved in the NAD+ metabolisms pathway, functioning as precursors in the biosynthesis process and enhancing NAD+ formation [47]. Therefore, their effects on mitochondria might be related to coenzyme NAD+ and its reduced form NAD(H) that play a role in cellular redox reactions as electron donors in the electron transport chain [48, 49]. In the study by Connell et al. [39], the intervention product was composed of a whey protein powder as a source of tryptophan, to which nicotinic acid (NA) and nicotinamide (NAM) were added. In total, the equivalent of 207.5 mg/day niacin equivalents of NAD+ precursors were supplemented, through L-Trp (210 mg/day at a 60:1 conversion rate), NA (4 mg/day), and NAM (200 mg/day). Subjects in the intervention group were orally supplemented dissolving the powder in 200 mL of orange juice, once a day. The protocol duration was 32 days [39].

All the interventions assessed in the studies included are summarized in Table 1.

Main findings in terms of muscle mitochondrial modifications

The effects of nutraceuticals on muscle mitochondrial modifications were assessed in 6 terms of: (a) mitochondrial oxidative capacity; (b) mitochondrial antioxidant capacity; (c) mitochondrial volume; (d) mitochondrial bioenergetic capacity; (e) mitochondrial transcriptome.

  1. (a)

    Mitochondrial oxidative capacity: Yoshino et al. [36] assessed mitochondrial function by microarray analyses. In particular, the study showed significant improvements (p < 0.05) in respiratory electron transport ATP synthesis, coupling proteins and heat production, respiratory electron transport, oxidative phosphorylation, and tricarboxylic acid cycle and respiratory electron transport after n-3 PUFA therapy compared with baseline [36]. On the other hand, Liu et al. [38] investigated the effect of urolithin A supplementation on the mean change in mitochondrial oxidative phosphorylation capacity assessed by maximal ATP production in the first dorsal interosseous muscle and in the tibialis anterior muscle, without reporting significant differences (p = NS) [38].

    The study by Elhassan et al. [34] assessed the high-resolution respirometry on muscle without reporting significant improvement in skeletal muscle oxidative phosphorylation and maximal respiratory capacity after NR supplementation (p = NS). Moreover, to assess sirtuin-mediated deacetylation activity, the western blot analysis was performed on skeletal muscle lysates without reporting a significant increase in muscle protein acetylation (p = NS) [34].

    Accordingly, the RCT by Connell et al. [39] assessed mitochondrial oxidative capacity by ex vivo high-resolution respirometry on permeabilized muscle fibers obtained from skeletal muscle biopsies. However, no significant differences between groups were underlined in terms of ADP-stimulated (State 3) respiration fueled by complex I linked (malate and octanoyl-carnitine) substrates (p = 0.882) and maximally uncoupled mitochondrial respiration (p = 0.495) [39]. Table 1 reported the results of the studies included in detail.

  2. (b)

    Mitochondrial antioxidant capacity: Rossman et al. [35] assessed mitochondrial-specific ROS-bioactivity showing a significant decrease ( – 35%; P < 0.05) after sodium nitrite supplementation. In contrast, there was no significant improvement after placebo intervention [35].

  3. (c)

    Mitochondrial volume: Rossmann et al. [35] assessed mitochondrial volume by MitoTracker fluorescence without reporting significant differences after sodium nitrite supplementation (p = NS) [35].

  4. (d)

    Mitochondrial bioenergetic capacity: Liu et al. [38] assessed the effect of urolithin A on biomarkers of mitochondrial health and inflammation, reporting a significant reduction in plasma acylcarnitine levels (p < 0.05) and a significant reduction in ceramide levels (p < 0.05) in the experimental group [38]. Elhassan et al. [34] found significant improvement in in the NAM methylation clearance pathways representing the NAD+ metabolome. More in datail, significant differences were underlined in methyl-nicotinamide (MeNAM) (Intervention Group-IG 1.45 pmol/mg vs Control Group-CG 0.35 pmol/mg; p = 0.006), N1-methyl-2-pyridone-5-carboxamide (IG 6.6 pmol/mg vs CG 1.1 pmol/mg; p < 0.001), and N1-methyl-4-pyridone-5-carboxamide (IG 1.6 pmol/mg vs CG 0.3 pmol/mg; p < 0.001) [34].

    Similarly, Connell et al. [39] reported skeletal MeNAM levels were significantly higher in subjects receiving NAD+ precursor supplementation (IG: 0.098 ± 0.063 compared with CG: 0.025 ± 0.014; p = 0.001), suggesting an increased NAD+ metabolism [39].

    Lastly, Zanini et al. [37] reported no significant differences in terms of NAD + /NADH ratio after HRW supplementation [37]. Table 1 shows the mitochondrial bioenergetic capacity outcomes in detail.

  5. (e)

    Mitochondrial transcriptome: Yoshino et al. [36] showed significant improvements in gene expression of UCP3 (~ 30%, p < 0.05) and UQCRC1 (~ 20%, p < 0.05) after n-3 PUFA therapy compared with baseline, suggesting potentials effects in mitochondrial biogenesis and function. In contrast, no significant improvements were reported in PPARGC1A, PPARA, PDHA1, CPT1B, CS, UQCRC2, COX4I1, and COX5B gene expression [36].

On the other hand, in the study of Elhassan et al. [34], the western blot analysis did not highlight significant improvements (p = NS) in the expression of selected mitochondrial proteins in skeletal muscle lysates (CV-ATP5A, CI-NDUFB8, CII-SDHB, CIII-UQCRC2, CIV-MTCO1) [34]. Further details are shown in Table 1.

Figure 2 summarized the statistically significant effects of dietary supplements on mitochondrial modifications.

Fig. 2
figure 2

Dietary supplements effects in mitochondrial modifications

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Quality assessment and risk of bias

According to the Jadad scale [32], all (n = 6, 100%) of the RCTs included [34,35,36,37,38,39] resulted in high-quality studies. Some concerns were present in two studies [36, 37] for the assessors’ blindness. Table 2 showed in detail the score of each subitem of the Jadad scale for the RCTs included.

Table 2 Quality assessment of the studies included in the present systematic review
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The risk of bias was assessed by RoBv2 [33], showing that 4 studies [38, 39, 42, 43] had a low risk of bias, while some concerns emerged for 1 RCTs [41], and 1 [40] showed a high risk of bias. The process showed that all studies (100%) [34,35,36,37,38,39] ensured a correct randomization, while Zanini et al. [37] (25%) showed some concerns in the second domain due to the lack of details on the blinding of study operators and assessors. All studies (n = 6, 100%) showed a low risk of bias in missing outcome data and outcome assessment [34,35,36,37,38,39]. One study (25%) [36] resulted in high risk of bias in selection of the reported results because it selected patients and controls from previous randomization [36]. See Fig. 3 for further details.

Fig. 3
figure 3

Risk of bias of the included studies according to the RoB2

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Discussion

Dietary supplements have been proposed in the last decade as a valid therapeutic option in the multidisciplinary approach to counteract the aging process[13, 14] and, to date, several benefits have been reported after dietary supplements administration on age-related functional and cognitive impairment [23,24,25, 50,51,52].

Although specific mechanisms underpinning cellular aging are far from being fully understood, mitochondria might play a key role in the complex framework regulating the senescence process, being a potential target of precise interventions aiming at slowing down the age-related cellular and subcellular modifications [6, 11, 15,16,17,18,19]. Despite these promising considerations, to the best of our knowledge, no previous systematic review assessed the effects of dietary supplements on mitochondria modifications in healthy older subjects.

In this context, the results of this systematic review of RCTs summarized the different effects of specific dietary supplements and their possible link to specific mitochondrial modifications (see Fig. 2). More in detail, sodium nitrite showed significant results in terms of antioxidant capacity, suggesting intriguing implications for this dietary supplement in targeting oxidative stressors pathways [35]. In this scenario, increased ROS production and concentration are major hallmarks of aging and strictly related to the progressive impairment of mitochondrial oxidative phosphorylation due to constitutive age-related changes in the mitochondrial structure [28, 29]. The supplementation of dietary antioxidants could be useful to counteract the damaging effects of ROS and to protect mitochondrial functions, limiting, for example, the loss of neuronal cells in neurodegenerative disorders [53, 54]. This correlation is due to the fact that mitochondrial functions are directly regulated by mitochondrial DNA. Thus, impairment in mitochondrial DNA due to ROS overproduction could trigger the pathogenesis of age-related pathological conditions, including neurodegenerative and metabolic disorders [55]. However, to date, data about the effects of nutraceuticals supplementation in these conditions are controversial and the reciprocal and multilevel interactions between nutraceuticals, mitochondria, and environmental stressors are far from being fully understood [56].

In addition, recent research is now focusing on specific pathological conditions and there is still a large gap of knowledge on the role of specific dietary supplements in healthy older adults.

On the other hand, it should be noted that our systematic review identified just one study [35] reporting significant improvement in mitochondrial-specific ROS-bioactivity after sodium nitrite supplementation. Therefore, the antioxidant activity of dietary supplements in healthy aging should be further studied to deeply characterize the role of specific nutraceuticals in mitochondria modifications of patients without pathology-induced organelle alterations.

Also the nutrigenomics might play a key role in counteracting the aging process [57, 58]. In this scenario, nutrition can be a key factor to delay the onset of age-related degenerative pathological mechanisms, also from a molecular point of view [59]. In addition, nutrigenomics has been proposed to play a crucial role in aging process and in particular in neuronal health [59]. It has been proposed that neuronal degeneration can be prevented by various foods rich in polyphenols or other bioactive compounds [60, 61]. These mechanisms are related to epigenetic regulation of anti-inflammatory and antioxidant pathways and regulation of gene expression. For example, as recently demonstrated by Uberti et al. [62], the increased ROS levels in the brain during aging (caused by oxidative stress) can be reduced by dietary supplementation with lipoic acid and vitamin D3. The same concept is also valid for suppressed H2O2-induced mitochondrial dissipation [62]. Interestingly, this study reported significant results in terms of mitochondrial transcriptome modifications only after fish oil-derived n-3 PUFA supplementations [36]. In addition, fish oil-derived n-3 PUFA supplementation seems to be effective in increasing mitochondrial oxidative capacity [36]. On the other hand, it should be noted that several studies supported macroscopical effects in terms of anabolic and function enhancement after n-3 PUFA supplementation [63,64,65]. Therefore, these data might partly explain the mechanisms underpinning macroscopical effects of n-3 PUFA supplementations through changes in key gene set pathways involved in muscle structure, growth and oxidative metabolism regulation [36]. Future research might clarify the optimal n-3 PUFA supplementation modality in the bioregulation of molecular pathways involved in the aging process to maximize the n-3 PUFA interaction with mitochondrial oxidative capacity and transcriptome regulation.

In the past few years, several papers suggested a possible association between aging and progressive decline in mitochondria bioenergetic capacity and energy homeostasis [66, 67]. In particular, mitochondrial electron transport chain dysfunction might reduce ATP production, thereby increasing anaerobic metabolism and free radical production [56]. In this scenario, it has been reported that mitochondrial ATP production might be increased using dietary supplements, as well as the removal of toxic metabolites and the exclusion of cellular defects such as deficiency of complex I, II, or III activity in electron transport chain [68]. Therefore, dietary supplements can be useful in the treatment of primary mitochondrial disease: the most commonly used ingredients are antioxidants, such as vitamin C, vitamin E and alpha-lipoic acid; electron donors and acceptors, such as CoQ10 and riboflavin; compounds that can be used as alternative energy sources, such as creatine [69] and compounds that can conjugate or bind mitochondrial toxins, such as carnitine [68]. On the other hand, previous data suggested that few dietary supplements could have a positive role in increasing mitochondria energy production in healthy older adults and significant results are reported only after supplementation with urolithin A [38], NR [34], and whey protein powder [38].

Similarly, the positive effects on mitochondria quality in older adults might be related to physical activity, suggesting a strict link between mitochondrial modifications and the progressive improvement of energy supply induced by exercise-induced conditioning [12]. Unfortunately, evidence from in vivo studies in healthy older adults is limited and few studies assessed the effects of physical activity in this context [66, 67]. Thus, future research might focus on the synergic proprieties between physical activity and dietary supplements to set up tailored multidisciplinary interventions aiming at optimizing the preventive and protective role of lifestyle intervention in the aging processes.

Although dietary supplements are designed only to supplement the diet and not to treat, mitigate or cure diseases [60, 61], several nutraceuticals are frequently prescribed in common clinical practice in wide range of pathological conditions. Thus, growing evidence is now focusing on the efficacy of dietary supplements in several pathological conditions but no previous systematic review focused on healthy older adults.

To date, mitochondria modifications are considered a cornerstone of the aging process and a key target for healthy aging. In the last few years, several dietary supplements have been proposed to treat this condition. Unfortunately, the currently available literature mainly focused on animal models, whereas the effects of dietary supplements on mitochondrial modifications in humans are still far from being deeply characterized. In light of these considerations, it should be noted that the low number of studies included reflect a large gap of knowledge about subcellular effects of dietary supplements in human subjects, although they could benefit from this kind of nutritional intervention. However, the improvement of clinical and research approaches to aging is strictly related to a deeper understanding of the pathophysiological mechanisms underpinning this condition. Therefore, the biological effects of these therapeutic interventions should be better investigated to fulfill the need for a translational approach to implement effective and tailored interventions to treat these subjects. In conclusion, despite macroscopical effects of dietary supplements and nutraceuticals have been previously described, controversies are still open in this field.

In this context, our findings might promote a better understanding of the biological effects of specific dietary supplements and could be considered as a catalyst for the development of precise treatment protocols targeting specific subcellular modifications that play a crucial role in the complex framework underpinning the aging process.

Despite these considerations, we are aware that the present work is not free from limitations. First, the few studies included might represent the main limitation of the present work. However, it should be noted that the strict eligibility criteria allow to provide specific data about the effectiveness of specific mitochondria modifications in healthy older adults. Moreover, the present work underlined a large gap of knowledge in this field, emphasizing the need for good-quality studies assessing the role of dietary supplements in healthy aging to pave the way to a precise prescription of dietary supplements targeting cellular and subcellular pathways involved in the aging process. Second, the large heterogeneity of the dietary supplements assessed is another study limitation. Nevertheless, it should be noted that the present study is the first systematic review of RCTs providing a broad overview on the effects of dietary supplements on mitochondria modifications in older adults. Lastly, a meta-analysis has not been performed due to the heterogeneity of dietary supplements considered, in accordance with the Cochrane Handbook for Systematic Review of Intervention [70].

Conclusion

In the recent years, a growing interest has been rising about the correlation among dietary supplements, mitochondria modifications and age-related functional impairment. However, to date, evidence supporting the role of single dietary supplements in targeting specific mitochondrial modification is still lacking. Taken together, the results of the present systematic review underlined potential effects of sodium nitrite, PUFA, NR, urolithin A and whey protein powder in modulating specific mitochondria modification. On the other hand, the few studies included by the present work severely limit the strength of our results. Thus, further good-quality studies are needed to better characterize the clinical implications of dietary supplements prescription to target specific cellular modifications involved in cellular aging processes.