**1. Introduction**

Iron is the most abundant transition metal in living organisms and is involved in multiple biochemical processes including oxygen binding and transport, energy production, regulation of cell growth and di fferentiation, and a variety of enzyme reactions. Most body iron is incorporated into haem proteins (e.g., haemoglobin, myoglobin, cytochromes, and haem thiolates). Non-haem iron

serves instead as an enzyme cofactor (i.e., atomic iron) or iron reserve (e.g., bound to cytosolic ferritin and haemosiderin) and is integral to electron transport chain complexes (i.e., iron-sulphur clusters) and transferrin (Tf) [1,2]. Approximately 5% of cellular iron exists as chelatable non-haem iron, referred to as labile iron pool. This iron fraction consists of both ferrous (Fe<sup>2</sup>+) and ferric (Fe<sup>3</sup>+) ions associated with a variety of small molecules, including organic anions, polypeptides, and phospholipids. Fe2<sup>+</sup> ions can participate in Fenton reactions thereby producing highly destructive radicals, which are thought to be major contributors to the generation of protein and DNA oxidative adducts [3–5]. Hence, a tight coordination encompassing iron absorption, uptake, e fflux, and sequestration is crucial to preserve cell homeostasis.

Circulating iron is bound and transported by Tf. However, in the setting of iron overload, the iron-binding capacity of plasma Tf can be exceeded and accumulation of non-Tf-bound iron (NTBI) occurs [6]. As such, NTBI needs to be adequately disposed. Fourteen divalent metal transporters belonging to the Zrt-Irt-like protein (ZIP) family, named ZIP1 to ZIP 14, have been identified [7]. Of them, ZIP8 and ZIP14 have similar amino acid sequences [8] and contribute to the import of several divalent ions, including iron [9]. In particular, Zip14 mediates, at least in part, NTBI uptake by hepatocytes in the context of iron overload [10].

Iron metabolism is modulated by the defensin-like hormone hepcidin [11] via binding and subsequently degrading of the iron exporter ferroportin at the level of key iron sources [i.e., duodenal enterocytes (absorption of dietary iron), splenic and hepatic macrophages (recycling iron from erythrophagocytosis), and hepatocytes (iron stores)] [12]. In particular, circulating iron concentrations decrease as a consequence of intestinal absorption and release of iron from recycling macrophages [11].

Skeletal muscle is a major reservoir of body iron, which is comprised by 60% of non-haem fraction [13]. Studies have shown that non-haem iron accumulates in muscle during ageing possibly causing oxidative damage to biomolecules and organelles, including mitochondria [14–18]. As such, iron dyshomeostasis is advocated as a mechanism involved in the pathogenesis of sarcopaenia of ageing and disuse-induced muscle atrophy [19].

Along with iron imbalance and mitochondrial dysfunction, chronic inflammation is a hallmark of ageing and a factor involved in functional decline [20,21]. A link between mitochondrial damage and chronic low-grade inflammation has recently been hypothesised [21,22]. However, little is known about the relationship among iron dyshomeostasis, inflamm-ageing, mitochondrial dysfunction, and physical performance in older adults. To provide an initial appraisal of the subject, the present study was undertaken to assess total iron content, the expression of selected iron transporters, and indexes of mitochondrial damage in muscle biopsies obtained from healthy young adults and older people with varying levels of physical performance. The relationship between muscle iron content and systemic inflammation was also explored.

#### **2. Materials and Methods**
