Journal of Signal Transduction such a checkpoint activation may be attenuated by p66ShcA deletion, which results in decreased intracellular ROS levels. With regard to prodiﬀerentiative eﬀects induced by ROS in this context, in addition to the already cited report by Lee et al. [137], it has been recently demonstrated that in a non skeletal-muscle cell, that is, vascular smooth mus- cle cells (VSMC), ROS increase their diﬀerentiation rate after quiescence through a p38 MAPK-dependent pathway [145]. Similarly, other studies focusing on ROS and muscle metabolism, diﬀerentiation, and growth unravelled some positive interactions with IGF-1 signaling (see below). Again, the most likely explanation for these opposite eﬀects is that cell fate may depend on the intracellular ROS type (i.e., which is the prevailing reactive species) and level. In fact, it is well known that ROS elicit a wide spectrum of cellular responses, depending on their intracellular level [146]. A low dose of ROS controls normal cellular signaling pathways while an intermediate dose results in either tem- porary or permanent growth arrest [147]. Obviously, a high dose of ROS causes cell death via either apoptotic or necrotic mechanisms [142]. 7. ROS and IGF-1 Signaling Growing evidence suggests that oxidative stress is responsi- ble, as a causal or a concausal factor, for the pathogenesis of many muscle diseases and muscle wasting [148, 149]. In muscle cells, IGF-1 is known to promote muscle welfare inducing muscle hypertrophy and stimulate muscle-cell pro- liferation, diﬀerentiation, and survival [123]. IGF-1 has also been found to contribute to oxidative balance and to mediate protective responses against iron-induced-lipid oxidative stress in vivo [150]. Accordingly, Yang and colleagues [151] demonstrated that IGF-1 displayed protective eﬀects on muscle cells after oxidative stress: indeed, pretreatment with IGF-1 protected muscle cells from H2O2-induced cell death and enhanced their survival through promotion of the antiapoptotic protein Bcl2. The same authors showed that protection was via an IGF-1 subpathway: PI3K/Akt and ERK1/2 MAPK pathways [151]. IGF-1 is a peptide hormone with a complex post-tra- scriptional regulation, generating distinct isoforms, namely, IGF-1Ea, IGF-1Eb, and IGF-1Ec (this latter also known as mechano growth factor, MGF) [152]. Mouse models have provided insights into the tissue-speciﬁc functions and responses to ROS of the diﬀerent IGF-1 isoforms [152–155]. For example, in murine models, the local muscle isoform of IGF-1 (mIGF-1, the orthologue of human MGF) has been shown not only to activate proliferation of myoblasts [156], but also to protect cardiomyocytes from oxidative stress via the Sirtuin 1 deacetylase activity [157]. As to physical activity, although its role in regulating the expression of speciﬁc IGF-1 isoforms has been widely studied, data in the literature regarding humans are often contradictory and are aﬀected by many uncontrolled vari- ables such as the lack of dietary control, heterogeneity of subjects, their physical ﬁtness, diﬀerences in proposed physical exercise, and time course of sampling [158–160]. 9 Similarly to other pathways, ROS may regulate either positively or negatively IGF-1 signaling [161]. Low levels of endogenous ROS—due to their reversible oxidative inhi- bition of protein tyrosine phosphatases (see also “ROS as multipurpose local regulators of muscle cell functions”)— induce the phosphorylation on speciﬁc tyrosine residues of insulin receptor (IR) and IR substrates (IRS) protein(s), thus facilitating the IGF-1 signaling. Indeed, the IRβ chain contains multiple sites for the phosphorylation of tyrosine that are sensitive targets of ROS such as H2O2 [162]. By contrast, higher ROS levels inhibit IGF-1 signaling cascades and recent evidence implicates ROS as downregulators of IGF-1 signaling and inducers of insulin resistance and its pathological sequelae [162]. However, ROS may be also involved in the activation of “insulin-like” metabolic eﬀects by activating other non- insulin-initiated signaling pathways: one of the most impor- tant examples is the stimulation of glucose transport in skeletal muscle during exercise [163, 164]. Skeletal muscle contraction stimulates, as well as insulin, glucose transport by up to 50-fold during maximal exercise in humans [165]. Adding exogenous ROS to skeletal muscle in vitro stimulates glucose transport [166] whereas NAC, a potent antioxidant, reduces contraction-mediated glucose uptake by about 50% [167]. This eﬀect of NAC was associated with a similar degree of inhibition of contraction-induced activation of AMPK. This kinase is a fundamental signaling kinase which, besides being involved in mitochondrial biogenesis (see “ROS signaling and myogenic diﬀerentiation”), is also known to upregulate the glucose uptake in muscle under conditions of high AMP/ATP ratio, like hypoxia and muscle contraction, forming a non-insulin-dependent pathway to increase muscle glucose utilization [158, 168–172]. Thus the proposed role of ROS in mediating the stimulation of glucose transport is related to skeletal muscle contraction, that increases superoxide anion production via mitochondrial respiration. Superoxide anion is rapidly converted to H2O2 by SOD, resulting in direct activation of AMPK, Glucose transporter 4 (Glut4) translocation to the plasma membrane, and an increase in glucose transport [173]. Moreover, in muscle cells, NAC antagonized ROS-mediated increase in glucose uptake in response to contraction, but not to insulin. Activation of AMPK in aerobic-exercise-induced glucose uptake is paradigmatic of ROS participation in physiologically-oriented signaling pathways relevant to the homeostasis of the entire organism. It has also been demonstrated that ROS regulate IGF- 1-induced myotube hypertrophy in vitro. It is well known that exercise-induced muscle hypertrophy mostly depends on the increased local production of IGF-1 via activation of the PI3K/Akt pathway [174, 175]; interestingly ROS, which are being overproduced during exercise, contribute in a feedforward manner to stimulate IGF1 net accumulation. Previous reports show that there are positive and negative interactions between ROS and IGF-1 synthesis in both skeletal and VSMCs [176–178]. Treating VSMCs with H2O2 or XO augments both IGF-1 mRNA and IGF-1 protein secreted into the cultured medium, indicating that ROS enhance the IGF-1 autocrine system in VSMCs [176]. By