The primary physiological role of the HO

The primary physiological role of the HO system is the protection of CGRP (rat) from oxidative stress. These cytoprotective effects are attributable both to the degradation of the pro-oxidant heme and to the positive contribution of the metabolites produced during this degradation, namely CO, BV, and BR. In particular, CO is an important gaseous transmitter that mediates vasodilatory, anti-apoptotic and anti-inflammatory effects [6]. The primary endogenous source of CO (nearly 80%) derives from oxidative degradation of heme by the HO system, while the remaining 20% is produced in conditions of high stress, through lipid peroxidation reactions of membrane phospholipids, oxidation of phenols, flavonoids, and halomethanes, and photo-oxidation of many other organic compounds [6]. Owing to its gaseous nature, CO can easily migrate to the extracellular compartment, where it acts as a messenger in numerous signaling pathways. The main effects include: i) anti-proliferative and vasorelaxant activities by activation of sGC; ii) ion channel modulation, such as K+, Ca2+-dependent channels; iii) anti-inflammatory and anti-apoptotic functions through the modulation of MAPK [6,7]. Heme catabolism also leads to the formation of free iron, a well-known pro-oxidant agent. Iron is an essential cofactor of numerous cellular enzymes and redox-dependent proteins, but an excess of its free form is cytotoxic and promotes the production of ROS, according to the well-known reactions of Fenton and Haber-Weiss. To avoid the toxicity caused by the production of radicals, the iron is rapidly removed thanks to the iron transporter (Fe-ATPase). Also, HO-1 induces the synthesis of ferritin, a globular protein which sequesters the iron produced by the heme degradation, transfers it into the endothelial cells, and renders the metal available for the endogenous biosynthesis of heme [8]. Finally, BV is oxidized by bilirubin reductase into BR, a powerful antioxidant, as it acts as a scavenger of ROS and RNS [9]. Several works have shown the beneficial effects of BR. Studies carried out using animal models have shown that BR perfusion induces a reduction in infarct damage and cardiac ischemia, while clinical data have shown that high levels of plasma BR reduce the risk of cardiovascular diseases [10]. Moreover, low concentrations of BR have been used in the treatment of peripheral vascular diseases, myocardial ischemia, congestive heart collapse, and sepsis [[11], [12], [13]]. Conversely, defective BR excretion, especially in newborns, and blood accumulation, are responsible for neonatal jaundice and potential neurological damage [14]. Besides its enzymatic role, HO-1 behaves as a signal molecule. Generally, HO-1 exerts its catalytic activity in the endoplasmic reticulum but HO-1, in shorten and inert form, can migrate into different subcellular compartments, e.g., nucleus and mitochondria. Its ability to translocate to the nucleus is due to a C-terminal cleavage, which is a region crucial for anchoring to the smooth endoplasmic reticulum membrane [15]. Through this migration, HO-1 can modulate gene expression, and translation of several proteins, including HO-2, cytochromes, transcription factors, such as Nrf2 and STAT3 and also can activate DNA repair agents [15]. Moreover, HO-1 regulates itself expression through prevention of Nrf2 phosphorylation and dependent proteolysis, mediated by ubiquitin. Consequently, Nrf2 accumulates in the nucleus and, in turn, regulates HO-1 expression [16]. The modulation of HO-1 expression is regulated by different transcription factors, including Nrf-2, one of the most important. Nrf-2 represents a key player in response to oxidative stress, implicated in the inducible expression of about 200 antioxidant genes, including HO-1 [17]. Nrf2/HO-1 axis contributes to the maintenance of cellular homeostasis, acting a physiological role in response to stress, inflammation, immune and vascular modulation [1]. Other transcription factors involved in HO-1 up-regulation include AP-1 [18], NF-κB [19] and different kinases, such as PKC [20]. Conversely, HO-1 expression is down-regulated by a variety of factors, such as angiotensin II and interferon-γ in the vascular system and glioblastoma cells, respectively, and TNF, as shown in peripheral monocytes [21]. The major physiological repressor of HO-1 expression is Bach-1, a transcription regulator protein, able to bind the HO-1 promoter site. Notably, Bach-1 binds MARE [22], where Nrf-2 modulates HO-1 gene and inhibits HO-1 expression under physiological conditions. In response to stressful stimuli, Bach-1 leaves the nucleus allowing Nrf-2 binding to DNA, where it acts as an inducer of this cytoprotective heme enzyme [23].