Clinical presentation of G6PD deficiency from classical drug/infection-induced hemolysis to current cellular effects
There are 400 different biochemical variants of the G6PD enzyme. These can be categorized into 5 classes (I, II, III, IV, and V) depending on the enzymatic activity in erythrocytes and the associated clinical presentation [22]. Class I variants are rare and individuals exhibit less than 10% of normal G6PD activity in their erythrocytes. It is associated with a chronic nonspherocytic hemolytic anemia (CNSHA). Some individuals experience repeated episodes of acute hemolysis and may require transfusion [23]. Class II variants are commonly found in Mediterranean and Asian countries. Similar to class I, individuals with class II variants display no more than 10% of normal G6PD activity in their erythrocytes. Class II variants are not associated with CNSHA. Individuals in this class often suffer from acute hemolysis due to infection as well as exposure to food (fava bean), chemicals (naphthalene mothballs), and certain drugs (antibiotics and antimalarial drugs) [24]. In these severe G6PD variants, extensive intravascular hemolysis can lead to acute kidney failure and acute tubular necrosis [25]. Class III variants can be found in Mediterranean and Asian countries. These moderately deficient individuals display 10-60% of normal G6PD activity in their erythrocytes. Individuals with class III variants have intermittent hemolysis caused by infection and oxidant exposure. Individuals with class IV variants have more than 60% of normal G6PD activity in their erythrocytes and present with milder pathological manifestations. Individuals with class V variants display higher G6PD activity in their erythrocytes compared to normal individuals [26]. These individuals are often asymptomatic and are unaware of having this condition.
Traditionally, G6PD studies have been focused on human red cells. G6PD in nucleated cells regulates cellular processes, including cell proliferation, cell death, autophagy, inflammation, and tumorigenesis. G6PD deficiency reduces replicative potential in human fibroblasts, leading to early-onset senescence [27]. Such premature senescence is most likely due to elevated oxidative stress rather than increased telomere shortening. Approaches using biochemical inhibitors or RNAi knockdown against G6PD in several cell lines demonstrate that decreased G6PD activity is associated with growth retardation [28]. The most common form of cell death caused by G6PD activity suppression is apoptosis.
The nitric oxide (NO) donor, sodium nitroprusside (SNP) at 50 µM, stimulates growth in human foreskin fibroblasts, whereas, at the same concentration, SNP causes apoptosis in G6PD-deficient foreskin cells [29]. Diamide is a GSH-depleting oxidant. Impaired GSH regeneration, membrane peroxidation, and abnormal aggregation of membrane-associated cytoskeletal proteins are found in diamide-treated G6PD-knockdown HepG2 cells. While diamide-induced oxidative damage may result in necrosis in G6PD-knockdown HepG2 cells, the antioxidant N-acetylcysteine (NAC) ameliorates diamide-induced cell death and oxidative stress [30]. Similarly, G6PD-knockdown HepG2 cells are highly susceptible to hydrogen peroxide-induced growth inhibition and apoptosis, whereas NAC reverses these impairments [31].
Redox homeostasis mediated by G6PD is implicated in the modulation of the immune response and inflammation. G6PD deficiency is correlated with an increased risk of neonatal sepsis [32–34]. Infants and trauma patients with G6PD deficiency display altered cytokine profiles [35–37]. Glucose overload-induced vascular inflammation in human aortic smooth muscle cells reveals that IL-1β enhances glucose transport and metabolism through the PPP, resulting in an increased pro-inflammatory response, including NF-κB and NOX activation and iNOS protein expression [38]. Blockade of G6PD with either the chemical inhibitor 6-aminonicotinamide, 6-AN, or siRNA against G6PD abolishes the pro-inflammatory response. G6PD deficiency can elevate inflammation through NF-κB-mediated pro-inflammatory cytokine upregulation. An in vitro HepG2 cell model of lipid-induced chronic hepatic inflammation indicates that G6PD knockdown enhances a pro-inflammatory cytokine response and ROS production [39]. Treatment with the anti-oxidative enzyme glutathione peroxidase or the anti-inflammatory agent curcumin in HepG2 cells inhibits the secretion and expression of the pro-inflammatory cytokine IL-8. These findings suggest that G6PD modulates the pro-inflammatory response in an induced and cell-dependent manner.
G6PD plays a role in the modulation of the inflammatory response in several immune cells. Peripheral mononuclear cells from G6PD-deficient individuals produce lower levels of the pro-inflammatory cytokines, IL-6, and IL-1β, compared with normal individuals [40]. G6PD-deficient granulocytes display a reduced respiratory burst resulting in diminished bactericidal activity and an increased susceptibility to infection [41,42]. G6PD gene and protein expression are increased in macrophages by free fatty acids and lipopolysaccharides (LPS) [43]. Upregulation of the macrophage G6PD gene in adipose tissue of obese mice is associated with increased levels of proinflammatory cytokines, including IL-6, IL-1β, and MCP-1. The prooxidative genes, including NOXs and iNOS, are also increased when accompanied by increased G6PD gene and protein expression. The increased pro-inflammatory cytokines and pro-oxidative genes are downregulated when the NF-κB and MAPK pathways are suppressed as well as if macrophage G6PD is reduced by chemical inhibitors (6-AN, DHEA) or siRNA [43].