Signaling Integration of Hydrogen Sulfide and Iron on Cellular Functions

Abstract

Significance:

Hydrogen sulfide (H₂S) is an endogenous signaling molecule, regulating numerous physiological functions from vasorelaxation to neuromodulation. Iron is a well-known bioactive metal ion, being the central component of hemoglobin for oxygen transportation and participating in biomolecule degradation, redox balance, and enzymatic actions. The interplay between H₂S and iron metabolisms and functions impacts significantly on the fate and wellness of different types of cells.

Recent Advances:

Iron level in vivo affects the production of H₂S via nonenzymatic reactions. On the contrary, H₂S quenches excessive iron inside the cells and regulates the redox status of iron.

Critical Issues:

Abnormal metabolisms of both iron and H₂S are associated with various conditions and diseases such as iron overload, anemia, oxidative stress, and cardiovascular and neurodegenerative diseases. The molecular mechanisms for the interactions between H₂S and iron are unsettled yet. Here we review signaling links of the production, metabolism, and their respective and integrative functions of H₂S and iron in normalcy and diseases.

Future Directions:

Physiological and pathophysiological importance of H₂S and iron as well as their therapeutic applications should be evaluated jointly, not separately. Future investigation should expand from iron-rich cells and tissues to the others, in which H₂S and iron interaction has not received due attention. Antioxid. Redox Signal. 36, 275–293.

Introduction

Iron and hydrogen sulfide (H₂S) are important chemical species that lie on the opposite ends of the redox spectrum. Where excess of iron creates oxidative stress, H₂S offers antioxidant protection. The metabolisms of H₂S and iron in mammalian cells are also intertwined. Through a nonenzymatic process, iron participates in H₂S production. On the contrary, H₂S regulates iron uptake, transport, and accumulation. The interaction between H₂S and iron affects cellular homeostasis and constitutes important signaling cross talk. Although with great efforts, our understanding of the intertwined metabolisms of H₂S and iron in mammalian cells and their respective and integrated roles in redox signaling has been limited.

This review article aims at identifying the relevant knowledge gaps in iron and H₂S biology. Multiple mechanisms are presented to interpret the reported metabolisms of H₂S and iron, respectively, and their impacts to each other under different physiological and pathophysiological conditions. We further reviewed differential impacts of iron and H₂S on cellular redox balance, considering one being a prooxidant and the other being an antioxidant. Finally, the functional consequences of the interaction between H₂S and iron are discussed, focusing on the cardiovascular system, the liver, and the brain.

H₂S Metabolism

H₂S is produced in eukaryotes, enzymatically from three main enzymes: cystathionine γ-lyase (CSE; EC 4.2.1.22), cystathionine β-synthase (CBS; EC 4.4.1.1), and 3-mercaptopyruvate sulfurtransferase (MST; EC 2.8.1.2). Where CSE and CBS can use both l-cysteine and homocysteine as the substrates, MST only uses 3-mercaptopyruvate (3-MP) as a substrate, which is a derivative of l-cysteine (176). Methionine is converted to homocysteine in one carbon metabolism with S-adenosylmethionine (SAM) and S-adenosyl-homocysteine (SAH) as intermediate products (72, 128). Entering the reverse-transsulfuration pathway, homocysteine converts to cystathionine catalyzed by CBS (154). Cystathionine is converted to l-cysteine using CSE. H₂S is released from l-cysteine in the presence of CSE or CBS (176). Similarly, cysteine can be converted to 3-MP using cysteine aminotransferase (CAT). MST catalyzes the conversion of 3-MP to H₂S (73). CBS is the main enzyme for H₂S production in the brain, whereas CSE is the main enzyme in cardiovascular and liver systems (21, 196). MST is mainly found in the heart and endothelium in both the cytoplasm and mitochondria (124, 146). Our laboratory has recently shown that MST is the main enzyme for H₂S production in chicken red blood cells (RBCs) (63). Figure 1 summarizes enzymatic and nonenzymatic H₂S production pathways in eukaryotes.

FIG. 1.

Summary of important enzymatic and nonenzymatic H₂S generation pathways. CBS and CSE can act on L-homocysteine to convert it to L-cystathionine releasing H₂S. CSE further converts L-cystathionine to l-cysteine and subsequently pyruvate and ammonia, along with H₂S. Cysteine can be converted to 3-MP by CAT, which can be further broken down to release H₂S by MST. Nonenzymatic H₂S production is conducted by reduction of elemental or bound sulfur, or reaction of ferric ions with l-cysteine in the presence of vitamin B6. 3-MP, 3-mercaptopyruvate; CAT, cysteine aminotransferase; CBS, cystathionine β-synthase; CSE, cystathionine γ-lyase; H₂S, hydrogen sulfide; MST, 3-mercaptopyruvate sulfurtransferase.

Nonenzymatic sources of H₂S in eukaryotes include cysteine reaction with iron and vitamin B6 (200) or reduction of elemental sulfur in which protein-bound sulfur or sulfur containing compounds releases H₂S (74, 98, 142). Vitamin B6 initiates a nucleophilic attack on cysteine, forming a cysteine-aldimine. Furthermore, free or heme-bound iron reacts with the cysteine-aldimine to form a cysteine-quinonoid. Ferric ions react with cysteine-quinonoid to eliminate the thiol group as H₂S and the resulting desulfurated aldimine undergo hydrolysis to yield vitamin B6, ammonia, and pyruvate (Fig. 2A). A classic example of nonenzymatic H₂S production is the reduction of elemental sulfur using NADPH (98). Searcy and Lee showed that RBC lysates can react with reduced glutathione (GSH), NADH, and NADPH to form H₂S nonenzymatically (142). They further showed that H₂S may be formed through inorganic sulfur using reducing equivalents from glucose oxidation in the phosphogluconate pathway in RBCs [Equation (1) in Fig. 2B]. The phosphogluconate pathway represents 5%–10% of glucose metabolism in RBCs but becomes more active in oxidative stress (142). Searcy and Lee predicted that 2 glucose molecules reacting with 6 elemental sulfur and 3 water molecules yield 3 lactate, 6 hydrogen sulfide (H₂S), and 3 carbon dioxide molecules [Equation (1) in Fig. 2B]. However, their empirical data showed a ratio of 6 H₂S to 2 carbon dioxide. They theorized this with the release of pyruvate instead of lactate but did not give a balanced reaction. A balanced equation would be achieved with 4 molecules of sedoheptulose [Equation (2) in Fig. 2B], a metabolite in the phosphogluconate pathway that coincidentally is also increased in oxidative stress (29, 192). This phenomenon also hints an antioxidative role of H₂S. Similarly, thiosulfates can undergo reduction by interacting with GSH to release H₂S (Fig. 2C). Leskova et al. showed this phenomenon in human umbilical vein endothelial cells (86).

FIG. 2.

Reactions leading to nonenzymatic H₂S formation in cells. Cysteine reacts with P5P to form a cysteine/aldimine complex, which reacts with Fe³⁺ to form cysteine-quinonoid. H₂S is released from cysteine-quinonoid via elimination, resulting in aldimine-pyridoxal that further dissociates back into P5P, pyruvate, and ammonia (A). Glucose or its reduced equivalent sedoheptulose reacts with elemental sulfur to form lactate or pyruvate, H₂S, and CO₂ (B). Equation (2) in (B) is speculative based on literature review. Thiosulfate reacts with glutathione (reduced) to form glutathione (oxidized), sulfite, and H₂S (C). GSSG, glutathione oxidized; P5P, pyridoxal 5 phosphate.

Where enzymatic production of H₂S is part of a complex series of reactions in cysteine metabolism and the expression of H₂S-producing enzymes can be induced or inhibited, nonenzymatic production of H₂S is less regulated via nonspecific reactions. There would be no difference in the biological functions of H₂S whether it is produced enzymatically or nonenzymatically as it is the same molecule. The functional differences, if any, may be accounted for by the differences in the concentrations of H₂S produced and the coproducts from different pathways. Nonenzymatically produced H₂S may contribute to basal levels of H₂S in the blood stream (200) where H₂S-producing enzymes are less expressed. Moreover, iron-dependent nonenzymatic production of H₂S may offer a protection mechanism against iron-induced oxidative stress. This mechanism is discussed in detail later in this article.

H₂S can be exhaled or passes through urine or feces as sulfate or thiosulfate (56, 177). H₂S can also be oxidized in mitochondria to thiosulfate and further to sulfite or sulfate catalyzed by sulfate detoxifying enzymes, such as rhodanese (20, 126). Methylation of H₂S to a nontoxic compound, dimethylsulfide, by thiol S-methyltransferase is another possible fate of H₂S (181). Lastly, H₂S can be scavenged by different proteins containing metallic groups or disulfide molecules, such as methemoglobin, which is a hemoglobin derivative with oxidized ferric irons (Fe³⁺) (165). The neutral H₂S molecule rapidly binds with Fe³⁺ in methemoglobin in vitro and in vivo with an association constant of 3.5 × 10³ M ⁻¹·s⁻¹. This association is pH independent, but its dissociation is pH dependent. The dissociation of H₂S increases with an increased acidity and the dissociation curve shifts to the right with an increase in pH (26). This is similar to Bohr's effect for oxygen, which governs loading and unloading of oxygen from hemoglobin depending on partial pressure of CO₂ and local pH. RBCs may unload oxygen and H₂S to tissues with increased metabolic activity to enhance local oxygen and increase vasodilation (59). Figure 3 summarizes transport of H₂S via methemoglobin. Other molecules, which may bind H₂S, include oxidized glutathione, catalase, and horseradish peroxidase (21, 27).

FIG. 3.

Transport of oxygen via hemoglobin (Fe²⁺) and H₂S via methemoglobin (Fe³⁺) in RBCs to metabolically active tissues. RBC, red blood cell. Color images are available online.

H₂S moves intracellularly by free diffusion. It can also diffuse out of the cell for paracrine signaling. Bearden et al. found the presence of extracellular H₂S, produced by CSE and CBS in the blood as part of plasma proteome (9). Moreover, H₂S can bind to oxidized, ferric hemoglobin (methemoglobin) and be transported in the circulation as a potential endocrine signaling molecule (59).

Iron Metabolism

Human body contains 2.5–4.0 g of iron, predominantly present in hemoglobin and in lesser quantities in myoglobin, and in various enzymes and transport proteins (16). Iron is involved in a wide array of physiological functions, such as erythropoiesis, oxygen transport, and degradation of lipids, proteins, and nucleic acids. It is also used in myocardial and muscle metabolism, function of thyroid gland, central nervous system, and immune system (57). Iron exists in multiple states and forms in vivo, including Fe³⁺ (ferric), Fe²⁺ (ferrous), bound iron in the heme group, or in iron/sulfur (Fe-S) clusters such as 2Fe2S or 4Fe4S (83, 172). Each form is tightly regulated by complex mechanisms. The most abundant form of iron is the one bound to heme, followed by transferrin and ferritin (1). Free ionic forms of iron are kept at a very low concentration to avoid toxicity by free radicals (32). Transferrin levels in plasma are sufficient to bind more than 99% of iron, which leaves the rare possibility of free iron. Any free iron is usually loosely complexed with molecules such as citrate or serum albumin. However, in diseases such as atransferrinemia, hemochromatosis, and thalassemia, plasma free iron levels overwhelm the transferrin binding capacity (131).

Free iron enters the cell as Fe²⁺ through ion channels or transporters such as divalent metal transporter 1 (DMT1) and ZRT/IRT-like protein 14 (ZIP14). Whereas DMT1 chiefly prefers and transports iron in the cell, it can also transport copper, zinc, and manganese. DMT1-deficient mice showed decreased iron concentrations in blood, liver, spleen, and heart, but not other metal ions (144). ZIP14 is a zinc transporter, but it transports iron as well (71). Moreover, iron may enter through L-type Ca²⁺ channels as well (45, 96, 119). Ionic Fe³⁺ gets reduced to Fe²⁺ by ferrireductases such as six-transmembrane epithelial antigen of prostate 2 (STEAP2), stromal cell derived receptor 2 (SDR-2), and duodenal cytochrome B (Dcytb) outside the cell (15). Dietary heme enters the enterocyte by heme carrier protein 1 (HCP1) transporter protein (145). Transferrin-bound iron binds to a transferrin receptor 1 (TfR1) and is internalized as an endosome. Furthermore, the iron is released from transferrin as Fe³⁺ and reduced to Fe²⁺ by the neighboring STEAP2 ferrireductase in the endosome before being transported into the cytoplasm through DMT1 (6). Free iron as Fe²⁺ constitutes the labile iron pool (LIP), which may increase through breakdown of heme by heme oxygenase (HO) (188). Furthermore, Fe²⁺ can bind to apoferritin to become ferritin and be stored as Fe³⁺ (6). Free iron inside the cell may be incorporated as a cofactor for iron-containing proteins involved in processes such as reactive oxygen species (ROS) generation and degradation of biomolecules (57, 81). Free iron can be exported out of the cell via ferroportin (Fpn) transporter along with ferroxidase enzymes, hephaestin, and ceruloplasmin. Ceruloplasmin converts Fe²⁺ to Fe³⁺ outside the cell (15). Most of the iron requirements for cellular metabolism and function are fulfilled by recycling existing iron. An example of this process is heme recycling by macrophages, which engulf senescent RBCs and break them down into free iron or heme, which is moved into the cytoplasm via natural resistance-associated macrophage protein (NRAMP1) or heme responsive gene 1 (HRG1) transporter (76, 201). The heme can be broken down in the cytoplasm by HO or exported out of the cell via feline leukemia virus subgroup C receptor-related protein 1a (FLVCR1a) (201).

H₂S can interact with a wide variety of ion channels (177). For example, H₂S is a known K_ATP channel opener through which it mediates vasorelaxation (61). One possible interaction between H₂S and iron is that H₂S can inhibit L-type Ca²⁺ channels by S-sulfhydration as observed in mouse pancreatic beta cells (159). Iron can enter the cell through L-type Ca²⁺ channels as observed in cardiomyocytes (118). Inhibition by H₂S may decrease the amount of iron entering in the cell.

Total iron concentration inside the body is largely controlled through the protein hepcidin. It is secreted by the liver into circulation when plasma iron concentration is high (Fig. 4). Hepcidin blocks Fpn by binding to it, causing its ubiquitination followed by internalization and degradation (134). This results in the blockade of iron movement into circulation from enterocytes, macrophages, or hepatocytes (43, 113). If there is an abnormal increase in hepcidin in circulation, such as in the case of inflammation, a higher level of hepcidin may lead to iron deficiency (113). Conversely, abnormally low hepcidin, as in the case of certain hereditary hemochromatosis (HH), leads to iron overload (91, 114). Human body loses 1–2 mg of iron daily by skin and enteric desquamation or through blood loss (171).

FIG. 4.

Uptake, transportation, and recycling of iron. Heme enters enterocytes after binding with the HCP1 receptor and releases iron via HO. DMT1, ZIP14, or L-type Ca²⁺ channels provide pathways for Fe²⁺ entering enterocytes. Fe³⁺ first gets reduced to Fe²⁺ by ferrireductase such as STEAP2/SDR-2/Dcytb or CP. Transferrin-bound iron is internalized via binding to TfR1 and endocytosis after which the iron in Fe³⁺ form is released and converted to Fe²⁺ by ferrireductase such as STEAP2 and transported to the cytoplasm via Fe²⁺ channels, such as DMT1 or ZIP14. Free iron inside the cell is in Fe²⁺ form, which constitutes LIP. Heme can be degraded to release Fe²⁺ via HO. Heme is shuttled among intracellular organelles via (i) transportation to the cytoplasm from mitochondria via FLVCR1b receptors; and (ii) moving to the cytoplasm via HRG1 transporter. Iron in the form of Fe²⁺ can be moved from intracellular organelles to cytoplasm through NRAMP1 transporter and moved out of the cell via Fpn, which is regulated by hepcidin secreted by the liver. Hepcidin blocks Fpn, leading to reduced release of iron from enterocytes or other types of cells. CP, ceruloplasmin; Dcytb, duodenal cytochrome B; DMT1, divalent metal transporter 1; FLVCR, feline leukemia virus subgroup C receptor-related protein; Fpn, ferroportin; HCP1, heme carrier protein 1; HO, heme oxygenase; HRG1, heme responsive gene 1; LIP, labile iron pool; NRAMP1, natural resistance-associated macrophage protein; SDR-2, stromal cell-derived receptor 2; STEAP2, six-transmembrane epithelial antigen of prostate 2; TfR, transferrin receptor; ZIP14, ZRT/IRT-like protein 14. Color images are available online.

Iron status in vivo can be determined by different parameters, such as hemoglobin concentration, serum iron, serum ferritin, tissue ferritin, liver iron concentration, and nontransferrin-bound iron (NTBI) levels in the blood or tissues. Each parameter changes with respect to different physiological conditions and can be used as an indicator for disease. However, careful considerations need to be made before making a conclusion based on iron values as they tend to be different between different age groups, gender, and disease models (18, 38, 44, 125, 162). Table 1 summarizes total iron concentration (free and bound irons) in different biological samples.

Table 1.

The Concentrations of Total Iron and Selective Iron Proteins in Different Organs

Aortic tissue iron concentration	Human	<300 mg iron/g dry tissue weight (140)
Aortic tissue iron concentration	Mouse (C57BL/6J)	<200 mg iron/g dry tissue weight (140)
Brain iron concentration	Human	28–122 mg iron/kg dry tissue weight (78)
	Mouse (C57BL/6J)	18.3–19.8 mg iron/kg dry tissue weight (52)
Liver iron concentration	Human	0.2–2 mg iron/g dry tissue weight (153)
	Mouse (C57BL/6J)	0.181 mg iron/g dry tissue weight (189)
Heart iron concentration	Human	0.09–78.64 mg iron/g dry tissue weight (24)
	Mouse (C57BL/6J)	0.513 mg iron/g dry tissue weight (189)
Plasma iron concentration	Human	10–30 μM (42)
Mouse (C57BL/6J)	17–23 μM ^a (27)
Serum ferritin concentration	Human	65–670 pM (111)
Mouse (C57BL/6J)	241 pM ^b (106)
Serum iron concentration	Human	9–32 μM (111)
Serum iron concentration	Mouse (C57BL/6J)	19–21 μM ^a (101)
Serum transferrin concentration	Human	2–3.6 g/L (111)
Serum transferrin concentration	Mouse (C57BL/6J)	2.6–3.2 g/L (88)

SI units were converted.

One milligram of ferritin is equal to 8 mg of stored iron (111).

Whereas hepcidin controls overall iron in the body, intracellular regulation of iron level depends on iron response elements (IRE) and iron response element binding proteins (IRE-BP). IREs are short sequences on untranslated regions (UTR) of messenger RNA (mRNA), which can bind to IRE-BP (83). For example, mRNA of ferritin contains an IRE near 5′ UTR and mRNA of TfR1 has multiple IREs near 3′ UTR. If the cellular iron content is low, IRE-BP becomes active and binds to IRE of ferritin mRNA to destabilize it, resulting in decreased storage and more free form of iron. Conversely, IRE-BP binding to IRE of TfR1 mRNA stabilizes it, resulting in increased expression of TfR1 and increased intake of iron. With high cellular iron concentrations, IRE-BP becomes inactive and results in the exact opposite effect on ferritin and TfR1 (83, 108).

Iron and H₂S in Redox Signaling

Oxidative stress is the accumulation of prooxidants, which overwhelms the body's natural antioxidant systems (50). The word “oxidative stress” has a frightening connotation, but events of oxidative stress also occur at a homeostatic level, which is essential for redox signaling, known as oxidative eustress (149). Redox signaling encompasses cellular processes involving reactions where electron transfer occurs through free radicals, redox-active metals, or related species. Overproduction of ROS and reactive nitrogen species in oxidative stress leads to widespread damage of biomolecules such as proteins, lipids, and nucleic acids (104). The reactive species of note include the superoxide (O₂ ^•), hydroxyl (^•OH), alkoxyl (RO^•), peroxyl (ROO^•), and nitroxyl radical (NO^•), along with hydrogen peroxide (H₂O₂), organic hydroperoxides (ROOH), and hypochlorous acid (HOCl) (152). The most notable source of ROS includes mitochondria (mitoROS), where the electron transport chain may generate O₂ ^•, ^•OH, and H₂O₂ (39). Complex I releases O₂ ^• when molecular O₂ reacts with reduced FMN (flavin mononucleotide) when there is a high NADH/NAD⁺ ratio in the matrix. Complex III produces O₂ ^• in the presence of respiratory inhibitors such as antimycin; and complex II produces O₂ ^• when complex I and II are inhibited (39). Other sources of mitoROS include enzymes such as the α-ketoglutarate dehydrogenase complex and pyruvate dehydrogenase complex (97, 155). Cytoplasmic production of ROS (cytoROS) includes ROS generation through the NADPH oxidase (NOX) family and nitric oxide synthases (eNOS, iNOS, and nNOS) (39).

Superoxide and H₂O₂ generated either in cytoplasm or mitochondria are freely membrane permeable and can react with iron to form the highly reactive and toxic hydroxyl radical through Fenton reaction (110). Under normal metabolic conditions, superoxide leads to the more favorable oxidation of Fe²⁺ to Fe³⁺ along with ROS generation. If superoxide levels are high, however, the less favorable reduction of Fe³⁺ to Fe²⁺ along with ROS generation takes place (25, 104). The source of the free Fe²⁺ or Fe³⁺ is the LIP in cytoplasm (cLIP) and mitochondria (mLIP) (17). Iron metabolism and formation of LIP are discussed in the previous section and summarized in Figure 4. There are several conditions that can lead to iron overload. HH is an autosomal recessive disorder that leads to increased absorption of dietary iron. There are four types of HH: type 1 due to a fault in HFE gene required for iron sensing; type 2 due to a fault in hemojuvelin, a regulator of hepcidin or hepcidin itself; type 3 due to a fault in TfR2; and type 4 due to a fault in Fpn (127). Downregulation of hepcidin is common in HH, which leads to iron overload (182). The increased iron is accumulated in parenchyma of organs such as the liver and heart and is responsible for widespread organ damage due to oxidative stress (182). Similar to HH, iron overload is also observed in diseases where repeated blood transfusion is required, such as thalassemia (49), sickle cell anemia (49), chronic kidney disease (133), liver disease (107), and certain cancer types such as colon-rectal (164) and liver cancer (54). Iron overload-catalyzed ROS can induce generalized damage to biomolecules such as carbohydrates, lipids, proteins, and membranous structures. A secondary effect of iron catalyzed-ROS is the induction of stress and inflammation signaling pathways such as Janus kinase/signal transducer and activator of transcription (JAK/STAT), nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB), and p38/mitogen-activated protein kinase (p38/MAPK) (19, 26, 33, 151).

Oxidative stress can be countered through enzymatic and nonenzymatic antioxidant mechanisms (14, 132). The enzymatic antioxidants include the enzymes superoxide dismutase (SOD), catalase, glutathione peroxidase, and HO, and redox proteins such as glutaredoxins, thioredoxins, and peroxiredoxins. The nonenzymatic antioxidants include low-molecular-weight compounds such as GSH, NADPH, and vitamin C and E along with trace metals such as selenium (14, 132). Any O₂ ^• that is converted to H₂O₂ by SOD can be reduced to water and oxygen by catalase or water only by glutathione peroxidase utilizing GSH molecules (183). Collectively, SOD, catalase, and glutathione peroxidase constitute first-line antioxidant defense, whereas other enzymes and molecules such as glutathione, ascorbic acid, and vitamins are considered second line of defense (55).

Another species critical to redox signaling are reactive sulfur species (RSS). Although sulfur is part of antioxidant mechanisms, as explained in subsequent paragraphs, its role in “reductive stress” has been recently brought forward (116). Sulfur has lower electronegativity than oxygen, which is why it can exist in more oxidation states. RSS can exist as hydrogen persulfide or polysulfide (H₂S₂ or H₂S_n); protein sulfide or polysulfide [PSH, PS(S)_nH, PS(S)_nP]; small-weight polysulfides [RS(S)_nH or RS(S)_nR]; sulfenic acids or nitrosothiols (RSOH or RSNO) (75). RSS generation involves a loss of electron from 2 H₂S molecules to form 2 HS^•, which can recombine to form persulfides or polysulfides (116). Whether the formation of persulfide can be categorized as a redox process remains to be determined. Recently, Akaike et al. have shown the strong potency of cysteinyl-tRNA (transfer RNA) synthetases in forming cysteine polysulfides in mitochondria and cytoplasm during redox signaling (3).

The key difference in ROS and RSS is their ability to be preserved. When peroxide (ROS) attacks a sulfhydryl group of cysteine in a protein, the peroxide is reduced to oxygen and water. When persulfide attacks the sulfhydryl group, however, the persulfide can be reversed by thiol exchange between the protein-RSS and a low-molecular-weight thiol group as carrier. Consequently, the original persulfide is preserved (116). Coincidentally, oxidation of H₂S produces a stronger antioxidant, that is, persulfide, which, if unchecked, may lead to reductive stress (116).

H₂S itself has a significant antioxidant capacity but is not as recognized. At high concentrations, H₂S can create reducing environments and cytotoxicity, but at low concentrations or from endogenous sources, it offers protective and beneficial effects (34, 176). H₂S can inhibit ROS via diverse mechanisms (92, 123, 156, 187, 195). It is known to directly scavenge O₂ ^•, H₂O₂, and other ROS (46, 184, 195). It can upregulate SOD and glutathione peroxidase, reducing oxidative stress in conditions such as diabetic nephropathy, and intestine or lung damage (30, 64, 90, 130). It also upregulates GSH as a potential mechanism to reduce oxidative stress (64). Pathways associated with antioxidant activity of H₂S include activation of nuclear factor erythroid 2-related factor 2 (NRF2) pathway involved in cardioprotection (22, 158), and inhibition of p38/MAPK and NF-κB pathways, lowering ROS, oxidative stress, and inflammation (28, 37, 80, 89, 173). These effects of H₂S are largely observed with exogenous H₂S-based therapies. CSE and CBS knockout (KO) mice show reduced GSH levels and are more prone to oxidative stress (137, 198, 204, 208), proving a relationship between oxidative stress and endogenous H₂S. Regarding the relationship between iron, H₂S, and oxidative stress, it has been recently shown by Zhou et al. that CBS is required for iron homeostasis in mice (208). They noticed that CBS-deficient mice had increased iron content in the liver, serum, heart, and spleen along with other markers of oxidative stress. Homozygous CBS-deficient mice could not survive past 4 weeks (208). Hepcidin was upregulated in the liver and circulation due to a decrease in transcriptional factor, NRF2. Increased hepcidin caused a loss of Fpn and increased iron retention, severely damaging the liver. This phenotype was reversed by reintroduction of CBS through the adenoviral vector (208). Figure 5 summarizes interaction between H₂S and Iron on cellular functions at a molecular level.

FIG. 5.

Interaction between H₂S and iron on cell functions. Iron overload causes production of ROS. This will activate NF-κB and p38/MAPK, leading to inflammatory reactions. Excess iron can also lead to nonenzymatic catalysis of cysteine to release H₂S. The latter quenches excess iron and inhibits NF-κB and p38/MAPK pathways and ROS production. The inhibition of L-type Ca²⁺ channels by H₂S decreases iron influx. H₂S also induces NRF2, activating anti-inflammatory genes, upregulating Fpn, and promoting the efflux of excess iron. NF-κB, nuclear factor kappa-light-chain-enhancer of activated B cells; NRF2, nuclear factor erythroid 2–related factor 2; p38/MAPK, p38/mitogen-activated protein kinase; ROS, reactive oxygen species. Color images are available online.

The Effects of Iron and H₂S on Erythrocytes

The human body produces ∼200 billion RBCs per day with each containing 300 million hemoglobin molecules with single iron moiety. This feat requires 20 mg of iron daily, which is mostly sourced through macrophage recycling of senescent RBCs, body iron stores, and dietary iron (161). Transport of oxygen by RBCs is the most prominent role of iron inside hemoglobin in the body. Low oxygen delivery is sensed by peritubular cells in the renal cortex and outer medulla in kidney, which release erythropoietin (EPO) in response (143). The EPO drives erythropoiesis by proliferation and differentiation of erythrocytes, which requires a large reserve of iron to produce hemoglobin (161). Transferrin-bound iron is kept high during each stage of nucleated erythroblast differentiation (141). Once the hemoglobin production ceases, iron uptake is reduced, and TfRs are released as final steps of erythroid differentiation (2).

Iron metabolism and erythropoiesis are linked, and defect in erythropoiesis often leads to iron overload (112, 136). Hepcidin plays an important role in regulating iron and its expression can be controlled by erythropoiesis (93). Hypoxia-induced factor (HIF) stimulates EPO and suppresses hepcidin, resulting in an increase in iron concentration. Liu et al. also found that hepcidin suppression indirectly requires EPO-induced erythropoiesis (93). Erythroferrone (ERFE), released from erythroblasts, can suppress hepcidin in hepatocytes, strengthening the link between erythropoiesis and iron metabolism (69).

H₂S and iron may interact on erythropoiesis in different ways. It has been recently demonstrated that chicken RBCs, which contain an intact nucleus and mitochondria, produce H₂S, mainly from MST enzyme (63). Human RBCs, which lack the nucleus and mitochondria, may produce limited H₂S by sulfur reduction nonenzymatically (12, 142, 170). Jennings demonstrated that deprotonated H₂S in the form of HS⁻ can be transported into the RBCs in exchange for a chloride ion through AE1 anion exchange protein as part of the Jacobs/Stewart cycle. The HS⁻ can be converted to H₂S in cells, which can take part in cellular functions or diffuse out (58).

H₂S is a regulator of erythropoiesis. CBS-deficient mice had systemic iron overload due to decreased erythropoiesis and iron utilization (204). The absence of CBS caused a lower expression of delta-aminolevulinate synthase 2, ferrochelatase, FLVCR, HIF-2α, and EPO, resulting in low erythropoiesis and anemia (204). Exogenous H₂S donor, GYY 4137, can stimulate erythropoiesis through HIF (85). Mouse lacking CSE had reduced EPO and hemoglobin in hypoxia, and patients with anemia associated with chronic kidney disease had low H₂S and EPO (84).

Regulation of Cardiovascular Functions by Iron and H₂S

Iron deficiency or overload has been associated with increased cardiovascular risk with different mechanisms (82). Iron deficiency can be caused by either underutilization of circulating iron or failure to absorb dietary iron (202). As a result of iron deficiency-induced anemia, heart workload increases along with elevated ischemia and myocardial cell death (82). Elevated renal ischemia can activate the renin/angiotensin system, causing increased salt and water retention and vasoconstriction, further putting stress on the already stressed out heart (51, 82). Anemic mouse models, IRP1/2 KO or TfR1 KO, showed decreased cardiac function and mitochondrial respiration, which were reversed with aggressive iron therapy (202). These findings have a connotation that iron deficiency due to underutilization or improper regulation may result in cardiovascular disturbance.

Iron overload can also produce detrimental effects on the cardiovascular system primarily through ROS generation and enhanced oxidative stress (82). After having tested adult and neonatal cardiomyocytes as well as H9c2 cells (rat myoblasts), Sweeney et al. demonstrated that iron overload increased ROS via mitochondria and xanthine oxidase. Moreover, iron overload decreased the autophagic reflux as seen by an increase in microtubule-associated protein 1A/1B-light chain 3-II (LC3-II) and p62 protein expression, which are markers for autophagy (160). This caused a decrease in insulin sensitivity in cardiomyocytes, which was confirmed in autophagy-deficient H9C2 cells prepared by overexpressing dominant negative form of autophagy-related 5 protein (Atg5) (157).

Júnior et al. measured effects of chronic iron overload on mesenteric resistance arteries isolated from rats. Chronic iron overload increased the wall to lumen ratio and thickness of the blood vessel, along with decreased lumen and external diameters. Decreased distensibility and increased stiffness and pulse wave velocity were also observed. With wire myography recording, Júnior et al. found that iron overload, induced by repeated injections of iron-dextran, increased the vasoconstrictor response in isolated mesenteric resistance arteries, but did not affect relaxation responses by acetylcholine and sodium nitroprusside. They concluded that iron overload-induced free radicals decreased nitric oxide bioavailability. This conclusion was supported by the observation that L-N^G-Nitro arginine methyl ester (L-NAME), an inhibitor of nitric oxide synthase, had no effect on vasoconstriction of mesenteric resistance arteries with iron overload, whereas L-NAME with SOD resulted in typical shifting of the phenylephrine contraction curve toward left. In vitro assays also showed a decrease in NO amount in iron overload tissues (65). Interestingly, the same group showed similar results with aortic tissues (103). Kuang et al. reported on the acute effects of iron overload on rat aortic tissues (77). They found that ferric ammonium citrate can transiently contract isolated aorta with an increased contraction amplitude at higher calcium concentrations. They also note that incubating tissues with ferric ammonium citrate impair vasoconstriction by phenylephrine, and this decrease is abolished by adding catalase or glutathione before ferric ammonium citrate. This suggests that iron overload-caused contraction dysfunction is due to accumulation of ROS (77). Sangartit et al. report an increase in blood pressure and impaired vascular function in iron-overload mice (139). Nitric oxide and glutathione levels in the blood were decreased by iron overload but reversed by tetrahydrocurcumin and deferiprone, compounds with antioxidant and anti-inflammatory properties (139). Iron overload increased calcification in human aortic smooth muscle cells and increased interleukin (IL)-24, thereby accelerating arteriosclerosis (70). Similarly, iron overload accelerated atherosclerosis and induced endothelial dysfunction in apolipoprotein E-deficient mice due to oxidative stress (102). Hypertension as a complication or comorbid factor is also found in patients with sickle cell anemia (4), thalassemia (41), kidney disease (168), and liver disease (36). Recent research has shown that hypertension in sickle cell anemia and thalassemia patients is due to the toxic effects of chronic iron overload rather than anemia (4, 41). An increased cardiac output, a decreased heart rate variability, and vascular damage due to oxidative stress were observed in both diseases (4, 41, 49, 185). These insights warrant further research into the mechanisms of iron overload and vascular damage.

ROS may induce vascular damage and activation of p38/MAPK pathway. The consequential endothelial dysfunction, elastin remodeling, and vascular stiffness can result in hypertension (105, 166, 179). Inhibitors of p38/MAPK are known to ameliorate vascular damage and reduce endothelial dysfunction and hypertension (179). Iron overload has been shown to stimulate ROS production and p38/MAPK activation in MC3TC-E1 preosteoblast cells (26). Another mediator of inflammation, NF-κB pathway, also increases vascular damage and is believed to be pivotal in the pathogenesis of hypertension and can be activated by iron overload and ROS (19, 33).

One of the most prominent effects of H₂S is vasorelaxation (198). The discovery that H₂S caused vasorelaxation invoked significant interest into H₂S biology worldwide. One of the earliest articles by our laboratory found that H₂S at physiologically relevant concentrations opens K_ATP channels, which leads to vasorelaxation (206). We subsequently demonstrated that mouse lacking CSE-gene had hypertensive phenotype because CSE is the major enzyme for H₂S generation in vasculature (198, 206). Vascular response to H₂S is also endothelium dependent, not inhibited by 1H-[1,2,4]oxadiazolo-[4,3-a]quinoxalin-1-one (ODQ) or NS2028, two potent inhibitors of guanylate cyclase pathway (205). H₂S can mediate the phosphorylation of endothelial nitric oxide synthetase through p38/MAPK and Akt-dependent pathway, increasing bioavailable NO for vasorelaxation (5). The interaction of H₂S and NO on vasorelaxation also involves the redox regulation of soluble guanylyl cyclase (sGC). Zhou et al. suggested that H₂S reduced ferric (Fe³⁺) heme center of sGC to its ferrous (Fe²⁺) form in cultured rat aortic smooth muscle cells. By doing so alone, H₂S would not activate sGC. In the presence of H₂S, however, sGC can be conditioned for activation by NO since the latter binds to ferrous (Fe²⁺) heme of sGC to increase its catalytic activity. Consequently, cGMP level was increased and vasorelaxation ensued (209).

H₂S also suppresses the development of atherosclerosis (100) by inhibiting adhesion molecule expression and vessel intimal proliferation. We also observed that decreased endogenous H₂S leads to atherosclerosis by vascular remodeling (99). Estrogen and H₂S both inhibit atherosclerosis in CSE-wild-type (WT) mice fed with atherosclerotic diet, but estrogen cannot inhibit atherosclerosis in CSE-KO mice fed with the same diet. Markers of atherosclerosis, such as intracellular adhesion molecule 1 (ICAM1) and NF-κB, also increased more in CSE-KO than CSE-WT on atherosclerotic diet. Finally, the observation that estrogen increases H₂S concentration in livers of CSE-WT but not CSE-KO leads to the conclusion that estrogen interacts with H₂S to inhibit atherosclerosis (87). We recently reported that insulin-like growth factor 1 receptor (IGF-1R) interacts with estrogen receptor-α (ER-α) to cause atherosclerosis, but this interaction is inhibited by H₂S, as the latter can inhibit IGF-1R by S-sulfhydration (147). Zheng et al. demonstrated that H₂S donor, GYY4137, decreased atherosclerotic plaques in aorta by decreasing ICAM1, vascular cell adhesion molecule 1 (VCAM1), and inflammatory markers such as tumor necrosis factor-α (TNF-α), monocyte chemoattractant protein 1 (MCP1), IL-6, and IL-1β in diabetes-accelerated atherosclerotic mice (207). It was concluded that H₂S inhibits NLR family pyrin domain containing 3 (NLRP3) inflammasome, which ultimately led to a decrease in atherosclerosis (207).

The role of H₂S as an inhibitor of smooth muscle proliferation and viability is well documented by our research team (100, 147, 148, 194, 197, 199) and others. The absence of CSE expression causes vascular smooth muscle cells (VSMCs) to overproliferate (197). Exogenous addition of 100 μM H₂S significantly increased VSMC apoptosis and phosphorylation of extracellular signal-regulated kinase (ERK) 1/2 in CSE-deficient mice. ERK regulates apoptosis, growth, and cellular response to stress. H₂S also decreased cyclin D1 and increased p21Cip/WAF-1 expression, which arrests cell cycle in G1 phase (197). These findings have great implications in the pathogenesis of atherosclerosis. We recently demonstrated that estrogen increased endogenous H₂S production to regulate smooth muscle cell proliferation (87). Iron is also associated with atherosclerosis, but this role of iron is often debated (191). Mueller et al. reported that the addition of ferrous ions decreased proliferation of smooth muscle cells (110). Conversely, Porreca et al. report that iron chelation and deficiency also inhibit proliferation of VSMCs (129). Although iron accumulation is observed in atherosclerotic plaques, epidemiological data are conflicting regarding the association between iron overload and atherosclerosis in patients (169). H₂S can offer protection against iron overload, and H₂S may be released as a consequence of iron overload. This may very well be the reason why it has been difficult to prove the link between iron and atherosclerosis, whereas a strong link between H₂S and antiatherosclerosis already exists. For example, it was found that iron overload can stimulate ERK1/2 in SH-SY5Y neuroblastoma cells (8). Whether the activation of ERK results from iron overload or from the consequential increase in H₂S is unsettled. Epidemiological data also suggest that females are at lesser risk of cardiovascular diseases and iron overload (31). The discovery that estrogen can induce H₂S production may offer a valid explanation on why females are at lesser risk of iron overload and cardiovascular diseases.

H₂S also regulates vascular endothelial proliferation and function (178). H₂S stimulates angiogenesis and endothelial repair (121). H₂S is also an endothelium-derived hyperpolarizing factor (EDHF), along with K_ATP channel opener, which establishes the role of H₂S in regulation of vascular tone (178). Jian et al. reported that iron deficiency is associated with increased angiogenesis due to upregulation of vascular endothelial growth factor (VEGF) and stabilization of hypoxia-inducible factor-1α (60). Iron overload, on the contrary, is associated with increased oxidative stress and MAPK activation (60). Interestingly, H₂S is proangiogenic under normoxic conditions through phosphorylation of PI3K/AKT, ERK, and p38/MAPK pathways, but it is antiangiogenic under hypoxic conditions by inhibiting translation of HIF-1α (177, 186). Liu et al. reported that H₂S promoted angiogenesis by stabilizing HIF-1α in hypoxia by using cobalt to induce a chemical hypoxia (94). The conflicting conclusions may be due to different models of hypoxia, along with differences in chronic and acute hypoxia. Regardless, there seems to be a link between iron deficiency-induced angiogenesis and H₂S-induced angiogenesis. Figure 6 summarizes potential avenues for interaction between H₂S and iron in the cardiovascular system.

FIG. 6.

Interactions between iron and H₂S in the cardiovascular system. Iron deficiency can increase heart workload, hypertension, angiogenesis, and erythropoiesis. Iron overload can increase oxidative stress and damage heart and blood vessels. On the contrary, H₂S offers cardioprotection, decreases oxidative stress, inhibits smooth muscle cell proliferation, and decreases hypertension. H₂S also increases angiogenesis and erythropoiesis. Color images are available online.

Molecular Mechanisms of Interaction Between H₂S and Iron

As summarized in the previous sections, iron causes oxidative stress and increases markers of inflammation, whereas H₂S can inhibit p38/MAPK, NF-κB, and ROS (38a, 156, 158, 173). These are all possible mechanisms where H₂S can potentially decrease the deleterious effects of iron overload and ameliorate hypertension. Cysteine can undergo catalysis under the influence of ferric ions to release H₂S as explained before (200). This nonenzymatic H₂S generation is believed to be a protective mechanism to decrease excess iron as H₂S can react with iron to form acid labile iron sulfide, quenching and inhibiting the excess iron from participating in oxidative stress (200). H₂S has been shown to have a cardioprotective role through activation of NRF2 (22), but this role is yet to be established in vasculature. The increased NRF2 upregulates Fpn1, which will lead to efflux of iron and ultimately decrease iron overload (40, 203). CBS KO mice showed increased iron content in the heart, decreased Trf1, Fpn1, and IRP1 by a decrease in hepcidin (208). The effect of H₂S on TfR1 and Fpn1 has only been demonstrated in macrophages and spleen tissue. Should the similar effect occur in the cardiovascular system, it may mitigate detrimental effects of anemia (203).

To reiterate, iron deficiency or overload causes cardiovascular diseases and H₂S can potentially ameliorate the detrimental effects of abnormal iron metabolism. H₂S can accomplish this through direct regulation of iron and transport and indirectly through suppressing downstream targets of iron dysfunction such as the renin/angiotensin system, ROS generation, p38/MAPK, and Nf-κB regulation. Furthermore, H₂S can offer enhanced protection through Nrf2 signaling to lower iron-induced oxidative stress.

Iron and H₂S Interaction in the Liver

The liver is the command center for iron metabolism, chiefly through expression of hepcidin. High levels of hepcidin can inhibit Fpn, decreasing iron release into the circulation through enterocytes and iron recycling by macrophages (13). The liver produces a significant amount of H₂S (21, 66) and excretes H₂S-producing enzymes, such as CSE and CBS, into the blood stream (9, 177). Iron and H₂S can interact through different pathways, including the BMP/SMAD pathway, which governs apoptosis, cellular differentiation, embryogenesis, organogenesis, and osteogenesis (115). Another important pathway is the JAK/STAT pathway, which can affect genes responsible for apoptosis, cell differentiation, proliferation, and migration (135). Hepcidin expression is controlled by BMP/SMAD signaling in the liver sinusoidal endothelial cells (150). Activin receptor like kinase 2 and 3 (ALK2 and ALK3) can activate BMP6 and BMP2, respectively, in response to increased tissue iron or basal expression of hepcidin. The BMP-SMAD pathway also interacts with JAK/STAT3 pathway, which regulates hepcidin produced due to the inflammatory response (150).

Hepcidin is also regulated by erythropoietic drive, which stimulates the kidney to produce EPO in response to hypoxia. This increased EPO results in erythroblast differentiation, leading them to secrete ERFE to suppress hepcidin (175). Although direct regulation of ERFE by H₂S is yet to be established, H₂S may increase it indirectly by increasing EPO. Similarly, inflammatory cytokine IL-6 can upregulate hepcidin by the JAK-STAT3 pathway interacting with SMAD1/5/8 pathway and Activin B (174). It has been shown that H₂S reduced hepcidin expression through JAK-STAT pathway by suppressing pSTAT3 leading to decreased IL-6 (203). A similar report revealed that H₂S promoted SIRT1 expression, which stabilizes SIRT1-STAT3, reducing STAT3 acetylation. The latter event lowered hepcidin level (190).

It is evident that hepcidin expression is affected by a JAK/STAT and BMP/SMAD axis. The fact is that H₂S can also affect these pathways, and ultimately, hepcidin expression points toward a key role of H₂S in regulating physiological iron levels.

Iron and H₂S Interaction in the Brain

As discussed earlier, iron is important for energy metabolism because it is bound to complexes in electron transport chain such as flavin adenine dinucleotide. Iron/sulfur clusters and heme also serve as cofactors for different enzymes. In the brain, iron serves as a cofactor in synthesis of neurotransmitters, such as tryptophan hydroxylase and tyrosine hydroxylase (109). Absorption of iron through the blood/brain barrier (BBB) is analogous to dietary iron absorption in the duodenum. Once crossing the BBB, it is oxidized to Fe³⁺, bound to transferrin, and released into the brain circulation. Transferrin-bound iron is thought to be either broken down in endosomes and released into the cytosol by DMT1 and exported by Fpn or to be transported into the endosomes from luminal to abluminal surface where endosomes release iron between brain endothelial cells and astrocytes. Furthermore, Fe²⁺ can bind to ATP or citrate to be transported as NTBI in oligodendrocytes and astrocytes, which do not express TfR1 (11). Iron concentration in the brain is tightly regulated. Abnormal iron concentrations are associated with neurodegenerative diseases, such as Parkinson's disease (PD), Alzheimer's disease (AD), Huntington disease, and neurodegeneration (62).

The physiological role of H₂S in the nervous system has evolved to include neuroprotection, neuromodulation, and nociception (95). H₂S also induces long-term potentiation and increases the sensitivity of NMDA receptor-mediated memory response. Administration of NaHS elicited an anti-inflammatory response to counter LPS-induced inflammation in microglia by inhibiting p38/MAPK (53). H₂S increases [Ca²]_i for neuronal health and function (95). Moreover, H₂S can inhibit NOX and decrease oxidative stress in the brain (138). Studies have also shown the importance of H₂S in peripheral nerve regeneration and degeneration, significant for Schwann cell proliferation and dedifferentiation (120). The role of H₂S in various neurodegenerative diseases has been reviewed by Paul and Snyder (122).

With PD, the substantia nigra region of brain has considerable dopaminergic neuronal cell death (153). Symptoms of this disease include postural instability, tremors, and rigidity. The neuropathological characteristics include Lewy bodies, which are intracytoplasmic inclusions, widely distributed in paralimbic and neocortical regions of substantia nigra. Neuromelanin level, associated with iron storage, decreases with age, leading to high iron accumulation and oxidative stress. DMT1 isoform was upregulated in PD with increased iron uptake and dopaminergic cell death. A decrease in ceruloplasmin in PD is also observed in substantia nigra, which leads to increased iron deposition (7). Iron chelation therapy has shown to be effective in decreasing neurotoxicity in PD (10, 68). H₂S offers a protective role in neurotoxin-induced PD in rat models by reducing oxidative stress and inflammation by regulating different pathways, including PKC, p38, JNK, PI3K/Akt, and ERK-MAPK (163). Endogenous H₂S was downregulated in substantia nigra by 6-hydroxydopamine (6-OHDA), whereas exogenous administration of H₂S reversed 6-OHDA-induced neuronal death, suggesting a protective role of H₂S in PD (23).

AD is a neurodegenerative disease with impairment of memory, reasoning, and perception as its prominent features. Pathological features include insoluble amyloid beta protein (ab) aggregates and neurofibrillary tangles (NFTs). NFTs contain hyperphosphorylated tau proteins, which can reduce Fe³⁺ to Fe²⁺, increasing NFT production (193). Amyloid precursor protein (APP) is a transmembrane protein 1 ubiquitously found in the brain. It is upregulated in the presence of iron and causes iron deposition in amyloid plaques. APP is like ceruloplasmin and aids in the efflux of iron and converts Fe²⁺ to Fe³⁺. High iron can upregulate paired amino acid cleaving enzyme, resulting in an increase in amyloidogenic peptides (117).

H₂S levels in brain are significantly decreased along with the expression of CBS in patients with AD (35). Exogenous H₂S administration slowed down AD progression in mouse and rat models (47, 48). In these animal models, H₂S lowered ab plaques in the hippocampus and proteins associated with AD progression. Also, H₂S was found to modulate inflammation and apoptosis of hippocampus cells and with mice of different ages with AD (167). H₂S may protect against AD by directly reducing HOCl-induced cytotoxicity, protein and lipid peroxidation, along with homocysteine-induced oxidative stress (180). There is a positive association with homocysteine and iron level and deposits in arterial plaques, suggesting iron/homocysteine interaction (50). H₂S has been reported to inhibit homocysteine-induced AD-like pathology (67). This raises an interesting question whether the iron/homocysteine/H₂S axis exists in regulating brain functions under physiological and pathophysiological conditions.

Conclusion

The interactions between iron and H₂S impact each other's metabolism and functions. Whereas excess iron produces ROS, H₂S can quench excess iron and ROS altogether. Conversely, whereas low iron causes anemia, H₂S can downregulate hepcidin to release more iron. The interaction is also seen at the molecular level where iron and H₂S can act on same cellular pathways but with opposite effects. For example, iron catalyzes ROS-activated p38/MAPK and NF-κB and H₂S can counter this effect. Given that excess iron can increase H₂S production nonenzymatically, the effect of iron on H₂S signaling pathways and targets is much less known. The interactions between iron and H₂S have been observed in numerous systems, including the brain, blood, liver, and cardiovascular system. Atherosclerosis is a case in point. Iron can increase H₂S and the latter would protect from atherosclerosis. On the contrary, iron overload accelerated the development of atherosclerosis. The net outcome of iron overload and induced H₂S production on atherosclerosis development is difficult to speculate yet. The similar dilemma also challenges the contributions of iron overload and H₂S level, respectively, and/or integrated, to the development of hypertension. While having acknowledged these challenges, there is no doubt that a better understanding of the interactions between iron and H₂S would shed light on the pathogenesis of and treatment strategies for many diseases. In the future, physiological and pathophysiological importance of H₂S and iron as well as their therapeutic applications should be evaluated jointly, not separately. Future investigation should also expand from iron-rich cells and tissues to the others in which H₂S and iron interaction has not received due attention.

Footnotes

Authors' Contributions

All the authors have equally contributed to the writing, editing, and revising of the article and figures.

Author Disclosure Statement

The authors declare that they have no competing interests.

Funding Information

This study has been supported by a Discovery Grant from the Natural Sciences and Engineering Research Council of Canada to R.W.

Abbreviations Used

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Signaling Integration of Hydrogen Sulfide and Iron on Cellular Functions

Abstract

Significance:

Recent Advances:

Critical Issues:

Future Directions:

Introduction

H2S Metabolism

Iron Metabolism

Iron and H2S in Redox Signaling

The Effects of Iron and H2S on Erythrocytes

Regulation of Cardiovascular Functions by Iron and H2S

Molecular Mechanisms of Interaction Between H2S and Iron

Iron and H2S Interaction in the Liver

Iron and H2S Interaction in the Brain

Conclusion

Footnotes

Authors' Contributions

Author Disclosure Statement

Funding Information

Abbreviations Used

References

H₂S Metabolism

Iron and H₂S in Redox Signaling

The Effects of Iron and H₂S on Erythrocytes

Regulation of Cardiovascular Functions by Iron and H₂S

Molecular Mechanisms of Interaction Between H₂S and Iron

Iron and H₂S Interaction in the Liver

Iron and H₂S Interaction in the Brain