Introduction

Renal ischemia-reperfusion (IR) injury, defined as the cessation and restoration of renal blood flow (RBF), is a major cause of acute kidney injury (AKI) [1, 2]. AKI is a worldwide health issue that is associated with a high rate of morbidity and mortality [3]. Renal IR injury is most commonly caused by transient or persistent renal hypoperfusion [4]. However, it may also be owing to temporary renal artery occlusion during suprarenal aortic aneurysm repair, nephrectomy, renal transplantation [5], cardiovascular surgery, hypovolemic, and septic shock [6, 7].

Interruption of RBF during the ischemic phase decreases medullary blood flow and reduces oxygen and glucose supply to tubular structures in this area, resulting in an imbalance in delivery, and demand [5]. Reduced O2 levels alter metabolism from aerobic to anaerobic, which cannot provide the requirement of aerobic tissues, resulting in fast depletion of intracellular ATP levels [8]. ATP depletion causes an increase in cytoplasm Ca2 + charge, hypoxanthine level inside the cells, and production of reactive oxygen species (ROS), which finally results in acidosis [9]. Apoptosis and necrosis are triggered by hypoxia, glucose deprivation, acidosis, and the formation of ROS [10]. In addition, reduced intracellular pH and ATP levels during the ischemia phase result in (1) lysosome membrane instability, followed by lysosome enzymes exudation and cell structure disturbance, and (2) inhibition of ionic pumps, especially the Na+/K+ ATPase [11].

Inhibition of Na+/K+ ATPase activity and Na+/H+ antiporter function, which attempts to correct the intracellular pH by pumping Na+ ions into the cell and protons out of the cell, results in increased intracellular Na+, water, and edema [12, 13]. Furthermore, because of ATP depletion, the cessation of Ca2+ pumping out of the cells and the suppression of Ca2+ reuptake into the endoplasmic reticulum raises intracellular Ca2+ levels. Furthermore, the cessation of Ca2+ pumping out of the cells and the suppression of Ca2+ reuptake into the endoplasmic reticulum due to ATP depletion raises intracellular Ca2+ levels [9].

The cytoplasmic Ca2+ overload activates Ca2+-dependent proteases like calpains, proteases, phospholipases, and caspases, which are inactive due to the acidic environment but can cause cell damage after pH normalization during reperfusion [9, 14]. Besides, Ca2+ overload leads to ROS production in mitochondria during ischemia, which is followed by the opening of the mitochondrial transition pore (mPTP) in reperfusion as a result of pH normalization, resulting in apoptosis and cell death [15]. Despite the fact that quick reperfusion recovers the oxygenation and substrates needed for aerobic ATP production and normalizes extracellular pH by washing out accumulated H+, it creates additional damage known as reperfusion damage [9, 16, 17]. Ca2+ overload causes activation of the calpains, ROS production, reduction of antioxidant capacity, mPTP opening, apoptosis, necrosis, endothelial dysfunction, alterations in outer medullary microcirculation, inflammation, and tubular injury [5, 18, 19].

TRPM2, the second TRPM channel subfamily member, enhances intracellular Ca2+, ROS generation, and oxidative stress, all of which are implicated in physiological and pathological processes [6, 20, 21]. TRPM2 has been found to play a deleterious role in IR injury in a variety of tissues, including the kidney, brain, and pancreas [22, 23]. Therefore, it is necessary to review the important pathophysiological mechanisms mediated by TRPM2 activation in renal IR injury. This review will focus on new results that provide light on the role of TRPM2 in IR-induced inflammation, oxidative stress, apoptosis, and AKI. Fig. 1 shows how TRPM2 channel opening activates oxidative stress, apoptosis, and inflammation signaling pathways in renal IR injury (Fig. 1).

Fig. 1.
figure 1

Schematic diagram of activation of inflammatory cytokines and cells, production of ROS, oxidative stress, and apoptosis in renal IR injury and the role of TRPM2 channels activation in these pathways.

VCAM: Vascular cell adhesion molecule, ICAM: Intercellular Adhesion Molecule, ROS: Reactive oxygen species, NUDT9-H: Nudix-like domain homology domain, PARG: Poly (ADP-Ribose) Glycohydrolase, PARP: Poly(ADP-Ribose) Polymerase, ADPR: Adenosine diphosphate ribose, NFkB: Nuclear Factor kappa B, ERK: Extracellular signal-regulated kinases, AIF: apoptosis-inducing factor.

Full size image

Materials and methods

For this review, we gathered data from a variety of sources, including PubMed, Scopus, Web of Science (WOS), and EMBASE. The following keywords and or their equivalents were used for the search strategy; TRPM2, acute kidney injury, oxidative stress, inflammation, and renal ischemia-reperfusion.

General structural features of TRPM2

Transient receptor potential (TRP) channels with six transmembrane domains belong to a superfamily of monovalent and divalent cation permeable ion channels [24]. TRP channels are implicated in many biological processes, i.e., regulating vascular tone, permeability, proliferation, cell volume, secretion, apoptosis, angiogenesis, and cell death [25].

Based on differences in amino acid sequence similarities between the different gene products, this superfamily with 28 members is classified into six subgroups: canonical (C), vanilloid (V), melastatin (M), mucolipin (ML), polycystin (P), and, ankyrin (A) [26]. TRPM1 to TRPM8 are members of the TRPM subfamily. TRPM1 is the name of the first member to be characterized (M, melastatin) [27].

TRPM2 was first isolated from the human brain and named LTRPC-2 or TRPC7. This channel was later classified as TRPM2 [28]. The TRPM2 gene is found on chromosome 21q22.3 in humans. Its 6.5-kilobyte transcript codes a 1503-amino-acid protein with a molecular weight of 170 kDa. It covers 90 kb which contains 32 exons. The trpm2 mouse gene has 34 exons and produces a 1507-amino-acid protein and is highly similar to human TRPM2 gene [23, 24].

The TRPM2 channel is a protein with 1503 amino acids with six transmembrane fragments, namely S1 to S6, a loop domain between S5 and S6 that forms pores, and intracellular N and C termini [29, 30]. TRPM2 contains four homologous domains and a calmodulin (CaM)-binding IQ-like motif at its N terminus, which is needed for channel activation [31]. The TRP box, coiled-coil domain (CCD), and NUDT9 (Nudix-like domain or NUDT9 homology domain) are all found at the C terminus (NUDT9-H) [30, 31]. The most common endogenous ligand for TRPM2 is adenosine diphosphate ribose (ADPR), which binds to the NUDT9-H domain and opens the channel, allowing Ca2+ inflow. The enzymatic activity of NUDT9H catalyzes the conversion of ADPR to ribose-5-phosphate (R5P), with adenine monophosphate (AMP) acting as a negative regulator for ADPR gating TRPM2 [21, 30, 32]. TRPM2 has both ADPR hydrolase enzymatic and ion channel gating activity, and is referred to as a “coenzyme” because of its ability to convert ADPR into AMP and R5P [25, 29, 33]. Calmodulin (CaM) has been demonstrated to have a key role in TRPM2 channel facilitation and activation; it interacts directly with the IQ-like motif and modifies TRPM2’s Ca2+dependent activation, increasing intracellular Ca2+ levels [34].

TRPM2 has been found in a variety of organs (kidney, brain, heart, lung, liver, spleen, bone marrow, and pancreas) and cell types (neurons, hematopoietic, cardiomyocytes, endothelial cells, and pancreatic β -cells) [25, 26, 35, 36]. TRPM2 is almost expressed in the cortex and outer medulla of the kidney’s proximal tubular epithelial cells. TRPM2 receptors were also shown to be intracellular without a clear plasma membrane localization, whereas glomeruli, peritubular endothelial cells, and interstitial cells were spared [25]. In the ischemic AKI, the TRPM2 channel is activated by different molecules, including cyclic ADPR (cADPR), hydrogen peroxide, and Ca2+ [37] that are implicated in oxidative stress, cell death, apoptosis, and inflammation [36, 38].

Regulation of TRPM2

TRPM2 activators

Although ADPR is the most common TRPM2 activator, various ADPR analogs, including ADPR-2′-phosphate, ADPR2′-O-acetyl-, and 2′-deoxy-ADPR can activate the channel [6]. Furthermore, it has been shown that TRPM2 can be activated by several second messengers associated with adenine nucleotides that are metabolically related to ADPR, including cyclic ADPR (cADPR), nicotinamide adenine dinucleotide (NAD), and nicotinic acid adenine dinucleotide phosphate (NAADP) [6, 29]. Other molecules like reactive nitrogen species (NOS), ROS, and H2O2 are triggers of the TRPM2 channels and bind to them in pathological conditions, including oxidative stress, inflammation, and cell death [25, 39]. Besides, TRPM2 is activated by tumor amyloid β-peptide, necrosis factor α (TNF-α), and concanavalin A. All of these extracellular signals cause ADPR production. ADPR binds to the TRPM2 NUDT9-H domain on the C-terminus and activates TRPM2, permitting substantial penetration of monovalent or divalent cations such as K+, Na+, Zn2+, and Ca2 + [6, 24].TRPM2 is also regulated by intracellular Ca2+. Therefore, when ADPR interacts with TRPM2, Ca2+ is released [36]. TRPM2 is entirely activated by Ca2+; thus, the loss of outer or inner Ca2+ prevents ADPR from inducing TRPM2 currents. This impact might be due to the canal’s increased sensitivity to ADPR [40, 41]. Ca2+ also acts as a concentration-dependent gate for the TRPM2 channel. This can be due to conformational changes induced by CaM interaction with the TRPM2 IQ-like motif [6]. Ca2+-bound CaM enhances the interaction between CaM and the IQ-like motif, providing positive feedback for TRPM2 stimulation [36].

TRPM2 inhibitors

The first TRPM2 channel inhibitor discovered was adenosine monophosphate (AMP), which is formed by hydrolysis of ADPR. AMP competes with ADPR by binding to binding sites located on the NUDT9-H domain of TRPM2 ion channels [42]. As a TRPM2 antagonist, 8-Br-ADPR can compete with ADPR to prevent TRPM2 activation [37]. Furthermore, protons and a variety of divalent heavy metal cations like Cu2+, Hg2+, Pb2+, Fe2+, Se2+, and Zn2+ inhibit TRPM2 currents, all of these factors are targets of the TRPM2 channel pore region. Some structurally unrelated pharmacological agents that have been found to block the TRPM2 activity include clotrimazole, flufenamic acid (a member of nonsteroidal anti-inflammatory drugs), N-(p-amylcinnamoyl) anthranilic acid, and 2-Aminoethoxydiphenyl borate (2-APB) [6].

The role of TRPM2 channels in renal IR injury

TRPM2 and inflammation in renal IR injury

The inflammatory process triggered by renal IR damage induces cascades of proinflammatory cytokines (IL-1, IL-6, IL-10, and TNF-α) [43], chemokines (MCP, MIP-2, and IL-8) [44, 45], expression of various adhesion molecules (ICAM (Intercellular adhesion molecule), VCAM (Vascular cell adhesion molecule), P selectin, and E selectin by endothelial and parenchymal renal cells [46, 47]. The combination of cytokines, chemokines, and adhesion molecules recruit leukocytes and neutrophils infiltration into the ischemic kidney, resulting in improved leukocyte-endothelial interactions, the generation of additional ROS and cytokines, and eventually significant progression of kidney damage [48, 49].

TRPM2 expression was found in tubular epithelial cells across the cortex and outer medulla using immunofluorescence. TRPM2 is also found in hematopoietic cells [31, 35]. Gao et al. have shown that mice Trpm2 knockout (Trpm2-KO) models are resistant to renal IR injury [31]. After IR injury, inflammatory cells like neutrophils entered the kidneys of Wild-Type (WT) mice, while this effect was less observed in Trpm2 KO animals. Chemokine production in monocytes is triggered by TRPM2-mediated Ca2+ influx, exacerbating neutrophil inflammatory properties [50, 51]. Because inflammation is an important mediator of IR injury, TRPM2 expression on hematopoietic cells could explain Trpm2-KO mice’s resistance to IR damage. Using bone marrow chimeric mice with Trpm2 KO in either hematopoietic or parenchymal cells, Gao et al. discovered that Trpm2 KO in parenchymal cells protected the kidney against IR injury, whereas Trpm2 KO in hematopoietic cells had no effect on ischemia-induced kidney damage. According to these data, TRPM2 in renal parenchymal cells appears to be a mediator in renal IR damage [31].

TRPM2-induced intracellular Ca2+ increase activates Ca2+-dependent signaling pathways such as the multi-subunit IB kinase (IKK) complex and extracellular signal-regulated kinase (ERK), which leads to nuclear factor B (NF-κB) activation [52, 53]. Studies identified the activation of NF-κB [54,55,56] and TRPM2 after renal IR damage [6, 31]. TRPM2 has been shown to modulate NF-κB signaling so that TRPM2 deficiency inhibited NF-κB signaling by blocking Jun N-terminal kinases (JNKs) signaling [57]. These findings imply that TRPM2 inhibiting and then preventing NF-κB activation can protect the kidney against inflammation.

Kurata et al. investigated the effects of TRPM2 on ischemic kidney in trpm2 KO mice by inducing bilateral IR at various ischemia periods (20, 25, and 30 min). A comparison of renal function in trpm2 KO and WT mice after 30 min of ischemia revealed a significant rise in plasma creatinine in KO mice, but not in 20 or 25 min of ischemia. In trpm2 KO, mRNA expression of kidney injury molecule 1 (Kim1) was considerably greater in 25 and 30 min of ischemia, and monocyte chemoattractant protein-1, (MCP-1 or Ccl2) was significantly higher in 20 and 25 min of ischemia. In contrast to the protective effect of TRPM2 deletion in the IR injury (28 min) described by Gao et al., trpm2 KO was deleterious in this investigation, as evidenced by a rise in plasma creatinine and higher mRNA expression levels of Kim1 and Ccl2 in several ischemia times. These disparities are most likely due to varied anesthetics and ischemia times in the two studies [58].

TRPM2 and oxidative stress in renal IR injury

When the balance between oxidants (free radicals) and antioxidants is disrupted, oxidative stress results, which, depending on the severity and duration, can cause tissue damage [59, 60]. Mammalian healthy cells produce a tiny amount of ROS when cellular metabolism is normal [61]. It is established that ROS moderate amounts affect many cellular signaling pathways, as well as proliferation, therefore playing a vital role in preserving cellular and tissue homeostasis [61].

It is known that several pathological conditions like IR injury, mitochondrial dysfunction, metabolism dysfunction, drug overdose, elevated Ca2+ levels, and aging increase the formation of ROS [62, 63]. Free radicals cause renal damage by DNA destruction, protein dysfunction, and lipid peroxidation ultimately leading to cell death [64].

The antioxidant defense systems deteriorate after renal IR injury, and the activity of antioxidant enzymes such as superoxide dismutase (SOD) and catalase (CAT) decreases, while the quantity of malondialdehyde (MDA), a lipid peroxidation product, rises [65]. Similar to oxidants, TRPM2 channels increase the cell membrane’s permeability to Ca2+, which promotes endothelial damage and ultimately, cell death [66, 67]. An increase in intracellular Ca2+ driven by Ca2+ entry through TRPM2 channels is part of the oxidant-induced breakdown of endothelial barrier function. Therefore, TRPM2 is considered a cellular redox sensor [68, 69]. As a result of these events (intracellular Ca2+ and oxidative stress), ADPR is generated and binds to the NUDT9-H domain, activating TRPM2 [70]. The production of ADPR takes place via two enzymes, including nuclear poly (ADPR) polymerases (PARPs) and poly (ADPR) glycohydrolases (PARGs) [71, 72]. PARP is triggered by oxidative stress-induced DNA damage and performs DNA repair tasks; whereas PARG hydrolyzes PARP chains to release free ADPR. Another avenue for ADPR formation in mitochondria is when oxidative stress results in the production of free ADPR [72, 73].

TRPM2 channel activation leads to a rise in Ca2+ entry and, as a result, a rise in Ca2+ adherence to calmodulin (CaM). CaM interaction with an IQ-like motif in the N-terminus of TRPM2 produces a positive response, activating TRPM2 channels and boosting the flow of Ca2+ through them. Increased intracellular Ca2+ activates Ca2+ dependent phospholipase A2, endonuclease, and proteases, ultimately leading to cell death in cells expressing TRPM2 channels [74,75,76]. While oxidative stress results in an excess of Ca2+, downregulating TRPM2-L, inhibiting TRPM2-S, or chelating Ca2+ might reduce TRPM2 activity and hence limit the rise in Ca2+ [28, 66, 77].

Furthermore, oxidative stress induces lipid peroxidation and the subsequent formation of highly electrophilic aldehydes such as 4-hydroxynonenal (4-HNE) that participate in renal IR injury [59, 60]. IR causes a significant rise in 4-HNE in WT mice kidneys, but Trpm2-KO mice have significantly lower 4-HNE levels [60]. The levels of H2O2, MDA, SOD, CAT and Glutathione (GSH) are increased in ischemic renal tissue while injection of 8 bromocyclic ADP ribose (8-BrcADPR) as an antagonist of cADPR, decreases the levels of all these enzymes[78]. Besides, renal IR injury increases TRPM2 expression while 8BrcADPR decreases it [37]. These findings show that cADPR is an excellent Ca2+ entry coreceptor via TRPM2 channels and it is possible that 8-Br‐cADPR altered oxidant and antioxidant enzymes levels (increase and decrease, respectively) by decreasing TRPM2 expression [37].

Tamm-Horsfall protein (THP) levels in AKI have been related to ROS production, and systemic oxidative stress. A HEK 293 recombinant cell line producing an inducible version of TRPM2 was used by La Favers et al. to demonstrate that THP suppressed TRPM2-mediated Ca2+ current and oxidative stress. The oxidative stress in vehicle-treated THP−/− animals was greater than in vehicle-treated THP+/+ animals. Inhibition of TRPM2 using 2-APB significantly reduced oxidative stress in THP+/+ and THP−/− mice exposed to IR injury. The disparity in oxidative stress found between THP+/+ and THP−/− animals was abolished after using 2-APB, showing that TRPM2 is a prime target for THP’s inhibitory impact on systemic oxidative stress [79].

TRPM2 and apoptosis in renal IR injury

In kidney cells of WT mice, IR damage increases apoptotic cell death by activating caspase-9 and caspase-3, reducing the expression of anti-apoptotic Bcl-xL and Bcl-2 proteins, which was reversed in Trpm2-KO animals exposed to IR [31]. This effect indicates that TRPM2 channels activate the apoptotic pathways after IR.

NADPH oxidase (NOX) is a transmembrane enzyme with several isoforms, including NOX oxidase 1 & 2, NOX1–5, NOX1 organizer, and NOX1 activator. The transfer of electrons from NADPH to O2 by NOX is the primary source of renal ROS in the cortex [80, 81].

IR damage stimulates NADPH oxidase and RAC1 (a key element of NAPDH oxidase activation) in the kidney of WT mice, while both of them decreased in Trpm2-KO animals [31]. The activated RAC1 interacts with TRPM2, which generates a possible positive feedback loop, resulting in TRPM2 membrane localization and greater oxidant injury [31, 37]. Treatment with NSC23766 as the RAC1 inhibitor before renal IR injury reduces NOX activation, PARP cleavage, and caspase-3 activity and increases the levels of anti-apoptotic proteins like Bcl-2 in mice subjected to IR injury [31].

ROS generated via activated NADPH oxidase causes DNA destruction and hence activates PARP, which functions as a modulator of cell death followed by IR injury [82]. The activated PARP increases the synthesis of PARP, which is converted into ADPR by PARG enzymes. ADPR stimulates Ca2+ influx through the TRPM2 channel and increases mitochondrial Ca2+ ion via NUDT9-H domain-TRPM2 C-terminal attachment [83]. The mitochondrial Ca2+ overload prompts the mitochondria’s swelling, destruction of the external membrane and the release of mitochondrial apoptosis factors into the cytosol, and induction of apoptosis [80, 84, 85]. PARP inhibitors restrict Ca2+ entrance through the TRPM2 channel by lowering ADPR synthesis, which is known to be the major activator of this channel [86].

TRPM2 channels are oxidative stress sensors, which means that oxidative stress either directly stimulates TRPM2 channels or increases cADPR levels [86]. The stimulation of TRPM2 channels by oxidative stress, as well as a rise in Ca2+ levels within the cell, indicated that it was linked to cell damage [38]. The TRPM2 blockade via ACA results in a decreased Ca2+ level within cells and consequently reduced cell death in H2O2 toxicity-exposed cells [68].

Intravenous injection of 2-APB as a TRPM2 inhibitor before ischemia protect the kidney against IR injury. This protective effect was shown by the reduction of apoptotic cells [87]. Cell apoptosis caused by nuclease, Ca2+-dependent phospholipase, and protease activation is recognized as one of the processes generating renal IR damage. Increased intracellular Ca2+ causes a rise in mitochondrial Ca2+ levels through the mitochondrial Ca2+ channels, and high mitochondrial Ca2+ load triggers irreversible processes that lead to cell apoptosis. 2-APB most likely protects the kidneys by reducing intracellular Ca2+ accumulation, which causes cell apoptosis following ischemia [87].

TRPM2 is activated by H2O2 under pathological circumstances like inflammation, oxidative stress and, cell death [88]. H2O2 also activates the TRPM2 channel directly and indirectly through NAD+-dependent ADPR production processes such as PARP activation, particularly PARP-1 and PARG enzymes in the nucleus [38]. H2O2 causes apoptosis by a variety of ways, including overexpression of Fas/Fas ligand, which activates the cell death pathway [89]. It also triggers the death of mitochondrial cells by modulating the mPTP [90]. H2O2 causes a rise in free Ca2+ inside cells, which leads to an increase in mitochondrial matrix Ca2+, which opens the mPTP [91]. When active, the mPTP decouples oxidative phosphorylation from ATP generation and promotes cytochrome c release into the cytosol. Cytochrome c attaches to Apaf-1(apoptotic peptidase activating factor 1) to create the apoptosome. After then, Caspase9, Caspase3, and Caspase7 are activated, leading to cell death [70].

TRPM2 and tissue damage in renal IR injury

Gao et al. investigated the function of the TRPM2 ion channel in kidney damage due to IR injury. Bilateral IR injury (28 min ischemia and 24 min reperfusion) caused histological damage to the kidney, comprising cast formation, an increase in the levels of blood urea nitrogen (BUN) and creatinine (Cr), and epithelial cell membrane damage in WT animal models while these effects were improved positively in TRPM2-KO animals [31]. TRPM2 deletion decreased renal dysfunction significantly, as demonstrated by lower serum Cr, BUN, and kidney injury molecule 1 (KIM-1) and lipocalin-2 (NGAL, a marker of kidney injury) levels [31, 57, 78]. Treatment with NSC23766 (RAC1 inhibitor), 8-Br‐cADPR, and 2-APB before ischemia, decreases ischemic damage in WT mice, while there was no additional amelioration in renal function in TRPM2-KO mice [78].

Conclusion

From this review it can be concluded that TRPM2 ion channels through various mechanisms such as ROS production, oxidative stress, Ca2+ overload, apoptosis, and inflammation are involved in AKI induced by IR injury. There is a crosstalk between the TRPM2 and its role in oxidative stress, inflammation, apoptosis, and inflammatory cells activation through Ca2+ overload.

The pharmacological inhibition of TRPM2 protects the kidney against IR damage. Therefore, the study of TRPM2 ion channels using their antagonists, agonists, modulators, as well as genetic deletion of TRPM2 and the use of animal models with manipulated TRPM2 channels can provide promising ideas for a better understanding of TRPM2 function under particular renal physiological and pathophysiological conditions and possibly a strategy for treating or even preventing the harmful effects of AKI caused by IR injury.