Alda-1, an Activator of ALDH2, Improves Postresuscitation Cardiac and Neurological Outcomes by Inhibiting Pyroptosis in Swine.

Aldehyde dehydrogenase 2 (ALDH2) has been proven to protect the heart and brain against regional ischemia/reperfusion injury, in which the protective role is related to the inhibition of pyroptosis. In the present study, we investigated whether an ALDH2 activator N-(1,3-benzodioxol-5-ylmethyl)-2,6-dichloro-benzamide (Alda-1) would improve postresuscitation cardiac and neurological outcomes in a clinically relevant swine model of cardiac arrest (CA) and resuscitation. The animal model was established by 8 min of untreated ventricular fibrillation and then 8 min of cardiopulmonary resuscitation (CPR). After restoring spontaneous circulation, the animals were randomly divided to receive either Alda-1 (0.88 mg/kg, n = 6) or saline (n = 5). Postresuscitation hemodynamic parameters, cardiac function, and cardiac and cerebral injuries were periodically measured for a total of 24 h. At 24 h postresuscitation, neurological function was evaluated, and then the animals were sacrificed, and cardiac and cerebral tissue samples were obtained for the measurements of oxidative stress, inflammation and pyroptosis. Consequently, postresuscitation cardiac and neurological dysfunction were significantly improved accompanied with significantly milder cardiac and cerebral injuries in the Alda-1 group compared with the CPR group. In addition, the increase in NLR family pyrin domain-containing 3 inflammasome expression and proinflammatory cytokine production, which indicated the occurrence of inflammatory response, were significantly less in the Alda1 group than in the CPR group. The expression level of gasdermin D used as a protein marker of pyroptosis was also significantly reduced in all resuscitated animals receiving Alda1 treatment. Moreover, the severity of oxidative stress indicated by the changes of 4-hydroxy-2-nonenal and malondialdehyde was significantly decreased in the heart and brain in all animals treated with Alda-1 compared to the CPR group. Thus, Alda-1 mitigated postresuscitation cardiac and neurological dysfunction and injuries possibly by inhibiting oxidative stress-mediated NLRP3 inflammasome activation and pyroptosis in a swine model of CA and resuscitation.

Mitochondrial aldehyde dehydrogenase (ALDH2) rescues cardiac contractile dysfunction in an APP/PS1 murine model of Alzheimer's disease via inhibition of ACSL4-dependent ferroptosis.

Alzheimer's disease (AD) is associated with high incidence of cardiovascular events but the mechanism remains elusive. Our previous study reveals a tight correlation between cardiac dysfunction and low mitochondrial aldehyde dehydrogenase (ALDH2) activity in elderly AD patients. In the present study we investigated the effect of ALDH2 overexpression on cardiac function in APP/PS1 mouse model of AD. Global ALDH2 transgenic mice were crossed with APP/PS1 mutant mice to generate the ALDH2-APP/PS1 mutant mice. Cognitive function, cardiac contractile, and morphological properties were assessed. We showed that APP/PS1 mice displayed significant cognitive deficit in Morris water maze test, myocardial ultrastructural, geometric (cardiac atrophy, interstitial fibrosis) and functional (reduced fractional shortening and cardiomyocyte contraction) anomalies along with oxidative stress, apoptosis, and inflammation in myocardium. ALDH2 transgene significantly attenuated or mitigated these anomalies. We also noted the markedly elevated levels of lipid peroxidation, the essential lipid peroxidation enzyme acyl-CoA synthetase long-chain family member 4 (ACSL4), the transcriptional regulator for ACLS4 special protein 1 (SP1) and ferroptosis, evidenced by elevated NCOA4, decreased GPx4, and SLC7A11 in myocardium of APP/PS1 mutant mice; these effects were nullified by ALDH2 transgene. In cardiomyocytes isolated from WT mice and in H9C2 myoblasts in vitro, application of Aß (20 µM) decreased cell survival, compromised cardiomyocyte contractile function, and induced lipid peroxidation; ALDH2 transgene or activator Alda-1 rescued Aß-induced deteriorating effects. ALDH2-induced protection against Aß-induced lipid peroxidation was mimicked by the SP1 inhibitor tolfenamic acid (TA) or the ACSL4 inhibitor triacsin C (TC), and mitigated by the lipid peroxidation inducer 5-hydroxyeicosatetraenoic acid (5-HETE) or the ferroptosis inducer erastin. These results demonstrate an essential role for ALDH2 in AD-induced cardiac anomalies through regulation of lipid peroxidation and ferroptosis.

Aldehyde Dehydrogenase 2 Activator Augments the Beneficial Effects of Empagliflozin in Mice with Diabetes-Associated HFpEF.

To ameliorate diabetes mellitus-associated heart failure with preserved ejection fraction (HFpEF), we plan to lower diabetes-mediated oxidative stress-induced 4-hydroxy-2-nonenal (4HNE) accumulation by pharmacological agents that either decrease 4HNE generation or increase its detoxification.A cellular reactive carbonyl species (RCS), 4HNE, was significantly increased in diabetic hearts due to a diabetes-induced decrease in 4HNE detoxification by aldehyde dehydrogenase (ALDH) 2, a cardiac mitochondrial enzyme that metabolizes 4HNE. Therefore, hyperglycemia-induced 4HNE is critical for diabetes-mediated cardiotoxicity and we hypothesize that lowering 4HNE ameliorates diabetes-associated HFpEF. We fed a high-fat diet to ALDH2*2 mice, which have intrinsically low ALDH2 activity, to induce type-2 diabetes. After 4 months of diabetes, the mice exhibited features of HFpEF along with increased 4HNE adducts, and we treated them with vehicle, empagliflozin (EMP) (3 mg/kg/d) to reduce 4HNE and Alda-1 (10 mg/kg/d), and ALDH2 activator to enhance ALDH2 activity as well as a combination of EMP + Alda-1 (E + A), via subcutaneous osmotic pumps. After 2 months of treatments, cardiac function was assessed by conscious echocardiography before and after exercise stress. EMP + Alda-1 improved exercise tolerance, diastolic and systolic function, 4HNE detoxification and cardiac liver kinase B1 (LKB1)-AMP-activated protein kinase (AMPK) pathways in ALDH2*2 mice with diabetes-associated HFpEF. This combination was even more effective than EMP alone. Our data indicate that ALDH2 activation along with the treatment of hypoglycemic agents may be a salient strategy to alleviate diabetes-associated HFpEF.

A role for aldehyde dehydrogenase (ALDH) 2 in angiotensin II-mediated decrease in angiogenesis of coronary endothelial cells.

Diabetes-induced coronary endothelial cell (CEC) dysfunction contributes to diabetic heart diseases. Angiotensin II (Ang II), a vasoactive hormone, is upregulated in diabetes, and is reported to increase oxidative stress in CECs. 4-hydroxy-2-nonenal (4HNE), a key lipid peroxidation product, causes cellular dysfunction by forming adducts with proteins. By detoxifying 4HNE, aldehyde dehydrogenase (ALDH) 2 reduces 4HNE mediated proteotoxicity and confers cytoprotection. Thus, we hypothesize that ALDH2 improves Ang II-mediated defective CEC angiogenesis by decreasing 4HNE-mediated cytotoxicity. To test our hypothesis, we treated the cultured mouse CECs (MCECs) with Ang II (0.1, 1 and 10 µM) for 2, 4 and 6 h. Next, we treated MCECs with Alda-1 (10 µM), an ALDH2 activator or disulfiram (2.5 µM)/ALDH2 siRNA (1.25 nM), the ALDH2 inhibitors, or blockers of angiotensin II type-1 and 2 receptors i.e. Losartan and PD0123319 respectively before challenging MCECs with 10 µM Ang II. We found that 10 µM Ang II decreased tube formation in MCECs with in vitro angiogenesis assay (P < .0005 vs control). 10 µM Ang II downregulated the levels of vascular endothelial growth factor receptor 1 (VEGFR1) (p < .005 for mRNA and P < .05 for protein) and VEGFR2 (p < .05 for mRNA and P < .005 for protein) as well as upregulated the levels of angiotensin II type-2 receptor (AT2R) (p < .05 for mRNA and P < .005 for protein) and 4HNE-adducts (P < .05 for protein) in cultured MCECs, compared to controls. ALDH2 inhibition with disulfiram/ALDH2 siRNA exacerbated 10 µM Ang II-induced decrease in coronary angiogenesis (P < .005) by decreasing the levels of VEGFR1 (P < .005 for mRNA and P < .05 for protein) and VEGFR2 (P < .05 for both mRNA and protein) and increasing the levels of AT2R (P < .05 for both mRNA and protein) and 4HNE-adducts (P < .05 for protein) relative to Ang II alone. AT2R inhibition per se improved angiogenesis in MCECs. Additionally, enhancing ALDH2 activity with Alda 1 rescued Ang II-induced decrease in angiogenesis by increasing the levels of VEGFR1, VEGFR2 and decreasing the levels of AT2R. In summary, ALDH2 can be an important target in reducing 4HNE-induced proteotoxicity and improving angiogenesis in MCECs. Finally, we conclude ALDH2 activation can be a therapeutic strategy to improve coronary angiogenesis to ameliorate cardiometabolic diseases.

A small-molecule activator of mitochondrial aldehyde dehydrogenase 2 reduces the severity of cerulein-induced acute pancreatitis.

Acute pancreatitis (AP) is one of the leading causes of hospital admission for gastrointestinal disorders. Although lipid peroxides are produced in AP, it is unknown if targeting lipid peroxides prevents AP. This study aimed to investigate the role of mitochondrial aldehyde dehydrogenase 2 (ALDH2), a critical enzyme for lipid peroxide degradation, in AP and the possible underlying mechanisms. Cerulein was used to induce AP in C57BL/6 J male mice and pancreatic acinar cells were used to elucidate underlying mechanisms in vitro. Pancreatic enzymes in the serum, lipid peroxidation products malondialdehyde (MDA) and 4-hydroxynonenal (4-HNE), and Bcl-2, Bax and cleaved caspase-3 were measured. ALDH2 activation with a small-molecule activator, Alda-1, reduced the levels of the pancreatic enzymes in the serum and the lipid peroxidation products MDA and 4-HNE. In addition, Alda-1 decreased Bax and cleaved caspase-3 expression and increased Bcl-2 expression in vivo and in vitro. In conclusion, ALDH2 activation by Alda-1 has a protective effect in cerulein-induced AP by mitigating apoptosis in pancreatic acinar cells by alleviating lipid peroxidation.

Aldehyde dehydrogenase 2 protects against sympathetic excitation-induced cardiac fibrosis.

Sympathetic stimulated-cardiac fibrosis imposes great significance on both disease progression and survival in the pathogenesis of many cardiovascular diseases. However, there are few effective therapies targeting it clinically. The cardioprotective effect of aldehyde dehydrogenase 2 (ALDH2) has been explored in many pathological conditions, whether it can exert benefit effects on chronic sympathetic stimulus-induced cardiac fibrosis remains unclear. In this study, we determined to explore the role of ALDH2 on isoproterenol (ISO)-induced cardiac fibroblasts (CF) proliferation and cardiac fibrosis. It was found that ALDH2 enzymatic activity was impaired in ISO-induced HCF proliferation and Aldh2 deficiency promoted mouse CF proliferation. Alda-1, an ALDH2 activator, exerted obvious suppressive effect on ISO-induced HCF proliferation, together with the induction of cell cycle arrest at G0/G1 phase and decreased expression of cyclin E1 and cyclin-dependent kinase 2 (CDK2). Mechanistically, the inhibitory role of Alda-1 on HCF proliferation was achieved by decreasing mitochondrial reactive oxygen species (ROS) production, which was partially reversed by rotenone, an inducer of ROS. In addition, wild-type mice treated with Alda-1 manifested with reduced fibrosis and better cardiac function after ISO pump. In summary, Alda-1 alleviates sympathetic excitation-induced cardiac fibrosis via decreasing mitochondrial ROS accumulation, highlighting ALDH2 activity as a promising drug target of cardiac fibrosis.

Aldehyde dehydrogenase-2 activation decreases acetaminophen hepatotoxicity by prevention of mitochondrial depolarization.

Oxidative stress contributes to acetaminophen (APAP) hepatotoxicity. Since lipid peroxidation produces reactive aldehydes, we investigated whether activation of mitochondrial aldehyde dehydrogenase-2 (ALDH2) with Alda-1 decreases liver injury after APAP. Male C57BL/6 mice fasted overnight received Alda-1 (20 mg/kg, i.p.) or vehicle 30 min before APAP (300 mg/kg, i.p.). Blood and livers were collected 2 or 24 h after APAP. Intravital multiphoton microscopy of rhodamine 123 (Rh123) and propidium iodide (PI) fluorescence was conducted 6 h after APAP administration to detect mitochondrial polarization status and cell death. 4-Hydroxynonenal protein adducts were present in 0.1% of tissue area without APAP treatment but increased to 7% 2 h after APAP treatment, which Alda-1 blunted to 1%. Serum alanine and aspartate aminotransferases increased to 7594 and 9768 U/L at 24 h respectively, which decreased ≥72% by Alda-1. Alda-1 also decreased centrilobular necrosis at 24 h after APAP from 47% of lobular areas to 21%. N-acetyl-p-benzoquinone imine protein adduct formation and c-Jun-N-terminal kinase phosphorylation increased after APAP as expected, but Alda-1 did not alter these changes. Without APAP, no mitochondrial depolarization was detected by intravital microscopy. At 6 h after APAP, 62% of tissue area showed depolarization, which decreased to 33.5% with Alda-1. Cell death as detected by PI labeling increased from 0 to 6.8 cells per 30× field 6 h after APAP, which decreased to 0.6 cells by Alda-1. In conclusion, aldehydes are important mediators of APAP hepatotoxicity. Accelerated aldehyde degradation by ALDH2 activation with Alda-1 decreases APAP hepatotoxicity by protection against mitochondrial dysfunction.

Design, synthesis and the structure-activity relationship of agonists targeting on the ALDH2 catalytic tunnel.

ALDH2, a key enzyme in the alcohol metabolism process, detoxifies several kinds of toxic small molecular aldehydes, which induce severe organ damages. The development of novel Alda-1 type ALDH2 activators was mostly relied on HTS but not rational design so far. To clarify the structure-activity relationship (SAR) of the skeleton of Alda-1 analogs by synthesis of the least number of analogs, we prepared 31 Alda-1 analogs and 3 isoflavone derivatives and evaluated for their ALDH2-activating activity. Among these, the ALDH2-activating activity of mono-halogen-substituted (Cl and Br) N-piperonylbenzamides 3b and 3 k, and non-aromatic amides 8a-8c, were 1.5-2.1 folds higher than that of Alda-1 at 20 µM. The relationship between binding affinity in computer aided molecular docking model and the ALDH2-activating activity assays were clarified as follows: for Alda-1 analogs, with the formation of halogen bonds, the enzyme-activating activity was found to follow a specific regression curve within the range between -5 kcal/mol and -4 kcal/mol. For isoflavone derivatives, the basic moiety on the B ring enhance the activating activity. These results provide a new direction of utilizing computer-aided modeling to design novel ALDH2 agonists in the future.

Alda-1, an Aldehyde Dehydrogenase 2 Agonist, Improves Cutaneous Wound Healing by Activating Epidermal Keratinocytes via Akt/GSK-3ß/ß-Catenin Pathway.

BACKGROUND: The cutaneous wound healing process mainly comprises re-epithelialization, fibrosis, and neovascularization. Impaired wound healing is common but tricky in plastic surgery. Aldehyde dehydrogenase 2 (ALDH2), the most effective subset of the ALDH enzyme family, is known to exert a major role in detoxification of aldehydes. Activation of ALDH2 by Alda-1 (a specific agonist) has been found to protect against cardiovascular diseases. However, no research has paid attention to the potential of ALDH2 activation in regulating wound healing. The previous studies suggested a high expression of ALDH2 in normal skin tissue. The aim of this study was to investigate if Alda-1 may ameliorate wound healing. METHODS: A full-thickness excisional wound model was established in vivo. Adult male C57BL/6 mice were randomly divided into DMSO and Alda-1 groups. Mice received an intraperitoneal injection of DMSO or 10 mg/mL Alda-1 (10 mg/kg body weight, dissolved in DMSO) for 7 days. The wound healing rate was measured at 0, 3, 5, and 7 days. Distribution of ALDH2 in wound tissue was showed. ALDH2 enzymatic activity was examined at 3, 5, and 7 days. The elongation of epithelial tongue was detected by hematoxylin-eosin staining, and collagen deposition was analyzed by Masson's trichrome staining at 7 days. Expressions of alpha-smooth muscle actin (alpha-SMA), transforming growth factor beta (TGF-beta), CD31, collagen 1, collagen 3, and elastin were stained by immunohistochemistry at 5 and 7 days. The HaCaT cell line was applied in vitro. Proliferation and migration were tested using CCK8 and wound healing assay separately. The level of TGF-ß was examined by ELISA. Protein levels of the Akt/glycogen synthase kinase-3 beta (GSK-3 beta)/beta-catenin pathway were determined by western blotting. RESULTS: Alda-1 accelerated wound healing rates. ALDH2 activity in wound sites was restored. Alda-1 promoted the length of the epithelial tongue, collagen deposition, as well as expressions of alpha-SMA, TGF-beta, collagen 1/3, elastin, but did not affect CD31. Proliferation, migration, and TGF-ß secretion were promoted by Alda-1 and deregulated by CVT-10216 (an ALDH2 inhibitor). Protein variations of the Akt/GSK-3ß/ß-catenin pathway were found to accord with ALDH2 changes. CONCLUSIONS: Alda-1, an ALDH2 agonist, improves cutaneous wound healing in a full-thickness excisional wound model. Alda-1 activates proliferation, migration, and TGF-ß secretion of HaCaT (epidermal keratinocytes) by regulating the Akt/GSK-3ß/ß-catenin pathway. NO LEVEL ASSIGNED: This journal requires that authors assign a level of evidence to each submission to which Evidence-Based Medicine rankings are applicable. This excludes Review Articles, Book Reviews, and manuscripts that concern Basic Science, Animal Studies, Cadaver Studies, and Experimental Studies. For a full description of these Evidence-Based Medicine ratings, please refer to the Table of Contents or the online Instructions to Authors www.springer.com/00266.

Aldehyde dehydrogenase 2 activation ameliorates CCl4 -induced chronic liver fibrosis in mice by up-regulating Nrf2/HO-1 antioxidant pathway.

Mitochondrial aldehyde dehydrogenase 2 (ALDH2) is critical in the pathogenesis of alcoholic liver cirrhosis. However, the effect of ALHD2 on liver fibrosis remains to be further elucidated. This study aimed to demonstrate whether ALDH2 regulates carbon tetrachloride (CCl4 )-induced liver fibrosis and to investigate the efficacy of Alda-1, a specific activator of ALDH2, on attenuating liver fibrosis. ALDH2 expression was increased after chronic CCl4 exposure. ALDH2 deficiency accentuated CCl4 -induced liver fibrosis in mice, accompanied by increased expression of collagen 1α1, α-SMA and TIMP-1. Moreover, ALDH2 knockout triggered more ROS generation, hepatocyte apoptosis and impaired mitophagy after CCl4 treatment. In cultured HSC-T6 cells, ALDH2 knockdown by transfecting with lentivirus vector increased ROS generation and α-SMA expression in an in vitro hepatocyte fibrosis model using TGF-ß1. ALDH2 overexpression by lentivirus or activation by Alda-1 administration partly reversed the effect of TGF-ß1, whereas ALDH2 knockdown totally blocked the protective effect of Alda-1. Furthermore, Alda-1 administration protected against liver fibrosis in vivo, which might be mediated through up-regulation of Nrf2/HO-1 cascade and activation of Parkin-related mitophagy. These findings indicate that ALDH2 deficiency aggravated CCl4 -induced hepatic fibrosis through ROS overproduction, increased apoptosis and mitochondrial damage, whereas ALDH2 activation through Alda-1 administration alleviated hepatic fibrosis partly through activation of the Nrf2/HO-1 antioxidant pathway and Parkin-related mitophagy, which indicate ALDH2 as a promising anti-fibrotic target and Alda-1 as a potential therapeutic agent in treating CCl4 -induced liver fibrosis.

Alda-1, an aldehyde dehydrogenase-2 agonist, causes deterioration in renal functions following ischemia-reperfusion injury due to crystalline nephropathy.

Renal ischemia-reperfusion injury (IRI) induces the production of aldehydes which are detoxified by aldehyde dehydrogenases (ALDHs). Alda-1 is a selective ALDH2 agonist and its protective effect was demonstrated in several conditions. The effect of Alda-1 on the kidney or on renal IRI was not investigated. We investigated the effect of Alda-1 on the renal dysfunction following IRI. Wistar rats underwent left IRI for 40 min. Group-Alda (n = 11) received Alda-1 starting 24 h before IRI and continued for 7 days thereafter when renal functions were measured. Group-Vx (n = 11) underwent similar protocol but received the dissolvent. Alda-1 did not affect renal blood flow or glomerular filtration rate in the left ischemic kidney in Group-Alda compared to Group-Vx (3.05 ± 0.50 vs. 3.53 ± 0.70, and 0.40 ± 0.06 vs. 0.51 ± 0.08, respectively, p > .05 for both). However, left renal fractional sodium excretion was higher in Group-Alda (2.80 ± 0.43 vs. 1.37 ± 0.36, p = .02). Alda-1 also adversely affected the gene expressions of kidney injury molecule-1 and neutrophil gelatinase-associated lipocalin (217 ± 38 vs. 99 ± 13 and 49 ± 13 vs. 20 ± 5, respectively, p < .05 for both) and the alterations in tumor necrosis factor-α, transforming growth factor-ß1, plasminogen activator inhibitor-1, fibronectin 1 and p53 (4.4 ± 0.9 vs. 2.1 ± 0.3, 1.5 ± 0.1 vs. 1.1 ± 0.1, 30.0 ± 2.7 vs. 11.7 ± 2.3, 3.6 ± 0.4 vs. 2.1 ± 0.2 and 1.3 ± 0.1 vs. 0.9 ± 0.07, respectively, p ≤ .05 for all). This was associated with intratubular crystal deposition suggestive of crystalline nephropathy. Alda-1 exacerbated the IRI-induced renal tubular dysfunction and alterations in markers of acute kidney injury, biomarkers of inflammation, fibrosis and apoptosis and this was associated with intratubular crystal deposition suggestive of crystalline nephropathy.

Pharmacological activation of aldehyde dehydrogenase 2 promotes osteoblast differentiation via bone morphogenetic protein-2 and induces bone anabolic effect.

Aldehyde dehydrogenases (ALDHs) are a family of enzymes involved in detoxifying aldehydes. Previously, we reported that an ALDH inhibitor, disulfiram caused bone loss in rats and among ALDHs, osteoblast expressed only ALDH2. Loss-of-function mutation in ALDH2 gene is reported to cause bone loss in humans which suggested its importance in skeletal homeostasis. We thus studied whether activating ALDH2 by N-(1, 3-benzodioxol-5-ylmethyl)-2, 6-dichlorobenzamide (alda-1) had osteogenic effect. We found that alda-1 increased and acetaldehyde decreased the differentiation of rat primary osteoblasts and expressions of ALDH2 and bone morphogenetic protein-2 (BMP-2). Silencing ALDH2 in osteoblasts abolished the alda-1 effects. Further, alda-1 attenuated the acetaldehyde-induced lipid-peroxidation and oxidative stress. BMP-2 is essential for bone regeneration and alda-1 increased its expression in osteoblasts. We then showed that alda-1 (40mg/kg dose) augmented bone regeneration at the fracture site with concomitant increase in BMP-2 protein compared with control. The osteogenic dose (40mg/kg) of alda-1 attained a bone marrow concentration that was stimulatory for osteoblast differentiation, suggesting that the tissue concentration of alda-1 matched its pharmacologic effect. In addition, alda-1 promoted modeling-directed bone growth and peak bone mass achievement, and increased bone mass in adult rats which reiterated its osteogenic effect. In osteopenic ovariectomized (OVX) rats, alda-1 reversed trabecular osteopenia with attendant increase in serum osteogenic marker (procollagen type I N-terminal peptide) and decrease in oxidative stress. Alda-1 has no effect on liver and kidney function. We conclude that activating ALDH2 by alda-1 had an osteoanabolic effect involving increased osteoblastic BMP-2 production and decreased OVX-induced oxidative stress.

Pharmacological enrollment of aldehyde dehydrogenase modulators to assist treating ischemia reperfusion-induced intestinal injury: is there a gap to be bridged?

This commentary highlights the research presented by Zhu et al. [1]. In this issue of the Clinical Science, the authors evaluated the protective effect of Alda-1 (a novel class of small molecule aldehyde dehydrogenase (ALDH2) activators) in the intestinal ischemia reperfusion (IR) injury. Remarkably, enhancing the ADLH2 activity by the use of Alda-1 can ameliorate several deleterious effects related to aldehydes, and may provide a better protection against an injury preestablished by IR. Together, an innovative metabolic strategy for treating patients with IR injury could be the use of ALDH modulators in a near future.

Pretreatment with the ALDH2 agonist Alda-1 reduces intestinal injury induced by ischaemia and reperfusion in mice.

Many studies demonstrate that activation of aldehyde dehydrogenase 2 (ALDH2) protects against oxidative stress via detoxification of cytotoxic aldehydes, and could attenuate cardiac, cerebral, lung and renal ischaemia-reperfusion (I/R) injuries. However, the effect of ALDH2 in intestinal I/R is unknown. The present study was set up to determine whether an ALDH2 agonist, Alda-1, could alleviate intestinal injury after gut I/R. In a mouse model of intestinal I/R injury, histological grading, proinflammatory cytokines, oxidative stress, cellular apoptosis, chemokine contents, ALDH2 activity, 4-hydroxy-trans-2-nonenal (4-HNE) and malondialdehyde (MDA) were evaluated. The results indicated that I/R treatment conferred elevation in pathological scores, proinflammatory cytokines, oxidative stress, cellular apoptosis and chemokine levels, accompanied by accumulated 4-HNE and MDA. No significant changes in ALDH2 activity were observed after I/R. However, Alda-1 pretreatment significantly decreased these injurious indicators, concomitant with up-regulated ALDH2 activity, and lessened 4-HNE and MDA accumulation. Taken together, our results implicate activation of ALDH2 by Alda-1 in the significant abatement intestinal I/R injury.

The impact of mitochondrial aldehyde dehydrogenase (ALDH2) activation by Alda-1 on the behavioral and biochemical disturbances in animal model of depression.

The etiology of depression remains still unclear. Recently, it has been proposed, that mitochondrial dysfunction may be associated with development of mood disorders, such as depression, bipolar disorder and anxiety disorders. Mitochondrial aldehyde dehydrogenase (ALDH2), an enzyme responsible for the detoxification of reactive aldehydes, is considered to exert protective function in mitochondria. We investigated the influence of Alda-1, a small-molecule activator of ALDH2, on depressive- and anxiety-like behaviors in an animal model of depression - the prenatally stressed rats - using behavioral, molecular and proteomic methods. Prolonged Alda-1 administration significantly increased the climbing time, tended to reduce the immobility time and increased the swimming time of the prenatally stressed rats in the forced swim test. Moreover, treatment of prenatally stressed rats with Alda-1 significantly increased number of entries into the open arms of the maze and the time spent therein, as assessed by elevated plus-maze test. Such actions were associated with reduction of plasma 4-HNE-protein content, decrease of TNF-α mRNA and increase of PGC-1α (regulator of mitochondrial biogenesis) mRNA level in the frontal cortex and hippocampus of the prenatally stressed rats as well as with normalization of peripheral immune parameters and significant changes in expression of 6 and 4 proteins related to mitochondrial functions in the frontal cortex and hippocampus, respectively. Collectively, ALDH2 activation by Alda-1 led to a significant attenuation of depressive- and anxiety-like behaviors in the prenatally stressed rats. The pattern of changes suggested mitoprotective effect of Alda-1, however the exact functional consequences of the revealed alterations require further investigation.

Pharmacological activation of aldehyde dehydrogenase 2 by Alda-1 reverses alcohol-induced hepatic steatosis and cell death in mice.

BACKGROUND & AIMS: Effective therapies for alcoholic liver disease are currently unavailable. The present study tested the efficacy of Alda-1, a specific aldehyde dehydrogenase 2 (ALDH2) activator, in treating alcoholic liver disease. METHODS: Male C57BL/6J mice were exposed to alcohol for a time-course study on aldehyde metabolism. The specificity and efficacy of Alda-1 on activating hepatic ALDH2 and aldehyde clearance were determined by acute treatments. Then, mice were fed alcohol for 8 weeks with Alda-1 administration for the last 10 days to test the therapeutic potential of Alda-1. Lastly, H4IIEC3 cells were treated with ethanol, acetaldehyde, or 4-hydroxynonenal to define the link between aldehydes and hepatotoxicity. RESULTS: Alcohol feeding for 8 weeks induced hepatic ALDH2 dysfunction and aldehyde accumulation. One dose of Alda-1 administration elevated hepatic ALDH activity, which was blocked by the specific ALDH2 inhibitor, daidzin. Alda-1 accelerated acetaldehyde clearance after acute alcohol intoxication. Alda-1 treatment in the 8-week alcohol feeding model reversed liver damage along with reduction of hepatic aldehydes. Alda-1 re-activated transcription factors, upregulated fatty acid oxidation enzymes, and reversed steatosis. Alcohol-induced endoplasmic reticulum stress and apoptotic cell death were also attenuated by Alda-1. Acetaldehyde or 4-hydroxynonenal treatment to H4IIEC3 cells inactivated transcription factors and induced endoplasmic reticulum stress and apoptosis, while ethanol per se showed limited effects. CONCLUSIONS: Pharmacological activation of ALDH2 by Alda-1 reversed alcoholic steatosis and apoptosis through accelerating aldehyde clearance. This study indicates that ALDH2 is a promising molecular target and Alda-1 has therapeutic potential for treating alcoholic liver disease.

The administration of Alda-1, an activator of ALDH2, inhibits relapse-like ethanol intake in female alcohol-preferring UChB rats.

AIMS: Alcohol relapse is a main limitation for the treatment of alcohol use disorders. Previous studies have shown that Alda-1, a pharmacological activator of ALDH2, inhibits both acquisition and chronic ethanol intake in rats; however, its effects on relapse-like ethanol intake are unknown. The aim of this study was to assess the effect of Alda-1 on post-deprivation and reaccess relapse-like ethanol intake in alcohol-preferring UChB rats. We also aimed to assess the possible mechanisms associated with the effects of Alda-1 by measuring the levels of glutamate transporter (GLT-1), oxidative stress and neuroinflammation markers in different regions of the mesocorticolimbic system. MAIN METHODS: In Experiment I, UChB female rats were exposed for 100 days to voluntary ethanol intake followed by 2-weeks of ethanol withdrawal and 1 week of ethanol reaccess. Alda-1 (25 mg/kg, intragastric, i.g) or vehicle was administered daily for 14 days during the withdrawal/re-access period. Experiment II was similar to Experiment I, but after the withdrawal period, ethanol re-access was not allowed, and Alda-1 was administered during the last week of withdrawal. At the end of both experiments, the levels of GLT-1, oxidative stress (GSH, MDA), and neuroinflammation markers (GFAP, Iba-1) were assessed in nucleus accumbens and/or hippocampus. KEY FINDINGS: The results showed that Alda-1 administration markedly blocked (90 %, p < 0.001) relapse-like ethanol intake in UChB rats. Alda-1 increased Iba-1 reactivity (microglial marker) in the NAc of ethanol-deprived rats. Alda-1 administration did not influence the levels of GLT-1, oxidative stress markers (MDA, GSH) or GFAP reactivity in the mesocorticolimbic system. SIGNIFICANCE: These preclinical findings support the use of activators of ALDH2, such as Alda-1, as a potential pharmacological strategy in the treatment of alcohol relapse.

Alda-1 ameliorates air embolism-induced acute lung injury.

OBJECTIVE: Evidence suggests that aldehyde dehydrogenase 2 (ALDH2) offers protection against damage caused by oxidative stress in diverse rodent models. Nonetheless, the effect of Alda-1, a compound that activates ALDH2, on acute lung injury (ALI) induced by air embolism (AE) remains unclear. The objective of this study was to explore the protective effects of Alda-1 in ALI induced by AE. METHODS: A rat model of in situ isolated perfused lung was established to investigate AE-induced ALI. Air was infused into the pulmonary artery at 0.25 mL/min for 1 minute. Before inducing AE, different doses (10, 20, or 30 mg/kg) of Alda-1 were given through intraperitoneal injection. Pathological changes in lung tissue were assessed using hematoxylin-eosin staining. We performed Western blot analysis to assess the protein levels of ALDH2,4-hydroxy-trans-2-nonenal (4-HNE), Bcl-2, caspase-3, phosphatidylinositol 3-kinase (PI3K), Akt, IκB-α, and nuclear NF-κB. RESULTS: Notably, AE results were demonstrated as harmful to the lungs, which is evidenced by intensified lung edema and disruption of lung tissue structure. Furthermore, AE caused a decrease in ALDH2 expression, increased accumulation of 4-HNE and MDA, infiltration of neutrophils, increased production of inflammatory cytokines, apoptosis, and upregulation of the PI3K/Akt and NF-κB signaling pathways within the lungs. Administration of a 20 mg/kg dose of Alda-1 alleviated the detrimental effects induced by AE. CONCLUSION: Alda-1 shows promise in mitigating AE-induced ALI, possibly through the upregulation of ALDH2 expression and suppression of the PI3K/Akt and NF-κB signaling pathways. Further research is warranted to validate these findings and to explore their translational potential in human subjects.

ALDH2 inhibition by lead and ethanol elicits redox imbalance and mitochondrial dysfunction in SH-SY5Y human neuroblastoma cell line: Reversion by Alda-1.

Lead (Pb), a common environmental contaminant, and ethanol (EtOH), a widely available drug of abuse, are well-known neurotoxicants. In vivo, experimental evidence indicates that Pb exposure affects oxidative EtOH metabolism with a high impact on living organisms. On these bases, we evaluated the consequences of combined Pb and EtOH exposure on aldehyde dehydrogenase 2 (ALDH2) functionality. In vitro exposure to 10 µM Pb, 200 mM EtOH, or their combination for 24 h reduced ALDH2 activity and content in SH-SY5Y human neuroblastoma cells. In this scenario, we observed mitochondrial dysfunction characterized by reduced mass and membrane potential, decreased maximal respiration, and spare capacity. We also evaluated the oxidative balance in these cells finding a significant increase in reactive oxygen species (ROS) production and lipid peroxidation products under all treatments accompanied by an increase in catalase (CAT) activity and content. These data suggest that ALDH2 inhibition induces the activation of converging cytotoxic mechanisms resulting in an interplay between mitochondrial dysfunction and oxidative stress. Notably, NAD+ (1 mM for 24 h) restored ALDH2 activity in all groups, while an ALDH2 enhancer (Alda-1, 20 µM for 24 h) also reversed some of the deleterious effects resulting from impaired ALDH2 function. Overall, these results reveal the crucial role of this enzyme on the Pb and EtOH interaction and the potential of activators such as Alda-1 as therapeutic approaches against several conditions involving aldehydes accumulation.

Impact of common ALDH2 inactivating mutation and alcohol consumption on Alzheimer's disease.

Aldehyde dehydrogenase 2 (ALDH2) is an enzyme found in the mitochondrial matrix that plays a central role in alcohol and aldehyde metabolism. A common ALDH2 polymorphism in East Asians descent (called ALDH2*2 or E504K missense variant, SNP ID: rs671), present in approximately 8% of the world's population, has been associated with a variety of diseases. Recent meta-analyses support the relationship between this ALDH2 polymorphism and Alzheimer's disease (AD). And AD-like pathology observed in ALDH2-/- null mice and ALDH2*2 overexpressing transgenic mice indicate that ALDH2 deficiency plays an important role in the pathogenesis of AD. Recently, the worldwide increase in alcohol consumption has drawn attention to the relationship between heavy alcohol consumption and AD. Of potential clinical significance, chronic administration of alcohol in ALDH2*2/*2 knock-in mice exacerbates the pathogenesis of AD-like symptoms. Therefore, ALDH2 polymorphism and alcohol consumption likely play an important role in the onset and progression of AD. Here, we review the data on the relationship between ALDH2 polymorphism, alcohol, and AD, and summarize what is currently known about the role of the common ALDH2 inactivating mutation, ALDH2*2, and alcohol in the onset and progression of AD.