PM2.5 toxicity in blood impairs cardiac redox balance and promotes mitochondrial dysfunction in rat heart that further aggravates ischemia reperfusion injury by modulating PI3K/AKT/mTOR/NF-kB signaling axis.

According to the pathophysiological mechanisms linking particulate matter (PM2.5) exposure and cardiovascular diseases, PM2.5 may directly translocate into the blood stream and remote target organs and thereby induce cardiovascular effects. The toxicity of PM2.5 is known to induce oxidative stress in pulmonary tissue, but its impact on the redox state in heart (distant organ) is unknown and how it modulates the cardiac response to ischemia reperfusion (IR) remains unclear. In the present study, we evaluated the toxic effect of PM2.5 on cardiac physiology in the presence and absence of IR after introducing PM2.5 into the blood. Female Wistar rats were injected with diesel particulate matter (DPM) via i.p & i.v routes at a concentration of 10 µg/ml. The toxic impact of PM2.5 not only adversely affects the cardiac ultra-structure (leading to nuclear infiltration, edema, irregularities in heart muscle and nuclear infiltration), but also altered the cellular redox balance, elevated inflammation and promoted the upregulation of proapoptotic mediator genes at the basal level of myocardium. The results showed alterations in cardiac ultrastructure, elevated oxidative stress and significant redox imbalance, increased inflammation and proapoptotic mediators at the basal level of myocardium. Moreover, the cardioprotective pro survival signaling axis was declined along with an increased NF-kB activation at the basal level. IR inflicted further injury with deterioration of cardiac hemodynamic indices (Heart rate [HR], Left ventricular developed pressure [LVDP], Left ventricular end-diastolic pressure [LVEDP] and rate pressure product [RPP]) along with prominent inactivation of signaling pathways. Furthermore, the levels of GSH/GSSG, NADH/NAD, NADPH/NADP were significantly low along with increased lipid peroxidation in mitochondria of PM2.5 treated IR rat hearts. This observation was supported by downregulation of glutaredoxin and peroxiredoxin genes in the myocardium. Similarly the presence of oxidative stress inducing metals was found at a higher concentration in cardiac mitochondria. Thus, the toxic impact of PM2.5 in heart augment the IR associated pathological changes by altering the physiological response, initiating cellular metabolic alterations in mitochondria and modifying the signaling molecules.

Investigating the temporal link between PM2.5 exposure and acceleration of myocardial ischemia reperfusion injury: Emphasizing the hazardous presence of metals in inhaled air.

Exposure to PM2.5 is widely acknowledged to induce cardiotoxic effects, leading to decreased myocardial tolerance to revascularization procedures and subsequent ischemia reperfusion injury (IR). However, the temporal relationship between PM2.5 exposure and vulnerability to IR, along with the underlying mechanisms, remains unclear and is the focus of this study. Female Wistar rats were exposed to PM2.5 at a concentration of 250 µg/m³ for 3 h daily over varying durations (7, 14, and 21 days), followed by IR induction. Our results demonstrated a significant increase in cardiac injury, as evidenced by increased infarct size and elevated cardiac injury markers, starting from day 14 of PM2.5 exposure, accompanied by declined cardiac function. These adverse effects were associated with apoptosis and impaired mitochondrial function, including reduced bioenergetics, mitochondrial DNA copy number and quality control mechanisms, along with inactivation of the PI3K/AKT/AMPK signalling pathways. Furthermore, analysis of myocardial tissue revealed elevated metal accumulation, particularly within mitochondria. Chelation of PM2.5 -associated metals using EDTA significantly mitigated the toxic effects on cardiac IR pathology, as confirmed in both rat myocardium and H9c2 cells. These findings suggest that metals in PM2.5 play a crucial role in inducing cardiotoxicity, impairing myocardial resilience to stress through mitochondrial accumulation and dysfunction.

PM2.5 Exposure-Linked Mitochondrial Dysfunction Negates SB216763-Mediated Cardio-Protection against Myocardial Ischemia-Reperfusion Injury.

GSK3ß is a promising target for treating various disease conditions, including myocardial ischemia-reperfusion injury (IR). This study investigated the potential of GSK3ß as a novel drug for managing IR in rats exposed to PM2.5 for 1 day and up to 21 days. Female Wistar rats were exposed to PM2.5 at a concentration of 250 µg/m3 for 3 h daily for either a single day or 21 days. After exposure, the isolated rat hearts underwent 30 min of ischemia followed by 60 min of reperfusion. GSK3ß inhibition effectively reduced IR injury in rat hearts from animals exposed to PM2.5 for 1 day but not in those exposed for 21 days. PM2.5 exposure disrupted the redox balance in mitochondria and reduced the gene expression of antioxidants (glutaredoxin and peroxiredoxin) and NRF2, which protects against oxidative stress. PM2.5 also impaired mitochondrial bioenergetics, membrane potential, and quality control, leading to mitochondrial stress. Importantly, PM2.5 increased the translocation of GSK3ß into mitochondria and compromised the overall mitochondrial function, particularly in the 21-day-exposed rat myocardium. The results indicate that extended exposure to PM2.5 leads to oxidative stress that disrupts mitochondrial function and diminishes the effectiveness of GSK3ß inhibitors in offering cardio-protection through mitochondria.

DNA hypomethylation by fisetin preserves mitochondria functional genes and contributes to the protection of I/R rat heart.

Myocardial I/R can alter the expression of different sets of cardiac genes that negatively influence the I/R outcome via epigenetic modifications. Fisetin is known to be cardioprotective against I/R, but its underlying epigenetic mode of action is not known and is addressed in the present study. Male Wistar rats were subjected to I/R by using the Langendorff perfusion system. Fisetin (20 mg/kg; i.p.) was administered before I/R induction, followed by the measurement of cardiac injury, hemodynamics, physiological indices, the differential expression of genes that regulate DNA methylation, and the function of mitochondria were performed. Fisetin administered I/R rat heart significantly reduced the global DNA hypermethylation and infarct size with an improved physiological recovery, measured via RPP (81%) and LVDP (82%) from the I/R control. Additionally, we noted decreased expression of the DNMT1 gene by 35% and increased expression of the TET1, TET2, and TET3 genes in fisetin-treated I/R rat hearts. Molecular docking analysis data reveals that the fisetin inhibits DNMT1 at the substrate binding site with minimum binding energy (- 8.2 kcal/mol) compared to the DNMT1 inhibitor, 5-azacytidine. Moreover, fisetin-treated I/R heart reversed the expression of the I/R-linked declined expression of bioenergetics genes (MT-ND1, MT-ND2, MT-ND4, MT-Cyt B, MT-COX1, MT-COX2, MT-ATP6), mitochondrial fission gene (Fis1), replication control genes PGC-1α, POLG, and TFAM to near-normal level. Based on the above findings, we demonstrated that fisetin possesses the ability to modulate the expression of different mitochondrial genes via influencing the global DNA methylation in cardiac tissue, which contributes significantly to the improved contractile function and thereby renders cardioprotection against I/R.

PM2.5-Induced Cardiac Structural Modifications and Declined Pro-Survival Signalling Pathways Are Responsible for the Inefficiency of GSK-3ß Inhibitor in Attenuating Myocardial Ischemia-Reperfusion Injury in Rats.

Circulatory GSK3ß is recognized as a biomarker and therapeutic target for diseases, including myocardial diseases. However, its potential as a target for myocardial ischemia-reperfusion injury (IR) in the presence of PM2.5 exposure is unclear. Wistar rats underwent IR following either a 21-day or single exposure to PM2.5 at a concentration of 250 µg/m3. The effects of GSK3ß inhibitor on cardiac physiology, tissue injury, mitochondrial function, and the PI3K/AKT/GSK3ß signalling axis were examined. The inhibitor was not effective in improving hemodynamics or reducing IR-induced infarction in the myocardium exposed to PM2.5 for 21 days. However, for a single-day exposure, the inhibitor showed potential in mitigating cardiac injury. In normal hearts undergoing IR, the inhibitor activated the PI3K/AKT signalling pathway, improved mitochondrial function, and reduced oxidative stress. These positive effects were not observed in PM2.5-exposed rats. Furthermore, the inhibitor stimulated autophagy in hearts exposed to PM2.5 for 21 days and subjected to IR, resulting in increased mTOR expression and decreased AMPK expression. In normal hearts and those exposed to a single dose of PM2.5, the inhibitor effectively activated the PI3K/Akt/AMPK axis. These findings suggest that GSK3ß may not be a reliable therapeutic target for IR in the presence of chronic PM2.5 exposure.

Exposure to real ambient particulate matter inflicts cardiac electrophysiological disturbances, vascular calcification, and mitochondrial bioenergetics decline more than diesel particulate matter: consequential impact on myocardial ischemia-reperfusion injury.

Particulate matter (PM) present in the air sample comprises different sizes and is derived from multiple sources, in particular from diesel engines. In the present study, we assessed the cardiotoxic effect of PM2.5 from real ambient air sample and diesel vehicular exhaust from a specific location and compared it with SRM-2975. Female Wistar rats were exposed to PM2.5 from real ambient PM (RA_PM), diesel particulate matter (DPM), and SRM-2975 for 3h daily for 21 days followed by cardiotoxicity assessment. Twenty-one days of daily PM2.5 exposure induced hypertrophy, vascular calcification, and alterations in cardiac electrophysiology, where the changes were more prominent in the animals exposed to RA_PM. The gross pathological changes were supported by altered mitochondrial function and increased oxidative stress in the myocardium. To evaluate the cardiac responsive ability, isolated rat hearts were subjected to ischemia-reperfusion injury (IR), and the results showed significantly low recovery in the RA_PM-exposed rat hearts. Chemical analysis of PM2.5 by ICPMS from different sources indicated the presence of additional metals like Cr, Ni, Ga, As, Rb, Cd, Ba, La, and Ce in the RA_PM sample. Additionally, the chelation of metals in the RA_PM enhanced the cell viability of H9c2 cells when compared to the non-chelated sample. Based on the above observations, we conclude that PM2.5 from the ambient air sample exhibited higher cardiovascular toxicity than DPM, emphasizing the contribution of non-diesel components of PM2.5 and the need for a comprehensive approach to tackle the PM2.5 in the air sample.

PM2.5 from diesel exhaust attenuated fisetin mediated cytoprotection in H9c2 cardiomyocytes subjected to ischemia reoxygenation by inducing mitotoxicity.

The impact of PM2.5 from diesel exhaust (termed as diesel particulate matter (DPM)) on ischemia re-oxygenation (IR) injury and the consequent effect of fisetin to attenuate this injury remains unclear. IR was induced in H9c2 cells after 24 hrs of fisetin treatment. The cells when incubated with 100 µg/mL of DPM followed by IR, induced 60% cell death which was escalated to 78% with DPM exposure. Fisetin significantly attenuated IR induced cytotoxicity, improved mitochondrial activity and reduced oxidative stress in normal cells but failed to render protection against IR in presence of DPM. Isolated mitochondria experiment confirmed the mitotoxic effect of DPM. Immunoblot analysis established the failure of fisetin to activate PI3K/Akt signaling pathway. Based on the above observations, we concluded that fisetin mediated protection against IR was abrogated with DPM exposure due to augmented mitochondrial dysfunction and inactivation of PI3K/Akt signaling pathway.

Consequential Impact of Particulate Matter Linked Inter-Fibrillar Mitochondrial Dysfunction in Rat Myocardium Subjected to Ischemia Reperfusion Injury.

A previous study has reported that exposure to PM2.5 from diesel exhaust (diesel particulate matter (DPM)) for 21 days can deteriorate the cardiac recovery from myocardial ischemia reperfusion injury (IR), where the latter is facilitated by the efficiency of mitochondrial subpopulations. Many investigators have demonstrated that IR impact on cardiac mitochondrial subpopulations is distinct. In the present study, we decipher the role of PM2.5 on IR associated mitochondrial dysfunction at the subpopulation level by administrating PM2.5 directly to isolated female rat hearts via KH buffer. Our results demonstrated that PM2.5 administered heart (PM_C) severely deteriorated ETC enzyme activity (NQR, SQR, QCR, and COX) and ATP level in both IFM and SSM from the normal control. Comparatively, the declined activity was prominent in IFM fraction. Moreover, in the presence of IR (PM_IR), mitochondrial oxidative stress was higher in both subpopulations from the normal, where the IFM fraction of mitochondria experienced elevated oxidative stress than SSM. Furthermore, we assessed the in vitro protein translation capacity of IFM and SSM and found a declined ability in both subpopulations where the inability of IFM was significant in both PM_C and PM_IR groups. In support of these results, the expression of mitochondrial genes involved in fission, fusion, and mitophagy events along with the DNA maintenance genes such as GUF1, LRPPRC, and HSD17-b10 were significantly altered from the control. Based on the above results, we conclude that PM2.5 administration to the heart inflicted mitochondrial damage especially to the IFM fraction, that not only deteriorated the cardiac physiology but also reduced its ability to resist IR injury.

Recent advances in potential of Fisetin in the management of myocardial ischemia-reperfusion injury-A systematic review.

BACKGROUND: The primary therapeutic strategy in managing ischemic heart diseases is to restore the perfusion of the myocardial ischemic area by surgical methods that often result in an unavoidable injury called ischemia-reperfusion injury (IR). Fisetin is an effective flavonoid with antioxidant and anti-inflammatory properties, proven to be cardioprotective against IR injury in both in-vitro and invivo models, apart from its promising health benefits against cancer, diabetes, and neurodegenerative ailments. PURPOSE: The potential of fisetin in attenuating myocardial IR is inconclusive as the effectiveness of fisetin needs more understanding in terms of its possible target sites and underlying different mechanisms. Considering the surge in recent scientific interests in fisetin as a pharmacological agent, this review not only updates the existing preclinical and clinical studies with fisetin and its underlying mechanisms but also summarizes its possible targets during IR protection. METHODS: We performed a literature survey using search engines Pubmed, PMC, Science direct, Google, and research gate published across the years 2006-2021. The relevant studies were extracted from the databases with the combinations of the following keywords and summarized: myocardial ischemia-reperfusion injury, natural products, flavonoid, fisetin, PI3K, JAK-STAT, Nrf2, PKC, JNK, autophagy. RESULTS: Fisetin is reported to be effective in attenuating IR injury by delaying the clotting time, preserving the mitochondrial function, reducing oxidative stress, and inhibiting GSK 3ß. But it failed to protect diseased cardiomyocytes challenged to IR. As discussed in the current review, fisetin not only acts as a conventional antioxidant and anti-inflammatory agent to exert its biological effect but may also exert modulatory action on the cellular metabolism and adaptation via direct action on various signalling pathways that comprise PI3K, JAK-STAT, Nrf2, PKC, JNK, and autophagy. Moreover, the dosage of fisetin and co-morbidities like diabetes and obesity are found to be detrimental factors for cardioprotection. CONCLUSION: For further evaluation and smooth clinical translation of the fisetin molecule in IR treatment, researchers should pay close attention to the potential of fisetin to possibly alter the key cardioprotective pathways and dosage, as the efficacy of fisetin is tissue and cell type-specific and varies with different doses.

Long-term administration of fisetin was not as effective as short term in ameliorating IR injury in isolated rat heart.

The current study aims to determine the comparative efficacy of fisetin in reducing myocardial ischemia-reperfusion injury (IR) in isolated rat hearts when the drug was given either oral or intraperitoneal (ip) for short-term and long-term administration. Rats treated with fisetin (20 mg/kg-oral/ip) for short (30 min prior to surgery) and long (15 days prior to surgery followed by 1-day washout) duration were subjected to myocardial IR using Langendorf perfusion system. Hemodynamics, cardiac injury, mitochondrial functional assessment, and fisetin levels were estimated. Unlike the long-term administration of fisetin, the short-term treated-rat heart exhibited significant cardioprotection, measured via hemodynamic indices (RPP in mmHg × beats/min × 10 ^ 4: IR - 4 ± 0.1, FIPS - 2.49 ± 0.18, FIPL - 1.87 ± 0.14), reduced infarct size (in % area of infarct: IR - 38 ± 5, FIPS - 17 ± 1, FOS - 14 ± 2), improved mitochondrial ETC enzyme activity (NQR activity in IFM: FIPS - 0.25 ± 0.016, FIPL - 0.20 ± 0.02), and declined oxidative stress (GSH in IFM: FIPS - 1.52 ± 0.14, FIPL - 1.25 ± 0.22). However, no significant difference in the protection was observed between the animals treated with oral or intraperitoneally administered fisetin. Single dose of fisetin administration before IR protocol was more effective than 15 days of fisetin-treated drug followed by 1-day washout, thus may not be suitable for long-term dietary supplement for post-surgical cardiac rehabilitation.

PM2.5 Exposure Lowers Mitochondrial Endurance During Cardiac Recovery in a Rat Model of Myocardial Infarction.

Many studies have reported the negative effect of PM2.5 exposure on heart function which is likely to impair postcardiac surgery rehabilitation that is involved in recovery and wound healing, yet the direct effects of PM2.5 from diesel exhaust (DPM) on cardiac recovery is unknown. To study the impact of DPM on cardiac recovery and repair, we utilized isoproterenol induced myocardial infarction (MI) model where female rats were exposed to DPM prior and after MI induction. The experimental groups comprise of normal, ISO control, DPM control (42 days of exposure), DPM exposed prior (21 days) and after (21 days) MI induction (D + I + D) and DPM exposed (21 days) after MI (I + D). Post-MI rat hearts from D + I + D group exhibited higher fibrosis, elevated cardiac injury and altered electrophysiology, where this pathology was also observed in I + D group animals which was mild. Loss of mitochondrial quality was evident in DPM exposed animals with and without MI, where severe mitochondrial damage persisted in D + I + D group. In addition, these animals showed striking decline in ETC enzyme activity, ATP levels, mitochondrial copy number and down regulation of PGC1-α, TFAM and POLG along with the genes involved in mitophagy and mitofusion. Besides, the MI associated inactivation of cardio protective signalling pathways like PI3K and Akt were persistent in D + I + D group. In fact, I + D group animals also showed a similar pattern of change, but in a mild form. Taken together, exposure to PM2.5 increases the risk, frequency or progression of MI by impairing the recovery potential of the myocardium.

Inhalation of PM2.5 from diesel exhaust promote impairment of mitochondrial bioenergetics and dysregulate mitochondrial quality in rat heart: implications in isoproterenol-induced myocardial infarction model.

Aim: Ambient exposure of PM2.5 from diesel exhaust (termed as diesel particulate matter [DPM]) can induce cardiotoxicity that can be manifested into myocardial ischemia/infarction, where the survival depends on mitochondrial function. The mechanism for DPM-induced mitochondrial dysfunction is yet to be elucidated and the consequential impact of impaired mitochondria on the severity of myocardial infarction (MI) has not been established.Materials and methods: Female Wistar rats were exposed to DPM (0.5 mg/ml) for 3 h daily (to achieve a PM2.5 concentration of 250 µg/m3) for 21 d trailed by an induction of MI using isoproterenol (ISO).Conclusion: DPM exposure altered the basal ECG pattern and increased heart weight (HW) to body weight (BW) ratio from control. Loss of mitochondrial quality in the cardiac tissue was observed in DPM exposed animals, measured via declined ETC enzyme activity, reduced ATP levels, high oxidative stress, low mitochondrial copy number, and low expression of the mitochondrial genes involved in mitophagy (PINK and PARKIN) and mitochondrial fusion (MFN-1). Subsequent induction of MI in DPM exposed animals (DPM + ISO) further deteriorated the normal sinus rhythm, accompanied by elevated plasma CK and LDH level, increased myocardial caspase activity, downregulation of Peroxisome proliferator-activated receptor-gamma coactivator (PGC1-α), transcription factor A (TFAM), DNA polymerase subunit gamma (POLG), and other mitochondrial quality control genes. Based on these results, we conclude that DPM alters the electrophysiology and ultrastructure of the heart that aggravates the MI-induced cardiotoxicity, where the diminished mitochondrial quality can be the potential contributor.

FIsetin Preserves Interfibrillar Mitochondria to Protect Against Myocardial Ischemia-Reperfusion Injury.

According to our previous study, fisetin (3,3',4',7-tetrahydroxyflavone), a bioactive phytochemical (flavonol), reportedly showed cardioprotection against ischemia-reperfusion injury (IRI) by reducing oxidative stress and inhibiting glycogen synthase kinase 3ß (GSK3ß) [1]. GSK3ß is said to exert a non-mitochondrial mediated cardioprotection; therefore, distinct mechanisms of GSK3ß on the regulatory effect of mitochondria need to be addressed. The two distinct mitochondrial subpopulations in the heart, namely interfibrillar mitochondria (IFM) and subsarcolemmal mitochondria (SSM), respond differently to disease states. The current study aimed to understand the effect of fisetin on the subpopulation-specific preservation of IFM and SSM while rendering cardioprotection against ischemia reperfusion (I/R). Rats were pre-treated with fisetin (20 mg/kg) intraperitoneally, and IRI was induced using Langendorff isolated heart perfusion technique. Hemodynamic parameters were recorded, and the cardiac injury was assessed using infarct size (IS), lactate dehydrogenase (LDH), and creatine kinase (CK) levels. Subpopulation-specific mitochondrial preservation was evaluated by electron transport chain (ETC), catalase, superoxide dismutase (SOD), and glutathione (GSH) activities. The bioavailability of fisetin in IFM and SSM was measured using the fluorescence method. The ability of fisetin to bind directly to the mitochondrial complex-1 and activating it through donating electrons to FMN was studied using molecular docking studies and further validated by in vitro rotenone sensitivity assay. Cardioprotective effects exhibited by fisetin were mainly mediated through IFM preservation. Mitochondrial bioavailability of fisetin is more in IFM than SSM in both ex vivo and in vitro conditions. Fisetin increased mitochondrial ATP production in I/R insult hearts by activating ETC complex 1. Inhibition of complex 1 prevents the ATP-producing capacity of fisetin. Our results provide evidence that fisetin plays a protective role in myocardial IRI, possibly by preserving the functional activities of IFM.

Diesel particulate matter exposure deteriorates cardiovascular health and increases the sensitivity of rat heart towards ischemia reperfusion injury via suppressing mitochondrial bioenergetics function.

Documents from previous studies do not sufficiently explain the pathophysiological alterations involved in rat hearts exposed to PM2.5 from diesel exhaust, termed as Diesel Particulate matter (DPM). In the present study, we explored the cardiovascular effect of DPM exposure on the recovery of heart from Ischemia reperfusion injury (IR) and explored the probable cause-effect relationship. Two groups of female Wistar rats were exposed to 0.5 mg/ml DPM for 1 h and 3 h durations daily for 21 days via a whole-body exposure system. At the end of 21st day, the animals were sacrificed and the heart was subjected to IR via Langendorff isolated rat heart perfusion system. 21 days of exposure altered cardiac electrophysiology and the ultra-structure of myocardium. Also, the same group of animals exhibited calcification in the vasculature. These changes were prominent in animals exposed to DPM for 3 h daily. Administration of DPM to H9C2 cells resulted in 15% and 36% cell death after 1hr and 3hrs of incubation, respectively. When the hearts were challenged to IR, both 1 h and 3 h exposed hearts exhibited a significant decline in IR recovery. At the sub-cellular level, DPM exposure reduced ATP levels, mitochondrial copy number, and increased oxidative stress after IR in both exposure groups. These changes were markedly seen in the interfibrillar mitochondrial fraction of the mitochondria. Hence, we conclude that exposure to PM2.5 from diesel exhaust alters electrophysiology and ultrastructure of heart and reduces the level of cellular mediators, thereby compromising the ability of heart to withstand IR injury.

Mitochondria and traffic-related air pollution linked coronary artery calcification: exploring the missing link.

The continuing increase in the exposure to Traffic-related air pollution (TRAP) in the general population is predicted to result in a higher incidence of non-communicable diseases like cardiovascular disease. The chronic exposure of air particulate matter from TRAP upon the vascular system leads to the enhancement of deposition of calcium in the vasculature leading to coronary artery calcification (CAC), triggered by inflammatory reactions and endothelial dysfunction. This calcification forms within the intimal and medial layers of vasculature and the underlying mechanism that connects the trigger from TRAP is not well explored. Several local and systemic factors participate in this active process including inflammatory response, hyperlipidemia, presence of self-programmed death bodies and high calcium-phosphate concentrations. These factors along with the loss of molecules that inhibit calcification and circulating nucleation complexes influence the development of calcification in the vasculature. The loss of defense to prevent osteogenic transition linked to micro organelle dysfunction that includes deteriorated mitochondria, elevated mitochondrial oxidative stress, and defective mitophagy. In this review, we examine the contributory role of mitochondria involved in the mechanism of TRAP linked CAC development. Further we examine whether TRAP is an inducer or trigger for the enhanced progression of CAC.

Fisetin Attenuates Myocardial Ischemia-Reperfusion Injury by Activating the Reperfusion Injury Salvage Kinase (RISK) Signaling Pathway.

Ischemia-reperfusion (I/R) injury is an unavoidable injury that occurs during revascularization procedures. In the previous study, we reported that fisetin is a natural flavonoid that attenuates I/R injury by suppressing mitochondrial oxidative stress and mitochondrial dysfunction. Though fisetin is reported as a GSK3ß inhibitor, it remains unclear whether it attenuates myocardial ischemia by activating the phosphoinositide 3-kinase/protein kinase B (PI3K/Akt) pathway, thereby inhibiting the downstream GSK3ß, or by directly interacting with GSK3ß while rendering its cardioprotection. In this study, the research team investigates the possible mechanism of action of fisetin while rendering its cardioprotective effect against myocardial I/R injury in rats. For this investigation, the team utilized two myocardial I/R models: Ligation of the left anterior descending artery and Langendorff isolated heart perfusion system. The latter has no neurohormonal influences. The PI3K inhibitor (Wortmannin, 0.015 mg/kg), GSK3ß inhibitor (SB216763, 0.7 mg/kg), and fisetin (20 mg/kg) were administered intraperitoneally before inducing myocardial I/R. The result of this study reveals that the administration of fisetin decreases the myocardial infarct size, apoptosis, lactate dehydrogenase, and creatine kinase in serum\perfusate of the rat hearts subjected to I/R. However, the inhibition of PI3K with Wortmannin significantly reduced the cardioprotective effect of fisetin both in the ex vivo and vivo models. The administration of GSK3ß inhibitor after the administration of fisetin and Wortmannin, re-establishing the cardioprotection, indicates the major role of PI3K in fisetin action. Changes in myocardial oxidative stress (level) and mitochondrial functional preservation of interfibrillar and subsarcolemmal mitochondria support the above findings. Hence, the team here reports that fisetin conferred its cardioprotection against I/R injury by activating the PI3K/Akt/GSK3ß signaling pathway in rat hearts.