MEK/ERK pathway mediates cytoprotection of salvianolic acid B against oxidative stress‐induced apoptosis in rat bone marrow stem cells

To improve the survival and/or differentiation of grafted BMSCs (bone marrow stem cells) represents one of the challenges for the promising cell‐based therapy. Considerable reports have implicated Sal B (salvianolic acid B), a potent aqueous extract of Salvia miltiorrhiza, in enhancing the survival of cells under various conditions. In this study, we investigated the effect of Sal B on H₂O₂‐induced apoptosis in rat BMSCs, focusing on the survival signalling pathways. Results indicated that the MEK [MAPK (mitogen‐activated protein kinase)/ERK (extracellular‐signal‐regulated kinase) kinase] inhibitor (PD98059) and 10 μM Sal B remarkably prevented BMSCs from H₂O₂‐induced apoptosis through attenuating caspase‐3 activation, which is accompanied by the significant up‐regulation of Bcl‐2. In addition, the ROS (reactive oxygen species) accumulation was also reduced after Sal B treatment. Furthermore, Sal B inhibited the ERK1/2 phosphorylations stimulated by H₂O₂. Taken together, our results showed that H₂O₂‐induced apoptosis in BMSCs via the ROS/MEK/ERK1/2 pathway and Sal B may exert its cytoprotection through mediating the pathway.     

Abamectin induces apoptosis and autophagy by inhibiting reactive oxygen species‐mediated PI3K/AKT signaling in MGC803 cells

Abamectin (ABA) is one of the most widely used compounds in agriculture and veterinary medicine. However, the cytotoxicity of ABA in human gastric cells is utterly unknown. In this study, ABA suppressed the proliferation of MGC803 cells by arresting the cell cycle at the G0/G1‐phase. Moreover, ABA induced mitochondrial‐mediated apoptosis by inducing the loss of mitochondrial membrane potential, upregulation of Bax/Bcl‐2, and activation of caspase‐3. ABA significantly improved the LC3‐II/LC3‐I ratio and reduced P62 protein expression in a dose‐dependent manner. Through detection of the reactive oxygen species (ROS) levels, we found ABA induced the accumulation of intracellular ROS and then reduced PI3K/AKT signaling activation related to MGC803 cell apoptosis and autophagy. Our results indicate that ABA exerts cytotoxic effects on human MGC803 cells through apoptosis and autophagy by inhibiting ROS‐mediated PI3K/AKT signaling. Furthermore, ABA may be a potential risk to human gastric health.     

Neuroprotective effects of astaxanthin against oxygen and glucose deprivation damage via the PI3K/Akt/GSK3β/Nrf2 signalling pathway in vitro

Astaxanthin (ATX), which is the most abundant flavonoid in propolis, has previously shown neuroprotective properties against cerebral ischaemia‐induced apoptosis. However, the mechanisms by which ATX mediates its therapeutic effects are unclear. At present, we explored the underlying mechanisms involved in the protective effects of ATX via the phosphoinositide 3‐kinase (PI3K)/Akt/glycogen synthase kinase 3 beta (GSK3β)/nuclear factor erythroid 2‐related factor 2 (Nrf2) signalling pathway in SH‐SY5Y cells. The PI3K/Akt inhibitor LY294002 and GSK3β inhibitor LiCl were employed in this study. Pre‐treatment with ATX for 24 hours significantly decreased the oxygen and glucose deprivation (OGD)‐induced viability loss, reduced the proportion of apoptosis and regulated OGD‐mediated reactive oxygen species (ROS) production. Furthermore, ATX suppressed OGD‐caused mitochondrial membrane potential and decomposition of caspase‐3 to cleaved caspase‐3, and heightened the B‐cell lymphoma 2 (Bcl‐2)/Bax ratio. PI3K/Akt/GSK3β/Nrf2 signalling pathway activation in SH‐SY5Y cells was verified by Western blot. ATX and LiCl treatment raised the protein levels of p‐Akt, p‐GSK3β, nucleus Nrf2 and haeme oxygenase 1 (HO‐1). However, these protein expression levels decreased by treatment of LY294002. The above in vitro data indicate that ATX can confer neuroprotection against OGD‐induced apoptosis via the PI3K/Akt/GSK3β/Nrf2 signalling pathway.     

LIGHT/IFN‐γ triggers β cells apoptosis via NF‐κB/Bcl2‐dependent mitochondrial pathway

LIGHT recruits and activates naive T cells in the islets at the onset of diabetes. IFN‐γ secreted by activated T lymphocytes is involved in beta cell apoptosis. However, whether LIGHT sensitizes IFNγ‐induced beta cells destruction remains unclear. In this study, we used the murine beta cell line MIN6 and primary islet cells as models for investigating the underlying cellular mechanisms involved in LIGHT/IFNγ – induced pancreatic beta cell destruction. LIGHT and IFN‐γ synergistically reduced MIN6 and primary islet cells viability; decreased cell viability was due to apoptosis, as demonstrated by a significant increase in Annexin V⁺ cell percentage, detected by flow cytometry. In addition to marked increases in cytochrome c release and NF‐κB activation, the combination of LIGHT and IFN‐γ caused an obvious decrease in expression of the anti‐apoptotic proteins Bcl‐2 and Bcl‐xL, but an increase in expression of the pro‐apoptotic proteins Bak and Bax in MIN6 cells. Accordingly, LIGHT deficiency led to a decrease in NF‐κB activation and Bak expression, and peri‐insulitis in non‐obese diabetes mice. Inhibition of NF‐κB activation with the specific NF‐κB inhibitor, PDTC (pyrrolidine dithiocarbamate), reversed Bcl‐xL down‐regulation and Bax up‐regulation, and led to a significant increase in LIGHT‐ and IFN‐γ‐treated cell viability. Moreover, cleaved caspase‐9, ‐3, and PARP (poly (ADP‐ribose) polymerase) were observed after LIGHT and IFN‐γ treatment. Pretreatment with caspase inhibitors remarkably attenuated LIGHT‐ and IFNγ‐induced cell apoptosis. Taken together, our results indicate that LIGHT signalling pathway combined with IFN‐γ induces beta cells apoptosis via an NF‐κB/Bcl2‐dependent mitochondrial pathway.     

BBR induces apoptosis in HepG2 cell through an Akt‐ASK1‐ROS‐p38MAPKs‐linked cascade

Berberine (BBR) has indicated significant antimicrobial activity against a variety of organisms including bacteria, viruses, and fungi. The mechanism by which BBR initiates apoptosis remains poorly understood. In the present study, we demonstrated that BBR exhibited significant cytotoxicity in human hepatoma HepG2 cells. Herein, we investigated cytotoxicity mechanism of BBR in HepG2 cells. The results showed that the induction of apoptosis in HepG2 cells by BBR was characterized by DNA fragmentation, an increased percentage of annexin V, and the activation of caspase‐3. The expressions of Bcl‐2 protein and pro‐caspase‐3 were reduced by BBR in HepG2 cells. However, Bax protein was increased in the cells. BBR‐induced apoptosis was preceded by increased generation of reactive oxygen species (ROS). NAC treatment, a scavenger of ROS, reversed BBR‐induced apoptosis effects via inhibition of Bax activation and Bcl‐2 inactivation. BBR‐induced, dose‐dependent induction of apoptosis was accompanied by sustained phosphorylation of MAP Kinases (JNK and p38 MAPK), ASK1, Akt, and p53. Furthermore, SB203580, p38 inhibitor, reduced the apoptotic effect of BBR, and blocks the generation of ROS and NO as well as activation of Bax. We found that the treatment of HepG2 cells with BBR triggers generation of ROS through Akt phosphorylation, resulting in dissociation of the ASK1‐mediated activation of JNK and p38 pathways. J. Cell. Biochem. 109: 329–338, 2010. © 2009 Wiley‐Liss, Inc.     

Triptolide improves spinal cord injury by promoting autophagy and inhibiting apoptosis

We investigated the effect of triptolide (TP) on spinal cord injury (SCI), and its underlying mechanism. Following the establishment of the SCI model using YFP H‐line transgenic mice, TP was intraperitoneally injected at a dose of 0.2 mg/kg once daily for 7 days. Behavioral tests, Nissl staining, and hematoxylin–eosin staining were employed to assess motor function recovery and neuronal cell death. Western blot and immunofluorescence staining were used to assess autophagy‐associated proteins (LC3B, p62, Beclin‐1) and the apoptosis‐associated proteins (Bcl‐2, caspase‐3, Bax). The TP‐treated group showed improved motor functions, and reduced neuronal cell death. Also, significant upregulation of Bcl‐2 and LC3B expressions, with the downregulation of p62, Bax and caspase‐3 expressions were found in the TP‐treated group. Additionally, phosphorylation of extracellular signal‐regulated protein kinases 1 and 2 (ERK1 and ERK2) was decreased in the TP‐treated group. TP mediates its protective effect in SCI by promoting the autophagic pathway while inhibiting the MAPK/ERK1/2 signaling pathway. These results demonstrate the therapeutic potential of TP in SCI.     

Phloroglucinol Derivatives from the Fruits of Eucalyptus globulus and Their Cytotoxic Activities

A new phloroglucinol derivative, named eucalyptin B ( 1 ), along with five related known compounds ( 2 – 6 ), was isolated from the fruits of Eucalyptus globulus. Their structures were elucidated by means of 1D‐ and 2D‐NMR spectroscopy, with the absolute configuration of 1 determined by electronic circular dichroism (ECD) calculations. All isolated compounds ( 1 – 6 ) were evaluated for their cytotoxic activities against lung (A549), breast (4T1), and skin (B16F10) cancer cell lines. On the basis of cell viability assay, the cytotoxic activity of eucalyptin B ( 1 ) was further confirmed by apoptosis assay. Additionally, after treatment with eucalyptin B ( 1 ), the apoptosis factor proteins (Bcl2 and Bax) and caspase‐3 levels in A549 cells were also determined by Western‐blot analysis. By cytotoxic assay, eucalyptin B ( 1 ) exhibited potent cytotoxicity against A549 cells with an IC₅₀ value of 1.51 μm and induced concentration dependent apoptosis of up to 49%. Additionally, eucalyptin B ( 1 ) inhibited 5‐fold and increased 10‐folds in the level of Bcl2 and Bax, respectively. Furthermore, the 11‐fold increase in the level of caspase‐3 confirmed eucalyptin B ( 1 ) activated caspase dependent apoptosis pathway. In conclusion, the isolated compound eucalyptin B ( 1 ) has promising cytotoxic activity in tumor cells, especially in A549.     

Platycodin D suppresses cisplatin‐induced cytotoxicity by suppressing ROS‐mediated oxidative damage,apoptosis, and inflammation in HEK‐293 cells

Cisplatin, a proven effective chemotherapeutic agent, has been used clinically to treat malignant solid tumors, whereas its clinical use is limited by serious side effect including nephrotoxicity. Platycodin D (PD), the major and marked saponin isolated from Platycodon grandiflorum, possesses many pharmacological effects. In this study, we evaluated its protective effect against cisplatin‐induced human embryonic kidney 293 (HEK‐293) cells injury and elucidated the related mechanisms. Our results showed that PD (0.25, 0.5, and 1 μM) can dose‐dependently alleviate oxidative stress by decreasing malondialdehyde and reactive oxygen species, while increasing the levels of glutathione, superoxide dismutase, and catalase. Moreover, the elevation of apoptosis including Bax, Bad, cleaved caspase‐3,‐9, and decreased protein levels of Bcl‐2, Bcl‐XL induced by cisplatin were reversed after PD treatment. Importantly, PD pretreatment can also regulate PI3K/Akt and ERK/JNK/p38 signaling pathways. Furthermore, PD was found to reduce NF‐κB‐mediated inflammatory relative proteins. Our finding indicated that PD exerted significant effects on cisplatin induced oxidative stress, apoptosis and inflammatory, which will provide evidence for the development of PD to attenuate cisplatin‐induced nephrotoxicity.     

The Nitric Oxide Prodrug JS‐K Induces Ca2+‐Mediated Apoptosis in Human Hepatocellular Carcinoma HepG2 Cells

Hepatocellular carcinoma is one of the most common and deadly forms of human malignancies. JS‐K, O²‐(2, 4‐dinitrophenyl) 1‐ [(4‐ethoxycarbonyl) piperazin‐1‐yl] diazen‐1‐ium‐1, 2‐diolate, has the ability to induce apoptosis of tumor cell lines. In the present study, JS‐K inhibited the proliferation of HepG2 cells in a time‐ and concentration‐dependent manner and significantly induced apoptosis. JS‐K enhanced the ratio of Bax‐to‐Bcl‐2, released of cytochrome c (Cyt c) from mitochondria and the activated caspase‐9/3. JS‐K caused an increasing cytosolic Ca²⁺ and the loss of mitochondrial membrane potential. Carboxy‐PTIO (a NO scavenger) and BAPTA‐AM (an intracellular Ca²⁺ chelator) significantly blocked an increasing cytosolic Ca²⁺ in JS‐K‐induced HepG2 cells apoptosis, especially Carboxy‐PTIO. Meanwhile, Carboxy‐PTIO and BAPTA‐AM treatment both attenuate JS‐K‐induced apoptosis through upregulation of Bcl‐2, downregulation of Bax, reduction of Cyt c release from mitochondria to cytoplasm and inactivation of caspase‐9/3. In summary, JS‐K induced HepG2 cells apoptosis via Ca²⁺/caspase‐3‐mediated mitochondrial pathway.     

Cell‐based high‐throughput screen for small molecule inhibitors of Bax translocation

Aberrant regulation of programmed cell death (PCD) has been tied to an array of human pathologies ranging from cancers to autoimmune disorders to diverse forms of neurodegeneration. Pharmacologic modulation of PCD signalling is therefore of central interest to a number of clinical and biomedical applications. A key component of PCD signalling involves the modulation of pro‐ and anti‐apoptotic Bcl‐2 family members. Among these, Bax translocation represents a critical regulatory phase in PCD. In the present study, we have employed a high‐content high‐throughput screen to identify small molecules which inhibit the cellular process of Bax re‐distribution to the mitochondria following commitment of the cell to die. Screening of 6246 Generally Recognized As Safe compounds from four chemical libraries post‐induction of cisplatin‐mediated PCD resulted in the identification of 18 compounds which significantly reduced levels of Bax translocation. Further examination revealed protective effects via reduction of executioner caspase activity and enhanced mitochondrial function. Consistent with their effects on Bax translocation, these compounds exhibited significant rescue against in vitro and in vivo cisplatin‐induced apoptosis. Altogether, our findings identify a new set of clinically useful small molecules PCD inhibitors and highlight the role which cAMP plays in regulating Bax‐mediated PCD.     

N‐terminal cleavage of Bax by calpain generates a potent proapoptotic 18‐kDa fragment that promotes Bcl‐2‐independent cytochrome C release and apoptotic cell death

Upon apoptosis induction, the proapoptotic protein Bax is translocated from the cytosol to mitochondria, where it promotes release of cytochrome c, a caspase‐activating protein. However, the molecular mechanisms by which Bax triggers cytochrome c release are unknown. Here we report that before the initiation of apoptotic execution by etoposide or staurosporin, an active calpain activity cleaves Bax at its N‐terminus, generating a potent proapoptotic 18‐kDa fragment (Bax/p18). Both the calpain‐mediated Bax cleavage activity and the Bax/p18 fragment were found in the mitochondrial membrane‐enriched fraction. Cleavage of Bax was followed by release of mitochondrial cytochrome c, activation of caspase‐3, cleavage of poly(ADP‐ribose) polymerase, and fragmentation of DNA. Unlike the full‐length Bax, Bax/p18 did not interact with the antiapoptotic Bcl‐2 protein in the mitochondrial fraction of drug‐treated cells. Pretreatment with a specific calpain inhibitor calpeptin inhibited etoposide‐induced calpain activation, Bax cleavage, cytochrome c release, and caspase‐3 activation. In contrast, transfection of a cloned Bax/p18 cDNA into multiple human cancer cell lines targeted Bax/p18 to mitochondria, which was accompanied by release of cytochrome c and induction of caspase‐3‐mediated apoptosis that was not blocked by overexpression of Bcl‐2 protein. Therefore, Bax/p18 has a cytochrome c–releasing activity that promotes cell death independent of Bcl‐2. Finally, Bcl‐2 overexpression inhibited etoposide‐induced calpain activation, Bax cleavage, cytochrome c release, and apoptosis. Our results suggest that the mitochondrial calpain plays an essential role in apoptotic commitment by cleaving Bax and generating the Bax/p18 fragment, which in turn mediates cytochrome c release and initiates the apoptotic execution. J. Cell. Biochem. 80:53–72, 2000. © 2000 Wiley‐Liss, Inc.     

Activated apoptotic and anti‐survival effects on rat hearts with fructose induced metabolic syndrome

Consumption of fructose has been linked to the development of metabolic syndrome, whereas the cardiomyopathic changes and cardiac apoptosis of dietary high‐fructose intake have not yet been clarified. The purpose of this study was to evaluate the effects of high‐fructose on cardiac apoptotic and survival pathways. Thirty‐two Wistar rats were randomly divided into a control group (CON), which received a standard chow diet, and a fructose‐induced metabolic syndrome group (FIMS), which received a 50% fructose‐content diet for 13 weeks. Histopathological analysis, TUNEL assays and Western blotting were performed on the excised hearts from both groups. The blood pressure, glucose, insulin, triglyceride and cholesterol levels were significantly increased in the FIMS group, compared with the CON group. The abnormal myocardial architecture, enlarged interstitial space and increased cardiac TUNEL‐positive apoptotic cells were observed in the FIMS group. The TNF‐α, TNF receptor 1, Fas ligand, Fas receptor, FADD, and activated caspase‐3 and 8 protein levels (Fas pathway) and the Bax, Bak, Bax/Bcl‐2, Bak/Bcl‐xL, cytosolic cytochrome c, and activated caspase‐3 and nine protein levels (mitochondria pathway) were increased in the FIMS group compared with those in the CON group. The IGFI, IGFI‐R, p‐PI3K, p‐Akt, Bcl‐2 and Bcl‐xL protein levels (survival pathway) were all significantly decreased in the FIMS group compared with those in the CON group. High‐fructose intake elevated blood pressure and glucose levels; moreover, high‐fructose diet activated cardiac Fas‐dependent and mitochondria‐dependent apoptotic pathways and suppressed the survival pathway, which might provide one possible mechanism for developing heart failure in patients with metabolic syndrome. Copyright © 2013 John Wiley & Sons, Ltd.     

ROS accumulation contributes to abamectin‐induced apoptosis and autophagy via the inactivation of PI3K/AKT/mTOR pathway in TM3 Leydig cells

Abamectin (ABA) as one of the worldwide used compounds in agriculture has raised safety concerns on nontarget organism toxicity. However, the study of male reproductive system damage caused by ABA remains unclear. Our aim is to investigate the effect of ABA‐induced cytotoxicity in TM3 Leydig cells and their underlying mechanisms. ABA inhibits TM3 cell viability and proliferation via cell cycle arrested in the G0/G1 phase. In addition, ABA‐induced mitochondrial depolarization leads to an imbalance in Bcl‐2 family expression, causing caspase‐dependent apoptosis in TM3 cells. The increased ratio of cells expression LC3 protein and LC3‐II to LC3‐I indicated the activation of autophagy potentially. Further experiments revealed ABA treatment reduced phosphatidylinositol 3‐kinase (PI3K), protein kinase B (AKT) phosphorylation, and mammalian target of rapamycin (mTOR) phosphorylation. Pretreatment with a PI3K/AKT inhibitor, LY294002, mimicked the ABA‐mediated effects on cytotoxicity. Pretreatment with a PI3K/AKT agonist, insulin‐like growth factor‐1, reversed the effects of ABA. ABA caused the accumulation of intracellular reactive oxygen species (ROS) by increased intensity of the ROS indicator. However, N‐acetylcysteine as ROS scavengers inhibited ABA‐induced apoptosis and autophagy and reversed these ABA‐mediated effects on PI3K/AKT/mTOR pathway. On the basis of the above results, it is suggested that ABA exposure induces apoptosis and autophagy in TM3 cells by ROS accumulation to mediate PI3K/AKT/mTOR signaling pathway suppression.     

Regulation of Survivin and Bcl‐2 in HepG2 Cell Apoptosis Induced by Quercetin

Quercetin, a widely distributed bioflavonoid, has been shown to induce growth inhibition in a variety of human cancer cells. However, the regulation of survivin and Bcl‐2 on the quercetin‐induced cell‐growth inhibition and apoptosis in cancer cells remains unclear. In the present study, we report that quercetin can inhibit proliferation and induce apoptosis in HepG2 cells in dose‐ and time‐dependent manner. Hoechst 33258 and acridine orange/ethidium bromide (AO/EB) staining showed that HepG2 cells underwent the typical morphologic changes of apoptosis characterized by nuclear shrinkage, chromatin condensation, or fragmentation after exposure to quercetin. Cell‐cycle analysis reveals a significant increase of the proportion of cells in G₀/G₁ phase. We also demonstrate that the levels of survivin and Bcl‐2 protein expression in HepG2 cells decreased concurrently, and the levels of p53 protein increased significantly after treatment with quercetin by immunocytochemistry analysis. Relative activity of caspase‐3 and caspase‐9 increased significantly. These data clearly indicate that quercetin‐induced apoptosis is associated with caspase activation, and the levels of survivin and Bcl‐2. Our results indicate that the expression of survivin may be associated with Bcl‐2 expression, and the inhibition expression of survivin, in conjunction with Bcl‐2, might cause more pronounced apoptotic effects. Together, concurrent down‐regulated survivin and Bcl‐2 play an important role in HepG2 cell apoptosis induced by quercetin.     

Apoptosis in zebrafish development

Apoptosis (programmed cell death) is important in normal biological processes and in pathogenesis in vertebrates. This review focuses on some of the prominent features of apoptosis during fish development. Caspases and other apoptosis-regulating genes have been cloned from zebrafish (Danio rerio) and other fish species. Elucidation of in vivo functions of apoptosis is focused on development, morphogenesis and sex differentiation. In an attempt to elucidate cause and effect relationships between caspase and development, transgenic zebrafish overexpressing procaspase-3 were generated. Stress-induced apoptosis in zebrafish embryos can be monitored by whole mount TUNEL staining and caspase assay. Thus, zebrafish is a useful experimental model animal for investigation of apoptosis in vivo.     

iTRAQ proteomics analysis reveals that PI3K is highly associated with bupivacaine‐induced neurotoxicity pathways

Bupivacaine, a commonly used local anesthetic, has potential neurotoxicity through diverse signaling pathways. However, the key mechanism of bupivacaine‐induced neurotoxicity remains unclear. Cultured human SH‐SY5Y neuroblastoma cells were treated (bupivacaine) or untreated (control) with bupivacaine for 24 h. Compared to the control group, bupivacaine significantly increased cyto‐inhibition, cellular reactive oxygen species, DNA damage, mitochondrial injury, apoptosis (increased TUNEL‐positive cells, cleaved caspase 3, and Bcl‐2/Bax), and activated autophagy (enhanced LC3II/LC3I ratio). To explore changes in protein expression and intercommunication among the pathways involved in bupivacaine‐induced neurotoxicity, an 8‐plex iTRAQ proteomic technique and bioinformatics analysis were performed. Compared to the control group, 241 differentially expressed proteins were identified, of which, 145 were up‐regulated and 96 were down‐regulated. Bioinformatics analysis of the cross‐talk between the significant proteins with altered expression in bupivacaine‐induced neurotoxicity indicated that phosphatidyl‐3‐kinase (PI3K) was the most frequently targeted protein in each of the interactions. We further confirmed these results by determining the downstream targets of the identified signaling pathways (PI3K, Akt, FoxO1, Erk, and JNK). In conclusion, our study demonstrated that PI3K may play a central role in contacting and regulating the signaling pathways that contribute to bupivacaine‐induced neurotoxicity.     

Kaempferol induces apoptosis in two different cell lines via Akt inactivation,Bax and SIRT3 activation,and mitochondrial dysfunction

Kaempferol (3,4′,5,7‐tetrahydroxyflavone) is a flavonoid with anti‐ and pro‐oxidant activity present in various natural sources. Kaempferol has been shown to posses anticancer properties through the induction of the apoptotic program. Here we report that treatment of the chronic myelogenous leukemia cell line K562 and promyelocitic human leukemia U937 with 50 µM kaempferol resulted in an increase of the antioxidant enzymes Mn and Cu/Zn superoxide dismutase (SOD). Kaempferol treatment induced apoptosis by decreasing the expression of Bcl‐2 and increasing the expressions of Bax. There were also induction of mitochondrial release of cytochrome c into cytosol and significant activation of caspase‐3, and ‐9 with PARP cleavage. Kaempferol treatment increased the expression and the mitochondria localization of the NAD‐dependent deacetylase SIRT3. K562 cells stably overexpressing SIRT3 were more sensitive to kaempferol, whereas SIRT3 silencing did not increase the resistance of K562 cells to kaempferol. Inhibition of PI3K and de‐phosphorylation of Akt at Ser473 and Thr308 was also observed after treating both K562 and U937 cells with kaempferol. In conclusion our study shows that the oxidative stress induced by kaempferol in K562 and U937 cell lines causes the inactivation of Akt and the activation of the mitochondrial phase of the apoptotic program with an increase of Bax and SIRT3, decrease of Bcl‐2, release of cytochrome c, caspase‐3 activation, and cell death. J. Cell. Biochem. 106: 643–650, 2009. © 2009 Wiley‐Liss, Inc.