Withaferin A enhances radiation-induced apoptosis in Caki cells through induction of reactive oxygen species, Bcl-2 downregulation and Akt inhibition

Withaferin A (Wit A), a natural compound derived from the medicinal plant Withania somnifera, has been reported for the anti-tumor effects, including the inhibition of tumor cell growth, metastasis and angiogenesis. In this study, we investigated the effect of Wit A on radiation-induced apoptosis in human renal cancer cells (Caki cells). Our results showed that, compared with Wit A or radiation alone, the combination of both resulted in a significant enhancement of apoptosis, showing the increase in the cleavage of caspase-3 and PARP as well as sub-G1 cell population. In addition, reactive oxygen species (ROS) generation was correlated with the enhancement of radiation-induced apoptosis by Wit A. Wit A downregulated Bcl-2 protein levels and ectopic expression of Bcl-2 in Caki cells attenuated the apoptosis induced by Wit A plus radiation. Taken together, these results indicate that Wit A enhanced radiation-induced apoptosis in Caki cells through ROS generation, down-regulation of Bcl-2 and Akt dephosphorylation. Thus, our study shows that Wit A may be used as an effective radiosensitizer in cancer therapy.     

Withaferin A-induced apoptosis in human breast cancer cells is mediated by reactive oxygen species

Withaferin A (WA), a promising anticancer constituent of Ayurvedic medicinal plant Withania somnifera, inhibits growth of MDA-MB-231 and MCF-7 human breast cancer cells in culture and MDA-MB-231 xenografts in vivo in association with apoptosis induction, but the mechanism of cell death is not fully understood. We now demonstrate, for the first time, that WA-induced apoptosis is mediated by reactive oxygen species (ROS) production due to inhibition of mitochondrial respiration. WA treatment caused ROS production in MDA-MB-231 and MCF-7 cells, but not in a normal human mammary epithelial cell line (HMEC). The HMEC was also resistant to WA-induced apoptosis. WA-mediated ROS production as well as apoptotic histone-associated DNA fragment release into the cytosol was significantly attenuated by ectopic expression of Cu,Zn-superoxide dismutase in both MDA-MB-231 and MCF-7 cells. ROS production resulting from WA exposure was accompanied by inhibition of oxidative phosphorylation and inhibition of complex III activity. Mitochondrial DNA-deficient Rho-0 variants of MDA-MB-231 and MCF-7 cells were resistant to WA-induced ROS production, collapse of mitochondrial membrane potential, and apoptosis compared with respective wild-type cells. WA treatment resulted in activation of Bax and Bak in MDA-MB-231 and MCF-7 cells, and SV40 immortalized embryonic fibroblasts derived from Bax and Bak double knockout mouse were significantly more resistant to WA-induced apoptosis compared with fibroblasts derived from wild-type mouse. In conclusion, the present study provides novel insight into the molecular circuitry of WA-induced apoptosis involving ROS production and activation of Bax/Bak.     

Arisostatins A induces apoptosis through the activation of caspase-3 and reactive oxygen species generation in AMC-HN-4 cells

A microbial secondary metabolite, arisostatins A (As-A), was originally discovered as a substance carrying the antibiotic activity against Gram-positive bacteria and shown to possess potent anti-tumor properties. The mechanism by which arisostatins A initiates apoptosis remains poorly understood. In the present report we investigated the effect of arisostatins A on activation of the apoptotic pathway in HN-4 cells. Arisostatins A was shown to be responsible for the inhibition of HN-4 cell growth by inducing apoptosis. Treatment with 4 microM arisostatins A for 24h produced morphological features of apoptosis and DNA fragmentation in HN-4 cells. Arisostatins A caused dose-dependent apoptosis and DNA fragmentation of HN-4 cells used as a model. Treatment with caspase inhibitor significantly reduced the arisostatins A-induced caspase 3 activation. In addition, arisostatins A-induced apoptosis was associated with the generation of reactive oxygen species (ROS), which was prevented by an antioxidant NAC (N-acetyl-cysteine). These data indicate that cytotoxic effect of arisostatins A on HN-4 cells is attributable to the induced apoptosis and that arisostatins A-induced apoptosis is mediated by caspase-3 activation pathway, loss of mitochondrial transmembrane potential (DeltaPsi(m)), and release of cytochrome c into cytosol.     

Artocarpin induces cell apoptosis in human osteosarcoma cells through endoplasmic reticulum stress and reactive oxygen species

Osteosarcoma is a malignant primary bone tumor that responds poorly to both chemotherapy and radiation therapy. However, because of side effects and drug resistance in chemotherapy and the insufficiency of an effective adjuvant therapy for osteosarcoma, it is necessary to research novel treatments. This study was the first to investigate the anticancer effects of the flavonoid derivative artocarpin in osteosarcoma. Artocarpin induced cell apoptosis in three human osteosarcoma cell lines—U2OS, MG63, and HOS. Artocarpin was also associated with increased intracellular reactive oxygen species (ROS). Mitochondrial dysfunction was followed by the release of cytochrome c from mitochondria and accompanied by decreased antiapoptotic Bcl-2 and Bcl-xL and increased proapoptotic protein Bak and Bax. Artocarpin triggered endoplasmic reticulum (ER) stress, as indicated by changes in cytosol calcium levels and increased glucose-regulated protein 78 and 94 expressions, and also increased calpains expression and activity. Animal studies revealed a dramatic 40% reduction in tumor volume after 18 days of treatment. This study demonstrated a novel anticancer activity of artocarpin against human osteosarcoma cells and in murine tumor models. In summary, artocarpin significantly induced cell apoptosis through ROS, ER stress, mitochondria, and the caspase pathway, and may thus be a novel anticancer treatment for osteosarcoma.     

2'-benzoyloxycinnamaldehyde induces apoptosis in human carcinoma via reactive oxygen species

2'-hydroxycinnamaldehyde (HCA) has been shown to have inhibitory effects on farnesyl protein transferase in vitro, angiogenesis, and tumor cell growth. However, mechanism for these inhibitions remains unknown. As a derivative of HCA, BCA (2'-benzoyl-oxycinnamaldehyde) was synthesized by replacing hydroxyl group with benzoyl-oxyl group. When p53-mutated cancer cell lines (MDA-MB-231 breast cancer cell and SW620 colon cancer cell) were treated with 10 microM HCA or BCA, it induced growth arrest and apoptosis of tumor cells. Markers of apoptosis such as degradations of chromosomal DNA and poly(ADP-ribose) polymerase and activation of caspase-3 were detected after HCA or BCA treatment. BCA-induced apoptosis was blocked by pretreatment of cells with anti-oxidants, glutathione, or N-acetyl-cysteine. In addition, BCA-induced activation of caspase-3 and degradation of poly(ADP-ribose) polymerase were abolished by pretreatment of cells with the anti-oxidants. These results suggest that reactive oxygen species are major regulator of BCA-induced apoptosis. HCA or BCA-induced accumulation of reactive oxygen species was detected by using DCF-DA, an intracellular probe of oxidative stress. Furthermore, when BCA (100 mg/kg) was administrated intraperitoneally or orally into a nude mouse, it inhibited >88 or 41% of tumor growth, respectively, without any detectable weight change. These results suggest that BCA is a good drug candidate for cancer therapy.     

Arsenite induces apoptosis in chinese hamster ovary cells by generation of reactive oxygen species

Arsenic, a human carcinogen, possesses a serious environmental threat but the mechanism of its toxicity remains unclear. Knowledge of how arsenic induces cell death and how cells escape the death path may help to understand arsenic carcinogenesis. We have investigated the nature of sodium arsenite-induced cell death in Chinese hamster ovary K1 cells. Following phosphate-citric acid buffer extraction, apoptotic cells with lower DNA content than the G1 cells were detected by flow cytometry. Immediately after 4 h of 40 μM arsenite treatment, no appreciable fraction of cells with sub-G1 DNA content was detected; however, the sub-G1 cell fraction increased with postarsenite incubation time, and detectable increase started at 8 h of incubation, whereas the intracellular peroxide level as measured by the fluorescent intensity of 2′,7′-dichlorofluorescein increased immediately following a 4-h arsenite treatment. Simultaneous treatment with arsenite plus antioxidant (N-acetyl-cysteine, Trolox, and Tempo); copper ion chelator (neocuproine); protein kinase inhibitor (H-7) or protein synthesis inhibitor (cycloheximide) reduced the fraction of sub-G1 cell and internucleosomal DNA degradation. Trolox, neocuproine, or cycloheximide given after arsenite treatment also effectively reduced apoptosis. These results lead to a working hypothesis that arsenite-induced apoptosis in CHO-K1 cells is triggered by the generation of hydrogen peroxide, followed by a copper-mediated Fenton reaction that catalyzes the production of hydroxyl radicals, which selectively activates protein kinase through de novo synthesis of macromolecules. © 1996 Wiley-Liss, Inc.     

2-Hydroxycurcuminoid induces apoptosis of human tumor cells through the reactive oxygen species-mitochondria pathway

2-Hydroxycinnamaldehyde (HCA) and curcumin have been reported to have antitumor effects against various human tumor cells in vitro and in vivo by generation of ROS. Aldehyde-free HCA analogs were synthesized based on the structure of curcumin, which we have called 2-hydroxycurcuminoids. The hydroxyl group of curcuminoids enhances the ability to generate ROS. 2-Hydroxycurcuminoid (HCC-7) strongly inhibited the growth of SW620 colon tumor cells with a GI₅₀ value of 7 μM, while the parent compounds, HCA and curcumin, displayed GI₅₀ values of 12 and 30 μM, respectively. HCC-7 was found to induce apoptosis through the reactive oxygen species-mitochondria pathway and cell cycle arrest at G2/M phase.     

Caudatin induces caspase-dependent apoptosis in human glioma cells with involvement of mitochondrial dysfunction and reactive oxygen species generation

Caudatin as one species of C-21 steroidal from Cynanchum bungei decne displays potential anticancer activity. However, the underlying mechanisms remain elusive. In the present study, the growth suppressive effect and mechanism of caudatin on human glioma U251 and U87 cells were evaluated in vitro. The results indicated that caudatin significantly inhibited U251 and U87 cell growth in both a time- and dose-dependent manner. Flow cytometry analysis revealed that caudatin-induced cell growth inhibition was achieved by induction of cell apoptosis, as convinced by the increase of Sub-G1 peak, PARP cleavage and activation of caspase-3, caspase-7 and caspase-9. Caudatin treatment also resulted in mitochondrial dysfunction which correlated with an imbalance of Bcl-2 family members. Further investigation revealed that caudatin triggered U251 cell apoptosis by inducing reactive oxygen species (ROS) generation through disturbing the redox homeostasis. Moreover, pretreatment of caspase inhibitors apparently weakens caudatin-induced cell killing, PARP cleavage and caspase activation and eventually reverses caudatin-mediated apoptosis. Importantly, caudatin significantly inhibited U251 tumour xenografts in vivo through induction of cell apoptosis involving the inhibition of cell proliferation and angiogenesis, which further validate its value in combating human glioma in vivo. Taken together, the results described above all suggest that caudatin inhibited human glioma cell growth by induction of caspase-dependent apoptosis with involvement of mitochondrial dysfunction and ROS generation.     

Bufalin induces autophagy-mediated cell death in human colon cancer cells through reactive oxygen species generation and JNK activation

Colorectal cancer is the second most common cause of cancer death in the world and about half of the patients with colorectal cancer require adjuvant therapy after surgical resection. Therefore, the eradication of cancer cells via chemotherapy constitutes a viable approach to treating patients with colorectal cancer. In this study, the effects of bufalin isolated from a traditional Chinese medicine were evaluated and characterized in HT-29 and Caco-2 human colon cancer cells. Contrary to its well-documented apoptosis-promoting activity in other cancer cells, bufalin did not cause caspase-dependent cell death in colon cancer cells, as indicated by the absence of significant early apoptosis as well as poly(ADP-ribose) polymerase and caspase-3 cleavage. Instead, bufalin activated an autophagy pathway, as characterized by the accumulation of LC3-II and the stimulation of autophagic flux. The induction of autophagy by bufalin was linked to the generation of reactive oxygen species (ROS). ROS activated autophagy via the c-Jun NH₂-terminal kinase (JNK). JNK activation increased expression of ATG5 and Beclin-1. ROS antioxidants (N-acetylcysteine and vitamin C), the JNK-specific inhibitor SP600125, and JNK2 siRNA attenuated bufalin-induced autophagy. Our findings unveil a novel mechanism of drug action by bufalin in colon cancer cells and open up the possibility of treating colorectal cancer through a ROS-dependent autophagy pathway.     

AGEs induces apoptosis and autophagy via reactive oxygen species in human periodontal ligament cells

The apoptosis of human periodontal ligament cells (HPDLCs) may be an important factor of the negative effect of advanced glycation end products (AGEs) on the periodontal tissue of diabetic patients. However, the pathways or potential effects of apoptosis in AGEs-treated HPDLCs have not been fully elucidated. Autophagy is closely related to apoptosis. Herein, we investigated the potential mechanism of apoptosis and autophagy in HPDLCs treated with AGEs via an in vitro model. We found that AGEs-treated HPDLCs showed a time- and concentration-dependent reduction in the cell survival rate. The mitochondrial-dependent apoptosis was induced in AGEs-treated HPDLCs, as confirmed by the mitochondrial membrane potential depolarization, decreased Bcl-2 expression, increased Bax expression, and increased caspase-3 and PARP cleavage. Autophagy was also induced in AGEs-treated HPDLCs, as indicated by the conversion of LC3-II/LC3-I and the presence of autophagosomes. Interestingly, our study results suggested that apoptosis and autophagy were related to reactive oxygen species (ROS) production. In addition, AGEs-induced autophagy acted as a latent factor in decreasing the generation of ROS in HPDLCs and protecting against the AGEs-induced apoptosis. In summary, our study shows that ROS are essential in AGEs-induced HPDLCs apoptosis and autophagy, which may be a molecular mechanism for the repairment of ROS-induced damage in HPDLCs treated with AGEs to promote cell survival. The present study might provide new insights into the therapeutic targeting of HPDLCs autophagy, which could be an additional strategy for periodontitis in patients with diabetes mellitus.     

A triazole-conjugated benzoxazone induces reactive oxygen species and promotes autophagic apoptosis in human lung cancer cells

Numerous approaches suggested that compounds with conjugated triazole moieties or benzoxazone pharmacores are effective to antagonize proliferation of human tumors. The current study reported that a synthetic triazole-conjugated benzoxazone, 4-((5-benzyl-1H-1,2,3-triazol-3-yl)-methyl)-7-methoxy-2H-benzo[b][1,4]-oxazin-3(4H)-one (BTO), inhibited growth rates of human non-small cell lung cancer cells. The cytotoxicity can be enhanced with increasing drug concentrations. More evidence supported that the induced reactive oxygen species lead to ultimate apoptotic cell death by recruiting autophagy. The mechanistic pathway as elucidated involved tumor suppressor p53 activation and LC3-1 conversion followed by PARP and procaspase-3 cleavage. Autophagy inhibition reverted apoptotic death and restored cell viabilities. BTO suppressed the development of A549 cell xenograft tumors by activating autophagy and apoptosis simultaneously. As an efficient tumor growth inhibitor with relatively small molecular weight, BTO is a viable addition to the existing list of lung cancer treatment.     

p53 regulates mitochondrial membrane potential through reactive oxygen species and induces cytochrome c-independent apoptosis blocked by Bcl-2.

Downstream mediators of p53 in apoptosis induction remain to be elucidated. We report that p53-induced apoptosis occurred in the absence of cytochrome c release into the cytosol. Although Bax was upregulated, it remained largely in the cytosol and there was no detectable translocation to the mitochondria. Bid was not activated as no cleavage could be detected. Thus, the absence of cytochrome c release may be due to the lack of Bax translocation to mitochondria and/or Bid inactivation. Nevertheless, p53-induced apoptosis is still caspase dependent because it could be abolished by z-VAD-fmk. To search for alternative downstream targets of p53, we detected production of reactive oxygen species (ROS) as well as mitochondrial membrane potential (Deltapsi). p53 induced ROS generation, which then caused a transient increase of Deltapsi followed by a decrease. Antioxidants could inhibit the alterations of Deltapsi, thereby preventing apoptosis. z-VAD-fmk was unable to abrogate Deltapsi elevation but inhibited Deltapsi decrease, indicating that Deltapsi elevation and its decrease are two independent events. Bcl-2 may abolish elevation as well as decrease of Deltapsi without interfering with ROS levels. Thus, the ROS-mediated disruption of Deltapsi constitutes a pivotal step in the apoptotic pathway of p53, and this pathway does not involve cytochrome c release.     

Heme oxygenase-1 attenuates the cisplatin-induced apoptosis of auditory cells via down-regulation of reactive oxygen species generation

Heme oxygenase-1 (HO-1), the rate-limiting enzyme of heme catabolism, is known to modulate various cellular functions, including cytokine production, cell proliferation, and apoptosis, in stress-related conditions. However, the role of HO-1 in the auditory system remains elusive. Herein, we demonstrate that pharmacologic induction of HO-1 along with catalytic activation significantly suppressed apoptosis of HEI-OC1 cells induced by cisplatin. Studies of ectopic expression of pcDNA3-HO-1 and siRNA of HO-1 further revealed the protective role of HO-1 against cisplatin in HEI-OC1 cells. Among the catabolic metabolites of HO-1, both carbon monoxide (CO) and bilirubin were directly involved in the protective role of HO-1 against cisplatin through inhibition of reactive oxygen species generation. Furthermore, pharmacological induction of HO-1 completely prevented the destruction of outer hair cell arrays by cisplatin through a CO-dependent mechanism in organotrophic culture of the rat primary organ of Corti explants. These results suggest that HO-1 may serve as a safeguard of auditory sensory hair cells against a variety of challenges of oxidative stress, including noise trauma, presbycusis, and ototoxic drugs, respectively.     

Apoptosis induction of U937 human leukemia cells by diallyl trisulfide induces through generation of reactive oxygen species

Background

Diallyl trisulfide (DATS) is one of the major constituents in garlic oil and has demonstrated various pharmacological activities, including antimicrobial, antihyperlipidemic, antithrombotic, and anticancer effects. However, the mechanisms of antiproliferative activity in leukemia cells are not fully understood. In this study, the apoptotic effects of DATS were investigated in human leukemia cells.

Results

Results of this study indicated that treatment with DATS resulted in significantly inhibited leukemia cell growth in a concentration- and time-dependent manner by induction of apoptosis. In U937 cells, DATS-induced apoptosis was correlated with down-regulation of Bcl-2, XIAP, and cIAP-1 protein levels, cleavage of Bid proteins, activation of caspases, and collapse of mitochondrial membrane potential. The data further demonstrated that DATS increased intracellular reactive oxygen species (ROS) generation, which was attenuated by pretreatment with antioxidant N-acetyl-l-cysteine (NAC), a scavenger of ROS. In addition, administration of NAC resulted in significant inhibition of DATS-induced apoptosis by inhibiting activation of caspases.

Conclusions

The present study reveals that the cytotoxicity caused by DATS is mediated by generation of ROS and subsequent activation of the ROS-dependent caspase pathway in U937 leukemia cells.     

A novel function for hydroxyproline oxidase in apoptosis through generation of reactive oxygen species

Proline and hydroxyproline are metabolized by distinct pathways. Proline is important for protein synthesis, as a source of glutamate, arginine, and tricarboxylic acid cycle intermediates, and for participating in a metabolic cycle that shuttles redox equivalents between mitochondria and cytosol. Hydroxyproline, in contrast, is not reutilized for protein synthesis. The first steps in the degradation of proline and hydroxyproline are catalyzed by proline oxidase (POX) and hydroxyproline oxidase (OH-POX), respectively. Because it is well documented that POX is induced by p53 and plays a role in apoptosis, we considered whether OH-POX also participates in the response to cytotoxic stress. In LoVo and RKO cells, which respond to adriamycin with a p53-mediated induction of POX and generation of reactive oxygen species, we found that adriamycin also induced OH-POX gene expression and markedly increased OH-POX catalytic activity, and this increase in activity was not observed in the cell lines HT29 and HCT15, which do not have a functional p53. We also observed an increase in reactive oxygen species generation and activation of caspase-9 with adriamycin in a hydroxyproline-dependent manner. Therefore, we hypothesize that OH-POX plays a role analogous to POX in growth regulation, ROS generation, and activation of the apoptotic cascade.     

Ornithine decarboxylase prevents dibenzoylmethane-induced apoptosis through repressing reactive oxygen species generation

Dibenzoylmethane (DBM) belongs to the flavonoid family and is a minor constituent of the root extract of licorice and the β-diketone analogue of curcumin. It exhibits antimutagenic, anticancer, and chemopreventive effects. Ornithine decarboxylase (ODC), the rate-limiting enzyme of the polyamine biosynthetic pathway, plays an important role in growth, proliferation, and transformation. Our previous studies showed ODC overexpression prevented etoposide-, paclitaxel-, and cisplatin-induced apoptosis. Here, we investigated one mechanism of DBM-induced apoptosis and the antiapoptotic effects of ODC during DBM treatment. We found that DBM induced apoptosis, promoted reactive oxygen species (ROS) generation, and disrupted the mitochondrial membrane potential (Δψ(m). N-acetylcysteine, a ROS scavenger, reduced DBM-induced apoptosis, which led to the loss of Δψ(m) due to reduced ROS. Overexpression of ODC in parental cells had the same effects as the ROS scavenger. The results demonstrated that DBM-induced apoptosis was a ROS-dependent pathway and ODC overexpression blocked DBM-induced apoptosis by inhibiting intracellular ROS production.     

Mebendazole induces apoptosis via Bcl-2 inactivation in chemoresistant melanoma cells

Most metastatic melanoma patients fail to respond to available therapy, underscoring the need for novel approaches to identify new effective treatments. In this study, we screened 2,000 compounds from the Spectrum Library at a concentration of 1 micromol/L using two chemoresistant melanoma cell lines (M-14 and SK-Mel-19) and a spontaneously immortalized, nontumorigenic melanocyte cell line (melan-a). We identified 10 compounds that inhibited the growth of the melanoma cells yet were largely nontoxic to melanocytes. Strikingly, 4 of the 10 compounds (mebendazole, albendazole, fenbendazole, and oxybendazole) are benzimidazoles, a class of structurally related, tubulin-disrupting drugs. Mebendazole was prioritized to further characterize its mechanism of melanoma growth inhibition based on its favorable pharmacokinetic profile. Our data reveal that mebendazole inhibits melanoma growth with an average IC(50) of 0.32 micromol/L and preferentially induces apoptosis in melanoma cells compared with melanocytes. The intrinsic apoptotic response is mediated through phosphorylation of Bcl-2, which occurs rapidly after treatment with mebendazole in melanoma cells but not in melanocytes. Phosphorylation of Bcl-2 in melanoma cells prevents its interaction with proapoptotic Bax, thereby promoting apoptosis. We further show that mebendazole-resistant melanocytes can be sensitized through reduction of Bcl-2 protein levels, showing the essential role of Bcl-2 in the cellular response to mebendazole-mediated tubulin disruption. Our results suggest that this screening approach is useful for identifying agents that show promise in the treatment of even chemoresistant melanoma and identifies mebendazole as a potent, melanoma-specific cytotoxic agent.     

Bcl-2 prevents hypoxia/reoxygenation-induced cell death through suppressed generation of reactive oxygen species and upregulation of Bcl-2 proteins

The function of bcl-2 in preventing cell death is well known, but the mechanisms whereby bcl-2 functions are not well characterized. One mechanism whereby bcl-2 is thought to function is by alleviating the effects of oxidative stress upon the cell. To examine whether Bcl-2 can protect cells against oxidative injury resulting from post-hypoxic reoxygenation (H/R), we subjected rat fibroblasts Rat-1 and their bcl-2 transfectants b5 to hypoxia (5% CO2, 95% N2) followed by reoxygenation (5% CO2, 95% air). The bcl-2 transfectants exhibited the cell viability superior to that of their parent non-transfectants upon treatment with reoxygenation after 24-, 48-, or 72-h hypoxia, but not upon normoxic serum-deprivation or upon serum-supplied hypoxic treatment alone. Thus bcl-2 transfection can prevent cell death of some types, which occurred during H/R but yet not appreciably until termination of hypoxia. The time-sequential events of H/R-induced cell death were shown to be executed via (1) reactive oxygen species (ROS) production at 1-12 h after H/R, (2) activation of caspases-1 and -3, at 1-3 h and 3-6 h after H/R, respectively, and (3) loss of mitochondrial membrane potential (DeltaPsi) at 3-12 h after H/R. These cell death-associated events were prevented entirely except caspase-1 activation by bcl-2 transfection, and were preceded by Bcl-2 upregulation which was executed as early as at 0-1 h after H/R for the bcl-2 transfectants but not their non-transfected counterpart cells. Thus upregulation of Bcl-2 proteins may play a role in prevention of H/R-induced diminishment of cell viability, but may be executed not yet during hypoxia itself and be actually operated as promptly as ready to go immediately after beginning of H/R, resulting in cytoprotection through blockage of either ROS generation, caspase-3 activation, or DeltaPsi decline.     

A specific cell-penetrating peptide induces apoptosis in SKOV3 cells by down-regulation of Bcl-2

Peptides are emerging as pharmaceutical agents in cancer therapy. The peptide, TLSGAFELSRDK (TLS) is a targeting ligand that can specifically triggers cellular uptake by binding to SKOV3 cells. Cell surface proteins and the C-terminal basic residues of the TLS are required for effective cell penetration, and the uptake process is energy-dependent. It inhibited the proliferation of SKOV3 cells and induced early-stage apoptosis by down-regulation of Bcl-2 expression mediated through a caspase-dependent pathway. The synergistic anti-proliferative effects of the peptide TLS and doxorubicin on SKOV3 cells were further investigated. Taken together, TLS, acting as a combination of a targeted ligand and a therapeutic agent, was a promising candidate for the development of peptide-based therapies in ovarian cancer.