• Reproductive and Developmental Biology
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• v.35 no.3
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• pp.209-214
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• 2011

Although the function and utility of RNA transcripts derived from matured spermatozoa remains unclear, they might play important roles in the establishment of a paternal genome and subsequently embryo development. Herein, we investigated the expression of X-chromosome linked RNA transcripts in matured bovine spermatozoa. The total RNA was extracted from the matured spermatozoa, and then converted to cDNA. Autosomal genes (ACT-${\beta}$ and H-2A) and X-chromosome linked genes (ANT3, HPRT, MeCP2, RPS4X, XIAP, XIST and ZFX) were analyzed for the characterization of X-chromosome linked RNA transcripts and compared to female fibroblasts by RT-PCR. The transcripts of autosomal genes (ACT-${\beta}$ and H2A) and X-chromosome linked genes (ANT3, HPRT, MeCP2, RPS4X and ZFX) were not detected in spermatozoa. However, XIAP (X-linked inhibitor of apoptosis protein) and XIST (X-chromosome inactive-specific transcript, a kind of paternal imprinted gene) transcripts were detected in spermatozoa, and relative levels of XIAP and XIST transcripts were similar and 0.5-fold lower when compared to female fibroblasts, respectively. Based on the findings, it is summarized that the presence of RNA transcripts of XIAP and XIST in the isolated spermatozoa may imply their role in inhibition of apoptosis and induction of X-chromosome inactivation in embryo development.

• Asian Pacific Journal of Cancer Prevention
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• v.15 no.23
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• pp.10483-10487
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• 2015

Heptaphylline derivatives are carbazoles in Clausena harmandiana, a medicinal plant that is utilized for headache, stomach ache, and other treatments of illness. The present study examined the effects of heptaphylline and 7-methoxyheptaphylline on apoptosis of human colon adenocarcinoma cells (HT-29 cell line). Quantification of cell viability was performed using cell proliferation assay (MTT assay) and of protein expression through immunoblotting. The results showed that only heptaphylline, but not 7-methoxyheptaphylline, significantly significantly activated cleaved of caspase-3 and poly (ADP-ribose) polymerase (PARP-1) which resulted in HT-29 cell death. We found that heptaphylline activated BH3 interacting-domain death agonist (Bid) and Bak, proapoptotic proteins. In contrast, it suppressed X-linked inhibitor-of-apoptosis protein (XIAP), Bcl-xL and survivin, inhibitors of apoptosis. In addition, heptaphylline inhibited activation of NF-${\kappa}B$/p65 (rel), a regulator of apoptotic regulating proteins by suppressing the activation of Akt and $IKK{\alpha}$, upstream regulators of p65. The findings suggested that heptaphylline induces apoptosis in human colon adenocarcinoma cells.

• Biomedical Science Letters
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• v.29 no.2
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• pp.66-74
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• 2023

Triglyceride (TG) accumulation can cause monocytic death and suppress innate immunity. However, the signaling pathways involved in this phenomenon are not fully understood. This study aimed to examine whether inducible nitric oxide synthase (iNOS) is involved in the TG-induced death of THP-1 monocytes. Results showed that iNOS was upregulated in TG-treated THP-1 monocytes, and iNOS inhibition blocked TG-induced monocytic death. In addition, TG-induced poly (ADP-ribose) polymerase (PARP) cleavage and caspase-3 and -7 activation were suppressed by iNOS inhibition. Furthermore, the expression of X-linked inhibitor of apoptosis protein (XIAP) and survivin, which inhibit caspase-3 and -7, was reduced in TG-treated THP-1 monocytes, but iNOS inhibition recovered the TG-induced downregulation of XIAP and survivin expression. Considering that TG-induced monocytic death is triggered by caspase2 and -8, we investigated whether caspase-2 and -8 are linked to the TG-induced expression of iNOS in THP-1 monocytes. When the activities of caspase-2 and -8 were inhibited by specific inhibitors, the TG-induced upregulation of iNOS and downregulation of XIAP and survivin were restored in THP-1 monocytes. These results suggest that TG-induced monocytic death is mediated by the caspase-2/caspase-8/iNOS/XIAP and survivin/executioner caspase/PARP pathways.

• Asian Pacific Journal of Cancer Prevention
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• v.16 no.6
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• pp.2453-2458
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• 2015

Background: X-linked inhibitor of apoptosis protein (XIAP) associated factor 1 (XAF1) exhibits aberrantly low or absent expression in various human malignancies, closely associated with anti-apoptosis and overgrowth of cancer cells. However, limited attention has been directed towards the contribution of XAF1 to invasion, apoptosis, and cisplatin (DDP)-resistance of epithelial ovarian cancer (EOC) cells. This study aimed to evaluate the potential effects of XAF1 on invasion, cell cycle, apoptosis, and cisplatin-resistance by overexpressing XAF1 in SKOV-3 and SKOV-3/DDP cells. Methods and Results: The pEGFP-C1-XAF1 plasmid was transfected into SKOV-3 and SKOV-3/DDP cells, and the expression of XAF1 at both mRNA and protein levels was analyzed by reverse transcription-PCR and Western blotting. Overexpression of XAF1 suppressed XIAP expression in both SKOV-3 and SKOV-3/DDP cells. Transwell invasion assays demonstrated that XAF1 exerted a strong anti-invasive effect in XAF1-overexpressing cells. Moreover, flow cytometry analysis revealed that XAF1 overexpression arrested the cell cycle at G0/G1 phase, and cell apoptosis analysis showed that overexpression of XAF1 enhanced apoptosis of SKOV-3 and SKOV-3/DDP cells apparently by activating caspase-9 and caspase-3. Furthermore, MTT assay confirmed a dose-dependent inhibitory effect of cisplatin in the tested tumor cells, and overexpression of XAF1 increased the sensitivity of SKOV-3 and SKOV-3/DDP cells to cisplatin-mediated antiproliferative effects. Conclusions: In summary, our data indicated that overexpression of XAF1 could suppress XIAP expression, inhibit invasion, arrest cell cycle, promote apoptosis, and confer cisplatin-sensitivity in SKOV-3 and SKOV-3/DDP cells. Therefore, XAF1 may be further assessed as a potential target for the treatment of both cisplatin-resistant and non-resistant EOCs.

• Journal of Chest Surgery
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• v.45 no.1
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• pp.1-10
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• 2012

Background: ${\alpha}$-Lipoic acid (${\alpha}$-LA) has been studied as an anticancer agent as well as a therapeutic agent for diabetes and obesity. We performed this study to evaluate the anticancer effects and mechanisms of ${\alpha}$-LA in a lung cancer cell line, A549. Materials and Methods: ${\alpha}$-LA-induced apoptosis of A549 cells was detected by fluorescence-activated cell sorting analysis and a DNA fragmentation assay. Expression of apoptosis-related genes was analyzed by western blot and reverse transcription.polymerase chain reaction analyses. Results: ${\alpha}$-LA induced apoptosis and DNA fragmentation in A549 cells in a dose- and time-dependent manner. ${\alpha}$-LA increased caspase activity and the degradation of poly (ADP-ribose) polymerase. It induced expression of endoplasmic reticulum (ER) stress-related genes, such as glucose-regulated protein 78, C/EBP-homologous protein, and the short form of X-box binding protein-1, and decreased expression of the anti-apoptotic protein, X-linked inhibitor of apoptosis protein. Reactive oxygen species (ROS) production was induced by ${\alpha}$-LA, and the antioxidant N-acetyl-L-cysteine decreased the ${\alpha}$-LA-induced increase in expression of apoptosis and ER stress-related proteins. Conclusion: ${\alpha}$-LA induced ER stress-mediated apoptosis in A549 cells via ROS. ${\alpha}$-LA may therefore be clinically useful for treating lung cancer.

• Tuberculosis and Respiratory Diseases
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• v.54 no.5
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• pp.495-509
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• 2003

Background : Neutrophil-mediated inflammation is usually self-limiting, because neutrophils have a remarkably short life span. Prolonged neutrophil survival, which is caused by decreased spontaneous apoptosis, leads to persistent inflammation in sepsis. Because many inflammatory cytokines, which generate signals that delay apoptosis, are regulated by nuclear factor-${\kappa}B$ transcription factor, we hypothesized that nuclear factor-${\kappa}B$ might be related to the reduced neutrophil apoptosis observed in sepsis. Methods : Neutrophils of healthy volunteers and sepsis patients were freshly isolated from venous blood. Neutrophil apoptosis was assayed with two approaches : by counting apoptotic cells under a microscope and by flow cytometry using Annexin V. The activity of nuclear factor-${\kappa}B$ was assessed by immunofluorescent staining or electrophoretic mobility shift assay. Expression of X-linked inhibitor of apoptosis was measured by western blot assay. Results : We confirmed reduced spontaneous neutrophil apoptosis in patients with sepsis. The number of apoptotic neutrophils in patients with sepsis increased to the level of that in healthy controls after cycloheximide treatment, suggesting that decreased spontaneous neutrophil apoptosis is dependent on de novo protein synthesis. In patients with sepsis, basal neutrophil nuclear factor-${\kappa}B$ was activated compared to the level in healthy controls. Moreover, a blockade of nuclear factor-${\kappa}B$ activity reversed the decreased spontaneous neutrophil apoptosis in sepsis patients. Meanwhile, X-linked inhibition of apoptosis expression, which is regulated by nuclear factor-${\kappa}B$, decreased 24 hours after incubation in healthy persons, but persisted for 24 hours in patients with sepsis. Conclusion : These observations suggest that the reduced spontaneous neutrophil apoptosis observed in patients with sepsis may be related to the induction of survival protein by nuclear factor-${\kappa}B$.

• Journal of Acupuncture Research
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• v.31 no.2
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• pp.87-98
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• 2014

Objectives : We investigated whether snake venom toxin(SVT) from Vipera lebetina turanica sensitizes HT29 human epithelial colorectal cancer cells to tumor necrosis factor(TNF)-related apoptosis-inducing ligand(TRAIL) induced apoptosis in cancer cells. Methods : Cell viability assay was used to assess the inhibitory effect of TRAIL on cell growth of HT29 human colorectal cancer cells. And 6-diamidino-2-phenylindole(DAPI), terminal deoxynucleotidyl transferase mediated dUTP nick end labeling assay(TUNEL) staining assay were used to evaluate cell-apoptosis. Western blot analysis were conducted to observe apoptosis related proteins and death receptor. To assess whether the synergized inhibitory effect of SVT and TRAIL on reactive oxygen species(ROS) generation was reversed by strong anti-oxidative agent. Results : SVT with TRAIL inhibited HT29 cell growth different from TRAIL alone. Consistent with cell growth inhibition, the expression of TRAIL receptors; Expression of death receptor(DR)4 and DR5 was significantly increased and intrinsic pro-apoptotic cleaved caspase-3, -9 was subsequently increased together with increase of Bax/Bcl-2 ratio and extrinsic pro-apototic caspase-8 was also activated. In addition, the expression of anti-apoptotic survival proteins, a marker of TRAIL resistance(eg, cFLIP, survivin, X-linked inhibitor of apoptosis protein(XIAP) and Bcl-2) was suppressed by the combination treatment of SVT and TRAIL. Pretreatment with the ROS scavenger N-acetylcysteine abolished the SVT and TRAIL-induced upregulation of DR4 and DR5 expression and expression of the intrinsic pro-apoptotic caspase-3 and-9. Conclusion : The collective results suggest that SVT facilitates TRAIL-induced apoptosis in $HT_{29}$ human epithelial colorectal cancer cells through up-regulation of the TRAIL receptors; DR4 and DR5 and consecutive induction of bilateral apoptosis via regulating apoptosis related proteins.

• Animal cells and systems
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• v.12 no.1
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• pp.15-21
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• 2008

Nitric oxide(NO) has been known to play important roles in numerous physiologic processes including neurotransmission, vasorelaxation, and cellular apoptosis. Using a mouse cDNA gene chip, we examined expression patterns and time course of NO-dependent genes in mouse macrophage RAW264.7 cells. Genes shown to be upregulated more than two fold or at least at two serial time points were further selected and validated by RT-PCR. Finally, 81 selected genes were classified by function as signaling, apoptosis, inflammation, transcription, translation, ionic homeostasis and metabolism. Among those, genes related with signaling, apoptosis and inflammation, such as guanylate cyclase 1, soluble, alpha3(Gucy1a3); protein kinase C, alpha($Pkc{\alpha}$); lymphocyte protein tyrosine kinase(Lck); BCL2/adenovirus E1B 19 kDa-interacting protein(Bnip3); apoptotic protease activating factor 1(Apaf1); X-linked inhibitor of apoptosis(Xiap); cyclin G1(Ccng1); chemokine(C-C motif) ligand 4(Ccl4); B cell translocation gene 2, anti-proliferative(Btg2); lysozyme 2(Lyz2); secreted phosphoprotein 1(Spp1); heme oxygenase(decycling) 1(Hmox1); CD14 antigen(Cd14); and granulin(Grn) may play important roles in NO-dependent responses in murine macrophages.

• Journal of Life Science
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• v.21 no.8
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• pp.1109-1119
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• 2011

In the present study, the pro-apoptotic effects of methanol extract of Prunus mume fruits (MEPM) in human leukemia U937 cells were investigated. It was found that exposure to MEPM resulted in growth inhibition in a concentration-dependent manner by inducing apoptosis. The induction of apoptotic cell death in U937 cells by MEPM was correlated with a down-regulation of inhibitor of apoptosis protein (IAP) family, such as X-linked inhibitor of apoptosis protein (XIAP) and survivin, anti-apoptotic Bcl-2, up-regulation of FasL and cleavage of Bid. MEPM treatment also induced the proteolytic activation of caspase-3, caspase-8 and caspase-9, and degradation of caspase-3 substrate proteins, such as poly (ADP-ribose) polymerase (PARP) and ${\beta}$-catenin. In addition, apoptotic cell death induced by MEPM was significantly inhibited by z-DEVD-fmk, a caspase-3 specific inhibitor, which demonstrates the important role of caspase-3 in the apoptotic process by MEPM in U937 cells. Taken together, these findings suggest that P. mume extracts may be a potential chemotherapeutic agent for the control of human leukemia cells and further studies will be needed to identify the active compounds.

• Asian Pacific Journal of Cancer Prevention
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• v.13 no.4
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• pp.1471-1476
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• 2012

Background and Aim: Currently available systemic therapies for malignant melanoma produce low response rates in patients, and more effective treatment modalities are clearly needed. The tumor necrosis factor (TNF)-related apoptosis-inducing ligand has a significant impact on therapy for patients with X-linked inhibitor of apoptosis protein-downregulation malignant melanoma. The primary objective of this study was to assess its therapeutic potential. Materials and Methods: We employed a conditionally replicating oncolytic adenoviral vector, named CRAd5.TRAIL/siXIAP, with the characteristics of over-expression of the therapeutic gene TRAIL and downregulation of XIAP in one vector. B16F10-luc cells were employed to detect anti-tumor activity of CRAd5.TRAIL/siXIAP in vitro and in vivo. Results: CRAd5.TRAIL/siXIAP enhanced caspase-8 activation and caspase-3 maturation in B16F10 cells in vitro. Furthermore, it more effectively infected and killed melanoma cells in vitro and in vivo than other adenoviruses. Conclusion: Taken together, the combination of upregulation of TRAIL and downregulation of siXIAP with one oncolytic adenoviral vector holds promise for development of an effective therapy for melanomas and other common cancers.