Backgroundβ-lapachone (β-lap) is a bioreductive agent that is activated by the two-electron reductase NAD(P)H quinone oxidoreductase 1 (NQO1). Although β-lap has been reported to induce apoptosis in various cancer types in an NQO1-dependent manner, the signaling pathways by which β-lap causes apoptosis are poorly understood.Methodology/Principal Findingsβ-lap-induced apoptosis and related molecular signaling pathways in NQO1-negative and NQO1-overexpressing MDA-MB-231 cells were investigated. Pharmacological inhibitors or siRNAs against factors involved in β-lap-induced apoptosis were used to clarify the roles played by such factors in β-lap-activated apoptotic signaling pathways. β-lap leads to clonogenic cell death and apoptosis in an NQO1- dependent manner. Treatment of NQO1-overexpressing MDA-MB-231 cells with β-lap causes rapid disruption of mitochondrial membrane potential, nuclear translocation of AIF and Endo G from mitochondria, and subsequent caspase-independent apoptotic cell death. siRNAs targeting AIF and Endo G effectively attenuate β-lap-induced clonogenic and apoptotic cell death. Moreover, β-lap induces cleavage of Bax, which accumulates in mitochondria, coinciding with the observed changes in mitochondria membrane potential. Pretreatment with Salubrinal (Sal), an endoplasmic reticulum (ER) stress inhibitor, efficiently attenuates JNK activation caused by β-lap, and subsequent mitochondria-mediated cell death. In addition, β-lap-induced generation and mitochondrial translocation of cleaved Bax are efficiently blocked by JNK inhibition.Conclusions/SignificanceOur results indicate that β-lap triggers induction of endoplasmic reticulum (ER) stress, thereby leading to JNK activation and mitochondria-mediated apoptosis. The signaling pathways that we revealed in this study may significantly contribute to an improvement of NQO1-directed tumor therapies.

Background/Aim: Colon cancer is prone to distant metastases to other sites and the risk of recurrence is relatively high. Therefore, the identification of liver metastasis-related factors is important for the diagnosis or treatment of colon cancer. The aim of this study was to identify the metastasis-related factors that are differentially expressed in synchronous solitary liver metastasis compared to primary colon cancer. Materials and Methods: Tissues of primary colon cancer and associated with liver metastases of five patients were used for mass spectrometry. Identified proteins were validated by western blotting. The in silico analysis was performed using the STRING database and GeneMANIA. Results: We identified 58 differentially expressed proteins (DEPs), including 51 under-expressed and 7 over-expressed proteins among a total of 164 identified proteins. Major hubs of protein-protein networks were ACTC1, PRDX6, TPI1, and ALDH1A1. DEPs were located in the extracellular region and cytoplasm and were involved in the regulation of enzymatic activity. The metabolic process was significantly enriched in biological processes and an involvement in the KEGG pathway. Conclusion: These DEPs can potentially be used as biomarkers for the diagnosis of liver metastasis and they may provide a new strategy for developing anti-metastatic liver drugs in colon cancer patients.

Original ArticlePurpose The 8th edition of gallbladder cancer staging in the American Joint Committee on Cancer (AJCC) staging system changed the T and N categories. Materials and MethodsIn order to validate the new staging system, a total of 348 surgically resected gallbladder cancers were grouped based on the 8th edition of the T and N categories and compared with patients' survival. ResultsSignificant differences were noted between T1b-T2a (p=0.003) and T2b-T3 (p < 0.001) tumors, but not between Tis-T1a, T1a-T1b, and T2a-T2b tumors. However, significant survival differences were observed both by the overall and pair-wise (T1-T2, T2-T3) comparisons (all, p < 0.001) without dividing T1/T2 subcategories. When cases with ! 6 examined lymph nodes were evaluated, significant survival differences were observed among the entire comparison (p < 0.001) and pair-wise comparisons of N0-N1 (p=0.001) and N1-N2 (p=0.039) lesions. When cases without nodal dissection (NX) were additionally compared, significant survival differences were observed between patients with N0-NX (p=0.001) and NX-N1 (p < 0.001) lesions. ConclusionThe T category in the 8th edition of the AJCC staging system did not completely stratify the prognosis of patients with gallbladder cancer. Modification by eliminating T subcategories can better stratify the prognosis. In contrast, the N category clearly determines patients' survival with ! 6 examined lymph nodes. The survival time in patients of gallbladder cancers without nodal dissection is between N0 and N1 cases. Therefore, close postoperative followed up is recommended for those patients.

Backgroundβ-lapachone (β-lap), has been known to cause NQO1-dependnet death in cancer cells and sensitize cancer cells to ionizing radiation (IR). We investigated the mechanisms underlying the radiosensitization caused by β-lap.Methodology/Principal Findingsβ-lap enhanced the effect of IR to cause clonogenic cells in NQO1+-MDA-MB-231 cells but not in NQO1−-MDA-MB-231 cells. β-lap caused apoptosis only in NQO1+ cells and not in NQO1− cells and it markedly increased IR-induced apoptosis only in NQO1+ cells. Combined treatment of NQO1+ cells induced ROS generation, triggered ER stress and stimulated activation of ERK and JNK. Inhibition of ROS generation by NAC effectively attenuated the activation of ERK and JNK, induction of ER stress, and subsequent apoptosis. Importantly, inhibition of ERK abolished ROS generation and ER stress, whereas inhibition of JNK did not, indicating that positive feedback regulation between ERK activation and ROS generation triggers ER stress in response to combined treatment. Furthermore, prevention of ER stress completely blocked combination treatment-induced JNK activation and subsequent apoptotic cell death. In addition, combined treatment efficiently induced the mitochondrial translocation of cleaved Bax, disrupted mitochondrial membrane potential, and the nuclear translocation of AIF, all of which were efficiently blocked by a JNK inhibitor. Caspases 3, 8 and 9 were activated by combined treatment but inhibition of these caspases did not abolish apoptosis indicating caspase activation played a minor role in the induction of apoptosis.Conclusions/Significanceβ-lap causes NQO1-dependent radiosensitization of cancer cells. When NQO1+ cells are treated with combination of IR and β-lap, positive feedback regulation between ERK and ROS leads to ER stress causing JNK activation and mitochondrial translocation of cleaved Bax. The resultant decrease in mitochondrial membrane leads to translocation of AIF and apoptosis.

The purpose of the present investigation was to study the effects of ionizing radiation on endothelial cells derived from diverse normal tissues. We first compared the effects of radiation on clonogenic survival and tube formation of endothelial cells, and then investigated the molecular signaling pathways involved in endothelial cell survival and angiogenesis. Among the different endothelial cells studied, human hepatic sinusoidal endothelial cells (HHSECs) were the most radio-resistant and human dermal microvascular endothelial cells were the most radio-sensitive. The radio-resistance of HHSECs was related to adenosine monophosphate-activated protein kinase and p38 mitogen-activated protein kinase-mediated expression of MMP-2 and VEGFR-2, whereas the increased radio-sensitivity of HDMECs was related to extracellular signal-regulated kinase-mediated generation of angiostatin. These observations demonstrate that there are distinct differences in the radiation responses of normal endothelial cells obtained from diverse organs, which may provide important clues for protection of normal tissue from radiation exposure.

BACKGROUND AND PURPOSE2,5-diaziridinyl-3-(hydroxymethyl)-6-methyl-1,4-benzoquinone (RH1) is a bioreductive agent that is activated by the two-electron reductase NAD(P)H quinone oxidoreductase 1 (NQO1). Although the cytotoxic efficacy of RH1 against tumours has been studied extensively, the molecular mechanisms underlying this anti-cancer activity have not yet been fully elucidated. EXPERIMENTAL APPROACH2,5-diaziridinyl-3-(hydroxymethyl)-6-methyl-1,4-benzoquinone-induced apoptosis and related signalling pathways in NQO1-negative and NQO1-overexpressing cells were evaluated. The role of p53 in RH1-induced cell death was investigated using parental and p53-deficient RKO human colorectal cancer cells by assaying clonogenic cell survival. Specific inhibitors and siRNAs targeting factors involved in RH1-induced apoptosis were used to clarify the roles played by such factors in RH1-activated apoptotic signalling pathways. KEY RESULTS2,5-diaziridinyl-3-(hydroxymethyl)-6-methyl-1,4-benzoquinone induced apoptosis and clonogenic death, dependent on NQO1 and p53. Treatment of NQO1-overexpressing cells with RH1 caused rapid disruption of mitochondrial membrane potential, nuclear translocation of apoptosis-inducing factor (AIF) and endonuclease G (Endo G) from mitochondria, and subsequent caspase-independent apoptotic cell death. siRNA targeting AIF and Endo G effectively attenuated RH1-induced apoptotic cell death. Moreover, RH1 induced cleavage of Bax, which targets mitochondria. RH1 significantly activated the c-Jun N-terminal kinase (JNK) pathway, and inhibition of this pathway suppressed RH1-induced mitochondria-mediated apoptosis. RH1-induced generation and mitochondrial translocation of cleaved Bax were blocked by the JNK inhibitor, SP600125. Inhibition of JNK with SP600125 attenuated the mitochondrial translocation of JNK. CONCLUSIONS AND IMPLICATIONS2,5-diaziridinyl-3-(hydroxymethyl)-6-methyl-1,4-benzoquinone activated JNK, resulting in mitochondria-mediated apoptotic cell death that was NQO1-dependent. AbbreviationsAIF, apoptosis-inducing factor; DiOC6(3), 3

Objective: To investigate whether the Minimal Clinically Important Difference (MCID) clinically defines improvement of Berg Balance Scale (BBS) scores in people with acute stroke in response to rehabilitation. Design: Retrospective study. Methods: Seventy-three participants with acute stroke participated in the study. Balance evaluation was performed using the BBS. All patients received rehabilitation with physical therapy for 4 weeks, 5 times a week, for 2 hours and 20 minutes a day. An anchor-based approach using the clinical global impression was used to determine the MCID of the BBS. The MCID was used to define the minimum change in the BBS total score (postintervention-preintervention) that was needed to perceive at least a 3-point improvement on the global rating of change. Receiver operating characteristic (ROC) curves was used to define the cutoff values of the optimal MCID of the BBS in order to discriminate between improvement and no improvement groups. Results: The optimal MCID cutoff point for the BBS change scores was 12.5 points for males with a sensitivity (Sn) of 0.62 and a specificity (Sp) of 0.89, and 12.5 points for females with a Sn of 0.69 and Sp of 0.85. The area under the curve of the ROC curve for all participants were 0.84 (95% confidence interval [CI], 0.72; 0.95, p<0.001), and 0.89 (95% CI, 0.77; 1.00, p<0.001), respectively. Conclusions: The MCID for improvement in balance as measured by the BBS was 13.5 points, indicating that the MCID does clinically detect changes in balance abilities in persons with stroke.

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