Chronic exposure of human bronchial epithelial BEAS-2B cells to hexavalent chromium [Cr(VI)] causes malignant cell transformation. Sirtuin-3 (SIRT3) regulates mitochondrial adaptive response to stress, such as metabolic reprogramming and antioxidant defense mechanisms. In Cr(VI)-transformed cells, SIRT3 was upregulated and mitochondrial adenosine triphosphate (ATP) production and proton leak were reduced. Knockdown of SIRT3 by its shRNA further decreased mitochondrial ATP production, proton leak, mitochondrial mass, and mitochondrial membrane potential, indicating that SIRT3 positively regulates mitochondrial oxidative phosphorylation and maintenance of mitochondrial integrity. Mitophagy is critical to maintain proper cellular functions. In Cr(VI)-transformed cells expressions of Pink 1 and Parkin, two mitophagy proteins, were elevated, and mitophagy remained similar as that in passage-matched normal BEAS-2B cells, indicating that in -Cr(VI)-transformed cells mitophagy is suppressed. Knockdown of SIRT3 induced mitophagy, suggesting that SIRT3 plays an important role in mitophagy suppression of Cr(VI)-transformed cells. In Cr(VI)-transformed cells, nuclear factor (erythroid-derived 2)-like 2 (Nrf2) was constitutively activated, and protein levels of p62 and p-p62Ser349 were elevated. Knockdown of SIRT3 or treatment with carbonyl cyanide m-chlorophenyl hydrazone (CCCP) decreased the binding of p-p62Ser349 to Keap1, resulting in increased binding of Keap1 to Nrf2 and consequently reduced Nrf2 activation. The results from CHIP assay showed that in Cr(VI)-transformed cells binding of Nrf2 to antioxidant response element (ARE) of SIRT3 gene promoter was dramatically increased. Knockdown of SIRT3 suppressed cell proliferation and tumorigenesis of Cr(VI)-transformed cells. Overexpression of SIRT3 in normal BEAS-2B cells exhibited mitophagy suppression phenotype and increased cell proliferation and tumorigenesis. The present study demonstrated that upregulation of SIRT3 causes mitophagy suppression and plays an important role in cell survival and tumorigenesis of Cr(VI)-transformed cells.

Prostate cancer is the most prevalent type of cancer in men. The etiology of prostate cancer development and the mechanisms underlying androgen-independent progression remains to be further investigated. There are many known targets for prostate cancer therapy including the androgen receptor (AR) axis, but resistance eventually develops in advanced disease suggesting the need to better understand mechanisms of resistance and consideration of multi-targeted therapy. Mechanisms contributing to resistance may include gene amplifications, gene mutations, AR splice variants, and changes in expression of androgen receptor co-regulatory proteins. Given the limitations of approved therapies, further study of additional potential targets is warranted. This review focuses on the roles of autophagy pathway, p62, Yes-associated protein (YAP), cancer stem cells, and epigenetics. Therapies targeting these potential mechanisms of resistance may interact with currently approved therapies either additively or synergistically. Thus, the study of combination therapy against multiple targets may be critically important to achieve more impact against lethal forms of prostate cancer resistant to all approved current therapies.

To test the hypothesis that p62 is an optimal target for autophagy inhibition and Verteporfin, a clinically available drug approved by FDA to treat macular degeneration that inhibits autophagy by targeting p62 protein, can be developed clinically to improve therapy for advanced prostate cancer. Results showed that Verteporfin inhibited cell growth and colony formation in PC-3 cells. Verteporfin generated crosslinked p62 oligomers, resulting in inhibition of autophagy and constitutive activation of Nrf2 as well as its target proteins, Bcl-2 and TNF-α. In normal prostate epithelial cells, forced expression of p62 caused constitutive Nrf2 activation and development of apoptosis resistance and Verteporfin treatment exhibited inhibitory effects. Verteporfin treatments also inhibited starvation-induced autophagic influx of these cells. Verteporfin inhibited tumorigenesis of both normal prostate epithelial cells with p62 expression and prostate cancer cells and decreased p62, constitutive Nrf2, and Bcl-xL in xenograft tumor tissues, indicating that p62 can be developed as a drug target against prostate cancer. So p62 has a high potential to be developed as a therapeutic target. Verteporfin represents a prototypical agent with therapeutic potential against prostate cancer through inhibition of autophagy by a novel mechanism of p62 inhibition. Citation Format: Lei Wang, Donghern Kim, James Wise, Xianglin Shi, Zhuo Zhang, Robert S. DiPaola. Verteporfin targets p62 in prostate cancer therapeutics [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 2895.

Retraction: [Wang, L, Kim, D, Wise, JT, Shi, X, Zhang, Z, DiPaola, RS. p62 as a therapeutic target for inhibition of autophagy in prostate cancer. The Prostate. 2018; 78: 390 to 400. https://doi.org/10.1002/pros.23483] The above article from The Prostate, published online on 26 January 2018 in Wiley Online Library (wileyonlinelibrary.com) and in Volume 78, pp. 390‐400 has been retracted by agreement between the journal Editor in Chief, John Isaacs, and Wiley Periodicals, Inc. The retraction has been agreed due to an investigation by the University of Kentucky which identified data fabrication as part of a larger pattern of research misconduct by co‐authors Kim, Shi, and Zhang.

Cr(VI) is a well known environmental carcinogen, but its mechanism of action and the measures required to mitigate its effects remain to be investigated. Our previous studies showed that exposure of human bronchial epithelial (BEAS-2B) cells to Cr(VI) caused malignant transformation, that these transformed cells progressed through tumorigenesis, and that luteolin, a natural compound, inhibited both of these processes. The present study investigates the underlying mechanisms by which luteolin protects cells against Cr(VI)-induced transformation and tumorigenesis. The present study shows that luteolin activates inducible Nrf2 to inhibit Cr(VI)-generated reactive oxygen species (ROS) in normal BEAS-2B cells. The decreased ROS level is likely responsible for the protective effect of luteolin against Cr(VI)-induced malignant cell transformation in normal cells. By contrast, in cells that have been transformed by Cr(VI), Nrf2 is constitutively activated, and its target proteins, heme oxygenase 1 (HO-1), NAD(P)H:quinone oxidoreductase 1 (NQO1), and superoxide dismutase 1/2 (SOD1/SOD2) are all constitutively activated, and ROS levels are low. Bcl-2, an anti-apoptotic protein and target protein of Nrf2 is elevated. Cr(VI)-transformed BEAS-2B cells develop apoptosis resistance, increasing the survival of these transformed cells. Luteolin decreases interactions between Nrf2 and the antioxidant response element sites of its target anti-apoptotic and antioxidant proteins, Bcl-2, Bcl-XL, and HO-1, which results in decreased constitutive Nrf2 activation. The decreased constitutive Nrf2 activation, decrease in Nrf2 target proteins and consequent apoptosis resistance by luteolin are possible mechanisms that mediate the protective effect of luteolin in Cr(VI)-transformed cells.

Hexavalent chromium (Cr(VI)) compounds are confirmed human carcinogens for lung cancer. Our previous studies has demonstrated that chronic exposure of human bronchial epithelial BEAS-2B cells to low dose of Cr(VI) causes malignant cell transformation. The acquisition of cancer stem cell-like properties is involved in the initiation of cancers. The present study has observed that a small population of cancer stem-like cells (BEAS-2B-Cr-CSC) exists in the Cr(VI)-transformed cells (BEAS-2B-Cr). Those BEAS-2B-Cr-CSC exhibit extremely reduced capability of generating reactive oxygen species (ROS) and apoptosis resistance. BEAS-2B-Cr-CSC are metabolic inactive as evidenced by reductions in oxygen consumption, glucose uptake, ATP production, and lactate production. Most importantly, BEAS-2B-Cr-CSC are more tumorigenic with high levels of cell self-renewal genes, Notch1 and p21. Further study has found that fructose-1,6-bisphosphatase (FBP1), an rate-limiting enzyme driving glyconeogenesis, was lost in BEAS-2B-Cr-CSC. Forced expression of FBP1 in BEAS-2B-Cr-CSC restored ROS generation, resulting in increased apoptosis, leading to inhibition of tumorigenesis. In summary, the present study suggests that loss of FBP1 is a critical event in tumorigenesis of Cr(VI)-transformed cells.

Apigenin is a natural flavonoid which possesses multiple anti-cancer properties such as anti-proliferation, anti-inflammation, and anti-metastasis in many types of cancers including colorectal cancer. Neural precursor cell expressed developmentally downregulated 9 (NEDD9) is a multi-domain scaffolding protein of the Cas family which has been shown to correlate with cancer metastasis and progression. The present study investigates the role of NEDD9 in apigenin-inhibited cell migration, invasion, and metastasis of colorectal adenocarcinoma DLD1 and SW480 cells. The results show that knockdown of NEDD9 inhibited cell migration, invasion, and metastasis and that overexpression of NEDD9 promoted cell migration and invasion of DLD1 cells and SW4890 cells. Apigenin treatment attenuated NEDD9 expression at protein level, resulting in reduced phosphorylations of FAK, Src, and Akt, leading to inhibition on cell migration, invasion, and metastasis of both DLD1 and SW480 cells. The present study has demonstrated that apigenin inhibits cell migration, invasion, and metastasis through NEDD9/Src/Akt cascade in colorectal cancer cells. NEDD9 may function as a biomarker for evaluation of cancer aggressiveness and for selection of therapeutic drugs against cancer progression.

Hexavalent chromium (Cr(VI))-containing compounds are well established environmental carcinogens. Most mechanistic investigations of Cr(VI)-induced carcinogenesis focus on oxidative stress and various cellular responses, leading to malignant cell transformation or the first stage of metal-induced carcinogenesis. The development of malignantly transformed cells into tumors that require angiogenesis is the second stage. This study focuses on the second stage, in particular, the role of EGF receptor (EGFR) signaling in angiogenesis and tumorigenesis of Cr(VI)-transformed cells. Our preliminary studies have shown that EGFR is constitutively activated in Cr(VI)-transformed cells, in lung tissue from Cr(VI)-exposed animals, and in lung tumor tissue from a non-smoking worker occupationally exposed to Cr(VI) for 19 years. Using in vitro and in vivo models, the present study has investigated the role of EGFR in angiogenesis of Cr(VI)-transformed cells. The results show that Cr(VI)-transformed cells are angiogenic. Hypoxia-inducible factor-1α, pro-angiogenic protein matrix metalloproteinase 1, and VEGF are all highly expressed in Cr(VI)-transformed cells, in lung tissue from animals exposed to Cr(VI), and in lung tumor tissue from a non-smoking worker occupationally exposed to Cr(VI) for 19 years. p38 MAPK is also activated in Cr(VI)-transformed cells and in human lung tumor tissue. Inhibition of EGFR reduces p38 MAPK, resulting in decreased expression of hypoxia-inducible factor-1α, metalloproteinase 1, and VEGF, leading to suppressions of angiogenesis and tumorigenesis. Overall, the present study has demonstrated that EGFR plays an important role in angiogenesis and tumorigenesis of Cr(VI)-transformed cells.

Nickel compounds are known as human carcinogens. Chronic environmental exposure to nickel is a worldwide health concern. Although the mechanisms of nickel-induced carcinogenesis are not well understood, recent studies suggest that stem cells/cancer stem cells are likely important targets. This study examines the role of cancer stem cells in nickel-induced cell transformation. The nontransformed human bronchial epithelial cell line (Beas-2B) was chronically exposed to nickel chloride for 12 months to induce cell transformation. Nickel induced Beas-2B cell transformation, and cancer stem-like cells were enriched in nickel-transformed cell (BNiT) population. The BNiT cancer stem-like cells demonstrated enhanced self-renewal and distinctive differentiation properties. In vivo tumorigenesis studies show that BNiT cancer stem-like cells possess a high tumor-initiating capability. It was also demonstrated that superoxide dismutase 1 was involved in the accumulation of cancer stem-like cells; the regulation of superoxide dismutase 1 expression was different in transformed stem-like cells and nontransformed. Overall, the accumulation of stem-like cells and their enhanced stemness functions contribute to nickel-induced tumorigenesis. Our study provides additional insight into the mechanisms by which metals or other chemicals can induce carcinogenesis.

Hexavalent chromium [Cr(VI)] is an important human carcinogen associated with pulmonary diseases and lung cancer. Inhibition of Cr(VI)-induced carcinogenesis by a dietary antioxidant is a novel approach. Quercetin is one of the most abundant dietary flavonoids widely present in many fruits and vegetables, possesses potent antioxidant and anticancer properties. MicroRNA-21 (miR-21) is a key oncomiR significantly elevated in the majority of human cancers that exerts its oncogenic activity by targeting the tumor suppressor gene programmed cell death 4 (PDCD4). The present study examined the effect of quercetin on the inhibition of Cr(VI)-induced malignant cell transformation and the role of miR-21-PDCD4 signaling involved. Our results showed that quercetin decreased ROS generation induced by Cr(VI) exposure in BEAS-2B cells. Chronic Cr(VI) exposure induced malignant cell transformation, increased miR-21 expression and caused inhibition of PDCD4, which were significantly inhibited by the treatment of quercetin in a dose dependent manner. Nude mice injected with BEAS-2B cells chronically exposed to Cr(VI) in the presence of quercetin showed reduced tumor incidence compared to Cr(VI) alone treated group. Stable knockdown of miR-21 and overexpression of PDCD4 or catalase in BEAS-2B cells suppressed Cr(VI)-induced malignant transformation and tumorigenesis. Taken together, these results demonstrate that quercetin is able to protect BEAS-2B cells from Cr(VI)-induced carcinogenesis by targeting miR-21-PDCD4 signaling.

Hexavalent chromium [Cr(VI)] is a well-known human carcinogen associated with an increased risk of lung cancer. However, the mechanisms underlying Cr(VI)-induced carcinogenesis remain unclear. MicroRNA-21 (miR-21) is a key regulator of oncogenic processes. Studies have shown that miR-21 exerts its oncogenic activity by targeting the tumor suppressor gene programmed cell death 4 (PDCD4). The present study examined the role of miR-21-PDCD4 signaling in Cr(VI)-induced cell transformation and tumorigenesis. Results showed that Cr(VI) induces ROS generation in human bronchial epithelial (BEAS-2B) cells. Chronic exposure to Cr(VI) is able to cause malignant transformation in BEAS-2B cells. Cr(VI) caused a significant increase of miR-21 expression associated with an inhibition of PDCD4 expression. Notably, STAT3 transcriptional activation by IL-6 is crucial for the Cr(VI)-induced miR-21 elevation. Stable knockdown of miR-21 or overexpression of PDCD4 in BEAS-2B cells significantly reduced the Cr(VI)-induced cell transformation. Furthermore, the Cr(VI) induced inhibition of PDCD4 suppressed downstream E-cadherin protein expression, but promoted β-catenin/TCF-dependent transcription of uPAR and c-Myc. We also found an increased miR-21 level and decreased PDCD4 expression in xenograft tumors generated with chronic Cr(VI)-exposed BEAS-2B cells. In addition, stable knockdown of miR-21 and overexpression of PDCD4 reduced the tumorogenicity of chronic Cr(VI)-exposed BEAS-2B cells in nude mice. Taken together, these results demonstrate that the miR-21-PDCD4 signaling axis plays an important role in Cr(VI)-induced carcinogenesis.

Chronic exposure to arsenic via drinking water is associated with an increased risk for development of type 2 diabetes mellitus (T2DM). This study investigates the role of mitochondrial oxidative stress protein Sirtuin 3 (Sirt3) and its targeting proteins in chronic arsenic-induced T2DM in mouse adipocytes and myotubes. The results show that chronic arsenic exposure significantly decreased insulin-stimulated glucose uptake (ISGU) in correlation with reduced expression of insulin-regulated glucose transporter type 4 (Glut4). Expression of Sirt3, a mitochondrial deacetylase, was dramatically decreased along with its associated transcription factor, forkhead box O3 (FOXO3a) upon arsenic exposure. A decrease in mitochondrial membrane potential (Δψm) was observed in both 3T3L1 adipocytes and C2C12 myotubes treated by arsenic. Reduced FOXO3a activity by arsenic exhibited a decreased binding affinity to the promoters of both manganese superoxide dismutase (MnSOD) and peroxisome proliferator-activated receptor-gamma coactivator (PGC)-1α, a broad and powerful regulator of reactive oxygen species (ROS) metabolism. Forced expression of Sirt3 or MnSOD in mouse myotubes elevated Δψm and restored ISGU inhibited by arsenic exposure. Our results suggest that Sirt3/FOXO3a/MnSOD signaling plays a significant role in the inhibition of ISGU induced by chronic arsenic exposure.

Extensive exposure of solar ultraviolet-B (UVB) radiation to skin induces oxidative stress and inflammation that play a crucial role in the induction of skin cancer. Photochemoprevention with natural products represents a simple but very effective strategy for the management of cutaneous neoplasia. In this study, we investigated whether blackberry extract (BBE) reduces chronic inflammatory responses induced by UVB irradiation in SKH-1 hairless mice skin. Mice were exposed to UVB radiation (100mJ/cm2) on alternate days for 10weeks, and BBE (10% and 20%) was applied topically a day before UVB exposure. Our results show that BBE suppressed UVB-induced hyperplasia and reduced infiltration of inflammatory cells in the SKH-1 hairless mice skin. BBE treatment reduced glutathione (GSH) depletion, lipid peroxidation (LPO), and myeloperoxidase (MPO) in mouse skin by chronic UVB exposure. BBE significantly decreased the level of pro-inflammatory cytokines IL-6 and TNF-α in UVB-exposed skin. Likewise, UVB-induced inflammatory responses were diminished by BBE as observed by a remarkable reduction in the levels of phosphorylated MAP Kinases, Erk1/2, p38, JNK1/2 and MKK4. Furthermore, BBE also reduced inflammatory mediators such as cyclooxygenase-2 (COX-2), prostaglandin E2 (PGE2), and inducible nitric oxide synthase (iNOS) levels in UVB-exposed skin. Treatment with BBE inhibited UVB-induced nuclear translocation of NF-κB and degradation of IκBα in mouse skin. Immunohistochemistry analysis revealed that topical application of BBE inhibited the expression of 8-oxo-7, 8-dihydro-2′-deoxyguanosine (8-oxodG), cyclobutane pyrimidine dimers (CPD), proliferating cell nuclear antigen (PCNA), and cyclin D1 in UVB-exposed skin. Collectively, these data indicate that BBE protects from UVB-induced oxidative damage and inflammation by modulating MAP kinase and NF-κB signaling pathways.

Arsenic is a known carcinogen to humans, and chronic exposure to environmental arsenic is a worldwide health concern. As a dietary factor, ethanol carries a well-established risk for malignancies, but the effects of co-exposure to arsenic and ethanol on tumor development are not well understood. In the present study, we hypothesized that ethanol would enhance the function of an environmental carcinogen such as arsenic through increase in COX-2 expression. Our in vitro results show that ethanol enhanced arsenic-induced COX-2 expression. We also show that the increased COX-2 expression associates with intracellular ROS generation, up-regulated AKT signaling, with activation of both NFAT and NF-κB pathways. We demonstrate that antioxidant enzymes have an inhibitory effect on arsenic/ethanol-induced COX-2 expression, indicating that the responsive signaling pathways from co-exposure to arsenic and ethanol relate to ROS generation. In vivo results also show that co-exposure to arsenic and ethanol increased COX-2 expression in mice. We conclude that ethanol enhances arsenic-induced COX-2 expression in colorectal cancer cells via both the NFAT and NF-κB pathways. These results imply that, as a common dietary factor, ethanol ingestion may be a compounding risk factor for arsenic-induced carcinogenesis/cancer development.

Occupational and environmental exposure to arsenic (III) and chromium VI (Cr(VI)) have been confirmed to cause lung cancer. Mechanisms of these metals carcinogenesis are still under investigation. Selection of cell lines to be used is essential for the studies. Human bronchial epithelial BEAS-2B cells are the cells to be utilized by most of scientists. However, due to p53 missense mutation (CCG→TCG) at codon 47 and the codon 72 polymorphism (CGC→CCC) in BEAS-2B cells, its usage has frequently been questioned. The present study has examined activity and expression of 53 and its downstream target protein p21 upon acute or chronic exposure of BEAS-2B cells to arsenic and Cr(VI). The results show that short-term exposure of BEAS-2B cells to arsenic or Cr(VI) was able to activate both p53 and p21. Chronic exposure of BEAS-2B cells to these two metals caused malignant cell transformation and tumorigenesis. In arsenic-transformed BEAS-2B cells reductions in p53 promoter activity, mRNA expression, and phosphorylation of p53 at Ser392 were observed, while the total p53 protein level remained the same compared to those in passage-matched parent ones. p21 promoter activity and expression were decreased in arsenic-transformed cells. Cr(VI)-transformed cells exhibit elevated p53 promoter activity, mRNA expression, and phosphorylation at Ser15, but reduced phosphorylation at Ser392 and total p53 protein level compared to passage-matched parent ones. p21 promoter activity and expression were elevated in Cr(VI)-transformed cells. These results demonstrate that p53 is able to respond to exposure of arsenic or Cr(VI), suggesting that BEAS-2B cells are an appropriate in vitro model to investigate arsenic or Cr(VI) induced lung cancer.

DNA methylation and histone modification promote opening and closure of chromatin structure, which affects gene expression without altering the DNA sequence. Epigenetic markers regulate the dynamic nature of chromatin structure at different levels: DNA, histone, noncoding RNAs, as well as the higher-order chromatin structure. Accumulating evidence strongly suggests that arsenic-induced carcinogenesis involves frequent changes in the epigenetic marker. However, progress in identifying arsenic-induced epigenetic changes has already been made using genome-wide approaches; the biological significance of these epigenetic changes remains unknown. Moreover, arsenic-induced changes in the chromatin state alter gene expression through the epigenetic mechanism. The current review provides a summary of recent literature regarding epigenetic changes caused by arsenic in carcinogenesis. We highlight the transgenerational studies needed to explicate the biological significance and toxicity of arsenic over a broad spectrum.

Hexavalent chromium (Cr(VI)) compounds are well-established lung carcinogens. Epidermal growth factor receptor (EGFR) is a tyrosine kinase transmembrane receptor that regulates cell survival, tumor invasion, and angiogenesis. Our results show that chronic exposure of human bronchial epithelial (BEAS-2B) cells to Cr(VI) is able to cause malignant cell transformation. These transformed cells exhibit apoptosis resistance with reduced poly ADP-ribose polymerase cleavage (C-PARP) and Bax expression and enhanced expressions of Bcl-2 and Bcl-xL. These transformed cells also exhibit reduced capacity of reactive oxygen species (ROS) generation along with elevated expression of antioxidant manganese superoxide dismutase 2 (SOD2). The expression of this antioxidant was also elevated in lung tumor tissue from a worker exposed to Cr(VI) for 19 years. EGFR was activated in Cr(VI)-transformed BEAS-2B cells, lung tissue from animals exposed to Cr(VI) particles, and human lung tumor tissue. Further study indicates that constitutive activation of EGFR in Cr(VI)-transformed cells was due to increased binding to its ligand amphiregulin (AREG). Inhibition of EGFR or AREG increased Bax expression and reduced Bcl-2 expression, resulting in reduced apoptosis resistance. Furthermore, inhibition of AREG or EGFR restored capacity of ROS generation and decreased SOD2 expression. PI3K/AKT was activated, which depended on EGFR in Cr(VI)-transformed BEAS-2B cells. Inhibition of PI3K/AKT increased ROS generation and reduced SOD2 expression, resulting in reduced apoptosis resistance with commitment increase in Bax expression and reduction of Bcl-2 expression. Xenograft mouse tumor study further demonstrates the essential role of EGFR in tumorigenesis of Cr(VI)-transformed cells. In summary, the present study suggests that ligand-dependent constitutive activation of EGFR causes reduced ROS generation and increased antioxidant expression, leading to development of apoptosis resistance, contributing to Cr(VI)-induced tumorigenesis.

Arsenic is an environmental carcinogen, its mechanisms of carcinogenesis remain to be investigated. Reactive oxygen species (ROS) are considered to be important. A previous study (Carpenter et al., 2011) has measured ROS level in human lung bronchial epithelial (BEAS-2B) cells and arsenic-transformed BEAS-2B cells and found that ROS levels were higher in transformed cells than that in parent normal cells. Based on these observations, the authors concluded that cell transformation induced by arsenic is mediated by increased cellular levels of ROS. This conclusion is problematic because this study only measured the basal ROS levels in transformed and parent cells and did not investigate the role of ROS in the process of arsenic-induced cell transformation. The levels of ROS in arsenic-transformed cells represent the result and not the cause of cell transformation. Thus question concerning whether ROS are important in arsenic-induced cell transformation remains to be answered. In the present study, we used expressions of catalase (antioxidant against H2O2) and superoxide dismutase 2 (SOD2, antioxidant against O2(-)) to decrease ROS level and investigated their role in the process of arsenic-induced cell transformation. Our results show that inhibition of ROS by antioxidant enzymes decreased arsenic-induced cell transformation, demonstrating that ROS are important in this process. We have also shown that in arsenic-transformed cells, ROS generation was lower and levels of antioxidants are higher than those in parent cells, in a disagreement with the previous report. The present study has also shown that the arsenic-transformed cells acquired apoptosis resistance. The inhibition of catalase to increase ROS level restored apoptosis capability of arsenic-transformed BEAS-2B cells, further showing that ROS levels are low in these cells. The apoptosis resistance due to the low ROS levels may increase cells proliferation, providing a favorable environment for tumorigenesis of arsenic-transformed cells.

Hexavalent chromium [Cr(VI)] is a well-known human carcinogen associated with the incidence of lung cancer. Inhibition of metal induced carcinogenesis by a dietary antioxidant is a novel approach. Luteolin, a natural dietary flavonoid found in fruits and vegetables, possesses potent antioxidant and anti-inflammatory activity. We found that short term exposure of human bronchial epithelial cells (BEAS-2B) to Cr(VI) (5μM) showed a drastic increase in ROS generation, NADPH oxidase (NOX) activation, lipid peroxidation, and glutathione depletion, which were significantly inhibited by the treatment with luteolin in a dose dependent manner. Treatment with luteolin decreased AP-1, HIF-1α, COX-2, and iNOS promoter activity induced by Cr(VI) in BEAS-2B cells. In addition, luteolin protected BEAS-2B cells from malignant transformation induced by chronic Cr(VI) exposure. Moreover, luteolin also inhibited the production of pro-inflammatory cytokines (IL-1β, IL-6, IL-8, TNF-α) and VEGF in chronic Cr(VI) exposed BEAS-2B cells. Western blot analysis showed that luteolin inhibited multiple gene products linked to survival (Akt, Fak, Bcl-2, Bcl-xL), inflammation (MAPK, NF-κB, COX-2, STAT-3, iNOS, TNF-α) and angiogenesis (HIF-1α, VEGF, MMP-9) in chronic Cr(VI) exposed BEAS-2B cells. Nude mice injected with BEAS-2B cells chronically exposed to Cr(VI) in the presence of luteolin showed reduced tumor incidence compared to Cr(VI) alone treated group. Overexpression of catalase (CAT) or SOD2, eliminated Cr(VI)-induced malignant transformation. Overall, our results indicate that luteolin protects BEAS-2B cells from Cr(VI)-induced carcinogenesis by scavenging ROS and modulating multiple cell signaling mechanisms that are linked to ROS. Luteolin, therefore, serves as a potential chemopreventive agent against Cr(VI)-induced carcinogenesis.

Skin cancer is one of the most commonly diagnosed cancers in the United States. Exposure to ultraviolet-B (UVB) radiation induces inflammation and photocarcinogenesis in mammalian skin. Cyanidin-3-glucoside (C3G), a member of the anthocyanin family, is present in various vegetables and fruits especially in edible berries, and displays potent antioxidant and anticarcinogenic properties. In this study, we have assessed the in vivo effects of C3G on UVB irradiation induced chronic inflammatory responses in SKH-1 hairless mice, a well-established model for UVB-induced skin carcinogenesis. Here, we show that C3G inhibited UVB-induced skin damage and inflammation in SKH-1 hairless mice. Our results indicate that C3G inhibited glutathione depletion, lipid peroxidation and myeloperoxidation in mouse skin by chronic UVB exposure. C3G significantly decreased the production of UVB-induced pro-inflammatory cytokines, such as IL-6 and TNF-α, associated with cutaneous inflammation. Likewise, UVB-induced inflammatory responses were diminished by C3G as observed by a remarkable reduction in the levels of phosphorylated MAP kinases, Erk1/2, p38, JNK1/2 and MKK4. Furthermore, C3G also decreased UVB-induced cyclooxygenase-2 (COX-2), PGE2 and iNOS levels, which are well-known key mediators of inflammation and cancer. Treatment with C3G inhibited UVB-induced nuclear translocation of NF-κB and degradation of IκBα in mice skin. Immunofluorescence assay revealed that topical application of C3G inhibited the expression of 8-hydroxy-2′-deoxyguanosine, proliferating cell nuclear antigen, and cyclin D1 in chronic UVB exposed mouse skin. Collectively, these data indicates that C3G can provide substantial protection against the adverse effects of UVB radiation by modulating UVB-induced MAP kinase and NF-κB signaling pathways.