Anti-Inflammatory Effect of Korean Soybean Sauce (Ganjang) on Mice with Induced Colitis.

Inflammatory bowel disease (IBD), characterized by chronic inflammation of the gut, is caused by several factors. Among these factors, microbial factors are correlated with the gut microbiota, which produces short-chain fatty acids (SCFAs) via anaerobic fermentation. Fermented foods are known to regulate the gut microbiota composition. Ganjang (GJ), a traditional fermented Korean soy sauce consumed worldwide, has been shown to exhibit antioxidant, anticancer, anti-colitis, and antihypertensive activities. However, its effects on the gut microbiota remain unknown. In the present study, we aimed to compare the anti-inflammatory effects of GJ manufactured using different methods and investigate its effect on SCFA production in the gut. To evaluate the anti-inflammatory effects of GJ in the gut, we performed animal experiments using a mouse model of dextran sulfate sodium (DSS)-induced colitis. All GJ samples attenuated DSS-induced colitis symptoms, including reduced colonic length, by suppressing the expression of inflammatory cytokines. In addition, GJ administration modulated SCFA production in the DSS-induced colitis model. Overall, GJ exerted anti-inflammatory effects by reducing DSS-induced symptoms via regulation of inflammation and modulation of SCFA levels in a DSS-induced colitis model. Thus, GJ is a promising fermented food with the potential to prevent IBD.

Angular dihydropyranocoumarins from the flowers of Peucedanum japonicum and their aldo-keto reductase inhibitory activities.

Twenty-one angular dihydropyranocoumarins and a linear furanocoumarin, including four previously undescribed compounds (1-4), were isolated from the flowers of Peucedanum japonicum (Umbelliferae). The structures of 1-4, along with their absolute stereochemistry, were determined to be (3'S,4'S)-3'-O-propanoyl-4'-O-(3‴-methyl-2‴-butenoyl)khellactone (1), (3'S,4'S)-3'-O-propanoyl-4'-O-(2‴-methyl-2‴Z-butenoyl)khellactone (2), (3'S,4'S)-3'-O-propanoyl-4'-O-(2‴-methylbutanoyl)khellactone (3), and (3'S,4'S)-3'-O-(2″-methylpropanoyl)-4'-O-(3‴-methyl-2‴-butenoyl)khellactone (4) using one- and two-dimensional nuclear magnetic resonance, high-resolution electrospray ionization mass spectroscopy, and electronic circular dichroism spectroscopy. In addition, the absolute configuration of the three angular dihydropyranocoumarins (5-7) was determined for the first time in this study. Among the previously reported compounds isolated in this study, 8 and 9 were isolated for the first time from the genus Peucedanum, whereas 10 and 11 were previously unreported and had not been isolated from P. japonicum to date. Furthermore, all isolated compounds were evaluated for their aldo-keto reductase 1C1 inhibitory activities on A549 human non-small-cell lung cancer cells. Compounds 10 and 12 exhibited substantial AKR1C1 inhibitory activities with IC50 values of 35.8 ± 0.9 and 44.2 ± 1.5 µM, respectively.

Antimetastatic activity of seongsanamide B in γ-irradiated human lung cancer.

Lung cancer, which has a high incidence and mortality rates, often metastasizes and exhibits resistance to radiation therapy. Seongsanamide B has conformational features that suggest it has therapeutic potential; however, its antitumor activity has not yet been reported. We evaluated the possibility of seongsanamide B as a radiation therapy efficiency enhancer to suppress γ-irradiation-induced metastasis in non-small cell lung cancer. Seongsanamide B suppressed non-small cell lung cancer cell migration and invasion caused by γ-irradiation. Furthermore, it suppressed γ-irradiation-induced upregulation of Bcl-XL and its downstream signaling molecules, such as superoxide dismutase 2 (SOD2) and phosphorylated Src, by blocking the nuclear translocation of phosphorylated STAT3. Additionally, seongsanamide B markedly modulated the γ-irradiation-induced upregulation of E-cadherin and vimentin. Consistent with the results obtained in vitro, while seongsanamide B did not affect xenograft tumor growth, it significantly suppressed γ-irradiation-induced metastasis by inhibiting Bcl-XL/SOD2/phosphorylated-Src expression and modulating E-cadherin and vimentin expression in a mouse model. Thus, seongsanamide B may demonstrate potential applicability as a radiation therapy efficiency enhancer for lung cancer treatment.

Effects of Cheonggukjang (Fermented Soybean) on the Development of Colitis-Associated Colorectal Cancer in Mice.

Colorectal cancer (CRC) is the third most common type of cancer and is caused by multiple factors. Chronic inflammation, known to cause inflammatory bowel disease (IBD), is closely associated with CRC. Cheonggukjang (CJ), a traditional Korean fermented soybean, is a functional food with anti-inflammatory effects in the intestines, but its anti-cancer effects have not yet been explored. In this study, we investigated the cancer-protective effects of cheonggukjang in an azoxymethane/DSS (AOM/DSS)-induced colitis-associated colorectal cancer (CAC) mouse model. The CJ alleviated AOM/DSS-induced pathological symptoms such as colonic shortening, increased spleen weight, tumor formation, and histological changes. It also modulated pro-inflammatory and anti-inflammatory cytokine levels via the suppression of NF-κB and inflammatory mediator signaling pathways. Furthermore, the CJ improved intestinal integrity by regulating mucin-associated and tight junction proteins. In addition, it suppressed tumor growth by regulating apoptosis and proliferation. These results highlight the anti-tumor effects of CJ in an AOM/DSS-induced CAC mouse model.

Phytochemical profile and anti-inflammatory activity of the hull of γ-irradiated wheat mutant lines (Triticum aestivum L.).

Wheat (Triticum aestivum Linn.; Poaceae) is the second most cultivated food crop among all global cereal crop production. The high carbohydrate content of its grains provides energy, multiple nutrients, and dietary fiber. After threshing, a substantial amount of wheat hull is produced, which serves as the non-food component of wheat. For the valorization of these by-products as a new resource from which functional components can be extracted, the hull from the seeds of cultivated wheat mutant lines bred after γ-irradiation were collected. Untargeted metabolite analysis of the hull of the original cultivar (a crossbreeding cultivar., Woori-mil × D-7) and its 983 mutant lines were conducted using ultra-performance liquid chromatography-electrospray ionization quadrupole time-of-flight mass spectrometry technique. A total of 55 molecules were tentatively identified, including 21 compounds found in the Triticum species for the first time and 13 compounds not previously described. Among them, seven flavonolignans with a diastereomeric structure, isolated as a single compound from the hull of T. aestivum in our previous study, were used as the standards in the metabolite analysis. The differences in their collision cross-section values were shown to contribute to the clear distinction between tricine-lignan stereoisomers. To select functionally active agents with anti-inflammatory activity among the identified compounds, the wheat hull samples were evaluated for their inhibitory effect on nitric oxide production in lipopolysaccharide-stimulated RAW 264.7 cells. As a result of multivariate analysis based on the results of chemical and biological profiles of the wheat hull samples, 10 metabolites were identified as key markers, contributing to the distinction between active and inactive mutant lines. Considering that one of the four key markers attributed to anti-inflammatory activity has been identified to be a flavonolignan, the wheat hull could be a valuable source of diverse tricin-lignan type compounds and used as a natural health-promoting product in food supplements.

Comparative Analysis of Coumarin Profiles in Different Parts of Peucedanum japonicum and Their Aldo-Keto Reductase Inhibitory Activities.

Peucedanum japonicum (Umbelliferae) is widely distributed throughout Southeast Asian countries. The root of this plant is used in traditional medicine to treat colds and pain, whereas the young leaves are considered an edible vegetable. In this study, the differences in coumarin profiles for different parts of P. japonicum including the flowers, roots, leaves, and stems were compared using ultra-performance liquid chromatography time-of-flight mass spectrometry. Twenty-eight compounds were tentatively identified, including three compounds found in the genus Peucedanum for the first time. Principal component analysis using the data set of the measured mass values and intensities of the compounds exhibited distinct clustering of the flower, leaf, stem, and root samples. In addition, their anticancer activities were screened using an Aldo-keto reductase (AKR)1C1 assay on A549 human non-small-cell lung cancer cells and the flower extract inhibited AKR1C1 activity. Based on these results, seven compounds were selected as potential markers to distinguish between the flower part versus the root, stem, and leaf parts using an orthogonal partial least-squares discriminant analysis. This study is the first to provide information on the comparison of coumarin profiles from different parts of P. japonicum as well as their AKR1C1 inhibitory activities. Taken together, the flowers of P. japonicum offer a new use related to the efficacy of overcoming anticancer drug resistance, and may be a promising source for the isolation of active lead compounds.

Content of Two Major Steroidal Glycoalkaloids in Tomato (Solanum lycopersicum cv. Micro-Tom) Mutant Lines at Different Ripening Stages.

Esculeoside A and tomatine are two major steroidal alkaloids in tomato fruit (Solanum lycopersicum) that exhibit anti-inflammatory, anticancer, and anti-hyperlipidemia activities. Tomatine contained in immature tomato fruit is converted to esculeoside A as the fruit matures. To develop new tomato varieties based on the content analysis of functional secondary metabolites, 184 mutant lines were generated from the original cultivar (S. lycopersicum cv. Micro-Tom) by radiation breeding. Ultra-performance liquid chromatography coupled with evaporative light scattering detector was used to identify the mutant lines with good traits by analyzing tomatine and esculeoside A content. Compared with the original cultivar, candidates for highly functional cultivars with high esculeoside A content were identified in the mature fruit of the mutant lines. The mutant lines with low and high tomatine content at an immature stage were selected as edible cultivars due to toxicity reduction and as a source of tomatine with various pharmacological activities, respectively. During the process of ripening from green to red tomatoes, the rate of conversion of tomatine to esculeoside A was high in the green tomatoes with a low tomatine content, whereas green tomatoes with a high tomatine content exhibited a low conversion rate. Using methanol extracts prepared from unripe and ripe fruits of the original cultivar and its mutant lines and two major compounds, we examined their cytotoxicity against FaDu human hypopharynx squamous carcinoma cells. Only tomatine exhibited cytotoxicity with an IC50 value of 5.589 µM, whereas the other samples did not exhibit cytotoxicity. Therefore, radiation breeding represents a useful tool for developing new cultivars with high quality, and metabolite analysis is applicable for the rapid and objective selection of potential mutant lines.

Protective Effects of Fermented Soybeans (Cheonggukjang) on Dextran Sodium Sulfate (DSS)-Induced Colitis in a Mouse Model.

Inflammatory bowel disease (IBD) is a chronic inflammatory disease, and the incidence of IBD is increasing every year owing to changes in dietary structure. Although the exact pathogenesis of IBD is still unclear, recent evidence suggests that gut dysbiosis is closely associated with IBD pathogenesis. Cheonggukjang is a traditional Korean fermented soybean paste produced using traditional and industrial methods, and contains probiotics, which affect the gut microbiota composition. However, the protective effect of Cheonggukjang against IBD is unknown. In this study, we investigated the bacterial community structure of traditional and commercial Cheonggukjang samples, as well as the protective effect of Cheonggukjang on a dextran sulfate sodium (DSS)-induced colitis mouse model. Traditional and commercial Cheonggukjang were found to contain various type of useful probiotics in their bacterial community structure. Cheonggukjang reduced the progression of DSS-induced symptoms, such as body weight loss, colonic shortening, disease activity index, and histological changes. Further, Cheonggukjang improved the intestinal epithelial barrier integrity on DSS-induced colitis mice. In addition, Cheonggukjang suppressed the expression of proinflammatory cytokines and inflammatory mediators through the inactivation of NF-κB and MAPK signaling pathways. These results indicate that Cheonggukjang exerts protective effects against DSS-induced colitis, suggesting its possible application as a functional food for improving inflammatory diseases.

Anti-Inflammatory Effects of Weigela subsessilis Callus Extract via Suppression of MAPK and NF-κB Signaling.

Weigela subsessilis is used in folk medicine to treat pain and allergic syndromes in Korea. However, the antibacterial and anti-inflammatory activities of W. subsessilis callus extract remain unexplored. In this study, we aimed to evaluate the W. subsessilis callus of pharmacological activity. Therefore, we first established in vitro calluses of W.subsessilis via plant tissue culture methods. We then evaluated the antioxidant and anti-inflammatory effects of W. subsessilis callus extract in lipopolysaccharide (LPS)-treated RAW264.7 macrophage cells. The W. subsessilis callus extract showed antioxidant and anti-inflammatory effects. These effects were regulated via suppression of mitogen-activated protein kinase signaling through LPS-induced translocation of nuclear factor kappa B (NF-κB) p65 from the cytoplasm to the nucleus. W. subsessilis callus extract also showed antibacterial and anti-inflammatory activities in Propionibacterium acnes-treated HaCaT keratinocyte cells. These results indicate that W. subsessilis callus extract has antioxidant, antibacterial and anti-inflammatory activities, suggesting its possible application in the treatment of inflammatory disorders.

Inhibition of A549 Lung Cancer Cell Migration and Invasion by Ent-Caprolactin C via the Suppression of Transforming Growth Factor-ß-Induced Epithelial-Mesenchymal Transition.

The epithelial-mesenchymal transition (EMT) of cancer cells is a crucial process in cancer cell metastasis. An Aquimarina sp. MC085 extract was found to inhibit A549 human lung cancer cell invasion, and caprolactin C (1), a new natural product, α-amino-ε-caprolactam linked to 3-methyl butanoic acid, was purified through bioactivity-guided isolation of the extract. Furthermore, its enantiomeric compound, ent-caprolactin C (2), was synthesized. Both 1 and 2 inhibited the invasion and γ-irradiation-induced migration of A549 cells. In transforming growth factor-ß (TGF-ß)-treated A549 cells, 2 inhibited the phosphorylation of Smad2/3 and suppressed the EMT cell marker proteins (N-cadherin, ß-catenin, and vimentin), as well as the related messenger ribonucleic acid expression (N-cadherin, matrix metalloproteinase-9, Snail, and vimentin), while compound 1 did not suppress Smad2/3 phosphorylation and the expression of EMT cell markers. Therefore, compound 2 could be a potential candidate for antimetastatic agent development, because it suppresses TGF-ß-induced EMT.

Dendrobine Inhibits γ-Irradiation-Induced Cancer Cell Migration, Invasion and Metastasis in Non-Small Cell Lung Cancer Cells.

The use of ionizing radiation (IR) during radiotherapy can induce malignant effects, such as metastasis, which contribute to poor prognoses in lung cancer patients. Here, we explored the ability of dendrobine, a plant-derived alkaloid from Dendrobium nobile, to improve the efficacy of radiotherapy in non-small cell lung cancer (NSCLC). We employed Western blotting, quantitative real-time (qRT)-PCR, transwell migration assays, and wound-healing assays to determine the effects of dendrobine on the migration and invasion of A549 lung cancer cells in vitro. Dendrobine (5 mm) inhibited γ-irradiation-induced migration and invasion of A549 cells by suppressing sulfatase2 (SULF2) expression, thus inhibiting IR-induced signaling. To investigate the inhibitory effects of dendrobine in vivo, we established a mouse model of IR-induced metastasis by injecting BALB/c nude mice with γ-irradiated A549 cells via the tail vein. As expected, injection with γ-irradiated cells increased the number of pulmonary metastatic nodules in mice (0 Gy/DPBS, 9.8 ± 1.77; 2 Gy/DPBS, 20.87 ± 1.42), which was significantly reduced with dendrobine treatment (2 Gy/Dendrobine, 10.87 ± 0.71), by prevention of IR-induced signaling. Together, these findings demonstrate that dendrobine exerts inhibitory effects against γ-irradiation-induced invasion and metastasis in NSCLC cells in vitro and in vivo at non cytotoxic concentrations. Thus, dendrobine could serve as a therapeutic enhancer to overcome the malignant effects of radiation therapy in patients with NSCLC.

Chemical and Biological Profiles of Dendrobium in Two Different Species, Their Hybrid, and Gamma-Irradiated Mutant Lines of the Hybrid Based on LC-QToF MS and Cytotoxicity Analysis.

The Dendrobium species (Orchidaceae) has been cultivated as an ornamental plant as well as used in traditional medicines. In this study, the chemical profiles of Dendrobii Herba, used as herbal medicine, Dendrobium in two different species, their hybrid, and the gamma-irradiated mutant lines of the hybrid, were systematically investigated via ultra-performance liquid chromatography coupled with quadrupole time-of-flight mass spectrometry (UPLC-QToF MS). Among the numerous peaks detected, 17 peaks were unambiguously identified. Gigantol (1), (1R,2R)-1,7-hydroxy-2,8-methoxy-2,3-dihydrophenanthrene-4(1H)-one (2), tristin (3), (-)-syringaresinol (4), lusianthridin (5), 2,7-dihydroxy-phenanthrene-1,4-dione (6), densiflorol B (7), denthyrsinin (8), moscatilin (9), lusianthridin dimer (10), batatasin III (11), ephemeranthol A (12), thunalbene (13), dehydroorchinol (14), dendrobine (15), shihunine (16), and 1,5,7-trimethoxy-2-phenanthrenol (17), were detected in Dendrobii Herba, while 1, 2, and 16 were detected in D. candidum, 1, 11, and 16 in D. nobile, and 1, 2, and 16 in the hybrid, D. nobile × candidum. The methanol extract taken of them was also examined for cytotoxicity against FaDu human hypopharynx squamous carcinoma cells, where Dendrobii Herba showed the greatest cytotoxicity. In the untargeted metabolite analysis of 436 mutant lines of the hybrid, using UPLC-QToF MS and cytotoxicity measurements combined with multivariate analysis, two tentative flavonoids (M1 and M2) were evaluated as key markers among the analyzed metabolites, contributing to the distinction between active and inactive mutant lines.

Phytochemicals and Bioactivities of Zingiber cassumunar Roxb.

Zingiber cassumunar Roxb. (Zingiberaceae), is an important medicinal plant known as "Plai (Phlai)" in Thailand, "Bangle" in Indonesia, and "Bulei" in China. Traditionally, this plant has been used to treat inflammation, pain, and respiratory problems. The rhizomes are the primary part of the plant that has been used for medicinal purposes due to their constituents with therapeutic properties, including phenylbutenoids, curcuminoids, and essential oils. Since the 1970s, many studies have been conducted on the phytochemicals and bioactivities of Z. cassumunar to establish fundamental scientific evidence that supports its use in traditional medicine. The accumulated biological studies on the extracts, solvent fractions, and constituents of Z. cassumunar have described their diverse medicinal properties, including antioxidant, anti-inflammatory, anticancer, neuroprotective/neurotrophic, cosmeceutical, and antifungal/antimicrobial bioactivities. In this review, we summarize information on the phytochemicals of Z. cassumunar and the bioactivities of its extracts and constituents.

Anti-Inflammatory Activities of an Extract of In Vitro Grown Adventitious Shoots of Toona sinensis in LPS-Treated RAW264.7 and Propionibacterium acnes-Treated HaCaT Cells.

Toona sinensis has been traditionally used to treat dysentery, enteritis, flatulence, and itchiness. However, the existence of anti-inflammatory effects of T. sinensis on Propionibacterium acnes-induced skin disease is unknown. In vitro cultures of plant cells and tissues produced under controlled conditions offer a continuous production platform for plant natural products including pigments and anti-inflammatory agents. In this study, we determine the anti-inflammatory activities of an extract of in vitro grown adventitious shoots of T. sinensis on P. acnes, the etiologic agent of skin inflammation. The extract of T. sinensis showed antioxidant and anti-inflammatory activity in LPS-treated RAW264.7 cells. It also had antibacterial activity and anti-inflammatory effects on P. acnes-treated HaCaT cells. In addition, these effects were regulated by suppression of the mitogen-activated protein kinase (MAPK) pathways. These results suggesting the potential application of adventitious shoots of T. sinensis grown with an in vitro proliferation system as a medicine for treating P. acnes-induced inflammatory skin disease.

Triterpenoids from the Leaves of Centella asiatica Inhibit Ionizing Radiation-Induced Migration and Invasion of Human Lung Cancer Cells.

Radiotherapy using ionizing radiation is a major therapeutic modality for advanced human lung cancers. However, ionizing radiation itself can induce malignant behaviors such as cancer cell migration and invasion, leading to local recurrence or distal metastasis. Therefore, safer and more effective agents that inhibit the metastatic behaviors of cancer cells in radiotherapy are needed. As a part of our ongoing search for new radiotherapy enhancers from medicinal herbs, we isolated the following triterpenoids from the ethanol extract of Centella asiatica: asiatic acid (1), madecassic acid (2), and asiaticoside (3). These compounds inhibited the ionizing radiation-induced migration and invasion of A549 human lung cancer cells at noncytotoxic concentrations. These results suggest that triterpenoids 1-3 isolated from C. asiatica are candidate natural compounds to enhance the effect of radiotherapy in patients with non-small-cell lung cancer.

Epithelial-mesenchymal transition gene signature is associated with prognosis and tumor microenvironment in head and neck squamous cell carcinoma.

In this study we assessed the clinical significance of an epithelial-mesenchymal transition (EMT) gene signature and explored its association with the tumor microenvironment related to immunotherapy in patients with head and neck squamous cell carcinoma (HNSCC). Genes were selected when mRNA levels were positively or negatively correlated with at least one well-known EMT marker. We developed an EMT gene signature consisting of 82 genes. The patients were classified into epithelial or mesenchymal subgroups according to EMT signature. The clinical significance of the EMT signature was validated in three independent cohorts and its association with several immunotherapy-related signatures was investigated. The mesenchymal subgroup showed worse prognosis than the epithelial subgroup, and significantly elevated PD-1, PD-L1, and CTLA-4 levels, and increased interferon-gamma, cytolytic, T cell infiltration, overall immune infiltration, and immune signature scores. The relationship between PD-L1 expression and EMT status in HNSCC after treatment with TGF-ß was validated in vitro. In conclusion, the EMT gene signature was associated with prognosis in HNSCC. Additionally, our results suggest that EMT is related to immune activity of the tumor microenvironment with elevated immune checkpoint molecules.

Moscatilin Induces Apoptosis in Human Head and Neck Squamous Cell Carcinoma Cells via JNK Signaling Pathway.

Dendrobii Herba is an herbal medicine that uses the stems of Dendrobium species (Orchidacea). It has been traditionally used to treat fever, hydrodipsomania, stomach disorders, and amyotrophia. In our previous study, a bibenzyl compound, moscatilin, which is isolated from Dendrobii Herba, showed potent cytotoxicity against a FaDu human pharyngeal squamous carcinoma cell line. Prompted by this finding, we performed additional studies in FaDu cells to investigate the mechanism of action. Moscatilin induced FaDu cell death by using 5 µM of concentration and by mediating apoptosis, whereas cell proliferation following treatment with 1 µM of moscatilin was not suppressed to the same levels as by the anti-cancer agent, cisplatin. Apoptosis-related protein expression (cleaved caspase-8, cleaved caspase-7, cytochrome c, cleaved caspase-9, cleaved caspase-3, and poly (ADP-ribose) polymerase (PARP) was increased by treating with 5 µM of moscatilin. This suggests that moscatilin-mediated apoptosis is associated with the extrinsic and intrinsic apoptotic signaling pathways. In addition, moscatilin-induced apoptosis was mediated by the c-Jun N-terminal kinase (JNK) signaling pathway. Overall, this study identified additional biological activity of moscatilin derived from natural products and suggested its potential application as a chemotherapeutic agent for the management of head and neck squamous cell carcinoma.

Identification of Two New Phenanthrenes from Dendrobii Herba and Their Cytotoxicity towards Human Hypopharynx Squamous Carcinoma Cell (FaDu).

Two new phenanthrenes, (1R,2R)-1,7-hydroxy-2,8-methoxy-2,3-dihydrophenanthrene-4(1H)-one (1) and 2,7-dihydroxy-phenanthrene-1,4-dione (2), were isolated from the ethyl acetate-soluble fraction of Dendrobii Herba, together with seven known phenanthrenes (3-9), two bibenzyls (10-12), and a lignan (13). Structures of 1 and 2 were elucidated by analyzing one-dimensional (1D) and two-dimensional (2D)-NMR and High-resolution electrospray ionization mass spectra (HR-ESI-MS) data. The absolute configuration of compound 1 was confirmed by the circular dichroism (CD) spectroscopic method. In cytotoxicity assay using FaDu human hypopharynx squamous carcinoma cell line, compounds 3-6, 8, 10, and 12 showed activities, with IC50 values that ranged from 2.55 to 17.70 µM.

Mitochondrial superoxide dismutase 2 mediates γ-irradiation-induced cancer cell invasion.

Sublethal doses of γ-rays promote cancer cell invasion by stimulating a signaling pathway that sequentially involves p53, sulfatase 2 (SULF2), ß-catenin, interleukin-6 (IL-6), signal transducer and activator of transcription 3 (STAT3), and Bcl-XL. Given that Bcl-XL can increase O2•- production by stimulating respiratory complex I, the possible role of mitochondrial reactive oxygen species (ROS) in γ-irradiation-induced cell invasion was investigated. Indeed, γ-irradiation promoted cell invasion by increasing mitochondrial ROS levels, which was prevented by metformin, an inhibitor of complex I. γ-Irradiation-stimulated STAT3 increased the expression of superoxide dismutase 2 (SOD2), a mitochondrial enzyme that catalyzes the conversion of O2•- to hydrogen peroxide (H2O2). In contrast to O2•-, H2O2 functions as a signaling molecule. γ-Irradiation consistently stimulated the Src-dependent invasion pathway in a manner dependent on both complex I and SOD2. SOD2 was also essential for the invasion of un-irradiated cancer cells induced by upregulation of Bcl-XL, an intracellular oncogene, or extracellular factors, such as SULF2 and IL-6. Overall, these data suggested that SOD2 is critical for the malignant effects of radiotherapy and tumor progression through diverse endogenous factors.

Genome-Wide Studies of Rho5-Interacting Proteins That Are Involved in Oxidant-Induced Cell Death in Budding Yeast.

Rho GTPases play critical roles in cell proliferation and cell death in many species. As in animal cells, cells of the budding yeast Saccharomyces cerevisiae undergo regulated cell death under various physiological conditions and upon exposure to external stress. The Rho5 GTPase is necessary for oxidant-induced cell death, and cells expressing a constitutively active GTP-locked Rho5 are hypersensitive to oxidants. Yet how Rho5 regulates yeast cell death has been poorly understood. To identify genes that are involved in the Rho5-mediated cell death program, we performed two complementary genome-wide screens: one screen for oxidant-resistant deletion mutants and another screen for Rho5-associated proteins. Functional enrichment and interaction network analysis revealed enrichment for genes in pathways related to metabolism, transport, and plasma membrane organization. In particular, we find that ATG21, which is known to be involved in the CVT (Cytoplasm-to-Vacuole Targeting) pathway and mitophagy, is necessary for cell death induced by oxidants. Cells lacking Atg21 exhibit little cell death upon exposure to oxidants even when the GTP-locked Rho5 is expressed. Moreover, Atg21 interacts with Rho5 preferentially in its GTP-bound state, suggesting that Atg21 is a downstream target of Rho5 in oxidant-induced cell death. Given the high degree of conservation of Rho GTPases and autophagy from yeast to human, this study may provide insight into regulated cell death in eukaryotes in general.