YD-10B Cells Research Articles

Quercetin Induces Cell Cycle Arrest and Apoptosis in YD10B and YD38 Oral Squamous Cell Carcinoma Cells.

Oral squamous cell carcinoma (OSCC) exhibits the highest lethality among head and neck cancers. Treatment for OSCC is limited due to diverse side effects. Quercetin is a natural flavonoid compound found in many kinds of plants and foods. Quercetin has been reported to be a modulator of proliferation and survival in various types of cancers due to its cytotoxic effects. We aimed to investigating chemopreventative roles of quercetin in YD10B and YD38 OSCC cells. For our study, two different types of OSCC cells were used. YD10B cells are tongue SCC cells with the p53 mutation and YD38 cells are lower gingiva SCC cells without the p53 mutation, respectively. The anticancer effects of quercetin were examined by cell viability, cell cycle, annexin-PI staining, and western blot. Our results showed that quercetin decreased cell viability and induced G1 cell cycle arrest in YD10B and YD38 OSCC cells. Moreover, quercetin remarkably decreased the expression of cell cycle upregulating proteins and increased the expression of a CDK inhibitor. Quercetin also significantly increased the number of annexin-V-positive cells in a dose-dependent manner in both types of OSCC cells. This apoptotic potential of quercetin triggered cleavage of PARP followed by activation of p38 MAPK signaling pathway. In conclusion, this study demonstrates that quercetin shows different anti-cancer responses in OSCC with and without p53 mutation, respectively. Despite different p53 status in OSCC cells, quercetin led to apoptotic signals in both cells. Quercetin repressed cell proliferation with G1 cell cycle arrest and apoptosis by activating the p38 signaling pathway in two OSCC cells with different p53 status. These findings might provide new strategy for OSCC therapy by quercetin.

Silibinin induces oral cancer cell apoptosis and reactive oxygen species generation by activating the JNK/c-Jun pathway.

Background: Oral cancer is one of the most prevalent malignant tumors worldwide. Silibinin has been reported to exert therapeutic effects in various cancer models. However, its mechanism of action in oral cancer remains unclear. We aimed to examine the molecular processes underlying the effects of silibinin in oral cancer in vitro and in vivo as well as its potential anticancer effects. Next, we investigated the molecular processes underlying both in vitro and in vivo outcomes of silibinin treatment on oral cancer. Methods: To investigate the effects of silibinin on the growth of oral cancer cells, cell proliferation and anchorage-independent colony formation tests were conducted on YD10B and Ca9-22 oral cancer cells. The effects of silibinin on the migration and invasion of oral cancer cells were evaluated using transwell assays. Flow cytometry was used to examine apoptosis, cell cycle distribution, and accumulation of reactive oxygen species (ROS). The molecular mechanism underlying the anticancer effects of silibinin was explored using immunoblotting. The in vivo effects of silibinin were evaluated using a Ca9-22 xenograft mouse model. Results: Silibinin effectively suppressed YD10B and Ca9-22 cell proliferation and colony formation in a dose-dependent manner. Moreover, it induced cell cycle arrest in the G0/G1 phase, apoptosis, and ROS generation in these cells. Furthermore, silibinin inhibited the migration and invasion abilities of YD10B and Ca9-22 cells by regulating the expression of proteins involved in the epithelial-mesenchymal transition. Western blotting revealed that silibinin downregulated SOD1 and SOD2 and triggered the JNK/c-Jun pathway in oral cancer cells. Silibinin significantly inhibited xenograft tumor growth in nude mice, with no obvious toxicity. Conclusions: Silibinin considerably reduced the development of oral cancer cells by inducing apoptosis, G0/G1 arrest, ROS generation, and activation of the JNK/c-Jun pathway. Importantly, silibinin effectively suppressed xenograft tumor growth in nude mice. Our findings indicate that silibinin may be a promising option for the prevention or treatment of oral cancer.

Effects of Sangju Honey on Oral Squamous Carcinoma Cells.

Since ancient times, honey has been used in traditional medicine owing to its pharmacological effects. It possesses anticancer properties. However, the therapeutic implications of Sangju honey in cancer remains unknown. Therefore, we aimed to demonstrate the potential anticancer effects of Sangju honey on human oral squamous cell carcinoma (OSCC), particularly focusing on epithelial-mesenchymal transition (EMT) and apoptotic and mitogen-activated protein kinase (MAPK) signaling pathways. Ca9-22 and YD-10B human OSCC cells were treated with 0.25% or 0.5% Sangju honey, and the cell viability was examined using the Cell Counting Kit-8 assay. Cell morphology studies were conducted to observe morphological changes, and the wound-healing assay was performed to evaluate the proliferation of honey-treated OSCC cells. Western blot analysis was conducted to investigate protein expression related to EMT and apoptotic and MAPK signaling pathways. Sangju honey reduced cell viability, induced morphological changes, and significantly suppressed the proliferation and migration of Ca9-22 and YD-10B cells. The expression of E-cadherin and N-cadherin was increased and decreased, respectively, in both OSCC cell lines. Moreover, Sangju honey stimulated apoptosis by increasing the expression of p21, p53, cleaved caspase 3, and caspase 9. Furthermore, it downregulated the expression of phospho (p)-extracellular signal-regulated kinases 1 and 2, p-c-Jun amino-terminal kinase, and p-p38 in Ca9-22 and YD-10B cells. Sangju honey inhibits Ca9-22 and YD-10B cell proliferation by regulating EMT, inducing apoptosis, and suppressing the MAPK signaling pathway. Thus, it is a potential anticancer agent for human OSCC.

5-Fluorouracil-Immobilized Hyaluronic Acid Hydrogel Arrays on an Electrospun Bilayer Membrane as a Drug Patch.

The hyaluronic acid (HA) hydrogel array was employed for immobilization of 5-fluorouracil (5-FU), and the electrospun bilayer (hydrophilic: polyurethane/pluronic F-127 and hydrophobic: polyurethane) membrane was used to support the HA hydrogel array as a patch. To visualize the drug propagating phenomenon into tissues, we experimentally investigated how FITC-BSA diffused into the tissue by applying hydrogel patches to porcine tissue samples. The diffusive phenomenon basically depends on the FITC-BSA diffusion coefficient in the hydrogel, and the degree of diffusion of FITC-BSA may be affected by the concentration of HA hydrogel, which demonstrates that the high density of HA hydrogel inhibits the diffusive FITC-BSA migration toward the low concentration region. YD-10B cells were employed to investigate the release of 5-FU from the HA array on the bilayer membrane. In the control group, YD-10B cell viability was over 98% after 3 days. However, in the 5-FU-immobilized HA hydrogel array, most of the YD-10B cells were not attached to the bilayer membrane used as a scaffold. These results suggest that 5-FU was locally released and initiated the death of the YD-10B cells. Our results show that 5-FU immobilized on HA arrays significantly reduces YD-10B cell adhesion and proliferation, affecting cells even early in the cell culture. Our results suggest that when 5-FU is immobilized in the HA hydrogel array on the bilayer membrane as a drug patch, it is possible to control the drug concentration, to release it continuously, and that the patch can be applied locally to the targeted tumor site and administer the drug in a time-stable manner. Therefore, the developed bilayer membrane-based HA hydrogel array patch can be considered for sustained release of the drug in biomedical applications.

Falcarindiol Stimulates Apoptotic and Autophagic Cell Death to Attenuate Cell Proliferation, Cell Division, and Metastasis through the PI3K/AKT/mTOR/p70S6K Pathway in Human Oral Squamous Cell Carcinomas.

Human oral squamous cell carcinomas (OSCCs) have high cancer mortality and a 5-year survival rate lower than that of most other carcinomas. New therapeutic strategies are required for the treatment and prevention against OSCCs. An approach to cancer therapy using plant-derived natural compounds has been actively in progress as a trend. Falcarindiol (FALC), or its isolated form Ostericum koreanum Kitagawa (O. koreanum), is present in many food and dietary plants, especially in carrots, and this compound has a variety of beneficial effects. However, biological activity of FALC has not been reported in OSCCs yet. This study aimed to demonstrate the antitumor effects of FALC against OSCCs, YD-10B cells. In this study, FALC was selected as a result of screening for compounds isolated from various natural products in YD-10B cells. FALC suppressed cell growth, and FALC-induced apoptotic cell death was mainly accompanied by the dephosphorylation of PI3K, AKT, mTOR, and p70S6K. The apoptotic cell death was also associated with autophagy as evidenced by the expression of Beclin-1, the conversion of LC3-II, and the formation of autophagosome. FALC-induced autophagy was accompanied by MAPKs including ERK1/2 and p38. Furthermore, FALC caused the antimetastatic effects by inhibiting the migration and invasion of YD-10B cells. Taken together, the findings suggest the potential value of FALC as a novel candidate for therapeutic strategy against OSCCs.

Anti-growth and pro-apoptotic effects of dasatinib on human oral cancer cells through multi-targeted mechanisms.

Dasatinib is an inhibitor of Src that has anti‐tumour effects on many haematological and solid cancers. However, the anti‐tumour effects of dasatinib on human oral cancers remain unclear. In this study, we investigated the effects of dasatinib on different types of human oral cancer cells: the non‐tumorigenic YD‐8 and YD‐38 and the tumorigenic YD‐10B and HSC‐3 cells. Strikingly, dasatinib at 10 µM strongly suppressed the growth and induced apoptosis of YD‐38 cells and inhibited the phosphorylation of Src, EGFR, STAT‐3, STAT‐5, PKB and ERK‐1/2. In contrast, knockdown of Src blocked the phosphorylation of EGFR, STAT‐5, PKB and ERK‐1/2, but not STAT‐3, in YD‐38 cells. Dasatinib induced activation of the intrinsic caspase pathway, which was inhibited by z‐VAD‐fmk, a pan‐caspase inhibitor. Dasatinib also decreased Mcl‐1 expression and S6 phosphorylation while increased GRP78 expression and eIF‐2α phosphorylation in YD‐38 cells. In addition, to its direct effects on YD‐38 cells, dasatinib also exhibited anti‐angiogenic properties. Dasatinib‐treated YD‐38 or HUVEC showed reduced HIF‐1α expression and stability. Dasatinib alone or conditioned media from dasatinib‐treated YD‐38 cells inhibited HUVEC tube formation on Matrigel without affecting HUVEC viability. Importantly, dasatinib's anti‐growth, anti‐angiogenic and pro‐apoptotic effects were additionally seen in tumorigenic HSC‐3 cells. Together, these results demonstrate that dasatinib has strong anti‐growth, anti‐angiogenic and pro‐apoptotic effects on human oral cancer cells, which are mediated through the regulation of multiple targets, including Src, EGFR, STAT‐3, STAT‐5, PKB, ERK‐1/2, S6, eIF‐2α, GRP78, caspase‐9/3, Mcl‐1 and HIF‐1α.

Transforming growth factor-β-regulated fractalkine as a marker of erosive bone invasion in oral squamous cell carcinoma.

Patients with oral squamous cell carcinoma (OSCC) bone invasion are surgically treated with bone resection, which results in severe physical and psychological damage. Here, we investigated the potential of fractalkine (CX3CL1), which is regulated by transforming growth factor (TGF-β), as a novel biomarker for correct prediction and early detection of OSCC-associated bone invasion. TGF-β knockdown and treatment with a TGF-β-neutralizing antibody decreased the level of fractalkine in the culture media of HSC-2 and YD10B OSCC cells. Treatment with a fractalkine-neutralizing antibody reduced TGF-β-stimulated invasion by HSC-2 and YD10B cells. Fractalkine treatment increased the viability, invasion, and uPA secretion of both OSCC cell lines. Furthermore, OSCC cell bone invasion was assessed following subcutaneous inoculation of wild-type or TGF-β knockdown OSCC cells in mouse calvaria. TGF-β knockdown prevented erosive bone invasion, reduced the number of osteoclasts at the tumor-bone interface, and downregulated fractalkine expression in mouse tumor tissues. Our results indicate that the production of fractalkine is stimulated by TGF-β and mediates TGF-β-induced cell invasion in several OSCC cell lines showing an erosive pattern of bone invasion. Fractalkine may be a useful predictive marker and therapeutic target for OSCC-induced bone destruction.

20(S)-Ginsenoside Rh2 Suppresses Oral Cancer Cell Growth by Inhibiting the Src-Raf-ERK Signaling Pathway.

20(S)-Ginsenoside Rh2 (G-Rh2) has demonstrated therapeutic effects in many types of cancers. We aimed to investigate the potential anticancer activity and underlying mechanisms of G-Rh2 in oral cancer cells. The antigrowth effect of G-Rh2 in oral cancer cells was stimulated by cell proliferation, soft agar colony formation, and migration and invasion assay. The cell cycle and apoptosis were detected by flow cytometry. The underlying mechanism of G-Rh2 in oral cancer cells was explored by immunoblotting. G-Rh2 significantly inhibited oral cancer cell growth by inducing apoptosis and cell cycle G0/G1-phase arrest. G-Rh2 inhibited oral cancer cell migration and invasion through regulation of epithelial-mesenchymal transition (EMT)-related proteins. G-Rh2 inhibited the Src/Raf/ERK signaling pathway in YD10B and Ca9-22 cells. G-Rh2 exerted anticancer activity in vitro by inhibiting the Src/Raf/ERK signaling pathway in oral cancer. G-Rh2 is a potential therapeutic drug for oral cancer treatment.

6]-Gingerol Suppresses Oral Cancer Cell Growth by Inducing the Activation of AMPK and Suppressing the AKT/mTOR Signaling Pathway.

[6]-Gingerol, a compound extracted from ginger, has been studied for its therapeutic potential in various types of cancers. However, its effects on oral cancer remain largely unknown. Here, we aimed to investigate the potential anticancer activity and underlying mechanisms of [6]-gingerol in oral cancer cells. We analyzed the antigrowth effects of [6]-gingerol in oral cancer cell lines by cell proliferation, colony formation, migration, and invasion assays. We detected cell cycle and apoptosis with flow cytometry and further explored the mechanisms of action by immunoblotting. [6]-Gingerol significantly inhibited oral cancer cell growth by inducing apoptosis and cell cycle G2/M phase arrest. [6]-Gingerol also inhibited oral cancer cell migration and invasion by up-regulating E-cadherin and down-regulating N-cadherin and vimentin. Moreover, [6]-gingerol induced the activation of AMPK and suppressed the AKT/mTOR signaling pathway in YD10B and Ca9-22 cells. [6]-Gingerol exerts anticancer activity by activating AMPK and suppressing the AKT/mTOR signaling pathway in oral cancer cells. Our findings highlight the potential of [6]-gingerol as a therapeutic drug for oral cancer treatment.

Resveratrol inhibits cell growth via targeting the Bmi-1 pathway in YD-10B human oral squamous cell carcinoma cells

Resveratrol has been reported to exert anticancer activity via modulation of multiple pathways and genes. In this study, we examined the effect of resveratrol on YD-10B human oral squamous cell carcinoma cells and its molecular mechanisms of action. We found that resveratrol inhibited the proliferation of YD-10B cells in a dose- and timedependent manner. The suppressive effect of resveratrol was accompanied by a reduction in Bmi-1 gene expression. We observed that silencing the Bmi-1 gene by small interfering RNA effectively downregulated the levels of GLUT1 mRNA and protein, which were also repressed by resveratrol. Bmi-1 silencing increased the number of YD-10B cells in S-phase arrest by approximately 2.3-fold compared with the control. In conclusion, the results of the present study demonstrate, for the first time, that resveratrol suppresses Bmi-1-mediated GLUT1 expression in human oral squamous cell carcinoma cells and suggest that the specific molecular targeting of Bmi-1 and/or GLUT1 expression can be combined with a chemotherapeutic strategy to improve the response of oral cancer cells to resveratrol.

Anticancer Effects of Cisplatin in Combination with Paeonia Japonica in YD-10B Cells

Anticancer Effects of Cisplatin in Combination with Paeonia Japonica in YD-10B Cells

Abstract 2860: RCP augments oral cancer cell invasion through Zeb1 and Slug expression

Abstract Background: Rab coupling protein (RCP), also called family interacting protein 1 (Rab11fip1) is a protein encoded by the RAB11FIP1 gene and has been known to augment cancer tumorigenesis, invasion, and metastasis. Recently, RCP was reported to aggravate cancer cell invasion and metastasis by stabilizing beta1 integrin and inducing epithelial-mesenchymal transition (EMT) in breast and ovarian cancer. Further, a high RCP expression was significantly correlated with shorter overall survival and disease-free survival of HNSCC patients. However, the molecule mechanisms by which RCP increases HNSCC invasion and metastasis are unclear. In the present study, we demonstrate the role of RCP in oral cancer cell invasiveness and its underlying mechanism. Methods: Oral cancer YD-10B and YD-38 cells were transiently transfected with RCP constructs. siRNAs were used to analyze the role of specific proteins for Slug and Zeb1 expression. Immunoblotting and immunofluorescence were used to analyze protein expression. Matrigel-coated in vitro insert and wound healing assay were used to analyze cancer cell invasion and migration, respectively. Results: We observed that ectopic expression of RCP induces invasion of oral cancer cells. Conversely, silencing of RCP expression increased E-cadherin expression and reduced oral cancer cell invasion. In addition, RCP upregulated Zeb1 and Slug expression that is important for RCP-induced oral cancer cell EMT and invasion. Furthermore, we identified that silencing of beta1 integrin efficiently attenuates RCP-induced Zeb1 and Slug expression. In addition, we observed that a selective inhibitor of EGFR kinase activity, Gefitinib, significantly reduces RCP-induced Zeb1 and Slug expression and that RasN17 recovers RCP-induced Zeb1 expression. Finally, RCP increased the viability of oral cancer cells against a chemotherapeutic agent cisplatin. Conclusion: Collectively, we demonstrate that RCP augments oral cancer cell EMT and invasiveness through a beta1 integrin/EGFR/Ras/Slug and Zeb1 signaling axis, providing potential biomarkers for oral cancer patients therapy. Citation Format: Jin Young Kim, Kyung Hwa Cho, Bo Young Jeong, Zi Hae Song, Chang Gyo Park, Hoi Young Lee. RCP augments oral cancer cell invasion through Zeb1 and Slug expression [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 2860.

Effect of STAT3 on Lysophosphatidic Acid-Induced Oral Cancer Cell Invasion

Background: Oral cancer has a high incidence worldwide and has been closely associated with smoking, alcohol, and infection by the human papillomavirus. Metastasis is highly important for oral cancer survival. Lysophosphatidic acid (LPA) is a bioactive lipid mediator that promotes various cellular processes, including cell survival, proliferation, metastasis, and invasion. Signal transducer and activator of transcription (STATs) are transcription factors that mediate gene expression. Among the seven types of STATs in mammals, STAT3 is involved in invasion and metastasis of numerous tumors. However, little is known about the role of STAT3 in oral tumor invasion. In the present study, we hypothesized that STAT3 mediates LPA-induced oral cancer invasion. Methods: Immunoblotting was performed to analyze LPA-induced STAT3 activation. 3-(4,5-Dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) assay was performed to assess the survival rates of YD-10B cells. STAT3 levels in LPA-treated oral tumor cells were evaluated by performing in vitro invasion assay. Results: To the best of our knowledge, this is the first study to demonstrate that LPA enhances STAT3 phosphorylation in oral cancer. In addition, treatment with WP1066, a selective inhibitor of STAT3, at a concentration that does not cause severe reduction in cell viability, significantly attenuated LPA-induced YD-10B cancer cell invasion. Conclusion: The results suggested that LPA induces oral tumor cells with greater invasive potential via STAT3 activation. Our findings provided important insights into the mechanisms underlying mouth neoplasms.

Meridianin C inhibits the growth of YD-10B human tongue cancer cells through macropinocytosis and the down-regulation of Dickkopf-related protein-3.

Meridianin C is a marine natural product known for its anti‐cancer activity. At present, the anti‐tumour effects of meridianin C on oral squamous cell carcinoma are unknown. Here, we investigated the effect of meridianin C on the proliferation of four different human tongue cancer cells, YD‐8, YD‐10B, YD‐38 and HSC‐3. Among the cells tested, meridianin C most strongly reduced the growth of YD‐10B cells; the most aggressive and tumorigenic of the cell lines tested. Strikingly, meridianin C induced a significant accumulation of macropinosomes in the YD‐10B cells; confirmed by the microscopic and TEM analysis as well as the entry of FITC‐dextran, which was sensitive to the macropinocytosis inhibitor amiloride. SEM data also revealed abundant long and thin membrane extensions that resemble lamellipodia on the surface of YD‐10B cells treated with meridianin C, pointing out that meridianin C‐induced macropinosomes was the result of macropinocytosis. In addition, meridianin C reduced cellular levels of Dickkopf‐related protein‐3 (DKK‐3), a known negative regulator of macropinocytosis. A role for DKK‐3 in regulating macropinocytosis in the YD‐10B cells was confirmed by siRNA knockdown of endogenous DKK‐3, which led to a partial accumulation of vacuoles and a reduction in cell proliferation, and by exogenous DKK‐3 overexpression, which resulted in a considerable inhibition of the meridianin C‐induced vacuole formation and decrease in cell survival. In summary, this is the first study reporting meridianin C has novel anti‐proliferative effects via macropinocytosis in the highly tumorigenic YD‐10B cell line and the effects are mediated in part through down‐regulation of DKK‐3.

Effect of Resveratrol on Oral Cancer Cell Invasion Induced by Lysophosphatidic Acid

The aim of the current study was to demonstrate the potential therapeutic efficacy of resveratrol in oral cancer patients. Lysophosphatidic acid (LPA) intensifies cancer cell invasion and metastasis, whereas resveratrol, a natural polyphenolic compound, possesses antitumor activity, suppressing cell proliferation and progression in various cancer cell lines (ovarian, gastric, oral, pancreatic, colon, and prostate cancer cells). In addition, resveratrol has been identified as an inhibitor of LPA-induced proteolytic enzyme expression and ovarian cancer invasion. Furthermore, resveratrol was shown to inhibit oral cancer cell invasion by downregulating hypoxia-inducible factor 1α and vascular endothelial growth factor expression. Recently, we demonstrated that LPA is important for the expression of transcription factors TWIST and SLUG during epithelial-mesenchymal transition (EMT) in oral squamous carcinoma cells. In this study, we treated serum-starved cultures of oral squamous carcinoma cell line YD-10B with resveratrol for 24 hours prior to stimulation with LPA. To identify an optimal resveratrol concentration that does not induce apoptosis in oral squamous carcinoma cells, we determined the toxicity of resveratrol in YD-10B cells by assessing their viability using the MTT assay. Another assay was performed using Matrigel-coated cell culture inserts to detect oral cancer cell invasion activity. Immunoblotting was applied for analyzing protein expression of SLUG, TWIST1, E-cadherin, and GAPDH. We demonstrated that resveratrol efficiently inhibited LPA-induced oral cancer cell EMT and invasion by downregulating SLUG and TWIST1 expression. Therefore, resveratrol may potentially reduce oral squamous carcinoma cell invasion and metastasis in oral cancer patients, improving their survival outcomes. In summary, we identified new targets for the development of therapies against oral cancer progression and characterized the therapeutic potential of resveratrol for the treatment of oral cancer patients.

Acetylshikonin suppresses invasion of Porphyromonasgingivalis‑infected YD10B oral cancer cells by modulating the interleukin-8/matrix metalloproteinase axis.

The development of pharmaceutical agents possessing anti-invasive and anti-metastatic abilities, as well as apoptotic activity, is important in decreasing the incidence and recurrence of oral cancer. Cancer cells are known to acquire invasiveness not only through epigenetic changes, but also from inflammatory stimuli within the tumor microenvironment. Accordingly, the identification of agents that can suppress the inflammation-promoted invasiveness of cancer cells may be important in treating cancer and improving the prognosis of patients with cancer. Acetylshikonin, a flavonoid with anti-inflammatory activity, inhibits proliferation and induces apoptosis of oral cancer cells. In the present study, the anti-invasive effect of acetylshikonin on YD10B oral cancer cells infected with Porphyromonas gingivalis, a major pathogen of chronic periodontitis, and the mechanisms involved were investigated. Firstly, we examined whether P. gingivalis infection increased the invasiveness of YD10B cells. Results suggested that YD10B oral cancer cells become more aggressive when they are infected with P. gingivalis. Secondly, acetylshikonin significantly inhibited the invasion of P. gingivalis-infected YD10B cells by suppressing IL-8 release and IL-8-dependent MMP release. These data suggest that acetylshikonin may be a useful preventive and therapeutic candidate for oral cancer that is chronically infected with periodontal pathogens.

MAPK/JNK1 activation protects cells against cadmium-induced autophagic cell death via differential regulation of catalase and heme oxygenase-1 in oral cancer cells

Antioxidant enzymes are related to oral diseases. We investigated the roles of heme oxygenase-1 (HO-1) and catalase in cadmium (Cd)-induced oxidative stress and the underlying molecular mechanism in oral cancer cells. Exposing YD8 cells to Cd reduced the expression levels of catalase and superoxide dismutase 1/2 and induced the expression of HO-1 as well as autophagy and apoptosis, which were reversed by N-acetyl-l-cysteine (NAC). Cd-exposed YD10B cells exhibited milder effects than YD8 cells, indicating that Cd sensitivity is associated with antioxidant enzymes and autophagy. Autophagy inhibition via pharmacologic and genetic modulations enhanced Cd-induced HO-1 expression, caspase-3 cleavage, and the production of reactive oxygen species (ROS). Ho-1 knockdown increased autophagy and apoptosis. Hemin treatment partially suppressed Cd-induced ROS production and apoptosis, but enhanced autophagy and CHOP expression, indicating that autophagy induction is associated with cellular stress. Catalase inhibition by pharmacological and genetic modulations increased Cd-induced ROS production, autophagy, and apoptosis, but suppressed HO-1, indicating that catalase is required for HO-1 induction. p38 inhibition upregulated Cd-induced phospho-JNK and catalase, but suppressed HO-1, autophagy, apoptosis. JNK suppression exhibited contrary results, enhancing the expression of phospho-p38. Co-suppression of p38 and JNK1 failed to upregulate catalase and procaspase-3, which were upregulated by JNK1 overexpression. Overall, the balance between the responses of p38 and JNK activation to Cd appears to have an important role in maintaining cellular homeostasis via the regulation of antioxidant enzymes and autophagy induction. In addition, the upregulation of catalase by JNK1 activation can play a critical role in cell protection against Cd-induced oxidative stress.

JNK-associated scattered growth of YD-10B oral squamous carcinoma cells while maintaining the epithelial phenotype

Cell scattering of epithelial carcinoma cancer cells is one of the critical event in tumorigenesis. Cells losing epithelial cohesion detach from aggregated epithelial cell masses and may migrate to fatal organs through metastasis. The present study investigated the molecular mechanism by which squamous cell carcinoma cells grow scattered at the early phase of transformation while maintaining the epithelial phenotype. We studied YD-10B cells, which are established from human oral squamous cell carcinoma, because the cells grow scattered without the development of E-cadherin junctions (ECJs) under routine culture conditions despite the high expression of functional E-cadherin. The functionality of their E-cadherin was demonstrated in that YD-10B cells developed ECJs, transiently or persistently, when they were cultured on substrates coated with a low amount of fibronectin or to confluence. The phosphorylation of JNK was up-regulated in YD-10B cells compared with that in human normal oral keratinocyte cells or human squamous cell carcinoma cells, which grew aggregated along with well-organized ECJs. The suppression of JNK activity induced the aggregated growth of YD-10B cells concomitant with the development of ECJs. These results indicate for the first time that inherently up-regulated JNK activity induces the scattered growth of the oral squamous cell carcinoma cells through down-regulating the development of ECJ despite the expression of functional E-cadherin, a hallmark of the epithelial phenotype.

YD-10B 인간구강암세포주에서 생강 유기용매 분획물의 항산화, 트롬빈억제 및 thrombin에 의해 처리된 암 침윤 및 전이 억제 효과

YD-10B 인간구강암세포주에서 생강 유기용매 분획물의 항산화, 트롬빈억제 및 thrombin에 의해 처리된 암 침윤 및 전이 억제 효과

Heme oxygenase-1-mediated apoptosis under cadmium-induced oxidative stress is regulated by autophagy, which is sensitized by tumor suppressor p53

Heme oxygenase-1 (HO-1) is a stress-inducible cytoprotective enzyme. It is often overexpressed in different types of cancers and promotes cell survival. However, the role of HO-1 and the underlying molecular mechanism of cadmium (Cd)-induced oxidative stress in cancer cells remain undefined. Here we show that the role of HO-1 under Cd-induced oxidative stress is dependent upon autophagy, which is sensitized by the tumor suppressor p53. The sensitivity to Cd was 3.5- and 14-fold higher in p53-expressing YD8 and H460 cells than in p53-null YD10B and H1299 cells, respectively. The levels of p53 in YD8 and H460 cells decreased in a Cd concentration-dependent manner, which was inhibited by pretreatment with N-acetylcysteine. In both cell lines, Cd exposure resulted in caspase-3-mediated PARP-1 cleavage and the induction of CHOP, LC3-II, and HO-1, which were limited in YD10B and H1299 cells exposed to high concentrations of Cd. Cd exposure to p53-overexpressing YD10B cells enhanced Cd-induced HO-1 and LC3-II levels, whereas genetic knockdown of p53 in YD8 cells resulted in the suppression of Cd-induced levels of HO-1 and LC3-II, indicating that p53 is required in the sensing of HO-1 and induction of autophagy. The inhibition of autophagy using small interfering RNA (siRNA) for the autophagy-related gene atg5 enhanced HO-1, CHOP, and PARP-1 cleavage induced by Cd. However, transfection with HO-1 siRNA increased Cd-induced LC3-II, and suppressed the expression of CHOP and cleavage of PARP-1. Collectively, the role of HO-1 in apoptosis could be modulated by autophagy, which is sensitized by p53 expression in human cancer cell lines.