Achaete-scute family bHLH transcription factor 2 activation promotes hepatoblastoma progression.

Achaete-scute family bHLH transcription factor 2 (ASCL2) is highly expressed in hepatoblastoma (HB) tissues, but its role remains unclear. Thus, biological changes in the HB cell line HepG2 in response to induced ASCL2 expression were assessed. ASCL2 expression was induced in HepG2 cells using the Tet-On 3G system, which includes doxycycline. Cell viability, proliferation activity, mobility, and stemness were evaluated using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide, colony-formation, migration, invasion, and sphere-formation assays. Quantitative reverse-transcription polymerase chain reaction was used to assess the expression of markers for proliferation (CCND1 and MYC), epithelial-mesenchymal transition (EMT; SNAI1, TWIST1, and ZEB1), mesenchymal-epithelial transition (CDH1), and stemness (KLF4, POU5F1, and SOX9). Compared with the non-induced HepG2 cells, cells with induced ASCL2 expression showed significant increases in viability, colony number, migration area (%), and sphere number on days 7, 14, 8, and 7, respectively, and invasion area (%) after 90 h. Furthermore, induction of ASCL2 expression significantly upregulated CCND1, MYC, POU5F1, SOX9, and KLF4 expression on days 2, 2, 3, 3, and 5, respectively, and increased the ratios of SNAI1, TWIST1, and ZEB1 to CDH1 on day 5. ASCL2 promoted the formation of malignant phenotypes in HepG2 cells, which may be correlated with the upregulation of the Wnt signaling pathway-, EMT-, and stemness-related genes. ASCL2 activation may therefore be involved in the progression of HB.

Resveratrol exhibits diverse anti-cancer activities through epigenetic regulation of E-cadherin and p21 in triple-negative breast cancer cells.

BACKGROUND: Triple-negative breast cancer (TNBC) has an aggressive phenotype and poor outcome, however no specific targeted therapy has been established for TNBC lacking germline BRCA1/2 pathogenic variants. To develop a novel therapeutic strategy, we explored the potential of resveratrol (RSV) for TNBC treatment. METHODS: We investigated the effects of RSV on malignant phenotypes of TNBC cells as well as on apoptosis induced by ABT263, a specific inhibitor of BCL-2 and BCL-xL, using morphological observation, migration assay, ß-galactosidase staining, and Hoechst staining. To elucidate the underlying mechanisms of RSV-mediated effects, expression levels and histone acetylation levels of cadherin 1 (CDH1, E-cadherin) and cyclin dependent kinase inhibitor 1A (CDKN1A, p21) were determined by RT-qPCR, western blotting, and chromatin immunoprecipitation. Furthermore, knockdown analysis was conducted to evaluate the involvement of E-cadherin and/or p21 in RSV potentiation on cytotoxic activity of ABT263. RESULTS: RSV treatment induced epithelial-like cellular morphology and suppressed the migration capacity in MDA-MB-231 and BT-549-Luc TNBC cells. ß-galactosidase-positive cells were increased after RSV treatment, indicating the induction of cellular senescence, in MDA-MB-231 cells but not in BT-549-Luc cells. RSV increased the expression and histone acetylation of CDH1 and CDKN1A in both cells. Interestingly, pre-treatment with RSV enhanced the induction of apoptosis in the ABT263-treated MDA-MB-231 and BT-549-Luc cells, and knockdown of CDKN1A decreased ABT263-induced apoptosis in RSV-treated MDA-MB-231 cells. CONCLUSIONS: RSV represses the metastatic capacity and enhances the cytotoxic activity of ABT263 in TNBC cells. Our results suggested that RSV can potentially be used as a repressor of metastasis or a sensitizer to ABT263 for TNBC treatment via up-regulation of CDH1 and CDKN1A through epigenetic mechanisms.

Bovine Lactoferrin Suppresses Tumor Angiogenesis through NF-κB Pathway Inhibition by Binding to TRAF6.

Tumor angiogenesis is essential for tumor progression. The inhibition of tumor angiogenesis is a promising therapy for tumors. Bovine lactoferrin (bLF) has been reported as an anti-tumor agent. However, bLF effects on tumor angiogenesis are not well demonstrated. This study evaluated the inhibitory effects of bLF on tumor angiogenesis in vivo and in vitro. Herein, tumor endothelial cells (TECs) and normal endothelial cells (NECs) were used. Proliferation, migration, tube formation assays, RT-PCR, flow cytometry, Western blotting, siRNA experiments and immunoprecipitation were conducted to clarify the mechanisms of bLF-induced effects. CD-31 immunoexpression was examined in tumor tissues of oral squamous cell carcinoma mouse models with or without Liposomal bLF (LbLF)-administration. We confirmed that bLF inhibited proliferation/migration/tube formation and increased apoptosis in TECs but not NECs. TNF receptor-associated factor 6 (TRAF6), p-p65, hypoxia inducible factor-α (HIF-1α) and vascular endothelial growth factor (VEGF) were highly expressed in TECs. In TECs, bLF markedly downregulated VEGF-A, VEGF receptor (VEGFR) and HIF-1α via the inhibition of p-p65 through binding with TRAF6. Since NECs slightly expressed p-p65, bLF-TRAF-6 binding could not induce detectable changes. Moreover, orally administrated LbLF decreased CD31-positive microvascular density only in TECs. Hence, bLF specifically suppressed tumor angiogenesis through p-p65 inhibition by binding to TRAF6 and suppressing HIF-1α activation followed by VEGF/VEGFR down-regulation. Collectively, bLF can be an anti-angiogenic agent for tumors.

Oncogenic Role of ADAM32 in Hepatoblastoma: A Potential Molecular Target for Therapy.

Outcomes of pediatric hepatoblastoma (HBL) have improved, but refractory cases still occur. More effective and safer drugs are needed that are based on molecular mechanisms. A disintegrin and metalloproteases (ADAMs) are expressed with high frequency in various human carcinomas and play an important role in cancer progression. In this study, we analyzed expression of ADAMs in HBL with a cDNA microarray dataset and found that the expression level of ADAM32 is particularly high. To investigate the role of ADAM32 in cancer, forced expression or knockdown experiments were conducted with HepG2 and HBL primary cells. Colony formation, cell migration and invasion, and cell viability were increased in HepG2 expressing ADAM32, whereas knockdown of ADAM32 induced a decrease in these cellular functions. Quantitative RT-PCR demonstrated an association between ADAM32 expression and the expression of genes related to cancer stem cells and epithelial-mesenchymal transition (EMT), suggesting a role of ADAM32 in cancer stemness and EMT. Furthermore, knockdown of ADAM32 increased cisplatin-induced apoptosis, and this effect was attenuated by a caspase-8 inhibitor, suggesting that ADAM32 plays a role in extrinsic apoptosis signaling. We conclude that ADAM32 plays a crucial role in progression of HBL, so it might be a promising molecular target in anticancer therapy.

p53 status modifies cytotoxic activity of lactoferrin under hypoxic conditions.

Lactoferrin (LF) is an iron binding glycoprotein of the transferrin family with a wide spectrum of biological effects, including anti-cancer activity. However, the detailed molecular mechanisms of anti-cancer activity of LF have not been fully determined. In this study, we tried to clarify cytotoxic functions of LF on various cell lines under hypoxic conditions and elucidate those molecular mechanisms. Cytotoxic activity of LF on cell lines was found to have a range of sensitivities. Hypoxia decreased sensitivity to LF in KD (lip fibroblast) but increased that in HSC2 (oral squamous cell carcinoma). Expression analyses further revealed that LF treatments increased hypoxic HIF-1α, -2α and p53 proteins in KD but attenuated them in HSC2 cells, and decreased HIF-1 target gene, DEC2, in KD but increased it in HSC2, suggesting a possible relationship between LF-modified DEC2 expression and HIF-α protein. MTT assay strikingly demonstrated that cells expressing mutant-type p53 (MT5) were more sensitive to LF than control HepG2 (hepatoma), suggesting an important role of the p53 signal. Knock-down of TP53 (p53 gene) interestingly reduced sensitivity to LF in HepG2, suggesting that p53 may be a target of LF cytotoxic activity. Further analyses with a ferroptosis promoter or inhibitor demonstrated that LF increased ACSL4 in hypoxic MT5, suggesting LF-induced ferroptosis in cells expressing mutant-type p53. In conclusion, hypoxia was found to regulate cytotoxic activities of LF differently among various cell lines, possibly through the p53 signaling pathway. LF further appeared to regulate ferroptosis through a modification of ACSL4 expression.

Low-Dose-Rate Irradiation Suppresses the Expression of Cell Cycle-Related Genes, Resulting in Modification of Sensitivity to Anti-Cancer Drugs.

The biological effects of low-dose-rate (LDR) radiation exposure in nuclear power plant accidents and medical uses of ionizing radiation (IR), although being a social concern, remain unclear. In this study, we evaluated the effects of LDR-IR on global gene expression in human cells and aimed to clarify the mechanisms. RNA-seq analyses demonstrated that relatively low dose rates of IR modify gene expression levels in TIG-3 cells under normoxic conditions, but those effects were attenuated under hypoxia-mimicking conditions. Gene set enrichment analysis demonstrated that LDR-IR significantly decreased gene expression related to cell division, cell cycle, mitosis, and the Aurora kinase B and FOXM1 pathways. Quantitative RT-PCR confirmed the down-regulation of AURKB and FOXM1 genes in TIG-3 cells with LDR-IR or hypoxia-mimicking treatments without any dose-rate effect. Knock-down experiments suggested that HIF-1α and HIF-2α, as well as DEC1, participated in down-regulation of AURKB and FOXM1 under DFOM treatments, but to a lesser extent under LDR-IR treatment. FACS and microscopic analyses demonstrated that LDR-IR induced G0/G1 arrest and increased micronucleus or chromosome condensation. Finally, MTT assays demonstrated that LDR-IR decreased sensitivity to paclitaxel or barasertib in TIG-3 cells but not in A549 cells. In conclusion, LDR-IR modifies global gene expression and cell cycle control, resulting in a reduction of sensitivity to anti-cancer chemotherapy in non-cancer cells and thus a reduction in untoward effects (GA).

The roles of Y-box-binding protein (YB)-1 and C-X-C motif chemokine ligand 14 (CXCL14) in the progression of prostate cancer via extracellular-signal-regulated kinase (ERK) signaling.

The cold-shock protein Y-box-binding protein (YB)-1 regulates the expression of various chemokines and their receptors at the transcriptional level. Expression of the orphan chemokine CXCL14 is repressed by EGF induced signaling. The possible links between EGF-mediated YB-1 and CXCL14 as well as the functions of critical kinase pathways in the progression of prostate cancer have remained unexplored. Here we examined the correlation between YB-1 and CXCL14, and the ERK/AKT/mTOR pathways in prostate cancer. Knockdown of YB-1 decreased cyclinD1 expression with an upregulation of cleaved-PARP in human prostate cancer cells. EGF treatment upregulated phospho-YB-1 expression in a time-dependent manner, while treatment with an ERK inhibitor completely silenced its expression in prostate cancer cells. EGF treatment stimulates CyclinD1 and YB-1 phosphorylation in an ERK-dependent pathway. Positive and negative regulation of YB-1 and CXCL14 was observed after EGF treatment in prostate cancer cells, respectively. EGF rescues cell cycle and apoptosis via the AKT and ERK pathways. Furthermore, YB-1 silencing induces G1 arrest and apoptosis, while knockdown of CXCL14 facilitates cell growth and inhibits apoptosis in prostate cancer cells. YB-1 and CXCL14 were inversely correlated in prostate cancer cells and tissues. A significant association between poor overall survival and High YB-1 expression was observed in human prostate cancer patients. In conclusion, our data reveal the functional relationship between YB-1 and CXCL14 in EGF mediated ERK signaling, and YB-1 expression is a significant prognostic marker to predict prostate cancer.

Inhibiting SARS-CoV-2 infection in vitro by suppressing its receptor, angiotensin-converting enzyme 2, via aryl-hydrocarbon receptor signal.

Since understanding molecular mechanisms of SARS-CoV-2 infection is extremely important for developing effective therapies against COVID-19, we focused on the internalization mechanism of SARS-CoV-2 via ACE2. Although cigarette smoke is generally believed to be harmful to the pathogenesis of COVID-19, cigarette smoke extract (CSE) treatments were surprisingly found to suppress the expression of ACE2 in HepG2 cells. We thus tried to clarify the mechanism of CSE effects on expression of ACE2 in mammalian cells. Because RNA-seq analysis suggested that suppressive effects on ACE2 might be inversely correlated with induction of the genes regulated by aryl hydrocarbon receptor (AHR), the AHR agonists 6-formylindolo(3,2-b)carbazole (FICZ) and omeprazole (OMP) were tested to assess whether those treatments affected ACE2 expression. Both FICZ and OMP clearly suppressed ACE2 expression in a dose-dependent manner along with inducing CYP1A1. Knock-down experiments indicated a reduction of ACE2 by FICZ treatment in an AHR-dependent manner. Finally, treatments of AHR agonists inhibited SARS-CoV-2 infection into Vero E6 cells as determined with immunoblotting analyses detecting SARS-CoV-2 specific nucleocapsid protein. We here demonstrate that treatment with AHR agonists, including FICZ, and OMP, decreases expression of ACE2 via AHR activation, resulting in suppression of SARS-CoV-2 infection in mammalian cells.

Tamoxifen resistance alters sensitivity to 5-fluorouracil in a subset of estrogen receptor-positive breast cancer.

Sequential treatment with endocrine or chemotherapy is generally used in the treatment of estrogen receptor (ER)-positive recurrent breast cancer. To date, few studies have investigated the effect of long-term endocrine therapy on the response to subsequent chemotherapy in ER-positive breast cancer. We examined whether a preceding endocrine therapy affects the sensitivity to subsequent chemotherapy in ER-positive breast cancer cells. Three ER-positive breast cancer cell lines (T47D, MCF7, BT474) and tamoxifen-resistant sublines (T47D/T, MCF7/T, BT474/T) were analyzed for sensitivity to 5-fluorouracil, paclitaxel, and doxorubicin. The mRNA levels of factors related to drug sensitivity were analyzed by RT-PCR. MCF7/T cells became more sensitive to 5-fluorouracil than wild-type (wt)-MCF7 cells. In addition, the apoptosis induced by 5-fluorouracil was significantly increased in MCF7/T cells. However, no difference in sensitivity to chemotherapeutic agents was observed in T47D/T and BT474/T cells compared with their wt cells. Dihydropyrimidine dehydrogenase (DPYD) mRNA expression was significantly decreased in MCF7/T cells compared with wt-MCF7 cells. The expression of DPYD mRNA was restored with 5-azacytidine treatment in MCF7/T cells. In addition, DPYD 3'-UTR luciferase activity was significantly reduced in MCF7/T cells. These data indicated that the expression of DPYD mRNA was repressed by methylation of the DPYD promoter region and post-transcriptional regulation by miRNA in MCF7/T cells. In the mouse xenograft model, capecitabine significantly reduced the tumor volume in MCF7/T compared with MCF7. The results of this study indicate that endocrine therapy could alter the sensitivity to chemotherapeutic agents in a subset of breast cancers, and 5-fluorouracil may be effective in tamoxifen-resistant breast cancers.

GLIS1, a novel hypoxia-inducible transcription factor, promotes breast cancer cell motility via activation of WNT5A.

We previously demonstrated that expression of a Krüppel-like zinc finger transcription factor, GLIS1, dramatically increases under hypoxic conditions via a transcriptional mechanism induced by HIF-2α cooperating with AP-1 members. In this study, we focused on the functional roles of GLIS1 in breast cancer. To uncover its biological function, the effects of altered levels of GLIS1 in breast cancer cell lines on cellular growth, wound-healing and invasion capacities were assessed. Knockdown of GLIS1 using siRNA in BT-474 cells resulted in significant growth stimulation under normoxia, while attenuation was found in the cell invasion assay under hypoxic conditions. In MDA-MB-231 cells expressing exogenous 3xFLAG-tagged GLIS1, GLIS1 attenuated cell proliferation and enhanced cell mobility and invasion capacities under normoxia. In addition, breast cancer cells expressing GLIS1 acquired resistance to irradiation. Whole transcriptome analysis clearly demonstrated that downstream signals of GLIS1 are related to various cellular functions. Among the genes with increased expression, we focused on WNT5A. Knockdown of WNT5A indicated that enhancement of acquired cell motility in the MDA-MB-231 cells expressing GLIS1 was mediated, at least in part, by WNT5A. In an analysis of publicly available data, patients with estrogen receptor-negative breast cancer showing high levels of GLIS1 expression showed much worse prognosis than those with low levels. In summary, hypoxia-induced GLIS1 plays significant roles in breast cancer cells via regulation of gene expression related to cell migration and invasion capacities, resulting in poorer prognosis in patients with advanced breast cancer.

Simulated microgravity enhances CDDP-induced apoptosis signal via p53-independent mechanisms in cancer cells.

Although the biological systems in the human body are affected by the earth's gravity, information about the underlying molecular mechanisms is limited. For example, apoptotic signaling is enhanced in cancer cells subjected to microgravity. We reasoned that signaling regulated by p53 may be involved because of its role in apoptosis. Therefore, we aimed to clarify the molecular mechanisms of modified cis-diamminedichloroplatinum (CDDP)-sensitivity under simulated microgravity by focusing on p53-related cell death mechanisms. Immunoblotting analyses indicated that, under microgravity, CDDP-induced ATM/p53 signaling increased and caspase-3 was cleaved earlier. However, microgravity decreased the levels of expression of p53 targets BAX and CDKN1A. Interestingly, microgravity increased the PTEN, DRAM1, and PRKAA1 mRNA levels. However, microgravity decreased the levels of mTOR and increased the LC3-II/I ratio, suggesting the activation of autophagy. The CDDP-induced cleavage of caspase-3 was increased during the early phase in Group MG (+), and cleaved caspase-3 was detected even in Group MG (+) with constitutive expression of a mutant type of p53 (hereafter, "+" indicates CDDP treatment). These results interestingly indicate that microgravity altered CDDP sensitivity through activation of caspase-3 by p53-independent mechanism.

Differentiated embryo chondrocyte plays a crucial role in DNA damage response via transcriptional regulation under hypoxic conditions.

Tumor hypoxia contributes to a biologically aggressive phenotype and therapeutic resistance. Recent studies have revealed that hypoxia reduces expression of several DNA damage recognition and repair (DRR) genes via both hypoxia-inducible factor (HIF)-independent and -dependent pathways, and this induced genomic instability in cancer cells. We show here that one of the HIF-target genes-differentiated embryo chondrocyte (DEC)-plays a role in DNA damage response via transcriptional repression. Comprehensive gene expression and database analyses have revealed systemic repression of DNA-DRR genes in cancer and non-cancer cells under hypoxic conditions. Hypoxic repression in typical cases was confirmed by quantitative RT-PCR and promoter reporter experiments, and knockdown experiments indicated the critical role of DEC2 in such repression. Assessment of histone H2AX phosphorylation revealed that recognition and repair of DNA double-strand breaks (DSBs) induced by bleomycin or γ-ray irradiation were attenuated; moreover, Cleaved Caspase-3 levels were decreased with pre-conditioning under hypoxia: opposing phenomena were ascertained by knockdown of DEC2. Finally, pre-conditioning under hypoxia decreased the sensitivity of cancer cells to DSBs, and knockdown of DEC2 increased γ-ray sensitivity. These data imply that a critical reduction of DNA-DRR occurs via DEC-dependent transcriptional repression and suggest that DEC is a potential molecular target for anti-cancer strategies.

A Chemical Modulator of p53 Transactivation that Acts as a Radioprotective Agonist.

Inhibiting p53-dependent apoptosis by inhibitors of p53 is an effective strategy for preventing radiation-induced damage in hematopoietic lineages, while p53 and p21 also play radioprotective roles in the gastrointestinal epithelium. We previously identified some zinc(II) chelators, including 8-quinolinol derivatives, that suppress apoptosis in attempts to discover compounds that target the zinc-binding site in p53. We found that 5-chloro-8-quinolinol (5CHQ) has a unique p53-modulating activity that shifts its transactivation from proapoptotic to protective responses, including enhancing p21 induction and suppressing PUMA induction. This p53-modulating activity also influenced p53 and p53-target gene expression in unirradiated cells without inducing DNA damage. The specificity of 5CHQ for p53 and p21 was demonstrated by silencing the expression of each protein. These effects seem to be attributable to the sequence-specific alteration of p53 DNA-binding, as evaluated by chromatin immunoprecipitation and electrophoretic mobility shift assays. In addition, 5-chloro-8-methoxyquinoline itself had no antiapoptotic activity, indicating that the hydroxyl group at the 8-position is required for its antiapoptotic activity. We applied this remarkable agonistic activity to protecting the hematopoietic and gastrointestinal system in mouse irradiation models. The dose reduction factors of 5CHQ in total-body and abdominally irradiated mice were about 1.2 and 1.3, respectively. 5CHQ effectively protected mouse epithelial stem cells from a lethal dose of abdominal irradiation. Furthermore, the specificity of 5CHQ for p53 in reducing the lethality induced by abdominal irradiation was revealed in Trp53-KO mice. These results indicate that the pharmacologic upregulation of radioprotective p53 target genes is an effective strategy for addressing the gastrointestinal syndrome. Mol Cancer Ther; 17(2); 432-42. ©2017 AACRSee all articles in this MCT Focus section, "Developmental Therapeutics in Radiation Oncology."

Genetics of the hypoxia-inducible factors in human cancers.

Professor Lorenz Poellinger (Karolinska Institute/Cancer Science Institute of Singapore) passed away in March 2016. Then hypoxic research field lost a noble and unique researcher when he died, since he had contributed very much to this field in a variety of aspects. We had been collaborating on a various research projects of genomic analyses of HIF-signaling pathway for a long time, and recently reported several interesting results with HIF-α genes (HIF1A, EPAS1 and HIF3A). Genomics/Genetics is a still growing field, with new technologies appearing often, and many groups have performed extensive genomic/genetic analyses. In this review, I thus focused on the genetics of HIF-α genes in human cancers. I deeply mourn Professor Poellinger's loss and dedicate this review to him.

Association between Functional Capacity Decline and Nutritional Status Based on the Nutrition Screening Initiative Checklist: A 2-Year Cohort Study of Japanese Community-Dwelling Elderly.

AIM: To assess whether nutritional status based on the Nutrition Screening Initiative Checklist is useful for predicting functional capacity decline in community-dwelling Japanese elderly. METHODS: This two-year observational cohort study included 536 community-dwelling Japanese (65 years and older at baseline) who were independent in both activities and instrumental activities of daily living. Demographic attributes, chronic illness, lifestyle-related habits, nutritional status, functional capacity, and anthropometric measurements were assessed, with decline in functional capacity used as the outcome measure. RESULTS: Subjects were classified into three groups as follows based on the Nutrition Screening Initiative Checklist: low (59.5%), moderate (23.7%), and high (16.8%) nutritional risk. Significant differences were found between nutritional status and the following four baseline variables: age, hypertension, cerebrovascular diseases, and current smoking. However, no significant differences were evident between nutritional status and sex, body mass index, diabetes, drinking habit, or exercise habit. Logistic regression analysis adjusted for age, sex, body mass index, hypertension, cerebrovascular diseases and smoking habit showed that the high nutritional risk group was significantly associated with a decline in both activities of daily living (odds ratio: 4.96; 95% confidence interval (CI): 1.59-15.50) and instrumental activities of daily living (OR: 2.58; 95% CI: 1.31-5.06) compared with the low nutritional risk group. CONCLUSIONS: Poor nutritional status based on the Nutrition Screening Initiative Checklist was associated with a decline in functional capacity over a 2-year period in community-dwelling Japanese elderly. These results suggest that the Nutrition Screening Initiative Checklist is a suitable tool for predicting functional capacity decline in community-dwelling elderly.

Suitable Food Textures for Videofluoroscopic Studies of Swallowing in Esophageal Cancer Cases to Prevent Aspiration Pneumonia.

AIMS: To determine suitable food textures for videofluoroscopic study of swallowing (VFSS), in order to predict and prevent subsequent aspiration pneumonia in esophageal cancer patients with dysphagia after surgery. MATERIALS AND METHODS: We evaluated 45 hospitalized esophageal cancer patients who underwent surgery between January 2012 and December 2013. The control group consisted of 43 patients treatmed from January 2010 until December 2011 and were not examined by VFSS. Test foods, which were presented in order of increasing thickness, included thin barium sulfate (Ba) liquid (3 or 10 ml), slightly thickened Ba liquid (3 or 10 ml), a spoonful of Ba jelly, and a spoonful of Ba puree. RESULTS: Patients could most safely swallow puree, followed by jelly. The 3mL samples of both the thin and thick liquids put patients at risk for aspiration pneumonia, with incidence rates of 13% and 11%, respectively. While 64.4% of patients could swallow all test foods and liquids safely, 35.6% were at risk for aspiration pneumonia when swallowing liquids. Even though >30% of patients were at risk, only 1 (2.2%) in the VFSS group developed aspiration pneumonia, which occurred at the time of admission. Following VFSS, no incidence of aspiration pneumonia was observed. However, aspiration pneumonia occurred in 4 (9.3%) control patients during hospitalization. CONCLUSIONS: Postoperative esophageal cancer patients were more likely to aspirate any kind of liquid than solid foods, such as jellies. VFSS is very useful in determining suitable food textures for postoperative esophageal cancer patients.

OASIS modulates hypoxia pathway activity to regulate bone angiogenesis.

OASIS/CREB3L1, an endoplasmic reticulum (ER)-resident transcription factor, plays important roles in osteoblast differentiation. In this study, we identified new crosstalk between OASIS and the hypoxia signaling pathway, which regulates vascularization during bone development. RT-PCR and real-time PCR analyses revealed significant decreases in the expression levels of hypoxia-inducible factor-1α (HIF-1α) target genes such as vascular endothelial growth factor A (VEGFA) in OASIS-deficient (Oasis(-/-)) mouse embryonic fibroblasts. In coimmunoprecipitation experiments, the N-terminal fragment of OASIS (OASIS-N; activated form of OASIS) bound to HIF-1α through the bZIP domain. Luciferase assays showed that OASIS-N promoted the transcription activities of a reporter gene via a hypoxia-response element (HRE). Furthermore, the expression levels of an angiogenic factor Vegfa was decreased in Oasis(-/-) osteoblasts. Immunostaining and metatarsal angiogenesis assay showed retarded vascularization in bone tissue of Oasis(-/-) mice. These results suggest that OASIS affects the expression of HIF-1α target genes through the protein interaction with HIF-1α, and that OASIS-HIF-1α complexes may play essential roles in angiogenesis during bone development.

The A Allele at rs13419896 of EPAS1 Is Associated with Enhanced Expression and Poor Prognosis for Non-Small Cell Lung Cancer.

Hypoxia-inducible factor-2α (HIF-2α, or EPAS1) is important for cancer progression, and is a putative biomarker for poor prognosis for non-small cell lung cancer (NSCLC). However, molecular mechanisms underlying the EPAS1 overexpression are not still fully understood. We explored a role of a single nucleotide polymorphism (SNP), rs13419896 located within intron 1 of the EPAS1 gene in regulation of its expression. Bioinformatic analyses suggested that a region including the rs13419896 SNP plays a role in regulation of the EPAS1 gene expression and the SNP alters the binding activity of transcription factors. In vitro analyses demonstrated that a fragment containing the SNP locus function as a regulatory region and that a fragment with A allele showed higher transactivation activity than one with G, especially in the presence of overexpressed c-Fos or c-Jun. Moreover, NSCLC patients with the A allele showed poorer prognosis than those with G at the SNP even after adjustment with various variables. In conclusion, the genetic polymorphism of the EPAS1 gene may lead to variation of its gene expression levels to drive progression of the cancer and serve as a prognostic marker for NSCLC.

Reversible hypopituitarism with pituitary tuberculoma.

A 50-year-old woman presented with a headache and nausea. A sellar and suprasellar mass was detected on MRI; the tumor was heterogeneously enhanced with gadolinium, and the pituitary stalk was slightly thickened. Laboratory tests revealed severe growth hormone, luteinizing hormone, follicle-stimulating hormone and thyroid-stimulating hormone deficiencies. A pathological examination of the tumor showed scattered granulomas with central necrosis and Langhans giant cells. Tuberculin skin and QuantiFERON TB-Gold tests (QFT-2G) were positive. Accordingly, we diagnosed the patient with pituitary tuberculoma presenting with pituitary dysfunction. Following treatment with antituberculous drugs, the pituitary hormone function normalized and the pituitary tuberculoma disappeared.

UCHL1 provides diagnostic and antimetastatic strategies due to its deubiquitinating effect on HIF-1α.

Hypoxia-inducible factor 1 (HIF-1) plays a role in tumour metastases; however, the genes that activate HIF-1 and subsequently promote metastases have yet to be identified. Here we show that Ubiquitin C-terminal hydrolase-L1 (UCHL1) abrogates the von Hippel-Lindau-mediated ubiquitination of HIF-1α, the regulatory subunit of HIF-1, and consequently promotes metastasis. The aberrant overexpression of UCHL1 facilitates distant tumour metastases in a HIF-1-dependent manner in murine models of pulmonary metastasis. Meanwhile, blockade of the UCHL1-HIF-1 axis suppresses the formation of metastatic tumours. The expression levels of UCHL1 correlate with those of HIF-1α and are strongly associated with the poor prognosis of breast and lung cancer patients. These results indicate that UCHL1 promotes metastases as a deubiquitinating enzyme for HIF-1α, which justifies exploiting it as a prognostic marker and therapeutic target of cancers.