Thyroid hormone upregulates LAMP2 expression and lysosome activity.

Thyroid hormone (T3)-induced autophagy and its biological significance have been extensively investigated in recent years. However, limited studies to date have focused on the important role of lysosomes in autophagy. In this study, we explored the effects of T3 on lysosomal protein expression and trafficking in detail. Our findings showed that T3 activates rapid lysosomal turnover and expression of numerous lysosomal genes, including TFEB, LAMP2, ARSB, GBA, PSAP, ATP6V0B, ATP6V0D1, ATP6V1E1, CTSB, CTSH, CTSL, and CTSS, in a thyroid hormone receptor-dependent manner. In a murine model, LAMP2 protein was specifically induced in mice with hyperthyroidism. T3-promoted microtubule assembly was significantly disrupted by vinblastine, resulting in accumulation of the lipid droplet marker PLIN2. In the presence of the lysosomal autophagy inhibitors bafilomycin A1, chloroquine and ammonium chloride, we observed substantial accumulation of LAMP2 but not LAMP1 protein. T3 further enhanced the protein levels of ectopically expressed LAMP1 and LAMP2. Upon knockdown of LAMP2, cavities of lysosomes and lipid droplets accumulated in the presence of T3, although the changes in LAMP1 and PLIN2 expression were less pronounced. More specifically, the protective effect of T3 against ER stress-induced death was abolished by knockdown of LAMP2. Our collective results indicate that T3 not only promotes lysosomal gene expression but also LAMP protein stability and microtubule assembly, leading to enhancement of lysosomal activity in digesting any additional autophagosomal burden.

Loss of core-fucosylation of SPARC impairs collagen binding and contributes to COPD.

Chronic obstructive pulmonary disease (COPD) is a progressive lung disease with high morbidity and mortality worldwide. Although several mechanisms to account for deleterious immune effects were proposed, molecular description for the underlying alveolar structural alterations for COPD is lacking. Here, silencing of α1,6-fucosyltransferase (Fut8), the enzyme for core-fucosylation and highly expressed in lung stem cells, resulted in alveolar structural changes in lung organoids, recapitulating COPD. Site-specific mass spectrometry analysis demonstrated that the secreted protein acidic and rich in cysteine (SPARC), which binds collagen, contains a core-fucosylation site in its VCSNDNcfK glycopeptide. Biacore assay showed markedly reduced collagen binding of SPARC lacking core fucosylation. Molecular dynamics analysis revealed that core fucosylation of SPARC-induced dynamic conformational changes in its N-glycan, allowing terminal galactose and N-acetylglucosamine to interact with K150, P261 and H264 residues, thereby promoting collagen binding. Site-specific mutagenesis of these residues also resulted in low affinity for collagen binding. Moreover, loss of collagen and decline of core fucosylation were observed in COPD lung tissues. These findings provide a new mechanistic insight into the role of core fucosylation of SPARC in cell-matrix communication and contribution to the abnormal alveolar structures in COPD.

Benzo[a]pyrene induces fibrotic changes and impairs differentiation in lung stem cells.

Human activities have generated air pollution, with extremely small particles (PM 2.5, particulate matter less than 2.5 µm in diameter) and liquid droplets, which become a menace to human health. Among the pollutants, polycyclic aromatic hydrocarbons (PAHs), which enhance the risks of pulmonary dysfunction and cancer development, have been extensively studied. Numerous studies have addressed the effects of PAHs on the respiratory system, whereas the effects on lung stem/progenitor cells remain unknown. Here, we provide evidence that benzo[a]pyrene (BaP), a major toxic PAH, induces fibrotic changes with a loss of α-1,6-fucosylation in CD54+CD157+CD45- cells (lung stem cells). In studies with aryl hydrocarbon receptor (AHR) antagonist, we found that these effects by BaP are independent of the canonical AHR pathway. In addition, these BaP-induced fibrotic changes are reduced by TGF-ß antagonist, suggesting an alternative pathway of BaP toxicity is different from other PAH/AHR signaling pathways. Finally, it was observed that BaP impairs the spheroid formation and the podoplanin expression of CD54+CD157+CD45- cells, indicating that BaP suppresses the differentiation of lung stem cells. Taken together, our findings reveal specific BaP-induced injuries in CD54+CD157+CD45- cells.

Negative modulation of the epigenetic regulator, UHRF1, by thyroid hormone receptors suppresses liver cancer cell growth.

The thyroid hormone, 3,3',5-triiodo-l-thyronine (T3 ), mediates several physiological processes, including embryonic development, cellular differentiation, metabolism and regulation of cell proliferation. Thyroid hormone (T3 ) and its receptor (TR) are involved in metabolism and growth. In addition to their developmental and metabolic functions, TRs play a tumor suppressor role, and therefore, their aberrant expression can lead to tumor transformation. Aberrant epigenetic silencing of tumor suppressor genes promotes cancer progression. The epigenetic regulator, Ubiquitin-like with PHD and ring finger domains 1 (UHRF1), is overexpressed in various cancers. In our study, we demonstrated that T3 negatively regulates UHRF1 expression, both in vitro and in vivo. Our results further indicate that UHRF1 regulation by T3 is indirect and mediated by Sp1. Sp1-binding elements of UHRF1 were identified at positions -664/-505 of the promoter region using the luciferase and chromatin immunoprecipitation assays. Notably, UHRF1 and Sp1 levels were elevated in subgroups of hepatocellular carcinoma patients and inversely correlated with TRα1 expression. Knockdown of UHRF1 expression should therefore provide a means to inhibit hepatoma cell proliferation. Expression of UHRF1 was downregulated by TRs, in turn, relieving silencing of the UHRF1 target gene, p21. Based on the collective findings, we propose that T3 /TR signaling induces hepatoma cell growth inhibition via UHRF1 repression.

Repression of microRNA-130b by thyroid hormone enhances cell motility.

BACKGROUND & AIMS: Thyroid hormone (T3) and its receptor (TR) are involved in cell growth and cancer progression. Although deregulation of microRNA (miRNA) expression has been detected in many tumor types, the mechanisms underlying functional impairment and specific involvement of miRNAs in tumor metastasis remain unclear. In the current study, we aimed to elucidate the involvement of deregulated miRNA-130b (miR-130b) and its target genes mediated by T3/TR in cancer progression. METHODS: Quantitative reverse transcription-PCR, luciferase and chromatin immunoprecipitation assays were performed to identify the miR-130b transcript and the mechanisms implicated in its regulation. The effects of miR-130b on hepatocellular carcinoma (HCC) invasion were further examined in vitro and in vivo. Clinical correlations among miR-130b, TRs and interferon regulatory factor 1 (IRF1) were examined in HCC samples using Spearman correlation analysis. RESULTS: Our experiments disclosed negative regulation of miR-130b expression by T3/TR. Overexpression of miR-130b led to marked inhibition of cell migration and invasion, which was mediated via suppression of IRF1. Cell migration ability was promoted by T3, but partially suppressed upon miR-130b overexpression. Furthermore, miR-130b suppressed expression of epithelial-mesenchymal transition (EMT)-related genes, matrix metalloproteinase-9, phosphorylated mammalian target of rapamycin (mTOR), p-ERK1/2, p-AKT and p-signal transducer and activator of transcription (STAT)-3. Notably, miR-130b was downregulated in hepatoma samples and its expression patterns were inversely correlated with those of TRα1 and IRF1. CONCLUSIONS: Our data collectively highlight a novel pathway interlinking T3/TR, miR-130b, IRF1, the EMT-related genes, p-mTOR, p-STAT3 and the p-AKT cascade, which regulates the motility and invasion of hepatoma cells.

Thyroid hormone enhanced human hepatoma cell motility involves brain-specific serine protease 4 activation via ERK signaling.

BACKGROUND: The thyroid hormone, 3, 3', 5-triiodo-L-thyronine (T3), has been shown to modulate cellular processes via interactions with thyroid hormone receptors (TRs), but the secretory proteins that are regulated to exert these effects remain to be characterized. Brain-specific serine protease 4 (BSSP4), a member of the human serine protease family, participates in extracellular matrix remodeling. However, the physiological role and underlying mechanism of T3-mediated regulation of BSSP4 in hepatocellular carcinogenesis are yet to be established. METHODS: The thyroid hormone response element was identified by reporter and chromatin immunoprecipitation assays. The cell motility was analyzed via transwell and SCID mice. The BSSP4 expression in clinical specimens was examined by Western blot and quantitative reverse transcription polymerase chain reaction. RESULTS: Upregulation of BSSP4 at mRNA and protein levels after T3 stimulation is a time- and dose-dependent manner in hepatoma cell lines. Additionally, the regulatory region of the BSSP4 promoter stimulated by T3 was identified at positions -609/-594. BSSP4 overexpression enhanced tumor cell migration and invasion, both in vitro and in vivo. Subsequently, BSSP4-induced migration occurs through the ERK 1/2-C/EBPß-VEGF cascade, similar to that observed in HepG2-TRα1 and J7-TRα1 cells. BSSP4 was overexpressed in clinical hepatocellular carcinoma (HCC) patients, compared with normal subjects, and positively associated with TRα1 and VEGF to a significant extent. Importantly, a mild association between BSSP4 expression and distant metastasis was observed. CONCLUSIONS: Our findings collectively support a potential role of T3 in cancer cell progression through regulation of the BSSP4 protease via the ERK 1/2-C/EBPß-VEGF cascade. BSSP4 may thus be effectively utilized as a novel marker and anti-cancer therapeutic target in HCC.

Stable isotope labeling with amino acids in cell culture (SILAC)-based quantitative proteomics study of a thyroid hormone-regulated secretome in human hepatoma cells.

The thyroid hormone, 3, 3',5-triiodo-l-thyronine (T(3)), regulates cell growth, development, differentiation, and metabolism via interactions with thyroid hormone receptors (TRs). However, the secreted proteins that are regulated by T(3) are yet to be characterized. In this study, we used the quantitative proteomic approach of stable isotope labeling with amino acids in cell culture coupled with nano-liquid chromatography-tandem MS performed on a LTQ-Orbitrap instrument to identify and characterize the T(3)-regulated proteins secreted in human hepatocellular carcinoma cell lines overexpressing TRα1 (HepG2-TRα1). In total, 1742 and 1714 proteins were identified and quantified, respectively, in three independent experiments. Among these, 61 up-regulated twofold and 11 down-regulated twofold proteins were identified. Eight proteins displaying increased expression and one with decreased expression in conditioned media were validated using Western blotting. Real-time quantitative RT-PCR further disclosed induction of plasminogen activator inhibitor-1 (PAI-1), a T(3) target, in a time-course and dose-dependent manner. Serial deletions of the PAI-1 promoter region and subsequent chromatin immunoprecipitation assays revealed that the thyroid hormone response element on the promoter is localized at positions -327/-312. PAI-1 overexpression enhanced tumor growth and migration in a manner similar to what was seen when T(3) induced PAI-1 expression in J7-TRα1 cells, both in vitro and in vivo. An in vitro neutralizing assay further supported a crucial role of secreted PAI-1 in T(3)/TR-regulated cell migration. To our knowledge, these results demonstrate for the first time that proteins involved in the urokinase plasminogen activator system, including PAI-1, uPAR, and BSSP4, are augmented in the extra- and intracellular space of T(3)-treated HepG2-TRα1 cells. The T(3)-regulated secretome generated in the current study may provide an opportunity to establish the mechanisms underlying T(3)-associated tumor progression and prognosis.

Emerging regulation and function of betatrophin.

Betatrophin, also known as TD26/RIFL/lipasin/ANGPTL8/C19orf80, is a novel protein predominantly expressed in human liver. To date, several betatrophin orthologs have been identified in mammals. Increasing evidence has revealed an association between betatrophin expression and serum lipid profiles, particularly in patients with obesity or diabetes. Stimulators of betatrophin, such as insulin, thyroid hormone, irisin and caloric intake, are usually relevant to energy expenditure or thermogenesis. In murine models, serum triglyceride levels as well as pancreatic cell proliferation are potently enhanced by betatrophin. Intriguingly, conflicting phenomena have also been reported that betatrophin suppresses hepatic triglyceride levels, suggesting that betatrophin function is mediated by complex regulatory processes. However, its precise physiological role remains unclear at present. In this review, we have summarized the current findings on betatrophin and their implications.

Effects of methylation and transcription factor YY1 on ID2 expression in non-small cell lung carcinoma cells.

The inhibitor of DNA-binding 2 (ID2) plays a major role in tumor dedifferentiation in non-small cell lung cancer (NSCLC). Studies have indicated an inverse correlation between ID2 expression and NSCLC cell invasiveness. However, the mechanisms through which ID2 activation is regulated are currently unclear. We overexpressed ID2 in H1299 cells and extensively characterized their cellular behaviors. By employing a serial deletion approach combined with a reporter assay, we pinpointed the basal promoter region of ID2. We also examined the DNA methylation status of the ID2 promoter to elucidate the epigenetic mechanisms driving ID2 regulation. Our results revealed that ID2 overexpression effectively inhibited the migration, invasion, proliferation, and colony formation abilities of H1299 cells. The region from -243 to +202 played a major role in driving the transcriptional activity of ID2. Sequence analysis results indicated that the transcription factor Yin Yang 1 (YY1) might be crucial in the regulation of ID2 expression. The ectopically expressed YY1 activated both the expression levels of ID2 and the transcriptional activity of the ID2 promoter, potentially contributing to its repressive activity on cancer cell growth. Furthermore, site-directed mutagenesis and chromatin immunoprecipitation assays revealed that YY1 may target the -120 and -76 sites of the ID2 promoter, thereby activating its transcriptional activity. The ID2 promoter regions were also fully methylated in CL1-5 cells, and the methylation level was correlated with the expression levels of the ID2 promoter. Moreover, the YY1-induced suppression of colony formation was counteracted by ID2 knockdown, which suggests that YY1 represses cell colony growth through the regulation of ID2. Our results indicate that YY1 plays a role in transactivating ID2 expression and might also contribute to the repression of colony growth through the regulation of ID2.

Thyroid hormone receptor inhibits hepatoma cell migration through transcriptional activation of Dickkopf 4.

Triiodothyronine (T3) is a potent form of thyroid hormone mediates several physiological processes including cellular growth, development, and differentiation via binding to the nuclear thyroid hormone receptor (TR). Recent studies have demonstrated critical roles of T3/TR in tumor progression. Moreover, long-term hypothyroidism appears to be associated with the incidence of human hepatocellular carcinoma (HCC), independent of other major HCC risk factors. Dickkopf (DKK) 4, a secreted protein that antagonizes the canonical Wnt signaling pathway, is induced by T3 at both mRNA and protein levels in HCC cell lines. However, the mechanism underlying T3-mediated regulation of DKK4 remains unknown. In the present study, the 5' promoter region of DKK4 was serially deleted, and the reporter assay performed to localize the T3 response element (TRE). Consequently, we identified an atypical direct repeat TRE between nucleotides -1645 and -1629 conferring T3 responsiveness to the DKK4 gene. This region was further validated using chromatin immunoprecipitation (ChIP) and electrophoretic mobility shift assay (EMSA). Stable DKK4 overexpression in SK-Hep-1 cells suppressed cell invasion and metastatic potential, both in vivo andin vitro, via reduction of matrix metalloproteinase-2 (MMP-2) expression. Our findings collectively suggest that DKK4 upregulated by T3/TR antagonizes the Wnt signal pathway to suppress tumor cell progression, thus providing new insights into the molecular mechanism underlying thyroid hormone activity in HCC.

Dickkopf 4 positively regulated by the thyroid hormone receptor suppresses cell invasion in human hepatoma cells.

UNLABELLED: Thyroid hormone (T(3)) mediates cellular growth, development, and differentiation by binding to the nuclear thyroid hormone receptor (TR). Recent studies suggest that long-term hypothyroidism is associated with human hepatocellular carcinoma (HCC) independent from other major HCC risk factors. Dickkopf (DKK) 4, a secreted protein, antagonizes the Wnt signal pathway. In this study, we demonstrate that T(3) may play a suppressor role by inducing DKK4 expression in HCC cells at both the messenger RNA (mRNA) and protein levels. DKK4 was down-regulated in 67.5% of HCC cancerous tissues. The decrease in DKK4 levels was accompanied by a concomitant decrease in TR protein levels in the matched cancerous tissues in 31% of tissues compared by immunoblotting with the adjacent noncancerous tissues. Further, TR and DKK4 expression levels were positively correlated in both normal and cancerous specimens by tissue array analysis. In function assays, stable DKK4 transfected into J7 or HepG2 cells decreased cell invasion in vitro. Conversely, knocking down DKK4 restores cell invasiveness. DKK4-expressing J7 clones showed increased degradation of ß-catenin, but down-regulation of CD44, cyclin D1, and c-Jun. To investigate the effect of DKK4 and TR on tumor growth in vivo, we established a xenograft of J7 cells in nude mice. J7-DKK4 and J7-TRα1 overexpressing mice, which displayed growth arrest, lower lung colony formation index, and smaller tumor size than in control mice, supporting an inhibitory role of DKK4 in tumor progression. CONCLUSION: Taken together, these data suggest that the TR/DKK4/Wnt/ß-catenin cascade influences the proliferation and migration of hepatoma cells during the metastasis process and support a tumor suppressor role of the TR.

Overexpression of ADP-ribosylation factor 1 in human gastric carcinoma and its clinicopathological significance.

Gastric cancer is the sixth leading cause of cancer-related death in Taiwan, and the identification of related factors is essential to increase patient survival. ADP-ribosylation factor 1 (ARF1) was initially identified using 2-D electrophoresis combined with MALDI-time-of-flight mass spectrometry. ADP-ribosylation factor 1 belongs to the Ras superfamily or GTP-binding protein family and has been shown to enhance cell proliferation. In the current study, we evaluated the potential of ARF1 as a biomarker for gastric cancer detection. ADP-ribosylation factor 1 mRNA was upregulated in tumor tissues (compared with adjacent non-tumor tissues, n = 55) in approximately 67.2% of gastric cancer patients. Expression of ARF1 protein was additionally observed using Western blot and immunohistochemistry (IHC) analyses. The clinicopathological correlations of ARF1 were further evaluated. Elevated ARF1 expression was strongly correlated with lymph node metastasis (P = 0.008), serosal invasion (P = 0.046), lymphatic invasion (P = 0.035), and pathological staging (P = 0.010). Moreover, the 5-year survival rate for the lower ARF1 expression group (n = 50; IHC score < 90) was higher than that of the higher expression group (n = 60; IHC score ≥ 90) (P = 0.0228, log-rank test). To establish the specific function of ARF1 in human gastric cancer, isogenic ARF1-overexpressing cell lines were prepared. Our results showed that ARF1-overexpressing clones display enhanced cell proliferation, migration, and invasion. Furthermore, ARF1-overexpression might contribute to poor prognosis of patients. These findings collectively support the utility of ARF1 as a novel prognostic marker for gastric cancer and its role in cell invasion.

Id2 exerts tumor suppressor properties in lung cancer through its effects on cancer cell invasion and migration.

Background: Despite advances in prognosis and treatment of lung adenocarcinoma (LADC), a notable non-small cell lung cancer subtype, patient outcomes are still unsatisfactory. New insight on novel therapeutic strategies for LADC may be gained from a more comprehensive understanding of cancer progression mechanisms. Such strategies could reduce the mortality and morbidity of patients with LADC. In our previous study, we performed cDNA microarray screening and found an inverse relationship between inhibitor of DNA binding 2 (Id2) expression levels and the invasiveness of LADC cells. Materials and Methods: To identify the functional roles of Id2 and its action mechanisms in LADC progression, we successfully established several Id2-overexpressing and Id2-silenced LADC cell clones. Subsequently, we examined in vitro the effects exerted by Id2 on cell morphology, proliferation, colony formation, invasive, and migratory activities and examined in vivo those exerted by Id2 on cell metastasis. The mechanisms underlying the action of Id2 were investigated using RNA-seq and pathway analyses. Furthermore, the correlations of Id2 with its target gene expression and clinical outcomes were calculated. Results: Our data revealed that Id2 overexpression could inhibit LADC cells' migratory, invasive, proliferation, and colony formation capabilities. Silencing Id2 expression in LADC cells reversed the aforementioned inhibitory effects, and knockdown of Id2 increased LADC cells' metastatic abilities in vivo. Bioinformatics analysis revealed that these effects of Id2 on cancer progression might be regulated by focal adhesion kinase (FAK) signaling and CD44/Twist expression. Furthermore, in online clinical database analysis, patients with LADC whose Id2 expression levels were high and FAK/Twist expression levels were low had superior clinical outcomes.Conclusion: Our data indicate that the Id2 gene may act as a metastasis suppressor and provide new insights into LADC progression and therapy.

Dysregulated FAM215A Stimulates LAMP2 Expression to Confer Drug-Resistant and Malignant in Human Liver Cancer.

Hepatocellular carcinoma (HCC) is one of the most common and aggressive human malignancies worldwide. Long non-coding (lnc) RNAs regulate complex cellular functions, such as cell growth, differentiation, metabolism, and metastasis. Although deregulation of lncRNA expression has been detected in HCC, many of the hepato-carcinogenesis-associated lncRNAs remain yet unidentified. Here, we aimed to investigate the involvement of a specific HCC-dysregulated lncRNA, FAM215A, and characterize its molecular regulation mechanism. We show for the first time that FAM215A is overexpressed in HCC, and its expression level correlates with tumor size, vascular invasion, and pathology stage. Overexpression of FAM215A accelerates cell proliferation and metastasis in HCC cells. According to Gene Expression Omnibus Dataset analysis, FAM215A is induced in doxorubicin (DOX)-resistant HCC cells. Overexpression of FAM215A increases DOX resistance in two HCC cell lines, and this is associated with enhanced expression of lysosome-associated membrane protein 2 (LAMP2). FAM215A interacts with LAMP2 to protect it from ubiquitination. Together, our results show that the lncRNA, FAM215A, is highly expressed in HCC, where it interacts with and stabilizes LAMP2 to increase tumor progression while decreasing doxorubicin sensitivity.

Thyroid hormone suppresses expression of stathmin and associated tumor growth in hepatocellular carcinoma.

Stathmin (STMN1), a recognized oncoprotein upregulated in various solid tumors, promotes microtubule disassembly and modulates tumor growth and migration activity. However, the mechanisms underlying the genetic regulation of STMN1 have yet to be elucidated. In the current study, we report that thyroid hormone receptor (THR) expression is negatively correlated with STMN1 expression in a subset of clinical hepatocellular carcinoma (HCC) specimens. We further identified the STMN1 gene as a target of thyroid hormone (T3) in the HepG2 hepatoma cell line. An analysis of STMN1 expression profile and mechanism of transcriptional regulation revealed that T3 significantly suppressed STMN1 mRNA and protein expression, and further showed that THR directly targeted the STMN1 upstream element to regulate STMN1 transcriptional activity. Specific knockdown of STMN1 suppressed cell proliferation and xenograft tumor growth in mice. In addition, T3 regulation of cell growth arrest and cell cycle distribution were attenuated by overexpression of STMN1. Our results suggest that the oncogene STMN1 is transcriptionally downregulated by T3 in the liver. This T3-mediated suppression of STMN1 supports the theory that T3 plays an inhibitory role in HCC tumor growth, and suggests that the lack of normal THR function leads to elevated STMN1 expression and malignant growth.

A novel small-form NEDD4 regulates cell invasiveness and apoptosis to promote tumor metastasis.

Despite numerous investigations on metastasis, the determinants of metastatic processes remain unclear. We aimed to identify the metastasis-associated genes in hepatocellular carcinoma (HCC). Potent metastatic SK-hep-1 (SK) cells, designated 'SKM', were generated using Transwell assay followed by selection in a mouse model. Genes expressed differentially in SKM and SK cells were identified via microarray analyses. A small form of Neural precursor cell-expressed developmentally downregulated 4 (sNEDD4) was identified to be overexpressed in SKM cells, which was confirmed as a novel transcript using liquid chromatography-mass spectrometry. In clinical specimens, sNEDD4 was significantly overexpressed in tumors and serves as a poor prognostic factor for male patients with HCC (P = 0.035). Upon subcutaneous introduction of sNEDD4-overexpressing SK cells into flanks of nude mice, tumors grew faster than those of the control group. Furthermore, sNEDD4-mediated promotion of tumor metastasis was demonstrated in the orthotopic mouse model. Overexpression of sNEDD4 increased the invasive ability of SK cells through upregulation of matrix metalloproteinase 9 and inhibited serum deprivation-induced apoptosis via upregulation of myeloid cell leukemia 1. In conclusion, sNEDD4 is a novel metastasis-associated gene, which prevents apoptosis under nutrient restriction conditions. The present findings clearly support the prognostic potential of sNEDD4 for HCC.

Potential prognostic, diagnostic and therapeutic markers for human gastric cancer.

The high incidence of gastric cancer (GC) and its consequent mortality rate severely threaten human health. GC is frequently not diagnosed until a relatively advanced stage. Surgery is the only potentially curative treatment. Thus, early screening and diagnosis are critical for improving prognoses in patients with GC. Gastroscopy with biopsy is an appropriate method capable of aiding the diagnosis of specific early GC tumor types; however, the stress caused by this method together with it being excessively expensive makes it difficult to use it as a routine method for screening for GC on a population basis. The currently used tumor marker assays for detecting GC are simple and rapid, but their use is limited by their low sensitivity and specificity. In recent years, several markers have been identified and tested for their clinical relevance in the management of GC. Here, we review the serum-based tumor markers for GC and their clinical significance, focusing on discoveries from microarray/proteomics research. We also review tissue-based GC tumor markers and their clinical application, focusing on discoveries from immunohistochemical research. This review provides a brief description of various tumor markers for the purposes of diagnosis, prognosis and therapeutics, and we include markers already in clinical practice and various forthcoming biomarkers.

Chromosome 19 open reading frame 80 is upregulated by thyroid hormone and modulates autophagy and lipid metabolism.

The thyroid hormone, T 3, regulates cell growth, differentiation and development through binding to the nuclear thyroid hormone receptor (THR), a member of the steroid/TR superfamily of ligand-dependent transcriptional factors. T 3 modulates lipid metabolism in liver, although the detailed molecular mechanisms are unclear at present. Here, by a microarray analysis, we identified a novel chromosome 19 open reading frame 80 (C19orf80) which was activated by T 3. T 3 stimulation led to upregulation of both mRNA and protein levels of C19orf80. Immunofluorescence analysis revealed a vesicle-like pattern of C19orf80 around lipid droplets or within the lysosome-associated compartment in cells. Furthermore, T 3 treatment as well as C19orf80 overexpression specifically activated the autophagic response and lipid metabolism, as observed from lipidated LC3 (LC3-II) and levels of oxygen consumption rate, respectively. Reciprocally, knockdown of C19orf80 obstructed T 3-activated autophagy and lipolysis. Moreover, treatment with autolysosome maturation inhibitors, ammonium chloride and chloroquine, not only suppressed the T 3-activated autophagic process but also lipid metabolism. Our results collectively suggested that T 3 regulates lipid metabolism through a C19orf80-activated autophagic process.

MicroRNA-196a/-196b promote cell metastasis via negative regulation of radixin in human gastric cancer.

MicroRNAs (miRNAs) play an important role to contribute carcinogenesis. The aim of the current study was to identify useful biomarkers from miRNAs. Differential miRNA profiles were analyzed using the miRNA qRT-PCR-based assay. Two of the most upregulated miRNAs were selected and validated. The miR-196a/-196b levels were significantly increased in gastric cancer (GC) tissues (n=109). Overexpression of miR-196a/-196b was significantly associated with tumor progression and poorer 5-year survival outcomes. Overexpression of miR-196a/-196b enhances GC cell migration and invasion. Further, radixin was identified as a target gene of miR-196a/-196b. Elevated miR-196a/-196b expression in GC cells led to reduced radixin protein levels and vice versa. Notably, an inverse correlation between miR-196a/-196b and radixin mRNA and protein expression was observed in GC tissues with in situ hybridization and immunohistochemistry analyses. Together, miR-196a/-196b inhibitory oligonucleotides or overexpression of the radixin may thus have therapeutic potential in suppressing GC metastasis.

Interleukin-32 increases human gastric cancer cell invasion associated with tumor progression and metastasis.

PURPOSE: The proinflammatory cytokine interleukin-32 (IL-32) is a novel tumor marker highly expressed in various human carcinomas, including gastric cancer. However, its effects on prognosis of patients with gastric cancer and cancer metastasis are virtually unknown at present. The main aim of this study was to explore the clinical significance of IL-32 in gastric cancer and further elucidate the molecular mechanisms underlying IL-32-mediated migration and invasion. EXPERIMENTAL DESIGN: Gastric cancer cells with ectopic expression or silencing of IL-32 were examined to identify downstream molecules and establish their effects on cell motility, invasion, and lung metastasis in vivo. RESULTS: IL-32 was significantly upregulated in gastric cancer and positively correlated with aggressiveness of cancer and poor prognosis. Ectopic expression of IL-32 induced elongated morphology and increased cell migration and invasion via induction of IL-8, VEGF, matrix metalloproteinase 2 (MMP2), and MMP9 expression via phosphor-AKT/phospho-glycogen synthase kinase 3ß/active ß-catenin as well as hypoxia-inducible factor 1α (HIF-1α) signaling pathways. Conversely, depletion of IL-32 in gastric cancer cells reversed these effects and decreased lung colonization in vivo. Examination of gene expression datasets in oncomine and staining of gastric cancer specimens demonstrated the clinical significance of IL-32 and its downstream molecules by providing information on their coexpression patterns. CONCLUSIONS: IL-32 contributes to gastric cancer progression by increasing the metastatic potential resulting from AKT, ß-catenin, and HIF-1α activation. Our results clearly suggest that IL-32 is an important mediator for gastric cancer metastasis and independent prognostic predictor of gastric cancer.