α-Hederin inhibits the platelet activating factor-induced metastasis of HCC cells through disruption of PAF/PTAFR axis cascaded STAT3/MMP-2 expression.

Metastasis remains a crucial obstacle to the clinical treatment of hepatocellular carcinoma (HCC). Investigating the potential anti-tumor compounds from medicinal herb against HCC metastasis is of particular interest. As a triterpenoid saponin, α-Hederin has been reported to exhibit cytotoxicity for diverse cancer cell lines by inducing mitochondrial related apoptosis or autophagic cell death. Nevertheless, little is known about the inhibitory effect of α-Hederin on the metastasis of HCC and its underlying mechanisms. Here, we integrated well-established target prediction webtool and molecular docking methods to predict the potential targets for α-Hederin, and finally focused on PTAFR, the receptor for platelet-activating factor (PAF). Activation of PAF/PTAFR pathways has been reported to be contribution to the initiation and progression of cancer. We showed for the first time that non-cytotoxic concentration of α-Hederin inhibited cell migration and invasion induced by PAF in HCC cells, as well as lung metastasis in vivo. Moreover, we demonstrated α-Hederin reduced the PAF-induced matrix metalloproteinase-2 expression through inhibiting the activation of STAT3 in PAF stimulated HCC cells. These findings suggest that α-Hederin functions as a prospective inhibitor of PTAFR and may be utilized as an optional candidate for treatment of HCC.

O-GlcNAcylation of RACK1 promotes hepatocellular carcinogenesis.

BACKGROUND & AIMS: Aberrant oncogenic mRNA translation and protein O-linked ß-N-acetylglucosaminylation (O-GlcNAcylation) are general features during tumorigenesis. Nevertheless, whether and how these two pathways are interlinked remain unknown. Our previous study indicated that ribosomal receptor for activated C-kinase 1 (RACK1) promoted chemoresistance and growth in hepatocellular carcinoma (HCC). The aim of this study is to examine the role of RACK1 O-GlcNAcylation in oncogene translation and HCC carcinogenesis. METHODS: The site(s) of RACK1 for O-GlcNAcylation was mapped by mass spectrometry analysis. HCC cell lines were employed to examine the effects of RACK1 O-GlcNAcylation on the translation of oncogenic factors and behaviors of tumor cells in vitro. Transgenic knock-in mice were used to detect the role of RACK1 O-GlcNAcylation in modulating HCC tumorigenesis in vivo. The correlation of RACK1 O-GlcNAcylation with tumor progression and relapse were analyzed in clinical HCC samples. RESULTS: We found that ribosomal RACK1 was highly modified by O-GlcNAc at Ser122. O-GlcNAcylation of RACK1 enhanced its protein stability, ribosome binding and interaction with PKCßII (PRKCB), leading to increased eukaryotic translation initiation factor 4E phosphorylation and translation of potent oncogenes in HCC cells. Genetic ablation of RACK1 O-GlcNAcylation at Ser122 dramatically suppressed tumorigenesis, angiogenesis, and metastasis in vitro and in diethylnitrosamine (DEN)-induced HCC mouse model. Increased RACK1 O-GlcNAcylation was also observed in HCC patient samples and correlated with tumor development and recurrence after chemotherapy. CONCLUSIONS: These findings demonstrate that RACK1 acts as key mediator linking O-GlcNAc metabolism to cap-dependent translation during HCC tumorigenesis. Targeting RACK1 O-GlcNAcylation provides promising options for HCC treatment. LAY SUMMARY: O-GlcNAcylation of ribosomal receptor for activated C-kinase 1 at the amino acid serine122 promotes its stability, ribosome localization and interaction with the protein kinase, PKCßII, thus driving the translation of oncogenes and tumorigenesis of hepatocellular carcinoma. Increased O-GlcNAcylation of ribosomal receptor for activated C-kinase 1 is positively correlated with tumor growth, metastasis and recurrence in patients with hepatocellular carcinoma.

RACK1-mediated translation control promotes liver fibrogenesis.

Activation of quiescent hepatic stellate cells (HSCs) is the central event of liver fibrosis. The translational machinery is an optimized molecular network that affects cellular homoeostasis and diseases, whereas the role of protein translation in HSCs activation and liver fibrosis is little defined. Our previous report suggests that up-regulation of receptor for activated C-kinase 1(RACK1) in HSCs is critical for liver fibrogenesis. In this study, we found that RACK1 promoted macrophage conditioned medium (MCM)-induced assembly of eIF4F and phosphorylation of eIF4E in primary HSCs. RACK1 enhanced the translation and expression of pro-fibrogenic factors collagen 1α1, snail and cyclin E1 induced by MCM. Administration of PP242 or knock-down of eIF4E suppressed RACK1-stimulated collagen 1α1 production, proliferation and migration in primary HSCs. In addition, depletion of eIF4E attenuated thioacetamide (TAA)-induced liver fibrosis in vivo. Our data suggest that RACK1-mediated stimulation of cap-dependent translation plays crucial roles in HSCs activation and liver fibrogenesis, and targeting translation initiation could be a promising strategy for the treatment of liver fibrosis.

PKCα promotes generation of reactive oxygen species via DUOX2 in hepatocellular carcinoma.

Hepatocellular carcinoma (HCC) remains the second leading cause of cancer-related death worldwide, and elevated rates of reactive oxygen species (ROS) have long been considered as a hallmark of almost all types of cancer including HCC. Protein kinase C alpha (PKCα), a serine/threonine kinase among conventional PKC family, is recognized as a major player in signal transduction and tumor progression. Overexpression of PKCα is commonly observed in human HCC and associated with its poor prognosis. However, how PKCα is involved in hepatocellular carcinogenesis remains not fully understood. In this study, we found that among the members of conventional PKC family, PKCα, but not PKCßI or ßII, promoted ROS production in HCC cells. PKCα stimulated generation of ROS by up-regulating DUOX2 at post-transcriptional level. Depletion of DUOX2 abrogated PKCα-induced activation of AKT/MAPK pathways as well as cell proliferation, migration and invasion in HCC cells. Moreover, the expression of DUOX2 and PKCα was well positively correlated in both HCC cell lines and patient samples. Collectively, our findings demonstrate that PKCα plays a critical role in HCC development by inducing DUOX2 expression and ROS generation, and propose a strategy to target PKCα/DUOX2 as a potential adjuvant therapy for HCC treatment.

Overexpression of Ras-GTPase-activating protein SH3 domain-binding protein 1 correlates with poor prognosis in gastric cancer patients.

AIMS: Ras-GTPase-activating protein SH3 domain-binding protein 1 (G3BP1) is a downstream effector of Ras signalling, and is overexpressed in several types of human malignancy. However, its role in gastric cancer remains unclear. The aim of this study was to investigate the prognostic significance of G3BP1 in gastric cancer. METHODS AND RESULTS: G3BP1 mRNA and protein levels in paired frozen tumour samples were detected by real-time polymerase chain reaction and western blotting, respectively. Paraffin-embedded tumour samples were used for immunohistochemistry. Gastric cancer cells were used to detect the tumorigenic role of G3BP1 in vitro. We found that G3BP1 protein expression was markedly increased in gastric cancer tissues as compared with corresponding non-malignant mucosa, whereas corresponding changes in mRNA levels were not observed. G3BP1 staining was positively correlated with tumour size, vascular invasion, T classification, lymph node metastasis, TNM stage, and reduced overall survival. Further analysis identified G3BP1 as an independent prognostic factor for poor prognosis, and combining G3BP1 with TNM stage generated a better predictive model for patient outcomes. G3BP1 also promoted proliferation, migration/invasion and extracellular signal-related kinase and AKT activation in gastric cancer cells. CONCLUSIONS: Our data define G3BP1 as a novel independent prognostic factor that is correlated with gastric cancer progression.

Activation of farnesoid X receptor attenuates hepatic injury in a murine model of alcoholic liver disease.

Alcoholic liver disease (ALD) is a common cause of advanced liver disease, and considered as a major risk factor of morbidity and mortality worldwide. Hepatic cholestasis is a pathophysiological feature observed in all stages of ALD. The farnesoid X receptor (FXR) is a member of the nuclear hormone receptor superfamily, and plays an essential role in the regulation of bile acid, lipid and glucose homeostasis. However, the role of FXR in the pathogenesis and progression of ALD remains largely unknown. Mice were fed Lieber-DeCarli ethanol diet or an isocaloric control diet. We used a specific agonist of FXR WAY-362450 to study the effect of pharmacological activation of FXR in alcoholic liver disease. In this study, we demonstrated that FXR activity was impaired by chronic ethanol ingestion in a murine model of ALD. Activation of FXR by specific agonist WAY-362450 protected mice from the development of ALD. We also found that WAY-362450 treatment rescued FXR activity, suppressed ethanol-induced Cyp2e1 up-regulation and attenuated oxidative stress in liver. Our results highlight a key role of FXR in the modulation of ALD development, and propose specific FXR agonists for the treatment of ALD patients.

FIB-SEM combined with proteomics and modification omics clarified mechanisms of lipids synthesis in organelles of Chlorella pyrenoidosa cells with high CO2 concentration.

In order to explain reasons why flue-gas CO2 (normally containing high CO2) enhanced carbon fixation and lipids synthesis with increased photochemical electron production in microalgae cells. Focused ion beam scanning electron microscopy (FIB-SEM) was combined with proteomics and phosphorylation modification mics to clarify mechanisms of lipids synthesis at protein and organelle levels in Chlorella pyrenoidosa cells cultivated with high CO2 concentration (15 % v/v). The volumes of chloroplast and endoplasmic reticulum in subcellular organelles increased by 47 % and 306 %, respectively, compared with the control, which improved conversion efficiency of starch grains to lipids (lipid content increased by 57 %). Proteomics and modifications omics revealed that protein translation and ribosome structure and biogenesis-related enzymes were significantly modified by phosphorylation, which regulated protein biological functions. Glycolysis, pentose phosphate pathway and other carbohydrate metabolic pathways were markedly enriched and promoted the expression of lipid synthase, which was consistent with enhanced carbon fixation in photosynthesis, expansion of subcellular organelles and improved lipids synthesis.

Astragalus Polysaccharide Promotes Doxorubicin-Induced Apoptosis by Reducing O-GlcNAcylation in Hepatocellular Carcinoma.

The toxicity and side effects of chemotherapeutic drugs remain a crucial obstacle to the clinical treatment of hepatocellular carcinoma (HCC). Identifying combination therapy from Chinese herbs to enhance the sensitivity of tumors to chemotherapeutic drugs is of particular interest. Astragalus polysaccharide (APS), one of the natural active components in Astragalus membranaceus, has been reported to exhibit anti-tumor properties in diverse cancer cell lines. The aim of this study was to determine the effect of APS on Doxorubicin (Dox)-induced apoptosis in HCC and the underlying mechanism. The results showed that APS dose-dependently promoted Dox-induced apoptosis and enhanced endoplasmic reticulum (ER) stress. Additionally, APS decreased the mRNA level and protein stability of O-GlcNAc transferase (OGT), and increased the O-GlcNAcase (OGA) expression. Furthermore, OGT lentiviral transfection or PugNAc (OGA inhibitor) treatment reversed the ER stress and apoptosis induced by the combination of Dox and APS. A xenograft tumor mouse model confirmed that the combination of APS and Dox showed an advantage in inhibiting tumor growth in vivo. These findings suggested that APS promoted Dox-induced apoptosis in HCC cells through reducing the O-GlcNAcylation, which led to the exacerbation of ER stress and activation of apoptotic pathways.

Functional metabolism pathways of significantly regulated genes in Nannochloropsis oceanica with various nitrogen/phosphorus nutrients for CO2 fixation.

To determine the optimal CO2 concentration for microalgal biomass cultivated with industrial flue gas and improve carbon fixation capacity and biomass production. Functional metabolism pathways of significantly regulated genes in Nannochloropsis oceanica (N. oceanica) with various nitrogen/phosphorus (N/P) nutrients for CO2 fixation were comprehensively clarified. At 100 % N/P nutrients, the optimum CO2 concentration was 70 % and the maximum biomass production of microalgae was 1.57 g/L. The optimum CO2 concentration was 50 % for N or P deficiency and 30 % for both N and P deficiency. The optimal combination of CO2 concentration and N/P nutrients caused significant up regulation of proteins related to photosynthesis and cellular respiration in the microalgae, enhancing photosynthetic electron transfer efficiency and carbon metabolism. Microalgal cells with P deficiency and optimal CO2 concentration expressed many phosphate transporter proteins to enhance P metabolism and N metabolism to maintain a high carbon fixation capacity. However, inappropriate combination of N/P nutrients and CO2 concentrations caused more errors in DNA replication and protein synthesis, generating more lysosomes and phagosomes. This inhibited carbon fixation and biomass production in the microalgae with increased cell apoptosis.

Sodium arsenite down-regulates the expression of X-linked inhibitor of apoptosis protein via translational and post-translational mechanisms in hepatocellular carcinoma.

X-linked inhibitor of apoptosis protein (XIAP) is a member of the inhibitors of apoptosis protein (IAP) family, and has been reported to exhibit elevated expression levels in hepatocellular carcinoma (HCC) and promote cell survival, metastasis and tumor recurrence. Targeting XIAP has proven effective for the inhibition of cancer cell proliferation and restoration of cancer cell chemosensitivity. Arsenic (or sodium arsenite) is a potent anti-tumor agent used to treat patients with acute promyelocytic leukemia (APL). Additionally, arsenic induces cell growth inhibition, cell cycle arrest and apoptosis in human HCC cells. In this study, we identified XIAP as a target for sodium arsenite-induced cytotoxicity in HCC. The exposure of HCC cell lines to sodium arsenite resulted in inhibition of XIAP expression in both a dose- and time-dependent manner. Sodium arsenite blocked the de novo XIAP synthesis and the activity of its internal ribosome entry site (IRES) element. Moreover, treatment with sodium arsenite decreased the protein stability of XIAP and induced its ubiquitin-proteasomal degradation. Overexpression of XIAP attenuated the pro-apoptotic effect of sodium arsenite in HCC. Taken together, our data demonstrate that sodium arsenite suppresses XIAP expression via translational and post-translational mechanisms in HCC.

Combined anti-tumor effects of IFN-α and sorafenib on hepatocellular carcinoma in vitro and in vivo.

Hepatocellular carcinoma (HCC) is among the most common and aggressive cancers worldwide, and novel therapeutic strategies are urgently required to improve clinical outcome. Interferon-alpha (IFN-α) and sorafenib are widely used as anti-tumor agents against various malignancies. In this study, we investigated the combined effects of IFN-α and sorafenib against HCC. We demonstrated that the combination therapy synergistically suppressed HCC cellular viability, arrested cell cycle propagation and induced apoptosis in HCC cells. Further research revealed that IFN-α and sorafenib collaboratively regulated the expression levels of cell cycle-related proteins Cyclin A and Cyclin B as well as the pro-survival Bcl-2 family proteins Mcl-1, Bcl-2 and Bcl-X(L). Moreover, sorafenib inhibited IFN-α induced oncogenic signaling of STAT3, AKT and ERK but not the activation of the tumor suppressor STAT1. Xenograft experiments also confirmed the combined effects of IFN-α and sorafenib on tumor growth inhibition and apoptosis induction in vivo. In conclusion, these results provide rationale for the clinical application of IFN-α and sorafenib combination therapy in HCC treatment.

Fibroblast growth factor 7 inhibits cholesterol 7α-hydroxylase gene expression in hepatocytes.

Cholesterol 7α-hydroxylase (CYP7A1) is the initial and rate-limiting enzyme for bile acid synthesis. Transcription of the CYP7A1 gene is regulated by bile acids, nuclear receptors and cytokines. Fibroblast growth factor 7 (FGF7) secreted from activated hepatic stellate cells (HSC) during chronic liver fibrosis regulates hepatocyte survival and liver regeneration. In the carbon tetrachloride (CCl(4))-induced fibrotic mouse liver, we demonstrated that the expression of CYP7A1 was largely decreased while the expression of FGF7 was significantly increased. We further demonstrated that FGF7 inhibited CYP7A1 gene expression in hepatocytes. Knockdown study by short interfering RNA, kinase inhibition and phosphorylation assays revealed that the suppression of CYP7A1 expression by FGF7 was mediated by FGFR2 and its downstream JNK signaling cascade. The FGF7 neutralizing antibody restored CYP7A1 expression in Hep3B cells treated with conditioned medium from HSC. In summary, the data suggest that FGF7 is a novel regulator of CYP7A1 expression in hepatocytes and may prevent hepatocytes from accumulating toxic bile acids during liver injury and fibrosis.

[The analysis of the influencing factors of mortality rate in the transplantation of mouse bone marrow].

The purpose of the study was to investigate the influencing factors of mortality rate in the bone marrow transplantation in mice. The recipient mice receiving whole-body irradiation of gamma-ray were infused with the same strain of bone marrow cells or the mixture of the bone marrow cells and splenocytes respectively. Experiments were carried out in four batches, with different strains of mice used, respectively. The manifestations and the appearance of graft-versus-host disease (GVHD) were observed, as well as the mortality rate within 35 d of the transplantation in the recipient mice. The mortality rate of the first group of recipient mice was the lowest, the mortality rate of the second group of recipient mice was the highest and the obvious GVHD performance was observed before death. In the third group, the mortality rate declined and there was statistical significance compared to that of the second group. The mortality rate of the fourth group of mice was higher than that of the third group, but still lower than that of the second group of mice and there is a statistical significance. This evidence suggested that mouse genetic purity, splenocytes, the ratio of the bone marrow cells and splenocytes and the week-old of the mouse could be the important influencing factors of the mortality rate in mouse bone marrow transplantation.

Enhancing microalgae production by installing concave walls in plate photobioreactors.

In order to optimize light distribution for promoting biomass growth rate of Chlorella pyrenoidosa, concave walls were installed in plate photobioreactors (PBR) to generate rotational flow field of microalgal solution circulated from top inlets to bottom outlets. Flow vortices in four corners of concave-wall PBR resulted in decreased mixing time and increased mass transfer coefficient. The CO2 bio-fixation by C. pyrenoidosa increased by 27% and chlorophyll-a concentration enhanced by 18.5% in concave-wall PBR compared to those in control (flat-wall) PBR. The concave walls diverge light rays to enhance frontal light exposure and supply more light photons into interior regions of PBRs. The promotion in light distribution and vortex flow field with concave walls enhanced light and nutrients utilization by microalgal cells, leading to an increased biomass growth rate by 21%.

Akebia saponin E, as a novel PIKfyve inhibitor, induces lysosome-associated cytoplasmic vacuolation to inhibit proliferation of hepatocellular carcinoma cells.

ETHNOPHARMACOLOGICAL RELEVANCE: Hepatocellular carcinoma (HCC) is an aggressive malignancy with increasing mortality in China. Screening and identifying effective anticancer compounds from active traditional Chinese herbs for HCC are in demand. Akebia trifoliata (Thunb) Koidz, with pharmacological anti-HCC activities in clinical, has been shown in previous research. In the present research, we elucidated a potential anticancer effect of Akebia saponin E (ASE), which is isolated from the immature seeds of Akebia trifoliata (Thunb.) Koidz, and revealed that ASE could induce severe expanded vacuoles in HCC cells. But the potential mechanism of vacuole-formation and the anti-HCC effects by ASE remain uncover. AIM OF THIS STUDY: To elucidate the potential mechanism of vacuole-formation and the proliferation inhibition effects by ASE in HCC cell lines. MATERIALS AND METHODS: MTT assay, colony formation assay and flow cytometry were performed to detect cell viability. Immunofluorescence analysis was used to examine the biomarkers of endomembrane. Cells were infected with tandem mRFP-GFP-LC3 lentivirus to assess autophagy flux. RNA-seq was conducted to analyze the genome-wide transcriptional between treatment cell groups. In vitro PIKfyve kinase assay is detected by the ADP-GloTM Kinase Assay Kit. RESULTS: ASE could inhibit the proliferation of HCC with severe expanded vacuoles in vitro, and could significantly reduce the size and weight of xenograft tumor in vivo. Further, the vacuoles induced by ASE were aberrant enlarged lysosomes instead of autophagosome or autolysosomes. With cytoplasmic vacuolation, ASE induced a mTOR-independent TFEB activation for lysosomal biogenesis and a decrement of cholesterol levels in HCC cells. Furthermore, ASE could reduce the activity of PIKfyve (phosphoinositide kinase containing a FYVE-type finger), causing aberrant lysosomal biogenesis and cholesterol dyshomeostasis which triggered the expanded vacuole formation. CONCLUSION: ASE can prospectively inhibit the kinase activity of PIKfyve to induce lysosome-associated cytoplasmic vacuolation, and may be utilized as an alternative candidate to treat human HCC.

Akebia trifoliata (Thunb.) Koidz Seed Extract inhibits human hepatocellular carcinoma cell migration and invasion in vitro.

ETHNOPHARMACOLOGICAL RELEVANCE: The high recurrence rate postoperative and extensive metastases have become the obstacle of Hepatocellular Carcinoma (HCC) efficacy improvements, which contribute to most of the patient mortality. Akebia trifoliata (Thunb.) Koidz has been shown pharmacological activities in clinical and anti-HCC biological activity in previous research, but its potential function of anti-metastasis remains unknown. AIM OF THIS STUDY: To make sure whether ATKSE inhibits migration and invasion in HCC cell lines in vitro and the potential mechanism. MATERIALS AND METHODS: A UHPLC-HRMS analysis was adopted to identify and control the quality of the ethanol extract of Akebia trifoliata (Thunb.) Koidz Seed (abbreviated ATKSE). Cell viability of three kinds of HCC cell lines (HEPG2, HUH7, and SMMC7721) was detected using MTT assay and Flow cytometry. Adhesion capacity was measured by cell-matrigel adhesion assay. Wounded healing and Matrigel-transwell invasion assays were performed to assess cell migration and invasion, respectively. Western blot assay was used to detect several metastasis-related protein molecules, including FAK adhesion signaling, cadherin molecules, and MMPs. ELISA assay was used to evaluate the secreted MMP9 level. RESULTS: ATKSE significantly suppressed HCC cells viability and proliferation (from 0.9 up to 3.0 mg/ml); then under sub-lethal concentration (from 0.25 up to 1.0 mg/ml), ATKSE inhibited cell adhesion, migration, and invasion in a way of dose-dependent. Several metastatic-related molecules or pathway, including FAK adhesion signaling, cadherin molecules, and MMPs, took part in this process. There are both differences and commonalities in various cell lines: typically such as p-FAK was down-regulated by ATKSE in both HEPG2 and SMMC7721, while was raised in HUH7; Further attempts on the combination of ATKSE and FAK inhibitors, provide us with the enhanced inhibitory effects of invasion and migration in HEPG2 and HUH7 cells, as well as antagonistic effects in SMMC7721. As a target or potential mechanism, it may be more valuable to concern FAK inhibition by ATKSE in HEPG2 cells than in the other two cells. CONCLUSIONS: These results suggest that ATKSE has anti-metastasis potency in HCC cells.

High expression of GFAT1 predicts unfavorable prognosis in patients with hepatocellular carcinoma.

Hepatocellular carcinoma (HCC) is the second leading cause of cancer-related deaths worldwide. As a branch of glucose metabolism, hexosamine biosynthesis pathway (HBP) has been reported to play a critical role in the insulin resistance and progression of cancer. Glutamine:fructose-6-phosphate amidotransferase (GFAT) is the rate-limiting enzyme of the HBP; nevertheless, the prognostic value of GFAT1 in HCC remains elusive. In this study, we found that high expression of GFAT1 was significantly associated with serum alpha-fetoprotein (AFP), serum alanine aminotransferase (ALT), tumor size, tumor encapsulation, T stage and TNM stage. High GFAT1 expression was identified as an independent prognostic factor which predicted poor overall survival (OS) and recurrence-free survival (RFS) in HCC patients. Incorporation of GFAT1 expression could improve the prognostic accuracy of traditional TNM stage system. Integration of GFAT1 expression with other independent prognosticators generated a predictive nomogram, which showed better prognostic efficiency for OS and RFS in HCC patients. In vitro studies also revealed that GFAT1 promoted the proliferation, cell cycle progression, migration and invasion of HCC cells. In conclusion, GFAT1 is a potential prognostic biomarker for overall survival and recurrence-free survival of HCC patients after surgery.

Loss of GFAT1 promotes epithelial-to-mesenchymal transition and predicts unfavorable prognosis in gastric cancer.

Gastric cancer remains the third leading cause of cancer-related mortality worldwide, and invasion and metastasis of gastric cancer represent the major reason for its poor prognosis. Glutamine: fructose-6-phosphate amidotransferase 1 (GFAT1) is the first and rate-limiting enzyme of hexosamine biosynthesis pathway (HBP). Nevertheless, the role of GFAT1 in gastric cancer is little investigated. In this study, we found that the expression of GFAT1 was decreased in gastric cancer. Low expression of GFAT1 was positively associated with vessel invasion, late T stage, lymph node metastasis, distant metastasis, advanced TNM stage and poor prognosis in patients with gastric cancer. Furthermore, in vitro and in vivo studies revealed that down-regulation of GFAT1 promoted epithelial-to-mesenchymal transition (EMT) and invasive activities in gastric cancer cells through inducing the expression of TGF-ß1. The GFAT1 expression also significantly correlated with EMT-related factors in gastric cancer patients. Together, these findings indicate that GFAT1 functions as a novel suppressor of EMT and tumor metastasis in gastric cancer.

E3 ubiquitin ligase CHIP interacts with C-type lectin-like receptor CLEC-2 and promotes its ubiquitin-proteasome degradation.

C-type lectin-like receptor 2 (CLEC-2) was originally identified as a member of non-classical C-type lectin-like receptors in platelets and immune cells. Activation of CLEC-2 is involved in thrombus formation, lymphatic/blood vessel separation, platelet-mediated tumor metastasis and immune response. Nevertheless, the regulation of CLEC-2 expression is little understood. In this study, we identified that the C terminus of Hsc70-interacting protein (CHIP) interacted with CLEC-2 by mass spectrometry analysis, and CHIP decreased the protein expression of CLEC-2 through lysine-48-linked ubiquitination and proteasomal degradation. Deleted and point mutation also revealed that CHIP controlled CLEC-2 protein expression via both tetratricopeptide repeats (TPR) domain and Ubox domain in a HSP70/90-independent manner. Moreover, reduced CHIP expression was associated with decreased CLEC-2 polyubiquitination and increased CLEC-2 protein levels in PMA-induced differentiation of THP-1 monocytes into macrophages. These results indicate that CLEC-2 is the target substrate of E3 ubiquitin ligase CHIP, and suggest that the CHIP/CLEC-2 axis may play an important role in the modulation of immune response.

Decreased expression of Calpain-9 predicts unfavorable prognosis in patients with gastric cancer.

Calpain-8 and calpain-9 belong to the family of calcium-dependent cysteine proteases, which are highly expressed in the stomach. However, the roles of calpain-8 and calpain-9 in gastric tumorigenesis remain little understood. Herein, we demonstrated that calpain-9 was generally decreased in gastric cancer cell lines and primary tumor tissues, while calpain-8 expression was not significantly altered. Calpain-9, but not calpain-8, induced cell cycle arrest in the G1 phase and cellular apoptosis in vitro, and it attenuated the growth of subcutaneous tumor xenografts in vivo. Low expression of calpain-9 was positively associated with male sex, late T stage, lymph node metastasis, and advanced TNM stage. Further analysis identified calpain-9 as an independent prognostic factor for poor prognosis, and combining calpain-9 with TNM stage generated a better predictive model for patient outcomes. In conclusion, calpain-9 is a tumor suppressor that can be regarded as a potential prognosis indicator for clinical outcomes in gastric cancer.