Exo-miR-1290-induced by COX-2 overexpression promotes cancer-associated fibroblasts activation and tumor progression by CUL3-Nrf2 pathway in lung adenocarcinoma.

BACKGROUND: Cancer-associated fibroblasts (CAFs) are critically involved in tumor progression by maintaining extracellular mesenchyma (ECM) production and improving tumor development. Cyclooxygenase-2 (COX-2) has been proved to promote ECM formation and tumor progression. However, the mechanisms of COX-2 mediated CAFs activation have not yet been elucidated. Therefore, we conducted this study to identify the effects and mechanisms of COX-2 underlying CAFs activation by tumor-derived exosomal miRNAs in lung adenocarcinoma (LUAD) progression. METHODS: As measures of CAFs activation, the expressions of fibroblasts activated protein-1 (FAP-1) and α-smooth muscle actin (α-SMA), the main CAFs markers, were detected by Western blotting and Immunohistochemistry. And the expression of Fibronectin (FN1) was used to analyze ECM production by CAFs. The exosomes were extracted by ultracentrifugation and exo-miRNAs were detected by qRT-PCR. Herein, we further elucidated the implicated mechanisms using online prediction software, luciferase reporter assays, co-immunoprecipitation, and experimental animal models. RESULTS: In vivo, a positive correlation was observed between the COX-2 expression levels in parenchyma and α-SMA/FN1 expression levels in mesenchyma in LUAD. However, PGE2, one of major product of COX-2, did not affect CAFs activation directly. COX-2 overexpression increased exo-miR-1290 expression, which promoted CAFs activation. Furthermore, Cullin3 (CUL3), a potential target of miR-1290, was found to suppress COX-2/exo-miR-1290-mediated CAFs activation and ECM production, consequently impeding tumor progression. CUL3 is identified to induce the Nuclear Factor Erythroid 2-Related Factor 2 (NFE2L2, Nrf2) ubiquitination and degradation, while exo-miR-1290 can prevent Nrf2 ubiquitination and increase its protein stability by targeting CUL3. Additionally, we identified that Nrf2 is direcctly bound with promoters of FAP-1 and FN1, which enhanced CAFs activation by promoting FAP-1 and FN1 transcription. CONCLUSIONS: Our data identify a new CAFs activation mechanism by exosomes derived from cancer cells that overexpress COX-2. Specifically, COX-2/exo-miR-1290/CUL3 is suggested as a novel signaling pathway for mediating CAFs activation and tumor progression in LUAD. Consequently, this finding suggests a novel strategy for cancer treatment that may tackle tumor progression in the future. Video Abstract.

CD36 deficiency ameliorates drug-induced acute liver injury in mice.

BACKGROUND: Acetaminophen (APAP) overdose causes hepatotoxicity and even acute liver failure. Recent studies indicate that sterile inflammation and innate immune cells may play important roles in damage-induced hepatocytes regeneration and liver repair. The scavenger receptor CD36 has its crucial functions in sterile inflammation. However, the roles of CD36 in APAP induced acute liver injury remain unclear and warrant further investigation. METHODS: WT C57BL/6 J and CD36-/- mice were intraperitoneally injected with APAP (300 mg/kg) after fasting for 16 h. Liver injury was evaluated by serum alanine aminotransferase (ALT) level and liver tissue hematoxylin and eosin (H&E) staining. Liver inflammatory factor expression was determined by real-time polymerase chain reaction (PCR). The protein adducts forming from the metabolite of APAP and the metabolism enzyme cytochrome P450 2E1 (CYP2E1) levels were measured by Western blot. Liver infiltrating macrophages and neutrophils were characterized by flow cytometry. RNA sequencing and Western blot were used to evaluate the effect of damage-associated molecular patterns (DAMP) molecule high mobility group B1 (HMGB1) on WT and CD36-/- macrophages. Moreover, PP2, a Src kinase inhibitor, blocking CD36 signaling, was applied in APAP model. RESULTS: The expression of CD36 was increased in the liver of mice after APAP treatment. Compared with WT mice, APAP treated CD36-/- mice show less liver injury. There was no significant difference in APAP protein adducts and CYP2E1 expression between these two strains. However, reduced pro-inflammatory factor mRNA expression and serum IL-1ß level were observed in APAP treated CD36-/- mice as well as infiltrating macrophages and neutrophils. Moreover, CD36 deficiency impaired the activation of c-Jun N-terminal kinase (JNK) caused by APAP. Interestingly, the lack of CD36 reduced the activation of extracellular regulated protein kinases (Erk) and v-akt murine thymoma viral oncogene homolog (Akt) induced by HMGB1. RNA transcription sequencing data indicated that HMGB1 has a different effect on WT and CD36-/- macrophages. Furthermore, treatment with PP2 attenuated APAP induced mouse liver injury. CONCLUSION: Our data demonstrated that CD36 deficiency ameliorated APAP-induced acute liver injury and inflammatory responses by decreasing JNK activation. CD36 might serve as a new target to reduce acute liver injury.

CPT1A-mediated succinylation of S100A10 increases human gastric cancer invasion.

Gastric cancer (GC) is a malignancy of the lining of the stomach and is prone to distant metastasis, which involves a variety of complex molecules. The S100 proteins are a family of calcium-binding cytosolic proteins that possess a wide range of intracellular and extracellular functions and play pivotal roles in the invasion and migration of tumour cells. Among these, S100A10 is known to be overexpressed in GC. Lysine succinylation, a recently identified form of protein post-translational modification, is an important regulator of cellular processes. Here, we demonstrated that S100A10 was succinylated at lysine residue 47 (K47), and levels of succinylated S100A10 were increased in human GC. Moreover, K47 succinylation of S100A10 was stabilized by suppression of ubiquitylation and subsequent proteasomal degradation. Furthermore, carnitine palmitoyltransferase 1A (CPT1A) was found to function as a lysine succinyltransferase that interacts with S100A10. Succinylation of S100A10 is regulated by CPT1A, while desuccinylation is regulated by SIRT5. Overexpression of a succinylation mimetic mutant, K47E S100A10, increased cell invasion and migration. Taken together, this study reveals a novel mechanism of S100A10 accumulation mediated by succinylation in GC, which promotes GC progression and is regulated by the succinyltransferase CPT1A and SIRT5-mediated desuccinylation.

CD36 deficiency attenuates immune-mediated hepatitis in mice by modulating the proapoptotic effects of CXC chemokine ligand 10.

The scavenger receptor CD36 recognizes a diverse set of ligands and has been implicated in a wide variety of normal and pathological processes, including lipid metabolism, angiogenesis, atherosclerosis, and phagocytosis. In particular, recent findings have demonstrated its crucial functions in sterile inflammation and tumor metastasis. However, the role of CD36 in immune-mediated hepatitis remains unclear. Concanavalin A (ConA)-induced liver injury is a well-established experimental T cell-mediated hepatitis. To understand the role of CD36 in hepatitis, we tested the susceptibility of CD36-deficient (CD36-/- ) mice to this model, evaluated by a liver enzyme test, terminal deoxynucleotidyl transferase dUTP nick-end labeling (TUNEL) assay, histological analysis, mononuclear cell (MNC) infiltration, and hepatic proinflammatory factor production. CD36-/- mice were less sensitive to ConA-induced hepatitis and had a significantly lower number of liver MNCs (LMNCs), including CD4+ cells, CD8+ T cells, natural killer cells, natural killer T cells, infiltrating macrophages, and neutrophils, as well as reduced expression of inflammatory mediators (tumor necrosis factor α, CXC chemokine ligand (CXCL) 10, interleukin (IL)-1α, monocyte chemotactic protein 1, and IL-6) compared with controls. Notably, we used bone marrow chimeric mice to demonstrate that CD36 expression on nonhematopoietic cells was required to drive ConA-induced liver injury. Furthermore, our data show that the CD36 receptor was essential for CXCL10-induced hepatocyte apoptosis and activation of IκB kinase, Akt, and Jun N-terminal kinase. Moreover, treatment of wild-type mice with genistein, a tyrosine kinase inhibitor that blocks CD36-Lyn signaling, attenuated ConA-induced liver injury and reduced the number of MNCs. CONCLUSIONS: Our findings suggest that CD36 plays an important proinflammatory role in ConA-induced liver injury by promoting hepatic inflammation and mediating the proapoptotic effect of chemokine CXCL10, and therefore, may be a potential therapeutic target for immune-mediated hepatitis. (Hepatology 2018;67:1943-1955).

Helicobacter pylori-induced exosomal MET educates tumour-associated macrophages to promote gastric cancer progression.

Helicobacter pylori (H. pylori) infection triggers chronic inflammation that has been associated with gastric cancer (GC). Exosomes are small extracellular vesicles that have become the key mediators of intercellular communication. In this study, we investigated exosome-mediated communication between H. pylori-infected GC cells and macrophages, focusing on the transfer of activated mesenchymal-epithelial transition factor (MET). We observed a significant decrease in MET protein expression in GC cells after infection with H. pylori, whereas MET mRNA levels remained unchanged. Intriguingly, MET expression, specifically the phosphorylated active form, was increased in exosomes released from H. pylori-infected GC cells. Confocal microscopy and Western blotting analyses showed that these exosomes containing MET were delivered to and internalized by macrophages. Indeed, in human GC tissues positive for H. pylori, we also observed that activated MET was highly expressed in tumour-infiltrating macrophages. After internalization, exosomal MET then appeared to educate the macrophages towards a pro-tumorigenesis phenotype. This included exosomal MET-mediated stimulation of proinflammatory cytokine secretion IL-1ß, which subsequently promoted tumour growth and progression in vitro and in vivo. Taken together, these data were the first to demonstrate H. pylori infection-induced upregulation of activated MET in exosomes and the pro-tumorigenic effect on tumour-associated macrophages.

AIM2 regulates autophagy to mitigate oxidative stress in aged mice with acute liver injury.

The cytoplasmic pattern recognition receptor, absent in melanoma 2 (AIM2), detects cytosolic DNA, activating the inflammasome and resulting in pro-inflammatory cytokine production and pyroptotic cell death. Recent research has illuminated AIM2's contributions to PANoptosis and host defense. However, the role of AIM2 in acetaminophen (APAP)-induced hepatoxicity remains enigmatic. In this study, we unveil AIM2's novel function as a negative regulator in the pathogenesis of APAP-induced liver damage in aged mice, independently of inflammasome activation. AIM2-deficient aged mice exhibited heightened lipid accumulation and hepatic triglycerides in comparison to their wild-type counterparts. Strikingly, AIM2 knockout mice subjected to APAP overdose demonstrated intensified liver injury, compromised mitochondrial stability, exacerbated glutathione depletion, diminished autophagy, and elevated levels of phosphorylated c-Jun N-terminal kinase (JNK) and extracellular signal-regulated kinase (ERK). Furthermore, our investigation revealed AIM2's mitochondrial localization; its overexpression in mouse hepatocytes amplified autophagy while dampening JNK phosphorylation. Notably, induction of autophagy through rapamycin administration mitigated serum alanine aminotransferase levels and reduced the necrotic liver area in AIM2-deficient aged mice following APAP overdose. Mechanistically, AIM2 deficiency exacerbated APAP-induced acute liver damage and inflammation in aged mice by intensifying oxidative stress and augmenting the phosphorylation of JNK and ERK. Given its regulatory role in autophagy and lipid peroxidation, AIM2 emerges as a promising therapeutic target for age-related acute liver damage treatment.

CD5L deficiency attenuate acetaminophen-induced liver damage in mice via regulation of JNK and ERK signaling pathway.

CD5 molecule like (CD5L), a member of the scavenger receptor cysteine-rich domain superfamily, plays a critical role in immune homeostasis and inflammatory disease. Acetaminophen (APAP) is a safe and effective antipyretic analgesic. However, overdose may cause liver damage or even liver failure. APAP hepatotoxicity is characterized by extensive necrotic cell death and a sterile inflammatory response, in which the role of CD5L remains to be investigated. In this study, we found that the expression of CD5L was increased in the livers of mice after APAP overdose. Furthermore, CD5L deficiency reduced the increase of alanine transaminase (ALT) level, histopathologic lesion area, c-Jun N-terminal kinase (JNK)/extracellular signal-regulated kinase (ERK) phosphorylation level, Transferase-Mediated dUTP Nick End-Labeling positive (TUNEL+) cells proportion, vascular endothelial cell permeability and release of inflammatory cytokines induced by excess APAP. Therefore, our findings reveal that CD5L may be a potential therapeutic target for prevention and treatment of APAP-induced liver injury.

Application of truncated immunodominant polypeptide from hepatitis E virus (HEV) ORF2 in an assay to exclude nonspecific binding in detecting anti-HEV immunoglobulin M.

The diagnosis of recent hepatitis E virus (HEV) infection depends on serologic testing for anti-HEV IgM; however, false-positive results may occur. In the present study, we cloned the ORF2 fragment of genotype 4 HEV and demonstrated that a subregion covering amino acids 459 to 607 in ORF2 forms the immunodominant B-cell epitopes, as it does in genotype 1 viruses. Truncation of several residues from either the N or C terminus of the polypeptide abolished the reactivity of anti-HEV from naturally infected persons. By the combination of high reactivity of the immunodominant polypeptide and poor reactivity of the truncated polypeptide, we established an indirect enzyme-linked immunosorbent assay (ELISA) to detect anti-HEV IgM. In this assay, all 37 sera that were HEV RNA positive reacted with the immunodominant polypeptide but not with the truncated one, and none of 159 sera from healthy persons reacted with either of the polypeptides. In retesting of 117 sera that originally tested positive for anti-HEV IgM, using a Genelabs kit, only 34 were positive and 83 were negative. Western blot analyses and other experiments strongly indicated that these 83 discordant sera were negative for anti-HEV IgM. Furthermore, among the 117 sera, 5 reacted with both the immunodominant and truncated polypeptides, with comparable optical densities at 450 nm. However, their reactivity was demonstrated to result from nonspecific binding. Together, the data indicate that the poor reactivity of a truncated ORF2 polypeptide can be used to exclude nonspecific binding in the detection of anti-HEV IgM.

Lysine-222 succinylation reduces lysosomal degradation of lactate dehydrogenase a and is increased in gastric cancer.

BACKGROUND: Lysine succinylation is an emerging posttranslational modification that has garnered increased attention recently, but its role in gastric cancer (GC) remains underexplored. METHODS: Proteomic quantification of lysine succinylation was performed in human GC tissues and adjacent normal tissues by mass spectrometry. The mRNA and protein levels of lactate dehydrogenase A (LDHA) in GC and adjacent normal tissues were analyzed by qRT-PCR and western blot, respectively. The expression of K222-succinylated LDHA was measured in GC tissue microarray by the K222 succinylation-specific antibody. The interaction between LDHA and sequestosome 1 (SQSTM1) was measured by co-immunoprecipitation (co-IP) and proximity ligation assay (PLA). The binding of carnitine palmitoyltransferase 1A (CPT1A) to LDHA was determined by co-IP. The effect of K222-succinylated LDHA on tumor growth and metastasis was evaluated by in vitro and in vivo experiments. RESULTS: Altogether, 503 lysine succinylation sites in 303 proteins were identified. Lactate dehydrogenase A (LDHA), the key enzyme in Warburg effect, was found highly succinylated at K222 in GC. Intriguingly, this modification did not affect LDHA ubiquitination, but reduced the binding of ubiquitinated LDHA to SQSTM1, thereby decreasing its lysosomal degradation. We demonstrated that CPT1A functions as a lysine succinyltransferase that interacts with and succinylates LDHA. Moreover, high K222-succinylation of LDHA was associated with poor prognosis in patients with GC. Finally, overexpression of a succinylation-mimic mutant of LDHA promoted cell proliferation, invasion, and migration. CONCLUSIONS: Our data revealed a novel lysosomal pathway of LDHA degradation, which is mediated by the binding of K63-ubiquitinated LDHA to SQSTM1. Strikingly, CPT1A succinylates LDHA on K222, which thereby reduces the binding and inhibits the degradation of LDHA, as well as promotes GC invasion and proliferation. This study thus uncovers a new role of lysine succinylation and the mechanism underlying LDHA upregulation in GC.

[Effects of inhibition of ANG-2 expression in Ishikawa cell line].

OBJECTIVE: To investigate the effect of RNA interference mediated angiopoietin-2 (ANG-2) gene silencing on human endometrial carcinoma cell line Ishikawa. METHODS: Short hairpin RNA (shRNA) targeting ANG-2 gene was designed and transfected into Ishikawa cells with Lipofectamine 2000. The mRNA and protein expression level of ANG-2, proliferation, morphological changes, apoptosis, cell cycle and invasive ability of the cells after transfection were analyzed. RESULTS: The shRNA targeting the human ANG-2 gene effectively decreased the expression of ANG-2 on both mRNA and protein level, the proliferation inhibition rate of the Ishikawa cells was 63.11%, cell apoptosis was induced, and the cell cycle was arrested in G1 phase. The apoptotic rate of the Ishikawa cells in the transfected group was significantly higher, and the invasive ability was decreased markedly, than that of control groups. CONCLUSION: The shRNA targeting human ANG-2 gene could reduce ANG-2 expression, inhibit cellular growth and invasion in Ishikawa cells in vitro.

Inhibition of lung cancer growth and metastasis by DHA and its metabolite, RvD1, through miR-138-5p/FOXC1 pathway.

BACKGROUND: Non small cell lung cancer (NSCLC) is one of the most common cancers in the world. DHA is known to be capable of suppressing NSCLC cell proliferation and metastasis. However, the mechanisms by which DHA exhibits its antitumor effects are unknown. Here we aimed to identify the effects and mechanisms of DHA and its metabolites on lung cancer cell growth and invasion. METHODS: As measures of cell proliferation and invasion ability, the cell viability and transwell assays were used in vitro. Transgenic mfat-1 mice, which convert ω-6 PUFAs to ω-3 PUFAs, were used to detect the effect of endogenous DHA on tumor transplantation. An LC - MS/MS analysis identified the elevation of several eicosanoid metabolites of DHA. By using qPCR miRNA microarray, online prediction software, luciferase reporter assays and Western blot analysis, we further elucidated the mechanisms. RESULTS: Addition of exogenous DHA inhibited the growth and invasion in NSCLC cells in vitro. Endogenously produced DHA attenuated LLC-derived tumor growth and metastasis in the transgenic mfat-1 mice. Among the elevation of DHA metabolites, resolvin D1 (RvD1) significantly contributed to the inhibition in cell growth and invasion. MiRNA microarray revealed that the level of miR-138-5p was significantly increased after RvD1 treatment. MiR-138-5p mimics decreased cell viability and invasion; while miR-138-5p inhibitor abolished RvD1-mediated suppression of cell viability and invasion. The expression of FOXC1 was significantly reduced upon overexpression of miR-138-5p while luciferase reporter assay showed that FOXC1 was a direct target of miR-138-5p. In vivo, endogenous DHA by the mfat-1 transgene enhanced miR-138-5p expression and decreased FOXC1 expression. Furthermore, overexpression of FOXC1 reversed the inhibition in cell viability and invasion induced by RvD1 treatment. CONCLUSIONS: These data identified the RvD1/miR-138-5p/FOXC1 pathway as a novel mechanism by DHA and its metabolite, RvD1, and the potential of targeting such pathway as a therapeutic strategy in treating NSCLC.

Tumor-derived lactate induces M2 macrophage polarization via the activation of the ERK/STAT3 signaling pathway in breast cancer.

Tumor-associated macrophages (TAM) are prominent components of tumor microenvironment (TME) and capable of promoting cancer progression. However, the mechanisms for the formation of M2-like TAMs remain enigmatic. Here, we show that lactate is a pivotal oncometabolite in the TME that drives macrophage M2-polarization to promote breast cancer proliferation, migration, and angiogenesis. In addition, we identified that the activation of ERK/STAT3, major signaling molecules in the lactate signaling pathway, deepens our molecular understanding of how lactate educates TAMs. Moreover, suppression of ERK/STAT3 signaling diminished tumor growth and angiogenesis by abolishing lactate-induced M2 macrophage polarization. Finally, research data of the natural compound withanolide D provide evidence for ERK/STAT3 signaling as a potential therapeutic strategy for the prevention and treatment of breast cancer. These findings suggest that the lactate-ERK/STAT3 signaling pathway is a driver of breast cancer progression by stimulating macrophage M2-like polarization and reveal potential new therapeutic targets for breast cancer treatment.

Chitinase 3-like 1-CD44 interaction promotes metastasis and epithelial-to-mesenchymal transition through ß-catenin/Erk/Akt signaling in gastric cancer.

BACKGROUND: Enzymatically inactive chitinase-like protein CHI3L1 drives inflammatory response and promotes tumor progression. However, its role in gastric cancer (GC) tumorigenesis and metastasis has not yet been fully elucidated. We determined the significance of CHI3L1 expression in patients with GC. We also explored an as-yet unknown receptor of CHI3L1 and investigated the involved signaling in GC metastasis. METHODS: CHI3L1 expression was evaluated by immunoblotting, tissue microarray-based immunohistochemistry analysis (n = 100), and enzyme linked immunosorbent assay (ELISA) (n = 150). The interactions between CD44 and CHI3L1 or Interleukin-13 receptor alpha 2 (IL-13Rα2) were analyzed by co-immunoprecipitation, immunofluorescence co-localization assay, ELISA, and bio-layer interferometry. The roles of CHI3L1/CD44 axis in GC metastasis were investigated in GC cell lines and experimental animal model by gain and loss of function. RESULTS: CHI3L1 upregulation occurred during GC development, and positively correlated with GC invasion depth, lymph node status, and tumor staging. Mechanically, CHI3L1 binding to CD44 activated Erk and Akt, along with ß-catenin signaling by phosphorylating ß-catenin at Ser552 and Ser675. CD44 also interacted with IL-13Rα2 to form a complex. Notably, CD44v3 peptide and protein, but not CD44v6 peptide or CD44s protein, bound to both CHI3L1 and IL-13Rα2. Our in vivo and in vitro data further demonstrated that CHI3L1 promoted GC cell proliferation, migration, and metastasis. CONCLUSIONS: CHI3L1 binding to CD44v3 activates Erk, Akt, and ß-catenin signaling, therefore enhances GC metastasis. CHI3L1 expression is a novel biomarker for the prognosis of GC, and these findings have thus identified CHI3L1/CD44 axis as a vital pathway and potential therapeutic target in GC.

An Ω-3 fatty acid desaturase-expressing gene attenuates prostate cancer proliferation by cell cycle regulation.

Previous studies have reported that Ω-6 and Ω-3 fatty acids have opposing effects on cancer development. Consuming high levels of long-chain Ω-3 polyunsaturated fatty acids (PUFAs) has been shown to reduce prostate cancer risk and increase chemotherapy sensitivity. The sdd17 gene encodes an Ω-3 fatty acid desaturase, which converts arachidonic acid into eicosapentaenoic acid (EPA). However, little is known regarding the function of the sdd17 gene in tumor cells in vitro. In the present study, prostate cancer cells were infected with the msdd17 gene, which allowed the endogenous production of Ω-3 PUFAs. The cells that expressed the msdd17 gene had high levels of long-chain Ω-3 PUFAs compared with the control cells. Expression of the msdd17 gene significantly inhibited prostate cancer cell proliferation. EPA exposure and msdd17 gene transfection each induced G2 cell cycle arrest and reduced E2F transcription factor 1 expression in prostate cancer cells. These results suggest that msdd17 gene transfection suppressed prostate cancer cell proliferation and induced G2 cell cycle arrest.

Oncometabolite succinate promotes angiogenesis by upregulating VEGF expression through GPR91-mediated STAT3 and ERK activation.

Altered cellular metabolism is now generally acknowledged as a hallmark of cancer cells, the resultant abnormal oncometabolites cause both metabolic and nonmetabolic dysregulation and potential transformation to malignancy. A subset of cancers have been found to be associated with mutations in succinate dehydrogenase genes which result in the accumulation of succinate. However, the function of succinate in tumorigenesis remains unclear. In the present study, we aim to investigate the role of oncometabolite succinate in tumor angiogenesis. Our data demonstrated the accumulation of markedly elevated succinate in gastric cancer tissues compared with that in paracancerous tissues. Moreover, succinate was able to increase the chemotactic motility, tube-like structure formation and proliferation of primary human umbilical vascular endothelial cells (pHUVECs) in vitro, as well as promoting the blood vessel formation in transgenic zebrafish. Our mechanistic studies reveal that succinate upregulates vascular endothelial growth factor (VEGF) expression by activation of signal transducer and activator of transcription 3 (STAT3) and extracellular regulated kinase (ERK)1/2 via its receptor GPR91 in a HIF-1α independent mechanism. Taken together, these data indicate an important role of the succinate-GPR91 axis in tumor angiogenesis, which may enable development of a novel therapeutic strategy that targets cancer metabolism.

IκB-kinase-ε in the tumor microenvironment is essential for the progression of gastric cancer.

The tumor microenvironment is critical for tumor growth and metastasis, but the underlying molecular mechanisms are poorly understood. Recent studies have shown that IκB-kinase-ε (IKKε) is involved in the proliferation and migration of certain cancers. However, the functional role of IKKε in the progression of gastric cancer (GC) remains unknown. In this study, we found that high levels of IKKε expression in GC tumors were correlated with more advanced disease and poor overall survival of patients. Silencing of IKKε effectively suppressed the migratory and invasive capabilities of human GC cells in vitro and tumorigenicity and metastasis in vivo. Further analysis revealed that IKKε was also highly expressed in tumor-infiltrating lymphocytes. Moreover, it was involved in tumor-infiltrating T-cell-mediated invasion and metastasis. Knockdown of IKKε elevated T-cell antitumor immunity. These findings suggest that IKKε may be a novel prognostic marker and a potential therapeutic target in human GCs.

Associations of plasma 8-isoprostane levels with the presence and extent of coronary stenosis in patients with coronary artery disease.

Oxidative stress may play a role in the development of atherosclerosis. The purpose of the present study was to explore the relationship between 8-isoprostaglandin F(2alpha) (8-iso-PGF(2alpha)) levels and the presence of coronary artery disease (CAD) and to also clarify whether 8-iso-PGF(2alpha) might add independently to measures of CAD extent. The study group consisted of 241 consecutive patients who were undergoing coronary angiography for suspected CAD. 8-iso-PGF(2alpha) levels were recorded for all participants. The analysis revealed a significant difference in 8-iso-PGF(2alpha) levels in patients with and without hypertension (P<0.001), in patients with diabetes relative to nondiabetic patients (P<0.05), and in males respect to females (P<0.001). A significant positive correlation was found between age and 8-iso-PGF(2alpha) levels (P<0.001). 8-iso-PGF(2alpha) levels correlated with the number of cardiovascular risk factors (P<0.001). 8-iso-PGF(2alpha) levels were higher in the CAD(+) respect to the CAD(-) groups (337.7+/-80.2 and 263.8+/-74.2 pg/ml and P<0.001). A stepwise elevation in the 8-iso-PGF(2alpha) levels was found depending on the number of affected vessels (P<0.001). The 8-iso-PGF(2alpha) levels showed a significant positive correlation with the numbers of >50 and >25% stenotic segments (P<0.001) and the extent score of coronary stenosis (P<0.001). The multivariate logistic regression analysis indicated 8-iso-PGF(2alpha) as an independent factor associated with CAD (odds ratio, 2.47 and P=0.001). The results suggested that 8-iso-PGF(2alpha) is associated with the presence of CAD in patients undergoing coronary angiography and is also related to the extent of coronary stenosis in Chinese population.

Inhibitive effect of genistein on hypoxia-induced basic fibroblast growth factor expression in human retinal pigment epithelium cells.

AIM: The time course changes of basic fibroblast growth factor (bFGF) expression induced by hypoxia and the effects of genistein on hypoxia-induced bFGF expression in the human retinal pigment epithelium (RPE) cells were studied. METHODS: The bFGF mRNA expression was examined by reverse transcription polymerase chain reaction. The bFGF protein expression was detected by Western blot. RESULTS: Hypoxia significantly increased bFGF mRNA expression. The maximal level detected at 24 h was approximately two times that at the start of treatment. With pretreatment of genistein (10, 20, 50, 100, and 200 microM) for 30 min, the elevated expression of bFGF mRNA was suppressed in a concentration-dependent manner. bFGF mRNA expression was reduced to 30.4% by 200 microM of genistein when compared with that untreated with genistein. Hypoxia treatment also remarkably increased the expression of bFGF protein. At 24 h after hypoxia, when the highest expression of bFGF protein was observed, it was approximately two times as much as that at the start of treatment. Genistein (10, 20, 50, 100, and 200 microM) could also suppress bFGF protein expression in a concentration-dependent manner. The highest suppression was observed when exposed to 200 microM of genistein, which was 43% of control. CONCLUSIONS: These results suggested that suppression of bFGF expression in RPE cells might partly account for the inhibitive effect of genistein on retinal neovascularization in vivo.

Clinical and Virological Characteristics of Chronic Hepatitis B Patients with Coexistence of HBsAg and Anti-HBs.

Coexistence of hepatitis B surface antigen (HBsAg) and antibody against HBsAg (anti-HBs) comprises an atypical serological profile in patients with chronic hepatitis B virus (HBV) infection. In this study, in total 94 patients with coexisting HBsAg and anti-HBs and 94 age- and sex-matched patients with positive HBsAg were characterized by quantitatively measuring HBsAg and HBV DNA, sequencing large S genes, and observing clinical features. Compared with common hepatitis B patients, the patients with coexisting HBsAg and anti-HBs had lower HBsAg and HBV DNA levels. These two groups had similar rate of pre-S deletion mutations. However, in patients with coexisting HBsAg and anti-HBs, more amino acid substitutions in the a determinant of S gene were observed in HBV genotype C, but not in genotype B. Fourteen patients with coexisting HBsAg and anti-HBs were followed up for an average of 15.5 months. There were no significant changes in the levels of HBsAg, anti-HBs, HBV DNA and ALT over the follow-up period. Compared with the baseline sequences, amino acid substitutions in the MHR of HBsAg occurred in 14.3% (2/14) patients. In conclusion, coexistence of HBsAg and anti-HBs may be associated with higher frequency of mutations in the a determinant of HBV genotype C.

Adiponectin Deficiency Leads to Female Subfertility and Ovarian Dysfunctions in Mice.

Adipose tissue plays an important role in regulating female fertility, owing to not only its energy stores but also the endocrine actions of secreted adipokines. As one of the adipokines, adiponectin is almost exclusively secreted from the fat, and its circulating concentration is paradoxically reduced in obesity. Although recent studies implied a purported positive role of adiponectin in ovarian functions, definitive in vivo evidence has been sorely lacking. We have consistently observed subfertility in female adiponectin null mice and therefore postulated a protective role of adiponectin in ovarian functions. Female adiponectin null mice displayed impaired fertility, reduced retrieval of oocytes, disrupted estrous cycle, elevated number of atretic follicles, and impaired late folliculogenesis. Analysis of their sera revealed a significant decrease in estradiol and FSH but an increase in LH and testosterone at proestrus. In addition, we found marked reduction of progesterone levels at diestrus, a significant decrease in LH receptor expression as well as in the number of GnRH immunoreactive neurons. Adiponectin deficiency also altered the peak concentrations of LH surge and led to lower expression of Cytochrome P450 family 11 subfamily A member 1 (P450scc), an enzyme critical for progesterone synthesis, as well as an increase in BCL2 associated X, apoptosis regulator and Insulin like growth factor binding protein 4 in atretic follicles. These physiological and molecular events were independent of insulin sensitivity. Thus, we have revealed a novel mechanism linking adiponectin and female fertility that entails regulation of reproductive hormone balance and ovarian follicle development.