A rare subpopulation of cancer cells, termed cancer stem cells (CSCs), may be responsible for tumor relapse and resistance to conventional chemotherapy. The development of a non-toxic, natural treatment for the elimination of CSCs is considered a strategy for cancer treatment with minimal side effects. In the present study, the potential for Sasa quelpaertensis leaf extract (SQE) and its two bioactive compounds, tricin and p-coumaric acid, to exert anti-CSC effects by suppressing cancer stemness characteristics were evaluated in colon cancer cells. CD133+CD44+ cells were isolated from HT29 and HCT116 cell lines using flow-activated cell sorting (FACs). SQE treatment was found to significantly suppress the self-renewal capacity of both cell lines. SQE treatment was also associated with the down-regulation of β-catenin and phosphorylated GSK3β, while significantly enhancing cell differentiation by up-regulating CK20 expression and blocking the expression of several stem cell markers, including DLK1, Notch1, and Sox-2. In vivo, SQE supplementation suppressed tumor growth in a xenograft model by down-regulating stem cell markers and β-catenin as well as HIF-1α signaling. Compared with two bioactive compounds of SQE, SQE exhibited the most effective anti-CSC properties. Taken together, these results provide evidence that SQE inhibits colon cancer by regulating the characteristics of CSCs.

Ischemia/reperfusion injury triggers acute kidney injury (AKI) by aggravating oxidative stress mediated mitochondria dysfunction. The peroxisome proliferator-activated receptor gamma coactivator 1α (PGC-1α) is a master player that regulates mitochondrial biogenesis and the antioxidant response. We postulated that PGC-1α functions as cytoprotective effector in renal cells and that its regulation mechanism is coordinated by nuclear factor erythroid 2-related factor 2 (Nrf-2). In this study, to understand the effect and molecular mechanisms of PGC-1α, we developed an empty vector or PGC-1α-overexpressing stable cell lines in HK-2 cells (Mock or PGC-1α stable cells). PGC-1α overexpression increased the viability of cells affected by H2O2 mediated injury, protected against H2O2-mediated apoptotic events and inhibited reactive oxygen species accumulation in the cytosol and mitochondria as compared to that in Mock cells. The cytoprotective effect of PGC-1α was related to Nrf-2 upregulation, which was counteracted by Nrf-2-specific knockdown. Using inhibitor of p38, we found that regulation of the p38/glycogen synthase kinase 3β (GSK3β)/Nrf-2 axis was involved in the protective effects of PGC-1α. Taken together, we suggest that PGC-1α protects human renal tubule cells from H2O2-mediated apoptotic injury by upregulating Nrf-2 via GSK3β inactivation mediated by activated p38.

Instances of sustained oxidative activity have been shown to involve dysregulation of Nrf2-mediated transcriptional induction; however, mechanisms warranting Nrf2-repression remain unclear. In this study, using primary rat hepatocytes, we have attempted to identify factors that may negatively influence Nrf2 survival pathway. Though studies indicate a conspicuous association between Akt and Nrf2, a confirmatory link between the two is unaddressed. On inhibiting PI3K/Akt pathway, we observed compromised activities of antioxidant and detoxification enzymes culminating in oxidative cytotoxicity. This was accompanied by reduced nuclear retention of Nrf2 and its ARE binding affinity, increased Nrf2 ubiquitination and concurrent decline in its downstream targets. Moreover, Akt inhibition enhanced nuclear translocation as well as phosphorylation of Fyn kinase, an enzyme linked to Nrf2 degradation, by relieving GSK3β from phosphorylation-mediated repression. The involvement of Akt and Fyn kinase in influencing Nrf2 signaling was further confirmed in oxidatively stressed hepatocytes by using tert-butyl hydroperoxide (tBHP). tBHP-induced decrease in Nrf2 levels was associated with enhanced Fyn kinase phosphorylation, Fyn kinase nuclear translocation and decreased levels of phosphorylated GSK3β(Ser9) in a time-dependent manner. Interestingly, tBHP induced site-specific deactivation of Akt as only Akt(Ser473) phosphorylation was observed to be affected. Further, protein expression as well as nuclear localization of PHLPP2, a phosphatase specific for Akt(Ser473), was found to be significantly enhanced in tBHP-stressed hepatocytes. Silencing of PHLPP2 not only resulted in considerable restoration of Nrf2 signaling, enhanced Nrf2-ARE binding and reduced Nrf2 ubiquitination but also significantly suppressed tBHP-induced ROS generation and alterations in mitochondrial permeability. We infer that cellular PHLPP2 levels may aggravate oxidative toxicity by suppressing Nrf2/ARE transcriptional regulation via Akt(Se473)/GSK3β/Fyn kinase axis. The study indicates that PHLPP2 could serve as a new target for developing strategies to manage pathological conditions exacerbated due to oxidative stress.

3-Methyladenine (3-MA) is one of the most commonly used inhibitors in autophagy research today. However, rather than inhibiting class III PI3K that is involved in autophagy suppression, 3-MA might also interfere with class I PI3K and consequently augment autophagy flux. In this study, we aim to get a thorough understanding on the action mechanisms of 3-MA in TLR4-mediated inflammatory responses in RAW264.7 macrophages and, moreover, to decipher the action of 3-MA in modulation of autophagy. We found that 3-MA could enhance LPS-induced NF-κB activation and production of TNF-α, inducible NO synthase (iNOS), cyclooxygenase-2, IL-1β, and IL-12. In contrast, 3-MA suppressed LPS-induced IFN-β production and STAT signaling. Studies revealed that 3-MA can, through inhibition of Akt as a result of class I PI3K interference, positively regulate p38, JNK, and p65, but negatively regulate TANK-binding kinase 1 and IFN regulatory factor 3 mediated by TLR4. As glycogen synthase kinase 3β (GSK3β) is an important Akt substrate, we further explored its involvement in the actions of 3-MA. 3-MA was found to enhance LPS-induced NF-κB activation, iNOS, and pro-IL-1β expression, and these actions were reversed by either GSK3β inhibitors or small interfering GSK3β. Lastly, we demonstrated that 3-MA acts as an autophagy inducer in RAW264.7 macrophages, but the stimulating effects on NF-κB activation and iNOS and cyclooxygenase-2 expression were not affected in LPS-stimulated macrophages with small interfering autophagy protein-5 treatment. These results not only shed new light on the action mechanisms of 3-MA to differentially regulate inflammatory outcomes derived from TLR4-mediated MyD88 and Toll/IL-1R domain-containing adapter inducing IFN-β pathways, but also highlight the necessity to check autophagy status upon taking 3-MA as a general autophagy inhibitor.

Nuclear factor kappa-light-chain-enhancer of activated B cells (NFκB) is regulated by a myriad of signaling cascades including glycogen synthase kinase (GSK) 3β and plays a Janus role in podocyte injury. In vitro, lipopolysaccharide (LPS) or adriamycin (ADR) elicited podocyte injury and cytoskeletal disruption, associated with NFκB activation and induced expression of NFκB target molecules, including pro-survival Bcl-xL and podocytopathic mediators like MCP-1, cathepsin L, and B7-1. Broad-range inhibition of NFκB diminished the expression of all NFκB target genes, restored cytoskeleton integrity, but potentiated apoptosis. In contrast, blockade of GSK3β by lithium or 4-benzyl-2-methyl-1,2,4-thiadiazolidine-3,5-dione (TDZD-8) mitigated the expression of podocytopathic mediators, ameliorated podocyte injury, but barely affected Bcl-xL expression or sensitized apoptosis. Mechanistically, GSK3β was sufficient and essential for RelA/p65 phosphorylation, specifically at serine 467, which specifies the expression of selective NFκB target molecules, including podocytopathic mediators, but not Bcl-xL. In vivo, lithium or TDZD-8 therapy improved podocyte injury and proteinuria in mice treated with LPS or ADR, concomitant with the suppression of podocytopathic mediators, but retained Bcl-xL in glomerulus. Broad-range inhibition of NFκB conferred similar but much weakened antiproteinuric and podoprotective effects accompanied with a blunted glomerular expression of Bcl-xL and marked podocyte apoptosis. Thus, the GSK3β-dictated fine-tuning of NFκB may serve as a novel therapeutic target for podocytopathy.

SUMO conjugation has emerged as a dynamic process in regulating protein function. Here we identify estrogen receptor β (ERβ) to be a new target of SUMO-1. ERβ SUMO-1 modification occurs on a unique nonconsensus sumoylation motif which becomes fully competent upon phosphorylation of its contained serine residue, which provides the essential negative charge for sumoylation. This process is further regulated by phosphorylation of additional adjacent serine residues by glycogen synthase kinase 3β (GSK3β), which maximizes ERβ sumoylation in response to hormone. SUMO-1 attachment prevents ERβ degradation by competing with ubiquitin at the same acceptor site and dictates ERβ transcriptional inhibition by altering estrogen-responsive target promoter occupancy and gene expression in breast cancer cells. These findings uncovered a novel phosphorylated sumoylation motif (pSuM), which consists of the sequence ψKXS (where ψ represents a large hydrophobic residue) and which is connected to a GSK3-activated extension that functions as a SUMO enhancer. This extended pSuM offers a valuable signature to predict SUMO substrates under protein kinase regulation.

Increasing evidence supports an association between Alzheimer's disease (AD) and diabetes. Rosiglitazone, a peroxisome proliferator-activated receptor-γ (PPARγ) agonist, which is an anti-diabetic agent against type 2 diabetes, is currently in Phase III clinical trials in AD patients because rosiglitazone reduces β-amyloid (Aβ) pathology and inflammation. However, few studies have investigated whether rosiglitazone affects tau phosphorylation, another critical pathological feature of AD. Thus, we investigated it using OLETF type 2 diabetic rats and streptozotocin-injected type 1 diabetic mice. Interestingly, rosiglitazone reduced tau phosphorylation only in the hippocampus of OLETF type 2 diabetes rats, and not in that of STZ-injected type 1 diabetes mice. The activity of JNK was reduced in the hippocampus of rosiglitazone-treated OLETF rats, correlating with a reduction in tau phosphorylation, however, which was not correlated with GSK3β activity. In human tau-transfected SH-SY5Y neuronal cell line, reduction of tau phosphorylation was also associated with reduction of JNK activity, not of GSK3β activity. Hence, rosiglitazone could be used in reducing tau phosphorylation through JNK inactivation for therapeutic effects in type 2 diabetes related Alzheimer's disease.

BACKGROUND

Neuroinflammation often results in enduring cognitive impairment and is a risk factor for postoperative cognitive dysfunction. There are currently no effective treatments for infection-induced cognitive impairment. Previous studies have shown that the iron chelator deferoxamine (DFO) can increase the resistance of neurons to injury and disease by stimulating adaptive cellular stress responses. However, the impact of DFO on the cognitive sequelae of neuroinflammation is unknown.

METHODS

A mouse model of lipopolysaccharide (LPS)-induced cognitive impairment was established to evaluate the neuroprotective effects of DFO against LPS-induced memory deficits and neuroinflammation. Adult C57BL/6 mice were treated with 0.5 μg of DFO 3 days prior to intracerebroventricular microinjection of 2 μg of LPS. Cognitive function was assessed using a Morris water maze from post-injection days 1 to 3. Animal behavioral tests, as well as pathological and biochemical assays were performed to evaluate the LPS-induced hippocampal damage and the neuroprotective effect of DFO.

RESULTS

Treatment of mice with LPS resulted in deficits in cognitive performance in the Morris water maze without changing locomotor activity, which were ameliorated by pretreatment with DFO. DFO prevented LPS-induced microglial activation and elevations of IL-1β and TNF-α levels in the hippocampus. Moreover, DFO attenuated elevated expression of caspase-3, modulated GSK3β activity, and prevented LPS-induced increases of MDA and SOD levels in the hippocampus. DFO also significantly blocked LPS-induced iron accumulation and altered expression of proteins related to iron metabolism in the hippocampus.

CONCLUSIONS

Our results suggest that DFO may possess a neuroprotective effect against LPS-induced neuroinflammation and cognitive deficits via mechanisms involving maintenance of less brain iron, prevention of neuroinflammation, and alleviation of oxidative stress and apoptosis.

Multiple organ systems, including the brain, which undergoes changes that may increase the risk of cognitive decline, are adversely affected by diabetes mellitus (DM). Here, we demonstrate that type 2 diabetes mellitus (T2DM) db/db mice exhibited hippocampus-dependent memory impairment, which might associate with a reduction in dendritic spine density in the pyramidal neurons of brain, Aβ1-42 deposition in the prefrontal cortex (PFC) and hippocampus, and a decreased expression of neurostructural proteins including microtubule-associated protein (MAP2), a marker of dendrites, and postsynaptic density 95 (PSD95), a marker of excitatory synapses. To investigate the effects of the ZiBuPiYin recipe (ZBPYR), a traditional Chinese medicine recipe, on diabetes-related cognitive decline (DACD), db/db mice received daily administration of ZBPYR over an experimental period of 6 weeks. We then confirmed that ZBPYR rescued learning and memory performance impairments, reversed dendritic spine loss, reduced Aβ1-42 deposition and restored the expression levels of MAP2 and PSD95. The present study also revealed that ZBPYR strengthened brain leptin and insulin signaling and inhibited GSK3β overactivity, which may be the potential mechanism or underlying targets of ZBPYR. These findings conclude that ZBPYR prevents DACD, most likely by improving dendritic spine density and attenuating brain leptin and insulin signaling pathway injury. Our findings provide further evidence for the effects of ZBPYR on DACD.

Neuroinflammation is a crucial mechanism related to many neurological diseases. Extensive studies in recent years have indicated that dysregulation of Glycogen Synthase Kinase 3 Beta (GSK3β) contributes to the development and progression of these disorders through regulating the neuroinflammation processes. Inhibitors of GSK3β have been shown to be beneficial in many neuroinflammatory disease models including Alzheimer's disease, multiple sclerosis and AIDS dem entia complex. Glial activation and elevated pro-inflammation cytokines (signs of neuroinflammation) in the spinal cord have been widely recognized as a pivotal mechanism underlying the development and maintenance of many types of pathological pain. The role of GSK3β in the pathogenesis of pain has recently emerged. In this review, we will first review the GSK3β structure, regulation, and mechanisms by which GSK3βregulates inflammation. We will then describe neuroinflammationin general and in specific types of neurological diseases and the potential beneficial effects induced by inhibiting GSK3β. Finally, we will provide new evidence linking aberrant levels of GSK3β in the development of pathological pain.

Spinocerebellar ataxia type 3 (SCA3) is an autosomal dominant neurodegenerative disorder caused by the expansion of a CAG trinucelotide repeat that encodes an abnormal polyglutamine (PolyQ) tract in the disease protein, ataxin-3. The formation of neuronal intranuclear inclusions in the specific brain regions is one of the pathological hallmarks of SCA3. Acceleration of the degradation of the mutant protein aggregates is proven to produce beneficial effects in SCA3 and other PolyQ diseases. Lithium is known to be neuroprotective in various models of neurodegenerative disease and can reduce the mutant protein aggregates by inducing autophagy. In this study, we explored the therapeutic potential of lithium in a SCA3 Drosophila model. We showed that chronic treatment with lithium chloride at specific doses notably prevented eye depigmentation, alleviated locomotor disability, and extended the median life spans of SCA3 transgenic Drosophila. By means of genetic approaches, we showed that co-expressing the mutant S9E, which mimicked the phosphorylated S9 state of Shaggy as done by lithium, also partly decreased toxicity of gmr-SCA3tr-Q78. Taken together, our findings suggest that lithium is a promising therapeutic agent for the treatment of SCA3 and other PolyQ diseases.

Rett syndrome (RTT) is a neurodevelopmental disorder caused by mutations in the methyl CpG binding protein 2 (MECP2) gene. There are currently no approved treatments for RTT. The expression of K⁺/Cl^- cotransporter 2 (KCC2), a neuron-specific protein, has been found to be reduced in human RTT neurons and in RTT mouse models, suggesting that KCC2 might play a role in the pathophysiology of RTT. To develop neuron-based high-throughput screening (HTS) assays to identify chemical compounds that enhance the expression of the KCC2 gene, we report the generation of a robust high-throughput drug screening platform that allows for the rapid assessment of KCC2 gene expression in genome-edited human reporter neurons. From an unbiased screen of more than 900 small-molecule chemicals, we have identified a group of compounds that enhance KCC2 expression termed KCC2 expression-enhancing compounds (KEECs). The identified KEECs include U.S. Food and Drug Administration-approved drugs that are inhibitors of the fms-like tyrosine kinase 3 (FLT3) or glycogen synthase kinase 3β (GSK3β) pathways and activators of the sirtuin 1 (SIRT1) and transient receptor potential cation channel subfamily V member 1 (TRPV1) pathways. Treatment with hit compounds increased KCC2 expression in human wild-type (WT) and isogenic MECP2 mutant RTT neurons, and rescued electrophysiological and morphological abnormalities of RTT neurons. Injection of KEEC KW-2449 or piperine in Mecp2 mutant mice ameliorated disease-associated respiratory and locomotion phenotypes. The small-molecule compounds described in our study may have therapeutic effects not only in RTT but also in other neurological disorders involving dysregulation of KCC2.

Lycium barbarum polysaccharide (LBP), an antioxidant from wolfberry, displays the antioxidative and anti-inflammatory effects on experimental models of insulin resistance in vivo. However, the effective mechanism of LBP on high-fat diet-induced insulin resistance is still unknown. The objective of the study was to investigate the mechanism involved in LBP-mediated phosphatidylinositol 3-kinase (PI3K)/AKT/Nrf2 axis against high-fat-induced insulin resistance. HepG2 cells were incubated with LBP for 12 hrs in the presence of palmitate. C57BL/6J mice were fed a high-fat diet supplemented with LBP for 24 weeks. We analyzed the expression of nuclear factor-E2-related factor 2 (Nrf2), Jun N-terminal kinases (JNK), and glycogen synthase kinase 3β (GSK3β) involved in insulin signaling pathway in vivo and in vitro. First, LBP significantly induced phosphorylation of Nrf2 through PI3K/AKT signaling. Second, LBP obviously increased detoxification and antioxidant enzymes expression and reduced reactive oxygen species (ROS) levels via PI3K/AKT/Nrf2 axis. Third, LBP also regulated phosphorylation levels of GSK3β and JNK through PI3K/AKT signaling. Finally, LBP significantly reversed glycolytic and gluconeogenic genes expression via the activation of Nrf2-mediated cytoprotective effects. In summary, LBP is novel antioxidant against insulin resistance induced by high-fat diet via activation of PI3K/AKT/Nrf2 pathway.

β-N-methylamino-l-alanine (L-BMAA) is a neurotoxic amino acid that has been related to various neurodegenerative diseases. The aim of this work was to analyze the biotoxicity produced by L-BMAA in vivo in rats, trying to elucidate its physiopathological mechanisms and to search for analogies between the found effects and pathologies like Amyotrophic Lateral Sclerosis (ALS). Our data demonstrated that the neurotoxic effects in vivo were dosage-dependent. For evaluating the state of the animals, a neurological evaluation scale was developed as well as a set of functional tests. Ultrastructural cell analysis of spinal motoneurons has revealed alterations both in endoplasmic reticulum and mitochondria. Since GSK3β could play a role in some neuropathological processes, we analyzed the alterations occurring in GSK3β levels in L-BMAA treated rats, we have observed an increase in the active form of GSK3β levels in lumbar spinal cord and motor cerebral cortex. On the other hand, (TAR)-DNA-binding protein 43 (TDP-43) increased in L-BMAA treated animals. Our results indicated that N-acetylaspartate (NAA) declined in animals treated with L-BMAA, and the ratio of N-acetylaspartate/choline (NAA/Cho), N-acetylaspartate/creatine (NAA/Cr) and N-acetylaspartate/choline+creatine (NAA/Cho+Cr) tended to decrease in lumbar spinal cord and motor cortex. This project offers some encouraging results that could help establishing the progress in the development of an animal model of sporadic ALS and L-BMAA could be a useful tool for this purpose.

Glucocorticoids (GCs) regulate energy supply in response to stress by increasing hepatic gluconeogenesis during fasting. Long-term GC treatment induces hepatic steatosis and weight gain. GC signaling is coordinated via the GC receptor (GR) GRα, as the GRβ isoform lacks a ligand-binding domain. The roles of the GR isoforms in the regulation of lipid accumulation is unknown. The purpose of this study was to determine whether GRβ inhibits the actions of GCs in the liver, or enhances hepatic lipid accumulation. We show that GRβ expression is increased in adipose and liver tissues in obese high-fat fed mice. Adenovirus-mediated delivery of hepatic GRβ overexpression (GRβ-Ad) resulted in suppression of gluconeogenic genes and hyperglycemia in mice on a regular diet. Furthermore, GRβ-Ad mice had increased hepatic lipid accumulation and serum triglyceride levels possibly due to the activation of NF-κB signaling and increased tumor necrosis factor α (TNFα) and inducible nitric-oxide synthase expression, indicative of enhanced M1 macrophages and the development of steatosis. Consequently, GRβ-Ad mice had increased glycogen synthase kinase 3β (GSK3β) activity and reduced hepatic PPARα and fibroblast growth factor 21 (FGF21) expression and lower serum FGF21 levels, which are two proteins known to increase during fasting to enhance the burning of fat by activating the β-oxidation pathway. In conclusion, GRβ antagonizes the GC-induced signaling during fasting via GRα and the PPARα-FGF21 axis that reduces fat burning. Furthermore, hepatic GRβ increases inflammation, which leads to hepatic lipid accumulation.

UNASSIGNED

To resolve the controversy as to whether periostin plays a role in myocardial regeneration after myocardial infarction (MI), we created a neonatal mouse model of MI to investigate the influence of periostin ablation on myocardial regeneration and clarify the underlying mechanisms.

UNASSIGNED

Neonatal periostin-knockout mice and their wildtype littermates were subjected to MI or sham surgery. In the wildtype mice after MI, fibrosis was detectable at 3 days and fibrotic tissue was completely replaced by regenerated myocardium at 21 days. In contrast, in the knockout mice, significant fibrosis in the infarcted area was present at even 3 weeks after MI. Levels of phosphorylated-histone 3 and aurora B in the myocardium, detected by immunofluorescence and western blotting, were significantly lower in knockout than in wildtype mice at 7 days after MI. Similarly, angiogenesis was decreased in the knockout mice after MI. Expression of both the endothelial marker CD-31 and α-smooth muscle actin was markedly lower in the knockout than in wildtype mice at 7 days after MI. The knockout MI group had elevated levels of glycogen synthase kinase (GSK) 3β and decreased phosphatidylinositol 3-kinase (PI3K), phosphorylated serine/threonine protein kinase B (p-Akt), and cyclin D1, compared with the wildtype MI group. Similar effects were observed in experiments using cultured cardiomyocytes from neonatal wildtype or periostin knockout mice. Administration of SB216763, a GSK3β inhibitor, to knockout neonatal mice decreased myocardial fibrosis and increased angiogenesis in the infarcted area after MI.

UNASSIGNED

Ablation of periostin suppresses post-infarction myocardial regeneration by inhibiting the PI3K/GSK3β/cyclin D1 signalling pathway, indicating that periostin is essential for myocardial regeneration.

The devastating prognosis of pancreatic ductal adenocarcinoma (PDAC) is partially attributed to chemotherapy resistance. Glutathione peroxidase-1 (GPx1) plays various roles in the development and progression of multiple tumors, with the exception of pancreatic cancer. Here, we tentatively explored the role of GPx1 in the malignant biological behavior and gemcitabine (GEM) resistance of PDAC. GPx1 levels were detected using tissue microarrays and were negatively correlated with the overall survival of patients with PDAC. GPx1 silencing induced a mesenchymal transition phenotype and increased GEM resistance in vitro and in vivo. Additionally, the activation of reactive oxygen species (ROS)-mediated Akt/glycogen synthase kinase 3β (GSK3β)/Snail signaling was involved in this process, as determined by RNA sequencing. Moreover, low GPx1 expression correlated with a worse survival rate in patients with PDAC who received GEM adjuvant chemotherapy, whereas this correlation was not detected in patients receiving fluoropyrimidine. Based on our results, GPx1 inhibits the epithelial-mesenchymal transition (EMT) and chemoresistance by regulating the Akt/GSK3β/Snail signaling axis in PDAC. Furthermore, GPx1 may be a potential predictive biomarker in GEM-treated PDAC patients.

The tumor microenvironment induces endoplasmic reticulum (ER) stress in tumor cells, an event that can promote progression, but it is unknown how tumor cells adapt to this stress. In this study, we show that the fibroblast growth factor FGF19, a gene frequently amplified in hepatocellular carcinoma (HCC), facilitates a survival response to ER stress. Levels of FGF19 expression were increased in stressed HCC cells in culture and in a mouse xenograft model. Induction of ER stress required the transcription factor ATF4, which directly bound the FGF19 promoter. In cells where ER stress was induced, FGF19 overexpression promoted HCC cell survival and increased resistance to apoptosis, whereas FGF19 silencing counteracted these effects. Mechanistic investigations implicated glycogen synthase kinase-3β (GSK3β) in regulating nuclear accumulation of the stress-regulated transcription factor Nrf2 activated by FGF19. Our findings show how FGF19 provides a cytoprotective role against ER stress by activating a FGFR4-GSK3β-Nrf2 signaling cascade, with implications for targeting this signaling node as a candidate therapeutic regimen for HCC management. Cancer Res; 77(22); 6215-25. ©2017 AACR.

The malignant neoplasm of the cervix, cervical cancer, has effects on the reproductive tract. Although infection with oncogenic human papillomavirus is essential for cervical cancer development, it alone is insufficient to explain the development of cervical cancer. Therefore, other risk factors such as host genetic factors should be identified, and their importance in cervical cancer induction should be determined. Although gene expression profiling studies in the last decade have made significant molecular findings about cervical cancer, adequate screening and effective treatment strategies have yet to be achieved. In the current study, meta-analysis was performed on cervical cancer-associated transcriptome data and reporter biomolecules were identified at RNA (mRNA, miRNA), protein (receptor, transcription factor, etc.), and metabolite levels by the integration of gene expression profiles with genome-scale biomolecular networks. This approach revealed already-known biomarkers, tumor suppressors and oncogenes in cervical cancer as well as various receptors (e.g. ephrin receptors EPHA4, EPHA5, and EPHB2; endothelin receptors EDNRA and EDNRB; nuclear receptors NCOA3, NR2C1, and NR2C2), miRNAs (e.g., miR-192-5p, miR-193b-3p, and miR-215-5p), transcription factors (particularly E2F4, ETS1, and CUTL1), other proteins (e.g., KAT2B, PARP1, CDK1, GSK3B, WNK1, and CRYAB), and metabolites (particularly, arachidonic acids) as novel biomarker candidates and potential therapeutic targets. The differential expression profiles of all reporter biomolecules were cross-validated in independent RNA-Seq and miRNA-Seq datasets, and the prognostic power of several reporter biomolecules, including KAT2B, PCNA, CD86, miR-192-5p and miR-215-5p was also demonstrated. In this study, we reported valuable data for further experimental and clinical efforts, because the proposed biomolecules have significant potential as systems biomarkers for screening or therapeutic purposes in cervical carcinoma.

The mechanisms of how signaling pathways are coordinated and integrated for the maintenance of the self-renewal of human embryonic stem cells (hESCs) and the acquisition of pluripotency in reprogramming are still only partly understood. CDK1 is a key regulator of mitosis. Recently, CDK1 has been shown to be involved in regulating self-renewal of stem cells, even though the mechanistic role of how CDK1 regulates pluripotency is unknown. In this report, we aim to understand how CDK1 can control pluripotency by reducing CDK1 activity to a level that has no effect on cell cycle progression. We demonstrated that high levels of CDK1 is associated with the pluripotency stage of hESCs; and decreased CDK1 activity to a level without perturbing the cell cycle is sufficient to induce differentiation. CDK1 specifically targets the phosphorylation of PDK1 and consequently the activity of PI3K/Akt and its effectors ERK and GSK3β. Evidence of the reversion of inactive CDK1-mediated differentiation by the inhibition of Akt signaling effectors suggests that the CDK1-PDK1-PI3K/Akt kinase cascade is a functional signaling pathway for the pluripotency of hESCs. Moreover, cyclin B1-CDK1 complexes promote somatic reprogramming efficiency, probably by regulating the maturation of induced pluripotent stem cells (iPSCs), as cyclin B1 stimulates a higher cellular level of LIN28A, suggesting that monitoring iPSC factors could be a new path for the enhancement of reprogramming efficiency. Together, we demonstrate an essential role for the CDK1-PDK1-PI3K/Akt kinase signaling pathway in the regulation of self-renewal, differentiation, and somatic reprogramming, which provides a novel kinase cascade mechanism for pluripotency control and acquisition.

Purpose Fucoidan, a complex, sulfated polysaccharide obtained from brown seaweed, exerted anticancer activity through the down-regulation of β-catenin signaling in mouse breast cancer cells in our previous study. This study examines the anti-cancer effects of fucoidan as well as its underlying molecular mechanisms in the human triple negative breast cancer (TNBC) cell line and in 7,12-dimethylbenz[a]anthracene (DMBA)-induced experimental mammary carcinogenesis in rats. Methods in vitro studies, fluorescent staining, flow cytometry and Western blotting were performed to analyze apoptosis and protein expression in human breast cancer MDA-MB-231 cells. In vivo intervention experiments were conducted with Sprague Dawley (SD) rats with DMBA-induced breast cancer. Tumor volumes and weights were measured. Results in vitro fucoidan treatment inhibited proliferation and induced apoptosis in MDA-MB-231 cells. Western blotting detected that Cyt C and Smac were released into the cell cytoplasm and that caspase-3 and caspase-9 were activated in MDA-MB-231 cells. The levels of AIF and EndoG were significantly increased in the cytoplasm and in the nuclei by fucoidan. These data show that fucoidan induced caspase-dependent and caspase-independent apoptosis. Moreover, fucoidan treatment down-regulated the expression of Bid, Bcl-2 and Bcl-xl and up-regulated the level of Bax. In vivo, fucoidan supplementation decreased the mean tumor weight.

CONCLUSIONS

Results from the in vivo and in vitro experiments both showed that fucoidan decreased the levels of p-PI3K, p-AKT and p-GSK-3β (Ser9) in breast cancer. The level of β-catenin was also decreased. These results suggest that fucoidan can inhibit MDA-MB-231 human breast cancer cells and DMBA-induced tumors in rats by down-regulating the PI3 K/AKT/GSK3β pathway. This study provides experimental evidence that elucidates the mechanism of antitumor effect of fucoidan and clarifies the mechanism of the effect of fucoidan on the regulation of β-catenin.

H19 is involved in tumor metastasis and associated with tumor progression. Enhancer of zest homolog 2 (EZH2) is overexpressed in multiple cancer types and correlates with tumor proliferation, epithelial-mesenchymal transition, and poor prognosis. However, the interaction between H19 and EZH2 to promote tongue squamous cell carcinoma (TSCC) progression remains largely uncharacterized. Insitu hybridization and quantitative reverse-transcription PCR (qRT-PCR) were performed to measure H19 expression in primary TSCC and adjacent normal tissues and cell lines. EZH2 expression was determined by immunohistochemistry in matched primary TSCC and adjacent normal tissues. The correlation between H19 and EZH2 expression and clinicopathological characteristics were analyzed. The roles of H19 in cell proliferation, apoptosis, and invasion were analyzed using a H19-targeted lentivirus. Western blot and qRT-PCR were carried out to detect downstream signal pathway changes. Expression levels of downstream signaling proteins in primary TSCC tissues and adjacent normal tissues were analyzed by immunohistochemistry. H19 and EZH2 were upregulated in TSCC tissues compared to matched normal tissues, and significantly correlated with WHO grade, lymph node metastasis, and poor prognosis. H19 silencing attenuated cell proliferation, apoptosis, and invasion in vitro. H19 knockdown inhibited the activation of β-catenin/GSK-3β/cyclin D1/c-myc, upregulated E-cadherin and zonula occludens-1 (ZO-1), and inhibited N-cadherin, vimentin, Snail1, Twist1, and ZEB1. Silencing H19 expression also inhibited tumor progression and lung metastasis in an animal model. Our findings indicate that H19 promotes TSCC progression through association with EZH2, and affects downstream β-Catenin/GSK3β/EMT signaling, suggesting that H19 inhibition might be a potential target for the treatment of TSCC.

The efficacy of antihypertensive agents in Alzheimer's disease (AD) is controversial. It has been tested here whether some antihypertensive drugs might influence AD through mechanisms independent of blood pressure-lowering activity. The effects of treatment with the antihypertensive propranolol on cognition and AD-related markers have been studied in the Tg2576 mouse model of AD. Propranolol, at a lower dose than that used as antihypertensive (5 mg/kg, 6 wk), attenuated cognitive impairments shown by Tg2576 mice aged 9 months in the novel object recognition and fear conditioning tests. Propranolol was also able to counteract the increases in hippocampal levels of Aβ(42) present in Tg2576 mice. This effect was accompanied by an increased expression of insulin degrading enzyme. Changes in markers of synaptic pathology, as shown by decreases in phosphorylation of Akt and in the expression of BDNF in Tg2676 mice, were also counteracted by propranolol treatment. Tau hyperphosphorylation shown by Tg2576 mice was also decreased in the hippocampus of propranolol-treated mice, an effect probably related to an increase of GSK3β phosphorylation (inactive form) and a decreased JNK1 expression. Overall, these data further strengthen the potential of propranolol as a therapeutic agent for AD.