Efficient nitrogen removal from municipal wastewater using an integrated fixed-film activated sludge process in a novel air-lifting loop reactor: A pilot-scale demonstration.

A novel air-lifting loop reactor combines anoxic, oxic, and settling zones to achieve organic and nutrient removal, as well as solid-liquid separation. To address sludge settling ability and operation stability issues caused by low dissolved oxygen in aerobic zones, this study proposes using modified polypropylene carriers to establish a fixed-film activated sludge (IFAS) system. A pilot-scale demonstration of the IFAS-based air-lifting loop reactor is conducted, and the results show successful operation for approximately 300 days. The pilot-scale reactor achieves a maximum aerobic granulation ratio of 16% in the bulk liquid. The IFAS system contributes to efficient removal of organic matter (96%) and nitrogen (94%) by facilitating simultaneous nitrification and denitrification, as well as fast solid-liquid separation with a low sludge volume index of 34 mL/g. Microbial analysis reveals enrichment of functional bacteria involved in nitrification, denitrification, and flocculation throughout the operation process.

Effects of Iron-Peptides Chelate Nanoliposomes on Iron Supplementation in Rats.

The objective of this study was to investigate the effects of iron nanoliposomes on iron supplementation and toxicity in SD rats induced by a low-iron diet. The size and infrared spectroscopy of a liposomal oral delivery system were investigated. The particle size of nanoliposomes embedded with chelates was increased. Infrared spectra proved that peptides-iron and blank nanoliposomes were bonded by interaction forces, including the fracture of hydrogen bonds, C = C bonds, hydrophobic interaction, and C-N bonds. We found that iron supplementation chelates had a certain protective effect on viscera after being embedded by nanoliposomes. After 10 days of treatment, the concentration of hemoglobin could be gradually increased. Nanoliposome encapsulated peptides-iron has a better effect than other groups. At the same time, SOD, MDA, and CAT reached normal levels after 20 days. Histological results showed that the sections of the nanoliposomes groups were clearer than those of the other groups. There was a little inflammation in the liver without obvious pathological changes, which also proved that the iron chelates embedded by nanoliposomes had no obvious side effects on iron supplementation in rats. Nanoliposome encapsulated peptides-iron has a small side effect and a significant curative effect of iron supplementation. It maybe has a good application prospect in the clinical medical field.

Knockdown of miR-150-5p reduces hypoxia-induced autophagy and epithelial-mesenchymal transition of endometriotic cells via regulating the PDCD4/NF-κB signaling pathway.

BACKGROUND: Hypoxia is an important microenvironmental factor that induces Endometriosis (EMs), but its mechanism remains unclear. Our study aims to investigate the mechanisms of miR-150-5p on hypoxia-induced EMs. METHODS: Ovarian endometriosis cyst wall stromal cell lines CRL-7566 cells were treated with hypoxia. Cell migration ability was measured by Transwell assay. qRT-PCR was performed to detect miR-150-5p and PDCD4 expression. The autophagy-related proteins (LC3-I, LC3-II, Beclin-1, and p62), epithelial-mesenchymal transition (EMT) related proteins (E-cadherin, N-cadherin, and Vimentin) and NF-κB signaling pathway related proteins p65 expression were measured by western blot. Dual-luciferase reporter gene assay verified the binding relationship between miR-150-5p and PDCD4. RESULTS: After hypoxia treatment, the miR-150-5p expression was up-regulated in CRL-7566 cells, while the expression of PDCD4 was down-regulated. In CRL-7566 cells, autophagy, migration and EMT were increased after hypoxia treatment. The autophagy inhibitor 3-MA inhibited hypoxia-induced the autophagy, migration and EMT of CRL-7566 cells. Hypoxia-induced autophagy and EMT of CRL-7566 cells were inhibited after knocking down miR-150-5p. Then miR-150-5p negatively regulated PDCD4 expression. PDCD4 knockdown reversed the inhibitory effect of miR-150-5p silencing on hypoxia-induced autophagy and EMT of CRL-7566 cells. Inhibiting the NF-κB signaling pathway weakened the effect of PDCD4 knockdown on hypoxia-induced autophagy and EMT of CRL-7566 cells. CONCLUSION: MiR-150-5p silencing inhibited hypoxia-induced autophagy and EMT of endometriotic cells by regulating the PDCD4/NF-κB signaling pathway.

Network Pharmacology and Molecular Docking Approach to Reveal the Immunotherapeutic Mechanism of Cuscutae Semen in Treating Thin Endometrium.

Objective: Thin endometrium is considered as a leading cause of infertility, recurrent pregnancy loss, and repeated implantation failure. The seed of Cuscutae Semen (CS) has been used to prevent aging and improve sexual function in Traditional Chinese Medicine. However, the pharmacological mechanism of CS in preventing and treating thin endometrium remains to be elucidated. Methods: Three public databases, TCMSP, GeneCards, and OMIM, were searched to collect the main active compounds and putative molecules of CS, as well as the targets of thin endometrium, respectively. The CS and thin endometrium common targets were subject to protein-protein interaction (PPI) analysis followed by functional enrichment analysis. The best binding mode of CS compounds and common target proteins was evaluated by molecular docking and analysis in the AutoDockTools. Results: In total, 11 main active compounds, 102 drug target proteins, and 70 CS and thin endometrium common targets were identified. There were 68 nodes with 722 edges in the PPI network; HIF1A, MYC, ESR1, and EGFR were the top 4 targets. After functional enrichment analysis, it was revealed that the therapeutic effects of active compounds of CS on thin endometrium were achieved through cellular response to chemical stress, transcription regulator, DNA-binding transcription factor binding, chemical carcinogenesis-receptor activation, lipid, and atherosclerosis. The molecular docking analysis revealed that the 3 active compounds of CS, quercetin, matrine, and isorhamnetin, have good binding ability with their targets, HIF1A, MYC, ESR1, and EGFR. Conclusion: Our study uncovers the main active compounds in CS and their corresponding targets related to thin endometrium which explains the pharmacological mechanism underlying therapeutic effects of CS on thin endometrium.

lncRNA EGFR-AS1 facilitates leiomyosarcoma progression and immune escape via the EGFR-MYC-PD-L1 axis.

AIM: this study aimed to investigate the role of long non-coding RNA (lncRNA) epidermal growth factor receptor antisense RNA 1 (EGFR-AS1), an antisense transcript of EGFR, in leiomyosarcoma (LMS) and the underlying mechanisms. METHODS: levels of EGFR-AS1 and programmed death ligand 1 (PD-L1) were measured in LMS tissues and cell lines using quantitative real-time PCR (qRT-PCR), as well as western blotting and/or immunohistochemical staining; flow cytometry was employed to validate the role of EGFR-AS1 in altering the activity of CD8+ T cells; interaction of EGFR-AS1 and EGFR was determined by fluorescent in situ hybridization (FISH) and RNA pull-down; regulation of MYC on the PD-L1 promoter was assessed by chromatin immunoprecipitation (ChIP); a xenograft in vivo tumor growth assay was applied to verify the EGFR-AS1/EGFR/MYC/PD-L1 axis in vivo. RESULTS: up-regulation of EGFR-AS1 and PD-L1 in LMS tissues was negatively correlated with CD8+ T-cell infiltration; EGFR-AS1 positively regulated PD-L1, thereby strengthening interaction of LMS cells and CD8+ T cells and triggering CD8+ T cell apoptosis via the PD-1/PD-L1 checkpoint; EGFR-AS1 co-localized and interacted with EGFR to promote MYC activity; MYC was identified as a transcriptional activator of PD-L1. CONCLUSION: lncRNA EGFR-AS1 was demonstrated to increase PD-L1 expression through the EGFR/MYC pathway in LMS cells, thereby repressing T-cell infiltration and contributing to immune escape.

Physicochemical, digestive and rheological properties of protein from tuna by subcritical dimethyl ether: Focus on process-related indexes.

The process-related physicochemical, digestive and rheological properties of protein prepared by subcritical dimethyl ether extraction (SDEE) were comprehensively investigated and compared with those obtained by pH-shift, to study the industrial potential of SDEE. Two different materials from tuna (meat and liver) were studied in parallel, and SDEE had similar effects on the proteins in them. The protein component was almost unchanged before and after SDEE, while the content of water-soluble protein and alkali-soluble protein was substantially reduced and increased after pH-shift, respectively. We also found that SDEE had superior ability to pH-shift to conserve light metals, remove lipids and heavy metals, and maintain protein structure. Furthermore, SDEE-produced protein powders were easier for humans to digest, and their gelation and emulsification were also superior to those prepared by pH-shift. The aforementioned results suggest that SDEE can remove more impurities, and the obtained protein has outstanding potential in industrial applications.

Mechanism and inhibition kinetics of peptide P13 as thrombin inhibitor.

Excessive coagulation can easily lead to arterial and venous thrombosis, which is the main reason for the evolution of myocardial infarction and cerebrovascular accidents. As a key coagulation factor for the coagulation pathway, thrombin has become a remarkable target for the control of thrombosis. The synthesized peptide P13 with amino acid sequence of N-RGDAGFAGDDAPR was expected to be an inhibitor with higher antithrombotic activity. The results showed that the IC50 (50% inhibition of thrombin activity) of the peptide P13 was determined by colorimetric method to be 115 µM. And enzyme kinetic experiments showed that P13 was a competitive inhibitor of thrombin with Ki = 106 µM. Fluorescence spectra and three-dimensional fluorescence showed that P13 could alter the secondary structure of thrombin and the microenvironment of certain chromogenic amino acids. P13 can spontaneously bind with thrombin exosite 1 in the form of 1:1 mainly through hydrogen bonding and van der Waals force. And the optimal docking mode of P13 and thrombin was revealed by molecular docking with "-CDOCKER_Energy" of 178.679 kcal mol-1. This study revealed P13 may become a potential anticoagulant drug widely used after further studies in preclinical and clinical trials.

Optimization of ultrasonic-assisted enzymatic extraction of polysaccharides from thick-shell mussel (Mytilus coruscus) and their antioxidant activities.

This study aimed to obtain the purified fractions of Mytilus coruscus polysaccharides (MCPs) and investigate their antioxidant activities. MCPs were prepared through ultrasonic-assisted enzymatic extraction optimized by employing the response surface methodology. A single-factor experiment was conducted using the Box-Behnken design to determine the optimum extraction conditions of MCPs. The ultrasonic power was 60 W, liquid-to-material ratio was 30 mL/g, extraction time was 36 min, extraction temperature was 64 °C, enzyme concentration was 3.2%, and polysaccharide extraction yield was 12.86% ±â€¯0.12%. A novel polysaccharide (MCP1-2) was obtained after the purification with AB-8 macroporous resin, DEAE Sepharose Fast Flow, and Sepharose CL-6B column. The molecular weight of MCP1-2 was estimated to be 134.9 kDa according to high-performance gel permeation chromatography. High-pressure liquid-phase chromatography results showed that MCP1-2 contained mannose, rhamnose, glucuronic acid, glucose, galactose, and L-Fuc at a molar ratio of 1.53:1:4.83:81.82:2.36:1.51. Infrared and NMR spectroscopies confirmed that MCP1-2 possessed α- and ß- configurations. The antioxidant activities of MCP1-2 were investigated in vitro, and the results showed that MCP1-2 had good antioxidant activity and can be used as a natural antioxidant in food.

Purification of an iron-binding peptide from scad (Decapterus maruadsi) processing by-products and its effects on iron absorption by Caco-2 cells.

This work was aimed at producing peptides containing iron-binding capabilities from scad (Decapterus maruadsi) processing by-product with alcalase hydrolysis. The chelating peptides were purified by ultrafiltration, immobilized-metal affinity chromatography, gel filtration chromatography, and reversed-phase high-performance liquid chromatography. A novel iron-binding peptide was purified with 1,386.63 Da molecular weight and amino acid sequence of QKGTYDDYVEGL. The peptide binds to iron mainly through carboxyl and hydroxyl oxygen bonds. The iron-binding peptide can significantly promote the absorption of inorganic iron in Caco-2 cells. These results have contributed to development of the peptide from scad processing by-products hydrolyzate in iron supplementations. PRACTICAL APPLICATIONS: Iron deficiency is one of the most common and widespread nutritional disorders in the world. Iron-peptide chelates may be suitable for iron-fortification. Our study shows that a peptide purified from scad processing by-product has iron-chelating activity, and significantly increases iron absorption by Caco-2 cells. Hence, this peptide has potential application as a novel carrier for enhancing iron absorption.

Effects of elevated ultraviolet-B radiation on root growth and chemical signaling molecules in plants.

Ozone layer depletion leads to elevated ultraviolet-B (UV-B) radiation, which affects plant growth; however, little is known about the relationship between root growth and signaling molecules in roots. Therefore, in this work, simulated UV-B radiation was used to study the effects of elevated UV-B radiation on root growth of soybean seedlings and changes in the content of signaling molecules in roots. The results showed that compared with the control, the 2.63 kJ m-2 d-1 and 6.17 kJ m-2 d-1 elevated UV-B radiation treatments inhibited root growth, and root growth parameters (total root length, root surface area, root volume, average diameter, root tip number, and root dry weight) all decreased. For root signaling molecules, the content of nitric oxide, reactive oxygen species, abscisic acid, salicylic acid, and jasmonic acid increased, and the content of auxin, cytokinin, and gibberellin decreased. The above indices changed more significantly under the 6.17 kJ m-2 d-1 treatment. After withdrawal of the exposure, the above indices could be restored to a certain extent. These data indicated that UV-B radiation interfered with root growth by affecting the content of signaling molecules in roots, and the degree of the effects was related to the intensity of UV-B radiation. The results from this study provide a theoretical basis for studying the preliminary mechanism of elevated UV-B radiation on root growth and possible pathways that can mitigate UV-B radiation damage for root growth. ONE SENTENCE SUMMARY: The effects of elevated UV-B on root growth of soybean seedlings were regulated by signaling molecules, and the degree of the effects was related to the intensity of UV-B radiation.

Casticin inhibits breast cancer cell migration and invasion by down-regulation of PI3K/Akt signaling pathway.

Casticin is one of the major active components isolated from Fructus viticis Increasing studies have revealed that casticin has potential anticancer activity in various cancer cells, but its effects on breast cancer cell migration and invasion are still not well known. Therefore, the ability of cell migration and invasion in the breast cancer MDA-MB-231 and 4T1 cells treated by casticin was investigated. The results indicated that casticin significantly inhibited cell migration and invasion in the cells exposed to 0.25 and 0.50 µM of casticin for 24 h. Casticin treatment reduced matrix metalloproteinase (MMP) 9 (MMP-9) activity and down-regulated MMP-9 mRNA and protein expression, but not MMP-2. Casticin treatment suppressed the nuclear translocation of transcription factors c-Jun and c-Fos, but not nuclear factor-κB (NF-κB), and decreased the phosphorylated level of Akt (p-Akt). Additionally, the transfection of Akt overexpression vector to MDA-MB-231 and 4T1 cells could up-regulate MMP-9 expression concomitantly with a marked increase in cell invasion, but casticin treatment reduced Akt, p-Akt, and MMP-9 protein levels and inhibited the ability of cell invasion in breast cancer cells. Additionally, casticin attenuated lung metastasis of mouse 4T1 breast cancer cells in the mice and down-regulated MMP-9 expression in the lung tissues of mice treated by casticin. These findings suggest that MMP-9 expression suppression by casticin may act through inhibition of the phosphatidylinositol 3-kinase (PI3K)/Akt signaling pathway, which in turn results in the inhibitory effects of casticin on cell migration and invasion in breast cancer cells. Therefore, casticin may have potential for use in the treatment of breast cancer invasion and metastasis.

Protein Hydrolysates as Promoters of Non-Haem Iron Absorption.

Iron (Fe) is an essential micronutrient for human growth and health. Organic iron is an excellent iron supplement due to its bioavailability. Both amino acids and peptides improve iron bioavailability and absorption and are therefore valuable components of iron supplements. This review focuses on protein hydrolysates as potential promoters of iron absorption. The ability of protein hydrolysates to chelate iron is thought to be a key attribute for the promotion of iron absorption. Iron-chelatable protein hydrolysates are categorized by their absorption forms: amino acids, di- and tri-peptides and polypeptides. Their structural characteristics, including their size and amino acid sequence, as well as the presence of special amino acids, influence their iron chelation abilities and bioavailabilities. Protein hydrolysates promote iron absorption by keeping iron soluble, reducing ferric iron to ferrous iron, and promoting transport across cell membranes into the gut. We also discuss the use and relative merits of protein hydrolysates as iron supplements.

Downregulation of glypican-3 expression increases migration, invasion, and tumorigenicity of human ovarian cancer cells.

Glypican-3 (GPC3) is a membrane of heparan sulfate proteoglycan family involved in cell proliferation, adhesion, migration, invasion, and differentiation during the development of the majority of mesodermal tissues and organs. GPC3 is explored as a potential biomarker for hepatocellular carcinoma screening. However, as a tumor-associated antigen, its role in ovarian cancer remains elusive. In this report, the expression levels of GPC3 in the various ovarian cancer cells were determined with quantitative reverse transcription-polymerase chain reaction (qRT-PCR), and GPC3 expression in ovarian cancer UCI 101 and A2780 cells was knocked down by siRNA transfection, and the effects of GPC3 knockdown on in vitro cell proliferation, migration, and invasion were respectively analyzed by 3-[4, 5-dimethylthiazol-2-yl]-2, 5-diphenyltetrazolium bromide (MTT) assay and Transwell migration assay. Additionally, the effect of GPC3 knockdown on in vivo tumorigenesis were investigated in athymic nude mice. The results indicated that GPC3 knockdown significantly promoted cell proliferation and increased cell migration and invasion by upregulation of matrix metalloproteinase (MMP)-2 and MMP-9 expression and downregulation of tissue inhibitor of metalloproteinase-1 expression. Additionally, GPC3 knockdown also increased in vivo tumorigenicity of UCI 101 and A2780 cells and final tumor weights and volumes after subcutaneous cell injection in the nude mice. The results of immunohistochemical staining and Western blotting both demonstrated a lower expression of GPC3 antigen in the tumors of GPC3 knockdown groups than that of negative control groups. Moreover, transforming growth factor-ß2 protein expression in the tumors of GPC3 knockdown groups was significantly increased, which at least contributed to tumor growth in the nude mice. Taken together, these findings suggest that GPC3 knockdown promotes the progression of human ovarian cancer cells by increasing their migration, invasion, and tumorigenicity, and suggest that GPC3 is a potential therapeutic target for ovarian cancer patients.

Combined effects of Lanthanum(III) and elevated Ultraviolet-B radiation on root nitrogen nutrient in soybean seedlings.

Rare earth element pollution and elevated ultraviolet-B (UV-B) radiation occur simultaneously in some regions, but the combined effects of these two factors on plants have not attracted enough attention. Nitrogen nutrient is vital to plant growth. In this study, the combined effects of lanthanum(III) and elevated UV-B radiation on nitrate reduction and ammonia assimilation in soybean (Glycine max L.) roots were investigated. Treatment with 0.08 mmol L(-1) La(III) did not change the effects of elevated UV-B radiation on nitrate reductase (NR), nitrite reductase (NiR), glutamine synthetase (GS), glutamate synthase (GOGAT), glutamate dehydrogenase (GDH), nitrate, ammonium, amino acids, or soluble protein in the roots. Treatment with 0.24 mmol L(-1) La(III) and elevated UV-B radiation synergistically decreased the NR, NiR, GS, and GOGAT activities as well as the nitrate, amino acid, and soluble protein levels, except for the GDH activity and ammonium content. Combined treatment with 1.20 mmol L(-1) La(III) and elevated UV-B radiation produced severely deleterious effects on all test indices, and these effects were stronger than those induced by La(III) or elevated UV-B radiation treatment alone. Following the withdrawal of La(III) and elevated UV-B radiation, all test indices for the combined treatments with 0.08/0.24 mmol L(-1) La(III) and elevated UV-B radiation recovered to a certain extent, but they could not recover for treatments with 1.20 mmol L(-1) La(III) and elevated UV-B radiation. In summary, combined treatment with La(III) and elevated UV-B radiation seriously affected nitrogen nutrition in soybean roots through the inhibition of nitrate reduction and ammonia assimilation.

Silymarin induces cell cycle arrest and apoptosis in ovarian cancer cells.

The polyphenolic flavonoid silymarin that is the milk thistle extract has been found to possess an anti-cancer effect against various human epithelial cancers. In this study, to explore the regulative effect of silymarin on human ovarian cancer line A2780s and PA-1 cells, 3-[4, 5-dimethylthiazol-2-yl]-2, 5-diphenyltetrazolium bromide assay and flow cytometry were respectively used to determine the inhibitory effect of silymarin on the both cell lines, and to measure their cell cycle progression. Apoptosis induction and mitochondrial membrane potential damage were separately detected by terminal deoxynucleotidyl transferase-mediated 2'-deoxyuridine 5'-triphosphate nick end labeling assay and 5,5',6,6'-tetrachloro-1,1',3,3'-tetraethylbenzimidazolylcarbocyanine iodide staining. Additionally, western blotting was applied to determine cytochrome C release and expression levels of p53, p21, p27, p16, CDK2, Bax, Bcl-2, procaspase-9, procaspase-3, cleaved caspase-9 and caspase-3 proteins. The activity of caspase-9 and caspase-3 was measured using Caspase-Glo-9 and Caspase-Glo-3 assay. The results indicated that silymarin effectively suppressed cell growth in a dose- and time-dependent manner, and arrested cell cycle progression at G1/S phase in A2780s and PA-1 cells via up-regulation of p53, p21, and p27 protein expression, and down-regulation of CDK2 protein expression. Additionally, silymarin treatment for 24h at 50 and 100µg/ml resulted in a reduction of mitochondrial membrane potential and cytochrome C release, and significantly induced apoptosis in A2780s and PA-1 cells by increasing Bax and decreasing Bcl-2 protein expression, and activation of caspase-9 and caspase-3. Therefore, silymarin is a possible potential candidate for the prevention and treatment of ovarian cancer.

Iron binding capacity of dephytinised soy protein isolate hydrolysate as influenced by the degree of hydrolysis and enzyme type.

Soy protein is increasingly used in extended meat products and dairy type products due to the presence of high quality proteins with excellent functional properties. However, it has been shown to inhibit iron bioavailability because of phytic acid present in the protein. This present study investigated the effects of dephytinise from soy protein isolate (SPI) on iron binding capacity and degree of hydrolysis. Also the effects of enzyme type and degree of hydrolysis on iron binding capacity were studied. It was demonstrated that phytase and anion exchange resin could remove effectively the phytate from SPI. The dephytinise would decrease the degree of hydrolysis of SPI. The enzyme type and degree of hydrolysis influenced significantly the iron binding capacity of the hydrolysate. Flavourzyme might be the best choice for producing peptides with iron binding capacity from SPI and middle degree of hydrolysis would be benefitable to this process.

Lanthanum (III) regulates the nitrogen assimilation in soybean seedlings under ultraviolet-B radiation.

Ultraviolet-B (UV-B, 280-320 nm) radiation has seriously affected the growth of plants. Finding the technology/method to alleviate the damage of UV-B radiation has become a frontal topic in the field of environmental science. The pretreatment with rare earth elements (REEs) is an effective method, but the regulation mechanism of REEs is unknown. Here, the regulation effects of lanthanum (La(III)) on nitrogen assimilation in soybean seedlings (Glycine max L.) under ultraviolet-B radiation were investigated to elucidate the regulation mechanism of REEs on plants under UV-B radiation. UV-B radiation led to the inhibition in the activities of the key enzymes (nitrate reductase, glutamine synthetase, glutamate synthase) in the nitrogen assimilation, the decrease in the contents of nitrate and soluble proteins, as well as the increase in the content of amino acid in soybean seedlings. The change degree of UV-B radiation at the high level (0.45 W m(-2)) was higher than that of UV-B radiation at the low level (0.15 W m(-2)). The pretreatment with 20 mg L(-1) La(III) could alleviate the effects of UV-B radiation on the activities of nitrate reductase, glutamine synthetase, glutamate synthase, and glutamate dehydrogenase, promoting amino acid conversion and protein synthesis in soybean seedlings. The regulation effect of La(III) under UV-B radiation at the low level was better than that of UV-B radiation at the high level. The results indicated that the pretreatment with 20 mg L(-1) La(III) could alleviate the inhibition of UV-B radiation on nitrogen assimilation in soybean seedlings.

Over-expression of Oct4 in human esophageal squamous cell carcinoma.

The Octamer 4 gene (Oct4) is a master pluripotency controller that has been detected in several types of tumors. Here, we examine the expression of Oct4 in human esophageal squamous cell carcinoma (ESCC). We found that punctate Oct4 protein was expressed in most (93.7%) ESCC samples but it was not observed in esophageal mucosa. Some ESCC cells had the capacity to form tumorospheres; those with an Oct4(+)-rich cell phenotype had increased proliferation and Oct4 mRNA levels compared to those of differentiated cells in culture or xenograft tumors. The over-expression of Oct4 in ESCCs suggests that it is a potential target for ESCC therapy. Oct4 could be a useful tumor marker in an immunohistochemical panel designed to differentiate between ESCC and esophageal mucosa. Expression of Oct4 in tumorospheres might indicate the presence of a population of ECSCs and its expression in xenograft tumors suggests that Oct4 is also associated with tumor metastasis.

Expression of the stem cell marker, Nanog, in human endometrial adenocarcinoma.

The embryonic stem cell self-renewal gene, Nanog, has been shown to be expressed in several tumor types and to regulate tumor development. The aim of this study was to carry out a detailed analysis of Nanog expression in human endometrial adenocarcinoma (EAC). Immunohistochemical analysis and reverse transcription-polymerase chain reaction were used to characterize Nanog, Sox2, and Oct4 expression in tissue arrays containing EAC, benign endometrium samples, and tumorosphere cells. Tumorosphere formation of EAC-derived cells in the stem cell culture medium was also analyzed. Nanog expression was then analyzed in secondary tumors initiated by the injection of tumorospheres or tumorosphere-derived differentiated cells into 15 female nude mice. Apoptosis and cell proliferation were detected in the fluorescence-activated cell sorter and 3-(4,5-dimethylthiazolyl-2)-2,5-diphenyltetrazolium bromide experiments, respectively. The Nanog protein was expressed in a majority of EAC samples (45 of 55, 81.8%), but not in benign endometrium samples (0 of 26, 0.0%). Oct4 and Sox2 were also commonly expressed in EAC samples (42 of 55, 76.4% and 39 of 55, 70.9%, respectively). Subsets of cancer cells from all EAC samples (15 of 15, 100%) exhibited the capacity to form Nanog-positive tumorospheres. The tumorospheres also expressed Nanog, Oct4, and Sox2 mRNA and showed a higher proliferation potential than differentiated cells. All 15 mice that were injected with tumorosphere cell-formed tumors, whereas only 3 of 15 mice injected with differentiated cells derived from tumorospheres developed tumors. All secondary xenograft tumors still expressed Nanog protein and Nanog, Oct4, and Sox2 mRNA, and had higher proliferation and lower apostosis than did differentiated cells. Overexpression of Nanog in EACs suggests that Nanog may represent a potential therapeutic target for EAC. In addition, Nanog may be useful as a biomarker in an immunohistochemical panel to differentiate between EAC and benign endometrial tissues. The expression of Nanog in tumorospheres may be indicative of the presence of a population of endometrial cancer stem cells, and its expression in xenograft tumors suggests that Nanog may also be associated with tumor metastasis.