Near-infrared-dye labeled tumor vascular-targeted dimer GEBP11 peptide for image-guided surgery in gastric cancer.

Introduction: Positive resection margins occur in about 2.8%-8.2% gastric cancer surgeries and is associated with poor prognosis. Intraoperative guidance using Nearinfrared (NIR) fluorescence imaging is a promising technique for tumor detection and margin assessment. The goal of this study was to develop a tumor-specific probe for real-time intraoperative NIR fluorescence imaging guidance. Methods: The tumor vascular homing peptide specific for gastric cancer, GEBP11, was conjugated with a near-infrared fluorophore, Cy5.5. The binding specificity of the GEBP11 probes to tumor vascular endothelial cells were confirmed by immunofluorescent staining. The ability of the probe to detect tumor lesions was evaluated in two xenograft models. An orthotopic gastric cancer xenograft model was used to evaluate the efficacy of the GEBP11 NIR probes in real-time surgical guidance. Results: In vitro assay suggested that both mono and dimeric GEBP11 NIR probes could bind specifically to tumor vascular epithelial cells, with dimeric peptides showed better affinity. In tumor xenograft mice, live imaging suggested that comparing with free Cy5.5 probe, significantly stronger NIR signals could be detected at the tumor site at 24-48h after injection of mono or dimeric GEBP11 probes. Dimeric GEBP11 probe showed prolonged and stronger NIR signals than mono GEBP11 probe. Biodistribution assay suggested that GEBP11 NIR probes were enriched in gastric cancer xenografts. Using dimeric GEBP11 NIR probes in real-time surgery, the tumor margins and peritoneal metastases could be clearly visualized. Histological examination confirmed the complete resection of the tumor. Conclusion: (GEBP11)2-ACP-Cy5.5 could be a potential useful probe for intraoperative florescence guidance in gastric cancer surgery.

Visualization and quantification of de novo lipogenesis using a FASN-2A-GLuc mouse model.

Background: De novo lipogenesis (DNL) is a dynamic process that converts excess carbohydrates into fatty acids to maintain cellular homeostasis. Dysregulation of DNL is associated with diverse obesity-related diseases and many tumor types. Therefore, monitoring DNL in real-time with high sensitivity should be highly beneficial when screening therapeutic agents for their potential use as obesity treatments. Methods: A sequence coding for Gaussia luciferase (GLuc) preceded by a 2A peptide was inserted into the murine fatty acid synthase (FASN) genetic locus by homologous recombination to generate FASN-2A-GLuc mice. The luciferase mouse model was evaluated in conditions of physical and pharmacological stimuli by in vivo and ex vivo imaging. Results: The distribution of bioluminescence signals in different organs was identical to the FASN expression: high in white fat, brown fat, and the lungs. In addition, the bioluminescence signals accurately recapitulated the dynamic change of FASN in response to fasting and refeeding conditions. Moreover, with this murine reporter model, we also discovered that fatostatin, a synthetic inhibitor of sterol regulatory element-binding proteins, effectively inhibited DNL in multiple organs, especially in adipose tissues under a high-carbohydrate diet. Conclusions: Our FASN-2A-GLuc reporter mouse model proved to be a sensitive visualization tool for monitoring both systemic and organ-specific DNL in real time.

Gut Microbiota and Metabolite Changes in Patients With Ulcerative Colitis and Clostridioides difficile Infection.

Background: Patients with ulcerative colitis (UC) are at an increased risk of developing Clostridioides difficile infection (CDI), which in turn leads to poor outcomes. The gut microbial structure and metabolites in patients with UC and CDI have been scarcely studied. We hypothesized that CDI changes the gut microbiota and metabolites of patients with UC. Materials and Methods: This study included 89 patients: 30 healthy controls (HC group), 29 with UC alone (UCN group), and 30 with UC and CDI (UCP group). None of the participants has been exposed to antibiotic treatments during the 3 months before stool collection. Stool samples were analyzed using 16S rRNA gene sequencing of the V3-V4 region and gas chromatography tandem time-of-flight mass spectrometry. Results: The UCN group displayed lower diversity and richness in gut microbiota and a higher relative abundance of the phylum Proteobacteria than the HC group. There were no significant differences between the UCN and UCP groups in the α-diversity indices. The UCP group contained a higher relative abundance of the genera Clostridium sensu stricto, Clostridium XI, Aggregatibacter, and Haemophilus, and a lower relative abundance of genera Clostridium XIVb and Citrobacter than the UCN group. In the UCP group, the increased metabolites included putrescine, maltose, 4-hydroxybenzoic acid, 4-hydroxybutyrate, and aminomalonic acid. Spearman's correlation analysis revealed that these increased metabolites negatively correlated with Clostridium XlVb and positively correlated with the four enriched genera. However, the correlations between hemoglobin and metabolites were contrary to the correlations between erythrocyte sedimentation rate and high-sensitivity C-reactive protein and metabolites. Conclusion: Our study identified 11 differential genera and 16 perturbed metabolites in patients with UC and CDI compared to those with UC alone. These findings may guide the design of research on potential mechanisms and specific treatments for CDI in patients with UC.

The role of morphology, shell composition and protein corona formation in Au/Fe3O4 composite nanoparticle mediated macrophage responses.

The great interest in using nanoparticles (NPs) for biomedical applications is transversal to various materials despite the poorly understood correlation between their physicochemical properties and effects on the immune system. NPs, such as gold and Fe3O4, are generally regarded as safe, but the immunotoxicological profile of Au/Fe3O4 composite NPs with different physicochemical properties is not well documented. This study investigated the biological impact of Au/Fe3O4 composite NPs with different morphologies (spherical core-shell and flower-like) and shell composition in vitro to analyze their potential cytotoxic effects and inflammatory responses on RAW 264.7 cells. Au/Fe3O4 composite NPs with a flower-like structure (FLNPs) induce a pronounced reduction in cell viability compared with Au/Fe3O4 composite NPs with a spherical core-shell structure (CSNPs). The increased production of reactive oxygen species, which damages cellular membranes, might contribute to the cytotoxicity effect of FLNPs. However, CSNPs presented more RAW 264.7 cell adhesion and uptake than FLNPs. Remarkably, a significant TNF-α release was observed with CSNP treated RAW 264.7 cells other than that of FLNPs. Protein corona analysis revealed the adsorption of a distinct amount and profile of proteins on the surface of CSNPs and FLNPs. Given the similar particle size and ζ-potential of CSNPs and FLNPs under the cell culture condition, results indicate that the impact of Au/Fe3O4 composite NPs on the macrophage activity highly depends on their morphology, shell composition and protein corona profile.

Targeted imaging of esophageal adenocarcinoma with a near-infrared fluorescent peptide.

BACKGROUND: Targeted optical imaging offers a noninvasive and accurate method for the early detection of gastrointestinal tumors, especially for flat appearances. In our previous study, a sequence of SNFYMPL (SNF) was identified as a specific peptide to bind to esophageal carcinoma using phage-display technology. This study aimed to evaluate the tumor-targeting efficacy of Cy5.5-conjugated SNF probe for imaging of esophageal carcinoma in vitro and in vivo. METHODS: The SNF-Cy5.5 probe was synthesized and then identified using High Performance Liquid Chromatography (HPLC) and mass spectrometry (MS). Confocal fluorescence imaging and Flow cytometry analysis were performed to evaluate the binding specificity and the receptor binding affinity of SNF-Cy5.5 to OE33. In vivo imaging was performed to evaluate the targeting ability of SNF-Cy5.5 to esophageal carcinoma. RESULTS: The confocal imaging and flow cytometry analysis showed that SNF-Cy5.5 bound specifically to the plasma membrane of OE33 cells with a high affinity. In vivo, for non-block group, SNF-Cy5.5 probe exhibited rapid OE33 tumor targeting during 24 h p.i. and excellent tumor-to-background contrast at 2 h p.i. For the block group, SNF-Cy5.5 was not observed in the mice after 4 h p.i. Ex vivo imaging also revealed that a higher fluorescent signal intensity value of the tumors was clearly observed in the non-block group than that in the block group (2.6 ± 0.32 × 109 vs. 0.8 ± 0.08 × 109, p < 0.05). CONCLUSIONS: SNF-Cy5.5 was synthesized and characterized with a high efficiency and purity. The higher affinity, specificity, and tumor targeting efficacy of SNF-Cy5.5 were confirmed by in vitro and in vivo tests. SNF-Cy5.5 is a promising optical probe for the imaging of esophageal adenocarcinoma.

Albumin-based fluorescence resonance energy transfer nanoprobes for multileveled tumor tissue imaging and dye release imaging.

Tumor tissue imaging and drug release imaging are both crucial for tumor imaging and image-guided drug delivery. It is urgent to develop a multileveled tumor imaging platform to realize the multiple imaging applications. In this work, we synthesized an albumin-based fluorescence resonance energy transfer (FRET) probe Cy5/7@HSA NPs containing two near-infrared cyanine dyes (CyBI5 and CyBI7) with high FRET efficiency (97 %). Excellent brightness and efficient FRET inside Cy5/7@HSA NPs enabled high-sensitive cell imaging and tumor imaging. This unique nanoprobe imaged 4T1 tumor-bearing mice with high sensitivity (TBR = 5.2) at 24 h post-injection and the dyes penetrated the tumor interior around 4 h post-injection. The release of dyes from nanoprobes was also tracked. This result shows the strong potential of this albumin-based FRET nanoprobe as multileveled tumor imaging platform for in vivo tumor imaging, drug delivery and image-guided surgery.

KRAS Mutation-Responsive miR-139-5p inhibits Colorectal Cancer Progression and is repressed by Wnt Signaling.

Introduction: Colorectal cancer (CRC) frequently harbors KRAS mutations that result in chemoresistance and metastasis. MicroRNAs (miRNAs) are usually dysregulated and play important regulatory roles in tumor progression. However, the KRAS mutation-responsive miRNA profile in CRC remains uninvestigated. Methods: miR-139-5p was identified and evaluated by small RNA sequencing, qRT-PCR and in situ hybridization. The roles of miR-139-5p in CRC cells with and without KRAS mutation were determined by Cell Counting Kit-8 (CCK-8), colony formation, flow cytometry and transwell assays in vitro and by tumorigenesis and metastasis assays in vivo. Microarrays followed by bioinformatic analyses, luciferase reporter assays and Western blotting were applied for mechanistic studies. Results: miR-139-5p was significantly downregulated in KRAS-mutated CRC cells and tissues compared with their wild-type counterparts. Low miR-139-5p expression was associated with aggressive phenotypes and poor prognosis in CRC patients. miR-139-5p overexpression inhibited CRC cell proliferation, migration and invasion in vitro, sensitized tumors to chemotherapy, and impaired tumor growth and metastasis in vivo. Transcriptomic profiling identified multiple modulators in the Ras (JUN and FOS) and Wnt (CTNNB1 and DVL1) signaling pathways and the epithelial-to-mesenchymal transition (EMT) process (ZEB1) as direct targets of miR-139-5p, and inverse correlations were confirmed in CRC clinical tissues. Aberrantly activated Wnt signaling in KRAS-mutant cells was demonstrated to transcriptionally repress miR-139-5p through TCF4, forming a miR-139-5p/Wnt signaling double-negative feedback loop. Conclusions: We identified miR-139-5p as a KRAS-responsive miRNA and demonstrated its involvement in CRC progression. KRAS mutation disrupted the miR-139-5p/Wnt signaling reciprocal negative feedback mechanism, which might cause miR-139-5p downregulation and derepression of oncogenic signaling pathways and EMT. These results reveal a transcriptional regulatory mode of KRAS-driven malignant transformation and highlight miR-139-5p as a novel regulator of crosstalk between the Ras and Wnt signaling pathways in CRC.

In Vivo and in Situ Activated Aggregation-Induced Emission Probes for Sensitive Tumor Imaging Using Tetraphenylethene-Functionalized Trimethincyanines-Encapsulated Liposomes.

The design and exploration of fluorescent probes with high-sensitivity and low-background are essential for noninvasive optical molecular imaging. The in vivo and in situ activated aggregation-induced emission (AIE) probes were found to be ideal for achieving higher signal-to-background ratios for tumor detections. We herein developed novel tetraphenylethene-encapsulated liposomes (TPE-LPs) constructed by loading TPE-trimethincyanine into liposomes for the first time, and the probes were applied to tumor bioimaging in vivo. TPE-functionalized trimethincyanines were synthesized with a new and efficient one-pot reaction. In TPE-LPs, TPE-functionalized bicarboxylic acids benzoindole trimethinecyanine (TPE-BICOOH) fluorophores were found to be well dispersed in lipid bilayers (with non-restricted rotation) during the blood circulation, and then aggregated (with restriction of intramolecular rotation) upon liposome rupture in the tumor tissue, achieving a low-background and high-target signal for tumor imaging. The in situ activated AIE probes not only had great accumulation at the tumor site after intravenous injection in 4T1 tumor-bearing mice but also demonstrated excellent signal-to-background ratios, as well as low cytotoxicity and excellent biocompatibility. The proposed strategy is believed to be a simple and powerful tool for the sensitive detection of tumors.

Forkhead box C1 promotes colorectal cancer metastasis through transactivating ITGA7 and FGFR4 expression.

Metastatic colorectal cancer (CRC) is one of the most common causes of cancer death worldwide; however, the molecular mechanism underlying CRC metastasis remains unknown. Using an integrated approach, we identified forkhead box C1 (FOXC1) as a novel regulator of CRC metastasis. Elevated expression of FOXC1 is significantly correlated with metastasis, recurrence and reduced survival. FOXC1 overexpression promotes CRC invasion and lung metastasis, whereas FOXC1 knockdown has the opposite effect. In addition, FOXC1 directly binds its target genes integrin α7 (ITGA7) and fibroblast growth factor receptor 4 (FGFR4) and activates their expression. Genetic epistasis analysis confirmed that ITGA7 and FGFR4 act downstream of FOXC1. Furthermore, pharmaceutical inhibition of FGFR4 can reverse CRC metastasis mediated by FOXC1 overexpression. These results suggest that FOXC1 is a prognostic biomarker in CRC patients and targeting the FGFR4 signaling pathway may provide a promising strategy for the treatment of FOXC1-driven CRC metastasis.

Novel peptide GX1 inhibits angiogenesis by specifically binding to transglutaminase-2 in the tumorous endothelial cells of gastric cancer.

The clinical application of GX1, an optimal gastric cancer (GC) targeting peptide, is greatly limited because its receptor in the GC vasculature is unknown. In this study, we screened the candidate receptor of GX1, transglutaminase-2(TGM2), by co-immunoprecipitation (co-IP) combined with mass spectrometry. We found that TGM2 was up-regulated in GC vascular endothelial cells and that GX1 receptor expression was suppressed correspondingly after TGM2 downregulation. A highly consistent co-localization of GX1 receptor and TGM2 was detected at both the cellular and tissue levels. High TGM2 expression was evident in GC tissues from patients with poor prognosis. After TGM2 downregulation, the GX1-mediated inhibition of proliferation and migration and the induction of the apoptosis of GC vascular endothelial cells were weakened or even reversed. Finally, we observed that GX1 could inhibit the GTP-binding activity of TGM2 by reducing its intracellular distribution and downregulating its downstream molecular targets (nuclear factor-kappa B, NF-κB; hypoxia-inducible factor 1-α, HIF1α) in GC vascular endothelial cells. Our study confirms that peptide GX1 can inhibit angiogenesis by directly binding to TGM2, subsequently reducing the GTP-binding activity of TGM2 and thereby suppressing its downstream pathway(NF-κB/HIF1α). Our conclusions suggest that GX1/TGM2 may provide a new target for the diagnosis and treatment of GC.

Improved Tumor Targeting and Longer Retention Time of NIR Fluorescent Probes Using Bioorthogonal Chemistry.

The traditional labeling method for targeted NIR fluorescence probes requires directly covalent-bonded conjugation of targeting domains and fluorophores in vitro. Although this strategy works well, it is not sufficient for detecting or treating cancers in vivo, due to steric hindrance effects that relatively large fluorophore molecules exert on the configurations and physiological functions of specific targeting domains. The copper-free, "click-chemistry"-assisted assembly of small molecules in living systems may enhance tumor accumulation of fluorescence probes by improving the binding affinities of the targeting factors. Here, we employed a vascular homing peptide, GEBP11, as a targeting factor for gastric tumors, and we demonstrate its effectiveness for in vivo imaging via click-chemistry-mediated conjugation with fluorescence molecules in tumor xenograft mouse models. This strategy showed higher binding affinities than those of the traditional conjugation method, and our results showed that the tumor accumulation of click-chemistry-mediated probes are 11-fold higher than that of directly labeled probes. The tracking life was prolonged by 12-fold, and uptake of the probes into the kidney was reduced by 6.5-fold. For lesion tumors of different sizes, click-chemistry-mediated probes can achieve sufficient signal-to-background ratios (3.5-5) for in vivo detection, and with diagnostic sensitivity approximately 3.5 times that of traditional labeling probes. The click-chemistry-assisted detection strategy utilizes the advantages of "small molecule" probes while not perturbing their physiological functions; this enables tumor detection with high sensitivity and specific selectivity.

Reduced silent information regulator 1 signaling exacerbates sepsis-induced myocardial injury and mitigates the protective effect of a liver X receptor agonist.

Myocardial injury and dysfunction are critical manifestations of sepsis. Previous studies have reported that liver X receptor (LXR) activation is protective during sepsis. However, whether LXR activation protects against septic heart injury and its underlying mechanisms remain elusive. This study was designed to determine the role of LXR activation in the septic heart with a focus on SIRT1 (silent information regulator 1) signaling. Male cardiac-specific SIRT1 knockout mice (SIRT1-/-) and their wild-type littermates were subjected to sepsis by cecal ligation and puncture (CLP) in the presence or absence of LXR agonist T0901317. The survival rate of mice was recorded during the 7-day period post CLP. Our results demonstrated that SIRT1-/- mice suffered from exacerbated mortality and myocardial injury in comparison with their wild-type littermates. Meanwhile, T0901317 treatment improved mice survival, accompanied by significant ameliorations of myocardial injury and dysfunction in wild-type mice but not in SIRT1-/- mice. Furthermore, the levels of myocardial inflammatory cytokines (TNF-α, IL-6, IL-1ß, MCP-1, MPO and HMGB1), oxidative stress (ROS generation, MDA), endoplasmic-reticulum (ER) stress (protein levels of CHOP, GRP78, GRP94, IRE1α, and ATF6), and cardiac apoptosis following CLP were inhibited by T0901317 treatment in wild-type mice but not in SIRT1-/- mice. Mechanistically, T0901317 enhanced SIRT1 signaling and the subsequent deacetylation and activation of antioxidative FoxO1 and anti-ER stress HSF1, as well as the deacetylation and inhibition of pro-inflammatory NF-ΚB and pro-apoptotic P53, thereby alleviating sepsis-induced myocardial injury and dysfunction. Our data support the promise of LXR activation as an effective strategy for relieving heart septic injury.

SIRT1 Activation by Resveratrol Alleviates Cardiac Dysfunction via Mitochondrial Regulation in Diabetic Cardiomyopathy Mice.

BACKGROUND: Diabetic cardiomyopathy (DCM) is a major threat for diabetic patients. Silent information regulator 1 (SIRT1) has a regulatory effect on mitochondrial dynamics, which is associated with DCM pathological changes. Our study aims to investigate whether resveratrol, a SRIT1 activator, could exert a protective effect against DCM. METHODS AND RESULTS: Cardiac-specific SIRT1 knockout (SIRT1KO) mice were generated using Cre-loxP system. SIRT1KO mice displayed symptoms of DCM, including cardiac hypertrophy and dysfunction, insulin resistance, and abnormal glucose metabolism. DCM and SIRT1KO hearts showed impaired mitochondrial biogenesis and function, while SIRT1 activation by resveratrol reversed this in DCM mice. High glucose caused increased apoptosis, impaired mitochondrial biogenesis, and function in cardiomyocytes, which was alleviated by resveratrol. SIRT1 deletion by both SIRT1KO and shRNA abolished the beneficial effects of resveratrol. Furthermore, the function of SIRT1 is mediated via the deacetylation effect on peroxisome proliferator-activated receptor gamma coactivator 1-alpha (PGC-1α), thus inducing increased expression of nuclear respiratory factor 1 (NRF-1), NRF-2, estrogen-related receptor-α (ERR-α), and mitochondrial transcription factor A (TFAM). CONCLUSIONS: Cardiac deletion of SIRT1 caused phenotypes resembling DCM. Activation of SIRT1 by resveratrol ameliorated cardiac injuries in DCM through PGC-1α-mediated mitochondrial regulation. Collectively, SIRT1 may serve as a potential therapeutic target for DCM.

Activation of Cannabinoid Receptor Type II by AM1241 Ameliorates Myocardial Fibrosis via Nrf2-Mediated Inhibition of TGF-ß1/Smad3 Pathway in Myocardial Infarction Mice.

AIMS: Myocardial interstitial fibrosis is a major histologic landmark resulting in cardiac dysfunction after myocardial infarction (MI). Activation of cannabinoid receptor type II (CB2 receptor) have been demonstrated to reduce fibrosis in hepatic cirrhotic rat. However, the anti-fibrotic effect of CB2 receptor activation in infarcted hearts was still unclear. In this study, we aimed to investigate the effects of a CB2 receptor selective agonist AM1241 on myocardial fibrosis post MI in mice. METHODS: Echocardiograph was conducted to assess cardiac function. Fibrosis markers such as type I and type III collagen, fibronectin, Plasminogen activator inhibitor(PAI)-1 and tissue inhibitor of metalloprotease(TIMP)-1 were examined by Western blot, while collagens were directly observed by Sirius-red staining. Primary cultured cardiac fibroblasts(CFs) were subjected to hypoxia/serum deprivation (H/SD) injury to simulate ischemic conditions in vivo. Nrf2 siRNA were applied to explore the role of Nrf2 and TGF-ß1/Smad3 pathway in this process. RESULTS: Echocardiography showed that AM1241 significantly improved cardiac function, suppressed the expression of fibrosis markers such as collagen I and collagen III, fibronectin, PAI-1 and TIMP-1 in mice with MI. In cardiac fibroblasts subjected to H/SD injury, AM1241 reduced the elevated levels of α-SMA, collagen I and collagen III, which were partially abrogated by the Nrf2 siRNA transfection. Furthermore, AM1241 not only activated and accelerated the translocation of Nrf2 to nucleus, but also inhibited TGF-ß1/ Smad3 pathway in an Nrf2 dependent manner. CONCLUSION: CB2 receptor agonist AM1241 alleviated myocardial interstitial fibrosis via Nrf2 -mediated down-regulation of TGF-ß1/Smad3 pathway, which suggested that CB2 receptor activation might represent a promising target for retarding cardiac fibrosis after MI.

Combined cell surface carbonic anhydrase 9 and CD147 antigens enable high-efficiency capture of circulating tumor cells in clear cell renal cell carcinoma patients.

Circulating tumor cells (CTCs) have emerged as promising tools for noninvasive cancer detection and prognosis. Most conventional approaches for capturing CTCs use an EpCAM-based enrichment strategy, which does not work well in cancers that show low or no expression of EpCAM, such as renal cell carcinoma (RCC). In this study, we developed a new set of cell surface markers including CA9 and CD147 as alternative CTC-capture antigens specifically designed for RCC patients. We showed that the expression of both CA9 and CD147 was prevalent in a RCC patient cohort (n=70) by immunohistochemical analysis, with both molecules in combination covering 97.1% of cases. The NanoVelcro platform combined with CA9-/CD147-capture antibodies demonstrated significantly higher efficiency for capturing both CTC-mimicking renal cancer cells and RCC CTCs in peripheral blood, compared to the conventional EpCAM-based method. Using immunofluorescence cytological validation at the single-cell level, we were able to identify bona fide CTCs in RCC patient blood following the well-accepted criteria in our CTC-capture system. We further demonstrated a significant association of CTC numbers as well as the CTC expression status of Vimentin, a mesenchymal marker, with disease progression, including pathologic features and clinical staging. These results provide new insights into developing novel, effective targets/approaches for capturing CTCs, making CTCs a valuable tool for improved cancer detection, prognosis and treatment in RCC.

Quaternized Chitosan/Alginate-Fe3O4 Magnetic Nanoparticles Enhance the Chemosensitization of Multidrug-Resistant Gastric Carcinoma by Regulating Cell Autophagy Activity in Mice.

Multidrug resistance (MDR) and targeted therapies present major challenges in tumor chemotherapy. Nanoparticles (NPs) hold promise for use in cancer theranostics due to their advantages in terms of tumor-targeted cytotoxicity and imaging. In this study, we developed N-((2-hydroxy-3-trimethylammonium) propyl) chitosan chloride (HTCC)/alginate-encapsulated Fe3O4 magnetic NPs (HTCC-MNPs) and applied them to MDR gastric cancer both in vivo and in vitro. HTCC-MNPs were fabricated from sodium alginate (ALG), Fe3O4 and HTCC using an ionic gelation method. The sizes and physical characteristics of the NPs were determined using dynamic light scattering, transmission electron microscopy (TEM) and zeta potential analysis. The HTCC-MNPs exhibited excellent water solubility and biocompatibility as well as significantly reduced cell viability in the drug-resistant cancer cell line SGC7901/ADR, but not in normal gastric cells (P < 0.05). An analysis of LC3 expression demonstrated the involvement of autophagy in HTCC-MNP cytotoxicity. Additionally, apoptosis was verified using a DNA content assay. HTCC-MNPs led to mitochondrial membrane potential loss, decreased ATP production and excessive reactive oxygen species (ROS) generation compared to a control group (P < 0.05). Magnetic resonance imaging showed enrichment of HTCC-MNPs in tumor-bearing mice. In vivo bioluminescence imaging and tumor volume measurements revealed that HTCC-MNPs markedly inhibited in vivo tumor growth (P < 0.05). In conclusion, HTCC-MNPs significantly inhibited MDR gastric tumor growth and reduced tumor volume via the induction of cellular autophagy and apoptosis, which was attributed to mitochondrial dysfunction and excessive ROS accumulation.

POU2F2-oriented network promotes human gastric cancer metastasis.

BACKGROUND AND AIMS: Aberrant upregulation of POU2F2 expression has been discovered in metastatic gastric cancer (GC). However, the mechanisms underlying the aberrant upregulation and the potential functions of POU2F2 remain uncertain. DESIGN: The role and mechanism of POU2F2 in GC metastasis were investigated in gastric epithelial cells, GC cell lines and an experimental metastasis animal model by gain of function and loss of function. Upstream and downstream targets of POU2F2 were selected by bioinformatics and identified by luciferase reporter assay, electrophoretic mobility shift assay and chromatin immunoprecipitation PCR. The influence of miR-218 on its putative target genes (POU2F2, ROBO1 and IKK-ß) and GC metastasis was further explored via in vitro and in vivo approaches. RESULTS: Increased POU2F2 expression was detected in metastatic GC cell lines and patient samples. POU2F2 was induced by the activation of nuclear factor (NF)-κB and, in turn, regulated ROBO1 transcription, thus functionally contributing to GC metastasis. Finally, miR-218 was found to suppress GC metastasis by simultaneously mediating multiple molecules in the POU2F2-oriented network. CONCLUSIONS: This study demonstrated that NF-κB and the SLIT2/ROBO1 interaction network with POU2F2 as the central part may exert critical effects on tumour metastasis. Blocking the activation of the POU2F2-oriented metastasis network using miR-218 precursors exemplified a promising approach that sheds light on new strategies for GC treatment.

In vivo molecular imaging of gastric cancer in human-murine xenograft models with confocal laser endomicroscopy using a tumor vascular homing peptide.

The early detection of premalignant lesions and cancers are very important for improving the survival of patients with gastric malignancies. Confocal laser endomicroscopy (CLE) is a novel imaging tool for achieving real-time microscopy during the ongoing endoscopy at subcellular resolution. In the present study, to evaluate the feasibility of real-time molecular imaging of GEBP11 by CLE in gastric cancer, CLE was performed on two types of tumor-bearing mice models, as well as surgical specimens of patients with gastric cancer, after the application of GEBP11. A whole-body fluorescent imaging device was first used to screen for the strongest specific fluorescent signal in xenograft models. Next, the tumor sites, as well as human tissues, were scanned with CLE. After this, targeted specimens were obtained for fluorescence microscopy and histology. We confirmed that GEBP11 could specifically bind to co-HUVECs by means of CLE in cell experiments. Thereafter, a specific signal was observed in both subcutaneous and orthotopic xenograft models in vivo after the injection of FITC-GEBP11 via tail vein, whereas the group injected with FITC-URP showed no fluorescent signals. In human tissues, a specific signal of GEBP11 was observed in 26/28 neoplastic specimens and in 8/28 samples of non-neoplastic specimens from the patients (p < 0.01). The findings from ex vivo immunofluorescence microscopy of cryostat sections correlated well with that obtained by CLE. These findings indicate that the peptide, GEBP11, might be a potential candidate for the molecular imaging of gastric cancer.