Identification of Metabolomic Markers in Frozen or Formalin-Fixed and Paraffin-Embedded Samples of Diffuse Glioma from Adults.

The aim of this study was to identify metabolomic signatures associated with the gliomagenesis pathway (IDH-mutant or IDH-wt) and tumor grade of diffuse gliomas (DGs) according to the 2021 WHO classification on frozen samples and to evaluate the diagnostic performances of these signatures in tumor samples that are formalin-fixed and paraffin-embedded (FFPE). An untargeted metabolomic study was performed using liquid chromatography/mass spectrometry on a cohort of 213 DG samples. Logistic regression with LASSO penalization was used on the frozen samples to build classification models in order to identify IDH-mutant vs. IDH-wildtype DG and high-grade vs low-grade DG samples. 2-Hydroxyglutarate (2HG) was a metabolite of interest to predict IDH mutational status and aminoadipic acid (AAA) and guanidinoacetic acid (GAA) were significantly associated with grade. The diagnostic performances of the models were 82.6% AUC, 70.6% sensitivity and 80.4% specificity for 2HG to predict IDH status and 84.7% AUC, 78.1% sensitivity and 73.4% specificity for AAA and GAA to predict grade from FFPE samples. Thus, this study showed that AAA and GAA are two novel metabolites of interest in DG and that metabolomic data can be useful in the classification of DG, both in frozen and FFPE samples.

Could immunoscore improve the prognostic and therapeutic management in patients with solid tumors?

The Immunoscore (ISc) is an emerging immune-based scoring system that has shown potential in improving the prognostic and therapeutic management of patients with solid tumors. The ISc evaluates the immune infiltrate within the tumor microenvironment (TME) and has demonstrated superior predictive ability compared to traditional histopathological parameters. It has been particularly promising in colorectal, lung, breast, and melanoma cancers. This review summarizes the clinical evidence supporting the prognostic value of the ISc and explores its potential in guiding therapeutic decisions, such as the selection of adjuvant therapies and recognizing patients likely to profit from immune checkpoint inhibitors (ICIs). The challenges and future directions of ISc implementation are also discussed, including standardization and integration into routine clinical practice.

Pilot Study on the Use of Untargeted Metabolomic Fingerprinting of Liquid-Cytology Fluids as a Diagnostic Tool of Malignancy for Thyroid Nodules.

Although it is the gold standard for assessing the malignancy of thyroid nodules (TNs) preoperatively, the cytological analysis of fine-needle aspiration cytology (FNAC) samples results in 20-30% of cases in indeterminate lesions (ITNs). As two-thirds of these lesions will appear benign after diagnostic surgery, improved preoperative diagnostic methods need to be developed. In this pilot study, we evaluate if the metabolomic profiles of liquid-based (CytoRich®) FNAC samples of benign and malignant nodules can allow the molecular diagnosis of TNs. We performed untargeted metabolomic analyses with CytoRich® FNAC in a monocentric retrospective study. The cohort was composed of cytologically benign TNs, histologically benign or papillary thyroid carcinomas (PTCs) cytologically ITNs, and suspicious/malignant TNs histologically confirmed as PTCs. The diagnostic performance of the identified metabolomic signature was assessed using several supervised classification methods. Seventy-eight patients were enrolled in the study. We identified 7690 peaks, of which 2697 ions were included for further analysis. We selected a metabolomic signature composed of the top 15 metabolites. Among all the supervised classification methods, the supervised autoencoder deep neural network exhibited the best performance, with an accuracy of 0.957 (0.842-1), an AUC of 0.945 (0.833-1), and an F1 score of 0.947 (0.842-1). Here, we report a promising new ancillary molecular technique to differentiate PTCs from benign TNs (including among ITNs) based on the metabolomic signature of FNAC sample fluids. Further studies with larger cohorts are now needed to identify a larger number of biomarkers and obtain more robust signatures.

BioNet: a large-scale and heterogeneous biological network model for interaction prediction with graph convolution.

MOTIVATION: Understanding chemical-gene interactions (CGIs) is crucial for screening drugs. Wet experiments are usually costly and laborious, which limits relevant studies to a small scale. On the contrary, computational studies enable efficient in-silico exploration. For the CGI prediction problem, a common method is to perform systematic analyses on a heterogeneous network involving various biomedical entities. Recently, graph neural networks become popular in the field of relation prediction. However, the inherent heterogeneous complexity of biological interaction networks and the massive amount of data pose enormous challenges. This paper aims to develop a data-driven model that is capable of learning latent information from the interaction network and making correct predictions. RESULTS: We developed BioNet, a deep biological networkmodel with a graph encoder-decoder architecture. The graph encoder utilizes graph convolution to learn latent information embedded in complex interactions among chemicals, genes, diseases and biological pathways. The learning process is featured by two consecutive steps. Then, embedded information learnt by the encoder is then employed to make multi-type interaction predictions between chemicals and genes with a tensor decomposition decoder based on the RESCAL algorithm. BioNet includes 79 325 entities as nodes, and 34 005 501 relations as edges. To train such a massive deep graph model, BioNet introduces a parallel training algorithm utilizing multiple Graphics Processing Unit (GPUs). The evaluation experiments indicated that BioNet exhibits outstanding prediction performance with a best area under Receiver Operating Characteristic (ROC) curve of 0.952, which significantly surpasses state-of-theart methods. For further validation, top predicted CGIs of cancer and COVID-19 by BioNet were verified by external curated data and published literature.

Ergosta-7,9(11),22-trien-3ß-ol Alleviates Intracerebral Hemorrhage-Induced Brain Injury and BV-2 Microglial Activation.

Intracerebral hemorrhage (ICH) is a devastating neurological disorder characterized by an exacerbation of neuroinflammation and neuronal injury, for which few effective therapies are available at present. Inhibition of excessive neuroglial activation has been reported to alleviate ICH-related brain injuries. In the present study, the anti-ICH activity and microglial mechanism of ergosta-7,9(11),22-trien-3ß-ol (EK100), a bioactive ingredient from Asian medicinal herb Antrodia camphorate, were evaluated. Post-treatment of EK100 significantly attenuated neurobehavioral deficit and MRI-related brain lesion in the mice model of collagenase-induced ICH. Additionally, EK100 alleviated the inducible expression of cyclooxygenase (COX)-2 and the activity of matrix metalloproteinase (MMP)-9 in the ipsilateral brain regions. Consistently, it was shown that EK100 concentration-dependently inhibited the expression of COX-2 protein in Toll-like receptor (TLR)-4 activator lipopolysaccharide (LPS)-activated microglial BV-2 and primary microglial cells. Furthermore, the production of microglial prostaglandin E2 and reactive oxygen species were attenuated by EK100. EK100 also attenuated the induction of astrocytic MMP-9 activation. Among several signaling pathways, EK100 significantly and concentration-dependently inhibited activation of c-Jun N-terminal kinase (JNK) MAPK in LPS-activated microglial BV-2 cells. Consistently, ipsilateral JNK activation was markedly inhibited by post-ICH-treated EK100 in vivo. In conclusion, EK100 exerted the inhibitory actions on microglial JNK activation, and attenuated brain COX-2 expression, MMP-9 activation, and brain injuries in the mice ICH model. Thus, EK100 may be proposed and employed as a potential therapeutic agent for ICH.

Proteomic analysis identified LBP and CD14 as key proteins in blood/biphasic calcium phosphate microparticle interactions.

Immediately upon implantation, scaffolds for bone repair are exposed to the patient's blood. Blood proteins adhere to the biomaterial surface and the protein layer affects both blood cell functions and biomaterial bioactivity. Previously, we reported that 80-200 µm biphasic calcium phosphate (BCP) microparticles embedded in a blood clot, induce ectopic woven bone formation in mice, when 200-500 µm BCP particles induce mainly fibrous tissue. Here, in a LC-MS/MS proteomic study we compared the differentially expressed blood proteins (plasma and blood cell proteins) and the deregulated signaling pathways of these osteogenic and fibrogenic blood composites. We showed that blood/BCP-induced osteogenesis is associated with a higher expression of fibrinogen (FGN) and an upregulation of the Myd88- and NF-κB-dependent TLR4 signaling cascade. We also highlighted the key role of the LBP/CD14 proteins in the TLR4 activation of blood cells by BCP particles. As FGN is an endogenous ligand of TLR4, able to modulate blood composite stiffness, we propose that different FGN concentrations modify the blood clot mechanical properties, which in turn modulate BCP/blood composite osteoactivity through TLR4 signaling. The present findings provide an insight at the protein level, into the mechanisms leading to an efficient bone reconstruction by blood/BCP composites. STATEMENT OF SIGNIFICANCE: Upon implantation, scaffolds for bone repair are exposed to the patient's blood. Blood proteins adhere to bone substitute surface and this protein layer affects both biomaterial bioactivity and bone healing. Therefore, for the best outcome for patients, it is crucial to understand the molecular interactions between blood and bone scaffolds. Biphasic calcium phosphate (BCP) ceramics are considered as the gold standard in bone reconstruction surgery. Here, using proteomic analyses we showed that the osteogenic properties of 80-200 µm BCP particles embedded in a blood clot is associated with a higher expression of fibrinogen. Fibrinogen upregulates the Myd88- and NF-κB-dependent TLR4 pathway in blood cells and, BCP-induced TLR4 activation is mediated by the LBP and CD14 proteins.

Tumor microenvironment affects exogenous sodium/iodide symporter expression.

For decades, sodium/iodide symporter NIS-mediated iodide uptake has played a crucial role in the radioactive ablation of thyroid cancer cells. NIS-based gene therapy has also become a promising tool for the treatment of tumors of extrathyroidal origin. But its applicability has been hampered by reduced expression of NIS, resulting in a moderated capacity to accumulate 131I and in inefficient ablation. Despite numerous preclinical enhancement strategies, the understanding of NIS expression within tumors remains limited. This study aims at a better understanding of the functional behavior of exogenous NIS expression in the context of malignant solid tumors that are characterized by rapid growth with an insufficient vasculature, leading to hypoxia and quiescence. Using subcutaneous HT29NIS and K7M2NIS tumors, we show that NIS-mediated uptake and NIS expression at the plasma membrane of cancer cells are impaired in the intratumoral regions. For a better understanding of the underlying molecular mechanisms induced by hypoxia and quiescence (separately and in combination), we performed experiments on HT29NIS cancer cells. Hypoxia and quiescence were both found to impair NIS-mediated uptake through mechanisms including NIS mis-localization. Modifications in the expression of proteins and metabolites involved in plasma membrane localization and in energy metabolism were found using untargeted proteomics and metabolomics approaches. In conclusion, our results provide evidence that hypoxia and quiescence impair NIS expression at the plasma membrane, and iodide uptake. Our study also shows that the tumor microenvironment is an important parameter for successful NIS-based cancer treatment.

Comparison of unsupervised machine-learning methods to identify metabolomic signatures in patients with localized breast cancer.

Genomics and transcriptomics have led to the widely-used molecular classification of breast cancer (BC). However, heterogeneous biological behaviors persist within breast cancer subtypes. Metabolomics is a rapidly-expanding field of study dedicated to cellular metabolisms affected by the environment. The aim of this study was to compare metabolomic signatures of BC obtained by 5 different unsupervised machine learning (ML) methods. Fifty-two consecutive patients with BC with an indication for adjuvant chemotherapy between 2013 and 2016 were retrospectively included. We performed metabolomic profiling of tumor resection samples using liquid chromatography-mass spectrometry. Here, four hundred and forty-nine identified metabolites were selected for further analysis. Clusters obtained using 5 unsupervised ML methods (PCA k-means, sparse k-means, spectral clustering, SIMLR and k-sparse) were compared in terms of clinical and biological characteristics. With an optimal partitioning parameter k = 3, the five methods identified three prognosis groups of patients (favorable, intermediate, unfavorable) with different clinical and biological profiles. SIMLR and K-sparse methods were the most effective techniques in terms of clustering. In-silico survival analysis revealed a significant difference for 5-year predicted OS between the 3 clusters. Further pathway analysis using the 449 selected metabolites showed significant differences in amino acid and glucose metabolism between BC histologic subtypes. Our results provide proof-of-concept for the use of unsupervised ML metabolomics enabling stratification and personalized management of BC patients. The design of novel computational methods incorporating ML and bioinformatics techniques should make available tools particularly suited to improving the outcome of cancer treatment and reducing cancer-related mortalities.

LC-MS based metabolomic profiling for renal cell carcinoma histologic subtypes.

Renal cell carcinomas (RCC) are classified according to their histological features. Accurate classification of RCC and comprehensive understanding of their metabolic dysregulation are of critical importance. Here we investigate the use of metabolomic analyses to classify the main RCC subtypes and to describe the metabolic variation for each subtype. To this end, we performed metabolomic profiling of 65 RCC frozen samples (40 clear cell, 14 papillary and 11 chromophobe) using liquid chromatography-mass spectrometry. OPLS-DA multivariate analysis based on metabolomic data showed clear discrimination of all three main subtypes of RCC (R2 = 75.0%, Q2 = 59.7%). The prognostic performance was evaluated using an independent cohort and showed an AUROC of 0.924, 0.991 and 1 for clear cell, papillary and chromophobe RCC, respectively. Further pathway analysis using the 21 top metabolites showed significant differences in amino acid and fatty acid metabolism between three RCC subtypes. In conclusion, this study shows that metabolomic profiling could serve as a tool that is complementary to histology for RCC subtype classification. An overview of metabolic dysregulation in RCC subtypes was established giving new insights into the understanding of their clinical behaviour and for the development of targeted therapeutic strategies.

Hirsutanol A Attenuates Lipopolysaccharide-Mediated Matrix Metalloproteinase 9 Expression and Cytokines Production and Improves Endotoxemia-Induced Acute Sickness Behavior and Acute Lung Injury.

Activated human monocytes/macrophages, which increase the levels of matrix metalloproteinases (MMPs) and pro-inflammatory cytokines, are the essential mechanisms for the progression of sepsis. In the present study, we determined the functions and mechanisms of hirsutanolA (HA), which is isolated from the red alga-derived marine fungus Chondrostereum sp. NTOU4196, on the production of pro-inflammatory mediators produced from lipopolysaccharide (LPS)-treated THP-1 cells. Our results showed that HA suppressed LPS-triggered MMP-9-mediated gelatinolysis and expression of protein and mRNA in a concentration-dependent manner without effects on TIMP-1 activity. Also, HA significantly attenuated the levels of TNF-α, IL-6, and IL-1ß from LPS-treated THP-1 cells. Moreover, HA significantly inhibited LPS-mediated STAT3 (Tyr705) phosphorylation, IκBα degradation and ERK1/2 activation in THP-1 cells. In an LPS-induced endotoxemia mouse model, studies indicated that HA pretreatment improved endotoxemia-induced acute sickness behavior, including acute motor deficits and anxiety-like behavior. HA also attenuated LPS-induced phospho-STAT3 and pro-MMP-9 activity in the hippocampus. Notably, HA reduced pathologic lung injury features, including interstitial tissue edema, infiltration of inflammatory cells and alveolar collapse. Likewise, HA suppressed the induction of phospho-STAT3 and pro-MMP-9 in lung tissues. In conclusion, our results provide pharmacological evidence that HA could be a useful agent for treating inflammatory diseases, including sepsis.

Theissenolactone C Exhibited Ocular Protection of Endotoxin-Induced Uveitis by Attenuating Ocular Inflammatory Responses and Glial Activation.

The aim of this study was to investigate the effects of a natural component, theissenolactone C (LC53), on the ocular inflammation of experimental endotoxin-induced uveitis (EIU) and its related mechanisms in microglia. Evaluation of the severity of anterior uveitis indicated that LC53 treatment significantly decreased iridal hyperemia and restored the clinical scores. Additionally, the deficient retina functions of electroretinography were improved by LC53. LC53 significantly reduced levels of tumor necrosis factor (TNF)-α, monocyte chemoattractant protein-1, protein leakage and activation of matrix metalloproteinases in the anterior section during EIU. Moreover, LC53 treatment decreased the oxidative stress as well as neuroinflammatory reactivities of GFAP and Iba-1 in the posterior section. Furthermore, LC53 decreased the phosphorylation of p65, expression of HSP90, Bax, and cleaved-caspase-3 in EIU. According to the microglia studies, LC53 significantly abrogated the productions of TNF-α, PGE2, NO and ROS, as well as inducible NO synthase and cyclooxygenase-2 expression in LPS-stimulated microglial BV2 cells. The microglial activation of IKKß, p65 phosphorylation and nuclear phosphorylated p65 translocation were strongly attenuated by LC53. On the other hand, LC53 exhibited the inhibitory effects on JNK and ERK MAPKs activation. Our findings indicated that LC53 exerted the ocular-protective effect through its inhibition on neuroinflammation, glial activation, and apoptosis in EIU, suggesting a therapeutic potential with down-regulation of the NF-κB signaling for uveitis and retinal inflammatory diseases.

[Advances in application of Jurkat cell model in research on infectious diseases].

Infectious diseases can be caused by multiple pathogens, which can produce specific immune response in human body. The immune response produced by T cells is cellular immunity, which plays an important role in the anti-infection process of human body, and can participate in immunological protection and cause immunopathology. The outcome of various infectious diseases is closely related to cellular immune function, especially the function of T cells. Jurkat cells belong to the human acute T lymphocyte leukemia cell line. Jurkat cell model can simulate the function T lymphocytes, so it is widely used in the in vitro studies of T cell signal transduction, cytokines, and receptor expression, and can provide reference and guidance for the treatment of various infectious diseases and the research on their pathogenesis. The Jurkat cell model has been widely used in the in vitro studies of viral diseases and atypical pathogens, but parasitic infection studies using the Jurkat cell model are still rare. This article reviews advances in the application of Jurkat cell model in the research on infectious diseases.

Phyllanthus emblica Linn. fruit extract potentiates the anticancer efficacy of mitomycin C and cisplatin and reduces their genotoxicity to normal cells in vitro.

OBJECTIVE: Fruit of Phyllanthus emblica Linn. (PE) is widely consumed as a functional food and used as a folk medicine due to its remarkable nutritional and pharmacological effects. Mitomycin C (MMC) and cisplatin (cDDP) are the most widely used forms of chemotherapeutic drug, but their clinical use is limited by their genotoxicity to normal cells. We aimed to determine whether PE has potential to reduce the genotoxicity, while improving the anticancer effect, of MMC and cDDP. METHODS: Cell proliferation was evaluated using the trypan blue exclusion assay and colony-forming assay. Genomic instability (GIN) was measured using the cytokinesis-block micronucleus assay. RESULTS: Co-treatment (72 h) with PE at 20-320 µg/ml significantly enhanced the efficacy of MMC (0.05 µg/ml) and cDDP (1 µg/ml) against Colo205 colorectal cancer cells (P<0.05), and at 80-320 µg/ml significantly decreased MMC- and cDDP-induced GIN and multinucleation in normal colonic NCM460 cells (P<0.05). PE significantly decreased the mitotic index (P<0.01), blocked mitotic progression (P<0.05), and promoted apoptosis (P<0.01) in MMC- and cDDP-treated NCM460 cells, suggesting that PE-mediated inhibition of mitosis and induction of apoptosis may limit the division and survival of highly damaged cells. Also, PE was found to inhibit the clonal expansion of MMC- and cDDP-treated NCM460 cells (P<0.05) and decrease the heterogeneity of the surviving clones. CONCLUSIONS: PE potentiates the anticancer efficacy of MMC and cDDP, while preventing their genotoxicity and inhibiting clonal expansions of unstable genomes in normal cells. These data suggest that PE has the potential to reduce the risk of secondary cancers induced by chemotherapeutics.

Analysis of 28 trace elements in the blood and serum antioxidant status in chickens under arsenic and/or copper exposure.

This study aimed to evaluate the 28 trace elements in the blood and serum antioxidant status in chickens under arsenic (As) and/or copper (Cu) exposure. A total of 200 1-day-old male Hy-Line chickens were fed either a commercial diet (C-group) or arsenic trioxide (30 mg/kg) and/or cupric sulfate (300 mg/kg) for 90 days. The 28 trace element levels in the blood were analyzed by inductively coupled plasma mass spectrometry (ICP-MS). The concentrations of As in the blood of chickens were elevated approximately 17.15-fold, 2.30-fold, and 13.37-fold in the As-group, Cu-group, and As + Cu-group, respectively, at 90 days. The concentrations of Cu did not change in the As-group and increased approximately 29.53 and 23.37% in the Cu-group and As + Cu-group, respectively, at 90 days. Moreover, As exposure caused ion profile disorders in the blood, including increased concentrations of Na, Mg, Si, K, Cr, Fe, and Se and reduced B, Ca, Ti, V, Mn, Co, Ni, Zn, Sr, and Mo. Cu exposure increased the contents of Mg, Si, Ca, Ti, V, Cr, Mn, Fe, Co, Zn, and Se and decreased the content of B, Ca, Al, Ni, and Mo. As + Cu exposure increased the contents of Mg, Si, Cr, Fe, Zn, and Se and decreased the content of B, Ca, Ti, Co, Ni, Sr, and Mo. Moreover, As and/or Cu exposure induced oxidative stress in the blood of chickens. In conclusion, the results indicated that the mixture of As and Cu caused a synergistic effect via disturbing homeostasis of trace elements and oxidative stress in the blood of chickens.

Assessment of 28 trace elements and 17 amino acid levels in muscular tissues of broiler chicken (Gallus gallus) suffering from arsenic trioxide.

The contents of 28 trace elements, 17 amino acid were evaluated in muscular tissues (wings, crureus and pectoralis) of chickens in response to arsenic trioxide (As2O3). A total of 200 one-day-old male Hy-line chickens were fed either a commercial diet (C-group) or an As2O3 supplement diet containing 7.5mg/kg (L-group), 15mg/kg (M-group) or 30mg/kg (H-group) As2O3 for 90 days. The elements content was analyzed by inductively coupled plasma mass spectrometry (ICP-MS). Under As2O3 exposure, the concentration of As were elevated 8.87-15.76 fold, 7.93-15.63 fold and 5.94-12.45 fold in wings, crureus and pectoralis compared to the corresponding C-group, respectively. 19 element levels (lithium (Li), magnesium (Mg), aluminum (Al), silicon (Si), kalium (K), vanadium (V), chromium (Cr), manganese (Mn), nickel (Ni), copper (Cu), selenium (Se), strontium (Sr), molybdenum (Mo), cadmium (Cd), tin (Sn), antimony (Sb), barium (Ba), mercury (Hg) and lead (Pb), 9 element levels (K, Co, Ni, Cu, As, Se, Sr, Sn, Ba and Hg) and 4 element levels (Mn, cobalt (Co), As, Sr and Ba) were significantly increased (P < 0.05) in wing, crureus and pectoralis, respectively. 2 element levels (sodium (Na) and zinc (Zn)), 5 element levels (Li, Na, Si, titanium (Ti and Cr), 13 element levels (Li, Na, Mg, K, V, Cr, iron (Fe), Cu, Zn, Mo, Sn, Hg and Pb) were significantly decreased (P < 0.05) in wing muscle, crureus and pectoralis, respectively. Additionally, in crureus and pectoralis, the content of total amino acids (TAA) was no significant alterations in L and M-group and then increased approximately 10.2% and 7.6% in H-group, respectively (P < 0.05). In wings, the level of total amino acids increased approximately 10% in L-group, whereas it showed unchanged in M and H-group compared to the corresponding C-group. We also observed that significantly increased levels of proline, cysteine, aspartic acid, methionine along with decrease in the tyrosine levels in muscular tissues compared to the corresponding C-group. In conclusion, the residual of As in the muscular tissues of chickens were dose-dependent and disrupts trace element homeostasis, amino acids level in muscular tissues of chickens under As2O3 exposure. Additionally, the response (trace elements and amino acids) were different in wing, thigh and pectoral of chick under As2O3 exposure. This study provided references for further study of heavy metal poisoning and may be helpful to understanding the toxicological mechanism of As2O3 exposure in muscular tissues of chickens.

Four microRNAs promote prostate cell proliferation with regulation of PTEN and its downstream signals in vitro.

BACKGROUND: Phosphatase and tensin homologue (PTEN), as a tumor suppressor, plays vital roles in tumorigenesis and progression of prostate cancer. However, the mechanisms of PTEN regulation still need further investigation. We here report that a combination of four microRNAs (miR-19b, miR-23b, miR-26a and miR-92a) promotes prostate cell proliferation by regulating PTEN and its downstream signals in vitro. METHODOLOGY/PRINCIPAL FINDINGS: We found that the four microRNAs (miRNAs) could effectively suppress PTEN expression by directly interacting with its 3' UTR in prostate epithelial and cancer cells. Under-expression of the four miRNAs by antisense neutralization up-regulates PTEN expression, while overexpression of the four miRNAs accelerates epithelial and prostate cancer cell proliferation. Furthermore, the expression of the four miRNAs could, singly or jointly, alter the expression of the key components in the phosphoinositide 3-kinase (PI3K)/Akt pathway, including PIK3CA, PIK3CD, PIK3R1 and Akt, along with their downstream signal, cyclin D1. CONCLUSIONS: These results suggested that the four miRNAs could promote prostate cancer cell proliferation by co-regulating the expression of PTEN, PI3K/Akt pathway and cyclin D1 in vitro. These findings increase understanding of the molecular mechanisms of prostate carcinogenesis and progression, even provide valuable insights into the diagnosis, prognosis, and rational design of novel therapeutics for prostate cancer.

[A study of inhibitory effect of chromogranin A derived peptide CGA(47 _ 66) on hyper - permeability of endothelium induced by serum of septic patient].

OBJECTIVE: To explore the role of chromogranin A ( CGA) derived peptide CGA47~ ( Chromfungin, CHR) on septic serum induced high permeability of vascular endothelial cells. METHODS: Human umbilical venous endothelial cell line (EA.hy926 cells) was exposed to CHR, serum of septic shock patient, and tumor necrosis factor-a (TNF-a) respectively. Methyl thiazolyl tetrazolium (MTT) method, Transwell assay and immunofluorescence were performed to determine cell viability (absorbance (A) value J, permeability of monolayer endothelial cells (A value) , and the morphological characteristic and distribution ofF -actin respectively. RESULTS: Compared with the blank control group, when EA.hy926 were exposed to CHR with 1, 10, 100 nmol/L the cell activity was not significantly affected (A value: 1.219 ± 0.253, 1.179 ± 0.065, 1.179 ± 0.062 vs. 1.306 ± 0.162, all P>0.05), while when the cells was exposed to CHR in 1 000 nmol/L the cell activity was significantly inhibited (A value: 1.049 ± 0.256 vs. 1.306 ± 0.162, t=-2.390, P=0.031 ). Compared with blank control group, when the cells were exposed to CHR of 1, 10, 100 nmol/L a significant decrease in permeability in EA.hy926 cells was observed (A value: 1.619 ± 0.324, 1.496 ± 0.356, 1.132 ± 0.280 vs. 2.315 ± 0.440, P<0.05 or P<0.01 ). Treatment of septic shock patient's serum or TNF-a to EA. hy926 produced an obvious increase in its permeability (septic serum group A value: 1.204 ± 0.248 vs. 0.277 ± 0.017, P<0.01; TNF-a group A value: 2.485 ± 0.113 vs. 1.602 ± 0.679, P<0.05). High-permeability induced by TNF-a or septic shock patient's serum was alleviated hy CHR in the concentration of 1, 10, 100 nmol/L in a dose-dependent manner (septic serum + CHR group A value: 0.299 ± 0.065, 0.224 ± 0.028, 0.131 ± 0.015 vs. 1.204 ± 0.248; TNF -a + CHR group A value: 1.995 ± 0.394, 1.920 ± 0.096, 1.744 ± 0.475 vs. 2.485 ± 0.113, P<0.05 or P<0.01 ). Under a laser scanning confocal microscope, it was found that the F-actin cytoskeleton of EA.hy926 cells was redistributed, and more stress fibers were found in the septic shock patient's serum group and TNF-α group, while CHR obviously alleviated the above effects induced by septic shock patient's serum or TNF-α. CONCLUSION: In a dose-dependent manner, CHR may inhibit increased permeability of vascular endothelial cells induced by septic shock patient's serum, its underlying mechanism may be related to inhibition of the effect of TNF-α.

Role and fate of SP100 protein in response to Rep-dependent nonviral integration system.

Previously, we studied an AAVS1 site-specific non-viral integration system with a Rep-donor plasmid and a plasmid containing adeno-associated virus integration element. Our earlier study focused on the plasmid vector itself, but the cellular response to the system was still unknown. SP100 is a member of the promyelocytic leukemia nuclear bodies. It is involved in many cellular processes such as transcriptional regulation and the cellular intrinsic immune response against viral infection. In this study, we revealed that SP100 inhibited the Rep-dependent nonviral integration. Conversely, transient expression of Rep78 increased the degradation of SP100. This degradation was inhibited by treatment with MG132, an inhibitor of the ubiquitin proteasome. SP100 and Rep78 are both located in the nucleolus, which provides the spatial possibility for their interaction. Rep78 was coimmunoprecipitated with the enhanced green fluorescent protein (EGFP)-SP100 fusion protein but not EGFP, which verified the interaction between Rep78 and SP100. These results have enriched our knowledge about the cellular protein SP100 and Rep-dependent nonviral integration. It may lead to an improvement in the application of Rep-related transgene integration method and in the selection of target cells.

MicroRNA-7 inhibits tumor growth and metastasis by targeting the phosphoinositide 3-kinase/Akt pathway in hepatocellular carcinoma.

UNLABELLED: MicroRNAs (miRNAs) are known to be involved in carcinogenesis and tumor progression in hepatocellular carcinoma (HCC). Recently, microRNA-7 (miR-7) has been proven to play a substantial role in glioblastoma and breast cancer, but its functions in the context of HCC remain unknown. Here, we demonstrate that miR-7 inhibits HCC cell growth and metastasis in vitro and in vivo. We first screened and identified a novel miR-7 target, phosphoinositide 3-kinase catalytic subunit delta (PIK3CD). Overexpression of miR-7 would specifically and markedly down-regulate its expression. miR-7-overexpressing subclones showed significant cell growth inhibition by G(0) /G(1) -phase cell-cycle arrest and significant impairment of cell migration in vitro. To identify the mechanisms, we investigated the phosphoinositide 3-kinase (PI3K)/Akt pathway and found that Akt, mammalian target of rapamycin (mTOR), and p70S6K were down-regulated, whereas 4EBP1 was up-regulated in miR-7-overexpressing subclones. We also identified two novel, putative miR-7 target genes, mTOR and p70S6K, which further suggests that miR-7 may be a key regulator of the PI3K/Akt pathway. In xenograft animal experiments, we found that overexpressed miR-7 effectively repressed tumor growth (3.5-fold decrease in mean tumor volume; n = 5) and abolished extrahepatic migration from liver to lung in a nude mouse model of metastasis (n = 5). The number of visible nodules on the lung surface was reduced by 32-fold. A correlation between miR-7 and PIK3CD expression was also confirmed in clinical samples of HCC. CONCLUSION: These findings indicate that miR-7 functions as a tumor suppressor and plays a substantial role in inhibiting the tumorigenesis and reversing the metastasis of HCC through the PI3K/Akt/mTOR-signaling pathway in vitro and in vivo. By targeting PIK3CD, mTOR, and p70S6K, miR-7 efficiently regulates the PI3K/Akt pathway. Given these results, miR-7 may be a potential therapeutic or diagnostic/prognostic target for treating HCC.