Nitazoxanide is an FDA-approved antiprotozoal drug. Our previous study found that oral administration of nitazoxanide inhibited Western diet (WD)-induced hepatic steatosis in ApoE^-/- mice. However, the specific mechanism remains to be elucidated. In the present study, we performed an untargeted metabolomics approach to reveal the effect of nitazoxanide on the liver metabolic profiles in WD-fed ApoE^-/- mice, and carried out the cellular experiments to elucidate the underlying mechanisms. UPLC-MS-based untargeted metabolomics analysis was used to investigate the effect of nitazoxanide on global metabolite changes in liver tissues. The differential metabolites were screened for enrichment analysis and pathway analysis. Hepatocytes were treated with tizoxanide, the metabolite of nitazoxanide, to investigate the underlying mechanism based on the findings in metabolomics study. The improvement of liver lipid metabolism disorders by nitazoxanide treatment in WD-fed ApoE^-/- mice was mainly through regulating glycerophospholipid metabolism, D-glutamine and glutamate metabolism, glutathione metabolism, and arginine biosynthesis metabolism. Tizoxanide, the active metabolite of nitazoxanide, increased glutathione (GSH) contents and glutamate-cysteine ligase catalytic subunit (Gcl-c) and glutathione reductase (Gsr) mRNA expressions in HepG2 cells. Tizoxanide increased cystathionine β-synthase (CBS) and phosphatidylethanolamine N-methyltransferase (PEMT) protein levels, inhibited lipid accumulation in hepatocytes induced by free fatty acid (FFA). Tizoxanide increased S-adenosyl-L-homocysteine hydrolase (SAHH) protein levels in HepG2 cells and mouse primary liver cells stimulated with free fatty acid (FFA). Tizoxanide increased N-acetyl glutamate synthase (Nags) and carbamoylphosphate synthetase 1 (Cps1) mRNA expressions in HepG2 cells. In conclusion, nitazoxanide improves WD-induced hepatic steatosis in ApoE^-/- mice and the underlying mechanisms include increasing CBS expression, GSH content, PEMT protein expression, Nags and Cps1 mRNA expression in hepatocytes.

BACKGROUND: Prenatal evaluation of fetal lung maturity (FLM) is a challenge, and an effective non-invasive method for prenatal assessment of FLM is needed. The study aimed to establish a normal fetal lung gestational age (GA) grading model based on deep learning (DL) algorithms, validate the effectiveness of the model, and explore the potential value of DL algorithms in assessing FLM. METHODS: A total of 7013 ultrasound images obtained from 1023 normal pregnancies between 20 and 41 + 6 weeks were analyzed in this study. There were no pregnancy-related complications that affected fetal lung development, and all infants were born without neonatal respiratory diseases. The images were divided into three classes based on the gestational week: class I: 20 to 29 + 6 weeks, class II: 30 to 36 + 6 weeks, and class III: 37 to 41 + 6 weeks. There were 3323, 2142, and 1548 images in each class, respectively. First, we performed a pre-processing algorithm to remove irrelevant information from each image. Then, a convolutional neural network was designed to identify different categories of fetal lung ultrasound images. Finally, we used ten-fold cross-validation to validate the performance of our model. This new machine learning algorithm automatically extracted and classified lung ultrasound image information related to GA. This was used to establish a grading model. The performance of the grading model was assessed using accuracy, sensitivity, specificity, and receiver operating characteristic curves. RESULTS: A normal fetal lung GA grading model was established and validated. The sensitivity of each class in the independent test set was 91.7%, 69.8%, and 86.4%, respectively. The specificity of each class in the independent test set was 76.8%, 90.0%, and 83.1%, respectively. The total accuracy was 83.8%. The area under the curve (AUC) of each class was 0.982, 0.907, and 0.960, respectively. The micro-average AUC was 0.957, and the macro-average AUC was 0.949. CONCLUSIONS The normal fetal lung GA grading model could accurately identify ultrasound images of the fetal lung at different GAs, which can be used to identify cases of abnormal lung development due to gestational diseases and evaluate lung maturity after antenatal corticosteroid therapy. The results indicate that DL algorithms can be used as a non-invasive method to predict FLM.
Algorithms ; Deep Learning ; Female ; Gestational Age ; Humans ; Infant ; Infant, Newborn ; Lung/diagnostic imaging* ; Neural Networks, Computer ; Pregnancy

Prohibitin (PHB), an evolutionarily conserved mitochondrial inner membrane protein, is highly expressed in cells that require strong mitochondrial function. Recently, we demonstrated that the deletion of Phb in spermatocytes results in impaired mitochondrial function. In addition, PHB expression in the mitochondrial sheath of human sperm has a significantly negative correlation with mitochondrial reactive oxygen species levels, but a positive one with mitochondrial membrane potential and sperm motility. These results suggest that mitochondrial PHB expression plays a role in sperm motility. However, the mechanism of PHB-mediated regulation of sperm motility remains unknown. Here, we demonstrate for the first time that PHB interacts with protein kinase B (AKT) and exists in a complex with phospho-PHB (pT258) and phospho-AKT in the mitochondrial sheath of murine sperm, as determined using colocalization and coimmunoprecipitation assays. After blocking AKT activity using wortmannin (a phosphatidylinositol 3-kinase [PI3K] inhibitor), murine sperm have significantly ( P < 0.05) decreased levels of phospho-PHB (pT258) and the total and progressive motility. Furthermore, significantly ( P < 0.05) lower levels of phospho-PI3K P85 subunit α+γ (pY199 and pY467) and phospho-AKT (pS473; pT308) are found in sperm from infertile asthenospermic and oligoasthenospermic men compared with normospermic subjects, which suggest a reduced activity of the PI3K/AKT pathway in these infertile subjects. Importantly, these sperm from infertile subjects also have a significantly ( P < 0.05) lower level of phospho-PHB (pT258). Collectively, our findings suggest that the interaction of PHB with AKT in the mitochondrial sheath is critical for sperm motility, where PHB phosphorylation (pT258) level and PI3K/AKT activity are key regulatory factors.

OBJECTIVETo validate those obtained immunogenic membrane antigens candidate of human pancreatic cancer in the performed research.

METHODSIn the pre-studies, serum IgG purified from clinically collected sera of pancreatic cancer patients underwent immunoblot with human pancreatic cancer cell line SW1990 membrane protein, totally obtained 9 positive protein spots. Number 5 and 6 positive dots of immunoblot were identified by matrix-assisted laser desorption/ionization-time of flight mass spectrometry and peptide mass fingerprinting matching. The candidate membrane antigens were further validated in cell lines by RT-PCR and real-time PCR. RNA of human normal pancreatic tissue and pancreatic cancer tissue was extracted respectively, different gene expression level of prohibitin 2 was studied by real-time PCR.

RESULTSNumber 5 and 6 positive dots were identified as prohibitin 2 and prohibitin. RT-PCR and real-time PCR all showed that gene of prohibitin 2 and prohibitin were expressed in the human pancreatic cancer cell line SW1990, AsPc and P3 respectively, especially in P3 cell with highest expression (t = 7.442, P < 0.01). In addition, gene expression level of prohibitin 2 was significant higher in human pancreatic cancer than that of normal pancreatic tissue (t = 0.893, P < 0.01).

CONCLUSIONSProhibitin 2 and prohibitin are both differently expressed in the pancreatic cancer cell line SW1990, AsPc and P3. Prohibitin 2 is obvious highly expressed in human pancreatic cancer tissue. Prohibitin 2 and prohibitin might be the candidate immunogenic membrane antigens of human pancreatic cancer.

Antigens, Neoplasm ; genetics ; metabolism ; Cell Line, Tumor ; Gene Expression Regulation, Neoplastic ; Humans ; Membrane Proteins ; genetics ; metabolism ; Pancreatic Neoplasms ; genetics ; immunology ; metabolism ; Proteomics ; methods ; Repressor Proteins ; genetics ; metabolism

OBJECTIVETo verify the obtained immunogenic membrane antigens candidate of pancreatic cancer in the performed research.

METHODSPancreatic cancer cell line SW1990 membrane protein underwent immunoblot with serum IgG purified from clinically collected sera of 66 pancreatic cancer patients. Number 3 and number 8 positive dots of immunoblot were identified by MALDI-TOF mass spectrometry and peptide mass fingerprinting matching. The candidate membrane antigens were further validated in cell lines by RT-PCR, real-time PCR and Western blot, and their different expression level of gene and protein in pancreatic cancer cell lines were contrastly studied.

RESULTSNumber 3 and number 8 positive dots were identified as: voltage-dependent anion channel (VDAC3) and catechol-o-methyltransferase (COMT). RT-PCR, real-time PCR and Western blot showed that gene and protein of VDAC3 and COMT were expressed in the pancreatic cancer cell line SW1990, AsPc and P3 respectively.

CONCLUSIONVDAC3 and COMT might be the candidate immunogenic membrane antigens of human pancreatic cancer, and their gene and protein are differently expressed in the pancreatic cancer cell line SW1990, AsPc and P3.

Antigens, Neoplasm ; analysis ; Cell Line, Tumor ; Humans ; Pancreatic Neoplasms ; immunology ; Proteomics

OBJECTIVETo investigate the genes concerning multidrug resistance (MDR) of pancreatic ductal adenocarcinoma with microarray analysis.

METHODSGene expression profile of pancreatic cancer cell line SW1990 and resistance subline SW1990/5-FU, SW1990/ADM, SW1990/GEM were screened in two independent replicates using oligonucleotide microarray (Affymetrix HG U133 2.0 plus) which contained 38,500 human genes. And advanced bioinformatics analysis was conducted.

RESULTSTotally, 165 genes and expressed sequence tags (ESTs), which were seldom reported to be related with drug resistance before,were statistically difference and the fold change was up- or down-regulated at least 2 folds in all 3 resistant sub-lines when compared with SW1990. According gene ontology, the genes related to oxidoreductase activity, apoptosis, cell cycle, signal transduction and cell adhesion might be some epigenetic changes for MDR development. Hierarchical clustering analysis, showed several interesting clusters, namely, TNKS2, PRDX4 and CCDC4.

CONCLUSIONSMDR of pancreatic cancer is a complicated and multifactorial process. In the present study, a widespread differential gene expression pattern was constructed in PDAC multidrug resistant cells. Advanced study will provide new targets for MDR research and cast insights into research of the molecular mechanism of MDR.

Adenocarcinoma ; genetics ; pathology ; Carcinoma, Pancreatic Ductal ; genetics ; pathology ; Cell Line, Tumor ; Cluster Analysis ; Drug Resistance, Multiple ; genetics ; Drug Resistance, Neoplasm ; genetics ; Drug Screening Assays, Antitumor ; Gene Expression Profiling ; methods ; Gene Expression Regulation, Neoplastic ; Humans ; Oligonucleotide Array Sequence Analysis ; Pancreatic Neoplasms ; genetics ; pathology

BACKGROUNDChemotherapy is the most frequently adopted adjuvant therapy of pancreatic ductal adenocarcinoma (PDAC), but the development of drug resistance reduces its effectiveness. Clarification of the mechanism of multidrug resistance (MDR) development in PDAC is needed to improve the therapeutic effect of chemotherapy. This study was aimed to investigate the molecular mechanism of MDR of PDAC and to identify genes associated with MDR development.

METHODSThe gene expression profiles of cell line SW1990 and three drug-selected pancreatic chemoresistant sub-lines, SW1990/5-Fu, SW1990/ADM and SW1990/GEM, were obtained using an oligonucleotide microarray (Affymetrix HG U133 2.0 plus) that contained approximately 38,000 human genes. The microarray results were validated by real-time quantitative polymerase chain reaction and Western blot analysis.

RESULTSThere were 165 genes and expressed sequence tags, some of which have never been linked to drug resistance, that were up- or down-regulated at least 2-fold in all resistant sub-lines when compared with SW1990. According to Gene Ontology annotation, differentially expressed genes related to MDR in pancreatic cancer belong to many functional families and with diverse biological processes. Genes related to antioxidant activity, apoptosis, the cell cycle, signal transduction and intracellular adhesion may undergo epigenetic changes preceding MDR development. A hierarchical clustering was conducted and several interesting clusters were discovered that may be primarily related to cell cycle and developmental regulation. A prediction rule was built from the expression profiles of 117 genes after support vector machine (SVM) analysis, and the prediction result was examined by cytotoxic testing. As a result, a differential gene expression pattern was constructed in multidrug resistant pancreatic cancer cells.

CONCLUSIONSThe findings of this study prove that construction of a chemoresistance prediction rule, based on gene expression patterns, is practical. These data provide new insights into the molecular mechanism of pancreatic cancer MDR development and may be useful for the detection and treatment of MDR in pancreatic cancer patients.

Cell Cycle Proteins ; genetics ; Cell Line, Tumor ; Computational Biology ; Drug Resistance, Multiple ; Drug Resistance, Neoplasm ; Gene Expression Profiling ; Glutathione Peroxidase ; genetics ; Glutathione Transferase ; genetics ; Humans ; Microtubule-Associated Proteins ; genetics ; Oligonucleotide Array Sequence Analysis ; methods ; Pancreatic Neoplasms ; drug therapy ; genetics ; Tankyrases ; genetics

OBJECTIVETo study the expression and significance of GCS gene in human pancreatic cancer cell line SW1990 and its drug-resistant sublines.

METHODSSW1990 and its drug-resistant sublines, SW1990/FU, SW1990/ADM and SW1990/GEM were cultured in vitro. CCK-8 (Cell Counting kit-8) was used to detect the drug resistance of the sublines. Relative quantitation of GCS mRNA expression was evaluated by real-time PCR and Western blot was adopted to evaluate the expression of GCS protein.

RESULTSThe drug resistance indexes of SW1990/FU, SW1990/ADM and SW1990/GEM were 339.7, 11.9 and 56.6, respectively. The GCS mRNA and protein were expressed in all SW1990 and its drug-resistant sublines. There was a higher expression of GCS mRNA in all the sublines and a significant difference of GCS protein expression was detected in SW1990/ADM and SW1990/GEM compared with SW1990.

CONCLUSIONSGCS gene is expressed in SW1990 and its drug-resistance sublines. The high expression of GCS protein in SW1990/ADM and SW1990/GEM might be one reason of resistance to ADM and GEM in the sublines.

Cell Line, Tumor ; Drug Resistance, Neoplasm ; genetics ; Gene Expression ; Glucosyltransferases ; biosynthesis ; genetics ; Humans ; Immunoblotting ; Pancreatic Neoplasms ; enzymology ; genetics ; pathology ; RNA, Messenger ; genetics ; Reverse Transcriptase Polymerase Chain Reaction

Child ; Humans ; Nephrotic Syndrome ; diagnosis ; physiopathology ; urine ; Proteinuria ; urine ; Severity of Illness Index

OBJECTIVESTo evaluate the safety and midterm efficacy of stent revascularization as treatment for renal artery stenosis.

METHODSPercutaneous transluminal renal angioplasty with stent (PTRA) was performed because of poorly controlled hypertension or preservation of renal function in 150 consecutive patients with severe renal artery stenosis, caused by atheroma (96 patients), arteritis (44 patients) and fibromuscular dysplasia (10 patients). All of them subsequently underwent 6-month clinical follow-up to observe the effect of the procedure on renal function, blood pressure control, number of antihypertensive medications.

RESULTAngiographic success was obtained in 148 (98.7%) of 150 patients after PTRA. At 6 months, both systolic and diastolic blood pressures significantly decreased (from 169.6 to 142.7 mm Hg and from 97.3 to 83.3 mm Hg, respectively; P < 0.001), and less antihypertensive medication was taken (from 2.7 to 1.9). The blood pressure became normal without taking any antihypertensive medications in 48 of 150 patients (32.0%), and the blood pressure control was more facile in 78 patients (52.0%), however, there were no improvement in 22 patients (16.0%). Creatinine level decreased in 34 patients (22.7%), remained stable in 112 patients (74.6%), and increased in 4 (2.7%). There was no statistical significance. No deaths occurred during 6-months follow-up.

CONCLUSIONSRenal artery stent revascularization had a beneficial effect on blood pressure control and a nondeleterious effect on renal function during 6-months follow-up. The long-term efficacy should be investigated. The procedure is safe in usual.

Adolescent ; Adult ; Aged ; Aged, 80 and over ; Female ; Follow-Up Studies ; Humans ; Kidney Function Tests ; Male ; Middle Aged ; Renal Artery ; surgery ; Renal Artery Obstruction ; surgery ; Stents ; Treatment Outcome ; Young Adult