Search

Gelsenicine, mainly isolated from Gelsemium elegans Benth., is one of the most toxic alkaloids. The lack of information on gelsenicine leads to inaccurate risk and poisoning evaluation. In this study, the metabolic profiling and toxicokinetics of gelsenicine was studied by ultra-high performance liquid chromatography (UPLC) with quadrupole time-of-flight (Q-ToF) and tandem mass spectrometry in rats after intraperitoneal (i.p., 40 μg/kg) and intragastric (i.g., 60 μg/kg) administration. After i.p. administration, the area under the curve (AUC), the apparent volume of distribution (V), and the total body clearance (CL/F) of gelsenicine in plasma were 3.79 μg/L h, 38.47 L/kg, and 11.87 mL/h kg, respectively. After i.g. administration, the corresponding values were slightly increased (5.49 μg/L h; 53.10 mL/kg, and 12.66 mL/h kg). The toxicokinetic results indicated that the hepatic first-pass effect was predominant after i.p. administration. The UPLC-Q-ToF-MS data revealed nine metabolites in plasma, urine, and bile which were largely obtained by demethylation, hydroxylation, acetylation and glycine conjugation. Metabolites were mainly excreted through urine and bile, most of which in urine was basically eliminated in 24 h. Molecular docking and liver microsome experiments further showed that gelsenicine was metabolized by cytochrome P450 3A4 and 3A5. Summarizing, the present study provides metabolic and toxicokinetic information on gelsenicine which in turn may help in future risk assessment and forensic identification after poisonings.

Introduction: Premature infants are exceptionally vulnerable to nutrition-related diseases, and the utilization of standardized feeding guidelines may reduce nutritional practice variation, which can promote growth. Nutritional risk screening is the first step for standardized nutrition advice. However, risk screening tools specific for following up preterm infants are scarce. Hence, our study aimed to develop and evaluate a standardized Nutritional Risk Screening Tool for Preterm Infants (NRSP subscale 1) from birth to corrected age four months old . Methods: This study was a two-phase (the development phase and evaluation phase) study. Initially, we used the Delphi expert consultation method to create NRSP subscale 1. Then, a professional panel interviewed the participated preterm infants using the screening tool, measured anthropometric parameters, and conducted an intellectual development test on the interview day and remeasured anthropometric parameters 2 weeks or 1 month after the first interview. In the development phase, we cross-tabulated the responses to the screening tool with the classifications of z-scores of the body weight, length, or head circumference to identify significant predictors of underweight, stunting, or microcephaly. We then combined significant predictors to produce models for predicting underweight, stunting, or microcephaly by multivariate logistic regression analysis. In the evaluation phase, the area under the curve (AUC), sensitivity, specificity, and correlation coefficient by Spearman’s correlation analysis (rs) between the risk classifications by NRSP subscale 1 and the classifications of the z-scores of the body weight, length, or head circumference were calculated to assess the validity of the screening tool. Intellectual development levels between high and low nutritional risk infants were statistically compared. Results: A total of 219 and 244 preterm infants were included to two phases, respectively. AUC was 0.936 (95% CI: 0.860–1.000, p < 0.001), sensitivity was 0.667, specificity was 0.941, rs = 0.407 (p < 0.001); AUC was 0.794 (95% CI: 0.638–0.951, p = 0.002), sensitivity was 0.500, specificity was 0.953, rs = 0.339 (p < 0.001); AUC was 0.831 (95% CI: 0.737–0.925, p = 0.001), sensitivity was 0.889, specificity was 0.643, rs = 0.215 (p = 0.001) in predicting underweight, stunting, and microcephaly on the interview day, respectively. AUC was 0.905 (95% CI: 0.826–0.984, p = 0.006), sensitivity was 0.500, specificity was 0.905, rs = 0.504 (p < 0.001); AUC was 0.738 (95% CI: 0.515–0.960, p = 0.034), sensitivity was 0.429, specificity was 0.848, rs = 0.382 (p < 0.001); AUC was 0.664 (95% CI: 0.472–0.856, p = 0.071), sensitivity was 0.455, specificity was 0.809, rs = 0.169 (p = 0.037) in predicting underweight, stunting, and microcephaly 2 weeks or 1 month after the first interview, respectively. Gross motor development quotients (DQs) (95.85 [32.87] vs. 86.29 [17.19], p = 0.022), fine motor DQs (115.77 [46.03] vs. 102.12 [20.27], p = 0.010), and verbal DQs (110.73 [35.27] vs. 100.63 [21.28], p = 0.042) were higher in low nutritional risk infants than high-risk ones. Conclusion: NRSP subscale 1 was acceptable and reliable in predicting underweight, but the validity in predicting stunting or microcephaly was slightly mild. Further investigations are required to authenticate NRSP subscale 1’s effectiveness.

Beige adipocytes have recently attracted attention for their potential as new therapeutic targets in the management of obesity and related metabolic disorders. MicroRNAs (miRNAs) have been reported as transcriptional regulators or biomarkers of brown and beige adipogenesis. Nevertheless, the effects of miRNAs involved in beige differentiation of human visceral adipocytes remain to be investigated. In this study, microarray screening showed that miR-1275 was significantly decreased during the differentiation of beige adipocytes induced by human omental adipose-derived stem cells (hASCs). Overexpression of miR-1275 suppressed the "brown-like" differentiation of hASCs by inhibiting the key transcriptional factor PR domain containing 16 (

Gestational diabetes mellitus (GDM) is a pathological condition of glucose intolerance associated with adverse pregnancy outcomes and increased risk of developing maternal type 2 diabetes later in life. Metabolomics is finding increasing use in the study of GDM. To date, GDM-specific metabolomic changes have not been completely elucidated.In this pilot study, metabolomics fingerprinting data, obtained by ultra-performance liquid chromatography quadrupole time-of-flight mass spectrometry (UHPLC/Q-TOF-MS), of 54 healthy pregnant women and 49 patients with GDM at the second and third gestational trimesters were analyzed. Multilevel statistical methods were used to process complex metabolomic data from the retrospective cohorts.Using univariate analysis (p 0.05), 41 metabolites were identified as having the most significant differences between these two groups. Lipid metabolites, particularly glycerophospholipids, were the most prevalent class of altered compounds. In addition, metabolites with previously unknown connection to GDM - such as monoacylglycerol, dihydrobiopterin, and 13S-hydroxyoctadecadienoic acid - were identified with strong discriminative power. The main metabolic pathways affected by GDM included glycerophospholipid metabolism, linoleic acid metabolism, and D-arginine and D-ornithine metabolism.Our data provide a comprehensive overview of metabolite changes at different stages of pregnancy, which offers further insights into the pathogenesis of GDM.

The clinical pharmacodynamics of tacrolimus in renal transplant patients has significant interindividual variability. T lymphocytes were selected to study the pharmacodynamic response of tacrolimus, which was significantly correlated with renal function and the outcome of renal transplant patients. Ultra-performance liquid chromatography–quadrupole time-of-flight mass spectroscopy (UPLC/Q-TOF-MS) was performed to obtain the metabolic profiles of 109 renal transplant patients. A partial least squares (PLS) model was constructed to screen potential biomarkers that could predict the efficacy of tacrolimus. Multinomial logistic regression analysis established a bridge that could quantify the relationship between the efficacy of tacrolimus and biomarkers. The results showed a good correlation between endogenous molecules and the efficacy of tacrolimus. Metabolites such as serum creatinine, mesobilirubinogen, L-isoleucine, 5-methoxyindoleacetate, eicosapentaenoic acid, N2-succinoylarginine, tryptophyl-arginine, and butyric acid were indicated as candidate biomarkers. In addition, the key biomarkers could correctly predict the efficacy of tacrolimus with an accuracy of 82.5%. Finally, we explored the mechanism of individual variation by pathway analysis, which showed that amino acid metabolism was significantly related to the efficacy of tacrolimus. Moreover, orthogonal partial least squares discriminant analysis (OPLS-DA) showed that there was no difference in key metabolites among different pharmacodynamic groups at 1 month and 3 months after dose adjustment, suggesting that pharmacometabonomics is a useful tool to predict individual differences in pharmacodynamics and thus to facilitate individualized drug therapy.

The aim of this study is to evaluate morphological features of corneal flap/cap and the correlations with corneal higher-order aberrations (HOAs) changes after femtosecond laser in situ keratomileusis (FS-LASIK) and small incision lenticule extraction (SMILE).This was a retrospective study. Pre- and postoperative (1 and 3 months) corneal HOAs were assessed with Pentacam HR. The corneal flap/cap thickness at 32 points (± 1.5 mm, ± 2 mm, ± 2.5 mm, and ± 3 mm from the corneal vertex on meridian 0°/45°/90°/135°) were measured using anterior segment optical coherence tomography at 3 months postoperatively. Morphological features of corneal flap/cap including predictability (P), uniformity (U), and symmetry (S) were calculated and used for correlation analysis with corneal HOAs changes.Eighty-six eyes (44 patients) and ninety-six eyes (50 patients) were involved in FS-LASIK and SMILE groups, respectively. Significant thicker corneal flap/cap than the predicted was observed at each measuring point and meridian in both groups (difference 2.225 μm, the within-subject standard deviation over 6-mm optical zone). There was no statistically significant difference in predictability of corneal flap/cap thickness, while UBoth FS-LASIK and SMILE had good predictability in flap or cap thickness, while the uniformity and symmetry of SMILE cap were better than FS-LASIK flap. The quality of flap/cap was closely associated with the changes of corneal HOAs.

Mammary epithelial cells (MECs) are specific target cells to study the underlying mechanism for mammary gland development and lactation, mammary gland bioreactor and breast cancer pathogenesis. However, the conventional isolation and purification methods of MECs have restricted the research and application of its related fields, because the isolated MECs population was always mixed with other cell types.In order to solve the problem of incomplete purification of MECs isolated in vitro, it is very necessary to establish the optimal culture method for its isolation and purification.In this study, a method were called the "selective secondary tissue attachment method", which were proved to be effective and accurate by the purity, cellular viability, biological characteristic, and transgenic efficiency of the purified MECs.The results showed that compared with traditional methods, this method could obtain 100% MECs population with high purity and high cell viability in vitro, and these purified cells showed high transgenic efficiency and a high number of positive clones. In addition, the early embryo development rate could significantly improve when the purified mammary epithelial cells were used as donor cells for nuclear transfer.Therefore, this study provides a versatile and effective method for the isolation of MECs with high purity, which can be used further for mammary-related research.It is most effective approach to isolate mammary epithelial cells by "selective secondary tissue attachment method".

Polycystic ovary syndrome (PCOS) is a complex reproductive endocrine disease characterized by polycystic ovary. The aim of the study was to assess microRNA-21 regulates granulosa cell apoptosis and proliferation in polycystic ovary syndrome through target toll-like receptor 8. Granulosa cells were collected from 30 PCOS patients and 30 normal patients with tubal or male factor infertility (control) during in vitro fertilization-Embryo Transfer (IVF-ET) and were flash frozen with liquid nitrogen for storage for subsequent use. PCOS diagnosis was based on the revised standards of the American Society of Reproductive Medicine (ASRM) and the Rotterdam criteria PCOS granulosa cells and control granulosa cells were cultured in DMEM/F12 medium containing 10% fetal bovine serum and 1% antibiotic. After this RT-PCR, Western blot assessment and Detection of apoptosis by flow cytometry were conducted. The results of qPCR showed that the mRNA and protein expression of TLR8 in PCOS granulosa cells were significantly increased compared with the normal group. The results of Western blot also showed that the expression of TLR8, IFN-γ, TNF-α and IL-12 gene protein in the transfected cells was significantly higher than that in the control cells. Here, we show that miR-21 and TLR8 significantly increased in PCOS granulosa cell as compared with normal granulosa cells, and miR-21 enhances the TLR8 mRNA translation and then promotes the IFN-γ, TNF-α, and IL-12 secretion. Our study demonstrates that miR-21/ TLR8 involved in the PCOS inflammation, it provides profound insights into pathogenesis of PCOS.

BackgroundAntenatal corticosteroids (ACS) treatment is critical to support survival and lung maturation in preterm infants, however, its effect on feeding and growth is unclear. Prior preterm delivery, it remains uncertain whether ACS treatment should be continued if possible (repeated course ACS), until a certain gestational age is reached. We hypothesized that the association of single-course ACS with feeding competence and postnatal growth outcomes might be different from that of repeated course ACS in very-low-birth-weight preterm infants.MethodsA multicenter retrospective cohort study was conducted in very-low-birth-weight preterm infants born at 23–37 weeks’ gestation in South China from 2011 to 2014. Data on growth, nutritional and clinical outcomes were collected. Repeated course ACS was defined in this study as two or more courses ACS (more than single-course). Infants were stratified by gestational age (GA), including GA ResultsA total of 841 infants were recruited. The results, just in very-low-birth-weight preterm infants born at 28–32 weeks’ gestation, showed both single and repeated course of ACS regimens had shorter intubated ventilation time compared to non-ACS regimen. Single-course ACS promoted the earlier application of amino acid and enteral nutrition, and higher rate of weight increase (15.71; 95%CI 5.54–25.88) than non-ACS after adjusting for potential confounding factors. No associations of repeated course ACS with feeding, mean weight and weight increase rate were observed.ConclusionsSingle-course ACS was positively related to feeding and growth outcomes in very-low-birth-weight preterm infants born at 28–32 weeks’ gestation. However, the similar phenomenon was not observed in the repeated course of ACS regimen.

Early pregnancy loss (EPL) is a common event worldwide. Previous studies show that mitochondrial DNA (mtDNA) copy number (CN) is associated with semen parameters and preimplantation embryo viability, indicating the predictive potential of mtDNA CN for ongoing pregnancy outcomes. However, no relevant study has assessed the relationship between mtDNA CN and EPL. Thus, we aimed to determine whether mtDNA CN and mtDNA 4977-bp deletion rate (DR) in chorionic villous tissue are associated with EPL. Chorionic villous tissue total DNA was extracted from 75 EPL cases and 75 healthy controls. Chromosomal analysis was conducted using copy number variation (CNV) sequencing. The mtDNA CN and DR were measured in samples without pathogenic CNVs. The association between mtDNA CN or DR and EPL risk were estimated using logistic regression. The EPL group had a significantly different mtDNA CN (P 0.001) and DR (P = 0.005) compared to the control group. Both biomarkers were independent risk factors for EPL (CN odds ratio 1.71, 95% confidence interval 1.17 to 2.49, P = 0.005; DR odds ratio 1.07, 95% confidence interval 1.02 to 1.12, P = 0.006). These results suggest that higher mtDNA CN and DR levels are strongly associated with EPL and represent independent risk factors for EPL. Further studies validating these findings and exploring the underlying biological mechanisms are warranted.

Background A complementary feeding (CF) period is necessary for nutritional and developmental reasons. Preterm children encounter more feeding problems than their term counterparts in the CF period. The goal of this study was to develop a nutritional risk screening tool specific to preterm children (the NRSP) in outpatient settings in the CF period, with the expectation of providing a standardised process to determine feeding problems and subsequently offering targeted nutritional advice. Methods This study was a 2-phase study consisting of the development and evaluation phases. In the development phase, the items of the NRSP were initially developed based on references and the Delphi expert consultation method. Second, 329 preterm individuals with corrected ages from 5 to 36 months were enrolled. The participating preterm children were interviewed with the NRSP and anthropometric measurements, and underwent intellectual developmental tests and biochemistry detection (haemoglobin, red blood cell count, mean corpuscular volume, mean corpuscular haemoglobin, mean corpuscular haemoglobin concentration, serum iron, vitamin D). Third, preterm children’s anthropometric parameters were remeasured 1 month (for infants whose corrected age was 5–11 months) or 3 months (for children whose corrected age was 12–36 months) after the interview. Data in the development phase were analysed via univariate and binary logistic regression analysis sequentially to assign scores for items of the NRSP and to generate the models to predict underweight, stunting, and microcephaly of the NRSP. In the evaluation phase, another 605 preterm individuals were recruited to undergo the interview, anthropometric measurements, intellectual developmental tests, and biochemistry detection as in the development phase. Interrater reliability, test-retest reliability, area under the curve (AUC), accuracy, sensitivity, specificity, the positive/negative predictive value (P/NPV), the positive/negative likelihood ratio (LR+/−), and the correlation coefficient by Spearman’s correlation analysis (rs) were used to assess the reliability and validity of the NRSP. Finally, anthropometric parameters, biochemistry levels, and intellectual development quotients (DQs) from the development and evaluation phases between the high- and low-risk groups classified by the NRSP were compared using a t-test. Results The κ coefficients of the interrater and test-retest reliability of the NRSP were all above 0.600, which meant that the reliability of the NRSP was moderate to substantial. The NRSP exhibited relatively higher efficiency in predicting underweight and stunting, with AUCs, accuracies, specificities, and NPVs near to or greater than 0.900, sensitivities above 0.600, PPVs above 0.400, LR + s near to or greater than 10, and rss above 0.400. On the other hand, the NRSP manifested a weaker ability in predicting microcephaly, with most of the values of validity indicators lower than those of underweight and stunting prediction. Z scores of body weight, body length and head circumference, as well as DQs, were all higher in the low-risk groups than in the high-risk groups. There were no significant differences with respect to biochemistry levels between the high- and low-risk groups. Conclusion The NRSP shows moderate to substantial reliability and validity in predicting underweight, stunting, and microcephaly. Health care staff should shed light on improving the feeding practices of preterm children with high nutritional risk classified by the NRSP to facilitate their physical growth and intellectual development. More research is expected to promote the NRSP models.

Early non-chromosome-related missed abortion (MA) is commonly associated with an altered immunological environment during pregnancy. Human decidual natural killer (dNK) cells, the most abundant lymphocyte population within the first-trimester maternal-fetal interface, are vital maternal regulators of immune tolerance mediating successful embryo implantation and placentation. Previous studies have shown that dNK cells may play a role in MA. However, the gene expression status and specific altered manifestations of dNK cells in patients with early MA remain largely unknown. Here, we show that MA dNK cells have distinct mRNA and lncRNA expression profiles through RNA sequencing, with a total of 276 mRNAs and 67 lncRNAs being differentially expressed compared with controls. Protein-protein interaction analysis of differentially expressed mRNAs was performed to identify hub genes and key modules. An lncRNA-mRNA regulatory network characterized by the small-world property was constructed to reveal the regulation of mRNA transcription by differential hub lncRNAs. Functional annotation of differentially expressed mRNAs and lncRNAs was performed to disclose their potential roles in MA pathogenesis. Our data highlight several enriched biological processes (immune response, inflammatory response, cell adhesion, and extracellular matrix [ECM] organization) and signaling pathways (cytokine-cytokine receptor interaction, ECM-receptor interaction, Toll-like receptor signaling pathway, and phosphatidylinositol signaling system) that may influence MA. This study is the first to demonstrate the involvement of altered mRNA and lncRNA expression profiles in the dNK cell pathogenesis of early MA, facilitating a better understanding of the underlying molecular mechanisms and the development of novel MA therapeutic strategies targeting key mRNAs and lncRNAs.

We previously demonstrated that Tetraticopeptide 4 (TTC4) inhibited apoptosis in vascular endothelial cells (VEC) deprived of serum and fibroblast growth factor 2 (FGF-2). In this study, we aimed to resolve the mechanism of TTC4 inhibiting VEC apoptosis. TTC4, predicted as a HSP70 co-chaperone protein, may regulate the fate of cells by affecting the activity of HSP70, however, there is no experimental evidence showing the interaction of TTC4 and HSP70. Using Co-immunoprecipitation (Co-IP), we demonstrated that TTC4 interacted with HSP70. If HSP70 was knockdown, TTC4 no longer suppressed apoptosis. Furthermore, we found ABO, an inhibitor of annexin A7 (ANXA7) GTPase, could promote the interaction of TTC4 and HSP70 and the translocation of ANXA7 to lysosome. At the same time, ABO inhibited the interaction of HSP70 and ANXA7. Moreover, Akt, as a downstream effector of HSP70 was upregulated, and ANXA7 translocating to lysosome protected the stability of lysosomal membrane. Here, we discovered a special mechanism by which TTC4 inhibited apoptosis via HSP70 in VECs. On the one hand, increasing TTC4 and HSP70 interaction upregulated Akt that inhibited apoptosis. On the other hand, decreasing HSP70 and ANXA7 interaction promoted the translocation of ANXA7 to lysosome, which inhibited apoptosis through protecting the lysosomal membrane stability.

Studies have shown that the use of artificial intelligence can reduce errors in medical image assessment. The diagnosis of breast cancer is an essential task; however, diagnosis can include ‘detection’ and ‘interpretation’ errors. Studies to reduce these errors have shown the feasibility of using convolution neural networks (CNNs). This narrative review presents recent studies in diagnosing mammographic malignancy investigating the accuracy and reliability of these CNNs. Databases including ScienceDirect, PubMed, MEDLINE, British Medical Journal and Medscape were searched using the terms ‘convolutional neural network or artificial intelligence’, ‘breast neoplasms [MeSH] or breast cancer or breast carcinoma’ and ‘mammography [MeSH Terms]’. Articles collected were screened under the inclusion and exclusion criteria, accounting for the publication date and exclusive use of mammography images, and included only literature in English. After extracting data, results were compared and discussed. This review included 33 studies and identified four recurring categories of studies: the differentiation of benign and malignant masses, the localisation of masses, cancer‐containing and cancer‐free breast tissue differentiation and breast classification based on breast density. CNN's application in detecting malignancy in mammography appears promising but requires further standardised investigations before potentially becoming an integral part of the diagnostic routine in mammography., This article explores the current research and developments in using convolutional neural networks for mammography diagnosis. Current developments are focused on four distinct categories; however, limitations of this artificial intelligence withhold it from clinical implementation.

A new mitochondria-targeted fluorescent probe RBC, constructed using a coumarin moiety which was selected as the donor and a benzothiazole derivative as the acceptor, for SO2 derivatives (HSO3-/SO32-) was presented. The probe designed on a new FRET platform showed high selectivity and a low detection limit. Importantly, the probe could respond to HSO3-/SO32- within 35 s. Furthermore, the probe could target mitochondria and was successfully used for fluorescence imaging of endogenous bisulfite in HepG2 with low cytotoxicity, which significantly assisted in cancer diagnosis.

Our previous study revealed that Homeobox containing 1 (HMBOX1), essential for the survival of vascular endothelial cells (VECs), was involved in the progression of atherosclerosis. Knockdown of HMBOX1 promoted apoptosis and inhibited autophagy through regulating intracellular free zinc level in cultured VECs. In current study, in order to investigate the roles of HMBOX1 in vivo and in endothelium, we generated a knockout (KO) mouse for HMBOX1 by using transcription activator-like effector nucleases (TALENs) technology. Herein, we reported that the protein level of HMBOX1 was gradually increased during mouse development. The HMBOX1 KO mouse was viable and fertile. There existed no differences in apoptosis and autophagy of aortic endothelial cells between wild type and KO mouse. Whereas, loss of HMBOX1 promoted apoptosis and inhibited autophagy of aortic endothelial cells under lipopolysaccharide (LPS) stimulation in mouse. We also demonstrated that HMBOX1 deletion had no influence on the secretion of inflammatory cytokines TNF-α and IL-6. Moreover, overexpression or knockdown of HMBOX1 failed to regulate multiple pro-apoptotic genes expression in vitro. In conclusion, HMBOX1 participated in the functional maintenance of mouse aortic endothelial cells, the aortic endothelial cells of HMBOX1 KO mouse showed increased apoptosis and decreased autophagy with LPS treatment.

Anemia is associated with neurodevelopmental delays and brain injury in infants and toddlers, but whether early anemia has a similar effect in neonatal preterm infants is largely unknown. Thus, this study aimed to determine the relationship of early anemia with neurodevelopment and brain injury in very-low-birth-weight (VLBW) preterm infants within the neonatal period. A prospective cohort study including 110 VLBW preterm infants was conducted in Southern China from 2016 to 2018. All participants were followed from birth to 1 month corrected age. Early anemia is defined as hemoglobin of ≤145 g/L within the first week after birth. The non-anemic group (control group, N = 55) was 1:1 matched with the early anemia group (N = 55) according to birth weight and gestational age. Neurodevelopment at 1 month corrected age and brain injury within 1 month corrected age were measured by neonatal behavioral neurological assessments (NBNA) and cranial ultrasound, respectively. Compared to the control group, the early anemia group had a lower score in behavioral ability in the NBNA test [11 (10–12) vs. 10 (9.5–11), p = 0.033]. Early anemia was negatively associated with the NBNA total score (β= −0.680, 95% CI: −1.300, −0.059), especially with the behavioral ability score (β= −0.504, 95% CI: −0.941, −0.067) after adjusting for the confounders. However, no association between early anemia and brain injury was observed. In conclusion, in VLBW preterm infants, early anemia is negatively correlated with neurodevelopment, especially with behavioral ability.

Age-related cell loss underpins many senescence-associated diseases. Apoptosis of lens epithelial cells (LECs) is the important cellular basis of senile cataract resulted from prolonged exposure to oxidative stress, although the specific mechanisms remain elusive. Our data indicated the concomitance of high autophagy activity, low SQSTM1/p62 protein level and apoptosis in the same LEC from senile cataract patients. Meanwhile, in primary cultured LECs model, more durable autophagy activation and more obvious p62 degradation under oxidative stress were observed in LECs from elder healthy donors, compared with that from young healthy donors. Using autophagy-deficiency HLE-B3 cell line, autophagy adaptor p62 was identified as the critical scaffold protein sustaining the pro-survival signaling PKCι-IKK-NF-κB cascades, which antagonized the pro-apoptotic signaling. Moreover, the pharmacological inhibitor of autophagy, 3-MA, significantly inhibited p62 degradation and rescued oxidative stress-induced apoptosis in elder LECs. Collectively, this study demonstrated that durable activation of autophagy promoted age-related cell death in LECs. Our work contributes to better understanding the pathogenesis of senescence-associated diseases.

Objectives The association of low or high birth weight (L/HBW) with obesity and hypertension in childhood remains unclear. We aimed to identify the secular trend of birth weight distribution and its relationship to obesity and hypertension in Southern Chinese children and adolescents. Methods 6,561 individuals (6–17-year-old) were enrolled by multistage cluster sampling to observed the trend of birth weight distribution and its associated factors. 1,218 were further selected by group matching to investigate the correlation between birth weight and obesity or hypertension. Results Between 1997 and 2008, a significant decline in the LBW rate and no significant change in the HBW rate was found. LBW was associated with maternal BMI2 (OR1.79, 95% CI 1.08–2.97) during pregnancy, while maternal BMI between 25.0 and 27.9 kg/m2 (OR1.62, 95% CI 1.04–2.52) and paternal BMI>28 kg/m2 (OR1.64, 95% CI 1.02–2.63) during pregnancy were associated with HBW. The prevalence of obesity was significantly higher with HBW than normal birth weight (NBW) or LBW (16.73, 6.25 and 5.50%, respectively). The prevalence rates of suspected hypertension were 1.62, 1.25 and 1.49% among LBW, NBW and HBW, respectively (p>0.05). LBW decreased the risks of childhood overweight (OR0.31, 95% CI 0.18–0.54), but had no effect on suspected hypertension. HBW increased the risks of childhood obesity (OR2.19, 95% CI 1.50–3.20), but decreased the risks of suspected hypertension (OR0.22, 95% CI 0.09–0.58). Conclusions HBW was positively associated with childhood obesity, and parental BMIs management might be one of the measurements to control birth weight to lessen childhood obesity.

The adipocyte precursors (APs) located in white adipose tissue (WAT) are functionally significant in adipose plasticity and browning. Modifying adipogenesis or WAT browning targeted on APs is a promising mechanism for anti-obesity drug. We herein explored the in vitro actions and mechanisms of glucose-dependent insulinotropic polypeptide (GIP), a gut-derived peptide, in human adipose-derived mesenchymal stem cells (hADSCs) isolated from omentum. The hADSCs were cotreated with 100 nM GIP with or without equimolar concentration of GIP3-42 (a GIP receptor antagonist), and subsequently examined in vitro. CCK-8, EdU incorporation, and flow cytometry assays were used to assess cellular proliferation. Annexin V FTIC/PI double stain, TUNEL staining, and Western blot were applied for apoptosis evaluation. Adipogenesis was reflected by Western blot, real-time PCR, Oil Red O staining, mitochondrial staining, and mitochondrial DNA analysis. Results showed that GIP promoted proliferation and inhibited apoptosis of hADSCs via pleiotropic effects. Besides, GIP facilitated de novo beige adipogenesis, by accelerating mitotic clonal expansion (MCE), upregulating core adipogenic regulators (C/EBPα and PPARγ), augmenting beige-related genes (UCP1, PGC1α, and PRDM16), increasing mitochondrial content and improving beige adipocyte functionalities. Above all, our study expands knowledge on the mechanisms of GIP modifying adipogenesis especially in inducing beige adipogenesis, and thus provides a theoretical support for clinical usage of GIP on obesity treatment.

To gain mechanistic insights into the functions and developmental dynamics of tumor-infiltrated immune cells, especially B-lymphocytes, here we combine single-cell RNA-sequencing and antigen receptor lineage analysis to characterize a large number of triple-negative breast cancer infiltrated immune cells and report a comprehensive atlas of tumor-infiltrated B-lymphocytes. The single-cell transcriptional profiles reveal significant heterogeneity in tumor-infiltrated B-cell subgroups. The single-cell antigen receptor analyses demonstrate that compared with those in peripheral blood, tumor-infiltrated B-cells have more mature and memory B-cell characteristics, higher clonality, more class switching recombination and somatic hypermutations. Combined analyses suggest local differentiation of infiltrated memory B-cells within breast tumors. The B-cell signatures based on the single-cell RNA-sequencing results are significantly associated with improved survival in breast tumor patients. Functional analyses of tumor-infiltrated B-cell populations suggest that mechanistically, B-cell subgroups may contribute to immunosurveillance through various pathways. Further dissection of tumor-infiltrated B-cell populations will provide valuable clues for tumor immunotherapy., Immune cells infiltrating the tumour microenvironment play critical roles in disease pathogenesis and the immune response. Here the authors present the characterisation of infiltrating B cells in breast tumours by the formation of an atlas created from paired RNA sequence and antigen receptor profiling.

PURPOSE To evaluate the influence of 2.2 mm clear corneal incision (CCI) features in surgically induced astigmatism (SIA) and higher-order aberrations (HOAs) after cataract surgery. METHODS Right eyes of 92 subjects receiving 2.2 mm incision cataract surgery were involved. A total of 38 eyes were categorized as the intact incision group, and 54 eyes were the defective incision group. Pre- and postoperative (1 month and 6 months) corneal astigmatism and HOAs on anterior and posterior corneal surfaces, corneal volume, and corneal thickness (CT) were measured using Pentacam. The CCI features including incision length (IL), incision angles, distance from incision to central cornea (Dis-En/Ex), and CT at incision site were quantified using AS-OCT. RESULTS The defective incision group showed shorter IL and larger incision angles [false discovery rate (FDR) - p

The lentivirus-short hairpin RNA (shRNA) system is a widely used tool for RNA interference. Multiple factors may affect the RNA interference efficiency during lentivirus production and transduction procedures. Thus, an optimized protocol is required to achieve high-titer lentivirus and efficient gene delivery. In the present study, lentivirus was produced by transfecting lentiviral transfer and packaging plasmids into HEK 293T cells. The factors affecting lentiviral titer were assessed, including lentiviral plasmid ratio, lentiviral transfer plasmid type, serum type for cell culture, transfection reagent-plasmid mixture incubation time, and the inoculation density of 293T cells for transfection. The high-titer lentivirus was achieved when plasmids were transfected at a molar ratio of 1:1:1:2, and the transfection reagent-plasmid mixture was replaced 6-8 h after transfection. The pLVX-shRNA2 lentiviral transfer plasmid was associated with the highest lentiviral titer, while both pLVX-shRNA2 and psi-LVRU6GP plasmids were associated with efficient RNA interference in target cells. The serum type for 293T cell culture affected the lentiviral titer significantly, while the inoculation density of 293T cells showed no influence on transfection efficiency or lentiviral titer. Moreover, the human primary fibroblasts infected with lentivirus, using the centrifugation method, achieved higher transduction efficiency than those infected with the non-centrifugation method. In conclusion, this study helped optimize lentiviral production and transduction procedures for more efficient gene delivery.

The miR-302s/367 family has the ability to induce mouse and human somatic cell reprogramming into induced pluripotent stem cells (iPSCs), inhibit the proliferation of several types of cancer cells, and even cause cancer cell apoptosis. However, the functions of the miR-302s/367 family in other mammals have not been explored. In the present study, the effects of miR-302s/367 on reprogramming, proliferation, and apoptosis in sheep fetal fibroblasts (SFFs) were evaluated by the delivery of a plasmid vector containing synthetic precursor miRNAs into cells, followed by the induction of mature miR-302s/367 expression. The results showed that miR-302s/367 could not reprogram SFFs into iPSCs; however, they could inhibit both the proliferation and apoptosis of SFFs by targeting CDK2, E2F1, E2F2, and PTEN in the cell cycle and PI3K-Akt pathways. Based on our findings, a novel mechanism was proposed in which the miR-302s/367 family functions in both the proliferation and apoptosis of somatic cells in mammals, suggesting that caution is needed when using miR-302s/367 as therapeutic agent.

Background We previously identified PIWIL1 as an oncogene involved in endometrial carcinogenesis. However, the mechanism of Piwil1 mediated regulation of tumorigenesis remains poorly understood. Methods The expression levels of target genes in endometrial cancer cells were detected by quantitative reverse transcription-PCR (RT-qPCR) and western blotting. Up- or down-regulation of ERα or PIWIL1 was achieved by transient transfection with expressing plasmids or short hairpin RNA (shRNA). Dual-luciferase reporter assays and chromatin immunoprecipitation (ChIP) were used to demonstrate the ERα bound to the half estrogen response element (half-ERE) located in PIWIL1 promoter. The expression of PIWIL1 and ERα in endometrial carcinoma tissues were investigated using immunohistochemistry and RT-qPCR. The proliferation ability of cancer cells were evaluated by MTT. Methylation status of the PIWIL1 promoter was detected by bisulfite sequencing PCR (BSP). Results In the present study, we found that PIWIL1 mediated E2-stimulated cancer cell proliferation. In ERα-positive endometrial cancer cells, we demonstrated that estrogen-ERα signaling significantly up-regulated the expression of PIWIL1, which was mediated by binding of the ERα onto the PIWIL1 promoter. Furthermore, we found that a half-ERE in the PIWIL1 promoter was essential for ERα binding. The PIWIL1 promoter was hypomethylated in ERα-positive endometrial cancer cells. Treatment with 5-aza-deoxycytidine (5-aza-dC) could up-regulate PIWIL1 expression. Conclusions These findings uncover a novel molecular mechanism by which estrogen-ERα signaling and DNA hypomethylation co-regulate PIWIL1 expression. These findings provide novel insights into the hormonal regulation of PIWIL1 in endometrial cancer and the PIWIL1’s role in estrogen-stimulated endometrial carcinogenesis.

To detect potential variant in an ethical Han Chinese pedigree affected with breast cancer.The proband and her relatives were subjected to next-generation sequencing using a target capture sequencing kit containing 121 cancer-related genes. Candidate variants were selected by analysis of their type, frequency in population, and segregation with the phenotype. Candidate variant was verified by Sanger sequencing and TA cloning.A c.2013_2014ins GT variant was detected in the BRCA1 gene among all breast cancer patients from this pedigree but not among healthy females. The variant was not recorded in the 1000 Genome Project database or the Exome Aggregation Consortium (ExAC) database. The frameshifting insertion was predicted to form an premature stop codon in gene transcript and can give rise to a truncated protein.The BRCA1 c.2013_2014ins GT variant probably underlies the pathogenesis of breast cancer in this Chinese pedigree.

ContextGraves’ orbitopathy (GO) causes infiltrative exophthalmos by inducing excessive proliferation, adipogenesis, and glycosaminoglycan production in orbital fibroblasts (OFs). Interference with OF autophagy is a potential therapy for proptosis.ObjectivesHere, we aimed to evaluate the effects of chloroquine (CQ) and hydroxychloroquine (HCQ), the autophagy inhibitors commonly used in clinical practice, on OFs.Design/Setting/ParticipantsOFs isolated from patients with GO (GO-OFs) or control individuals (non-GO-OFs) were cultured in proliferation medium (PM) or subjected to differentiation medium. OFs were treated with CQ or HCQ (0, 0.5, 2, and 10 μM), and subsequently examined in vitro.Main Outcome MeasuresCCK-8, EdU incorporation, and flow cytometry assays were used to assess cellular viability. Adipogenesis was assessed with Western blot analysis, real-time polymerase chain reaction (PCR) , and Oil Red O staining. Hyaluronan production was determined by real-time PCR and enzyme-linked immunosorbent assay. Autophagy flux was detected through red fluorescent protein (RFP)-green fluorescent protein (GFP)-LC3 fluorescence staining and Western blot analyses.ResultsCQ/HCQ halted proliferation and adipogenesis in GO-OFs in a concentration-dependent manner through blockage of autophagy, phenotypes that were not detected in non-GO-OFs. The inhibitory effect of CQ/HCQ on hyaluronan secretion of GO-OFs was also concentration dependent, mediated by downregulation of hyaluronan synthase 2 rather than hyaluronidases. Moreover, CQ (10 μM) induced GO-OF apoptosis without aggravating oxidative stress.ConclusionsThe antimalarials CQ/HCQ affect proliferation, adipogenesis, and hyaluronan generation in GO-OFs by inhibiting autophagy, providing evidence that they can be used to treat GO as autophagy inhibitors.

A promising near-infrared emissive and mitochondria-targeted fluorescence probe (SNB) for the ratiometric detection of sulfur dioxide derivatives with a novel reaction mechanism was developed on the basis of FRET and the ICT platform. Probe SNB showed favorable fluorescence properties, including an active NIR emission signal (671 nm), ultra-large Stokes shift (251 nm) and an ultra-broad emission band gap (193 nm). Furthermore, SNB was applied to detect sulfur dioxide derivatives with a low detection limit (17 nM), excellent sensitivity and exceptional selectivity. Of greater significance, SNB also successfully implemented the real-time and ratiometric surveillance for exogenous or endogenous sulfur dioxide derivatives in living HeLa, L-O2 and HepG2 cells. Interestingly, SNB possessed a subcellular organelle targeting ability with a preferred co-localization coefficient of 0.91 for mitochondria, which revealed its promising potential in the detection of subcellular sulfur dioxide derivatives.

Abdominal obesity is considered a major factor in the development of metabolic disorders. Glucagon-like peptide-1 (GLP-1) has been reported to have positive effects on improving body metabolism and to reducing insulin resistance. However, it remains less clear whether GLP-1 plays a role in the adipogenesis process of visceral fat. Here, we analyzed the in vitro actions and probable mechanisms of Exendin-4, a GLP-1 receptor agonist, on human adipose-derived stromal cells (hADSCs) isolated from omentum. Our results demonstrated that Exendin-4 improved cell viability via promoting proliferation and inhibiting apoptosis in hADSCs isolated from omentum. Mechanistically, the activation of MAPK/ ERK1/2, Akt/GSK-3β, and PKA/CREB pathways and downstream consequences induced are involved in the proliferative and anti-apoptotic roles of Exendin-4. More intriguingly, Exendin-4 could promote the differentiation of omental hADSCs. Underlying mechanisms of the differentiation of hADSCs are associated with the upregulation of the expression of pro-adipogenic genes and downregulation of the expression of anti-adipogenic genes. Our data demonstrate that Exendin-4 modifies adipogenesis of hADSCs isolated from omentum through multiple mechanisms, these effects could contribute to the protective actions of GLP-1 receptor agonist body metabolism and insulin sensitivity.

Aims/Introduction To explore angiopoietin‐like protein 8 (ANGPTL‐8) levels, and its association with hepatocellular lipid content (HCL) and insulin resistance in patients with different extents of non‐alcoholic fatty liver disease (NAFLD). Materials and Methods In 48 adults were recruited, of which 12 had no NAFLD (HCL < 5.5%; group 1), 18 had mild NAFLD (5.5% ≤ HCL < 10.0%; group 2) and 18 had moderate‐to‐severe NAFLD (HCL ≥ 10.0%; group 3). The peripheral insulin sensitivity of all participants was monitored by a hyperinsulinemic‐euglycemic clamp (M value), as well as the magnetic resonance image of HCL. Serum ANGPTL‐8, blood glucose levels and lipid profiles were also recorded in the study. Results Group 3 had a worse metabolic profile, and had the highest ANGPTL‐8 level (1,129 ± 351 pg/mL vs 742 ± 252 pg/mL, 765 ± 301 pg/mL, P = 0.001) compared with those in group 1 and group 2. In all metabolic profiles, HCL positively correlated the strongest with ANGPTL‐8 (r = 0.436, P = 0.042). Multivariate stepwise linear regression analysis showed ANGPTL‐8 and alanine aminotransferase were independent determinants of HCL (P = 0.002, P < 0.001, respectively), and these two indexes explained 67.4% of the variation of HCL (P < 0.001). Conclusions ANGPTL‐8 was positively correlated with hepatocellular lipid content independent of obesity and insulin resistance, indicating that ANGPTL‐8 might be a new and important important predictor of the severity of NAFLD.

Aims/Introduction Optimal glycemic targets during short-term intensive insulin therapy in patients with newly diagnosed type 2 diabetes are not standardized. The present study was carried out to determine the optimal glycemic targets during therapy by analyzing the impacts of glucose levels on therapeutic outcomes. Materials and Methods A total of 95 individuals with newly diagnosed type 2 diabetes were enrolled. Short-term intensive insulin therapy was carried out using an insulin pump to achieve and maintain glycemic targets (fasting blood glucose ≤6.0 mmol/L, 2-h postprandial blood glucose ≤7.8 mmol/L) for 14 days, with daily eight-point capillary blood glucose profiles recorded. Patients were followed up for 1 year after discharge. Results In most participants, the mean blood glucose and glycemic excursion parameters during the therapy were controlled within the normal range. Mean blood glucose was independently associated with amelioration of acute insulin response (r = −0.25, P = 0.015) and 1-year remission (odds ratio 0.12, 95% confidence interval 0.034–0.426), but negatively associated with more level 1 hypoglycemia (r = −0.34, P = 0.001), although major hypoglycemia was rare. Among mean blood glucose tertiles, patients in the middle (68.7%) and lower (75.0%) tertiles had a higher 1-year remission rate compared with the upper tertile (32.3%, both P < 0.001), whereas only the middle tertile did not have increased hypoglycemia compared with the upper tertile (8.1 ± 5.4 vs 7.2 ± 3.9 events/person, P = 0.48). Conclusions Stricter glycemic control during short-term intensive insulin therapy produced more remission despite self-manageable hypoglycemia. Based on glycemic parameters in the middle mean tertile, we propose new glycemic targets that are approximately 0.4 mmol/L lower than current the targets, as long-term benefit outweighs short-term risks.

Background: Little is known about the population pharmacokinetics (PPK) of daptomycin in kidney transplant patients. The present study established a pharmacokinetic model for daptomycin in kidney transplant patients in China and examinee the important factors affecting the pharmacokinetic parameters of daptomycin. Methods: The study population included 49 kidney transplant patients with 537 daptomycin concentrations. The PPK model of daptomycin was developed using a nonlinear mixed-effects model, a two-compartment structural model, and a mixed residual error model. The stability and predictive ability of the final model were evaluated based on bootstrapping, visual prediction checks and normalized prediction distribution errors. Results: Glomerular filtration rate (GFR) and total body weight significantly affected clearance, and body weight influenced the central volume of distribution. The average clearance of the population was 0.316 L/h, the central volume of distribution was 6.04 L, the intercompartmental clearance was 2.31 L/h, and the peripheral volume of distribution was 2.46 L. Based on the established model and the target of area under curve (AUC0-24h)/minimum inhibition concentration (MIC) ≥666, we developed a recommended dose regimen for kidney transplant patients according to their renal function and weight. The daily doses were 4.0±0.31, 4.7±0.36, 5.1±0.40, 5.5±0.43, 5.8±0.45, and 6.1±0.48 mg/kg when the GFRs were 15, 30, 45, 60, 75, and 90 ml/min/1.73 m2, respectively. Conclusion: This study provides a reference for individualized daptomycin administration in kidney transplant recipients, and it is a valuable resource for improving the treatment effect and reducing the toxic effects of daptomycin.

We explored whether red blood cell distribution width (RDW), a routinely checked item of complete blood cell counts, was an indicator of long-term euglycemia remission in patients with type 2 diabetes after short-term continuous subcutaneous insulin infusion (CSII).We analyzed the original data of patients enrolled in three randomized control trials from 2002 to 2014. CSII was administered to drug-naїve patients with newly diagnosed type 2 diabetes to achieve and maintain euglycemia for 2weeks.A total of 185 patients were involved and 98 patients (52.97%) who achieved and maintained euglycemia for at least 12months were classified as the remission group, and the others as the non-remission group. Patients in remission group had a relatively lower value for baseline RDW (38.82±2.76vs 39.89±2.78fL, p=0.017) compared with those in non-remission group. A graded decrease of remission rate (67.50%, 55.00%, 53.66% and 30.77% for Quartile 1 to Quartile 4 respectively, P0.05) was observed with the increasing of RDWs. The risk of hyperglycemic relapse was significantly increased for those in the highest quartile compared with the lowest (hazard ratio=2.68; 95% CI, 1.38-5.22). Those who achieved euglycemia within 7days or obtained a better fasting glucose after therapy had preferable remission rates.Patients with lower baseline RDWs are more likely to maintain a one-year euglycemia remission after short-term CSII. A faster normalization of glucose during treatment and a lower fasting glucose after therapy are correlated with a long-term glucose control.

ObjectiveTo discover diverse genotype-phenotype associations affiliated with Type 2 Diabetes Mellitus (T2DM) via genome-wide association study (GWAS) and phenome-wide association study (PheWAS), more cases (T2DM subjects) and controls (subjects without T2DM) are required to be identified (e.g., via Electronic Health Records (EHR)). However, existing expert based identification algorithms often suffer in a low recall rate and could miss a large number of valuable samples under conservative filtering standards. The goal of this work is to develop a semi-automated framework based on machine learning as a pilot study to liberalize filtering criteria to improve recall rate with a keeping of low false positive rate.Materials and MethodsWe propose a data informed framework for identifying subjects with and without T2DM from EHR via feature engineering and machine learning. We evaluate and contrast the identification performance of widely-used machine learning models within our framework, including k-Nearest-Neighbors, Naïve Bayes, Decision Tree, Random Forest, Support Vector Machine and Logistic Regression. Our framework was conducted on 300 patient samples (161 cases, 60 controls and 79 unconfirmed subjects), randomly selected from 23,281 diabetes related cohort retrieved from a regional distributed EHR repository ranging from 2012 to 2014.ResultsWe apply top-performing machine learning algorithms on the engineered features. We benchmark and contrast the accuracy, precision, AUC, sensitivity and specificity of classification models against the state-of-the-art expert algorithm for identification of T2DM subjects. Our results indicate that the framework achieved high identification performances (~0.98 in average AUC), which are much higher than the state-of-the-art algorithm (0.71 in AUC).DiscussionExpert algorithm-based identification of T2DM subjects from EHR is often hampered by the high missing rates due to their conservative selection criteria. Our framework leverages machine learning and feature engineering to loosen such selection criteria to achieve a high identification rate of cases and controls.ConclusionsOur proposed framework demonstrates a more accurate and efficient approach for identifying subjects with and without T2DM from EHR.

Vascular endothelial cells (VECs) that form the inner wall of blood vessels can be injured by high glucose-induced autophagy and apoptosis. Although the role of long noncoding RNA in regulating cell fate has received widespread attention, long noncoding RNAs (lncRNAs) that can both regulate autophagy and apoptosis need to be discovered. In this study, we identified that a small chemical molecule, 3-benzyl-5-([2-nitrophenoxy] methyl)-dihydrofuran-2(3H)-one (3BDO), synthesized by us, could inhibit VEC autophagy and apoptosis induced by a high concentration of glucose. To find new lncRNAs that regulate autophagy and apoptosis in VECs, we performed lncRNA microarray analysis. We found and verified an upregulated lncRNA named CA7-4 that was induced by a high concentration of glucose could be downregulated by 3BDO most obviously among all of the detected lncRNAs. Meanwhile, we investigated the mechanism of CA7-4 in regulating VEC autophagy and apoptosis. The results showed that CA7-4 facilitated endothelial autophagy and apoptosis as a competing endogenous RNA (ceRNA) by decoying MIR877-3P and MIR5680. Further study elucidated that MIR877-3P could trigger the decrease of CTNNBIP1 (catenin beta interacting protein 1) by combining with its 3ʹ UTR and then upregulating CTNNB1 (catenin beta 1); MIR5680 inhibited the phosphorylation of AMP-activated protein kinase (AMPK) by targeting and decreasing DPP4 (dipeptidyl peptidase 4). Therefore, CA7-4, MIR877-3P and MIR5680 represent new signal pathways that regulate VEC autophagy and apoptosis under the high-glucose condition. Abbreviations: 3BDO: 3-benzyl-5-([2-nitrophenoxy] methyl)-dihydrofuran-2(3H)-one; 3ʹ UTR: 3ʹ untranslated region; AGO2: argonaute RISC catalytic component 2; AMPK: AMP-activated protein kinase/protein kinase AMP-activated; BAX/BCL2L4: BCL2 associated X, apoptosis regulator; BCL2: BCL2 apoptosis regulator; CASP3: caspase 3; ceRNA: competing endogenous RNA; CTNNB1: catenin beta 1; CTNNBIP1/ICAT: catenin beta interacting protein 1; DPP4: dipeptidyl peptidase 4; FGF2/FGF-2: fibroblast growth factor 2; HG: high concentration glucose (30 mM glucose); lncRNA: long noncoding RNA; MAP1LC3B/LC3B: microtubule associated protein 1 light chain 3 beta; miRNA: microRNA; MIR4778-3P: microRNA 4778-3p; MIR561-3P: microRNA 561-3p; MIR5680: microRNA 5680; MIR877-3P: microRNA 877-3p; MTOR: mechanistic target of rapamycin kinase; Mut: mutant; NC: negative control; NG: normal concentration glucose (5.5 mM glucose); PARP1: poly(ADP-ribose) polymerase 1; qPCR: quantitative real-time PCR; RNA-FISH: RNA-fluorescence in situ hybridization; ROS: reactive oxygen species; RT-PCR: reverse transcription polymerase chain reaction; siRNA: small interfering RNA; SQSTM1: sequestosome 1; TGFB2-OT1: TGFB2 overlapping transcript 1; TUNEL: terminal deoxynucleotidyl transferase dUTP nick end labeling; VECs: vascular endothelial cells; WT: wild type

More and more studies reported that diverse biological roles of long noncoding RNAs were usually dependent on their subcellular location. In our previous study, long noncoding RNA CERNA1 was identified both located in cytoplasm and nucleus of vascular endothelial cells (VECs). And CERNA1 in cytoplasm, which functioned as competitive endogenous RNA (ceRNA), alleviated the apoptosis of VECs. However, the function of CERNA1 in nucleus was still unclear. In this study, we found that nuclear CERNA1 positively regulated BCL2L10, which accelerated the serum and FGF-2 starvation-induced apoptosis of VECs, by enhancing the histone modification level of H3K9ac and H3K4me3 in BCL2L10 promoter region. Furthermore, due to the paradoxical function, we investigated the variation of CERNA1 subcellular location in VECs. The results showed that, as the change of apoptosis status, CERNA1 altered the cellular distribution in VECs. And the annexin A7 inhibitor, ABO (6-amino-2,3-dihydro-3-hydroxymethyl-1,4-benzoxazine), not only increased the expression of CERNA1 by TIA-1, but also specifically improved its cytoplasm distribution proportion so as to inhibit the apoptosis of VECs. This evidence suggested that the subcellular location of CERNA1 played an important role in the VECs apoptosis and ABO might be a potential chemical molecule for therapy of VECs apoptosis related cardiovascular diseases.

Apoptosis of vascular endothelial cells (VEC) is the main form of vascular injury that is closely linked to numerous cardiovascular diseases. Therefore, it is important to find new factors that can suppress VEC apoptosis. By using long noncoding RNA (lncRNA) microarray analysis, we found a new read-through lncRNA, MROH7-TTC4, which acted as an apoptosis inhibitor in VECs. Furthermore, by using the inhibitor (ABO) of annexin A7 (ANXA7) GTPase, we discovered that ANXA7 translocated into nucleus and interacted with 5'→3' exoribonuclease (XRN2). The decreased XRN2 phosphorylation induced by ANXA7 GTPase activity inhibition, promoted MROH7-TTC4 expression. Moreover, T-cell intracellular antigen-1 (TIA1), a binding protein of MROH7-TTC4, processed it into MROH7 and TTC4 that could inhibit VEC apoptosis. Here, we conclude that inhibiting ANXA7 GTPase activity promotes the interaction of ANXA7 and XRN2 in nucleus, which regulates the read-through transcription of MROH7-TTC4, and TIA1 is responsible for the process of MROH7-TTC4 that inhibits apoptosis through MROH7 and TTC4.

Atherosclerosis is predicted to be the primary cause of death in the world by 2020. Changes in atherosclerotic plaque composition will lead to acute coronary syndromes. Although the studies on the molecular mechanisms of long noncoding RNA (lncRNA) are in-depth in molecular and cell levels, the in vivo research which studied the knowledge about lncRNAs in the regulation of plaque composition is still sparse. In this study, in order to investigate how a new lncRNA, CERNA1, regulates the composition of atherosclerotic plaques, we overexpressed CERNA1 in apolipoprotein E-/- (Apo E-/-) mice and analyzed the role of CERNA1 in atherosclerotic plaque stabilization. The results showed that CERNA1 inhibited the apoptosis of VSMCs and anti-inflammatory macrophages through increasing API5 level and further stabilized the atherosclerotic plaques. This discovery provided a novel therapeutic target for atherosclerosis.

Overexpression of stathmin (STMN1) is closely linked to tumor metastases and poor prognosis in endometrial carcinoma (EC). However, the underlying mechanism is little known. In the present study, we investigated the expression of STMN1 in EC. Subsequently, we assessed the role of STMN1 in EC cell proliferation and migration. Our data show that STMN1 is upregulated in EC, and elevated expression of STMN1 is correlated positively with tumor stage and lymph node metastasis. In vitro, forced expression of STMN1 promoted cell invasion and migration. In contrast, knockdown of STMN1 inhibited cell aggressive behaviors. Moreover, the expression and the activity changes of matrix metalloproteinases (MMP)-2/9 were observed in EC cells after the cells being silenced or overexpression of STMN1. In conclusion, STMN1 is an oncogene and it enhances the growth and invasion of EC possibly by mediating the secretion and activation of MMP2 and MMP9 protein.

Sclerostin domain containing protein 1 (SOSTDC1) is down-regulated and acts as a tumor suppressor in some kinds of cancers. However, the expression pattern and biological significance of SOSTDC1 in thyroid cancer are largely unknown. We demonstrated that SOSTDC1 was significantly down-regulated in thyroid cancer. Ectopic over-expression of SOSTDC1 inhibited proliferation and induced G1/S arrest in thyroid cancer cells. Moreover, SOSTDC1 over-expression suppressed the growth of tumor xenografts in nude mice. We also found that elevated SOSTDC1 led to inhibition of cyclin A2 and cyclin E2. Together, our results demonstrate that SOSTDC1 is down-regulated in thyroid cancer and might be a potential therapeutic target in the treatment of thyroid cancer.

NISCH encodes the imidazoline receptor Nischarin and is a known tumor suppressor in many human malignancies; however, its roles in ovarian cancer are still largely unknown. Here, we aim to investigate the biologic functions of NISCH in ovarian cancer. We found that NISCH was significantly downregulated, which correlated considerably with advanced tumor stage, poor differentiation, lymph node metastasis, and the serous/mucinous subtypes in a panel of ovarian cancer tissues. Moreover, NISCH gene silencing was mainly the product of promoter hypermethylation, which could be reversed by treatment with 5-aza-dC. In vitro, NISCH overexpression suppressed cell proliferation and colony formation by hindering cell-cycle progression, whereas the opposite was observed in NISCH knockdown counterparts. In vivo, abundant NISCH expression hindered the growth of HO8910 xenografts, whereas NISCH knockdown accelerated the growth of SKOV3 xenografts. In addition, NISCH significantly attenuated cell invasion by inhibiting the phosphorylation of FAK and ERK, which could be neutralized by PF-562271 (a FAK/Pyk2 inhibitor). Accordingly, NISCH knockdown xenografts exhibited increased peritoneal/pelvic metastases that were not present in counterparts treated with PF-562271. Furthermore, NISCH expression in primary ovarian cancer cells predicted a cellular resistance to PF-562271. In conclusion, we showed that NISCH was frequently silenced by promoter hypermethylation in human ovarian cancer. NISCH manipulated cellular proliferation and invasion by arresting cell cycle and inhibiting the FAK signal. Our findings revealed the biologic functions of NISCH in ovarian cancer, and might be useful for treating patients with aberrant expression of NISCH. Mol Cancer Ther; 14(5); 1202–12. ©2015 AACR.

Introduction: Neurosarcoidosis accounts for approximately 5% of the sarcoidosis, which develops exclusively in the nervous system and is always difficult to diagnose. We describe a rare case of isolated neurosarcoidosis mimicking as multifocal meningiomas. A 27-year-old male was admitted to our hospital with a history of unconsciousness and convulsion 1 month ago, which was suspected as a seizure. The results showed no abnormalities in complete blood count; serum electrolytes; erythrocyte sedimentation rate and ultrasonography of the liver, pancreas, spleen, kidney and parotid gland, and so on. Chest radiograph and electroencephalogram were also normal. Serum-angiotensin-converting enzyme slightly increased. Normal opening pressure was shown in cerebrospinal fluid sampling, which includes 8/μL white blood cells, 0.93 g/L protein, and 3.03 mmol/L glucose. Enhanced magnetic resonance imaging revealed multifocal enhancement lesions, including left sphenoid wing region, left temporal and bilateral occipitoparietal region, which were suspected as multiple “meningioma”. A left frontotemporal craniotomy was further performed. Both necrotizing and non-necrotizing granulomas were revealed in the pathological specimen, most of which were associated with multinucleated giant cells and macrophages. We could also see the fibrosis and inflammatory reaction in the sample composed of lymphocytes, histiocytes, and plasma cells. Histopathological examination showed that the cells were positive for human CD68 (KP1), CD68 (PGM1), and CD163; however, they were negative for the AF, epithelial membrane antigen, and glial fibrillary acidic protein. Tuberculosis-deoxyribonucleic acid test and special stains for acid-fast bacilli and fungi were negative. The diagnosis was finally made as isolated neurosarcoidosis. Then the patient was treated with additional corticosteroid therapy. Serial imaging examination 4 months later revealed that the lesions extremely decreased. Conclusion: The diagnosis of isolated central nervous system sarcoidosis was still difficult because of limitations of available diagnostic tests. So neurosarcoidosis in the clinical work should never be neglected.

Long non-coding RNAs (lncRNAs) are emerging as key molecules in human cancer. Homeobox (HOX) transcript antisense intergenic RNA (HOTAIR), a long non-coding RNA (lncRNA), is associated with a variety of human cancers, such as breast, liver and lung cancer. However, whether HOTAIR can function as a molecular marker in endometrial carcinoma (EC) remains unknown. In the present study, the expression of HOTAIR in 66 EC tissues from patients with EC and 30 normal tissues from healthy age-matched control subjects was determined using quantitative reverse transcription PCR. Furthermore, using in situ hybridization, we measured HOTAIR expression in 129 formalin-fixed paraffin-embedded (FFPE) tissue sections, which included 96 tissues that matched the frozen cases, 21 other EC tissues and 12 atypical hyperplasia tissues. Correlations between HOTAIR expression and the clinicopathological characteristics of patients were analyzed. Our results revealed that HOTAIR expression in the EC tissues was significantly upregulated compared with normal tissues (p

Purpose. This study aimed to explore the relationship among insulin sensitivity and ectopic fat depots in participants with different glucose status.Methods. Fifty-nine men and women were enrolled in this study: 29 with normal glucose tolerance (NGT), 17 with impaired glucose tolerance (IGT), and 13 with type 2 diabetes mellitus (T2DM). All participants underwent a hyperinsulinemic-euglycemic clamp to assess the insulin sensitivity index (Mvalue) and magnetic resonance imaging to measure the hepatocellular lipid content (HCL), skeletal muscle fat content including intramyocellular lipid (IMCL) and extramyocellular lipid (EMCL) of tibialis anterior (ta), and soleus muscle (sol).Results. TheMvalue of NGT group was higher than those of IGT and T2DM groups (P=0.001). Participants with T2DM had the highest HCL and IMCL (ta) compared with those in NGT and IGT groups (P=0.001). TheMvalue had an inverse relationship with HCL (r=-0.789,P=0.001), IMCL (sol) (r=-0.427,P=0.002), and IMCL (ta) (r=-0.419,P=0.002). Stepwise linear regression analysis showed that HCL (standardizedβ=-0.416;P=0.001) had an independent relationship withMvalue.Conclusions. Hepatocellular lipid content deposition happens earlier than skeletal muscle fat deposition. HCL is an independent risk factor for insulin resistance and may be used to evaluate the risk of developing T2DM as a noninvasive marker of insulin sensitivity index.

Purpose: Bardoxolone methyl, a novel synthetic triterpenoid and antioxidant inflammation modulator, potently induces Nrf2 and inhibits NF-κB and Janus-activated kinase/STAT signaling. This first-in-human phase I clinical trial aimed to determine the dose-limiting toxicities (DLT), maximum tolerated dose (MTD), and appropriate dose for phase II studies; characterize pharmacokinetic and pharmacodynamic parameters; and assess antitumor activity. Experimental Design: Bardoxolone methyl was administered orally once daily for 21 days of a 28-day cycle. An accelerated titration design was employed until a grade 2–related adverse event occurred. A standard 3 + 3 dose escalation was then employed until the MTD was reached. Single dose and steady-state plasma pharmacokinetics of the drug were characterized. Assessment of Nrf2 activation was examined in peripheral blood mononuclear cells (PBMC) by measuring NAD(P)H:quinone oxidoreductase (NQO1) mRNA levels. Immunohistochemical assessment of markers of inflammation, cell cycle, and apoptosis was carried out on tumor biopsies. Results: The DLTs were grade 3 reversible liver transaminase elevations. The MTD was established as 900 mg/d. A complete tumor response occurred in a mantle cell lymphoma patient, and a partial response was observed in an anaplastic thyroid carcinoma patient. NQO1 mRNA levels increased in PBMCs, and NF-κB and cyclin D1 levels decreased in tumor biopsies. Estimated glomerular filtration rate (eGFR) was also increased. Conclusions: Bardoxolone methyl was well tolerated with an MTD of 900 mg/d. The increase in eGFR suggests that bardoxolone methyl might be beneficial in chronic kidney disease. Objective tumor responses and pharmacodynamic effects were observed, supporting continued development of other synthetic triterpenoids in cancer. Clin Cancer Res; 18(12); 3396–406. ©2012 AACR.

The pharmacologic utility of lengthy peptides can be hindered by loss of bioactive structure and rapid proteolysis, which limits bioavailability. For example, enfuvirtide (Fuzeon, T20, DP178), a 36-amino acid peptide that inhibits human immunodeficiency virus type 1 (HIV-1) infection by effectively targeting the viral fusion apparatus, has been relegated to a salvage treatment option mostly due to poor in vivo stability and lack of oral bioavailability. To overcome the proteolytic shortcomings of long peptides as therapeutics, we examined the biophysical, biological, and pharmacologic impact of inserting all-hydrocarbon staples into an HIV-1 fusion inhibitor. We find that peptide double-stapling confers striking protease resistance that translates into markedly improved pharmacokinetic properties, including oral absorption. We determined that the hydrocarbon staples create a proteolytic shield by combining reinforcement of overall α-helical structure, which slows the kinetics of proteolysis, with complete blockade of peptide cleavage at constrained sites in the immediate vicinity of the staple. Importantly, double-stapling also optimizes the antiviral activity of HIV-1 fusion peptides and the antiproteolytic feature extends to other therapeutic peptide templates, such as the diabetes drug exenatide (Byetta). Thus, hydrocarbon double-stapling may unlock the therapeutic potential of natural bioactive polypeptides by transforming them into structurally fortified agents with enhanced bioavailability.

Ixabepilone is an epothilone, a novel class of non-taxane microtubule stabilizing agents. A phase I/II and pharmacokinetic trial of ixabepilone was conducted in patients with recurrent high-grade gliomas. Adult patients received ixabepilone as a 1-h infusion daily for 5 days every 3 weeks. A modified continual reassessment method was used to escalate doses, beginning at 5.0 mg/m(2), in patients stratified by use or non-use of enzyme inducing antiepileptic drugs (EIAED). In the phase I study, the maximum tolerated dose (MTD) and pharmacokinetics of ixabepilone were determined for each group. The phase II study used a two-stage design to evaluate response rate. Secondary endpoints were survival and 6-month progression free survival. In the phase I trial, 38 patients (median age 54 years) were enrolled. The MTD was 6.8 mg/m(2) for patients not taking EIAEDs and 9.6 mg/m(2) for those taking EIAEDs. The dose limiting toxicities in both groups were hematologic. Twenty-three patients (median age 54 years) were enrolled in the first stage of the phase II trial. No objective responses were observed. Median overall survival was 5.8 (95% CI, 5.0-8.6) months and 6-month PFS rate was 4% (95% CI, 0-22%). The overall mean total body clearance for ixabepilone was significantly higher (P = 0.003) in patients receiving EIAEDs (36 ± 11 l/h/m(2)) than those not (24 ± 9.2 l/h/m(2)). Patients on EIAEDs had a substantially higher MTD likely due to induction of cytochrome P450. Ixabepilone had no activity in patients with recurrent high-grade gliomas.

Purpose: Determining whether potentially therapeutic drug exposure is achieved within brain tumors in an exploratory clinical investigation would provide a rational basis for selecting agents for evaluation in phase II trials. This study investigated the use of microdialysis to assess intratumoral drug distribution in patients with recurrent high grade gliomas (HGG). Patients and Methods: Microdialysis catheters were placed during surgery for residual HGG 1-day before giving methotrexate (MTX) 12-g/m2 by 4-h i.v. infusion. MTX was measured by Liquid Chromatography/Mass Spectrometry (LC/MS) in plasma and microdialysate during the infusion and for 24-h thereafter. Blood brain barrier (BBB) permeability of tissue in which the microdialysis probe was located was determined by digitally fusing brain CT and contrast enhanced MRI images. Results: The microdialysis probe was located in contrast enhancing tumor in two patients and nonenhancing tissue in two others. Cerebral drug penetration, as indicated by the ratio of the area under the MTX concentration–time curves in brain extracellular fluid and plasma, was considerably greater in contrast enhancing tumor (0.28–0.31) than nonenhancing tissue (0.032–0.094). Nevertheless, MTX concentrations in ECF exceeded 2-μM, the average concentration for 50% cell kill against glioma cell lines in vitro, for 20–26 h in both regions of the tumor. Conclusions: Microdialysis is a very informative technique for characterizing the intratumoral pharmacokinetics of drugs, such as MTX, that do not freely penetrate the BBB. Establishing the catheter probe location relative to areas of BBB disruption is required to properly assess the significance of microdialysis data in this context.

3,4-Dihydro-2,2-dimethyl-2H-naphthol[1,2-b]pyran-5,6-dione (ARQ 501; beta-lapachone) showed promising anticancer activity in phase I clinical trials as monotherapy and in combination with cytotoxic drugs. ARQ 501 is currently in multiple phase II clinical trials. In vitro incubation in fresh whole blood at 37 degrees C revealed that ARQ 501 is stable in plasma but disappears rapidly in whole blood. Our data showed that extensive metabolism in red blood cells (RBCs) was mainly responsible for the rapid disappearance of ARQ 501 in whole blood. By comparison, covalent binding of ARQ 501 and/or its metabolites to whole blood components was a minor contributor to the disappearance of this compound. Sequestration of intact ARQ 501 in RBCs was not observed. Cross-species metabolite profiles from incubating [(14)C]ARQ 501 in freshly drawn blood were characterized using a liquid chromatography-mass spec-trometry-accurate radioactivity counter. The results show that ARQ 501 was metabolized more rapidly in mouse and rat blood than in dog, monkey, and human blood, with qualitatively similar metabolite profiles. Six metabolites were identified in human blood using ultra-high performance liquid chromatography/time-of-flight mass spectrometry, and the postulated structure of five metabolites was confirmed using synthetic standards. We conclude that the primary metabolic pathway of ARQ 501 in human blood involved oxidation of the two adjacent carbonyl groups to produce dicarboxylic and monocarboxylic metabolites, elimination of a carbonyl group to form a ring-contracted metabolite, and lactonization to produce two metabolites with a pyrone ring to form a ring-contracted metabolite. Metabolism by RBCs may play a role in clearance of ARQ 501 from the blood compartment in cancer patients.

Background. Women with ovarian cancer who experience disease progression during or within 6 months of first-line treatment with platinum-based anticancer drugs are considered to have platinum-resistant tumors. These patients have an unfavorable prognosis, and they frequently seek complementary and alternative therapies (CAM). Historically, this represents an understudied and underreported component of ovarian cancer treatment. Case. This report describes the case of a woman with rapidly progressive, platinum-resistant ovarian cancer. Upon initiating self-directed treatment with Haelan951®, a commercially available fermented soy beverage, she entered into a phase of prolonged disease stabilization including improvement in the serum tumor marker CA-125. Conclusion. Fermented soy products are known to contain high concentrations of the isoflavone, genistein, and other compounds that exhibit anticancer activity in preclinical models. This case report supports the prospective evaluation of alternative therapies such as these in patients with platinum-refractory ovarian cancer.