Stimuli directed towards activation of apoptosis mechanisms are an attractive approach to eliminate evasion of apoptosis, a ubiquitous cancer hallmark. In these in vitro studies, kinetics and electric field thresholds for several apoptosis characteristics are defined in E4 squamous carcinoma cells (SCC) exposed to ten 300 ns pulses with increasing electric fields. Cell death was >95% at the highest electric field and coincident with phosphatidylserine externalization, caspase and calpain activation in the presence and absence of cytochrome c release, decreases in Bid and mitochondria membrane potential (Δψm) without apparent changes reactive oxygen species levels or in Bcl2 and Bclxl levels. Bid cleavage was caspase-dependent (55-60%) and calcium-dependent (40-45%). Intracellular calcium as an intrinsic mechanism and extracellular calcium as an extrinsic mechanism were responsible for about 30 and 70% of calcium dependence for Bid cleavage, respectively. The results reveal electric field-mediated cell death induction and progression, activating pro-apoptotic-like mechanisms and affecting plasma membrane and intracellular functions, primarily through extrinsic-like pathways with smaller contributions from intrinsic-like pathways. Nanosecond second pulsed electric fields trigger heterogeneous cell death mechanisms in E4 SCC populations to delete them, with caspase-associated cell death as a predominant, but not an unaccompanied event.

Purpose. To study the characteristics and prognostic significance of preoperative magnetic resonance imaging- (MRI-) assessed circumferential margin (CRM) in rectal cancer. Methods. Patients underwent preoperative high resolution pelvic MRI, followed by resection of primary tumor. The relationship between MRI-assessed CRM and pathological CRM (pCRM) was studied, and survival analysis was used to determine the prognostic significance of MRI-assessed CRM. Results. Of all the 203 patients, the total accuracy of MRI-assessed CRM for predicting involvement of pCRM was 84.2%, sensitivity was 50%, and specificity was 86.8%. Anterior tumors were more possible to assess involvement of CRM by MRI, while the false positive rate was significantly higher than lateral or posterior tumor (87.5% versus 50%, p = 0.0002). The 3-year local recurrence, disease-free survival, and overall survival rates were 35.6%, 58.1%, and 85.2% in patients with involved mrCRM, compared with 8.9%, 78.9%, and 92.3% in patients with clear mrCRM. In multivariate analysis, MRI-assessed CRM found an independent risk factor for local recurrence, with a hazard ratio of 3.49 (p = 0.003). Conclusions. High resolution MRI was accurate to assess CRM preoperatively, while anterior tumor should be assessed more cautiously. Involvement of mrCRM was significantly associated with local recurrence regardless of pCRM status.

The successful development of safe and highly effective nanoprobes for targeted imaging and simultaneous therapy of in vivo gastric cancer is a great challenge. Herein we reported for the first time that anti-α-subunit of ATP synthase antibody, HAI-178 monoclonal antibody-conjugated fluorescent magnetic nanoparticles, was successfully used for targeted imaging and simultaneous therapy of in vivo gastric cancer. A total of 172 specimens of gastric cancer tissues were collected, and the expression of α-subunit of ATP synthase in gastric cancer tissues was investigated by immunohistochemistry method. Fluorescent magnetic nanoparticles were prepared and conjugated with HAI-178 monoclonal antibody, and the resultant HAI-178 antibody-conjugated fluorescent magnetic nanoparticles (HAI-178-FMNPs) were co-incubated with gastric cancer MGC803 cells and gastric mucous GES-1 cells. Gastric cancer-bearing nude mice models were established, were injected with prepared HAI-178-FMNPs via tail vein, and were imaged by magnetic resonance imaging and small animal fluorescent imaging system. The results showed that the α-subunit of ATP synthase exhibited high expression in 94.7% of the gastric cancer tissues. The prepared HAI-178-FMNPs could target actively MGC803 cells, realized fluorescent imaging and magnetic resonance imaging of in vivo gastric cancer, and actively inhibited growth of gastric cancer cells. In conclusion, HAI-178 antibody-conjugated fluorescent magnetic nanoparticles have a great potential in applications such as targeted imaging and simultaneous therapy of in vivo early gastric cancer cells in the near future.

In the past few years, therapies targeted at the von Hippel-Lindau (VHL) and hypoxia-inducible factor (HIF) pathways, such as sunitinib and sorafenib, have been developed to treat clear cell renal cell carcinoma (ccRCC). However, the majority of patients will eventually show resistance to antiangiogenesis therapies. The purpose of our study was to identify novel pathways that could be potentially used as targets for new therapies. Whole transcriptome sequencing (RNA-Seq) was conducted on eight matched tumor and adjacent normal tissue samples. A novel RUNX1-RUNX1T1 pathway was identified which was upregulated in ccRCC through gene set enrichment analysis (GSEA). We also confirmed the findings based on previously published gene expression microarray data. Our data shows that upregulated of the RUNX1-RUNX1T1 gene set maybe an important factor contributing to the etiology of ccRCC.

The main objective of this study was to demonstrate the proof-of-principle for the hypothesis that conjugated linoleic acid-paclitaxel conjugate (CLA-PTX), a novel fatty acid modified anti-cancer drug conjugate, could self-assemble forming nanoparticles. The results indicated that a novel self-assembling nanomedicine, CLA-PTX@PEG NPs (about 105 nm), with Cremophor EL (CrEL)-free and organic solvent-free characteristics, was prepared by a simple precipitation method. Being the ratio of CLA-PTX:DSPE-PEG was only 1:0.1 (w/w), the higher drug loading CLA-PTX@PEG NPs (about 90%) possessed carrier-free characteristic. The stability results indicated that CLA-PTX@PEG NPs could be stored for at least 9 months. The safety of CLA-PTX@PEG NPs was demonstrated by the MTD results. The anti-tumor activity and cellular uptake were also confirmed in the in vitro experiments. The lower crystallinity, polarity and solubility of CLA-PTX compared with that of paclitaxel (PTX) might be the possible reason for CLA-PTX self-assembling forming nanoparticles, indicating a relationship between PTX modification and nanoparticles self-assembly. Overall, the data presented here confirm that this drug self-delivery strategy based on self-assembly of a CLA-PTX conjugate may offer a new way to prepare nanomedicine products for cancer therapy involving the relationship between anticancer drug modification and self-assembly into nanoparticles.

In this study, new clindamycin (CLIN) artificial antigens were prepared and used to produce broad-specificity monoclonal antibodies. Based on the as-produced mAbs, a heterologous ELISA was developed to detect CLIN and lincomycin (LIN) residues in edible animal tissues. The IC50 values of the developed assay were 0.3ng/mL (CLIN) and 1.2ng/mL (LIN) in buffer, respectively. The detection limits were estimated to be 1.8μg/kg (CLIN) and 6.8μg/kg (LIN) in bovine, chicken, porcine and fish muscles. In the spike and recovery tests, the mean recovery rate ranged from 76% to 112% at different spiked levels, and the intra-/inter-assay coefficients of variation were in the range of 7.1% to 13.2%. This method was verified using LC-MS/MS with a correlation coefficient >0.97. The developed ELISA is therefore well suited for simultaneous determination of CLIN and LIN residues in bovine, chicken, porcine and fish muscles.

Cadherin is crucial for cell-cell adhesion and N-glycosylation of N-cadherin has been implicated in the process of mammary, renal, and ovarian carcinogenesis. However, whether N-glycosylation of N-cadherin plays a role in glioma remains unknown. Previous studies had indicated that N-glycosylation could occur at three asparagine residues of N-cadherin. By generating and over-expressing N-glycosylation-deficient N-cadherin mutants in the human glioma cell lines SHG66 and U87, we found that mutation of N402 but not of the other potentially N-glycosylated residues destabilized N-cadherin and led to its ubiquitylation and subsequent proteasomal degradation. Furthermore, destabilized N-cadherin inhibited cadherin-mediated cell-cell adhesion and promoted cell migration. Our findings reveal that N-glycosylation controls N-cadherin stability and plays a role in glioma migration. J. Cell. Biochem. 118: 1423-1431, 2017. © 2016 Wiley Periodicals, Inc.

Gait phase is widely used for gait trajectory generation, gait control and gait evaluation on lower-limb exoskeletons. So far, a variety of methods have been developed to identify the gait phase for lower-limb exoskeletons. Angular sensors on lower-limb exoskeletons are essential for joint closed-loop controlling; however, other types of sensors, such as plantar pressure, attitude or inertial measurement unit, are not indispensable.Therefore, to make full use of existing sensors, we propose a novel gait phase recognition method for lower-limb exoskeletons using only joint angular sensors. The method consists of two procedures. Firstly, the gait deviation distances during walking are calculated and classified by Fisher's linear discriminant method, and one gait cycle is divided into eight gait phases. The validity of the classification results is also verified based on large gait samples. Secondly, we build a gait phase recognition model based on multilayer perceptron and train it with the phase-labeled gait data. The experimental result of cross-validation shows that the model has a 94.45% average correct rate of set (CRS) and an 87.22% average correct rate of phase (CRP) on the testing set, and it can predict the gait phase accurately. The novel method avoids installing additional sensors on the exoskeleton or human body and simplifies the sensory system of the lower-limb exoskeleton.

GADD45B is a member of the growth arrest DNA damage-inducible gene family associated with cell growth control, apoptosis, and DNA damage repair response. The aim of this study is to detect the role of GADD45B in colorectal carcinoma (CRC); the area not studied in depth to date.

Medium spiny neurons (MSNs) in the nucleus accumbens (NAc) undergo persistent alterations in their biological and physiological characteristics upon exposure to drugs of abuse. Previous studies demonstrated that the biochemical, morphological, and intrinsic physiological properties of MSNs are heterogeneous and provided new insights into the physiological and molecular roles of individual MSNs in addictive behaviors. However, it remains unclear whether MSNs in the NAc shell (NAcSh), an important region for mediating behavioral sensitization, are electrophysiologically heterogeneous and how such heterogeneity is relevant to neuroadaptation associated with drug addiction. Here, the membrane properties, i.e., the intrinsic excitability and spike adaptation, of MSNs in the NAcSh from saline- or morphine-treated rats were investigated in vitro by whole-cell recording. In saline-treated rats, three distinct cell types were identified by their membrane properties: type I neurons showed high levels of intrinsic excitability and rapid spike adaptation; type II neurons showed moderate levels of intrinsic excitability and relatively slow spike frequency adaptation; type III neurons showed low levels of intrinsic excitability and putative strong spike adaptation. MSNs in rats undergoing withdrawal from chronic morphine treatment (10-14 days after the last injection) also exhibited the typical firing behaviors of these three types of neurons. However, the membrane properties of the MSNs were differentially altered after withdrawal. There was an enhancement in intrinsic excitability in type II MSNs and a promotion of spike adaptation in type I MSNs. The apamin-sensitive afterhyperpolarization current (I(AHP)) and the apamin-insensitive I(AHP) of the NAcSh MSNs were attenuated after chronic morphine withdrawal. These findings suggest that individual MSNs in the NAcSh manifest unique electrophysiological properties, which might contribute to psychostimulant-induced neuroadaptation.

Colorectal cancer is the leading cause of cancer-related deaths worldwide. The disease is curable when detected at an early stage. However, the compliance rate with current screening recommendations remains poor. An accurate, minimally invasive blood test that has the potential for greater patient compliance would be a welcome addition to the current methods. Recent data have shown that gene expression profile of peripheral blood cells can reflect disease states and thus have diagnostic value. In this study, genome-wide gene expression profiling of peripheral blood cells from 20 healthy controls and 20 colorectal cancer patients were performed using PAXgene™ technology and Affymetrix GeneChip® microarrays. We identified a list of 1,469 genes that were differentially expressed between the healthy controls and cancer patients. Gene annotation and functional enrichment analysis revealed that those genes are mainly related to immune functions. Particularly, a set of genes belonging to the Toll-Like Receptor pathways were up-regulated in the colorectal cancer patients. These findings provide a new understanding of blood gene expression profile in colorectal cancer. Our result may serve as the basis for further development of blood biomarkers for the diagnosis and treatment of colorectal cancer.

Although the allocation of brain functions across the two cerebral hemispheres has aroused public interest over the past century, asymmetric interhemispheric cooperation under attentional modulation has been scarcely investigated. An example of interhemispheric cooperation is visual spatial perception. During this process, visual information from each hemisphere is integrated because each half of the visual field predominantly projects to the contralateral visual cortex. Both egocentric and allocentric coordinates can be employed for visual spatial representation, but they activate different areas in primate cerebral hemispheres. Recent studies have determined that egocentric representation affects the reaction time of allocentric perception; furthermore, this influence is asymmetric between the two visual hemifields. The egocentric-allocentric incompatibility effect and its asymmetry between the two hemispheres can produce this phenomenon. Using an allocentric position judgment task, we found that this incompatibility effect was reduced, and its asymmetry was eliminated on an attentional task rather than a neutral task. Visual attention might activate cortical areas that process conflicting information, such as the anterior cingulate cortex, and balance the asymmetry between the two hemispheres. Attention may enhance and balance this interhemispheric cooperation because this imbalance may also be caused by the asymmetric cooperation of each hemisphere in spatial perception.

Liver metastasis is common in patients diagnosed with colorectal cancer (CRC), and is also correlated with poor outcome. In this study we screened the different expression profiles of microRNAs (miRNAs) on the development of liver metastasis in CRC patients. miR-99b-5p was found to be more than 6-fold higher in primary tumors than in matched liver metastases (P = 0.007). Expression of miR-99b-5p in primary tumors of patients with stage III CRC without liver metastases was higher than in CRC patients with liver metastases (P = 0.028). Up-regulated miR-99b-5p was associated with longer overall survival (P = 0.01). Besides, miR-99b-5p silencing in miR-99b-5p-positive CRC cell lines promoted cell migration and up-regulated mTOR, and vice versa. In addition, luciferase assays demonstrated that miR-99b-5p functioned as a tumor suppressor by targeting mTOR. Taken together, our results demonstrate thatmiR-99b-5p is differently expressed in primary CRC and liver metastasis and functions as a tumor-suppressive microRNA in metastatic CRC. The miR-99b-5p-mTOR axis may serve as a prognostic factor and therapeutic target for anti-metastatic therapy in CRC patients.

Like the anti-angiogenic strategy, anti-vascular mimicry is considered as a novel targeting strategy for glioma. In the present study, we used NGR as a targeting ligand and prepared NGR-modified liposomes containing combretastatin A4 (NGR-SSL-CA4) in order to evaluate their potential targeting of glioma tumor cells and vasculogenic mimicry (VM) formed by glioma cells as well as their anti-VM activity in mice with glioma tumor cells. NGR-SSL-CA4 was prepared by a thin-film hydration method. The in vitro targeting of U87-MG (human glioma tumor cells) by NGR-modified liposomes was evaluated. The in vivo targeting activity of NGR-modified liposomes was tested in U87-MG orthotopic tumor-bearing nude mice. The anti-VM activity of NGR-SSL-CA4 was also investigated in vitro and in vivo. The targeting activity of the NGR-modified liposomes was demonstrated by in vitro flow cytometry and in vivo biodistribution. The in vitro anti-VM activity of NGR-SSL-CA4 was indicated in a series of cell migration and VM channel experiments. NGR-SSL-CA4 produced very marked anti-tumor and anti-VM activity in U87-MG orthotopic tumor-bearing mice in vivo. Overall, the NGR-SSL-CA4 has great potential in the multi-targeting therapy of glioma involving U87-MG cells, and the VM formed by U87-MG cells as well as endothelial cells producing anti-U87-MG cells, and anti-VM formed by U87-MG cells as well as anti-endothelial cell activity.

The mangiferin-berberine (MB) salt was synthesized by ionic bonding of mangiferin (M) and berberine (B) at an equal molecular ratio. This study aimed to investigate the activities of MB salt in modulating lipid and glucose metabolisms in HepG2 cells. After 24 h treatment of the studying compounds, cellular AMP-activated protein kinase α (AMPKα)/acetyl-CoA carboxylase (ACC) protein levels and carnitine palmitoyltransferase (CPT) 1 activities, intracellular lipid contents, mRNA expression levels of target genes, glucose consumption, and glucose production amounts were determined. Compound C (CC) was used in the blocking experiments. Our results showed that MB salt increased p-AMPKα (Thr172)/p-ACC (Ser79) levels and CPT1 activity and suppressed oleic acid- (OA-) induced lipid accumulation and upregulation of lipogenic genes potently in HepG2 cells. The above activities of MB salt were AMPK dependent and were superior to those of M or B when administered at an equal molar concentration. MB salt enhanced basal and insulin-stimulated glucose consumption and suppressed gluconeogenesis more potently than M or B alone. The inhibiting activity of MB salt on cellular gluconeogenesis was AMPK dependent. Our results may support MB salt as a new kind of agent for the development of novel lipid or glucose-lowering drugs in the future.

ADP-ribosylation is a covalent posttranslational modification of proteins that is catalyzed by various types of ADP-ribosyltransferase (ART) enzymes, including members of the poly(ADP-ribose) polymerase (PARP) family. ADP-ribose (ADPR) modifications can occur as mono(ADP-ribosyl)ation, oligo(ADP-ribosyl)ation, or poly(ADP-ribosyl)ation, depending on the particular ART enzyme catalyzing the reaction, as well as the specific reaction conditions. Understanding the biology of ADP-ribosylation requires facile and robust means of generating and detecting the modification in all of its forms. Here we describe how to generate protein-linked mono(ADP-ribose), oligo(ADP-ribose), and poly(ADP-ribose) (MAR, OAR, and PAR, respectively) in vitro as an automodification of PARPs 1 or 3. First, epitope-tagged PARP-1 (a PARP polyenzyme) and PARP-3 (a PARP monoenzyme) are expressed individually in insect cells using baculovirus expression vectors, and purified using immunoaffinity chromatography. Second, the purified recombinant PARPs are incubated individually in the presence of different concentrations of NAD+ (as a donor of ADPR groups) and sheared DNA (to activate their catalytic activities) resulting in various forms of auto-ADP-ribosylation. Third, the products are confirmed using ADPR detection reagents that can distinguish among MAR, OAR, and PAR. Finally, if desired, the OAR and PAR can be deproteinized. The protein-linked and free MAR, OAR, and PAR generated in these reactions can be used as standards, substrates, or binding partners in a variety of ADPR-related assays.

The recombinant carbonyl reductase from Rhodococcus erythropolis WZ010 (ReCR) demonstrated strict (S)-stereoselectivity and catalyzed the irreversible reduction of N-Boc-3-piperidone (NBPO) to (S)-N-Boc-3-hydroxypiperidine [(S)-NBHP], a key chiral intermediate in the synthesis of ibrutinib. The NAD(H)-specific enzyme was active within broad ranges of pH and temperature and had remarkable activity in the presence of higher concentration of organic solvents. The amino acid residue at position 54 was critical for the activity and the substitution of Tyr54 to Phe significantly enhanced the catalytic efficiency of ReCR. The kcat/Km values of ReCR Y54F for NBPO, (R/S)-2-octanol, and 2-propanol were 49.17 s-1 mM-1, 56.56 s-1 mM-1, and 20.69 s-1 mM-1, respectively. In addition, the (S)-NBHP yield was as high as 95.92% when whole cells of E. coli overexpressing ReCR variant Y54F catalyzed the asymmetric reduction of 1.5 M NBPO for 12 h in the aqueous/(R/S)-2-octanol biphasic system, demonstrating the great potential of ReCR variant Y54F for practical applications.

Prostate cancer is still considered a significant health care challenge worldwide due in part to the distinct transformation of androgen-dependent prostate cancer (ADPC) into treatment-refractory castration-resistant prostate cancer (CRPC). Consequently, there is an urgent need to explore novel molecular mechanisms underlying treatment resistance in ADPC. Although numerous studies have alluded to the role of miR-200a in several cancers, the biological significance of miR-200a in prostate cancer remains unknown. After performing microarray analysis and reanalysis of the publicly available Memorial Sloan Kettering Cancer Center dataset, miR-200a expression was found higher in ADPC tissues and its expression was positively associated with survival of CRPC patients. In vitro studies showed that miR-200a overexpression in CRPC cells markedly suppressed cellular proliferation and facilitated apoptosis. In vivo studies indicated that overexpression of miR-200a inhibited growth and metastasis of prostate cancer. The luciferase reporter assay demonstrated that BRD4 is a direct target gene of miR-200a and it could reverse miR-200a-mediated biological effects in prostate cancer cells. Most importantly, our findings indicated that miR-200a suppresses the progression of CRPC by inhibiting the activation of BRD4-mediated AR signaling. This finding provides the foundation for the development of more personalized therapeutic approaches for CRPC patients.

To investigate the impact of Ramipril (RAM) on the expressions of insulin-like growth factor-1 (IGF-1) and renal mesangial matrix (RMM) in rats with diabetic nephropathy (DN).

Pseudogenes play a crucial role in cancer progression. However, the role of pituitary tumour-transforming 3, pseudogene (PTTG3P) in gastric cancer (GC) remains unknown. Here, we showed that PTTG3P expression was abnormally up-regulated in GC tissues compared with that in normal tissues both in our 198 cases of clinical samples and the cohort from The Cancer Genome Atlas (TCGA) database. High PTTG3P expression was correlated with increased tumour size and enhanced tumour invasiveness and served as an independent negative prognostic predictor. Moreover, up-regulation of PTTG3P in GC cells stimulated cell proliferation, migration and invasion both in vitro in cell experiments and in vivo in nude mouse models, and the pseudogene functioned independently of its parent genes. Overall, these results reveal that PTTG3P is a novel prognostic biomarker with independent oncogenic functions in GC.