Glioblastomas are the most common primary brain tumour in adults. While the prognosis for patients is poor, gene expression profiling has detected signatures that can sub-classify GBMs relative to histopathology and clinical variables. One category of GBM defined by a gene expression signature is termed ProNeural (PN), and has substantially longer patient survival relative to other gene expression-based subtypes of GBMs. Age of onset is a major predictor of the length of patient survival where younger patients survive longer than older patients. The reason for this survival advantage has not been clear.

Protein kinases have been found to possess two characteristic conformations in their activation-loops: the active DFG-in conformation and the inactive DFG-out conformation. Recently, it has been very interesting to develop type-II inhibitors which target the DFG-out conformation and are more specific than the type-I inhibitors binding to the active DFG-in conformation. However, solving crystal structures of kinases with the DFG-out conformation remains a challenge, and this seriously hampers the application of the structure-based approaches in development of novel type-II inhibitors. To overcome this limitation, here we present a computational approach for predicting the DFG-out inactive conformation using the DFG-in active structures, and develop related conformational selection protocols for the uses of the predicted DFG-out models in the binding pose prediction and virtual screening of type-II ligands. With the DFG-out models, we predicted the binding poses for known type-II inhibitors, and the results were found in good agreement with the X-ray crystal structures. We also tested the abilities of the DFG-out models to recognize their specific type-II inhibitors by screening a database of small molecules. The AUC (area under curve) results indicated that the predicted DFG-out models were selective toward their specific type-II inhibitors. Therefore, the computational approach and protocols presented in this study are very promising for the structure-based design and screening of novel type-II kinase inhibitors.

Curcumin has been shown to significantly improve spatial memory impairment induced by HIV-1 gp120 V3 in rats, but the electrophysiological mechanism remains unknown. Using extracellular microelectrode recording techniques, this study confirmed that the gp120 V3 loop could suppress long-term potentiation in the rat hippocampal CA1 region and synaptic plasticity, and that curcumin could antagonize these inhibitory effects. Using a Fura-2/AM calcium ion probe, we found that curcumin resisted the effects of the gp120 V3 loop on hippocampal synaptosomes and decreased Ca(2+) concentration in synaptosomes. This effect of curcumin was identical to nimodipine, suggesting that curcumin improved the inhibitory effects of gp120 on synaptic plasticity, ameliorated damage caused to the central nervous system, and might be a potential neuroprotective drug.

Curcumin improves the learning and memory deficits in rats induced by the gp120 V3 loop. The present study cultured rat hippocampal neurons with 1 nM gp120 V3 loop and 1 μM curcumin for 24 hours. The results showed that curcumin inhibited the gp120 V3 loop-induced mitochondrial membrane potential decrease, reduced the mRNA expression of the pro-apoptotic gene caspase-3, and attenuated hippocampal neuronal injury.

Membranous nephropathy (MN) is the major cause of adult nephrotic syndrome, which severely affects patients' quality of life. Currently, percutaneous renal biopsy is required to definitively diagnose MN. However, this technique is invasive and may cause severe complications. Therefore, an urgent clinical need exists for dynamic noninvasive monitoring of the renal state. In-depth molecular imaging studies could assist in finding a solution. Membrane attack complex C5b-9 is the key factor in the development of MN, and this protein primarily deposits in the glomerulus. The present study bound polyclonal antibodies to C5b-9 with ultrasmall superparamagnetic iron oxide (USPIO) nanoparticles to obtain C5b-9-targeted magnetic resonance molecular imaging probes. The probes were injected intravenously into rats with Heymann nephritis, a classic disease model of MN. The signal intensity in the T2*-weighted imaging of kidneys in vivo using 7.0 Tesla magnetic resonance imaging decreased significantly 24 hours after injection compared to the untargeted and control groups. This signal change was consistent with the finding of nanoparticle deposits in pathological glomeruli. This study demonstrated a novel molecular imaging technique for the assessment of MN.

Excessive reactive oxygen species (ROS) play an important role in the development of osteoporosis. Ophiopogonin D (OP-D), isolated from the traditional Chinese herbal agent Radix Ophiopogon japonicus, is a potent anti-oxidative agent. We hypothesized that OP-D demonstrates anti-osteoporosis effects via decreasing ROS generation in mouse pre-osteoblast cell line MC3T3-E1 subclone 4 cells and a macrophage cell line RAW264.7 cells. We investigated OP-D on osteogenic and osteoclastic differentiation under oxidative status. Hydrogen peroxide (H2O2) was used to establish an oxidative damage model. In vivo, we established a murine ovariectomized (OVX) osteoporosis model. Then, we searched the molecular mechanism of OP-D against osteoporosis. Our results revealed that OP-D significantly promoted the proliferation of MC3T3-E1 cells and improved some osteogenic markers. Moreover, OP-D reduced TRAP activity and the mRNA expressions of osteoclastic genes in RAW264.7 cells. OP-D suppressed ROS generation in both MC3T3-E1 and RAW264.7 cells. OP-D treatment reduced the activity of serum bone degradation markers, including CTX-1 and TRAP. Further research showed that OP-D displayed anti-osteoporosis effects via reducing ROS through the FoxO3a-β-catenin signaling pathway. In summary, our results indicated that the protective effects of OP-D against osteoporosis are linked to a reduction in oxidative stress via the FoxO3a-β-catenin signaling pathway, suggesting that OP-D may be a beneficial herbal agent in bone-related disorders, such as osteoporosis.

Memory type 1 T helper (T(H)1) cells are characterized by the stable expression of interferon (IFN)-γ as well as by the epigenetic imprinting of the IFNG locus. Among innate cells, NK cells play a crucial role in the defense against cytomegalovirus (CMV) and represent the main source of IFN-γ. Recently, it was shown that memory-like features can be observed in NK cell subsets after CMV infection. However, the molecular mechanisms underlying NK cell adaptive properties have not been completely defined. In the present study, we demonstrated that only NKG2Chi NK cells expanded in human CMV (HCMV) seropositive individuals underwent epigenetic remodeling of the IFNG conserved non-coding sequence (CNS) 1, similar to memory CD8(+) T cells or T(H)1 cells. The accessibility of the CNS1 was required to enhance IFN-γ transcriptional activity in response to NKG2C and 2B4 engagement, which led to consistent IFN-γ production in NKG2C(hi) NK cells. Thus, our data identify epigenetic imprinting of the IFNG locus as selective hallmark and crucial mechanism driving strong and stable IFN-γ expression in HCMV-specific NK cell expansions, providing a molecular basis for the regulation of adaptive features in innate cells.

An important therapeutic method of glioblastoma, the most common primary brain tumor, is radiotherapy. However, several studies reported recently that radiation could also promote the invasion and migration of malignant tumor. Herein, we have identified that a significant increase of migration and invasiveness of human glioma U251 cells undergoing X-ray was observed compared to controls, accompanied by the increase of cathepsin L (CTSL), which is a lysosomal cysteine protease overexpressed and secreted by tumor cells. To verify if there was a relationship between CTSL and the X-ray-induced glioma invasion, a CTSL specific inhibitor Z-FY-CHO or a short hairpin RNA interference was used to pretreat U251 cells. As a result, the cell invasion and migration was impaired via down-regulation of CTSL. Additionally, a marked reduction of the cell-signaling molecules Rho kinase was also detected compared with controls. We also found that CTSL is involved in EMT progress: both in vitro and in clinical specimens. Overall, our findings show that CTSL is an important protein which mediates cell invasion and migration of human glioma U251 cells induced by X-ray, and the inhibition of CTSL expression might diminish the invasion of U251 cells by reducing the activity of RhoA and CDC42 as well as EMT positive markers.

Based on the hybrid pharmacophore design concept, a novel series of dual diaryl urea and N-acylhydrazone derivatives were synthesized and evaluated for their in vitro cytotoxicity by the standard MTT assay. The pharmacological results indicated that most compounds exhibited moderate to excellent activity. Moreover, compound 2g showed the most potent cytotoxicity against HL-60, A549 and MDA-MB-231 cell lines, with IC50 values of 0.22, 0.34 and 0.41 μM, respectively, which was 3.8 to 22.5 times more active than the reference compounds sorafenib and PAC-1. The promising compound 2g thus emerges as a lead for further structural modifications.

Wnt/β-catenin signaling pathway is frequently dysregulated in human tumors and plays a critical role in tumorigenesis; however, the roles of microRNAs in mediating Wnt/β-catenin pathway are not well understood. Herein, we show that miR-30a-5p is activated by Wnt/β-catenin pathway through direct binding of β-catenin/TCF4 to two sites in the promoter region of miR-30a-5p. We also found that miR-30a-5p represses neural cell adhesion molecule (NCAM) expression by directly targeting two sites in the 3' untranslated region (3'-UTR) of NCAM mRNA. Moreover, Wnt/β-catenin pathway represses NCAM expression in glioma cells, which depends on miR-30a-5p. Finally, we found that miR-30a-5p promotes glioma cell growth invasion by repressing NCAM. Our findings demonstrate a novel Wnt/β-catenin-miR-30a-5p-NCAM regulatory axis which plays important roles in controlling glioma cell invasion and tumorigenesis.

Breast cancer (BC) is still a lethal threat to women worldwide. An accurate screening and diagnosis strategy performed in an easy-to-operate manner is highly warranted in clinical perspective. Besides the routinely focused protein markers, blood is full of small molecular metabolites with diverse structures and properties. This study aimed to screen metabolite markers with BC diagnosis potentials.

RNA Polymerase II Elongation Factor (ELL)-associated factor 2 (EAF2) is a tumor suppressor frequently down-regulated in human prostate cancer. We previously reported that its binding partner ELL1 can enhance EAF2 protein stability and activity. Here we show that EAF2 can be polyubiquitinated and its degradation blocked by proteasome inhibitor. Co-immunoprecipitation detected EAF2 binding to SIAH2, an E3 ligase, and SIAH2 overexpression enhanced polyubiquitination of EAF2. Co-transfection of EAF2 binding partner ELL1 blocked EAF2 ubiquitination, providing a mechanism for EAF2 stabilization. Finally, EAF2K81R mutant, which exhibits reduced polyubiquitination and increased stability, was more potent than wild-type EAF2 in apoptosis induction. These findings suggest that SIAH2 is an E3 ligase for EAF2 polyubiquitination and ELL1 can enhance EAF2 level and function by blocking its polyubiquitination.

HSV-1 infection in the cornea could lead to blindness. The infected cell polypeptide 4 (ICP4) of herpes simplex virus 1 (HSV-1) is a regulator of viral transcription that is required for productive infection. It has been previously demonstrated that miR-H6 encoded from HSV-1 genome targets ICP4 to help maintain latency. In this study, synthesized miR-H6 mimics were transfected into HSV-1-infected human cornea epithelial (HCE) cells. The inhibition of HSV-1 replication and viral ICP4 expression in miR-H6-transfected HCE was confirmed by plaque assay, immunofluorescence, and Western blot. Compared to nontransfection or mock, miR-H6 produced a low-titer HSV-1 and weak ICP4 expression. In addition, miR-H6 can decrease the interleukin 6 released into the medium, which was determined by ELISA. Taken together, the data suggests that miR-H6 targeting of ICP4 inhibits HSV-1 productive infection and decreases interleukin 6 production in HCE, and this may provide an approach to prevent HSV-1 lytic infection and inhibit corneal inflammation.

Gliomas are the most common and aggressive primary brain tumors for which unfortunately no effective treatment modalities exist despite advances in molecular biology as the knowledge base to unravel the extremely complex molecular mechanisms of tumorigenesis is limited. In this study an attempt has been made to understand the molecular pathological basis of tumorigenesis which led to an identification of an oncogene, CDC2, and an epigenetic strategy has been evaluated to control the tumorigensis by downregulating this oncogene.

To validate a next-generation sequencing (NGS)-based companion diagnostic using the MiSeqDx® sequencing instrument to simultaneously detect 56 RAS mutations in DNA extracted from formalin-fixed paraffin-embedded metastatic colorectal cancer (mCRC) tumor samples from the PRIME study. The test's ability to identify patients with mCRC likely to benefit from panitumumab treatment was assessed.

The development of biomaterials with the potential to accelerate wound healing is a great challenge in biomedicine. In this study, four types of samples including pepsin soluble collagen sponge (PCS), acid soluble collagen sponge (ACS), bovine collagen electrospun I (BCE I) and bovine collagen electrospun II (BCE II) were used as wound dressing materials. We showed that the PCS, ACS, BCE I and BCE II treated rats increased the percentage of wound contraction, reduced the inflammatory infiltration, and accelerated the epithelization and healing. PCS, ACS, BCE I, and BCE II significantly enhanced the total protein and hydroxyproline level in rats. ACS could induce more fibroblasts proliferation and differentiation than PCS, however, both PCS and ACS had a lower effect than BCE I and BCE II. PCS, ACS, BCE I, and BCE II could regulate deposition of collagen, which led to excellent alignment in the wound healing process. There were similar effects on inducing the level of cytokines including EGF, FGF, and vascular endothelial marker CD31 among these four groups. Accordingly, this study disclosed that collagens (PCS and ACS) from tilapia skin and bovine collagen electrospun (BCE I and BCE II) have significant bioactivity and could accelerate wound healing rapidly and effectively in rat model.

As human phosphodiesterase (PDE) proteins are attractive drug targets, a large number of selective PDE inhibitors have been developed. However, since the catalytic sites of PDE isoforms are conserved in sequence and structure, it remains unclear how these inhibitors discriminate PDE isoforms in a selective manner. Here we perform long-time scale molecular dynamics (MD) simulations to investigate the spontaneous association processes of a highly selective PDE2A inhibitor (BAY60-7550) with the catalytic pockets of six PDE isoforms. We found that the free-energy landscapes of PDE:BAY60-7550 interactions on the PDE surfaces are very different between various PDE isoforms; and the free-energy landscape of PDE2A forms a favorable low-energy pathway that not only drives BAY60-7550 toward the target binding site, but also guides BAY60-7750 to adopt its native binding conformation known from crystal structure. Thus, this study reveals that the inhibitor interactions with the PDE surface residues play an important role in its high selectivity for PDE2A, and thereby provides new fundamental insights into the PDE isoform-specific inhibitor selectivity.

Acute aortic dissection (AAD) is a life-threatening disease. Despite the higher risk of mortality, currently there are no effective therapies that can ameliorate AAD development or progression. Identification of meaningful clusters of co-expressed genes or representative biomarkers for AAD may help to identify new pathomechanisms and foster development of new therapies. To this end, we performed a weighted gene co-expression network analysis (WGCNA) and calculated module-trait correlations based on a public microarray dataset (GSE 52093) and discovered 9 modules were found to be related to AAD. The module which has the strongest positive correlation with AAD was further analyzed and the top 10 hub genes SLC20A1, GINS2, CNN1, FAM198B, MAD2L2, UBE2T, FKBP11, SLMAP, CCDC34, and GALK1 were identified. Furthermore, we validated the data by qRT-PCR in an independent sample set originated from our study center. Overall, the qRT-PCR results were consistent with the results of the microarray analysis. Intriguingly, the highest change was found for FKBP11, a protein belongs to the FKBP family of peptidyl-prolyl cis/trans isomerases, which catalyze the folding of proline-containing polypeptides. In congruent with the gene expression analysis, FKBP11 expression was induced in cultured endothelial cells by angiotensin II treatment and endothelium of the dissected aorta. More importantly we show that FKBP11 provokes inflammation in endothelial cells by interacting with NF-kB p65 subunit, resulting in pro-inflammatory cytokines production. Accordingly, siRNA mediated knockdown of FKBP11 in cultured endothelial cells suppressed angiotensin II induced monocyte transmigration through the endothelial monolayer. Based on these data, we hypothesize that pro-inflammatory cytokines elicited by FKBP11 overexpression in the endothelium under AAD condition could facilitate transendothelial migration of the circulating monocytes into the aorta, where they differentiate into active macrophages and secrete MMPs and other extracellular matrix (ECM) degrading proteins, contributing to sustained inflammation and AAD. Taken together, our data identify important role of FKBP11 which can serve as biomarker and/or therapeutic target for AAD.

On⁻off⁻on fluorescent sensors based on emerging carbon nanoparticles (CNPs) or carbon dots (CDs) have attracted extensive attention for their convenience and efficiency. In this study, dumped silkworm excrement was used as a novel precursor to prepare fluorescent nitrogen-doped CNPs (N-CNPs) through hydrothermal treatment. The obtained N-CNPs showed good photoluminescent properties and excellent water dispersibility. Thus, they were applied as fluorescence “on⁻off⁻on” probes for the detection of Fe(III) and biothiols. The “on⁻off” process was achieved by adding Fe(III) into N-CNP solution, which resulted in the selective fluorescence quenching, with the detection limit of 0.20 μM in the linear range of 1⁻500 μM. Following this, the introduction of biothiols could recover the fluorescence efficiently, in order to realize the “off⁻on” process. By using glutathione (GSH) as the representative, the linear range was in the range of 1⁻1000 μM, and the limit of detection was 0.13 μM. Moreover, this useful strategy was successfully applied for the determination of amounts of GSH in fetal calf serum samples.

Magnetic micromachines as wireless end-effectors have been widely applied for drug discovery and regenerative medicine. Yet, the magnetic assembly of arbitrarily shaped cellular microstructures with high efficiency and flexibility still remains a big challenge. Here, a novel clamp-shape micromachine using magnetic nanoparticles was developed for the indirect untethered bioassembly. With a multi-layer template, the nickel nanoparticles were mixed with polydimethylsiloxane (PDMS) for mold replication of the micromachine with a high-resolution and permeability. To actuate the micromachine with a high flexibility and large scalable operation range, a multi-pole electromagnetic system was set up to generate a three-dimensional magnetic field in a large workspace. Through designing a series of flexible translations and rotations with a velocity of 15mm/s and 3 Hz, the micromachine realized the propel-and-throw strategy to overcome the inevitable adhesion during bioassembly. The hydrogel microstructures loaded with different types of cells or the bioactive materials were effectively assembled into microtissues with reconfigurable shape and composition. The results indicate that indirect magnetic manipulation can perform an efficient and versatile bioassembly of cellular micromodules, which is promising for drug trials and modular tissue engineering.