The dopamine transporter (DAT) plays a critical role in terminating the action of dopamine by rapid reuptake into the presynaptic neuron. Previous studies have revealed that the DAT carboxyl terminus (DAT-CT) can directly interact with other cellular proteins and regulate DAT function and trafficking.

DNA methylation, the only known covalent modification of mammalian DNA, occurs primarily in CpG dinucleotides. 51% of CpGs in the human genome reside within repeats, and 25% within Alu elements. Despite that, no method has been reported for large-scale ascertainment of CpG methylation in repeats. Here we describe a sequencing-based strategy for parallel determination of the CpG-methylation status of thousands of Alu repeats, and a computation algorithm to design primers that enable their specific amplification from bisulfite converted genomic DNA. Using a single primer pair, we generated amplicons of high sequence complexity, and derived CpG-methylation data from 31 178 Alu elements and their 5' flanking sequences, altogether representing over 4 Mb of a human cerebellum epigenome. The analysis of the Alu methylome revealed that the methylation level of Alu elements is high in the intronic and intergenic regions, but low in the regions close to transcription start sites. Several hypomethylated Alu elements were identified and their hypomethylated status verified by pyrosequencing. Interestingly, some Alu elements exhibited a strikingly tissue-specific pattern of methylation. We anticipate the amplicons herein described to prove invaluable as epigenome representations, to monitor epigenomic alterations during normal development, in aging and in diseases such as cancer.

All antipsychotics work via dopamine D2 receptors (D2Rs), suggesting a critical role for D2Rs in psychosis; however, there is little evidence for a change in receptor number or pharmacological nature of D2Rs. Recent data suggest that D2Rs form dimers in-vitro and in-vivo, and we hypothesized that schizophrenia, as well as preclinical models of schizophrenia, would demonstrate altered dimerization of D2Rs, even though the overall number of D2Rs was unaltered.

TGF-beta has been postulated to play an important role in the maintenance of epithelial homeostasis and the development of epithelium-derived cancers. However, most of previous studies are mainly focused on the function of TGF-beta in immune cells to the development of allergic asthma and how TGF-beta signaling in airway epithelium itself in allergic inflammation is largely unknown. Furthermore, the in vivo TGF-beta function specifically in the airway epithelium during lung cancer development has been largely elusive.

Septic shock has a clinical mortality rate approaching fifty percent. The major clinical manifestations of sepsis are due to the dysregulation of the host's response to infection rather than the direct consequences of the invading pathogen. Central to this initial immunologic response is the activation of leukocytes and microvascular endothelium resulting in cardiovascular instability, lung injury and renal dysfunction. Due to the primary role of leukocyte activation in the sepsis syndrome, a synthetic biomimetic membrane, called a selective cytopheretic device (SCD), was developed to bind activated leukocytes. The incorporation of the SCD along an extracorporeal blood circuit coupled with regional anticoagulation with citrate to lower blood ionized calcium was devised to modulate leukocyte activation in sepsis.

The translocator protein 18 kDa (TSPO) is an attractive target for molecular imaging of neuroinflammation and tumor progression. [(18)F]PBR06, a fluorine-18 labeled form of PBR06, is a promising PET TSPO radioligand originally developed at NIMH. [(11)C]PBR06, a carbon-11 labeled form of PBR06, was designed and synthesized for the first time. The standard PBR06 was synthesized from 2,5-dimethoxybenzaldehyde in three steps with 71% overall chemical yield. The radiolabeling precursor desmethyl-PBR06 was synthesized from 2-hydroxy-5-methoxybenzaldehyde in five steps with 12% overall chemical yield. The target tracer [(11)C]PBR06 was prepared by O-[(11)C]methylation of desmethyl-PBR06 with [(11)C]CH(3)OTf in CH(3)CN at 80°C under basic condition and isolated by HPLC combined with SPE purification with 40-60% decay corrected radiochemical yield and 222-740 GBq/μmol specific activity at EOB. On the similar grounds, [(18)F]PBR06 was also designed and synthesized. The previously described Br-PBR06 precursor was synthesized from 2,5-dimethoxybenzaldehyde in two steps with 78% overall chemical yield. A new radiolabeling precursor tosyloxy-PBR06, previously undescribed tosylate congener of PBR06, was designed and synthesized from ethyl 2-hydroxyacetate, 4-methylbenzene-1-sulfonyl chloride, and N-(2,5-dimethoxybenzyl)-2-phenoxyaniline in four steps with 50% overall chemical yield. [(18)F]PBR06 was prepared by the nucleophilic substitution of either new tosyloxy-PBR06 precursor or known Br-PBR06 precursor in DMSO at 140°C with K[(18)F]F/Kryptofix 2.2.2 for 15 min and HPLC combined with SPE purification in 20-60% decay corrected radiochemical yield, >99% radiochemical purity, 87-95% chemical purity, and 37-222 GBq/μmol specific activity at EOB. Radiosynthesis of [(18)F]PBR06 using new tosylated precursor gave similar radiochemical purity, and higher specific activity, radiochemical yield and chemical purity in comparison with radiosynthesis using bromine precursor.

Receptor protein tyrosine phosphatase α (RPTPα)-mediated Src activation is required for survival of tested human colon and oestrogen receptor-negative breast cancer cell lines. To explore whether mutated RPTPα participates in human carcinogenesis, we sequenced RPTPα cDNAs from five types of human tumours and found splice mutants in ∼30% of colon, breast, and liver tumours. RPTPα245, a mutant expressed in all three tumour types, was studied further. Although it lacks any catalytic domain, RPTPα245 expression in the tumours correlated with Src tyrosine dephosphorylation, and its expression in rodent fibroblasts activated Src by a novel mechanism. This involved RPTPα245 binding to endogenous RPTPα (eRPTPα), which decreased eRPTPα-Grb2 binding and increased eRPTPα dephosphorylation of Src without increasing non-specific eRPTPα activity. RPTPα245-eRPTPα binding was blocked by Pro210 → Leu/Pro211 → Leu mutation, consistent with the involvement of the structural 'wedge' that contributes to eRPTPα homodimerization. RPTPα245-induced fibroblast transformation was blocked by either Src or eRPTPα RNAi, indicating that this required the dephosphorylation of Src by eRPTPα. The transformed cells were tumourigenic in nude mice, suggesting that RPTPα245-induced activation of Src in the human tumours may have contributed to carcinogenesis.

The rhizome of Smilax glabra has been used for a long time as both food and folk medicine in many countries. The present study focused on the active constituents from the rhizome of S. glabra, which possess potential anti-inflammatory activities. As a result, nine known compounds were isolated from the rhizome of S. glabra with the bioassay-guiding, and were identified as syringaresinol (1), lasiodiplodin (2), de-O-methyllasiodiplodin (3), syringic acid (4), 1,4-bis(4-hydroxy-3,5-dimethoxyphenyl)-2,3-bis(hydroxymethyl)-1,4-butanediol (5), lyoniresinol (6), trans-resveratrol (7), trans-caffeic acid methyl ester (8), and dihydrokaempferol (9). Among these compounds, 2 and 3 were isolated for the first time from S. glabra. In addition, the potential anti-inflammatory activities of the isolated compounds were evaluated in vitro in lipopolysaccharide- (LPS-) induced RAW264.7 cells. Results indicated that 4 and 7 showed significant inhibitory effects on NO production of RAW264.7 cells, and 1, 2, 3, and 5 showed moderate suppression effects on induced NO production. 1, 7, and 5 exhibited high inhibitory effects on TNF-α production, with the IC50 values less than 2.3, 4.4, and 16.6 μM, respectively. These findings strongly suggest that compounds 1, 2, 3, 4, 5, 7, and 9 were the potential anti-inflammatory active compositions of S. glabra.

Ornithuromorpha is the most inclusive clade containing extant birds but not the Mesozoic Enantiornithes. The early evolutionary history of this avian clade has been advanced with recent discoveries from Cretaceous deposits, indicating that Ornithuromorpha and Enantiornithes are the two major avian groups in Mesozoic. Here we report on a new ornithuromorph bird, Archaeornithura meemannae gen. et sp. nov., from the second oldest avian-bearing deposits (130.7 Ma) in the world. The new taxon is referable to the Hongshanornithidae and constitutes the oldest record of the Ornithuromorpha. However, A. meemannae shows few primitive features relative to younger hongshanornithids and is deeply nested within the Hongshanornithidae, suggesting that this clade is already well established. The new discovery extends the record of Ornithuromorpha by five to six million years, which in turn pushes back the divergence times of early avian lingeages into the Early Cretaceous.

Hybrid anticancer drugs are of great therapeutic interests as they can potentially overcome the deficiencies of conventional chemotherapy drugs and improve the efficacy. Many studies have revealed that the combination of histone deacetylase inhibitors (HDACi) and alkylating agents have synergistic effects. We reported a novel hybrid NL-101, in which the side chain of bendamustine was replaced with the hydroxamic acid of HDACi vorinostat (SAHA). NL-101 exhibited efficient anti-proliferative activity on myeloid leukemia cells especially Kasumi-1 and NB4 cells, accompanied by S phase arrest and caspase-3 dependent apoptosis. Importantly, it presented both the properties of HDAC inhibition and DNA damaging, as assessed by the acetylation of histone H3 and DNA double-strand breaks marker γ-H2AX. NL-101 also down-regulated the expression of anti-apoptotic protein Bcl-xL which was involved in the mitochondrial death pathway. Meanwhile, NL-101 induced apoptosis and DNA damage in primary cells from acute myeloid leukemia (AML) patients. NL-101 treatment could significantly prolong the survival time of t(8;21) leukemia mice with enhanced efficacy than bendamustine. These data demonstrate that NL-101 could be a potent and selective agent for leukemia treatment.

Renal ischemia and reperfusion (I/R) injury, which commonly occurs in kidney transplantation, is the leading cause of acute kidney injury. Picroside II possesses a wide range of pharmacological effects, including anti-apoptosis effects. In the present study, the ability of picroside II to attenuate apoptosis in a rat model of renal I/R injury was investigated. Sprague-Dawley rats were subjected to 45 min of ischemia followed by 24 h of reperfusion. Prior to reperfusion, the rats were treated with picroside II or an equal volume of phosphate-buffered saline. It was observed that renal function was significantly improved by the treatment with picroside II. Morphological analysis indicated that picroside II markedly reduced tissue damage and the expression of cleaved caspase-3. Reverse transcription-quantitative polymerase chain reaction and western blotting revealed that the expression levels of Bax and poly(ADP-ribose) polymerase-1 (PARP-1) were upregulated in the I/R group, whereas those of Bcl-2 were downregulated. However, the treatment with picroside II inhibited these changes induced by renal I/R injury. In conclusion, picroside II has potent anti-apoptotic activity against renal I/R injury.

Ischemia-reperfusion is a common injury of clinical ischemic disease and surgical lesions. Ischemic postconditioning (IPO) improves the ability of organs subjected to ischemia to tolerate injury. However, renal IPO studies have been based on animal models. In order to gain insights into IPO-induced alterations at the cellular level, an in vitro model for IPO was designed using the rat proximal tubular cell line NRK-52 E. This model was established by placing NRK-52 E cells in ischemic conditions for 3 h, then exposing cells to three cycles of reperfusion for 10 min and finally to ischemic conditions for 10 min (postconditioning). The cells were cultured further in reperfusion conditions for 3, 6, 12 and 24 h. Flow cytometry and Hoechst were used to assess apoptosis. The protein expression of B-cell lymphoma 2 (Bcl-2), Bcl-2-associated X protein (Bax), caspase-3, cleaved caspase-3 and caspase-8 were analyzed by western blotting. The results demonstrated that apoptosis occurred in cells subjected to ischemia/reperfusion (I/R) alone or with postconditioning following reperfusion for 24 h. Cells subjected to I/R demonstrated increased expression of Bax, cleaved caspase-3 and caspase-8 at the end of reperfusion. However, the levels of Bax, cleaved caspase-3 and caspase-8 were significantly attenuated in cells, which had undergone IPO. In conclusion, apoptosis was observed in cells subjected to 3 h of ischemia-reperfusion injury and IPO was able to inhibit this apoptosis. IPO inhibited apoptosis by inhibiting the caspase pathway thereby exerting protective effects.

Members of the microRNA (miR)-30 family have been reported to promote adipogenesis and inhibit osteogenesis, yet their role in the regulation of thermogenesis remains unknown. In this study, we show that miR-30b/c concentrations are greatly increased during adipocyte differentiation and are stimulated by cold exposure or the β-adrenergic receptor activator. Overexpression and knockdown of miR-30b and -30c induced and suppressed, respectively, the expression of thermogenic genes such as UCP1 and Cidea in brown adipocytes. Forced expression of miR-30b/c also significantly increased thermogenic gene expression and mitochondrial respiration in primary adipocytes derived from subcutaneous white adipose tissue, demonstrating a promoting effect of miRNAs on the development of beige fat. In addition, knockdown of miR-30b/c repressed UCP1 expression in brown adipose tissue in vivo. miR-30b/c targets the 3'-untranslated region of the receptor-interacting protein 140 (RIP140), and overexpression of miR-30b/c significantly reduced RIP140 expression. Consistent with RIP140 as a target of miR-30b/c in regulating thermogenic gene expression, overexpression of RIP140 greatly suppressed the promoting effect of miR-30b/c on the expression of UCP1 and Cidea in brown adipocytes. Taken together, the data from our study identify miR-30b/c as a key regulator of thermogenesis and uncover a new mechanism underlying the regulation of brown adipose tissue function and the development of beige fat.

Application of oxaliplatin for the treatment of pancreatic cancer (PC) is restricted owing to its toxic side effects and drug resistance. We investigated how withaferin A (WA), a bioactive component isolated from the medicinal plant Withania somnifera, acts synergistically with oxaliplatin on human PC in vitro and in vivo. We found that WA enhanced oxaliplatin-induced growth suppression and apoptosis in PC cells dramatically through a mechanism involving mitochondrial dysfunction and inactivation of the PI3K/AKT pathway. Combination treatment resulted in significant accumulation of intracellular reactive oxygen species (ROS). Pretreatment of cells with the ROS scavenger N-acetylcysteine completely blocked the apoptosis induced by combination treatment, and recovered expression of AKT inactivation, which revealed the important role of ROS in apoptosis and AKT regulation. In vivo, combination therapy showed the strongest anti-tumor effects compared with single agents, without obvious additional toxicity. These results support the notion that combination treatment with oxaliplatin and WA could facilitate development of an effective strategy for PC treatment.

Epithelial-mesenchymal transition (EMT) is associated with characteristics of breast cancer stem cells, including chemoresistance and radioresistance. However, it is unclear whether EMT itself or specific EMT regulators play causal roles in these properties. Here we identify an EMT-inducing transcription factor, zinc finger E-box binding homeobox 1 (ZEB1), as a regulator of radiosensitivity and DNA damage response. Radioresistant subpopulations of breast cancer cells derived from ionizing radiation exhibit hyperactivation of the kinase ATM and upregulation of ZEB1, and the latter promotes tumour cell radioresistance in vitro and in vivo. Mechanistically, ATM phosphorylates and stabilizes ZEB1 in response to DNA damage, ZEB1 in turn directly interacts with USP7 and enhances its ability to deubiquitylate and stabilize CHK1, thereby promoting homologous recombination-dependent DNA repair and resistance to radiation. These findings identify ZEB1 as an ATM substrate linking ATM to CHK1 and the mechanism underlying the association between EMT and radioresistance.

We characterized a novel group of HCV variants that are genetically related but distinct from each other belonging to genotype 6 (HCV-6). From 26 infected Austronesian-descended aborigines on Hainan Island, China, HCV sequences were determined followed by genetic analyses. Six nearly full-length genomes and 20 E1 sequences of HCV were obtained, which differ from each other and from all known HCV lineages by nucleotides above the intra-subtype level of 13%. Together with subtypes 6g and 6w, they constitute a phylogenetic group sharing a common ancestor dating from the end of the 12th century.

Knowledge of the distribution patterns of soil organic carbon (SOC) and factors that influence these patterns is crucial for understanding the carbon cycle. The objectives of this study were to determine the spatial distribution pattern of soil organic carbon density (SOCD) and the controlling factors in arid desert grasslands of northwest China. The above- and belowground biomass and SOCD in 260 soil profiles from 52 sites over 2.7×10(4) km2 were investigated. Combined with a satellite-based dataset of an enhanced vegetation index during 2011-2012 and climatic factors at different sites, the relationships between SOCD and biotic and abiotic factors were identified. The results indicated that the mean SOCD was 1.20 (SD:+/- 0.85), 1.73 (SD:+/- 1.20), and 2.69 (SD:+/- 1.91) kg m(-2) at soil depths of 0-30 cm, 0-50 cm, and 0-100 cm, respectively, which was smaller than other estimates in temperate grassland, steppe, and desert-grassland ecosystems. The spatial distribution of SOCD gradually decreased from the southeast to the northwest, corresponding to the precipitation gradient. SOCD increased significantly with vegetation biomass, annual precipitation, soil moisture, clay and silt content, and decreased with mean annual temperature and sand content. The correlation between BGB and SOCD was closer than the correlation between AGB and SOCD. Variables could together explain about 69.8%, 74.4%, and 78.9% of total variation in SOCD at 0-30 cm, 0-50 cm, and 0-100 cm, respectively. In addition, we found that mean annual temperature is more important than other abiotic factors in determining SOCD in arid desert grasslands in our study area. The information obtained in this study provides a basis for accurately estimating SOC stocks and assessing carbon (C) sequestration potential in the desert grasslands of northwest China.

We describe the landscape of somatic genomic alterations of 66 chromophobe renal cell carcinomas (ChRCCs) on the basis of multidimensional and comprehensive characterization, including mtDNA and whole-genome sequencing. The result is consistent that ChRCC originates from the distal nephron compared with other kidney cancers with more proximal origins. Combined mtDNA and gene expression analysis implicates changes in mitochondrial function as a component of the disease biology, while suggesting alternative roles for mtDNA mutations in cancers relying on oxidative phosphorylation. Genomic rearrangements lead to recurrent structural breakpoints within TERT promoter region, which correlates with highly elevated TERT expression and manifestation of kataegis, representing a mechanism of TERT upregulation in cancer distinct from previously observed amplifications and point mutations.

Mitogen-activated protein kinase kinase kinases (MAPKKKs; MAP3Ks) are important components of MAPK cascades, which are highly conserved signal transduction pathways in animals, yeast and plants, play important roles in plant growth and development. MAPKKKs have been investigated on their evolution and expression patterns in limited plants including Arabidopsis, rice and maize.

Maintaining the self-renewal of embryonic stem cells (ESCs) could be achieved by activating the extrinsic signaling, i.e., the use of leukemia inhibitory factor (LIF), or blocking the intrinsic differentiation pathways, i.e., the use of GSK3 and MEK inhibitors (2i). Here we found that even in medium supplemented with LIF, mESCs still tend to differentiate toward meso-endoderm lineages after long-term culture and the culture spontaneously secretes vascular endothelial growth factors (VEGFs). Blocking VEGF signaling with sunitinib, an anti-cancer drug and a receptor tyrosine kinase (RTK) inhibitor mainly targeting VEGF receptors (VEGFRs), is capable of maintaining the mESCs in the undifferentiated state without the need for feeder cells or LIF. Sunitinib facilitates the derivation of mESCs from blastocysts, and the mESCs maintained in sunitinib-containing medium remain pluripotent and are able to contribute to chimeric mice. Sunitinib also promotes iPSC generation from MEFs with only Oct4. Knocking down VEGFR2 or blocking it with neutralizing antibody mimicks the effect of sunitinib, indicating that blocking VEGF/VEGFR signaling is indeed beneficial to the self-renewal of mESCs. We also found that hypoxia-inducible factor alpha (HIF1α) and endoplasmic reticulum (ER) stress are involved in the production of VEGF in mESCs. Blocking both pathways inhibits the expression of VEGF and prevents spontaneous differentiation of mESCs. Interestingly, LIF may also exert its effect by downregulating HIF1α and ER stress pathways and subsequent VEGF expression. These results indicate the existence of an intrinsic differentiation pathway in mESCs by activating the autocrine VEGF signaling. Blocking VEGF signaling with sunitinib or other small molecules help to maintain the mESCs in the ground state of pluripotency.