The genomic origins of the Bronze Age Tarim Basin mummies.

The identity of the earliest inhabitants of Xinjiang, in the heart of Inner Asia, and the languages that they spoke have long been debated and remain contentious1. Here we present genomic data from 5 individuals dating to around 3000-2800 BC from the Dzungarian Basin and 13 individuals dating to around 2100-1700 BC from the Tarim Basin, representing the earliest yet discovered human remains from North and South Xinjiang, respectively. We find that the Early Bronze Age Dzungarian individuals exhibit a predominantly Afanasievo ancestry with an additional local contribution, and the Early-Middle Bronze Age Tarim individuals contain only a local ancestry. The Tarim individuals from the site of Xiaohe further exhibit strong evidence of milk proteins in their dental calculus, indicating a reliance on dairy pastoralism at the site since its founding. Our results do not support previous hypotheses for the origin of the Tarim mummies, who were argued to be Proto-Tocharian-speaking pastoralists descended from the Afanasievo1,2 or to have originated among the Bactria-Margiana Archaeological Complex3 or Inner Asian Mountain Corridor cultures4. Instead, although Tocharian may have been plausibly introduced to the Dzungarian Basin by Afanasievo migrants during the Early Bronze Age, we find that the earliest Tarim Basin cultures appear to have arisen from a genetically isolated local population that adopted neighbouring pastoralist and agriculturalist practices, which allowed them to settle and thrive along the shifting riverine oases of the Taklamakan Desert.

Satb2 regulates the development of dopaminergic neurons in the arcuate nucleus by Dlx1.

Dopaminergic (DA) neurons in the arcuate nucleus (ARC) of the hypothalamus play essential roles in the secretion of prolactin and the regulation of energy homeostasis. However, the gene regulatory network responsible for the development of the DA neurons remains poorly understood. Here we report that the transcription factor special AT-rich binding protein 2 (Satb2) is required for the development of ARC DA neurons. Satb2 is expressed in a large proportion of DA neurons without colocalization with proopiomelanocortin (POMC), orexigenic agouti-related peptide (AgRP), neuropeptide-Y (NPY), somatostatin (Sst), growth hormone-releasing hormone (GHRH), or galanin in the ARC. Nestin-Cre;Satb2flox/flox (Satb2 CKO) mice show a reduced number of ARC DA neurons with unchanged numbers of the other types of ARC neurons, and exhibit an increase of serum prolactin level and an elevated metabolic rate. The reduction of ARC DA neurons in the CKO mice is observed at an embryonic stage and Dlx1 is identified as a potential downstream gene of Satb2 in regulating the development of ARC DA neurons. Together, our study demonstrates that Satb2 plays a critical role in the gene regulatory network directing the development of DA neurons in ARC.

Post-last glacial maximum expansion of Y-chromosome haplogroup C2a-L1373 in northern Asia and its implications for the origin of Native Americans.

OBJECTIVES: Subbranches of Y-chromosome haplogroup C2a-L1373 are founding paternal lineages in northern Asia and Native American populations. Our objective was to investigate C2a-L1373 differentiation in northern Asia and its implications for Native American origins. MATERIALS AND METHODS: Sequences of rare subbranches (n = 43) and ancient individuals (n = 37) of C2a-L1373 (including P39 and MPB373), were used to construct phylogenetic trees with age estimation by BEAST software. RESULTS: C2a-L1373 expanded rapidly approximately 17.7,000-14.3,000 years ago (kya) after the last glacial maximum (LGM), generating numerous sublineages which became founding paternal lineages of modern northern Asian and Native American populations (C2a-P39 and C2a-MPB373). The divergence pattern supports possible initiation of differentiation in low latitude regions of northern Asia and northward diffusion after the LGM. There is a substantial gap between the divergence times of C2a-MPB373 (approximately 22.4 or 17.7 kya) and C2a-P39 (approximately 14.3 kya), indicating two possible migration waves. DISCUSSION: We discussed the decreasing time interval of "Beringian standstill" (2.5 ky or smaller) and its reduced significance. We also discussed the multiple possibilities for the peopling of the Americas: the "Long-term Beringian standstill model," the "Short-term Beringian standstill model," and the "Multiple waves of migration model." Our results support the argument from ancient DNA analyses that the direct ancestor group of Native Americans is an admixture of "Ancient Northern Siberians" and Paleolithic communities from the Amur region, which appeared during the post-LGM era, rather than ancient populations in greater Beringia, or an adjacent region, before the LGM.

Analysis of the water-soluble polysaccharides from Camellia japonica pollen and their inhibitory effects on galectin-3 function.

Pollen has been defined as dietary supplement used to supplement the diet in many countries, but the primary structure and activity of Camellia japonica pollen polysaccharide remain unclear. In this study, the water-soluble polysaccharide extracted from Camellia japonica pollen (WCPP) was fractionated into one neutral fraction (WCPP-N) and two acidic fractions (WCPP-A1 and WCPP-A2) by DEAE-cellulose column, and WCPP-A2 was further fractionated into two homogeneous sub-fractions (WCPP-A2a and WCPP-A2b) by Sepharose CL-6B column. Monosaccharide composition results showed that WCPP-N might mainly contain starch-like glucan as well as some arabinogalactan, while WCPP-A1, WCPP-A2 and its sub-fractions might mainly composed of rhamnogalacturonan I (RG-I) pectic polysaccharide domain backbone with some different types of side chains, including arabinan, galactan, and/or arabinogalactan. The primary structure analysis of WCPP-A2a by NMR spectra analysis suggested that WCPP-A2a was an RG-I-like pectic polysaccharide, branched at the O-4 of Rha residues in the backbone, with α-(1 â†’ 3,5)-L-arabinan as well as type-II arabinogalactan side chain to which were attached. The results of galectin-3-mediated hemagglutination assay indicated that WCPP-A2a exhibited the strongest inhibitory effect on galectin-3 with MIC value around 0.27 µg/mL. These results suggested the potential use of Camellia japonica pollen polysaccharide as a galectin3 inhibitor in functional foods.

Simultaneous quantification of chlorogenic acid and taurocholic acid in human plasma by LC-MS/MS and its application to a pharmacokinetic study after oral administration of Shuanghua Baihe tablets.

An LC-MS/MS method was developed and validated for the simultaneous quantification of chlorogenic acid (CGA) and taurocholic acid (TCA) in human plasma using hydrochlorothiazide as the internal standard. The chromatographic separation was achieved on a Hedera ODS-2 column with a gradient elution using 10 mmol·L(-1) of ammonium acetate buffer solution containing 0.5% of formic acid - acetonitrile as mobile phase at a flow rate of 300 µL·min(-1). The detection was performed on a triple quadrupole tandem mass spectrometer by multiple reaction monitoring in negative ESI mode. The method was fully validated over the concentration ranges of 0.1-10 ng·mL(-1) for CGA and 2-150 ng·mL(-1) for TCA. It was successfully applied to a pharmacokinetic study of CGA and TCA in healthy Chinese volunteers after oral administration of Shuanghua Baihe tablets (SBTs). In the single-dose study, the maximum plasma concentration (Cmax), time to reach Cmax (Tmax) and elimination half-life (t1/2) of CGA were (0.763 8 ± 0.542 0) ng·mL(-1), (1.0 ± 0.5) h, and (1.3 ± 0.6) h, respectively. In the multiple-dose study, the Cmax, Tmax and t1/2 of CGA were (0.663 7 ± 0.583 3) ng·mL(-1), (1.1 ± 0.5) h, and (1.4 ± 0.7) h, respectively. For TCA, no significant characteristic increasing plasma TCA concentration-time curve was found in the volunteers after oral administration of SBTs, indicating its complicated process in vivo as an endogenous ingredient.

Simultaneous determination of corynoline and acetylcorynoline in human urine by LC-MS/MS and its application to a urinary excretion study.

Corynoline and acetycorynoline, the major active components derived from Corydalis bungeana Herba, showed multiple pharmacological activities. However, quantification of the two compounds in human urine has not been reported. A simple liquid chromatography with tandem mass spectrometry method for the simultaneous determination of corynoline and acetycorynoline in human urine has been developed and fully validated. The analytes were extracted from urine samples by simple liquid-liquid extraction. Chromatographic separation was achieved on a Hedera ODS-2C18 column with the mobile phase of water (containing 0.5% formic acid) and acetonitrile (28:72, v/v) at a flow rate of 0.4mL/min. A tandem mass spectrometric detection was conducted using multiple reaction monitoring via an electrospray ionization source in positive mode. The monitored ion transitions were m/z 368.1→289.1 for corynoline, m/z 410.2→289.2 for acetycorynoline and m/z 380.2→243.2 for donepezil (internal standard), respectively. The calibration curves were linear (correlation coefficients>0.9970) over the concentration ranges of 3.0-3000pg/mL for corynoline and 3.0-1000pg/mL for acetycorynoline. The established method was highly sensitive with the lower limit of quantification (LLOQ) of 3.0pg/mL for both analytes. The intra- and inter-day precision was lower than 10% in terms of relative standard deviation for the low, medium, and high quality control samples, and lower than 16% for the LLOQ samples of the analytes. The accuracy was within ±10% in terms of relative error for both analytes. The method was successfully applied to a urinary excretion study after oral administration of the Chinese medicine formula Shuanghua Baihe tablets in healthy volunteers. The urinary excretion profiles of corynoline and acetycorynoline in human were first reported. The results of this study suggest that renal excretion was not the main excretion pathway of corynoline and acetycorynoline in humans.

LC-MS/MS determination and urinary excretion study of seven alkaloids in healthy Chinese volunteers after oral administration of Shuanghua Baihe tablets.

An LC-MS/MS method was developed and validated for the simultaneous determination of magnoflorine, berberrubine, jatrorrhizine, coptisine, epiberberine, palmatine and berberine in human urine. The sample preparation procedure involved the four-fold dilution of the urine samples with acetonitrile/water (1:3, v/v). The chromatographic separation was achieved on a Hedera ODS-2 column under gradient elution at a flow rate of 0.4 mL/min with acetonitrile and water containing 0.5% formic acid as the mobile phase. The mass detection was performed in the positive mode. Calibration curves of the seven alkaloids showed good linearity (correlation coefficients>0.9973) over their concentration ranges. To meet the requirements of urinary excretion study for each alkaloid in human, the lower limit of quantification was set at different values from 0.05063 ng/mL to 2.034 ng/mL for the seven alkaloids, respectively. The intra- and inter-batch precision and accuracy were all within ± 15%. No matrix effect was observed for the analytes. The validated method was applied to the excretion study for the seven alkaloids in healthy Chinese volunteers after oral administration of Shuanghua Baihe tablets. The average 72 h cumulative urinary excretion of magnoflorine, berberrubine, jatrorrhizine, coptisine, epiberberine, palmatine and berberine accounted for 1.81%, 0.27%, 0.29%, 0.046%, 0.027%, 0.010% and 0.021% of the respective administered dose.

Highly sensitive method for simultaneous determination of nine alkaloids of Shuanghua Baihe tablets in human plasma by LC-MS/MS and its application.

Shuanghua Baihe tablets (SBT) is a traditional Chinese medicinal formula which has been used to treat recurrent aphthous stomatitis for many years. To study the pharmacokinetic profiles of berberine, epiberberine, coptisine, palmatine, jatrorrhizine, magnoflorine, berberrubine, corynoline and acetylcorynoline in human after administration of SBT, a sensitive liquid chromatography-tandem mass spectrometry method was developed and fully validated for the simultaneous quantification of these nine alkaloids in human plasma. After protein precipitation, the nine alkaloids in human plasma sample was separated on a Hanbon C18 (150mm×2.1mm, 5µm) column with gradient elution using methanol and 0.5% formic acid water solution, and detected by a triple quadrupole mass spectrometer with an electrospray ionization source. It is a challenge to design different calibration ranges for different analytes in a bioanalytical method for simultaneous determination of multi-analytes in bio-samples. To ensure that each alkaloid in the plasma was determined accurately by the simultaneous quantitation method, the upper limits of quantification for the nine alkaloids were designed at 100, 300, 800, 1800 and 5000pg/mL, respectively, according to the maximum plasma concentration value of each alkaloid obtained from the pilot pharmacokinetic study. The lower limit of quantification was 15pg/mL for berberine, epiberberine, coptisine, magnoflorine, berberrubine, corynoline and acetylcorynoline, while for palmatine and jatrorrhizine it was 1.5pg/mL. This method was successfully applied to investigate the pharmacokinetic profiles of the nine alkaloids in healthy Chinese volunteers after a single oral administration of SBT.

Inhibition of MITF and tyrosinase by paeonol-stimulated JNK/SAPK to reduction of phosphorylated CREB.

Tyrosinase and its transcriptional regulator microphthalmia-associated transcription factor (MITF) play critical roles in regulation of melanogenesis, and are required for environmental cues or agents in modulation of melanin synthesis. Identifying the signals regulating tyrosinase and MITF is crucial to understanding how pigmentation responds to extracellular stimuli. In this report, we discovered that paeonol down-regulated melanin production via decreasing MITF expression and consequent mRNA and protein levels of tyrosinase. We also found that paeonol reduced phosphorylation of a cAMP responsive element binding protein (phospho-CREB), which binds and activates MITF. A selective inhibitor of c-jun N-terminal or stress-activated protein kinases (JNK/SAPK)-SP600125 significantly reversed paeonol-induced down-regulation of melanogenesis. Inhibition of cAMP/PKA pathway intensified the hypopigmentation response to paeonol. These results identify a mechanism in which paeonol induces the down-regulation of melanogenesis through inhibition of CREB phosphorylation, leading to the expression reduction of MITF and subsequently tyrosinase. The key kinase mediating the effects of paeonol on melanogenesis in B16F10 cells is JNK/SAPK. Additionally, the cAMP/PKA pathway may take part in this process.

Down-regulation of melanin synthesis and transfer by paeonol and its mechanisms.

Down-regulation of melanin synthesis and\or melanin transfer are/is required for recovery of pigmentary disorders. It is known that direct inhibitors of tyrosinase, the key enzyme in melanin synthesis, such as hydroquinone with a phenol structure, suppress melanin synthesis. We screened some herbal monomers using human melanocytes and found that paeonol, a major phenolic component of Moutan Cortex, down-regulated melanin synthesis. The melanin synthesis and tyrosinase activity were inhibited by paeonol in a dose-dependent manner. The expression levels of tyrosinase mRNA and protein were also reduced by paeonol. We further studied the inhibitory effects of paeonol on melanin transfer in co-culture of melanocytes and keratinocytes. More than 50% of inhibition of melanin transfer was observed at concentration of 200 microM of paeonol and the increased melanin transfer induced by SLIGRL, the PAR-2 activating peptide, was also reduced by paeonol. However, paeonol did not influence the expression level of PAR-2 mRNA in co-culture cells. These results indicate that the depigmenting effect of paeonol might be due to its down-regulation of melanogenesis and melanin transfer.

Triptolide impairs dendritic cell migration by inhibiting CCR7 and COX-2 expression through PI3-K/Akt and NF-kappaB pathways.

Inhibition of dendritic cell (DC) migration into tissues and secondary lymphoid organs is an efficient way to induce immunosuppression and tolerance. CCR7 and PGE(2) are critical for DC migration to secondary lymphoid organs where DC initiate immune response. Triptolide, an active component purified from the medicinal plant Tripterygium Wilfordii Hook F., is a potent immunosuppressive drug capable of prolonging allograft survival in organ transplantation by inhibiting T cell activation and proliferation. Considering the essential role in T cell tolerance of DC migration to secondary lymphoid organs, here we demonstrate that triptolide can significantly inhibit LPS-triggered upregulation of CCR7 expression and PGE(2) production by inhibiting cyclooxygenase-2 (COX-2) expression in DC, thus impairing DC migration towards CCR7 ligand CCL19/MIP-3betain vitro. Moreover, triptolide-treated DC display impaired migration into secondary lymphoid organs and in vivo administration of triptolide also inhibits DC migration. Further studies show that the triptolide-mediated inhibitory effects of LPS-induced activation of phosphatidylinositol-3 kinase (PI3-K)/Akt and nuclear NF-kappaB activation are involved in down-regulation of COX-2 and CCR7 expression resulting in impaired migration to secondary lymphoid organs of DC. Therefore, inhibition of DC migration through decreasing COX-2 and CCR7 expression via PI3-K/Akt and NF-kappaB signal pathways provides additional mechanistic explanation for triptolide's immunosuppressive effect.

Immunosuppressant triptolide inhibits dendritic cell-mediated chemoattraction of neutrophils and T cells through inhibiting Stat3 phosphorylation and NF-kappaB activation.

Triptolide, an active component purified from the medicinal plant Tripterygium wilfordii Hook F., is potent in anti-inflammation and immunosuppression. Dendritic cells (DC), one of important targets of immunosuppressants, play crucial roles in linking the innate immunity and adaptive immunity. However, the effects of triptolide on DC have not been fully elucidated. Chemoattraction of neutrophils and T cells by DC may favor their interactions and initiation of immune response. Here we demonstrate that triptolide significantly impairs DC-mediated chemoattraction of neutrophils and T cells both in vitro and in vivo by suppressing DC production of CC and CXC chemokines including MIP-1alpha, MIP-1beta, MCP-1, RANTES, TARC, and IP-10 in response to LPS. Furthermore, triptolide-mediated inhibition of NF-kappaB activation, Stat3 phosphorylation and increase of SOCS1 expression in DC may be involved in the inhibitory effect of triptolide. Our study provides a novel mechanistic explanation for the anti-inflammatory and immunosuppressive activities of triptolide.

Triptolide (PG-490) induces apoptosis of dendritic cells through sequential p38 MAP kinase phosphorylation and caspase 3 activation.

Dendritic cells (DCs) are the most potent antigen-presenting cells that play crucial roles in the regulation of immune response. Triptolide, an active component purified from the medicinal plant Tripterygium wilfordii Hook F., has been demonstrated to act as a potent immunosuppressive drug capable of inhibiting T cell activation and proliferation. However, little is known about the effects of triptolide on DCs. The present study shows that triptolide does not affect phenotypic differentiation and LPS-induced maturation of murine DCs. But triptolide can dramatically reduce cell recovery by inducing apoptosis of DCs at concentration as low as 10ng/ml, as demonstrated by phosphatidylserine exposure, mitochondria potential decrease, and nuclear DNA condensation. Triptolide induces activation of p38 in DCs, which precedes the activation of caspase 3. SB203580, a specific kinase inhibitor for p38, can block the activation of caspase 3 and inhibit the resultant apoptosis of DCs. Our results suggest that the anti-inflammatory and immunosuppressive activities of triptolide may be due, in part, to its apoptosis-inducing effects on DCs.