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Tea (Camellia sinensis [L.] O. Kuntze) tree is a perennial plant in which winter dormancy is an important biological adaptation to environmental changes. We discovered and reported a novel tea tree cultivar that can generate tender shoots in winter several years ago, but the molecular mechanism for this unique phenotype remains unknown . Here, we conducted comparative transcriptomics, proteomics and metabolomics along with phytohormone quantitation between the winter and spring tender shoots to investigate the physiological basis and putative regulatory mechanisms of its evergrowing character during winter. Our multi-omics study has led to the following findings. Gibberellin (GA) levels and key enzymes for GA biosynthesis and the signal transduction pathway were increased in the winter shoots, causing the ABA/GA content ratio to decrease, which might play a key regulatory role in maintaining normal growth during winter. The abundance of proteins, genes and metabolites involved in energy metabolism was all increased in winter shoots, indicating that energy is critical for continuous growth under the relatively weak-light and low-temperature environment. Abiotic resistance-related proteins and free amino acids were also increased in abundance in the winter shoots, which possibly represents an adaptation response to winter conditions. These results allowed us to hypothesize a novel molecular mechanism of adaptation for this unique tender shoot evergrowing in winter.

Circular RNAs (circRNAs) play an important role in cancer development and progression by regulating gene expression. The present study aimed to investigate the function of circRNA_100859 in colon cancer. circRNA expression profiles from a human circRNAs chip were analyzed. The effects of circRNA_100859 on cell proliferation and apoptosis were assessed in vitro and interactions between circRNA_100859 and its micro (mi)RNA and target genes were analyzed. The diagnostic and prognostic significance of circRNA_100859 was also investigated. It was identified that circRNA_100859 was overexpressed in colon cancer tissues and promoted cell proliferation and inhibited cell apoptosis. Additionally, bioinformatics and a dual-luciferase reporter assay confirmed that circRNA_100859 acted as a miR-217 sponge, and miR-217 directly targeted hypoxia-inducible factor (HIF)-1α. Rescue assays demonstrated that HIF-1α protein and mRNA expression levels and cell proliferation were regulated by the circRNA_100859/miR-217 axis (P

P2X7 receptor (P2X7R) is an ATP-gated non-selective cation channel which mediates ATP-induced inflammation in macrophages. Transient receptor potential (TRP) receptors are nociceptors in cellular membrane which can perceive the stimuli of environmental irritant. The interaction between TRP channels and P2X7R has been found while the details about inflammation are still unclear. In this study, we suggested that transient receptor potential ankyrin 1 (TRPA1), a member of TRP superfamily, participates in ATP-induced oxidative stress and inflammation in human acute monocytic leukemia cell line (THP-1)-derived macrophage. The co-localization between TRPA1 and P2X7R was detected using immunofluorescence in THP-1-derived macrophage and transfected human embryonic kidney cell line (HEK293T). The mechanism by which ATP or 3'-O-(4-Benzoylbenzoyl)-ATP (BzATP) induces the activation of macrophages was verified by calcium imaging, mitochondrial reactive oxygen species (mtROS) detection, mitochondrial membrane potential (∆Ψm) measurement, flow cytometry, enzyme-linked immunosorbent assay (ELISA), western blotting, CCK-8 assay, and the lactate dehydrogenase (LDH) release cytotoxic assay. The BzATP and ATP induced calcium overload, mitochondria injury, interleukin-1β (IL-1β) secretion, and cytotoxicity can be inhibited by TRPA1 antagonists. These results indicated that TRPA1 can co-localize with P2X7R and mediate ATP-induced oxidative stress and inflammation. Therefore, the inhibition of TRPA1 may provide a potential therapy for ATP-elicited inflammatory diseases, including atherosclerosis.

Transient receptor potential ankyrin 1 (TRPA1), the non-selective cation channel, was found that can mediate the generation of multiple sclerosis, while the mechanism is still controversial. Lysophosphatidylcholine (LPC) is a critical trigger of multiple sclerosis which results from the syndrome of neuronal inflammation and demyelination. In this work, we suggested that TRPA1 can mediate the LPC-induced oxidative stress and cytotoxicity in OLN-93 oligodendrocyte. The expression of TRPA1 in OLN-93 was detected by using quantitative real-time PCR (qRT-PCR) and immunofluorescence. The calcium overload induced by LPC via TRPA1 was detected by calcium imaging. The mechanism of LPC-induced mitochondrial reactive oxygen species (mtROS) generation, mitochondria membrane depolarization, nitric oxide (NO) increase, and development of superoxide production via TRPA1 was verified by using confocal imaging. The cell injury elicited by LPC via TRPA1 was confirmed by both CCK-8 and LDH cytotoxicity detection. These results indicate that TRPA1 plays an important role of the LPC-induced oxidative stress and cell damage in OLN-93 oligodendrocyte. Therefore, inhibition of TRPA1 may protect the LPC-induced demyelination.

Lysophosphatidylcholine (LPC) is a major atherogenic lipid that stimulates an increase in mitochondrial reactive oxygen species (mtROS) and the release of cytokines under inflammasome activation. However, the potential receptors of LPC in macrophages are poorly understood. Members of the transient receptor potential (TRP) channel superfamily, which is crucially involved in transducing environmental irritant stimuli into nociceptor activity, are potential receptors of LPC. In this study, we investigated whether LPC can induce the activation of transient receptor potential ankyrin 1 (TRPA1), a member of the TRP superfamily. The functional expression of TRPA1 was first detected by quantitative real-time polymerase chain reaction (qRT-PCR), western blotting and calcium imaging in human acute monocytic leukemia cell line (THP-1)-derived macrophages. The mechanism by which LPC induces the activation of macrophages through TRPA1 was verified by cytoplasmic and mitochondrial calcium imaging, mtROS detection, a JC-1 assay, enzyme-linked immunosorbent assay, the CCK-8 assay and the lactate dehydrogenase (LDH) cytotoxic assay. LPC induced the activation of THP-1-derived macrophages via calcium influx, and this activation was suppressed by potent and selective inhibitors of TRPA1. These results indicated that TRPA1 can mediate mtROS generation, mitochondrial membrane depolarization, the secretion of IL-1β and cytotoxicity through cellular and mitochondrial Ca2+ influx in LPC-treated THP-1-derived macrophages. Therefore, the inhibition of TRPA1 may protect THP-1-derived macrophages against LPC-induced injury.

This paper reported a simple hydrothermal method to prepare MoS2 with different surface affinity. By adjusting the reaction time and Mo/S proportion of hydrothermal synthesis, MoS2 powder with a face-to-edge ratio of 1.19 can be obtained on the surface. Interestingly, for the first time, MoS2 with such ratio can function as an intermediary similar to surfactant. It tends to distribute on the interface between water and n-hexane, and further adsorb methylene blue from water to the interface. By this route, it has a great implication for the interaction of incompatible molecules and the interfacial process in future applications.

Vitamin D plays a major role in the regulation of calcium homeostasis and affects bone metabolism. There is currently limited detailed knowledge about the vitamin D endocrine system in human bone cells. Here, we investigated the direct effects of 1α, 25-dihydroxyvitamin D₃ (1α, 25-(OH)₂D₃ or 'VD3') on osteogenesis of human umbilical cord mesenchymal stem cells (HUCMSCs). We also studied the impact of VD3 on intracellular Ca2+ concentrations in osteogenic cells. The results of alizarin red staining and alkaline phosphatase activity tests showed that VD3 could not induce osteogenic differentiation in HUCMSCs. However, addition of VD3 to the osteogenic differentiationinducing medium could promote HUCMSC to differentiate into osteoblasts. Calcium imaging showed that the addition of VD3 increased intracellular Ca2+ concentrations in osteogenic HUCMSCs. Thus, we concluded that adding VD3 increased intracellular Ca2+ concentrations in osteogenic HUCMSCs and promoted their osteogenesis.

Additional file 5: Supplementary Table 2. Antibodies used for immunocytochemistry.

Additional file 4 Supplementary Table 1. This iPSCs were negative for mycoplasma test.

Dravet syndrome is an epileptic encephalopathy largely due to haploinsufficiency of the voltage-gated sodium channel Nav1.1 that is expressed primarily in GABAergic neurons. In order to distinguish the different subtypes, we used gene editing to introduce tdTomato gene into the genome of iPSCs to label the GABAergic neurons in the differentiated neuronal networks. The gene-edited cell line demonstrates normal karyotype, expresses the main pluripotency markers, and shows the presence of differentiation into the three embryonic germ layers in teratomas.

BackgroundEsophageal squamous cell carcinoma (ESCC) is a leading cause of cancer death worldwide. Although the exposure of esophageal mucosa to heat stimuli has long been recognized as an important risk for the initiation and development of ESCC, its underlying mechanisms remain uncharacterized. MethodsWestern blotting and immunofluorescence were used to detect the expression and localization of transient receptor potential vanilloid receptor 2 (TRPV2) in the ESCC cells. The cancerous behaviors of the ESCC cells were evaluated by a single-cell culturing assay, a wound healing assay, a 3D culturing assay and a tube formation assay respectively. In vivo tumorigenicity and metastasis of the ESCC cells were examined via xenograft nude mouse models. Western blotting and IHC were performed to determine the expression profiles of TRPV2 in the ESCC patient tissues. TRPV2 knocked-out ESCC cell line was established using CRISPR-Cas9 gene editing technique.ResultsHere, we found that the expression of TRPV2, one of the thermally sensitive TRP family members, was upregulated in both ESCC cells and clinical samples. We further showed that activation of TRPV2 by recurrent acute thermal stress (54°C, a temperature unexpectedly much lower than those in many dietary modalities) or O1821 (20 μM), a TRPV2 agonist, promoted cancerous behaviors in ESCC cells. The proangiogenic capacity of the heat-challenged ESCC cells was also found to be enhanced profoundly in the tube formation assay; both tumor formation and metastasis that originated from the cells were substantially promoted in nude mouse models upon the activation of TRPV2. These effects were inhibited significantly by tranilast (120 μM), a TRPV2 inhibitor, and abolished by TRPV2 knock-out using CRISPR-Cas9 gene editing. Conversely, overexpression of TRPV2 by transfection of TRPV2 DNA into NE2 cells, could switch the cells to tumorigenesis upon activation of TRPV2. Mechanistically, the driving role of TRPV2 in the progression of ESCC is mainly regulated by the PI3K/Akt/mTOR signaling pathway. Application of a pan-PI3K/mTOR inhibitor and/or a PTEN activator resulted in markedly reduced ESCC cell proliferation. Conclusions Our study first proved that TRPV2 plays an important role in the tumorigenesis of ESCC upon thermal stress. We revealed that TRPV2-PI3K/Akt/mTOR is a novel and promising target for the prevention and treatment of ESCC.

BackgroundNeural stem cells（NSCs）therapy remains one of the most potential approaches for neurological disorders treatment. The discovery of human induced pluripotent stem cells (hiPSCs) and the establishment of hiPSC-derived human neural stem cells (hiNSCs) have revolutionized our technique to cell therapy. Meanwhile, it is often required that NSCs are stored and transported long distances for research or treatment. Although high survival rates could be maintained, conventional methods of cell transport (dry ice or liquid nitrogen) are inconvenient and expensive. Therefore，the establishment of a safe, affordable, and frequent obtained hiPSCs and hiNSCs, with characteristics that match fetal hNSCs and a simple, low-cost way to store and transport, are incredibly urgent. MethodsWe reprogrammed human urinary cells to iPSCs using a virus-free technique and differentiated the iPSCs toward iNSCs/neurospheres and neurons, under Good Manufacturing Practice (GMP)-compatible conditions. The pluripotency of iPSCs and iNSCs was characterized by a series of classical methods (surface markers, karyotype analysis and in vitro and in vivo differentiation capabilities, etc).ResultsHere, our results showed that we successfully generated hiNSCs/neurospheres from more available, non-invasive, and more acceptable urinary cells by a virus-free technique and their differentiation into neural networks. Moreover，hiNSCs survived longer as neurospheres at ambient temperature than those cultured in a monolayer. Approximately 7 days, the neural viability remained at > 80%, while hiNSCs cultured in a monolayer died almost immediately. Neurospheres exposed to ambient temperature that were placed under standard culture conditions (37 ℃, 5% CO2) recovered their typical morphology, and retained their ability to proliferate and differentiate. ConclusionsIn this study, we provided a simple method for the storage of NSCs as neurospheres at ambient temperature as an alternative to more costly and inconvenient traditional methods of cryopreservation. This will enable hiNSCs to be transported over long distances at ambient temperature and facilitate the therapeutic application of NSCs as neurospheres without any further treatment.

Background Neural stem cell (NSC) therapy remains one of the most potential approaches for the treatment of neurological disorders. The discovery of human induced pluripotent stem cells (hiPSCs) and the establishment of hiPSC-derived human neural stem cells (hiNSCs) have revolutionized the technique of cell therapy. Meanwhile, it is often required that NSCs are stored and transported to a long distance for research or treatment purposes. Although high survival rates could be maintained, conventional methods for cell transportation (dry ice or liquid nitrogen) are inconvenient and expensive. Therefore, the establishment of a safe, affordable, and low-cost strategy to store and transport easily accessible hiPSCs and hiNSCs, with characteristics that match fetal hNSCs, is incredibly urgent. Methods We reprogrammed human urinary cells to iPSCs using a non-integrating, virus-free technique and differentiated the iPSCs toward iNSCs/neurospheres and neurons, under Good Manufacturing Practice (GMP)-compatible conditions. The pluripotency of iPSCs and iNSCs was characterized by a series of classical methods (surface markers, karyotype analysis, and in vitro as well as in vivo differentiation capabilities, etc.). Results Here, our results showed that we successfully generated hiNSCs/neurospheres from more available, non-invasive, and more acceptable urinary cells by a virus-free technique. Next, we demonstrated that the iNSCs differentiated into mature cerebral cortical neurons and neural networks. Interestingly, hiNSCs survived longer as neurospheres at ambient temperature (AT) than those cultured in a monolayer. Within 7 days approximately, the neural viability remained at > 80%, while hiNSCs cultured in a monolayer died almost immediately. Neurospheres exposed to AT that were placed under standard culture conditions (37 °C, 5% CO2) recovered their typical morphology, and retained their proliferation and differentiation abilities. Conclusions In this study, we provided a simple method for the storage of NSCs as neurospheres at AT as an alternative method to more costly and inconvenient traditional methods of cryopreservation. This will enable hiNSCs to be transported over long distances at AT and facilitate the therapeutic application of NSCs as neurospheres without any further treatment.

Epilepsy is a neurological disorder, characterized by recurrent (two or more) epileptic seizures resulting from excessive and abnormal cortical neural activity.Fibroblasts were collected from a 10-year-old male with antecedent febrile seizures (PEFS+) and carrying a heterozygous A > G mutation of Nav1.1 α subunit gene. The induced USTCi001-A retained the mutation, expressed pluripotent markers, showed normal karyotype, and displayed in vitro differentiation potential toward cells of the three embryonic germ layers.

Platelet-derived growth factor BB (PDGF-BB) is a potential biomarker of tumor angiogenesis. For the first time, we developed a highly sensitive aptasensor for PDGF-BB with an enhanced test line signal by using two different gold nanoparticles (AuNPs). Herein, we describe a highly sensitive biosensor for PDGF-BB detection that combines biotinylated aptamer on a sample pad and poly thymine-Cy3-AuNP-monoclonal antibody complexes against PDGF-BB immobilized on conjugate pad A. Streptavidin (SA) and rabbit anti-mouse polyclonal antibody were also immobilized in the nitrocellulose membrane at the test and control zones, respectively. When the target PDGF-BB protein was added, it first bound the aptamer, and later the monoclonal antibody to form a biotinylated complex that was captured by SA, resulting in a visual red line on the test zone. In addition, to enhance the sensitivity, another monoclonal antibody against Cy3 was conjugated on AuNP B and immobilized on conjugate pad B to form a AuNPs (A&B)-antibody-(PDGF-BB-Cy3)-aptamer-biotin-SA complex on the test line when a loading buffer was subsequently added. This approach showed a linear response to PDGF-BB from 3 ng mL−1 to 300 ng mL−1 with a limit of detection as low as 1 ng mL−1 obtained in 10 minutes. Our biosensor displayed results through red lines readable by the naked eye. Interestingly, our approach has been successfully applied for real sample verification, proving its applicability for cancer monitoring and diagnosis.

With the strategy of constructing new power system with new energy as the main body put forward, the installed capacity of new energy in Hunan area will usher in explosive growth during the 14th five-year Plan period. The safe and stable operation of new energy stations to power grid will also be increasingly intensified. The paper briefly introduces the working principle, working mode and application of SVG(Static Var Generator) device supporting new energy stations. Based on relevant standards, a test scheme suitable for new energy stations in Hunan province is proposed. Taking the SVG device supporting a new energy station in Hunan province as an example, the grid-connection performance test is carried out to comprehensively and scientifically evaluate the network-related performance of SVG devices in the new energy station.

Autism spectrum disorder (ASD) is a neurological disorder with complex etiologies. In this study, urine cells were collected from a 16-year-old male with ASD and reprogrammed with the human SKOM transcription factors. The patient has a heterozygous C > T mutation of FCGR1B gene that was confirmed by sequencing analysis. The pluripotency was verified by gene expression and capacity of differentiation towards the three germ layers. This kind of iPSC will be valuable for further understanding the pathogenesis of ASD and help to develop drugs for treating ASD.

Vitamin D plays a major role in the regulation of calcium homeostasis and affects bone metabolism. There is currently limited detailed knowledge about the vitamin D endocrine system in human bone cells. Here, we investigated the direct effects of 1α, 25-dihydroxyvitamin D₃ (1α, 25-(OH)₂D₃ or 'VD3') on osteogenesis of human umbilical cord mesenchymal stem cells (HUCMSCs). We also studied the impact of VD3 on intracellular Ca

Piezoelectric cantilever is one of the simplest configurations for energy harvesting. Many reports have shown that the resonances frequency of the piezoelectric energy harvesting (PEH) can be essentially shifted by the proof masses to the lower working frequencies in practice, and its high electric capacity is required to decrease its output electric impedance to match the electric load resistant. However, there are few reports to investigate the mechanical vibration damping ratio of PEH affected by the proof masses and output electric loads. In this study, the mechanical damping ratio (ζm) of PEHBC with zero proof mass, and the electric damping ratio (ζe) related to the electric load, the mechanical vibration damping ratio with proof mass and the electric load are measured and discussed for the piezoelectric bimorph cantilever energy harvesting (PEHBC). The damping ratio was calculated by the time dependence of tip displacement or velocity amplitude of PEHBC using a laser vibrometer. A theoretical analys...

Primers used in real-time RT-PCR for genes of differentially accumulated proteins in winter shoots and spring shoots. (DOC 23Â kb)

Although tea leaves are rich in secondary metabolites, not enough transcriptomic information is available to aid understanding of the molecular mechanisms underlying plant growth, development, and secondary metabolite production. In this study, a total of 437,908 reads were generated from the tea leaf transcriptome using 454 sequencing. De novo assembly yielded 25,637 unigenes, 22,872 of which were annotated by BLAST searches against public databases. Most of these unigenes mapped to carbohydrate metabolism, energy metabolism and secondary metabolite biosynthetic pathways. Some abundant transcripts related to photomorphogenesis and development in plants, including ubiquitin/26S proteasome, lipid transfer protein, PPR-containing protein, small GTPase, expansin, transport inhibitor response 1 and thioredoxin, were identified in the transcriptome. Most of the genes encoding the main enzymes involved in flavonoid, caffeine and theanine biosynthesis were also found, and six MYB and two bHLH genes known to regulate flavonoid synthesis were identified. ABC transporter and glutathione S-transferase, generally responsible for secondary metabolite transport, and CYP450, broadly involved in oxidation steps in secondary metabolism, were also present in a large number of unigenes. Additionally, 3,767 EST-SSRs were identified as potential molecular markers in our unigenes. A total of 100 PCR primer pairs used in initial screening tests among 20 tea genotypes successfully identified 36 polymorphic loci. Overall, the tea leaf transcriptome sequences generated in this study reveal novel gene expression profiles and offer important clues for further study of the molecular mechanism of tea leaf growth, development and secondary metabolite synthesis. The thousands of EST-SSR markers identified will facilitate marker-assisted selection in tea breeding.

Electrostrictive materials exhibit high induced strain under dc electric fields and this effect is important in generator application. This paper studied the energy-harvesting performances of La-doped PMN-PT electrostrictive ceramics that have rarely been reported. A multilayer structured electrostrictive energy harvester with high dielectric constant and electrostrictive coefficient is proposed. With a strain Sm = 7 × 10−5 (m/m), a peak current value of 630 µA is obtained across a small matched resistance of 450 Ω under a dc electrical field of 1.5 V/µm. The working principle of the device is reviewed and improved to predict the current and power generated by the harvester. Experimental results are compared to the theoretic behaviors of the improved model. A good agreement is observed by the two sets of data. The study on the efficiency of the energy conversion showed that a positive balance between the consumed power and the harvested power was presented by the device. The result demonstrates the potential perspective of the electrostrictive device in energy harvesting applied to low-power electronics.

In this paper, a super version of the Hopf quiver theory is developed. The notion of Hopf superquivers is introduced. It is shown that only the path supercoalgebras of Hopf superquivers admit graded Hopf superalgebra structures. A complete classification of such graded Hopf superalgebras is given. A superquiver setting for general pointed Hopf superalgebras is also built up. In particular, a super version of the Gabriel type theorem and the Cartier-Gabriel decomposition theorem is given.

Summary In the present study, we employed high performance liquid chromatography with an amide-C16 column to determine the eighteen major active ingredients in black tea, including theanine, gallic acid, four purine alkaloids, eight catechins and four theaflavins. The method was successfully used to analyse two new kinds of black teas from the leaves of Camellia ptilophylla and Camellia kucha in China and several other world- famous black teas. Forty percentage ethanol was chosen as the extraction solvent for preparing tea extracts. All of the eighteen compounds could be separated within 86 min with a gradient elution system. Excellent linearity was observed for all the standard calibration curves, and correlation coefficients were above 0.9991. The developed method is accurate and sensitive enough for the determination of active components in black tea.

Camellia ptilophylla Chang is a wild tea tree containing theobromine and is caffeine-free. Lots of researches have been conducted for its domestication since it was discovered to contain theobromine in its shoots. First, its pharmacological and physiological effects have been studied, demonstrating that it can be used as a new resource of tea as daily and healthy beverage. Cocoa tea differs from traditional tea in that it does not excite the nervous system. Second, various ways of propagation have been investigated, and sexless cutting has been the method adopted currently. Third, through selection and breeding, plantation of cocoa tea can be set up to cultivate new varieties, and cocoa tea of different flavors such as green cocoa tea, oolong cocoa tea, and black cocoa tea can be processed. Thus, cocoa tea will become a choice in the tea market.