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1.  A novel ethanol gas sensor based on TiO2/Ag0.35V2O5 branched nanoheterostructures 
Scientific Reports  2016;6:33092.
Much greater surface-to-volume ratio of hierarchical nanostructures renders them attract considerable interest as prototypical gas sensors. In this work, a novel resistive gas sensor based on TiO2/Ag0.35V2O5 branched nanoheterostructures is fabricated by a facile one-step synthetic process and the ethanol sensing performance of this device is characterized systematically, which shows faster response/recovery behavior, better selectivity, and higher sensitivity of about 9 times as compared to the pure TiO2 nanofibers. The enhanced sensitivity of the TiO2/Ag0.35V2O5 branched nanoheterostructures should be attributed to the extraordinary branched hierarchical structures and TiO2/Ag0.35V2O5 heterojunctions, which can eventually result in an obvious change of resistance upon ethanol exposure. This study not only indicates the gas sensing mechanism for performance enhancement of branched nanoheterostructures, but also proposes a rational approach to design nanostructure based chemical sensors with desirable performance.
doi:10.1038/srep33092
PMCID: PMC5018879  PMID: 27615429
2.  A comprehensive estimate of recent carbon sinks in China using both top-down and bottom-up approaches 
Scientific Reports  2016;6:22130.
Atmospheric inversions use measurements of atmospheric CO2 gradients to constrain regional surface fluxes. Current inversions indicate a net terrestrial CO2 sink in China between 0.16 and 0.35 PgC/yr. The uncertainty of these estimates is as large as the mean because the atmospheric network historically contained only one high altitude station in China. Here, we revisit the calculation of the terrestrial CO2 flux in China, excluding emissions from fossil fuel burning and cement production, by using two inversions with three new CO2 monitoring stations in China as well as aircraft observations over Asia. We estimate a net terrestrial CO2 uptake of 0.39–0.51 PgC/yr with a mean of 0.45 PgC/yr in 2006–2009. After considering the lateral transport of carbon in air and water and international trade, the annual mean carbon sink is adjusted to 0.35 PgC/yr. To evaluate this top-down estimate, we constructed an independent bottom-up estimate based on ecosystem data, and giving a net land sink of 0.33 PgC/yr. This demonstrates closure between the top-down and bottom-up estimates. Both top-down and bottom-up estimates give a higher carbon sink than previous estimates made for the 1980s and 1990s, suggesting a trend towards increased uptake by land ecosystems in China.
doi:10.1038/srep22130
PMCID: PMC4770414  PMID: 26924637
3.  Complement component C3a plays a critical role in endothelial activation and leukocyte recruitment into the brain 
Background
The complement system is becoming increasingly recognized as a key participant in many neurodegenerative diseases of the brain. Complement-deficient animals exhibit reduced neuroinflammation.
Methods
In the present study, we administered intracerebroventricularly lipopolysaccharide (LPS) to mimic local infection of the brain and investigated the role of key complement component C3 in brain vasculature endothelial activation and leukocyte recruitment. The degree of neutrophil infiltration was determined by esterase staining. Leukocyte-endothelial interactions were measured using intravital microscopy. Cerebral endothelial activation was evaluated using real-time PCR and Western blotting.
Results
Neutrophil infiltration into the brain cortex and hippocampus was significantly reduced in C3−/− mice and C3aR−/− mice but not in C6−/− mice. We detected markedly attenuated leukocyte-endothelial interactions in the brain microvasculature of C3−/− mice. Accordingly, in response to LPS administration, the brain microvasculature in these mice had decreased expression of P-selectin, E-selectin, intercellular cell adhesion molecule 1 (ICAM-1), and vascular cell adhesion molecule 1 (VCAM-1). Depletion of C3 from the circulation also caused reduction in VCAM-1 and E-selectin expression and leukocyte recruitment, suggesting that C3 in the circulation contributed to brain endothelial activation. Furthermore, C3−/− mice exhibited decreased leukocyte recruitment into the brain upon tumor necrosis factor-α (TNF-α) stimulation. C3a activated the phosphorylation of p38 mitogen-activated protein kinase (MAPK) and nuclear factor-κB (NF-κB) and induced the upregulation of VCAM-1 and ICAM-1 expression in murine primary cerebral endothelial cells in vitro.
Conclusions
Our study provides the first evidence that C3a plays a critical role in cerebral endothelial activation and leukocyte recruitment during inflammation in the brain.
Electronic supplementary material
The online version of this article (doi:10.1186/s12974-016-0485-y) contains supplementary material, which is available to authorized users.
doi:10.1186/s12974-016-0485-y
PMCID: PMC4731990  PMID: 26822321
Complement; CNS inflammation; Intravital microscopy; Adhesion molecule; Leukocyte recruitment
4.  14-3-3γ regulates cell viability and milk fat synthesis in lipopolysaccharide-induced dairy cow mammary epithelial cells 
Our previous study demonstrated that 14-3-3γ overexpression was able to inhibit the production of lipopolysaccharide (LPS)-induced cytokines in dairy cow mammary epithelial cells (DCMECs) by inhibiting the activation of nuclear factor-κB (NF-κB) signaling pathways. However, the association between 14-3-3γ overexpression and milk fat synthesis in LPS-induced DCMECs remains unclear. Therefore, the present study investigated the effect of 14-3-3γ on cell viability and milk fat synthesis in LPS-induced DCMECs. The results of the MTT assay and lactate dehydrogenase activity assay demonstrated that 14-3-3γ overexpression was able to attenuate LPS-induced cytotoxicity in DCMECs, and increase the viability of the cells. In addition, the results of reverse transcription-quantitative polymerase chain reaction suggested that mRNA expression levels of genes associated with milk fat synthesis, including sterol regulatory element binding protein (SREBP1), peroxisome proliferator-activated receptor-γ (PPARG), cluster of differentiation 36, acetyl-coA carboxylase (ACC), fatty acid synthase (FAS) and fatty acid binding protein-3, were significantly upregulated in cells overexpressing the 14-3-3γ protein. In addition, as compared with the LPS-treated group, the activities of FAS and ACC were significantly increased. Furthermore, western blotting demonstrated that 14-3-3γ overexpression enhanced the protein expression levels of phosphorylated SREBP1 and PPARG. These results suggested that high levels of 14-3-3γ protein were able to attenuate LPS-induced cell damage and promote milk fat synthesis in LPS-induced DCMECs by increasing the cell viability and upregulating the expression levels of transcription factors associated with milk fat synthesis.
doi:10.3892/etm.2016.3029
PMCID: PMC4812431  PMID: 27073437
14-3-3γ; dairy cow mammary epithelial cells; cell viability; milk fat synthesis; lipopolysaccharide
5.  Nano-TiO2 Is Not Phytotoxic As Revealed by the Oilseed Rape Growth and Photosynthetic Apparatus Ultra-Structural Response 
PLoS ONE  2015;10(12):e0143885.
Recently nano-materials are widely used but they have shown contrasting effects on human and plant life. Keeping in view the contrasting results, the present study has evaluated plant growth response, antioxidant system activity and photosynthetic apparatus physiological and ultrastructural changes in Brassica napus L. plants grown under a wide range (0, 500, 2500, 4000 mg/l) of nano-TiO2 in a pot experiment. Nano-TiO2 has significantly improved the morphological and physiological indices of oilseed rape plants under our experimental conditions. All the parameters i-e morphological (root length, plant height, fresh biomass), physiological (photosynthetic gas exchange, chlorophyll content, nitrate reductase activity) and antioxidant system (Superoxide dismutase, SOD; Guaiacol peroxidase, POD; Catalase, CAT) recorded have shown improvement in their performance by following nano-TiO2 dose-dependent manner. No significant chloroplast ultra-structural changes were observed. Transmission electron microscopic images have shown that intact & typical grana and stroma thylakoid membranes were in the chloroplast, which suggest that nano-TiO2 has not induced the stressful environment within chloroplast. Finally, it is suggested that, nano-TiO2 have growth promoting effect on oilseed rape plants.
doi:10.1371/journal.pone.0143885
PMCID: PMC4666630  PMID: 26624621
6.  Therapeutic window of globular adiponectin against cerebral ischemia in diabetic mice: the role of dynamic alteration of adiponectin/adiponectin receptor expression 
Scientific Reports  2015;5:17310.
Recent studies have demonstrated that adiponectin (APN) attenuates cerebral ischemic/reperfusion via globular adiponectin (gAD). However, the therapeutic role of gAD in cerebral ischemic injury in type 1 diabetes mellitus (T1DM) remains unclear. Our results showed that gAD improved neurological scores and reduced the infarct volumes in the 8-week T1DM (T1DM-8W) mice, but not in the 2-week T1DM (T1DM-2W) mice. Moreover, the ischemic penumbra APN levels increased and peaked in T1DM-2W mice, and reduced to normal in T1DM-8W mice, while the APN receptor 1 (AdipoR1) expression change was the opposite. Administration of rosiglitazone in T1DM-2W mice up-regulated the expression of AdipoR1 and restored the neuroprotection of gAD, while intracerebroventricular injection of AdipoR1 small interfering RNA (siRNA) in T1DM-8W mice reversed it. Furthermore, the expression of p-PERK, p-IRE1 and GRP78 were increased whereas the expressions of CHOP and cleaved caspase-12 as well as the number of apoptotic neurons were decreased after gAD treatment in T1DM-8W mice. These beneficial effects of gAD were reversed by pretreatment with AdipoR1 siRNA. These results demonstrated a dynamic dysfunction of APN/AdipoR1 accompanying T1DM progression. Interventions bolstering AdipoR1 expression during early stages and gAD supplementation during advanced stages may potentially reduce the cerebral ischemic injury in diabetic patients.
doi:10.1038/srep17310
PMCID: PMC4661424  PMID: 26611106
7.  Neuroimmune and Neuropathic Responses of Spinal Cord and Dorsal Root Ganglia in Middle Age 
PLoS ONE  2015;10(8):e0134394.
Prior studies of aging and neuropathic injury have focused on senescent animals compared to young adults, while changes in middle age, particularly in the dorsal root ganglia (DRG), have remained largely unexplored. 14 neuroimmune mRNA markers, previously associated with peripheral nerve injury, were measured in multiplex assays of lumbar spinal cord (LSC), and DRG from young and middle-aged (3, 17 month) naïve rats, or from rats subjected to chronic constriction injury (CCI) of the sciatic nerve (after 7 days), or from aged-matched sham controls. Results showed that CD2, CD3e, CD68, CD45, TNF-α, IL6, CCL2, ATF3 and TGFβ1 mRNA levels were substantially elevated in LSC from naïve middle-aged animals compared to young adults. Similarly, LSC samples from older sham animals showed increased levels of T-cell and microglial/macrophage markers. CCI induced further increases in CCL2, and IL6, and elevated ATF3 mRNA levels in LSC of young and middle-aged adults. Immunofluorescence images of dorsal horn microglia from middle-aged naïve or sham rats were typically hypertrophic with mostly thickened, de-ramified processes, similar to microglia following CCI. Unlike the spinal cord, marker expression profiles in naïve DRG were unchanged across age (except increased ATF3); whereas, levels of GFAP protein, localized to satellite glia, were highly elevated in middle age, but independent of nerve injury. Most neuroimmune markers were elevated in DRG following CCI in young adults, yet middle-aged animals showed little response to injury. No age-related changes in nociception (heat, cold, mechanical) were observed in naïve adults, or at days 3 or 7 post-CCI. The patterns of marker expression and microglial morphologies in healthy middle age are consistent with development of a para-inflammatory state involving microglial activation and T-cell marker elevation in the dorsal horn, and neuronal stress and satellite cell activation in the DRG. These changes, however, did not affect the establishment of neuropathic pain.
doi:10.1371/journal.pone.0134394
PMCID: PMC4524632  PMID: 26241743
9.  14-3-3γ Regulates Lipopolysaccharide-Induced Inflammatory Responses and Lactation in Dairy Cow Mammary Epithelial Cells by Inhibiting NF-κB and MAPKs and Up-Regulating mTOR Signaling 
As a protective factor for lipopolysaccharide (LPS)-induced injury, 14-3-3γ has been the subject of recent research. Nevertheless, whether 14-3-3γ can regulate lactation in dairy cow mammary epithelial cells (DCMECs) induced by LPS remains unknown. Here, the anti-inflammatory effect and lactation regulating ability of 14-3-3γ in LPS-induced DCMECs are investigated for the first time, and the molecular mechanisms responsible for their effects are explored. The results of qRT-PCR showed that 14-3-3γ overexpression significantly inhibited the mRNA expression of tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6), interleukin-1β (IL-1β) and inducible nitric oxide synthase (iNOS). Enzyme-linked immunosorbent assay (ELISA) analysis revealed that 14-3-3γ overexpression also suppressed the production of TNF-α and IL-6 in cell culture supernatants. Meanwhile, CASY-TT Analyser System showed that 14-3-3γ overexpression clearly increased the viability and proliferation of cells. The results of kit methods and western blot analysis showed that 14-3-3γ overexpression promoted the secretion of triglycerides and lactose and the synthesis of β-casein. Furthermore, the expression of genes relevant to nuclear factor-κB (NF-κB) and mitogen-activated protein kinase (MAPKs) and lactation-associated proteins were assessed by western blot, and the results suggested that 14-3-3γ overexpression inactivated the NF-κB and MAPK signaling pathways by down-regulating extracellular signal regulated protein kinase (ERK), p38 mitogen-activated protein kinase (p38MAPK) and inhibitor of NF-κB (IκB) phosphorylation levels, as well as by inhibiting NF-κB translocation. Meanwhile, 14-3-3γ overexpression enhanced the expression levels of β-casein, mammalian target of rapamycin (mTOR), ribosomal protein S6 kinase 1 (S6K1), serine/threonine protein kinase Akt 1 (AKT1), sterol regulatory element binding protein 1 (SREBP1) and peroxisome proliferator-activated receptor gamma (PPARγ). These results suggest that 14-3-3γ was able to attenuate the LPS-induced inflammatory responses and promote proliferation and lactation in LPS-induced DCMECs by inhibiting the activation of the NF-κB and MAPK signaling pathways and up-regulating mTOR signaling pathways to protect against LPS-induced injury.
doi:10.3390/ijms160716622
PMCID: PMC4519969  PMID: 26204835
14-3-3γ; inflammatory responses; dairy cow mammary epithelial cells; NF-κB; MAPKs; mTOR signaling pathway
11.  Detection and evolutionary analysis of picobirnaviruses in treated wastewater 
Microbial Biotechnology  2014;8(3):474-482.
Wastewater contains numerous viruses. In this study, picobirnaviruses (PBVs) were detected in the stream of a wastewater treatment plant in Changsha, Hunan province, China, and evolutionary analysis of the isolated PBVs was performed. The phylogenetic tree revealed that the PBVs were highly divergent and could be classified into six distinct groups according to their hosts. Among these groups, pairwise comparison of the six groups revealed that the nucleotide distance of group 4 (bootstrap value = 0.92; nucleotide identity = 94%) was the largest. Thus, group 4 might represent a new division of PBVs. Comprehensive analysis of the obtained PBV sequences to investigate their evolutionary history and phylodynamics revealed that group 5 (PBVs from monkey) exhibited maximum polymorphism (K = 30.582, S = 74, η = 98, Pa = 47) and lowest nucleotide substitutions per site per year (6.54E-3 subs per site per year), except group 4. Maximum clade credibility tree indicated that group 5 appeared earlier than the other groups. In conclusion, this study detected PBVs in treated wastewater in China, and identified a new PBV group. Furthermore, among these PBVs, group 5 was found to survive longer and present a balance between PBVs and their monkey host.
doi:10.1111/1751-7915.12239
PMCID: PMC4408179  PMID: 25546400
12.  Single Cell Assay for Molecular Diagnostics and Medicine: Monitoring Intracellular Concentrations of Macromolecules by Two-photon Fluorescence Lifetime Imaging 
Theranostics  2015;5(9):919-930.
Molecular organization of a cell is dynamically transformed along the course of cellular physiological processes, pathologic developments or derived from interactions with drugs. The capability to measure and monitor concentrations of macromolecules in a single cell would greatly enhance studies of cellular processes in heterogeneous populations. In this communication, we introduce and experimentally validate a bio-analytical single-cell assay, wherein the overall concentration of macromolecules is estimated in specific subcellular domains, such as structure-function compartments of the cell nucleus as well as in nucleoplasm. We describe quantitative mapping of local biomolecular concentrations, either intrinsic relating to the functional and physiological state of a cell, or altered by a therapeutic drug action, using two-photon excited fluorescence lifetime imaging (FLIM). The proposed assay utilizes a correlation between the fluorescence lifetime of fluorophore and the refractive index of its microenvironment varying due to changes in the concentrations of macromolecules, mainly proteins. Two-photon excitation in Near-Infra Red biological transparency window reduced the photo-toxicity in live cells, as compared with a conventional single-photon approach. Using this new assay, we estimated average concentrations of proteins in the compartments of nuclear speckles and in the nucleoplasm at ~150 mg/ml, and in the nucleolus at ~284 mg/ml. Furthermore, we show a profound influence of pharmaceutical inhibitors of RNA synthesis on intracellular protein density. The approach proposed here will significantly advance theranostics, and studies of drug-cell interactions at the single-cell level, aiding development of personal molecular medicine.
doi:10.7150/thno.11863
PMCID: PMC4493531  PMID: 26155309
Fluorescence lifetime imaging (FLIM); two-photon excited fluorescence; macromolecular crowding; protein concentration; nuclear organization; nucleoplasm; nucleolus; nuclear speckles.
13.  Ideotype Population Exploration: Growth, Photosynthesis, and Yield Components at Different Planting Densities in Winter Oilseed Rape (Brassica napus L.) 
PLoS ONE  2014;9(12):e114232.
Rapeseed is one of the most important edible oil crops in the world and the seed yield has lagged behind the increasing demand driven by population growth. Winter oilseed rape (Brassica napus L.) is widely cultivated with relatively low yield in China, so it is necessary to find the strategies to improve the expression of yield potential. Planting density has great effects on seed yield of crops. Hence, field experiments were conducted in Wuhan in the Yangtze River basin with one conventional variety (Zhongshuang 11, ZS11) and one hybrid variety (Huayouza 9, HYZ9) at five planting densities (27.0×104, 37.5×104, 48.0×104, 58.5×104, 69.0×104 plants ha–1) during 2010–2012 to investigate the yield components. The physiological traits for high-yield and normal-yield populations were measured during 2011–2013. Our results indicated that planting densities of 58.5×104 plants ha–1 in ZS11 and 48.0×104 plants ha–1 in HYZ9 have significantly higher yield compared with the density of 27.0×104 plants ha–1for both varieties. The ideal silique numbers for ZS11 and HYZ9 were ∼0.9×104 (n m–2) and ∼1×104 (n m-2), respectively, and ideal primary branches for ZS11 and HYZ9 were ∼250 (n m–2) and ∼300 (n m–2), respectively. The highest leaf area index (LAI) and silique wall area index (SAI) was ∼5.0 and 7.0, respectively. Moreover, higher leaf net photosynthetic rate (Pn) and water use efficiency (WUE) were observed in the high-yield populations. A significantly higher level of silique wall photosynthesis and rapid dry matter accumulation were supposed to result in the maximum seed yield. Our results suggest that increasing the planting density within certain range is a feasible approach for higher seed yield in winter rapeseed in China.
doi:10.1371/journal.pone.0114232
PMCID: PMC4269386  PMID: 25517990
14.  Plasmon resonance enhanced multicolour photodetection by graphene 
Nature communications  2011;2:579.
Graphene has the potential for high-speed, wide-band photodetection, but only with very low external quantum efficiency and no spectral selectivity. Here we report a dramatic enhancement of the overall quantum efficiency and spectral selectivity that enables multicolour photodetection, by coupling graphene with plasmonic nanostructures. We show that metallic plasmonic nanostructures can be integrated with graphene photodetectors to greatly enhance the photocurrent and external quantum efficiency by up to 1,500%. Plasmonic nanostructures of variable resonance frequencies selectively amplify the photoresponse of graphene to light of different wavelengths, enabling highly specific detection of multicolours. Being atomically thin, graphene photodetectors effectively exploit the local plasmonic enhancement effect to achieve a significant enhancement factor not normally possible with traditional planar semiconductor materials.
doi:10.1038/ncomms1589
PMCID: PMC4235953  PMID: 22146398
15.  A systematic study of atmospheric pressure chemical vapor deposition growth of large-area monolayer graphene† 
Journal of materials chemistry  2012;22(4):1498-1503.
Graphene has attracted considerable interest as a potential material for future electronics. Although mechanical peel is known to produce high quality graphene flakes, practical applications require continuous graphene layers over a large area. The catalyst-assisted chemical vapor deposition (CVD) is a promising synthetic method to deliver wafer-sized graphene. Here we present a systematic study on the nucleation and growth of crystallized graphene domains in an atmospheric pressure chemical vapor deposition (APCVD) process. Parametric studies show that the mean size of the graphene domains increases with increasing growth temperature and CH4 partial pressure, while the density of domains decreases with increasing growth temperature and is independent of the CH4 partial pressure. Our studies show that nucleation of graphene domains on copper substrate is highly dependent on the initial annealing temperature. A two-step synthetic process with higher initial annealing temperature but lower growth temperature is developed to reduce domain density and achieve high quality full-surface coverage of monolayer graphene films. Electrical transport measurements demonstrate that the resulting graphene exhibits a high carrier mobility of up to 3000 cm2 V−1 s−1 at room temperature.
doi:10.1039/C1JM14272K
PMCID: PMC4235958  PMID: 25414547
16.  Increasing Burn Severity in Mice is Associated with Delayed Mobilization of Circulating Angiogenic Cells 
Angiogenesis is an important component of wound healing. Mobilization of circulating angiogenic cells (CACs) has been observed in patients with cutaneous burn wounds, but a systematic exploration of the phenomenon in an animal model has not been carried out. Using a murine model, in which burn depth can be varied precisely, and a validated culture method for quantifying circulating CACs, we found that increasing burn depth resulted in a progressive delay in the time to mobilization of circulating CACs. This delay in CAC mobilization was associated with a delay in perfusion and vascularization of the burn wound tissue. Analysis of CACs in the peripheral blood of human burn patients, using the same culture assay, confirmed results previously obtained by flow cytometry, that CAC levels peak early after the burn wound, and point to the clinical relevance of findings from the murine model.
doi:10.1001/archsurg.2009.285
PMCID: PMC4209728  PMID: 20231626
17.  Impaired Angiogenesis and Mobilization of Circulating Angiogenic Cells in HIF-1α Heterozygous-Null Mice after Burn Wounding 
Hypoxia-inducible factor 1 (HIF-1) is a transcription factor that controls vascular responses to hypoxia and ischemia. In this study, mice that were heterozygous for a null allele at the locus encoding the HIF-1α subunit (HET mice) and their wild type (WT) littermates were subjected to thermal injury involving 10% of body surface area. HIF-1α protein levels were increased in burn wounds of WT but not of HET mice on day 2. Serum levels of stromal-derived factor 1α, which binds to CXCR4, were increased on day 2 in WT but not in HET mice. Circulating angiogenic cells were also increased on day 2 in WT but not in HET mice and included CXCR4+Sca1+ cells. Laser Doppler perfusion imaging demonstrated increased blood flow in burn wounds of WT but not HET mice on day 7. Immunohistochemistry on day 7 revealed a reduced number of CD31+ vessels at the healing margin of burn wounds in HET as compared to WT mice. Vessel maturation was also impaired in wounds of HET mice as determined by the number of α-smooth muscle actin-positive vessels on day 21. The remaining wound area on day 14 was significantly increased in HET mice compared to WT littermates. The percentage of healed wounds on day 14 was significantly decreased in HET mice. These data delineate a signaling pathway by which HIF-1 promotes angiogenesis during burn wound healing.
doi:10.1111/j.1524-475X.2010.00570.x
PMCID: PMC4206187  PMID: 20163569
18.  Endothelial Na+/H+ exchanger NHE1 participates in redox-sensitive leukocyte recruitment triggered by methylglyoxal 
Background
Excessive levels of methylglyoxal (MG) encountered in diabetes foster enhanced leukocyte-endothelial cell interactions, mechanisms of which are incompletely understood. MG genomically upregulates endothelial serum- and glucocorticoid-inducible kinase 1 (SGK1) which orchestrates leukocyte recruitment by regulating the activation and expression of transcription factors and adhesion molecules. SGK1 regulates a myriad of ion channels and carriers including the Na+/H+ exchanger NHE1. Here, we explored the effect of MG on SGK1-dependent NHE1 activation and the putative role of NHE1 activation in MG-induced leukocyte recruitment and microvascular hyperpermeability.
Methods
Using RT-PCR and immunoblotting, we analyzed NHE1 mRNA and protein levels in murine microvascular SVEC4-10EE2 endothelial cells (EE2 ECs). NHE1 phosphorylation was detected using a specific antibody against the 14-3-3 binding motif at phospho-Ser703. SGK in EE2 ECs was silenced using targeted siRNA. ROS production was determined using DCF-dependent fluorescence. Leukocyte recruitment and microvascular permeability in murine cremasteric microvasculature were measured using intravital microscopy. The expression of endothelial adhesion molecules was determined by immunoblotting and confocal imaging analysis.
Results
MG treatment significantly upregulated NHE1 mRNA and dose-dependently increased total- and phospho-NHE1. Treatment with SGK1 inhibitor GSK650394, antioxidant Tempol and silencing SGK all blunted MG-triggered phospho-NHE1 upregulation in EE2 ECs. NHE1 inhibitor cariporide attenuated MG-triggered ROS production, leukocyte adhesion and emigration and microvascular hyperpermeability, without affecting leukocyte rolling. Cariporide treatment did not alter MG-triggered upregulation of P- and E-selectins, but reduced endothelial ICAM-1 expression.
Conclusion
MG elicits SGK1-dependent activation of endothelial Na+/H+ exchanger NHE1 which participates in MG-induced ROS production, upregulation of endothelial ICAM-1, leukocyte recruitment and microvascular hyperpermeability. Pharmacological inhibition of NHE1 attenuates the proinflammatory effects of excessive MG and may, thus, be beneficial in diabetes-associated inflammation.
doi:10.1186/s12933-014-0134-7
PMCID: PMC4193979  PMID: 25270604
Methylglyoxal; NHE1; Oxidative stress; SGK1; Leukocyte recruitment
19.  Strengthening the Skin with Topical Delivery of Keratinocyte Growth Factor-1 Using a Novel DNA Plasmid 
Molecular Therapy  2014;22(4):752-761.
Fragile skin, susceptible to decubitus ulcers and incidental trauma, is a problem particularly for the elderly and for those with spinal cord injury. Here, we present a simple approach to strengthen the skin by the topical delivery of keratinocyte growth factor-1 (KGF-1) DNA. In initial feasibility studies with the novel minimalized, antibiotic-free DNA expression vector, NTC8385-VA1, the reporter genes luciferase and enhanced green fluorescent protein were delivered. Transfection was documented when luciferase expression significantly increased after transfection. Microscopic imaging of enhanced green fluorescent protein–transfected skin showed green fluorescence in hair follicles, hair shafts, and dermal and superficial epithelial cells. With KGF-1 transfection, KGF-1 mRNA level and protein production were documented with quantitative reverse transcriptase–polymerase chain reaction and immunohistochemistry, respectively. Epithelial thickness of the transfected skin in the KGF group was significantly increased compared with the control vector group (26 ± 2 versus 16 ± 4 µm) at 48 hours (P = 0.045). Dermal thickness tended to be increased in the KGF group (255 ± 36 versus 162 ± 16 µm) at 120 hours (P = 0.057). Biomechanical assessment showed that the KGF-1–treated skin was significantly stronger than control vector–transfected skin. These findings indicate that topically delivered KGF-1 DNA plasmid can increase epithelial thickness and strength, demonstrating the potential of this approach to restore compromised skin.
doi:10.1038/mt.2014.2
PMCID: PMC3982499  PMID: 24434934
20.  Electroacupuncture decreases cognitive impairment and promotes neurogenesis in the APP/PS1 transgenic mice 
Background
Alzheimer’s disease (AD) is a severe neurodegenerative disease for which there is currently no effective treatment. The purpose of this study was to investigate whether repeated electroacupuncture (EA) stimulation would improve cognitive function and the pathological features of AD in amyloid precursor protein (APP)/presenilin 1 (PS1) double transgenic mice.
Methods
Cognitive function of APP/PS1 double transgenic mice was assessed using the Morris water maze test before and after EA treatment. Levels of amyloid β-peptide (Aβ) deposits in the hippocampus and cortex were evaluated by immunofluorescence, western blot and enzyme-linked immunosorbent assay. Expression of brain-derived neurotrophic factor (BDNF) was also examined by immunofluorescence and western blot. The neurogenesis was labeled by the DNA marker bromodeoxyuridine.
Results
EA stimulation significantly ameliorated the learning and memory deficits of AD mice by shortening escape latency and increasing the time spent in the target zone during the probe test. Additionally, decreased Aβ deposits and increased BDNF expression and neurogenesis in the hippocampus and cortex of EA-treated AD mice were detected. The same change was detected in wild-type mice after EA treatment compared with wild-type mice without EA treatment.
Conclusions
Repeated EA stimulation may improve cognitive function, attenuate Aβ deposits, up-regulate the expression of BDNF and promote neurogenesis in the APP/PS1 double transgenic mice. This suggests that EA may be a promising treatment for AD.
doi:10.1186/1472-6882-14-37
PMCID: PMC3907495  PMID: 24447795
Alzheimer’s disease; Electroacupuncture; Neurogenesis; BDNF; Aβ deposits
21.  RECENT PROGRESS IN MULTIFOCAL MULTIPHOTON MICROSCOPY 
Journal of innovative optical health sciences  2012;5(3):10.1142/S1793545812500186.
Multifocal multiphoton microscopy (MMM) has recently become an important tool in biomedicine for performing three-dimensional fast fluorescence imaging. Using various beamsplitting techniques, MMM splits the near-infrared laser beam into multiple beamlets and produces a multifocal array on the sample for parallel multiphoton excitation and then records fluorescence signal from all foci simultaneously with an area array detector, which significantly improves the imaging speed of multiphoton microscopy and allows for high efficiency in use of the excitation light. In this paper, we discuss the features of several MMM setups using different beamsplitting devices, including a Nipkow spinning disk, a microlens array, a set of beamsplitting mirrors, or a diffractive optical element (DOE). In particular, we present our recent work on the development of an MMM using a spatial light modulator (SLM).
doi:10.1142/S1793545812500186
PMCID: PMC3868482  PMID: 24363782
Multifocal multiphoton microscopy (MMM); microlens array; beamsplitter; diffractive optical element (DOE); spatial light modulator (SLM)
22.  High Yield Chemical Vapor Deposition Growth of High Quality Large-Area AB Stacked Bilayer Graphene 
ACS nano  2012;6(9):8241-8249.
Bernal stacked (AB stacked) bilayer graphene is of significant interest for functional electronic and photonic devices due to the feasibility to continuously tune its band gap with a vertical electrical field. Mechanical exfoliation can be used to produce AB stacked bilayer graphene flakes but typically with the sizes limited to a few micrometers. Chemical vapor deposition (CVD) has been recently explored for the synthesis of bilayer graphene but usually with limited coverage and a mixture of AB and randomly stacked structures. Herein we report a rational approach to produce large-area high quality AB stacked bilayer graphene. We show that the self-limiting effect of graphene growth on Cu foil can be broken by using a high H2/CH4 ratio in a low pressure CVD process to enable the continued growth of bilayer graphene. A high temperature and low pressure nucleation step is found to be critical for the formation of bilayer graphene nuclei with high AB stacking ratio. A rational design of a two-step CVD process is developed for the growth of bilayer graphene with high AB stacking ratio (up to 90 %) and high coverage (up to 99 %). The electrical transport studies demonstrated that devices made of the as-grown bilayer graphene exhibit typical characteristics of AB stacked bilayer graphene with the highest carrier mobility exceeding 4,000 cm2/V·s at room temperature, comparable to that of the exfoliated bilayer graphene.
doi:10.1021/nn302918x
PMCID: PMC3493488  PMID: 22906199
bilayer graphene; band gap; AB stacking; chemical vapor deposition; copper foil
23.  Methylglyoxal modulates endothelial nitric oxide synthase-associated functions in EA.hy926 endothelial cells 
Background
Increased levels of the sugar metabolite methylglyoxal (MG) in vivo were shown to participate in the pathophysiology of vascular complications in diabetes. Alterations of endothelial nitric oxide synthase (eNOS) activity by hypophosphorylation of the enzyme and enhanced monomerization are found in the diabetic milieu, and the regulation of this still remains undefined. Using various pharmacological approaches, we elucidate putative mechanisms by which MG modulates eNOS-associated functions of MG-stimulated superoxide O2•- production, phosphorylation status and eNOS uncoupling in EA.hy926 human endothelial cells.
Methods
In cultured EA.hy926 endothelial cells, the effects of MG treatment, tetrahydrobiopterin (BH4; 100 μM) and sepiapterin (20 μM) supplementation, NOS inhibition by NG-nitro-L-arginine methyl ester (L-NAME; 50 μM), and inhibition of peroxynitrite (ONOO-) formation (300 μM Tempol plus 50 μM L-NAME) on eNOS dimer/monomer ratios, Ser-1177 eNOS phosphorylation and 3-nitrotyrosine (3NT) abundance were quantified using immunoblotting. O2•-–dependent fluorescence was determined using a commercially available kit and tissue biopterin levels were measured by fluorometric HPLC analysis.
Results
In EA.hy926 cells, MG treatment significantly enhanced O2•- generation and 3NT expression and reduced Ser-1177 eNOS phosphorylation, eNOS dimer/monomer ratio and cellular biopterin levels indicative of eNOS uncoupling. These effects were significantly mitigated by administration of BH4, sepiapterin and suppression of ONOO- formation. L-NAME treatment significantly blunted eNOS-derived O2•- generation but did not modify eNOS phosphorylation or monomerization.
Conclusion
MG triggers eNOS uncoupling and hypophosphorylation in EA.hy926 endothelial cells associated with O2•- generation and biopterin depletion. The observed effects of the glycolysis metabolite MG presumably account, at least in part, for endothelial dysfunction in diabetes.
doi:10.1186/1475-2840-12-134
PMCID: PMC4015749  PMID: 24050620
Methylglyoxal; eNOS uncoupling; Superoxide; Tyrosine nitration; Biopterins; eNOS phosphorylation
24.  Scalable fabrication of self-aligned graphene transistors and circuits on glass 
Nano Letters  2011;12(6):2653-2657.
High frequency graphene transistors with the intrinsic cut-off frequency up to 300 gigahertz (GHz) have been demonstrated for radio frequency (RF) applications. However, functional graphene RF circuits such as frequency doublers and mixers operating in the gigahertz range is yet to demonstrated. Here we report a scalable approach to fabricate self-aligned graphene transistors and circuits that can operate in gigahertz regime. The devices are fabricated through a self-aligned aligned process on glass substrate using chemical vapor deposition (CVD) grown graphene and a dielectrophoretic assembled nanowire gate array. The self-aligned process allows to achieving unprecedented performance in CVD graphene transistors with a highest transconductance of 0.36 mS/μm. With the minimization of parasitic capacitance on insulating substrate, the resulting graphene transistors exhibit a record high extrinsic cut-off frequency (> 50 GHz) achieved in graphene transistors to date. The excellent extrinsic cut-off frequency readily allows configuring the graphene transistors into frequency doubling or mixing circuits functioning in the 1–10 GHz regime, a significant advancement over previous report (~20 MHz). The studies open a pathway to scalable fabrication of high speed graphene transistors and functional circuits, and represent a significant step forward to graphene based radio frequency devices.
doi:10.1021/nl201922c
PMCID: PMC3269556  PMID: 21648419
graphene transistors; self-aligned gate; cut-off frequency; RF mixers
25.  Domain Wall Motion in Synthetic Co2Si Nanowires 
Nano letters  2012;12(4):1972-1976.
We report the synthesis of single crystalline Co2Si nanowires, and electrical transport studies of single Co2Si nanowire devices at low temperature. The butterfly-shaped magnetoresistance shows interesting ferromagnetic features including negative magnetoresistance, hysteretic switch fields and step-wise drops in magnetoresistance. The non-smooth step-wise magnetoresistance response is attributed to magnetic domain wall pinning and de-pinning motion in the Co2Si nanowires probably at crystal defects or morphology defects. The temperature dependence of the domain wall de-pinning field is observed and is described by a model based on thermally assisted domain wall de-pinning over a single energy barrier.
doi:10.1021/nl204510p
PMCID: PMC3493485  PMID: 22469009
nanowires; magnetoresistance; domain wall; de-pinning field

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