Determination of human COVID-19 total antibodies in serum using a time-resolved fluorescence immunoassay.

Coronavirus disease 2019 (COVID-19) caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is spreading rapidly around the world. Antibody detection plays an important role in the diagnosis of COVID-19. Here, we established a new time-resolved fluorescence immunoassay (TRFIA) to determine COVID-19 total antibodies. A double-antigen sandwich TRFIA was optimized and established: recombinant nucleocapsid phosphoprotein (N protein) and spike protein (S protein) of COVID-19 immobilized on 96-well plates captured human COVID-19 antibodies and then banded together with the N/S proteins labeled with europium(III) (Eu3+ ) chelates, and finally, time-resolved fluorometry was used to measure the fluorescence values. We successfully established a TRFIA method for the detection of human COVID-19 total antibodies, and the cutoff value was 2.02. There was no cross-reactivity with the negative reference of the National Reference Panel for IgM and IgG antibodies to COVID-19. The CV of the precision assay was 3.19%, and the assay could be stored stably for 15 days at 37°C. Compared with that of the colloidal gold method and chemiluminescence method, the sensitivity of the TRFIA method was higher, and the false positive/negative rate was lower. This established TRFIA has high sensitivity, accuracy, and specificity, which indicates that this method provides a new detection method for the high-throughput routine diagnosis of COVID-19.

Spatiotemporal variation of nitrogen and phosphorus and its main influencing factors in Huangshui River basin.

The foundation of managing excess nutrients in river is the identification of key physical processes and the control of decisive influencing factors. The existing studies seldom consider the influence of rainfall-runoff relationship and only focus on a few anthropogenic activities and natural attributes factors. To address this issue, a comprehensive set of influencing factors including rainfall-runoff relationship (represented by runoff coefficient), basic physical and chemical parameters of water quality, land use types, landscape patterns, topography, and socioeconomic development was constructed in this study. M-K test and cluster analysis were conducted to identify the temporal mutation and spatial clustering characteristics of NH3-N and TP in Huangshui River basin, respectively. Partial least squares regression was used to elucidate the linkages between water contaminants and the factors. As shown in the results, the temporal mutations of NH3-N and TP were obvious in the middle reaches, with 4 out of 7 catchments in the middle reaches have a larger number of mutations of NH3-N than other catchments. The cluster analysis results of NH3-N and TP among catchments were similar. This study also indicated that although the Huangshui River basin was located in the upper reaches of the Yellow River, the influences of rainfall-runoff relationship on spatiotemporal changes of NH3-N and TP in its sub-basins were limited. Only the temporal change of NH3-N in Jintan catchment in the upstream area was significantly affected by runoff coefficient. The indexes of proportion of water area (PWA), proportion of impervious area (PIA), and proportion of primary industry (PPI) were the top three influencing factors of temporal variation of NH3-N and TP for most catchments in the middle reaches. The temporal change of NH3-N in Jintan catchment in the upstream area was obviously affected by runoff coefficient. The spatial variation of NH3-N and TP were all affected by PWA and proportion of secondary industry significantly. The results of this study can provide theoretical basis and technical support for the control and management of nitrogen and phosphorus pollution in upper reaches of rivers.

Multifunctional Branched Nanostraw-Electroporation Platform for Intracellular Regulation and Monitoring of Circulating Tumor Cells.

Downstream analysis of circulating tumor cells (CTCs) has provided new insights into cancer research. In particular, the detection of CTCs, followed by the regulation and monitoring of their intracellular activities, can provide valuable information for comprehensively understanding cancer pathogenesis and progression. However, current CTC detection techniques are rarely capable of in situ regulation and monitoring of the intracellular microenvironments of cancer cells over time. Here, we developed a multifunctional branched nanostraw (BNS)-electroporation platform that could effectively capture CTCs and allow for downstream regulation and monitoring of their intracellular activities in a real-time and in situ manner. The BNSs possessed numerous nanobranches on the outer sidewall of hollow nanotubes, which could be conjugated with specific antibodies to facilitate the effective capture of CTCs. Nanoelectroporation could be applied through the BNSs to nondestructively porate the membranes of the captured cells at a low voltage, allowing the delivery of exogenous biomolecules into the cytosol and the extraction of cytosolic contents through the BNSs without affecting cell viability. The efficient delivery of biomolecules (e.g., small molecule dyes and DNA plasmids) into cancer cells with spatial and temporal control and, conversely, the repeated extraction of intracellular enzymes (e.g., caspase-3) for real-time monitoring were both demonstrated. This technology can provide new opportunities for the comprehensive understanding of cancer cell functions that will facilitate cancer diagnosis and treatment.

[Determination of six active ingredients in different parts of Belamcanda chinensis and Iris tectorum and their anti-inflammatory activity].

In order to explore the anti-inflammatory activity and active ingredient basis from the leaves of the Belamcanda chinensis and Iris tectorum, we established an HPLC method for simultaneous determination of six anti-inflammatory active ingredient contents in the root of the B. chinensis and I. tectorum as well as their leaves with different dry methods, and the anti-inflammatory effects of the extract were studied by the mouse ear swelling experiment. The HPLC analysis was performed on an Agilent WondaSil© C18-WR(4.6 mm×250 mm，5 µm)，with isocratic elution of acetonitrile-0.1% ortho-phosphoric acid solution at a flow rate of 1. 0 mL·min⁻¹ and the detection was carried out at 265 nm. The chemical compositions of the B. chinensis and I. tectorum are similar but the contents of them are obviously different. Both rhizome and leaf extract of B. chinensis and I. tectorum had inhibitory effects on inflamed mice induced by dimethylbenzene and had anti-inflammatory effects by animal experiment, which could lay the material and active foundation for the development of the non-medicinal parts of the B. chinensis and I. tectorum.

Decreased miR-144 expression as a non-invasive biomarker for acute myeloid leukemia patients.

MicroRNAs are found to be stable in blood and they demonstrated tissue specific expression patterns. Thus, they may be used as potential non-invasive biomarkers of specific cancers. In the current study, we mainly focused on miR-144, which has never been studied in acute myeloid leukemia (AML). The expression of miR-144 was explored in the bone marrow and peripheral blood of AML patients and healthy control. The correlation between peripheral blood miR-144 level and key clinical parameters, including overall survival and prognostic value, was further explored. We showed that miR-144 was markedly reduced in both the peripheral blood and bone marrow of AML patients compared with healthy controls. Further study revealed that there is a significant correlation between peripheral blood miR-144 level and FAB classification (p=0.0023) and cytogenetics (p=0.001). More importantly, a lower expression of peripheral blood miR-144 level was found to be positively correlated with poorer overall survival rate. In summary, peripheral blood miR-144 may be utilized as a potential novel non-invasive biomarker for AML screening.

[A study of immunocyte subsets and serum cytokine profiles before and after immunal suppression treatment in patients with immune thrombocytopenia].

OBJECTIVE: To explore the clinical significance of a series of cytokines and peripheral blood immunocyte subsets before and after immunosuppressive therapy in patients with immune thrombocytopenia (ITP). METHODS: The percentages of immunocyte subsets in the peripheral blood of 20 patients with ITP and 20 healthy controls were detected by flow cytometry, including CD3(+), CD4(+), CD8(+), CD4(+)/CD8(+), CD19(+). ELISA was applied to detect the level of serum TNFα, IL-2, IL-6, IL-4, IL-10, IL-11, IL-17, IL-27, transforming growth factor ß (TGFß), thrombopoietin (TPO) of 20 patients with ITP and 20 healthy controls. RESULTS: The percentage of CD3(+) T lymphocyte, CD4(+) T lymphocyte and the ratio of CD4(+)/CD8(+) T lymphocyte in patients with ITP were lower than those in healthy controls[(62.66 ± 6.58)% vs (69.93 ± 4.81)%, (29.46 ± 5.02)% vs (39.08 ± 3.50)%, 0.97 ± 0.35 vs 1.56 ± 0.26, all P<0.05]. After immunosuppressive therapy, the percentage of CD3(+) T lymphocyte, CD4(+) T lymphocyte and the ratio of CD4(+)/CD8(+) T lymphocyte [(71.49 ± 5.16)%, (39.25 ± 3.21)% and 1.56 ± 0.28] recovered to the same levels in healthy controls. The percentage of CD8(+) T lymphocyte and CD19(+) B lymphocyte in patients with ITP were higher than those in the healthy controls [(30.28 ± 4.63)% vs (25.90 ± 3.06)%, (18.92 ± 4.27)% vs (13.17 ± 3.64)%, all P<0.05]. After treatment of immunosuppressive therapy, the percentage of CD8(+) T lymphocyte and CD19(+) B lymphocyte [(25.16 ± 3.45)% and (11.98 ± 3.68)%] recovered to the similar levels in healthy controls. The serum levels of IL-4, IL-6, IL-11, IL-17 and TPO in patients with ITP were significantly higher than those in healthy controls. While TGFß level was significantly decreased. There was no significant difference of IL-27 between ITP patients and healthy controls. After the treatment of immunosuppressive therapy, IL-4, IL-6, IL-11, IL-17, TPO and TGFß were down-regulated while IL-27 was up-regulated. There was no significant difference of IFNγ, TNFα, IL-2 and IL-10 among ITP patients before or after immunosuppressive therapy and healthy controls. CONCLUSIONS: The present study suggests that the aberrant immunocyte subsets and cytokines are involved in the pathogenesis of ITP. Hyper-function of Th2 and Th17, dysfunction of Treg cells, up-regulation of IL-27, IL-11, TPO and other factors may contribute to the pathogenesis of ITP.

Drought-flood abrupt alteration events over China.

Drought-flood abrupt alternation (DFAA) is characterized by a period of persistent drought followed by sudden heavy precipitation at a certain level, with impacts on ecosystems and socioeconomic environment. At present, previous studies have mainly focuses on the monthly scale and regional scale. However, this study proposed a multi-indicator daily-scale method for identifying the DFAA occurrence, and explored the DFAA events over China from 1961 to 2018. The DFAA events mainly occurred in the center and southeast of China, especially in the Yangtze River Basin, Pearl River Basin, Huai River Basin, Southeast Rivers Basin, and south part of the Southwest Rivers Basin. The spatial coverage has a statistically significant (p < 0.05) increasing trend over China, of 0.355 %/decade. The occurrence and spatial coverage of DFAA events increased by decades, and were mainly concentrated in summer (around 85 %). The possible formation mechanisms were closely related to global warming, atmospheric circulation index anomalies, soil properties (e.g., soil field capacity), etc.

Continued decline of global soil moisture content, with obvious soil stratification and regional difference.

Soil is an important component connecting atmosphere and vegetation, and is an important 'regulator' of slope hydrological process. Global warming accelerates the global water cycle, and Soil Moisture Content (SMC) will change, but this change is not yet clear. Here, we study the global trend of SMC at different depths over the past 70 years and the next 70 years, based on the GLDAS-NOAH025 dataset and precipitation and temperature data from 15 CMIP6 models. We found that compared with the long-term average of 70 years, the global 0-200 cm SMC is decreasing at a rate of 1.284 kg/m2 per year from 2000 to 2020, and the area showing a significant decreasing trend accounts for 31.67 % of the global. Over the past decade, 0-200 cm SMC reduction rate (2.251 kg/m2) doubled. Global warming and precipitation reduction are the main reasons for the attenuation of SMC at different depths in the global from 2000 to 2020. Under the SSP126, SSP245, SSP370 and SSP585 scenarios, the global 0-200 cm SMC will continue to decay in the future, and the area showing a significant reduction trend accounts for 22.73-49.71 % of the global, but the stratified soil and regional differences are obvious. The attenuation of SMC will further aggravate the global water cycle and enhance the variability of extreme meteorological disasters. We will face more severe soil drought problems.

Lubricant-entrenched slippery surface-based nanocarriers to avoid macrophage uptake and improve drug utilization.

INTRODUCTION: Reducing the protein adsorption of nanoparticles (NPs) as drug carriers to slow their rapid clearance by macrophages uptake is a critical challenge for NPs clinical translational applications. Despite extensive research efforts to inhibit cellular uptake, including covering biological agents or surface chemical coatings to impart "stealth" properties to NPs, their stability remains insufficient. OBJECTIVES: Developed a novel surface modification technology based on a physical infusion engineering approach to achieve persistent inhibition of protein adhesion and cellular uptake by nanocarriers. METHODS: The nanoparticles were prepared based on conventional drug carrier mesoporous silica NPs through a two-step process. A functional nanoscale slippery surface was formed by grafting "liquid-like" brushes on the particles surface, and then a lubricant-entrenched slippery surfaces (LESS) was formed by infusing silicone oil lubricant into the entire surface. Co-incubation with macrophages (in vitro and in vivo) was used to examine the anti-uptake properties of modified NPs. The anti-adhesion properties of LESS coating surfaces to various liquids, proteins and cells were used to analyze the anti-uptake mechanism. Loaded with drugs, combined with tumor models, to evaluate the drug utilization of modified NPs. RESULTS: Relying on the stable and slippery LESS coating, the modified surface could prevent the adhesion of various liquids and effectively shield against the adhesion of proteins and cells, as well as remarkably reduce macrophage cellular uptake in vitro and in vivo. In addition, the LESS coating does not affect cell activity and allows NPs to be loaded with drugs, significantly improving the utilization of drugs in vitro and in vivo. This allows the NPs to reach to the target tumor site for drug delivery without active clearance by macrophages. CONCLUSION: Our research introduces a new nanocarrier technology to improve anti-biofouling performance and stealth efficiency that will facilitate the development of nanomedicines for clinical transformation applications.

Oligomannurarate sulfate sensitizes cancer cells to doxorubicin by inhibiting atypical activation of NF-κB via targeting of Mre11.

Aberrant regulation of nuclear factor kappa B (NF-κB) transcription factor is involved in cancer development, progression and resistance to chemotherapy. JG3, a marine-derived oligomannurarate sulfate, was reported as a heparanase and NF-κB inhibitor to significantly block tumor growth and metastasis in various animal models. However, the detailed functional mechanism remains unclear. Here, we report that JG3 inhibits NF-κB activation by specifically antagonizing the doxorubicin-triggered Ataxia-telangiectasia-mutated kinase (ATM) and the sequential MEK/ERK/p90Rsk/IKK signaling pathway but does not interfere with TNF-α-mediated NF-κB activation. This selective inactivation of the specific NF-κB cascade is attributed to the binding capacity of JG3 for Mre11, a major sensor of DNA double-strand breaks (DSB). Based on this selective mechanism, JG3 showed synergistic effect with doxorubicin in a panel of tumor cells and did not affect immune system function as shown in the in vivo delayed-type hypersensitivity (DTH) and hemolysis assays. All these highlight the clinical potential of JG3 as a favorable sensitizer in cancer therapy. In addition, identification of Mre11 as a potential target in the development of NF-κB inhibitors provides a platform for the further development of effective anticancer agents.

Newly discovered angiogenesis inhibitors and their mechanisms of action.

In the past decade, the success of angiogenesis inhibitors in clinical contexts has established the antiangiogenic strategy as an important part of cancer therapy. During that time period, we have discovered and reported 17 compounds that exert potent inhibition on angiogenesis. These compounds exhibit tremendous diversity in their sources, structures, targets and mechanisms. These studies have generated new models for further modification and optimization of inhibitory compounds, new information for mechanistic studies and a new drug candidate for clinical development. In particular, through studies on the antiangiogenic mechanism of pseudolaric acid B, we discovered a novel mechanism by which the stability of hypoxia-inducible factor 1α is regulated by the transcription factor c-Jun. We also completed a preclinical study of AL3810, a compound with the potential to circumvent tumor drug resistance to a certain extent. All of these findings will be briefly reviewed in this article.

[The role of CXCR3 and its ligand I-TAC in the pathogenesis of immune thrombocytopenic purpura].

OBJECTIVE: To investigate the roles of the chemokine receptor CXCR3 and its ligand I-TAC in the pathogenesis of immune thrombocytopenic purpura (ITP). METHODS: A total of 48 ITP patients were enrolled in this study: 30 with newly diagnosed or relapse ITP and 18 in remission after treatment, and 24 healthy volunteers were as controls. IFNγ and I-TAC in plasma were detected by ELISA. The mRNA expression of CXCR3 in the peripheral blood mononuclear cells (PBMNCs) was determined by quantitative RT-PCR. RESULTS: The IFNγ level in the plasma of ITP patients before the treatment was obviously increased than those in the remission group and controls [(71.45 ± 17.62) ng/L vs (36.94 ± 14.86) ng/L and (25.28 ± 12.85) ng/L, all P < 0.05] and those in the remission group was higher than in the controls (P < 0.05). In contrast, there were no statistic differences of the levels of I-TAC among the three groups [(455.56 ± 144.70) ng/L, (488.24 ± 164.70) ng/L and (382.97 ± 167.43) ng/L, P > 0.05]. Both ITP patients before the treatment and remission groups expressed more CXCR3 mRNA [6.76 (3.03, 37.00), 1.76 (0.45, 14.18) vs 0.12 (0.04, 0.28), P < 0.05]. After effective therapy, CXCR3 mRNA expression decreased, while it was still higher than that in the controls. CONCLUSIONS: Our data demonstrate that Th1 cytokine (IFNγ) dominance is reflected in ITP. Simultaneously, the CXCR3(+) cell may play a role in cell-mediated immunity through chemotaxis in ITP.

A data set of global river networks and corresponding water resources zones divisions v2.

The scale and topological relationship of river networks (RN) and water resources zones (WRZ) directly affect the simulation results of global multi-scale hydrological cycle and the accuracy of water resource refined evaluation. However, few existing global hydrological data sets take account of both aspects simultaneously. Here, we constructed a new hydrologic data set with a spatial resolution of 90 m as an upgraded version of the GRNWRZ V1.0. This data set had proper grading and partitioning thresholds and clear coding of topological relationships. Based on maintaining the accuracy of river networks in the GRNWRZ V1.0, we determined the more refined thresholds and created a new coding rule, which made the grading RN and partitioning WRZ more precise and the topological relationship more intuitive. Supported by this data set, the accuracy and efficiency of the large-scale hydrological simulation can be guaranteed. This data set provides fundamental data support for global water resources governance and global hydrological modeling under climate change.

Interleukin-27 enhances the production of tumour necrosis factor-α and interferon-γ by bone marrow T lymphocytes in aplastic anaemia.

Aplastic anaemia (AA) is considered as an immune-mediated bone marrow failure syndrome. The mechanism is involved with a variety of immune molecules including interferon-γ (IFN-γ), tumour necrosis factor-α (TNF-α) and interleukins (ILs). IL-27 is a novel member of the IL-12 family, which mediates T cell response and enhances the production of IFN-γ. However, little is known about the role of IL-27 in the development of AA. This study investigated the role of IL-27 and its receptor IL-27R in the pathogenesis of AA. Both the mRNA expression of IL-27/IL-27R subunits in the bone marrow mononuclear cells (BMMNCs) and the levels of IL-27 in the marrow plasma in AA were found to be higher than in controls. Increased IL-27 correlated with the disease severity of AA. Subsequently, we stimulated marrow T lymphocytes with recombinant human (rh)IL-27 and found that rhIL-27 enhanced the production of TNF-α and IFN-γ in both CD4(+) and CD8(+) T lymphocytes from AA patients. We also detected increased TNF-α and IFN-γ in the supernatants of BMMNCs from AA patients after IL-27 stimulation. In conclusion, our data suggest that elevated IL-27 and IL-27-induced TNF-α and IFN-γ overproduction might be involved in the pathogenesis of AA.

D11, a novel glycosylated diphyllin derivative, exhibits potent anticancer activity by targeting topoisomerase IIα.

Glycosylated natural products are reliable platforms for the development of anticancer drugs, simply due to the important features added by sugar appendages to the shape and the stereoelectronic properties of natural scaffolds. Herein, we indentified D11, a novel diphyllin glycoside with acetylated D-quinovose sugar moiety, as a potent topoisomerase IIα (Topo IIα) inhibitior. This peculiar sugar moiety endows D11 an optimal conformation with a high binding affinity for Topo IIα via hydrogen bonding to the entrance of ATPase pocket, thereby helping achieve more potent Topo II inhibition activity compared to the aglycon diphyllin. Further biochemical insights manifested that D11 significantly inhibited Topo IIα ATPase-catalyzed ATP hydrolysis in an ATP-dependent, but a DNA-independent manner. All these underlie the consequent superiority of D11 in the in vitro proliferation inhibition, apoptosis induction and the in vivo remarkable antitumor potency in xenograft mouse model. Moreover, D11 treatment also displayed no obvious body weight loss and other apparent toxicities, indicative of the restricted side effects by the virtue of sugar attachment. Taken together, a defined glycosylated manipulation of diphyllin may be a promising alternative approach in the development of novel Topo II inhibitors.

[The immunocyte subsets and their clinical significance in the peripheral blood of 35 patients with immune thrombocytopenic purpura].

OBJECTIVE: To explore the clinical significance of immunocyte subsets before and after immunosuppressive therapy in the peripheral blood of patients with immune thrombocytopenic purpura (ITP). METHODS: The percentages of immunocyte subsets in the peripheral blood of 35 patients with ITP and 20 healthy controls were detected by flow cytometry, including CD(3)(+), CD(4)(+), CD(8)(+), CD(+)(56), CD(19)(+) lymphocytes and CD(4)(+)/CD(8)(+). RESULTS: The percentages of CD(3)(+) T lymphocyte (61.58 ± 6.45)%, CD(4)(+) T lymphocyte (28.38 ± 4.89)% and the ratio of CD(4)(+)/CD(8)(+) 0.99 ± 0.22 in patients with ITP were lower than those in healthy controls [(67.85 ± 4.68)%, (38.00 ± 3.37)%, 1.54 ± 0.13, all P < 0.05]. After immunosuppressive therapy, the percentages of CD(3)(+)T lymphocyte (69.41 ± 5.03)%, CD(4)(+)T lymphocyte (38.17 ± 3.18)% and the ratio of CD(4)(+)/CD(8)(+) 1.60 ± 0.15 recovered to control levels. The percentages of CD(8)(+)T lymphocyte (29.20 ± 4.50)% and CD(19)(+)B lymphocyte (17.74 ± 4.14)% were higher than those in healthy controls [(24.82 ± 2.93)% and (12.09 ± 3.51)%, all P < 0.05]. After the immunosuppressive therapy, the percentages of CD(8)(+)T lymphocyte (24.06 ± 3.02)% and CD(19)(+)B lymphocyte (10.90 ± 3.55)% recovered to control levels. There were no significant difference of the percentage of CD(56)(+) lymphocyte among ITP patients (15.80 ± 2.85)%, ITP patients after immunosuppressive therapy (15.16 ± 2.77)% and healthy controls (16.36 ± 2.75)%. CONCLUSION: The aberrant immunocyte subsets are involved in the pathogenesis of ITP, and detection of immunocyte subsets might be helpful for the diagnosis and determination of therapeutic outcome of ITP.

Agapetesheana, a new species of A.ser.Longifiles (Ericaceae) from Yunnan, China.

Agapetesheana Y. H. Tong & J. D. Ya (Ericaceae), a new species from Lüchun Xian, Yunnan Province, China is described and illustrated. This new species is assigned to Agapetessect.Agapetesser.Longifiles Airy Shaw. It is closest to A.inopinata Airy Shaw and A.oblonga Craib, but differs in having bead-like tubers, leaf blade with a wholly serrulate margin, subulate and much longer calyx lobes, much larger corollas that are carmine, green at the apex and maroon on angles, and longer stamens without spurs on the back.

Increased accumulation of hypoxia-inducible factor-1α with reduced transcriptional activity mediates the antitumor effect of triptolide.

BACKGROUND: Hypoxia-inducible factor-1α (HIF-1α), a critical transcription factor to reduced O2 availability, has been demonstrated to be extensively involved in tumor survival, aggressive progression, drug resistance and angiogenesis. Thus it has been considered as a potential anticancer target. Triptolide is the main principle responsible for the biological activities of the Traditional Chinese Medicine tripterygium wilfordii Hook F. Triptolide possesses great chemotherapy potential for cancer with its broad-spectrum anticancer, antiangiogenesis, and drug-resistance circumvention activities. Numerous biological molecules inhibited by triptolide have been viewed as its possible targets. However, the anticancer action mechanisms of triptolide remains to be further investigated. Here we used human ovarian SKOV-3 cancer cells as a model to probe the effect of triptolide on HIF-1α. RESULTS: Triptolide was observed to inhibit the proliferation of SKOV-3 cells, and meanwhile, to enhance the accumulation of HIF-1α protein in SKOV-3, A549 and DU145 cells under different conditions. Triptolide did not change the kinetics or nuclear localization of HIF-1α protein or the 26 S proteasome activity in SKOV-3 cells. However, triptolide was found to increase the levels of HIF-1α mRNA. Unexpectedly, the HIF-1α protein induced by triptolide appeared to lose its transcriptional activity, as evidenced by the decreased mRNA levels of its target genes including VEGF, BNIP3 and CAIX. The results were further strengthened by the lowered secretion of VEGF protein, the reduced sprout outgrowth from the rat aorta rings and the inhibitory expression of the hypoxia responsive element-driven luciferase reporter gene. Moreover, the silencing of HIF-1α partially prevented the cytotoxicity and apoptosis triggered by triptolide. CONCLUSIONS: The potent induction of HIF-1α protein involved in its cytotoxicity, together with the suppression of HIF-1 transcriptional activity, indicates the great therapeutic potential of triptolide as an anticancer drug. Meanwhile, our data further stress the possibility that HIF-1α functions in an unresolved nature or condition.

Intracellular recording of cardiomyocyte action potentials by nanobranched microelectrode array.

Electrophysiological study that records the action potential of cardiomyocyte served as excellent tool to explore cardiology and neuroscience, disease investigation and pharmacological screening. Advances of micro/nanotechnologies promote the development of three-dimensional (3D) nanodevices to record high-quality intracellular recordings by various perforation approaches of cells, however, the complicated fabrication processes limited their large-scale manufacture. In this work, a unique nanobranched microelectrode array (NBMEA) platform is developed to achieve high-quality intracellular recording of cultured cardiomyocytes in a minimally invasive manner. The NBMEA is consisting of high aspect ratio conductive nanobranches fabricated on patterned microelectrodes combining hydrothermal growth and standard microfabrication. The 3D structure of nanobranches enables the electrode to form tight coupling with cardiomyocytes to achieve the low voltage cell electroporation and high-quality intracellular recording. The recorded intracellular action potentials of cardiomyocytes by NBMEA exhibited significant enhancement on amplitude (~5 mV), signal-to-noise ratio (SNR) (~67.47 dB), recording duration (up to 105 min), and recording yield (69.5 ± 17.8%). This NBMEA platform is a promising and powerful tool for electrophysiology that opens up new opportunities for high-quality and stable intracellular recording of cardiomyocytes.

Antibody-free isolation and regulation of adherent cancer cells via hybrid branched microtube-sandwiched hydrodynamic system.

The detection of circulating tumor cells (CTCs) has achieved promising progress for early diagnosis and disease analysis. Microfluidic chip techniques have recently promoted the technologies of CTC sorting and analysis, yet seldom can the microfluidic chips for CTC enrichment via antibody-free capture provide in situ regulation of both extracellular and intracellular activity, which would be advantageous for cell-based pharmaceutical therapeutics and screening. Herein, we have demonstrated a hybrid TiO2/ZnO branched microtube array (HBMTA)-sandwiched hydrodynamic device that integrates the multiple functions of selective enrichment of adherent tumor cells in an antibody-free manner and in situ delivery to the extracellular and intracellular spaces of the enriched tumor cells. More than 90% cancer cells were enriched on the device due to their preferential adhesion with the nano-branches of HBMTA, while more than 91% blood cells were eliminated from the device by constant hydrodynamic fluid shearing. For in situ regulation, temporally and spatially controlled extracellular delivery to the enriched tumor cells could be precisely achieved through the hollow structures of the HBMTA. In addition, reagents (e.g. propidium iodide) could be delivered into the intracellular spaces of enriched tumor cells by coupling an electric field to nondestructively perforate the cell membrane. Our study not only offers a promising and facile strategy for antibody-free isolation of tumor cells, but also provides unique opportunities to facilitate cancer research, including antitumor drug screening and personalized therapeutics.