TOPK Inhibition Enhances the Sensitivity of Colorectal Cancer Cells to Radiotherapy by Reducing the DNA Damage Response.

OBJECTIVE: Abnormal expression of T-lymphokine-activated killer cell-originated protein kinase (TOPK) was reported to be closely related to the resistance of prostate cancer to radiotherapy and to targeted drug resistance in lung cancer. However, the role of TOPK inhibition in enhancing radiosensitivity of colorectal cancer (CRC) cells is unclear. This study aimed to evaluate the radiosensitization of TOPK knockdown in CRC cells. METHODS: The expression of TOPK was detected in CRC tissues by immunohistochemistry, and the effect of TOPK knockdown was detected in CRC cells by Western blotting. CCK-8 and clonogenic assays were used to detect the growth and clonogenic ability of CRC cells after TOPK knockdown combined with radiotherapy in CRC cells. Furthermore, proteomic analysis showed that the phosphorylation of TOPK downstream proteins changed after radiotherapy. DNA damage was detected by the comet assay. Changes in the DNA damage response signaling pathway were analyzed by Western blotting, and apoptosis was detected by flow cytometry. RESULTS: The expression of TOPK was significantly greater in CRC tissues at grades 2-4 than in those at grade 1. After irradiation, CRC cells with genetically silenced TOPK had shorter comet tails and reduced expression levels of DNA damage response-associated proteins, including phospho-cyclin-dependent kinase 1 (p-CDK1), phospho-ataxia telangiectasia-mutated (p-ATM), poly ADP-ribose polymerase (PARP), and meiotic recombination 11 homolog 1 (MRE11). CONCLUSIONS: TOPK was overexpressed in patients with moderately to poorly differentiated CRC. Moreover, TOPK knockdown significantly enhanced the radiosensitivity of CRC cells by reducing the DNA damage response.

MARK3 kinase: Regulation and physiologic roles.

Microtubule affinity-regulating kinase 3 (MARK3), a member of the MARK family, regulates several essential pathways, including the cell cycle, ciliated cell differentiation, and osteoclast differentiation. It is important to understand the control of their activities as MARK3 contains an N-terminal serine/threonine kinase domain, ubiquitin-associated domain, and C-terminal kinase-associated domain, which perform multiple regulatory functions. These functions include post-translational modification (e.g., phosphorylation) and interaction with scaffolding and other proteins. Differences in the amino acid sequence and domain position result in different three-dimensional protein structures and affect the function of MARK3, which distinguish it from the other MARK family members. Recent data indicate a potential role of MARK3 in several pathological conditions, including congenital blepharophimosis syndrome, osteoporosis, and tumorigenesis. The present review focuses on the physiological and pathological role of MARK3, its regulation, and recent developments in the small molecule inhibitors of the MARK3 signalling cascade.

Inhibiting ALK-TOPK signaling pathway promotes cell apoptosis of ALK-positive NSCLC.

T-LAK cell-oriented protein kinase (TOPK) is a potential therapeutic target in tumors. However, its role in anaplastic lymphoma kinase (ALK)-positive non-small cell lung cancer (NSCLC) has not been reported. Here, we found that TOPK was highly expressed in ALK-positive NSCLC. Additionally, ALK was identified as another upstream kinase of TOPK by in vitro kinase assay screening. Then, it was proven that ALK phosphorylated TOPK at Y74 in vitro and ex vivo, and the pathways downstream of ALK-TOPK were explored by phosphoproteomic analysis. Subsequently, we demonstrated that inhibiting TOPK enhanced tumor sensitivity to alectinib (an ALK inhibitor). The combination of alectinib and HI-032 (a TOPK inhibitor) suppressed the growth and promoted the apoptosis of ALK-positive NSCLC cells ex vivo and in vivo. Our findings reveal a novel ALK-TOPK signaling pathway in ALK-positive NSCLC. The combination of alectinib and HI-032 might be a promising therapeutic strategy for improving the sensitivity of ALK-positive NSCLC to targeted therapy.

Effects of straw mulching and nitrogen application rates on crop yields, fertilizer use efficiency, and greenhouse gas emissions of summer maize.

Although straw mulching and nitrogen applications are extensively practiced in the agriculture sector, large uncertainties remain about their impacts on crop yields and especially the environment. The responses of summer maize yields, fertilizer use efficiency, and greenhouse gas (GHG) emissions including carbon dioxide (CO2), methane (CH4), and nitrous oxide (N2O) in the North China Plain (NCP) to two straw management practices (S0: no straw and S1: straw mulching) and two nitrogen application rates (N1: 180 and N2: 210 kg N ha-1) were investigated in field tests in 2018, 2019, and 2020. The highest yields and partial factor productivity (PFP) were obtained by S1N1, followed by S1N2, S0N1, and S0N2. S1N2 had the highest CO2 emissions and greatest CH4 uptake, S0N1 had the lowest CO2 emissions, and S0N2 had the smallest CH4 uptake. The highest and lowest N2O emissions were found in S0N1 and S1N1, respectively. The S1N2 treatment, an extensively applied practice, had the greatest global warming potential (GWP), which was 70.3 % larger than S1N1 and two times more than S0N1 and S0N2. The largest GHG emission intensity (GHGI) of 19.4 was found in the S1N2 treatment, while the other three treatments, S0N1, S0N2, and S1N1, had a GHGI of 10.1, 10.7, and 10.7, respectively according to three tested results. In conclusion, S1N1 treatment achieved a better trade-off between crop yields and GHG emissions of summer maize in NCP.

Effects of no-tillage on greenhouse gas emissions in maize fields in a semi-humid temperate climate region.

Agricultural tillage practices have a significant impact on the generation and consumption of greenhouse gases (GHGs), the primary causes of global warming. Two tillage systems, conventional tillage (CT) and no-tillage (NT), were compared to evaluate their effects on GHG emissions in this study. Averaged from 2018 to 2020, significant decreases of CO2 and N2O emissions by 7.4% and 51.1% were observed in NT as compared to those of CT. NT was also found to inhibit the soil CH4 uptake. In this study, soil was a source of CO2 and N2O but a sink for CH4. The effect of soil temperature on the fluxes of CO2 was more pronounced than that of soil moisture. However, soil temperature and soil moisture had a weak correlation with CH4 and N2O flux variations. As compared to CT, NT did not affect maize yields but significantly reduced global warming potential (GWP) by 8.07%. For yield-scaled GWP, no significant difference was observed in NT (9.63) and CT (10.71). Taken together, NT was an environment-friendly tillage practice to mitigate GHG emissions in the soil under the tested conditions.

A Self-Detecting and Self-Cleaning Biomimetic Porous Metal-Based Hydrogel for Oil/Water Separation.

Porous materials with super-wetting surfaces (superhydrophilic/underwater superoleophobic) are ideal for oil/water separation. However, the inability to monitor the pollution degree and self-cleaning during the separation process limits their application in industrial production. In this study, a porous metal-based hydrogel is proposed, inspired by the porous structure of wood. Porous copper foam with nano-Cu(OH)2 is used as the skeleton, and its surface is coated with a polyvinyl alcohol, tannic acid, and multiwalled carbon nanotube cross-linked hydrogel coating. The hydrogel has superhydrophilicity and excellent oil/water separation efficiency (>99%) and can adapt to various environments. This approach can also realize hydrogel pollution degree self-detection according to the change in the electrical signal generated during the oil/water separation process, and the hydrogel can also be recovered by soaking to realize self-cleaning. This study will provide new insights into the application of oil/water separation materials in practical industrial manufacturing.

Convergent Adaptation of Ootheca Formation as a Reproductive Strategy in Polyneoptera.

Insects have evolved numerous adaptations and colonized diverse terrestrial environments. Several polyneopterans, including dictyopterans (cockroaches and mantids) and locusts, have developed oothecae, but little is known about the molecular mechanism, physiological function, and evolutionary significance of ootheca formation. Here, we demonstrate that the cockroach asymmetric colleterial glands produce vitellogenins, proline-rich protein, and glycine-rich protein as major ootheca structural proteins (OSPs) that undergo sclerotization and melanization for ootheca formation through the cooperative protocatechuic acid pathway and dopachrome and dopaminechrome subpathway. Functionally, OSP sclerotization and melanization prevent eggs from losing water at warm and dry conditions, and thus effectively maintain embryo viability. Dictyopterans and locusts convergently evolved vitellogenins, apolipoprotein D, and laminins as OSPs, whereas within Dictyoptera, cockroaches and mantids independently developed glycine-rich protein and fibroins as OSPs. Highlighting the ecological-evolutionary importance, convergent ootheca formation represents a successful reproductive strategy in Polyneoptera that promoted the radiation and establishment of cockroaches, mantids, and locusts.

Astragalus Polysaccharides Attenuate Ovalbumin-Induced Allergic Rhinitis in Rats by Inhibiting NLRP3 Inflammasome Activation and NOD2-Mediated NF-κB Activation.

Allergic rhinitis (AR) is an IgE-mediated chronic inflammatory disease of the allergic nasal mucosa. It has a significant effect on quality life; most patients with AR also suffer from sleep disorders, mood disorders, and deterioration in social relationships. As increasing numbers of medicinal plants show productive anti-inflammatory activity against inflammatory diseases, there is growing interest in natural medicinal plant ingredients. To this end, we selected Astragalus polysaccharides (APS) to evaluate its anti-inflammatory effect on ovalbumin-induced AR rats, and we further explored its impact on NLRP3 inflammasome activation and NOD2-mediated NF-κB activation. We found that APS can alleviate the nasal symptom of AR rats and attenuate pathological alterations. APS also reduced the inflammatory cytokine levels. APS not only inhibited the NLRP3 inflammasome activation but also inhibited NF-κB activation by decreasing NOD2 expression and blocking the phosphorylation of NF-κB (p65). In conclusion, APS can effectively improve the inflammatory symptoms of nasal mucosa in AR rats, which may be mediated by the inhibition of NLRP3 inflammasome activation and NOD2-mediated NF-κB activation. These findings indicate that APS has the potential to be used as a therapeutic agent for AR.

Simulation Study on the Dynamic Behaviors of Water-in-Oil Emulsified Droplets on Coalescing Fibers.

Although increasing superwetting membranes have been developed for separating oil-water emulsions based on the "size-sieving" mechanism, their pores are easily blocked and fouled by the intercepted emulsified droplets, which would result in a severe membrane fouling issue and a sharp decline in flux. Instead of droplet interception, a fiber-based coalescer separates oil/water emulsions by inducing the emulsified droplets to coalesce and transform into layered oil/water mixtures, exhibiting an ability to work continuously for a long time with high throughput, which makes it a promising technology for emulsion treatment. However, the underlying mechanism of the separation process is not well understood, which makes it difficult to further improve the separation performance. Hence, in this work, the dynamic behaviors of water-in-oil emulsified droplets on the surface of the coalescing fiber were numerically investigated based on the phase-field model. The attachment, transport, and detachment behaviors of droplets on fibers were directly observed, and the effects of fiber wettability, orientation, arrangement, and fluid speed were studied in detail. First, it was observed that the droplets will move downstream along the fiber surface under the effect of fluid shear, and the large droplets tend to coalesce with their downstream small droplets on the same fiber surface because they move faster compared to the small droplets. Second, it was found that the emulsified droplet will spontaneously transport to the intersection of two angled fibers under the drive of asymmetric Laplace pressure, which demonstrated that the emulsified droplets tend to gather at the intersection of fibers when permeating through a coalescing medium. Third, it was found that the detachment behaviors of droplets from the fiber surface are strongly affected by their size, fiber wettability, and fluid velocity. In addition, the results of our simulation show that the backside of two closely attached fibers can further inhibit the detachment of droplets. We truly believe that our research results are of significance to optimize the parameters of a fiber-based coalescer for separating oil-water emulsions and to develop novel oil/water separators.

Under-oil superhydrophilic salt particle filter for the efficient separation of water-in-oil emulsions.

In this study, a surfactant stabilized water-in-oil emulsion has been successfully separated by using only NaCl particles as a filter. This novel strategy is suitable for continuous filtration of a large quantity of water-in-oil emulsion with a volume of up to 1500 mL. Moreover, a filtration flux of up to 40 000 L m-2 h-1 is reported, which is around ten times higher than the conventional filtration methods.

Supertough and Transparent Poly(lactic acid) Nanostructure Blends with Minimal Stiffness Loss.

This contribution is an attempt to explore the effectiveness of a series of newly obtained thermoplastic elastomers (TPEs) as a toughening agent for modifying poly(lactic acid) (PLA). The TPEs, including ionically modified isotactic polypropylene-graft-PLA (iPP-g-PLA) copolymers with explicit graft length, graft density, and ionic group content, and an iPP-g-PLA copolymer with a very high molecular weight and explicit graft density, were elaborately designed and synthesized. The semicrystal or rubbery copolymer backbone originated from iPP was designed to improve the toughness and maintain a relatively high strength, while the grafted PLA side chain was to ensure a high level of compatibility with the PLA matrix. To obtain further enhancement in interfacial reinforcement, the imidazolium-based ionic group was also added during graft onto reaction. All of these graft copolymers were identified with randomly distributed PLA branches, bearing a very high molecular weight ((33-398) × 104) and very high PLA content (57.3-89.3 wt %). Unprecedentedly, with a very small amount of newly designed TPE, the modified PLA blends exhibited a significantly increased elongation at break (up to about 190%) and simultaneously retained the very high stiffness and excellent transparency. The nanometer-scale phase-separated particles with good compatibility and refractive index matching to the PLA matrix were demonstrated to play a crucial role in the excellent performance. The findings suggested that the newly designed iPP-g-PLA copolymers are very economic, promising, and effective modifying agents for developing highly transparent and tough PLA-based sustainable materials.

The Influence of Blood Collection Tubes in Biomarkers' Screening by Mass Spectrometry.

PURPOSE: Mass spectrometry is one of the rapidly developing bio-analytical techniques in recent years, and it shows that the results of biomarkers' screening can be influenced by pre-analytical process. The selection of the blood collection tubes is one of the most significant steps of pre-analytical process which is often neglected by researchers. So, it is urgent to define the influence of blood collection tubes clearly in biomarkers' screening. EXPERIMENTAL DESIGN: Two types of blood collection tubes, non-additive tubes and coagulant activator tubes, are used to collect serum samples from patients and healthy controls. All samples are analyzed using matrix-assisted laser desorption ionization-time of flight mass spectrum in this study. RESULTS: The serum protein profile changes while using coagulant tubes whether for patients or healthy controls. It is found that the effect of coagulant on serum protein of patients is smaller than that of control group. There are 27 significantly different peaks between the control group and the control coagulant group. However, between patient group and patient coagulant group, only one differential peak is obtained. Coagulant changes the expression differences between patients and healthy controls, making the differences expand, shrink or reverse, and most of the polypeptides are small molecule, which will change the results of biomarker's screening. CONCLUSIONS AND CLINICAL RELEVANCE: This research suggested that different types of blood collection tubes would influence the final laboratory results. So it's important for clinicians to choose the proper tubes to detect biomarkers and make correct diagnoses.

Co-delivery of paclitaxel and tanespimycin in lipid nanoparticles enhanced anti-gastric-tumor effect in vitro and in vivo.

Combined administration regimens are commonly used in cancer therapy to reduce cell toxicity and drug resistance. In this study, we use solid lipid nanoparticles (SLNs) as drug carriers and sought to investigate the effect of combined administration of paclitaxel (PTX) and tanespimycin (17-AAG) in gastric cancer. The SLNs loaded with paclitaxel and tanespimycin were prepared using the solvent injection method. The effect of encapsulated SLNs on cell viability and colony formation were measured in three human gastric cell lines. Cell apoptosis assay was carried out in MKN45 cells and xenograft model was used to investigate the effect of encapsulated SLNs in vitro and in vivo. The expression levels of proteins involved in oxidative stress and apoptosis were measured by western blotting analysis. The encapsulated SLNs reduced cell viabilities and colony formation in gastric cell lines. These SLNs could also induce apoptosis in MKN45 cells, inhibit growth of xenograft and influence the protein levels of Hsp90, MnSOD, Cleaved caspase 3 and Cleaved PARP. The effect of encapsulated SLNs exceeded that of single treatment of PTX or 17-AAG. The combination administration of PTX or 17-AAG resulted in a synergetic anti-cancer effect, probably via an increased oxidative stress and apoptosis levels.

ICPMS-Based Specific Quantification of Phosphotyrosine: A Gallium-Tagging and Tyrosine-Phosphatase Mediated Strategy.

Low-abundance tyrosine phosphorylation is crucial to not only normal but also aberrant life processes. We designed and synthesized a photocleavable magnetic nanoparticle-based gallium tag for tagging and enrichment as well as UV-release of the phosphate-bearing molecules/ions in cells. HPLC/71Ga species-unspecific isotope dilution (71Ga-SUID) ICPMS was subsequently developed for specific and absolute quantification of phosphotyrosine (pY) under the assistance of a protein tyrosine phosphatase-1B (PTP-1B). pY quantification was thus achieved via determination of Ga in the Ga-phosphate complexes that come exclusively from the Ga-tagged pY. In this way, the method detection limit of pY reached down to 30 amol with the RSD lower than 5.70% (n = 5 at pmol level). Feasibility of this proposed method was validated using VNQIGTLSEpYIK, VNQIGTLpSEpYIK, and extracellular regulated protein kinase 1 peptide (-pTEpY-) standards with the recovery of more than 96% (n = 5). It was applied to the absolute quantification of pY in human breast cancer MCF-7 cells, indicating that pY increased by 1.60 nmol (61.1%) in 3.0 × 106 MCF-7 cells after 100 nM insulin stimulation. We believe that, not limited to pY quantification, this element-tagging and protease-specific reaction mediated ICPMS methodology will pave a simple path for ever more applications of ICPMS to the studies of quantitative protein post-translational modifications (PTMs) when suitable element-tags are designed and specific proteases are available toward targeted PTMs.

Hippocampal cAMP/PKA/CREB is required for neuroprotective effect of acupuncture.

Acupuncture has beneficial effects in vascular dementia (VaD) patients. The underlying mechanism, however, remains unknown. The present study was designed to investigate whether the cAMP/PKA/CREB cascade is involved in the mechanism of acupuncture in cerebral multi-infarction rats. In this study, cerebral multi-infarction was modeled in adult Wistar rats by homologous blood clot emboli. After a two-week acupuncture treatment at Zusanli (ST36), hippocampal-dependent memory was tested by employing a radial arm maze test. The hippocampus was isolated for analyses of cAMP concentration, phosphodiesterase (PDE) activity and CREB/pCREB and ERK/pERK expressions. The Morris water maze (MWM) task and CREB phosphorylation were evaluated in the presence of PKA-selective peptide inhibitor (H89). The radial arm maze test results demonstrated that acupuncture treatment at ST36 reversed hippocampal-dependent memory in impaired animals. Compared to those of the impaired group, cAMP concentration, PKA activity and pCREB and pERK expressions were increased following acupuncture therapy. Finally, the blockade of PKA reversed the increase in CREB phosphorylation and the improvement in recognitive function induced by acupuncture treatment. These results suggest that acupuncture could improve hippocampus function by modulating the cAMP/PKA/CREB signaling pathway, which represents a molecular mechanism of acupuncture for recognitive function in cerebral multi-infarction rats.

Click chemistry mediated Eu-tagging: activity-based specific quantification and simultaneous activity evaluation of CYP3A4 using 153Eu species-unspecific isotope dilution inductively coupled plasma mass spectrometry.

P450 3A4 (CYP3A4) is one of the most important isoforms in the human cytochrome P450 superfamily. It was used as an example in this proof-of-concept study in order to demonstrate an activity-based labeling and then click chemistry (CC) mediated element-tagging strategy for simultaneously specific quantification and activity measurement of an enzyme using species-unspecific isotope dilution inductively coupled plasma mass spectrometry (SUID ICPMS). A dual functional hexynylated 17α-ethynylestradiol activity-based probe was synthesized for specifically labeling CYP3A4 and then CC-mediated Eu-tagging with an azido-DOTA-Eu complex for CYP3A4 quantification and activity measurement in human liver microsome and serum samples using (153)Eu SUID ICPMS. The LOD (3σ) of CYP3A4 reached 20.3 fmol when monitoring (151/153)Eu ICPMS signals, in addition to the merits of specificity and simultaneous activity measurement achieved. We believe that this activity-based CC-mediated element-tagging strategy will liberate more potential advantages of ICPMS in bioanalysis.

[Rapid determination of pesticide multiresidues in porphyra by dispersive solid-phase extraction coupled with online gel permeation chromatography-gas chromatography-mass spectrometry].

A rapid method for the simultaneous identification and quantification of pesticide multiresidues in porphyra was developed using gel permeation chromatography (GPC) coupled to gas chromatography-mass spectrometry (GPC-GC/MS). Nineteen pesticides (organochlorines, organophosphoruses, triazines and pyrethroids) were selected as the target analytes. The pretreatment method was applied consisting of organic solvent extraction followed by dispersive solid-phase extraction with graphitized carbon black (GCB) and primary secondary amine (PSA) adsorbents. GPC was also employed online to remove the large molecules such as pigments and lipids. The quantitative analysis was carried out by external standard method using gas chromatography coupled with mass spectrometry in selective ion monitoring (SIM) mode. Moreover, a large volume of sample was allowed to be injected using the program of GPC programmed-temperature vaporizer of gas chromatography to improve the sensitivity of measurements. The results showed that the calibration curves were linear (r > 0.995) in the range of 10-1,000 µg/L for all the pesticides. The limits of detection (LODs) for the pesticides in porphyra were from 0.005 to 0.03 mg/kg, and the average recoveries were between 70% and 120%. The advantages of the method are simple, sensitive and shorter operation time for analysis of pesticide residues in porphyra samples.

Highly sensitive detection of organophosphorus pesticides by acetylcholinesterase-coated thin film bulk acoustic resonator mass-loading sensor.

An acetylcholinesterase-coated thin film bulk acoustic resonator has been developed for the detection of organophosphorus pesticides. The thin film bulk acoustic resonator acts as a robust mass-sensitive transducer for bio-sensing. This device works in thickness shear mode with a resonance at 1.97 GHz. The detection is based on the inhibitory effects of organophosphorus compounds on the enzymatic activity of the acetylcholinesterase immobilized on one of the faces of the acoustic resonator. The enzyme reaction in the substrate solution and the inhibitory effect is observed are real time by measuring the frequency shift. The presence of organophosphorus pesticides can be detected from the diminution of the frequency shift compared with the levels found in their absence. The device exhibits linear responses, good reproducibility, simple operation, portability and a low detection limit of 5.3×10(-11) M for paraoxon. The detection results of organophosphorus pesticide residues in practical samples show that the proposed sensor has the feasibility and sensing accuracy comparable to gas chromatography.