Noble nanomaterials also present special physical and chemical properties and, therefore, they provide model building blocks in modifying nanoscale structures for various applications, ranging from nanomedicine to catalysis and optics. The introduction of noble metal nanoparticles (NPs)  into chemically derived graphene is important in opening new avenues for both materials in different fields where they can provide hybrid materials with exceptional performance due to the synergistical result of the specific properties of each of the materials. This review presents the different synthetic procedures for preparing Pt, Ag, Pd and Au NP/graphene oxide (GO) and reduced graphene oxide (rGO) composites.In recent years, research has focused on developing materials exhibiting outstanding mechanical, electrical, thermal, catalytic, magnetic and optical properties such as graphene/polymer, graphene/metal nanoparticles and graphene/ceramic nanocomposites. Two-dimensional sp Considering its superior electronic characteristics, graphene sheets offer potential solutions for the manufacturing of nanocomposites . Th. Th13]. Abdulhusain et al.  prepared3O4-decorated graphene oxide and Au-Mn3O4 nanocomposite  is reacted for 24 h at 180 °C inside a high-pressure reactor. Afterward, a formulated (BNrGO)5-(CNT)5 support material is inserted into the ethylene–glycol mixture containing K2PtCl4, and additional processing at 120 °C is performed for a duration of 12 h in order to obtain the final product. Another ration of BNrGO-CNT was also prepared. The results obtained through ICP-MS determined the Pt loadings for the Pt/(BNrGO)3-(CNT)7, Pt/(BNrGO)5-(CNT)5, Pt/(BNrGO)7-(CNT)3, Pt/(BNrGO)9-(CNT)1, Pt/rGO, Pt/CNT and Pt/C catalysts at 18.20, 18.80, 18.70, 19.40, 18.40, 19.00 and 19.60 wt%, respectively. Pt particles show interplanar spacings of 0.223 nm and 0.195 nm, accurately corresponding to the (111) and (200) planes of face-centered cubic Pt crystals. Through TEM it can be observed that the Pt/BNrGO-CNT hybrid is composed mainly of the four types of elements , with these four components being uniformly distributed throughout the whole sheets. XRD reveals that the aforementioned diffraction peak appears at approximately 2θ = 25.0° for Pt/rGO and Pt/BNrGO-CNT. The three characteristic diffraction peaks of metallic Pt correspond to the (111), (200) and (220) crystal planes within the cubic Pt structure. The intensity ration between D and G (ID/IG) of the GO, rGO and Pt/rGO samples was determined as 0.87, 1.04 and 0.96, respectively, proving the presence of several topological flaws in the carbon structures. Conversely, the ID/IG of Pt/BNrGO-CNT proved a low 0.76, hinting towards the fact that the incorporation of low-defect CNTs into the rGO skeletons can be used as a means of reducing the density of defects of the hybrid system. The specific surface areas are 226 m2 g−1 and 240 m2 g−1 for Pt/BNrGO-CNT and BNrGO-CNT hybrids, respectively.Xue et al. used the thermally assisted method to anchor Pt nanocrystals onto three-dimensional (3D) porous boron and nitrogen double-doped reduced graphene oxide–carbon nanotube frameworks (Pt/BNrGO-CNT) . They us2 + Ar) mixture [2 solution in diluted HCl by wet impregnation, resulting in 10 wt.% palladium on rGO. The target Pd-Au/rGO with metal concentrations of 7.5 wt.% Pd and 2.5 wt.% Au was obtained similarly through wet impregnation, employing a mixture comprising PdCl2 and HAuCl4 aqueous solutions. In both cases, the impregnated samples were dried at ambient temperature followed by thermal reduction in Ar. Afterward, it was thermal treated in a (H2 + Ar) mixture (10 vol.% H2) for 30 min at 250 °C. The resulting surface areas determined using the BET method are 210 m2/g for Pd/rGO and 206 m2/g for Pd-Au/rGO. The XRD profiles for both of the catalysts display the (002) graphene reflexion situated at 23° for Pd/rGO and at 23.7° for the Pd-Au bimetallic catalyst, as well as the metal reflexions: Pd (111) at 39.9°, Pd (200) at 46.4° for Pd/rGO and one metal reflexion situated at 38.9° for Pd-Au/rGO. Size of the metal crystallite calculated by XRD is 5 nm for Pd/rGO and 3.5 nm for Pd-Au/rGO. The bimetallic catalyst displays a distance between the carbon layers of 0.378 nm, whereas the Pd/rGO catalyst displays a distance of 0.386 nm, with the medium number of graphene layers for Pd/rGO being 4 and for Pd-Au/rGO being 5.5. The burning temperature for the palladium containing catalyst is 517 °C, while the Pd-Au bimetallic has a much lower value at 447 °C. The ID/IG intensity ratio of the D and G bands is 0.90 for Pd/rGO and 0.85 for Pd-Au/rGO; values were obtained from Raman spectra [Grad et al. prepared Pd/rGO and Pd-Au/rGO catalysts through wet impregnation of GO using an aqueous mixture of metal salts. The resulting mixture of GO and metal ions was then processed through thermal treatment in  presented by the Raman spectra of the investigated samples proved to be equal to 0.60 for GO, 0.63 for rGO and 0.81 for Au/rGO, respectively. The obtained ID/IG values are directly proportional with the number of structural defects in the sample.Wojnicki et al. synthesized Au/rGO. They first dissolved metallic gold in aqua regia to obtain the Au(III) chloride complex . A MagnuGold nanoparticles decorated on rGO were prepared using the microwave-assisted process (MW). The procedure has demonstrated remarkable advantages as eco-friendly method for Au/rGOs obtaining with simultaneous reduction in graphene oxide and formation of gold nanoparticles by an innovative one-step process. The characterization of prepared samples demonstrated good chemical stability and controllable morphology. The samples were used for membrane electrode assembly development and tested in operation of proton exchange membrane fuel cells. The electrochemical stability of the innovative Au/rGO-based cathode was analyzed using several accelerated stress tests (ASTs) by considering the cycling potential protocol. The electrochemical analysis considering the I-V study, cyclic and linear voltammetry has provided improved performances in comparison with the standard commercial cathode. The aggressive AST indicated an excellent stability; thus, the authors reported an improved electrocatalyst for oxygen reduction reaction with higher stability and durability for fuel cells. Moreover, the paper indicates the possibility of extending the protocol using the microwave-assisted process for obtaining other noble metal nanoparticles supported on rGO ,37,38,39A single-step route to obtain platinum/platinum-cobalt uniformly distributed nanoparticles supported on reduced graphene oxide was developed recently. This route provides significant advantages such as its low cost, low time-consuming nature and high yield in comparison to state-of-the-art chemical methods used to prepare efficient Pt/rGO catalyst. The morphology of prepared samples has been evaluated by specific techniques, while the electro catalytic durability has been evaluated using the electrochemical performances in fuel cells ,41. SignThe ultrasonic method  leads to−1~3200 cm−1 [Li et al. synthesized the AuPs/rGO through a mixture of 4 mg/mL GO suspension and 0.48 mg/mL AuPs solution with a volume ratio of 1:1; it was then sonicated for 1 h. After the AuPs/rGO was reduced by hydroiodic acid, it was washed with deionized water and then dried in air. The images of AuPs/rGO composite by SEM showed the uniform distribution of the AuPs on the surface of rGO. XRD measurements and the corresponding calculated results are consistent with the SEM images. The AuPs composites are inserted into the layered graphene sheets. The diffraction peak is 2θ = 8.84° for rGO to 2θ = 7.86° for AuPs-rGO. XRD measurements are consistent with the SEM images. The Raman spectra of both the AuPs-rGO and pure rGO film show a wide band at 2400 cm200 cm−1 .3 was added. Centrifugation at 12,000 rpm was used to separate the final Ag/GO nanocomposites, which were then washed with double-distilled water [−1 for hydroxyl -OH, carboxyl -COOH, epoxy C-O-C and alkoxyl C-O. After anchoring with AgNPs, the epoxy stretching mode at 1226 cm−1 is no longer present, while peaks that are indicative of different oxygen functional groups remain well-preserved. Nanocomposites of GO and Ag/GO reveal a diffraction peak at 2θ of 10.1° in the XRD patterns relating to the (002) crystal plane of GO nanosheets. In addition to the distinctive diffraction peak of GO, the Ag/GO nanocomposites also exhibit a number of separate peaks at 2θ of 38.1, 44.2, 64.5 and 77.5 degrees, which can be attributed to the (111), (200), (220) and (311) facets of typical fcc metallic Ag (JCPDS No. 04-0783), respectively. The patterns revealed by XRD demonstrate the effective adhesion of AgNPs to GO nanowires. The ID/IG ratio of the GO increased from 0.87 to 0.92 when the GO was anchored with AgNPs. The C/O atomic ratio found in the Ag/GO nanocomposites is greater than that of the GO, with the values obtained by XPS analysis being 2.6 for Ag/GO and 2.0 for GO. FTIR showed the characteristic peaks for Ag/GO centered at 284.8, 287.0 and 288.5 eV, respectively, for C=C, C-O and C=O. XPS analyses determined an atomic percentage of Ag of 2.46.Tran et al. prepared the Ag/GO nanocomposites through the ultrasonication method. Ag/GO nanocomposites were synthesized with GO, double-distilled water and sonicated for a duration of 10 min, after which AgNOed water . The FTI+-decorated PCN-222  using ZrCl4, meso-tetra(4-carboxyphenyl) porphine, benzoic acid and AgNO3. The solution was ultrasonically dispersed and dissolved in a mixture of N, N-dimethylformamide and acetic acid. Preparing the Ag+-decorated PCN-222@EDTA-GO-CS (CS-chitosan) foam required AgNO3, ZrCl4, H2TCPP and benzoic acid that were ultrasonically solvable in the N, N-dimethylformamide [+-decorated PCN-222 showed clear crystal lattices and the TEM-EDS mappings revealed the even dispersion of Ag, C, N, O and Zr elements. The characteristic peaks of PCN-222 at 1709, 1640, 1603, 1549, 1402, 967, 801 and 719 cm−1 were obtained through the use of FT-IR spectra. The peaks in binding energies at 367.5 eV and 373.5 eV arose from the presence of Ag+ ions.Bi et al. prepared PCN-222 and Agormamide . The Ag+2PtCl6 was added to the solution. In the XRD patterns, the characteristic peaks of (002) obtained at 11.0° (d002 = 0.85 nm), 28.9° and 24.0° were for GO, CQDs and Pt/CQDs/rGO, respectively. After the reduction process, the diffraction peaks are observed at, respectively, 46.9°, 55.7° and 81.6°, corresponding to (111), (200) and (220) planes of the face-centered cubic (fcc) structure of Pt. The crystallite sizes of GO, CQDs and Pt/CQDs@rGO are 54 nm, 17 nm and 38 nm, respectively. The large crystallite size of Pt/CQDs@rGO shows the aggregation of Pt nanoparticles. D and G bands have been detected using Raman spectroscopy at 1350 cm−1 and 1580 cm−1 for graphene oxide, 1350 cm−1 and 1580 cm−1 in the case of GQDs and at 1357 cm−1 and 1590 cm−1 for Pt/N-CQDs@rGO nanocomposite. The values of the ratios between the intensities of the D and G bands (ID/IG) are 0.88 and 0.94 and have been calculated for GO and Pt/CQDs@rGO, respectively. The EDX investigation exhibits all essential elements such as C, O, Na, Cl and Pt individually from Pt/CQDs@rGO nanocomposite.Mehmandoust et al. prepared Pt/CQDs@rGO nanocomposite (CQDs—carbon quantum dots) by ultrasonication method . The Pt/4 as reducing agents through the ultrasonication method [3 and glucose are added. The same experimental procedure is repeated for vitamin C and NaBH4. The Ag/rGO samples have peaks at 2θ values of 38.13°, 44.34°, 66.44° and 77.44°, which agree with the cubic crystal structure of the Ag NPs. The grain sizes of the Ag NPs are 28 nm, 25 nm and 22 nm for GAg_G, GAg_V and GAg_S, respectively. Through Raman spectroscopy, a graphitic band (G band) at 1590 cm−1 and a disorder band (D band) at 1365 cm−1 can be observed. The absorption bands at 257 nm for GAg_G, 268 nm for GAg_S and 270 nm for GAg_V. ID/IG ratios have been measured at 1.26 for GAg_S, 1.19 for GAg_V and 0.94 GAg_G.Mariappan et al. presented the study of Ag/rGO prepared with glucose, vitamin C and NaBHn method . The Ag/Aljafari et al. prepared Pd/GO using the sonication technique. This process used GO, palladium acetate and glucose. The pH was adjusted using NaOH . The dif3 and cetyl trimethylammonium bromide. The final product is graphene oxide sheets that are decorated by silver nanoparticles. The peaks at 38.31°, 44.41°, 63.51° and 77.71° can be assigned to the (111), (200), (220) and (311) crystalline planes of silver, respectively, which shows that the silver nanoparticles are composed of pure crystalline silver. The particle diameter of silver nanoparticles is about 10 nm. The band of silver nanoparticles is at about 414 nm.Mao et al.  synthesiFor the preparation of graphene materials doped with metal nanomaterials, the synthesis methods are very important to ensure the best dispersion of the metal particles and the narrowest distribution of their size, because both significantly affect the electrocatalytic activity.In In recent years, different methods have been proposed for the synthesis of nanoparticles deposited on a graphene support. The choice of the most suitable method has the greatest importance in terms of the structure and catalytic efficiency of the catalysts. In conclusion, the most valuable method among the preparation methods of graphene-deposited nanomaterial catalysts is microwave field irradiation, especially due to the short synthesis time, the fast and uniform heating and the significant challenge in controlling uniformity of the metal nanoparticle’s decoration on the graphene surface. By applying irradiation in the microwave field, under the influence of temperature, homogeneous reaction centers are formed in the reaction medium at the interface between the irradiation-sensitive graphene support and the metal precursor. Additionally, the presence of a reducing agent in the reaction medium means that the precursor can be converted to its metallic form by microwave irradiation.The qualities of noble metals have demonstrated a special efficiency in the electrocatalytic activity and the electrochemical stability of compounds based on carbon and graphene oxide. In order to improve the oxygen reduction reaction (ORR) and the quality of hydrogen adsorption and desorption, a higher electrochemical active surface area (ECSA) of the catalyst based on noble metals is necessary. The intrinsic increase in the active surface is proportional to the metal content in the chemical compound and to the dispersion of metal nanoparticles on the rGO sheets. The uniform distribution and surface morphology of noble metal nanoparticles on rGO have an effect on the ORR. An excessive reaction energy can cause an agglomeration of the noble metal nanoparticles, leading to particle sizes over 10 nm and the suppression of catalytic activity by reducing the active surface. This review presents the most used and up-to-date methods for the synthesis of graphene functionalized with noble metals  as well as the relevant methods for the characterization of catalysts. The potential of capitalizing on the improved catalytic properties of graphene functionalized with noble metals was also discussed. The studies presented in this review were carried out in order to understand how the metal–support interaction drives chemical catalysis. The preparation technique, the type and amount of metal, the nature of the support, the type of dopant and the technique of applying the catalyst, all these are dependent on the metal–support relationship. Following this review, it was found that the noble metals demonstrated a special efficiency in the electrocatalytic activity and the electrochemical stability of the compounds based on carbon and graphene oxide. It was also observed that to improve the oxygen reduction reaction (ORR) and the quality of hydrogen adsorption and desorption, a higher electrochemical active surface area (ECSA) of the noble metal catalyst is required. The uniform distribution and surface morphology of noble metal nanoparticles on rGO were found to have an effect on the ORR. Most applications of noble metal functionalized graphene are in fuel cells, renewable energy  and supercapacitor applications.


Pseudomonas aeruginosa are global antimicrobial resistance threats. Cefepime-taniborbactam (FTB) is an investigational β-lactam/ β-lactamase inhibitor combination that is active against Enterobacterales and Pseudomonas aeruginosa expressing serine and metallo-β-lactamases. CERTAIN-1 (Cefepime Rescue with Taniborbactam in cUTI) evaluated FTB efficacy and safety in the treatment of cUTI.Carbapenem-resistant Enterobacterales and multidrug resistant CERTAIN-1 was a randomized, double blind, double dummy, Phase 3 study comparing FTB to meropenem (MEM) in adults hospitalized with cUTI or AP. The primary endpoint was the composite  response at the test of cure (TOC) visit in the microITT population. Patients were randomized 2:1 to FTB 2.5g IV q8h or MEM 1g IV q8h for 7 days or up to 14 days in patients with bacteremia. The non-inferiority margin was -15.0% and a pre-specified test for superiority for the primary endpoint was performed following confirmation of non-inferiority.> 65 years represented 38.0% of the microITT population and bacteremia was present in 13.1% of patients. Composite success was achieved in 70.0% and 58.0% of FTB and MEM patients, respectively . FTB was statistically superior to MEM for the primary endpoint at the TOC  and statistical superiority was sustained at the late follow up visit (LFU). Analyses of secondary endpoints and subgroups were consistent with the primary efficacy analysis. Treatment-emergent adverse events (TEAEs) were observed in 35.5% of FTB patients and 29.0% of MEM patients. Serious adverse events occurred in 2.0% and 1.8% of FTB and MEM patients, respectively. The most common TEAEs were headache  and diarrhea .A total of 661 patients were randomized and 436 patients (66.0%) were included in the microITT population, including 42.2% with AP and 57.8% with cUTI. Patients Following 7 days of therapy, FTB was statistically superior to MEM for the primary endpoint at the TOC. Composite success for FTB remained statistically superior to MEM at the LFU visit. FTB was safe and well-tolerated with a safety profile similar to MEM.Paul C. McGovern, MD, Paratek Pharmaceuticals: Stocks/Bonds|Venatorx Pharmaceuticals: employee|Venatorx Pharmaceuticals: Stocks/Bonds Florian Wagenlehner, MD, Glaxo Smith Kline: Advisor/Consultant|Spero Pharmaceuticals: Advisor/Consultant|Venatorx Pharmaceuticals: Advisor/Consultant Leanne Gasink, MD, MSCE, Amplyx Pharmaceuticals: Advisor/Consultant|CSL Behring: Advisor/Consultant|Pfizer: Advisor/Consultant|Spero Therapeutics: Advisor/Consultant|Venatorx Pharmaceuticals: Advisor/Consultant|Vera Therapeutics: Advisor/Consultant Greg Moeck, PhD, Venatorx Pharmaceuticals: Employee|Venatorx Pharmaceuticals: Stocks/Bonds Patrick L. McLeroth, MD, Labcorp: Full time employee|Labcorp: Stocks/Bonds Mary Beth Dorr, PhD, Merck: Past employee|Merck: Stocks/Bonds|Pfizer: Stocks/Bonds|Venatorx: Employee Aaron Dane, MSc, Amplyx: Advisor/Consultant|AN2 therapeutics: Advisor/Consultant|Artizan: Advisor/Consultant|Cidara: Advisor/Consultant|ContraFect: Advisor/Consultant|Correvio: Advisor/Consultant|Davolterra: Advisor/Consultant|Destiny Pharma: Advisor/Consultant|Entasis: Advisor/Consultant|F2G Limited: Advisor/Consultant|GSK: Advisor/Consultant|Humanigen: Advisor/Consultant|Kymab: Advisor/Consultant|Modis: Advisor/Consultant|Orca: Advisor/Consultant|Pfizer: Advisor/Consultant|Phico: Advisor/Consultant|Pled Pharma: Advisor/Consultant|Rare Thyroid: Advisor/Consultant|Roche: Advisor/Consultant|Scynexis: Advisor/Consultant|Sinovent: Advisor/Consultant|Spero Therapeutics: Advisor/Consultant|Transcrip: Advisor/Consultant|Venatorx: Advisor/Consultant Tim Henkel, MD, PhD, Venatorx Pharmaceuticals: Stocks/Bonds.


Guidelines recommend regular physical activity (PA) and decreased sedentary time (SED) for patients after myocardial infarction (MI). Therefore, valid self-assessment of PA is vital in clinical practice. The purpose of this study was to assess the convergent validity of commonly used PA and SED questions recommended by the National Board of Health and welfare (NBHW) and national SWEDEHEART-registry using accelerometers as the reference method in patients after MI.Data were obtained 2017–2021 among Swedish men and women (180 assessments). Participants answered five commonly used PA and SED-questions (by NBHW and SWEDEHEART) and wore an accelerometer (Actigraph GT3X) for seven days. Convergent validity was assessed gradually by; Kruskall Wallis-, Sperman rho, Weighted Kappa- and ROC-analyses. Misclassification was explored by Chi-square analyses with Benjamini–Hochberg adjustment.The strongest correlation (r = 0.37) was found for the SED-GIH question (NBHW). For PA, no specific question stood out, with correlations of r = 0.31 (NBWH), and r = 0.24–0.30 (SWEDEHEART). For all questions (NBHW and SWEDEHEART), there was a high degree of misclassification (congruency 12–30%) affecting the agreement (0.09–0.32) between self-report and accelerometer assessed time. The SED-GIH, PA-index and SWEDEHEART-VPA had the strongest sensitivity for identifying individuals with high SED (0.72) or low PA (0.77 and 0.75).The studied PA and SED questions may provide an indication of PA and SED level among patients with MI in clinical practice and could be used to form a basis for further dialogue and assessment. Further development is needed, since practical assessment tools of PA and SED are desirable.The online version contains supplementary material available at 10.1186/s13102-022-00509-y. Cardiovascular disease (CVD) is the major cause of death in Europe . There iInternational guidelines strongly recommend individualised PA and SED recommendations for patients with CVD , 7. Ther“The Swedish National Board of Health and Welfare” (NBHW) recommends two PA questions (NBHW-PA questions) to assess levels of everyday PA, exercise, and a combination of these . They alThus, the current questions seem to have an acceptable predictive validity for identifying individuals with a higher risk of rehospitalization and all-cause mortality , 3. HoweThis was a single center study performed at the Department of Cardiology at Danderyd’s hospital, Stockholm, Sweden. Individuals were informed of the study and invited to participate by a nurse at the first routine follow-up visit, six to eight weeks after MI.Data were collected via questionnaires, accelerometers, diaries, and the national quality registry SWEDEHEART between October 2017 and May 2021. The participants answered a questionnaire concerning PA and SED both at six to eight weeks and at 10–12 months post MI. Immediately after the visits, an accelerometer with a diary were posted to the participant, with instructions on how to wear the device and to start the accelerometer assessment directly. After the measuring period, participants returned the accelerometer and the diary by prepaid postage to the Swedish School of Sport and Health Science. Covariates were collected from a sub-register of SWEDEHEART, the SEPHIA-registry, focusing on outpatient cardiac rehabilitation up to one year after MI from the responsible cardiac department . The folIndividuals aged 18–80 years with newly diagnosed MI (ICD I21) registered in the SWEDEHEART registry were included in the study. Individuals who were physically disabled (wheelchair dependent) or with reduced ability to answer the questions were excluded by not being asked for participation.Six commonly used PA and SED questions were included in the study .The NBHW-PA and SED questions use different time frames. The NBHW-PA questions focus on PA in an ordinary week while the SED question focuses on an ordinary day.Everyday PA: “During a regular week, how much time (in minutes) are you physically active in ways that are not exercise, for example walks, bicycling, or gardening? Add together all activities lasting at least 10 min”. Seven fixed answers were available; “no time”, ” < 30”, “30–60”, “60–90”, “90–150”, “150–300” and “ > 300”.Physical exercise: During a regular week, how much time do you spend exercising on a level that makes you out of breath, for example running, fitness class, or ball games? The questions had six fixed answers; “no time”, “ < 30”, “30–60”, “60–90”, “90–120” and “ > 120”.The questions of everyday PA and physical exercise formed a validated PA-index (3–19 points) of total PA level . This waSedentary time: “How much time (in hours) do you sit during a normal day, excluding sleep?” . There wSWEDEHEART-MVPA: “Number of physical activity sessions of at least 30 min (two 15-min sessions can be combined into one 30-min session) in the last 7 days, with a minimal intensity of fast walking”.The PA questions in the SWEDEHART registry focus on PA during the previous week and are listed below. The individuals can choose between 0–7 days/week SWEDEHEAR: How many days in the last week did you do at least 30 min, total time (at least 10 min at a time) of physical activity that made you slightly out of breath and gave slightly elevated heart rate?SWEDEHEART-MPASWEDEHEART-VPA: How many days during the last week did you do some form of continuous vigorous physical activity /exercise, (at least 20 min), that made you out of breath and gave you elevated heart rate?The following two questions separate moderate and vigorous intensity levels of PA.An accelerometer provides information on body movements, which can be translated to data on PA, using validation algorithms. Such translation gives reasonably valid data of duration, frequency, and absolute intensity of PA and SED , 17. AccThe accelerometers and data files were processed using the software Actilife, version 6.13.4 . Data were collected tri-axially using a sampling rate of 30 Hz and, after extraction, data were down-sampled and saved as 60 s epochs, with activity intensity based on vector magnitude. Normal frequency filter was applied. Additionally, the Choi algorithm was used for validate non-wear time , definedCut-points were used to describe the daily PA behaviour, using the following components: time spent in sedentary , light physical activity , moderate PA (MPA 2690–6166) and vigorous physical activity  . The meaTo be included in the analyses, individuals had to answer the questionnaire and have valid accelerometer data for at least seven days defined as ≥ 600 min of data per day after non-wear time had been excluded.The descriptive data are presented as median and 25th and 75th percentiles (Q1–Q3), or number and proportions. Before Spearman and weighted Kappa analysis, the continuous accelerometer data were divided into the same categories as the different answer options for PA  and SED questions. For the PA-index, the average number of minutes in MVPA was kept as a continuous variable.Convergent validity was explored gradually:Multiple linear regressions were performed to control for repeated measurements among individuals with two assessments. The study-id was used as a covariate.Correlations between the categorised accelerometer data and the PA and SED-GIH questions were calculated using Spearman’s rho (r). The associations were interpreted as weak (r < 0.10), modest (r 0.1–0.3), moderate (r 0.3–0.5), strong (r 0.5–0.8) or very strong (r 0.8–1.0) . To explWeighted kappa analyses were conducted to assess the agreement between PA and SED-GIH questions with the categorized accelerometer data (n.a. for the PA-index). The agreement (Kappa) was interpreted as poor (< 0.20), fair (0.21–0.40), moderate (0.41–0.60), substantial (0.61–0.80) or almost perfect (0.81–1.00) .Receiver operating characteristics (ROC) curves were calculated. The answer options were kept as ordinal variables and the accelerometer data were used as state variable (golden standard) and dichotomised at a cut-off value. The cut-off for achieving the PA recommendations was ≥ 150 min a week of MPA for the everyday PA question, ≥ 75 min a week of VPA for the exercise question and ≥ 150 min of MVPA for the PA-index (NBHW-PA questions). For the SWEDEHEART questions the accelerometer cut-off was set at ≥ 20 min of total VPA for at least 3 days a week and ≥ 30 min of total MPA or MVPA for at least 5 days a week. Individuals with ≥ 9.5 h of SED per day were categorized as high SED, while individuals with less time spent sedentary were categorized as low SED. This cut-off was chosen based on international PA , 7 and SKruskal–Wallis analyses with Bonferroni correction were performed to investigate the differences in median accelerometer derived time between the different categories of the PA and SED questions.Lastly, calculations in Excel were performed to assess how well the PA and SED-GIH questions corresponded to the accelerometer derived data, either congruent, or over- and underestimating. Then, Chi square analyses with Benjamini–Hochberg adjustments were used to assess differences in congruence, under-, and overestimation between gender, age, LVEF, systolic blood pressure, diabetes, and BMI. All statistical analyses were performed using SPSS 27.0 software .n = 2), (n = 3) everyday-PA and exercise (n = 2) questions.A total of 123 individuals answered the questionnaire and provided complete accelerometer data were included  in accelerometer collected time for different categories of the NBHW-PA and SED questions Fig. a. For evIn the multiple linear regression , but the agreement was weak (0.09). However, the ability to classify individuals sitting ≥ 9.5 h was good, with an acceptable level of both sensitivity 72%) and specificity (62%)  and no specific question had a significantly stronger correlation (p > 0.47). In general, the weighted kappa analyses showed a poor to fair agreement between the answer categories and categorised accelerometer data (agreement 0.10–0.32) with the strongest agreement (fair) for the SWEDEHART-MVPA and the NBHW everyday PA questions. Results from the ROC analyses are presented in Table 4–0.31 anCongruence between individuals’ self-reported PA and SED levels compared to the accelerometer measured data varied between 12 and 30% .There were few other factors affecting the reporting pattern. Older individuals over-reported everyday PA to a higher degree than younger 63% vs 44%). Men under-reported the number of PA sessions in MVPA (SWEDEHEART) compared to women (39% vs 18%)  frequently used in clinical practice, there was a high degree of misclassification (over- and underreporting), with modest to moderate correlation and poor to fair agreement. Nonetheless, they may provide some broadly indication about PA and SED level, which could form the basis for further discussion and assessment on an individual level in clinical practice.The updated European guidelines of CVD prevention highlight the importance of decreasing SED; therefore it is important that the convergent validity of the SED-GIH question has an acceptable level. The correlation to accelerometer assessed SED (r = 0.37) was somewhat lower compared to a study in a general population that showed a correlation between the SED-GIH question and total stationary time (r = 0.48) . ConvergThe agreement between the SED-GIH question and SED time assessed by accelerometer was weak. This might be due to eight out of ten participants underestimating their sedentary time causing misclassification. The difficulty to estimate SED has been shown in other studies, with misclassifications of minus 85 min per day  and undeFor PA, the correlation for the NBHW and SWEDEHEART questions with accelerometer derived data was modest to moderate. There were no significant differences in correlation for the NBHW-PA questions compared to the SWEDEHEART ones. The correlation of NBHW PA-index with accelerometers as a criterion method showed similar results to a study in a general population, r = 0.27 (vs 0.31) for MVPA min/day . To our For the PA questions, agreement with time assessed by accelerometer was poor to fair (0.10–0.32) with the strongest agreement for the SWEDEHEART-MVPA and the NBHW everyday PA question. As with a previous study , there wROC-curve analysis of the PA-index was in line with a study in a general population (0.66) . TheretoThe most important strength of this study is that it includes a large cohort of CVD patients, focusing on individuals with MI. It also includes a higher proportion of older individuals compared to previous studies , 11. GenA limitation is the lack of consensus for cut-off values of accelerometer data for individuals with CVD. This is troublesome and the time spent at different levels of PA intensity should therefore be interpreted with caution, especially in patients with reduced cardiorespiratory fitness, which leads to differences in absolute and relative intensity . In thisAccelerometers were used as the criterion method. However, accelerometer measurements do have disadvantages. For example, information about relative intensity and activities such as bicycling, strength training, and swimming, all common types of exercise within cardiac rehabilitation , are omiAnother limitation is that participants may have become more conscious about their PA behaviour during the study, affecting how they answered the questions and their PA behaviour during the measurement period. The participants answered the questionnaire prior to wearing the accelerometer, this may be a limitation when the SWEDEHEART-PA questions focus at PA the last week, however several studies indicate that accelerometer data collected over seven days use to be consistent between weeks (high reliability) –31. Non-Exploring an individual’s total PA behaviour is complex and these questions have a limited convergent validity and low, precision. Therefore, a combination of both accelerometer derived data and questionnaires could be recommended , 9.The present study is clinically important as it focuses on commonly used SED and PA questions to patients with CVD, where regular PA and low SED both have a central role in cardiac rehabilitation. The convergent validity of the SED and PA questions is poor to moderate compared to accelerometer assessed data. The SED-GIH question had the strongest correlation. However, for PA we could not identify any preferable question. In spite of the risk of misclassification when using questionnaires, the questions seem to be a practical and acceptable method that may provide some indication about PA-level, which could form the basis for further discussion and assessment on an individual level.Additional file 1. Correlations between self-rated PA level, SED and study-id with accelerometer assessed PA and SED time.Additional file 2. Graphical results of the ROC analyses for the specificity and sensitivity of the PA and SED questions compared to accelerometer collected data.Additional file 3. Convergent validity for self-reported estimates of physical activity and sedentary using accelerometer assessed time as reference for time point one (n=123) and two (n=56).Additional file 4. Congruence between self-reported PA and SED compared to the accelerometer assessed time for different strata.


Bone defects and fractures heal slowly compared with injuries to other tissues, creating a heavy burden for patients, their families, and society. Alongside conventional treatment methods for fractures and bone defects, adjuvant therapies play an important but underappreciated role. In a previous study, we found that systemic administration of flufenamic acid promoted osteogenesis in vivo, but its side effects limited the application of our findings. In the present study, we assess the effects of external butyl flufenamate ointment on the healing of cranial defects in mice. We found that application of butyl flufenamate ointment on the surface of the skin accelerated the healing of cranial defects in mice by promoting BMP2 secretion from mouse-skin mesenchymal stem-cells. These findings indicate that butyl flufenamate ointment has potential therapeutic value for treating superficial fractures or bone defects while avoiding the toxicity and side effects of systemic medication, representing a safe and convenient adjuvant therapy to promote healing of superficial bone defects and fractures. Bone defects caused by fractures, tumors, congenital malformation, trauma, and infection are common in the clinic and can cause serious harm to patients’ physical and mental health . Since tLocal administration has several benefits compared with systemic administration, including decreased side effects and toxicity, greater convenience, and improved patient compliance ,5. Thus,As a member of the non-steroidal anti-inflammatory drug family, flufenamic acid (FFA) is a representative of fenamic acid. It is commonly used in the clinic to treat inflammatory diseases, such as rheumatic arthritis ,9. Oral In the present study, we aimed to explore whether the topical application of butyl flufenamate ointment on the skin can promote healing of cranial defects in mice. Our results showed that topical application of butyl flufenamate ointment accelerated the healing of cranial defects in mice. Mechanistically, butyl flufenamate ointment accelerated healing by promoting BMP2 secretion from mouse-skin mesenchymal stem-cells (mSMSCs). This study provides valuable data on the potential application of butyl flufenamate ointment to treat superficial bone defects, representing a safe and convenient adjuvant therapy to promote healing.All animals used in this study were obtained from Vital River Corporation . The mice were kept in a pathogen-free facility on a 12:12-h light: dark cycle with water and food provided ad libitum. The Institutional Animal Care and Use Committee of the Peking University Health Science Center (LA2019019) approved the in vivo experiments in the present study, and all experiments were performed under relevant guidelines.Collage I, dispase V, fetal bovine serum (FBS), modified Eagle’s medium (MEM), and 100× penicillin/streptomycin solution were purchased from Gibco . Other reagents and materials were obtained from Sigma-Aldrich  unless stated otherwise.2 in area was carefully removed. The skin was then disinfected with 75% ethanol and washed with PBS. Then, the skin was cut as finely as possible and digested using a mixture of 2 mL collagenase I (4 mg/mL) and 2 mL dispase V (8 mg/mL) at 37 °C for approximately 1.5 h. After adding 4 mL MEM, the mixture was centrifuged at 1500 rpm for 5 min. The sediment was then resuspended in medium, filtered through a 70 μm sieve, and seeded in 100 mm dishes. The cells were then cultured in a 5% CO2 atmosphere at 37 °C in proliferation medium (PM) consisting of MEM, penicillin/streptomycin, and 20% (v/v) FBS. After 24 h, the cells were washed twice with PBS, and new PM was added. Cells from passages 2–4 were used in subsequent experiments.Eight-week-old female C57BL6 mice were used to isolate mSMSCs. After the mice were sacrificed, the skin on their backs was prepared and disinfected. An area of skin on the back approximately 1.5 × 1.5 cmv/v) FBS, penicillin/streptomycin, 10 mM β-glycerophosphate, 10 nM dexamethasone, and 0.2 mM L-ascorbic acid. The adipogenic medium (AM) was prepared by mixing MEM, 10% (v/v) FBS, penicillin/streptomycin, 100 nM dexamethasone, 200 μM indomecin, 10 μM insulin, and 500 μM 3-isobutyl-1-methylxanthine. The osteogenic medium (OM) was prepared by mixing MEM, 10% (v/v) FBS, penicillin/streptomycin, 10 nM dexamethasone, 10 mM β-glycerophosphate, and 0.2 mM L-ascorbic acid. The chondrogenic medium (CM) was prepared by mixing MEM, 10% (v/v) FBS, penicillin/streptomycin, 1 μg/mL ascorbic acid, 100 nM dexamethasone, 1% sodium pyruvate, 10% insulin–transferrin–selenium (ITS-Premix), and 10 ng/mL mouse transforming growth factor β1 . T. T20]. Togenesis e. The rebmp2 but not bmp4 -β protein superfamily . BMP2 haOur study still has several limitations. We used a mouse cranial defect model to assess the effects of external use of butyl flufenamate ointment; different bone defect/fracture models should be used to verify these effects. While we showed that butyl flufenamate ointment promoted BMP2 secretion in mSMSCs, further studies are required to comprehensively determine the mechanism of its effects.Overall, our results demonstrated that the external use of butyl flufenamate ointment promoted the healing of cranial defects in mice, suggesting that it might have therapeutic value as an adjuvant for external use to promote healing of superficial bone defects and fractures.The present study showed that topical application of butyl flufenamate ointment accelerates the healing of cranial defects in mice by promoting BMP2 secretion in mSMSCs. These findings suggest that butyl flufenamate ointment may have therapeutic value for treating superficial fractures or bone defects while avoiding the side effects of systemic medication, representing a safe and convenient adjuvant therapy.


LC and SN integrity was assessed using neuromelanin-sensitive MRI. Response inhibition was measured using fMRI with a stop-signal task. In healthy controls, LC (but not SN) integrity correlated with the stopping-related activity of the right inferior frontal gyrus (IFG) and right subthalamic nucleus (STN), which further correlated with stop-signal reaction time (SSRT). PD patients showed reduced LC integrity, longer SSRT, and lower stopping-related activity over the right IFG, pre-supplementary motor area, and right caudate nucleus than healthy controls. In PD patients, the relationship between SSRT and the fronto-subthalamic pathway was preserved. However, LC integrity no longer correlated with the stopping-related right IFG or right STN activity. No contribution of SN integrity was found during stopping. In conclusion, LC (but not SN) might modulate inhibitory functions of the right IFG-STN pathway. Damage to the LC might impact the right IFG-STN pathway during stopping, leading to response disinhibition in PD.A long-running debate concerns whether dopamine or noradrenaline deficiency drives response disinhibition in Parkinson’s disease (PD). This study aimed to investigate whether damage to the locus coeruleus (LC) or substantia nigra (SN) might impact inhibitory functions of the fronto-subthalamic hyperdirect or fronto-striatal indirect pathway. Patients with PD ( ResponsA long-held debate concerns whether dopamine or noradrenaline deficiency (or both) drives response disinhibition in PD. A series of human and animal studies using the selective noradrenaline reuptake inhibitor atomoxetine has shown the role of noradrenaline in response inhibition. In rats, atomoxetine can produce a dose-dependent speeding of SSRT . In healin vivo using neuromelanin-sensitive MRI. Neuromelanin is a by-product of catecholamine synthesis, existing in LC noradrenaline neurons and SN dopamine neurons , pramipexole (n = 16), piribedil (n = 8), rasagiline (n = 8), selegiline (n = 2), and amantadine (n = 5).We recruited 29 patients with idiopathic PD at the Zhongshan Hospital Department of Neurology between 2020 and 2021. Inclusion criteria were (a) PD diagnosed by senior neurologists according to the Movement Disorder Society clinical diagnostic criteria for PD , (b) Hoen = 2), violation of the race model for computing SSRT (n = 1), excessive head motion (n = 6), or feeling of discomfort during scanning (n = 1).2.2We recruited 29 healthy controls (HCs). Inclusion criteria were (a) age 50–75 years, (d) education > 6 years, and (c) right-handed. Exclusion criteria were (a) a history of significant neurological or psychiatric diseases, (b) alcohol or drug abuse, (c) possible current depression, (d) possible mild cognitive impairment (MoCA-B < 26/30), or (e) contraindications to MRI. Three additional HCs were excluded due to a violation of the race model for computing SSRT.2.3The stop-signal task was designed, analyzed, and reported following a recent consensus . The tas2.4We controlled behavioral data quality by monitoring whether the task performance met the independence assumption of the race model: (a) p is between 0.25 and 0.75, and (b) the reaction time of failed Stop is numerically shorter than the reaction time of Go . All inct tests .We estimated SSRT using a modified integration method that replaces Go omissions with the maximum reaction time, given the presence of Go omissions . We repo2.5Neuroimaging data were acquired on a Siemens Tim Trio 3T MRI scanner with a 32-channel head coil at the Brain Imaging Center of the Center for Excellence in Brain Science and Intelligence Technology.2, resolution 1 × 1 × 1 mm3).Structural T1-weighted images used a magnetisation-prepared rapid gradient-echo sequence .Neuromelanin-sensitive T1-weighted images used a fast spin-echo sequence with magnetization transfer pulse .Functional T2*-weighted images used a standard echo-planar imaging sequence . The first three images were discarded to allow magnetization equilibration. All other images were corrected for slice acquisition time difference, realigned to a mean functional image, registered to the high-resolution T1-weighted image, normalized to the Montreal Neurological Institute (MNI) coordinate system, smoothed with a Gaussian kernel of 4-mm full-width half-maximum, and filtered with a 128-sec high-pass filter.fMRI data were processed using SPM12 .We controlled fMRI data quality by monitoring the total displacement  and spatversus correct Go (SS > Go). A whole-brain two-sample t test was conducted at the group level .First, we computed the stopping-related regional activation. The general linear model convolved a design matrix with a canonical hemodynamic response function at the subject level. The design matrix included successful Stop, failed Stop, correct Go, Go commission errors, and Go omissions as separate regressors. Each trial was time-locked to its onset. The total displacement was included as a nuisance regressor. Classical parameter estimation was applied with a one-lag autoregressive model. The stopping effect was defined as successful Stop t tests .Second, we examined whether PD patients showed lower stopping-related activity than HCs in the fronto-subthalamic or fronto-striatal pathway. Regions of interest were derived from a meta-analysis of 99 stop-signal task fMRI studies (NeuroSynth) , includip < 0.05), and (b) whether the stopping-related activity of the fronto-subthalamic pathway correlated with LC or SN integrity or levodopa equivalent daily dose .Third, we explored relationships between LC integrity, fronto-subthalamic pathway, and SSRT. We examined: (a) whether SSRT correlated with the stopping-related activity of the fronto-subthalamic or fronto-striatal pathway . They manually identified the LC and SN in the native space to avoid potential distortion in normalization following previous studies .2 each) and pontine (20 mm2) were identified adjacent to the fourth ventricle in three contiguous slices. The LC contrast-to-noise ratio (CNR) was the difference between the mean LC signal intensity and mean pontine signal intensity divided by the SD of the pontine signal intensity . The SN area was summed up across sides.The SN was identified through a semi-automatical procedure to minimize subjective bias . The SN r = 0.98, p < 0.001; SN, r = 0.98, p < 0.001). Their mean measurements were used for statistical analysis. We examined whether PD patients showed reduced LC or SN integrity than HCs using one-tailed two-sample t tests .Measurements of the two raters were highly consistent  = −2.92, p = 0.002) and SN integrity than HCs (t(55) = −5.33, p < 0.001). The group difference in LC integrity was confirmed by an atlas-based analysis (see supplement)  .Fig. 2Lo3.2p = 0.42) or Geriatric Depression Scale score (p = 0.97).3.3t test). The IFG, preSMA, and caudate nucleus were more activated for successful Stop than correct Go. The STN did not show significant stopping-related regional activation at the group level, probably due to a larger inter-individual variability. Given the importance of the fronto-subthalamic pathway, we applied region-of-interest analysis to the STN.t tests, t(56) = −1.81, p = 0.038), preSMA (t(56) = −1.99, p = 0.025), and right caudate nucleus (t(56) = −2.25, p = 0.015), and marginally in the right STN (t(56) = −1.50, p = 0.07). However, these tests were only significant using one-tailed t tests and not corrected for multiple comparisons.PD patients showed lower stopping-related activity than HCs in the fronto-subthalamic and fronto-striatal pathways B, includ3.4F = 5.04, p = 0.004, R2 = 0.23) included the stopping-related right IFG , preSMA  and right STN activity  but excluded the stopping-related caudate activity (p = 0.78). Four outliers were analyzed separately (see supplement). Participants with higher stopping-related preSMA and right STN activity and lower stopping-related right IFG activity showed shorter SSRT.First, SSRT correlated with the stopping-related activity of the right IFG, preSMA, and right STN across groups C. A stepF = 4.76, p = 0.038, R2 = 0.15) included the LC CNR  but excluded the SN area (p = 0.757). A stepwise regression model for the stopping-related right STN activity  = 5.81, p = 0.023, R2 = 0.18) also included the LC CNR  but not the SN area (p = 0.466). HCs with better LC integrity showed higher stopping-related right IFG and right STN activity. However, no such correlation was found for PD patients (ps > 0.27).Second, LC (but not SN) integrity correlated with the stopping-related activity of the right IFG and right STN in HCs A. A stepF = 4.33, p = 0.024, R2 = 0.26) included the daily levodopa dose  and levodopa equivalent daily dose of D2/3 receptor agonists  but excluded the LC CNR and SN area (ps > 0.38). PD patients with daily exposure to more levodopa and fewer receptor agonists showed higher stopping-related preSMA activity.Third, daily doses of dopaminergic drugs correlated with the stopping-related preSMA activity in PD patients B. A step4Cognitive decline in PD correlates with the progression of α-synuclein pathology . Causal Damage to the LC may impact the right IFG-STN hyperdirect pathway during stopping. In PD patients B, the re4.1The STN is a strong candidate for inhibiting an initiated response within hundreds of milliseconds as the conduction velocity of ipsilateral cortico-subthalamic axons is ∼ 7 m/s, much faster than that of the cortico-striatal axons (∼1.5 m/s) . The STNBeyond the STN, we observed a complex regulation of response inhibition in frontal regions projecting along the hyperdirect pathway. In healthy adults and PD patients, higher stopping-related preSMA activity and lower stopping-related right IFG activity correlated with faster SSRT. This observation is in line with diffusion MRI findings that both right IFG-STN and preSMA-STN white matter tracts are engaged in response inhibition . HoweverIt remains unclear which fronto-subthalamic pathway is dominant or how the two fronto-subthalamic pathways interact during stopping. Recent human studies with invasive EEG and single-unit recordings have emphasized the importance of the right IFG-STN pathway . Chen etIn contrast, human imaging studies suggested that the right IFG modulates the excitatory projection from the preSMA to STN . Rae et 4.2Our novel finding is that greater LC integrity correlated with greater stopping-related right IFG and right STN activity in HCs. This finding thus fills an important gap in the literature. Human imaging studies have shown that the elevation of extracellular noradrenaline levels  can enhance stopping-related right IFG activity . Populat4.3SN integrity did not contribute to the preSMA-STN pathway . However, levodopa and D2/3 receptor agonists may impact the stopping-related preSMA activity in PD patients. PD patients with daily exposure to more levodopa and fewer D2/3 receptor agonists showed higher stopping-related preSMA activity, potentially leading to faster SSRT. This observation is consistent with previous findings that both D1 and D2 receptors can modulate SSRT in humans and animals , and tha5n = 58). Causal evidence is needed to confirm the model or to investigate hidden factors driving the correlations.First, LC and STN are small structures, but the spatial resolution of 3T MRI was limited. Further research with ultra-high field MRI or brain stimulation is needed to confirm our findings and determine the contribution of different LC and STN subregions . Second,6This study demonstrated that damage to the LC (but not SN) might impact the fronto-subthalamic hyperdirect pathway during stopping. In HCs, LC integrity correlated with the stopping-related right IFG and right STN activity, which further correlated with SSRT. PD patients showed reduced LC integrity, longer SSRT, and lower stopping-related activity over the right IFG and preSMA than HCs. In PD patients, the relationship between SSRT and the fronto-subthalamic pathway was preserved. However, LC integrity no longer correlated with the stopping-related right IFG or right STN activity. No contribution of SN integrity was found during stopping. LC might modulate inhibitory functions of the right IFG-STN pathway. Damage to the LC might impact the right IFG-STN pathway during stopping, leading to response disinhibition in PD.Biman Xu: Methodology, Investigation, Formal analysis, Writing – original draft. Tingting He: Investigation, Writing – review & editing. Yuan Lu: Investigation, Formal analysis, Writing – review & editing. Jia Jia: Investigation, Writing – review & editing. Barbara J. Sahakian: Writing – review & editing. Trevor W. Robbins: Writing – review & editing. Lirong Jin: Conceptualization, Resources, Writing – review & editing, Funding acquisition. Zheng Ye: Conceptualization, Methodology, Formal analysis, Writing – original draft, Funding acquisition.The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.


We propose a concept of terahertz waveguide plasmon amplifier based on a metal groove with active graphene. It is shown that the power amplification factor of the longitudinal-section magnetic (LSM) waveguide plasmon  near the cut-off frequency of this mode can exceed the amplification factor of the transverse magnetic (TM) plasmon in a layered graphene structure by more than four orders of magnitude for the same frequency. This is caused by the increase of the LSM plasmon wavelength near the cut-off frequency, smaller energy velocity of the LSM mode, and greater energy release from graphene for the LSM plasmon due to stronger lateral confinement of the LSM waveguide plasmon as compared to the TM plasmon in a layered graphene structure. We show that the enhancement of the LSM plasmon amplification factor near the cut-off frequency is a stronger effect than that due to screening of graphene. Creation of the inverse population of free charge carriers in graphene at THz frequencies3 as well as the development of tools for excitation of highly confined graphene plasmons1 resulted in possibility of compensation losses and amplification of THz electromagnetic field in active graphene structures giving rise to various concepts of THz graphene lasers5 and plasmonic amplifiers8. It is known that efficiency of the optical pumping of graphene is rather low because graphene absorbs less than 2.3% of the incident optical pump power9. A concept of the diffusion pumping of graphene allows for creating the free carrier population inversion in graphene with greater efficiency8. The diffusion pumping of graphene through a narrow-gap black-As substrate can prevent heating of graphene11. Creation of the population inversion in graphene via the electron and hole injection in graphene was studied in12. It was shown that electron–hole injection in graphene can substantially reduce the pumping threshold13. The possibility of plasmon amplification and necessary conditions for the plasmon lasing regime in graphene pumped by the electron–hole injection was discussed in14.Graphene is an atom thick monolayer of graphite possessing unique properties such as high mobility, linear energy spectrum of free charge carriers and zero energy band-gap15. The influence of embedded graphene layer on the guided modes of optical layered waveguides was investigated in16. Dispersion characteristics and damping of waveguide modes17, and the magnetoplasmon-polariton excitations18 in the layered structures with gated two-dimensional electron systems and graphene were studied theoretically.Propagation of THz plasmons in layered structures and metal waveguides with two-dimensional electron systems, particularly with graphene, has been actively studied theoretically and experimentally throughout the last decade. The effect of a graphene monolayer on the propagation of transverse electric modes in a rectangular metal waveguide was theoretically studied in6 and diffusion8 pumping was theoretically discussed. Amplification of THz plasmons in a pair of parallel active graphene layers was theoretically studied in19. We will show below in this paper that the amplification of plasmons in a waveguide with active graphene could be more efficient than that in the layered graphene structures due to strong transverse confinement of the plasmon field in a waveguide.Active guiding of THz surface plasmons in layered structure with inversely populated graphene with optical21 and experimentally realized22. The transition region between bound and leaky mode ranges in a metal groove waveguide was explored in23. Amplification of plasmons in waveguide structures with inversely populated graphene has not been studied so far.Open plasmonic waveguides are promising for transferring the electromagnetic signals in THz plasmonic integrated circuits. Waveguide plasmons are at an advantage because the field of such plasmons is fully confined in the plane transverse to the direction of the plasmon propagation. Propagation of the channel plasmons and their field structure in rectangular and triangular metal grooves were theoretically discussedd, placed in the bottom of a rectangular metal groove of width L and infinite depth. As a matter of fact, we assume that above graphene there is a hollow  graphene, schematically shown in Fig. 24 unlike the case of layered structures or hollow metal waveguides with transverse magnetic (TM) and transverse electric (TE) modes. We focus on the longitudinal-section magnetic (LSM) plasmon with the ac electric and magnetic fields 1, 2, and 3 corresponding to three different groove widths L = 1 μm, 2 μm, and 5 μm, respectively. Curve 4 in Fig. 18. Dispersion of the TM plasmon in a layered graphene structure and its increment were calculated using dispersion equation presented in25. One can see t hat, for large metal groove width, the LSM plasmon dispersion curve merges with the TM plasmon dispersion curve at high THz frequencies. This occurs because at high THz frequencies the influence of the side metal walls of the groove becomes negligible and the wavevector component transverse to the plasmon propagation direction becomes much smaller as compared to the longitudinal one: In a metal waveguide partially filled with dielectric, the eigenmodes of the waveguide split into the longitudinal-section magnetic (LSM) modes and the longitudinal-section electric (LSE) modesL as in Fig. In Fig. entclass1pt{minimaentclass1pt{minimaL as in Figs. Let us discuss the physical origins of significant increase of the LSM plasmon power amplification factor. First, the longitudinal component of the LSM plasmon wavevector entclass1pt{minima19, where 26We wonder which factors influence the increment of the modes. According to the energy conservation law, the spatial growths of the plasmon energy flux defined asAs can be seen from Eq. , the powL as in the previous figures in comparison with the energy velocity of the TM plasmon. One can see that the energy velocity of the TM plasmon monotonically increases with decreasing the plasmon frequency. The energy velocity of the LSM plasmon is smaller than the energy velocity of the TM plasmon and markedly decreases near the cut-off frequencies. Also note that the energy velocity of the LSM plasmon becomes smaller for smaller width of the groove. At the cut-off frequency, the plane wave oscillates strictly across the side walls of the groove waveguide being homogeneous  along the waveguide. As a result, the wave does not transfer the energy along the waveguide so that the energy velocity becomes zero in this case. In general, the cut-off frequency of a specific metallic waveguide mode is determined by the transverse wavevector of this mode24 which in turn depends on the width of the metal groove in our case.We calculated the energy velocities of the LSM plasmon presented in Fig. The ratio 25. Comparison the effect of vertical and lateral plasmon field confinement on the power increment of the LSM and TM plasmon modes is shown in Fig. d = 10 μm (curve 1) near its cut-off frequency is greater than the power increment of the TM plasmon in the layered graphene structure with a much closer metal screen placed at distance d = 0.01 μm from graphene (curve 3). It means that the enhancement of the LSM plasmon power increment near its cut-off frequency is a stronger effect as compared to that due to screening of graphene.It is known that graphene screening by a nearby metal plate (gate) enhances the amplification of THz plasmons as compared to the case of unscreened plasmons due to stronger vertical confinement of the screened plasmon field near graphene26. However, the effect of losses (gain) increases near the cut-off frequencies since the imaginary part of the wavevector become equal and can even exceed its real part (the latter tends to zero at the cut-off frequency). As a result, the group velocity loses definite physical meaning and senselessly differs from the energy velocity in this case. That is why we use only the energy velocity quantity in our paper.Note that the energy velocity introduced in our paper well coincides with the group velocity in the systems with small losses z axis) and y and z axes, where the first superscript indicates the direction along the y axis, the second index does for the z axis). We use the following boundary conditions. The components of the electric field tangential to the metal surfaces are set to zero at the walls and bottom of the groove waveguide: z axis) wavevector component , ε0 is the electric constant, 6:e is the elementary charge, T are the mean free time and temperature of the charge carriers in graphene, respectively, F and –EF for electrons and holes, respectively), τ. The remaining terms in Eq.  near the cut-off frequency of this mode can exceed that of the TM plasmon in a layered graphene structure by more than four orders of magnitude for the same frequency. This is caused by the increase of the LSM plasmon wavelength (up to tens of micrometer) and stronger interaction of the LSM plasmon with active graphene as compared to the TM plasmon due to smaller energy velocity of the LSM mode and greater energy release from graphene for the LSM plasmon near its cut-off frequency. The plasmon energy velocity exceeds 10


LINE-1 or L1) comprises about 17% of the human genome. L1 retrotransposition is known to cause genomic instability and related disorders, and resveratrol suppresses this retrotransposition; however, the underlying mechanism is still not elucidated. Recent observations showed that low-molecular-weight compounds might induce L1 retrotransposition through unknown mechanisms. This study aimed to determine polyphenol resveratrol (RV)’s effect on L1-RTP (retrotransposition) in somatic cells. Surprisingly, RV completely blocked L1-RTP. Experiments using the PPARα inhibitor GW6471 or siRNA-mediated PPARα depletion showed that RV-mediated L1-RTP’s inhibition depended on peroxisome proliferator-activated receptor α (PPARα). We demonstrated that RV inhibits p38 and cAMP response element binding protein phosphorylation, which are involved in MAPK signaling, and the L1-ORF1 protein’s chromatin recruitment. Furthermore, RV increased the expression of sirtuin-6 (SIRT6), which inhibited the activation of L1. The sirtuins family, SIRT1, SIRT6, and SIRT7, but not SIRT3, are involved in RV-mediated inhibition of L1-RTP. Overall, our findings suggest that RV directly modulates PPARα-mediated L1-RTP in somatic cells and that MAPK signaling interacts with SIRT6 closely and may play a role in preventing human diseases such as cancer.The retroelement long interspersed element-1 ( LINE-1 or L1), a highly active autonomous and non-long terminal repeat retrotransposon, is the most abundant endogenous retroelement in humans and accounts for approximately 17% of the human genome3. L1 can retrotranspose to new genomic loci via a “copy and paste” mechanism. Notably, the L1 element can retrotranspose itself as well as other retroelements, such as Alu and SVA (SINE-VNTR-Alu: short interspersed element-variable number tandem repeat-Alu). A single human cell contains more than 5 × 105 copies of L1, most of which are functionally inactive. However, 80–100 copies are capable of active retrotransposition. By analyzing the blood coagulation factor VIII gene in hemophilia A, it was found to be potentially capable of retrotransposistion in human genome4. Moreover, genetic mutation by L1 insertion has been reported, and a relationship with disease development is known. L1 insertions were reported in various human tumors, especially in cancer-related genes such as c-myc and APC in breast and colon carcinomas, respectively6. Recently, genome instability was a contributing factor to many age-related diseases. L1 drives IFN in senescent cells and developed age-related diseases7. L1-RTP is induced by disease development and environmental pollutants such as benzo[a]pyrene (B[a]P).Long interspersed element-1 (9. ORF2 is a multidomain protein with endonuclease and reverse transcriptase functions11. ORF1 and ORF2 perform L1-RTP (retrotransposition), which proceeds via three steps: transcription, reverse transcription, and insertion of the newly synthesized L1 DNA into the host genome via target-site primed reverse transcription13.The L1 genomic sequence is ~ 6 kb in length and contains two open reading frames (ORF1 and ORF2), with molecular masses of ~ 40 and 150 kDa, respectively. ORF1 is a basic protein with a nucleic acid chaperone activity and is present within cytoplasmic ribonucleoprotein complexes or stress granules in the cytoplasm14. We also found that L1-RTP is induced by 2-amino-1-methyl-6-phenylimidazopyridine (PhIP) and 2-amino-3,8-dimethyl-imidazoquinoxaline (MeIQx), heterocyclic amines (HCAs) formed during the cooking of red meat as food-borne carcinogens, as well as benzo[a]pyrene (B[a]P), environmental pollutants16. Moreover, we reported that L1-RTP was induced during two-stage skin carcinogenesis initiated by 7,12-dimethylbenz[a]anthracene (DMBA) and 12-O-tetradecanoylphorbol-13-acetate17 and abused drugs including methamphetamine, cocaine, morphine, and fentanyl citrate19. We suggested that many carcinogenic and substance abuse drugs induced L1-RTP and genome instability, but we did not identify any compounds that inhibited L1-RTP.We recently reported that 6-formylindolocarbazole (FICZ), a tryptophan photoproduct, acts as a potent inducer of L1-RTP20. Several studies have shown pleiotropic health benefits of RV, suggesting that it may prevent the progression of various age-related diseases23.Conversely, this study focuses on resveratrol (RV) , a polyphenolic phytoalexin abundant in several plants and is also found in grapes, pine, and red wine+-dependent deacylases that play a crucial role in transcription and L1 resistant factor24. It has been reported that the expression of SIRT6 is increased when cells are exposed to RV. SIRT6 knockout mice drive inflammation and show strong L1 activation, and cells display excess genomic instability25. Moreover, SIRT7 mediated chromatin form regulation, Lamin A/C, and repression of L1 element transcription level26. Collectively, we took particular note of the sirtuins family and assessed the mechanism of L1-RTP inhibition by RV. We investigated if L1-RTP inhibition involves PPARa and SIRT6, which interact with SNF2H, a component of the SWI chromatin remodeling complex. SNF2H is a member of the SWI/SNF chromatin remodeling complex family known to regulate target genes’ transcription by altering chromatin structure. SIRT6 and SNF2H are chromatin remodeling factors involved in repair during DNA damage, suggesting that they are also involved in the aging process.Our focus in this study was on peroxisome proliferator-activated receptor α (PPARα) as a ligand for RV, and sirtuins (SIRT) 6 are NADL1 retrotransposition also occurs in somatic cells28, in which deregulation of the process may trigger various disease states owing to gene deletions. RV has been reported to inhibit L1 methylation and RTP, but the mechanism of this is unclear29. Therefore, we analyzed the L1 repressive mechanism of RV using the L1-RTP inducers we have previously reported. Elucidating the mechanisms that inhibit L1 function in RV could contribute to the aging process and the extension of a healthy life span.These observations suggest that human somatic cells are susceptible to genome instability by L1-RTP, but it is unclear whether L1-RTP inhibits cellular activity by altering genomic stability or if external stimuli mechanistically induce it. However, other studies have provided evidence that 14. Briefly, HeLa cells were transfected with pCEP4/L1 mneol/ColE1  and selected for 2 days with hygromycin. RV for additional 2 days, cells were grown on neomycin for 3–4 weeks  colonies. As shown in Fig. L1-RTP was detected by a colony formation assay, as described15. We next examined the effect of RV on L1-RTP that is induced by these compounds. As shown in Fig. 30. The p38 pathway and a transcription factor, cAMP response element binding protein (CREB), are present downstream of MAPK31. We have previously reported that FICZ, MeIQx, and PhIP induce L1-RTP by phosphorylating the p38 pathway and CREB. Therefore, we analyzed whether the phosphorylation of the p38 pathway and CREB by FICZ, MeIQx, and PhIP was inhibited by RV. To confirm this, we performed western blotting (WB) analysis, showing that RV markedly inhibited the phosphorylation of p38 and CREB by FICZ, MeIQx and PhIP  gene family30. SIRT1 and SIRT7 exist in the nucleus and nucleolus, respectively, while SIRT3 exists in the mitochondria. To further demonstrate SIRT1, SIRT3, SIRT6, and SIRT7 dependency, we described that two different siRNAs sequences . DSBs formation is associated with its chromatin incorporation regulated by SIRT6 and SNF2H. To examine whether SNF2H is involved in RV inhibited L1-RTP, and to further demonstrate this SNF2H dependency, we found that two F2H Fig. b. These F2H Fig. c. SIRT6 F2H Fig. d. RV-indF2H Fig. e. These 34. To confirm this, we performed L1-RTP analysis after treatment with RV or N-acetyl cysteine (NAC), an antioxidant compound serving as a positive control, which was found to have a minimal effect on L1-RTP  a protein encoded by human immunodeficiency virus type-1 (HIV-1)41. We searched among the compounds that regulate L1-RTP but were unable to find compounds that inhibit L1-RTP. This study found that RV inhibited L1-RTP in HeLa cells and HuH-7 cells and investigated the molecular mechanism involved. We discovered that RV, a polyphenol with antioxidant properties, inhibited L1-RTP at concentrations ranging from 5 to 20 μM, which is one-half of the vasodilator effect of RV (30–40 μM)42. Under physiological conditions, RV is described as being present at the order of a few μM, so RV-based inhibition of L1-RTP could occur within an in vivo range.Recent studies have shown that the insertion of human L1 into genes occurs in the germline during early development, and in somatic cells such as cancer cells17. As these compounds exist within our surroundings, we are frequently exposed to them. RV can reportedly regulate L1 methylation29, whereas the mechanism by which RV regulates the metastatic potential of L1 remains elusive. Therefore, we investigated whether RV inhibits compounds that induce L1-RTP and clarified the analysis of the mechanism. We found that RV inhibited FICZ-, MeIQx-, and PhIP-induced L1-RTP in HeLa cells and HuH-7 cells. RV inhibited the phosphorylation of p38 and CREB by FICZ and HCA. Moreover, FICZ- or HCA-induced chromatin recruitment of ORF1 was blocked by RV. Taken together, our data suggested that RV inhibited L1-RTP by inhibiting p38 and CREB phosphorylation as well as by blocking the chromatin recruitment of the L1-ORF1 protein. Recently it was reported that L1 activity dependent on and integrated with cellular phosphorylation regulatory cascades significantly increased our understanding of interactions between L1 and the host44. We suggested that L1-RTP required MAPK phosphorylation and its downstream transcription factor in previous articles. Several articles reported that RV is a MAPK inhibitor; thus, RV inhibits the L1-RTP inducer. Collectively, L1-RTP required phosphorylation of ORF1 protein and downstream cascade factor or environmental transcription factor in host cells.We previously showed that environmental carcinogens, tryptophan photoproducts, and HCAs, originating from the cooking of red meat, induced L1-RTP45; thus, we suggested that RV first interacts with and binds to PPARα to initiate the L1-RTP inhibition step. PPARα inhibitors reportedly induce L1 activation37, suggesting that PPARα is involved in the pathway that induces L1-RTP.To determine the molecular mechanism of RV-mediated L1-RTP inhibition, we examined whether RV-based inhibition of L1-RTP depended on its antioxidant effect. The effect of RV was compared to that of NAC, which was found not to inhibit L1-RTP; thus, RV acts independently of its antioxidant effect. We further tested this using an inhibitor and siRNA-based depletion of the PPARα, the first target factor identified for RV. Interestingly, biochemical analysis revealed that RV inhibited L1-RTP in a PPARα-dependent manner. The 4’-hydroxyl group of resveratrol is critical for the direct activation of PPARα33. SIRT6 was found to repress L1 activity by ribosylating Kruppel-associated box (KRAB)-associated protein 1 (KAP1) of transcription inhibition factor. SIRT6 binds to the 5ʹ-UTR of the L1 promoter and ribosylates KAP125. The SIRT6-mediated KAP1 mono-ADP-ribosylation (MARylation) promotes the interaction between KAP1 and SIRT6, thereby promoting the recruitment to the L1 5ʹ-UTR24. L1-RTP is facilitated by the chemical modification of the Poly (ADP-ribose) Polymerase 2 recruitment of replication protein A to other DNA repair proteins. Moreover, RV was shown to increase SIRT genes’ expression. Here, we confirmed that RV increased SIRT6 protein expression. Combined with the finding that SIRT6 interacts with L1, this strongly suggests that it mediates inhibition of L1-RTP. The cell signal is PPARα as target factor and involves the regulatory function of SIRT6 and L1. SNF2H is involved in RV-mediated inhibition of L1-RTP, but SNF2H did not interact with ORF1 by IP-WB. We confirmed that expression of SIRT6 increased by RV is inhibited by PPARα and SNF2H siRNA.Many studies have recently identified potential restriction factors for L146. We suggested that RV inhibiting L1-RTP by two cell signaling pathways. First, RV directly modulated the PPARα receptor and SNF2H upregulated SIRT6 expression inhibited L1; second, ORF1 interacted with SIRT6 and PPARα blocked MAPK signaling , compounds that induce autoimmune diseases and cancer development. The effect of RV is essential from the perspective of preventive medicine—we are exposed to ultraviolet light and carcinogens in the environment regularly. We reported previously that the picomolar levels of FICZ (3 pM) could induce L1-RTP14; notably, about 8 pM FICZ is generated after 24-h exposure of tissue-culture medium to ordinary laboratory light47. Moreover, picomolar levels of HCAs induced L1-RTP, comparable to the PhIP concentration detected in human breast milk48. These data suggest that somatic cells are exposed to factors that promote genome instability in the typical life environment. As people age, genetic instability is stimulated by exposure to environmental and intrinsic factors. Activation of transposable elements occurs due to environmental factors and can extend to loss of neuronal function50.In our current study, we showed that RV directly modulates the PPARα receptor to inhibit L1-RTP. Furthermore, we suggest a new signaling pathway in which SIRT6 interacts with p38 in MAPK signaling, with RV inhibiting L1-RTP via suppression of MAPK signaling. Cea et al. reported that SIRT6 binds DNA damage sites and recruits and blocks MAPK signalinging Fig. . SIRT1 aing Fig. . These f51. RV can reverse this process in a SIRT1 dependent manner. SIRT7 mediated H3K18 deacetylation regulates the L1 expression and promotes L1 association with elements of the nuclear lamina26. As SIRT6 plays a crucial role in silencing L1 activity, SIRT6 activator as RV may prove effective at counteracting the age-related loss of L1 silencing. Our results demonstrate that SIRT1, SIRT6, and SIRT7 were associated with the L1 element and PPARα, so RV inhibited genome instability, including L1. RV appears to promote genomic stability, limiting the occurrence of genetic mutations.Sirtuins protect against many of the various aging-associated conditions, including skin damaged by UV radiation of SIRT1 in human keratinocytesIn this study, RV-mediated inhibition of L1-RTP depended on PPARα, SIRT1, 6, 7, and SNF2H. RV inhibits p38 and CREB phosphorylation, which are involved in MAPK signaling, and chromatin recruitment of the L1-ORF1 protein. Furthermore, RV increased the expression of SIRT6, which inhibited L1 activation. We suggest that RV directly modulates PPARα-mediated L1-RTP in somatic cells and that MAPK signaling closely interacts with SIRT6 preventing human diseases, such as cancer and sporadic cases of inborn errors.N-acetylcysteine (NAC) were procured from Wako Pure Chemical Industries, Ltd . Acetoxy-forms of HCAs  were synthesized . FICZ is a tryptophan photoproduct . GW6471 PPARα inhibitor was purchased from Wako . Neomycin (Wako) and hygromycin  were used after checking the cytotoxic effects in each cell line. Antibodies against PPARα , sirtuin 1 (SIRT1), sirtuin 3 (SIRT3), sirtuin 6 (SIRT6), sirtuin 7 (SIRT7), p38, phosphorylated p38, CREB, phosphorylated CREB, and SNF2H , H2AX (Millipore), enhanced green fluorescent protein , Anti-FLAG Tag M2 antibody , and glyceraldehyde 3-phosphate dehydrogenase (GAPDH)  were used as primary antibodies. A rabbit polyclonal antibody against human ORF1 was generated using the peptide MGKKQNRKTGNSKTQSAC as an immunogen 14. Anti-mouse and anti-rabbit secondary IgG conjugated to horseradish peroxidase were obtained from DAKO Japan .Protease and phosphatase inhibitors were purchased from Roche Diagnostics . Resveratrol and L1 retrotransposition was assayed as described previously53 using reporter vectors based on pCEP4/L1 mneoI/ColE1 (pL1-NeoR). The construct contained all components of human L1 in a single transcriptional unit, with a neomycin-resistance gene in the reverse orientation. When L1 retrotransposes, the intron within the neomycin  resistance gene is spliced out, resulting in a neomycin-resistant cell. Constructs were transfected into ~ 2.0 × 106 cells using Lipofectamine 2000 (Life Technologies), with selection for 3 days on 25 μg/mL hygromycin. Transformants were then transferred to 100-mm plates at ~ 1 × 105 per six plates, treated for an additional 2 days with the indicated amounts of chemicals, and finally grown on 300 (HeLa cell) or 250 (HuH-7 cell) μg/mL neomycin to select for cells in which retrotransposition occurred. After 2–3 weeks, colonies were stained with methylene blue and counted. Plating efficiency was examined to exclude cytotoxic effects of RV and observed that uM of RV. HeLa cells were treated for 2 days with RV at doses ranging from 200 to 1.25 μM, and 500 cells of each test group were plated onto 6-cm plates. After additional 7 days, cells were fixed and stained with methylene blue.19. Cells were washed with phosphate-buffered saline and resuspended in 50 mM Tris–HCl pH 7.6, 150 mM NaCl, 1 mM EDTA, 0.1% SDS, 0.5% deoxycholic acid, 1% NP-40, and protease inhibitors. Cells were ultrasonicated for 12.5 min  at medium power (250 W) at 4 °C with a Bioruptor , and soluble cellular extracts were recovered after centrifugation for 10 min at 16,000×g. Protein concentration was determined using the BCA Protein Assay Reagent Kit , and extracts were analyzed by WB. Blots were probed with primary antibodies, labeled with secondary antibodies conjugated to horseradish peroxidase, and visualized using Pierce Western Blotting Substrate Plus (Thermo Scientific). The original data obtained using WB and the result of adjusting the brightness using Photoshop are shown in the supplementary information.WB was analyzed as described previously15. We transfected 10 nM of siRNA using Lipofectamine RNAiMAX in HeLa cells and HuH-7 in 2.0 × 106 cells. The nucleotide sequences of each siRNA are shown in Table R to evaluate the inhibitory effects of the siRNAs on L1-RTP induction. Two days later, the cells were replated, incubated for 2 days with RV, and analyzed. As a control, Silencer Negative Control siRNAs  were used.Two small interfering RNAs (siRNAs) were prepared (Thermo Scientific), and their functions were evaluated by transfection into cells followed by WB analysis15, which encodes a chimeric protein of ORF1 protein A and the calmodulin-binding protein. On day 2, after transfection of pORF1-TAP into HeLa cells, FICZ or HCAs was added to the culture medium before RV, and cell extracts were prepared on the following day. The chromatin-enriched fraction (chromatin fraction) was isolated using the Subcellular Protein Fractionation Kit (Thermo Fisher Scientific) with micrococcal nuclease as described previously15. Detection of ORF1-TAP was performed by probing with a horseradish peroxidase-conjugated α-rabbit IgG antibody. H2AX served as a positive control for the chromatin fraction. Fractionation and detection of H2AX were evaluated using previously described methods15. A subcellular fractionation analysis revealed ORF1 after transfection of expression vectors encoding a chimeric ORF1-TAP protein.We used the pORF1-TAP (tandem affinity purification) construct15. HEK293T cells were transfected with each plasmid by Lipofectamine 2000. To express ORF1-EGFP, flag-tagged SIRT6 flag-tagged ovalbumin (OVA), and ST (streptavidin-tagged)-EGFP, we constructed pORF1-EGPF, pFlag-SIRT6, pFlag-OVA, and pST-EGFP. Cells were suspended in IP buffer composed of 20 mM Tris–HCl (pH 7.6), 5 mM EDTA, 150 mM NaCl, 0.5% NP-40, 10% glycerol. 500 μg of cell extract were reacted with 4 μg of αFlag or EGFP or αPPARα and then recovered with Dynabeads protein G beads (Thermo scientific). About one-tenth of each extract subjected to IP was loaded onto SDS-PAGE and simultaneously analyzed as an input sample. The immunohistochemical analysis was done by the reported method15. We used the TrueBlot  secondary antibody, which masks the detection of the heavy and light IgG chains, and thus, it displayed them as thin or undetectable bands.Immunoprecipitation (IP) assay was carried out, according to the reported methodU-test, with an n value of more than four. P-values of less than 0.05 were considered statistically significant.Statistical significance was evaluated using the Mann–Whitney Supplementary Figures.


A high-performance electro-optic Mach–Zehnder modulator (MZM) with outstanding characteristics is proposed. The MZM is in a push-pull configuration that is constructed using an ITO/graphene-based silicon waveguide. A novel idea for engineering of the plasma dispersion effect in an ITO/graphene-based waveguide is proposed so that the modulation characteristics of the MZM are highly improved. Plasma dispersion effects of ITO and graphene layers are tailored in such a way that a large difference between real parts of guided mode effective index of the two arms is achieved while their corresponding imaginary parts are equal. As a result, a very low  Optical modulators have already been investigated by many research groups20. The demand for high speed, low power consumption, high modulation depth and compact optical modulators still continues to support rapidly growing data in integrated optical systems.Optical modulators are the key components of photonic communication networks and processing systems. The input information of the modulators can be transferred to the amplitude, phase or polarization of the optical carrier through electro-optic22. However, pure silicon optical modulators are mainly based on the weak plasma dispersion effect, which hinders the realization of compact and integrated interconnections7. As a result, new materials and structures have already been explored to mitigate the drawbacks of silicon optical modulators. One of the outstanding materials that could revolutionize silicon photonics is graphene.Silicon photonics is a promising technology that has received tremendous interest for realizing efficient integrated optical devices such as modulators23. The superb optical characteristics of graphene have made it an amazing choice for various applications such as photodetectors25, logic gates27, polarizers28, plasmonic tweezers29, isolators30, mode-locked lasers32, switches and modulators37. The complementary metal-oxide-semiconductor (CMOS) compatibility and great tenability of graphene are of exceptional importance for optical modulators. In addition, high carrier mobility of more than 19. The modulator was performed by applying an external voltage across the graphene layers and changing the Fermi levels so that the absorption mechanisms were tuned. However, the modulation bandwidth was limited to 1 GHz. In 2016, a broadband graphene-based optical modulator was proposed where a modulation depth of 2 dB and insertion loss of 0.9 dB at a very high bandwidth of 35 GHz were reported38. However, the high bandwidth was achieved at the cost of high drive voltage of 25 V. It should be emphasized that most of the previously proposed graphene modulators have been based on the EA effects while the electro-refractive effect in graphene may also lead to high-performance modulators and phase shifters40.Graphene is a two-dimensional (2D) sheet of carbon atoms arranged in a honeycomb lattice that exhibits unique electrical, mechanical, thermal and optical properties43. The ITO-based EA modulators work within the α-dominant region of ITO permittivity where the optical loss is changed by modulating voltage. In contrast, the n-dominant region of ITO permittivity is of interest for phase shifters and electro-refractive modulators. However, according to Kramers–Kronig relations44, the real part variations of ITO refractive index is inevitably accompanied by the variations of optical loss α. Therefore, it is important to design electro-refractive modulators with low required voltage and simultaneously low insertion loss. In recent years, different kinds of electro-refractive modulators based on ITO have been studied. In 2018, an ITO-based Mach–Zehnder modulator (MZM) was proposed where a half-wave voltage and active device length product of 45. However, the switching speed was undesirably limited to 1 kHz. In 2018, another research group presented a lower 46. However, such a 47. It was experimentally shown that the 48.Indium tin oxide (ITO) as a transparent conductive oxide (TCO) is another material for next-generation optical modulators. The carrier concentration in ITO can be several orders of magnitude higher than silicon which leads to significant index modulation. Furthermore, the permittivity of ITO can be effectively tuned through an external bias voltage so that its real part can cross the zero point known as epsilon-near zero (ENZ). Various EA optical modulators using the ENZ behavior of ITO have been investigatedIn this paper, a very high performance ITO/graphene-based electro-optic MZM is proposed. The modulator is made of an engineered silicon waveguide based on two graphene sheets and two ITO layers as active materials whose characteristics are modified by an external voltage. The free-carrier dispersion effects in ITO and graphene are engineered so that large real part variations of guided mode effective indices in the two arms are obtained while their imaginary parts are the same. As a result, a 49. A graphene sheet is transferred to the hBN and an ITO layer is subsequently grown that are spaced by The perspective view of the proposed MZM is schematically shown in Fig. D) of aSi is firstly assumed to be 100 nm but it will be optimized in the following section. It should be emphasized that in practice graphene sheets cannot be laminated on sharp corners and on 90° waveguide edges practically. However, we first assume an ideal waveguide with rectangular cross section. The practical aspects of the modulator including the waveguide with trapezoidal cross section is investigated at the end of the paper.The total width and height of the ridge are respectively taken as As it is shown in Fig. In order to gain a better insight into the operation of the proposed waveguide, the effects of graphene sheets and ITOs on the guided mode should be investigated, separately. Hence, the stacks of the waveguide are theoretically separated to form the hypothetical waveguides shown in Fig. 50 and theoretical52 works where chemical potentials of 0.7 eV to 1 eV have been reported. However, our proposed modulator can also operate with lower voltages and chemical potentials. Furthermore, assuming 53, a breakdown voltage of 6.5 V is obtained for the In order to calculate the guided mode effective index of the waveguide shown in Fig. entclass1pt{minimaAs it is shown in Fig. According to Fig. As a general rule, a large real part change of the effective index is of interest for an electro-refractive modulator while the imaginary parts at the two states of modulator should be nearly equal. Therefore, as it is shown in Fig. The effective index for the guided mode of the structure shown in Fig. As it is shown in Fig. According to Fig. mentclass2pt{minimAs it is shown in Fig. d as Eq.  (see “Med as Eq. ”. When tAs it is shown in Fig. entclass1pt{minimaIn order to find the optimum The characteristics of the guided mode at the optimum initial doping and as a function of the applied voltage is plotted in Fig. D in Fig. D are calculated and shown in Fig. D leads to more confinement of guided mode and as a result higher light-matter interaction so that a larger phase shift is acquired. On the other side, as it is expected, the IL of the modulator increases from 2 dB at One of the geometrical parameters that can affect the modulator characteristics is the thickness of aSi (Another phenomenon that should be noticed is the electrostatic effect of the upper graphene sheet on the carrier concentration of the lower ITO layer as a consequence of thin aSi. As it is shown in Fig. The important point that should be taken into account is that it is not practically possible to place the stack of layers on the Si ridge with vertical side walls. Therefore, a trapezius with the side wall angle of It is instructive to investigate the effect of electrodes on the device operation. The metal electrodes are located at the two sides of the waveguide to apply the required external voltages. Such metal electrodes can possibly increase the IL of the device. The electrodes are assumed to be 1200 nm away from each other in our simulations. The guided modes of the proposed waveguide in the presence of the electrodes and without electrodes are calculated and respectively shown in Fig. Now, the operation of the modulator with the designed waveguide is investigated. The transmission spectra of the MZM in the on and off states are shown in Fig. For the sake of completeness, the effect of applied voltage errors on the performance of the modulator is studied. In this regard, the transmission spectra of the modulator in the off state for various pull voltage errors at Finally, other electrical characteristics of the proposed modulator are calculated. One of the most important parameters is the bandwidth of the modulator which is limited by the RC time constant of the modulator. The total capacitance of the proposed waveguide is the parallel combination of the two oxide capacitances (g to Eq.  (see “Meg to Eq. ”, the qu55, and a graphene-contact resistance of 56, the total ohmic resistance is obtained as The total resistance of the waveguide is also obtained by adding the graphene sheet resistance to the graphene- and ITO-contact resistances. Assuming a sheet resistance of mentclass2pt{minim61:62e is the electron charge, 61 and The simulation results in “63. Here, 43:The dielectric constant of ITO is calculated from the Drude model where the ITO carrier concentration is required. The induced carrier concentration in ITO is calculated according to:Then, the calculated chemical potential is applied to Eq.  and finaRegarding the analysis of the MZM, the transfer matrices of input Y-junction, the arms and output Y-junction are respectively expressed as:The output power of the MZM can be obtained by:In summary, we proposed a very high-performance electro-optic ITO/graphene-based MZM. According to the presented analysis, the contributions of the plasma dispersion effects in graphene and ITO layers are engineered so that a large difference between


Moreover, dimethylated arsenicals did not adsorb to BC at all, and in the case of FeBC, qm values of dimethylarsinic acid (DMA(V)), dimethylmonothioarsinic acid (DMMTA(V)), and dimethyldithioarsinic acid (DMDTA(V)) were calculated to be 7.08, 0.43, and 0.28 mg/g, respectively. This was due to the formation of iron oxide  on the surface of BC. Linear combination fitting using As K-edge X-ray absorption near edge structure spectra confirmed that all chemical forms of dimethylated arsenicals adsorbed on the two-line ferrihydrite were DMA(V). Thus, FeBC could retain highly mobile and toxic arsenicals such as DMMTA(V) and DMDTA(V)) in the environment, and transform them into DMA(V) with relatively low toxicity.In this study, the adsorption characteristics of dimethylated arsenicals to rice husk biochar (BC) and Fe/biochar composite (FeBC) were assessed through isothermal adsorption experiments and X-ray absorption spectroscopy analysis. The maximal adsorption capacities (q Daphnia magna were 9.5, <30, 1.7, and 6.5 mg/L, respectively  was 58 mg [Yoon et al.  studied as 58 mg , whilst An X-ray absorption near edge structure (XANES) analysis was performed to evaluate the chemical forms of dimethylated arsenicals adsorbed onto FeBC. As the main cause of the adsorption of dimethylated arsenicals onto FeBC was presumed to be the two-line ferrihydrite formed on the FeBC surface, the adsorption characteristics of dimethylated arsenicals on FeBC were studied by using the As K-edge XANES spectra of two-line ferrihydrite samples adsorbed with DMA(V), DMMTA(V), and DMDTA(V). In contrast, except for the two-line ferrihydrite adsorbed with As(V), the absorption energy of all samples adsorbed with DMA(V), DMMTA(V), and DMDTA(V) were the same as that of the reference compound . Daphnia magna was reported to be 9.5, <30, 1.7, and 6.5 mg/L, respectively [DMMTA(V) has been shown to exhibit high human- and eco-toxicity compared to DMA(V) and DMDTA(V). Cytotoxicity levels (IC50) of As(III), DMA(V), DMMTA(V), and DMDTA(V) in bladder cancer EJ-1 cells was reported to be 75 μM, ≥1 mM, 17 μM, and ≥1 mM, respectively , whilst ectively . In thisIn this study, the adsorption characteristics of As(V) and dimethylated arsenicals (DMA(V), DMMTA(V), and DMDTA(V)) on BC and FeBC were assessed. As BC was modified with two-line ferrihydrite, the maximum adsorption capacity of As(V) increased approximately 5-fold, from 1.28 to 6.32 mg/g. This may have been due to the Fe oxide modification, thereby increasing adsorption sites on the FeBC surface and decreasing the electrostatic repulsion. Dimethylated arsenicals did not adsorb onto BC, whereas on FeBC, DMA(V), DMMTA(V), and DMDTA(V) showed maximum adsorption capacities at 7.08, 0.43, and 0.28 mg/g, respectively. As such, dimethylated arsenicals were adsorbed through the reaction with two-line ferrihydrite presented on the surface of FeBC. Similar to observations in previous studies, adsorption inhibition was observed with an increase in thiol-group content. The XANES-LCF analysis confirmed that the main form of dimethylated arsenicals adsorbed to two-line ferrihydrite was DMA(V). This indicated that FeBC could be utilized to lower the mobility of dimethylated arsenicals in the environment and to transform them into less hazardous forms.