Functional fabrics have gained attention as an environmentally-friendly synthesis route. In the current study, novelty bamboo pulp fabrics with thermal conductivity properties were prepared by coating the fabric with graphene and cellulose nanocrystal (G/CNC) solutions. The influences of G and CNC concentrations on properties of fabrics were studied. The viscosities of the G/CNC solutions increased with an increase of G contents. G had an obvious thickening effect. Furthermore, compounded fabrics with different G and CNC contents (GCBPFs) were prepared and extensively characterized in terms of thermal and mechanical properties, and morphology. The ultimate thermal conductivity, bursting strength, and tensile strength of the GCBPF were 0.136 W/m·K, 1.514 MPa, and 25.8 MPa, with 4 wt.% CNC and 3 wt.% G contents, respectively. The results demonstrated that the as-fabricated GCBPFs with favorable thermal conductivity could be applied as a novel fast cooling textile for the clothing industry. Energy consumption was one of the major issues for humans in the 21st century . The conFirst efforts for personal thermal management techniques to regulate the thermal conductivity of fabrics focused on thickness , aerial Bamboo has emerged as the ultimate green, cost-efficient textile material that meets the definition of a renewable and sustainable raw material, with little harmful impact to the environment due to its fast growing speed  and abundant quantity in China ,14,15. C−1) [For fibrous textile materials, conduction, convection, and radiation are the three main routes to processing heat transfer, among which the most significant way is conduction . Therefo−1) . Coated −1) , polyeth−1) , sodium −1) , sodium −1) , and cel−1)  have beeIn this study, G material was dispersed in a cellulose nanocrystalline (CNC) aqueous solution to make a G/CNC solution. The solution was coated onto bamboo pulp fabrics to the compound thermal conductive textile by a dip-coating method. The thermal and mechanical properties of the G/CNC-coated bamboo pulp fabrics (GCBPFs) were evaluated by the thermal conductivity, tensile strength, bursting strength, viscosity, and crystallinity.Plain weave fabric made from 100% bamboo viscose  was purchased from Jilin Chemical Fiber Co. Ltd. . The bamboo fabric was bleached using a standard process to remove inherent impurities and additives. All treated fabrics were subjected to the same scoring and beaching process before coating.The multilayer graphene-based carbon nano-materials were provided by the Department of Sustainable Bio-products, Mississippi State University . The graphene nanoplates consisted of several sheets (with a range of 2–30 layers) of graphene with an overall thickness of approximately 1–10 nm . The morThe cellulose nano-crystalline (CNC) suspension was prepared through the sulfuric acid hydrolysis of microcrystalline celluloses . The obtThe prepared G/CNC solutions were applied to impregnate the bamboo pulp fabrics simply by a dip-coating method. The fabrics were first dipped into the G/CNC solutions and then padded through a rolling device to remove the excessive solutions. The above procedure was repeated for several times until the resistance of the fabric became constant. The obtained G/CNC coated bamboo pulp fabrics (GCBPFs) were dried at 45 °C for 2 h before being tested. For comparison, CNC suspensions coated fabric samples (CBPFs) were also prepared.−1. The surface morphologies of G, CNC, and fabrics were tested by Quanta 200 scanning electron microscope  and transmission electron microscope , respectively.For measurement of the viscosity of dipping solution before and after the addition of G, an NDJ-79 rotational viscometer was used with a rotational speed of 750 r·minThe X-ray diffraction (XRD) was performed using Cu kα radiation source  in an Ultima IV XRD instrument . The tensile strength of fabrics was determined according to ASTM D2256 by an HY-932CS micro-computer control electron universal testing machine, with a tensile rate of 1 mm/min. The bursting strength of samples was evaluated according to ASTM D3787-07 using a bursting strength tester . Five duplicates were repeated in each mechanical test group.T2 (mV/s), 2). A heat conduction coefficient tester  was employed to record the heat conductivity of samples. The test standard was according to GB/T 11048-2018. The thermal conductivity  for the pure CNC suspension and G/CNC solutions. The Cr of pure CNC suspension was calculated at 83%, and three concentration levels of CNC suspensions  had the same Cr values as shown in Cr values of the G/CNC solutions were lower than that of pure CNC, and decreased with an increase of G contents. The lower the CNC suspension concentrations, the faster the Cr values descend in range of G/CNC solutions with the increase of G contents. The lowest Cr value of the G/CNC solution was 19% with 3 wt.% G and 1 wt.% CNC. This phenomenon may be the continuous crystallization of CNC in the G/CNC solution which was organized by graphene sheets [Cr in the G/CNC dispersion. In addition, the interaction between graphene and CNC may also lead to the disappearance of the periodic interval between graphene layers, thus greatly improving the dispersion of graphene in CNC suspension [Cr of G/CNC solution. From the TEM images  also had the lower diffraction peak values than the CBPF one.The thermal conductivities of CBPFs ranged from 0.046 to 0.049 W/m·K tested by the thermal conductivity meter using a steady plate method as shown in The GCBPF sample with a higher CNC concentration of 4 wt.% had higher thermal conductivity with different graphene contents from 1 wt.% to 3 wt.%. Lower thermal conductivity for low contents of CNC might be due to situations such as the coating solution (to some extent) blocking the interstices and pores, and water droplets spreading in/on the fabric . FurtherCr (The results of tensile and elongation at break were plotted in Cr  and incrp < 0.05) with the addition of graphene from 1 wt.% to 3 wt.% compared with CPBF, with the values of TS in GCBPFs ranging from 18.2 MPa to 25.8 MPa. The larger decrease of TS in GCBPFs than CBPFs might be contribute to the addition of graphene destroying the complete self-assembly of CNC and the stronger electron scattering by graphene resulting in GCBPFs yielding brighter structures than CBPFs [A gradual increase in the tensile strength (TS) of CPBFs was observed (from 24.9 MPa to 29.0 MPa) in an CBPFs .p < 0.05). The ultimate strength of the GCPBF was up to 1.514 MPa, which was slightly lower than the CPBF (1.696 MPa). This indicates that GCPBFs inherited the excellent mechanical properties of CPBFs, and the dip coating treatment did not change the inner structure of fibers; the complex of graphene and hydroxyl groups in cellulose makes cellulose molecules cross-linked [The bursting strength (BS) of CPBFs and GCPBFs is shown in s-linked . MeanwhiThe structures of the fabrics and G/CNC-coated bamboo viscose fabrics (GCBPFs) were investigated by SEM as shown in In summary, the GCBPFs were synthesized by impregnating BPFs into the G/CNC solutions through a wet coating process in this study. The structure, and mechanical and thermal properties were studied subsequently. The graphene was found to uniformly coat bamboo pulp fiber surfaces due to the dispersion of CNC suspension, which enhanced the thermal and mechanical properties of the fabric at the same time. The GCBPF sample with a higher CNC concentration of 4 wt.% and G of 3 wt.% had excellent thermal conductivity and mechanical properties. The G/CNC coatings formed a firmly linked thermal conductive network, which made the originally insulating BPF thermally conductive. The results of this study indicate that the prepared GCBPFs have great application potentials for cooling textiles, conductive fabrics, wearable electronics, etc. Moreover, the durability of the G/CNC coating, i.e., the washing experiment, color fastness, and wear test, will be evaluated in order to validate the practical interest of this solution in our future work.


There are many epidemiological pieces of evidence that show IPF patients have the highest risk of lung cancer. We conducted a systematic review of all published data to define the characteristics of lung cancer that develops in IPF by performing a meta-analysis.2 were used. The review protocol has been registered in PROSPERO with ID: CRD42018094037.This study was performed based on the PRISMA guideline. Documents gathered by searching through the Web of Sciences, Scopus, PubMed/Medline, OVID, and COCHRANE databases which published before 03/25/2018 that related to lung cancer in IPFs’ patients. Articles were searched using standard keywords as well as Mesh and Mesh Entry and all probabilistic combinations of words using Boolean operators. Data searching, extracting and quality appraising were done by two researchers, independently. At last, Random-effects size based on Cochrane test and IBased on the meta-analysis conducted in 35 (0.18%) included studies, the total sample size of patients with IPF was estimated 131947 among whom 6384 had LC. The total rate of LC prevalence in IPF patients was estimated to be 13.54% (95% CI: 10.43–17.4) that was significantly 9 times higher in men vs. Women and smoker vs. non-smoker. Highest to lowest prevalence of cellular  subtypes of lung cancer in IPF were SQCC (37.82%), ADC (30.79%), SmCC (20.48%), LCC (5.21%), and ADQC (4.81%), respectively. The highest and lowest stage of lung cancer in IPF patients was estimated at III and II, respectively. The highest involvement location of lung cancer in IPF patients was in the Peripheral. Also, the prevalence of the tumor region involved from the highest to the lowest was estimated to be in the RLL, LLL, RUL and LUL regions.Lung cancer in IPF, most commonly SQCC, presents in elderly heavy smokers with a male, locating in peripheral regions and the lower part of lung predominance. Idiopathic Pulmonary Fibrosis (IPF), known as Cryptogenic Fibrosing Alveolitis (CFA), diffuse Fibrosing Alveolitis (diff FA), and Usual Interstitial Pneumonitis (UIP) is one of the most common forms of Interstitial Lung Disease (ILD) over the years –6. IPF iFamily history of disease 16, , 19, 20,Among radiographic features used in screening IPF patients, chest x-ray (CXR) can be mentioned, which the diagnosis and clinical manifestations may show bilateral interstitial opacities with the possibility of occurrence in peripheral and lower lung zones. This demonstration may not be visible in 2 to 10% of patients , 22. AlsAmong, lung cancer(LC), pulmonaThere are many epidemiological pieces of evidence that show IPF patients have the highest risk of lung cancer, 40, thaDespite previously published studies about the prevalence of LC in IPF patients in different countries (primary study) , 50–79, S1 File) including design and search strategy, a collection of articles and their systematic review, evaluation of inclusion and exclusion criteria, qualitative evaluation and statistical analysis of data. All steps were carried out by two researchers independently and, any encounters were assessed by a specialist. The review protocol has been registered in PROSPERO: International Prospective Register of Systematic Reviews (https://www.crd.york.ac.uk/PROSPERO/) Identifier: CRD42018094037 [The present study is based on the Meta-analysis of Observational Studies in Epidemiology guideline  and it h18094037 , 90(S2 FS1 Table) on each database without time limit until 03/25/2018. And, a manual search was also done as reviewing the reference list of related articles. The important point in searching the databases was the high-sensitivity searching, and also the search was conducted by the researchers and a specialist which is expert in searching databases (A.R).An advanced relevant search was conducted in international databases, such as Web of Sciences, Scopus, PubMed/Medline, OVID, and COCHRANE, to collect all of the studies which were related to LC in IPF patients. Articles were searched using standard keywords as well as Mesh and Mesh Entry and all probabilistic combinations of words using Boolean operators combined in accordance with the search syntax  Intervention: Surgical resection or radiological and pathologically confirmed cancer; (3) Comparison: That can show the rate of LC incidence in relation to non-LC in patients with IPF, which is called prevalence rate; (4) Outcome: Estimate the overall rate of LC in IPF patients and other risk factors.(1) (1) Review articles, Letters, Comments, Case reports , or Conference proceedings; (2) Studies that did not focus on the prevalence of LC in IPF patients; (3) Duplicated papers; (4) Non-English full text; (5) Non-accessible full text.All patients after initial diagnosis of IPF based on CT-Scan and pathological criteria by examining the dossier or attending the clinic, according to the American Thoracic Society (ATS) and the European Respiratory Society (ERS), 92, 93 To diagnose the LC in patients, after presuming IPF, the pathology and cytology reports were evaluated and categorized into two groups of patients with and without LC. Cellular  subtypes and clinical staging and region and location of LC were also evaluated in patients, 55, 96.After the end of the search, the papers were entered into the EndNote, reference management software and, after “Find References Updates”, duplicates were removed. After blinding studies , each study was evaluated by two researchers independently in the screening stage, with skimming and scanning the study titles, to evaluate the inclusion and exclusion criteria and IPF and LC detection criteria (the eligibility stage). In the event of disagreement between the two researchers, the specialist researcher made the final decision .After excluding irrelevant studies in the screening and eligibility stages, the quality of the final studies was examined. At this stage, the Newcastle-Ottawa Scale (NOS) checklisAt first, a checklist was designed according to the aims of the study and studying other available resources. Designed checklist includes items: Author name, Year, Place, Sample Size(SS), Periods of time, SS of IPF, SS of LC, Prevalence (LC in IPF), Cellular  Subtypes (ADC(Adenocarcinoma (%)), SQCC(Squamous-cell carcinoma (%)), SmCC), LCC(large-cell carcinoma (%)), ADSQC(Adeno-Squamous carcinoma (%)), Others), Clinical Staging , Prevalence separated by Sex, Prevalence separated by Smoking status, FEV1(Forced Expiratory Volume in the first second) (Mean±SD), FVC1 (Mean±SD), FEV1/FVC1 (Mean±SD), Age (Mean±SD), DLco(Diffusing capacity for carbon monoxide) (Mean±SD), Region (RUL(Right Upper Lobe), LUL(Left Upper Lobe), RLL(Right Lower Lobe), LLL(Left Lower Lobe), Upper, Center, Lower) and Location , which were extracted by two independent researchers and blind for the name of the author, institute and journal. In necessary cases, further information and raw data were requested by contacting the author (the first author or responsible or the authors' department).2 index were used. The I2 index less than 25% is low heterogeneity, between 75% -25% is the average heterogeneity and more than 75% are considered as heterogeneous[In each study, after considering the prevalence rate of LC in IPF patients as a binomial distribution probability, its variance was calculated by binomial distribution and for evaluating the heterogeneity of the studies Cochran test (Q) and Irogeneous, 98. AccIn this systematic study, based on performed searches, 667 articles were identified and after conclusive investigation and evaluation according to the checklist, 35 (0.18%) articles, 50–79SFig 1) w wFig 1) The overall prevalence rate of LC in patients with IPF was calculated at 13.54% : 10.43–17.4, P = 0.000). The highest and lowest prevalence rate was observed in Matsushita et al. (1995) from Japan with a prevalence rate of 48.2% (95% CI: 37.69–58.87) and Jeune et al. (2007) from the UK with a prevalence rate of 2.7% (95% CI: 1.88–3.86), respectively .Among the studies in 8 countries, the highest and lowest prevalence of LC in IPF patients was in Japan with 12 studies and Denmark with 1 study that were estimated 22.12% (95% CI: 11.35–38.64) and 5.79% (95% CI: 2.79–11.64) which were statistically significant (P = 0.000) .Sensitivity analysis of LC prevalence in IPF patients and the confidence interval of each study was calculated with a 95% confidence interval, and results showed that before and after exclusion of each study there had been no significant effect on the overall prevalence rate of LC in IPF patients . CumulatThe Meta-Regression test shows that the prevalence of LC in IPF patients has been decreased by the year of publication, but this relationship is not statistically significant (Mixed effects regression (Method of moments); Slope = -0.025), Intercept = 49.68), P = 0.451).The prevalence rate of LC in IPF patients with female gender was estimated to be 9.48% (77 out of 890) (95% CI: 7.49–11.94) and in the male gender 91.22% (813 out of 890) (95% CI: 88.72–93.2) that this difference  was statistically significant (P = 0.000) .The prevalence of smoking in LC-IPF patients were estimated to be 90.74% (95% CI: 87.07–93.44) .The prevalence of SQCC in IPF patients with LC was estimated 37.82% (95% CI: 32.65–43.29) , which tThe highest and lowest Stage of LC in IPF patients were estimated in III and II with a prevalence of 30.72% (95% CI: 22.68–40.14) and 13.33% (95% CI: 8.74–19.81), respectively (P = 0.000). The highest density was observed in Stage III and IV .The involvement location of LC in IPF patients was more pronounced in the peripheral area with a prevalence of 84.03% (95% CI: 75.47–89.99), which is statistically significant (P = 0.004) . The tumThe publication bias was also evaluated by Begg and Egger’s tests and was estimated at P = 0.516 and P = 0.0521, respectively. In this test, the probability of publication bias was not statistically significant .The total sample size of patients with IPF was estimated 131947 among whom 6384 had LC. The total rate of LC prevalence in IPF patients based on the meta-analysis review was estimated to be 13.54% (95% CI: 10.43–17.4) that was significantly 9 times higher in men than women. Also, the prevalence of smoking in LC-IPF patients is estimated to be 9 times higher. Highest to lowest prevalence of cellular  subtypes of lung cancer in IPF were SQCC (37.82%), ADC (30.79%), SmCC (20.48%), LCC (5.21%), and ADQC (4.81%), respectively. The highest and lowest stage of lung cancer in IPF patients was estimated at III and II, respectively. The highest involvement location of lung cancer in IPF patients was in the Peripheral region with highest to the lowest RLL, LLL, RUL and LUL regions respectively, that totally the highest to lowest was estimated to be in the lower, upper and middle regions.Age and smoking status are also known to be the effective factors in the development of lung cancer in IPF patients , 70. NeaRecent studies found that the progression of lung cancer in lower lobes is higher in IPF patients with, 100. Ascell[Evidence represent that IPF and LC have common pathological features. Anticancell–111. Alscell–105.In another study, epigenetic and genetic changes, abnormal expression of microRNAs , like different responses to regulatory signals, apoptosis, delay or decrease in cell-to-cell correlation along with activation of specific signal transmission pathways lead to pathogenic features of both LC-IPF. Likewise, genetic analysis has shown that harmful and deleterious mutations in A1 or A2-surfactant proteins cause familial idiopathic interstitial pneumonia and lung cancer–114.2) in the present study was calculated at 90.71%, which is in the line of studies with high heterogeneity. It is assumed that the observed differences are due to various samplings and also the difference in the measured parameter in different societies[The heterogeneity rate  by the publication year. Even though the studies are in different countries and in years, but these findings cannot represent the reality in all countries, so further studies are needed to be conducted in this regard.One of the main limitations of this meta-analysis to be mentioned is the inclusion of studies with different inclusion and exclusion criteria, and there is no consensus definition of IPF expressed. Selection Bias is more discussed which can limit the generalization of these findings because the type of lung cancer in a country can be different with the other countries and could be related to descent diversities. Due to lack of resources and very few studies have investigated the survival rate and causes of mortality and the precise methods of treatment, according to the aim of the present study, we did not focus on these factors. Also, data were accessed by using Guilan University of Medical Sciences’ -Iran Ministry of Health & Medical Education- VPN which some databases are not fully accessible.In conclusion, the high prevalence of the LC-IPF with 13.5% is more observed in older men who smoke, and is more evident in the progression of cancer, SQCC, and SmCC, and mostly affects the peripheral regions and the lower part of the lung. Studies have been conducted in limited countries, such as Japan, Korea, and UK and USA, which the weakness of a unit study of LC-IPF in different countries investigating the factors and important risk factors and reaching to a consensus and preparing a comprehensive global database for clinical decision-making is felt and is an essential need.S1 File(DOC)Click here for additional data file.S2 File(PDF)Click here for additional data file.S3 File(PDF)Click here for additional data file.S1 Table(DOCX)Click here for additional data file.S2 Table(XLSX)Click here for additional data file.S1 Fig(TIF)Click here for additional data file.S2 Fig(TIF)Click here for additional data file.S3 Fig(TIF)Click here for additional data file.S4 Fig(TIF)Click here for additional data file.S5 Fig(TIF)Click here for additional data file.S6 Fig(TIF)Click here for additional data file.S7 Fig(TIF)Click here for additional data file.S8 Fig(TIF)Click here for additional data file.S9 Fig(TIF)Click here for additional data file.S10 Fig(TIF)Click here for additional data file.S11 Fig(TIF)Click here for additional data file.S12 Fig(TIF)Click here for additional data file.S13 Fig(TIF)Click here for additional data file.S14 Fig(TIF)Click here for additional data file.S15 Fig(TIF)Click here for additional data file.S16 Fig(TIF)Click here for additional data file.


Based on the structural characteristics of the anodes of lithium-ion batteries, an improved Hummers’ method is proposed to recycle the anode materials of spent lithium-ion batteries into graphene. In order to effectively separate the active material from the copper foil, water was selected as an ultrasonic solvent in this experiment. In order to further verify whether lithium ions exist in the active material, carbon powder, it was digested by microwave digestion. ICP-AES was then used to analyse the solution. It was found that lithium ions were almost non-existent in the carbon powder. In order to further increase the added value of the active material, graphene oxide was obtained by an improved Hummers’ method using the carbon powder. The graphene material was also reduced by adding vitamin C as a reducing agent through a chemical reduction method using graphene oxide. Meanwhile, the negative graphite, graphite oxide and graphene samples were characterized by XRD, SEM, FTIR and TEM. The conductivity of the negative graphite, graphite oxide and graphene was tested. The results show that graphene prepared by a redox method has a better layered structure, less impurities and oxygen groups in its molecular structure, wider interlayer spacing and smaller resistivity. Because silicon-based anodes have huge reserves and super high theoretical capacity (4200 mAh g−1 for Li4.4Si), which is approximately 10 times higher than the state-of-the-art graphitic anode (372 mAh g−1 for LiC6), Silicon (Si) has been considered as one of the most promising anode material for the next generation lithium-ion batteries (LIBs)6. With the continuous development and application of silicon-based lithium-ion batteries, all-solid-state lithium-ion batteries, aluminum-air batteries and other batteries, the output of batteries has increased year after year10. In addition, the total quantity and weight of these exhausted batteries in China will exceed 25 billion units and 500,000 tons in 2020, as surveyed by Zeng et al.11. The recovery and treatment of spent lithium-ion batteries mainly concentrate on the separation, recovery and refining of cobalt and lithium as cathode and precious metals. However, little research has been done on the separation, recovery and recycling of anode active materials13. Spent lithium-ion battery anodes that have not been reasonably recycled lead to the release of pollutants into the natural environment, which may threaten human health. However, the carbon powder adhering to copper foil in the anode electrode of spent lithium-ion batteries has very large utilization potential and high recovery value. Therefore, the effective separation of the anode electrode components of spent lithium-ion batteries would play an important role in maximizing their resource utilization and eliminating the corresponding impact on the environment15. In order to alleviate the serious resource shortage and environmental pollution caused by rapid economic development, it has become a global consensus to recycle all components of spent lithium-ion batteries.Lithium-ion batteries have become ideal energy sources in the 21st century due to their lightweight, small volume, high specific energy, small self-discharge and long cycle life17. The second method based on a thermal treatment of graphite to recycle and reuse as high-performance cathode18. And the final investigated approach uses a liquid carbon dioxide or supercritical carbon dioxide as extractant to recycle electrolyte19.The recovery of the anodes of spent lithium-ion batteries is mainly concentrated on copper and carbon powder. Copper (~35%) separated from the anodes of spent lithium-ion batteries can be reused directly. It is an important raw material widely used in production. However, most of the carbon powder adhering to it (~60% content) has been abandoned and has not been rationally utilized. Usually, there are also some methods concentrating on the recovery of anode electrodes in combination with three different concepts. The first method is based on a separation or leaching process in order to recovery copper foil and lithium23, pyrometallurgy24 and mechanical physics26. However, most of these methods are complex, time-consuming and expensive to operate. The redox method forconverting carbon powder into graphene proposed in this study is expected to be a more suitable alternative. The method has the mainadvantages of simple operation, low costand high yield. Furthermore, themethod also shows that the great potential ofrecycling spent battery anode materials.In recent years, several traditional methods for recycling spent lithium-ion batteries have been developed, including hydrometallurgy27, high elastic modulus, low thermal expansion coefficient and high thermal fatigue and creep resistance are considered to be revolutionary for materials in the future30.Graphene is a single layer of carbon atom surface material, which is stripped from graphite. It is a two-dimensional structure of carbon and is deemed a “super material”. Its hardness is greater than that of diamond, and it has the same elasticityas rubber. Its electric and thermal conductivity are higher than copper, and its weight is almost negligible compared with copper. A crystalline film of graphene is only 0.335 nm thick. The discovery of graphene was a landmark in nanotechnology. Graphene has excellent properties in optics, electricity, mechanics, thermology and other fields. Graphene has many advantages in nano-electronic devices, highly sensitive sensors, transparent conductive films, functional composites, energy storage, catalysis and other fields. Its unique atomic structure, high electrical conductivity31, chemical vapor deposition32, epitaxial growth33 and oxidation-reduction34. Through consulting a large number of literature studies, it is found that the graphene oxide reduction method is easier to operate than the other methods. The production cost is greatly reduced, graphene can be prepared on a large scale, and functional graphene oxide can be prepared while graphene is prepared. Most carbon powders in the anode electrodes of spent lithium-ion batteries have been discarded, and have not been rationally recycled. This not only wastes carbon resources, but also causes environmental pollution. Here, for the first time, a new method is proposed to prepare graphene using an oxidation-reduction method to recover the anode material of spent lithium-ion batteries. The structures of the products were analyzed by X-ray diffraction (XRD), scanning electron microscopy (SEM), transmission electron microscopy (TEM) and Fourier transform infrared spectroscopy (FTIR). This research not only focuses on the serious problems of resource shortages and environmental pollution caused by rapid economic development, but also maximizes the resource utilization of spent lithium-ion batteries.Graphene has unique physical and chemical properties. The main preparation methods are mechanical strippingThe spent lithium-ion batteries needed for the experiment were collected from various places, and the rated capacity ranged from 550 to 1250 mA·H. The spent lithium-ion batteries were mechanically disassembled and the cathode, anode and diaphragm of the batteries were separated. The anode material was collected and cut it into 1 cm × 5 cm pieces for use. Concentrated phosphoric acid, hydrochloric acid (5%), N-methyl-2-pyrrolidone (NMP) and acetone were purchased from the Tianda Chemical Reagent Factory . Ethylene glycol, concentrated sulfuric acid, potassium permanganate, ammonia water, hydrogen peroxide and vitamin C were purchased from the Shanghai National Pharmaceutical Reagent Factory . All the above reagents were analytically pure in this experiment. An ultrapure water system  was used to produce the ultrasonic solvent with a specific conductivity of 0.055 μS/cm.First, the packaging and shell of the spent lithium-ion batteries were peeled off by means of mechanical tools, and the cores were taken out. The cathode and anode materials were then separated. The anode material was cut into small pieces of 1 cm × 5 cm, and the carbon in the anode material was separated from the copper foil by an ultrasonic leaching method. Ultrasound leaching is an environmentally-friendly and efficient method for the separation of copper and carbon powder, because it does not require chemical reagents, only uses water and has low energy consumption. A quantity of 0.25 g of anode material was weighed and put it in an electrothermal drying chamber at a constant temperature of 120 °C for 1 h. After cooling, the cathode material and copper foil were separated completely at normal room temperature after 2 min of ultrasound treatment. The separated copper foil and carbon powder were dried in an electrothermal drying oven at a constant temperature of 120 °C for 1 h. The carbon powder content in the anode material was calculated as 77%. The separation efficiency of the active substances in the anode material was calculated as follows (1):−1 for 3 min. The lower solids of the centrifugal tube were taken, and the products were washed with hydrochloric acid (5% volume fraction) and distilled water until the pH was ~7. Finally, the obtained graphite oxide was dispersed in water, then the graphite oxide was ultrasonically cleaned for 8 h with an ultrasonic cleaner, placed in a vacuum drying box and dried for later use.Concentrated sulfuric acid and phosphoric acid with a volume ratio of 9:1 and 1 g of carbon powder were put into a small beaker, and then 6 g of potassium permanganate was added in succession to prevent a violent reaction. The solution was stirred in an iced water bath for 1 h, then placed into a small beaker, whereupon the solution was stirred and maintained at a temperature of 50 °C and reacted at this temperature for 12 h. Iced water was added to the product, and hydrogen peroxide (30% volume fraction) was added dropwise while stirring until the color of the solution turned golden. Then, the centrifugation was started, and the centrifugation time was 4 000 r minA stable graphite oxide dispersion solution can be obtained by dispersing 10 mg of graphite oxide in a 25 ml aqueous solution, and then ultrasonically dispersing for 1 h until it becomes a brown suspension. Graphene oxide dispersions were moved into a 100 ml beaker and ammonia water was added to the beaker to make the pH ~8. After that, 250 mg of vitamin C was added to the reaction solution, stirred for 0.5 h, then the homogeneous solution was placed in a magnetic stirrer and stirred at 90 °C for 1 h. Next, the stable black dispersions were dried in a vacuum drying chamber at 60 °C for 48 h, and the graphene samples were finally obtained. After the sample was fully dried, the agate mortar was used to grind the powder for reserve. As shown in Fig. −1. Diffraction patterns were collected using an Advance D8 X-ray diffractometer with a scanning range of 10–80°. The appearance and internal morphology of the samples were analyzed with a Quanta 200 scanning electron microscope and Hitachi HT7700 transmission electron microscope. The resistivity of the samples was measured by a RTS-9 four-probe tester.The samples were analyzed by a Tensor 27 Fourier transform infrared spectrometer using the KBr pressing method, with a scanning wavenumber ranging from 400 to 4000 cm2O, acetone and ethaneiol were used as ultrasonic solvents, which could be used to separate the anode materials efficiently during the course of the experiment. Table In order to obtain the optimal conditions for the separation of anode materials, NMP, H−1, which belongs to the stretching vibration absorption peak of the C=C bond in the sp2 structure of the graphite crystal35. In the high frequency region at ~3430 cm−1, it belongs to the stretching vibration of OH. The wider spectrum peaks in the range of 3000–3700 cm−1 comes from the water molecules  were grafted onto the surface of the graphene sheet, and the resistivity increased. However, after vitamin C reduction, a large number of oxygen-containing groups on the graphene surface were removed, and the sheets were opened in a large area. The structure of graphene was restored and the conductivity was improved. The resistivity of acetylene black (d = 1 cm), graphite oxide (d = 1 cm) and graphene (d = 1 cm) were measured by a four-probe tester. Three areas were selected for the surface measurement of the sample film, as shown in Table The potential value of spent lithium-ion battery anode materials is constantly being explored. In this experiment, the ultrasonic leaching method was used. After optimizing the experimental conditions, water was chosen as the ultrasonic solvent to recover carbon powder from the negative material of spent lithium-ion batteries. The improved Hummers’ method was used to convert carbon powder into graphite oxide. Finally, graphene was prepared with vitamin C as a reducing agent. XRD analysis showed that most of the oxygen-containing functional groups of graphene were removed by a reduction reaction. FTIR characterization showed that a large number of oxygen-containing groups on the surface of graphene were removed, the structure of graphene was restored, and the reducibility of vitamin C was very good. Graphene was observed by SEM and TEM. The obvious layered structure of graphene was observed, which proved that graphene was obtained. The resistivity of graphene was measured by a four-probe tester. It was shown that the oxygen-containing groups on the surface of graphene were largely removed and the structure of graphene was restored. Moreover, the whole process cost is very low, which provides a new method and way for the recovery and utilization of spent lithium-ion batteries.


Recent advances in MRI have made it easier to collect data for studying human structural and functional connectivity networks. Computational methods can reveal complex spatiotemporal dynamics of the human developing brain. In this paper, we propose a Developmental Meta-network Decomposition (DMD) method to decompose a series of developmental networks into a set of Developmental Meta-networks (DMs), which reveal the underlying changes in connectivity over development. DMD circumvents the limitations of traditional static network decomposition methods by providing a novel exploratory approach to capture the spatiotemporal dynamics of developmental networks. We apply this method to structural correlation networks of cortical thickness across subjects at 3–20 years of age, and identify four DMs that smoothly evolve over three stages, i.e., 3–6, 7–12, and 13–20 years of age. We analyze and highlight the characteristic connections of each DM in relation to brain development. Understanding normal brain development is critical for basic developmental neuroscience. Recent advances in MRI have made it easier to collect data for studying human structural and functional connectivity networks. For example, the multi-site NIH MRI Study of Normal Brain Development has collected developmental structural MRI of over a thousand of children, ranging from infancy to young adulthood. The rich spatial and temporal information afforded in such datasets calls for sophisticated computational methods to capture the dynamic changes in brain connectivity during the course of development.To date, most developmental studies are mainly focused on charting network characteristics, such as small-world properties, global efficiency, local efficiency, and etc. Menon, . They trdevelopmental meta-networks (DMs) and propose an approach for Developmental Meta-network Decomposition (DMD). DMD decomposes the developmental networks into a set of temporally smooth DMs that capture the underlying connectivity patterns across developmental stages to adapt to the cognitive development. DMD not only automatically identifies the developmental stages and the number of DMs, but also uncovers the evolution of DMs. In this study, we apply DMD to developmental networks constructed based on inter-regional cortical thickness correlation across subjects for age spanning 3–20 years.To study temporal changes in brain connectivity, various matrix decomposition methods, especially principal component analysis (PCA), independent component analysis (ICA), and non-negative matrix factorization (NMF) have been applied to developmental structural and functional networks to identify the intrinsic components , τ is the number of time points. Assume that the τ developmental networks consist of G developmental stages, let t ∈ ) denote the tth developmental stage, then X can be reorganized as X = [X1X2 ⋯ XG]. Our aim is to decompose a set of DMs (denoted as U) and their developmental trajectories (denoted as V) from X. Note that we allow each DM to evolve across different stages, instead of remaining unchanged through development.Let Xt. It is an observation of the tth developmental stage that consists of two parts. One is the true developmental networks of the tth stage (Xt. We assume that p DMs at the tth stage (Ut), whose contributions are indicated by their trajectories (Vt), i.e., We start from considering the decomposition of a single developmental stage Xt, the second term penalizes the nuclear norm of UtTUt = I avoids finding overlapped connectivity patterns. Besides, the non-negativity constraints on Ut and Vt facilitate their neurobiological interpretation as DMs and developmental trajectories. To simplify the formulation, we replace Ut and VtT in Equation (1) and have the following objective function.The first term in Equation (1) minimizes the difference between Next, we take all the developmental stages into consideration. According to existing neural network models, nervous systems can change smoothly by slowly changing connectivity patterns and strength  indicates the adjacency between the kth and lth developmental stages. The second term of V across different time points, where WV measures the adjacency between the ith and jth time points.In Equation (4), the first term of Since it has been proved that minimizing the nuclear norm of the product of two non-negative matrices is equivalent to minimizing half of their squared Frobenius norm  times with random initializations and choosing the one with the least cost. The empirical convergence of DMD method is shown on Figure where The number of DMs is determined by maximizing Minimal Trajectory Distance (MTD), which computes the minimal distance between pairwise developmental trajectories. The larger MTD, The more separable the developmental trajectories, the more distinct and biologically significant the corresponding DMs.stable connections and rapidly-changing connections, for analysis.On the basis of developmental meta-network decomposition, we characterize two types of connections in DMs, i.e., stable connections constitute the “bones” of DMs that keep unchanged through development, while their contributions in the developmental networks are indicated by the developmental trajectories. We select the stable connections with the least normalized divergence  for each DM, and exclude those insignificant connections with average strength (r). Take the rth DM (denoted as Ur) for example,The Ur) in Equation (8) computes the normalized divergence of the connections in the the rth DM, where tth state of Ur. The threshold δr is adaptively determined by the quadratic mean  of Ur, where GM is the number of connections in Ur.Ω and even-numbered ages . The reproducibility is quantified by computing similarity between the corresponding DMs , which are independently decomposed from the two splits and matched using the Hungarian algorithm Kuhn, . We use Parameter Influence.We apply DMD method to the developmental structural correlation networks and obtain four DMs, which evolve across three identified developmental stages, including ages 3–6, 7–12, and 13–20, respectively Figure . For theindirect connection pattern, which looks like “<>,” between the prefrontal and occipital areas through interlaying temporal regions. The trajectory of the first DM shows that the indirect connection pattern decreases quickly at first but then slows down after the age of 13. In comparison, the stable connections of the second and third DMs are characterized with the long direct connections between the prefrontal and occipital areas. The trajectories of these direct connections, after going through ups and downs, reach higher values at the age of 20 than at the age of 3 years. Combined together, they indicate the gradual replacement of the indirect connection pattern by the direct connection pattern from age 3 to 20 years. This is consistent with the development of functional network architecture, where regional interactions change from being predominantly anatomically local in children to interactions spanning longer cortical distances in young adults  between the prefrontal and temporal regions  and MOG.L . Since IFGoperc.L play a role in processing word phonology  in the first language acquisition  to the third (ages 13–20), the first DM is still featured with decreased connection strength and decreased stage contribution Figure . Its rapTo summarize, we have three major findings from the characteristic connections of DMs, which are validated in Figure indirect connections between the prefrontal and occipital regions gradually, to some extent, get replaced by the direct connections from 3 to 20 years of age.The emotion-related regions  are significantly increased from 6 to 12 years of age.The connections with the language-related regions (IFGoperc.L and IFGoperc.R) are significantly decreased from 13 to 20 years of age.The connections with the G), the developmental stage partition at first keeps very robust in consensus clustering, but then exhibits growing instability when G becomes larger than 3. Therefore, in this study we choose the most robust developmental stage partition in the finest granularity, i.e., three stages including 3–6, 7–12, and 13–20 years of age. Note that although we do not utilize any temporal information, the developmental stage partitions still group the networks at adjacent time points together, indicating the smooth change of the developmental networks.We first study the influence of stage number on the quality of the developmental stage partition Figure . With thNext we show the influence of the DM number on the separability of developmental trajectories Figure . With thWe also examine the reproducibility of DMs and their developmental trajectories under different settings of DM number Figure . The repWe propose the DMD method to decompose developmental networks into a set of DMs and their developmental trajectories. One important advantage of our approach is that it goes beyond current static network decomposition strategies to capture dynamic temporal patterns of developmental networks. Moreover, it automatically identifies the partition of developmental stages as well as the number of DMs. Our analysis of normal developmental brain networks shows that the decomposed DMs contain not only stable connections but also rapidly-changing connections. Capturing variability in the connectivity patterns via DMs moves us closer to the clinic of the future.DMD helps to find significant underlying changes in connectivity over development. For instance, the rapidly-changing connections of DMs reveal that (1) the connections with the emotion-related regions  are significantly increased after 7 years of age, and (2) the connections with the language-related regions  are significantly decreased after 13 years of age. These findings are consistent with the existing behavioral studies that emotion control gets enhanced at the age of 7 years  or Functional MRI (fMRI), the main challenge lies in that different sets of DMs will be derived from different within-subject longitudinal networks. Therefore, it is necessary to extend DMD to decompose a representative set of DMs from multiple within-subject longitudinal networks. Then the representative set of DMs can be applied to single subjects for early disease diagnosis. We believe that DMD method will ultimately pave the way to much more refined representation and understanding of brain network development, diseases, and other longitudinal biological phenomena.This study was carried out in accordance with the recommendations of Name of Guidelines, Name of Committee with written informed consent from all subjects. All subjects gave written informed consent in accordance with the Declaration of Helsinki. The protocol was approved by the Name of Committee.PH contributed to designing and analyzing the experiments, summarizing and visualizing the results, and writing the manuscript. XX contributed to developing and implementing the method, visualizing and analyzing the results. HZ, GL, and P-TY contributed to the critical revision of the manuscript and the discussion of the work. JN contributed to preparing the data and revising the manuscript. DS contributed to directing the whole project.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


TDP-43 accumulates in nerve cells of nearly all cases of amyotrophic lateral sclerosis  and in the majority of Tau-negative frontotemporal lobar degeneration (FTLD). There is currently no biochemical test or marker of disease activity for ALS or FTLD, and the clinical diagnosis depends on the opinion of an experienced neurologist. TDP-43 has a key role in the pathogenesis of ALS/FTLD. Measuring TDP-43 in easily accessible biofluids, such as blood or cerebrospinal fluid, might reduce diagnostic delay and offer a readout for use in future drug trials. However, attempts at measuring disease-specific forms of TDP-43 in peripheral biofluids of ALS and FTLD patients have not yielded consistent results, and only some of the pathological biochemical features of TDP-43 found in human brain tissue have been detected in clinical biofluids to date. Reflecting on the molecular pathology of TDP-43, this review provides a critical overview on biofluid studies and future directions to develop a TDP-43-based clinical biomarker for ALS and FTLD. TDP-43 has been identified as binding to the transactivation-responsive region of the HIV-1 gene, acting as transcriptional regulator . AmyotroCytoplasmic aggregation of ubiquitinated, phosphorylated, and truncated TDP-43 is a unifying pathologic observation across the clinical spectrum of ALS and FTLD , 7. TDP-Under physiological conditions, TDP-43 is a mainly nuclear protein. In most cases of ALS and FTLD, it is found translocated to the cytoplasm where it changes formation and forms aggregates Fig. . WhetherTARDBP) gene in hereditary forms of ALS, clinically and pathologically identical to sporadic cases [TARDBP mutations linked to FTD without ALS have been described [Support for a direct mechanistic link between TDP-43 and neurodegeneration came from the identification of mutations in the TAR-DNA binding protein  and additional C-terminal fragments at 18–24 kDa, which were not detected in control brains. By extraction of those phosphorylation-dependent bands, subsequent mass spectrometric analysis revealed several intrinsically cleaved peptides, N- and C-terminal peptides, and modifications of sarkosyl-insoluble TDP-43. Almost all modifications were localized in the Gly-rich C-terminal half. Importantly, several novel phosphorylation, deamidation, and cleavage sites were identified by this approach. Common modifications focused on the 180–330 residue region, suggesting that this region of TDP-43 is affected in disease [Cell-specific differences in TDP-43 distribution and in the propensity to form inclusions within regions affected by early TDP-43 pathology have been reported. In α-motoneurons of the spinal cord, loss of normal nuclear TDP-43 is always accompanied by the formation of cytoplasmic TDP-43 deposits, either granular or compact which form phosphorylated skein-like TDP-43 inclusion bodies in advanced stages. While in large Betz cells of the motor cortex, only sparse cytoplasmic deposits can be found, which are not phosphorylated . This fiThe distribution of the pathological isoforms of TDP-43 has been well established from human postmortem tissue. All TDP-43 antibodies applied in biofluid studies so far bind to aa1–260, 205–222, or 256–296 of TDP-43 . While iDisease-specific biomarkers have been highly successful in other neurodegenerative diseases such as Alzheimer’s disease, where measurement of neuron-specific proteins such as protein tau and amyloid-ß have provided a high sensitivity and specificity of diagnostic tests . HoweverThe pathological signature of TDP-43 seen in immunoblots of postmortem tissue has been only partially reproduced in CSF from patients with ALS and FTLD . A polycSOD1 mutation confirming that this subgroup of ALS is associated with a different molecular pathway.Given that it is a ubiquitously expressed protein, other studies have pursued the possibility of TDP-43 as a biomarker in blood lymphocytes . Blood fThe TDP-43 biochemical profile detected in both blood and CSF was investigated to see whether it reflected brain-derived TDP-43, or predominantly represents the physiological full-length form derived from blood . The stuGiven the indications of altered TDP-43 in biofluids, subsequent studies have attempted to quantify the degree of TDP-43 and the pathological altered TDP-43 using either a monoclonal or polyclonal TDP-43 antibody against amino acid regions 1–260, 205–222, and 256–296 or phosphorylation-dependent antibodies to aa409/410 of TDP-43. Nearly all studies used enzyme-linked immunosorbent assay (ELISA) to measure TDP-43 in CSF and plasma of different patient groups. An overview of all ELISA studies is given in Table C9orf72 repeat expansion carriers of both subgroups [n = 10) relative to the FTD patients (n = 50). In contrast to previous studies, a positive correlation with male gender was observed. However, it should also be noted that the study also reported a high inter-assay coefficient of variation.The first quantification of TDP-43 in the CSF of ALS patients used an ELISA technique against full-length TDP-43 with a commercially available chemiluminescent substrate to enhance the TDP-43 signal . With a ubgroups . TDP-43 GRN, C9orf72, and MAPT) or clinically. Remarkably, this is the first study reporting lower TDP-43 levels in patients with neurodegenerative diseases of FTLD-TDP and AD compared to controls. Thus, in lumbar CSF, full-length TDP-43 was significantly lower in FTLD-TDP compared to controls, but with high group variability, while FTLD-Tau was not different from controls or FTLD-TDP. In lumbar CSF, phosphorylated TDP-43 was not detectable. In autopsy-obtained ventricular CSF samples, lower levels of full-length TDP-43 were detected in FTLD-TDP and AD compared to controls, and non-significantly lower levels in FTLD-Tau were observed. Phosphorylated TDP-43 levels were lower in FTLD-Tau and AD, and a trend towards lower levels was observed in FTLD-TDP. This study might suggest that lower levels of TDP-43 are a feature of neurodegeneration in general, as lower levels were not restricted to TDP-43 proteinopathies. Of note, a possible breakdown of the CSF barrier postmortem is likely to have an influence on the quantification of TDP-43 in the ventricular CSF samples such as impaired antibody binding through higher IgG and albumin contamination from blood [Another study group used an ELISA with a full-length TDP-43 coating antibody for ventricular CSF obtained postmortem at the time of autopsy and lumbar CSF samples, applying a biotin-streptavidin detection method for the latter . For eacom blood .The first study to quantify the full-length TDP-43 isoform in plasma included an initial patient cohort clinically diagnosed with FTD and Alzheimer’s disease (AD) . In thisTARDBP mutations, were increased compared to healthy control subjects. A positive correlation with age was shown. A subgroup of patients had longitudinal samples at different time points but no conclusion could be drawn due to the heterogeneity of TDP-43 levels.In a separate study, plasma levels of TDP-43 in a large cohort of 219 ALS patients were measured by ELISA using a chemiluminescent substrate . By thisC9orf72 repeat expansion, GRN mutation carriers, and healthy controls [C9orf72 and GRN mutation carriers compared to FTD non-mutation carriers and healthy controls. Interestingly, full-length TDP-43 levels were found to be negatively correlated with phosphorylated TDP-43, meaning that it was lower in mutation carriers compared to non-mutation carriers and controls. Both TDP-43 forms in plasma correlated negatively with age, which was taken into account in the statistical analysis. No difference was found when investigating a smaller CSF sample group of 20 FTD patients and 5 FTD mutation carriers compared to controls.Another study measured full-length and phosphorylated TDP-43 in plasma in FTD non-mutation carriers compared to a small subgroup of FTD patients with a controls . This stA different methodology aimed to detect TDP-43 with an atomic force microscopy-based biopanning protocol, utilizing phage-displayed, single-chain antibody fragments directed against TDP-43 from an established library , 81. PhaThe pathologic specimens of TDP-43 have been well established from human postmortem studies; however, the quantification of TDP-43 isoforms in the CSF and plasma of ALS/FTLD patients has been contradicting. Although a range of antibodies can be used to detect TDP-43 in biofluids , an unacA limitation of all reviewed ELISA studies on plasma and CSF TDP-43 levels is the use of the common commercially available antibodies for TDP-43. These are restricted to a certain peptide region or phosphorylation sites of TDP-43. Immunoblotting shows that the identification of TDP-43 in CSF and serum by the commonly applied antibody combinations used for quantification represent a 45-kDa full-length form of TDP-43, rather than disease-specific truncated forms of TDP-43 while even more concerning commonly applied phosphorylation-dependent antibodies have not yet revealed a convincing immunoreaction in blood or CSF , 69. ProMonoclonal antibodies which selectively recognize the pathological forms of TDP-43 will be of critical importance for the monitoring of disease status and to assess future treatment options (such as immunization studies). An extensive study generated new monoclonal antibodies by immunization of recombinant full-length, N-terminal, and C-terminal TDP-43 peptides recognizing epitopes distributed over the entire length of TDP-43. Although many of these antibodies are suitable for common applications , there was only one antibody reported “mab138” against aa182–261 , which appeared to detect cytoplasmic, pathological TDP-43, independent of its phosphorylation status and not endogenous TDP-43 [postmortem tissue such as phosphorylation and ubiquitination are potential candidates to enrich pathological TDP-43 forms. Serial fractionation of postmortem tissue enables to isolate and enrich insoluble TDP-43 with the identification of a disease-specific pattern [To apply disease-specific TDP-43 antibodies in future studies will be essential to detect brain-derived TDP-43 and to limit the detection of normal TDP-43, which may derive from blood . These s pattern , 8, 47.  pattern , 82, and pattern . Only th pattern –89. As s pattern , 91. The pattern . Recombi pattern . The cha pattern , 93, 94. pattern , 96. New pattern . TDP-43  pattern , 95.TDP-43 is central to ALS and FTLD pathology, and the frontrunner for a suitably specific biomarker that may also reflect individual disease activity. While the simple measurement of TDP-43 from blood and CSF has failed so far, only continued advances in understanding the localization, protein interactions, and immunology of its various modified forms will deliver this full potential.


SP263 and SP142 showed strong concordance (k = 0.788) with combined+ rates of 33.3% (42/126) and 34.9% (44/126), respectively. In DLBCL‐NOS, TC+ by SP263 preferred to non‐GCB and immunoblastic variant DLBCL‐NOS (P = 0.029 and P = 0.004). Combined+ (SP263 and SP142) were associated with more than one extranodal site involved , higher ECOG PS scores , high IPI risk , and poor treatment response . IC+ by SP263 and SP142 were both independent risk factors . 9p24.1 locus amplification and gain were identified in 4.3% and 7.6% DLBCL‐NOS and indicated shorter overall survival (P = 0.004). Positive rate of PD‐L1 by RNAscope was 36.5%, while no clinical significance shown. PD‐L1 positive rates were all higher in PMBCL and DHL than in DLBCL‐NOS by SP263, SP142, RNAscope, and FISH . In conclusion, combined PD‐L1 expression by IHC was potentially reliable and convenient as a predicting biomarker. SP263 staining was easier to evaluate and recognized more PD‐L1‐stained cells, but SP142 presented a better prognostic indicator. FISH and RNAscope could be used as supplementary assays. PMBCL itself was a sensitive cohort for immunotherapy.The expression of programmed cell death ligand 1 (PD‐L1) is a biomarker for immunotherapy, but approved detection method is absent in diffuse large B‐cell lymphoma (DLBCL). Here, we performed three methods including immunohistochemistry (IHC) (clone SP263 and SP142), RNAscope, and fluorescence in situ hybridization (FISH) to evaluate PD‐L1 status on a cohort of DLBCL including 94 of DLBCL‐NOS, 25 of primary mediastinal large B‐cell lymphoma (PMBCL) and 7 of double‐hit lymphoma (DHL). SP263 with 25% for immune cell (IC) or combined cell and SP142 with 10% for tumor cell (TC), 20% for both of IC and combined cell were proved to have corresponding survival prognostic. Combined Currently, the main biomarkers for the selection of patient cohorts include PD‐L1 overexpression, microsatellite instability (MSI), and high tumor mutational burden (TMB).In the described study, the protein expression of PD‐L1 was detected by IHC with the antibodies SP263 and SP142, the mRNA level was measured by RNAscope, and the PD‐L1 genetic locus alteration of 9p24.1 was analyzed by fluorescence in situ hybridization (FISH) in a cohort of DLBCL, including DLBCL‐NOS, PMBCL, and double‐hit lymphoma (DHL). The status of PD‐L1 from different molecular levels was determined by the three methods and was compared. At the same time, the corresponding prognostic roles of PD‐L1 detected by each of the methods were analyzed. The investigation aimed to identify a more practical and reliable method for PD‐L1 detection for potentially predicting the response of DLBCL to PD‐1/PD‐L1 inhibitors in clinical practice.22.1One hundred and twenty‐six cases of large B‐cell lymphomas were selected from the files of the Department of Pathology, Peking University First Hospital. Among the 126 cases, 101 cases of DLBCL‐NOS were obtained from 2011 to 2015, while 25 cases with PMBCL were obtained from 2008 to 2017. All the 126 cases were reclassified according to the 2017 World Health Organization (WHO) classification of tumors of hematopoietic and lymphoid tissues. DLBCL‐NOS was defined as cases with diffuse proliferation of neoplastic large B cells but that could not be classified as any other specific subtype of DLBCL, and EBV‐positive cases were also excluded. As a new entity, cases with high‐grade B‐cell lymphoma, with rearrangements of MYC and BCL2 and/or BCL6, which was also called “double‐hit” or “triple‐hit” lymphoma (THL), were selected out from cases initially diagnosed as DLBCL‐NOS. PMBCL was confirmed based on the site of involvement (mediastinum), morphology (diffuse infiltration of large cells and fibrotic background), and distinctive immunophenotype . Six cases of reactive hyperplasia lymph nodes were selected as the control group. All 126 cases had adequate specimens of formalin‐fixed paraffin‐embedded (FFPE) samples, of which 30 DLBCL‐NOS, 9 PMBCL, and 3 DHL were embedded into six TMA blocks with a manual tissue arrayer . The clinical information of the patients was obtained by reviewing the medical digital database; 117 of the 126 cases had available clinical data, and 110 of the 126 cases had complete follow‐up data.2.2To assay the PD‐L1 protein expression level, two antibodies  were used for the 126 cases. IHC with SP263 was performed on freshly cut 4‐μm‐thick FFPE tissue sections using the BenchMark ULTRA automated staining platform  with an Optiview DAB IHC Detection Kit according to the manufacturer's instructions. Every section stained with the PD‐L1 rabbit monoclonal primary antibody  was set up with a matched rabbit immunoglobulin G negative control and a positive control from the placenta. IHC of SP142 was carried out with PD‐L1/PAX5 immunohistochemical double staining according to a previous procedure.+, and TPS  was estimated; for ICs, predominantly macrophages and lymphocytes  was approved by FDA in the utilized of 22C3 antibody in gastric and gastroesophageal junction adenocarcinoma.+  in DLBCL. The percentage of PD‐L1 staining cells in our study was mainly scored by two experienced hematopathologists, and the mean percentage of positive cells was used for analysis. The disputed cases were confirmed by a third hematopathologist. The determination of positive cutoff value for SP263, SP142, and the detailed reasons is described in the Results section.The scoring algorithm of PD‐L1 expression in our study was performed as reported.The antibodies used for classified cell‐of‐origin (COO) and double‐expressor lymphoma (DEL) were anti‐CD10 , anti‐BCL6 , anti‐MUM1 , anti‐BCL2 , anti‐c‐Myc . Immunohistochemical staining was performed on 10% formalin‐fixed, deparaffinized 4‐um sections using routine methods. Color development was carried out using 3,3′‐diaminobenzidine tetrahydrochloride. We use a cutoff of 50% with BCL‐2 expression and 40% with c‐Myc expression to define DEL.2.3FISH was performed on 126 DLBCL FFPE samples according to the manufacturer's instructions. PD‐L1 9p24.1/CD274) Dual‐Color probe  was performed as previously reported..1/CD274 MYC (8q24) break apart rearrangement probe ; BCL2/IGH fusion translocation t  probe  and BCL6 (3q27) break apart rearrangement probe  were performed and evaluated according to the manufacturer's instructions, and the threshold all was 20%.2.4PD‐L1 mRNA levels by ISH with RNAscope were detected in 58 samples, including 30 cases of DLBCL‐NOS, 25 cases of PMBCL, and 3 cases of DHL (six TMA samples and the remaining 16 samples were PMBCL FFPE puncture cases). An RNAscope assay  with the probes Hs‐PAX5‐C1  and Hs‐CD274‐C2 , positive control probes Hs‐PPIB‐C1/POLR2A‐C2 , negative control probe DapB , and RNAscope® 2.5 HD Duplex Reagent kit  was performed following the manufacturer's instructions. In brief, 5‐μm‐thick paraffin‐embedded TMA sections were baked for 60 minutes, deparaffinized, and then boiled with the pretreatment reagent for 15 minutes. Protease digestion was carried out at 40°C for 30 minutes, followed by probe hybridization for 2 hours at 40°C with target probes against PD‐L1 and PAX5. Hybridized signals were amplified and visualized with the RNAscope 2.5 HD Duplex Reagent kit. PD‐L1‐positive signals showed as red punctate dots in the cytoplasm or nucleus, and green punctate dots (PAX‐5) distinguished tumor cells from immune cells. PD‐L1 mRNA expression levels were categorized into 5 grades according to the manufacturer's scoring guideline: score 0, no staining or < 1 dot per 10 cells; score 1, 1‐3 dots per cell; score 2, 4‐9 dots per cell and no dot clusters; score 3, 10‐15 dots per cell and < 10% of positive cells contain dot clusters; score 4, >15 dots per cell and > 10% of positive cells contain dot clusters. Scores of 0, 1, and 2 were classified as negative, and 3 and 4 were positive.2.5P‐values involved in this study are 2 sided, and a P‐value < 0.05 was considered significant. All statistical analyses were performed with the SPSS 20.0 software.The interobserver concordance and concordance between the results of PD‐L1 presence detected by IHC, RNAscope, and FISH was evaluated by the Kappa value. A Kappa value ≤ 0.40 was poor concordance; 0.41‐0.75 was moderate concordance; >0.75 was strong concordance. The relationship between PD‐L1 alterations and clinicopathological characteristics of the patients was compared using the chi‐squared test or Fisher's exact test. The treatment outcomes were measured by overall survival (OS), which was defined as the time from the day of diagnosis to the day of death or the day of the last follow‐up. Estimates of OS were calculated by the Kaplan‐Meier method, and the log‐rank test was used to compare the difference between the two groups. Cox regression models were used to estimate the prognostic risk factors. All 33.1The 126 patients included 66 males and 60 females with ages ranging from 15 to 89 years (median 59 years). According to the 2017 WHO classification, 94 of cases were reclassified into DLBCL‐NOS, 25 were PMBCL and 7 were DHL. All the 126 patients had available samples and presented optimal results for IHC, with SP263 100% (126/126) and SP142 100% (126/126).PD‐L1 expression assessed by SP263 showed generally stronger and more complete membrane staining  cases, including 37 cases with positive expression and 77 cases with negative expression  cases were diagnosed as GCB type DLBCL, and 63 (67.0%) cases were non‐GCB type DLBCL. DEL was diagnosed in 27 (28.7%) cases. As to histologic classification, 12 (12.8%) cases were grouped as immunoblastic variant subtype, and 82 (87.2%) cases were centroblastic variant subtype. TC3.3P = 0.006, P = 0.042, respectively), higher ECOG PS scores , high IPI risk , and poor treatment response . Besides, patients with positive SP263 expression were more likely to relapse (P = 0.029). Owing to the small size of the cohort, patients with PMBCL and DHL showed no relationship with clinical features (data not shown).A total of 117 patients had available and complete clinical information, of which 91 patients were DLBCL‐NOS, 19 cases were PMBCL and 7 cases were DHL. The clinical features of 91 DLBCL‐NOS patients and the association with PD‐L1 alterations are summarized Table . Among 9+ DLBL‐NOS by IHC had inferior OS  patients died due to DLBCL‐NOS progression; 18.8% (3/16) of patients died due to PMBCL; 66.7% (4/6) of patients died due to DHL; and 5 patients died for other reasons. About 18.2% (16/88) patients with DLBCL‐NOS experienced recurrence. The five‐year survival rate of patients with DLBCL‐NOS was 71.6%. In DLBCL‐NOS, the Kaplan‐Meier analysis showed that patients with combined/88 patie3.4P < 0.001, Table P < 0.001, P < 0.001, P < 0.001, respectively; Figure FISH was performed on all 126 samples, and 119 samples yielded usable results. Among them, 15 cases were identified with 9p24.1 locus amplifications (12.6%) and 17 cases with gains 14.3%)  of the results were available, including 29 cases of DLBCL‐NOS, 21 cases of PMBCL, and 2 cases of DHL. High PD‐L1 expression score = 4) was presented in 5 (9.6%) cases; score 3 was presented in 14 (26.9%) cases; score 2 was presented in 7 (13.5%) cases; score 1 was presented in 16 (30.8%) cases; and score 0 was presented in 10 (19.2%) cases. Positive PD‐L1 mRNA overexpression (score = 3‐ 4) was detected in 36.5% (19/52) cases  can all lead to upregulation of PD‐L1.PD‐L1 mRNA expression level has been measured in solid tumors, including gastric cancer and non‐small‐cell lung cancer by using RNA‐seq or ISH with RNAscope, and it was found to be highly consistent with their IHC detection results.Previous study showed that in tumors with high PD‐L1 amplification occurrence and/or PD‐L1 overexpression, patients seemed to be more susceptible to PD‐1/PD‐L1 blockade,In conclusion, the detection of PD‐L1 by IHC could be a reliable and easy method in predicting the response to PD‐1/PD‐L1 blockade for DLBCL‐NOS, especially the combined PD‐L1 staining recording could be used in practice. The cutoff for SP142 and SP263 should be different. The staining of SP263 was easier to evaluate and was more sensitive, while SP142 was a more accurate biomarker for assessing the prognosis of patients with DLBCL‐NOS. FISH and RNAscope could also be considered as a supplementary diagnostic assay. In PMBCL, the high frequency of PD‐L1 locus amplification and PD‐L1 overexpression probably indicated their good response to immunotherapy. Nevertheless, a concrete cutoff of PD‐L1 expression and detected method chosen should be determined by their relation to the outcome or response to immunotherapy in clinical practice; hence, our study was more a suggestion than a principle in PD1/PD‐L1 blockade prediction, and treatment response and larger cohort were needed in the future study.No conflict of interest disclaimed. Click here for additional data file. Click here for additional data file. Click here for additional data file. Click here for additional data file.


The companion diagnostic test for checkpoint inhibitor immune therapy is an immunohistochemical test for PD-L1. The test has been shown to be reproducible for expression in tumor cells, but not in immune cells. Immune cells were used in the IMpassion130 trial which showed PD-L1 expression was associated with a better outcome. Two large studies have been done assessing immune cell PD-L1 expression in lung cancer. Here, we reanalyze one of those studies, to show that, even with an easier scoring method, there is still only poor agreement between assays and pathologist for immune cell PD-L1 expression. Companion diagnostic testing has gained increased importance of the last few years. The earliest companion tests were immunohistochemistry (IHC) based (estrogen receptor and HER2). These have recently been followed by a series of molecular, mutation-based tests (EGFR and BRAF) and most recently, another IHC test for PD-L1. When the FDA clears or approves companion diagnostic tests, it is widely assumed that these tests are accurate, reproducible, and robust. In fact, the SSED (Summary of Safety and Effectiveness Documents) released by the FDA provide the evidence to justify the assumption that the tests are worthy of consumer, payer, and physician confidence. Examination of the SSEDs for the PD-L1 tests shows that the FDA clears assays after review by only 2 or 3 pathologists, often showing high overall percent agreement (OPA) that may not reflect real-world outcomes. In fact, when PD-L1 assays were assessed by multiple observers, some FDA-approved categories were found to be unreproducible, specifically including immune cell expression of PD-L1 [In October of 2018, Schmid and colleagues from Genentech reported the results of the IMpassion 130 trial in first-line metastatic setting in breast cancer , 5. As sHere, we reanalyzed the data from NCCN study  using thThe low agreement between the assays is likely attributable to previously demonstrated lower SP142 sensitivity compared to other FDA-approved and laboratory-developed test (LDT) assays [In summary, this analysis raises a significant concern for pathologists who need to provide accurate and reproducible companion diagnostic results for PD-L1. While the NCCN study data presented here are from lung cancer, not breast cancer tissue, there is no evidence that the biochemistry of the interaction has any difference between the tumor sites. While the lung cancer pathologists in the NCCN study were not trained to read IC scores, the Blueprint 2 study included 1.5 days of training for 15 pathologists and found very low concordance, suggesting that training will not solve this problem. We look forward to Genentech’s help in solving this problem. A potential solution would be a reanalysis using the SP263 assay (produced by the same vendor as the SP142 assay) or a bridging study between the SP142 assay and the SP263 assay using the IMpassion 130 tissues.


According to RM, truthful memories of actual events originate in perceptual experience and are embedded in the context of time and space. As such, they are expected to include more spatial and temporal contextual attributes  and perceptual attributes  than do false memories, which originate in self-generated thought or imagination. Derived from this prediction, the traditional use of richness in detail as an indicator of deception is based on the number of perceptual and contextual details in the interviewee's accounts. However, as a memory source-monitoring theory, RM does not take into consideration the intention of liars to deceive and consequently cannot explain the full scope of richness in detail in the field of deception  theory , is richeception . In contmories”; ]. Liars mories”; –5, for e details , 7. Affeach VA; , 8] appl applrichquality of perceptual and contextual details than it does from consideration of their quantity alone.The VA  for lie liars' dilemma hypothesis. Specifically, liars perceive richness in detail as an indicator of truthfulness . However, their strategy leaves traces in the quality of their accounts, in terms of verifiability. By assessing the quality  rather than the quantity of details provided, it is possible to reveal the liars' strategy, and thereby indicate their lies.In the last years, the validity of the VA, which was originally developed and tested in police interview setting e.g., , 12, 13], 1312, 1The first approach was recently proposed by the authors of the current paper Nisin and Nahari (in preparation), who suggest that the qualitative differences between perceptual and contextual details can serve as a potential generator of deception strategy. According to this approach, while perceptual information is actually experienced, and acquired directly by the senses, contextual information is virtual in its nature, and based on semantic knowledge and relative conceptualizations. For instance, we experience the perceptual aspects of an interaction with a friend through our senses: we see the friend and the clothes he is wearing, hear his speech, and feel his touch. Meanwhile, the contextual aspects of this interaction, such as its length and specific location, are based on conceptualization and knowledge. In fact, the contextual attributes are imposed on the perceptual details and frame them in time and space. Accordingly, the perceptual details  can be regarded as primary data, and the contextual details —as meta-data. Obviously, the truthfulness of perceptual details can be checked only when they are given by the interviewee within the framework of contextual information regarding time and space. Thus, the contextual details are those that confer the status of verifiability upon perceptual details. Considering the differences between the two types of details in light of the VA , liars wresolution of the verifiable details provided: names increase the resolution of the details. The mention of an identifiable person without a name leaves the interviewees with degrees of freedom, at least temporarily, such that the verifiability of the details is neither immediate nor easy (relatively speaking) to check. Importantly, these degrees of freedom also range, as mentioning an unnamed uncle, for example, leaves less degrees of freedom than mentioning an unnamed acquaintance. These assumptions lead to the expectation that liars, when they do provide verifiable details, will strategically prefer low-resolution over high-resolution details, while truth-tellers will prefer high-resolution details. Again, as with the contextual details, the number of low-resolution verifiable details in an account can serve as a verbal lie indicator.The second approach involves the resolution of the verifiable details provided, as determined by the immediacy in which the information they incorporate can be verify. A good example of such resolution involves the use of names, which already found significance for lie detection . Accordidetail quantity while sharpening the differences in detail quality between truthful and fabricated accounts, we stress that research should go beyond the surface of content, to look for strategies that activate verbal behaviors among liars, define the qualitative manifestations of their strategies, and then—to exploit these manifestations in indicating their lies. The VA, a new and promising paradigm for deception detection is an outcome of this research approach. Having presented two new, un-studied, strategy-based approaches to further demonstrate and extend this line of research, we propose that following this path will benefit the field both theoretically and practically.The current paper is a call for more strategy-based research in the field of verbal lie detection. By demonstrating how strategies blur the differences in GN: conceptualization and writing the first draft, revising and editing. ZN: conceptualization and review and editing.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest. The reviewer BK declared a past collaboration with one of the authors GN to the handling editor.