Patient suicidality is a frequently experienced topic for psychotherapists. Especially adolescents with borderline personality pathology (BPP) often exhibit suicidal tendencies. Previous research which examined therapists’ countertransference towards suicidal patients suggested that therapists are negatively affected and distressed by them. We hypothesize that this emotional response of the therapists is related to specific sessions in which suicidality came up as a topic. Accordingly, the objective of this study consists in examining therapists’ emotional state on a session level of analysis.N = 21 adolescents (age 13–19 years) with BPD or subthreshold BPD. Therapists’ emotional states were measured in n = 418 sessions using the Session Evaluation Questionnaire. Principal component analysis was used to reduce dimensionality of the therapist response. The emotional states were compared depending on whether suicidality has been addressed in the session (SS) or not (NSS).The sample consisted of Discussion: Suicidality does not always have to be a burden for therapists: Both a “distress” and an “eustress” component occur in this context from which the latter is supposed to help clinicians master a difficult situation. Since countertransference feelings are often not fully conscious, it is necessary to do research on therapists’ emotional states after sessions in which suicidality is addressed. This is crucial to both prevent the therapeutic process from being endangered and preserve clinicians’ mental health. Clinical implications and limitations are discussed.Two components could be identified. Firstly, therapists were more aroused, excited, afraid, angry and uncertain after SS than after NSS. Secondly, therapists were more aroused, excited, definite and pleased after SS than after NSS.  Suicidality is defined as “the risk of suicide, usually indicated by suicidal ideation or intent, especially as evident in the presence of a well-elaborated suicidal plan” . 50 to 9Suicidality plays an important role in Borderline Personality Disorder (BPD). 60 to 70% of patients with BPD commit a suicide attempt at some point in their life and the rate of completed suicide is 50 times higher in patients with BPD than in the general population . DespiteThe term countertransference is of great historical importance for therapeutic work since it has heightened therapists’ awareness of their own feelings and has been coined in different ways. In the last decades, it has received more attention since it was acknowledged having great impact on treatment process and outcome , 12. TheIn association with suicidal patients, the term “countertransference hate” was introduced , which cScientific evidence about therapist’ countertransference towards suicidal patients is broadly consistent and suggest that it involves negative emotions such as anxiety, anger and higher self-reported distress , 6, 14. Since previous research used a between-subject design comparing therapist responses towards suicidal versus towards non-suicidal patients, the question remains whether specific suicidal statements or general personality structure mediated the correlation with therapist response. Moreover, research has mostly focused on therapist traits rather than states such as affect. Therefore, it is important to examine within-therapist variables to further the understanding of therapist factors . The curSome of the method passages are adopted from Zimmermann et al. , 29 wherAll methods were carried out in accordance with relevant guidelines and regulations. All experimental protocols were approved by the ‘Ethikkommission Nordwest- und Zentralschweiz’. Ethical approval was obtained from the local ethics committee ‘Ethikkommission Nordwest- und Zentralschweiz’. All adolescents, their parents and the therapists provided their informed consent.clinicaltrials.gov (NCT02518906). The study aimed at showing non-inferiority of Adolescent Identity Treatment vs Dialectic Behavior Therapy as the outcome research part but, additionally, aims at answering a number of psychotherapy-process questions  participated in this study. For each psychotherapy 25 sessions of Adolescent Identity Treatment were planned  years. Fifteen patients presented with BPD and six with sub-threshold-BPD (three or four fulfilled BPD criteria in SKID-II). Six patients dropped out of treatment but were included in this study. Nine recordings of therapeutic sessions were missing due to technical difficulties or human failure. Figure A total of ed . To inced ; and iore > 60 , 35;). TEight clinicians were involved in the current study of whom six were female (75%). All therapists were psychologists or psychiatrists who underwent or were currently undergoing psychotherapy training to obtain the Swiss specialist degree “Fachpsychologe/in für Psychotherapie FSP” or "Facharzt für Psychiatrie und Psychotherapie FMH". They were either focused on psychodynamic or systemic psychotherapeutic approaches for children and adolescent. Additionally, the therapists underwent training and supervision in Adolescent Identity Treatment which is a manualized therapeutic approach . “AIT After each session, the clinicians had to note information on essential moments in the last session and on suicidality . Sessions are treated as “sessions in which suicidality was addressed” (SS) if suicidal tendency had either been mentioned as an essential moment or if the question on current suicidality had been answered with yes. All the other sessions were labeled as “sessions in which suicidality was not addressed” (NSS). The term suicidality comprises current suicidal ideation, a concrete suicide plan and suicide attempt and excludes non-suicidal self-injury behavior. Figure Therapist response was measured using the Session Evaluation Questionnaire SEQ; –39), For, For39])Data analysis was performed with R . PrincipN = 11 patients (52%) have talked in at least one session about suicidal ideation, the wish to kill themselves or about a concrete suicide plan. On average, the participating patients (N = 21) reported a lifetime suicide attempt history with M = 2.86 attempts (SD = 6.85). In the sample in which suicidality had been addressed during psychotherapy (N = 11), lifetime suicide attempt history measured M = 4.82 attempts (SD = 9.43), ranging from 0 to 30 attempts. The remaining sample (N = 10) had a lifetime suicide attempt history with M = 0.70 attempts (SD = 0.68) with a range from 0 to 2 attempts.N = 42 sessions were labeled as suicidality-sessions  and n = 376 as non-suicidality-sessions . From these 11 patients, talking about suicidality appeared in M = 20% of the sessions, ranging from 4 to 69%. Five patients (46%) mentioned suicidality twice during psychotherapy, two patients (18%) three times, one patient once, one patient four times, one patient ten and one eleven times .This step was used to extract principal components from the SEQ data as a preparatory step to later correlate the extracted components with observed suicidality  = 2.40, p < 0.05, ICC = 0.19, σ2 = 3.85, deviance = 1784.2) and the PC2 therapist response pattern  = 0.95, p < 0.001, ICC = 0.18, σ2 = 1.73, deviance = 1450.4). The hypothesis that therapists are more aroused, excited, afraid, uncertain and angry after SS than after NSS  was significantly positively correlated with both the PC1 . We expected that therapists are more afraid, excited, aroused, uncertain and angry after SS than after NSS. Therapist response was measured by Session Evaluation Questionnaire (SEQ) which had to be filled out by all therapists after each session. Results showed that SS are accompanied by higher therapist arousal. PC1 reflects that SS are associated with higher clinician excitement, anger, fear, arousal and uncertainty than NSS. Moreover, PC2 explained 23% of the variance in the therapist response items predicted by suicidality. This pattern is also characterized by higher arousal and excitement but demonstrates as well satisfaction and definiteness.Previous research on this topic has especially dealt with therapists’ emotional responses towards “suicidal patients”. For example, a study with suicidal adolescents has shown that due to therapists' countertransference and due to trait-like suicidal ideation, therapists are unwilling to treat those patients at risk for suicide . In patiPC1 reflects therapists’ negative affect and arousal and is to be summarized in the term “distress”. This is consistent with qualitative research , 6 and sIn contrast to this, PC2 seems to point towards a positive kind of arousal whereby therapists feel definite and pleased. PC2 shows similarities with the concept of “eustress” and will be referred to as ‘eustress-component’. In the APA Dictionary of Psychology, eustress is understood as a “positive stress response, involving optimal levels of stimulation: a type of stress that results from challenging but attainable and enjoyable […] tasks […]. It has a beneficial effect by generating a sense of fulfillment or achievement and facilitating growth, development, mastery, and high levels of performance” . This stOf course, causality cannot be inferred from this correlation of patient suicidality and therapists’ emotional state after session in which suicidality is addressed. For example, an alternative hypothesis could be stated that therapists’ emotional state during the session has an influence on the patients in whether or not they raise the issue of suicidality. Another variant could be that the mood of the therapists has nothing to do with the session per se and thus represents a finding by chance. Moreover, Krause & Lutz  point ouAlthough countertransference is a complex concept with a meaningful historical background, it offers a good possibility to do justice to the multifaceted feelings therapists are confronted with during psychotherapy – both consciously and unconsciously. Since working with suicidal patients can be a very challenging and anxiety-provoking task for psychotherapists, their awareness of concrete emotional patterns in sessions where patients seem at risk for suicide is essential and should discourage the temptation of acting out unconscious feelings or conflicts . AvoidinSecondly, we proposed that addressing suicidal ideation during psychotherapy is associated with both distress and eustress in therapists. The question remains of what distinguishes the therapeutic “distress” reaction from the “eustress” reaction and how the former can be transformed into the latter. In this regard, our results suggest that the main difference between these two emotional responses is that the “eustress” response includes a sense of definiteness and satisfaction and the absence of anxiety, anger and uncertainty which in turn facilitates development and the mastery of a concrete situation. Therefore, it is crucial that therapists feel definite, self-effective and act in the feeling that they are part of the therapy process. By being aware of possible conflictive emotions, a stable therapeutic relationship can be established which in turn leads to a promising outcome. A focus for how therapists can be supported to feel comfortable in challenging situations and a clinical culture that allows for errors and uncertainties should be established. Again, supervision can be helpful for clinicians to gain understanding of a difficult moment in the therapeutic process.Thirdly, it should be kept in mind that suicidal ideation is not always reason for an alarm and that it can overshadow other important issues that should be discussed in psychotherapy . AccordiN = 21 dyads. Only 11 of these dyads discussed the topic of suicidality. However, in terms of investigated sessions, the sample size was 418 sessions of which 211 sessions stemmed from dyads who discussed suicidality. The sample size in the current study is based on a power analysis which was aimed to sufficiently power an outcome comparison . It wasSecondly, the patients’ suicidality was not documented with a validated scale. The therapists documented whether suicidality was addressed or not after each session. Our rationale is that the discussion of suicidality is a salient event and determining whether the event occurred or not should be simple and straightforward. However, a validated scale might be helpful in getting more differentiated results to determine, for instance, the perceived intensity of addressing the topic or the topic's actual acuteness for the patient. Thirdly, therapist response was measured with the Session Evaluation Questionnaire (SEQ) which is a self-rating questionnaire that is not constructed as a stress-test. Although this seems adequate for a subjective description of post-session mood, it should be complemented by a validated stress questionnaire and by psychophysiological measures such as saliva cortisol level or electrodermal skin response to measure the reaction at the level of the autonomous nervous system –57.Two different therapist responses were identified after sessions in which young female patients with BPD or subthreshold BPD talked about their suicidal ideation or about a more concrete suicide plan. Therapists showed an emotional response pattern consisting firstly of arousal and negative affect (“distress”) and secondly of arousal and positive affect (“eustress”). These two types of arousal are distinguished by being angry, uncertain and afraid versus feeling pleased and definite. First of all, this implies that patient suicidality does not always have to be emotionally burdening for therapists and moreover, that clinicians need to feel definite and self-effective which in turn enables the mastery of a challenging situation. If therapists become aware of their “suicidality-countertransference” and their anxiety about patient suicide contained, this can on one hand enable a good therapy process and outcome (including prevent discontinuation of the therapeutic process) and on the other hand lead to clinicians’ well-being. Conducting further research on how therapists’ mental health can be preserved seems necessary in order to ensure appropriate and good care for patients with suicidal tendencies.


The PET and PLA sensors have response and recovery times under 5 min and limited hysteresis. However, the paper and starch sensors have response and recovery times closer to 20 min, with significant hysteresis around 100%. The PET and starch sensors are temperature independent, while the PLA and paper sensors display thermal drift that increases with temperature.Flexible and biodegradable sensors are advantageous for their versatility in a range of areas from smart packaging to agriculture. In this work, we characterize and compare the performance of interdigitated electrode (IDE) humidity sensors printed on different biodegradable substrates. In these IDE capacitive devices, the substrate acts as the sensing layer. The dielectric constant of the substrate increases as the material absorbs water from the atmosphere. Consequently, the capacitance across the electrodes is a function of environmental relative humidity. Here, the performance of polylactide (PLA), glossy paper, and potato starch as a sensing layer is compared to that of nonbiodegradable polyethylene terephthalate (PET). The capacitance across inkjet-printed silver electrodes is measured in environmental conditions ranging from 15 to 90% relative humidity. The sensitivity, response time, hysteresis, and temperature dependency are compared for the sensors. The relationship between humidity and capacitance across the sensors can be modeled by exponential growth with an R Humidity sensing is critical to a wide range of applications, from manufacturing to healthcare. The need for cheap and versatile humidity sensors is extensive  and can Smart packaging utilizes sensors fabricated directly into packaging material in order to track information about its contents and environmental history, such as pH, temperature, gas levels, light exposure, and relative humidity ,8,9. SchAnother use for biodegradable humidity sensors is in high density spatial and temporal monitoring of growing conditions. Extensive data on a crop’s condition allow for more efficient use of resources, and the use of biodegradable sensors can reduce the impact of electrical and plastic waste, as well as the labor required to retrieve sensors at the end of a crop cycle ,12.Currently, it is challenging to compare the performance of humidity sensors printed on different substrates because of a lack of standard sensor characterization and fabrication. Not only are sensors made in different ways ,13, but Humidity can be quantified in many ways, two of which are absolute and relative humidity (RH). Relative humidity is the most common measurement for environmental humidity and is the ratio of actual water vapor partial pressure to the saturation vapor pressure. The saturation vapor pressure is the point at which condensation occurs. Saturation vapor pressure is an exponential function of temperature, in that hotter air can hold more water vapor. The mass of water vapor per volume of air is called absolute humidity and, unlike relative humidity, it is not dependent on temperature . This mePrinted humidity sensors typically take the form of interdigitated electrodes (IDEs), with capacitance measured between the two interwoven electrodes. The capacitance of an IDE depends on the number of fingers, finger width, gap width, and finger overlap, as well as the dielectric constant and thickness of the substrate on which it is printed. An IDE sensor indirectly measures humidity as a function of water vapor absorbed by the sensor’s substrate. In humid environments, the substrate absorbs water, which has a relatively high dielectric constant compared with that of the substrate. As a result, the dielectric constant of the material increases, causing a proportional increase in the capacitance reading across the sensor. When relative humidity drops, water vapor evaporates out of the substrate to achieve equilibrium, and the capacitance decreases.While IDE capacitance varies linearly as a function of substrate dielectric constant , the absSubstrate choice is fundamental to the behavior of IDE capacitive humidity sensors. This paper characterizes and directly compares sensors printed on four different materials: planarized polyethylene terephthalate (PET), polylactic acid (PLA), glossy photo paper, and a starch and polypropylene (PP) composite.PET is a polar polymer that is commonly used for packaging of foods and liquids. It is also a standard material that can be optimized for printed, flexible electronics, and has been studied as a base substrate for humidity sensors ,24. UnliPLA is also a polar polymer, but is made from plant matter and is biodegradable. It was chosen for this study because it is highly ubiquitous, and often considered a biodegradable alternative to PET. PLA is a popular material for packaging and degradable medical devices and is one of the most common filaments for 3D printing. PLA can be degraded by hydrolysis, photo, microbial, and enzymatic degradation. Under ambient conditions in air and soil, it degrades slowly ,26,27.Paper is another popular material for humidity sensors as it is versatile and affordable. A potential application for printed humidity sensors is monitoring shipments of food or medicine, in which sensors can be directly printed onto paper labels or cardboard boxes. Furthermore, many sensors for environmental monitoring are already paper-based, allowing for the potential development of an array of environmental sensors printed on a single sheet of paper. Humidity sensors could be integrated with already established air quality monitoring and electrochemical detecting paper sensors . Paper cFinally, the starch and PP composite is a polar, commercially available bioplastic sold in the form of “Tater Ware”. It is comprised primarily of potato starch and used to make cutlery and other tableware. Starch can be combined with a variety of petroleum-based or biopolymers to make inexpensive, unique polymer combinations. For this reason, starch has recently become popular for use in developing new sustainable plastics , and thuIn this paper, the substrate itself functions as the “sensing layer”. While some capacitive humidity sensors depend on an additional sensing layer deposited on top of the IDE ,23,24,31In order for a humidity sensor to be widely applicable, it must have high sensitivity over a range of humidity, quick response times, little hysteresis, temperature independence, a simple structure, and low costs to manufacture . Thus, eFour types of materials were used as a substrate for the sensors: 50 μm thick planarized PET from Kimoto Tech, 300 μm thick PLA from Tamiya, 300 μm thick glossy coated paper from Kelly Paper, and 500 μm thick starch and PP from Biodegradable Food Service. Prior to printing, the starch substrate was sanded to improve surface uniformity.Sensor electrodes were printed with a drop spacing of 33 μm onto the substrates using the Dimatix Materials Printer DMP-2850  in silver DGP 40LT-15C ink from ANP . The platen was heated to 60 °C for printing on all four substrates to reduce ink spreading . The PET2 squares of ink were also printed onto each substrate. The squares were cured under the same conditions as the sensors, and their conductivity was measured using a Signatone 4 point probe.In addition to the sensors, 1 × 1 cm2 square were placed inside of an environmental chamber  and subjected to a series of humidity and temperature tests. The experimental setup can be seen in the 2 square was measured.Printed sensors were trimmed to size and clipped into a flexible flat cable connector and connected to a precision LCR meter  to take capacitance readings of the sensor. Capacitance measurements were taken every 12 s at 5 kHz frequency. The printed sensor, a commercially available humidity sensor sensor , and the associated 1 × 1 cmSensors underwent three types of tests, as plotted in Data are presented and analyzed for a single sensor on each material; however, additional data are available in the Sensor data can be represented along a temporal x-axis or can be plotted to show the relationship between relative humidity and capacitance. The relationship between relative humidity and capacitance is typically non-linear for capacitive humidity sensors ,32,33, aTotal sensitivity can be compared between sensors by looking at capacitive output range at a certain temperature. The paper- and starch-based sensors have the highest sensitivity, with capacitance increasing by a factor of 6 and 2.5, respectively, as relative humidity goes from a low of 15% to a high of 90%. High sensitivity gives paper and starch an advantage over the lower sensitivity PET- and PLA-based sensors when monitoring small changes in relative humidity. In comparison, PET and PLA only show an increase in capacitance by a factor of 1.6 and 1.15, respectively.Response time is defined as the time it takes for the sensor capacitance to increase from a local minimum to within 90% of the average maximum capacitance across the high relative humidity region. Similarly, recovery time is defined as the time it takes for the sensor capacitance to decrease from a local maximum to within 10% of the average minimum capacitance across the low relative humidity region . HoweverResponse and recovery time depends on three physical processes of permeation. When humidity increases, water vapor absorbs on the surface of material, dissolves through the surface, and diffuses throughout the material. Dissolution depends on a variety of factors such as the interaction energy between the material and water vapor, the concentration of environmental water vapor, and the saturation of the material. Diffusion is driven by a concentration gradient within the material and depends on factors including material porosity, free volume, and substrate thickness. Desorption of water vapor from the substrate occurs when the environmental humidity is low enough that it cannot maintain saturation of the material. The rate of desorption is a function of relative humidity and is not simply the reverse of absorption ,19. TherThe PLA-based sensor has a near instant response to the humidity step, with a response time less than 1 min. A fast response time can be an important characteristic of humidity sensors when sudden environmental changes need to be detected. Meanwhile, the PET-, paper-, and starch-based sensors show a gradual response resembling a logrithmic growth curve, and response times of 21, 24, and 38 min, respectively. Both the paper and PET substrate have a protective film on their surface, which likely slows the rate at which water vapor can dissolve into the bulk of the material, and consequently increases the response time. The starch substrate has a very slow response time because it is both thicker and more porous than the others. Both characteristics allow it to absorb more water than the other substrates, which is reflected in starch’s high sensitivity, but this process takes time.It is expected that the polar materials will have a longer recovery time than response time because of interactions between the material and the absorbed water molecules. This is only true in the case of the PLA-based sensor, which has a recovery time of 3 min. Meanwhile, the recovery time for the PET-, paper-, and starch-based sensors was shorter than their response time at 1, 13, and 13 min, respectively. Although PET is a polar material, it is possible that the planarized coating interferes with absorption, leading to a slow response time relative to recovery time. Paper is not polar, thus a slower recovery time is not expected. Another interesting feature of the paper sensor is the large spike in capacitance right as humidity is dropped from high to low. This may be due to condensation on the surface of the substrate.Hysteresis occurs when the capacitive output depends on whether relative humidity is increasing or decreasing to the point of measurement. The repeatability and accuracy of a sensor’s measurement depend largely on the amount of hysteresis, thus hysteresis should be minimized.If the relative humidity around a sensor drops, but the sensor is slow to release absorbed water, the capacitance of the sensor will remain high until enough time has passed for water to desorb completely . This caiC corresponds to capacitance during the ith humidity step, and the maximum and minimum capacitance values are found for a single step. The numerator is the largest capacitive difference between the curves generated by the humidity increase and decrease. The denominator captures the range of capacitance values, or the full scale deflection, in order to normalize the hysteresis percentage. Normalization is necessary when comparing between humidity steps because hysteresis is dependent on the maximum humidity reached during the step.Hysteresis was tested by cycling between a constant low (15%) and progressively increasing relative humidity. Varying the relative humidity can reveal the extent to which sensors are affected by previous humidity exposures. Hysteresis percentage was calculated for the 30%, 60%, and 90% steps using Equation (2) ,36, wherThe PET- and PLA-based sensors show the least amount of hysteresis, with relative hysteresis decreasing as the humidity steps increase. Small hysteresis curves can be distinguished when zooming in on the left side of the plot, but the amount of hysteresis is low compared with the capacitance range and sensitivity. Meanwhile, the paper- and starch-based sensors show considerable humidity-independent hysteresis as a result of their long response and recovery time. The difference between response and recovery rates results in asymmetric hysteresis loops, where the high capacitance recovery curve has a more gradual slope than the low capacitance response curve. Because response times are longer, most of the capacitance increase occurs in the vertical region of the plot where relative humidity is held constant. The spike in capacitance of the paper sensor as relative humidity goes from 95% to 15% is noticeable in To test temperature dependency, sensors were exposed to relative humidities between 15% and 95% over three distinct temperatures . Temperature impacts the permeation properties of the sensing layer, which can explain a sensor’s temperature dependency. Specifically, a rise in temperature corresponds to an increase in the material’s diffusion coefficient (rate of molecular diffusion) and maximum saturation concentration. This is because higher temperature leads to thermal expansion and to an increased mobility of the water molecules.All four sensors show an increase in thermal drift as relative humidity increases. This suggests that sensitivity at high relative humidity is significantly temperature-dependent, which can be explained by the changes in material permeation as a function of temperature. PET and starch appear to be the least affected by temperature changes, with little variance in sensitivity and base capacitance throughout the three temperature cycles. Paper is affected by temperature, with slight increases in curve slope corresponding to increased temperatures. Although thermal drift is relatively high for this material, paper has a large operating range compared with the polymer materials. Meanwhile, the effects of temperature on the permeation properties and thus sensitivity of the sensors are most evident for the PLA-based sensor, where the curve slope and maximum capacitance increase significantly with temperature.Examining sensor temperature dependence is necessary to establish if sensors are measuring relative humidity or something else. A common misconception is that these humidity sensors only measure the amount of water vapor in the air, or the absolute humidity. At standard pressure, the capacitance output for 15 °C air at 90% relative humidity, 25 °C air at 50% relative humidity, and 40 °C air at 24% relative humidity should be the same regardless of the temperature difference, as these conditions have the same absolute humidity. However, the star markers in 2 printed test square was put in the humidity chamber alongside the corresponding sensor, and its conductivity was measured after each test cycle using a 10 μm source current. In order to determine whether short-term humidity exposure degrades the integrity of the sensors, the 1 × 1 cmRecent innovations in smart packaging for manufacturer to consumer tracking, as well as high density monitoring of crop conditions, require the development of biodegradable humidity sensors. A widely applicable sensor has high sensitivity, fast response times, little hysteresis, and temperature independence. Each of the four studied sensors excel in some, but not all of these categories, and thus would work best for specific rather than general applications. Although this paper supports the viability of biodegradable humidity sensors for short-term use and establishes a baseline performance for newly fabricated devices, future work could look at the change in performance over long-scale testing. It can be expected that sensing behavior will change as the substrate begins to break down. The method of degradation, either through hydrolysis, bacterial activity, or other methods, might also impact long-term sensor behavior. In addition to extended tracking of sensor performance, future tests could include examining a wider range of temperature stability, using fully biocompatible inks, and integrating the sensor with transducers for easier data collection.


We combine Density Functional Theory (DFT) and classical Molecular Dynamics (MD) simulations to study graphene–boron nitride (BN) hybrid monolayers spanning a wide range of sizes (from 2 nm to 100 nm). Our simulations show that the elastic properties depend on the fraction of BN contained in the monolayer, with Young's modulus values decreasing as the BN concentration increases. Furthermore, our calculations reveal that the mechanical properties are weakly anisotropic. We also analyze the evolution of the stress distribution during our MD simulations. Curiously, we find that stress does not concentrate on the graphene–BN interface, even though fracture always starts in this region. Hence, we find that fracture is caused by the lower strength of C–N and C–B bonds, rather than by high local stress values. Still, in spite of the fact that the weaker bonds in the interface region become a lower fraction of the total as size increases, we find that the mechanical properties of the hybrid monolayers do not depend on the size of the structure, for constant graphene/BN concentrations. Our results indicate that the mechanical properties of the hybrid monolayers are independent of scale, so long as the graphene sheet and the h-BN nanodomain decrease or increase proportionately. We combine Density Functional Theory and Molecular Dynamics to study graphene–BN monolayers ranging from 2 nm to 100 nm. We find that the mechanical properties are independent of scale when we increase graphene and the BN domain proportionately. I.1 Within the family of 2D materials, those composed of sp2-hybridized atoms arranged in a honeycomb-like hexagonal lattice stand out, with graphene2–4 and hexagonal boron nitride (h-BN)5–7 being the two most notable examples. Graphene in its pristine form is a semimetal8 with the highest Young's modulus (≈1 TPa) and ultimate tensile strength (≈130 ± 10 GPa) ever measured.9–12 On the other hand, h-BN in its pristine form is an insulator (>4 eV) with high resistance to oxidation.13,14 The mechanical properties of h-BN are also outstanding, with a Young's modulus of ≈0.665 TPa and an ultimate tensile strength of ≈70.5 ± 5.5 GPa.9,15–17 Both graphene and h-BN are materials with a promising future for theoretical/experimental studies and practical applications in nanotechnology.Over the past few decades, two-dimensional (2D) materials have been an active research topic due to their exceptional physical and chemical properties, which promise applications in many modern technologies.18–21 The reason for this is that the band gap of graphene (h-BN) is too small (large), leading to devices that do not work properly in an on-off current regime. Given this scenario, continuous efforts have been made in order to predict and synthesize alternative 2D structures. Examples include the BxCyNz hexagonal hybrid sheets (h-BxCyNz sheets), which are structures composed of carbon, boron, and nitrogen that have already been synthesized.22–24 These materials can be thought of as being a graphene with part of their C atoms replaced by B and N atoms. The properties of h-BxCyNz sheets are strongly dependent on the arrangement of the C, B, and N atoms, as well as the particular stoichiometry.25–29 These sheets exhibit a range of band gaps (<2 eV) that are intermediate between those found in graphene and h-BN.26–31 The adjustable band gap of these hybrid 2D materials indicate that they are better candidates than either graphene or h-BN for the production of modern nanoelectronic devices.In spite of their excellent properties, there is evidence indicating that graphene and h-BN are not suited for the construction of certain nanoelectronic devices.xCyNz sheets are those composed of h-BN nanodomains embedded within a larger graphene sheet (graphene–BN sheets). Structures of this type were synthetized for the first time by Ci et al. using the thermal catalytic CVD method.22 The reported HRTEM images revealed that most samples had two or three layers, and included large h-BN nanodomains with irregular geometric forms. These novel 2D hybrid materials have physical–chemical properties that can be tailored by controlling the size of the h-BN nanodomains. After this discovery, other experimental methods were developed for the growth of this type of structure.23,24,32–36 Concurrently, Manna et al. performed DFT calculations to investigate hybrid sheets including h-BN nanodomains of varying size and geometry .37 The authors found that the boundary between the h-BN nanodomain and graphene governs the electronic properties of these structures. Depending on the number of C–B and C–N bonds, the graphene–BN sheets may be metallic, semimetallic, or semiconducting. Similar results were found in other reports.38–42An interesting subset of the Bvia controlled mechanical deformation.43–47 This can be accomplished, for example, through the application of tensile stress.48–51 Depending on the direction and magnitude of this stress, the material can be bent, wrinkled, stretched, or even broken. Experimental methods to introduce tensile stress include: (i) depositing the 2D structure on a flexible substrate that can be bent, elongated or shrunk;52–54 (ii) depositing the 2D material on top of a hole in a substrate, and then pushing it down with an atomic force microscope (AFM) tip.55 All of these findings have contributed to the emergence of a new research area called “straintronics”, where strain engineering methods and strain-induced physical effects are used to develop devices for new technologies.56,57 Finally, note that the mechanical properties of a material, such as stiffness and tensile strength, can also be determined through the application of a tensile stress.Another common approach to modulate the physical–chemical properties of 2D materials is to use strain engineering, where the properties of a material are modified et al. investigated the mechanical properties of graphene–BN sheets using molecular dynamics (MD) simulations.58 These authors considered different sizes, shapes, and amounts of h-BN nanodomains embedded in graphene sheets, and found that hybrid sheets displayed strong plasticity behavior. They also found that the Young's modulus of the hybrid sheets presented values intermediate between those of graphene and h-BN, decreasing as the concentration of h-BN increased. It is hoped that future experimental studies can confirm this prediction. Recently, Azevedo and Kaschny employed DFT calculations to study the mechanical properties of one of the structures considered by Zhao et al., namely, a graphene sheet containing a hexagonal h-BN nanodomain in its center.59 In addition to investigating the mechanical properties of the graphene–BN sheets, these authors also studied how the electronic properties of the hybrid sheets varied with increasing strain. Additionally, MD60 and DFT studies61 investigated the mechanical properties of BxCyNz hexagonal sheets with atomic arrangements including a large amount of B–C and N–C bonds. The reported values of stiffness and ultimate tensile strength were lower than those reported for structures composed of h-BN nanodomains embedded in graphene sheets.Many theoretical studies have investigated the mechanical properties of 2D materials. For example, Zhao xNz sheets with sizes ranging from 2 nm to 100 nm, using molecular dynamics simulations. Furthermore, in order to assess the reliability of the classical potential considered, we performed DFT calculations for the smallest structures considered here and compared the DFT and MD results. Our calculations show that the MD results are reliable, and that the mechanical properties do not depend on the scale of the considered structures, so long as the size of the h-BN nanodomain and the graphene sheet are increased by the same factor.Still, up to now all simulation studies have only considered small model structures, far smaller than those investigated in experimental studies. And have not considered whether the small unit cell size could affect the obtained results. Note, for instance, that the fraction of B–C and N–C bonds decreases as the structure size increases. In order to verify possible size effects, here we investigated the mechanical properties of square graphene–BII.Lx = Lz = L), with side lengths (L) ranging from 2 to 100 nm  and by the molecular formula BxNx, which specifies the number of atoms in the domain. For example, L2nm-B3N3 stands for the hybrid sheet illustrated in γ), which is determined by the following equation:nB and nN follow the previous definition, and nT is the total number of atoms. γ values for the investigated structures vary from 0.01 to 0.34, as shown in We combined first principles calculations and MD simulations to investigate the mechanical properties of graphene nanosheets containing h-BN nanodomains (graphene–BN sheets). Some of the structures considered here are shown in 0 nm see  and 2. R0 nm see  and circ0 nm see . We denox) and zigzag (z) directions, as illustrated in To determine the mechanical properties of the graphene–BN sheets, we applied a tensile strain along one direction and then calculated the resulting tensile stress to obtain a stress–strain curve. The methodology used to obtain the stress differed in the DFT and MD simulations. In the former, a derivative of the total energy was used in the calculation, while in the latter the forces acting on the atoms were used. Details can be found in the ESI.62 as implemented in the SIESTA code.63,64 The exchange-correlation energy is expressed within the generalized gradient approximation (GGA),65 in the form of the Perdew–Burke–Ernzerhof (PBE) functional. We used the double-ζ polarized basis set (DZP), with core electrons described by norm-conserving Troullier–Martins pseudopotentials,66 in the Kleinman–Bylander factorized form.67 The optimization of atomic positions was allowed to proceed until the force on each atom was less than 0.1 eV Å−1. A convergence criterion was adopted where self-consistency is achieved when the maximum difference between the output and the input of each element of the density matrix, in a self-consistent field cycle, is less than 10−4 eV. Charge density was represented in the real space by a cutoff of 150 Ry for the grid integration. The Brillouin zone was sampled using a 10 × 1 × 10 k-point mesh within the Monkhorst and Pack scheme. We adopted a rectangular unit cell. When the tensile strain is applied to one direction, periodic boundary conditions are imposed in the other directions. A vacuum region of 100 Å was added along the z direction to avoid artificial interaction between neighboring images.The first principles calculations are based on density functional theory (DFT),68 The interactions between B, C, and N atoms were described with the Tersoff potential,69 using the parameters adjusted by Kinaci et al.70 Our MD simulations were performed using a reasonably small timestep of 0.1 fs, and proceeded in three steps:The MD simulations were carried out using the Large-Scale Atomic/Molecular Massively Parallel Simulator (LAMMPS) code.5 steps in the NPT ensemble. Nose–Hoover thermostats and barostats71–73 were used to set temperature and pressure values to 10 K and 0 Pa.(1) We first evolved the system for 2 × 105 steps in the NPH ensemble, using the same barostat described above to set the pressure to 0 Pa. Note that the barostat was only applied to the planar direction that is not strained in the following simulation step.(2) The thermostat was turned off and then the system was evolved for 2 × 106 steps using a strain rate of 10−6 fs−1 .(3) In the final step we maintained the barostat configuration described in Step 2, and then pulled the system for 3 × 10L = 2 nm) shown in L ≤ 50 nm) shown in L is kept constant and the graphene/BN concentration is variable. In (b), we illustrate the case in which L is variable and the graphene/BN concentration is kept constant. For the latter, we also performed MD simulations using a larger structure with L = 100 nm. For all structures, we obtained stress–strain curves by applying a tensile strain along the zigzag or the armchair direction. Next, for the various stress–strain curves we calculated the slope of the linear region to determine the Young's modulus (Y) of each structure. The tensile strength (σ) and the ultimate strain (ε) are taken at the point where the stress reaches its maximum value. The calculated values of Y, σ, and ε are presented in Initially, we performed DFT calculations and MD simulations to determine the mechanical properties of the small hybrid sheets L =  nm shownIII.L2nm-B3N3, L2nm-B12N12, and L2nm-B27N27, illustrated in 43,44We analyze first the mechanical properties of L2nm-B3N3, L2nm-B12N12, and L2nm-B27N27 are summarized in 74 and graphene.75 In the case of graphene, this behavior was attributed to the hexagonal structure of its unit cell.75 Our results also show that Young's modulus (Y) and tensile strength (σ) values decrease with increasing concentrations of h-BN, regardless of the direction of applied strain. These results are in good agreement with those reported by Zhao et al.58 and Azevedo and Kaschny.59 Moreover, our DFT and MD results show that the hybrid sheets present Y and σ values intermediate between those of graphene and h-BN.9,10,15 Finally, our simulation results reveal that the tensile strength of the hybrid sheets is about 22.2% lower than that of graphene (130 GPa) and 23.6% higher than that h-BN (70.5 GPa).The mechanical properties of L2nm-B3N3, L2nm-B12N12, and L2nm-B27N27 reaches a maximum for strain values around 21% for the armchair and 15% for the zigzag direction. For higher strain values, the material undergoes fracture  using the following equation:E and E0 are the total energies calculated for stretched and unstretched hybrid sheets, respectively. In 9,59To finish the discussion of the DFT results, note that it is possible to obtain the strain energy (L ≤ 50 nm), which are shown in L = 40 nm) but varying domain size , but varying size (10 ≤ L ≤ 50 nm). The stress–strain curves obtained for case (i) are shown in L10nm-B376N376 is an example). However, for other structures, such as L20nm-B1503N1503, the tensile strength is still higher in the armchair direction. Finally, note that the stress suddenly drops after reaching its maximum value, due to the fracture of the hybrid sheet.Let us now discuss the mechanical properties of the larger hybrid sheets (10 ≤ Y) is sensitive to the graphene/BN concentration, but not to the size of the structure (for constant γ). Also, notice that Y values are higher in the armchair than in the zigzag direction, for all sheet sizes considered. Still, the difference is small, ranging from 0.78% to 11.86%. And, as expected, the tensile strength of the hybrid sheets is considerably lower than that of graphene.The mechanical properties of the larger hybrid sheets are summarized in γ). Note that Y decreases as γ increases for the range of investigated values. The same general behavior was also observed for the smaller hybrid sheets, as shown in Y values differ by ∼10%. Finally, note that we also performed calculations using different temperatures and strain rates, to verify the validity of these results. Overall, we observed that the trend remained the same for all temperatures and strain rates, although we also found that the Young's modulus decreased as the temperature and the strain rate increased. These results are detailed in the ESI.†Let us now analyze results for the case where the concentration is changed while the sheet size is kept constant. Y is equal to ∼907 GPa in the armchair and ∼898 GPa in the zigzag direction for γ ≈ 0.19. To confirm that the Young's modulus does not depend on the size of the hybrid sheet, we also considered a very large hybrid sheet, with L = 100 nm. Results for this structure confirm that the Young's modulus does not depend on the size of the considered sheet for constant γ  indicate good agreement between the DFT and MD results. For all the considered structures and methods, we found that the Young's modulus decreased as the h-BN concentration increased. In addition, simulation results showed that the tensile strength of the hybrid sheets are ∼15–20% lower than that of graphene. Furthermore, we found that the ultimate strain of the graphene–BN sheets was lower than that of graphene or h-BN, but still greater than 10%. This result indicates that the graphene–BN sheets are well-suited for strain engineering, as they can withstand large strain values. We also found that the mechanical properties of the hybrid sheets are weakly anisotropic. By analyzing the spatial distribution of stress in the hybrid sheets before fracture, we found a rather uniform stress distribution, with slightly lower values in carbon atoms near the h-BN nanodomain. In spite of that, fracture always started at the graphene–BN interface, as the weaker B–C and N–C bonds required lower stress values to break. Finally, we found that the Young's modulus does not depend on the scale of the considered structure, remaining the same so long as the graphene/BN concentration remained constant. This finding indicates that the results described here should remain valid for even larger graphene–BN sheets, like those synthesized in experiments.In summary, we combined DFT and MD simulations to investigate the mechanical properties of graphene sheets containing h-BN nanodomains (graphene–BN sheets). We considered hybrid sheets of varying atomic composition and size (ranging from 2 nm to 100 nm). The stress–strain curves obtained for the smaller structures (There are no conflicts to declare.RA-011-D1RA00316J-s001


RSC Advances article due to concerns affecting the reliability of the data.We, the named authors, hereby wholly retract this et al., RSC Adv., 2019, 9, 23843–23855), however, the experimental procedures of these two papers are different, with different foam ratio and graphene content, so the same picture cannot be used to illustrate different samples.Fig. 3d is exactly the same as Fig 3d in our related paper “A versatile strategy for alternately arranging the foam ratio layers of multilayer graphene/thermoplastic polyurethane composite foams towards lightweight and broadband electromagnetic wave absorption” –(d) represent 2–5 layers of TPU/graphene multilayer samples, respectively. However, all the images are taken from a picture of the same electron microscope image under different magnification rates, which cannot prove the microscopic morphology of the samples with different layers. At the same time, the size labeling is incorrect. The magnification ratios of (a)–(d) are different, so the size labeling cannot be all 3 mm.On page 21864 of the article, it states “The graphene, selectively dispersed over the cellular walls by foaming, formed a three-dimensional conductive network structure, as shown in the Fig. 6b”. However, in Fig. 6b, the three-dimensional conductive network structure and graphene’s selective dispersion and alignment in the pore structures cannot be seen very clearly, because it is only a cell structure diagram, and therefore does not prove the existence of a three-dimensional conductive path.As the above data errors and misuse of pictures have greatly influenced the reliability of the conclusions, we are retracting this article to protect the integrity and accuracy of the scientific record.Signed: Chaozhi Wang, Jiang Li and Shaoyun Guoth April 2021Date: 5RSC AdvancesRetraction endorsed by Laura Fisher, Executive Editor,


Desirable carbon allotropes such as graphene oxide (GO) have entered the field with several biomedical applications, owing to their exceptional physicochemical and biological features, including extreme strength, found to be 200 times stronger than steel; remarkable light weight; large surface-to-volume ratio; chemical stability; unparalleled thermal and electrical conductivity; and enhanced cell adhesion, proliferation, and differentiation properties. The presence of functional groups on graphene oxide (GO) enhances further interactions with other molecules. Therefore, recent studies have focused on GO-based materials (GOBMs) rather than graphene. The aim of this research was to highlight the physicochemical and biological properties of GOBMs, especially their significance to biomedical applications. The latest studies of GOBMs in biomedical applications are critically reviewed, and in vitro and preclinical studies are assessed. Furthermore, the challenges likely to be faced and prospective future potential are addressed. GOBMs, a high potential emerging material, will dominate the materials of choice in the repair and development of human organs and medical devices. There is already great interest among academics as well as in pharmaceutical and biomedical industries. Trauma, aging, and disease cause damage to tissue, which can result in loss of function. In the early stages, self-healing can be effective, but with substantial injury, the organ cannot restore itself sufficiently; thus, organ repair or replacement  is recommended, albeit with some challenges. The most critical challenges facing organ replacements are organ shortages, grafts, transplant rejection, high cost, inflammation, infection, and also death in some cases . AlternaCarbon-based nanomaterials have recently boomed in biomaterial applications. Graphene is an important new addition to these carbon family materials due to its unique properties. Researchers in tissue engineering have already extensively investigated graphene-containing structures, specifically for bone, neuronal, cardiac, skin, cartilage, and dental tissue regeneration, as highlighted in graphical abstract. Carbon-based nanomaterials have recently boomed in biomaterial applications. Graphene is an important new addition to these carbon family materials due to its unique properties. Researchers in tissue engineering have already extensively investigated graphene-containing structures, specifically for bone, neuronal, cardiac, skin, cartilage, and dental tissue regeneration, as highlighted in 2-hybridized carbon atoms arranged in a hexagonal lattice-like honeycomb. Valuable research has completely described the synthesis method, but in brief, the synthesis approach was divided into top-down and bottom-up strategies. In the top-down approach, graphite is treated by exfoliation . The bottom-up approach is a developed approach, which contains epitaxial growth, chemical vapor deposition, and the dry ice method for graphene material production [In this review, graphene oxide (GO), reduced graphene oxide (rGO), and functionalized GO (FGO) are noted among the various allotropic forms of carbons due to their significant surface area, strength, light weight, chemical stability, enhanced cell adhesion, proliferation, differentiation, and application in the repair of tissue ,5. The boduction ,7,8  graphene lattice structures have some shortcomings, such as an unstable chemical structure and limited active sites for interacting with other molecules or nanomaterials; hence, it has some incompatibility. The chemical modification of graphene and the production of GO resolves some of these problems. GO maintains graphene’s atomic configuration and only has carboxyl groups (-OOH) on the edge of its structure, as well as epoxy (-O) and hydroxyl (-OH) groups on the basal plane. By reducing the amount of oxygen in GO through thermal, chemical, or UV exposure processes, rGO is produced. Most often, the reduction of GO does not complete, and some oxygen groups remain because not all sptructure . rGO protructure . Thus, t2 with the addition of π orbitals. In each unit cell of graphene, two π orbitals exist that are dispersed to form two π bonds, both of which could be known as bonding and antibonding [2 reduces its mechanical, electrical, and thermal properties [The distinctive properties of graphene are derived from its particular crystal lattice structure. Within this, the bonding between each carbon atom is hybridized spibonding . This aribonding . The graibonding ,14. Grapibonding . In GO, operties ,14. Althoperties .P. aeruginosa and S. aureus). GO and rGO, owing to the existence of functionalized groups, can alter the partial pressure of intracellular oxygen, which results in oxidative damage that destroys the bacterial cell internal composition, particularly E. coli, through the deactivation of their proteins and lipids, which eventually leads to cell death. GO has illustrated synergistic effects with laser energy; hence, it has been used for photothermal therapy, directly enhancing its antibacterial activity [2 carbons play an essential role in scavenging radicals by radical adduct formation and electron transfer. Because of this characteristic, these biomaterials can effectively scavenge radicals and protect cells from high levels of oxidative stress [It has been proven that most GO and derivatives are cytocompatible in vitro and in vivo. However, the physicochemical properties of 2D materials, such as structure, shape, size, surface functionality, concentration, and aggregation state have an essential impact on cellular behavior. Graphene, with its sharp edge properties, has the potential to cause cell damage during the penetration of cell membranes. Its aggregation can also lead to cytotoxicity. Graphene at the nanoscale, when <100 nm, results in cytotoxicity, inflammation, and even genotoxicity (due to facing less steric hindrance). In contrast, graphene with functionalized groups  is easily internalized by cells , in addition to causing more irregular cell membrane perturbation ,17. GO aactivity . Anothere stress . Graphene stress . It is re stress ,22,23.GO and its derivatives have been added to other biomaterials either as filler to enhance their properties or as a conjugate during synthesis. The latter makes the nanocomposite materials much stronger. Below are examples of the applications of graphene derivatives in the repair and replacement of human organs .Having drastic proliferation and differentiation of stem cells into the specific tissue lineage, considered as the most critical features of an ideal scaffold. As mentioned before, GBMs have considerable potential for cells stimulating in stem cell-based therapy. In −2 S/m [0.25Co1.257W0.25S3 hybridized with GO (MCWS/GO), was prepared, which led to successful nerve regeneration and recovery within 2–5 weeks after MCWS/GO transplantation [Electrical signals and external stimuli enhance tissue regeneration for excitable tissue, specifically nerve, muscle, and cardiac tissue. Depolarization and repolarization take place between the two sides of both nerve and muscle cell membranes under an action potential, which leads to their contracting activity and response to electrical signals. Conductive materials, such as GOBMs, with similar electrical conductivity to native tissue, are considered to be promising scaffolds that stimulate cell proliferation and differentiation in stem-cell-based therapy, with decreased cytotoxicity and improved mechanical properties. They are better electron transmitters in comparison to other electronic materials, such as carbon nanotubes . Much ef−2 S/m . The LBLantation .Similar to the nerves, muscles  have an electrical stimuli-responsive characteristic; thus, the utilization of conductive biomaterials was proposed. There was no difference in cell adhesion and cellular growth in a random and aligned GO electrospun mat, which was modified with oxygen plasma . GO-poly3); hence, conductive biomaterials  are potential candidates that can mimic the native extracellular matrix (ECM) for cell growth, promote electrical signal conduction, and preserve electrical coupling between the myocardium cells [As mentioned, cardiac muscles are electrically conductive . The results indicated rGO enhances bone regeneration, although when the rGO percentage exceeded a certain threshold level (≥100 μg/mL), it became severely cytotoxic, [The bone has a prominent ability to regenerate; however, the human skeleton has a limited capacity of self-regeneration when bone defects are large enough or critical sized. Several different approaches have been utilized for bone tissue engineering (BTE); the most common were those that tried to mimic the natural process of bone repair using 3D osteoconductive scaffolds. Recently, the potential of GOBMs has gained tremendous attention to facilitate and improve BTE ,57,58,59totoxic,  which cototoxic, ,28,60,64totoxic, . A key ctotoxic, .The skin acts as a vital barrier, blocking the entrance of dangerous foreign matter into the body . BecauseCartilage is an avascular tissue that has limited self-healing potential compared to other tissues . Thus, u2) and GO in dental implants resulted in healthier surrounding tissue and fewer bacteria growth, in addition to possessing enhanced mechanical features  [2O2) and GOBM nanocomposites. This combination enhanced the whitening effect of H2O2 and decreased treatment time [Most studies in dental tissue engineering have emphasized the role of GOBMs in the osteogenic differentiation of stem cells after dental implantation, which is an essential function in regenerative dentistry. The potential of GO was demonstrated to induce the odontoblastic or osteogenic differentiation of dental pulp stem cells (DPSCs) without the use of any chemical inductors . However by 41%) . The com by 41%) . Barrier by 41%) . The lasent time .Graphene oxide has raised considerable interest in tissue engineering and regenerative medicine due to its specific characteristics, e.g., exceptional mechanical properties and electrical conductivity, as well as physiochemical, antibacterial, and biological capabilities. The biological features of GOBMs depend on many parameters, including concentration, size, shape, and surface chemistry. Within GOBMs, FGO and rGO exhibited the greatest antibacterial capability. Moreover, GOBMs in various forms of 2D and 3D structures could stimulate the proliferation and differentiation of cells to a specific lineage, designated to chemical reactions with biomolecules. Growth factors, small molecules, and ECM proteins force cell differentiation via GOBMs’ π–π and hydrophobic interactions. The main challenges associated with utilizing GOBMs are toxicity, the lack of a detailed mechanism of GOBMs’ biological features, and the explanation of the biological pathway, which has still not been clarified in depth. Moreover, most research has concentrated on toxicity at the cellular level instead of at the genetic level. GOBMs can interact with different biomolecules, especially the DNA; therefore, studies are needed to elucidate the apoptosis pathway. ROS production, which causes cell death, has been studied more, along with lung, liver, intestine, and kidney tissues, than other tissues, e.g., nerve, muscle, cardiac, bone, skin, cartilage, and dental tissue. The application of GOBMs in biomedicine is summarized in The long-term toxicity of GOBMs, known as a primary hurdle, has limited experimental data, and further research investigating their effects is necessary . This un


The physiologic function of tripartite motif protein 56 (TRIM56), a ubiquitously expressed E3 ligase classified within the large TRIM protein family, remains elusive. Gene knockdown studies have suggested TRIM56 as a positive regulator of the type I interferon (IFN-I) antiviral response elicited via the Toll-like receptor 3 (TLR3) and cyclic GMP–AMP synthase (cGAS)–stimulator of interferon genes (STING) pathways, which detect and respond to danger signals—extracellular double-stranded (ds) RNA and cytosolic dsDNA, respectively. However, to what extent these pathways depend on TRIM56 in human cells is unclear. In addition, it is debatable whether TRIM56 plays a part in controlling the expression of IFN-stimulated genes (ISGs) resulting from IFN-I based antiviral treatment. In this study, we created HeLa-derived TRIM56 null cell lines by gene editing and used these cell models to comprehensively examine the impact of endogenous TRIM56 on innate antiviral responses. Our results showed that TRIM56 knockout severely undermined the upregulation of ISGs by extracellular dsRNA and that loss of TRIM56 weakened the response to cytosolic dsDNA. ISG induction and ISGylation following IFN-α stimulation, however, were not compromised by TRIM56 deletion. Using a vesicular stomatitis virus-based antiviral bioactivity assay, we demonstrated that IFN-α could efficiently establish an antiviral state in TRIM56 null cells, providing direct evidence that TRIM56 is not required for the general antiviral action of IFN-I. Altogether, these data ascertain the contributions of TRIM56 to TLR3- and cGAS–STING-dependent antiviral pathways in HeLa cells and add to our understanding of the roles this protein plays in innate immunity. Tripartite motif protein 56 (TRIM56) is a member of the large TRIM protein family of E3 ligases that are involved in a broad array of host processes, including, but not limited to, proliferation, differentiation, development and, recently, immunity ,5,6,7. Aper se is moderately upregulated by IFN-I [In addition to its reported antiviral activities, TRIM56 has also been implicated in regulating innate immune-signaling pathways that culminate in the induction of type I interferon (IFN-I) response, a hallmark of the intrinsic, immediate defense mechanisms of mammalian hosts against viral infections. In keeping with this, TRIM56 expression by IFN-I ,5,9, as by IFN-I , HeLa [5by IFN-I . SeparatThere also is a discrepancy regarding whether TRIM56 has a role in modulating the induction of IFN-stimulated genes (ISGs), whose products act in concert to execute the antiviral actions of IFNs. In immunoblotting experiments, Shen et al. found that upregulation of representative ISGs by IFN-α was normal in TRIM56 knockdown cells . The latIn this study, we set out to create TRIM56-deficient cell lines by CRISPR/Cas9 gene editing, and to address the requirements for this protein in various innate immune pathways in human cells. We report that TRIM56 deletion severely compromises antiviral gene expression induced via the TLR3 signaling pathway and that its loss weakens the response through the cytosolic DNA-sensing pathway. By contrast, our data do not support a significant role, if any, of TRIM56 in regulating ISG induction downstream of the IFN-I receptors or impacting the establishment of a general antiviral state by IFN-I.The retroviral vector pCX4pur-FH-TRIM56 encoding N-terminally Flag- and HA-tandem tagged human TRIM56 (FH-T56) in the pCX4pur backbone has been described previously . The twoHeLa and Vero cells  were maintained in Dulbecco’s modified Eagle medium supplemented with 10% fetal bovine serum (FBS), 100 U/mL penicillin, and 100 μg/mL streptomycin. To create TRIM56 null cell lines by CRISPR/Cas9 gene editing, HeLa cells were co-transfected with TRIM56_172121 and TRIM56_172123 constructs at a 1:1 ratio. Forty-eight hours later, cells were dissociated from the culture plate via trypsin digestion to form single cell suspension and subsequently reseeded onto 96-well plates at a cell density of ~0.5 cells per well. The plates were returned to culture and inspected periodically for wells that contained a single colony. After ~3 weeks, individual cell colonies were expanded and screened for TRIM56 expression, in comparison with parental HeLa cells, by Western blot. Two independent cell lines, clone 17 (referred to as T56-KO#1) and clone 3 (referred to as T56-KO#2), were found to completely lack TRIM56 protein and selected for further analyses.To stably express Flag- and HA-tandem tagged TRIM56 (FH-T56) in TRIM56 null cells, we transduced T56-KO#1 and T56-KO#2 cells with a replication-incompetent retrovirus packaged from pCX4pur-FH-TRIM56. Following selection in puromycin-containing medium, surviving cell colonies were pooled, designated HeLa-T56-KO#1-FHT56 and HeLa-T56-KO#2-FHT56 cells, respectively, and used for analyses.Poly(I:C), poly(dA:dT), Calf Thymus DNA (C-T DNA), and recombinant human IFN-α 2b were obtained from Sigma . VSV-Luc, a recombinant, firefly luciferase (FLuc)-encoding vesicular stomatitis virus, was a gift from Sean Whelan . VSV-LucCells were stimulated by various PRR ligands for the indicated time periods to elicit innate immune responses—specifically for this study, ISG expression. To engage the TLR3 pathway, poly(I:C) was added directly into culture medium at a final concentration of 20–50 μg/mL. To activate the cytosolic DNA-sensing pathway, poly(dA:dT) or C-T DNA was transfected into cells at 3 μg per well of a 6-well plate after being complexed with Lipofectamine 2000 at a 1:1 (μg:μL) ratio. To determine ISG induction by IFN-α, cells were incubated with recombinant IFN-α at the indicated concentration for 6 and 12 h, respectively.To directly gauge the efficacy of IFN-mediated establishment of antiviral state, cells were mock-stimulated or stimulated by IFN-α at indicated concentrations for 16 h, followed by challenge with VSV-Luc at an MOI of 0.1. At 8 h post infection, cells were lysed for firefly luciferase assay as a readout for VSV replication.Total RNA was extracted from cells following various treatments using TRIzol (Invitrogen) as per the manufacturer’s protocol. Complementary DNA synthesis by reverse transcription and SYBR green-based quantitative PCR (qPCR) were implemented as described previously ,14. BrieE. coli) [® 680RD or goat anti-rabbit IgG IRDye® 800CW , protein bands were imaged with an Odyssey infrared imaging system (LI-COR Biosciences). To compare target protein expression levels across different samples in immunoblotting, the signal intensity of protein band(s) of interest was determined by Image Studio Lite (LI-COR Biosciences) and normalized to that of a loading control, as indicated.Cell lysates were prepared in RIPA buffer, quantified for protein concentrations, and subjected to SDS-PAGE and immunoblotting as previously described ,16. The E. coli) . Followit-test . A p value of <0.05 was regarded as significant.All results are presented as means ± standard deviations. Statistical differences between two groups were analyzed using a two-tailed Student’s To clarify the roles of TRIM56 expressed at physiologic levels in innate immune signaling, we performed CRISPR/Cas9-mediated gene editing to eliminate TRIM56 expression in HeLa cells. HeLa was chosen as the founder to create TRIM56 KO cells because this well-characterized cell line (1) has been widely used as a cell culture model to study innate immunity against viral infections, (2) possesses intact dsRNA- and dsDNA-sensing antiviral pathways including those dependent on TLR3 ,18, and Next, we investigated the effect of TRIM56 deficiency on TLR3 signaling by comparing HeLa-T56-KO#1 and -KO#2 with parental HeLa cells for ISG induction following poly(I:C) stimulation via the extracellular route, which engages specifically the TLR3 pathway . ImmunobWe considered the remote chance that the profound impairment in TLR3 response of the two TRIM56 null cell lines was intrinsic to the cell clones selected, independent of TRIM56 deletion. To exclude this possibility, we reconstituted TRIM56 expression in HeLa-T56-KO#1 and -KO#2 cells by retroviral gene transfer of FLAG- and HA-tandem tagged human TRIM56 (FH-T56) and compared the reconstituted cells with their untransduced counterparts for ISG induction by extracellular poly(I:C). We chose an early time point (8 h) post stimulation, when the response was still in its climbing phase, for immunoblotting detection of two representative ISGs, IFIT3 and IFIT1. This strategy avoided the plateau phase of the TLR3 response, ensuring that any difference between T56-KO and -reconstituted cells would be captured. Consistent with our earlier data, there was little induction of either ISG by poly(I:C) in T56-KO#1 B. ConsisSeveral previous studies have shown that RNAi-mediated knockdown of TRIM56 undermines the IFN response triggered by cytosolic delivery of DNA ,5,6, whiNext, we determined whether cells with and without endogenous TRIM56 expression differed in their response to IFN-I. HeLa and the two T56-KO cell lines were stimulated by high (100 U/mL) and low (10 U/mL) concentrations of IFN-α for 6 and 12 h, respectively, and subsequently lysed for RNA extraction and RT-qPCR analysis of the expression for three representative ISGs, MDA5, IFIT1, and OAS1. As shown in ISGylation, characterized by a ubiquitination-like process covalently linking ISG15 to a subset of protein targets, contributes to IFN-mediated antiviral protection against some, but not all, viruses . One memTo directly gauge the overall impact, if any, that TRIM56 may have on IFN-mediated establishment of an antiviral state, we conducted VSV challenge experiments in cells with and without prior IFN-α stimulation. VSV is a negative-strand RNA virus highly sensitive to the antiviral action of IFNs. As such, it has been widely used as a tool virus for IFN bioactivity assays. We took advantage of VSV-Luc, a recombinant VSV whose replication can be conveniently and quantitatively monitored by measuring activities of the firefly luciferase reporter the virus encodes, in infected cells. HeLa, HeLa-T56-KO#1, and -KO#2 cells were pre-incubated with low (10 U/mL) and high (100 U/mL) concentrations of IFN-α for 16 h, respectively, or left unstimulated, followed by infection with VSV-Luc (MOI = 0.1) for 8 h. As shown in In this study, we have created two independent TRIM56 null cell lines from HeLa cells by CRISPR/Cas9 gene editing. Using these TRIM56-deficient cell models and their parental wild-type counterpart, we systematically investigated the influence of physiologic level of TRIM56 on innate antiviral responses, which TRIM56 has been suggested to regulate by several recent studies based on gene knockdown experiments. As discussed below, data described herein help clarify and add to our understanding of the various roles that TRIM56 plays in host intrinsic immune mechanisms fending off viruses.In both TRIM56 knockout cell lines, the induction of multiple ISGs following exposure to extracellular dsRNA, which is predominantly mediated via the endocytic TLR3-TRIF pathway , was proWe also observed impaired ISG expression in TRIM56 null cells following stimulation by cytosolic dsDNA , which iThe upregulation of ISGs with antiviral activities is an important mechanism by which IFNs hinder viral replication in infected cells and protect uninfected surrounding cells . In an e


In the new era of modern flexible and bendable technology, graphene-based materials have attracted great attention. The excellent electrical, mechanical, and optical properties of graphene as well as the ease of functionalization of its derivates have enabled graphene to become an attractive candidate for the construction of flexible devices. This paper provides a comprehensive review about the most recent progress in the synthesis and applications of graphene-based composites. Composite materials based on graphene, graphene oxide (GO), and reduced graphene oxide (rGO), as well as conducting polymers, metal matrices, carbon–carbon matrices, and natural fibers have potential application in energy-harvesting systems, clean-energy storage devices, and wearable and portable electronics owing to their superior mechanical strength, conductivity, and extraordinary thermal stability. Additionally, the difficulties and challenges in the current development of graphene are summarized and indicated. This review provides a comprehensive and useful database for further innovation of graphene-based composite materials.  It is well known that materials play an important role in the development of science and technology, because the realization of a new technology often requires the support of novel materials. Therefore, exploring materials with excellent properties has always been an important subject of scientific research. A remarkable material, graphene, has attracted widespread attention since it was first exfoliated from graphite by Andre Geim and Konstantin Novoselov in 2004. As a result of its prominent performances, graphene can be used in various fields, such as energy storage, biosensing, optoelectronics, flexible electronics, electrochemical sensing, robotics, textile industry, and so on ,2,3,4. TGraphene with a thickness of a single carbon atom is arranged in a honeycomb lattice. It is very solid and can be fashioned into 0D, 1D, 3D forms  6]. In . In 6]. Graphene and its derivatives have their own unique advantages and can be used in many domains applying different techniques, such as thermal chemical vapor deposition (CVD), self-assembly technique, spin coating, vacuum filtration, thermal decomposition, solution dispersion technique, and chemical decomposition polymer processing technique ,13,14,15There are many reports about the synthesis of GO, and the structures of the obtained products are slightly different . One of Several attempts have been adopted to study the synthesis methods of carbon-based materials. The very first attempt dates back to 1962. Boehm et al. prepared soot composed of thin-layer graphite-intercalated compounds by the reduction and combustion of graphite oxide. In 1944, these products were named as graphene platelets and had a single carbon layer . In 2004Micromechanical cleavage is a process in which the bonds in graphite crystal are broken by mechanical energy so that graphene sheets are peeled from a silicon substrate. Exfoliation can be done in solution by intercalating graphite and exfoliating to a single carbon sheet . GO prepThe CVD technique is another common method used to synthesize graphene. Certain carrier gases and carbon-based precursors like camphor and methane are injected into a CVD chamber at a specific temperature. Then, the carbon precursor is decomposed to form graphene on transition metal sheets such as a nickel foam . In addi4, ascorbic acid, and HI can be used. Among these, ascorbic acid is essential for the scalable production of rGO. The chemical procedure to obtain rGO using ascorbic acid as a reducing agent is shown in GO prepared by Hummer’s method consists of a-few-layer carbon platelets decorated with oxygen containing functional groups. The removal of some oxygen-based groups by reducing agents or thermal treatment can yield rGO . The mai4 was added to an NaOH solution under stirring followed by heating at 160 °C for 3 h. Next, the solution was treated with 2 M sulphuric acid and ethyl glycol and then was electrochemically deposited on a graphene-filtrated carbon cloth/graphene paper substrate. It is worth noting that this flexible composite material is expected to be used in energy storage, because the square shape of the corresponding electrochemical graph indicates excellent capacitive properties [With the evolution of science and technology, more and more novel materials with fascinating properties have been discovered and can be applied in many domains. Among these novel materials, graphene has received a lot of attention because of its excellent properties, such as high mechanical strength, stability, charge storage capacity, etc. Furthermore, graphene has very good flexibility and shows excellent application prospects in some flexible composite materials. For instance, a flexible composite consisting of polyethylene–ioxythiophene–graphene was fabricated by the following method. First, PtCloperties . Likewisoperties . 2 on Zn2SnO4 (ZTO) nanowires grown on carbon microfibers. This material can be used in supercapacitor electrodes, whose composite analysis suggests a long cycle life [2 combined with light-weight carbon nanotubes (CNTs) formed an ultra-flexible thin-film composite, which has been used for various energy storage devices as a robust electrode, as it holds extraordinary mechanical properties with superb electrochemical activities when fabricated by the chemical co-precipitation method [Many additional related reports on the preparation and application of other flexible graphene-based composites have been published . For exacle life . A typiccle life . Graphenn method . 3 solution, which helped to control the diameter of fiber, finally obtaining graphene/PPy fibers [2 can also be used for the fabrication of this composite due to its high specific capacitance. A 3D graphene/MnO2 composite foam to be used as a negative electrode for asymmetric supercapacitors was fabricated by the solution casting method. GO was reduced on Ni foam and then subjected to electrodeposition of MnO2 to obtain an asymmetric supercapacitor, showing excellent cyclic stability /graphene, polyethylene terephthalate/graphene, and polycarbonate/graphene nanocomposites have been fabricated through in situ intercalative polymerization, solution intercalation, as well as melt intercalation ,45. In aeduction . Beside eduction . Likewispacitors . To attal method .Although the material choice for certain application remains crucial, the choice of the substrate has a great effect on flexibility. Textile fibers, carbon cloth, and paper pulp have evolved as excellent substrate choices for various graphene-based composites. For example, a graphene-based carbon cloth composite fabricated by the simple brush coating technique showed great properties as an electrode material . Light wThe composite of vanadium oxide with graphene paper is binder-free and shows versatility. The fabrication follows an alkaline deoxygenation process, which is more suitable than the chemical reduction of GO to graphene. This flexible composite paper membrane possesses remarkable advantages for double-layered and pseudocapacitive electrodes . In ordeMoreover, combing graphene with some common substances in nature can lead to composites with outstanding performances. As shown in A nano nickel oxide/graphene PANI composite with enhanced cyclic stability and a high specific capacitance of 92% after 2500 charge–discharge cycles can be applied in the fabrication of energy storage devices . Consequ2 composite membrane could be used as a filtration membrane for the removal of water impurities. The composite was fabricated by vacuum filtration and allowed a moderate water purification [The designed flexible composites should have not only excellent flexibility but also a certain strength which would enable the composites to withstand external environmental factors. Recently, a layered composite of GO/PVC with large mechanical strength fabricated by the vacuum filtration method was reported . These ffication . Althougfication . Yarns wfication .2O4 and polyvinylidene fluoride fabricated by a simple sonication method turned out to be a good absorber of harmful microwaves in the electromagnetic spectrum [2O5/polyindole and activated carbon cloth were used as cathodic and anodic electrodes of an asymmetric supercapacitor. V2O5 nanostructures were constructed on a carbon cloth by in situ polymerization and showed good cyclic stability on testing  is widely studied as electrode material for supercapacitors. Recently, a W18O49 nanowires (NWs)/rGO nanocomposite, which can act as the negative electrode in asymmetric supercapacitor devices, was prepared from the precursors WCl6 and GO by the solvothermal method [18O49 NWs-rGO//rGO showed high specific capacitance and excellent cycling stability. For the fabrication of paper-based electrode, the incorporation of celluloses and pulps is desirable to attain flexibility and stability. GO-based nanocomposite of nanocrystalline cellulose acetate fabricated via stirring and a solvent casting method showed high thermal stability and good mechanical strength [2, whereas further reduction of the composite to rGO/TiO2 was obtained by stirring and drying. Anatase TiO2 exhibits higher power and energy density than other conventional metal oxides [Organic substrates have been widely used in recent studies, but metallic substrates remain an important choice. Copper metallic foil acted as a substrate for the growth of rGO/Cupacitors . Howeverpacitors . The choechnique . Chong eechnique  preparedl method . The asystrength . Likewisstrength . An outsl oxides . In compl oxides . AltogetWith the fast development of portable, wearable, and lightweight electronic devices, highly efficient and flexible energy strategies are urgently needed. In order to achieve this goal, it is crucial to explore novel materials. Graphene has attracted tremendous attention in the field of material science due to its outstanding properties since it was first exfoliated in 2004. This review aimed to outline the different fabrication methods and applications of graphene-based materials, especially for flexible, portable, environment-friendly, and cost-effective energy storage and conversion devices. Some representative methods used to prepare the composites based on graphene and the corresponding applications are listed in


We hypothesized that we could leverage social media to recruit learners to a gamification-infused ID knowledge competition, and entice them to explore additional online educational resources.We created the ID Fellows Cup, a knowledge-based trivia competition, to engage Infectious Diseases fellows. The game was crafted via Kaizen-Education, a software platform developed at the University of Alabama at Birmingham, that uses gamification to engage learners. Multiple choice questions including figures and/or text are presented to learners, followed by detailed teaching explanations. 60 questions emphasizing high-yield concepts were delivered over 4 weeks. Questions were written by fellows and reviewed by faculty at three programs. Elements of gamification  were included to enhance engagement. Recruitment strategies included Twitter, program director emails, and peer-to-peer. We measured game statistics and participation. Learners were invited to complete a post-game survey about their experience. Table 1 shows our game statistics with broad geographic reach including 42 programs. Most fellows matriculated in 2019 or 2020; the number of US ID fellows equaled 17% of those completing ID in-training exam. Recruitment sources included 44% co-fellow, 42% Twitter, and 15% Program Director. Through 20 days with questions, we had 155 daily average users. Overall, fellows answered 11,419 total questions, representing 89% of all released questions. Of 103 responses to post-game survey (table 2) 97% would participate again and all felt the game was a good use of their time. Over 80% of participants reported some engagement with linked resources included in the answer explanations. In general, 78% felt engagement with online resources increased subsequent to participating in the game, including learning about at least one new online resource. We leveraged social media and gamification to effectively engage, and stimulate ID learners to explore additional online educational resources. Technology enriched learning, helps supplement and globalize ID education, making it as diverse and engaging as our field.Todd P. McCarty, MD, Cidara (Grant/Research Support)GenMark T2 Biosystems (Consultant) Prathit A. Kulkarni, M.D., Vessel Health, Inc. (Grant/Research Support)