Current application and future perspective of CRISPR/cas9 gene editing system mediated immune checkpoint for liver cancer treatment.

Liver cancer, which is well-known to us as one of human most prevalent malignancies across the globe, poses a significant risk to live condition and life safety of individuals in every region of the planet. It has been shown that immune checkpoint treatment may enhance survival benefits and make a significant contribution to patient prognosis, which makes it a promising and popular therapeutic option for treating liver cancer at the current time. However, there are only a very few numbers of patients who can benefit from the treatment and there also exist adverse events such as toxic effects and so on, which is still required further research and discussion. Fortunately, the clustered regularly interspaced short palindromic repeat/CRISPR-associated nuclease 9 (CRISPR/Cas9) provides a potential strategy for immunotherapy and immune checkpoint therapy of liver cancer. In this review, we focus on elucidating the fundamentals of the recently developed CRISPR/Cas9 technology as well as the present-day landscape of immune checkpoint treatment which pertains to liver cancer. What's more, we aim to explore the molecular mechanism of immune checkpoint treatment in liver cancer based on CRISPR/Cas9 technology. At last, its encouraging and powerful potential in the future application of the clinic is discussed, along with the issues that already exist and the difficulties that must be overcome. To sum up, our ultimate goal is to create a fresh knowledge that we can utilize this new CRISPR/Cas9 technology for the current popular immune checkpoint therapy to overcome the treatment issues of liver cancer.

Neurophysiological effective network connectivity supports a threshold-dependent management of dynamic working memory gating.

To facilitate goal-directed actions, effective management of working memory (WM) is crucial, involving a hypothesized WM "gating mechanism." We investigate the underlying neural basis through behavioral modeling and connectivity assessments between neuroanatomical regions linked to theta, alpha, and beta frequency bands. We found opposing, threshold-dependent mechanisms governing WM gate opening and closing. Directed beta band connectivity in the parieto-frontal and parahippocampal-occipital networks was crucial for threshold-dependent WM gating dynamics. Fronto-parahippocampal connectivity in the theta band was also notable for both gating processes, although weaker than that in the beta band. Distinct roles for theta, beta, and alpha bands emerge in maintaining information in WM and shielding against interference, whereby alpha band activity likely acts as a "gatekeeper" supporting processes reflected by beta and theta band activity. The study shows that the decision criterion for WM gate opening/closing relies on concerted interplay within neuroanatomical networks defined by beta and theta band activities.

Recent research progress based on ferroptosis-related signaling pathways and the tumor microenvironment on it effects.

The existing therapies for cancer are not remote satisfactory due to drug-resistance in tumors that are malignant. There is a pressing necessity to take a step forward to develop innovative therapies that can complement current ones. Multiple investigations have demonstrated that ferroptosis therapy, a non-apoptotic modality of programmed cell death, has tremendous potential in face of multiple crucial events, such as drug resistance and toxicity in aggressive malignancies. Recently, ferroptosis at the crosswalk of chemotherapy, materials science, immunotherapy, tumor microenvironment, and bionanotechnology has been presented to elucidate its therapeutic feasibility. Given the burgeoning progression of ferroptosis-based nanomedicine, the newest advancements in this field at the confluence of ferroptosis-inducers, nanotherapeutics, along with tumor microenvironment are given an overview. Here, the signaling pathways of ferroptosis-related were first talked about briefly. The emphasis discussion was placed on the pharmacological mechanisms and the nanodrugs design of ferroptosis inducing agents based on multiple distinct metabolism pathways. Additionally, a comprehensive overview of the action mechanisms by which the tumor microenvironment influences ferroptosis was elaborately descripted. Finally, some limitations of current researches and future research directions were also deliberately discussed to provide details about therapeutic avenues for ferroptosis-related diseases along with the design of anti-drugs.

Inhibitory control in WM gate-opening: Insights from alpha desynchronization and norepinephrine activity under atDCS stimulation.

Our everyday activities require the maintenance and continuous updating of information in working memory (WM). To control this dynamic, WM gating mechanisms have been suggested to be in place, but the neurophysiological mechanisms behind these processes are far from being understood. This is especially the case when it comes to the role of oscillatory neural activity. In the current study we combined EEG recordings, and anodal transcranial direct current stimulation (atDCS) and pupil diameter recordings to triangulate neurophysiology, functional neuroanatomy and neurobiology. The results revealed that atDCS, compared to sham stimulation, affected the WM gate opening mechanism, but not the WM gate closing mechanism. The altered behavioral performance was associated with specific changes in alpha band activities (reflected by alpha desynchronization), indicating a role for inhibitory control during WM gate opening. Functionally, the left superior and inferior parietal cortices, were associated with these processes. The findings are the first to show a causal relevance of alpha desynchronization processes in WM gating processes. Notably, pupil diameter recordings as an indirect index of the norepinephrine (NE) system activity revealed that individuals with stronger inhibitory control (as indexed through alpha desynchronization) showed less pupil dilation, suggesting they needed less NE activity to support WM gate opening. However, when atDCS was applied, this connection disappeared. The study suggests a close link between inhibitory controlled WM gating in parietal cortices, alpha band dynamics and the NE system.

Synergistic anti-tumor effect of dual drug co-assembled nanoparticles based on ursolic acid and sorafenib.

Both ursolic acid (UA) and sorafenib (Sora) have been generally utilized in cancer treatment, and the combination of the two has also shown a good anti-tumor effect. However, single-agent therapy for Hepatocellular carcinoma (HCC) has the disadvantages of multi-drug resistance, poor water solubility and low bioavailability, and the application of traditional nanocarrier materials is limited due to their low drug loading and low carrier-related toxicity. Therefore, we prepared US NPs with different proportions of UA and Sora by solvent exchange method for achieving synergistic HCC therapy. US NPs had suitable particle size, good dispersibility and storage stability, which synergistically inhibited the proliferation of HepG2 cells, SMMC7721 cells and H22 cells. In addition, we also proved that US NPs were able to suppress the migration of HepG2 cells and SMMC7721 cells and reduce the adhesion ability and colony formation ability of these cells. According to the results, US NPs could degrade the membrane potential of mitochondrial, participate in cell apoptosis, and synergistically induce autophagy. Collectively, the carrier-free US NPs provide new strategies for HCC treatment and new ideas for the development of novel nano-drug delivery systems containing UA and Sora.

Research progress and application of the CRISPR/Cas9 gene-editing technology based on hepatocellular carcinoma.

Hepatocellular carcinoma (HCC) is now a common cause of cancer death, with no obvious change in patient survival over the past few years. Although the traditional therapeutic modalities for HCC patients mainly involved in surgery, chemotherapy, and radiotherapy, which have achieved admirable achievements, challenges are still existed, such as drug resistance and toxicity. The emerging gene therapy of clustered regularly interspaced short palindromic repeat/CRISPR-associated nuclease 9-based (CRISPR/Cas9), as an alternative to traditional treatment methods, has attracted considerable attention for eradicating resistant malignant tumors and regulating multiple crucial events of target gene-editing. Recently, advances in CRISPR/Cas9-based anti-drugs are presented at the intersection of science, such as chemistry, materials science, tumor biology, and genetics. In this review, the principle as well as statues of CRISPR/Cas9 technique were introduced first to show its feasibility. Additionally, the emphasis was placed on the applications of CRISPR/Cas9 technology in therapeutic HCC. Further, a broad overview of non-viral delivery systems for the CRISPR/Cas9-based anti-drugs in HCC treatment was summarized to delineate their design, action mechanisms, and anticancer applications. Finally, the limitations and prospects of current studies were also discussed, and we hope to provide comprehensively theoretical basis for the designing of anti-drugs.

Three-pronged attacks by hybrid nanoassemblies involving a natural product, carbon dots, and Cu2+ for synergistic HCC therapy.

Tumor microenvironment (TME) stimuli-responsive nanoassemblies are emerging as promising drug delivery systems (DDSs), which acquire controlled release by structural transformation under exogenous stimulation. However, the design of smart stimuli-responsive nanoplatforms integrated with nanomaterials to achieve complete tumor ablation remains challenging. Therefore, it is of utmost importance to develop TME-based stimuli-responsive DDSs to enhance drug-targeted delivery and release at tumor sites. Herein, we proposed an appealing strategy to construct fluorescence-mediated TME stimulus-responsive nanoplatforms for synergistic cancer therapy by assembling photosensitizers (PSs) carbon dots (CDs), chemotherapeutic agent ursolic acid (UA), and copper ions (Cu2+). First, UA nanoparticles (UA NPs) were prepared by self-assembly of UA, then UA NPs were assembled with CDs via hydrogen bonding force to obtain UC NPs. After combining with Cu2+, the resulting particles (named UCCu2+ NPs) exhibited quenched fluorescence and photosensitization due to the aggregation of UC NPs. Upon entering the tumor tissue, the photodynamic therapy (PDT) and the fluorescence function of UCCu2+ were recovered in response to TME stimulation. The introduction of Cu2+ triggered the charge reversal of UCCu2+ NPs, thereby promoting lysosomal escape. Furthermore, Cu2+ resulted in additional chemodynamic therapy (CDT) capacity by reacting with hydrogen peroxide (H2O2) as well as by consuming glutathione (GSH) in cancer cells through a redox reaction, hence magnifying intracellular oxidative stress and enhancing the therapeutic efficacy due to reactive oxygen species (ROS) therapy. In summary, UCCu2+ NPs provided an unprecedented novel approach for improving the therapeutic efficacy through the three-pronged (chemotherapy, phototherapy, and heat-reinforced CDT) attacks to achieve synergistic therapy.

Auricular Transcutaneous Vagus Nerve Stimulation Specifically Enhances Working Memory Gate Closing Mechanism: A System Neurophysiological Study.

Everyday tasks and goal-directed behavior involve the maintenance and continuous updating of information in working memory (WM). WM gating reflects switches between these two core states. Neurobiological considerations suggest that the catecholaminergic and the GABAergic are likely involved in these dynamics. Both of these neurotransmitter systems likely underlie the effects to auricular transcutaneous vagus nerve stimulation (atVNS). We examine the effects of atVNS on WM gating dynamics and their underlying neurophysiological and neurobiological processes in a randomized crossover study design in healthy humans of both sexes. We show that atVNS specifically modulates WM gate closing and thus specifically modulates neural mechanisms enabling the maintenance of information in WM. WM gate opening processes were not affected. atVNS modulates WM gate closing processes through the modulation of EEG alpha band activity. This was the case for clusters of activity in the EEG signal referring to stimulus information, motor response information, and fractions of information carrying stimulus-response mapping rules during WM gate closing. EEG-beamforming shows that modulations of activity in fronto-polar, orbital, and inferior parietal regions are associated with these effects. The data suggest that these effects are not because of modulations of the catecholaminergic (noradrenaline) system as indicated by lack of modulatory effects in pupil diameter dynamics, in the inter-relation of EEG and pupil diameter dynamics and saliva markers of noradrenaline activity. Considering other findings, it appears that a central effect of atVNS during cognitive processing refers to the stabilization of information in neural circuits, putatively mediated via the GABAergic system.SIGNIFICANCE STATEMENT Goal-directed behavior depends on how well information in short-term memory can be flexibly updated but also on how well it can be shielded from distraction. These two functions were guarded by a working memory gate. We show how an increasingly popular brain stimulation techniques specifically enhances the ability to close the working memory gate to shield information from distraction. We show what physiological and anatomic aspects underlie these effects.

Neurophysiological principles of inhibitory control processes during cognitive flexibility.

Inhibitory control plays an indispensable role in cognitive flexibility. Nevertheless, the neurophysiological principles underlying this are incompletely understood. This owes to the fact that the representational dynamics, as coded in oscillatory neural activity of different frequency bands has not been considered until now-despite being of conceptual relevance. Moreover, it is unclear in how far distinct functional neuroanatomical regions are concomitantly involved in the processing of representational dynamics. We examine these questions using a combination of EEG methods. We show that theta-band activity plays an essential role for inhibitory control processes during cognitive flexibility across informational aspects coded in distinct fractions of the neurophysiological signal. It is shown that posterior parietal structures and the inferior parietal cortex seem to be the most important cortical region for inhibitory control processes during cognitive flexibility. Theta-band activity plays an essential role in processes of retrieving the previously inhibited representations related to the current task during cognitive flexibility. The representational content relevant for inhibitory processes during cognitive flexibility is coded in the theta frequency band. We outline how the observed neural mechanisms inform recent overarching cognitive frameworks on how flexible action control is accomplished.

The neural stability of perception-motor representations affects action outcomes and behavioral adaptation.

Actions can fail - even though this is well known, little is known about what distinguishes neurophysiological processes preceding errors and correct actions. In this study, relying on the Theory of Event Coding, we test the assumption that only specific aspects of information coded in EEG activity are relevant for understanding processes leading to response errors. We examined N = 69 healthy participants who performed a mental rotation task and combined temporal EEG signal decomposition with multivariate pattern analysis (MVPA) and source localization analyses. We show that fractions of the EEG signal, primarily representing stimulus-response translation (event file) processes and motor response representations, are essential. Stimulus representations were less critical. The source localization results revealed widespread activity modulations in structures including the frontopolar, the middle and superior frontal, the anterior cingulate cortex, the cuneus, the inferior parietal cortex, and the ventral stream regions. These are associated with differential effects of the neural dynamics preceding correct/erroneous responses. The temporal-generalization MVPA showed that event file representations and representations of the motor response were already distinct 200 ms after stimulus presentation and this lasted till around 700 ms. The stability of this representational content was predictive for the magnitude of posterror slowing, which was particularly strong when there was no clear distinction between the neural activity profile of event file representations associated with a correct or an erroneous response. The study provides a detailed analysis of the dynamics leading to an error/correct response in connection to an overarching framework on action control.

A ventral stream-prefrontal cortex processing cascade enables working memory gating dynamics.

The representation of incoming information, goals and the flexible processing of these are required for cognitive control. Efficient mechanisms are needed to decide when it is important that novel information enters working memory (WM) and when these WM 'gates' have to be closed. Compared to neural foundations of maintaining information in WM, considerably less is known about what neural mechanisms underlie the representational dynamics during WM gating. Using different EEG analysis methods, we trace the path of mental representations along the human cortex during WM gate opening and closing. We show temporally nested representational dynamics during WM gate opening and closing depending on multiple independent neural activity profiles. These activity profiles are attributable to a ventral stream-prefrontal cortex processing cascade. The representational dynamics start in the ventral stream during WM gate opening and WM gate closing before prefrontal cortical regions are modulated. A regional specific activity profile is shown within the prefrontal cortex depending on whether WM gates are opened or closed, matching overarching concepts of prefrontal cortex functions. The study closes an essential conceptual gap detailing the neural dynamics underlying how mental representations drive the WM gate to open or close to enable WM functions such as updating and maintenance.

Protocol to decode representations from EEG data with intermixed signals using temporal signal decomposition and multivariate pattern-analysis.

The electroencephalogram (EEG) is one of the most widely used techniques in cognitive neuroscience. We present a protocol showing how to combine a temporal signal decomposition approach (RIDE, Residue iteration decomposition) with multivariate pattern analysis (MVPA) to obtain insights into the temporal stability of representations coded in distinct informational fractions of the EEG signal. In this protocol, we describe pre-processing of human EEG data, followed by the set-up and use of MATLAB-based toolboxes for RIDE and MVPA analysis. For complete details on the use and execution of this protocol, please refer to Petruo et al. (2021).

How low working memory demands and reduced anticipatory attentional gating contribute to impaired inhibition during acute alcohol intoxication.

High-dose alcohol intoxication is commonly associated with impaired inhibition, but the boundary conditions, as well as associated neurocognitive/neuroanatomical changes have remained rather unclear. This study was motivated by the counterintuitive finding that high-dose alcohol intoxication compromises response inhibition performance when working memory demands were low, but not when they were high. To investigate whether this is more likely to be caused by deficits in cognitive control processes or in attentional processes, we examined event-related (de)synchronization processes in theta and alpha-band activity and performed beamforming analyses on the EEG data of previously published behavioral findings. This yielded two possible explanations: There may be a selective decrease of working memory engagement in case of relatively low demand, which boosts response automatization, ultimately putting more strain on the remaining inhibitory resources. Alternatively, there may be a decrease in proactive preparatory and anticipatory attentional gating processes in case of relatively low demand, hindering attentional sampling of upcoming stimuli. Crucially, both of these interrelated mechanisms reflect differential alcohol effects after the actual motor inhibition process and therefore tend to be processes that serve to anticipate future response inhibition affordances. This provides new insights into how high-dose alcohol intoxication can impair inhibitory control.

A role of the norepinephrine system or effort in the interplay of different facets of inhibitory control.

Inhibitory control has multiple facets, and one possible distinction can be made between 'inhibition of interferences' and the 'inhibition of actions'. Both facets of inhibitory control show an interdependency. Even though some neurophysiological processes underlying this interdependency have been examined, the role of neuro-modulatory processes in their interplay are not understood. In the current study, we examine the role of the norepinephrine (NE) system in these processes. We did so by combining a Go/Nogo and Simon task. We recorded the EEG and pupil diameter data as an indirect index of NE system activity during the task. EEG theta band activity data and pupil diameter data were then integrated after conducting a temporal signal decomposition of the EEG data. We show that particularly theta band activity coding stimulus-response translation processes associated with middle frontal cortices, but not stimulus-driven processes are modulated by the interplay between the 'inhibition of interferences' and the 'inhibition of actions'. Modulations in stimulus-response translation processes were systematically correlated with pupil-diameter responses. The pattern of correlations suggests that phasic NE system activity particularly modulates stimulus-response mapping processes during conflict monitoring in incongruent Nogo trials, which may explain behavioral performance effects. Phasic NE system activity reflects essential modulators of the interplay between the 'inhibition of interferences' and the 'inhibition of actions'.

Superior frontal regions reflect the dynamics of task engagement and theta band-related control processes in time-on task effects.

Impairment of cognitive performance is often observed in time-on tasks. Theoretical considerations suggest that especially prefrontal cortex cognitive control functions is affected by time-on-task effects, but the role of effort/task engagement is not understood. We examine time-on-task effects in cognitive control on a neurophysiological level using a working-memory modulated response inhibition task and inter-relate prefrontal neuroanatomical region-specific theta-band activity with pupil diameter data using EEG-beamforming approaches. We show that task performance declines with time-on tasks, which was paralleled by a concomitant decreases of task-evoked superior frontal gyrus theta-band activity and a reduction in phasic pupil diameter modulations. A strong relation between cognitive control-related superior frontal theta-band activity and effort/task engagement indexed by phasic pupil diameter modulations was observed in the beginning of the experiment, especially for tasks requiring inhibitory controls and demanding high working memory. This strong relation vanished at the end of the experiment, suggesting a decoupling of cognitive control resources useable for a task and effort invested that characterizes time-on-task effects in prefrontal cortical structures.

Time-On-Task Effects on Working Memory Gating Processes-A Role of Theta Synchronization and the Norepinephrine System.

Performance impairment as an effect of prolonged engagement in a specific task is commonly observed. Although this is a well-known effect in everyday life, little is known about how this affects central cognitive functions such as working memory (WM) processes. In the current study, we ask how time-on-task affects WM gating processes and thus processes regulating WM maintenance and updating. To this end, we combined electroencephalography methods and recordings of the pupil diameter as an indirect of the norepinephrine (NE) system activity. Our results showed that only WM gate opening but not closing processes showed time-on-task effects. On the neurophysiological level, this was associated with modulation of dorsolateral prefrontal theta band synchronization processes, which vanished with time-on-task during WM gate opening. Interestingly, also the modulatory pattern of the NE system, as inferred using pupil diameter data, changed. At the beginning, a strong correlation of pupil diameter data and theta band synchronization processes during WM gate opening is observed. This modulatory effect vanished at the end of the experiment. The results show that time-on-task has very specific effects on WM gate opening and closing processes and suggests an important role of NE system in the time-on-task effect on WM gate opening process.

Working Memory for Spatial Sequences: Developmental and Evolutionary Factors in Encoding Ordinal and Relational Structures.

Sequence learning is a ubiquitous facet of human and animal cognition. Here, using a common sequence reproduction task, we investigated whether and how the ordinal and relational structures linking consecutive elements are acquired by human adults, children, and macaque monkeys. While children and monkeys exhibited significantly lower precision than adults for spatial location and temporal order information, only monkeys appeared to exceedingly focus on the first item. Most importantly, only humans, regardless of age, spontaneously extracted the spatial relations between consecutive items and used a chunking strategy to compress sequences in working memory. Monkeys did not detect such relational structures, even after extensive training. Monkey behavior was captured by a conjunctive coding model, whereas a chunk-based conjunctive model explained more variance in humans. These age- and species-related differences are indicative of developmental and evolutionary mechanisms of sequence encoding and may provide novel insights into the uniquely human cognitive capacities.SIGNIFICANCE STATEMENT Sequence learning, the ability to encode the order of discrete elements and their relationships presented within a sequence, is a ubiquitous facet of cognition among humans and animals. By exploring sequence-processing abilities at different human developmental stages and in nonhuman primates, we found that only humans, regardless of age, spontaneously extracted the spatial relations between consecutive items and used an internal language to compress sequences in working memory. The findings provided insights into understanding the origins of sequence capabilities in humans and how they evolve through development to identify the unique aspects of human cognitive capacity, which includes the comprehension, learning, and production of sequences, and perhaps, above all, language processing.

Event-related synchronization/desynchronization and functional neuroanatomical regions associated with fatigue effects on cognitive flexibility.

Cognitive flexibility is an essential prerequisite for goal-directed behavior, and daily observations already show that it deteriorates when one is engaged in a task for a (too) long time. Yet, the neural mechanisms underlying such fatigability effect in cognitive flexibility are poorly understood. We examined how theta, alpha, and beta frequency event-related synchronization and desynchronization processes during cued memory-based task switching are modulated by time-on-task effects. We put special emphasis on the examination of functional neuroanatomical regions being associated with these modulations, using EEG beamforming. We show clear declines in task switching performance (increased switch costs) with time on task. For processes occurring before rule switching or repetition processes, we show that anticipatory attentional sampling and selection mechanisms associated with fronto-parietal structures are modulated by time-on-task effects but sensory areas (occipital cortex) also show fatigability-dependent modulations. After target stimulus presentation, the allocation of processing resources for response selection as reflected by theta-related activity in parietal cortices is compromised with time on task and similarly a concomitant increase in alpha and beta band-related attentional processing or gating mechanisms in frontal and occipital regions. Yet, considering the behavioral data showing an apparent decline in performance, this probably compensatory increase is still insufficient to allow reasonable performance. The same is likely the case for processes occurring before rule switching or repetition processes. Comparative analyses show that modulations of alpha band activity are as strongly modulated by fatigability as theta band activity. Implications of these findings for theoretical concepts on fatigability are discussed.NEW & NOTEWORTHY We examine the neurophysiological and functional neuroanatomical basis of fatigability in cognitive flexibility. We show that alpha and theta modulations in fronto-parietal and primary sensory areas are central for the understanding of fatigability effects in cognitive flexibility.