To summarize the results of adult obesity neuroimaging studies  published from 2010, with a focus on the treatment of sex as an important biological variable in the analysis, and identify gaps in sex difference research.Neuroimaging studies have shown obesity-related changes in brain structure, function, and connectivity. However, relevant factors such as sex are often not considered.n = 10; 5%) or providing single-sex/disaggregated data ; the remaining studies controlled for sex  or did not consider sex in the analysis . Synthesizing sex-based results, obesity-related parameters  may be generally associated with more robust morphological alterations in men and more robust structural connectivity alterations in women. Additionally, women with obesity generally expressed increased reactivity in affect-related regions, while men with obesity generally expressed increased reactivity in motor-related regions; this was especially true under a fed state. The keyword co-occurrence analysis indicated that sex difference research was especially lacking in intervention studies. Thus, although sex differences in the brain associated with obesity are known to exist, a large proportion of the literature informing the research and treatment strategies of today has not specifically examined sex effects, which is needed to optimize treatment.We conducted a systematic review and keyword co-occurrence analysis. Literature searches identified 6281 articles, of which 199 met inclusion criteria. Among these, only 26 (13%) considered sex as an important variable in the analysis, directly comparing the sexes (The online version contains supplementary material available at 10.1007/s13679-023-00498-0. In the USA, the prevalence of obesity among adults was 42.4% in 2017–2018 . For exa0.00001) .Numerous neuroimaging studies have shown associations between obesity and measures of various brain features, such as morphology, resting-state activity and connectivity, white matter microstructure, and task activation. However, the effects may vary according to sex. Although research has confirmed the involvement of the brain in obesity, a comprehensive multimodal summary of the effects of both obesity and sex on the brain is lacking. Therefore, this review aimed to summarize the results of adult obesity neuroimaging studies published from 2010, with a focus on the treatment of sex in the analysis. Specifically, studies were categorized according to whether the analysis comprised direct comparisons between men and women (direct sex); used data from only male or only female participants or separately analyzed male and female participants, providing disaggregated data and allowing qualitative comparisons (single/within sex); controlled for sex, disallowing comparisons (sex adjusted); or did not consider sex, disallowing comparisons (no sex). We considered the first two categories to adequately treat sex as an important biological variable in the analyses. Additionally, we performed a keyword co-occurrence analysis of the selected articles, as a snapshot of popular topics/concepts in the field of adult obesity neuroimaging their relationship to the topic of sex differences.We searched three online electronic databases, PubMed, Embase, and Web of Science, for obesity neuroimaging studies published from 2010 to the present. This search was conducted using a predetermined set of words related to obesity, neuroimaging, and sex/gender. The search terms, selected in collaboration with a research librarian, are indicated in Supplemental Table Two stages of screening with predetermined inclusion and exclusion criteria were performed. The authors were contacted as needed. All studies considered for eligibility were published in a peer-reviewed journal.In the first stage, we examined the title and abstract of each article for the following criteria: (1) investigation of obesity, (2) adult human participants, and (3) reporting of brain MRI data. Reasons for exclusion were as follows: (1) used non-human models; (2) included infant, child, or adolescent participants; (3) imaged non-brain tissues; (4) no structural magnetic resonance imaging (MRI), resting-state MRI (rsMRI), task/stimulus functional MRI (fMRI), or diffusion tensor imaging (DTI) was performed; (5) no measure of obesity was reported; (6) in treatment studies, no pre-treatment MRI data was reported; (7) investigated patients with anorexia nervosa or other eating disorders; (8) review article/protocol study/abstract only; and (9) duplicate article that was not previously identified.In the second screening stage, we read the entire article, paying particular attention to the methods, results, and discussion sections. Articles that were found to violate the abovementioned study criteria upon full-text review were excluded.After screening, we extracted data on the treatment of sex in the analyses, participant characteristics , modality(s) used, and main findings. Articles were categorized according to how the researchers analyzed sex. The first category was “direct sex,” which was applied to studies that directly compared men and women in the analysis. The second category was “single/within sex,” which was applied to studies that included only male or only female participants or separately reported results for male and female participants, without a direct comparison, but allowing qualitative comparisons. The third category was “sex adjusted,” which was applied to studies that controlled for sex differences in the analysis, disallowing comparisons between the sexes. The fourth category was “no sex,” which was applied to studies that did not consider sex in the analysis , disallowing comparisons between the sexes. The first two categories were considered to adequately treat sex as an important biological variable in the analyses. Subsequently, we organized the findings by the type of brain imaging performed, namely, DTI, structural MRI, rsMRI, and task/stimulus fMRI, with multimodal studies represented in multiple sections.https://vosviewer.com) using LinLog normalization and modularity clustering techniques [A keyword co-occurrence network analysis of the included articles was conducted to examine popular topics/concepts in the field of adult obesity neuroimaging linked and not linked to the topic of sex differences. Keywords comprised those assigned to the article by the authors and/or the database , with preference given to the keywords assigned by the Web of Science database for articles in multiple databases. Keyword co-occurrence networks represent each keyword as a node and each co-occurrence of a pair of words as a link, with the number of times that a pair of words co-occurs in multiple articles as the weight of the link connecting the pair. Keyword co-occurrence networks can be constructed to visualize the cumulative knowledge of a research area, aiding in uncovering meaningful knowledge and insights based on the patterns and strength of links between keywords that appear in the literature . The citchniques . Other sn = 327) were removed. Thus, 6281 articles remained for further screening. After stage 1 screening, 5997 articles were excluded, and 284 articles were selected for full-text screening.The PRISMA diagram illustrating the study selection process is shown in Fig. After second-stage screening, an additional 85 articles were excluded. The reasons for exclusion are indicated in Supplemental Table In total, 12 studies reported on sex differences/commonalities and single/within-sex obesity-related alterations in regional and total gray matter volume (GMV), regional white matter volume (WMV), and cortical thickness. Although most of these studies reported associations between obesity (based on BMI cut-off points), or higher values in measures of body size  or total body fat, and lower GMV in both sexes, the spatial extent of the alterations may be more widespread in men than in women –9, suggeAmong patients with cerebral small vessel disease, reductions in hippocampal volume over a 9-year period were more prominent in men than in women with larger baseline waist circumference . In contA female-specific study found a positive association between high BMI and GMV in the left nucleus accumbens , unlike n = 48) reported on obesity-related alterations in morphological parameters, as well as changes before/after intervention. Similar to the direct-sex and single/within-sex studies, sex-adjusted studies generally support obesity/high-BMI/larger waist circumference as related to lower total GMV [Sex-covariate studies from 2010 to present  largely focused on the impact of bariatric surgery and other interventions on morphological parameters. These studies generally suggest that interventions can at least partially ameliorate obesity-related effects on these parameters [No-sex structural studies from 2010 (rameters –42.n = 4) reported on regional alterations in functional centrality (a graph theory parameter) and the amplitude and frequency distribution of spontaneous fluctuations. A graph theory study reported that increased early life adversity (ELA) was associated with increased centrality in reward and emotion regulation regions in women with high BMI and in somatosensory regions in men with high BMI [Direct-sex and single/within-sex rsMRI studies  have mostly reported on alterations in intrinsic connectivity between regions or within a network, not centrality or frequency distribution changes, rendering it difficult to compare the results to those in direct-sex and single/within-sex studies. However, several studies reported obesity-related increases in intrinsic connectivity or involving regions with increased centrality or altered frequency distribution in direct-sex and single/within-sex studies, including the ventral striatum, amygdala, and insula [Sex-adjusted rsMRI studies (d insula –49.n = 13) largely reported on the impact of interventions on rsMRI parameters [No-sex resting-state studies  investigated obesity-related responses to high and low energy–dense foods, under fed and fasting conditions. In a fed state, men with obesity showed greater connectivity between the amygdala and subgenual ACC and between the ventral striatum and bilateral supplementary and primary motor areas, left postcentral gyrus, and left precuneus, and greater activation in bilateral supplementary motor areas (motor control), while women with obesity showed greater connectivity between the amygdala and the left angular gyrus and primary motor areas, and greater activation in the dorsal ACC, in response to high energy–dense food cues [Direct-sex and single/within-sex task-fMRI studies  investigated responses to a wider range of stimuli (food and odor cues) and tasks  than direct-sex and single/within-sex studies. Studies using visual food-cue stimuli found greater obesity-related reactivity in the amygdala, insula, hippocampus, striatum, hypothalamus, ventral posterior cingulate, angular gyrus, putamen, and frontal cortex, including the dorsolateral PFC (in a fasting state) [Sex-adjusted task-fMRI studies (g state) –67; someg state) –58.n = 28) largely reported on the impact of interventions on reactivity to food-related cues [No-sex task-fMRI studies  reported on common and sex-specific associations between BMI/body fat and DTI measures of fractional anisotropy (FA), mean diffusivity (MD), and axial diffusivity (AD), as well as structural centrality. Specifically, high BMI/total body fat was associated with decreased AD in the corpus callosum [Direct-sex and single/within-sex DTI studies  mainly reported associations between obesity/high BMI/adiposity and FA. One study reported high BMI as related to a reduction in the global FA [Sex-adjusted studies  reported on the impact of exercise intervention on diffusivity parameters [No-sex DTI studies  published in early (2010–2015) and later (after 2015) portions of the study period. Ten (12.8%) of 78 studies published in the early study period and 16 (13.2%) of 121 studies published in the later study period considered sex as an important variable. Accordingly, the majority of studies that considered sex as an important variable were published in the late study period .In the keyword co-occurrence network analysis of neuroimaging articles on adult obesity from 2010, three main clusters were identified: a reward/reactivity cluster, including keywords related to the extended reward system  and reward-related functions , and brain reactivity to food-related cues and food intake; a disease-state cluster ; and a weight-loss intervention cluster Fig. a. AlthouIn this review, we categorized adult obesity neuroimaging studies according to the treatment of sex in the analyses and found that only ~ 13% of the studies treated sex as an important variable. There was a general lack of studies that examined sex differences, or at least presented disaggregated data, which could be useful in developing sex-specific biomarkers and interventions. Only 10 studies (5%) directly compared the sexes, and 16 (8%) limited the analysis to a single sex or analyzed the sexes separately, allowing qualitative comparisons, which may be a first step in understanding the impact of sex. This is in stark contrast to the 120 studies (60%) that controlled for sex and 53 studies (27%) that did not consider sex in the analysis, disallowing any comparisons. Moreover, the few single-sex studies were disproportionately focused on women rather than on men. This reveals the need for more research on sex differences and men in the field of obesity neuroimaging. In order to better understand and treat obesity, differences in obesity effects on the brain between the two sexes need to be elucidated, especially as the present review indicates that such differences do exist.The literature to date indicates that obesity is associated with numerous alterations in brain structure and responses to food-related stimuli; however, the extent and predominate regions involved differ between men and women with obesity, raising concerns regarding conclusions drawn from sex-adjusted and no-sex studies, as ignoring sex or adjusting for sex can mask important differences. Direct-sex studies generally showed more robust morphological alterations in subcortical and emotion-related regions in men –13 and mInvestigations of sex effects were especially lacking in intervention studies. This was clearly apparent in the keyword co-occurrence, as the keyword, sex differences, did not show any links with intervention-related keywords, such as bariatric surgery and weight loss. The keyword co-occurrence analysis revealed other major areas of research with limited consideration of sex differences, which included disease states such as dementia, Alzheimer’s disease, and insulin resistance. Many of the no-sex studies focused on the impact of various interventions on brain measures in the general obese population and in specific patient populations. Because sex was ignored, its relative contribution to differences in the neural response to treatment is unclear. However, non-imaging studies suggest sex differences in obesity intervention effectiveness , and thep-values [Some of the studies reporting “sex-based” or “sex-specific” effects did not directly compare the sexes; rather, the researchers reported obesity-related effects in men and women considered separately. This approach (testing within each sex separately) can inflate the probability of concluding that a sex-specific effect is present compared to that when directly comparing the sexes . In the p-values •, 97••, The earliest articles covered in this review were published nearly a decade after the Institute of Medicine reported on the importance of considering sex in the exploration of the biological contributions to human health in 2001 . DespiteSupplementary Materials: The full bibliography of reviewed articles according to the treatment of sex in the analysis.Supplementary file1 (DOCX 34 KB)Below is the link to the electronic supplementary material.


Three-dimensional (3D) printing is a novel innovation in the field of craniomaxillofacial surgery, however, a lack of evidence exists related to the comparison of the trueness of skull models fabricated using different technology-based printers belonging to different cost segments.A study was performed to investigate the trueness of cone-beam computed tomography-derived skull models fabricated using different technology based on low-, medium-, and high-cost 3D printers. Following the segmentation of a patient’s skull, the model was printed by: (i) a low-cost fused filament fabrication printer; (ii) a medium-cost stereolithography printer; and (iii) a high-cost material jetting printer. The fabricated models were later scanned by industrial computed tomography and superimposed onto the original reference virtual model by applying surface-based registration. A part comparison color-coded analysis was conducted for assessing the difference between the reference and scanned models. A one-way analysis of variance (ANOVA) with Bonferroni correction was applied for statistical analysis.The model printed with the low-cost fused filament fabrication printer showed the highest mean absolute error (Both stereolithography and material jetting based printers, belonging to the medium- and high-cost market segment, were able to replicate the skeletal anatomy with optimal trueness, which might be suitable for patient-specific treatment planning tasks in craniomaxillofacial surgery. In contrast, the low-cost fused filament fabrication printer could serve as a cost-effective alternative for anatomical education, and/or patient communication. Recent advancements in additive manufacturing (AM), also known as three-dimensional 3D) printing and rapid prototyping (RP), have led to an ever-increasing impact on the field of craniomaxillofacial surgery . The manD printinCurrently, a wide variety of 3D printers exist in the market for printing maxillofacial skeletal models. These printers can be further classified as low-cost desktop/consumer grade and high-cost professional 3D printers . Fused fprinting .For preoperative planning and clinical training, craniomaxillofacial 3D models are typically manufactured via in-house or by commercial external printing service centers utilizing high-cost professional-grade 3D printers. The expertise of operators, cost and delivery time of the models might influence the patient’s treatment process, consequently, limiting their general applicability , 13. In This research was conducted in compliance with the World Medical Association Declaration of Helsinki on medical research. The study was approved by the Ethical Review Board of the University Hospitals Leuven, Leuven, Belgium (reference number: S64493) for retrospectively collecting and using patient imaging data. Informed consent was obtained from all participants.A 32-year-old female patient’s CBCT image consisting of a normal complete skull (craniomaxillary complex and mandible) without any pathological condition or artefacts was retrospectively obtained from the Dentomaxillofacial Radiology Center . The scanning was performed using NewTom VGI evo  at The DICOM images were imported into Mimics 22.0 , where thresholding-based semi-automatic segmentation of the skeletal structures was performed Fig. d. The spThree different printing technologies and printers were utilized for the fabrication of the model: a low-cost Prusa i3 MK3S printer  Fig. f, a mediFigure The fabricated models were scanned with an industrial CT  at Mean error, mean absolute error (MAE), root mean square (RMS) and volumetric values were calculated in 3-Matic 14.0 , where the mean error refers to positive or negative deviation error, while MAE is the overall magnitude of error as shown in Eq.  as shown in the Eq. . It allop value of Data were analyzed using IBM SPSS Statistics for Windows, version 21.0 .The Shapiro-Wilk test was used to investigate assumptions of normality. A one-way analysis of variance (ANOVA) with Bonferroni correction was applied for multiple comparisons between different printers and The printing process of all printers went smoothly without any issues. However, qualitative observation of the models following post-processing stage revealed that the low-cost FFF model exhibited noticeable rough patches at the right temporal region and left skull base. No other flaws were detected with the other models. Moreover, FFF model costed a fraction compared to other models, nevertheless, it required approximately 5 times longer to print.Taking into account the RMS values, the high-cost MJ, medium-cost SLA and low-cost FFF models observed an average discrepancy of entclass1pt{minimaBased on the mean error, the FFF model had the highest overall surface error of According to the RVD of printed anatomical structures Table , the higIn the era of personalized precision medicine, patient-specific 3D printed skeletal models derived from medical imaging datasets have become a standard tool . An anatDue to the multi-dimensionality and complex nature of a human skull, its accurate anatomical representation is vital in all areas of craniomaxillofacial surgery . The truA previous study showed that a dimensional linear error within the range of 2% variability could be considered as clinically acceptable for the production of maxillofacial skeletal models . SimilarThe low-cost FFF printer utilized in the present study showed a higher amount of discrepancy compared to other studies, whereas both consumer-grade and professional printers showed comparable trueness for printing skeletal models , 16, 25.trueness . The infmentclasspt{minimaa et al. . Even wiThe trueness of the SLA model was in accordance with other studies, where SLA was found to be optimal for fabricating skeletal structures with intricate details, given its superior resolution, ultimately related to the laser positional accuracy . LikewisThe main strength of the study was the first-time fabrication and trueness assessment of the craniomaxillary complex using a low-cost printer which has not been previously investigated. The findings highlight the potential of a low-cost printer for fabricating anatomical models which could be useful for treatment planning, surgical simulation, and anatomical education. However, it is crucial to conduct further testing and optimization before establishing their application in a routine clinical setting. The study also had certain limitations. Firstly, the study was limited to a small sample of three printers with different technologies and materials which cannot be generalized to all the printers. Secondly, the selection of material could have influenced the trueness. Hence, it is important to investigate the impact of different materials and additive components on material conversion and properties . ThirdlyBoth stereolithography and multi-jetting were able to replicate the skeletal anatomy on a medium- and high-cost printer, respectively, with the least amount of error, thereby confirming their applicability for clinical application, such as pre-bending plates and fabricating implants. Desktop/consumer grade FFF printer offered the highest discrepancy which might not be optimal for clinical applications, however, it could serve as a cost-effective alternative for surgical simulation, anatomical education, and/or patient communication.


Language lateralization in patients with focal epilepsy frequently diverges from the left-lateralized pattern that prevails in healthy right-handed people, but the mechanistic explanations are still a matter of debate. Here, we debate the complex interaction between focal epilepsy, language lateralization, and functional neuroimaging techniques by introducing the case of a right-handed patient with unaware focal seizures preceded by aphasia, in whom video-EEG and PET examination suggested the presence of focal cortical dysplasia in the right superior temporal gyrus, despite a normal structural MRI. The functional MRI for language was inconclusive, and the neuropsychological evaluation showed mild deficits in language functions. A bilateral stereo-EEG was proposed confirming the right superior temporal gyrus origin of seizures, revealing how ictal aphasia emerged only once seizures propagated to the left superior temporal gyrus and confirming, by cortical mapping, the left lateralization of the posterior language region. Stereo-EEG-guided radiofrequency thermocoagulations of the (right) focal cortical dysplasia not only reduced seizure frequency but led to the normalization of the neuropsychological assessment and the “restoring” of a classical left-lateralized functional MRI pattern of language. This representative case demonstrates that epileptiform activity in the superior temporal gyrus can interfere with the functioning of the contralateral homologous cortex and its associated network. In the case of presurgical evaluation in patients with epilepsy, this interference effect must be carefully taken into consideration. The multimodal language lateralization assessment reported for this patient further suggests the sensitivity of different explorations to this interference effect. Finally, the neuropsychological and functional MRI changes after thermocoagulations provide unique cues on the network pathophysiology of focal cortical dysplasia and the role of diverse techniques in indexing language lateralization in complex scenarios. Indeed, right hemispheric dominance or bilateral patterns of either receptive, expressive, or both language aspects are reported in up to one-third of patients with focal epilepsy , a prevaLanguage assessment in epilepsy has evolved through a bunch of recording techniques, from the most traditional intracarotid amobarbital (Wada) test for language lateralization . For lanNeuroimaging techniques emerged in the last decades providing non-invasive methods  for langMagnetic and electrophysiological recordings also contributed tools for the estimation of language lateralization , especiaIn the case of epilepsy, understanding the network between language organization and epileptic interictal/ictal discharges in the EZ would help to refine the estimation of language dominance, with fundamental implications for pre-surgical procedures and cognitive prognosis.One missing piece of information is whether the effects of epilepsy on language organization are, to some extent, reversible once the epileptic activity is mitigated or removed. An ideal framework to tackle this issue is represented by stereo-EEG-guided radiofrequency thermocoagulations (RFTC), i.e., controlled focal lesions applied at the end of stereo-EEG monitoring to reduce the number and intensity of seizures . This prCombining the potential of RFTC with the issue of language lateralization and organization in epileptic patients, two major questions remain unanswered. The first concerns whether RFTC can improve the functioning of language networks previously conditioned by epileptic activity arising from connected territories outside the language network. In parallel, an intriguing question is whether RFTC may also alter the estimation of language lateralization via neuroimaging techniques.We address these issues by introducing the case of a patient with seizure-associated language impairment, thus suggesting epilepsy arising from the language-dominant hemisphere but presenting with a challenging definition of language lateralization. According to a neurological, neuroimaging, and neuropsychological assessment. Of note, functional MRI for language failed to indicate any clear lateralization. The subsequent stereo-EEG allowed identifying a right temporal EZ, propagating ictal activity to a left-dominant language network determining the language ictal disturbances. The EZ was thermocoagulated, and the patient showed improved language functions documented by neuropsychological tests. Notably, the functional MRI investigation repeated after the thermocoagulation revealed a clear left lateralization of language, indicating how epileptic activity can, in part reversibly, reduce the reliability of language lateralization by metabolic imaging.2.A right-handed 31-year-old woman, a finance worker with drug-resistant focal epilepsy, was evaluated for epilepsy surgery in May 2017. Neurological examination was normal. From the age of five, she presented seizures that started with an acoustic illusion (“voices and noises overlapping in both ears”) accompanied by fear and warmth sensation, followed by an impairment of language (both comprehension and expression) and, subsequently, loss of awareness, and oroalimentary automatisms. After the seizure, language disturbance persisted for at least 10 min. She continued having weekly seizures.Prolonged video-EEG showed interictally bilateral asynchronous anterior and mid-temporal spikes. One seizure with usual semiology was recorded, suggesting an ictal onset in the right temporal region with early contralateral spreading. Brain MRI (1.5 Tesla) resulted normal, while a Positron Emission Tomography and Computed Tomography with 18F-fluorodeoxyglucose (18FDG-PET/CT) showed a right temporal hypometabolism, more marked at the level of the temporal pole and superior temporal gyrus .2.1.The neuropsychological evaluation  showed aA functional MRI was performed in an Achieva® 1.5 T magnet . The acquisition consisted of a T2*-weighted gradient echo planar imaging sequence sensitive to blood oxygen level-dependent (BOLD) contrast. Three language paradigms- comprehension, association, and fluency - were acoustically administered . The patThe functional MRI did not display any BOLD hemispheric activation during auditory comprehension and association tasks and only a slight activation of the right middle and inferior frontal gyrus during the fluency task. A right cerebellar activation was observed during all language tasks .Then, Diffusion Tensor Imaging (DTI) based fiber tracking was performed . A probaThe study showed the bilateral presence of a white matter tract connecting the inferior frontal gyrus and the superior temporal gyrus , with asymmetry and a prevalence of the left-sided one . These l2.2.The inconclusive findings regarding the limits of a firmly suspected right temporal EZ made the patient eligible for an invasive study with intracerebral electrodes (stereo-EEG) , 2017. TThe stereo-EEG interictal recording showed an altered background and continuous epileptiform discharges suggestive of type II focal cortical dysplasia (FCD)  in the p2.3.After recording spontaneous seizures on stereo-EEG, we proceeded to perform an ESM and analyze the gamma activity induced during language tasks.On the left side, 50 Hz stimulations  in the posterior part of the superior temporal gyrus interrupted fluent speech, associated with an auditory illusion. In the same site, stimulations at 9 Hz   induced z-score) to the interval  before the stimulus presentation. Significance among homologous contacts located in the left and right inferior frontal gyrus, and separately among left and right superior temporal gyrus, were tested with a two-tailed paired t-test (p < 0.001)   induced < 0.001) .The gamma activation was significantly left lateralized in both the listening and the read-and-repeat condition, both in the superior temporal and inferior frontal gyrus. A remarkable activation during reading was observed in the left inferior frontal gyrus, congruent with the clinical slowing in reading observed during ESM .Cortico-cortical evoked potentials were visually analyzed using a low-frequency stimulation protocol  applied through all electrode contacts . They do2.4.The patient underwent stereo-EEG-guided RFTC applied through the electrode contacts exploring the posterior right superior temporal gyrus and the ipsilateral temporal pole. She remained seizure-free for 2 months. Then, seizures restarted with a lower frequency than before the procedure.Three months after RFTC, a new neuropsychological evaluation revealed improved naming and verbal memory  and stable phonemic fluency .The functional MRI for language was repeated with the same protocol, revealing a left-hemispheric activation in all three tasks. On the contrary, right cerebellar activation remained similar to that unveiled during the preoperative exam .The EEG was performed 6 months after surgery and only slow activity in the right temporal derivations was present, in total absence of epileptiform abnormalities.The patient was operated with a tailored resection within the limits of the right temporal lobe, including the superior gyrus, pole, and mesial structures . She didOne year after the resection, the neuropsychological evaluation showed increased phonemic fluency and confirmed the previously observed improvements (after RFTC) in verbal memory and naming .3.In the present study, we introduced the case of a patient whose epileptiform activity impacted language functions and the capacity of non-invasive explorations to identify her language dominance. The longitudinal and multimodal set of investigations conducted on the patient and the positive clinical outcome following stereo-EEG enabled us to document a complex interaction between epilepsy and the function of the language networks.A right temporal origin of the seizures was firmly supported by video-EEG and 18FDG-PET/CT findings. However, this posed an issue with hemispheric language lateralization. Indeed, ictal aphasia could be explained by the propagation of seizures to a left-dominant hemisphere or by an “atypical” pattern of right hemispheric language lateralization. The neuropsychological evaluation supported the second hypothesis, as language and verbal memory impairments are more frequent in patients whose epilepsy originates in the dominant hemisphere . The funDifferent clinical conditions may lead to false-negative results during the functional MRI assessment of language lateralization. For instance, in tumor surgery, the weakness of the BOLD signal in regions adjacent to the tumor may lead to false negatives  due to ide facto generating a false negative finding about cortical language lateralization.In physiological conditions, neural metabolism and blood flow are tightly linked (“neurovascular coupling”). The increase in neural activity and metabolic demand when performing a task thus implies an increase in regional blood flow, oxygen supply, and subsequent BOLD signal in functional MRI . On the An exceptional opportunity for causally testing this hypothesis came from repeating the functional MRI examinations and neuropsychological tests after the thermocoagulation of the FCD. Indeed, not only did the fMRI activation during language tasks recover the left-lateralized pattern coherent with the language dominance of the patient, but she also showed an improvement in the neuropsychological assessment. Starting from the previous observation that RFTC interferes with adjacent but also distant areas connected with the ablated cortex , we concThe case described here provides valuable insights for the non-invasive study of language lateralization in daily clinical practice. We propose that language lateralization in epileptic patients cannot be investigated without accounting for the topography of the EZ and the propagation of epileptic activity, which could alter or mask the language-related contrast. Beyond using EEG-fMRI approaches that model the interictal epileptiform discharges into the functional MRI signal analysis , two parThe first derives from the (right) cerebellar functional MRI activation stability across all the language tasks in pre- and post-thermocoagulation recordings. The right cerebellar hemisphere plays a role in different language processing components through crossed cerebro-cerebellar connections . Likewis. In the present case, this asymmetry was more informative than the initial functional MRI lateralization, suggesting that the alterations driven by epileptic activity are larger at the functional level  than the structural one (diffusion tractography). Although previous studies have proposed a coincidence between grades of lateralization of BOLD activations in functional MRI and features of the arcuate bundle in DTI, the coincidence in patients whose language in functional MRI is not left-lateralized, and the underlying mechanistic explanation, is still a matter of debate  may significantly affect cortical activation on functional MRI, the etiology of epilepsy must be considered during the investigation of language lateralization. In case of inconclusive functional MRI cortical contrasts, we suggest incorporating in clinical practice the quantification of cerebellar activations and arcuate fasciculus asymmetry by DTI.The original contributions presented in the study are included in the article/The studies involving humans were approved by A.S.S.T. Grande Ospedale Metropolitano Niguarda. The studies were conducted in accordance with the local legislation and institutional requirements. The participants provided their written informed consent to participate in this study. Written informed consent was obtained from the individual(s) for the publication of any potentially identifiable images or data included in this article. Written informed consent was obtained from the participant/patient(s) for the publication of this case report.BA: Conceptualization, Writing – original draft, Writing – review & editing. VM: Conceptualization, Writing – original draft, Writing – review & editing. MR: Formal analysis, Software, Writing – original draft, Investigation. LB: Formal analysis, Software, Writing – original draft, Investigation. PS: Methodology, Writing – original draft, Investigation. FZ: Data curation, Formal analysis, Writing – original draft, Investigation. SS: Formal analysis, Writing – original draft, Investigation. PB: Resources, Writing – original draft, Investigation. Pd’O: Formal analysis, Software, Writing – original draft, Investigation. FrC: Software, Writing – original draft, Investigation. MD: Software, Writing – original draft, Formal analysis, Investigation. FaC: Writing – review & editing, Investigation. PA: Funding acquisition, Methodology, Supervision, Writing – original draft, Writing – review & editing. IS: Supervision, Writing – original draft, Writing – review & editing, Methodology.


There have been substantial changes in the simulation technology landscape, in particular virtual reality (VR), during the past decade, which have resulted in increased abundance and decreased cost. We therefore updated a previous meta-analysis conducted in 2011, aiming to quantify the impact of digital technology-enhanced simulation (T-ES) compared with traditional teaching in physicians, physicians-in-training, nurses, and nursing students.We conducted a meta-analysis consisting of randomized controlled trials published in English between January 2011 and December 2021 in peer-reviewed journals indexed in seven databases. Moderators for study duration, instruction, type of healthcare worker, type of simulation, outcome measure, and study quality rated by Medical Education Research Study Quality Instrument (MERSQI) score were included in our model and used to calculate estimated marginal means (EMMs).The overall effect of T-ES was positive across the 59 studies included in the analysis compared with traditional teaching [overall effect size 0.80 ]. This indicates that T-ES is effective in improving outcomes across a wide variety of settings and participants. The impact of T-ES was found to be greatest for expert-rated product metrics such as procedural success, and process metrics such as efficiency, compared with knowledge and procedure time metrics.The impacts of T-ES training on the outcome measures included in our study were greatest in nurses, nursing students and resident physicians. T-ES was strongest in studies featuring physical high-fidelity mannequins or centers, compared with VR sensory environment T-ES, though there was considerable uncertainty in all statistical analyses. Further high-quality studies are required to assess direct effects of simulation training on patient and public health outcomes. Technology-enhanced simulation (T-ES) allows learners to develop their knowledge and skills without exposing real patients to potential harm. T-ES enables clinicians to train for low-frequency, high-intensity events where other forms of medical education may not provide a sufficiently realistic experience. T-ES also allows healthcare professionals to gain insight into clinical practice from multiple perspectives, including that of the patient. For example, a nationwide study in the United States concluded that high-quality simulation experiences could substitute up to 50% of traditional clinical hours across the prelicensure nursing curriculum . This isThere are numerous potential benefits to the adoption of T-ES: simulation could contribute to increasing both quality and capacity of healthcare training, provided it is sufficiently realistic with opportunities to apply learning, both of which are key themes when frontline healthcare professionals are surveyed . Aside fquasi-experimental studies or randomized studies with no intervention control groups.The main finding of the Cook et al.  study waToday, the technology landscape is markedly different than what existed when Cook et al.  was condThe Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA)  guidanceWe based our search strategy on Cook et al. , and madn results = 14,876), Embase , Scopus , Web of Science , PsycINFO , CINAHL , and ERIC (n = 681), which was conducted in April and May 2022 , of which 52.6% was due to between-study heterogeneity and 32% was from heterogeneity between the individual effect sizes. A moderator test for publication bias and time-lag bias was conducted, which was not statistically significant . Moderator analysis also confirmed that there was no statistically significant impact of study quality (MERSQI score) on the overall effect size , which means that lower-quality studies did not contribute to an unduly strong effect size.Total heterogeneity was considerable ] and residents , and smallest in medical students  and physicians in practice . High-fidelity mannequins or physical environments  were then compared to studies featuring simulated VR environments . VP case-based T-ES demonstrated the greatest EMMs of any modality , though this result should be interpreted with caution due to a small sample size and below-average MERSQI scores (range 9.5–11.5). Lastly, average scores were greater when T-ES training was carried out over more than 1 day [0.94 ], compared with single-day exercises [0.58 ], which may suggest a dose-response relationship where a greater duration of T-ES training leads to greater scores in our included outcome measures.The impact of T-ES across our included outcome measures was found to be greatest in nurses and nursing students  shows a strong immediate effect of T-ES training when translated to knowledge scores, clinical settings involving real patients, and dexterous surgical tasks involving cadaver or porcine models. These results support the consensus that high-fidelity simulation is best suited to refining performance as opposed to knowledge, with the largest effects shown for process and product outcomes as opposed to time or knowledge. Broadly, this suggests that T-ES can contribute to improving patient care and prepare healthcare trainees for unfamiliar situations.In comparison with the Cook et al.  study, wEvidence has suggested that low-fidelity simulation is most impactful in building knowledge, whilst higher-fidelity simulation is best used to develop performance and action . It is iDue to limited randomized studies with suitable control groups, we lacked the statistical power to make definitive comparisons between types of simulation or healthcare professionals using EMMs. This was apparent when examining a difference in outcomes of T-ES between physicians at each stage of training, or between physicians and nurses, as almost three-quarters of included studies evaluated physicians-in-training, it was difficult to draw statistically powerful conclusions, something that is also highlighted by a relative lack of reviews evaluating simulation in practicing physicians.We used a broad search strategy to synthesize 59 generally high-quality randomized studies to contribute to the evidence base for simulation in diverse healthcare settings.We found that skills and knowledge developed during T-ES are generally transferable to other settings across physicians and nurses at all stages of training, types of high-fidelity simulation modality and numerous clinical specialties. However, this must be considered with considerable uncertainty and heterogeneity taken into account. Differing inclusion criteria, study designs and statistical methodology mean that fair and direct comparisons with Cook et al.  are diffWith a tendency to focus on novel, rapidly advancing technologies such as VR, existing simulation modalities such as low-fidelity bench-top models or simulated patients may perhaps not receive equal consideration. Lower-fidelity methods can still achieve the desired learning outcomes, especially in more inexperienced trainees. There remains a clear need to identify whether simulation is effective in improving the quality and scalability of medical education, and care delivered to patients, relative to lower-fidelity, less expensive interventions.The original contributions presented in the study are included in the article/AM devised the study concept, conducted the literature searches, and drafted the first version of the manuscript. AM and EI-C conducted the meta-analysis and redrafted the manuscript. Both authors contributed to the article and approved the submitted version.


Locally advanced pancreatic ductal adenocarcinoma (PDAC), accounting for about 30% of PDAC patients, is difficult to cure by radical resection or systemic chemotherapy alone. A multidisciplinary strategy is required and our TT-LAP trial aims to evaluate whether triple-modal treatment with proton beam therapy (PBT), hyperthermia, and gemcitabine plus nab-paclitaxel is a safe and synergistically effective treatment for patients with locally advanced PDAC.This trial is an interventional, open-label, non-randomized, single-center, single-arm phase I/II clinical trial organized and sponsored by the University of Tsukuba. Eligible patients who are diagnosed with locally advanced pancreatic cancer, including both borderline resectable (BR) and unresectable locally advanced (UR-LA) patients, and selected according to the inclusion and exclusion criteria will receive triple-modal treatment consisting of chemotherapy, hyperthermia, and proton beam radiation. Treatment induction will include 2 cycles of chemotherapy (gemcitabine plus nab-paclitaxel), proton beam therapy, and 6 total sessions of hyperthermia therapy. The initial 5 patients will move to phase II after adverse events are verified by a monitoring committee and safety is ensured. The primary endpoint is 2-year survival rate while secondary endpoints include adverse event rate, treatment completion rate, response rate, progression-free survival, overall survival, resection rate, pathologic response rate, and R0 (no pathologic cancer remnants) rate. The target sample size is set at 30 cases.The TT-LAP trial is the first to evaluate the safety and effectiveness (phases1/2) of triple-modal treatment comprised of proton beam therapy, hyperthermia, and gemcitabine/nab-paclitaxel for locally advanced pancreatic cancer.This protocol was approved by the Tsukuba University Clinical Research Review Board (reference number TCRB22-007). Results will be analyzed after study recruitment and follow-up are completed. Results will be presented at international meetings of interest in pancreatic cancer plus gastrointestinal, hepatobiliary, and pancreatic surgeries and published in peer-reviewed journals.https://jrct.niph.go.jp/en-latest-detail/jRCTs031220160.Japan Registry of Clinical Trials, jRCTs031220160. Registered 24 th June 2022,  Becauser types) . AdditioWhile diverse clinical trials have reported treatment strategies for LAPC, a gold standard regimen remains to be established as multidisciplinary chemo/radiotherapy combinations have failed to improve 5-year OS rates. This was seen in previously reported chemoradiation therapy studies where overall survival did not change but progression-free survival did improve . Thus, mWe recently published a retrospective study about the feasibility and effectiveness of a triple-modal strategy that combines gemcitabine-based chemotherapy, proton beam therapy (PBT), and local hyperthermia for UR-LA patients with major-artery invasion . In thisThis trial is an interventional, open-label, non-randomized, single-center, single-arm phase I and II clinical trial organized and sponsored by the University of Tsukuba. This single-center study will be conducted at the University of Tsukuba Hospital. Eligible patients who are diagnosed with locally advanced pancreatic cancer and selected according to the inclusion and exclusion criteria listed below will receive triple-modal treatment, including PBT, hyperthermia, and chemotherapy.The aim of this trial is to assess the safety of the triple-modal treatment comprised of PBT, hyperthermia, and gemcitabine with nab-paclitaxel as the phase I part and the efficacy and safety of this strategy as the phase II part.The primary outcome for phase I is drug-limiting toxicity (DLT) and the 2-year survival rate is the phase II primary outcome. The primary and the secondary outcomes for each phase are summarized in Table This trial includes patients who are diagnosed with locally advanced pancreatic cancer, including both borderline resectable with arterial invasion (BR-A) and unresectable locally advanced (UR-LA), according to National Comprehensive Cancer Network (NCCN) guidelines Version 1.2021. The diagnosis of primary tumors will be performed by thin-slice (slice thickness of 2.5 mm or less) dynamic CT and board-certified radiologists, surgeons, physicians and radiation oncologists in clinical concert at the Tsukuba Pancreato-Biliary Tumor Center. Patients with distant organ metastases, including para-aortic lymph node metastasis and/or peritoneal dissemination, will be excluded from this trial. The evaluation of distant metastases will be conducted by lung CT scans and/or liver MRI with gadoxetate disodium as contrast agent , with FDG-PET (18F-fluorodeoxyglycose positron emission tomography) considered if the diagnosis of distant metastases is difficult. Eligible patients will be screened according to the following criteria:A)Histological or cytological diagnosis of pancreatic ductal adenocarcinoma (PDAC).B)Radiologically diagnosed as BR (borderline resectable) or UR-LA  according to NCCN guidelines.C)Performance Status (PS): 0-1 (ECOG criteria).D)3, platelet ≥ 75,000/mm3, total bilirubin ≤ 3.0mg/dl, AST ≤ ULN x 5, ALT ≤ ULN x 5, and creatinine clearance > 40 mL/min (according to the Cockroft-Gult formula)Patients with normal major organ function  reflected in the following lab values: hemoglobin ≥ 9.0g/dl, neutrocyte ≥ 1,500/mmE)Between ages 20 and 80.F)Patients who can receive proton beam therapy.G)Patients who give written, informed consent.Inclusion criteria:A)Patients with distant metastases, including para-aortic lymph node metastasis and/or peritoneal dissemination.B)Patients with severe drug-induced hypersensitivity syndrome.C)Patients with other, concurrent invasive cancers with less than 5 years of expected disease-free survival.D)Patients with allergies for contrast materials and difficult-to-diagnose pancreatic ductal adenocarcinomas.E)Past treatment history for pancreatic cancer and resistance for any treatmentsF)Patients who received dose overlap with previous irradiation or for whom this cannot be ruled out.G)Patients with comorbidities that carry contraindications to radiation therapy .H)Patients with clear endoscopic exposure of cancer in the mucosa of the gastrointestinal tract.I)Patients with HCV RNA virus-positive or HBs antigen-positive hepatitis statuses.J)Patients on dialysis with chronic renal failure or Child-Pugh B or higher-class liver cirrhosis.K)Patients who are, may be, or wish to become pregnant plus those who are breastfeeding.L)Patients with severe mental disabilities.Exclusion criteria:This is a phase I/II study to evaluate the safety and efficacy of gemcitabine/nab-paclitaxel therapy in combination with PBT and local hyperthermia in patients with locally advanced pancreatic cancer Fig. . Two rouThe TT-LAP trial includes safety evaluations in 5 initial patients as phase I Fig. . After tDose-limiting toxicity (DLT) is defined as an adverse event (AE) that is at least potentially related to the protocol treatment and meets the criteria. Each criterion is listed in Table For the purpose of evaluating the safety of the study, the first 5 patients were assigned to Phase 1, which was a safety part of the study. The total number of patients in the study was set at 30 for feasibility and for the following reasons. The results of a single-center, retrospective, observational study on multimodality therapy using proton beams and hyperthermia with GEM alone  showed aThis number of cases meets the threshold response rate of 30% for 2-year survival when conventional chemoradiotherapy is administered, an expected response rate of 55%, and power set to 80% when the two-sided significance level is set at 0.05. This assumes a higher expected response rate to intensified chemotherapy and local hyperthermia than the 50% response rate for PBT and chemotherapy.Efficacy will be evaluated by blood samples to measure tumor markers and contrast-enhanced CT scans of the thorax and abdomen, which will be scheduled every 3 months, to evaluate the presence of primary tumors and neoplastic lesions. For efficacy, overall survival will be analyzed by the Kaplan-Meier method to obtain a 2-year survival rate. The analysis of the primary efficacy endpoint will be based on the total analysis of the population (FAS).This study is funded by the Grant for Implementation of Advanced Medicine (GIAM) of the University of Tsukuba Hospital.This study was approved by the Tsukuba University Clinical Research Review Board based on the Clinical Trials Act of Japan and the Declaration of Helsinki or its equivalents. This trial is registered in the Japan Registry of Clinical Trials (jRCTs031220160). The results from this study will be analyzed and published in peer-reviewed journals.Treatment of LAPC is extremely confounding since heterogenous disease progression creates unique treatment response profiles and high case-to-case variability. Compounding this difficulty is exact quantification of tumor and metastatic parameters only by currently available imaging studies. In addition, the high biological malignancy of pancreatic cancer cells and their resistance to chemotherapy and radiotherapy are also contributing factors. Thus, effective and recommended treatment strategies remain undefined. LAPC, in these cases, must be approached from the standpoint of not just solid tumors but also malignancies in active transition to micro-metastasis. Because of their multispectral targeting ability, intensive and synergistic chemotherapies (such as FOLFIRINOX) should therefore be considered as a mainstay for systemic treatment of LAPC since the previously reported median OS of 24.2 months is superior to gemcitabine for LAPC . HoweverHence, the aim of this clinical trial (TT-LAP) is to evaluate the safety and effectiveness of our triple-modal treatment, comprised of gemcitabine with nab-paclitaxel (standard for advanced PDAC), PBT, and local hyperthermia.The TT-LAP trial includes an initial phase 1 safety assessment due to a lack of evidence when combining PBT with local hyperthermia and gemcitabine with nab-paclitaxel infusion for pancreatic cancer. In our previous report on gemcitabine-based, triple-modal treatment for LPAC , a gastrSeveral studies have reported the feasibility and effectiveness of PBT as the ideal radiation therapy for pancreatic cancer , 13–15. Hyperthermia has been widely applied to breast cancer and other malignant tumors of the body surface as well as to the multidisciplinary treatment of pancreatic cancer. A review by Horst et al. showed that hyperthermia can enhance the therapeutic effect of combinations of chemoradiotherapy or radiotherapy for pancreatic cancer . In addiClinical trials have been conducted for unresectable/regionally advanced pancreatic cancer using PBT in combination with chemotherapy to determine radiation dose . InvestiAlthough the number of facilities that can provide PBT and hyperthermia is limited, several trials have reported that PBT for locally advanced pancreatic cancer is well tolerated and has a high local control effect. A multimodality approach combining PBT, hyperthermia, and modern chemotherapy  has the potential to achieve even stronger local control and control of micro-metastases.The TT-LAP trial is funded by the Advanced Medical Care Promotion Support Program, University of Tsukuba Hospital (Reference Number: Senshin-99), which support the data management and monitoring of the trial.


The nerve-sparing (NS) effect of robot-assisted radical prostatectomy (RARP) on patients with a high-risk prostate cancer remains unclear. The objective of this study was to compare the urinary continence, erectile function and oncology outcomes of the nerve-sparing and non-nerve-sparing (NNS) group during RARP surgeries.We systematically searched databases including PubMed, Embase, Cochrane Library and Web of Science to identify relevant studies published in English up to December 2022. Newcastle-Ottawa Scale (NOS) was used as a quality evaluation tool to evaluate the quality of the literature parameters involved, including urinary continence, erectile function and oncologic outcomes, which were compared using the Stata 15.1 software .A total of 8 cohort studies involving 2499 patients were included. A meta-analysis of results showed that the NS group was beneficial to the recovery of urinary continence  and erectile function  12 months after surgeries, which showed a better oncological outcome .The current study results indicate that intraoperative NS during RARP is beneficial to long-term postoperative functional recovery and tumor prognosis of patients with high-risk prostate cancers. Due to interstudy interferences, the results should be interpreted with caution.https://www.crd.york.ac.uk/PROSPERO/, identifier: CRD42022384647. Prostate cancer (Pca) is the second most common solid tumor for men . In the Nerve sparing (NS) is a topic well worth exploring in the surgical management of patients with high-risk Pca, and many studies have confirmed that NS helps to improve the functional prognosis of patients in radical prostatectomy –10. SeveTherefore, we conducted a meta-analysis of the published clinical studies on the outcomes of NS in RARP to provide the latest evidence for clinicians to make decisions.Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) (https://www.crd.york.ac.uk/PROSPERO/) with a registration number of CRD42022384647.We conducted a systematic review and meta-analysis following the guideline of the (PRISMA) . A relevThe databases Pubmed, Cochrane Library, Web of Science and Embase were searched, and the deadline for literature search was December 8, 2022. According to the PICOS standard definition of search terms, literature search was conducted by combining the topic with free terms: (((high-risk) AND ((prostate cancer) OR (prostate neoplasm))) AND ((NS) OR (neurovascular bundle sparing))) AND  OR (RARP)).The literature to be searched was written in English language only, so we also manually searched and reviewed the relevant references to avoid any omission.We used the PICOS approach to define the inclusion criteria. P (patients): all the patients were diagnosed with a high-risk prostate cancer  and undergone RARP; I (intervention): NS, bilateral nerve-sparing (BNS), unilateral nerve-sparing (UNS) or partial nerve-sparing (PNS); C (comparator): non-nerve-sparing; O (outcome): one or more of the following outcomes: EF, urinary continence and oncologic outcomes; S (study type): cohort studies. Exclusion criteria (1): non-comparative studies; (2) editorial comments, meeting abstracts, case reports, unpublished studies, or reviews; (3) studies with unavailable data for analyses; (4) other studies that did not meet the inclusion criteria.Data extraction was carried out independently by the two reviewers, which were as follows: (1) general information: the first author extracted the years of publication and countries; (2) population characteristics: the number of patients, age, sample size, criteria to the assess recovery of urinary continence and EF; (3) outcomes: continence recovery (defined as the use of no pad or one safety pad/day), EF recovery , oncologic outcomes: positive surgical margins (PSM). Any discrepancy was resolved through a consensus or consultation with a third reviewer.https://www.ohri.ca//programs/clinical_epidemiology/oxford.asp); the quality of data extracted from each study was assessed based on case selection, comparability and outcome reporting. A meta-analysis was performed using the Stata 15.1 software . Risk ratio (RR) was used as a dichotomous variable. Statistical heterogeneity was evaluated based on I2 statistics; I2 ≥ 50% indicated a significant heterogeneity, and a random effects model was employed; when I2 < 50%, a fixed effects model was employed; p ≤ 0.05 was the threshold for statistical significance   scale. EF recovery was defined as SHIM≥17 or an erectile hardness meeting sexual intercourse requirements. The results were evaluated  studies –22. Howe studies , 21, so  studies Figure 3The meta-analysis showed that the NSS group presented higher positive surgical margin (PSM) rates compared with the NS group  Figure 4We performed a meta-analysis on PSM according to the type of NS. The results showed that the BNS group presented lower PSM rates compared with the NNS group . There was no significant difference in the PSM of the NNS group and the U(P)NS group . The BNS group showed lower PSM rates compared with the U(P)NS group  Figure 5The result showed no apparent asymmetry, which indicated no obvious publication bias Figure 6Some significant findings on the urinary continence, EF and oncologic outcomes of the present study warrant in-depth discussions.via reattachment of the tendon arch . Secondly, due to limited data, the situation of 1, 3, 6 and 24 months after surgeries was not further discussed. In addition, although a comparison of baseline data between the 2 treatment groups was described in the 3 studies included, fewer details were described. Therefore, it is unknown whether our interpretation of results would be affected. We expect that in follow-up studies, more attention will be paid to this point. Moreover, we have no consensus on the classification of NS, nor is there a standard classification of NS at present. Postoperative outcomes may be different due to different types of NS, and the result depends on its percentage , 46. TheDue to the differences in postoperative follow-ups, surgical techniques and medical levels in different regions among the studies included, as well as the inevitable selective bias in the NSS group in terms of oncology characteristics. Our results need to be carefully interpreted and verified through high-quality studies.NS is feasible during RARP surgeries, which is beneficial for the long-term (12 months after surgeries) urination continence and EF recovery of patients with high-risk Pca and is associated with better oncological outcomes. It is encouraging to find that NS may significantly improve the life quality of patients high-risk Pca after surgeries, which thus actively cooperates with treatments. However, due to interference factors in studies, it is not clear whether this advantage is caused by NS, and clinical decisions should be combined with various considerations on whether to perform NS.The original contributions presented in the study are included in the article/supplementary material. Further inquiries can be directed to the corresponding author.YL, X-ZD, JQ and X-SY: Protocol development, data collection and management, data analysis and manuscript writing. ZW, YJ and JH: Data management, data analysis and manuscript writing, data management, data analysis and manuscript writing. C-JW, LW, K-PL and J-HW: Data management and manuscript writing. All authors contributed to the article and approved the submitted version.


Over the last few decades, small molecule cancer drug discovery and development have focused on personalized medicine, emphasizing molecularly targeted drugs. Because of their small size , small molecules have been successfully used to target the extracellular, cell surface ligand-binding receptors and the intracellular proteins that play key roles in transducing downstream signalling for cancer cell survival, proliferation, and metastasis. Small molecule cancer drugs cover a wide range of inhibitors, including kinase inhibitors, proteasome inhibitors, matrix metalloproteinase inhibitors, etc. Research on molecularly targeted cancer drug discovery has resulted in some small molecule drugs being successfully introduced in the clinic for cancer treatment. Although many small molecules cancer drugs have been developed and are currently being utilized in the clinic, many challenges remain, such as a low response rate and drug resistance.The current Research Topic would like to raise such issues for discussion; and how we could overcome the problems by understanding the mechanisms underlying the effect of small molecule cancer drugs. This Research Topic is under the Pharmacology of Anti-Cancer Drugs section of the journal Frontiers in Pharmacology, a sum of seven contributing articles  have been included, and we would like to summarize their findings in this editorial article. The works can be divided into two categories, based on their experimental approaches.Qu et al. aimed to study the known anti-tumor compound, calycosin, on thyroid cancer. Calycosin is one of the main components of astragalus, which could induce the apoptosis via PI3K/AKT/mTOR signaling pathway. However, its anti-cancer mechanism on papillary thyroid cancer has not been fully described. Authors underwent the microarray analysis on the B-CPAP cells and, confirmed that calycosin could influence the apoptotic genes and p70S6 Kinase via the Sestrin2/AMP-activated protein kinase/mammalian target of rapamycin complex (SESN2/AKT/mTOR) pathway.In this Research Topic, three types of cancers have been studied by various compounds. Otkur et al. aimed to find out if the orphan G_protein coupled receptor GPR35 antagonists could serve as a good candidate in inhibiting the colorectal cancer. They showed that the GPR35 could stimulate the YAP/TAZ activity in Hippo signaling pathway through the Rho-GTPase. Besides, they identified that the GPR35 antagonist CID-2745687 could suppress the hyperactivation of YAP/TAZ in colorectal cancer.A study on colorectal cancer by Hu et al. here purposed the anti-tumor feature of N-linoleyltyrosine (NlTyr), an anandamide analog, in A549 cells. They found that NlTyr could active the endocannabinoid receptors via the PI3K and ERK pathways, which resulted in cell cycle arrest, inhibited migration, and induced apoptosis in the non-small cell lung cancer cells.Non-small cell lung cancer is the major types of lung cancers that has a very low 5-year overall survival rate. Li et al. summarized the current findings on the PAK4 mediated singaling pathways, and their roles in cancers. The article described the potential anti-tumor abilities of PAK4 inhibitors, and classified them into ATP competitive inhibitors, and allosteric inhibitors. They further investigated the preclinical and clinical studies of two selected inhibitors, PF-3758309 and KPT-9274, which was terminated and in phase I clinical trials, respectively. The article ended with the upcoming challenges, limitations, and prospective on PAK4 usage.One review paper on the potential use of p21-activated kinase 4 (PAK4) inhibitors was published in this Research Topic. Chen et al. applied the network pharmacology-based prediction and in vitro cell line models on the coumarin derivatives and demonstrated that the compound 6e could suppress migration, invasion, and induce mitochondria dependent apoptosis via the PI3K/AKT-mediated BCL-2 signaling pathway.Network pharmacology is a powerful tool to characterize the action mechanisms of undeveloped or complicated derivatives in drug discovery. Taghvaei et al. performed the in silico database screening that targeting on the Sentrin-specific protease 1 (SENP1). SENP1 participates in cancer development by inhibit the apoptosis, and stimulate the cell proliferation and migration. Besides, it promotes the angiogenesis that shape it as a common target for cancer drugs. The group screened for 84,000 compounds and selected final 20 compounds after the molecular docking analysis. They then identified that resveratrol and ZINC33916875 could act as the SENP1 inhibitor after the essential dynamics and molecular dynamics analysis.Cai et al. aimed to understand the clinical benefit of different regiments (double-drug or triple-drug) for left-sided RAS wild-type metastatic colorectal cancer patients. They performed a network meta-analysis by using the published data and real-world data, and concluded that a double-drug regimen plus cetuximab/panitumumab is more effective than double-drug- and triple-drug-based regimens with bevacizumab for left-sided RAS wild-type metastatic colorectal cancer. Besides, they claimed that the double-drug regimen plus cetuximab/panitumumab is safer than triple-drug regimen with bevacizumab via the safety analysis.To summarize, the Research Topic has provided several new potential anti-cancer compounds by various approaches, such as performing meta-analysis on patient data to identify the clinical benefit of treatments; biological experiments in cell models to identify the molecular mechanism of the compounds; computational analysis on compounds’ structure and docking abilities. We hope that the Research Topic could provide some directions to researchers for searching the new anti-cancer drugs.


Klebsiella pneumoniae are a leading cause of healthcare-associated infections worldwide. In particular, strains expressing extended-spectrum β-lactamases (ESBLs) and carbapenemases pose serious treatment challenges, leading the World Health Organization (WHO) to designate ESBL and carbapenem-resistant Enterobacteriaceae as ‘critical’ threats to human health. Research efforts to combat these pathogens can be supported by accessibility to diverse and clinically relevant isolates for testing novel therapeutics. Here, we describe a panel of 100 diverse K. pneumoniae isolates that are publicly available to assist the research community in this endeavour. Whole-genome sequencing (WGS) was performed on 3878 K. pneumoniae clinical isolates housed at the Multidrug-Resistant Organism Repository and Surveillance Network. The isolates were cultured from 63 facilities in 19 countries between 2001 and 2020. Core-genome multilocus sequence typing and high-resolution single-nucleotide polymorphism-based phylogenetic analyses captured the genetic diversity of the collection and were used to select the final panel of 100 isolates. In addition to known multidrug-resistant (MDR) pandemic lineages, the final panel includes hypervirulent lineages and isolates with specific and diverse resistance genes and virulence biomarkers. A broad range of antibiotic susceptibilities, ranging from pan-sensitive to extensively drug-resistant isolates, are described. The panel collection, and all associated metadata and genome sequences, are available at no additional cost and will be an important resource for the research community and for the design and development of novel antimicrobial agents and diagnostics against this important pathogen. K. pneumoniae panel isolates described in this study are available at BEI resources (https://www.beiresources.org/) at no cost under catalogue #NR-55604. Metadata for the 100 panel isolates can be accessed in Table S1 (available in the online version of this article) and on Microreact: A Panel of Diverse Klebsiella pneumoniae Clinical Isolates for Research and Development .Data pertinent to this study have been deposited in National Center for Biotechnology Information (NCBI) BioProject PRJNA717739. The 100 Klebsiella pneumoniae is a major cause of healthcare-associated infections that are increasingly difficult to treat due to the emergence of multidrug-resistant strains. In particular, strains expressing extended-spectrum β-lactamases (ESBLs) and carbapenemases have attained global notoriety, with the World Health Organization (WHO) listing these strains as a ‘critical priority’ for the development of new therapeutics. Access to a diverse collection of strains for testing is critical for this endeavour, but few resources currently exist. Similarly, pivotal research on the genetic determinants underlying the pathogenesis of hypervirulent lineages is hampered by the lack of standardized comparator strains. Herein we describe a panel of 100 diverse K. pneumoniae constructed to maximize genetic and phenotypic diversity from a repository of over 3800 clinical isolates collected over 19 years. The panel, and all associated metadata and genome sequences, are provided at no cost and will greatly assist efforts by academic, government and industry research groups.Klebsiella pneumoniae are a leading cause of nosocomial infections resulting in pneumonia, bacteraemia, surgical site and urinary tract infections , metabolite transporter peg-344, genotoxic polyketide colibactin (clb), and regulators of mucoviscosity and capsular polysaccharide (rmpA and rmpA2)]  . While trminants . This corminants .K. pneumoniae population is diverse, consisting of over 250 clonal phylogenetic lineages and an estimated accessory genome of >100 000 protein coding sequences  to colistin by broth microdilution (BMD). As the CLSI guidelines [mgrB (761 403 and 375 436) were assigned an intermediate interpretation , as reported previously [Carbapenemase genes encoding IMP, KPC, NDM, OXA-48-like and VIM enzymes were present in 24 isolates. Eleven isolates produced OXA-48-like ctamases , all stridelines  do not reviously .armA, rmtB1, rmtF1, or rmtH), with the exception of MRSN 430405 for which no acquired RMTase gene was identified (Table S1). Specifically, five of the pan-aminoglycoside-resistant isolates  carried rmtH, rmtF1, or rmtB1, and the remaining three  carried armA. MRSN 752729 also carried armA but was susceptible to amikacin and gentamicin. Reduced susceptibility was confirmed by BMD . Further analysis of the armA sequence revealed a missense mutation at nucleotide position 617 (A to T) resulting in an amino acid substitution of isoleucine to lysine.Forty-two isolates were susceptible to all three aminoglycosides tested  while nine isolates were pan-resistant to all aminoglycosides. Of the latter, all carried a 16S rRNA methyltransferase  named carbapenem-resistant and ESBL-producing Enterobacterales as ‘urgent’ and ‘serious’ threats, respectively . As a rebaumanii  and P.ginosa . Herein K. pneumoniae strains exist, but they mainly focus on the identification of antibiotic resistance mechanisms and were not designed to encompass the diversity of the species. For example, the US CDC and Food and Drug Administration (FDA) have collaborated to produce the AR Isolate Bank that contains multiple isolate panels for a range of bacterial pathogens and resistance mechanisms (https://wwwn.cdc.gov/arisolatebank/Overview), and this panel has proven to be an excellent resource to test the activity of antibiotic combinations [Klebsiella phages is the external capsular polysaccharide, although recent work suggests that phages also attach to other outer-membrane structures below the capsule, including the O-antigen [Klebsiella phages. Besides therapeutics, the understanding of K. pneumoniae pathogenesis is rapidly evolving, in particular the understanding of virulence factors that can accurately predict the pathogenic potential of strains. For example, not all hvKp strains are equally virulent in murine models of infection despite carrying well-characterized virulence biomarkers [Other panels and characterized inations . Howeverinations . The mai-antigen . The pan-antigen  and 8 pr-antigen , providiomarkers . Herein K. pneumoniae over the past two decades has been characterized by widely geographically distributed ‘high-risk’ clones and the constant emergence and dissemination of new clonal groups [blaCTX-M-15, and five genomic convergent lineages that have not been previously described . Furthermore, close attention was paid to selecting rare clones that cause localized epidemics in different regions of the world. Clones ST-43, ST-268, ST-340 and ST-392 are all represented in the panel and have been reported previously as harbouring NDM carbapenemases and circulating in hospitalized patients in Iran [The epidemiology of l groups . This pa in Iran . Similar in Iran , and inf in Iran  and Sout in Iran . Similar in Iran . In our blaSHV-27 ESBL had a non-ESBL phenotype. However, this discrepancy is most likely due to a base pair substitution (A to C) in the promoter region that was previously reported in SHV-27-producing isolates susceptible to cephalosporins [armA RMTase, had increased susceptibility to all aminoglycoside antibiotics. Previous studies report that point mutations in armA can result in the inability to bind to the 16S rRNA and consequently block methylation, resulting in aminoglycoside susceptibility [K. pneumoniae to have minimal to no carbapenemase activity [Notably, a strong association between antibiotic susceptibility and the presence of AMR genes and/or mutations was observed, with few exceptions. For example, isolates carrying the osporins . Similartibility . Finallytibility , yet stuactivity , consistWhile strains in this panel originated from around the globe, all were isolated from the clinic, which likely explains the scarcity of hvKp isolates that are usually associated with community-acquired infections. As such, the final panel does not contain a prototypical ST-23 hvKp isolate. Instead, strains from hvKp lineages found globally , as well as an alarming ST-23 convergent isolate, were chosen. More generally, other region-specific or recently emerging clones may not be represented in the final panel, which was capped at 100 isolates to provide a standardized, pragmatic and easy-to-use resource for the community.K. pneumoniae isolates from an extensive collection of over 3800 K. pneumoniae isolates collected from across the globe. The panel encompasses the diversity of the species, includes both antibiotic-susceptible and non-susceptible isolates, and captures known epidemic clones as well as sporadic ones. Furthermore, this panel captures diverse genomic convergent and hvKp strains that are rapidly emerging worldwide and are of considerable concern [In summary, this study describes the construction of a panel of 100 unique  concern . 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Intracellular liquid–liquid phase separation (LLPS) is a critical process involving the dynamic association of biomolecules and the formation of non-membrane compartments, playing a vital role in regulating biomolecular interactions and organelle functions. A comprehensive understanding of cellular LLPS mechanisms at the molecular level is crucial, as many diseases are linked to LLPS, and insights gained can inform drug/gene delivery processes and aid in the diagnosis and treatment of associated diseases. Over the past few decades, numerous techniques have been employed to investigate the LLPS process. In this review, we concentrate on optical imaging methods applied to LLPS studies. We begin by introducing LLPS and its molecular mechanism, followed by a review of the optical imaging methods and fluorescent probes employed in LLPS research. Furthermore, we discuss potential future imaging tools applicable to the LLPS studies. This review aims to provide a reference for selecting appropriate optical imaging methods for LLPS investigations. I.in vivo.Cellular compartmentalization ensures that various essential cellular biochemical activities are carried out in an orderly, efficient, and precise manner. Classical membrane-enclosed compartmentalizations, such as mitochondria, Golgi apparatus, endoplasmic reticulum, and cell nucleus chloroplasts, play crucial roles in this process.2 response.LLPS is intimately linked to the pathogenesis of various diseases, including neurodegenerative disease such as amyotrophic lateral sclerosis (ALS),in vitro remodeling .Common research techniques in LLPS studies include computational approach using sequence analysis (to identify LLPS protein),II.2/O2 levels, light exposure, crowding agents, pH, and ATP, among others , 868 proteins are associated with LLPS.According to PhaSepDB2.1, a database dedicated to phase-separation-related proteins  by LLPS, while in late spermatids, FXR1 activates the translation of mRNAs by recruiting translational machinery.Furthermore, LLPS of the coronavirus SARS-CoV-2 nucleocapsid (N) protein and genomic RNA not only facilitates their assembly and transcription but also diminishes stress granule formation and suppresses the innate antiviral immune response in host cells.III.A.in vitro, making it widely used in the study of the properties, functions, and kinetics of LLPS in vitro.Differential interference contrast (DIC) is an imaging technique that converts the optical path gradient of a specimen into an amplitude difference image. A DIC microscope requires a polarizer, an analyzer, and a Wollaston prism, as illustrated in et al. studied the phase separation of RNA molecules in vitro using DIC microscopy and hyperspectral stimulated Raman microscopy.2+. The Mg2+ regulation of RNA droplets may be closely related to the general aging process of RNA-containing membrane-less organelles. As shown in 2+ concentration. In addition, DIC microscopy is a common tool for exploring the mechanism of protein Tau phase separation in vitro.in vitro.Tang in vitro. It is particularly effective for characterizing LLPS due to its ability to provide higher contrast than bright field microscopy. This high contrast originates from the differences in refractive indices within the specimen. Phase-separated droplets are typically transparent, rendering small droplets undetectable under bright field microscopy. However, with DIC microscopy, these droplets exhibit better contrast because of the refractive indices difference. DIC microscopy lacks the specificity required to capture small LLPS droplets inside cells. The complexity of intracellular components necessitates the use of specific fluorescent labels to study intracellular biological events. Therefore, fluorescence microscopy is a more efficient tool for investigating the phase separation process in cells compared to DIC microscopy.DIC microscopy is a valuable technique for studying the morphological changes of phase-separated droplets B.in vivo and in vitro. Fluorescence microscopy provides several advantages for studying LLPS, including high controllability, adjustable environmental conditions, and good repeatability in vitro. Additionally, it offers intuitive visualization, non-invasiveness, high spatiotemporal resolution, and reversibility in vitro. However, due to the large illumination depth of wide-field fluorescence microscope, sample signals in out-of-focus areas are also collected, significantly reducing the signal-to-noise ratio of imaging. To solve this problem, confocal fluorescence microscopy imaging technology has been proposed.Fluorescence microscopy is a widely used technique that relies on autofluorescence or fluorescent tags to image samples. It enables the examination of cells or tissues that have been specifically labeled with fluorescent markers. The structure of fluorescence wide-field microscopy is demonstrated in in vitro or in vivo can be captured with high efficiency by confocal microscopy.et al. showed that mammalian oocytes can store mRNAs in mitochondria-associated membrane-less compartments.et al. investigated the formation of phase-separated droplets by FXR1 in spermatids in vivo and in vitro using confocal microscopy.Over the past 30 years, confocal microscopy has become an important tool in biological studies. In confocal microscopy, the focused beam replaces the wide field beam as the excitation light source . The sigConfocal microscopy can produce high-resolution images by finely sectioning biological samples. In particular, it is a powerful tool to study the formation or properties of LLPS. However, its limitations in acquisition speed render it unsuitable for imaging highly dynamic processes. The point excitation characteristic of confocal microscopy results in greater phototoxicity compared to wide-field illumination. Furthermore, achieving single-molecule level imaging with confocal microscopy is challenging. Therefore, capturing the dynamics at the early stage of phase separation droplet formation, such as the transition process from monomeric molecules to micrometer-scale phase separation droplets, remains a challenge for confocal microscopy.To enhance acquisition speed and reduce phototoxicity, a modified version of the conventional confocal microscope known as the spinning disk confocal microscope has been developed.C.et al. utilized TIRF-M to study the LLPS of Escherichia coli single-stranded DNA-binding protein (EcSSB).et al. used TIRF microscopy to study the formation process of P62/SQSTM1,Total internal reflection fluorescence (TIRF) microscopy (TIRF-M) is a widely used optical technique in cell biology, particularly for analyzing cell membranes, studying the localization and dynamics of single molecules.However, TIRF-M is limited to imaging near the surface, and the droplet size formed by LLPS can be smaller than the optical diffraction limit in the early stages. This makes it challenging to accurately characterize and quantify droplets using traditional fluorescence microscopes. Thus, the fluorescence microscope with higher resolution (super-resolution fluorescence microscope) should be used to study the characteristics and formation kinetics of LLPS.D.Biological events occurring at nanometer scales (5–100 nm) are of great interest for understanding biological dynamics, functions, and structures.Photo-activated localization microscopy (PALM) and stochastic optical reconstruction microscopy (STORM) are two commonly used single molecule localization super-resolution microscopy (SMLM) techniques that have been applied in many biological studies.STORM has a similar operating principle to PALM. However, compared with PALM, STORM improves the rate of super-resolution image acquisition. The photobleaching of active fluorophores in PALM is considered an additional time-consuming step. Thus, STORM replaces this step with light-switchable fluorophores, which can be activated (turned on) and deactivated (turned off) at will . This imet al. presented a comprehensive protocol regarding the use of super-resolution-microscopy to study LLPS  is a super-resolution microscopy technique that utilizes interference patterns created between the sample and the illumination pattern.E.in vivo and in vitro.Another method for measuring the kinetics of fluorescently labeled molecules is fluorescence correlation spectroscopy (FCS), which is a single-molecule fluorescence detection technique.et al. captured the formation of nanoscale condensates using a dual color fluorescence cross correlation spectroscopy (dcFCCS) method thiazol-3-ium trifluoroacetate (TASG), which not only binds to the core SG protein G3BP1 and its complexes but also exhibits minimal fluorescence in the absence of SG.There are also some small-molecule fluorescent probes available for LLPS live-cell imaging. Shao in situ and monitor the maturation of assembled amyloid fibrils. To summarize, while immunostaining involves complex processes and lacks real-time monitoring capabilities, it offers advantages such as high affinity, high sensitivity, and high accuracy.Immunostaining is also used to image proteins and droplets of LLPS in fixed cells and tissue sections using antibodies. In addition, Congo Red,in vitro, recombinant proteins are expressed using procaryotic expression system and labeled with amine-reactive dyes. For instance, Zhang et al. labeled Δtau187 with Alexa Fluor 488 NHS Ester and studied the droplet formation process in the presence of poly(A) RNA.in vitro.To remodel LLPS In the context of LLPS, using fluorescent probes presents two main challenges. The first challenge concerns the effect of viscosity on the fluorescence quantum yield of the probe.V.The LLPS-induced cellular condensates possess multiple biochemical and biological functions along with unique biophysical properties.When conducting LLPS studies, it is important to carefully select the appropriate microscopy based on the research object, and correlative optical imaging can be used if necessary . For exaChoosing the appropriate optical imaging method for LLPS studies involves several factors, including sample size and thickness, imaging speed, and potential phototoxicity and photobleaching effects. For example, DIC, wide-field microscopy, and confocal microscopy are suitable for imaging thicker samples, while TIRF microscopy is ideal for thin samples. High-resolution imaging can be achieved using techniques such as SMLM, SIM, and TIRF. Some imaging techniques may cause phototoxicity or photobleaching, which can affect the sample over time. Methods, such as FCS and Capflex imaging, are less likely to cause such issues. Cost is also a consideration when selecting an imaging technique. In summary, the choice of the imaging method for LLPS studies will depend on various factors. It may be necessary to use multiple techniques to obtain a comprehensive understanding of LLPS behavior.In recent years, real-time 3D single molecule tracking has emerged as a powerful tool for studying molecular dynamical interactions, thanks to its high spatiotemporal precision, high sensitivity, and large imaging field.