Multimaterial decomposition in dual-energy CT for characterization of clots from acute ischemic stroke patients.

BACKGROUND: Nowadays, there is no method to quantitatively characterize the material composition of acute ischemic stroke thrombi prior to intervention, but dual-energy CT (DE-CT) offers imaging-based multimaterial decomposition. We retrospectively investigated the material composition of thrombi ex vivo using DE-CT with histological analysis as a reference. METHODS: Clots of 70 patients with acute ischemic stroke were extracted by mechanical thrombectomy and scanned ex vivo in formalin-filled tubes with DE-CT. Multimaterial decomposition in the three components, i.e., red blood cells (RBC), white blood cells (WBC), and fibrin/platelets (F/P), was performed and compared to histology (hematoxylin/eosin staining) as reference. Attenuation and effective Z values were assessed, and histological composition was compared to stroke etiology according to the Trial of ORG 10172 in Acute Stroke Treatment (TOAST) criteria. RESULTS: Histological and imaging analysis showed the following correlation coefficients for RBC (r = 0.527, p < 0.001), WBC (r = 0.305, p = 0.020), and F/P (r = 0.525, p < 0.001). RBC-rich thrombi presented higher clot attenuation in Hounsfield units than F/P-rich thrombi (51 HU versus 42 HU, p < 0.01). In histological analysis, cardioembolic clots showed less RBC (40% versus 56%, p = 0.053) and more F/P (53% versus 36%, p = 0.024), similar to cryptogenic clots containing less RBC (34% versus 56%, p = 0.006) and more F/P (58% versus 36%, p = 0.003) than non-cardioembolic strokes. No difference was assessed for the mean WBC portions in all TOAST groups. CONCLUSIONS: DE-CT has the potential to quantitatively characterize the material composition of ischemic stroke thrombi. RELEVANCE STATEMENT: Using DE-CT, the composition of ischemic stroke thrombi can be determined. Knowledge of histological composition prior to intervention offers the opportunity to define personalized treatment strategies for each patient to accomplish faster recanalization and better clinical outcomes. KEY POINTS: • Acute ischemic stroke clots present different recanalization success according to histological composition. • Currently, no method can determine clot composition prior to intervention. • DE-CT allows quantitative material decomposition of thrombi ex vivo in red blood cells, white blood cells, and fibrin/platelets. • Histological clot composition differs between stroke etiology. • Insights into the histological composition in situ offer personalized treatment strategies.

Validating a minipig model of reversible cerebral demyelination using human diagnostic modalities and electron microscopy.

BACKGROUND: Inflammatory demyelinating diseases of the central nervous system, such as multiple sclerosis, are significant sources of morbidity in young adults despite therapeutic advances. Current murine models of remyelination have limited applicability due to the low white matter content of their brains, which restricts the spatial resolution of diagnostic imaging. Large animal models might be more suitable but pose significant technological, ethical and logistical challenges. METHODS: We induced targeted cerebral demyelinating lesions by serially repeated injections of lysophosphatidylcholine in the minipig brain. Lesions were amenable to follow-up using the same clinical imaging modalities (3T magnetic resonance imaging, 11C-PIB positron emission tomography) and standard histopathology protocols as for human diagnostics (myelin, glia and neuronal cell markers), as well as electron microscopy (EM), to compare against biopsy data from two patients. FINDINGS: We demonstrate controlled, clinically unapparent, reversible and multimodally trackable brain white matter demyelination in a large animal model. De-/remyelination dynamics were slower than reported for rodent models and paralleled by a degree of secondary axonal pathology. Regression modelling of ultrastructural parameters (g-ratio, axon thickness) predicted EM features of cerebral de- and remyelination in human data. INTERPRETATION: We validated our minipig model of demyelinating brain diseases by employing human diagnostic tools and comparing it with biopsy data from patients with cerebral demyelination. FUNDING: This work was supported by the DFG under Germany's Excellence Strategy within the framework of the Munich Cluster for Systems Neurology (EXC 2145 SyNergy, ID 390857198) and TRR 274/1 2020, 408885537 (projects B03 and Z01).

Fluorescein-stained confocal laser endomicroscopy versus conventional frozen section for intraoperative histopathological assessment of intracranial tumors.

BACKGROUND: The aim of this clinical trial was to compare Fluorescein-stained intraoperative confocal laser endomicroscopy (CLE) of intracranial lesions and evaluation by a neuropathologist with routine intraoperative frozen section (FS) assessment by Neuropathology. METHODS: In this phase II non-inferiority, prospective, multicenter, non-randomized, off-label clinical trial (Eudra-CT: 2019-004512-58), patients above the age of 18 years with any intracranial lesion scheduled for elective resection were included. The diagnostic accuracies of both CLE and FS referenced with the final histopathological diagnosis were statistically compared in a non-inferiority analysis, representing the primary endpoint. Secondary endpoints included the safety of the technique and time expedited for CLE and FS. RESULTS: 210 patients were included by 3 participating sites between November 2020 and June 2022. Most common entities were high grade gliomas (37.9%), metastases (24.1%), and meningiomas (22.7%), A total of 6 serious adverse events in 4 (2%) patients were recorded. For the primary endpoint, the diagnostic accuracy for CLE was inferior with .87 versus .91 for FS, resulting in a difference of .04 (95% confidence interval -.10; .02; p=.367). The median time expedited until intraoperative diagnosis was 3 minutes for CLE and 27 minutes for FS, with a mean difference of 27.5 minutes (standard deviation 14.5; p<.001). CONCLUSION: CLE allowed for a safe and time-effective intraoperative histological diagnosis with a diagnostic accuracy of 87% across all intracranial entities included. The technique achieved histological assessments in real-time with a tenfold reduction of processing time compared to FS, which may invariably impact surgical strategy on the fly.

Enhanced Sensitivity to ALDH1A3-Dependent Ferroptosis in TMZ-Resistant Glioblastoma Cells.

Temozolomide (TMZ) is standard treatment for glioblastoma (GBM); nonetheless, resistance and tumor recurrence are still major problems. In addition to its association with recurrent GBM and TMZ resistance, ALDH1A3 has a role in autophagy-dependent ferroptosis activation. In this study, we treated TMZ-resistant LN229 human GBM cells with the ferroptosis inducer RSL3. Remarkably, TMZ-resistant LN229 clones were also resistant to ferroptosis induction, although lipid peroxidation was induced by RSL3. By using Western blotting, we were able to determine that ALDH1A3 was down-regulated in TMZ-resistant LN229 cells. Most intriguingly, the cell viability results showed that only those clones that up-regulated ALDH1A3 after TMZ withdrawal became re-sensitized to ferroptosis induction. The recovery of ALDH1A3 expression appeared to be regulated by EGFR-dependent PI3K pathway activation since Akt was activated only in ALDH1A3 high clones. Blocking the EGFR signaling pathway with the EGFR inhibitor AG1498 decreased the expression of ALDH1A3. These findings shed light on the potential application of RSL3 in the treatment of glioblastoma relapse.

Multiscale quantification of morphological heterogeneity with creation of a predictor of longer survival in glioblastoma.

Intratumor heterogeneity is a main cause of the dismal prognosis of glioblastoma (GBM). Yet, there remains a lack of a uniform assessment of the degree of heterogeneity. With a multiscale approach, we addressed the hypothesis that intratumor heterogeneity exists on different levels comprising traditional regional analyses, but also innovative methods including computer-assisted analysis of tumor morphology combined with epigenomic data. With this aim, 157 biopsies of 37 patients with therapy-naive IDH-wildtype GBM were analyzed regarding the intratumor variance of protein expression of glial marker GFAP, microglia marker Iba1 and proliferation marker Mib1. Hematoxylin and eosin stained slides were evaluated for tumor vascularization. For the estimation of pixel intensity and nuclear profiling, automated analysis was used. Additionally, DNA methylation profiling was conducted separately for the single biopsies. Scoring systems were established to integrate several parameters into one score for the four examined modalities of heterogeneity (regional, cellular, pixel-level and epigenomic). As a result, we could show that heterogeneity was detected in all four modalities. Furthermore, for the regional, cellular and epigenomic level, we confirmed the results of earlier studies stating that a higher degree of heterogeneity is associated with poorer overall survival. To integrate all modalities into one score, we designed a predictor of longer survival, which showed a highly significant separation regarding the OS. In conclusion, multiscale intratumor heterogeneity exists in glioblastoma and its degree has an impact on overall survival. In future studies, the implementation of a broadly feasible heterogeneity index should be considered.

Alpha-synuclein fibrils amplified from multiple system atrophy and Parkinson's disease patient brain spread after intracerebral injection into mouse brain.

Parkinson's disease (PD), multiple system atrophy (MSA), and dementia with Lewy bodies (DLB) are neurodegenerative disorders with alpha-synuclein (α-syn) aggregation pathology. Different strains of α-syn with unique properties are suggested to cause distinct clinical and pathological manifestations resulting in PD, MSA, or DLB. To study individual α-syn spreading patterns, we injected α-syn fibrils amplified from brain homogenates of two MSA patients and two PD patients into the brains of C57BI6/J mice. Antibody staining against pS129-α-syn showed that α-syn fibrils amplified from the brain homogenates of the four different patients caused different levels of α-syn spreading. The strongest α-syn pathology was triggered by α-syn fibrils of one of the two MSA patients, followed by comparable pS129-α-syn induction by the second MSA and one PD patient material. Histological analysis using an antibody against Iba1 further showed that the formation of pS129-α-syn is associated with increased microglia activation. In contrast, no differences in dopaminergic neuron numbers or co-localization of α-syn in oligodendrocytes were observed between the different groups. Our data support the spreading of α-syn pathology in MSA, while at the same time pointing to spreading heterogeneity between different patients potentially driven by individual patient immanent factors.

A rare case of H3K27-altered diffuse midline glioma with multiple osseous and spinal metastases at the time of diagnosis.

BACKGROUND: H3K27-altered diffuse midline gliomas are uncommon central nervous system tumors with extremely poor prognoses. CASE PRESENTATION: We report the case of a 24-year-old man patient with multiple, inter alia osseous metastases who presented with back pain, hemi-hypoesthesia, and hemi-hyperhidrosis. The patient underwent combined radio-chemotherapy and demonstrated temporary improvement before deteriorating. CONCLUSIONS: H3K27-altered diffuse midline glioma presents an infrequent but crucial differential diagnosis and should be considered in cases with rapid neurological deterioration and multiple intracranial and intramedullary tumor lesions in children and young adults. Combined radio-chemotherapy delayed the neurological deterioration, but unfortunately, progression occurred three months after the diagnosis.

Design and Transition of an Emergency E-Learning Pathology Course for Medical Students-Evaluation of a Novel Course Concept.

BACKGROUND: Around the world, the emergency brought about by the COVID-19 pandemic forced medical schools to create numerous e-learning supplements to provide instruction during this crisis. The question now is to determine a way in which to capitalize on this momentum of digitization and harness the medical e-learning content created for the future. We have analyzed the transition of a pathology course to an emergency remote education online course and, in the second step, applied a flipped classroom approach including research skills training. METHODS: In the summer semester of 2020, the pathology course at the Technical University of Munich was completely converted to an asynchronous online course. Its content was adapted in winter 2021 and incorporated into a flipped classroom concept in which research skills were taught at the same time. RESULTS: Screencasts and lecture recordings were the most popular asynchronous teaching formats. Students reported developing a higher interest in pathology and research through group work. The amount of content was very challenging for some students. CONCLUSION: Flipped classroom formats are a viable option when using pre-existing content. We recommend checking such content for technical and didactic quality and optimizing it if necessary. Content on research skills can be combined very well with clinical teaching content.

ALDH1-Mediated Autophagy Sensitizes Glioblastoma Cells to Ferroptosis.

The fatal clinical course of human glioblastoma (GBM) despite aggressive adjuvant therapies is due to high rates of recurrent tumor growth driven by tumor cells with stem-cell characteristics (glioma stem cells, GSCs). The aldehyde dehydrogenase 1 (ALDH1) family of enzymes has been shown to be a biomarker for GSCs, and ALDH1 seems to be involved in the biological processes causing therapy resistance. Ferroptosis is a recently discovered cell death mechanism, that depends on iron overload and lipid peroxidation, and it could, therefore, be a potential therapeutic target in various cancer types. Since both ALDH1 and ferroptosis interact with lipid peroxidation (LPO), we aimed to investigate a possible connection between ALDH1 and ferroptosis. Here, we show that RSL3-induced LPO and ferroptotic cell death revealed RSL3-sensitive and -resistant malignant glioma cell lines. Most interestingly, RSL3 sensitivity correlates with ALDH1a3 expression; only high ALDH1a3-expressing cells seem to be sensitive to ferroptosis induction. In accordance, inhibition of ALDH1a3 enzymatic activity by chemical inhibition or genetic knockout protects tumor cells from RSL3-induced ferroptotic cell death. Both RSL-3-dependent binding of ALDH1a3 to LC3B and autophagic downregulation of ferritin could be completely blocked by ALDH inhibition. Therefore, ALDH1a3 seems to be involved in ferroptosis through the essential release of iron by ferritinophagy. Our results also indicate that ferroptosis induction might be a particularly interesting clinical approach for targeting the highly aggressive cell population of GSC.

Five material tissue decomposition by dual energy computed tomography.

The separation of mixtures of substances into their individual components plays an important role in many areas of science. In medical imaging, one method is the established analysis using dual-energy computed tomography. However, when analyzing mixtures consisting of more than three individual basis materials, a physical limit is reached that no longer allows this standard analysis. In addition, the X-ray attenuation coefficients of chemically complicated basis materials may not be known and also cannot be determined by other or previous analyses. To address these issues, we developed a novel theoretical approach and algorithm and tested it on samples prepared in the laboratory as well as on ex-vivo medical samples. This method allowed both five-material decomposition and determination or optimization of the X-ray attenuation coefficients of the sample base materials via optimizations of objective functions. After implementation, this new multimodal method was successfully tested on self-mixed samples consisting of the aqueous base solutions iomeprol, eosin Y disodiumsalt, sodium chloride, and pure water. As a first proof of concept of this technique for detailed material decomposition in medicine we analyzed exact percentage composition of ex vivo clots from patients with acute ischemic stroke, using histological analysis as a reference standard.

Primary Bone Tumors of the Spine-Proposal for Treatment Based on a Single Centre Experience.

This study reports a large single-center series of primary bone tumors of the spine (PBTs). We aimed to review the concepts for management, as this kind of tumor represents a very rare entity, and also propose a new treatment algorithm. Retrospective analysis revealed 92 patients receiving surgery for PBTs from 2007 to 2019 at our center. They were analyzed based on surgical management and the course of the disease. A total of 145 surgical procedures were performed (50 cervical, 46 thoracic, 28 lumbar, and 21 sacral). Complete tumor resection was achieved in 65%, of which 22% showed tumor recurrence during follow-up (mean time to recurrence 334 days). The five-year mortality rate was significantly lower after complete resection (3% versus 25% after subtotal resection). Most of the patients improved in their symptoms through surgery. Regarding the tumor entity, the most common PBTs were vertebral hemangiomas (20%), osteoid osteomas (15%), and chordomas (16%). The Enneking graduation system showed a good correlation with the risk of recurrence and mortality. Complete resection in PBTs increased survival rates and remains the method of choice. Thus, quality of life-especially with a higher extent of resection-should be considered.

Progressive multifocal leukoencephalopathy associated with chemotherapy induced lymphocytopenia in solid tumors - case report of an underestimated complication.

Background: JC virus reactivation causing progressive multifocal leukoencephalopathy (PML) occurs preferentially in human immunodeficiency virus (HIV) positive individuals or patients suffering from hematologic neoplasms due to impaired viral control. Reactivation in patients suffering from solid malignancies is rarely described in published literature. Case Presentation: Here we describe a case of PML in a male patient suffering from esophageal cancer who underwent neoadjuvant radiochemotherapy and surgical resection in curative intent resulting in complete tumor remission. The radiochemotherapy regimen contained carboplatin and paclitaxel (CROSS protocol). Since therapy onset, the patient presented with persistent and progredient leukopenia and lymphopenia in absence of otherwise known risk factors for PML. Symptom onset, which comprised aphasia, word finding disorder, and paresis, was apparent 7 months after therapy initiation. There was no relief in symptoms despite standard of care PML directed supportive therapy. The patient died two months after therapy onset. Conclusion: PML is a very rare event in solid tumors without obvious states of immununosuppression and thus harbors the risk of unawareness. The reported patient suffered from lymphopenia, associated with systemic therapy, but was an otherwise immunocompetent individual. In case of neurologic impairment in patients suffering from leukopenia, PML must be considered - even in the absence of hematologic neoplasia or HIV infection.

Histology of Cerebral Clots in Cryptogenic Stroke Varies According to the Presence of a Patent Foramen Ovale.

Although a pathophysiological impact remains difficult to prove in individual patient care, a patent foramen ovale (PFO) is currently considered of high relevance for secondary prophylaxis in selected patients with cryptogenic ischemic stroke. By quantification of histological clot composition, we aimed to enhance pathophysiological understanding of PFO attributable ischemic strokes. Retrospectively, we evaluated all cerebral clots retrieved by mechanical thrombectomy for acute ischemic stroke treatment between 2011 and 2021 at our comprehensive stroke care center. Inclusion criteria applied were cryptogenic stroke, age (≤60 years), and PFO status according to transesophageal echocardiography, resulting in a study population of 58 patients. Relative clot composition was calculated using orbit image analysis to define the ratio of main histologic components (fibrin/platelets (F/P), red blood cell count (RBC), leukocytes). Cryptogenic stroke patients with PFO (PFO+, n = 20) displayed a significantly higher percentage of RBC (0.57 ± 0.17; p = 0.002) and lower percentage of F/P (0.38 ± 0.15; p = 0.003) compared to patients without PFO (PFO-, n = 38) (RBC: 0.41 ± 0.21; F/P: 0.52 ± 0.20). In conclusion, histologic clot composition in cryptogenic stroke varies depending on the presence of a PFO. Our findings histologically support the concept that a PFO may be of pathophysiological relevance in cryptogenic ischemic stroke.

Glioblastoma Relapses Show Increased Markers of Vulnerability to Ferroptosis.

Background: Despite the availability of various therapy options and being a widely focused research area, the prognosis of glioblastoma (GBM) still remains very poor due to therapy resistance, genetic heterogeneity and a diffuse infiltration pattern. The recently described non-apoptotic form of cell death ferroptosis may, however, offer novel opportunities for targeted therapies. Hence, the aim of this study was to investigate the potential role of ferroptosis in GBM, including the impact of treatment on the expression of the two ferroptosis-associated players glutathione-peroxidase 4 (GPX4) and acyl-CoA-synthetase long-chain family number 4 (ACSL4). Furthermore, the change in expression of the recently identified ferroptosis suppressor protein 1 (FSP1) and aldehyde dehydrogenase (ALDH) 1A3 was investigated. Methods: Immunohistochemistry was performed on sample pairs of primary and relapse GBM of 24 patients who had received standard adjuvant treatment with radiochemotherapy. To identify cell types generally prone to undergo ferroptosis, co-stainings of ferroptosis susceptibility genes in combination with cell-type specific markers including glial fibrillary acidic protein (GFAP) for tumor cells and astrocytes, as well as the ionized calcium-binding adapter molecule 1 (Iba1) for microglial cells were performed, supplemented by double stains combining GPX4 and ACSL4. Results: While the expression of GPX4 decreased significantly during tumor relapse, ACSL4 showed a significant increase. These results were confirmed by analyses of data sets of the Cancer Genome Atlas. These profound changes indicate an increased susceptibility of relapsed tumors towards oxidative stress and associated ferroptosis, a cell death modality characterized by unrestrained lipid peroxidation. Moreover, ALDH1A3 and FSP1 expression also increased in the relapses with significant results for ALDH1A3, whereas for FSP1, statistical significance was not reached. Results obtained from double staining imply that ferroptosis occurs more likely in GBM tumor cells than in microglial cells. Conclusion: Our study implies that ferroptosis takes place in GBM tumor cells. Moreover, we show that recurrent tumors have a higher vulnerability to ferroptosis. These results affirm that utilizing ferroptosis processes might be a possible novel therapy option, especially in the situation of recurrent GBM.

The Impact of Postoperative Tumor Burden on Patients With Brain Metastases.

Background: Brain metastases were considered to be well-defined lesions, but recent research points to infiltrating behavior. Impact of postoperative residual tumor burden (RTB) and extent of resection are still not defined enough. Patients and Methods: Adult patients with surgery of brain metastases between April 2007 and January 2020 were analyzed. Early postoperative MRI (<72 h) was used to segment RTB. Survival analysis was performed and cutoff values for RTB were revealed. Separate (subgroup) analyses regarding postoperative radiotherapy, age, and histopathological entities were performed. Results: A total of 704 patients were included. Complete cytoreduction was achieved in 487/704 (69.2%) patients, median preoperative tumor burden was 12.4 cm3 (IQR 5.2-25.8 cm3), median RTB was 0.14 cm3 (IQR 0.0-2.05 cm3), and median postoperative tumor volume of the targeted BM was 0.0 cm3 (IQR 0.0-0.1 cm3). Median overall survival was 6 months (IQR 2-18). In multivariate analysis, preoperative KPSS (HR 0.981982, 95% CI, 0.9761-0.9873, p < 0.001), age (HR 1.012363; 95% CI, 1.0043-1.0205, p = 0.0026), and preoperative (HR 1.004906; 95% CI, 1.0003-1.0095, p = 0.00362) and postoperative tumor burden (HR 1.017983; 95% CI; 1.0058-1.0303, p = 0.0036) were significant. Maximally selected log rank statistics showed a significant cutoff for RTB of 1.78 cm3 (p = 0.0022) for all and 0.28 cm3 (p = 0.0047) for targeted metastasis and cutoff for the age of 67 years (p < 0.001). (Stereotactic) Radiotherapy had a significant impact on survival (p < 0.001). Conclusions: RTB is a strong predictor for survival. Maximal cytoreduction, as confirmed by postoperative MRI, should be achieved whenever possible, regardless of type of postoperative radiotherapy.

Quantifying Heterogeneity in Tumors: Proposing a New Method Utilizing Convolutional Neuronal Networks.

Heterogeneity is a hallmark of glioblastoma (GBM), the most common malignant brain tumor, and a key reason for the poor survival rate of patients. However, establishing a clinically applicable, cost-efficient tool to measure and quantify heterogeneity is challenging. We present a novel method in an ongoing study utilizing two convolutional neuronal networks (CNN). After digitizing tumor samples, the first CNN delimitates GBM from normal tissue, the second quantifies heterogeneity within the tumor. Since neuronal networks can detect and interpret underlying and hidden information within images and have the ability to incorporate different information sets (i.e. clinical data and mutational status), this approach might venture towards a next level of integrated diagnosis. It may be applicable to other tumors as well and lead to a more precision-based medicine.

Age-adjusted Charlson comorbidity index in recurrent glioblastoma: a new prognostic factor?

BACKGROUND: For recurrent glioblastoma (GB) patients, several therapy options have been established over the last years such as more aggressive surgery, re-irradiation or chemotherapy. Age and the Karnofsky Performance Status Scale (KPSS) are used to make decisions for these patients as these are established as prognostic factors in the initial diagnosis of GB. This study's aim was to evaluate preoperative patient comorbidities by using the age-adjusted Charlson Comorbidity Index (ACCI) as a prognostic factor for recurrent GB patients. METHODS: In this retrospective analysis we could include 123 patients with surgery for primary recurrence of GB from January 2007 until December 2016 (43 females, 80 males, mean age 57 years (range 21-80 years)). Preoperative age, sex, ACCI, KPSS and adjuvant treatment regimes were recorded for each patient. Extent of resection (EOR) was recorded as a complete/incomplete resection of the contrast-enhancing tumor part. RESULTS: Median overall survival (OS) was 9.0 months (95% CI 7.1-10.9 months) after first re-resection. Preoperative KPSS > 80% (P < 0.001) and EOR (P = 0.013) were associated with significantly improved survival in univariate analysis. Including these factors in multivariate analysis, preoperative KPSS < 80 (HR 2.002 [95% CI: 1.246-3.216], P = 0.004) and EOR are the only significant prognostic factor (HR 1.611 [95% CI: 1.036-2.505], P = 0.034). ACCI was not shown as a prognostic factor in univariate and multivariate analyses. CONCLUSION: For patients with surgery for recurrent glioblastoma, the ACCI does not add further information about patient's prognosis besides the well-established KPSS and extent of resection.

Socio-Organizational Impact of Confocal Laser Endomicroscopy in Neurosurgery and Neuropathology: Results from a Process Analysis and Expert Survey.

During brain tumor resection surgery, it is essential to determine the tumor borders as the extent of resection is important for post-operative patient survival. The current process of removing a tissue sample for frozen section analysis has several shortcomings that might be overcome by confocal laser endomicroscopy (CLE). CLE is a promising new technology enabling the digital in vivo visualization of tissue structures in near real-time. Research on the socio-organizational impact of introducing this new methodology to routine care in neurosurgery and neuropathology is scarce. We analyzed a potential clinical workflow employing CLE by comparing it to the current process. Additionally, a small expert survey was conducted to collect data on the opinion of clinical staff working with CLE. While CLE can contribute to a workload reduction for neuropathologists and enable a shorter process and a more efficient use of resources, the effort for neurosurgeons and surgery assistants might increase. Experts agree that CLE offers huge potential for better diagnosis and therapy but also see challenges, especially due to the current state of experimental use, including a risk for misinterpretations and the need for special training. Future studies will show whether CLE can become part of routine care.