Determinants of cerebral blood flow and arterial transit time in healthy older adults.

Cerebral blood flow (CBF) and arterial transit time (ATT), markers of brain vascular health, worsen with age. The primary aim of this cross-sectional study was to identify modifiable determinants of CBF and ATT in healthy older adults (n = 78, aged 60-81 years). Associations between cardiorespiratory fitness and CBF or ATT were of particular interest because the impact of cardiorespiratory fitness is not clear within existing literature. Secondly, this study assessed whether CBF or ATT relate to cognitive function in older adults. Multiple post-labelling delay pseudo-continuous arterial spin labelling estimated resting CBF and ATT in grey matter. Results from multiple linear regressions found higher BMI was associated with lower global CBF (ß = -0.35, P = 0.008) and a longer global ATT (ß = 0.30, P = 0.017), global ATT lengthened with increasing age (ß = 0.43, P = 0.004), and higher cardiorespiratory fitness was associated with longer ATT in parietal (ß = 0.44, P = 0.004) and occipital (ß = 0.45, P = 0.003) regions. Global or regional CBF or ATT were not associated with processing speed, working memory, or attention. In conclusion, preventing excessive weight gain may help attenuate age-related declines in brain vascular health. ATT may be more sensitive to age-related decline than CBF, and therefore useful for early detection and management of cerebrovascular impairment. Finally, cardiorespiratory fitness appears to have little effect on CBF but may induce longer ATT in specific regions.

Protocol: revolutionizing central nervous system tumour diagnosis in low- and middle-income countries: an innovative observational study on intraoperative smear and deep learning.

OBJECTIVE: The aim of this study is to assess the feasibility and implementation of a novel approach for intraoperative brain smears within the operating room, which is augmented with deep learning technology. Materials and methods: This study is designed as an observational to evaluate the feasibility and implementation of using an innovative approach to intraoperative brain smears within the operating room, augmented with deep learning technology. The study will be conducted at Aga Khan University Hospital in Karachi, Pakistan, from May 2024 to July 2026, with an estimated sample size of 258. A neurosurgical trainee, trained by the study neuropathologist, will prepare and examine the smears under a microscope in the operating room. The findings of the trainee will be documented and compared to routine intraoperative consultations (smear and/or frozen section) and final histopathology results obtained from the pathology department. Additionally, the study will incorporate artificial intelligence tools to assist with the interpretation of smear and a telepathology interface to enable consultation from an off-site neuropathologist. CONCLUSIONS: The results of this study will hold significant potential to revolutionise neurosurgery practices in lowand middle-income countries by introducing a cost-effective, efficient, and high-quality intraoperative consultation method to settings that currently lack the necessary infrastructure and expertise. The implementation of this innovative approach has the potential to improve patient outcomes and increase access to intraoperative diagnosis, thereby addressing a significant unmet need in LMICs.

Consensus guidelines for the management of vestibular schwannoma for lowand middle-income countries.

Vestibular Schwannoma (VS), previously known as acoustic neuroma, constitutes the majority of tumours found in the cerebellopontine angle (CPA). Most guidelines for managing CPA tumours have been developed by high-income countries (HICs). However, these guidelines often fall short in addressing the unique challenges encountered in low- and middle-income countries (LMICs), such as Pakistan. In LMICs, issues related to a limited healthcare workforce, inadequate infrastructure, and constrained financial resources hinder the effective implementation of these HIC-derived guidelines. Additionally, it has been observed that VS tends to present at a larger size in LMICs compared to HICs. Given that VS is the predominant type of CPA tumour and other types are covered under separate guidelines, this article aims to provide practical, contextspecific recommendations for the screening, diagnosis, and management of Vestibular Schwannoma in LMIC settings. Our focus is to bridge the gap in care strategies and adapt them to the resource constraints and clinical realities of LMICs.

Consensus guidelines for the management of pediatric medulloblastoma in low and middle-income countries.

The management of medulloblastoma, a pediatric brain tumor, has evolved significantly with the advent of genomic subgrouping, yet morbidity and mortality remain high in LMICs like Pakistan due to inadequate multidisciplinary care infrastructure. This paper aims to establish evidence-based guidelines tailored to the constraints of such countries. An expert panel comprising neuro-oncologists, neurosurgeons, radiologists, radiation oncologists, neuropathologists, and pediatricians collaborated to develop these guidelines, considering the specific challenges of pediatric brain tumor care in Pakistan. The recommendations cover various aspects of medulloblastoma treatment, including pre-surgical workup, neurosurgery, neuropathology, chemotherapy, radiation therapy, and supportive care. They offer both minimum required and additional optional protocols for more advanced centers, ensuring comprehensive patient management with attention to complications and complexities encountered in Pakistan. The paper's consensus guidelines strive for uniformity in healthcare delivery and address significant gaps in diagnosis, treatment, and follow-up of pediatric medulloblastoma patients.

Consensus guidelines for the management of adult high-grade gliomas for low and middle-income countries.

High-grade glioma (HGG), a formidable and often incurable disease, presents an even greater challenge in low- and middle-income countries (LMICs) where resources and medical expertise are scarce. This scarcity not only exacerbates the suffering of patients but also contributes to poorer clinical outcomes. Particularly in LMICs, the underrepresentation of the population in clinical trials and the additional hurdles posed by financial constraints underscore an urgent need for contextspecific management strategies. In response, we have rigorously evaluated recent guidelines from leading medical societies, adapting them to suit the specific needs and limitations of the local context in Pakistan. This effort, undertaken in collaboration with local physicians, aims to provide a comprehensive, standardised approach to diagnose, treat, and follow-up with HGG patients. By focussing on the best available clinical evidence and judicious use of limited resources, we strive to improve patient care and outcomes in these challenging settings.

Consensus guidelines for the management of primary central nervous system lymphoma for low and middle-income countries.

Primary lymphoma of the central nervous system (PCNSL) is a rare and aggressive form of extranodal non-Hodgkin lymphoma primarily involving the brain, spinal cord, cerebrospinal fluid, and eyes. The role of surgical intervention in PCNSL is currently limited to biopsy and decompression of critical structures if needed - extended resection is debated. Chemotherapy is the mainstay of treatment. In lower and middle-income countries (LMICs), issues like delayed diagnosis and resource constraints are widespread. These guidelines provide a framework for addressing PCNSL in LMICs, emphasizing the importance of early diagnosis, tailored treatment approaches, and ongoing patient monitoring to improve outcomes for this rare and aggressive disease.

Consensus guidelines for the management of adult low-grade gliomas for low and middle-income countries.

Low-grade gliomas (LGG) are brain tumors of glial cells origin. They are grade 1 and grade 2 tumors according to the WHO classification. Diagnosis of LGG is made through imaging, histopathological analysis, and use of molecular markers. Imaging alone does not establish the grade of the tumor and thus a histopathological examination of tissue is crucial in establishing the definite histopathological diagnosis. Clinical presentation varies according to the location and size of the tumor. Surgical resection is strongly recommended in LGG over observation to improve overall survival as surgery leads to greater benefit due to progression-free survival. Radiation has shown benefits in LGG patients in randomized controlled trials and chemotherapy with temozolomide has also shown good results. This paper covers the principles of low-grade gliomas management and summarizes the recommendations for the LMICs.

Consensus guidelines for the management of pineal region tumours for low- and middle-income countries.

Pineal region tumours are rare and mainly arise at a younger age. They can be categorized into various types: germ cell tumours (GCT), pineal parenchymal tumours (PPT), meningiomas, gliomas, pineoblastoma, pineal parenchymal tumours of intermediate differentiation, papillary tumours of the pineal region, and SMARCB1- mutant desmoplastic myxoid tumour. Within GCT, germinomas are the most prevalent, comprising the majority of tumours in this region, while nongerminomatous GCTs are also present. In rare instances, metastases from other sites may manifest. These tumours often lead to obstructive hydrocephalus and commonly exhibit symptoms related to mass effect, including headache, nausea, vomiting, and impaired gait stability. Different subtypes of pineal region tumours exhibit distinct radiological characteristics, thus imaging remains the primary diagnostic tool. Histologic diagnosis necessitates biopsy, unless in cases of germ cell tumours, particularly germinomas, which can be identified through elevated levels of tumour markers like alpha-fetoprotein (AFP) and human chorionic gonadotropin (HCG) in both cerebrospinal fluid (CSF) and serum. While benign tumours might be effectively treated with radical resection alone, malignant tumours demand additional chemotherapy and radiotherapy following surgical removal.

Consensus guidelines for the management of intracranial ependymoma for low- and middle-income countries.

This paper presents comprehensive consensus guidelines for the management of intracranial ependymoma, neoplasms arising from ependymal cells in the central nervous system's ventricular system, in low- and middleincome countries (LMICs). Acknowledging the distinct epidemiological patterns of ependymomas, notably their higher incidence in paediatric patients, and variable survival rates, these guidelines emphasize tailored management approaches for different age groups. An expert panel, comprising specialists in neuro-oncology, convened to address gaps in diagnosis and management within LMICs, considering the varying clinical presentation based on tumour size and location. Emphasizing surgical intervention as the cornerstone of treatment, the guidelines also address challenges such as intraoperative bleeding and tumour location impacting complete resection. The role of molecular subgrouping in stratifying treatment and predicting prognosis is highlighted, alongside a careful consideration of radiotherapy timing, dose, and volume based on risk factors. Chemotherapy's role, especially in paediatric cases, is explored. The paper synthesizes current research and expert opinions, including the need for standardisation, genetic testing, and exploration of less invasive treatment modalities, to address the unique healthcare infrastructure challenges in LMICs. The guidelines also emphasize multidisciplinary teams, aiming to bridge the care gap between high-income countries and LMICs, and improve survival rates and quality of life for patients with intracranial ependymoma. This article serves as a valuable resource for clinicians, researchers, and policymakers in Pakistan and beyond, facilitating the development of evidence-based strategies in diverse healthcare settings.

Consensus guidelines for the management of brain stem and diffuse midline glioma for low- and middle-income countries.

The understanding of brainstem gliomas and diffuse midline gliomas has significantly increased in the last decade. However, the management paradigm remains a dilemma. The critical location is the foremost factor dictating the outcome. Recent advancements in the field of neuro-oncology are pushing the boundaries of optimal care in the developed world nevertheless, the strategies in low- and middle-income countries (LMICs) need to be tailored according to the resources to improve outcome. The objective of these guidelines is to provide an algorithm-based management plan to cater challenges for healthcare providers in LMICs.

Consensus guidelines for the management of craniopharyngioma in low- and middle-income countries.

Craniopharyngiomas are benign, extra-axial epithelial tumours originating from the pituitary stalk, impacting areas such as the hypothalamus, optic chiasm, and various cranial nerves. These tumours present unique surgical challenges due to their proximity to critical neurovascular structures. Management typically involves maximal safe resection as the primary approach. However, in low- and middle-income countries (LMICs), factors like late presentation, higher risks of endocrine and visual complications, frequent recurrence, and potential for incomplete resection complicate treatment. These challenges are exacerbated by limited access to specialised expertise and surgical equipment, increasing the risk of damage during surgery compared to High- Income Countries. This manuscript outlines management guidelines tailored for LMICs, emphasizing that a combination of surgical resection and chemoradiation therapy, as advised by a neuro-oncology tumour board, often yields the best outcomes.

AB059. Molecular signatures for survival prediction in glioma: a prospective, real-world data analysis.

BACKGROUND: Glioma characterization and follow-up are underreported from low-and-middle-income country centers within the literature. With the recent emphasis on molecular markers for survival prediction, there is a need for robust data exploring molecular epidemiology in these countries. In Pakistan particularly, there is a significant gap in glioma outcomes reporting and survival analysis. METHODS: One hundred and sixty-five consecutive glioma patients were enrolled from 2019 onwards; histopathological and molecular analysis was performed on archived formalin-fixed paraffin-embedded (FFPE) blocks for isocitrate dehydrogenase (IDH), P53, α-thalassemia retardation X-linked (ATRX) and Ki-67 immunohistochemical (IHC) markers. Survival analysis was calculated using the Kaplan-Meier method; hazard ratios are reported through a multivariate Cox regression model. RESULTS: Fifty-seven (35%) histopathological diagnoses were revised according to the updated criteria; 30% (n=16) glioblastoma were converted to a new category on re-analysis. IDH wild type (IDH-WT) gliomas had a significantly worse overall survival (log-rank =0.002), with a 2-year survival rate of 60% for IDH-mutant (IDH-M) and 38% for IDH-WT. Significant survival differences were seen for the Ki-67 index (log-rank =0.001) and methylguanine methyltransferase (MGMT) promotor methylation [log-rank =0.027, 2-year survival rate: 100% (methylation detected), 33% (methylation not detected)]. On Cox proportional hazards regression, gross total resection (P<0.001), IDH mutation (P<0.001), and updated histopathological diagnosis (P<0.001) were significant predictors of survival, with good sensitivity and specificity as seen on receiver operating characteristic (ROC) analysis [area under the curve (AUC) =0.86]. CONCLUSIONS: In our cohort, the revised World Health Organization (WHO) classification shows significant implications on prognosis and implications for treatment. Although these markers are not commonly used in low-and-middle-income country centers, our results strongly support their greater implementation for improved prognostication and reclassification.

Non-granulomatous meningoencephalitis with Balamuthia mandrillaris mimicking a tumor: First confirmed case from Pakistan.

Background: Free-living amoebae rarely instigate intracranial infections that may resemble neoplastic conditions on imaging. Naegleria fowleri precipitates an acute, swiftly fatal meningoencephalitis, whereas Acanthamoeba and Balamuthia species typically manifest with a less aggressive onset but carry equally dire consequences. Case Description: The case describes a 33-year-old woman with subacute encephalitis caused by Balamuthia mandrillaris. She experienced 2 months of back pain, 1 month of headaches, and 2 weeks of vomiting without fever, recent travel, aquatic activities, or animal exposure. Brain magnetic resonance imaging revealed a sizable, heterogeneous enhancing mass in the right temporal and frontal lobes, accompanied by vasogenic edema and midline shift. Histopathology showed marked inflammation and damage to blood vessels with amoebic trophozoites present. The trophozoites displayed specific characteristics, leading to the diagnosis of amoebic meningoencephalitis. Polymerase chain reaction and Sanger sequencing confirmed B. mandrillaris infection while testing for N. fowleri and Acanthamoeba was negative. Despite antibiotic treatment, the patient's condition deteriorated rapidly, resulting in death within 2 weeks of presentation. Conclusion: This is the first confirmed case of B. mandrillaris central nervous system (CNS) infection from Pakistan. The incidence of this disease is expected to rise due to increasing temperatures due to climate change and the deteriorating quality of the water supply. Balamuthia meningoencephalitis should, therefore be on the differential for non-neoplastic CNS lesions. Furthermore, an atypical histopathologic picture, including the absence of granulomatous inflammation, needs to be recognized.

Pediatric neuropathology practice in a low- and middle-income country: capacity building through institutional twinning.

Background: Accurate and precise diagnosis is central to treating central nervous system (CNS) tumors, yet tissue diagnosis is often a neglected focus in low- and middle-income countries (LMICs). Since 2016, the WHO classification of CNS tumors has increasingly incorporated molecular biomarkers into the diagnosis of CNS tumors. While this shift to precision diagnostics promises a high degree of diagnostic accuracy and prognostic precision, it has also resulted in increasing divergence in diagnostic and management practices between LMICs and high-income countries (HICs). Pathologists and laboratory professionals in LMICs lack the proper training and tools to join the molecular diagnostic revolution. We describe the impact of a 7-year long twinning program between Canada and Pakistan on pathology services. Methods: During the study period, 141 challenging cases of pediatric CNS tumors initially diagnosed at Aga Khan University Hospital (AKUH), Karachi, were sent to the Hospital for Sick Children in Toronto, Canada (SickKids), for a second opinion. Each case received histologic review and often immunohistochemical staining and relevant molecular testing. A monthly multidisciplinary online tumor board (MDTB) was conducted to discuss the results with pathologists from both institutions in attendance. Results: Diagnostic discordance was seen in 30 cases. Expert review provided subclassification for 53 cases most notably for diffuse gliomas and medulloblastoma. Poorly differentiated tumors benefited the most from second review, mainly because of the resolving power of specialized immunohistochemical stains, NanoString, and targeted gene panel next-generation sequencing. Collaboration with expert neuropathologists led to validation of over half a dozen immunostains at AKUH facilitating diagnosis of CNS tumors. Conclusions: LMIC-HIC Institutional twinning provides much-needed training and mentorship to pathologists and can help in infrastructure development by adopting and validating new immunohistochemical stains. Persistent unresolved cases indicate that molecular techniques are indispensable in for diagnosis in a minority of cases. The development of affordable alternative molecular techniques may help with these histologically unresolved cases.

Immunohistochemical and Histopathological Characterization of Spina Bifida Defect Tissues Removed After Prenatal and Postnatal Surgical Repair.

Background: Myelomeningocele or spina bifida is an open neural tube defect that is characterized by protrusion of the meninges and the spinal cord through a deformity in the vertebral arch and spinous process. Myelomeningocele of post-natal tissue is well described; however, pre-natal tissue of this defect has no known previous histologic characterization. We compared the histology of different forms of pre-natal myelomeningocele and post-natal myelomeningocele tissue obtained via prenatal intrauterine and postnatal surgical repairs. Methods: Pre-and post-natal tissues from spina bifida repair surgeries were obtained from lipomyelomeningocele, myeloschisis, and myelomeningocele spina bifida defects. Tissue samples were processed for H&E and immunohistochemical staining (KRT14 and p63) to assess epidermal and dermal development. Results: Prenatal skin near the defect site develops with normal epidermal, dermal, and adnexal structures. Within the grossly cystic specimens, histology shows highly dense fibrous connective tissue with complete absence of a normal epidermal development with a lack of p63 and KRT14 expression. Conclusion: Tissues harvested from prenatal and postnatal spina bifida repair surgeries appear as normal skin near the defect site. However, cystic tissues consist of highly dense fibrous connective tissue with complete absence of normal epidermal development.

Accelarated immune ageing is associated with COVID-19 disease severity.

BACKGROUND: The striking increase in COVID-19 severity in older adults provides a clear example of immunesenescence, the age-related remodelling of the immune system. To better characterise the association between convalescent immunesenescence and acute disease severity, we determined the immune phenotype of COVID-19 survivors and non-infected controls. RESULTS: We performed detailed immune phenotyping of peripheral blood mononuclear cells isolated from 103 COVID-19 survivors 3-5 months post recovery who were classified as having had severe (n = 56; age 53.12 ± 11.30 years), moderate (n = 32; age 52.28 ± 11.43 years) or mild (n = 15; age 49.67 ± 7.30 years) disease and compared with age and sex-matched healthy adults (n = 59; age 50.49 ± 10.68 years). We assessed a broad range of immune cell phenotypes to generate a composite score, IMM-AGE, to determine the degree of immune senescence. We found increased immunesenescence features in severe COVID-19 survivors compared to controls including: a reduced frequency and number of naïve CD4 and CD8 T cells (p < 0.0001); increased frequency of EMRA CD4 (p < 0.003) and CD8 T cells (p < 0.001); a higher frequency (p < 0.0001) and absolute numbers (p < 0.001) of CD28-ve CD57+ve senescent CD4 and CD8 T cells; higher frequency (p < 0.003) and absolute numbers (p < 0.02) of PD-1 expressing exhausted CD8 T cells; a two-fold increase in Th17 polarisation (p < 0.0001); higher frequency of memory B cells (p < 0.001) and increased frequency (p < 0.0001) and numbers (p < 0.001) of CD57+ve senescent NK cells. As a result, the IMM-AGE score was significantly higher in severe COVID-19 survivors than in controls (p < 0.001). Few differences were seen for those with moderate disease and none for mild disease. Regression analysis revealed the only pre-existing variable influencing the IMM-AGE score was South Asian ethnicity ([Formula: see text] = 0.174, p = 0.043), with a major influence being disease severity ([Formula: see text] = 0.188, p = 0.01). CONCLUSIONS: Our analyses reveal a state of enhanced immune ageing in survivors of severe COVID-19 and suggest this could be related to SARS-Cov-2 infection. Our data support the rationale for trials of anti-immune ageing interventions for improving clinical outcomes in these patients with severe disease.

Histological and neuroimaging comparison of SMART syndrome versus focal neuronal gigantism.

Two of the rarest radiation-induced adverse effects are focal neuronal gigantism (FNG) and SMART syndrome (stroke-like migraine attacks after radiation therapy). Both conditions develop years, and sometimes decades, after receipt of therapeutic radiation to the brain. To date, there are only 3 previously reported cases of FNG, all of which describe cortical thickening, enlarged "hypertrophic" neurons, and neuronal cytological changes. No detailed studies exist of histological features of SMART or the comparison between FNG and SMART. In this study, we contrast histological and neuroimaging features of 3 FNG vs. 4 SMART cases, the latter diagnosed by a neuroradiologist, neurooncologist, and/or neurosurgeon. We confirm the cortical thickening, dyslamination, neuronal cytomegaly, and gliosis in FNG vs. cortical architectural preservation and normal neuronal cytology in SMART, although both showed gliosis, scattered neurons with cytoplasmic accumulation of tau and neurofibrillary protein and variable co-existence of other radiation-induced lesions. Both conditions lacked significant inflammation or consistent small vessel hyalinization throughout the entire resection specimen. The absence of pathognomonic histologic alterations in SMART cases suggests underlying vascular dysregulation. Despite differing histology, some overlap may exist in neuroimaging features. Molecular assessment conducted in 2 cases of FNG was negative for significant alterations including in the MAPK pathway.

Unveiling the potential application of intraoperative brain smear for brain tumor diagnosis in low-middle-income countries: A comprehensive systematic review.

Background: Immediate intraoperative histopathological examination of tumor tissue is indispensable for a neurosurgeon to track surgical resection. A brain smear is a simple, rapid, and cost-effective technique, particularly important in the diagnosis of brain tumors. The study aims to determine the effectiveness of intraoperative brain smear in the diagnosis of brain tumors in low- and middle-income countries (LMICs), while also evaluating its sensitivity, specificity, positive predictive value (PPV), negative predictive value (NPV), and overall accuracy. Methods: A comprehensive search of the literature was conducted using PubMed, Scopus, and Google Scholar. The retrieved articles were independently screened by two reviewers. The data was extracted, processed, and organized using Microsoft Excel. Results: A total of 59 out of 553 articles screened were included in the final analysis. The sensitivity and specificity of the intraoperative smear of brain tumors were found to be over 90% in most studies. The PPV was consistently above 90% in 11 studies, reaching 100% in one study and the NPV varied, ranging from 63% to 100%, and the accuracy was found to be >80% in most studies. One recurrent theme in the majority of the included studies was that an intraoperative brain smear is a cost-effective, quick, accessible, and accurate method of diagnosing brain tumors, requiring minimal training and infrastructure. Conclusion: Intraoperative brain smear is a simple, rapid, cost-effective, and highly sensitive diagnostic modality for brain tumors. It can be a viable and accessible alternative to more traditional methods such as frozen sections and can be incorporated into neurosurgical practice in LMICs as a reliable and efficient diagnostic tool.

Integration of single-nuclei RNA-sequencing, spatial transcriptomics and histochemistry defines the complex microenvironment of NF1-associated plexiform neurofibromas.

Plexiform neurofibroma (PN) is a leading cause of morbidity in children with the genetic condition Neurofibromatosis Type 1 (NF1), often disfiguring or threatening vital structures. During formation of PN, a complex tumor microenvironment (TME) develops, with recruitment of neoplastic and non-neoplastic cell types being critical for growth and progression. Due to the cohesive cellularity of PN, single-cell RNA-sequencing is difficult and may result in a loss of detection of critical cellular subpopulations. To bypass this barrier, we performed single-nuclei RNA-sequencing (snRNA-seq) on 8 frozen PN samples, and integrated this with spatial transcriptomics (ST) in 4 PN samples and immunohistochemistry to provide morphological context to transcriptomic data. SnRNA-seq analysis definitively charted the heterogeneous cellular subpopulations in the PN TME, with the predominant fraction being fibroblast subtypes. PN showed a remarkable amount of inter-sample homogeneity regarding cellular subpopulation proportions despite being resected from a variety of anatomical locations. ST analysis identified distinct cellular subpopulations which were annotated using snRNA-seq data and correlated with histological features. Schwann cell/fibroblast interactions were identified by receptor/ligand interaction analysis demonstrating a high probability of Neurexin 1/Neuroligin 1 (NRXN1/NLGN1) receptor-ligand cross-talk predicted between fibroblasts and non-myelinated Schwann cells (NM-SC) and subtypes, respectively. We observed aberrant expression of NRXN1 and NLGN1 in our PN snRNA-seq data compared to a normal mouse sciatic nerve single-cell RNA-seq dataset. This pathway has never been described in PN and may indicate a clear and direct communication pathway between putative NM-SC cells of origin and surrounding fibroblasts, potentially driving disease progression. SnRNA-seq integrated with spatial transcriptomics advances our understanding of the complex cellular heterogeneity of PN TME and identify potential novel communication pathways that may drive disease progression, a finding that could provide translational therapy options for patients with these devastating tumors of childhood and early adulthood.