Biology subjects

Poulsen, F. R.

Publications and source records attributed to Poulsen, F. R..

2 recordsLinked to original sources

bioRxiv2025

Isolation and characterization of synaptic structures from human neural organoids

Human neural organoids (NOs) provide a powerful platform for investigating synaptic development and dysfunction during early neurodevelopment. However, methodologies for isolating functional synaptic structures from these models remain limited. Here, we present a differential centrifugation protocol enabling the enrichment of growth cone particles (GCPs) and immature synaptosomes from air-liquid interface cerebral organoids (ALI-COs) at distinct developmental stages (day 90 and 150). Notably, the method avoids density gradients, requires minimal starting material while maintaining reproducibility across human and murine tissues. Quantitative proteomic profiling revealed significant enrichment of growth cone markers (e.g. GAP43) and classical synaptosomal proteins (e.g. PCLO, BSN, SYN1). Transmission electron microscopy (TEM) confirmed the presence of membrane-enclosed GCPs with fibrous content and mitochondria in day 90 isolates, and immature synaptosomes containing synaptic vesicles on day 150. Functional viability of both types of synaptic structures was demonstrated through KCl-induced depolarization, which triggered phosphorylation changes in growth cone proteins (GAP43, MARCKS, MARCKSL1), cytoskeletal regulators (DCLK1, SHTN1, MARK4, MAP1B) and protein kinases (CAMK2G, PRKCE) in day 90 GCPs, as well as classical synaptic vesicle cycle proteins (SYN1, DNM1, RPH3A) at day 150. Overall, this study establishes a centrifugation-based protocol for isolating growth cones and immature synapses from human organoids, capturing key stages of synaptic development and enabling scalable, patient-compatible models to study synaptic function and dysfunction in neurodevelopmental and neurodegenerative disorders.

neuroscience↗

bioRxiv2023

Insight into spatial intratumoral genomic evolution in glioblastoma

Glioblastoma undergoes a complex and dynamic evolution involving genetic and epigenetic changes. Understanding the mechanisms underlying this evolution is vital for the development of efficient therapeutic strategies. Although treatment resistance is associated with intratumoral heterogeneity in glioblastoma, it remains uncertain whether hypometabolic and hypermetabolic lesions observed through positron emission tomography (PET) imaging are influenced by spatial intratumoral genomic evolution. In this study, we precisely isolated autologous hypometabolic and hypermetabolic lesions from glioblastoma using advanced neurosurgical and brain tumor imaging technologies, followed by comprehensive whole-genome exome and transcriptome analyses. Our findings revealed that hypermetabolic lesions evolved from hypometabolic lesions, harbored shrewd focal amplifications and deletions, and exhibited a higher frequency of critical genomic alterations linked to increased aggressiveness, upregulated APOBEC3 and hypoxic genes, and downregulated putative tumor suppressors. This study highlights spatial genomic evolution with diagnostic implications and unveils the obstacles and possibilities that should be considered in the development of novel therapeutic strategies. Statement of significanceGlioblastoma is a multifaceted disease that is difficult to treat, and insights into the metabolic gradient observed in imaging and the underlying role of genomic evolution are lacking. This study is the first to investigate the molecular basis of hypermetabolic tumor lesions in glioblastoma using precise three-dimensional biopsy isolation, whole genome/exome, and mRNA sequencing. These findings have diagnostic significance, provide insights into therapeutic resistance, and shed light on the obstacles encountered by precision therapeutics for glioblastoma.

cancer biology↗