Integrative spatial analysis reveals a multi-layered organization of glioblastoma.

Glioma contains malignant cells in diverse states. Here, we combine spatial transcriptomics, spatial proteomics, and computational approaches to define glioma cellular states and uncover their organization. We find three prominent modes of organization. First, gliomas are composed of small local environments, each typically enriched with one major cellular state. Second, specific pairs of states preferentially reside in proximity across multiple scales. This pairing of states is consistent across tumors. Third, these pairwise interactions collectively define a global architecture composed of five layers. Hypoxia appears to drive the layers, as it is associated with a long-range organization that includes all cancer cell states. Accordingly, tumor regions distant from any hypoxic/necrotic foci and tumors that lack hypoxia such as low-grade IDH-mutant glioma are less organized. In summary, we provide a conceptual framework for the organization of cellular states in glioma, highlighting hypoxia as a long-range tissue organizer.

Multi-sensory Gamma Stimulation Ameliorates Alzheimer's-Associated Pathology and Improves Cognition.

We previously reported that inducing gamma oscillations with a non-invasive light flicker (gamma entrainment using sensory stimulus or GENUS) impacted pathology in the visual cortex of Alzheimer's disease mouse models. Here, we designed auditory tone stimulation that drove gamma frequency neural activity in auditory cortex (AC) and hippocampal CA1. Seven days of auditory GENUS improved spatial and recognition memory and reduced amyloid in AC and hippocampus of 5XFAD mice. Changes in activation responses were evident in microglia, astrocytes, and vasculature. Auditory GENUS also reduced phosphorylated tau in the P301S tauopathy model. Furthermore, combined auditory and visual GENUS, but not either alone, produced microglial-clustering responses, and decreased amyloid in medial prefrontal cortex. Whole brain analysis using SHIELD revealed widespread reduction of amyloid plaques throughout neocortex after multi-sensory GENUS. Thus, GENUS can be achieved through multiple sensory modalities with wide-ranging effects across multiple brain areas to improve cognitive function.

A metastasis map of human cancer cell lines.

Most deaths from cancer are explained by metastasis, and yet large-scale metastasis research has been impractical owing to the complexity of in vivo models. Here we introduce an in vivo barcoding strategy that is capable of determining the metastatic potential of human cancer cell lines in mouse xenografts at scale. We validated the robustness, scalability and reproducibility of the method and applied it to 500 cell lines1,2 spanning 21 types of solid tumour. We created a first-generation metastasis map (MetMap) that reveals organ-specific patterns of metastasis, enabling these patterns to be associated with clinical and genomic features. We demonstrate the utility of MetMap by investigating the molecular basis of breast cancers capable of metastasizing to the brain-a principal cause of death in patients with this type of cancer. Breast cancers capable of metastasizing to the brain showed evidence of altered lipid metabolism. Perturbation of lipid metabolism in these cells curbed brain metastasis development, suggesting a therapeutic strategy to combat the disease and demonstrating the utility of MetMap as a resource to support metastasis research.

Glioblastoma-cortical organoids recapitulate cell state heterogeneity and intercellular transfer.

Glioblastoma is characterized by heterogeneous malignant cells that are functionally integrated within the neuroglial microenvironment. Here, we model this ecosystem by growing glioblastoma into long-term cultured human cortical organoids that contain the major neuroglial cell types found in the cerebral cortex. Single-cell RNA-seq analysis suggests that, compared to matched gliomasphere models, glioblastoma cortical organoids (GCO) more faithfully recapitulate the diversity and expression programs of malignant cell states found in patient tumors. Additionally, we observe widespread transfer of glioblastoma transcripts and GFP proteins to non-malignant cells in the organoids. Mechanistically, this transfer involves extracellular vesicles and is biased towards defined glioblastoma cell states and astroglia cell types. These results extend previous glioblastoma-organoid modeling efforts and suggest widespread intercellular transfer in the glioblastoma neuroglial microenvironment.

Multiplexed and scalable super-resolution imaging of three-dimensional protein localization in size-adjustable tissues.

The biology of multicellular organisms is coordinated across multiple size scales, from the subnanoscale of molecules to the macroscale, tissue-wide interconnectivity of cell populations. Here we introduce a method for super-resolution imaging of the multiscale organization of intact tissues. The method, called magnified analysis of the proteome (MAP), linearly expands entire organs fourfold while preserving their overall architecture and three-dimensional proteome organization. MAP is based on the observation that preventing crosslinking within and between endogenous proteins during hydrogel-tissue hybridization allows for natural expansion upon protein denaturation and dissociation. The expanded tissue preserves its protein content, its fine subcellular details, and its organ-scale intercellular connectivity. We use off-the-shelf antibodies for multiple rounds of immunolabeling and imaging of a tissue's magnified proteome, and our experiments demonstrate a success rate of 82% (100/122 antibodies tested). We show that specimen size can be reversibly modulated to image both inter-regional connections and fine synaptic architectures in the mouse brain.