Biology subjects

Ravi, V. M.

Publications and source records attributed to Ravi, V. M..

8 recordsLinked to original sources

bioRxiv2026

Spatially confined niches support hypoxia-associated transcriptional plasticity contributing to malignant progression in IDH-mutant gliomas

BackgroundWhile hypoxia is a well-established driver of glioblastoma progression, its role in IDH-mutant gliomas, characterized by localized hypoxic microenvironments rather than overt necrosis, remains poorly understood. Here, we investigate how hypoxia and microenvironmental-adaptations shape cellular heterogeneity and transcriptional plasticity in these tumors. MethodsWe integrated bulk, single-cell, and spatial-transcriptomics datasets from IDH-mutant glioma patients (Astrocytomas and Oligodendrogliomas) to characterize cellular-states and map the localization of hypoxic niches. To uncover tumor microenvironment effects, we established co-culture models using primary IDH-mutant glioma cells with human microglia and astrocytes, maintained under hypoxic and normoxic conditions, followed by bulk RNA-sequencing. ResultsWe identified a hypoxia-associated astrocyte-like (AC-like) program that defines a quiescent, non-cycling population with a distinct transcription factor profile indicative of functional plasticity in IDH-mutant gliomas. These cells harbor glioma stem cell (GSC)-like features and are poised for a quiescent-to-activated (Q-to-A) transition that drives tumor progression. Mechanistically, co-culture models reveal that microglia promote this Q-to-A transition by enhancing HBEGF/EGFR paracrine signaling. Spatial transcriptomics uncovers the co-localization of hypoxic niches within quiescent AC-like cells, whereby the activated subpopulation forms discrete niches defined by localized HBEGF/EGFR communication gradients. Notably, tumors exhibiting elevated EGFR-driven activation signatures correlate with higher histological grade and poorer patient survival, implicating the Q-to-A transition as a critical driver of malignant progression. ConclusionQ-to-A transition within the hypoxic niche represents a critical driver of malignant progression in IDH-mutant gliomas, providing a microenvironment-driven mechanism for the transition to higher-grade disease and identifying targetable-vulnerabilities for therapeutic intervention. Key pointsO_LIHypoxic niches spatially confine quiescent, astrocyte-like cellular state characterized by glioma stem cell features in IDH-mutant gliomas. C_LIO_LIMicroglia triggers the quiescent-to-activated (Q-to-A) transition via paracrine EGFR signaling crosstalk. C_LIO_LIEGFR-driven Q-to-A plasticity serves as a microenvironmentally-regulated driver of malignant progression and adverse patient survival. C_LI Importance of the studyWhile high-grade glioblastomas feature well-defined hypoxic and necrotic regions that drive tumor progression and therapy resistance, IDH-mutant gliomas lack these distinct hallmarks, instead exhibiting disorganized hypoxic niches. Consequently, the functional contribution of these niches to tumor progression has remained poorly understood. Although genetic and epigenetic alterations are established drivers of progression in IDH-mutant gliomas, we provide an additional, essential layer of complexity: microenvironment-driven transcriptional plasticity. By uncovering the relationship between tumor hypoxia and heterogeneous glioma cell states, we uncover that these niches are essential for such plasticity. We also show that the tumor microenvironment provides critical molecular cues necessary to initiate a transcriptional shift enabling glioma cells to transition from a quiescent reservoir into an activated state primed for rapid proliferation. By identifying this paracrine mechanism, our work uncovers a targetable vulnerability that dictates how dormant cell reservoirs are mobilized to fuel malignant transformation.

cancer biology↗

bioRxiv2026

Spatial analysis reveals the evolving organization of low-grade and high-grade IDH-mutant glioma

Adult diffuse gliomas are comprised of malignant cell states interwoven with the non-malignant brain microenvironment. Here we combine spatial transcriptomics and spatial proteomics of IDH-mutant gliomas to define organizational principles across histological grades. In low-grade tumors, spatial organization arises from underlying nonmalignant brain structures. For example, we classify low-grade tumor regions as embedded into white matter and identify a sharp white-grey matter junction that restricts cortical invasion and is associated with marked changes in tumor composition and cellular phenotypes. This junction is preferentially traversed by oligodendrocyte progenitor (OPC)-like malignant cells, which may drive tumor expansion. In contrast, intermediate-grade tumors are largely disorganized, with few recurring pairwise interactions between cancer cell states and TME cell types. In high-grade tumors, hypoxia/necrosis-associated global structure begins to emerge, reminiscent of IDH-wildtype glioblastoma. Together, these findings reveal two independent axes of glioma spatial organization--from brain anatomy-driven organization in low-grade tumors to hypoxia-associated structure in high-grade tumors--and establishes a framework that links tumor grade to recurrent spatial associations between cell states and cell types.

cancer biology↗

bioRxiv2026

Hypoxia and Associated Acidosis Generate Cell-Type Specific Myeloid Responses in Glioblastoma

Hypoxia is a defining feature of glioblastoma (GBM), yet how it cooperates with hypoxia-associated acidosis to shape microglia and infiltrating monocyte-derived macrophages (MDM) remains poorly understood. We integrated cyclic immunohistochemistry, single-cell RNA sequencing, spatial transcriptomics, in vitro cell cultures, and DNA methylation profiling to outline hypoxia-driven responses in up to 136 GBMs. These hypoxic niches were selectively enriched for MDMs that activated carbonic anhydrase (CA) mediated pH buffering and other metabolic adaptation programs, enabling survival in acidic hypoxia, increasingly interacted with cancer cells, and show polarization toward immunosuppressive myeloid-derived suppressor cell (MDSC)-like states. In contrast, microglia were depleted in hypoxic areas, lacked compensatory CA isoenzymes, and developed TNF-linked stress responses and loss of homeostatic identity in acidic hypoxia. These findings identify metabolic adaptation to hypoxia-associated microenvironmental stress as a key determinant of GBM immune architecture, driving myeloid cell fates, spatial TME reorganization and the emergence of immunosuppressive tumor ecosystems.

cancer biology↗

bioRxiv2025

Iron-Associated Mesenchymal Plasticity and Tumor Invasion in Glioblastoma

Glioblastoma (GBM) recurrence is driven by tumor cells that infiltrate surrounding brain tissue and evade surgical and therapeutic eradication. Although dysregulated iron handling is a recognized feature of the necrotic and hemorrhagic GBM tumor core, its relationship to invasive tumor cell states remains incompletely defined. Here, we investigated how iron-associated microenvironments relate to transcriptional programs of invasion in human GBM. Using multi-regional single-nucleus RNA sequencing of human GBM specimens encompassing tumor core, invasive front, and infiltrated cortex, we found that malignant cells from the tumor core exhibit coordinated upregulation of iron uptake and storage pathways together with invasion-associated gene programs. At the single-cell level, iron metabolism and invasion signatures were strongly correlated, defining a distinct core-enriched malignant subpopulation with mesenchymal-like transcriptional identity, stress-adaptive features, and angiogenic signaling. These iron-high/invasion-high cells aligned with mesenchymal-and astrocyte-like GBM states and were associated with unfavorable patient survival. Despite elevated oxidative stress and ferroptosis-associated transcriptional pressure, this population concurrently expressed anti-apoptotic and anti-ferroptotic regulators, consistent with an iron-tolerant invasive state. To assess functional consequences of iron exposure, we modeled iron-rich conditions using non-cytotoxic particulate iron in patient-derived GBM cell lines and human organotypic cortical slice cultures. Iron exposure induced intracellular iron accumulation, oxidative stress responses, increased tumor cell motility in vitro, and enhanced invasion within intact human brain tissue. Collectively, these findings demonstrate that iron-rich tumor core niches are closely associated with mesenchymal plasticity and invasive behavior in GBM and support a role for iron-associated microenvironmental pressure in shaping invasive tumor cell states. Key PointsO_LITranscriptional profiling reveals that the GBM tumor core harbors malignant cells that strictly couple active iron metabolism with invasive programs. C_LIO_LIThis iron-accumulating, Mesenchymal-like subpopulation is distinctively characterized by angiogenesis and stress resistance. C_LIO_LIIron supplementation in vitro and human ex vivo models are sufficient to drive mesenchymal transition and significantly enhance tumor migration and tissue invasion. C_LI Importance of the StudyGlioblastoma (GBM) recurrence is driven by highly invasive tumor cells that evade resection and resist therapy. Yet the microenvironmental pressures that push GBM cells into an invasive, therapy-resistant state remain poorly defined. Although iron dysregulation has been implicated across cancers, its role as a microenvironmental determinant of GBM invasion has never been demonstrated in physiologically relevant human systems. By integrating multi-regional single-nucleus RNA sequencing with functional validation in patient-derived GBM lines and human organotypic cortical slice cultures, we uncover a core-enriched, iron-associated mesenchymal program that co-segregates with invasion, stress adaptation, and angiogenic signaling. We further show that physiologically relevant iron exposure is sufficient to induce mesenchymal transition, enhance motility, and accelerate tissue invasion within human cortical architecture. These findings position iron as a selective driver that links the hemorrhagic, necrotic GBM core to the emergence of invasive subpopulations that seed recurrence. The data identify iron handling and stress-response pathways as actionable therapeutic vulnerabilities, providing a foundation for strategies that target metabolic resilience and iron-dependent invasive states in GBM. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=157 SRC="FIGDIR/small/695920v1_ufig1.gif" ALT="Figure 1"> View larger version (35K): org.highwire.dtl.DTLVardef@141cc11org.highwire.dtl.DTLVardef@bcb055org.highwire.dtl.DTLVardef@1dcd332org.highwire.dtl.DTLVardef@ad7cad_HPS_FORMAT_FIGEXP M_FIG C_FIG

cancer biology↗

bioRxiv2021

Spatiotemporal heterogeneity of glioblastoma is dictated by microenvironmental interference

Glioblastomas are highly malignant tumors of the central nervous system. Evidence suggests that these tumors display large intra- and inter-patient heterogeneity hallmarked by subclonal diversity and dynamic adaptation amid developmental hierarchies1-3. However, the source for dynamic reorganization of cellular states within their spatial context remains elusive. Here, we in-depth characterized glioblastomas by spatially resolved transcriptomics, metabolomics and proteomics. By deciphering exclusive and shared transcriptional programs across patients, we inferred that glioblastomas develop along defined neural lineages and adapt to inflammatory or metabolic stimuli reminiscent of reactive transformation in mature astrocytes. Metabolic profiling and imaging mass cytometry supported the assumption that tumor heterogeneity is dictated by microenvironmental alterations. Analysis of copy number variation (CNV) revealed a spatially cohesive organization of subclones associated with reactive transcriptional programs, confirming that environmental stress gives rise to selection pressure. Deconvolution of age-dependent transcriptional programs in malignant and non-malignant specimens identified the aging environment as the major driver of inflammatory transformation in GBM, suggesting that tumor cells adopt transcriptional programs similar to inflammatory transformation in astrocytes. Glioblastoma stem cells implanted into human neocortical slices of varying age levels, independently confirmed that the ageing environment dynamically shapes the intratumoral heterogeneity towards reactive transcriptional programs. Our findings provide insights into the spatial architecture of glioblastoma, suggesting that both locally inherent tumor as well as global alterations of the tumor microenvironment shape its transcriptional heterogeneity. Global age-related inflammation in the human brain is driving distinct transcriptional transformation in glioblastomas, which requires an adjustment of the currently prevailing glioma models.

cancer biology↗

bioRxiv2020

Inferring spatially transient gene expression pattern from spatial transcriptomic studies

Spatial transcriptomic is a technology to provide deep transcriptomic profiling by preserving the spatial organization. Here, we present a framework for SPAtial Transcriptomic Analysis (SPATA, https://themilolab.github.io/SPATA), to provide a comprehensive characterization of spatially resolved gene expression, regional adaptation of transcriptional programs and transient dynamics along spatial trajectories.

bioinformatics↗

bioRxiv2020

Lineage and Spatial Mapping of Glioblastoma-associated Immunity

The diversity of molecular states and cellular plasticity of immune cells in the glioblastoma environment is still poorly understood. Here, we performed scRNA sequencing of the immune compartment and mapped potential cellular interactions leading to an immunosuppressive microenvironment and dysfunction of T cells. Through inferring the dynamic adaptation during T cell activation, we identified three different terminal states with unique transcriptional programs. Modeling of driver genes for terminal T cell fate identified IL-10 signaling alterations in a subpopulation of HAVCR2(+) T cells. To explore in depth cellular interactions, we established an in-silico model by the integration of spatial transcriptomic and scRNA-sequencing, and identified a subset of HMOX1+ myeloid cells defined by IL10 release leading to T cell exhaustion. We found a spatial overlap between HMOX(+) myeloid and HAVCR2(+) T cells, suggesting that myeloid-lymphoid interaction causes immunosuppression present in tumor regions with enriched mesenchymal gene expression. Using human neocortical GBM model, coupled with patient-derived T cells, we confirmed that the functional interaction between myeloid and lymphoid cells, leads to a dysfunctional state of T cells. This IL-10 driven T cell exhaustion was found to be rescued by JAK/STAT inhibition. A comprehensive understanding of the cellular states and plasticity of lymphoid cells in GBM will aid towards successful immunotherapeutic approaches.

cancer biology↗

bioRxiv2017

When a rose is not a rose: Pyramidal neurons respond differently in healthy and epileptic human neocortex

Epilepsy affects a huge number of patients by severe disruption of brain functions and is characterised by recurrent seizures, sometimes hard to be treated by medications. Seizure induced cellular consequences in ionic gradient and homeostasis are expected to result in electrophysiological differences between epileptic and non-epileptic neurons.\n\nIn the following work we demonstrate these differences in layer III cortical pyramidal neurons sourced from epileptic and non-epileptic human patient tissue. Although visually indistinguishable and featuring similar membrane potentials and latency to first spikes upon whole cell patch stimulation, epileptic pyramidal neurons display a larger rheobase and a smaller membrane resistance, responding less efficiently to electrical stimulation than their peri-tumorous equivalents. This decreased excitability contradicts results in comparable animal models of epilepsy and was further corroborated by detailed analysis of spiking characteristics and phase plot analysis of these events. Both point to an overexpression of K+ channels trying to compensate for the hyperexcited, epileptic network state. A computational model of a pyramidal neuron was utilized to give an estimate of the needful relative changes in K+ and Na+ conductances.

neuroscience↗