Comparing vibrissal morphology and infraorbital foramen area in pinnipeds

Pinniped vibrissae are well-adapted to sensing in an aquatic environment, by being morphologically diverse and more sensitive than those of terrestrial species. However, it is both challenging and time-consuming to measure vibrissal sensitivity in many species. In terrestrial species, the infraorbital foramen (IOF) area is associated with vibrissal sensitivity and increases with vibrissal number. While pinnipeds are thought to have large IOF areas, this has not yet been systematically measured before. We investigated vibrissal morphology, IOF area, and skull size in 16 species of pinniped and 12 terrestrial Carnivora species. Pinnipeds had significantly larger skulls and IOF areas, longer vibrissae, and fewer vibrissae than the other Carnivora species. IOF area and vibrissal number were correlated in Pinnipeds, just as they are in terrestrial mammals. However, despite pinnipeds having significantly fewer vibrissae than other Carnivora species, their IOF area was not smaller, which might be due to pinnipeds having vibrissae that are innervated more. We propose that investigating normalized IOF area per vibrissa will offer an alternative way to approximate gross individual vibrissal sensitivity in pinnipeds and other mammalian species. Our data show that many species of pinniped, and some species of felids, are likely to have strongly innervated individual vibrissae, since they have high values of normalized IOF area per vibrissa. We suggest that species that hunt moving prey items in the dark will have more sensitive and specialized vibrissae, especially as they have to integrate between individual vibrissal signals to calculate the direction of moving prey during hunting

Abstract 331: Mitochondrial transfer from astrocytes drives glioblastoma tumorigenicity

Abstract Mitochondrial transfer in the central nervous system occurs from astrocytes to neurons in stroke. Mitochondrial exchange has also been reported among tumor cells in glioblastoma (GBM), the most common primary brain tumor. However, the role of mitochondrial transfer from non-neoplastic cells in the surrounding microenvironment to GBM remains poorly understood. We hypothesized that mitochondrial transfer from these non-neoplastic to GBM cells supports tumor metabolism and growth. Using transgenic mice expressing fluorophore-tagged mitochondria, we found that ~50% of orthotopically-implanted mouse GBM cells acquire host mitochondria. Brain-resident cells, mainly astrocytes, but not infiltrating immune cells were the primary mitochondrial donors in vivo and in vitro. Mitochondrial transfer also occurred from immortalized human astrocytes to a broad array of patient-derived xenograft (PDX) models of GBM in vitro at rates of 15-35%. GBM cells that acquired mitochondria expressed higher levels of the ATP-synthase subunit ATP5A and produced more ATP, while metabolomics revealed upregulated amino acid metabolism in recipient cells. In vivo, mouse GBM cells that acquired mitochondria were more likely to be in G2/M proliferative cell cycle phases. We observed a similar effect in PDX that acquired astrocyte mitochondria from co-cultures in vitro. To mechanistically link increased proliferation specifically to mitochondrial transfer, we isolated astrocyte mitochondria by differential centrifugation and found that addition and uptake of cell-free mitochondria in human GBM cells recapitulated the increased proliferation. Using sorted mouse and human GBM cells with/without astrocyte mitochondrial acquisition, we further found that mitochondrial transfer promoted in vitro self-renewal and in vivo tumorigenicity, leading to significant reduction in survival and increased penetrance in orthotopic GBM models. Transfer in mouse and human systems was contact-dependent and was abrogated by physical separation of donor and recipient cells by transwell inserts. We visualized contact-dependent transfer across actin-based intercellular connections consistent with previously reported microtubes. We confirmed the critical role of actin and the actin-associated protein, growth-associated protein 43 (GAP43) in facilitating mitochondrial transfer by showing that pharmacologic inhibition and genetic knockdown (respectively) significantly decreased the rate of mitochondrial transfer. Taken together, mitochondrial transfer comprises a fundamental, protumorigenic mechanism of GBM, enhancing metabolic activity and driving tumor cell proliferation. Further elucidating the molecular machinery regulating astrocyte mitochondrial transfer and its downstream protumorigenic effects will lead to therapeutic opportunities targeting this understudied tumor microenvironment interaction. Citation Format: Dionysios C. Watson, Defne Bayik, Simon Storevik, Shannon S. Moreino, Samuel S. Sprowls, Gauravi Deshpande, Palavalasa Sravya, Costas A. Lyssiotis, Daniel R. Wahl, Hrvoje Miletic, Justin D. Lathia. Mitochondrial transfer from astrocytes drives glioblastoma tumorigenicity [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 331

Chemotherapeutic Activity of Pitavastatin in Vincristine Resistant B-Cell Acute Lymphoblastic Leukemia

B-cell acute lymphoblastic leukemia (ALL) is derived from an accumulation of malignant, immature B cells in the bone marrow and blood. Relapse due, in part, to the emergence of tumor cells that are resistant to front line standard chemotherapy is associated with poor patient outcomes. This challenge highlights the need for new treatment strategies to eliminate residual chemoresistant tumor cells. Based on the use of pitavastatin in acute myeloid leukemia (AML), we evaluated its efficacy in an REH ALL cell line derived to be resistant to vincristine. We found that pitavastatin inhibited the proliferation of both parental and vincristine-resistant REH tumor cells at an IC50 of 449 nM and 217 nM, respectively. Mitochondrial bioenergetic assays demonstrated that neither vincristine resistance nor pitavastatin treatment affected cellular oxidative phosphorylation, beta-oxidation, or glycolytic metabolism in ALL cells. In a co-culture model of ALL cells with bone marrow stromal cells, pitavastatin significantly decreased cell viability more robustly in the vincristine-resistant ALL cells compared with their parental controls. Subsequently, NSG mice were used to develop an in vivo model of B-cell ALL using both parental and vincristine-resistant ALL cells. Pitavastatin (10 mg/kg i.p.) significantly reduced the number of human CD45+ REH ALL cells in the bone marrow of mice after 4 weeks of treatment. Mechanistic studies showed that pitavastatin treatment in the vincristine-resistant cells led to apoptosis, with increased levels of cleaved PARP and protein-signaling changes for AMP-activated protein kinase/FoxO3a/Puma. Our data suggest the possible repurposing of pitavastatin as a chemotherapeutic agent in a model of vincristine-resistant B-cell ALL

Chemotherapeutic Activity of Pitavastatin in Vincristine Resistant B-Cell Acute Lymphoblastic Leukemia

B-cell acute lymphoblastic leukemia (ALL) is derived from an accumulation of malignant, immature B cells in the bone marrow and blood. Relapse due, in part, to the emergence of tumor cells that are resistant to front line standard chemotherapy is associated with poor patient outcomes. This challenge highlights the need for new treatment strategies to eliminate residual chemoresistant tumor cells. Based on the use of pitavastatin in acute myeloid leukemia (AML), we evaluated its efficacy in an REH ALL cell line derived to be resistant to vincristine. We found that pitavastatin inhibited the proliferation of both parental and vincristine-resistant REH tumor cells at an IC50 of 449 nM and 217 nM, respectively. Mitochondrial bioenergetic assays demonstrated that neither vincristine resistance nor pitavastatin treatment affected cellular oxidative phosphorylation, beta-oxidation, or glycolytic metabolism in ALL cells. In a co-culture model of ALL cells with bone marrow stromal cells, pitavastatin significantly decreased cell viability more robustly in the vincristine-resistant ALL cells compared with their parental controls. Subsequently, NSG mice were used to develop an in vivo model of B-cell ALL using both parental and vincristine-resistant ALL cells. Pitavastatin (10 mg/kg i.p.) significantly reduced the number of human CD45+ REH ALL cells in the bone marrow of mice after 4 weeks of treatment. Mechanistic studies showed that pitavastatin treatment in the vincristine-resistant cells led to apoptosis, with increased levels of cleaved PARP and protein-signaling changes for AMP-activated protein kinase/FoxO3a/Puma. Our data suggest the possible repurposing of pitavastatin as a chemotherapeutic agent in a model of vincristine-resistant B-cell ALL