Spatial architecture of high-grade glioma reveals tumor heterogeneity within distinct domains.

Background: High-grade gliomas (HGGs) are aggressive primary brain cancers with poor response to standard regimens, driven by immense heterogeneity. In isocitrate dehydrogenase (IDH) wild-type HGG (glioblastoma, GBM), increased intratumoral heterogeneity is associated with more aggressive disease. Methods: Spatial technologies can dissect complex heterogeneity within the tumor ecosystem by preserving cellular organization in situ. We employed GeoMx digital spatial profiling, CosMx spatial molecular imaging, Xenium in situ mapping and Visium spatial gene expression in experimental and validation patient cohorts to interrogate the transcriptional landscape in HGG. Results: Here, we construct a high-resolution molecular map of heterogeneity in GBM and IDH-mutant patient samples to investigate the cellular communities that compose HGG. We uncovered striking diversity in the tumor landscape and degree of spatial heterogeneity within the cellular composition of the tumors. The immune distribution was diverse between samples, however, consistently correlated spatially with distinct tumor cell phenotypes, validated across tumor cohorts. Reconstructing the tumor architecture revealed two distinct niches, one composed of tumor cells that most closely resemble normal glial cells, associated with microglia, and the other niche populated by monocytes and mesenchymal tumor cells. Conclusions: This primary study reveals high levels of intratumoral heterogeneity in HGGs, associated with a diverse immune landscape within spatially localized regions.

Persistence of improved glucose homeostasis in Gclm null mice with age and cadmium treatment.

Antioxidant signaling/communication is among the most important cellular defense and survival pathways, and the importance of redox signaling and homeostasis in aging has been well-documented. Intracellular levels of glutathione (GSH), a very important endogenous antioxidant, both govern and are governed by the Nrf2 pathway through expression of genes involved in its biosynthesis, including the subunits of the rate-limiting enzyme (glutamate cysteine ligase, GCL) in GSH production, GCLC and GCLM. Mice homozygous null for the Gclm gene are severely deficient in GSH compared to wild-type controls, expressing approximately 10% of normal GSH levels. To compensate for GSH deficiency, Gclm null mice have upregulated redox-regulated genes, and, surprisingly, are less susceptible to certain types of oxidative damage. Furthermore, young Gclm null mice display an interesting lean phenotype, resistance to high fat diet-induced diabetes and obesity, improved insulin and glucose tolerance, and decreased expression of genes involved in lipogenesis. However, the persistence of this phenotype has not been investigated into old age, which is important in light of studies which suggest aging attenuates antioxidant signaling, particularly in response to exogenous stimuli. In this work, we addressed whether aging compromises the favorable phenotype of increased antioxidant activity and improved glucose homeostasis observed in younger Gclm null mice. We present data showing that under basal conditions and in response to cadmium exposure (2 mg/kg, dosed once via intraperitoneal injection), the phenotype previously described in young (<6 months) Gclm null mice persists into old age (24+ months). We also provide evidence that transcriptional activation of the Nrf2, AMPK, and PPARγ pathways underlie the favorable metabolic phenotype observed previously in young Gclm null mice.

Genetic determinants of susceptibility to silver nanoparticle-induced acute lung inflammation in mice.

Silver nanoparticles (AgNPs) are employed in a variety of consumer products; however, in vivo rodent studies indicate that AgNPs can cause lung inflammation and toxicity in a strain- and particle type-dependent manner, but mechanisms of susceptibility remain unclear. The aim of this study was to assess the variation in AgNP-induced lung inflammation and toxicity across multiple inbred mouse strains and to use genome-wide association (GWA) mapping to identify potential candidate susceptibility genes. Mice received doses of 0.25 mg/kg of either 20-nm citrate-coated AgNPs or citrate buffer using oropharyngeal aspiration. Neutrophils in bronchoalveolar lavage fluid (BALF) served as markers of inflammation. We found significant strain- and treatment-dependent variation in neutrophils in BALF. GWA mapping identified 10 significant single-nucleotide polymorphisms (false discovery rate, 15%) in 4 quantitative trait loci on mouse chromosomes 1, 4, 15, and 18, and Nedd4l (neural precursor cell expressed developmentally downregulated gene 4-like; chromosome 18), Ano6 (anocatmin 6; chromosome 15), and Rnf220 (Ring finger protein 220; chromosome 4) were considered candidate genes. Quantitative RT-PCR revealed significant inverse associations between mRNA levels of these genes and neutrophil influx. Nedd4l, Ano6, and Rnf220 are candidate susceptibility genes for AgNP-induced lung inflammation that warrant additional exploration in future studies.-Scoville, D. K., Botta, D., Galdanes, K., Schmuck, S. C., White, C. C., Stapleton, P. L., Bammler, T. K., MacDonald, J. W., Altemeier, W. A., Hernandez, M., Kleeberger, S. R., Chen, L.-C., Gordon, T., Kavanagh, T. J. Genetic determinants of susceptibility to silver nanoparticle-induced acute lung inflammation in mice.

The pulmonary inflammatory response to multiwalled carbon nanotubes is influenced by gender and glutathione synthesis.

Inhalation of multiwalled carbon nanotubes (MWCNTs) during their manufacture or incorporation into various commercial products may cause lung inflammation, fibrosis, and oxidative stress in exposed workers. Some workers may be more susceptible to these effects because of differences in their ability to synthesize the major antioxidant and immune system modulator glutathione (GSH). Accordingly, in this study we examined the influence of GSH synthesis and gender on MWCNT-induced lung inflammation in C57BL/6 mice. GSH synthesis was impaired through genetic manipulation of Gclm, the modifier subunit of glutamate cysteine ligase, the rate-limiting enzyme in GSH synthesis. Twenty-four hours after aspirating 25µg of MWCNTs, all male mice developed neutrophilia in their lungs, regardless of Gclm genotype. However, female mice with moderate (Gclm heterozygous) and severe (Gclm null) GSH deficiencies developed significantly less neutrophilia. We found no indications of MWCNT-induced oxidative stress as reflected in the GSH content of lung tissue and epithelial lining fluid, 3-nitrotyrosine formation, or altered mRNA or protein expression of several redox-responsive enzymes. Our results indicate that GSH-deficient female mice are rendered uniquely susceptible to an attenuated neutrophil response. If the same effects occur in humans, GSH-deficient women manufacturing MWCNTs may be at greater risk for impaired neutrophil-dependent clearance of MWCNTs from the lung. In contrast, men may have effective neutrophil-dependent clearance, but may be at risk for lung neutrophilia regardless of their GSH levels.

Susceptibility to quantum dot induced lung inflammation differs widely among the Collaborative Cross founder mouse strains.

Quantum dots (QDs) are engineered semiconductor nanoparticles with unique physicochemical properties that make them potentially useful in clinical, research and industrial settings. However, a growing body of evidence indicates that like other engineered nanomaterials, QDs have the potential to be respiratory hazards, especially in the context of the manufacture of QDs and products containing them, as well as exposures to consumers using these products. The overall goal of this study was to investigate the role of mouse strain in determining susceptibility to QD-induced pulmonary inflammation and toxicity. Male mice from 8 genetically diverse inbred strains (the Collaborative Cross founder strains) were exposed to CdSe-ZnS core-shell QDs stabilized with an amphiphilic polymer. QD treatment resulted in significant increases in the percentage of neutrophils and levels of cytokines present in bronchoalveolar lavage fluid (BALF) obtained from NOD/ShiLtJ and NZO/HlLtJ mice relative to their saline (Sal) treated controls. Cadmium measurements in lung tissue indicated strain-dependent differences in disposition of QDs in the lung. Total glutathione levels in lung tissue were significantly correlated with percent neutrophils in BALF as well as with lung tissue Cd levels. Our findings indicate that QD-induced acute lung inflammation is mouse strain dependent, that it is heritable, and that the choice of mouse strain is an important consideration in planning QD toxicity studies. These data also suggest that formal genetic analyses using additional strains or recombinant inbred strains from these mice could be useful for discovering potential QD-induced inflammation susceptibility loci.

In vivo approaches to assessing the toxicity of quantum dots.

The small size and heavy metal composition of quantum dots (QDs) combined with their growing consumer product and biomedical research applications have generated concern over their safety. In an occupational setting where QD-enabled products are being manufactured, inhalation is a likely route of exposure. Since current research indicates that QDs could cause inflammation and toxicity in the respiratory tract, it is important that a variety of methods be available to further characterize this potential respiratory hazard. This chapter focuses primarily on in vivo methods for modeling the inhalation and assessing the pulmonary toxicity of QDs.