Craniofacial studies in chicken embryos confirm the pathogenicity of human FZD2 variants associated with Robinow syndrome.

Robinow syndrome is a rare disease caused by variants of seven WNT pathway genes. Craniofacial features include widening of the nasal bridge and jaw hypoplasia. We used the chicken embryo to test whether two missense human FZD2 variants (1301G>T, p.Gly434Val; 425C>T, p.Pro142Lys) were sufficient to change frontonasal mass development. In vivo, the overexpression of retroviruses with wild-type or variant human FZD2 inhibited upper beak ossification. In primary cultures, wild-type and variant human FZD2 significantly inhibited chondrogenesis, with the 425C>T variant significantly decreasing activity of a SOX9 luciferase reporter compared to that for the wild type or 1301G>T. Both variants also increased nuclear shuttling of ß-catenin (CTNNB1) and increased the expression of TWIST1, which are inhibitory to chondrogenesis. In canonical WNT luciferase assays using frontonasal mass cells, the variants had dominant-negative effects on wild-type FZD2. In non-canonical assays, the 425C>T variant failed to activate the reporter above control levels and was unresponsive to exogenous WNT5A. This is the first single amino acid change to selectively alter ligand binding in a FZD receptor. Therefore, FZD2 missense variants are pathogenic and could lead to the altered craniofacial morphogenesis seen in Robinow syndrome.

BRAF V600E in a preclinical model of pleomorphic Xanthoastrocytoma: Analysis of the tumor microenvironment and immune cell infiltration dynamics in vivo.

Low-grade glioma (LGG) is the most common brain tumor affecting pediatric patients (pLGG) and BRAF mutations constitute the most frequent genetic alterations. Within the spectrum of pLGGs, approximately 70%-80% of pediatric patients diagnosed with transforming pleomorphic xanthoastrocytoma (PXA) harbor the BRAF V600E mutation. However, the impact of glioma BRAF V600E cell regulation of tumor-infiltrating immune cells and their contribution to tumor progression remains unclear. Moreover, the efficacy of BRAF inhibitors in treating pLGGs is limited compared with their impact on BRAF-mutated melanoma. Here we report a novel immunocompetent RCAS-BRAF V600E murine glioma model. Pathological assessment indicates this model seems to be consistent with diffuse gliomas and morphological features of PXA. Our investigations revealed distinct immune cell signatures associated with increased trafficking and activation within the tumor microenvironment (TME). Intriguingly, immune system activation within the TME also generated a pronounced inflammatory response associated with dysfunctional CD8+ T cells, increased presence of immunosuppressive myeloid cells and regulatory T cells. Further, our data suggests tumor-induced inflammatory processes, such as cytokine storm. These findings suggest a complex interplay between tumor progression and the robust inflammatory response within the TME in preclinical BRAF V600E LGGs, which may significantly influence animal survival.

Meningioma: current updates on genetics, classification, and mouse modeling.

Meningiomas, the most common primary brain tumors in adults, are often benign and curable by surgical resection. However, a subset is of higher grade, shows aggressive growth behavior as well as brain invasion, and often recurs even after several rounds of surgery. Increasing evidence suggests that tumor classification and grading primarily based on histopathology do not always accurately predict tumor aggressiveness and recurrence behavior. The underlying biology of aggressive treatment-resistant meningiomas and the impact of specific genetic aberrations present in these high-grade tumors is still only insufficiently understood. Therefore, an in-depth research into the biology of this tumor type is warranted. More recent studies based on large-scale molecular data such as whole exome/genome sequencing, DNA methylation sequencing, and RNA sequencing have provided new insights into the biology of meningiomas and have revealed new risk factors and prognostic subtypes. The most common genetic aberration in meningiomas is functional loss of NF2 and occurs in both low- and high-grade meningiomas, whereas NF2-wildtype meningiomas are enriched for recurrent mutations in TRAF7, KLF4, AKT1, PI3KCA, and SMO and are more frequently benign. Most meningioma mouse models are based on patient-derived xenografts and only recently have new genetically engineered mouse models of meningioma been developed that will aid in the systematic evaluation of specific mutations found in meningioma and their impact on tumor behavior. In this article, we review recent advances in the understanding of meningioma biology and classification and highlight the most common genetic mutations, as well as discuss new genetically engineered mouse models of meningioma.

Haploinsufficiency of phosphodiesterase 10A activates PI3K/AKT signaling independent of PTEN to induce an aggressive glioma phenotype.

Glioblastoma is universally fatal and characterized by frequent chromosomal copy number alterations harboring oncogenes and tumor suppressors. In this study, we analyzed exome-wide human glioblastoma copy number data and found that cytoband 6q27 is an independent poor prognostic marker in multiple data sets. We then combined CRISPR-Cas9 data, human spatial transcriptomic data, and human and mouse RNA sequencing data to nominate PDE10A as a potential haploinsufficient tumor suppressor in the 6q27 region. Mouse glioblastoma modeling using the RCAS/tv-a system confirmed that Pde10a suppression induced an aggressive glioma phenotype in vivo and resistance to temozolomide and radiation therapy in vitro. Cell culture analysis showed that decreased Pde10a expression led to increased PI3K/AKT signaling in a Pten-independent manner, a response blocked by selective PI3K inhibitors. Single-nucleus RNA sequencing from our mouse gliomas in vivo, in combination with cell culture validation, further showed that Pde10a suppression was associated with a proneural-to-mesenchymal transition that exhibited increased cell adhesion and decreased cell migration. Our results indicate that glioblastoma patients harboring PDE10A loss have worse outcomes and potentially increased sensitivity to PI3K inhibition.

Tumor associated microglia/macrophages utilize GPNMB to promote tumor growth and alter immune cell infiltration in glioma.

Tumor-associated microglia and blood-derived macrophages (TAMs) play a central role in modulating the immune suppressive microenvironment in glioma. Here, we show that GPNMB is predominantly expressed by TAMs in human glioblastoma multiforme and the murine RCAS-PDGFb high grade glioma model. Loss of GPNMB in the in vivo tumor microenvironment results in significantly smaller tumor volumes and generates a pro-inflammatory innate and adaptive immune cell microenvironment. The impact of host-derived GPNMB on tumor growth was confirmed in two distinct murine glioma cell lines in organotypic brain slices from GPNMB-KO and control mice. Using published data bases of human glioma, the elevated levels in TAMs could be confirmed and the GPNMB expression correlated with a poorer survival.

A comparative life cycle assessment on recycled concrete aggregates modified by accelerated carbonation treatment and traditional methods.

Recycling of construction and demolition wastes contributes to achieve carbon summit and carbon neutrality early in the construction industry. Accelerated carbonation is a promising new technology for enhancing the properties of recycled concrete aggregates (RCAs) as well as mitigating global warming. This study performed a comparative life cycle assessment on RCAs modified by accelerated carbonation treatment and traditional methods. The effect of different treatment methods on environmental impacts of concrete was evaluated. The key contributors of environmental impacts for concrete incorporating carbonated RCAs were identified. Moreover, a sensitivity analysis on the transport distance of concrete incorporating carbonated RCAs was conducted. Results demonstrated that incorporating carbonated RCAs could significantly reduce the energy demand, environmental impacts and environmental cost compared with natural aggregate concrete. Accelerated carbonation treatment exhibited greater potential than the normal two-stage crushing and heating treatment in mitigating environmental burden, especially for the global warming potential. Cement production and transportation were the primary contributors to environmental impacts of concrete incorporating carbonated RCAs. Sensitivity analysis indicated incorporating carbonated RCAs as alternatives of natural aggregates contributes to lower the environmental impacts of concrete when the natural aggregates are far from urban areas while the recycling center is near the city.

Tumor-targeted delivery of lnc antisense RNA against RCAS1 by live-attenuated tryptophan-auxotrophic Salmonella inhibited 4T1 breast tumors and metastasis in mice.

Emerging chemo- and radiotherapy resistance exacerbated the cancer risk and necessitated novel treatment strategies. Although RNA therapeutics against pro-oncogenic genes are highly effective, tumor-specific delivery remains a barrier to the implementation of this valuable tool. In this study, we report a tryptophan-auxotrophic Salmonella typhimurium strain as an onco-therapeutic delivery system with tumor-targeting ability using 4T1 mice breast-cancer model. The receptor-binding cancer antigen expressed on SiSo cell (RCAS1) is a cancer-specific protein that induces the apoptosis of peripheral lymphocytes and confers tumor immune evasion. We designed a long non-coding antisense-RNA against RCAS1 (asRCAS1) and delivered by Salmonella using a non-antibiotic, auxotrophic-selective, eukaryotic expression plasmid, pJHL204. After in vivo tumor-to-tumor passaging, the JOL2888 (ΔtrpA, ΔtrpE, Δasd + asRCAS1) strain exhibited high sustainability in tumors, but did not last in healthy organs, thereby demonstrating tumor specificity and safety. RCAS1 inhibition in the tumor was confirmed by western blotting and qPCR. In mice, JOL2888 treatment reduced tumor-associated macrophages, improved the T cell population, elicited cell-mediated immunity, and suppressed cancer-promoting genes. Consequently, the JOL2888 treatment significantly decreased the tumor volume by 80%, decreased splenomegaly by 30%, and completely arrested lung metastasis. These findings highlight the intrinsic tumor-targeting ability of tryptophan-auxotrophic Salmonella for delivering onco-therapeutic macromolecules.

Investigating the effects of dermal exposure to in-vivo animal models on the riot-control properties of a powder formulation of Tragia involucrata leaf hair lining.

Purpose: Riot control agents (RCAs) such as CS, CN, CR, PAVA, and OC, etc., are already in use and has produced numerous health risks, including skin burns, dermatitis, gastrointestinal issues, impairment of respiratory variables, conjunctivitis, etc., and even prolonged and repeated exposure may cause death. Therefore, there is a demand and need for non-lethal, non-toxic RCAs that can effectively control riots without resulting in fatal outcomes. This study was carried out to evaluate the health risks related to a novel formulation made from isolated Tragia involucrata leaf hair lining, that can be used as the best suitable non-lethal RCAs.Methods: According to the OECD guidelines, studies on acute dermal toxicity, dermal irritation/corrosion, and skin sensitisation were carried out. Wistar rats were used in an acute dermal toxicity study, and the results indicated no mortality, morbidity, or abnormal food-and-water intake, biochemical parameters, or histopathological examination findings. A study on dermal irritation in Rabbits produced moderate erythema and the effect was instantaneous and resolved within 72 hrs of post-exposure. A skin sensitisation test was conducted on Guinea pig.Results: The results showed that the formulation had moderate skin-sensitizing properties after the application of the challenge dose. Patchy erythema was seen, and it went away 30 hrs after the gauze patch was removed.Conclusion: The preclinical results did not produce any indication of severe toxicity which supports it to be used as a natural RCAs in the future.

Genetically modified IL2 bone-marrow-derived myeloid cells reprogram the glioma immunosuppressive tumor microenvironment.

Gliomas are one of the leading causes of cancer-related death in the adolescent and young adult (AYA) population. Two-thirds of AYA glioma patients are affected by low-grade gliomas (LGGs), but there are no specific treatments. Malignant progression is supported by the immunosuppressive stromal component of the tumor microenvironment (TME) exacerbated by M2 macrophages and a paucity of cytotoxic T cells. A single intravenous dose of engineered bone-marrow-derived myeloid cells that release interleukin-2 (GEMys-IL2) was used to treat mice with LGGs. Our results demonstrate that GEMys-IL2 crossed the blood-brain barrier, infiltrated the TME, and reprogrammed the immune cell composition and transcriptome. Moreover, GEMys-IL2 extended survival in an LGG immunocompetent mouse model. Here, we report the efficacy of an in vivo approach that demonstrates the potential for a cell-mediated innate immunotherapy designed to enhance the recruitment of activated effector T and natural killer cells within the glioma TME.

Chicken HOXC8 and HOXC10 genes may play a role in the altered skull morphology associated with the Crest phenotype.

One of the most intriguing traits found in domestic chickens is the Crest phenotype. This trait, characterized by a tuft of elongated feathers sprouted from the head, is found in breeds such as Polish chickens and Silkie chickens. Moreover, some crested chicken breeds also exhibit a protuberance in their anterodorsal skull region. Previous studies have strived to identify the causative factors of this trait. This study aimed to elucidate the role of chicken HOXC8 and HOXC10 in the formation of the Crest phenotype. We explored the effect of ectopic expression of HOXC8 or HOXC10 on the chicken craniofacial morphology using the RCAS retrovirus transformation system. Microcomputed tomography scanning was conducted to measure the 3D structure of the cranial bone of transgenic embryos for geometric morphometric analysis. We found that the ectopic expression of HOXC8 or HOXC10 in chicken heads caused mild morphological changes in the skull compared with the GFP-transgenic control group. Geometric morphometric analysis showed that HOXC8 and HOXC10 transgenic groups expressed a mild upward shape change in the frontal region of the skull compared with the control group, which is similar to what is seen in the crested chicken breeds. In conclusion, this study supports findings in previous studies in which HOX genes play a role in the formation of the altered skull morphology related to the Crest phenotype. It also supports that mutations in HOX genes may contribute to intra- and inter-specific variation in morphological traits in vertebrates.

Novel genetically engineered H3.3G34R model reveals cooperation with ATRX loss in upregulation of Hoxa cluster genes and promotion of neuronal lineage.

Background: Pediatric high-grade gliomas (pHGGs) are aggressive pediatric CNS tumors and an important subset are characterized by mutations in H3F3A, the gene that encodes Histone H3.3 (H3.3). Substitution of Glycine at position 34 of H3.3 with either Arginine or Valine (H3.3G34R/V), was recently described and characterized in a large cohort of pHGG samples as occurring in 5-20% of pHGGs. Attempts to study the mechanism of H3.3G34R have proven difficult due to the lack of knowledge regarding the cell-of-origin and the requirement for co-occurring mutations for model development. We sought to develop a biologically relevant animal model of pHGG to probe the downstream effects of the H3.3G34R mutation in the context of vital co-occurring mutations. Methods: We developed a genetically engineered mouse model (GEMM) that incorporates PDGF-A activation, TP53 loss and the H3.3G34R mutation both in the presence and loss of Alpha thalassemia/mental retardation syndrome X-linked (ATRX), which is commonly mutated in H3.3G34 mutant pHGGs. Results: We demonstrated that ATRX loss significantly increases tumor latency in the absence of H3.3G34R and inhibits ependymal differentiation in the presence of H3.3G34R. Transcriptomic analysis revealed that ATRX loss in the context of H3.3G34R upregulates Hoxa cluster genes. We also found that the H3.3G34R overexpression leads to enrichment of neuronal markers but only in the context of ATRX loss. Conclusions: This study proposes a mechanism in which ATRX loss is the major contributor to many key transcriptomic changes in H3.3G34R pHGGs. Accession number: GSE197988.

Novel cancer gene discovery using a forward genetic screen in RCAS-PDGFB-driven gliomas.

BACKGROUND: Malignant gliomas, the most common malignant brain tumors in adults, represent a heterogeneous group of diseases with poor prognosis. Retroviruses can cause permanent genetic alterations that modify genes close to the viral integration site. METHODS: Here we describe the use of a high-throughput pipeline coupled to the commonly used tissue-specific retroviral RCAS-TVA mouse tumor model system. Utilizing next-generation sequencing, we show that retroviral integration sites can be reproducibly detected in malignant stem cell lines generated from RCAS-PDGFB-driven glioma biopsies. RESULTS: A large fraction of common integration sites contained genes that have been dysregulated or misexpressed in glioma. Others overlapped with loci identified in previous glioma-related forward genetic screens, but several novel putative cancer-causing genes were also found. Integrating retroviral tagging and clinical data, Ppfibp1 was highlighted as a frequently tagged novel glioma-causing gene. Retroviral integrations into the locus resulted in Ppfibp1 upregulation, and Ppfibp1-tagged cells generated tumors with shorter latency on orthotopic transplantation. In human gliomas, increased PPFIBP1 expression was significantly linked to poor prognosis and PDGF treatment resistance. CONCLUSIONS: Altogether, the current study has demonstrated a novel approach to tagging glioma genes via forward genetics, validating previous results, and identifying PPFIBP1 as a putative oncogene in gliomagenesis.

Multiparametric Longitudinal Profiling of RCAS-tva-Induced PDGFB-Driven Experimental Glioma.

Glioblastomas are incurable primary brain tumors harboring a heterogeneous landscape of genetic and metabolic alterations. Longitudinal imaging by MRI and [18F]FET-PET measurements enable us to visualize the features of evolving tumors in a dynamic manner. Yet, close-meshed longitudinal imaging time points for characterizing temporal and spatial metabolic alterations during tumor evolution in patients is not feasible because patients usually present with already established tumors. The replication-competent avian sarcoma-leukosis virus (RCAS)/tumor virus receptor-A (tva) system is a powerful preclinical glioma model offering a high grade of spatial and temporal control of somatic gene delivery in vivo. Consequently, here, we aimed at using MRI and [18F]FET-PET to identify typical neuroimaging characteristics of the platelet-derived growth factor B (PDGFB)-driven glioma model using the RCAS-tva system. Our study showed that this preclinical glioma model displays MRI and [18F]FET-PET features that highly resemble the corresponding established human disease, emphasizing the high translational relevance of this experimental model. Furthermore, our investigations unravel exponential growth dynamics and a model-specific tumor microenvironment, as assessed by histology and immunochemistry. Taken together, our study provides further insights into this preclinical model and advocates for the imaging-stratified design of preclinical therapeutic interventions.

Expression of CD44v6 and RCAS1 in Uterine Cervical Carcinoma Infected with Human Papillomavirus and Its Effect on Cell Proliferation and Differentiation.

OBJECTIVE: To investigate the expression of CD44v6 and RCAS1 and the presence of HPV in cervical cancer tissues, to determine serum RCAS1 levels, and to evaluate these components in correlation with clinicopathologic features and survival. METHODS: A total of 52 patients consisting of 28 squamous cell carcinoma (SCC) and 24 adenocarcinoma cases, were studied. RCAS1 and CD44v6 expression was evaluated using immunohistochemical staining. HPV 16 and 18 E6 genes were detected using PCR, and serum RCAS1 concentrations were measured using ELISA. Associations between these factors and clinicopathologic features and survival were analyzed. RESULTS: CD44v6 expression was significantly higher in SCC compared with that in adenocarcinoma (P<0.001). It also showed a significant relation to histologic grade (P<0.001) and tumor size (P=0.03). RCAS1 expression was higher in adenocarcinoma than in SCC (P=0.001), and it showed a borderline relation with histological grade (P=0.057). Overall survival was not significantly different in both CD44v6 and RCAS1 expression; however, FIGO stage (P=0.025) and tumor size (P=0.042) resulted statistically different. The pre-surgical treatment serum RCAS1 levels were not associated with any clinicopathological variables. The presence of HPV 16 E6 was higher in SCC, while the presence of HPV 18 E6 was higher in adenocarcinoma (P<0.001). Detection of HPV 16 E6 was significantly associated with expression of CD44v6. The presence of HPV both HPV 16 E6 and HPV 18 E6 was found in cancer tissues with RCAS1 expression, but without any statistical significance. CONCLUSION: CD44v6 and RCAS1 expression seems to be involved in tumor proliferation and differentiation, but it is not implicated in the progression and invasion of cervical cancer infected by HPV. Pre-treatment levels of serum RCAS1 in cervical cancer are not a diagnostic and predictive biomarker.

Functional Analysis of Rare Genetic Variants in the Negative Regulator of Intracellular Calcium Signaling RCAS/SLC10A7.

The solute carrier family 10 member SLC10A7 is a negative regulator of intracellular calcium signaling (RCAS). In cell culture, SLC10A7 expression is negatively correlated with store-operated calcium entry (SOCE) via the plasma membrane. SLC10A7-deficient cells have significantly increased calcium influx after treatment with thapsigargin for depletion of ER calcium stores, whereas SLC10A7/RCAS overexpression limits calcium influx. Genetic variants in the human SLC10A7 gene are associated with skeletal dysplasia and amelogenesis imperfecta and reveal loss of function on cellular calcium influx. More recently, an additional disease-related genetic variant (P303L) as well as some novel genetic variants (V235F, T221M, I136M, L210F, P285L, and G146S) have been identified. In the present study, these variants were expressed in HEK293 cells to study their subcellular localization and their effect on cellular calcium influx. All variants were properly sorted to the ER compartment and closely co-localized with the STIM protein, a functional component of SOCE. The variants P303L and L210F showed significantly reduced effects on cellular calcium influx compared to the wild type but still maintained some degree of residual activity. This might explain the milder phenotype of patients bearing the P303L variant and might indicate disease potential for the newly identified L210F variant. In contrast, all other variants behaved like the wild type. In conclusion, the occurrence of variants in the SLC10A7 gene should be considered in patients with skeletal dysplasia and amelogenesis imperfecta. In addition to the already established variants, the present study identifies another potential disease-related SLC10A7/RCAS variant, namely, L210F, which seems to be most frequent in South Asian populations.

Overexpression of OsERF83, a Vascular Tissue-Specific Transcription Factor Gene, Confers Drought Tolerance in Rice.

Abiotic stresses severely affect plant growth and productivity. To cope with abiotic stresses, plants have evolved tolerance mechanisms that are tightly regulated by reprogramming transcription factors (TFs). APETALA2/ethylene-responsive factor (AP2/ERF) transcription factors are known to play an important role in various abiotic stresses. However, our understanding of the molecular mechanisms remains incomplete. In this study, we identified the role of OsERF83, a member of the AP2/ERF transcription factor family, in response to drought stress. OsERF83 is a transcription factor localized to the nucleus and induced in response to various abiotic stresses, such as drought and abscisic acid (ABA). Overexpression of OsERF83 in transgenic plants (OsERF83OX) significantly increased drought tolerance, with higher photochemical efficiency in rice. OsERF83OX was also associated with growth retardation, with reduced grain yields under normal growth conditions. OsERF83 is predominantly expressed in the vascular tissue of all organs. Transcriptome analysis revealed that OsERF83 regulates drought response genes, which are related to the transporter (OsNPF8.10, OsNPF8.17, OsLH1), lignin biosynthesis (OsLAC17, OsLAC10, CAD8D), terpenoid synthesis (OsTPS33, OsTPS14, OsTPS3), cytochrome P450 family (Oscyp71Z4, CYP76M10), and abiotic stress-related genes (OsSAP, OsLEA14, PCC13-62). OsERF83 also up-regulates biotic stress-associated genes, including PATHOGENESIS-RELATED PROTEIN (PR), WALL-ASSOCIATED KINASE (WAK), CELLULOSE SYNTHASE-LIKE PROTEIN E1 (CslE1), and LYSM RECEPTOR-LIKE KINASE (RLK) genes. Our results provide new insight into the multiple roles of OsERF83 in the cross-talk between abiotic and biotic stress signaling pathways.

CRISPR/Cas Technologies Applied to Pseudogenes.

Pseudogenes have been considered as nonfunctional copies of their parental genes for a long time. Indeed, they have been often defined "junk DNA" or "transcriptional noise." However, with the identification of their involvement in several biological processes, the necessity of their study is inevitably growing up. The manipulation of pseudogene expression is complicated by their high homology with parental genes and by the fact that most of them work at the transcriptional level as noncoding RNAs. With the advent of CRISPR/Cas technology, these problems can be overcome. Particularly, as we describe in this chapter, it is possible: To perform genome editing, obtaining the complete elimination of the pseudogene genomic sequence (knock-out), preventing pseudogene transcription, introducing specific mutations in the pseudogene sequence, or introducing a specific sequence (knock-in). To positively or negatively manipulate pseudogene transcription. To target pseudogene RNA and negatively regulate its expression. To edit pseudogene DNA and RNA and alter a specific sequence. Moreover, CRISPR/Cas technology can be used as an RNA Binding Protein system for molecular biology techniques (such as RNA immunoprecipitation and pull-down), as well as for transcript tracking and live imaging.

Elevated fibroblast growth factor-inducible 14 expression transforms proneural-like gliomas into more aggressive and lethal brain cancer.

High-grade gliomas (HGGs) are aggressive, treatment-resistant, and often fatal human brain cancers. The TNF-like weak inducer of apoptosis (TWEAK)/fibroblast growth factor-inducible 14 (Fn14) signaling axis is involved in tissue repair after injury and constitutive signaling has been implicated in the pathogenesis of numerous solid cancers. The Fn14 gene is expressed at low levels in the normal, uninjured brain but is highly expressed in primary isocitrate dehydrogenase wild-type and recurrent HGGs. Fn14 signaling is implicated in numerous aspects of glioma biology including brain invasion and chemotherapy resistance, but whether Fn14 overexpression can directly promote tumor malignancy has not been reported. Here, we used the replication-competent avian sarcoma-leukosis virus/tumor virus A system to examine the impact of Fn14 expression on glioma development and pathobiology. We found that the sole addition of Fn14 to an established oncogenic cocktail previously shown to generate proneural-like gliomas led to the development of highly invasive and lethal brain cancer with striking biological features including extensive pseudopalisading necrosis, constitutive canonical and noncanonical NF-κB pathway signaling, and high plasminogen activator inhibitor-1 (PAI-1) expression. Analyses of HGG patient datasets revealed that high human PAI-1 gene (SERPINE1) expression correlates with shorter patient survival, and that the SERPINE1 and Fn14 (TNFRSF12A) genes are frequently co-expressed in bulk tumor tissues, in tumor subregions, and in malignant cells residing in the tumor microenvironment. These findings provide new insights into the potential importance of Fn14 in human HGG pathobiology and designate both the NF-κB signaling node and PAI-1 as potential targets for therapeutic intervention. MAIN POINTS: This work demonstrates that elevated levels of the TWEAK receptor Fn14 in tumor-initiating, neural progenitor cells leads to the transformation of proneural-like gliomas into more aggressive and lethal tumors that exhibit constitutive NF-κB pathway activation and plasminogen activator inhibitor-1 overexpression.

Leveraging the replication-competent avian-like sarcoma virus/tumor virus receptor-A system for modeling human gliomas.

Gliomas are the most common primary intrinsic brain tumors occurring in adults. Of all malignant gliomas, glioblastoma (GBM) is considered the deadliest tumor type due to diffuse brain invasion, immune evasion, cellular, and molecular heterogeneity, and resistance to treatments resulting in high rates of recurrence. An extensive understanding of the genomic and microenvironmental landscape of gliomas gathered over the past decade has renewed interest in pursuing novel therapeutics, including immune checkpoint inhibitors, glioma-associated macrophage/microglia (GAMs) modulators, and others. In light of this, predictive animal models that closely recreate the conditions and findings found in human gliomas will serve an increasingly important role in identifying new, effective therapeutic strategies. Although numerous syngeneic, xenograft, and transgenic rodent models have been developed, few include the full complement of pathobiological features found in human tumors, and therefore few accurately predict bench-to-bedside success. This review provides an update on how genetically engineered rodent models based on the replication-competent avian-like sarcoma (RCAS) virus/tumor virus receptor-A (tv-a) system have been used to recapitulate key elements of human gliomas in an immunologically intact host microenvironment and highlights new approaches using this model system as a predictive tool for advancing translational glioma research.

Quantitative volumetric analysis of the Golgi apparatus following X-ray irradiation by super-resolution 3D-SIM microscopy.

To obtain quantitative volumetric data for the Golgi apparatus after ionizing radiation (IR) using super-resolution three-dimensional structured illumination (3D-SIM) microscopy. Normal human retinal pigment epithelial (RPE) cells were irradiated with X-rays (10 Gy), followed by immunofluorescence staining of the Golgi marker RCAS1. 3D-SIM imaging was performed using DeltaVision OMX version 4 and SoftWoRx 6.1. Polygon rendering and spot signal identification were performed using Imaris 8.1.2. Differences between groups were assessed by Welch's t test. RCAS1 signals in untreated cells were located adjacent to nuclei and showed a reticular morphology. Upon IR, the area of RCAS1 signals expanded while retaining the reticular morphology. Polygon rendering imaging revealed that the volume of RCAS1 at 48 h post-IR was greater than that for unirradiated cells (93.7 ± 19.0 µm3 vs. 33.0 ± 4.2 µm3, respectively; P < 0.001): a 2.8-fold increase. Spot signal imaging showed that the number of RCAS1 spot signals post-IR was greater than that for unirradiated cells [3.4 ± 0.8 (× 103) versus 1.3 ± 0.2 (× 103), respectively; P < 0.001]: a 2.7-fold increase. This is the first study to report quantitative volumetric data of the Golgi apparatus in response to IR using super-resolution 3D-SIM microscopy.