Conditioned Medium From Reactive Astrocytes Inhibits Proliferation, Resistance, and Migration of p53-Mutant Glioblastoma Spheroid Through GLI-1 Downregulation.

Glioblastoma (GBM) aggressiveness is partly driven by the reactivation of signaling pathways such as Sonic hedgehog (SHH) and the interaction with its microenvironment. SHH pathway activation is one of the phenomena behind the glial transformation in response to tumor growth. The reactivation of the SHH signaling cascade during GBM-astrocyte interaction is highly relevant to understanding the mechanisms used by the tumor to modulate the adjacent stroma. The role of reactive astrocytes considering SHH signaling during GBM progression is investigated using a 3D in vitro model. T98G GBM spheroids displayed significant downregulation of SHH (61.4 ± 9.3%), GLI-1 (6.5 ± 3.7%), Ki-67 (33.7 ± 8.1%), and mutant MTp53 (21.3 ± 10.6%) compared to the CONTROL group when incubated with conditioned medium of reactive astrocytes (CM-AST). The SHH pathway inhibitor, GANT-61, significantly reduced previous markers (SHH = 43.0 ± 12.1%; GLI-1 = 9.5 ± 3.4%; Ki-67 = 31.9 ± 4.6%; MTp53 = 6.5 ± 7.5%) compared to the CONTROL, and a synergistic effect could be observed between GANT-61 and CM-AST. The volume (2.0 ± 0.2 × 107 µm³), cell viability (80.4 ± 3.2%), and migration (41 ± 10%) of GBM spheroids were significantly reduced in the presence of GANT-61 and CM-AST when compared to CM-AST after 72 h (volume = 2.3 ± 0.4 × 107 µm³; viability = 92.2 ± 6.5%; migration = 102.5 ± 14.6%). Results demonstrated that factors released by reactive astrocytes promoted a neuroprotective effect preventing GBM progression using a 3D in vitro model potentiated by SHH pathway inhibition.

GANT-61 Induces Autophagy and Apoptosis in Glioblastoma Cells despite their heterogeneity.

Glioblastoma (GBM) is the most common adult primary tumor of the CNS characterized by rapid growth and diffuse invasiveness into the brain parenchyma. The GBM resistance to chemotherapeutic drugs may be due to the presence of cancer stem cells (CSCs). The CSCs activate the same molecular pathways as healthy stem cells such as WNT, Sonic hedgehog (SHH), and Notch. Mutations or deregulations of those pathways play a key role in the proliferation and differentiation of their surrounding environment, leading to tumorigenesis. Here we investigated the effect of SHH signaling pathway inhibition in human GBM cells by using GANT-61, considering stem cell phenotype, cell proliferation, and cell death. Our results demonstrated that GANT-61 induces apoptosis and autophagy in GBM cells, by increasing the expression of LC3 II and cleaved caspase 3 and 9. Moreover, we observed that SHH signaling plays a crucial role in CSC phenotype maintenance, being also involved in the epithelial-mesenchymal transition (EMT) phenotype. We also noted that SHH pathway modulation can regulate cell proliferation as revealed through the analysis of Ki-67 and c-MYC expressions. We concluded that SHH signaling pathway inhibition may be a promising therapeutic approach to treat patients suffering from GBM refractory to traditional treatments.

Neuromechanisms of SARS-CoV-2: A Review.

Recent studies have suggested the neuroinvasive potential of severe acute respiratory coronavirus 2 (SARS-CoV-2). Notably, neuroinvasiveness might be involved in the pathophysiology of coronavirus disease 2019 (COVID-19). Some studies have demonstrated that synapse-connected routes may enable coronaviruses to access the central nervous system (CNS). However, evidence related to the presence of SARS-CoV-2 in the CNS, its direct impact on the CNS, and the contribution to symptoms suffered, remain sparse. Here, we review the current literature that indicates that SARS-CoV-2 can invade the nervous system. We also describe the neural circuits that are potentially affected by the virus and their possible role in the progress of COVID-19. In addition, we propose several strategies to understand, diagnose, and treat the neurological symptoms of COVID-19.

Cyclopamine sensitizes glioblastoma cells to temozolomide treatment through Sonic hedgehog pathway.

AIM: Glioblastoma is an extremely aggressive glioma, resistant to radio and chemotherapy usually performed with temozolomide. One of the main reasons for glioblastoma resistance to conventional therapies is due to the presence of cancer stem-like cells. These cells could recapitulate some signaling pathways important for embryonic development, such as Sonic hedgehog. Here, we investigated if the inhibitor of the Sonic hedgehog pathway, cyclopamine, could potentiate the temozolomide effect in cancer stem-like cells and glioblastoma cell lines in vitro. MAIN METHODS: The viability of glioblastoma cells exposed to cyclopamine and temozolomide treatment was evaluated by using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay while the induction of apoptosis was assessed by western blot. The stemness properties of glioma cells were verified by clonogenic and differentiation assay and the expression of stem cell markers were measured by fluorescence microscopy and western blot. KEY FINDINGS: The glioblastoma viability was reduced by cyclopamine treatment. Cyclopamine potentiated temozolomide treatment in glioblastoma cell lines by inducing apoptosis through activation of caspase-3 cleaved. Conversely, the combined treatment of cyclopamine and temozolomide potentiated the stemness properties of glioblastoma cells by inducing the expression of SOX-2 and OCT-4. SIGNIFICANCE: Cyclopamine plays an effect on glioblastoma cell lines but also sensibilize them to temozolomide treatment. Thus, first-line treatment with Sonic hedgehog inhibitor followed by temozolomide could be used as a new therapeutic strategy for glioblastoma patients.

Secondary glioblastoma metastasis outside the central nervous system in a young HIV-infected patient.

Glioblastoma is the most common adult primary brain tumor that occurs in the central nervous system and is characterized by rapid growth and diffuse invasiveness with respect to the adjacent brain parenchyma, which renders surgical resection inefficient. Although it is a highly infiltrative tumor, it is rarely disseminated beyond the central nervous system, wherein extracranial metastasis is a unique but rare manifestation of this kind of tumor. It is very common for acquired immunodeficiency syndrome (AIDS) patients to be infected with the human immunodeficiency virus (HIV), which suggests that a possible association between HIV infection and tumor development exists. In this paper, we present a new case of a young patient's HIV-associated glioblastoma, with glioblastoma metastasis within the T9 vertebral body and lymph nodes in the anterior neck tissue. Initially, the patient was diagnosed with a grade III plastic astrocytoma. The patient lived a normal life for a year while being treated with temozolomide, radiotherapy, and highly active antiretroviral therapy. However, the tumor quickly evolved into a glioblastoma. We believe that the drastic progression of the tumor from a grade III anaplastic astrocytoma to a metastatic glioblastoma is due to the HIV infection that the patient had acquired, which contributed to a weakened immune system, thus accelerating progression of the cancer.

A highlight on Sonic hedgehog pathway.

Hedgehog (Hh) signaling pathway plays an essential role during vertebrate embryonic development and tumorigenesis. It is already known that Sonic hedgehog (Shh) pathway is important for the evolution of radio and chemo-resistance of several types of tumors. Most of the brain tumors are resistant to chemotherapeutic drugs, consequently, they have a poor prognosis. So, a better knowledge of the Shh pathway opens an opportunity for targeted therapies against brain tumors considering a multi-factorial molecular overview. Therefore, emerging studies are being conducted in order to find new inhibitors for Shh signaling pathway, which could be safely used in clinical trials. Shh can signal through a canonical and non-canonical way, and it also has important points of interaction with other pathways during brain tumorigenesis. So, a better knowledge of Shh signaling pathway opens an avenue of possibilities for the treatment of not only for brain tumors but also for other types of cancers. In this review, we will also highlight some clinical trials that use the Shh pathway as a target for treating brain cancer.

LPA-primed astrocytes induce axonal outgrowth of cortical progenitors by activating PKA signaling pathways and modulating extracellular matrix proteins.

Lysophosphatidic acid (LPA) is one of the main membrane-derived lysophospholipids, inducing diverse cellular responses like cell proliferation, cell death inhibition, and cytoskeletal rearrangement, and thus is important in many biological processes. In the central nervous system (CNS), post-mitotic neurons release LPA extracellularly whereas astrocytes do not. Astrocytes play a key role in brain development and pathology, producing various cytokines, chemokines, growth factors, and extracellular matrix (ECM) components that act as molecular coordinators of neuron-glia communication. However, many molecular mechanisms underlying these events remain unclear-in particular, how the multifaceted interplay between the signaling pathways regulated by lysophospholipids is integrated in the complex nature of the CNS. Previously we showed that LPA-primed astrocytes induce neuronal commitment by activating LPA1-LPA2 receptors. Further, we revealed that these events were mediated by modulation and organization of laminin levels by astrocytes, through the induction of the epidermal growth factor receptor (EGFR) signaling pathway and the activation of the mitogen-activated protein (MAP) kinase (MAPK) cascade in response to LPA (Spohr et al., 2008, 2011). In the present work, we aimed to answer whether LPA affects astrocytic production and rearrangement of fibronectin, and to investigate the mechanisms involved in neuronal differentiation and maturation of cortical neurons induced by LPA-primed astrocytes. We show that PKA activation is required for LPA-primed astrocytes to induce neurite outgrowth and neuronal maturation and to rearrange and enhance the production of fibronectin and laminin. We propose a potential mechanism by which neurons and astrocytes communicate, as well as how such interactions drive cellular events such as neurite outgrowth, cell fate commitment, and maturation.

Gliomas and the vascular fragility of the blood brain barrier.

Astrocytes, members of the glial family, interact through the exchange of soluble factors or by directly contacting neurons and other brain cells, such as microglia and endothelial cells. Astrocytic projections interact with vessels and act as additional elements of the Blood Brain Barrier (BBB). By mechanisms not fully understood, astrocytes can undergo oncogenic transformation and give rise to gliomas. The tumors take advantage of the BBB to ensure survival and continuous growth. A glioma can develop into a very aggressive tumor, the glioblastoma (GBM), characterized by a highly heterogeneous cell population (including tumor stem cells), extensive proliferation and migration. Nevertheless, gliomas can also give rise to slow growing tumors and in both cases, the afflux of blood, via BBB is crucial. Glioma cells migrate to different regions of the brain guided by the extension of blood vessels, colonizing the healthy adjacent tissue. In the clinical context, GBM can lead to tumor-derived seizures, which represent a challenge to patients and clinicians, since drugs used for its treatment must be able to cross the BBB. Uncontrolled and fast growth also leads to the disruption of the chimeric and fragile vessels in the tumor mass resulting in peritumoral edema. Although hormonal therapy is currently used to control the edema, it is not always efficient. In this review we comment the points cited above, considering the importance of the BBB and the concerns that arise when this barrier is affected.

Neuron-astroglial interactions in cell-fate commitment and maturation in the central nervous system.

Neuron-astroglia interactions play a key role in several events of brain development, such as neuronal generation, migration, survival, and differentiation; axonal growth; and synapse formation and function. While there is compelling evidence of the effects of astrocyte factors on neurons, their effects on astrocytes have not been fully determined. In this review, we will focus on the role of neurons in astrocyte generation and maturation. Further, we highlight the great heterogeneity and diversity of astroglial and neural progenitors such as radial glia cells, and discuss the importance of the variety of cellular interactions in controlling the structural and functional organization of the brain. Finally, we present recent data on a new role of astrocytes in neuronal maturation, as mediators of the action of biolipids in the cerebral cortex. We will argue that the functional architecture of the brain depends on an intimate neuron-glia partnership, by briefly discussing the emerging view of how neuron-astrocyte dysfunctions might be associated with neurodegenerative diseases and neurological disorders.

Sphingosine 1-phosphate-primed astrocytes enhance differentiation of neuronal progenitor cells.

Sphingosine 1-phosphate (S1P) is a bioactive signaling lysophospholipid. Effects of S1P on proliferation, survival, migration, and differentiation have already been described; however, its role as a mediator of interactions between neurons and glial cells has been poorly explored. Here we describe effects of S1P, via the activation of its receptors in astrocytes, on the differentiation of neural progenitor cells (NPC) derived from either embryonic stem cells or the developing cerebral cortex. S1P added directly to NPC induced their differentiation, but S1P-primed astrocytes were able to promote even more pronounced changes in maturation, neurite outgrowth, and arborization in NPC. An increase in laminin by astrocytes was observed after S1P treatment. The effects of S1P-primed astrocytes on neural precursor cells were abrogated by antibodies against laminin. Together, our data indicate that S1P-treated astrocytes are able to induce neuronal differentiation of NPC by increasing the levels of laminin. These results implicate S1P signaling pathways as new targets for understanding neuroglial interactions within the central nervous system.