ABSTRACT
Glioma-initiating cells (GIC) are considered the underlying cause of recurrences of aggressive glioblastomas, replenishing the tumor population and undermining the efficacy of conventional chemotherapy. Here we report the discovery that inhibiting T-type voltage-gated Ca2+ and KCa channels can effectively induce selective cell death of GIC and increase host survival in an orthotopic mouse model of human glioma. At present, the precise cellular pathways affected by the drugs affecting these channels are unknown. However, using cell-based assays and integrated proteomics, phosphoproteomics, and transcriptomics analyses, we identified the downstream signaling events these drugs affect. Changes in plasma membrane depolarization and elevated intracellular Na+, which compromised Na+-dependent nutrient transport, were documented. Deficits in nutrient deficit acted in turn to trigger the unfolded protein response and the amino acid response, leading ultimately to nutrient starvation and GIC cell death. Our results suggest new therapeutic targets to attack aggressive gliomas. Cancer Res; 77(7); 1741-52. ©2017 AACR.
Subject(s)
Amino Acids/metabolism , Brain Neoplasms/drug therapy , Calcium Channel Blockers/pharmacology , Calcium Channels, T-Type/physiology , Glioma/drug therapy , Potassium Channels, Calcium-Activated/antagonists & inhibitors , Unfolded Protein Response/drug effects , Animals , Biological Transport , Brain Neoplasms/metabolism , Brain Neoplasms/pathology , Cell Death , Cell Line, Tumor , Dihydropyridines/pharmacology , Glioma/metabolism , Glioma/pathology , Humans , Mice , Mycotoxins/pharmacology , Neoplastic Stem Cells/pathology , Proteomics , Sodium/metabolismABSTRACT
Tissue-engineered tracheal transplants have been successfully performed clinically. However, before becoming a routine clinical procedure, further preclinical studies are necessary to determine the underlying mechanisms of in situ tissue regeneration. Here we describe a protocol using a tissue engineering strategy and orthotopic transplantation of either natural decellularized donor tracheae or artificial electrospun nanofiber scaffolds into a rat model. The protocol includes details regarding how to assess the scaffolds' biomechanical properties and cell viability before implantation. It is a reliable and reproducible model that can be used to investigate the crucial aspects and pathways of in situ tracheal tissue restoration and regeneration. The model can be established in <6 months, and it may also provide a means to investigate cell-surface interactions, cell differentiation and stem cell fate.
Subject(s)
Guided Tissue Regeneration/methods , Tissue Engineering/methods , Tissue Scaffolds/chemistry , Trachea/physiology , Animals , Biomechanical Phenomena , Colorimetry , Nanofibers/therapeutic use , Rats , Trachea/transplantationABSTRACT
Neural circuits in the spinal cord transform instructive signals from the brain into well-coordinated locomotor movements by virtue of rhythm-generating components. Although evidence suggests that excitatory interneurons are the essence of locomotor rhythm generation, their molecular identity and the assessment of their necessity have remained unclear. Here we show, using larval zebrafish, that V2a interneurons represent an intrinsic source of excitation necessary for the normal expression of the locomotor rhythm. Acute and selective ablation of these interneurons increases the threshold of induction of swimming activity, decreases the burst frequency, and alters the coordination of the rostro-caudal propagation of activity. Thus, our results argue that V2a interneurons represent a source of excitation that endows the spinal circuit with the capacity to generate locomotion.
