Maja Žugec, Borut Furlani, Maria J Castañon, Boštjan Rituper, Irmgard Fischer, Giuseppe Broggi, Rosario Caltabiano, Giuseppe M V Barbagallo, Michelino Di Rosa, Daniele Tibullo, Rosalba Parenti, Nunzio Vicario, Saša Simčič, Victorio Martin Pozo Devoto, Gorazd B Stokin, Gerhard Wiche, Jernej Jorgačevski, Robert Zorec, Maja Potokar
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引用次数: 0
Abstract
Background: The expression of aquaporin 4 (AQP4) and intermediate filament (IF) proteins is altered in malignant glioblastoma (GBM), yet the expression of the major IF-based cytolinker, plectin (PLEC), and its contribution to GBM migration and invasiveness, are unknown. Here, we assessed the contribution of plectin in affecting the distribution of plasmalemmal AQP4 aggregates, migratory properties, and regulation of cell volume in astrocytes.
Methods: In human GBM, the expression of glial fibrillary acidic protein (GFAP), AQP4 and PLEC transcripts was analyzed using publicly available datasets, and the colocalization of PLEC with AQP4 and with GFAP was determined by immunohistochemistry. We performed experiments on wild-type and plectin-deficient primary and immortalized mouse astrocytes, human astrocytes and permanent cell lines (U-251 MG and T98G) derived from a human malignant GBM. The expression of plectin isoforms in mouse astrocytes was assessed by quantitative real-time PCR. Transfection, immunolabeling and confocal microscopy were used to assess plectin-induced alterations in the distribution of the cytoskeleton, the influence of plectin and its isoforms on the abundance and size of plasmalemmal AQP4 aggregates, and the presence of plectin at the plasma membrane. The release of plectin from cells was measured by ELISA. The migration and dynamics of cell volume regulation of immortalized astrocytes were assessed by the wound-healing assay and calcein labeling, respectively.
Results: A positive correlation was found between plectin and AQP4 at the level of gene expression and protein localization in tumorous brain samples. Deficiency of plectin led to a decrease in the abundance and size of plasmalemmal AQP4 aggregates and altered distribution and bundling of the cytoskeleton. Astrocytes predominantly expressed P1c, P1e, and P1g plectin isoforms. The predominant plectin isoform associated with plasmalemmal AQP4 aggregates was P1c, which also affected the mobility of astrocytes most prominently. In the absence of plectin, the collective migration of astrocytes was impaired and the dynamics of cytoplasmic volume changes in peripheral cell regions decreased. Plectin's abundance on the plasma membrane surface and its release from cells were increased in the GBM cell lines.
Conclusions: Plectin affects cellular properties that contribute to the pathology of GBM. The observed increase in both cell surface and released plectin levels represents a potential biomarker and therapeutic target in the diagnostics and treatment of GBMs.
期刊介绍:
The Journal of Biomedical Science is an open access, peer-reviewed journal that focuses on fundamental and molecular aspects of basic medical sciences. It emphasizes molecular studies of biomedical problems and mechanisms. The National Science and Technology Council (NSTC), Taiwan supports the journal and covers the publication costs for accepted articles. The journal aims to provide an international platform for interdisciplinary discussions and contribute to the advancement of medicine. It benefits both readers and authors by accelerating the dissemination of research information and providing maximum access to scholarly communication. All articles published in the Journal of Biomedical Science are included in various databases such as Biological Abstracts, BIOSIS, CABI, CAS, Citebase, Current contents, DOAJ, Embase, EmBiology, and Global Health, among others.