title: Diversified expression of NG2/CSPG4 isoforms in glioblastoma and human foetal brain identifies pericyte subsets.
abstract: NG2/CSPG4 is a complex surface-associated proteoglycan (PG) recognized to be a widely expressed membrane component of glioblastoma (WHO grade IV) cells and angiogenic pericytes. To determine the precise expression pattern of NG2/CSPG4 on glioblastoma cells and pericytes, we generated a panel of >60 mouse monoclonal antibodies (mAbs) directed against the ectodomain of human NG2/CSPG4, partially characterized the mAbs, and performed a high-resolution distributional mapping of the PG in human foetal, adult and glioblastoma-affected brains. The reactivity pattern initially observed on reference tumour cell lines indicated that the mAbs recognized 48 immunologically distinct NG2/CSPG4 isoforms, and a total of 14 mAbs was found to identify NG2/CSPG4 isoforms in foetal and neoplastic cerebral sections. These were consistently absent in the adult brain, but exhibited a complementary expression pattern in angiogenic vessels of both tumour and foetal tissues. Considering the extreme pleomorphism of tumour areas, and with the aim of subsequently analysing the distributional pattern of the NG2/CSPG4 isoforms on similar histological vessel typologies, a preliminary study was carried out with endothelial cell and pericyte markers, and with selected vascular basement membrane (VBM) components. On both tumour areas characterized by 'glomeruloid' and 'garland vessels', which showed a remarkably similar cellular and molecular organization, and on developing brain vessels, spatially separated, phenotypically diversified pericyte subsets with a polarized expression of key surface components, including NG2/CSPG4, were disclosed. Interestingly, the majority of the immunolocalized NG2/CSPG4 isoforms present in glioblastoma tissue were present in foetal brain, except for one isoform that seemed to be exclusive of tumour cells, being absent in foetal brain. The results highlight an unprecedented, complex pattern of NG2/CSPG4 isoform expression in foetal and neoplastic CNS, discriminating between phenotype-specific and neoplastic versus non-neoplastic variants of the PG, thus opening up vistas for more selective immunotherapeutic targeting of brain tumours.
authors.affiliation: Department of Basic Medical Sciences, Neurosciences and Sensory Organs, University of Bari School of Medicine, Bari, Italy.
authors.forename: Francesco
authors.initials: F
authors.lastname: Girolamo
authors.affiliation: COMT - Centre for Molecular and Translational Oncology and Department of Biosciences, University of Parma, Parma, Italy.
authors.forename: Alice
authors.initials: A
authors.lastname: Dallatomasina
authors.affiliation: Department of Basic Medical Sciences, Neurosciences and Sensory Organs, University of Bari School of Medicine, Bari, Italy.
authors.forename: Marco
authors.initials: M
authors.lastname: Rizzi
authors.affiliation: Department of Basic Medical Sciences, Neurosciences and Sensory Organs, University of Bari School of Medicine, Bari, Italy.
authors.forename: Mariella
authors.initials: M
authors.lastname: Errede
authors.affiliation: Brain Research Institute, University of Zurich, Department of Health Sciences and Technology, Swiss Federal Institute of Technology (ETH) Zurich, Zurich, Switzerland ; Department of Neurosurgery, University Hospital Zurich, Zurich, Switzerland.
authors.forename: Thomas
authors.initials: T
authors.lastname: Wälchli
authors.affiliation: S.O.C. for Experimental Oncology 2, The National Cancer Institute Aviano, CRO-IRCCS, Aviano, Italy.
authors.forename: Maria Teresa
authors.initials: MT
authors.lastname: Mucignat
authors.affiliation: Department of Neurosurgery, University Hospital Zurich, Zurich, Switzerland.
authors.forename: Karl
authors.initials: K
authors.lastname: Frei
authors.affiliation: Department of Basic Medical Sciences, Neurosciences and Sensory Organs, University of Bari School of Medicine, Bari, Italy.
authors.forename: Luisa
authors.initials: L
authors.lastname: Roncali
authors.affiliation: COMT - Centre for Molecular and Translational Oncology and Department of Biosciences, University of Parma, Parma, Italy ; S.O.C. for Experimental Oncology 2, The National Cancer Institute Aviano, CRO-IRCCS, Aviano, Italy.
authors.forename: Roberto
authors.initials: R
authors.lastname: Perris
authors.affiliation: Department of Basic Medical Sciences, Neurosciences and Sensory Organs, University of Bari School of Medicine, Bari, Italy.
authors.forename: Daniela
authors.initials: D
authors.lastname: Virgintino
mesh_terms: D000328:Adult; D000818:Animals; D058846:Antibodies, Monoclonal, Murine-Derived / Q000737:chemistry; D000912:Antibodies, Neoplasm / Q000737:chemistry; D001921:Brain / Q000378:metabolism* / Q000473:pathology; D001932:Brain Neoplasms / Q000378:metabolism* / Q000473:pathology; D045744:Cell Line, Tumor; D011508:Chondroitin Sulfate Proteoglycans / Q000096:biosynthesis*; D005260:Female; D005333:Fetus / Q000378:metabolism* / Q000473:pathology; D015972:Gene Expression Regulation, Neoplastic; D005909:Glioblastoma / Q000378:metabolism* / Q000473:pathology; D006801:Humans; D008297:Male; D008565:Membrane Proteins / Q000096:biosynthesis*; D051379:Mice; D008807:Mice, Inbred BALB C; D009363:Neoplasm Proteins / Q000096:biosynthesis*; D020286:Pericytes / Q000378:metabolism* / Q000473:pathology; D020033:Protein Isoforms / Q000096:biosynthesis; D000099216:Chondroitin Sulfate Proteoglycan 4
chemical_list: D058846:Antibodies, Monoclonal, Murine-Derived; D000912:Antibodies, Neoplasm; D011508:Chondroitin Sulfate Proteoglycans; D008565:Membrane Proteins; D009363:Neoplasm Proteins; D020033:Protein Isoforms; C101390:CSPG4 protein, human; D000099216:Chondroitin Sulfate Proteoglycan 4