A Nanoscale Interface Promoting Molecular and Functional Differentiation of Neural Cells.

Tamara Posati,Emanuela Saracino,Marco Caprini,Francesco Formaggio,Francesco Valle,Michele Muccini,Morena Nocchetti,Assunta Pistone,Maria Grazia Mola,Marianna Barbalinardo,Roberto Zamboni,Grazia Paola Nicchia,Valentina Benfenati,Anna Sagnella,Simone Bonetti,Elisabetta Troni

doi:10.1038/srep31226

Abstract

Potassium channels and aquaporins expressed by astrocytes are key players in the maintenance of cerebral homeostasis and in brain pathophysiologies. One major challenge in the study of astrocyte membrane channels in vitro, is that their expression pattern does not resemble the one observed in vivo. Nanostructured interfaces represent a significant resource to control the cellular behaviour and functionalities at micro and nanoscale as well as to generate novel and more reliable models to study astrocytes in vitro. However, the potential of nanotechnologies in the manipulation of astrocytes ion channels and aquaporins has never been previously reported. Hydrotalcite-like compounds (HTlc) are layered materials with increasing potential as biocompatible nanoscale interface. Here, we evaluate the effect of the interaction of HTlc nanoparticles films with primary rat neocortical astrocytes. We show that HTlc films are biocompatible and do not promote gliotic reaction, while favouring astrocytes differentiation by induction of F-actin fibre alignment and vinculin polarization. Western Blot, Immunofluorescence and patch-clamp revealed that differentiation was accompanied by molecular and functional up-regulation of both inward rectifying potassium channel Kir 4.1 and aquaporin 4, AQP4. The reported results pave the way to engineering novel in vitro models to study astrocytes in a in vivo like condition.

Highlights

The capability of the Central Nervous System (CNS) to receive, integrate and compute information from and to the body is dependent on the maintenance of an electrochemical gradient of ions, organic molecules and osmotically driven water across the plasma-membrane of neural cells
Scanning electron microscopy (SEM) and atomic force microscopy (AFM) imaging were performed to provide an insight into the surface morphology, organization and roughness of Hydrotalcite-like compounds (HTlc) films
Since morphological alteration occurs upon gliosis[55,56], we evaluated the distribution and expression level of Glial Fibrillary Acid Protein (GFAP), a well known marker of astrogliotic cells in vivo and in vitro, by performing confocal imaging of immunofluorescent staining (Fig. 6a) and immunoblotting (Fig. 6b) of GFAP in astrocytes grown on PDL and on HTlc film, after 5 div

Summary

Introduction

The capability of the Central Nervous System (CNS) to receive, integrate and compute information from and to the body is dependent on the maintenance of an electrochemical gradient of ions, organic molecules and osmotically driven water across the plasma-membrane of neural cells. Astrocytes, by contacting neurons and cells lining the fluid-filled compartments (microvasculature, ventricular and subarachnoid spaces) are strategic players in the homeostatic regulation of the brain They are endowed with ion and water transmembrane channel proteins that are localized in specific plasma-membrane microdomains facing diverse liquid spaces. In a mouse model of Alzheimer Disease (Tg-ArcSwe mouse) changes in AQP4 expression are evident at the early stages of plaque formation, indicating that a loss of astrocytic AQP4 polarization could play a key role in the pathogenesis of the disease[6] All these pieces of evidence indicate that generating models intended for studying, manipulating and controlling of astrocytes Kir4.1 and AQP4 activity are highly desirable

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Results

Conclusion