Enhanced GLT-1 mediated glutamate uptake and migration of primary astrocytes directed by fibronectin-coated electrospun poly-L-lactic acid fibers.
Bioengineered fiber substrates are increasingly studied as a means to promote regeneration and remodeling in the injured central nervous system (CNS). Previous reports largely focused on the ability of oriented scaffolds to bridge injured regions and direct outgrowth of axonal projections. In the present work, we explored the effects of electrospun microfibers on the migration and physiological properties of brain astroglial cells. Primary rat astrocytes were cultured on either fibronectin-coated poly-L-lactic acid (PLLA) films, fibronectin-coated randomly oriented PLLA electrospun fibers, or fibronectin-coated aligned PLLA electrospun fibers. Aligned PLLA fibers strongly altered astrocytic morphology, orienting cell processes, actin microfilaments, and microtubules along the length of the fibers. On aligned fibers, astrocytes also significantly increased their migration rates in the direction of fiber orientation. We further investigated if fiber topography modifies astrocytic neuroprotective properties, namely glutamate and glutamine transport and metabolism. This was done by quantifying changes in mRNA expression (qRT-PCR) and protein levels (Western blotting) for a battery of relevant biomolecules. Interestingly, we found that cells grown on random and/or aligned fibers increased the expression levels of two glutamate transporters, GLAST and GLT-1, and an important metabolic enzyme, glutamine synthetase, as compared to the fibronectin-coated films. Functional assays revealed increases in glutamate transport rates due to GLT-1 mediated uptake, which was largely determined by the dihydrokainate-sensitive GLT-1. Overall, this study suggests that aligned PLLA fibers can promote directed astrocytic migration, and, of most importance, our in vitro results indicate for the first time that electrospun PLLA fibers can positively modify neuroprotective properties of glial cells by increasing rates of glutamate uptake.
Center for Biotechnology and Interdisciplinary Studies, Rensselaer Polytechnic Institute, Troy, NY 12180, USA; Department of Biomedical Engineering, Rensselaer Polytechnic Institute, Troy, NY 12180, USA.
Jonathan M
JM
Zuidema
Center for Neuropharmacology and Neuroscience, Albany Medical College, Albany, NY 12208, USA.
María C
MC
Hyzinski-García
Center for Biotechnology and Interdisciplinary Studies, Rensselaer Polytechnic Institute, Troy, NY 12180, USA; Department of Biology, Rensselaer Polytechnic Institute, Troy, NY 12180, USA.
Kristien
K
Van Vlasselaer
Center for Biotechnology and Interdisciplinary Studies, Rensselaer Polytechnic Institute, Troy, NY 12180, USA; Department of Biomedical Engineering, Rensselaer Polytechnic Institute, Troy, NY 12180, USA.
Nicholas W
NW
Zaccor
Center for Biotechnology and Interdisciplinary Studies, Rensselaer Polytechnic Institute, Troy, NY 12180, USA; Department of Biology, Rensselaer Polytechnic Institute, Troy, NY 12180, USA.
George E
GE
Plopper
Center for Neuropharmacology and Neuroscience, Albany Medical College, Albany, NY 12208, USA.
Alexander A
AA
Mongin
Center for Biotechnology and Interdisciplinary Studies, Rensselaer Polytechnic Institute, Troy, NY 12180, USA; Department of Biomedical Engineering, Rensselaer Polytechnic Institute, Troy, NY 12180, USA. Electronic address: gilber2@rpi.edu.
Ryan J
RJ
Gilbert
D000818:Animals; D001253:Astrocytes / Q000166:cytology / Q000378:metabolism*; D002448:Cell Adhesion; D002478:Cells, Cultured; D027342:Excitatory Amino Acid Transporter 2 / Q000502:physiology*; D005353:Fibronectins / Q000737:chemistry*; D018698:Glutamic Acid / Q000378:metabolism*; D019344:Lactic Acid / Q000737:chemistry*; D011091:Polyesters; D011108:Polymers / Q000737:chemistry*; D051381:Rats; D017207:Rats, Sprague-Dawley; D020133:Reverse Transcriptase Polymerase Chain Reaction; D054457:Tissue Scaffolds
D027342:Excitatory Amino Acid Transporter 2; D005353:Fibronectins; D011091:Polyesters; D011108:Polymers; C493395:Slc1a2 protein, rat; D019344:Lactic Acid; D018698:Glutamic Acid; C033616:poly(lactide)
Aligned microfibers; Astrocytes; Glutamate metabolism; Glutamate transport; Migration; Poly-l-lactic acid