Cellular and Molecular NeuroscienceResearch PaperImmortalization and functional characterization of rat arachnoid cell lines
Research highlights
▶This is the first report of an immortalized arachnoid cell line derived from normal primary cells. ▶We created immortalized rat arachnoid cell lines using two retroviral constructs. ▶Cell lines were characterized on the basis of phenotypic markers, growth properties, and physiology. ▶This paper has direct relevance to modeling hydrocephalus.
Section snippets
Primary culture of rat arachnoid cells
Arachnoid cells were harvested from 21 to 23 day old female Sprague–Dawley rats. The animals were euthanized with CO2 according to an approved VA Medical Center animal protocol. Carcasses were thoroughly cleaned with 70% ethanol. The cranium and cervical spine were exposed and the brain and upper spinal cord was removed en bloc. The leptomeninges were removed by gentle dissection from the brainstem and finely minced with a #15 scalpel, then placed in Dulbecco's modified Eagle's medium (DMEM,
Results
The aim of this study was developing and partially characterizing immortal arachnoid cell lines which could be utilized for physiological studies in modeling normal CSF flow, and which could be modified or treated with various substances to study pathological conditions such as hydrocephalus, arachnoid cysts, or other disorders. As a starting point, the following basic facts needed to be established: (1) that the cells had been stably transduced with the intended gene constructs; (2) that the
Discussion
In this study, we report the first immortalization of primary arachnoid cells, as an initial step toward manufacture of arachnoid cell bioscaffolds for modeling cellular transport properties. We utilized both a one-step and a two-step retroviral transduction with LTAg and hTERT. If desired, the two-step approach can be adapted to human source material with amphotropic MMLV-based viral packaging. Irrespective of approach, we observed similar functional properties of the ST and DT cells, which
Acknowledgments
This work was supported by the University of Minnesota Institute of Engineering in Medicine and the University of Minnesota Stem Cell Institute. Additional support came from the Minneapolis VAMC Research Advisory Group and the National Endowment for Alzheimer's Research. This work received the 2010 Hydrocephalus Association Award for meritorious applied research in hydrocephalus.
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