IN VITRO CULTURE OF VARIOUS SPECIES OF MICROSPORIDIA CAUSING KERATITIS: EVALUATION OF THREE IMMORTALIZED CELL LINES

Object. Brain tumors, including gliomas, develop several months after rats are exposed in utero to N-ethyl-N-nitrosourea (ENU). Although pathological changes cannot be detected until these animals are several weeks old, the process that eventually leads to glioma formation must begin soon after exposure given the rapid clearance of the carcinogen and the observation that transformation of brain cells isolated soon after exposure occasionally occurs. This model can therefore potentially provide useful insights about the early events that precede overt glioma formation. The authors hypothesized that future glioma cells arise from stem/progenitor cells residing in or near the subventricular zone (SVZ) of the brain. Methods. Cells obtained from the SVZ or corpus striatum in ENU-exposed and control rats were cultured in an epidermal growth factor (EGF)-containing, chemically defined medium. Usually, rat SVZ cells cultured in this manner (neurospheres) are nestin-positive, undifferentiated, and EGF-dependent and undergo cell senescence. Consistent with these prior observations, control SVZ cells undergo senescence by the 12th to 15th doubling (20 of 20 cultures). In contrast, three of 15 cultures of cells derived from the SVZs of individual ENU-treated rats continue to proliferate for more than 60 cell passages. Each of these nestin-expressing immortalized cell lines harbored a common homozygous deletion spanning the INK4a/ARF locus and was unable to differentiate into neural lineages after exposure to specific in vitro stimuli. Nevertheless, unlike the rat C6 glioma cell line, these immortalized cell lines demonstrate EGF dependence and low clonogenicity in soft agar and did not form tumors after intracranial transplantation. Conclusions. Data in this study indicated that immortalized cells may represent glioma precursors that reside in the area of the SVZ after ENU exposure that may serve as a reservoir for further genetic and epigenetic hits that could eventually result in a full glioma phenotype.

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Blood-Brain Barrier In Vitro Model Utilizing Immortalized Cell Lines

IFMBE Proceedings - 25th Southern Biomedical Engineering Conference 2009, 15 – 17 May 2009, Miami, Florida, USA ◽

10.1007/978-3-642-01697-4_92 ◽

2009 ◽

pp. 263-264

Author(s):

Z. Zhang ◽

E. Crumpler ◽

C. Li

Keyword(s):

Blood Brain Barrier ◽

Cell Lines ◽

In Vitro Model ◽

Brain Barrier ◽

Immortalized Cell Lines ◽

Blood Brain ◽

Vitro Model ◽

Immortalized Cell

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Establishment of murine in vitro blood-brain barrier models using immortalized cell lines: co-cultures of brain endothelial cells, astrocytes, and neurons

10.1101/435990 ◽

2018 ◽

Cited By ~ 1

Author(s):

Fakhriedzwan Idris ◽

Siti Hanna Muharram ◽

Zainun Zaini ◽

Suwarni Diah

Keyword(s):

Endothelial Cells ◽

Blood Brain Barrier ◽

Cell Lines ◽

Brain Barrier ◽

Brain Endothelial Cells ◽

Immortalized Cell Lines ◽

Culture Model ◽

Immortalized Cell

AbstractBlood-brain barrier (BBB) is a selective barrier formed by the endothelial cells that line cerebral microvessels. It serves as a physical barrier due to the presence of complex tight junctions between adjacent endothelial cells which limits the paracellular movement of most molecules across the BBB. Many in vitro models of the BBB have been established to mimic these in vivo properties with limited success. In this study, we described the properties of a cell-based murine in vitro BBB model in five configurations constructed using immortalized cell lines in a 12-well format Transwell system: murine brain endothelial cells (bEnd.3) grown in a monoculture, or as co-culture in contact with astrocytes, or without contact with astrocytes or neurons, and triple co-culture combining the three cell lines. We found that only contact and triple co-culture model closely mimic the in vivo BBB tightness as evaluated by apparent permeability (Papp) of sucrose and albumin producing the lowest Papp values of 0.56 ± 0.16 × 10−6 cms−1 and 3.30 ± 0.51 × 10−6 cms−1, respectively, obtained in triple co-culture model. Co-culturing of bEnd.3 with astrocytes increased the expression of occludin as shown by western blot analysis, and immunohistochemistry showed an increase in peripheral localization of occludin and claudin-5. In addition, we found conditioned media were able to increase in vitro BBB model tightness through the modulation of tight junction proteins localization. We conclude that the presence of astrocytes and neurons in close proximity to brain endothelial cells is essential to produce a tight in vitro BBB model.

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Replacement of human primary dermal fibroblasts in in vitro toxico pharmacological models by immortalized cell lines

Journal of Dermatological Science ◽

10.1016/s0923-1811(98)83389-6 ◽

1998 ◽

Vol 16 ◽

pp. S66

Author(s):

Katja Herzhoff ◽

Salvatore Maniglia ◽

Beate Eckes ◽

Betty Nusgens ◽

Thomas Krieg ◽

...

Keyword(s):

Cell Lines ◽

Dermal Fibroblasts ◽

Immortalized Cell Lines ◽

Immortalized Cell

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Cell Type-Specific Adhesion and Migration on Laser-Structured Opaque Surfaces

International Journal of Molecular Sciences ◽

10.3390/ijms21228442 ◽

2020 ◽

Vol 21 (22) ◽

pp. 8442

Author(s):

Jörn Schaeske ◽

Elena Fadeeva ◽

Sabrina Schlie-Wolter ◽

Andrea Deiwick ◽

Boris N. Chichkov ◽

...

Keyword(s):

Cell Migration ◽

Cell Lines ◽

Adhesion Formation ◽

Focal Adhesions ◽

Primary Cells ◽

Migration Assay ◽

Immortalized Cell Lines ◽

Immortalized Cell

Cytocompatibility is essential for implant approval. However, initial in vitro screenings mainly include the quantity of adherent immortalized cells and cytotoxicity. Other vital parameters, such as cell migration and an in-depth understanding of the interaction between native tissue cells and implant surfaces, are rarely considered. We investigated different laser-fabricated spike structures using primary and immortalized cell lines of fibroblasts and osteoblasts and included quantification of the cell area, aspect ratio, and focal adhesions. Furthermore, we examined the three-dimensional cell interactions with spike topographies and developed a tailored migration assay for long-term monitoring on opaque materials. While fibroblasts and osteoblasts on small spikes retained their normal morphology, cells on medium and large spikes sank into the structures, affecting the composition of the cytoskeleton and thereby changing cell shape. Up to 14 days, migration appeared stronger on small spikes, probably as a consequence of adequate focal adhesion formation and an intact cytoskeleton, whereas human primary cells revealed differences in comparison to immortalized cell lines. The use of primary cells, analysis of the cell–implant structure interaction as well as cell migration might strengthen the evaluation of cytocompatibility and thereby improve the validity regarding the putative in vivo performance of implant material.

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