Article
IL-17A, IL-22, IL-23 and CCL20 – influence on migration and invasion of rat glioblastoma cell lines using in vitro techniques and a rat organotypic brain slice co-culture (OBSC)
IL-17A, IL-22, IL-23 und CCL20 – Einfluss auf Migration und Invasion von Ratten Glioblastomzelllinien unter Verwendung von in vitro Methoden und organotypischen Rattenhirnschnitt-Co-Kulturen (OBSC)
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Published: | May 25, 2022 |
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Objective: Tumor cell invasion has been demonstrated to be enhanced by inflammatory cytokines such as IL-17A, IL-22, IL-23 and CCL20 in various tumor entities other than glioblastoma (GBM). We have tested the impact of these factors on GBM migration and invasion utilizing four different in vitro techniques.
Methods: For avoiding xenogenous effects in the rat organotypic brain slice co-culture (OBSC), we use the rat glioblastoma cell lines 9L and S635 and as a non-neoplastic control cell line we used primary rat astrocytes. We employed a Boyden Chamber Invasion assay (transwell with matrigel), spheroid migration assay (4000 cell spheroid in round bottom well) and a wound healing migration assay. Moreover we have adopted and evolved rat OBSC to test for pro-migratory effects of the candidate cytokines: we use 6-12d rat brains, cut them in 350µm slices, put them on a 0.4µm insert swimming on culture medium. Next, we placed 4000 PKH67 fluorescence-labeled rat glioblastoma cells as spheroid-like structures in the area of the hippocampus next to the lateral ventricle and quantified the migration for 7 days.
Results: IL-22 shows enhancement of invasion and migration in our tested glioblastoma cell lines but not in the normal astrocytes. IL-17A shows enhancement of migration only on 9L cell lines. The establishment of the rat OBSC was feasible in our set-up and viability of the co-culture was shown over the course of 7 days, yet the effect of the above mentioned factors was not reproducible in the OBSC.
Conclusion: The effect of enhanced migration and/or invasion was not reproducible with our rat OBSC, probably because of minor discriminatory power due to our method of measuring the 2D extent of fluorescence. Refinement of measurement techniques might lead to more conclusive data in our rat OBSC. The downstream pathways leading to the effect with IL-22 and the effect on human cell lines remain to be examined.