Effects of chemotherapeutics on organotypic corticostriatal slice cultures identified by a panel of fluorescent and immunohistochemical markers.
Effects of chemotherapeutics on glioma cell lines and spheroids are usually investigated without evaluating the effects of chemotherapeutics on normal brain tissue. To perform such investigations, the aim of this study was to establish a panel of markers for detection of general cell death and more specific neuronal and glial degeneration induced by chemotherapeutics in organotypic rat corticostriatal slice cultures. The slice cultures were exposed to the alkylating agents temozolomide (TMZ) and nimustine (ACNU), the tyrosine kinase inhibitor imatinib mesylate (IM) and the microtubule-destabilizing agent vincristine (VCR). Densitometric measurements of uptake of the fluorescent dye propidium iodide (PI) were used for quantifying cellular degeneration. Moreover, paraffin sections were hematoxylin eosine stained and immunostained for the neuronal marker microtubule-associated protein 2 (MAP2), the astroglial marker glial fibrillary acidic protein (GFAP), and the oligodendroglial marker p25α. The results showed that the supposed clinically relevant drug concentrations were non-toxic. However, a time dependent increase in PI uptake was observed for high drug concentrations, except for TMZ, where no toxicity was observed. Corresponding immunostaining showed loss of MAP2 and increased expression of GFAP and p25α for cultures exposed to 1,000 nM VCR. Cultures exposed to high concentrations of ACNU and IM disintegrated, leaving no tissue for histology. In conclusion, corticostriatal slice cultures and the established panel of markers represent an excellent tool for detecting toxicity induced by chemotherapeutics. Toxicity was not detected at clinical concentrations, but high concentrations with toxic effects were identified suggesting that some of the earlier identified anti-cancer effects are general cytotoxic effects and not specific anti-cancer effects.
Department of Pathology, Odense University Hospital, Winsløwparken 15, 5000, Odense C, Denmark.
Annette
A
Nørregaard
Stine Skov
SS
Jensen
Jesper
J
Kolenda
Charlotte
C
Aaberg-Jessen
Karina Garnier
KG
Christensen
Poul Henning
PH
Jensen
Henrik Daa
HD
Schrøder
Bjarne Winther
BW
Kristensen
D000818:Animals; D000970:Antineoplastic Agents / Q000494:pharmacology*; D001549:Benzamides; D015415:Biomarkers / Q000032:analysis; D002352:Carrier Proteins / Q000032:analysis*; D016923:Cell Death / Q000187:drug effects; D003606:Dacarbazine / Q000031:analogs & derivatives / Q000494:pharmacology; D005904:Glial Fibrillary Acidic Protein / Q000032:analysis*; D000068877:Imatinib Mesylate; D007150:Immunohistochemistry; D007202:Indicators and Reagents; D008869:Microtubule-Associated Proteins / Q000032:analysis*; D009457:Neuroglia / Q000187:drug effects*; D009474:Neurons / Q000187:drug effects*; D015376:Nimustine / Q000494:pharmacology; D010879:Piperazines / Q000494:pharmacology; D011419:Propidium; D011743:Pyrimidines / Q000494:pharmacology; D051381:Rats; D000077204:Temozolomide; D014750:Vincristine / Q000494:pharmacology
D000970:Antineoplastic Agents; D001549:Benzamides; D015415:Biomarkers; D002352:Carrier Proteins; D005904:Glial Fibrillary Acidic Protein; D007202:Indicators and Reagents; C489787:MAP2 protein, rat; D008869:Microtubule-Associated Proteins; D010879:Piperazines; D011743:Pyrimidines; C539090:TPPP protein, rat; D015376:Nimustine; D011419:Propidium; D014750:Vincristine; D003606:Dacarbazine; D000068877:Imatinib Mesylate; D000077204:Temozolomide