Abstracts

Brain damage may lead to neuronal cell death, gliosis or both. In a variety of pathologies including epilepsy, these glial changes tend to be permanent. In epileptic brain pronounced astrocytosis is frequently observed in pathological samples. Furthermore, [ldquo]epileptic[rdquo] astrocytes have been shown to express abnormal levels of multiple drug resistance proteins. We hypothesized that expression of multiple drug resistance is a consequence of altered mechanisms of apoptotic cell death. This was supported by previous findings by others, suggesting that mutant p53 cooperates to up regulate human multiple drug resistance genes. To test this, we used a combination of immunocytochemical and protein analysis techniques.
To explore a possible link between expression of multiple drug resistance genes and tumorigenesis, we studied expression of apoptotic genes in cells isolated from human brain resections through Western blotting techniques. Astrocyte and endothelial cell cultures were established from human cerebral cortical tissue of patients undergoing temporal lobectomies (n= 6) to relieve medically intractable seizures, one aneursym and one astrocytoma. Immunoblot and reverse transcriptase-polymerase chain reaction (RT-PCR) experiments were performed on confluent passage three cultures, which were 100% pure according to morphological criteria and positive staining for GFAP. To investigate the expression of multidrug resistance proteins in human tissues, we used mouse monoclonal anti-P-Glycoprotein, anti-MRP and anti-LRP antibodies.
Expression of MDR1 mRNA was observed in all cultures tested at the protein and mRNA level. Protein analysis demonstrated a higher expression of MDR1 in astrocytes isolated from epileptic brain compared to astrocytes from non-pathological samples. Low-grade astrocytoma isolated cells expressed levels of MDR1 comparable to epileptic specimens. p53 and p21 expression was induced in endothelial cells and normal astrocytes after [quot]in vitro[quot] culturing. Epileptic glia failed to express p53/p21 and loss of p53 coincided with MDR expression. This was also supported by previous findings obtained in tumor cells, suggesting that mutant p53 cooperates to up regulate human multiple drug resistance genes.
We conclude that: 1) Epileptic astrocytes express abnormal levels of MDR1; 2) These glia appear to lack normal mechanisms of cell apoptosis; 3) Incapacity to undergo programmed cell death may be a characteristic of [quot]epileptic[quot] glia, and 4) Anti-apoptotic transformation in glia is not exclusive property of gliomas.
[Supported by: HL51614, NS43284, NS38195]