gms | German Medical Science

Objective: It was previously demonstrated that the anti-proliferative effect of carnosine on glioblastoma cell lines was associated with increased expression of mRNA encoding pyruvate dehydrogenase kinase (PDK) 4 and reduced expression of PDK1 mRNA. In order to analyze whether these changes can also be observed in primary cultures and are also detectable at the protein level, further qRT-PCR and Western Blot experiments were performed. In addition, we analyzed whether carnosine does affect the activity of the pyruvate dehydrogenase complex (PDC).

Method: Cells from the glioblastoma cell lines T98G, U87 and LN405 and cells from 17 primary cultures, established after surgery of glioblastoma, were incubated for 24 hours in the absence and presence of 50 mM carnosine. Then, qRT-PCR experiments, Western Blot analysis and functional assays for the determination of PDC activity were performed.

Results: QRT-PCR analysis of the lines U87, T98G and LN405 and 17 primary cultures revealed enhanced expression of PDK4 mRNA after a 24 hour cultivation in 50 mM carnosine in all lines and in 6 primary cultures. One primary culture exhibited a reduced expression of PDK4 whereas no change was observed in the remaining 10 cultures. In addition, expression of PDK1 was reduced in all cell lines and 16 out of the 17 primary cultures (factors of reduction from 1.43 ± 0.15 to 10.2 ± 0.27). In order to analyze whether the effects observed at the level of mRNA are reflected by corresponding changes at the level of protein, Western Blot experiments were performed with cells from the three lines. These experiments revealed that in all lines the level of PDK1 protein expression significantly correlated with ist mRNA expression (p=0.0389). Surprisingly, there was no correlation between the expression of PDK4 protein and ist mRNA. In addition, despite the significant change of expression of PDK1 at the level of the protein, no significant change in the activity of the pyruvate dehydrogenase complex was observed.

Conclusions: Reduced expression of mRNA encoding PDK1 is more frequently observed in glioblastoma cells exposed to carnosine than the previously described enhanced expression of PDK4. In addition, no correlation between expression at the level of the protein and the mRNA were observed in the case of PDK4. Surprisingly, although a significant correlation was observed with regard to PDK1, a change of expression was not associated with a modified activity of the PDC.