LDH-A Knockdown: Changes in The LDH Isoenzyme Profile and Variability in Glioma Response

Abstract

Background: Lactate metabolism in tumors is now recognized as a major energy source and a major gluconeogenic precursor for many tumors, as well as shown to exhibit signaling properties. There is less information on the role of the LDH/lactate axis in brain tumors, although lactate formation in gliomas is associated with poor survival. Methods: Three murine glioma cell lines (GL261, CT2A, and ALTS1C1) were transduced to knockdown (KD) expression of the murine LDH-A gene. The effects of the LDH-A KD were compared to those in control (NC) cells and tumors. Results: Differences in the expression of LDH-A and LDH-B mRNA, protein, and enzymatic activity were observed in the six cell lines. LDH zymography showed a major difference in LDH subunit distribution between GL261 LDH-A KD and NC tumors, whereas little or no effect of LDH-A KD was observed in CT2A and ALTS1C1 tumors. Tumors LDH-A and LDH-B immunohistochemistry and a Weka segmentation analysis were consistent with isoenzyme patterns and the above analyses. An “inverse” LDH-A/LDH-B staining relationship (high vs low) was observed in many local GL261 tumor regions. In contrast, CT2A tumors showed a more “direct” local LDH-A/LDH-B staining relationship. LDH-A KD prolonged the doubling time of GL261 cells in culture and prevented the formation of subcutaneous flank tumors in immune-competent C57BL/6 mice (GL261 NC tumors had a prolonged growth delay). In nude mice, both LDH-A KD and NC GL261 tumors grew more rapidly than GL261 NC tumors in C57BL/6 mice. No differences between NC and KD cell proliferation (in vitro) and tumor growth in C57BL/6 mice (doubling time) were observed for CT2A and ALTS1C1 cells and tumors, consistent with the absence of a difference in their LDH isoenzyme profiles. Conclusions: These results show the combined impact of a genetic alteration (LDH-A depletion) on the LDH isoenzyme profile, expression of LDH-A vs LDH-B and LDH enzymatic activity, and the immune system (C57BL/6 vs nude mice) on the growth of s.c. located tumors.