Related Experiment Video
Updated: Sep 23, 2025
Metabolic Mapping: Quantitative Enzyme Cytochemistry and Histochemistry to Determine the Activity of Dehydrogenases in Cells and Tissues
Published on: May 26, 2018
LDH-A-Modulation and the Variability of LDH Isoenzyme Profiles in Murine Gliomas: A Link with Metabolic and Growth
Masahiro Shindo1,2,3, Masatomo Maeda1,2,3, Ko Myat1,2,4
1Department of Neurology, Memorial Sloan Kettering Cancer Center, 1275 York Avenue, Box 52, New York, NY 10065, USA.
Abstract:
Three murine glioma cell lines (GL261, CT2A, and ALTS1C1) were modified to downregulate the expression of the murine LDH-A gene using shRNA, and compared to shRNA scrambled control (NC) cell lines. Differences in the expression of LDH-A and LDH-B mRNA, protein and enzymatic activity, as well as their LDH isoenzyme profiles, were observed in the six cell lines, and confirmed successful LDH-A KD. LDH-A KD (knock-down) resulted in metabolic changes in cells with a reduction in glycolysis (GlycoPER) and an increase in basal respiratory rate (mitoOCR). GL261 cells had a more limited ATP production capacity compared to CT2A and ALTS1C1 cells. An analysis of mRNA expression data indicated that: (i) GL261 LDH-A KD cells may have an improved ability to metabolize lactate into the TCA cycle; and (ii) that GL261 LDH-A KD cells can upregulate lipid metabolism/fatty acid oxidation pathways, whereas the other glioma cell lines do not have this capacity. These two observations suggest that GL261 LDH-A KD cells can develop/activate alternative metabolic pathways for enhanced survival in a nutrient-limited environment, and that specific nutrient limitations have a variable impact on tumor cell metabolism and proliferation. The phenotypic effects of LDH-A KD were compared to those in control (NC) cells and tumors. 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, whereas GL261 NC tumors had a prolonged growth delay in C57BL/6 mice. In nude mice, both LDH-A KD and NC GL261 tumors grew rapidly (more rapidly than GL261 NC tumors in C57BL/6 mice), demonstrating the impact of an intact immune system on GL261 tumor growth. No differences between NC and KD cell proliferation (in vitro) or tumor growth in C57BL/6 mice (doubling time) were observed for CT2A and ALTS1C1 cells and tumors, despite the small changes to their LDH isoenzyme profiles. These results suggest that GL261 glioma cells (but not CT2A and ALTS1C1 cells) are pre-programmed to have the capacity for activating different metabolic pathways with higher TCA cycle activity, and that this capacity is enhanced by LDH-A depletion. We observed that the combined impact of LDH-A depletion and the immune system had a significant impact on the growth of subcutaneous-located GL261 tumors.
Insights
Downregulating lactate dehydrogenase-A (LDH-A) in GL261 glioma cells alters metabolism, enhancing survival pathways and reducing tumor growth, especially in immune-competent hosts. This highlights LDH-A
Area of Science:
- Oncology
- Cancer Metabolism
- Immunology
Background:
- Lactate dehydrogenase-A (LDH-A) plays a crucial role in cancer cell metabolism, particularly in glycolysis.
- Understanding LDH-A's role in glioma is vital for developing targeted therapies.
Purpose of the Study:
- To investigate the metabolic and phenotypic consequences of LDH-A downregulation in murine glioma cell lines.
- To assess the impact of LDH-A knockdown on tumor growth in immunocompetent and immunodeficient mouse models.
Main Methods:
- Three murine glioma cell lines (GL261, CT2A, ALTS1C1) were engineered for LDH-A knockdown (KD) using shRNA.
- Metabolic analyses included glycolysis (GlycoPER) and respiration (mitoOCR) assays.
- Tumor growth was evaluated in C57BL/6 (immune-competent) and nude (immunodeficient) mice.
Main Results:
- LDH-A KD reduced glycolysis and increased respiration in glioma cells.
- GL261 LDH-A KD cells showed enhanced capacity for lactate metabolism and lipid oxidation, suggesting alternative survival pathways.
- LDH-A KD prevented GL261 tumor formation in C57BL/6 mice and significantly delayed growth, while nude mice showed rapid tumor growth for both KD and control cells.
- CT2A and ALTS1C1 cells showed minimal phenotypic changes post-LDH-A KD.
Conclusions:
- GL261 glioma cells possess a latent capacity for metabolic pathway activation, enhanced by LDH-A depletion.
- LDH-A knockdown significantly impacts GL261 tumor growth, particularly in the presence of an intact immune system.
- Tumor cell metabolism and proliferation are variably affected by nutrient limitations and genetic modifications, with immune status playing a critical role.
