Lactic acid and acidification inhibit TNF secretion and glycolysis of human monocytes.

High concentrations of lactic acid (LA) are found under various pathophysiological conditions and are accompanied by an acidification of the environment. To study the impact of LA on TNF secretion, human LPS-stimulated monocytes were cultured with or without LA or the corresponding pH control. TNF secretion was significantly suppressed by low concentrations of LA (< or = 10 mM), whereas only strong acidification had a similar effect. This result was confirmed in a coculture model of human monocytes with multicellular tumor spheroids. Blocking synthesis of tumor-derived lactate by oxamic acid, an inhibitor of lactate dehydrogenase, reversed the suppression of TNF secretion in this coculture model. We then investigated possible mechanisms underlying the suppression. Uptake of [3-(13)C]lactate by monocytes was shown by hyphenated mass spectrometry. As lactate might interfere with glycolysis, the glycolytic flux of monocytes was determined. We added [1,2-(13)C(2)]glucose to the culture medium and measured glucose uptake and conversion into [2,3-(13)C(2)]lactate. Activation of monocytes increased the glycolytic flux and the secretion of lactate, whereas oxygen consumption was decreased. Addition of unlabeled LA resulted in a highly significant decrease in [2,3-(13)C(2)]lactate secretion, whereas a mere corresponding decrease in pH exerted a less pronounced effect. Both treatments increased intracellular [2,3-(13)C(2)]lactate levels. Blocking of glycolysis by 2-deoxyglucose strongly inhibited TNF secretion, whereas suppression of oxidative phosphorylation by rotenone had little effect. These results support the hypothesis that TNF secretion by human monocytes depends on glycolysis and suggest that LA and acidification may be involved in the suppression of TNF secretion in the tumor environment.

Warburg phenotype in renal cell carcinoma: high expression of glucose-transporter 1 (GLUT-1) correlates with low CD8(+) T-cell infiltration in the tumor.

Many tumor cells are characterized by a dysregulated glucose metabolism associated with increased glycolysis in the presence of oxygen ("Warburg Effect"). Here, we analyzed for the first time a possible link between glucose metabolism and immune cell infiltration in renal cell carcinoma (RCC). RCC specimens revealed a highly significant increase in the expression of lactate dehydrogenase A (LDHA) and glucose-transporter 1 (GLUT-1) compared to the corresponding normal kidney tissue on mRNA level. Accordingly, tumor cell lines of different origin such as RCC, melanoma and hepatocellular carcinoma strongly expressed LDHA and GLUT-1 compared to their nonmalignant counterparts. In line with this finding, tumor cells secreted high amounts of lactate. High expression of GLUT-1 and LDH5, a tetramer of 4 LDHA subunits, was confirmed by tissue microarray analysis of 249 RCC specimens. Overall, 55/79 (69.6%) and 46/71 (64.7%) cases of clear cell carcinoma showed a constitutive, but heterogeneous expression of GLUT-1 and LDH5, respectively. The number of CD3(+), CD8(+) and FOXP3(+) T cells was significantly elevated in RCC lesions compared to normal kidney epithelium, but effector molecules such as granzyme B and perforin were decreased in tumor infiltrating T cells. Of interest, further analysis revealed an inverse correlation between GLUT-1 expression and the number of CD8(+) T cells in RCC lesions. Together, our data suggest that an accelerated glucose metabolism in RCC tissue is associated with a low infiltration of CD8(+) effector T cells. Targeting the glucose metabolism may represent an interesting tool to improve the efficacy of specific immunotherapeutic approaches in RCC.

Lactate promotes glioma migration by TGF-beta2-dependent regulation of matrix metalloproteinase-2.

Lactate dehydrogenase type A (LDH-A) is a key metabolic enzyme catalyzing pyruvate into lactate and is excessively expressed by tumor cells. Transforming growth factor-beta2 (TGF-beta2) is a key regulator of invasion in high-grade gliomas, partially by inducing a mesenchymal phenotype and by remodeling the extracellular matrix. In this study, we tested the hypothesis that lactate metabolism regulates TGF-beta2-mediated migration of glioma cells. Small interfering RNA directed against LDH-A (siLDH-A) suppresses, and lactate induces, TGF-beta2 expression, suggesting that lactate metabolism is strongly associated with TGF-beta2 in glioma cells. Here we demonstrate that TGF-beta2 enhances expression, secretion, and activation of matrix metalloproteinase-2 (MMP-2) and induces the cell surface expression of integrin alpha(v)beta(3) receptors. In spheroid and Boyden chamber migration assays, inhibition of MMP-2 activity using a specific MMP-2 inhibitor and blocking of integrin alpha(v)beta(3) abrogated glioma cell migration stimulated by TGF-beta2. Furthermore, siLDH-A inhibited MMP2 activity, leading to inhibition of glioma migration. Taken together, we define an LDH-A-induced and TGF-beta2-coordinated regulatory cascade of transcriptional regulation of MMP-2 and integrin alpha(v)beta(3). This novel interaction between lactate metabolism and TGF-beta2 might constitute a crucial mechanism for glioma migration.

LDHA-Associated Lactic Acid Production Blunts Tumor Immunosurveillance by T and NK Cells.

Elevated lactate dehydrogenase A (LDHA) expression is associated with poor outcome in tumor patients. Here we show that LDHA-associated lactic acid accumulation in melanomas inhibits tumor surveillance by T and NK cells. In immunocompetent C57BL/6 mice, tumors with reduced lactic acid production (Ldhalow) developed significantly slower than control tumors and showed increased infiltration with IFN-γ-producing T and NK cells. However, in Rag2-/-γc-/- mice, lacking lymphocytes and NK cells, and in Ifng-/- mice, Ldhalow and control cells formed tumors at similar rates. Pathophysiological concentrations of lactic acid prevented upregulation of nuclear factor of activated T cells (NFAT) in T and NK cells, resulting in diminished IFN-γ production. Database analyses revealed negative correlations between LDHA expression and T cell activation markers in human melanoma patients. Our results demonstrate that lactic acid is a potent inhibitor of function and survival of T and NK cells leading to tumor immune escape.