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. 2019 Jul 9;156(6):1137–1148. doi: 10.1016/j.chest.2019.05.038

Figure 4.

Figure 4

5′-Methylthioadenosine (MTA)-related polyamine metabolism was upregulated by combinatorial treatment of rapamycin and chloroquine in TSC2-deficient 621-101 cells derived from a patient with lymphangioleiomyomatosis. A, Heatmap of significantly upregulated or downregulated metabolites (rows) across treatment conditions (columns). 621-101 cells (passage 24) were treated for 24 hours with 20 nM rapamycin, 5 μM chloroquine, a combination of 20 nM rapamycin and 5 μM chloroquine, or dimethylsulfoxide (DMSO) control. Many intermediate metabolites in polyamine metabolism were upregulated by the combinatorial treatment with rapamycin and chloroquine, including MTA, arginine, and S-adenosyl-l-homocysteine. B, Volcano plot shows that polyamine metabolites were upregulated by treatment with rapamycin and chloroquine (CQ) compared with DMSO control, including MTA and arginine (labeled). Dots on the right sides represent metabolites upregulated by the treatment, and dots on the left side represent metabolites downregulated by the treatment. Metabolites in the polyamine metabolism pathway are depicted as green dots. The x axis represents fold change; the y axis represents negative log10-transformed P values. The dashed red line indicates a false discovery rate (FDR) cutoff of 0.05. Same experiment as described in (A). C, Arginine, but not MTA, was significantly upregulated by rapamycin. Metabolites in the polyamine pathway are represented as green dots. The horizontal dashed red line indicates an FDR cutoff of 0.05. Same experiment as described in (A). D, Arginine was significantly upregulated by the treatment with rapamycin and by the combinatorial treatment with rapamycin and CQ (FDR <0.05). Combinatorial treatment of rapamycin and chloroquine significantly decreased arginine compared with rapamycin treatment alone. E, MTA was significantly upregulated by the combinatorial treatment with rapamycin and CQ, but not by either rapamycin or chloroquine alone. No significant difference is observed between rapamycin treatment alone and combinatorial treatment of rapamycin and chloroquine (denoted as ns). ns = not significant; Rapa = rapamycin.