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. 2023 Jun 10;299(7):104909. doi: 10.1016/j.jbc.2023.104909

Figure 4.

Figure 4

SHMT2-/MTHFD2-mediated mitochondrial folate metabolic pathway sustains TGF-β1 signaling.A, compartmentalization of FA-mediated 1C metabolism. B, qPCR detection of one-carbon metabolism gene expression in primary human HSC datasets from GSE67664. C, Western blot performed on cell lysates for α-SMA and collagen α1(I) after LX-2 cells were pretreated with DMSO or SHIN1 (1.25, 2.5, 5, 10 μM) or LY345899 (2.5, 5, 10, 20 μM) for 6 h, then stimulated with TGF-β1(5 ng/ml) for 24 h. D-H, Western blot for collagen α1(I) proteins expression in knockdown SHMT2, MTHFD2, MTHFD1L, SHMT1, MTHFD1 LX-2 cells treated with TGF-β1(5 ng/ml) for 36 h. I, LX-2 cells were stimulated multiple times by TGF-β1(5 ng/ml) at 0 h, 24 h, 48 h and were pretreated with DMSO or SHIN1 (5 μM) or LY345899 (5 μM) at 24 h. Flow cytometry analysis for collagen α1(I) expression at 0 h, 24 h, 48 h, 72 h. J, knockdown SHMT2 and MTHFD2 LX-2 cells were stimulated multiple times by TGF-β1 (5 ng/ml) at 0 h, 24 h, 48 h. Flow cytometry analysis for collagen α1(I) expression at 0, 24, 48, 72 h. The data are shown as mean ± SEM. The statistical significance (∗p < 0.05, ∗∗p < 0.01, ∗∗∗p < 0.001) was tested by unpaired t test. 1C, one-carbon; FA, folic acid; HSC, hepatic stellate cell; MTHFD, methylenetetrahydrofolate dehydrogenase; SHMT, serine hydroxymethyltransferase.