Journal: Nature Communications

Article Title: Mitochondrial one-carbon metabolism is required for TGF-β-induced glycine synthesis and fibrotic responses

doi: 10.1038/s41467-025-64320-2

Figure Lengend Snippet: a Western blot analysis of collagen and α-smooth muscle actin (α-SMA) protein expression in human lung fibroblasts (HLFs) transfected with siRNA targeting MTHFD2 or nontargeting siRNA. Cells were treated with TGF-β for the indicated intervals. b Quantification of Collagen 1 and α-SMA levels in MTHFD2 knockdown HLFs relative to control cells at the indicated intervals. c Western blot analysis of GCN2 and S6-kinase phosphorylation and Collagen 1 expression in control and MTHFD2 knockdown HLFs. Cells were cultured in the presence or absence of extracellular glycine and were treated with TGF-β for the indicated intervals. Blots are representative of 4 separate experiments. d Western blot analysis of puromycin incorporation into newly translated proteins. Control and MTHFD2 knockdown HLFs were treated with TGF-β for 48 hours or left untreated. Cells were pulse labeled with puromycin for the indicated intervals. e qRT-PCR analysis of COL1A1 , ACTA2 , and COL3A1 mRNA expression in control and MTHFD2 knockdown HLFs either left untreated or treated with TGF-β for 48 hours. f Heatmap analysis of TGF-β-induced gene expression (differentially expressed genes between control knockdown untreated and control knockdown TGF-β, P < 0.01) in control and MTHFD2 knockdown HLFs treated with TGF-β for 48 hours. g Volcano plot showing differentially expressed genes between control knockdown TGF-β and combined MTHFD2 knockdown TGF-β-treated HLFs. h Proliferation analysis of control and MTHFD2 knockdown HLFs. i , j Analysis of cellular migration of control and MTHFD2 knockdown HLFs. k , l Analysis of contraction of control and MTHFD2 knockdown HLFs embedded in agarose and treated with TGF-β for 24 hours before gel release. All plots are presented as mean ± SD. Significance was calculated by 1-way ANOVA with Dunnett’s post-test ( b , n = 3 independent experiments), ( h , n = 4 biological replicates), ( j , n = 16 biological replicates), ( l , n = 4 biological replicates), 2-way ANOVA with Tukey’s post-test ( e , n = 3 biological replicates), and quasi-likelihood F-test ( f , n = 3 biological replicates). Source data are provided as a Source Data file.

Article Snippet: Mice receiving MTHFD2 inhibitor were IP injected with DS18561882 (MedChemExpress, HY-130251) beginning on day 8 after final bleomycin instillation.

Techniques: Western Blot, Expressing, Transfection, Knockdown, Control, Phospho-proteomics, Cell Culture, Labeling, Quantitative RT-PCR, Gene Expression, Migration

Journal: Nature Communications

Article Title: Mitochondrial one-carbon metabolism is required for TGF-β-induced glycine synthesis and fibrotic responses

doi: 10.1038/s41467-025-64320-2

Figure Lengend Snippet: a Schematic representation of metabolite labeling downstream of 2,3,3-D3-Serine. Human lung fibroblasts (HLFs) were transfected with siRNA targeting MTHFD2 or nontargeting siRNA. Cells were labeled with 2,3,3-D3-Serine and treated with TGF-β for 48 hours or left untreated. b – g Gas chromatography/mass spectrometry analysis of cellular metabolites. b Analysis of cellular glycine labeling from 2,3,3-D3-Serine in control and MTHFD2 knockdown HLFs. c Relative levels of M + 1 glycine from ( b ). d Analysis of cellular serine after labeling with 2,3,3-D3-Serine in control and MTHFD2 knockdown HLFs. e Relative levels of M + 1 and M + 2 serine from ( d ). f Analysis of cellular proline labeling from 2,3,3-D3-Serine in control and MTHFD2 knockdown HLFs. g Relative levels of M + 1 and proline from ( f ). h Relative levels of media formate content in NHLFs treated with TGF-β or left untreated. i – k Liquid chromatography/mass spectrometry analysis of cellular metabolites. i Analysis of cellular GTP labeling from 2,3,3-D3-Serine in control and MTHFD2 knockdown HLFs. j Analysis of cellular ATP labeling from 2,3,3-D3-Serine in control and MTHFD2 knockdown HLFs. k Analysis of cellular SAM labeling from 2,3,3-D3-Serine in control and MTHFD2 knockdown HLFs. Bar graphs represent mean ± SD, n = 3 biological replicates ( b – g , i – k ), n = 4 biological replicates ( h ). Significance was calculated by 2-way ANOVA with Tukey’s post-test. Source data are provided as a Source Data file.

Article Snippet: Mice receiving MTHFD2 inhibitor were IP injected with DS18561882 (MedChemExpress, HY-130251) beginning on day 8 after final bleomycin instillation.

Techniques: Labeling, Transfection, Gas Chromatography, Mass Spectrometry, Control, Knockdown, Liquid Chromatography

Journal: Nature Communications

Article Title: Mitochondrial one-carbon metabolism is required for TGF-β-induced glycine synthesis and fibrotic responses

doi: 10.1038/s41467-025-64320-2

Figure Lengend Snippet: a Uniform manifold approximation and projection (UMAP) clustering of lineage labeled alveolar fibroblast populations from mice as annotated by Tsukui et al. . b UMAP representation of the expression of Mthfd2 in lineage labeled alveolar fibroblasts at the indicated day after bleomycin instillation. c Dot plot representation of the expression of 1 C metabolic enzymes in total lineage labeled alveolar fibroblasts at the indicated day after bleomycin instillation. d Dot plot representation of the expression of 1 C metabolic enzymes in fibroblast subpopulations as in ( a ) on Day 21 after bleomycin instillation. e UMAP clustering of lung fibroblast populations from 10 control lungs and 20 lungs from patients with pulmonary fibrosis as defined by Habermann et al. . f UMAP representation of the expression of MTHFD2 in human lung fibroblasts as in ( e ). g Dot plot representation of the expression of 1 C metabolic enzymes in fibroblast subpopulations as in ( e ). h UMAP representation of subclustered alveolar fibroblasts and myofibroblasts from ( e ) showing alveolar, inflammatory, and fibrotic groups. i UMAP representation of the expression of MTHFD2 in human lung fibroblasts as in ( h ). j Dot plot representation of the expression of 1 C metabolic enzymes in fibroblast subpopulations as in ( h ). k Histological analysis of Mthfd2 protein expression in mouse lung tissue 21 days after instillation of either bleomycin or saline (vehicle). l Histological analysis of MTHFD2 protein expression in lung tissue from a control donor, or a patient with IPF. m Pearson correlation of MTHFD2 gene expression with two independent measures of lung function (FVC-pre-BD and DLCO) in patients with IPF. Chart represents best fit linear regression with 95% confidence interval. Data is from GSE32537 .

Article Snippet: Mice receiving MTHFD2 inhibitor were IP injected with DS18561882 (MedChemExpress, HY-130251) beginning on day 8 after final bleomycin instillation.

Techniques: Labeling, Expressing, Control, Saline, Gene Expression

Journal: Theranostics

Article Title: Identification of theranostic factors for patients developing metastasis after surgery for early-stage lung adenocarcinoma

doi: 10.7150/thno.53176

Figure Lengend Snippet: The functional assays of biomarker knockdown in A549 lung cancer cells using shRNAs against ADAM9, MTHFD2 , SLC2A1, and RRM2 individually or simultaneously (4G KD). (A) Western blots of control (shVOID), individual gene knockdown (shADAM9, shMTHFD2, shSLC2A1, and shRRM2), and 4G KD in A549 cells. (B) Colony formation of individual single gene knockdown and 4G KD A549 cells. Colonies were detected with crystal violet staining of the cell culture after 8 days (left) and quantified (right). (C) Cell migration ability of control, individual single gene knockdown, and 4G KD A549 cells. Cell migration was detected by time-lapse video microscopy (left) and quantified (right). (D) Western blots of control, individual gene knockdown, and 4G KD KD CL1-0 lung cancer cells. (E) Colony formation assay of control, individual gene knockdown, and 4G KD CL1-0 cells on day 11. (F) Cell migration ability of control, individual gene knockdown, and 4G KD CL1-0 cells. GAPDH served as the internal control. Error bars represent the mean ± SD of triplicate experiments. Statistical differences were analyzed using the unpaired t test ( * P < 0.05, ** P < 0.01).

Article Snippet: After blocking the sections with 2.5% goat or horse sera, primary antibodies against ADAM9 (AF949, R&D Systems, MN, USA; at 1:1000 dilution), RRM2 (HPA056994, Sigma, MO, USA; at 1:300 dilution), MTHFD2 (H00010797-M01, Abnova, Taiwan; at 1:1500 dilution), and SLC2A1 (07-14011, Millipore, Billerica, MA, USA; at 1:3000 dilution) were added and incubated overnight at 4 °C.

Techniques: Functional Assay, Biomarker Assay, Knockdown, Western Blot, Control, Staining, Cell Culture, Migration, Microscopy, Colony Assay

Journal: Theranostics

Article Title: Identification of theranostic factors for patients developing metastasis after surgery for early-stage lung adenocarcinoma

doi: 10.7150/thno.53176

Figure Lengend Snippet: The functional assays of biomarker overexpression in A549 and CL1-0 cells using transiently transfection of plasmids expressing ADAM9, SLC2A1, MTHFD2, and RRM2 proteins individually or combined plasmids (4G OE). (A) Western blots of vector control, individual gene overexpression, and 4G OE in A549 cells. (B) Colony formation of control, individual single gene, and 4G OE A549 cells on day 8. (C) Cell migration ability of control, individual single gene, and 4G OE A549 cells. (D) Western blots of control, individual single gene, and 4G OE CL1-0 cells. (E) Colony formation assay of control, individual single gene, and 4G OE CL1-0 cells on day 8. (F) Cell migration ability of control, individual single gene, and 4G OE CL1-0 cells. Tubulin served as the internal control. Error bars represent the mean ± SD of triplicate experiments. Statistical differences were analyzed using the unpaired t test ( * P < 0.05, ** P < 0.01).

Article Snippet: After blocking the sections with 2.5% goat or horse sera, primary antibodies against ADAM9 (AF949, R&D Systems, MN, USA; at 1:1000 dilution), RRM2 (HPA056994, Sigma, MO, USA; at 1:300 dilution), MTHFD2 (H00010797-M01, Abnova, Taiwan; at 1:1500 dilution), and SLC2A1 (07-14011, Millipore, Billerica, MA, USA; at 1:3000 dilution) were added and incubated overnight at 4 °C.

Techniques: Functional Assay, Biomarker Assay, Over Expression, Transfection, Expressing, Western Blot, Plasmid Preparation, Control, Migration, Colony Assay

Journal: bioRxiv

Article Title: Metabolic immunity to infection is driven by mitochondrial one-carbon metabolism

doi: 10.1101/2025.05.07.652698

Figure Lengend Snippet: ( A ) Immunoblot (IB) analyses of lysates from WT and eIF2α S51A AN3-12 mouse embryonic stem cells that were either uninfected, infected with Toxoplasma , or tunicamycin-treated (3 μg/ml) for 24 hours: ATF4, ∼50 kDa; phospho-EIF2α (Ser51), ∼38 kDa; α-Tubulin (TUBA), ∼55 kDa; and Toxoplasma GAP45 (TgGAP45), ∼45 kDa. ( B ) Heatmap of mRNA levels of ATF4 target genes in uninfected, infected and tunicamycin (3 μg/ml) treated ES-2 cells; n=3 independent cultures; SHMT2 and MTHFD2 are boxed. ( C and D ) WT and ATF4 KO ES-2 cells were uninfected or infected with Toxoplasma for 24 h and analyzed by qPCR for SHMT2 and MTHFD2 . Transcript levels were normalized to ACTB and are relative to WT uninfected. Data are mean ± SD of n=7 independent cultures. ****p<0.0001 for uninfected versus infected by means of two-way ANOVA analysis. ( E ) IB of ES-2 cells of the indicated genotype: MTHFD2, ∼32 kDa; α-Tubulin (TUBA), ∼55 kDa. ( F, G and H ) ES-2 cells of the indicated genotype were analyzed for mtDNA by qPCR for DLOOP , mt -ND1 and mt -CTYB normalized to RUNX2 levels and relative to WT uninfected. Data are mean ± SD of n=4 independent cultures. *p<0.05; for uninfected versus Toxoplasma infected, and #p<0.05; ##p<0.01 for WT versus MTHFD2 KO by means of two-way ANOVA analysis. ( I ) IB analysis of lysates from ES-2 cells that were uninfected or infected with indicated Toxo strains (MOI: 4) at 24 hpi: ATF4, ∼50 kDa; Vinculin (VCL), ∼124 kDa; and Toxoplasma GAP45 (TgGAP45), ∼45 kDa. ( J and K) Cells treated as in ( I ) were analyzed by qPCR for MTHFD2 and ASNS . Transcript levels were normalized to ACTB levels and are relative to uninfected cells. Data are mean ± SD of n=4 independent cultures, *p<0.05; **p<0.01 by means of one-way ANOVA analysis. ( L ) WT and ATF4 KO ES-2 were infected with Toxoplasma ± ISRIB (200nM) and analyzed 24 hpi by means of flow cytometry for Toxoplasma burden (mCherry median FI). Data are mean ± SEM of n=3 biological experiments, **p< 0.01 for DMSO versus ISRIB, #### p < 0.0001 for WT versus ATF4 KO by means of two-way ANOVA analysis. ( B - L ) ES-2 cells used for all experiments.

Article Snippet: The following antibodies were used: ATF4 (CST #11815), VCL (CST #4650), pS6K (CST #97596), TUBA (Proteintech 66031-1-Ig), MTHFD2 (CST #41377), SHMT1 (CST #80715), MTHFD1L (Invitrogen #PA5-100158), MTHFD1 ( Sigma #HPA015006), phospho-eIF2a (Ser51) (CST #9721), ACTB (CST #4970), TgGra45 (Dr. D Soldati; U. of Geneva) and TgGRA7 (Dr. J Boothroyd; Stanford University).

Techniques: Western Blot, Infection, Flow Cytometry

Journal: bioRxiv

Article Title: Metabolic immunity to infection is driven by mitochondrial one-carbon metabolism

doi: 10.1101/2025.05.07.652698

Figure Lengend Snippet: ( A ) Schematic of parasite and host one-carbon metabolism as determined by incorporation of 2 H from [2,3,3- 2 H] serine-labelling into dTMP and dTTP. Contribution of cytosolic-1C (cyto-1C) metabolism to dTMP production results in M+2 2 H-labelleled dTMP (2 blue dots), whereas its production from 1C units generated in mitochondria produces M+1 2 H-labelled dTMP (1 red dot). Toxoplasma has a cytosolic SHMT and thus parasite-1C metabolism generates M+2 2 H-labelleled dTMP. ( B and C ) dTMP and dTTP labeling for 24h during infection in WT and two clones of ATF4 KO ES-2 cells uninfected or infected with WT Toxoplasma and Δ cpsII (Carbamoyl phosphate synthase II) parasites. Data are mean ± SD of n=3 independent cultures, ****p<0.0001 for uninfected versus infected and ####p<0.0001 for WT versus ATF KO clones by means of two-way ANOVA analysis. ( D ) ES-2 cells with indicated genotypes were uninfected or infected with WT Toxoplasma or Δ cpsII parasites and harvested at 24 hpi for analysis by immunoblotting for ATF4, ∼50 kDa; Vinculin (VCL), ∼124 kDa; and Toxoplasma Gap45 (TgGAP45), ∼45 kDa. ( E ) Abundance of Toxoplasma dTMP levels in WT- Toxoplasma infected cells and ATF4 KO-infected cells for 24hpi. Data are mean ± SD of n=7 independent cultures, dTTP not detected in two cultures, *p<0.05; **p<0.01; for WT versus ATF4 KO by means of unpaired t-test. ( F ) Total levels of folate represented as (%) were measured by UPLC-MS/MS in uninfected and Toxoplasma -infected ES-2 cells at MOI=4 at 24 hpi and normalized to cell number. DHF: dihydrofolate; THF: tetrahydrofolate; CH=‘THF: 5,10-methenyl-tetrahydrofolate; CH 2 -THF: 5,10-methylene-tetrahydrofolate; CH 3 -THF: 5-methyl-tetrahydrofolate; CHO-THF: 10-formyl-tetrahydrofolate. Data are ± SD of n=4 independent cultures *p<0.05; ***p<0.001 for uninfected versus infected by means of multiple unpaired t-test. ( G ) WT and ATF4 KO ES-2 cells were infected with Toxoplasma ± MTX (200 nM) and analyzed 24 hpi by means of flow cytometry for Toxoplasma burden (mCherry median FI). Data are mean ± SEM of three biological experiments, ***p < 0.001 for DMSO versus MTX, and # p < 0.05 for WT versus ATF4 KO by means of two-way ANOVA analysis. ( H ) WT, MTHFD2 KO, and SHMT1 knockdown (KD) ES-2 cells were infected with Toxoplasma and analyzed as in ( G ). Data are mean ± SEM of five biological experiments, *p<0.05; ****p< 0.001; by means of one-way ANOVA; IB analyses of lysates from WT and SHMT1 KD cells: SHMT1, ∼55 kDa; VCL, ∼124 kDa. ( B-H ) ES-2 cells used for all experiments.

Article Snippet: The following antibodies were used: ATF4 (CST #11815), VCL (CST #4650), pS6K (CST #97596), TUBA (Proteintech 66031-1-Ig), MTHFD2 (CST #41377), SHMT1 (CST #80715), MTHFD1L (Invitrogen #PA5-100158), MTHFD1 ( Sigma #HPA015006), phospho-eIF2a (Ser51) (CST #9721), ACTB (CST #4970), TgGra45 (Dr. D Soldati; U. of Geneva) and TgGRA7 (Dr. J Boothroyd; Stanford University).

Techniques: Generated, Labeling, Infection, Clone Assay, Western Blot, Tandem Mass Spectroscopy, Flow Cytometry, Knockdown

Journal: bioRxiv

Article Title: Metabolic immunity to infection is driven by mitochondrial one-carbon metabolism

doi: 10.1101/2025.05.07.652698

Figure Lengend Snippet: ( A ) WT and SLC19A KO cells were analyzed by qPCR for SLC19A1 . Transcripts were normalized to ACTB . Data are mean ± SD of n=3 independent cultures. ( B ) WT and SLC19A1 knockdown (KD) ES-2 cells were cultured for 4 days in RPMI medium lacking folic acid and uninfected, infected with Toxoplasma (MOI: 4) cells or supplemented with folic acid (2 μM). After 24h, samples were analyzed by immunoblotting for the indicated antibodies MTHFD1L, ∼106 kDa; MTHFD2, ∼32 kDa; MTHFD1, ∼40 kDa and α-Actin (ACTB), ∼35 kDa. ( C ) WT and S LC19A1 KD ES-2 cells were infected with Toxoplasma and analyzed 24 hpi by means of flow cytometry for Toxoplasma burden [mCherry median FI]. Data are mean ± SEM of three biological experiments, *p < 0.05 for WT versus SLC19A1 KO, by means of one-way ANOVA analysis. ( D ) Total abundance of dTMP and dTTP of WT and S LC19A1 KO ES-2 uninfected and Toxo -infected cells. Data are mean ± of SED n=3 independent cultures, and normalized by cell number, *p<0.05; ****p<0.0001 for uninfected versus infected cells and ####p < 0.0001 for WT versus SLC19A1 KO cells by means of two-way ANOVA analysis.

Article Snippet: The following antibodies were used: ATF4 (CST #11815), VCL (CST #4650), pS6K (CST #97596), TUBA (Proteintech 66031-1-Ig), MTHFD2 (CST #41377), SHMT1 (CST #80715), MTHFD1L (Invitrogen #PA5-100158), MTHFD1 ( Sigma #HPA015006), phospho-eIF2a (Ser51) (CST #9721), ACTB (CST #4970), TgGra45 (Dr. D Soldati; U. of Geneva) and TgGRA7 (Dr. J Boothroyd; Stanford University).

Techniques: Knockdown, Cell Culture, Infection, Western Blot, Flow Cytometry

Journal: Redox Report : Communications in Free Radical Research

Article Title: MTHFD2-mediated redox homeostasis promotes gastric cancer progression under hypoxic conditions

doi: 10.1080/13510002.2024.2345455

Figure Lengend Snippet: MTHFD2 is highly expressed and predicts poor prognosis in GC. (A) Schematic representation of the function of MTHFD1/2 in folate metabolism. (B) Quantitative polymerase chain reaction (qPCR) analysis of MTHFD1/2 expression in 36 paired GC tissues. (C–D) MTHFD2 expression profiling in multiple GC microarray datasets from the Oncomine (C) and TCGA (D) databases. (E) Immunohistochemical analyses of 96 human GC specimens and their paired adjacent normal gastric tissues were performed. MTHFD2 in tumor tissues is higher than in adjacent tissues. Representative staining images are shown (scale bar = 100 μm). (F) Immunohistochemical staining scores of MTHFD2 in paired primary GC tumor and paired normal tissues. (G-H) Kaplan–Meier analysis of 5-year survival (G), overall survival and disease-free survival (H) for GC patients with low vs. high MTHFD2 expression. MTHFD1/2 = methylene tetrahydrofolate dehydrogenase 1/2; N = normal tissues; ANT = adjacent normal tissues; T = tumor. Data are presented as mean ± SD, Student’s t test (non-parametric comparisons for B and D, parametric comparison for C and F) or log-rank test (G-H). ns, not significant, ** P < 0.01.

Article Snippet: The following antibodies were used for immunoblotting or IHC analysis: MTHFD2 (1:200 for IHC and 1:1000 for immunoblotting, sc-100750) (Santa Cruz, CA, U.S.A.); cleaved caspase-3 (1:1000, #96664S), c-PARP (1:1000, #9185), β-actin (1:1000, #3700) (Cell Signaling, Beverly, U.S.A.); and Ki67 (1:200, #ZM-0167) (ZSGB-BIO, Beijing, China).

Techniques: Real-time Polymerase Chain Reaction, Expressing, Microarray, Immunohistochemical staining, Staining, Comparison

Journal: Redox Report : Communications in Free Radical Research

Article Title: MTHFD2-mediated redox homeostasis promotes gastric cancer progression under hypoxic conditions

doi: 10.1080/13510002.2024.2345455

Figure Lengend Snippet: MTHFD2 suppression inhibits GC cell proliferation. (A–B) qPCR and immunoblotting analysis of MTHFD2 expression levels in gastric cancer cell lines. (C) Immunoblotting analysis of MTHFD2 protein levels in paired GC and normal tissues. (D) Immunoblotting evaluating the knockdown efficiency of MTHFD2 with two unique shRNAs (#1, #2) in HGC27 and MKN74 cells. (E) Proliferation of sh-Control and sh-MTHFD2 MKN74 and HGC27 cells. (F-G) Colony formation assays in MKN74 and HGC27 cells after knockdown of MTHFD2. β-Actin was included as a loading control. Data are presented as mean ± SD, one-way ANOVA (non-parametric comparisons for A and G) or two-way ANOVA (E). ns, not significant, ** P < 0.01.

Article Snippet: The following antibodies were used for immunoblotting or IHC analysis: MTHFD2 (1:200 for IHC and 1:1000 for immunoblotting, sc-100750) (Santa Cruz, CA, U.S.A.); cleaved caspase-3 (1:1000, #96664S), c-PARP (1:1000, #9185), β-actin (1:1000, #3700) (Cell Signaling, Beverly, U.S.A.); and Ki67 (1:200, #ZM-0167) (ZSGB-BIO, Beijing, China).

Techniques: Western Blot, Expressing, Knockdown, Control

Journal: Redox Report : Communications in Free Radical Research

Article Title: MTHFD2-mediated redox homeostasis promotes gastric cancer progression under hypoxic conditions

doi: 10.1080/13510002.2024.2345455

Figure Lengend Snippet: MTHFD2 knockdown impairs NADPH homeostasis and accelerates GC cell death under hypoxia. (A) Gene set enrichment score and distribution of ROS-related genes along the rank of MTHFD2 up vs. MTHFD2 down available from The Cancer Genome Atlas GC database. (B–D) Cellular ROS (B), NADPH/NADP + (C) and GSH/GSSG (D) levels were measured in indicated GC cells under hypoxic conditions for 24 h. (E) Cell death was measured by Annexin-V/PI assays in the indicated GC cells under normal and hypoxic conditions for 48 h (red numbers indicate the subpopulation of cells positive for Annexin V/PI). (F) Quantification of cell death in the indicated cells cultured under hypoxic conditions for 72 h (with or without 5 mM N-acetyl-L-cysteine). (G) Immunoblotting analysis of cleaved poly ADP-ribose polymerase (PARP) in the indicated GC cells. GAPDH was included as a loading control. Nor = normoxia. Data are presented as mean ± SD, one-way ANOVA (non-parametric comparisons). ns, not significant, ** P < 0.01.

Article Snippet: The following antibodies were used for immunoblotting or IHC analysis: MTHFD2 (1:200 for IHC and 1:1000 for immunoblotting, sc-100750) (Santa Cruz, CA, U.S.A.); cleaved caspase-3 (1:1000, #96664S), c-PARP (1:1000, #9185), β-actin (1:1000, #3700) (Cell Signaling, Beverly, U.S.A.); and Ki67 (1:200, #ZM-0167) (ZSGB-BIO, Beijing, China).

Techniques: Knockdown, Cell Culture, Western Blot, Control

Journal: Redox Report : Communications in Free Radical Research

Article Title: MTHFD2-mediated redox homeostasis promotes gastric cancer progression under hypoxic conditions

doi: 10.1080/13510002.2024.2345455

Figure Lengend Snippet: MTHFD2 knockdown suppresses GC tumorigenesis in vivo. (A–C) A xenograft model was established in nude mice subcutaneously implanted with MTHFD2-knockdown and control MKN74 cells ( N = 5). Tumor images are presented (A). Tumor volumes were calculated (B) Tumor weights were obtained (C). (D–F) Patient-derived xenograft (PDX) tumors were implanted into the flanks of mice. Mice were treated with either PBS or si-MTHFD2. Tumor images are shown (D). Tumor volumes were calculated (E). Tumor weights were obtained (F). (G) Paraffin-embedded tumor sections derived from the indicated group were stained with hematoxylin and eosin (HE) or MTHFD2, Ki67 and cleaved caspase 3 antibodies or TUNEL assay. Ki67 levels are lower, whereas cleaved caspase 3 and TUNEL levels are higher, in tumor tissues with MTHFD2 knockdown. Representative staining images are shown (scale bar = 100 μm). H. The proliferation index (Ki67 staining), apoptotic index (cleaved caspase 3) and TUNEL index in tumor sections were quantified. Data are presented as mean ± SD, one-way ANOVA (parametric comparisons, C and F), Student’s t test (non-parametric comparison, H) or two-way ANOVA (B and E). ns, not significant, ** P < 0.01.

Article Snippet: The following antibodies were used for immunoblotting or IHC analysis: MTHFD2 (1:200 for IHC and 1:1000 for immunoblotting, sc-100750) (Santa Cruz, CA, U.S.A.); cleaved caspase-3 (1:1000, #96664S), c-PARP (1:1000, #9185), β-actin (1:1000, #3700) (Cell Signaling, Beverly, U.S.A.); and Ki67 (1:200, #ZM-0167) (ZSGB-BIO, Beijing, China).

Techniques: Knockdown, In Vivo, Control, Derivative Assay, Staining, TUNEL Assay, Comparison

Journal: Redox Report : Communications in Free Radical Research

Article Title: MTHFD2-mediated redox homeostasis promotes gastric cancer progression under hypoxic conditions

doi: 10.1080/13510002.2024.2345455

Figure Lengend Snippet: DS18561882 exerts antitumor effects against GC in vivo. (A–C) A xenograft model was established in nude mice subcutaneously implanted with MTHFD2-knockdown and control HGC27 cells ( N = 6). Tumor images are shown (A). Tumor volumes were calculated (B). Tumor weights were obtained (C). (D–F) Patient-derived xenograft (PDX) tumors were implanted into the flanks of mice. Mice were treated either with PBS or DS18561882. Tumor images are shown (D). Tumor volumes were calculated (E). Tumor weights were obtained (F). (G) Paraffin-embedded tumor sections derived from the indicated group were stained with hematoxylin and eosin (HE) or MTHFD2, Ki67, and cleaved caspase 3 antibodies or TUNEL assay. Ki67 levels are lower, whereas cleaved caspase 3 levels are higher, in tumor tissues with MTHFD2 inhibition. Representative staining images are shown (scale bar = 100 μm). H. The proliferation index (Ki67 staining), apoptotic index (cleaved caspase 3) and TUNEL index in tumor sections were quantified. Data are presented as mean ± SD, Student’s t test (parametric comparisons for C and F, non-parametric comparison for H) or two-way ANOVA (B and E). ns, not significant, ** P < 0.01.

Article Snippet: The following antibodies were used for immunoblotting or IHC analysis: MTHFD2 (1:200 for IHC and 1:1000 for immunoblotting, sc-100750) (Santa Cruz, CA, U.S.A.); cleaved caspase-3 (1:1000, #96664S), c-PARP (1:1000, #9185), β-actin (1:1000, #3700) (Cell Signaling, Beverly, U.S.A.); and Ki67 (1:200, #ZM-0167) (ZSGB-BIO, Beijing, China).

Techniques: In Vivo, Knockdown, Control, Derivative Assay, Staining, TUNEL Assay, Inhibition, Comparison