Journal: Communications Biology

Article Title: L-cystine alleviates necrotizing enterocolitis by regulating ferroptosis and Th17 cell differentiation via the IL-6/STAT3 pathway

doi: 10.1038/s42003-025-09438-1

Figure Lengend Snippet: A The standard chemical structure of L-cystine. B Cell viability measured by CCK-8 assay. C IL-6 mRNA and protein levels analyzed by qRT-PCR and Western blot. D IL-6 concentration in cell supernatant quantified by ELISA. E ROS generation assessed via flow cytometry. F Intracellular Fe²⁺ levels. G 4-HNE expression detected by immunofluorescence. Scale bar = 25 μm. H , I MDA, GSH, and SOD levels. J Western blot analysis of STAT3 phosphorylation and NCOA4/FTH1 protein levels. n = 3 biologically independent samples. Data are presented as mean ± SD; * p < 0.05, ** p < 0.01, *** p < 0.001.

Article Snippet: For therapeutic intervention, pups received daily oral L-cystine (200 mg/mL in saline solution; C6H12N2O4S2, 56-89-3, MCE, Monmouth Junction, NJ, USA) and intraperitoneal injections of the STAT3 inhibitor S3I-201 (0.5 mg/mL in DMSO and 20% cyclodextrin; 501919-59-1, MCE) twice weekly .

Techniques: CCK-8 Assay, Quantitative RT-PCR, Western Blot, Concentration Assay, Enzyme-linked Immunosorbent Assay, Flow Cytometry, Expressing, Immunofluorescence, Phospho-proteomics

Journal: Communications Biology

Article Title: L-cystine alleviates necrotizing enterocolitis by regulating ferroptosis and Th17 cell differentiation via the IL-6/STAT3 pathway

doi: 10.1038/s42003-025-09438-1

Figure Lengend Snippet: A Cell viability assessed by CCK-8. B Fe²⁺ levels. C 4-HNE expression detected by immunofluorescence. Scale bar = 25 μm. D ROS generation via flow cytometry. E MDA, GSH, and SOD levels. F STAT3 phosphorylation and NCOA4/FTH1 levels using Western blot. n = 3 biologically independent samples. Data are presented as mean ± SD; * p < 0.05, ** p < 0.01, *** p < 0.001.

Article Snippet: For therapeutic intervention, pups received daily oral L-cystine (200 mg/mL in saline solution; C6H12N2O4S2, 56-89-3, MCE, Monmouth Junction, NJ, USA) and intraperitoneal injections of the STAT3 inhibitor S3I-201 (0.5 mg/mL in DMSO and 20% cyclodextrin; 501919-59-1, MCE) twice weekly .

Techniques: CCK-8 Assay, Expressing, Immunofluorescence, Flow Cytometry, Phospho-proteomics, Western Blot

Journal: Communications Biology

Article Title: L-cystine alleviates necrotizing enterocolitis by regulating ferroptosis and Th17 cell differentiation via the IL-6/STAT3 pathway

doi: 10.1038/s42003-025-09438-1

Figure Lengend Snippet: A Cell viability measured by CCK-8. B ROS generation via flow cytometry. C Fe²⁺ levels. D 4-HNE expression detected by immunofluorescence. Scale bar = 25 μm. E MDA, GSH, and SOD levels. F IL-6 concentration in supernatant by ELISA. G STAT3 phosphorylation and NCOA4/FTH1 levels using Western blot. n = 3 biologically independent samples. Data are presented as mean ± SD; * p < 0.05, ** p < 0.01, *** p < 0.001.

Article Snippet: For therapeutic intervention, pups received daily oral L-cystine (200 mg/mL in saline solution; C6H12N2O4S2, 56-89-3, MCE, Monmouth Junction, NJ, USA) and intraperitoneal injections of the STAT3 inhibitor S3I-201 (0.5 mg/mL in DMSO and 20% cyclodextrin; 501919-59-1, MCE) twice weekly .

Techniques: CCK-8 Assay, Flow Cytometry, Expressing, Immunofluorescence, Concentration Assay, Enzyme-linked Immunosorbent Assay, Phospho-proteomics, Western Blot

Journal: Communications Biology

Article Title: L-cystine alleviates necrotizing enterocolitis by regulating ferroptosis and Th17 cell differentiation via the IL-6/STAT3 pathway

doi: 10.1038/s42003-025-09438-1

Figure Lengend Snippet: A Flow cytometry sorting of native CD4 + T cells from patient peripheral blood. B , C Native CD4 + T cells treated with NCM-460-conditioned medium (CM) and anti-IL-6 antibody (900 nM) for 48 h. Flow cytometry analysis of Treg/Th17 populations and ratios. D , E Native CD4 + T cells treated with gradient concentrations of recombinant IL-6 (rIL-6). Flow cytometry analysis of Treg/Th17 populations and ratios. F , G Native CD4 + T cells treated with NCM-460-CM, rIL-6 (20 ng/mL), and STAT3 inhibitor S3I-201 (50 µM) for 48 h. Flow cytometry analysis of Treg/Th17 populations and ratios. H Western blot analysis of STAT3 phosphorylation. n = 3 biologically independent samples. Data are presented as mean ± SD; * p < 0.05, ** p < 0.01, *** p < 0.001.

Article Snippet: For therapeutic intervention, pups received daily oral L-cystine (200 mg/mL in saline solution; C6H12N2O4S2, 56-89-3, MCE, Monmouth Junction, NJ, USA) and intraperitoneal injections of the STAT3 inhibitor S3I-201 (0.5 mg/mL in DMSO and 20% cyclodextrin; 501919-59-1, MCE) twice weekly .

Techniques: Flow Cytometry, Recombinant, Western Blot, Phospho-proteomics

Journal: Communications Biology

Article Title: L-cystine alleviates necrotizing enterocolitis by regulating ferroptosis and Th17 cell differentiation via the IL-6/STAT3 pathway

doi: 10.1038/s42003-025-09438-1

Figure Lengend Snippet: A MDA, GSH, and SOD levels. B Flow cytometry analysis of ROS generation. C Fe²⁺ levels. D Mitochondrial morphology in colon tissues using TEM. Scale bar = 500 nm. E , F Immunofluorescence showing 4-HNE expression in colon tissues. Scale bar = 25 μm. G Western blot analysis of STAT3 phosphorylation and NCOA4/FTH1 levels. n = 3 animals (both sexes). Data are presented as mean ± SD; * p < 0.05, ** p < 0.01, *** p < 0.001.

Article Snippet: For therapeutic intervention, pups received daily oral L-cystine (200 mg/mL in saline solution; C6H12N2O4S2, 56-89-3, MCE, Monmouth Junction, NJ, USA) and intraperitoneal injections of the STAT3 inhibitor S3I-201 (0.5 mg/mL in DMSO and 20% cyclodextrin; 501919-59-1, MCE) twice weekly .

Techniques: Flow Cytometry, Immunofluorescence, Expressing, Western Blot, Phospho-proteomics