Journal: Frontiers in Nutrition

Article Title: Reduced hepatic GLUL expression accompanies altered RIP3/MLKL-pathway protein abundance in early cholestatic liver injury following bile duct ligation

doi: 10.3389/fnut.2026.1912372

Figure Lengend Snippet: Experimental assessment of candidate genes and necroptosis-associated markers by RT-qPCR. Relative mRNA expression levels of (A) Glul , (B) Ifngr1 , (C) Sqstm1 , (D) Ripk1 , (E) Ripk3 , and (F) Mlkl in SH and BDL liver tissues. Gapdh served as the internal control. Data are presented as mean ± SD ( n = 6 per group). ** p < 0.01.

Article Snippet: Membranes were blocked with 5% non-fat milk for 1 h at room temperature and incubated overnight at 4 °C with primary antibodies against glutamine synthetase (GLUL), receptor-interacting protein kinase 3 (RIP3), phospho-RIP3 (p-RIP3), mixed lineage kinase domain-like protein (MLKL), phospho-mixed lineage kinase domain-like protein (p-MLKL), β-tubulin, and GAPDH (Proteintech, Wuhan, China).

Techniques: Quantitative RT-PCR, Expressing, Control

Journal: Frontiers in Nutrition

Article Title: Reduced hepatic GLUL expression accompanies altered RIP3/MLKL-pathway protein abundance in early cholestatic liver injury following bile duct ligation

doi: 10.3389/fnut.2026.1912372

Figure Lengend Snippet: Western blot assessment of GLUL and RIP3/MLKL-pathway proteins. (A) Representative immunoblots of GLUL, β-tubulin, RIP3, p-RIP3, MLKL, p-MLKL, and GAPDH. Three representative samples from each group are shown. Quantification of (B) GLUL normalized to β-tubulin and (C) RIP3, (D) p-RIP3, (E) MLKL, and (F) p-MLKL normalized to GAPDH. (G) p-RIP3/RIP3 ratio. (H) p-MLKL/MLKL ratio. Ratios were calculated for each biological replicate from the corresponding GAPDH-normalized values. Data are presented as mean ± SD ( n = 6 per group). * p < 0.05; ** p < 0.01; *** p < 0.001.

Article Snippet: Membranes were blocked with 5% non-fat milk for 1 h at room temperature and incubated overnight at 4 °C with primary antibodies against glutamine synthetase (GLUL), receptor-interacting protein kinase 3 (RIP3), phospho-RIP3 (p-RIP3), mixed lineage kinase domain-like protein (MLKL), phospho-mixed lineage kinase domain-like protein (p-MLKL), β-tubulin, and GAPDH (Proteintech, Wuhan, China).

Techniques: Western Blot

Journal: Frontiers in Nutrition

Article Title: Reduced hepatic GLUL expression accompanies altered RIP3/MLKL-pathway protein abundance in early cholestatic liver injury following bile duct ligation

doi: 10.3389/fnut.2026.1912372

Figure Lengend Snippet: Hypothesis-generating framework summarizing concurrent changes observed during early cholestatic liver injury. Bile duct ligation was accompanied by biochemical and histological liver injury, transcriptomic enrichment of metabolic processes, reduced Glul mRNA expression and GLUL protein abundance, increased GAPDH-normalized RIP3, p-RIP3, MLKL, and p-MLKL abundance, and significant increases in both p-RIP3/RIP3 and p-MLKL/MLKL. Solid arrows indicate concurrent observations in the 24-h BDL model and do not imply causal or temporal relationships. GLUL activity, glutamine-related metabolites, necroptotic flux, and direct pathway dependence were not measured.

Article Snippet: Membranes were blocked with 5% non-fat milk for 1 h at room temperature and incubated overnight at 4 °C with primary antibodies against glutamine synthetase (GLUL), receptor-interacting protein kinase 3 (RIP3), phospho-RIP3 (p-RIP3), mixed lineage kinase domain-like protein (MLKL), phospho-mixed lineage kinase domain-like protein (p-MLKL), β-tubulin, and GAPDH (Proteintech, Wuhan, China).

Techniques: Ligation, Expressing, Quantitative Proteomics, Activity Assay