Fig 2: The silencing of ARG2 alleviated IL-1β-induced senescence and apoptosis in NP cells. (A,B) SA-β-gal staining and the ratio of SA-β-gal-positive cells. (C–E) The protein level of senescent marker p21 and P16INKa were detected via western blotting. (C,F,G) The protein level of apoptosis marker Cleaved-caspase-3, BCL-2, and BAX were detected via western blotting. (H,I) Mitochondrial membrane potential (MIMP) reflecting the early stage of apoptosis was assessed to detect apoptosis via JC1 staining (Scale bar: 100 μm). (J,K) The apoptosis rate was assessed with TUNEL staining (Scale bar: 100 μm). Data are represented as the mean ± SD. Two-group comparisons were analyzed by Student’s t-test. Significant differences between the treatment and control groups are indicated as *P < 0.05, **P < 0.01, and ***P < 0.001, n = 3.

Fig 3: Role of ARG2 in IL-1β-induced oxidative stress, inflammatory response, and apoptosis of NPCs via the activation of NF–κB pathway. (A–C) The effect of ARG2 overexpression on NF-κB. (D) The levels of ARG2, NF-κB and apoptosis related proteins in the human NPCs were measured by western blotting. (F–I) Quantitative analysis of western blotting results. (J) The intracellular localization of p65 was detected by immunofluorescence (Scale bar: 100 μm). (J,K) The levels of IL-6 and TNFα were detected by ELISA. (K) The level of ROS was detected (Scale bar: 100 μm). Data are represented as the mean ± SD. Two-group comparisons were analyzed by Student’s t-test. Significant differences between the treatment and control groups are indicated as *P < 0.05, **P < 0.01, and ***P < 0.001, n = 3.

Fig 4: The effect of ARG2 silencing on the expression of matrix metalloproteinases (MMPs), type II collagen, and aggrecan in NPCs. (A) The silencing efficiency of ARG2 was selected. (B) The efficacy of small interfering RNA (siRNA)-ARG2 in NPCs treated with IL-1β. (C,D) The effect of ARG2 downregulation on the mRNA levels of MMP3 and aggrecan of NPCs treated with IL-1β. (E) MMP13 and type II collagen levels were detected by immunofluorescence (Scale bar: 100 μm). (F–K) The effect of ARG2 downregulation on the protein levels of ARG2, type II collagen, aggrecan, MMP3, and MMP13 of NPCs treated with IL-1β. Data are represented as the mean ± SD. Two-group comparisons were analyzed by Student’s t-test. Significant differences between the treatment and control groups are indicated as *P < 0.05, **P < 0.01, and ***P < 0.001, n = 3.

Fig 5: Effect of L-Cit supplementation on NO synthesis and arginase activity in proximal small intestine of female mice with FFC-induced NASH. Levels of nitrite in (A) lumen content, and (B) tissue obtained from proximal small intestine, (C) densitometric analysis of 3-NT staining in proximal small intestine, (D) arginase activity in proximal small intestine, (E) mRNA expression of Arg2 in proximal small intestine, (F) relative expression of ARG2 protein in proximal small intestine with representative western blot. Data are presented as mean ± SEM, n = 8, except for nitrite in lumen content n = 5. Unpaired t-test was used to compare FFC and FFC + L-Cit after 13 weeks of feeding, *p ≤ 0.05. 3-NT, 3-nitrotyrosine; Arg2, arginase 2; C, control diet; L-Cit, l-citrulline; FFC, fat-, fructose- and cholesterol-rich diet; NASH, non-alcoholic steatohepatitis.