Depleting apolipoprotein C3 alleviates acute pancreatitis in hamsters but not in mice

Figure 1 Depletion of apolipoprotein C3 does not ameliorate caerulein-induced acute pancreatitis in mice. A: Experimental timeline: 8- to 9-week-old male mice received intraperitoneal injections of caerulein (50 μg/kg) once per hour for a total of 10 injections. Pancreatic tissues and blood samples were collected 24 hours after the first injection; B-D: Comparison of plasma triglyceride levels (B), amylase activity (C), and lipase activity (D) between wild type (WT) and apolipoprotein C3 knockout (ApoC3^-/-) mice, n = 4; E: Representative hematoxylin and eosin staining of pancreatic tissues from WT and ApoC3^-/- mice, with enlarged images shown below; F-I: Quantitative histological scoring of pancreatic tissue: Oedema (F), inflammation (G), necrosis (H), and total histology score (I) in WT and ApoC3^-/- mice, n = 4. Data are presented as the mean ± SEM. ^aP < 0.05, ^bP < 0.001. ApoC3: Apolipoprotein C3; AP: Acute pancreatitis; NS: Not significant; W: Week; WT: Wild type; ApoC3^-/-: Apolipoprotein C3 knockout; HE: Hematoxylin and eosin.

Figure 2 Inactivation of apolipoprotein C3 alleviates caerulein-induced acute pancreatitis in hamsters. A: Time-course curves of plasma amylase and lipase activities in wild type (WT) hamsters; B: Experimental timeline: 8- to 9-week-old male hamsters were intraperitoneally injected with caerulein (50 μg/kg) every hour for a total of 10 times, and samples were collected 24 hours after the first injection; C-F: Plasma biochemical parameters among different treatment groups, including triglycerides (C), NEFA (D), amylase activity (E), and lipase activity (F), n = 3-4; G: Representative hematoxylin and eosin staining of pancreatic tissue; enlarged views of selected areas are shown below; H-K: Quantitative histological scoring of pancreatic tissue: Oedema (H), inflammation (I), necrosis (J), and total pathology score (K), n = 3-4; L-P: Relative mRNA expression levels of inflammation- and apoptosis-related genes in the pancreas: Tnf-α (L), Il-6 (M), Nlrp3 (N), Bax (O), and Il-18 (P), n = 3-4; Q and R: Representative immunofluorescence staining of MPO (green) with DAPI (blue) for nuclear counterstaining (Q), and quantitative analysis of relative MPO-positive areas (R) in pancreatic tissue of WT and apolipoprotein C3 knockout (ApoC3^-/-) hamsters. White arrowheads indicate MPO-positive staining; scale bar: 75 μm; S and T: Immunofluorescence staining of CD68 (green) with DAPI (blue) (S), and quantification of relative CD68-positive areas (T) in pancreatic tissue of WT and ApoC3^-/- hamsters. White arrowheads indicate CD68-positive staining; scale bar: 75 μm. For each group, three animals were examined. Two fields of view per animal were quantified, and each field was considered an independent observation (n = 6 per group) for statistical comparisons. Data are presented as the mean ± SEM. ^aP < 0.05, ^bP < 0.01, ^cP < 0.001, and ^dP < 0.0001. ApoC3: Apolipoprotein C3; ApoC3^-/-: Apolipoprotein C3 knockout; AP: Acute pancreatitis; HE: Hematoxylin and eosin; NEFA: Non-esterified fatty acids; Tnf-α: Tumor necrosis factor-α; Il-6: Interleukin-6; Nlrp3: NLR family pyrin domain containing 3; Bax: BCL2-associated X protein; Il-18: Interleukin-18; DAPI: 4’,6-diamidino-2-phenylindole; MPO: Myeloperoxidase; CD68: Cluster of differentiation 68.

Figure 3 Apolipoprotein C3 deficiency alleviates ethanol + palmitoleic acid-induced acute pancreatitis in hamsters. A: Schematic of the experimental protocol: 8- to 9-week-old male hamsters were intraperitoneally injected with ethanol (EtOH) (1.35 g/kg) and palmitoleic acid (POA) (150 mg/kg) to induce pancreatitis. Blood samples were collected at 0, 6, 9, 12, and 24 hours, and the animals were sacrificed at 24 hours; B: Plasma triglyceride levels were measured across treatment groups, n = 3-5; C: Plasma amylase activity was assessed at multiple time points in wild type and apolipoprotein C3 knockout hamsters with or without EtOH + POA, n = 3-5; D: Representative hematoxylin and eosin-stained pancreatic tissue sections, with enlarged views shown below; E: Quantitative histological scoring of pancreatic tissue, evaluating oedema, inflammation, necrosis, and total pathology score, n = 3-5. Data are presented as the mean ± SEM. ^aP < 0.05, ^bP < 0.01 and ^cP < 0.001. ApoC3: Apolipoprotein C3; ApoC3^-/-: Apolipoprotein C3 knockout; AP: Acute pancreatitis; EtOH: Ethanol; POA: Palmitoleic acid; HE: Hematoxylin and eosin.

Figure 4 Loss of apolipoprotein C3 alleviates caerulein-induced acute pancreatitis under high-fat diet conditions in hamsters. A: Schematic of the experimental protocol: 8-week-old male hamsters were fed a high-fat diet (HFD) for 2 months. At 16 weeks of age, they received hourly intraperitoneal injections of caerulein (50 μg/kg) for a total of 10 times and were sacrificed 24 hours after the first injection; B and C: Plasma triglyceride (B) and NEFA (C) levels were measured under CD and HFD conditions in the indicated treatment groups, n = 4-6; D and E: Plasma amylase (D) and lipase (E) activities were assessed across treatment groups; F: Representative hematoxylin and eosin-stained pancreatic tissue sections from WT and apolipoprotein C3 knockout (ApoC3^-/-) hamsters; enlarged views are shown below, n = 4-6; G-J: Histological scoring of pancreatic tissue from WT and ApoC3^-/- hamsters, including oedema (G), inflammation (H), necrosis (I), and total pathology score (J), n = 4-6; K-O: Relative mRNA expression levels of inflammation- and apoptosis-related genes in pancreatic tissue from WT and ApoC3^-/- hamsters, including Tnf-α (K), Il-6 (L), Nlrp3 (M), Bax (N), and Il-18 (O) , n = 4; P and Q: Representative immunofluorescence staining of MPO (green) with DAPI (blue) nuclear counterstaining (P), and quantification of the relative MPO-positive area in pancreatic tissue of WT and ApoC3^-/- hamsters (Q). White arrowheads indicate MPO-positive staining; scale bar: 75 μm; R and S: Representative immunofluorescence staining of CD68 (green) with DAPI (blue) (R), and quantification of the relative CD68-positive area in the same groups (S). White arrowheads indicate CD68-positive staining; scale bar: 100 μm. For each group, three animals were used; two fields of view per animal were quantified as independent observations (n = 6 per group). Data are presented as the mean ± SEM. ^aP < 0.05, ^bP < 0.01, ^cP < 0.001, and ^dP < 0.0001. ApoC3: Apolipoprotein C3; ApoC3^-/-: Apolipoprotein C3 knockout; AP: Acute pancreatitis; CD: Chow diet; HFD: High-fat diet; TG: Triglyceride; NEFA: Non-esterified fatty acid; WT: Wild type; Tnf-α: Tumor necrosis factor-α; Il-6: Interleukin-6; Nlrp3: NLR family pyrin domain containing 3; Bax: BCL2-associated X protein; Il-18: Interleukin-18; DAPI: 4’,6-diamidino-2-phenylindole; MPO: Myeloperoxidase; CD68: Cluster of differentiation 68.