Journal: Diabetes & Metabolism Journal

Article Title: Interleukin 33 Promotes Liver Sinusoidal Endothelial Cell Dysfunction and Hepatic Fibrosis in Diabetic Mice

doi: 10.4093/dmj.2024.0532

Figure Lengend Snippet: Diabetic liver fibrosis is alleviated in interleukin 33 (IL33)-deficient mice. (A) Schematic diagram of the experimental procedure, mice were fed with normal diet (ND) or high-fat diet (HFD) for a total of 26 weeks. After 12 weeks, HFD-fed mice were consecutively injected with streptozotocin (STZ, 50 mg/kg) for 5 days. Diabetic phenotype was validated by fasting blood glucose ≥11.1 mmol/L at 7 days later (14 weeks). HFD-fed mice were consecutively injected with αIL33, immunoglobulin G (IgG), recombinant IL33 (rIL33), or vehicle for 12 weeks. (B) Serum IL33 level in control (Ctrl) mice and diabetic (DM) mice assessed by enzyme-linked immunosorbent assay (ELISA) ( n =12). (C) Western blot and measurement for IL33 expression of whole liver ( n =6). (D) Representative images of H&E and Sirius Red staining of liver sections from DM mice and IL33 knockout (KO) diabetic mice. Scale bar, 100 μm. (E) A positive area of Sirius Red staining is used to quantify liver fibrosis (right) ( n =4). (F) Hepatic hydroxyproline levels in mice ( n =4). (G, H) Representative images and quantify of immunohistochemical (IHC) staining of alpha smooth muscle actin (αSMA) in liver sections ( n =4). Scale bar, 100 μm. Data was shown as mean±standard error of the mean. ST2, suppression of tumorigenicity 2; i.p., intraperitoneal; PBS, phosphate buffered saline; GAPDH, glyceraldehyde-3-phosphate dehydrogenase. a P <0.01.

Article Snippet: Primary antibodies targeting IL33 (Proteintech, Rosemont, IL, USA; 1:1,000; 12372-1-AP), ST2 (Proteintech, 1:1,000; 11920-1-AP), P62 (Cell Signaling Technology [CST], Danvers, MA, USA; 1:1,000; 5114T), microtubule-associated protein 1 light chain 3 B (CST, 1:1,000; 3868S), phosphor-extracellular signal-regulated kinase (p-ERK) (CST, 1:1,000; 4370T), ERK (CST, 1:2,000; 9108S), and glyceraldehyde-3-phosphate dehydrogenase (GAPDH) (CST, 1:2,000; 2118S).

Techniques: Injection, Recombinant, Control, Enzyme-linked Immunosorbent Assay, Western Blot, Expressing, Staining, Knock-Out, Immunohistochemical staining, Immunohistochemistry, Saline

Journal: Diabetes & Metabolism Journal

Article Title: Interleukin 33 Promotes Liver Sinusoidal Endothelial Cell Dysfunction and Hepatic Fibrosis in Diabetic Mice

doi: 10.4093/dmj.2024.0532

Figure Lengend Snippet: Interleukin 33 (IL33) promotes liver sinusoidal endothelial cell (LSEC) dysfunction in diabetic mice. (A) Representative scanning electron micrographs of livers of mice. Scale bar, 5 μm. (B) Hepatic nitric oxide (NO) levels ( n =8). (C, D) Representative immunofluorescent images and analysis of lymphatic vessel endothelial hyaluronan receptor 1 (LYVE-1) and CD31 of liver sections ( n =4) (scale bar, 100 μm). (E) Relative mRNA abundance of vascular cell adhesion molecule 1 (Vcam1), intercellular adhesion molecule 1 (Icam1), nitric oxide synthase 2 (Nos2), endothelin 1 (Edn-1), nitric oxide synthase 3 (Nos3), and KLF transcription factor 2 (Klf2) of liver tissue ( n =8). Data was shown as mean±standard error of the mean. Ctrl, control; DM, diabetes mellitus; IgG, immunoglobulin G; PBS, phosphate buffered saline; rIL33, recombinant IL33; NS, no significant. a P <0.05, b P <0.01.

Article Snippet: Primary antibodies targeting IL33 (Proteintech, Rosemont, IL, USA; 1:1,000; 12372-1-AP), ST2 (Proteintech, 1:1,000; 11920-1-AP), P62 (Cell Signaling Technology [CST], Danvers, MA, USA; 1:1,000; 5114T), microtubule-associated protein 1 light chain 3 B (CST, 1:1,000; 3868S), phosphor-extracellular signal-regulated kinase (p-ERK) (CST, 1:1,000; 4370T), ERK (CST, 1:2,000; 9108S), and glyceraldehyde-3-phosphate dehydrogenase (GAPDH) (CST, 1:2,000; 2118S).

Techniques: Control, Saline, Recombinant

Journal: Diabetes & Metabolism Journal

Article Title: Interleukin 33 Promotes Liver Sinusoidal Endothelial Cell Dysfunction and Hepatic Fibrosis in Diabetic Mice

doi: 10.4093/dmj.2024.0532

Figure Lengend Snippet: Interleukin 33 (IL33) amplifies palmitic acid and high glucose-induced liver sinusoidal endothelial cell (LSEC) dysfunction in vitro. (A) Representative fluorescent images of adhered monocytes (upper), and microscopy images of reactive oxygen species (ROS) in LSEC (lower). (B) Analysis of adherent monocytes (upper) and average fluorescent intensity of ROS (lower) ( n =4) (scale bar, 25 μm). (C) Nitric oxide (NO) levels of culture medium from LSEC administrated with recombinant IL33 in vitro (0, 1, 5, 10, 50, or 100 ng/mL) for 24 and 48 hours in the presence of palmitic acid (PA) plus high glucose (PAHG; 0.2 mM PA and 30 mM glucose) ( n =4). (D) Schematic representation of Transwell coculture system. (E, F) Representative fluorescent images and analysis of alpha smooth muscle actin (αSMA) in LX-2 co-cultured with LSEC ( n =4) (scale bar, 25 μm). (G) Western blot images and quantification of suppression of tumorigenicity 2 (ST2) from L02 cells transfected with small interfering RNA targeting ST2 (siST2) or scramble RNA. (H) Representative fluorescent images of adhered monocytes (upper), and microscopy images of ROS in LSEC (lower) after transfected with siST2 or scramble RNA ( n =3) (scale bar, 25 μm). (I) Representative fluorescent images and analysis of αSMA in LSEC transfected with siST2 or scramble RNA ( n =3) (scale bar, 25 μm). (J) Representative fluorescent images and analysis of αSMA in LX-2 cultured with culture medium collected from LSECs transfected with siST2 or scramble RNA ( n =3) (scale bar, 25 μm). Data was shown as mean±standard error of the mean. NG, normal glucose; NS, no significant; HSC, hepatic stellate cell; DAPI, 4’,6-diamidino-2-phenylindole; wt, wild-type; GAPDH, glyceraldehyde-3-phosphate dehydrogenase. a P <0.05, b P <0.01.

Article Snippet: Primary antibodies targeting IL33 (Proteintech, Rosemont, IL, USA; 1:1,000; 12372-1-AP), ST2 (Proteintech, 1:1,000; 11920-1-AP), P62 (Cell Signaling Technology [CST], Danvers, MA, USA; 1:1,000; 5114T), microtubule-associated protein 1 light chain 3 B (CST, 1:1,000; 3868S), phosphor-extracellular signal-regulated kinase (p-ERK) (CST, 1:1,000; 4370T), ERK (CST, 1:2,000; 9108S), and glyceraldehyde-3-phosphate dehydrogenase (GAPDH) (CST, 1:2,000; 2118S).

Techniques: In Vitro, Microscopy, Recombinant, Cell Culture, Western Blot, Transfection, Small Interfering RNA

Journal: Diabetes & Metabolism Journal

Article Title: Interleukin 33 Promotes Liver Sinusoidal Endothelial Cell Dysfunction and Hepatic Fibrosis in Diabetic Mice

doi: 10.4093/dmj.2024.0532

Figure Lengend Snippet: Interleukin 33 (IL33) drives liver sinusoidal endothelial cell (LSEC) dysfunction by blocking autophagy in vitro. (A) Western blot and measurement for light chain 3 (LC3) and p62 of LSEC ( n =4). (B) Nitric oxide (NO) levels of culture medium from LSEC cultured with different administration ( n =4). (C) Representative fluorescent images of adhered monocytes (upper), and microscopy images of reactive oxygen species (ROS) in LSEC (lower) (scale bar, 25 μm). (D) Analysis of adherent monocytes number (left) ( n =4) and average fluorescent intensity of ROS in LSEC (right) ( n =4). (E, F) Representative fluorescent images and analysis of alpha smooth muscle actin (αSMA) in LX-2 (scale bar, 25 μm). Data was shown as mean±standard error of the mean. PAHG, palmitic acid plus high glucose; Rapa, rapamycin; GAPDH, glyceraldehyde-3-phosphate dehydrogenase; NG, normal glucose; NS, no significant; DAPI, 4’,6-diamidino-2-phenylindole. a P <0.05, b P <0.01.

Article Snippet: Primary antibodies targeting IL33 (Proteintech, Rosemont, IL, USA; 1:1,000; 12372-1-AP), ST2 (Proteintech, 1:1,000; 11920-1-AP), P62 (Cell Signaling Technology [CST], Danvers, MA, USA; 1:1,000; 5114T), microtubule-associated protein 1 light chain 3 B (CST, 1:1,000; 3868S), phosphor-extracellular signal-regulated kinase (p-ERK) (CST, 1:1,000; 4370T), ERK (CST, 1:2,000; 9108S), and glyceraldehyde-3-phosphate dehydrogenase (GAPDH) (CST, 1:2,000; 2118S).

Techniques: Blocking Assay, In Vitro, Western Blot, Cell Culture, Microscopy

Journal: Diabetes & Metabolism Journal

Article Title: Interleukin 33 Promotes Liver Sinusoidal Endothelial Cell Dysfunction and Hepatic Fibrosis in Diabetic Mice

doi: 10.4093/dmj.2024.0532

Figure Lengend Snippet: Interleukin 33 (IL33) elicits liver sinusoidal endothelial cell (LSEC) dysfunction by activating the extracellular signal-regulated kinase 1 (ERK1)/mitogen-activated protein kinase (MAPK) pathway. (A, B) Western blot and measurement for p-ERK, ERK, light chain 3 (LC3), and p62 of LSEC ( n =3). (C) Nitric oxide (NO) levels of culture medium from LSEC cultured with different administration ( n =4). (D) Representative fluorescent images and analysis of adhered monocytes (upper), and microscopy images of reactive oxygen species (ROS) in LSEC (lower) (scale bar, 25 μm). (E) Representative fluorescent images and analysis of alpha smooth muscle actin (αSMA) in LX-2 cultured with culture medium collected from different groups of LSECs ( n =4) (scale bar, 25 μm). (F) Representative fluorescent images and analysis of αSMA in LSEC cultured with different administration ( n =4) (scale bar, 25 μm). Data was shown as mean±standard error of the mean. GAPDH, glyceraldehyde-3-phosphate dehydrogenase; NG, normal glucose; NS, no significant; PAHG, palmitic acid plus high glucose; DAPI, 4’,6-diamidino-2-phenylindole. a P <0.05, b P <0.01.

Article Snippet: Primary antibodies targeting IL33 (Proteintech, Rosemont, IL, USA; 1:1,000; 12372-1-AP), ST2 (Proteintech, 1:1,000; 11920-1-AP), P62 (Cell Signaling Technology [CST], Danvers, MA, USA; 1:1,000; 5114T), microtubule-associated protein 1 light chain 3 B (CST, 1:1,000; 3868S), phosphor-extracellular signal-regulated kinase (p-ERK) (CST, 1:1,000; 4370T), ERK (CST, 1:2,000; 9108S), and glyceraldehyde-3-phosphate dehydrogenase (GAPDH) (CST, 1:2,000; 2118S).

Techniques: Western Blot, Cell Culture, Microscopy