Journal: Nature communications

Article Title: VSIG4 inhibits proinflammatory macrophage activation by reprogramming mitochondrial pyruvate metabolism.

doi: 10.1038/s41467-017-01327-4

Figure Lengend Snippet: Fig. 1 Vsig4−/−mice are more susceptible to HFD-induced obesity with insulin resistance. Eight-week-old male Vsig4−/−mice and age-matched C57BL/6 WT controls were fed a HFD. a Body weight was measured and compared. The obese mice were sacrificed after 10 weeks of HFD feeding. b Fat distribution was detected by μCT. Yellow indicates subcutaneous fat and brown indicates that visceral fat. c Measurement of abdominal wall fat, perirenal fat, serum triglyceride, cholesterol, and free fatty acid. d Representative liver H&E staining (left), and intrahepatic triglyceride contents (right), scale bar = 20 μm, n = 10 per group. e Representative the architecture of adipose tissues stained by H&E (left), adipocyte size and cell numbers was calculated (right), scale bar = 20 μm, n = 10 per group. f The 15-h-fasting blood glucose levels and 5-h-fasting serum insulin levels. g GTT and ITT were performed in theses obese mice, n = 6 per group. h Western blot of the AKT, p-Aktser473, and p-IRS-1 in VAT, muscle and liver tissues of obese mice after 4 min of insulin administration, n = 4 per group. ATMs were isolated from obese mice. i Flow cytometry analyzing pro-IL-1β, IFN-γ, and TNF. Cytokines in VAT were detected by j qRT-PCR and k western blot. Error bar, s.e.m. *p < 0.05, **p < 0.01 and ***p < 0.001 (Student’s t-test). Data are representative of five (a) and three (c–i) independent experiments

Article Snippet: ELISA Kits, including TNF (#EK0527), IL-6 (#EK0411), IL-1β (#EK0394), and IL-12 p40 (#EK0932) were from Boster Ltd. (Wuhan, China).

Techniques: Staining, Western Blot, Isolation, Flow Cytometry, Quantitative RT-PCR

Journal: Nature communications

Article Title: VSIG4 inhibits proinflammatory macrophage activation by reprogramming mitochondrial pyruvate metabolism.

doi: 10.1038/s41467-017-01327-4

Figure Lengend Snippet: Fig. 2 Vsig4 deficiency exacerbates MHV-3-induced fulminant hepatitis. The Vsig4−/−mice and age-matched C57BL/6 WT littermates were infected with MHV-3 (100 PFU/mouse) via i.p. injection. a The survival was monitored. b H&E staining of liver, and TUNEL staining of cell apoptosis, scale bar = 20 μm, n = 5–8 per group, arrow indicated positive cells. c Serum ALT and AST levels at 0 h and 48 h post infection (PI), n = 5–8 per group. d Plaque assay of virus titers in livers at 48 h PI. e qRT-PCR analyzing proinflammatory cytokines in PEMs at 12 h and in liver tissues at 72 h of MHV-3 infection. f Flow cytometry analyzing TNF, pro-IL1-β, and IL-6 from PEMs after 12 h of virus infection. g Western blot analyzing proinflammatory cytokines in infected livers at 24 h and 48 h PI, n = 4 per group. h ELISA of serum concentration of proinflammatory mediators, n = 5–10 per group. Error bar, s.e.m. *p < 0.05, **p < 0.01, ***p < 0.001 and NS, p > 0.05. a was analyzed by log-rank test and others are calculated by Student’s t-test. Data are representative of six (a) and three (b–f, h) independent experiments

Article Snippet: ELISA Kits, including TNF (#EK0527), IL-6 (#EK0411), IL-1β (#EK0394), and IL-12 p40 (#EK0932) were from Boster Ltd. (Wuhan, China).

Techniques: Infection, Injection, Staining, TUNEL Assay, Plaque Assay, Virus, Quantitative RT-PCR, Flow Cytometry, Western Blot, Enzyme-linked Immunosorbent Assay, Concentration Assay

Journal: Nature communications

Article Title: VSIG4 inhibits proinflammatory macrophage activation by reprogramming mitochondrial pyruvate metabolism.

doi: 10.1038/s41467-017-01327-4

Figure Lengend Snippet: Fig. 3 VSIG4 impedes LPS-induced macrophage M1 polarization in vitro. PEMs were treated with LPS (2 μg/ml), a qRT-PCR analysis of Il-1β and Tnf transcripts. b ELISA of cytokines in cultured supernatants. c Western blot analyzing cytokine protein expression. d Flow cytometry analyzing surface expression of activation markers. RAW264.7 cells stably infected with lentiviral control vectors (Len-cont.) or vectors encoding Vsig4 (Len-Vsig4), cells were further treated with LPS (2 μg/ml), e qRT-PCR analysis of Il-1β, Il-6, and Tnf transcripts. f ELISA detecting cytokines in cultured supernatant. g Flow cytometry analyzing surface expression of CD40. h C3−/−BMDMs were tranfected to overexpress VSIG4, and cells were further treated with LPS (2 μg/ ml), the secretion of IL-6 and IL-1β was detected by ELISA. Error bar, s.e.m. *p < 0.05, **p < 0.01, ***p < 0.001 and NS, p > 0.05 (Student’s t-test). Data are representative of three independent experiments

Article Snippet: ELISA Kits, including TNF (#EK0527), IL-6 (#EK0411), IL-1β (#EK0394), and IL-12 p40 (#EK0932) were from Boster Ltd. (Wuhan, China).

Techniques: In Vitro, Quantitative RT-PCR, Enzyme-linked Immunosorbent Assay, Cell Culture, Western Blot, Expressing, Flow Cytometry, Activation Assay, Stable Transfection, Infection, Control

Journal: Nature communications

Article Title: VSIG4 inhibits proinflammatory macrophage activation by reprogramming mitochondrial pyruvate metabolism.

doi: 10.1038/s41467-017-01327-4

Figure Lengend Snippet: Fig. 5 VSIG4 triggers PDK2 expression in macrophages. Macrophages from WT and Vsig4−/−mice were collected. a qRT-PCR detection of 4 Pdk isoforms in BMDMs. b Western blot analyzing PDK2, p-PDH-E1αs300, p-PDH-E1αs293, and total PDH. c The location of p-PDH-E1αs300 in mitochondria was analyzed by immunofluoresence double staining, scale bar = 20μm. d Western blot of PDK2, p-PDH-E1αs300, p-PDH-E1αs293 in liver tissues at 0 h and 48 h PI. RAW264.7 cells were transfected to expression of Vsig4, and cells were further treated with LPS (2 μg/ml), e Western blot analysis of PDK2 and p-PDH- E1αs300. f PDH activity analysis, n = 6 per group. The expression of Pdk2 in RAW264.7 cells was silenced by shRNA or enhancing Pdk2 expression by lentivirus infection. g Seahorse analysis of OCR after 2 h of LPS treatment (up), and basal and maximal OCR of the indicated conditions was plotted in bar graphs (down), n = 5 per group. h Flow cytometric assay of mtROS secretion after LPS administration. i ELISA of IL-6 and TNF in cultured supernatants, n = 4 per group. j Flow cytometric assay of LPS-caused CD40 expression at 6 h. Error bar, s.e.m. *p < 0.05,**p < 0.01, ***p < 0.001 and NS, p > 0.05 (Student’s t-test). Data are representative of three independent experiments

Article Snippet: ELISA Kits, including TNF (#EK0527), IL-6 (#EK0411), IL-1β (#EK0394), and IL-12 p40 (#EK0932) were from Boster Ltd. (Wuhan, China).

Techniques: Expressing, Quantitative RT-PCR, Western Blot, Double Staining, Transfection, Activity Assay, shRNA, Infection, Flow Cytometry, Enzyme-linked Immunosorbent Assay, Cell Culture

Journal: Nature communications

Article Title: VSIG4 inhibits proinflammatory macrophage activation by reprogramming mitochondrial pyruvate metabolism.

doi: 10.1038/s41467-017-01327-4

Figure Lengend Snippet: Fig. 8 Forced overexpression of Vsig4 improves MHV-3-induced hepatitis. C57BL/6 WT mice were infected with lentivirus (107 PFU/mouse) to induce the expression of Vsig4 in vivo, these mice were further infected with MHV-3 at day 6. a Liver Vsig4 gene transcription was analyzed by qRT-PCR at day 6, n = 5 per group. b Western blotting for PDK2, p-PDH-E1αs300, FGL2, and proinflammatory cytokines TNF, IL-6 and IL-1β in liver tissues at 72 h of MHV-3 infection, n = 4 per group. c The architecture of the liver tissues at 72 h of infection was compared by H&E staining, scale bar = 20 μm, n = 5 per group. d The survival was monitored for a total of 20 days. Error bar, s.e.m. a *p < 0.05 was analyzed by Student’s t-test, and d was analyzed by log-rank test. Data are representative of three independent experiments

Article Snippet: ELISA Kits, including TNF (#EK0527), IL-6 (#EK0411), IL-1β (#EK0394), and IL-12 p40 (#EK0932) were from Boster Ltd. (Wuhan, China).

Techniques: Over Expression, Infection, Expressing, In Vivo, Quantitative RT-PCR, Western Blot, Staining

Journal: Journal of the American Heart Association: Cardiovascular and Cerebrovascular Disease

Article Title: Reduced Plasma Kallistatin Is Associated With the Severity of Coronary Artery Disease, and Kallistatin Treatment Attenuates Atherosclerotic Plaque Formation in Mice

doi: 10.1161/JAHA.118.009562

Figure Lengend Snippet: Kallistatin inhibited low‐shear stress–induced carotid artery plaque formation. A , Representative magnetic resonance images in cross‐sectional views through the carotid arteries. The red arrow indicates the cross section of the left carotid artery, and the blue arrow indicates the cross section of the right carotid artery. B , Quantitative analysis of left carotid artery diameter in the Ad. HKS and Ad.Null groups. L‐ NAME and NAM blocked the effects of kallistatin (n=10, * P <0.05 vs the Ad.Null‐, L‐ NAME ‐, or NAM ‐treated groups). C , Quantitative analysis of left carotid artery plaque volume in the Ad. HKS and Ad.Null groups. L‐ NAME and NAM blocked the effect of kallistatin; n=10 (* P <0.05 vs the Ad.Null‐, L‐ NAME ‐ or NAM ‐treated groups). D , Plasma MDA levels in apoE −/− mice. E , Plasma TNF ‐α levels in apoE −/− mice. F , Distribution of mouse kallistatin expression in atherosclerotic lesions and normal vascular tissue. Data are presented as the mean± SEM ; n=10, * P <0.05 vs the Ad.Null‐, L‐ NAME ‐, or NAM ‐treated groups. Ad.HKS indicates adenovirus containing kallistatin cDNA; Ad.Null, adenovirus containing null cDNA; L‐NAME, N ω ‐nitro‐L‐arginine methyl ester; MDA, malondialdehyde; NAM, nicotinamide; SEM, standard error of the mean; TNF‐α, tumor necrosis factor‐α.

Article Snippet: Plasma samples were used for the analysis of TNF‐α using a Mouse TNF‐α ELISA (Proteintech, Rosemount, IL) according to the manufacturer's protocol.

Techniques: Shear, Clinical Proteomics, Expressing

Journal: Journal of the American Heart Association: Cardiovascular and Cerebrovascular Disease

Article Title: Reduced Plasma Kallistatin Is Associated With the Severity of Coronary Artery Disease, and Kallistatin Treatment Attenuates Atherosclerotic Plaque Formation in Mice

doi: 10.1161/JAHA.118.009562

Figure Lengend Snippet: Kallistatin led to morphological changes in the atherosclerotic plaques of apoE −/− mice. A , Representative images of HE staining. Original magnification is ×10 (n=5 in each group). B , Oil red O staining of carotid specimens of mice in the Ad.Null group, the Ad. HKS group, and L‐ NAME ‐ or NAM ‐treated groups (n=5 in each group). C , Representative images of superoxide formation labeled by the red fluorescence dye dihydroethidium in the carotid artery of Ad.Null mice, Ad. HKS mice, Ad. HKS +L‐ NAME mice, and Ad. HKS + NAM mice (n=5 in each group). D , Representative images of CD 68 immunostaining in lesion areas of the left carotid artery (n=5 in each group). E , Immunohistochemical assessments of the protein level of TNF ‐α in carotid plaque (n=5 in each group). F, Quantitative analyses of CD 68‐positive cells in the 4 groups (n=5 in each group, * P <0.05 vs the Ad.Null‐, L‐ NAME ‐, or NAM ‐treated groups). G , Fluorescence intensity was measured by a fluorescence microscope and quantified with Image software (n=5 in each group, * P <0.05 vs the Ad.Null‐, L‐ NAME ‐, or NAM ‐treated groups). H , Colocalization of PE red fluorescence ( SIRT 1) and FITC green fluorescence ( CD 31) within plaques (×40 magnification) in the carotid arteries of the Ad.Null, Ad. HKS , Ad. HKS +L‐ NAME , and Ad. HKS + NAM groups with labeled cell nuclei (blue) (n=5 in each group). Regions shown at a higher magnification (far right) are indicated by yellow boxes. Endothelial cell‐specific SIRT 1 fluorescence was measured on the luminal side of the internal elastic lamina only and expressed in mean pixel intensity per area (n=5 in each group, # P <0.05 vs the Ad.Null‐ and NAM ‐treated groups). I , Colocalization of PE red fluorescence ( eNOS ) and FITC green fluorescence ( CD 31) within carotid artery plaques (×40 magnification) of the 4 groups with labeled cell nuclei (blue). Regions shown at a higher magnification (far right) are indicated by yellow boxes. Endothelial cell‐specific eNOS fluorescence was measured on the luminal side of the internal elastic lamina only and expressed in mean pixel intensity per area (* P <0.05 vs the Ad.Null‐, L‐ NAME ‐, or NAM ‐treated groups). We observed that red fluorescence ( SIRT 1, eNOS ) and green fluorescence ( CD 31) signals were colocalized in the Ad. HKS group. Scale bar=50 μm. Ad.HKS indicates adenovirus containing kallistatin cDNA; Ad.Null, adenovirus containing null cDNA; DAPI, 4′,6‐diamidino‐2‐phenylindole; DHE, dihydroethidium; eNOS, endothelial nitric oxide synthase; FITC, fluoroisothiocyanate; HE, hematoxylin and eosin; L‐NAME, N ω ‐nitro‐L‐arginine methyl ester; NAM, nicotinamide; PE, phycoerythrin; SIRT1, sirtuin 1; TNF‐α, tumor necrosis factor‐α.

Article Snippet: Plasma samples were used for the analysis of TNF‐α using a Mouse TNF‐α ELISA (Proteintech, Rosemount, IL) according to the manufacturer's protocol.

Techniques: Staining, Labeling, Fluorescence, Immunostaining, Immunohistochemical staining, Microscopy, Software

Journal: Journal of the American Heart Association: Cardiovascular and Cerebrovascular Disease

Article Title: Reduced Plasma Kallistatin Is Associated With the Severity of Coronary Artery Disease, and Kallistatin Treatment Attenuates Atherosclerotic Plaque Formation in Mice

doi: 10.1161/JAHA.118.009562

Figure Lengend Snippet: Effect of kallistatin protein ( KS ) on TNF ‐α–induced HUVEC s. NADPH oxidase activity ( A ), intracellular ROS accumulation ( B ), relative SIRT 1 mRNA expression levels ( C ), relative eNOS mRNA expression levels ( D ), and SIRT 1 and eNOS protein expression ( E ). Data are presented as the mean± SEM (n=5, * P <0.05 vs the TNF ‐α, TNF ‐α+ KS + NAM , and TNF ‐α+ KS + L‐ NAME groups. # P <0.05 vs the TNF ‐α and TNF ‐α+ KS + NAM groups). eNOS indicates endothelial nitric oxide synthase; HUVEC, human umbilical vein endothelial cells; KS, kallistatin; L‐NAME, N ω ‐nitro‐L‐arginine methyl ester; NAM, nicotinamide; ROS, reactive oxygen species; SIRT1, sirtuin 1; TNF‐α, tumor necrosis factor‐α.

Article Snippet: Plasma samples were used for the analysis of TNF‐α using a Mouse TNF‐α ELISA (Proteintech, Rosemount, IL) according to the manufacturer's protocol.

Techniques: Activity Assay, Expressing

Journal: Journal of Nanobiotechnology

Article Title: HA-coated collagen nanofibers for urethral regeneration via in situ polarization of M2 macrophages

doi: 10.1186/s12951-021-01000-5

Figure Lengend Snippet: The polarization of macrophages to M2 phenotype is related to the elongated cell shape. A Fluorescence micrographs of Raw 264.7 macrophages immune-stained for arginase-1 (green), iNOS (red), and nuclear counterstain (blue) on cell plate (control), collagen and HA-collagen nanofibrous films. Scale bars: 15 μm. B Representative Western blot of arginase-1, iNOS, and tubulin of control, collagen and HA-collagen nanofibrous films and quantification of average across three separate experiments. Quantified TNF-α C and IL-10 D secretion from macrophages cultured on different nanofibrous scaffolds or culture plates using ELISA assay. n = 3, ** p < 0.01, *** p < 0.001

Article Snippet: The supernatant was collected, centrifuged, and used for ELISA after 24, 48, and 72 h. According to the manufacturer’s protocol, TNF-α and IL-10 were quantified by ELISA Kit (FEK0527, EK0417, Boster).

Techniques: Fluorescence, Staining, Control, Western Blot, Cell Culture, Enzyme-linked Immunosorbent Assay

Journal: European Journal of Medicinal Chemistry Reports

Article Title: Kolaviron ameliorates chronic colitis induced by prolonged oral administration of Dextran Sulphate Sodium in Balb/c mice

doi: 10.1016/j.ejmcr.2022.100071

Figure Lengend Snippet: Fig. 3. Effect of Kolaviron (KOL) on pro-inflammatory proteins in Colitis Chronicity. Tumor Necrosis Factor Alpha (TNF-α) (A) Interleukin-1β (IL-1 β) (B) Inducible Nitric Oxide Synthase (iNOS) (C) Cyclooxygenase-2 (COX-2) (D). Each bar represents mean SD of ten mice. a: Values differ significantly from control (p < 0.05). b: Values differ significantly from KOL þ DSS (p < 0.05).

Article Snippet: Anti-iNOS and Anti-COX-2 mice monoclonal antibodies, tumor necrosis factor-alpha (TNF-α), and interleukin-1β (IL-1β), ELISA kits and were purchased from CUSABIO Life Science Inc. (Wuhan, China).

Techniques: Control