Journal: Scientific Reports

Article Title: Expression of FABP4, adipsin and adiponectin in Paneth cells is modulated by gut Lactobacillus

doi: 10.1038/srep18588

Figure Lengend Snippet: ( a ) Schematic representation (right) and staining (left) of the Paneth cells and putative intestinal stem cells. Paneth cells (lysozyme positive) and putative intestinal stem cells (LGR5 positive) localized at the crypt base. Intestinal tissues were sliced and stained using pooled isotypic control (NC) with FITC-labeled (NC-FITC) or PE-labeled (NC-PE) secondary antibodies, anti-lysozyme (lysozyme) and anti-LGR5 (LGR5) antibodies followed by fluorescence-labeled second antibodies. Green indicated lysozyme or LGR5 positive. Nuclei were stained by DAPI (blue). ( b ) Immunostaining of FABP4, adipsin, adiponectin (adipon) and FABP2 in ileum section from WT mice. Red and green indicated respectively FABP4, adipsin or adiponectin (adipon) and FABP2. Scale bar, 20μm. ( c ) Double immunostaining of FABP4, adipsin and adiponectin (adipon) with lysozyme in ileum section from WT mice. Scale bar, 40μm. ( d ) Analysis of confocal microscope of FABP4, adipsin and adiponectin (adipon) with lysozyme in ileum section from WT mice after double staining. Scale bar, 5 μm. ( e ) Double immuno-staining of adipsin, adiponectin and FABP4 with CD24 in the colon fragments of mice. Colon tissues were sliced and stained using anti-adipsin, anti-adiponectin and anti-FABP4 with anti-CD24 antibodies followed by fluorescence-labeled second antibodies. Green indicated CD24; Red indicated adipsin, adiponectin or FABP4. Nuclei were stained by DAPI (blue). Scale bar, 40 μm.

Article Snippet: Commercial kits for FABP4 (CY-8077, CycLex), adipsine (KA3822, Abnova) and adiponectin (A05187, bertinpharma) were used to quantitate mouse serum FABP4, adipsin and adiponecin.

Techniques: Staining, Control, Labeling, Fluorescence, Immunostaining, Double Immunostaining, Microscopy, Double Staining

Journal: Scientific Reports

Article Title: Expression of FABP4, adipsin and adiponectin in Paneth cells is modulated by gut Lactobacillus

doi: 10.1038/srep18588

Figure Lengend Snippet: ( a,b ) qRT-PCR ( a ) and immunoblot analyses ( b ) of intestinal crypts with and without dithizone treatments. The mice were intravenously injected by dithizone and killed at the indicated time (male, n = 6 or indicated number). R. E., relative expression. ( c ) Serum levels of FABP4, adipsin and adiponectin in mice with (6h, 12h and 6 days after dithizone) and without (0h) dithizone treatments. ( d ) Serum levels of glucose (Glu), triglyceride (TG) and cholesterol (CHO) in fasting mice with (6h, 12h and 6 days after dithizone) and without (0h) dithizone treatments. * P < 0.05 and ** P < 0.01 ( t -test, mean ± SD in a; Mann-Whitney U test in c,d). The data were a representative of three independent experiments.

Article Snippet: Commercial kits for FABP4 (CY-8077, CycLex), adipsine (KA3822, Abnova) and adiponectin (A05187, bertinpharma) were used to quantitate mouse serum FABP4, adipsin and adiponecin.

Techniques: Quantitative RT-PCR, Western Blot, Injection, Expressing, MANN-WHITNEY

Journal: Scientific Reports

Article Title: Expression of FABP4, adipsin and adiponectin in Paneth cells is modulated by gut Lactobacillus

doi: 10.1038/srep18588

Figure Lengend Snippet: ( a ) qRT-PCR and immunoblot analyses of epithelial crypts in ileum fragments of germ-free (GF) mice (n = 10 (male, 5; female, 5)) and feces transplanted GF mice (GF + feces, n = 10 (male (M), 5; female (F), 5)). Actin, a loading control, was detected by anti-β-actin antibody. R. E., relative expression. ( b ) Immuno-staining of ileum fragments of germ-free (GF) mice and GF mice transplanted with WT mouse feces (GF + feces). Red and green indicated respectively FABP4, adipsin or adiponectin (adipon) and FABP2. Scale bar, 40 μm. The result is a representative of 6 samples. ( c ) RT-PCR and immunoblot analyses of ileum epithelial crypts in mice after in vivo transplantation with (Lac, N = 10 (male, 5; female, 5)) or without (NC, N = 10 (male, 5; female, 5)) Lacbacillus NK6. Actin, a loading control. R. E., relative expression. ( d ) qRT-PCR and immunoblot analyses of ileum epithelial crypts after in vitro stimulation with (Lac) or without (NC) Lacbacillus NK6. Actin, a loading control. ( e ) Serum levels of FABP4, adipsin and adiponectin (adipon) after transplantation with ( WT + Lac, N = 10 (male (M), 5; female ( F ), 5)) or without ( WT , N = 10 (male (M), 5; female (F), 5)) Lacbacillus NK6. ( f ) Serum levels of glucose (Glu), triglyceride (TG) and cholesterol (CHO) in fasting mice after transplantation with ( WT + Lac, N = 10 (male (M), 5; female(F), 5)) or without ( WT , N = 10 (male(M), 5; female(F), 5)) Lacbacillus NK6. * P < 0.05 and ** P < 0.01 ( t -test, mean ± SD in a, c, d; Mann-Whitney U test in e, f). The data were representative of three independent experiments.

Article Snippet: Commercial kits for FABP4 (CY-8077, CycLex), adipsine (KA3822, Abnova) and adiponectin (A05187, bertinpharma) were used to quantitate mouse serum FABP4, adipsin and adiponecin.

Techniques: Quantitative RT-PCR, Western Blot, Control, Expressing, Immunostaining, Reverse Transcription Polymerase Chain Reaction, In Vivo, Transplantation Assay, In Vitro, MANN-WHITNEY

Journal: Scientific Reports

Article Title: Expression of FABP4, adipsin and adiponectin in Paneth cells is modulated by gut Lactobacillus

doi: 10.1038/srep18588

Figure Lengend Snippet: ( a ) qRT-PCR (upper) and immunoblot (lower) of FABP4, adipsin, adiponectin and FABP2 in the jejunum epithelial crypt cells of WT and TLR2 deficient (−/−) mice (n = 6). R. E., relative expression. ( b ) qRT-PCR (upper) and immunoblot (lower) of FABP4, adipsin, adiponectin and FABP2 in the ileum epithelial crypt cells of WT and NF-κB deficient (−/−) mice (n = 6). R. E., relative expression. ( c ) qRT-PCR and immunoblot of FABP4, adipsin, adiponectin and FABP2 in the ileum epithelial crypts of WT , TLR2 and NF-κB deficient (−/−) mice with (Lac) or without (NC) Lacbacillus NK6 stimulation. R. E., relative expression. ( d ) Phosphorylation analyses of p65, p38, JNK, and ERK in the intestinal crypts after exposed to Lacbacillus NK6. Murine intestinal crypts were stimulated with Lacbacillus NK6 and lysed at the indicated time. Phosphor-p65, p38, -ERK and -JNK were detected using anti-phosphor-p-65, p38, -ERK or –JNK antibodies. Actin, a loading control, was detected by anti-β-actin antibody. ( e ) Analyses of TRAF2 ubiquitination in intestinal crypt cells after exposed to Lacbacillus NK6. Immunoblot analysis of ubiqitinated TRAF2 and TRAF2 immunoprecipitated from intestinal crypt cells stimulated with Lacbacillus NK6 at the indicated time (upper blots), and TRAF2, IκBα and actin (below blot) in the same cells without immunoprecipitation. IP, immunoprecipitation; IB, immunoblot assay. ( f ) Analyses of TRAF6 ubiquitination in intestinal crypt cells after exposed to Lacbacillus NK6. Immunoblot analysis of K63-linked ubiquitination (K63-Ub) (middle blot) of endogenouse TRAF6 immunoprecipitated from intestinal crypts stimulated with Lacbacillus NK6, and immunoblot analysis of Ub-TRAF6 (top blot), TRAF6, IkBα and actin (below blots) in the same cells without immunoprecipitation. PolyUbi., polyubiquitination; IP, immunoprecipitation; IB, immunoblot assay. * P < 0.05 and ** P < 0.01 ( t -test in a, b, c, mean ± SD). The data were at least a representative of three independent experiments.

Article Snippet: Commercial kits for FABP4 (CY-8077, CycLex), adipsine (KA3822, Abnova) and adiponectin (A05187, bertinpharma) were used to quantitate mouse serum FABP4, adipsin and adiponecin.

Techniques: Quantitative RT-PCR, Western Blot, Expressing, Phospho-proteomics, Control, Ubiquitin Proteomics, Immunoprecipitation