Journal: Gut Pathogens

Article Title: Effect of crude polysaccharide from seaweed, Dictyopteris divaricata (CDDP) on gut microbiota restoration and anti-diabetic activity in streptozotocin (STZ)-induced T1DM mice

doi: 10.1186/s13099-022-00512-1

Figure Lengend Snippet: Crude polysaccharide (CDDP) enhanced the expression of: A Mucin-2 (MUC-2) and B Zonula occludens-1 (ZO-1) in STZ-induced T1DM mice colon by immunohistochemistry technique after 4-weeks treatment. Inflammatory cells (demonstrated by red arrow), goblet cells (demonstrated by yellow arrow). Mucin expression (demonstrated by green arrow) and Zonula occludens-1 expression (demonstrated by orange arrow) were shown. The picture is the representative of colon of six different treatment groups. Original magnification: 20x, scale bar: 100 µm

Article Snippet: The primary antibodies [goat antirabbit: β-actin, Mucin-2 (MUC-2), Claudin-1, Occludin, Zonula occludens-1 (ZO-1) and insulin receptor substrate-1 (IRS-1)], secondary antibodies and the radioimmunoprecipitation assay (RIPA) buffer were purchased from Proteintech (Wuhan, China).

Techniques: Expressing, Immunohistochemistry

Journal: Gut Pathogens

Article Title: Effect of crude polysaccharide from seaweed, Dictyopteris divaricata (CDDP) on gut microbiota restoration and anti-diabetic activity in streptozotocin (STZ)-induced T1DM mice

doi: 10.1186/s13099-022-00512-1

Figure Lengend Snippet: Effect of CDDP treatment on tight junction proteins (TJs) expression in the colon epithelium. A The relative expression of Occludin and Claudin-1 in STZ-induced T1DM mice were analyzed by western blotting technique after 4-weeks treatment using β-actin as an internal control. Western blots are the representative of three experiments. B Bar graph represent the intensity of relative protein band against β-actin as an internal control, quantified by NIH image J software. Data obtained from three experiments are presented as Mean ± SEM (n = 6). * p < 0.05, ** p < 0.01, *** p < 0.001 and **** p < 0.001, model group compared with control group and CDDP and metformin treatment groups compared with model group

Article Snippet: The primary antibodies [goat antirabbit: β-actin, Mucin-2 (MUC-2), Claudin-1, Occludin, Zonula occludens-1 (ZO-1) and insulin receptor substrate-1 (IRS-1)], secondary antibodies and the radioimmunoprecipitation assay (RIPA) buffer were purchased from Proteintech (Wuhan, China).

Techniques: Expressing, Western Blot, Control, Software

Journal: Cellular and Molecular Gastroenterology and Hepatology

Article Title: Enhanced Utilization of Induced Pluripotent Stem Cell–Derived Human Intestinal Organoids Using Microengineered Chips

doi: 10.1016/j.jcmgh.2017.12.008

Figure Lengend Snippet: Table of Antibodies Used in Study

Article Snippet: Mucin-2 (CCP58) , 1:500 , NBP2-25221 , Mouse IgG1 , Novus Biologicals.

Techniques:

Journal: Journal of Oral Microbiology

Article Title: Aggregatibacter actinomycetemcomitans exacerbates colitis and perturbs the gut microbiota in a murine model​

doi: 10.1080/20002297.2026.2613536

Figure Lengend Snippet: Oral administration of A. actinomycetemcomitans was associated with aggravated intestinal barrier impairment and elevated levels of inflammatory mediators in mice with colitis. (A) Relative mRNA expression levels of pro-inflammatory cytokines in colon tissue were normalized to Gapdh (2 -ΔΔCt method). (B) Serum concentrations of IL-6, IL-1β, and TNF- α measured by ELISA. (C) Representative immunofluorescence images of colon sections stained for the tight junction protein ZO-1 (green) and the mucin MUC-2 (red). Nuclei were counterstained with DAPI (blue). Scale bars: 200 μm. Quantification of ZO-1 and MUC-2 signal intensity was performed on at least 9 non-overlapping fields per mouse using ImageJ software. Data are representative of two independent experiments. Data are shown as mean ± SEM. Statistical analysis was performed using two-sided one-way ANOVA followed by the least significant difference (LSD) post–hoc test or two-sided unpaired Student’s t-test and Dunnett T3 (uneven variance). * p < 0.05, ** p < 0.01, *** p < 0.001.

Article Snippet: The ZO-1 and MUC-2 protein-targeting rabbit antibodies (Servicebio, China) were applied to the 4 μm-thick tissue sections, which were subsequently incubated overnight at 4 °C, washed with phosphate-buffered saline, and then treated with anti-rabbit fluorescent secondary antibodies (Servicebio, China) at 37 °C for 1 h. Following washing, the tissue nuclei were re-stained with 4-methyl-6-methyl-2-phenylindole (DAPI).

Techniques: Expressing, Enzyme-linked Immunosorbent Assay, Immunofluorescence, Staining, Software

Journal: Scientific Reports

Article Title: Prevention of Shiga toxin 1-caused colon injury by plant-derived recombinant IgA

doi: 10.1038/s41598-022-22851-4

Figure Lengend Snippet: Activation of caspase-3 and depletion of secretory granules in the colon epithelium upon intrarectal exposure to Stx1. Frozen sections ( a,c ) or paraffin sections ( b ) of colon tissue from Stx1-exposed or control (PBS) mice were stained with anti-activated caspase-3 antibodies (red in a ), Alcian blue dye (blue in b ), or anti-MUC2 antibodies (red in c ). Nuclei were visualized by staining with DAPI (blue in a,c ), and with hematoxylin (violet in b ). Arrows indicate signals from activated caspase-3 ( a ). Fixation of tissues was performed with formalin-PBS ( a , c ) or Carnoy’s fixative ( b ). Bars represent 50 μm. Each mouse was exposed to 100 ng of Stx1. ( d ) Quantification of secretory granules in the colon tissues in response to the amount of Stx1 (abscissa) exposed to each mouse. The ordinate indicates the percentage of the MUC2-positive area divided by the total area of the mucous epithelium (mean ± SEM) from four independent mice. There were no exclusions of data points.

Article Snippet: Restriction enzymes Nde I, Sac I-HF, Not I-HF, Sac II and Nsi I-HF were purchased from New England BioLabs (Ipswich, MA, USA); Sma I, Kpn I, Sal I, binary vector pRI201-AN, Klenow fragment, a DNA Ligation Kit and a DNA Ligation Kit from Takara Bio (Shiga, Japan); Hin dIII from Nippon Gene (Tokyo, Japan); a protease inhibitor cocktail for plant cell and tissue extracts, myeloma proteins TEPC 15 (mouse IgA, κ) and MOPC 21 (mouse IgG1, κ), and Alcian Blue 8GX from Sigma-Aldrich (St. Louis, MO, USA); 2,2'-Azino-bis (3-ethylbenzothiazoline-6-sulfonic acid) (ABTS), Mayer's Hematoxylin Solution, Murashige and Skoog (MS) plant salt mixture, and gellan gum from Wako Pure Chemical Industries, Ltd. (Osaka, Japan); Vero cells (an African Green Monkey kidney-derived cell line) from the American Type Culture Collection (Manassas, VA, USA); fetal bovine serum (FBS) from Hyclone (South Logan, UT, USA); a Cell Counting Kit-8 from DOJINDO (Kumamoto, Japan); 4,6-Diamidino-2-phenylindole Dihydrochloride (DAPI) from Nacalai Tesque (Kyoto, Japan); a PierceTM BCA Protein Assay Kit, HiMarkTM Pre-Stained Protein Standards, MagicMarkTM XP Western Protein Standards, SuperSignalTM West Pico PLUS Chemiluminescent Substrate, Medium 199 and Alexa594-goat anti-rabbit IgG from Thermo Fisher Scientific (Waltham, MA, USA); goat anti-mouse κ, horseradish peroxidase (HRP)-goat anti-mouse IgA, HRP-goat anti-mouse IgG, HRP-donkey anti-goat IgG and HRP-goat anti-rabbit IgG from Southern Biotech (Birmingham, AL, USA); rabbit anti-mouse J chain from Proteintech (Wuhan, China); rabbit anti-human/mouse activated-caspase 3 from BD Biosciences (Franklin Lakes, NJ, USA); and rabbit anti-mouse MUC2 from Santa Cruz Biotechnology (Santa Cruz, CA, USA).

Techniques: Activation Assay, Control, Staining

Journal: Scientific Reports

Article Title: Prevention of Shiga toxin 1-caused colon injury by plant-derived recombinant IgA

doi: 10.1038/s41598-022-22851-4

Figure Lengend Snippet: Prevention of Stx1-caused colon injury by an IgA plantibody specific for Stx1. Sixty ng of Stx1 was treated with the leaf extract of an IgA transgenic plant (Stx1 + IgA-Tg) containing 8 μg IgA (1.6 mg total proteins), that of a wild-type plant (Stx1 + WT) containing 1.6 mg of total protein, or PBS (indicated as Stx1 only) for 1 h. Mice were then exposed to each sample containing Stx1 or to PBS alone (indicated as PBS) through intrarectal administration. After 16 h, colon tissues were collected and embedded in O.C.T. compound. Frozen sections of the colon tissues were stained with anti-activated caspase-3 ( a ) or anti-MUC2 ( b,c ) antibodies. Nuclei were counterstained with DAPI ( a,b ). Arrows indicate signals from activated caspase-3 ( a ). Bars represent 50 μm. The percentages of the MUC2-positive area divided by the total area of the mucous epithelium are plotted ( c ). Horizontal bars represent the means for individual groups and error bars are SD ( n = 4). There were no exclusions of data points. Statistical significance was analyzed by ANOVA followed by the Tukey test. ** P < 0.01, **** P < 0.0001, ns P > 0.05.

Article Snippet: Restriction enzymes Nde I, Sac I-HF, Not I-HF, Sac II and Nsi I-HF were purchased from New England BioLabs (Ipswich, MA, USA); Sma I, Kpn I, Sal I, binary vector pRI201-AN, Klenow fragment, a DNA Ligation Kit and a DNA Ligation Kit from Takara Bio (Shiga, Japan); Hin dIII from Nippon Gene (Tokyo, Japan); a protease inhibitor cocktail for plant cell and tissue extracts, myeloma proteins TEPC 15 (mouse IgA, κ) and MOPC 21 (mouse IgG1, κ), and Alcian Blue 8GX from Sigma-Aldrich (St. Louis, MO, USA); 2,2'-Azino-bis (3-ethylbenzothiazoline-6-sulfonic acid) (ABTS), Mayer's Hematoxylin Solution, Murashige and Skoog (MS) plant salt mixture, and gellan gum from Wako Pure Chemical Industries, Ltd. (Osaka, Japan); Vero cells (an African Green Monkey kidney-derived cell line) from the American Type Culture Collection (Manassas, VA, USA); fetal bovine serum (FBS) from Hyclone (South Logan, UT, USA); a Cell Counting Kit-8 from DOJINDO (Kumamoto, Japan); 4,6-Diamidino-2-phenylindole Dihydrochloride (DAPI) from Nacalai Tesque (Kyoto, Japan); a PierceTM BCA Protein Assay Kit, HiMarkTM Pre-Stained Protein Standards, MagicMarkTM XP Western Protein Standards, SuperSignalTM West Pico PLUS Chemiluminescent Substrate, Medium 199 and Alexa594-goat anti-rabbit IgG from Thermo Fisher Scientific (Waltham, MA, USA); goat anti-mouse κ, horseradish peroxidase (HRP)-goat anti-mouse IgA, HRP-goat anti-mouse IgG, HRP-donkey anti-goat IgG and HRP-goat anti-rabbit IgG from Southern Biotech (Birmingham, AL, USA); rabbit anti-mouse J chain from Proteintech (Wuhan, China); rabbit anti-human/mouse activated-caspase 3 from BD Biosciences (Franklin Lakes, NJ, USA); and rabbit anti-mouse MUC2 from Santa Cruz Biotechnology (Santa Cruz, CA, USA).

Techniques: Transgenic Assay, Staining

Journal: Molecular Medicine

Article Title: Evaluation of two laboratory model methods for diarrheal irritable bowel syndrome

doi: 10.1186/s10020-022-00599-x

Figure Lengend Snippet: The primers used in this experiment

Article Snippet: Antibodies zonulae occluden-1 (ZO-1) and mucin 2 (Muc2) were purchased from Boster Bioengineering Co., LTD. (PB9234, BM5029).

Techniques: Sequencing

Journal: Molecular Medicine

Article Title: Evaluation of two laboratory model methods for diarrheal irritable bowel syndrome

doi: 10.1186/s10020-022-00599-x

Figure Lengend Snippet: Intensity of ZO-1 and MUC2 (green) in ileal and colonic tissues of rats in each group. Nuclei stained with DAPI (blue) (200×) ( A ), mRNA expression levels of ZO-1 ( B ), MUC2 ( C ), OCLN ( D ), CLDN4 ( E ) in each group of rats.* p < 0.05, ** p < 0.01, *** p < 0.001

Article Snippet: Antibodies zonulae occluden-1 (ZO-1) and mucin 2 (Muc2) were purchased from Boster Bioengineering Co., LTD. (PB9234, BM5029).

Techniques: Staining, Expressing