Journal: Pediatric research

Article Title: Galectin-3 Expression and Effect of Supplementation in Neonatal Mice with Disseminated Candida albicans Infection

doi: 10.1038/s41390-019-0279-x

Figure Lengend Snippet: Tissue fungal burden and galectin-3 concentrations in adult mice with disseminated candidiasis. Mice (n=7) were infected via tail-vein injection with C. albicans and euthanized at 48 h after injection. Panel A: Tissue fungal burden. Panel B: Mean gal3 concentration in tissue homogenates compared to animals receiving saline (n=10). Error bars represent standard deviation. No differences in tissue gal3 concentration were detected by ANOVA with inter-group comparisons by the Student-Newman-Keuls Method.

Article Snippet: In a subsequent experiment, pups were delivered and randomized to receive 5 μg carrier-free recombinant mouse gal3 (R&D Systems, Minneapolis, MN) or saline in 20 μl i.p. injection prior to infection with C. albicans as described above.

Techniques: Infection, Injection, Concentration Assay, Saline, Standard Deviation

Journal: Pediatric research

Article Title: Galectin-3 Expression and Effect of Supplementation in Neonatal Mice with Disseminated Candida albicans Infection

doi: 10.1038/s41390-019-0279-x

Figure Lengend Snippet: Tissue fungal burden and galectin-3 concentrations in neonatal mice with disseminated candidiasis. Two-day-old mouse pups (n=14) were infected via intraperitoneal injection with C. albicans and euthanized at 24 h after injection. Panel A: Tissue fungal burden. Panel B: Mean gal3 concentration in tissue homogenates compared to pups receiving saline (n=14). Error bars represent standard deviation. Mean gal3 concentration was reduced in the spleen of infected pups relative to controls based on ANOVA with inter-group comparisons by the Student-Newman-Keuls Method (p=0.04).

Article Snippet: In a subsequent experiment, pups were delivered and randomized to receive 5 μg carrier-free recombinant mouse gal3 (R&D Systems, Minneapolis, MN) or saline in 20 μl i.p. injection prior to infection with C. albicans as described above.

Techniques: Infection, Injection, Concentration Assay, Saline, Standard Deviation

Journal: Pediatric research

Article Title: Galectin-3 Expression and Effect of Supplementation in Neonatal Mice with Disseminated Candida albicans Infection

doi: 10.1038/s41390-019-0279-x

Figure Lengend Snippet: Survival curve of neonatal mice with disseminated candidiasis after pretreatment with recombinant galectin-3. Two-day-old mouse pups were given intraperitoneal injections of either saline or recombinant gal3, 2 h prior to infection with C. albicans . Uninfected pups receiving gal3 only were included as a control. Pretreatment with gal3 reduced mortality in infected compared to saline treated pups based on log-rank test (p=0.02).

Article Snippet: In a subsequent experiment, pups were delivered and randomized to receive 5 μg carrier-free recombinant mouse gal3 (R&D Systems, Minneapolis, MN) or saline in 20 μl i.p. injection prior to infection with C. albicans as described above.

Techniques: Recombinant, Saline, Infection, Control

Journal: Pediatric research

Article Title: Galectin-3 Expression and Effect of Supplementation in Neonatal Mice with Disseminated Candida albicans Infection

doi: 10.1038/s41390-019-0279-x

Figure Lengend Snippet: Tissue fungal burden in neonatal mice with disseminated candidiasis pretreated with galectin-3. Two-day-old mouse pups were given intraperitoneal injections of either saline or recombinant gal3, 2 h prior to infection with C. albicans (n=10 pups per group). Tissues were collected at the time of death or at 72 h in surviving animals. Fungal burden is depicted with the bars representing median values. P values were derived based on analysis using a negative binomial model to account for the variability in these data which are not normally distributed.

Article Snippet: In a subsequent experiment, pups were delivered and randomized to receive 5 μg carrier-free recombinant mouse gal3 (R&D Systems, Minneapolis, MN) or saline in 20 μl i.p. injection prior to infection with C. albicans as described above.

Techniques: Saline, Recombinant, Infection, Derivative Assay

Journal: Pediatric research

Article Title: Galectin-3 Expression and Effect of Supplementation in Neonatal Mice with Disseminated Candida albicans Infection

doi: 10.1038/s41390-019-0279-x

Figure Lengend Snippet: Tissue fungal burden in neonatal mice with disseminated candidiasis at early time points following infection and pretreatment with galectin-3. Two-day-old mouse pups were given intraperitoneal injections of either saline or recombinant gal3, 2 h prior to infection with C. albicans (n=5 pups per group). Pups were euthanized and tissues were collected at 24 and 36 h following infection. Fungal burden is depicted with the bars representing median values. P values were derived based on analysis using a negative binomial model to account for the variability in these data which are not normally distributed. NS – not significant.

Article Snippet: In a subsequent experiment, pups were delivered and randomized to receive 5 μg carrier-free recombinant mouse gal3 (R&D Systems, Minneapolis, MN) or saline in 20 μl i.p. injection prior to infection with C. albicans as described above.

Techniques: Infection, Saline, Recombinant, Derivative Assay

Journal: Investigative Ophthalmology & Visual Science

Article Title: Transmembrane Mucin 1 Blocks Fluorescein Ingress to Corneal Epithelium

doi: 10.1167/iovs.63.2.31

Figure Lengend Snippet: (A) The expression of MUC16 protein in HCECs separately transduced with two lentiviral-based shRNA vectors (shMUC16-1 or shMUC16-2) were examined through a Western blot analysis. The a-tubulin was used as a loading control. (B) Representative flow cytometric analyses of fluorescence intensity and MUC16 expressions in transduced HCECs followed by fluorescein staining were shown in contour plots. (C) Representative flow cytometric analyses of fluorescence intensity and MUC16 expression in HCECs cultured in KSFM ( left ) or MPM ( right ) followed by fluorescein staining were presented in contour plots. (D) The frozen tissue sections prepared from fluorescein-stained corneal epithelia were stained with GAL3 ( red , immunofluorescence), and the cell nuclei were counterstained with DAPI ( blue , immunofluorescence).

Article Snippet: To detect the expression of GAL3, cell suspensions were fixed and incubated with anti-galectin-3 (GAL3) mouse antibody (no. A3A12; Novus Biologicals).

Techniques: Expressing, Transduction, shRNA, Western Blot, Control, Fluorescence, Staining, Cell Culture, Immunofluorescence

Journal: Science Advances

Article Title: Galectin-3 exacerbates autoimmune diabetes by limiting regulatory T cell differentiation and function

doi: 10.1126/sciadv.adz7916

Figure Lengend Snippet: ( A ) Dynamic circulating levels of Galectin-3 in NOD and their control BALB/c mice at different ages ( n = 6). ( B ) The mRNA abundance of Galectin-3 in the pancreatic islets of NOD and BALB/c mice, expressed as an arbitrary unit after normalization for β-actin mRNA levels, relative to the levels of 4-week-old BALB/c mice ( n = 4). ( C ) The content of Galectin-3 in the pancreatic islet lysates of NOD and BALB/c mice ( n = 3). ( D and E ) Representative patterns and median fluorescence intensity (MFI) of Galectin-3 expressed by CD45 + leukocytes (D) and CD45 + F4/80 + CD11b + macrophages (E) in pancreatic islets of 4- and 12-week-old NOD and BALB/c mice ( n = 4). ( F ) Galectin-3 expression in pancreatic islet macrophages of NOD mice at different ages ( n = 3 to 4) from the National Center for Biotechnology Information database ( GSE141782 ). ( G ) Representative patterns and MFI of Galectin-3 expressed by insulin + pancreatic islet β cells of 4- and 12-week-old NOD and BALB/c mice ( n = 4). Data are expressed as mean ± SEM. ns, no significance. *P < 0.05, **P < 0.01, ***P < 0.001, and ****P < 0.0001. 4w, 4 weeks; 12w, 12 weeks.

Article Snippet: Hematoxylin and eosin (H&E) staining analysis of pancreatic sections showed more severe islet destruction and much higher insulitis scores in 12-week-old Galectin-3 +/+ NOD mice compared to Galectin-3 −/− NOD mice ( ).

Techniques: Control, Fluorescence, Expressing

Journal: Science Advances

Article Title: Galectin-3 exacerbates autoimmune diabetes by limiting regulatory T cell differentiation and function

doi: 10.1126/sciadv.adz7916

Figure Lengend Snippet: ( A ) Diabetes incidence in Galectin-3 +/+ NOD ( n = 20) and Galectin-3 −/− NOD mice ( n = 16) at different ages. ( B ) Glucose excursion curve after receiving intraperitoneal glucose tolerance test (IPGTT) and the quantification of area under the curve in IPGTT ( n = 8 to 9). ( C ) Representative images of H&E analysis for pancreatic sections and ( D ) insulitis scores in 12-week-old female Galectin-3 +/+ NOD and Galectin-3 −/− NOD mice ( n = 4). Scale bar, 50 μm. ( E ) Representative images of immunohistochemistry (IHC) staining of TUNEL (green) and insulin (red) in pancreas of 12-week-old female Galectin-3 +/+ NOD and Galectin-3 −/− NOD mice ( n = 6). Scale bar, 20 μm, with magnification of ×400. Data are expressed as mean ± SEM. *P < 0.05, **P < 0.01, and ***P < 0.001.

Article Snippet: Hematoxylin and eosin (H&E) staining analysis of pancreatic sections showed more severe islet destruction and much higher insulitis scores in 12-week-old Galectin-3 +/+ NOD mice compared to Galectin-3 −/− NOD mice ( ).

Techniques: Immunohistochemistry, TUNEL Assay

Journal: Science Advances

Article Title: Galectin-3 exacerbates autoimmune diabetes by limiting regulatory T cell differentiation and function

doi: 10.1126/sciadv.adz7916

Figure Lengend Snippet: Pancreatic islets from 12-week-old nondiabetic Galectin-3 +/+ NOD and Galectin-3 −/− NOD mice were harvested and subject to flow cytometry analysis ( n = 5). Frequencies of CD4 + and CD8 + T cells ( A ) and T reg cells (CD4 + CD25 + FOXP3 + ) ( B ) are shown as representative fluorescence-activated cell sorting plots. ( C to E ) Frequencies of T H 1 (CD4 + IFN-γ + ) (C), T H 2 (CD4 + IL-4 + ) (D), and T H 17 (CD4 + IL-17 + ) (E) subsets among CD45 + cells in islets of 12-week-old Galectin-3 +/+ NOD and Galectin-3 −/− NOD mice. ( F to J ) Frequencies of perforin + CD8 + (F), Tc1 (CD8 + IFN-γ + ) (G), Tc2 (CD8 + IL-4 + ) (H), Tc17 (CD8 + IL-17 + ) (I), and granzyme B + CD8 + (J) subsets among CD45 + cells in islets of 12-week-old Galectin-3 +/+ NOD and Galectin-3 −/− NOD mice. Data are expressed as mean ± SEM. *P < 0.05.

Article Snippet: Hematoxylin and eosin (H&E) staining analysis of pancreatic sections showed more severe islet destruction and much higher insulitis scores in 12-week-old Galectin-3 +/+ NOD mice compared to Galectin-3 −/− NOD mice ( ).

Techniques: Flow Cytometry, Fluorescence, FACS

Journal: Science Advances

Article Title: Galectin-3 exacerbates autoimmune diabetes by limiting regulatory T cell differentiation and function

doi: 10.1126/sciadv.adz7916

Figure Lengend Snippet: ( A ) Schematic diagram showing the protocol of TD139 treatment. ( B ) Incidence of diabetes expressed as percentage of diabetic mice at different ages ( n = 20). ( C ) Glucose excursion curve after receiving IPGTT ( n = 8). ( D ) Serum insulin concentration at 0 and 15 min during the IPGTT in 12-week-old TD139 or vehicle-treated NOD mice ( n = 7 to 8). ( E ) Representative H&E staining of pancreas from 12-week-old NOD mice treated with TD139 or vehicle ( n = 4). ( F ) Insulitis scores calculated on the basis of histological evaluation of pancreatic section as in (E). ( G ) Frequencies of CD4 + and CD8 + T cells in total CD3 + T cells from islets of 12-week-old NOD mice treated with TD139 or vehicle ( n = 4). ( H ) Frequencies of T reg cells (CD4 + CD25 + FOXP3 + ) from islets of 12-week-old NOD mice treated with TD139 or vehicle ( n = 4). ( I to K ) Frequencies of perforin + CD8 + T, Tc1 (CD8 + IFN-γ + ), Tc2 (CD8 + IL-4 + ), Tc17 (CD8 + IL-17 + ), and granzyme B + CD8 + T subsets from islets of 12-week-old NOD mice treated with TD139 or vehicle ( n = 3 to 4). ( L ) Frequencies of T H 1 (CD4 + IFN-γ + ), T H 2 (CD4 + IL-4 + ), and T H 17 (CD4 + IL-17 + ) from islets of 12-week-old NOD mice treated with TD139 or vehicle ( n = 4). Data are expressed as mean ± SEM. *P < 0.05 and **P < 0.01.

Article Snippet: Hematoxylin and eosin (H&E) staining analysis of pancreatic sections showed more severe islet destruction and much higher insulitis scores in 12-week-old Galectin-3 +/+ NOD mice compared to Galectin-3 −/− NOD mice ( ).

Techniques: Concentration Assay, Staining

Journal: Arteriosclerosis, Thrombosis, and Vascular Biology

Article Title: Flanking Recipient Vasculature, Not Circulating Progenitor Cells, Contributes to Endothelium and Smooth Muscle in Murine Allograft Vasculopathy

doi: 10.1161/atvbaha.110.221184

Figure Lengend Snippet: Figure 5. The majority of blood-derived cells in AV lesions were found to be macrophages. Shown is a BALB/c allograft that was inserted into a nonfluorescent isograft in an eGFPapoE/ B6 mouse (see Figure 1c). Most eGFP cells stained positive for the macrophage marker Mac2 (red in a and c). Green indicates eGFP; blue, nuclei; a, merged color channels; b, green channel; c, red channel; gray, DIC; L, lumen; M, tunica media. Scale bars50 m.

Article Snippet: The Mac2 epitope was stained as a marker for plaque macrophages by using rat anti-mouse Mac2 antibody (CL8942AP, Cedarlane Labs, 1:500) followed by Texas Red–conjugated goat anti-rat secondary antibody (Jackson Immunoresearch, 1:200).

Techniques: Derivative Assay, Staining, Marker

Journal: Frontiers in Cellular and Infection Microbiology

Article Title: Autophagy Receptor Tollip Facilitates Bacterial Autophagy by Recruiting Galectin-7 in Response to Group A Streptococcus Infection

doi: 10.3389/fcimb.2020.583137

Figure Lengend Snippet: Tollip is recruited to GAS-containing vacuoles before GAS damages the endosomal membrane. (A) HeLa cells transfected with mCherry-Tollip and EmGFP-LC3 were infected with Group A Streptococcus (GAS) for 4 h, and immunostained with anti-Group A Streptococcus carbohydrate (GAC) to stain GAS. Immunofluorescence analysis was performed to determine the localization of mCherry-Tollip and EmGFP-LC3. Cellular and bacterial DNA were stained with DAPI. Insets: enlarged boxed areas. All images are representative of at least 3 independent experiments. Scale bar, 10 μm. (B) Percentages of cells containing Tollip-positive GAS and cells containing Tollip-positive autophagosomes; cells were manually counted using immunofluorescence confocal microscopy. In each experiment, >60 autophagosomes were evaluated per sample; bars: means ± SD of 3 independent experiments. (C) HeLa cells were transfected with mCherry-Tollip and infected with GAS or SLO-deletion-mutant (ΔSLO) GAS for 4 h, and then stained for endogenous galectin-3. Immunofluorescence microscopy was used to examine the localization of mCherry-Tollip and endogenous galectin-3. Insets: enlarged boxed areas. All images are representative of at least 3 independent experiments. Scale bar, 10 μm.

Article Snippet: The following primary and secondary antibodies were used in the study: mouse monoclonal anti-galectin-3 (1:250, B2C10; BD Biosciences, 556904), rabbit polyclonal anti-galectin-7 (1:250 dilution for immunofluorescence, 1:1000 dilution for Western blot, Abcam, ab10482), mouse monoclonal anti-LAMP-1 (1:250, H4A3; Santa Cruz Biotechnology, sc-20011), rabbit polyclonal anti-Tollip (1:1000, Abcam, ab187198), rabbit monoclonal anti-NBR1 (1;250, D2E6; Cell Signaling, 9891), mouse monoclonal anti-p62 (1:250, D-3; Santa Cruz Biotechnology, sc-28359), rabbit monoclonal anti-TAX1BP1 (1:250, D1D5; Cell Signaling, 5105), mouse monoclonal anti-streptolysin (1:2000, 6D11; Abcam, ab23501), mouse monoclonal anti-GFP (1:1000, GF200; Nacalai Tesque, 04363‐24), mouse monoclonal anti-GAPDH (1:1000, 6C5; Santa Cruz Biotechnology, sc-32233), and mouse monoclonal anti-FLAG (1:1000, M2; Sigma-Aldrich, A2220).

Techniques: Transfection, Infection, Staining, Immunofluorescence, Confocal Microscopy, Mutagenesis, Microscopy