goat-derived anti-gal3  (R&D Systems)

Journal: International Journal of Molecular Sciences

Article Title: Gal3 Plays a Deleterious Role in a Mouse Model of Endotoxemia

doi: 10.3390/ijms23031170

Figure Lengend Snippet: Gal3 knockout mice are more resistant to septic shock. WT and Gal3KO mice were subjected to an intraperitoneal injection of LPS (5 mg/kg of body weight) or saline solution ( N = 12 animals per group). The mortality rate was monitored regularly for 80 h and represented as percentage of survival. The statistical analysis was performed using the Log-Rank Test. Abbreviations: WT, wild type mice; KO, Gal3 knockout mice; WTLPS, wild type mice treated with LPS; KOLPS, Gal3 knockout mice treated with LPS. *, p < 0.05.

Article Snippet: Primary antibodies used were goat-derived anti-Gal3 (R&D Systems, Minneapolis, MN, USA; 1:100), rat-derived anti-CD4 (Santa Cruz Biotechnologies, Santa Cruz, CA, USA; 1:100), and mouse-derived anti-CD68 (Invitrogen, Waltham, MA, USA; 1:100).

Techniques: Knock-Out, Injection, Saline

Journal: International Journal of Molecular Sciences

Article Title: Gal3 Plays a Deleterious Role in a Mouse Model of Endotoxemia

doi: 10.3390/ijms23031170

Figure Lengend Snippet: Determination of Gal3 on immune cells of peripheral blood. Levels of Gal3 were determined by flow cytometry in peripheral blood of mice ( N = 5 animals). Briefly, after blood collection, erythrocytes were lysed using an ammonium chloride lysis solution. Cells were washed and stained with surface marker antibodies for 20 min on ice. Gal-3 expression was analyzed in B-cells, CD4 + T-cells, CD8 + T-cells, double negative T-cells, dendritic cells, neutrophils, and macrophages. Results are expressed as median ± IQR. Statistical significance was calculated using the Mann–Whitney U test. Abbreviations: DCs, dendritic cells; Macro, macrophages; Monoc, monocytes; Neutro, neutrophils. *, p < 0.01.

Article Snippet: Primary antibodies used were goat-derived anti-Gal3 (R&D Systems, Minneapolis, MN, USA; 1:100), rat-derived anti-CD4 (Santa Cruz Biotechnologies, Santa Cruz, CA, USA; 1:100), and mouse-derived anti-CD68 (Invitrogen, Waltham, MA, USA; 1:100).

Techniques: Flow Cytometry, Lysis, Staining, Marker, Expressing, MANN-WHITNEY

Journal: International Journal of Molecular Sciences

Article Title: Gal3 Plays a Deleterious Role in a Mouse Model of Endotoxemia

doi: 10.3390/ijms23031170

Figure Lengend Snippet: Determination of Gal3 and TLR4. ( A )The expression of Gal3 in serum was measured by ELISA. Blood samples were collected from the heart of mice 1 h after LPS/saline injection. Results are mean ± SD of N = 4–6 animals, expressed as ng/mLof the analyzed protein and relative to the WT group. Statistical significance (two tailed Student’s t test): p < 0.001. Using RT-PCR, the mRNA expression of Gal3 was measured in the liver ( B ), spleen ( C ), and peritoneal macrophages ( D ). Using RT-PCR, the mRNA expression of TLR4 was measured in the liver ( E ), spleen ( F ), and peritoneal macrophages ( G ). For PCR analysis, animals were culled 1 h after LPS/saline injection. Results are mean ± SD of N = 3–10 animals, normalized to β-actin and expressed as relative expression to the WT group. Statistical significance (one-way ANOVA followed by the LSD post hoc test for multiple comparisons): *—compared with WT group; #—compared with KO group; $—compared with WTLPS group; p < 0.001 for ( B – D ), p < 0.05 for ( G ). Abbreviations: WT, wild type mice; KO, Gal3 knockout mice; WTLPS, wild type mice treated with LPS; KOLPS, Gal3 knockout mice treated with LPS.

Article Snippet: Primary antibodies used were goat-derived anti-Gal3 (R&D Systems, Minneapolis, MN, USA; 1:100), rat-derived anti-CD4 (Santa Cruz Biotechnologies, Santa Cruz, CA, USA; 1:100), and mouse-derived anti-CD68 (Invitrogen, Waltham, MA, USA; 1:100).

Techniques: Expressing, Enzyme-linked Immunosorbent Assay, Saline, Injection, Two Tailed Test, Reverse Transcription Polymerase Chain Reaction, Knock-Out

Journal: International Journal of Molecular Sciences

Article Title: Gal3 Plays a Deleterious Role in a Mouse Model of Endotoxemia

doi: 10.3390/ijms23031170

Figure Lengend Snippet: Gal3 expression in the liver, lung, and spleen of mice from the different treatments assayed. ( A ) Section from liver showing some Gal3 positive cells in a WT animal. ( B ) Immunoreactivity of Gal3 in a liver section from a WT animal injected i.p. with LPS. ( F ) Section from spleen showing some Gal3 positive cells in a WT animal. ( G ) Immunoreactivity of Gal3 in a spleen section from a WT animal injected i.p. with LPS. Again, a strong reaction can be seen. ( I ) Section from lung showing some Gal3 positive cells in a WT animal. ( G ) Immunoreactivity of Gal3 in a lung section from a WT animal injected i.p. with LPS, showing a strong reaction. As expected, in the case of Gal3KO animals, immunoreactivity of Gal3 was not found in any treatment assayed ( C , D ). Scale bar: ( A – D ), 100 μm; ( F – J ), 20 μm. Quantification of the density of Gal3 positive cells in liver ( E ), spleen ( H ), and lung ( K ) from the different treatments assayed. Results are mean ± SD of N = 3–4 animals, expressed as number of cells per mm 2 . Statistical significance (two tailed Student- t test): * p < 0.01; ** p < 0.05. Abbreviations: WT, wild type mice; WTLPS, wild type mice treated with LPS; KO, Gal3 knockout mice; KOLPS, Gal3 knockout mice treated with LPS.

Article Snippet: Primary antibodies used were goat-derived anti-Gal3 (R&D Systems, Minneapolis, MN, USA; 1:100), rat-derived anti-CD4 (Santa Cruz Biotechnologies, Santa Cruz, CA, USA; 1:100), and mouse-derived anti-CD68 (Invitrogen, Waltham, MA, USA; 1:100).

Techniques: Expressing, Injection, Two Tailed Test, Knock-Out

Journal: International Journal of Molecular Sciences

Article Title: Gal3 Plays a Deleterious Role in a Mouse Model of Endotoxemia

doi: 10.3390/ijms23031170

Figure Lengend Snippet: Double immunofluorescence of Iba1 andGal3 expression. ( A ) Iba1 staining in the liver of a WT animal. Virtually, no monocytes/macrophages are infiltrated in control animals. ( B ) Section from liver showing some Gal3 positive cells in a WT animal. ( C ) Merge image showing co-localization of Iba1 + cells and Gal3 + cells. ( D ) Immunoreactivity of Iba1 in a liver section from a WT animal injected i.p. with LPS. A strong reaction can be seen. ( E ) Immunoreactivity of Gal3 in WT animals injected with LPS. ( F ) Merge image showing co-localization of Iba1 + cells and Gal3 + cells (arrows). Most Gal3 + cells co-localize with Iba1 + cells. Scale bar: 50 μm.

Article Snippet: Primary antibodies used were goat-derived anti-Gal3 (R&D Systems, Minneapolis, MN, USA; 1:100), rat-derived anti-CD4 (Santa Cruz Biotechnologies, Santa Cruz, CA, USA; 1:100), and mouse-derived anti-CD68 (Invitrogen, Waltham, MA, USA; 1:100).

Techniques: Immunofluorescence, Expressing, Staining, Injection

Journal: International Journal of Molecular Sciences

Article Title: Gal3 Plays a Deleterious Role in a Mouse Model of Endotoxemia

doi: 10.3390/ijms23031170

Figure Lengend Snippet: Effect of Gal3 deletion on the expression of TNF-α , iNOS , IL-1β , IL-6 , YM1 , IL-10 , and arginase mRNAs in the liver, spleen, and macrophages of mice from the different treatments assayed, measured by RT-PCR ( A – U ). Animals were culled 1 h after LPS/saline injection. Results are mean ± SD of N = 3–10 animals, normalized to β-actin and expressed as relative expression to the WT group. Statistical significance (one-way ANOVA followed by the LSD post hoc test for multiple comparisons): *—compared with WT group; #—compared with KO group; $—compared with WTLPS group; p < 0.001 for ( A – F , H , J – M , O , P , T ); p < 0.01 for ( G , N , S ); p < 0.05 for ( Q , R , U ). Abbreviations: WT, wild type mice; KO, Gal3 knockout mice; WTLPS, wild type mice treated with LPS; KOLPS, Gal3 knockout mice treated with LPS.

Article Snippet: Primary antibodies used were goat-derived anti-Gal3 (R&D Systems, Minneapolis, MN, USA; 1:100), rat-derived anti-CD4 (Santa Cruz Biotechnologies, Santa Cruz, CA, USA; 1:100), and mouse-derived anti-CD68 (Invitrogen, Waltham, MA, USA; 1:100).

Techniques: Expressing, Reverse Transcription Polymerase Chain Reaction, Saline, Injection, Knock-Out

Journal: International Journal of Molecular Sciences

Article Title: Gal3 Plays a Deleterious Role in a Mouse Model of Endotoxemia

doi: 10.3390/ijms23031170

Figure Lengend Snippet: CD68 expression in liver. Representative immunostaining from sections of the different treatments assayed. WT ( A ) and KO ( C ) animals show a normal pattern of CD68 expression; however, the treatment with LPS produces a strong induction of CD68-positive cells in WT animals ( B ). Absence of Gal3 clearly reduces this effect ( D ). Scale bar: 100 μm. ( E ) Quantification of the density of CD68 positive cells in liver. Results are mean ± SD of N = 3 animals, expressed as number of cells per mm 2 . Statistical significance (one-way ANOVA followed by the LSD post hoc test for multiple comparisons): *—compared with WT group; #—compared with KO group; $—compared with WTLPS group; p < 0.01. Abbreviations: WT, wild type mice; WTLPS, wild type mice treated with LPS; KO, Gal3 knockout mice; KOLPS, Gal3 knockout mice treated with LPS.

Article Snippet: Primary antibodies used were goat-derived anti-Gal3 (R&D Systems, Minneapolis, MN, USA; 1:100), rat-derived anti-CD4 (Santa Cruz Biotechnologies, Santa Cruz, CA, USA; 1:100), and mouse-derived anti-CD68 (Invitrogen, Waltham, MA, USA; 1:100).

Techniques: Expressing, Immunostaining, Knock-Out

Journal: International Journal of Molecular Sciences

Article Title: Gal3 Plays a Deleterious Role in a Mouse Model of Endotoxemia

doi: 10.3390/ijms23031170

Figure Lengend Snippet: CD4 expression in liver. Representative immunostaining from sections of the different treatments assayed. WT ( A ) and KO ( C ) animals show a normal pattern of CD4 expression; however, the treatment with LPS produces a strong induction of CD4-positive cells in WT animals ( B ). Absence of Gal3 clearly reduces this effect ( D ). Scale bar: 100 μm. ( E ) Quantification of the density of CD4 positive cells in liver. Results are mean ± SD of N = 3 animals, expressed as number of cells per mm 2 . Statistical significance (one-way ANOVA followed by the LSD post hoc test for multiple comparisons): *—compared with WT group; #—compared with KO group; $—compared with WTLPS group; p < 0.001. Abbreviations: WT, wild type mice; WTLPS, wild type mice treated with LPS; KO, Gal3 knockout mice; KOLPS, Gal3 knockout mice treated.

Article Snippet: Primary antibodies used were goat-derived anti-Gal3 (R&D Systems, Minneapolis, MN, USA; 1:100), rat-derived anti-CD4 (Santa Cruz Biotechnologies, Santa Cruz, CA, USA; 1:100), and mouse-derived anti-CD68 (Invitrogen, Waltham, MA, USA; 1:100).

Techniques: Expressing, Immunostaining, Knock-Out

Journal: International Journal of Molecular Sciences

Article Title: Gal3 Plays a Deleterious Role in a Mouse Model of Endotoxemia

doi: 10.3390/ijms23031170

Figure Lengend Snippet: Determination of TNF-α and IL-6 levels in serum. TNF-α ( A ) and IL-6 ( B ) levels in serum of WT and Gal3KO animals. Blood samples were collected from the heart of mice 1 h after LPS/saline injection. Results are mean ± SD of N = 4–10 animals, expressed as ng/mLof the analyzed protein and relative to the WT group. Statistical significance (one-way ANOVA followed by the LSD post hoc test for multiple comparisons): *—compared with WT group; #—compared with KO group; $—compared with WTLPS group; p < 0.001. Abbreviations: WT, wild type mice; KO, Gal3 knockout mice; WTLPS, wild type mice treated with LPS; KOLPS, Gal3 knockout mice treated with LPS.

Article Snippet: Primary antibodies used were goat-derived anti-Gal3 (R&D Systems, Minneapolis, MN, USA; 1:100), rat-derived anti-CD4 (Santa Cruz Biotechnologies, Santa Cruz, CA, USA; 1:100), and mouse-derived anti-CD68 (Invitrogen, Waltham, MA, USA; 1:100).

Techniques: Saline, Injection, Knock-Out

Journal: International Journal of Molecular Sciences

Article Title: Gal3 Plays a Deleterious Role in a Mouse Model of Endotoxemia

doi: 10.3390/ijms23031170

Figure Lengend Snippet: Histopathology of the livers of mice from the different treatments assayed. ( A ) Hepatic lobule with Remak cords of apparently normal hepatocytes (arrow). ( B ) Detail of hepatocyte with abundant rough endoplasmic reticulum (RER) and mitochondria (circle). ( C ) Hepatic parenchyma, appreciating abundant unilocular and multilocular steatosis (arrow). ( D ) Detail of hepatocyte with abundant diffuse fat (circle). ( E ) Detail of Remak cords with steatosis in the hepatocytes (circle). ( F ) Hepatocytes with vacuolations of its membranous system (arrow). ( G ) Histological score showing a semiquantitative analysis of steatosis in the liver. The pathology scores were as follows: 0, without significant injuries (0%); 1, minimum (<10%); 2, mild (11–25%); 3, moderate (26–50%); 4, marked (51–75%); 5, severe (>75%). Histopathological evaluation is performed in a blind manner by two highly experienced pathologists. Results are mean ± SD of N = 3 animals.Statistical significance (two tailed Student- t test): * p < 0.001. Abbreviations: WT, wild type mice; WTLPS, wild type mice treated with LPS; KO, Gal3 knockout mice; KOLPS, Gal3 knockout mice treated with LPS; ND, not damaged. ( A , C , E ), optical microscopy. Scale bars: 100 µm. ( B , D , F ), ultrastructural observations. Scale bars: ( B ), 10 µm; ( D , F ), 5 µm.

Article Snippet: Primary antibodies used were goat-derived anti-Gal3 (R&D Systems, Minneapolis, MN, USA; 1:100), rat-derived anti-CD4 (Santa Cruz Biotechnologies, Santa Cruz, CA, USA; 1:100), and mouse-derived anti-CD68 (Invitrogen, Waltham, MA, USA; 1:100).

Techniques: Histopathology, Two Tailed Test, Knock-Out, Microscopy

Journal: International Journal of Molecular Sciences

Article Title: Gal3 Plays a Deleterious Role in a Mouse Model of Endotoxemia

doi: 10.3390/ijms23031170

Figure Lengend Snippet: Histopathology of the spleens of mice from the different treatments assayed. ( A ) Apparently normal spleen. Normal white pulp (lymphoid follicles) (circle). ( B ) Marginal zone of the apparently normal spleen. ( C ) Hypertrophy and hyperplasia of lymphoid follicles very marked (circle). ( D ) Detail of lymphoid follicle with abundant lymphoblasts (LB) and lymphocytes (L), and some red blood cells (RBC). ( E ) Detail of spleen with hypertrophy and especially hyperplasia of the lymphoid follicles (circle). ( F ) Detail of lymphoid follicle with lymphocytes (L), lymphoblasts (LB) and reticular cells (RC). ( G ) Histological score showing a semiquantitative analysis of hypertrophy and hyperplasia of lymphoid follicles in the spleen. The pathology scores were as follows: 0, without significant injuries (0%); 1, minimum (<10%); 2, mild (11–25%); 3, moderate (26–50%); 4, marked (51–75%); 5, severe (>75%). Histopathological evaluation is performed in a blind manner by two highly experienced pathologists. Results are mean ± SD of N = 3 animals. Statistical significance (two tailed Student- t test), * p < 0.001. Abbreviations: WT, wild type mice; WTLPS, wild type mice treated with LPS; KO, Gal3 knockout mice; KOLPS, Gal3 knockout mice treated with LPS; ND, not damaged. ( A , C , E ), optical microscopy. Scale bars: 100 µm. ( B , D , F ), ultrastructural observations. Scale bars: ( B , D ), 10 µm; ( F ), 5 µm.

Article Snippet: Primary antibodies used were goat-derived anti-Gal3 (R&D Systems, Minneapolis, MN, USA; 1:100), rat-derived anti-CD4 (Santa Cruz Biotechnologies, Santa Cruz, CA, USA; 1:100), and mouse-derived anti-CD68 (Invitrogen, Waltham, MA, USA; 1:100).

Techniques: Histopathology, Two Tailed Test, Knock-Out, Microscopy

Journal: International Journal of Molecular Sciences

Article Title: Gal3 Plays a Deleterious Role in a Mouse Model of Endotoxemia

doi: 10.3390/ijms23031170

Figure Lengend Snippet: Histopathology of the lungs of mice from the different treatments assayed. ( A ) Lung in which an apparently normal bronchus (BR) and respiratory lobules (RL) stand out. ( B ) Detail of capillary and pulmonary alveolus separated by the respiratory barrier (circle) formed by the endothelial cell (EC) and pneumocyteI (NI). ( C ) Detail of lung that shows a marked atelectasis in both respiratory lobule (RL) and alveoli (ALV). ( D ) Detail of lung with capillary hyperemia (circle) and hypertrophy of pneumocytes II (NII). ( E ) Lung detail showing atelectasis zones (circle) and emphysema (arrow). ( F ) Septal hypertrophy of pneumocytes II (NII). ( G ) Histological score showing a semiquantitative analysis of atelectasis in the lung. The pathology scores were as follows: 0, without significant injuries (0%); 1, minimum (<10%); 2, mild (11–25%); 3, moderate (26–50%); 4, marked (51–75%); 5, severe (>75%). Histopathological evaluation is performed in a blind manner by two highly experienced pathologists. Results are mean ± SD of N = 3 animals. Statistical significance (two tailed Student- t test), * p < 0.001. Abbreviations: WT, wild type mice; WTLPS, wild type mice treated with LPS; KO, Gal3 knockout mice; KOLPS, Gal3 knockout mice treated with LPS; ND, not damaged. ( A , C , E ), optical microscopy. Scale bars: 100 µm. ( B , D , F ), ultrastructural observations. Scale bars: ( B , F ), 2 µm; ( D ), 5 µm.

Article Snippet: Primary antibodies used were goat-derived anti-Gal3 (R&D Systems, Minneapolis, MN, USA; 1:100), rat-derived anti-CD4 (Santa Cruz Biotechnologies, Santa Cruz, CA, USA; 1:100), and mouse-derived anti-CD68 (Invitrogen, Waltham, MA, USA; 1:100).

Techniques: Histopathology, Two Tailed Test, Knock-Out, Microscopy

Journal: International Journal of Molecular Sciences

Article Title: Gal3 Plays a Deleterious Role in a Mouse Model of Endotoxemia

doi: 10.3390/ijms23031170

Figure Lengend Snippet: Histopathology of the brains of mice from the different treatments assayed. ( A ) Detail of cerebral cortex with abundant neurons. ( B ) Detail of apparently normal neuron and oligodendrocyte (circle). ( C ) Detail of cerebral cortex, showing mobilization of glia cells (circle). ( D ) Detail of a damaged neuron densified and vacuolized. ( E ) Detail of cerebral cortex with a tenuous hyperemia (circle). ( F ) Detail of neuron with lipofucsin precursor granules (Gr). ( G ) Histological score showing brain damage. The pathology scores were as follows: 0, without significant injuries (0%); 1, minimum (<10%); 2, mild (11–25%); 3, moderate (26–50%); 4, marked (5–75%); 5, severe (>75%). Histopathological evaluation is performed in a blind manner by two highly experienced pathologists. Results are mean ± SD of N = 3 animals. Statistical significance (two tailed Student- t test), * p < 0.001. Abbreviations: WT, wild type mice; WTLPS, wild type mice treated with LPS; KOLPS, Gal3 knockout mice treated with LPS; ND, not damaged. ( A , C , E ), optical microscopy. Scale bars: 100 µm. ( B , D , F ), ultrastructural observations. Scale bars: ( B , F ), 5 µm; ( D ), 2 µm. ( H ) Immunofluorescence of Iba1 showing a normal patter in microglial cells staining. ( I ) When animals were treated with LPS microglial cells activate and proliferate. Scale bars ( H , I ): 100 µm.

Article Snippet: Primary antibodies used were goat-derived anti-Gal3 (R&D Systems, Minneapolis, MN, USA; 1:100), rat-derived anti-CD4 (Santa Cruz Biotechnologies, Santa Cruz, CA, USA; 1:100), and mouse-derived anti-CD68 (Invitrogen, Waltham, MA, USA; 1:100).

Techniques: Histopathology, Two Tailed Test, Knock-Out, Microscopy, Immunofluorescence, Staining

Journal: International Journal of Molecular Sciences

Article Title: Gal3 Plays a Deleterious Role in a Mouse Model of Endotoxemia

doi: 10.3390/ijms23031170

Figure Lengend Snippet: Sense and antisense sequences of the primers used for the analysis of mRNA expression by qPCR.

Article Snippet: Primary antibodies used were goat-derived anti-Gal3 (R&D Systems, Minneapolis, MN, USA; 1:100), rat-derived anti-CD4 (Santa Cruz Biotechnologies, Santa Cruz, CA, USA; 1:100), and mouse-derived anti-CD68 (Invitrogen, Waltham, MA, USA; 1:100).

Techniques: Expressing