Journal: Cellular and Molecular Immunology

Article Title: Specific contribution of mannose-binding lectin murine isoforms to brain ischemia/reperfusion injury

doi: 10.1038/s41423-019-0225-1

Figure Lengend Snippet: Neuroscore and ischemic volume after 48 h of reperfusion in MBL −/− , MBL-A −/− , and MBL-C −/− mice. a Only MBL −/− mice had fewer behavioral deficits than WT mice when rated by the composite neuroscore (the sum of focal and general deficits). The data are shown as bars + SDs ( n = 7 for MBL −/− , n = 10 for MBL-A −/− , and n = 6 for MBL-C −/− ); Mann–Whitney test, *** p < 0.001. b Representative images of cresyl violet-stained sections, scale bar 1 mm. MBL −/− and MBL-A −/− mice had a smaller ischemic volume than WT mice. The data are shown as bars with individual values ± SDs ( n = 7 for MBL −/− , n = 10 for MBL-A −/− , and n = 6 for MBL-C −/− ); t test, ** p < 0.01, * p < 0.05

Article Snippet: Samples were then incubated for 1 h 30 min at room temperature (RT) with anti-MBL-A or anti-MBL-C antibodies (both 1 µg/mL, Hycult, # HM1035 and #HM1038, respectively).

Techniques: MANN-WHITNEY, Staining

Journal: Cellular and Molecular Immunology

Article Title: Specific contribution of mannose-binding lectin murine isoforms to brain ischemia/reperfusion injury

doi: 10.1038/s41423-019-0225-1

Figure Lengend Snippet: Neuronal count after 48 h of reperfusion in MBL −/− , MBL-A −/− , and MBL-C −/− mice. a Positioning of the cortical (outlines) and striatal (dotted outlines) regions of interest for calculating neuronal cell viability (CC: contralateral cortex; CS: contralateral striatum; IC: ipsilateral cortex; IS: ipsilateral striatum). The regions of interest were designed to include only the lesion area (pale cresyl violet staining) throughout the experimental groups. b , c Representative high-magnification images of cresyl violet-stained sections (scale bar, 50 µm) and neuronal counts showing that MBL −/− mice had more preserved neurons in the striatum and cortex and MBL-A −/− mice had more preserved neurons in the striatum than WT mice. The data are shown as bars with individual values ± SDs ( n = 7 for MBL −/− , n = 10 for MBL-A −/− , and n = 5 for MBL-C −/− ); t test, * p < 0.05. Compared with WT mice, MBL-C −/− mice showed no protection from ischemic injury

Article Snippet: Samples were then incubated for 1 h 30 min at room temperature (RT) with anti-MBL-A or anti-MBL-C antibodies (both 1 µg/mL, Hycult, # HM1035 and #HM1038, respectively).

Techniques: Staining

Journal: Cellular and Molecular Immunology

Article Title: Specific contribution of mannose-binding lectin murine isoforms to brain ischemia/reperfusion injury

doi: 10.1038/s41423-019-0225-1

Figure Lengend Snippet: Summary of the outcome measures in the MBL −/− , MBL-A −/− , or MBL-C −/− ischemic mice. Neuronal count represent the global value relative to striatum and cortex

Article Snippet: Samples were then incubated for 1 h 30 min at room temperature (RT) with anti-MBL-A or anti-MBL-C antibodies (both 1 µg/mL, Hycult, # HM1035 and #HM1038, respectively).

Techniques:

Journal: Cellular and Molecular Immunology

Article Title: Specific contribution of mannose-binding lectin murine isoforms to brain ischemia/reperfusion injury

doi: 10.1038/s41423-019-0225-1

Figure Lengend Snippet: Hepatic gene expression of Mbl1 and Mbl2 after 48 h of reperfusion in MBL −/− , MBL-A −/− , and MBL-C −/− mice. a , The time course of Mbl1 ( a ) and Mbl2 ( a’ ) hepatic expression in WT mice showed overexpression of Mbl1 at 24 h and 48 h and overexpression of Mbl2 at 48 h and 4 d after tMCAo. The data are shown as bars with individual values ± SDs ( n = 5–6); one-way ANOVA followed by Dunnett’s post hoc test, * p < 0.05, ** p < 0.01, *** p < 0.001 vs. naive. b At 48 h after tMCAo, Mbl1 hepatic expression in MBL-C −/− mice was induced to the same extent as that in WT mice ( b ). The data are shown as bars with individual values ± SDs ( n = 4–6); two-way ANOVA followed by Sidak’s post hoc test, * p < 0.05, ** p < 0.01. At 48 h after tMCAo, Mbl2 hepatic expression in MBL-A −/− mice was induced to the same extent as that in WT mice ( b' ). The data are shown as bars with individual values ± SDs ( n = 4–6), Welch-corrected ANOVA for unequal variances, * p < 0.05

Article Snippet: Samples were then incubated for 1 h 30 min at room temperature (RT) with anti-MBL-A or anti-MBL-C antibodies (both 1 µg/mL, Hycult, # HM1035 and #HM1038, respectively).

Techniques: Expressing, Over Expression

Journal: Cellular and Molecular Immunology

Article Title: Specific contribution of mannose-binding lectin murine isoforms to brain ischemia/reperfusion injury

doi: 10.1038/s41423-019-0225-1

Figure Lengend Snippet: Plasma levels of MBL-A and MBL-C and systemic activation of the complement system after 48 h of reperfusion in MBL −/− , MBL-A −/− , and MBL-C −/− mice. a At 48 h after tMCAo, MBL-A was consumed to the same extent in MBL-C −/− mice as in WT mice ( a ). The data are shown as bars with individual values ± SDs ( n = 5–6); Welch-corrected ANOVA for unequal variances, * p < 0.05 vs. naive WT mice, # p < 0.05 vs. naive MBL-C −/− mice. At 48 h after tMCAo, MBL-C was consumed to the same extent in MBL-A −/− mice as in WT mice ( a' ). The data are shown as bars with individual values ± SDs ( n = 5–6); two-way ANOVA followed by Sidak’s post hoc test, * p < 0.05 vs. naive WT mice, # p < 0.05 vs. naive MBL-C −/− mice. b The in vitro assay for LP activation on mannans showed no activation in plasma from MBL −/− and MBL-A −/− mice but some activation in plasma from MBL-C −/− mice. c The in vitro assay for LP activation on acBSA showed similar activation in plasma from mice of each genotype. The data in b and c refer to pools of plasma from 5–6 mice per group, and plasma concentrations are reported on a logarithmic scale

Article Snippet: Samples were then incubated for 1 h 30 min at room temperature (RT) with anti-MBL-A or anti-MBL-C antibodies (both 1 µg/mL, Hycult, # HM1035 and #HM1038, respectively).

Techniques: Activation Assay, In Vitro

Journal: Cellular and Molecular Immunology

Article Title: Specific contribution of mannose-binding lectin murine isoforms to brain ischemia/reperfusion injury

doi: 10.1038/s41423-019-0225-1

Figure Lengend Snippet: Gene and protein expression levels of MBL isoforms and complement system activation in the brain after ischemia/reperfusion in WT, MBL −/− , MBL-A −/− , and MBL-C −/− mice. a Basal gene expression (fragments per kilobase million, FKPM) of Mbl1 , Mbl2 , and HSE-MSF (C3) in Mus musculus brain cell populations. Oligodendrocyte precursors (OPCs, expressing NG2 ), endothelial cells (ECs, expressing PECAM ), microglia (expressing Itgam ), neurons (expressing RbFox3 ), and astrocytes (expressing GFAP ) do not express Mbl1 or Mbl2 . Microglia are the only brain cell population expressing HSE-MSF (C3). Data were obtained from single-cell RNA-seq databases as described in the Materials and methods section. b Expression changes (microarray analysis) of Mbl1 , Mbl2 , and HSE-MSF (C3) in the brain following brain ischemia/reperfusion in WT mice. Mbl1 and Mbl2 gene expression was not induced at 3 h or 24 h after tMCAo. C3 expression was significantly upregulated at 24 h after tMCAo. The data are expressed as the log 2 fold change compared with untreated mice, with a Benjamini–Hochberg multiple comparisons test for P value adjustment, * p < 0.05 vs. untreated mice ( n = 4 for each experimental group). Data were obtained from microarray databases as described in the Materials and methods section. c MBL-A and MBL-C protein presence in the brain cortices of WT, MBL-A −/− , and MBL-C −/− mice 48 h after tMCAo. MBL-A was present in WT and MBL-C −/− mice to a similar extent. MBL-C was present to a higher extent in MBL-A −/− mice than in WT mice. The data are expressed as the differences between the ipsilateral and contralateral optical densities (OD 450 nm) and are shown as bars with individual values ± SDs ( n = 6); two-way ANOVA followed by Sidak’s post hoc test, ** p < 0.01 and *** p < 0.001. d Representative high-magnification images of C3 immunolabeling at 48 h after tMCAo, scale bar 50 µm. e MBL −/− and MBL-A −/− but not MBL-C −/− mice had less C3 deposition in the cortex than WT mice. The data are shown as bars with individual values ± SDs ( n = 6–10); t test, ** p < 0.01, * p < 0.05

Article Snippet: Samples were then incubated for 1 h 30 min at room temperature (RT) with anti-MBL-A or anti-MBL-C antibodies (both 1 µg/mL, Hycult, # HM1035 and #HM1038, respectively).

Techniques: Expressing, Activation Assay, RNA Sequencing Assay, Microarray, Immunolabeling