Hypoxia-ischemia or excitotoxin-induced tissue plasminogen activator- dependent gelatinase activation in mice neonate brain microvessels.

Hypoxia-ischemia (HI) and excitotoxicity are validated causes of neonatal brain injuries and tissue plasminogen activator (t-PA) participates in the processes through proteolytic and receptor-mediated pathways. Brain microvascular endothelial cells from neonates in culture, contain and release more t-PA and gelatinases upon glutamate challenge than adult cells. We have studied t-PA to gelatinase (MMP-2 and MMP-9) activity links in HI and excitotoxicity lesion models in 5 day-old pups in wild type and in t-PA or its inhibitor (PAI-1) genes inactivated mice. Gelatinolytic activities were detected in SDS-PAGE zymograms and by in situ fluorescent DQ-gelatin microscopic zymographies. HI was achieved by unilateral carotid ligature followed by a 40 min hypoxia (8%O₂). Excitotoxic lesions were produced by intra parenchymal cortical (i.c.) injections of 10 µg ibotenate (Ibo). Gel zymograms in WT cortex revealed progressive extinction of MMP-2 and MMP-9 activities near day 15 or day 8 respectively. MMP-2 expression was the same in all strains while MMP-9 activity was barely detectable in t-PA⁻/⁻ and enhanced in PAI-1⁻/⁻ mice. HI or Ibo produced activation of MMP-2 activities 6 hours post-insult, in cortices of WT mice but not in t-PA⁻/⁻ mice. In PAI-1⁻/⁻ mice, HI or vehicle i.c. injection increased MMP-2 and MMP-9 activities. In situ zymograms using DQ-gelatin revealed vessel associated gelatinolytic activity in lesioned areas in PAI-1⁻/⁻ and in WT mice. In WT brain slices incubated ex vivo, glutamate (200 µM) induced DQ-gelatin activation in vessels. The effect was not detected in t-PA⁻/⁻ mice, but was restored by concomitant exposure to recombinant t-PA (20 µg/mL). In summary, neonatal brain lesion paradigms and ex vivo excitotoxic glutamate evoked t-PA-dependent gelatinases activation in vessels. Both MMP-2 and MMP-9 activities appeared t-PA-dependent. The data suggest that vascular directed protease inhibition may have neuroprotection potential against neonatal brain injuries

Age-Dependent Neonatal Intracerebral Hemorrhage in Plasminogen Activator Inhibitor 1 Knockout Mice

International audienc

Creating a National Biodiversity Database in Gabon and the Challenges of Mobilizing Natural History Data for Francophone Countries

Language is a major barrier for researchers wanting to digitize and publish collection data in Africa. Despite being the fifth most spoken language on Earth and the second most common in Africa, resources in French about digitization, data management, and publishing are lacking. Furthermore, French-speaking regions of Africa (primarily Central/West Africa and Madagascar) host some of the highest biodiversity on the continent and therefore are of great importance to scientists and decision-makers. Without having representation in online portals like the Global Biodiversity Information Facility (GBIF) and Integrated Digitized Biocollections (iDigBio), these important collections are effectively invisible. Producing relevant/applicable resources about digitization in French will help shine a light on these valuable natural history records and allow the data-holders in Africa to retain the autonomy of their collections. Awarded a GBIF-BID (Biodiversity Information for Development) grant in 2021, an international, multilingual network of partners has undertaken the important task of digitizing and mobilizing Gabon’s vertebrate collections. There are an estimated 13,500 vertebrate specimens housed in five institutions in different parts of Gabon. To date, the group has mobilized >4,600 vertebrate records to our recently launched Gabon Biodiversity Portal (https://gabonbiota.org/). The portal also hosts French guides for using Symbiota-based portals to manage, georeference, and publish natural history databases. These resources can provide much-needed guidance for other Francophone countries⁠—in Africa and beyond⁠—working to maximize the accessibility and value of their biodiversity collections.

Quantification of gelatinase activities in cortical extracts 6 hours after insults in P5 mice.

<p>MMP-2 (A,C,E) and MMP-9 (B,D,F) gelatinolytic activities measured in hypoxia-ischemia (A,B), ibotenate excitotoxic paradigm (C,D) and PBS injection trauma (E,F). Data are expressed relative to the average densitometry in WT mice. * indicate significant difference with activity in respective controls, i.e. WT sham operated (A,B), PBS injected (C,D) or non-treated animals (E,F). *p<0.05; **p<0.01; ***p<0.001 vs WT controls,^##p<0.001 and ^###p<0.001 vs PAI-1^−/− controls. Numbers in parentheses indicate the number of animals used. ND; non detected upon standard incubation.</p

<i>In situ</i> and <i>in vitro</i> gelatinolytic activity 5 days after PBS injection in WT and PAI-1^−/− mice.

<p>Effect of PBS intracortical injection in WT (A–C) or PAI-1^−/− (D–F) 5 days post-insult. Gelatinolytic activity remained elevated in PAI-1^−/− mice in cell nuclei, in extracellular spots and in vessels. (<b>G</b>) Gel zymogram of gelatinase activity in PAI-1^−/− mouse cortex 5 days after PBS injection at P5 and non-injected control. (<b>H</b>) Quantification of MMP-2 and MMP-9 activities in gels. Arrowheads point to spots of high activity; arrows point to microvessels. Numbers in parentheses indicate the number of animals used. *p<0.05 compared to corresponding controls, according to Student’<i>t</i> test.</p

<i>In situ</i> gelatinolytic activity labeling 6 h after hypoxic-ischemic procedure.

<p>Gelatinolytic activity (green) and vessel labeling by IB4 (red) were obtained in WT (A–D), t-PA^−/− (E–H) and PAI-1^−/− (I–L) mice. Low magnification observation of gelatinolytic activity (A,E,I). Note gross tissue alteration in PAI-1^−/− and low level of fluorescence in t-PA^−/− mice sections. Higher magnification allows to visualizing micro-vascularization. Gelatinolytic activity is hardly detectable on vessels in WT (B–D), while it is undetectable in t-PA^−/− (F–H) and high in PAI-1^−/− (J–L) mice. Arrowheads point to spots of high activity; arrows point to microvessels. ISZ; <i>in situ</i> zymographic activity; Ib4; isolectin B4 vessels labeling.</p

Vascular gelatinolytic activity labeling in 5 day-old mice brain sections exposed to glutamate and hrt-PA.

<p>Gelatinolytic activity (green) and vessel labeling by IB4 (red) were obtained in WT (A–F) or t-PA^−/− (G–L) mouse brain sections (250 µm thick) after 3 hours <i>ex vivo</i> exposure to control medium (A–C), glutamate 200 µM alone (D–I) or in association with 20 µg/mL hrt-PA (J–L). Arrows point out gelatinolytic activity in vessels. Bar = 200 µm.</p

<i>In situ</i> gelatinolytic activity labeling 6 h after ibotenate injection.

<p>Ibotenate was injected in WT (A–C) and in t-PA^−/− (D–F) mice and PBS (control) was injected in WT (G–I) and PAI-1^−/− (J–L) mice. Ibotenate induced high gelatinolytic activity spots in WT but not in t-PA^−/− mice (A,D), but did not induce activity in microvessels (C,F). PBS controls in WT did not exhibit enhanced gelatinolytic ativity (G–I) but a strong intracellular activity together with vessel activity in PAI-1^−/− (J–L) mice. Arrowheads point to spots of high activity; arrows point to microvessels. ISZ; <i>in situ</i> zymographic activity; Ib4; isolectin B4 vessels labeling.</p