Biology subjects

Guimbal, S.

Publications and source records attributed to Guimbal, S..

6 recordsLinked to original sources

bioRxiv2026

Extracellular methylglyoxal; the passage across brain endothelial cells and the effect on barrier function

BackgroundMethylglyoxal (MGO), a highly reactive by-product of glycolysis, has been associated with cognitive decline and Alzheimers disease, though the mechanistic role of MGO remains unclear. Moreover, conflicting findings exist regarding MGOs toxicity on the blood-brain barrier (BBB). This study investigated whether MGO can cross the BBB under physiologically relevant conditions and whether MGO affects BBB permeability. MethodsMice were intravenously injected with highly purified home-made MGO or PBS, and MGO concentration was measured at five timepoints in the cerebral cortex up to 4 hours after injection. MGO toxicity was screened on a human brain endothelial cell line (hCMEC/D3) using a live/dead assay prior to the study of selected MGO concentrations and on hiPSC-derived brain microvascular endothelial cells (EECM-BMECs). EECM-BMECs were cultured on Transwell(R) inserts, and barrier function was assessed by sodium fluorescein permeability and transendothelial passage of 13C3-MGO quantified by UPLC-MS/MS. ResultsMGO levels in the mouse cortex did not increase post-injection. MGO was not toxic to hCMEC/D3 cells, and it had no impact on barrier properties of EECM-BMECs. After 1-hour exposure, [~]13% of total 13C3-MGO was recovered in its free form, and only [~]1% of supplemented MGO was recovered from the abluminal side. ConclusionMGO does not cross the BBB in vivo and does not affect barrier properties of a human in vitro model of the BBB. In vitro MGO passage across the BBB is minimal. These findings suggest that circulating MGO is unlikely to directly affect neuronal function via BBB disruption or enter the brain in its free from.

neuroscience↗

bioRxiv2020

Mast cells participate in the development of diastolic dysfunction in diabetic obese mice

Rational Heart failure with preserved ejection fraction (HFpEF) is a growing epidemiologic issue. However, to date, its pathophysiology remains poorly understood.Objective Our goal was to investigate the role of microvessel disease in the pathophysiology of diastolic dysfunction.Findings To do so, we used Leptin receptor deficient (Leprdb/db) female mice as a model of diastolic dysfunction. In these mice, the increased end diastolic pressure (EDP) signing diastolic dysfunction is associated with vascular leakage, endothelial cell activation and leucocyte infiltration. Strikingly, a RNA sequencing analysis of the cardiac vascular fraction of both Leprdb/db and control female mice confirmed endothelial dysfunction and systemic inflammation but also revealed a strong increase in several mast cell markers (notably FceR1a, Tryptase and Chymase). We then histologically confirmed an accumulation of activated mast cells in the heart of Leprdb/db mice. Importantly, mast cell degranulation inhibition reduced EDP, vascular leakage and leucocyte infiltration in Leprdb/db mice.Conclusion Mast cells play a critical role in the development of cardiac microvessel disease and diastolic dysfunction.Competing Interest StatementThe authors have declared no competing interest.View Full Text

physiology↗

bioRxiv2020

Desert Hedgehog-driven endothelium integrity is enhanced by Gas1 but negatively regulated by Cdon

Evidences accumulated within the past decades, identified Hedgehog (Hh) signaling as a new regulator of micro-vessel integrity. More specifically, we recently identified Desert Hedgehog (Dhh) as a downstream effector of Klf2 in endothelial cells (ECs). ObjectiveThe purpose of this study is to investigate whether Hh co-receptors Gas1 and Cdon may be used as therapeutic targets to modulate Dhh signaling in ECs. Methods and resultsWe demonstrated that both Gas1 and Cdon are expressed in adult ECs and relied on either siRNAs or EC specific conditional KO mice to investigate their role. We found that Gas1 deficiency mainly photocopies Dhh deficiency especially by inducing VCAM-1 and ICAM-1 overexpression while Cdon deficiency has opposite effects by promoting endothelial junction integrity. At a molecular level, Cdon prevents Dhh binding to Ptch1 and thus acts a decoy receptor for Dhh, while Gas1 promotes Dhh binding to Smo and as a result potentiates Dhh effects. Since Cdon is overexpressed in ECs treated by inflammatory cytokines including TNF and Il1{beta}, we then tested whether Cdon inhibition would promote endothelium integrity in acute inflammatory conditions and found that both fibrinogen and IgG extravasation were decreased in association with an increased Cdh5 expression in the brain cortex of EC specific Cdon KO mice administered locally with Il1{beta}. ConclusionAltogether these results demonstrate that Gas1 is a positive regulator of Dhh in ECs while Cdon is a negative regulator. Interestingly Cdon blocking molecules may then be used to promote endothelium integrity at least in inflammatory conditions.

cell biology↗

bioRxiv2020

Genetic disruption of the Blood Brain Barrier leads to protective barrier formation at the Glia Limitans

Recent work demonstrated that Central Nervous System (CNS) inflammation induces endothelial Blood Brain Barrier (BBB) opening as well as the formation of a tight junction barrier between reactive astrocytes at the Glia Limitans. We hypothesized that these two barriers may be reciprocally regulated by each other state and further, that the CNS parenchyma may acquire protection from the reactive astrocytic Glia Limitans not only in neuro-inflammation but also when BBB integrity is compromised under resting condition, without pathology. Previous studies identified Sonic hedgehog (Shh) astrocytic secretion as implicated in stabilizing the BBB during neuropathology and we recently demonstrated that desert hedgehog (Dhh) is expressed at the BBB in adults. Here we unraveled the role of the morphogen Dhh in maintaining BBB tightness and, using endothelial Dhh knockdown as a model of permeable BBB, we demonstrated that a double barrier system comprising both the BBB and Glia Limitans, is implemented in the CNS and regulated by a crosstalk going from endothelial cell to astrocytes. First, we showed that, under neuro-inflammatory conditions, Dhh expression is severely down regulated at the BBB and that Dhh is necessary for endothelial intercellular junction integrity as Dhh knockdown leads to CNS vascular leakage. We then demonstrated that, in Dhh endothelial knockout (DhhECKO) mice which display an open BBB, astrocytes are reactive and express the tight junction Claudin 4 (Cldn4) and showed that astrocytes can respond to signals secreted by the permeable endothelial BBB by becoming reactive and expressing Cldn4. To examine the consequences of the above results on disease severity, we finally induced multiple sclerosis in DhhECKO mice versus control littermates and showed that the pathology is less severe in the knockout animals due to Glia Limitans tightening, in response to BBB leakage, which drives inflammatory infiltrate entrapment into the perivascular space. Altogether these results suggest that genetic disruption of the BBB generates endothelial signals capable of driving the implementation of a secondary barrier at the Glia Limitans to protect the parenchyma. The concept of a reciprocally regulated CNS double barrier system has implications for treatment strategies in both the acute and chronic phases of multiple sclerosis pathophysiology.

neuroscience↗

bioRxiv2020

Endothelial cell response to Hedgehog ligands depends on their processing

RationaleThe therapeutic potential of Hedgehog (Hh) signaling agonists for vascular diseases is of growing interest. However, molecular and cellular mechanisms underlying the role of the Hh signaling in vascular biology remain poorly understood. ObjectiveThe purpose of the present paper is to clarify some conflicting literature data. FindingsWith this goal we have demonstrated that, unexpectedly, ectopically administered N-terminal Sonic Hedgehog (N-Shh) and endogenous endothelial-derived Desert Hedgehog (Dhh) induce opposite effects in endothelial cells. (ECs). Notably, endothelial Dhh acts under its full-length soluble form (FL-Dhh) and activates Smoothened in ECs, while N-Shh inhibits it. At molecular level, N-Shh prevents FL-Dhh binding to Patched-1 demonstrating that N-Shh acts as competitive antagonist to FL-Dhh. Besides, we found that even though FL-Hh ligands and N-Hh ligands all bind Patched-1, they induce specific Patched-1 localization. Finally, we confirmed that in a pathophysiological setting i.e. brain inflammation, astrocyte-derived N-Shh act as a FL-Dhh antagonist. ConclusionThe present study highlights for the first time that FL-Dhh and N-Hh ligands have antagonistic properties especially in ECs, and demonstrates that Hh ligands or forms of Hh ligands cannot be used instead of another for therapeutic purposes.

physiology↗

bioRxiv2019

Dock5 is a new regulator of microtubule dynamic instability through GSK3β inhibition in osteoclasts

Background informationOsteoclast resorption is dependent on a podosome-rich structure, called sealing zone, which is stabilized by acetylated microtubules. It tightly attaches the osteoclast to the bone creating a favorable acidic microenvironment for bone degradation. We already established that Rac activation by Dock5 is necessary for osteoclast resorption. Indeed, inhibition of Dock5 in osteoclasts results in Rac1 decreased activity associated to impaired podosome assembly into sealing zones and resorbing activity.\n\nResultsIn this report, we show that Dock5 knockout osteoclasts also present a reduced acetylated tubulin level leading to a decreased length and duration of microtubule growth phases whereas their growth speed remains unaffected. Dock5 does not act by direct interaction with the polymerized tubulin but through inhibition of the microtubules destabilizing kinase GSK3{beta} downstream of Akt activation. Interestingly, we ruled out the implication of Rac1 in this process using specific inhibitors.\n\nConclusionOur data involve Dock5 as a new regulator of microtubule dynamic instability in osteoclast.\n\nSignificanceThe fact that Dock5 is a regulator of both actin cytoskeleton and microtubule dynamics makes it an interesting therapeutic target for osteolytic pathologies because of its dual role on sealing zone formation and stabilization.

cell biology↗