Mesenchymal stem cells (MSCs) protect tissues against cell death induced by ischemia/reperfusion insults. This therapeutic effect seems to be controlled by physiological cues released by the local microenvironment following injury. Recent lines of evidence indicate that MSC can communicate with their microenvironment through bidirectional exchanges of mitochondria. In particular, in vitro and in vivo studies report that MSCs rescue injured cells through delivery of their own mitochondria. However, the role of mitochondria conveyed from somatic cells to MSC remains unknown. By using a co-culture system consisting of MSC and distressed somatic cells such as cardiomyocytes or endothelial cells, we showed that mitochondria from suffering cells acted as danger-signaling organelles that triggered the anti-apoptotic function of MSC. We demonstrated that foreign somatic-derived mitochondria were engulfed and degraded by MSC, leading to induction of the cytoprotective enzyme heme oxygenase-1 (HO-1) and stimulation of mitochondrial biogenesis. As a result, the capacity of MSC to donate their mitochondria to injured cells to combat oxidative stress injury was enhanced. We found that similar mechanisms – activation of autophagy, HO-1 and mitochondrial biogenesis – occurred after exposure of MSC to exogenous mitochondria isolated from somatic cells, strengthening the idea that somatic mitochondria alert MSC of a danger situation and subsequently promote an adaptive reparative response. In addition, the cascade of events triggered by the transfer of somatic mitochondria into MSC was recapitulated in a model of myocardial infarction in vivo. Specifically, MSC engrafted into infarcted hearts of mice reduced damage, upregulated HO-1 and increased mitochondrial biogenesis, while inhibition of mitophagy or HO-1 failed to protect against cardiac apoptosis. In conclusion, our study reveals a new facet about the role of mitochondria released from dying cells as a key environmental cue that controls the cytoprotective function of MSC and opens novel avenues to improve the effectiveness of MSC-based therapies.

M itochondria and the endoplasmic reticulum (ER) are separately considered key players in cell death signaling.1 Mitochondria and ER are interconnected organelles and form an endomembrane network. The contact points through which ER communicates with mitochondria are referred to as mitochondria-associated membranes (MAM).2 MAM are enriched in phospholipid-and glycosphingolipid-synthesis enzymes, as well as chaperone proteins, which transport lipids and exchange calcium between these 2 organelles.1 Several recent studies have identified new proteins enriched at the ER-mitochondria interface, highlighting the emerging understanding of the role of this region within the cell.3,4 One of them has identified a macromolecular complex composed of VDAC1, Grp75, and IP3R1 that regulates direct Ca 2+ transfer from ER to mitochondria. 5 Indeed, ER-mitochondria junctions are aligned with mitochondrial contact points where VDAC1 is abundantly present, thus creating a hot spot for the Ca 2+ transfer from the ER. 6 Although the role of this organelle cross talk is beginning to be understood in cell physiology, MAM involvement in cardiac pathologies remains unknown. Clinical Perspective on p 1565Calcium signaling is central for heart function through its physiological role in excitation-contraction coupling and the detrimental impact of Ca 2+ overload during heart failure and myocardial ischemia/reperfusion. During this latter condition, it is well accepted that the cytosolic accumulation of Ca 2+ subsequently results in mitochondrial Ca 2+ overload,Background-Under physiological conditions, Ca 2+ transfer from the endoplasmic reticulum (ER) to mitochondria might occur at least in part at contact points between the 2 organelles and involves the VDAC1/Grp75/IP3R1 complex. Accumulation of Ca 2+ into the mitochondrial matrix may activate the mitochondrial chaperone cyclophilin D (CypD) and trigger permeability transition pore opening, whose role in ischemia/reperfusion injury is well recognized. We questioned here whether the transfer of Ca 2+ from ER to mitochondria might play a role in cardiomyocyte death after hypoxia-reoxygenation. Methods and Results-We report that CypD interacts with the VDAC1/Grp75/IP3R1 complex in cardiomyocytes. Genetic or pharmacological inhibition of CypD in both H9c2 cardiomyoblasts and adult cardiomyocytes decreased the Ca 2+ transfer from ER to mitochondria through IP3R under normoxic conditions. During hypoxia-reoxygenation, the interaction between CypD and the IP3R1 Ca 2+ channeling complex increased concomitantly with mitochondrial Ca 2+ content. Inhibition of either CypD, IP3R1, or Grp75 decreased protein interaction within the complex, attenuated mitochondrial Ca 2+ overload, and protected cells from hypoxia-reoxygenation. Genetic or pharmacological inhibition of CypD provided a similar effect in adult mice cardiomyocytes. Disruption of ER-mitochondria interaction via the downregulation of Mfn2 similarly reduced the interaction between CypD and the IP3R1 complex and protected against hypoxia-reo...

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Objective: To assess the relationship between matrix metalloproteinase 9 (MMP-9), a proteolytic enzyme involved in the breakdown of the blood-brain barrier, and infarct growth and hemorrhagic transformation in acute ischemic stroke (AIS) with large vessel occlusion (LVO) in the era of mechanical thrombectomy (MT) using the kinetics of MMP-9 and sequential magnetic resonance imaging (MRI).Methods: HIBISCUS-STROKE is a cohort study including AIS patients with LVO treated with MT following admission MRI. Patients underwent sequential assessment of MMP-9, follow-up CT at day 1, and MRI at day 6. The CT scan at day 1 classified any hemorrhagic transformation according to the European Co-operative Acute Stroke Study-II (ECASS II) classification. Infarct growth was defined as the difference between final Fluid-Attenuated Inversion Recovery volume and baseline diffusion-weighted imaging volume. Conditional logistic regression analyses were adjusted for main confounding variables including reperfusion status.Results: One hundred and forty-eight patients represent the study population. A high MMP-9 level at 6 h from admission (H6) (p = 0.02), a high glucose level (p = 0.01), a high temperature (p = 0.04), and lack of reperfusion (p = 0.02) were associated with infarct growth. A high MMP-9 level at H6 (p = 0.03), a high glucose level (p = 0.03) and a long delay from symptom onset to groin puncture (p = 0.01) were associated with hemorrhagic transformation. Conclusions:In this MT cohort study, MMP-9 level at H6 predicts infarct growth and hemorrhagic transformation.

Rationale & aim: Various types of cell therapies are currently under investigation for the treatment of ischemic stroke patients. To bridge the gap between cell administration and therapeutic outcome, there is a need for non-invasive monitoring of these innovative therapeutic approaches. Spectral photon counting computed tomography (SPCCT) is a new imaging modality that may be suitable for cell tracking. SPCCT is the next generation of clinical CT that allows the selective visualization and quantification of multiple contrast agents. The aims of this study are: (i) to demonstrate the feasibility of using SPCCT to longitudinally monitor and quantify therapeutic cells, i.e. bone marrow-derived M2-polarized macrophages transplanted in rats with brain damage; and (ii) to evaluate the potential of this approach to discriminate M2-polarized macrophages from their encapsulating scaffold. Methods: Twenty one rats received an intralesional transplantation of bone marrow-derived M2-polarized macrophages. In the first set of experiments, cells were labeled with gold nanoparticles and tracked for up to two weeks post-injection in a monocolor study via gold K-edge imaging. In the second set of experiments, the same protocol was repeated for a bicolor study, in which the labeled cells are embedded in iodine nanoparticle-labeled scaffold. The amount of gold in the brain was longitudinally quantified using gold K-edge images reconstructed from SPCCT acquisition. Animals were sacrificed at different time points post-injection, and ICP-OES was used to validate the accuracy of gold quantification from SPCCT imaging. Results: The feasibility of therapeutic cell tracking was successfully demonstrated in brain-damaged rats with SPCCT imaging. The imaging modality enabled cell monitoring for up to 2 weeks post-injection, in a specific and quantitative manner. Differentiation of labeled cells and their embedding scaffold was also feasible with SPCCT imaging, with a detection limit as low as 5,000 cells in a voxel of 250 × 250 × 250 µm in dimension in vivo. Conclusion: Multicolor SPCCT is an innovative translational imaging tool that allows monitoring and quantification of therapeutic cells and their encapsulating scaffold transplanted in the damaged rat brain.

ObjectiveTo assess whether interleukin-6 (IL-6) level is a marker of futile reperfusion in acute ischemic stroke (AIS) patients with large vessel occlusion (LVO) treated with mechanical thrombectomy (MT).MethodsHIBISCUS-STROKE (CoHort of Patients to Identify Biological and Imaging markerS of CardiovascUlar Outcomes in Stroke) includes AIS patients treated with MT after magnetic resonance imaging (MRI). We performed a sequential assessment of IL-6 (admission, 6 hours, 24 hours, 48 hours and 3 months from admission). Among patients with successful reperfusion (thrombolysis in cerebral infarction scale 2b/3), reperfusion was considered as effective if 3-month modified Rankin scale (mRS) score was 0–2 and futile if 3-month mRS score was 3–6. Our model was adjusted for the main confounding variables.ResultsOne hundred and sixty-four patients represent the study population. One hundred and thirty-three patients had successful reperfusion (81.1%) while in 46 (34.6%), this one was classified as futile. In single-variable analyses, high IL-6 levels at 6, 24 and 48 hours, in combination with a higher age, a pre-stroke mRS score >2, a past history of hypertension or diabetes, current smoking, a higher baseline National Institute of Health Stroke Scale Score, the absence of associated intravenous thrombolysis, an intracranial internal carotid artery or a tandem occlusion and an increased infarct growth were associated with futile reperfusion. Following multivariable analyses, a high IL-6 level at 24 hours (odds ratio 6.15, 95% confidence interval 1.71–22.10) remained associated with futile reperfusion.ConclusionsIL-6 is a marker of futile reperfusion in the setting of MT.

SummaryAn original model of organo-specific, immortalized and stabilized endothelial cell lines was used to delineate the part played by some chemokines (CCL21, CX3CL1, CCL5 and CXCL12) and their receptors in endothelium organo-specificity. Chemokine receptor expression and chemokine presentation were investigated on organo-specific human endothelial cell lines. Although the chemokines showed distinct binding patterns for the various endothelial cell lines, these were not correlated with the expression of the corresponding receptors (CX3CR1, CXCR4, CCR5 and CCR7). Experiments with CCL21 on peripheral lymph node endothelial cells demonstrated that the chemokine did not co-localize with its receptor but was associated with extracellular matrix components. The specific activity of chemokines was clearly shown to be related to the endothelial cell origin. Indeed, CX3CL1 and CCL21 promoted lymphocyte recruitment by endothelial cells from the appendix and peripheral lymph nodes, respectively, while CX3CL1 pro-angiogenic activity was restricted to endothelial cells from the appendix and skin. The high specificity of the chemokine/endothelium interaction allowed the design of a direct in vitro endothelial cell targeting assay. This unique cellular model demonstrated a fundamental role for chemokines in conferring on the endothelium its organo-specificity and its potential for tissue targeting through the selective binding, presentation and activation properties of chemokines.