The biological sample collection of the OFSEP French MS registry: An essential tool dedicated to researchers.

Today's medicine strives to be personalized, preventive, predictive and participatory. This implies to have access to multimodal data to better characterize patients groups and to combine clinical and imaging data with high-quality biological samples. Collecting such data is one of the objectives of the Observatoire français de la sclérose en plaques (OFSEP), the French MS registry. On December 2022, the OFSEP biocollection includes 4,888 patients with scientific characteristics and about 90,000 samples. Thanks to its richness, this biocollection open for the scientific community, contributes to address unmet needs in MS through identification of multiomics determinants of MS activity, progression and secondary effects.

A lower admission level of interleukin-6 is associated with first-pass effect in ischemic stroke patients.

BACKGROUND: First-pass effect (FPE) defined as a complete or near-complete reperfusion achieved after a single thrombectomy pass is predictive of favorable outcome in acute ischemic stroke (AIS) patients. We aimed to assess whether admission levels of inflammatory markers are associated with FPE. METHODS: HIBISCUS-STROKE (CoHort of Patients to Identify Biological and Imaging markerS of CardiovascUlar Outcomes in Stroke) includes AIS patients with large vessel occlusion treated with mechanical thrombectomy following brain MRI. C-reactive protein, interleukin (IL)-6, IL-8, IL-10, monocyte chemoattractant protein-1, soluble tumor necrosis factor receptor I, soluble form suppression of tumorigenicity 2, matrix metalloproteinase-9 (MMP-9), soluble P-selectin, and vascular cellular adhesion molecule-1 were measured in admission sera using an ELISA assay. FPE was defined as a complete or near-complete reperfusion (thrombolysis in cerebral infarction scale (TICI) 2c or 3) after the first pass. A multivariate logistic regression analysis was performed to assess independent factors associated with FPE. RESULTS: A total of 151 patients were included. Among them, 43 (28.5%) patients had FPE. FPE was associated with low admission levels of IL-6, MMP-9, and platelet count, an older age, lack of hypertension, lack of tandem occlusion, a shorter thrombus length, and a reduced procedural time. Following multivariate analysis, a low admission level of IL-6 was associated with FPE (OR 0.66, 95% CI 0.46 to 0.94). Optimal cut-off of IL-6 level for distinguishing FPE from non-FPE was 3.0 pg/mL (sensitivity 92.3%, specificity 42.3%). CONCLUSION: A lower admission level of IL-6 is associated with FPE.

Colchicine for Left Ventricular Infarct Size Reduction in Acute Myocardial Infarction: A Phase II, Multicenter, Randomized, Double-Blinded, Placebo-Controlled Study Protocol - The COVERT-MI Study.

Inflammatory processes have been identified as key mediators of ischemia-reperfusion injury in ST-segment elevation myocardial infarction (STEMI). They add damage to the myocardium and are associated with clinical adverse events (heart failure and cardiovascular death) and poor myocardial recovery. Colchicine is a well-known alkaloid with potent anti-inflammatory properties. In a proof-of-concept phase II trial, colchicine has been associated with a significant 50% reduction of infarct size (assessed by creatine kinase levels) in comparison to placebo in acute STEMI patients referred for primary percutaneous coronary intervention (PPCI). The Colchicine in STEMI Patients Study (COVERT-MI) is an ongoing confirmative prospective, multicenter, randomized, double-blind trial testing whether a short course oral treatment with colchicine versus placebo decreases myocardial injury in patients presenting with STEMI referred for PPCI. Adult patients, with a first STEMI episode and an initial TIMI flow ≤1, referred for PPCI, will be randomized (n = 194) in a 1:1 ratio to receive an oral bolus of colchicine of 2 mg followed by 0.5 mg b.i.d. treatment during 5 days or matching placebo. The primary endpoint will be the reduction in infarct size as assessed by cardiac magnetic resonance at 5 ± 2 days between both groups. The main secondary endpoints will be tested between groups in hierarchical order with left ventricular ejection fraction at 5 days, microvascular obstruction presence at 5 days, and absolute adverse left ventricular remodeling between 5 days and 3 months. This academic study is being financed by a grant from the French Ministry of Health (PHRCN-16-0357). Results from this study will contribute to a better understanding of the complex pathophysiology underlying myocardial injury after STEMI. The present study describes the rationale, design, and methods of the trial.

Association of myocardial hemorrhage and persistent microvascular obstruction with circulating inflammatory biomarkers in STEMI patients.

INTRODUCTION: Myocardial hemorrhage (IMH) and persistent microvascular obstruction (MVO) are associated with impaired myocardial recovery and adverse clinical outcomes in STEMI patients. However, their relationship with circulating inflammatory biomarkers is unclear in human patients. METHODS AND RESULTS: Twenty consecutive patients referred for primary percutaneous coronary intervention of first STEMI were included in a prospective study. Blood sampling was performed at admission, 4, 12, 24, 48 hours, 7 and 30 days after reperfusion for inflammatory biomarker (C reactive protein, fibrinogen, interleukin-6 (IL-6) and neutrophils count) assessment. At seven days, cardiovascular magnetic resonance (CMR) was performed for infarct size, MVO and IMH assessment. Median infarct size was 24.6% Interquartile range (IQR) [12.0-43.5] of LV mass and edema was 13.2% IQR [7.7-36.1] of LV mass. IL-6 reached a peak at H24 (5.6 pg/mL interquartile range (IQR) [2.5-17.5]), CRP at H48 (11.7 mg/L IQR [7.1-69.2]), fibrinogen one week after admission (4.4 g/L IQR [3.8-6.7]) and neutrophils at H12 (9.0 G/L IQR [6.5-12.7]). MVO was present in 11 patients (55% of the study population) and hemorrhage in 7 patients (35%). Patients with IMH had significantly higher IL-6, CRP, fibrinogen, and neutrophils levels compared to patients without IMH. Patients with persistent MVO had significantly higher CRP, fibrinogen and neutrophils level compared to patients without MVO, but identical IL-6 kinetics. CONCLUSION: In human patients with acute myocardial infarction, intramyocardial hemorrhage appears to have a stronger relationship with inflammatory biomarker release compared to persistent MVO. Attenuating myocardial hemorrhage may be a novel target in future adjunctive STEMI treatments.

Association of Interleukin-6 Levels and Futile Reperfusion After Mechanical Thrombectomy.

OBJECTIVE: To assess whether interleukin-6 (IL-6) level is a marker of futile reperfusion in patients with acute ischemic stroke (AIS) with large vessel occlusion treated with mechanical thrombectomy (MT). METHODS: The Cohort of Patients to Identify Biological and Imaging Markers of Cardiovascular Outcomes in Stroke (HIBISCUS-STROKE) includes patients with AIS treated with MT after 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 effective if 3-month modified Rankin Scale (mRS) score was 0 to 2 and futile if 3-month mRS score was 3 to 6. Our model was adjusted for the main confounding variables. RESULTS: One hundred sixty-four patients represent the study population. One hundred thirty-three patients had successful reperfusion (81.1%), while in 46 (34.6%), reperfusion 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 prestroke mRS score >2, a history of hypertension or diabetes, lack of current smoking, a higher baseline NIH 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. After 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. CONCLUSIONS: IL-6 is a marker of futile reperfusion in the setting of MT.

Successful Leptospira genotyping strategy on DNA extracted from canine biological samples.

Leptospirosis is an emerging worldwide zoonosis with a changing epidemiology responsible for an acute disease in humans and dogs. A better knowledge of the responsible bacterium Leptospira and in particular its various serovars and serogroups prevalence is essential for better diagnosis and prevention of the disease. The gold standard for leptospirosis diagnosis is the Microscopic Agglutination Test (MAT) but it requires long and fastidious laboratory work and sometimes results in controversial data. For these reasons, PCR-based techniques for detection of pathogenic leptospiral DNA in biological samples are currently replacing the MAT. However, these strategies do not provide any information regarding the infecting serovar or serogroup. In this study, an optimized genotyping method is described to allow the identification of Leptospira ssp. directly at serovars level using DNA extracted from canine blood and urine. 16S rDNA, Variable Number Tandem Repeat (VNTR) and Multispacer Sequence Typing (MST) protocols were adapted to biological samples. Eighty-eight DNA samples were analyzed from 72 different European canine clinical cases of leptospirosis confirmed by real-time PCR. 92% of DNA samples with Ct values below 34 were fully typed, and typing success decreased to about 30% for the other samples. Typing failure also showed a specie-specific correlation, with 63% of complete typing for L. interrogans and only 40% for L. kirschneri. Additionally, an exact match was observed between serological and molecular data for the few investigated cases where MAT data were available. This methodology is a suitable alternative to the MAT for determining the infecting serovar when Leptospira DNA from blood or urine is detected at Ct values below 34, contributing to clinical surveillance of leptospirosis.

Matrix Metalloproteinase-9 Relationship With Infarct Growth and Hemorrhagic Transformation in the Era of Thrombectomy.

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.

Matrix Metalloproteinase-9 and Monocyte Chemoattractant Protein-1 Are Associated With Collateral Status in Acute Ischemic Stroke With Large Vessel Occlusion.

BACKGROUND AND PURPOSE: In ischemic stroke, inflammatory status may condition the development of collateral circulation. Here we assessed the relationship between systemic inflammatory biomarkers and collateral status in large vessel occlusion before mechanical thrombectomy. METHODS: HIBISCUS-STROKE is a cohort study including acute ischemic stroke patients with large vessel occlusion treated with mechanical thrombectomy following admission magnetic resonance imaging. MMP-9 (matrix metalloproteinase-9) and MCP-1 (monocyte chemoattractant protein-1) were measured on blood sampling collected at admission. Collateral status was assessed on pretreatment Digital subtraction angiography and categorized into poor (Higashida score, 0-2) and good (Higashida score, 3-4). A multiple logistic regression model was performed to detect independent markers of good collateral status. RESULTS: One hundred and twenty-two patients were included, of them 71 patients (58.2%) had a good collateral status. In univariate analysis, low MMP-9 levels (P=0.01), high MCP-1 levels (P<0.01), a low National Institute of Health Stroke Score (P=0.046), a high diastolic blood pressure (P=0.049), the absence of tandem occlusion (P=0.046), a high Alberta Stroke Program Early CT Score (P<0.01) and a low volume on the diffusion-weighted imaging (P<0.01) were associated with good collateral status. Following multivariate analysis, low MMP-9 levels (P=0.02) and high MCP-1 levels (P<0.01) remained associated with good collateral status. CONCLUSIONS: Low MMP-9 and high MCP-1 levels were associated with good pretreatment collateral status in patients with acute ischemic stroke with large vessel occlusion. These results might suggest a relationship between collateral status and inflammation.

Neprilysin levels at the acute phase of ST-elevation myocardial infarction.

BACKGROUND: Several preliminary analyses suggested an association between neprilysin (NEP) levels and myocardial infarction. HYPOTHESIS: The objective was to assess whether NEP plasma levels following reperfusion might be a surrogate for infarct size (IS) or predict adverse outcomes in acute ST-segment elevation myocardial infarction (STEMI) patients. METHODS: We measured NEP levels in a prospective cohort of 203 patients with STEMI referred for primary percutaneous coronary intervention. Circulating soluble NEP was measured by enzyme-linked immunosorbent assay at admission (t0) and 4 hours later (t4) following reperfusion and on 7 times points (t0, t4, t12, t24, t48, day 7 and day 30) in a subset of 21 patients. IS and left ventricular ejection fraction (LVEF) were measured at 1 month by cardiac magnetic resonance. Adverse cardiovascular outcomes were collected at 12-month follow-up. RESULTS: Median t0 and t4 NEP levels in 203 patients were respectively 88.3 pg/mL (interquartile range [IQR] [14; 375.4]) and 101.5 pg/mL (IQR [18.5; 423.8]). These levels remained unchanged over 1 month (P = 0.70). NEP levels did not correlate significantly with IS (P = 0.51) or LVEF (P = 0.34). There was no correlation between NEP and troponin, creatine kinase and interleukin-6 levels at h0 and h4. NEP levels above the median were not associated with adverse outcomes at follow-up (hazard ratio = 1.28, 95% confidence interval [0.69; 2.37]; P = 0.42). CONCLUSIONS: NEP serum levels were widely distributed and did not change significantly in the first hours and 1-month period following reperfusion in STEMI patients. There was no significant relationship with markers of infarct size and inflammation, and 1-year adverse outcomes.

Early kinetics of serum Interleukine-17A and infarct size in patients with reperfused acute ST-elevated myocardial infarction.

BACKGROUND: Recently, it was shown that interleukin-17A (IL-17A) is involved in the pathophysiology of reperfusion injury and associated with infarct size (IS) in experimental models of myocardial infarction. Our aim was to evaluate whether the IL-17A serum level and the IL-17A active fraction was correlated with IS in humans. METHODS: 101 patients presenting with a ST-elevated Myocardial Infarction (STEMI) referred for primary percutaneous coronary intervention (PPCI) and 10 healthy controls were included. For each participant, blood samples at admission (H0) and 4 hours after admission (H4) were collected. IL-17A serum levels were assessed using ELISA and the active fraction was assessed with a functional test. IS was determined by peak troponin and peak CK levels for every patient and by contrast-enhanced cardiac magnetic resonance (ce-CMR) for 20 patients. RESULTS: The IL-17A serum level was significantly increased in STEMI patients compared to healthy controls, (0.9 pg/mL IQR [0.0-3.2] at H0 and 1.0 pg/mL IQR [0.2-2.8] at H4 versus 0.2 pg/mL IQR [0.0-0.7] for healthy controls; p<0.005). At either time points, IL-17A levels did not correlate with IS as measured by peak troponin, peak CK pr ce-CMR. Also, no correlation was found between the active fraction of IL-17A and IS. CONCLUSION: Serum IL-17A level is significantly increased in patients at the early phase of acute MI compared to healthy controls. However, the level of IL-17A in the early hours after reperfusion does not correlate with IS.

Use of flow cytometry for rapid and accurate enumeration of live pathogenic Leptospira strains.

Enumeration of Leptospira, the causative agent of leptospirosis, is arduous mainly because of its slow growth rate. Rapid and reliable tools for numbering leptospires are still lacking. The current standard for Leptospira cultures is the count on Petroff-Hausser chamber under dark-field microscopy, but this method remains time-consuming, requires well-trained operators and lacks reproducibility. Here we present the development of a flow-cytometry technique for counting leptospires. We showed that upon addition of fluorescent dyes, necessary to discriminate the bacterial population from debris, several live Leptospira strains could be enumerated at different physiologic states. Flow cytometry titers were highly correlated to counts with Petroff-Hausser chambers (R2>0.99). Advantages of flow cytometry lie in its rapidity, its reproducibility significantly higher than Petroff-Hausser method and its wide linearity range, from 104 to 108leptospires/ml. Therefore, flow cytometry is a fast, reproducible and sensitive tool representing a promising technology to replace current enumeration techniques of Leptospira in culture. We were also able to enumerate Leptospira in artificially infected urine and blood with a sensitivity limit of 105leptospires/ml and 106leptospires/ml, respectively, demonstrating the feasibility to use flow cytometry as first-line tool for diagnosis or bacterial dissemination studies.

Rabies DNA vaccine in the horse: strategies to improve serological responses.

In order for DNA vaccines to become a practical alternative to conventional vaccines their ability to induce antibody responses in large mammals needs to be improved. We used DNA vaccination against rabies in the horse as a model to test the potential of two different strategies to enhance antibody responses in a large mammalian species. The administration of the DNA vaccine in the presence of aluminum phosphate improved both the onset and the intensity of serological responses but was not potent enough to achieve seroconversion in all vaccinated ponies. However, when the DNA vaccine was formulated with the cationic lipid DMRIE-DOPE instead of aluminum phosphate, a very strong impact on both onset and intensity of serological responses was observed. This latter strategy ensured excellent seroconversion in all vaccinated ponies after a primary course of two injections, demonstrating a clear improvement of the homogeneity of the induced responses. These data indicate that rabies DNA vaccination is feasible in horses and further suggests that properly formulated DNA vaccines can generate immune responses in large veterinary species at a level comparable to the responses achieved with conventional vaccines.