Lacunar infarcts represent one of the most frequent subtypes of ischemic strokes and may represent the first recognizable manifestation of a progressive disease of the small perforating arteries, capillaries, and venules of the brain, defined as cerebral small vessel disease. The pathophysiological mechanisms leading to a perforating artery occlusion are multiple and still not completely defined, due to spatial resolution issues in neuroimaging, sparsity of pathological studies, and lack of valid experimental models. Recent advances in the endovascular treatment of large vessel occlusion may have diverted attention from the management of patients with small vessel occlusions, often excluded from clinical trials of acute therapy and secondary prevention. However, patients with a lacunar stroke benefit from early diagnosis, reperfusion therapy, and secondary prevention measures. In addition, there are new developments in the knowledge of this entity that suggest potential benefits of thrombolysis in an extended time window in selected patients, as well as novel therapeutic approaches targeting different pathophysiological mechanisms involved in small vessel disease. This review offers a comprehensive update in lacunar stroke pathophysiology and clinical perspective for managing lacunar strokes, in light of the latest insights from imaging and translational studies.

The link atom (LA) and the generalized hybrid orbital (GHO) quantum mechanical/molecular mechanics (QM/MM) boundary treatment methods are compared, in the context of the acylation process (the rate-limiting step) involving the NS3/NS4A HCV serine protease and its NS5A/5B natural substrate. The potential energy surface was calculated, and the free energy along the selected reaction coordinate was obtained from umbrella sampling molecular dynamics simulations, at the AM1/CHARMM27 level. The LA and GHO methods, when properly applied, lead to similar chemical behavior, although the agreement is not quantitative. The choice of QM/MM partitioning is dictated to some extent by the nature of the two different methods, and this influences the results. The free energy profiles obtained by umbrella sampling suggest that the GHO approach is better suited for this system, because it provides a consistent description of the reaction in both the forward and backward directions. This is probably a consequence of the different QM/MM partitioning required by the two different methods (i.e., different numbers of atoms have to be included in the QM region). This finding is therefore likely to be system dependent, so careful testing should be considered for each enzyme application.

Background and Purpose: Acute ischemic stroke and large vessel occlusion can be concurrent with the coronavirus disease 2019 (COVID-19) infection. Outcomes after mechanical thrombectomy (MT) for large vessel occlusion in patients with COVID-19 are substantially unknown. Our aim was to study early outcomes after MT in patients with COVID-19. Methods: Multicenter, European, cohort study involving 34 stroke centers in France, Italy, Spain, and Belgium. Data were collected between March 1, 2020 and May 5, 2020. Consecutive laboratory-confirmed COVID-19 cases with large vessel occlusion, who were treated with MT, were included. Primary investigated outcome: 30-day mortality. Secondary outcomes: early neurological improvement (National Institutes of Health Stroke Scale improvement ≥8 points or 24 hours National Institutes of Health Stroke Scale 0–1), successful reperfusion (modified Thrombolysis in Cerebral Infarction grade ≥2b), and symptomatic intracranial hemorrhage. Results: We evaluated 93 patients with COVID-19 with large vessel occlusion who underwent MT (median age, 71 years [interquartile range, 59–79]; 63 men [67.7%]). Median pretreatment National Institutes of Health Stroke Scale and Alberta Stroke Program Early Computed Tomography score were 17 (interquartile range, 11–21) and 8 (interquartile range, 7–9), respectively. Anterior circulation acute ischemic stroke represented 93.5% of cases. The rate modified Thrombolysis in Cerebral Infarction 2b to 3 was 79.6% (74 patients [95% CI, 71.3–87.8]). Thirty-day mortality was 29% (27 patients [95% CI, 20–39.4]). Early neurological improvement was 19.5% (17 patients [95% CI, 11.8–29.5]), and symptomatic intracranial hemorrhage was 5.4% (5 patients [95% CI, 1.7–12.1]). Patients who died at 30 days exhibited significantly lower lymphocyte count, higher levels of aspartate, and LDH (lactate dehydrogenase). After adjustment for age, initial National Institutes of Health Stroke Scale, Alberta Stroke Program Early Computed Tomography score, and successful reperfusion, these biological markers remained associated with increased odds of 30-day mortality (adjusted odds ratio of 2.70 [95% CI, 1.21–5.98] per SD-log decrease in lymphocyte count, 2.66 [95% CI, 1.22–5.77] per SD-log increase in aspartate, and 4.30 [95% CI, 1.43–12.91] per SD-log increase in LDH). Conclusions: The 29% rate of 30-day mortality after MT among patients with COVID-19 is not negligible. Abnormalities of lymphocyte count, LDH and aspartate may depict a patient’s profiles with poorer outcomes after MT. REGISTRATION: URL: https://www.clinicaltrials.gov ; Unique identifier: NCT04406090.

Combined quantum mechanics and molecular mechanics (QM/MM) calculations were carried out to characterize the reaction mechanism of the NS3 protease with its preferred substrate (NS5A/5B). The main purpose of this study was to locate the barrier states and intermediates along the distinguished coordinate path (DCP) involved in this process. These structures, and in particular the one corresponding to the first barrier state and intermediate (B1 and I1), could be a starting point for the synthesis of inhibitors of this protease, which could be used to treat hepatitis C. The two first steps of the reaction mechanism were studied, i.e., the acylation step and the breaking of the peptide bond. The first step takes place through a tetracoordinated intermediate, as suggested from previous works on other Serine proteases. The importance of the different amino acid residues was also considered (perturbation study where the MM charges of each residue were set to zero independently). The residues of the oxyanion hole were confirmed as the most important for the electrostatic stabilization of the tetracoordinate intermediate. Moreover, the role of other residues, e.g., Arg-155 and Asp-79, was also explained.

The reaction mechanism of the NS3/NS4A protease with the NS4B/5A and NS4A/4B natural substrates has been investigated using the QM/MM (quantum mechanics/molecular mechanics) approach, and some calculations have been performed on the reaction with the NS5A/5B natural substrate. This study widely extends a previous contribution of our group on the reaction mechanism with the NS5A/5B substrate, the main goal here being to understand the differences found between the reaction mechanism of each natural substrate and the role played by the enzymatic residues in the catalytic cycle. This knowledge will ultimately help in developing new NS3/NS4A protease inhibitors. The two first steps of the mechanism have been considered: Acylation and breaking of the peptide bond, with emphasis on the former one (rate limiting process). Energy and free energy profiles for both steps have been calculated at the AM1/MM level and corrected by means of MP2 ab initio calculations, being evident the importance of correlation energy. Acylation is the rate limiting step in all cases and occurs through a tetracoordinated intermediate, as previously suggested for other serine proteases. Specificities in the NS4B/5A mechanism can be attributed to the presence of a Proline residue in the substrate P2 position. The analysis of structures and energies confirm the importance of the oxyanion hole in the electrostatic stabilization of the tetracoordinated intermediate. Finally, the role of other residues, e.g., Arg-155 and Asp-79, has been explained, and the viability of Arg-155 mutants and its resistance to some protease inhibitors has been understood thanks to virtual mutation studies.