Unusual arterial thrombotic events in Covid-19 patients.

INTRODUCTION: COVID-19 infection is commonly complicated with pro-thrombotic state and endothelial dysfunction. While several studies reported a high incidence of venous thromboembolic events. The occurrence of arterial thromboses are yet rarely described and could be underestimated. OBJECTIVES: To describe the clinical and biological characteristics of COVID-19 patients presenting with an associated arterial thromboembolic event. MATERIAL AND METHODS: We performed a retrospective multicentric study in 3 centers between France and Italy. All patients with a confirmed SARS-CoV-2 infection and arterial thromboembolic events were included in the analysis. RESULTS: From March 8th to April 25th 2020, we identified 20 patients (24 events) with arterial thromboembolic events over 209 admitted patients (9.6%) with severe COVID-19 infection. Arterial thrombotic events included acute coronary occlusions (n = 9), stroke (n = 6), limb ischemia (n = 3), splenic infarcts (n = 3), aortic thrombosis (n = 2) and occlusive mesenteric ischemia (n = 1). At the time of the event, 10/20 (50%) of patients received thromboprohylaxis, 2/20 (10%) were receiving treatment dose anticoagulation and 5/20 (25%) were receiving antiplatelet therapy. CONCLUSION: Our observations suggest that serious arterial thrombotic events might occur in Covid-19 patients. However, the exact incidence of such events and the best way to prevent them yet remains to be investigated.

Leukoencephalopathy with calcifications and cysts (LCC): 5 cases and literature review.

INTRODUCTION: Leukoencephalopathy with calcifications and cysts (LCC) is a rare autosomal recessive cerebral angiomatous-like microangiopathy characterized by diffuse and asymmetric white-matter lesions associated with multiple calcifications and cysts. The disease is caused by SNORD118 mutations. The entire clinical spectrum of LCC is not yet fully determined. MATERIAL AND METHODS: To define the clinical spectrum of LCC, we analyzed data from recently diagnosed cases and from the litterature. Both clinical and imaging features from our five LCC cases harboring compound heterozygous SNORD118 mutations were presented and all cases reported in the litterature reviewed. RESULTS: Ninety-two LCC cases including our five patients were identified. Consanguinity was rare (4%), and 97% of cases were symptomatic. Mean age of first clinical manifestations was 16.1±16.1 years (range 1 month-71 years) and was earlier in men (10.3±14.3 years) than in women (20.2±22.8 years) (P=0.02). The main inaugural symptoms were seizures (36%; mean age at onset: 5.2±9.5 years) and progressive neurological symptoms including ataxia, dystonia and spasticity (26%; 27.8±23.6 years). Intracranial hypertension was less frequently observed (14%), mostly in adults (mean age 31.5±13.2 years). Ischemic or hemorrhagic strokes were inaugural symptoms in two adults (2%). During follow-up, most patients developed progressive extrapyramidal, cerebellar and pyramidal signs (83%), cognitive decline (56%), seizures (37%), intracranial hypertension (30%) or stroke (2%). CONCLUSION: In LCC, the clinical spectrum is largely heterogeneous and the course of the disease appears highly variable in contrast to other hereditary cerebral small vessel diseases.

Endovascular management of extracranial occlusions at the hyperacute phase of stroke with tandem occlusions.

BACKGROUND AND PURPOSE: The management of cervical artery occlusions in hyperacute stroke with tandem cervical/intracranial occlusions has not yet become standardized, especially when the circle of Willis is effective. METHODS: We retrospectively analyzed the safety and accuracy of current approaches to manage the cervical occlusion in consecutive patients with tandem occlusions addressed for intracranial mechanical thrombectomy (MT) in our department from January 2012 to May 2017. The different approaches that could be performed in a same patient during the same procedure or hospitalization were analyzed separately. RESULTS: We reported 64 approaches to manage the cervical occlusion in 49 patients with tandem occlusion (14% of MT): medical treatment alone in 16/64 (25%), stenting/angioplasty in 16/64 (25%), occlusion with coils in 12/64 (19%), angioplasty alone in 9/64 (14%), stent-retriever in 8/64 (12%), and/or thromboaspiration in 3/64 (5%). Early ipsilateral embolic recurrence occurred after 9/64 (14%) of them. It was strongly associated with the presence of a cervical intraluminal thrombus (P=0.001) and was then lower after occlusion with coils and stent-retriever compared to medical treatment alone and thromboaspiration (P=0.002). Occlusion with coils had a lower rate of radiological intracranial hemorrhage at 48-hour compared to other approaches (P=0.009). The 3-month rates of favorable outcome (P=0.806) and mortality (P=0.878) were similar. One delayed stroke was imputable to an occlusion with coils, for a median (Q1-Q3) follow-up of 10 (3-20) months. CONCLUSIONS: Cervical occlusion with coils and thrombectomy with stent-retrievers may be relevant to prevent early embolic recurrence in cervical occlusions with intraluminal thrombus. Stent-retrievers should be further assessed as a first-line approach, since delayed stroke may occur following occlusion with coils. Medical treatment alone may be sufficient when no cervical intraluminal thrombus is present, the Willis polygon is effective, and the cervical occlusion can be crossed easily to perform the intracranial thrombectomy.

Competition of Thermomyces lanuginosus lipase with its hydrolysis products at the oil-water interface.

Lipase-catalyzed hydrolysis of triglycerides yields glycerol and free fatty-acids, provided that the enzyme is non-regioselective. For an Sn-1,3 regioselective enzyme, such as lipase from Thermomyces lanuginosus, the final product is no longer glycerol but Sn-2 monoglyceride instead. However, surface active molecules generated by lipolysis may have a detrimental effect on the interfacial biocatalysis since it is known that low molecular weight surfactants can displace proteins from interfaces. By using drop profile analysis tensiometry, we evaluated the interfacial properties of the lipase-generated molecules and their competitive effect on the adsorption behavior of the lipase and on the proceeding lipolysis. Our results show that even at concentration ratios of 8.64×10-4M (Sn-2 monoglyceride) to 2.5×10-7M (lipase), the final interfacial pressure values are very similar as for the system containing the lipase alone (i.e. ∼26 mN/m). This is a strong indication that monoglycerides, as the most interfacially active products generated during regioselective lipolysis, are expelled from the oil-water interface by the lipase. We attribute this effect to intermolecular lipase-lipase interactions, resulting in a low desorption probability of the lipase. For low oleic acid concentrations, the interfacial tension is solely determined by the lipase, while for higher concentrations, lipase and oleic acid both contribute to the tension values. We propose a hypothesis based on the preferential interaction of oleic acid molecules with hydrophobic sites on the lipase. The pH dependence of the adsorption rate and the interfacial activity of the lipase were also investigated.

Ellipsometric study of molecular orientations of Thermomyces lanuginosus lipase at the air-water interface by simultaneous determination of refractive index and thickness.

Ellipsometric studies of very thin organic films suffer from the low refractive index contrast between layer and bulk substrate. We demonstrate that null ellipsometry can not only provide detailed information about the adsorption kinetics and surface excess values, but in addition on layer thicknesses with submonolayer resolution of a lipase from Thermomyces lanuginosus at the air-water interface. While measuring very close to the Brewster angle, refractive indices and layer-thicknesses can both be determined with a precision that is sufficiently high to make conclusions on the density and orientation of the molecules at the interface. The orientation was found to be concentration- and pH value-dependent. At the isoelectric point, the lipase was almost vertically oriented with respect to the surface, while for pure distilled water and low lipase concentration a rather horizontal alignment was found. Further experiments, varying the size of the interfacial area in a Langmuir trough, confirm the different layer structures.

[Chorea, lupus and antiphospholipid antibodies]. / Chorée, lupus et antiphospholipides.

Chorea may occur in patients with SLE with a frequency estimated at 1 to 3% in adults and up to 9% in paediatric lupus. Chorea is frequently a presenting feature, and is strongly related to the presence of antiphospholipid antibodies. A treatment with antiplatelet agents and hydroxychloroquine is generally sufficient. During follow-up, the patients with chorea have a significant higher risk to develop thrombotic events (mainly arterial). They also have an excess risk of obstetric morbidity and valvular disease. The prescription of antiplatelet agents and adequate management, especially during pregnancy, can probably reduce this risk.

[An unrecognized cause of myelopathy associated with copper deficiency: the use of denture cream]. / Une cause méconnue de myélopathie par carence en cuivre: l'utilisation de pâte adhésive dentaire.

We report two patients with myelopathy associated with copper deficiency and pancytopenia. Excessive intake of zinc can lead to a severe deficiency of copper reducing the absorption of ingested copper. The patients had in common consumption of denture adhesive paste containing zinc. In both patients, laboratory tests showed a combination of copper deficiency, hyperzincemia and increased urinary zinc level. The use of a denture cream was stopped. Copper supplementation, initially subcutaneously then oral corrected the copper deficiency and pancytopenia. Clinically, the pain faded but the gait disturbance persisted. Copper deficiency associated with the use of denture cream rich in zinc is an unrecognized cause of myelopathy associated with pancytopenia which should be diagnosed early to establish appropriate therapeutic measures to minimize neurological complications.

General practitioners' needs for ongoing support for the interpretation of spirometry tests.

BACKGROUND: Although one out of three general practitioners (GPs) carries out spirometry, the diagnostic interpretation of spirometric test results appears to be a common barrier for GPs towards its routine application. METHODS: Multivariate cross-sectional analysis of a questionnaire survey among 137 GPs who participated in a spirometry evaluation programme in the Netherlands. We identified characteristics of GPs and their practice settings associated with GPs' need for ongoing support for spirometry interpretation. RESULTS: Response rate on the survey questionnaire was 98%. The need for ongoing support among the participating GPs was 69%. GPs' recent spirometry training showed a statistically significant association with the need for ongoing support for the interpretation of spirometry (odds ratio 0.43, 95% CI 0.20-0.92). CONCLUSION: There is a need for ongoing support for spirometry interpretation among GPs. Recent spirometry training partially diminished this need.

beta-Amyloid efflux mediated by p-glycoprotein.

A large body of evidence suggests that an increase in the brain beta-amyloid (Abeta) burden contributes to the etiology of Alzheimer's disease (AD). Much is now known about the intracellular processes regulating the production of Abeta, however, less is known regarding its secretion from cells. We now report that p-glycoprotein (p-gp), an ATP-binding cassette (ABC) transporter, is an Abeta efflux pump. Pharmacological blockade of p-gp rapidly decrease extracellular levels of Abeta secretion. In vitro binding studies showed that addition of synthetic human Abeta1-40 and Abeta1-42 peptides to hamster mdr1-enriched vesicles labeled with the fluorophore MIANS resulted in saturable quenching, suggesting that both peptides interact directly with the transporter. Finally, we were able to directly measure transport of Abeta peptides across the plasma membranes of p-gp enriched vesicles, and showed that this phenomenon was both ATP- and p-gp-dependent. Taken together, our study suggests a novel mechanism of Abeta detachment from cellular membranes, and represents an obvious route towards identification of such a mechanism in the brain.

Localization of the cGMP-dependent protein kinases in relation to nitric oxide synthase in the brain.

The distributions of the type I and type II isoforms of cGMP-dependent protein kinase were determined in the rat brain using immunohistochemistry and in situ hybridization, and compared with the localization of NO synthase determined with NADPH-diaphorase histochemistry. The type I cGMP-dependent protein kinase was highly expressed in the Purkinje cells of the cerebellar cortex, where it was closely associated with the NO synthase containing granule and basket cells. This kinase was also found in neurons in the dorsomedial nucleus of the hypothalamus, where it may be regulated by NO or atriopeptides. The type I kinase was not detected in other central neurons. In contrast, the type II kinase was widely distributed in the brain. In particular, it was highly expressed in the olfactory bulb, cortex, septum, thalamus, tectum and various brainstem nuclei. Many regions expressing this kinase also contained, or received innervation from NO synthase positive neurons. These results indicate that type I cGMP-dependent protein kinase may act as a downstream effector for NO only in the cerebellar cortex and the dorsomedial hypothalamus. The type II cGMP-dependent protein kinase appears to be a major mediator of NO actions in the brain.

Noradrenaline excites non-cholinergic laterodorsal tegmental neurons via two distinct mechanisms.

Cholinergic neurons of the laterodorsal tegmental nucleus have been hypothesized to play a critical role in the-generation and maintenance of rapid eye movement sleep. Less is known about the function of non-cholinergic laterodorsal tegmental nucleus neurons. As part of our ongoing studies of the brainstem circuitry controlling behavioral state, we have begun to investigate the functional properties of these neurons. In the course of these experiments, we have observed a novel response to the neurotransmitter noradrenaline. Whole-cell patch-clamp recordings of laterodorsal tegmental nucleus neurons were carried out in 21- to 35-day-old rat brain slices. A subpopulation of laterodorsal tegmental nucleus cells responded to a 30-s application of 50 microM noradrenaline with depolarization and a decrease in input resistance which lasted several minutes. Following return to resting membrane potential, these cells invariably exhibited barrages of excitatory postsynaptic potentials which lasted at least 12 min. These excitatory postsynaptic potentials were reversibly abolished by bath application of tetrodotoxin, as well as by the non-N-methyl-D-aspartate receptor antagonist 6-cyano-7-nitroquinoxaline-2,3-dione, but were insensitive to application of the N-methyl-D-aspartate receptor antagonist 2-amino-5-phosphonopentanoic acid. To examine whether these neurons were cholinergic, the recorded cells were labeled with biocytin and tested for co-localization with reduced nicotinamide adenine dinucleotide phosphate-diaphorase, a marker for laterodorsal tegmental nucleus cholinergic neurons. In every instance, neurons with these properties were non-cholinergic. However, they were always located in close proximity to reduced nicotinamide adenine dinucleotide phosphate-diaphorase-positive laterodorsal tegmental nucleus cells. The present data indicate that noradrenaline, in addition to directly inhibiting cholinergic cells of the laterodorsal tegmental nucleus, also results in the direct and indirect excitation of non-cholinergic cells of the laterodorsal tegmental nucleus. The indirect excitation is long lasting and mediated by glutamatergic mechanisms. Our working hypothesis is that these non-cholinergic cells are local circuit inhibitory interneurons and that prolonged excitation of these neurons by noradrenaline may serve as a mechanism for inhibition of cholinergic laterodorsal tegmental nucleus cells during wakefulness, when noradrenaline tone is high.

Vasoactive intestinal polypeptide excites medial pontine reticular formation neurons in the brainstem rapid eye movement sleep-induction zone.

Although it has long been known that microinjection of the cholinergic agonist carbachol into the medial pontine reticular formation (mPRF) induces a state that resembles rapid eye movement (REM) sleep, it is likely that other transmitters contribute to mPRF regulation of behavioral states. A key candidate is the peptide vasoactive intestinal polypeptide (VIP), which innervates the mPRF and induces REM sleep when injected into this region of the brainstem. To begin understanding the cellular mechanisms underlying this phenomenon, we examined the effects of VIP on mPRF cells using whole-cell patch-clamp recordings in the in vitro rat brainstem slice. VIP directly depolarized cells via activation of an inward current; these effects were attenuated and potentiated in low-sodium and low-calcium medium, respectively. The depolarization induced by VIP was slower in onset and longer-lived than that evoked by carbachol. The VIP-induced depolarization was reduced in a dose-dependent manner by a competitive antagonist of VIP receptors. Effects of VIP were attenuated in the presence of guanosine 5'-O-(2-thiodiphosphate, 2'5'dideoxyadenosine, and PKI15-24 and were nonadditive in the presence of 8-bromo-cAMP. We conclude that VIP excites mPRF neurons by activation of a sodium current. This effect is mediated at least in part by G-protein stimulation of adenylyl cyclase, cAMP, and protein kinase A. These data suggest that VIP may play a physiological role in REM induction by its actions on mPRF neurons.

Dynamic properties of corticothalamic excitatory postsynaptic potentials and thalamic reticular inhibitory postsynaptic potentials in thalamocortical neurons of the guinea-pig dorsal lateral geniculate nucleus.

The properties of postsynaptic potentials evoked by stimulation of cortical, retinal and GABAergic thalamic afferents were examined in vitro in thalamocortical neurons of the guinea-pig dorsal lateral geniculate nucleus. Brief trains of stimulation (2-10 stimuli) delivered to corticothalamic fibers led to a frequency-dependent increase in excitatory postsynaptic potential amplitude associated with an increase in activation of both N-methyl-D-aspartate and non-N-methyl-D-aspartate glutamate receptors. In addition, repetitive stimulation of corticothalamic fibers also gave rise to a slow excitatory postsynaptic potential that was blocked by local application of the glutamate metabotropic receptor antagonist alpha-methyl-4-carboxyphenylglycine. In contrast, repetitive stimulation of optic tract fibers resulted in monosynaptic excitatory postsynaptic potentials that did not potentiate and were not followed by the generation of a slow excitatory postsynaptic potential. Repetitive activation of the optic radiation also evoked both GABA(A) and GABA(B) receptor-mediated inhibitory postsynaptic potentials. These inhibitory postsynaptic potentials exhibited frequency-dependent depression during repetitive activation. The presence of frequency-dependent facilitation of corticothalamic excitatory postsynaptic potentials and frequency-dependent decrement of inhibitory postsynaptic potentials, as well as the ability of corticothalamic fibers to activate glutamate metabotropic receptors, suggests that sustained activation of corticothalamic afferents in vivo may result in postsynaptic responses in thalamocortical cells that are initially dominated by GABAergic inhibitory postsynaptic potentials followed by prominent monosynaptic excitatory postsynaptic potentials as well as a slow depolarization of the membrane potential.Therefore, the corticothalamic system may inhibit or enhance the excitability and responsiveness of thalamocortical neurons, based both on the spatial and temporal features of thalamocortical interactions.

Novel effects of FCCP [carbonyl cyanide p-(trifluoromethoxy)phenylhydrazone] on amyloid precursor protein processing.

Amyloidogenic processing of the beta-amyloid precursor protein (APP) has been implicated in the pathology of Alzheimer's disease. Because it has been suggested that catabolic processing of the APP holoprotein occurs in acidic intracellular compartments, we studied the effects of the protonophore carbonyl cyanide p-(trifluoromethoxy)phenylhydrazone (FCCP) and the H+-ATPase inhibitor bafilomycin A1 on APP catabolism in human embryonic kidney 293 cells expressing either wild-type or "Swedish" mutant APP. Unlike bafilomycin A1, which inhibits beta-amyloid production in cells expressing mutant but not wild-type APP, FCCP inhibited beta-amyloid production in both cell types. Moreover, the effects of FCCP were independent of alterations in total cellular APP levels or APP maturation, and the concentrations used did not alter either cellular ATP levels or cell viability. Bafilomycin A1, which had no effect on beta-amyloid production in wild-type cells, inhibited endocytosis of fluorescent transferrin, whereas concentrations of FCCP that inhibited beta-amyloid production in these cells had no effect on endosomal function. Thus, in wild-type-expressing cells it appears that the beta-amyloid peptide is not produced in the classically defined endosome. Although bafilomycin A1 decreased beta-amyloid release from cells expressing mutant APP but not wild-type APP, it altered lysosomal function in both cell types, suggesting that in normal cells beta-amyloid is not produced in the lysosome. Although inhibition of beta-amyloid production by bafilomycin A1 in mutant cells may occur via changes in endosomal/lysosomal pH, our data suggest that FCCP inhibits wild-type beta-amyloid production by acting on a bafilomycin A1-insensitive acidic compartment that is distinct from either the endosome or the lysosome.

Regulation of amyloid precursor protein cleavage.

Multiple lines of evidence suggest that increased production and/or deposition of the beta-amyloid peptide, derived from the amyloid precursor protein, contributes to Alzheimer's disease. A growing list of neurotransmitters, growth factors, cytokines, and hormones have been shown to regulate amyloid precursor protein processing. Although traditionally thought to be mediated by activation of protein kinase C, recent data have implicated other signaling mechanisms in the regulation of this process. Moreover, novel mechanisms of regulation involving cholesterol-, apolipoprotein E-, and stress-activated pathways have been identified. As the phenotypic changes associated with Alzheimer's disease encompass many of these signaling systems, it is relevant to determine how altered cell signaling may be contributing to increasing brain amyloid burden. We review the myriad ways in which first messengers regulate amyloid precursor protein catabolism as well as the signal transduction cascades that give rise to these effects.

Mitogen-activated protein kinase is involved in N-methyl-D-aspartate receptor regulation of amyloid precursor protein cleavage.

Glutamate is the principal excitatory neurotransmitter in the mammalian brain. Several lines of evidence suggest that glutamatergic hypoactivity exists in the Alzheimer's disease brain, where it may contribute to both brain amyloid burden and cognitive dysfunction. Although metabotropic glutamate receptors have been shown to alter cleavage of the amyloid precursor protein, little attention has been paid to the role of N-methyl-D-aspartate receptors in this process. We now report that activation of N-methyl-D-aspartate receptors in transiently transfected human embryonic kidney 293 cells increases production of the soluble amyloid precursor protein derivative. Moreover, using both pharmacological and gene transfer techniques, we show that this effect is largely due to activation of the mitogen-activated protein kinase cascade, specifically the pathway leading to activation of extracellular signal-regulated protein kinase but not other mitogen-activated protein kinases. These observations further our understanding of the pathways that regulate amyloid precursor protein cleavage, and buttress the notion that regulation of amyloid precursor protein cleavage is critically dependent upon the mitogen-activated protein kinase cascade.

Nitric oxide regulates cyclic GMP-dependent protein kinase phosphorylation in rat brain.

Nitric oxide (NO) acts via soluble guanylyl cyclase to increase cyclic GMP (cGMP), which can regulate various targets including protein kinases. Western blotting showed that type II cGMP-dependent protein kinase (cGK II) is widely expressed in various brain regions, especially in the thalamus. In thalamic extracts, the phosphorylation of several proteins, including cGK II, was increased by exogenous NO or cGMP. In vivo pretreatment with a NO synthase inhibitor reduced the phosphorylation of cGK II, and this could be reversed by exogenous NO or cGMP. Conversely, brainstem electrical stimulation, which enhances thalamic NO release, caused a NO synthase-dependent increase in the phosphorylation of thalamic cGK II. These results indicate that endogenous NO regulates cGMP-dependent protein phosphorylation in the thalamus. The activation of cGKII by NO may play a role in thalamic mechanisms underlying arousal.

Monitoring neuronal NO release in vivo in cerebellum, thalamus and hippocampus.

A variety of methods has been developed based on in vivo microdialysis which allow one to examine the NO/cGMP signal transduction system in action in behaving animals. The extracellular levels of cGMP, the NO oxidative products nitrate and nitrite, and NO itself can all be determined. Using these methods changes in NO and cGMP production in response to pharmacological manipulations can be examined in vivo. In addition, it has been discovered that the activity of this system varies with the behavioral state of the animal. NO and cGMP appear to act via distinct downstream effectors in different brain regions. This opens up the possibility of selectively manipulating NO and cGMP signaling in discrete neuronal populations.

Nitric oxide production in rat thalamus changes with behavioral state, local depolarization, and brainstem stimulation.

Since its discovery as a putative neurotransmitter in the CNS, several functional roles have been suggested for nitric oxide (NO). However, few studies have investigated the role of NO in natural physiology. Because NO synthase (NOS) has been localized in regions believed to be important for attention and arousal, we hypothesized that NO production would be state-dependent. To test this hypothesis, we used in vivo microdialysis, coupled with the hemoglobin-trapping technique, to monitor extracellular NO concentrations in rat thalamus during wake, slow-wave sleep (SWS), and rapid eye movement (REM) sleep. The thalamus is known to receive a massive innervation from the NOS/cholinergic neurons in the mesopontine brainstem, which have been suggested to play a key role in EEG desynchronized states. To test whether thalamic NO output was sensitive to neuronal-dependent changes in the mesopontine brainstem, we measured thalamic NO concentration in response to electrical stimulation in the laterodorsal tegmentum (LDT) of anesthetized rats. Finally, the calcium dependence of NO release was tested by local depolarization with a high potassium dialysate or by addition of a calcium chelator. The results showed that (1) extracellular NO concentrations in the thalamus were high during wake and REM sleep and significantly lower during SWS, (2) thalamic NO release increased in response to LDT stimulation in both a site-specific and tetrodotoxin (TTX)-dependent manner, and (3) NO production was calcium-dependent. These data suggest that thalamic NO production may play a role in arousal.