Neurological manifestations of COVID-19 in adults and children

Different neurological manifestations of coronavirus disease 2019 (COVID-19) in adults and children and their impact have not been well characterized. We aimed to determine the prevalence of neurological manifestations and in-hospital complications among hospitalized COVID-19 patients and ascertain differences between adults and children. We conducted a prospective multicentre observational study using the International Severe Acute Respiratory and emerging Infection Consortium (ISARIC) cohort across 1507 sites worldwide from 30 January 2020 to 25 May 2021. Analyses of neurological manifestations and neurological complications considered unadjusted prevalence estimates for predefined patient subgroups, and adjusted estimates as a function of patient age and time of hospitalization using generalized linear models. Overall, 161 239 patients (158 267 adults; 2972 children) hospitalized with COVID-19 and assessed for neurological manifestations and complications were included. In adults and children, the most frequent neurological manifestations at admission were fatigue (adults: 37.4%; children: 20.4%), altered consciousness (20.9%; 6.8%), myalgia (16.9%; 7.6%), dysgeusia (7.4%; 1.9%), anosmia (6.0%; 2.2%) and seizure (1.1%; 5.2%). In adults, the most frequent in-hospital neurological complications were stroke (1.5%), seizure (1%) and CNS infection (0.2%). Each occurred more frequently in intensive care unit (ICU) than in non-ICU patients. In children, seizure was the only neurological complication to occur more frequently in ICU versus non-ICU (7.1% versus 2.3%, P < 0.001). Stroke prevalence increased with increasing age, while CNS infection and seizure steadily decreased with age. There was a dramatic decrease in stroke over time during the pandemic. Hypertension, chronic neurological disease and the use of extracorporeal membrane oxygenation were associated with increased risk of stroke. Altered consciousness was associated with CNS infection, seizure and stroke. All in-hospital neurological complications were associated with increased odds of death. The likelihood of death rose with increasing age, especially after 25 years of age. In conclusion, adults and children have different neurological manifestations and in-hospital complications associated with COVID-19. Stroke risk increased with increasing age, while CNS infection and seizure risk decreased with age

Inhibition of p42 and p44 MAP kinase does not alter smooth muscle contraction in swine carotid artery

Caldesmon inhibits myosin ATPase activity; phosphorylation of caldesmon reverses the inhibition. The caldesmon kinase is believed to be mitogen-activated protein (MAP) kinase. MAP kinases are activated during vascular stimulation, but a cause-and-effect relationship between kinase activity and contraction has not been established. We examined the role of MAP kinase in contraction using PD-098059, an inhibitor of MAP kinase kinase (MEK). MAP kinase activity was assessed using an anti-active MAP kinase antibody and direct measurement of MAP kinase catalyzed phosphorylation of myelin basic protein, MBP-(95—98). MAP kinase phosphorylation, stimulated by histamine (50 μM) or phorbol 12,13-dibutyrate (PDBu, 0.1 μM), was inhibited by PD-098059 (100 μM). PD-098059 did not alter the sensitivity or the maximal level of force in smooth muscle stimulated by histamine or PDBu, nor did PD-098059 affect contraction of β-escin-permeabilized tissue. Our data suggest that p44 and p42 MAP kinases are not involved in regulation of vascular smooth muscle contraction. These results do not, however, preclude a role for other isoforms of the MAP kinase family.</jats:p

Abstract 780: Atorvastatin Activates A Novel Nbs1-dependent Mechanism Of Accelerating Dna Repair In Atherosclerosis

There is increasing evidence that reactive oxidant species (ROS) and DNA damage promote the development and complications of atherosclerosis. Although statin therapy reduces both ROS and DNA damage in atherosclerosis, the mechanism of this effect is unknown. We first examined expression of DNA damage and repair markers in vascular smooth muscle cells (VSMCs) of human atherosclerotic plaques. With increasing disease severity, there was increased VSMC expression of the DNA repair markers P-ATM/ATR substrate and P-H2AX from 2.7%±2.2 and 0.5±0.71 [mean±SEM] (AHA Grade I/II), to 21%±3.5 and 36.5±2.1 (Grade III) lesions, and 86.5%±0.7 and 69.3±7.6 (Grade IV/V). Cultured plaque VSMCs also showed a 1.5 fold increased oxidant stress; a 4.4 fold increased double-stranded DNA breaks, and expression of P-H2AX by Western blots. ROS analogues induced a robust DNA damage response in VSMCs, characterised by lengthening of tails on COMET assay, and activation of ATM and P-H2AX, with completion of repair by 6 hours. Atorvastatin pre-treatment accelerated DNA repair by approximately 2 hours without inhibiting ROS induction or DNA damage, and markedly accelerated the kinetics of nibrin (NBS-1) and P-H2AX activation, both proteins recruited to sites of DNA damage, by preventing degradation of NBS-1. Atorvastatin induced phosphorylation of HDM2, an E3 ligase and putative regulator of NBS-1 stability, and siRNA knockdown of HDM2 replicated the effect of atorvastatin on NBS-1. The ability of atorvastatin to accelerate repair was completely dependent upon NBS-1, as atorvastatin was ineffective in cells either null or expressing constitutively active NBS-I. In summary, we have demonstrated a novel NBS-1-dependent mechanism by which statins accelerate DNA repair in atherosclerosis, through HDM2 phosphorylation and stabilisation of NBS-1. We believe that both NBS-1 and HDM2 are critical to DNA repair in atherosclerosis.</jats:p

LPP, a LIM protein highly expressed in smooth muscle

An 80-kDa protein, prominently expressed in smooth muscle, was microsequenced and identified as LPP, the product of the lipoma-preferred partner gene (Petit MMR, Mols R, Schoenmakers EFPM, Mandahl N, and Van de Ven WJM. Genomics 36: 118–129, 1996). Using a specific anti-LPP antibody, we showed, in Western blots and with immunofluorescence microscopy, the selective expression of LPP in vascular and visceral smooth muscles (∼0.5–1 ng/μg total protein). In other mature (noncultured) tissues, including heart and skeletal muscle, the protein is present only in trace amounts and is closely correlated with the levels of the smooth muscle marker α-actin. In freshly isolated guinea pig bladder smooth muscle cells, immunofluorescence images showed LPP as linear arrays of punctate, longitudinally oriented staining superimposed with vinculin staining on the plasma membrane surface. A corresponding pattern of periodic labeling at the membrane in transverse sections of bladder smooth muscle suggested an association of LPP with peripheral dense bodies. In cultured rat aortic smooth muscle cells, LPP colocalized with vinculin at focal adhesions but not with p120 catenin or α-actinin. Overexpression of the protein increased EGF-stimulated migration of vascular smooth muscle cells in Transwell assays, suggesting the participation of LPP in cell motility. The Rho-kinase inhibitor Y-27632 dissociated focal adhesions and LPP staining at the cell periphery and enhanced the nuclear accumulation of LPP induced by leptomycin B, indicating that LPP has a potential for relocating to the nucleus through a shuttling mechanism that is sensitive to inhibition of Rho-kinase.</jats:p

Regulation by GDI of RhoA/Rho-kinase-induced Ca²⁺sensitization of smooth muscle myosin II

We characterized the role of guanine nucleotide dissociation inhibitor (GDI) in RhoA/Rho-kinase-mediated Ca2+sensitization of smooth muscle. Endogenous contents (∼2–4 μM) of RhoA and RhoGDI were near stoichiometric, whereas a supraphysiological GDI concentration was required to relax Ca2+sensitization of force by GTP and guanosine 5′- O-(3-thiotriphosphate) (GTPγS). GDI also inhibited Ca2+sensitization by GTP · G14V RhoA, by α-adrenergic and muscarinic agonists, and extracted RhoA from membranes. GTPγS translocated Rho-kinase to a Triton X-114-extractable membrane fraction. GTP · G14V RhoA complexed with GDI also induced Ca2+sensitization, probably through in vivo dissociation of GTP · RhoA from the complex, because it was reversed by addition of excess GDI. GDI did not inhibit Ca2+sensitization by phorbol ester. Constitutively active Cdc42 and Rac1 inhibited Ca2+sensitization by GTP · G14V RhoA. We conclude that 1) the most likely in vivo function of GDI is to prevent perpetual “recycling” of GDP · RhoA to GTP · RhoA; 2) nucleotide exchange (GTP for GDP) on complexed GDP · RhoA/GDI can precede translocation of RhoA to the membrane; 3) activation of Rho-kinase exposes a hydrophobic domain; and 4) Cdc42 and Rac1 can inhibit Ca2+sensitization by activated GTP · RhoA.</jats:p

Phosphorylation of telokin by cyclic nucleotide kinases and the identification of in vivo phosphorylation sites in smooth muscle

AbstractThe Ca2+-independent acceleration of dephosphorylation of the regulatory light chain of smooth muscle myosin and relaxation of smooth muscle by telokin are enhanced by cyclic nucleotide-activated protein kinase(s) [Wu et al. (1998) J. Biol. Chem. 273, 11362–11369]. The purpose of this study was to determine the in vivo site(s) and in vitro rates of telokin phosphorylation and to evaluate the possible effects of sequential phosphorylation by different kinases. The in vivo site(s) of phosphorylation of telokin were determined in rabbit smooth muscles of longitudinal ileum and portal vein. Following stimulation of ileum with forskolin (20 μM) the serine at position 13 was the only amino acid to exhibit increased phosphorylation. Rabbit portal vein telokin was phosphorylated on both Ser-13 and -19 as a result of forskolin and GTPγS stimulation in vivo. Point mutation of Ser-13 (to Ala or Asp) abolished in vitro phosphorylation by cyclic nucleotide-dependent protein kinases