Preferential delivery of lipid-ligand conjugated DNA/RNA heteroduplex oligonucleotide to ischemic brain in hyperacute stage.

Antisense oligonucleotide (ASO) is a major tool used for silencing pathogenic genes. For stroke in the hyperacute stage, however, the ability of ASO to regulate genes is limited by its poor delivery to the ischemic brain owing to sudden occlusion of the supplying artery. Here we show that, in a mouse model of permanent ischemic stroke, lipid-ligand conjugated DNA/RNA heteroduplex oligonucleotide (lipid-HDO) was unexpectedly delivered 9.6 times more efficiently to the ischemic area of the brain than to the contralateral non-ischemic brain and achieved robust gene knockdown and change of stroke phenotype, despite a 90% decrease in cerebral blood flow in the 3 h after occlusion. This delivery to neurons was mediated via receptor-mediated transcytosis by lipoprotein receptors in brain endothelial cells, the expression of which was significantly upregulated after ischemia. This study provides proof-of-concept that lipid-HDO is a promising gene-silencing technology for stroke treatment in the hyperacute stage.

The Role of Long Noncoding RNA MALAT1 in Diabetic Polyneuropathy and the Impact of Its Silencing in the Dorsal Root Ganglion by a DNA/RNA Heteroduplex Oligonucleotide.

Diabetic polyneuropathy (DPN) is the most common complication of diabetes, yet its pathophysiology has not been established. Accumulating evidence suggests that long noncoding RNA metastasis-associated lung adenocarcinoma transcript 1 (MALAT1) plays pivotal roles in the regulation of cell growth and survival during diabetic complications. This study aimed to investigate the impact of MALAT1 silencing in dorsal root ganglion (DRG) sensory neurons, using an α-tocopherol-conjugated DNA/RNA heteroduplex oligonucleotide (Toc-HDO), on the peripheral nervous system of diabetic mice. We identified MALAT1 upregulation in the DRG of chronic diabetic mice that suggested either a pathological change or one that might be protective, and systemic intravenous injection of Toc-HDO effectively inhibited its gene expression. However, we unexpectedly noted that this intervention paradoxically exacerbated disease with increased thermal and mechanical nociceptive thresholds, indicating further sensory loss, greater sciatic-tibial nerve conduction slowing, and additional declines of intraepidermal nerve fiber density in the hind paw footpads. Serine/arginine-rich splicing factors, which are involved in pre-mRNA splicing by interacting with MALAT1, reside in nuclear speckles in wild-type and diabetic DRG neurons; MALAT1 silencing was associated with their disruption. The findings provide evidence for an important role that MALAT1 plays in DPN, suggesting neuroprotection and regulation of pre-mRNA splicing in nuclear speckles. This is also the first example in which a systemically delivered nucleotide therapy had a direct impact on DRG diabetic neurons and their axons.

Effective silencing of miR-126 after ischemic stroke by means of intravenous α-tocopherol-conjugated heteroduplex oligonucleotide in mice.

Brain endothelial cells (BECs) are involved in the pathogenesis of ischemic stroke. Recently, several microRNAs (miRNAs) in BECs were reported to regulate the endothelial function in ischemic brain. Therefore, modulation of miRNAs in BECs by a therapeutic oligonucleotide to inhibit miRNA (antimiR) could be a useful strategy for treating ischemic stroke. However, few attempts have been made to achieve this strategy via systemic route due to lack of efficient delivery-method toward BECs. Here, we have developed a new technology for delivering an antimiR into BECs and silencing miRNAs in BECs, using a mouse ischemic stroke model. We designed a heteroduplex oligonucleotide, comprising an antimiR against miRNA-126 (miR-126) known as the endothelial-specific miRNA and its complementary RNA, conjugated to α-tocopherol as a delivery ligand (Toc-HDO targeting miR-126). Intravenous administration of Toc-HDO targeting miR-126 remarkably suppressed miR-126 expression in ischemic brain of the model mice. In addition, we showed that Toc-HDO targeting miR-126 was delivered into BECs more efficiently than the parent antimiR in ischemic brain, and that it was delivered more effectively in ischemic brain than non-ischemic brain of this model mice. Our study highlights the potential of this technology as a new clinical therapeutic option for ischemic stroke.

Restoration of clean water supply and toilet hygiene reduces infectious diseases in post-disaster evacuation shelters: A multicenter observational study.

After a massive disaster, many residents in affected areas are forced to temporarily stay in evacuation shelters. The exact impact of the state of resource supply and infrastructure in evacuation shelters on the health status of evacuees has not been sufficiently studied. Two weeks after the 2011 Great East Japan Earthquake (GEJE), comprehensive surveillance related to the health status and hygiene level was performed for all evacuation shelters (328 shelters with 46,480 evacuees at the peak) in one of the most devastating medical zones after the tsunami hit the area (Ishinomaki City). The joint relief team regularly visited all evacuation shelters across the area to assess the situation of resource supply levels, infrastructural damage, rapid need of resources, and the health status of the evacuees. In this cross-sectional observational study, we evaluated the relationship between the resource supply levels and health status among evacuees in two time periods (days 14-19 and 20-25). Among the evaluated vital resources, clean tap water supply was among the most disrupted by the disaster, and was not fully restored in most shelters during the assessment period. The cross-sectional relationship between resource supplies and morbidity was inconsistent between the two assessment periods, reflecting the multifactorial nature of health status in evacuation shelters. The clean tap water supply level at the first assessment showed a strong negative correlation with the subsequent prevalence of respiratory or gastrointestinal infectious conditions at the second assessment. Restorations in the clean tap water supply and toilet hygiene correlated each other, and both correlated with a decrease in the prevalence of gastrointestinal infectious conditions. In conclusion, disrupted clean tap water supply and inadequate toilet hygiene after a massive disaster would jointly harm the health status of those in shelters. Prompt assessments using quick visual assessment and restorations of these key resources have validity with suppressed environmental health risks among evacuees.

Convexity Subarachnoid Hemorrhage Soon after Starting a Direct Oral Anticoagulant in 2 Patients with Acute Infarction.

Convexity subarachnoid hemorrhage (cSAH) is typically due to head trauma, but it rarely occurs subsequent to acute ischemic stroke. Direct oral anticoagulants (DOACs) have favorable bleeding profiles as compared with warfarin, and, to our knowledge, no DOAC has been regarded as a causative agent for cSAH. Here, we reported 2 patients with cSAH apparently caused by starting DOAC therapy. No hemorrhage had been evident just prior to treatment initiation, but cSAH occurred so soon after DOAC therapy began. Each of our patients had occlusion or severe stenosis of a major artery due to emboligenic disease, and cSAH occurred in the territory of the affected artery. Reperfusion and dynamic changes in perfusion pressure due may trigger cSAH. Clinicians should remain alert for cSAH when starting DOAC for treatment of embolic ischemic stroke during the acute phase.

Dupilumab for Atopic Dermatitis, a Possible Risk Factor of Juvenile Ischemic Stroke: A Case Report.

Dupilumab, a dual inhibitor of IL-4 and IL-13 cytokine signaling, is indicated for the treatment of moderate-to-severe atopic dermatitis, which leads to the control of atopic dermatitis. The cytokines IL-4 and IL-13 are related to vascular inflammation, which is mediated by vascular endothelial cells. We report the case of a 20-year-old man with atopic dermatitis treated with dupilumab for half a year, who presented with sudden onset of dizziness, nausea, and slight cerebellar ataxia. Brain magnetic resonance imaging revealed acute infarction in the bicerebellar hemispheres. No risk factors known to be associated with ischemic stroke in young adults were detected. We suspected this ischemic stroke might be related to dupilumab. The administration of dupilumab was discontinued, and he had no recurrence subsequently. IL-4 and IL-13, anti-inflammatory cytokines secreted from T helper 2 cells, suppress proinflammatory cytokines. Therefore, dupilumab, a dual inhibitor of IL-4 and IL-13 cytokine signaling, leads to the promotion of coagulation and thrombosis. We speculate that the activation of proinflammatory cytokines in vascular endothelial cells by the inhibition of IL-4 and IL-13 signaling by dupilumab led to ischemic stroke even at a young age.

Practical guide to choosing dabigatran 150 mg twice daily or apixaban 5 mg twice daily for patients with atrial fibrillation.

Based on previous reports, we propose a practical guide to choose dabigatran 150 mg twice daily or apixaban 5 mg twice daily for patients with atrial fibrillation. We recommend the use of dabigatran 150 mg twice daily for patients with atrial fibrillation who have a high risk of embolism (e.g., ischemic stroke on other oral anticoagulants, presence of left atrial appendage thrombus) and a low risk of bleeding. However, the prevalence of such patients with atrial fibrillation is considered low because patients with atrial fibrillation with a high risk of embolism usually have a high risk of bleeding. In most other patients with atrial fibrillation, the use of apixaban 5 mg twice daily should be considered.

Intrathecal IgG Synthesis and Persistent Inflammation Are Associated with White Matter Lesions in HIV-negative Patients with Cryptococcal Meningoencephalitis.

Objective Cryptococcal meningoencephalitis (CM) causes significant morbidity and mortality in human immunodeficiency virus (HIV)-negative and HIV-positive populations. White matter lesions (WMLs) have been reported in both populations of CM patients; however, the mechanisms underlying WML formation remain unknown. We herein report the relationship between the intrathecal immune response and the development of WMLs in HIV-negative patients with CM. Methods Eleven consecutive HIV-negative patients with CM who presented at one of three emergency hospitals in Japan from April 2001 to March 2018 were enrolled. For all patients, we retrospectively assessed the relationships between clinical and laboratory information and the presence of WMLs. Results At presentation, 6 patients had WMLs on magnetic resonance imaging (MRI). The cerebrospinal fluid immunoglobulin G (CSF IgG) index was significantly higher in the patients with WMLs than in those without WMLs (mean, 1.34 vs. 0.70, p=0.017). The time from the symptom onset to initial neuroimaging was also significantly longer in the patients with WMLs than in those without WMLs (median, 31.5 vs. 7.0 days; p=0.008). The clinical outcome was comparable among the patients with and without WMLs. Conclusion In HIV-negative patients with CM, a persistent, aberrant immune response to Cryptococcus, such as intrathecal IgG synthesis, may induce WML formation.

Bi-allelic CSF1R Mutations Cause Skeletal Dysplasia of Dysosteosclerosis-Pyle Disease Spectrum and Degenerative Encephalopathy with Brain Malformation.

Colony stimulating factor 1 receptor (CSF1R) plays key roles in regulating development and function of the monocyte/macrophage lineage, including microglia and osteoclasts. Mono-allelic mutations of CSF1R are known to cause hereditary diffuse leukoencephalopathy with spheroids (HDLS), an adult-onset progressive neurodegenerative disorder. Here, we report seven affected individuals from three unrelated families who had bi-allelic CSF1R mutations. In addition to early-onset HDLS-like neurological disorders, they had brain malformations and skeletal dysplasia compatible to dysosteosclerosis (DOS) or Pyle disease. We identified five CSF1R mutations that were homozygous or compound heterozygous in these affected individuals. Two of them were deep intronic mutations resulting in abnormal inclusion of intron sequences in the mRNA. Compared with Csf1r-null mice, the skeletal and neural phenotypes of the affected individuals appeared milder and variable, suggesting that at least one of the mutations in each affected individual is hypomorphic. Our results characterized a unique human skeletal phenotype caused by CSF1R deficiency and implied that bi-allelic CSF1R mutations cause a spectrum of neurological and skeletal disorders, probably depending on the residual CSF1R function.

Remarkable improvement in progressive multifocal leukoencephalopathy following acute pyelonephritis with bacteremia.

Progressive multifocal leukoencephalopathy (PML) is caused by John Cunningham (JC) virus in immunocompromized patients such as those with human immunodeficiency virus (HIV) infection, hematological malignancy, autoimmune disorder, and immunodeficiency disorder as well as those undergoing chemotherapy or immunosuppressive therapy. No effective treatments have been established for PML, which commonly causes severe neurological sequelae. We describe the first case of PML in a patient without HIV infection who exhibited remarkable improvement following acute pyelonephritis with Escherichia coli bacteremia.

Effects of propofol versus sevoflurane on cerebral circulation time in patients undergoing coiling for cerebral artery aneurysm: a prospective randomized crossover study.

Many neuroendovascular treatments are supported by real-time anatomical and visual hemodynamic assessments through digital subtraction angiography (DSA). Here we used DSA in a single-center prospective randomized crossover study to assess the intracranial hemodynamics of patients undergoing coiling for cerebral aneurysm (n = 15) during sevoflurane- and propofol-based anesthesia. Color-coded DSA was used to define time to peak density of contrast medium (TTP) at several intravascular regions of interest (ROIs). Travel time at a particular ROI was defined as the TTP at the selected ROI minus TTP at baseline position on the internal carotid artery (ICA). Travel time at the jugular bulb on the anterior-posterior view was defined as the cerebral circulation time (CCT), which was divided into four segmental circulation times: ICA, middle cerebral artery (MCA), microvessel, and sinus. When bispectral index values were kept between 40 and 60, CCT (median [interquartile range]) was 10.91 (9.65-11.98) s under propofol-based anesthesia compared with 8.78 (8.32-9.45) s under sevoflurane-based anesthesia (P < 0.001). Circulation times for the ICA, MCA, and microvessel segments were longer under propofol-based anesthesia than under sevoflurane-based anesthesia (P < 0.05 for all). Our results suggest that, relative to sevoflurane, propofol decreases overall cerebral perfusion.

Progressive Encephalomyelitis with Rigidity and Myoclonus Resolving after Thymectomy with Subsequent Anasarca: An Autopsy Case.

Progressive encephalomyelitis with rigidity and myoclonus (PERM) is an autoimmune disorder involving the brainstem and spinal cord and is sometimes associated with thymoma. We encountered a 75-year-old woman with typical PERM features, glycine receptor antibody, and thymoma. Her neurologic symptoms improved after thymectomy, but she unexpectedly developed anasarca with massive pleural effusions and hypoalbuminemia and finally succumbed to death. The autopsy showed edema and mononuclear infiltration in the pleura but no neuropathological findings typical of PERM. Effective treatment of PERM can reverse the neuropathological signs of encephalomyelitis. The autoimmune nature of anasarca is possible but not proven.

A multi-institutional study of secondary check of treatment planning using Clarkson-based dose calculation for three-dimensional radiotherapy.

PURPOSE: As there have been few reports on quantitative analysis of inter-institutional results for independent monitor unit (MU) verification, we performed a multi-institutional study of verification to show the feasibility of applying the 3-5% action levels used in the U.S. and Europe, and also to show the results of inter-institutional comparisons. METHODS: A total of 5936 fields were collected from 12 institutions. We used commercial software employing the Clarkson algorithm for verification after a validation study of measurement and software comparisons was performed. The doses generated by the treatment planning systems (TPSs) were retrospectively analyzed using the verification software. RESULTS: Mean ±â€¯two standard deviations of all locations were 1.0 ±â€¯3.6%. There were larger differences for breast (4.0 ±â€¯4.0%) and for lung (2.5 ±â€¯5.8%). A total of 80% of the fields with differences over 5% of the action level involved breast and lung targets, with 7.2 ±â€¯5.4%. Inter-institutional comparisons showed various systematic differences for field shape for breast and differences in the fields were attributable to differences in reference point placement for lung. The large differences for breast and lung are partially attributable to differences in the methods used to correct for heterogeneity. CONCLUSIONS: The 5% action level may be feasible for verification; however, an understanding of larger differences in breast and lung plans is important in clinical practice. Based on the inter-institutional comparisons, care must be taken when determining an institution-specific action level from plans with different field shape settings and incorrectly placed reference points.

A multi-institutional study of independent calculation verification in inhomogeneous media using a simple and effective method of heterogeneity correction integrated with the Clarkson method.

In inhomogeneous media, there is often a large systematic difference in the dose between the conventional Clarkson algorithm (C-Clarkson) for independent calculation verification and the superposition-based algorithms of treatment planning systems (TPSs). These treatment site-dependent differences increase the complexity of the radiotherapy planning secondary check. We developed a simple and effective method of heterogeneity correction integrated with the Clarkson algorithm (L-Clarkson) to account for the effects of heterogeneity in the lateral dimension, and performed a multi-institutional study to evaluate the effectiveness of the method. In the method, a 2D image reconstructed from computed tomography (CT) images is divided according to lines extending from the reference point to the edge of the multileaf collimator (MLC) or jaw collimator for each pie sector, and the radiological path length (RPL) of each line is calculated on the 2D image to obtain a tissue maximum ratio and phantom scatter factor, allowing the dose to be calculated. A total of 261 plans (1237 beams) for conventional breast and lung treatments and lung stereotactic body radiotherapy were collected from four institutions. Disagreements in dose between the on-site TPSs and a verification program using the C-Clarkson and L-Clarkson algorithms were compared. Systematic differences with the L-Clarkson method were within 1% for all sites, while the C-Clarkson method resulted in systematic differences of 1-5%. The L-Clarkson method showed smaller variations. This heterogeneity correction integrated with the Clarkson algorithm would provide a simple evaluation within the range of -5% to +5% for a radiotherapy plan secondary check.

Dynamic spinal compression revealed by computed tomography myelography in overshunting-associated myelopathy: A case report.

RATIONALE: OSAM is a rare ventriculoperitoneal (VP) shunt complication where cervical spinal cord compression by epidural venous plexus engorgement is caused by cerebrospinal fluid (CSF) overdrainage. Symmetrically indented deformity of the upper cervical spinal cord and surrounding epidural venous engorgement are characteristic radiological findings. Both of them are typically detected on magnetic resonance imaging (MRI) and enhanced computed tomography (CT). PATIENT CONCERNS: The 77-year-old man who underwent the placement of a VP shunt without an antisiphon device to treat post-subarachnoid hemorrhage (SAH) hydrocephalus presented with progressive quadriplegia 10 years postoperatively. DIAGNOSIS: MRI revealed a symmetrically indented spinal cord from the craniocervical junction (CCJ) to the C2 level and enhanced CT showed the epidural venous engorgement, which were characteristic radiological findings of overshunting-associated myelopathy (OSAM). However, MRI atypically failed to detect the engorged epidural vein and showed no compressive lesion around the spinal cord. INTERVENTION: In order to reveal how the cervical spinal cord was deformed and compressed by engorged epidural vein, CT myelography was performed. OUTCOMES: CT myelography proved that the epidural vein dynamically engorged and compressed the cervical spinal cord immediately after rotation and extension of the neck. LESSONS: CT myelography combined with neck rotation and extension revealed the dynamic change of the epidural venous engorgement, and is useful for evaluation and diagnosis of OSAM especially when epidural venous engorgement was not detectable on MRI.

Sphingosine-1-Phosphate Receptor 1 Activation Enhances Leptomeningeal Collateral Development and Improves Outcome after Stroke in Mice.

BACKGROUND: Development of collateral circulation after acute ischemic stroke is triggered by shear stress that occurs in pre-existing arterioles. Recently, sphingosine-1-phosphate receptor 1 (S1P1) on endothelial cells was reported to sense shear stress and transduce its signaling pathways. METHODS: BALB/c mice (n = 118) were subjected to permanent middle cerebral artery occlusion (pMCAO) or sham operation. We investigated the effect of an S1P1-selective agonist SEW2871 on leptomeningeal collateral arteries and neurological outcome after pMCAO. RESULTS: Immunohistochemistry showed that without treatment, the expression of S1P1 on endothelial cells of leptomeningeal arteries and capillaries increased early after pMCAO, peaking at 6 hours, whereas a significant increase in the expression of S1P1 in neurons was seen from 24 hours later. After intraperitoneal administration of SEW2871 for 7 days after pMCAO, the number of leptomeningeal collateral arteries was significantly increased, cerebral blood flow improved, infarct volume was decreased, and neurological outcome improved compared with the controls. Significantly increased phosphorylation of endothelial nitric oxide synthase (eNOS) as early as 6 hours after pMCAO and higher expression of tight junction proteins at postoperative day 3 were observed with SEW2871 treatment as assessed by Western blot. Daily administration of SEW2871 also increased capillary density in peri-infarct regions and promoted monocyte/macrophage mobilization to the surface of ischemic cortex at 7 days after pMCAO. CONCLUSIONS: An S1P1-selective agonist enhanced leptomeningeal collateral circulation via eNOS phosphorylation and promoted postischemic angiogenesis with reinforced blood-brain barrier integrity in a mouse model of acute ischemic stroke, leading to smaller infarct volume and better neurological outcome.

Myasthenia Gravis Complicated with Peripheral T-cell Lymphoma, Not Otherwise Specified (PTCL-NOS), Following Thymectomy and Longstanding Tacrolimus Therapy.

Myasthenia gravis (MG), a neuromuscular junction autoimmune disease, sometimes complicates second malignancies; however, T-cell lymphoproliferative disorders have rarely been reported. A 55-year-old man, who received oral tacrolimus and prednisolone for MG for 16 years after thymectomy, presented with left abdominal pain, lymphadenopathy, and splenomegaly. A lymph node biopsy revealed peripheral T-cell lymphoma, not otherwise specified (PTCL-NOS). This is the first report of oral tacrolimus leading to a T-cell lymphoproliferative disorder in patient without a history of transplantation. Physicians should be aware of the possibility of rare T-cell lymphoproliferative disorders, such as PTCL-NOS, occurring as complications in MG patients on immunosuppressive regimens after thymectomy.