Human Herpesvirus-6 corneal Endotheliitis after intravitreal injection of Ranibizumab.

BACKGROUND: To report the first case of human herpesvirus-6 (HHV-6) corneal endotheliitis that developed after intravitreal ranibizumab injections. CASE PRESENTATION: A 63-year-old man with a medical history of diabetes and systemic steroid treatment for bullous pemphigoid had been receiving intravitreal injections of ranibizumab in the left eye for 2 years according to a Pro Re Nata treatment regimen for macular edema associated with branch retinal vein occlusion. Twenty days after the last injection, the patient presented with pain and decreased visual acuity in his left eye. His best corrected visual acuity in the left eye was 2/200, and intraocular pressure was 45 mmHg with edema of the central stromal cornea, mild conjunctival injection, intermediate keratic precipitates, and mild anterior chamber reaction. HHV-6 DNA was detected in the aqueous humor using multiplex strip polymerase chain reaction, and it was identified as variant A, HHV-6A. A diagnosis of HHV-6A-associated corneal endotheliitis was made. Oral valganciclovir and topical ganciclovir therapy was initiated with good resolution of all symptoms and signs. CONCLUSIONS: HHV-6A can be a possible complication of intravitreal ranibizumab therapy. To the best of our knowledge, this is the first reported case of HHV-6A corneal endotheliitis following intravitreal ranibizumab injection.

Pediatric brain repair from endogenous neural stem cells of the subventricular zone.

There is great interest in the regenerative potential of the neural stem cells and progenitors that populate the germinal zones of the immature brain. Studies using animal models of pediatric brain injuries have provided a clearer understanding of the responses of these progenitors to injury. In this review, we have compared and contrasted the responses of the endogenous neural stem cells and progenitors of the subventricular zone in animal models of neonatal cerebral hypoxia-ischemia, neonatal stroke, congenital cardiac disease, and pediatric traumatic brain injury. We have reviewed the dynamic shifts that occur within this germinal zone with injury as well as changes in known signaling molecules that affect these progenitors. Importantly, we have summarized data on the extent to which cell replacement occurs in response to each of these injuries, opportunities available, and obstacles that will need to be overcome to improve neurological outcomes in survivors.

Abnormal RNA structures (RNA foci) containing a penta-nucleotide repeat (UGGAA)n in the Purkinje cell nucleus is associated with spinocerebellar ataxia type 31 pathogenesis.

Spinocerebellar ataxia type 31 (SCA31) is an autosomal-dominant cerebellar ataxia showing a Purkinje cell (PC)-predominant neurodegeneration in humans. The mutation is a complex penta-nucleotide repeat containing (TGGAA)n , (TAGAA)n , (TAAAA)n and (TAGAATAAAA)n inserted in an intron shared by two different genes BEAN1 and TK2 located in the long arm of the human chromosome 16. Previous studies have shown that (TGGAA)n is the critical component of SCA31 pathogenesis while the three other repeats, also present in normal Japanese, are not essential. Importantly, it has been shown that BEAN1 and TK2 are transcribed in mutually opposite directions in the human brain. Furthermore, abnormal RNA structures called "RNA foci" are observed by a probe against (UAGAAUAAAA)n in SCA31 patients' PC nuclei, indicating that the BEAN1-direction mutant transcript appears instrumental for the pathogenesis. However, it is not known whether the critical repeat (TGGAA)n contributes to the formation of RNA foci, neither do we understand how the RNA foci formation is relevant to the pathogenesis. To address these issues, we investigated two SCA31 cerebella by fluorescence in situ hybridization using a probe against (UGGAA)n . We also asked whether the mutant BEAN1-transcript containing (UGGAA)n exerts toxicity compared to the other three repeats in cultured cells. Histopathologically, we confirm that the PC is the main target of SCA31 pathogenesis. We find that the RNA foci containing (UGGAA)n are indeed observed in PC nuclei of both SCA31 patients, whereas similar foci were not observed in control individuals. In both transiently and stably expressed cultured cell models, we also find that the mutation transcribed in the BEAN1-direction yields more toxicity than control transcripts and forms RNA foci detected with probes against (UGGAA)n and (UAGAAUAAAA)n . Taking these findings together, we conclude that the RNA foci containing BEAN1-direction transcript (UGGAA)n are associated with PC degeneration in SCA31.

Prevalence of inositol 1, 4, 5-triphosphate receptor type 1 gene deletion, the mutation for spinocerebellar ataxia type 15, in Japan screened by gene dosage.

Spinocerebellar ataxia type 15 (SCA15) is an autosomal dominant neurodegenerative disorder clinically characterized by late-onset, slowly progressive pure cerebellar ataxia. This disease is caused by a heterozygous deletion of the inositol 1, 4, 5-triphosphate receptor type 1 (ITPR1) gene, suggesting that haploinsufficiency of the receptor function is the plausible disease mechanism. To clarify the prevalence of SCA15 in Japan, we designed four sets of probes and primers in different regions of ITPR1 and performed TaqMan PCR assay to search for gene deletions in 226 index SCA patients excluded for repeat expansion disorders. Deletion was found in only one patient, in whom gait ataxia started at 51 years of age and progressed to show cerebellar ataxia. This study demonstrates a simple but efficient method for screening ITPR1 deletion. We also conclude that ITPR1 gene deletions are much rare in Japan than in Europe, comprising only 0.3% in all SCAs in Japan.

Reduced brain-derived neurotrophic factor (BDNF) mRNA expression and presence of BDNF-immunoreactive granules in the spinocerebellar ataxia type 6 (SCA6) cerebellum.

Spinocerebellar ataxia type 6 (SCA6) is an autosomal-dominant neurodegenerative disorder caused by a small expansion of tri-nucleotide (CAG) repeat encoding polyglutamine (polyQ) in the gene for α(1A) voltage-dependent calcium channel (Ca(v) 2.1). Thus, this disease is one of the nine neurodegenerative disorders called polyQ diseases. The Purkinje cell predominant neuronal loss is the characteristic neuropathology of SCA6, and a 75-kDa carboxy-terminal fragment (CTF) of Ca(v) 2.1 containing polyQ, which remains soluble in normal brains, becomes insoluble in the cytoplasm of SCA6 Purkinje cells. Because the suppression of the brain-derived neurotrophic factor (BDNF) expression is a potentially momentous phenomenon in many other polyQ diseases, we implemented BDNF expression analysis in SCA6 human cerebellum using quantitative RT-PCR for the BDNF mRNA, and by immunohistochemistry for the BDNF protein. We observed significantly reduced BDNF mRNA levels in SCA6 cerebellum (n = 3) compared to controls (n = 6) (Mann-Whitney U-test, P = 0.0201). On immunohistochemistry, BDNF protein was only weakly stained in control cerebellum. On the other hand, we found numerous BDNF-immunoreactive granules in dendrites of SCA6 Purkinje cells. We did not observe similar BDNF-immunoreactive granules in other polyQ diseases, such as Huntington's disease or SCA2. As we often observed that the 1C2-positive Ca(v) 2.1 aggregates existed more proximally than the BDNF-positive granules in the dendrites, we speculated that the BDNF protein trafficking in dendrites may be disturbed by Ca(v) 2.1 aggregates in SCA6 Purkinje cells. We conclude that the SCA6 pathogenic mechanism associates with the BDNF mRNA expression reduction and abnormal localization of BDNF protein.

Safety evaluation of a clinical focused ultrasound system for neuronavigation guided blood-brain barrier opening in non-human primates.

An emerging approach with potential in improving the treatment of neurodegenerative diseases and brain tumors is the use of focused ultrasound (FUS) to bypass the blood-brain barrier (BBB) in a non-invasive and localized manner. A large body of pre-clinical work has paved the way for the gradual clinical implementation of FUS-induced BBB opening. Even though the safety profile of FUS treatments in rodents has been extensively studied, the histological and behavioral effects of clinically relevant BBB opening in large animals are relatively understudied. Here, we examine the histological and behavioral safety profile following localized BBB opening in non-human primates (NHPs), using a neuronavigation-guided clinical system prototype. We show that FUS treatment triggers a short-lived immune response within the targeted region without exacerbating the touch accuracy or reaction time in visual-motor cognitive tasks. Our experiments were designed using a multiple-case-study approach, in order to maximize the acquired data and support translation of the FUS system into human studies. Four NHPs underwent a single session of FUS-mediated BBB opening in the prefrontal cortex. Two NHPs were treated bilaterally at different pressures, sacrificed on day 2 and 18 post-FUS, respectively, and their brains were histologically processed. In separate experiments, two NHPs that were earlier trained in a behavioral task were exposed to FUS unilaterally, and their performance was tracked for at least 3 weeks after BBB opening. An increased microglia density around blood vessels was detected on day 2, but was resolved by day 18. We also detected signs of enhanced immature neuron presence within areas that underwent BBB opening, compared to regions with an intact BBB, confirming previous rodent studies. Logistic regression analysis showed that the NHP cognitive performance did not deteriorate following BBB opening. These preliminary results demonstrate that neuronavigation-guided FUS with a single-element transducer is a non-invasive method capable of reversibly opening the BBB, without substantial histological or behavioral impact in an animal model closely resembling humans. Future work should confirm the observations of this multiple-case-study work across animals, species and tasks.

Neuroregenerative and protective functions of Leukemia Inhibitory Factor in perinatal hypoxic-ischemic brain injury.

Neonatal hypoxic-ischemic encephalopathy remains the most important neurological problem of the newborn. Delays in diagnosing perinatal brain injuries are common, preventing access to acute therapies. Therefore, there is a critical need for therapeutic strategies that are beneficial when delivered beyond 24 h after birth. Here we show that Leukemia Inhibitory Factor (LIF) functions as an essential injury-induced neurotrophic cytokine in the CNS and that non-invasively administering LIF as late as 3 days after a hypoxic-ischemic insult improves neurological function. Using a mouse model of late preterm brain injury we show that astroglial and microglial/macrophage reactivity to hypoxia-ischemia was diminished at 3 days of recovery, but then exacerbated at 2 weeks of recovery in LIF haplodeficient mice. There also were significantly more CD68+/Iba-1+ cells in the ipsilateral striatum in LIF-Het mice compared to WT mice at 2 weeks of recovery. This desynchronized glial response was accompanied by increased neuronal cell death in the striatum and neocortex (Fluorojade C), hypomyelination (reduced MBP staining and thinner external capsule), increased extent of brain damage (Nissl) and diminished neurological function on sensorimotor tests. To our surprise, injured LIF-Het mice had ~7-fold higher IGF-1 levels than injured WT mice at 3 days after HI injury. Intranasally administered LIF activated the Jak-Stat-3 pathway both within the subventricular zone and the neocortex at 30 min after administration. When delivered with a delay of 3 days after the insult, intranasal LIF reduced the extent of brain injury by ~60%, attenuated astrogliosis and microgliosis in striatum, improved subcortical white matter thickness, increased numbers of Olig2+ cells in corpus callosum and improved performance on sensorimotor tests at 2 weeks of recovery. These studies provide key pre-clinical data recommending LIF administration as a neuroprotectant and regenerative cytokine and they highlight the feasibility of pursuing new therapeutics targeting the tertiary phase of neurodegeneration for hypoxic-ischemic encephalopathies.

A case of childhood glaucoma with a combined partial monosomy 6p25 and partial trisomy 18p11 due to an unbalanced translocation.

Background: Chromosomal deletion involving the 6p25 region results in a clinically recognizable syndrome characterized by anterior eye chamber anomalies with risk of glaucoma and non-ocular malformations (6p25 deletion syndrome). We report a newborn infant case of childhood glaucoma with a combination of partial monosomy 6p25 and partial trisomy 18p11 due to an unbalanced translocation.Materials and methods: The patient was a 0-year-old girl. Both eyes showed aniridia and left eye Peters anomaly with multiple malformations. To identify the chromosomal aberrations in the patient with clinically suspected 6p25 deletion syndrome, we performed cytogenetic analysis (G-banding and multicolor fluorescent in-situ hybridization) and array-based comparative genomic hybridization (array-CGH) analysis.Results: Cytogenetic analyses revealed a derivative chromosome 6 with its distal short arm replaced by an extra copy of the short arm of chromosome 18. Array-CGH analysis detected a 4.6-Mb deletion at 6pter to 6p25.1 and 8.9-Mb duplication at 18pter to 18p11.22. To determine the breakpoint of the unbalanced rearrangement at the single-base level, we performed a long-range PCR for amplifying the junctional fragment of the translocation breakpoint. By sequencing the junctional fragment, we defined the unbalanced translocation as g.chr6:pter_4594783delinschr18:pter_8911541.Conclusions: A phenotype corresponding to combined monosomy 6p25 and trisomy 18p11 presented as childhood glaucoma associated with non-acquired (congenital) ocular anomalies consist of aniridia and Peters anomaly and other systemic malformations. To the best of our knowledge, this is the first report which demonstrated the breakpoint sequence of an unbalanced translocation in a Japanese infant with childhood glaucoma.

[Molecular genetic approach to spinocerebellar ataxias].

Spinocerebellar ataxia (SCA) is a group of degenerative ataxias with autosomal dominant inheritance. The most common form of mutation that causes SCA is the expansion of trinucleotide (CAG) repeat encoding polyglutamine. These "polyglutamine disorders" are, SCA1, SCA2, Machado-Joseph disease, SCA6, SCA7, SCA17 and DRPLA. Another dynamic mutation, yet a non-coding one, has been identified as the cause of SCA8, SCA10 and SCA12. This mutation includes, trinucleotide (CAG/CTG) expansion causing SCA8 and SCA12, and pentanuclotide (ATTCT) expansion leading SCA10. In addition to these dynamic mutations, static mutations, such as missense mutations and deletions, have been identified to cause SCA5, SCA11, SCA13, SCA14, SCA15 and SCA27. Since 1992, authors have been involved in identifying the mutation (s) of autosomal dominant cerebellar ataxia with rather pure cerebellar syndrome (ADCAIII). About a half of our cohort with ADCAIII were SCA6, caused by a small CAG repeat expansion in the alpha1A-voltage-dependent calcium channel gene. Recent study in patients' brains suggested that a small polyglutamine expansion leads a portion of this channel protein to aggregate in the Purkinje cell. Another type of ADCAIII is the chromosome 16q22.1-linked ADCA. By a comprehensive positional cloning strategy, we have found a genetic change that segregate with the disease. Identifying the mutation of 16q-ADCA is imperative for understanding molecular basis of this disease.

Focused ultrasound enhanced intranasal delivery of brain derived neurotrophic factor produces neurorestorative effects in a Parkinson's disease mouse model.

Focused ultrasound-enhanced intranasal (IN + FUS) delivery is a noninvasive approach that utilizes the olfactory pathway to administer pharmacological agents directly to the brain, allowing for a more homogenous distribution in targeted locations compared to IN delivery alone. However, whether such a strategy has therapeutic values, especially in neurodegenerative disorders such as Parkinson's disease (PD), remains to be established. Herein, we evaluated whether the expression of tyrosine hydroxylase (TH), the rate limiting enzyme in dopamine catalysis, could be enhanced by IN + FUS delivery of brain-derived neurotrophic factor (BDNF) in a toxin-based PD mouse model. Mice were put on the subacute dosing regimen of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP), producing bilateral degeneration of the nigrostriatal pathway consistent with early-stage PD. MPTP mice then received BDNF intranasally followed by multiple unilateral FUS-induced blood-brain barrier (BBB) openings in the left basal ganglia for three consecutive weeks. Subsequently, mice were survived for two months and were evaluated morphologically and behaviorally to determine the integrity of their nigrostriatal dopaminergic pathways. Mice receiving IN + FUS had significantly increased TH immunoreactivity in the treated hemisphere compared to the untreated hemisphere while mice receiving only FUS-induced BBB opening or no treatment at all did not show any differences. Additionally, behavioral changes were only observed in the IN + FUS treated mice, indicating improved motor control function in the treated hemisphere. These findings demonstrate the robustness of the method and potential of IN + FUS for the delivery of bioactive factors for treatment of neurodegenerative disorder.

Unilateral Focused Ultrasound-Induced Blood-Brain Barrier Opening Reduces Phosphorylated Tau from The rTg4510 Mouse Model.

The neuropathological hallmarks of Alzheimer's disease include amyloid plaques and neurofibrillary tangles. Tau pathology correlates well with impaired neuronal activity and dementia. Focused ultrasound coupled with systemic administration of microbubbles has previously been shown to open the blood-brain barrier and induce an immune response, which, in an amyloid AD mouse model, resulted in the reduction of the amyloid brain load. Methods: In this study, we investigated the effect of focused ultrasound at the early stages of tau pathology (pre-tangle) in the rTg4510 mouse model. Results: Reduction of phosphorylated tau from the hippocampal formation processes, and particularly the pyramidal CA1 neurons, was noted in the ultrasound-treated brains without an associated increase in the phosphorylated tau-affected cell somas, typically associated with disease progression. Attenuation of the pathology was found to correlate well with the ultrasound-initiated immune response without compromising neuronal integrity. Unilateral ultrasound application resulted in a bilateral effect indicating a broader reduction of the phosphorylated tau. Conclusion: Findings presented herein reinforce the premise of ultrasound in reducing tau pathology and thus curbing the progression of Alzheimer's disease.

Elderly onset vitreous opacities as the initial manifestation in hereditary transthyretin (ATTR Val30Met) carries.

Familial amyloid polyneuropathy (FAP) is a systemic disorder transmitted as an autosomal dominant trait and is characterized by deposits of protein fibrils in various organs leading to physiologic dysfunction. In cases with FAP in Japanese endemic foci, their signs and symptoms often develop before the age of 40 years. We report on two elderly patients (an 80-year-old woman and an 83-year-old man) with progressive vitreous opacities (VOs) as the initial manifestation of hereditary transthyretin (ATTR Val30Met) carries, who had no evidence of systemic involvement or family history of amyloidosis and lived in non-endemic areas. Therapeutic vitrectomy with extensive vitreous removal combined with cataract surgery was performed. Clinicians should consider the possibility of hereditary transthyretin carries in cases presenting with VOs of undetermined etiology to avoid misdiagnosis.

Cytoplasmic location of α1A voltage-gated calcium channel C-terminal fragment (Cav2.1-CTF) aggregate is sufficient to cause cell death.

The human α1A voltage-dependent calcium channel (Cav2.1) is a pore-forming essential subunit embedded in the plasma membrane. Its cytoplasmic carboxyl(C)-tail contains a small poly-glutamine (Q) tract, whose length is normally 4∼19 Q, but when expanded up to 20∼33Q, the tract causes an autosomal-dominant neurodegenerative disorder, spinocerebellar ataxia type 6 (SCA6). A recent study has shown that a 75-kDa C-terminal fragment (CTF) containing the polyQ tract remains soluble in normal brains, but becomes insoluble mainly in the cytoplasm with additional localization to the nuclei of human SCA6 Purkinje cells. However, the mechanism by which the CTF aggregation leads to neurodegeneration is completely elusive, particularly whether the CTF exerts more toxicity in the nucleus or in the cytoplasm. We tagged recombinant (r)CTF with either nuclear-localization or nuclear-export signal, created doxycyclin-inducible rat pheochromocytoma (PC12) cell lines, and found that the CTF is more toxic in the cytoplasm than in the nucleus, the observations being more obvious with Q28 (disease range) than with Q13 (normal-length). Surprisingly, the CTF aggregates co-localized both with cAMP response element-binding protein (CREB) and phosphorylated-CREB (p-CREB) in the cytoplasm, and Western blot analysis showed that the quantity of CREB and p-CREB were both decreased in the nucleus when the rCTF formed aggregates in the cytoplasm. In human brains, polyQ aggregates also co-localized with CREB in the cytoplasm of SCA6 Purkinje cells, but not in other conditions. Collectively, the cytoplasmic Cav2.1-CTF aggregates are sufficient to cause cell death, and one of the pathogenic mechanisms may be abnormal CREB trafficking in the cytoplasm and reduced CREB and p-CREB levels in the nuclei.

[Dissecting molecular mechanism of spinocerebellar ataxia type 31].

Spinocerebellar ataxia is a group of neurodegenerative disorders clinically presenting adult onset cerebellar ataxia. To date, 21 different genes (SCA1, 2, 3, 5, 6, 7, 8, 10, 11, 12, 13, 14, 15, 17, 23, 27, 28, 31, 35, 36 and DRPLA) and additionally 10 different gene loci (SCA4, 18, 19, 20, 21, 25, 26, 29, 30 and 32) are identified. Among these, SCA6 and SCA31 are the two common diseases clinically presenting as a relatively predominant cerebellar syndrome, whereas Machado-Joseph disease/SCA3, DRPLA, SCA1 and SCA2 are SCAs often associated with extra-cerebellar manifestations. SCA31 is a late-onset purely cerebellar ataxia caused by a complex pentanucleotide repeat containing (TGGAA)(n) lying in an intronic region shared by two genes, BEAN (brain expressed, associated with NEDD4) and TK2 (thymidine kinase 2). In situ hybridization analysis in patients' Purkinje cells demonstrated that pentanucleotide repeats transcribed in BEAN direction form RNA aggregates ("RNA foci"), and essential splicing factors, SFRS1 and SFRS9, bind to (UGGAA)(n), the transcript of (TGGAA)(n)in vitro. Our preliminary data also demonstrated that (UGGAA)(n) is toxic when expressed in cultured cells. These findings may imply that RNA-mediated pathogenesis is involved in SCA31. Further studies are needed to explore precise mechanism of this disease.

[Spinocerebellar ataxia type 31].

Spinocerebellar ataxia type 31 (SCA31) is a relatively common degenerative ataxia in Japan. We recently discovered SCA31 mutation as a complex pentanucleotide repeat containing (TAAAA)(n), (TAGAA)(n), and (TGGAA)(n). The size of this repeat ranged from 2.8 to 3.5 kilo-base pairs (kb). Among these repeats, (TGGAA)(n) repeat appears crucial for SCA31 pathogenesis. The length of this complex repeat inversely correlated with ages of onset in patients. The mutation lies in an intron shared by two different genes, BEAN (brain expressed, associated with NEDD4) and TK2 (thymidine kinase 2), which are transcribed in opposite directions. Thus, the complex pentanucleotide sequence is predicted to be transcribed in both directions, but not necessarily translated into proteins. In situ hybridization analysis in patients' Purkinje cells demonstrated that pentanucleotide repeats transcribed in BEAN direction form RNA aggregates ("RNA foci"). We further found that splicing factors, SFRS1 and SFRS9, binds to (UGGAA)(n), the transcript of (TGGAA)(n) in vitro. These findings may imply that SCA31 conforms to pathogenic mechanisms underlying non-coding repeat disorders, such as myotonic dystrophies (DM1 & DM2), and that SFRS1 and SFRS9 are involved in SCA31 pathogenesis.