Restless legs syndrome in Parkinson's disease: relationship with quality of life and medication.

OBJECTIVES: We aimed to disclose the relationship between restless leg syndrome (RLS) and antiparkinsonian treatment, and its effect on quality of life (QoL) in patients with Parkinson's disease (PD). BACKGROUND: Previous studies documented the prevalence of RLS among patients with PD to be higher than in the general population, but conclusions regarding the aetiology and impact were contradictory. METHODS: We examined 101 patients with idiopathic PD. All participants completed the five-dimension/five-level-EuroQoL questionnaire (EQ-5D-5L) and the International Restless-Legs-syndrome-study-group rating Scale (IRLS). RESULTS: The prevalence of RLS was 22.77 %. There were no statistically significant differences in levodopa or dopamine agonists (DA) doses between RLS-positive and negative participants. However, the use of levodopa as the last night-time medication was connected with a higher risk of RLS (OR=2.049, p=0.041). There was significantly lower prevalence of RLS in patients after surgical treatment for PD (p=0.024). Participants with RLS were at a greater risk for sleep disturbances (OR=3.866, p=0.023) and excessive daytime sleepiness (OR=7.202, p<0.001). Greater RLS symptoms were associated with worse QoL (higher IRLS score predicted higher EQ5D5L score, p=0.023). CONCLUSION: RLS is prevalent among PD patients and night-time dopaminergic over-excitation with levodopa plays an important role in its pathogenesis. Since the symptoms of RLS are associated with decreased QoL, early accurate diagnosis and appropriate adjustment of dopaminergic therapy can lead to immediate relief from RLS symptoms and to QoL improvement (Tab. 4, Fig. 1, Ref. 34).

Presynaptic GABAA Receptors Modulate Thalamocortical Inputs in Layer 4 of Rat V1.

Fast inhibitory GABAergic transmission plays a fundamental role in neural circuits. Current theories of cortical function assume that fast GABAergic inhibition acts via GABAA receptors on postsynaptic neurons, while presynaptic effects of GABA depend on GABAB receptor activation. Manipulations of GABAA receptor activity in vivo produced different effects on cortical function, which were generally ascribed to the mode of action of a drug, more than its site of action. Here we show that in rodent primary visual cortex, α4-containing GABAA receptors can be located on subsets of glutamatergic and GABAergic presynaptic terminals and decrease synaptic transmission. Our data provide a novel mechanistic insight into the effects of changes in cortical inhibition; the ability to modulate inputs onto cortical circuits locally, via presynaptic regulation of release by GABAA receptors.

Prevalent placement error of deep brain stimulation electrode in movement disorders (technical considerations).

BACKGROUND: Deep brain stimulation is an effective and safe technique. Displacement of the electrode relative to the optimal stimulation site can lead to insufficient effect and sometimes to the need of operative electrode re-position. OBJECTIVE: This study was aimed to analyse targeting accuracy of deep brain stimulation electrode implantation to subthalamic nucleus (STN) and globus pallidus internus (Gpi). It detected possible causes of inaccuracy and prevalent shift to certain direction. METHODS: Targeting accuracy was analysed in 47 patients with Parkinson´s disease (PD) and 11 patients with dystonia with bilateral implantation of deep brain stimulation electrodes between years 2009 and 2016. RESULTS: A shift of electrode to prevalent direction was observed on the left side to medial and posterior and on the right side to lateral direction. Greater shift was observed on the left side and in a higher angulation of trajectory laterally. Movement of the electrode, because of its traction in anchoring device, was identified as a possible factor for prevalent electrode shift. Calibration of stereotactic coordinates to correct prevalent shift was used. CONCLUSION: Targeting inaccuracy is the result of accumulation of errors in individual steps of electrode implantation. Direction of the shift can be random or it can be toward a prevalent direction. A correction of prevalent error can prevent a suboptimal electrode placement (Tab. 3, Fig. 11, Ref. 29).

Graft-Infiltrating Macrophages Adopt an M2 Phenotype and Are Inhibited by Purinergic Receptor P2X7 Antagonist in Chronic Rejection.

Macrophages exhibit diverse phenotypes and functions; they are also a major cell type infiltrating chronically rejected allografts. The exact phenotypes and roles of macrophages in chronic graft loss remain poorly defined. In the present study, we used a mouse heart transplant model to examine macrophages in chronic allograft rejection. We found that treatment of C57BL/6 mice with CTLA4 immunoglobulin fusion protein (CTLA4-Ig) prevented acute rejection of a Balb/c heart allograft but allowed chronic rejection to develop over time, characterized by prominent neointima formation in the graft. There was extensive macrophage infiltration in the chronically rejected allografts, and the graft-infiltrating macrophages expressed markers associated with M2 cells but not M1 cells. In an in vitro system in which macrophages were polarized into either M1 or M2 cells, we screened phenotypic differences between M1 and M2 cells and identified purinergic receptor P2X7 (P2x7r), an adenosine triphosphate (ATP)-gated ion channel protein that was preferentially expressed by M2 cells. We further showed that blocking the P2x7r using oxidized ATP (oATP) inhibited M2 induction in a dose-dependent fashion in vitro. Moreover, treatment of C57BL/6 recipients with the P2x7r antagonist oATP, in addition to CTLA4-Ig treatment, inhibited graft-infiltrating M2 cells, prevented transplant vasculopathy, and induced long-term heart allografts survival. These findings highlight the importance of the P2x7r-M2 axis in chronic rejection and establish P2x7r as a potential therapeutic target in suppression of chronic rejection.

Cytoplasmic retention of Xenopus nuclear factor 7 before the mid blastula transition uses a unique anchoring mechanism involving a retention domain and several phosphorylation sites.

Xenopus nuclear factor 7 (xnf7) is a maternally expressed protein that belongs to the B-box zinc finger gene family consisting of transcription factors, protooncogenes, and ribonucleoproteins. Its function is regulated by retention in the cytoplasm from oocyte maturation until the mid blastula transition (MBT) when it reenters the nucleus. We defined a 22-amino acid cytoplasmic retention domain (CRD) in xnf7 that functioned cooperatively with two phosphorylation sites within the xnf7 molecule to retain the protein in the cytoplasm until the MBT. Deletion of this region or mutations in the phosphorylation sites resulted in the early entry of xnf7 into the nucleus. A mutation changing one of the phosphorylation sites to a glutamic acid resulted in the prolonged retention of the xnf7 protein in the cytoplasm until stages 9-10, well past the MBT. Additionally, a mutant form of xnf7 possessing a second nuclear localization signal at the COOH terminus was retained in the cytoplasm. This suggests that retention of xnf7 was not due to the masking of its NLS as is the case with NFkB and dorsal but was due to a novel anchoring mechanism in which the CRD interacts with an anchor protein. The CRD sequence is also found in another B-box zinc finger protein that is also retained in the cytoplasm until the MBT in the newt. Therefore, we believe that this may be an important mechanism whereby the function of a number of nuclear proteins is regulated during development.

Delocalization of Vg1 mRNA from the vegetal cortex in Xenopus oocytes after destruction of Xlsirt RNA.

The Xlsirts are a family of transcribed repeat sequence genes that do not code for protein. Xlsirt RNAs become localized to the vegetal cortex of Xenopus oocytes early in oogenesis, before the localization of the messenger RNA Vg1, which encodes a transforming growth factor-beta-like molecule involved in mesoderm formation, and coincident with the localization of Xcat2 transcripts, which encode a nanos-like molecule. Destruction of the localized Xlsirts by injection of antisense oligodeoxynucleotides into stage 4 oocytes resulted in the release of Vg1 transcripts but not Xcat2 transcripts from the vegetal cortex. Xlsirt RNAs, which may be a structural component of the vegetal cortex, are a crucial part of a genetic pathway necessary for the proper localization of Vg1 that leads to subsequent normal pattern formation.

Translocation of repetitive RNA sequences with the germ plasm in Xenopus oocytes.

Xlsirts are a family of interspersed repeat RNAs from Xenopus laevis that contain from 3 to 13 repeat units (each 79 to 81 nucleotides long) flanked by unique sequences. They are homologous to the mammalian Xist gene that is involved in X chromosome inactivation. Xlsirt RNA appears first in the mitochondrial cloud (Balbiani body) in stage 2 oocytes and is then translocated as island-like structures to the vegetal cortex at early stage 3 coincident with the localization of the germ plasm. Exogenous Xlsirt RNA injected into oocytes translocates to the location of the endogenous RNA at that particular stage. The Xlsirt RNA repeat sequences are required for translocation and can cause the translocation of heterologous unique RNAs to the vegetal cortex.

Donor and host photoreceptors engage in material transfer following transplantation of post-mitotic photoreceptor precursors.

Photoreceptor replacement by transplantation is proposed as a treatment for blindness. Transplantation of healthy photoreceptor precursor cells into diseased murine eyes leads to the presence of functional photoreceptors within host retinae that express an array of donor-specific proteins. The resulting improvement in visual function was understood to be due to donor cells integrating within host retinae. Here, however, we show that while integration occurs the majority of donor-reporter-labelled cells in the host arises as a result of material transfer between donor and host photoreceptors. Material transfer does not involve permanent donor-host nuclear or cell-cell fusion, or the uptake of free protein or nucleic acid from the extracellular environment. Instead, RNA and/or protein are exchanged between donor and host cells in vivo. These data require a re-evaluation of the mechanisms underlying rescue by photoreceptor transplantation and raise the possibility of material transfer as a strategy for the treatment of retinal disorders.

RNA localization and germ cell determination in Xenopus.

In many organisms the proper development of the embryo depends on the asymmetrical distribution of maternal RNAs and proteins in the egg. Although the Xenopus oocyte is radially symmetrical it contains distinct populations of maternal RNAs that are localized either in the animal or vegetal pole. The process of localization of RNAs in Xenopus oocytes occurs during the long period of oocyte differentiation and growth that is accompanied by the elaboration of oocyte polarity. Some of the vegetally localized RNAs, such as Vg1, VegT, and Xwnt11, are involved in axial patterning and germ layer specification. Others, such as Xdazl and Xcat2, which are located in the germ plasm, are likely to play a role in the specification of germ cell fate. We will discuss the different aspects of RNA localization in Xenopus in the context of the differentiation of the germ cells and the development of the oocyte polarity.

Apparent continuity between the messenger transport organizer and late RNA localization pathways during oogenesis in Xenopus.

The localization of RNAs at the vegetal cortex in Xenopus oocytes is a complex process, involving at least two different pathways. The early, or messenger transport organizer (METRO), pathway, localizes RNAs such as Xlsirts, Xcat2 and Xwnt11 during stages 1 and 2 of oogenesis, while the late pathway localizes RNAs such as Vg1 during stages 2-4. We demonstrate that the onset of Vg1 localization is characterized by its microtubule-independent binding to a subdomain of the endoplasmic reticulum (ER). The formation of this unique ER structure is intimately associated with the movement of the mitochondrial cloud toward the vegetal cortex. In addition, we demonstrate that the mitochondrial cloud contains a gamma-tubulin-positive structure that may function as a microtubule organizing center for establishing microtubule tracks for Vg1 localization. These data, support, although they do not prove, a model in which the development of the late pathway machinery relies upon the prior functioning of the early pathway.

The cloning and characterization of a localized maternal transcript in Xenopus laevis whose zygotic counterpart is detected in the CNS.

We have cloned a cDNA (xlan4) from a Xenopus laevis oocyte cDNA library whose cognate mRNA is localized in the animal pole region of full grown oocytes. The cDNA can be translated in vitro to produce a predicted size protein of 35 kDa and, is also expressed in E. coli as a fusion protein. The conceptual protein encoded by the xlan4 cDNA is 17.5% proline rich and possesses several PEST sequences found in proteins with short half-lives. The xlan4 mRNA is 2.6 kb and during early development its titer decreases until the neurula stage after which it begins to reaccumulate. Northern blots on dissected embryos and in situ hybridization revealed that the zygotic expression is limited to the dorsal axial structures consisting primarily of the CNS. UV irradiation of the vegetal pole region immediately following fertilization that produces ventralized embryos results in a loss of zygotic xlan4 expression. In the adult, xlan4 mRNA is limited primarily to the brain. The presence of this mRNA in animal pole region which contributes to the future neural cell lineages suggests that this gene product may function either in the specification of neural cell types or in a neural specific function.

The vegetally localized mRNA fatvg is associated with the germ plasm in the early embryo and is later expressed in the fat body.

Vegetally localized RNAs in Xenopus oocytes have been implicated in the establishment of the primary germ layers and the formation and development of the primordial germ cells. fatvg mRNA is localized through the late pathway to the vegetal cortex. Like Vg1 mRNA fatvg is distributed throughout the entire cortex; however, unlike Vg1 there is a small fraction of the fatvg mRNA that is associated with the mitochondrial cloud. In early cleavage stage embryos, fatvg mRNA is associated with the germ plasm located at the tips of the vegetal blastomeres of the embryo. While several localized RNAs that follow the Message Transport Organizer (METRO) pathway have been found in the germ plasm in embryos, fatvg is a late pathway RNA that is associated with the germ plasm. In tadpoles, fatvg mRNA shows a novel pattern of expression which is distinct from the germ cell lineage and is detected at the dorso-anterior margin of the endodermal mass along the midline in two clusters of cells. fatvg mRNA expression is also detected later in the developing fat bodies, the major adipose tissues of the frog.

Contribution of METRO pathway localized molecules to the organization of the germ cell lineage.

To elucidate the potential role of localized components in the specification of the germ cell lineage we analyzed the composition of the germ plasm in Xenopus laevis oocytes and early embryos with respect to the vegetally-localized RNAs. We focused on Xlsirts, Xcat2, and Xwnt11 transcripts that are localized to the vegetal cortex through a region of the mitochondrial cloud called the messenger transport organizer (METRO) that also contains the nuage or germ plasm. At the ultrastructural level Xcat2 mRNA was detected on germinal granules while Xlsirts and Xwnt11 were associated with a fibrillar network of the germ plasm in stage-1 and stage-4 oocytes. In embryos, we found that all three RNAs remained associated with the germ plasm. Vg1 mRNA, a transcript localized through the late pathway, was excluded from the germ plasm in oocytes and embryos. Addtionally, we detected the protein spectrin within 16 cell nests of germ cells, in a structure reminiscent of the Drosophila spectrosome. Spectrin was detected in the mitochondrial cloud and was found in the germ plasm during embryogenesis. These data indicate that the various RNAs found within METRO and the protein spectrin are integral components of the Xenopus germ plasm with the RNAs being associated with different subcellular structures. They also suggest that the pathway through which RNAs are localized during oogenesis may be an important factor in biasing their distribution into specific cell lineages. The presence of Xwnt11 in the germ cell lineage suggests that a wnt-directed signaling pathway may be involved in germ cell specification. differentiation or migration.

Id gene activity during Xenopus embryogenesis.

The activity of bHLH transcription factors that are involved in cell determination and differentiation is inhibited by Ids, HLH proteins lacking the basic amino acid sequence element. In order to determine the role of Id during development, we have isolated and characterized the Id genes expressed in Xenopus embryos. Three cDNAs were characterized: XIdIa and XIdIb, which are transcribed from one gene but differentially spliced in the 3' untranslated part, and XIdII which is transcribed from a second copy of the gene. One of the two forms of the differentially spliced mRNAs exhibits, 30 nucleotides upstream from the AATAAA site, a sequence box homologous to the cytoplasmic polyadenylation element (CPE) which is present also in Id2 and Id3 mRNAs from higher vertebrates. This raises the question of whether this CPE-like element may link Id mRNA polyadenylation and translation to the cell cycle metabolism. The Xenopus Id gene is transcribed at low level in oocytes and at high level in embryos, after midblastula transition, in a large number of tissues, including the notochord, neural tube, eye, ear, neural crest cells, presomitic mesoderm, myotomes, tailbud and dorsal fin. In myotomes, expression is high in the areas of proliferating myoblasts and decreases as terminal differentiation proceeds, consistent with a function in cell determination and differentiation and possibly also in cell cycle regulation.

Extrachromosomal DNA and its activity in RNA synthesis in oogonia and oocytes in the pupal ovary of Creophilus maxillosus (Staphylinidae, Coleoptera-Polyphaga).

The participation of extrachromosomal DNA (extra DNA) in RNA synthesis in the nuclei of terminal oogonial cells and oocytes in the pupal ovary of Creophilus maxillosus (Staphylinidae, Coleoptera) was examined by autoradiography. It was found that extra DNA in the nuclei of terminal oogonial cells, although predominantly in a condensed and heterochromatic state, produces numerous nucleoli and incorporates 3H-uridine during the interphases between successive differential divisions. Moreover, it was shown that extra DNA is active in RNA synthesis at the same stage of pupal development in which it is synthesized and accumulated, i.e. in the nuclei of terminal oogonial cells. As soon as the oocyte forms RNA synthesis ceases in the extrachromosomal DNA body cells showed that nucleolar material does not disappear during division but remains, at least partly, connected with the extra DNA body.

Membrane-bound or soluble truncated RT1.Aa rat class I major histocompatibility antigens induce specific alloimmunity.

Transfectants that express membrane-bound (MB) or secrete soluble truncated (TR) rat class I RT1.Aa major histocompatibility (MHC) antigens induce alloimmunity in vivo. The MB-RT1.Aa was produced by transfecting the full-length RT1.Aa cDNA, including the alpha 1, alpha 2, and alpha 3, transmembrane and intracellular domains. The TR-RT1.Aa cDNA insert included only the extracellular alpha 1, alpha 2, and alpha 3 domains; a stop codon was placed in front of the transmembrane domain. Following full-length sequencing, MB-RT1.Aa and TR-RT1.Aa cDNAs were translated in vitro into glycosylated MB-RT1.Aa (45 kDa) and TR-RT1.Aa (36 kDa) proteins, respectively. Each cDNA construct was individually subcloned into the pSG5 vector before transfection into Buffalo (BUF; RT1b) hepatoma cells. FACscan analysis with anti-RT1.Aa-specific R2/15S monoclonal antibody (MAb) confirmed surface expression of RT1.Aa molecules on the MB-RT1.Aa, but not on the TR-RT1.Aa, transfectants. In contrast, enzyme-linked immunoadsorbent assays documented the presence of soluble RT1.Aa molecules in supernates from cells transfected with the TR-RT1.Aa, but not from cells transfected with the MB-RT1.Aa, cDNA. Subcutaneous injection of MB-RT1.Aa or TR-RT1.Aa transfectants to BUF or Wistar Furth (WF; RT1u) rats induced accelerated rejection of ACI (RT1a) but not third-party Brown Norway (RT1n) heart allografts. Furthermore, supernates of TR-RT1.Aa, but not of MB-RT1.Aa, transfectants immunized WF hosts toward ACI hearts. Thus, both intact MB-RT1.Aa and soluble TR-RT1.Aa class I alloantigens induce potent sensitization against alloantigens.

Ribosomal gene amplification in the oocytes of Creophilus maxillosus (Staphylinidae, Coleoptera-polyphaga)--an insect with telotrophic ovaries.

We have shown that the extrachromosomal DNA body present in the oocyte nucleus of Creophilus maxillosus contains amplified copies of ribosomal DNA and that multiple nucleoli associated with extrachromosomal DNA body contain ribosomal RNA. In addition, we assessed the level of rDNA amplification in Creophilus oocytes. The amount of DNA in single extra DNA body corresponds to 86% of total nuclear DNA content or 617% of chromosomal DNA content. In Creophilus, there are two phases of transcriptional activity of extrachromosomal rDNA, first in oogonia, and second in growing oocytes, separated by a short period of transcriptional quiescence in the ovary of newly hatched imago. We have described the organization and activity of extra DNA body in RNA synthesis and production of multiple nucleoli in previtellogenic stages of oocyte growth in imaginal ovariole of Creophilus.

Extrachromosomal rDNA and polarity of pro-oocytes during ovary development in Creophilus maxillosus (Coleoptera, Staphylinidae).

In telotrophic ovary of Creophilus maxillosus, the differentiation of the oocyte and nurse cells takes place within the linear clusters of sister oogonial cells. The amplification of rDNA occurs in the nuclei of pro-oocytes which are the most posterior cells of the clusters. During the consecutive oogonial divisions extrachromosomal rDNA segregates preferentially to the pro-oocyte of the next generation. We analyzed the ultrastructure of pro-oocytes and pro-nurse cells in the early and late phase of rDNA amplification in pupal ovary of Creophilus maxillosus. We found that pro-oocytes of the same generation contain variable amounts of extrachromosomal rDNA and that the presence of extra DNA is not limited to the nuclei of pro-oocytes; extra DNA is also present in the nuclei of some pro-nurse cells. Pro-oocytes can experience partial loss of extrachromosomal DNA during early oogonial divisions which is caused by the imprecise segregation of this material to the posterior pole. We believe that this imperfect segregation is a source of extrachromosomal DNA present in the nuclei of pro-nurse cells. Ultrastructural analysis showed that multiple nucleoli do not disperse in oogonial mitoses but remain associated with extrachromosomal chromatin and segregate with it to the posterior pole of the pro-oocyte. We also analyzed the ultrastructure of the germ plasm--a cytoplasmic structure present at the posterior pole of pro-oocytes. We have found that this structure contains spectrin and at the ultrastructural level is strikingly similar to the spectrosome which is present in germline cells of Drosophila. We also found spectrin in the intercellular bridges which connect oogonial cells and are known to contain fusomes.