Generation of mouse lines with conditionally or constitutively inactivated Snca gene and Rosa26-stop-lacZ reporter located in cis on the mouse chromosome 6

α-Synuclein is involved in many important molecular processes in neuronal cells and their synapses, and its malfunction has been linked to the development of Parkinson’s and certain other neurodegenerative diseases. Animal models allowing tightly monitored conditional inactivation of the encoding gene, Snca, are indispensible for studies aimed at understanding normal function of α-synuclein in various neuronal populations and its role in pathogenesis of neurodegenerative diseases. We have recently reported the production of several novel mouse lines for manipulating expression of the endogenous Snca gene, including a line for Cre-recombinase-driven conditional inactivation of the gene (mice with floxed Snca) and a new line with a constitutive knockout of α-synuclein. Rosa26-stop-lacZ reporter cassette is commonly used for monitoring efficiency of Cre-recombination but in mouse genome Snca and Rosa26 loci are located on the same chromosome. Here we describe production of lines with a modified Snca locus, either floxed or constitutively inactivated and the Rosa26-stop-lacZ reporter cassette located in cis on the mouse chromosome 6. These new mouse lines are invaluable for fast identification of cells with inactivation of Snca by Cre-recombination and represent useful tools for in vivo studies of α-synuclein function and dysfunctio

Mullite production: phase transformations of kaolinite, thermodynamics of the process

The growing demand for mullite raw materials, which meet industrial requirements originates the search for new and alternative sources, as well as efficient technologies for obtaining the target products (nanocomposites). The article suggests a method for obtaining mullite from kaolinite experimentally (Vezhayu-Vorykvinsky deposit, Russia). Structural kaolinite transformations (Al-Si-O-Me system), mineral phases transformations, and thermodynamics of the process have been studied. Based on the estimation of the thermodynamics of the reactions, the preferable reaction of mullite formation was determined. The article shows, that formation of the target product, mullite nanocomposite, has several intermediate phases (metakaolinite, pseudomullite). The transformations of the initial kaolinite structure include the removal of structural water and separation of the silica-oxygen tetrahedral and alumina-oxygen octahedral layers, the decomposition into free oxides, breaking of bonds between the silica-oxygen tetrahedrons and the partial increase in the coordination number of aluminium ions, the formation of mullite and cristobalite from free oxides. The proposed approach controls the ratio of Al2O3 and SiO2 phases at certain stages, which will further improve the mechanical and other properties of the matrix of the obtained raw materials for the target prototypes of industrial products

A novel resource for studying function and dysfunction of α-synuclein: mouse lines for modulation of endogenous Snca gene expression

Pathological modification of α-synuclein is believed to be an important event in pathogenesis of Parkinson’s disease and several other neurodegenerative diseases. In normal cells this protein has been linked to many intracellular processes and pathways. However, neither normal function of α-synuclein in neuronal and certain other types of cells nor its exact role in the disease pathogenesis is well understood, which is largely due to limitations of animal models used for studying this protein. We produced and validated several novel mouse lines for manipulating expression of the endogenous Snca gene coding for α-synuclein. These include a line for conditional Cre-recombinase-driven inactivation of the gene; a line for conditional Flp-driven restoration of a neo-cassete-blocked α-synuclein expression; a new line with a “clean” constituent knockout of the gene as well as a line carrying this knockout locus and Rosa26-stop-lacZ reporter locus linked at the same mouse chromosome 6. Altogether these lines represent a set of new useful tools for studies of α-synuclein normal function and the role of this protein in disease pathogenesis

Efficacy of targeted screening for familial hypercholesterolemia in children and adults aged <44 years: retrospective, prospective observational study

BACKGROUND: Considering the high cardiovascular risk in patients with familial hypercholesterolemia (FH), early diagnosis and treatment are the basis for the prevention of vascular accidents. In the absence of universal screening, the results of more targeted diagnostics are of interest — the search for lipid metabolism disorders in young relatives of patients with atherosclerotic vascular diseases. OBJECTIVE: To determine the effectiveness of targeted screening for identifying patients with FH, which will justify its widespread use in clinical practice. MATERIALS AND METHODS: At the start of the observational study, 584 patients were identified from the vascular centers of the Krasnoyarsk Regional Hospital, Krasnoyarsk Interdistrict Clinical Hospital No. 20 named after. I.S. Berzon, who had lipid profile data and early cardiovascular events (all forms of coronary heart diseases), including reconstructive vascular interventions up to 55 years in men and up to 60 years in women in the case of first-degree relatives and up to 50 years in second-degree relatives. The study examined patients’ relatives aged 44 years. To diagnose FH in participants aged 16 years, the scale developed by a network of Dutch lipid clinics was used; in participants aged 16 years, the S. Broome criteria were used. Targeted screening and lipid parameters in individuals with probable, possible, and definite FH were analyzed. Data processing was performed using Microsoft Excel and Statistica v. 12.0 (USA). Parametric (calculation of the median and 25th and 75th quartiles) and nonparametric (statistical significance of differences was determined using the Mann–Whitney and χ2 tests) methods were used. RESULTS: The study examined 70 children and 104 adults aged 44 years whose first- and/or second-degree relatives suffered premature vascular events. In the general group of participants, 42 (24.14%) patients had FH (of which 18 were 17 years old) and 37 (21.2%) had hyperlipoproteinemia (a). Definite FH was documented in 16 (9.1%) patients, probable in another 16, and possible in 9. Hyperlipoproteinemia (a) was found in 37 (21.2% of general group) patients, including 12 children (17.1% of pediatric group). CONCLUSION: Not more than 30% of the population are aware of the hereditary nature of lipid metabolism disorders and atherosclerotic diseases. Targeted screening is the least expensive and most effective tool for identifying patients with FH and hyperdipoproteinemia (a)

Low level of expression of C-terminally truncated human FUS causes extensive changes in the spinal cord transcriptome of asymptomatic transgenic mice

A number of mutations in a gene encoding RNA-binding protein FUS have been linked to the development of a familial form of amyotrophic lateral sclerosis known as FUS-ALS. C-terminal truncations of FUS by either nonsense or frameshift mutations lead to the development of FUS-ALS with a particularly early onset and fast progression. However, even in patients bearing these highly pathogenic mutations the function of motor neurons is not noticeably compromised for at least a couple of decades, suggesting that until cytoplasmic levels of FUS lacking its C-terminal nuclear localisation signal reaches a critical threshold, motor neurons are able to tolerate its permanent production.In order to identify how the nervous system responds to low levels of pathogenic variants of FUS we produced and characterised a mouse line, L-FUS[1-359], with a low neuronal expression level of a highly aggregation-prone and pathogenic form of C-terminally truncated FUS. In contrast to mice that express substantially higher level of the same FUS variant and develop severe early onset motor neuron pathology, L-FUS[1-359] mice do not develop any clinical or histopathological signs of motor neuron deficiency even at old age. Nevertheless, we detected substantial changes in the spinal cord transcriptome of these mice compared to their wild type littermates. We suggest that at least some of these changes reflect activation of cellular mechanisms compensating for the potentially damaging effect of pathogenic FUS production. Further studies of these mechanism might reveal effective targets for therapy of FUS-ALS and possibly, other forms of ALS

Dimebon Does Not Ameliorate Pathological Changes Caused by Expression of Truncated (1–120) Human Alpha-Synuclein in Dopaminergic Neurons of Transgenic Mice

Background: Recent clinical studies have demonstrated that dimebon, a drug originally designed and used as a non-selective antihistamine, ameliorates symptoms and delays progress of mild to moderate forms of Alzheimer’s and Huntington’s diseases. Although the mechanism of dimebon action on pathological processes in degenerating brain is elusive, results of studies carried out in cell cultures and animal models suggested that this drug might affect the process of pathological accumulation and aggregation of various proteins involved in the pathogenesis of proteinopathies. However, the effect of this drug on the pathology caused by overexpression and aggregation of alpha-synuclein, including Parkinson’s disease (PD), has not been assessed. Objective: To test if dimebon affected alpha-synuclein-induced pathology using a transgenic animal model. Methods: We studied the effects of chronic dimebon treatment on transgenic mice expressing the C-terminally truncated (1–120) form of human alpha-synuclein in dopaminergic neurons, a mouse model that recapitulates several biochemical, histopathological and behavioral characteristics of the early stage of PD. Results: Dimebon did not improve balance and coordination of aging transgenic animals or increase the level of striatal dopamine, nor did it prevent accumulation of alpha-synuclein in cell bodies of dopaminergic neurons. Conclusion: Our observations suggest that in the studied model of alpha-synucleinopathy dimebon has very limited effect on certain pathological alterations typical of PD and related diseases

C9ORF72 hexanucleotide repeat expansion in ALS patients from the Central European Russia population

Cohorts of amyotrophic lateral sclerosis (ALS) patients and control individuals of Caucasian origin from the Central European Russia (Moscow city and region) were analyzed for the presence of hexanucleotide repeat GGGGCC expansion within the first intron of the C9ORF72 gene. The presence of a large (>40) repeat expansion was found in 15% of familial ALS cases (3 of 20 unrelated familial cases) and 2.5% of sporadic ALS cases (6 of 238) but in none of control cases. These results suggest that the frequency of C9ORF72 hexanucleotide repeats expansions in the Central Europea

Dynamics of Dopamine and Other Monoamines Content in Rat Brain after Single Low-Dose Carbon Nuclei Irradiation

Space radiation, presented primarily by high-charge and -energy particles (HZEs), has a substantial impact on the central nervous system (CNS) of astronauts. This impact, surprisingly, has not only negative but also positive effects on CNS functions. Despite the fact that the mechanisms of this effect have not yet been elucidated, several studies indicate a key role for monoaminergic networks underlying these effects. Here, we investigated the effects of acute irradiation with 450 MeV/n carbon (12C) nuclei at a dose of 0.14 Gy on Wistar rats; a state of anxiety was accessed using a light–dark box, spatial memory in a Morris water maze, and the dynamics of monoamine metabolism in several brain morphological structures using HPLC. No behavioral changes were observed. Irradiation led to the immediate suppression of dopamine turnover in the prefrontal cortex, hypothalamus, and striatum, while a decrease in the level of norepinephrine was detected in the amygdala. However, these effects were transient. The deferred effect of dopamine turnover increase was found in the hippocampus. These data underscore the ability of even low-dose 12C irradiation to affect monoaminergic networks. However, this impact is transient and is not accompanied by behavioral alterations

iii EXPRESSION AND FUNCTION OF THE SMALL HEAT SHOCK PROTEIN HSP27

The members of the Committee appointed to examine the thesis of ALEXEY USTYUGOV find it satisfactory and recommend that it be accepted

Hsp27 is persistently expressed in zebrafish skeletal and cardiac muscle tissues but dispensable for their morphogenesis

Constitutive expression of Hsp27 has been demonstrated in vertebrate embryos, especially in developing skeletal and cardiac muscle. Results of several previous studies have indicated that Hsp27 could play a role in the development of these tissues. For example, inhibition of Hsp27 expression has been reported to cause defective development of mammalian myoblasts in vitro and frog embryos in vivo. In contrast, transgenic mice lacking Hsp27 develop normally. Here, we examined the distribution of Hsp27 protein in developing and adult zebrafish and effects of suppressing Hsp27 expression using phosphorodiamidate morpholino oligonucleotides (PMO) on zebrafish development. Consistent with our previous analysis of hsp27 messenger RNA expression, we detected the protein Hsp27 in cardiac, smooth, and skeletal muscle of both embryonic and adult zebrafish. However, embryos lacking detectable Hsp27 after injection of antisense hsp27 PMO exhibited comparable heart beat rates to that of control embryos and cardiac morphology was indistinguishable in the presence or absence of Hsp27. Loss of Hsp27 also had no effect on the structure of the skeletal muscle myotomes in the developing embryo. Finally, embryos injected with antisense hsp27 and scrambled control PMO displayed equal motility. We conclude that Hsp27 is dispensable for zebrafish morphogenesis but could play a role in long-term maintenance of heart and muscle tissues