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Multiple complexes of 20S proteasomes with accessory factors play an essential role in proteolysis in eukaryotic cells. In this report, several forms of 20S proteasomes from extracts of Spodoptera frugiperda (Sf9) cells were separated using electrophoresis in a native polyacrylamide gel and examined for proteolytic activity in the gel and by Western blotting. Distinct proteasome bands isolated from the gel were subjected to liquid chromatography-tandem mass spectrometry and identified as free core particles (CP) and complexes of CP with one or two dimers of assembly chaperones PAC1-PAC2 and activators PA28γ or PA200. In contrast to the activators PA28γ and PA200 that regulate the access of protein substrates to the internal proteolytic chamber of CP in an ATP-independent manner, the 19S regulatory particle (RP) in 26S proteasomes performs stepwise substrate unfolding and opens the chamber gate in an ATP-dependent manner. Electron microscopic analysis suggested that spontaneous dissociation of RP in isolated 26S proteasomes leaves CPs with different gate sizes related presumably to different stages in the gate opening. The primary structure of 20S proteasome subunits in Sf9 cells was determined by a search of databases and by sequencing. The protein sequences were confirmed by mass spectrometry and verified by 2D gel electrophoresis. The relative rates of sequence divergence in the evolution of 20S proteasome subunits, the assembly chaperones and activators were determined by using bioinformatics. The data confirmed the conservation of regular CP subunits and PA28γ, a more accelerated evolution of PAC2 and PA200, and especially high divergence rates of PAC1. • 20S particle forms a core in multiple proteasome complexes in Spodoptera cells. • Complexes with chaperones PAC1, PAC2 and activators PA28γ, PA200 are isolated. • Detachment of RP in 26S proteasomes leaves core particles with different gate sizes. • Evolution rates of 20S proteasome subunits, chaperones and activators are different. [ABSTRACT FROM AUTHOR]

The protein VCP/p97 (also named CDC48 and TER94) belongs to a type II subfamily of the AAA+ATPases and controls cellular proteostasis by acting upstream of proteasomes in the ubiquitin-proteasome protein degradation pathway. The function of VCP/p97 in the baculovirus infection cycle in insect cells remains unknown. Here, we identified VCP/p97 in the fall armyworm Spodoptera frugiperda (Sf9) cells and analyzed the replication of the Autographa californica multiple nucleopolyhedrovirus, AcMNPV, in Sf9 cells in which the VCP/p97 function was inhibited. The specific allosteric inhibitor of the VCP/p97 ATPase activity, NMS-873, did not deplete VCP/p97 in infected cells but caused a dose-dependent inhibition of viral DNA synthesis and efficiently suppressed expression of viral proteins and production of budded virions. NMS-873 caused accumulation of ubiquitinated proteins in a manner similar to the inhibitor of proteasome activity, Bortezomib. This suggests the essential function of VCP/p97 in the baculovirus infection cycle might be associated, at least in part, with the ubiquitin-proteasome system. [ABSTRACT FROM AUTHOR]

Spasticity is a symptom occurring in many neurological conditions including stroke, multiple sclerosis, hypoxic brain damage, traumatic brain injury, tumours and heredodegenerative diseases. It affects large numbers of patients and may cause major disability. So far, spasticity has merely been described as part of the upper motor neurone syndrome or defined in a narrowed neurophysiological sense. This consensus organised by IAB—Interdisciplinary Working Group Movement Disorders wants to provide a brief and practical new definition of spasticity—for the first time—based on its various forms of muscle hyperactivity as described in the current movement disorders terminology. We propose the following new definition system: Spasticity describes involuntary muscle hyperactivity in the presence of central paresis. The involuntary muscle hyperactivity can consist of various forms of muscle hyperactivity: spasticity sensu strictu describes involuntary muscle hyperactivity triggered by rapid passive joint movements, rigidity involuntary muscle hyperactivity triggered by slow passive joint movements, dystonia spontaneous involuntary muscle hyperactivity and spasms complex involuntary movements usually triggered by sensory or acoustic stimuli. Spasticity can be described by a documentation system grouped along clinical picture (axis 1), aetiology (axis 2), localisation (axis 3) and additional central nervous system deficits (axis 4). Our new definition allows distinction of spasticity components accessible to BT therapy and those inaccessible. The documentation sheet presented provides essential information for planning of BT therapy. [ABSTRACT FROM AUTHOR]

Botulinum toxin (BT) therapy is an established treatment of spasticity due to stroke. For multiple sclerosis (MS) spasticity this is not the case. IAB-Interdisciplinary Working Group for Movement Disorders formed a task force to explore the use of BT therapy for treatment of MS spasticity. A formalised PubMed literature search produced 55 publications (3 randomised controlled trials, 3 interventional studies, 11 observational studies, 2 case studies, 35 reviews, 1 guideline) all unanimously favouring the use of BT therapy for MS spasticity. There is no reason to believe that BT should be less effective and safe in MS spasticity than it is in stroke spasticity. Recommendations include an update of the current prevalence of MS spasticity and its clinical features according to classifications used in movement disorders. Immunological data on MS patients already treated should be analysed with respect to frequencies of MS relapses and BT antibody formation. Registration authorities should expand registration of BT therapy for spasticity regardless of its aetiology. MS specialists should consider BT therapy for symptomatic treatment of spasticity. [ABSTRACT FROM AUTHOR]

Baculoviruses are large DNA viruses that infect insect species such as Lepidoptera and are used in biotechnology for protein production and in agriculture as insecticides against crop pests. Baculoviruses require activity of host proteasomes for efficient reproduction, but how they control the cellular proteome and interact with the ubiquitin proteasome system (UPS) of infected cells remains unknown. In this report, we analyzed possible changes in the subunit composition of 26S proteasomes of the fall armyworm, Spodoptera frugiperda (Sf9), cells in the course of infection with the Autographa californica multiple nucleopolyhedrovirus (AcMNPV). 26S proteasomes were purified from Sf9 cells by an immune affinity method and subjected to 2D gel electrophoresis followed by MALDI-TOF mass spectrometry and Mascot search in bioinformatics databases. A total of 34 homologues of 26S proteasome subunits of eukaryotic species were identified including 14 subunits of the 20S core particle (7 α and 7 β subunits) and 20 subunits of the 19S regulatory particle (RP). The RP contained homologues of 11 of RPN-type and 6 of RPT-type subunits, 2 deubiquitinating enzymes (UCH-14/UBP6 and UCH-L5/UCH37), and thioredoxin. Similar 2D-gel maps of 26S proteasomes purified from uninfected and AcMNPV-infected cells at 48 hpi confirmed the structural integrity of the 26S proteasome in insect cells during baculovirus infection. However, subtle changes in minor forms of some proteasome subunits were detected. A portion of the α5(zeta) cellular pool that presumably was not associated with the proteasome underwent partial proteolysis at a late stage in infection. [ABSTRACT FROM AUTHOR]

Treatment of dystonias is generally symptomatic. To produce sufficient therapy effects, therefore, frequently a multimodal and interdisciplinary therapeutic approach becomes necessary, combining botulinum toxin therapy, deep brain stimulation, oral antidystonic drugs, adjuvant drugs and rehabilitation therapy including physiotherapy, occupational therapy, re-training, speech therapy, psychotherapy and sociotherapy. This review presents the recommendations of the IAB-Interdisciplinary Working Group for Movement Disorders Special Task Force on Interdisciplinary Treatment of Dystonia. It reviews the different therapeutic modalities and outlines a strategy to adapt them to the dystonia localisation and severity of the individual patient. Hints to emerging and future therapies will be given. [ABSTRACT FROM AUTHOR]

Abstract: Baculovirus AcMNPV causes proteotoxicity in Sf9 cells as revealed by accumulation of ubiquitinated proteins and aggresomes in the course of infection. Inhibition of proteasomes by lactacystin increased markedly the stock of ubiquitinated proteins indicating a primary role of proteasomes in detoxication. The proteasomes were present in Sf9 cells as 26S and 20S complexes whose protease activity did not change during infection. Proteasome inhibition caused a delay in the initiation of viral DNA replication suggesting an important role of proteasomes at early stages in infection. However, lactacystin did not affect ongoing replication indicating that active proteasomes are not required for genome amplification. At late stages in infection (24–48hpi), aggresomes containing the ubiquitinated proteins and HSP/HSC70s showed gradual fusion with the vacuole-like structures identified as lysosomes by antibody to cathepsin D. This result suggests that lysosomes may assist in protection against proteotoxicity caused by baculoviruses absorbing the ubiquitinated proteins. [Copyright &y& Elsevier]

Abstract: Here we examined the mechanism of propagation of variation potential (VP) induced by burning in wheat leaves. Participation of hydraulic and chemical mechanisms in VP transmission was analyzed by optical coherent tomography and a radioactive tracer method, respectively. The speed of the hydraulic signal considerably exceeded the VP velocity. Investigation of a chemical substance spreading from the zone of local wounding was based on experimental data for radioactive marker transmission derived with a one-dimensional diffusion equation. The speed of the marker transmission was in accordance with VP velocity. The elimination of the potential transmission of a chemical signal by a timed severing of the leaf between the burn site and the recorded site blocked VP propagation. We suggest that a VP is formed by the transmission of a wound substance, the velocity of which is likely increased by hydraulic wave propagation. [Copyright &y& Elsevier]

Abstract: Eight members of the HSP/HSC70 family were identified in Spodoptera frugiperda Sf9 cells infected with Autographa californica multiple nucleopolyhedrovirus (AcMNPV) by 2D electrophoresis followed by mass spectrometry (MALDI/TOF) and a Mascot search. The family includes five HSP70s induced by AcMNPV-infection and three constitutive cognate HSC70s that remained abundant in infected cells. Confocal microscopy revealed dynamic changes in subcellular localization of HSP/HSC70s in the course of infection. At the early stages (4 to 10 hpi), a fraction of HSPs is localized in distinct speckles in cytoplasm. The speckles contained ubiquitinylated proteins suggesting that they may be aggresomes where proteins targeted by ubiquitin are sequestered or processed for proteolysis. S. frugiperda HSP90 was identified in the 2D gels by Western blotting. Its amount was unchanged during infection. A selective inhibitor of HSP90, 17-AAG, decreased the rate of viral DNA synthesis in infected cells suggesting a supportive role of HSP90 in virus replication. [Copyright &y& Elsevier]

Abstract: Background: Cytology-based screening has significantly decreased incidence of and mortality from cervical cancer. High sensitivity of high-risk human papillomavirus (hrHPV) test for the detection of high-grade cervical intraepithelial neoplasia (CIN) makes it a useful screening tool, especially in settings where cytology is not available or not quality assured. The study aimed to determine the prevalence of hrHPV and associated cervical lesions in Russian women over 30 years of age and to assess usefulness of HPV test for cervical screening in Russia. Methods: Consecutive women aged 30–65 years (n =823) receiving routine gynaecological care, not pregnant and not treated for high-grade CIN, were recruited. Oncogenic HPV types were detected using Hybrid Capture 2 (HC2) assay and genotyped with direct DNA sequencing. Women with cytological abnormalities higher than atypical squamous cells of undetermined significance (ASCUS) and women positive for hrHPV were referred to colposcopy. Results: HPV infection was present in 107 (13%) women. Cytological abnormalities were found in 81 (9.8%) patients, including 59 (7.2%) with ASCUS, 21 (2.5%) with low-grade squamous intraepithelial lesions (LSIL), and one (0.1%) with high-grade SIL. Only 11 (18.6%) patients with ASCUS were positive for hrHPV. Histological diagnoses were obtained for 63 women. Relative sensitivities and positive predictive values of the HPV test and cytology for the detection of high-grade CIN were 100% (6/6) and 9.5% (6/63), and 83.3% (5/6) and 27.8% (5/18), respectively. The most prevalent hrHPV types were 16 (3.9%), 31 (2.8%), 52 (1.7%), and 33 (1.3%). Cytological abnormalities and symptoms of a urogenital infection were strongly associated with hrHPV infection. Conclusions: The study provides data on the prevalence of hrHPV types in association with cervical lesions, as well as on hrHPV determinants, in an unvaccinated population of Russian women. Our results indicate that HPV test might be a useful screening tool in Russia. [Copyright &y& Elsevier]

Abstract: The aim of this study was to evaluate the efficacy and safety of intramuscular (IM) administration of botulinum toxin type A (Dysport®, Ipsen Biopharm Ltd.) for the treatment of cervical dystonia (CD) and the long-term safety and efficacy of repeated treatments. During the randomized, double-blind, placebo-controlled phase patients were randomized to 500 units Dysport (n = 55) or placebo (n = 61). Efficacy assessments included the Toronto Western Spasmodic Torticollis Rating Scale (TWSTRS) total and subscale scores, visual analogue scale (VAS) for pain, subject/investigator’s VAS for symptom assessments. Patients completing the double-blind treatment could enter an open-label extension phase and receive up to 4 additional Dysport treatments. Dysport produced a significant decrease from baseline in mean (±SE) TWSTRS total scores compared with placebo at Week 4 (primary efficacy endpoint; −15.6 ± 2.0 vs. −6.7 ± 2.0; p < 0.001) with significant improvements sustained to Week 12 (p = 0.019). Dysport also produced significant improvements in TWSTRS subscale scores, VAS pain scores, and subject/investigator’s VAS symptom assessments compared to placebo. The mean duration of open-label study participation was 51.9 weeks (range 3.9–94.0 weeks). During open-label treatment, all treatment cycles resulted in improvements in mean TWSTRS total and subscale scores at Week 4 post-treatment; greatest improvement was seen in cycle 1. The mean duration between treatment cycles was 15–17 weeks. Dysport demonstrated a good long-term safety profile; most adverse events were mild or moderate and typical of the known safety profile of Dysport in this indication. These results confirm that Dysport (500 units) is safe, effective, and well-tolerated in patients with CD. [Copyright &y& Elsevier]

Bemethyl is an actoprotector, an antihypoxant, and a moderate psychostimulant. Even though the therapeutic effectiveness of bemethyl is well documented, there is a gap in knowledge regarding its metabolic products and their quantitative and qualitative characteristics. Since 2018, bemethyl is included to the Monitoring Program of the World Anti-Doping Agency, which highlights the challenge of identifying its urinary metabolites. The objective of the study was to investigate the biotransformation pathways of bemethyl using a combination of liquid chromatography-high-resolution mass spectrometry and in silico studies. Metabolites were analyzed in a 24 h rat urine collected after oral administration of bemethyl at a single dose of 330 mg/kg. The urine samples were prepared for analysis by a procedure developed in the present work and analyzed by high performance liquid chromatography–tandem mass spectrometry. For the first time, nine metabolites of bemethyl with six molecular formulas were identified in rat urine. The most abundant metabolite was a benzimidazole–acetylcysteine conjugate; this biotransformation pathway is associated with the detoxification of xenobiotics. The BioTransformer and GLORY computational tools were used to predict bemethyl metabolites in silico. The molecular docking of bemethyl and its derivatives to the binding site of glutathione S-transferase has revealed the mechanism of bemethyl conjugation with glutathione. The findings will help to understand the pharmacokinetics and pharmacodynamics of actoprotectors and to improve antihypoxant and adaptogenic therapy. [ABSTRACT FROM AUTHOR]

Probiotic bacteria protect against gastroenteritis, promote toxin reduction, cholesterol reduction, improved digestion and good function, mineral bioavailability enhancement. Bacterial strains, including Lactobacillus and Bifidobacterium, are generally used in probiotic food production. However, they are weakly resistant to the stomach's aggressive environment and high temperature, which can result in significant loss of viable probiotic microorganisms. Application of spore-forming bacteria Bacillus coagulans MTCC 5856 allows developing an innovative natural dairy fermented product with high probiotic microorganism viability (p < 0.05) during the technological process, storage period, at different pH values, and in the presence of bile salts when examining in vitro. Due to the low caseolytic activity of Bacillus coagulans , the proper ratio and fermentation temperature with the use of auxiliary starter culture Streptococcus thermophilus were established. The addition of milk protein improves the rheological characteristics of the product. Bacillus coagulans and Streptococcus thermophilus maintained more than 8 Log 10 CFU/g of viable microorganisms during the product storage within 60 days. The use of Bacillus coagulans allows increasing the fermented product's antioxidant activity versus the antioxidant activity of the product when Streptococcus thermophilus is used alone. The results showed that the fermented dairy product itself to be an excellent carrier of Bacillus coagulans. • The method for applying B. coagulans in fermented dairy products is developed for the first time. • B. coagulans coupled with S. thermophilus produce a dairy product of improved quality. • Milk protein at 2% (w/v) improves the rheology, biological value, and taste of the product. • The product has a high antioxidant activity enhancing its therapeutic value. • Surprisingly, the storage period of the dairy beverage is determined as 60 days at 4 ± 2 °C. [ABSTRACT FROM AUTHOR]

• The sonication results in changing the rheological properties of the dough. • Sonication leads to a 2-fold increase in the dough volume. • The proposed technology has an additional beneficial economic effect. • The ultrasound-assisted technology provides sterilization of final product. Nowadays celiac disease is becoming more common. It is the autonomic genetic disease that is accompanied by damage to the intestines due to a reaction to eating some proteins. People who are suffering from celiac disease cannot eat food containing gluten, including dough made from gluten-containing seeds. But the gluten-free dough has commonly bad rheological properties and cannot be used for automatic molding the dumplings. In this article, we propose the ultrasonic-assisted technology to fabricate the gluten-free dough with improved rheological properties acceptable for automatic molding of the dumplings. Application of ultrasonic treatment at a frequency of 35 kHz during the dough preparation leads to the homogenization of the dough structure and changing the rheological properties of the dough. The ultrasound induces mechanical, physical and chemical/biochemical changes of the dough components through cavitation. The sonication causes a doubled dough volume increase followed by an additional mass yield of the dumplings equal 2–10% per kilogram of dough. Besides extra beneficial economic effect, our technology provides an additional sterilization effect of the fabricated dough. [ABSTRACT FROM AUTHOR]