The HSPB1-p62/SQSTM1 functional complex regulates the unconventional secretion and transcellular spreading of the HD-associated mutant huntingtin protein.

Conformational diseases, such as Alzheimer, Parkinson and Huntington diseases, are part of a common class of neurological disorders characterized by the aggregation and progressive accumulation of proteins bearing aberrant conformations. Huntington disease (HD) has autosomal dominant inheritance and is caused by mutations leading to an abnormal expansion in the polyglutamine (polyQ) tract of the huntingtin (HTT) protein, leading to the formation of HTT inclusion bodies in neurons of affected patients. Interestingly, recent experimental evidence is challenging the conventional view by which the disease pathogenesis is solely a consequence of the intracellular accumulation of mutant protein aggregates. These studies reveal that transcellular transfer of mutated huntingtin protein is able to seed oligomers involving even the wild-type (WT) forms of the protein. To date, there is still no successful strategy to treat HD. Here, we describe a novel functional role for the HSPB1-p62/SQSTM1 complex, which acts as a cargo loading platform, allowing the unconventional secretion of mutant HTT by extracellular vesicles. HSPB1 interacts preferentially with polyQ-expanded HTT compared with the WT protein and affects its aggregation. Furthermore, HSPB1 levels correlate with the rate of mutant HTT secretion, which is controlled by the activity of the PI3K/AKT/mTOR signalling pathway. Finally, we show that these HTT-containing vesicular structures are biologically active and able to be internalized by recipient cells, therefore providing an additional mechanism to explain the prion-like spreading properties of mutant HTT. These findings might also have implications for the turn-over of other disease-associated, aggregation-prone proteins.

[Reduction in the need for glucocorticoids on the background of therapy with biologic disease-modifying antirheumatic drugs and Janus kinase inhibitors in rheumatoid arthritis: evidence from real clinical practice].

BACKGROUND: Clinical guidelines for the treatment of rheumatoid arthritis (RA) recommend reducing the use of glucocorticoids (GCs) due to the high risk of associated complications. AIM: To determine the frequency of GC cancellations and dose reductions in real clinical practice, while taking into account active RA therapy. MATERIALS AND METHODS: The study group consisted of 303 patients with RA reliable according to ACR/EULAR criteria (women 79.9%, age 52.8±13.3, disease duration 9 [4; 16] years, DAS-28-CRP 4.9±1.0, RF seropositivity 77.4%, ACPA seropositivity 70.3%), who were prescribed or changed therapy with disease-modifying antirheumatic drugs (DMARDs), biologic disease-modifying antirheumatic drugs (bDMARDs) or Janus kinase inhibitors (iJAK) due to disease exacerbation and ineffectiveness of previous treatment. All patients initially received GC (7.7±3.8 mg/day equivalent of prednisolone). After adjustment of therapy, 42.9% of patients received methotrexate, 27.6% leflunomide, 2.5% sulfasalazine, hydroxychloroquine, or a combination with an Non-steroidal anti-inflammatory drugs, 63.7% bDMARDs, and 7.2% iJAK. The need for GC intake was assessed by a telephone survey conducted 6 months after the start of follow-up. RESULTS: Telephone survey was possible in 274 (90.4%) persons. There was a significant decrease in pain intensity (numerical rating scale, NRS 0-10) from 6.3±1.4 to 4.3±2.4 (p<0.001), fatigue (NRS) from 6.7±2.3 to 5.2±2.1 (p<0.001), and functional impairment (NRS) from 5.4±2.1 to 3.9±2.0 (p<0.001). A positive PASS index (symptom status acceptable to patients) was noted in 139 (50.7%) patients. GC cancellation was noted in 19.7%, dose reduction in 25.9%, maintaining the same dose in 42.7%, and dose increase in 11.7%. CONCLUSION: Against the background of intensive RA therapy, including combination of DMARDs with bDMARDs or iJAK, complete withdrawal or reduction of GC dose was achieved in less than half (45.6%) of patients after 6 months.

[Dihydroquercetin influence on clinical and biochemical blood parameters of pigs under conditions of stress load].

In modern society, distress has become a widespread condition that negatively affects the functioning of all systems of the human organism. The study of biological mechanisms and changes in the organism under the influence of stress, as well as methods of their leveling, are relevant in medicine, animal science and veterinary medicine. Pigs are an excellent biological model that is closest to humans. The aim of the research was to study the hematological and biochemical parameters of pigs out of and under stress, including against the background of daily consumption of the flavonoid dihydroquercetin (DHQ) with feed. Material and methods. The research was conducted in the experimental yard of the L.K. Ernst Federal Science Center for Animal Husbandry on 3 groups of pigs [F2 hybrid (large white×Landrace)×Duroc] with an initial body weight of 30-35 kg (n=27). Group 1K consisted of control animals not exposed to stress (n=9); group 2K - control animals subjected to simulated stress by the rearrangement of animals (n=9); group 3O - experimental animals subjected to simulated stress and fed throughout the entire experiment DHQ (32 mg per 1 kg of feed) (n=9). On days 0, 42, and 76, blood was collected from the animals and their hematological and biochemical parameters were studied using conventional methods. Results. The positive effect of using DHQ in pigs' nutrition on enhancing the oxidizing function of blood, metabolic intensity, and increasing the endurance of animals under stress conditions has been manifested in maintaining leukocyte level with a higher content of erythrocytes and hematocrit. In animals fed DHQ, alanine aminotransferase activity was lower than in animals not receiving DHQ. Stress led to a significant increase in lactate dehydrogenase activity in group 2K on the 46th day, which was not observed in animals treated with DHQ. Conclusion. Long-term intake DHQ (up to 72 days inclusive) against the background of stress contributed to the preservation of blood values at the control level (without stress), within the physiological norm.

The incidence of dry chlorhexidine gluconate transfer from skin to surgical gloves: a simulation and in vitro study.

BACKGROUND: To prevent alcohol-based chlorhexidine from reaching the cerebrospinal fluid, it is recommended that the antiseptic solution be allowed to dry before skin palpation or puncture. However, no guidelines specify a drying time interval. Manufacturers recommend 3 min of air drying, based upon the isopropyl alcohol component. Therefore, to fill this knowledge gap, we designed a simulation study to investigate the incidence of primary chlorhexidine transfer from skin to gloves following three drying time intervals. We also investigated the incidence of secondary chlorhexidine transfer from gloves to another surface following one drying time interval. METHODS: An alcohol-based chlorhexidine antiseptic solution with dye, ChloraPrep®, was applied to the skin of the lumbar region of 20 volunteers. Cotton-tipped applicators wrapped in material from gloves were taken from the application area at 3, 4, 5, and 10 min following application. Transfer of chlorhexidine from skin to gloves, and gloves to another medium, was assessed through a chemical assay that produced a color change when chlorhexidine was present on the sample. RESULTS: The incidence of primary chlorhexidine transfer from skin to gloves at 3, 4 and 10 min following application was 99.5%, 99.4%, and 99.6%, respectively. The incidence of secondary chlorhexidine transfer from gloves to another surface was 68.9%. CONCLUSION: Gloves are routinely contaminated with chlorhexidine during central neuraxial blockade. The high incidence of secondary transfer in our simulation suggests a pathway by which chlorhexidine may gain access to the neuraxial space.

Correlative light-electron microscopy reveals the tubular-saccular ultrastructure of carriers operating between Golgi apparatus and plasma membrane.

Transport intermediates (TIs) have a central role in intracellular traffic, and much effort has been directed towards defining their molecular organization. Unfortunately, major uncertainties remain regarding their true structure in living cells. To address this question, we have developed an approach based on the combination of the green fluorescent protein technology and correlative light-electron microscopy, by which it is possible to monitor an individual carrier in vivo and then take a picture of its ultrastructure at any moment of its life-cycle. We have applied this technique to define the structure of TIs operating from the Golgi apparatus to the plasma membrane, whose in vivo dynamics have been characterized recently by light microscopy. We find that these carriers are large (ranging from 0.3-1.7 microm in maximum diameter, nearly half the size of a Golgi cisterna), comprise almost exclusively tubular-saccular structures, and fuse directly with the plasma membrane, sometimes minutes after docking to the fusion site.

TRPML1 links lysosomal calcium to autophagosome biogenesis through the activation of the CaMKKß/VPS34 pathway.

The lysosomal calcium channel TRPML1, whose mutations cause the lysosomal storage disorder (LSD) mucolipidosis type IV (MLIV), contributes to upregulate autophagic genes by inducing the nuclear translocation of the transcription factor EB (TFEB). Here we show that TRPML1 activation also induces autophagic vesicle (AV) biogenesis through the generation of phosphatidylinositol 3-phosphate (PI3P) and the recruitment of essential PI3P-binding proteins to the nascent phagophore in a TFEB-independent manner. Thus, TRPML1 activation of phagophore formation requires the calcium-dependent kinase CaMKKß and AMPK, which increase the activation of ULK1 and VPS34 autophagic protein complexes. Consistently, cells from MLIV patients show a reduced recruitment of PI3P-binding proteins to the phagophore during autophagy induction, suggesting that altered AV biogenesis is part of the pathological features of this disease. Together, we show that TRPML1 is a multistep regulator of autophagy that may be targeted for therapeutic purposes to treat LSDs and other autophagic disorders.

[Fatty acid composition of cells and lipopolysaccliarides of Mannheimia haemolytica, Mannheimia glucosida and Bibersteinia trehalosi strains].

The Mannheimia haemolytica, Mannheimia glucosida and Bibersteinia trehalosi strains and the similar fatty acid composition of cells with domination of C(16:1) and C(16:0), which were in almost equal quantities, C(14:0 and C(18:1) + C(18:2). The fatty acid composition of lipopolysaccharides (LPS) of the studied bacteria had no essential differences too. It was mainly represented by C(14:0) and 3-OH-C(14:0) which consisted of more than 80% of all LPS fatty acids. C(12:0), C(16:1) and C(16:0) were presented in LPS in small quantities. The M. haemolytica, M. glucosida and B. trehalosi strains did not differ essentially by fatty acid compositions of cells and LPS from earlier studied strains of genera Pasteurella (P. multocida), Haemophilus (H. influenzae and other species), Actinobacillus (A. pleuropneumoniae). This shows the close phylogenetic relationship of the mentioned bacteria and significance of investigated signs as chemotaxonomic markers for differentiation of taxons of the above genus level. The paper is presented in Russian.

[Cellular fatty acid composition of the Burkholderia mallei and Burkholderia pseudomallei strains as generic feature of Burkholderia].

The cellular fatty acid compositions of studied two strains of Burkholderia mallei and a strain of Burkholderia pseudomallei are represented by saturated and monounsaturated straight chain fatty acids with 14-18 carbon atoms, cyclopropane fatty acids C(17 inverted delta) and C(19 inverted delta), and hydroxy acids 3-OH-C14:0, 2-OH-C(16:0), and 3-OH-C(16:0). The strain variation of cyclopropane and unsaturated fatty acid levels was observed. The cellular fatty acid spectra of studied bacteria did not depend essentially on growth medium. The levels of cyclopropane fatty acids increased and those of unsaturated ones decreased with culture age, a tendency to increasing the levels of hydroxy fatty acids was observed too.

[Fatty-acid composition of cells and lipopolysaccharides in different Yersinia species under the conditions of growth at low temperature].

Y. pestis, Y. pseudotuberculosis, Y. enterocolitica, Y. frederiksenii, Y. intermedia, Y. kristensenii and Y. ruckeri grown at 4 degrees C were characterized by fatty acid composition with a high content of C16:1 and C18:1, as well as the proportion of saturated to nonsaturated fatty acids equal to, on the average, 2.0. In Yersinia lipopolysaccharides a relatively high level of C16:1 and C12:0 was observed with the prevalence of 3-OH-C14:0. In the fatty-acid spectra of both cells and lipopolysaccharides no essential difference was noted. Thus, during growth at low temperature differences, earlier detected in the studied Yersinia species grown at 37 degrees C and making it possible to divide 7 Yersinia species into 2 groupes, were completely leveled. These results confirmed the close phylogenetic relationship between the Yersinia species under study and were indicative of more pronounced biological community of Yersinia under the conditions of growth at low temperature.

Coalescence of Golgi fragments in microtubule-deprived living cells.

The process of stack coalescence, an important mechanism of Golgi recovery from mitosis, was examined using novel experimental paradigms. In living cells with disrupted (by nocodazole) microtubules, galactosyl transferase-GFP-labelled Golgi fragments constantly appeared, grew, sometimes moved with a speed of 1-2 microns/min, coalesced or gradually diminished and disappeared. The rate of Golgi fragment turnover and coalescence was highly balanced to maintain a constant number of Golgi units per cell. Moreover some Golgi islands appear and some received new GalTase-GFP after photobleaching of cell cytoplasm. Short tubules extending from the rims of scattered Golgi fragments frequently formed bridges between ministacks, inducing their coalescence. The frequency of coalescence could also be inhibited by disruption of actin microfilaments. After the Golgi redistribution into endoplasmic reticulum induced by brefeldin A, either the growth of small Golgi fragments or their coalescence leads to compartmentalized stack formation without the participation of microtubules. These results demonstrate that this coalescence between isolated Golgi stacks is microtubule-independent and could thus be mediated by membranous tubules.

Antimony biomethylation by Scopulariopsis brevicaulis: characterization of intermediates and the methyl donor.

The filamentous fungus Scopulariopsis brevicaulis biomethylates inorganic antimony(III) compounds to trimethylstibine, that can be detected in culture headspace gases. Dimethylantimony and trimethylantimony species have been detected in the medium of these cultures, but the origin of these species was controversial. We now show that the dimethylantimony species is a true intermediate on the pathway to trimethylstibine (rather than arising from trimethylstibine oxidation or as an analytical artifact) because no dimethylantimony species are formed on trimethylstibine oxidation, as determined by using HG-GC-AAS. Furthermore, the dimethylantimony and trimethylantimony species can be separated, by using anion exchange chromatography, and so the dimethylantimony species is not an analytical artifact, formed during the hydride generation process. The antimony biomethylation mechanism was further probed by measuring incorporation of the methyl group, from 13CD3-L-methionine and CD3-D-methionine, into methylantimony species and, for comparison, into methylarsenic species. The percentage incorporation of the labeled methyl group into methylarsenic and methylantimony species was not significantly different. The incorporation from 13CD3-L-methionine was 54% and 47% for antimony and arsenic, respectively. The incorporation from CD3-D-methionine was 20% and 16% for antimony and arsenic, respectively. It appears that the biomethylation of arsenic and antimony occur by very similar, perhaps identical, mechanisms.

Effect of ion adsorption on the electrokinetic properties of erythrocytes.

The microelectrophoresis technique was used to determine the dependence of human erythrocyte surface potential on the concentration of various cations and anions. The interpretation of the results is based on the Gouy--Chapman--Stern theory. Values of pK, characterizing the binding of ions to the external surface of erythrocytes, as well as numbers of binding sites per unit area were determined. The affinities of ions for the red cell membrane were shown to decrease in the sequence: H+ greater than Ca2+ greater than Sr2+ greater than Mg2+ greater than Ba2+ greater than Li+ greater than Na+ congruent to congruent to K+ congruent to NH4+ and trinitrophenol greater than IO4- greater than CIO4- greater than salicylate congruent to I- greater than greater than SCN- greater than H2PO4- greater than Br- greater than Cl- greater than HPO4(2-). Changes in the ionic strength of the medium resulted in changes in numbers of exposed ion-binding sites. This phenomenon is interpreted in terms of ionic strength-dependent structural transformations of the cell surface coat.

[Selective action of argon laser radiation in breast cancer]. / Selektivnoe deistvie izlucheniia argonovogo lazera pri rake molochnoi zhelezy.

The attempts were made to apply the argon laser radiation (the green region of light spectrum) for the treatment of breast carcinoma. The 46 post-operative skin metastases of breast carcinoma in 15 women were irradiated with no photosensitizers used. As a result of the treatment (4 to 50 sessions) the complete resolution of metastatic tumours was achieved in 13 patients. In contrast to the methods of laser coagulation treatment of tumours, used till now, the described approach provides for selective elimination of tumour cells without normal tissue alteration. The involvement of photothermic or photochemical mechanisms in this selective effect is discussed.

[Structural changes in the contractile proteins of muscle fiber studied by polarization ultraviolet fluorescence microscopy. IX. The effect of the pH and ionic strength of the solution on the conformational restructurings of F-actin induced by the binding of heavy meromyosin]. / Izuchenie strukturnykh izmenenii sokratitel'nykh belkov myshechnogo volokna s pomoshch'iu poliarizatsionnoi ul'trafioletovoi fluorestsentnoi mikroskopii. IX. Vliianie pH i ionnoi sily rastvora na konformatsionnye perestroiki F-aktina, indutsirovannye sviazyvaniem tiazhelogo meromiozina.

The dependence of F-actin conformational changes induced by the F-actin-HMM complex on pH and ionic strength was found by polarized ultraviolet fluorescence microscopy. It is discovered that pH affects sufficiently the cooperativity of F-actin structural changes, while the ionic strength affects their depth. The actomyosin complex was supposed to be at least in two structural states, differing in their orientation as well as in flexibility of F-actin monomers.

[Use of tonic contracture of strips of frog myocardium to study sodium-calcium exchange in the sarcolemma of myocardiocytes]. / Ispol'zovanie tonicheskoi kontraktury polosok miokarda liagushki dlia izucheniia natrii-kal'tsievogo obmena v sarkolemme kardiomiotsitov.

The mechanism of tonic component of the frog myocardium potassium contraction was studied. The degree of contraction depended on the potassium, sodium and calcium ions concentrations in extracellular medium. The tonic component of hyperpotassium contraction, as well as the additional contractions due to hyposodium and hypercalcium medium in the tonic component phase seems to stem from activation of sodium--calcium exchange. The latter through the sarcolemma of the heart cells was 3Na: 1Ca. The effect of some mono- and bivalent cations on sodium-calcium exchange in respect ot their inhibiting activity was as follows: Cd+2 greater than Sr+2 greater than Ba+2 greater than Mn+2 much greater than Mg+2 greater than Li+ greater than Cs+.

[Treatment of skin tumors with a pulse laser]. / Lechenie opukholei kozhi impul'snym lazerom.

The clinical laser complex including neodymium and CO2--lasers is described, and the methods of irradiation depending on the type of the laser and tumor are given. The results of the treatment in 559 patients with different malignant and benign skin tumors are analysed. The data of the follow-up (up to 7 years) indicated that 15 of 48 patients with primary melanomas developed metastases, and 12 of them died. Among 105 patients with skin cancers 4 persons had recurrences and were irradiated repeatedly. The main merits of the method compared with surgery and X-ray therapy (the absence of side effects, promptness of the treatment, sterility and precise localization of the effect, vascular thrombosis contributing to an ablastic operation) are discussed. The results obtained with neodymium and CO2--lasers are compared.

[Laser therapy of cervix dysplasia]. / Lazernaia terapiia displazii sheiki matki.

Irradiation from a continuous wave CO2-laser was used in the treatment of 1,234 cases of cervical epithelial dysplasia. Diagnosis was established on the findings of extended colposcopy, aiming biopsy and cytological examination of smears. Such advantages over diathermocoagulation as complication-free recovery and a low incidence of recurrences (2.6%) were shown. The end results were studied.

[The immunomodulating properties of the filtrates of Klebsiella culture broth and the nature of the immunomodulating factor]. / Immunomoduliruiushchie svoistva fil'tratov kul'tural'noi zhidkosti klebsiell i priroda immunomoduliruiushchego faktora.

Influence of broth culture filtrates (BCF) of Klebsiella on delayed type hypersensitivity (DTN) in mice has been studied. BCF of plasmid-containing virulent strains and non-plasmid avirulent strains have been found to suppress DTH to xenogenic splenocytes. Gel filtration data have shown that immunosuppressive factor possesses molecular weight about 180-800 kDa. BCF treated with etanol-ether mixture got to soluble fraction that evidenced for its lipid nature. This factor can be inactivated or reactivated by the treatment with trichloracetic acid, phenol and other chemicals. A new factor is found in reactivated BCF. It induces the ability of E. coli O55 to suppress DTH in mice and looks like Shigella factor. It is supposed that Klebsiella immunosuppressive factors are connected with lipopolysaccharide.

[Clinico-microbiological observations in specialized neonatology departments]. / Kliniko-mikrobiologicheskie nabliudeniia v spetsializirovannykh neonatologicheskikh otdeleniiakh.

Results of dynamic microbiological control and clinical observation in specialized departments of newborns pathologies and nursing of premature children for 1989-1994 are presented. Criteria of gastrointestinal colonization, clinical and bacteriological indices of the risk of appearance of suppurative-septic infections in newborns, the leading role of gram-negative bacteria and the increasing role of staphylococci in the development of these infections have been established. Sensitivity of the hospital strains of microorganisms isolated from newborns and from the environment to antibacterial preparations has been studied. Their multiple drug resistance and uniformity according to antibiotic program which permits supposing intrahospital origin of these strains has been established.