Proteomic Investigation of Murine Neuronal α7-Nicotinic Acetylcholine Receptor Interacting Proteins

The α7-nicotinic acetylcholine receptor (α7-nAChR) is a ligand-gated ion channel that is expressed widely in vertebrates and is the principal high-affinity α-bungarotoxin (α-bgtx) binding protein in the mammalian CNS. α7-nAChRs associate with proteins that can modulate its properties. The α7-nAChR interactome is the summation of proteins interacting or associating with α7-nAChRs in a protein complex. To identify an α7-nAChR interactome in neural tissue, we isolated α-bgtx-affinity protein complexes from wild-type and α7-nAChR knockout (α7 KO) mouse whole brain tissue homogenates using α-bgtx-affinity beads. Affinity precipitated proteins were trypsinized and analyzed with an Orbitrap Fusion mass spectrometer. Proteins isolated with the α7-nAChR specific ligand, α-bgtx, were determined to be α7-nAChR associated proteins. The α7-nAChR subunit and 120 additional proteins were identified. Additionally, 369 proteins were identified as binding to α-bgtx in the absence of α7-nAChR expression, thereby identifying nonspecific proteins for α7-nAChR investigations using α-bgtx enrichment. These results expand on our previous investigations of α7-nAChR interacting proteins using α-bgtx-affinity bead isolation by controlling for differences between α7-nAChR and α-bgtx-specific proteins, developing an improved protein isolation methodology, and incorporating the latest technology in mass spectrometry. The α7-nAChR interactome identified in this study includes proteins associated with the expression, localization, function, or modulation of α7-nAChRs, and it provides a foundation for future studies to elucidate how these interactions contribute to human disease

Proteomic Investigation of Murine Neuronal α7-Nicotinic Acetylcholine Receptor Interacting Proteins

The α7-nicotinic acetylcholine receptor (α7-nAChR) is a ligand-gated ion channel that is expressed widely in vertebrates and is the principal high-affinity α-bungarotoxin (α-bgtx) binding protein in the mammalian CNS. α7-nAChRs associate with proteins that can modulate its properties. The α7-nAChR interactome is the summation of proteins interacting or associating with α7-nAChRs in a protein complex. To identify an α7-nAChR interactome in neural tissue, we isolated α-bgtx-affinity protein complexes from wild-type and α7-nAChR knockout (α7 KO) mouse whole brain tissue homogenates using α-bgtx-affinity beads. Affinity precipitated proteins were trypsinized and analyzed with an Orbitrap Fusion mass spectrometer. Proteins isolated with the α7-nAChR specific ligand, α-bgtx, were determined to be α7-nAChR associated proteins. The α7-nAChR subunit and 120 additional proteins were identified. Additionally, 369 proteins were identified as binding to α-bgtx in the absence of α7-nAChR expression, thereby identifying nonspecific proteins for α7-nAChR investigations using α-bgtx enrichment. These results expand on our previous investigations of α7-nAChR interacting proteins using α-bgtx-affinity bead isolation by controlling for differences between α7-nAChR and α-bgtx-specific proteins, developing an improved protein isolation methodology, and incorporating the latest technology in mass spectrometry. The α7-nAChR interactome identified in this study includes proteins associated with the expression, localization, function, or modulation of α7-nAChRs, and it provides a foundation for future studies to elucidate how these interactions contribute to human disease

Clinical characteristics of Polish patients with ANCA-associated vasculitides-retrospective analysis of POLVAS registry

Objective Antineutrophil cytoplasmic antibody (ANCA)-associated vasculitides (AAV) are rare small to medium-size vessel systemic diseases. As their clinical picture, organ involvement, and factors influencing outcome may differ between countries and geographical areas, we decided to describe a large cohort of Polish AAV patients coming from several referral centers-members of the Scientific Consortium of the Polish Vasculitis Registry (POLVAS). Methods We conducted a systematic multicenter retrospective study of adult patients diagnosed with AAV between Jan 1990 and Dec 2016 to analyze their clinical picture, organ involvement, and factors influencing outcome. Patients were enrolled to the study by nine centers (14 clinical wards) from seven Voivodeships populated by 22.3 mln inhabitants (58.2% of the Polish population). Results Participating centers included 625 AAV patients into the registry. Their distribution was as follows: 417 patients (66.7%) with GPA, 106 (17.0%) with MPA, and 102 (16.3%) with EGPA. Male-to-female ratios were almost 1:1 for GPA (210/207) and MPA (54/52), but EGPA was twice more frequent among women (34/68). Clinical manifestations and organ involvement were analyzed by clinical phenotype. Their clinical manifestations seem very similar to other European countries, but interestingly, men with GPA appeared to follow a more severe course than the women. Fifty five patients died. In GPA, two variables were significantly associated with death: permanent renal replacement therapy (PRRT) and respiratory involvement (univariate analysis). In multivariate analysis, PRRT (OR = 5.3; 95% confidence interval (CI) = 2.3–12.2), respiratory involvement (OR = 3.2; 95% CI = 1.06–9.7), and, in addition, age > 65 (OR = 2.6; 95% CI = 1.05–6.6) were independently associated with death. In MPA, also three variables were observed to be independent predictors of death: PRRT (OR = 5.7; 95% CI = 1.3–25.5), skin involvement (OR = 4.4; 95% CI = 1.02– 19.6), and age > 65 (OR = 6.3; 95% CI = 1.18–33.7). Conclusions In this first multicenter retrospective study of the Polish AAV patients, we have shown that their demographic characteristics, disease manifestations, and predictors of fatal outcome follow the same pattern as those from other European countries, with men possibly suffering from more severe course of the disease

Zmiany w zawartosci biomasy zywych mikroorganizmow w glebach zanieczyszczonych weglowodorami monoaromatycznymi

W pracy przedstawiono wyniki badań, w których oceniano wpływ wybranych węglowodorów aromatycznych na biomasę żywych drobnoustrojów w dwóch, różnych typach gleb, z uwzględnieniem wielkości dawki użytego węglowodoru i czasu inkubacji. W doświadczeniu wykorzystano dwie gleby (piasek gliniasty lekki pylasty oraz glinę lekką pylastą). Próby glebowe o masie 300 g mieszano z wybranym węglowodorem: benzenem, toluenem, etylenem lub ksylenem w dawce 100, 1000, 10 000 mg·kg⁻¹ s.m. gleby. Próbę kontrolną stanowiła gleba bez dodatku węglowodoru. Gleby doprowadzono do 50% maksymalnej pojemności wodnej i inkubowano w temperaturze pokojowej przez 112 dni. Oznaczenie biomasy przeprowadzono metodą SIR (ang. substrate induced respiration). Uzyskane wartości przedstawiono w mg CO₂ zawartego w biomasie żywych drobnoustrojów w przeliczeniu na 100 g s.m. gleby. Na podstawie badań stwierdzono, że wprowadzone do gleby węglowodory aromatyczne wpływały na ilość biomasy żywych mikroorganizmów. W zależności od rodzaju węglowodoru, jego dawki oraz czasu oddziaływania, wpływ ten miał różny charakter (zarówno stymulacji, jak i inhibicji), a obserwowane zmiany często utrzymywały się przez cały okres inkubacji. Węglowodory monoaromatyczne w dniu wprowadzenia do gleby piaszczystej, niezależnie od dawki, powodowały zmniejszenie ilości biomasy żywych mikroorganizmów średnio o 50%. Podobną sytuację obserwowano w glebie gliniastej. W glebie piaszczystej w dniu skażenia na ogół najsilniej redukowały biomasę węglowodory w dawce 10000 mg·kg⁻¹ s.m. gleby (z wyjątkiem benzenu). W glebie gliniastej po wprowadzeniu toluenu i etylobenzenu inhibicja utrzymywała się przez cały okres doświadczenia, natomiast benzen i ksylen w tym stężeniu powodowały gwałtowny wzrost ilości biomasy.The paper presents study results upon the evaluation of influence of selected aromatic hydrocarbons on living microorganism biomass in two different soil types including the dose of hydrocarbon used and incubation time. Two soil types (light dusty loamy sand and light dusty loam) were used in the experiments. Soil samples (300 g each) were mixed with selected hydrocarbon: benzene, toluene, ethylbenzene or xylene at the rates of 100, 1000, 10 000 mg·kg⁻¹ soil DM. The control was the soil with no hydrocarbon addition. Soils were adjusted to 50% of MPW and incubated at ambient temperature for 112 days. The biomass determination was performed by the means of SIR method (substrate induced respiration). The achieved results were expressed in mg of CO₂ contained in biomass of living microorganisms recalculated onto 100 g of soil DM. The studies revealed that aromatic hydrocarbons introduced into the soil affected the number of living organism biomass. Depending on the hydrocarbon used, its dose and interaction time, the influence was of various character (both stimulation and incubation), and the observed changes often lasted for the whole incubation period. Monoaromatic hydrocarbons on the day of introduction into the sandy soil, regardless the rate, caused the decrease of living organism biomass by 50%, on the average. Similar situation was observed in loamy soil. On the day of sandy soil pollution, in general, hydrocarbons at 10 000 mg·kg⁻¹ soil DM rate strongest reduced the biomass (except from benzene). In loamy soil, the inhibition maintained for the whole experiment after the introduction of toluene and ethylobenzene; benzene and xylene caused a sudden increase of biomass amount at the same concentration