Zakażenie wirusowe : internalizacja i transport wewnątrzkomórkowy

Błona komórkowa stanowi podstawową barierę dla infekcji wirusowej, a obecność konkretnych białek na jej powierzchni determinuje specyficzność tkankową i gatunkową wirusa. Aby ją przekroczyć, wirusy wykształciły liczne mechanizmy, których wykorzystanie zależne jest od budowy wirusa oraz komórki docelowej. Wirusy opłaszczone generalnie ulegają fuzji z błoną komórkową, co skutkuje przeniesieniem nukleoproteiny do wnętrza komórki. Proces ten może nastąpić na powierzchni komórki, jednak bardzo często jest poprzedzony internalizacją, w której wykorzystywane są naturalne mechanizmy komórkowe służące w warunkach fizjologicznych między innymi do odżywiania się, czy odbierania cząsteczek sygnałowych. Przedmiotem prezentowanych badań są dwa wirusy wywołujące choroby u człowieka. Pierwszy z nich, ludzki koronawirus OC43 (HCoV-OC43, od ang. human coronavirus OC43), jest częstą przyczyną chorób górnych i dolnych dróg oddechowych o cięższym przebiegu u dzieci, osób starszych oraz z niedoborem odporności. Drugim wirusem wybranym do badań jest wirus Zika (ZIKV, od ang. Zika virus). Choć choroba związana z tym flawiwirusem jest łagodna lub asymptomatyczna, zakażenie może prowadzić do rozwoju zespołu Guillaina-Barrégo (autoimmunologicznego uszkodzenia nerwów) u dorosłych i małogłowia u noworodków. Choć każdy z tych wirusów wykorzystuje inną ścieżkę wejścia, to ostatecznie obydwu udaje się przedostać do wczesnych endosomów, a fuzja ich otoczek z błoną pęcherzyków komórki gospodarza silnie zależy od niskiego pH. W ramach przeprowadzonych badań opisano drogi wejścia obydwu wirusów do komórki gospodarza. Podczas gdy HCoV-OC43 inicjuje klasyczną, stosunkowo wolną (około 90 min) drogę kaweolinozależną, ZIKV wnika do komórek drogą klatrynozależną i ulega fuzji już po 10-15 min. Zastosowanie mikroskopii konfokalnej umożliwiło obserwację kolokalizacji białek wirusowych z białkami markerowymi poszczególnych przedziałów komórkowych. W efekcie byliśmy w stanie śledzić trasę pojedynczych wirionów w początkowej fazie zakażenia komórki. Znaczenie poszczególnych białek w tym procesie zostało następnie potwierdzone poprzez zahamowanie ich aktywności z wykorzystaniem selektywnych inhibitorów lub poprzez wyciszenie wyrażania poszczególnych białek przy pomocy małych interferujących RNA. W związku z tym, że proces internalizacji obydwu wirusów jest wrażliwy na zmiany pH, inkubacja komórek z czynnikami zwiększającymi wewnątrzkomórkowe pH (bafilomycyna A1 oraz NH4Cl) również prowadziła do zahamowania infekcji. Bafilomycyna A1 jest inhibitorem pompy protonowej i hamuje obniżanie pH w przedziałach wewnątrzkomórkowych, natomiast chlorek amonu działa buforująco, neutralizując zakwaszenie endosomu. Oba te związki są złotym standardem stosowanym w badaniach nad endocytozą i uważa się, że ich działanie polega na hamowaniu procesu fuzji poprzez zablokowanie zmian strukturalnych indukowanych niskim pH. W takim scenariuszu w obu przypadkach powinno dochodzić do zatrzymania procesu fuzji, a wirusy powinny wejść na ścieżkę degradacji i zostać usunięte z komórki. Przeprowadzone badania wykazały, że w przypadku bafilomycyny A1 faktycznie realizowany jest ten scenariusz, jednak w obecności chlorku amonu uzyskane wyniki były niezgodne z teorią. Bezsprzecznie wykazaliśmy, że chlorek amonu działa w sposób całkowicie odmienny od opisanego, zmieniając transport wewnątrzkomórkowy i prowadząc do usunięcia wirionów zaraz po wejściu do przedziału endosomalnego poprzez szybki recykling zależny od Rab35. Uzyskane wyniki podważają jeden z paradygmatów w wirusologii i biologii komórki i pozwalają sądzić, że duża część badań dotyczących wczesnych etapów zakażenia wymaga weryfikacji. Przeprowadzone badania pozwoliły lepiej zrozumieć proces zakażenia wirusami HCoV-OC43 i ZIKV, a w przyszłości mogą przyczynić się do opracowania lepszych strategii terapeutycznych w zakresie opracowania inhibitorów wejścia.Although plasma membrane is the primary barrier to viral infection, its composition and the presence of particular proteins on the cell surface determine the possibility of virus entry and, in consequence, tissue and species specificity of the virus. Enveloped viruses have developed different strategies of fusion with host cell membranes, which allows them to deliver their genetic material to the replication site. Subsequent to binding, some viruses are able to fuse with the host membrane immediately on the cell surface, while others need to be delivered to a precisely defined intracellular location. In the latter case, viruses hijack inward transport machinery, that in physiological conditions serves the cell for nutrition and communication. The main goal of this study was to identify and characterize the entry pathways of two viruses that pose a significant risk to human health. Human coronavirus OC43 (HCoV-OC43) frequently causes upper and lower respiratory tract disease, which may take a more severe course in children, the elderly and immunocompromised people. Zika virus (ZIKV) is a mosquito-borne flavivirus, that is usually associated with mild infections manifested by a set of unspecific symptoms, such as fever, rash, conjunctivitis, muscle and joint pain, malaise and headache. However, in some cases neurologic complications may occur, leading to the development of Guillain-Barré syndrome in adults and lethal microcephaly in fetuses. HCoV-OC43 was found to initiate infection via relatively slow (~90 min) caveolin-dependent pathway, whereas ZIKV was found to follow clathrin-dependent endocytosis and to undergo fusion as soon as 10-15 min post inoculation. Using confocal microscopy, we were able to observe co-localization between viral proteins and marker proteins of different intracellular compartments, which allowed us to track single virions during the early phase of infection. The results were further verified using small interfering RNA to transiently silence the expression of proteins involved in the chosen internalization processes, and also by using selective chemical inhibitors of these pathways. As far as the internalization process of both viruses is endocytosis-dependent and thus is sensitive to pH changes, incubation of the virus-overlaid cells with pH-increasing agents hampered the infection development. Bafilomycin A1 is a proton pump inhibitor which suppresses the pH decrease in intracellular compartments, whereas NH4Cl exhibits a simple buffering function and in this way neutralizes the acidification of endosomes. Both of the compounds are golden standards in the research of endocytosis process; by now, it has been commonly believed that their inhibitory function relies on the blocking of structural changes that in physiological (low pH) conditions induce fusion process. In such a scenario, both agents would inhibit the fusion between the viral envelope and the host membrane, which would further lead the virus to the degradation pathway. Although such a pattern has been indeed observed in the case of bafilomycin A1-treated cells, the results obtained for NH4Cl are significantly different. We have demonstrated that NH4Cl reprograms the endocytic hub so that virions are immediately ejected from endosomal compartment via Rab35-driven fast recycling pathway. These data call into question one of the paradigms in virology and cell biology and suggest that much of the research on the early stages of infection requires verification. The complex research on mechanisms of HCoV-OC43 and ZIKV trafficking presented in this study shed light on the regulation of intracellular transport and helped to understand viral strategies of host cells deception. In a broader perspective, identification of cellular factors indispensable for the early steps of infection is going to contribute to the development of novel antiviral therapies

Demony kobiece w literaturze bizantyńskiej

The thesis’ aim is to demonstrate not only the evolution, but also means of representation and various functions of Byzantine female demons. The thesis discusses a selection of texts (from antiquity to late Byzantine times) concerning female demons, notorious for killing newborn babies and harming women during pregnancy, labour or immediately after it, as well as demonesses, whose main task was to tempt men. Although these female demons had originated from beliefs in different cultures, had different names, they possessed identical attributes, e.g. they appeared in similar situations, had also common traits of appearance and skills: polymorphism or flying. The first chapter serves as a background for the next two chapters. It is a presentation of the female demons of the ancient world: Mesopotamia, Israel, Greece and Rome. The second chapter is an analysis of a late antique text, the so called Testament of Solomon. The third chapter is dedicated to Gello – a demoness, whose origins may be traced back to no less than VII century BC (where her name was mentioned for the first time in a poem by Sappho), and who enjoyed great popularity in Byzantium, remains well-known and is still present in tchem folklore of many countries, whose culture is genealogically connected with the Byzantine/Orthodox tradition

Die Entwicklung der Sehhilfenverordnung für Sehbehinderte in den Jahren 1959-1988. Eine retrospektive Studie anhand der Patientenakten der Lupenbrillen-Sprechstunde von Frau Professor Elfriede Aulhorn der Universitäts-Augenklinik Tübingen

Background: In the last century, the variety of low vision aids was limited and only few low vision services were available. The University Eye Hospital in Tuebingen, Germany, offered a “magnifier service” run by Professor Elfriede Aulhorn. This study evaluates retrospectively the spectrum and development of visual aids for low vision patients of this service over 3 decades. Methods: The reports of 1927 low vision patients were evaluated through 3 decades (1: 1959-1968, 2: 1969-1978, 3: 1979-1988) regarding patients` history and complaints, visual acuity, magnification need, perimetry, diagnosis, treatment and rehabilitation measures. Results: Mean age was 51.5 years (SD 25.8, range: 5 months – 101 years). The size of the age group > 65 years increased during the decades.The prevalence of age-related macular degeneration increased from 18.3 % in decade 1 to 24.2% in decade 3. Visual aids were supplied in the 1st decade mostly as Kepler and Galilei systems (55.6%) or magnifying spectacles (36.5%). In the 2nd decade, mostly magnifying spectacles (57%), mostly bifocal ones, were dispensed. During the 3rd decade, the hand-held magnifiers (12,3%) and electronic devices (CCTV monitors) were provided more and more (10.1%). Significance: The increase of age-related eye diseases, the improving technology of optical aids and the availability of the first electronic devices influenced the supply with visual aids over time, along with decreasing percentage of complex telescopic systems and an increasing percentage of magnifying spectacles and electronic devices. This development continued in later decades

Levels of tissue factor pathway inhibitor in patients with inflammatory bowel disease

Endothelial dysfunction has been reported to be involved in the pathogenesis of inflammatory bowel disease (IBD) and concomitant thromboembolic complications. Inflammation stimulates the expression of tissue factor and tissue factor pathway inhibitor (TFPI) by endothelial cells. This study assessed the relationship between TFPI levels and disease activity in patients with IBD. A total of 50 consecutive adult patients with ulcerative colitis (UC), 50 patients with Crohn disease (CD), and 50 healthy controls were enrolled to the study. Plasma levels of total TFPI, free TFPI, and von Willebrand factor were measured. Associations among these levels, disease activity, and inflammatory marker levels were assessed. Total TFPI levels were higher in patients with IBD (median, 68.5 [IQR, 60.2-80.1] ng/ml) than in controls (median, 61.1 ng/ml [IQR, 54.3-74.2]; P = 0.01). Free TFPI levels were higher in patients with active UC (median, 12.8 ng/ml [IQR, 11.1-15.4]), inactive UC (median, 9.9 ng/ml [IQR, 7.3-11.5]), active CD (median, 11.7 [IQR, 9.7-14.4] ng/ml), and inactive CD (median,], 9.7 ng/ml [IQR, 8.6-11.6]) than in controls (median, 5.5 ng/ml [IQR, 4.3-7.2]; P <0.001). In the CD and UC groups, free TFPI levels correlated with the levels of inflammatory markers and disease activity. The von Willebrand factor level was higher in patients with UC (median, 143.4 IU/dl [IQR, 115.5–170.4]) and those with CD (median, 151.8 IU/dl [IQR, 112.8-189.4]) than in controls (85.1 IU/dl [IQR, 77.1-101.5]; P <0.001 for both comparisons). The anticoagulant TFPI pathway is activated during remissions and flares in patients with IBD. The free TFPI level correlates with biochemical markers of inflammation and disease activity

Endocrine Disrupting Compounds – Problems and Challenges

In this chapter, information about some of the estrogenic compounds and their environmental fate and biological influence can be found. Special attention is paid to the review of the analytical approaches used at the stages of detection and determination of Endocrine Disrupting Compounds (EDCs) in the environmental samples. Also, a brief characterization of both cellular and non-cellular bioassays is presented

Serum concentration of selected biochemical markers of endothelial dysfunction and inflammation in patients with the varying activity of inflammatory bowel disease

Introduction Endothelial dysfunction leads to an increased expression of cell adhesion molecules, leukocyte diapedesis, vascular smooth‑muscle tone, excessive permeability of vascular walls, and increased procoagulant activity. Objectives We investigated whether serum levels of several endothelial and platelet activation markers correlated with disease activity in patients with inflammatory bowel disease (IBD). Patients and methods This study included 56 patients with ulcerative colitis, 66 with Crohn disease, and 40 healthy controls. We measured the complete blood count and levels of fibrinogen, C‑reactive protein, albumin, interleukin 6, tumor necrosis factor α, E‑selectin, P‑selectin, monocyte chemoattractant protein 1 (MCP‑1), soluble CD40 ligand (sCD40L), and microparticles. Results There were no significant differences in the median levels of E‑selectin, P‑selectin, MCP‑1, sCD40L, and microparticles between patients with active IBD, those with inactive IBD, and healthy controls. The clinical disease activity assessed with the Mayo scale in the ulcerative‑colitis group was weakly, positively correlated with sCD40L (R = 0.32, P = 0.02), P‑selectin (R = 0.32, P = 0.02), and inflammatory marker levels. The clinical disease activity index in the Crohn disease group was positively correlated with the markers of inflammation yet not with the markers of endothelial activity. Conclusions E‑selectin, P‑selectin, sCD40L, MCP‑1, and microparticle levels do not significantly differ between patients with the varying activity of IBD. However, due to the observed correlations, further studies of a larger patient group should be conducted to confirm our observations

Insulin-like growth factor system in remission and flare of inflammatory bowel diseases

Insulin‑like growth factor 1 (IGF‑1) is involved in the modulation of immunity and inflammation. It also plays a role in regulating the migration of endothelial cells and production of vasoactive agents. This study assessed the concentrations of IGF‑1 and insulin‑like growth factor-binding protein 3 (IGFBP‑3) and their relationships to disease activity in patients with inflammatory bowel disease (IBD). A total of 129 adult patients with IBD (69 with Crohn disease [CD] and 60 with ulcerative colitis [UC]) were involved in the study. The control group consisted of 31 healthy volunteers. Biochemical serum analyses were performed and the associations of IGF‑1 and IGFBP‑3 with inflammatory markers and disease activity were assessed. IGF‑1 levels were decreased in patients with active UC compared with those with nonactive UC (mean [SD], 78.3 [22.7] ng/ml and 96.2 [24.5] ng/ml, respectively; P = 0.02) and controls (94.5 [26.5] ng/ml; P = 0.03). The IGF‑1 level was lower in patients with active CD compared with those with nonactive CD (mean [SD], 79.2 [24.9] ng/ml and 110.1 [43.4] ng/ml, respectively; P <0.001). The IGFBP‑3 level was lower in patients with active UC compared with those with nonactive UC (P = 0.04) and controls (P = 0.04). IGF‑1 correlated negatively with C‑reactive protein (CRP) levels (P <0.01), disease activity (P <0.05), and disease duration (P <0.05). IGFBP‑3 levels correlated negatively with CRP levels (P <0.05). The IGF system is disrupted in patients with IBD. Systemic levels of the IGF axis components are related to disease activity and duration

Oral ulceration in patient with active Crohn’s disease

Choroba Leśniowskiego-Crohna (ChLC) jest przewlekłym procesem zapalnym, który może umiejscawiać się w każdym odcinku przewodu pokarmowego. Wbrew wcześniejszym opiniom uważa się, że zmiany patologiczne w obrębie jamy ustnej występują dość często, lecz rzadko są rozpoznawane. Przedstawiono 30-letniego chorego, u którego w czasie zaostrzenia objawów ChLC wystąpiło owrzodzenie jamy ustnej. Za pomocą badania histologicznego błony śluzowej z otoczenia wrzodu zdiagnozowano zapalenie ziarniniakowe. Zmiany typowe dla ChLC obejmowały dystalny odcinek jelita cienkiego z przetoką między pętlami jelita cienkiego oraz esicę i prostnicę, a potwierdzone były badaniem kolonoskopowym i histologicznym wycinków błony śluzowej oraz badaniem radiologicznym jelita cienkiego. Autorzy zwracają uwagę na przydatność badania stomatologicznego u chorego z zaostrzeniem ChLC. Umożliwia ono rozpoznanie zmian patologicznych błony śluzowej i potwierdzenie w ocenie histopatologicznej nieswoistego zapalenia ziarniniakowego. U chorych na ChLC owrzodzenia w jamie ustnej są przewlekłe i trudno poddają się leczeniu miejscowemu. Podstawowe znaczenie w tych przypadkach ma standardowe leczenie aktywnej ChLC.Crohn’s disease (CD) is a chronic inflammatory process with pathological changes which can involve any part of the gastrointestinal tract. Contrary to previous opinions, involvement of the oral cavity is frequent, but is rarely diagnosed. A thirty-years-old patient with deep ulceration of the oral mucosa which appeared during exacerbation of CD is presented. Granulomatous inflammation of the oral mucosa was confirmed by histopathology. Inflammatory changes typical for CD were present in the distal small bowel with ileo-ileal fistula and in the distal colon, and were confirmed by colonoscopy, histopathological examination of mucosal biopsies and double-contrast radiographic examination of the small bowel. Dental examination of a patient with active CD is helpful in the diagnosis of oral manifestations of the disease and mucosal biopsies can easily be taken for histopathological confirmation of granulomatous inflammation. In patients with CD the oral ulcerations are chronic and difficult to treat. The standard treatment of active CD plays the main role in the approach to these patients