Interpain A, a Cysteine Proteinase from Prevotella intermedia, Inhibits Complement by Degrading Complement Factor C3

Periodontitis is an inflammatory disease of the supporting structures of the teeth caused by, among other pathogens, Prevotella intermedia. Many strains of P. intermedia are resistant to killing by the human complement system, which is present at up to 70% of serum concentration in gingival crevicular fluid. Incubation of human serum with recombinant cysteine protease of P. intermedia (interpain A) resulted in a drastic decrease in bactericidal activity of the serum. Furthermore, a clinical strain 59 expressing interpain A was more serum-resistant than another clinical strain 57, which did not express interpain A, as determined by Western blotting. Moreover, in the presence of the cysteine protease inhibitor E64, the killing of strain 59 by human serum was enhanced. Importantly, we found that the majority of P. intermedia strains isolated from chronic and aggressive periodontitis carry and express the interpain A gene. The protective effect of interpain A against serum bactericidal activity was found to be attributable to its ability to inhibit all three complement pathways through the efficient degradation of the α-chain of C3—the major complement factor common to all three pathways. P. intermedia has been known to co-aggregate with P. gingivalis, which produce gingipains to efficiently degrade complement factors. Here, interpain A was found to have a synergistic effect with gingipains on complement degradation. In addition, interpain A was able to activate the C1 complex in serum, causing deposition of C1q on inert and bacterial surfaces, which may be important at initial stages of infection when local inflammatory reaction may be beneficial for a pathogen. Taken together, the newly characterized interpain A proteinase appears to be an important virulence factor of P. intermedia

Acquisition of Complement Inhibitor Serine Protease Factor I and Its Cofactors C4b-Binding Protein and Factor H by Prevotella intermedia

Infection with the Gram-negative pathogen Prevotella intermedia gives rise to periodontitis and a growing number of studies implies an association of P. intermedia with rheumatoid arthritis. The serine protease Factor I (FI) is the central inhibitor of complement degrading complement components C3b and C4b in the presence of cofactors such as C4b-binding protein (C4BP) and Factor H (FH). Yet, the significance of complement inhibitor acquisition in P. intermedia infection and FI binding by Gram-negative pathogens has not been addressed. Here we show that P. intermedia isolates bound purified FI as well as FI directly from heat-inactivated human serum. FI bound to bacteria retained its serine protease activity as shown in degradation experiments with 125I-labeled C4b. Since FI requires cofactors for its activity we also investigated the binding of purified cofactors C4BP and FH and found acquisition of both proteins, which retained their activity in FI mediated degradation of C3b and C4b. We propose that FI binding by P. intermedia represents a new mechanism contributing to complement evasion by a Gram-negative bacterial pathogen associated with chronic diseases

Image of Donbas in ukrainian literature after 1991 year

Głównym przedmiotem niniejszej pracy jest analiza obrazu Donbasu w literaturze ukraińskiej po 1991 roku, próba omówienia dominujących w jego przedstawianiu tendencji, motywów i symboli. Praca ta jest również próbą odpowiedzi na pytanie, czy z omówionych utworów wyłania się spójny obraz Donieckiego Zagłębia Węglowego we współczesnej literaturze ukraińskiej, czy można mówić o powstawaniu ukraińskiego tekstu donbaskiego, analogicznego do tekstu donbaskiego literatury rosyjskojęzycznej. Do tego celu posłużyła mi twórczość prozatorska pisarzy młodszego pokolenia, debiutujących po roku 1991, takich jak Serhij Żadan i Ołeksij Czupa.The topic of this work is the image of Donbas in ukrainian literature after 1991 year - analyze of main symbols, motives and tendencies in this image, based on the works of Oleksiy Chupa and Serhiy Zhadan

Passenger trip patterns changes after launch of Suburban Railway in Małopolska

Szybka Kolej Aglomeracyjna (SKA) jest istotnym elementem obsługi transportowej Krakowa i województwa małopolskiego. Jej uruchomienie i ciągły rozwój istotnie wpłynęły na zmianę zachowań komunikacyjnych mieszkańców Małopolski. W artykule zostały przedstawione wyniki badań ankietowych przeprowadzonych wśród pasażerów SKA oraz obserwacji parkingów Park and Ride przy stacjach i przystankach kolejowych. Na podstawie wyników badań ankietowych określono w jaki sposób i jak długo pasażerowie SKA docierają do przystanków początkowych, jaki środek transportu wykorzystywali przed uruchomieniem połączeń SKA oraz czy zmienili miejsce zamieszkania lub pracy wskutek pojawienia się nowej usługi. Najwięcej pasażerów SKA podróżuje do przystanku początkowego pieszo (około 40%), niezależnie od badanej linii. Ponad połowa ankietowanych dociera do przystanku początkowego w czasie krótszym niż 10 minut. Niemal 1/3 pasażerów już wcześniej podróżowała koleją. Pozostali zrezygnowali z samochodu osobowego (26,3%), prywatnych przewoźników (18,3%) lub komunikacji miejskiej w Krakowie (14,1%). Pojawiły się również podróże wcześniej niewykonywane. Zmiana środka transportu wskutek uruchomienia połączeń SKA jest uzależniona od badanej linii. W artykule oszacowano również zmniejszenie liczby pojazdów, które wjeżdżają do Krakowa oraz oszczędności czasów i kosztów podróży wskutek rezygnacji części pasażerów z samochodu osobowego na rzecz SKA.Suburban Railway (SKA) is an important element of Krakow and Małopolska region transport services. Its development changed inhabitants trip patterns significantly. In the article results of questionnaire survey among SKA passengers and observations of Park and Ride facilities at railway stations are presented. Based on survey results transport mode and access time to initial railway stops were analysed. Moreover transport modes used before SKA launching were identified. Additionally passengers were asked if they had changed place of living or working. Most passengers (around 40%) go to the railway station by foot. More than half of inquired passengers need less than 10 minutes to reach railway station. Almost one third of passengers were travelling by rail before SKA launching. Others have abandoned travelling by their own cars (26,3%), private public transport carriers (18,3%) or municipal public transport in Krakow (14,1%). There were also induced trips observed. Change of transport mode is dependent on the SKA line. As a result of the SKA development reduction of numbers of vehicles entering Krakow was estimated as well as reduction of travel time and travel cost of passengers who changed from the car to rail

Biphasic Effect of Gingipains from Porphyromonas gingivalis on the Human Complement System.

Periodontitis is an inflammatory disease of the supporting structures of the teeth and is caused by, among other agents, Porphyromonas gingivalis. P. gingivalis is very resistant to killing by human complement, which is present in a gingival fluid at 70% of the serum concentration. We found that the incubation of human serum with purified cysteine proteases of P. gingivalis (gingipains) or P. gingivalis wild-type strains W83 and W50 resulted in a drastic decrease of the bactericidal activity of the serum. In contrast, serum treated with P. gingivalis mutants lacking gingipains (particularly strains without HRgpA) maintained significant bactericidal activity. To understand in detail the mechanism by which gingipains destroy the serum bactericidal activity, we investigated the effects of gingipains on the human complement system. We found that all three proteases degraded multiple complement components, with arginine-specific gingipains (HRgpA and RgpB) being more efficient than lysine-specific gingipain (Kgp). Interestingly, all three proteases at certain concentrations were able to activate the CI complex in serum, which resulted in the deposition of C1q on inert surfaces and on bacteria themselves. It is therefore plausible that P. gingivalis activates complement when present at low numbers, resulting in a local inflammatory reaction and providing the bacteria with a colonization opportunity and nutrients. At later stages of infection the concentration of proteases is high enough to destroy complement factors and thus render the bacteria resistant to the bactericidal activity of complement

Biphasic effect of gingipains from Porphyromonas gingivalis on the human complement system

Periodontitis is an inflammatory disease of the supporting structures of the teeth and is caused by, among other agents, Porphyromonas gingivalis. P. gingivalis is very resistant to killing by human complement, which is present in a gingival fluid at 70% of the serum concentration. We found that the incubation of human serum with purified cysteine proteases of P. gingivalis (gingipains) or P. gingivalis wild-type strains W83 and W50 resulted in a drastic decrease of the bactericidal activity of the serum. In contrast, serum treated with P. gingivalis mutants lacking gingipains (particularly strains without HRgpA) maintained significant bactericidal activity. To understand in detail the mechanism by which gingipains destroy the serum bactericidal activity, we investigated the effects of gingipains on the human complement system. We found that all three proteases degraded multiple complement components, with arginine-specific gingipains (HRgpA and RgpB) being more efficient than lysine-specific gingipain (Kgp). Interestingly, all three proteases at certain concentrations were able to activate the CI complex in serum, which resulted in the deposition of C1q on inert surfaces and on bacteria themselves. It is therefore plausible that P. gingivalis activates complement when present at low numbers, resulting in a local inflammatory reaction and providing the bacteria with a colonization opportunity and nutrients. At later stages of infection the concentration of proteases is high enough to destroy complement factors and thus render the bacteria resistant to the bactericidal activity of complement

Binding of complement inhibitor C4b-binding protein contributes to serum resistance of Porphyromonas gingivalis.

The periodontal pathogen Porphyromonas gingivalis is highly resistant to the bactericidal activity of human complement, which is present in the gingival crevicular fluid at 70% of serum concentration. All thirteen clinical and laboratory P. gingivalis strains tested were able to capture the human complement inhibitor C4b-binding protein (C4BP), which may contribute to their serum resistance. Accordingly, in serum deficient of C4BP, it was found that significantly more terminal complement component C9 was deposited on P. gingivalis. Moreover, using purified proteins and various isogenic mutants, we found that the cysteine protease high molecular weight arginine-gingipain A (HRgpA) is a crucial C4BP ligand on the bacterial surface. Binding of C4BP to P. gingivalis appears to be localized to two binding sites: on the complement control protein 1 domain and complement control protein 6 and 7 domains of the alpha-chains. Furthermore, the bacterial binding of C4BP was found to increase with time of culture and a particularly strong binding was observed for large aggregates of bacteria that formed during culture on solid blood agar medium. Taken together, gingipains appear to be a very significant virulence factor not only destroying complement due to proteolytic degradation as we have shown previously, but was also inhibiting complement activation due to their ability to bind the complement inhibitor C4BP