Complete immunization against Trypanosoma cruzi verified in individual mice by complement-mediated lysis

Experimental systems to assay immunity against Trypanosoma cruzi usually demonstrate partial resistance without excluding the establishment of sub-patent infections in protected animals. To test whether Swiss mice immunized with attenuated parasites might develop complete resistance against virulent T. cruzi, experiments were performed involving challenge with low numbers of parasites, enhancement of local inflammation and the combination of natural and acquired resistance. Absence of infection was established after repeated negative parasitological tests (including xenodiagnosis and hemoculture), and lack of lytic antibody was tested by complement mediated lysis. Immunization with 10(7) attenuated epimastigotes conferred protection against the development of high levels of parasitemia after challenge with Tulahuen strain, but was unable to reduce the number of infected animals. However, when a strong, delayed-type hypersensitivity reaction was triggered at the site of infection by injecting a mixture of virulent and attenuated T. cruzi, a significant proportion of immunized animals remained totally free of virulent infection. The same result was obtained when the immunization experiment was performed in four month old Swiss mice, displaying a relatively high natural resistance and challenged with wild, vector-borne parasites. These experiments demonstrate that complete resistance against T. cruzi can be obtained in a significant proportion of animals, under conditions which replicate natural, vector delivered infection by the parasite.

Complement facilitates early prion pathogenesis.

New-variant Creutzfeldt-Jakob disease and scrapie are typically initiated by extracerebral exposure to the causative agent, and exhibit early prion replication in lymphoid organs. In mouse scrapie, depletion of B-lymphocytes prevents neuropathogenesis after intraperitoneal inoculation, probably due to impaired lymphotoxin-dependent maturation of follicular dendritic cells (FDCs), which are a major extracerebral prion reservoir. FDCs trap immune complexes with Fc-gamma receptors and C3d/C4b-opsonized antigens with CD21/CD35 complement receptors. We examined whether these mechanisms participate in peripheral prion pathogenesis. Depletion of circulating immunoglobulins or of individual Fc-gamma receptors had no effect on scrapie pathogenesis if B-cell maturation was unaffected. However, mice deficient in C3, C1q, Bf/C2, combinations thereof or complement receptors were partially or fully protected against spongiform encephalopathy upon intraperitoneal exposure to limiting amounts of prions. Splenic accumulation of prion infectivity and PrPSc was delayed, indicating that activation of specific complement components is involved in the initial trapping of prions in lymphoreticular organs early after infection.

The complement inhibitor low molecular weight dextran sulfate prevents TLR4-induced phenotypic and functional maturation of human dendritic cells.

Low molecular weight dextran sulfate (DXS) has been reported to inhibit the classical, alternative pathway as well as the mannan-binding lectin pathway of the complement system. Furthermore, it acts as an endothelial cell protectant inhibiting complement-mediated endothelial cell damage. Endothelial cells are covered with a layer of heparan sulfate (HS), which is rapidly released under conditions of inflammation and tissue injury. Soluble HS induces maturation of dendritic cells (DC) via TLR4. In this study, we show the inhibitory effect of DXS on human DC maturation. DXS significantly prevents phenotypic maturation of monocyte-derived DC and peripheral myeloid DC by inhibiting the up-regulation of CD40, CD80, CD83, CD86, ICAM-1, and HLA-DR and down-regulates DC-SIGN in response to HS or exogenous TLR ligands. DXS also inhibits the functional maturation of DC as demonstrated by reduced T cell proliferation, and strongly impairs secretion of the proinflammatory mediators IL-1beta, IL-6, IL-12p70, and TNF-alpha. Exposure to DXS leads to a reduced production of the complement component C1q and a decreased phagocytic activity, whereas C3 secretion is increased. Moreover, DXS was found to inhibit phosphorylation of IkappaB-alpha and activation of NF-kappaB. These findings suggest that DXS prevents TLR-induced maturation of human DC and may therefore be a useful reagent to impede the link between innate and adaptive immunity.

Ultrastructure of the membrane attack complex of complement: detection of the tetramolecular C9-polymerizing complex C5b-8.

The ultrastructure of the membrane attack complex (MAC) of complement had been described as representing a hollow cylinder of defined dimensions that is composed of the proteins C5b, C6, C7, C8, and C9. After the characteristic cylindrical structure was identified as polymerized C9 [poly(C9)], the question arose as to the ultrastructural identity and topology of the C9-polymerizing complex C5b-8. An electron microscopic analysis of isolated MAC revealed an asymmetry of individual complexes with respect to their length. Whereas the length of one boundary (+/- SEM) was always 16 +/- 1 nm, the length of the other varied between 16 and 32 nm. In contrast, poly(C9), formed spontaneously from isolated C9, had a uniform tubule length (+/- SEM) of 16 +/- 1 nm. On examination of MAC-phospholipid vesicle complexes, an elongated structure was detected that was closely associated with the poly(C9) tubule and that extended 16-18 nm beyond the torus of the tubule and 28-30 nm above the membrane surface. The width of this structure varied depending on its two-dimensional projection in the electron microscope. By using biotinyl C5b-6 in the formation of the MAC and avidin-coated colloidal gold particles for the ultrastructural analysis, this heretofore unrecognized subunit of the MAC could be identified as the tetramolecular C5b-8 complex. Identification also was achieved by using anti-C5 Fab-coated colloidal gold particles. A similar elongated structure of 25 nm length (above the surface of the membrane) was observed on single C5b-8-vesicle complexes. It is concluded that the C5b-8 complex, which catalyzes poly(C9) formation, constitutes a structure of discrete morphology that remains as such identifiable in the fully assembled MAC, in which it is closely associated with the poly(C9) tubule.

A monoclonal antibody which blocks the function of factor D of human complement.

Factor D is an essential enzyme for activation of complement by the alternative pathway (AP). It has been difficult to obtain mouse monoclonal antibodies (Mabs) which block the function of factor D. We have developed a strategy to obtain such Mabs using a double screening procedure of the initial clones. We selected the clone whose supernatant had the lowest level of anti-factor D Ab by ELISA and abolished factor D haemolytic activity. Addition of this Mab to human serum was shown to abolish conversion of C3 by cobra venom factor, haemolysis of rabbit erythrocytes, and activation of C3 and C5 by cuprophane dialysis membranes.

Levels of complement C3 and C4 components in Amerindians living in an area with high prevalence of tuberculosis

The levels of complement C3 and C4 components were determined in non-indigenous (creoles) and indigenous (Warao) populations, the latter with an extremely high tuberculosis (TB) rate. Serum samples from 209 adults were studied and classified in 4 groups taking into account tuberculin skin tests (TST): (1) the group of Warao patients (58 positive for the TST, WP TST+ and 9 negative for the TST, WP TST-), (2) the group of creole patients (34 positive for the TST, CP TST+ and 9 negative for the TST, CP TST-), (3) the group of healthy Warao controls (38 positive and 14 negative for TST, WC TST+ and WC TST-, respectively), (4) the creole controls (26 positive and 21 negative for the TST, CC TST+ and CC TST-, respectively). With respect to the results concerning the measurement of both complement C3 and C4 components with the exception of the WC TST and the CC groups, the WP TST+ and WP TST- as well as WC TST+ groups showed a significant frequency of individuals with decreased levels of complement C3 component (20.6, 33.3, and 26.3%, respectively) and also C4 component (12.0, 11.1, and 13.3%, respectively) in comparison to both creole patients (CP TST+, 8.82% and CP TST-, 0% and CP TST+, 5.88% and CP TST-, 0%) for C3 and C4, respectively. The study of these parameters carried out in 15 Warao subjects with active infection, before and after anti-TB chemotherapy,statisticallyconfirmedthat the effective chemotherapy did not restore normal levels of the complement C3 and C4 components among Warao patients. Aditional tests for hepatitis B or hepatitis C infection, and the profile of the hepatic proteins were not associated to the deficiency in production of the complement components.In conclusion, the results show that within the Warao population, a high percentage of subjects exhibit decreased levels of both complement C3 and C4 components independent of latent or active infection and the status of TST.

Implementació d'un sistema de generació i tramesa automàtica d'avisos de retorn per correu electrònic com a complement del mòdul de gestió de préstec

SAVIA (Servei d'AVIsos Adhoc) és un sistema compost per una sèrie de programes en PERL desenvolupats per la Biblioteca de la UOC que gestionen la tramesa per correu electrònic dels avisos de retorn de préstec dels documents de la Biblioteca, mitjançant els programes de l'aplicatiu VTLS destinats a la creació de les notificacions impreses. Aquests programes flexibilitzen la comunicació entre la Biblioteca i els usuaris donat que permeten el tractament individualitzat de les trameses per tipologies d'usuari i la incorporació d'addendes temporals als missatges, definides i controlades per la persona propietària del procés del préstec.

Genotype-phenotype correlation of age-related macular degeneration : influence of complement factor H polymorphism

RAPPORT DE SYNTHESE La dégénérescence maculaire liée à l'âge (DMLA) est une maladie très fréquente qui représente la cause principale de cécité légale chez les sujets de plus de 50 ans en Occident. Bien que l'étiologie exacte de cette affection ne soit pas complètement connue, des facteurs environnementaux et génétiques influencent sa survenue et son évolution. A ce sujet, de récentes recherches ont notamment montré une association marquée à une variante du gène CFH, Y402H. Nous ignorons toutefois si le polymorphisme Y402H est associé à un phénotype particulier de la maladie. Cette étude a pour but d'établir si cette variante du gène CFH est associée à certaines caractéristiques phénotypiques précoces. L'étude porte sur quatre cent vingt patients atteints de DMLA qui ont été phénotypés sur la base de photographie du fond d'oeil (International Classification and Grading system for age- related macular degeneration) et génotypés à partir d'ADN leucocytaire à la recherche de la variante Y402H du gène CFH. Ces données ont ensuite fait l'objet d'une analyse statistique de l'association génotype-phénotype (le génotype de 50 sujets-contrôle a été utilisé pour confirmer l'association du polymorphisme avec la DMLA). Les résultats obtenus, corrigés pour l'âge et le sexe, montrent un odds ratio (OR) de développer une DMLA de 2.95 en présence d'au moins un allèle à risque C et de 9.05 pour les homozygotes CC. Par contre, aucune influence n'est observée sur les stades de la maladie (précoce-tardif)? Une association significative entre le génotype CC et la présence de druses périphériques (p=0.028), ainsi que la localisation centrale des druses (p=0,049) a été mise en évidence. Aucune autre tendance n'a été dégagée concernant les critères restants (taille, surface totale recouverte, localisation nasale des druses) ou les changements pigmentaires. Cette étude a permis de confirmer l'association entre la variante Y4G2H et la DMLA dans la population suisse et de conclure que la variante Y402H du gène CFH présente une association géno-phénotypique pour certaines caractéristiques des druses. Il est probable que d'autres facteurs génétiques encore influencent le phénotype de la DMLA. De nouvelles recherches seront nécessaires pour préciser l'influence de ces autres facteurs génétiques ou environnementaux, en vue d'une meilleure compréhension de la pathogenèse de cette affection, et pour développer des mesures thérapeutiques et prophylactiques adaptées.

Navigator-gated coronary magnetic resonance angiography using steady-state-free-precession: comparison to standard T2-prepared gradient-echo and spiral imaging.

RATIONALE AND OBJECTIVES: Recent developments of magnetic resonance imaging enabled free-breathing coronary MRA (cMRA) using steady-state-free-precession (SSFP) for endogenous contrast. The purpose of this study was a systematic comparison of SSFP cMRA with standard T2-prepared gradient-echo and spiral cMRA. METHODS: Navigator-gated free-breathing T2-prepared SSFP-, T2-prepared gradient-echo- and T2-prepared spiral cMRA was performed in 18 healthy swine (45-68 kg body-weight). Image quality was investigated subjectively and signal-to-noise ratio (SNR), contrast-to-noise ratio (CNR) and vessel sharpness were compared. RESULTS: SSFP cMRA allowed for high quality cMRA during free breathing with substantial improvements in SNR, CNR and vessel sharpness when compared with standard T2-prepared gradient-echo imaging. Spiral imaging demonstrated the highest SNR while image quality score and vessel definition was best for SSFP imaging. CONCLUSION: Navigator-gated free-breathing T2-prepared SSFP cMRA is a promising new imaging approach for high signal and high contrast imaging of the coronary arteries with improved vessel border definition.

Prospective comparison of MR enteroclysis with multidetector spiral-CT enteroclysis: interobserver agreement and sensitivity by means of "sign-by-sign" correlation.

Our objective was a prospective comparison of MR enteroclysis (MRE) with multidetector spiral-CT enteroclysis (MSCTE). Fifty patients with various suspected small bowel diseases were investigated by MSCTE and MRE. The MSCTE was performed using slices of 2.5 mm, immediately followed by MRE, obtaining T1- and T2-weighted sequences, including gadolinium-enhanced acquisition with fat saturation. Three radiologists independently evaluated MSCTE and MRE searching for 12 pathological signs. Interobserver agreement was calculated. Sensitivities and specificities resulted from comparison with pathological results ( n=29) and patient's clinical evolution ( n=21). Most pathological signs, such as bowel wall thickening (BWT), bowel wall enhancement (BWE) and lymphadenopathy (ADP), showed better interobserver agreement on MSCTE than on MRE (BWT: 0.65 vs 0.48; BWE: 0.51 vs 0.37; ADP: 0.52 vs 0.15). Sensitivity of MSCTE was higher than that of MRE in detecting BWT (88.9 vs 60%), BWE (78.6 vs 55.5%) and ADP (63.8 vs 14.3%). Wilcoxon signed-rank test revealed significantly better sensitivity of MSCTE than that of MRE for each observer ( p=0.028, p=0.046, p=0.028, respectively). Taking the given study design into account, MSCTE provides better sensitivity in detecting lesions of the small bowel than MRE, with higher interobserver agreement.

Algoritmo para el diagnóstico precoz de la deficiencia de vitamina b12 en ancianos

BACKGROUND: The elderly population is particularly at risk for developing vitamin B12-deficiency. Serum cobalamin does not necessarily reflect a normal B12 status. The determination of methylmalonic acid is not available in all laboratories. Issues of sensitivity for holotranscobalamin and the low specificity of total homocysteine limit their utility. The aim of the present study is to establish a diagnostic algorithm by using a combination of these markers in place of a single measurement. METHODS: We compared the diagnostic efficiency of these markers for detection of vitamin B12 deficiency in a population (n = 218) of institutionalized elderly (median age 80 years). Biochemical, haematological and morphological data were used to categorize people with or without vitamin B12 deficiency. RESULTS: In receiver operating curves characteristics for detection on vitamin B12 deficiency using single measurements, serum folate has the greatest area under the curve (0.87) and homocysteine the lowest (0.67). The best specificity was observed for erythrocyte folate and methylmalonic acid (100% for both) but their sensitivity was very low (17% and 53%, respectively). The highest sensitivity was observed for homocysteine (81%) and serum folate (74%). When we combined these markers, starting with serum and erythrocyte folate, followed by holotranscobalamin and ending by methylmalonic acid measurements, the overall sensitivity and specificity of the algorithm were 100% and 90%, respectively. CONCLUSION: The proposed algorithm, which combines erythrocyte folate, serum folate, holotranscobalamin and methylmalonic acid, but eliminate B12 and tHcy measurements, is a useful alternative for vitamin B12 deficiency screening in an elderly institutionalized cohort.

Complement Factor B Polymorphism and the Phenotype of Early Age-related Macular Degeneration.

Abstract Purpose: Age-related macular degeneration (AMD) has been associated with a number of polymorphisms in genes in the complement pathway. We examined the potential genotype-phenotype correlation of complement factor B (CFB) (R32Q) polymorphisms in Caucasian patients with AMD. Methods: Data from a Central European cohort of 349 patients with early AMD in at least one eye were analyzed for potential associations of the CFB (R32Q/rs641153) polymorphism with phenotypic features of early AMD. Early AMD was classified according to the International Classification and Grading System into predominant drusen size, largest drusen, drusen covered surface, central or ring-like location, peripheral drusen, and pigmentary changes. The potential association with single nucleotide polymorphisms on CFB (R32Q/rs641153) was evaluated for all patients, corrected for age, sex, and the polymorphisms of CFH (Y402H) and ARMS2 (A69S). Results: CFB (R32Q) polymorphisms showed a significant association with smaller drusen size (largest drusen ≤250 µm, p = 0.021, predominant drusen ≤125 µm, p = 0.016), with smaller surface covered by drusen (≤10%; p = 0.02), and with more frequent occurrence of peripheral drusen (p = 0.007). No association was found for pigmentary changes. Conclusions: The CFB (R32Q) polymorphism was associated with AMD characterized by small drusen only, and appeared to be protective of large drusen (OR 0.48/0.45) and of larger drusen covered area (OR 0.34). Furthermore, peripheral drusen were more frequently found (OR 2.27). This result supports the role of complement components and their polymorphisms in drusen formation and may enable a better understanding of AMD pathogenesis.