Preventative immunotherapeutic treatments have been an area of great interest to combat infectious disease because of the ability to stimulate the hosts immune system which prepares the host to fight pathogenic microbes. The immunotherapeutic approach requires the use of an immune stimulating molecule that is able to boost the hosts immune response. A major problem exists that these immune stimulating molecules are often very expensive and require a large dose to be effective. To reduce the cost of using these molecules, a delivery system can be used which is able to lower the effective dose of the immune stimulant while not causing any toxic effects towards the hosts health. In this study, the immune stimulating molecules synthetic unmethylated cytidine-phosphate-guanosine oligodeoxynucleotides were attached non-covalently to multi-walled carbon nanotubes. The use of carbon nanotubes as a delivery mechanism could result in a lower effective dose able to stimulate a protective immune response in a chicken model. In this study, we first assessed which of the non-covalant linkages was ideal for linking the immune stimulant to the carbon nanotubes. This was conducted by looking at which method of linkage would allow the best cellular proliferation and transcriptional activation of selected innate immune genes. Once an appropriate linkage method had been selected, cellular uptake studies were conducted to establish that cytidine-phosphate-guanosine oligodeoxynucleotides were delivered to intracellular target receptors. After cellular uptake was demonstrated, it was important that the carbon nanotubes linked to the immune stimulant do not cause toxicity towards the host. To measure toxicity, in vitro studies were conducted to observe cell viability post treatment with carbon nanotube linked immune stimulant. Further studies were conducted on any alterations to the immune stimulants ability to activate immune cells by studying the pathway of macrophage activation. The protective ability of the molecules was then measured by the ability to protect chickens from a lethal challenge with S. typhimurium. Once the protective nature of the molecules was established, the mechanism of immune stimulation was examined by in vivo cell recruitment and in vitro cytokine production. These studies indicate that linking cytidine-phosphate-guanosine oligodeoxynucleotides to carbon nanotubes can lower the effective dose of the immune stimulant without altering the biological function of the molecule.

The purpose of this research was to investigate selected methods of mucosal immunization for commercial chickens. Induction of mucosal immunity in commercial chickens through the use of orally administered subunit vaccines or through immunomodulation of the host??s response to live vaccines may be a viable means to control enteric infections in commercial poultry. In the present investigations we evaluated a means for delivering protein antigen in the drinking water and the use of CpG-ODNs, a recently reported mucosal adjuvant, in order to both improve this response and to modulate the host??s immune response when vaccinated with field strains of Eimeria acervulina and Eimeria tenella. In order to evaluate the efficacy of immunizing commercial poultry with subunit vaccines through the drinking water we chose the model antigen Bovine Serum Albumin (BSA). Chicks were administered BSA via intraperitoneal (I.P.) injection, oral crop gavage, or orally through the addition of BSA to the drinking water. These experiments demonstrated the efficacy of drinking water administration to induce antibodyproduction in the serum, intestine, and bile. When BSA was co-administered with CpGODNs we observed a modest increase in this response dependent upon dose. To evaluate the immunomodulation of the host response to live parasite using CpG-ODNs we used three administration models. The first was a single dose of CpGODNs with a trickle immunization regime of Eimeria acervulina. The second was coadministration of CpG-ODNs with a clinical dose of Eimeria acervulina or tenella. The third was pre-administration of CpG-ODNs 24 hours prior to the clinical dose of either species. These studies demonstrate that the first and third models were effective in reducing lesions and improving performance.

Bacterial DNA is immunostimulatory, and the motifs responsible for this activity are unmethylated CpG dinucleotides. Following cellular uptake, CpG-containing oligodeoxynucleotides (CpG ODN) are trafficked to the endosome where they bind Toll-like receptor 9 (TLR9) to initiate a signaling cascade that culminates in the release of numerous pro-inflammatory cytokines. Because of their immunostimulatory properties, CpG ODN are being clinically evaluated as treatments and vaccine adjuvants for infectious diseases, cancer, and allergic disorders. α2-Macroglobulin (α2M) is a human plasma protein that binds and modulates the activity of a variety of cytokines, growth factors, enzymes, and antigens. Upon proteolytic activation, α2M is converted to its receptor recognized form, α2M*, and rapidly binds to and is internalized by immune competent cells expressing the α2M* endocytic receptor, LRP, and is then trafficked to the endosome. Based on these interactions, α2M seems to play an important role at sites of infection and inflammation by controlling the level of proteinase activity, modulating cytokine signals, and enhancing antigen processing for the adaptive immune response. Here, we report the first evidence that α2M* binds and forms stable complexes with nucleic acids. We have characterized the mechanisms and stoichiometry of this interaction, examined the pH and temperature stability of these complexes, and identified structural variables in the nucleic acids, namely length, base composition, and chemical modifications, that affect the nature of this interaction. We hypothesized that CpG ODN incorporation into α2M* may alter their immunostimulatory properties. Murine macrophages (MΦs) treated with α2M*-ODN complexes respond more rapidly and produce a greater cytokine response than those treated with free CpG ODN alone. Treating human PBMCs with α2M*-ODN complexes likewise demonstrated their enhanced ability to elicit immune responses. This was due to more rapid uptake and CpG ODN protection from degradation by extracellular nucleases. Co-incorporation of both protein ligands and CpG ODN into α2M* yields ternary complexes; these may permit the simultaneous delivery of both protein antigens and adjuvants to immune competent cells, potentially greatly enhancing the adaptive immune response and protective immunity. Based on the findings that incorporation into α2M* confers enhanced immunostimulatory activity of CpG ODN, this technology may be exploited to improve CpG ODN-based therapeutics by increasing efficacy, minimizing side effects, reducing dosing requirements, and reducing cost. / Dissertation

The objective of this investigation was to evaluate the effects of administering CpG-ODN to commercial strain chickens as a potential adjuvant to vaccination against Salmonella, Eimeria spp., and Newcastle disease virus, or immunization to bovine serum albumin (BSA). During Experiment 1, which evaluated the dual application of CpG-ODN and a Newcastle disease virus vaccine, in the first of three replicate trials, on day 28 of the experiment, animals in the Vaccine + CpG 1& 14 experimental group were observed to have the highest levels of (p<0.05) anti-NDV IgG in serum. These levels were elevated above levels in animals from all other experimental groups. This suggestion for an adjuvant effect associated with CpG-ODN administration was not supported in the remaining two trials of experiment 1. Experiment 2 evaluated the potential for CpG-ODN to adjuvant a commercial live oocyst coccidial vaccine when applied by an oral route to neonatal broiler chickens. Overall, when body weight gain during challenge, development of intestinal lesions, and anti-Eimeria IgG levels were evaluated, vaccine administration alone was demonstrated to provide the best measure of protection among animals in all experimental groups, including those receiving either CpG-ODN or Non CpG-ODN. Experiment 3 investigated the simultaneous administration of CpG-ODN or Non-CpG ODN and a commercially acquired Salmonella typhimurium vaccine to SCWL chickens. Similar to experiments 1 and 2, antigen specific IgG responses in serum and indices of protection against field strain Salmonella challenge were variable and inconsistent. Anti-BSA IgG levels were compared in broiler and SCWL chickens immunized against BSA by a drinking water route of administration alone, or in combination with two different concentrations of CpG-ODN or Non CpG-ODN in experiment 4. The only observation where CpG-ODN and BSA co-administration resulted in anti-BSA IgG levels that were elevated above BSA alone immunized chickens was measured in broilers at day 19 post-final immunization. Taken together, given the variable results reported in this investigation related to the co-administration of ODN and vaccine or protein antigen, these data are largely inconclusive for suggesting that CpG-ODN can effectively adjuvant humoral immune responses in commercial strain chickens.

No description available.

In der vorliegenden Arbeit wurde die Auswirkung der Behandlung muriner Mikrogliazellen mit Agonisten von TLR2, TLR4 und TLR9 untersucht. Zu diesem Zweck wurden murine Mikrogliazellkulturen angelegt. Als TLR - Agonisten dienten Pam3Cys und HKAL (TLR2), Pneumolysin und LPS (TLR4) sowie CPG (TLR9). Die Stimulation muriner Mikrogliazellen mit den verschiedenen TLR - Agonisten führte zur Freisetzung von NO und TNF-α. Durch den Einsatz unterschiedlicher Konzentrationen der TLR - Agonisten konnten Dosis - Wirkungs - Kurven für die Freisetzung von NO und TNF-α erstellt werden. Anhand der EC50 wurde die Potenz der TLR - Liganden beurteilt. Für die Freisetzung von NO wies LPS die höchste stimulatorische Potenz auf, gefolgt von Pneumolysin, CpG und Pam3Cys. Für die TNF-α - Freisetzung besaß ebenfalls LPS die höchste stimulatorische Potenz, auch hier folgten Pneumolysin und CpG. Das verwendete Pam3Cys löste sich nicht optimal, vermutlich aus diesem Grund wurde durch die Pam3Cys - Gabe keine maximale Stimulation erreicht. Darum konnte die EC50 für die TNF-α - Freisetzung fur Pam3Cys nicht ermittelt werden. Die EC50 für die TNF-a - Freisetzung war jeweils höher als die entsprechende EC50 für die Freisetzung von NO. Die Behandlung mit HKAL führte zur starken NO - und TNF-a - Freisetzung. Ein direkter Vergleich der Potenz von HKAL mit der der anderen Liganden ist nicht möglich, da die Konzentration von HKAL in Zellzahl pro ml gemessen wird. Die Konzentrationen von Pam3Cys, Pneumolysin und LPS werden jedoch in µg/l gemessen.  Die Stimulation von Mikrogliazellen über verschiedene TLR hatte eine relativ gleich starke Sezernierung von NO und TNF-a zur Folge. Die Costimulation der Mikrogliazellen mit Konzentrationen von zwei unterschiedlichen TLR - Agonisten, welche allein jeweils zur maximalen NO - Produktion geführt hatten, resultierte nicht in einer weiteren Erhöhung der NO - Freisetzung.  Niedrig dosiert zeigte die Pneumolysinbehandlung einen immunstimulatorischen Effekt. Das Maximum an Stimulation, gemessen an der Zunahme der NO - Produktion, wurde bei einer Pneumolysin - Konzentration von 0,3 µg/ml (6nM) beobachtet. Eindeutige zytotoxische Effekte anhand der signifikant geringeren NO - Freisetzung waren bei Konzentrationen von 3 µg/ml bzw. 10 µg/ml (60 nM bzw. 200 nM) nachweisbar. Durch die Isolectin-B4 - Färbung wurden bei diesen Konzentrationen pneumolysinbedingte Zellschäden dargestellt. Bei den anderen Substanzen wurde in hohen Konzentrationen keine Zytotoxität beobachtet Die Behandlung TLR4 - defizienter Mikrogliazellen mit den spezifischen TLR4 - Agonisten Pneumolysin und LPS führte zu einer signifikant geringeren Freisetzung von NO im Vergleich zu Wildtypzellen. Schlussfolgernd kann gesagt werden: Die Stimulation von Mikrogliazellen über unterschiedliche TLR resultiert in einer relativ einheitlichen Freisetzung von NO und TNF-α. Die gleichzeitige Stimulation mit zwei jeweils niedrigdosierten TLR - Agonisten führt zu einem additiven oder supraadditiven Effekt. Nicht nur bakterielle Substanzen, sondern auch endogene Stoffe sind Agonisten an TLR - Rezeptoren. Der additive Effekt durch die simultane Stimulation mehrerer TLR erhöht nicht nur die Sensitivität von Mikroglia während Infektionen, sondern kann ebenfalls Wechselwirkungen zwischen exogenen und endogenen Agonisten von TLR zur Folge haben. Dies kann ein Grund für die Exazerbation oder Induktion autoimmuner Krankheiten durch Infektionen sein.

Submitted by Anderson Silva (avargas@icict.fiocruz.br) on 2012-05-21T19:40:25Z No. of bitstreams: 1 alexandre_santos_silva_ioc_bp_0011_2010.pdf: 1220834 bytes, checksum: e61580079e61e5f06f901abb6feddfb9 (MD5) / Made available in DSpace on 2012-05-21T19:40:25Z (GMT). No. of bitstreams: 1 alexandre_santos_silva_ioc_bp_0011_2010.pdf: 1220834 bytes, checksum: e61580079e61e5f06f901abb6feddfb9 (MD5) Previous issue date: 2010 / Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Rio de Janeiro, RJ, Brasil. / A hepatite A é uma doença endêmica no Brasil e na América Latina. A prevalência da infecção tem correlação com precárias condições de higiene e saneamento. Em países em desenvolvimento, um saneamento inadequado resulta em maior transmissão desta doença, principalmente entre crianças e jovens. Atualmente, devido às melhorias das condições sanitárias, o perfil epidemiológico da doença está se deslocando para idades mais avançadas, o que facilita a ocorrência de surtos epidemiológicos. Os kits comerciais para detecção de anti-HAV total normalmente utilizam imunoglobulina G (IgG) de mamíferos no período convalescente da doença para a produção dos anticorpos de captura e do conjugado. Uma alternativa à aplicação dos anticorpos de mamíferos no diagnóstico é o uso da imunoglobulina Y (IgY), encontrada no soro e gema dos ovos de aves e répteis. Essas proteínas têm varias vantagens quando comparadas com IgG: alta resposta contra antígenos de mamíferos, redução da cor de fundo em ensaios imunoenzimáticos e de serem obtidas por um método não-invasivo (coleta da gema dos ovos). O objetivo deste trabalho foi a obtenção de anticorpos IgY anti-HAV produzidos em galinhas imunizadas contra o vírus da Hepatite A (HAV) e o desenvolvimento de um ensaio imunoenzimático para detecção de anti-HAV total utilizando IgY anti-HAV como imunoglobulinas de captura e conjugado. Cinco grupos de galinhas foram imunizadas com diferentes inóculos contendo: vacina com e sem o adjuvante CpG-ODN, HAV com adjuvante incompleto de Freund (IFA) com e sem o adjuvante CpG-ODN e um grupo controle com IFA. Os ovos foram coletados e a gema foi purificada pela precipitação com polietileno glicol. A solução purificada contendo IgY anti-HAV foi avaliada para determinação da concentração da IgY anti-HAV por espectrofotometria e sua especificidade e título foram determinados à partir de um teste imunoenzimático. Os anticorpos foram conjugados com a peroxidase e foi estabelecida a diluição ideal para os anticorpos de captura e conjugado. Para avaliar o ensaio imunoenzimático “in-house” com IgY anti-HAV, foi avaliado um painel composto de 100 amostras positivas e 100 amostras negativas para anti- HAV-total. A presença da IgY anti-HAV nas gemas dos ovos foi confirmada por SDS-PAGE e Western Blotting, e após a purificação, a média da concentração de proteínas nas gemas dos ovos foi de 8,7406 mg /mL. O grupo imunizado com HAV, IFA e CPG-ODN apresentou os maiores títulos de anticorpo. O ensaio “in-house” apresentou sensibilidade de 84%, especificidade de 79% e eficiência de 81,5%. Os métodos utilizados para a produção de IgY anti-HAV e sua conjugação com peroxidase foram eficientes e o ensaio imunoenzimático “in-house” IgY anti-HAV demonstrou uma boa sensibilidade e especificidade. A produção de anti-HAV IgY apresenta vantagens quando comparado com obtenção da IgG anti-HAV. O teste imunoenzimático “in house” com IgY anti- HAV pode ser uma alternativa a utilização da IgG nos ensaios imunoenzimáticos. / Hepatitis A is an endemic disease in Brazil and Latin America. Prevalence of this infection is related to the low degree of hygiene and sanitation. In developing countries, inadequate sanitation results in larger transmission of the disease mostly in children and young people. Nowadays, due to better sanitation conditions, the epidemiological profile of disease is changing to older ages resulting in the occurrence of outbreaks. Diagnostic kits for detection of total anti-HAV generally use mammals immunoglobulin G (IgG) in the convalescent period of disease for production of capture and conjugated antibodies. One alternative to the application of mammals antibodies in the diagnosis is the use of immunoglobulin Y (IgY), encountered in birds and reptiles. These proteins have several advantages when compared to IgG: high response against mammals antigens, reduction of the background in imunoenzymatic assays and it is obtained by a non-invasive method (harvest of the egg yolks). The objective of this work was the acquisition of anti-HAV IgY antibodies produced in immunized chickens against Hepatitis A virus and the development of an immunoenzymatic assay for total anti-HAV detection using IgY anti-HAV as capture and conjugated immunoglobulins. Five groups of chickens were immunized with different inocula containing: vaccine with and without CpG-ODN adjuvant, HAV with incomplete Freund adjuvant (IFA) with and without CpG-ODN and one control group with IFA. The eggs were harvested and the yolk was purified by precipitation with polyethylene glycol. The purified solution containing anti-HAV IgY was evaluated by espectrofotometry and their specificity and title were determined by an immunoenzymatic assay. These antibodies were conjugated with peroxidase and was estabilized the ideal dilution for capture and conjugated antibodies. For evaluation of the immunoenzymatic “in-house” assay with IgY anti-HAV, a panel composed of 100 positive samples and 100 negative samples for total anti-HAV was assessed. The presence of IgY anti-HAV in egg yolks was established by SDS-PAGE and Western Blotting, and after the purification, the average of the proteins concentrations in the egg yolks was of 8,7406 mg/mL. The group immunized with HAV, IFA and CpG-ODN demonstrate the higher titer of antibodies. The “in-house” assay showed sensibility of 84%, specificity of 79% and efficiency of 81,5%. The methods used for anti-HAV IgY production and conjugation with peroxidase were efficient and the “in-house” immunoenzymatic assay IgY anti-HAV demonstrated a good sensitivity and specificity. The production of IgY anti-HAV showed advantages when compared to the acquisition of IgG anti-HAV. The immunoenzymatic “in-house” assay IgY anti- HAV can be an alternative to the utilization of IgG in immunoenzymatic assays.

Vaccines have become an important approach for the treatment of cancer. Cancer vaccines help the immune system to detect and eradicate tumor cells. Also, cancer vaccines are designed to stimulate an effective immune response that can create long-term immune memory to prevent tumor recurrence. This treatment approach involves the administration of a vaccine comprising or encoding an antigen and can often be combined with an adjuvant to further promote the immune response. The goal of this research was to study the effect of the polyanhydride composition of prophylactic cancer vaccine formulations on the tumor-specific immune response. To achieve this goal, three different amphiphilic polyanhydride copolymers were generated comprising different ratios of 1,6-bis-(p-carboxyphenoxy)-hexane (CPH) and 1,8-bis-(p-carboxyphenoxy)-3,6-dioxaoctane (CPTEG) or sebacic anhydride (SA) monomers. These copolymers were used to fabricate particles encapsulating a model antigen, ovalbumin (OVA), using a double emulsion solvent evaporation technique. The ability of the three different compositions of amphiphilic polyanhydride copolymers (50:50 CPTEG:CPH, 20:80 CPTEG:CPH, and 20:80 CPH:SA) encapsulating OVA to elicit immune responses was investigated. Further, the impact of soluble unmethylated oligodeoxynucleotides containing deoxycytidyl-deoxyguanosine dinucleotides (CpG ODN), an immunologic adjuvant, on the immune response to the three formulations was also studied. The immune response to cancer vaccines was measured after treatment of C57BL/6J mice with two subcutaneous injections, seven days apart, of 50 μg OVA encapsulated in particles composed of different polyanhydride copolymers with or without 25 μg CpG ODN. In vivo studies showed that 20:80 CPTEG:CPH particles encapsulating OVA significantly stimulated the highest level of CD8+ T lymphocytes, generated the highest serum titers of OVA-specific IgG antibodies, and produced longer survival in comparison to formulations involving the other polyanhydride copolymers. The results also revealed that supplementing the vaccine formulations with CpG ODN did not enhance the immunogenicity of OVA. These results accentuate the crucial role of the copolymer composition of polyanhydrides in stimulating the immune response and improving cancer vaccine efficacy.

Toll-like receptors (TLRs) recognize microbial components as danger signals and induce immune responses. TLR’s are expressed in many tissues of the host that are involved in immune responses including the intestines where they are abundantly expressed. This situation presents a challenge in the gastrointestinal tract which is constantly exposed to a wide variety of commensal organisms. Therefore, innate immune recognition in the intestine must be tightly regulated to prevent unwanted inflammation against harmless commensal micro-organisms and yet allow for the induction of protective immunity to invading pathogens. A dysregulation of this balance can result in intestinal inflammation. Peyer’s patches (PP) are the primary site for the induction of immune responses in the intestine and abundantly express TLRs. It is not known how PP regulate microbial signals from commensal bacteria and yet mount vigorous immune responses against dangerous pathogens. CpG DNA, an agonist for TLR9, can strongly activate immune cells in blood, lymph nodes and spleen. However, CpG very poorly activates immune cells from Peyer’s patches, although these cells express TLR9 [1, 2]. Understanding how TLR responses are regulated in PP cells will unveil important information on how immune responses are regulated in the intestine. Investigations from our laboratory have revealed a B cell population (CD5-CD11c-CD21+) in PP that spontaneously secrete high levels of IL-10 which in turn down regulates TLR9 induced IFN and IL-12 production. These IL-10-secreting PP B cells represent a novel subset of the recently proposed regulatory B cells (Bregs) in the intestine [1, 3]. Bregs may have a role in maintaining tolerance to commensal bacteria thereby achieving intestinal homeostasis. The overall goal of the work described in this thesis was to improve our understanding of the immunobiology of Bregs. We performed several experiments to achieve this goal. First, we studied the development of regulatory B cells in lambs of different ages. Jejunal PP were collected from 3-4 month old, neonatal and fetal lambs and the production of IL-10 (the immunoregulatory cytokine secreted by Bregs) was assayed. We found that IL-10 was secreted by CD21+ B cells from the PP in all the three age groups, confirming that Bregs develop prior to birth. We then wondered whether our CD21+ B cells might be contaminated with other cells or activated when using MACS to enrich B cells. To address this issue, we prepared very highly purified CD21+ B cell population using high speed cell sorting to negatively enrich for B cells. We also sorted DCs and assayed IL-10 production in both cell populations. Only the PP B cells spontaneously secreted IL-10. In contrast, dendritic cells, T cells, macrophages, neutrophils and NK cells did not secrete detectable IL-10. Since B cells exist as regulatory and effector populations in mice, we wondered whether an effector B cell population also existed in ovine PP that secreted the pro-inflammatory cytokines IFN-, IFN- and IL-12. Therefore, ovine PP B cells were fractionated into CD72+CD21+and CD72+CD21- subpopulations to assess their capacity to secrete pro-inflammatory cytokines. Interestingly, the CD72+CD21- B cell population secreted the cytokines IFN-, IFN- and IL-12 suggesting there was an effector population. We then surveyed for Bregs in different mucosal and peripheral tissues in sheep. We observed the Bregs frequency varied among the different lymphoid tissues. Finally, we investigated whether Bregs were present in PP of other ruminant species. We identified Bregs exist in PP of neonatal calves. In conclusion, our investigations reveal that ovine Bregs develop in utero prior to antigen exposure, and are present in a variety of mucosal and peripheral tissues. We also report the novel observation that two distinct B cell sub-populations are present in ovine jejunal PP’s: Regulatory and effector B cells.

Escherichia coli ist eine der Hauptursachen von Meningitis und Meningoencephalitis in älteren und immunsupprimierten Patienten sowie von der durch Gram-negative Bakterien verursachten Meningitis bei Säuglingen. In der vorliegenden Arbeit untersuchten wir den Beitrag der neutrophilen Granulozyten und der TLR-Signalkaskade zur Resistenz adulter Mäuse gegen eine intrazerebrale Escherichia-coli-K1-Infektion, anhand von Mäusen, deren neutrophile Granulozyten durch den Anti-Ly-6G-Antikörper depletiert wurden sowie mit Hilfe von MyD88-defizienten und TRIF-defizienten Mäusen. Ein Mangel an MyD88-Adapter-Proteinen reduzierte dramatisch das Überleben der Tiere, während das Fehlen von TRIF-Adapter-Proteinen keine Einschränkung im Überleben nach sich zog im Vergleich zu den Wildtypmäusen. Die Depletion der neutrophilen CD11b+Ly-6G+Ly-6Cint-Granulozyten durch intraperitoneale Injektion des Anti-Ly-6G-Antikörpers führte zu höheren bakteriellen Titern im Kleinhirn und in der Milz und schließlich zu einer erhöhten Letalität im Vergleich zu den mit dem Isotyp behandelten Mäusen. Unsere Ergebnisse zeigen, dass der den Toll-like-Rezeptoren nachgeschaltete MyD88-Signalweg und die neutrophilen Granulozyten wichtige Elemente in der Immunabwehr während der Frühphase einer Escherichia-coli-Meningitis sind. Wie bereits in verschiedenen Experimenten gezeigt wurde, können CpG-ODNs Tiere vor verschiedenen Infektionen schützen (Elkins et al. 1999; Barrier et al.2006). Vor diesem Hintergrund prüften wir, ob eine prophylaktische Behandlung mit CpG-ODN ebenfalls vor einer experimentell erzeugten Infektion des Zentralen Nervensystems schützt. Dazu wurden Mäuse, denen mit Hilfe des Anti-Ly-6G-Antikörpers eine Neutropenie induziert wurde, sowie immunkompetente Mäuse und TLR9-defiziente Mäuse jeweils intrakraniell mit Escherichia coli K1 infiziert. Drei Tage vor der Infektion bekamen die Mäuse der CpG-Gruppen eine einmalige intraperitoneale Injektion von 100 μg CpG-ODN. Hierbei zeigte sich, dass die Behandlung mit CpG-ODN die Letalität nach einer intrazerebralen Infektion von 66 % auf 23 % bei den neutropenischen Mäusen senkte (P = 0,0002, Log-Rank-Test). Zusätzlich wiesen die mit CpG-ODN behandelten Mäuse 42 Stunden nach Infektion geringere bakterielle Titer im Kleinhirn und in der Milz auf, als es in den mit Puffer behandelten Tieren der Fall war (P = 0,01 und P = 0,04, Mann-Whitney-U-Test). In den immunkompetenten Mäusen war die Letalität der mit CpG-ODN behandelten Mäuse leicht niedriger im Vergleich zur Puffergruppe, es konnte jedoch kein statistisch signifikanter Unterschied gefunden werden. TLR9-defiziente Mäuse erhielten keinen schützenden Effekt durch die prophylaktische Behandlung mit CpG-ODN. Dies zeigt, dass die prophylaktische Behandlung mit CpG-ODN nicht nur während einer systemischen Infektion einen Vorteil bringt, sondern ebenfalls die Resistenz von neutropenischen Mäusen gegen eine ZNS-Infektion mit Escherichia coli erhöht. Somit könnte CpG-ODN in Zukunft ein Mittel bieten, um neutropenische Patienten prophylaktisch vor einer Escherichia-coli-K1-Meningitis zu schützen und ihnen somit schwere Verlaufsformen und Komplikationen der Krankheit zu ersparen.