Macrophages as effector cells of protective immunity in murine schistosomiasis: macrophage activation in mice vaccinated with radiation-attenuated cercariae

Cell-mediated immune responses contributing to macrophage activation were compared in mice that demonstrated partial resistance to challenge Schistosoma mansoni infection as a result of vaccination with radiation-attenuated cercariae or of ongoing low-grade primary infection. Vaccinated mice developed significant delayed hypersensitivity reactions to soluble schistosome antigens in vivo. Splenocytes from vaccinated animals responded to in vitro culture with various specific antigens (soluble adult worm extract, living or disrupted schistosomula) by proliferation and production of macrophage-activating lymphokines as did lymphocytes from S. mansoni-infected animals. Macrophage-activating factors produced by spleen cells from vaccinated mice upon specific antigen stimulation eluted as a single peak on Sephadex G-100 with a molecular weight of approximately 50,000 and contained gamma interferon activity. Moreover, peritoneal macrophages with larvicidal and tumoricidal activity were recovered from vaccinated mice after intraperitoneal challenge with soluble schistosome antigens, a procedure also observed to elicit activated macrophages in S. mansoni-infected animals. These observations demonstrate that vaccination with irradiated cercariae stimulates many of the same cellular responses observed after primary S. mansoni infection, and suggest that lymphokine-activated macrophages may participate in the effector mechanism of vaccine-induced and concomitant immunity to challenge schistosome infection. This is the first demonstration of a potential immune effector mechanism in the irradiated vaccine model.     

Extracellular matrix in hepatic granulomas of mice infected with Schistosoma mansoni. Qualitative and quantitative analysis

To investigate the role of extracellular matrix molecules in the granulomatous inflammation, cercariae of Schistosoma mansoni were injected subcutaneously into BALB/c mice. Well-organized granulomas consisting mainly of stimulated macrophages and epithelioid cells developed in the liver at 11 weeks after infection, thereafter showing a tendency to heal. Fibronectin and heparan sulfate proteoglycan deposition appeared around parasite eggs, then increased distinctly at 11 weeks after infection, and subsequently diminished. Quantities of glycosaminoglycans and hydroxyproline in the egg lesion increased significantly at 11 weeks after infection. Thereafter, the amounts of glycosaminoglycans decreased, whereas hydroxyproline content did not. The data suggest that fibronectin and other macromolecules interact to form granuloma extracellular matrix, and that these extracellular events participate in the development of granulomatous inflammation and subsequent fibrosis induced by schistosome eggs.     

Cutaneous basophil hypersensitivity and inhibited macrophage migration in guinea-pigs with schistosomiasis

In guinea-pigs infected with schistosomes, delayed cutaneous reactions rich in basophils (CBH) were found to characterize skin test responses to schistosome egg antigens. In addition, strong contact hypersensitivity-like skin eruptions with large basophil infiltrates resulted from skin penetration challenge by live cercariae (larvae) in these animals. Oedema and diminished basophil granule staining were noted around schistosomula which had penetrated the skin of sensitized animals. CBH responses to egg antigens and to live cercarial challenges were also noted after immunization with a single injection of dead cercariae.

Using peritoneal exudates from guinea-pigs immunized with dead cercariae or infected with schistosomes, direct macrophage migration inhibition with schistosome antigens was found only in animals with infections. Thus, CBH correlated with intradermal exposure to schistosome cercarial antigens, while MMI correlated with live infections. It is suggested that cutaneous basophil responses may play a role in protection from re-infection with schistosomes, and that dead cercarial vaccines might stimulate this beneficial response, without immunizing for potentially harmful granulomatous hypersensitivity.

     

Autoanti-allotype antibodies and idiotypic network regulating allotype production in rabbits.

A panel of T-cell clones reactive to the soluble egg antigen (SEA) of Schistosoma mansoni is described. The proliferative responses of the primary immune lymph node (LN) cells and T-cell clones to different schistosome antigen preparations (cercarial extract (CE), live or dead schistosomula, adult worm extract, and soluble egg antigen) were compared. Primary immune LN cells could not distinguish between these schistosome antigen preparations. However, when a total of 22 T-cell clones was analysed, a complex pattern of both stage-specific and common (or cross-reactive) antigen reactivity was observed. These patterns have been grouped into four types: (i) five clones were reactive to all different schistosome antigen preparations; (ii) two clones were reactive only to SEA, CE and schistosomula; (iii) three clones were reactive only to SEA and cercariae; (iv) twelve clones were reactive to SEA antigen alone. The T-cell clones were identified as Thy-1-positive cells. All the clones were able to respond to exogenous IL-2 after antigenic stimulation. However, there was a variable degree of IL-2 responsiveness when compared with antigen-specific stimulation. Four T clones were selected for further studies on the genetic control of the proliferative response to schistosome antigens. One of the proliferative T clones was restricted by the IA subregion of the major histocompatibility complex (MHC), and the other three clones are restricted by the IE (AeE alpha) subregion.     

Prime-boost and recombinant protein vaccination strategies using Sm-p80 protects against Schistosoma mansoni infection in the mouse model to levels previously attainable only by the irradiated cercarial vaccine

Advent of an effective schistosome vaccine would contribute significantly toward reducing the disease spectrum and transmission of schistosomiasis. We have targeted a functionally important antigen, Sm-p80, as a vaccine candidate because of its consistent immunogenicity, protective and antifecundity potentials, and important role in the immune evasion process. In this study, we report that using two vaccination approaches (prime boost and recombinant protein), Sm-p80-based vaccine formulation(s) confer up to 70% reduction in worm burden in mice. Animals immunized with the vaccine exhibited a decrease in egg production by up to 75%. The vaccine elicited strong immune responses that included IgM, IgA, and IgG (IgG1, IgG2a, IgG2b, and IgG3) in vaccinated animals. Splenocytes proliferated in response to Sm-p80 produced Th1 and Th17 response enhancing cytokines. These results again emphasize the potential of Sm-p80 as a viable vaccine candidate for schistosomiasis.     

Influence of vaccine deposition site on post-vaccinal viraemia and vaccine efficacy in broiler chickens following in ovo vaccination against Marek's disease

In ovo vaccination against Marek's disease is a widely used technology in the broiler industry.A series of experiments was carried out to determine the site of vaccine deposition in the egg during automated in ovo vaccination, and the effect of vaccine deposition site and dose on vaccine responses following vaccination with cell-associated herpesvirus of turkeys in commercial broiler chickens. Vaccine deposition site following automated in ovo vaccination was principally influenced by the age of embryo, with egg size having a smaller effect. The frequency of vaccine deposition inside the embryo body increased as incubation progressed from day 17.5 to 19.5. In experiments using manual vaccine deposition intra-embryonically (IE) or extra-embryonically (EE) at day 18.5, EE vaccine deposition resulted in a significantly delayed development of post-vaccinal viraemia relative to both IE vaccination and subcutaneous vaccination at hatch. There were no effects of vaccine dose (2000, 4000 or 8000 plaque forming units) on the timing of post-vaccinal viraemia. The timing of post-vaccinal viraemia was found to be a good indicator of the level of protection provided by the vaccine against challenge with earlier viraemia associated with better protection. IE vaccine deposition induced significantly greater protection than EE deposition against challenge with a virulent strain of Marek's disease virus. IE deposition consistently produced a high level of protection (68 to 84%) irrespective of vaccine dose or challenge day, while EE vaccine deposition produced no or low levels of protection (0 to 27%) depending on the vaccine dose and day of challenge. The growth of challenged chickens was also affected by site of vaccine deposition, with significantly higher live weights at day 56 of age in IE compared with EE vaccinated groups. These data suggest that the site of vaccine deposition within the embryo is an important determinant of the success of in ovo vaccination.     

Evaluation of protective immune response in mice by vaccination the recombinant adenovirus for expressing Schistosoma japonicum inhibitor apoptosis protein

Schistosomiasis is a worldwide parasitic disease, and while it can be successfully treated with chemotherapy, this does not prevent reinfection with the parasite. Adenovirus vectors have been widely used for vaccine delivery, and a vaccination approach has the potential to prevent infection with Schistosoma. Here, we developed a recombinant adenoviral vector that expresses Schistosoma japonicum inhibitor apoptosis protein (Ad-SjIAP) and assessed its immunoprotective functions against schistosomiasis in mice. Murine immune responses following vaccination were investigated using enzyme-linked immunosorbent assays (ELISA), lymphocyte proliferation, and cytokine assays. The protective immunity in mice was evaluated by challenging with S. japonicum cercariae. Our results indicated that immunization with the Ad-SjIAP in mice induced a strong serum IgG response against IAP including IgG1, IgG2a, and IgG2b. In addition, lymphocyte proliferation experiments showed that mice treated with Ad-SjIAP significantly increased the lymphocyte response upon stimulation with recombinant Schistosoma japonicum inhibitor apoptosis protein (rSjIAP). Moreover, cytokine assays indicated that vaccination of Ad-SjIAP significantly increased the production of interferon (IFN)-γ and IL-2 as compared to the corresponding control group. Furthermore, following the challenge with S. japonicum cercariae, the vaccine conferred moderate protection, with an average rate of 37.95 % for worm reduction and 31.7 % for egg reduction. Taken together, our preliminarily results suggested that schistosoma IAP may be a potential vaccine against S. japonicum and that adenoviral vectors may serve as an alternative delivery vehicle for schistosome vaccine development.     

Vaccination of the Vervet Monkey (Cercopithecus aethiops) against Infection with Schistosoma mansoni

Pilot experiments were carried out to assess the immunizing potential of radiation-attenuated cercariae of S. mansoni. Groups of 4 monkeys each were vaccinated 4-5 times at 3-5-week intervals using cercariae which had received 10, 20, 40 or 60 krad of gamma radiation. Animals were vaccinated with 1000 or 2000 cercariae per kilogram of body weight. Overall the difference in trans burdens between the vaccinated and unvaccinated groups was highly significant ( P > 0.01). The highest level of protection achieved was 44.4%. This was in monkeys which were immunized live times with 2000, 20-krad cercariae at 3–4-week intervals. Protection levels of 33.3%, 36.6% and 37.0% were achieved in groups which had received, respectively, 1000 20-krad cercariae, 1000 10-krad cercariae, 2000 40-krad cercariae and 2000 60-krad cercariae. Vaccination reduced faecal egg counts markedly and intestinal tissue egg cotints by 20–40%. Antischistosomular antibody was detectable in vitro 3 weeks after the first vaccination. Schistosomule kill rates of up to 60% were observed in vitro.     

Prepubertal vaccination of mice against experimental infection of the genital tract with type 2 herpes simplex virus

Summary Pre-pubertal immunisation of mice with a formalin-inactivated type 1 and 2 herpes simplex virus vaccine conferred a level of life-long protection against primary type 2 genital infection. Protection levels were better with type 1 vaccine and strikingly influenced by vaccine dosage where a one-hundred-fold reduction from the standard vaccine dosage diminished protection to insignificant levels. Vaccine efficacy was not significantly affected by the method of virus inactivation, the number of immunisations or the age of the mouse at immunisation. Vaccination conferred better protection than previous type 2 genital infection; this may be a consequence of a higher antigenic dose, more acceptable antigenic presentation or to a perversion of the immune response in a latently infected animal to homologous virus challenge.With 6 Figures     

Immunopathology of experimental schistosome (S. Mansoni) egg granulomas in mice--possible defence mechanisms mediated by local immune complexes

A ten month experimental S. Mansoni infection was followed in mice by studying schistosome egg granulomatous inflammation, antigen and immunoglobulins deposits in the granulomata and the circulating antibodies. During the experimental infection it was demonstrated that while the granulomatous reaction matures there is a progressive restriction of the antigens to the miracidia. Furthermore, immunoglobulins, particularly IgG and C3 deposits were demonstrated in the granulomata. Our data support the concept that schistosome egg granuloma is a hypersensitivity high turnover granuloma that is particularly efficient in walling off the antigen. Whereas the granuloma is built up largely by a cell mediated immune reaction, antibody deposits are also found, probably functioning as a local humoral antibody barrier which allows a slow and progressive neutralisation of the antigens. Viewed as a whole, experimental schistosome egg granuloma might be considered as a mixed, chiefly cell but also humoral antibody mediated immune reaction.     

Lack of host gut microbiota alters immune responses and intestinal granuloma formation during schistosomiasis

Fatalities from schistosome infections arise due to granulomatous, immune‐mediated responses to eggs that become trapped in host tissues. Schistosome‐specific immune responses are characterized by initial T helper type 1 (Th1) responses and our previous studies demonstrated that myeloid differentiation primary response gene 88 (Myd88)‐deficient mice failed to initiate such responses in vivo. Paradoxically, schistosomal antigens fail to stimulate innate cells to release proinflammatory cytokines in vitro. Since Schistosoma mansoni infection is an intestinal disease, we hypothesized that commensal bacteria could act as bystander activators of the intestinal innate immune system to instigate Th1 responses. Using a broad spectrum of orally administered antibiotics and anti‐mycotics we analysed schistosome‐infected mice that were simultaneously depleted of gut bacteria. After depletion there was significantly less inflammation in the intestine, which was accompanied by decreased intestinal granuloma development. In contrast, liver pathology remained unaltered. In addition, schistosome‐specific immune responses were skewed and faecal egg excretion was diminished. This study demonstrates that host microbiota can act as a third partner in instigating helminth‐specific immune responses.     

Human influenza vaccines and assessment of immunogenicity

Influenza is responsible for the infection of approximately 20% of the population every season and for an annual death toll of approximately half a million people. The most effective means for controlling infection and thereby reducing morbidity and mortality is vaccination by injection with an inactivated vaccine, or by intranasal administration of a live-attenuated vaccine. Protection is not always optimal and there is a need for the development of new vaccines with improved efficacy and for the expansion of enrollment into vaccination programs. An overview of old and new vaccines is presented. Methods of monitoring immune responses such as hemagglutination-inhibition, ELISA and neutralization tests are evaluated for their accuracy in the assessment of current and new-generation vaccines.     

Immunization with a Toll-like receptor 7 and/or 8 agonist vaccine adjuvant increases protective immunity against Leishmania major in BALB/c mice

Activation of Toll-like receptors (TLRs) on antigen-presenting cells of the innate immune system initiates, amplifies, and directs the antigen-specific acquired immune response. Ligands that stimulate TLRs therefore represent potential vaccine adjuvants. In the present study, we determined whether imiquimod and its related compound R848, which are TLR7 and/or TLR8 agonists, represent potential vaccine adjuvants when delivered topically, subcutaneously, or intramuscularly. Using the Leishmania major infection model in BALB/c mice, vaccination with crude Leishmania antigen was not protective against subsequent challenge infection unless it was administered with R848 or a topical application of imiquimod containing cream on the skin. Subcutaneous vaccination with these adjuvants mediated a TH1 response against L. major antigen, which appeared to suppress the TH2 response following a challenge infection. Protective immunity was generated following subcutaneous vaccination but not intramuscular vaccination. These observations suggest that topically administered imiquimod or subcutaneously injected R848 represent potential vaccine adjuvants to enhance the TH1 response, which can be used with existing or new vaccine formulations.     

Isolation and Characterization of 44.6 kDa Protein from Schistosoma japonicum Male Worm

Schistosomes are hermaphrodite. The male worm has somespecific antigen different from the female′s [1,2]. Explo ration and characterization of the male worm antigen com ponents may be of great importance to investigation andprevention of schistosomiasis…     

The acquisition and loss of antigen-specific cellular immune responsiveness in acute and chronic schistosomiasis in man

To characterize the development and evolution of cellular immune responsiveness in individuals infected with the parasite Schistosoma mansoni, we studied fifteen patients with acute, subacute and chronic schistosomiasis. Lymphocytes from the three acutely infected patients responded vigorously to schistosome antigens in an in vitro blastogenic assay. By contrast, cells from nine chronically infected individuals were essentially unreactive to these same antigens. Patients infected for an intermediate period of time (9 months) generated responses between those of acute and chronic patients.

The diminished responsiveness of chronically infected individuals was specific for schistosome antigens and did not extend to humoral immune responses. Following treatment of the infection with niridazole, these patients temporarily regained responsiveness to schistosome antigens.

From these data we speculate that during the course of this parasitic helminth infection there develops a progressive and specific modulation of antigen recognition and proliferation by lymphocytes to schistosome antigens, and that such diminished immune reactivity may be important in maintaining the unique biological relationship which exists between a host and its parasites.

     

Cellular and humoral immune responses and protection against schistosomes induced by a radiation-attenuated vaccine in chimpanzees

The radiation-attenuated Schistosoma mansoni vaccine is highly effective in rodents and primates but has never been tested in humans, primarily for safety reasons. To strengthen its status as a paradigm for a human recombinant antigen vaccine, we have undertaken a small-scale vaccination and challenge experiment in chimpanzees (Pan troglodytes). Immunological, clinical, and parasitological parameters were measured in three animals after multiple vaccinations, together with three controls, during the acute and chronic stages of challenge infection up to chemotherapeutic cure. Vaccination induced a strong in vitro proliferative response and early gamma interferon production, but type 2 cytokines were dominant by the time of challenge. The controls showed little response to challenge infection before the acute stage of the disease, initiated by egg deposition. In contrast, the responses of vaccinated animals were muted throughout the challenge period. Vaccination also induced parasite-specific immunoglobulin M (IgM) and IgG, which reached high levels at the time of challenge, while in control animals levels did not rise markedly before egg deposition. The protective effects of vaccination were manifested as an amelioration of acute disease and overall morbidity, revealed by differences in gamma-glutamyl transferase level, leukocytosis, eosinophilia, and hematocrit. Moreover, vaccinated chimpanzees had a 46% lower level of circulating cathodic antigen and a 38% reduction in fecal egg output, compared to controls, during the chronic phase of infection.     

Acetylcholinesterase from Schistosoma mansoni: immunological characterization

The enzyme acetylcholinesterase (AChE) is present in the trematode Schistosoma mansoni, which infects humans and causes a severe disease called schistosomiasis or Bilharzia. We have purified this enzyme and raised polyclonal antibodies against it. The specificity of these antibodies against the schistosome enzyme was demonstrated by their capacity to precipitate exclusively AChE activity from cercariae extract and to recognize the 8S molecular form of the parasite's AChE. On the other hand, they did not cross-react at all with AChE from human erythrocytes. By employing immunogold electron microscopy, AChE was located on the surface, in the membranal bodies of the tegument and in the muscles of schistosomula. The antibodies raised against the purified AChE of S. mansoni are of protective value, as they led to efficient complement-mediated killing of schistosomula in vitro. It was also demonstrated that antibodies specific towards S. mansoni AChE are present in the sera of mice and of human patients infected with the parasite, suggesting that this enzyme partakes in the immune response towards the parasite during infection. These cumulative data, particularly the schistosomicidal activity of the antibodies and their lack of cross-reactivity with human AChE, are of significance in the consideration of the S. mansoni AChE for vaccination purposes.     

Current issues with influenza vaccination in egg allergy

Influenza infection, with its accompanying morbidity and mortality, represents a major public health concern yearly to the elderly and high-risk groups, including asthmatic patients. Active prevention with vaccination consistently falls short of reaching optimal immunization rates in all risk groups. Asthmatic patients and others with concomitant egg allergy might be denied active immunization because of the risk of inducing adverse reactions with a vaccine derived from egg embryo tissue. Evidence supports the relatively safe administration of influenza vaccine to individuals with egg allergy in whom vaccination is indicated when specific protocols are followed under the supervision of experienced physicians. A practical protocol that includes incremental dosing of influenza vaccine is presented to guide clinicians in influenza vaccination in this high-risk group.     

The search for a vaccine against schistosomiasis - a difficult path hut an achievable goal

Summary: The search for an effective vaccine against schistosomiasis, a parasitic disease currently affecting over 200 million people, remains a desirable but as yet challenging and elusive goal. Progress in the area has been relatively slow but research demonstrating the ability of humans to acquire natural immunity to schistosome infection, together with the successful use in animals of attenuated vaccines, supplemented with encouraging results obtained with defined antigens, suggests that development of a vaccine is achievable. Noteworthy also are recent immune correlate findings which shed light on the complex, putatively protective immune responses in buinans, which have improved the prospects of success. With the first human clinical trial having been completed with a schistosome vaccine candidate, this review examines current progress aimed at achieving the objective of a safe and effective vaccine for widespread use against schistosomiasis. The review emphasises work undertaken in the author's laboratory and those of his chief collaborators in the search for a vaccine against schistosomiasis japonica, a disease of major public health significance in The People's Republic of China and The Philippines. Schistosomiasis vaccines should not be considered as the panacea for schistosomiasis control as, when available, it is generally envisaged that they would be used as one component of an integrated strategy complementing currently available and effective tools such as chemotherapy, improvements to sanitation, piped water supply, effective sewage draining and health education.