α-Enolase as a novel vaccine candidate against Streptococcus dysgalactiae infection in cobia (Rachycentron canadum L.).

Streptococcus dysgalactiae is an important pathogenic bacterium that has caused economic loss for the cobia industry in Taiwan, ROC. This study presents a highly effective subunit vaccine composed of a moonlight protein, α-enolase, for the prevention of S. dysgalactiae infection. First, α-enolase was cloned, transformed, and expressed in E. coli for production of recombinant protein. Then, the protective efficacies of α-enolase recombinant protein were evaluated in combination with either a pro-inflammatory cytokine, TNF-α, or an oil adjuvant, ISA 763 AVG. The results showed that the combination of α-enolase and ISA 763 AVG was highly protective (RPS = 88.89%), while a negative effect was found in the group immunised with α-enolase adjuvanted with TNF-α (RPS = 22.22%). A further study was conducted with double dose of ISA 763 AVG, which led to an increased RPS value of 97.37%. Moreover, immunised cobia exhibited significantly greater lysozyme activity, antibody responses, and expression of certain immune-related genes post-challenge. Altogether, our results demonstrated that a combination of α-enolase recombinant protein with ISA 763 AVG adjuvant is a promising vaccine that can be employed for protection of cobia against S. dysgalactiae infection.

Immunization with recombinant enolase of Sporothrix spp. (rSsEno) confers effective protection against sporotrichosis in mice.

In recent years, research has focused on the immunoreactive components of the Sporothrix schenckii cell wall that can be relevant targets for preventive and therapeutic vaccines against sporotrichosis, an emergent worldwide mycosis. In a previous study, we identified a 47-kDa enolase as an immunodominant antigen in mice vaccinated with an adjuvanted mixture of S. schenckii cell wall proteins. Here, we sought to assess the protective potential of a Sporothrix spp. recombinant enolase (rSsEno) formulated with or without the adjuvant Montanide Pet-GelA (PGA) against the S. brasiliensis infection in mice. Mice that were immunized with rSsEno plus PGA showed increased antibody titters against rSsEno and increased median survival time when challenged with S. brasiliensis as compared with mice that had not been immunized or that were immunized with rSsEno alone. Immunization with rSsEno plus PGA induced a predominantly T-helper 1 cytokine pattern after in vitro stimulation of splenic cells with rSsEno: elevated levels of IFN-γ and IL-2, as well as of other cytokines involved in host defense against sporotrichosis, such as TNF-alpha, IL-6, and IL-4. Furthermore, we show for the first time the presence of enolase in the cell wall of both S. schenckii and S. brasiliensis. As a whole, our results suggest that enolase could be used as a potential antigenic target for vaccinal purposes against sporotrichosis.

Neuron-specific enolase and response to initial therapy are important prognostic factors in patients with small cell lung cancer

Purpose. The prognostic factors for the survival of small cell lung cancer (SCLC) patients are still widely debated. The aim of this study was to identify the clinical features and prognostic factors in SCLC patients. Methods. A retrospective study was conducted on SCLC patients who were treated in our hospital between July 2010 and July 2015. Comparison of overall survival (OS) was performed using the Kaplan-Meier method. Prognostic factors for OS were identified by multivariate Cox regression models. Results. A total of 523 patients with complete data and ECOG 0-2 were enrolled in our study. A total of 383 patients (73.2%) were diagnosed with ES-SCLC (extensive-stage SCLC) and 140 patients (26.8%) were diagnosed with LS-SCLC (limited-stage SCLC). In all patients, early disease stage, good ECOG, normal neuron-specific enolase (NSE), thoracic radiotherapy, ≥4 cycles of chemotherapy, prophylactic cranial irradiation, good response to initial therapy were independent favorable prognostic factors for OS, along with gender, age, CEA and CA125. In LS-SCLC patients, normal NSE, normal CEA, good response to initial therapy and surgery were independent favorable prognostic factors for OS. In ES-SCLC patients, good ECOG, normal NSE, thoracic radiotherapy, ≥4 cycles of chemotherapy, prophylactic cranial irradiation and good response to initial therapy were independent favorable prognostic factors for OS. Remarkably, NSE and response to initial therapy were independent prognostic factors for OS in all SCLC patients, LS-SCLC patients and ES-SCLC patients. Conclusion. The normal NSE and good response to initial therapy predicted a better survival for SCLC patients, regardless of disease stage (AU)

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A cell wall protein-based vaccine candidate induce protective immune response against Sporothrix schenckii infection.

Sporotrichosis is a subcutaneous mycosis caused by several closely related thermo-dimorphic fungi of the Sporothrix schenckii species complex, affecting humans and other mammals. In the last few years, new strategies have been proposed for controlling sporotrichosis owning to concerns about its growing incidence in humans, cats, and dogs in Brazil, as well as the toxicity and limited efficacy of conventional antifungal drugs. In this study, we assessed the immunogenicity and protective properties of two aluminum hydroxide (AH)-adsorbed S. schenckii cell wall protein (ssCWP)-based vaccine formulations in a mouse model of systemic S. schenckii infection. Fractioning by SDS-PAGE revealed nine protein bands, two of which were functionally characterized: a 44kDa peptide hydrolase and a 47kDa enolase, which was predicted to be an adhesin. Sera from immunized mice recognized the 47kDa enolase and another unidentified 71kDa protein, whereas serum from S. schenckii-infected mice recognized both these proteins plus another unidentified 9.4kDa protein. Furthermore, opsonization with the anti-ssCWP sera led to markedly increased phagocytosis and was able to strongly inhibit the fungus' adhesion to fibroblasts. Immunization with the higher-dose AH-adjuvanted formulation led to increased ex vivo release of IL-12, IFN-γ, IL-4, and IL-17, whereas only IL-12 and IFN-γ were induced by the higher-dose non-adjuvanted formulation. Lastly, passive transference of the higher-dose AH-adjuvanted formulation's anti-ssCWP serum was able to afford in vivo protection in a subsequent challenge with S. schenckii, becoming a viable vaccine candidate for further testing.

Prophylactic Injection of Recombinant Alpha-Enolase Reduces Arthritis Severity in the Collagen-Induced Arthritis Mice Model.

OBJECTIVE: To evaluate the ability of the glycolytic enzyme alpha-enolase (ENO1) or its immunodominant peptide (pEP1) to reduce the severity of CIA in DBA/1 mice when injected in a prophylactic way. METHODS: Mice were treated with mouse ENO1 or pEP1 one day prior to collagen II immunization. Clinical assessment was evaluated using 4 parameters (global and articular scores, ankle thickness and weight). Titers of serum anti-ENO1, anti-cyclic citrullinated peptides (anti-CCP) and anti-CII (total IgG and IgG1/IgG2a isotypes) antibodies were measured by ELISA at different time-points. Disease activity was assessed by histological analysis of both anterior and hind paws at the end of experimentation. RESULTS: Prophylactic injection of 100 µg of ENO1 reduced severity of CIA. Serum levels of anti-CII antibodies were reduced in ENO1-treated mice. Concordantly, ENO1-treated mice joints presented less severe histological signs of arthritis. ENO1 did not induce a shift toward a Th2 response since IgG1/IgG2a ratio of anti-CII antibodies remained unchanged and IL-4 serum levels were similar to those measured in the control group. CONCLUSIONS: Pre-immunization with ENO1 or its immunodominant peptide pEP1 reduces CIA severity at the clinical, immunological and histological levels. Effects of pEP1 were less pronounced. This immunomodulatory effect is associated with a reduction in anti-CII antibodies production but is not due to a Th1/Th2 shift.

Oral delivery of Bacillus subtilis spore expressing enolase of Clonorchis sinensis in rat model: induce systemic and local mucosal immune responses and has no side effect on liver function.

Caused by the consumption of raw or undercooked freshwater fish containing infective metacercariae of Clonorchis sinensis, human clonorchiasis remains a major public health problem in China. In previous study, we had expressed enolase from C. sinensis (CsENO) on the surface of Bacillus subtilis spore and the recombinant spore induced a pronounced protection in terms of reduced worm burden and eggs per gram feces, suggesting B. subtilis spore as an ideal vehicle for antigen delivery by oral treatment and CsENO as a promising vaccine candidate against clonorchiasis. In the current study, we detected CsENO-specific IgG and IgA levels both in serum and in intestinal mucus from rats orally administrated with B. subtilis spore surface expressing CsENO by ELISA. Lysozyme levels in serum and in intestinal mucus were analyzed too. In addition, IgA-secreting cells in intestine epithelium of the rats were detected by immunohistochemistry assay. The intestinal villi lengths of duodenum, jejunum, and ileum were also measured. Rats orally treated with B. subtilis spore or normal saline were used as controls. Our results showed that, compared with the control groups, oral administration of B. subtilis spore expressing CsENO induced both systemic and local mucosal immune response. The recombinant spores also enhanced non-specific immune response in rats. The spores had no side effect on liver function. Moreover, it might facilitate food utilization and digestion of the rats. Our work will pave the way to clarify the involved mechanisms of protective efficacy elicited by B. subtilis spore expressing CsENO and encourage us to carry out more assessment trails of the oral treated spore to develop vaccine against clonorchiasis.

Α1-giardin based live heterologous vaccine protects against Giardia lamblia infection in a murine model.

Giardia lamblia is a leading protozoan cause of diarrheal disease worldwide, yet preventive medical strategies are not available. A crude veterinary vaccine has been licensed for cats and dogs, but no defined human vaccine is available. We tested the vaccine potential of three conserved antigens previously identified in human and murine giardiasis, α1-giardin, α-enolase, and ornithine carbamoyl transferase, in a murine model of G. lamblia infection. Live recombinant attenuated Salmonella enterica Serovar Typhimurium vaccine strains were constructed that stably expressed each antigen, maintained colonization capacity, and sustained total attenuation in the host. Oral administration of the vaccine strains induced antigen-specific serum IgG, particularly IgG(2A), and mucosal IgA for α1-giardin and α-enolase, but not for ornithine carbamoyl transferase. Immunization with the α1-giardin vaccine induced significant protection against subsequent G. lamblia challenge, which was further enhanced by boosting with cholera toxin or sublingual α1-giardin administration. The α-enolase vaccine afforded no protection. Analysis of α1-giardin from divergent assemblage A and B isolates of G. lamblia revealed >97% amino acid sequence conservation and immunological cross-reactivity, further supporting the potential utility of this antigen in vaccine development. Together. These results indicate that α1-giardin is a suitable candidate antigen for a vaccine against giardiasis.

Immunisation with the glycolytic enzyme enolase confers effective protection against Candida albicans infection in mice.

Candida albicans is an opportunistic human fungal pathogen that continues to be a leading cause of candidal infections in immunocompromised hosts. Enolase, an important glycolytic enzyme located on the cell wall of C. albicans, was cloned, purified, and characterized by molecular cloning, affinity chromatography and Western blotting. C57BL/6J mice were immunized with recombinant enolase subcutaneously every two weeks, and the protective effect against systemic challenge evaluated by fungal burdens in target organs, titres of specific antibodies to enolase, and by levels of Th1/2 cytokines in serum. After challenge with C. albicans strains SC5314 and 3630, fungal burdens in the liver, kidney, brain, spleen and lung were significantly decreased in immunized mice. Histopathological assessment demonstrated that enolase protected the tissue structure, and decreased the infiltration of inflammatory cells. The titres of enolase-specific IgG1 and IgG2a in the immune serum reached up to 1:51200. Furthermore, opsonization with immune serum resulted in enhanced killing of both 3630 and SC5314 by murine neutrophils. Levels of IL-12 and IL-8 in the immune serum increased, whereas the concentration of the Th2 cytokine, IL-10, was significantly higher in immunized mice compared to the control group. It was concluded that recombinant enolase effectively protected mice against disseminated candidiasis, and may be a promising target for vaccination against different strains of C. albicans.

Molecular mimicry between neurons and an intracerebral pathogen induces a CD8 T cell-mediated autoimmune disease.

To identify basic mechanisms of how infections may induce a neuron-specific autoimmune response, we generated mice expressing OVA as neuronal autoantigen under control of the neuron-specific enolase promoter (NSE-OVA mice). Intracerebral, but not systemic, infection with attenuated Listeria monocytogenes-secreting OVA induced an atactic-paretic neurological syndrome in NSE-OVA mice after bacterial clearance from the brain, whereas wild-type mice remained healthy. Immunization with attenuated Listeria monocytogenes-secreting OVA before intracerebral infection strongly increased the number of intracerebral OVA-specific CD8 T cells aggravating neurological disease. T cell depletion and adoptive transfer experiments identified CD8 T cells as decisive mediators of the autoimmune disease. Importantly, NSE-OVA mice having received OVA-specific TCR transgenic CD8 T cells developed an accelerated, more severe, and extended neurological disease. Adoptively transferred pathogenic CD8 T cells specifically homed to OVA-expressing MHC class I(+) neurons and, corresponding to the clinical symptoms, approximately 30% of neurons in the anterior horn of the spinal cord became apoptotic. Thus, molecular mimicry between a pathogen and neurons can induce a CD8 T cell-mediated neurological disease, with its severity being influenced by the frequency of specific CD8 T cells, and its induction, but not its symptomatic phase, requiring the intracerebral presence of the pathogen.

Immunogenicity and protective effect of recombinant enolase of Candida albicans in a murine model of systemic candidiasis.

Enolase, a 46-kDa glycolytic enzyme, is an immunodominant antigen of the opportunistic pathogen Candida albicans. A recombinant 6 x His-tagged enolase was studied, in conjunction with interleukin-12 (IL-12), as an adjuvant for cytokine induction favouring protection in a murine model of haematogenous candidiasis. Mice immunized with enolase plus IL-12 showed increased antibody titres against enolase, as well as increased median survival time and decreased fungal burden in kidneys, in comparison to non-immunized or IL-12-treated mice. This increased survival was attributable to enolase-induced cell-mediated immunity as it also occurred in B-cell-deficient mice. Enolase immunization stimulated a predominant T-helper-1 (Th1) cytokine pattern in splenic cells and induced production of interferon-gamma (IFN-gamma) and interleukin-2 (IL-2) by purified CD4+ T cells. However, despite the elevation of immunogenicity, recombinant enolase induced only a modest protection against disseminated candidiasis, suggesting a form of protection likely attributable to the induction of a Th1 cell-mediated immune response.

La afectación ganglionar en los carcinomas escamosos de pulmón.Interés de la proteína oncogénica erbB2 y el Cyfra 21.1 / Lymph nodes involvement from lung squamous carcinomas. Interest of erbB2 oncogenic protein and Cyfra 21.1

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Inhalant allergies to fungi: reactions to bakers' yeast (Saccharomyces cerevisiae) and identification of bakers' yeast enolase as an important allergen.

Forty-seven subjects diagnosed as having inhalant allergies to fungi were tested for allergic sensitivity to bakers' yeast. Skin prick tests with yeast extract showed that 35 subjects responded with wheal reactions that were at least 3 mm while 32 subjects were regarded as clearly RAST-positive to bakers' yeast antigens. Skin and RAST testing with purified enolase from bakers' yeast and comparisons with the whole yeast extract showed that the enzyme is a major allergenic component of the extract. This conclusion was supported by results of electroblotting studies. RAST inhibition experiments demonstrated allergenic cross-reactivity between bakers' yeast, bakers' yeast enolase and Candida albicans.