Improvement of the T-cell response to a non immunogenic peptide by its tandem association with a highly efficient T-helper peptide.

The 45-69 peptide, an helper T-cell epitope derived from the HIV nef protein is strongly immunogenic. A T-cell proliferative response was observed following immunization of Lou/M rats with 45-69 peptide administered in low dose and without any adjuvant. It is already known that the T-cell response to the 115-131 peptide of Sm28GST antigen, a protein of the parasite Schistosoma mansoni, requires the presence of a carrier of the use of peptidic constructs. We demonstrate here that a T-cell response against the 115-131 peptide can be obtained in the absence of adjuvant using peptidic constructs (115-45 and 45-115 peptides) resulting from tandem synthesis of 115-131 and 45-69 peptides. A covalent association of both peptides is necessary, since the coinjection of 45-69 and 115-131 peptides is not sufficient to induce a detectable anti-115-131 T-cell response. The mutual orientation between the respective tandem peptides (45-115 and 115-45) is critical for the T-cell response. These peptidic constructs possess distinct properties of antigenicity and immunogenicity but both allowed to reveal the existence of a specific T-cell response normally undetectable using 115-131 peptide alone. This immunopharmacological approach should be useful in the rational design and construction of vaccines.

Effect of a lipopeptidic formulation on macrophage activation and peptide presentation to T cells.

We studied a 45-69 lipopeptide obtained by N-terminal modification with a N epsilon-palmitoyl lysine residue of the 45-69 peptide derived from the nef protein of HIV. T cells from animals immunized intraperitoneally with 45-69 lipopeptide proliferated in vitro in the presence of 45-69 peptide while no response was obtained after intraperitoneal immunization with 45-69 peptide. The efficiency of the 45-69 lipopeptide is supported by the covalent association to the N epsilon-palmitoyl lysine moiety. The immunogenicity of the 45-69 lipopeptide or of the unmodified peptide is dependent on the route of immunization but is not related to a mitogenic effect on cells or to an increase of the peptide antigenicity. Moreover, only 45-69 lipopeptide induces the secretion of cytokines such as IL-1, IL-6 and TNF-alpha by peritoneal macrophages. Finally, the use of 45-69 lipopeptide permits the activation of highly purified T cells without the addition of antigen-presenting cells. These results have implications for the formulation of synthetic vaccines.

Improvement of the T-cell response to a non-immunogenic peptide by its tandem association with a highly efficient T-helper peptide.

The 45-69 peptide, an helper T-cell epitope derived from HIV nef protein, is strongly immunogenic. A T-cell proliferative response was observed following immunization of Lou/M rats with 45-69 peptide administered in low dose and without any adjuvant. It is already known that the T-cell response to the 115-131 peptide of Sm28GST antigen, a protein of the parasite Schistosoma mansoni, requires the presence of a carrier or the use of peptidic constructs. We demonstrate here that a T-cell response against the 115-131 peptide can be obtained in the absence of adjuvant using peptidic constructs (115-45 and 45-115 peptides) resulting from tandem synthesis of 115-131 and 45-69 peptides. A covalent association of both peptides is necessary, since the co-injection of 45-69 and 115-131 peptides is not sufficient to induce a detectable anti-115-131 T-cell response. The mutual orientation between the respective tandem peptides (45-115 and 115-45) is critical for the T-cell response. These peptidic constructs possess distinct properties of antigenicity and immunogenicity but both allowed to reveal the existence of a 115-131 specific T-cell response normally undetectable using 115-131 peptide alone. This immunopharmacological approach should be useful in the rational design and construction of vaccines.

La detección de IgA para diagnosticar las infecciones retrovíricas en el Caribe / IgA testing for diagnosis of retroviral infections in the Caribbean

En la isla caribeña de Guadalupe, donde hay una alta prevalencia de infección por el virus linfotrópico T humano del tipo 1 (HTLV-1), los métodos de diagnóstico dan lugar a muchos resultados positivos falsos que hay que verificar mediante pruebas adicionales largas y costosas. En el presente artículo se describe una manera simple de solucionar este problema. Consiste, primero, en extraer del suero de un paciente toda la IgG y sus inmunocomplejos circulantes (ICC) mediante un tratamiento con proteína G, para después detectar la presencia de IgA antivírica en el suero tratado usando las pruebas comerciales de ELISA y Western Blot con ligeras modificaciones

La detección de IgA para diagnosticar las infecciones retrovíricas en el Caribe / IgA testing for diagnosis of retroviral infections in the Caribbean

En la isla caribeña de Guadalupe, donde hay una alta prevalencia de infección por el virus linfotrópico T humano del tipo 1 (HTLV-1), los métodos de diagnóstico dan lugar a muchos resultados positivos falsos que hay que verificar mediante pruebas adicionales largas y costosas. En el presente artículo se describe una manera simple de solucionar este problema. Consiste, primero, en extraer del suero de un paciente toda la IgG y sus inmunocomplejos circulantes (ICC) mediante un tratamiento con proteína G, para después detectar la presencia de IgA antivírica en el suero tratado usando las pruebas comerciales de ELISA y Western Blot con ligeras modificaciones

Disponible en inglés en Bull Pan Am. Health Organ 25(3):201-06, 1991

IgA testing for diagnosis of retroviral infections in the Caribbean.

In Guadeloupe, a Caribbean island with a high prevalence of HTLV-I infected subjects, the percentage of false positive results for HIV IgG is high, requiring additional time and expense for confirmatory tests. This article describes a simple way of overcoming this problem. First, all IgG and IgG immune complexes are removed by protein G treatment, and then IgA enzyme-linked immunosorbent assay (ELISA) and the Western blot test are performed with minor modification of commercially available kits.

PIP: Researchers wanted to determine whether they could do a more specific ELISA HIV test directed at IgA rather than IgG antibodies. So they 1st used streptococcal protein G to remove IgG from serum samples of 10 follow up patients from Guadeloupe who previously tested positive via ELISA for HIV-1/2 IgG and HTLV-I IgG. They did this to test sera for IgA. The protein G treatment resulted in a 99.9% reduction in the IgG concentration (12.21.7 mg/ml to .007-.017 mg/ml). After IgG depletion, the researchers noted no ELISA positive samples for HIV-1/2 or HTLV-I IgG. In fact, all the values were well below the cutoff levels. This result indicated that streptococcal protein G treatment followed by IgA ELISA is more specific than testing for IgG. The sera of 8 patients tested negative for HIV-1/2 IgA, but all tested positive for HTLV-1 IgA. This result showed that IgA may be more useful than IgG for also confirming retroviral infections. The sera of 2 patients were 44% and 27% above the cutoff level indicating that these patients had HIV-1/2 IgA antibodies. The researchers observed that protein G treatment suppressed all HIV-1/2 and HTLV-I IgG Western blot reactivity which confirmed the effectiveness of protein G treatment. Further the HIV IgG bands in the 8 false positive sera did not materialize at all suggesting that the IgG reactivity observed was most likely nonspecific. These results indicated that nonspecific HIV-1/2 IgG caused the false positive HIV-1/2 IgG ELISA results. In conclusion, in Caribbean countries where HTLV-1 infection can be as high as 13%, the more specific HIV IgA ELISA test should be used since its use saves time and money. Further it can do generalized testing for HTLV antibodies.

IgA testing for diagnosis of retroviral infections in the Caribbean

In Guadeloupe, a Caribbean island with a high prevalence of HTLV-I infected subjects, the percentage of false positive results for HIV IgG is high, requiring additional time and expense for confirmatory tests. This article describes a simple way of overcoming this problem. First, all IgG and IgG immune complexes are removed by protein G treatment, and then IgA enzyme-linked immudosorbent assay (ELISA) and the Western blot test are performed with minor modification of commercially available kits

IgA testing for diagnosis of retroviral infections in the Caribbean

In Guadeloupe, a Caribbean island with a high prevalence of HTLV-I infected subjects, the percentage of false positive results for HIV IgG is high, requiring additional time and expense for confirmatory tests. This article describes a simple way of overcoming this problem. First, all IgG and IgG immune complexes are removed by protein G treatment, and then IgA enzyme-linked immudosorbent assay (ELISA) and the Western blot test are performed with minor modification of commercially available kits

Available in spanish in Bol. Oficina Sanit. Panam 112(1):12-18, 1992

[Antigens of Toxoplasma gondii with a diagnostic and potential immunoprophylactic value: new strategies of identification]. / Antigènes de Toxoplasma gondii d'intérêt diagnostique et immunoprophylactique potentiel: nouvelles stratégies d'identification.

Toxoplasma gondii is an ubiquitous protozoan parasite which induces severe pathology in in utero infected children and in immunosuppressed patients (particularly in the case of AIDS). Previous work that focused on toxoplasma somatic antigens failed to demonstrate an efficient protection against highly virulent T. gondii strains. The authors therefore first studied the role of parasite excreted-secreted (ES) antigens in the immune response. They describe here the preparation of excreted-secreted antigens in cell-free medium from tachyzoites, the intracellular proliferative stage present during acute infection. Major ES antigens have Mr of 108 K, 97 K, 86 K, 57 K, 42 K, 39 K, 28.5 K, 27 K and 21 K. The protective role of ES antigens has been demonstrated using congenitally athymic (Nu/Nu) rats that are highly sensitive to T. gondii infection (+/+ Fischer rats are resistant). The humoral and cellular components of this protection have been studied by the passive transfer either of sera or of T lymphocytes from ES-immunized +/+ Fischer rats into Nu/Nu rats. Adoptive transfers were carried out 24 hours before infection with the highly virulent T. gondii RH strain. Based on the concept of concomitant immunity, the authors have characterized antigens from tachyzoites and bradyzoites (the encysted stage persisting during chronic infection) which share common epitopes. Four tachyzoite antigens, P63, GP43, P39 and GP 28.5 have been shown by immunoprecipitation to cross-react with bradyzoite antigens. Two monoclonal antibodies raised against ES antigens permitted to demonstrate the localization of the 28.5 K and 27 K antigens inside the dense granules of tachyzoites and bradyzoites.(ABSTRACT TRUNCATED AT 250 WORDS)

Protective role of IgE in immunocompromised rat toxoplasmosis.

In contrast to euthymic adult Fischer rats, immunocompromised Nu/Nu animals develop a lethal infection when inoculated with the RH strain of the protozoan Toxoplasma gondii. However, a significant period of survival is obtained when Nu/Nu rats are passively transferred with sera from 28-day infected Fischer +/+ (euthymic) animals. Specific IgE are involved since IgE-depleted sera are unable to afford such a protection. Only excreted/secreted Ag or living tachyzoites are able to induce a significant protective IgE response in intact animals. In addition, platelets or, to a lesser extent, eosinophil-rich populations from Toxoplasma infected or excreted-secreted Ag-immunized euthymic animals bear surface IgE and are cytotoxic for the parasite in vitro. Also, adoptive transfer of immune platelets confers a significant degree of protection to Toxoplasma-infected Nu/Nu animals. Our results clearly show the key role of Ag present in both living parasites and excreted-secreted Ag to induce, in this model, a protective IgE response. In addition, as in other parasitic infections, platelets and probably eosinophils are the effector cells involved in controlling parasitic dissemination during Toxoplasma infection in immunocompromised rats.

Killer cells in human cutaneous leishmaniasis.

In French Guiana, American cutaneous leishmaniasis is localized in the skin. The host response appears to be effective since few extra- or intracellular organisms can be found in tissue lesions, and we never observed any cutaneous dissemination or visceral involvement. However, this response is not fully effective since lesions may last for months. By using immunoperoxidase techniques and monoclonal antibodies directed against various cell populations, we examined the local immune response in skin biopsies. We found a high percentage of cells with the K/NK phenotype, a variable but usually high percentage of cells with the T cell phenotype bearing TAC receptors, and moderate numbers of monocytes and B cells. These results suggest that K/NK cells could play a role in the local control of parasite dissemination.

[Changes in immunologic and biochemical functions after a mission to a tropical forest]. / Modification des fonctions immunologiques et biochimiques après une mission en forêt tropicale.

A biochemical and immunological survey of a selected military group has been performed, before and after a jungle raid. Biochemical (total proteins, albumin, proteins electrophoresis, transferrin, haptoglobin, ceruloplasmin, orosomucoid, alpha-1 antitrypsin and alpha-2 macroglobin) and immunological (Ig G., Ig A., C 3c, C 4, specific antibodies against Leishmania) analysis of sera have been studied as well as the level of cellular immunity (Multitest scoring, macrophage cytotoxicity, interleukin-1 and prostaglandin E-2, macrophage production). The results show some modifications as a consequence of the work in tropical forest, particularly the tendency to some cellular immunity deficiency. This can be linked to several factors: nutritional balance, cutaneous infections, sustained physical activity and excessive sweating.

Macrophage function in patients with American cutaneous leishmaniasis: in vitro cytotoxicity and interleukin-1 production.

Monocyte-derived blood macrophages of untreated patients with Leishmania braziliensis or Leishmania mexicana amazonensis infections show anomalies in their nonspecific immune functions. Their ability to kill HeLa cells or to produce interleukin-1 in vitro in response to lipopolysaccharide plus Candida albicans is lower than controls indicating that acquired or innate macrophage deficiencies may be involved in the course of the disease.

Interleukin-1 released by blood-monocyte-derived macrophages from patients with leprosy.

In highly purified blood-monocyte-derived macrophages collected from patients with leprosy and from healthy individuals and cultured in vitro with mycobacterial antigens such as Mycobacterium bovis BCG or Mycobacterium leprae, we nonspecifically induced the synthesis of interleukin-1. Normally, all supernatants from cultured macrophages of all subjects tested produced similar amounts of interleukin-1. However, only in patients with lepromatous leprosy, M. leprae, but not BCG, induced high-level synthesis of prostaglandin E2, which acted as a suppressor factor in the mouse thymocyte proliferative assay used to measure the interleukin-1 content of the supernatants. Normal interleukin-1 content of those supernatants was demonstrated by blocking the prostaglandin E2 synthesis by the addition of indomethacin to the medium throughout the experimental procedure. We also tested the efficiency of a combination of BCG and M. leprae in reducing the prostaglandin E2 synthesis, but with the methodology used, we did not observe any beneficial effect of such a combination. These results demonstrate the possible role of M. leprae in the induction of at least one of the suppressive monokines and are additional arguments for the involvement of macrophages in the suppression of the specific cell-mediated immunity to M. leprae observed in lepromatous leprosy.

Cell-mediated immunity in mice treated with Mycobacterium leprae or with macrophages harbouring M. leprae.

Following treatment of BALB/c or C3H/HeN mice in the hind footpads with irradiated Mycobacterium leprae, a marked enhancement of natural killer (NK) activity was observed in cells from the draining popliteal lymph node or from the spleen. NK activity was further enhanced when the treatment consisted of killed M. leprae which had been incorporated into mouse peritoneal macrophages. This effect was noted as early as 2 weeks after treatment and persisted for at least 9 weeks. Lymphoblastic transformation in response to suboptimal doses of the T-cell mitogen transformation in response to suboptimal doses of the T-cell mitogen concanavalin A or to M. leprae antigen was assayed in parallel in cells from the draining popliteal lymph node and from the spleen. In contrast to NK assays, treatment with M. leprae alone moderately altered the response to mitogen. However, there was a prominent enhancement of the T-cell response when treatment consisted of M. leprae-laden macrophages.

Detection of circulating antigens in onchocerciasis.

This report describes the presence of circulating Onchocerca volvulus antigens (COA) in sera of patients with onchocerciasis. By using a double diffusion immunoelectrophoresis method, COA could be detected in 24 of 77 sera analyzed (31%). In contrast, when more sensitive assays such as the radioimmunoprecipitation-PEG assay or sandwich radioimmunoassay were used to detect COA, about 75% of the sera from O. volvulus-infected patients were found positive; moreover, a highly significant correlation between the two assays was observed. The parasite specificity of the COA was demonstrated directly by identity reaction with a component of O. volvulus somatic antigens. COA was never found when hyperimmune antisera against other parasite antigenic extracts were used instead of anti-O. volvulus hyperimmune serum. However, when anti-O, volvulus hyperimmune serum was used against sera obtained from patients infected with various other helminths we found a cross-reactivity between COA and the circulating antigens of other human filarids (Wuchereria bancrofti, Loa loa, Brugia malayi), but not with other nematode or trematode parasites (Ascaris lumbricoides, Schistosoma mansoni, Fasciola hepatica). Further immunoelectrophoretic studies demonstrated one precipitin are localized in the cathodic region which seemed specific for COA, which raises the possibility of preparing a monospecific hyperimmune serum to circumvent cross-reactivities.