[Comparison of adjuvant activities of glucosaminyl-muramyl dipeptide and of the gene coding for granulocyte-macrophage colony-stimulating factor in DNA immunization against herpes simplex virus].

Adjuvant activities of granulocyte-macrophage colony-stimulating factor (GM-CSF) and synthetic glucosaminyl-muramyl dipeptide (GMDP) were studied in immunization against type 1 herpes simplex virus (HSV1). Gene encoding the gD HSV1 protein (pDNAgD) was used as an immunogen. Gene encoding GM-CSF in pDNAGM-CSF plasmid, which was developed for eukaryotic expression, and GM-DP were used as immune response modulators. GMDP and plasmid DNA with inserted GM-CSF gene enhanced T-cell immune response to HSV1 after a single injection (pDNAGM-CSF) or 24 h before (GMDP) immunization with the gD HSV1 gene. Both adjuvants increased protective effect of DNA-immunization by a virus gene with 63 up to 100% after injection of two genes and up to 96% after the viral gene was inoculated 24 h after GMDP. These high effects indicate that further investigation of anti-HSV1 DNA-based vaccines used with genetic and peptide adjuvant is prospective.

[New acellular pertussis vaccine, containing glucosaminylmuramyldipeptide]. / Sozdanie novoi beskletochnoi kokliushnoi vaktsiny, vkliuchaiushchei gliukozaminilmuramildipeptid.

As shown in this work, the synthetic immunomodulator glucosaminylmuramyldipeptide (GMDP) can be included into acellular pertussis vaccine (APV). The optimal doses of GMDP, ranging from 0.001 to 0.0001 microg, have been found. These doses enhance the protective activity of APV, especially its low-active doses. GMDP decrease the manifestations of toxic, anaphylactogenic and pyrogenic properties of APV, which may lead to the decrease of the antigenic load of APV on the body of the vaccines and thus to lessening the side-effects of vaccination. GMDP has been shown to considerably increase, in comparison with common pertussis vaccine and APV, the percentage of phagocytizing leukocytes by day 14. The immunization of mice with APV with and without GMDP in doses of 0.01 and 0.001 microg leads to a change in T-lymphocyte/B-lymphocyte ratio in the population of spleen lymphocytes.

Immune response modulation by recombinant peptides expressed in virus-like particles.

Aspergillus fumigatus, a ubiquitous fungus, is implicated in the pathogenesis of a number of clinically different allergic diseases in man, including allergic bronchopulmonary aspergillosis. Peptide-based immunotherapy may offer an alternative treatment strategy for the management of allergic disease. The objective of this study was to alter the allergen-specific immune response using dominant T cell epitopes of a major A. fumigatus allergen, Asp f2, expressed in yeast as virus-like particles (VLP). The T cell epitopes of Asp f2, recognized in mice with an H-2d background, were determined by producing T-cell hybridomas. Two dominant T cell epitopes, aa60--71 and aa235--249, were identified and expressed in a yeast VLP system. To induce tolerance VLP-peptides were injected subcutaneously into mice previously immunized with recombinant Asp f2. The T cell immune response was abrogated totally in 3 weeks following a single injection of VLP but was restored 2 months later following intranasal antigen exposure. T-cell depletion resulted in the reduction of 20-30% of all antigen-specific immunoglobulin classes. Thus, recombinant peptides expressed in the VLP system can be used successfully in the modulation of Asp f2-induced immune response in mice, although a single administration is not sufficient to maintain a state of tolerance for a long period of time.

Endogenous tumour necrosis factor-alpha sensitise melanoma cells to glucosaminylmuramyl dipeptide.

Flow cytometry was used to demonstrate that cultured human melanoma BRO cells expressed membrane-bound tumour necrosis factor-alpha (TNF-alpha) and were able to release TNF-alpha upon treatment with glucosaminylmuramyl dipeptide (GMDP). The released TNF-alpha was shown to prime melanoma cells, previously unable to respond to GMDP by increasing expression of melanoma-associated antigens, making them sensitive to GMDP treatment.

Biochemical characterization of glucosaminylmuramyldipeptide binding sites of murine macrophages.

By using radioligand analysis, murine peritoneal macrophages were shown to express several hundred high-affinity cell surface GMDP-binding sites (Ka 350 pM). Photoaffinity labeling followed by SDS-PAGE enabled us to identify 32-34 and 38 kDa proteins inside these cells that bound GMDP specifically.

Dituftsin and polytuftsin induce an anti-peptide IgG response to non-immunogenic peptides in mice.

The effect of covalently attaching multiple forms of the immunomodulating tetrapeptide tuftsin to normally non-immunogenic peptides was studied in BALB/c and C57Bl/6 mice. The peptides: (NANP)3 from the Plasmodium falciparum circumsporozoite protein and peptides 136-152 and 205-213 derived from the capsid protein of foot-and-mouth disease virus were coupled to polytuftsin or dituftsin. Anti-peptide IgG titers were determined after two immunizations. All of these three non-immunogenic peptides coupled to polytuftsin or dituftsin induced anti-peptide antibody production in mice while peptides alone did not elicit IgG. In addition, a conjugate of (NANP)3 and polytuftsin with built-in glycopeptide adjuvant elicited an anti-peptide response comparable in magnitude with that of a peptide-KLH conjugate. The data suggest that when non-immunogenic peptides are synthesized in tandem with dituftsin or conjugated to polytuftsin a significant immune response to the peptides may be elicited. This approach may be employed in synthetic vaccine design.

[The effect of the immunomodulator glucosaminylmuramyl dipeptide on hematopoiesis in mice with experimental cytopenia]. / Vliianie immunomoduliatora gliukozaminilmuramildipeptida na krovetvorenie myshei s éksperimental'noi tsitopeniei.

Simulation of cytopenia by injection of cyclophosphamide (100 mg/kg) or by exposure to ionizing radiation (4 Gy) was shown to cause in mice similar, by the severity and rate of development, transient inhibition of haemopoiesis which was somewhat more persistent after irradiation. Glucosaminylmuramyl dipeptide applied after the above effects promoted the haemopoiesis recovery.

[Activation of cellular immunity in mice under normal conditions and in tumor growth during treatment with glucosaminyl muramyl dipeptide]. / Aktivatsiia kletochnogo immuniteta u myshei v norme i pri opukholevom roste pod deistviem gliukozaminilmuramildipeptida.

In vitro stimulation of mice spleen cells by means of glucosaminyl muramyl dipeptide (GMDP) was accompanied by development of tumor necrosis factor and of interleukin-I. The factor was detected in blood serum only after administration of GMDP simultaneously with lipopolysaccharide. GMDP activated peritoneal macrophages; the phenomenon was evaluated by means of the macrophages ability to kill tumoral cells P815 as well as by interleukin-I production after additional stimulation with lipopolysaccharide. At the same time, an increase in proliferating activity of spleen and bone marrow cells was observed. An increase of middle lifetime and recovery of 24% mice of C57BL/6 strain with leukosis EL-4 were observed after complex treatment of the animals with GMDP, lipopolysaccharide, cyclophosphane and indomethacin.

[Immunostimulating effects of muramyl dipeptide, glucosaminyl muramyl dipeptide and their synthetic derivatives in vitro]. / Immunostimuliruiushchee deistvie muramildipeptida, gliuko- zaminilmuramildipeptida i ikh sinteticheskikh proizvodnykh v sisteme in vitro.

The effect of muramyldipeptide (MDP), glucosaminylmuramyldipeptide (GMDP) and their six synthetic derivatives on production of tumor necrosis factor (TNF), interleukin-1 (IL-1) and interleukin-2 (IL-2) by murine spleen cells in vitro was studied. MDP induced insignificant TNF production and did not stimulate production of IL-1 by the murine splenocytes within a 24-hour cultivation period whereas in combination with lipopolysaccharide (LPS) it induced significant production of both the cytokins. GMDP induced marked production of TNF (54 per cent cytotoxic index) and IL-1 (stimulation index 8). Addition of LPS in an amount of 10 ng/ml increased production of TNF by the murine splenocytes under the effect of GMDP but had no effect on production of IL-1. Neither MDP nor GMDP even in combination with LPS induced production of IL-2 by splenocytes of mice DVA/2 and C57B1/6 at activation for 24 hours. All the synthetic derivatives of MDP and GMDP except the MDP polymer activated TNF production by the murine spleen cells. GMDP lysine had the highest effect: 67 per cent cytotoxic index. In combination with LPS its cytotoxic index amounted to 87 per cent. The TNF activity was always higher when LPS in an amount of 10 ng/ml was added to the glycopeptides.

Glucosaminylmuramyl dipeptide-induced changes in murine macrophage metabolism.

The purpose of the study was to investigate influence of glucosaminyl muramyl dipeptide (GMDP) and its derivatives on adenosine deaminase (ADA, CE 3.5.4.4.) and 5'-nucleotidase (5-N, CE 3.1.3.5) activity in murine macrophages in vitro. The intensity of superoxide radicals (O2-) formation by these cells has been also studied. GMDP incubated with macrophages was found to inhibit substantially the activity of 5-N, without affecting the activity of ADA in these cells. The maximal effect on 5-N activity was noted following 24 h of co-culture and was accompanied by a higher intensity of O2- formation. GMDP added in doses ranging from 0.01 to 1 microgram/ml induced a gradual decrease in 5-N activity, with an increase in activity of the O2- -generating system. The GMDP analog with double dipeptide link GM(DP)2 has demonstrated the same activating effect as GMDP. The presence of dipeptide alanyl-D-isoglutamine in the GMDP structure is necessary for realization of the drug activating effect, as N-acetylglucosaminyl-N-acetylmuramyl failed to influence macrophage activity. Neither D-D nor L-L isomers of the drug affect the 5-N activity and O2- formation in macrophages. The mechanism of macrophages activation induced by GMDP may include the inhibition of 5-N activity and the stimulation in production of superoxide radicals.