Cyclophosphamide-immunosuppressed Mice Research Articles

Shrimp peptide hydrolysate modulates the immune response in cyclophosphamide immunosuppressed mice model.

Bioactive peptides are naturally found in various foods and were shown to have various distinct physiological as well as medicinal benefits. In this study shrimp peptide hydrolysate (SPH) was prepared to investigate its immunomodulatory effect against cyclophosphamide (CTX) induced immunosuppressed mice. The SPH effect was also analyzed on murine macrophage (RAW264.7 cells). The findings show that SPH stimulates macrophages to form multiple pseudopodia, has no cytotoxic effect, and increases phagocytic activity in RAW264.7 cells. Furthermore, the immunosuppressed in-vivo model illustrates the improvement in various aspects, that is body weight, escalation in immune organ index, and ameliorates histopathological transformation of thymus along with the spleen. SPH enhances cell-mediated immunity by facilitating splenocyte proliferation and inhibit excessive apoptosis. Moreover, the significant outcome had been observed with the upregulation of cytokines interferon-gamma (IFN-ϒ), interleukin-2 (IL-2) level and simultaneously downregulate certain genes include interleukin-4 (IL-4) and interleukin-10 (IL-10). Additionally, SPH expedites cellular immunity by enhancing the regulation of immunoglobulin A (IgA) and immunoglobulin M (IgM). However, these findings support the hypothesis that SPH is an effective immunomodulatory agent capable of preventing immune system hypofunction. It is necessary to investigate the detailed mechanism to rule out any unforeseen effects of SPH in future research. PRACTICAL APPLICATIONS: Chemotherapy medications, despite their dominating detrimental effects of damaging immunological organs such as the spleen and thymus, extend the treatment process as well as the destruction of the self-immune system. This study found that SPH is an effective immunomodulatory agent capable of avoiding immune organ hypofunction and improving cell mediate immunity by enhancing macrophage activation, phagocytosis, spleenocyte proliferation, suppressing apoptosis, and elevating cytokines and antibodies. As a result, SPH can be utilized as a nutritional and functional dietary supplement to boost immunological modulation in combination with chemotherapy medications in order to lessen their adverse effects.

Solidified Reverse Micellar Solution- (SRMS-) Based Microparticles for Enhanced Oral Bioavailability and Systemic Antifungal Efficacy of Miconazole Nitrate in Immunocompromised Mice.

Purpose To assess the improvement in oral bioavailability and efficacy in systemic candidiasis treatment of miconazole nitrate (MN) formulations in murine models of candidiasis. Methods Selected formulations containing 5% of Softisan + Phospholipon 90H lipid matrix with 3% of MN (A1), 5% of stearic acid + Phospholipon 90H lipid matrix with 3% of MN (B1), and 5% Softisan + stearic acid + Phospholipon 90H with 3% of MN (C1) from the in vitro investigation were used for the study. Their acute toxicity was assessed using Lorke's method (with slight modification) while bioavailability was determined using the bioassay method. The optimized batch (A1) was tested in murine systemic candidiasis induced in cyclophosphamide-immunosuppressed mice. The mice were treated with a single oral dose (100 mg/kg) of the formulations for five days. Serum fungal counts (cfu/mL) were determined on days 1, 3, and 5 of the treatment period. Haematological assessments were done. Results The lipid formulations were safer than MN powder with LD50 values of 3162.8 and 1118.3 mg/kg. Bioavailability determination revealed a higher area under the curve (AUC) value for formulations A1 (6.11 μg/hr/mL) and B1 (4.91 μg/hr/mL) while formulation C1 (1.80 μg/hr/mL) had a lower AUC than MN (4.46 μg/hr/mL). Fungi were completely cleared from the blood of animals treated with the optimized formulation by day 3 as opposed to the controls (MN and Tween® 20) which still had fungi on day 5. No significant increase (p > 0.05) in haematological parameters was observed in mice treated with A1. Conclusion Formulation A1 successfully cleared Candida albicans from the blood within a shorter period than miconazole powder. This research has shown the potential of orally administered MN-loaded SRMS-based microparticles in combating systemic candidaemia.

Improvement of Myelopoiesis in Cyclophosphamide-Immunosuppressed Mice by Oral Administration of Viable or Non-Viable Lactobacillus Strains.

Myelosuppression is the major dose-limiting toxicity of cancer chemotherapy. There have been many attempts to find new strategies that reduce myelosuppression. The dietary supplementation with lactic acid bacteria (LAB) improved respiratory innate immune response and the resistance against respiratory pathogens in immunosupressed hosts. Although LAB viability is an important factor in achieving optimal protective effects, non-viable LAB are capable of stimulating immunity. In this work, we studied the ability of oral preventive administration of viable and non-viable Lactobacillus rhamnosus CRL1505 or L. plantarum CRL1506 (Lr05, Lr05NV, Lp06V or Lp06NV, respectively) to minimize myelosuppressive and immunosuppressive effects derived from chemotherapy. Cyclophosphamide (Cy) impaired steady-state myelopoiesis in lactobacilli-treated and untreated control mice. Lr05V, Lr05NV and Lp06V treatments were the most effective to induce the early recovery of bone marrow (BM) tissue architecture, leukocytes, myeloid, pool mitotic and post-mitotic, peroxidase positive, and Gr-1Low/High cells in BM. We selected the CRL1505 strain for being the one capable of maintaining its myelopoiesis-enhancing properties in its non-viable form. Although the CRL1505 treatments do not modify the Cy ability to induce apoptosis, both increased the incorporation of BrdU in BM cells. Consequently, Lr05NV and Lr05V treatments were able to promote early recovery of LSK cells (Lin-Sca-1+c-Kit+ cells), multipotent progenitors (Lin-Sca-1+c-Kit+CD34+ cells), and myeloid cells (Gr-1+Ly6G+Ly6C- cells) with respect to the untreated Cy control. In addition, these treatments were able to increase the frequency of IL17A-producing innate lymphoid cells in the intestinal lamina propria (IL-17A+RORγt+CD4-NKp46+ cells) after Cy injection. These results were correlated with an increase in the IL-17A serum levels, a GM-CSF high expression and a CXCL12 lower expression in BM. Therefore, both Lr05V and Lr05NV treatments are able to activate beneficially the IL-17A/GM-CSF axis and accelerate the recovery of Cy-induced immunosuppression by increasing BM myeloid precursors. We demonstrated for the first time the beneficial effect of CRL1505 strain on myelopoiesis affected by a chemotherapeutic drug. Furthermore, Lr05NV could be a good and safe resource for reducing chemotherapy-induced leukopenia. The results are a starting point for future research and open up broad prospects for future applications of the immunobiotics.

Ma-Nuo-Xi decoction has an immunostimulating effect in cyclophosphamide-immunosuppressed mice

Ma-Nuo-Xi decoction (MNXD), as well as its hundreds of derivative preparations, has been used in Tibetan medicine since the 14th century. MNXD is in accordance with the theory of treatment determination based on syndrome differentiation. This study aimed to compare the effect of the auxiliary MNXD prescription (MNXD-AD) with that of the basic MNXD prescription (MNXD-BD) on the immunostimulating activity of MNXD. The immunopotentiation of MNXD, MNXD-BD, and MNXD-AD was evaluated using a cyclophosphamide (CTX)-immunosuppressed mouse model. Their influences on non-specific and specific immunity were evaluated using immune organ indexes, peripheral white blood cell (WBC) count, red blood cell (RBC) count, platelet count, phagocytosis, macrophage-secreted nitric oxide (NO) and cytokines, natural killer (NK) cytotoxic activity, lymphocyte proliferation, serum cytokines, splenic T-lymphocyte subpopulations, and quantitative hemolysis of sheep red blood cell (QHS SRBC) assays. MNXD, MNXD-BD, and MNXD-AD increased the spleen and thymus indexes, as well as the peripheral WBC, RBC, and platelet counts. They also promoted phagocytosis, NO and cytokine secretion from macrophages, NK cytotoxic activity, and lymphocyte proliferation, and also raised the CD4+ /CD8+ T-cell ratio, serum cytokine concentrations, and haemolysin formation in CTX-treated immunosuppressed mice. Compared with MNXD-BD and MNXD-AD, MNXD was superior in restoring the phagocytic index, concanavalin A (ConA)-induced T-lymphocyte proliferation, NO secretion from macrophages, and haemolysin formation, as well as the levels of interleukin 1 beta (IL-1β), and serum interleukin-2 (IL-2) and interferon gamma (INF-γ). MNXD, MNXD-BD, and MNXD-AD have excellent immunostimulating and myelosuppression-restoring activities on CTX-immunosuppressed mice. Among them, MNXD-AD might be an immunomodulator, which may happen to be in line with the clinical experience of Tibetan medicine physicians of using it to promote the efficacy of MNXD-BD.

Screening and structure study of active components of Astragalus polysaccharide for injection based on different molecular weights

As a special traditional Chinese medicine, Astragalus polysaccharides for injection (APS, batch no. Zhunzi Z20040086) includes complex polysaccharide macromolecules that may increase the risk upon application. Although fingerprints for quality control are available, the specific active ingredients are unclear. Identifying the active components is the key to reduce the risk of adverse reactions of the drug. In this work, APS was mainly separated into two components, namely, macromolecular component (APS-I) and small molecular components (APS-II). The molecular weight measurement revealed that the average molecular weight of APS-I exceeded 500 kDa, and that of APS-II was 10 kDa. Monosaccharide-composition analysis revealed that APS-I consisted of glucose, galactose, arabinose, rhamnose, and galacturonic acid, with a ratio of approximately 1.5:1:5.4:0.08:0.1. Meanwhile, APS-II consisted of glucose, galactose, arabinose, rhamnose, and galacturonic acid, with a molar ratio of 9:1:1.4:0.04:0.001. Methylation, FT-IR, and NMR analysis indicated that the APS-I monosaccharide residue was linked as follows: D-Glcp-(1→, →4)-D-Glcp-(1→, →2)-L-Rhap-(1→, D-Araf-(1→, →5)-D-Araf-(1→, →2,5)-D-Araf-(1→, →4)-D-Galp-(1 → . Meanwhile, the APS-II monosaccharide residue was connected as follows: α-D-Glcp-(1→, →4)-α-D-Glcp-(1→, →6)-α-D-Glcp-(1→, →4,6)-α-D-Glcp-(1→, →3,4,6)-α-D-Glcp-(1→, →2)-α-L-Rhap-(1→, α-D-Araf-(1→, →5)-α-D-Araf-(1→, →4)-β-D-Galp-(1 → . Screening experiments on their in vitro immunological activity showed that APS-II had stronger effect on innate and adaptive immunities than APS-I. In vivo animal experiments showed that APS-II can increase the leukocyte level of cyclophosphamide immunosuppressed mice and improve their immunomodulatory ability. Therefore, APS-II is the main active ingredient of APS and is expected to become a new generation of APS products.

Immunostimulating effect of polysaccharides isolated from Ma-Nuo-Xi decoction in cyclophosphamide-immunosuppressed mice

Ma-Nuo-Xi Decoction (MNXD) is well-known in Tibetan medicine to be in line with the theory of treatment determination based on syndrome differentiation. However, the components responsible for its immunomodulating effect are unknown. In this study, three polysaccharide components—MNXD-P, MNXD-BD-P, and MNXD-AD-P—were isolated from MNXD and its basic and auxiliary prescription decoctions, of which MNXD-BD-P is composed of β-(1,4)-d-glucan and RG-I pectin, MNXD-AD-P contains mainly α-(1,4)-d-glucan and some amount of arabinogalactan and/or arabinorhamnogalactan, and MNXD-P contains components of both MNXD-BD-P and MNXD-AD-P. And treatment with these polysaccharides could significantly improve the host's specific and non-specific immunity, including cellular and humoral immunities, as well as promote recovery from myelosuppression in cyclophosphamide (CTX)-immunosuppressed mice. To our knowledge, this is the first report on chemical and immunoactivity study on polysaccharides from traditional Tibetan medicine compounds, which may provide a new idea for development of carbohydrate drugs from them.

Successful Aspects of the Coadministration of Sterol 14α-Demethylase Inhibitor VFV and Benznidazole in Experimental Mouse Models of Chagas Disease Caused by the Drug-Resistant Strain of Trypanosoma cruzi.

Up to now, no vaccines are available for Chagas disease, and the current therapy is largely unsatisfactory. Novel imidazole-based scaffolds of protozoan sterol 14α-demethylase (CYP51) inhibitors have demonstrated potent antiparasitic activity with no acute toxicity. Presently our aim was to investigate the effectiveness of the experimental 14α-demethylase inhibitor VFV in the mouse models of Trypanosoma cruzi infection using a naturally drug-resistant Colombiana strain, under monotherapy and in association with the reference drug, benznidazole (Bz). The treatment with VFV resulted in complete parasitemia suppression and 100% animal survival when administered orally (given in 10% DMSO plus 5% Arabic gum) at 25 mg/kg (bid) for 60 days. However, as parasite relapse was found using VFV alone under this treatment scheme, the coadministration of VFV with Bz was assayed giving simultaneously (for 60 days, bid) by oral route, under two different drug vehicles (10% DMSO plus 5% Gum Arabic with or without 3% Tween 80). All tested mice groups resulted in >99.9% of parasitemia decrease and 100% animal survival. qPCR analysis performed on cyclophosphamide immunosuppressed mice revealed that, although presenting lack of cure, VFV given as monotherapy was 14-fold more active than Bz, and the coadministration of Bz plus VFV (given simultaneously, using 10% DMSO plus 5% Gum Arabic as vehicle) resulted in 106-fold lower blood parasitism as compared to the monotherapy of Bz. Another interesting finding was the parasitological cure in 70% of the animals treated with Bz and VFV when the coadministration was given using the VFV suspension in 10% DMSO + Arabic gum + Tween 80 (a formulation that we have found to provide a better pharmacokinetics), even after immunosuppression using cyclophosphamide cycles, supporting the promising aspect of the drug coadministration in improving the efficacy of therapeutic arsenal against T.cruzi.

Tarin stimulates granulocyte growth in bone marrow cell cultures and minimizes immunosuppression by cyclo-phosphamide in mice.

Chemotherapeutic drugs, such as cyclophosphamide, cause severe immunosuppression and patients become susceptible to infections. Based on this, the immunomodulatory potential of tarin, a lectin from Colocasia esculenta, was evaluated in bone marrow cell cultures and in cyclophosphamide-immunosuppressed mice. Tarin promoted maintenance of hematopoietic progenitors and repopulation of Gr1 cells in vitro which was supported by in vivo results. In immunosuppressed mice, tarin increased bone marrow cell numbers and altered cell profile distribution by enhancing the frequency of Gr1+ progenitors, including Ly6-CintLy6-Glo, and anticipating their proliferation/differentiation in mature cells, especially Ly6-CloLy6-Ghi. Bone marrow cells harvested from tarin-treated immunosuppressed mice proliferated in response to GM-CSF or G-CSF in vitro and, the low numbers of bone marrow cells in the G0 phase, combined with a high number cells undergoing apoptosis confirmed that tarin promoted a faster and intense proliferation/differentiation, even in the presence of CY-induced toxicity. As a result, tarin minimized leukopenia in immunosuppressed mice promoting a faster recovery of peripheral leucocytes and protected erythroid bone marrow cells from CY-cytotoxicity in a dose-dependent manner. Data suggest that tarin could be considered a potential adjuvant to decrease leukopenia and possibly ameliorate anemia, if carefully evaluated in human cancer cell lineages and in clinical trials.

Corticosteroids Impair Granulocyte Transfusion Therapy By Targeting NET Formation and Neutrophil Antifungal Functions Via ROS/Dectin1 Pathways

Introduction:Invasive pulmonary Aspergillus fumigatus infections cause high morbidity and mortality in neutropenic patients. Granulocyte transfusions have been tested as an alternative therapy for the management of high-risk neutropenic patients with invasive A. fumigatus infections. To increase the granulocyte yield for transfusion, donors are treated with corticosteroids. Yet, the efficacy of granulocyte transfusion and functional defense mechanisms of granulocytes collected from corticosteroid treated donors remain largely elusive. Therefore, we investigated the efficacy of granulocyte transfusion and functional defense mechanisms of corticosteroid treated granulocytes using mouse models.Methods:To determine the effects of corticosteroids on granulocytes to control A. fumigatus infections, we performed granulocyte adoptive cell transfers using in vivo mouse models, in vitro human and mouse granulocyte and A. fumigatus functional co-culture experiments in combination with flow cytometry, cytokine analysis, fluorescence and electron microscopy.Results:Transfusion of granulocytes from corticosteroid treated mice did not protect cyclophosphamide immunosuppressed mice against lethal A. fumigatus infection in contrast to granulocytes from untreated mice. Upon infection increased levels of inflammatory cytokines helped to recruit granulocytes to the lungs without any recruitment defects in corticosteroid treated and infected mice or in cyclophosphamide immunosuppressed and infected mice that had received the granulocytes from corticosteroid treated mice. However, corticosteroid treated human or mouse neutrophils failed to form neutrophil extracellular traps (NETs) under in vitro and in vivo conditions. Furthermore, corticosteroid treated granulocytes exhibited impaired ROS production against A. fumigatus. Notably, corticosteroids impaired the β-glucan receptor Dectin-1 (CLEC7A) on mouse and human granulocytes to efficiently recognize and phagocytize A. fumigatus, which markedly impaired fungal killing.Conclusions:We conclude that corticosteroid treatment of granulocyte donors for increasing neutrophil yields or patients with ongoing corticosteroid treatment could result in deleterious effects on granulocyte antifungal functions, thereby limiting the benefit of granulocyte transfusion therapies against invasive fungal infections. DisclosuresEinsele:Celgene: Consultancy, Honoraria, Speakers Bureau; Janssen: Consultancy, Honoraria, Speakers Bureau; Novartis: Consultancy, Honoraria; Amgen: Consultancy, Honoraria, Speakers Bureau.

Dynamic Immune Cell Recruitment After Murine Pulmonary Aspergillus fumigatus Infection under Different Immunosuppressive Regimens.

Humans are continuously exposed to airborne spores of the saprophytic fungus Aspergillus fumigatus. However, in healthy individuals pulmonary host defense mechanisms efficiently eliminate the fungus. In contrast, A. fumigatus causes devastating infections in immunocompromised patients. Host immune responses against A. fumigatus lung infections in immunocompromised conditions have remained largely elusive. Given the dynamic changes in immune cell subsets within tissues upon immunosuppressive therapy, we dissected the spatiotemporal pulmonary immune response after A. fumigatus infection to reveal basic immunological events that fail to effectively control invasive fungal disease. In different immunocompromised murine models, myeloid, notably neutrophils, and macrophages, but not lymphoid cells were strongly recruited to the lungs upon infection. Other myeloid cells, particularly dendritic cells and monocytes, were only recruited to lungs of corticosteroid treated mice, which developed a strong pulmonary inflammation after infection. Lymphoid cells, particularly CD4+ or CD8+ T-cells and NK cells were highly reduced upon immunosuppression and not recruited after A. fumigatus infection. Moreover, adoptive CD11b+ myeloid cell transfer rescued cyclophosphamide immunosuppressed mice from lethal A. fumigatus infection but not cortisone and cyclophosphamide immunosuppressed mice. Our findings illustrate that CD11b+ myeloid cells are critical for anti-A. fumigatus defense under cyclophosphamide immunosuppressed conditions.

Encephalitozoonosis in pharmacologically immunosuppressed mice

Encephalitozoon cuniculi is a parasite that has been identified as a cause of opportunistic infections in immunocompromised individuals. This study was performed to evaluate E. cuniculi infection in pharmacologically immunosuppressed mice. Mice were immunosuppressed with cyclophosphamide (100mg/kg twice a week, IP) or cyclosporin (10mg/kg daily, IP) and inoculated with 107E. cuniculi spores IP. The E. cuniculi spores were cultivated in MDCK cells. E. cuniculi identification was performed by light microscopy studies using Gram-Chromotrope, Hematoxylin–Eosin and Toluidine blue–fuchsin staining techniques, as well as by PCR at 15, 30 and 45days post-inoculation (DPI). Cyclophosphamide-immunosuppressed mice have greatly reduced amounts of CD8+, CD4+ and CD3+ T cells and CD19+ B cells. The cells from these mice were analyzed by FACS and showed acute disseminated and fatal encephalitozoonosis. Mice treated with ciclosporin, which is both antiparasitic and immunosuppressive, have a milder, chronic, non-lethal infection and showed a significant reduction only in CD3+ and CD4+ T cell numbers. Our results support the role of CD8+ T cells in controlling infection by E. cuniculi and show that preventive measures are essential for preventing this zoonosis in individuals undergoing chemotherapy for cancer or other immunosuppressive therapies.

Immunomodulatory activity of Zingiber officinale Roscoe, Salvia officinalis L. and Syzygium aromaticum L. essential oils: evidence for humor- and cell-mediated responses.

The immunomodulatory effect of ginger, Zingiber officinale (Zingiberaceae), sage, Salvia officinalis (Lamiaceae) and clove, Syzygium aromaticum (Myrtaceae), essential oils were evaluated by studying humor- and cell-mediated immune responses. Essential oils were administered to mice (once a day, orally, for a week) previously immunized with sheep red blood cells (SRBCs). Clove essential oil increased the total white blood cell (WBC) count and enhanced the delayed-type hypersensitivity (DTH) response in mice. Moreover, it restored cellular and humoral immune responses in cyclophosphamide-immunosuppressed mice in a dose-dependent manner. Ginger essential oil recovered the humoral immune response in immunosuppressed mice. Contrary to the ginger essential oil response, sage essential oil did not show any immunomodulatory activity. Our findings establish that the immunostimulatory activity found in mice treated with clove essential oil is due to improvement in humor- and cell-mediated immune response mechanisms.

Immunomodulatory activity of Zingiber officinale Roscoe, Salvia officinalis L. and Syzygium aromaticum L. essential oils: evidence for humor- and cell-mediated responses

Abstract Objectives The immunomodulatory effect of ginger, Zingiber officinale (Zingiberaceae), sage, Salvia officinalis (Lamiaceae) and clove, Syzygium aromaticum (Myrtaceae), essential oils were evaluated by studying humor- and cell-mediated immune responses. Methods Essential oils were administered to mice (once a day, orally, for a week) previously immunized with sheep red blood cells (SRBCs). Key findings Clove essential oil increased the total white blood cell (WBC) count and enhanced the delayed-type hypersensitivity (DTH) response in mice. Moreover, it restored cellular and humoral immune responses in cyclophosphamide-immunosuppressed mice in a dose-dependent manner. Ginger essential oil recovered the humoral immune response in immunosuppressed mice. Contrary to the ginger essential oil response, sage essential oil did not show any immunomodulatory activity. Conclusions Our findings establish that the immunostimulatory activity found in mice treated with clove essential oil is due to improvement in humor- and cell-mediated immune response mechanisms.

Treatment of transplanted adriamycin-resistant ovarian cancers in mice by combination of adriamycin and ultrasound exposure

Ovarian cancer models were established in cyclophosphamide immunosuppressed mice by subrenal capsular cell fibrin clot transplantation. SKOV 3 cancers were treated by adriamycin alone, or adriamycin combined with ultrasound exposure. SKOV 3 /ADR cancers were treated with adriamycin, as well as verapamil and insonation were administrated alone or concurrently. The results were: (1) Insonation alone could not suppress growth of tumours. (2) In SKOV 3 cancers, ultrasound exposure potentiated the efficiency of adriamycin. (3) In SKOV 3 /ADR cancers, insonation reversed adriamycin resistance, but verapamil was not effective and no synergism existed between it and ultrasound. These findings revealed that ultrasound exposure enhanced the efficiency of adriamycin to both chemosensitive and chemoresistant ovarian cancers in vivo. Mechanisms were discussed.

Treatment of transplanted adriamycin-resistant ovarian cancers in mice by combination of adriamycin and ultrasound exposure

Ovarian cancer models were established in cyclophosphamide immunosuppressed mice by subrenal capsular cell fibrin clot transplantation. SKOV 3 cancers were treated by adriamycin alone, or adriamycin combined with ultrasound exposure. SKOV 3/ADR cancers were treated with adriamycin, as well as verapamil and insonation were administrated alone or concurrently. The results were: (1) Insonation alone could not suppress growth of tumours. (2) In SKOV 3 cancers, ultrasound exposure potentiated the efficiency of adriamycin. (3) In SKOV 3/ADR cancers, insonation reversed adriamycin resistance, but verapamil was not effective and no synergism existed between it and ultrasound. These findings revealed that ultrasound exposure enhanced the efficiency of adriamycin to both chemosensitive and chemoresistant ovarian cancers in vivo. Mechanisms were discussed.

Activities of the triazole derivative SCH 56592 (posaconazole) against drug-resistant strains of the protozoan parasite Trypanosoma (Schizotrypanum) cruzi in immunocompetent and immunosuppressed murine hosts.

We have studied the in vivo activity of the new experimental triazole derivative SCH 56592 (posaconazole) against a variety of strains of the protozoan parasite Trypanosoma (Schizotrypanum) cruzi, the causative agent of Chagas' disease, in both immunocompetent and immunosuppressed murine hosts. The T. cruzi strains used in the study were previously characterized as susceptible (CL), partially resistant (Y), or highly resistant (Colombiana, SC-28, and VL-10) to the drugs currently in clinical use, nifurtimox and benznidazole. Furthermore, all strains are completely resistant to conventional antifungal azoles, such as ketoconazole. In the first study, acute infections with the CL, Y, and Colombiana strains in both normal and cyclophosphamide-immunosuppressed mice were treated orally, starting 4 days postinfection (p.i.), for 20 consecutive daily doses. The results indicated that in immunocompetent animals SCH 56592 at 20 mg/kg of body weight/day provided protection (80 to 90%) against death caused by all strains, a level comparable or superior to that provided by the optimal dose of benznidazole (100 mg/kg/day). Evaluation of parasitological cure revealed that SCH 56592 was able to cure 90 to 100% of the surviving animals infected with the CL and Y strains and 50% of those which received the benznidazole- and nifurtimox-resistant Colombiana strain. Immunosuppression markedly reduced the mean survival time of untreated mice infected with any of the strains, but this was not observed for the groups which received SCH 56592 at 20 mg/kg/day or benznidazole at 100 mg/kg/day. However, the overall cure rates were higher for animals treated with SCH 56592 than among those treated with benznidazole. The results were confirmed in a second study, using the same model but a longer (43-dose) treatment period. Finally, a model for the chronic disease in which oral treatment was started 120 days p.i. and consisted of 20 daily consecutive doses was investigated. The results showed that SCH 56592 at 20 mg/kg/day was able to induce a statistically significant increase in survival of animals infected with all strains, while benznidazole at 100 mg/kg/day was able to increase survival only in animals infected with the Colombiana strain. Moreover, the triazole was able to induce parasitological cures in 50 to 60% of surviving animals, irrespective of the infecting strain, while no cures were obtained with benznidazole. Taken together, the results demonstrate that SCH 56592 has in vivo trypanocidal activity, even against T. cruzi strains naturally resistant to nitrofurans, nitroimidazoles, and conventional antifungal azoles, and that this activity is retained to a large extent in immunosuppressed hosts.

Effects of lysozyme dimer on humoral response to sheep erythrocytes in non-treated and cyclophosphamide-immunosuppressed mice.

The effects of lysozyme dimer on humoral response to sheep erythrocytes (SRBC) and restoration of the response impaired by a single cyclophosphamide dose (200 mg/kg) were tested on mice. The effect of lysozyme dimer on the humoral response to SRBC in non-treated with cyclophosphamide mice was determined in relation to doses (0.2, 2, 20 or 200 micrograms/kg) and the time of the drug administration with respect to the antigen before or after SRBC immunization. Moreover, the effect of lysozyme dimer on the humoral response in cyclophosphamide-treated mice was studied depending on the dose applied and time of exposure to the drug in relation to SRBC. It has been found that lysozyme dimer potentiates the humoral response to SRBC in mice, resulting in an increased number of splenocytes producing haemolytic antibodies (PFC) and the total and 2-mercaptoethanol resistant level of anti-SRBC antibodies. A single exposure to lysozyme dimer gave the strongest stimulating action on SRBC when the doses of 2 or 20 micrograms/kg were administered 2 h prior to the antigen. The potentiating effect of the drug was reduced when it was administered 24 h before the antigen and also when single doses were as high as 200 micrograms/kg and as low as 2 micrograms/kg. Exposure to four doses of lysozyme dimer at 24 h intervals was more activating than a single injection. A strong potentiating effect on the specific response to SRBC was noted after four injections of lysozyme dimer at doses from 0.2 to 20 micrograms/kg. The effect of the drug did not depend on the time of exposure to the antigen. It has also been found that lysozyme dimer significantly reduces the suppressive effect of a high cyclophosphamide dose (200 mg/kg) on the humoral response of SRBC-immunized mice. The protective action of lysozyme dimer was dose- and time-dependent. The strongest protection was observed after three doses of 20 micrograms/kg administered prior to pharmacological immunosuppression. Reduction in the dose to 2 micrograms/kg and shorter treatment resulted in reduced protective effects. We have also found that the protective action of three doses of lysozyme dimer (2 or 20 micrograms/kg each) administered between cyclophosphamide injection and the antigen, or after antigen administration is weaker than such a treatment prior to cyclophosphamide immunosuppression.

The effect of DTC on humoral response restoration and thymocyte subpopulations in cyclophosphamide-immunosuppressed mice.

The effects of sodium diethyldithiocarbamate (DTC) on humoral response to SRBC and restoration of the response impaired by a single cyclophosphamide dose (200 mg/kg) were tested on mice. Moreover, the effect of DTC (20 mg/kg) on thymocyte subpopulations was tested on non-immunized mice previously treated with cyclophosphamide. It was found that DTC (20 mg/kg) administered to the immunized mice enhanced humoral response to SRBC, which was reflected in the increased number of PFC and (7S) serum hemagglutinin titer. In contrast, partial restoration of the primary humoral response after DTC injection was observed in the mice administered a single cyclophosphamide dose. The effect of DTC was stronger after 6 days following cyclophosphamide injection, i.e. at the time when spontaneous restoration of T lymphocytes begins. In addition, it was found that DTC given to non-immunized mice treated with cyclophosphamide had a modulating effect on thymocyte subpopulations. Depending on the time of exposure to cyclophosphamide, DTC either increased the percentage of CD4 thymocytes or decreased the percentage of CD8, which subsequently led increased CD4/CD8 coefficient. DTC did not change the suppressing action of cyclophosphamide on the percentage of double-positive thymocytes.

Hypoxanthine Derivatives in Experimental Infections

In vivo treatment with parenterally administered hypoxanthine derivatives, notably ST 789, was able to protect cyclophosphamide‐immunosuppressed mice against experimental infections with both bacterial and fungal pathogens. However, the mechanisms accounting for these effects of hypoxanthine derivatives remain to be fully established. In fact, only the treatment with ST 789 resulted in a clear enhancement of the primary antibody production as well as macrophage phagocytic activity, whereas T lymphocyte responsiveness to mitogens and both macrophage‐ and natural killer‐dependent cytotoxicity were not significantly affected. These data, together with the recently shown ability of ST 789 to increase interleukin‐6 production, suggest that monocyte/macrophages are likely to be the main cellular target of the immunomodulating activity of ST 789. Finally, in the presentln vivo study, hypoxanthine derivatives did not enhance the mean survival time of tumour‐bearing immunosuppressed mice.

Immunostimulating effects of 2 hypoxanthine derivatives (ST 789 and ST 657) in cyclophosphamide-immunosuppressed mice

Immunostimulating effects of 2 hypoxanthine derivatives (ST 789 and ST 657) in cyclophosphamide-immunosuppressed mice