Cyclophosphamide-induced Immunosuppression Research Articles

Synergistic immunostimulating activity of pidotimod and red ginseng acidic polysaccharide against cyclophosphamide-induced immunosuppression

We investigated the synergistic effect of combined treatment with red ginseng acidic polysaccharide (RGAP) from Panax ginseng C.A. Meyer and pidotimod in cyclophosphamide-treated mice. The combination of pidotimod and RGAP restored concanavalin A-induced splenic T cell proliferation and LPS-stimulated B cell proliferation significantly. The production of nitric oxide from peritoneal macrophages was increased by the combinations. NK cell activity was increased by RGAP alone or in combination with pidotimod. A synergistic increase in the level of serum IL-12 and interferongamm was observed when the combination of the two was used. RGAP alone or in combination with pidotimod modulated the level of serum C-reactive protein to a near-normal level. These results indicate that combinations of pidotimod and RGAP are synergistic and suggest that combination therapy using pidotimod and RGAP for improving immune activity may provide an additional benefit over the use of the two drugs by themselves.

Effect of AC II, an Herbal Formulation in Cyclophosphamide-Induced Immunosuppression in BALB/c Mice—Implication in HIV Treatment

Effect of AC II, herbal drug formulation in reducing immunosuppression caused by administration of cyclophosphamide was studied. Mice were injected cyclophosphamide (CTX) 50 mg/kg b.wt. for 14 days with or without the drug and total WBC, bone marrow cellularity and α-esterase positive cells were determined. On day 15, total WBC count in cyclophosphamide treated mice was 1500 ± 420 cells/mm3, while in AC II-treated mice it was 7658 ± 376 cells/mm3. On day 16, administration of cyclophosphamide reduced bone marrow cellularity to 3.42 ± 0.38 × 106cells/femur from the normal value of 13.83 ± 0.96 × 106 cells/femur. In AC II treated group bone marrow cellularity was increased to 8.05 ± 0.7 × 106 cells/femur. The number of α-esterase positive cells was found to be reduced to 177 ± 25 cells per 4000 cells in CTX treated groups. But in AC II-treated group the number of α-esterase positive cells were raised to 843 ± 86 cells per 4000 cells, which was closer to that of normal (710 ± 49 cells per 4000 cells). Results indicate the usefulness of AC II to combat immunosuppression induced by chemical and biological agents.

Antioxidant activities of Sophora subprosrate polysaccharide in immunosuppressed mice

It is known that the reactive oxygen species (ROS) and free radicals play multiple roles in some pathological events. Sophora subprosrate polysaccharide (SSP1), a natural polysaccharide with a mean molecular weight of 2.24 × 10 4, isolated from the roots of Sophora subprosrate was studied for its antioxidant activities in mice. The in vivo investigation demonstrated that intraperitoneal administration of SSP1 was able to overcome cyclophosphamide-induced immunosuppression and significantly raised glutathione level, superoxidase dismutase activity, total antioxidant capacity, thymus and spleen indices in a dose-dependent manner in mice. The results showed that the SSP1, possessing pronounced free radical scavenging and antioxidant activities, may play an important role in prevention of oxidative damage in immunological system.

Effects of polysaccharide peptide (PSP) from Coriolus versicolor on the pharmacokinetics of cyclophosphamide in the rat and cytotoxicity in HepG2 cells

Polysaccharide peptide (PSP), isolated from Coriolus versicolor COV-1, has been shown to restore the immunological effects against cyclophosphamide-induced immuno-suppression, although the mechanism(s) involved remain uncertain. This study investigated the PSP-cyclophosphamide interaction by studying the effects of PSP on the pharmacokinetic of cyclophosphamide in the rat and the effect of PSP on the cytotoxic effects of cyclophosphamide on a cancer cell line (HepG2 cells). In the pharmacokinetic studies in the rat, acute pre-treatment of PSP (4 μmol/kg/day, i.p.) decreased the clearance (CL) of cyclophosphamide by 31%, with a concomitant increase in the area under concentration–time curve (AUC) by 44%, and prolongation of the plasma half-life ( T 1/2) by 43%. Sub-chronic pre-treatment of PSP (2 μmol/kg/day, i.p., 3 days) decreased the CL of cyclophosphamide by 33%, with a concomitant increase in the AUC by 50%, and prolongation of the plasma T 1/2 by 34%. In cytotoxicity studies using HepG2 cells, non-toxic dose of PSP (1–10 μM) enhanced the cytotoxicity of cyclophosphamide. PSP at 10 μM further decreased HepG2 cell viability by 22% compared to when cyclophosphamide was present alone. In summary, PSP enhanced the cytotoxic effect of cyclophosphamide on a cancer cell line in vitro and altered the pharmacokinetics of cyclophosphamide in vivo in the rat. Both of these effects may be beneficial in the use of PSP as an adjunct to cyclophosphamide treatment.

Preventive and therapeutic effects of sugar cane extract on cyclophosphamide-induced immunosuppression in chickens

Effects of oral administration of sugar cane extract (SCE) on immunosuppression in chickens treated with cyclophosphamide (CPA) were evaluated. Three-week-old inbred chickens were inoculated into the crop with SCE (500 mg/kg/day) for three consecutive days before or after injection of CPA 12 or 20 mg/chicken. At the last day of SCE or CPA treatment, all chickens were immunized intravenously with sheep red blood cells (SRBC) and Brucella abortus (BA). Chickens administered SCE showed a significant increase in body weight, gain in body weight/day, relative weight of the bursa of Fabricius and antibody responses to SRBC and BA than untreated control chickens. Chickens injected with CPA alone showed significantly decreased body weight, gain in body weight/day, relative weight of the bursa and antibody responses to SRBC and BA, showing immunosuppression in the bursa-dependent immune system. All chickens administered SCE before or after the treatment with CPA showed significantly higher values in body weight, gain in body weight/day, relative bursal weight and antibody responses to both antigens, when compared to chickens treated with CPA alone. In histological examination, chickens administered SCE showed a typical bursa with well constituted follicles, although chickens treated with CPA alone showed a severely atrophied bursa with rudimentary follicles and enormous proliferation of interfollicular connective tissue. Chickens treated with SCE and CPA showed a well-reconstituted bursa with almost normal structure. These results suggest that SCE has functionally and morphologically reconstituting effects on the bursa-dependent immune system in immunosuppressed chickens induced by injection of CPA.

In vitro and in Vivo inhibition of immunoglobulin secretion by the immunosuppressive compound HR325 is reversed by exogenous uridine.

The objective was to demonstrate that the immunosuppressive agent HR325 (an inhibitor of dihydroorotate dehydrogenase, DHODH) inhibits immunoglobulin (Ig) secretion both in vitro and in vivo and that this effect can be reversed with exogenous uridine. In vitro, Ig secretion from mouse splenocytes was induced by lipopolysaccharide (LPS) for 5 days. HR325 inhibited the secretion of IgM and IgG with IC50 values of 2.5 and 2 microm, respectively. Adding uridine (50 microm) increased these values to 70 and 60 microm, respectively. Similarly, the IC50 values of another DHODH inhibitor, brequinar sodium, were also attenuated by uridine from 0.04 to 1 microm for IgM, and 0.012 to 10 microm for IgG. HR325 (and a structural analogue A771726) inhibited LPS-induced kappa light-chain cell surface expression on 70Z/3 cells, a property also reversed by uridine. In vivo, the secondary anti-sheep red blood cell (SRBC) antibody response (unaffected by uridine alone) was inhibited by HR325 and brequinar with respective ID50 values of 38 and 0.6 mg/kg per oral (p.o.). Immunosuppression with HR325 (50 mg/kg) and brequinar (1 mg/kg) was abrogated by uridine. Uridine had no effect on cyclophosphamide-induced (10 mg/kg p.o.) immunosuppression. These data are consistent with the immunosuppressive mechanism of HR325 being the result of pyrimidine depletion in vitro and in vivo.

Subtractive immunization: a tool for the generation of discriminatory antibodies to proteins of similar sequence

Antibodies specific for a protein of interest are invaluable tools for monitoring the protein's structure, location and activity. Due to the tendency of an immune system to mount a response toward the abundant, immunodominant epitopes in a protein mixture, difficulties are inherent in the isolation of antibodies specific for proteins that are rare or poorly immunogenic. Likewise, isolation of antibodies specific for a protein with significant sequence similarity to other proteins, such as those derived from protein engineering, may be challenging. Subtractive immunization is a technique proven to facilitate efforts to produce monoclonal antibodies specific for antigens that are present in low abundance in a protein mixture, poorly immunogenic and/or similar in sequence or structure to other proteins. This protocol provides a detailed, stepwise procedure for the isolation of antibodies specific for a protein with sequence similarity to other proteins. As an example, we describe methods established to isolate antibodies specific to a methionine-enriched variant of soybean vegetative storage protein β (VSPβ-Met) that shares 91.8% amino acid sequence identity to the wild-type protein (VSPβ-WT). These methods include cyclophosphamide-induced immunosuppression of mice for the wild-type protein followed by immunization with VSPβ-Met. As a result of this procedure, mouse polyclonal antibodies that exhibited 10-fold greater reactivity with VSPβ-Met than VSPβ-WT in an ELISA were generated. It is anticipated that this strategy will have utility for generating antibodies specific to protein variants derived from protein engineering.

Cyclophosphamide-induced immunosuppression protects cardiac noradrenergic nerve terminals from damage by Trypanosoma cruzi infection in adult rats

Trypanosoma cruzi-infected juvenile rats develop severe cardiac sympathetic denervation in parallel with acute myocarditis. This aspect has not been studied in adult rats, thought to be resistant to this infection. The mechanism involved in T. cruzi-induced neuronal damage remains to be completely elucidated. In juvenile rats, the mortality during the acute phase depends on T. cruzi populations, ranging from 30% to 100%. Therefore, studies of mechanisms through hazardous procedures such as immunosuppression are restricted. The current paper shows that adult rats infected with T. cruzi (Y strain) develop severe acute myocarditis and cardiac sympathetic denervation, despite null mortality and virtual absence of patent parasitaemia followed by negative haemoculture. Recovery from the myocarditis and denervation occurred but PCR studies showed persistence of parasite DNA at least until day 111 post inoculation. Immunosuppression by cyclophosphamide treatment increased the parasitaemia, prevented the acute myocarditis and the sympathetic denervation without significant alteration of the myocardial parasitism. These results argue against a direct role for parasite-derived products and implicate the inflammatory cells in the denervation process. As previous studies in juvenile animals have discarded an essential role for radiosensitive cells, the macrophages remain as the possible effectors for the T. cruzi-induced neuronal damage.

Studies on immunomodulatory activity of Withania somnifera (Ashwagandha) extracts in experimental immune inflammation

The immunomodulatory activities of an Indian Ayurvedic medicinal preparation, i.e. extracts from Ashwagandha, Withania somnifera (L.) Dunal (Solanaceae), namely WST and WS2, were studied in mice for immune inflammation: active paw anaphylaxis and delayed type hypersensitivity (DTH). Immunomodulatory effect was assessed in If IgE-mediated anaphylaxis as reduction of ovalbumin-induced paw edema, in animals treated with WS2 at doses of 150 and 300 mg/kg, and the results were compared with the standard drug disodium chromoglycate. In the DTH model, the modulatory effect was assessed as potentiation or suppression of the reaction, revealing an increase or decrease in mean foot pad thickness, respectively. Potentiation of the DTH reaction was observed in animals treated with cyclophosphamide at a dose of 20 mg/kg, WST at a dose of 1000 mg/kg and WS2 at a dose of 300 mg/kg. On the other hand, cyclophosphamide-induced potentiation of DTH reaction was suppressed in animals treated with WST and WS2. A significant increase in white blood cell counts and platelet counts was observed in animals treated with WST. A protective effect in cyclophosphamide-induced myelosuppression was observed in animals treated with WST and WS2, revealing a significant increase in white blood cell counts and platelet counts. Cyclophosphamide-induced immunosuppression was counteracted by treatment with WS2, revealing significant increase in hemagglutinating antibody responses and hemolytic antibody responses towards sheep red blood cells.

Therapeutic efficacy of BO-3482, a novel dithiocarbamate carbapenem, in mice infected with methicillin-resistant Staphylococcus aureus.

The in vivo activity of BO-3482, which has a dithiocarbamate chain at the C-2 position of 1beta-methyl-carbapenem, was compared with those of vancomycin and imipenem in murine models of septicemia and thigh infection with methicillin-resistant Staphylococcus aureus (MRSA). Because BO-3482 was more susceptible than imipenem to renal dehydropeptidase I in a kinetic study of hydrolysis by this renal enzyme, the therapeutic efficacy of BO-3482 was determined during coadministration with cilastatin. In the septicemia models, which involved two homogeneous MRSA strains and one heterogeneous MRSA strain, the 50% effective doses were, respectively, 4.80, 6.06, and 0.46 mg/kg of body weight for BO-3482; 5.56, 2.15, and 1.79 mg/kg for vancomycin; and >200, >200, and 15.9 mg/kg for imipenem. BO-3482 was also as effective as vancomycin in an MRSA septicemia model with mice with cyclophosphamide-induced immunosuppression. In the thigh infection model with a homogeneous MRSA strain, the bacterial counts in tissues treated with BO-3482-cilastatin were significantly reduced in a dose-dependent manner compared with the counts in those treated with vancomycin and imipenem-cilastatin (P < 0.001). These results indicate that BO-3482-cilastatin is as effective as vancomycin in murine systemic infections and is more bactericidal than vancomycin in local-tissue infections. The potent in vivo activity of BO-3482-cilastatin against such MRSA infections can be ascribed to the good in vitro anti-MRSA activity and improved pharmacokinetics in mice when BO-3482 is combined with cilastatin and to the bactericidal nature of the carbapenem.

The protective role of the immunomodulator MECH against cyclophosphamide-induced immunosuppression in mice

The protective role of the immunomodulator MECH against cyclophosphamide-induced immunosuppression in mice

The protective role of the immunomodulator MECH against cyclophosphamide-induced immunosuppression in mice

The protective role of the immunomodulator MECH against cyclophosphamide-induced immunosuppression in mice

Immunopathogenic role of T-cell subsets in Borna disease virus-induced progressive encephalitis.

Borna disease is an immunopathological virus-induced encephalopathy comprising severe inflammation and degenerative brain cell lesions which results in organ atrophy and chronic debility in rats. CD4+ and CD8+ T cells have been reported to be involved in the development of this disease of the central nervous system. A virus-specific homogeneous T-cell line, established in vitro after immunization of rats with the recombinant 24-kDa virus-specific protein, showed antigen-specific proliferation in the presence of the 24-kDa but not the 38-kDa Borna disease virus-specific protein, another major virus-specific antigen. This T-cell line, P205, was found to exhibit characteristics of a T-helper cell: CD4+ CD8- IL-2- IL-4- IFN-gamma+ IL-6+ IL-10+. Furthermore, this T-cell line expressed the alpha/beta T-cell receptor and the alpha 4 integrin (VLA-4). Adoptive transfer of this helper cell resulted in an increase of antibody titers and two different types of disease in virus-infected rats after cyclophosphamide-induced immunosuppression. (i) Rats receiving T cells between 10 and 18 days after treatment with cyclophosphamide showed an acute lymphoproliferative disease in the gut and lungs within 9 days after adoptive transfer and died. (ii) Passive transfer within the first 5 days after immunosuppressive treatment resulted in typical Borna disease associated with neurological symptoms such as ataxia and paresis starting 14 to 16 days after transfer. Immunohistological analysis of the brains of rats with Borna disease uniformly revealed the presence of CD8+ T cells in encephalitic lesions in addition to CD4+ cells that were found in the brains of recipients of the virus-specific CD4+ T-cell line, irrespective of whether neurological symptoms developed or not. However, recipient rats treated with antibodies against CD8+ T cells developed neither encephalitis nor disease. Therefore, CD4+ T cells appear to accumulate in the brain and cause perivascular inflammatory lesions which alone obviously do not cause disease. In contrast, the presence of CD8+ cells apparently directly correlates with the development of neurological symptoms.

A trypsin‐sensitive avian reovirus: Isolation and experimental infection of poults and chicks

A reovirus (strain TR1) was isolated from the hock joint of a turkey with arthritis. It was rarely excreted in the faeces of poults after oral or intranasal inoculation and only occasionally following footpad inoculation. The virus did not localise in the hock joints or cause lesions. Cyclophosphamide-induced immunosuppression of poults did not modify the infection. Oral inoculation of chicks with a high dose of strain TR1 resulted in joint localisation, of virus and erosive arthritis. Footpad inoculation caused deaths in 13 of 15 chicks by 8 days pi due to hepatic necrosis. Intranasal inoculation of chicks resulted in a delayed and limited faecal excretion. However, joint lesions were most marked in chicks infected via this route. The infectivity of strain TR1 in chick embryo liver cell monolayers was reduced by almost 3.0 log(10) after treatment with 0.01% trypsin for 30 min, but the infectivity of strain R2 was unaffected. Bile salts had no effect on either virus. This trypsin-sensitive reovirus was much more pathogenic for chicks than for poults. The respiratory route may be important in the dissemination of trypsin-sensitive avian reoviruses such as this.

Antibody production by different sites and cyclophosphamide-induced immunosuppression of the TNP-LPS response in the grass frog, [formula omitted][formula omitted

Antibody production by different sites and cyclophosphamide-induced immunosuppression of the TNP-LPS response in the grass frog, [formula omitted][formula omitted

Resistance of peripheral autonomic neurons to in vivo productive infection by herpes simplex virus mutants deficient in thymidine kinase activity.

We used a model involving acute and latent herpes simplex virus (HSV) infections of mouse superior cervical ganglia to assess in vivo neuronal infections with two thymidine kinase-deficient (TK-) mutants of HSV type 1. Despite replication of the TK- HSV strains at the site of inoculation in the eyes, little if any viral replication occurred in the superior cervical ganglia, as assessed by the viral titers of ganglion homogenates, viral antigens in tissue sections, and histopathological evidence of cytopathology or inflammation. Cyclophosphamide-induced immunosuppression and treatment with 6-hydroxydopamine, which enhanced productive infections of superior cervical ganglia with TK+ HSV did not induce TK- HSV ganglionic infections. Latent TK- HSV infections were not detected by cocultivation of ganglion explants. Efforts to infect ganglia in culture after they were removed from animals indicated that superior cervical ganglia, and particularly their neuronal elements, resisted productive TK- HSV infection. These results supported the hypothesis that viral thymidine kinase facilitates acute and reactivated productive HSV infections of neurons.

Cyclophosphamide-Induced Immunosuppression Demonstrated in Pasteurella multocida-Vaccinated Chickens

SUMMARY Vaccinated cyclophosphamide-treated chickens were more susceptible to challenge infection than were vaccinated untreated chickens. Antibody titers were greatly reduced in cyclophosphamidetreated chickens. The results suggest that the presence of specific antibody is important in determining the ability of chickens to resist Pasteurella infection.

Immunological reactivity of the lung: VII. Effect of corticosteroids and cyclophosphamide on the Fc receptor function of alveolar macrophages

The effects of various in vitro and in vivo regimens of either corticosteroid or cyclophosphamide administration on guinea pig alveolar macrophages were studied. Corticosteroid- and cyclophosphamide-induced immunosuppression was assessed by the effect of drug administration on the capacity of alveolar macrophages to attach to and/or ingest antibody-coated sheep red blood cells (SRBC). In vitro hydrocortisone (up to 20 μg/ml) had no effect on either the binding or ingestion of antibody-coated SRBC. Two separate regimens of in vivo corticosteroids were given: a single dose of iv hydrocortisone (100 mg/kg), which is a short-acting soluble preparation, and sc doses of cortisone acetate (100 mg/kg for 7 days), which is a depot preparation resulting in sustained levels of plasma cortisol of the magnitude of that found for a brief period of time following iv injection of hydrocortisone. Both regimens resulted in similar degrees of peripheral blood lymphocytopenia and monocytopenia 4 and 24 hr, respectively, following injection. The regimen of hydrocortisone has previously been reported to have no effect on alveolar macrophage cytotoxic effector function in antibody-dependent cellular cytotoxicity (ADCC), whereas the cortisone acetate regimen markedly suppressed ADCC. In the present study, hydrocortisone had no effect on either the binding or ingestion of antibody-coated SRBC by alveolar macrophages. In contrast, cortisone acetate caused a marked decrease in both the binding and ingestion of antibody-coated SRBC. This suppressive effect was maximal at suboptimal concentrations of antibody on the SRBC and could be overcome by increasing the concentrations of anti-SRBC antibody. Alveolar macrophages from animals treated with daily cyclophosphamide (a regimen which suppresses ADCC) were capable of binding and ingesting antibody-coated SRBC normally. Thus, prolonged exposure to corticosteroids in vivo causes an alteration in membrane Fc receptor function of alveolar macrophages, which can explain this impaired ability to kill target cells. Since cyclophosphamide therapy did not interfere with the binding and ingestion of antibody-coated target cells, it is concluded that the impairment in killing of target cells by alveolar macrophages is not directly related to an alteration of Fc receptor function but to a defect in the actual killing process.

Effect of n-acetylcysteine on some aspects of cyclophosphamide-induced toxicity and immunosuppression

Cyclophosphamide (CPA)-induced bladder toxicity and lethality were inhibited by N-acetylcysteine (NAC) when given systemically at a 4 : 1 ratio prior to CPA. This dose of NAC did not affect the production or the formation of free alkylating agents from CPA in vivo or in vitro. The immunosuppressive effect of CPA against T cell response systems, such as graft vs host reaction, antibody to sheep red blood cells and (PHA) phytohemagglutin stimulation was unaffected by NAC. It is concluded that the metabolic products of CPA for cytotoxicity as expressed as cystitis and lethality are different from the alkylating agents, which appear to affect immunological phenomena.

Divergent Effects of Cyclophosphamide Administration on Mononuclear Killer Cells: Quantitative Depletion of Cell Numbers Versus Qualitative Suppression of Functional Capabilities

The effects of various regimens of cyclophosphamide administration on guinea pig peripheral blood leukocytes were studied. Cyclophosphamide-induced immunosuppression was assessed by the effect of drug administration on the proportions and absolute numbers of leukocyte populations, and by the effect on functional capabilities of unfractionated and adherent cell-depleted mononuclear cell suspensions as measured by the PHA-induced cellular cytotoxicity and antibody-dependent cellular cytotoxicity assays against chicken erythrocyte targets. Intraperitoneal administration of five daily doses of cyclophosphamide (5 mg/kg) caused a modest absolute leukopenia but no change in cytotoxic effector function of the mononuclear cells remaining in the circulation. As the dosage of cyclophosphamide was increased to 20 mg/kg/day to produce a pronounced leukopenia, a profound neutropenia (less than 300 polymorphonuclear leukocytes/mm3) together with a marked decrease in mononuclear cell effector function was noted. A single i.p. injection of cyclophosphamide (100 mg/kg), which produced identical degrees of leukopenia of each leukocyte class as did daily administration of cyclophosphamide (20 mg/kg/day), caused no change in mononuclear cell effector function when compared to saline controls. Complement receptor-bearing and Fc-receptor bearing mononuclear cells were decreased to the same degree by both regimens of cyclophosphamide administration. Removal of adherent cells from mononuclear cell suspensions by column purification resulted in a marked decrease in cytotoxic effector function at low effector to target ratios. At higher effector to target ratios there was no difference in cytotoxic effector function between unfractionated and column-purified cells. In contrast, the functional defect in mononuclear cell suspensions from animals that received five daily doses of cyclophosphamide (20 mg/kg) could not be compensated for at higher effector to target ratios, indicating that this functional defect was not an artifact of relative depletion of monocytes by cyclophosphamide, but was due to an actual suppression of the effector functional capabilities of the killer cells. This study indicates that, dependent on the particular regimen of drug administration, the quantitative depletion of mononuclear cell populations by cyclophosphamide administration can be clearly distinguished from the qualitative effect on certain functional capabilities of surviving cells.