Histopathology of murine toxoplasmosis under treatment with dialyzable leukocyte extract.

BACKGROUND: Dialyzable leukocyte extracts (DLEs) contain molecules smaller than 10 kDa with biological activity in receptor organisms. Primarily, they participate in the regulation of the Th1 immune response, which is essential for the control of several intracellular infections, such as toxoplasmosis. This disease is associated with congenital infection, encephalitis or systemic infections in immunocompromised individuals. The clinical course of this infection fundamentally depends on a well-regulated immune response and timely treatment with the appropriate drugs. OBJECTIVE: The aim of this study was to evaluate the effect of treatment with a leukocyte extract, derived from crocodile lymphoid tissue, on the histopathology and brain parasite load in NIH mice that had been infected with cysts of Toxoplasma gondii (ME-49 strain). METHODS: The treatment was applied during the acute and chronic stages of the infection. Histopathological changes were evaluated in the ileum, liver and spleen at one, four and eight weeks after infection and in the brain at week 8. The parasite load was evaluated by counting the cysts of T. gondii found in the brain. FINDINGS: Compared to the control mouse group, the mice infected with T. gondii and under treatment with DLE showed less tissue damage, mainly at the intestinal, splenic and hepatic levels. In addition, a greater percentage of survival was observed, and there was a considerable reduction in the parasite load in the brain. CONCLUSIONS: The results suggest that DLE derived from crocodile is a potential adjunctive therapy in the conventional treatment of toxoplasmosis.

Histopathology of murine toxoplasmosis under treatment with dialyzable leukocyte extract

BACKGROUND Dialyzable leukocyte extracts (DLEs) contain molecules smaller than 10 kDa with biological activity in receptor organisms. Primarily, they participate in the regulation of the Th1 immune response, which is essential for the control of several intracellular infections, such as toxoplasmosis. This disease is associated with congenital infection, encephalitis or systemic infections in immunocompromised individuals. The clinical course of this infection fundamentally depends on a well-regulated immune response and timely treatment with the appropriate drugs. OBJECTIVE The aim of this study was to evaluate the effect of treatment with a leukocyte extract, derived from crocodile lymphoid tissue, on the histopathology and brain parasite load in NIH mice that had been infected with cysts of Toxoplasma gondii (ME-49 strain). METHODS The treatment was applied during the acute and chronic stages of the infection. Histopathological changes were evaluated in the ileum, liver and spleen at one, four and eight weeks after infection and in the brain at week 8. The parasite load was evaluated by counting the cysts of T. gondii found in the brain. FINDINGS Compared to the control mouse group, the mice infected with T. gondii and under treatment with DLE showed less tissue damage, mainly at the intestinal, splenic and hepatic levels. In addition, a greater percentage of survival was observed, and there was a considerable reduction in the parasite load in the brain. CONCLUSIONS The results suggest that DLE derived from crocodile is a potential adjunctive therapy in the conventional treatment of toxoplasmosis.

[Transfer factors in medical therapy]. / Factores de transferencia en la terapéutica médica.

Transfer factor (TF) consists of messenger peptides produced by activated T lymphocytes as part of cellular immunity, and it acts in virgin lymphocytes through TF inducers, suppressors and specific antigens. TF is not immunogenic because it is not species-specific, since it contains a consensus sequence of amino acids LLYAQDL/VEDN. TF extracted from leukocytes can transfer immunity from a human to another species. TF extracts are complex, containing more than 200 molecules with molecular weights ranging from 1 to 20 kDa. The antigen specific transfer factors (STF) have molecular weights between 3,5 and 5 kDa. TF is easy to prepare and well tolerated. It does not contain HL-A antigens against potential receptors and it can used as adjuvant therapy in several diseases.

Preparation and properties of the specific anti-influenza virus transfer factor.

Specific anti-influenza virus and normal transfer factors prepared in an experimental animal model, the pig, have been tested for their components, characteristics, and activity of known specificity. Two transfer factors are small molecular mixture which consist entirely or partly of polypeptides and polynucleosides. Moreover, the biological activity of transfer factors could be approved by Rosettes test and specific skin test. The study would lay a foundation for the research and development of other specific transfer factor.

Transfer factors: identification of conserved sequences in transfer factor molecules.

BACKGROUND: Transfer factors are small proteins that "transfer" the ability to express cell-mediated immunity from immune donors to non-immune recipients. We developed a process for purifying specific transfer factors to apparent homogeneity. This allowed us to separate individual transfer factors from mixtures containing several transfer factors and to demonstrate the antigen-specificity of transfer factors. Transfer factors have been shown to be an effective means for correction of deficient cellular immunity in patients with opportunistic infections, such as candidiasis or recurrent Herpes simplex and to provide prophylactic immunity against varicella-zoster in patients with acute leukemia. MATERIALS AND METHODS: Transfer factors of bovine and murine origin were purified by affinity chromatography and high performance liquid chromatography. Cyanogen bromide digests were sequenced. The properties of an apparently conserved sequence on expression of delayed-type hypersensitivity by transfer factor recipients were assessed. RESULTS: A novel amino acid sequence, LLYAQDL/VEDN, was identified in each of seven transfer factor preparations. These peptides would not transfer expression of delayed-type hypersensitivity to recipients, which indicates that they are not sufficient for expression of the specificity or immunological properties of native transfer factors. However, administration of the peptides to recipients of native transfer factors blocked expression of delayed-type hypersensitivity by the recipients. The peptides were not immunosuppressive. CONCLUSIONS: These findings suggest that the peptides may represent the portion of transfer factors that binds to the "target cells" for transfer factors. Identification of these cells will be helpful in defining the mechanisms of action of transfer factors.

[The isolation of a transfer factor of the delayed-hypersensitivity type to the antigenic substances of Staphylococcus aureus in guinea pigs]. / Oderzhannia faktora perenosu hiperchutlyvosti spovil'nenoho typu mors'kykh svynok do antyhennykh substantsiï Staphylococcus aureus.

Immunoactivating effects of guinea pig's staphylococcal Transfer factor (TF) were studied in homologous and heterologous (human leukocytes) systems. In vivo (skin tests) and in vitro (Inhibition of leukocytes migration) tests showed that our TF is able to activate intact as well as sensibilized cells. Antigen-specific properties of TF also were showed.

Isolation and purification of HSV-1 specific transfer factor produced by HSV-1 immunized goat leukocyte dialysate.

A herpes simplex virus type 1 (HSV-1)-specific transfer factor (TF), was separated and purified from the leukocyte dialysate of goats immunized with HSV-1 using affinity chromatography on antigen-sorbent and reversed phase high performance liquid chromatography (RP-HPLC). The antigen-specific activities of the starting dialysate and the isolated TF component (s) were examined by 51Cr-labelled leukocyte adherence inhibition (51Cr LAI) assay. The analytical hydrophobic interaction HPLC (HI-HPLC) and isoelectric focusing (IEF) techniques were employed to evaluate the purity and the isoelectric point (PI) of isolated TF component(s). The experiments provided a two-step procedure for purifying the TF material from the starting dialysate. It seems that the purified active TF component (PTFC) was specific for HSV-1. The specific PTFC activity was increased 10,000-fold as compared with the activity of the dialysate. The active moiety appeared as a single band in the IEF gel as demonstrated by silver staining; it was hydrophilic and its PI was pH 4.48.

Purification of transfer factors.

Transfer factor activities have been studied in both clinical and basic science settings for several decades. Until now, highly purified transfer factors that are suitable for molecular analysis have not been available. This has impeded progress towards understanding the molecular and cellular basis of the activities of these important inducers of cell-mediated immune responses. Murine transfer factors with specificities for chicken egg albumin or horse spleen ferritin were purified to virtual homogeneity using a combination of affinity chromatography and reversed-phase and polytypic high performance liquid chromatography (hplc). Transfer factors prepared by this methodology were recovered in high yield and in biologically-active, antigen-specific forms. The purified materials were further analyzed using sodium dodecyl sulfate polyacrylamide gel electrophoresis, chromatographic methods and an in vivo assay for immunological activity. For the first time definitions for unit transfer factor activity and specific activity are introduced. The results of these experiments indicate that transfer factors are a family of highly polar, hydrophilic molecules of low molecular weight (approximately 5,000) which are produced in small quantities by lymphoid cells and which have potent biological activity. The availability of purified transfer factors should facilitate definitive studies into the nature and mechanisms of production and action of these molecules.

Amino acid analysis of selected reversed-phase high performance liquid chromatographic peaks of crude and partially purified lysed human leukocyte ultrafiltrate.

Analytic reversed-phase high performance liquid chromatography (RP-HPLC) was performed to separate from the crude lysed human leukocyte ultrafiltrate (LLU) its partially purified most immunoactive subfraction P2/II in vivo. Under conditions used, the highest degree of segregation of both. LLU and P2/II could be observed in the first, as well as in the last two fifth of the water-methanol gradient. The comparison of the RP-HPLC traces of LLU and P2/II suggests that probably some hydrophilic components of LLU have been removed or--at least--diminished. The preliminary amino acid analysis (AA) of the selected peaks showed that none of them lacks Gly, Ser, and Glu. Of the basic amino acid residues Lys has been found with relatively many peaks while hydrophobic as well as aromatic amino acids have been represented very modestly. Further study is warranted in order to determine better the bearings of presented findings for the in vivo situation.

[Delayed hypersensitivity to the antigens of the herpes simplex virus and herpetic resistance induced in mice by the administration of xenogeneic lymphocytic ultrafiltrates]. / Giperchuvstvitel'nost' zamedlennogo tipa k antigenam virusa prostogo gerpesa i protivogerpeticheskaia rezistentnost', indutsirovannye u myshei vvedeniem ksenogennykh limfotsitarnykh ul'trafil'tratov.

Experiments on mice have shown that ultrafiltrates, prepared from lymphocytes obtained from the spleen of horses immunized with herpes vaccine and from human tonsils, contain herpes-specific transfer factor inducing delayed hypersensitivity. The antiherpetic resistance of mice has been found to achieve its maximum if herpes simplex antigens are introduced after the injection of the preparation of transfer factor.

Transfer factor therapy of thoracic sarcoidosis.

The authors repeatedly treated 59 patients with thoracic sarcoidosis with transfer factor (TF) since 1976. They utilized this therapy with TF from human tonsil lymphocytes (TFh) on account of the ineffectiveness of the corticosteroid treatment, because of the side effects of the corticosteroids, and as primary TF therapy, and to test an animal TF preparation from pig tonsil lymphocytes (TFp). In their observations only fraction II of the dialysable leukocyte extract was sufficient. Differences in the effectiveness between TFh and TFp do not exist on the whole. Our conclusion is that TF can stimulate the immunosystem of the patients, and can be an important mode of treatment. The mode of action is not clear.

Adult chronic granulomatosis disease-like neutrophil granulocyte disorder corrected by dialysable leukocyte extract.

A 47 year old female presented with a septic clinical picture including fever, abscesses, late cachexia, and unmanageable by disease. Similar characteristics to chronic granulomatosis disease (CGD) seriously decreased intracellular killing activity and chemiluminescence, granulomas in the histology, and the role of genetic factors were found, suggesting that our case is CGD-like disorder, manifested in an adult. Dialysable leukocyte extract (DLE) therapy, complemented with fresh normal plasma, resulted in a striking clinical improvement and there was an increase in the in vitro PMNL intracellular killing activity, too. Although it is generally accepted that DLE derives from monocytes and lymphocytes, it is possible that DLE is a family of DNA-oligopeptide molecules, including factors derived from PMNLs which are capable of influencing PMNL function, transferring information from normal cells. Our results also suggest that it would be worth trying DLE in patients with classic CGD.

De novo initiation of specific cell-mediated immune responsiveness in chickens by transfer factor (specific immunity inducer) obtained from bovine colostrum and milk.

Transfer factors (TF) were prepared from colostrum and milk of bovines previously immunized with antigens obtained from Coccidioides immitis, infectious bovine rhinotracheitis virus, or from the viral agents responsible for avian Newcastle disease, laryngotracheitis disease or infectious bursal disease. The ability of bovine TF to transfer specific cell-mediated immune responsiveness to a markedly xenogenic species was studied using specific pathogen free (SPF) and standard commercial (SC) chickens as model recipients. Cell-mediated immune responsiveness was documented using one or more of the following for each antigen (organism) studied: (a) an in vitro chicken leukocyte (heterophil) migration inhibition assay; (b) delayed-wattle reactivity; or (c) protection from clinical disease. Chicken TFs obtained from spleens of immune donors were evaluated in parallel to bovine TF's in selected comparative studies. Bovine TF also referred to as specific immunity inducer (SII), and chicken TF were found to initiate antigen-specific cell-mediated immunity de novo in previously non-immune SPF chickens as well as in SC chickens despite the presence of maternally acquired humoral antibody which may serve as a "barrier" to immunization of SC chickens when commercially available vaccines are administered by parenteral routes. Bovine TF's specific for laryngotracheitis virus or infectious bursal disease virus afforded protection equal to that found for commercially available vaccines. Bovine TF's action was rapid (less than a day) and of relatively long duration at least 35 days.

Chromatographic and enzymatic effects on transfer factor-like activity from human leukocytes and porcine spleen dialysate.

1. The effect of dialysable transfer factor (TFd), derived from human leukocytes or porcine spleen cells, was measured using Listeria resistance in mice. 2. The molecular weight range of substance(s) containing TF-like activity is in the less than 3500 MW dialysis fraction on the basis of the capacity of peritoneal macrophages to produce superoxide anion (O2-). This biological activity is removed by heating at 56 degrees C. 3. After Sephadex G-10 chromatography of dialysates the significant activities are found in fractions III and IV of human leukocyte dialysate and in fractions of II and III of porcine spleen dialysate. 4. From enzymatic studies, most of the protective activity of both human leukocyte and porcine spleen dialysate is based on the action of small-molecular weight structures containing peptides and/or polynucleotides.

[Ultrafiltration characteristics of immunobiological preparations]. / Nekotorye osobennosti ul'trafil'tratsii immunobiologicheskikh preparatov.

The removal of ammonium sulfate from the bulk product of fermented antitoxic serum by continuous diafiltration was not accompanied by changes in the stability of the solution. To concentrate immunoglobulin, eluted from DEAE cellulose, by diafiltration, the stabilization of the solution by adding sodium chloride at high concentration was necessary. The use of membranes purchased from different manufacturers and having similar selectivity characteristics permitted obtaining transfer factor preparations somewhat differing in their biological activity. The process of ultrafiltration, carried out in the atmosphere of compressed carbon dioxide, made it possible to obtain such preparations from donor blood plasma.

[Studies of an assay system for dialyzable leukocyte extracts (DLE)--influence of DLE on leukocyte migration inhibition test by HBsAg].

In order to examine the suitability of leukocyte migration inhibition test (LMIT) in the capacity of in vitro assay system for dialyzable leukocyte extracts (DLE), the effect of DLE on hepatitis B and its antigen-specificity, the migration inhibitory activities to purified hepatitis B surface antigen (HBsAg) was measured using the leukocyte MIF test with DLEs obtained from HBsAb-positive or HBsAb-negative blood. The direct LMIT using agarose plate was modified according to the technique of Clausen et al. In spite of our assay system was dose-dependent for PPD, a significant response for purified HBsAg was not observed. However, some meaningful migration inhibition appeared when HBsAg and DLE were added simultaneously to the migration cells. From these results, it is concluded that DLE has antigen-specific and/or antigen non specific influences to the cell-mediated immunity for HBsAg Though some problems remain, we think our results are interesting, since the assay system for DLE has not been established and our study is closely related to the effect of DLE concerning hepatitis B.

Effect of human leukocyte and other tissue dialysates on Listeria resistance and phagocytosis in mice.

The effect of dialyzable transfer factor (TFd) on Listeria resistance was measured by survival studies and by assessing phagocytic capacity of peritoneal macrophages. Unfractionated dialysates from human leukocytes (DLE), bovine liver, porcine spleen and kidney as well as saline were injected i.p. into NMRI mice 72 h before the i.p. injection of 1-3 x 10(6) Listeria organisms. The results show that DLE, porcine spleen and bovine liver dialysate increased the LD50 5-20 times. Porcine kidney dialysate had no effect on the survival of the mice. After the fractionation of porcine spleen dialysate on Sephadex G-10 column, a significant activity was found in two fractions, II and IX. When active fractions were given together (II + IX) i.p. three days prior to the infection with listeria organisms, the survival of mice increased significantly, whereas no effect was seen when the fractions were given i.v. and the bacteria i.p. Also the treatment with active fractions increased significantly the phagocytic capacity of peritoneal macrophages. Taken together, our results suggest that the Listeria protective substances seem to operate via monocyte activation.

Effect of long-term therapy with transfer factor in rheumatoid arthritis.

Specific immunotherapy with transfer factor (TF) was used in a chronic experiment in a group of 50 female patients with rheumatoid arthritis (RA) stage I-III. The patients were followed up for 24 months, clinical and biologic examinations being repeated every 3 months. In this period the patients received beside the basic nonsteroid antiinflammatory therapy, one unit TF every week over a period of 6 months then one until TF every month (10 patients) to the end of experiment. Of the 50 patients 15 (30%) did not respond to the therapy and the experiments had to be interrupted after 6 months. Excellent, very good and good results were obtained in 35 patients (70%). In 12 patients the response was good but the dose of TF had to be increased to two units/week in the first 6 months. In 13 patients the results obtained were very good and therapy with nonsteroid products + TF was continued even after the first 6 months. In 10 patients with RA stage I the results obtained were excellent and after 6 months the nonsteroid therapy could be interrupted and the therapy was continued only with one unit TF every month. The study confirmed the fact that specific immunotherapy with TF represents an important adjuvant in the treatment of rheumatoid arthritis (RA).