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Urbanization with reduced microbial exposure is associated with an increased burden of asthma and atopic symptoms. Conversely, environmental exposure to endotoxins in childhood can protect against the development of allergies. Our study aimed to investigate whether the renaturation of the indoor environment with aerosolized radiation-detoxified lipopolysaccharide (RD-LPS) has a preventative effect against the development of ragweed-induced Th 2 -type airway inflammation. To explore this, cages of six-week-old BALB/c mice were treated daily with aerosolized native LPS (N-LPS) or RD-LPS. After a 10-week treatment period, mice were sensitized and challenged with ragweed pollen extract, and inflammatory cell infiltration into the airways was observed. As dendritic cells (DCs) play a crucial role in the polarization of T-cell responses, in our in vitro experiments, the effects of N-LPS and RD-LPS were compared on human monocyte-derived DCs (moDCs). Mice in RD-LPS-rich milieu developed significantly less allergic airway inflammation than mice in N-LPS-rich or common environments. The results of our in vitro experiments demonstrate that RD-LPS-exposed moDCs have a higher Th 1 -polarizing capacity than moDCs exposed to N-LPS. Consequently, we suppose that the aerosolized, non-toxic RD-LPS applied in early life for the renaturation of urban indoors may be suitable for the prevention of Th 2 -mediated allergies in childhood.

Stem cell-based therapies raise hope for cell replacement and provide opportunity for cardiac regenerative medicine and tumor therapy. Extracellular vesicles are a membrane-enclosed intercellular delivery system with the potential to improve the therapeutic efficacy of the treatment of a variety of disorders. As the incidence of breast cancer continues to rise, radiotherapy has emerged as a leading treatment modality. Radiotherapy also increases the risk of coronary heart disease and cardiac mortality. In a chest-irradiated mouse model of cardiac injury, we investigated the effects of local irradiation. We found an increased lethality after 16 Gy irradiation. Importantly, radio-detoxified LPS (RD-LPS) treatment prolonged the survival significantly. By flow cytometry, we demonstrated that upon administration of RD-LPS, the number of bone marrow-derived endothelial progenitor cells increased in the bone marrow and, in particular, in the circulation. Furthermore, mass spectrometry analysis showed that RD-LPS altered the proteomic composition of bone marrow cell-derived small extracellular vesicles (sEVs). RD-LPS treatment increased interferon-induced transmembrane protein-3 (IFITM3) expression markedly both in bone marrow cells and in bone marrow cell-derived small extracellular vesicles. This is the first study to demonstrate that radio-detoxified LPS treatment induces an increase of circulating endothelial progenitor cells (EPCs) in parallel with a reduced radiotherapy-related mortality. While the total number of bone marrow-derived extracellular vesicles was significantly increased 24 h after treatment in the RD-LPS groups, the number of endothelial progenitor cells was reduced in animals injected with GW4896 (a chemical inhibitor of exosome biogenesis) as compared with controls. In contrast to these in vivo results, in vitro experiments did not support the effect of sEVs on EPCs. Our data raise the intriguing possibility that IFITM3 may serve as a marker of the radio-detoxified LPS treatment.

The observations on the protective effect of bacterial endotoxin in farm-derived cow's milk on childhood asthma and allergy are contradictory. The aim of this study was to determine the endotoxin levels in 'farm-derived whole raw' and 'processed shop' sources of cow's milk, and to test how the temperature and storing conditions might alter their endotoxin concentrations. Milk was collected from farms and shops. The level of endotoxin was measured by micro (gel-clot) Limulus amebocyte lysate test expressed as EU/ml. The concentration ranges of endotoxin were much higher and more widely scattered in the samples of whole raw farm milk than in the processed shop milk. Cold storage or heating increased the endotoxin concentrations in all samples of farm milk, but not in the processed shop milk. These results show that elevated levels of endotoxin in raw farm milk samples can occur from the cowshed or be formed during storage. In processed shop milk, storage does not cause any changes in the amount of endotoxin. Therefore, it is consistent that the handling and storage of raw milk alters the endotoxin concentrations, which may explain previous contradictory findings regarding the beneficial modulating effects on innate immunity toward allergy prevention in early childhood., (© The Author(s) 2014 Reprints and permissions: sagepub.co.uk/journalsPermissions.nav.)

It is well demonstrated that serial endotoxin injections produce endotoxin tolerance and elevate the natural immunity/resistance. However, such injections may also have harmful effects such as high fever, hypotension and abortion. For this reason endotoxin (LPS) injections are not suitable to enhance nonspecific resistance in endotoxin-sensitive species like man. Various techniques have been designed (physical, chemical, etc.) for the detoxification of endotoxins while the beneficial effects are maintained. Perhaps one of the best detoxification techniques is the treatment with ionizing radiation. The irradiation of LPS with 60Co (150 kGy) decreased its toxicity in a dose-dependent manner. Such radio-detoxified endotoxin (RD-LPS) preparations show decreased toxicity whereas the beneficial effects were preserved. Irradiation causes marked chemical alterations in LPS, such as a decrease of glucosamine, ketodeoxyoctonic and fatty acids. A single parenteral RD-LPS injection prevents various forms of shock in experimental animals. This preparation has a membrane-stabilizing effect, and thereby it can prevent the membrane-damaging effect of LPS and of some cytostatic agents. Unlike endotoxin, RD-LPS has little hypotensive effects, and the pretreatment with this preparation can prevent practically all the hemodynamic changes induced by LPS. LPS plays an important role in the pathogenesis of intestinal syndrome of radiation disease, which may be prevented by RD-LPS pretreatment up to 70% in rats. RD-LPS retains the adjuvant activity of LPS, and it serves as a good adjuvant for inactivated virus vaccines. RD-LPS can also evoke the regeneration of the immune system in irradiated animals. The decrease of nonspecific resistance in immunodeficient or immunosuppressed patients is the most important cause of opportunistic infections that may lead to sepsis like in endotoxaemia and pneumonia. Organ transplant recipients commonly die of septicaemia. Antilymphocyte serum (ALS) is used in such patients as an immunosuppressant. The augmentation of natural resistance and the induction of endotoxin tolerance are of major significance in such patients. In ALS-treated rats RD-LPS induces also tolerance against the lethal dose of LPS. This demonstrates that in spite of the suppressive effect of ALS on T-lymphocytes the induction of LPS tolerance (the enhancement of natural resistance) remains normal. Facultative pathogenic organisms may flourish and cause disease when specific and nonspecific resistance is impaired. RD-LPS can produce a significant proliferation of lymphoid cells in germ-free animals which are immunodeficient. Many other beneficial effects are preserved by RD-LPS preparations, such as the activation of macrophages and of the reticuloendothelial system and antitumor activity. On the basis of these favorable experimental results, RD-LPS has been tested on 350 surgical patients suffering from gastrointestinal tumors, patients suffering from acquired immunodeficiency syndrome (AIDS) and cancer patients treated with CYSPLATIN. RD-LPS treatment prevented sepsis and activated the bone marrow function in these patients.

The discovery of the physico-chemical host defence is closely connected with the endotoxin research. It is well known that the toxic effects of endotoxins under experimental conditions can be induced only when they are administered parenterally. However, in naturally occurring entero-endotoxemic diseases (e.g. septic and various shocks, etc.), the endotoxin is absorbed from the intestinal tract. The cause and mode of translocation have been unknown. The generally used experimental shock models differ from natural diseases only in the mode by which endotoxin enters the blood circulation. If the common bile duct of rats was chronically canulated (bile-deprived animals) orally administered endotoxin was absorbed from the intestinal tract into blood circulation and provoked endotoxin shock. This translocation of endotoxins and the consequent shock can be prevented by sodium deoxycholate or natural biles. The bile acids split the endotoxin macromolecule into atoxic fragments. A similar detoxifying detergent action plays a significant role in host defence against infectious agents with outer lipoprotein structure (e.g. so-called 'big' viruses). This defence mechanism of macroorganisms based on the detergent activity of bile acids (end-products of the cholesterol metabolism) is called as physico-chemical defence system. Therefore, bile deficiency and the consequent endotoxemia are important components in the pathogenesis of certain diseases (e.g. sepsis, intestinal syndrome of radiation disease, hepato-renal syndrome, parvovirus infection, herpes, psoriasis, atherosclerosis, etc.). Bile acids may be used for the prevention and/or therapy of the above mentioned clinical conditions.

The authors have tested the hypothesis that the deficiency of bile acids and the consequent endotoxin translocation might play a role in the pathogenesis of psoriasis. Under normal conditions the bile acids act as detergents (physico-chemical defense) and can protect the body against enteric endotoxins by splitting them into nontoxic fragments and thus preventing the consequent release of cytokines [Persp. Biol. Med. 21 (1977) 70]. A total of 800 psoriasis patients participated in the study and 551 were treated with oral bile acid (dehydrocholic acid) supplementation for 1-8 weeks. The efficacy of the treatment was evaluated clinically and also by means of the Psoriasis Area Severity Index (PASI score). During this treatment, 434 patients (78.8%) became asymptomatic. Of 249 psoriatics receiving the conventional therapy, only 62 (24.9%) showed clinical recovery during the same period of time (P<0.05). The curative effect of bile acid supplementation was more pronounced in the acute form of psoriasis (95.1% of the patients became asymptomatic). Two years later, 319 out of the 551 acute and chronic psoriasis patients treated with bile acid (57.9%) were asymptomatic, compared to only 15 out of the 249 patients (6.0%) receiving the conventional treatment (P<0.05). At the end of the 2-year follow-up, only 10 out of 139 acute psoriasis patients (7.2%) receiving the conventional therapy and 147 out of 184 bile acid treated patients (79.9%) were asymptomatic (P<0.01).To conclude, the results obtained suggest that psoriasis can be treated with success by oral bile acid supplementation presumably affecting the microflora and endotoxins released and their uptake in the gut.

The authors demonstrated significant curative effect of bile acids (Suprachol; Acidum dehydrocholicum) in 551 psoriatic patients. The clinical efficiancy was evaluated by means of PASI-score (Psoriasis Area Severity Index). During this treatment (1-8 weeks) 434 patients (78.8 per cent) became asymptomatic. However, the traditional therapy resulted in 62 patients (24.9 per cent) of 249 sick persons a recovery (p < 0.05). In acute form of psoriasis (184 patients) this curative effect of bile acids was elevated (95.1 per cent). Two years later 319 patients (57.9 per cent) of bile treated 551 people were asymptomatic in contrast with 15 people (6.0 per cent) of 249 traditional treated patients (p < 0.05). In same time among the patients which were treated in acute form of psoriasis 10 (7.2 per cent) of 139 controls and 147 (79.9 per cent) of 184 bile-treated individuals were asymptomatic (p < 0.01). On the basis of their clinical observations (digestive disorders, ultrasonical confirmed gallbladder complaints, etc.) authors supposed that the deficiency of bile acids and the consecutive endotoxin translocation might play a role in the pathogenesis of psoriasis. In normal conditions the bile acids as detergent (physico-chemical defense) can protect the body against enteral endotoxins while split them to atoxic fragments and so preventing consecutive cytokin liberation.

Innate resistance is mediated by non-immune defense and by natural immunity. Non-immune defense includes diverse mechanisms (e.g., physico-chemical defense by bile acids). Natural killer (NK) cells, gamma delta T lymphocytes and CD5+ B lymphocytes are key mediators of natural immunity. These cells utilize germ-line coded receptors that recognize highly conserved, homologous epitopes (homotopes). Typically, it is not the antigen, but cytokines and hormones that regulate the level of NK-mediated cytotoxicity. These include interleukin-2, interferons, prolactin and growth hormone. Less is known about gamma delta T lymphocytes. CD5+ B lymphocytes produce germ-line coded antibodies (predominantly IgM) that are polyspecific, and able to recognize a great variety of microorganisms, cancer-cells and self-components. Antigen is not an effective stimulus for natural antibody (NAb), but bacterial lipopolysaccharide (LPS) is. During the acute phase response (febrile illness) the T-cell-regulated adaptive immune response is switched off and natural immune mechanisms are amplified several hundred to a thousand times within 24-48 hours (immunoconversion). This immunoconversion is initiated by immune-derived cytokines, and involves profound neuroendocrine and metabolic changes, all in the interest of host defense. Immune recognition is assured by natural antibodies and by some liver-derived acute phase proteins, such as C-reactive protein or endotoxin-binding protein, the level of which is elevated in the serum. Thus, natural immunity is essential for a first and last line of defense and the neuroendocrine system is an important promoter of this activity.

Simultaneous HHV-6A infection can activate HIV-1 latency and promote AIDS progression, but in this process the effects of HHV-6A induced soluble mediators on HIV-1 have not been studied yet. Recently, supernatants of HSB-2 cultures infected with HHV-6A and/or treated with endotoxin have been filtered virus free at time intervals until the cytopathic effect developed. Biological activity of some cytokines which might participate in HIV-1 activation was quantitated. Filtered supernatants were mixed into CEM-ss cultures, which had been HIV-1 infected at 1:1 cell:virus ratio, subsequently HIV-1 replication was quantitated and compared to controls. Supernatants filtered during the first 96 hours of HHV-6A replication without visible cytopathic effect augmented HIV-1 syncytium formation by tenfold, reverse transcriptase activity by threefold, p24 antigen production by 6-fold. Filtered supernatants obtained at onset of HHV-6A cytopathic effect did not modify HIV-1 replication. HSB-2 cultures produced no IL-2, and IFN-gamma induced by endotoxin diminished HIV-1 replication. HHV-6A delayed IFN-gamma release. An increase in the tumour necrosis factor activity upon the effect of HHV-6A and endotoxin was not parallel to HIV-1 activation. The putative mediator, different from those above which characterisation is in progress, might transmit similar transactivating effects between immune cells of lymph nodes and circulation.

Using ionizing radiation the author and co-workers produced a detoxified endotoxin preparation (Tolerin) which seems to be a suitable product for the increase of natural immunity (nonspecific resistance)-including activation of bone marrow in immunosuppressions, immunodeficiencies-protection against various types of shocks-radiation injury, septic/endotoxic shock, etc.- and increase of immunogen effect of antigens (e.g. inactivated virus vaccines) as an immunoadjuvant in human beings and experimental animals.

The toxic effects of endotoxin--the cell wall component of Gram negative intestinal bacteria--under experimental conditions can be induced only when they are administered parenterally. However, in naturally occurring enteroendotoxemic diseases (e.g. septic and various shocks, etc.), the endotoxin absorbs from the intestinal tract. The cause and mode of translocation was unknown. The generally used experimental shock models differ from natural diseases only in the mode by which endotoxin enters the blood circulation. If the common bile ducts of rats were chronically cannulated (bile deprived animals) perorally administered endotoxin was absorbed from the intestinal canal into blood circulation and provoked endotoxin shock. The translocation of endotoxins and consequent shock can be prevented by sodium deoxycholate or natural biles. The bile acids can split the endotoxin macromolecule (atoxic fragments). A similar destructive detergent action might will be a significant factor against potential infectious agents with lipoprotein outer structure (e.g. so-called "big" viruses). This defense mechanism of macrooganisms based on the detergent activity of bile acids is called as physico-chemical defense system. On the basis of this knowledge the bile acids might be used in the prevention and therapy of some clinical processes (e.g. hepatorenal syndrome; psoriasis).

The plasma level of endotoxin was determined in 116 healthy blood donors in this laboratory. After a routine physical and laboratory investigations the endotoxin level was determined with Limulus amebocyte lysate assay (LAL test) by the chromogenic kinetic method of Bio-Whittaker Co. (USA). Its sensitivity was 0.005-50 EU/ml. The plasma level of endotoxin in most of healthy donors was less than 1 EU/ml (in the range of 0.01-1.0 EU/ml), but always measurable. The average +/- S. D. was 0.128 +/- 0.215 EU/ml. Because of the high standard deviation and high range of values, the data were distributed to two groups with the means of 0.05 +/- 0.022 EU/ml and 0.294 +/- 0.186 EU/ml. The difference between the groups was significant found (p < 0.001). In conclusion, endotoxin can be measured in plasma of healthy individuals.

The lipopolysaccharide endotoxin macromolecules are cell wall's components of the Gram negative bacteria. The endotoxins are produced by Gram negative bacteria of intestinal flora. If the endotoxins are translocated from the intestinal tract to the circulation or injected into bloodstream, they elicit (depending from the quantity of endotoxin), slight or serious effects (e.g. endotoxin shock). In the effects of endotoxin certain cell populations (e.g. thrombocytes, macrophages, leukocytes, etc.), certain organs and organ-systems (e.g. liver, spleen, bone marrow, endocrine and lymphoreticular systems etc.) are involved. Effects of endotoxin are produced by mediators (e.g. endotoxin binding proteins, cytokines, prostaglandins, prostacyclins, NO etc.). The endotoxin sensitivity of vertebrate organisms is dependent from the phylogenetical status of the species. Most sensitive species is the human. Generally accepted that endotoxin has an important role in the pathogenesis of septic shock. In other pathological processes (e.g. intestinal syndrome of radiation disease, Gram negative infections, various shock forms etc.) are supposed or proved the role of endotoxins. Lead acetate induced endotoxin hypersensitivity or LAL methods are good tools for demonstration of the role of endotoxin in the pathogenesis of various processes. For this reason, the experimental endotoxin shock is used a model of septic and other shocks.

The effect of trace element combination, Béres Drop Plus (BDP) on the immune response of rats following single treatment with cytostatic drug (5-fluorouracil, 5-FU) was tested. Animals were treated with 5-FU and SRBC simultaneously, but separately. Rats were pretreated with BDP for 21 days. The body and spleen weight, furthermore the number of spleen cells and antibody producing cells were determined. The antibody titres in blood serum were also measured. The immune system of rats, 5 days following the 5-FU treatment, showed considerable regeneration. Pretreatment of rats with BDP had a beneficial effect on all parameters investigated.

The stress situations, including medical intervention (e.g. operations, antitumor drugs, irradiation, etc.) decrease the nonspecific resistance of the body. In these situations patients people have greater chance to get an opportunistic infection than healthy ones. The restoration or elevation of the activity of immune system in injured patients is a very important task of medicine. Minute amounts of bacterial endotoxin (LPS)--given parenterally--can elevate the nonspecific resistance. Unfortunately this beneficial influence is associated with noxious properties. Irradiation (60 Co-gamma; 150 kGy) is a good technique for the detoxification of LPS. The radiodetoxified endotoxin (RD-LPS) preparation (so-called TOLERIN) is less toxic but its beneficial properties is preserved. On the basis of animal experiments and clinical trials TOLERIN could be a suitable preparation for regeneration of the lymphoreticular-immune system and elevation of nonspecific resistance.

It has long been known that the toxic effects of endotoxins under experimental conditions can be induced only when they are administered parenterally. However, in naturally occurring enteroendotoxemic diseases (e. g. septic and intestinal ischemic shocks) the endotoxins--which are produced by gram negative members of intestinal flora-, absorb from the intestinal tract to the blood circulation and can elicit pathological processes. It is an important distinction between natural and experimental endotoxin shock. If the common bile duct of rats were chronically cannulated a significant amount of perorally administered endotoxin was absorbed into the blood. This endotoxin shock can be prevented by bile acids. The physiological surfactants, the bile acids, are important facts in the defense of macroorganisms against endotoxins (physico-chemical defense). The production and passage of bile acids depend from the function of liver and the cholecystokinine (CCK) synthesis of small intestine wall. If the bile (bile acid) content of the intestinal canal decreases the endotoxin can translocate to the body and elicits toxic symptoms. So most important parts of defense against endotoxins in natural conditions are the CCK and bile acids. The consequence of damage of liver (place of bile acid synthesis) or small intestine (place of CCK synthesis) is the absorption of endotoxins.

We studied the effect of a trace element combination (Béres Drop Plus; BDP) on the immune response of rats, with intact and with injured immune system. Rats were treated with different doses of BDP for 26 days. The immune system was injured by 7.0 Gy whole body gamma irradiation on the first day of BDP treatment. Rats were immunized on the 31st day of the BDP treatment with sheep red blood cells. In the spleen the cell count and the number of antibody producing cells were determined. In the serum the titre of hemolysin was tested. In rats with intact immune system all used doses of BDP induced elevation of the immune response. The moderate high doses were especially effective. In rats with injured immune system the effect was less. Only the 250 microliters/kg body weight dose could elevate the immune response, and the highest (500 microliters/kg) dose reduced the response.

Restoration of immune functions through promoting cell cycle might delay acquired immunodeficiency syndrome development. Therefore, stimulation of peripheral lymphocytes of human immunodeficiency virus-1 infected patients in successive clinical stages was studied by phytohaemagglutinin and other stimulants. In vitro blastogenesis was quantitated by 3H-thymidine uptake. Stimulation by phytohaemagglutinin decreased in patients with AIDS related complex to 63.1%, with AIDS to 13.6% of control values. Small amount of recombinant interleukin-2 or indomethacin solely not promoting lymphocytes, increased response to phytohaemagglutinin minimally. Alone ineffective methyl-ester and methyl-phosphonate inosine derivatives augmented phytohaemagglutinin-response of controls and patients with AIDS related complex by approx. 1.5-fold, but the effect in the case of AIDS patients was minimal. Radio-detoxified endotoxin alone or in combination with phytohaemagglutinin stimulated lymphocytes of both controls and patients with AIDS related complex slightly. Lymphocyte stimulation of patients with AIDS related complex was augmented in concentration-dependent manner, and by synergic effect it approached phytohaemagglutinin-stimulated blastogenesis of controls. Anergy due to human immunodeficiency virus-1 infection damages synchronisation of secondary messenger systems induced on cell surface receptors, therefore their selective influence by recombinant interleukin-2 or indomethacin is less efficient. Inosine derivatives promote cell cycle by inhibiting cyclic adenosine 3',5'-monophosphate production. In the early stage of virus infection, radio-detoxified endotoxin might bind to receptors of immature T cells and facilitate cell cycle through cyclic guanosine 3',5'-monophosphate stimulation. The clinical trials of radio-detoxified endotoxin (Tolerin) have already been launched.

The membrane bound form of the catalytic subunit of adenylate cyclase of chicken embryo brain has been found earlier to be rather radioresistant [28]. The radiation induced changes of G proteins of membrane preparations of 19 day old chicken embryo brains were investigated in this work. The activation of catalytic subunit of adenylate cyclase by G protein dependent activators (Gpp/NH/p and NaF) was found elevated at lower radiation doses (0-400 Gy), while both basal enzyme activity (measured without any activator) and the activity measured in the presence of activators decreased at higher doses (above 800 Gy). Heterogeneity of GTP binding sites measured with 3-H-Gpp/NH/p a GTP-ase resistant GTP analogue was observed in the case of control (with Kd1 = 0.0663 +/- 0.034 mumol/l, Bmax = 0.0079 +/- 0.0022 nmol/ml and Kd2 = 2.038 +/- 0.4779 mumol/l, Bmax2 = 0.0291 +/- 0.0017 nmol/ml). The lower affinity high capacity binding sites seemed to be more radiosensitive, than the higher affinity sites. A marked decrease was observed in the number of low affinity binding sites above 200 Gy and these low affinity binding sites practically disappeared after irradiation with 400 Gy. At high doses (above 1600 Gy) the catalytic subunit was damaged, too. On the basis of the decrease of low affinity binding sites together with an increase in activation of the catalytic subunit via G proteins one can conclude that it is caused by radiation induced damage of Gi protein that can be more radiosensitive, than Gs protein.