Immunological models in high dilution research following M Bastide.

In 1994, Madeleine Bastide described experimental models in immunology that were used during the 1980s to investigate high dilution effects on several biological systems. She classified the available papers in four categories: High dilutions of antigens; High dilutions of thymus, bursa and other hormones; High dilutions of cytokines; Immunopharmacological activity of silica. The studies about high dilutions of antigens were not continued after this period, but gave rise to a long process of a series of in vitro models on antigens and histamine dilutions, that led to the demonstration of the biological modulation effects of these preparations on basophil degranulation. During this process, a multi-centre study was performed, with a high degree of reproducibility among different independent laboratories. The studies about high diluted cytokines, thymulin and other hormones opened a new line of scientific investigation, about the regulatory properties of endogenous substances prepared according to homeopathic methods. The most frequently studied substance, thymulin, when administered to mice at 5cH potency, is able to improve the activity of phagocytes in different experimental situations, such as viral, bacterial and parasitic infections. The immunopharmacological activity of silica was demonstrated, at that time, as an in vivo illustration of the homeopathic 'similia principle'. More recently, studies on silica have assumed another focus: the putative role of silica as active contaminant present in high dilutions. This paper presents a follow-up summary on these items, considering the evolution of discoveries from 1994 to 2014.

[Difference of hypaconitine concentration in serum between cold-deficiency and normal mice].

OBJECTIVE: To investigate the difference of hypaconitine concentration in serum between normal and cold-deficiency mice after administration of aconite decoction. To analyze how the toxic dose of aconite decoction correlate to the metabolic environment. METHOD: Prepared cold-deficiency mice model, treated normal and cold-deficiency mice with aconite decoction for 14 days continuously, and then detected hypaconitine concentration in serum by HPLC along with survival ratio of mice on the first, seventh and fourteenth day. RESULT: After administration of aconite decoction for 14 days, the hypaconitine concentration in serum of cold-deficiency mice is close to that in normal mice. It showed aconite decoction has the ability of regulating metabolism environment, the hypaconitine concentration in serum of normal mice was higher on the seventh and fourteenth day than that on first day. It showed that aconite decoction can disturb metabolism environment of normal mice. It was also been observed that the range of variation of hypaconitine concentration in cold-deficiency mice was minor than that in normal mice during the fourteen days' administration. CONCLUSION: The difference of serum concentration in normal and cold-deficiency mice showed that there were different metabolic environments in two mice models, and the metabolic environment changed during administration. These results showed that the different toxic doses of aconite decoction were partially due to the different metabolic environments.

Cordycepin induced eryptosis in mouse erythrocytes through a Ca2+-dependent pathway without caspase-3 activation.

Cordyceps sinensis is a prized traditional Chinese medicine and its major component cordycepin is found to have anti-leukemia activities. However, its cytotoxicity in erythrocytes was unclear. To examine the effect of cordycepin on the induction of eryptosis (an apoptosis-like process in enucleated erythrocytes), flow cytometric assays based on membrane integrity and asymmetry were employed. For comparison, analyses were performed in parallel with two other anti-leukemia agents, indirubin 3'-monoxime (IDM) and As2O3. We found that at the IC50 against leukemia HL-60, cordycepin elicited eryptosis while IDM and As2O3 showed no erythrotoxicity in mouse erythrocytes. Mechanistically, cordycepin increased the [Ca2+]i and activated mu-calpain protease in a dose-dependent manner. Yet, no caspase-3 activation was observed in the cordycepin-treated erythrocytes. When extracellular Ca2+ was depleted, both the cordycepin-induced eryptosis and mu-calpain cleavage were suppressed. Our study therefore demonstrated for the first time that cordycepin induces eryptosis through a calcium-dependent pathway in the absence of mitochondria and caspase-3 activation.

Role of vitamin D3-binding protein in activation of mouse macrophages.

When mouse peritoneal nonadherent (lymphocytes) cells were treated with lysophosphatidylcholine (lyso-Pc) and cultured with adherent cells (macrophages) in 1% fetal calf serum (FCS)- or adult mouse serum (AMS)-supplemented medium for 3 h, markedly enhanced phagocytic and superoxide-generating capacities of macrophages were observed. Stepwise cultivation of lyso-Pc-treated B cells and untreated T cells with an FCS-supplemented medium generated a macrophage-activating factor (MAF), whereas cultivation of lyso-Pc-treated B cells alone in AMS-supplemented medium was sufficient to generate the MAF. The accumulated evidence suggests that lyso-Pc-inducible beta-galactosidase of B lymphocytes and the Neu-1 sialidase of T lymphocytes modified the bovine serum vitamin D3-binding protein (DBP) to yield the MAF, a protein with N-acetylgalactosamine as the remaining sugar. In contrast, the lyso-Pc-inducible beta-galactosidase of B cells alone modified mouse DBP to yield the MAF. These observations led us to conclude that bovine DBP carries a trisaccharide composed of N-acetylgalactosamine, galactose, and sialic acid, whereas mouse DBP carries a disaccharide composed of N-acetylgalactosamine and galactose. Thus, macrophages of a T-cell-deficient nude (BALB/c nu/nu) mouse and a T-cell Neu-1 sialidase-deficient SM/J mouse were efficiently activated by administration of lyso-Pc.

Binding of mouse red blood cells (MRBC) by human thymocytes: augmentation of rosette formation by phospholipase C treatment.

The formation of rosettes with MRBC is not an exclusive property of chronic lymphocytic leukemia cells and of human B-lymphocytes. Human thymus cells and the HD-MAR T-cell line were also found to be capable of forming rosettes with MRBC. The binding of MRBC by thymocytes differed from that of sheep (SRBC), rabbit (RRBC) and human (HRBC) erythrocytes by a number of parameters: monoclonal antibodies against the E-receptor inhibited the attachment of SRBC, RRBC, and HRBC, but not of MRBC to thymus cells. The presence of EDTA had the opposite effect, abrogating only the attachment of MRBC to thymus cells but not of RBC from the other sources. Exposure of thymus cells to pronase eliminated their capacity to bind SRBC, RRBC, and HRBC, but only slightly inhibited the attachment of MRBC. Treatment with Vibrio cholerae neuraminidase enhanced the formation of rosettes with SRBC, RRBC, and HRBC, but had no effect on the formation of rosettes with MRBC. Exposure of thymus cells and of the HD-MAR cells to phospholipase C enhanced the proportion of rosettes formed with MRBC, but had no effect on the binding of other RBC. Treatment with either phospholipase A2 or phospholipase D had no such effect. The binding of MRBC by Raji cells was not increased by phospholipase C treatment. The present study indicates that the binding of MRBC by human thymus cells is mediated by receptors distinct from those involved in the binding of SRBC, RRBC, and HRBC and also from those mediating the binding of MRBC to human B-cells.

Autologous transplantation with circulating hemopoietic stem cells.

Circulating stem cells exist in sufficient numbers in mouse, dog, and man to allow collection and transplantation after ablative treatment. Preclinical studies in the mouse have shown a low concentration, with a transplantation potential ratio of bone marrow to blood of 1:100. The ratio of circulating stem cells to bone marrow stem cells is more favorable in the dog (1:10-20). Recent pilot studies carried out in different centers with 10 patients have shown that this approach is feasible in man, too. It appears that 5 X 10(8) mononuclear cells/kg of body weight collected by seven or eight leukapheresis procedures of about 4 hrs each is sufficient for fast hemopoietic recovery after marrow ablative treatment. Potential advantages of the use of blood stem cells over bone marrow stem cells are the decreased likelihood of contamination with malignant cells, the avoidance of general anesthesia, and the infusion of immunocompetent cells, which might hasten immunorecovery in the autologous setting.