Immunomodulating properties of mistletoe extracts

Effects of mistletoe extracts on immunocompetent cells of the non-specific and specific immune system such as NK cells, monocytes, macrophages, antigen-presenting cells, T-lymphocytes (T-cells) including T-helper cells, neutrophil and eosinophilic granulocytes, dendritic cells and a number of cytokines has been well established.

Originally, mistletoe lectin I (ML I) was thought to be responsible for the various immunological reactions, but today it is known that, in addition to other mistletoe lectins such as ML III and chitin-binding ML, other constituents such as viscotoxins, oligo- and polysaccharides also have an immunomodulating effect [175, 194, 195, 211, 212].

Activation of the non-specific immune system upon subcutaneous injection causes an acute-phase reaction with an increase in neutrophil granulocytes, while activation of the specific immune system is characterised by the production of antibodies against mistletoe constituents (activation of B-cells), an increase in the number of T-helper cells and an increase in eosinophilic granulocytes in the blood.

Dendritic cells (DC) are potent antigen-presenting cells. They transmit the information obtained from dendrites to T- and B-lymphocytes and activate them. In vitro experiments have shown that ML-containing mistletoe extracts can accelerate the maturation of dendritic cells and neutralise the immunosuppressive effect of tumour cell signals on dendritic cells [213].

 

Last update: August 10th, 2020/AT

Effects on cells of the non-specific immune system

The activation of cells of the non-specific immune system such as NK cells, macrophages and granulocytes by mistletoe extracts by an acute phase reaction has now been well documented by numerous in vitro and in vivo studies. Similarly, an increase in inflammatory cytokines such as tumour necrosis factor (TNF-alpha), IL-1 or IL-6 was observed under exposure to mistletoe extracts both in vitro and in tumour patients as well as in healthy volunteers, both for lectin-rich and lectin-poor or viscotoxin-rich extracts [211, 214, 215, 216].

In a randomised double-blind placebo-controlled study of 47 healthy volunteers who received Iscador Qu special (ML-rich), Iscador P (ML-poor) or placebo (physiological saline solution) for 12 weeks, it was shown that the administration of ML-rich extract significantly increased the production of granulocyte/macrophage-colony stimulating factor (GM-CSF) by peripheral blood mononuclear cells (PBMC) [215]. GM-CSF is responsible for the maturation, recruitment and activation of granulocyte and monocyte precursor cells from the bone marrow and is an important factor for the release of eosinophilic granulocytes from the bone marrow [217].

Therefore, the increased release of GM-CSF could explain both the activation of cells of the non-specific immune system and the increase in leukocytes (especially neutrophils) and eosinophilic granulocytes observed during therapy [211, 212]. There are also indications that the administration of mistletoe extracts prevents postoperative suppression of granulocytes and NK cells [218, 219].

 

Last update: August 10th, 2020/AT

Influence on monocytes and macrophages

The monocyte/macrophage system can be activated directly and indirectly by various mistletoe agents (total extracts, isolated mistletoe lectins, Kuttan's peptides, oligosaccharides, polysaccharides) in different ways. Antigen-presenting functions are stimulated, co-stimulating molecules are more strongly expressed while phagocytosis activity and cytotoxicity are increased. Furthermore, cytokines are induced and the antitumoural efficacy of macrophages/monocytes is augmented [220].

Various investigations have shown that mistletoe lectins preferably bind to monocytes/macrophages. Binding causes an increase in intracellular calcium and stimulates the synthesis and release of cytokines (TNF-α, IL-1α, IL-1β, IL-6) both by single lectins and by isolated A and B chains [221]. When macrophages were incubated with mistletoe lectins, a dose-dependent increase in their phagocytosis capability was recorded [202].

The antitumoural effects of mistletoe extracts have frequently been observed in connection with an increase in the number of monocytes/macrophages. A single administration of mistletoe activated macrophages was sufficient to induce antitumoural effects in animal models [175].

In addition, the transfer of in vivo activated macrophages from healthy mice to tumour-bearing animals doubled the survival time of tumour-bearing mice. This effect could not be achieved with non-activated macrophages [202]. It is also assumed that the anti-angiogenetic effect of mistletoe extracts is caused by the release of TNF-α from macrophages [175].

 

Last update: August 10th, 2020/AT

Influence on granulocytes

Mistletoe extracts can directly activate granulocytes in vitro. Here the viscotoxins have proven to be particularly effective by significantly enhancing both phagocytosis and oxidative burst in human granulocytes. All viscotoxins and also various total mistletoe extracts were effective in this way, but not isolated mistletoe lectins.

Viscotoxin-free mistletoe extracts can also significantly activate granulocytes (oxidative burst and phagocytosis). If granulocytes were incubated with mistletoe extracts and viscotoxins, an additive increase in granulocyte activation occurred, with two different routes of granulocyte activation seeming possible: On the one hand, granulocytes seem to be activated by mistletoe extract independently of viscotoxins or mistletoe lectins; on the other hand, another activation pathway seems to be induced by the total mistletoe extract and viscotoxin [175, 202, 222].

Influence on NK cells

The immunomodulating effect of mistletoe constituents (total extracts, polysaccharides, lectins, Kuttan's peptides, viscotoxins) is also characterised by an increase in the number and activity of NK cells, which can kill their targets, such as tumour cells, virus-infected cells or microorganisms, by releasing cytotoxic substances and inducing apoptosis.

Mistletoe activated NK cells also show a stronger antitumoural effect than non-mistletoe activated NK cells in animal experiments [175, 202, 223]. For example, in an experiment on mice injected with melanoma cells, a strong decrease in metastasis was observed in animals who were injected with in vivo activated spleen cells from healthy mistletoe treated mice four hours after tumour cell injection. In addition, their survival time was significantly increased [224].

Furthermore, the same group of researchers could demonstrate that spleen cells activated with mistletoe extract in vitro significantly reduced metastasis and prolonged survival time of the mice. In contrast, the administration of normal, non-mistletoe-activated splenocytes had no effect [224, 225, 226].

Single constituents of mistletoe also have an effect on NK cells in vitro. For example, the polysaccharide rhamnogalacturonan isolated from the mistletoe plant led to an increase in the cytotoxicity of NK cells via a specific bridge bond between effector cells and tumour cells.

Similarly, oligosaccharides indirectly increased the cytotoxicity of CD56+ NK cells by releasing lymphokines [202].

In a non-cytotoxic concentration, isolated viscotoxins also led to a significant increase in the cytotoxicity of NK cells compared to various tumour cell lines, whereas the mechanism has not yet been explained [227].

In contrast to the other mistletoe constituents, the cytotoxic properties of isolated mistletoe lectins predominate against single lymphocyte subpopulations. Thus, in an analysis of the lymphocyte subsets after 72 hours of incubation, a significant selective decrease in the NK cell fraction (CD16+ / CD56+ CD3) was observed in cell culture already at a mistletoe lectin concentration of 1ng/ml in the sequence ML III > ML II > ML I. Within the other T-cell subsets, however, no significant changes occurred. Likewise, the ratio between T- and B-cells remained the same. The preferential killing of NK cells could be completely suppressed by anti-mistletoe lectin serum.

Thus, among lymphoid cells NK cells appear to be a particularly mistletoe lectin-sensitive population [228]. In a study involving 40 patients with an operable mamma or colorectal carcinoma, the effect of mistletoe therapy on the absolute number of NK cells and their activity against both K562 cells and the patient's own tumour cells was investigated. The study also examined whether there was a correlation with the clinical course and quality of life of the patients. The absolute number of NK cells in peripheral blood increased. Patients without progression had a significantly higher NK cell activity against K562 cells than patients with progression. Within the group with progression only patients with stage IV showed a reduced cell function. The NK cell activity against K562 cells was not related to the number of NK cells, but a negative correlation between NK cell activity and tumour progression was found. A correlation between NK cell function and clinical course or quality of life was not found [229].

Last update: August 10th, 2020/AT

Effects on cells of the specific immune system

Mistletoe extract-specific antibody production and lymphocyte proliferation

As early as by the end of the 1980s it could be shown that volunteers who received a subcutaneous injection of mistletoe extract formed antibodies against ML I [230], since the injection of mistletoe extracts confronts the immune system with foreign antigens.

It was also shown that therapy with mistletoe extracts leads to a specific proliferation of lymphocytes, i.e. both the humoral and cellular immune systems are activated. This induction of a specific immune response was initially mainly attributed to ML I. However, it is now known that ML II, ML III or viscotoxins can also stimulate lymphocyte proliferation and antibody formation [212, 231, 232].

These antibodies occur in tumour patients under mistletoe therapy and also in healthy volunteers who have been treated with mistletoe extracts over a longer period of time. The antibodies are predominantly of the IgG isotype (IgG1 and IgG3), but the anti-ML I and anti-ML III antibodies may in rare cases also be of the IgE type [211, 212, 215, 231]. 

In later years another lectin, the so-called chitin-binding mistletoe lectin (cbML), was identified from mistletoe extracts. Antibodies belonging solely to the IgG class are also induced against this lectin during mistletoe therapy in tumour patients and healthy volunteers.

An increase in proliferation could also be observed when incubating lymphocytes with cbML in vitro. An interesting observation was that antibodies against cbML could also be detected in individuals who had never been in contact with mistletoe extracts. This is in contrast to the immunological reactions to ML I, ML III or viscotoxins which have been detected only in individuals who have had contact with mistletoe extracts. Probably cross-reactions are responsible for this [211, 212, 233].

 

Last update: August, 10th, 2020/AT

Influence on dendritic cells

Dendritic cells are the most important antigen-presenting cells of the human immune system, building a bridge between the unspecific and specific immune system. They play an important role in the management of tumour disease and are produced either from monocytes or from myeloid precursor cells [234, 235].

In vitro studies with human blood cells have shown that ML-containing mistletoe extracts can accelerate the maturation of dendritic cells [236] and that lymphocytes co-incubated with dendritic cells were stimulated by them [234].

In addition, it has been shown in vitro that mistletoe extracts can neutralise the immunosuppressive effect of tumour cell factors on dendritic cells [236], which might suggest a basic principle how mistletoe preparations could act against tumours via the immune system.

In the course of mistletoe therapy B-cells activated by dendritic cells produce antibodies against the various antigens of the mistletoe extract such as ML antibodies, viscotoxin antibodies etc. [232], which partially neutralise the effect and thus the local reactions may decrease over the course of a constant mistletoe therapy [237].

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