Azadirachta Indica as a herbal medicine

NEEM EXTRACT MODULATE THE CYTOKINE NETWORKS OF THE IMMUNE AND REPRODUCTIVE SYSTEMS

Extracts of Neem leaves, bark and seeds have a long history as folk remedies for a number of diseases. Neem oil extracted from seeds could prevent pregnancy in rodents and monkeys. Because of the cytokine interactions between the immune system and pregnancy, this led to the suggestion that Neem influences immune cytokine networks. This was strengthened when it was shown that in vivo treatment with Neem oil resulted in higher IFNγ levels when spleen cells were subsequently stimulated with Con A in culture. IFN γ is a central component of the Th1 cytokine pattern, secreted by the TH1 subset of CD4 T cells that carry out cell-mediated immune reactions. In contrast, TH2 cells secrete 1L4, 1L5 and IL1O, and provide help for antibody production. TH1 and TH2 responses cross-inhibit each other, mainly via their characteristic cytokines.
TH1 cytokines are harmful for pregnancy, but enhance the immune response against a number of intracellular pathogens. Neem may act by directly or indirectly enhancing the production of TH1 cytokines, modifying both pregnancy and disease resistance.

1. Neem extract Effects on Cytokine Synthesis, B Cell Proliferation and Parasite Infection:
a). Neem extract preferentially enhances synthesis of the Th1 cytokine IFNγ by spleen cells.
b) Neem extract enhances GM-CSF synthesis.
c) Neem extract stimulate B cell proliferation and antibody production.
d) Neem extract stimulate production of cytokines and other mediators by macrophages.
e) Neem extract effects on Leishmania infection in vitro and in vivo.

2. Analysis of the mode of action of the active component in Neem on immune cells:
a) Mechanism of action of Neem on cytokine synthesis and differentiation of T cells.
b) Neem effects on macrophage pro-and anti-inflammatory mediator synthesis.
c) Neem stimulation of B cell proliferation and antibody production and switching.
3. Neem effects on infections and fertility:
a) Neem enhancement/inhibition of Leishmania infection.
b) Neem effects on placental cytokine production in vitro and in vivo.
c) Neem effects Pre implantation regulated by immune cytokines.

Neem effects on placental cytokine production in vitro and in vivo. The effects of Neem on placental cytokine production will also be investigated. Previous experiments demonstrated substantial levels of TH2 cytokines in placental tissue. The cytokine synthesis ceases immediately after tissue disruption and in vitro culture, and so two approaches will be taken to evaluate placental responses to Neem. In situ hybridization and/or antibody staining will be used to determine the location of TH2 (and possibly Th1)
cytokine synthesis. The possibility of establishing culture conditions for placental cells or tissue that allow continued cytokine synthesis. Possible modifications to the basic culture system include the use of explants instead of single-cell cultures, culture of defined cell types (e.g. trophoblast or uterine epithelial cells), and the addition of cytokines and hormones to mimic
the in vivo environment. If culture conditions can be found that allow the same pattern of cytokine expression as in vivo, this will provide a very powerful system for future characterization of the influences of Neem and other compounds on the placental cytokine regulatory networks.
Neem effects in Pre-implantation: Krishnan has recently found that Leishmania infection significantly compromises concurrent pregnancy. The C57B116 TH1 response can increase the resorption rate, consistent with previous studies showing that IFN γ and NK cells were deleterious to the maintainance of pregnancy. A more unexpected finding was that the early response of C57B1/6 mice against Leishmania could also cause a block in
pregnancy at some stage before implantation. It is tempting to speculate that this effect could also be due to cytokine regulatory changes. Interestingly, others have shown Neem oil prevented pregnancy at a stage before implantation, provided the Neem was administered at an early time. Although it is difficult to compare the effects of Neem seed oil with the water-soluble Neem extract. The
hypothesis that fertilization or implantation is susceptible to the same cytokine regulatory pathways as the maintenance of pregnancy after implantation. This will be tested by first comparing the water soluble extract with seed oil for their effects on fertilization/implantation. In vivo treatments with anti-IFNγ and anti-TNF will be used to determine whether these TH1 cytokines mediate the Neem-induced inhibition of the phase of reproduction. In vivo (the work of two students, Li Li and Yew Hon Lai) the effect of exogenous IL1O for potential blocking of Neem-induced reduction of implantations.