Objective:
Evaluate the role of TNF' and IL-6 in the observed anti-inflammatory effects of two local hepatic herbal remedies (Urtica pilulifera and Hypericum triquetrifolium) in communities of Israel and elsewhere in the Middle East to help develop novel therapies that specifically interfere with the overproduction of the pro-inflammatory mediators TNF' and IL-6 while preserving their beneficial regenerative effects.

Approach:
The effects of the plant extracts will be investigated in the first phase using cocultures of cells from human hepatocyte cell line (HepG2) and from the human monocytes cell line THP1. In the second phase of our proposed study we will apply primary rat hepatocytes and primary rat Kupffer cells. The cocultures will be maintained under "in vivo like", three-dimensional cell culture conditions using DegraPol-foams as a three dimensional cell carrier. Lipopolysaccharides stimulated cells will be treated with various concentrations of plant extracts for 24h and 48h. The following end points will be measured: Cell viability and cytotoxicity; cell proliferation; the secretion of Il-6, TNFa 'and nitric oxide as well as the secretion of albumin. Positive plant extracts will be further fractionated and their effects will be estimated as mentioned before using primary isolated cells. Effects of plant extracts will be also assessed in the absence and presence of macrophage inhibitor. Since TNFa and IL-6 are involved in both hepatic inflammation and liver regeneration, the implementation of anti-cytokine therapy requires cautions. For example, adequate levels of TNFa and IL-6 are critical for liver regeneration. Inhibition of all TNFa or IL-6 activities in a patient with severe liver disease could potentially inhibit recovery. We have the following four specific objectives: 1. To evaluate the role of TNFa 'and IL-6 in anti-inflammatory effects of Urtica pilulifera and Hypericum triquetrifolium. Therefore, nontoxic concentration of plant extracts will be used. 2. To develop novel therapy that specifically interferes with the overproduction of TNFa and IL-6 while preserving their beneficial regenerative effects. 3. To perform in vitro studies to assess the absence of toxicity of the two plants. 4. The creation of new in vitro test system, in which the cell-to-cell interactions is considered. Therefore, cells will be cultured in multi-layers in a three dimensional pattern. This new in vitro test system will be used for the studying of mechanisms and toxicity of cell-active xenobiotecs.