h1

%0 Thesis
%A Zhou-Stache, Jingfang
%T Investigation of the effects of chinese herb medicine on TNF-[alpha] induced apoptosis in Huvec and Jurkat cells and their antioxidative effects
%C Aachen
%I Publikationsserver der RWTH Aachen University
%M RWTH-CONV-118957
%P III, 84 S. : Ill., graph. Darst.
%D 2002
%Z Aachen, Techn. Hochsch., Diss., 2002
%X The Chinese herb, Radix Salviae Miltiorrhizae (RSM), is being used in traditional Chinese medicine as a treatment for cardiovascular and cerebrovascular diseases. Several components of the plant extract from Salvia Mitorrhiza Bunge have been determined previously, two of which are protocatechuic acid (PAC) and protocatechuic aldehyde (PAL). Since anti-apoptotic therapies have been proposed to limit tissue damage in cardiovascular and cerebrovascular diseases, PAC and PAL effects on cell protection from apoptosis were investigated in this thesis. XTT assays were first used to quantify cell viability. We found that PAC and PAL inhibited TNF-alpha induced apoptosis of human umbilical vein endothelial cells (Huvec) and Jurkat cells in a concentration of 100 µM and 1mM respectively, when applied 2 hours prior to TNF-alpha exposition. To investigate molecular consequences on cellular signal transduction pathways, NF-kappaB reporter gene and DNA binding activities were investigated by luciferase assay and gel shift assay. Degradation of IkBalpha was determined by western blotting. The molecular studies revealed that PAC activated NF-kappaB with a maximal effect after 30 min of treatment. Inhibition of NF-kappaB action by MG132 and NF-kappaB inhibitory peptide suppressed the anti-apoptotic effect of PAC. Further, degradation of IkBalpha occurred in response to PAC treatment. Our results provide evidence that activation of NF-kappaB plays an important role in mediating the anti-apoptotic effect of PAC on HUVEC and Jurkat cells. Nevertheless, PAL protection from apoptosis triggered by TNF-alpha could not prevented by the treatment of MG132. Additionally, PAL did not induce IkappaBalpha degradation. The results implied that the anti-apoptotic action of PAL may be mediated by a molecular mechanism other than activation of NF-kappaB. In addition, a computer controlled flow channel system (Elias-c-) was tested in this thesis to study antioxidative capacities of herbal medicine. Effects of herbal extracts and single components on oxidatively impaired red blood cell (RBC) stiffness and relaxation time as well as on oxidative damage (H2O2, 2 mM) induced RBC-endothelial cell (EC) adhesion were investigated. Following H2O2 treatment (20 min), RBC became significantly stiffer and the relaxation time was reduced as compared with control. These changes were irreversible after re-incubating oxidatively damaged RBC in HEPES buffer. However, when oxidatively damaged RBC were re-incubated with 88.5 µM Tetramethylpyrazine (TMP), the H2O2-induced reduction in relaxation time turned back to control levels whereas the RBC stiffening did not. As mechanism we hypothesize, that TMP applied to hydrogen peroxide damaged RBC may partially reverse the hydrogen peroxide mediated formation of hemoglobin-spectrin complexes possibly leading to a normalization of the membrane viscosity. In the RBC-EC adhesion tests, a maximum (about three-fold as compared to control) increase in RBC-EC adhesion was obtained when both RBC and EC were treated with H2O2. This increase in adhesion was almost completely inhibited by 50 µM 3,4-dihydroxyphenyl lactate (Dhpl), when applied either prior to or after treatment with H2O2. The mechanism of the protection may be addressed in part to a direct free radical scavenging effect of antioxidants dissolved in the EC membrane. From a methodological point of view, the Elias-c in combination with appropriate cell types and experimental designs can be seen as a cellular bioassays to test herbal extracts with high laboratory efficacy.
%F PUB:(DE-HGF)11
%9 Dissertation / PhD Thesis
%U https://publications.rwth-aachen.de/record/56880