Relation between activation of NF-kappa B and chemotherapy induced apoptosis of leukemic cells / 中华肿瘤杂志

<p><b>OBJECTIVE</b>To analyze the relation between activation of NF-kappa B and chemotherapy induced apoptosis of leukemic cells and the effect of vincristine (VCR) on them.</p><p><b>METHODS</b>Electrophoretic mobility shift assay (EMSA) was used to detect the activation of NF-kappa B and tunel DNA electrophoresis was adopted to observe the apoptosis induced by cytosine arabinoside (Ara-C) and etopside (Vp-16) in P388 leukemic cells.</p><p><b>RESULTS</b>The activation of NF-kappa B induced by Ara-C and Vp-16 was obviously correlated to apoptosis in P388 cells. VCR (0.1 micromol/L) could suppress activation of NF-kappa B by 52% and 63% and significantly increase the apoptosis by 89% and 123% as induced by Ara-C (100 micromol/L) and Vp-16 (100 micromol/L). The activity of NF-kappa B could be found in P388 cells before being exposed to chemotherapeutic agent.</p><p><b>CONCLUSION</b>Chemotherapeutic agents can induce apoptosis and activation of NF-kappa B of P388 cells. The mechanism of VCR potentiating chemotherapeutics induction of leukemia cell apoptosis may be related to its suppression of the NF-kappa B activity in the P388 cells.</p>

Novel multi-probe RNase protection assay set for detection of endotoxin associated receptors gene expression / 中华创伤杂志(英文版)

<p><b>OBJECTIVE</b>To construct the multi-probe ribonuclease protection assay (RPA) template set to be used for detecting expression patterns of MD-2, TLR4, CD14 mRNAs in human peripheral blood mononuclear cells.</p><p><b>METHODS</b>The designed cDNA fragments of the three genes were generated by polymerase chain reaction (PCR) using specific primers and directionally cloned into EcoR I and Hind III sites of expression plasmid pSP72 containing the T7 promoter, the linearized plasmids was used as template to synthesize anti-sense RNA probes. Then we extracted total RNA from peripheral blood mononuclear cells (PBMC) and detected the dynamic expression patterns of the three genes with RPA method.</p><p><b>RESULTS</b>The proper sequence and orientation of the template set were confirmed by sequencing and the template set was successfully used to assay TLR4, MD-2 and CD14 mRNAs in human PBMC. The results showed that the three detected genes decreased transiently 1-3 hours after 100 ng/ml LPS stimulation.</p><p><b>CONCLUSIONS</b>These new RPA multi-probe set provided valuable tool for the simultaneous quantitative determination of expression of TLR4, CD14 and MD-2 mRNAs in both constitutive and inducible types.</p>