EFFECTS OF 2ND MESSENGER SYSTEM ACTIVATION ON FUNCTIONAL EXPRESSION OF TYROSINE-HYDROXYLASE FUSION GENE CONSTRUCTS IN NEURONAL AND NONNEURONAL CELLS

A genomic clone for rat tyrosine hydroxylase (TH) was isolated and a fragment containing 503 bp upstream of the transcription start site was sequenced. The BamHI/AluI fragment was inserted into a plasmid carrying the coding sequence for bacterial chloramphenicol acetyltransferase (CAT). Another construct with the 5' sequence truncated to -151 bp also was prepared. When these were introduced into several mammalian cell lines, including C6 glioma, BE(2) neuroblastoma, CV-1 or Ltk- fibroblasts, different basal levels of CAT expression were observed. In the fibroblast lines, THCAT constructs were not expressed unless the cells were treated with forskolin or TPA. However, the low basal expression was not correlated to endogenous expression as THCAT constructs expressed comparably in BE(2)C, HeLa, and C6 glioma. Treatment of any of the cell lines with forskolin, TPA, or a combination of the two agents stimulated the expression by at least two-fold in all cell lines and the maximally induced levels were at least 10-fold over promoterless controls. These data indicate that the essential promoter elements as well as those conferring responsivity to cyclic AMP reside within 151 bp of the transcription start site. However, the array of elements regulating cell-type expression lie, at least in part, beyond the 500-bp region examined. Further, a role for phosphorylation in the regulation of basal and induced transcription of TH is suggested.X1128sciescopu

In vitro Demonstration of Cancer Inhibiting Properties from Stratified Self-Organized Plasma-Liquid Interface

Abstract Experiments on plasma-liquid interaction and formation of thinly stratified self-organized patterns at plasma-liquid interface have revealed a nontrivial cancer-inhibiting capability of liquid media treated at self-organized interfacial patterns. A pronounced cancer suppressing activity towards at least two cancer cells, breast cancer MDA-MB-231 and human glioblastoma U87 cancer lines, was demonstrated in vitro. After a short treatment at the thinly stratified self-organized plasma-liquid interface pattern, the cancer inhibiting media demonstrate pronounced suppressing and apoptotic activities towards tumor cells. Importantly, this would have been impossible without interfacial stratification of plasma jet to thin (of several µm) current filaments, which plays a pivotal role in building up the cancer inhibition properties. Furthermore, thinly stratified, self-organized interfacial discharge is capable to efficiently control the ROS and RNS concentrations in the cancer-inhibiting media. In particular, abnormal ROS/RNS ratios are not achievable in discharges since they do not form stratified thin-filament patterns. Our findings could be tremendously important for understanding the cancer proliferation problem and hence, the potential of this approach in tackling the challenges of high cancer-induced mortality should be explored