Polymer-encapsulated Schwannoma cells expressing human nerve growth factor promote the survival of cholinergic neurons after a fimbria-fornix transection.

Many investigators have recently used genetically modified primary fibroblasts of fibroblast cell lines (e.g., 3T3, 208F, or BHK cells) to deliver recombinant nerve growth factor (NGF) into the CNS. In the current study, SCT-1 cells, a Schwannoma cell line derived from a transgenic mouse, were transfected with a human NGF (hNGF) cDNA. After selection, these cells were encased within a polymer capsule and implanted into the ventricles of fimbria-fornix lesioned rats. Encapsulated, non-transfected cells served as controls. Results demonstrated that the hNGF transgene is expressed for at least 3 weeks after implantation. Moreover, the cells did not overgrow the capsule. Recombinant hNGF was able to save > 70% of lesioned cholinergic neurons, as assessed by NGF-receptor (NGFr) and choline acetyltransferase (ChAT) immunohistochemistry, from cell death. The number of cholinergic neurons in animals that received control capsules (i.e., nontransfected SCT-1 cells) was similar to lesion only animals (i.e., approximately 27% and approximately 33% for NGFr- and ChAT-positive neurons, respectively. These results show that SCT-1 cells can be used to deliver biologically active hNGF into the lesioned rat brain.

Clonal insulinoma cell line that stably maintains correct glucose responsiveness.

A number of pancreatic beta-tumor cell (beta TC) lines have been derived from insulinomas arising in transgenic mice expressing the SV40 T antigen gene under control of the insulin promoter. Some of these lines secrete insulin in response to physiological glucose concentrations. However, this phenotype is unstable. After propagation in culture, these nonclonal lines become responsive to subphysiological glucose levels and/or manifest reduced insulin release. Here we report the use of soft-agar cloning to isolate single-cell clones from a beta TC line, which give rise to sublines that maintain correct glucose responsiveness and high insulin production and secretion for > 55 passages (over a year) in culture. One of these clonal lines, denoted beta TC6-F7, was characterized in detail. beta TC6-F7 cells expressed high glucokinase and low hexokinase activity, similarly to normal islets. In addition, they expressed mRNA for the GLUT2 glucose transporter isotype and no detectable GLUT1 mRNA, as is characteristic of normal beta-cells. These results demonstrate that transformed beta-cells can maintain a highly differentiated phenotype during prolonged propagation in culture, which has implications for the development of continuous beta-cell lines for transplantation therapy of diabetes.

Two PC12 pheochromocytoma lines sealed in hollow fiber-based capsules tonically release L-dopa in vitro.

Two PC12 cell-derived lines have been studied following encapsulation into polymer-based hollow fibers with respect to secreted catecholamines and their metabolites. Cellular encapsulation provides a chronic microperfusion environment within which basally secreted PC12 products can be readily measured. Encapsulated PC12 cells grown and held under the conditions specified in this report basally release amounts exceeding their total cellular stores of the dopamine precursor L-DOPA and the electrochemically active dopamine metabolites DOPAC and HVA during 45-min static incubations. Under these same conditions, these cells release less than 0.1% of their total cellular store of dopamine. Depolarizing incubations enhance dopamine secretion eightyfold and enhance secretion of L-DOPA, HVA, and DOPAC about twofold. The relative composition of products basally secreted differs between PC12-derived cell lines, and an inverse relationship exists between basal release of L-DOPA and total cellular store of dopamine. These results further indicate that selected PC12 cell lines are potentially a source of both dopamine and L-DOPA in therapeutic cellular replacement applications.