The role of calcium channels in the growth regulation of normal and neoplastic neuroendocrine lung cells

Author

Barbara Jane Sheppard

Date of Award

5-1999

Degree Type

Dissertation

Degree Name

Doctor of Philosophy

Major

Comparative and Experimental Medicine

Major Professor

Hildegar M. Schuller

Committee Members

David O. Slauson, Kevin Hahn, George Kabalka

Abstract

The binding of nicotine or nicotine-derived carcinogenic nitrosamines at the alpha7 nicotinic acetylcholinergic receptor (α₇nAChR) on the surface of human small cell lung carcinomas (SCLC) and pulmonary neuroendocrine cells (PNEC) stimulates release of the autocrine growth factor 5-HT. The release and reuptake of 5-HT leads to cell proliferation. Thus, nicotinic agonists acting at the cell surface α₇nAChR, which is a functional Ca²⁺ channel, initiate a mitogenic autocrine loop with growing evidence of downstream activation of intracellular kinase-driven signaling cascades involving the raf kinases . The central role of Ca²⁺ as a second messenger in kinase-driven signaling pathways strongly suggests that cellular tools for controlling Ca²⁺ movement, such as VOCC, are upregulated in SCLC cells derived from chronic smokers.

It was shown using flow cytometry that nicotine and the nicotine-derived carcinogenic nitrosamine nitrosamine 4-(methylnitrosamino)-1-(3-pyridyl)-1- butanone (NNK) treatment ofSCLC-derived NCI-H69 cells stimulated the influx of extracellular Ca²⁺ through α₇nAChR followed by Ca²⁺ movement through voltage operated calcium channels (VOCC). The maximal Ca²⁺ influx was observed with much lower doses of NNK (100pm) compared to nicotine (1μΜ) indicating a much greater affinity ofNNK for the α₇nAChR. PNEC exposed to nicotine or NNK demonstrated a much smaller amount of Ca²⁺ influx indicating that the neoplastic SCLC cells exhibit an increased responsiveness to both compounds.

This work demonstrated, using relative quantitative reverse transcription - polymerase chain reaction (RQ RT-PCR), high expression levels by untreated NCIH69 cells of all three "L, N, and P" VOCC subtype cDNA targets examined. In contrast, fetal hamster PNEC demonstrated a significant expression of only the "P" subtypes. The NCI-H69 exhibited this greatly enhanced expression of all the VOCC subtypes due to the massive upregulation of the serotonergic autocrine mitogenic pathway following years of exposure to nicotine and nicotine-derived carcinogenic nitrosamines such as NNK and to the hypoxia of concurrent chronic lung disease. It was also shown that NCI-H69 display a subtype-dependent alteration in VOCC subtype expression levels when subchronically exposed to constant uninterrupted nicotine, NNK or hypoxia.

These findings strongly support the hypothesis that the NCI-H69 cell line derived from the SCLC of a chronic smoker demonstrated a massively upregulated ability to manipulate Ca²⁺ levels using the a.rnAChR Ca²⁺ channel and VOCCs. Unique levels or patterns of VOCC expression by SCLC may provide clinically useful targets in the search for better diagnostic and therapeutic options with which to combat this aggressive tumor type.

Recommended Citation

Sheppard, Barbara Jane, "The role of calcium channels in the growth regulation of normal and neoplastic neuroendocrine lung cells. " PhD diss., University of Tennessee, 1999.
https://trace.tennessee.edu/utk_graddiss/8961