preproenkephalin and Pheochromocytoma

Most adrenal chromaffin cells synthesize opioids derived from proenkephalin but not from prodynorphin. However, human pheochromocytomas and the PC12 rat pheochromocytoma cell line synthesize dynorphins. The aim of this study was to confirm the presence of the authentic prodynorphin transcript and its dynorphin product in PC12 cells. We have found that the sequence of a 458 bp cDNA fragment derived from RT-PCR amplification of total PC12 RNA was in complete accordance with the published sequence of the equivalent region of the prodynorphin gene. It encodes the potent endogenous kappa opioid agonists alpha-neo-endorphin, dynorphin A and dynorphin B. Furthermore, immunoaffinity-purified PC12 cell extracts were subjected to RP-HPLC. Most of its IR-dynorphin eluted on a peak exhibiting the retention time of similarly treated rat anterior pituitary. The expression of the prodynorphin gene in pheochromocytomas can be explained as either the result of (a) the process of dedifferentiation of chromaffin cells to pheochromocytoma which may thus cause the expression of a previously unexpressed prodynorphin or that (b) those pheochromocytomas expressing the prodynorphin gene derive from the few, centrally located chromaffin cells, which express this gene even under normal conditions.

Topics: Adrenal Medulla; Animals; Chromatography, Affinity; Chromatography, High Pressure Liquid; Dynorphins; Endorphins; Enkephalins; Gene Expression; PC12 Cells; Pheochromocytoma; Pituitary Gland; Polymerase Chain Reaction; Protein Precursors; Rats; Sequence Analysis

Topics: Adrenal Gland Neoplasms; Dexamethasone; Enkephalins; Humans; In Vitro Techniques; Pheochromocytoma; Protein Precursors; RNA, Messenger; Tumor Cells, Cultured

We demonstrated the presence and the secretion in vivo and in vitro of immunoreactive preproenkephalin B-derived opioid peptides (alpha-neoendorphin, dynorphin and leumorphin) in human phaeochromocytomas. In seventeen human phaeochromocytomas and two human adrenal medullas, the tissue contents of immunoreactive preproenkephalin B-derived opioid peptides (alpha-neoendorphin, dynorphin and leumorphin) and leu-enkephalin were studied by specific RIAs. Compared with a remarkable wide distribution in amounts of immunoreactive leu-enkephalin (1063 +/- 437 pg/mg, mean +/- SE), small amounts of immunoreactive alpha-neoendorphin (22.6 +/- 6.4 pg/mg) and dynorphin (8.5 +/- 1.2 pg/mg) were detected in all seventeen human phaeochromocytomas and the two human adrenal medullas. Leumorphin-like immunoreactivity was detected in only four tumours. Gel chromatographic studies revealed the presence of preproenkephalin B-derived peptides and their high molecular forms. A significant positive correlation between the tumour tissue contents of immunoreactive alpha-neoendorphin and of dynorphin was observed. Nicotine (10(-5), 10(-4) mol/l) significantly stimulated the secretion of immunoreactive alpha-neoendorphin and dynorphin as well as leu-enkephalin and catecholamines from cultured human phaeochromocytoma cells. Administration of 1 mg of glucagon to a patient with medullary phaeochromocytoma induced a rapid increase in the plasma concentration of immunoreactive alpha-neoendorphin with a concomitant increase in plasma catecholamines. These results indicate the presence of preproenkephalin B-derived opioid peptides in human phaeochromocytomas and human adrenal medullas and their secretion in human phaeochromocytomas.

Topics: Adolescent; Adrenal Gland Neoplasms; Adult; Cells, Cultured; Dynorphins; Endorphins; Enkephalins; Epinephrine; Female; Glucagon; Humans; Male; Middle Aged; Nicotine; Norepinephrine; Pheochromocytoma; Protein Precursors; Radioimmunoassay

Tissue contents of immunoreactive met-enkephalin (Met-Enk), leu-enkephalin (Leu-Enk) and met-enkephalin-Arg-Gly-Leu (Met-Enk-AGL) were determined by specific RIAs in 17 pheochromocytomas and the levels of preproenkephalin A mRNA were compared in some of these tissues by Northern blot hybridization. Remarkably wide distributions in the amounts of immunoreactive Met-Enk, Leu-Enk and Met-Enk-AGL were observed in 17 pheochromocytomas. Medullary pheochromocytomas contained significantly larger amounts of immunoreactive Met-Enk, Leu-Enk and Met-Enk-AGL than did the extramedullary ones. Significant positive correlations of tumor tissue immunoreactivities between Met-Enk and Leu-Enk (r = 0.97, P less than 0.01) and between Met-Enk and Met-Enk-AGL (r = 0.97, P less than 0.01) were observed. No difference in size of mRNA of preproenkephalin A was observed in the pheochromocytomas. Medullary pheochromocytomas contained higher amounts of preproenkephalin A mRA than did the extramedullary ones. The quantities of preproenkephalin A mRNA in nine pheochromocytoma tissues significantly correlated with those of immunoreactive Met-Enk in these tissues (r = 0.94, P less than 0.01). These results indicate that the remarkable difference in tissue immunoreactivities of preproenkephalin A-derived opioid peptides is decided at the transcriptional level but not at the posttranslational level.

Topics: Adrenal Gland Neoplasms; Adult; Enkephalin, Leucine; Enkephalin, Methionine; Enkephalins; Female; Humans; Male; Middle Aged; Nucleic Acid Hybridization; Pheochromocytoma; Protein Precursors; Radioimmunoassay; RNA, Messenger