ANP secretion from small cell lung cancer cell lines: a potential model of ANP release.

Although atrial distension is widely accepted as the primary stimulus for atrial natriuretic peptide (ANP) release, a number of agonists are also known to induce its secretion. The mechanisms underlying these processes are not well understood. Studies of this nature are hampered by the inherent difficulty in culturing homogeneous populations of cardiac myocytes in sufficient quantities to perform molecular investigations. For this reason, we have examined the possibility of using other cell types as a model of ANP release. It has been reported that a number of tumor samples from small cell lung cancer (SCLC) patients express the ANP gene. Characterization of a large number of cell lines derived from SCLC tumor samples indicated that two of these cell lines, OS-A and SHP-77, secrete ANP at rates of approximately 10(-20) g.cell-1.min-1. This is a sufficient quantity to facilitate secretion studies using a perifusion system. We have demonstrated that ANP is released through regulated secretory pathways, as the Ca2+ ionophore A-23187, arginine vasopressin (AVP), and the sodium ionophore, monensin, were capable of modifying secretion rates. High-pressure liquid chromatography (HPLC) analysis indicated that the primary secretory product is ANP-(99-126), the circulating form of this hormone. Intracellularly, both ANP-(99-126) and ANP-(1-126) were present, suggesting the synthesis and appropriate cleavage of pro-ANP-(1-126). Because both of these cell lines have doubling times in the range of 3-5 days, they could serve as a rapidly proliferating and easily maintainable supply of homogeneous tissue for release studies.(ABSTRACT TRUNCATED AT 250 WORDS)

Formation of N-methyl protoporphyrin in chemically-induced protoporphyria. Studies with a novel porphyrogenic agent.

1-[4-(3-Acetyl-2,4,6-trimethylphenyl)-2,6-cyclohexanedionyl]-O-eth yl propionaldehyde oxime (for short ATMP) is a novel porphyrogenic agent causing hepatic protoporphyria in the mouse. Mice given a single dose of the drug showed 24 h later a 70% inhibition of liver ferrochelatase and marked accumulation of protoporphyrin. These changes were not seen in similarly treated rats, guinea pigs, hamsters or chick embryos. A green pigment was isolated from the liver of mice treated with ATMP and identified by its electronic absorption spectrum and chromatographic properties on HPLC as N-methyl protoporphyrin. The ATMP pigment markedly inhibited the enzyme ferrochelatase in vitro, thus supporting its identification as N-methyl protoporphyrin. Two inhibitors of liver cytochrome P450, compound SKF 525-A and piperonyl butoxide, when given before ATMP, afforded protection against ATMP-induced porphyria and production of N-methyl protoporphyrin, suggesting a role of cytochrome P450 in the induction of the metabolic disorder. The most likely interpretation for these findings is therefore that ATMP is metabolized in the mouse to a reactive species, which in turn alkylates the haem moiety of liver cytochrome P450, thus producing N-methyl protoporphyrin. This inhibits ferrochelatase and, as a secondary response, protoporphyrin accumulates. This pathway of metabolism to the postulated reactive metabolite presumably does not occur to a significant extent in the other species examined and hence is the likely basis for the species difference in protoporphyria.

Isolation of two N-monosubstituted protoporphyrins, bearing either the whole drug or a methyl group on the pyrrole nitrogen atom, from liver of mice given griseofulvin.

1. A hepatic green pigment with inhibitory properties towards the enzyme ferrochelatase has been isolated from the liver of mice treated with griseofulvin and identified as N-methylprotoporphyrin. 2. All four structural isomers of N-methylprotoporphyrin have been demonstrated to be present, NA, where ring A of protoporphyrin IX is N-methylated, being the predominant isomer. 3. In addition to N-methylprotoporphyrin, a second green pigment, present in far greater amounts, was also isolated from the liver of griseofulvin-treated mice. This second green pigment is also an N-monosubstituted protoporphyrin, but in this case the substituent on the pyrrole nitrogen atom appears to be intact griseofulvin rather than a methyl group. 4. The fragmentation of this adduct in tandem m.s. studies suggests that griseofulvin is bound to the pyrrole nitrogen through one of its carbon atoms and further suggests that N-methylprotoporphyrin may arise as a secondary product from the major griseofulvin pigment.