George Feuer

Action of Pregnancy and Various Progesterones on Hepatic Microsomal Activities

(1979). Action of Pregnancy and Various Progesterones on Hepatic Microsomal Activities. Drug Metabolism Reviews: Vol. 9, No. 1, pp. 147-169.

Failure of various drugs to induce drug-metabolizing enzymes in extrahepatic tissues of the rat

Abstract The effect of a tranquilizer (chlorpromazine HCl), a diuretic (hydrochlorthiazide), an antihydroxylation agent (metyrapone), and known inducers of hepatic microsomal drug-metabolizing enzymes (phenobarbital, 3-methylcholanthrene, 4-methylcoumarin) on drug metabolism in various organs of the rat has been studied. In all organs tested (liver, brain, kidneys, intestines, adrenals) various drug-metabolizing enzymes (aminopyrine demethylase, aniline hydroxylase, coumarin 3-hydroxylase, hexobarbital oxidase) could be demonstrated at various levels. However, the liver was the only organ in which drug metabolizing enzymes were induced by these compounds in various degrees. Even the hydroxylase inhibitor, metyrapone, increased hepatic hydroxylation. There was no change in the activity of these enzymes in any other organs. It appears, therefore, that the specific pharmacologic action of various drugs on target organs may not be connected with induction of drug metabolism.

3-Hydroxylation of coumarin or 4-methylcoumarin by rat-liver microsomes and its induction by 4-methyl-coumarin given orally

A new drug-metabolizing enzyme that hydroxylates coumarin in position 3 of the ring has been demonstrated in the liver of the rat. The enzyme is localized in the microsomal fraction. It requires O2 and reduced coenzyme (either NADH or NADPH). The pH optimum is 7.4, Km for coumarin is 9 · 10−4 and 4-methylcoumarin is 2.8 · 10−3M; SKF 525A is an inhibitor. Treatment of the microsomes with Triton X-100 or phospholipase A reduced the enzyme activity. The enzyme was induced in rats by the oral administration of 4-methylcoumarin but coumarin had no inducing effect. The activity returned to normal within 2 weeks of cessation of treatment. Actinomycin D prevented the inducing effect of 4-methyl-coumarin.

The effect of phenobarbital and carbon tetrachloride on fatty acid content and composition of phospholipids from the endoplasmic reticulum of rat liver

Abstract The fatty acid content and composition of hepatic microsomes of separated smooth and rough components and of isolated phosphatidylcholine and phosphatidylethanolamine fractions were studied in male albino rats treated with phenobarbital or carbon tetrachloride. Both test compounds significantly altered the fatty acid composition of the endoplasmic reticulum. The total amount was significantly raised by phenobarbital and reduced by carbon tetrachloride. Phenobarbital enhanced palmitic, stearic, arachidic, palmitoleic, linoleic, eicosenoic, eicosadienoic, eicosatrienoic, eicosapentenoic, docosatrienoic, and docosahexenoic acids. Carbon tetrachloride diminished all these, excluding palmitic and palmitoleic acids. The fatty acid content of rough microsomes was significantly increased by phenobarbital and decreased by carbon tetrachloride, while in smooth microsomes fatty acids were raised by phenobarbital but mainly unaffected by carbon tetrachloride. In microsomal phosphatidylcholine fractions, phenobarbital significantly elevated oleic, linoleic, eicosatrienoic, arachidonic, eicosapentenoic, docosapentenoic, and docosahexenoic acids, whereas all these were significantly reduced with carbon tetrachloride. In phosphatidylethanolamine fractions, phenobarbital increased palmitoleic, oleic, linoleic, and arachidonic acids; carbon tetrachloride elicited opposite effects on these acids. Phenobarbital increased and carbon tetrachloride reduced the fatty acid content in the phosphatidylcholine fraction of rough membranes. Opposite effects were seen in oleic, linoleic, arachidonic, and eicosapentenoic acids. Both test compounds brought about similar changes in the fatty acid composition of the phosphatidylethanolamine fractions of rough microsomes. In smooth microsomes, phosphatidylcholine fatty acids were significantly enhanced by phenobarbital and reduced by carbon tetrachloride. The fatty acid content of phosphatidylethanolamine was increased by phenobarbital, mainly manifesting in palmitoleic, oleic, linoleic, arachidonic, docosapentenoic, and docosahexenoic acids. Carbon tetrachloride elicited no major change in this fraction. Phenobarbital increased the production of unsaturated fatty acids, whereas carbon tetrachloride elevated the relative amount of saturated fatty acids. The saturated/unsaturated fatty acids ratio was reduced by phenobarbital and increased by carbon tetrachloride, and thus may indicate a selective difference between an inducer and hepatotoxin on fatty acid synthesis of the hepatic endoplasmic reticulum.

Progesterone binding by the hepatic endoplasmic reticulum of the female rat

Abstract Progesterone-binding by hepatic microsomal fractions of normal female rats was measured by using saturation analysis and by competition studies applying the charcoal adsorption technique. Scatchard analysis revealed the presence of a single binding component, with an equilibrium association constant varying between 0.17 and 0.27 × 109 l/mol. The concentration of binding sites showed variations from 1.4 to 3.4 pmol/mg protein. The binding of [3H]-progesterone to microsomes was highly specific. Relative binding affinities of cortisol, estradiol, 3β-hydroxy-5α-pregnan-20-one, 16α-hydroxyprogesterone and testosterone were low, between 1.8 and 9.1%. The microsomal progesterone receptor was characterized by the assessment of temperature stability and the effect of pH changes. The binding of [3H]-progesterone by microsomes represents a direct association without the involvement of a cytosol receptor and transfer processes. Since progesterone is mainly metabolized by hepatic microsomes, the existence of the progesterone-microsome complex may represent an intermediate in the mechanism that regulates the interaction between drugs and steroids in the drug-metabolizing enzyme system.

Morphological and biochemical changes in the liver of various species in experimental phospholipidosis after diethylaminoethoxyhexestrol treatment

Abstract The mechanism of druginduced experimental phospholipidosis was studied in several species by the administration of diethylaminoethoxyhexestrol. Rabbits, rats, mice, dogs, and guinea pigs developed microscopic and biochemical abnormalities, while hamsters were less affected. In the liver of affected species characteristic subcellular changes were found, accompanied by phospholipid accumulation. Hepatic lesions consisted of concentric lamellar bodies with varying degrees of osmic affinity, representing secondary lysosomes characterized by cytochemical methods. Accumulation of these bodies was also seen in Kupffer, endothelial, and biliary epithelial cells. The intensity of the changes was related to species susceptibility. Biochemical studies revealed an overall increase of total phospholipids in the affected species, together with changes in the relative distribution of individual phospholipids and the appearance of unidentified components. The activity of microsomal drug metabolizing enzymes and microsomal phospholipid synthesis were diminished. The lesions closely resembled those observed in man after treatment with diethylaminoethoxyhexestrol and are related to altered phospholipid metabolism with subsequent changes in microsomal drug metabolizing enzyme activity.

The effect of phenobarbital and carbon tetrachloride on fatty acid content and composition of phospholipids from rat liver.

Abstract The administration of phenobarbital or carbon tetrachloride to rats caused various changes in hepatic fatty acid content and composition. Phenobarbital elicited no effect on the total amount of fatty acids but significantly decreased myristic, pentadecanoic, and arachidonic acids and increased eicosatrienoic, eicosapentenoic, lignoceric, and docosatrienoic acid. In contrast, carbon tetrachloride enhanced significantly the total content and several components such as pentadecanoic, palmitic, palmitoleic, oleic, linoleic, arachidic, eicosenoic, eicosadienoic, eicosatrienoic, docosapentenoic, lignoceric and docosahexenoic acids. It elicited no effect on arachidonic acid. Unsaturated fatty acid moieties participating in the structure of these phosphatides were increased by phenobarbital and diminished by carbon tetrachloride. Phenobarbital caused a reduction in the ratio of saturated/unsaturated fatty acids mainly because of the decreased palmitic and increased oleic, linoleic, eicosatrienoic, arachidonic, docosapentenoic, and docosahexenoic acids. The significant variation brought about by phenobarbital and carbon tetrachloride on tissue fatty acids and in particular on fatty acid composition of phosphatidylcholine and phosphatidylethanolamine fractions reflects the opposing effects of these compounds on the liver cell. The major action of phenobarbital and carbon tetrachloride is associated with changes of the endoplasmic reticulum. Thus, their contrasting effect on fatty acid composition and metabolism may suggest that the disposition of lipid constituents plays a determinant role in the hepatic action of foreign compounds.

Differential action of progesterones on hepatic microsomal activities in the rat.

Abstract A significant reduction was found in the activity of drug-metabolizing enzymes (aminopyrine N -demethylase and coumarin 3-hydroxylase) and glucose 6-phosphatase in hepatic microsomes after the administration of reduced derivatives of progesterone (5α-pregnane-3β,-ol-20-one, 5β-pregnane-3α-ol-20-one, 5α-pregnane-3β,20β-diol and 5β-pregnane-3α,20α-diol) to rats. These steroids slightly raised inosine diphosphatase activity. On the other hand, 16α-hydroxyprogesterone and pregnenolone-16α-carbonitrile significantly increased drug metabolism and slightly elevated glucose 6-phosphatase. The contrasting action of the different progesterone derivatives was associated with changes in microsomal phospholipid synthesis. Pregnanolone and pregnanediol significantly decreased the de novo incorporation of [ 14 C-Me]- l -methionine into microsomal phospholipids, mainly manifesting in phosphatidylcholine, phosphatidylethanolamine and lysophosphatidylcholine fractions; reduced the activity of S -adenosyl- l -methionine:microsomal-phosphatidylethanolamine methyl transferase; and caused a reduction of total microsomal phosphatidylcholine:phosphatidylethanolamine ratio. In contrast, 16α-hydroxy-progesterone and pregnenolone-16α-carbonitrile increased the de novo synthesis of microsomal phospholipids, methyl transferase activity and the ratio of total microsomal phosphatidylcholine: phosphatidylethanolamine. Treatment of rats with reduced progesterone derivatives diminished microsomal progesterone hydroxylation in the 16α- and 6β-position and raised progesterone Δ 4 -5α-dehydrogenase activity measured in vitro . On the other hand, 16α-hydroxyprogesterone and pregnenolone-16α-carbonitrile elevated progesterone hydroxylation. Considering these opposite effects it can be postulated that in the rat the induction of drug-metabolizing activity of the hepatic endoplasmic reticulum might be controlled by a balance displayed in the synthesis and metabolism of various progesterone derivatives.

Antagonistic effect of foreign compounds on microsomal enzymes of the liver of the rat

Abstract The effect of various foreign compounds, given to rats, on the activity of hepatic microsomal enzymes has been studied. Carbon tetrachloride, coumarin, ethionine, and thioacetamide brought about an impairment of phosphatases (substrates: glucose 6-phosphate, inorganic pyrophosphate, inosine diphosphate, guanosine diphosphate, or uridine diphosphate) and slight reduction of drug-metabolizing enzymes (hexobarbital oxidase, butylated hydroxytoluene oxidase, coumarin 3-hydroxylase, nitroanisole demethylase, or codeine demethylase). Hexobarbital, phenobarbital, 3-methylcholanthrene, 4-methylcoumarin, butylated hydroxytoluene (BHT), BHT-alcohol and BHT-alcohol- O -methyl ether had no effect on phosphatases but induced the synthesis of drug-metabolizing enzymes. Antagonism was found between coumarin and 4-methylcoumarin, carbon tetrachloride and 3-methylcholanthrene or phenobarbital in their effect on microsomal enzymes when pairs of these compounds were administered simultaneously, indicating a common site in their action on the endoplasmic reticulum.

Drug Control of Steroid Metabolism by the Hepatic Endoplasmic Reticulum

In this review evidence is provided for the interaction between various drugs and steroid hormones in man and between drugs and progestogens in experimental animals. The mechanism by which these drug interactions occur are of fundamental biochemical and pharmacological interest. The importance of practical clinical considerations of drug-steroid interactions has also been discussed. In particular, considering the present tendency to lower the dose of progestogen and estrogen in most contraceptive preparations, any factor that reduces the bioavailability of the steroid hormones becomes very important. Other drugs and environmental chemicals may interact with these steroids and thereby diminish their efficacy. Clinical studies have reported that the most important interfering drugs are some anticonvulsants and antibiotics, and the antituberculosis compound rifampicin. Anticonvulsants and antituberculotics affect microsomal enzyme induction in the liver or interfere with enzyme systems in the gut wall. The action of antibiotics is connected with the pharmacokinetics of contraceptive steroids by an interaction with their enterohepatic circulation. Some environmental factors such as smoking, alcohol, and other dietary variations, and concurrent hepatic disease may modify the disposition of circulating endogenous steroids and exogenous contraceptive steroids. These effects may alter their response accordingly. In our studies drug treatments of rats reduced serum progesterone level irrespective of whether a potent inducer (phenobarbital, 4-methylcoumarin) or a hepatotoxin (carbon tetrachloride, coumarin, alpha-naphthylisothiocyanate) was administered. These treatments affected hepatic progesterone content. Phenobarbital and carbon tetrachloride reduced serum level, but the hepatic incorporation was enhanced by phenobarbital and reduced by carbon tetrachloride. The opposite actions were selective; phenobarbital raised the oxidative pathway of progesterone metabolism but did not modify the reductive pathway. This drug also enhanced progesterone 16 alpha-, 6 beta-, and 20 alpha-hydroxylase, but did not alter delta 4 - 5 alpha-reductase. In contrast, carbon tetrachloride inhibited hydroxylase and enhanced reductase activities. The effects of these test compounds on progesterone-metabolizing enzymes in isolated microsomes in vitro were similar to the in vivo results. It is concluded that the action of various drugs on serum and liver progesterone levels and metabolism is probably related to changes manifest in the function of the hepatic endoplasmic reticulum.