Regular ArticleA Specific Loss of Growth Hormone Abolished Sex-Dependent Expression of Hepatic Cytochrome P450 in Dwarf Rats: Reversal of the Profiles by Growth Hormone-Treatment
Abstract
A spontaneous dwarf rat derived from a colony of Sprague–Dawley (SD) strain has no detectable level of growth hormone (GH) in pituitary, although it contained other hormones like prolactin and ACTH. Hepatic profile of cytochrome P450 (P450) differed clearly between dwarf and normal SD rats. A male-specific form of P450, CYP2C11, was detected in dwarf male rat livers, while the level was one-third of the normal SD livers. This P450 was also detected in dwarf females. Other male-specific CYP3A2 and CYP3A18 were also contained in both sexes of dwarf rats, whereas a female-specific form, CYP2C12, was not detectable in dwarf females. Phenobarbital-inducible CYP2B1 and CYP2B2 were constitutively expressed in dwarf rats, although substantially absent in normal SD rats. To assess the role of GH on hepatic P450 expression, GH was given to dwarf rats for 7 to 9 days. The intermittent injection (mimicking the male secretory pattern) resulted in the elevation of CYP2C11 to a level as observed in normal SD males. Continuous infusion of GH (mimicking the female secretory pattern) evoked CYP2C12 in livers of both sexes of dwarf rats, whereas the treatment decreased levels of CYP3A2, CYP3A18, and CYP2B1. These results clearly demonstrate that specific defect of GH, but not pituitary, cause the clear changes in hepatic P450 forms including sex-specific forms. The present study provides evidence further to strengthen the principal role of GH on the regulation of expression of P450 in rat livers.
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Cytochrome P450 is regulated by noradrenergic and serotonergic systems
2011, Pharmacological ResearchThe aim of the present study was to ascertain whether the noradrenergic or serotonergic systems may affect the expression of liver cytochrome P450 (CYP). Rats were injected intraperitoneally with N-(2-chloroethyl)-N-ethyl-2-bromobenzylamine (DSP-4, a noradrenergic neurotoxin) or p-chloroamphetamine (PCA, a serotonergic neurotoxin) or p-chlorophenylalanine (PCPA, an inhibitor of serotonin synthesis). One week after neurotoxin injection the levels of neurotransmitters (noradrenaline, dopamine, serotonin) and their metabolites were measured in brain structures, and the activity and protein levels of CYP isoforms were measured in the liver. In the brain, DSP-4 or PCA and PCPA selectively decreased noradrenaline or serotonin levels, respectively. In the liver, the applied neurotoxins evoked decrease in the activity of CYP2B, CYP2C11 and CYP3A (DSP-4, PCA, PCPA) and increase in the activity of CYP1A (PCA, PCPA), while the activity of CYP2A, CYP2C6 and CYP2D was not affected by the applied neurotoxins. Since the affected isoforms (CYP1A/2B/2C11/3A) are regulated by endogenous hormones (growth hormone, corticosterone, thyroid hormones), the latter being under control of the central nervous system, it is postulated that the brain noradrenergic and serotonergic systems are involved in the physiological regulation of liver CYP expression.
Enduring effects of severe developmental adversity, including nutritional deprivation, on cortisol metabolism in aging Holocaust survivors
2009, Journal of Psychiatric ResearchIn animal models, early life exposure to major environmental challenges such as malnutrition and stress results in persisting cardiometabolic, neuroendocrine and affective effects. While such effects have been associated with pathogenesis, the widespread occurrence of ‘developmental programming’ suggests it has adaptive function. Glucocorticoids may mediate ‘programming’ and their metabolism is known to be affected by early life events in rodents. To examine these relationships in humans, cortisol metabolism and cardiometabolic disease manifestations were examined in Holocaust survivors in relation to age at exposure and affective dysfunction, notably lifetime posttraumatic stress disorder (PTSD).
Fifty-one Holocaust survivors and 22 controls without Axis I disorder collected 24-h urine samples and were evaluated for psychiatric disorders and cardiometabolic diagnoses. Corticosteroids and their metabolites were assayed by gas chromatography–mass spectroscopy (GC–MS); cortisol was also measured by radioimmunoassay (RIA).
Holocaust survivors showed reduced cortisol by RIA, and decreased levels of 5α-tetrahydrocortisol (5α-THF) and total glucocorticoid production by GC–MS. The latter was associated with lower cortisol metabolism by 5α-reductase and 11β-hydroxysteroid dehydrogenase (11β-HSD) type-2. The greatest decrements were associated with earliest age of Holocaust exposure and less severe PTSD symptomatology. Cardiometabolic manifestations were associated with decreased 11β-HSD-2 activity. In controls, 5α-reductase was positively associated with trauma-related symptoms (i.e., to traumatic exposures unrelated to the Holocaust).
Extreme malnutrition and related stress during development is associated with long-lived alterations in specific pathways of glucocorticoid metabolism. These effects may be adaptive and link with lower risks of cardiometabolic and stress-related disorders in later life.
Simultaneous expression of plural forms of human cytochrome P450 at desired ratios in HepG2 cells: Adenovirus-mediated tool for cytochrome P450 reconstitution
2009, Drug Metabolism and PharmacokineticsHuman hepatocyte culture is widely used to predict human drug metabolism for new drug development. The limited supply and lot-to-lot (donor-to-donor) variations in enzymatic activity, however, hamper its applicability. In the present study, we explore a new cell system with adenovirus-mediated expression of cytochrome P450s (P450s) as an alternative for hepatocytes. In this system, P450 apoprotein levels and catalytic activity increased depending on the amounts of adenoviruses infected for the individual expression of CYP3A4 or CYP2C19 in HepG2 cells. Similar results were observed in the system with co-expression of CYP3A4 and CYP2C19. When HepG2 cells were infected with adednovirus for CYP3A4 and that for CYP2C19 simultaneously at a ratio of 10:1, the ratio of their apoprotein levels was similar to that observed in human hepatocytes and the metabolic profile of diazepam in the system was almost identical to that observed in hepatocyets. These results indicate that this adenovirus-mediated system makes it possible to reproducibly prepare cells expressing multiple P450s at a desired ratio, suggesting a possible use of this system in preclinical metabolic tests for a drug candidate(s), particularly to assess the influence of inter-individual variation in P450 activity.
Influence of recipient gender on intrasplenic fetal liver tissue transplants in rats: Cytochrome P450-mediated monooxygenase functions
2004, ToxicologyRat livers display a sex-specific cytochrome P450 (P450) isoforms expression pattern with consecutive differences in P450-mediated monooxygenase activities, which have been shown to be due to a differential profile of growth hormone (GH) secretion. Parallel to previous investigations on P450 isoforms expression, the aim of the present study was to elucidate the influence of recipient gender on P450-mediated monooxygenase activities in intrasplenic liver tissue transplants in comparison to orthotopic liver. Fetal liver tissue suspensions of mixed gender were transplanted into the spleen of adult male or female syngenic recipients. Four months after grafting transplant-recipients and age-matched controls were treated with β-naphthoflavone (BNF), phenobarbital (PB), dexamethasone (DEX) or the vehicles and sacrificed 24 or 48 h thereafter. P450-dependent monooxygenase activities were assessed by a series of model reactions for different P450 subtypes in liver and spleen 9000g supernatants. In spleens of male and female control rats only very low monooxygenase activities were detectable, whereas with most model reactions distinct activities were observed in transplant-containing organs. Livers and transplant-containing spleens from male rats displayed higher basal ethoxycoumarin O-deethylase and testosterone 2α-, 2β-, 6β-, 14α-, 15α-, 15β-, 16α-, 16β- and 17-hydroxylase activities than those from females. On the other hand, like the respective livers, spleens from female transplant-recipients demonstrated more pronounced p-nitrophenol- and testosterone 6α- and 7α-hydroxylase activities than those from male hosts. With nearly all model reactions gender-specific differences in inducibility by BNF, PB or DEX could be demonstrated in livers as well as in transplant-containing spleens. These results further confirm that the P450 system of intrasplenic liver tissue transplants and the respective orthotopic livers is similarly influenced by recipient gender.
Biochemical background of toxic interaction between tiamulin and monensin
2004, Chemico-Biological InteractionsCitation Excerpt :Several P450 enzymes contribute to N-demethylation of ethylmorphine and aminopyrine to different extent, however, CYP3A1/2 and CYP2C11/12/13 are the main enzymes involved in these N-demethylase activities [31]. CYP3A2 and CYP2C11 are male specific enzymes in rats [32,33]. In untreated adult rats, CYP3A2 and CYP2C11 are constitutively expressed in males, while are undetectable in females.
Tiamulin, a diterpene antibiotic, is used for treatment of pulmonary and gastrointestinal infections in swine and poultry. Combined administration of tiamulin and ionophores (e.g. monensin) to farm animals may lead to intoxication manifested in severe clinical symptoms. Tiamulin metabolite complex with cytochrome P450 has been suggested to be the basis of drug-interactions. However, the formation of metabolic intermediate complex is questionable. The effect of tiamulin-treatment on cytochrome P450 activities was investigated in rats. Ethylmorphine and aminopyrine N-demethylation activities as well as monensin metabolism (O-demethylation) increased in liver microsomes of tiamulin-treated (200 mg/kg) animals. CYP3A1 induction caused by tiamulin was confirmed by the results of Western blot analysis. To test metabolic intermediate complex formation as a result of tiamulin treatment, cytochrome P450 activities were also determined in the presence of potassium ferricyanide. The findings together with those of in vitro complex formation suggested that formation of metabolic intermediate complexes of tiamulin with cytochrome P450 could be excluded. On the other hand, the results of inhibition studies showed significant decrease of ethylmorphine or aminopyrine as well as monensin demethylation in the presence of tiamulin. Our results proved that tiamulin has dual effect on cytochromes P450. It is able to induce and directly inhibit CYP3A enzymes, which are predominantly responsible for monensin O-demethylation. The direct effect of tiamulin as an inhibitor might play a more important role in toxicity than its putative effect as a chemical inducer of CYP3A enzymes.
Influence of recipient gender on cytochrome P450 isoforms expression in intrasplenic fetal liver tissue transplants in rats
2003, ToxicologyRat livers display a sex-specific cytochrome P450 (P450) isoforms expression pattern which is regulated by a differential profile of growth hormone (GH) secretion. The aim of the present study was to elucidate whether liver cell transplants at an ectopic site are also subject to this influence. Fetal liver tissue suspensions of mixed gender were transplanted into the spleen of adult male or female syngenic recipients. Four months after grafting transplant recipients and age-matched controls were treated with β-naphthoflavone (BNF), phenobarbital (PB), dexamethasone (DEX) or the solvents and sacrificed 24 or 48 h thereafter. Livers and intrasplenic transplants were evaluated for the expression of the P450 subtypes 1A1, 2B1, 2E1, 3A2 and 4A1 by means of immunohistochemistry. The livers of both male and female rats displayed nearly no P450 1A1, but a distinct P450 2B1, 2E1, 3A2 and 4A1 expression. Whereas no sex differences were seen in the P450 1A1 expression, the immunostaining for P450 2B1, 3A2 and 4A1 was stronger in males and that for P450 2E1 in females. Similarly, in the intrasplenic liver cell transplants almost no P450 1A1, but a noticeable P450 2B1, 2E1, 3A2 and 4A1 expression was observed. Like in the respective livers, the immunostaining for P450 2B1, 3A2 and 4A1 was stronger in the transplants hosted by male than by female rats, whereas the opposite was the case for the P450 2E1 expression. Both in livers and transplants with some sex-specific differences P450 1A1 and 2E1 expression was induced by BNF, that of P450 2B1 by BNF and PB, and that of P450 3A2 by PB and DEX. These results indicate that the P450 system of ectopically transplanted liver cells is influenced by the gender of the recipient organism like that of the orthotopic livers.
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