Effect of biliary obstruction and internal biliary drainage on hepatic cytochrome P450 isozymes in rats

AIM： To investigate the total cytochrome P450 （CYP） content, microsomal mixed-function oxidase （MFO） activity, and expression of mRNAs for various CYP isozymes in a simple rat model of reversible obstructive jaundice. METHODS： Obstructive jaundice was created in male rats by causing bile duct obstruction with polyester tape. In another group of rats, bile duct obstruction was followed by internal biliary drainage after releasing the tape. The expression of various CYP isozyme mRNAs was semi-quantitatively assessed by competitive RT- PCR. RESULTS： The total CYP content and microsomal MFO activity showed a significant decrease after biliary obstruction, but returned to respective control levels after biliary drainage. A marked reduction in the expression of CYP1A2, 2B1/2, 2C11, 2E1, 3A1, and 3A2 mRNA was detected during biliary obstruction, while expression increased significantly toward the control level after biliary drainage. Although expression of CYP4A1 mRNA showed no reduction during biliary obstruction, it still increased significantly after biliary drainage. CONCLUSION： These results suggest that not only obstructive jaundice, but also the subsequent internal biliary drainage may affect regulatory medications of the synthesis of individual CYP isozymes differently.     

Effect of ethanol on cytochrome P450 in the rat brain.

After a single dose of ethanol (0.8 ml/kg) administered intraperitoneally, the P450 content of the rat brain increased from 62 +/- 19 to 230 +/- 97 pmol/g (wet weight) of tissue (mean +/- SD, n = 5). Most of this increase could be accounted for by a 10- to 20-fold increase in the olfactory lobes and hypothalamic preoptic area. The P450s were identified by Western blot analysis and by microsequencing of the N-terminal ends after resolution of the proteins on SDS gels. They were identified as P450 2C7, 2C11, 2E1, 4A3, 4A8, and a member of the P450 2D family. In P450 extracted from the brains of control rats, P450 2C and 4A were also detectable but at a much lower concentration. P450 1A1, 2A1, 2B1, or 3A was not detected in the brains of either control or ethanol-treated rats. Oral administration of the same dose of ethanol resulted in a similar increase in the whole brain but smaller effects in the olfactory lobes. This effect of ethanol on the P450 in the brain has implications for the mechanism of toxicity and the development of tolerance to ethanol and for the effects of other drugs and environmental pollutants that act on the central nervous system.     

Effects of portal vein ligation on sex hormone metabolism in male rats: relationship to lowered hepatic cytochrome P450 levels

Hepatic cytochrome P450 levels in male rats fall after portal vein ligation, a procedure that produces total hepatic bypass of portal blood. The present study was undertaken to examine whether changes in sex hormone metabolism could account for these lowered cytochrome P450 levels. Portal vein ligation resulted in testicular atrophy and low serum testosterone concentrations. Serum luteinizing hormone levels were also reduced, suggesting that testicular atrophy was secondary to suppression of the hypothalamic-pituitary-gonadal axis. Serum estrone and estradiol concentrations were significantly increased after portal vein ligation, while the magnitude and delayed onset of increases in urinary total estrogen excretion suggested that this was due largely to increased estrogen production. In male rats, both castration (at 12 wk) and exogenous estrogen administration resulted in changes in hepatic cytochrome P450 levels and ethylmorphine N-demethylase activity that were qualitatively similar to those seen after portal vein ligation. In female and castrated male rats, however, cytochrome P450 was not affected by portal vein ligation. Testosterone supplementation corrected the changes of cytochrome P450 levels in castrated male rats but did not have this effect in portal vein-ligated male rats. It is concluded that changes in sex hormone metabolism do occur after portal vein ligation and may contribute to alterations in cytochrome P450 and drug-metabolizing enzyme activity. Decreased levels of serum testosterone, however, do not alone account for the changes in hepatic drug metabolism in this model, and suppression of a hypothalamic-pituitary factor appears to be important.     

肝细胞色素P450 2E1在实验性肝纤维化组织中的表达

目的 研究肝细胞色素Ｐ４５０ ２Ｅ１在实验性肝纤维化组织中的表达。方法 ８只Ｗｉｓｔａｒ大鼠,建立四氯化碳（ＣＣｌ４）肝纤维化模型,用免疫组织化学方法观察肝纤维化模型肝组织中肝细胞色素Ｐ４５０ ２Ｅ１。结果 正常肝组织内,ＣＹＰ ２Ｅ１表达仅见于中央静脉周围区,主要表达于肝腺泡Ⅲ区,２～３层细胞厚,肝细胞浆和细胞膜可见ＣＹＰ ２Ｅ１表达。在ＣＣｌ４模型肝组织中,ＣＹＰ ２Ｅ１表达的强度增加,分布的     

Expression of cytochrome P450 enzymes in hepatic organoid reconstructed by rat small hepatocytes

BACKGROUND AND AIMS: Small hepatocytes (SH), which are hepatic progenitor cells, were isolated from an adult rat liver. SH in a colony sometimes change their shape from small to large and from flat to rising/piled-up. The morphological changes of SH may be correlated with hepatic maturation. Cytochrome P450s (CYP) are drug-metabolizing enzymes and the expression is one of hepatic differentiated functions. However, it is well known that the re-expression and maintenance of CYP activity are very difficult in cultured hepatocytes. We investigated the expression of CYP and the enzymatic activities in long-term cultured SH. METHODS: SH were isolated from adult rat livers and SH colonies were collected, replated on new dishes, and then cultured. CYP1A1/2, CYP2B1, CYP3A2, CYP4A1, and CYP2E1 were induced by the addition of 3-methylcholanthrene, phenobarbital, pregnenolone-16alpha-carbonitrile, clofibric acid, and ethanol, respectively. Immunocytochemistry, immunoblots, and enzyme activities were examined. RESULTS: SH could differentiate into mature hepatocytes by the addition of Matrigel and re-express constitutive CYPs. The expression of CYP1A1/2, CYP2B1, CYP3A2, and CYP4A1 dose-dependently increased and the amounts gradually increased with time in culture, especially in the cells treated with Matrigel. Activities of CYP1A, CYP2B, CYP3A and CYP2E in SH treated with Matrigel induced by each of the inducers were approximately 120-fold, 2.8-fold, 6.4-fold and 0.8-fold higher than in the control. CONCLUSION: The matured SH could re-express the constitutive CYP and recover inducibility, not only of protein expression but also of enzyme activities.     

Metabolism of melatonin by cytochrome P450s in rat liver mitochondria and microsomes

Abstract: In the present study we provide direct evidence for the involvement of rat microsomal cytochrome P450s in melatonin O‐demethylation and hydroxylation at two different positions: 2 and 6, as well as generation of N¹‐acetyl‐N²‐formyl‐5‐methoxy‐kynuramine (AFMK) and two unknown products. Moreover, we found that mitochondrial cytochrome P450s also converts melatonin into AFMK, N‐acetylserotonin, 2‐hydroxymelatonin, 6‐hydroxymelatonin and the same two unknown products. Eadie–Hofstee plots for 6‐hydroxylation and O‐demethylation reactions were curvilinear for all tested fractions, suggestive of involvement of at least two components, one with a high affinity and low capacity, and another with a low affinity and high capacity. Mitochondrial cytochrome P450s exhibited higher affinity (suggesting lower K_m value) and higher V_max for melatonin 6‐hydroxylation and O‐demethylation for both high‐affinity and low‐affinity components as compared with microsomal enzymes. The intrinsic clearance for melatonin hydroxylation by high‐ and low‐affinity components displayed the highest values in all tested fractions, indicating that both mitochondrial and microsomal cytochrome P450s metabolize melatonin principally by 6‐hydroxylation, with O‐demethylation representing a minor metabolic pathway.     

Measurement and diagnostic use of hepatic cytochrome P450 metabolism of oleic acid in liver disease

Background: Oleic acid is a major systemically circulating fatty acid in humans with atheroprotective and immunomodulatory properties. As of today, the contribution of individual cytochrome P450 (CYP) mono‐oxygenases to the epoxidation of this fatty acid is unknown. Furthermore, the extent of the oleic acid oxidation product cis‐9,10‐epoxyoctadecanoic acid (cis‐EODA) in humans and its plasma levels in patients with impaired liver function are not known. Patients and methods: We studied cis‐EODA in plasma of patients suffering from chronic liver diseases, a condition that often displays impaired liver CYP enzyme activities. Fifteen CYP mono‐oxygenases were investigated in vitro as a potential source of cis‐EODA. Results: Strikingly, plasma levels of cis‐EODA were significantly repressed (P<0.0005) when patients with liver impairment (n=16) were compared with healthy subjects (n=14). Production of cis‐EODA was catalysed by CYP in the following order: 2C8, 2C9, 2C19, 3A4, 1A2 and CYP3A7. Conclusion: cis‐EODA plasma concentrations are decreased in hepatic disease with impaired liver function. Oleic acid is primarily oxidized to oleic acid oxide (cis‐EODA) by CYP2C and CYP3A mono‐oxygenases. The liver is the major organ responsible for the oxidation of oleic acid to cis‐EODA, and thus, cis‐EODA may be a suitable biomarker to assess liver function.     

Effect of amphotericin B on hepatic cytochrome P-450 and glucose-6-phosphatase in the rat.

The effect of amphotericin B on hepatic microsomal cytochrome P-450 (P-450) concentration was measured in vitro, in vivo and ex vivo in the rat. In vitro, both amphotericin B (0-500 micrograms/ml) and its vehicle, sodium deoxycholate (0-410 micrograms/ml), caused similar dose-dependent decreases of P-450 concentrations and glucose-6-phosphatase activity. Intravenous amphotericin B (3 mg/kg) given daily for 3 days decreased antipyrine clearance from control values of 1.24 +/- 0.24 ml/min to 0.67 +/- 0.12 ml/min (p less than 0.001); whereas antipyrine clearance was unchanged by sodium deoxycholate. The P-450 concentration on the third day was reduced from 0.74 +/- 0.14 nmol/mg protein in control rats to 0.33 +/- 0.09 nmol/mg protein in rats treated with amphotericin B (p less than 0.001). Sodium deoxycholate had no effect on P-450 concentration. In contrast, amphotericin B had no effect on either antipyrine clearance or P-450 concentration following enzyme induction by phenobarbital. Amphotericin B had no effect on microsomal glucose-6-phosphatase activity in vivo. Neither amphotericin B nor sodium deoxycholate induced lipid peroxidation, measured as malondialdehyde production. These results show that amphotericin B decreases hepatic cytochrome P-450 content and function in the rat. These effects can not be observed in the enzyme induced state. Amphotericin B has no effect on glucose-6-phosphatase in vivo, the key enzyme of the gluconeogenesis, indicating selective effects on hepatic microsomal function.     

Effect of cyclosporine on liver regeneration after orthotopic reduced-size hepatic transplantation in the rat

These experiments were undertaken to study the effects of cyclosporine A (CsA) on liver regeneration after an isogeneic orthotopic reduced-size hepatic transplantation (RSHT) in rats. Male Wistar rats were treated with or without a daily injection of CsA beginning 24 hr before surgery and were subjected to a 68% partial hepatectomy. A isogeneic orthotopic reduced-size hepatic transplantation was performed in recipient rats pretreated with or without CsA. A daily injection of CsA was continued until the recipient rats were sacrificed. Animals were sacrificed at various time points (12, 24, 36, 48, and 72 hr) postoperatively. The incorporation of bromodeoxyuridine (BrdU) into the DNA of the remnant hepatocytes was evaluated by immunohistochemical staining with a monoclonal antibody against BrdU. CsA (10 mg/kg/day) significantly augmented BrdU incorporation into hepatocytes after hepatectomy. The maximum labeling index (LI) was observed at 24 hr after hepatectomy. In contrast, the maximum LI in the recipient rats not receiving CsA was seen at 36 hr after RSHT, and 10 mg/kg/day of CsA decreased the LI at 36 hr after RSHT. A lower dose of CsA (3 mg/kg/day), however, significantly increased the LI in the recipient rats (P<0.01), and it reached a peak at 24 hr after RSHT when compared to the transplant recipients not receiving CsA. The time course of the increase in the LI in the transplant recipient rats receiving 3 mg/kg/day of CsA was similar to that observed in the rats after hepatectomy. This dosage improved the delay in the reduced-size hepatic transplant LI reaching its peak. These findings suggest that after RSHT the liver graft is more sensitive to both hepatotrophic and hepatotoxic effects of CsA.This work was supported in part by Mika Fund.     

Effect of tamoxifen on hepatic regeneration in male rats

A number of metabolic changes within the liver occur concurrent with hepatic regeneration. These processes suggest that the administration of an antiestrogen might alter the rate of hepatic regeneration. To examine this question, male Wistar rats were treated with tamoxifen (0.1 mg/rat/day or 1.0 mg/rat/day) or vehicle for three days prior to and after partial hepatectomy, and the anatomic and biochemical process of hepatic regeneration was assessed. Tamoxifen administration caused a dose-dependent decrease in the hepatic cytosolic estrogen receptor activity and, conversely, a dose-dependent increase in cytosolic androgen receptor activity. Despite these changes in baseline hepatic sex steroid receptor status, all receptor activities were comparable between the three groups within 24 hr of partial hepatectomy. Moreover, no differences in any of the the parameters assessing hepatic regeneration following partial hepatectomy were evident: liver-body ratio, ornithine decarboxylase activity, and thymidine kinase activity. This lack of effect of tamoxifen treatment on hepatic regeneration suggests either that estrogens do not play a role in the modulation of liver growth after partial hepatectomy or that, once initiated, the regenerative process per sedetermines a series of events that regulate hepatocellular sex hormone receptor status independent of extrahepatic stimuli.This work was supported in part by the Veteran's Administration and grants from NIAAA AA06601, NIAAA AA04425, and NIDDK AM 32556.     

Porcine gonadal and placental isozymes of aromatase cytochrome P450: sub-cellular distribution and support by NADPH-cytochrome P450 reductase

Functional differences exist between the porcine gonadal and placental aromatase cytochrome P450 (P450arom) isozymes that are encoded by separate genes. The present experiments investigated the sub-cellular location of these isozymes, their dependence on the redox partner protein NADPH-cytochrome P450 reductase (P450 reductase) for catalytic activity and the release of steroid intermediates during the aromatization of androgens to estrogen. After differential centrifugation, similar levels of gonadal and placental porcine P450arom were found along with P450 reductase in the microsomal compartment using activity and immunoblot analyses. Activity was stimulated much more by recombinant P450 reductase addition, and higher levels of 19-hydroxy and 19-oxo intermediates accumulated during androstenedione and testosterone metabolism, in cells expressing the gonadal compared to the placental isozyme. No other steroid products were identified by HPLC. Thus, the porcine gonadal P450arom is more sensitive to P450 reductase deprivation than is the placental P450arom, accounting in part for catalytic differences between these isozymes.     

Effect of cigarette smoking on caffeine and trimethadione N-demethylation catalyzed by different cytochrome P450 isozymes in patients with cirrhosis

It has been reported that caffeine (CA) metabolism in humans is catalyzed by cytochrome P450(CYP)1A2. Recently, we reported that human CYP3A4 and 2C, but not 1A2, contribute to trimethadione (TMO) N-demethylation in the human liver. In this study we simultaneously administered two indicators, CA (2 mg/kg) and TMO (4 mg/kg), which have different patterns of metabolism, to patients with cholelithiasis (control) or cirrhosis. The serum levels of CA and TMO, and their individual metabolites, were determined by high-performance liquid chromatography and gas chromatography, respectively. The metabolism of CA and TMO was significantly inhibited in patients with cirrhosis as compared with controls. The ratios of serum paraxanthine (PX)/CA and DMO/TMO at 4 h after administration of the two probe drugs were also significantly decreased in patients with cirrhosis. The PX/CA ratios of the smoking patients with cirrhosis were higher than those of non-smokers. No differences in serum DMO/TMO ratios were observed between the smokers and non-smokers. These results suggest that different estimations of hepatic oxidizing capacity, catalysed mainly by CYP2C and 3A4, are possible in humans.     

Halothane-induced liver injury in guinea-pigs: Importance of cytochrome P450 enzyme activity and hepatic blood flow

The basis for susceptibility to halothane-induced liver necrosis in guinea-pigs was examined. In hepatic microsomes, the following were similar in susceptible and resistant animals: total cytochrome (CYP) P450 (P450), phenobarbital-inducible pathways of mixed function oxidation (androstenedione 6β- and 16β-hydroxylase activities) and the CYP2E1-catalysed pathway of N-nitrosodimethylamine N-demethylase activity. Similarly, immunohistochemical staining of CYP2E1 protein was equivalent in livers from susceptible and resistant guinea-pigs. Prior treatment with the P450-inhibitors, metyrapone and SKF-525A ameliorated halothane-induced liver damage in susceptible animals. Conversely, in resistant guinea-pigs, stimulation of hepatic CYP2E1 activity by treatment with 4-methylpyrazole produced severe hepatotoxicity after re-exposure to halothane. These results confirm the conclusions of others, that P450-mediated metabolism produces halothane-induced liver necrosis in the guinea-pig model but, as in other work, the data fail to explain why no difference in activity of these enzymes could be found between susceptible and resistant guinea-pigs. To establish whether a differential effect on hepatic blood flow between susceptible and resistant guinea-pigs could explain this paradox, studies were performed using a radiolabelled microsphere technique. The effect of halothane on lowering cardiac output was identical in both groups of animals and halothane significantly reduced hepatic arterial but not portal blood flow. The effect on arterial blood flow was more profound in susceptible guinea-pigs (0.67 ± 0.17% of injected microspheres) than in resistant animals (0.99 ± 0.13%; P< 0.005). It is concluded that P450-catalysed metabolism and reduced hepatic blood flow are both necessary to produce halothane-induced liver injury in susceptible guinea-pigs, but it is the effect of halothane on hepatic arterial blood flow that differs between susceptible and resistant animals.     

非酒精性脂肪肝大鼠肝细胞色素P450 1A1基因及表达变化的意义

目的研究非酒精性脂肪肝大鼠肝细胞色素P450 1A1基因和表达变化及其意义.方法雄性Wistar大鼠40只,随机分为两组,正常对照组8只,高脂饮食组2、4、8、12各8只,紫外分光光度计法检测P450 1A1活性(7-乙氧异恶唑0-脱乙基酶,EROD);免疫组织化学和Western blot方法测定肝细胞色素P450 1A1表达变化,逆转录聚合酶链反应测定肝细胞色素P450 1A1 mRNA表达变化.结果脂肪肝2、4、8和12周EROD活性分别为325.07±59.68、345.25±49.28、468.95±55.28和548.68±43.25 nmol·mg-1·min-1,与正常对照组260.42±35.32 nmol·mg-1·min-1比较明显增高(P＜0.01),肝细胞色素P450 1A1基因及蛋白表达随着脂肪肝程度的加重明显增强.结论非酒精性脂肪肝大鼠肝细胞色素P450 1A1基因和表达变化与脂肪肝引起的肝脏损害程度密切相关.     

Effect of amphotericin B on hepatic cytochrome P-450 and glucose-6-phosphatase in the rat

The effect of amphotericin B on hepatic microsomal cytochrome P-450 (P-450) concentration was measured in vitro, in vivo and ex vivo in the rat. In vitro, both amphotericin B (0–500 μg/ml) and its vehicle, sodium deoxycholate (0–410 μg/ml), caused similar dose-dependent decreases of P-450 concentrations and glucose-6-phosphatase activity. Intravenous amphotericin B (3 mg/kg) given daily for 3 days decreased antipyrine clearance from control values of 1.24 ± 0.24 ml/min to 0.67 ± 0.12 ml/min (p < 0.001); whereas antipyrine clearance was unchanged by sodium deoxycholate. The P-450 concentration on the third day was reduced from 0.74 ± 0.14 nmol/mg protein in control rats to 0.33 ± 0.09 nmol/mg protein in rats treated with amphotericin B (p < 0.001). Sodium deoxycholate had no effect on P-450 concentration. In contrast, amphotericin B had no effect on either antipyrine clearance or P-450 concentration following enzyme induction by phenobarbital. Amphotericin B had no effect on microsomal glucose-6-phosphatase activity in vivo. Neither amphotericin B nor sodium deoxycholate induced lipid peroxidation, measured as malondialdehyde production. These results show that amphotericin B decreases hepatic cytochrome P-450 content and function in the rat. These effects can not be observed in the enzyme induced state. Amphotericin B has no effect on glucose-6-phosphatase in vivo, the key enzyme of the gluconeogenesis, indicating selective effects on hepatic microsomal function.     

Ursodeoxycholic acid prevents hepatic cytochrome P450 isozyme reduction in rats with deoxycholic acid-induced liver injury

BACKGROUND/AIMS: Hydrophobic bile acids, such as deoxycholic acid produce cholestatic liver injury. Ursodeoxycholic acid has been shown to be useful in the treatment of cholestatic liver disease. METHODS: In this study, we investigated the effects of deoxycholic acid or ursodeoxycholic acid (1% of diet, for 14 days) and their combination (1% each) on expression of hepatic cytochrome P450 isozymes, their related enzyme activities and mRNA level in rats. RESULTS: Adding 1% deoxycholic acid to chow caused a marked increase in serum total bilirubin (47-fold) and total bile acid (8-fold) concentrations and in alkaline phosphatase (2.5-fold, p<0.01) and alanine aminotransferase activities (23.5-fold, p<0.01). Adding the same dose of ursodeoxycholic acid along with the deoxycholic acid mitigated both the rise in serum total bilirubin and bile acid concentrations and that in alkaline phosphatase and alanine aminotransferase activities, although the use of ursodeoxycholic acid alone did not affect any of the above. Feeding 1% deoxycholic acid caused a decrease (48% of control) in total cytochrome P450 content in hepatic microsomes. Addition of 1% ursodeoxycholic acid along with the 1% deoxycholic acid completely prevented the decrease in total cytochrome P450 content. Feeding ursodeoxycholic acid alone did not affect the total cytochrome P450 content. The expression of cytochrome P450 2B1, 2E1, 3A2, 2C6, 2C11 and 4A1 proteins in hepatic microsomes was decreased by deoxycholic acid (44, 51, 23, 59, 30 and 74% of control, respectively). Likewise, the activities of cytochrome P450 2B1 (pentoxyresorufin O-depentylation), 2E1 (aniline p-hydroxylation) and 3A2 (testosterone 6beta-hydroxylation) isozymes and the 3A2 mRNA levels in liver were decreased by deoxycholic acid. Addition of 1% ursodeoxycholic acid to 1% deoxycholic acid also prevented the decrease in these cytochrome P450 proteins, related enzyme activities and mRNA levels in liver. CONCLUSIONS: These results indicate that, in rats with deoxycholic acid-induced liver injury, ursodeoxycholic acid prevents the decrease in hepatic cytochrome P450 isozymes and suggest that ursodeoxycholic acid is useful for the treatment of liver injury in terms of aiding the normalization of the hepatic drug-metabolizing system.     

Effect of beta-carotene on hepatic cytochrome P-450 in ethanol-fed rats

BACKGROUND: Hepatotoxicity of ethanol is increased by beta-carotene in both rodents and nonhuman primates. Furthermore, in smokers who are also drinkers, beta-carotene increases the incidence of pulmonary cancer. The hepatotoxicity was associated with proliferation of the membranes of the smooth endoplasmic reticulum, suggesting the involvement of cytochromes P-450. Therefore, the aim of the present study was to assess the effect of beta-carotene and ethanol treatment on rodent hepatic cytochromes P-450. METHODS AND RESULTS: Weanling male Sprague-Dawley rats were pair-fed beta-carotene (56.5 mg/l of diet) for 8 weeks, with and without ethanol (Lieber-DeCarli, 1994 liquid diet). As expected, ethanol increased CYP2E1 (measured by Western blots) from 67 +/- 8 to 317 +/- 27 densitometric units (p < 0.001). Furthermore, beta-carotene potentiated the ethanol induction to 442 +/- 38 densitometric units (p < 0.01) with a significant interaction (p = 0.012). The rise was confirmed by a corresponding increase in the hydroxylation of p-nitrophenol, a specific substrate for CYP2E1, and by the inhibition with diethyl dithiocarbamate (50 microM). Beta-carotene alone also significantly induced CYP4A1 protein (328 +/- 49 vs. 158 +/- 17 densitometric units, p < 0.05). The corresponding CYP4A1 mRNA (measured by Northern blots) was also increased (p < 0.05) and there was a significant interaction of the two treatments (p = 0.015). The combination of ethanol and beta-carotene had no significant effect on either total cytochrome P-450 or CYP1A1/2, CYP2B, CYP3A, and CYP4A2/3 contents. CONCLUSIONS: Beta-carotene potentiates the CYP2E1 induction by ethanol in rat liver and also increases CYP4A1, which may, at least in part, explain the associated hepatotoxicity.     

The role of sex steroids in the regulation of insulin sensitivity and serum lipid concentrations during male puberty: a prospective study with a P450-aromatase inhibitor

OBJECTIVE: Our purpose was to study the sex steroid-mediated changes in serum insulin and lipid concentrations in boys during puberty. DESIGN AND METHODS: We treated boys with constitutional delay of puberty either with testosterone plus placebo or with testosterone plus an aromatase inhibitor, letrozole, which inhibits the conversion of androgens to oestrogens. We demonstrated previously that during treatment with testosterone plus letrozole the increase in testosterone concentration was more than 5-fold higher than during treatment with testosterone plus placebo. The concentrations of 17beta-oestradiol, IGF-I and IGF-binding protein-3 increased during testosterone-plus-placebo treatment, but during testosterone-plus-letrozole treatment the concentrations remained unchanged. These divergent changes in the two groups enabled us to study the effects of sex steroids and GH on insulin sensitivity and lipid concentrations. RESULTS: The insulin concentration in the testosterone-plus-placebo-treated group did not change. In contrast, in the testosterone-plus-letrozole-treated group, the concentration decreased during letrozole treatment, indicating improved insulin sensitivity. Changes in insulin and IGF-I concentrations within 12 and 18 months were correlated. In the testosterone-plus-placebo-treated group, the high-density lipoprotein cholesterol concentration did not change but in the testosterone-plus-letrozole-treated group the concentration decreased. The concentrations of low-density lipoprotein cholesterol (LDL-cholesterol) and triglycerides did not change in either of the groups. CONCLUSIONS: The findings indicate that androgens do not directly alter insulin sensitivity in boys during puberty. In contrast, the observations suggest tight regulation of glucose--insulin homeostasis by GH in boys at this stage. Furthermore, our findings indicate that sex steroids do not significantly participate in the regulation of serum concentrations of LDL-cholesterol or triglycerides in boys during early and mid-puberty.