Agneta Blanck

Genetic regulation of the cytochrome P-450 system in Drosophila melanogaster. I. Chromosomal determination of some cytochrome P-450-dependent reactions.

The genetic regulation of some cytochrome P-450-dependent enzyme activities has been studied in adult Drosophila. Strains having genetically determined high or low enzyme activities were crossed with a marker strain and the metabolism was analyzed in microsomes from hybrids carrying different combinations of chromosomes from the strain under test. High p-nitroanisole (PNA) N-demethylation, biphenyl 3-hydroxylation and an increased amount of a protein with an apparent mol. wt. of 54 000, after SDS-gel electrophoresis of the microsomes in insecticide-resistant Drosophila strains, are inherited as dominant second chromosome traits. A low capacity for benzo[a]pyrene (BP) hydroxylation and 7-ethoxycoumarin O-deethylation in the Hikone R strain is semidominantly inherited in both cases and determined by gene(s) on the third chromosome. A semidominantly inherited high 4-hydroxylation of biphenyl and a high amount of a protein with an apparent mol. wt. of 56 000 in the Oregon R strain are also localized to the second chromosome. The results indicate that several other cytochrome P-450-dependent activities are not regulated by the genes mentioned above. In conclusion, at least three genes regulating the cytochrome P-450 system in Drosophila have been identified.

Gene Expression Profiling of the Effects of Castration and Estrogen Treatment in the Rat Uterus

Abstract The development and functions of female reproductive tissues are regulated by the actions of two major sex steroid hormones, estrogen and progesterone. To investigate estrogen-dependent gene expression in the rat uterus, we studied the effect of ovariectomy with or without estrogen treatment on the uterine expression of 3000 genes using cDNA microarrays. Many genes were regulated by either treatment, but only few were reciprocally regulated by these contrasting treatments. The present study confirms previous findings and identifies several genes with expressions not previously known to be influenced by estrogen. These genes include follistatin-related protein, Thy-1 glycoprotein, α-fodrin, CD24, immediate early response 5, insulin-like growth factor-binding protein 2, growth response protein CL-6 (INSIG-1), ladinin1, class I major histocompatibility complex heavy chain, lactadherin, ezrin, and Fas-activated serine/threonine kinase. Because of their function as regulators of proliferation and apoptosis, CD24, insulin-like growth factor-binding protein 2, and Fas/Fas ligand were examined further by immunohistochemical expression and tissue localization analysis. Our analysis confirms a contrasting regulation of these gene products by ovariectomy and estrogen treatment. The present study identifies novel mediators of estrogen actions in the uterus and provides genome-wide expression data from which novel hypotheses regarding uterine function can be generated.

Apoptosis, cellular proliferation and expression of p53 in human uterine leiomyomas and myometrium during the menstrual cycle and after menopause

Background. Cell proliferation, apoptosis and expression of p53 proteins were studied in human uterine leiomyomas and myometrium during the menstrual cycle and after menopause.

Expression of Bcl-2, Bcl-x, Mcl-1, Bax and Bak in human uterine leiomyomas and myometrium during the menstrual cycle and after menopause

To investigate the expression of Bcl-2, Bcl-x, Mcl-1, Bax and Bak proteins in human uterine leiomyomas and homologous myometrium during the menstrual cycle and after menopause. The expression of Bcl-2, Bcl-x, Mcl-1, Bax and Bak in leiomyomas (n=24) and myometrial samples (n=22) from women with leiomyomas was measured by immunohistochemistry and Western blot. Measured by immunohistochemistry, a significant difference between leiomyomas and myometrium was observed only for the Bax protein, in tissues obtained from women in the secretory phase of the menstrual cycle. The Bcl-2 staining was more abundant in leiomyomas than in myometrium only in tissues obtained in the proliferative phase of the cycle. Bcl-2 was more abundant in leiomyomas from women of fertile age than in leiomyomas from menopausal women. No significant differences were observed for the Bcl-x or Bak proteins, whereas the Mcl-1 protein was significantly less abundant in secretory phase leiomyomas than in leiomyomas from menopausal women. Western blot analysis based on pools of tissue extracts from the different groups essentially confirmed the data obtained by immunohistochemistry. Bcl-2 family proteins are expressed in leiomyomas and myometrium in different phases related to and influenced by gonadal steroids. These proteins are suggested to interact with each other in the regulation of programmed cell death, apoptosis, but their specific role in growth control of uterine leiomyomas remains to be investigated.

Differences in mRNA expression of endothelin-1, c-fos and c-jun in placentas from normal pregnancies and pregnancies complicated with preeclampsia and/or intrauterine growth retardation

A defect in placental function has been suggested to be associated both with preeclampsia (PE), with or without concomitant intrauterine growth retardation (IUGR), and with IUGR as a single entity. Our aim was to compare the mRNA expression of the growth-related protooncogenes c-fos and c-jun and the vasoconstrictor and growth factor endothelin-1 (ET-1) in placentas from normal pregnancies with those of PE and IUGR. The mRNA expression of c-jun was significantly higher in all groups of complicated pregnancies while ET-1 and c-fos mRNA expression was significantly higher only in the group with IUGR. These results support the concept that an aberrant placental mRNA expression of the ET-1, c-fos and c-jun genes is part of an altered pattern of gene expression in pathological pregnancies.

Placental expression of endothelial constitutive nitric oxide synthase mRNA in pregnancy complicated by preeclampsia

BACKGROUND The role of nitric oxide (NO) in normal pregnancy and pregnancy complicated with preeclampsia (PE) and/or intrauterine growth restriction (IUGR) still remains questionable. The placenta lacks autonomic innervation, therefore blood flow must be regulated by humoral and endothelium derived factors. NO is a potent vasodilator released by endothelial cells. It is synthesized by the catalytic action of the endothelial constitutive nitric oxide synthase (ecNOS). Moreover, the synthesis of NO in normal human placental vasculature has already been established and impairment of the uteroplacental blood flow in pregnancies complicated by PE and/or IUGR has been demonstrated. DESIGN AND METHODS The aim of the present study was to compare the expression of ecNOS mRNA in placenta from women with complicated and normal pregnancies. Placenta was collected from women with PE (n = 13) or small for gestational age (SGA) (n = 8), both PE and SGA (n = 7) and normal pregnancies (n = 41). Total nucleic acids were prepared and a solution hybridization technique was used for mRNA analysis. RESULT The mRNA expression of ecNOS was significantly higher (p<0.05) in all groups of complicated pregnancies compared to normal pregnancies. CONCLUSION Our findings indicate that the increased placental expression of ecNOS mRNA might reflect a compensatory mechanism in the disturbed uterine circulation seen in PE and/or SGA.

Metabolism of Xenobiotics in Hepatocyte Nodules

Hepatocyte nodules that develop during the process of hepatocarcinogenesis in the rat, using a multitude of different experimental protocols, are all resistant to the toxic effects of several xenobiotic compounds (29, 30). This phenotypic property is obvious both irz viw, when nodule-bearing animals arc treated with different toxic substances (32, 89), and itt vitro by treating nodular cells in tissue culture (13, 14, 48). The biological significance of this tolerance to xenobiotic substances is not known in detail, but in the hepatocarcinogenic protocols where mitoinhibitory or other toxic drugs are used in the promotion phase, the resistance certainly seems to be important, permitting the selective proliferation of initiated cells to foci and hepatocyte nodules. Even in the models not using chemical agents as promoters, i.e., the choline-methionine deficient diet model (36, 59, 92) and the orotic acid model (49, 67), resistance to endogenously produced cytotoxic factors like peroxides (64) and free radicals might be important for the development of preneoplastic lesions. The resistance to endogenously produced toxic products might, in these models, be mediated by the same phenotypic changes that are involved in cellular multi-drug resistance. It is generally believed that the development of the “resistance phenotype,” presumably as a result of initiation in the experimental models for chemical hepatocarcinogenesis, is the key to clonal proliferation, manifestation of foci, and their growth into hepatocyte nodules in these models. Therefore, the biochemical investigation of the nature of this resistance seems to be important in the understanding of the mechanisms of chemical hepatocarcino-

Expression of basic fibroblast growth factor (bFGF), FGF receptor 1 and FGF receptor 2 in uterine leiomyomas and myometrium during the menstrual cycle, after menopause and GnRHa treatment

Background. To investigate whether basic fibroblast growth factor (bFGF) is involved in the growth regulation of human uterine leiomyomas the expression of bFGF and its receptors was measured in leiomyomas and myometrium obtained under different endocrine conditions.

Correlation between nortriptyline and debrisoquine hydroxylation in the human liver

The benzylic hydroxylation of nortriptyline (NT) and debrisoquine (D) by isolated human liver microsomes from eight subjects was studied. There was a strong correlation between the 10-hydroxylation of NT and the 4-hydroxylation of D (r = 0.96). The ability to hydroxylate D was also measured in vivo as the ratio between D and 4-OH-D in urine after oral administration of the drug to four subjects. This estimate of hydroxylation capacity agreed with the in vitro measurements. Liver microsomes from a subject defined as a poor in vivo oxidizer of D hydroxylated NT and D unusually slowly. Separation of microsomal proteins by SDS-gel electrophoresis indicated a relative lack of a cytochrome P-450 isozyme with a molecular weight of 54,500 in the liver from the poor oxidizer.

Sex differences and endocrine regulation of morphine oxidation in rat liver

The present study was designed to investigate whether N-demethylation of morphine is regulated via the hypothalamo-pituitary-liver axis in a way similar to that previously described for metabolism of 4-androstene-3,17-dione (androstenedione)