Enzymatic sulfation of steroids. XI. The extensive purification and some properties of hepatic sulfotransferase III from female rats

1980 ◽  
Vol 58 (8) ◽  
pp. 660-666 ◽  
Author(s):  
Sanford S. Singer ◽  
Lawrence Bruns
Keyword(s):  
Protein Band ◽  
Female Rats ◽  
Male Rats ◽  
Ph Optimum ◽  
Single Protein ◽  
Sequential Mechanism ◽  
Liver Homogenates ◽  
Inhibition Studies ◽  
Estradiol 17Β ◽  
Ordered Sequential Mechanism

Our earlier studies showed that livers from female rats contained three glucocorticoid sulfating enzymes we named sulfotransferases I, II, and III, (STI, STII, and STIII, respectively). In this report STIII from female Charles River CD rats was purified 1010- to 1300-fold compared with liver homogenates. The most highly purified STIII fraction electrophoresed as a single protein band. The molecular weight of STIII was 68 300 ± 4900. Its pH optimum for cortisol sulfation was pH 6.0 ± 0.1. However, it was routinely assayed at pH 6.8 for reasons enumerated in the text. Cortisol sulfation by STIII proceeded by either an ordered sequential mechanism or by an Iso Theorell-Chance mechanism at pH 6.8. The Km's for cortisol and the reaction coenzyme, 3′-phosphoadenosine-5′-phosphosulfate were 6.48 ± 0.78 and 6.78 ± 1.26 μM, respectively. Comparison of the ability of the enzyme to sulfate 40 μM cortisol, estradiol-17β, testosterone, deoxycorticosterone, and dehydroepiandrosterone, showed that the glucocorticoid was sulfated preferentially. Interestingly, its cortisol sulfotransferase activity was inhibited by a number of steroids. p-Hydroxymercuribenzoate inhibition studies indicated the presence of essential sulfhydryl groups in STIII. The enzyme was activated by divalent Ba, Ca, Co, Cr, Mg, Mn, and Ni salts. It was inactivated by Zn2+ and Cd2+ salts. The text compares STIII from female rats with other steroid sulfotransferases including the major glucocorticoid sulfotransferase from male rats.

Enzymatic sulfation of steroids. V. Partial purification and some properties of sulfotransferase III, the major glucocorticoid sulfotransferase of liver cytosols from male rats

1978 ◽  
Vol 56 (11) ◽  
pp. 1028-1035 ◽  
Author(s):  
Sanford S. Singer ◽  
James Gebhart ◽  
Edward Hess
Keyword(s):  
Molecular Weight ◽  
Sulfhydryl Groups ◽  
Male Rats ◽  
Partial Purification ◽  
Ph Optimum ◽  
Fold Enrichment ◽  
Competitive Inhibitors ◽  
Sequential Mechanism ◽  
Inhibition Studies ◽  
Estradiol 17Β

This manuscript describes purification of sulfotransferase III (STIII), the major hepatic glucocorticoid sulfotransferase of male rats, 77.8 ± 16 fold from cytosol. This represents a probable 250–345 fold enrichment, compared with homogenates. Purified STIII has a molecular weight of 61 500 ± 2500 from Sephadex G-100 chromatography. It is markedly activated by 5 mM divalent Ba, Ca, Co, Cr, Mg, Mn, and Ni salts; inhibited strongly by 5 mM divalent Zn and Cd; and unaffected by 8 mM ADP, ATP, and AMP. Comparison of the ability of purified STIII to sulfate equimolar Cortisol, estradiol-17β, testosterone, and dehydroepiandrosterone suggests that the enzyme may sulfate glucocorticoids preferentially. However, its Cortisol sulfotransferase activity is inhibited by a variety of steroids. Of these, dehydroepiandrosterone, dexamethasone, and progesterone were tested extensively. They were found to be competitive inhibitors. STIII has a sharp pH optimum at pH 6.0 ± 0.1. However, it is routinely assayed at pH 6.8, as explained in the text. It exhibits a sequential mechanism and Km values of 6.82 ± 1.2 and 6.28 ± 0.64 μM for Cortisol and 3′-phosphoadenosine-5′-phosphosulfate, respectively. It also possesses essential sulfhydryl groups, as shown by p-hydroxymercuribenzoate inhibition studies.

Gender difference in the Oatp1-mediated tubular reabsorption of estradiol 17β-d-glucuronide in rats

2002 ◽  
Vol 282 (6) ◽  
pp. E1245-E1254 ◽  
Author(s):  
Yasumasa Gotoh ◽  
Yukio Kato ◽  
Bruno Stieger ◽  
Peter J. Meier ◽  
Yuichi Sugiyama
Keyword(s):  
Gender Difference ◽  
Urinary Excretion ◽  
Glomerular Filtration ◽  
Hormonal Regulation ◽  
Organic Anion ◽  
Female Rats ◽  
Male Rats ◽  
Control Male ◽  
Male And Female Rats ◽  
Estradiol 17Β

The gender difference in the urinary excretion of estradiol-17β-glucuronide (E2-17βG) was examined in rats. The urinary clearance of E2-17βG was >250 times lower in male than in female rats. No such major gender difference was observed in its biliary excretion or metabolism in kidney homogenate. Both plasma protein binding and inulin clearance were comparable in male and female rats, suggesting that this gender difference cannot be explained by glomerular filtration. The urinary clearance with respect to the plasma unbound E2-17βG in male rats was <1% of the glomerular filtration rate, indicating its potential reabsorption by the kidney, and this increased to a level comparable with that found in female rats when dibromosulfophthalein was coinfused. A marked increase in E2-17βG urinary excretion was also observed in male rats that had undergone orchidectomy. Testosterone injections given to female rats reduced the urinary excretion to a level comparable with that of control male rats. The concomitant change in the expression of the gene product for organic anion-transporting polypeptide Oatp1, of which E2-17βG is a typical substrate, was found in the kidney membrane fractions after these treatments. These results suggest that urinary E2-17βG excretion is subject to hormonal regulation and that the large gender difference can be explained by regulation in Oatp1-mediated reabsorption.

Purine catabolism in man: inhibition of 5′-phosphomonoesterase activities from placental microsomes

1976 ◽  
Vol 54 (12) ◽  
pp. 1055-1060 ◽  
Author(s):  
Irving H. Fox ◽  
Pamela J. Marchant
Keyword(s):  
Alkaline Phosphatase ◽  
Product Inhibition ◽  
Maximum Activity ◽  
Ph Optimum ◽  
Phosphomonoesterase Activity ◽  
Competitive Inhibitors ◽  
Sequential Mechanism ◽  
Mechanism Of Interaction ◽  
Noncompetitive Inhibitor ◽  
Inhibition Studies

The 5′-phosphomonoesterase activity of 5′-nucleotidase (EC 3.1.3.5) and alkaline phosphatase (EC 3.1.3.1) participates in the catabolism of purine ribonucleotides to uric acid in humans.Initial velocity studies of 5′-nucleotidase suggest a sequential mechanism of interaction between AMP and MgCl2, with a Km of 14 and 3 μM, respectively. With product inhibition studies the apparent Ki's for adenosine, inosine, cytidine, and inorganic phosphate were 0.4, 3.0, 5.0, and 42 mM, respectively. A large number of nucleoside mono-, di-, and tri-phosphate compounds were inhibitors of the enzyme. Allopurinol ribonucleotide, ADP, or ATP were competitive inhibitors when AMP was the substrate, with a Ki slope of 10, 20, or 54 μM, respectively. GTP was a noncompetitive inhibitor, with a Ki slope of 120 μM.The phosphomonoesterase activity of human placental microsomal alkaline phosphatase had a pH optimum of 10.0 and had only 18% of maximum activity at pH 7.4. Substrates and inhibitors included almost any phosphorylated compound. The Km for AMP was 0.4 mM and the apparent Ki for Pi was 0.6 mM. Activity was increased only 19% by 5 mM MgCl2.These observations suggest that 5′-nucleotidase and alkaline phosphatase may be inhibited by ATP and Pi, respectively, under normal intracellular conditions, and that AMP may be preferentially hydrolyzed by 5′-nucleotidase.

Differences in cholesterol oxidation and biosynthesis in liver of male and female rats

1961 ◽  
Vol 200 (3) ◽  
pp. 519-522 ◽  
Author(s):  
David Kritchevsky ◽  
Ezra Staple ◽  
Joseph L. Rabinowitz ◽  
Michael W. Whitehouse
Keyword(s):  
Rat Liver ◽  
Sodium Acetate ◽  
Liver Mitochondria ◽  
Female Rats ◽  
Male Rats ◽  
Cholesterol Oxidation ◽  
Male Rat ◽  
Rat Liver Mitochondria ◽  
Oxidized Cholesterol ◽  
Liver Homogenates

Female rat liver mitochondria oxidized cholesterol-26-C14 and sodium pyruvate-2-C14 to C14O2 to a much greater extent (per mg N) than did male rat liver mitochondria. Mitochondrial preparations from livers of castrated, estrogenized or castrated-estrogenized male rats all oxidized cholesterol-26-C14 to a greater extent than did liver preparations from normal male rats. No differences were observed in the oxidation of sodium octanoate-1-C14. The serum and liver cholesterol levels of the feminized rats were higher than those of the intact males. Biosynthesis of cholesterol from sodium acetate-2-C14 by male rat liver homogenates was significantly lower than biosynthesis by liver homogenates from normal female rats or gonadectomized rats of both sexes. The rate of biosynthesis from mevalonic acid-2-C14 by liver homogenates from castrated male rats was much higher than in homogenates of oophorectomized females or intact males or females. Differences in sex or gonadectomy had no effect on biosynthesis of fatty acids from sodium acetate-2-C14.

Isolation and Characterization of Plasminogen Activator from Pig Leucocytes

1974 ◽  
Vol 31 (01) ◽  
pp. 072-085 ◽  
Author(s):  
M Kopitar ◽  
M Stegnar ◽  
B Accetto ◽  
D Lebez
Keyword(s):  
Molecular Weight ◽  
Plasminogen Activator ◽  
Gel Electrophoresis ◽  
Gel Filtration ◽  
Ph Optimum ◽  
Isolation And Characterization ◽  
Ph Range ◽  
Inhibition Studies ◽  
Final Purification

SummaryPlasminogen activator was isolated from disrupted pig leucocytes by the aid of DEAE chromatography, gel filtration on Sephadex G-100 and final purification on CM cellulose, or by preparative gel electrophoresis.Isolated plasminogen activator corresponds No. 3 band of the starting sample of leucocyte cells (that is composed from 10 gel electrophoretic bands).pH optimum was found to be in pH range 8.0–8.5 and the highest pH stability is between pH range 5.0–8.0.Inhibition studies of isolated plasminogen activator were performed with EACA, AMCHA, PAMBA and Trasylol, using Anson and Astrup method. By Astrup method 100% inhibition was found with EACA and Trasylol and 30% with AMCHA. PAMBA gave 60% inhibition already at concentration 10–3 M/ml. Molecular weight of plasminogen activator was determined by gel filtration on Sephadex G-100. The value obtained from 4 different samples was found to be 28000–30500.

INDUCTION OF GOITRE BY PTU OR KClO4 IN MALE AND FEMALE RATS. EFFECT OF GONADECTOMY

1973 ◽  
Vol 74 (1) ◽  
pp. 88-104 ◽  
Author(s):  
T. Jolín ◽  
M. J. Tarin ◽  
M. D. Garcia
Keyword(s):  
Iodine Content ◽  
High Plasma ◽  
Disc Electrophoresis ◽  
Female Rats ◽  
Male Rats ◽  
Adult Rats ◽  
Male And Female ◽  
Glucose Levels ◽  
Male And Female Rats ◽  
Tsh Levels

ABSTRACT Male and female rats of varying ages were placad on a low iodine diet (LID) plus KClO4 or 6-propyl-2-thiouracil (PTU) or on the same diet supplemented with I (control rats). Goitrogenesis was also induced with LID plus PTU in gonadectomized animals of both sexes. The weight of the control and goitrogen treated animals, and the weight and iodine content of their thyroids were determined, as well as the plasma PBI, TSH, insulin and glucose levels. The pituitary GH-like protein content was assessed by disc electrophoresis on polyacrylamide gels. If goitrogenesis was induced in young rats of both sexes starting with rats of the same age, body weight (B.W.) and pituitary growth hormone (GH) content, it was found that both the males and females developed goitres of the same size. On the contrary, when goitrogenesis was induced in adult animals, it was found that male rats, that had larger B.W. and pituitary GH content than age-paired females, developed larger goitres. However, both male and female rats were in a hypothyroid condition of comparable degree as judged by the thyroidal iodine content and the plasma PBI and TSH levels. When all the data on the PTU or KClO4-treated male and female rats of varying age and B.W. were considered together, it was observed that the weights of the thyroids increased proportionally to B.W. However, a difference in the slope of the regression of the thyroid weight over B.W. was found between male and female rats, due to the fact that adult male rats develop larger goitres than female animals. In addition, in the male rats treated with PTU, gonadectomy decreased the B.W., pituitary content of GH-like protein and, concomitantly, the size of the goitre decreased; an opposite effect was induced by ovariectomy on the female animals. However, when goitrogenesis was induced in weight-paired adult rats of both sexes, the male animals still developed larger goitres than the females. Among all the parameters studied here, the only ones which appeared to bear a consistent relationship with the size of the goitres in rats of different sexes, treated with a given goitrogen, were the rate of body growth and the amount of a pituitary GH-like protein found before the onset of the goitrogen treatment. Moreover, though the pituitary content of the GH-like protein decreased as a consequence of goitrogen treatment, it was still somewhat higher in male that in female animals. The present results suggest that GH may somehow be involved in the mechanism by which male and female rats on goitrogens develop goitres of different sizes, despite equally high plasma TSH levels.

OXYDATION DES ANABOLIKUMS 1-METHYL-ANDROST-1-EN-17β-OL-3-ON MIT WASSERSTOFFTRANSFER AUF ÖSTRON

1971 ◽  
Vol 67 (3) ◽  
pp. 517-530 ◽  
Author(s):  
Martin Wenzel
Keyword(s):  
Rat Liver ◽  
Anabolic Steroid ◽  
Female Rats ◽  
Male Rats ◽  
Female Rat ◽  
Liver Slices ◽  
Androgen Effects ◽  
Biochemical Difference

ABSTRACT With the aid of metenolon-17α-T a tritium-transfer to oestrone in rat liver slices was demonstrated. This tritium-transfer from metenolon17α-T to oestrone yielding tritium-labelled oestradiol had a higher efficiency in male than in female rat liver. Correspondingly in the presence of metenolon the relation of oestrone to oestradiol is changed more in male than in female rat liver. Looking for biochemical differences between the anabolic steroid metenolon and testosterone the oxydation at C17 was measured in different organs of the rat using 17α-T-labelled steroids. The highest oxydation rate was found for both steroids in the liver. In the sexual organs of male rats the oxydation rate of testosterone was 50–10 times higher than that of the anabolic steroid. This difference was less in sexual organs of female rats. This result of a greater biochemical difference between both steroids in males than in females leads to the question, whether the dissociation between the anabolic and the androgen effects is higher in males than in females.

The effect of dexamethasone on mucosal immunity of uterine tissue during pregnancy in rat

2019 ◽  
Vol 20 (1) ◽  
pp. 75-84
Keyword(s):  
Early Pregnancy ◽  
Histopathological Examination ◽  
Early Period ◽  
Interleukin 10 ◽  
Treated Group ◽  
Female Rats ◽  
Male Rats ◽  
Mrna Levels ◽  
Uterine Tissue ◽  
Leukocytic Infiltration

Disturbances in early pregnancy immunity affect embryo development, endometrial receptivity, placental development, fetal growth and lead to subfertility, dexamethasone is a synthetic glucocorticoid used for treatment of various complications. Immune cells and cytokines were examined during the early pregnancy in twenty-four female rats and six male rats for mating. Rats were grouped into two group control and dexamethasone treated by a dose of 50µgm/kgm body weight daily starting from one week before mating and persisted for one week after pregnancy. Blood samples were collected from each rat at 5hrs and at 1,3,7 day of pregnancy. Extracted RNA was subjected to real time PCR to determine mRNA levels for immune related genes interleukin1a(IL1A) and interleukin 10(IL10). Histopathological examination was done to uterus in order to detect leukocyte infiltration in uterine tissue. Results showed that significant increase in white blood cell count mainly eosinophil at 5hrs and lymphocyte at three and seven day of pregnancy of dexamethasone treated group. Moreover, TNF, C-reactive protein and progesterone were increased mainly at seven day of pregnancy of dexamethasone treated group. Similarly, interleukin 1alpha and interleukin 10 significantly increased at 5hrs and one day of pregnancy of dexamethasone treated group. In contrast, serum levels of total antioxidant capacity and estrogen were decreased significantly at 5hrs and seven day in dexamethasone treated group. Histopathological examination of uterus revealed leukocytic infiltration especially neutrophil and few eosinophils at five hours and one day of gestation then eosinophil become absent at 3day and seven day of dexamethasone group. Epithelial height and uterine gland diameter significantly increased at 5hrs, three day and seven days of gestation of dexamethasone treated group. The present investigation demonstrated that using of dexamethasone by dose of 50µgm/kgm during early pregnancy had a conflicting impact on some immune cytokines and parameters and may reflect a harmful response of immune system toward early period of pregnancy

Female rats display higher methamphetamine-primed reinstatement and c-Fos immunoreactivity than male rats

2021 ◽  
pp. 173089
Author(s):  
Steven T. Pittenger ◽  
Shinnyi Chou ◽  
Nathen J. Murawski ◽  
Scott T. Barrett ◽  
Olivia Loh ◽  
...  
Keyword(s):  
Female Rats ◽  
Male Rats

scholarly journals Kidney Injury Caused by Preeclamptic Pregnancy Recovers Postpartum in a Transgenic Rat Model

2021 ◽  
Vol 22 (7) ◽  
pp. 3762
Author(s):  
Sarah M. Kedziora ◽  
Kristin Kräker ◽  
Lajos Markó ◽  
Julia Binder ◽  
Meryam Sugulle ◽  
...  
Keyword(s):  
Rat Model ◽  
Renal Damage ◽  
Kidney Injury ◽  
Tissue Growth ◽  
Female Rats ◽  
Male Rats ◽  
Renal Diseases ◽  
Transgenic Rat ◽  
Increased Risk ◽  
Before And After

Preeclampsia (PE) is characterized by the onset of hypertension (≥140/90 mmHg) and presence of proteinuria (>300 mg/L/24 h urine) or other maternal organ dysfunctions. During human PE, renal injuries have been observed. Some studies suggest that women with PE diagnosis have an increased risk to develop renal diseases later in life. However, in human studies PE as a single cause of this development cannot be investigated. Here, we aimed to investigate the effect of PE on postpartum renal damage in an established transgenic PE rat model. Female rats harboring the human-angiotensinogen gene develop a preeclamptic phenotype after mating with male rats harboring the human-renin gene, but are normotensive before and after pregnancy. During pregnancy PE rats developed mild tubular and glomerular changes assessed by histologic analysis, increased gene expression of renal damage markers such as kidney injury marker 1 and connective-tissue growth factor, and albuminuria compared to female wild-type rats (WT). However, four weeks postpartum, most PE-related renal pathologies were absent, including albuminuria and elevated biomarker expression. Only mild enlargement of the glomerular tuft could be detected. Overall, the glomerular and tubular function were affected during pregnancy in the transgenic PE rat. However, almost all these pathologies observed during PE recovered postpartum.

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