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1.
Identification and partial purification of a unique phenolic steroid sulphotransferase in rat liver cytosol. 总被引:1,自引:0,他引:1
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Y Sugiyama A Stolz M Sugimoto J Kuhlenkamp T Yamada N Kaplowitz 《The Biochemical journal》1984,224(3):947-953
Phenolic steroid sulphotransferase activity for both oestradiol and oestrone was identified in male rat liver cytosol in the 30 000-40 000 Mr fractions on gel filtration when activity was assayed at pH 5.5 (pH optimum 5.5-6.0). Activity for oestradiol but not oestrone was found in the 60 000-70 000-Mr range when assayed at pH 8.0 (pH optimum biphasic, 5.5-6.0 and 7.0-8.0). Km for oestradiol (1.3 microM) was lower than published values for hydroxysteroid sulphotransferases (15-35 microM) and previously reported oestradiol sulphotransferases (71-85 microM). At above 2 microM-oestradiol phenolic sulphotransferase activity exhibited substrate inhibition. The phenolic steroid sulphotransferase activity was found to be distinct in chromatofocusing from organic-anion-binding and bile acid-binding proteins previously identified in this Mr range. Further purification on hydroxyapatite yielded a 44-fold enriched fraction that contained two monomeric bands, Mr 32 500 and 29 500. 相似文献
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Christine E. Conner Rodney W. Kelly Robert Hume 《Histochemistry and cell biology》2001,116(4):313-319
Specialisation of the respiratory portion of human fetal lung commences around 20-24 weeks gestation. In contrast, human fetal lung in vitro has the capacity to self-differentiate from 12 weeks gestation when grown in media devoid of growth factors or hormones, suggesting activation of autocrine or paracrine factors in vitro, or removal of the fetus from in utero inhibitory mechanisms. Prostaglandins play a key role during in vitro human fetal lung development and are synthesised by prostaglandin H synthase-1 (PGHS-1) and inactivated by 15-hydroxyprostaglandin dehydrogenase (PGDH) with formation of inactive 13,14-dihydro-15-keto-prostaglandins. We have used quantitative immunohistochemistry to determine expression and localisation of PGHS-1, PGDH, PGE2 and 13,14-dihydro-15-keto-PGE2 (PGEM) in human fetal lung with in situ hybridisation to localise PGHS-1 and PGDH mRNA. For the catabolic enzyme PGDH, amounts of mRNA, protein and enzyme product PGEM are increased within epithelium of distal as compared to more proximal airways. For PGHS-1, comparable amounts of mRNA, protein and enzyme product PGE2 are found in proximal and distal lung epithelium. Catabolism by PGDH is a sensitive mechanism for regulating bioavailability of prostaglandins and we propose that active catabolism of prostaglandins within human fetal lung epithelium is an inhibitory mechanism retarding epithelial differentiation in utero. 相似文献
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Differential responsiveness of prostatic acid phosphatase and prostate-specific antigen mRNA to androgen in prostate cancer cells 总被引:3,自引:0,他引:3
Androgens regulate the expression of both human prostatic acid phosphatase (PAcP) and prostate-specific antigen (PSA), two major prostate epithelium-specific differentiation antigens. Due to the important role of these two enzymes as prostate epithelium differentiation markers, we investigated their regulation of expression at the mRNA level in LNCaP human prostate carcinoma cells. Interestingly, phenol red, a pH indicator in the culture medium, promoted cell growth. To eliminate this non-specific effect, a phenol red-free, steroid-reduced medium was utilized. When high-density cells were grown in that medium, 5alpha-dihydrotestosterone (DHT) suppressed PAcP but stimulated PSA. However, tumor promoter phorbol ester 12-o-tetradecanoyl phorbol-13-acetate (TPA) functioned as a potent inhibitor of both PAcP and PSA expression. Prolonged treatment with DHT as well as TPA resulted in a similar down-regulation of protein kinase C and cellular PAcP activities. Thus, the levels of PAcP and PSA mRNA are differentially regulated by androgens in LNCaP cells. 相似文献
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Brockhausen I 《Biochemical Society transactions》2003,31(2):318-325
This review summarizes the occurrence, properties and role in mucin O-glycosylation pathways of the various members of glycoprotein sulphotransferase families. Although a number of sulphotransferases have been cloned that act on mucin-type substrates in vitro, it is still difficult to determine exactly which enzymes are responsible for mucin sulphation in vivo. Sulphotransferases play a critical role in determining the chemical, physical and biological properties of mucins. Several of these enzymes have been shown to differ in expression and activity in cancer and inflammation. 相似文献
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Linda W. Gonzales Robert Gonzalez Anne Marie Barrette Ping Wang Leland Dobbs Philip L. Ballard 《The journal of histochemistry and cytochemistry》2015,63(12):908-921
The membrane protein carcinoembryonic antigen cell adhesion molecule (CEACAM6) is expressed in the epithelium of various tissues, participating in innate immune defense, cell proliferation and differentiation, with overexpression in gastrointestinal tract, pancreatic and lung tumors. It is developmentally and hormonally regulated in fetal human lung, with an apparent increased production in preterm infants with respiratory failure. To further examine the expression and cell localization of CEACAM6, we performed immunohistochemical and biochemical studies in lung specimens from infants with and without chronic lung disease. CEACAM6 protein and mRNA were increased ~4-fold in lungs from infants with chronic lung disease as compared with controls. By immunostaining, CEACAM6 expression was markedly increased in the lung parenchyma of infants and children with a variety of chronic lung disorders, localizing to hyperplastic epithelial cells with a ~7-fold elevated proliferative rate by PCNA staining. Some of these cells also co-expressed membrane markers of both type I and type II cells, which is not observed in normal postnatal lung, suggesting they are transitional epithelial cells. We suggest that CEACAM6 is both a marker of lung epithelial progenitor cells and a contributor to the proliferative response after injury due to its anti-apoptotic and cell adhesive properties. 相似文献
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Wan H Luo F Wert SE Zhang L Xu Y Ikegami M Maeda Y Bell SM Whitsett JA 《Development (Cambridge, England)》2008,135(15):2563-2572
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A Radominska K A Comer P Zimniak J Falany M Iscan C N Falany 《The Biochemical journal》1990,272(3):597-604
The sulphation of bile acids is an important pathway for the detoxification and elimination of bile acids during cholestatic liver disease. A dehydroepiandrosterone (DHEA) sulphotransferase has been purified from male and female human liver cytosol using DEAE-Sepharose CL-6B and adenosine 3',5'-diphosphate-agarose affinity chromatography [Falany, Vazquez & Kalb (1989) Biochem. J. 260, 641-646]. Results in the present paper show that the DHEA sulphotransferase, purified to homogeneity, is also reactive towards bile acids, including lithocholic acid and 6-hydroxylated bile acids, as well as 3-hydroxylated short-chain bile acids. The highest activity towards bile acids was observed with lithocholic acid (54.3 +/- 3.6 nmol/min per mg of protein); of the substrates tested, the lowest activity was detected with hyodeoxycholic acid (4.2 +/- 0.01 nmol/min per mg of protein). The apparent Km values for the enzyme are 1.5 +/- 0.31 microM for lithocholic acid and 4.2 +/- 0.73 microM for taurolithocholic acid. Lithocholic acid also competitively inhibits DHEA sulphation by the purified sulphotransferase (Ki 1.4 microM). No evidence was found for the formation of bile acid sulphates by sulphotransferases different from the DHEA sulphotransferase during purification work. The above results suggest that a single steroid sulphotransferase with broad specificity encompassing neutral steroids and bile acids exists in human liver. 相似文献
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Blaisdell CJ Morales MM Andrade AC Bamford P Wasicko M Welling P 《American journal of physiology. Lung cellular and molecular physiology》2004,286(2):L420-L426
Normal lung morphogenesis is dependent on chloride-driven fluid transport. The molecular identity of essential fetal lung chloride channel(s) has not been elucidated. CLC-2 is a chloride channel, which is expressed on the apical surface of the developing respiratory epithelium. CLC-2-like pH-dependent chloride secretion exists in fetal airway cells. We used a 14-day fetal rat lung submersion culture model to examine the role of CLC-2 in lung development. In this model, the excised fetal lung continues to grow, secrete fluid, and become progressively cystic in morphology (26). We inhibited CLC-2 expression in these explants, using antisense oligonucleotides, and found that lung cyst morphology was disrupted. In addition, transepithelial voltage (V(t)) of lung explants transfected with antisense CLC-2 was inhibited with V(t) = -1.5 +/- 0.2 mV (means + SE) compared with -3.7 +/- 0.3 mV (means + SE) for mock-transfected controls and -3.3 +/- 0.3 mV (means + SE) for nonsense oligodeoxynucleotide-transfected controls. This suggests that CLC-2 is important for fetal lung fluid production and that it may play a role in normal lung morphogenesis. 相似文献
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Yu ZY McKay K van Asperen P Zheng M Fleming J Ginn SL Kizana E Latham M Feneley MP Kirkland PD Rowe PB Lumbers ER Alexander IE 《The journal of gene medicine》2007,9(6):429-439
BACKGROUND: Development of effective and durable gene therapy for treatment of the respiratory manifestations of cystic fibrosis remains a formidable challenge. Obstacles include difficulty in achieving efficient gene transfer to mature airway epithelium and the need to stably transduce self-renewing epithelial progenitor cells in order to avoid loss of transgene expression through epithelial turnover. Targeting the developing airway epithelium during fetal life offers the prospect of circumventing these challenges. METHODS: In the current study we investigated vesicular stomatitis virus glycoprotein (VSVg)-pseudotyped HIV-1-derived lentivirus vector-mediated gene transfer to the airway epithelium of mid-gestation fetal lambs, both in vitro and in vivo. In the in vitro studies epithelial sheet explants and lung organ culture were used to examine transduction of the proximal and more distal airway epithelium, respectively. For the in vivo studies, vector was delivered directly into the proximal airway. RESULTS: We found that even during the early pseudoglandular and canalicular phases of lung development, occurring through mid-gestation, the proximal bronchial airway epithelium was relatively mature and highly resistant to lentivirus-mediated transduction. In contrast, the more distal bronchiolar airway epithelium was relatively permissive for transduction although the absolute levels achieved remained low. CONCLUSION: This result is promising as the bronchiolar airway epithelium is a major site of pathology in the cystic fibrosis airway, and much higher levels of transduction are likely to be achieved by developing strategies that increase the amount of vector reaching the more distal airway after intratracheal delivery. 相似文献
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Compartmentalization of vascular endothelial growth factor to the epithelial surface of the human lung 总被引:17,自引:0,他引:17
BACKGROUND: Based on assessment of mRNA expression, the lung is a major site of expression of the vascular endothelial growth factor (VEGF) gene, largely from type II alveolar epithelial cells. With the knowledge that VEGF can function to induce vascular leak, we hypothesized that to protect the lung from pulmonary edema, the VEGF produced in the lung must be compartmentalized from the pulmonary endothelium, and thus must be compartmentalized to the surface of the respiratory epithelium. MATERIAL AND METHODS: To assess this hypothesis, we quantified the levels of VEGF in human respiratory epithelial lining fluid recovered by bronchoalveolar lavage from normal individuals. RESULTS: Strikingly, human respiratory epithelial lining fluid contains 11 +/- 5 ng/mL as quantified by ELISA, a 500-fold greater concentration than plasma (22 +/- 10 pg/mL, p < 0.0005). Western analysis of BAL fluid proteins showed the major VEGF isoform in respiratory epithelial lining fluid is VEGF165. CONCLUSIONS: With the knowledge that proteins of molecular mass like VEGF (34 to 46 kDa) slowly diffuse across the alveolar epithelium, it is likely that this high level "reservoir" of VEGF protein on the respiratory epithelial surface plays a role in normal lung endothelial biology. However, this compartmentalized VEGF reservoir may also be a "Damocles sword" poised to induce lung endothelial permeability in conditions of acute lung injury when the integrity of the alveolar epithelial barrier is breached. 相似文献
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G. S. I. M. Jansen G. F. J. M. Vrensen G. M. J. Van Kempen 《Journal of neurochemistry》1974,23(2):329-335
—The intracellular localization of phenol sulphotransferase in rat brain was studied The distribution pattern found after differential centrifugation closely resembles that of lactate dehydrogenase and does not change during postnatal development. The distribution of the enzyme in discontinuous and continuous sucrose gradients, however, shows a deviation from the lactate dehydrogenase pattern and a shift towards a higher sucrose concentration during development. In the adult the phenol sulphotransferase coincides with monoamine oxidase, succinate dehydrogenase and β-glucuronidase. Disruption experiments, purification of mitochondria and electron microscopy exclude localization of phenol sulphotransferase in mitochondria. These studies support the idea of phenol sulphotransferase as a cytoplasmic enzyme with a preferential binding to or localization in oligodendroglial cells or, more probably, a specific type of synaptosomes. 相似文献
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Properties of estrogen and hydroxysteroid sulphotransferases in human mammary cancer 总被引:2,自引:0,他引:2
Partial purification (approximately x 140-fold) of estrogen sulphotransferase (EC 2.8.2.4) in human mammary estrogen receptor positive cancer tissue was achieved by affinity chromatography on adenosine-3',5'-diphosphate-agarose. It had a Mr of approximately 70,000 by gel filtration and upon electrophoresis on concave gradient polyacrylamide gels, showed a major (Mr 70,000) and a minor (Mr 200,000) peak of activity. Kinetics of this preparation (estradiol-17 beta and estrone as substrates), and also that of hydroxysteroid sulphotransferase (EC 2.8.2.2) contained in the cytosol of human mammary cancer MCF-7 cells (5-androstene-3 beta,17 beta-diol and dehydroepiandrosterone as substrates), were compared. The enzymes showed very similar behaviour, characterized by high affinity for their steroid substrates (low nM range) and co-operativity in their binding. For hydroxysteroid sulphotransferase, the adrenal-derived estrogen 5-androstene-3 beta,17 beta-diol was the preferred substrate compared to dehydro-epiandrosterone in the 0-40 nM concentration range. Such properties of the enzymes might be designed to limit the exposure of nuclear receptor to free ligand. Alternatively, a defined subcellular location would perhaps involve the enzymes in the elimination of estrogen after processing of the ligand-bound receptor. 相似文献
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Tanbir Najrana Anshu Mahadeo Rasha Abu-Eid Elena Kreienberg Victoria Schulte Alper Uzun Christoph Schorl Laura Goldberg Peter Quesenberry Juan Sanchez-Esteban 《Journal of cellular physiology》2020,235(11):8210-8223
The underlying mechanism of normal lung organogenesis is not well understood. An increasing number of studies are demonstrating that extracellular vesicles (EVs) play critical roles in organ development by delivering microRNAs (miRNA) to neighboring and distant cells. miRNAs are important for fetal lung growth; however, the role of miRNA–EVs (miRNAs packaged inside the EVs) during fetal lung development is unexplored. The aim of this study was to examine the expression of miRNA–EVs in MLE-12, a murine lung epithelial cell line subjected to mechanical stretch in vitro with the long-term goal to investigate their potential role in the fetal lung development. Both cyclic and continuous mechanical stretch regulate miRNA differentially in EVs released from MLE-12 and intracellularly, demonstrating that mechanical signals regulate the expression of miRNA–EVs in lung epithelial cells. These results provide a proof-of-concept for the potential role that miRNA–EVs could play in the development of fetal lung. 相似文献