Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:3.1.4.1 (phosphodiesterase)
 18,767 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Human Sertoli cell parameters, namely lactate, estradiol-17 beta, and transferrin production, were determined after a 24-hour incubation with either human follicle stimulating hormone (FSH) or dbcAMP in the presence or absence of testosterone plus a phosphodiesterase inhibitor (1-methyl-3-isobutylxanthine; MIX). Testicular tissues were obtained from 10 young patients (mean age, 29 years); using a 3-step enzymatic treatment, Sertoli cell enriched preparations (> 92%) were studied after 4 days as primary cultures. No significant changes in lactate, estradiol-17 beta, and transferrin outputs have been observed according to age in patients ranging in age from 16 years to 47 years. Sertoli cell production of the compounds is controlled by testosterone plus MIX; FSH (or dbcAMP) treatment only slightly improves their synthesis. It is suggested that human Sertoli cell function, as far as the parameters measured in this study are concerned, is likely regulated by cAMP-dependent and independent pathways.
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PMID:Human Sertoli cells in vitro. Lactate, estradiol-17 beta and transferrin production. 142 17

We have used successive density gradient centrifugation with vesicles prepared from a human hepatoma Hep G2 post nuclear supernatant to obtain a highly enriched preparation of early endosomes. A monoclonal antibody (8E4) raised against this early endosome preparation recognizes a single polypeptide highly enriched in light vesicle membranes. The antigen has a molecular weight of 195 kDa by SDS-PAGE in the presence or absence of a reducing agent. Western blot analysis shows that the 8E4 antigen is detectable only in light vesicle membranes and not among heavy membranes, whole cytosol, or nuclear pellet proteins. The 8E4 antigen appears to be an integral membrane protein as it is precipitated by Triton X-114. The distribution of the 8E4 antigen in a Nycodenz density gradient fractionation of light vesicle membranes is identical to the distribution of 125I-ASOR-labeled early endosomes but distinct from the distribution of the plasma membrane enzyme, alkaline phosphodiesterase. In addition, incubation of cells with a horseradish peroxidase-transferrin conjugate followed by 3,3'-diaminobenzidine cytochemistry specifically quenches 8E4 antigen detection by protein dot blot analysis. These data strongly suggest that the 8E4 antigen is an integral membrane protein primarily located in endocytic vesicles.
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PMID:Identification of an endosome-specific antigen. 184 26

Patients with Pelizaeus-Merzbacher disease (PM), hemizygous mice with the jimpy mutation (jp/Y), and hemizygous rats with X-linked myelin deficiency (md/Y) share a profound lack of proteolipid protein (PLP) in their central nervous systems (CNS). The peripheral nervous system is normal. These X-linked disorders are associated with or actually caused by the lack of normal oligodendrocytes. Vibratome sections of brain were incubated with antisera to myelin basic protein (MBP), myelin-associated glycoprotein (MAG), 2':3'-cyclic-nucleotide 3'-phosphodiesterase (CNP) (EC 3.1.4.37), PLP, a synthetic PLP-peptide, glial fibrillary acidic protein (GFAP), and transferrin. Reaction product was developed by sequential incubation with biotinylated second antibodies, the avidin-biotin-peroxidase complex (ABC), and diaminobenzidine (DAB) plus hydrogen peroxide as chromogenic substrates. In PM, jp/Y and md/Y, islands of myelin-like structures were revealed by antisera to MBP, MAG, and CNP. Reaction product after application of anti-PLP was absent. Reaction product after anti-PLP-peptide was restricted to infrequent bizarre cells possibly representing abnormal oligodendroglia. The lack of oligodendrocytes in jp/Y and md/Y could also be confirmed by immunocytochemistry for transferrin.
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PMID:Comparative immunocytochemistry of Pelizaeus-Merzbacher disease, the jimpy mouse, and the myelin-deficient rat. 245 99

The synthesis of carnosine (beta-Ala-His) by astroglia-rich primary cultures was much higher if the cells were cultivated in Ham's nutrient mixture F-12 than if they were grown in Dulbecco's modified Eagle's medium. Carnosine synthesis was not affected by the presence of insulin, transferrin, phorbol myristate acetate, or dexamethasone. However, dibutyryl cyclic AMP and other agents that can, directly or indirectly, activate cyclic AMP-dependent protein kinases strongly lower the rate of carnosine synthesis. The depression of carnosine synthesis was dependent on the concentration of dibutyryl cyclic AMP. The effect was maximal (approximately 80% inhibition) in cultures preincubated with 1 mM dibutyryl cyclic AMP for 4 days. The adenylate cyclase activator forskolin, the phosphodiesterase inhibitor 3-isobutyl-1-methylxanthine, and 8-bromo-cyclic AMP caused the same depression as dibutyryl cyclic AMP, whereas neither butyrate nor dibutyryl cyclic GMP elicited any effect.
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PMID:Regulation by dibutyryl cyclic AMP of carnosine synthesis in astroglia-rich primary cultures kept in serum-free medium. 246 17

We have studied the regulation of inhibin secretion by rat Sertoli cells grown on extracellular matrix-impregnated porous filters in a twin chamber assembly. Previous studies have established that rat Sertoli cells cultured under these conditions reproduce the morphological and functional polarization observed in the Sertoli cell in situ. Sertoli cells isolated from 18- to 22-day-old Wistar rats were cultured for up to 8 days with daily changes of fully defined supplemented Eagle's Minimum Essential Medium (MEM). Rat inhibin was measured by RIA and pituitary cell bioassay, and transferrin by RIA. Inhibin measured by immunoassay or bioassay was always readily detectable in the upper, but not the lower, chamber. Inhibin secretion into the upper chamber exhibited a dose-dependent stimulation of up to 3.7-fold by ovine FSH, with a medium effective dose of 2.2 micrograms/liter and a constant bio- to immunoreactive ratio (3.6 +/- 0.4). Apically directed secretion accounted for over 80% of inhibit output under basal conditions and over 94% with FSH stimulation. Insulin also stimulated upper chamber inhibin secretion at a high dose (5 mg/liter) but not at lower doses or in conjunction with FSH exposure of Sertoli cells. Testosterone augmented FSH-induced stimulation of inhibin secretion, but was ineffective without FSH exposure. In contrast to inhibin secretion, for which FSH is the principal regulator, transferrin secretion by Sertoli cells is more evenly bidirectional (overall mean upper to lower chamber ratio of 1.5) and requires exposure to other stimuli (insulin, retinoic acid, and testosterone) in addition to FSH to achieve maximal secretion. Both submaximal and maximal FSH stimulation of inhibin output were augmented by a phosphodiesterase inhibitor, isobutylmethylxanthine, and these effects were fully reproduced by forskolin, which suggests the involvement of cAMP in the vectorial secretion of inhibin. The marked polarization of Sertoli cell inhibin secretion in vitro could not be explained by restricted transmembrane passage of inhibin. It is, therefore, suggested that the bulk of inhibin secretion by the immature rat Sertoli cell in vivo may be directed primarily into the seminiferous tubular lumen. Thus, in addition to its role in endocrine negative feedback signaling to the pituitary, inhibin may also have important functions in seminiferous tubular function and the support of spermatogenesis.
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PMID:Highly polarized secretion of inhibin by Sertoli cells in vitro. 254 46

Electrotonic coupling between pairs of sympathetic neurons dissociated from superior cervical ganglia of neonatal rats is rare when cells are cultured for 2 weeks in a nutrient medium plus serum and is common when cells are cultured for the same period in serum-free defined medium. This defined medium is the same nutrient medium with five added factors (progesterone, transferrin, putrescine, insulin, and selenium). When added singly to serum-containing medium, insulin and, to a lesser extent, selenium promote the development of electrotonic and dye coupling. The insulin effect is obtained with doses as low as 0.01 microgram/ml and is maximal after exposures from 3 to 5 days. The incidence of electrotonic coupling is also enhanced by exposure of cells to dibutyryl cAMP. This effect is obtained with doses as low as 0.1 mM, is faster (being maximal at approximately equal to 12 hr exposure), and is prolonged in the presence of the phosphodiesterase inhibitor caffeine. Butyrate itself promotes coupling to a small extent, but cAMP involvement is confirmed by similar effects of other membrane permeant analogues. Endogenous levels of cAMP are significantly elevated in cultures grown in the defined medium but not in those in serum-containing medium to which insulin or selenium are added. We conclude that the promotion of coupling by cAMP and by insulin or selenium are independent. The development of coupling in the defined medium thus seems to be a consequence of the addition of promoting substances (insulin, selenium) and the removal of an inhibitory effect of serum on cAMP levels.
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PMID:Determination of synaptic phenotype: insulin and cAMP independently initiate development of electrotonic coupling between cultured sympathetic neurons. 609 Nov 44

Regulation of the synthesis of alpha-fetoprotein (AFP), albumin, and transferrin was studied in temperature-sensitive (ts), simian virus 40 (SV40) tsA mutant-transformed rat fetal liver RLA209-15 cells. The intracellular cyclic AMP (cAMP) levels in RLA209-15 cells grown at the nonpermissive temperature (40 degrees C, the temperature at which the cells exhibit the differentiated phenotype) were higher than the levels in these cells grown at the permissive temperature (33 degrees C, the temperature at which these cells exhibit the transformed phenotype). We have shown previously that differentiation of RLA209-15 cells at 40 degrees C was accompanied by increases in AFP, albumin and transferrin. In the present study, we found that 8 bromo-cAMP (8 BrcAMP), cholera toxin and methylisobutylxanthine (MIX), agents that increase intracellular cAMP levels, greatly stimulated the production of AFP, albumin and transferrin. The phosphodiesterase inhibitor MIX was the most effective inducer of AFP and albumin. The kinetics of AFP induction in the presence of 8BrcAMP, cholera toxin and MIX were similar to the kinetics of albumin induction. The positive correlation of cAMP levels and the differentiated state of these cells indicate that cAMP may be one of the factors that maintains differentiation of fetal liver cells in vitro.
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PMID:Role of cyclic adenosine 3',5'-monophosphate in differentiation of fetal liver cells in vitro. 618 95

Cultured granulosa cells from intact immature rats produced large amounts of progestins in response to phosphodiesterase inhibitor, 3-isobutyl-1-methylxanthine (MIX). MIX alone stimulated up to 9 times higher amounts of 20 alpha-hydroxy-4-pregnen-3-one (20 alpha-OH-P) when compared with FSH (100 ng/ml)-stimulated steroidogenesis. Combined action of MIX and FSH did not yield more activity than MIX action alone. MIX activity was demonstrable exclusively in serum-free culture medium (Dulbecco modified Eagle's medium/F-12) supplemented with insulin, transferrin, hydrocortisone, and fibronectin. Serum severely inhibited MIX induction of 20 alpha-OH-P production. MIX also caused dramatic cell shape changes which occurred within 14 h of exposure to the drug. The cells assumed a spherical shape and remained so as long as MIX was maintained in the culture medium. Upon removal of the drug, the cells respread and acquired the morphology of luteinized cells. In contrast to FSH action, MIX failed to induce the appearance of functional LH receptors. However, similar to FSH effect on immature granulosa cells, MIX successfully induced aromatase enzyme throughout 12 days in culture. MIX alone caused only a minute increase in the accumulation of cAMP. cAMP content in cells induced with FSH (100 ng/ml) was 46 times higher than the cyclic nucleotide levels measured under maximal effective doses of MIX. The discrepancy between the intensive steroidogenic activity of MIX and its inability to substantially raise cAMP content suggests a possible cAMP-independent mechanism for steroid production in the ovarian cells.
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PMID:Cyclic nucleotide phosphodiesterase inhibitor, 3-isobutyl-1-methylxanthine, induces cytodifferentiation of follicular granulosa cells cultured in serum-free medium. 620 21

The response of lamb Sertoli cells to follicle stimulating hormone (FSH) was investigated by measuring transferrin secretion in seminiferous tubule cell cultures throughout the non-pubertal and the prepubertal periods. Cells could be cultured from birth until they attained a testicular weight of 19 g. The characteristics of individual dose-response curves were compared according to the breed, season of birth and testicular weight of the lambs. At the same season of birth and within a given testis weight range, dose-response curves of Romanov and Ile-de-France lambs were similar. Within a given testis weight range, spring-born animals exhibited a higher maximal transferrin secretion than autumn-born lambs, but the ED50 was similar. The main factor of variation of the dose-response curve parameters was the testicular weight of the lambs: the amplitude of FSH response increased 3-fold from a testicular weight of 6 g onwards, i.e. from the appearance of spermatogonia in seminiferous tubules. The ED50 increased 5-fold from 11 g onwards, i.e. from the beginning of the prepubertal period. Thus, Sertoli cells become less sensitive to FSH as spermatogenesis develops in seminiferous tubules. This phenomenon is largely the result of higher phosphodiesterase activity and is greatly reduced by 1-methyl-3-isobutyl-xanthine (MIX).
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PMID:Secretion of transferrin in ovine seminiferous tubule cell cultures in response to FSH: influence of breed, season of birth and age of lambs. 769 24

The effect of hormones on the morphology and cell surface polarity of the epithelial cell line MDCK was examined. When MDCK cells were seeded in high densities in media containing FCS a regular monolayer was formed. However, in serum-free medium supplemented with insulin, transferrin, prostaglandin E1, hydrocortisone and triiodothyronine, the development of a multilayer with intercellular lumina was observed. In hormone-depletion studies we identified PGE1 as the inducer of these multilayers. Since dibutyryl cyclic AMP and the phosphodiesterase inhibitor isobutyl methylxanthine could substitute for PGE1, we conclude that an elevated intracellular cAMP level resulted in formation of the multilayer. Further analysis by electron microscopy and immunocytochemistry revealed a polarized organization of the multilayered cells. Junctional complexes, enclosing microvilli-rich membrane domains, were found at the apices of adjacent cells facing the medium and those surrounding the intercellular lumina. Surprisingly, cells participating in the formation of both the free surface and the surface of the intercellular lumen, exhibited two distinct membranes with microvilli, each separated by junctional complexes. Immunolocalization of membrane marker proteins demonstrated that an apical 114 kDa membrane protein was localized to the free cell surfaces, the same membrane domains where extensive microvilli were also observed. The distribution of a basolateral 58 kDa membrane protein was restricted to sites of cell contact. These results provided evidence that nontransformed epithelial MDCK cells form multilayers in response to elevated cAMP levels; however, they retain the potential of developing cell surface polarity.
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PMID:Elevated cAMP levels induce multilayering of MDCK cells without disrupting cell surface polarity. 839 Oct 11


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