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Biomedical subjects

B Musicki

Publications and source records attributed to B Musicki.

At least 19 recordsLinked to original sources
PubMed2006

Endothelial dysfunction in diabetic erectile dysfunction.

Erectile dysfunction (ED) is highly prevalent in diabetes mellitus. Pathophysiological mechanisms underlying diabetes-associated ED are in large part due to endothelial dysfunction, which functionally refers to the inability of the endothelium to produce vasorelaxing messengers and to maintain vasodilation and vascular homeostasis. The precise mechanisms leading to endothelial dysfunction in the diabetic vasculature, including the penis, are not yet fully understood. Hyperglycemia affects endothelial nitric oxide synthase activity and nitric oxide production/bioavailability, nitric oxide-independent relaxing factors, oxidative stress, production and/or action of hormones, growth factors and/or cytokines, and generation and activity of opposing vasoconstrictors. Considering recent advances in the field of vascular biology and diabetes, the emphasis in this review is placed on the mechanisms of hyperglycemia-induced endothelial dysfunction in the pathophysiology of diabetes-associated ED.

Diabetes Complications↗
PubMed2005

Comparisons of brain, uterus, and liver mRNA expression for cytochrome p450s, DNA methyltransferase-1, and catechol-o-methyltransferase in prepubertal female Sprague-Dawley rats exposed to a mixture of aryl hydrocarbon receptor agonists.

Non-ortho polychlorinated biphenyls (PCBs), polychlorinated dibenzodioxins (PCDDs), and polychlorinated dibenzofurans (PCDFs) are ubiquitous environmental contaminants that exert their toxicity mostly through activation of the aryl-hydrocarbon receptor (AhR), and are referred to as AhR agonists. The objective was to study, by real time reverse-transcriptase-polymerase chain reaction (RT-PCR), the effects of postnatal exposure to a reconstituted mixture of AhR agonists present in breast milk (3 non-ortho PCBs, 6 PCDDs, and 7 PCDFs, referred to here-in-after as AhRM) on mRNA expression of estrogen receptor (ERalpha), enzymes involved with the metabolism of estrogens [catechol-o-methyltransferase (Comt), cytochrome P450 (Cyp)1A1, 1B1 and 2B1], and DNA methyltransferase-1 (Dnmt1), in brain areas, liver and uterus of immature female rats. Neonates were exposed by gavage during postnatal day (PND) 1-20 with dosages equivalent to 1, 10, 100, and 1000 times the estimated average human exposure level, and were sacrificed at PND 21. None of the end points were affected in uterine cross-sections, or in samples of uterine tissue layers collected by laser capture microdissection. At 1000x, the AhRM reduced Dnmt1 mRNA abundance to 28% and 32% of control in the liver and hypothalamus, respectively. In the brain, Cyp1A1 was increased (409%) but ERalpha was reduced (66%). Similarly, mRNA abundance for Comt isoforms was reduced in the liver (45%) and brain areas (55-70%). AhRM at 100x, the lowest effective dose, exerted a 220% increase in brain cortex Comt [membrane bound (Mb)], a 219% increase in hepatic Cyp1B1, and a 63% decrease in hepatic Comt (soluble (S)+Mb). These results support the possibility that early exposure to environmental contaminants could lead to effects mediated by changes in DNA methylation and/or estrogen metabolism and signaling.

Animals↗
PubMed2003

Effects of postnatal exposure to mixtures of non-ortho-PCBs, PCDDs, and PCDFs in prepubertal female rats.

There are concerns that postnatal exposure to organochlorines present in breast milk could lead to adverse health effects. We reconstituted four mixtures of aryl-hydrocarbon receptor (AhR) agonists (3 non-ortho polychlorinated biphenyls [PCBs], 6 polychlorinated dibenzodioxins [PCDDs], 7 polychlorinated dibenzofurans [PCDFs], or all 16 chemicals together [referred to as AhRM]) based on their concentrations in breast milk, and examined their effects following exposure by gavage from day 1 until day 20 of age. Female neonates received dosages of AhRM equivalent to 1, 10, 100, or 1000 times the amount consumed by an infant over the first 24 days of life. Other groups received the PCBs, the PCDDs, or the PCDFs at the 1000x level. All rats were sacrificed at 21 days of age. Changes in ethoxyresorufin-o-deethylase hepatic activity, thymus and body weights, and serum thyroxin were linked to the 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) toxic equivalents (TEQ) of the four mixtures (1000x-AhRM > PCDDs > PCBs > PCDFs). To test for AhRM antiestrogenicity, two additional groups received 1.5 microg/kg of 17alpha-ethynyl estradiol (EE) with or without the 1000x-AhRM. The AhRM had no effect on uterine weight or EE-stimulated uterine growth. The actions of the combined EE and AhRM treatments suggest additive effects in decreasing pentoxyresorufin-o-deethylase activity and spleen weight, but nonadditive/antagonistic effects on adrenal weight and serum thyroxin. In conclusion, (1) 10x-AhRM had no detectable effects, (2) TEQ values relate to observed toxicities, even when testing complex mixtures of AhR agonists, and (3) indications of tissue-specific additive and nonadditive/antagonistic effects, but no synergism, were observed when doses of AhRM were increased, or combined with EE.

Animals↗
PubMed2000

Improved antibacterial activities of coumarin antibiotics bearing 5',5'-dialkylnoviose: biological activity of RU79115.

A new series of coumarin inhibitors of DNA gyrase B bearing a N-propargyloxycarbamate at C-3' of various 5',5'-dialkylnoviose, including RU79115, were synthesised and their antibacterial activities have been delineated. Introduction of dialkyl substituents at 5'5'-position of noviose leads to coumarin analogues with improved in vitro and in vivo antibacterial activity.

Anti-Bacterial Agents↗
PubMed2000

Coumarin inhibitors of gyrase B with N-propargyloxy-carbamate as an effective pyrrole bioisostere.

The synthesis and biological profile in vitro of a series of coumarin inhibitors of gyrase B bearing a N-propargyloxycarbamate at C-3' of noviose is presented. Replacement of the 5-methylpyrrole-2-carboxylate of coumarin drugs with an N-propargyloxycarbamate bioisostere leads to analogues with improved antibacterial activity. Analysis of crystal structures of coumarin antibiotics with the 24 kDa N-terminal domain of the gyrase B protein provides a rational for the excellent inhibitory potency of C-3' N-alkoxycarbamates.

Anti-Bacterial Agents↗
PubMed1999

Synthesis and biological evaluation of coumarincarboxylic acids as inhibitors of gyrase B. L-rhamnose as an effective substitute for L-noviose.

A series of novobiocin-like coumarincarboxylic acids has been prepared bearing the L-rhamnosyl moiety as the sugar portion of the molecule. The similar DNA gyrase inhibitory activity of the novel class of coumarins to that of novobiocin demonstrates that L-rhamnose can effectively replace L-noviose. Introduction of alkyl side-chains at C-5 of coumarin leads to improved in vitro antibacterial properties in the novel series.

Anti-Bacterial Agents↗
PubMed1999

Synthesis and in vitro evaluation of novel highly potent coumarin inhibitors of gyrase B.

The design, synthesis, and in vitro biological activity of a series of novel coumarin inhibitors of gyrase B is presented. Replacement of the 3-acylamino residue (3-NHCOR) of coumarin drugs with reversed isosteres C(=O)R, C(=N-OR)R', COOR, CONHR and CONHOR leads to highly potent analogues which displayed excellent inhibition of the negative supercoiling of the relaxed DNA and antibacterial activity.

Anti-Bacterial Agents↗
PubMed1997

Functional differentiation of placental syncytiotrophoblasts during baboon pregnancy: developmental expression of chorionic somatomammotropin messenger ribonucleic acid and protein levels.

The objective of the present study was to determine whether, in addition to the onset of chorionic somatomammotropin (CS) production previously shown to result from the morphological differentiation of cytotrophoblasts into syncytiotrophoblasts, there is a further developmental increase in the capacity of syncytiotrophoblasts to produce CS with advancing stages of baboon pregnancy. Placentas were obtained from baboons in early (days 48-62), mid (days 97-110), and late (days 161-175) gestation (term = 184 days), and CS messenger ribonucleic acid (mRNA) and protein levels were determined in a syncytiotrophoblast-rich cell fraction isolated by Percoll gradient centrifugation. CS mRNA levels in syncytiotrophoblasts, expressed as a ratio of beta-actin, exhibited a progressive increase from early (0.04 +/- 0.04 relative arbitrary units) to mid (2.37 +/- 0.33; P < 0.001) to late (3.66 +/- 0.39; P < 0.05) gestation. Levels of the 22-kDa CS protein were very low on days 48-55 (0.83 +/- 0.09 arbitrary units), increased 10-fold (P < 0.001) on days 57-60 (8.11 +/- 0.68), and increased (P < 0.001) to a maximum of 14.58 +/- 0.58 near term. CS mRNA levels in whole placental villous tissue increased (P < 0.05) between early (0.89 +/- 0.48) and mid (2.97 +/- 0.47) gestation, then remained constant. CS protein exhibited a similar increase (P < 0.001) in villous tissue between early (2.32 +/- 0.40) and mid (6.07 +/- 0.24) gestation, then remained constant. The increase in mRNA and protein levels of CS in the placenta was accompanied by a progressive (P < 0.001) rise in serum CS. We conclude that in addition to the morphological differentiation of cytotrophoblasts into syncytiotrophoblasts that has been well established to result in the onset of CS biosynthesis, villous syncytiotrophoblasts undergo functional/biochemical differentiation thereafter, manifested as an increase in the capacity for the synthesis of CS.

Animals↗
PubMed1996

Endocrine regulation of ascorbic acid transport and secretion in luteal cells.

Luteal ascorbic acid depletion by LH and prostaglandin (PG) F2 alpha is well known, but how such depletion occurs is not. We therefore investigated the nature and regulation of ascorbic acid uptake and depletion in the rat CL and luteal cells. In vivo studies showed that blockade of steroidogenesis by aminoglutethimide prevented ascorbate depletion by LH, but not PGF2 alpha. Also, the time course for half-maximal depletion of ascorbic acid in vivo in response to PGF2 alpha was extremely rapid (2-3 min) compared to that known for LH (60 min). Thus, ascorbate depletion by LH and PGF2 alpha appears to occur by different mechanisms. In luteal cells, ascorbate uptake was energy-, sodium-, and microfilament-dependent with a Michaelis constant (Km) of 33 microM, similar to that reported for other cells. In contrast to findings for other cells, PGF2 alpha was found to be a potent and rapid inhibitor of ascorbate uptake with a half-maximal inhibition (IC50) of about 5 nM in luteal cells. Ascorbate uptake was unaffected by LH, PGE2, glucose, bromo-cAMP, progesterone, phorbol ester, ionomycin, hydrogen peroxide (H2O2), or aminoglutethimide. Also novel was the finding that luteal cell secretion of ascorbic acid was rapidly and potently stimulated by PGF2 alpha (IC50 about 5 nM), an effect mimicked by LH, H2O2, generators of reactive oxygen, calcium ionophore, and cytochalasin B. Basal release of ascorbic acid was energy-dependent, as secretion was blocked by a mitochondrial uncoupler and lowered temperature. Phorbol ester, bromo-cAMP, progesterone, aminoglutethimide, and ouabain had no effect on ascorbic acid secretion in luteal cells. These findings indicate that the secretion of ascorbic acid induced by PGF2 alpha, and possibly LH, may be mediated by calcium, reactive oxygen, and cytoskeletal changes. The ability of PGF2 alpha to inhibit ascorbate transport and to stimulate secretion implicates these processes as the basis for the rapid depletion of ascorbic acid in the CL. Ascorbate depletion by LH is associated with stimulation of steroidogenesis and an increase in ascorbic acid secretion.

Aminoglutethimide↗
PubMed1994

Inhibition of protein synthesis and hormone-sensitive steroidogenesis in response to hydrogen peroxide in rat luteal cells.

Hydrogen peroxide (H2O2) is generated in the corpus luteum at functional luteal regression and produces rapid antigonadotropic effects in rat luteal cells. However, the mechanism by which peroxide interrupts LH- and cAMP-sensitive progesterone synthesis is unknown. The post-cAMP site of H2O2 action is due to the reduced cholesterol availability in mitochondria, and this process is well known to be dependent on protein synthesis. Therefore, we examined whether H2O2 may interfere with protein and RNA synthesis, and whether such responses may be associated with inhibition of steroidogenesis. Incorporation of radiolabeled amino acids into luteal proteins was inhibited in response to H2O2 in a time- and dose-dependent manner, and these doses are similar to those that inhibit progesterone synthesis, shown earlier in the identical paradigm. The inhibitory effect of H2O2 on amino acid incorporation was not due to increased protein degradation, impaired transport of amino acids, or depletion of cellular ATP levels. H2O2 also inhibited RNA synthesis, increased RNA degradation, and impaired the efficiency of mRNA as a translation template. The time course for the inhibitory effect of H2O2 on protein and RNA synthesis was very rapid and coincident with inhibition of steroidogenesis. Inhibition of protein and RNA synthesis and steroidogenesis were reversed by preincubation of cells with the cell-permeable metal chelator o-phenanthroline, which implicates metal-dependent radical generation as the probable mediator of these actions of H2O2. We conclude that the target of the post-cAMP site of peroxide-induced inhibition of cAMP-dependent steroidogenesis is the inhibition of rapidly inducible proteins that are known to mediate translocation of cholesterol within mitochondria, where it is used as a substrate for pregnenolone synthesis.

Amino Acids↗
PubMed1993

Light-emitting properties of recombinant semi-synthetic aequorins and recombinant fluorescein-conjugated aequorin for measuring cellular calcium.

15 kinds of recombinant semi-synthetic aequorins and a recombinant fluorescein-conjugated aequorin were prepared and their properties in Ca(2+)-triggered luminescence were studied. The semi-synthetic aequorins showed a wide range of Ca(2+)-sensitivity. The luminescence intensity of a high-sensitivity type (hcp-aequorin) was greater than 10(4)-times that of a low-sensitivity type (n-aequorin) at pCa 6.0-6.5. The fluorescein-conjugated aequorin exhibited fluorescence in addition to the Ca(2+)-triggered luminescence, thus it can be used to visualize the diffusion and distribution of aequorin in cells. The data obtained, particularly the Ca(2+)-sensitivity curves, are useful in selecting a suitable semi-synthetic aequorin for an experiment.

Aequorin↗
PubMed1993

Metal chelators reverse the action of hydrogen peroxide in rat luteal cells.

In rat luteal cells hydrogen peroxide (H2O2) interferes with the functional coupling of the luteinizing hormone (LH) receptor and blocks cAMP-dependent progesterone production. To test if this action of H2O2 is dependent on the generation of hydroxyl radicals, the effects of metal chelators and hydroxyl radical scavengers were evaluated. The heavy metal chelator o-phenanthroline prevented H2O2 inhibition of LH-sensitive cAMP and progesterone accumulation and depletion of ATP. Tetrakis(2-pyridylmethyl)ethylenediamine (TPEN) partially reversed inhibition of cAMP accumulation by H2O2 and completely prevented H2O2-induced ATP depletion, but had no effect on H2O2 inhibition of progesterone synthesis. Three other heavy metal chelators, deferoxamine, bathocuproinedisulfonic acid (BA) and penicillamine, as well as hydroxyl radical scavengers ethanol, thiourea and N-(2-mercaptopropionyl)glycine (MPG), had no effect on the luteolytic actions of H2O2. Differential effects of the chelators were probably due to differences in their cell permeability and subcellular compartmentalization. We conclude that metal chelators block the luteolytic actions of H2O2 by a mechanism probably linked to inhibition of hydroxyl radical generation.

Animals↗
PubMed1990

Recombinant aequorin and recombinant semi-synthetic aequorins. Cellular Ca2+ ion indicators.

Properties of a recombinant aequorin were investigated in comparison with those of natural aequorin. In chromatographic behaviour the recombinant aequorin did not match any of ten isoaequorins tested, although it was very similar to aequorin J. Its sensitivity to Ca2+ was found to be higher than that of any isoaequorin except aequorin D. The recombinant aequorin exhibited no toxicity when tested in various kinds of cells, even where samples of natural aequorin had been found to be toxic. Properties of four recombinant semi-synthetic aequorins (fch-, hcp-, e- and n-types), prepared from the recombinant apo-aequorin and synthetic analogues of coelenterazine, were approximately parallel with those of corresponding semi-synthetic aequorins prepared from natural apo-aequorin. Both recombinant e-aequorin and natural e-aequorin J luminesced with high values of the luminescence intensity ratio I400/I465, although the ratios were not pCa-dependent. The recombinant aequorin and recombinant semi-synthetic aequorins are highly suited for monitoring cellular Ca2+.

Aequorin↗
PubMed1990

The antigonadotropic actions of prostaglandin F2 alpha and phorbol ester are mediated by separate processes in rat luteal cells.

Protein kinase-C (PKC) has been suggested as a possible mediator of the antigonadotropic action of prostaglandin F2 alpha (PGF2 alpha) in luteal cells. To examine this possibility, we evaluated the effects of phorbol ester [12-O-tetradecanoylphorbol-13-acetate (TPA)] in relation to those of PGF2 alpha on cAMP accumulation and ATP levels as well as on the subcellular distribution of PKC activity in rat luteal cell cultures. Treatment of luteal cells for 1 h with TPA or PGF2 alpha produced a dose-dependent inhibition of LH-stimulated cAMP accumulation. Maximal inhibition produced by PGF2 alpha was about 35% greater than that produced by TPA. Moreover, PGF2 alpha produced a further inhibition of LH action when the cells were maximally inhibited by TPA. Staurosporine, a PKC inhibitor, reversed inhibition of LH-dependent cAMP accumulation produced by TPA, but had no effect on the response to PGF2 alpha. Furthermore, cells in which PKC was persistently activated by prolonged TPA treatment lost their responsiveness to additional TPA, but continued to show inhibition of cAMP accumulation by PGF2 alpha. TPA also produced a dose-dependent decrease in cell levels of ATP in contrast to PGF2 alpha. Finally, TPA produced a rapid redistribution of PKC activity from the cytosolic to the particulate fraction, whereas PGF2 alpha produced only a slight redistribution. We conclude that the acute antigonadotropic action of PGF2 alpha in rat luteal cells occurs via mechanisms other than phorbol ester-sensitive PKC activation.

Adenosine Triphosphate↗
PubMed1989

Semi-synthetic aequorins with improved sensitivity to Ca2+ ions.

Thirty-seven coelenterazine analogues were synthesized and incorporated into apo-aequorin, yielding 30 semi-synthetic aequorins that have the capacity to emit a significant amount of light in the presence of Ca2+. The properties of resultant photoproteins were investigated. The most prominent feature of those photoproteins was the wide range in their sensitivities to Ca2+ concentration. The relative intensity of Ca2+-triggered luminescence of the photoproteins ranged from 0.01 to 190 when compared with natural aequorin (relative intensity 1.0) at pCa 6 for the cases where the relative intensity is less than 1 and at pCa 7 for the cases where the relative intensity is higher than 1. Eight of the semi-synthetic aequorins belonged to the class of e-aequorin. With two of those photoproteins, the degree of dependence of the luminescence intensity ratio I400/I465 on pCa was greater than that with e-aequorin, suggesting that these two photoproteins are possibly superior to e-aequorin in measuring Ca2+ concentration by the ratio method.

Aequorin↗
PubMed1988

Semi-synthetic aequorin. An improved tool for the measurement of calcium ion concentration.

The photoprotein aequorin isolated from the jellyfish Aequorea emits blue light in the presence of Ca2+ by an intramolecular process that involves chemical transformation of the coelenterazine moiety into coelenteramide and CO2. Because of its high sensitivity to Ca2+, aequorin has widely been used as a Ca2+ indicator in various biological systems. We have replaced the coelenterazine moiety in the protein with several synthetic coelenterazine analogues, providing semi-synthetic Ca2+-sensitive photoproteins. One of the semi-synthetic photoproteins, derived from coelenterazine analogue (II) (with an extra ethano group), showed highly promising properties for the measurement of Ca2+, namely (1) the rise time of luminescence in response to Ca2+ was shortened by approx. 4-fold compared with native aequorin and (2) the luminescence spectrum showed two peaks at 405 nm and 465 nm and the ratio of their peak heights was dependent on Ca2+ concentration in the range of pCa 5-7, thus allowing the determination of [Ca2+] directly from the ratio of two peak intensities. Coelenterazine analogue (I) (with a hydroxy group replaced by an amino group) was also incorporated into apo-aequorin, yielding a Ca2+-sensitive photoprotein, which indicates that an electrostatic interaction between the phenolate group in the coelenterazine moiety and some cationic centre in apo-aequorin is not important in native aequorin, contrary to a previous suggestion.

Aequorin↗
PubMed1988

Selective amplification of luteinizing hormone by adenosine in rat luteal cells.

Adenosine amplification of LH-stimulated cAMP accumulation in rat luteal cells is rapid and dependent on mitochondrial ATP production. The objective of the present studies was to determine if this effect of adenosine is specific for LH and to gain information on the mechanism of the ATP-dependent amplification of LH action in rat luteal cells. Adenosine significantly amplified maximum cAMP accumulation in response to LH, isoproterenol, forskolin, and cholera toxin. However, amplification of this response by adenosine was significantly greater for LH than for the other agonists. The relative order of amplification by adenosine was LH greater than isoproterenol greater than forskolin greater than cholera toxin; the relative magnitudes of amplification by adenosine were 1, 0.6, 0.2, and 0.2, respectively. Neither LH, isoproterenol, forskolin, nor cholera toxin had any effect on cellular levels of ATP, and adenosine produced a similar rate of increase and maximal levels of ATP in the presence of all agonists. Ionomycin, a calcium ionophore, inhibited LH- and cholera toxin-stimulated cAMP accumulation and produced a dose-dependent depletion of ATP. Adenosine reversed the inhibitory effect of ionomycin on LH-stimulated cAMP accumulation and cellular levels of ATP. However, adenosine did not reverse the inhibitory effect of ionomycin on cholera toxin-stimulated cAMP accumulation, although its effects on cellular ATP levels were identical to those on LH. Thus, the selective amplification of LH by adenosine is not merely a substrate effect on adenylate cyclase activity. The nature of adenylate cyclase activation by cholera toxin and forskolin and the weak amplification by adenosine of these agonists compared to that of LH indicate that the site of the ATP-dependent action of adenosine appears to be before or on the G-protein of adenylate cyclase. We suggest that adenosine, by an ATP-dependent process, either increases the availability of functional LH receptors or increases coupling between the LH receptor and adenylate cyclase.

Adenosine↗