In vitro reprotoxicity of carboxyl-functionalised single- and multi-walled carbon nanotubes on human spermatozoa.

Reproductive toxicity of carboxyl-functionalised carbon nanotubes (CNT-COOH), as the most commonly used form of water-soluble CNTs, is not clearly studied. The aim of this study was to investigate in vitro toxicity of carboxylated single-walled and multi-walled CNTs (SWCNT-COOH and MWCNT-COOH) against human spermatozoa. Sperm cells from healthy donors were incubated with 0.1-100 µg/ml of SWCNT-COOH or MWCNT-COOH at 37°C for up to 5 hr. Viability of sperm cells was assessed using MTT test, and sperm motility was evaluated following World Health Organization guideline. Production of reactive oxygen species (ROS) and nitric oxide (NO) in sperm was also assessed. We showed that both MWCNT-COOH and SWCNT-COOH following incubation in vitro with human spermatozoa did not exert negative effect on viability while motility was significantly (p < .05) dropped in a dose-dependent manner. Moreover, there was no significant effect of the type, dose and exposure time of the CNT-COOH on NO production. Exposure of sperm cells to both examined types of CNTs at concentrations as low as 0.1 µg/ml caused a significant increase in ROS levels. In conclusion, carboxylated forms of CNTs seem to be harmful for human spermatozoa. Further studies, especially using in vivo models, are needed to decide about reprotoxicity of carboxylated forms of CNTs.

Ellagic acid protects against arsenic toxicity in isolated rat mitochondria possibly through the maintaining of complex II.

Chronic arsenic exposure has been linked to many health problems including diabetes and cancer. In the present study, we assessed the protective effect of ellagic acid (EA) against toxicity induced by arsenic in isolated rat liver mitochondria. Reactive oxygen species (ROS) and mitochondrial membrane potential decline were assayed using dichlorofluorescein diacetate and rhodamine 123, respectively, and dehydrogenase activity obtained by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide conversion assay. Arsenic increased ROS levels and mitochondrial dysfunction, which led to a reduction in mitochondrial total dehydrogenase activity. Mitochondria pretreated with EA exposed to arsenic at various concentrations led to a reversal of ROS production and mitochondrial damage. Our results showed that mitochondria were significantly affected when exposed to arsenic, which resulted in excessive ROS production and mitochondrial membrane disruption. Pretreatment with EA, reduced ROS amounts, mitochondrial damage, and restored total dehydrogenase activity specifically associated with mitochondrial complex II. EA protective characteristics may be accomplished particularly throughout the mitochondrial maintenance either directly by its antioxidant property or indirectly through its maintaining of complex II. These findings also suggest a potential role for EA in treating or preventing mitochondria associated disorders.

Effect of beta carotene on lipid peroxidation and antioxidant status following renal ischemia/reperfusion injury in rat.

BACKGROUND: The production of free radicals and reactive oxygen species are important factors contributing to ischemia/reperfusion injury. Thus scavenging of the excess free radicals can be an important therapeutic approach. Beta carotene, a carotenoid pigment has a potent antioxidant property. The present study examined the effect of beta carotene administration on the level of renal content of antioxidants and lipid peroxidation following ischemia/reperfusion injury in the rat kidney. METHODS: Male adult Wistar rats (250-300 g) were exposed to 45 min of renal ischemia followed by 4 h of reperfusion. Beta carotene (10, 30 and 100 mg kg(-1)) or vehicle was administered for 5 days prior to ischemia. Renal content of antioxidants and the level of lipid peroxidation were measured after the reperfusion period. RESULTS: Our results showed that ischemia/reperfusion injury increased lipid peroxidation (p < 0.001) and decreased antioxidant (p < 0.001) in renal tissue. Pre-administration of beta carotene could attenuate these alterations (p < 0.05-p < 0.001), although not at all doses. Since beta carotene administration improved renal lipid peroxidation and antioxidants, it seems that beta carotene protects renal tissue against ischemia/reperfusion-induced oxidative damage.

Protective effect of beta carotene pretreatment on renal ischemia/reperfusion injury in rat.

Renal ischemia/reperfusion injury is a major cause of acute renal failure. The production of free radicals and reactive oxygen species are important factors contributing to ischemia/reperfusion injury. Thus, scavenging of the excess free radicals can be an important therapeutic approach. The present study examined the protective effect of beta carotene against renal ischemia/reperfusion injury in rat. Male adult Wistar rats (250-300 g) were exposed to 45 min of renal ischemia followed by 4 h of reperfusion. Beta carotene (10, 30 and 100 mg kg(-1)) or vehicle was administered for 5 days prior to ischemia. Renal function was assessed by plasma and urinary analysis. Present results showed that ischemia/reperfusion injury increased (p < 0.05-p < 0.001) serum urea and creatinine levels, as well as urinary excretion of protein and calcium and fractional excretion of sodium, while decreased glomerular filtration rate and potassium excretion. However, alterations in these biochemical indices due to ischemia/reperfusion injury were attenuated by beta carotene pretreatment (p < 0.05-p < 0.001), although not by all doses. Since, beta carotene administration improved renal function, it seems that beta carotene protects renal tissue against ischemia/reperfusion-induced oxidative damage.

New thiol and sulfodiimine metalloproteinase inhibitors and their effect on human microvascular endothelial cell growth.

Matrix metalloproteinases (MMPs, matrixins) are a family of homologous zinc endopeptidases that may play a very important role in many physiological and pathological processes, e.g., the initiation of angiogenesis. Two new matrixin inhibitors were synthesized and characterized. A thiol inhibitor MAG-283 had IC(50) values of 480, 3, 280, 14, 1.1, and 2.3 nM against human interstitial collagenase (MMP-1), gelatinase A (MMP-2), stromelysin (MMP-3), matrilysin (MMP-7), neutrophil collagenase (MMP-8), and gelatinase B (MMP-9), respectively. A sulfodiimine inhibitor YLL-224 had IC(50) values of 180, 63, 4500, 210, 5.9, and 44 nM against MMP-1, -2, -3, -7, -8, and -9, respectively. Human skin microvascular endothelial cells were treated with these two compounds in culture. These inhibitors at very low micromolar concentrations suppressed proliferation of the endothelial cells stimulated by acidic fibroblast growth factor and vascular endothelial growth factor. They also partially blocked cell invasion through type IV collagen. These results suggested a correlation between the anti-metalloenzyme activity and the effects of these inhibitors on the growth and invasion of endothelial cells.

Synthesis and tissue distribution of substituted [125I]iodophenylamine derivatives: possible brain imaging agents.

The synthesis and brain uptake in mice of the radioiodinated derivatives of N,N-dimethyl-N'-(iododimethoxyphenyl)-1,3-propanediamine, as well as the N-substituted derivatives of (iodoalkylphenyl)isopropyl, iodoalkylphenylethylamine and 3,4-(methylenedioxy)phenyl-amphetamine (MDA) are described. These compounds contain structural features of both IMP and HIPDM, the cerebral perfusion agents currently in clinical use. The radiolabeled analogs were obtained via the [125I]I exchange method, or by [125I]NaI treatment of the iodo-free precursor in the presence of an oxidant. Following intravenous injection in mice, all compounds showed important radioactivity concentrations in the lungs and kidneys. The N-substituted (iodoalkylphenyl)isopropyl and iodoalkylphenyl-ethylamine derivatives displayed a high initial brain uptake (> 10% IDg-1) followed by a rapid clearance phase, resulting in lower brain-to-blood ratios as those reported for IMP and HIPDM. In contrast, N,N-dimethyl-N'-(iododimethyphenyl)-1,3-propanediamine derivatives featuring the iodo substituent on the benzene carbon adjacent to the methyl amine group and the methoxy substituents on the 2,5- or 2,4-positions, showed low but more persistant brain uptake. Combined with fast blood clearance, this resulted in high brain-to-blood ratios at later time points. Among all compounds tested, the highest brain-to-blood ratio was observed with compound N,N-dimethyl-N'-(6-iodo-3,4-dimethyoxyphenyl)-1,3-propanediamine (27e), reaching a maximum of > 20 at 12 h post-injection.

Importance of oxygen functions in the biological hydroxylation of flavonoids by Absidia blackesleeana.

1. The synthesis and microbiological transformation of 2-phenyl-1-tetralone (compound 3, 1-deoxyisoflavanone), 3-phenyl-1-tetralone (compound 4, 1-deoxyflavanone), 2-phenylchroman (compound 7, 4-deoxyflavanone), 3-phenylchroman (compound 8, 4-deoxyisoflavanone) and 1,2-dihydro-3-phenylnaphthalene (compound 10, 1,4-dideoxy-dehydroflavanone) by Absidia blackesleeana are described. 2. Compounds 3, 4, 7 and 8 were hydroxylated at the 4'-position while compound 10 was not utilized as a substrate. The two phenylchroman analogues 7 and 8 gave approximately the same yield (22% and 26%, respectively) of the 4'-hydroxylation products, while the phenyltetralone analogues 3 and 4 showed significant differences in 4'-hydroxylation (2% and 47%, respectively).

Inhibition of human collagenases by sulfur-based substrate analogs.

A series of sulfhydryl and novel sulfur-based substrate-analog inhibitors has been synthesized and tested against human fibroblast and neutrophil collagenases. Absolute stereospecific synthesis of several sulfhydryl inhibitors establishes that it is the diastereomers with the R-configuration of the P'1 residues, which correspond to the unnatural D-amino acid analogs, that are the most potent inhibitors. The corresponding disulfide, sulfonate, sulfinate, sulfide, sulfoxide and sulfone analogs exhibit widely variable levels of potency, but all less than the sulfhydryl compounds. No correlation between inhibitor potency and any single structural feature of these new compounds is apparent. However, differences in potency can be ascribed to the different affinities of these functional groups for zinc coordination and hydrogen bonding to nearby active site residues.

Synthesis, receptor binding, and tissue distribution of 7 alpha- and 11 beta-substituted (17 alpha,20E)- and (17 alpha,20Z)-21-[125I]iodo-19-norpregna-1,3,5(10),20-tetraene-3,17-diols.

The 11 beta-methoxy, 11 beta-ethoxy, and 7 alpha-methyl derivatives of the isomeric (17 alpha,20E)- and (17 alpha,20Z)-(iodovinyl)estradiols 3 and 6, and their no-carrier-added [125I]iodovinyl analogues, were evaluated for their relative target-tissue retention and binding affinity for the estrogen receptor. The isomeric iodovinyl and [125I]iodovinyl derivatives were prepared via destannylation of the corresponding tributylstannyl precursors in the presence of H2O2 or chloramine-T, with retention of configuration. The 20Z isomers 6 exhibited slightly higher receptor binding affinities than the 20E isomers 3, with all eight isomeric products giving relative binding affinity values in the 20-50 range. The 11 beta- and 7 alpha-substituted (iodovinyl)estradiols gave substantially higher estrogen receptor-mediated uterus uptake as compared to the nonsubstituted parent molecule. Synergism between the effect of 11 beta- or 7 alpha-substituents and the configuration of the iodovinyl group was evident from the in vivo distribution pattern of [125I]-3 and -6. The best uterus uptake was observed, at 2 h postinjection, with the 20E isomer of 11 beta-methoxy derivative 3b. However, at 5 h postinjection the 20Z isomer 6b reached higher uterus concentrations than the 20E isomer 3b, and furthermore, these values are now comparable to those observed with the 20Z isomer of the 11 beta-ethoxy derivative 6c. In the case of the 7 alpha-methyl derivatives the differences in in vivo stability between the 20E and 20Z isomers was less pronounced, whereas the 20Z isomer 6d reached somewhat higher uterus to blood as well as nontarget ratios.

Reaction of thiol nucleophiles with 1,2-epoxy- and 4,5-epoxy-estrene-3-one-17 beta-ols.

Four ring A steroidal epoxyenones as probable intermediate in the formation of catechol estrogens were synthesized. The isomeric 1 alpha,2 alpha-epoxy-17 beta-hydroxyestr-4-en-3-one (9) and 1 beta,2 beta-epoxy-17 beta-hydroxyestr-4-en-3-one (8) were synthesized from 17 beta-hydroxy-5 alpha-estra-3-one. The isomeric 4 alpha,5 alpha-epoxy-17 beta-hydroxyestr-1-en-3-one (11) and 4 beta,5 beta-epoxy-17 beta-hydroxyestr-1-en-3-one (10) were prepared from 19-nortestosterone. The reaction of 9 and 10 with sodium/ethanethiol resulted in the formation of three types of reactions leading to multiple products: 1,4-addition, opening of epoxide, and epoxide opening followed by dehydration. Reaction of 8 with ethanethiol gave only one compound identified as 2-ethanethio-1,4-estradien-17 beta-ol-3-one, while reaction of 9 with ethanethiol gave an unusual product identified as 4-estren-1 alpha,17 beta-diol-3-one. Unlike reaction of ethanethiol with 9 and 10, reaction with N-acetylecysteine or glutathione results in epoxide opening followed by dehydration leading to the formation of estradiol-4-thioethers.

Infarcted heart uptake and biodistribution of radiolabelled anti-myosin monoclonal antibody in rat and dog myocardial infarct models.

A new mouse monoclonal antibody that recognizes alpha- and beta-heavy chains of human atrial and ventricular myosin and beta-heavy chain of human slow skeletal muscle myosin was obtained. The 125I- and 111In-labelled antibody, and its F(ab')2 and Fab fragments localize in isoproterenol induced infarcted rat heart, with the F(ab')2 fragment showing the highest uptake. Comparison with 99Tc-pyrophosphate uptake in infarcted dog heart, induced by selective obstruction of a coronary artery, suggest that the 111In-labelled F(ab')2 localizes specifically in infarcted myocardium only.

Synthesis, receptor binding, and tissue distribution of (17 alpha,20E)- and (17 alpha,20Z)-21-[125I]iodo-19-norpregna-1,3,5(10), 20-tetraene-3,17-diol .

The isomeric (17 alpha,20E)- and (17 alpha,20Z)-(iodovinyl)estradiol derivatives 3 and 6, and their no-carrier-added (nca) [125I]iodovinyl analogues, were tested for their relative target tissue retention and binding affinity for the estrogen receptor. The (iodovinyl)estradiols 3 and 6 were prepared via destannylation of the (17 alpha,20E)- and (17 alpha,20Z)-tributylstannyl precursors 2 and 4 with retention of configuration. Selective formation of the E or Z isomers 2 and 4 during the reaction of 17 alpha-ethynylestradiol 1a with tri-n-butyltin hydride was controlled by the presence or absence of the catalyst, the polarity of the solvent, and the reaction temperature. The nca [125I]iodovinyl analogues [125I]-3a and [125I]-6a were obtained in good radiochemical yield and high purity by treatment of 2a and 4a with [125I]NaI in the presence of H2O2 and chloroamine-T, respectively. Of the two isomeric iodovinyl derivatives 3 and 6, the 20Z isomer 6a exhibited the highest receptor binding affinity and the [125I]-6a gave the highest in vivo receptor-mediated target tissue uptake.

Regioselective A-ring iodination of estradiol diacetates.

Treatment of estrone or estradiol acetate with thallium trifluoroacetate in TFA and subsequent reaction with KI gave the 2-iodoestrogens as the major product. In the case of estradiol diacetate, treatment of the thallation product with [125I]NaI, using the same reaction conditions, gave exclusively the 2-iodo isomer. Thus, regioselectivity combined with rapidity, renders this procedure particularly suitable for A-ring radioiodination of estradiol with short-lived isotopes.