Immunohistochemical localization of serotonin transporter in normal and colchicine treated rat brain

The activity of liver microsomal CYP2E1 is commonly measured as the rate of 5-chloro-2-benzoxazolone (chlorzoxazone) 6-hydroxylation, which requires separation of 6-hydroxychlorzoxazone and chlorzoxazone by high pressure liquid chromatography (HPLC). In the present study, we describe a solvent extraction (non-HPLC) assay for measuring CYP2E1 activity, based on the 6-hydroxylation of [14C]chlorzoxazone. When [14C]chlorzoxazone was incubated with human or rat liver microsomes in the presence of NADPH, the major product formed was 6-[14C]hydroxychlorzoxazone. Unreacted [14C]chlorzoxazone was quantitatively extracted from the incubation mixture with dichloromethane under conditions that resulted in approximately 45% extraction of 6-[14C]hydroxychlorzoxazone. The amount of 6-[14C]hydroxychlorzoxazone remaining in the aqueous incubation mixture ( approximately 55% of the total amount formed) was quantified by liquid scintillation spectrometry. The limit of detection for this assay was 100 pmol of 6-[14C]hydroxychlorzoxazone. The solvent extraction procedure was validated by comparing the rates of formation of 6-[14C]hydroxychlorzoxazone with those determined by HPLC under a variety of experimental conditions. The close correspondence between the two analytical methods suggests that the extraction procedure for measuring 6-[14C]hydroxychlorzoxazone provides a simple, sensitive, and rapid alternative to the HPLC procedure for measuring CYP2E1 activity. In rats, the assay is not specific for CYP2E1 because CYP1A1 also catalyzes the 6-hydroxylation of chlorzoxazone. Recombinant human CYP1A1 also catalyzed the 6-hydroxylation of chlorzoxazone (at (1)/(5) the rate of CYP2E1), although CYP1A1 is not expressed in human liver microsomes. The non-HPLC assay was used to investigate the postulated role of CYP1A2 in the 6-hydroxylation of chlorzoxazone by human liver microsomes. Recombinant CYP1A2 did not catalyze the 6-hydroxylation of chlorzoxazone, and studies with 1-[(3,4-dimethoxyphenyl)methyl]-6,7-dimethoxyisoquinoline, which inhibits CYP1A2 but not CYP2E1, indicated that, in human liver microsomes, the 6-hydroxylation of chlorzoxazone is catalyzed by CYP2E1 with little or no contribution from CYP1A2 enzymes over a wide range of substrate concentrations.     

Doxycycline hyclate treatment of experimental canine ehrlichiosis followed by challenge inoculation with two Ehrlichia canis strains

These is increasing evidence to suggest that central noradrenergic mechanisms may contribute to the central nervous system manifestations of acute liver failure. To further elucidate this possibility, extracellular brain concentrations of the monoamines, noradrenaline (NA), dopamine (DA), and serotonin, were measured by high-performance liquid chromatography with electrochemical detection in microdialysates from the extracellular compartment of frontal cortex in rats with acute (ischemic) liver failure at various times during the progression of encephalopathy and brain edema, as well as in obligate control groups of animals. In addition, binding sites for the noradrenergic receptor subtype ligands, [3H]-prazosin (alpha1 sites), [3H]-RX821002 (alpha2 sites), and [125]I-iodopindolol (beta sites), were assessed using quantitative receptor autoradiography in regions of the brains of rats at coma stage of acute liver failure and of control groups of animals. Coma stages of encephalopathy in acute liver failure were associated with selectively increased noradrenaline concentrations (P < .05) and a concomitant selective loss of alpha1 and beta1 sites in frontal cortex and thalamus. These findings add to a growing body of evidence that central noradrenergic function is modified in acute liver failure and suggest that alpha1/beta1 receptor-mediated noradrenergic mechanisms may play a role in the pathogenesis of brain edema and encephalopathy in this condition.     

Contribution of glycogen and exogenous glucose to glucose metabolism during ischemia in the hypertrophied rat heart

Although hypertrophied hearts have increased rates of glycolysis under aerobic conditions, it is controversial as to whether glucose metabolism during ischemia is altered in the hypertrophied heart. Because endogenous glycogen stores are a key source of glucose during ischemia, we developed a protocol to label the glycogen pool in hearts with either [3H]glucose or [14C]glucose, allowing for direct measurement of both glycogen and exogenous glucose metabolism during ischemia. Cardiac hypertrophy was produced in rats by banding the abdominal aorta for an 8-week period. Isolated hearts from aortic-banded and sham-operated rats were initially perfused under substrate-free conditions to decrease glycogen content to 40% of the initial pool size. Resynthesis and radiolabeling of the glycogen pool with [3H]glucose or [14C]glucose were accomplished in working hearts by perfusion for a 60-minute period with 11 mmol/L [3H]glucose or [14C]glucose, 0.5 mmol/L lactate, 1.2 mmol/L palmitate, and 100 mumol/mL insulin. Although glycolytic rates during the aerobic perfusion were significantly greater in hypertrophied hearts compared with control hearts, glycolytic rates from exogenous glucose were not different during low-flow ischemia. The contribution of glucose from glycogen was also not different in hypertrophied hearts compared with control hearts during ischemia (1314 +/- 665 versus 776 +/- 310 nmol.min-1.g dry wt-1, respectively). Glucose oxidation rates decreased during ischemia but were not different between the two groups. However, in both hypertrophied and control hearts, the ratio of glucose oxidation to glycolysis was greater for glucose originating from glycogen than from exogenous glucose. Our data demonstrate that glycogen is a significant source of glucose during low-flow ischemia, but the data do not differ between hypertrophied and control hearts.     

Estimation of glucose turnover in rats in vivo with tritium labeled glucoses

Starved and starved-refed rats were injected intravenously with labelled glucose (a mixture of [2-3H]-, [3-3H]- and [U-14C]glucose with either [5-3H]- or [6-3H]glucose), and the decay of the specific activity of [14C]glucose followed. Glucose was degraded to obtain the 3H/14C ratios for 3 isotope combinations in the same sample. The apparent rates of replacements, apparent carbon recycling, and the body glucose mass were calculated for the different tracers. The 3H/14C ratio from [2-3H, -U-14C]glucose declined much faster than that of the other tracers. Apparent recycling as calculated in fasted rats was 28% for [2-3H, U-14C]- 18% for [5-3H,-U-14C]- 17% for [3-3H, U-14C]- and 14% for [6-3H,U-14C]glucoses. The values in fed rats showed a similar pattern. We estimate that in fasted rats 85 to 90% of the 3HOH liberated from injected [2-3H]glucose is formed by catabolism in the periphery and the rest by recycling in the liver between glucose and glucose 6-P. Detritiation of other labels by hepatic recycling accounts for a very small fraction of the total 3HOH yield.     

Synthesis of deoxyglucose-1-phosphate, deoxyglucose-1,6-bisphosphate, and other metabolites of 2-deoxy-D-[14C]glucose in rat brain in vivo: influence of time and tissue glucose level

When the kinetics of interconversion of deoxy[14C]glucose ([14C]DG) and [14C]DG-6-phosphate ([14C]DG-6-P) in brain in vivo are estimated by direct chemical measurement of precursor and products in acid extracts of brain, the predicted rate of product formation exceeds the experimentally measured rate. This discrepancy is due, in part, to the fact that acid extraction regenerates [14C]DG from unidentified labeled metabolites in vitro. In the present study, we have attempted to identify the 14C-labeled compounds in ethanol extracts of brains of rats given [14C]DG. Six 14C-labeled metabolites, in addition to [14C]DG-6-P, were detected and separated. The major acid-labile derivatives, DG-1-phosphate (DG-1-P) and DG-1,6-bisphosphate (DG-1,6-P2), comprised approximately 5 and approximately 10-15%, respectively, of the total 14C in the brain 45 min after a pulse or square-wave infusion of [14C]DG, and their levels were influenced by tissue glucose concentration. Both of these acid-labile compounds could be synthesized from DG-6-P by phosphoglucomutase in vitro. DG-6-P, DG-1-P, DG-1,6-P2, and ethanol-insoluble compounds were rapidly labeled after a pulse of [14C]DG, whereas there was a 10-30-min lag before there was significant labeling of minor labeled derivatives. During the time when there was net loss of [14C]DG-6-P from the brain (i.e., between 60 and 180 min after the pulse), there was also further metabolism of [14C]DG-6-P into other ethanol-soluble and ethanol-insoluble 14C-labeled compounds. These results demonstrate that DG is more extensively metabolized in rat brain than commonly recognized and that hydrolysis of [14C]DG-1-P can explain the overestimation of the [14C]DG content and underestimation of the metabolite pools of acid extracts of brain. Further metabolism of DG does not interfere with the autoradiographic DG method.     

The association of non-insulin-dependent diabetes mellitus and cognitive function in an older cohort

This is the first demonstration of the use of accelerator mass spectrometry (AMS) as a tool for the measurement of 3H with attomole (10(-18) mol) sensitivity in a biological study. AMS is an analytical technique for quantifying rare isotopes with high sensitivity and precision and has been most commonly used to measure 14C in both the geosciences and more recently in biomedical research. AMS measurement of serially diluted samples containing a 3H-labeled tracer showed a strong correlation with liquid scintillation counting. The mean coefficient of variation of 3H AMS based upon the analysis of separately prepared aliquots of these samples was 12%. The sensitivity for 3H detection in tissue, protein, and DNA was approximately 2-4 amol/mg of sample. This high sensitivity is comparable to detection limits for 14C-labeled carcinogens using 14C AMS and demonstrates the feasibility of 3H AMS for biomedical studies. One application of this technique is in low-dose, dual-isotope studies in conjunction with 14C AMS. We measured the levels of 3H-labeled 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) and 14C-labeled 2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline (MeIQx) in rat liver tissue and bound to liver DNA and protein 4.5 h following acute administration of individual or coadministered doses in the range of 4-5100 pmol/kg of body weight. Levels of PhIP and MeIQx in whole tissue and bound to liver protein were dose-dependent. MeIQx-protein and -DNA adduct levels were higher than PhIP adduct levels, which is consistent with their respective carcinogenicity in this organ. Coadministration of PhIP and MeIQx did not demonstrate any measurable synergistic effects compared to administration of these compounds individually. These studies demonstrate the application of AMS for the low-level detection of 3H in small biological samples and for its use in conjunction with 14C AMS for dual-labeling studies.     

Expression, purification, and characterization of a recombinant 5-lipoxygenase from potato tuber

The involvement of mdr1a P-glycoprotein (P-gP) on the tissue distribution of the multidrug resistance-reversing agent SDZ PSC 833 was assessed by use of mdr1a (-/-) mice. The mdr1a (-/-) and wild-type mdr1a (+/+) mice received a 4-h constantrate i.v. infusion (2 micrograms/min) of [14C]SDZ PSC 833. Mice were sacrificed at 0, 0.5, 1, 2 and 4 h during infusion and at 0.5, 1, 3, 8 and 24 h after stopping the infusion. Blood and tissues were analyzed on total (14C) and parental SDZ PSC 833 concentrations. Mdr1a (-/-) mice exhibited increased SDZ PSC 833 accumulation in cerebrum, cerebellum and somewhat in testes and small intestine compared with the wild-type mice. The difference between mdr1a (-/-) and (+/+) brain (cerebrum and cerebellum) penetration depended on SDZ PSC 833 blood concentrations, because this cyclosporin analog apparently governs its own brain penetration by inhibiting the P-glycoprotein pump in mdr1a (+/+) mice. Thus the mdr1a (-/-)/(+/+) ratio of brain concentrations tended to decrease and increase at high and low blood concentrations, respectively. These findings clearly demonstrate the interaction of SDZ PSC 833 with the P-glycoprotein present at the blood-brain barrier. The SDZ PSC 833 distribution in other mdr1a P-glycoprotein-expressed tissues, as well as its metabolism and elimination, was not affected by the mdr1a gene disruption. This suggests that factors other than mdr1a P-gP are involved in the disposition of this multidrug resistance-reversing agent.     

Endogenous loss of leucine and methionine in adult male rats

1. The fractional rate of loss of 14C and body-weight was measured in adult male rats after giving 14C-labelled methionine or leucine and maintaining rats for 30 d on a low-protein or a specific methionine+cystine-free diet: carcasses were then analysed for protein and fat 14C radioactivity. 2. The fractional loss of 14CO2 from [14C]methionine or [14C]leucine between day 20 and day 30 was always greater than the fractional loss of body-weight. 3. Carcass protein 14C radioactivity after giving [14C]leucine was higher than after giving [14C]methionine, but fat 14C radioactivity after either 14C-labelled amino acid was only a small proportion of the total body 14C radioactivity. 4. After correction of the fractional loss of 14CO2 for urinary 14C loss, but not body-weight loss, absolute amino acid loss was calculated using published values for methionine and leucine content of rats. 5. The best estimates of endogenous amino acid loss obtained using I-14C-labelled amino acids, expressed as mg/kg body-weight 0.75 per day were leucine 79, methionine 38.     

Effects of in vivo hypoxia on acetylcholine synthesis by rat brain synaptosomes

Synaptosomes from normoxic and hypoxic rats were incubated aerobically in the presence and absence of veratridine. In the absence of veratridine, no significant difference was observed between the two types of preparation regarding either ATP/ADP ratio or 14CO2 or [14C]acetylcholine synthesis from D-[U-14C]glucose. However, in the presence of veratridine, significant reductions in the output of 14CO2 and [14C]acetylcholine by synaptosomes from hypoxic rats were apparent. It was concluded that irreversible metabolic lesions occur at the synapse as a result of hypoxia, which are apparent only when the metabolism of the preparation is accelerated to a level comparable with the maximal rate occurring in vivo. The presence of such lesions is further evidenced by the significant reductions in ATP/ADP ratio, 14CO2 output, and [14C]acetylcholine synthesis that occur in synaptosomes from hypoxic rats made anoxic in vitro and permitted to recover. Such decreases are not seen when synaptosomes from normoxic rats are similarly treated.     

Tissue distribution of macromolecular conjugate, adriamycin linked to oxidized dextran, in rat and mouse bearing tumor cells

Tissue distribution of the radioactivities after intravenous administration of [14C]adriamycin ([14C]ADM) or [14C]ADM linked to oxidized dextran ([14C]ADM-OXD) in mouse bearing Lewis lung carcinoma (LLC) and rat bearing Walker 256 carcinosarcoma was studied. ADM conjugated with OXD increased plasma half-life and gave high area under the plasma concentration-time curve (AUC). The AUC values were 13.0 and 5.8 times higher than those of the [14C]ADM group in mice and rats, respectively. In the tumor tissues, AUC values of the [14C]ADM-OXD group were also respectively 1.6 and 1.9 times higher than those of the [14C]ADM group. However, the AUC values in the heart of the [14C]ADM-OXD group were about half those of [14C]ADM group in both animals. Thus the distribution of ADM was changed by the conjugation with OXD. The excretion profile of ADM was also changed by the conjugation. During 6 h after administration, [14C]ADM-OXD was mainly excreted into rat urine at 45.2% of the original dose, but in the [14C]ADM group recovery in urinary excretion was 4.2%. Using [14C]ADM-OXD and ADM-[14C]OXD, the respective tissue distribution of ADM and OXD portions in the ADM-OXD was studied in rats bearing Walker 256. The radioactivities of both [14C]ADM-OXD and ADM-[14C]OXD groups increased in tumor and liver within 1 h after administration. In the liver, both radioactivities were retained for 24 h, which suggested that ADM and OXD were retained as conjugated form, however, different behavior was observed between the two groups in tumor tissues.(ABSTRACT TRUNCATED AT 250 WORDS)     

Surgical treatment of Ebstein''s anomaly

Up to 88% of a single dose of methyl[14C]methacrylate in rats is expired as 14CO2 in 10 days (65% in 2 h), irrespective of the route of administration and of the specific labelling of the propylene residue of the molecule. The implications of this observation, and of the excretion of small amounts of [14C]methylmalonate, [14C]-succinate and probably of [14C]beta-hydroxyisobutyrate and 2-formylpropionate, and of the formation of [14C] normal, physiological metabolites that may be accounted for by anabolism both from 14CO2 and from [14C]acetate emergent from the citric acid cycle, are that the metabolic pathway concerned involves intermediary metabolism and relates to mitochondrial function. Present findings are discussed in relation to the imputations of a report of carcinogenic risk.     

P53 genes and malignant hematologic diseases

The primary objective of the present study was to compare the rates of plasma clearance and hepatic utilization of stearic (18:0), myristic (14:0) and linoleic (18:2) acids, as introduced via chylomicrons. Lymph chylomicrons were specifically labeled in vivo with [14C]stearic and (SA), [14C]myristic acid (MA), or [14C]linoleic acid (LA) by infusing donor rats intraduodenally with the labeled fatty acids in a lipid emulsion. Following intravenous injection of recipient rats with the labeled chylomicrons, the rates of plasma clearance and incorporation of the label in triglycerides (TG), phospholipids (PL) and other lipids in the liver were compared at 5, 15 and 30 min. [14C]SA was cleared at a slightly faster rate (t1/2 = 7.0 min) than [14C]MA (t1/2 = 8.1 min) and [14C]LA (t1/2 = 8.0 min) (P < 0.05). [14C]SA was accumulated in the liver at a significantly faster rate than [14C]MA and [14C]LA. At the peak (15 min) of hepatic uptake, 30.3% of [14C]SA, 26.2% of [14C]LA and 21.9% of [14C]MA were recovered in the liver. At 30 min, 33.5% of [14C]SA was taken up by the liver, whereas 27.8% of [14]LA and only 15.2% of [14C]MA were removed. In the liver, the percentage of [14C]SA incorporated into PL steadily increased with time, whereas the percent-age incorporated into TG decreased. [14C]SA was preferentially incorporated into PL at all time intervals, as compared with [14C]MA and [14C]LA. At 30 min, 38.6% of [14C]SA was found in PL, and only 5.2% of [14C]MA and 12.0% of [14C]LA were present in PL.(ABSTRACT TRUNCATED AT 250 WORDS)     

Hospital administration--the new order; the medical administrator''s point of view

Urine from rats and sheep given single doses of [14C]propachlor contained 14C metabolites in which the chlorine of propachlor was replaced by a methylsulfonyl group. Methylsulfonyl-containing metabolites were also isolated from the urine of rats given an intraperitoneal dose of the cysteine conjugate of [14C]propachlor; this indicated that the methylsulfonyl-containing metabolites resulted from metabolic reactions subsequent to the mercapturic acid pathway.     

Rates of excretion of cotinine, nicotine glucuronide, and 3-hydroxycotinine glucuronide in rat bile

A new HPLC assay was adapted for radiometric detection of nicotine metabolites in rat bile. Two glucuronides were identified as the principal biliary metabolites of nicotine. In addition to nicotine glucuronide and 3-hydroxycotinine glucuronide, cotinine was also detected in bile after administration to rats of a single subcutaneous dose of (-)-S-nicotine (0.2 or 1.0 mg/kg) that contained a tracer dose of rac-[pyrrolidine-2'-14C]nicotine (20 microCi). Biliary metabolites accounted for only 3% of the [14C]nicotine dose, but phenobarbital pretreatment (100 mg/kg ip for 3 days) increased the amount of [14C]nicotine-derived radioactivity recovered in bile to 8% and also accelerated rates of biliary excretion of all three nicotine metabolites. Dose-dependency of nicotine metabolism occurred: less nicotine glucuronide was excreted at the low dose than at the high dose.     

Utilization of 1-14C- glucose, 1-14C-acetate and 1-14C-palmitate during lipid synthesis in vitro in the liver and skeletal muscles of pigs at different ontogenic stages

The specific radioactivity of lipids synthesized in the liver and quadriceps muscle of 110-day embryos, and 1-, 30-, 60-, 90-day and 12-month pigs in vitro from [1-14C]acetate, [1-14C] glucose and [1-14C] palmitate was investigated. The lipid synthesis from most of the above compounds in the fetal liver developed at a greater rate and in the fetal skeletal muscles at a smaller rate than in livers and skeletal muscles of newborn piglets. During the first months after birth the lipid synthesis from [1-14C] acetate, [1-14C] glucose and [1-14C] palmitate in the liver increased gradually. At the same time the lipid synthesis in skeletal muscles from [1-14C]glucose and [1-14C]palmitate decreased and from [1-14C] acetate increased. The utilization of the above compounds in the lipid synthesis of the liver and quadriceps muscle of pigs follows the decreasing order: [1-14C] palmitate greater than [1-14C]acetate greater than [1-14C]glucose.     

Synthesis and excretion profile of 1,4-[14C]phenylenebis(methylene)selenocyanate in the rat

1,4-Phenylenebis(methylene)selenocyanate (p-XSC) inhibits chemically induced tumors in several laboratory animal models. To understand its mode of action, we synthesized p-[14C]XSC, examined its excretion pattern in female CD rats and also the nature of its metabolites. p-[14C]XSC was synthesized from alpha,alpha-dibromo-p-[ring-14C]xylene in 80% yield. The excretion profile of p-[14C]XSC (15.8 mg/kg body wt, 200 microCi/rat, oral administration, in 1 ml corn oil) in vivo was monitored by measuring radioactivity and selenium content. On the basis of radioactivity, approximately 20% of the dose was excreted in the urine and 68% in the feces over 3 days. The cumulative percentages of the dose excreted over 7 days were 24% in urine and 75% in feces, similar to excretion rates of selenium. According to selenium measurement, <1% of the dose was detected in exhaled air; radioactivity was not detected. Only 15% of the dose was extractable from the feces with EtOAc and was identified as tetraselenocyclophane (TSC). Most of the radioactivity remained tightly bound to the feces. Approximately 10% of this bound material converted to TSC on reduction with NaBH4. Organic soluble metabolites in urine did not exceed 2% of the dose; sulfate (9 % of urinary metabolites) and glucuronic acid (19.5% of urinary metabolites) conjugates were observed but their structural identification is still underway. Co-chromatography with a synthetic standard led to the detection of terephthalic acid (1,4-benzenedicarboxylic acid) as a minor metabolite. The major urinary conjugates contained selenium. Despite the low levels of selenium in the exhaled air, the reductive metabolism of p-XSC to H2Se cannot be ruled out. Identification of TSC in vivo indicates that a selenol may be a key intermediate responsible for the chemopreventive action of p-XSC.     

Antibodies to the food mutagens, 2-amino-1-methyl-6-phenylimidazo [4,5-b]pyridine and 2-amino-3,4,8-trimethylimidazo[4,5-f]quinoxaline: useful for immunoassay and immunoaffinity chromatography of biological samples

Monoclonal mouse IgG1 and IgG3 antibodies were developed to the food mutagens, 2-amino-1-methyl-6-phenylimidazo[4,5-b] pyridine (PhIP) and 2-amino-3,4,8-trimethylimidazo[4,4-f] quinoxaline (4,8-DiMeIQx) in order to make specific and sensitive detection and purification systems suitable for biological samples. The antibodies were developed with the strategy that cross-reaction with analogues modified in the N2-position was desirable. Competitive enzyme-linked immunosorbent assays (ELISA) with 50% inhibition by 0.4-6 pmol food mutagen were developed. The epitopes recognized by the antibodies have been characterized by ELISA using 52 synthetic analogues and metabolites of PhIP, 4,8-DiMeIQx, and other food mutagens. One of the anti-PhIP antibodies only recognizes PhIP and those PhIP-analogues which have minor modifications in the N2-amino group, whereas the other, 7B7-1, is less stringent and also recognizes several other modified metabolites, including bulky adducts at the N2-amino group e.g. the major guanine and deoxyguanosine adducts isolated from PhIP-modified DNA. The antibodies to DiMeIQx also recognize the food mutagens 2-amino-3,4-dimethylimidazo[4,5-f]quinoxaline (4-MeIQx), 2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline (8-MeIQx), and the corresponding quinolines (4-MeIQ and 8-MeIQ). Two of these antibodies only bind analogues with minor modifications in the free amino group, whereas analogues with major modifications in this position, including a deoxyguanosine adduct, react with the third antibody. Urine samples and faecal extracts from 3H-PhIP or 2-14C-DiMeIQx dosed rats were analysed by these ELISA assays, and high correlations between radioactivity and response in the ELISA assays were observed. Urine samples and faecal extracts from 3H-PhIP-dosed rats were purified on an affinity column containing the less stringent anti-PhIP antibody, 7B7-1. The affinity column was found by high performance liquid chromatography (HPLC) analysis to concentrate exclusively labelled material. This affinity column also bound PhIP-related materials from dilute samples of acid hydrolysed PhIP-DNA with high efficiency. Only approximately 40% of the 4,8-DiMeIQx related materials found in dilute acid hydrolysed samples of 4,8-DiMeIQx-DNA was bound by an affinity column containing the less stringent anti-4,8-DiMeIQx antibody, 2C5-1. We conclude that our anti-PhIP and anti-DiMeIQx antibodies can be used to determine the presence of these food mutagens and some of their activated or conjugated metabolites in complex biological samples.     

Opioid-immune interactions in autism: behavioural and immunological assessment during a double-blind treatment with naltrexone

The chemical derivatization of biologically active microbial metabolites continues to be a promising approach to the identification of new drugs. We recently synthesized the novel antiproliferative compound SDZ 281-977, 5-[2-(2,5-dimethoxy-phenyl)ethyl]-2-hydroxy-benzoic acid methylester, a derivative of the EGF receptor tyrosine kinase inhibitor lavendustin A. Here we report on our studies of the anticancer efficacy and the mode of action of SDZ 281-977. The growth of both the human pancreatic tumor cells MIA PaCa-2 and the human vulvar carcinoma cells A431 was inhibited in the low micromolar range. Tumors from these cells were induced in nude mice and were shown to respond to orally or intravenously administered SDZ 281-977. In contrast, no antitumor effect was detected in rats bearing dimethylbenzanthracene-induced mammary tumors. Studies in mice indicated that SDZ 281-977 was neither immunosuppressive nor hematosuppressive at doses effectively inhibiting tumor growth. Surprisingly, the mode of action of SDZ 281-977 apparently does not involve inhibition of EGF receptor tryosine kinase, because, in contrast to lavendustin A, SDZ 281-977 failed to inhibit this enzyme in a cell-free assay. The mechanism of the antiproliferative effect can be explained on a cellular level by the ability of the compound to arrest cells in mitosis. SDZ 281-977 is thus the first example of an antimitotic agent derived from the potent tyrosine kinase inhibitor lavendustin A. The therapeutic potential of SDZ 281-977 is enhanced by the fact that it is not subject to multidrug resistance, because tumor cells expressing the multidrug resistance phenotype were as sensitive to SDZ 281-977 as their nonresistant counterparts. In conclusion, SDZ 281-977 represents a novel lavendustin A derivative with potent antiproliferative properties in vitro and in vivo that may be explained on the basis of its antimitotic effects. SDZ 281-977 may be a candidate drug for the treatment of selected cancers, including those expressing the multidrug resistance phenotype.     

In vivo imaging of brain incorporation of fatty acids and of 2-deoxy-D-glucose demonstrates functional and structural neuroplastic effects of chronic unilateral visual deprivation in rats

Regional cerebral 'incorporation coefficients' k* of each of 3 labeled long-chain fatty acids -[9,10-3H]palmitate ([3H]PA), [1-14C]arachidonate ([14C]AA) and [1-14C]docosahexaenoate ([14C]DHA)-were measured using quantitative autoradiography in 11 bilateral brain visual areas of 3.5-month-old awake, hooded, Long-Evans rats, and were compared with regional cerebral metabolic rates for glucose (rCMRglc). The rats, which had undergone unilateral orbital enucleation at 15 days of age, were studied either in the dark with eyelids of the intact eye sutured, or when stimulated in a light box with the intact eye open. rCMRglc did not differ between homologous contralateral and ipsilateral visual areas in the dark or during stimulation, but was elevated bilaterally by 25% or more in many visual areas during stimulation compared with dark. Contralateral compared with ipsilateral k* was lower for each fatty acid tracer in superficial gray of the superior colliculus (in dark and during stimulation) and dorsal nucleus of lateral geniculate body (during stimulation). In the dark, k* for [3H]PA was correlated significantly with rCMRglc for the 22 visual areas studied, whereas during stimulation k* for [14C]AA was correlated with rCMRglc. These results suggest that central neuroplastic changes following chronic unilateral enucleation are accompanied by reduced incorporation of [3H]PA, [14C]AA and [14C]DHA into contralateral brain ares that normally receive crossed retinofugal fibers, and by symmetry of rCMRglc in the dark but increased bilateral symmetrical responsiveness of rCMRglc to visual stimulation of the intact eye.