Immunological methods for dosimetry of heterocyclic amines.
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Adult↗
Biomedical subjects
H Frandsen
Publications and source records attributed to H Frandsen.
Biotransformation of the cyclopenta-fused polycyclic aromatic hydrocarbon benz[j]aceanthrylene in isolated rat liver cell: identification of nine new metabolites.
The biotransformation of benz[j]aceanthrylene (B[j]A) was studied in suspensions of hepatocytes isolated from Aroclor 1254-treated or untreated rats. Using radiolabeled cofactors and metabolic inhibitors combined with UV, mass and 1H-NMR spectroscopy, we have detected five known metabolites and characterized nine new metabolites: metabolite 1 was tentatively assigned as B[j]A-1,2-dihydrodiol-8-sulfate; metabolite 2, B[j]A-1,2,9,10-tetrahydrotetrol; metabolite 3, B[j]A-1,2-dihydrodiol-10-O-glucuronide; metabolite 4, B[j]A-1-one-8-sulfate; metabolite 5, B[j]A-1,2-dihydrodiol-10-sulfate; metabolite 6, the sulfate conjugate of B[j]A-dihydrodiol-phenol; peak 7 in the chromatogram is a mixture of one glutathione conjugate and two sulfate conjugates of a B[j]A-metabolite; metabolite 8, B[j]A-10-O-glucuronide; metabolite 8', B[j]A-1,2-dihydrodiol; metabolite 9, B[j]A-10-sulfate; metabolite 9', B[j]A-9,10-dihydrodiol and metabolite 10, B[j]A-9,10-dihydro-9-hydroxy-10-sulfate. The metabolites identified support the notion that epoxidation at the cyclopenta region is an important activation step of B[j]A. Furthermore, sulfation appears to play a very important role in the conversion of hydroxylated B[j]A metabolites into more polar excretable products.
Animals↗
Reproductive performance and changes in blood lipids in breeding females and in growing Watanabe heritable hyperlipidaemic and New Zealand White rabbits.
The aim of the study was to compare the reproductive performance of homozygous and heterozygous WHHL and NZW females, to record the changes in blood lipids due to gestation and lactation in the heterozygous WHHL and NZW females, and to investigate the changes in blood lipids from 4 to 11 weeks of age in homozygous and heterozygous WHHL and NZW rabbits. The conception rate of homozygous WHHL females was 14% lower than that of NZW and heterozygous WHHL females. The litter size and the weaning rate of homozygous WHHL females were significantly lower than in NZW and heterozygous WHHL females. In heterozygous WHHL and NZW females the total cholesterol was lower during the gestation and lactation while the triglycerides were higher during the gestation and were the lowest during lactation when compared to the levels at mating. In growing homozygous WHHLs of both sexes the total cholesterol did not change from 4 to 11 weeks of age. The triglycerides remained unchanged in males but decreased in females at 11 weeks of age. At this age the triglycerides were significantly lower in females than in males. In growing heterozygous WHHLs of both sexes the total cholesterol and triglycerides decreased with age. At 11 weeks of age the blood lipids were significantly higher in females than in males. The triglyceride levels in homozygous and heterozygous WHHL males and females were comparable at 4 and 6 weeks of age but significantly lower in heterozygous WHHLs at 11 weeks of age. In growing NZW rabbits of both sexes the blood lipids decreased significantly with age but no sex difference was recorded. The blood lipids in the heterozygous WHHLs of both sexes were higher than in NZWs during weaning. At 11 weeks of age the blood lipids of the males and triglycerides of the females of both strains were comparable. Only the total cholesterol remained higher in heterozygous WHHL females. The decrease with age in blood lipids in growing rabbits should be kept in mind when designing experiments beginning in animals younger than 3 months of age.
Aging↗
DNA-binding and disposition of 2-amino-1-methyl-6-phenylimidazo[4,5-b] pyridine (PhIP) in the rat.
Untreated and Aroclor 1254-pretreated male Wistar rats were given a single dose of 1.0 mg/kg body weight of randomly tritium-labelled 2-amino-1-methyl-6-phenylimidazo[4,5-b] pyridine (3H-PhIP) by oral intubation. Urine and faeces were collected at 24, 48 and 72 hours after dosing, and total radioactivity determined. At 2, 4, 6, 16, 26, 48 and 72 h, animals were killed and several organs, including liver, bladder, lungs, kidney, stomach, large and small intestines, heart, thigh muscles, spleen and blood were collected for DNA extraction and for determination of total radioactivity. Highest total radioactivity at 2 h was not unexpectedly observed in the stomach, small intestines and bladder, whereas radiolabels corresponding to approximately 2.5 nmol PhIP/g of kidney and liver showed the highest levels observed at 24 h. Several tissues, including blood, plasma, liver and muscles had a slightly bimodal time-distribution of radioactivity showing a second peak at 16-24 h. At 72 h after a single dose of PhIP, highest radioactivity was observed in the liver and the large intestine (0.4 nmol PhIP/g tissue), whereas most other organs, irrespective of pretreatment had levels at approximately 0.2 nmol/g of tissue. At earlier time points, Aroclor 1254-treated rats had lower amounts of radiolabel in all tissues. Radioactivity bound to DNA was determined by high sensitivity scintillation counting. In contrast to total radioactivity, DNA-associated radioactivity was generally higher in the Aroclor 1254-treated rats, most notably in the heart, but levels had decreased to approximately the same level in controls and in Aroclor 1254-treated rats at 72 h. DNA-binding was highest at 2-6 h after dosing, highest in the heart of Aroclor 1254-treated animals at 6 h (120 adducts/10(8) bases) followed by thigh muscle at 4-6 h (approximately 50 adducts/10(8) bases, irrespective of pretreatment). Levels were approximately 1.5-3 times lower in other organs at 2-6 h after dosing. At 72 h, radioactivity associated with DNA was again highest in the heart of Aroclor 1254-treated rats (20 adducts/10(8) bases) and 5-10 times lower in most other organs, approaching the detection limit. Total DNA was extracted from the livers of PhIP dosed rats at 4 ad 72 h. DNA was hydrolysed, affinity-concentrated, and analysed by liquid chromatography. A radiolabelled peak had identical retention time and UV-spectral characteristics as peaks isolated by affinity chromatography and HPLC of acid-hydrolysed synthetic PhIP-DNA and PhIP-deoxyguanosine adduct.(ABSTRACT TRUNCATED AT 400 WORDS)
Animals↗
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.
Animals↗
Repair of DNA lesions induced by ultraviolet irradiation and aromatic amines in normal and repair-deficient human lymphoblastoid cell lines.
A host cell reactivation (HCR) assay was employed to study the capacity of a normal and three repair-deficient human lymphoblastoid cell lines to repair DNA damage induced by UV irradiation and the aromatic amines 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) and N-acetyl-2-aminofluorene (AAF) respectively. The cell line belonging to xeroderma pigmentosum complementation group C (XP-C) removed all three types of damage less efficiently than the normal cell line, but more efficiently than the cell line belonging to xeroderma pigmentosum complementation group D (XP-D). The cell line belonging to complementation group B of Cockayne's syndrome (CS-B) showed reduced host cell reactivation. Fibroblasts from CS-B patients have reduced gene-specific DNA repair, but normal total genomic DNA repair, thus our data suggest that the HCR assay measures the capacity for gene-specific DNA repair. In the XP-D cell line, which had practically no DNA repair capacity, AAF adducts had a more potent inhibitory effect on gene expression than UV and PhIP adducts. When corrected for this inhibitory effect, the wild-type, XP-C and CS-B cell lines repaired low levels of AAF and UV adducts with similar efficiencies, however, PhIP adducts were repaired less efficiently.
2-Acetylaminofluorene↗
(-)-anipamil retards atherosclerosis in Watanabe heritable hyperlipidemic rabbits.
Calcium antagonists have been reported to limit atherosclerosis in cholesterol fed rabbits. The purpose of this study was to examine the effect of the calcium antagonist (-)-anipamil on the spontaneous development of atherosclerosis in homozygote WHHL rabbits. From the age of 7 weeks, three groups of rabbits received 0 (control, n = 11), 1 mg/kg body weight (n = 11), or 10 mg/kg body weight (n = 12) of (-)-anipamil for 17 weeks. Serum cholesterol at start of the investigation was 18.8 +/- 1.2 (mean +/- SEM), 20.5 +/- 1.8, and 19.5 +/- 0.8 mM, respectively, in the three groups. No significant differences were found in serum lipids (i.e., VLDL, IDL, LDL, HDL) in the study period among the three groups. Plasma anipamil at the end of the study was 0.23 +/- 6, and 202 +/- 19 ng/ml, respectively, in the three treatment groups. The degree of atherosclerosis in the abdominal aorta was significantly lower (p < 0.02) in the 10 mg/kg body weight (-)-anipamil group compared with the two other groups. No significant difference was found in the thoracic aorta. It is concluded that (-)-anipamil retards development of atherosclerosis in the abdominal aorta in WHHL rabbits.
Animals↗
Altered peripheral vasodilator profile of nitroglycerin during long-term infusion of N-acetylcysteine.
OBJECTIVES: The aim of this study was to compare the short- and long-term effects of intravenous nitroglycerin plus placebo and nitroglycerin plus N-acetylcysteine on peripheral arteries, veins and microcirculation in humans. BACKGROUND: The thiol donor N-acetylcysteine may potentiate the hemodynamic response to nitrates in nitrate-tolerant and nontolerant patients. The vascular changes responsible for this effect are not clear. METHODS: Eight male volunteers were treated with nitroglycerin (0.1 microgram/kg per min) combined with N-acetylcysteine (2 g intravenously, followed by 5 mg/kg per h) or placebo for 23 h in a double-blind, randomized, crossover study. Venous volume, the diameter of the radial and temporal arteries, calf blood flow and subcutaneous blood flow were measured at baseline and repeated after 1 and 23 h of infusion. RESULTS: Prolonged coadministration of N-acetylcysteine and nitroglycerin potentiated the acute venodilator effect of nitroglycerin as estimated by changes in venous volume (nitroglycerin plus N-acetylcysteine, 4.45 +/- 0.36 ml/100 g; nitroglycerin plus placebo, 3.65 +/- 0.46 ml/100 g, mean +/- SEM, p < 0.05) and prevented development of tolerance as seen after 23 h of treatment with nitroglycerin plus placebo (4.35 +/- 0.25 vs. 3.47 +/- 0.41 ml/100 g, p < 0.05). N-acetylcysteine had no effect on nitroglycerin-induced changes in arterial diameters (p > 0.05) but significantly increased microcirculatory subcutaneous blood flow after 1 h (nitroglycerin plus N-acetylcysteine: 6.3 +/- 1.3 ml/100 g per min vs. nitroglycerin plus placebo: 3.5 +/- 0.3 ml/100 g per min, p < 0.05) and after 23 h (4.4 +/- 0.6 vs. 3.1 +/- 0.5 ml/100 g per min, p < 0.05). CONCLUSIONS: The results suggest that coadministration of nitroglycerin and N-acetylcysteine in humans 1) potentiates and preserves nitroglycerin-induced venodilation and 2) augments the effect of nitroglycerin on small resistance vessels (regulating subcutaneous blood flow) without affecting the response to nitroglycerin in middle-sized arteries. Both the development of nitrate tolerance and the administration of N-acetylcysteine significantly change the normal vasodilator profile of nitroglycerin in humans.
Acetylcysteine↗
In vitro formation and degradation of 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) protein adducts.
Adduct formation between the food mutagen 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) and rat serum albumin (RSA) was studied in vitro using hepatic microsomes isolated from polychlorinated biphenyl-induced rats. With 1-methyl-2-nitro-6-phenylimidazo[4,5-b]pyridine (2-nitro-PhIP) as starting material, four main products were formed. Pretreatment of RSA with beta-mercaptoethanol markedly increased the yield of one of them. In this adduct, the C-2 of PhIP was linked to cysteine of RSA at position 34 in a C-S linkage. With N2-acetoxy-PhIP as starting material, unstable conjugates were formed with RSA as well as with glutathione (GSH) and cysteine. The suggested structures of the GSH and cysteine conjugates, GSH-S-N2-PhIP and cysteine-S-N2-PhIP respectively, are based on mass spectra and UV spectra. The degradation of the conjugates of GSH and cysteine as well as of the protein adduct were monitored. They all resulted in the same degradation product, identified as 2-amino-5-hydroxy-1-methyl-6-phenylimidazo[4,5-b]pyridine (5-hydroxy-PhIP).
Animals↗
Formation of DNA adducts by the food mutagen 2-amino-3,4,8-trimethyl-3H-imidazo[4,5-f]quinoxaline (4,8-DiMeIQx) in vitro and in vivo. Identification of a N2-(2'-deoxyguanosin-8-yl)-4,8-DiMeIQx adduct.
The covalent binding of the mutagenic N2-hydroxy metabolite of the food mutagen 2-amino-3,4,8-trimethyl-3H-imidazo[4,5-f]quinoxaline (4,8-DiMeIQx) to 2'-deoxynucleosides and DNA was investigated in vitro and in vivo. N2-Hydroxy-4,8-DiMeIQx reacted to a small extent spontaneously with 2-deoxyguanosine. However, acetylation of N2-hydroxy-4,8-DiMeIQx with acetic anhydride to form the N2-acetoxy derivative prior to reaction with 2-deoxyguanosine resulted in much higher yield of adduct. N2-Acetoxy-4,8-DiMeIQx did not form adducts with 2'-deoxyadenosine, 2'-deoxycytidine or 2'-deoxythymidine. The adduct formed between the N2-OH metabolite of 4,8-DiMeIQx and 2-deoxyguanosine was analysed by mass spectrometry and NMR spectroscopy and the structure of the adduct was shown to be N2-(2'-deoxyguanosin-8-yl)-4,8-DiMeIQx. N2-Acetoxy-4,8-DiMeIQx reacted with calf thymus DNA and formed a covalently bound 4,8-DiMeIQx residue, which could not be removed by repeated precipitations or solvent extractions. The 4,8-DiMeIQx-DNA was hydrolysed enzymatically with nuclease P1/acid phosphatase and HPLC analysis showed that 70% of the bound mutagen was recovered as N2-(2'-deoxyguanosin-8-yl)-4,8-DiMeIQx. An additional minor adduct accounting for approximately 15% of the bound mutagen showed UV spectral characteristics similar to N2-(2'-deoxyguanosin-8-yl)-4,8-DiMeIQx and is probably an undigested oligomer. 32P-Postlabelling analysis of calf thymus DNA modified with 4,8-DiMeIQx in vitro and liver DNA from rats dosed with 50 mg/kg 4,8-DiMeIQx showed a similar adduct pattern. In both samples N2-(2'-deoxyguanosin-8-yl)-4,8-DiMeIQx accounted for 60-70% of the bound mutagen. Thus, these results show that 4,8-DiMeIQx similar to other heterocyclic amines form adducts with C-8 of guanine both in vitro and in vivo via its N2-OH metabolite.
Acetylation↗
Microsomal metabolism of the food mutagen 2-amino-3,4,8-trimethyl-3H-imidazo[4,5-f]-quinoxaline to mutagenic metabolites.
Heterocyclic aromatic amines are formed in the crust of meat during ordinary cooking. These aromatic amines are potent bacterial mutagens and also potent rodent carcinogens. 2-Amino-3,4,8-trimethyl-3H-imidazo[4,5-f]quinoxaline (DiMeIQx) is one of the more abundant heterocyclic aromatic amines, accounting for approximately 20% of the mutagenic material found in cooked meat. DiMeIQx is metabolically activated, by hepatic microsomes from PCB treated rats, to two major and three minor metabolites. One major and one minor metabolite were identified as 2-hydroxyamino-3,4,8-trimethyl-3H-imidazo[4,5-f]quinoxaline and 3,4,8-trimethyl-2-nitro-3H-imidazo[4,5-f]quinoxaline, respectively, confirmed by comparison of HPLC retention times, and UV and mass spectra of synthetic standards. Both metabolites were mutagenic in Salmonella typhimurium TA98 without metabolic activation. The other major metabolite was identified as 2-amino-8-hydroxymethyl-3,4-dimethyl-3H-imidazo[4,5-f]quinoxaline by mass and NMR spectral analysis. The two remaining minor metabolites were identified as the 2-hydroxyamino- and 2-nitro- derivatives of 2-amino-8-hydroxymethyl-3,4-dimethyl-3H-imidazo]4,5-f]quinoxaline by UV and mass spectral analysis. Both of these metabolites were mutagenic in S. typhimurium TA98 without metabolic activation.
Animals↗
Atherosclerosis in Watanabe heritable hyperlipidaemic rabbits. Evaluation by macroscopic, microscopic and biochemical methods and comparison of atherosclerosis variables.
The spontaneous development of atherosclerotic disease in 38 homozygous and 34 heterozygous Watanabe heritable hyperlipidaemic rabbits was evaluated by qualitative and quantitative light microscopy in aorta, coronary, pulmonary and renal arteries, by naked eye and macroscopic morphometric estimation of aortic atherosclerosis extent and by biochemical analysis of aortic cholesterol content. No noteworthy atherosclerosis was demonstrated within 19 months in heterozygous rabbits. In homozygous rabbits, atherosclerotic lesions were seen from the age of 4 months and progressed with age. All 19-month-old rabbits had severe atherosclerotic disease. As much as 64% of the variation in atherosclerosis extent/severity could be explained by serum cholesterol and age. A highly significant correlation between the various methods for quantitation of atherosclerosis extent and/or severity was demonstrated, suggesting that quantitative microscopy, macroscopic morphometry and determination of aortic cholesterol content may be equally valid as a measure of atherosclerosis in WHHL rabbits and are therefore interchangeable.
Animals↗
Bioactivation of 2-amino-1-methyl-6-phenylimidazo[4,5-b]-pyridine by liver microsomes from three different rat strains.
The biotransformation of 2-amino-1-methyl-6-phenylimidazo[4,5-b]-pyridine (PhIP) and the protein binding of PhIP and 2-amino-3-methylimidazo[4,5-f]quinoline (IQ) was studied using microsomes from PCB-pretreated or untreated male rats of the strains, Wistar, Fischer and Sprague-Dawley. The microsomal monooxygenases, P450IA1 and IA2, which are important for the biotransformation of heterocyclic amines, were quantified by immunoblots. The two metabolites detected, 2-amino-1-methyl-6-(4'-hydroxyphenyl)imidazo[4,5-b]pyridine (4'OH-PhIP) and 2-hydroxyamino-1-methyl-6-phenylimidazo[4,5-b]pyridine (N2-OH-PhIP) were formed in similar amounts whereas no minor metabolites were found in our highly sensitive radiochemical assay. Irrespective of the rat strain used, pretreatment with PCB significantly induced both the activation and the detoxication in all three rat strains. Except for a significantly higher concentration of P450IA2 in microsomes from control and PCB induced Wistar rats, no major differences between the strains were found.
Animals↗
Reaction of the N2-acetoxy derivative of 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) with 2'-deoxyguanosine and DNA. Synthesis and identification of N2-(2'-deoxyguanosin-8-yl)-PhIP.
The direct acting mutagenic N2-hydroxylated metabolite of the food mutagen 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) does not react with DNA. Upon acetylation of the N2-hydroxy-PhIP with acetic anhydride two products could be detected. Mass spectrometric analysis showed that both products were monoacetyl derivatives of N2-hydroxy-PhIP. One of the products did not show any reactivity towards DNA and is probably the N-acetyl derivative of N2-hydroxy-PhIP. The other product which is most likely to be N2-acetoxy-PhIP reacted with DNA and 2'-deoxyguanosine but not with 2'-deoxycytidine, 2'-deoxyadenosine or 2'-deoxythymidine. The PhIP-2'-deoxyguanosine adduct was purified and characterized by mass spectral, 1H and [13C]NMR analysis, showing that PhIP like the other cooked food mutagen 2-amino-3-methylimidazo[4,5-f]quinoline, had reacted with C-8 of guanine forming N2-(2'-deoxyguanosin-8-yl)-PhIP. HPLC analysis of enzymatically hydrolyzed calf thymus DNA which had been reacted with N2-acetoxy-PhIP showed one adduct which was chromatographically and spectroscopically identical to N2-(2'-deoxyguanosin-8-yl)-PhIP. HPLC separation followed by liquid scintillation counting of hydrolyzed liver DNA from a rat dosed with [3H]PhIP showed that radioactivity coeluted with the hydrolysis product of the synthetic PhIP-2-deoxyguanosine adduct, indicating that PhIP in vivo also forms an N2-(2'-deoxyguanosin-8-yl)-PhIP adduct.
Animals↗
Effects of alpha-adrenoceptor and of combined sympathetic and parasympathetic blockade on cardiac performance and vascular resistance.
1. Cardiac performance and vascular resistance was studied in seven healthy men by radionuclide cardiography and venous plethysmography before and after alpha-adrenoceptor blockade with phentolamine and after combined alpha-adrenoceptor, beta-adrenoceptor (propranolol) and parasympathetic (atropine) blockade. 2. During alpha-adrenoceptor blockade heart rate and cardiac output increased considerably and left ventricular ejection fraction increased because of increased contractility. Systemic vascular resistance fell both during alpha-adrenoceptor blockade alone and during combined blockade. The increase in calf blood flow was of the same magnitude after combined blockade and after alpha-adrenoceptor blockade alone, and was considerably higher than the fall in systemic vascular resistance. Plasma catecholamine concentrations increased after phentolamine, but the changes were blunted when propranolol and atropine were added. 3. These results indicate that peripheral vasoconstriction especially that exerted by alpha-adrenoceptor nervous tone in skeletal muscle restricts left ventricular emptying of the intact heart. During pharmacologic blockade of the sympathetic and parasympathetic nervous system at rest the chronotropic state is augmented, whereas preload and inotropy are unaffected.
Adrenergic alpha-Antagonists↗
Metabolic formation, synthesis and genotoxicity of the N-hydroxy derivative of the food mutagen 2-amino-1-methyl-6-phenylimidazo (4,5-b) pyridine (PhIP).
Hepatic microsomes from rats pretreated with PCB were found to metabolize the food mutagen 2-amino-1-methyl-6-phenylimidazo(4,5-b)pyridine (PhIP) to two major metabolites, one of which was identified as the N-hydroxy derivative, 2-hydroxy-amino-1-methyl-6-phenylimidazo(4,5-b)pyridine (N-OH-PhIP). This identification was based on mass spectral (MS), UV and HPLC data by comparison with N-OH-PhIP prepared by chemical synthesis, as well as the specific activity of the compound in the Ames Salmonella test. Synthetic N-OH-PhIP was prepared by catalytic reduction of the nitro derivative of PhIP, which was synthesized from PhIP by diazotization and reaction with sodium nitrite. N-OH-PhIP was mutagenic to Salmonella typhimurium TA98 without metabolic activation and had a specific mutagenic activity of 2700 revertants/nmol. N-OH-PhIP thus seems to be a proximate mutagenic metabolite of PhIP. Other direct acting mutagens were not detected in the microsomal incubation mixture after HPLC separation. N-OH-PhIP also induced sister chromatid exchange (SCE) in Chinese hamster ovary cells (CHO cells) without metabolic activation. The specific activity of N-OH-PhIP in this assay was approximately 3 times higher than the activity of PhIP with microsomal activation.
Animals↗
Plasma volume changes during hypoglycaemia: the effect of autonomic blockade.
To investigate the role of the autonomic nervous system in changes in blood volume and composition induced by acute hypoglycaemia in man, seven healthy subjects participated in three experiments on separate days: insulin-induced hypoglycaemia with concomitant alpha-adrenoceptor blockade; insulin-induced hypoglycaemia with total autonomic blockade (alpha-adrenoceptor blockade combined with beta-adrenoceptor blockade and atropine); and insulin-induced hypoglycaemia without any autonomic blockade. In the experiments without autonomic blockade the peripheral venous hematocrit increased, plasma volume decreased, intravascular albumin content decreased and the transcapillary escape rate of albumin increased. In both experiments with autonomic blockade the increase in venous haematocrit was abolished, yet plasma volume decreased, intravascular albumin content decreased and the transcapillary escape rate of albumin increased in these experiments. Thus, the changes in plasma volume and composition in response to hypoglycaemia are due to the combined actions of adrenaline and of insulin.
Adult↗
Effect of insulin-induced hypoglycaemia on absorption of unmodified insulin after subcutaneous or intramuscular injection.
The effect of insulin-induced hypoglycaemia on the absorption of iodine-125 labelled unmodified insulin (10 U) from thigh after subcutaneous or intramuscular injection was studied in eight immobilized, supine, normal subjects. Ultrasonic determination of the subcutaneous thickness was used to accurately localize the site for insulin injection. Insulin absorption was studied twice during hypoglycaemia or normoglycaemia in random order. Insulin absorption was similar after subcutaneous and intramuscular injections (residual activity at 5 h: SC 59.3 +/- 5.0 (+/- SE) %; IM 55.2 +/- 3.7%). Hypoglycaemia did not change the disappearance rate of iodine-125 insulin after either subcutaneous (55.0 +/- 4.4%) or intramuscular injection (52.6 +/- 5.7%), despite a plasma glucose nadir of 1.7 +/- 0.2 mmol I-1. In a controlled study under standardized conditions hypoglycaemia has no effect on insulin absorption rate.(ABSTRACT TRUNCATED AT 250 WORDS)
Absorption↗