Iron concentrations in liver and kidney of cadmium-exposed human subjects

Iron (Fe) concentrations in liver and kidney of cadmium (Cd)-exposed people (11 Itai-Itai patients, 13 persons requiring observation and 4 Cd-exposed persons) were compared with those of nonexposed people. No significant difference in Fe levels in liver was observed between the groups. This fact indicates that anaemia in environmentally Cd-exposed people was not due to iron deficiency in contrast to findings in animal experiments.     

Methylation of captopril in human liver,kidney and intestine

1. The methylation of captopril was studied in the microsomal fraction from 20 human liver, 12 kidney, and 14 intestinal rnucosa specimens.

2. The hepatic methyltransferase activity (mean ± SD) was 477 ± 204 pmol/min per mg. Renal and intestinal methyltransferase activities were 3 and 8 times lower, respectively, than hepatic activity.

3. The kinetics of methyltransferase with captopril as substrate were studied in four specimens of liver, kidney and intestine. The maximum velocities of reaction (mean ± SD; pmol/min per?mg) were 697 ± 219 (liver), 456 ± 120 (renal cortex), 264 ± 77 (renal medulla) and 101 ± 28 (ileum mucosa). K_m values (mean ± SD; mM) were 5.2±2.3 (liver) 4.3±1.7 (renal cortex) 4.1±1.5 (renal medulla) and 5.3 ± 2.0 mM (ileum mucosa). V_max is subjected to a marked tissue dependence whereas K_m is similar in all tissues.

4. Liver is the primary site of captopril methylation whereas the intestine plays only a minor role. Kidney may contribute substantially to the hepatic methylation of captopril.     

Lauric acid hydroxylation in human liver and kidney cortex microsomes

The ω- and (ω- 1 )-hydroxylation of the medium-chain fatty acid, dodecanoic or lauric acid, was studied in liver and kidney cortex microsomes from seven human cadavers. The rates of laurate hydroxylation in human liver microsomes were found to exceed the rates recorded in human kidney cortex microsomes by 4-to 30-fold. The mean specific activity of laurate hydroxylation from the seven human kidneys was six to fourteen times lower than the specific activities found in pig, rat or hamster kidney microsomes. The effects of several known inhibitors of the liver microsomal cytochrome P-450-dependent mono-oxygenase system were also studied. Metyrapone preferentially inhibited the (ω- 1)-hydroxylase activity of human liver microsomes, but did not affect the ω-hydroxylation reaction. In the presence of 7,8-benzoflavone, the human liver microsomal (ω- 1 )-hydroxylase activity was stimulated, but an inhibitory effect was observed on the ω-hydroxylation reaction. 2-Diethylaminoethyl-2,2-diphenylvaIerate (SKF 525A) inhibited both hydroxylase activities in human liver microsomes. Neither metyrapone nor SKF 525A inhibited the laurate hydroxylation reactions catalyzed by human kidney microsomes. These studies indicate that the cytochrome P-450-mediated hydroxylations of medium chain fatty acids in human kidney cortex microsomes are much less active than in kidneys of other species investigated. The effects of the inhibitors, metyrapone and SKF 525A, on ω- and (ω- 1)-hydroxylation of laurate in human liver and kidney microsomes were similar to the effects reported in other mammalian species.     

Captopril methylation in human liver and kidney: interindividual variability

1. The methylation of captopril was studied in the microsomal fraction obtained from 87 specimens of human liver and 70 specimens of human renal cortex.

2. The rate of captopril methylation ranged over one order of magnitude in the liver and kidney. In the human liver, the mean (± SD) rate of captopril methylation (pmol/min/mg) was significantly (p < 0·001) greater women (199 ± 97) than in men (126 ± 88), whereas in the kidney no sex-difference was observed, and the mean (± SD) of all cases was 47 ± 23 pmol/min/mg.

3. In the kidney, the statistical analysis revealed the presence of two subgroups in the rate of captopril methylation and their mean (± SD) estimates were 42·5 ± 13·9 and 90·3 ± 12·0 pmol/min/mg (p < 0·05). Of the population, 84 % fell in the former and the remaining 16 % in the latter subgroup.

4. Captopril is mainly eliminated by metabolism and its bioavailability is 65 %. Methylation is one of the metabolic routes of captopril and its variability may contribute, to some extent, to modulate the intracellular concentration of this drug.     

Methylation of captopril in human liver, kidney and intestine.

1. The methylation of captopril was studied in the microsomal fraction from 20 human liver, 12 kidney, and 14 intestinal mucosa specimens. 2. The hepatic methyltransferase activity (mean +/- SD) was 477 +/- 204 pmol/min per mg. Renal and intestinal methyltransferase activities were 3 and 8 times lower, respectively, than hepatic activity. 3. The kinetics of methyltransferase with captopril as substrate were studied in four specimens of liver, kidney and intestine. The maximum velocities of reaction (mean +/- SD; pmol/min per mg) were 697 +/- 219 (liver), 456 +/- 120 (renal cortex), 264 +/- 77 (renal medulla) and 101 +/- 28 (ileum mucosa). Km values (mean +/- SD; mM) were 5.2 +/- 2.3 (liver) 4.3 +/- 1.7 (renal cortex) 4.1 +/- 1.5 (renal medulla) and 5.3 +/- 2.0 mM (ileum mucosa). Vmax is subjected to a marked tissue dependence whereas Km is similar in all tissues. 4. Liver is the primary site of captopril methylation whereas the intestine plays only a minor role. Kidney may contribute substantially to the hepatic methylation of captopril.     

Glucuronidation of thyroxine in human liver, jejunum, and kidney microsomes.

Glucuronidation of thyroxine is a major metabolic pathway facilitating its excretion. In this study, we characterized the glucuronidation of thyroxine in human liver, jejunum, and kidney microsomes, and identified human UDP-glucuronosyltransferase (UGT) isoforms involved in the activity. Human jejunum microsomes showed a lower K(m) value (24.2 microM) than human liver (85.9 microM) and kidney (53.3 microM) microsomes did. Human kidney microsomes showed a lower V(max) value (22.6 pmol/min/mg) than human liver (133.4 pmol/min/mg) and jejunum (184.6 pmol/min/mg) microsomes did. By scaling-up, the in vivo clearances in liver, intestine, and kidney were estimated to be 1440, 702, and 79 microl/min/kg body weight, respectively. Recombinant human UGT1A8 (108.7 pmol/min/unit), UGT1A3 (91.6 pmol/min/unit), and UGT1A10 (47.3 pmol/min/unit) showed high, and UGT1A1 (26.0 pmol/min/unit) showed moderate thyroxine glucuronosyltransferase activity. The thyroxine glucuronosyltransferase activity in microsomes from 12 human livers was significantly correlated with bilirubin O-glucuronosyltransferase (r = 0.855, p < 0.001) and estradiol 3-O-glucuronosyltransferase (r = 0.827, p < 0.0001) activities catalyzed by UGT1A1, indicating that the activity in human liver is mainly catalyzed by UGT1A1. Kinetic and inhibition analyses suggested that the thyroxine glucuronidation in human jejunum microsomes was mainly catalyzed by UGT1A8 and UGT1A10 and to a lesser extent by UGT1A1, and the activity in human kidney microsomes was mainly catalyzed by UGT1A7, UGT1A9, and UGT1A10. The changes of activities of these UGT1A isoforms via inhibition and induction by administered drugs as well as genetic polymorphisms may be a causal factor of interindividual differences in the plasma thyroxine concentration.     

Immunoquantitation of FMO1 in human liver, kidney, and intestine.

To determine the level of FMO1 protein present in human liver tissues, a monospecific antibody was prepared and a sensitive Western blotting procedure with enhanced chemiluminescence detection was developed. Human FMO1, purified from insect cells expressing the recombinant protein, was used as a protein standard for absolute quantification. The average concentrations of FMO1 in microsomes prepared from human liver, kidney, intestine, and fetal liver were found to be <1, 47 +/- 9, 2.9 +/- 1.9, and 14.4 +/- 3.5 pmol/mg, respectively. Quantitation in intestinal microsomes was complicated by variable degrees of proteolytic degradation of FMO1, not seen in microsomes prepared from liver or kidney. Recombinant human FMO1 and detergent-solubilized human duodenal microsomes both metabolized p-tolyl methyl sulfide stereoselectively to the (R)-sulfoxide, indicating the expression of functional FMO1 in human intestine. The relatively high levels of immunoquantifiable FMO1 in human kidney and fetal liver complement our previous catalytic studies in these tissues, which also demonstrated preferential (R)-p-tolyl methyl sulfoxide formation. These data demonstrate a profound ontogenic change in expression of hepatic FMO1 in humans, such that in adult life FMO1 is exclusively an extrahepatic drug-metabolizing enzyme. The marked expression levels of FMO1 found in human kidney coupled to the high catalytic activity of this isoform toward a diverse array of sulfides and tertiary amines suggest the possibility that human renal FMO1 is a significant contributor to the metabolic clearance of drugs and other xenobiotics bearing these functionalities.     

The isolation and characterization of glycosaminoglycans in normal human serum

The optimization of conditions for the isolation and characterization of human serum glycosaminoglycans (GAG) is described, together with studies of the accuracy and reproducibility of the method. The principle of the method is proteolytic digestion of serum using papain followed by precipitation of GAGs from the digested sample with cetyl pyridinium chloride (CPC). The uronic acid level and electrophoretic separations can be obtained from a 5 ml serum sample. The mean CPC-precipitable uronic acid level in pooled normal serum was 10.8 mg l(-1) serum. Using enzymatic and chemical analysis the major serum GAG was shown to be chondroitin sulphate (CS). Two distinct electrophoretic fractions were identified both consisting of CS but differing in their degree of sulphation. Dermatan sulphate, heparan sulphate and hyaluronic acid were not detected.     

Alterations in mouse liver monooxygenases by benzothiadiazoles

Administration of 1,2,3-benzothiadiazoles to mice had a biphasic effect on liver microsomal monooxygenases. During the first 15 hr of treatment, an inhibition of the in vivo metabolism of hexobarbital, as well as of the in vitro hydroxylation of naphthalene and N-demethylation of aminopyrine, was observed. An apparent decrease in cytochrome P-450 and in the activity of the NADPH-cytochrome c reductase also occurred. The levels of cytochrome b₅ and NADH-cytochrome c reductase activity were only slightly affected. A shift to 452 nm in the carbon monoxide difference spectrum was obtained with dithionite-reduced microsomes and this was not modified by ferricyanide. After the initial inhibitory phase, an enhancement of drug-metabolizing activities in vivo and in vitro and in the levels of some components of the mixed function oxidase system was observed. The carbon monoxide difference spectra of dithionite-reduced microsomes returned to a maximal absorption at 450 nm. The stimulatory effect on monooxygenase activity, elicited by benzothiadiazoles, was prevented completely by actinomycin D and was accompanied by increases in liver weight, microsomal protein, and incorporation of labeled amino acids into microsomal protein, as well as by proliferation of smooth and rough endoplasmic reticulum. Acrylamide gel analysis of liver microsomes from mice, given a single dose of 6-chloro-1, 2,3-benzothiadiazole 48 hr prior to being killed, showed preferential induction of cytochrome P-450 apoproteins of 50,000, 52,000 and 53,000 molecular weight.     

Alterations in activity and mRNA expression of acetylcholinesterase in the liver,kidney and gill of common carp exposed to atrazine and chlorpyrifos

Insecticides and herbicides are widely used in modern agricultural production. The intensive use of insecticide chlorpyrifos (CPF) and herbicide atrazine (ATR) has resulted in serious environmental problems. Herein, we investigated alteration in activity and mRNA levels of AChE in the liver, kidney and gill from common carp after 40 d exposure to CPF and ATR alone or in combination and 20 d recovery treatment. Results indicated that activity and mRNA levels of AChE at all high-dose groups have been significantly decreased after CPF and ATR alone or ATR/CPF mixture exposure, and the changes were improved in the end of recovery tests in varying degrees, the activity and gene expression of AChE in the joint toxicity of ATR and CPF groups were significantly lower than that in the single toxicant group. Our study suggests that the decrease of AChE activity observed at all high-dose groups (CPF and ATR alone or in combination) may be directly related to a lower AChE expression, and the joint toxicity of ATR and CPF is higher than ATR and CPF alone.     

人胎肾上腺,肝和肾内谷胱甘肽相关酶活性

目的：通过谷胱甘肽相关酶活性及其在亚细胞分布，了解胎肾上腺在发育期间对活性代谢产物的解毒处理能力。方法：制备肾上腺、肝亚细胞组分。测定谷胱甘肽转移酶（ＧＳＴ）、还原酶、过氧化物酶。结果：ＧＳＴ在胎肾上腺微粒体、线粒体、胞浆中含量分别是肝各亚细胞组分中的３７３％、２７０％和民６７％、肾上腺微粒体ＧＳＴ活性与细胞色素Ｐ－４５０、与氨基比林脱甲基酶活性皆呈正相关，肾上腺线粒体谷胱甘肽还原酶、过氧化物分别     

Glutathione-related enzyme activities in human fetal adrenal,liver,and kidney

目的：通过测定谷胱甘肽相关酶活性及其在亚细胞分布，了解胎肾上腺在发育期间对活性代谢产物的解毒处理能力．方法：制备肾上腺、肝亚细胞组分．测定谷胱甘肽转移酶（ＧＳＴ）、还原酶、过氧化物酶．结果：ＧＳＴ在胎肾上腺微粒体、线粒体、胞浆中含量分别是肝各亚细胞组分中的３７３％、２７０％和１６７％．肾上腺微粒体ＧＳＴ活性与细胞色素Ｐ４５０、与氨基比林脱甲基酶活性皆呈正相关．肾上腺线粒体谷胱甘肽还原酶、过氧化物酶分别是肝线粒体中的５０６％和４８２％．结论：胎肾上腺有比胎肝更大的解毒能力．提示胎肾上腺兼有药物代谢器官的功能．     

Glucuronidation of codeine and morphine in human liver and kidney microsomes: effect of inhibitors

Glucuronidation of codeine was detected and compared with that of morphine in microsomes from human livers and kidneys. Vmax values for codeine-6-glucuronide (C6G) were 0.54 +/- 0.24 and 0.74 +/- 0.35 nmol/mg/min. in the livers and 0.10 and 0.13 nmol/mg/min. in the kidney, respectively, when codeine and UDP-glucuronic acid (UDPGA) were incubated with microsomal preparation. The corresponding Km values were 2.21 +/- 0.68 and 1.41 +/- 0.36 mM in the livers and 6.69 and 4.12 mM in the kidney. The average codeine glucuronyltransferase (GT) activity was 14-fold lower in the six kidneys than in the 11 livers. Higher GT activities were observed in liver microsomes from patients who had been exposed to enzyme inducers. Rates of glucuronide formation from morphine correlated significantly with those from codeine in both human liver and kidney microsomes. Morphine, amitriptyline, diazepam, probenecid and chloramphenicol inhibited codeine glucuronidation with Ki values of 3.6, 0.13, 0.18, 1.7 and 0.27 mM, respectively.     

Glutathione conjugation of trichloroethylene in human liver and kidney: kinetics and individual variation.

Isolated human hepatocytes exhibited time-, trichloroethylene (Tri) concentration-, and cell concentration-dependent formation of S-(1, 2-dichlorovinyl)glutathione (DCVG) in incubations in sealed flasks with 25 to 10,000 ppm Tri in the headspace, corresponding to 0.011 to 4.4 mM in hepatocytes. Maximal formation of DCVG (22.5 +/- 8.3 nmol/120 min per 10(6) cells) occurred with 500 ppm Tri. Time-, protein concentration-, and both Tri and GSH concentration-dependent formation of DCVG were observed in liver and kidney subcellular fractions. Two kinetically distinct systems were observed in both cytosol and microsomes from pooled liver samples, whereas only one system was observed in subcellular fractions from pooled kidney samples. Liver cytosol exhibited apparent Km values (microM Tri) of 333 and 22.7 and Vmax values (nmol DCVG formed/min per mg protein) of 8.77 and 4.27; liver microsomes exhibited apparent Km values of 250 and 29.4 and Vmax values of 3.10 and 1.42; kidney cytosol and microsomes exhibited apparent Km values of 26.3 and 167, respectively, and Vmax values of 0.81 and 6.29, respectively. DCVG formation in samples of liver cytosol and microsomes from 20 individual donors exhibited a 6.5-fold variation in microsomes but only a 2.4-fold variation in cytosol. In coincubations of pooled liver cytosol and microsomes, addition of an NADPH-regenerating system produced marked inhibition of DCVG formation, but addition of GSH had no effect on cytochrome P-450-catalyzed formation of chloral hydrate. These results indicate that both human kidney and liver have significant capacity to catalyze DCVG formation, indicating that the initial step of the GSH-dependent pathway is not limiting in the formation of nephrotoxic and nephrocarcinogenic metabolites.     

The glucuronidation of mycophenolic acid by human liver, kidney and jejunum microsomes

AIMS: To estimate the relative contribution of liver, kidney and jejunum to MPA elimination via glucuronidation from in vitro kinetic data. METHODS: The kinetics of MPA glucuronidation by human liver, kidney and jejunum microsomes were characterized. Mycophenolic acid glucuronide (MPAG) concentrations in microsomal incubations were determined using a specific h.p.l.c. procedure. Non-specific microsomal binding of MPA was excluded using an equilibrium dialysis approach. RESULTS: Microsomes from all three tissues catalysed the conversion of MPA to MPAG. Mean microsomal intrinsic clearances for MPAG formation by liver, kidney and jejunum microsomes were 46.6, 73.5 and 24.5 microl (min mg)(-1), respectively. When extrapolated to the whole organ, however, hepatic intrinsic clearance was 21- and 38-fold higher than the respective intrinsic clearances for kidney and small intestine. CONCLUSIONS: The data suggest that the liver is the organ primarily responsible for the systemic clearance of MPA, with little contribution from the kidney, and that the small intestine would be expected to contribute to first-pass extraction to a minor extent only.     

氯化两面针碱在体外对肝细胞L-O2和肾细胞293增殖的影响

目的:观察氯化两面针碱在体外对人胃癌细胞7901、人鼻咽癌细胞cne1-1、人肾癌细胞ketr-3、人肝癌细胞BEL 7404的增殖抑制作用和对人胚肝细胞L-O2和人胚肾细胞293的增殖影响。方法:采用MTT法检测不同浓度氯化两面针碱对4种肿瘤细胞和人胚肝细胞L-O2及人胚肾细胞293增殖的影响,比较其IC50值;采用紫外分光光度法测定氯化两面针碱作用后肝、肾细胞培养上清液中SOD,MDA,LDH值的变化。结果:氯化两面针碱在体外对肿瘤细胞具有较强的增殖抑制作用,同时对人胚肝细胞L-O2和人胚肾细胞293有一定的增殖抑制作用。结论:氯化两面针碱能抑制多种肿瘤细胞的增殖,对肿瘤细胞有明显细胞毒性作用,同时可抑制人胚肝细胞L-O2和人胚肾细胞293的增殖,对人体肝、肾细胞有一定的毒性。     

Alterations in the rat kidney associated with sodium saccharin feeding

Sodium saccharin has previously been demonstrated to induce hyperplasia and tumors of the urothelium of the rat urinary bladder. It was fed as 5% of the diet to male F344 rats for 2 years. In the present experiment, mild simple hyperplasia of the urinary bladder epithelium was again frequently observed, and a marked nodular and papillary hyperplasia of the urothelium of the kidney pelvis was also found in approximately half of the sodium saccharin-fed rats. This was infrequently associated with focal calcification of the renal papilla and pelvis. In contrast, rats fed sodium saccharin had a significantly reduced incidence of the interstitial nephritis frequently observed in older rats. No significant incidence was observed of lesions of tissues other than those of the urinary tract.