A new device for the determination of microsomal cytochrome P-450 in renal tissue preparations from various species contaminated with mitochondria and hemoglobin.

Ohno, Yasuo; Kawanishi, Toru; Takahashi, Atsushi; Kasuya, Yutaka; Omori, Yoshihito

doi:10.1254/jjp.32.679

Cited by 9 publications

(3 citation statements)

References 16 publications

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“…Preliminary experiments showed minor spectral interference in the dithionite difference spectra, and therefore bias in the CYP content measurements and subsequent MPPGK estimates was unlikely when considered alongside the inter-assay variability (Matsubara et al, 1976). Furthermore, the dog kidney cortex microsomal CYP content measured in the current study using the dithionite difference method (Table 3) were comparable with a value reported using a customised spectral method (0.223 nmol/mg protein; (Ohno et al, 1982)). CYP content measured in dog liver were also in good agreement with previously published values (Smith et al, 2008).…”

Section: Discussionsupporting

confidence: 83%

“…Ensuring complete homogenization of kidney tissue, while also limiting contamination of microsomes with other sources of haemoproteins such as mitochondria, can be challenging. When measuring CYP content in kidney cortex, the low CYP levels, and potential for spectral interference from contaminating haemoproteins, makes accurate quantification challenging (Jakobsson and Cinti, 1973; Ohno et al, 1982). Preliminary experiments showed minor spectral interference in the dithionite difference spectra, and therefore bias in the CYP content measurements and subsequent MPPGK estimates was unlikely when considered alongside the inter-assay variability (Matsubara et al, 1976).…”

Section: Discussionmentioning

confidence: 99%

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Microsomal and Cytosolic Scaling Factors in Dog and Human Kidney Cortex and Application for In Vitro-In Vivo Extrapolation of Renal Metabolic Clearance

Scotcher¹,

Billington²,

Brown³

et al. 2017

Drug Metab Dispos

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In vitro-in vivo extrapolation of drug metabolism data obtained in enriched preparations of subcellular fractions rely on robust estimates of physiologically relevant scaling factors for the prediction of clearance in vivo. The purpose of the current study was to measure the microsomal and cytosolic protein per gram of kidney (MPPGK and CPPGK) in dog and human kidney cortex using appropriate protein recovery marker and evaluate functional activity of human cortex microsomes. Cytochrome P450 (CYP) content and glucose-6-phosphatase (G6Pase) activity were used as microsomal protein markers, whereas glutathione-S-transferase activity was a cytosolic marker. Functional activity of human microsomal samples was assessed by measuring mycophenolic acid glucuronidation. MPPGK was 33.9 and 44.0 mg/g in dog kidney cortex, and 41.1 and 63.6 mg/g in dog liver (n = 17), using P450 content and G6Pase activity, respectively. No trends were noted between kidney, liver, and intestinal scalars from the same animals. Species differences were evident, as human MPPGK and CPPGK were 26.2 and 53.3 mg/g in kidney cortex (n = 38), respectively. MPPGK was 2-fold greater than the commonly used in vitro-in vivo extrapolation scalar; this difference was attributed mainly to tissue source (mixed kidney regions versus cortex). Robust human MPPGK and CPPGK scalars were measured for the first time. The work emphasized the importance of regional differences (cortex versus whole kidney–specific MPPGK, tissue weight, and blood flow) and a need to account for these to improve assessment of renal metabolic clearance and its extrapolation to in vivo.

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Section: Discussionsupporting

confidence: 83%

Section: Discussionmentioning

confidence: 99%

Microsomal and Cytosolic Scaling Factors in Dog and Human Kidney Cortex and Application for In Vitro-In Vivo Extrapolation of Renal Metabolic Clearance

Scotcher¹,

Billington²,

Brown³

et al. 2017

Drug Metab Dispos

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“…Enzyme Activity Assays, GSH and Total Glutathione Determinations Kidney cytochrome P-450 was measured as described by Omura and Sato [16] as modified by Ohno et al [17] who corrected for hemoglobin and mitochondria contaminations. p-Nitrophenol glucuronyl transferase was estimated in the microsomal suspension by the method of Burchell and Weatherill [18].…”

Section: Animal Treatment and Microsome Preparationmentioning

confidence: 99%

Cisplatin Nephrotoxicity in Cadmium-Pretreated Rats

Bompart¹,

Orfila²,

Manuel³

1991

Nephron

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Treatment of rats with cisplatin or with cisplatin after chronic pre-exposure to cadmium induced a decrease in kidney cytochrome P-450 and glutathione levels, and in glutathione peroxidase and reductase activities. Furthermore, cadmium and cisplatin enhanced lipid peroxidation, oxidized glutathione and N-glucuronyl transferase activity. Glutathione S-transferase (substrate: 1-chloro-2,4-dinitrobenzene) was increased and decreased by cadmium and cisplatin respectively. On morphological observation, cadmium nephrotoxicity was characterized by tubular proximal damage with mitochondrial and lysosomal changes and a widespread vesiculation of tubular cells. A marked focal tubular necrosis associated with cyst formation was observed in cisplatin nephrotoxicity. On the basis of the measured biochemical, functional and histological parameters, it is concluded that cadmium pretreatment did not potentiate the nephrotoxic effect of cisplatin.

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Cisplatin-induced changes in cytochrome P-450, lipid peroxidation and drug-metabolizing enzyme activities in rat kidney cortex

Bompart¹

1989

Toxicology Letters

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A new device for the determination of microsomal cytochrome P-450 in renal tissue preparations from various species contaminated with mitochondria and hemoglobin.

Abstract: Using a spectrophotometer connected to a microcomputer, the carbon monoxide difference spectrum of renal microsomal dithionite reduced cytochrome P-450 was measured avoiding the effects of the

Cited by 9 publications

References 16 publications

Microsomal and Cytosolic Scaling Factors in Dog and Human Kidney Cortex and Application for In Vitro-In Vivo Extrapolation of Renal Metabolic Clearance

Microsomal and Cytosolic Scaling Factors in Dog and Human Kidney Cortex and Application for In Vitro-In Vivo Extrapolation of Renal Metabolic Clearance

Cisplatin Nephrotoxicity in Cadmium-Pretreated Rats

Cisplatin-induced changes in cytochrome P-450, lipid peroxidation and drug-metabolizing enzyme activities in rat kidney cortex

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