Improved genomic/nuclear DNA extraction for 8-hydroxydeoxyguanosine analysis of small amounts of rat liver tissue

Nakae, Dai; Mizumoto, Yasushi; Kobayashi, Eisaku; Noguchi, Osamu; Konishi, Yoichi

doi:10.1016/0304-3835(95)03980-b

Cited by 152 publications

(74 citation statements)

References 18 publications

Supporting

Mentioning

Contrasting

Unclassified

Order By: Relevance

“…Although a tendency for elevated 8-OHdG levels in the brain was seen in the work of Hirano et al (1996), it was not clear, and "low" levels in the preceding studies were always around a single 8-OHdG within 10 5 dGs, corresponding to "high" in the present study. Substantial decline of background noises can be achieved by improving the HPLC-ECD method with introduction of a reliable DNA extraction using a commercially available kit (Helbock et al, 1998;1999;Nakae et al, 1995). Representative 8-OHdG immunohistochemistry in the digestive system: Liver at 18 weeks (A, ϫ50); pancreas at 6 weeks (B, ϫ66); glandular stomach at 6 weeks (C, ϫ40); and intestine at 6 weeks (D, ϫ40).…”

Section: Discussionmentioning

confidence: 99%

“…Portions weighing 50 -100 mg were pulverized in liquid nitrogen, and DNA was extracted by our chaotropic NaI isolation method using a DNA Extractor WB kit (Wako Pure Chemical Industries, Limited, Kyoto, Japan) (Nakae et al, 1995) with a slight modification (Helbock et al, 1998 and1999). DNA hydrolysis and microfiltration of the resultant samples were subsequently conducted by the method of Helbock et al (1999).…”

Section: Hplc-ecd Assay For Nuclear 8-ohdg Levelsmentioning

confidence: 99%

See 1 more Smart Citation

Age and Organ Dependent Spontaneous Generation of Nuclear 8-Hydroxydeoxyguanosine in Male Fischer 344 Rats

Nakae

Akai

Kishida

et al. 2000

Laboratory Investigation

Self Cite

View full text Add to dashboard Cite

SUMMARY: 8-Hydroxydeoxyguanosine (8-OHdG) is a major oxidative DNA adduct playing roles in senescence, carcinogenesis and various disease processes. High-performance liquid chromatography with an electrochemical detection (HPLC-ECD) method has been widely used to assess organ levels of 8-OHdG, and a recently introduced immunohistochemical approach has made it possible to clarify intra-organ localization. In the present study, these methods were employed to reveal age-dependent changes in nuclear 8-OHdG within various tissues of male Fischer 344 rats between 18 fetal days and 104 weeks of age. 8-OHdG was detected in the nuclei of cerebellar small granule and small cortical cells, cerebral nerve cells, and choroid plexus epithelia of the brain and ependymal cells of the spinal cord; parenchymal cells in the anterior lobe of the pituitary and adrenal glands (mainly cortex); bronchial epithelium of the lung; intra-hepatic bile duct, pancreatic duct, glandular gastric and intestinal epithelial cells; renal tubular epithelial cells (mainly medulla); and spermatogonia and spermatocytes of the testis and seminal vesicle epithelia. The nuclear 8-OHdG levels were high (more than two lesions per 10 6 deoxyguanosines) from 7 days to 104 weeks of age in the brain, 3 to 6 weeks in the adrenal gland, 6 to 104 weeks in the lung, and 3 to 52 weeks in the testis. In the other organs, the nuclear 8-OHdG levels remained low throughout. These findings provide a basis for research dealing with oxidative stress by indicating organ-specific and age-but not aging-dependent changes in the localization of spontaneously generated nuclear 8-OHdG in intact rats. The immunohistochemical approach has advantages for assessing variation of 8-OHdG formation at the cellular level not accessible to the HPLC-ECD method. (Lab Invest 2000, 80:249-261).

show abstract

Section: Discussionmentioning

confidence: 99%

Section: Hplc-ecd Assay For Nuclear 8-ohdg Levelsmentioning

confidence: 99%

Age and Organ Dependent Spontaneous Generation of Nuclear 8-Hydroxydeoxyguanosine in Male Fischer 344 Rats

Nakae

Akai

Kishida

et al. 2000

Laboratory Investigation

Self Cite

View full text Add to dashboard Cite

show abstract

“…The 8-OHdG levels of cerebral cortex were measured with a method previously described (23,24). Briefly, nuclear DNA was isolated with a DNA extractor WB kit (Wako Pure Chemical Industries, Osaka, Japan) and digested into deoxynucleotides by treatment with nuclease P1 enzyme (Yamasa Corp., Chiba, Japan) and alkaline phosphatase (Sigma-Aldrich Chemical Co., St. Louis, MO).…”

Section: Methodsmentioning

confidence: 99%

Edaravone Inhibits DNA Peroxidation and Neuronal Cell Death in Neonatal Hypoxic-Ischemic Encephalopathy Model Rat

et al. 2009

View full text Add to dashboard Cite

Neonatal hypoxic-ischemic encephalopathy (HIE) is the most frequent neurologic disease in the perinatal period. Its major cause is oxidative stress, which induces DNA peroxidation and apoptotic neuronal death. We examined 8-hydroxy-2Ј-deoxyguanosine (8-OHdG) expression to evaluate brain damage in neonatal HIE and the therapeutic effect of edaravone, a free radical scavenger. Using HPLC and immunohistochemistry, the 8-OHdG levels of neonatal HIE model Sprague-Dawley rats that were subjected to left common carotid artery ligation and 2-h hypoxia significantly increased after 24 -48 h of hypoxic-ischemic (HI) insult, but decreased after 72 h. Moreover, the number of apoptotic cells with terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling and karyorrhexis significantly increased after 24 -72 h of HI insult. In a therapeutic experiment, edaravone was administered i.p.

show abstract

“…The mononuclear cells were separated in Mono-poly resolving medium (Dainippon Seiyaku, Osaka, Japan). DNA was extracted from mononuclear cells using WB DNA extractor kits (Wako Chemical, Tokyo, Japan) [6]. This chaotrophic DNA isolation method was reported to produce the lowest and least variable 8-oxodG value [7].…”

Section: Methodsmentioning

confidence: 99%

Oxidative DNA damage in diabetes mellitus: its association with diabetic complications

et al. 1999

View full text Add to dashboard Cite

Hyperglycaemia, a key clinical manifestation of diabetes mellitus, not only generates reactive oxygen species (ROS), but also attenuates anti-oxidative mechanisms by scavenging enzymes and antioxidant substances [1]. As ROS cause strand breaks in DNA and base modifications including the oxidation of guanine residues to 8-oxo, 2 ¢-deoxyguanosine (8-oxodG), 8-oxodG can serve as a sensitive biomarker of oxidative DNA damage [2]. 8-OxodG was increased in the kidneys of diabetic rats, and insulin treatment reduced both urinary albumin excretion and 8-oxodG formation in the kidney [3]. A recent study reported an increase in the 8-oxodG content in mononuclear cells and ROS level in Type I (insulin-dependent) and Type II (non-insulin-dependent) diabetic patients when compared with control subjects [4]. Another study reported that urinary 8-oxodG excretion was higher in Type II diabetic patients than in the control subjects [5]. Urinary 8-oxodG excretion correlated with glycated haemoglobin [5]. We speculated that diabetes-associated modifications of DNA by ROS might contribute to the diabetic complications. Diabetologia (1999) Abstract Aims/hypothesis. Augmented oxidative stress induced by hyperglycaemia possibly contributes to the pathogenesis of diabetic complications. Oxidative stress is known to increase the conversion of deoxyguanosine to 8-oxo, 2 ¢-deoxyguanosine in DNA. To investigate the possible contribution of oxidative DNA damage to the pathogenesis of diabetic complications, we measured the content of 8-oxo, 2 ¢-deoxyguanosine in the urine and the blood mononuclear cells of Type II (non-insulin-dependent) diabetic patients. Methods. We studied 53 Type II diabetic patients and 39 age-matched healthy control subjects. We assayed 8-oxo, 2 ¢-deoxyguanosine by HPLC-electrochemical detection method. Results. The content of 8-oxo, 2 ¢-deoxyguanosine in the urine and the mononuclear cells of the Type II diabetic patients was much higher than that of the control subjects. Urinary 8-oxo, 2 ¢-deoxyguanosine excretion and the 8-oxo, 2 ¢-deoxyguanosine content in the mononuclear cells from the diabetic patients with complications were higher than those from the diabetic patients without complications. Urinary excretion of 8-oxo, 2 ¢-deoxyguanosine was significantly correlated with the 8-oxo, 2 ¢-deoxyguanosine content in the mononuclear cells. The 8-oxo, 2 ¢-deoxyguanosine content in the urine and mononuclear cells was correlated with the haemoglobin A 1 c value. Conclusion/interpretation. This is the first report of a direct association between oxidative DNA damage and the complications of diabetes. The augmented oxidative DNA damage in diabetes is speculated to contribute to the pathogenesis of diabetic complications. [Diabetologia (1999) 42: 995±998]

show abstract

Improved genomic/nuclear DNA extraction for 8-hydroxydeoxyguanosine analysis of small amounts of rat liver tissue

Cited by 152 publications

References 18 publications

Age and Organ Dependent Spontaneous Generation of Nuclear 8-Hydroxydeoxyguanosine in Male Fischer 344 Rats

Age and Organ Dependent Spontaneous Generation of Nuclear 8-Hydroxydeoxyguanosine in Male Fischer 344 Rats

Edaravone Inhibits DNA Peroxidation and Neuronal Cell Death in Neonatal Hypoxic-Ischemic Encephalopathy Model Rat

Oxidative DNA damage in diabetes mellitus: its association with diabetic complications

Contact Info

Product

Resources

About