Biomedical subjects

D Gradinaru

Publications and source records attributed to D Gradinaru.

4 recordsLinked to original sources

PubMed2004

Redox state alteration modulates astrocyte glucuronidation.

We have investigated the effects of mild oxidative conditions on drug-metabolizing enzyme activity in rat cultured astrocytes. These experimental conditions promoting an oxidative environment were obtained by short exposure to a low concentration of menadione (5 microM) for a short duration (15 min). This resulted in the rapid and transient production of reactive oxygen species (+130%), associated with a decrease in GSH cellular content (-24%), and an increase in total protein oxidation (+26%), but promoted neither PGE(2) nor NO production. This treatment induced a rapid and persistent decrease in astrocyte glucuronidation activities, which was totally prevented by N-acetyl-l-cysteine. These oxidative conditions also affected the specific UGT1A6 activity measured in transfected V79-1A6 cells. Finally, the subsequent recovery of astrocyte glucuronidation activity may result from upregulation of UGT1A6 expression (+62%) as shown by RT-PCR and gene reporter assay. These results show that the catalytic properties and expression of cerebral UGT1A6 are highly sensitive to the redox environment. The protective effect of N-acetyl-l-cysteine suggests both a direct action of reactive oxygen species on the protein and a more delayed action on the transcriptional regulation of UGT1A6. These results suggest that cerebral metabolism can be altered by physiological or pathological redox modifications.

Animals↗

PubMed2001

Rat olfactory bulb and epithelium UDP-glucuronosyltransferase 2A1 (UGT2A1) expression: in situ mRNA localization and quantitative analysis.

UDP-glucuronosyltransferases (UGTs) form a multigenic family of enzymes involved in the biotransformation and elimination of numerous endo- and xenobiotic compounds. Beside the diverse UGT isoforms present in the liver as well as in other tissues, the UGT2A1 isoform, also called olfactory UGT, was initially thought to be expressed in the nasal epithelium only. In this work, we demonstrate the UGT2A1 mRNA expression in the olfactory bulb, using in situ hybridization and quantitative reverse transcription-polymerase chain reaction (RT-PCR) techniques. Within the epithelium, UGT2A1 mRNA is mainly found in the sustentacular cells and to a lesser extent in Bowman's gland cells. Moreover, in situ hybrization staining reveals UGT2A1 mRNA expression in the olfactory sensory neuron nuclei. Neuronal localization of UGT2A1 mRNA within the olfactory bulb is mainly found in the deeper granular cells. The development of the quantitative multistandard RT-PCR method firstly required characterization of the mouse Ugt2A1 cDNA by rapid amplification of cDNA ends (RACE)-PCR. UGT2A1 mRNA levels appear quantitatively six-fold lower in the olfactory bulb than in the epithelium, in both the rat and mouse. The expression of UGT2A1 in the olfactory bulb, which directly connects the nasal epithelium to the brain, emphasizes the potential role of this enzyme in the protection of the brain against airborne hazardous chemicals.

Air Pollutants↗

PubMed1999

UDP-glucuronosyltransferase in the rat olfactory bulb: identification of the UGT1A6 isoform and age-related changes in 1-naphthol glucuronidation.

Xenobiotic glucuronidation represents a major metabolic protection of the brain against chemical aggressions at blood-brain interfaces. We previously observed that glucuronidation of 1-naphthol was very effective in olfactory bulb, which is a pathway for the entry of foreign molecules into the brain. In this work, we showed that 1-naphthol glucuronidation varied according to age. It was very high at birth, then decreased markedly in 3-month-old rats and increased again significantly during aging. By Western blot and reverse transcription-polymerase chain reaction (RT-PCR), we demonstrated the presence in the olfactory bulb of the UDP-glucuronosyltransferase (UGT) 1A6 isoform, which catalyzes the glucuronidation of phenols, such as 1-naphthol. Quantitative RT-PCR indicated that the mRNA levels encoding UGT1A6 did not significantly change according to age, thus suggesting that other differently regulated UGT isoforms were present and would account for the variations of 1-naphthol glucuronidation observed.

Aging↗

PubMed1998

Identification of the uridine diphosphate glucuronosyltransferase isoform UGT1A6 in rat brain and in primary cultures of neurons and astrocytes.

The expression of a phenol uridine diphosphate glucuronosyltransferase (UGT) was investigated in rat brain homogenate and in primary cultures of astrocytes and neurons, by means of model substrates (1-naphthol and 4-methylumbelliferone) assays, Western blot analysis and reverse transcription-polymerase chain reaction (RT-PCR) experiments. Glucuronidation of these substances occurred in cerebral cell or brain homogenates, although to different extents. The specific activity was the highest in astrocytes, with values more than 10- and 100-fold those found in neurons or total brain, respectively. Using antibodies able to recognize several rat liver UGT isoforms, only one protein with an apparent molecular mass of 54 kDa was detected in astrocyte and neuron homogenates and brain microsomes. RT-PCR experiments run with primers specifically designed for the rat liver UGT1A6 revealed amplificons of the expected sizes in accordance with the presence of UGT1A6 mRNA. The nucleotide sequence of the 330-base pair product was 100% homologous to that of exon 1 of rat liver isoform UGT1A6. In conclusion, this work allowed us to identify for the first time a constitutive cerebral UGT isoform identical to rat liver UGT1A6, which glucuronidates planar phenolic substances in cultured astrocytes, neurons, and the entire brain.

Animals↗