[Show abstract][Hide abstract] ABSTRACT: One hypothesis for the etiology of behavioral disorders is that infection by a virus induces neuronal cell dysfunctions resulting in a wide range of behavioral abnormalities. However, a direct linkage between viral infections and neurobehavioral disturbances associated with human psychiatric disorders has not been identified. Here, we show that transgenic mice expressing the phosphoprotein (P) of Borna disease virus (BDV) in glial cells develop behavioral abnormalities, such as enhanced intermale aggressiveness, hyperactivity, and spatial reference memory deficit. We demonstrate that the transgenic brains exhibit a significant reduction in brain-derived neurotrophic factor and serotonin receptor expression, as well as a marked decrease in synaptic density. These results demonstrate that glial expression of BDV P leads to behavioral and neurobiological disturbances resembling those in BDV-infected animals. Furthermore, the lack of reactive astrocytosis and neuronal degeneration in the brains indicates that P can directly induce glial cell dysfunction and also suggests that the transgenic mice may exhibit neuropathological and neurophysiological abnormalities resembling those of psychiatric patients. Our results provide a new insight to explore the relationship between viral infections and neurobehavioral disorders.
Proceedings of the National Academy of Sciences 08/2003; 100(15):8969-74. DOI:10.1073/pnas.1531155100 · 9.67 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The effects of hypophysectomy on hepatic and extrahepatic UDP-glucuronosyltransferase activities in adult male rats were observed. UDP-glucuronosyltransferase activities toward 1-naphthol decreased to 20-30% of control in the liver, kidney, lung, and testis. The mRNA of UGT1A6, which is an isoform contributing to the glucuronidation of various phenolic xenobiotics such as 1-naphthol, were decreased drastically in the liver, kidney, and testis by hypophysectomy. However, while bilirubin UDP-glucuronosyltransferase activity in the liver intensified, there was only a slight increase in the activity in the kidney and no alteration in the lung. The mRNA of UGT1A1, which is an isoform contributing to the glucuronidation of bilirubin, increased significantly in the liver and slightly in the kidney after hypophysectomy. These inductions and reductions in enzymatic activities and mRNA levels in each tissue were restored to control levels by intermittent injections of rat growth hormone. Interestingly, while hepatic UGT activity toward bisphenol A remained constant in hypophysectomized rats, the testicular UGT activity declined to 10-15% of control but returned to normal levels following growth hormone treatment, suggesting that an unknown UGT isoform (s) mediates bisphenol A glucuronidation in the testis. These results indicate that the expression of extrahepatic UGT is isoform-specific and regulated differentially in tissues by the pituitary gland.
Journal of Biochemistry 09/2002; 132(2):265-70. DOI:10.1093/oxfordjournals.jbchem.a003220 · 2.58 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The latency-associated transcript (LAT) promoter of pseudorabies virus (PrV) is unique among the many promoters of the viral genome in that it remains active during the latent state. The regulatory mechanism of PrV LAT gene expression is complex and different between latency and lytic infection of cultured cells. Although two different sequences, LAP1 and LAP2, are thought to be involved in LAT gene expression, the function of the upstream region of the LAT promoter (LAP1 and LAP2) remains an enigma, even in cultured cells. To analyze the function of the upstream region, it is necessary to examine the effects of the upstream sequence on LAT gene expression in the absence of other viral proteins. Transient expression assays were performed by employing a series of reporter plasmids in which various sequences upstream of the LAT promoter (from nucleotide positions -592 to +423 relative to the transcriptional start site of the large latency transcript (LLT)) were linked to the chloramphenicol acetyltransferase (CAT) gene in cells of neuronal and non-neuronal origin. We identified a region (from nucleotide positions -3606 to -1386) that was capable of repressing the LAT promoter activity in Vero cells by analyzing CAT gene expression of the series of reporter plasmids. This effect was not observed in Neuro-2a cells. We have also shown that the LAT promoter activity of the reporter plasmid containing the upstream region was repressed by the immediate-early gene product IE180 in Vero cells, but not in Neuro-2a cells. These results suggest that the upstream region of the LAT promoter may have a role in repressing LAT gene expression in cultured non-neuronal cells.
[Show abstract][Hide abstract] ABSTRACT: The 92 kDa type VI collagenase (matrix metalloproteinase-9 (MMP-9)) activities on zymography assay were found to be 1–6 times higher in benign tumor breast tissues of 12 canines and 4–26 times higher in adenocarcinoma breast tissues of nine canines than that of control tissues, respectively. A full-length canine MMP-9 cDNA was cloned from the adenocarcinoma tissue by reverse transcription-PCR and 5′- and 3′-RACE. The isolated cDNA contained an open reading frame coding for a polypeptide of 704 amino acids. The predicted protein sequence displayed extensive similarity to that of known MMP-9s and contained a putative signal sequence, a propeptide, an active site with three zinc-binding histidine residues, a calcium-binding domain, a hemopexin region, and three key cysteine residues. Western blotting using MMP-9-specific antibodies prepared against the peptide corresponding to Arg642–Asp704 of canine MMP-9 and Northern blotting using a MMP-9-specific cDNA fragment as a probe confirmed that MMP-9 (the 92 kDa protein band) was highly expressed in canine mammary adenocarcinoma tissues. Higher levels of MMP-9 activity were found in the sera of canines with mammary adenocarcinoma. The results indicated that MMP-9 plays an important role in the progression of a canine mammary tumor and that assay of serum MMP-9 is helpful for early diagnosis as progress of adenocarcinoma.
Biochimica et Biophysica Acta (BBA) - General Subjects 11/2001; 1568(1):7-12. DOI:10.1016/S0304-4165(01)00192-1 · 4.38 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Pseudorabies virus (PRV) early protein 0 (EP0) consisting of 410 amino acids is a transactivator of viral genes. A mutant consisting of amino acids 1–113 exhibits dominant-negative properties. In order to assess the antiviral potential of the EP0 mutant, Vero cells were transformed with the EP0 mutant gene expressed in a tetracycline-regulated system. The transformed cell lines showed marked resistance to PRV infection when expression of the EP0 mutant gene was induced. In the transformed cell line infected with PRV, synthesis of the immediate-early protein (IE180) and of EP0 was inhibited, whereas the levels of IE and EP0 messenger RNA (mRNA) were not decreased, as compared with those of the control cell line. The present results suggest that the EP0 mutant may not alter the efficiency of the viral gene transcription but rather translation efficiency of the viral mRNA.
[Show abstract][Hide abstract] ABSTRACT: A chimeric gene encoding a fusion protein consisting of the DNA-binding domain of the immediate-early (IE) protein of pseudorabies virus (PRV) and a tail-truncated VP16 of herpes simplex virus 1, lacking the transcription activation domain, has been shown to repress transcription of the PRV IE gene, resulting in the inhibition of PRV growth in vitro. To assess the antiviral potential of the fusion protein in vivo, transgenic mice containing the chimeric gene under the control of the virus- and interferon-inducible Mx 1 promoter were generated. A transgenic mouse line showed marked resistance to PRV infection when the mice were challenged intranasally with PRV. Inhibition of PRV replication was also observed in monolayers of embryonic cells prepared from the transgenic mice. In the cells infected with PRV, transcription of the PRV IE gene was repressed. The present results indicate that the chimeric gene is able to exert a significant antiviral effect against PRV infection in vivo.
[Show abstract][Hide abstract] ABSTRACT: Bisphenol A, an environmental oestrogenic chemical, was found to conjugate highly with glucuronic acid in male rat liver microsomes studied in vitro. In the various isoforms tested (1A1, 1A3, 1A5, 1A6, 1A7 and 2B1), glucuronidation of bisphenol A and of diethylstilboestrol, a synthetic crystalline compound possessing oestrogenic activity and known to be glucuronidated by liver microsomes, was catalysed by an isoform of UDP-glucuronosyltransferase (UGT), namely UGT2B1, which glucuronidates some endogenous androgens. UGT activity towards bisphenol A in liver microsomes and in UGT2B1 expressed in yeast AH22 cells (22.9 and 0.58 nmol/min per mg of microsomal proteins respectively) was higher than that towards diethylstilboestrol (75.0 and 4.66 pmol/min per mg of microsomal proteins respectively). UGT activities towards both bisphenol A and diethylstilboestrol were distributed mainly in the liver but were also observed at substantial levels in the kidney and testis. Northern blot analysis disclosed the presence of UGT2B1 solely in the liver, and about 65% of the male rat liver microsomal UGT activities towards bisphenol A were absorbed by the anti-UGT2B1 antibody. These results indicate that bisphenol A, in male rat liver, is glucuronidated by UGT2B1, an isoform of UGT.
[Show abstract][Hide abstract] ABSTRACT: A phenol UDPglucuronosyltransferase (UGT) was partially purified, and the cDNA encoding the isoform was cloned and sequenced from sheep small intestine. The purified preparation containing a one major band (57 kDa) and one minor band (50 kDa) revealed high activities toward xenobiotics such as 1-naphthol (1-NA), 4-nitrophenol, and 4-methylumbelliferone. The preparation, however, had only little activity toward 4-hydroxybiphenyl and no activity toward bilirubin, suggesting that the preparation contains UGT1 isoforms. The NH2-terminal amino acid sequence of the major band was determined to be Gly-Lys-Leu-Leu-Val-Val-Pro-Met-Asp-Gly-Ser. A full-length UGT cDNA was obtained by reverse transcription-polymerase chain reaction with the degenerated 5'-primer from the NH2-terminal amino acid sequence of the purified major one and rapid amplification of cDNA ends from sheep small intestine. The cloned cDNA named sheUGT1A07 by amino acid similarity has a NH2-terminus sequence identical to that of the purified major one. Another phenol UGT cDNA named sheUGT1A6 was also cloned from sheep liver. sheUGT1A6 was expressed mainly in the liver, whereas sheUGT1A07 mRNA was expressed almost only in the alimentary organs, suggesting that sheUGT1A6 plays a role as a general drug metabolizing UGT isoform in the liver and sheUGT1A07 plays important role in the xenobiotics glucuronidation in the sheep small intestine.
Archives of Biochemistry and Biophysics 05/1999; 364(2):143-52. DOI:10.1006/abbi.1999.1123 · 3.02 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: In the rat intestine, UDP-glucuronosyltransferase (UGT) isoforms were highly induced by oral administration of 2-naphthoflavone, as shown by intestinal UGT activity toward 1-naphthol (1-NA). The greatest increase in UGT activity occurred in the duodenum. Using UGT1A6 cDNA as a probe, we obtained three types of clones corresponding to UGT1A2, UGT1A6 and UGT1A7, in a ratio of 1:1:8, from a cDNA library constructed from the 2-naphthoflavone-treated rat intestine. The induction of each isoform was evaluated by means of Northern blotting with isoform-specific probes. The mRNAs of UGT1A6 (glucuronizing various phenolic xenobiotics) and the mRNAs of UGT1A7 (glucuronizing the ultimate carcinogenic metabolite of benzo(a)pyrene) were expressed constitutively and were highly induced in the duodenum and proximal jejunum. S1 mapping showed that induction of the isoforms of the UGT1 family was more pronounced in the liver than in the small intestine and that UGT1A7 was the major UGT1 isoform in the small intestine of vehicle-treated rats and in that of 2-naphthoflavone-treated rats. These results indicate that, in rats, UGT1A7 is expressed constitutively and is particularly inducible in the small intestine. In the light of these results, we believe that the UGT1A7 isoform would play an important role in glucuronidation in the small intestinal mucosa of rats.
European Journal of Biochemistry 01/1999; 258(3):948-55. DOI:10.1046/j.1432-1327.1998.2580948.x · 3.58 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Phenol UDP-glucuronosyltransferase activity was highly induced in the microsomes of the kidney medulla of rats by beta-naphthoflavone treatment. In the medulla, phenol UDP-glucuronosyltransferase and its mRNA were greatly increased in both immunoblotting and Northern blot analyses following beta-naphthoflavone treatment of the rats. In untreated rat kidneys, phenol UDP-glucuronosyltransferase was detected by immunohistochemical analysis only in proximal convolution tubular cells located in the cortex. After beta-naphthoflavone treatment of the rats, UDP-glucuronosyltransferase appeared in the epithelial cells in the straight portion of the distal tubules located in the medulla. In conclusion, the medullary distal tubular cells have high latent glucuronidation activity and are thought to play an important role in drug excretion.