Stimulation of RNA polymerase II elongation by hepatitis delta antigen.

Transcription elongation by RNA polymerase II (RNAPII) is negatively regulated by the human factors DRB-sensitivity inducing factor (DSIF) and negative elongation factor (NELF). A 66-kilodalton subunit of NELF (NELF-A) shows limited sequence similarity to hepatitis delta antigen (HDAg), the viral protein required for replication of hepatitis delta virus (HDV). The host RNAPII has been implicated in HDV replication, but the detailed mechanism and the role of HDAg in this process are not understood. We show that HDAg binds RNAPII directly and stimulates transcription by displacing NELF and promoting RNAPII elongation. These results suggest that HDAg may regulate RNAPII elongation during both cellular messenger RNA synthesis and HDV RNA replication.

Survival of retinal ganglion cells after transection of the optic nerve in adult cats: a quantitative study within two weeks.

We have previously reported that a small number of retinal ganglion cells (RGCs) of adult cats survive 2 months after transection of the optic nerve (ON) and that alpha cells have the greatest ability to survive among different types of RGCs (Watanabe et al., 1995). Here we report the time course of RGC survival within 15 days after ON transection using retrograde labeling with DiI injected into the bilateral lateral geniculate nuclei of cats. The density of DiI-labeled RGCs in the central retina as well as in the periphery did not change until day 3 after ON transection, then decreased rapidly, to 43% of the original density on day 7, and falling to 19% by day 14. We then intracellularly injected Lucifer yellow into the DiI-labeled RGCs to examine the difference in the time course between surviving alpha and beta cells. Similar to the density change in total surviving RGCs, the proportion of surviving beta cells did not change until day 3, then decreased rapidly to 65% of the original density on day 4, falling to 12% by day 14. By contrast, 64% of alpha cells survived for 14 days after axotomy. Analysis of regression lines for survival time courses indicated that death of beta cells was characterized with a rapid period phase from day 3 to day 7 after axotomy whereas that of alpha cells lacked it. Axon-like sprouting from surviving beta cells was first recognized in the nerve fiber layer on day 3, and were later more conspicuous.

SPT genes: key players in the regulation of transcription, chromatin structure and other cellular processes.

Suppressor of Ty (SPT) genes were originally identified through a genetic screen for mutations in the yeast Saccharomyces cerevisiae that restore gene expression disrupted by the insertion of the transposon Ty. Classic members of the SPT gene family, SPT11, SPT12, and SPT15, encode for the histones H2A and H2B, and for TATA-binding protein (TBP), respectively. Over the past few years, molecular complexes and cellular functions in which other SPT gene products involve have been discovered through genetic and biochemical studies in yeast and several other organisms: Key regulators of transcription and chromatin structure, such as DSIF, SAGA, and FACT, all contain SPT gene products as essential subunits. In addition, accumulating evidence suggests that SPT gene products play more diverse roles, including roles in DNA replication, DNA recombination and developmental regulation. Here we review the current understanding of the functions and roles of the SPT genes, with special emphasis on the role of SPT5 in transcript elongation and in neuronal development in vertebrates.

Environmental light enhances survival and axonal regeneration of axotomized retinal ganglion cells in adult cats.

A segment of peripheral nerve was transplanted to the cut stump of the optic nerve to facilitate axonal regeneration of retinal ganglion cells (RGCs) in adult cats. The cats were reared under different light environment: 12 h light-12 h darkness, additional flash light under conventional light cycle, or 24 h darkness. After 60 days, the density and morphology of RGCs with regenerated axons were examined with retrograde labeling by fluoro-ruby and intracellular injections of Lucifer Yellow. In the retina of cats reared in darkness, densities of RGCs with regenerated axons were 11-42% of those in the retina of cats reared under conventional light and dark cycle. More than half of the labeled RGCs were degenerative in the retina of cats reared in darkness, while most RGCs were normal under conventional environment or flash light. We conclude that environmental light is essential for the survival and axonal regeneration of axotomized RGCs.

Lipid peroxide levels and superoxide-scavenging abilities of Sera obtained from hotbred (Thoroughbred) horses.

Hotbred (Thoroughbred) horses were grouped into three classes according to the levels of constant physical exercise (foals, 6 months old; racing horses, 5 years old; horses for breeding, 6-10 years old), and lipid peroxide levels in their sera were measured as thiobarbituric acid-reactive substances. No significant differences were observed among them. The superoxide-scavenging abilities of sera were measured next; to examine the antioxidative properties of hotbreds, and were found to be highest in the racing horses. The higher scavenging ability of the racing horses might contribute to keep their lipid peroxide levels as low as those of the other two groups. HPLC analysis of substances in sera suggested that the presence of albumin-bound bilirubin was one of the reasons for the high superoxide-scavenging ability of sera of the racing horses. When the hotbreds were compared with coldbred (crossbred) horses, the lipid peroxide levels of hotbreds was higher (7.0 +/- 1.2 nmol/ml) than that of coldbreds (2.6 +/- 0.7 nmol/ml). Comparison of the superoxide-scavenging abilities of sera between hotbreds and coldbreds showed that the hotbreds possessed higher scavenging ability than the coldbreds. These results indicated that the lipid peroxide level in sera of hotbreds was higher than that of coldbreds regardless of the higher superoxide-scavenging abilities of sera.

The effects of alpha-phenyl-tert-butyl nitrone (PBN) on copper-induced rat fulminant hepatitis with jaundice.

In the present study we demonstrated the protective effects of the spin-trapping agent alpha-phenyl-tert-butyl nitrone (PBN) against fulminant hepatitis with jaundice in LEC rats. In LEC rats an excess amount of copper is accumulated in the liver and causes hepatitis with severe jaundice. PBN was subcutaneously administered every 2 d at the concentration of 128 mg/kg, beginning with 13-week-old rats and continuing for 17 weeks. PBN prevented the loss of body weight, reduced death rate, and suppressed the increase in GTP and GOT values reflecting hepatic cell destruction. Ocular inspection also confirmed the suppressive effects of PBN on jaundice. In parallel with these phenomena, the amounts of thiobarbituric acid-reactive substances (TBARS) in livers of PBN-administered rats were found to be lower than those of non-PBN-administered rats. Little histological changes were observed in PBN-administered rats in comparison with non-PBN-administered rats. The protective effect of PBN on the formation of oxidative damage in liver DNA was observed but not so remarkable as that on lipid peroxidation. From these results, it was concluded that PBN had the liver-protective effects against fulminant hepatitis with jaundice. This suggested that free radicals play an important role in abnormally accumulated copper-induced liver injury and that PBN potentially has therapeutic value for the treatment of hepatitis.

The suppression of age-related accumulation of lipid peroxides in rat brain by administration of Rooibos tea (Aspalathus linearis).

The protective effects of Rooibos tea (RT), Aspalathus linearis, against damage to the central nervous system (CNS) accompanying aging were examined by both the thiobarbituric acid reaction (TBA) and magnetic resonance imaging (MRI) methods in brains of chronically RT-treated rats. Ad libitum administration of RT was begun with 3-month-old Wistar female rats and continued for 21 months. The contents of TBA reactive substances (TBARS) in the frontal cortex, occipital cortex, hippocampus and cerebellum in 24-month-old rats after administration with water were significantly higher than those in young rats (5 weeks old). However, no significant increase of TBARS was observed in RT-administered aged rats. When MR images of the brains of 24-month-old rats with and without RT as well as 5-week-old rats were taken, a decrease of the signal intensity was observed in the cerebral cortex, hippocampus and cerebellum in MR images of aged rats without RT, whereas little change of the signal intensity was observed in MR images of the same regions of 24-month-old rats treated with RT, whose images were similar to those of young rats. These observations suggested that (1) the age-related accumulation of lipid peroxides in the brain was closely related to the morphological changes observed by MRI, and (2) chronic RT-administration prevented age-related accumulation of lipid peroxides in several regions of rat brain.

In vitro photosensitized lysis of red blood cells by an antifungal drug griseofulvin.

Human red blood cells (RBCs) were lysed by in vitro irradiation in the presence of the antifungal drug griseofulvin (GF). Effects of UVA fluence and GF concentration on photohaemolysis were examined under aerobic conditions. The photohaemolysis occurred at much lower fluence than that necessary for oxidation of the membrane lipids. UVA-irradiated solution of GF did not cause haemolysis. The photohaemolysis was colloid osmotic in nature because it was preceded by K+ leakage from the cells and was delayed in the presence of 30 mM sucrose in the medium. Even under anaerobic conditions, RBCs were lysed by irradiation with higher fluence than that required for aerobic photohaemolysis. Therefore, some phototoxic mechanism other than photosensitized oxidation is also involved in the photohaemolysis.

Possible involvement of oxidation of lipids in inducing griseofulvin photosensitivity.

Griseofulvin-sensitized photo-oxidation of lipids was examined in vitro. Iodometric determination and thiobarbituric acid assay proved peroxidation of squalene, cholesterol and red blood cell membrane lipids by UVA irradiation in the presence of griseofulvin. Simultaneous progression of Type I (non-singlet oxygen) and Type II (singlet oxygen) processes is likely because the oxidation products of cholesterol through both processes were identified by thin layer chromatography. These results suggest that lipids, especially membrane lipids, are one of the possible targets for induction of griseofulvin photosensitivity.

[Mode of action of fluoropyrimidines, in relation to their clinical application].

From results which we obtained in experiments on 5-fluoropyrimidine analogs, it was demonstrated that the mode of action of these compounds was as follows: In human cancer, in contrast to experimental animal tumors, it was demonstrated that thymidine phosphorylase (Thd Pase) activity was dominant compared to that of uridine phosphorylase (Urd Pase). This observation would indicate that 5-fluorouracil (5-FU) was mainly metabolized to produce 5-fluoro-2'-deoxyuridine-5'-monophosphate (FdUMP), via 5-flu fluoro-2'-deoxyuridine(FdUR). FdUMP would inhibit DNA synthesis by blocking thymidilate synthetase activity, resulting in the death of cancer cells. Thd Pase activity by which 5-FU is converted to FdUR would participate greatly in the antitumor effect of the compounds because of the fact that FdUR has 1,000 times greater antitumor activity than 5-FU. Moreover, some of the fluoropyrimidines are activated by this enzyme to produce 5-FU. It should be emphasized that Thd Pase activity is higher in tumor tissue than in normal tissues. This finding would mean that the compounds would exert a selective (antitumor) activity on cancer cells. In addition, a clinical trial of fluoropyrimidines showed them to be effective when administered to patients in order to maintain a lower effective level of 5-FU in tumor tissues for a longer period of time. Such experiences would also lend more weight the possible of the mode of action of the of fluoropyrimidines mentioned above.

New sulfamoylphenethylamines, potent alpha 1-adrenoceptor antagonists.

The (+)-isomer of amosulalol, a combined alpha- and beta-adrenoceptor antagonist, was one log unit order more potent and less potent than the (-)-isomer in blocking alpha 1- and beta 1-adrenoceptors, respectively, in anaesthetized rats. Nine newly synthesized desoxy compounds derived from amosulalol and its analogues were found to possess potent alpha 1-adrenoceptor blocking activity and to be practically devoid of beta 1-adrenoceptor blocking activity. Among the desoxy derivatives, YM-12617 was more potent than prazosin in blocking alpha 1-adrenoceptors in anaesthetized rats and in reducing blood pressure, total peripheral resistance and left ventricular work in anaesthetized dogs.